ovalbumin and Bronchial-Hyperreactivity

House dust mite (HDM) is the most pervasive indoor aeroallergen source worldwide. Allergens derived from HDM are associated with sensitization and allergic asthma. Allergic asthma is an immunologically driven disease characterized by a Th2-polarized immune response, eosinophilic inflammation, airway hyperreactivity, and remodeling. Animal models of asthma utilizing ovalbumin (OVA) exposure have afforded us considerable insight with respect to the mediators and cell types involved in allergic airway inflammation. However, OVA preparations and HDM are two vastly different materials. This chapter is specifically concerned with modeling responses to HDM exposure in mice. These studies have furnished new information and unlocked new lines of inquiry regarding biological responses to common aeroallergens. The complexity of HDM as an allergen source, with its plethora of protein and nonprotein immunogenic components, may influence the mechanisms underlying sensitization, inflammation and remodeling. Here, we will discuss this issue, along with giving critical thought to the use of experimental models.

Topics: Animals; Antigens, Dermatophagoides; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Humans; Hypersensitivity; Mice; Ovalbumin; Pyroglyphidae; Th2 Cells

Allergic bronchial asthma is characterized by chronic inflammation of the airways, development of airway hyperreactivity and recurrent reversible airway obstruction. Target and effector cells responsible for airway hyperresponsiveness and airway obstruction include sensory and motor neurons as well as epithelial and smooth muscle cells. Although it is well established that the inflammatory process is controlled by T-helper-2 (Th2) cells, the mechanisms by which immune cells interact with neurons, epithelial cells or smooth muscle cells still remain uncertain. Due to growing evidence for extensive communication between neurons and immune cells, the mechanisms of this neuroimmune crosstalk in lung and airways of asthmatic patients are becoming the focus of asthma research. Neurotrophins represent molecules potentially responsible for regulating and controlling the crosstalk between the immune and peripheral nervous system. They are constitutively expressed by resident lung cells and produced in increasing concentrations by immune cells invading the airways under pathological conditions. Neurotrophins modify the functional activity of sensory and motor neurons, leading to enhanced and altered neuropeptide and tachykinin production. These effects are defined as neuronal plasticity. The consequences are the development of neurogenic inflammation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Humans; Mice; Mice, Knockout; Nerve Growth Factors; Neurogenic Inflammation; Neuronal Plasticity; Ovalbumin; Rats; Receptor, Nerve Growth Factor; Receptors, Nerve Growth Factor; T-Lymphocytes, Helper-Inducer; Tachykinins; Time Factors

Nerve growth factor (NGF), in addition to its essential role in neuronal growth and survival, may also act as an inflammatory mediator. As several animal studies have shown, NGF appears to play a part in the development of airway hyperresponsiveness and in the increased sympathetic and sensory innervation of the lung. It also has a profound effect on airway inflammation and asthma-related symptoms. Sources of NGF in the airways are numerous: inflammatory cells infiltrated into the bronchial mucosa, and structural cells including lung fibroblasts, airway epithelial and smooth muscle cells. These cells, by releasing more NGF in inflammatory conditions, may contribute to the increased NGF levels observed in bronchoalveolar lavage fluid and serum from patients with asthma. Taken together, these results suggest that NGF is an important mediator in both inflammation and asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Disease Models, Animal; Dose-Response Relationship, Drug; Fibroblasts; Humans; Inflammation; Interleukin-1; Lung; Muscle, Smooth; Nerve Growth Factor; Neuroimmunomodulation; Ovalbumin; Trachea

Eclipta prostrata (L.) L. is a medicinal plant used by many ethnic groups in Brazil to treat respiratory diseases, hepatitis and the bites of venomous animals. A methanolic extract of E. prostrata (MEEP), the major components of which are wedelolactone (WED) and demethylwedelolactone (DMW), exhibited anti-inflammatory activity in acute asthma models but the effects on lung inflammation and the mechanisms of action of MEEP in a chronic asthma model are not known.. To study the effects of MEEP in vivo using a chronic ovalbumin (OVA)-induced allergic asthma model in mice.. The identities of WED and DMW in MEEP were confirmed and the concentrations determined by liquid chromatography and tandem mass spectrometry. Male Balb/c mice were sensitized and challenged with OVA and experimental animals were treated with MEEP (100, 250 or 500 mg/kg) while control animals were treated with dexamethasone (2 mg/kg) or normal saline. Bronchial hyperresponsiveness, total and differential cell counts in bronchoalveolar lavage (BAL), and the production of Th2 cytokines in lung homogenates were assessed. Lung inflammation and mucus production were evaluated by histological analysis while nuclear factor kappa-B (NF-κB) activation was assessed immunohistochemically.. Concentrations of WED and DMW in MEEP were 5.12% and 1.04%, respectively. Treatments with MEEP (250 or 500 mg/kg) significantly decreased bronchial hyperresponsiveness, reduced total cell and eosinophil counts in BAL and IL-4 concentrations in lung homogenate, and inhibited NF-κB activation. Treatment with MEEP at 500 mg/kg reduced the level of IL-5 in lung homogenates but did not decrease IL-13 concentration or mucus production.. MEEP attenuated bronchial hyperresponsiveness and decreased lung and airway inflammation in a chronic asthma model in mice. The mechanism of action involves inhibition of NF-κB activation, most likely associated with the presence of the coumestans WED and DMW. These results support the ethnopharmacological evidence for the use of E. prostrata against asthma and other respiratory inflammatory diseases.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eclipta; Hypersensitivity; Inflammation; Lung; Methanol; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin

Stachytarpheta cayennensis (Verbenaceae) has been used in Brazilian traditional medicine to treat asthma and other respiratory diseases.. To investigate the effects of different doses of standardized hydro-ethanolic (SCH) and aqueous (SCA) extracts of aerial parts of S. cayennensis using a murine ovalbumin (OVA)-induced asthma model.. The major constituents of the plant extracts were identified and standardized by ultra-performance liquid chromatography coupled with mass spectrometry. Balb/c mice were challenged with OVA solution and treated concomitantly by intraperitoneal injection of standardized SCH or SCA extracts at 50, 100, and 200 mg/kg concentrations. OVA-challenged control animals were treated with either dexamethasone (OVA-DEX) or saline solution (OVA-SAL). After challenge, we assessed in vivo bronchial hyperresponsiveness, airway inflammation (number of cells), peribronchial inflammation (histological analysis) and production of OVA-specific IgE and interleukin (IL)-4, IL-5, and IL-13 (ELISA).. Acteoside, isoacteoside, and ipolamiide were the major constituents of SCH and SCA. The respective concentrations of acteoside in SCH and SCA were 78 and 98 μg/mL, while those of ipolamiide were 30 and 19 μg/mL. Treatment with 200 mg/kg of SCH or SCA decreased IL-4, IL-5, and IL-13 in lung homogenates. These reductions were accompanied by a lower influx of inflammatory cells (eosinophils, lymphocytes, and macrophages) to the airways and lungs. In addition to the anti-inflammatory effects, administration of SCA, but not SCH, ameliorated the parameters of bronchial hyperresponsiveness and decreased levels of circulating OVA-specific IgE.. The results presented herein demonstrate for the first time the anti-asthmatic activity of S. cayennensis extracts in a murine model, thereby supporting the ethnopharmacological uses of the plant.

Topics: Animals; Anti-Asthmatic Agents; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Immunoglobulin E; Inflammation; Interleukin-13; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Verbenaceae

Over the past 50 years, the prevalence of allergic respiratory diseases has been increasing. The Hygiene hypothesis explains this progression by the decrease in the bio-diversity of early microbial exposure. This study aims to evaluate the effect of early-life farm exposure on airway hyperresponsiveness and cough hypersensitivity in an allergic airway inflammation rabbit model.. A specific environment was applied to pregnant rabbits and their offspring until six weeks after birth. Rabbits were housed in a pathogen-free zone for the control group and a calf barn for the farm group. At the end of the specific environmental exposure, both groups were then housed in a conventional zone and then sensitized to ovalbumin. Ten days after sensitization, the rabbit pups received ovalbumin aerosols to provoke airway inflammation. Sensitization to ovalbumin was assessed by specific IgE assay. Cough sensitivity was assessed by mechanical stimulation of the trachea, and bronchial reactivity was assessed by methacholine challenge. The farm environment was characterized by endotoxin measurement.. A total of 38 rabbit pups were included (18 in the farm group). Endotoxin levels in the farm environment varied from 30 to 1854 EU.m-3. There was no significant difference in specific IgE values to ovalbumin (p = 0.826) between the two groups. The mechanical threshold to elicit a cough did not differ between the two groups (p = 0.492). There was no difference in the number of cough (p = 0.270) or the intensity of ventilatory responses (p = 0.735). After adjusting for age and weight, there was no difference in respiratory resistance before and after methacholine challenge.. Early exposure to the calf barn did not affect cough sensitivity or bronchial reactivity in ovalbumin-sensitized rabbits. These results suggest that not all farm environments protect against asthma and atopy. Continuous exposure to several sources of microbial diversity is probably needed.

Topics: Animals; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cough; Dust; Endotoxins; Farms; Immunoglobulin E; Inflammation; Methacholine Chloride; Ovalbumin; Rabbits

Bronchial asthma (BA) is one of the most common chronic inflammatory disease of airways. There are huge experimental data indicating that Th2-cytokines IL-4 and IL-13 play a key role in BA pathogenesis. They are implicated in the IgE synthesis, eosinophil infiltration to the lungs and in the development of airway hyperreactivity (AHR), that makes these cytokines the promising targets. Neutralization of IL-4 and IL-13 or its common receptor chain (IL-4Rα) by monoclonal antibodies substantially reduce asthma symptoms. RNA interference provides a novel method for regulation of gene expression by siRNA molecules. In this study we evaluated whether the siRNA targeted to IL-4 and IL-13 reduce BA symptoms in mice model. Experimental BA was induced in BALB/c mice by sensitization to ovalbumin allergen followed by intranasal challenge. The intranasal delivery of siRNAs targeted to IL-4 and IL-13 inhibited the lung expression of these cytokines by more than 50% that led to the attenuation of AHR and pulmonary inflammation; the quantity of eosinophils in lungs which are one of the major inflammatory cells involved in allergic asthma pathogenesis decreased by more than 50% after siRNA treatment. These data support the possibility of a dual IL-4 and IL-13 inhibition by locally delivered siRNAs which in turn leads to the suppression of allergen-induced pulmonary inflammation and AHR.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophils; Immunoglobulin E; Inflammation; Interleukin-13; Interleukin-4; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; RNA, Small Interfering

Control groups received treatment with normal saline or dexamethasone (2 mg/kg) on the same day. We assessed. MCHA significantly improved airway hyperresponsiveness near baseline levels. MCHA administration significantly improved airway and lung inflammation, demonstrated by decreased total and inflammatory cells in BAL, lower levels of IL-5 and IL-13 in lung homogenate, and fewer inflammatory cells in lung tissue. Additionally, MCHA significantly diminished goblet cells in lung tissue.. Administration of a hydroethanolic extract of

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Humans; Inflammation; Lung; Mice; Mice, Inbred BALB C; Momordica charantia; Ovalbumin

Bronchial asthma (BA) is a heterogeneous chronic inflammatory disease of the airways. The majority of patients with mild to moderate BA develop Th2-biased eosinophilic pulmonary inflammation and respond well to corticosteroid treatment. However up to 10% of BA patients develop severe pathology, which is associated with neutrophilic inflammation and resistant to conventional corticosteroid therapy. Contrary to eosinophil-predominant airway inflammation neutrophilic BA is developed through Th1- and Th17-immune responses. However, the etiology of corticoid insensitive neutrophilic BA is still remains unclear. Therefore, in the current study we developed a mouse model of BA with predominant neutrophilic rather than eosinophilic pulmonary inflammation. BALB/c mice were immunized with the mixture of the ovalbumin allergen and Freund's adjuvant, followed by aerosol challenge with the same allergen mixed with E. coli lipopolysaccharide. As a result, mice developed the main BA manifestations: production of allergen specific IgE, development of airway hyperreactivity, airway remodeling and pulmonary neutrophilic inflammation. Moreover, this pathology developed through Th1- and Th17-dependent mechanisms and mice with induced neutrophilic BA phenotype responded poorly to dexamethasone treatment, that coincide to clinical observations. The established mouse model could be useful both for studying the pathogenesis and for testing novel approaches to control neutrophilic BA.

Topics: Adrenal Cortex Hormones; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Escherichia coli; Humans; Inflammation; Lung; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Pneumonia; Steroids

Asthma currently affects more than 339 million people worldwide. In the present preliminary study, we examined the efficacy of a new, inhalable soluble epoxide hydrolase inhibitor (sEHI), 1-trifluoromethoxyphenyl-3-(1-propionylpiperidin-4-yl) urea (TPPU), to attenuate airway inflammation, mucin secretion, and hyper-responsiveness (AHR) in an ovalbumin (OVA)-sensitized murine model. Male BALB/c mice were divided into phosphate-buffered saline (PBS), OVA, and OVA+TPPU (2- or 6-h) exposure groups. On days 0 and 14, the mice were administered PBS or sensitized to OVA in PBS. From days 26-38, seven challenge exposures were performed with 30 min inhalation of filtered air or OVA alone. In the OVA+TPPU groups, a 2- or 6-h TPPU inhalation preceded each 30-min OVA exposure. On day 39, pulmonary function tests (PFTs) were performed, and biological samples were collected. Lung tissues were used to semi-quantitatively evaluate the severity of inflammation and airway constriction and the volume of stored intracellular mucosubstances. Bronchoalveolar lavage (BAL) and blood samples were used to analyze regulatory lipid mediator profiles. Significantly (p < 0.05) attenuated alveolar, bronchiolar, and pleural inflammation; airway resistance and constriction; mucosubstance volume; and inflammatory lipid mediator levels were observed with OVA+TPPU relative to OVA alone. Cumulative findings indicated TPPU inhalation effectively inhibited inflammation, suppressed AHR, and prevented mucosubstance accumulation in the murine asthmatic model. Future studies should determine the pharmacokinetics (i.e., absorption, distribution, metabolism, and excretion) and pharmacodynamics (i.e., concentration/dose responses) of inhaled TPPU to explore its potential as an asthma-preventative or -rescue treatment.

Topics: Aerosols; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Epoxide Hydrolases; Humans; Inflammation; Lipids; Male; Mice; Mice, Inbred BALB C; Ovalbumin

Asthma is a lung disease that has influenced more than 350 million people worldwide. Airway smooth muscle (ASM) spasm leads to airway hyperresponsiveness (AHR) and bronchial obstruction, which are clinical manifestations of an asthma attack. Botulinum toxin (BTX) is a bacteria toxin that acts as muscle relaxant and may have therapeutic effects on AHR and asthma.. In this study, the effect of BTX on AHR and related gene expressions was evaluated.. An asthma mice model was developed which was treated with BTX in two ways: intranasally (IN) and via nebulization (N) (0.01, 0.1, and 1 U/mL and 10 U/mL, respectively) on days 25, 27 and 29. AHR was evaluated on days 24, 26, 28, and 30, and gene expressions were evaluated for TrkA, TrkB, M1-M5, α7nAChR, TNF-α, and extracellular signal-regulated kinase 2 (ERK2) proteins. For histopathology of the lungs, perivascular and peribronchial inflammation, production of mucus, and goblet cell hyperplasia were studied.. On day 24, treatment with BTX (for all doses) had no significant effect on AHR, but on days 26 and 28, AHR was decreased and this continued up to day 30 for all treated groups. Treatment with BTX had no significant effect on the gene expressions of TrkA, TrkB, M1-M5, α7nAChR, TNF-α, and ERK2 proteins, perivascular inflammation, peribronchial inflammation, hyperplasia of the goblet cell and production of mucus. Besides, mice administered with 10 mg/mL BTX perished. The BTX therapy controlled asthma attacks by decreasing AHR and relaxation of ASMs.. However, BTX had no significant effect on airway inflammation and production of mucus. While using BTX, it is necessary to prescribe safe doses in order to prevent adverse reactions.

Topics: alpha7 Nicotinic Acetylcholine Receptor; Animals; Asthma; Botulinum Toxins; Bronchi; Bronchial Hyperreactivity; Disease Models, Animal; Humans; Hyperplasia; Inflammation; Mice; Mice, Inbred BALB C; Myocytes, Smooth Muscle; Ovalbumin; Signal Transduction; Tumor Necrosis Factor-alpha

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Immunoglobulin E; Mice; Mice, Inbred BALB C; Ovalbumin; Vitamin E

Asthma has been the most prevalent chronic respiratory disease (Mensah et al. J Allergy Clin Immunol 142:744-748, 2018). To explore pathogenic mechanism or new treatments of asthma, mice have been utilized to model the disease. Eosinophilic airway inflammation, allergen specific-IgE, and airway hyperresponsiveness have been characteristic features of allergic asthma (Drake et al. Pulm Ther 5:103-115, 2019). In mouse models, airway hyperresponsiveness to inhaled broncho-constrictor agents such as methacholine chloride (MCh) has been a key disease marker (Alessandrini et al. Front Immunol 11:575936, 2020). A variety of systems to assess airway reactivity in mice are currently available. Here, three distinct systems are described as these have been used in many publications. In the first system, an invasive system in which mice are anesthetized and intubated followed by mechanical ventilation, lung resistance (R), dynamic compliance (C), and other respiratory parameters with MCh challenge are measured. In the second system, a noninvasive system equipped with a chamber in which mice can move freely and spontaneously breathe, changes in airways with MCh challenge are measured as enhanced pause (Penh) values. In the third system, in vitro airway smooth muscle (ASM) reactivity is monitored in an extracted mouse tracheal duct with a cholinergic agonist challenge or electrical stimulation. Each of these systems has unique features, benefits, or disadvantages.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophilia; Immunoglobulin E; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Respiration Disorders; Respiratory Hypersensitivity

Extracellular acidification is a major component of tissue inflammation, including airway inflammation. The extracellular proton-sensing mechanisms are inherent in various cells including airway structural cells, although their physiological and pathophysiological roles in bronchial smooth muscles (BSMs) are not fully understood. In the present study, to explore the functional role of extracellular acidification on the BSM contraction, the isolated mouse BSMs were exposed to acidic pH under contractile stimulation.. The RT-PCR analyses revealed that the proton-sensing G protein-coupled receptors were expressed both in mouse BSMs and cultured human BSM cells. In the mouse BSMs, change in the extracellular pH from 8.0 to 6.8 caused an augmentation of contraction induced by acetylcholine. Interestingly, the acidic pH-induced BSM hyper-contraction was further augmented in the mice that were sensitized and repeatedly challenged with ovalbumin antigen. In this animal model of asthma, upregulations of G protein-coupled receptor 68 (GPR68) and GPR65, that were believed to be coupled with Gq and Gs proteins respectively, were observed, indicating that the acidic pH could cause hyper-contraction probably via an activation of GPR68. However, psychosine, a putative antagonist for GPR68, failed to block the acidic pH-induced responses.. These findings suggest that extracellular acidification contributes to the airway hyperresponsiveness, a characteristic feature of bronchial asthma. Further studies are required to identify the receptor(s) responsible for sensing extracellular protons in BSM cells.

Topics: Acetylcholine; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Humans; Hydrogen-Ion Concentration; Inflammation; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Protons; Psychosine; Receptors, G-Protein-Coupled

Asthma is a T helper 2 (Th2) cell-associated chronic inflammatory diseases characterized with airway obstruction, increased mucus production, and eosinophil infiltration. Conventional medications for asthma treatment cannot fully control the symptoms, and potential side effects are also the concerns. Thus, complement or alternative medicine (CAM) became a new option for asthma management. Ding Chuan Tang (DCT) is a traditional Chinese herbal decoction applied mainly for patients with coughing, wheezing, chest tightness, and asthma. Previously, DCT has been proved to improve children airway hyperresponsiveness (AHR) in a randomized and double-blind clinical trial. However, the mechanisms of how DCT alleviates AHR remain unclear. Since asthmatic features such as eosinophil infiltration, IgE production, and mucus accumulation are relative with Th2 responses, we hypothesized that DCT may attenuate asthma symptoms through regulating Th2 cells. Ovalbumin (OVA) was used as a stimulant to sensitize BALB/c mice to establish an asthmatic model. AHR was detected one day before sacrifice. BALF and serum were collected for immune cell counting and antibody analysis. Splenocytes were cultured with OVA in order to determine Th2 cytokine production. Lung tissues were collected for histological and gene expression analyses. Our data reveal that DCT can attenuate AHR and eosinophil accumulation in the 30-day sensitization asthmatic model. Histological results demonstrated that DCT can reduce cell infiltration and mucus production in peribronchial and perivascular site. In OVA-stimulated splenocyte cultures, a significant reduction of IL-5 and IL-13 in DCT-treated mice suggests that DCT may alleviate Th2 responses. In conclusion, the current study demonstrates that DCT has the potential to suppress allergic responses through the reduction of mucus production, eosinophil infiltration, and Th2 activity in asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Down-Regulation; Eosinophils; Female; Immunization; Immunoglobulin E; Interleukin-13; Interleukin-5; Mice, Inbred BALB C; Mucus; Ovalbumin; Plant Extracts; Pneumonia; Spleen

CpG-oligodeoxynucleotides (CpG-ODNs) constitute an attractive alternative for asthma treatment. However, very little evidence is available from studies on the oral administration of CpG-ODNs in animals. Previously, we developed acid-resistant particles (named ODNcap) as an oral delivery device for ODNs. Here, we showed that free feeding of an ODNcap-containing feed prophylactically attenuates allergic airway inflammation, hyperresponsiveness, and goblet cell hyperplasia in an ovalbumin-induced asthma model. Using transcriptomics-driven approaches, we demonstrated that injury of pulmonary vein cardiomyocytes accompanies allergen inhalation challenge, but is inhibited by ODNcap feeding. We also showed the participation of an airway antimicrobial peptide (Reg3γ) and fecal microbiota in the ODNcap-mediated effects. Collectively, our findings suggest that daily oral ingestion of ODNcap may provide preventive effects on allergic bronchopulmonary insults

Topics: Administration, Oral; Animals; Antimicrobial Peptides; Bronchial Hyperreactivity; Female; Gastrointestinal Microbiome; Hypersensitivity; Mice; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Ovalbumin; Pancreatitis-Associated Proteins; Pneumonia

Callicarpa japonica Thunb., as an herbal medicine has been used for the treatment of inflammatory diseases in China and Korea.. Ultra performance liquid chromatography-photodiode array-quadrupole time-of-flight mass spectrometer (UPLC-PDA-QTof MS) was used to detect the major phenylethanoid glycosides in the C. japonica extract. BALB/c mice were intraperitoneally sensitized by ovalbumin (OVA) (on days 0 and 7) and challenged by OVA aerosol (on days 11-13) to induce airway inflammatory response. The mice were also administered with C. japonica Thunb. (CJT) (20 and 40 mg/kg Per oral) on days 9-13. CJT pretreatment was conducted in lipopolysaccharide (LPS)-stimulated RAW264.7 or phorbol 12-myristate 13-acetate (PMA)-stimulated A549 cells.. CJT administration significantly reduced the secretion of Th2 cytokines, TNF-α, IL-6, immunoglobulin E (IgE) and histamine, and the recruitment of eosinophils in an OVA-exposed mice. In histological analyses, the amelioration of inflammatory cell influx and mucus secretion were observed with CJT. The OVA-induced airway hyperresponsiveness (AHR), iNOS expression and NF-κB activation were effectively suppressed by CJT administration. In addition, CJT led to the upregulation of HO-1 expression. In an in vitro study, CJT pretreatment suppressed the LPS-induced TNF-α secretion in RAW264.7 cells and attenuated the PMA-induced IL-6, IL-8 and MCP-1 secretion in A549 cells. These effects were accompanied by downregulated NF-κB phosphorylation and by upregulated HO-1 expression.. These results suggested that CJT has protective activity against OVA-induced airway inflammation via downregulation of NF-κB activation and upregulation of HO-1, suggesting that CJT has preventive potential for the development of allergic asthma.

Topics: A549 Cells; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Callicarpa; Cytokines; Disease Models, Animal; Female; Heme Oxygenase-1; Humans; Lung; Membrane Proteins; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Plant Extracts; RAW 264.7 Cells; Signal Transduction; Up-Regulation

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Inflammation; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory System

2,6-Bis-(4-hydroxyl-3-methoxybenzylidine) cyclohexanone (BHMC), a synthetic curcuminoid analogue, has been shown to exhibit anti-inflammatory properties in cellular models of inflammation and improve the survival of mice from lethal sepsis. We further evaluated the therapeutic effect of BHMC on acute airway inflammation in a mouse model of allergic asthma. Mice were sensitized and challenged with ovalbumin (OVA), followed by intraperitoneal administration of 0.1, 1, and 10 mg/kg of BHMC. Bronchoalveolar lavage fluid, blood, and lung samples were collected, and the respiratory function was measured. OVA sensitization and challenge increased airway hyperresponsiveness (AHR) and pulmonary inflammation. All three doses of BHMC (0.1-10 mg/kg) significantly reduced the number of eosinophils, lymphocytes, macrophages, and neutrophils, as well as the levels of Th2 cytokines (IL-4, IL-5 and IL-13) in bronchoalveolar lavage fluid (BALF) as compared to OVA-challenged mice. However, serum level of IgE was not affected. All three doses of BHMC (0.1-10 mg/kg) were effective in suppressing the infiltration of inflammatory cells at the peribronchial and perivascular regions, with the greatest effect observed at 1 mg/kg which was comparable to dexamethasone. Goblet cell hyperplasia was inhibited by 1 and 10 mg/kg of BHMC, while the lowest dose (0.1 mg/kg) had no significant inhibitory effect. These findings demonstrate that BHMC, a synthetic nonsteroidal small molecule, ameliorates acute airway inflammation associated with allergic asthma, primarily by suppressing the release of inflammatory mediators and goblet cell hyperplasia to a lesser extent in acute airway inflammation of allergic asthma.

Topics: Acute Disease; Animals; Asthma; Bronchial Hyperreactivity; Curcumin; Cyclohexanones; Cytokines; Goblet Cells; Immunoglobulin E; Leukocytes; Male; Mice; Mice, Inbred BALB C; Ovalbumin

In this study we assessed the effects of antigen exposure in mice pre-sensitized with allergen following viral infection on changes in lung function, cellular responses and tight junction expression. Female BALB/c mice were sensitized to ovalbumin and infected with influenza A before receiving a second ovalbumin sensitization and challenge with saline, ovalbumin (OVA) or house dust mite (HDM). Fifteen days post-infection, bronchoalveolar inflammation, serum antibodies, responsiveness to methacholine and barrier integrity were assessed. There was no effect of infection alone on bronchoalveolar lavage cellular inflammation 15 days post-infection; however, OVA or HDM challenge resulted in increased bronchoalveolar inflammation dominated by eosinophils/neutrophils or neutrophils, respectively. Previously infected mice had higher serum OVA-specific IgE compared with uninfected mice. Mice previously infected, sensitized and challenged with OVA were most responsive to methacholine with respect to airway resistance, while HDM challenge caused significant increases in both tissue damping and tissue elastance regardless of previous infection status. Previous influenza infection was associated with decreased claudin-1 expression in all groups and decreased occludin expression in OVA or HDM-challenged mice. This study demonstrates the importance of the respiratory epithelium in pre-sensitized individuals, where influenza-infection-induced barrier disruption resulted in increased systemic OVA sensitization and downstream effects on lung function.

Topics: Airway Resistance; Animals; Bronchial Hyperreactivity; Claudin-1; Down-Regulation; Female; Influenza A virus; Methacholine Chloride; Mice; Mice, Inbred BALB C; Orthomyxoviridae Infections; Ovalbumin; Pyroglyphidae; Treatment Outcome

Allergic diseases and especially allergic asthma are widespread diseases with high prevalence in childhood, but also in adults. Acid sphingomyelinase (ASM) is a key regulator of the sphingolipid pathway. Previous studies defined the association of ASM with the pathogenesis of T. To determine the role of Asm under baseline conditions, wild-type (WT) and Asm. At baseline, Asm. Asm deficiency could induce higher numbers of T

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Sphingomyelin Phosphodiesterase; Th2 Cells

Myxopyrum serratulum A. W. Hill. (Oleaceae) is a traditionally used Indian medicinal plant for the treatment of cough, asthma and many other inflammatory diseases.. In this study, the protective effects of M. serratulum on airway inflammation was investigated in ovalbumin (OVA)-induced murine model of allergic asthma and lipopolysaccharide (LPS)-stimulated inflammation in RAW 264.7 murine macrophages, and the possible mechanisms were elucidated.. The phytochemicals present in the methanolic leaf extract of M. serratulum (MEMS) were identified by reverse phase high performance liquid chromatography (RP-HPLC) analysis. In vitro anti-inflammatory activity of MEMS were evaluated by estimating the levels of nitric oxide (NO), reactive oxygen species (ROS) and cytokines (IL-1α, IL-1β, IL-2, IL-4, IL-6, IL-10, IL-12, IL-17A, IFN-γ, TNF-α, G-CSF and GM-CSF) in LPS-stimulated RAW 264.7 macrophages. In vivo anti-asthmatic activity of MEMS was studied using OVA-induced murine model. Airway hyperresponsiveness (AHR), was measured; total and differential cell counts, eosinophil peroxidase (EPO), prostaglandin E2 (PGE2), NO, ROS, and cytokines (IL-4, IL-5 and IL-13), were estimated in bronchoalveolar lavage fluid (BALF). Serum total IgE level was measured; and the histopathological changes of lung tissues were observed. The expressions of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase (iNOS) in lung tissue homogenates were detected by Western blot.. The chromatographic analysis of MEMS identified the presence of gallic acid, protocatechuic acid, catechin, ellagic acid, rutin, p-coumaric acid, quercetin, naringenin and apigenin. MEMS (125 and 250 μg/mL) dose-dependently reduced the levels of NO, ROS and pro-inflammatory cytokines in LPS-stimulated RAW 264.7 macrophages. MEMS (200 and 400 mg/kg, p.o.) significantly (p < 0.05) alleviated AHR; number of inflammatory cells, EPO, PGE2, NO, ROS, and cytokines (IL-4, IL-5 and IL-13) in BALF; serum total IgE and the histopathological changes associated with lung inflammation. Western blot studies showed that MEMS substantially suppressed COX-2 and iNOS protein expressions in the lung tissues of OVA-sensitized/challenged mice.. The present study corroborates for the first time the ameliorative effects of MEMS on airway inflammation by reducing the levels of oxidative stress, pro-inflammatory cytokines and inhibiting COX-2, iNOS protein expressions, thereby validating the ethnopharmacological uses of M. serratulum.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Cytokines; Disease Models, Animal; Female; Inflammation Mediators; Lipopolysaccharides; Lung; Macrophages; Mice; Mice, Inbred BALB C; Nitric Oxide; Oleaceae; Ovalbumin; Plant Extracts; Plant Leaves; RAW 264.7 Cells; Reactive Oxygen Species

This study is to determine the role and mechanism of cryptotanshinone (CTS) in allergic airway inflammation. Asthma induced by OVA was established in BALB/c mice. We found increased airway hyperresponsiveness (AHR), increased inflammatory cell infiltration, elevated levels of TNF-α, interleukin-1β (IL-1β), IL-4, IL-5, IL-6 and IL-13, decreased interferon gamma (IFN-γ) in lung tissue, increased content of total immunoglobulin E (IgE), OVA specific IgE, Eotaxin, ICAM-1, VCAM-1, nuclear factor-kappaB (NF-κB) and phosphorylation of p38 MAPK in lung tissue. However, the administration of CTS significantly decreased AHR in asthmatic mice, reduced inflammation around the bronchioles and inflammatory cells around airway, regulated cytokine production, reduced the total IgE and OVA-specific IgE levels, and inhibited NF-κB activation and p38 MAPK phosphorylation. In vitro experiments in 16 HBE cells revealed that CTS attenuated CAM-1 and IL-6 expression. These results indicate that CTS alleviates allergic airway inflammation by modulating p38 MAPK phosphorylation and NF-κB activation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemotaxis, Leukocyte; Cytokines; Drugs, Chinese Herbal; Female; Hypersensitivity; Immunoglobulin E; Inflammation; Lung; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; p38 Mitogen-Activated Protein Kinases; Phenanthrenes; Phosphorylation

The present study explored the function and regulatory mechanism of High mobility group box 1 (HMGB1) in asthma.. OVA (ovalbumin)-induced asthmatic mice model and LPS-treated cellular model were established in this study. Airway inflammation was measured through detecting the expression of IL-4, IL-5, IL-13 and Interferon-γ (IFN-γ) in serum and BALF (bronchoalveolar lavage fluid) by ELISA kits. Bioinformatics predictive analysis, ChIP assays, Luciferase reporter assay and Western blotting were used to explore the relation between HMGB1 and HSF1 (Heat shock factor 1).. HMGB1 expression was increased in OVA-induced asthmatic mice. Silencing HMGB1 attenuated the increasing of IgE, inflammatory factors (IL-4, IL-5 and IL-13), and airway hyperresponsiveness that induced by OVA. In addition, our study found that HSF1 directly bind with the HMGB1 promoter and negatively regulation of HMGB1. HSF-1 were upregulated in OVA-induced asthmatic mice, and knockdown of HSF1 aggravated the OVA-induced airway inflammation and airway hyperreactivity in mice may through promoting the expression of HMGB1 and the activation of the Toll-like receptor 4 (TLR4)/Myeloid differentiation primary response 88 (MyD88)/Nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) signal pathway.. The expression of HMGB1 could be negatively regulated by HSF1, and the TLR4/MyD88/NF-κB signal pathway was involved in HSF1/HMGB1-mediated regulation of asthma.

Topics: Animals; Apoptosis; Asthma; Base Sequence; Bronchial Hyperreactivity; Cytokines; Heat Shock Transcription Factors; HEK293 Cells; HMGB1 Protein; Humans; Inflammation; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Myeloid Differentiation Factor 88; NF-kappa B; Ovalbumin; Promoter Regions, Genetic; Signal Transduction; Toll-Like Receptor 4

Innate immune activation through exposure to indoor and outdoor pollutants is emerging as an important determinant of asthma severity. For example, household levels of the bacterial product lipopolysaccharide (LPS) are associated with increased asthma severity. We hypothesized that activation of the innate immune receptor TLR5 by its bacterial ligand flagellin will exacerbate airway inflammation and asthma symptoms.. We determined the effect of flagellin co-exposure with ovalbumin in a murine model of allergic asthma. We evaluated the presence of flagellin activity in house dust of asthma patients. Finally, we analyzed the association of a dominant-negative polymorphism in TLR5 (rs5744168) with asthma symptoms in patients with asthma.. We showed that bacterial flagellin can be found in the house dust of patients with asthma and that this bacterial product exacerbates allergic airway inflammation in an allergen-specific mouse model of asthma. Furthermore, a dominant-negative genetic polymorphism in TLR5, the receptor for flagellin, is associated with decreased symptoms in patients with asthma.. Together, our results reveal a novel genetic protective factor (TLR5 deficiency) and a novel environmental pollutant (microbial flagellin) that influence asthma severity. (Clinical trials NCT01688986 and NCT01087307).

Topics: Adult; Animals; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Case-Control Studies; Cross-Sectional Studies; Cytokines; Disease Models, Animal; Female; Flagellin; HEK293 Cells; Humans; Lung; Male; Mice, Inbred C57BL; Middle Aged; Ovalbumin; Polymorphism, Single Nucleotide; Signal Transduction; Th1 Cells; Toll-Like Receptor 5

This study was to investigate of the mechanism by which histone deacetylase (HDAC) 8 inhibitor ameliorated airway hyperresponsiveness (AHR) and allergic airway inflammation.. Mice were sensitized and then treated with budesonide (BUD) or PCI-34051 (PCI) prior to exposing to normal saline (NS) or ovalbumin (OVA). The raw264.7 cells were treated with interleukin (IL)-4 and PCI or shRNA alone. Repetitive measurements of enhanced pause (Penh) were executed by increasing concentrations of acetyl-β-methacholine chloride (0 - 50 mg/ml). Cells in bronchoalveolar lavage fluid (BALF) and pathological changes of lungs were examined, respectively. The expression levels of HDAC8, Galecitn (Gal)-3, CD68, CD86, CD163, Arg1 and NOS2 in lungs were measured. Co-regulation of HDAC8 and Gal-3 proteins was observed by immunofluorescence staining and co-immunoprecipitation assay (Co-IP).. Significant increases in Penh and IL-4 level were detected with a large inflammatory infiltrate, comprised predominantly of macrophages and eosinophils, into the BALF in OVA-exposed lungs. HDAC8, Gal-3, CD68, CD86, CD163, Arg1 and NOS2 proteins were over-expressed with the significant changes in the Arg1 and NOS2 mRNA levels in the lungs and the IL-4-treated cells. PCI intervention obviously reduced the counts of CD163. The HDAC8 inhibitor ameliorates AHR and airway inflammation in animal model of allergic asthma through reducing HDAC8-Gal-3 interaction and M2 macrophage polarization.

Topics: Animals; Bronchial Hyperreactivity; Cell Polarity; Female; Galectin 3; Histone Deacetylases; Hydroxamic Acids; Indoles; Lung; Macrophages; Mice; Mice, Inbred BALB C; Ovalbumin; Random Allocation; RAW 264.7 Cells

Bronchial asthma is a chronic disease characterized by inflammation, obstruction, and hyperresponsiveness of the airways. There is currently no curative therapy for asthma. Type 2 helper T cell response plays a critical role in the pathogenesis of the disease. Protein S is a glycoprotein endowed with anticoagulant, anti-inflammatory, and anti-apoptotic properties. Whether protein S can suppress bronchial asthma and be useful for its therapy is unknown.. To address this question here we compared the development of allergen-associated bronchial asthma between wild type and protein S-overexpressing transgenic mice. Mice were sensitized and challenged with ovalbumin. We also evaluated the circulating levels of total and active protein S in patients with bronchial asthma and healthy controls.. The circulating level of total protein S and of its active form was significantly decreased in patients with bronchial asthma compared to controls. Allergic protein S transgenic mice showed a significant reduction of airway hyperresponsiveness, lung tissue inflammatory cell infiltration, lung levels of Th2 cytokines and IgE compared to their wild-type counterparts. Administration of exogenous human protein S also decreased airway hyperresponsiveness and Th2-mediated lung inflammation in allergic wild-type mice compared with their untreated mouse counterparts. Human protein S significantly shifted the Th1/Th2 balance to Th1 and promoted the secretion of Th1 cytokines (IL-12, tumor necrosis factor-α) from dendritic cells.. These observations suggest the strong protective activity of protein S against the development of allergic bronchial asthma implicating its potential usefulness for the disease treatment.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Humans; Immunoglobulin E; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Protein S; Th2 Cells

Androgen levels inversely correlate with the incidence, susceptibility and severity of asthma. However, whether male sex hormones such as 5α-dihydrotestosterone (DHT) have beneficial effects on asthma symptoms and/or could affect asthma susceptibility have not been investigated. DHT administration to female mice, during the sensitization phase, abrogates the sex bias in bronchial hyperreactivity. This effect correlates with inhibition of leukotriene biosynthesis in the lung. DHT significantly inhibits also other asthma-like features such as airway hyperplasia and mucus production in sensitized female mice. Conversely, DHT does not affect plasma IgE levels as well as CD3

Topics: Androgens; Animals; Asthma; Bronchial Hyperreactivity; Cell Line; Dihydrotestosterone; Female; Immunologic Factors; Male; Mast Cells; Mice; Mice, Inbred BALB C; Ovalbumin; Sex Characteristics

Titanium dioxide nanoparticles (TiO. According to the results, the mice receiving OVA alone or OVA plus TiO. Based on the findings of the current study, intranasally or inhalation exposure to high-dose nanosized TiO

Topics: Administration, Intranasal; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Eosinophils; Female; Inhalation Exposure; Mice; Mice, Inbred BALB C; Nanoparticles; Ovalbumin; Respiratory Function Tests; T-Lymphocytes, Helper-Inducer; Titanium

Asthma is a common chronic inflammatory airway disease. Recent studies have reported that interleukin (IL)‑33 is a potential link between the airway epithelium and Th2‑type inflammatory responses, which are closely related to the progression of asthma. The IL‑33 receptor, ST2, is highly expressed in group 2 innate lymphoid cells (ILC2s), Th2 cells, mast cells, eosinophils and natural killer (NK) cells. Cnidii Fructus is a Chinese herb with a long history of use in the treatment of asthma in China. Osthole is one of the major components of Cnidii Fructus. The present study examined the anti‑asthmatic effects of osthole in mice and aimed to elucidate the underlying mechanisms involving the IL‑33/ST2 pathway. BALB/c mice were sensitized and challenged with ovalbumin and then treated with an intraperitoneal injection of osthole (25 and 50 mg/kg). Subsequently, the airway hyper‑responsiveness (AHR) and inflammation of the lungs were evaluated. The amounts of IL‑4, IL‑5, IL‑13, interferon (IFN)‑γ and IL‑33 in the bronchoalveolar lavage fluid (BALF) were measured by Luminex assay and their mRNA levels in the lungs were measured by reverse transcription‑quantitative PCR. The histopathology of the lungs was performed with H&E, PAS and Masson's staining. The expression of ST2 in the lungs was evaluated by immunohistochemistry. The data demonstrated that osthole markedly reduced AHR and decreased the number of eosinophils and lymphocytes in BALF. It was also observed that osthole significantly inhibited the release of Th2‑type cytokines (IL‑4, IL‑5 and IL‑13) and upregulated the IFN‑γ level in BALF. Moreover, osthole significantly attenuated the IL‑33 and ST2 expression in the lungs of asthmatic mice. On the whole, osthole attenuated ovalbumin‑induced lung inflammation through the inhibition of IL‑33/ST2 signaling in an asthmatic mouse model. These results suggest that osthole is a promising target for the development of an asthma medication.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Coumarins; Disease Models, Animal; Drugs, Chinese Herbal; Female; Gene Expression Regulation; Inflammation; Interferon-gamma; Interleukin-1 Receptor-Like 1 Protein; Interleukin-33; Interleukins; Lung; Lymphocyte Count; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Pulmonary Eosinophilia; Random Allocation; RNA, Messenger; Signal Transduction

Asthma is an inflammatory disease of the airways of the lungs, which is characterized by airflow obstruction and bronchospasms. Glabridin is a major flavonoid, especially found in root of Glycyrrhiza glabra, and has several pharmacological activities, including antioxidant and anti-inflammatory effects. The anti-asthmatic effect and possible mechanism of glabridin, however, have not been revealed so far. The aim of this study is to investigate the effects and possible mechanisms of glabridin against ovalbumin (OVA)-induced airway hyperresponsiveness (AHR) and inflammation in mice. In male BALB/c mice, asthma was induced by intraperitoneal (i.p) injection of OVA mixed with 2 mg aluminium hydroxide on days 0, 14 and boosted with OVA aerosol challenge on days 21, 22, and 23. Mice were either treated with dexamethasone (i.p, 1 mg/kg) or glabridin (10, 20, and 30 mg/kg) from days 18-23. Pulmonary function parameters such as peak inspiratory flow, peak expiratory flow, tidal volume, expiratory volume, the frequency of breathing, enhanced pause values were evaluated by using whole-body plethysmography. Measurements were performed at baseline and following methacholine (50 mg/mL) challenges. In addition, white blood cells (WBC) count, total protein, and IgE levels were measured in bronchial alveolar lavage fluid (BALF), lung, and serum, respectively. Glabridin (20 or 30 mg/kg) significantly attenuated (p < 0.05) OVA-induced alteration in respiratory parameters. Elevated counts of total WBC, differential WBC (neutrophils, lymphocytes, monocytes, and eosinophils) in BALF and the total protein in lungs and BALF were significantly decreased (p < 0.05) by glabridin (20 or 30 mg/kg). It also significantly attenuated the increased serum IgE levels (p < 0.05). As glabridin reduces the level of serum IgE, the total protein and the count of WBC and improves respiratory function, it may be a novel therapeutic agent in asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Inflammation; Isoflavones; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Phenols

Topics: Animals; Anti-Inflammatory Agents; Asthma; beta-N-Acetylhexosaminidases; Body Weight; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Capsaicin; Cordyceps; Cytokines; Disease Models, Animal; Eosinophil Peroxidase; Female; Histamine Release; Immunization; Immunoglobulin E; Mast Cells; Methacholine Chloride; Mice, Inbred BALB C; Mycelium; Nasal Lavage; Ovalbumin; Rats, Sprague-Dawley; Rhinitis, Allergic; Skin; Spleen; Trachea

Asthma is a leading allergic disease worldwide, demonstrating an ever‑increasing prevalence over the past two decades. Asthma is characterized by allergen‑associated airway hyperresponsiveness (AHR) that primarily results from T helper 2 (Th2) cell inflammation, in which dendritic cells (DCs) serve an important role in determining T cell development after encountering an antigen. Atractylodin (ATL), a polyethene alkyne extracted from Atractylodis rhizoma (also known as Cangzhu), has proven effective in treating digestive disorders, rheumatic disease and influenza. In addition, ATL was discovered to alleviate mouse collagen‑induced arthritis via regulating DC maturation. The present study aimed to investigate the effect of ATL on asthma given that DCs serve an essential role in Th2‑mediated inflammation in asthma. Mouse model of asthma was induced by ovalbumin (OVA). OVA‑induced airway hyperresponsiveness (AHR) and inflammatory cells in bronchoalveolar lavage fluid (BALF) were detected. The production of IgE and IgG1 in serum and cytokines in BALF were detected by ELISA. The effects of ATL on dendritic cells maturation and T cell expansion were detected by flow cytometry analysis and 3H‑thymidine incorporation. Using a model of OVA‑induced asthma, it was demonstrated that ATL ameliorated AHR and decreased the levels of IL‑4, IL‑5 and IL‑13 in bronchoalveolar lavage fluid (BALF), and OVA‑specific IgE and IgG1 in the serum. OVA‑stimulated splenocytes were used to demonstrated that ATL decreased cell expansion and the production of IL‑4, IL‑5 and IL‑13 in the culture medium. In order to determine the cellular mechanism of ATL in asthma, splenic DCs were isolated and it was subsequently observed that ATL downregulated the expression levels of CD40 and CD80. Furthermore, OVA‑stimulated CD4+ T cells were co‑cultured with splenic DCs, which revealed that ATL‑treated splenic DCs led to impaired cellular proliferation and the production of IL‑4, IL‑5 and IL‑13 in OVA‑stimulated T cells. In conclusion, these results indicated that ATL may suppress antigen‑specific Th2 responses in an OVA‑induced allergic asthma model via regulating DCs. Therefore, ATL may exhibit therapeutic potential in the management of asthma and other allergic diseases presenting with Th2 inflammation.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; China; Cytokines; Dendritic Cells; Disease Models, Animal; Furans; Immunoglobulin E; Immunologic Factors; Inflammation; Interleukin-13; Interleukin-4; Interleukin-5; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes, Regulatory; Th2 Cells

B cells could convert naïve T cells into regulatory T cells (so-called Treg-of-B cells) which have the ability to treat animal models of inflammatory diseases, including allergic asthma, collagen-induced arthritis and colitis; however, the mechanisms of Treg-of-B cell generation remain unclear. In this study, we investigated the role of STAT6 in the generation of Treg-of-B (P) cells, which Treg cells were generated by Peyer's patch B cells (P stands for Peyer's patch). CD4+CD25- T cells from wild type, STAT6 knockout and IL-4 knockout mice were cocultured with wild type Peyer's patch B cells for Treg-of-B (P) cell generation. A murine asthmatic model was used to analyze the

Topics: Adoptive Transfer; Animals; Antigens, CD; Apoptosis; Apoptosis Regulatory Proteins; B-Lymphocytes; Bronchial Hyperreactivity; Coculture Techniques; Cytokines; Disease Models, Animal; Gene Expression Regulation; Interleukin-4; Lung; Lymphocyte Activation Gene 3 Protein; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Peyer's Patches; Phosphorylation; Protein Processing, Post-Translational; STAT Transcription Factors; STAT6 Transcription Factor; T-Lymphocytes, Regulatory

The respiratory system is exposed to various allergens via inhaled and intranasal routes. Murine models of allergic lung disease have been developed to clarify the mechanisms underlying inflammatory responses and evaluate the efficacy of novel therapeutics. However, there have been no comparative studies on differences in allergic phenotypes following inhaled vs. intranasal allergen challenge. In this study, we compared the asthmatic features of mice challenged via different routes following allergen sensitization and investigated the underlying mechanisms.. To establish ovalbumin (OVA)-induced allergic asthma models, BALB/c mice were sensitized to 20 μg OVA with 1 mg aluminum hydroxide by the intraperitoneal route and then challenged by inhalation or intranasal administration with 5% OVA for 3 consecutive days. Cellular changes and immunoglobulin (Ig) E levels in bronchoalveolar lavage fluid (BALF) and serum, respectively, were assessed. Histological changes in the lungs were examined by hematoxylin and eosin (H&E) and periodic acid Schiff (PAS) staining. Levels of T helper (Th)2 cytokines including interleukin (IL)-4, -5, and -13 in BALF and epithelial cytokines including IL-25 and -33 in BALF and lung tissues were measured by enzyme-linked immunosorbent assay and western blotting. Airway hyperresponsiveness (AHR) was evaluated by assessing airway resistance (Rrs) and elastance (E) via an invasive method.. OVA-sensitized and challenged mice showed typical asthma features such as airway inflammation, elevated IgE level, and AHR regardless of the challenge route. However, H&E staining showed that inflammation of pulmonary vessels, alveolar ducts, and alveoli were enhanced by inhaled as compared to intranasal OVA challenge. PAS staining showed that intranasal OVA challenge induced severe mucus production accompanied by inflammation in bronchial regions. In addition, Th2 cytokine levels in BALF and AHR in lung were increased to a greater extent by inhalation than by intranasal administration of OVA. Epithelial cytokine expression, especially IL-25, was increased in the lungs of mice in the inhaled OVA challenge group.. OVA-sensitized mice exhibit different pathophysiological patterns of asthma including expression of epithelial cell-derived cytokines depending on the OVA challenge route. Thus, some heterogeneous phenotypes of human asthma can be replicated by varying the mode of delivery after OVA sensitization.

Topics: Administration, Inhalation; Administration, Intranasal; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Interleukin-17; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Phenotype; T-Lymphocytes

Allergic asthma causes morbidity in many subjects, and novel precision-directed treatments would be valuable.. We sought to examine the role of a novel innate molecule, repulsive guidance molecule b (RGMb), in murine models of allergic asthma.. In models of allergic asthma using ovalbumin or cockroach allergen, mice were treated with anti-RGMb or control mAb and examined for airway inflammation and airway hyperreactivity (AHR), a cardinal feature of asthma. The mechanisms by which RGMb causes airways disease were also examined.. We found that blockade of RGMb by treatment with anti-RGMb mAb effectively blocked the development of airway inflammation and AHR. Importantly, blockade of RGMb completely blocked the development of airway inflammation and AHR, even if treatment occurred only during the challenge (effector) phase. IL-25 played an important role in these models of asthma because IL-25 receptor-deficient mice did not develop disease after sensitization and challenge with allergen. RGMb was expressed primarily by innate cells in the lungs, including bronchial epithelial cells (known producers of IL-25), activated eosinophils, and interstitial macrophages, which in the inflamed lung expressed the IL-25 receptor and produced IL-5 and IL-13. We also found that neogenin, the canonical receptor for RGMb, was expressed by interstitial macrophages and bronchial epithelial cells in the inflamed lung, suggesting that an innate RGMb-neogenin axis might modulate allergic asthma.. These results demonstrate an important role for a novel innate pathway in regulating type 2 inflammation in patients with allergic asthma involving RGMb and RGMb-expressing cells, such as interstitial macrophages and bronchial epithelial cells. Moreover, targeting this previously unappreciated innate pathway might provide an important treatment option for allergic asthma.

Topics: Allergens; Animals; Antibodies, Monoclonal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Adhesion Molecules, Neuronal; Cockroaches; Female; Interleukin-1 Receptor-Like 1 Protein; Macrophages; Membrane Proteins; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Programmed Cell Death 1 Ligand 2 Protein; Receptors, Interleukin

Topics: Animals; Bronchial Hyperreactivity; Chickens; Dust; Inhalation Exposure; Male; Mice; Mice, Inbred C57BL; Organic Agriculture; Ovalbumin; Swine

Glucagon has been shown to be beneficial as a treatment for bronchospasm in asthmatics. Here, we investigate if glucagon would prevent airway hyperreactivity (AHR), lung inflammation, and remodeling in a murine model of asthma. Glucagon (10 and 100 µg/Kg, i.n.) significantly prevented AHR and eosinophilia in BAL and peribronchiolar region induced by ovalbumin (OVA) challenge, while only the dose of 100 µg/Kg of glucagon inhibited subepithelial fibrosis and T lymphocytes accumulation in BAL and lung. The inhibitory action of glucagon occurred in parallel with reduction of OVA-induced generation of IL-4, IL-5, IL-13, TNF-α, eotaxin-1/CCL11, and eotaxin-2/CCL24 but not MDC/CCL22 and TARC/CCL17. The inhibitory effect of glucagon (100 µg/Kg, i.n.) on OVA-induced AHR and collagen deposition was reversed by pre-treatment with indomethacin (10 mg/Kg, i.p.). Glucagon increased intracellular cAMP levels and inhibits anti-CD3 plus anti-CD28-induced proliferation and production of IL-2, IL-4, IL-10, and TNF- α from TCD4

Topics: Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cell Proliferation; Chemokine CCL24; Cytokines; Glucagon; Lung; Mice, Inbred Strains; Ovalbumin; Pneumonia; Receptors, Glucagon

Asthma is a chronic inflammatory disease that involves a variety of cytokines and cells. Interleukin-16 (IL-16) is highly expressed during allergic airway inflammation and is involved in its development. However, its specific mechanism of action remains unclear. In the present study, we used an animal model of ovalbumin (OVA)-induced allergic asthma with mice harboring an IL-16 gene deletion to investigate the role of this cytokine in asthma, in addition to its underlying mechanism. Increased IL-16 expression was observed during OVA-induced asthma in C57BL/6J mice. However, when OVA was used to induce asthma in IL-16

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Disease Models, Animal; Female; Immunoglobulin E; Interleukin-16; Lung; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Signal Transduction; Spleen; Th17 Cells; Th2 Cells

Ovalbumin (OVA) sensitization has limitations in modelling asthma. Thus, we examined the value of allergic sensitization using a purified natural allergen, house dust mite (HDM), over the sensitization performed with OVA. Mice were sham-treated, or sensitized with OVA- or HDM with identical chronology. Airway resistance, tissue damping and elastance were assessed under control conditions and after challenging the animals with methacholine (MCh) and the specific allergen. Inflammatory profile of the bronchoalveolar lavage fluid was characterized and lung histology was performed. While no difference in the lung responsiveness to the specific allergen was noted, hyperresponsiveness to MCh was observed only in the HDM-sensitized animals in the lung peripheral parameters. Lung inflammation differed between the models, but excessive bronchial smooth muscle remodelling occurred only with OVA. In conclusion, we demonstrate that a purified natural allergen offers a more relevant murine model of human allergic asthma by expressing the key features of this chronic inflammatory disease both in the lung function and structure.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Lung; Methacholine Chloride; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Pneumonia; Pyroglyphidae; Respiratory Mechanics

V-domain immunoglobulin suppressor of T-cell activation (VISTA) is a novel immune checkpoint receptor and ligand that regulates T-cell activation. We investigated the functional involvement of VISTA in Th2 cell-mediated immune responses using an ovalbumin (OVA)-induced allergic asthma model. Treatment with an anti-VISTA monoclonal antibody (mAb) during allergen sensitization increased the production of antibodies, including total IgE, OVA-specific IgG1 and IgG2a and allergen-specific IL-5 and IL-13; it also increased the expression of IL-13 by splenic CD4+ T cells. However, treatment with the anti-VISTA mAb during sensitization did not accelerate asthmatic responses, including airway hyper-responsiveness (AHR) or the number of eosinophils in bronchoalveolar lavage (BAL) fluid. In contrast, treatment with the anti-VISTA mAb during allergen challenge significantly augmented AHR and BAL fluid eosinophilia. This treatment also increased the production of IL-5 and IL-13 in BAL fluid and the expression of IL-13 by CD4+ T cells in draining lymph nodes. These results suggest that VISTA is involved in the regulation of Th2 cell generation and Th2 cell-mediated antibody production and regulates asthmatic responses, especially in the effector phase.

Topics: Allergens; Animals; Antibodies, Monoclonal; Antigen-Antibody Reactions; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Disease Models, Animal; Eosinophils; Female; Flow Cytometry; Membrane Proteins; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells

Uncaria tomentosa (Willd. Ex Schult) DC is used by indigenous tribes in the Amazonian region of Central and South America to treat inflammation, allergies and asthma. The therapeutic properties of U. tomentosa have been attributed to the presence of tetracyclic and pentacyclic oxindole alkaloids and to phenolic acids.. To characterize aqueous bark extracts (ABE) and aqueous leaf extracts (ALE) of U. tomentosa and to compare their anti-inflammatory effects.. Constituents of the extracts were identified by ultra performance liquid chromatography-mass spectrometry. Anti-inflammatory activities were assessed in vitro by exposing lipopolysaccharide-stimulated macrophage cells (RAW264.7-Luc) to ABE, ALE and standard mitraphylline. In vivo assays were performed using a murine model of ovalbumin (OVA)-induced asthma. OVA-sensitized animals were treated with ABE or ALE while controls received dexamethasone or saline solution. Bronchial hyperresponsiveness, production of Th1 and Th2 cytokines, total and differential counts of inflammatory cells in the bronchoalveolar lavage (BAL) and lung tissue were determined.. Mitraphylline, isomitraphylline, chlorogenic acid and quinic acid were detected in both extracts, while isorhyncophylline and rutin were detected only in ALE. ABE, ALE and mitraphylline inhibited the transcription of nuclear factor kappa-B in cell cultures, ALE and mitraphylline reduced the production of interleukin (IL)-6, and mitraphylline reduced production of tumor necrosis factor-alpha. Treatment with ABE and ALE at 50 and 200 mg kg. The results clarify for the first time the anti-inflammatory activity of U. tomentosa in a murine model of asthma. Although ABE and ALE exhibited distinct chemical compositions, both extracts inhibited the production of pro-inflammatory cytokines in vitro. In vivo assays revealed that ABE was more effective in treating asthmatic inflammation while ALE was more successful in controlling respiratory mechanics. Both extracts may have promising applications in the phytotherapy of allergic asthma.

Topics: Acids, Carbocyclic; Allergens; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cat's Claw; Cell Survival; Cytokines; Disease Models, Animal; Indole Alkaloids; Lung; Mice; Ovalbumin; Phytotherapy; Plant Bark; Plant Extracts; Plant Leaves; RAW 264.7 Cells

The purpose of this study is to explore the influence of ozone repeated short exposures on airway/lung inflammation, airway hyperresponsiveness (AHR) and airway hypersecretion in ovalbumin (OVA) sensitized/challenged asthmatic mouse model.. OVA sensitization was performing by intraperitoneal injection. Ozone exposures (3ppm for 3hours) were given one hour after aerosolized OVA challenges (once every other day, 4 times totally). Methacholine (MCH) bronchial provocation tests, Liu's staining of BALF cell smears, hematoxylin-eosin (HE) staining and Periodic Acid-Schiff (PAS) staining of lung tissue were performed. Interleukins (ILs; IL-4, IL-13, IL-1β, and IL-18) protein (ELISA) and mRNA expression levels (RT-qPCR) in murine lung, 8-hydroxy-2'-deoxyguanosine (8-OHdG, ELISA), malondialdehyde (MDA, thiobarbituric acid assay), reduced glutathione (GSH, spectrophotometric method) in bronchoalveolar lavage fluid (BALF), and GSH1 mRNA relative expression levels (RT-qPCR) in lung tissue were analyzed.. Repeated ozone exposures down-regulated the AHR to MCH in mice undergoing OVA sensitization and challenge, however not all parameters associated with asthma were decreased since obvious mucus hypersecretion was induced and airway inflammation increased slightly, especially around small airways. Following ozone co-exposure, the increase of IL-4 and IL-13 levels in murine lung caused by OVA sensitization/challenge were reversed. Instead, levels of IL-1β in BALF remained, higher than negative control group. Ozone repeated short exposures also induced significant increase of 8-OHdG in BALF in OVA sensitized and challenged mice.. For asthmatic mice undergoing ozone exposures, AHR is not an accurate indicator of the severity of asthma. Repeated short ozone exposures increase mucus hypersecretion, possibly via an increase in oxidative stress and immune dysregulation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Interleukin-13; Interleukin-1beta; Interleukin-4; Male; Mice; Mice, Inbred C57BL; Mucus; Ovalbumin; Oxidative Stress; Ozone; Pneumonia; Reverse Transcriptase Polymerase Chain Reaction; Time Factors

Asthma is a common chronic airway inflammation disease and is considered as a major public health problem. Fisetin (3,3',4',7-tetrahydroxyflavone) is a naturally occurring flavonoid abundantly found in different vegetables and fruits. Fisetin has been reported to exhibit various positive biological effects, including anti-proliferative, anticancer, anti-oxidative and neuroprotective effects. We evaluated the effects of fisetin on allergic asthma regulation in mice. Mice were first sensitized, then airway-challenged with ovalbumin (OVA). Whether fisetin treatment attenuated OVA-induced airway inflammation was examined via inflammation inhibition through MyD88-related NF-κB (p65) signaling pathway. Mice were divided into the control (Con), OVA-induced asthma (Mod), 40 (FL) and 50 (FH) mg/kg fisetin-treated OVA-induced asthma groups. Our results found that OVA-induced airway inflammation in mice caused a significant inflammatory response via the activation of MyD88 and NF-κB signaling pathways, leading to release of pro-inflammatory cytokines. In contrast, fisetin-treated mice after OVA induction inhibited activation of MyD88 and NF-κB signaling pathways, resulting in downregulation of pro-inflammatory cytokine secretion. Further, fisetin significantly ameliorated the airway hyperresponsiveness (AHR) towards methacholine (Mch). In addition, fisetin reduced the number of eosinophil, monocyte, neutrophil and total white blood cell in the bronchoalveolar lavage fluid (BALF) of OVA-induced mice. The serum and BALF samples obtained from the OVA-induced mice with fisetin showed lower levels of pro-inflammatory cytokines. The results of our study illustrated that fisetin may be a new promising candidate to inhibit airway inflammation response induced by OVA.

Topics: Airway Resistance; Animals; Asthma; Bronchial Hyperreactivity; Cell Count; Cytokines; Disease Models, Animal; Flavonoids; Flavonols; Inflammation Mediators; Lipopolysaccharides; Lung; Male; Mice, Inbred C57BL; Myeloid Differentiation Factor 88; NF-kappa B; Ovalbumin; Pneumonia; Signal Transduction; Toll-Like Receptor 5

Ras, a small GTPase protein, is thought to mediate Th2-dependent eosinophilic inflammation in asthma. Ras requires cell membrane association for its biological activity, and this requires the posttranslational modification of Ras with an isoprenyl group by farnesyltransferase (FTase) or geranylgeranyltransferase (GGTase). We hypothesized that inhibition of FTase using FTase inhibitor (FTI)-277 would attenuate allergic asthma by depleting membrane-associated Ras. We used the OVA mouse model of allergic inflammation and human airway epithelial (HBE1) cells to determine the role of FTase in inflammatory cell recruitment. BALB/c mice were first sensitized then exposed to 1% OVA aerosol or filtered air, and half were injected daily with FTI-277 (20 mg/kg per day). Treatment of mice with FTI-277 had no significant effect on lung membrane-anchored Ras, Ras protein levels, or Ras GTPase activity. In OVA-exposed mice, FTI-277 treatment increased eosinophilic inflammation, goblet cell hyperplasia, and airway hyperreactivity. Human bronchial epithelial (HBE1) cells were pretreated with 5, 10, or 20 μM FTI-277 prior to and during 12 h IL-13 (20 ng/ml) stimulation. In HBE1 cells, FTase inhibition with FTI-277 had no significant effect on IL-13-induced STAT6 phosphorylation, eotaxin-3 peptide secretion, or Ras translocation. However, addition of exogenous FPP unexpectedly augmented IL-13-induced STAT6 phosphorylation and eotaxin-3 secretion from HBE1 cells without affecting Ras translocation. Pharmacological inhibition of FTase exacerbates allergic asthma, suggesting a protective role for FTase or possibly Ras farnesylation. FPP synergistically augments epithelial eotaxin-3 secretion, indicating a novel Ras-independent farnesylation mechanism or direct FPP effect that promotes epithelial eotaxin-3 production in allergic asthma.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Disease Models, Animal; Enzyme Inhibitors; Eosinophils; Epithelial Cells; Farnesyltranstransferase; Humans; Inflammation; Lung; Male; Methionine; Mice; Mice, Inbred BALB C; Ovalbumin; Polyisoprenyl Phosphates; ras Proteins; Sesquiterpenes; Signal Transduction

To examine the effect of metanol extract of Petiveria alliacea (PM) on airway inflflammation in a murine model of chronic asthma.. Two-month-old male BALB/c mice (n=6-8/group) were sensitized on days 0 and 14 by intraperitoneal injection of 20 μg ovalbumin (OVA). On day 25, the mice received an airway challenge with OVA (3%, w/v, in phosphate buffered saline). PM was administered orally by oral gavage to mice at doses of 100, 200 and 400 mg/kg body weight once daily from days 18 to 23. Control mice were orally administered phosphate buffered saline (PBS) to induce a model of asthma. At the end of the test, respiratory reactivity was assayed, the total cell number, interleukin-4 (IL-4), IL-5, IL-13, tumor necrosis factor-alpha (TNF-α) and reactive oxygen species (ROS) in the bronchoalveolar lavage fluid (BALF) were determined and the levels of serum IgE, intercellular cell adhesion molecule 1 (ICAM-1) and eotoxin were measured. In addition, lung tissue was used to qualify the IL-4, IL-5, IL-13, TNF-α and transforming growth factor beta 1 (TGF-β1). Histologic examination was performed to observe inflammatory cellular infiltration.. The administration of PM in comparison with the OVA-only treated group signifificantly attenuated the infifiltration of eosinophils and other inflflammatory cells (P<0.01). Airway resistance (RI) in the OVA-only induced group was significantly higher than that of the PBS control group (P<0.01) when methacholine was added. TNF-α, IgE, TGF-β1 and cytokine levels IL-4, IL-5, IL-13 in the BALF decreased compared to control mice (P<0.01 or P<0.05). PM treatment also inhibited the production of chemokines, eotaxin and ICAM-1 in BALF (P<0.01), which improved lung function. Histopathological examination revealed that the sensitized treated PM groups had significant lower in inflammatory scores similar to dexamethasone treatments and the untreated group.. Administration of PM could inhibit airway inflammation, regulate cytokines, chemokines and enhance pulmonary conditions in allergic murine model of asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Disease Models, Animal; Immunoglobulin E; Inflammation; Lung; Male; Methanol; Mice, Inbred BALB C; Mucus; Ovalbumin; Phytolaccaceae; Phytotherapy; Plant Extracts; Reactive Oxygen Species; Th2 Cells

Ovalbumin-induced allergic lung inflammation (ALI) is a condition believed to be mediated by cytokines, extracellular matrix remodeling, and redox imbalance. In this study, we evaluated pulmonary function together with inflammatory markers as interleukin-4 (IL-4), myeloperoxidase (MPO), eosinophil cells, and redox markers in the lungs of BALB/c mice after ovalbumin (OVA) sensitization and challenge. Our results showed an increase in bronchial hyperresponsiveness stimulated by methacholine (Mch), inflammatory cell influx, especially eosinophils together with an increase of high mobility group box 1 (HMGB1) and altered lipid peroxidation (LP) and antioxidant defenses in the OVA group compared to the control group (p ≤ 0.5). Thus, we demonstrated that OVA-induced ALI altered redox status concomitantly with impaired lung function, which was associated with HMGB1 expression and proteolytic remodeling. Taken together all results found here, we may suggest HMGB1 is an important therapeutic target for asthma, once orchestrates the redox signaling, inflammation, and remodeling that contribute to the disease development.

Topics: Animals; Asthma; Biomarkers; Bronchial Hyperreactivity; Eosinophils; HMGB1 Protein; Inflammation; Lipid Peroxidation; Mice; Mice, Inbred BALB C; Ovalbumin; Oxidative Stress; Proteolysis

Lipopolysaccharide (LPS) contributes to asthma exacerbations and development of inhaled corticosteroid insensitivity. Complete resistance to systemic corticosteroids is rare, and most patients lie on a continuum of steroid responsiveness. This study aimed to examine the sensitivity of combined ovalbumin- (Ova) and LPS-induced functional and inflammatory responses to inhaled and systemic corticosteroid in conscious guinea pigs to test the hypothesis that the route of administration affects sensitivity. Guinea pigs were sensitized to Ova and challenged with inhaled Ova alone or combined with LPS. Airway function was determined by measuring specific airway conductance via whole-body plethysmography. Airway hyper-responsiveness to histamine was determined before and 24 hours post-Ova challenge. Airway inflammation and underlying mechanisms were determined from bronchoalveolar lavage cell counts and lung tissue cytokines. Vehicle or dexamethasone was administered by once-daily i.p. injection (5, 10, or 20 mg/kg) or twice-daily inhalation (4 or 20 mg/ml) for 6 days before Ova challenge or Ova with LPS. LPS exacerbated Ova-induced responses, elongating early asthmatic responses (EAR), prolonging histamine bronchoconstriction, and further elevating airway inflammation. Intraperitoneal dexamethasone (20 mg/kg) significantly reduced the elongated EAR and airway inflammation but not the increased bronchoconstriction to histamine. In contrast, inhaled dexamethasone (20 mg/ml), which inhibited responses to Ova alone, did not significantly reduce functional and inflammatory responses to combined Ova and LPS. Combined Ova and LPS-induced functional and inflammatory responses are insensitive to inhaled, but they are only partially sensitive to systemic, dexamethasone. This finding suggests that the route of corticosteroid administration may be important in determining corticosteroid sensitivity of asthmatic responses.

Topics: Administration, Inhalation; Adrenal Cortex Hormones; Animals; Asthma; Bronchial Hyperreactivity; Drug Administration Routes; Drug Combinations; Guinea Pigs; Injections, Intraperitoneal; Lipopolysaccharides; Male; Ovalbumin; Respiratory Hypersensitivity

Epidemiological evidence suggests that formaldehyde (FA) exposure may influence the prevalence and severity of allergic asthma. However, the role of genetic background in FA-induced asthma-like responses is poorly understood. In the present study, we investigated the nature and severity of asthma-like responses triggered by exposure to different doses of FA together with or without ovalbumin (OVA) in two genetically different mouse strains-BALB/c and C57BL/6. Both mouse strains were divided into two main groups: the non-sensitized group and the OVA-sensitized group. All the groups were exposed to 0, 0.5 or 3.0 mg/m3 FA for 6 h/day over 25 consecutive days. At 24 h after the final FA exposure, the pulmonary parameters were evaluated. We found that FA exposure induced Th2-type allergic responses in non-sensitized BALB/c and C57BL/6 mice. In addition, FA-induced allergic responses were significantly more prominent in BALB/c mice than in C57BL/6 mice. In sensitized BALB/c mice, however, FA exposure suppressed the development of OVA-induced allergic responses. Exposure to 3.0 mg/m3 FA in sensitized C57BL/6 mice also led to suppressed allergic responses, whereas exposure to 0.5 mg/m3 FA resulted in exacerbated allergic responses to OVA. Our findings suggest that FA exposure can induce differential airway inflammation and bronchial hyperresponsiveness in BALB/c and C57BL/6 mice.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Formaldehyde; Humans; Inflammation; Inhalation Exposure; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Respiratory Hypersensitivity; Th2 Cells

Topics: Animals; Antibodies; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Differentiation; Cell Polarity; Female; Hypersensitivity; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Pneumonia; Receptors, Tumor Necrosis Factor, Type II; Signal Transduction; Th17 Cells; Th2 Cells; Tumor Necrosis Factor-alpha

Asthma is associated with exposure to a wide variety of allergens and adjuvants. The extent to which overlap exists between the cellular and molecular mechanisms triggered by these various agents is poorly understood, but it might explain the differential responsiveness of patients to specific therapies. In particular, it is unclear why some, but not all, patients benefit from blockade of TNF. Here, we characterized signaling pathways triggered by distinct types of adjuvants during allergic sensitization. Mice sensitized to an innocuous protein using TLR ligands or house dust extracts as adjuvants developed mixed eosinophilic and neutrophilic airway inflammation and airway hyperresponsiveness (AHR) following allergen challenge, whereas mice sensitized using proteases as adjuvants developed predominantly eosinophilic inflammation and AHR. TLR ligands, but not proteases, induced TNF during allergic sensitization. TNF signaled through airway epithelial cells to reprogram them and promote Th2, but not Th17, development in lymph nodes. TNF was also required during the allergen challenge phase for neutrophilic and eosinophilic inflammation. In contrast, TNF was dispensable for allergic airway disease in a protease-mediated model of asthma. These findings might help to explain why TNF blockade improves lung function in only some patients with asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cell Differentiation; Cytokines; Disease Models, Animal; Eosinophils; Hypersensitivity; Inflammation; Interleukin-17; Ligands; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; Ovalbumin; Respiratory Hypersensitivity; Signal Transduction; Th17 Cells; Th2 Cells; Toll-Like Receptors; Tumor Necrosis Factor-alpha

Genipin is a natural compound isolated from the fruit of Gardenia jasminoides with various pharmacological effects. In this study, we investigated whether genipin effectively alleviates allergic responses in a murine model of ovalbumin (OVA)-induced asthma. The mice were administered an intraperitoneal injection of OVA on day 0 and 14 to boost the immune response; genipin was then administered from day 18 to 23 by oral gavage. On days 21 to 23, mice were OVA-challenged using am ultrasonic nebulizer, and airway hyperresponsiveness (AHR) was determined on day 24 by plethysmography. Genipin significantly reduced the inflammatory cell count in bronchoalveolar lavage fluids (BALF) and AHR, which were accompanied by lower interleukin-5 (IL-5), IL-13 and OVA-specific immunoglobulin (Ig) E levels in the BALF or serum from OVA-induced asthmatic mice. In histology, genipin significantly decreased airway inflammation and mucus hypersecretion in OVA-induced asthmatic mice. Additionally, genipin inhibited OVA-induced increases in the expression of inducible nitric oxide synthase and cyclooxygenase-2 proteins. Further, genipin reduced the activity and protein levels of matrix metalloproteinase-9 in lung tissue from OVA induced asthmatic mice. Overall, genipin effectively alleviated the asthmatic inflammatory response in an OVA-induced asthmatic model. Therefore, our results suggest that genipin has therapeutic potential for treating asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Gardenia; Hypersensitivity; Immunoglobulin E; Inflammation; Interleukin-13; Interleukin-5; Iridoids; Lung; Matrix Metalloproteinase 9; Mice; Mice, Inbred BALB C; Nitric Oxide Synthase Type II; Ovalbumin

Selaginella uncinata (Desv.) Spring, known as "Cuiyuncao", is a perennial herb widely distributed in the Southeast Asian countries. In the folk medicine, the local minority commonly use it to treat cough and asthma for centuries.. This study was carried out to investigate the protective mechanisms of total flavonoids from S. uncinata (SUF) on airway hyperresponsiveness, cytokine release and bitter taste receptors (T2Rs) signaling with emphasis on inflammatory responses in a rat model of ovalbumin (OVA)-induced asthma.. Rats were sensitized and challenged with OVA to induce typical asthmatic reactions. Pathological changes of lung tissue were examined by HE staining. The serum levels of T cell-associated cytokines (IFN-γ, IL-4, IL-5 and IL-13), total IgE and OVA-specific IgE were determined by enzyme-linked immunosorbent assay (ELISA). Gene expressions of T2R10, IP3R1 and Orai1 in lung tissue were assayed by fluorescence quantitative real-time polymerase chain reaction (FQ-PCR) while protein expressions of NFAT1 and c-Myc were assayed by western blot analysis. The activation of SUF was investigated on tansgentic T2R10-GFP HEK293 cells.. SUF treatment attenuated airway hyperresponsiveness and goblet cell hyperplasia compared with OVA-challenged asthmatic rats. The serum levels of IL-4, IL-5 and IL-13 as well as total and OVA-specific IgE were decreased while serum IFN-γ was increased in SUF-treated rats. SUF treatment significantly up-regulated T2R10 gene expression, down-regulated IP3R1 and Orai1 gene expression. SUF further suppressed eotaxin, NFAT1 and c-Myc protein expression in lung tissues of OVA-challenged rats.. These results imply that SUF exerts anti-inflammatory function through the T2R10/IP3R1/NFAT1 dependent signaling pathway, and may warrant further evaluation as a possible agent for the treatment of asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Bronchodilator Agents; Cytokines; Disease Models, Animal; Flavonoids; HEK293 Cells; Humans; Immunoglobulin E; Inositol 1,4,5-Trisphosphate Receptors; Lung; Male; NFATC Transcription Factors; ORAI1 Protein; Ovalbumin; Phytotherapy; Plant Extracts; Plants, Medicinal; Proto-Oncogene Proteins c-myc; Rats, Sprague-Dawley; Receptors, G-Protein-Coupled; Selaginellaceae; Transfection

Eclipta prostrata (L.) L. (Asteraceae) has been used in Brazilian traditional medicine to treat asthma and other respiratory illnesses.. To investigate the effects of different doses of a standardized extract of E. prostrata using a murine model of allergen induced asthma.. Balb/c mice were sensitized twice with ovalbumin (OVA) administered intraperitoneally and challenged over four alternate days with nasal instillations of OVA solution. The standardized methanol extract of E. prostrata was administered in doses of 100, 250 and 500mgkg. The concentrations of chemical markers in the standardized methanol extract were 0.02% oroboside, 1.69% demethylwedelolactone and 1.71% wedelolactone. Treatment with 250mgkg. The results presented herein demonstrate for the first time the anti-inflammatory activity of E. prostrata in a murine model of asthma, thereby supporting the ethnopharmacological uses of the plant.

Topics: Airway Remodeling; Allergens; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Brazil; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Eclipta; Eosinophils; Male; Medicine, Traditional; Mice; Mice, Inbred BALB C; Ovalbumin; Plant Extracts; Th2 Cells

Mandevilla longiflora, popularly known as "velame" in central Brazil, is a subshrub widely distributed in South America. Its xylopodium is used in the form of a decoction or infusion to treat inflammation and other ailments.. This study aimed to evaluate the anti-inflammatory potential of M. longiflora in an in vivo model of ovalbumin-induced immediate hypersensitivity, identifying its effects on leukocyte infiltration, IgE and LTB. The hydroethanolic extract 70% of M. longiflora (HEMI) was obtained by maceration of the plant xylopodium. Swiss mice were sensitized by i.p. injection OVA-aluminium hydroxide on days 1 and 10. Nine days after the last sensitisation animals were challenged for 6 consecutive days with OVA solution for 20min daily in a closed chamber under continuous flow of aerosol. The animals were treated with HEMl (20, 50 and 200mg/kg p.o.), 2 times per day, and euthanized 24h later. Animals treated with vehicle (2% Tween-20) or dexamethasone were used as negative and positive controls, respectively. The recruitment of inflammatory cells into the pulmonary cavity was evaluated by counting cells present in broncho-alveolar lavage fluid (BALF). Lung tissue was also collected for histopathology and infiltration analysis. Quantification of IL-4, IL-5 and IL-13 from the BALF, and IgE, and LTB. The HEMl attenuated leukocyte migration into the airways, which was evidenced by a decrease in eosinophils, neutrophils and mononuclear cells, both in BALF quantification and by histopathological analysis, as well as decreasing the concentrations of IL-4, IL-5, IL-13, IgE and LTB

Topics: Animals; Anti-Inflammatory Agents; Apocynaceae; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Inflammation; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Plant Extracts

Leukotriene E4 (LTE4) that plays a key role in airway inflammation is expressed on platelets and eosinophils. We investigated whether blocking of the P2Y12 receptor can suppress eosinophilic inflammation in a mouse model of asthma because platelets and eosinophils share this receptor to be activated. BALB/c mice were sensitized by intraperitoneal injection of ovalbumin (OVA), followed by OVA nebulization. On each challenge day, clopidogrel, a P2Y12 antagonist was administered 30 min. before each challenge. Forty-eight hours after the last OVA challenge, mice were assessed for airway hyperresponsiveness (AHR), cell composition and cytokine levels, including chemokine ligand 5 (CCL5), in bronchoalveolar lavage (BAL) fluid. EOL cells were treated with LTE4, with or without clopidogrel treatment, and intracellular and extracellular eosinophil cationic protein (ECP) expressions were measured to find the inhibiting function of P2Y12 antagonist on eosinophilic activation. The levels of P2Y12 expression were increased markedly in the lung homogenates of OVA-sensitized and -challenged mice after platelet depletion. Administration of clopidogrel decreased AHR and the number of airway inflammatory cells, including eosinophils, in BAL fluid following OVA challenge. These results were associated with decreased levels of Th2 cytokines and CCL5. Histological examination showed that inflammatory cells as well as mucus-containing goblet cells were reduced in clopidogrel-administered mice compared to vehicle-treated mice. Clopidogrel inhibited extracellular ECP secretion after LTE4 stimulation in EOL-1 cells. Clopidogrel could prevent development of AHR and airway inflammation in a mouse model of asthma. P2Y12 can be a novel therapeutic target to the suppression of eosinophils in asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophils; Female; Inflammation; Mice; Mice, Inbred BALB C; Ovalbumin; Receptors, Purinergic P2Y12

We investigated the effect of long-term treatment with azithromycin on the pathogenesis of chronic asthma with airway remodeling.. Six-week-old-BALB/c mice were sensitized with ovalbumin (OVA) combined with lipopolysaccharide (LPS) for 1 month, then challenged with OVA for 3 months. Azithromycin at 75 mg/kg was administered via oral gavage five times a week during the challenge period. Inflammatory cells, T helper 2 cytokines in bronchoalveolar lavage fluid (BAL) fluid, and airway hyperresponsiveness (AHR) were measured. Parameters related to airway remodeling were evaluated. The levels of neutrophil elastase, Interleukin (IL)-8, and BRP-39 (human homologue YKL-40) were assessed. The expression of MAPK and NF-κB signaling were investigated.. Long-term treatment with azithromycin improved AHR and airway inflammation compared with the OVA and the OVA/LPS groups. The concentrations of IL-5 and IL-13 in the OVA/LPS group decreased significantly after azithromycin administration. The levels of neutrophil elastase and IL-8, as surrogate markers of neutrophil activation, were reduced in the azithromycin group compared with the OVA/LPS group. Goblet cell hyperplasia and the smooth muscle thickening of airway remodeling were attenuated after azithromycin treatment. The expression of MAPK/NF-kappaB signal and the level of BRP-39 in the lung decreased remarkably in the OVA/LPS with azithromycin-treated group.. This study suggests that in a murine model of chronic asthma, long-term azithromycin treatment ameliorates not only airway inflammation but also airway remodeling by influencing on neutrophilc-related mediators, BRP-39 and MAPK/NF-κB signal pathways. Macrolide therapy might be an effective adjuvant therapy in a chronic, severe asthma with remodeling airway.

Topics: Animals; Anti-Bacterial Agents; Asthma; Azithromycin; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Female; Interleukins; Leukocyte Elastase; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; Ovalbumin; Pneumonia; T-Lymphocytes, Helper-Inducer

Asthma is a chronic airway inflammatory disorder and progresses mainly due to airway remodeling. Chrysin, a natural flavonoid, has been reported to possess multiple biologic activities, including anti-inflammation, anti-oxidation and anti-proliferation. The present study aimed to investigate whether chrysin could relieve allergic airway inflammation and remodeling in a murine model of chronic asthma and the mechanism involved. The female BALB/c mice sensitized and challenged with ovalbumin (OVA) successfully developed airway hyperresponsiveness (AHR), inflammation and remodeling. The experimental data showed that chrysin could alleviate OVA-induced AHR. Chrysin could also reduce OVA-induced increases in the number of inflammatory cells, especially eosinophils, interleukin (IL) -4, and IL-13 in bronchoalveolar lavage fluid (BALF) and total IgE in serum. The decreased interferon-γ (IFN-γ) level in BALF was also upregulated by chrysin. In addition, inflammatory cell infiltration, goblet cell hyperplasia and the expression of α-smooth muscle actin (α-SMA) around bronchioles were suppressed by chrysin. Furthermore, the phosphorylation levels of Akt and extracellular signal-regulated kinase (ERK) could be decreased by chrysin, which are associated with airway smooth muscle cell (ASMC) proliferation. These results indicate the promising therapeutic effect of chrysin on chronic asthma, especially the progression of airway remodeling.

Topics: Acetylcholine; Airway Remodeling; Allergens; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Extracellular Signal-Regulated MAP Kinases; Female; Flavonoids; Immunoglobulin E; Lung; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Proto-Oncogene Proteins c-akt

Severe bronchospasm refractory to β-agonists is a challenging aspect of asthma therapy, and novel therapeutics are needed. β-agonist-induced airway smooth muscle (ASM) relaxation is associated with increases in the phosphorylation of the small heat shock-related protein (HSP) 20. We hypothesized that a transducible phosphopeptide mimetic of HSP20 (P20 peptide) causes relaxation of human ASM (HASM) by interacting with target(s) downstream of the β2-adrenergic receptor (β2AR) pathway. The effect of the P20 peptide on ASM contractility was determined in human and porcine ASM using a muscle bath. The effect of the P20 peptide on filamentous actin dynamics and migration was examined in intact porcine ASM and cultured primary HASM cells. The efficacy of the P20 peptide in vivo on airway hyperresponsiveness (AHR) was determined in an ovalbumin (OVA) sensitization and challenge murine model of allergic airway inflammation. P20 peptide caused dose-dependent relaxation of carbachol-precontracted ASM and blocked carbachol-induced contraction. The β2AR inhibitor, (±)-1-[2,3-(dihydro-7-methyl-1H-inden-4-yl)oxy]-3-[(1-methylethyl)amino]-2-butanol hydrochloride (ICI 118,551), abrogated isoproterenol but not P20 peptide-mediated relaxation. The P20 peptide decreased filamentous actin levels in intact ASM, disrupted stress fibers, and inhibited platelet-derived growth factor-induced migration of HASM cells. The P20 peptide treatment reduced methacholine-induced AHR in OVA mice without affecting the inflammatory response. These results suggest that the P20 peptide decreased airway constriction and disrupted stress fibers through regulation of the actin cytoskeleton downstream of β2AR. Thus, the P20 peptide may be a potential therapeutic for asthma refractory to β-agonists.

Topics: Actins; Adrenergic beta-2 Receptor Antagonists; Allergens; Animals; Bronchial Hyperreactivity; Carbachol; Cell Movement; Constriction; HSP20 Heat-Shock Proteins; Humans; Inflammation; Lung; Mice; Muscle Contraction; Muscle Relaxation; Muscle, Smooth; Myocytes, Smooth Muscle; Ovalbumin; Peptides; Receptors, Adrenergic, beta-2; Stress Fibers; Sus scrofa

Asthma is a highly prevalent chronic inflammatory lung disease characterized by airway hyperresponsiveness to allergens, airway edema, and increased mucus secretion. Such mucus can be liquefied by recombinant human deoxyribonuclease (rhDNase), in which efficacy of rhDNase has been well documented in patients with cystic fibrosis, but little studied in asthma. In the present study, we investigated whether rhDNase intranasal administration improved inflammation and pulmonary function in an experimental model of asthma.. Mice were sensitized by two subcutaneous injections of ovalbumin (OVA), on days 0 and 7, followed by three intranasal challenges with OVA on days 14, 15, and 16. A control group, replacing OVA by DPBS, was included. On days 15 and 16, after 2 hours of OVA challenge, mice received 1 mg/mL of intranasal rhDNase.. We showed that rhDNase decreased significantly the airway resistance and reduced EETs formation and globet cells hyperplasia.. Our results suggest that extracellular DNA in mucus play a role in lower airways obstruction in OVA asthma protocol and that the treatment with rhDNase improved lung function and DNA extracellular traps, with no direct cellular anti-inflammatory effects.

Topics: Administration, Intranasal; Airway Obstruction; Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Deoxyribonucleases; Disease Models, Animal; DNA; Extracellular Traps; Female; Humans; Lung; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Recombinant Proteins

In allergen-induced asthma, activated mast cells start the lung inflammatory process with degranulation, cytokine synthesis, and mediator release. Bruton's tyrosine kinase (Btk) activity is required for the mast cell activation during IgE-mediated secretion.. This study characterized a novel inhaled Btk inhibitor RN983 in vitro and in ovalbumin allergic mouse models of the early (EAR) and late (LAR) asthmatic response.. RN983 potently, selectively, and reversibly inhibited the Btk enzyme. RN983 displayed functional activities in human cell-based assays in multiple cell types, inhibiting IgG production in B-cells with an IC50 of 2.5 ± 0.7 nM and PGD2 production from mast cells with an IC50 of 8.3 ± 1.1 nM. RN983 displayed similar functional activities in the allergic mouse model of asthma when delivered as a dry powder aerosol by nose-only inhalation. RN983 was less potent at inhibiting bronchoconstriction (IC50(RN983) = 59 μg/kg) than the β-agonist salbutamol (IC50(salbutamol) = 15 μg/kg) in the mouse model of the EAR. RN983 was more potent at inhibiting the antigen induced increase in pulmonary inflammation (IC50(RN983) = <3 μg/kg) than the inhaled corticosteroid budesonide (IC50(budesonide) = 27 μg/kg) in the mouse model of the LAR.. Inhalation of aerosolized RN983 may be effective as a stand-alone asthma therapy or used in combination with inhaled steroids and β-agonists in severe asthmatics due to its potent inhibition of mast cell activation.

Topics: Administration, Inhalation; Adrenergic beta-2 Receptor Agonists; Agammaglobulinaemia Tyrosine Kinase; Albuterol; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Bronchoconstriction; Bronchodilator Agents; Budesonide; Cell Degranulation; Cells, Cultured; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Dry Powder Inhalers; Glucocorticoids; Humans; Immunoglobulin G; Lung; Male; Mast Cells; Mice, Inbred BALB C; Ovalbumin; Phthalazines; Pneumonia; Prostaglandin D2; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Pyridazines

Phosphodiesterases (PDEs) are enzymes involved in the degradation of cAMP and cGMP. Selective PDE4 inhibitors (e.g., roflumilast) are effective in therapy of chronic obstructive pulmonary disease associated with neutrophil inflammation. The aim of this study was to evaluate the effects of a selective PDE4 inhibitor, YM976, on citric acid-induced cough, in vivo and in vitro airway smooth muscle reactivity to histamine, and on inflammatory mediators in ovalbumin-sensitized guinea pigs, with experimentally induced eosinophil inflammation. The YM976 was administered intraperitoneally at a dose of 1.0 mg/kg once daily for 7 days. Sensitization with ovalbumin led to a significant increase in the number of coughs, and in vivo and in vitro airway reactivity. Also, increased plasma levels of IL-4, IL-5, and PAF were observed, with a significant increase in the differential count of eosinophils in both blood and bronchoalveolar lavage fluid. The YM976 suppressed the number of coughs, the airway reactivity in tracheal tissue strips, and the IL-4 level. The findings indicate that PDE4 inhibition by YM976 exerts antitussive and anti-inflammatory effects in guinea pigs with ovalbumin-induced eosinophilic inflammation.

Topics: Animals; Bronchial Hyperreactivity; Cough; Cyclic Nucleotide Phosphodiesterases, Type 4; Guinea Pigs; Male; Ovalbumin; Phosphodiesterase 4 Inhibitors; Pyridines; Pyrimidinones; Respiratory System

Guang-Pheretima, the live form of the earthworm Pheretima aspergillum, is a traditional Chinese medicine commonly used for the treatment of asthma, cough, stroke, epilepsy and other diseases due to its anti-inflammatory, anti-asthmatic, anti-seizure, thrombolytic and diuretic properties. Although Guang-Pheretima is effective in the relief of asthma, its pharmacological activity and the underlying molecular mechanisms are not fully understood. Hence, we investigated the effects of a Pheretima aspergillum decoction (PAD) against inflammation in a model of ovalbumin (OVA)-induced asthma in BALB/c mice, as well as the nuclear factor-κB (NF-κB) pathway involved in this process.. OVA was used to sensitize and challenge the airway of the mice, and PAD was administrated by gavage. We measured airway hyperresponsiveness (AHR) in the mice 24h following a final methacholine challenge with whole-body plethysmography. The bronchoalveolar lavage fluid (BALF), serum and pulmonary tissues were collected 48h after the last challenge. The levels of inflammatory factors and the related mRNAs were determined by enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (RT-PCR), respectively. The number of differential inflammatory cells in the BALF was counted. Serum total and OVA-specific IgE levels were measured with ELISA. The activation of NF-κB signaling in the lung was detected by western blotting. In addition, the lung tissues were stained with hematoxylin and eosin or periodic acid Schiff stain for histopathological examination.. PAD treatment significantly alleviated AHR in the asthmatic mice, decreased the mRNA and protein levels of IL-4, IL-5 and IL-13 and downregulated IgE. In addition, PAD treatment attenuated mucus secretion and infiltration of inflammatory cells in the lung while inhibiting the activation of NF-κB signaling.. PAD effectively inhibited the activation of NF-κB signaling in the lungs of mice with OVA-induced asthma, and mitigated AHR and Th2 type inflammatory reactions. Therefore, PAD may serve as a drug candidate for asthma treatment.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cytokines; Disease Models, Animal; Down-Regulation; Female; Immunoglobulin E; Inflammation Mediators; Mice, Inbred BALB C; NF-kappa B; Oligochaeta; Ovalbumin; RNA, Messenger; Signal Transduction; Th2 Cells; Time Factors; Tissue Extracts

Imperatorin is a furanocoumarin compound which exists in many medicinal herbs and possesses various biological activities. Herein, we investigated the antiallergic effects of imperatorin in asthmatic mice and explored the immunomodulatory actions of imperatorin on immune cells. We used a murine model of ovalbumin (OVA)-induced asthma to evaluate the therapeutic potential of imperatorin. Additionally, bone marrow-derived dendritic cells (DCs; BMDCs) were used to clarify whether imperatorin exerts an antiallergic effect through altering the ability of DCs to regulate T cells. Oral administration of imperatorin to OVA-sensitized and -challenged mice decreased serum OVA-specific immunoglobulin E (IgE) production, attenuated the airway hyperresponsiveness (AHR), and alleviated airway inflammation in a dose-dependent manner. Notably, secretions of Th2 cytokines and chemokines were reduced, and numbers of interleukin (IL)-10-producing regulatory T cells (Tregs) increased in imperatorin-treated mice. Imperatorin inhibited proinflammatory cytokines and IL-12 production but enhanced IL-10 secretion by lipopolysaccharide (LPS)-stimulated BMDCs. Compared to fully mature DCs, imperatorin-treated DCs expressed high levels of the inducible costimulatory ligand (ICOSL) and Jagged1 molecules, and had the regulatory capacity to promote the generation of IL-10-producing CD4(+) T cells in vitro. Additionally, imperatorin directly suppressed activated CD4(+) T-cell proliferation and cytokine production. Imperatorin may possess therapeutic potential against Th2-mediated allergic asthma not only via stimulating DC induction of Tregs but also via direct inhibition of Th2 cell activation. These findings provide new insights into how imperatorin affects the Th2 immune response and the development of imperatorin as a Treg-type immunomodulatory agent to treat allergic asthma.

Topics: Animals; Anti-Allergic Agents; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Cell Proliferation; Cells, Cultured; Dendritic Cells; Disease Models, Animal; Female; Furocoumarins; Immunoglobulin E; Inducible T-Cell Co-Stimulator Ligand; Interleukin-10; Jagged-1 Protein; Lung; Lymphocyte Activation; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Phenotype; Signal Transduction; T-Lymphocytes, Regulatory; Th2 Cells; Time Factors

Silica nanoparticles (SNPs) can easily enter in respiratory system via inhalation because of their low molecular weight and ease of dispersion. Toxicity and adverse effects of SNPs vary according to the physical characteristics of the particle.. To evaluate the toxic and adjuvant effects of 3 types of SNPs in the airway system, six-week-old female BALB/c mice were intranasally administered 3 types of SNPs (spherical [S-SNP], mesoporous [M-SNP], and polyethylene glycol-conjugated [P-SNP]) alone or SNPs/ovalbumin (OVA), three times weekly for 2 weeks. Airway hyper-responsiveness (AHR), bronchoalveolar lavage fluid (BALF), cytokine levels, and histology of the lungs were analyzed.. The S-SNPs/OVA group and M-SNPs/OVA group showed significant AHR, compared to the control group. Among all SNP-treated groups, the group administered SNPs/OVA showed greater inflammatory cell infiltration in BALF, extensive pathological changes, and higher cytokine levels (IL-5, IL-13, IL-1β, and IFN-γ) than those administered SNPs alone or saline/OVA.. Exposure to SNPs alone and SNPs/OVA induced toxicity in the respiratory system. SNPs alone showed significant toxic effects on the airway system. Meanwhile, SNPs/OVA exerted adjuvant effects to OVA of inducing allergic airway inflammation. In particular, M-SNPs showed the most severe airway inflammation in both direct toxicity and adjuvant effect assays. P-SNPs induced less inflammation than the other types of SNPs in both models.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cytokines; Disease Models, Animal; Female; Inflammation Mediators; Inhalation Exposure; Lung; Mice, Inbred BALB C; Nanoparticles; Ovalbumin; Pneumonia; Polyethylene Glycols; Porosity; Silicon Dioxide

Phosphodiesterases (PDEs) represent a super-family of 11 enzymes hydrolyzing cyclic nucleotides into inactive 5' monophosphates. Inhibition of PDEs leads to a variety of cellular effects, including airway smooth muscle relaxation, inhibition of cellular inflammation, and immune responses. In this study we focused on theophylline, a known non-selective inhibitor of PDEs. Theophylline has been used for decades in the treatment of chronic inflammatory airway diseases. It has a narrow therapeutic window and belongs to the drugs whose plasma concentration should be monitored. Therefore, the main goal of this study was to evaluate the plasma theophylline concentration and to determine its relevance to pharmacological effects after single and longer term (7 days) administration of theophylline at different doses (5, 10, 20, and 50 mg/kg) in guinea pigs. Airway hyperresponsiveness was assessed by repeated exposure to ovalbumin. Theophylline reduced specific airway resistance in response to histamine nebulization, measured in a double chamber body plethysmograph. A decrease in tracheal smooth muscle contractility after cumulative doses of histamine and acetylcholine was confirmed in vitro. A greater efficacy of theophylline after seven days long treatment indicates the predominance of its anti-inflammatory activity, which may be involved in the bronchodilating action.

Topics: Airway Resistance; Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchoconstriction; Bronchodilator Agents; Guinea Pigs; Inflammation; Male; Muscle Contraction; Ovalbumin; Plethysmography, Whole Body; Theophylline

The catalytical isoforms p110γ and p110δ of phosphatidylinositide 3-kinase γ (PI3Kγ) and PI3Kδ play an important role in the pathogenesis of asthma. Two key elements in allergic asthma are increased levels of eosinophils and IgE. Dual pharmacological inhibition of p110γ and p110δ reduces asthma-associated eosinophilic lung infiltration and ameliorates disease symptoms, whereas the absence of enzymatic activity in p110γKOδD910A mice increases IgE and basal eosinophil counts. This suggests that long-term inhibition of p110γ and p110δ might exacerbate asthma. Here, we analysed mice genetically deficient for both catalytical subunits (p110γ/δ-/-) and determined basal IgE and eosinophil levels and the immune response to ovalbumin-induced asthma. Serum concentrations of IgE, IL-5 and eosinophil numbers were significantly increased in p110γ/δ-/- mice compared to single knock-out and wildtype mice. However, p110γ/δ-/- mice were protected against OVA-induced infiltration of eosinophils, neutrophils, T and B cells into lung tissue and bronchoalveolar space. Moreover, p110γ/δ-/- mice, but not single knock-out mice, showed a reduced bronchial hyperresponsiveness. We conclude that increased levels of eosinophils and IgE in p110γ/δ-/- mice do not abolish the protective effect of p110γ/δ-deficiency against OVA-induced allergic airway inflammation.

Topics: Animals; B-Lymphocytes; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Class I Phosphatidylinositol 3-Kinases; Eosinophilia; Eosinophils; Goblet Cells; Hypersensitivity; Immunoglobulin E; Interleukin-5; Lung; Metaplasia; Mice, Inbred C57BL; Neutrophils; Ovalbumin; Pneumonia; T-Lymphocytes

Asthma is a heterogeneous disease characterized by chronic airway inflammation. CD4(+) T-helper 9 (Th9) cells are closely linked to asthma, helping to regulate inflammation and immunity. Epidemiological studies showed that mycobacteria infections are negatively associated with asthma. Our previous research showed that inactivated Mycobacterium phlei nebulization alleviated the airway hyperresponsiveness and inflammation of asthma. However, the relationship between Th9 cells and mycobacteria remains unknown. Here, we evaluated the relationship between Mycobacterium vaccae nebulization and Th9 cells in asthmatic mice. Eighteen Balb/c mice were randomized into 3 groups of 6 mice each (normal control group, asthma control group, and nebulization asthma group [Neb. group]). The Neb. group was nebulized with M. vaccae one month before establishment of the asthmatic model with ovalbumin (OVA) sensitization, and the normal and asthma control groups were nebulized with phosphate-buffered saline. The hyperresponsiveness of the mouse airways was assessed using a non-invasive lung function machine. Lung airway inflammation was evaluated by hematoxylin and eosin and periodic acid-Schiff staining. Cytokine interlukin-9 (IL-9) concentration and OVA-specific IgE in the bronchoalveolar lavage fluid were measured by enzyme-linked immunosorbent assays. The percentages of γδTCR+ CD3+, IL-9+CD3+, IL-10+CD3+ lymphocytes, and IL9+γδT and IL-10+γδT cells were detected by flow cytometry. The airway inflammation and concentration of IL-9 and OVA-specific IgE were significantly reduced in the Neb. group compared to the asthma control group. The Neb. group had lower airway hyperresponsiveness, percentages of γδTCR+CD3+ and IL-9+CD3+ lymphocytes, and IL9+γδT cells, and higher percentages of IL-10+CD3+ lymphocytes and IL-10+γδT cells compared to the asthma control group. Thus, mouse bronchial asthma could be prevented by M. vaccae nebulization. The mechanism could involve M. vaccae-mediated effects on induction of IL-9 secretion and suppression of IL-10 secretion from γδT cells. γδT cells showed prominent IL-10 expression, indicating that they possibly belong to the Th9 family.

Topics: Administration, Inhalation; Animals; Asthma; Bacterial Vaccines; Bronchial Hyperreactivity; Disease Models, Animal; Female; Inflammation; Interleukin-9; Mice; Mice, Inbred BALB C; Mycobacterium; Nebulizers and Vaporizers; Ovalbumin; T-Lymphocytes, Helper-Inducer

Allergic airway inflammation is one of the primary features of allergic asthma. Interleukin-33 (IL-33) is recognized as a key pro-inflammatory cytokine that mediates allergic airway inflammation, and its expression is elevated in this condition, but little is known about the regulatory mechanisms underlying IL-33 induction. Here, we show that the RNA binding protein Mex-3B plays a critical role in the induction of IL-33 in the development of allergic airway inflammation. We generated Mex3b(-/-) mice and found that they develop significantly less airway inflammation than wild-type mice due to reduced induction of IL-33. Furthermore, we show that Mex-3B directly upregulates IL-33 expression by inhibiting miR-487b-3p-mediated repression of IL-33. Moreover, we show that inhalation of an antisense oligonucleotide targeting Mex-3B suppresses allergic airway inflammation. Our data identify a signaling pathway that post-transcriptionally regulates IL-33 expression and suggest that Mex-3B could be a promising molecular target for the treatment of allergic asthma.

Topics: Animals; Base Sequence; Bronchial Hyperreactivity; Epithelial Cells; Gene Expression Regulation; Humans; Interleukin-33; Killer Cells, Natural; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Knockout; MicroRNAs; Nucleic Acid Conformation; Oligonucleotides, Antisense; Ovalbumin; Pulmonary Alveoli; Respiratory Mucosa; RNA-Binding Proteins; Signal Transduction; Th2 Cells

Angelicin, a furocoumarin found in Psoralea corylifolia L. fruit, has been reported to have anti-inflammatory activity. The purpose of this study was to determine the protective effects of angelicin on allergic asthma induced by ovalbumin (OVA) in mice. Mice were sensitized to OVA (on days 0 and 14) and challenged with OVA three times (on days 21 to 23). Angelicin (2.5, 5, 10 mg/kg) was given intraperitoneally 1 h before OVA treatment after the initial OVA sensitization. The production of IL-4, IL-5, and IL-13 in BALF and IgE in the serum were measured by ELISA. Lung histological changes were detected by using hematoxylin and eosin (H&E) stain. The results showed that angelicin significantly inhibited inflammatory cells infiltration into the lungs. Histological studies showed that angelicin significantly attenuated OVA-induced lung injury. Meanwhile, treatment of angelicin dose-dependently inhibited OVA-induced the production of IL-4, IL-5, and IL-13 in BALF and IgE in the serum. Furthermore, angelicin was found to inhibit airway hyperresponsiveness and NF-kB activation. In conclusion, our results suggested that angelicin inhibited allergic airway inflammation and hyperresponsiveness by inhibiting NF-kB activation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Dose-Response Relationship, Drug; Furocoumarins; Immunoglobulin E; Inflammation; Lung; Mice; NF-kappa B; Ovalbumin

Compelling evidences point out a crucial role for extracellular nucleotides such as adenosine triphosphate (ATP) during inflammatory conditions. Once released into the extracellular space, ATP modulates migration, maturation and function of various inflammatory cells via activating of purinergic receptors of the P2Y- and P2X- family. P2RX4 is an ATP-guided ion channel expressed on structural cells such as alveolar epithelial and smooth muscle cells as well as inflammatory cells including macrophages, dendritic cells (DCs) and T cells. P2RX4 has been shown to interact with P2RX7 and promote NLRP3 inflammasome activation. Although P2RX7 has already been implicated in allergic asthma, the role of P2RX4 in airway inflammation has not been elucidated yet. Therefore, we used a selective pharmacological antagonist and genetic ablation to investigate the role of P2RX4 in an ovalbumin (OVA) driven model of allergen-induced airway inflammation (AAI). Both, P2RX4 antagonist 5-BDBD treatment and P2rx4 deficiency resulted in an alleviated broncho alveolar lavage fluid eosinophilia, peribronchial inflammation, Th2 cytokine production and bronchial hyperresponsiveness. Furthermore, P2rx4-deficient bone marrow derived DCs (BMDCs) showed a reduced IL-1ß production in response to ATP accompanied by a decreased P2rx7 expression and attenuated Th2 priming capacity compared to wild type (WT) BMDCs in vitro. Moreover, mice adoptively transferred with P2rx4-deficient BMDCs exhibit a diminished AAI in vivo. In conclusion our data suggests that P2RX4-signaling contributes to AAI pathogenesis by regulating DC mediated Th2 cell priming via modulating IL-1ß secretion and selective P2RX4-antagonists might be a new therapeutic option for allergic asthma.

Topics: Adenosine Triphosphate; Adoptive Transfer; Allergens; Animals; Benzodiazepinones; Bone Marrow Cells; Bone Marrow Transplantation; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cells, Cultured; Coculture Techniques; Cytokines; Dendritic Cells; Disease Models, Animal; Genetic Predisposition to Disease; Humans; Lung; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Phenotype; Pneumonia; Purinergic P2X Receptor Agonists; Purinergic P2X Receptor Antagonists; Pyroglyphidae; Receptors, Purinergic P2X4; Th2 Cells

Glibenclamide has a newly discovered role in inflammation regulation besides its antidiabetic effect. As an inhibitor of ATP-sensitive potassium (KATP) channel, glibenclamide antagonizes the relaxation of the tracheal smooth muscle. This indicates that glibenclamide might attenuate airway inflammation while aggravate airway hyperresponsiveness (AHR) in asthmatics. Clinically, many diabetics with asthma are prescribed with glibenclamide to control blood glucose. However, whether glibenclamide could exert any effects on asthmatic inflammation remains unknown. Using an ovalbumin (OVA)-induced mouse model of asthma, we evaluated the effects of glibenclamide on the AHR and inflammation. Interestingly, glibenclamide reduced all the cardinal features of asthma in OVA-challenged mice, including AHR, airway inflammation, and T-helper type 2 (Th2) cytokines. Glibenclamide also downregulated OVA-induced expressions of vascular cell adhesion molecule 1 (VCAM-1) and phosphorylated signal transducer and activator of transcription 6 (p-STAT6) in the lung. In addition, increased sulfonylurea receptor 1 (SUR1) expression in the lung was observed after the OVA challenge. These findings suggest that the classic sulfonylurea glibenclamide plays an important protective role in the development of asthma, which not only provides the evidence for the safety of prescribed glibenclamide in diabetics combined with asthma but also indicates a possible new therapeutic for asthma via targeting glibenclamide-related pathways.

Topics: Animals; Bronchial Hyperreactivity; Cells, Cultured; Chickens; Cytokines; Glyburide; Hypoglycemic Agents; Inflammation; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Protective Agents

Sphingosine-1-phosphate (S1P) has been widely associated with inflammation-based lung pathologies. Because B cells play a critical role as antigen-presenting and/or Ig-producing cells during asthmatic conditions, we wanted to dissect the role of these cells in S1P-dependent airway hyperreactivity and inflammation. Mice were sensitized to ovalbumin or exposed to S1P. Ovalbumin sensitization caused airway hyperreactivity coupled to an increased lung infiltration of B cells, which was significantly reduced after the inhibition of sphingosine kinases I/II. Similarly, the sole administration of S1P increased bronchial reactivity compared with vehicle and was accompanied by a higher influx of B cells in a time-dependent manner. This effect was associated with higher levels of IL-13, transforming growth factor-β, IL-10, and T regulatory cells. In addition, isolated S1P-derived lung B cells increased CD4(+) and CD8(+) T cell proliferation in vitro, and their suppressive nature at Day 14 was associated with the higher release of transforming growth factor-β and IL-10 when they were cocultured. Therefore, to prove the role of B cells in S1P-mediated airway inflammation, and because CD20 expression, contrary to major hystocompatibility complex I and major hystocompatibility complex II, was up-regulated at Day 14, CD20(+) B cells were depleted by means of a specific monoclonal antibody. The absence of CD20(+) B cells increased airway reactivity and inflammation in S1P-treated mice compared with control mice. These data imply that sphingosine kinase/S1P-mediated airway inflammation is countered by B cells via the induction of an immune-suppressive environment to reduce asthma-like outcomes in mice.

Topics: Animals; Antibodies, Monoclonal; Antigens, CD20; B-Lymphocytes; Bronchial Hyperreactivity; Bronchoconstriction; Cell Proliferation; Chemotaxis, Leukocyte; Disease Models, Animal; Female; Inflammation Mediators; Interleukin-10; Interleukin-13; Lung; Lymphocyte Activation; Lysophospholipids; Mice, Inbred BALB C; Ovalbumin; Phosphotransferases (Alcohol Group Acceptor); Pneumonia; Protein Kinase Inhibitors; Sphingosine; T-Lymphocytes, Regulatory; Time Factors; Transforming Growth Factor beta

Nitric oxide (NO) effects in airways are influenced by the activity of NO-synthase isoforms and NO metabolism. Inducible NO-synthase (iNOS), which produces large amounts of NO, is active during the inflammatory process. NO quickly reacts, producing reactive oxygen species (ROS). In this study we attempted to detect the expression of iNOS and markers of ROS in the airway hyperreactivity (AHR) condition. The study was performed in guinea pigs, divided into four groups. Two groups were treated with the non-selective inhibitor of NO-synthase L-NAME. The other two groups were used as controls. Exhaled NO was monitored in vivo, AHR was assessed both in vivo and in vitro, and the expression of iNOS in lung homogenate, and oxidative stress markers were measured in the venous blood. L-NAME significantly affected the AHR only in in vitro condition, blocked the expression of iNOS in control but not in sensitized animals, and decreased the level of exhaled NO. The results concerning the oxidative stress markers are equivocal. The study confirmed that NO is involved in the regulation of AHR; the effects being mediated via iNOS and ROS activity.

Topics: Animals; Bronchial Hyperreactivity; Enzyme Inhibitors; Exhalation; Gene Expression; Guinea Pigs; Lung; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase Type II; Ovalbumin; Oxidative Stress; Reactive Oxygen Species

Chronic inflammatory diseases, associated with airway obstruction and cough, are usually treated with bronchodilating and anti-inflammatory drugs. Inhibition of phosphodiesterases (PDE) leads to both of these effects and influences apoptosis of immune cells. In chronic obstructive pulmonary disease, roflumilast, a selective PDE4 inhibitor, has been recently approved for pharmacotherapy. The aim of this study was to evaluate the effect of long-term administration of roflumilast in experimental allergic inflammation in guinea pigs. Male adult guinea pigs were used in the study. There were four experimental groups sensitized with ovalbumin for 14 days and thereafter treated per os, by inhalation, and intraperitoneally for 7 days with roflumilast or vehicle. A control group was left without sensitization. Roflumilast reduced specific airway resistance after nebulization of histamine, as measured in a double-chamber whole-body plethysmograph. This effect was confirmed in in vitro organ bath, with significant decreases in tracheal and lung smooth muscle contractility after cumulative doses of histamine. Suppression of hematological and immunological markers of inflammation and enhanced apoptosis in animals treated with roflumilast points to the possibility of a beneficial effect of roflumilast in allergic inflammation.

Topics: Administration, Inhalation; Administration, Oral; Airway Resistance; Aminopyridines; Animals; Apoptosis; Benzamides; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cyclopropanes; Guinea Pigs; Histamine; Leukocytes, Mononuclear; Lung; Male; Muscle Contraction; Muscle, Smooth; Ovalbumin; Phosphodiesterase 4 Inhibitors; Plethysmography, Whole Body; Primary Cell Culture; Tissue Culture Techniques

High-conductive calcium-sensitive potassium channels (BK+Ca) and ATP-sensitive potassium (K+ATP) channels play a significant role in the airway smooth muscle cell and goblet cell function, and cytokine production. The present study evaluated the therapeutic potential of BK+Ca and K+ATP openers, NS 1619 and pinacidil, respectively, in an experimental model of allergic inflammation. Airway allergic inflammation was induced with ovalbumine in guinea pigs during 21 days, which was followed by a 14-day treatment with BK+Ca and K+ATP openers. The outcome measures were airway smooth muscle cells reactivity in vivo and in vitro, cilia beating frequency and the level of exhaled NO (ENO), and the level of pro-inflammatory cytokines in the plasma and bronchoalveolar lavage fluid. The openers of both channels decreased airway smooth muscle cells reactivity, cilia beating frequency, and cytokine levels in the serum. Furthermore, NS1619 reduced ENO and inflammatory cells infiltration. The findings confirmed the presence of beneficial effects of BK+Ca and K+ATP openers on airway defence mechanisms. Although both openers dampened pro-inflammatory cytokines and mast cells infiltration, an evident anti-inflammatory effect was provided only by NS1619. Therefore, we conclude that particularly BK+Ca channels represent a promising new drug target in treatment of airway's allergic inflammation.

Topics: Administration, Inhalation; Airway Resistance; Animals; Benzimidazoles; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cilia; Cytokines; Exhalation; Guinea Pigs; KATP Channels; Large-Conductance Calcium-Activated Potassium Channels; Lung; Male; Membrane Transport Modulators; Muscle, Smooth; Nitric Oxide; Ovalbumin; Pinacidil; Plethysmography, Whole Body; Tissue Culture Techniques

Asthma is an inflammatory disease of the lung characterized by airways hyper-responsiveness (AHR), inflammation, and mucus hyperproduction. Current mainstream therapies include bronchodilators that relieve bronchoconstriction and inhaled glucocorticoids to reduce inflammation. The small molecule hormone and neurotransmitter serotonin has long been known to be involved in inflammatory processes; however, its precise role in asthma is unknown. We have previously established that activation of serotonin 5-hydroxytryptamine (5-HT)(2A) receptors has potent anti-inflammatory activity in primary cultures of vascular tissues and in the whole animal in vasculature and gut tissues. The 5-HT(2A) receptor agonist, (R)-2,5-dimethoxy-4-iodoamphetamine [(R)-DOI] is especially potent. In this work, we have examined the effect of (R)-DOI in an established mouse model of allergic asthma. In the ovalbumin mouse model of allergic inflammation, we demonstrate that inhalation of (R)-DOI prevents the development of many key features of allergic asthma, including AHR, mucus hyperproduction, airways inflammation, and pulmonary eosinophil recruitment. Our results highlight a likely role of the 5-HT2 receptors in allergic airways disease and suggest that 5-HT2 receptor agonists may represent an effective and novel small molecule-based therapy for asthma.

Topics: 5-Hydroxytryptophan; Amphetamines; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Enzyme Activation; Eosinophils; Immunoglobulin E; Inflammation; Lung; Male; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Pulmonary Eosinophilia; Receptors, Serotonin, 5-HT2; Serotonin 5-HT2 Receptor Agonists

We reported that DNA-dependent protein kinase (DNA-PK) is critical for the expression of nuclear factor κB-dependent genes in TNF-α-treated glioblastoma cells, suggesting an involvement in inflammatory diseases.. We sought to investigate the role of DNA-PK in asthma.. Cell culture and ovalbumin (OVA)- or house dust mite-based murine asthma models were used in this study.. DNA-PK was essential for monocyte adhesion to TNF-α-treated endothelial cells. Administration of the DNA-PK inhibitor NU7441 reduced airway eosinophilia, mucus hypersecretion, airway hyperresponsiveness, and OVA-specific IgE production in mice prechallenged with OVA. Such effects correlated with a marked reduction in lung vascular cell adhesion molecule 1 expression and production of several cytokines, including IL-4, IL-5, IL-13, eotaxin, IL-2, and IL-12 and the chemokines monocyte chemoattractant protein 1 and keratinocyte-derived chemokine, with a negligible effect on IL-10/IFN-γ production. DNA-PK inhibition by gene heterozygosity of the 450-kDa catalytic subunit of the kinase (DNA-PKcs(+/-)) also prevented manifestation of asthma-like traits. These results were confirmed in a chronic model of asthma by using house dust mite, a human allergen. Remarkably, such protection occurred without causing severe combined immunodeficiency. Adoptive transfer of TH2-skewed OT-II wild-type CD4(+) T cells reversed IgE and TH2 cytokine production but not airway hyperresponsiveness in OVA-challenged DNA-PKcs(+/-) mice. DNA-PK inhibition reduced IL-4, IL-5, IL-13, eotaxin, IL-8, and monocyte chemoattractant protein 1 production without affecting IL-2, IL-12, IFN-γ, and interferon-inducible protein 10 production in CD3/CD28-stimulated human CD4(+) T cells, potentially by blocking expression of Gata3. These effects occurred without significant reductions in T-cell proliferation. In mouse CD4(+) T cells in vitro DNA-PK inhibition severely blocked CD3/CD28-induced Gata3 and T-bet expression in CD4(+) T cells and prevented differentiation of TH1 and TH2 cells under respective TH1- and TH2-skewing conditions.. Our results suggest DNA-PK as a novel determinant of asthma and a potential target for the treatment of the disease.

Topics: Adoptive Transfer; Allergens; Animals; Asthma; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Adhesion; Cytokines; Disease Models, Animal; DNA-Activated Protein Kinase; Eosinophils; Epithelial Cells; GATA3 Transcription Factor; Gene Expression; Genetic Heterogeneity; Humans; Immunoglobulin E; Lymphocyte Activation; Male; Mice; Mice, Knockout; Organ Size; Ovalbumin; Phenotype; Plasma Cells; Pyroglyphidae; Receptors, Antigen, T-Cell; Respiratory Mucosa; Severe Combined Immunodeficiency; Spleen; T-Lymphocyte Subsets; Th2 Cells; Tumor Necrosis Factor-alpha; Vascular Cell Adhesion Molecule-1

A long-term imbalance between pro- and anti-inflammatory mediators leads to airway remodelling, which is strongly correlated to most of the symptoms, severity and progression of chronic lung inflammation. The Angiotensin-(1-7) [Ang-(1-7)]/Mas receptor axis of the renin-angiotensin system is associated with attenuation of acute and chronic inflammatory processes. In this study, we investigated the effects of Ang-(1-7) treatment in a model of chronic allergic lung inflammation.. Mice were sensitized to ovalbumin (OVA; 4 injections over 42 days, 14 days apart) and were challenged three times per week (days 21-46). These mice received Ang-(1-7) (1 μg·h(-1) , s.c.) by osmotic mini-pumps, for the last 28 days. Histology and morphometric analysis were performed in left lung and right ventricle. Airway responsiveness to methacholine, analysis of Ang-(1-7) levels (RIA), collagen I and III (qRT-PCR), ERK1/2 and JNK (Western blotting), IgE (elisa), cytokines and chemokines (elisa multiplex), and immunohistochemistry for Mas receptors were performed.. Infusion of Ang-(1-7) in OVA-sensitized and challenged mice decreased inflammatory cell infiltration and collagen deposition in the airways and lung parenchyma, and prevented bronchial hyperresponsiveness. These effects were accompanied by decreased IgE and ERK1/2 phosphorylation, and decreased pro-inflammatory cytokines. Mas receptors were detected in the epithelium and bronchial smooth muscle, suggesting a site in the lung for the beneficial actions of Ang-(1-7).. Ang-(1-7) exerted beneficial attenuation of three major features of chronic asthma: lung inflammation, airway remodelling and hyperresponsiveness. Our results support an important protective role of Ang-(1-7) in lung inflammation.

Topics: Airway Remodeling; Angiotensin I; Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchoconstriction; Collagen; Cytokines; Disease Models, Animal; Hypertrophy, Right Ventricular; Immunoglobulin E; Inflammation Mediators; Lung; Male; Mice, Inbred BALB C; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Ovalbumin; Peptide Fragments; Phosphorylation; Pneumonia; Proto-Oncogene Mas; Proto-Oncogene Proteins; Receptors, G-Protein-Coupled; Respiratory Hypersensitivity; Signal Transduction

Zerumbone is a sesquiterpene compound isolated from the rhizome of wild ginger, Zingiber zerumbet Smith. The rhizomes of the plant are used as a spice and traditional medicine. Zerumbone was shown to possess anticarcinogenic, anti-inflammatory, and antioxidant properties. However, the antiallergic activity and the underlying mechanism of zerumbone have not been reported. Herein, we investigated the immunomodulatory effects of zerumbone on antigen-presenting dendritic cells (DCs) in vitro and its potential therapeutic effects against ovalbumin (OVA)-induced T helper 2 (Th2)-mediated asthma in mice. In the presence of zerumbone, lipopolysaccharide-activated bone marrow-derived DCs enhanced T cell proliferation and Th1 cell polarization in an allogeneic mixed lymphocyte reaction. In animal experiments, mice were sensitized and challenged with OVA, and were orally treated with different doses of zerumbone after sensitization. Circulating titers of OVA-specific antibodies, airway hyperresponsiveness to methacholine, histological changes in lung tissues, the cell composition and cytokine levels in bronchoalveolar lavage fluid, and cytokine profiles of spleen cells were assessed. Compared to OVA-induced hallmarks of asthma, oral administration of zerumbone induced lower OVA-specific immunoglobulin E (IgE) and higher IgG2a antibody production, attenuated airway hyperresponsiveness, prevented eosinophilic pulmonary infiltration, and ameliorated mucus hypersecretion. Zerumbone treatment also reduced the production of eotaxin, keratinocyte-derived chemokine (KC), interleukin (IL)-4, IL-5, IL-10, and IL-13, and promoted Th1 cytokine interferon (IFN)-γ production in asthmatic mice. Taken together, these results suggest that zerumbone exhibits an antiallergic effect via modulation of Th1/Th2 cytokines in an asthmatic mouse model.

Topics: Animals; Anti-Allergic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Dendritic Cells; Female; Immunoglobulin E; Immunoglobulin G; Lung; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Sesquiterpenes; Spleen; Th1 Cells; Th2 Cells

CD38 is a cell-surface protein involved in calcium signaling and contractility of airway smooth muscle. It has a role in normal airway responsiveness and in airway hyperresponsiveness (AHR) developed following airway exposure to IL-13 and TNF-α but appears not to be critical to airway inflammation in response to the cytokines. CD38 is also involved in T cell-mediated immune response to protein antigens. In this study, we assessed the contribution of CD38 to AHR and inflammation to two distinct allergens, ovalbumin and the epidemiologically relevant environmental fungus Alternaria. We also generated bone marrow chimeras to assess whether Cd38(+/+) inflammatory cells would restore AHR in the CD38-deficient (Cd38(-/-)) hosts following ovalbumin challenge. Results show that wild-type (WT) mice develop greater AHR to inhaled methacholine than Cd38(-/-) mice following challenge with either allergen, with comparable airway inflammation. Reciprocal bone marrow transfers did not change the native airway phenotypic differences between WT and Cd38(-/-) mice, indicating that the lower airway reactivity of Cd38(-/-) mice stems from Cd38(-/-) lung parenchymal cells. Following bone marrow transfer from either source and ovalbumin challenge, the phenotype of Cd38(-/-) hosts was partially reversed, whereas the airway phenotype of the WT hosts was preserved. Airway inflammation was similar in Cd38(-/-) and WT chimeras. These results indicate that loss of CD38 on hematopoietic cells is not sufficient to prevent AHR and that the magnitude of airway inflammation is not the predominant underlying determinant of AHR in mice.

Topics: Administration, Inhalation; ADP-ribosyl Cyclase 1; Allergens; Animals; Bone Marrow; Bone Marrow Transplantation; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Chemokines; Chimera; Lung; Methacholine Chloride; Mice, Inbred C57BL; Ovalbumin; Pneumonia; Respiratory Hypersensitivity

Asthma exacerbations contribute to corticosteroid insensitivity. LPS is ubiquitous in the environment. It causes bronchoconstriction and airway inflammation and may therefore exacerbate allergen responses. This study examined whether LPS and ovalbumin co-administration could exacerbate the airway inflammatory and functional responses to ovalbumin in conscious guinea pigs and whether these exacerbated responses were insensitive to inhaled corticosteroid treatment with fluticasone propionate (FP).. Guinea pigs were sensitized and challenged with ovalbumin and airway function recorded as specific airway conductance by whole body plethysmography. Airway inflammation was measured from lung histology and bronchoalveolar lavage. Airway hyper-reactivity (AHR) to inhaled histamine was examined 24 h after ovalbumin. LPS was inhaled alone or 24 or 48 h before ovalbumin and combined with ovalbumin. FP (0.05-1 mg·mL(-1) ) or vehicle was nebulized for 15 min twice daily for 6 days before ovalbumin or LPS exposure.. Ovalbumin inhalation caused early (EAR) and late asthmatic response (LAR), airway hyper-reactivity to histamine and influx of inflammatory cells into the lungs. LPS 48 h before and co-administered with ovalbumin exacerbated the response with increased length of the EAR, prolonged response to histamine and elevated inflammatory cells. FP 0.5 and 1 mg·mL(-1) reduced the LAR, AHR and cell influx with ovalbumin alone, but was ineffective when guinea pigs were exposed to LPS before and with ovalbumin.. LPS exposure exacerbates airway inflammatory and functional responses to allergen inhalation and decreases corticosteroid sensitivity. Its widespread presence in the environment could contribute to asthma exacerbations and corticosteroid insensitivity in humans.

Topics: Administration, Inhalation; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Drug Resistance; Fluticasone; Guinea Pigs; Histamine; Inflammation; Lipopolysaccharides; Lung; Male; Ovalbumin; Plethysmography, Whole Body

Neurturin (NTN) was previously described for its neuronal activities, but recently, we have shown that this factor is also involved in asthma physiopathology. However, the underlying mechanisms of NTN are unclear. The aim of this study was to investigate NTN involvement in acute bronchial Th2 responses, to analyze its interaction with airway structural cells, and to study its implication in remodeling during acute and chronic bronchial inflammation in C57BL/6 mice. We analyzed the features of allergic airway inflammation in wild-type and NTN(-/-) mice after sensitization with two different allergens, OVA and house dust mite. We showed that NTN(-/-) dendritic cells and T cells had a stronger tendency to activate the Th2 pathway in vitro than similar wild-type cells. Furthermore, NTN(-/-) mice had significantly increased markers of airway remodeling like collagen deposition. NTN(-/-) lung tissues showed higher levels of neutrophils, cytokine-induced neutrophil chemoattractant, matrix metalloproteinase 9, TNF-α, and IL-6. Finally, NTN had the capacity to decrease IL-6 and TNF-α production by immune and epithelial cells, showing a direct anti-inflammatory activity on these cells. Our findings support the hypothesis that NTN could modulate the allergic inflammation in different mouse asthma models.

Topics: Airway Remodeling; Animals; Asthma; Blotting, Western; Bronchial Hyperreactivity; Coculture Techniques; Dendritic Cells; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Inflammation; Mice; Mice, Inbred C57BL; Mice, Knockout; Neurturin; Ovalbumin; Real-Time Polymerase Chain Reaction; Th2 Cells

In asthma, airflow obstruction is thought to result primarily from inflammation-triggered airway smooth muscle (ASM) contraction. However, anti-inflammatory and smooth muscle-relaxing treatments are often temporary or ineffective. Overproduction of the mucin MUC5AC is an additional disease feature that, while strongly associated pathologically, is poorly understood functionally. Here we show that Muc5ac is a central effector of allergic inflammation that is required for airway hyperreactivity (AHR) to methacholine (MCh). In mice bred on two well-characterized strain backgrounds (C57BL/6 and BALB/c) and exposed to two separate allergic stimuli (ovalbumin and Aspergillus extract), genetic removal of Muc5ac abolishes AHR. Residual MCh responses are identical to unchallenged controls, and although inflammation remains intact, heterogeneous mucous occlusion decreases by 74%. Thus, whereas inflammatory effects on ASM alone are insufficient for AHR, Muc5ac-mediated plugging is an essential mechanism. Inhibiting MUC5AC may be effective for treating asthma and other lung diseases where it is also overproduced.

Topics: Allergens; Animals; Aspergillus oryzae; Asthma; Bronchial Hyperreactivity; Female; Immunohistochemistry; Inflammation; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Mucin 5AC; Mucus; Ovalbumin; Species Specificity

Asthma is a serious health problem causing significant mortality and morbidity globally. Persistent airway inflammation, airway hyperresponsiveness, increased immunoglobulin E (IgE) levels and mucus hypersecretion are key characteristics of the condition. Asthma is mediated via a dominant T-helper 2 (Th2) immune response, causing enhanced expression of Th2 cytokines. These cytokines are responsible for the various pathological changes associated with allergic asthma. To investigate the anti-asthmatic potential of afzelin, as well as the underlying mechanisms involved, its anti-asthmatic potential were investigated in a murine model of asthma. In the present study, BALB/c mice were systemically sensitized using ovalbumin (OVA) followed by aerosol allergen challenges. The effect of afzelin on airway hyperresponsiveness, eosinophilic infiltration, Th2 cytokine and OVA‑specific IgE production in a mouse model of asthma were investigated. It was found that afzelin‑treated groups suppressed eosinophil infiltration, allergic airway inflammation, airway hyperresponsiveness, OVA-specific IgE and Th2 cytokine secretion. The results of the present study suggested that the therapeutic mechanism by which afzelin effectively treats asthma is based on reduction of Th2 cytokine via inhibition of GATA-binding protein 3 transcription factor, which is the master regulator of Th2 cytokine differentiation and production.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Eosinophils; GATA3 Transcription Factor; Gene Expression Regulation; Humans; Immunoglobulin E; Mannosides; Mice; Ovalbumin; Proanthocyanidins; Th2 Cells

Previous studies investigating the role of toll-like receptors (TLRs) in asthma have been inconclusive. It has remained elusive whether the toll-like receptors (TLR2)/mycloid differentiation factor 88 (MyD88)/nuclear factor (NF)-κB signaling pathway is involved in lipoxin A4 (LXA4)-induced protection against asthma. Therefore, the present study investigated whether ovalbumin (OVA)-induced airway inflammation is mediated by upregulation of the TLR2/MyD88/NF-κB signaling pathway, and whether it proceeds via the inhibition of the activation of the LXA4 receptor and anti-interleukin (IL)-1β antibodies. Mice with airway inflammation induced by OVA administration were treated with or without a LXA4 receptor agonist, BML-111 and anti-IL-1β antibody. Serum levels of IL-1β, IL-4, IL-8 and interferon-γ (IFN-γ) were assessed, and levels of IL-1β, IL-4, IL-8 and OVA-immunoglobulin (Ig)E, as well as leukocyte counts in the bronchoalveolar lavage fluid (BALF) were measured. Pathological features and expression of TLR2, MyD88 and NF-κB in the lungs were analyzed. Expression of TLR2 and MyD88, and activation of NF-κB in leukocytes as well as levels of IL-4, IL-6 and IL-8 released from leukocytes exposed to IL-1β were assessed. OVA treatment increased the levels of IL-1β, IL-4 and IL-8 in the serum and BLAF, the number of leukocytes and the levels of OVA-IgE in the BALF, the expression of TLR2 and MyD88, and the activation of NF-κB in the lung. These increments induced by OVA were inhibited by treatment with BML-111 and anti-IL-1β antibodies. Treatment of the leukocytes with BML-111 or TLR2 antibody, or MyD88 or NF-κB inhibitor, all blocked the IL-1β-triggered production of IL-4, IL-6 and IL-8 and activation of NF-κB. Treatment of the leukocytes with BML-111 or TLR2 antibody suppressed IL-1β-induced TLR2 and MyD88 expression. The present study therefore suggested that OVA-induced airway inflammation is mediated by the TLR2/MyD88/NF-κB pathway. IL-1β has a pivotal role in the airway inflammation and upregulation of the TLR2/MyD88/NF-κB pathway induced by OVA. BML-111 and anti-IL-1β antibody restrains the OVA-induced airway inflammation via downregulation of the TLR2/MyD88/NF-κB pathway.

Topics: Animals; Anti-Asthmatic Agents; Antibodies, Monoclonal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Gene Expression Regulation; Heptanoic Acids; Interleukin-1beta; Interleukin-4; Interleukin-6; Interleukin-8; Lipoxins; Male; Mice; Mice, Inbred BALB C; Myeloid Differentiation Factor 88; NF-kappa B; Ovalbumin; Receptors, Formyl Peptide; Signal Transduction; Toll-Like Receptor 2

Steroid-resistant asthma is a major clinical problem that is linked to activation of innate immune cells. Levels of IFN-γ and LPS are often increased in these patients. Cooperative signaling between IFN-γ/LPS induces macrophage-dependent steroid-resistant airway hyperresponsiveness (AHR) in mouse models. MicroRNAs (miRs) are small noncoding RNAs that regulate the function of innate immune cells by controlling mRNA stability and translation. Their role in regulating glucocorticoid responsiveness and AHR remains unexplored.. IFN-γ and LPS synergistically increase the expression of miR-9 in macrophages and lung tissue, suggesting a role in the mechanisms of steroid resistance. Here we demonstrate the role of miR-9 in IFN-γ/LPS-induced inhibition of dexamethasone (DEX) signaling in macrophages and in induction of steroid-resistant AHR.. MiRNA-9 expression was assessed by means of quantitative RT-PCR. Putative miR-9 targets were determined in silico and confirmed in luciferase reporter assays. miR-9 function was inhibited with sequence-specific antagomirs. The efficacy of DEX was assessed by quantifying glucocorticoid receptor (GR) cellular localization, protein phosphatase 2A (PP2A) activity, and AHR.. Exposure of pulmonary macrophages to IFN-γ/LPS synergistically induced miR-9 expression; reduced levels of its target transcript, protein phosphatase 2 regulatory subunit B (B56) δ isoform; attenuated PP2A activity; and inhibited DEX-induced GR nuclear translocation. Inhibition of miR-9 increased both PP2A activity and GR nuclear translocation in macrophages and restored steroid sensitivity in multiple models of steroid-resistant AHR. Pharmacologic activation of PP2A restored DEX efficacy and inhibited AHR. MiR-9 expression was increased in sputum of patients with neutrophilic but not those with eosinophilic asthma.. MiR-9 regulates GR signaling and steroid-resistant AHR. Targeting miR-9 function might be a novel approach for the treatment of steroid-resistant asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Dexamethasone; Disease Models, Animal; Egg Hypersensitivity; Eosinophils; Gene Expression Regulation; Genes, Reporter; Glucocorticoids; Humans; Interferon-gamma; Lipopolysaccharides; Luciferases; Macrophages, Alveolar; Mice; Mice, Inbred BALB C; MicroRNAs; Neutrophils; Oligonucleotides; Ovalbumin; Primary Cell Culture; Protein Phosphatase 2; Receptors, Glucocorticoid; Signal Transduction

Comorbid asthma in sickle cell disease (SCD) confers higher rates of vaso-occlusive pain and mortality, yet the physiological link between these two distinct diseases remains puzzling. We used a mouse model of SCD to study pulmonary immunology and physiology before and after the induction of allergic airway disease (AAD). SCD mice were sensitized with ovalbumin (OVA) and aluminum hydroxide by the intraperitoneal route followed by daily, nose-only OVA-aerosol challenge to induce AAD. The lungs of naive SCD mice showed signs of inflammatory and immune processes: (1) histologic and cytochemical evidence of airway inflammation compared with naive wild-type mice; (2) bronchoalveolar lavage (BAL) fluid contained increased total lymphocytes, %CD8+ T cells, granulocyte-colony stimulating factor, interleukin 5 (IL-5), IL-7, and chemokine (C-X-C motif) ligand (CXCL)1; and (3) lung tissue and hilar lymph node (HLN) had increased CD4+, CD8+, and regulatory T (Treg) cells. Furthermore, SCD mice at AAD demonstrated significant changes compared with the naive state: (1) BAL fluid with increased %CD4+ T cells and Treg cells, lower %CD8+ T cells, and decreased interferon gamma, CXCL10, chemokine (C-C motif) ligand 2, and IL-17; (2) serum with increased OVA-specific immunoglobulin E, IL-6, and IL-13, and decreased IL-1α and CXCL10; (3) no increase in Treg cells in the lung tissue or HLN; and (4) hyporesponsiveness to methacholine challenge. In conclusion, SCD mice have an altered immunologic pulmonary milieu and physiological responsiveness. These findings suggest that the clinical phenotype of AAD in SCD mice differs from that of wild-type mice and that individuals with SCD may also have a unique, divergent phenotype perhaps amenable to a different therapeutic approach.

Topics: Anemia, Sickle Cell; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophils; Female; Hemizygote; Hypersensitivity; Inflammation; Leukocyte Count; Lung; Mice, Inbred C57BL; Mucus; Ovalbumin; T-Lymphocytes

We previously demonstrated that antigen sensitization increases vulnerability to airway hyperreactivity induced by the organophosphorus pesticide (OP) parathion. Sensitization also changes the mechanism of parathion-induced airway hyperreactivity to one that is dependent on IL-5. To determine whether this effect can be generalized to other OPs, and to other classes of pesticides, we measured airway responsiveness to vagal stimulation or intravenous acetylcholine in nonsensitized and ovalbumin-sensitized guinea pigs 24 hours after a single subcutaneous injection of the OPs diazinon or chlorpyrifos, or the pyrethroid permethrin. Sensitization exacerbated the effects of chlorpyrifos on bronchoconstriction in response to vagal stimulation or intravenous acetylcholine. Pretreatment with function-blocking IL-5 antibody prevented chlorpyrifos-induced airway hyperreactivity in sensitized, but not in nonsensitized, guinea pigs. In sensitized guinea pigs, blocking IL-5 decreased eosinophil activation, as measured by decreased eosinophil major basic protein in the trachea. In contrast, sensitization did not alter diazinon-induced airway hyperreactivity, and permethrin did not cause airway hyperreactivity in either nonsensitized or sensitized guinea pigs. None of the pesticides affected inflammatory cells in the bronchoalveolar lavage fluid or blood. We have previously shown that three different OPs cause airway hyperreactivity via loss of neuronal M2 muscarinic receptor function. Similar to parathion, but unlike diazinon, the mechanism of chlorpyrifos-induced airway hyperreactivity is changed by sensitization. Thus, OP-induced airway hyperreactivity is dependent on sensitization status and on the OP used, which may influence therapeutic approaches.

Topics: Acetylcholine; Animals; Antibodies, Neutralizing; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Chlorpyrifos; Diazinon; Eosinophils; Female; Guinea Pigs; Immunization; Injections, Intravenous; Injections, Subcutaneous; Insecticides; Interleukin-5; Ovalbumin; Permethrin; Trachea; Vagus Nerve

Thalidomide is known to have anti-inflammatory and immunomodulatory actions. However, the effect and the anti-asthmatic mechanism of thalidomide in the pathogenesis of asthmatic airways are not fully understood.. This study is designed to determine the effect and the potential mechanism of thalidomide in the pathogenesis of asthmatic airways using animal model of allergic asthma.. Six-week-old female BALB/C mice were sensitized with alum plus ovalbumin (OVA) and were exposed to OVA via intranasal route for 3 days for challenge. Thalidomide 200 mg/kg was given via gavage twice a day from a day before the challenge and airway hyperresponsivenss (AHR), airway inflammatory cells, and cytokines in bronchoalveolar lavage fluids (BALF) were evaluated. The expression levels of pro-inflammatory cytokines and other mediators were evaluated using ELISA, real time (RT)-qPCR, and flow cytometry. CRL-2456, alveolar macrophage cell line, was used to test the direct effect of thalidomide on the activation of macrophages in vitro.. The mice with thalidomide treatment showed significantly reduced levels of allergen-induced BALF and lung inflammation, AHR, and the expression of a number of pro-inflammatory cytokines and mediators including Th2 related, IL-17 cytokines, and altered levels of allergen-specific IgG1/IgG2a. Of interesting note, thalidomide treatment significantly reduced expression levels of allergen- or Th2 cytokine-stimulated alternative activation of macrophages in vivo and in vitro.. These studies highlight a potential use of thalidomide in the treatment of allergic diseases including asthma. This study further identified a novel inhibitory effect of thalidomide on alternative activation of macrophages as a potential mechanism of anti-asthmatic effect of thalidomide.

Topics: Allergens; Alum Compounds; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Immunoglobulin G; Inflammation; Interleukin-17; Macrophages, Alveolar; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells; Thalidomide

Asthma development and pathogenesis are influenced by the interactions of airway epithelial cells and innate and adaptive immune cells in response to allergens. Oxidative stress is an important mediator of asthmatic phenotypes in these cell types. Nuclear erythroid 2-related factor 2 (Nrf2) is a redox-sensitive transcription factor that is the key regulator of the response to oxidative and environmental stress. We previously demonstrated that Nrf2-deficient mice have heightened susceptibility to asthma, including elevated oxidative stress, inflammation, mucus, and airway hyperresponsiveness (AHR) (Rangasamy T, Guo J, Mitzner WA, Roman J, Singh A, Fryer AD, Yamamoto M, Kensler TW, Tuder RM, Georas SN, Biswal S. J Exp Med 202: 47-59, 2005). Here we dissected the role of Nrf2 in lung epithelial cells and tested whether genetic or pharmacological activation of Nrf2 reduces allergic asthma in mice. Cell-specific activation of Nrf2 in club cells of the airway epithelium significantly reduced allergen-induced AHR, inflammation, mucus, Th2 cytokine secretion, oxidative stress, and airway leakiness and increased airway levels of tight junction proteins zonula occludens-1 and E-cadherin. In isolated airway epithelial cells, Nrf2 enhanced epithelial barrier function and increased localization of zonula occludens-1 to the cell surface. Pharmacological activation of Nrf2 by 2-trifluoromethyl-2'-methoxychalone during the allergen challenge was sufficient to reduce allergic inflammation and AHR. New therapeutic options are needed for asthma, and this study demonstrates that activation of Nrf2 in lung epithelial cells is a novel potential therapeutic target to reduce asthma susceptibility.

Topics: Adaptor Proteins, Signal Transducing; Animals; Asthma; Bronchial Hyperreactivity; Cadherins; Chalcones; Cytokines; Cytoprotection; Cytoskeletal Proteins; Epithelial Cells; Inflammation; Kelch-Like ECH-Associated Protein 1; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; NF-E2-Related Factor 2; Ovalbumin; Oxidative Stress; Respiratory Mucosa; Th2 Cells; Tight Junctions; Zonula Occludens-1 Protein

Bronchial asthma, one of the most common allergic diseases, is characterized by airway hyperresponsiveness (AHR), inflammation, and remodeling. The anti-oxidant flavone aglycone diosmetin ameliorates the inflammation in pancreatitis, but little is known about its impact on asthma. In this study, the effects of diosmetin on chronic asthma were investigated with an emphasis on the modulation of airway remodeling in BALB/c mice challenged with ovalbumin (OVA). It was found that diosmetin significantly relieved inflammatory cell infiltration, goblet cell hyperplasia, and collagen deposition in the lungs of asthmatic mice and notably reduced AHR in these animals. The OVA-induced increases in total cell and eosinophil counts in bronchoalveolar lavage fluid were reversed, and the level of OVA-specific immunoglobulin E in serum was attenuated by diosmetin administration, implying an anti-Th2 activity of diosmetin. Furthermore, diosmetin remarkably suppressed the expression of smooth muscle actin alpha chain, indicating a potent anti-proliferative effect of diosmetin on airway smooth muscle cells (ASMCs). Matrix metallopeptidase-9, transforming growth factor-β1, and vascular endothelial growth factor levels were also alleviated by diosmetin, suggesting that the remission of airway remodeling might be attributed to the decline of these proteins. Taken together, our findings provided a novel profile of diosmetin with anti-remodeling therapeutic benefits, highlighting a new potential of diosmetin in remitting the ASMC proliferation in chronic asthma.

Topics: Airway Remodeling; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Chronic Disease; Disease Models, Animal; Flavonoids; Mice; Mice, Inbred BALB C; Ovalbumin

Carbon monoxide (CO) levels in expired gas are higher in patients with bronchial asthma than in healthy individuals. Heme oxygenase-1 (HO-1) is a rate-limiting enzyme that catalyzes the degradation of heme to yield biliverdin, CO and free iron. Thus, HO-1 is implicated in the pathogenesis of bronchial asthma. However, whether HO-1 expression and activity in lung tissue are related to allergic airway inflammation remains unclear. We investigated whether expression of HO-1 is related to allergic airway inflammation in lungs and whether HO-1 could influence airway hyperresponsiveness and eosinophilia in mice sensitized to ovalbumin (OVA).. C57BL/6 mice immunized with OVA were challenged thrice with an aerosol of OVA every second day for 8 days. HO-1-positive cells were identified by immunostaining in lung tissue, and zinc protoporphyrin (Zn-PP), a competitive inhibitor of HO-1, was administered intraperitoneally to OVA-immunized C57BL/6 mice on day 23 (day before inhalation of OVA) and immediately before inhalation on the subsequent 4 days (total five doses). Mice were analyzed for effects of HO-1 on AHR, inflammatory cell infiltration and cytokine levels in lung tissue. Ethical approval was obtained from the concerned institutional review board.. Number of HO-1-positive cells increased in the subepithelium of the bronchi after OVA challenge, and HO-1 localized to alveolar macrophages. Zn-PP clearly inhibited AHR, pulmonary eosinophilia and IL-5 and IL-13 expression in the lung tissue.. Expression of HO-1 is induced in lung tissue during attacks of allergic bronchial asthma, and its activity likely amplifies and prolongs allergic airway inflammation.

Topics: Acetylcholine; Animals; Asthma; Bronchial Hyperreactivity; CD4 Lymphocyte Count; Disease Models, Animal; Eosinophilia; Heme Oxygenase-1; Inflammation; Lung; Macrophages; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Protoporphyrins

Given the relationship between allergic rhinitis (AR) and asthma, it can be hypothesized that reducing upper airway inflammation by targeting oligodeoxynucleotides with CpG motifs (CpG-ODN) specifically to the upper airway via intranasal administration in a small volume (10 μL) might improve lower airway (asthma) outcomes. The goal of this study was to investigate the therapeutic efficacy of 10 μL of intranasal versus intradermal administration of CpG-ODN in suppressing lower airway inflammation and methacholine-induced airway hyperreactivity (AHR) in mice subjected to ovalbumin (OVA)-induced combined allergic rhinitis and asthma syndrome (CARAS). OVA-sensitized BALB/c mice were subjected to upper-airway intranasal OVA exposure three times per week for 3 weeks. Then, CpG-ODN was administered to a subset of these mice 1h after intranasal OVA exposure, followed by five days of OVA aerosol challenges, thereby targeting OVA to the lower airways. Immunologic variables and nasal symptoms were evaluated. The results showed that the CARAS mice exhibited significant increases in bronchoalveolar lavage fluid (BALF) and splenocytes Th2-associated cytokine production, OVA-specific serum IgE, and AHR, as well as nose and lung pathologies. Intranasal administration of CpG-ODN significantly reduced Th2-associated cytokine production, the percentage of eosinophils in the BALF, the IL-4 and IL-5 concentrations in the supernatants of cultured OVA-challenged splenic lymphocytes, the serum OVA-specific IgE levels, the peribronchial inflammation score in the lungs, and the severity of nose pathology and nasal symptoms. However, intradermal administration of CpG-ODN did not significantly reduce the aforementioned parameters. In conclusion, intranasal treatment with CpG-ODN attenuated AR and significantly alleviated lower airway inflammation and AHR in the CARAS model. CpG-ODN therapy was more effective when administered intranasally than when administered intradermally. The current study supports the development of CpG-ODN nasal spray as a novel therapeutic agent for CARAS.

Topics: Administration, Intranasal; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophilia; Female; Immunoglobulin E; Lymphocytes; Mice, Inbred BALB C; Nasal Mucosa; Oligodeoxyribonucleotides; Ovalbumin; Rhinitis, Allergic; Spleen; Syndrome

An important portion of asthmatics do not respond to current therapies. Thus, the need for new therapeutic drugs is urgent. We have demonstrated a critical role for PARP in experimental asthma. Olaparib, a PARP inhibitor, was recently introduced in clinical trials against cancer. The objective of the present study was to examine the efficacy of olaparib in blocking established allergic airway inflammation and hyperresponsiveness similar to those observed in human asthma in animal models of the disease.. We used ovalbumin (OVA)-based mouse models of asthma and primary CD4(+) T cells. C57BL/6J WT or PARP-1(-/-) mice were subjected to OVA sensitization followed by a single or multiple challenges to aerosolized OVA or left unchallenged. WT mice were administered, i.p., 1 mg/kg, 5 or 10 mg/kg of olaparib or saline 30 min after each OVA challenge.. Administration of olaparib in mice 30 min post-challenge promoted a robust reduction in airway eosinophilia, mucus production and hyperresponsiveness even after repeated challenges with ovalbumin. The protective effects of olaparib were linked to a suppression of Th2 cytokines eotaxin, IL-4, IL-5, IL-6, IL-13, and M-CSF, and ovalbumin-specific IgE with an increase in the Th1 cytokine IFN-γ. These traits were associated with a decrease in splenic CD4(+) T cells and concomitant increase in T-regulatory cells. The aforementioned traits conferred by olaparib administration were consistent with those observed in OVA-challenged PARP-1(-/-) mice. Adoptive transfer of Th2-skewed OT-II-WT CD4(+) T cells reversed the Th2 cytokines IL-4, IL-5, and IL-10, the chemokine GM-CSF, the Th1 cytokines IL-2 and IFN-γ, and ovalbumin-specific IgE production in ovalbumin-challenged PARP-1(-/-)mice suggesting a role for PARP-1 in CD4(+) T but not B cells. In ex vivo studies, PARP inhibition by olaparib or PARP-1 gene knockout markedly reduced CD3/CD28-stimulated gata-3 and il4 expression in Th2-skewed CD4(+) T cells while causing a moderate elevation in t-bet and ifn-γ expression in Th1-skewed CD4(+) T cells.. Our findings show the potential of PARP inhibition as a viable therapeutic strategy and olaparib as a likely candidate to be tested in human asthma clinical trials.

Topics: Adoptive Transfer; Animals; Antigens, CD; Asthma; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cytokines; Disease Models, Animal; Eosinophilia; GATA3 Transcription Factor; Gene Knockout Techniques; Humans; Immunoglobulin E; Mice, Inbred C57BL; Mucus; Ovalbumin; Phthalazines; Piperazines; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Spleen; T-Box Domain Proteins; Th1 Cells; Th2 Cells

Naringin, a well known component isolated from Exocarpium Citri Grandis, has significant antitussive effects. Recently, Naringin exhibited novel anti-inflammatory effect in chronic inflammatory diseases. In this work, we firstly evaluated the effects of naringin on enhanced cough, airway hyper-responsiveness (AHR), and airway inflammation in an ovalbumin-induced experimental cough-variant asthma (CVA) model in guinea pigs. We investigated the effect of naringin (18.4 mg/kg, per os, single dose or consecutively) on cough to inhaled capsaicin after challenge with an aerosolized antigen in actively sensitized guinea pigs. The effect of naringin on AHR to inhaled methacholine was evaluated 24 h after cough determination. Airway inflammation was assessed via bronchoalveolar lavage fluid (BALF) cytology and lung histopathology. Naringin, given consecutively, significantly reduced ovalbumin-induced enhanced cough and AHR, inhibited the increases in the leukocytes, interleukin-4 (IL-4), IL-5, and IL-13 in BALF compared with the model group. Moreover, the pathologic changes in lung tissues were clearly ameliorated by naringin treatment. These results suggest that naringin may be a beneficial agent for CVA treatment.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Capsaicin; Cough; Disease Models, Animal; Flavanones; Guinea Pigs; Inflammation; Male; Ovalbumin

Compelling evidence suggests that hydrogen sulfide represents an important gaseous transmitter in the mammalian respiratory system. In the present study, we have evaluated the role of mast cells in hydrogen sulfide-induced effects on airways in a mouse model of asthma. Mice were sensitized to ovalbumin and received aerosol of a hydrogen sulfide donor (NaHS; 100 ppm) starting at day 7 after ovalbumin challenge. Exposure to hydrogen sulfide abrogated ovalbumin-induced bronchial hypereactivity as well as the increase in lung resistance. Concomitantly, hydrogen sulfide prevented mast cell activity as well as FGF-2 and IL-13 upregulation. Conversely, pulmonary inflammation and the increase in plasmatic IgE levels were not affected by hydrogen sulfide. A lack of hydrogen sulfide effects in mast cell deficient mice occurred. Primary fibroblasts harvested from ovalbumin-sensitized mice showed an increased proliferation rate that was inhibited by hydrogen sulfide aerosol. Furthermore, ovalbumin-induced transdifferentiation of pulmonary fibroblasts into myofibroblasts was reversed. Finally, hydrogen sulfide did abrogate in vitro the degranulation of the mast cell-like RBL-2H3 cell line. Similarly to the in vivo experiments the inhibitory effect was present only when the cells were activated by antigen exposure. In conclusion, inhaled hydrogen sulfide improves lung function and inhibits bronchial hyper-reactivity by modulating mast cells and in turn fibroblast activation.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cell Transdifferentiation; Disease Models, Animal; Hydrogen Sulfide; Immunoglobulin E; Lung; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin

We have previously shown that T helper type 2 (Th2)-polarized airway inflammation can facilitate priming to new antigens in the lungs, which we called "collateral priming". To investigate whether allergic skin inflammation can also facilitate priming toward new antigens, we developed an allergic skin inflammation model based on an allergic lung inflammation model. Mice were sensitized intraperitoneally toward the primary antigen, ovalbumin. Challenge was subsequently performed intranasally or epicutaneously with ovalbumin and a secondary antigen, keyhole limpet hemocyanin (KLH). Re-challenge consisted of local application of either antigen alone. Analysis of KLH-specific antibody responses, KLH-specific cytokines, and local inflammation demonstrated tolerance induction toward the secondary antigen in the skin, whereas in the lung priming had occurred. Flow-cytometric analysis revealed increased numbers of regulatory T cells (Tregs), increased cytotoxic T lymphocyte antigen-4 (CTLA-4) expression, and an enhanced suppressive capacity of Tregs from skin-draining lymph nodes when compared with Tregs from the lung-draining lymph nodes. Furthermore, depletion of Tregs resulted in restoration of collateral priming in the skin. These results demonstrate crucial local differences between the Treg function in the skin and lung to repetitive antigen exposure, which can decisively influence the immune response toward new antigens.

Topics: Analysis of Variance; Animals; Bronchial Hyperreactivity; Cytokines; Dermatitis; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immune Tolerance; Immunization; Lung; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Ovalbumin; Random Allocation; Skin; T-Lymphocytes, Regulatory; Th2 Cells

Viral infections are a common cause of asthma exacerbation. These maladies are sometimes complicated by bacterial infections. Toll-like receptors (TLRs) are in the forefront of our microbial defence, with TLR3 responding to viral and TLR4 to bacterial stimulation. The present study was designed to evaluate the effect of concomitant TLR3 and TLR4 stimulation in a murine model of allergic asthma.BALB/c mice were stimulated intranasally with a combination of poly(I:C) and LPS activating TLR3 and TLR4, respectively. This resulted in the development of airway hyperresponsiveness (AHR) in the proximal part of the lung, along with signs of neutrophilic inflammation. Analysis of the bronchioalveolar lavage fluid (BALF) revealed a marked increase in TNFα. In contrast, the allergic airway inflammation induced by ovalbumin administration to sensitized mice caused AHR in the whole lung along with an increase in eosinophils and lymphocytes in the BALF and lung.When poly(I:C) + LPS were given to mice with an ongoing allergic airway inflammation induced by ovalbumin, the AHR was further increased in the peripheral lung and neutrophils appeared together with eosinophils and lymphocytes in the BALF and lung. Treatment with the TNFα-blocking antibody infliximab blunted the AHR increase, without affecting the cells influx in BALF.To conclude; a combined TLR3- and TLR4-stimulation, representing a concomitant viral and bacterial infection, causes an AHR that is further exaggerated during an ongoing allergic inflammation. The airway stabilizing effect of infliximab indicates the possible future use of TNFα blockade in treatment of microbial induced exacerbations of allergic asthma.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Disease Models, Animal; Disease Progression; Eosinophils; Female; Immunity, Innate; Infliximab; Lipopolysaccharides; Lung; Lymphocytes; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Pneumonia; Poly I-C; Signal Transduction; Toll-Like Receptor 3; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

The anti-malaria drug artesunate has been shown to attenuate experimental allergic asthma via inhibition of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway. This study was to further determine the effects of artesunate on cigarette smoke and ovalbumin (OVA) concurrent exposure-induced airway inflammation, the related mechanism, and glucocorticoid insensitivity.. In vivo: Female BALB/c mice concurrently exposed to cigarette smoke and OVA developed mixed eosinophilic and neutrophilic airway inflammation. Airway hyper-responsiveness, total and differential cell counts, and pro-inflammatory cytokine levels (interleukin (IL)-4, IL-8, IL-13 and tumor necrosis factor (TNF)-α) in bronchoalveolar lavage fluid (BALF) were measured. Lung tissue sections were stained for histological analysis, and proteins were extracted for Western blotting. Artesunate reduced methacholine-induced airway hyper-responsiveness, suppressed pulmonary inflammation cell recruitment and IL-4, IL-8, IL-13 and TNF-α levels, selectively inhibited PI3Kδ/Akt pathway, and restored HDAC2 activity. In vitro: BEAS-2B cells were exposed to cigarette smoke extract (CSE) for 6h and then stimulated with TNF-α overnight. Glucocorticoid sensitivity was evaluated by the inhibition of TNF-α-induced IL-8 production by dexamethasone. CSE reduced the effects of dexamethasone on TNF-α-induced IL-8 production in BEAS-2B cells, while artesunate reversed CSE-induced glucocorticoid insensitivity and restored HDAC2 deactivation induced by CSE.. Artesunate ameliorated cigarette smoke and OVA concurrent exposure-induced airway inflammation, inhibited the PI3Kδ/Akt pathway, restored HDAC2 activity, and reversed CSE-induced glucocorticoid insensitivity in BEAS-2B cells. These findings indicate that artesunate might play a protective role in asthma induced by cigarette smoke and glucocorticoid insensitivity.

Topics: Animals; Antimalarials; Artemisinins; Artesunate; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Line; Cytokines; Dexamethasone; Drug Resistance; Eosinophils; Female; Glucocorticoids; Inflammation; Lung; Mice; Mice, Inbred BALB C; Nicotiana; Ovalbumin; Respiratory Tract Diseases; Smoke

Gata5 is a transcription factor expressed in the lung, but its physiological role is unknown. To test whether and how Gata5 regulates airway constrictor responsiveness, we studied Gata5(-/-), Gata5(+/-), and wild-type mice on the C57BL/6J background. Cholinergic airway constrictor responsiveness was assessed invasively in mice without and with induction of allergic airway inflammation through ovalbumin sensitization and aerosol exposure. Gata5-deficient mice displayed native airway constrictor hyperresponsiveness (AHR) in the absence of allergen-induced inflammation. Gata5-deficient mice retained their relatively greater constrictor responsiveness even in ovalbumin-induced experimental asthma. Gata5 deficiency did not alter the distribution of cell types in bronchoalveolar lavage fluid, but bronchial epithelial mucus metaplasia was more prominent in Gata5(-/-) mice after allergen challenge. Gene expression profiles revealed that apolipoprotein E (apoE) was the fifth most down-regulated transcript in Gata5-deficient lungs, and quantitative RT-PCR and immunostaining confirmed reduced apoE expression in Gata5(-/-) mice. Quantitative RT-PCR also revealed increased IL-13 mRNA in the lungs of Gata5-deficient mice. These findings for the first time show that Gata5 regulates apoE and IL-13 expression in vivo and that its deletion causes AHR. Gata5-deficient mice exhibit an airway phenotype that closely resembles that previously reported for apoE(-/-) mice: both exhibit cholinergic AHR in native and experimental asthma states, and there is excessive goblet cell metaplasia after allergen sensitization and challenge. The Gata5-deficient phenotype also shares features that were previously reported for IL-13-treated mice. Together, these results indicate that Gata5 deficiency induces AHR, at least in part, by blunting apoE and increasing IL-13 expression.

Topics: Animals; Apolipoproteins E; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Disease Models, Animal; GATA5 Transcription Factor; Gene Expression Regulation; Genotype; Goblet Cells; Interleukin-13; Lung; Metaplasia; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocytes, Smooth Muscle; Ovalbumin; Phenotype; Pneumonia

Intravenous immunoglobulin (IVIg) is a polyclonal IgG preparation with potent immunomodulating properties. Our laboratory demonstrated that IVIg significantly increases numbers of forkhead box protein 3-positive regulatory T (Treg) cells through generation of tolerogenic dendritic cells (DCs) in an allergic airways disease model.. We sought to investigate potential receptors on DCs mediating these events.. C57BL/6 mice were either sensitized to ovalbumin (OVA) intraperitoneally or through adoptive transfer of OVA-primed DCs and then challenged with intranasal OVA. IVIg was fractionated into sialic acid-enriched IVIg (SA-IVIg) and sialic acid-depleted IVIg (non-SA-IVIg). Dendritic cell immunoreceptor (DCIR) constructs in CHO cells or on DCs were examined by using fluorescent microscopy and flow cytometry.. Administration of SA-IVIg, but not non-SA-IVIg, to OVA-sensitized and OVA-challenged mice induced Treg cells and attenuated airway hyperresponsiveness (AHR) and inflammation comparably with IVIg. Bone marrow-derived dendritic cells cultured with SA-IVIg or IVIg adoptively transferred to mice before OVA challenge induced Treg cells and inhibited AHR. IVIg-treated bone marrow-derived dendritic cells from Fcγ receptor knockout mice inhibited AHR, suggesting IVIg's action was not caused by Fcγ receptor-mediated events. Fluorescently labeled IVIg or SA-IVIg bound DCs and colocalized specifically to the C-type lectin DCIR. IVIg binding to DCIR induced phosphorylation of Src homology domain 2-containing protein tyrosine phosphatase (SHP) 2 and Src homology domain 2-containing inositol phosphatase 1 (SHIP-1) and internalization of IVIg into DCs. Inhibition of IVIg binding to DCIR by small interfering RNA completely blocked induction of Treg cells. Inhibition of SHP-2 or abrogation of IgG internalization through clatherin inhibitors rendered IVIg ineffective.. IVIg alleviates allergic airways disease through interaction of SA-IgG with DCIR. DCIR is a novel receptor for IVIg, mediating interaction of innate and adaptive immunity in tolerogenic responses.

Topics: Animals; Bronchial Hyperreactivity; CHO Cells; Cricetulus; Dendritic Cells; Immunoglobulin G; Immunoglobulins, Intravenous; Inositol Polyphosphate 5-Phosphatases; Lectins, C-Type; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Ovalbumin; Phosphatidylinositol-3,4,5-Trisphosphate 5-Phosphatases; Phosphoric Monoester Hydrolases; Protein Tyrosine Phosphatase, Non-Receptor Type 11; Receptors, Immunologic; T-Lymphocytes, Regulatory

The cytosolic nucleotide oligomerization domain (NOD)-like receptors NOD1 and NOD2 are important contributors to the intracellular recognition of pathogens including Chlamydophila pneumoniae, but little is known about their influence on allergen-induced airway inflammation. In BALB/c mice, we observed that infection with C. pneumoniae before systemic sensitization with ovalbumin (OVA) and local OVA airway exposure diminished airway hyperresponsiveness (AHR). Thus, the impact of the NOD1 agonist FK156 and the NOD2 agonist muramyl dipeptide given 6 hours before each sensitization or airway challenge was evaluated regarding AHR, OVA-specific plasma immunoglobulins, bronchoalveolar lavage fluid differentials, and cytokines. Spleen dendritic cells of FK156-treated mice were isolated and cocultured with OVA-specific T cells isolated from DO11.10 mice, and T-cell proliferation was quantified after OVA restimulation. T-cell proliferation was investigated in vivo in lungs and lymph nodes of FK156-treated and OVA-exposed DO11.10 mice. FK156, but not muramyl dipeptide, reduced AHR and pulmonary eosinophilic infiltration if given before OVA sensitization or challenge, whereas T-helper (Th)2 cytokines were not diminished. Dendritic cells from FK156-treated mice evoked less OVA-specific T-cell proliferation as compared with solvent-treated controls. Similarly, antigen-specific T-cell activation in lung tissue was diminished after FK156 treatment. We conclude that NOD1 activation reduced AHR in allergen-induced lung inflammation, which was accompanied by a reduction of allergen-specific T-cell proliferation.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cell Proliferation; Chlamydophila pneumoniae; Cytokines; Dendritic Cells; Female; Immunoglobulins; Inflammation; Ligation; Lung; Lymph Nodes; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Nod1 Signaling Adaptor Protein; Nod2 Signaling Adaptor Protein; Ovalbumin; Respiratory System; Th2 Cells

Pothos scandens L. was used in Indian traditional medicine as an antiasthmatic drug. The ethanolic and aqueous extracts were prepared with aerial parts of P. scandens (PSE & PSA). ESI MS/MS of PSE ethanolic extract was carried out for the determination of chemical constituents. CP1 is isolated from the PSE, structurally confirmed with NMR and LCMS/MS. PSE, PSA and CP1 are evaluated against ovalbumin (OVA) induced airway hyperresponsiveness (AHR) in balb/c mice. The test drugs are administered p.o. prior to challenge with aerosolized 2.5% w/v OVA. Total and differential leucocyte count, nitrite (NO2), nitrate (NO3), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and interleukin-13 (IL-13) are estimated in bronchoalveolar lavage fluid (BALF). Similarly, myeloperoxidase (MPO), malonaldehyde (MDA) and total lung protein (TLP) are estimated in the lungs. The results reveal a significant increase in total and differential leucocyte count, NO2, NO3, TNF-α, IL-6, and IL-13 in OVA induced AHR. However, these parameters are significantly decreased in PSE and PSA tested doses (PSE 100 & 200mg/kg). While, treatment with CP1 is less effective at 5 & 10mg/kg doses. Similar observations obtain for MPO and MDA in lungs. However, the mean value indicated that the PSE at 200mg/kg showed a significant restoration in all the parameters. Pro-inflammatory mediators are known to be responsible for AHR. Histopathology revealed justifies the effectiveness. The present investigations suggest PSE are interesting molecules for further research for asthma, with an approach through pro-inflammatory inhibitory pathway. P. scandens is a potential herbal medicine for allergy induced asthma.

Topics: Allergens; Animals; Araceae; Bronchial Hyperreactivity; Cells, Cultured; Cytokines; Down-Regulation; Female; Humans; Inflammation Mediators; Leukocytes; Lung; Male; Malondialdehyde; Medicine, Traditional; Mice; Mice, Inbred BALB C; Nitrates; Ovalbumin; Peroxidase; Phytotherapy; Plant Extracts

Aspergillus fumigatus is often associated in asthmatic patients with the exacerbation of asthma symptoms. The pathomechanism of this phenomenon has not been fully understood. Here, we evaluated the immunological mechanisms and the role of the prostaglandin D2 / Chemoattractant Receptor-Homologous Molecule Expressed on Th2 Cells (CRTH2) pathway in the development of Aspergillus-associated asthma exacerbation. We studied the effects of A. fumigatus on airway inflammation and bronchial hyper-responsiveness in a rat model of chronic asthma. Inhalation delivery of A. fumigatus conidia increased the airway eosinophilia and bronchial hyper-responsiveness in ovalbumin-sensitized, challenged rats. These changes were associated with prostaglandin D2 synthesis and CRTH2 expression in the lungs. Direct inflammation occurred in ovalbumin-sensitized, challenged animals, whereas pre-treatment with an antagonist against CRTH2 nearly completely eliminated the A. fumigatus-induced worsening of airway eosinophilia and bronchial hyper-responsiveness. Our data demonstrate that production of prostaglandin D2 followed by eosinophil recruitment into the airways via a CRTH2 receptor are the major pathogenic factors responsible for the A. fumigatus-induced enhancement of airway inflammation and responsiveness.

Topics: Animals; Anti-Inflammatory Agents; Aspergillus fumigatus; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophils; Lung; Male; Ovalbumin; Prostaglandin D2; Pulmonary Aspergillosis; Pulmonary Eosinophilia; Rats; Rats, Wistar; Receptors, Immunologic; Receptors, Prostaglandin; Signal Transduction

Exposure to dogs in early infancy has been shown to reduce the risk of childhood allergic disease development, and dog ownership is associated with a distinct house dust microbial exposure. Here, we demonstrate, using murine models, that exposure of mice to dog-associated house dust protects against ovalbumin or cockroach allergen-mediated airway pathology. Protected animals exhibited significant reduction in the total number of airway T cells, down-regulation of Th2-related airway responses, as well as mucin secretion. Following dog-associated dust exposure, the cecal microbiome of protected animals was extensively restructured with significant enrichment of, amongst others, Lactobacillus johnsonii. Supplementation of wild-type animals with L. johnsonii protected them against both airway allergen challenge or infection with respiratory syncytial virus. L. johnsonii-mediated protection was associated with significant reductions in the total number and proportion of activated CD11c(+)/CD11b(+) and CD11c(+)/CD8(+) cells, as well as significantly reduced airway Th2 cytokine expression. Our results reveal that exposure to dog-associated household dust results in protection against airway allergen challenge and a distinct gastrointestinal microbiome composition. Moreover, the study identifies L. johnsonii as a pivotal species within the gastrointestinal tract capable of influencing adaptive immunity at remote mucosal surfaces in a manner that is protective against a variety of respiratory insults.

Topics: Animals; Bronchial Hyperreactivity; Dogs; Dust; Environmental Exposure; Flow Cytometry; Fluorescence; Gastrointestinal Tract; Lactobacillus; Lung; Mice; Mice, Inbred BALB C; Microbiota; Ovalbumin; Respiratory Syncytial Virus Infections; Reverse Transcriptase Polymerase Chain Reaction; Th2 Cells

Endogenous mediators, such as RvE1 (resolvin E1), promote resolution of an inflammatory response and have potential as novel therapeutic agents. In the present study, we investigated the activity of RvE1 in a model of an acute exacerbation of chronic allergic asthma in mice. Animals sensitized to OVA (ovalbumin) received controlled low-level challenge with aerosolized antigen for 4 weeks, followed by a single moderate-level challenge to simulate an allergen-induced exacerbation of asthmatic inflammation. Induction of an exacerbation was associated with rapid recruitment of neutrophils, lymphocytes and eosinophils, together with increased levels of Th2 and pro-inflammatory cytokines. When administered before the final moderate-level challenge, RvE1 had only a modest effect on airway inflammation. To assess its effects when administered after induction of an exacerbation, we first characterized the cellular and molecular events associated with spontaneous resolution of airway inflammation over the following 96 h. Subsequently, we showed that administration of RvE1 at 2 and 8 h after the final challenge accelerated this process significantly. Specifically, RvE1 promoted a decline in the number of inflammatory cells, concentration of cytokines in lavage fluid and expression of mRNA for cytokines by macrophages, confirming its pro-resolution activity. In vitro, RvE1 had no apparent effect on lymphocytes, but suppressed significantly cytokine production by pulmonary macrophages, with evidence of down-regulation of the nuclear translocation of NF-κB (nuclear factor κB) p65 in these cells. The present study provides novel evidence that RvE1 can facilitate resolution of airway inflammation in a clinically relevant model of an acute exacerbation of asthma, possibly via its effects on activated pulmonary macrophages.

Topics: Active Transport, Cell Nucleus; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Line; Cytokines; Disease Models, Animal; Eicosapentaenoic Acid; Female; Hypersensitivity; Immunohistochemistry; Inflammation; Leukocytes; Macrophages; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; RNA, Messenger; Time Factors

Most asthma begins in the first years of life. This early onset cannot be attributed merely to genetic factors because the prevalence of asthma is increasing. Epidemiologic studies have indicated roles for prenatal and early childhood exposures, including exposure to diesel exhaust. However, little is known about the mechanisms. This is largely due to a paucity of animal models.. We aimed to develop a mouse model of asthma susceptibility through prenatal exposure to diesel exhaust.. Pregnant C57BL/6 female mice were given repeated intranasal applications of diesel exhaust particles (DEPs) or PBS. Offspring underwent suboptimal immunization and challenge with ovalbumin (OVA) or received PBS. Pups were examined for features of asthma; lung and liver tissues were analyzed for transcription of DEP-regulated genes.. Offspring of mice exposed to DEPs were hypersensitive to OVA, as indicated by airway inflammation and hyperresponsiveness, increased serum OVA-specific IgE levels, and increased pulmonary and systemic TH2 and TH17 cytokine levels. These cytokines were primarily produced by natural killer (NK) cells. Antibody-mediated depletion of NK cells prevented airway inflammation. Asthma susceptibility was associated with increased transcription of genes known to be specifically regulated by the aryl hydrocarbon receptor and oxidative stress. Features of asthma were either marginal or absent in OVA-treated pups of PBS-exposed mice.. We created a mouse model that linked maternal exposure to DEPs with asthma susceptibility in offspring. Development of asthma was dependent on NK cells and associated with increased transcription from aryl hydrocarbon receptor- and oxidative stress-regulated genes.

Topics: Air Pollutants; Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Disease Susceptibility; Female; Gene Expression Profiling; Gene Expression Regulation; Humans; Immunoglobulin E; Interleukin-17; Killer Cells, Natural; Maternal Exposure; Mice; Mice, Inbred C57BL; Ovalbumin; Pregnancy; Prenatal Exposure Delayed Effects; Receptors, Aryl Hydrocarbon; Transcription, Genetic; Vehicle Emissions

The natural product oleanolic acid is commonly found in a variety of medicinal plants. It is a triterpenoid compound known for its anti-inflammatory effects as well as various other pharmacological properties. The aim of the current study was to use a mouse model of allergic asthma to investigate whether oleanolic acid has anti-asthmatic effects, and if so, to determine the mechanism of these effects. Oleanolic acid suppressed eosinophil infiltration, allergic airway inflammation, and Penh, which occurred by suppressing the production of IL-5, IL-13, IL-17, and ovalbumin-specific IgE through the upregulation of T-bet and Foxp3, and the downregulation of GATA-3 and RORγt. The therapeutic effect of oleanolic acid was due to suppression of Th2 cytokines (IL-5 and IL-13), B cell-dependent production of OVA-specific IgE, and Gr-1 expression through the T-bet, GATA-3, retinoic acid-related orphan receptor γ t (RORγ t) and forkhead box p3 (Foxp3) transcription pathways. It is therefore reasonable to suggest that the anti-inflammatory and anti-asthmatic effects of oleanolic acid may be exerted through inhibition of the GATA-3 and RORγt pathways.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cell Line, Tumor; Cytokines; Eosinophils; Female; Forkhead Transcription Factors; GATA3 Transcription Factor; Lung; Mice; Mice, Inbred BALB C; Neutrophils; Nuclear Receptor Subfamily 1, Group F, Member 3; Oleanolic Acid; Ovalbumin; Pneumonia; T-Box Domain Proteins; Th2 Cells

Activation of poly(ADP-ribose) polymerases (PARPs) is considered a key event in the molecular and cellular processes leading from acute asthma attacks to bronchial hyper-reactivity, leucocyte recruitment, chronic inflammation, airway remodelling and lung damage. The present investigation has been carried out to investigate the action of hydroxyl-dimethylaminomethyl-thieno[2,3-c]isoquinolin-5(4H)-one (HYDAMTIQ), a new potent PARP inhibitor, in the process leading from asthma-like events to airway damage. Ovalbumin-sensitized guinea pigs exposed two times to allergen inhalation were treated for 8 days with vehicle or HYDAMTIQ. Asthma-like signs, bronchial hyper-reactivity to methacholine, cytokine production, histamine release from mast cells, airway remodelling, collagen deposition and lung damage were evaluated. Repeated HYDAMTIQ administration (1-10 mg/kg/day i.p.) reduced lung PARP activity, delayed the appearance and reduced the severity of allergen-induced cough and dyspnoea and dampened the increased bronchial responses to methacholine. HYDAMTIQ-treated animals presented reduced bronchial or alveolar abnormalities, lower number of eosinophils and other leucocytes in the lung and decreased smooth muscle or goblet cell hyperplasia. The treatment also reduced lung oxidative stress markers, such as malondialdehyde or 8-hydroxy-2'-deoxyguanosine and the lung content of pro-inflammatory cytokines (TNF-α, interleukin (IL)-1β, IL-5, IL-6 and IL-18). Finally, mast cells isolated from the peritoneal or pleural cavities of sensitized, HYDAMTIQ-treated animals had a reduced ability to release histamine when exposed to ovalbumin in vitro. Our findings support the proposal that PARP inhibitors could have a therapeutic potential to reduce chronic lung inflammation, airway damage and remodelling in severe unresponsive asthmatic patients.

Topics: Airway Remodeling; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Enzyme Inhibitors; Guinea Pigs; Histamine Release; Inflammation; Isoquinolines; Leukocytes; Lung; Mast Cells; Ovalbumin; Oxidative Stress; Poly Adenosine Diphosphate Ribose; Poly(ADP-ribose) Polymerase Inhibitors; Poly(ADP-ribose) Polymerases; Thiophenes

Allergic asthma is a complex disease characterized by acute inflammation of the airways that over time leads to the development of significant structural changes termed remodeling. TNF-related apoptosis-inducing ligand (TRAIL) has an important regulatory role in acute allergic airways inflammation through up-regulation of the E3 ubiquitin ligase Midline-1 (MID-1), which limits protein phosphatase 2A (PP2A) activity and downstream dephosphorylation of proinflammatory signaling molecules. The relevance of TRAIL in the development of airways remodeling has yet to be determined. In this study, the lungs of wild-type (WT) BALB/c and Tnfsf10 knockout (TRAIL-/-) mice were chronically exposed to ovalbumin (OVA) for 12 weeks to induce hallmark features of chronic allergic airways disease, including airways hyperreactivity (AHR), subepithelial collagen deposition, goblet cell hyperplasia, and smooth muscle hypertrophy. TRAIL-/- mice were largely protected from the development of AHR and peribronchial eosinophilia and had reduced levels of mast cells in the airways. This correlated with lower levels of cytokines, including IL-4, -5, -10, and -13, and with lower levels of proinflammatory chemokines from cultured cells isolated from the draining lymph nodes. TRAIL-/- mice were also protected from the characteristic features of airways remodeling, including peribronchial fibrosis, smooth muscle hypertrophy, and mucus hypersecretion, which correlated with reduced TGF-β1 levels in the lungs. MID-1 expression was reduced in TRAIL-/- mice and up-regulated in allergic WT mice. Raising PP2A activity using 2-amino-4-(4-heptyloyphenol)-2-methylbutan-1-ol in allergic WT mice reduced eosinophilia, TGF-β1, and peribronchial fibrosis. This study shows that TRAIL promotes airways remodeling in an OVA-induced model of chronic allergic airways disease. Targeting TRAIL and its downstream proinflammatory signaling pathway involving PP2A may be of therapeutic benefit in reducing the hallmark features of airways remodeling observed in chronic allergic airways inflammation.

Topics: Airway Remodeling; Animals; Apoptosis; Blotting, Western; Bronchial Hyperreactivity; Cytokines; Immunoenzyme Techniques; Mice; Mice, Inbred BALB C; Mice, Knockout; Mucus; Muscle, Smooth; Ovalbumin; Protein Phosphatase 2; Proteins; Pulmonary Eosinophilia; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; TNF-Related Apoptosis-Inducing Ligand; Transforming Growth Factor beta1; Ubiquitin-Protein Ligases

Low-level laser therapy (LLLT) controls bronchial hyperresponsiveness (BHR) associated with increased RhoA expression as well as pro-inflammatory mediators associated with NF-kB in acute lung inflammation. Herein, we explore if LLLT can reduce both BHR and Th2 cytokines in allergic asthma. Mice were studied for bronchial reactivity and lung inflammation after antigen challenge. BHR was measured through dose-response curves to acetylcholine. Some animals were pretreated with a RhoA inhibitor before the antigen. LLLT (660 nm, 30 mW and 5.4 J) was applied on the skin over the right upper bronchus and two irradiation protocols were used. Reduction of BHR post LLLT coincided with lower RhoA expression in bronchial muscle as well as reduction in eosinophils and eotaxin. LLLT also diminished ICAM expression and Th2 cytokines as well as signal transducer and activator of transduction 6 (STAT6) levels in lungs from challenged mice. Our results demonstrated that LLLT reduced BHR via RhoA and lessened allergic lung inflammation via STAT6.

Topics: Airway Remodeling; Amides; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoconstriction; Cytokines; Enzyme Inhibitors; Hypersensitivity; Low-Level Light Therapy; Lung; Male; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Pneumonia; Pyridines; rho GTP-Binding Proteins; rhoA GTP-Binding Protein; STAT6 Transcription Factor

Infants born prematurely are often treated with supplemental oxygen, which can increase their risk for airway hyper-responsiveness (AHR), asthma, reduced lung function, and altered responses to respiratory viral infections later in childhood. Likewise, exposure of newborn mice to hyperoxia alters baseline pulmonary mechanics and the host response to influenza A virus infection in adult mice. Here, we use this mouse model to test the hypothesis that neonatal hyperoxia also promotes AHR and exacerbated allergen-induced symptoms in adult mice.. Baseline lung mechanics and AHR measured by methacholine provocation were assessed in adult male and female mice exposed to room air or 100% oxygen (hyperoxia) between post-natal days 0-4. AHR and lung inflammation were evaluated after adult female mice were sensitized with ovalbumin (OVA) plus alum and challenged with aerosolized OVA.. Baseline lung compliance increased and resistance decreased in adult female, but not male, mice exposed to neonatal hyperoxia compared with siblings exposed to room air. Neonatal hyperoxia significantly enhanced methacholine-induced AHR in female mice, but did not affect allergen-induced AHR to methacholine or lung inflammation.. Increased incidence of AHR and asthma is reported in children born prematurely and exposed to supplemental oxygen. Our findings in adult female mice exposed to hyperoxia as neonates suggest that this AHR reported in children born prematurely may reflect non-atopic wheezing due to intrinsic structural changes in airway development.

Topics: Age Factors; Airway Resistance; Animals; Animals, Newborn; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Bronchoconstrictor Agents; Disease Models, Animal; Female; Hyperoxia; Lung; Lung Compliance; Male; Methacholine Chloride; Mice, Inbred C57BL; Molecular Sequence Data; Ovalbumin; Pneumonia; Risk Factors; Sex Factors

The flavonoid naringenin has been shown to attenuate airway inflammation and airway hyper‑reactivity in acute murine models of asthma. The purpose of this study was to investigate the effects of naringenin in allergen‑induced airway remodeling in mice. Ovalbumin (OVA)‑sensitized mice were challenged with OVA for 8 weeks to produce a model of chronic asthma. Airway hyper-responsiveness (AHR), inflammation and remodeling were evaluated in mice receiving naringenin prior to OVA challenge. Compared to OVA-sensitized and -challenged mice, those treated with naringenin showed markedly attenuated chronic inflammation, persistent AHR and airway remodeling. In addition, naringenin treatment caused a significant reduction in the levels of total serum IgE and of T helper 2 (Th2) cytokines in the bronchoalveolar lavage fluid (BALF). Naringenin may thus delay the progression of airway remodeling, providing a potential treatment for asthma.

Topics: Airway Remodeling; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Differentiation; Chronic Disease; Cytokines; Disease Models, Animal; Female; Fibrosis; Flavanones; Immunoglobulin E; Inflammation; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Th2 Cells

Orosomucoid-like (ORMDL)3 has been strongly linked with asthma in genetic association studies. Because allergen challenge induces lung ORMDL3 expression in wild-type mice, we have generated human ORMDL3 zona pellucida 3 Cre (hORMDL3(zp3-Cre)) mice that overexpress human ORMDL3 universally to investigate the role of ORMDL3 in regulating airway inflammation and remodeling. These hORMDL3(zp3-Cre) mice have significantly increased levels of airway remodeling, including increased airway smooth muscle, subepithelial fibrosis, and mucus. hORMDL3(zp3-Cre) mice had spontaneously increased airway responsiveness to methacholine compared to wild-type mice. This increased airway remodeling was associated with selective activation of the unfolded protein response pathway transcription factor ATF6 (but not Ire1 or PERK). The ATF6 target gene SERCA2b, implicated in airway remodeling in asthma, was strongly induced in the lungs of hORMDL3(zp3-Cre) mice. Additionally, increased levels of expression of genes associated with airway remodeling (TGF-β1, ADAM8) were detected in airway epithelium of these mice. Increased levels of airway remodeling preceded increased levels of airway inflammation in hORMDL3(zp3-Cre) mice. hORMDL3(zp3-Cre) mice had increased levels of IgE, with no change in levels of IgG, IgM, and IgA. These studies provide evidence that ORMDL3 plays an important role in vivo in airway remodeling potentially through ATF6 target genes such as SERCA2b and/or through ATF6-independent genes (TGF-β1, ADAM8).

Topics: Activating Transcription Factor 6; Airway Remodeling; Allergens; Animals; Antibody Specificity; Asthma; Bronchial Hyperreactivity; Chemokines, CC; Chemokines, CXC; Cytokines; Disease Models, Animal; eIF-2 Kinase; Eosinophils; Gene Expression; Gene Order; Gene Targeting; Humans; Immunoglobulin E; Inflammation; Lung; Membrane Proteins; Methacholine Chloride; Mice; Mice, Transgenic; Ovalbumin; Th2 Cells; Transgenes; Unfolded Protein Response

Astragaloside IV is the chief ingredient of Radix Astragali, which has been used in the Traditional Chinese Medicine as a major component of many polyherbal formulations for the repair and regeneration of injured organ and tissues. We tested the anti-asthmatic effects of AST IV and the possible mechanisms. BALB/c mice that were sensitized and challenged to ovalbumin (OVA) were treated with AST IV (40mg/kg and 20mg/kg) 1h before they were challenged with OVA. Our study demonstrated that AST IV inhibited OVA-induced increases in eosinophil count; interleukin (IL)-4 level were recovered in bronchoalveolar lavage fluid increased IFN-γ and IL-10 levels in bronchoalveolar lavage fluid. Histological studies demonstrated that AST IV substantially inhibited OVA-induced eosinophilia in lung tissue. Flow cytometry studies demonstrated that AST IV substantially increased CD4(+)CD25(+)Foxp3 T cells (Treg). Furthermore quantitative real-time (qPCR) studies demonstrated that AST IV substantially enhanced Foxp3 mRNA expression in lung tissue. These findings suggest that AST IV may effectively ameliorate the progression of airway inflammation and could be used as a therapy for patients with allergic inflammation.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Drugs, Chinese Herbal; Eosinophilia; Female; Flow Cytometry; Forkhead Transcription Factors; Inflammation; Interferon-gamma; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Interleukin-4; Lung; Mice, Inbred BALB C; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; Saponins; T-Lymphocytes, Regulatory; Th1 Cells; Th2 Cells; Triterpenes

The IgE Fcε3 domain is an active immunotherapeutic target for asthma and other allergic diseases. However, previous methods for preparing IgE fusion protein vaccines are complex. Antigen 43 (Ag43) is a surface protein found in Escherichia coli that contains α and β subunits (the α subunit contains multiple T epitopes). Here we constructed a novel Ag43 surface display system (Ag43 system) to express Ag43 chimeric proteins to disrupt immune tolerance against IgE. The Ag43 system was constructed from the E. coli strain Tan109, in which the Ag43 gene was deleted and a recombinant plasmid (pETAg43) expressing a partial Ag43 gene was introduced. The Fcε3 domain of the IgE gene was then subcloned into plasmid pETAg43, resulting in a recombinant plasmid pETAg43/Fcε3, which was used to transform Tan109 for Ag43/Fcε3 surface expression. Thereafter, Ag43/Fcε3 was investigated as an asthma vaccine in a mouse model. Ag43/Fcε3 was expressed on and could be separated from the bacterial surface by heating to 60° while retaining activity. Ag43/Fcε3, as a protein vaccine, produced neutralizing autoantibodies to murine IgE, induced significant anti-asthma effects, and regulated IgE and T helper cytokines in a murine asthma model. Data show that Ag43/Fcε3 chimeric protein is a potential model vaccine for asthma treatment, and that the Ag43 system may be an effective tool for novel vaccine preparation to break immune tolerance to other self-molecules.

Topics: Adhesins, Escherichia coli; Adoptive Transfer; Animals; Antibodies, Neutralizing; Asthma; Autoantibodies; Bronchial Hyperreactivity; Bronchoconstriction; Cells, Cultured; Cloning, Molecular; Cytokines; Disease Models, Animal; Histamine; Immune Tolerance; Immunoglobulin E; Male; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Ovalbumin; Receptors, IgE; Recombinant Fusion Proteins; T-Lymphocytes, Helper-Inducer; Time Factors; Vaccines, Synthetic

Asthma is thought to result from the generation of T helper type 2 (Th2) responses, leading to bronchial inflammation. IFN-λ1 (also known as IL-29) is a recently described member of the IFN-λ family and has been shown to decrease production of Th2 cytokines in vitro. However, the role and mechanism of IFN-λ1 in asthma remain unknown.. The aim of this study was to clarify the importance of IFN-λ1 in allergen-induced airway hyperresponsiveness (AHR) and inflammation.. We used a murine model for ovalbumin (OVA)-induced asthma to examine the effect of intranasal delivery of recombinant adenovirus expressing human IFN-λ1 (Ad-hIFN-λ1) on AHR and allergic airway inflammation.. Intranasal instillation of Ad-hIFN-λ1 before airway antigen challenge in OVA-immunized mice significantly decreased the severity of AHR and numbers of eosinophils and levels of IL-4, IL-5, and IL-13, but not IL-10 and IFN-γ; both in vivo, in the bronchoalveolar lavage fluid and in vitro, following stimulation of lymphocytes from spleens with OVA, compared with administration of a control virus (Ad-mock). Furthermore, Ad-hIFN-λ1 treatment inhibited serum IgE secretion and increased numbers of splenic CD4(+)CD25(+)FOXP3(+) Treg cells. Histological studies showed that Ad-hIFN-λ1 attenuated OVA-induced lung tissue eosinophilia.. These results demonstrate that delivery of the Ad-hIFN-λ1 can mitigate allergic airway inflammation in experimental asthma. The potent immunoregulatory action of IFN-λ1 may offer a novel therapeutic approach to treat allergic asthma.

Topics: Adenoviridae; Administration, Intranasal; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Cytokines; Disease Progression; Humans; Immunoglobulin E; Interferons; Interleukins; Mice; Mice, Inbred BALB C; Models, Animal; Ovalbumin; Pneumonia; T-Lymphocytes, Regulatory

Allergic asthma is a chronic inflammatory airway disease arising from an aberrant immune response following exposure to environmental stimuli in genetically susceptible persons. The complement component 5 (C5)/C5a Receptor (C5aR/CD88) signaling pathway has been implicated in both experimental allergic asthma and human asthmatic disease. Targeting the C5a/C5aR signaling pathway in rodent models has been shown to either enhance or reduce allergic asthma consequences. Treatment with a recombinant humanized monoclonal antibody directed against C5 has shown unclear results in patients with asthma. The objective of this proof-of-concept animal study was to determine whether the low molecular weight C5aR peptidomimetic antagonist, PMX205, would reduce experimental allergic asthma consequences in mice. PMX205 or vehicle control was administered subcutaneously to BALB/c mice prior to and during standard ovalbumin (OVA) allergen sensitization and aerosolized challenge phases. PMX205 substantially reduced OVA-induced total cell (60%), neutrophil (66%) and eosinophil (65%) influxes in lavage fluid sampling. There were also significant reductions in OVA-induced lavage fluid IL-13 protein and lung Th2 cytokine gene expression with PMX205 administration. PMX205 treatment also diminished OVA-induced lung parenchyma cellular infiltration. PMX205 administration did not reduce OVA-induced serum IgE levels or epithelial mucous/goblet cell generation. There was no evidence of toxicity observed with PMX205 treatment in saline or OVA-challenged animals. These data provide evidence that pharmacologic blockade of C5aR by a low molecular weight antagonist (PMX205) reduces airway inflammatory cell and cytokine responses in experimental allergic asthma, and suggests that PMX205 might represent a novel therapeutic agent for reducing asthmatic outcomes.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophils; Epithelial Cells; Female; Goblet Cells; Immunoglobulin E; Inflammation; Interleukin-13; Lung; Male; Mice; Mice, Inbred BALB C; Mucus; Neutrophils; Ovalbumin; Peptides, Cyclic; Receptor, Anaphylatoxin C5a; Th2 Cells

This study is aimed to evaluate the effects of Psoraleae fructus (PF) on Th2 responses in a rat model of asthma in vivo and psoralen, a major constituent in PF, on Th2 responses in vitro. A rat model of asthma was established by sensitization and challenged with ovalbumin (OVA). Airway hyperresponsiveness was detected by direct airway resistance analysis. Lung tissues were examined for cell infiltration and mucus hypersecretion. Bronchoalveolar lavage fluid (BALF) was assessed for cytokine levels. In vitro study, Th2 cytokine production was evaluated in the culture supernatant of D10.G4.1 (D10 cells) followed by the determination of cell viability, meanwhile Th2 transcription factor GATA-3 expression in D10 cells was also determined. The oral administration of PF significantly reduced airway hyperresponsiveness (AHR) to aerosolized methacholine and decreased IL-4 and IL-13 levels in the BALF. Histological studies showed that PF markedly inhibited inflammatory infiltration and mucus secretion in the lung tissues. In vitro study, psoralen significantly suppressed Th2 cytokines of IL-4, IL-5 and IL-13 by ConA-stimulated D10 cells without inhibitory effect on cell viability. Furthermore, GATA-3 protein expression was also markedly reduced by psoralen. This study demonstrated that PF exhibited inhibitory effects on hyperresponsiveness and airway inflammation in a rat model of asthma, which was associated with the suppression of Th2 response. Psoralen, a major constituent of PF, has immunomodulatory properties on Th2 response in vitro, which indicated that psoralen might be a critical component of PF for its therapeutic effects.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Ficusin; Inflammation Mediators; Male; Ovalbumin; Phytotherapy; Psoralea; Rats; Rats, Sprague-Dawley; Specific Pathogen-Free Organisms; Th2 Cells

Observational as well as experimental studies support that prenatal challenges seemed to be associated with an increased risk for allergic airway diseases in the offspring. However, insights into biomarkers involved in mediating this risk are largely elusive. We here aimed to test the association between endogenous and exogenous factors documented in pregnant women, including psychosocial, endocrine, and life style parameters, and the risk for allergic airway diseases in the children later in life. We further pursued to functionally test identified factors in a mouse model of an allergic airway response. In a prospectively designed pregnancy cohort (n = 409 families), women were recruited between the 4th and 12th week of pregnancy. To investigate an association between exposures during pregnancy and the incidence of allergic airway disease in children between 3 and 5 years of age, multiple logistic regression analyses were applied. Further, in prenatally stressed adult offspring of BALB/c-mated BALB/c female mice, asthma was experimentally induced by ovalbumin (OVA) sensitization. In addition to the prenatal stress challenge, some pregnant females were treated with the progesterone derivative dihydrodydrogesterone (DHD). In humans, we observed that high levels of maternal progesterone in early human pregnancies were associated with a decreased risk for an allergic airway disease (asthma or allergic rhinitis) in daughters (adjusted OR 0.92; 95% confidence interval [CI] 0.84 to 1.00) but not sons (aOR 1.02, 95% CI 0.94-1.10). In mice, prenatal DHD supplementation of stress-challenged dams attenuated prenatal stress-induced airway hyperresponsiveness exclusively in female offspring. Reduced levels of maternal progesterone during pregnancy-which can result from high stress perception-increase the risk for allergic airway diseases in females but not in males. Key messages: Lower maternal progesterone during pregnancy increases the risk for allergic airway disease only in female offspring. Prenatal progesterone supplementation ameliorates airway hyperreactivity in prenatally stressed murine offspring.

Topics: Adult; Allergens; Animals; Bronchial Hyperreactivity; Child, Preschool; Cytokines; Disease Models, Animal; Female; Germany; Humans; Infant; Infant, Newborn; Male; Mice, Inbred BALB C; Odds Ratio; Ovalbumin; Pregnancy; Progesterone; Prospective Studies; Respiratory Hypersensitivity; Risk; Sex Factors; Stress, Psychological

Epidemiologic studies suggest that increased concentrations of airborne spores of Aspergillus fumigatus closely relate to asthma aggravation. Chronic exposure to A. fumigatus aggravates airway inflammation, remodeling, and airway hyperresponsiveness in asthmatic rats. The effects of chronic exposure to A. fumigatus on epidermal growth factor receptor (EGFR) expression in the airway epithelial cells of asthmatic rats remain unclear. This study aimed to investigate the effects of chronic exposure to A. fumigatus on injury and shedding of airway epithelium, goblet cell metaplasia, and EGFR expression in the airway epithelial cells of asthmatic rats. A rat model of chronic asthma was established using ovalbumin (OVA) sensitization and challenge. Rats with chronic asthma were then exposed to long-term inhalation of spores of A. fumigatus, and the dynamic changes in injury and shedding of airway epithelium, goblet cell metaplasia, and EGFR expression were observed and analyzed. Chronic exposure to A. fumigatus could aggravate airway epithelial cell damage, upregulate the expression of EGFR and its ligands EGF and TGF-α, promote goblet cell metaplasia, and increase airway responsiveness in rats with asthma. Chronic exposure to A. fumigatus upregulates the expression of EGFR and its ligands in asthmatic rats. The EGFR pathway may play a role in asthma aggravation induced by exposure to A. fumigatus.

Topics: Animals; Aspergillosis; Aspergillus fumigatus; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Epidermal Growth Factor; Epithelial Cells; ErbB Receptors; Goblet Cells; Male; Metaplasia; Ovalbumin; Pneumonia; Rats; Rats, Wistar

Linker for activation of T cells (LAT) is a key adaptor in the T cell receptor (TCR) signaling pathway. The expression of LAT is lower in asthmatic patients than that in healthy people, but there is little knowledge about the mechanism underlying this phenomenon. This study was aimed to determine whether LAT-PLC-γ1 interaction was involved in the development of asthma. It was shown that the phosphorylation of PLC-γ1 decreased in the asthmatic mouse model and Th2 cell differentiated CD4(+) T cells. In addition, depleted endogenous PLC-γ1 promoted CD4(+) T cells to differentiate into IL-4-Productor. It was therefore concluded that the low level of LAT-PLC-γ1 interaction was associated with Th2 polarized differentiation, and this may contribute to the etiology of asthma.

Topics: Adaptor Proteins, Signal Transducing; Animals; Asthma; Bronchial Hyperreactivity; Cell Differentiation; Cells, Cultured; Female; Interleukin-4; Lymphocyte Activation; Membrane Proteins; Mice; Mice, Inbred BALB C; Ovalbumin; Phospholipase C gamma; Phosphoproteins; Phosphorylation; Signal Transduction; Th2 Cells

Periostin, a secreted extracellular matrix protein, has been localized to deposits of subepithelial fibrosis in asthmatic patients, and periostin levels have been linked to increases in IL-13.. We hypothesized that periostin is required for airway inflammatory responses to a physiologic aeroallergen, house dust mite (HDM).. We studied F4-F6 B6;129-Postn(tm1Jmol)/J wild-type (Postn(+/+)) and null (Postn(-/-)) mice, as well as C57BL/6 mice treated with either IgM or OC-20 periostin neutralizing antibody. Mice were exposed to 5 doses of HDM intranasally over a 16-day period.. HDM increased airways responsiveness in Postn(+/+) but not Postn(-/-) mice. In addition, HDM-treated C57BL/6 mice injected with OC-20 had lower airways responsiveness than HDM-treated mice injected with IgM. Compared with Postn(+/+) mice, Postn(-/-) mice showed decreases in HDM-induced inflammation and mucous metaplasia, as well as reduced IL-4, IL-25, CD68, Gob5, and periostin mRNA expression. OC-20 antibody produced similar results. HDM exposure increased periostin expression in the airway epithelium, subepithelium, smooth muscle and inflammatory cells. OC-20 blocked the HDM-induced IgE response, and T cells incubated with dendritic cells (DCs) from Postn(-/-) mice or treated with OC-20 showed deficient DNA synthesis and IL-13 responses compared with T cells incubated with wild-type DCs. Finally, adoptive transfer of bone marrow-derived DCs from Postn(+/+) mice was sufficient to promote allergic responses in F6 Postn(-/-) littermates.. In mice, periostin is required for maximal airways hyperresponsiveness and inflammation after HDM sensitization and challenge. Periostin is required for maximal HDM-induced T-cell responses.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cell Adhesion Molecules; Complex Mixtures; Dendritic Cells; Dermatophagoides pteronyssinus; Immunoglobulin E; Lung; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Pneumonia; Respiratory Hypersensitivity; RNA, Messenger; T-Lymphocytes

Mycoplasma pneumoniae causes a range of airway and extrapulmonary pathologies in humans. Clinically, M. pneumoniae is associated with acute exacerbations of human asthma and a worsening of experimentally induced asthma in mice. Recently, we demonstrated that Community Acquired Respiratory Distress Syndrome (CARDS) toxin, an ADP-ribosylating and vacuolating toxin synthesized by M. pneumoniae, is sufficient to induce an asthma-like disease in BALB/cJ mice. To test the potential of CARDS toxin to exacerbate preexisting asthma, we examined inflammatory responses to recombinant CARDS toxin in an ovalbumin (OVA) murine model of asthma. Differences in pulmonary inflammatory responses between treatment groups were analyzed by histology, cell differentials and changes in cytokine and chemokine concentrations. Additionally, assessments of airway hyperreactivity were evaluated through direct pulmonary function measurements. Analysis of histology revealed exaggerated cellular inflammation with a strong eosinophilic component in the CARDS toxin-treated group. Heightened T-helper type-2 inflammatory responses were evidenced by increased expression of IL-4, IL-13, CCL17 and CCL22 corresponding with increased airway hyperreactivity in the CARDS toxin-treated mice. These data demonstrate that CARDS toxin can be a causal factor in the worsening of experimental allergic asthma, highlighting the potential importance of CARDS toxin in the etiology and exacerbation of human asthma.

Topics: Animals; Asthma; Bacterial Proteins; Bacterial Toxins; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL17; Chemokine CCL22; Eosinophils; Humans; Inflammation; Interleukin-13; Interleukin-4; Mice; Mice, Inbred BALB C; Ovalbumin; Recombinant Proteins; Respiratory System; Th2 Cells

It is well-established that bacterial and viral infections have an exacerbating effect on allergic asthma, particularly aggravating respiratory symptoms, such as airway hyperresponsiveness (AHR). The mechanism by which these infections alter AHR is unclear, but some studies suggest that Toll-like receptors (TLRs) play a role. In this study, we investigated the impact of TLR3 and TLR4 ligands on AHR and airway inflammation in a model of pre-established allergic inflammation. Female BALB/c mice were sensitised and challenged intranasally (i.n.) with either PBS or ovalbumin (OVA) and subsequently i.n. challenged with poly (I:C) (TLR3) or LPS (TLR4) for four consecutive days. The response to methacholine was measured in vivo; cellular and inflammatory mediators were measured in blood, lung tissue and broncheoalveolar lavage fluid (BALF). OVA challenge resulted in an increase in AHR to methacholine, as well as increased airway eosinophilia and TH2 cytokine production. Subsequent challenge with TLR agonists resulted in a significant increase in AHR, but decreased TLR-specific cellular inflammation and production of immune mediators. Particularly evident was a decline in LPS-induced neutrophilia and neutrophil-associated cytokines following LPS and poly (I:C) treatment. The present data indicates that TLRs may play a pivotal role in AHR in response to microbial infection in allergic lung inflammation. These data also demonstrate that aggravated AHR occurs in the absence of an exacerbation in airway inflammation and that allergic inflammation impedes a subsequent inflammatory response to TLRs. These results may parallel clinical signs of microbial asthma exacerbation, including an extended duration of illness and increased respiratory symptoms.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Female; Inflammation; Ligands; Mice; Ovalbumin; Poly I-C; Respiratory System; Toll-Like Receptor 3; Toll-Like Receptor 4

Bronchial asthma is characterized by chronic lung inflammation, airway hyperresponsiveness, and airway remodelling. Astragaloside IV (3-O-β-D-xylopyranosyl-6-O-β-D-glucopyranosyl-cycloastragenol, AST), the primary pure saponin isolated from the root of Astragalus membranaceus, is an effective compound with distinct pharmacological effects including anti-inflammation, immunoregulation, and antifibrosis. However, the effect of AST on asthma remains unclear. In the present study, in the murine model of asthma, the airway hyperresponsiveness was relieved after treatment with AST, accompanied by a reduction of inflammatory cells. In addition, the levels of IL-4 and IL-5 decreased, while the IFN-γ level increased, in bronchoalveolar lavage fluid. The compound also significantly inhibited the synthesis of GATA-3-encoding mRNA and protein in addition to increasing the synthesis of T-bet-encoding mRNA and protein in both lung tissues and CD4+ T cells. Our findings indicate that AST treatment inhibits ovalbumin-induced airway inflammation by modulating the key master switches GATA-3 and T-bet, which results in committing T helper cells to a Th1 phenotype.

Topics: Animals; Asthma; Astragalus propinquus; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; GATA3 Transcription Factor; Interferon-gamma; Interleukin-4; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; RNA, Messenger; Saponins; Triterpenes

Asthma develops from injury to the airways/lungs, stemming from airway inflammation (AI) and airway remodeling (AWR), both contributing to airway hyperresponsiveness (AHR). Airway epithelial damage has been identified as a new etiology of asthma but is not targeted by current treatments. Furthermore, it is poorly studied in currently used animal models of AI and AWR. Therefore, this study aimed to incorporate epithelial damage/repair with the well-established ovalbumin (OVA)-induced model of chronic allergic airway disease (AAD), which presents with AI, AWR, and AHR, mimicking several features of human asthma. A 3-day naphthalene (NA)-induced model of epithelial damage/repair was superimposed onto the 9-week OVA-induced model of chronic AAD, before 6 weeks of OVA nebulization (NA+OVA group), during the second last OVA nebulization period (OVA/NA group) or 1 day after the 6-week OVA nebulization period (OVA+NA group), using 6-8-week-old female Balb/c mice (n=6-12/group). Mice subjected to the 9-week OVA model, 3-day NA model or respective vehicle treatments (saline and corn oil) were used as appropriate controls. OVA alone significantly increased epithelial thickness and apoptosis, goblet cell metaplasia, TGF-β1, subepithelial collagen (assessed by morphometric analyses of various histological stains), total lung collagen (hydroxyproline analysis), and AHR (invasive plethysmography) compared with that in saline-treated mice (all P<0.05 vs saline treatment). NA alone caused a significant increase in epithelial denudation and apoptosis, TGF-β1, subepithelial, and total lung collagen compared with respective measurements from corn oil-treated controls (all P<0.01 vs corn oil treatment). All three combined models underwent varying degrees of epithelial damage and AWR, with the OVA+NA model demonstrating the greatest increase in subepithelial/total lung collagen and AHR (all P<0.05 vs OVA alone or NA alone). These combined models of airway epithelial damage/AAD demonstrated that epithelial damage is a key contributor to AWR, fibrosis and related AHR, and augments the effects of AI on these parameters.

Topics: Animals; Apoptosis; Asthma; Bronchi; Bronchial Hyperreactivity; Collagen; Disease Models, Animal; Epithelium; Female; Fibrosis; Lung; Metaplasia; Mice; Mice, Inbred BALB C; Naphthalenes; Ovalbumin

Phospholipase Cε (PLCε) is an effector of Ras and Rap small GTPases and expressed in non-immune cells. It is well established that PLCε plays an important role in skin inflammation, such as that elicited by phorbol ester painting or ultraviolet irradiation and contact dermatitis that is mediated by T helper (Th) 1 cells, through upregulating inflammatory cytokine production by keratinocytes and dermal fibroblasts. However, little is known about whether PLCε is involved in regulation of inflammation in the respiratory system, such as Th2-cells-mediated allergic asthma.. We prepared a mouse model of allergic asthma using PLCε+/+ mice and PLCεΔX/ΔX mutant mice in which PLCε was catalytically-inactive. Mice with different PLCε genotypes were immunized with ovalbumin (OVA) followed by the challenge with an OVA-containing aerosol to induce asthmatic response, which was assessed by analyzing airway hyper-responsiveness, bronchoalveolar lavage fluids, inflammatory cytokine levels, and OVA-specific immunoglobulin (Ig) levels. Effects of PLCε genotype on cytokine production were also examined with primary-cultured bronchial epithelial cells.. After OVA challenge, the OVA-immunized PLCεΔX/ΔX mice exhibited substantially attenuated airway hyper-responsiveness and broncial inflammation, which were accompanied by reduced Th2 cytokine content in the bronchoalveolar lavage fluids. In contrast, the serum levels of OVA-specific IgGs and IgE were not affected by the PLCε genotype, suggesting that sensitization was PLCε-independent. In the challenged mice, PLCε deficiency reduced proinflammatory cytokine production in the bronchial epithelial cells. Primary-cultured bronchial epithelial cells prepared from PLCεΔX/ΔX mice showed attenuated pro-inflammatory cytokine production when stimulated with tumor necrosis factor-α, suggesting that reduced cytokine production in PLCεΔX/ΔX mice was due to cell-autonomous effect of PLCε deficiency.. PLCε plays an important role in the pathogenesis of bronchial asthma through upregulating inflammatory cytokine production by the bronchial epithelial cells.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Epithelial Cells; Female; Gene Expression Regulation; Immunoglobulin E; Immunoglobulin G; Male; Mice; Mice, Knockout; Ovalbumin; Phosphoinositide Phospholipase C; Primary Cell Culture; rap GTP-Binding Proteins; ras GTPase-Activating Proteins; Respiratory Mucosa; Signal Transduction; Th1-Th2 Balance; Th2 Cells; Tumor Necrosis Factor-alpha

Allergic asthma is a complex disease with a strong genetic component where mast cells play a major role by the release of proinflammatory mediators. In the mouse, mast cell protease-6 (mMCP-6) closely resembles the human version of mast cell tryptase, β-tryptase. The gene that encodes mMCP-6, Tpsb2, resides close by the H-2 complex (MHC gene) on chromosome 17. Thus, when the original mMCP-6 knockout mice were backcrossed to the BALB/c strain, these mice were carrying the 129/Sv haplotype of MHC (mMCP-6(-/-)/H-2bc). Further backcrossing yielded mMCP-6(-/-) mice with the BALB/c MHC locus. BALB/c mice were compared with mMCP-6(-/-) and mMCP-6(-/-)/H-2bc mice in a mouse model of experimental asthma. Although OVA-sensitized and challenged wild type mice displayed a striking airway hyperresponsiveness (AHR), mMCP-6(-/-) mice had less AHR that was comparable with that of mMCP-6(-/-)/H-2bc mice, suggesting that mMCP-6 is required for a full-blown AHR. The mMCP-6(-/-)/H-2bc mice had strikingly reduced lung inflammation, IgE responses, and Th2 cell responses upon sensitization and challenge, whereas the mMCP-6(-/-) mice responded similarly to the wild type mice but with a minor decrease in bronchoalveolar lavage eosinophils. These findings suggest that inflammatory Th2 responses are highly dependent on the MHC-haplotype and that they can develop essentially independently of mMCP-6, whereas mMCP-6 plays a key role in the development of AHR.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chromosomes, Mammalian; Crosses, Genetic; Eosinophils; Female; Gene Expression Regulation; H-2 Antigens; Haplotypes; Immunoglobulin E; Major Histocompatibility Complex; Male; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Signal Transduction; Th2 Cells; Tryptases

CD86-CD28 interaction has been suggested as the principal costimulatory pathway for the activation and differentiation of naïve T cells in allergic inflammation. However, it remains uncertain whether this pathway also has an essential role in the effector phase. We sought to determine the contribution of CD86 on dendritic cells in the reactivation of allergen-specific Th2 cells.. We investigated the effects of the downregulation of CD86 by short interfering RNAs (siRNAs) on Th2 cytokine production in the effector phase in vitro and on asthma phenotypes in ovalbumin (OVA)-sensitized and -challenged mice.. Treatment of bone marrow-derived dendritic cells (BMDCs) with CD86 siRNA attenuated LPS-induced upregulation of CD86. CD86 siRNA treatment impaired BMDCs' ability to activate OVA-specific Th2 cells. Intratracheal administration of CD86 siRNA during OVA challenge downregulated CD86 expression in the airway mucosa. CD86 siRNA treatment ameliorated OVA-induced airway eosinophilia, airway hyperresponsiveness, and the elevations of OVA-specific IgE in the sera and IL-5, IL-13, and CCL17 in the bronchoalveolar lavage fluid, but not the goblet cell hyperplasia.. These results suggest that local administration of CD86 siRNA during the effector phase ameliorates lines of asthma phenotypes. Targeting airway dendritic cells with siRNA suppresses airway inflammation and hyperresponsiveness in an experimental model of allergic asthma.

Topics: Animals; Asthma; B7-2 Antigen; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cells, Cultured; Coculture Techniques; Cytokines; Dendritic Cells; Disease Models, Animal; Immunoglobulin E; Lung; Lymphocyte Activation; Mice, Inbred BALB C; Ovalbumin; Phenotype; Pulmonary Eosinophilia; RNA Interference; RNAi Therapeutics; Th2 Cells; Transfection

Epidemiological studies have revealed that intrauterine growth retardation (IUGR) or low birth weight is linked to the later development of asthma. Epigenetic regulatory mechanisms play an important role in the fetal origins of adult disease. However, little is known regarding the correlation between epigenetic regulation and the development of asthma following IUGR.. An IUGR and ovalbumin (OVA)-sensitization/challenge rat model was used to study whether epigenetic mechanisms play a role in the development of asthma following IUGR.. Maternal nutrient restriction increased histone acetylation levels of the endothelin-1 (ET-1) gene promoter in lung tissue of offspring, but did not cause significant alterations of DNA methylation. The effect was maintained until 10 weeks after birth. Furthermore, these epigenetic changes may have induced IUGR individuals to be highly sensitive to OVA challenge later in life, resulting in more significant changes related to asthma.. These findings suggest that epigenetic mechanisms might be closely associated with the development of asthma following IUGR, providing further insight for improved prevention of asthma induced by environmental factors.

Topics: Acetylation; Age Factors; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; DNA Methylation; Endothelin-1; Epigenesis, Genetic; Female; Fetal Growth Retardation; Gene Expression Regulation; Genetic Predisposition to Disease; Histones; Maternal Nutritional Physiological Phenomena; Nutritional Status; Ovalbumin; Pregnancy; Promoter Regions, Genetic; Rats, Sprague-Dawley; Risk Factors

Atopic march refers to the typical transition from a food allergy in early childhood to allergic asthma in older children and adults. However the precise interplay of events involving gut, skin and pulmonary inflammation in this process is not completely understood.. To develop a mouse model of mixed food and respiratory allergy mimicking the atopic march and better understand the impact of food allergies on asthma.. Food allergy to ovalbumin (OVA) was induced through intra-peritoneal sensitization and intra-gastric challenge, and/or a respiratory allergy to house dust mite (HDM) was obtained through percutaneous sensitization and intra-nasal challenges with dermatophagoides farinae (Der f) extract. Digestive, respiratory and systemic parameters were analyzed.. OVA-mediated gut allergy was associated with an increase in jejunum permeability, and a worsening of Der f-induced asthma with stronger airway hyperresponsiveness and pulmonary cell infiltration, notably eosinophils. There was overproduction of the pro-eosinophil chemokine RANTES in broncho-alveolar lavages associated with an enhanced Th2 cytokine secretion and increased total and Der f-specific IgE when the two allergies were present. Both AHR and lung inflammation increased after a second pulmonary challenge.. Gut sensitization to OVA amplifies Der f-induced asthma in mice.

Topics: Animals; Antigens, Dermatophagoides; Arthropod Proteins; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Chemokine CCL5; Disease Models, Animal; Female; Food Hypersensitivity; Immunoglobulin E; Intestinal Mucosa; Intestines; Lung; Mice, Inbred BALB C; Ovalbumin; Permeability; Pneumonia; Pulmonary Eosinophilia; Th2 Cells; Time Factors

In recent studies, SPA0355, a thiourea analog, has been demonstrated to possess strong anti-inflammatory activity. However, the mechanisms underlying the effects of SPA0355 on immune-mediated diseases have not been fully defined. The present study was designed to investigate the immunological and molecular mechanisms by which SPA0355 modulates cluster of differentiation of (CD4)(+) T-cell-mediated immune responses in allergic airway inflammation. In vitro studies have shown that SPA0355 suppresses CD4(+) T-cell activation, proliferation, and differentiation via modulation of T-cell receptor (TCR) signal transduction and cytokine-induced Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling. Next, we investigated the efficacy of SPA0355 in ovalbumin (OVA)-induced allergic airway inflammation. Intraperitoneal administration of SPA0355 inhibited inflammatory cell recruitment into the airways as well as the production of Th2 cytokines in bronchoalveolar fluid and suppressed OVA-induced IgE production in serum. Additionally, SPA0355 suppressed mucin production and smooth muscle hypertrophy and prevented the development of airway hyperresponsiveness. Given that allergic airway inflammation is mainly driven by Th2 cell responses, it is highly possible that the defects in CD4(+) T-cell activation and Th2 cell differentiation in the draining lymph nodes and suppressed NF-κB activation in the lungs of SPA0355-treated mice illustrate an immunological mechanism of the preventive effect of SPA0355 on the aforementioned asthmatic characteristics. Collectively, our results suggest that SPA0355 directly modulates Th1 and Th2 responses through the suppression of multiple signaling pathways triggered by TCR or cytokine receptor stimulation, and that SPA0355 has protective effects in a murine model of allergic airway inflammation.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Benzoxazines; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Chemotaxis, Leukocyte; Cytokines; Disease Models, Animal; Immunosuppressive Agents; In Vitro Techniques; Lymphocyte Activation; Male; Mice; Mice, Inbred BALB C; Mucins; NF-kappa B; Ovalbumin; Receptors, Antigen, T-Cell; Signal Transduction; Thiourea

This study examines the immunological responses in rats following inhalation to titanium dioxide nanoparticles (TiO2 NPs), in naïve rats and in rats with induced allergic airway disease. The responses of two different inbred rat strains were compared: the Dark Aguoti (DA), susceptible to chronic inflammatory disorders, and the Brown Norwegian (BN), susceptible to atopic allergic inflammation. Naïve rats were exposed to an aerosol of TiO2 NPs once daily for 10 days. Another subset of rats was sensitized to the allergen ovalbumin (OVA) in order to induce airway inflammation. These sensitized rats were exposed to TiO2 NPs before and during the allergen challenge. Naïve rats exposed to TiO2 NPs developed an increase of neutrophils and lymphocytes in both rat strains. Airway hyperreactivity and production of inflammatory mediators typical of a T helper 1 type immune response were significantly increased, only in DA rats. Sensitization of the rats induced a prominent OVA-specific-IgE and IgG response in the BN rat while DA rats only showed an increased IgG response. Sensitized rats of both strains developed airway eosinophilia following allergen challenge, which declined upon exposure to TiO2 NPs. The level of neutrophils and lymphocytes increased upon exposure to TiO2 NPs in the airways of DA rats but remained unchanged in the airways of BN rats. In conclusion, the responses to TiO2 NPs were strain-dependent, indicating that genetics play a role in both immune and airway reactivity. DA rats were found to be higher responder compared to BN rats, both when it comes to responses in naïve and sensitized rats. The impact of genetically determined factors influencing the inflammatory reactions pinpoints the complexity of assessing health risks associated with nanoparticle exposures.

Topics: Aerosols; Animals; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Genetic Variation; Genotype; Immunoglobulin E; Immunoglobulin G; Inflammation Mediators; Inhalation Exposure; Lung; Male; Metal Nanoparticles; Neutrophils; Ovalbumin; Phenotype; Pneumonia; Pulmonary Eosinophilia; Rats, Inbred BN; Risk Factors; Species Specificity; Th1 Cells; Titanium

IL-4 and IL-13 comprise promising targets for therapeutic interventions in asthma and other Th2-associated diseases, but agents targeting either IL-4 or IL-13 alone have shown limited efficacy in human clinical studies. Because these cytokines may involve redundant function, dual targeting holds promise for achieving greater efficacy. We describe a bifunctional therapeutic targeting IL-4 and IL-13, developed by a combination of specific binding domains. IL-4-targeted and IL-13-targeted single chain variable fragments were joined in an optimal configuration, using appropriate linker regions on a novel protein scaffold. The bifunctional IL-4/IL-13 antagonist displayed high affinity for both cytokines. It was a potent and efficient neutralizer of both murine IL-4 and murine IL-13 bioactivity in cytokine-responsive Ba/F3 cells, and exhibited a half-life of approximately 4.7 days in mice. In a murine model of ovalbumin-induced ear swelling, the bifunctional molecule blocked both the IL-4/IL-13-dependent early-phase response and the IL-4-dependent late-phase response. In the ovalbumin-induced lung inflammation model, the bifunctional IL-4/IL-13 antagonist reduced the IL-4-dependent rise in serum IgE titers, and reduced IL-13-dependent airway hyperresponsiveness, lung inflammation, mucin gene expression, and serum chitinase responses. Taken together, these findings demonstrate the effective dual blockade of IL-4 and IL-13 with a single agent, which resulted in the modulation of a more extensive range of endpoints than could be achieved by targeting either cytokine alone.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Binding Sites; Bronchial Hyperreactivity; CHO Cells; Cricetinae; Cross-Linking Reagents; Ear; Female; Half-Life; Immunoglobulin E; Interleukin-13; Interleukin-13 Receptor alpha2 Subunit; Interleukin-4; Mice; Mice, Inbred C57BL; Molecular Conformation; Neutralization Tests; Ovalbumin; Pneumonia; Protein Binding; Protein Interaction Domains and Motifs; Single-Chain Antibodies

Recent studies have suggested that upper airway inflammation has a strong impact on lower airway diseases. The purpose of this study was to assess whether nasal inflammation could exacerbate allergic asthma in a mouse model.. Mice were assigned to 4 groups: control (Cont), either rhinosinusitis (R) or allergic asthma (A) alone, and both rhinosinusitis and allergic asthma (R&A). Mice underwent induction of nasal inflammation (R and R&A) or sham surgery (Cont and A) on day 1. Mice in the A and R&A groups were sensitized to ovalbumin on days 1, 7, and 14, followed by aerosol challenge on days 18 to 20, whereas in the Cont and R groups only saline was administered. All mice were assessed for airway hyperresponsiveness (AHR) and were euthanized on day 21. The sera, bronchoalveolar lavage fluids (BALFs), and nasal and lung tissues were collected for further analyses.. Histology findings confirmed upper and lower airway inflammation in experimental mice. Significantly increased AHR and total serum immunoglobulin E (IgE) were observed in the R&A group when compared with those of the Cont, R, and A groups. Responses to IgG2a induction were also found in sera and BALFs from mice with rhinosinusitis (R and R&A). Higher levels of interleukin 4 (IL-4) and IL-13, and increased eosinophilic inflammation were detected in BALFs and lung tissues from the experimental groups when compared with those from the Cont group.. Our results confirm that upper airway inflammation could exacerbate allergic asthma, and provide support to the concept of "one airway, one disease.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Immunoglobulin E; Immunoglobulin G; Interleukin-13; Interleukin-4; Leukocyte Count; Lung; Male; Mice; Mice, Inbred BALB C; Nasal Cavity; Ovalbumin; Rhinitis; Sinusitis

The present study aimed to determine the protective effects and the underlying mechanisms of curcumin on ovalbumin (OVA)-induced allergic inflammation in a mouse model of allergic asthma. Asthma mice model was established by ovalbumin. A total of 60 mice were randomly assigned to six experimental groups: control, model, dexamethasone (2 mg/kg), and curcumin (50 mg/kg, 100 mg/kg, 200 mg/kg). Airway resistance (Raw) was measured by the forced oscillation technique, differential cell count in BAL fluid (BALF) was measured by Wright-Giemsa staining, histological assessment was measured by hematoxylin and eosin (HE) staining, BALF levels of Treg/Th17 cytokines were measured by enzyme-linked immunosorbent assay, Treg cells and Th17 cells were evaluated by flow cytometry (FCM). Our study demonstrated that curcumin inhibited OVA-induced increases in eosinophil count; interleukin (IL)-17A level were recovered in bronchoalveolar lavage fluid increased IL-10 level in bronchoalveolar lavage fluid. Histological studies demonstrated that curcumin substantially inhibited OVA-induced eosinophilia in lung tissue. Flow cytometry (FCM) studies demonstrated that curcumin remarkably inhibited Th17 cells and significantly increased Treg cells. The results in vivo show ovalbumin-induced significantly broke Treg/Th17 balance; curcumin treatments markedly attenuated the inflammatory in asthma model by regulating Treg/Th17 balance. Our findings support the possible use of curcumin as a therapeutic drug for patients with allergic asthma.

Topics: Airway Resistance; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4 Antigens; Curcuma; Curcumin; Eosinophilia; Eosinophils; Female; Inflammation; Interleukin-10; Interleukin-17; Interleukin-2 Receptor alpha Subunit; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plant Extracts; T-Lymphocytes, Regulatory; Th17 Cells

Several studies suggest that the N/OFQ (nociceptin/orphanin FQ)-NOP (N/OFQ peptide) receptor pathway is involved in airway physiology. We previously demonstrated a modulation of the endogenous N/OFQ levels in allergen-sensitized mice. Here, we investigated the effects of NOP receptor activation in allergen sensitization using a murine model of allergen-induced airway hyperresponsiveness (AHR). BALB/c mice were intraperitoneally treated with the NOP receptor agonist UFP-112, either during the sensitization phase (30 min before ovalbumin administration) or at the end of sensitization process (15 min before bronchopulmonary reactivity evaluation). At day 21 from the first allergen exposure, bronchopulmonary reactivity and total and differential cell count in bronchoalveolar lavage fluid were evaluated. In a separate set of experiments cell proliferation in lymphocytes, cytokine levels, IgE serum levels, and the effect of UFP-112 on IL-13-induced AHR were evaluated. Pretreatment with UFP-112, during the sensitization phase, caused a significant reduction in allergen-induced AHR and total cell lung infiltration. No effect on allergen-induced AHR was observed when the treatment was performed at the end of sensitization process, on tissues harvested from OVA-sensitized mice and on IL-13-induced AHR. The in vitro proliferative response of lymphocytes was significantly reduced by pretreatment during the sensitization phase with UFP-112. This effect was paralleled by a significant modulation of cytokine secretion in pulmonary tissues and lymphocytes. In conclusion, we demonstrated a role for the NOP receptor and N/OFQ pathway in the AHR induced by allergen, probably through a modulation of the immune response that triggers the development of AHR that involves pro- and anti-inflammatory cytokines.

Topics: Allergens; Animals; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Proliferation; Female; Immunoglobulin E; Interleukin-13; Lung; Lymphocytes; Mice; Mice, Inbred BALB C; Nociceptin; Nociceptin Receptor; Opioid Peptides; Ovalbumin; Receptors, Opioid

Hygiene hypothesis demonstrates that the lack of microbial exposure would promote the development of allergic airway disease (AAD). Therefore, the gut microbiota, including Escherichia coli (E. coli), would probably offer a potential strategy for AAD.. To investigate whether E. coli infection is able to suppress the induction of AAD and to elucidate the underlying mechanisms.. Nonpathogenic E. coli ATCC 25922 was infected by gavage before AAD phase in three patterns: 10(8) or 10(6) CFU in neonates or 10(8) CFU in adults. Then mice were sensitized and challenged with ovalbumin (OVA) to induce allergic inflammation in both the upper and lower airways. Hallmarks of AAD, in terms of eosinophil infiltration and goblet cell metaplasia in subepithelial mucosa, Th2 skewing of the immune response, and levels of T regulate cells (Tregs), were examined by histological analysis, ELISA, and flow cytometry, respectively.. E. coli, especially neonatally infected with an optimal dose, attenuated allergic responses, including a decrease in nasal rubbing and sneezing, a reduction in eosinophil inflammation and goblet cell metaplasia in subepithelial mucosa, decreased serum levels of OVA-specific IgE, and reduced Th2 (IL-4) cytokines. In contrast, this effect came with an increase of Th1 (IFN-r and IL-2) cytokines, and an enhancement of IL-10-secreting Tregs in paratracheal lymph nodes (PTLN).. E. coli suppresses allergic responses in mice, probably via a shift from Th1 to Th2 and/or induction of Tregs. Moreover, this infection is age- and dose-dependent, which may open up novel possibilities for new therapeutic interventions.

Topics: Animals; Antibody Specificity; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Escherichia coli; Female; Immunoglobulin E; Immunomodulation; Inflammation; Lymph Nodes; Mice; Ovalbumin; T-Lymphocytes, Regulatory

Chemoattractant receptor homologous molecule expressed on T helper type 2 cells (CRTH2) is a PGD2 receptor found on eosinophils, basophils, and Th2 type T cells which exhibits chemotaxis and functions in activation cascades. However, while a number of CRTH2 antagonists, including ramatroban, are known to exert activity in certain animal models, activity in a guinea pig model of EA-induced airway hyperresponsiveness has not been demonstrated. The newly developed CRTH2 antagonist ASP5642 has shown antagonistic activity against human and guinea pig CRTH2 in previous studies and has also been found effective in treating guinea pig models of airway inflammation and airway hyperresponsiveness. While previous studies have used animals such as rats and mice to evaluate CRTH2 antagonist effects, ours is the first attempt to evaluate CRTH2 function in a guinea pig asthma model, which may prove useful in evaluating the compound's effects in humans, given the comparable airway function between the two species taken together, these data from the present study strongly suggest the utility of ASP5642 in investigating the role of CRTH2 in inflammatory responses and as a drug treatment for human asthma.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Antigens; Benzhydryl Compounds; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carbazoles; Cell Count; Eosinophilia; Guinea Pigs; HEK293 Cells; Humans; K562 Cells; Male; Ovalbumin; Pneumonia; Prostaglandin D2; Pyridazines; Receptors, Immunologic; Receptors, Prostaglandin; Sulfonamides

The inducible T cell kinase (ITK) regulates type 2 (Th2) cytokines that provide defense against certain parasitic and bacterial infections and are involved in the pathogenesis of lung inflammation such as allergic asthma. Activation of ITK requires the interaction of its SH3 domain with the poly-proline region of its signaling partner, the SH2 domain containing leukocyte phosphoprotein of 76 kilodaltons (SLP-76). The specific disruption of the ITK-SH3/SLP-76 poly-proline interaction in vitro by a cell-permeable competitive inhibitor peptide (R9-QQP) interferes with the activation of ITK and the transduction of its cellular functions in T lymphocytes. In the present investigation, we assessed the effects of R9-QQP treatment on the induction of an in vivo immune response as represented by lung inflammation in a murine model of allergic asthma. We found that mice treated with R9-QQP and sensitized and challenged with the surrogate allergen ovalbumin (OVA) display significant inhibition of lung inflammation in a peptide-specific manner. Thus, parameters of the allergic response, such as airway hyper-responsiveness, suppression of inflammatory cell infiltration, reduction of bronchial mucus accumulation, and production of relevant cytokines from draining lymph nodes were significantly suppressed. These findings represent the first demonstration of the biological significance of the interaction between ITK and SLP-76 in the induction of an immune response in a whole animal model and specifically underscore the significance of the ITK-SH3 domain interaction with the poly-proline region of SLP-76 in the development of an inflammatory response. Furthermore, the experimental approach of intracellular peptide-mediated inhibition might be applicable to the study of other important intracellular interactions thus providing a paradigm for dissecting signal transduction pathways.

Topics: Amino Acid Sequence; Animals; Bronchial Hyperreactivity; Bronchioles; Cell-Penetrating Peptides; Cytokines; Eosinophilia; Female; Goblet Cells; Immunity; Lymph Nodes; Lymphocytes; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Molecular Sequence Data; Mucus; Ovalbumin; Pneumonia; Protein-Tyrosine Kinases; Signal Transduction; Th2 Cells

Viral exacerbations of allergen-induced pulmonary inflammation in pre-clinical models reportedly reduce the efficacy of glucocorticoids to limit pulmonary inflammation and airways hyper-responsiveness to inhaled spasmogens. However, exacerbations of airway obstruction induced by allergen challenge have not yet been studied. hPIV-3 (human parainfluenza type 3 virus) inoculation of guinea-pigs increased inflammatory cell counts in BAL (bronchoalveolar lavage) fluid and caused hyper-responsiveness to inhaled histamine. Both responses were abolished by treatment with either dexamethasone (20 mg/kg of body weight, subcutaneous, once a day) or fluticasone propionate (a 0.5 mg/ml solution aerosolized and inhaled over 15 min, twice a day). In ovalbumin-sensitized guinea-pigs, allergen (ovalbumin) challenge caused two phases of airway obstruction [measured as changes in sGaw (specific airways conductance) using whole body plethysmography]: an immediate phase lasting between 4 and 6 h and a late phase at about 7 h. The late phase, airway hyper-responsiveness to histamine and inflammatory cell counts in BAL were all significantly reduced by either glucocorticoid. Inoculation of guinea-pigs sensitized to ovalbumin with hPIV-3 transformed the allergen-induced airway obstruction from two transient phases into a single sustained response lasting up to 12 h. This exacerbated airway obstruction and airway hyper-responsiveness to histamine were unaffected by treatment with either glucocorticoid whereas inflammatory cell counts in BAL were only partially inhibited. Virus- or allergen-induced pulmonary inflammation, individually, are glucocorticoid-sensitive, but in combination generate a phenotype where glucocorticoid efficacy is impaired. This suggests that during respiratory virus infection, glucocorticoids might be less effective in limiting pulmonary inflammation associated with asthma.

Topics: Administration, Inhalation; Allergens; Androstadienes; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dexamethasone; Drug Resistance; Fluticasone; Glucocorticoids; Guinea Pigs; Histamine; Humans; Male; Ovalbumin; Parainfluenza Virus 3, Human; Pneumonia; Respiratory Hypersensitivity; Respirovirus Infections

Airway hyperreactivity and inflammation are important factors in the aggravation of lung function. Suplatast tosilate (IPD) is a novel and unique anti‑asthma clinical compound. However, the mechanisms of IPD action in the inhibition of asthma remain to be elucidated. The present study aimed to investigate the role of the GATA binding protein 3 (GATA‑3)/interleukin (IL)‑5 signaling pathway in IPD‑induced inhibition of asthma. Sprague‑Dawley rats were sensitized by intraperitoneal injection with ovalbumin (OVA) to establish an animal model of asthma. IPD was administered continuously (C‑IPD) or at a later stage (L‑IPD). Budesonide (BUD) was used as a positive control. Airway resistance and the expression of genes at the mRNA and protein levels were measured. Morphological changes in lung tissue and the percentage of eosinophils (EOS) in peripheral blood were observed and correlation analysis was performed. The results revealed that sensitization by OVA significantly increased airway resistance and the percentage of EOS in peripheral blood and induced significant inflammatory changes in lung tissue, as demonstrated by thick epithelium, goblet cell hyperplasia and submucosal cell infiltration. In addition, sensitization by OVA was found to markedly upregulate IL‑5 mRNA and protein expression. Airway resistance was found to positively correlate with the expression of IL‑5 in the rat lung tissues. Sensitization by OVA was also observed to markedly enhance GATA‑3 protein expression and GATA‑3 levels were found to positively correlate with airway resistance and IL‑5 levels. Similar to the effect of BUD, treatment with C‑IPD or L‑IPD was found to significantly attenuate OVA‑induced increases in airway resistance and the percentage of EOS in peripheral blood. Notably, treatment with C‑IPD or L‑IPD markedly reduced the OVA-induced expression of IL‑5 and GATA‑3. In the present study, IPD intervention was demonstrated to ameliorate airway hyperreactivity and inflammation and the mechanisms may involve inhibition of the GATA‑3/IL‑5 signaling pathway.

Topics: Airway Resistance; Animals; Arylsulfonates; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophils; GATA3 Transcription Factor; Histamine Antagonists; Inflammation; Interleukin-5; Leukocyte Count; Lung; Male; Ovalbumin; Rats; Signal Transduction; Sulfonium Compounds

This study was designed to investigate whether the protective effects of Lactobacillus rhamnosus (Lcr35) on allergic asthma are associated with the adoptive transfer of dendritic cells (DCs) and regulatory T cells (Tregs), using a mouse experimental model of asthma.. BALB/c mice were orally administered Lcr35 or intravenously treated with in vivo Lcr35-treated DCs daily and were then sensitized and challenged with ovalbumin (OVA) in accordance with a model of asthma protocol. Both the oral application of Lcr35 and intravenous administration of Lcr35-treated DCs suppressed all aspects of the asthmatic response, including bronchial hyperresponsiveness (BHR), total cell counts in the bronchoalveolar lavage (BAL) fluid, the production of OVA-specificimmunoglobulin E (IgE), and pulmonary eosinophilic inflammation. The mechanism of action of Lcr35 is related to Tregs, which suppress the Th2 response in the respiratory organs, and this is mediated by DCs in the mouse model of asthma.. These results confirm that the mechanism underlying the effects of Lcr35 on asthma involves the adoptive transfer of DCs.. This finding broadens the possibility that Lcr35 has potential as an alternative therapeutic approach to the treatment of human asthma.

Topics: Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dendritic Cells; Disease Models, Animal; Female; Lacticaseibacillus rhamnosus; Mice; Mice, Inbred BALB C; Ovalbumin; Probiotics

Taraxasterol was isolated from the Chinese medicinal herb Taraxacum officinale which has been frequently used as a remedy for inflammatory diseases. In the present study, we determined the in vivo protective effect of taraxasterol on allergic asthma induced by ovalbumin (OVA) in mice.. Mice were sensitized and challenged with OVA, and were orally treated daily with taraxasterol at 2.5, 5 and 10mg/kg from day 23 to 27 after sensitization. The number of inflammatory cells in bronchoalveolar lavage fluid (BALF) was determined. Th2 cytokine interleukin-4 (IL-4), interleukin-5 (IL-5), interleukin-13 (IL-13) production in BALF and OVA-specific immunoglobulin E (IgE) production in sera were measured using ELISA. Histological changes in lung tissues were examined using hematoxylin and eosin (H&E) and periodic acid-Schiff staining (PAS). Airway hyperresponsiveness (AHR) to inhaled methacholine was assessed.. Taraxasterol dramatically decreased the total inflammatory cell and main inflammatory cell counts, reduced the production of Th2 cytokine IL-4, IL-5, IL-13 in BALF and OVA-specific IgE in sera, and suppressed AHR in a dose-dependent manner. Histological studies demonstrated that taraxasterol substantially suppressed OVA-induced inflammatory cells infiltration into lung tissues and goblet cell hyperplasia in airways.. This finding suggests that taraxasterol protects against OVA-induced allergic asthma in mice.

Topics: Allergens; Animals; Anti-Allergic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Female; Immunoglobulin E; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Sterols; Triterpenes

Increased airway smooth muscle (ASM) mass is believed to underlie the relatively fixed airway hyperresponsiveness (AHR) in asthma. Developments of therapeutic approaches to reverse airway remodeling are impeded by our lack of insight on the mechanisms behind the increase in mass of contractile ASM cells. Increased expression of laminin, an extracellular matrix protein, is associated with asthma. Our studies investigate the role of laminin-induced ASM survival signals in the development of increased ASM and AHR. Antagonizing laminin integrin binding using the laminin-selective competing peptide, YIGSR, and mimicking laminin with exogenous α2-chain laminin, we show that laminin is both necessary and sufficient to induce ASM cell survival, concomitant with the induction of ASM contractile phenotype. Using siRNA, we show that the laminin-binding integrin α7β1 mediates this process. Moreover, in laminin-211-deficient mice, allergen-induced AHR was not observed. Notably, ASM cells from asthmatic airways express a higher abundance of intracellular cell survival proteins, consistent with a role for reduced rates of cell apoptosis in development of ASM hyperplasia. Targeting the laminin-integrin α7β1 signaling pathway may offer new avenues for the development of therapies to reduce the increase in mass of contractile phenotype ASM cells that underlie AHR in asthma.

Topics: Animals; Asthma; bcl-2-Associated X Protein; Biomarkers; Bronchial Hyperreactivity; Cell Line; Cell Survival; Female; Humans; Integrins; Laminin; Mice; Mice, Knockout; Muscle Contraction; Muscle, Smooth; Ovalbumin; RNA, Small Interfering; Signal Transduction; Thionucleotides

One feature of allergic asthma, the EAR (early allergic reaction), is not present in the commonly used mouse models. We therefore investigated the mediators involved in EAR in a guinea-pig in vivo model of allergic airway inflammation. Animals were sensitized using a single OVA (ovalbumin)/alum injection and challenged with aerosolized OVA on day 14. On day 15, airway resistance was assessed after challenge with OVA or MCh (methacholine) using the forced oscillation technique, and lung tissue was prepared for histology. The contribution of mast cell mediators was investigated using inhibitors of the main mast cell mediators [histamine (pyrilamine) and CysLTs (cysteinyl-leukotrienes) (montelukast) and prostanoids (indomethacin)]. OVA-sensitized and challenged animals demonstrated AHR (airway hyper-responsiveness) to MCh, and lung tissue eosinophilic inflammation. Antigen challenge induced a strong EAR in the sensitized animals. Treatment with a single compound, or indomethacin together with pyrilamine or montelukast, did not reduce the antigen-induced airway resistance. In contrast, dual treatment with pyrilamine together with montelukast, or triple inhibitor treatment, attenuated approximately 70% of the EAR. We conclude that, as in humans, the guinea-pig allergic inflammation model exhibits both EAR and AHR, supporting its suitability for in vivo identification of mast cell mediators that contribute to the development of asthma. Moreover, the known mast cell mediators histamine and leukotrienes were major contributors of the EAR. The data also lend further support to the concept that combination therapy with selective inhibitors of key mediators could improve asthma management.

Topics: Acetates; Animals; Asthma; Bronchial Hyperreactivity; Constriction, Pathologic; Cyclopropanes; Disease Models, Animal; Guinea Pigs; Histamine Antagonists; Hypersensitivity; Indomethacin; Leukotriene Antagonists; Lung; Mast Cells; Ovalbumin; Prostaglandin Antagonists; Pyrilamine; Quinolines; Sulfides

Vitamin D may be essential for restricting the development and severity of allergic diseases and asthma, but a direct causal link between vitamin D deficiency and asthma has yet to be established. We have developed a 'low dose' model of allergic airway disease induced by intraperitoneal injection with ovalbumin (1 µg) and aluminium hydroxide (0.2 mg) in which characteristics of atopic asthma are recapitulated, including airway hyperresponsiveness, antigen-specific immunoglobulin type-E and lung inflammation. We assessed the effects of vitamin D deficiency throughout life (from conception until adulthood) on the severity of ovalbumin-induced allergic airway disease in vitamin D-replete and -deficient BALB/c mice using this model. Vitamin D had protective effects such that deficiency significantly enhanced eosinophil and neutrophil numbers in the bronchoalveolar lavage fluid of male but not female mice. Vitamin D also suppressed the proliferation and T helper cell type-2 cytokine-secreting capacity of airway-draining lymph node cells from both male and female mice. Supplementation of initially vitamin D-deficient mice with vitamin D for four weeks returned serum 25-hydroxyvitamin D to levels observed in initially vitamin D-replete mice, and also suppressed eosinophil and neutrophil numbers in the bronchoalveolar lavage fluid of male mice. Using generic 16 S rRNA primers, increased bacterial levels were detected in the lungs of initially vitamin D-deficient male mice, which were also reduced by vitamin D supplementation. These results indicate that vitamin D controls granulocyte levels in the bronchoalveolar lavage fluid in an allergen-sensitive manner, and may contribute towards the severity of asthma in a gender-specific fashion through regulation of respiratory bacteria.

Topics: Aerosols; Allergens; Animals; Asthma; Bacteria; Bacterial Load; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dendritic Cells; Disease Models, Animal; Female; Granulocytes; Immunoglobulin E; Immunoglobulin G; Leukocyte Count; Lung; Lymph Nodes; Male; Mice; Mice, Inbred BALB C; Ovalbumin; RNA, Messenger; T-Lymphocytes; Vitamin D; Vitamin D Deficiency

Asthma is a chronic inflammatory disease of the small airways, with airway hyperresponsiveness (AHR) and inflammation as hallmarks. Recent studies suggest a role for arginase in asthma pathogenesis, possibly because arginine is the substrate for both arginase and NO synthase and because NO modulates bronchial tone and inflammation. Our objective was to investigate the importance of increased pulmonary arginase 1 expression on methacholine-induced AHR and lung inflammation in a mouse model of allergic asthma. Arginase 1 expression in the lung was ablated by crossing Arg1(fl/fl) with Tie2Cre(tg/-) mice. Mice were sensitized and then challenged with ovalbumin. Lung function was measured with the Flexivent. Adaptive changes in gene expression, chemokine and cytokine secretion, and lung histology were quantified with quantitative PCR, ELISA, and immunohistochemistry. Arg1 deficiency did not affect the allergic response in lungs and large-airway resistance, but it improved peripheral lung function (tissue elastance and resistance) and attenuated adaptive increases in mRNA expression of arginine-catabolizing enzymes Arg2 and Nos2, arginine transporters Slc7a1 and Slc7a7, chemokines Ccl2 and Ccl11, cytokines Tnfa and Ifng, mucus-associated epithelial markers Clca3 and Muc5ac, and lung content of IL-13 and CCL11. However, expression of Il4, Il5, Il10, and Il13 mRNA; lung content of IL-4, IL-5, IL-10, TNF-α, and IFN-γ protein; and lung pathology were not affected. Correlation analysis showed that Arg1 ablation disturbed the coordinated pulmonary response to ovalbumin challenges, suggesting arginine (metabolite) dependence of this response. Arg1 ablation in the lung improved peripheral lung function and affected arginine metabolism but had little effect on airway inflammation.

Topics: Airway Resistance; Animals; Arginase; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Chemokines; Cytokines; Dendritic Cells; Female; Gene Expression Profiling; Hypersensitivity; Immunoenzyme Techniques; Lung; Macrophages; Male; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Myeloid Cells; Ovalbumin; Pneumonia; Real-Time Polymerase Chain Reaction; Respiratory System; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Epidemiological studies suggest that maternal exposure to environmental hazards, such as particulate matter, is associated with increased incidence of asthma in childhood. We hypothesized that maternal exposure to combustion derived ultrafine particles containing persistent free radicals (MCP230) disrupts the development of the infant immune system and results in aberrant immune responses to allergens and enhances asthma severity.. Pregnant C57/BL6 mice received MCP230 or saline by oropharyngeal aspiration on gestational days 10 and 17. Three days after the second administration, blood was collected from MCP230 or saline treated dams and 8-isoprostanes in the serum were measured to assess maternal oxidative stress. Pulmonary T cell populations were assayed in the infant mice at six days, three and six weeks of postnatal age. When the infant mice matured to adults (i.e. six weeks of age), an asthma model was established with ovalbumin (OVA). Airway inflammation, mucus production and airway hyperresponsiveness were then examined.. Maternal exposure to MCP230 induced systemic oxidative stress. The development of pulmonary T helper (Th1/Th2/Th17) and T regulatory (Treg) cells were inhibited in the infant offspring from MCP230-exposed dams. As the offspring matured, the development of Th2 and Treg cells recovered and eventually became equivalent to that of offspring from non-exposed dams. However, Th1 and Th17 cells remained attenuated through 6 weeks of age. Following OVA sensitization and challenge, mice from MCP230-exposed dams exhibited greater airway hyperresponsiveness, eosinophilia and pulmonary Th2 responses compared to offspring from non-exposed dams.. Our data suggest that maternal exposure to MCP230 enhances postnatal asthma development in mice, which might be related to the inhibition of pulmonary Th1 maturation and systemic oxidative stress in the dams.

Topics: Age Factors; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Dinoprost; Female; Gestational Age; Inflammation Mediators; Inhalation Exposure; Lung; Maternal Exposure; Mice, Inbred C57BL; Ovalbumin; Oxidative Stress; Particulate Matter; Pregnancy; Prenatal Exposure Delayed Effects; Pulmonary Eosinophilia; Severity of Illness Index; T-Lymphocytes, Regulatory; Th1 Cells; Th17 Cells; Th2 Cells

Our group recently reported the strong anti-inflammatory effects of geniposide (Gen), a bioactive iridoid glucoside derived from gardenia jasminoides, in a mouse acute lung injury model. Herein, we hypothesized that Gen might also have potential therapeutic benefits in treatment of asthma, which was tested in a mouse model of ovalbumin (Ova)-induced allergic airway inflammation. Ova-sensitized and -challenged BALB/c mice, as compared with control animals, displayed airway hyperresponsiveness (AHR), bronchoalveolar lavage eosinophilia, mucus hypersecretion, and increased T help 2 (Th2)-associated cytokine and chemokine amounts, as well as serum Ova-specific immunoglobulin E (IgE) level. Being compared with the Ova-induced hallmarks of asthma, intraperitoneal Gen treatment prevented eosinophilic pulmonary infiltration, attenuated the increases in interleukin (IL)-4, IL-5, and IL-13, and reduced eotaxin and vascular cell adhesion molecule 1 (VCAM-1) expression. Also, Gen significantly ameliorated the Ova-driven airway hyperresponsiveness, mucus hypersecretion, and allergen-specific IgE level, which are the cardinal pathophysiological symptoms in allergic airway diseases. In addition, the efficacy of Gen was comparable to that of dexamethasone (Dex), a currently available anti-asthmatic drug. Collectively, our findings reveal that the development of immunoregulatory strategies based on Gen may be considered as an effective adjuvant therapy for allergic asthma.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Iridoids; Lung; Mice; Mice, Inbred BALB C; Ovalbumin

Second-hand smoke (SHS) exposure in utero exacerbates adult responses to environmental irritants. We tested the hypothesis that effects of in utero SHS exposure on modulating physiological and transcriptome responses in BALB/c mouse lungs after ovalbumin (OVA) challenge extend well into adulthood, and that the responses show a sex bias. We exposed BALB/c mice in utero to SHS or filtered air (AIR), then sensitized and challenged all offspring with OVA from 19 to 23 weeks of age. At the end of the adult OVA challenge, we evaluated pulmonary function, examined histopathology, analyzed bronchoalveolar lavage fluid (BALF), and assessed gene expression changes in the lung samples. All groups exhibited lung inflammation and inflammatory cell infiltration. Pulmonary function testing (airway hyperresponsiveness [AHR], breathing frequency [f]) and BALF (cell differentials, Th1/Th2 cytokines) assessments showed significantly more pronounced lung responses in the SHS-OVA groups than in AIR-OVA groups (AHR, f; eosinophils, neutrophils; IFN-γ, IL-1b, IL-4, IL-5, IL-10, IL-13, KC/CXCL1, TNF-α), with the majority of responses being more pronounced in males than in females. SHS exposure in utero also significantly altered lung gene expression profiles, primarily of genes associated with inflammatory responses and respiratory diseases, including lung cancer and lung fibrosis. Altered expression profiles of chemokines (Cxcl2, Cxcl5, Ccl8, Ccl24), cytokines (Il1b, Il6, Il13) and acute phase response genes (Saa1, Saa3) were confirmed by qRT-PCR. In conclusion, in utero exposure to SHS exacerbates adult lung responses to OVA challenge and promotes a pro-asthmatic milieu in adult lungs; further, males are generally more affected by SHS-OVA than are females.

Topics: Acute-Phase Reaction; Animals; Bronchial Hyperreactivity; Chemokines; Cytokines; Female; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Pregnancy; Prenatal Exposure Delayed Effects; Sex Characteristics; Tobacco Smoke Pollution; Up-Regulation

Bronchial asthma is a chronic inflammatory disease of the airway. Apoptosis signal-regulating kinase 1 (ASK1), a member of the mitogen-activated protein kinase kinase kinase family, is activated by environmental stress and plays a crucial role in the induction of apoptosis and inflammation. To examine whether ASK1 is involved in the induction of bronchial asthma, we investigated the role of ASK1 using a genetic approach in the production of cytokines, as well as the development of airway hyperreactivity (AHR) and antibody responses using a murine airway inflammation model.. ASK1-deficient (ASK1(-/-)) and control wild-type (WT) mice were immunized with ovalbumin (OVA) without alum intraperitoneally, followed by intranasal administration of OVA. Airway infiltration of inflammatory cells, cytokine production, AHR and antibody production were assayed. The asthmatic phenotype was assessed following intranasal administration of IL-13 or TNF-α.. ASK1(-/-) mice sensitized with OVA displayed an impaired inflammatory cell infiltration into airways and a decreased AHR relative to WT mice. Moreover, the production of OVA-specific IgE antibodies and proasthmatic cytokines (IL-5, IL-13 and TNF-α) was substantially reduced in OVA-stimulated ASK1(-/-) mice. Intranasal administration of IL-13 and OVA enhanced the accumulation of inflammatory cells in OVA-primed ASK1(-/-) mice. The OVA-induced AHR in response to methacholine was enhanced by IL-13 in WT mice but not ASK1(-/-) mice.. The ASK1 signaling pathway regulates the OVA-induced asthmatic phenotype, specifically AHR sensitivity and cytokine production. Therefore, the ASK1 signaling pathway is a promising target for therapeutic intervention in some asthmatic patients.

Topics: Animals; Apoptosis; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Goblet Cells; Immunoglobulin E; Inflammation; Interleukin-13; Interleukin-5; Lung; MAP Kinase Kinase Kinase 5; MAP Kinase Signaling System; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophil Infiltration; Ovalbumin; Tumor Necrosis Factor-alpha

Allergic asthma is characterized by Th2 type inflammation, leading to airway hyperresponsivenes, mucus hypersecretion and tissue remodeling. S-Nitrosoglutathione reductase (GSNOR) is an alcohol dehydrogenase involved in the regulation of intracellular levels of S-nitrosothiols. GSNOR activity has been shown to be elevated in human asthmatic lungs, resulting in diminished S-nitrosothiols and thus contributing to increased airway hyperreactivity. Using a mouse model of allergic airway inflammation, we report that intranasal administration of a new selective inhibitor of GSNOR, SPL-334, caused a marked reduction in airway hyperreactivity, allergen-specific T cells and eosinophil accumulation, and mucus production in the lungs in response to allergen inhalation. Moreover, SPL-334 treatment resulted in a significant decrease in the production of the Th2 cytokines IL-5 and IL-13 and the level of the chemokine CCL11 (eotaxin-1) in the airways. Collectively, these observations reveal that GSNOR inhibitors are effective not only in reducing airway hyperresponsiveness but also in limiting lung inflammatory responses mediated by CD4(+) Th2 cells. These findings suggest that the inhibition of GSNOR may provide a novel therapeutic approach for the treatment of allergic airway inflammation.

Topics: Administration, Intranasal; Alcohol Dehydrogenase; Allergens; Animals; Anti-Inflammatory Agents, Non-Steroidal; Benzoates; Bronchial Hyperreactivity; Cell Movement; Chemokine CCL11; Enzyme Inhibitors; Eosinophils; Female; Glutathione Reductase; Humans; Interleukin-13; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Mice, Transgenic; Ovalbumin; Pneumonia; Pyrimidinones; Th2 Cells

Asthma is a chronic disease characterized by airways hyperresponsiveness, inflammation and airways remodelling involving reversible bronchial obstruction. Omega-3 fatty acids and their derivatives are known to reduce inflammation in several tissues including lung.. The effects of eicosapentaenoic acid monoacylglyceride (MAG-EPA), a newly synthesized EPA derivative, were determined on the resolution of lung inflammation and airway hyperresponsiveness in an in vivo model of allergic asthma.. Ovalbumin (OVA)-sensitized guinea-pigs were treated or not with MAG-EPA administered per os. Isometric tension measurements, histological analyses, homogenate preparation for Western blot experiments or total RNA extraction for RT-PCR were performed to assess the effect of MAG-EPA treatments.. Mechanical tension measurements revealed that oral MAG-EPA treatments reduced methacholine (MCh)-induced bronchial hyperresponsiveness in OVA-sensitized guinea-pigs. Moreover, MAG-EPA treatments also decreased Ca(2+) hypersensitivity of bronchial smooth muscle. Histological analyses and leucocyte counts in bronchoalveolar lavages revealed that oral MAG-EPA treatments led to less inflammatory cell recruitment in the lung of OVA-sensitized guinea-pigs when compared with lungs from control animals. Results also revealed a reduction in mucin production and MUC5AC expression level in OVA-sensitized animals treated with MAG-EPA. Following MAG-EPA treatments, the transcript levels of pro-inflammatory markers such as IL-5, eotaxin, IL-13 and IL-4 were markedly reduced. Moreover, per os MAG-EPA administrations reduced COX2 over-expression in OVA-sensitized animals.. We demonstrate that MAG-EPA reduces airway hyperresponsiveness and lung inflammation in OVA-sensitized animals, a finding consistent with a decrease in IL-4, IL-5, IL-13, COX-2 and MUC5AC expression levels in the lung. The present data suggest that MAG-EPA represents a new potential therapeutic strategy for resolving inflammation in allergic asthma.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Cyclooxygenase 2; Disease Models, Animal; Fatty Acids; Female; Guinea Pigs; Lung; Male; Monoglycerides; Mucins; Ovalbumin; Receptors, Chemokine

Allergic disorders have increased substantially in recent years. Asthma is characterized by airway damage and remodeling. Reprogramming induced pluripotent stem cells (iPSCs) from adult somatic cells transfected by Oct-4/Sox-2/Klf-4, but not c-Myc, has shown the potential of embryonic-like cells. These cells have potential for multilineage differentiation and provide a resource for stem cell-based utility. However, the therapeutic potential of iPSCs without c-Myc (iPSC-w/o-c-Myc) in allergic diseases and airway hyperresponsiveness has not been investigated. The aim of this study was to evaluate the therapeutic effect of iPSC-w/o-c-Myc transplantation in a murine asthma model.. BALB/c mice were sensitized with alum-adsorbed ovalbumin (OVA) and then challenged with aerosolized OVA. Phosphate-buffered saline or iPSC-w/o-c-Myc was then intravenously injected after inhalation. Serum allergen-specific antibody levels, airway hyperresponsiveness, cytokine levels in spleen cells and bronchoalveolar lavage fluid (BALF), and cellular distribution in BALF were then examined.. Treatment with iPSC-w/o-c-Myc effectively suppressed both Th1 and Th2 antibody responses, which was characterized by reduction in serum allergen-specific IgE, IgG, IgG1, and IgG2a levels as well as in interleukin-5 and interferon-γ levels in BALF and in OVA-incubated splenocytes. Meanwhile, regulatory cytokine, interleukin-10, was enhanced. Transplantation of iPSC-w/o-c-Myc also significantly attenuated cellular infiltration in BALF and allergic airway hyperresponsiveness. However, no tumor formation was observed 6 months after transplantation.. Administration of iPSC-w/o-c-Myc not only inhibited Th1 inflammatory responses but also had therapeutic effects on systemic allergic responses and airway hyperresponsiveness. iPSC-w/o-c-Myc transplantation may be a potential modality for treating allergic reactions and bronchial asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Induced Pluripotent Stem Cells; Inflammation Mediators; Kruppel-Like Factor 4; Kruppel-Like Transcription Factors; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Octamer Transcription Factor-3; Ovalbumin; Proto-Oncogene Proteins c-myc; SOXB1 Transcription Factors; Spleen; Th1 Cells; Th2 Cells; Time Factors; Transfection

Allergic asthma is a chronic inflammatory airway disease with increasing prevalence around the world. Current asthma therapy includes drugs that usually cause significant side effects, justifying the search for new anti-asthmatic drugs. Curine is a bisbenzylisoquinoline alkaloid that modulates calcium influx in many cell types; however, its anti-allergic and putative toxic effects remain to be elucidated. Our aim was to investigate the effects of curine on eosinophil activation and airway hyper-responsiveness (AHR) and to characterize its potential toxic effects. We used a mouse model of allergic asthma induced by sensitization and challenge with ovalbumin (OVA) to evaluate the anti-allergic effects of oral treatment with curine. The oral administration of curine significantly inhibited eosinophilic inflammation, eosinophil lipid body formation and AHR in animals challenged with OVA compared with animals in the untreated group. The curine treatment also reduced eotaxin and IL-13 production triggered by OVA. Verapamil, a calcium channel antagonist, had similar anti-allergic properties, and curine pre-treatment inhibited the calcium-induced tracheal contractile response ex-vivo, suggesting that the mechanism by which curine exerts its effects is through the inhibition of a calcium-dependent response. A toxicological evaluation showed that orally administered curine did not significantly alter the biochemical, hematological, behavioral and physical parameters measured in the experimental animals compared with saline-treated animals. In conclusion, curine showed anti-allergic activity through mechanisms that involve inhibition of IL-13 and eotaxin and of Ca(++) influx, without inducing evident toxicity and as such, has the potential for the development of anti-asthmatic drugs.

Topics: Administration, Oral; Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Calcium; Disease Models, Animal; Eosinophils; Inflammation; Interleukin-13; Isoquinolines; Male; Menispermaceae; Mice; Mice, Inbred BALB C; No-Observed-Adverse-Effect Level; Ovalbumin; Rats; Rats, Wistar; Verapamil

This study investigates whether allergen exposure elevates the risk of diabetes or cardiovascular diseases using acute OVA (Ovalbumin) allergen exposure model. We hypothesize that exposure to allergen can induce adipose tissue inflammation and affect adiponectin levels. An intranasal challenge with OVA male C57BL/6 mice was performed at dose of 6.25, 12.5, 25, 50 and 100μg, and compared to which challenge with PBS (phosphate buffered saline). Results showed that acute OVA exposure did not only cause airway inflammation in study mice, but also decreased serum adiponectin levels with a dose-response effect. When examining the gonadal adipose tissues, there was no significantly difference of adiponectin mRNA in OVA challenged mice compared to those PBS challenged, but lower inguinal adiponectin mRNA expression was found compared to those PBS-challenged, and had a good relationship with the serum adiponectin. Inguinal adipose tissues of OVA challenged mice, had significantly lower adipose tissue weight, and higher TNF-α expression without statistical significance. Our data indicate that acute OVA exposure appears to affect the characteristics of adipose tissues, and change the adiponectin levels in serum and adipose tissues. Allergen exposure may be considered a potential risk factor for presenting diabetes or cardiovascular diseases.

Topics: Adiponectin; Adipose Tissue; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Inflammation; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Real-Time Polymerase Chain Reaction; RNA

The relative contributions of large and small airways to hyperresponsiveness in asthma have yet to be fully assessed. This study used a mouse model of chronic allergic airways disease to induce inflammation and remodelling and determine whether in vivo hyperresponsiveness to methacholine is consistent with in vitro reactivity of trachea and small airways. Balb/C mice were sensitised (days 0, 14) and challenged (3 times/week, 6 weeks) with ovalbumin. Airway reactivity was compared with saline-challenged controls in vivo assessing whole lung resistance, and in vitro measuring the force of tracheal contraction and the magnitude/rate of small airway narrowing within lung slices. Increased airway inflammation, epithelial remodelling and fibrosis were evident following allergen challenge. In vivo hyperresponsiveness to methacholine was maintained in isolated trachea. In contrast, methacholine induced slower narrowing, with reduced potency in small airways compared to controls. In vitro incubation with IL-1/TNFα did not alter reactivity. The hyporesponsiveness to methacholine in small airways within lung slices following chronic ovalbumin challenge was unexpected, given hyperresponsiveness to the same agonist both in vivo and in vitro in tracheal preparations. This finding may reflect the altered interactions of small airways with surrounding parenchymal tissue after allergen challenge to oppose airway narrowing and closure.

Topics: Airway Remodeling; Allergens; Animals; Antibody Specificity; Bronchial Hyperreactivity; Bronchial Provocation Tests; Calcium; Dinoprostone; Disease Models, Animal; Female; Immunoglobulin E; Inflammation; Methacholine Chloride; Mice; Ovalbumin

Phenotypes of lung smooth muscle cells in health and disease are poorly characterized. This is due, in part, to a lack of methodologies that allow for the independent and direct isolation of bronchial smooth muscle cells (BSMCs) and vascular smooth muscle cells (VSMCs) from the lung. In this paper, we describe the development of a bi-fluorescent mouse that permits purification of these two cell populations by cell sorting. By subjecting this mouse to an acute allergen based-model of airway inflammation that exhibits many features of asthma, we utilized this tool to characterize the phenotype of so-called asthmatic BSMCs. First, we examined the biophysical properties of single BSMCs from allergen sensitized mice and found increases in basal tone and cell size that were sustained ex vivo. We then generated for the first time, a comprehensive characterization of the global gene expression changes in BSMCs isolated from the bi-fluorescent mice with allergic airway inflammation. Using statistical methods and pathway analysis, we identified a number of differentially expressed mRNAs in BSMCs from allergen sensitized mice that code for key candidate proteins underlying changes in matrix formation, contractility, and immune responses. Ultimately, this tool will provide direction and guidance for the logical development of new markers and approaches for studying human lung smooth muscle.

Topics: Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Cell Size; Disease Models, Animal; Fluorescence; Gene Expression; Gene Expression Profiling; Humans; Immunization; Inflammation; Mice; Mice, Transgenic; Muscle, Smooth, Vascular; Myocytes, Smooth Muscle; Ovalbumin; Phenotype; Proteome; RNA, Messenger; Single-Cell Analysis

Fish oil (FO) is rich in n-3 polyunsaturated fatty acids (PUFA), which have been suggested to be anti-inflammatory and are associated with improvement of several inflammatory diseases. In this study, we investigated the influence of FO on allergen-induced lung inflammation and airway hyperreactivity in mice.. Male A/J mice were fed either a standard-chow (SC) or a FO diet (FO) for 8 weeks. After 4 weeks, each group was further randomized for ovalbumin (SC-OVA and FO-OVA) or saline (SC-SAL and FO-SAL) challenge. Resistance and elastance were measured at baseline and after aerosolized methacholine, 24h after the last challenge. Bronchoalveolar lavage (BAL) was performed for leukocyte counts. Lung tissue mucus deposition, peribronchiolar matrix deposition and eosinophil infiltration were quantified. Serum immunoglobulin E (IgE) and IgG1 (ref 2.2), lung IL-4, IL-5, IL-10, IL-13, IL-17, INFγ and eotaxin-1 and 2 were detected by ELISA and nuclear factor kappa B (NFκB), GATA-3 and peroxisome proliferator-activated receptor gamma (PPARγ) expression was measured by Western blot.. Levels of serum IgE and IgG1 were significantly higher in OVA sensitized mice. OVA challenge resulted in increased eosinophil infiltration, increased inflammatory cytokine production, peribronchiolar matrix and mucus deposition and airway hyperreactivity to aerosolized methacholine. Elevated lung NFκB and GATA-3 expression was noted in OVA-challenged mice. These changes were attenuated in mice fed with FO diet. Higher PPARγ expression was also detected in the lungs from the FO-fed groups.. Our results demonstrate that FO intake attenuated classical asthma features by suppressing the systemic sensitization, thus providing evidence that FO might be a prophylactic alternative for asthma prevention.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Cytokines; Fatty Acids, Unsaturated; Fish Oils; Immunoglobulin E; Immunoglobulin G; Inflammation; Leukocytes; Lung; Male; Methacholine Chloride; Mice; Ovalbumin; Pancreatic Elastase

To investigate whether eosinophilic bronchitis without airway hyperresponsiveness will develop bronchial asthma in allergic mice.. Mice were sensitized with OVA on days 0, 7, and 14, challenged on days 21 to 23 (1(st) OVA challenge), and re-challenged on days 46 to 48 (2(nd) OVA challenge), intranasally with 10 (the EB group) and 200 (the AS group) μg OVA. Lung resistance (RL) was assessed 24 h after each challenge and on day 45 followed by analysis of leukocyte distribution in the bronchoalveolar lavage (BAL) fluid and histological examination.. Twenty-four hours after the 1(st) OVA challenge, aerosolized methacholine caused a dose-dependent increase in RL in all groups. At doses ≥1.56 mg/mL, RL in the AS group was significantly higher than that of the NS-1 group (P<0.01 or 0.05) and at doses ≥12.5 mg/mL, RL was markedly higher in the AS group than that of the EB group (P<0.01). The percentage of eosinophils in both the EB group and the AS group was markedly higher than that of the control group. Twenty-four hours after the 2(nd) OVA challenge, at doses ≤12.5 mg/mL, there was no significant difference in RL among all groups (P>0.05). At doses ≥12.5 mg/mL, RL in the AS group was significantly higher than that of the control group and EB group (P<0.01 or 0.05). The percentage of eosinophils in the AS group was noticeably higher than that of the EB group(P<0.05). Furthermore, there was apparent infiltration by inflammatory cells, predominantly eosinophils, into the sub-epithelial region of the bronchus and the bronchioles and around the vessels in the EB and AS group.. Re-challenge with low doses of ovalbumin did not increase airway reactivity and failed to induce bronchial asthma in mice with ovalbumin-induced EB.

Topics: Administration, Intranasal; Aerosols; Animals; Asthma; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophils; Female; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Pulmonary Eosinophilia

Asthma is a chronic disease that is characterized by airway hyperresponsiveness and airway remodeling. The underlying mechanisms that mediate the pathological processes are not fully understood. Abl is a non-receptor protein tyrosine kinase that has a role in the regulation of smooth muscle contraction and smooth muscle cell proliferation in vitro. The role of Abl in airway hyperresponsiveness and airway remodeling in vivo is largely unknown.. To evaluate the role of Abl in asthma pathology, we assessed the expression of Abl in airway tissues from the ovalbumin sensitized and challenged mouse model, and human asthmatic airway smooth muscle cells. In addition, we generated conditional knockout mice in which Abl expression in smooth muscle was disrupted, and then evaluated the effects of Abl conditional knockout on airway resistance, smooth muscle mass, cell proliferation, IL-13 and CCL2 in the mouse model of asthma. Furthermore, we determined the effects of the Abl pharmacological inhibitors imatinib and GNF-5 on these processes in the animal model of asthma.. The expression of Abl was upregulated in airway tissues of the animal model of asthma and in airway smooth muscle cells of patients with severe asthma. Conditional knockout of Abl attenuated airway resistance, smooth muscle mass and staining of proliferating cell nuclear antigen in the airway of mice sensitized and challenged with ovalbumin. Interestingly, conditional knockout of Abl did not affect the levels of IL-13 and CCL2 in bronchoalveolar lavage fluid of animals treated with ovalbumin. However, treatment with imatinib and GNF-5 inhibited the ovalbumin-induced increase in IL-13 and CCL2 as well as airway resistance and smooth muscle growth in animals.. These results suggest that the altered expression of Abl in airway smooth muscle may play a critical role in the development of airway hyperresponsiveness and airway remodeling in asthma. Our findings support the concept that Abl may be a novel target for the development of new therapy to treat asthma.

Topics: Airway Remodeling; Animals; Asthma; Benzamides; Bronchi; Bronchial Hyperreactivity; Cells, Cultured; Chemokine CCL2; Disease Models, Animal; Female; Humans; Imatinib Mesylate; In Vitro Techniques; Interleukin-13; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocytes, Smooth Muscle; Ovalbumin; Piperazines; Proliferating Cell Nuclear Antigen; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-abl; Pyrimidines

The current paradigm describing asthma pathogenesis recognizes the central role of abnormal epithelial function in the generation and maintenance of the disease. However, the mechanisms responsible for the initiation of airway remodeling, which contributes to decreased lung function, remain elusive. Therefore, we aimed to determine the role of altered pulmonary gene expression in disease inception and identify proremodeling mediators.. Using an adenoviral vector, we generated mice overexpressing smad2, a TGF-β and activin A signaling molecule, in the lung. Animals were exposed to intranasal ovalbumin (OVA) without systemic sensitization.. Control mice exposed to inhaled OVA showed no evidence of pulmonary inflammation, indices of remodeling, or airway hyper-reactivity. In contrast, local smad2 overexpression provoked airway hyper-reactivity in OVA-treated mice, concomitant with increased airway smooth muscle mass and peribronchial collagen deposition. Pulmonary eosinophilic inflammation was not evident, and there was no change in serum IgE or IgG1 levels. The profound remodeling changes were not mediated by classical pro-inflammatory Th2 cytokines. However, uric acid and interleukin-1β levels in the lung were increased. Epithelial-derived endothelin-1 and fibroblast growth factor were also augmented in smad2-expressing mice. Blocking endothelin-1 prevented these phenotypic changes.. Innate epithelial-derived mediators are sufficient to drive airway hyper-reactivity and remodeling in response to environmental insults in the absence of overt Th2-type inflammation in a model of noneosinophilic, noninflammed types of asthma. Targeting potential asthma therapies to epithelial cell function and modulation of locally released mediators may represent an effective avenue for therapeutic design.

Topics: Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Endothelin-1; Female; Gene Expression; Inflammation Mediators; Mice; Mice, Transgenic; Muscle, Smooth; Ovalbumin; Respiratory Mucosa; Smad2 Protein

Airway inflammation and airway remodeling are the key contributors to airway hyperresponsiveness (AHR), a characteristic feature of asthma. Both processes are regulated by Transforming Growth Factor (TGF)-β. Caveolin 1 (Cav1) is a membrane bound protein that binds to a variety of receptor and signaling proteins, including the TGF-β receptors. We hypothesized that caveolin-1 deficiency promotes structural alterations of the airways that develop with age will predispose to an increased response to allergen challenge.. AHR was measured in Cav1-deficient and wild-type (WT) mice 1 to 12 months of age to examine the role of Cav1 in AHR and the relative contribution of inflammation and airway remodeling. AHR was then measured in Cav1-/- and WT mice after an ovalbumin-allergen challenge performed at either 2 months of age, when remodeling in Cav1-/- and WT mice was equivalent, and at 6 months of age, when the Cav1-/- mice had established airway remodeling.. Cav1-/- mice developed increased thickness of the subepithelial layer and a correspondingly increased AHR as they aged. In addition, allergen-challenged Cav1-/- mice had an increase in AHR greater than WT mice that was largely independent of inflammation. Cav1-/- mice challenged at 6 months of age have decreased AHR compared to those challenged at 2 months with correspondingly decreased BAL IL-4 and IL-5 levels, inflammatory cell counts and percentage of eosinophils. In addition, in response to OVA challenge, the number of goblet cells and α-SMA positive cells in the airways were reduced with age in response to OVA challenge in contrast to an increased collagen deposition further enhanced in absence of Cav1.. A lack of Cav1 contributed to the thickness of the subepithelial layer in mice as they aged resulting in an increase in AHR independent of inflammation, demonstrating the important contribution of airway structural changes to AHR. In addition, age in the Cav1-/- mice is a contributing factor to airway remodeling in the response to allergen challenge.

Topics: Actins; Aging; Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Caveolin 1; Collagen; Disease Models, Animal; Female; Interleukin-4; Interleukin-5; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Pneumonia; Transforming Growth Factor beta

Ovarian cancer G protein-coupled receptor 1 (OGR1) stimulation by extracellular protons causes the activation of G proteins and subsequent cellular functions. However, the physiological and pathophysiological roles of OGR1 in airway responses remain largely unknown. In the present study, we show that OGR1-deficient mice are resistant to the cardinal features of asthma, including airway eosinophilia, airway hyperresponsiveness (AHR), and goblet cell metaplasia, in association with a remarkable inhibition of Th2 cytokine and IgE production, in an ovalbumin (OVA)-induced asthma model. Intratracheal transfer to wild-type mice of OVA-primed bone marrow-derived dendritic cells (DCs) from OGR1-deficient mice developed lower AHR and eosinophilia after OVA inhalation compared with the transfer of those from wild-type mice. Migration of OVA-pulsed DCs to peribronchial lymph nodes was also inhibited by OGR1 deficiency in the adoption experiments. The presence of functional OGR1 in DCs was confirmed by the expression of OGR1 mRNA and the OGR1-sensitive Ca(2+) response. OVA-induced expression of CCR7, a mature DC chemokine receptor, and migration response to CCR7 ligands in an in vitro Transwell assay were attenuated by OGR1 deficiency. We conclude that OGR1 on DCs is critical for migration to draining lymph nodes, which, in turn, stimulates Th2 phenotype change and subsequent induction of airway inflammation and AHR.

Topics: Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Calcium; Cell Movement; Dendritic Cells; Female; Gene Expression Regulation; Goblet Cells; Immunoglobulin E; Lung; Lymph Nodes; Mice; Mice, Knockout; Ovalbumin; Pulmonary Eosinophilia; Receptors, CCR7; Receptors, G-Protein-Coupled; Signal Transduction; Th1-Th2 Balance; Th2 Cells

Multipotent mesenchymal stromal cells (MSCs) possess reparative and immunoregulatory properties, making them attractive candidates for cellular therapy. However, the majority of MSCs administered i.v. encounter a pulmonary impasse and soon disappear from the lungs, raising the question of how they induce such durable immunosuppressive effects. Using a mouse model of allergic asthma, we show that administration of MSCs isolated from human bone marrow, umbilical cord, or adipose tissue provoked a pronounced increase in alveolar macrophages and inhibited hallmark features of asthma, including airway hyperresponsiveness, eosinophilic accumulation, and Th2 cytokine production. Importantly, selective depletion of this macrophage compartment reversed the therapeutic benefit of MSC treatment on airway hyperresponsiveness. Our data demonstrate that human MSCs exert cross-species immunosuppressive activity, which is mediated by alveolar macrophages in allergic asthma. As alveolar macrophages are the predominant immune effector cells at the air-tissue interface in the lungs, this study provides a compelling mechanism for durable MSC effects in the absence of sustained engraftment.

Topics: Adipose Tissue; Animals; Asthma; Bone Marrow Cells; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Clodronic Acid; Eosinophilia; Female; Genes, Reporter; Graft Survival; Heterografts; Humans; Immunization; Immunosuppression Therapy; Interleukin-10; Lung; Lymphokines; Macrophages, Alveolar; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Methacholine Chloride; Mice; Mice, Inbred BALB C; Organ Specificity; Ovalbumin; Species Specificity; Specific Pathogen-Free Organisms; Th2 Cells; Transduction, Genetic; Umbilical Cord

Trefoil factor 2 (TFF2) is a small peptide with an important role in mucosal repair. TFF2 is up-regulated in asthma, suggesting a role in asthma pathogenesis. Given its known biological role in promoting epithelial repair, TFF2 might be expected to exert a protective function in limiting the progression of airway remodeling in asthma. The contribution of TFF2 to airway remodeling in asthma was investigated by examining the expression of TFF2 in the airway and lung, and evaluating the effects of recombinant TFF2 treatment on established airway remodeling in a murine model of chronic allergic airways disease (AAD). BALB/c mice were sensitized and challenged with ovalbumin (OVA) or saline for 9 weeks, whereas mice with established OVA-induced AAD were treated with TFF2 or vehicle control (intranasally for 14 d). Effects on airway remodeling, airway inflammation, and airway hyperresponsiveness were then assessed, whereas TFF2 expression was determined by immunohistochemistry. TFF2 expression was significantly increased in the airways of mice with AAD, compared with expression levels in control mice. TFF2 treatment resulted in reduced epithelial thickening, subepithelial collagen deposition, goblet-cell metaplasia, bronchial epithelium apoptosis, and airway hyperresponsiveness (all P < 0.05, versus vehicle control), but TFF2 treatment did not influence airway inflammation. The increased expression of endogenous TFF2 in response to chronic allergic inflammation is insufficient to prevent the progression of airway inflammation and remodeling in a murine model of chronic AAD. However, exogenous TFF2 treatment is effective in reversing aspects of established airway remodeling. TFF2 has potential as a novel treatment for airway remodeling in asthma.

Topics: Actins; Airway Resistance; Animals; Annexin A5; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; ErbB Receptors; Female; Immunohistochemistry; Lung; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Inbred BALB C; Mucins; Muscle Proteins; Ovalbumin; Peptides; Recombinant Proteins; Respiratory Mucosa; Transforming Growth Factor beta1; Trefoil Factor-2

Phosphodiesterases (PDEs) are enzymes responsible for degradation of cAMP and cGMP in cells. Thus, PDE inhibitors may have significant clinical benefit in respiratory diseases associated with inflammation. The aim of the present study was to evaluate the effects of selective PDE4 (rolipram, ROL) and PDE7 inhibitors (BRL50481, BRL) on citric acid-induced cough, in vivo and in vitro airway smooth muscle reactivity in both healthy and ovalbumin sensitized guinea pigs. The drugs tested were administered intraperitoneally to male guinea pigs once daily for 7 days - ROL 1 mg/kg, BRL 1 mg/kg, and ROL+BRL 0.5 mg/kg. Double chamber whole body plethysmography was used for the evaluation of citric acid (0.6 M)-induced cough and specific airway resistance. An organ bath method was used for the measurement of tracheal and lung tissue strip contractions evoked by cumulative doses (10(-8)-10(-3) mol/L) of acetylcholine (ACH) and histamine (HIS). In healthy guinea pigs, the only significant relaxation was observed after ROL in ACH-induced contractions in vitro and the effect on cough was negligible. In ovalbumin-sensitized animals, more pronounced in vitro relaxing effects of BRL in HIS-induced contractions and of combination (ROL+BRL) in ACH-induced contractions were observed, with similar results in vivo, and no significant change in the number of cough efforts was observed in any of the groups tested. The results suggest that PDE4 and PDE7 inhibitors have stronger anti-inflammatory effects compared with direct effects on smooth muscle and cough, with a potential benefit of their concomitant administration.

Topics: Acetylcholine; Airway Resistance; Animals; Bronchial Hyperreactivity; Bronchoconstriction; Citric Acid; Cough; Cyclic Nucleotide Phosphodiesterases, Type 4; Cyclic Nucleotide Phosphodiesterases, Type 7; Guinea Pigs; Histamine; Male; Muscle Contraction; Muscle, Smooth; Ovalbumin; Phosphodiesterase 4 Inhibitors; Plethysmography, Whole Body; Respiratory System; Rolipram

The interest in L-arginine metabolism was triggered primarily by the discovery of nitric oxide (NO) synthesis in mammals and its remarkable biological roles. The real role of L-arginine in the airway hyperreactivity (AHR) has not been established yet. Therefore, we studied whether supplementation of L-arginine can influence the experimental AHR evoked by two different triggers - allergen and exogenous irritant (toluene vapours). Male TRIK strain guinea pigs were used in the study. We used two patterns of pretreatment with L-arginine in vivo, short- and long-term, in a dose of 300 mg/kg administered i.p., after which we studied reactivity of airway smooth muscles in vitro. Pretreatment with L-arginine for 3 days decreased the airway smooth muscle reactivity induced by toluene vapour, whereas pretreatment for 17 days was without any additional effect on smooth muscle reactivity. The short-term pretreatment in ovalbumin-induced hyperreactivity caused an increase in airway smooth muscle reactivity to lower concentrations of both bronchoconstrictors. On the other side, this pretreatment significantly decreased smooth muscle reactivity to high concentrations of both bronchoconstrictors. Supplementation of L-arginine resulted in a modification of the airway smooth muscle response. The effect of supplementation was different depending on the AHR trigger, airway region and pretreatment duration. The results also underscore the importance of an optimal L-arginine level for the control of bronchial tone.

Topics: Animals; Arginine; Bronchial Hyperreactivity; Bronchoconstriction; Dietary Supplements; Guinea Pigs; Lung; Male; Muscle Contraction; Muscle, Smooth; Nitric Oxide; Ovalbumin; Respiratory Hypersensitivity; Respiratory System; Toluene

The aim of the study was to investigate the potential anti-inflammatory effects in -experimental allergic asthma of natural polyphenolic compounds or their single major components. The experiment was performed after 21-days sensitization of guinea pigs with ovalbumin suspension. Changes in airway reactivity after the long-term treatment with the polyphenolic compounds Provinol and Flavin-7 and their single major components quercetin and resveratrol during were assessed using a whole body plethysmography. Reactivity of tracheal smooth muscle was studied in vitro in response to cumulative doses of the bronchoconstrictive mediators histamine and acetylcholine. Furthermore, concentrations of the inflammatory cytokines IL-4 and IL-5 were measured in bronchoalveolar lavage fluid. The results demonstrate significant anti-inflammatory effects of Provinol and Flavin-7 exerted in the airways. In contrast, chronic treatment with quercetin and resveratrol, single components of the two polyphenols, did not show such activity. We conclude that polyphenolic compounds are more effective in the anti-inflammatory effects in the airways than their separate components.

Topics: Acetylcholine; Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Bronchoconstrictor Agents; Guinea Pigs; Histamine; Interleukin-4; Interleukin-5; Lung; Muscle, Smooth; Ovalbumin; Plethysmography, Whole Body; Polyphenols; Quercetin; Respiratory System; Resveratrol; Stilbenes; Trachea

Sphingosine-1-phosphate (S1P), which is produced by 2 sphingosine kinase (SphK) isoenzymes, SphK1 and SphK2, has been implicated in IgE-mediated mast cell responses. However, studies of allergic inflammation in isotype-specific SphK knockout mice have not clarified their contribution, and the role that S1P plays in vivo in a mast cell- and IgE-dependent murine model of allergic asthma has not yet been examined.. We used an isoenzyme-specific SphK1 inhibitor, SK1-I, to investigate the contributions of S1P and SphK1 to mast cell-dependent airway hyperresponsiveness (AHR) and airway inflammation in mice.. Allergic airway inflammation and AHR were examined in a mast cell-dependent murine model of ovalbumin (OVA)-induced asthma. C57BL/6 mice received intranasal delivery of SK1-I before sensitization and challenge with OVA or only before challenge.. SK1-I inhibited antigen-dependent activation of human and murine mast cells and suppressed activation of nuclear factor κB (NF-κB), a master transcription factor that regulates the expression of proinflammatory cytokines. SK1-I treatment of mice sensitized to OVA in the absence of adjuvant, in which mast cell-dependent allergic inflammation develops, significantly reduced OVA-induced AHR to methacholine; decreased numbers of eosinophils and levels of the cytokines IL-4, IL-5, IL-6, IL-13, IFN-γ, and TNF-α and the chemokines eotaxin and CCL2 in bronchoalveolar lavage fluid; and decreased pulmonary inflammation, as well as activation of NF-κB in the lungs.. S1P and SphK1 play important roles in mast cell-dependent, OVA-induced allergic inflammation and AHR, in part by regulating the NF-κB pathway.

Topics: Amino Alcohols; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Chemokine CCL2; Female; Goblet Cells; Humans; Hyperplasia; Immunoglobulin E; Inflammation; Interferon-gamma; Interleukins; Lung; Lysophospholipids; Mast Cells; Methacholine Chloride; Mice; Mice, Inbred C57BL; NF-kappa B; Ovalbumin; Phosphotransferases (Alcohol Group Acceptor); Sphingosine; Tumor Necrosis Factor-alpha

Asthma is a chronic inflammatory disease characterized by airways hyperresponsiveness (AHR), reversible airflow obstruction, airway remodeling, and episodic exacerbations caused by air pollutants, such as particulate matter (PM; PM <2.5 μm in diameter [PM(2.5)]) and ozone (O(3)). Spleen tyrosine kinase (Syk), an immunoregulatory kinase, has been implicated in the pathogenesis of asthma.. We sought to evaluate the effect of Syk inhibition on AHR in a chronic mouse model of allergic airways inflammation and pollutant exposure.. We used a 12-week chronic ovalbumin (OVA) sensitization and challenge mouse model of airways inflammation followed by exposure to PM(2.5) plus O(3). Respiratory mechanics and methacholine (MCh) responsiveness were assessed by using the flexiVent system. The Syk inhibitor NVP-QAB-205 was nebulized intratracheally by using a treatment-based protocol 15 minutes before assessment of MCh responsiveness.. Syk expression increased significantly in the airway epithelia of OVA-sensitized and OVA-challenged (OVA/OVA) mice compared with OVA-sensitized but PBS-challenged (OVA/PBS) control mice. OVA/OVA mice exhibited AHR to MCh, which was attenuated by a single administration of NVP-QAB-205 (0.3 and 3 mg/kg). PM(2.5) plus O(3) significantly augmented AHR to MCh in the OVA/OVA mice, which was abrogated by NVP-QAB-205. Total inflammatory cell counts were significantly higher in the bronchoalveolar lavage fluid from OVA/OVA than OVA/PBS mice and were unaffected by PM(2.5) plus O(3) or NVP-QAB-205.. NVP-QAB-205 reduced AHR and the enhanced response to PM(2.5) plus O(3) to normal levels in an established model of chronic allergic airways inflammation, suggesting that Syk inhibitors have promise as a therapy for asthma.

Topics: Air Pollutants; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Disease Models, Animal; Female; Humans; Inflammation; Intracellular Signaling Peptides and Proteins; Keratinocytes; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Respiratory Mucosa; Syk Kinase; Vascular Endothelial Growth Factor A

Asthma is a chronic airway inflammatory disease characterized by eosinophilic infiltration and airway hyperresponsiveness. The over-activated Th2 and lung epithelium cells express many different cytokines, and chemokines mainly contribute to the severity of lung inflammation. Clara cell 10 kD protein (CC10) is highly expressed in airway epithelium cells and exhibits anti-inflammatory and immunomodulatory effects. Adeno-associated virus (AAV) 2/9 vector, composed of AAV2 rep and AAV9 cap genes, can efficiently and specifically target lung epithelium cells. Thus, AAV2/9 vector might carry therapeutic potential gene sequences for the treatment of asthma. This study tested whether AAV2/9 vector carrying CC10 could reduce inflammatory and asthmatic responses in OVA-induced asthmatic mouse model. The results showed that AAV2/9-CC10 vector virus significantly reduced airway hyperresponsiveness, CCL11, interleukin (IL)-4, IL-5, IL-6, IL-13, and eosinophilia in the lungs of sensitized mice. CC10 level in OVA-sensitized mice was rescued with the administration of AAV2/9-CC10 vector virus. Lung tissue remodeling, including collagen deposition and goblet cell hyperplasia, was also alleviated. However, serum levels of OVA-specific IgG1 and IgE as well as Th2 cytokine levels in OVA-stimulated splenocyte culture supernatants were at the comparable levels to the sensitized control group. The results demonstrate that AAV2/9-CC10 vector virus relieved local inflammatory and asthmatic responses in lung. Therefore, we propose that AAV2/9-CC10 vector virus guaranteed sufficient CC10 expression and had an anti-inflammatory effect in asthmatic mice. It might be applied as a novel therapeutic approach for asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dependovirus; DNA Primers; Genetic Therapy; Genetic Vectors; Immunoglobulin E; Immunoglobulin G; Immunohistochemistry; Interleukins; Lung; Mice; NIH 3T3 Cells; Ovalbumin; Treatment Outcome; Uteroglobin

Nonspecific airway hyperresponsiveness (AHR) is one of the cardinal features of bronchial asthma. Early AHR is caused by chemical mediators released from pulmonary mast cells activated in an IgE-dependent way. However, the mechanism of late AHR remains unclear.. Features of airway allergic inflammation were analyzed, including antigen-induced AHR, using a murine model of asthma. The model was suitable for examining the sequential early molecular events occurring after the initial airway exposure to antigen.. AHR increased at 10-12 h after airway challenge, followed by the second-phase response, which was larger and broader in resistance at 18-30 h. Pretreatment of sensitized animals with anti-tumor necrosis factor (TNF) before airway challenge or induction of allergic asthma in TNF(-/-) mice resulted in abrogation of the first-phase late AHR. Intratracheal instillation of TNF induced a single peak of AHR at 10 h. IgE and IgG immune complexes induced the development of the first-phase late AHR by TNF production. Pretreatment with cytosolic phospholipase inhibitor and 5-lipoxygenase inhibitors abolished the first-phase late AHR as well as the leukotriene B(4) levels in the airway. CpG-oligodeoxynucleotide (ODN) pretreatment reduced airway levels of Th2 cytokines, eosinophil infiltration and second-phase late AHR. However, CpG-ODN did not reduce TNF levels or the magnitude of first-phase late AHR.. Biphasic late AHR occurs in a murine model of asthma. First- and second-phase late AHR is caused by TNF and Th2 response, respectively.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Immunoblotting; Inflammation; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Th2 Cells; Tumor Necrosis Factor-alpha

Airway epithelial injury is the hallmark of various respiratory diseases and therapeutic targeting of epithelial injury could be an effective strategy for controlling these diseases. We recently reported that a novel cinnamate, ethyl 3',4',5'-trimethoxythionocinnamate (ETMTC) derived from Piper longum derivative, was most potent among various cinnamate derivatives in inhibiting inflammatory cell adhesion molecules (CAMs). In this study, we investigated the effects of ETMTC on the features of allergic asthma and epithelial injury in a murine model. ETMTC treatment to ovalbumin sensitized and challenged mice during ovalbumin challenge reduced airway hyperresponsiveness, and airway inflammation. This attenuation of asthma features was associated with the reduction in the expressions of various CAMs, NF-κB activation, Th2 cytokines, eotaxin and 8-isoprostane that were estimated in lung homogenates. Further, it increased activities of mitochondrial complexes I and IV in lung mitochondria and reduced cytochrome c and caspase 9 activities in lung cytosol. In addition, it reduced the levels of oxidative DNA damage marker in bronchoalveolar lavage fluid and DNA fragmentation of bronchial epithelia in lung sections. Further, ETMTC not only increased the levels of 15-(S)-hydroxyeicosatetraenoic acid, suppressor of airway remodeling, but also inhibited goblet cell metaplasia and sub-epithelial fibrosis. These results demonstrate that ETMTC reduces epithelial injury and mitochondrial dysfunction associated with allergic asthma and thus ETMTC could be useful to develop efficient therapeutic molecule against asthma.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Asthma; Bronchi; Bronchial Hyperreactivity; Cinnamates; Cytokines; DNA Damage; Immunoglobulin E; Immunoglobulin G; Male; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Respiratory Mucosa

Asthma is characterized as a chronic inflammatory disorder of the airways associated with an enhanced TH2 response to inhaled allergens. CD4+ T regulatory (Treg) cells are controlled by the master transcription factor FoxP3 and strictly maintain peripheral immunotolerance. Epigenetic regulation of FoxP3 by DNA methyltransferase inhibitors, such as 5-azacytidine (Aza), can generate a steady supply of functional Treg cells. Therefore, we propose that Aza can augment Treg cells in vivo to prevent the pathogenesis of asthma.. BALB/c mice were sensitized with chicken ovalbumin (OVA) and treated with different doses of Aza. Airway hyperresponsiveness to methacholine, eosinophilia in bronchoalveolar lavage fluid, circulating titers of OVA-specific IgG1 and IgE, and stimulating levels of TH2 cytokines from splenocytes were then determined. Cellular populations were examined by flow cytometry. PC61 antibody, which depletes CD25+ cells, was used to verify the role of CD25+ cells in Aza-induced tolerance.. Administration of Aza to OVA-sensitized mice diminished airway hyperreactivity, pulmonary eosinophilia, levels of OVA-specific IgG1 and IgE in serum, and secretion of TH2 cytokines from OVA-stimulated splenocytes in a dose-dependent manner. Percentages of CD25+ and FoxP3+ cells in the CD4+ cell population were notably increased in Aza-treated mice compared to sensitized control mice. Furthermore, the major symptoms of asthma were exacerbated by depleting CD25+ cells in Aza-treated mice.. Aza may have applications as a novel clinical strategy to increase the production of Treg cells in order to modulate the airway inflammation associated with asthma.

Topics: Animals; Asthma; Azacitidine; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4 Antigens; DNA Methylation; Eosinophilia; Forkhead Transcription Factors; Immunoglobulin E; Immunoglobulin G; Interleukin-13; Interleukin-2 Receptor alpha Subunit; Interleukin-4; Interleukin-5; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory System; Spleen; T-Lymphocytes, Regulatory

Astragalus membranaceus (AM), a traditional Chinese medicinal herb, has been widely used for centuries to treat asthma in China. Previous studies demonstrated that AM had inhibitory effects on airway hyperresponsiveness, inflammation and airway remodeling in murine models of asthma. However, it remained unclear whether the beneficial effects of AM on asthma were associated with CD4(+)CD25(+)Foxp3(+) Treg cells; this issue is the focus of the present work. An asthma model was established in Sprague-Dawley (SD) rats that were sensitized and challenged with ovalbumin. Bronchoalveolar lavage fluid (BALF) was assessed for inflammatory cell counts and cytokine levels. Airway hyperresponsiveness was detected by direct airway resistance analysis. Lung tissues were examined for cell infiltration, mucus hypersecretion and airway remodeling. CD4(+)CD25(+)Foxp3(+) Treg cells in the BALF and Foxp3 mRNA expression in lung tissues were examined. The oral administration of AM significantly reduced airway hyperresponsiveness to aerosolized methacholine and inhibited eosinophil counts and reduced IL-4, IL-5 and IL-13 levels and increased INF-γ levels in the BALF. Histological studies showed that AM markedly decreased inflammatory infiltration, mucus secretion and collagen deposition in the lung tissues. Notably, AM significantly increased population of CD4(+)CD25(+)Foxp3(+) Treg cells and promoted Foxp3(+) mRNA expression in a rat model of asthma. Together, these results suggest that the antiasthmatic effects of AM are at least partially associated with CD4(+)CD25(+)Foxp3(+) Tregs.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Astragalus propinquus; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Forkhead Transcription Factors; Male; Methacholine Chloride; Ovalbumin; Phytotherapy; Plant Extracts; Rats; Rats, Sprague-Dawley; RNA, Messenger; T-Lymphocytes, Regulatory

Although the contribution of basophils as inducers or amplifiers of Th2 responses is still debated, prolonged basophil/CD4 T cell interactions were observed in lungs but not lymph nodes (LNs) of parasite-infected mice. However, the impact of basophils on the function of tissue CD4 effector T cells remains unknown.. Basophils were purified from the lungs of ovalbumin (OVA)-sensitized and OVA-challenged (OVA-immunized) mice or human peripheral blood for in vivo and in vitro functional studies. Pulmonary basophils were adoptively transferred to OVA-sensitized hosts to assess airway inflammation in bronchoalveolar lavage fluid (BALF) and Th2 responses in lung explants and draining LNs. Basophils were co-cultured with effector T cells or Ag-specific naïve T cells alone or in combination with dendritic cells (DCs); IL-4 production was determined by flow cytometry and ELISA.. Basophils accumulated in lungs of OVA-immunized mice. Adoptive transfer of basophils to OVA-sensitized hosts enhanced lung IL-4 and IL-13 release while co-administration of OVA further aggravated airway inflammation and Th2 responses in LNs. Mechanistic in vitro studies revealed that pulmonary basophils interacted with lung CD4 effectors, in the absence of DCs, to increase T cell survival and Th2 cytokine expression at the single cell level but amplified OVA-loaded DC-driven Th2 differentiation. Finally, human basophils augmented in vitro IL-4 expression in effector memory CD4 T cells that include CRTH2(+) cells through IL-4 and TCR-independent pathways.. Basophils may worsen Th2 inflammatory disorders through direct interactions with pathogenic CD4 T cells as well as by enhancing DC-induced Th2 cell development.

Topics: Adoptive Transfer; Animals; Asthma; Basophils; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cells, Cultured; Coculture Techniques; Cytokines; Disease Models, Animal; Female; Flow Cytometry; Humans; Immunity, Innate; Immunoglobulin E; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Ovalbumin; Random Allocation

Since CD4+ T cells play a pivotal role in the development of airway inflammation and hyperresponsiveness, targeting activated CD4+ T cell subsets and increasing the cells with regulatory function would be a logical therapeutic approach. We showed that this outcome can be achieved by local therapy with Tim-3, which is a negative regulator of CD4+ T cells. Tim-3 expression was up-regulated by ovalbumin (OVA) induction. Attenuating Tim-3 expression by RNA interference suppressed allergen-induced immune responses. Intranasal application of Tim-3 shRNA diminished airway inflammation and hyperresponsiveness. Multiple mechanisms were involved in the inhibitory effects, including regulation the imbalance of Th1/Th17 and increasing Treg cell expression. Our results indicate that the Tim-3 pathway is highly involved in the regulation of asthma. Targeting Tim-3 by siRNA may hold therapeutic potential in preventing the development of allergic asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Female; Hepatitis A Virus Cellular Receptor 2; Inflammation; Leukocytes, Mononuclear; Mice; Mice, Inbred BALB C; Ovalbumin; Receptors, Virus; Respiratory Hypersensitivity; RNA Interference; RNA, Small Interfering; T-Lymphocytes, Regulatory; Th1 Cells; Th17 Cells

Asthma is characterized by airway hyperresponsiveness and remodeling. Pravastatin and atorvastatin are used clinically as cholesterol-lowering agents but also exhibit anti-inflammatory and immunomodulating properties.. To investigate the therapeutic effect of oral statins on airway hyperresponsiveness and allergic reaction.. BALB/c mice received intraperitoneal sensitization and aerosol inhalation with ovalbumin consequently. One week after ovalbumin aerosol challenge, pravastatin, atorvastatin, or phosphate-buffered saline were given by intragastric gavage daily for 2 weeks. Airway hyperresponsiveness, serum allergen specific antibody levels, cytokine production by splenocytes, and bronchoalveolar lavage fluid were examined.. Both pravastatin and atorvastatin effectively reduced airway hyperresponsiveness. Pravastatin effectively suppressed both T(H)1- and T(H)2-mediated antibody responses, reducing serum specific IgE, IgG, IgG1, and IgG2a levels. Pravastatin also effectively reduced interleukin (IL) 4, IL-5, and interferon γ production but significantly enhanced IL-10 levels in splenocytes and BALF. Similarly, atorvastatin effectively attenuated production of specific IgE, IgG1, and IgG2a antibodies. It also significantly attenuated IL-4, interferon γ, and increased IL-10 concentration in bronchoalveolar lavage fluid and splenocytes.. Oral administration of pravastatin or atorvastatin not only was able to inhibit T(H)1 inflammatory responses but also had therapeutic effects on airway hyperresponsiveness and T(H)2 allergic responses. These results seem to suggest that these drugs have potential as a nonimmunosuppressive therapy for asthma and allergic diseases.

Topics: Administration, Oral; Animals; Asthma; Atorvastatin; Bronchial Hyperreactivity; Cytokines; Female; Heptanoic Acids; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Hypersensitivity; Immunoglobulin G; Mice; Mice, Inbred BALB C; Ovalbumin; Pravastatin; Pyrroles; Rats, Sprague-Dawley; Th1 Cells

Airway hyperreactivity (AHR) is a key feature of bronchial asthma, and inhalation of irritants may facilitate development of nonallergic AHR. Swimmers exposed to hypochlorite (ClO(-))-containing water show a higher risk of developing AHR. We developed a mouse model in which instillation of ClO(-) before ovalbumin (OVA) induces AHR without bronchial inflammatory cells.. To investigate the mechanisms of ClO(-)-OVA-induced nonallergic AHR.. The involvement of the transient receptor potential ankyrin (TRPA)1 channel was checked in vivo by the use of TRPA1(-/-) mice and in vitro by Ca(2+) imaging experiments. The role of substance P (SP) was investigated by pretreating animals with the receptor antagonist RP67580, by replacing ClO(-) with SP in vivo, and by immunofluorescent staining of large airways of exposed mice. The role of mast cells was evaluated by exposing mast cell-deficient Kit(Wh)/Kit(Wsh) mice to ClO(-)-OVA with or without mast cell reconstitution.. ClO(-)-OVA did not induce AHR in TRPA1(-/-) mice, and ClO(-) generates a Ca(2+) influx in TRPA1-transfected cells. Pretreatment with RP67580 reduces ClO(-)-OVA-induced AHR, although no increased SP expression was shown in the airways. SP-OVA exposure resulted in the same AHR as induced by ClO(-)-OVA. Kit(Wsh)/Kit(Wsh) mice did not develop AHR in response to ClO(-)-OVA unless they were reconstituted with bone marrow-derived mast cells.. Induction of AHR by exposure to ClO(-)-OVA depends on a neuroimmune interaction that involves TRPA1-dependent stimulation of sensory neurons and mast cell activation.

Topics: Animals; Bronchial Hyperreactivity; Cells, Cultured; Hypochlorous Acid; Irritants; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Knockout; Neuroimmunomodulation; Nociceptors; Ovalbumin; Substance P; Transient Receptor Potential Channels; TRPA1 Cation Channel

Astragalus membranaceus from traditional Chinese herbal medicines previously showed that it possesses a strong anti-inflammatory activity. The purpose of this study was to elucidate the effect of astragalus on allergen-induced airway inflammation and airway hyperresponsiveness and investigate its possible molecular mechanisms.. Female BALB/c mice sensitized and challenged with ovalbumin (OVA) developed airway inflammation. Bronchoalveolar lavage fluid was assessed for total and differential cell counts and cytokine and chemokine levels. In vivo airway responsiveness to increasing concentrations of methacholine was measured 24 hours after the last OVA challenge using whole-body plethysmography. The expression of inhibitory κB-α and p65 in lung tissues was measured by Western blotting.. Astragalus extract attenuated lung inflammation, goblet cell hyperplasia and airway hyperresponsiveness in OVA-induced asthma and decreased eosinophils and lymphocytes in bronchoalveolar lavage fluid. In addition, astragalus extract treatment reduced expression of the key initiators of allergic T(H)2-associated cytokines (interleukin 4, interleukin 5) (P < 0.05). Furthermore, astragalus extract could inhibit nuclear factor κB (NF-κB) expression and suppress NF-κB translocation from the cytoplasm to the nucleus in lung tissue samples.. Taken together, our current study demonstrated a potential therapeutic value of astragalus extract in the treatment of asthma and it may act by inhibiting the expression of the NF-κB pathway.

Topics: Animals; Asthma; Astragalus Plant; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Disease Models, Animal; Female; Hyperplasia; Lung; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Plant Extracts; Plethysmography; Pneumonia; Signal Transduction

The effect of increased age on the induction of oral tolerance by low-dose antigen feeding and its effect on the response to antigen airway challenge in aged mice have not been well characterized.. To determine whether oral tolerance can be induced in aged mice and its impact on the development of allergic airway inflammation.. Younger (6 weeks old) and aged (18 months old) mice were fed ovalbumin (OVA) prior to sensitization to induce antigen tolerance. Serum antigen-specific immunoglobulins (Igs), bronchoalveolar lavage fluid (BALF), lung histology, enumeration of CD4 + Foxp3+ Treg cells, and airway hyperresponsiveness (AHR) were determined after the final antigen challenge.. Feeding antigen to aged mice prior to sensitization induced oral tolerance as determined by a decrease in antigen-specific IgE and IgG(1); however, the effect was greater in younger mice. Induction of oral tolerance was associated with a greater increase in airway Treg cells in the younger mice. Despite these differences, oral tolerance significantly suppressed features of asthma in aged mice, including BALF total cell and eosinophil numbers, cytokine production, and AHR.. Aged mice developed oral tolerance to antigen, which suppressed several features of allergic airway inflammation.

Topics: Administration, Oral; Age Factors; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Female; Histocytochemistry; Immune Tolerance; Immunoglobulin E; Immunoglobulin G; Mice; Mice, Inbred BALB C; Ovalbumin; Statistics, Nonparametric; T-Lymphocytes, Regulatory

To investigate the effect of Tripterygium polyglycosid on establishing airway eosinophil infiltration and related airway hyperresponsiveness of asthmatic mice.. A mature murine asthmatic model was made with ovabulmin sensitized and challenged C57BL/6 mice. Forty mice were divided into four groups with 10 mice in each group: mice sensitized and challenged with saline (WS group), mice sensitized and challenged with ovalbumin (WO group), mice sensitized and challenged with ovalbumin and treated with Tripterygium polyglycosid (TP group) and Dexamethasone (DXM group). The mice were intraperitoneally injected with 20 μg chicken ovabulmin emulsified in injected alum on days 0 and 14, then were challenged with an aerosol generated from 1% ovabulmin on days 24, 25 and 26. Tripterygium polyglycosid was injected intraperitoneally at 50 mg/kg on days 25, 26 and 27 after ovabulmin challenge. Dexamethasone was administrated to mice at 2 mg/kg on day 21, 23 before ovabulmin challenge. The airway hyperresponsiveness, mucus production, eosinophils in parabronchial area and bronchoalveolar lavage fluid and the level of interleukin-5, granulo-macrophage clone stimulating factor in bronchoalveolar lavage fluid were measured as indexes of inflammation.. Tripterygium polyglycosid treatment after ovabulmin challenge completely inhibited eosinophil infiltration in bronchoalveolar lavage fluid [(0.63 ± 0.34)× 10(4) vs. (75.0 ± 14.8)× 10(4), P<0.05] and the peribrochial area (12.60 ± 3.48 mm(2) vs. 379.0 ± 119.3 mm(2), P<0.05), mucus overproduction in airway (2.8 ± 1.7 vs. 7.1±5.6, P<0.05), and increased interleukin-5 levels in bronchoalveolar lavage fluid (28.8 ± 2.8 pg/mL vs. 7.5 ± 3.5 pg/mL, P<0.05). Meanwhile, Tripterygium polyglycosid treatment after ovabulmin challenge also partially inhibited airway hyperresponsiveness. The level of granulo-macrophage clone stimulating factor in bronchoalveolar lavage fluid didn't change with drugs intervention.. The administration of Tripterygium polyglycosid could inhibit the established airway inflammation and reduce the airway hyperresponsiveness of allergic asthmatic mice. It provides a possible alternative therapeutic for asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Dexamethasone; Drugs, Chinese Herbal; Eosinophils; Lung; Mice; Mice, Inbred C57BL; Mucus; Ovalbumin; Plant Extracts; Pneumonia; Tripterygium

Chronic asthma is often associated with neutrophilic infiltration in the airways. Neutrophils contain elastase, a potent secretagogue in the airways, nonetheless the role for neutrophil elastase as well as neutrophilic inflammation in allergen-induced airway responses is not well defined. In this study, we have investigated the impact of neutrophil elastase inhibition on the development of allergic airway inflammation and airway hyperresponsiveness (AHR) in previously sensitized and challenged mice.. BALB/c mice were sensitized and challenged (primary) with ovalbumin (OVA). Six weeks later, a single OVA aerosol (secondary challenge) was delivered and airway inflammation and airway responses were monitored 6 and 48 hrs later. An inhibitor of neutrophil elastase was administered prior to secondary challenge.. Mice developed a two-phase airway inflammatory response after secondary allergen challenge, one neutrophilic at 6 hr and the other eosinophilic, at 48 hr. PAR-2 expression in the lung tissues was enhanced following secondary challenge, and that PAR-2 intracellular expression on peribronchial lymph node (PBLN) T cells was also increased following allergen challenge of sensitized mice. Inhibition of neutrophil elastase significantly attenuated AHR, goblet cell metaplasia, and inflammatory cell accumulation in the airways following secondary OVA challenge. Levels of IL-4, IL-5 and IL-13, and eotaxin in BAL fluid 6 hr after secondary allergen challenge were significantly suppressed by the treatment. At 48 hr, treatment with the neutrophil elastase inhibitor significantly reduced the levels of IL-13 and TGF-β1 in the BAL fluid. In parallel, in vitro IL-13 production was significantly inhibited in spleen cells from sensitized mice.. These data indicate that neutrophil elastase plays an important role in the development of allergic airway inflammation and hyperresponsiveness, and would suggest that the neutrophil elastase inhibitor reduced AHR to inhaled methacholine indicating the potential for its use as a modulator of the immune/inflammatory response in both the neutrophil- and eosinophil-dominant phases of the response to secondary allergen challenge.

Topics: Allergens; Animals; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Chemokines; Cytokines; Female; Flow Cytometry; Glycine; Immunohistochemistry; Inflammation; Intercellular Signaling Peptides and Proteins; Leukocyte Elastase; Mice; Mice, Inbred BALB C; Neutrophil Infiltration; Ovalbumin; Proteinase Inhibitory Proteins, Secretory; Receptor, PAR-2; Respiratory Hypersensitivity; Sulfonamides

Cystathionine γ-lyase (CSE) is one of the major enzymes producing hydrogen sulfide (H2S) in lungs, participating in the regulation of respiratory functions. The role of CSE-derived H2S in eosinophil-dominant inflammation in allergic diseases has been unclear. The objective of this study was to explore the protective role of H2S against allergen-induced airway hyperresponsiveness (AHR) and inflammation. CSE expression and H2S production rate were assessed in mouse lung tissues with ovalbumin (OVA)-induced acute asthma. AHR, airway inflammation, and Th2 response in wild-type (WT) mice were compared with those in CSE gene knockout (KO) mice. The effect of NaHS, an exogenous H2S donor, was also evaluated on these parameters. CSE expression was absent and H2S production rate was significantly lower in the lungs of CSE KO mice when compared with WT littermates. OVA challenge decreased lung CSE expression and H2S production in WT mice. CSE deficiency resulted in aggravated AHR, increased airway inflammation, and elevated levels of Th2 cytokines such as IL-5, IL-13, and eotaxin-1 in bronchoalveolar lavage fluid after OVA challenge. The aforementioned alterations were reversed by exogenous H2S treatment. More importantly, NaHS supplement rescued CSE KO mice from the aggravated pathological process of asthma. The CSE/H2S system plays a critical protective role in the development of asthma. A new therapeutic potential for asthma via targeting CSE/H2S metabolism is indicated.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Survival; Cystathionine gamma-Lyase; Cytokines; Disease Models, Animal; Inflammation; Lung; Lymphocytes; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Sulfides; Th2 Cells

Hesperidin, a flavanone glycoside comprised of the flavanone hesperetin and the disaccharide rutinose, is a plentiful and inexpensive by-product of citrus cultivation. It has been reported to exert a wide range of pharmacological effects that include antioxidant, anti-inflammatory, and anticarcinogenic properties. In this study, we attempt to determine whether hesperidin inhibits inflammatory mediators in the mouse allergic asthma model. Mice were sensitized and challenged by ovalbumin (OVA) to induce chronic airway inflammation and airway remodeling. The administration of hesperidin significantly decreased the number of infiltrating inflammatory cells and Th2 cytokines in bronchoalveolar lavage (BAL) fluid compared with the OVA-induced group of mice. In addition, hesperidin reduced OVA-specific IgE levels in serum. Hesperidin markedly alleviated the OVA-induced airway hyperresponsiveness (AHR) to inhaled methacholine. Based on lung histopathological studies using hematoxylin and eosin and alcian blue-periodic acid-Schiff staining, hesperidin inhibited inflammatory cell infiltration and mucus hypersecretion compared with the OVA-induced group of mice. These findings provide new insight into the immunopharmacological role of hesperidin in terms of its effects in a murine model of asthma.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Hesperidin; Immunoglobulin E; Inflammation; Interleukin-13; Interleukin-4; Interleukin-5; Leukocytes; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory System; Th2 Cells

Aqueous extract of Kalanchoe pinnata (Kp) have been found effective in models to reduce acute anaphylactic reactions. In the present study, we investigate the effect of Kp and the flavonoid quercetin (QE) and quercitrin (QI) on mast cell activation in vitro and in a model of allergic airway disease in vivo. Treatment with Kp and QE in vitro inhibited degranulation and cytokine production of bone marrow-derived mast cells following IgE/FcɛRI crosslinking, whereas treatment with QI had no effect. Similarly, in vivo treatment with Kp and QE decreased development of airway hyperresponsiveness, airway inflammation, goblet cell metaplasia and production of IL-5, IL-13 and TNF. In contrast, treatment with QI had no effect on these parameters. These findings demonstrate that treatment with Kp or QE is effective in treatment of allergic airway disease, providing new insights to the immunomodulatory functions of this plant.

Topics: Animals; Basophil Degranulation Test; Bronchial Hyperreactivity; Goblet Cells; Interleukin-13; Interleukin-5; Kalanchoe; Mast Cells; Metaplasia; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plant Extracts; Quercetin; Tumor Necrosis Factor-alpha

Asthma is characterized by reversible bronchoconstriction and airway hyperreactivity. Although M(3) muscarinic receptors mediate bronchoconstriction, non-selective muscarinic receptor antagonists are not currently recommended for chronic control of asthma. We tested whether selective blockade of M(3) receptors, at the time of antigen challenge, blocks subsequent development of airway hyperreactivity in antigen-challenged guinea-pigs.. Ovalbumin-sensitized guinea-pigs were pretreated with 1 µg·kg(-1) of a kinetically selective M(3) receptor antagonist, tiotropium, or 1 mg·kg(-1) of a non-selective muscarinic receptor antagonist, atropine, and challenged with inhaled ovalbumin. Animals were anaesthetized, paralyzed, ventilated and vagotomized 24 h later. We measured vagally mediated bronchoconstriction and i.v. ACh-induced bronchoconstriction.. Electrical stimulation of both vagus nerves induced frequency-dependent bronchoconstriction in sensitized animals that was significantly increased after antigen challenge. Antigen-induced hyperreactivity was completely blocked by tiotropium pretreatment but only partially blocked by atropine pretreatment. Surprisingly, although tiotropium blocked bronchoconstriction induced by i.v. ACh, it did not inhibit vagally-induced bronchoconstriction in sensitized controls, suggesting that tiotropium does not block hyperreactivity by blocking receptors for vagally released ACh. Rather, tiotropium may have worked through an anti-inflammatory mechanism, since it inhibited eosinophil accumulation in the lungs and around nerves.. These data confirm that testing M(3) receptor blockade with exogenous ACh does not predict vagal blockade. Our data also suggest that selective blockade of M(3) receptors may be effective in asthma via mechanisms that are separate from inhibition of bronchoconstriction.

Topics: Acetylcholine; Animals; Asthma; Atropine; Bronchial Hyperreactivity; Bronchoconstriction; Disease Models, Animal; Eosinophils; Female; Guinea Pigs; Inflammation; Ovalbumin; Receptor, Muscarinic M3; Scopolamine Derivatives; Tiotropium Bromide; Vagus Nerve

IL-13 is a pleiotropic Th2 cytokine considered likely to play a pivotal role in asthma. Here we describe the preclinical in vitro and in vivo characterization of CAT-354, an IL-13-neutralizing IgG4 monoclonal antibody (mAb), currently in clinical development.. In vitro the potency, specificity and species selectivity of CAT-354 was assayed in TF-1 cells, human umbilical vein endothelial cells and HDLM-2 cells. The ability of CAT-354 to modulate disease-relevant mechanisms was tested in human cells measuring bronchial smooth muscle calcium flux induced by histamine, eotaxin generation by normal lung fibroblasts, CD23 upregulation in peripheral blood mononuclear cells and IgE production by B cells. In vivo CAT-354 was tested on human IL-13-induced air pouch inflammation in mice, ovalbumin-sensitization and challenge in IL-13 humanized mice and antigen challenge in cynomolgus monkeys.. CAT-354 has a 165 pM affinity for human IL-13 and functionally neutralized human, human variant associated with asthma and atopy (R130Q) and cynomolgus monkey, but not mouse, IL-13. CAT-354 did not neutralize human IL-4. In vitro CAT-354 functionally inhibited IL-13-induced eotaxin production, an analogue of smooth muscle airways hyperresponsiveness, CD23 upregulation and IgE production. In vivo in humanized mouse and cynomolgus monkey antigen challenge models CAT-354 inhibited airways hyperresponsiveness and bronchoalveolar lavage eosinophilia.. CAT-354 is a potent and selective IL-13-neutralizing IgG4 mAb. The preclinical data presented here support the trialling of this mAb in patients with moderate to severe uncontrolled asthma.

Topics: Adolescent; Animals; Antibodies, Monoclonal; Antigens; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Line, Tumor; Disease Models, Animal; Female; Human Umbilical Vein Endothelial Cells; Humans; Immunoglobulin E; Inflammation; Interleukin-13; Macaca fascicularis; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Receptors, IgE; Severity of Illness Index; Species Specificity; Up-Regulation

Experimental asthma increases eosinophil and collagen deposition in the lungs of sickle cell disease (SCD) mice to a greater extent than in control mice. However, the effects of asthma on inflammation and airway physiology remain unclear. To determine effects of asthma on pulmonary inflammation and airway mechanics in SCD mice, hematopoietic stem cell transplantation was used to generate chimeric SCD and hemoglobin A mice. Experimental asthma was induced by sensitizing mice with ovalbumin (OVA). Airway mechanics were assessed using forced oscillation techniques. Mouse lungs were examined histologically and physiologically. Cytokine, chemokine, and growth factors in bronchoalveolar lavage fluid were determined by multiplex. IgE was quantified by ELISA. LDH was quantified using a colorimetric enzymatic assay. At baseline (nonsensitized), chimeric SCD mice developed hemolytic anemia with sickled red blood cells, mild leukocytosis, and increased vascular endothelial growth factor and IL-13 compared with chimeric hemoglobin A mice. Experimental asthma increased perialveolar eosinophils, plasma IgE, and bronchoalveolar lavage fluid IL-1β, IL-4, IL-6, and monocyte chemotactic protein 1 in chimeric hemoglobin A and SCD mice. IFN-γ levels were reduced in both groups. IL-5 was preferentially increased in chimeric SCD mice but not in hemoglobin A mice. Positive end-expiratory pressures and methacholine studies revealed that chimeric SCD mice had greater resistance in large and small airways compared with hemoglobin A mice at baseline and after OVA sensitization. SCD alone induces a baseline lung pathology that increases large and small airway resistance and primes the lungs to increased inflammation and airway hyperresponsiveness after OVA sensitization.

Topics: Airway Resistance; Anemia, Sickle Cell; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Colorimetry; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophils; Hemoglobin A; Hemoglobin, Sickle; Humans; Immunoglobulin E; Inflammation Mediators; L-Lactate Dehydrogenase; Lung; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Transgenic; Ovalbumin; Pneumonia; Positive-Pressure Respiration; Vascular Endothelial Growth Factor A

ASP3258 is a potent and selective PDE4 inhibitor and exerts a wide-range of anti-inflammatory effects with low emetic potential, a major adverse effect of PDE4 inhibitors. Here, we investigated the anti-asthmatic potency of ASP3258 as compared with those of two representative PDE4 inhibitors: roflumilast and cilomilast. Orally administered ASP3258, roflumilast, and cilomilast all inhibited ovalbumin (OVA)-induced eosinophil infiltration into the airway of sensitized Brown Norway rats with ED(50) values of 0.81, 0.46, and 4.4 mg/kg, respectively. Histological examination also revealed a decreasing trend in inflammatory cell infiltration into the lung following ASP3258 administration. In vitro investigation of bronchodilatory activities showed that these compounds (10(-8)-10(-6) M) concentration-dependently inhibited OVA-induced contraction of trachea isolated from sensitized guinea pigs but had no effect on spasmogen-precontracted tracheal tension prepared from non-sensitized guinea pigs up to 10(-6) M. In vivo experiments using sensitized guinea pigs showed that these orally administered compounds inhibited OVA-induced increases in airway resistance with ED(50) values of 2.2, 0.35, and 12 mg/kg, respectively. Further, orally administered ASP3258 (0.1 and 1 mg/kg), roflumilast (0.1 and 1 mg/kg), and cilomilast (10 mg/kg) significantly suppressed airway hyperresponsiveness caused by OVA exposure. ASP3258's potent inhibition of antigen-induced bronchoconstriction and airway hyperresponsiveness, two characteristic symptoms of bronchial asthma, suggests that this compound will be useful in treating asthma.

Topics: Aminopyridines; Animals; Anti-Asthmatic Agents; Benzamides; Bronchial Hyperreactivity; Bronchoconstriction; Cyclohexanecarboxylic Acids; Cyclopropanes; Disease Models, Animal; Eosinophils; Female; Guinea Pigs; In Vitro Techniques; Lung; Male; Naphthyridines; Nitriles; Ovalbumin; Phosphodiesterase 4 Inhibitors; Rats; Trachea

Allergen-specific IgE has long been regarded as a major molecular component of allergic asthma. Although IgE plays a central role in the early asthmatic response, its roles in the chronic phase, such as the late asthmatic response, airway hyperresponsiveness (AHR), and airway remodeling (goblet cell hyperplasia and subepithelial fibrosis) have not yet been defined well. In this study, we investigated the hypothesis that chronic responses could be induced by IgE-dependent mechanisms. BALB/c mice passively sensitized with an ovalbumin (OVA)-specific IgE monoclonal antibody (mAb) were repeatedly challenged with intratracheal administration of OVA. The first challenge induced early phase airway narrowing without any late response, but the fourth challenge caused not only an early but also a late phase response, AHR, and goblet cell hyperplasia. Macrophages, lymphocytes and neutrophils, but not eosinophils, were significantly increased in the lung 24h after the fourth challenge. Interestingly, levels of OVA-specific IgG1 in serum increased by multiple antigen challenges. A C3a receptor antagonist inhibited the late asthmatic response, AHR, and infiltration by neutrophils. In contrast, no late response, goblet cell hyperplasia, inflammatory cells, or production of IgG1 was observed in severe combined immunodeficient mice. On the other hand, seven challenges in BALB/c mice induced subepithelial fibrosis associated with infiltration by eosinophils. In conclusion, the allergic asthmatic responses induced by passive sensitization with IgE mAb can provide a useful model system to study the pathological roles of IgE in acute and chronic phases of allergic asthma.

Topics: Allergens; Animals; Antigen-Antibody Complex; Arginine; Asthma; Benzhydryl Compounds; Bronchial Hyperreactivity; Cell Movement; Chronic Disease; Disease Models, Animal; Goblet Cells; Humans; Hyperplasia; Immunization; Immunoglobulin E; Immunoglobulin G; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Receptors, Complement

Bronchial asthma is an inflammatory disease of the airways. Thrombin-activatable fibrinolysis inhibitor (TAFI) is a carboxypeptidase that besides inhibiting fibrinolysis, also regulates inflammatory processes. The only validated substrate known for TAFI is fibrin. In the present study we evaluated the role of TAFI in bronchial asthma by comparing the development of allergic bronchial asthma between wild-type (WT) and TAFI-deficient mice (KO).. Asthmatic inflammation was induced by sensitization and challenge with ovalbumin in WT (WT/OVA) and TAFI KO (KO/OVA) mice. WT mice (WT/SAL) and TAFI KO (KO/SAL) were used as controls. Cytokines, markers of inflammation, and coagulation were measured in bronchoalveolar lavage fluid (BALF).. Airway hyperresponsiveness was worse in KO/OVA mice than in WT/OVA mice or control mice. Markers of lung injury were significantly increased in BALF from KO/OVA mice compared to WT/OVA mice. Airway hyperresponsiveness and the BALF concentrations of IL-5 and osteopontin were significantly increased in KO/OVA mice compared to WT/OVA mice. Treatment of WT/OVA and KO/OVA mice with a C5a receptor antagonist significantly decreased hyperresponsiveness along with the BALF concentrations of total protein and C5a compared to untreated asthmatic mice.. The results of this study suggest that TAFI plays a protective role in the pathogenesis of allergic inflammation probably by inhibiting the complement system.

Topics: Airway Resistance; Animals; Asthma; Biomarkers; Blood Coagulation; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carboxypeptidase B2; Complement C5a; Fibrinolysis; Interleukin-5; L-Lactate Dehydrogenase; Lung Injury; Mice; Mice, Inbred C57BL; Mice, Knockout; Osteopontin; Ovalbumin; Plasminogen Activator Inhibitor 1; Receptors, Complement

Aiming the extract of Cordyceps sinensis significantly inhibits airway inflammation, airway hyperresponsiveness, and the infiltration of eosinophils in the airway of rats and may be related to the modulation of T helper (Th)1 and Th2 cells functions. The mechanisms of C. sinensis involved in modulation of suppression inflammation are not yet determined. In this study, the mechanism involved in the extract of C. sinensis-C.S.3-modulated suppression of inflammation was investigated in vivo and in vitro systems. The results showed that C.S.3 reduced airway inflammation in ovalbumin-induced allergic mice. Furthermore, we found C.S.3 could decrease extracellular signal-regulated kinase 1/2 signaling pathway to suppress activity of nuclear factor-κB in lung cells and cultured airway smooth muscle cells. Conclusion C.S.3 may provide clinical applications for asthma in the future.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cordyceps; Eosinophils; Extracellular Signal-Regulated MAP Kinases; Hypersensitivity; Inflammation; Male; Mice; Mice, Inbred BALB C; Myocytes, Smooth Muscle; NF-kappa B; Ovalbumin; Rats; Respiratory System

Asthma is a persistent inflammatory disease characterized by airway obstruction and hyperresponsiveness in association with airway inflammation. In the current research, we studied the anti-inflammatory and anti-asthmatic effects of tiarellic acid (TA) isolated from Tiarella polyphylla, based on asthmatic parameters, such as immunoglobulin E (IgE) level, cytokine release, eosinophilia, airway hyperresponsiveness (AHR), reactive oxygen species (ROS) and mucus hypersecretion, in an ovalbumin (OVA)-sensitized/challenged mouse model. TA significantly inhibited increases in IgE, levels of ROS and T helper cytokines, such as interleukin (IL)-4, IL-5, TNF-α, and IL-13, in bronchoalveolar lavage fluid (BALF), and effectively suppressed airway hyperresponsiveness, eosinophilia, and mucus hypersecretion in the asthmatic mouse model. In addition, we found that administration of TA attenuated ovalbumin-induced increases in NF-κB activity in lungs. The efficacy of TA was comparable to that of montelukast, a currently available anti-asthmatic drug. Our results support the utility of TA as a herbal medicine for asthma treatment and may have application in the development of anti-inflammatory and anti-asthmatic drugs.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophilia; Female; Immunoglobulin E; Inflammation; Leukocyte Count; Mice; Mice, Inbred BALB C; Oleanolic Acid; Ovalbumin; Phytotherapy; Transcription Factor RelA; Triterpenes

Activation of the alternative pathway of complement plays a critical role in the development of allergen-induced airway hyperresponsiveness (AHR) and inflammation in mice. Endogenous factor H, a potent inhibitor of the alternative pathway, is increased in the airways of sensitized and challenged mice, but its role in regulating inflammation or AHR has been unknown. We found that blocking the tissue-binding function of factor H with a competitive antagonist increased complement activation and tissue inflammation after allergen challenge of sensitized mice. Conversely, administration of a fusion protein that contains the iC3b/C3d binding region of complement receptor 2 linked to the inhibitory region of factor H, a molecule directly targeting complement-activating surfaces, protected mice in both primary and secondary challenge models of AHR and lung inflammation. Thus, although endogenous factor H does play a role in limiting the development of AHR, strategies to deliver the complement-regulatory region of factor H specifically to the site of inflammation provide greater protection than that afforded by endogenous regulators. Such an agent may be an effective therapy for the treatment of asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Complement Factor H; Complement Pathway, Alternative; Female; Inflammation; Inflammation Mediators; Mice; Mice, Inbred C57BL; Ovalbumin

The development of chronic allergic dermatitis in early life has been associated with increased onset and severity of allergic asthma later in life. However, the mechanisms linking these two diseases are poorly understood. In this study, we report that the development of oxazolone-induced chronic allergic dermatitis, in a mouse model, caused enhanced OVA-induced allergic asthma after the resolution of the former disease. Our findings show that oxazolone-induced dermatitis caused a marked increase in tissue mast cells, which persisted long after the resolution of this disease. Subsequent OVA sensitization and airway challenge of mice that had recovered from dermatitis resulted in increased allergic airway hyperreactivity. The findings demonstrate that the accumulation of mast cells during dermatitis has the detrimental effect of increasing allergic airway hypersensitivity. Importantly, our findings also show that exposure to a given allergen can modify the immune response to an unrelated allergen.

Topics: Animals; Bronchial Hyperreactivity; Cell Count; Chronic Disease; Dermatitis, Atopic; Disease Models, Animal; Disease Progression; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Ovalbumin; Oxazolone; Respiratory Hypersensitivity; Tissue Distribution

Histamine is detected in high concentrations in the airways during an allergic asthma response. In a murine model of allergic asthma, JNJ 7777120, an antagonist at the histamine H(4) receptor, reduces asthmatic symptoms, while the histamine H(1) receptor-selective antagonist mepyramine is virtually without effect. In the present study, we analyzed the effect of combined antagonism at the histamine H(1) and H(4) receptors in a murine asthma model in relation to the timing of their application, i.e. sensitization or provocation.. Asthma was induced in mice by sensitization and provocation with ovalbumin. JNJ 7777120 and/or mepyramine were injected subcutaneously either during sensitization or during provocation, and typical asthma parameters were analyzed. JNJ 7777120, but not mepyramine, reduced serum concentrations of anti-OVA IgE, inflammatory infiltrations in lung tissue, and eosinophilia in bronchoalveolar-lavage (BAL)-fluids independently of the timing of application. Upon application of JNJ 7777120 plus mepyramine in combination during provocation, mepyramine inhibited the effects of JNJ 7777120. In contrast, when applied during sensitization, mepyramine enhanced the disease-ameliorating effects of JNJ 7777120.. Our study indicates that both histamine H(1) and H(4) receptors play important roles in the course of murine experimental asthma. Unexpectedly, the contribution of these receptors to the pathogenesis differs between the two phases, sensitization or provocation. Since in human asthma, repeated contact to the allergen is not only provocation but also a boost of sensitization, a combined pharmacological targeting of histamine H(1) and H(4) receptors could be taken into consideration as an option for the prevention of asthma and maybe other allergic diseases.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Drug Administration Schedule; Drug Therapy, Combination; Enzyme-Linked Immunosorbent Assay; Female; Flow Cytometry; Histamine Antagonists; Histamine H2 Antagonists; Humans; Immunization; Immunoglobulin E; Indoles; Mice; Mice, Inbred BALB C; Ovalbumin; Piperazines; Pyrilamine; Receptors, G-Protein-Coupled; Receptors, Histamine; Receptors, Histamine H4; Time Factors; Treatment Outcome

Skullcapflavone II is a flavonoid derived from Scutellaria baicalensis, a widely used herbal medicine in anti-inflammatory and anticancer therapy in Korea. Skullcapflavone II antagonized the bradykinin receptor more potently than any of the other flavonoids derived from this plant. Here, we were investigated its therapeutic effects in a mouse model of ovalbumin (OVA)-induced allergic asthma. Administration of skullcapflavone II significantly reduced airway hyperresponsiveness (AHR), airway eosinophilia, Th2 cytokine production, and increased transforming growth factor-β1 (TGF-β1) levels in bronchoalveolarlavage (BAL) fluids and lungs from OVA-sensitized and -challenged mice. Skullcapflavone II administration also significantly suppressed subepithelial collagen deposition and goblet cell hyperplasia, elevated Smad7 expression and suppressed pSmad2/3 levels. Collectively, these findings indicate that skullcapflavone II, a potential bradykinin antagonist, reduced the major pathophysiological features of allergic asthma, at least in part by acting on TGF-β1/Smad signaling pathways. Thus, skullcapflavone II may have therapeutic potential for the treatment of allergic asthma.

Topics: Alanine Transaminase; Allergens; Animals; Anti-Inflammatory Agents; Aspartate Aminotransferases; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Collagen; Cytokines; Disease Models, Animal; Female; Flavonoids; Immunoglobulin E; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Smad Proteins

BACKGROUND AND PURPOSE Airway sensory nerves play a key role in respiratory cough, dyspnoea, airway hyper-responsiveness (AHR), all fundamental features of airway diseases [asthma and chronic obstructive pulmonary disease (COPD)]. Vagally mediated airway reflexes such as cough, bronchoconstriction and chest tightness originate from stimulation of airway sensory nerve endings. The transient receptor potential vanilloid 1 receptor (TRPV1) is present on peripheral terminals of airway sensory nerves and modulation of its activity represents a potential target for the pharmacological therapy of AHR in airway disease. EXPERIMENTAL APPROACH As guinea pig models can provide some of the essential features of asthma, including AHR, we have established the model with some classical pharmacological agents and examined the effect of the TRPV1 antagonists, SB-705498 and PF-04065463 on AHR to histamine evoked by ovalbumin (OA) in unanaesthetized sensitized guinea pigs restrained in a double chamber plethysmograph. Specific airway conductance (sGaw) derived from the airflow was calculated as a percentage of change from baseline. KEY RESULTS Cetirizine and salbutamol significantly inhibited OA-evoked bronchoconstriction [sGaw area under the curve (AUC): 70 and 78%, respectively]. Atropine, SB-705498 and PF-04065463 significantly inhibited OA-evoked AHR to histamine in unanaesthetized, OA-sensitized guinea pigs (sGaw AUC: 94%, 57% and 73%, respectively). Furthermore, this effect was not related to antagonism of histamine's activity. CONCLUSION AND IMPLICATIONS These data suggest that TRPV1 receptors located on airway sensory nerves are important in the development of AHR and that modulation of TRPV1-receptor activity represents a potential target for the pharmacological therapy of AHR in airway disease.

Topics: Adrenergic beta-2 Receptor Agonists; Albuterol; Allergens; Animals; Atropine; Bronchial Hyperreactivity; Bronchoconstriction; Cetirizine; Cyclopropanes; Guinea Pigs; Histamine; Histamine H1 Antagonists, Non-Sedating; Male; Muscarinic Antagonists; Ovalbumin; Pyrrolidines; Sulfonamides; TRPV Cation Channels; Urea

Protein kinase C variants (PKCs) have been involved in the control of airway smooth muscle (ASM) tone, and abnormalities in PKC-dependent signaling have been associated with respiratory diseases such as asthma. In this study, the role of atypical PKCζ in airway hyperresponsiveness was investigated, using an in-vitro model of TNFα-treated human bronchi and an in vivo guinea pig model of chronic asthma. Our results demonstrated that PKCζ-specific inhibition produced a significant increase in isoproterenol sensitivity in TNFα-treated bronchi and ovalbumin (OVA)-sensitized guinea pig bronchi. The role of epoxy-eicosanoids, known to exert anti-inflammatory effects in lung, on PKCζ expression and activity in these models was evaluated. An enhanced PKCζ protein expression was delineated in TNFα-treated bronchi when compared with control (untreated) and epoxy-eicosanoid-treated bronchi. Measurements of Ca(2+) sensitivity, performed in TNFα-treated bronchi, demonstrated that treatment with myristoylated (Myr) PKCζ peptide inhibitor resulted in significant reductions of pCa-induced tension. Epoxy-eicosanoid treatments had similar effects on Ca(2+) sensitivity in TNFα-treated bronchi. In control and epoxy-eicosanoid-treated bronchi, the phosphorylated forms of p38MAPK and CPI-17 were significantly decreased compared with the TNFα-treated bronchi. An enhanced expression of PKCζ was ascertained in our in-vivo model of allergic asthma. Hence an increased Ca(2+) sensitivity could be explained by the phosphorylation of p38-MAPK, which in turn leads to phosphorylation and activation of the CPI-17 regulatory protein. This process was reversed upon treatment with the Myr-PKCζ-peptide inhibitor. The present data provide relevant evidence regarding the role of PKCζ in human and rodent models of airways inflammation.

Topics: Adrenergic beta-2 Receptor Agonists; Animals; Arachidonic Acids; Asthma; Biomechanical Phenomena; Bronchi; Bronchial Hyperreactivity; Bronchodilator Agents; Calcium; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Guinea Pigs; Humans; Intracellular Signaling Peptides and Proteins; Male; Muscle Contraction; Muscle Proteins; Muscle Relaxation; Muscle, Smooth; Ovalbumin; p38 Mitogen-Activated Protein Kinases; Phosphoprotein Phosphatases; Phosphorylation; Pneumonia; Protein Kinase C; Protein Kinase Inhibitors; Tissue Culture Techniques; Tumor Necrosis Factor-alpha; Up-Regulation

The dose-response of the pleiotropic effects of statins on airway inflammation has not yet been established and may differ from that of their cholesterol-lowering effects. High oral doses of statins may have adverse effects, and it may be possible to overcome the side effects and low clinical efficacy by administering statins via inhalation. In this study, we hypothesize that simvastatin is a potential anti-inflammatory drug with biological and pharmacokinetic properties suitable for delivery by the inhaled route. Mice were immunized with ovalbumin (OVA) and then challenged with aerosol OVA. Simvastatin was locally delivered by inhalation (i.h.) and intratracheal injection (i.t.) or systematically delivered by intraperitoneal injection (i.p.) and gavage (i.g.) during the OVA challenge. In a mouse model of asthma, i.h. simvastatin significantly and dose-dependently attenuated airway inflammation, remodeling and hyperresponsiveness in a RhoA-dependent pathway. Upon comparing the pharmacodynamics, i.h. simvastatin had a more potent effect than that of i.g. and i.p. simvastatin, and the i.h. or i.t. delivery routes led to a higher drug concentration in local lung tissue and a lower drug concentration in the plasma than that obtained by the i.g. These results suggest that simvastatin is a potential anti-inflammatory drug for airway inflammatory diseases with properties suitable for delivery by inhalation, which will probably reduce the side effects and increase clinical efficacy.

Topics: Administration, Inhalation; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cytokines; Disease Models, Animal; Female; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Mice; Mice, Inbred BALB C; Ovalbumin; RNA, Messenger; Simvastatin

The prevalence of asthma in industrialized countries has been increasing dramatically and asthma is now the most common chronic disease of children in the United States. The rapidity of the increase strongly suggests that changes in environmental exposures are the likely cause of this epidemic. Further, the early onset of allergic manifestations suggests that these exposures may act on the prenatal development of the immune system. We have focused on the potential effects of bisphenol A (BPA), a chemical pollutant with one of the largest productions, on the development of childhood asthma. We have reported that perinatal BPA exposure promotes the development of allergic asthma in a mouse model. The current study was designed to identify a critical period of BPA exposure and to begin elucidating the mechanisms for this susceptibility.. Female BALB/c mice received 10 micro g/ml BPA in their drinking water from one week before pregnancy until the end of the study. Some of the pups were transferred in the first 48 h of life from their BPA-loaded mother to an unexposed mother, or vice versa. Half of the pups were sensitized with a low dose of the experimental allergen ovalbumin (OVA), the rest received PBS as an unsensitized controls. On day 22, the pups were challenged by inhalations of ovalbumin or PBS followed by quantification of eosinophils in and hyperreactivity of their airways, major indicators of experimental asthma in this classical mouse model. Hepatic expression of two isoforms of UDP-glucuronosyltransferase (Ugt) was quantified by quantitative RT-PCR at various ages.. Pups exposed to BPA in utero and through breast milk, or in utero only, displayed an asthma phenotype in response to their "suboptimal" allergic sensitization, whereas, pups only exposed to BPA postnatally from breast milk, did not. The expression of Ugt2b1, an isoform related to BPA clearance in rats, was not detectable in mouse fetuses and newborn pups, but increased by day 5 and approached adult levels by day 25.. Prenatal exposures that produce environmentally relevant burdens of BPA, followed by postnatal allergic sensitization and challenges, promote the development of experimental allergic asthma. Delayed expression of BPA-metabolizing enzymes may explain, at least in part, the enhanced fetal susceptibility to this common environmental contaminant.

Topics: Animals; Animals, Newborn; Asthma; Benzhydryl Compounds; Bronchial Hyperreactivity; Environmental Pollutants; Enzyme-Linked Immunosorbent Assay; Eosinophils; Female; Glucuronosyltransferase; Liver; Lung; Maternal Exposure; Mice; Mice, Inbred BALB C; Ovalbumin; Phenols; Pregnancy; Prenatal Exposure Delayed Effects; Real-Time Polymerase Chain Reaction; Risk Factors; RNA, Messenger

A hallmark of atopic asthma is development of chronic airways hyper-responsiveness (AHR) that persists in the face of ongoing exposure to perennial aeroallergens. We investigated underlying mechanisms in sensitized rats focusing on a strain expressing the high-allergen-responder phenotype characteristic of human atopic asthmatics, and find that their high susceptibility to aeroallergen-induced persistent AHR is associated with deficiencies in the immunoregulatory and mucosal trafficking properties of inducible T-regulatory cells (iTregs). Counterintuitively, AHR susceptibility was inversely related to aeroallergen exposure level, high exposures conferring protection. We demonstrate that underlying this AHR-susceptible phenotype is reduced capacity of airway mucosal dendritic cells (AMDCs) for allergen sampling in vivo; this defect is microenvironmentally acquired, as allergen uptake by these cells in vitro is normal. Moreover, intranasal transfer of in vitro aeroallergen-loaded AMDC from naïve animals into AHR-susceptible animals during prolonged aerosol challenge markedly boosts subsequent accumulation of iTregs in the airway mucosa and rapidly resolves their chronic AHR, suggesting that compromised antigen surveillance by AMDC resulting in defective functional programming of iTreg may be causally related to AHR susceptibility.

Topics: Adoptive Transfer; Allergens; Animals; Antigen Presentation; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Dendritic Cells; Disease Models, Animal; Disease Susceptibility; Humans; Immunologic Surveillance; Immunomodulation; Ovalbumin; Rats; Rats, Inbred BN; Respiratory Mucosa; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory

The p38 mitogen-activated protein kinase (MAPK) appears to play an important role in various pathophysiological responses and has been suggested to be involved in many processes considered critical to the inflammatory response and tissue remodeling. Bronchial asthma is a chronic inflammatory disorder of the airway accompanied by increased vascular permeability. Vascular endothelial growth factor (VEGF) is a potent stimulator of bronchial inflammation, airway remodeling, and physiologic dysregulation that augments antigen sensitization and T-helper type 2 cell (Th2)-mediated inflammation in allergic airway diseases. However, there are little data on the relationship between p38 MAPK signaling and VEGF expression in allergic airway disease.. This study aimed to investigate the role of p38 MAPK on the pathogenesis of allergic airway disease, more specifically in VEGF expression.. Using ovalbumin (OVA)-inhaled mice and a selective p38 MAPK inhibitor, SB 239063, the involvement of p38 MAPK in allergen-induced VEGF expression in the airway was evaluated.. The increases of phosphorylation of p38 MAPK, VEGF protein expression, and vascular permeability in the lung after OVA inhalation were decreased substantially by the administration of SB 239063. In addition, SB 239063 significantly reduced the increase of Th2 cytokines and OVA-specific IgE. The inhibition of p38 MAPK or VEGF signaling prevented and also decreased the increases in the number of inflammatory cells and airway hyperresponsiveness in OVA-induced allergic airway disease.. These results indicate that inhibition of p38 MAPK may attenuate allergen-induced airway inflammation and vascular leakage through modulation of VEGF expression in mice.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Endothelial Growth Factors; Female; Imidazoles; Immunoglobulin E; Mice; Mice, Inbred C57BL; Ovalbumin; p38 Mitogen-Activated Protein Kinases; Peptides, Cyclic; Protein Kinase Inhibitors; Pyrimidines; Vascular Endothelial Growth Factor A

During methacholine challenge tests of airway responsiveness, it is invariably assumed that the administered dose of agonist is accurately reflected in the dose that eventually reaches the airway smooth muscle (ASM). However, agonist must traverse a variety of tissue obstacles to reach the ASM, during which the agonist is subjected to both enzymatic breakdown and removal by the bronchial and pulmonary circulations. This raises the possibility that a significant fraction of the deposited agonist may never actually make it to the ASM. To understand the nature of this effect, we measured the time course of changes in airway resistance elicited by various durations of methacholine aerosol in mice. We fit to these data a computational model of a dynamically contracting airway responding to agonist that diffuses through an airway compartment, thereby obtaining rate constants that reflect the diffusive barrier to methacholine. We found that these barriers can contribute significantly to the time course of airway narrowing, raising the important possibility that alterations in the diffusive barrier presented by the airway wall may play a role in pathologically altered airway responsiveness.

Topics: Administration, Inhalation; Aerosols; Airway Resistance; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Bronchoconstrictor Agents; Computer Simulation; Diffusion; Disease Models, Animal; Dose-Response Relationship, Drug; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Models, Biological; Muscle, Smooth; Ovalbumin; Permeability

The glucocorticoid receptor (GR) is a transcription factor able to support either target gene activation via direct binding to DNA or gene repression via interfering with the activity of various proinflammatory transcription factors. An improved therapeutic profile for combating chronic inflammatory diseases has been reported through selectively modulating the GR by only triggering its transrepression function. We have studied in this paper the activity of Compound A (CpdA), a dissociated GR modulator favoring GR monomer formation, in a predominantly Th2-driven asthma model. CpdA acted similarly to the glucocorticoid dexamethasone (DEX) in counteracting OVA-induced airway hyperresponsiveness, recruitment of eosinophils, dendritic cells, neutrophils, B and T cells, and macrophages in bronchoalveolar lavage fluid, lung Th2, Tc2, Th17, Tc17, and mast cell infiltration, collagen deposition, and goblet cell metaplasia. Both CpdA and DEX inhibited Th2 cytokine production in bronchoalveolar lavage as well as nuclear translocation of NF-κB and its subsequent recruitment onto the IκBα promoter in the lung. By contrast, DEX but not CpdA induces expression of the GR-dependent model gene MAPK phosphatase 1 in the lung, confirming the dissociative action of CpdA. Mechanistically, we demonstrate that CpdA inhibited IL-4-induced STAT6 translocation and that GR is essential for CpdA to mediate chemokine repression. In conclusion, we clearly show in this study the anti-inflammatory effect of CpdA in a Th2-driven asthma model in the absence of transactivation, suggesting a potential therapeutic benefit of this strategy.

Topics: Acetates; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Dexamethasone; Disease Models, Animal; Drug Evaluation, Preclinical; Dual Specificity Phosphatase 1; Enzyme Induction; Gene Expression Regulation; Goblet Cells; Inflammation; Leukocytes; Lung; Mast Cells; Metaplasia; Mice; Mice, Inbred BALB C; Ovalbumin; Quaternary Ammonium Compounds; Receptors, Glucocorticoid; STAT6 Transcription Factor; Transcriptional Activation; Tyramine

The root of Peucedanum praeruptorum Dunn (PPD) is a commonly used traditional Chinese medicine for the treatment of asthma. Its major constituents, coumarins, were presumed to be responsible for its efficacy.. The potential of coumarins from PPD (CPPD) as anti-asthma agent was investigated.. Female BALB/c mice were sensitized and challenged with ovalbumin (OVA) to induce allergic airway inflammation. CPPD was administered intragastrically before every OVA challenge. Airway reactivity to the intravenous administration of acetylcholine chloride was measured 48h after final OVA inhalation. Airway inflammation was evaluated by leukocyte counts of bronchoalveolar lavage fluid (BALF) and histopathological analysis of lung lesions. Levels of interleukin (IL)-4, IL-5, IL-10, IL-13 and IFN-γ in BALF and OVA-specific immunoglobulin (Ig) E in serum, and activity of eosinophil peroxidase (EPO) in lung was measured. The percentage of CD4(+)CD25(+)Foxp3(+) regulatory T cells among CD4(+) T cells in spleen was analyzed by flow cytometry.. Compared with model group, CPPD significantly reduced airway hyperreactivity and airway eosinophilic inflammation, improved pathologic lesion of the lungs, reduced levels of IL-4, IL-5, IL-13 in BALF and OVA-specific IgE in serum, inhibited the activities of EPO in lung, and up-regulated levels of IL-10 and IFN-γ in BALF as well as the percentage of CD4(+)CD25(+)Foxp3(+) regulatory T cells in spleen.. CPPD can significantly suppress OVA-induced airway inflammation, airway hyperreactivity and Th2 predominant response in mice, showing great therapeutic potential for the treatment of allergic asthma.

Topics: Acetylcholine; Animals; Anti-Allergic Agents; Anti-Asthmatic Agents; Antigens; Apiaceae; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Coumarins; Disease Models, Animal; Drugs, Chinese Herbal; Female; Flow Cytometry; Inflammation Mediators; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plant Roots; Plants, Medicinal; Pulmonary Eosinophilia; T-Lymphocytes, Regulatory; Th2 Cells

Airway remodeling in bronchial asthma results from chronic, persistent airway inflammation. The effects of the reversal of airway remodeling by drug interventions remain to be elucidated. We investigated the effects of ONO-1301, a novel prostacyclin agonist with thromboxane inhibitory activity, on the prevention and reversibility of airway remodeling in an experimental chronic asthma model. Mice sensitized and challenged to ovalbumin (OVA) three times a week for 5 consecutive weeks were administered ONO-1301 or vehicle twice a day from the fourth week of OVA challenges. Twenty-four hours after the final OVA challenge, airway hyperresponsiveness (AHR) was assessed, and bronchoalveolar lavage was performed. Lung specimens were excised for staining to detect goblet-cell metaplasia, airway smooth muscle, and submucosal fibrosis. Mice administered ONO-1301 showed limited increases in AHR compared with mice administered the vehicle. The histological findings of airway remodeling were improved in ONO-1301-treated mice compared with vehicle-treated mice. Presumably, these therapeutic effects of ONO-1301 are attributable to the up-regulation of production of hepatocyte growth factor (HGF) in lung tissue, because the neutralization of HGF by antibodies prevented the effects of ONO-1301 on AHR and airway remodeling. Mice administered ONO-1301 showed similar levels of AHR and airway remodeling as mice administered montelukast, a cysteinyl-leukotriene-1 receptor antagonist, and lower levels were observed in mice administered dexamethasone. These data suggest that ONO-1301 exerts the effect of reversing airway remodeling, at least in part through an elevation of HGF in the lungs, and may be effective as an anti-remodeling drug in the treatment of asthma.

Topics: Acetates; Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cyclopropanes; Dexamethasone; Epoprostenol; Female; Goblet Cells; Hepatocyte Growth Factor; Inflammation; Lung; Metaplasia; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Pulmonary Fibrosis; Pyridines; Quinolines; Receptors, Leukotriene; Sulfides; Thromboxanes; Up-Regulation

Chaperonin 60.1 from Mycobacterium tuberculosis suppressed allergic lung inflammation and bronchial hyperresponsiveness in mice by a mechanism that is yet to be clarified. To investigate the possible antiinflammatory mechanism(s) of action of Cpn60.1 in a model of allergic lung inflammation, ovalbumin (OVA)-allergic mice were pretreated with Cpn60.1 intranasally 20 minutes before each OVA aerosol challenge in a total of three treatments. Readouts were performed 24 hours after last challenge. Pretreatment with Cpn60.1 (1.0-0.001 μg) significantly inhibited the number of eosinophils in bronchoalveolar lavage fluid (OVA, 49.2 ± 12.3 versus Cpn60.1 [1 μg dose], 90.4 ± 2.3 × 10(4) cells/ml) and IL-5 release (OVA, 43 ± 8.5 versus Cpn60.1 [1 μg dose], 3 ± 11 pg/ml) but increased IL-12 levels (OVA, 50 ± 24 versus Cpn60.1 [1 μg dose], 103 ± 13 pg/ml). The effect of Cpn60.1 on cell recruitment and IL-5, but not IL-12, release was abolished in TLR-4 knockout mice. Intravital microscopy demonstrated that Cpn60.1 reduced chemokine-mediated leukocyte rolling and transmigration across the vessel wall (rolling cells: eotaxin, 11.7 ± 1.1 versus Cpn60.1 [1 μg dose], 2.8 ± 1 cells in 30 s). Similarly, Cpn60.1 reduced eotaxin-induced leukocyte migration in vitro (eotaxin, 17.3 ± 3.3 versus Cpn60.1 [0.1 μg dose], 3.3 ± 0.4 cells × 10(4)/ml). Immunostaining demonstrated that Cpn60.1 inhibits VCAM-1 and increases vascular endothelial-cadherin expression in lung vascular tissue, suggesting that the antiinflammatory effect of Cpn60.1 is partly mediated by altering the expression of adhesion molecules. This study shows that Cpn60.1 inhibits leukocyte diapedesis by a TLR-4 and an adhesion molecule-dependent mechanism in allergic inflammation in mice.

Topics: Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Adhesion Molecules; Chaperonin 60; Disease Models, Animal; Eosinophils; Female; Interleukin-12; Interleukin-5; Leukocytes; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mycobacterium tuberculosis; Ovalbumin; Pneumonia; Toll-Like Receptor 4

Cissampelos sympodialis Eichl. (Menispermaceae) is a plant found in Northeastern and Southeast of Brazil and hot water infusion of C. sympodialis root bark is largely used in the indigenous and folk medicine to treat several inflammatory disorders, including asthma. Asthma is a chronic inflammatory allergic disease characterized by airway hyperreactivity (AHR), eosinophil tissue infiltration and lung remodeling. The aim of this study was to evaluate the therapeutic effect of C. sympodialis and its isolated alkaloid warifteine on allergen triggered airway hyperreactivity (AHR) and lung remodeling in murine model of asthma.. The oral pre-treatment with C. sympodialis or warifteine inhibited allergen-induced AHR to inhaled methacholine and IL-13 levels in the bronchoalveolar lavage (BAL). In order to investigate the therapeutic potential of C. sympodialis and warifteine, animals were treated 1h after the last ovalbumin (OVA) challenge in sensitized animals. Similarly to the pre-treatment, post-treatment with warifteine was effective to inhibit significantly AHR to inhaled methacholine and to reduce IL-13 levels in the BAL. In addition, oral pre- or post-treatments with C. sympodialis or warifteine reduced OVA-induced eosinophil tissue infiltration, mucus production and subepithelial fibrosis to values similar to nonallergic controls.. Our data show the anti-allergic and immunoregulatory properties of C. sympodialis, acting mostly through the active compound warifteine, to inhibit the airway hyperreactivity and lung remodeling through a mechanism at least partially dependent of IL-13 and eosinophil inhibition. Therefore placing warifteine as an interesting therapeutic candidate in allergic inflammation and corroborating the folk medicine use of C. sympodialis as anti-allergic plant.

Topics: Alkaloids; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cissampelos; Collagen; Disease Models, Animal; Eosinophils; Female; Humans; Interleukin-13; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plants, Medicinal; Respiratory Hypersensitivity

We investigated whether roxithromycin reduces ovalbumin-specific allergic asthma symptoms in mice, and we further investigated the inhibitory mechanism of roxithromycin in ovalbumin-specific allergic asthma.. Mice were divided into five groups (n = 10 for each): control group, roxithromycin-treated groups (5, 20 and 40 mg/kg) and ovalbumin-challenged group. We measured the recruitment of inflammatory cells into the bronchoalveolar lavage fluid (BALF) or the lung tissues by Kwik-Diff and hematoxylin and eosin (H&E) staining, goblet cell hyperplasia by alcian blue-periodic acid-Schiff (AB-PAS) staining, airway hyperresponsiveness (AHR) by whole-body plethysmograph chamber, cytokine and immunoglobulin E (IgE) levels by ELISA, and the activation of mitogen-activated protein (MAP) kinases and nuclear factor-kappa B (NF-κB) in the lung tissues by Western blotting.. Treatment with roxithromycin resulted in fewer inflammatory cells in the BALF and peribronchial areas, and decreased AHR, goblet cell hyperplasia, IgE levels and inflammatory cytokines, as well as MAP kinases and NF-κB activation, which are increased in lung tissues of mice with ovalbumin-induced allergic asthma.. Our data suggest that oral administration of roxithromycin suppresses ovalbumin-induced airway inflammation and AHR by regulating the inflammatory cytokines via MAP kinases/NF-κB pathway in inflammatory cells. Based on these results, we suggest that roxithromycin may be used as a therapeutic agent for allergy-induced asthma.

Topics: Allergens; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cell Count; Cell Differentiation; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Lung Compliance; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; NF-kappa B; Ovalbumin; Roxithromycin

The downstream of kinase (DOK)-1 is involved in the protein tyrosine kinase (PTK) pathway in mast cells, but the role of DOK-1 in the pathogenesis of asthma has not been defined. In this study, we have demonstrated a novel regulatory role of DOK-1 in airway inflammation and physiologic responses in a murine model of asthma using lentiviral vector containing DOK-1 cDNA or DOK-1-specific ShRNA. The OVA-induced inflammatory cells, airway hyperresponsiveness, Th2 cytokine expression, and mucus response were significantly reduced in DOK-1 overexpressing mice compared to OVA-challenged control mice. The transgenic introduction of DOK-1 significantly stimulated the activation and expression of STAT-4 and T-bet, while impressively inhibiting the activation and expression of STAT-6 and GATA-3 in airway epithelial cells. On the other hand, DOK-1 knockdown mice enhanced STAT-6 expression and its nuclear translocation compared to OVA-challenged control mice. When viewed in combination, our studies demonstrate DOK-1 regulates allergen-induced Th2 immune responses by selective stimulation and inhibition of STAT-4 and STAT-6 signaling pathways, respectively. These studies provide a novel insight on the regulatory role of DOK-1 in allergen-induced Th2 inflammation and airway responses, which has therapeutic potential for asthma and other allergic diseases.

Topics: Animals; Asthma; Bronchial Hyperreactivity; DNA-Binding Proteins; GATA3 Transcription Factor; Lentivirus; Mice; Ovalbumin; Phosphoproteins; RNA-Binding Proteins; STAT4 Transcription Factor; STAT6 Transcription Factor; T-Box Domain Proteins

Ovalbumin (OVA) is the most frequently used allergen in animal models of asthma. Lipopolysaccharide (LPS) contaminating commercial OVA may modulate the evoked airway inflammatory response to OVA. However, the effect of LPS in OVA on airway remodeling, especially airway smooth muscle (ASM) has not been evaluated. We hypothesized that LPS in commercial OVA may enhance allergen-induced airway inflammation and remodeling. Brown Norway rats were sensitized with OVA on day 0. PBS, OVA, or endotoxin-free OVA (Ef-OVA) was instilled intratracheally on days 14, 19, 24. Bronchoalveolar lavage (BAL) fluid, lung, and intrathoracic lymph node tissues were collected 48 h after the last challenge. Immunohistochemistry for α-smooth muscle actin, Periodic-Acid-Schiff staining, and real-time qPCR were performed. Airway hyperresponsiveness (AHR) was also measured. BAL fluid macrophages, eosinophils, neutrophils, and lymphocytes were increased in OVA-challenged animals, and macrophages and neutrophils were significantly lower in Ef-OVA-challenged animals. The ASM area in larger airways was significantly increased in both OVA and Ef-OVA compared with PBS-challenged animals. The mRNA expression of IFN-γ and IL-13 in lung tissues and IL-4 in lymph nodes was significantly increased by both OVA and Ef-OVA compared with PBS and were not significantly different between OVA and Ef-OVA. Monocyte chemoattractant protein (MCP)-1 in BAL fluid and AHR were significantly increased in OVA but not in Ef-OVA. LPS contamination in OVA contributes to the influx of macrophages and MCP-1 increase in the airways and to AHR after OVA challenges but does not affect OVA-induced Th1 and Th2 cytokine expression, goblet cell hyperplasia, and ASM remodeling.

Topics: Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL2; Chemokine CXCL1; Disease Models, Animal; Drug Contamination; Eosinophils; ErbB Receptors; Hyperplasia; Inflammation; Interferon-gamma; Interleukin-13; Lipopolysaccharides; Lymph Nodes; Lymphocytes; Macrophages; Male; Muscle, Smooth; Neutrophils; NF-kappa B; Ovalbumin; Rats; Rats, Inbred BN; Th1 Cells; Th2 Cells

Chrysin, a flavonoid obtained from various natural sources, has been reported to possess anti-inflammatory, antitumor, antioxidant and anti-allergic activities. However, its anti-inflammatory and immunoregulatory activities in asthma animal models are poorly understood. In the present study, we examined the effects of chrysin on airway inflammation and the possible mechanisms through which it acts in a murine model of allergic asthma. BALB/c mice sensitized and challenged to ovalbumin (OVA) were administered intragastrically with chrysin at a dose of 50 mg/kg daily. Chrysin significantly suppressed OVA-induced airway hyperresponsiveness (AHR) to acetylcholine chloride (Ach). Chrysin administration significantly inhibited the total inflammatory cell and eosinophil counts in bronchoalveolar lavage fluid (BALF) and total immunoglobulin E (IgE) levels in serum. Histological examination of lung tissue demonstrated that chrysin significantly attenuated allergen-induced lung eosinophilic inflammation and mucus-producing goblet cells in the airway. In addition, chrysin triggered a switch of the immune response to allergens towards a T-helper type 1 (Th1) profile by modulating the transcription factors T-bet and GATA-3 in allergic mice. These data suggest that chrysin exhibits anti-inflammatory and immunoregulatory properties and provides new insights into the immunopharmacological role of chrysin in terms of its effects in a murine model of asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Flavonoids; GATA3 Transcription Factor; Immunoglobulin E; Inflammation; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; T-Box Domain Proteins; Th1 Cells; Th2 Cells

Phosphodiesterase-4 (PDE-4) is responsible for metabolizing adenosine 3',5'-cyclic monophosphate that reduces the activation of a wide range of inflammatory cells including eosinophils. PDE-4 inhibitors are under development for the treatment of respiratory diseases such as asthma and chronic obstructive pulmonary disease. Herein, we report a novel PDE-4 inhibitor, PDE-423 (3-[1-(3-cyclopropylmethoxy-4-difluoromethoxybenzyl)-1H-pyrazol-3-yl]-benzoic acid), which shows good in vitro and in vivo oral activities. PDE-423 exhibited in vitro IC(50)s of 140 nM and 550 nM in enzyme assay and cell-based assay, respectively. In vivo study using ovalbumin-induced asthmatic mice revealed that PDE-423 reduced methacholine-stimulated airway hyperreactivity in a dose-dependent manner by once daily oral administration (ED(50)=18.3 mg/kg), in parallel with decreased eosinophil peroxidase activity and improved lung histology. In addition, PDE-423 was effective in diminishing lipopolysaccharide-induced neutrophilia in vivo as well as in vitro. Oral administration of PDE-423 (100 mg/kg) had no effect on the duration of xylazine/ketamine-induced anesthesia and did not induce vomiting incidence in ferrets up to the dose of 1000 mg/kg. The present study indicates that a novel PDE-4 inhibitor, PDE-423, has good pharmacological profiles implicating this as a potential candidate for the development of a new anti-asthmatic drug.

Topics: Administration, Oral; Animals; Anti-Asthmatic Agents; Asthma; Benzoates; Bronchial Hyperreactivity; Disease Models, Animal; Dose-Response Relationship, Drug; Eosinophil Peroxidase; Female; Ferrets; Guinea Pigs; Inhibitory Concentration 50; Lipopolysaccharides; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Phosphodiesterase 4 Inhibitors; Pyrazoles; Rats; Rats, Sprague-Dawley

Lipopolysaccharide (LPS) can exacerbate asthma; however, the mechanisms are not fully understood. This study investigated the effect of LPS on antigen-stimulated mast cell degranulation and the underlying mechanisms. We found that LPS enhanced degranulation in RBL-2H3 cells and mouse peritoneal mast cells upon FcεRI activation, in a dose- and time-dependent manner. Parallel to the alteration of degranulation, LPS increased FcεRI-activated Ca(2+) mobilization, as well as Ca(2+) entry through store-operated calcium channels (SOCs) evoked by thapsigargin. Blocking Ca(2+) entry through SOCs completely abolished LPS enhancement of mast cell degranulation. Consistent with functional alteration of SOCs, LPS increased mRNA and protein levels of Orai1 and STIM1, two major subunits of SOCs, in a time-dependent manner. In addition, LPS increased the mRNA level of Toll-like receptor 4 (TLR4) in a time-dependent manner. Blocking TLR4 with Cli-095 inhibited LPS, increasing transcription and expression of SOC subunits. Concomitantly, the effect of LPS enhancement of Ca(2+) mobilization and mast cell degranulation was largely reduced by Cli-095. Administration of LPS (1 μg) in vivo aggravated airway hyperreactivity and inflammatory reactions in allergic asthmatic mice. Histamine levels in serum and bronchoalveolar lavage fluid were increased by LPS treatment. In addition, Ca(2+) mobilization was enhanced in peritoneal mast cells isolated from LPS-treated asthmatic mice. Taken together, these results imply that LPS enhances mast cell degranulation, which potentially contributes to LPS exacerbating allergic asthma. Lipopolysaccharide increases Ca(2+) entry through SOCs by upregulating transcription and expression of SOC subunits, mainly through interacting with TLR4 in mast cells, resulting in enhancement of mast cell degranulation upon antigen stimulation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Calcium Channel Blockers; Calcium Channels; Calcium Signaling; Calcium-Transporting ATPases; Cell Degranulation; Cell Line, Tumor; Disease Models, Animal; Dose-Response Relationship, Drug; Enzyme Inhibitors; Female; Histamine; Lipopolysaccharides; Mast Cells; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; ORAI1 Protein; Ovalbumin; Rats; Receptors, IgE; RNA, Messenger; Stromal Interaction Molecule 1; Time Factors; Toll-Like Receptor 4

Resident alveolar macrophages (rAMs) residing in the bronchoalveolar lumen of the airways play an important role in limiting excessive inflammatory responses in the respiratory tract. High phagocytic activity along with hyporesponsiveness to inflammatory insults and lack of autonomous IFN-β production are crucial assets in this regulatory function. Using a mouse model of asthma, we analyzed the fate of rAMs both during and after allergic bronchial inflammation. Although nearly indistinguishable phenotypically from naïve rAMs, postinflammation rAMs exhibited a strongly reduced basal phagocytic capacity, accompanied by a markedly increased inflammatory reactivity to Toll-like receptors TLR-3 (poly I:C), TLR-4 [lipopolysaccharide (LPS)], and TLR-7 (imiquimod). Importantly, after inflammation, rAMs exhibited a switch from an IFN-β-defective to an IFN-β-competent phenotype, thus indicating the occurrence of a new, inflammatory-released rAM population in the postallergic lung. Analysis of rAM turnover revealed a rapid disappearance of naïve rAMs after the onset of inflammation. This inflammation-induced rAM turnover is critical for the development of the hyperinflammatory rAM phenotype observed after clearance of bronchial inflammation. These data document a novel mechanism of innate imprinting in which noninfectious bronchial inflammation causes alveolar macrophages to acquire a highly modified innate reactivity. The resulting increase in secretion of inflammatory mediators on TLR stimulation implies a role for this phenomenon of innate imprinting in the increased sensitivity of postallergic lungs to inflammatory insults.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; Cell Differentiation; Cells, Cultured; Cytokines; Disease Progression; Female; Inflammation Mediators; Interferon-beta; Macrophages, Alveolar; Mice; Mice, Inbred C57BL; Ovalbumin; Phagocytosis; Phenotype; Toll-Like Receptors

The intestinal microbiota has important effects on host immune responses and feeding with certain commensal organisms has anti-inflammatory effects in a variety of diseases, including experimental asthma. The aim of the current study was to examine how robust the effects of feeding with the commensal strain, Bifidobacterium longum (Bif) were on the pulmonary responses to allergen sensitization and challenge.. BALB/c mice were given two intraperitoneal injections of ovalbumin (10 μg in alum) on days 0 and 7 and were fed daily with Bif or vehicle from days 0-14. Challenges with ovalbumin (10 μg) were administered intra-nasally once on day 14 or three times on days 14, 15 and 16 and the lung inflammatory response was assessed one day later.. Bif feeding attenuated airway inflammation following a single ovalbumin challenge, reducing bronchoalveolar lavage (BAL) eosinophilia, BAL fluid IL-4 protein and BAL cell IL-4 and IFN-γ mRNA levels. However, BAL fluid IL-5 protein was increased. There was an accompanying increase in lung regulatory T cells assessed by flow cytometry. Responses to triple challenge with ovalbumin were much less affected by Bif feeding, including unchanged cytokine levels, ovalbumin-specific IgE and airway hyperresponsiveness to methacholine.. These results show modest immunoregulatory effects of oral feeding with Bif with inhibition of certain components of allergen-induced airway inflammation that is associated with the expansion of regulatory T cells in the lungs but that is overcome by repeated allergen exposure.

Topics: Allergens; Animals; Bifidobacterium; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Inflammation Mediators; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Probiotics

Protein-S-glutathionylation (PSSG) is an oxidative modification of reactive cysteines that has emerged as an important player in pathophysiological processes. Under physiological conditions, the thiol transferase, glutaredoxin-1 (Glrx1) catalyses deglutathionylation. Although we previously demonstrated that Glrx1 expression is increased in mice with allergic inflammation, the impact of Glrx1/PSSG in the development of allergic airways disease remains unknown. In the present study we examined the impact of genetic ablation of Glrx1 in the pathogenesis of allergic inflammation and airway hyperresponsiveness (AHR) in mice. Glrx1(-/-) or WT mice were subjected to the antigen, ovalbumin (OVA), and parameters of allergic airways disease were evaluated 48 h after three challenges, and 48 h or 7 days after six challenges with aerosolized antigen. Although no clear increases in PSSG were observed in WT mice in response to OVA, marked increases were detected in lung tissue of mice lacking Glrx1 48 h following six antigen challenges. Inflammation and expression of proinflammatory mediators were decreased in Glrx1(-/-) mice, dependent on the time of analysis. WT and Glrx1(-/-) mice demonstrated comparable increases in AHR 48 h after three or six challenges with OVA. However, 7 days postcessation of six challenges, parameters of AHR in Glrx1(-/-) mice were resolved to control levels, accompanied by marked decreases in mucus metaplasia and expression of Muc5AC and GOB5. These results demonstrate that the Glrx1/S-glutathionylation redox status in mice is a critical regulator of AHR, suggesting that avenues to increase S-glutathionylation of specific target proteins may be beneficial to attenuate AHR.

Topics: Animals; Bronchial Hyperreactivity; Glutaredoxins; Glutathione; Lung; Lung Diseases; Metaplasia; Mice; Mucus; Ovalbumin; Pneumonia; Proteins

In traditional Chinese medicine, arsenous compounds, including arsenic trioxide (ATO), are often used to treat many diseases, which are safe and effective. Recently, studies have indicated that Th17- IL-17 involved in the pathogenesis and development of asthma. The goal of this study was to investigate the effect and mechanism of ATO on asthma, especially the Th17- IL-17 axis.We used oval bumin (OVA)-immunized mice as a model for asthma and treated mice with ATO or dexamethasone. The mice were then monitored airway responsiveness, airway inflammation, mucus production, IL-17 levels in BALF and the positive rate of Th17 cells. In vitro, CD4+ T cells from splenic cell suspensions were separated and purified. We measured the expression of IL-17 and caspase-12 protein in purified CD4+ T cells, and detected IL-17 levels in CD4+ T lymphocyte culture solution with or without ATO. Moreover, apoptosis, mitochondrial membrane potential, cytosolic calcium were analyzed. We found that ATO could reduce airway responsiveness, airway inflammation, mucus hyperplasia, the expression of IL-17 in BALF and the positive rate of Th17 cells at a level comparable to treatment with DXM. In vitro data suggested that ATO can induce CD4+ T cells apoptosis, cause mitochondrial dysfunction, Ca2+ overload and promote caspase-12 activation. Our study suggested that ATO had potential medical value for the treatment of human asthma..

Topics: Animals; Anti-Asthmatic Agents; Apoptosis; Arsenic Trioxide; Arsenicals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Calcium; Caspase 12; Cells, Cultured; Dexamethasone; Disease Models, Animal; Female; Interleukin-17; Lung; Membrane Potential, Mitochondrial; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Oxides; Signal Transduction; Th17 Cells

Triphala (TRP), a herbal extract from Tibetan medicine, has been shown to affect lymphocytes and natural killer T (NKT) cell function. We hypothesize that TRP could ameliorate bronchial hyperreactivity through immune-cell modulations.. Asthma mouse models were generated through intraperitoneal (IP) injections of ovalbumin (OVA)/2 weeks followed by repeated intranasal OVA challenges. Mice were then treated with normal saline (OVA/NS) or Triphala (OVA/TRP). Data were compared with mice treated with inhaled budesonide. All groups were assessed for allergen-induced hyperreactivity; lymphocytes from lungs, livers and spleens were analyzed for OVA-induced proliferation and their alterations were determined by flow cytometry. Oxidative reactivity using chemiluminescence, serum anti-OVA antibodies level and lung histology were assessed.. Both TRP and budesonide significantly ameliorated functional and histological OVA-induced bronchial hyperreactivity. TRP had no effect on serum anti-OVA antibodies as compared with decreased levels following budesonide treatment. Furthermore, a significant increase in lung and spleen CD4 counts and a decrease in the liver were noted after TRP treatments. Bronchoalveolar fluid from TRP-treated animals but not from the budesonide-treated animals showed anti-oxidative effects.. TRP and budesonide caused a significant decrease in bronchial reactivity. TRP treatment altered immune-cell distributions and showed anti-oxidative properties. These findings suggest that immune-cell modulation with TRP can ameliorate lung injury.

Topics: Administration, Inhalation; Alanine Transaminase; Animals; Anti-Asthmatic Agents; Antioxidants; Asthma; Biomarkers; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Budesonide; CD4-Positive T-Lymphocytes; Cell Proliferation; Cells, Cultured; Disease Models, Animal; Flow Cytometry; Immunity, Humoral; Liver; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Oxidative Stress; Plant Extracts; Spleen

Interleukin (IL)-33, which mediates the T(h)2 allergic pathway, may play a key role in allergic airway inflammation. This study was conducted to evaluate the therapeutic potential of anti-IL-33 antibody for treatment of allergic inflammation of the lower airway in a murine model.. Twenty-four BALB/c mice were used in this study. Saline was used for sensitization and challenge of mice in Group A (control group, n = 6). Mice in Group B (ovalbumin (OVA) group, n = 6) received intraperitoneal (ip) and intranasal OVA challenge. In Group C (control IgG group, n = 6), mice received ip injection with control IgG prior to OVA challenge. Mice in Group D (anti-IL-33 group, n = 6) received an ip injection of anti-IL-33 prior to challenge. Measurements of serum total and OVA-specific IgE and the number of eosinophils, neutrophils, and lymphocytes in bronchoalveolar lavage (BAL) fluid were performed. We performed histopathologic examination to evaluate the degree of eosinophilic infiltration in lung tissue. Airway hyperreactivity was measured according to change of enhanced pause (Penh).. A significant decrease in serum total and OVA-specific IgE and the number of eosinophils and neutrophils in BAL fluid was observed in Group D, compared with Group B or Group C (p < .05). In Group D, treatment with anti-IL-33 resulted in a significant decrease in eosinophilic infiltration in lung tissue, compared with Group B and Group C (p < .05). Degree of airway hyperreactivity, measured by Penh, showed a significant decrease in the anti-IL-33 treatment group, compared with the OVA group or the control IgG treatment group (p < .01, at 50 mg/mL of methacholine).. Anti-IL-33 has therapeutic potential for treatment of allergic inflammation of the lower airway.

Topics: Animals; Antibodies; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Female; Immunoglobulin E; Interleukin-33; Interleukins; Mice; Mice, Inbred BALB C; Ovalbumin

Inhaled heparin appears to provide benefit in the management of airway hyper-reactivity and inflammation. The pharmacodynamics of inhaled heparin are however transient. Providing sustained heparin concentrations in the respiratory tract should provide for an extended duration of action. We examined the in-vivo efficacy of a nebulised controlled-release microparticle formulation of heparin in modifying antigen-induced airway hyper-reactivity (AHR) and lung inflammation. Heparin-loaded biodegradable poly (D,L-lactide-co-glycolide) microparticles were prepared by spray-drying. Aerosol properties for both nebulised heparin solution and heparin microparticles displayed characteristics consistent with heparin delivery to the respiratory tract. In vitro release assays showed heparin to be released from the microparticles over 8-12 h and for the heparin to remain functional. Temporal pharmacodynamic responses were studied in an ovalbumin-sensitised in vivo model exhibiting AHR and airway inflammation. Despite a reduced total dose of heparin deposited in the airways following nebulisation with heparin microparticles, this treatment led to a more sustained inhibitory effect upon AHR and airway inflammation than equivalent doses of nebulised heparin solution. The work supports extended-release heparin as an inhalation dosing strategy in experimental therapeutic applications aimed at improving the pharmacodynamics of heparin in the treatment of AHR and lung inflammation.

Topics: Administration, Inhalation; Animals; Antigens; Bronchial Hyperreactivity; Delayed-Action Preparations; Drug Carriers; Guinea Pigs; Heparin; Lactic Acid; Lung; Male; Nebulizers and Vaporizers; Ovalbumin; Pneumonia; Polyglycolic Acid; Polylactic Acid-Polyglycolic Acid Copolymer; Solutions

Exposure to ultrafine particles exerts diverse harmful effects including aggravation of pulmonary diseases like asthma. Recently we demonstrated in a mouse model for allergic airway inflammation that particle-derived oxidative stress plays a crucial role during augmentation of allergen-induced lung inflammation by ultrafine carbon particle (UfCP) inhalation. The mechanisms how particle inhalation might change the inflammatory balance in the lungs, leading to accelerated inflammatory reactions, remain unclear. Lipid mediators, known to be immediately generated in response to tissue injury, might be strong candidates for priming this particle-triggered change of the inflammatory balance.. We hypothesize that inhalation of UfCP may disturb the balance of pro- and anti-inflammatory lipid mediators in: i) a model for acute allergic pulmonary inflammation, exposing mice for 24 h before allergen challenge to UfCP inhalation (51.7 nm, 507 μg/m3), and ii) an in-vitro model with primary rat alveolar macrophages (AM) incubated with UfCP (10 μg/1 x 106 cells/ml) for 1 h. Lungs and AM were analysed for pro- and anti-inflammatory lipid mediators, namely leukotriene B4 (LTB4), prostaglandin E2 (PGE2), 15(S)-hydroxy-eicosatetraenoic acid (15(S)-HETE), lipoxin A4 (LXA4) and oxidative stress marker 8-isoprostane by enzyme immunoassays and immunohistochemistry.. In non-sensitized mice UfCP exposure induced a light non-significant increase of all lipid mediators. Similarly but significantly in rat AM all lipid mediators were induced already within 1 h of UfCP stimulation. Also sensitized and challenge mice exposed to filtered air showed a partially significant increase in all lipid mediators. In sensitized and challenged mice UfCP exposure induced highest significant levels of all lipid mediators in the lungs together with the peak of allergic airway inflammation on day 7 after UfCP inhalation. The levels of LTB4, 8-isoprostane and PGE2 were significantly increased also one day after UfCP exposure. Immunohistochemistry localized highest concentrations of PGE2 especially in AM one day after UfCP exposure.. Our results suggest that UfCP exposure affects the balance between pro- and anti-inflammatory lipid mediators. In allergic mice, where the endogenous balance of pro- and anti-inflammatory mediators is already altered, UfCP exposure aggravates the inflammation and the increase in anti-inflammatory, pro-resolving lipid mediators is insufficient to counterbalance the extensive inflammatory response. This may be a contributing mechanism that explains the increased susceptibility of asthmatic patients towards particle exposure.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Carbon; Cells, Cultured; Disease Models, Animal; Female; Inflammation Mediators; Inhalation Exposure; Lipid Peroxidation; Lung; Macrophages; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Particle Size; Particulate Matter; Pulmonary Alveoli; Rats; Rats, Inbred WKY

Adiponectin is an adipose derived hormone that declines in obesity. We have previously shown that exogenous administration of adiponectin reduces allergic airways responses in mice. T-cadherin (T-cad; Cdh13) is a binding protein for the high molecular weight isoforms of adiponectin. To determine whether the beneficial effects of adiponectin on allergic airways responses require T-cad, we sensitized wildtype (WT), T-cadherin deficient (T-cad(-/-)) and adiponectin and T-cad bideficient mice to ovalbumin (OVA) and challenged the mice with aerosolized OVA or PBS. Compared to WT, T-cad(-/-) mice were protected against OVA-induced airway hyperresponsiveness, increases in BAL inflammatory cells, and induction of IL-13, IL-17, and eotaxin expression. Histological analysis of the lungs of OVA-challenged T-cad(-/-) versus WT mice indicated reduced inflammation around the airways, and reduced mucous cell hyperplasia. Combined adiponectin and T-cad deficiency reversed the effects of T-cad deficiency alone, indicating that the observed effects of T-cad deficiency require adiponectin. Compared to WT, serum adiponectin was markedly increased in T-cad(-/-) mice, likely because adiponectin that is normally sequestered by endothelial T-cad remains free in the circulation. In conclusion, T-cad does not mediate the protective effects of adiponectin. Instead, mice lacking T-cad have reduced allergic airways disease, likely because elevated serum adiponectin levels act on other adiponectin signaling pathways.

Topics: Adiponectin; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Cadherins; Crosses, Genetic; Inflammation; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Ovalbumin; Protein Binding; Signal Transduction

Bisphenol A (BPA) is a high-production volume chemical classified as an environmental estrogen and used primarily in the plastics industry. BPA's increased usage correlates with rising BPA levels in people and a corresponding increase in the incidence of asthma. Due to limited studies, the contribution of maternal BPA exposure to allergic asthma pathogenesis is unclear. Using two established mouse models of allergic asthma, we examined whether developmental exposure to BPA alters hallmarks of allergic lung inflammation in adult offspring. Pregnant C57BL/6 dams were gavaged with 0, 0.5, 5, 50, or 500 μg BPA/kg/day from gestational day 6 until postnatal day 21. To induce allergic inflammation, adult offspring were mucosally sensitized with inhaled ovalbumin containing low-dose lipopolysaccharide or ip sensitized using ovalbumin with alum followed by ovalbumin aerosol challenge. In the mucosal sensitization model, female offspring that were maternally exposed to ≥ 50 μg BPA/kg/day displayed enhanced airway lymphocytic and lung inflammation, compared with offspring of control dams. Peritoneally sensitized, female offspring exposed to ≤ 50 μg BPA/kg/day presented dampened lung eosinophilia, compared with vehicle controls. Male offspring did not exhibit these differences in either sensitization model. Our data demonstrate that maternal exposure to BPA has subtle and qualitatively different effects on allergic inflammation, which are critically dependent upon route of allergen sensitization and sex. However, these subtle, yet persistent changes due to developmental exposure to BPA did not lead to significant differences in overall airway responsiveness, suggesting that early life exposure to BPA does not exacerbate allergic inflammation into adulthood.

Topics: Administration, Oral; Animals; Asthma; Benzhydryl Compounds; Bronchial Hyperreactivity; Environmental Pollutants; Female; Lymphocytes; Male; Maternal Exposure; Mice; Mice, Inbred C57BL; Ovalbumin; Phenols; Pregnancy; Prenatal Exposure Delayed Effects; Sex Factors

Blockage of nerve growth factor (NGF) by anti-NGF antibodies can inhibit allergic airway hyper-responsiveness in mice. This study was aimed at determining the mechanisms underlying the action of anti-NGF in vivo.. BALB/c mice were sensitized with ovalbumin (OVA) and treated with anti-NGF. At 1 day after the last challenge, their airway responsiveness and inflammation were examined and the levels of cytokine and transcription factor mRNA transcripts in the lungs and cytokines in the bronchoalveolar lavage fluid were determined. The frequency of different functional T cells and the levels of serum OVA-specific antibodies were measured.. OVA challenge induced severe airway resistance, inflammation, higher levels of IL-4, TNFα, IL-17A, TGFβ, GATA-3 and RORγT expression and increased Th2 and Th17 cells and IgE responses, but decreased IFNγ and IL-10 responses, T-bet and Foxp3 expression and Th1 and Tregs. Treatment with anti-NGF significantly reduced allergic airway resistance and inflammation, up-regulated IFNγ, IL-10, TGFβ, T-bet, and Foxp3 expression, increased Th1 and Tregs, but down-regulated IL-4, TNFα, IL-17A, RORγT and GATA-3 expression and reduced Th2 and Th17 cells, accompanied by increased serum IgG2a.. Anti-NGF inhibits allergic airway inflammation by modulating the balance of pro- and anti-asthmatic T cell responses in the lungs of mice.

Topics: Allergens; Animals; Antibodies; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Female; Forkhead Transcription Factors; GATA3 Transcription Factor; Immunoglobulin E; Immunoglobulin G; Mice; Mice, Inbred BALB C; Nerve Growth Factor; Nuclear Receptor Subfamily 1, Group F, Member 3; Ovalbumin; Pneumonia; RNA, Messenger; T-Box Domain Proteins; T-Lymphocytes

The receptor for advanced glycation end products (RAGE) is a multiligand receptor that has been shown to contribute to the pathogenesis of diabetes, atherosclerosis, and neurodegeneration. However, its role in asthma and allergic airway disease is largely unknown. These studies use a house dust mite (HDM) mouse model of asthma/allergic airway disease. Respiratory mechanics were assessed and compared between wild-type and RAGE knockout mice. Bronchovascular architecture was assessed with quantitative scoring, and expression of RAGE, immunoglobulins, and relevant cytokines was assessed by standard protein detection methods and/or quantitative RT-PCR. The absence of RAGE abolishes most assessed measures of pathology, including airway hypersensitivity (resistance, tissue damping, and elastance), eosinophilic inflammation, and airway remodeling. IL-4 secretion, isotype class switching, and antigen recognition are intact in the absence of RAGE. In contrast, normal increases in IL-5, IL-13, eotaxin, and eotaxin-2 production are abrogated in the RAGE knockouts. IL-17 indicates complex regulation, with elevated baseline expression in RAGE knockouts, but no induction in response to allergen. Treatment of WT mice with an inhibitor of RAGE markedly reduces inflammation in the HDM model, suggesting that RAGE inhibition may serve as a promising therapeutic strategy. Finally, the results in the HDM model are recapitulated in an ovalbumin model of asthma, suggesting that RAGE plays a role in asthma irrespective of the identity of the allergens involved.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Chemokine CCL24; Eosinophilia; Immunoglobulin G; Interleukin-13; Interleukin-4; Interleukin-5; Lung; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Protein Transport; Pyroglyphidae; Receptor for Advanced Glycation End Products; Receptors, Immunologic

Allergic lung inflammation is mediated by allergen-specific T responses, which are negatively regulated by regulatory T cells (Tregs). Previous studies have reported that inoculation of indigenous Clostridium species in the early lives of mice can induce Tregs that colonize the colon. However, whether inoculation of C leptum alone in adult mice could induce systemic Treg responses and inhibit allergic airway inflammation remains unclear.. To investigate the effect of oral administration of C leptum on systemic Treg responses and allergic airway inflammation in a mouse model of asthma.. Adult BABL/c mice were injected with ovalbumin to induce asthma and treated orally with C leptum or vehicle daily for 2 weeks. The numbers of Foxp3(+)CD4(+)CD25(+) Tregs in both the spleen and mediastinal lymph nodes were examined by flow cytometry. After allergen challenge, the airway hyperresponsiveness of individual mice was measured, and the numbers of inflammatory infiltrates and the levels of cytokines in bronchoalveolar lavage fluids ere determined.. Oral feeding with C leptum increased the percentage and total number of Tregs in the spleens and mediastinal lymph nodes at 14 days after inoculation and attenuated allergen-induced airway hyperresponsiveness and inflammation by inhibiting inflammatory cytokine production but enhancing interleukin 10 and transforming growth factor β1 production in the lungs.. Oral treatment with C leptum can attenuate induced allergic airway inflammation in adult mice.

Topics: Administration, Oral; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Clostridium; Cytokines; Disease Models, Animal; Female; Hypersensitivity; Inflammation; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes, Regulatory

Caveolae are flask-shaped plasma membrane invaginations expressing the scaffolding caveolin proteins. Although caveolins have been found in endothelium and epithelium (where they regulate nitric oxide synthase activity), their role in smooth muscle is still under investigation. We and others have previously shown that caveolae of human airway smooth muscle (ASM), which express caveolin-1, contain Ca(2+) and force regulatory proteins and are involved in mediating the effects of inflammatory cytokines such as TNF-α on intracellular Ca(2+) concentration responses to agonist. Accordingly, we tested the hypothesis that in vivo, absence of caveolin-1 leads to reduced airway hyperresponsiveness, using a knockout (KO) (Cav1 KO) mouse and an ovalbumin-sensitized/challenged (OVA) model of allergic airway hyperresponsiveness. Surprisingly, airway responsiveness to methacholine, tested by use of a FlexiVent system, was increased in Cav1 KO control (CTL) as well as KO OVA mice, which could not be explained by a blunted immune response to OVA. In ASM of wild-type (WT) OVA mice, expression of caveolin-1, the caveolar adapter proteins cavins 1-3, and caveolae-associated Ca(2+) and force regulatory proteins such as Orai1 and RhoA were all increased, effects absent in Cav1 KO CTL and OVA mice. However, as with WT OVA, both CTL and OVA Cav1 KO airways showed signs of enhanced remodeling, with high expression of proliferation markers and increased collagen. Separately, epithelial cells from airways of all three groups displayed lower endothelial but higher inducible nitric oxide synthase and arginase expression. Arginase activity was also increased in these three groups, and the inhibitor nor-NOHA (N-omega-nor-l-arginine) enhanced sensitivity of isolated tracheal rings to ACh, especially in Cav1 KO mice. On the basis of these data disproving our original hypothesis, we conclude that caveolin-1 has complex effects on ASM vs. epithelium, resulting in airway hyperreactivity in vivo mediated by altered airway remodeling and bronchodilation.

Topics: Animals; Bronchial Hyperreactivity; Bronchoconstriction; Bronchoconstrictor Agents; Calcium; Caveolin 1; Disease Models, Animal; Methacholine Chloride; Mice; Mice, Inbred Strains; Mice, Knockout; Muscle, Smooth; Ovalbumin; Receptors, Cell Surface; Receptors, Tumor Necrosis Factor, Type I; Respiratory Mucosa

Heat shock proteins (HSPs), produced in response to stress, are suppressive in disease models. We previously showed that Mycobacterium leprae HSP65 prevented development of airway hyperresponsiveness and inflammation in mice. Our goal in this study was to define the mechanism responsible for the suppressive effects of HSP. In one in vivo approach, BALB/c mice were sensitized to OVA, followed by primary OVA challenges. Several weeks later, HSP65 was administered prior to a single, provocative secondary challenge. In a second in vivo approach, the secondary challenge was replaced by intratracheal instillation of allergen-pulsed bone marrow-derived dendritic cells (BMDCs). The in vitro effects of HSP65 on BMDCs were examined in coculture experiments with CD4(+) T cells. In vivo, HSP65 prevented the development of airway hyperresponsiveness and inflammation. Additionally, Th1 cytokine levels in bronchoalveolar lavage fluid were increased. In vitro, HSP65 induced Notch receptor ligand Delta1 expression on BMDCs, and HSP65-treated BMDCs skewed CD4(+) T cells to Th1 cytokine production. Thus, HSP65-induced effects on allergen-induced airway hyperresponsiveness and inflammation were associated with increased Delta1 expression on dendritic cells, modulation of dendritic cell function, and CD4(+) Th1 cytokine production.

Topics: Animals; Bacterial Proteins; Bronchial Hyperreactivity; Cells, Cultured; Chaperonin 60; Coculture Techniques; Cytokines; Dendritic Cells; Disease Models, Animal; Female; Inflammation; Mice; Mice, Inbred BALB C; Mycobacterium leprae; Ovalbumin; Th1 Cells

α- and γ-Mangostin, which are the major xanthones purified from a Mangosteen, Garcinia mangostana Linn., exhibit a wide range of anticancer, antioxidant, and anti-inflammatory activities. Here, we assessed their therapeutic effects in a mouse model of ovalbumin (OVA)-induced allergic asthma. Animals were treated with α- and γ-mangostins orally for 3 days at doses of 10 and 30 mg/kg daily, 1h before the OVA challenge. Administration of α- and γ-mangostins significantly reduced the major pathophysiological features of allergic asthma, including inflammatory cell recruitment into the airway, airway hyperresponsiveness (AHR), and increased levels of Th2 cytokines. In addition, α- and γ-mangostins attenuated the increases in phosphoinositide 3-kinase (PI3K) activity, phosphorylation of Akt, and NF-κB in nuclear protein extracts after OVA challenge. In conclusion, α- and γ-mangostin may have therapeutic potential for the treatment of allergic asthma.

Topics: Administration, Oral; Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Garcinia mangostana; Immunoglobulin E; Interleukins; Lung; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins c-akt; PTEN Phosphohydrolase; Transforming Growth Factor beta1; Xanthones

A disintegrin and metalloproteinase with thrombospondin motifs (ADAMTS) constitute a family of endopeptidases related to matrix metalloproteinases. These proteinases have been largely implicated in tissue remodeling associated with pathological processes. Among them, ADAMTS12 was identified as an asthma-associated gene in a human genome screening program. However, its functional implication in asthma is not yet documented. The present study aims at investigating potential ADAMTS-12 functions in experimental models of allergic airways disease. Two different in vivo protocols of allergen-induced airways disease were applied to the recently generated Adamts12-deficient mice and corresponding wild-type mice. In this study, we provide evidence for a protective effect of ADAMTS-12 against bronchial inflammation and hyperresponsiveness. In the absence of Adamts12, challenge with different allergens (OVA and house dust mite) led to exacerbated eosinophilic inflammation in the bronchoalveolar lavage fluid and in lung tissue, along with airway dysfunction assessed by increased airway responsiveness following methacholine exposure. Furthermore, mast cell counts and ST2 receptor and IL-33 levels were higher in the lungs of allergen-challenged Adamts12-deficient mice. The present study provides, to our knowledge, the first experimental evidence for a contribution of ADAMTS-12 as a key mediator in airways disease, interfering with immunological processes leading to inflammation and airway hyperresponsiveness.

Topics: ADAM Proteins; ADAMTS Proteins; Animals; Antigens, Dermatophagoides; Bronchial Hyperreactivity; Disease Models, Animal; Humans; Immunophenotyping; Inflammation Mediators; Male; Mice; Mice, 129 Strain; Mice, Knockout; Ovalbumin

Sulfur dioxide (SO(2)) is a common air pollutant that triggers asthmatic symptoms, but its toxicological mechanisms are not fully understood. Specifically, it is unclear how airborne SO(2) affects airway hyperresponsiveness (AHR) - a hallmark feature of asthma. To this end, we investigated the effects of chronic exposure to SO(2) on AHR, airway inflammation, tissue remodeling, cell stiffness (G') and contractility of the airway smooth muscle cell (ASMC). Newborn Sprague-Dawley (SD) rats sensitized to ovalbumin (OVA) was used as the model to mimic asthmatic symptoms. The experimental results show that exposure to SO(2): (1) significantly increased Penh (an indicator of AHR) in the OVA-sensitized rats (p<0.01) but not in the normal rats (p>0.05), which correlated with the increase of airway smooth muscle mass; (2) increased IL-4 production in BALF of both the normal (p<0.05) and OVA-sensitized rats (p<0.001), but decreased IFN-γ in BALF of only the normal rats, and in serum only increased IL-4 production of the OVA-sensitized rats (p<0.001); (3) increased ASMC stiffness (G') and contractility only in the OVA-sensitized rats (p<0.001, p<0.05, respectively). Taken together, these results demonstrate that SO(2) may be a universal airway inflammatory factor, but more importantly, specific to exacerbating AHR in asthmatics only. These findings uncover a potential mechanism of SO(2)-induced health effects and may provide a basis for therapeutic targets.

Topics: Air Pollutants; Airway Remodeling; Airway Resistance; Animals; Animals, Newborn; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Interleukin-4; Male; Muscle Contraction; Muscle, Smooth; Ovalbumin; Plethysmography, Whole Body; Rats; Rats, Sprague-Dawley; Respiratory Function Tests; Respiratory Physiological Phenomena; Sulfur Dioxide

Allergic asthma is a chronic inflammatory disease, characterized by airway hyperresponsiveness (AHR), inflammation, and tissue remodeling, in which mast cells play a central role. In the present study, we analyzed how mast cell numbers and localization influence the AHR in a chronic murine model of asthma. C57BL/6 (wild-type) and mast cell-deficient B6.Cg-Kit(W-sh) mice without (Wsh) and with (Wsh+MC) mast cell engraftment were sensitized to and subsequently challenged with ovalbumin for a 91-day period. In wild-type mice, pulmonary mast cells were localized in the submucosa of the central airways, whereas the more abundant mast cells in Wsh+MC mice were found mainly in the alveolar parenchyma. In Wsh+MC, ovalbumin challenge induced a relocation of mast cells from the perivascular space and central airways to the parenchyma. Allergen challenge caused a similar AHR in wild-type and Wsh mice in the resistance of the airways and the pulmonary tissue. In Wsh+MC mice the AHR was more pronounced. The elevated functional responses were partly related to the numbers and localization of connective tissue-type mast cells in the peripheral pulmonary compartments. A mast cell-dependent increase in IgE and IL-33 together with impairment of the IL-23/IL-17 axis was evoked in Wsh and Wsh+MC mice by allergen challenge. This study shows that within the same chronic murine asthma model the development of AHR can be both dependent and independent of mast cells. Moreover, the spatial distribution and number of pulmonary mast cells determine severity and localization of the AHR.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Chronic Disease; Disease Models, Animal; Female; Immunoglobulin E; Interleukin-17; Interleukin-23; Interleukin-33; Interleukins; Lung; Mast Cells; Mice; Mice, Inbred C57BL; Ovalbumin; Severity of Illness Index

Pre-clinical evaluation of asthma therapies requires animal models of chronic airways inflammation, airway hyperresponsiveness (AHR) and lung remodelling that accurately predict drug effectiveness in human asthma. However, most animal models focus on acute allergen challenges where chronic inflammation and airway remodelling are absent. Chronic allergen challenge models have been developed in mice but few studies use guinea-pigs which may be more relevant to humans. We tested the hypothesis that a chronic rather than acute pulmonary inflammation model would best predict clinical outcome for asthma treatments. Guinea-pigs sensitized with ovalbumin (OVA) received single (acute) or nine OVA inhalation challenges at 48 h intervals (chronic). Airways function was recorded as specific airways conductance (sG(aw)) in conscious animals for 12 h after OVA challenge. AHR to inhaled histamine, inflammatory cell influx and lung histology were determined 24 h after the single or 9th OVA exposure. The inhaled corticosteroid, fluticasone propionate (FP), the phosphodiesterase 4 inhibitor, roflumilast, and the inducible nitric oxide synthase (iNOS) inhibitor, GW274150, orally, were administered 24 and 0.5 h before and 6 h after the single or final chronic OVA exposure. Both models displayed early (EAR) and late (LAR) asthmatic responses to OVA challenge, as falls in sG(aw), AHR, as increased histamine-induced bronchoconstriction, and inflammatory cell influx. Tissue remodelling, seen as increased collagen and goblet cell hyperplasia, occurred after multiple OVA challenge. Treatment with FP and roflumilast inhibited the LAR, cell influx and AHR in both models, and the remodelling in the chronic model. GW274150 also inhibited the LAR, AHR and eosinophil influx in the acute model, but not, together with the remodelling, in the chronic model. In the clinical setting, inhaled corticosteroids and phosphodiesterase 4 inhibitors are relatively effective against most features of asthma whereas the iNOS inhibitor GW274150 was ineffective. Thus, while there remain certain differences between our data and clinical effectiveness of these antiasthma drugs, a chronic pulmonary inflammation guinea-pig model does appear to be a better pre-clinical predictor of potential asthma therapeutics than an acute model.

Topics: Acute Disease; Administration, Inhalation; Administration, Oral; Aminopyridines; Androstadienes; Animals; Anti-Asthmatic Agents; Asthma; Benzamides; Bronchial Hyperreactivity; Chronic Disease; Cyclopropanes; Disease Models, Animal; Fluticasone; Guinea Pigs; Histamine; Inflammation; Male; Ovalbumin; Sulfides; Time Factors

Although the concept of "global airway allergy" has become widely accepted during recent years, nasobronchial interaction and its mechanisms remain incompletely understood. The experimental study of the effect of nasal allergen deposition on lower airway pathology is hampered by the difficulty of avoiding lower airway penetration of the allergens. In ovalbumin-sensitized mice with experimental airway allergy, nasal allergen provocations were performed after complete anatomical separation of upper and lower airways by means of a tracheotomy. A canula was inserted in the trachea, and the trachea was ligated, thus inhibiting any passage of allergens from upper to lower airways. Mice showed bronchial hyperresponsiveness to methacholine as early as 4 hours after nasal allergen provocation in the absence of recruitment of inflammatory cells. An increased substance P (SP) concentration in the bronchial lumen was found, as well as an increased number of SP-positive pulmonary nerves. Treatment with a neurokinin (NK) 1 receptor antagonist abolished the allergen-induced bronchial hyperresponsiveness. Moreover, endobronchial administration of SP caused NK1 receptor-dependent bronchial hyperresponsiveness in mice with airway allergy. Nasal allergen provocation rapidly induces bronchial hyperresponsiveness via pulmonary up-regulation of SP and activation of NK1 receptors.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Immunization; Leukocyte Count; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Nasal Provocation Tests; Neurokinin-1 Receptor Antagonists; Ovalbumin; Receptors, Neurokinin-1; Substance P; Tryptophan; Up-Regulation

Inhalation of the local anaesthetic lidocaine has been suggested to be beneficial for asthmatics, but airway anaesthesia is unpleasant and may exacerbate bronchoconstriction. Our previous study showed that inhalation of the lidocaine analogue JMF2-1 can elicit the anti-inflammatory properties of lidocaine without anaesthesia. This prompted further research on the mechanism of action and putative therapeutic application of JMF2-1.. We tested the hypothesis that JMF2-1 would prevent allergen-induced lung inflammation and airway hyperresponsiveness (AHR) by modulating T cell function in vivo and in vitro. Methods Local and systemic changes in leucocyte levels, cytokine production and lung mechanics were examined in a murine model of lung inflammation. JMF2-1 (0.05-2%) or saline was aerosolized twice a day during the ovalbumin (OVA)-provocation period (19-21 days post-sensitization). Analyses were performed 24 h after the final challenge. Primary cultured lymph node cells were used to assess the effects of JMF2-1 (100-600 μm) at the cellular level.. OVA challenge resulted in lung recruitment of CD4(+) T cells and eosinophils, increased generation of inflammatory cytokines and AHR to inhaled methacholine within 24 h. These changes were prevented by JMF2-1 nebulization, and occurred in parallel with an increase in the number of apoptotic cells in the lung. JMF2-1 treatment did not alter levels of CD4(+) or CD8(+) T cells in the thymus or lymph nodes of naïve mice, although it inhibited OVA-induced IL-13 production and the lymphocyte proliferative response in vitro. It also induced apoptosis of OVA-activated lymphocytes in a mechanism sensitive to z-VAD, indicating that JMF2-1 mediates caspase-dependent apoptosis.. Inhalation of JMF2-1 prevents the cardinal features of asthma by reducing T(H) 2 cytokine generation and lung eosinophilic inflammatory infiltrates via local inhibition of T cell function and survival. JMF2-1 may represent a novel therapeutic alternative for asthma control with distinct advantages over local anaesthetics.

Topics: Animals; Anti-Inflammatory Agents; Apoptosis; Bronchial Hyperreactivity; Cytokines; Dexamethasone; Inflammation; Lidocaine; Mice; Mice, Inbred BALB C; Mice, Transgenic; Ovalbumin; T-Lymphocytes

Astragaloside IV (AST) is the main active constituent of Radix Astragali, a Chinese herb traditionally used to prevent asthma attack from chronic asthma patients. Its efficacy and action mechanisms in asthma attack prevention remain nonetheless to be further explored. In this study, chronic asthma was induced exposing ovalbumin (OVA) sensitized mice to repeated OVA challenges twice every two weeks for 12 weeks. Mice were treated with AST for 4 weeks just after the final challenge. In this murine model of chronic asthma, the airway dysfunction and remodeling remained severe and was accompanied with suppression of the IFN-gamma level in the bronchoalveolar lavage fluid (BALF) even four weeks after the final challenge, indicating that the airway structural changes continued to develop even after interruption of OVA challenges. However, after AST treatment, the airway hyperresponsiveness was sharply relieved, accompanied by the reduction of collagen deposition and mucus production, meanwhile the inflammatory cells were decreased but the IFN-gamma level increased in BALF. In conclusion, AST could prevent the development of chronic asthma, thus reducing asthma attacks. Our results indicated that it should be used as a supplementary therapy on preventing asthma attacks from chronic asthma patients.

Topics: Animals; Asthma; Astragalus Plant; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chronic Disease; Collagen; Disease Models, Animal; Drugs, Chinese Herbal; Female; Immune System; Inflammation; Interferon-gamma; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Phytotherapy; Plant Roots; Respiratory System; Saponins; Triterpenes

Mouse models of allergic asthma are characterized by airway hyperreactivity (AHR), Th2-driven eosinophilic airway inflammation, high allergen-specific IgE (anti-OVA IgE) levels in serum, and airway remodeling. Because asthma susceptibility has a strong genetic component, we aimed to identify new asthma susceptibility genes in the mouse by analyzing the asthma phenotypes of the Leishmania major resistant (lmr) recombinant congenic (RC) strains. The lmr RC strains are derived from C57BL/6 and BALB/c intercrosses and carry congenic loci on chromosome 17 (lmr1) and 9 (lmr2) in both backgrounds. Whereas the lmr2 locus on chromosome 9 contributes to a small background-specific effect on anti-OVA IgE and AHR, the lmr1 locus on chromosome 17 mediates a strong effect on Th2-driven eosinophilic airway inflammation and background-specific effects on anti-OVA IgE and AHR. The lmr1 locus contains almost 600 polymorphic genes. To narrow down this number of candidate genes, we performed genome-wide transcriptional profiling on lung tissue from C.lmr1 RC mice and BALB/c control mice. We identified a small number of differentially expressed genes located within the congenic fragment, including a number of Mhc genes, polymorphic between BALB/c and C57Bl/6. The analysis of asthma phenotypes in the C.B10-H2b RC strain, carrying the C57Bl/6 haplotype of the Mhc locus in a BALB/c genetic background, reveals a strikingly similar asthma phenotype compared with C.lmr1, indicating that the differentially expressed genes located within the C.B10-H2b congenic fragment are the most likely candidate genes to contribute to the reduced asthma phenotypes associated with the C57Bl/6 allele of lmr1.

Topics: Airway Remodeling; Animals; Asthma; Biomarkers; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chromosome Mapping; Disease Models, Animal; Eosinophilia; Eosinophils; Female; Gene Expression Profiling; Immunoglobulin E; Inflammation; Leishmania major; Leishmaniasis; Major Histocompatibility Complex; Male; Mice; Mice, Congenic; Mice, Inbred BALB C; Mice, Inbred C57BL; Oligonucleotide Array Sequence Analysis; Ovalbumin; Phenotype; Polymorphism, Single Nucleotide; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Asthma is a chronic inflammatory disease of the lungs, characterized by increased sensitivity to bronchoconstriction associated with infiltration of immune cells, mucus hypersecretion and structural remodelling of the airways. In South Africa, the indigenous plant Siphonochilus aethiopicus, is used by traditional health practitioners to treat colds, wheezing of the chest, coughs, influenza, sinus problems and mild asthma. In this study we aimed to investigate the potential anti-inflammatory and anti-allergic properties of S. aethiopicus in vitro and its efficacy in a mouse model of allergic asthma.. The dried and powdered S. aethiopicus plant material was extracted separately with organic solvents (diethyl ether, ethanol) and water. Dried extracts as well as a purified furanoterpenoid compound present in the extracts were screened in vitro in a glucocorticoid and histamine H(1) receptor binding assay and a phosphodiesterase IV enzyme inhibition assay. Extracts were also evaluated for efficacy against ovalbumin (OVA)-induced allergic airway disease in mice.. Biological assaying of extracts of the plant and the isolated furanoterpenoid showed significant in vitro inhibition of glucocorticoid and histamine H(1) receptor binding and phosphodiesterase IV activity, supporting a possible anti-inflammatory, anti-allergic and bronchodilatory effect. Administration of S. aethiopicus extracts to OVA-sensitized and challenged mice significantly reduced lung inflammation and the percentage of eosinophils in bronchoalveolar lavage fluid but did not influence airway hyperreactivity.. This study provides evidence that S. aethiopicus has anti-inflammatory and anti-allergic properties in vitro and in vivo. These findings may support anecdotal accounts of its effectiveness against asthma, sinusitis, colds and flu.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Ethnopharmacology; Humans; In Vitro Techniques; Lung; Medicine, African Traditional; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plant Extracts; Plants, Medicinal; South Africa; Zingiberaceae

Adult bone marrow-derived mesenchymal stem cells (MSC) possess potent immune modulatory effects which support their possible use as a therapy for immune-mediated disease. MSC induce regulatory T cells (T(reg)) in vitro although the in vivo relevance of this is not clear.. This study addressed the hypothesis that adult bone marrow derived-MSC would prevent the pathology associated with allergen-driven airway inflammation, and sought to define the effector mechanism.. The influence of allogeneic MSC was examined in a model system where T(reg) induction is essential to prevent pathology. This was tested using a combination of a model of ovalbumin-driven inflammation with allogeneic MSC cell therapy.. Systemic administration of allogeneic MSC protected the airways from allergen-induced pathology, reducing airway inflammation and allergen-specific IgE. MSC were not globally suppressive but induced CD4(+) FoxP3(+) T cells and modulated cell-mediated responses at a local and systemic level, decreasing IL-4 but increasing IL-10 in bronchial fluid and from allergen re-stimulated splenocytes. Moderate dose cyclophosphamide protocols were used to differentially ablate T(reg) responses; under these conditions the major beneficial effect of MSC therapy was lost, suggesting induction of T(reg) as the key mechanism of action by MSC in this model. In spite of the elimination of T(reg) , a significant reduction in airway eosinophilia persisted in those treated with MSC.. These data demonstrate that MSC induce T(reg) in vivo and reduce allergen-driven pathology. Multiple T(reg) dependent and independent mechanisms of therapeutic action are employed by MSC.

Topics: Animals; Antigens; Bronchial Hyperreactivity; Female; Inflammation; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes, Regulatory

Th2 cytokines and their downstream Janus kinase (JAK)-signal transducer and activation of transcription (STAT) pathways play a critical role in allergic asthma. We studied the effects of a pan-JAK inhibitor, pyridone 6 (P6), on asthmatic responses in a mouse model and investigated the mechanism for its biological effects. Mice were sensitized and challenged by ovalbumin (OVA). P6 treatment during the challenge phase suppressed eosinophilia in bronchoalveolar lavage (BAL) fluids but did not affect airway hyperresponsiveness (AHR). To improve the efficacy of the JAK inhibitor, P6 was encapsulated in polylactic-coglycolic acid nanoparticles (P6-PLGA). P6-PLGA treatment just before OVA challenge suppressed both airway eosinophilia and AHR. Although the IL-13 levels in BAL fluids and the OVA-specific IgE levels in serum after the challenge phase treatment with P6-PLGA were similar to those after a sham treatment, the eotaxin levels in BAL fluids and lung mCLCA3/Gob-5 expression were decreased in P6-PLGA-treated mice. Interestingly, the local IL-13 levels and serum OVA-specific IgE were decreased, while IL-17-producing T cells were increased by P6-PLGA treatment during the sensitization plus challenge phases. In vitro, P6 strongly suppressed the differentiation of Th2 from naive CD4 T cells, but it partly enhanced Th17 differentiation. P6 potently suppressed IL-13-mediated STAT6 activation and mCLCA3/Gob-5 expression in mouse tracheal epithelial cells. These findings suggest that the JAK inhibitor P6 suppresses asthmatic responses by inhibiting Th2 inflammation and that application of PLGA nanoparticles improves the therapeutic potency of P6.

Topics: Animals; Asthma; Benzimidazoles; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Capsules; Chloride Channels; Eosinophilia; Interleukin-13; Janus Kinases; Lactic Acid; Lung; Mice; Mucoproteins; Nanoparticles; Ovalbumin; Polyglycolic Acid; Polylactic Acid-Polyglycolic Acid Copolymer; Protein Kinase Inhibitors; Pyridones; STAT6 Transcription Factor; Th2 Cells

Asthma leads to structural changes in the airways, including the modification of extracellular matrix proteins such as tenascin-C. The role of tenascin-C is unclear, but it might act as an early initiator of airway wall remodelling, as its expression is increased in the mouse and human airways during allergic inflammation. In this study, we examined whether Th1 or Th2 cells are important regulators of tenascin-C in experimental allergic asthma utilizing mice with impaired Th1 (STAT4-/-) or Th2 (STAT6-/-) immunity.. Balb/c wildtype (WT), STAT4-/- and STAT6-/- mice were sensitized with intraperitoneally injected ovalbumin (OVA) followed by OVA or PBS airway challenge. Airway hyperreactivity (AHR) was measured and samples were collected. Real time PCR and immunohistochemistry were used to study cytokines and differences in the expression of tenascin-C. Tenascin-C expression was measured in human fibroblasts after treatment with TNF-α and IFN-γ in vitro.. OVA-challenged WT mice showed allergic inflammation and AHR in the airways along with increased expression of TNF-α, IFN-γ, IL-4 and tenascin-C in the lungs. OVA-challenged STAT4-/- mice exhibited elevated AHR and pulmonary eosinophilia. The mRNA expression of TNF-α and IFN-γ was low, but the expression of IL-4 was significantly elevated in these mice. OVA-challenged STAT6-/- mice had neither AHR nor pulmonary eosinophilia, but had increased expression of mRNA for TNF-α, IFN-γ and IL-4. The expression of tenascin-C in the lungs of OVA-challenged STAT4-/- mice was weaker than in those of OVA-challenged WT and STAT6-/- mice suggesting that TNF-α and IFN-γ may regulate tenascin-C expression in vivo. The stimulation of human fibroblasts with TNF-α and IFN-γ induced the expression of tenascin-C confirming our in vivo findings.. Expression of tenascin-C is significantly attenuated in the airways of STAT4-/- mice, which may be due to the impaired secretion of TNF-α and IFN-γ in these mice.

Topics: Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Disease Models, Animal; Female; Fibroblasts; Humans; Interferon-gamma; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; RNA, Messenger; STAT4 Transcription Factor; STAT6 Transcription Factor; Tenascin; Th1 Cells; Th2 Cells; Tumor Necrosis Factor-alpha; Up-Regulation

Erlotinib, an EGFR tyrosine kinase inhibitor, can inhibit the proliferation and survival of cancer cells. It has been widely used to treat non-small cell lung cancer. This study aimed to evaluate the effects of erlotinib on bronchial hyperresponsiveness, airway inflammation, and airway remodeling in sensitized, ovalbumin-challenged rats. Two experimental groups of Brown-Norway rats were sensitized and repeatedly challenged by breathing aerosolized ovalbumin. Since Day 1, one group was given oral erlotinib (OA-erlotinib group) while the other group was given only oral saline (OA-saline group). The control group was sensitized and challenged using saline. All were anesthetized and paralyzed, and pulmonary function tests conducted at baseline and after provocation with varying doses of acetylcholine. Lung tissues were examined for airway inflammation, airway remodeling, and Th2-related cytokine mRNA expression. Results showed that the OA-erlotinib group had better pulmonary function and less airway inflammation, Th2-related cytokines and their mRNA expression, and airway remodeling compared to the OA-saline group. In conclusion, erlotinib effectively prevents bronchial hyperreactivity, airway inflammation, Th2-related cytokine mRNA expression, and airway remodeling after sensitization and repeated allergen challenge in Brown-Norway rats.

Topics: Airway Remodeling; Allergens; Animals; Bronchial Hyperreactivity; Erlotinib Hydrochloride; Immunohistochemistry; Ovalbumin; Pneumonia; Protein Kinase Inhibitors; Quinazolines; Rats; Rats, Inbred BN; Reverse Transcriptase Polymerase Chain Reaction

Thymic stromal lymphopoietin (TSLP) is an essential cytokine for the initiation and development of allergic inflammation. In this study, we have investigated the role of TSLP in the breakdown of immune tolerance and generation of inducible regulatory T cells (iTregs). Our results demonstrated that TSLP diverted airway tolerance against OVA to Th2 sensitization and inhibited the generation of OVA-specific iTregs. TSLP exerted a direct inhibitory effect on both human and mouse iTreg development in vitro. Low doses of TSLP were capable of inhibiting iTreg induction without significantly promoting Th2 development, indicating that these two functions of TSLP are separable. Moreover, the TSLP-mediated inhibition of iTreg generation was only partially dependent on IL-4 and Stat6, and was effective when TSLP was present for the first 24 h of T cell activation. These results define a novel role for TSLP in regulating the balance of airway tolerance and allergic inflammation.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Epitopes, T-Lymphocyte; Gene Knock-In Techniques; Growth Inhibitors; Humans; Immune Tolerance; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Ovalbumin; Stromal Cells; T-Lymphocytes, Regulatory; Th2 Cells; Thymic Stromal Lymphopoietin; Thymus Gland

Intravenous immunoglobulin (IVIG) has potent anti-inflammatory and immune-modulating properties. IVIG has been utilized as a steroid-sparing agent in severe asthma, but the results of clinical trials have been conflicting.. To determine whether IVIG is able to attenuate bronchial reactivity, pulmonary inflammation and T cell function using a murine model of allergic airways disease.. BALB/c or C57BL/6 mice were sensitized to ovalbumin (OVA) or a phosphate-buffered saline control using local nasal sensitization, and then received five intranasal challenges on days 28-32 before sacrifice. Mice were treated intraperitoneally with either IVIG (1-2 g/kg) or equivalent human serum albumin 24 h before the first OVA challenge. Bronchial reactivity to methacholine was examined using the FlexiVent small animal ventilator. We evaluated pulmonary histology, mRNA from lung digests for T-helper type 2 (Th2)-related genes and bronchoalveolar lavage for cell counts and cytokines. Splenocytes were utilized to study OVA-induced cell proliferation, cytokine production and dendritic cell maturation.. IVIG markedly attenuated the perivascular and peribronchial pulmonary inflammation, and decreased bronchial hyperresponsiveness to methacholine. IVIG treatment of splenocytes from sensitized animals diminished cellular proliferation to OVA, whereas IVIG treatment in vivo markedly attenuated OVA-driven splenocyte proliferation. This is accompanied by diminished IL-13 and TNF-α levels in splenocyte culture, decreased expression of Jagged-1, increased Delta-4 and decreased GATA-3 mRNA levels, signs that IVIG has suppressed the expected Th2 response that accompanies repeated allergen exposure. Increased regulatory T cells were found in draining pulmonary lymph nodes in IVIG-treated mice but not in controls.. IVIG was effective in ameliorating allergic airway disease in our model. IVIG may be a promising adjunct therapy requiring further study for patients with severe asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Humans; Immunoglobulins, Intravenous; Injections, Intraperitoneal; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Models, Immunological; Ovalbumin; Serum Albumin; T-Lymphocytes, Regulatory

Glucocorticoids (GCs), which activate GC receptor (GR) signaling and thus modulate gene expression, are widely used to treat asthma. GCs exert their therapeutic effects in part through modulating airway smooth muscle (ASM) structure and function. However, the effects of genes that are regulated by GCs on airway function are not fully understood. We therefore used transcription profiling to study the effects of a potent GC, dexamethasone, on human ASM (HASM) gene expression at 4 and 24 hours. After 24 hours of dexamethasone treatment, nearly 7,500 genes had statistically distinguishable changes in expression; quantitative PCR validation of a 40-gene subset of putative GR-regulated genes in 6 HASM cell lines suggested that the early transcriptional targets of GR signaling are similar in independent HASM lines. Gene ontology analysis implicated GR targets in controlling multiple aspects of ASM function. One GR-regulated gene, the transcription factor, Kruppel-like factor 15 (Klf15), was already known to modulate vascular smooth and cardiac muscle function, but had no known role in the lung. We therefore analyzed the pulmonary phenotype of Klf15(-/-) mice after ovalbumin sensitization and challenge. We found diminished airway responses to acetylcholine in ovalbumin-challenged Klf15(-/-) mice without a significant change in the induction of asthmatic inflammation. In cultured cells, overexpression of Klf15 reduced proliferation of HASM cells, whereas apoptosis in Klf15(-/-) murine ASM cells was increased. Together, these results further characterize the GR-regulated gene network in ASM and establish a novel role for the GR target, Klf15, in modulating airway function.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cell Line; Cell Proliferation; Dexamethasone; DNA-Binding Proteins; Female; Gene Expression Regulation; Glucocorticoids; Humans; Kruppel-Like Transcription Factors; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Nuclear Proteins; Oligonucleotide Array Sequence Analysis; Ovalbumin; Signal Transduction; Transcription Factors

Asthma is an inflammatory disease of the lungs that is characterised by increased inflammatory cell infiltration into the airways and poor respiratory function. Ivermectin is a semi-synthetic derivative of a family of macrocyclic lactones that shows broad-spectrum anti-parasitic activity. This drug has been shown to possess anti-inflammatory activity, but whether it can be used in asthma treatment has not yet been investigated. In this study, we aimed to investigate the inhibitory effects of ivermectin on allergic asthma symptoms in mice.. We used a mouse asthma model, in which allergic airway inflammation and airway remodelling were induced by ovalbumin (OVA) sensitisation and challenge. Ivermectin or PBS treatment was administered 1 h before OVA challenge. Ivermectin at 2 mg/kg significantly diminished recruitment of immune cells, production of cytokines in the bronchoalveolar lavage fluids and secretion of OVA-specific IgE and IgG1 in the serum. Histological studies indicated that ivermectin suppressed mucus hypersecretion by goblet cells in the airway.. This is the first study to demonstrate that ivermectin is an effective suppressor of inflammation and may be efficacious in the treatment of non-infectious airway inflammatory diseases such as allergic asthma.

Topics: Animals; Anti-Allergic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cytokines; Dexamethasone; Disease Models, Animal; Female; Immunoglobulin E; Immunoglobulin G; Ivermectin; Lung; Mice; Mice, Inbred BALB C; Ovalbumin

Eosinophils are found in the lungs of humans with allergic asthma, as well as in the lungs of animals in models of this disease. Increasing evidence suggests that these cells are integral to the development of allergic asthma in C57BL/6 mice. However, the specific function of eosinophils that is required for this event is not known. In this study, we experimentally validate a dynamic computational model and perform follow-up experimental observations to determine the mechanism of eosinophil modulation of T cell recruitment to the lung during development of allergic asthma. We find that eosinophils deficient in IL-13 were unable to rescue airway hyperresponsiveness, T cell recruitment to the lungs, and Th2 cytokine/chemokine production in ΔdblGATA eosinophil-deficient mice, even if Th2 cells were present. However, eosinophil-derived IL-13 alone was unable to rescue allergic asthma responses in the absence of competence of other IL-13-producing cells. We further computationally investigate the role of other cell types in the production of IL-13, which led to the various predictions including early and late pulses of IL-13 during airway hyperresponsiveness. These experiments suggest that eosinophils and T cells have an interdependent relationship, centered on IL-13, which regulates T cell recruitment to the lung and development of allergic asthma.

Topics: Airway Resistance; Animals; Bronchial Hyperreactivity; Cell Movement; Computer Simulation; Disease Models, Animal; Eosinophils; Inflammation; Interleukin-13; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Models, Immunological; Ovalbumin; Random Allocation; Respiratory Hypersensitivity; Th2 Cells

Arginase overexpression contributes to airways hyperresponsiveness (AHR) in asthma. Arginase expression is further augmented in cigarette smoking asthmatics, suggesting that it may be upregulated by environmental pollution. Thus, we hypothesize that arginase contributes to the exacerbation of respiratory symptoms following exposure to air pollution, and that pharmacologic inhibition of arginase would abrogate the pollution-induced AHR.. To investigate the role of arginase in the air pollution-induced exacerbation of airways responsiveness, we employed two murine models of allergic airways inflammation. Mice were sensitized to ovalbumin (OVA) and challenged with nebulized PBS (OVA/PBS) or OVA (OVA/OVA) for three consecutive days (sub-acute model) or 12 weeks (chronic model), which exhibit inflammatory cell influx and remodeling/AHR, respectively. Twenty-four hours after the final challenge, mice were exposed to concentrated ambient fine particles plus ozone (CAP+O₃), or HEPA-filtered air (FA), for 4 hours. After the CAP+O₃ exposures, mice underwent tracheal cannulation and were treated with an aerosolized arginase inhibitor (S-boronoethyl-L-cysteine; BEC) or vehicle, immediately before determination of respiratory function and methacholine-responsiveness using the flexiVent®. Lungs were then collected for comparison of arginase activity, protein expression, and immunohistochemical localization.. Compared to FA, arginase activity was significantly augmented in the lungs of CAP+O₃-exposed OVA/OVA mice in both the sub-acute and chronic models. Western blotting and immunohistochemical staining revealed that the increased activity was due to arginase 1 expression in the area surrounding the airways in both models. Arginase inhibition significantly reduced the CAP+O₃-induced increase in AHR in both models.. This study demonstrates that arginase is upregulated following environmental exposures in murine models of asthma, and contributes to the pollution-induced exacerbation of airways responsiveness. Thus arginase may be a therapeutic target to protect susceptible populations against the adverse health effects of air pollution, such as fine particles and ozone, which are two of the major contributors to smog.

Topics: Animals; Arginase; Asthma; Blotting, Western; Boronic Acids; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Disease Models, Animal; Enzyme Inhibitors; Female; Immunohistochemistry; Inflammation Mediators; Inhalation Exposure; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Oxidative Stress; Ozone; Particulate Matter; Up-Regulation

Tumor Necrosis Factor (TNF) is a pleiotropic cytokine consisting of soluble and transmembrane forms, with distinct roles in inflammation and immunity. TNF is an important factor in allergic airway inflammation. However, the disparate functions of soluble (sol) and transmembrane (tm) TNF in lung pathology are not well understood. Our aim was to assess the activities of solTNF and tmTNF in murine models of allergic airway disease, and to evaluate the efficacy of solTNF-selective inhibition. We used ovalbumin sensitization and challenge of TNF knockout, tmTNF knockin, and wild-type C57BL/6 mice to distinguish differences in airway inflammation and hyperreactivity mediated by solTNF and tmTNF. Functions of solTNF and tmTNF in hyperresponsive, wild-type Balb/c mice were assessed by comparing dominant-negative anti-TNF biologics, which antagonize solTNF yet spare tmTNF, to etanercept, a nonselective inhibitor of both TNF forms. Responses in transgenic C57BL/6 mice demonstrated that solTNF, and not tmTNF, is necessary to drive airway inflammation. In Balb/c mice, dominant-negative TNF biologics administered during immunization decreased the recruitment of eosinophils and lymphocytes into the bronchoalveolar space and lung parenchyma, reduced specific serum IgE, goblet-cell hyperplasia, and eosinophilic inflammation, and suppressed methacholine-induced airway hyperreactivity. Concentrations of IL-5, CCL5/RANTES, CCL11/eotaxin, and CCL17/TARC were also reduced in bronchoalveolar lavage. Dominant-negative TNFs reduced lung eosinophilia, even when given only during antigen challenge. The selective inhibition of soluble TNF suppresses inflammation, hyperreactivity, and remodeling in transgenic and wild-type murine models of allergic airway disease, and may offer safety advantages in therapies that preserve the immunoprotective functions of transmembrane TNF.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Biological Products; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Etanercept; Goblet Cells; Humans; Hyperplasia; Immunoglobulin E; Immunoglobulin G; Inflammation Mediators; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Mutation; Ovalbumin; Pneumonia; Pulmonary Eosinophilia; Receptors, Tumor Necrosis Factor; Tumor Necrosis Factor-alpha

Airway remodelling, characterised by increased airway smooth muscle (ASM) mass, subepithelial fibrosis, goblet cell hyperplasia and mucus gland hypertrophy, is a feature of chronic asthma. Increased arginase activity could contribute to these features via increased formation of polyamines and l-proline downstream of the arginase product l-ornithine, and via reduced nitric oxide synthesis. Using the specific arginase inhibitor 2(S)-amino-6-boronohexanoic acid (ABH), we studied the role of arginase in airway remodelling using a guinea pig model of chronic asthma. Ovalbumin-sensitised guinea pigs were treated with ABH or PBS via inhalation before each of 12 weekly allergen or saline challenges, and indices of arginase activity, and airway remodelling, inflammation and responsiveness were studied 24 h after the final challenge. Pulmonary arginase activity of repeatedly allergen-challenged guinea pigs was increased. Allergen challenge also increased ASM mass and maximal contraction of denuded tracheal rings, which were prevented by ABH. ABH also attenuated allergen-induced pulmonary hydroxyproline (fibrosis) and putrescine, mucus gland hypertrophy, goblet cell hyperplasia, airway eosinophilia and interleukin-13, whereas an increased l-ornithine/l-citrulline ratio in the lung was normalised. Moreover, allergen-induced hyperresponsiveness of perfused tracheae was fully abrogated by ABH. These findings demonstrate that arginase is prominently involved in allergen-induced airway remodelling, inflammation and hyperresponsiveness in chronic asthma.

Topics: Airway Remodeling; Allergens; Aminocaproates; Animals; Anti-Asthmatic Agents; Arginase; Asthma; Boron Compounds; Bronchial Hyperreactivity; Chronic Disease; Citrulline; Eosinophilia; Exocrine Glands; Goblet Cells; Guinea Pigs; Interleukin-13; Lung; Male; Muscle Contraction; Muscle, Smooth; Ornithine; Ovalbumin; Pulmonary Fibrosis; Trachea

Despite their increasing use in autoimmune, inflammatory, and allergic conditions, the mechanism of action of i.v. Igs (IVIg) is poorly understood. On the basis of the critical role of invariant NKT (iNKT) cells in allergic airway inflammation (AAI) and their constitutive expression of the low-affinity IgG receptor FcγRIIIA, we surmised that IVIg targets iNKT cells to exert their anti-inflammatory effect. We found that IVIg treatment significantly inhibited AAI in OVA-sensitized C57BL/6 mice and downregulated α-galactosylceramide-induced iNKT cell activation and cytokine production. Allergic responses were restored in iNKT cell-deficient mice by transferring iNKT cells from PBS- but not from IVIg-treated mice, suggesting that IVIg acts directly on activated iNKT cells that have a critical role in AAI. The inhibitory effects of IVIg on both iNKT cell activation/function and OVA-driven AAI were lost in FcγRIIIA(-/-) mice. Our data unravel an FcγRIIIA-dependent inhibitory effect of IVIg on activated iNKT cells that confers protection in AAI.

Topics: Adoptive Transfer; Allergens; Animals; Anti-Inflammatory Agents, Non-Steroidal; Bronchial Hyperreactivity; Cytokines; Galactosylceramides; Immunoglobulins, Intravenous; Inflammation Mediators; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Natural Killer T-Cells; Ovalbumin; Receptors, IgG; Respiratory Hypersensitivity; Spleen

Epidemiologic studies have associated higher dietary consumption of soy isoflavones with decreased self-report of cough and allergic respiratory symptoms, but the pharmacodynamic effects of soy isoflavone on asthmatic model have not been well-described. Here, we hypothesized that soy isoflavone may have potential effects on airway hyperresponsiveness, inflammation and airway remodeling in a murine of asthma. Mice sensitized and challenged with ovalbumin developed airway inflammation. Bronchoalveolar lavage fluid was assessed for inflammatory cell counts, and for cytokine levels. Lung tissues were examined for cell infiltration, mucus hypersecretion and airway remodeling, and for the expression of inflammatory biomarkers. Airway hyperresponsiveness was monitored by direct airway resistance analysis. Oral administration of soy isoflavone significantly reduced ovalbumin-induced airway hyperresponsiveness to intravenous methacholine, and inhibited ovalbumin-induced increases in eosinophil counts. RT-PCR analysis of whole lung lysates revealed that soy isoflavone markedly suppressed ovalbumin-induced mRNA expression of eotaxin, interleukin(IL)-5, IL-4 and matrix metalloproteinase-9, and increased mRNA expression of interferon (IFN)-γ and tissue inhibitor of metalloproteinase-1 in a dose-dependent manner. Soy isoflavone also substantially recovered IFN-γ/IL-4 (Th1/Th2) levels in bronchoalveolar lavage fluid. In addition, histologic studies showed that soy isoflavone dramatically inhibited ovalbumin-induced lung tissue eosinophil infiltration, airway mucus production and collagen deposition in lung tissues. Our findings suggest that soy isoflavone as nutritional supplement may provide a novel means for the treatment of airway inflammatory disease.

Topics: Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Collagen; Eosinophils; Female; Glycine max; Hypersensitivity; Inflammation; Interferon-gamma; Interleukin-4; Interleukin-5; Isoflavones; Lung; Matrix Metalloproteinase 9; Methacholine Chloride; Mice; Mice, Inbred ICR; Mucus; Ovalbumin; Peroxidase; Respiratory System; Superoxide Dismutase; Tissue Inhibitor of Metalloproteinase-1

Zinc is an essential element required for the cell metabolism, including gene transcription, signal transduction, immunity, and apoptosis. The pathophysiological role of zinc in asthma, however, is not entirely clear. Mast cells have been implicated in atopic asthma, and zinc deprivation has been reported to reduce mast cell activation. Here, we investigate the effects of a zinc chelator, N,N,N',N'-tetrakis (2-pyridyl-methyl) ethylenediamine (TPEN), on asthmatic responses in mouse models of ovalbumin (OVA)-induced airway hyperresponsiveness and allergic airway inflammation.. Mice were sensitized with OVA with or without the adjuvant aluminum hydroxide (alum) and subjected to OVA exposure with or without treatment of TPEN. Cell profiles and cytokine levels in bronchoalveolar lavage (BAL) fluids, airway responsiveness to inhaled acetylcholine, and goblet cell hyperplasia after allergen exposure were assessed.. In mice sensitized to OVA without alum, TPEN significantly suppressed airway hyperresponsiveness and eosinophilia in BAL fluids. TPEN also attenuated the upregulation of TNFα, IL-13 and IL-4 in BAL fluids and goblet cell hyperplasia after OVA exposure. By contrast, in mice sensitized to OVA with alum, TPEN suppressed eosinophilia in BAL fluids but not airway hyperresponsiveness and goblet cell hyperplasia.. In pulmonary allergic inflammation induced in mice immunized with antigen without alum, zinc chelator inhibits airway inflammation and hyperresponsiveness. These findings suggest that zinc may be a therapeutic target of allergic asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chelating Agents; Cytokines; Disease Models, Animal; Eosinophilia; Ethylenediamines; Goblet Cells; Hyperplasia; Immunoglobulin E; Inflammation; Interleukin-13; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Respiratory System; Zinc

Inhaled short acting β2-agonists (SABA), e.g. albuterol, are used for quick reversal of bronchoconstriction in asthmatics. While SABA are not recommended for maintenance therapy, it is not uncommon to find patients who frequently use SABA over a long period of time and there is a suspicion that long term exposure to SABA could be detrimental to lung function. To test this hypothesis we studied the effect of long-term inhaled albuterol stereoisomers on immediate allergic response (IAR) and airway hyperresponsiveness (AHR) in mouse models of asthma.. Balb/C mice were sensitized and challenged with ovalbumin (OVA) and then we studied the IAR to inhaled allergen and the AHR to inhaled methacholine. The mice were pretreated with nebulizations of either racemic (RS)-albuterol or the single isomers (S)- and (R)-albuterol twice daily over 7 days prior to harvest.. We found that all forms of albuterol produced a significant increase of IAR measured as respiratory elastance. Similarly, we found that AHR was elevated by albuterol. At the same time a mouse strain that is intrinsically hyperresponsive (A/J mouse) was not affected by the albuterol isomers nor was AHR induced by epithelial disruption with Poly-L-lysine affected by albuterol.. We conclude that long term inhalation treatment with either isomer of albuterol is capable of precipitating IAR and AHR in allergically inflamed airways but not in intrinsically hyperresponsive mice or immunologically naïve mice. Because (S)-albuterol, which lacks affinity for the β2-receptor, did not differ from (R)-albuterol, we speculate that isomer-independent properties of the albuterol molecule, other than β2-agonism, are responsible for the effect on AHR.

Topics: Administration, Inhalation; Adrenergic beta-2 Receptor Agonists; Albuterol; Analysis of Variance; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Bronchodilator Agents; Disease Models, Animal; Drug Administration Schedule; Female; Isomerism; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Nebulizers and Vaporizers; Ovalbumin; Receptors, Adrenergic, beta; Respiratory Mechanics; Time Factors

Inducible co-stimulator ligand (ICOSL) is a rather newly defined co-stimulatory molecule, which, through interaction with ICOS expressed on T cells, plays an important role in T-cell activation, differentiation and function. T(h)2-type immune responses are critical for the development and maintenance of allergic responses including asthma. Using knockout (KO) mice, we have assessed the role of ICOSL in allergic airway inflammation and responsiveness using a standard mouse asthma model induced by ovalbumin (OVA) sensitization and challenge. Our data show that OVA-treated ICOSL KO mice exhibit significantly less lung eosinophilic infiltration, histopathology, mucus production and virtually no airway hyperresponsiveness in contrast to wild-type (Wt) counterparts. Serum antibody analysis showed that antigen-specific IgG1, IgG2a and IgE titers in ICOSL KO mice were significantly lower than those of Wt controls. Also, CD4(+) T cells isolated from ICOSL KO mice produced less T(h)2 cytokines (IL-4, IL-5, IL-10 and IL-13) but more T(h)1 (IFN-γ) and IL-17 than their Wt controls. Taken together, we conclude that ICOSL plays an important role in predisposing individuals to allergic airway hyperresponsiveness by enhancing IgE antibody class switching and T(h)2 cytokine production and diminishing the T(h)17 response and airway eosinophilia.

Topics: Animals; Bronchial Hyperreactivity; Cells, Cultured; Cytokines; Disease Models, Animal; Female; Humans; Immunity, Humoral; Immunization; Immunoglobulin E; Inducible T-Cell Co-Stimulator Ligand; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Proteins; Respiratory Hypersensitivity; Th17 Cells

Inflammation and oxidative stress are associated with airway diseases. There is growing evidence that Atorvastatin could be used as a therapy for these conditions.. On these bases, we evaluated Atorvastatin as a protective and reversal treatment for the allergic airway diseases in mice model. We also looked at the possible interaction with the currently used effective medication.. Mice were sensitized and challenged with ovalbumin (OVA) to develop features of allergic airway diseases mainly of bronchial inflammation. Atorvastatin was injected during or after the sensitization and challenge process to evaluate its protective or reversal effects, respectively. Total and differential cells in the BAL fluids together with IL-4, IL-5 and IL-10 cytokine levels were evaluated. Total IgE and cholesterol levels in serum were studied.. In the protective phase, Atorvastatin inhibited the OVA-induced cellular infiltration of lung bronchi, decreased IL-4 and IL-5 and prevented the increase in IL-10 cytokine levels. Also, it reduced the OVA-induced high serum total IgE level. Injection of Atorvastatin after challenge was not effective in reversing the inflammatory process, with no major contribution towards augmenting the actions of Dexamethasone. The cholesterol lowering effect was marked in the protective phase while less effective for the reversal phase.. Our results indicate that Atorvastatin reduced the allergic inflammatory features in mice and it could be useful towards developing a better therapeutic regimen for the treatment of allergic diseases.

Topics: Animals; Anti-Inflammatory Agents; Atorvastatin; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cholesterol; Cytokines; Dexamethasone; Drug Interactions; Heptanoic Acids; Immunoglobulin E; Inflammation; Leukocytes; Male; Mice; Ovalbumin; Pyrroles; Respiratory System

Most of the studies investigating the effectiveness of blocking the leukotriene B4 (LTB4) receptor 1 (BLT1) have been performed in models of primary or acute allergen challenge. The role of the LTB4-BLT1 pathway in secondary challenge models, where airway hyperresponsiveness (AHR) and airway inflammation have been established, has not been defined. We investigated the effects of blocking BLT1 on early- and late-phase development of AHR and airway inflammation in previously sensitized and challenged mice. Female BALB/c mice were sensitized (Days 1 and 14) and challenged (primary, Days 28-30) with ovalbumin. On Day 72, mice were challenged (secondary) with a single OVA aerosol, and the early and late phases of AHR and inflammation were determined. Specific blockade of BLT1 was attained by oral administration of a BLT1 antagonist on Days 70 through 72. Administration of the antagonist inhibited the secondary ovalbumin challenge-induced alterations in airway responses during the late phase but not during the early phase, as demonstrated by decreases in AHR and in bronchoalveolar lavage neutrophilia and eosinophilia 6 and 48 hours after secondary challenge. The latter was associated with decreased levels of KC protein, macrophage inflammatory protein 2, and IL-17 in the airways. These data identify the importance of the LTB4-BLT1 pathway in the development of late-phase, allergen-induced airway responsiveness after secondary airway challenge in mice with established airway disease.

Topics: Animals; Anti-Asthmatic Agents; Antibodies; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Chemokine CXCL1; Chemokine CXCL2; Disease Models, Animal; Female; Inflammation Mediators; Interleukin-17; Leukotriene Antagonists; Lung; Mice; Mice, Inbred BALB C; Neutrophil Infiltration; Ovalbumin; Pneumonia; Pulmonary Eosinophilia; Receptors, Leukotriene B4; Time Factors

Allergic asthma is characterized by hyperresponsiveness and inflammation of the airway with increased expression of inducible nitric oxide synthase (iNOS) and overproduction of nitric oxide (NO). Grape seed proanthocyanidin extract (GSPE) has been proved to have antioxidant, antitumor, anti-inflammatory, and other pharmacological effects. The purpose of this study was to examine the role of GSPE on airway inflammation and hyperresponsiveness in a mouse model of allergic asthma. BALB/c mice, sensitized and challenged with ovalbumin (OVA), were intraperitoneally injected with GSPE. Administration of GSPE remarkably suppressed airway resistance and reduced the total inflammatory cell and eosinophil counts in BALF. Treatment with GSPE significantly enhanced the interferon (IFN)- γ level and decreased interleukin (IL)-4 and IL-13 levels in BALF and total IgE levels in serum. GSPE also attenuated allergen-induced lung eosinophilic inflammation and mucus-producing goblet cells in the airway. The elevated iNOS expression observed in the OVA mice was significantly inhibited by GSPE. In conclusion, GSPE decreases the progression of airway inflammation and hyperresponsiveness by downregulating the iNOS expression, promising to have a potential in the treatment of allergic asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophils; Female; Grape Seed Extract; Immunoglobulin E; Inflammation; Interferon-gamma; Interleukin-13; Interleukin-4; Lung; Lymphocytes; Macrophages; Mice; Mice, Inbred BALB C; Neutrophils; Nitric Oxide Synthase Type II; Ovalbumin; Proanthocyanidins; Random Allocation; Vitis

In this study the effects of non-selective PDE inhibitors (theophylline and theobromine) and selective inhibitors of PDE 1, 3, 4 and 5 on cough, induced by citric acid, were evaluated. Inhalation of citric acid aerosol was used for cough provocation in healthy and ovalbumin-sensitized guinea pigs and the number of cough efforts was registered after visual and acoustic control by a skilled observer, with subsequent evaluation of airflow changes in a double chamber whole body plethysmograph. The pre-treatment with theophylline and theobromine (10 mg/kg b.w. intraperitoneally) decreased the number of cough efforts evoked by inhalation of citric acid aerosol (0.6 mol/l) in both healthy and ovalbumin-sensitized animals. The selective inhibitors (all 1 mg/kg b.w. intraperitoneally) of PDE1 (vinpocetin), PDE3 (cilostazol), and PDE4 (citalopram) showed antitussive effects in healthy guinea pigs. Conversely, the antitussive potential of PDE1 (vinpocetin), PDE4 (citalopram), and PDE5 (zaprinast) was observed in ovalbumin-sensitized animals. We conclude that the administration of non-selective PDE inhibitors influenced the citric acid-induced cough both in healthy and ovalbumin-sensitized guinea pigs, indicating the participation of a bronchodilating action and suppression of airway hyperreactivity in the cough suppression. With selective inhibitors, PDE4 inhibition seems to be the most effective in cough suppression, confirming its positive effects tested in chronic airway inflammatory diseases associated with bronchoconstriction and cough (Fig. 6, Ref. 27).

Topics: Animals; Antitussive Agents; Bronchial Hyperreactivity; Citric Acid; Cough; Guinea Pigs; Male; Ovalbumin; Phosphodiesterase Inhibitors; Reflex; Theobromine; Theophylline

The competition between arginases and NO synthases (NOS) for their common substrate L-arginine can be important in the airways hyperreactivity. We investigated the effect of the simultaneous modulation of arginase and NOS activities in allergen-induced airways hyperreactivity. We analysed the response of tracheal and lung tissue smooth muscle to histamine or acetylcholine after administration N(ω)-nitro-L-arginine methyl ester (L-NAME), aminoguanidine (AG) and N(ω)-hydroxy-L-arginine (NOHA) in the combinations in in vitro conditions. The results show the decrease of ovalbumin-induced hyperreactivity after inhibition of arginase activity with NOHA. A supplementation of L-arginine caused favourable effect on the airway smooth muscle response. We found the airway reactivity decrease on the whole if we used the combination of NOS and arginase inhibitors. The inhibition of both types of enzymes caused more expressive effect in tracheal smooth muscles. We recorded the difference in the response to histamine or acetylcholine. The simultaneous inhibition of iNOS (with AG) and arginase (with NOHA) evoked the most expressive effect. Results show the importance of competition of both types enzymes - NOS and arginase for the balance of theirs activities in the control of airways bronchomotoric tone in the conditions of the airways hyperreactivity.

Topics: Acetylcholine; Allergens; Animals; Arginase; Arginine; Bronchial Hyperreactivity; Enzyme Inhibitors; Guanidines; Guinea Pigs; Histamine; Lung; Male; Muscle, Smooth; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Ovalbumin; Substrate Specificity; Trachea

Viscum coloratum Nakai is used in traditional Chinese medicine to treat various diseases, including hemorrhage, hypertension, and inflammatory diseases. A previous study demonstrated a partially purified extract (PPE-SVC) and viscolin from Viscum coloratum Nakai inhibited phosphodiesterase activity. In this study, we evaluated the anti-asthmatic effects of PPE-SVC and viscolin, from Viscum coloratum Nakai, in OVA-sensitized mice.. Female BALB/c mice were sensitized and challenged with ovalbumin (OVA). The mice were randomized into groups and treated with PPE-SVC, viscolin, or rolipram by intraperitoneal injection on 1h before each inhalation of OVA and airway hyperresponsiveness (AHR).. PPE-SVC and viscolin suppressed AHR and reduced eosinophil infiltration of the lungs in OVA-sensitized mice. Moreover, PPE-SVC and viscolin inhibited chemokines, including CCL11 and CCL24, and Th2-associated cytokines in bronchoalveolar lavage fluid. However, PPE-SVC and viscolin could not decrease IL-4, IL-5, and IL-13 levels in cultures of OVA-activated spleen cells.. PPE-SVC and viscolin attenuate airway inflammation and eosinophil infiltration in OVA-sensitized mice.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Biphenyl Compounds; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Cytokines; Eosinophils; Female; Inflammation; Lung; Medicine, Chinese Traditional; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plant Extracts; Propane; Random Allocation; Rolipram; Spleen; Viscum

There have been few reports on the role of Fc receptors (FcRs) and immunoglobulin G (IgG) in asthma. The purpose of this study is to clarify the role of inhibitory FcRs and antigen presenting cells (APCs) in pathogenesis of asthma and to evaluate antigen-transporting and presenting capacity by APCs in the tracheobronchial mucosa.. In FcγRIIB deficient (KO) and C57BL/6 (WT) mice, the effects of intratracheal instillation of antigen-specific IgG were analysed using the model with sensitization and airborne challenge with ovalbumin (OVA). Thoracic lymph nodes instilled with fluorescein-conjugated OVA were analysed by fluorescence microscopy. Moreover, we analysed the CD11c+ MHC class II+ cells which intaken fluorescein-conjugated OVA in thoracic lymph nodes by flow cytometry. Also, lung-derived CD11c+ APCs were analysed by flow cytometry. Effects of anti-OVA IgG1 on bone marrow dendritic cells (BMDCs) in vitro were also analysed. Moreover, in FcγRIIB KO mice intravenously transplanted dendritic cells (DCs) differentiated from BMDCs of WT mice, the effects of intratracheal instillation of anti-OVA IgG were evaluated by bronchoalveolar lavage (BAL).. In WT mice, total cells and eosinophils in BAL fluid reduced after instillation with anti-OVA IgG1. Anti-OVA IgG1 suppressed airway inflammation in hyperresponsiveness and histology. In addition, the number of the fluorescein-conjugated OVA in CD11c+ MHC class II+ cells of thoracic lymph nodes with anti-OVA IgG1 instillation decreased compared with PBS. Also, MHC class II expression on lung-derived CD11c+ APCs with anti-OVA IgG1 instillation reduced. Moreover, in vitro, we showed that BMDCs with anti-OVA IgG1 significantly decreased the T cell proliferation. Finally, we demonstrated that the lacking effects of anti-OVA IgG1 on airway inflammation on FcγRIIB KO mice were restored with WT-derived BMDCs transplanted intravenously.. Antigen-specific IgG ameliorates allergic airway inflammation via FcγRIIB on DCs.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD11c Antigen; Cells, Cultured; Dendritic Cells; Disease Models, Animal; Female; Flow Cytometry; Immunoglobulin G; Lymph Nodes; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Microscopy, Fluorescence; Ovalbumin; Receptors, IgG; Th2 Cells

Previous studies have demonstrated that chronic mechanical strain produced by continuous positive airway pressure (CPAP) reduces in vivo airway reactivity in rabbits and ferrets. For CPAP to potentially have a therapeutic benefit for asthmatic subjects, the reduction in airway responsiveness would need to persist for 12-24 h after its discontinuation, require application for only part of the day, and be effective in the presence of atopic airway inflammation. In the present study, airway responsiveness to acetylcholine or methacholine was measured during mechanical ventilation following three different protocols in which active, nonanesthetized, tracheotomized rabbits were treated with High vs. Low CPAP (6 vs. 0 cmH(2)O). 1) High CPAP was applied continuously for 4 days followed by 1 day of Low CPAP; 2) High CPAP was applied at night and Low CPAP during the daytime for 4 days, and 3) High CPAP was applied for 4 days in animals following ovalbumin (Ova) sensitization and challenge. For all three protocols, treatment with High CPAP resulted in significantly reduced airway responsiveness compared with treatment with Low CPAP. Cumulatively, our in vivo results in rabbits suggest that high CPAP, even when applied only at night, produces a persistent reduction of airway responsiveness. In addition, CPAP reduces airway responsiveness even in the presence of atopic airway inflammation.

Topics: Acetylcholine; Airway Resistance; Allergens; Animals; Asthma; Biomechanical Phenomena; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Continuous Positive Airway Pressure; Hypersensitivity, Immediate; Immunohistochemistry; Male; Ovalbumin; Rabbits

Phosphodiesterase 3 (PDE3) inhibitors and sevoflurane are both known to have bronchodilator properties and the combination of these two agents may be synergistic. We tested this hypothesis in a model of airway hyperresponsiveness using ovalbumin-sensitised guinea pigs.. Animals were randomised into five groups: control, sevoflurane, sevoflurane/theophylline, sevoflurane/milrinone and sevoflurane/olprinone. Total lung resistance (RL) and dynamic lung compliance (CL) were recorded and the dose-response curves for acetylcholine (Ach) of RL and CL were used to evaluate the bronchodilator effect.. The dose-response curve for Ach of RL was elevated that for and CL was depressed significantly in the ovalbumin-sensitised animals compared to normal control guinea pigs. Among the five sensitised groups, RL was higher in the order of control > sevoflurane > sevoflurane/theophylline > sevoflurane/milrinone > sevoflurane/olprinone with increasing Ach concentration. Sevoflurane/olprinone treatment attenuated the bronchoconstriction induced by the highest dose of Ach with RL being significantly lower (0.318 ± 0.056 cmH2O ml(-1) s(-1)) than those observed in the control group (0.437 ± 0.061 cmH2O ml(-1) s(-1)), sevoflurane group (0.378 ± 0.052 cmH2O ml(-1) s(-1)) and in the sevoflurane/theophylline group (0.374 ± 0.073 cmH2O ml(-1) s(-1)).. Combined use of PDE inhibitors with a volatile anaesthetic had a synergic bronchodilator effect in ovalbumin-sensitised guinea pigs. A greater bronchodilator effect can be obtained by using the selective PDE3 inhibitor olprinone with the volatile anaesthetic sevoflurane.

Topics: Acetylcholine; Airway Resistance; Anesthetics, Inhalation; Animals; Bronchial Hyperreactivity; Bronchoconstriction; Bronchodilator Agents; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Synergism; Guinea Pigs; Imidazoles; Lung Compliance; Male; Methyl Ethers; Milrinone; Ovalbumin; Phosphodiesterase 3 Inhibitors; Pyridones; Sevoflurane; Theophylline; Time Factors

Many studies have found that smoking reduces lung function, but the relationship between cigarette smoke and allergic asthma has not been clearly elucidated, particularly the role of mast cells. This study aimed to investigate the effects of smoke exposure on allergic asthma and its association with mast cells.. BALB/c mice were sensitized and challenged by OVA to induce asthma, and bone marrow-derived mast cells (BMMCs) were stimulated with antigen/antibody reaction. Mice or BMMCs were exposed to cigarette smoke or CSE solution for 1 mo or 6 h, respectively. The recruitment of inflammatory cells into BAL fluid or lung tissues was determined by Diff-Quik or H&E staining, collagen deposition by Sircol assay, penh values by a whole-body plethysmography, co-localization of tryptase and Smad3 by immunohistochemistry, IgE and TGF-β level by ELISA, expressions of Smads proteins, activities of signaling molecules, or TGF-β mRNA by immunoblotting and RT-PCR.. Cigarette smoke enhanced OVA-specific IgE levels, penh values, recruitment of inflammatory cells including mast cells, expressions of smad family, TGF-β mRNA and proteins, and cytokines, phosphorylations of Smad2 and 3, and MAP kinases, co-localization of tryptase and Smad3, and collagen deposition more than those of BAL cells and lung tissues of OVA-induced allergic mice. CSE solution pretreatment enhanced expressions of TGF-β, Smad3, activities of MAP kinases, NF-κB/AP-1 or PAI-1 more than those of activated-BMMCs.. The data suggest that smoke exposure enhances antigen-induced mast cell activation via TGF-β/Smad signaling pathways in mouse allergic asthma, and that it exacerbates airway inflammation and remodeling.

Topics: Animals; Antigen-Antibody Reactions; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cells, Cultured; Collagen; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Immunoglobulin E; Immunohistochemistry; Lung; Mast Cells; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; NF-kappa B; Ovalbumin; Phosphorylation; Plasminogen Activator Inhibitor 1; Plethysmography, Whole Body; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; RNA, Messenger; Signal Transduction; Smad Proteins; Smad2 Protein; Smad3 Protein; Smoking; Time Factors; Transcription Factor AP-1; Transforming Growth Factor beta; Tryptases

Chrysin, a flavonoid obtained from various natural sources, has been reported to act as an anti-inflammatory and antioxidant agent. However, its anti-allergic action is not fully understood.. In this study, we investigated the in vivo anti-asthmatic activity of chrysin.. The effects of chrysin were evaluated using ovalbumin (OVA) (two subcutaneous 1 mL injections of 20 μg) to induce bronchoalveolar hyperresponsiveness in rats. Chrysin, when administered at 3, 10, and 30 mg/kg, p.o., respectively, before OVA challenge, reduced inflammatory cell (total and differential cell count) infiltration into the lungs measured from bronchoalveolar lavage fluid as supported by lung histology.. The total lung injury score was reduced in a dose-dependent manner, evaluated in six different categories (infiltration of leucocytes, type of inflammatory exudates, status of bronchi, perivascular status of lung blood vessels, integrity of alveoli and activation of alveolar macrophages). Various cellular injury parameters such as alkaline phosphatase, lactate dehydrogenase, and total protein were estimated and found to be reduced by chrysin pretreatment. Further, chrysin was found to reduce nitrite concentration (NO) and lipid peroxidation, suggesting its antioxidant activity.. Chrysin showed anti-asthmatic potential, probably due to the alteration of Th1/Th2 polarization via the suppression of inducible nitric oxide synthase, nuclear factor-κB, and activation protein.

Topics: Animals; Anti-Asthmatic Agents; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cell Polarity; Dose-Response Relationship, Drug; Flavonoids; Lung; Organ Size; Ovalbumin; Oxidative Stress; Rats; Rats, Wistar

The role of traffic-related air pollution in the development of allergic diseases is still unclear. We therefore investigated if NO₂, an important constituent of traffic-related air pollution, promotes allergic sensitization to the allergen ovalbumin (OVA). We also examined if NO₂ influenced the allergy adjuvant activity of diesel exhaust particles (DEP). For this purpose, mice were exposed intranasally to OVA with or without DEP present, immediately followed by exposure to NO₂ (5 or 25 parts per million [ppm]) or room air for 4 h in whole body exposure chambers. Eighteen hours after the last of three exposures, the lungs of half of the animals were lavaged with saline and markers of lung damage and lung inflammation in the bronchoalveolar lavage fluid (BALF) were measured. Three weeks later, after intranasal booster immunizations with OVA, the levels of OVA-specific IgE and IgG2a antibodies in serum were determined. Both NO₂ (25 ppm) and DEP gave lung damage, measured as increased total protein concentration in BALF, whereas only NO₂ seemed to stimulate release of the proinflammatory cytokine tumor necrosis factor alpha (TNF-α). In contrast, only DEP significantly increased the number of neutrophils. Furthermore, DEP in combination with OVA stimulated the production of serum allergen-specific IgE antibodies. NO₂, however, neither increased the production of allergen-specific IgE antibodies, nor influenced the IgE adjuvant activity of DEP. Thus, based on our findings, NO₂ seems to be of less importance than combustion particles in the development of allergic diseases after exposure to traffic-related air pollution.

Topics: Adjuvants, Immunologic; Administration, Intranasal; Air Pollutants; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Drug Interactions; Female; Inflammation; Inhalation Exposure; Mice; Mice, Inbred BALB C; Nitrogen Dioxide; Ovalbumin; Vehicle Emissions

Neurotrophins such as nerve growth factor and brain-derived neurotrophic factor have been described to be involved in the pathogenesis of asthma. Neurturin (NTN), another neurotrophin from the glial cell line-derived neurotrophic factor family, was shown to be produced by human immune cells: monocytes, B cells, and T cells. Furthermore, it was previously described that the secretion of inflammatory cytokines was dramatically stimulated in NTN knockout (NTN(-/-)) mice. NTN is structurally similar to TGF-β, a protective cytokine in airway inflammation. This study investigates the implication of NTN in a model of allergic airway inflammation using NTN(-/-) mice. The bronchial inflammatory response of OVA-sensitized NTN(-/-) mice was compared with wild-type mice. Airway inflammation, Th2 cytokines, and airway hyperresponsiveness (AHR) were examined. NTN(-/-) mice showed an increase of OVA-specific serum IgE and a pronounced worsening of inflammatory features. Eosinophil number and IL-4 and IL-5 concentration in the bronchoalveolar lavage fluid and lung tissue were increased. In parallel, Th2 cytokine secretion of lung draining lymph node cells was also augmented when stimulated by OVA in vitro. Furthermore, AHR was markedly enhanced in NTN(-/-) mice after sensitization and challenge when compared with wild-type mice. Administration of NTN before challenge with OVA partially rescues the phenotype of NTN(-/-) mice. These findings provide evidence for a dampening role of NTN on allergic inflammation and AHR in a murine model of asthma.

Topics: Animals; Antibodies; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Eosinophils; Flow Cytometry; Glial Cell Line-Derived Neurotrophic Factor Receptors; Lung; Lymph Nodes; Mice; Mice, Inbred C57BL; Mice, Knockout; Neurturin; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; Th2 Cells

Proliferation of activated CD4+ T lymphocytes is inhibited by indoleamine 2,3-dioxygenase (IDO).. We undertook the present study to test the hypothesis that IDO-expressing immature DCs (imDCs) can restore immune tolerance in mice suffering from allergic airway inflammation.. imDCs were generated from murine bone marrow cells using granulocyte-macrophage colony-stimulating factor.The imDCs were subsequently transfected with an IDO expression vector (pEGFP-N1-IDO). Surface marker expression, including CD11c, MHCII, CD80, and CD86, was analyzed using flow cytometry. IDO-expressing imDCs were injected into the trachea of ovalbumin (OVA)-sensitized mice, and lung histopathology and cytokine expression in bronchoalveolar lavage fluid were assessed. The splenic CD4+ T cells of OVA-sensitized mice were isolated and co-cultured with pEGFP-N1-IDO-expressing imDCs, and apoptosis of CD4+ T cells was detected using the terminal deoxynucleotidyl transferase dUTP nick end labeling assay.. Expression of IDO in imDCs did not alter cell surface molecule expression. We observed marked lung inflammation, elevated total cell and eosinophil count, and altered cytokine levels in OVA-sensitized mice. These parameters improved upon inoculation with IDO-expressing imDCs. Co-culture with IDO-expressing imDCs also induced apoptosis, inhibited IL-4 and IL-5 expression, and upregulated IFN-gamma expression in CD4+ T cells.. IDO-expressing imDCs induced T(H)2 cell apoptosis and reduced T(H)2 cell activation and allergic airway inflammation in OVA-sensitized mice. Thus, upregulation of IDO expression may provide a novel immunointervention strategy for asthma treatment.

Topics: Animals; Apoptosis; Asthma; Bone Marrow Cells; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cytokines; Dendritic Cells; Eosinophils; Female; Flow Cytometry; Genes, MHC Class II; Granulocyte-Macrophage Colony-Stimulating Factor; Immune Tolerance; Indoleamine-Pyrrole 2,3,-Dioxygenase; Interferon-gamma; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells; Up-Regulation

Some epidemiologic studies have indicated that attendance to chlorinated swimming pools is associated with airway hyperreactivity (AHR), allergies and asthma.. To investigate the effects of sodium hypochlorite (NaClO), the main pool disinfectant, on allergic sensitization and airway inflammation in mice.. In a first series of experiments, mice were sensitized to ovalbumin (OVA), followed by OVA aerosols with or without prior nasal instillation of NaClO (3ppm active chlorine). In a second series, naïve mice received 1-7 nasal instillations of OVA, 10min after instillations of NaClO or water. After 1, 3, 5 and 7 exposures airway reactivity to methacholine, cellular inflammation in bronchoalveolar lavage (BAL), serum OVA-specific IgEs and lung Th2 cytokines were measured.. In the first mouse model, airway allergy parameters were not significantly altered upon NaClO administration. However in the second model, NaClO exposure prior to OVA did induce AHR, already after 1 combined application. Combined NaClO+OVA exposure did not lead to an influx of inflammatory cells in BAL fluid or production of anti-OVA IgEs. No AHR developed when OVA was heat-denatured, pre-chlorinated, or replaced by bovine serum albumin or lipopolysaccharide.. Nasal instillation of NaClO prior to OVA induces AHR without allergic sensitization. This response is OVA-specific.

Topics: Animals; Bronchial Hyperreactivity; Hypersensitivity; Hypochlorous Acid; Male; Mice; Mice, Inbred BALB C; Ovalbumin

The prostaglandin D(2) (PGD(2))/CRTH2 pathway is important for eosinophil trafficking in vitro; however, genetic deficiency of CRTH2 does not suppress in vivo eosinophilic airway inflammation in acute models of asthma, and the role of CRTH2 in the pathogenesis of asthma is still ambiguous. Therefore, in the present study we explored whether the PGD(2)/CRTH2 pathway could affect the phenotypes of chronic asthma. Either CRTH2-deficient (CRTH2-/-) or wild-type mice were sensitized and exposed to ovalbumin (OVA) for 3 days (acute model) or 6 weeks (chronic model). While the magnitude of the acute eosinophilic inflammation was equivalent between CRTH2-/- and wild-type mice, the number of inflammatory cells and eosinophils in bronchoalveolar lavage fluid after chronic OVA exposure was significantly reduced in CRTH2-/- mice (18.0 ± 2.6 × 10(4) cells and 2.0 ± 0.5 × 10(4) cells) compared to wild-type mice (27.9 ± 2.5 × 10(4) cells and 6.8 ± 1.1 × 10(4) cells, p < 0.001). On the contrary, no difference was observed between CRTH2-/- and wild-type mice in terms of airway hyperresponsiveness or remodeling (goblet cell hyperplasia) in the chronic model of asthma. In conclusion, CRTH2 that mediates PGD(2) activity is essential for sustained eosinophilic inflammation in the airways, and its antagonists could exert an anti-inflammatory effect in chronic asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Goblet Cells; Interleukin-13; Interleukin-5; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Knockout; Mucins; Ovalbumin; Pulmonary Eosinophilia; Receptors, Immunologic; Receptors, Prostaglandin; Vaccination

Little is known about the role of the cysteinyl leukotriene (cysLT) 2 receptor in the pathophysiology of asthma. The aim of this study is to investigate the effects of a cysLT1 receptor antagonist (montelukast) and a dual cysLT1/2 receptor antagonist (BAY-u9773) on airway hypersensitivity and airway inflammation induced by antigen challenge in ovalbumin (OVA)-sensitized guinea pigs.. Male Hartley guinea pigs sensitized with OVA were intraperitoneally administered 0.1, 1, or 10 mg/kg of montelukast or 0.1 mg/kg of BAY-u9773 and then challenged with inhaled OVA. Airway reactivity to acetylcholine, inflammatory cells in bronchoalveolar lavage (BAL) fluid, and eosinophil infiltration in airway walls after OVA challenge were evaluated.. Pretreatment with 1 or 10 mg/kg, but not 0.1 mg/kg, of montelukast significantly suppressed airway hypersensitivity and eosinophil infiltration into the BAL fluid. Moreover, 0.1 mg/kg of BAY-u9773 significantly suppressed the development of these markers. The suppressive effects of BAY-u9773, although not significantly different, trended toward being greater than those of montelukast. Although all of the doses of montelukast tested and 0.1 mg/kg of BAY-u9773 significantly suppressed eosinophil infiltration in airway walls, the suppressive effect of BAY-u9773 was significantly greater than that of 0.1 mg/kg of montelukast.. Signaling may contribute to the pathophysiology of asthma via the cysLT1/2 receptor.

Topics: Acetates; Acetylcholine; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cyclopropanes; Eosinophils; Guinea Pigs; Inflammation; Leukotriene Antagonists; Lung; Male; Ovalbumin; Quinolines; Receptors, Leukotriene; SRS-A; Sulfides

We have previously shown that the allergic sensitization to ovalbumin does not represent a superantigen-like immune response. In gene-targeted mice (ΔD-iD) with a single modified Diversity gene segment (D(H)) of the immunoglobulin heavy chain, enriched for charged amino acids, the asthma phenotype in a murine model was markedly alleviated compared to wild-type animals.. We now sought to determine whether the confinement to a single D(H) gene segment alone leads to a reduced allergic phenotype.. We examined another gene-targeted mouse strain (ΔD-DFL) with a single D(H) gene segment which encodes for neutral amino acids, thus reflecting the preferential repertoire in wild-type mice. Mice were sensitized intraperitoneally to ovalbumin.. Despite the constraint to a single D(H) gene segment, ΔD-DFL mice mounted high total and allergen-specific IgG(1) and IgE serum levels after sensitization to ovalbumin. The affinity constants of allergen-specific IgG(1) antibodies did not differ between ΔD-DFL and wild type. Following challenge with aerosolized allergen, a marked local T(H)2 cytokine response and an eosinophilic airway inflammation developed. Quantitative histology revealed increased mucus production and intense goblet cell metaplasia which were identical to those in wild type. Moreover, ΔD-DFL mice developed an airway hyperreactivity to methacholine and to the specific allergen, which both did not differ from those in wild-type animals.. A single D(H) gene segment is sufficient for the establishment of the asthma phenotype in a murine model of allergic airway inflammation. Thus, the allergic phenotype depends on the amino acid composition and not on the diversity of the classical antigen-binding site.

Topics: Allergens; Animals; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Inflammation; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin

Transient receptor potential vanilloid-1 (TRPV1) may modulate allergic airway inflammation because it is expressed not only on the nerve endings but also on several cells of the immune system. We wanted to know the characteristics of airway and systemic responses against sensitization and challenge with allergens in TRPV1 receptor gene knockout mice (TRPV1(-/-)).. TRPV1(-/-) and their wild-type counterparts (TRPV1(+/+)) were sensitized with either house dust mite (HDM) or ovalbumin (OVA) via intranasal (i.n.) or intraperitoneal (i.p.) route before the final i.n. challenge with the corresponding allergen. One day after the final challenge, serum IgE levels, cytokine levels in the bronchoalveolar lavage fluid (BALF), and the number of BALF cells were examined after measuring bronchial hyperresponsiveness against methacholine.. Compared to TRPV1(+/+), TRPV1(-/-) showed enhanced Th2-biased response after i.n. HDM or OVA sensitization, including increased levels of serum IgE, interleukin 4 (IL-4) and eosinophils in the BALF. By contrast, when sensitized via i.p. route, the response against OVA or HDM was almost similar between TRPV1(+/+) and TRPV1(-/-).. TRPV1 receptor may downregulate Th2-biased immune response when sensitized via airways, although this was not the case when sensitized systemically.

Topics: Administration, Intranasal; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Immunoglobulin E; Injections, Intraperitoneal; Lung; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Pyroglyphidae; Respiratory Hypersensitivity; Th2 Cells; TRPV Cation Channels

Interleukin (IL)-25 (IL-17E) is a potent inducer of the type-2 immune effector response. Previously we have demonstrated that a neutralizing anti-IL-25 antibody, given during the establishment of ovalbumin-specific lung allergy, abrogates airways hyperreactivity.. Blocking IL-25 results in the suppression of IL-13, a cytokine known to exacerbate pulmonary inflammation, and an unexpected reciprocal increase in IL-17A. The role of IL-17A in asthma is complex with reports of both pro-inflammatory and anti-inflammatory functions. Our aim was to determine the influence of IL-17A in regulating IL-25-dependent lung allergy.. Neutralizing antibodies to IL-25 and/or IL-17A were administered during an experimental model of allergic asthma. Bronchoalveolar cell infiltrates and lung cytokine production were determined to assess lung inflammation. Invasive plethysmography was undertaken to measure lung function.. Neutralization of IL-25 correlated with a decrease in IL-13 levels and an increase in IL-17A production, and an accompanying prevention of airway hyperresponsiveness (AHR). Notably, the blocking of IL-17A reversed the protective effects of treating with anti-IL-25 antibodies, resulting in the re-expression of several facets of the lung inflammatory response, including IL-13 and eotaxin production, eosinophilia and AHR. Using mice over-expressing IL-13 we demonstrate that treatment of these mice with anti-IL-25 fails to suppress IL-13 levels and in turn IL-17A levels remain suppressed.. IL-13 is known to be an important inducer of lung inflammation, causing goblet cell hyperplasia and promoting airways hyperreactivity. Our data now demonstrate that IL-13 also plays an important role in the genesis of lung inflammation downstream of IL-25 by suppressing a protective IL-17A response. These findings also highlight the important reciprocal interplay of the IL-17 family members, IL-25 and IL-17A, in regulating allergic lung responses and suggest that the balance of IL-17A, together with IL-25, will be an important consideration in the treatment of allergic asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Female; Humans; Interleukin-13; Interleukin-17; Lung; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Pneumonia; Respiratory Hypersensitivity; Th2 Cells

The syndrome of allergic asthma features reversible bronchoconstriction, airway inflammation and hyperresponsiveness as well as airway remodelling, including goblet cell hyperplasia. Managing severe asthma is still a clinical challenge. Numerous studies report that furosemide, an inhibitor of Na(+)-K(+)-Cl(-) cotransporter (NKCC) reduces airway hyperresponsiveness (AHR) in asthmatic patients. However, the mechanism by which furosemide exerts anti-asthmatic action remains unclear.. This study sought to investigate the cellular profile of NKCC1 expression in the lung and examine the effects of furosemide on several outcome measurements in a mouse model of allergic asthma.. Mice were sensitized and challenged with ovalbumin (OVA). Before challenge, the OVA-sensitized mice were treated with furosemide (4.0 mg/kg/day, via daily intraperitoneal injection for 5 days). Outcome measurements in naïve, OVA-exposure, furosemide-treated naïve and furosemide-treated OVA-exposed mice included the slope of the relationship between inhaled methacholine (MCh) concentration and respiratory system resistance (Slope·R(RS)), bronchoalveolar lavage (BAL) cell counts and immunohistochemical and immunoblotting assays of lung tissues.. NKCC1 immunoreactivity was observed in airway epithelial cells (AECs) and alveolar type II (ATII) cells of the control mice. OVA exposure enhanced the expression of NKCC1 in AECs and ATII cells, and increased the infiltration of NKCC1-expressing T lymphocytes in the lung. NKCC1 immunoreactivity was not detected in the airway smooth muscle (ASM) cells. Furosemide treatment reduced the Slope·R(RS) in both naïve and OVA-exposed mice by about 50%. Furosemide treatment also increased T lymphocyte infiltration to the lung in OVA-exposed mice by approximately 53%, but had no effect on pulmonary goblet cell hyperplasia.. Furosemide decreases basal airway responsiveness, thereby reducing the extent of allergen-induced AHR. However, the same treatment also increases T lymphocytes infiltration in the course of allergic asthma. Further studies are necessary to address the usefulness of furosemide in the clinical treatment of asthma.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Furosemide; Goblet Cells; Hypersensitivity; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Sodium-Potassium-Chloride Symporters; Solute Carrier Family 12, Member 2; T-Lymphocytes; Treatment Outcome

The features of asthma are airway inflammation, reversible airflow obstruction, and an increased sensitivity to bronchoconstricting agents, termed airway hyperresponsiveness (AHR), excess production of Th2 cytokines, and eosinophil accumulation in the lungs. To investigate the antiasthmatic potential of hesperidin as well as the underlying mechanism involved, we studied the inhibitory effect and anti-inflammatory effect of hesperidin (HPN) on the production of Th2 cytokines, eotaxin, IL-17, -OVA-specific IgE in vivo asthma model mice.. In this paper, BALB/c mice were systemically sensitized to ovalbumin (OVA) followed intratracheally, intraperitoneally, and by aerosol allergen challenges. We investigated the effect of HPN on airway hyperresponsiveness, pulmonary eosinophilic infiltration, various immune cell phenotypes, Th2 cytokine production and OVA-specific IgE production in a mouse model of asthma.. In BALB/c mice, we found that HPN-treated groups had suppressed eosinophil infiltration, allergic airway inflammation, and AHR, and these occurred by suppressing the production of IL-5, IL-17, and OVA-specific IgE.. Our data suggest that the therapeutic mechanism by which HPN effectively treats asthma is based on reductions of Th2 cytokines (IL-5), eotaxin, OVA-specific IgE production, and eosinophil infiltration via inhibition of GATA-3 transcription factor.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchodilator Agents; Cytokines; Disease Models, Animal; Eosinophils; Ethanolamines; Female; Formoterol Fumarate; Hesperidin; Humans; Interleukins; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells

It is known that the late asthmatic response (LAR), a characteristic feature of asthma, is closely associated with CD4+ Th2 cell-mediated allergic inflammation. Airway remodeling is also a pathogenesis of asthma, but literature reporting roles of CD4+ cells in the remodeling is controversial. There has been no study that simultaneously assessed the roles of CD4+ cells in both LAR and airway remodeling. Sensitized mice were intratracheally challenged with ovalbumin 4 times. Treatment with an anti-CD4 monoclonal antibody (mAb) before the 1st challenge almost completely abolished increase in CD4+ cells in the tissues after the 4th challenge. The late phase increase in airway resistance after the 4th challenge was also completely inhibited by anti-CD4 mAb. Parameters of airway remodeling, subepithelial fibrosis and epithelial thickening were attenuated by treatment, whereas the inhibition was only 30% - 40%. Bronchial smooth muscle thickening was not affected. Because interleukin (IL)-5 production as well as eosinophilia was effectively suppressed by anti-CD4 mAb, the effect of anti-IL-5 mAb was also examined, resulting in no inhibition of airway remodeling. Collectively, although the LAR was completely dependent on CD4+ cell activation, airway remodeling was only partially dependent on the cell.

Topics: Airway Remodeling; Airway Resistance; Animals; Antibodies, Monoclonal; Asthma; Bronchi; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Dexamethasone; Eosinophilia; Epithelial Cells; Fibrosis; Inflammation; Interleukin-5; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Th2 Cells

S-nitrosoglutathione (GSNO) is an endogenous bronchodilator present in micromolar concentrations in airway lining fluid. Airway GSNO levels decrease in severe respiratory failure and asthma, which is attributable to increased metabolism by GSNO reductase (GSNOR). Indeed, we have found that GSNOR expression and activity correlate inversely with lung S-nitrosothiol (SNO) content and airway hyperresponsiveness (AHR) to methacholine (MCh) challenge in humans with asthmatic phenotypes (Que LG, Yang Z, Stamler JS, Lugogo NL, Kraft M. Am J Respir Crit Care Med 180: 226-231, 2009). Accordingly, we hypothesized that local aerosol delivery of GSNO could ameliorate AHR and inflammation in the ovalbumin-sensitized and -challenged (OVA) mouse model of allergic asthma. Anesthetized, paralyzed, and tracheotomized 6-wk-old male control and OVA C57BL/6 mice were administered a single 15-s treatment of 0-100 mM GSNO. Five minutes later, airway resistance to MCh was measured and SNOs were quantified in bronchoalveolar lavage (BAL). Duration of protection was evaluated following nose-only exposure to 10 mM GSNO for 10 min followed by measurements of airway resistance, inflammatory cells, and cytokines and chemokines at up to 4 h later. Acute delivery of GSNO aerosol protected OVA mice from MCh-induced AHR, with no benefit seen above 20 mM GSNO. The antibronchoconstrictive effects of GSNO aerosol delivered via nose cone were sustained for at least 4 h. However, administration of GSNO did not alter total BAL cell counts or cell differentials and had modest effects on cytokine and chemokine levels. In conclusion, in the OVA mouse model of allergic asthma, aerosolized GSNO has rapid and sustained antibronchoconstrictive effects but does not substantially alter airway inflammation.

Topics: Administration, Inhalation; Aerosols; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Humans; Inflammation; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mucin 5AC; Ovalbumin; S-Nitrosoglutathione; Tracheotomy

To examine the preventive effect of a hydro-ethanolic extract of Nigella sativa on the tracheal responsiveness and white blood cell count in the lung lavage fluid of sensitized guinea pigs.. Three groups of guinea pigs sensitized to intraperitoneally injected and inhaled ovalbumin were given drinking water alone (group S), drinking water containing a low concentration of N. sativa extract (group S+LNS) or drinking water containing a high concentration of N. sativa extract (group S+HNS). The tracheal responses of control animals (group C) and the three groups of sensitized guinea pigs (n = 7 for all groups) to methacholine were measured by the assessment of the tracheal smooth muscle response to increasing concentrations of methacholine, and the effective concentration causing 50% of the maximum response (EC50) was determined. Tracheal responses to 0.1% ovalbumin and white blood cell counts in the lung lavage fluid were also examined.. The tracheal response of the group S guinea pigs to both methacholine and ovalbumin was significantly higher than the response of the controls (p<0.01 for both cases). The tracheal responses of the S+LNS and S+HNS groups to both methacholine and ovalbumin were significantly decreased compared to those of the S group (p<0.05 to p<0.01). The total white blood cell and eosinophil counts in the lung lavage fluid of group S were significantly higher than those of group C (p<0.01). The white blood cell counts in both treated groups showed significant improvements (p<0.01 for both cases).. These results demonstrate the preventive effect of the N. sativa extract on the tracheal response and lung inflammation in sensitized guinea pigs.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Bronchodilator Agents; Cell Count; Dose-Response Relationship, Drug; Guinea Pigs; Male; Methacholine Chloride; Nigella sativa; Ovalbumin; Plant Extracts; Pneumonia; Trachea

BALB/c mice received saline (SAL groups) or ovalbumin (OVA groups) intraperitoneally (days 1, 3, 5, 7, 9, 11 and 13). After 27 days, a burst of intratracheal OVA or SAL (days 40, 43 and 46) was performed. Animals were then divided into four groups (N=8, each) and intranasally instilled with saline (SAL-SAL and OVA-SAL) or residual oil fly ash (SAL-ROFA and OVA-ROFA). 24h later, total, initial and difference resistances (Rtot, Rinit, Rdiff) and static elastance (Est) were measured. Lung responsiveness to methacholine was assessed as slope and sensitivity of Est, Rtot, Rinit, and Rdiff. Lung morphometry (collapsed and normal areas and bronchoconstriction index) and cellularity (polymorphonuclear, mononuclear and mast cells) were determined. OVA or ROFA similarly impaired lung mechanics and increased the amount of polymorphonuclear cells and collapsed areas. OVA-ROFA showed even higher hyperresponsiveness, bronchoconstriction and mast cell infiltration. Thus, we concluded that ROFA exposure may add an extra burden to hyperresponsive lungs.

Topics: Air Pollutants; Air Pollution; Animals; Bronchial Hyperreactivity; Coal Ash; Hypersensitivity; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Function Tests

The prevalence of allergic diseases has increased dramatically during the last four decades and is paralleled by a striking increase in iron intake by infants in affluent societies. Several studies have suggested a link between increased iron intake and the marked increase in prevalence of allergic diseases. We hypothesized that the increased iron intake by infants offers an explanation for the increased prevalence of allergic disease in industrialized societies during the past four decades. A well-established mouse model of ovalbumin (OVA)-driven allergic asthma was used to test the effects of differences in iron intake and systemic iron levels on the manifestations of allergic asthma. Surprisingly, iron supplementation resulted in a significant decrease in airway eosinophilia, while systemic iron injections lead to a significant suppression of both allergen-induced airway eosinophilia and hyperreactivity compared to placebo. In contrast, mice fed on an iron-deprived diet did not show any difference in developing experimentally induced allergic asthma when compared to those fed on an iron-sufficient control diet. In contrast to our hypothesis, airway manifestations of allergic asthma are suppressed by both increased levels of iron intake and systemic iron administrations in the mouse model.

Topics: Allergens; Animals; Asthma; Biomarkers; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophilia; Humans; Immunoglobulin E; Infant; Injections, Intraperitoneal; Iron; Iron-Dextran Complex; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Phenanthrolines; Plethysmography

Airborne exposure to microbial cell wall lipids such as lipopolysaccharide triggers innate immune responses that regulate susceptibility to allergic airway inflammation. α-Glycosylceramides represent another widespread class of microbial lipids that directly stimulate innate-like, IL-4- and IL-13-producing, CD1d-restricted NKT cells. In this study, we demonstrate that NKT cells constitutively accumulate and reside in the microvasculature of the mouse lung. After a single airborne exposure to lipid antigen, they promptly extravasate to orchestrate the formation of peribronchiolar and interstitial lymphohistiocytic granulomas containing numerous eosinophils. Concomitant airborne exposure to ovalbumin (OVA) induces the priming of OVA-specific Th2 cells and IgE antibodies by the same dendritic cell coexpressing CD1d and MHC class II. Although NKT cell activation remains confined to the lipid-exposed lung and draining lymph nodes, Th2 cells recirculate and seed the lung of a parabiotic partner, conferring susceptibility to OVA challenge months after the initial exposure, in a manner independent of NKT cells and CD1d. Thus, transient recruitment and activation of lung-resident intravascular NKT cells can trigger long-term susceptibility to allergic airway inflammation.

Topics: Administration, Inhalation; Animals; Antigen-Presenting Cells; Antigens; Antigens, CD1d; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Dendritic Cells; Genes, MHC Class II; Lipids; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Natural Killer T-Cells; Ovalbumin; Pneumonia

Chronic innocuous aeroallergen exposure attenuates CD4(+) T cell-mediated airways hyperresponsiveness in mice; however, the mechanism(s) remain unclear. We examined the role of airway mucosal dendritic cell (AMDC) subsets in this process using a multi-OVA aerosol-induced tolerance model in sensitized BALB/c mice. Aeroallergen capture by both CD11b(lo) and CD11b(hi) AMDC and the delivery of OVA to airway draining lymph nodes by CD8α(-) migratory dendritic cells (DC) were decreased in vivo (but not in vitro) when compared with sensitized but nontolerant mice. This was functionally significant, because in vivo proliferation of OVA-specific CD4(+) T cells was suppressed in airway draining lymph nodes of tolerized mice and could be restored by intranasal transfer of OVA-pulsed and activated exogenous DC, indicating a deficiency in Ag presentation by endogenous DC arriving from the airway mucosa. Bone marrow-derived DC Ag-presenting function was suppressed in multi-OVA tolerized mice, and allergen availability to airway APC populations was limited after multi-OVA exposure, as indicated by reduced OVA and dextran uptake by airway interstitial macrophages, with diffusion rather than localization of OVA across the airway mucosal surface. These data indicate that inhalation tolerance limits aeroallergen capture by AMDC subsets through a mechanism of bone marrow suppression of DC precursor function coupled with reduced Ag availability in vivo at the airway mucosa, resulting in limited Ag delivery to lymph nodes and hypoproliferation of allergen-specific CD4(+) T cells.

Topics: Administration, Inhalation; Allergens; Amino Acid Sequence; Animals; Bone Marrow Cells; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Proliferation; Cells, Cultured; Dendritic Cells; Disease Models, Animal; Epitopes, T-Lymphocyte; Female; Immune Tolerance; Mice; Mice, Inbred BALB C; Mice, Transgenic; Molecular Sequence Data; Ovalbumin; Peptide Fragments; Respiratory Mucosa; Stem Cells

CD8+ T cells participate in airway hyperresponsiveness (AHR) and allergic pulmonary inflammation that are characteristics of asthma. CXCL10 by binding to CXCR3 expressed preferentially on activated CD8+ T cells, attracts T cells homing to the lung. We studied the contribution and limitation of CXCR3 to AHR and airway inflammation induced by ovalbumin (OVA) using CXCR3 knockout (KO) mice.. Mice were sensitized and challenged with OVA. Lung histopathological changes, AHR, cellular composition and levels of inflammatory mediators in bronchoalveolar lavage (BAL) fluid, and lungs at mRNA and protein levels, were compared between CXCR3 KO mice and wild type (WT) mice.. Compared with the WT controls, CXCR3 KO mice showed less OVA-induced infiltration of inflammatory cells around airways and vessels, and less mucus production. CXCR3 KO mice failed to develop significant AHR. They also demonstrated significantly fewer CD8+ T and CD4+ T cells in BAL fluid, lower levels of TNFα and IL-4 in lung tissue measured by real-time RT-PCR and in BAL fluid by ELISA, with significant elevation of IFNγ mRNA and protein expression levels.. We conclude that CXCR3 is crucial for AHR and airway inflammation by promoting recruitment of more CD8+ T cells, as well as CD4+ T cells, and initiating release of proinflammatory mediators following OVA sensitization and challenge. CXCR3 may represent a novel therapeutic target for asthma.

Topics: Animals; Antigens; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Female; Inflammation Mediators; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Receptors, CXCR3

Allergic diseases, including asthma and food allergies, are an increasing health concern. Immunotherapy is an effective therapeutic approach for many allergic diseases but requires long dose escalation periods and has a high risk of adverse reactions, particularly in food allergy. New methods to safely induce Ag-specific tolerance could improve the clinical approach to allergic disease. We hypothesized that Ag-specific tolerance induced by the i.v. injection of Ags attached to the surface of syngeneic splenic leukocytes (Ag-coupled splenocytes [Ag-SPs]) with the chemical cross-linking agent ethylene-carbodiimide, which effectively modulate Th1/Th17 diseases, may also safely and efficiently induce tolerance in Th2-mediated mouse models of allergic asthma and food allergy. Mice were tolerized with Ag-SP before or after initiation of OVA/alum-induced allergic airway inflammation or peanut-induced food allergy. The effects on disease pathology and Th2-directed cytokine and Ab responses were studied. Ag-SP tolerance prevented disease development in both models and safely tolerized T cell responses in an Ag-specific manner in presensitized animals. Prophylactically, Ag-SP efficiently decreased local and systemic Th2 responses, eosinophilia, and Ag-specific IgE. Interestingly, Ag-SP induced Th2 tolerance was found to be partially dependent on the function of CD25(+) regulatory T cells in the food allergy model, but was regulatory T cell independent in the model of allergic airway inflammation. We demonstrate that Ag-SP tolerance can be rapidly, safely, and efficiently induced in murine models of allergic disease, highlighting a potential new Ag-specific tolerance immunotherapy for Th2-associated allergic diseases.

Topics: Allergens; Animals; Arachis; Bronchial Hyperreactivity; Desensitization, Immunologic; Disease Models, Animal; Humans; Immune Tolerance; Inflammation; Leukocytes; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Multiple Sclerosis; Ovalbumin; Peanut Hypersensitivity; Spleen; Th2 Cells

To investigate the inflammatory changes and the airway hyper-responsiveness in the asthma mouse model infected by respiratory syncytial virus and elucidate the relationship between the infection and the effect of glucocorticoid.. 60 BALB/c mice were randomly divided into 6 groups. One of these is the control group; the others are the OVA/sham group, the OVA/sham +Dex group, the PBS/RSV group, the OVA/RSV group and the OVA/RSV+Dex group. The airway resistance was measured using a sealed body plethysmograph. Pathological slides were stained with hematoxylin-eosin, and the peribronchial inflammation was observed microscopically. The concentrations of IL-4, IFN-gamma, TGF-beta1 in lung tissues were detected by ELISA.. Compared with the control group, the degree of the airway inflammation and hyperresponsiveness and the concentrations of IL-4/IFN-gamma, TGF-beta1 in all four OVA groups increased significantly. And there was a statistically significant difference between the OVA/sham group and the OVA/sham+Dex group, and between the OVA/RSV group and the OVA/RSV+Dex group respectively. Compared with the OVA/RSV group, there was an obvious aggravation of airway inflammation and hyper-responsiveness in the OVA/RSV+Dex group.. Glucocorticoid significantly reduces airway inflammation and hyper-responsiveness induced by repetitive OVA challenge in the mouse model of asthma. However, the significant decrease in Th1 and increase in Th2 inflammation and aggravation of airway hyper-responsiveness in the mice in OVA/RSV group show that they are not sensitive to glucocorticoid. The effects of infection with RSV on the mouse model of asthma could be the cause of the glucocorticoid resistance during the therapy.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cell Line; Dexamethasone; Disease Models, Animal; Disease Progression; Female; Humans; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Plethysmography; Respiratory Syncytial Virus Infections; Respiratory Syncytial Viruses; Th1-Th2 Balance

Hesperetin was reported to selectively inhibit phosphodiesterase 4 (PDE4). While hesperetin-7,3'-O-dimethylether (HDME) is a synthetic liposoluble hesperetin. Therefore, we were interested in investigating its selectivity on PDE4 and binding ability on high-affinity rolipram-binding sites (HARBs) in vitro, and its effects on ovalbumin-induced airway hyperresponsiveness in vivo, and clarifying its potential for treating asthma and chronic obstructive pulmonary disease (COPD).. PDE1~5 activities were measured using a two-step procedure. The binding of HDME on high-affinity rolipram-binding sites was determined by replacing 2 nM [3H]-rolipram. AHR was assessed using the FlexiVent system and barometric plethysmography. Inflammatory cells were counted using a hemocytometer. Cytokines were determined using mouse T helper (Th)1/Th2 cytokine CBA kits, and total immunoglobulin (Ig)E or IgG2a levels were done using ELISA method. Xylazine (10 mg/kg)/ketamine (70 mg/kg)-induced anesthesia was performed.. HDME revealed selective phosphodiesterase 4 (PDE4) inhibition with a therapeutic (PDE4H/PDE4L) ratio of 35.5 in vitro. In vivo, HDME (3~30 μmol/kg, orally (p.o.)) dose-dependently and significantly attenuated the airway resistance (RL) and increased lung dynamic compliance (Cdyn), and decreased enhanced pause (Penh) values induced by methacholine in sensitized and challenged mice. It also significantly suppressed the increases in the numbers of total inflammatory cells, macrophages, lymphocytes, neutrophils, and eosinophils, and levels of cytokines, including interleukin (IL)-2, IL-4, IL-5, interferon-γ, and tumor necrosis factor-α in bronchoalveolar lavage fluid (BALF) of these mice. In addition, HDME (3~30 μmol/kg, p.o.) dose-dependently and significantly suppressed total and ovalbumin-specific immunoglobulin (Ig)E levels in the BALF and serum, and enhanced IgG2a level in the serum of these mice.. HDME exerted anti-inflammatory effects, including suppression of AHR, and reduced expressions of inflammatory cells and cytokines in this murine model, which appears to be suitable for studying the effects of drugs on atypical asthma and COPD, and for screening those on typical asthma. However, HDME did not influnce xylazine/ketamine-induced anesthesia. Thus HDME may have the potential for use in treating typical and atypical asthma, and COPD.

Topics: Animals; Asthma; Binding Sites; Blood Cell Count; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cyclic Nucleotide Phosphodiesterases, Type 1; Cyclic Nucleotide Phosphodiesterases, Type 3; Cyclic Nucleotide Phosphodiesterases, Type 4; Cytokines; Disease Models, Animal; Female; Guinea Pigs; Hesperidin; Immunoglobulins; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Phosphodiesterase 4 Inhibitors; Pulmonary Disease, Chronic Obstructive; Rolipram

Glucocorticoids are the mainstream drugs used in the treatment and control of inflammatory diseases such as asthma. Annexin-1 (ANXA1) is an anti-inflammatory protein which has been described as an endogenous protein responsible for some anti-inflammatory glucocorticoid effects. Previous studies have identified its importance in other immune diseases such as rheumatoid arthritis and cystic fibrosis. ANXA1-deficient ((-/-)) mice are Th2 biased, and ANXA1 N-terminus peptide exhibits anti-inflammatory activity in a rat model of pulmonary inflammation.. ANXA1 protein is found in bronchoalveolar lavage fluid from asthmatics. However, the function of ANXA1 in the pathological development of allergy or asthma is unclear. Thus, in this study we intended to examine the effect of ANXA1 deficiency on allergen-specific antibody responses and airway responses to methacholine (Mch).. ANXA1(-/-) mice were sensitized with ovalbumin (OVA) and challenged with aerosolized OVA. Airway resistance, lung compliance and enhanced pause (PenH) were measured in naïve, sensitized and saline or allergen-challenged wild-type (WT) and ANXA1(-/-) mice. Total and allergen-specific antibodies were measured in the serum.. We show that allergen-specific and total IgE, IgG2a and IgG2b levels were significantly higher in ANXA1(-/-) mice. Furthermore, naïve ANXA1(-/-) mice displayed higher airway hypersensitivity to inhaled Mch, and significant differences were also observed in allergen-sensitized and allergen-challenged ANXA1(-/-) mice compared with WT mice.. In conclusion, ANXA1(-/-) mice possess multiple features characteristic to allergic asthma, such as airway hyperresponsiveness and enhanced antibody responses, suggesting that ANXA1 plays a critical regulatory role in the development of asthma.. We postulate that ANXA1 is an important regulatory factor in the development of allergic disease and dysregulation of its expression can lead to pathological changes which may affect disease progression.

Topics: Allergens; Animals; Annexin A1; Annexins; Antibody Specificity; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophils; Female; Immunity, Humoral; Immunoglobulin E; Interleukin-4; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin

Danggui Buxue Tang (DBT), an herbal formula containing Angelica sinensis (AS) and Astragalus membranaceus (AM) (AS:AM = 1:5, designated as DBT1 here), has been used in Chinese medicine to enhance qi and blood circulation. In addition, DBT has served as a treatment for atopic dermatitis in dogs in Taiwan. It also may improve fibrosis in a rat model of pulmonary fibrosis.. In this study, we evaluated the effect of oral administration of DBT1 in asthma in ovalbumin (OVA)-sensitized mice.. Female BALB/c mice were sensitized and challenged with OVA and fed with DBT1 or modified formulas of DBT1, designated as DBT2 (AS:AM = 1:1) and DBT3 (AS:AM = 5:1), from days 21 to 27.. DBT1 suppressed airway hyperresponsiveness and eosinophil infiltration in bronchoalveolar lavage fluid (BALF) and lung, and Th2-associated cytokines and chemokines were inhibited in BALF. In addition, levels of OVA-immunoglobulin E (IgE) also were suppressed in serum. However, treatment with DBT2 or DBT3 showed no improved effects relative to DBT1 in treating asthmatic symptoms.. These results suggest that orally administered DBT (DBT1) can reduce allergic reactions in OVA-sensitized mice.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Drugs, Chinese Herbal; Female; Immunoglobulin E; Immunohistochemistry; Mice; Mice, Inbred BALB C; Ovalbumin; Random Allocation

The presented studies were aimed on experimental confirmation of Althaea officinalis polysaccharide rhamnogalacturonan antitussive effect and its changes in conditions of allergic inflammation.. We have tested whether rhamnogalacturonan inhibits cough reflex and modulates airways reactivity of guinea pigs in vivo. The cough in guinea pigs was induced by 0.3 M citric acid (CA) aerosol for 3 min interval, in which total number of cough efforts (sudden enhancement of expiratory flow accompanied by cough movement and sound) was counted. Specific airway resistance and its changes induced by citric acid aerosol were considered as an indicator of the in vivo reactivity changes.. 1) Althaea officinalis polysaccharide rhamnogalacturonan dose- dependently inhibits cough reflex in unsensitized guinea pigs. Simultaneously, plant polysaccharide shortened the duration of antitussive effect when it was been tested in inflammatory conditions. 2) Rhamnogalacturonan did not influence airways reactivity in vivo conditions expressed as specific resistance values neither sensitized nor unsensitized groups of animals. 3) The antitussive activity of codeine (dose 10 mg.kg(-1) b.w. orally) tested under the same condition was comparable to higher dose of rhamnogalacturonan in unsensitized animals. 4) The characteristic cellular pattern of allergic airways inflammation was confirmed by histopathological investigations.. Rhamnogalacturonan isolated from Althaea officinalis mucilage possesses very high cough suppressive effect in guinea pigs test system, which is shortened in conditions of experimentally induced airways allergic inflammation (Tab. 1, Fig. 4, Ref. 25). Full Text in free PDF www.bmj.sk.

Topics: Airway Resistance; Althaea; Animals; Antitussive Agents; Bronchial Hyperreactivity; Cough; Dose-Response Relationship, Drug; Guinea Pigs; Inflammation; Lung; Ovalbumin; Plant Extracts; Polysaccharides; Reflex; Trachea

BLT2 is a low-affinity receptor for leukotriene B(4) (LTB(4)), a potent lipid mediator of inflammation generated from arachidonic acid via the 5-lipoxygenase pathway. Unlike BLT1, a high-affinity receptor for LTB(4), no clear physiological function has yet been identified for BLT2, especially with regard to the pathogenesis of asthma. The aim of this study was to investigate whether BLT2 plays a role in the pathogenesis of asthma. A murine model of allergic asthma was used to evaluate the role of BLT2 in ovalbumin-induced airway inflammation and airway hyperresponsiveness. The levels of BLT2 mRNA and its ligand, LTB(4), in the lung airway were highly elevated after ovalbumin challenge, and down-regulation of BLT2 with antisense BLT2 oligonucleotides markedly attenuated airway inflammation and airway hyperresponsiveness. Further analysis, aimed at identifying mediators downstream of BLT2, revealed that BLT2 activation led to elevation of reactive oxygen species and subsequent activation of NF-kappaB, thus inducing the expression of vascular cell adhesion molecule-1, which is known to be involved in eosinophil infiltration into the lung airway. Together, our results suggest that BLT2 plays a pivotal, mediatory role in the pathogenesis of asthma, acting through a "reactive oxygen species-NF-kappaB"-linked inflammatory signaling pathway.

Topics: Animals; Asthma; Biopsy; Bronchi; Bronchial Hyperreactivity; Cell Movement; Gene Expression Regulation; Humans; Lung; Mice; NF-kappa B; Ovalbumin; Pneumonia; Reactive Oxygen Species; Receptors, Leukotriene B4; RNA, Antisense; RNA, Messenger; Signal Transduction; T-Lymphocytes; Tetrazoles; Vascular Cell Adhesion Molecule-1

In previous studies, the relaxant, anticholinergic (functional antagonism) and antihistaminic, effects of Nigella sativa have been demonstrated on guinea pig tracheal chains. In the present study, the prophylactic effect of thymoquinone (one of the constituents of Nigella sativa) on tracheal responsiveness and white blood cell (WBC) count in lung lavage of sensitized guinea pigs was examined. Four groups of sensitized guinea pigs to ovalbumin (OA) were given drinking water alone (group S), drinking water containing low or high concentrations of thymoquinone (S + LTQ and S + HTQ groups) or inhaled fluticasone propionate (FP 250 microg) twice a day (positive control group) (n = 7, for all groups). Tracheal responses of control and four groups of sensitized animals to methacholine at an effective concentration causing 50 % of maximum response (EC(50) M) were measured. Tracheal responses to 0.1 % OA, relative to contraction induced by 10 microM methacholine were also examined. Total WBC and its differential count in lung lavage were also measured. The tracheal responsiveness to methacholine, OA and WBC of S guinea pigs was significantly higher than those of controls (p < 0.001 for all cases). Tracheal responsiveness in S + LTQ, S + HTQ, and FP groups to both methacholine (p < 0.05 to p < 0.001) and OA (p < 0.001 for all cases) was significantly decreased compared to that of the S group. Total WBC was also decreased in all experimental groups compared to that of the S group (p < 0.001 for all groups). There was an increase in eosinophils and a decrease in neutrophils, lymphocytes and monocytes in the S animals compared to the controls (p < 0.001 for all cases). Treatment with both concentrations of thymoquinone and FP variably improved differential WBC count changes compared to the S animals (nonsignificant to p < 0.001). The improvement in tracheal responsiveness, total WBC, eosinophils and lymphocytes changes in the S animals treated with both concentrations of TQ were significantly greater than those of FP (p < 0.05 to p < 0.001). These results showed a preventive effect of thymoquinone, one constituent of N. sativa, on tracheal responsiveness and inflammatory cells of lung lavage of sensitized guinea pigs which was comparable or even greater than that of the inhaled steroid.

Topics: Androstadienes; Animals; Anti-Inflammatory Agents; Benzoquinones; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Bronchoconstrictor Agents; Female; Fluticasone; Guinea Pigs; Leukocyte Count; Leukocytes; Lung; Lymphocytes; Male; Methacholine Chloride; Nigella sativa; Ovalbumin; Phytotherapy; Plant Extracts; Trachea

Evidence demonstrating an important role of the intestinal microbiota in the incidence of allergic disorders has led to the concept of using probiotics as possible antiallergic therapy. This study aimed to select a bacterial strain with the best antiallergic treatment effects from a panel of 6 bacterial strains in a mouse model of ovalbumin(OVA)-allergic asthma.. OVA-sensitized BALB/c mice were orally administered the bacterial strains Bifidobacterium breve M-16V, B. infantis NumRes251, B. animalis NumRes252 and NumRes253, Lactobacillus plantarum NumRes8 and L. rhamnosus NumRes6. After challenge by OVA inhalation in the lungs, the response to methacholine was measured. Pulmonary inflammation was assessed by analyzing bronchoalveolar lavage fluid for the presence of eosinophils, neutrophils, macrophages and lymphocytes and for interleukin 4, interleukin 5, interleukin 10 and interferon-gamma. OVA-specific IgE, IgG1 and IgG2a were measured in serum. Next, the effect on acute allergic skin reaction was measured after treatment with B. breve M-16V and L. plantarum NumRes8.. Of the panel of 6 strains, B. breve M-16V and L. plantarum NumRes8 inhibited (1) the response to methacholine, (2) reduced the number of eosinophils in the bronchoalveolar lavage fluid, (3) reduced both OVA-specific IgE and (4) OVA-specific IgG1, whereas the other strains did not affect all these parameters simultaneously. B. breve M-16V but not L. plantarum NumRes8 reduced interleukin 4, interleukin 5 and interleukin 10. Furthermore, B. breve M-16V but not L. plantarum NumRes8 reduced acute allergic skin reactions to OVA.. B. breve M-16V was identified as the most potent antiallergic strain.

Topics: Administration, Oral; Animals; Asthma; Bifidobacterium; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cell Count; Immunoglobulin E; Immunoglobulin G; Interferon-gamma; Interleukin-10; Interleukin-4; Interleukin-5; Lacticaseibacillus rhamnosus; Lactobacillus plantarum; Lymphocytes; Macrophages, Alveolar; Male; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Probiotics; Pulmonary Eosinophilia; Skin Tests; Specific Pathogen-Free Organisms; Vaccination

We investigated the efficacy of Viola mandshurica W. Becker (VM) ethanolic (EtOH) extract in the treatment of bronchial asthma in an ovalbumin (OVA)-induced asthmatic BALB/c mouse model.. Female BALB/c mice were sensitized with intraperitoneal (i.p.) ovalbumin (OVA) on days 0 and 14, and were next given intranasal OVA on days 28-30. Randomized treatment groups of sensitized mice received VM EtOH extract, dexamethasone, or placebo, orally, from days 28 to 30.. VM EtOH extract significantly inhibited increases in total immunoglobulin E (IgE) and cytokines IL-4 and IL-13 levels in serum and bronchoalveolar lavage fluid (BALF), and also effectively suppressed airway hyperresponsiveness (AHR), eosinophilia, and mucus hypersecretion, in mice with OVA-induced asthma.. The results suggest that VM EtOH extract and allied extracts could be useful herbal medicines for asthma treatment, and that VM may also be a valuable lead material for anti-asthma drug development.

Topics: Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophilia; Ethanol; Female; Hypersensitivity, Delayed; Immunoglobulin E; Inflammation; Interleukins; Korea; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Phytotherapy; Plant Extracts; Plants, Medicinal; Random Allocation; Viola

The migration of dendritic cells (DCs) from the lungs to the regional lymph nodes is necessary for the development of allergic airway disease. Following activation, mast cells release a variety of stored or de novo-produced inflammatory mediators, several of them being capable of activating DCs. In this study, the role of mast cells on DC migration from the lungs to the thoracic lymph nodes was investigated in sensitized mice.. Mast cell-deficient mice (Kit(W-sh/W-sh)) and their wild-type counterparts were sensitized intraperitoneally with ovalbumine (OVA) in saline and challenged by a single intranasal administration of OVA labeled with a fluorescent dye (OVA-Alexa).. Following challenge, the relative and absolute amount of OVA- Alexa-positive DCs was clearly increased in sensitized wild-type mice compared to nonsensitized mice. In contrast, sensitized Kit(W-sh/W-sh) showed no increase in OVA-Alexa-positive DCs compared to nonsensitized mast cell-deficient animals. In sensitized Kit(W-sh/W-sh) mice reconstituted with bone marrow-derived mast cells (BMMCs), the number of OVA- Alexa-positive DCs was comparable to that in sensitized wild-type animals. However, transfer of allergen-exposed BMMCs to sensitized mice prior to airway challenge augmented airway inflammation similarly in wild-type and mast cell-deficient mice. In line with this, sensitization with allergen-pulsed DCs induced allergic airway disease independently of mast cells.. This study shows an interaction between mast cells and DCs following allergen challenge in sensitized hosts. However, the function of mast cells can be bypassed in models utilizing activated allergen-exposed DCs to initiate the development of allergic airway disease.

Topics: Adoptive Transfer; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Separation; Chemotaxis, Leukocyte; Dendritic Cells; Flow Cytometry; Mast Cells; Mice; Ovalbumin

The herbal Petasites hybridus (butterbur) extract (Ze 339, PET) is known to have leukotriene inhibiting properties, and therefore might inhibit allergic diseases.. The effect of PET was investigated in ovalbumin (OVA) immunized BALB/c mice given intranasally together with antigen challenge in the murine model of allergic airway disease (asthma) with the analysis of the inflammatory and immune parameters in the lung.. PET given with the antigen challenge inhibited the allergic response. PET inhibited airway hyperresponsiveness (AHR) and eosinophil recruitment into the bronchoalveolar lavage (BAL) fluid upon allergen challenge, but had no effect in the saline control mice. Eosinophil recruitment was further assessed in the lung by eosinophil peroxidase (EPO) activity at a concentration of 100 microg PET. Microscopic investigations revealed less inflammation, eosinophil recruitment and mucus hyperproduction in the lung with 100 microg PET. Diminution of AHR and inflammation was associated with reduced IL-4, IL-5 and RANTES production in the BAL fluid with 30 microg PET, while OVA specific IgE and eotaxin serum levels remained unchanged.. PET, which has been reported to inhibit leukotriene activity, reduced allergic airway inflammation and AHR by inhibiting the production of the Th2 cytokines IL-4 and IL-5, and RANTES.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL5; Disease Models, Animal; Drug Evaluation, Preclinical; Eosinophil Peroxidase; Eosinophils; Immunoglobulin E; Interleukin-4; Interleukin-5; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Petasites; Phytotherapy; Plant Extracts; Th2 Cells

Airway hyperresponsiveness (AHR) in asthmatics includes a variable component that persists following an allergen challenge. This may be dissociated from inflammatory cell recruitment, implying a role for resident pulmonary cells in regulating the response.. Using improved methods of assessing AHR in a mouse model of allergic airway disease, to investigate the basis of the development of prolonged AHR.. BALB/c mice were systemically sensitized and then challenged with aerosolized ovalbumin (OVA). Airway and tissue responsiveness were measured at baseline and at 1 day, and 1, 2 and 3 weeks after the last OVA challenge. Inflammatory cell numbers in BALF and levels of mRNA for eotaxin-1 and -2, IFN-gamma, IL-5 and -13 in the lung were measured at each time-point. In further experiments, the roles of IFN-gamma and of CCR3(+) and CD4(+) cells in the development of prolonged AHR were assessed by blockade or depletion with monoclonal antibodies. The role of pulmonary macrophages was assessed by selective chemical depletion of these cells.. Airway responsiveness was increased above baseline at 1 day after the last OVA challenge, and this was sustained for 1 week. In contrast, tissue-specific responsiveness was only significantly increased above baseline at 1 day. Development of prolonged AHR was inhibited by neutralization of IFN-gamma or by depletion of pulmonary macrophages, but not by depletion of either CD4(+) T cells or CCR3(+) eosinophils.. An interaction between IFN-gamma and pulmonary macrophages contributed to the prolongation of airway hyperresponsiveness. In contrast, T cells and eosinophils did not contribute to prolongation of AHR. These findings emphasize the importance of the innate host response in the development of manifestations of asthma, as well as its potential relevance as a target for therapeutic intervention.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Disease Models, Animal; Immunization; Interferon-gamma; Macrophages, Alveolar; Mice; Mice, Inbred BALB C; Ovalbumin

Asthma is a chronic disease characterized by airway inflammation caused by the dysregulated production of cytokines secreted by allergen-specific type 2 T helper (Th2) cells. Antrodia camphorata is a commonly used fungus in Asian folk medicine, and A. camphorata polysaccharides are reported to possess anti-cancer activities. In this study, the immunomodulatory effects of purified fractionated polysaccharides (GF2) from A. camphorata on dendritic cells (DCs) and their potential preventive effects against ovalbumin (OVA) -induced asthma were investigated. In the presence of GF2, lipopolysaccharide (LPS) -activated DCs exhibited up-regulated expression of major histocompatibility complex (MHC) class II and co-stimulatory molecules, as well as enhanced interleukin-10 (IL-10) and IL-12 production. GF2 treatment on LPS-activated DCs suppressed naïve CD4(+) T-cell proliferation and Th2 cell polarization with IL-10 production in an allogeneic mixed lymphocyte reaction. In animal experiments, a high dose of GF2 efficiently reduced expression levels of OVA-specific immunoglobulin G1 (IgG1) and IgE. However, lower doses of GF2 significantly enhanced OVA-specific IgG2a production. Our data also showed that administration of GF2 dose-dependently inhibited the development of airway hyperresponsiveness, airway eosinophilia and Th2 responses. OVA-specific CD4(+) T cells from higher doses of GF2-treated mice had significantly lower proliferative capacities compared with control mice. Moreover, treatment with GF2 significantly increased the high levels of IL-10 and low levels of interferon-gamma produced by T cells. Taken together, these data indicate that administration of A. camphorata polysaccharides (GF2) may have therapeutic potential when used as an adjuvant for the immunomodulatory treatment of allergic asthma.

Topics: Allergens; Animals; Antrodia; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cell Differentiation; Cell Proliferation; Cell Survival; Cytokines; Dendritic Cells; Female; Gene Expression; Immunoglobulin E; Immunoglobulin G; Interleukin-10; Interleukin-12; Lipopolysaccharides; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Polysaccharides; Pulmonary Eosinophilia; Spleen; T-Lymphocytes; Th2 Cells; Vaccination

Matrix metalloproteinase-9 (MMP-9) is hypothesized to facilitate leukocyte extravasation and extracellular remodeling in asthmatic airways. Careful descriptive studies have shown that MMP-9 levels are higher in the sputum of asthmatics; however, the consequence of increased MMP-9 activity has not been determined in this disease. We induced asthma in transgenic mice that express human MMP-9 in the murine lung tissue macrophage to determine the direct effect of human MMP-9 expression on airway inflammation. Transgenic (TG) and wild-type (WT) mice were immunized and challenged with ovalbumin. Forty-eight hours after the ovalbumin challenge, airway hyperresponsiveness (AHR) was measured, and inflammatory cell infiltration was evaluated in bronchoalveolar lavage fluid (BALF) and lung tissue. Baseline levels of inflammation were similar in the TG and WT groups of mice, and pulmonary eosinophilia was established in both groups by sensitization and challenge with ovalbumin. There was a significant reduction in AHR in sensitized and challenged trangenics compared with WT controls. Although total BALF cell counts were similar in both groups, the lymphocyte number in the lavage of the TG group was significantly diminished compared with the WT group (0.25 +/- 0.08 vs. 0.89 +/- 0.53; P = 0.0032). In addition, the draining lymphocytes were found to be larger in the TG animals compared with the WT mice. Equal numbers of macrophages, eosinophils, and neutrophils were seen in both groups. IL-13 levels were found to be lower in the sensitized TG compared with the WT mice. These results demonstrate an inverse relationship between human MMP-9 and AHR and suggest that MMP-9 expression alters leukocyte extravasation by reducing lymphocyte accumulation in the walls of asthmatic airways.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD3 Complex; Cell Movement; Cytokines; Disease Models, Animal; Humans; Lymph Nodes; Lymphocytes; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Transgenic; Ovalbumin

We studied repeatedly the development of bronchial hyperreactivity (BHR) and bronchoalveolar lavage fluid (BALF) in rats undergoing different modes of ovalbumin exposures. Treatment was two intraperitoneal injections of ovalbumin in Groups 1-3, followed by one ovalbumin aerosolization in Groups 2 and 3, while rats in Group 4 received repeated ovalbumin aerosols after one single intraperitoneal injection. BHR was assessed longitudinally on day 0 (before treatment) and on day 14 (Groups 1 and 2) or 20 (Groups 3 and 4) and cellular influx was estimated from BALF. No BHR or change in BALF cellular profile was detected in Groups 1-3. However, the infiltration of inflammatory cells, associated with BHR (PC(100) 8.9+/-1.3 microg/kg vs. 4.2+/-1.1 microg/kg), was observed in Group 4. The BHR was always associated with increased number of eosinophils in the BALF. The substantial interindividual variability confirmed the need for a technique that permits follow-up of lung responsiveness and BALF profile. This approach evidenced strong associations between the severity of BHR and the eosinophilia.

Topics: Airway Resistance; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cell Count; Disease Models, Animal; Dose-Response Relationship, Drug; Eosinophils; Methacholine Chloride; Neutrophils; Ovalbumin; Rats; Rats, Wistar; Respiratory Mechanics; Time Factors

Nerve growth factor (NGF) contributes to airway inflammation and bronchoconstriction in allergic asthma. The Src homology 2beta/serine/threonine kinase (SH2-Bbeta/Akt) pathway is one of the avenues through which NGF regulates the biological activity of pheochromocytoma (PC)12 cells. It has also been reported that NGF upregulates the expression of SH2-Bbeta in the lung tissue of asthmatic mice. The present study investigated the effects of NGF and SH2-Bbeta on Akt activation during allergic airway challenge.. BALB/c mice were sensitized and challenged with ovalbumin. The effects of NGF and SH2-Bbeta on Akt in allergic airway challenge were assessed by intravenously administering anti-NGF antibody or a mutant of SH2-Bbeta (R555E) to these mice. Pulmonary histological changes were then assessed and the inflammatory cells in the BAL fluid (BALF) were counted. Additionally, phosphorylated Akt (p-Akt) expression was determined by fluorescence microscopy, western blotting and quantitative RT-PCR. Airway resistance was also measured using closed-type body plethysmography.. We observed p-Akt overexpression in the lungs after allergen challenge by fluorescence microscopy, Western blotting and RT-PCR, as compared with the control. However, after treatment with anti-NGF or R555E, p-Akt levels and allergen-induced airway inflammation were reduced in comparison with those of allergen-challenged mice. Anti-NGF and R555E also decreased airway hyperresponsiveness caused by allergen challenge in response to methacholine (MCH).. These results suggest that SH2-Bbeta regulation of Akt partly participates in the NGF-mediated development of allergic airway challenge.

Topics: Adaptor Proteins, Signal Transducing; Airway Resistance; Animals; Asthma; Bronchial Hyperreactivity; Constriction, Pathologic; Female; Methacholine Chloride; Mice; Mice, Inbred BALB C; Nerve Growth Factor; Ovalbumin; Proto-Oncogene Proteins c-akt

A possible role of 5-hydroxytryptamine (5-HT) in the origin of antigen-induced airway hyperresponsiveness (AI-AHR) has been scarcely investigated.. To explore the participation of different 5-HT receptors in the development of AI-AHR in guinea-pigs.. Lung resistance was measured in anaesthetized guinea-pigs sensitized to ovalbumin (OVA). Dose-response curves to intravenous (i.v.) acetylcholine (ACh) were performed before and 1 h after antigenic challenge and expressed as the 200% provocative dose (PD(200)). Organ bath experiments, confocal microscopy and RT-PCR were additionally used. The 5-HT content in lung homogenates was measured by HPLC.. Antigenic challenge significantly decreased PD(200), indicating the development of AI-AHR. This hyperresponsiveness was abolished by a combination of methiothepin (5-HT(1)/5-HT(2)/5-HT(5)/5-HT(6)/5-HT(7) receptors antagonist) and tropisetron (5-HT(3)/5-HT(4) antagonist). Other 5-HT receptor antagonists showed three different patterns of response. Firstly, WAY100135 (5-HT(1A) antagonist) and ondansetron (5-HT(3) antagonist) did not modify the AI-AHR. Secondly, SB269970 (5-HT(7) antagonist), GR113808 (5-HT(4) antagonist), tropisetron or methiothepin abolished the AI-AHR. Thirdly, ketanserin (5-HT(2A) antagonist) produced airway hyporresponsiveness. Animals with bilateral vagotomy did not develop AI-AHR. Experiments in tracheal rings showed that pre-incubation with LP44 or cisapride (agonists of 5-HT(7) and 5-HT(4) receptors, respectively) induced a significant increase of the cholinergic contractile response to the electrical field stimulation. In sensitized lung parenchyma strips, ketanserin diminished the contractile responses to ACh. Sensitization was associated with a ninefold increase in the 5-HT content of lung homogenates. Confocal microscopy showed that sensitization enhanced the immunolabelling and co-localization of nicotinic receptor and 5-HT in airway epithelium, probably located in pulmonary neuroendocrine cells (PNECs). RT-PCR demonstrated that neither sensitization nor antigen challenge modified the 5-HT(2A) receptor mRNA levels.. Our results suggested that 5-HT was involved in the development of AI-AHR to ACh in guinea-pigs. Specifically, 5-HT(2A), 5-HT(4) and 5-HT(7) receptors seem to be particularly involved in this phenomenon. Participation of 5-HT might probably be favoured by the enhancement of the PNECs 5-HT content observed after sensitization.

Topics: Animals; Bronchial Hyperreactivity; Chromatography, High Pressure Liquid; Disease Models, Animal; Dose-Response Relationship, Drug; Guinea Pigs; Male; Ovalbumin; Receptor, Serotonin, 5-HT2A; Receptors, Serotonin; Receptors, Serotonin, 5-HT4; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sensitivity and Specificity; Serotonin; Serotonin 5-HT2 Receptor Antagonists; Serotonin 5-HT4 Receptor Antagonists

ONO-1301 is a novel drug that acts as a prostacyclin agonist with thromboxane A(2) (TxA(2)) synthase inhibitory activity. We investigated the effect of ONO-1301 on development of airway allergic inflammation.. Mice sensitized and challenged to ovalbumin (OVA) received ONO-1301, OKY-046 (TxA(2) synthase inhibitor), beraprost, a prostacyclin receptor (IP) agonist, ONO-1301 plus CAY10449 (selective IP antagonist) or vehicle during the challenge period. Twenty-four hours after the OVA challenge, airway hyperresponsiveness (AHR) to methacholine was assessed and bronchoalveolar lavage was performed. Lung specimens were excised for goblet cell staining and analysis of lung dendritic cells (DCs). Bone marrow-derived dendritic cells (BMDCs) were generated, in the presence or absence of drugs, for analysis of DC function.. Mice that received ONO-1301 showed significantly lower AHR, airway eosinophilia, T-helper type 2 cytokine levels, mucus production and lung DCs numbers than vehicle-treated mice. These effects of ONO-1301 were mostly reversed by CAY10449. BMDCs treated with ONO-1301 alone showed lower DC functions, such as expression of costimulatory factors or stimulation to spleen T cells.. These data suggest that ONO-1301 may suppress AHR and airway allergic inflammation through modulation of DCs, mainly mediated through the IP receptor. This agent may be effective as an anti-inflammatory drug in the treatment of asthma.

Topics: Animals; Bronchial Hyperreactivity; Dendritic Cells; Disease Models, Animal; Epoprostenol; Female; Inflammation; Methacrylates; Mice; Mice, Inbred BALB C; Ovalbumin; Pyridines; Thromboxane-A Synthase; Thromboxanes

Several previous studies have demonstrated that some helminth infections can inhibit allergic reactions, but the examination on the effect of live Schistosoma japonicum (SJ) infection on allergic inflammation remains limited. The aim of this study was to examine the effect and mechanism of chronic SJ infection on airway allergic inflammation in a murine model. The data showed that chronic SJ infection suppressed airway eosinophilia, mucus production and antigen-specific IgE responses induced by ovalbumin (OVA) sensitization and challenge. Cytokine production analysis showed that chronic SJ infection reduced allergen-driven interleukin (IL)-4 and IL-5 production, but had no significant effect on IFN-gamma production. More importantly, we found that the adoptive transfer of dendritic cells (DCs) from SJ-infected mice dramatically decreased airway allergic inflammation in the recipients, which was associated with significant decrease of IL-4/IL-5 production and increase of IL-10 production. The results suggest that SJ infection may inhibit the development of allergy and that DCs may be involved in the process of helminth infection-mediated modulation of allergic inflammation.

Topics: Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Dendritic Cells; Eosinophilia; Female; Humans; Immunomodulation; Inflammation; Interleukins; Lung; Mice; Mucus; Ovalbumin; Schistosoma japonicum; Schistosomiasis japonica; Trachea

The signal transducer and activator of transcription (STAT) family of molecules play a critical role in the signaling of many cytokines. In addition to STAT6, implication of STAT1 and STAT3 in the development of airway hyperresponsiveness (AHR) has also been suggested in allergic bronchial asthma. However, there is little information whether or not antigen challenge really causes the in vivo activation of these STAT molecules in bronchial smooth muscles (BSMs). In the present study, the activations of these STATs were examined in BSMs of mice with allergic bronchial asthma. Male BALB/c mice were sensitized and repeatedly challenged with ovalbumin (OA) antigen. Total protein samples of the left main bronchi were prepared at 3 after the last OA challenge, and Western blot analyses for total and tyrosine-phosphorylated STATs molecules were conducted. In addition to the phosphorylation of STAT6, a significant increase in the level of phosphorylated STAT1 was also observed after the antigen exposure. In contrast, no significant increase in the level of phosphorylated STAT3 was observed in this mouse model of allergic bronchial asthma. The antigen exposure did not change the protein expressions of these STATs themselves. These findings suggest that STAT6 and STAT1, but not STAT3, might be crucial signal transducers in the development of BSM hyperresponsiveness, one of the causes of AHR in asthmatics.

Topics: Administration, Inhalation; Animals; Antigens; Asthma; Bronchi; Bronchial Hyperreactivity; Male; Mice; Mice, Inbred BALB C; Models, Animal; Muscle, Smooth; Ovalbumin; Phosphorylation; Signal Transduction; STAT1 Transcription Factor; STAT3 Transcription Factor; STAT6 Transcription Factor

The prevalence of asthma continues to increase in westernized countries, and optimal treatment remains a significant therapeutic challenge. Recently, CD1d-restricted invariant NKT (iNKT) cells were found to play a critical role in the induction of airway hyperreactivity (AHR) in animal models and are associated with asthma in humans. To test whether iNKT cell-targeted therapy could be used to treat allergen-induced airway disease, mice were sensitized with OVA and treated with di-palmitoyl-phosphatidyl-ethanolamine polyethylene glycol (DPPE-PEG), a CD1d-binding lipid antagonist. A single dose of DPPE-PEG prevented the development of AHR and pulmonary infiltration of lymphocytes upon OVA challenge, but had no effect on the development of OVA-specific Th2 responses. In addition, DPPE-PEG completely prevented the development of AHR after administration of alpha-galactosylceramide (alpha-GalCer) intranasally. Furthermore, we demonstrate that DPPE-PEG acts as antagonist to alpha-GalCer and competes with alpha-GalCer for binding to CD1d. Finally, we show that DPPE-PEG completely inhibits the alpha-GalCer-induced phosphorylation of ERK tyrosine kinase in iNKT cells, suggesting that DPPE-PEG specifically blocks TCR signaling and thus activation of iNKT cells. Because iNKT cells play a critical role in the development of AHR, the inhibition of iNKT activation by DPPE-PEG suggests a novel approach to treat iNKT cell-mediated diseases such as asthma.

Topics: Allergens; Animals; Antigens, CD1d; Binding, Competitive; Bronchial Hyperreactivity; Cell Line; Disease Models, Animal; Female; Galactosylceramides; Humans; Immunosuppressive Agents; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Natural Killer T-Cells; Ovalbumin; Phosphatidylethanolamines; Polyethylene Glycols

Anoectochilus formosanus HAYATA, a Chinese herb, is a valued folk medicine for fever, pain, and diseases of the lung and liver. Allergic asthma is characterized by increased serum IgE level and inflammation of the airways with high levels of interleukin (IL)-4 and IL-5 in bronchoalveolar lavage fluids (BALF). Constriction of airway smooth muscle and development of airway hyperresponsiveness (AHR) are the most important symptoms of allergic asthma. In our previous study, a standardized aqueous extract of A. formosanus (SAEAF) was used to modulate innate immunity of normal mice. In this study, airway inflammatory infiltrations, including T cell differentiation, cytokine modulation, allergic antibodies estimation, pulmonary pathology, and enhanced pause (Penh) of AHR were used to evaluate SAEAF treatment of an ovalbumin (OVA)-inhaled airway allergic murine model. The resulting cytokine profiles demonstrated that SAEAF can significantly reduce Th2 polarization after administration of SAEAF in OVA inhalation. These results also suggest that SAEAF modulates cytokine secretion in allergic asthma. Modulated natural T regulatory cells (CD25+/CD4+, Treg) were also shown to increase immuno-suppression in the allergic lung inflammation and further down-regulate airway inflammatory infiltration in eosinophils and macrophages. Finally, decreased airway anti-OVA IgE secretion and reduced AHR were observed. Our results indicate that the administration of SAEAF can modulate cytokines and T cell subpopulation by regulating inflammatory cell infiltration and modulating the allergic response.

Topics: Animals; Anti-Inflammatory Agents; Antigens; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Drugs, Chinese Herbal; Eosinophils; Hypersensitivity; Immunoglobulin E; Immunosuppressive Agents; Lung; Macrophages; Male; Mice; Mice, Inbred BALB C; Orchidaceae; Ovalbumin; Phytotherapy; Reference Values; T-Lymphocytes, Regulatory; Th2 Cells

The herbal Isatis tinctoria extract (ITE) inhibits the inducible isoform of cyclooxygenase (COX-2) as well as lipoxygenase (5-LOX) and therefore possesses anti-inflammatory properties. The extract might also be useful in allergic airway diseases which are characterized by chronic inflammation.. ITE obtained from leaves by supercritical carbon dioxide extraction was investigated in ovalbumin (OVA) immunised BALB/c mice given intranasally together with antigen challenge in the murine model of allergic airway disease (asthma) with the analysis of the inflammatory and immune parameters in the lung.. ITE given with the antigen challenge inhibited in a dose related manner the allergic response. ITE diminished airway hyperresponsiveness (AHR) and eosinophil recruitment into the bronchoalveolar lavage (BAL) fluid upon allergen challenge, but had no effect in the saline control mice. Eosinophil recruitment was further assessed in the lung by eosinophil peroxidase (EPO) activity at a dose of 30 microg ITE per mouse. Microscopic investigations revealed less inflammation, eosinophil recruitment and mucus hyperproduction in the lung in a dose related manner. Diminution of AHR and inflammation was associated with reduced IL-4, IL-5, and RANTES production in the BAL fluid at the 30 microg ITE dose, while OVA specific IgE and eotaxin serum levels remained unchanged.. ITE, which has been reported inhibiting COX-2 and 5-LOX, reduced allergic airway inflammation and AHR by inhibiting the production of the Th2 cytokines IL-4 and IL-5, and RANTES.

Topics: Allergens; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Eosinophil Peroxidase; Eosinophils; Hypersensitivity; Immunoglobulin E; Inflammation; Inflammation Mediators; Isatis; Lung; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Phytotherapy; Plant Extracts; Plant Leaves

The development of nanotechnology has increased the risk of environmental exposure to types of particles other than those derived from combustion, namely, industrial nanomaterials. Patients with bronchial asthma are sensitive to inhaled substances, including particulate matter. This study examined the effects of pulmonary exposure to a type of nano-sized carbon nanotube (single-walled nanotubes (SWCNT)) on allergic airway inflammation and sought their cellular mechanisms. In the in vivo experiments, ICR mice were divided into four experimental groups that were repeatedly administered vehicle, SWCNT (50 microg/animal), ovalbumin (OVA; an allergen), or OVA + SWCNT through an intratracheal route and thereafter assayed. SWCNT aggravated allergen-induced pulmonary inflammation with mucus hyperplasia. SWCNT with allergen amplified lung protein levels of T helper (Th) cytokines and chemokines related to allergy and exhibited adjuvant activity for allergen-specific IgG(1) (and IgE) compared with allergen alone. SWCNT accentuated the level/activity of oxidative stress-related biomarkers in the airways in the presence of allergen. In vitro, SWCNT can partially promote/strengthen the maturation/activation/function of bone marrow-derived dendritic cells (DC). Together, these results suggest that SWCNT can exacerbate murine allergic airway inflammation via enhanced activation of Th immunity and increased oxidative stress. In addition, this exacerbation may be partly through the inappropriate activation of antigen-presenting cells, including DC.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Progression; Environmental Exposure; Hyperplasia; Immunity, Cellular; Immunoglobulin E; Immunoglobulin G; Inflammation; Lung; Male; Mice; Mice, Inbred ICR; Nanotubes, Carbon; Ovalbumin; Oxidative Stress; Respiratory Mucosa

Allergic asthma is characterized by reversible airway obstruction, lung inflammation, and airway hyperresponsiveness (AHR). Previous studies using leukotriene B(4) (LTB(4)) receptor 1-deficient mice and adoptive transfer experiments have suggested that LTB(4) plays a role in lung inflammation and AHR.. In this study, we used a leukotriene A(4) hydrolase (LTA(4)H) inhibitor as a pharmacological tool to directly examine the role of LTB(4) in a mast cell-dependent murine model of allergic airway inflammation.. We used the forced oscillation technique to test the effects of an LTA(4)H inhibitor dosed during the challenge phase on AHR. Lung tissue and lavage were collected for analysis.. Treatment with an LTA(4)H inhibitor improved multiple parameters encompassing AHR and lung function. Significant decreases in inflammatory leukocytes, cytokines, and mucin were observed in the lung lumen. Serum levels of antigen-specific IgE and IgG1 were also decreased. Labeled antigen uptake by lung dendritic cells and subsequent trafficking to draining lymph nodes and the lung were decreased on LTA(4)H inhibitor treatment. Provocatively, inhibition of LTA(4)H increased lipoxin A(4) levels in lung lavage fluid.. These data suggest that LTB(4) plays a key role in driving lung inflammation and AHR. Mechanistically, we provide evidence that inhibition of LTA(4)H, affects recruitment of both CD4(+) and CD8(+) T cells, as well as trafficking of dendritic cells to draining lymph nodes, and may beneficially modulate other pro- and antiinflammatory eicosanoids in the lung. Inhibition of LTA(4)H is thus a potential therapeutic strategy that could modulate key aspects of asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cytokines; Epoxide Hydrolases; Immunoglobulin E; Leukotriene B4; Mast Cells; Mice; Mice, Inbred BALB C; Mucins; Ovalbumin

During asthma, lung DC capture and process antigens to initiate and maintain allergic Th2 cell responses to inhaled allergens. The aim of the study was to investigate whether allergen-specific IgG, generated during sensitization, can potentiate the acute airway inflammation through Fcgamma receptor (FcgammaR)-mediated antigen uptake and enhance antigen presentation resulting in augmented T-cell proliferation. We examined the impact of antigen presentation and T-cell stimulation on allergic airway hyperresponsiveness and inflammation using transgenic and gene-deficient mice. Both airway inflammation and eosinophilia in bronchoalveolar lavage fluid were markedly reduced in sensitized and challenged FcgammaR-deficient mice. Lung DC of WT, but not FcgammaR-deficient mice, induced increased antigen-specific CD4+ T-cell proliferation when pulsed with anti-OVA IgG immune complexes. Intranasal application of anti-OVA IgG immune complexes resulted in enhanced airway inflammation, eosinophilia and Th2 cytokine release, mediated through enhanced antigen-specific T-cell proliferation in vivo. Finally, antigen-specific IgG in the serum of sensitized mice led to a significant increase of antigen-specific CD4+ T-cell proliferation induced by WT, but not FcgammaR-deficient, lung DC. We conclude that FcgammaR-mediated enhanced antigen presentation and T-cell stimulation by lung DC has a significant impact on inflammatory responses following allergen challenge in asthma.

Topics: Adoptive Transfer; Animals; Antigen Presentation; Asthma; Bronchial Hyperreactivity; Dendritic Cells; Female; Immunoglobulin G; Inflammation; Lung; Lymphocyte Activation; Lymphokines; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Peptide Fragments; Receptors, IgG; Th2 Cells; Toll-Like Receptor 4

Mouse models of asthma suffer from the necessity to prime the animals by injections before respiratory exposure. Our aim was to develop a mouse model that mimics the progression of human allergic disease upon low-dose inhaled allergen exposure.. Mice were primed intraperitoneally to ovalbumin (OVA) before they were exposed repeatedly to aerosols of either OVA, ryegrass (Lolium perenne) pollen extract, or both concomitantly. The sensitization to ryegrass pollen proteins was evaluated by measurement of specific serum antibody, by the respiratory response to a challenge with ryegrass pollen extract and by lung cytokine production after challenge.. Inhalation of ryegrass pollen extract alone did not result in sensitization. Sensitization to inhaled ryegrass pollen proteins, however, did occur in mice that had been sensitized to OVA by intraperitoneal injections and were then exposed to inhaled ryegrass pollen extract and OVA simultaneously. T and B cell priming was ascertained by ryegrass pollen-specific IgG1 and IgE antibody production and by induction of airway inflammation and of Th2 cytokine mRNA transcripts in the lungs upon airway challenge with ryegrass pollen extract. A progressive spread of the IgE/IgG1 response to different ryegrass pollen proteins could be visualized in immunoblots by comparing antibody patterns at day 56 and 86.. Low-dose inhalatory allergen exposure results in sensitization when airways are exposed at the same time to another allergen to which the animals are already sensitized. This model can help to unravel the mechanisms that underlie the development and progression of respiratory allergic diseases.

Topics: Airway Resistance; Animals; Antigens, Plant; Blotting, Western; Bronchial Hyperreactivity; Bronchial Provocation Tests; Disease Models, Animal; Gene Expression; Immunization; Immunoglobulin E; Immunoglobulin G; Inflammation; Interleukin-13; Interleukin-4; Interleukin-5; Lolium; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Plant Extracts; Plant Proteins; Pollen; Respiratory Hypersensitivity; Th2 Cells

In this study, we explored whether prolonged ovalbumin (OVA) exposure in sensitized mice created an environment suitable for Th17 differentiation.. BALB/c and C57BL/6 mice (n = 36), after intraperitoneal OVA sensitization on days 0 and 12, received prolonged OVA aerosol challenges up to day 55. Airway inflammatory cell levels, cytokine profiles, and Th17 cell infiltration were evaluated after sacrifice.. Prolonged OVA challenge caused inflammation that was characterized by raised influxes of airway macrophages and neutrophils. Following long-term exposure, Th17 cytokines and Th17 cell numbers progressively increased in the lung (P < 0.05) along with increased production of Th17 polarization-related factors, including TGF-beta, IL-6, and IL-23. The lineage-specific transcription factor for Th17 subsets, RORgammat, displayed similar upregulation throughout the OVA challenge (P < 0.05).. Prolonged OVA challenge induces an environment that facilitates Th17 polarization.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Differentiation; Cell Polarity; Cytokines; Interleukin-23; Interleukin-6; Lung; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Nuclear Receptor Subfamily 1, Group F, Member 3; Ovalbumin; T-Lymphocyte Subsets; T-Lymphocytes, Helper-Inducer; Transforming Growth Factor beta

Asthma is an epithelial disorder in which T helper 2 (Th2)-type inflammation has a prominent role. Recent studies indicated that a cytokine, thymic stromal lymphopoietin (TSLP), is essential for the development of antigen-induced asthma. The authors used ovalbumin (OVA) sensitization and challenge to induce a murine asthmatic model. The model was confirmed by airway hyperresponsiveness, serum levels of total and OVA-specific immunoglobulin (IgE), histological analysis of lung tissues. The authors found that expression of TSLP was significantly increased in both mRNA and protein levels in mice lungs treated with OVA. The expression of CD40, CD80, and CD86 in bronchoalveolar lavage fluid (BALF) was increased in mice with OVA. Tight correlation between TSLP mRNA and interleukin (IL)-4, IL-5, and IL-13 in BALF was identified. Furthermore, treating mice with TSLP-neutralizing antibody reduced the expression of TSLP mRNA of lungs, CD40, CD80, and CD86 on dendritic cells, and IL-4, IL-5, and IL-13 in the OVA group. This study indicates that TSLP is increased in the airway epithelium in mice treated with OVA. In the lung inflammation model, TSLP activates dendritic cells (DCs) via up-regulation of CD40, CD80, and CD86, then induces the differentiation of prime naive CD4(+) T cells to become proinflammatory Th2 cells. Blocking TSLP is capable of inhibiting the production of Th2 cytokines, thus presents a promising strategy for the treatment of asthma.

Topics: Animals; Antibodies; Asthma; B7-1 Antigen; B7-2 Antigen; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD40 Antigens; Cytokines; Dendritic Cells; Gene Expression; Immunization; Immunoglobulin E; Inflammation; Interleukin-13; Interleukin-4; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Mucosa; Th2 Cells; Thymic Stromal Lymphopoietin

The mechanisms by which cannabinoid receptors CB(1) and CB(2) modulate immune function are not fully elucidated. Critical tools for the determination of the role of both receptors in the immune system are CB(1)/CB(2) double null mice (CB(1)/CB(2) null), and previous studies have shown that CB(1)/CB(2) null mice exhibit exaggerated responses to various immunological stimuli. The objective of these studies was to determine the magnitude to which CB(1)/CB(2) null mice responded to the respiratory allergen ovalbumin (OVA) as compared with wild-type C57BL/6 mice. The authors determined that in the absence of adjuvant, both wild-type and CB(1)/CB(2) null mice mounted a marked response to intranasally instilled OVA as assessed by inflammatory cell infiltrate in the bronchoalveolar lavage fluid (BALF), eosinophilia, induction of mucous cell metaplasia, and IgE production. Many of the endpoints measured in response to OVA were similar in wild-type versus CB(1)/CB(2) null mice, with exceptions being modest reductions in OVA-induced IgE and attenuation of BALF neutrophilia in CB(1)/CB(2) null mice as compared with wild-type mice. These results suggest that T-cell responses are not universally exaggerated in CB(1)/CB(2) null mice.

Topics: Adjuvants, Immunologic; Administration, Intranasal; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Enzyme-Linked Immunosorbent Assay; Female; Immunoglobulin E; Immunoglobulin G; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Receptor, Cannabinoid, CB1; Receptor, Cannabinoid, CB2; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Expression of the metalloprotease ADAM8 is increased in patients with asthma, but the functional significance of elevated ADAM8 expression in the context of asthma pathogenesis remains elusive.. To study development of asthma in ADAM8-deficient mice.. Ovalbumin-induced asthma was studied in wild-type, ADAM8-deficient, and ADAM8-chimeric mice. Lung inflammation was assessed by histology, analysis of bronchoalveolar lavage, and airway hyperresponsiveness.. ADAM8-deficient mice are highly resistant to the development of ovalbumin-induced airway inflammation and hyperresponsiveness. ADAM8 expression was induced in both hematopoietic cells and the nonhematopoietic microenvironment after induction of asthma, and ADAM8 expression in both cell populations was required for the full manifestation of asthma. Interestingly, loss of ADAM8 on T cells alone was sufficient to significantly decrease the asthma response. The attenuated response was not due to an intrinsic defect in antigen presentation or cytokine production but reflected an impaired migration of T cells, eosinophils, CD11b(+) CD11c(-), and CD11c(+) cells from blood vessels to the lung and alveolar space, suggesting a general hematopoietic cell deficiency in the absence of ADAM8.. The results show that ADAM8 plays a proinflammatory role in airway inflammation. The milder disease outcome in the absence of ADAM8 suggests that this protein might be an interesting new target in treatment of this, and potentially other, inflammatory diseases in which recruitment of inflammatory cells is an essential part of pathogenesis.

Topics: ADAM Proteins; Animals; Antigens, CD; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Differentiation; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Inflammation; Membrane Proteins; Mice; Mice, Inbred C57BL; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction

Probiotics have been shown to be effective in reducing allergic symptoms. However, there are few studies to evaluate the therapeutic effects of lactobacilli on allergen-induced airway inflammation.. We investigated whether three heat-killed lactobacilli, Lactobacillus plantarum, Lactobacillus curvatus and Lactobacillus sakei subsp. sakei, isolated from kimchi, exerted inhibitory effects on airway hyper-responsiveness in a murine asthma model.. Heat-killed lactic acid bacteria were orally administered into BALB/c mice, followed by challenge with aerosolized ovalbumin, after which allergic symptoms were evaluated.. Airway inflammation was suppressed in the L. plantarum- and L. curvatus-treated mice. Interleukin (IL)-4 and IL-5 levels were significantly lower in the L. plantarum- and L. curvatus-treated mice than in those treated with L. sakei subsp. sakei. Importantly, heat-killed L. plantarum administration induced Foxp3 expression in intestinal lamina propria cells, and heat-killed L. curvatus induced IL-10 as a way of inducing tolerance.. Specific strains of lactobacilli isolated from kimchi can effectively suppress airway hyper-responsiveness.

Topics: Administration, Oral; Animals; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Cytokines; Forkhead Transcription Factors; Hot Temperature; Immunization; Lactobacillus; Medicine, Korean Traditional; Mice; Mice, Inbred BALB C; Mucous Membrane; Ovalbumin

The prevalence of asthma has increased worldwide. The reasons for this rise remain unclear. Oxidative stress plays an important role in the pathogenesis of asthma. Glutathione (GSH) is the major representative of the class of nonprotein thiols and plays a pivotal role in a variety of enzymatic and nonenzymatic reactions that protect tissues against oxidative stress. In antioxidative reactions, GSH is converted into its oxidized form, glutathione disulfide (GSSG) that in its turn is enzymatically reduced into GSH to maintain a physiological redox balance. We used a guinea pig model of asthma to assess whether the early asthmatic reaction is associated with decreased lung levels of glutathione, and whether decreased glutathione is implicated in the increased airway smooth muscle reactivity that is associated with exposure of the lungs to allergen. Lung glutathione levels were decreased immediately after the onset of the early asthmatic reaction in vivo and associated with the release of 8-iso-PGF(2alpha), an indicator for oxidative stress. Glutathione ethylester, a glutathione precursor, blunted the airway obstruction during an early asthmatic reaction in a perfusion model and glutathione depletion rendered the airways hyperreactive. Glutathione ethyl ester in the buffer prevented this hyperreactivity. These results indicate that glutathione can modulate the early asthmatic reaction as well as the airway hyperresponsiveness.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Glutathione; Guinea Pigs; Histamine; Male; Ovalbumin; Oxidative Stress; Time Factors

The mechanisms leading to airway hyper-responsiveness (AHR) in asthma are still not fully understood. AHR could be produced by hypersensitivity of the airway smooth muscle or hyperreactivity of the airways. This study was conducted to ascertain whether AHR in a murine model of asthma is produced by changes at the level of the airway smooth muscle. Airway smooth muscle responses were characterised in vitro in isolated trachea spirals from naive mice and from an acute ovalbumin (OVA) challenge model of allergic asthma. AHR was investigated in vivo in conscious, freely moving mice. Inflammatory cell influx into the lungs and antibody responses to the antigen were also measured. In vitro study of tracheal airway smooth muscle from naïve mice demonstrated concentration-related contractions to methacholine and 5-HT, but no responses to histamine or adenosine or its stable analogue, 5'-N-ethyl-carboxamidoadenosine. The contractions to 5-HT were inhibited by ketanserin and alosetron indicating involvement of 5-HT(2A) and 5-HT(3) receptors, respectively. In an acute model of allergic asthma, OVA-treated mice were shown to be atopic by inflammatory cell influx to the lungs after OVA challenge, increases in total IgE and OVA-specific IgG levels and contractions to OVA in isolated trachea. In the asthmatic model, AHR to methacholine was demonstrated in conscious, freely moving mice in vivo and in isolated trachea in vitro 24 and 72h after OVA challenge. No AHR in vitro was seen for 5-HT, histamine or adenosine. These results suggest that, in our mouse model of asthma, changes occur at the level of the muscarinic receptor transduction pathway of coupling to airway smooth muscle contraction. These changes are maintained when tissues are removed from the inflammatory environment and for at least 3 days.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Immunoglobulin E; Male; Mice; Mice, Inbred BALB C; Muscle Contraction; Muscle, Smooth; Ovalbumin; Receptor, Serotonin, 5-HT2A; Receptors, Muscarinic; Receptors, Serotonin, 5-HT3; Time Factors; Trachea

Engagement of T cell receptor (TCR) triggers signaling pathways that mediate activation, proliferation, and differentiation of T lymphocytes. Such signaling events are mediated by reactive oxygen species (ROS), including hydrogen peroxide and lipid peroxides, both of which are reduced by glutathione peroxidase 1 (GPx1). We have now examined the role of GPx1 in the activation, differentiation, and functions of CD4(+) T helper (Th) cells. TCR stimulation increased the intracellular ROS concentration in Th cells in a time-dependent manner, and such TCR-induced ROS generation was found to promote cell proliferation. GPx1-deficient Th cells produced higher levels of intracellular ROS and interleukin-2 than wild-type Th cells and proliferated at a faster rate than did wild-type cells. Moreover, differentiation of GPx1-deficient Th cells was biased toward Th1, and Th17 cell development was also impeded by GPx1 depletion. Consistent with these findings, GPx1-null mice were protected from the development of ovalbumin-induced allergic asthma. Eosinophil infiltration, goblet cell hyperplasia, collagen deposition, and airway hyperresponsiveness were thus all attenuated in the lungs of GPx1-null mice. These data indicate that GPx1-dependent control of intracellular ROS accumulation is important not only for regulation of Th cell proliferation but for modulation of differentiation into Th1, Th2, and Th17 cells.

Topics: Allergens; Animals; Antibodies; Antioxidants; Bronchial Hyperreactivity; CD3 Complex; Cell Differentiation; Cell Proliferation; Cells, Cultured; Collagen; Cytokines; Eosinophils; Gene Expression; Glutathione Peroxidase; Glutathione Peroxidase GPX1; Goblet Cells; Homeodomain Proteins; Interferon-gamma; Interleukin-2; Interleukin-4; Lung; Lymph Nodes; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Reactive Oxygen Species; Spleen; T-Box Domain Proteins; T-Lymphocytes, Helper-Inducer; Th1 Cells; Th17 Cells; Th2 Cells

Endogenous adenosine is released in asthmatic patients' lungs by inhaled allergen, however, its exact role in asthmatic responses or the receptors mediating these responses has not been determined. Our hypothesis was that adenosine released during allergen challenge contributes to the airways responses to inhaled allergen. The effects of selective antagonists of the four adenosine receptor subtypes were investigated on the airways responses of sensitized guinea-pigs to inhaled ovalbumin to ascertain the role of adenosine in these allergen responses, and compared with a corticosteroid, dexamethasone. Early (EAR) and late asthmatic responses (LAR) to inhaled ovalbumin (10 microg/ml) of sensitized, conscious guinea-pigs were recorded by whole body plethysmography following administration of selective adenosine receptor antagonists. Airway reactivity to inhaled histamine (1 mM) and inflammatory cell influx in bronchoalveolar lavage fluid were also determined 24 h after ovalbumin challenge. ZM241385 (A(2A) receptor antagonist) did not affect these responses, whereas DPCPX (A(1) receptor antagonist) exerted a small inhibition only of the LAR. MRS1706 (A(2B) receptor antagonist) inhibited the airways hyperreactivity and cellular influx and enhanced the EAR. MRS1220 (A(3) receptor antagonist) inhibited the airways hyperreactivity and cellular influx without affecting EAR and LAR. Dexamethasone inhibited the ovalbumin-induced late asthmatic responses, airways hyperreactivity and cellular influx. The blockade of airway hyperreactivity and inflammatory cell influx by A(2B) and A(3) receptor antagonists suggests that endogenous adenosine is released by inhaled allergen and these responses are mediated via A(2B) and A(3) receptors in guinea-pigs. The adenosine released by allergen inhalation does not, however, appear to be involved in the EAR, but it may contribute to the LAR via A(1) receptors.

Topics: Adenosine; Adenosine A2 Receptor Antagonists; Adenosine A3 Receptor Antagonists; Animals; Bronchial Hyperreactivity; Dexamethasone; Glucocorticoids; Guinea Pigs; Male; Ovalbumin; Plethysmography, Whole Body; Purinergic P1 Receptor Antagonists; Receptor, Adenosine A2B; Receptor, Adenosine A3; Receptors, Purinergic P1; Time Factors

Acute exacerbations of asthma represent a common clinical problem with major economic impact. Air pollutants including ozone have been shown to contribute to asthma exacerbation, but the mechanisms underlying ozone-induced asthma exacerbation are only partially understood. The present study aimed to develop a mouse model to gain insight into the development of airway hyperresponsiveness (AHR) to methacholine (MCh) in mice after exposure to both allergen and ozone. Mice were exposed for 20 min per day for 10 consecutive days to an aerosol of 1% ovalbumin (OVA) or saline followed by a single 3-h exposure to clean air or 100, 250, or 500 ppb ozone. Ozone induced AHR in mice previously exposed to OVA when compared to non-exposed (saline) control mice. After a 10-d exposure to OVA, a single exposure to a low (100 ppb) ozone concentration was sufficient to induce AHR. The AHR response was associated with goblet-cell metaplasia. Even the lowest concentration of ozone tested, 100 ppb, which may be exceeded in urban environments and in the workplace, resulted in a significant increase in AHR, most prominent 24 h after exposure in the OVA-exposed mice.

Topics: Air Pollutants; Allergens; Animals; Asthma; Biomarkers; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Drug Interactions; Inhalation Exposure; Leukocyte Count; Leukocytes; Lung; Methacholine Chloride; Mice; Mucus; Ovalbumin; Ozone; Periodic Acid-Schiff Reaction; Respiratory Function Tests

Allergen-induced bronchial asthma is a chronic airway disease that involves the interplay of various genes with environmental factors triggering different inflammatory pathways.. The aim of this study was to identify possible mediators of airway inflammation (AI) in a model of allergic AI via microarray comparisons and to analyse one of these mediators, Lipocalin2 (Lcn2), for its role in a murine model of allergic airway disease.. Gene microarrays were used to identify genes with at least a twofold increase in gene expression in the lungs of two separate mouse strains with high and low allergic susceptibility, respectively. Validation of mRNA data was obtained by Western blotting, followed by functional analysis of one of the identified genes, Lcn2, in mice with targeted disruption of specific gene expression. Epithelial cell cultures were undertaken to define induction requirements and possible mechanistic basis of the results observed in the Lcn2 knock-out mice.. Lcn2 was up-regulated upon allergen sensitization and airway challenges in lung tissues of both mouse strains and retraced on the protein level in bronchoalveolar lavage fluids. Functional relevance was assessed in mice genetically deficient for Lcn2, which showed enhanced airway resistance and increased AI associated with decreased apoptosis of lung inflammatory cells, compared with wild-type controls. Similarly, application of Lcn2-blocking antibodies before airway challenges resulted in increased inflammation and reduced apoptosis.. These data indicate a protective role for Lcn2 in allergic airway disease, suggesting a pro-apoptotic effect as the underlying mechanism.

Topics: Acute-Phase Proteins; Alveolar Epithelial Cells; Animals; Apoptosis; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cytokines; Disease Models, Animal; Female; Gene Expression Profiling; Inflammation Mediators; Lipocalin-2; Lipocalins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Oligonucleotide Array Sequence Analysis; Oncogene Proteins; Ovalbumin; RNA, Messenger; Time Factors; Up-Regulation

The number of amphiregulin (AR)-positive mast cells in the bronchial mucosa and the levels of AR in sputum from asthmatic patients have been reported to be increased. In addition, AR can promote mucin gene expression in human epithelial cells, suggesting that AR contributes to the pathogenesis of allergic asthma.. To elucidate the role of AR in the pathogenesis of asthma, we immunized AR-deficient mice with ovalbumin (OVA) and then induced airway inflammation in them after OVA inhalation. The OVA-induced airway inflammation was assessed on the basis of the lung histology, number of leukocytes in the bronchoalveolar lavage (BAL) fluid, Th2 cytokine levels in the BAL fluid and OVA-specific IgG1 and IgE levels in the serum and compared between AR-sufficient and -deficient mice.. The OVA-induced airway inflammation was comparable in the AR-sufficient and -deficient mice.. Amphiregulin is not essential for induction of acute airway inflammation by OVA in mice.

Topics: Amphiregulin; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; EGF Family of Proteins; Glycoproteins; Humans; Immunization; Immunoglobulin E; Immunoglobulin G; Intercellular Signaling Peptides and Proteins; Lung; Mice; Mice, Knockout; Ovalbumin

Clara cell protein (CC16), the main secretory product of bronchiolar Clara cells, plays an important protective role in the respiratory tract against oxidative stress and inflammation. The purpose of the study was to investigate the role of elemental carbon ultrafine particles (EC-UFP)-induced oxidative stress on Clara cells and CC16 in a mouse model of allergic lung inflammation.. Ovalbumin (OVA)-sensitized mice were exposed to EC-UFP (507 microg/m(3) for 24 h) or filtered air immediately prior to allergen challenge and systemically treated with N-acetylcysteine (NAC) or vehicle prior and during EC-UFP inhalation. CC16 was measured up to one week after allergen challenge in bronchoalveolar lavage fluid (BALF) and in serum. The relative expression of CC16 and TNF-alpha mRNA were measured in lung homogenates. A morphometrical analysis of mucus hypersecretion and electron microscopy served to investigate goblet cell metaplasia and Clara cell morphological alterations.. In non sensitized mice EC-UFP inhalation caused alterations in CC16 concentration, both at protein and mRNA level, and induced Clara cell hyperplasia. In sensitized mice, inhalation of EC-UFP prior to OVA challenge caused most significant alterations of BALF and serum CC16 concentration, BALF total protein and TNF-alpha relative expression compared to relevant controls; their Clara cells displayed the strongest morphological alterations and strongest goblet cell metaplasia occurred in the small airways. NAC strongly reduced both functional and morphological alterations of Clara cells.. Our findings demonstrate that oxidative stress plays an important role in EC-UFP-induced augmentation of functional and morphological alterations of Clara cells in allergic lung inflammation.

Topics: Air Pollutants; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Gene Expression; Lung; Mice; Ovalbumin; Oxidative Stress; Particulate Matter; Tumor Necrosis Factor-alpha; Uteroglobin

The dust mite Blomia tropicalis is an important source of aeroallergens in tropical areas. Although a mouse model for B. tropicalis extract (BtE)-induced asthma has been described, no study comparing different mouse strains in this asthma model has been reported. The relevance and reproducibility of experimental animal models of allergy depends on the genetic background of the animal, the molecular composition of the allergen and the experimental protocol.. This work had two objectives. The first was to study the anti-B. tropicalis allergic responses in different mouse strains using a short-term model of respiratory allergy to BtE. This study included the comparison of the allergic responses elicited by BtE with those elicited by ovalbumin in mice of the strain that responded better to BtE sensitization. The second objective was to investigate whether the best responder mouse strain could be used in an experimental model of allergy employing relatively low BtE doses.. Groups of mice of four different syngeneic strains were sensitized subcutaneously with 100 microg of BtE on days 0 and 7 and challenged four times intranasally, at days 8, 10, 12, and 14, with 10 microg of BtE. A/J mice, that were the best responders to BtE sensitization, were used to compare the B. tropicalis-specific asthma experimental model with the conventional experimental model of ovalbumin (OVA)-specific asthma. A/J mice were also sensitized with a lower dose of BtE.. Mice of all strains had lung inflammatory-cell infiltration and increased levels of anti-BtE IgE antibodies, but these responses were significantly more intense in A/J mice than in CBA/J, BALB/c or C57BL/6J mice. Immunization of A/J mice with BtE induced a more intense airway eosinophil influx, higher levels of total IgE, similar airway hyperreactivity to methacholine but less intense mucous production, and lower levels of specific IgE, IgG1 and IgG2 antibodies than sensitization with OVA. Finally, immunization with a relatively low BtE dose (10 microg per subcutaneous injection per mouse) was able to sensitize A/J mice, which were the best responders to high-dose BtE immunization, for the development of allergy-associated immune and lung inflammatory responses.. The described short-term model of BtE-induced allergic lung disease is reproducible in different syngeneic mouse strains, and mice of the A/J strain was the most responsive to it. In addition, it was shown that OVA and BtE induce quantitatively different immune responses in A/J mice and that the experimental model can be set up with low amounts of BtE.

Topics: Administration, Intranasal; Allergens; Animals; Antigens, Plant; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Dose-Response Relationship, Immunologic; Immunoglobulin E; Injections, Subcutaneous; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred CBA; Ovalbumin; Pulmonary Eosinophilia; Pyroglyphidae; Rats; Rats, Wistar; Species Specificity; Time Factors

Natural killer T (NK T) cells have been shown to play an essential role in the development of allergen-induced airway hyperresponsiveness (AHR) and/or airway inflammation in mouse models of acute asthma. Recently, NK T cells have been reported to be required for the development of AHR in a virus induced chronic asthma model. We investigated whether NK T cells were required for the development of allergen-induced AHR, airway inflammation and airway remodelling in a mouse model of chronic asthma. CD1d-/- mice that lack NK T cells were used for the experiments. In the chronic model, AHR, eosinophilic inflammation, remodelling characteristics including mucus metaplasia, subepithelial fibrosis and increased mass of the airway smooth muscle, T helper type 2 (Th2) immune response and immunoglobulin (Ig)E production were equally increased in both CD1d-/- mice and wild-type mice. However, in the acute model, AHR, eosinophilic inflammation, Th2 immune response and IgE production were significantly decreased in the CD1d-/- mice compared to wild-type. CD1d-dependent NK T cells may not be required for the development of allergen-induced AHR, eosinophilic airway inflammation and airway remodelling in chronic asthma model, although they play a role in the development of AHR and eosinophilic inflammation in acute asthma model.

Topics: Acute Disease; Airway Remodeling; Airway Resistance; Allergens; Animals; Antigens, CD1d; Asthma; Bronchial Hyperreactivity; Bronchitis; Chronic Disease; Disease Models, Animal; Female; Fibrosis; Immunoglobulin E; Male; Metaplasia; Mice; Mice, Inbred BALB C; Mice, Knockout; Muscle, Smooth; Natural Killer T-Cells; Ovalbumin; Pulmonary Eosinophilia; Th2 Cells

The aim was to optimize antigen challenge for induction of airway hyperresponsiveness (AHR) and inflammation in BALB/c mice sensitized to ovalbumin (OVA). Comparisons were made between mice challenged with OVA either as an aerosol or intranasally. The protocol that induced maximal AHR in BALB/c mice was thereafter tested in C57BL/6 mice.. Methacholine responsiveness was measured using the flexiVent® system to assess AHR. Inflammatory responses were investigated by histology and cell counts in bronchoalveolar lavage (BAL) fluid.. 48 h after challenge with 1 or 6% OVA aerosols, there were similar increments in AHR and BAL cells, predominantly eosinophils. When comparing the effect of 1% OVA aerosol on AHR and cell infiltration at 24 and 48 h after challenge, the responses were similar. At 24 h, intranasal OVA administration (20-200 μg) caused a dose-dependent increase in AHR. BAL cells were increased by all intranasal OVA doses and to a greater extent than after 1% OVA aerosol challenge but without any dose dependency. Histological examination confirmed that there was an increase of eosinophils in lung tissue following either challenge. In C57BL/6 mice, baseline tissue elastance was the only functional outcome that was increased after intranasal OVA challenge. Even though the AHR response was negligible in C57BL/6 mice, a similar infiltration of BAL cells was observed in both strains.. Intranasal challenge was more effective than aerosol challenge at inducing both AHR and airway inflammation in BALB/c mice. Although intranasal challenge caused airway inflammation in C57BL/6 mice, this strain is not optimal for studying AHR.

Topics: Administration, Intranasal; Aerosols; Animals; Bronchial Hyperreactivity; Disease Models, Animal; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin

nTregs prevent autoimmunity and modulate immune and inflammatory responses to foreign antigens. CD4(+)Foxp3(+) nTregs from DO11.10 mice were expanded ex vivo, and their effectiveness in suppressing the development of lung inflammatory responses, elicited by differentiated CD4(+) T cells following antigen inhalation, was examined. Effector DO11.10 CD4(+) Th2 cells, when adoptively transferred into BALB/c mice that subsequently inhaled OVA, elicited a pronounced pulmonary, eosinophilic inflammation. Surprisingly, the cotransfer of expanded nTregs failed to suppress the Th2-mediated airway inflammation. Nevertheless, expanded OVA-specific CD4(+)Foxp3(+) nTregs were highly effective at inhibiting the polarization of naïve CD4(+) T cells into a Th2 phenotype. This suppression was reversed by an antibody to GITR but was not affected by the presence of the soluble OX40L. Further analysis revealed that although nTregs also failed to inhibit the lung neutrophilic inflammation induced by effector CD4(+) Th1 cells, they markedly suppressed pulmonary inflammation elicited by CD4(+) Th17 cells but not AHR. The suppression of the Th17-mediated response was evident from a striking reduction in the proportion of OVA-specific T cells expressing IL-17 and the numbers of neutrophils present in the airways of Th17 recipient mice. Collectively, these results demonstrate that expanded nTregs clearly limit the Th2 polarization process and that Th17-mediated inflammatory responses are particularly prone to the immunoregulatory properties of nTregs. These findings thus indicate that expanded nTregs are restrictive in their ability to suppress airway inflammatory processes and AHR.

Topics: Animals; Bronchial Hyperreactivity; CD4 Antigens; CD4-Positive T-Lymphocytes; Cell Differentiation; Cell Polarity; Cell Proliferation; Epitopes; Forkhead Transcription Factors; Interleukin-17; Interleukin-2 Receptor alpha Subunit; Interleukin-4; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; T-Lymphocytes, Regulatory; Th1 Cells; Th2 Cells

The authors have recently demonstrated that alveolar macrophages (AMs) are important in protecting against early phase reactions and airway hyperresponsiveness following allergen challenge. To further understand the mechanisms involved, the authors investigated the capacity of AMs to modulate airway inflammation and cytokine levels in bronchoalveolar lavage (BAL). AMs from allergy-susceptible Brown Norway (BN) rats or allergy-resistant Sprague-Dawley (SD) rats were transferred into AM-depleted BN rats 24 hours prior to allergen challenge. Methacholine-induced airway hyperresponsiveness was examined 24 hours following ovalbumin challenge. Total cells, cell types, and cytokine levels (tumor necrosis factor [TNF], interleukin [IL]-4, IL-10, IL-12 and IL-13) in BAL were measured 24 hours after allergen challenge. The transfer of AMs from SD rats into AM-depleted BN rats 24 hours before allergen challenge eliminated methacholine-induced airway hyperresponsiveness, but did not modify the number and the type of inflammatory cells in BAL. Levels of IL-13 and TNF were significantly higher in BAL of BN rats compared with SD rats. Interestingly, IL-13 and TNF levels were significantly increased and inhibited, respectively, in BN rats that received AMs from SD rats compared with BN rats. Our data suggest that AM modulation of cytokine milieu is involved in the reduction of airway hyperresponsiveness.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cytokines; Disease Models, Animal; Interleukins; Macrophages, Alveolar; Male; Methacholine Chloride; Ovalbumin; Rats; Rats, Inbred BN; Rats, Sprague-Dawley; Time Factors; Tumor Necrosis Factor-alpha

Adoptive cell transfer of an ovalbumin (OVA)-specific T(h)17-polarized cell population from transgenic DO11.10 mice into BALB/c mice followed by OVA inhalation caused airway hyperresponsiveness (AHR) with severe neutrophilia. The transferred T(h)17 cell population-previously polarized in vitro with IL-6, transforming growth factor-beta and IL-23-contained negligible numbers of IFN-gamma-producing cells; however, during T(h)17-cell-dependent airway inflammation, significant numbers of IFN-gamma-producing cells-including cells producing both IL-17 and IFN-gamma and cells producing only IFN-gamma-were detected in the lung in addition to cells producing only IL-17. Using T(h)17-polarized cell populations derived from IL-17(-/-) or IFN-gamma(-/-) mice, it was demonstrated that IL-17 is essential for inducing neutrophilic airway inflammation and that IFN-gamma is required for the AHR elevation. IFN-gamma appeared to be derived from cells producing both IL-17 and IFN-gamma and/or from cells producing only IFN-gamma, which were converted from the transferred T(h)17-polarized cell population. We also found that mAbs that neutralize IL-12 significantly suppressed the conversion of the T(h)17-polarized cell population toward IFN-gamma producers in the lung; concomitantly, with this decreased conversion, IL-12 neutralization also attenuated the AHR elevation in the lung. IL-12-dependent conversion of the transferred T(h)17-polarized cell population into IFN-gamma producers in the lung thus appeared to be a crucial process for inducing AHR elevation in T(h)17-cell-dependent airway inflammation.

Topics: Adoptive Transfer; Animals; Antibodies, Blocking; Bronchial Hyperreactivity; Cell Movement; Cells, Cultured; Disease Models, Animal; Interferon-gamma; Interleukin-12; Interleukin-17; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Neutrophils; Ovalbumin; Receptors, Antigen, T-Cell; T-Lymphocyte Subsets; T-Lymphocytes, Helper-Inducer

Respiratory viral infections have been implicated in exacerbations of allergic asthma, characterized by a Th2-biased immune response. Respiratory viruses target airway epithelial cells and dendritic cells (DCs). Their activation is, at least in part, mediated by the TLR3-dependent recognition of virus-derived dsRNA. To elucidate the role of epithelial cells and DCs and the implication of TLR3/Toll/IL-1R domain-containing adaptor-inducing IFN-beta (TRIF) pathway, we developed a mouse model of lung allergic exacerbation. The effect of intranasal administration of dsRNA in OVA-sensitized wild-type mice and TRIF(-/-) mice was evaluated on airway hyperresponsiveness and pulmonary inflammation. Our data demonstrated that treatment with dsRNA significantly increased the airway hyperresponsiveness, the lung inflammation, and the OVA-specific Th2 response. This was associated with an infiltrate of eosinophils, myeloid DCs, and T lymphocytes. TRIF activation was required for the development of dsRNA-induced exacerbation of the allergic reaction. Intratracheal transfer of IL-4/dsRNA/OVA-pretreated DCs also triggered exacerbation of the allergic reaction, whereas cells primed with dsRNA/OVA had a more limited effect. dsRNA-induced production of CCL20 by airway epithelium was associated with DC recruitment. In vivo and in vitro treatment with dsRNA amplified airway epithelial production of the pro-Th2 chemokines CCL11 and CCL17, their secretion being enhanced by Th2 cytokines. In conclusion, dsRNA derived from respiratory viruses trigger exacerbation of the pulmonary allergic reaction through TLR3/TRIF-dependent pathway. Moreover, Th2 cytokines participate in this process by modulating the response of airway epithelium and DCs to dsRNA.

Topics: Adaptor Proteins, Vesicular Transport; Allergens; Animals; Bronchial Hyperreactivity; Dendritic Cells; Disease Models, Animal; Female; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Respiratory Hypersensitivity; Respiratory Mucosa; RNA, Double-Stranded; RNA, Viral; Signal Transduction; Toll-Like Receptor 3

Asthma is characterized by chronic eosinophilic inflammation and hyperresponsiveness of the airways. We hypothesized that thalidomide, which has numerous immunomodulatory properties, may have anti-inflammatory effects in allergic asthma. BALB/c mice sensitized and challenged with ovalbumin (OVA) were treated orally with thalidomide (30, 100, or 300 mg/kg) or a vehicle. When thalidomide was administered to OVA-challenged mice, the number of eosinophils in bronchoalveolar lavage fluid (BALF) was significantly decreased. The numbers of inflammatory cells other than eosinophils were not reduced by thalidomide. Thalidomide inhibited the elevated levels of interleukin-5 (IL-5) and tumor necrosis factor-alpha (TNF-alpha) in BALF by OVA challenges. Histological analysis of the lung revealed that both the infiltration of inflammatory cells and the hyperplasia of goblet cells were significantly suppressed by thalidomide treatment. Furthermore, thalidomide significantly inhibited the response to methacholine induced by OVA challenges. Taken together, thalidomide treatment decreased airway inflammation and hyperresponsiveness in a murine model of allergic asthma. These results might provide an opportunity for the development of novel therapeutics to treat severe asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophils; Female; Goblet Cells; Hyperplasia; Inflammation; Interleukin-5; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Thalidomide; Tumor Necrosis Factor-alpha

CC chemokine receptor 4 (CCR4) is generally recognized as a preferential marker for T helper 2 cells, and we have previously reported morpholine-derivative CCR4 antagonists, RS-1154 and RS-1269. Here, we investigate the pharmacological profiles of a novel pyrimidine-derivative CCR4 antagonist, 2-{4-[2-(diethylamino)ethoxy]phenyl}-N-(2,4-difluorobenzyl)-5-fluoropyrimidin-4-amine (RS-1748), which showed potency to inhibit the bindings of [(125)I]CCL17 and [(35)S]GTPgammaS to human CCR4-expressing Chinese hamster ovary (CHO) cells with IC(50) values of 59.9 nM and 18.4 nM, respectively. Furthermore, RS-1748 inhibited ovalbumin-induced airway inflammation in guinea pigs at a dose of 10 mg/kg. These results indicate that RS-1748 would be a promising lead compound for developing a therapeutic agent against asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Chemokine CCL17; CHO Cells; Cricetinae; Cricetulus; Guanosine 5'-O-(3-Thiotriphosphate); Guinea Pigs; Humans; Inflammation; Inhibitory Concentration 50; Male; Ovalbumin; Pyrimidines; Receptors, CCR4

Helper T (Th) cells are deeply involved in the pathophysiology of bronchial asthma, such as eosinophilic inflammation, bronchial hyperresponsiveness (BHR), airflow limitation and remodeling. It is still unclear whether Th cells contribute to BHR independently of eosinophilic inflammation. The double GATA (dblGATA) site is a high-affinity GATA-binding site in the GATA-1 promoter. dblGATA site-deficient (Delta dblGATA) mice lack eosinophils.. Ovalbumin (OVA)-reactive Th clones were transferred into Delta dblGATA and wild-type (WT) mice of BALB/c background. The number of eosinophils in the bronchoalveolar lavage fluid (BALF) and bronchial responsiveness to methacholine were examined after OVA challenge.. The number of BALF eosinophils was significantly increased in WT mice, but not detectable in Delta dblGATA mice. BHR was also induced in WT mice, but significantly attenuated in Delta dblGATA mice.. Eosinophils are involved in T-cell-mediated BHR.

Topics: Airway Resistance; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cell Count; Eosinophils; GATA1 Transcription Factor; Lymphocytes; Macrophages; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Mutant Strains; Monocytes; Neutrophils; Ovalbumin; T-Lymphocytes, Helper-Inducer

Allergen sensitization through a disrupted skin barrier appears to play a prominent role in the development of atopic diseases, including allergic asthma. The role of the genetic background in immunological and physiological phenotypes induced by epicutaneous sensitization is undetermined.. BALB/c and C57BL/6 mice were sensitized either epicutaneously by patch application of ovalbumin (OVA) or systemically by intraperitoneal injection of OVA with alum before exposure to aerosolized OVA. The concentrations of OVA-specific immunoglobulin in serum and cytokines in bronchoalveolar lavage fluid (BALF) were measured by enzyme-linked immunosorbent assay. The severity of airway inflammation was evaluated by cell counts in BALF, and bronchial responsiveness to methacholine was measured by the flexiVent system.. The production of OVA-specific IgG1 and IgE was greater in the epicutaneously sensitized BALB/c than C57BL/6 mice. In contrast, both eosinophilic airway inflammation and bronchial responsiveness to methacholine were more prominent in the C57BL/6 than in the BALB/c mice. The concentrations of interleukin-4 increased significantly in the BALF from C57BL/6 mice only. No between-strain differences were observed after intraperitoneal sensitization.. The C57BL/6 mouse is a more appropriate model than the BALB/c mouse to study the relationship between skin barrier dysfunction and the pathogenesis of allergic asthma.

Topics: Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cell Count; Eosinophils; Immunization; Immunoglobulin E; Immunoglobulin G; Inflammation; Interferon-gamma; Interleukins; Leukocytes; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Phenotype

Invariant NKT cells (iNKT cells) play a pivotal role in the development of allergen-induced airway hyperresponsiveness (AHR) and inflammation. However, it is unclear what role they play in the initiation (sensitization) phase as opposed to the effector (challenge) phase. The role of iNKT cells during sensitization was examined by determining the response of mice to intratracheal transfer of OVA-pulsed or OVA-alpha-galactosylceramide (OVA/alphaGalCer)-pulsed bone marrow-derived dendritic cells (BMDCs) prior to allergen challenge. Wild-type (WT) recipients of OVA-BMDCs developed AHR, increased airway eosinophilia, and increased levels of Th2 cytokines in bronchoalveolar lavage fluid, whereas recipients of OVA/alphaGalCer BMDCs failed to do so. In contrast, transfer of these same OVA/alphaGalCer BMDCs into IFN-gamma-deficient (IFN-gamma(-/-)) mice enhanced the development of these lung allergic responses, which was reversed by exogenous IFN-gamma treatment following OVA-BMDC transfer. Further, Jalpha18-deficient recipients, which lack iNKT cells, developed the full spectrum of lung allergic responses following reconstitution with highly purified WT liver or spleen iNKT cells and transfer of OVA-BMDCs, whereas reconstituted recipients of OVA/alphaGalCer BMDCs failed to do so. Transfer of iNKT cells from IFN-gamma(-/-) mice restored the development of these responses in Jalpha18-deficient recipients following OVA-BMDC transfer; the responses were enhanced following OVA/alphaGalCer BMDC transfer. iNKT cells from these IFN-gamma(-/-) mice produced higher levels of IL-13 in vitro compared with WT iNKT cells. These data identify IFN-gamma as playing a critical role in dictating the consequences of iNKT cell activation in the initiation phase of the development of AHR and airway inflammation.

Topics: Adoptive Transfer; Allergens; Animals; Bone Marrow Cells; Bronchial Hyperreactivity; Cells, Cultured; Coculture Techniques; Dendritic Cells; Female; Galactosylceramides; Inflammation Mediators; Interferon-gamma; Intubation, Intratracheal; Ligands; Liver; Mice; Mice, Inbred C57BL; Mice, Knockout; Natural Killer T-Cells; Ovalbumin; Respiratory Hypersensitivity

Asthma is recognized throughout the world as a chronic airway inflammatory disease. In this study, we investigated the effect of probiotics in response to antigen challenge in an ovalbumin (OVA)-sensitized asthma model in BALB/c mice. Lactobacillus salivarius PM-A0006 was orally administered to mice before antigen challenge. After antigen challenge, serum OVA-specific antibody levels, airway responsiveness to methacholine, influx of inflammatory cells to the lung, and cytokine levels in bronchoalveolar lavage (BAL) fluid and splenocytes were assessed. Oral treatment with live L. salivarius PM-A0006 significantly attenuated the influx of eosinophils to the airway lumen and reduced the levels of serum OVA-specific IgE and eotaxin in BAL fluid of antigen-challenged animals. Furthermore, L. salivarius PM-A0006 also decreased allergen-induced airway hyperresponsiveness and elevated the levels of IFN-gamma. These results showed that oral treatment with L. salivarius PM-A0006 could have therapeutic potential in the treatment of allergic airway disease.

Topics: Administration, Oral; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Enzyme-Linked Immunosorbent Assay; Female; Immunoglobulin E; Immunoglobulin G; Lactobacillus; Mice; Mice, Inbred BALB C; Ovalbumin; Probiotics

Our previous report demonstrated that the oral administration of short-term high dose Gynostemma pentaphyllum extract (5 g/kg per day for 7days) decreased allergic reactions in ovalbumin (OVA)-sensitized mice. The aim of this study was to determine whether long-term oral administration of G. pentaphyllum attenuated airway inflammation in OVA-sensitized mice. Mice were sensitized and challenged with normal saline or OVA. OVA-sensitized mice were fed with 1.75 g/kg (low dose, GPL) or 5 g/kg (high dose, GPH) G. pentaphyllum extract, five days a week for 4 weeks. The airway hyperresponsiveness (AHR) and eosinophilia in bronchoalveolar lavage fluid (BALF) were examined. The cytokine levels or antibodies in BALF, serum and spleen cell culture supernatants were also determined. Both high and low dose extracts reduced AHR, serum OVA-IgE, and Th2-associated cytokine levels in spleen cell supernatants and BALF in OVA-sensitized mice. These results show that long-term orally administered G. pentaphyllum extract reduced allergic reactions in OVA-sensitized mice.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Eosinophils; Female; Gynostemma; Immunoglobulin E; Immunoglobulin G; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Plant Extracts; Respiratory Hypersensitivity; Spleen; Th2 Cells

Pulmonary function has been reported in mice using negative pressure-driven forced expiratory manoeuvres (NPFE) and the forced oscillation technique (FOT). However, both techniques have always been studied using separate cohorts of animals or systems. The objective of this study was to obtain NPFE and FOT measurements at baseline and following bronchoconstriction from a single cohort of mice using a combined system in order to assess both techniques through a refined approach.. Groups of allergen- or sham-challenged ovalbumin-sensitized mice that were either vehicle (saline) or drug (dexamethasone 1 mg/kg ip)-treated were studied. Surgically prepared animals were connected to an extended flexiVent system (SCIREQ Inc., Montreal, Canada) permitting NPFE and FOT measurements. Lung function was assessed concomitantly by both techniques at baseline and following doubling concentrations of aerosolized methacholine (MCh; 31.25 - 250 mg/ml). The effect of the NPFE manoeuvre on respiratory mechanics was also studied.. The expected exaggerated MCh airway response of allergic mice and its inhibition by dexamethasone were detected by both techniques. We observed significant changes in FOT parameters at either the highest (Ers, H) or the two highest (Rrs, RN, G) MCh concentrations. The flow-volume (F-V) curves obtained following NPFE manoeuvres demonstrated similar MCh concentration-dependent changes. A dexamethasone-sensitive decrease in the area under the flow-volume curve at the highest MCh concentration was observed in the allergic mice. Two of the four NPFE parameters calculated from the F-V curves, FEV0.1 and FEF50, also captured the expected changes but only at the highest MCh concentration. Normalization to baseline improved the sensitivity of NPFE parameters at detecting the exaggerated MCh airway response of allergic mice but had minimal impact on FOT responses. Finally, the combination with FOT allowed us to demonstrate that NPFE induced persistent airway closure that was reversible by deep lung inflation.. We conclude that FOT and NPFE can be concurrently assessed in the same cohort of animals to determine airway mechanics and expiratory flow limitation during methacholine responses, and that the combination of the two techniques offers a refined control and an improved reproducibility of the NPFE.

Topics: Adrenal Cortex Hormones; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Bronchoconstrictor Agents; Dexamethasone; Disease Models, Animal; Female; Forced Expiratory Flow Rates; Forced Expiratory Volume; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Oscillometry; Ovalbumin; Respiratory Function Tests; Respiratory Mechanics; Time Factors; Vital Capacity

Chronic allergic asthma is the result of a T-helper type 2 (Th2)-biased immune status. Current asthma therapies control symptoms in some patients, but a long-lasting therapy has not been established. Anti-Asthma Simplified Herbal Medicine Intervention (ASHMI™), a Chinese herbal formula, improved symptoms and lung function, and reduced Th2 responses in a controlled trial of patients with persistent moderate to severe asthma.. We evaluated the persistence of ASHMI™ beneficial effects following therapy in a murine model of chronic asthma and the immunological mechanisms underlying such effects. Methods BALB/c mice sensitized intraperitoneally with ovalbumin (OVA) received 3 weekly intratracheal OVA challenges to induce airway hyper-reactivity (AHR) and inflammation (OVA mice). Additionally, OVA mice were treated with ASHMI™ (OVA/ASHMI™) or water (OVA/sham) for 4 weeks, and then challenged immediately and 8 weeks post-therapy. In other experiments, OVA mice received ASHMI™ treatment with concomitant neutralization of IFN-γ or TGF-β. Effects on airway responses, cytokine- and OVA-specific IgE levels were determined 8 weeks post-therapy.. Before treatment, OVA mice exhibited AHR and pulmonary eosinophilic inflammation following OVA challenge, which was almost completely resolved immediately after completing treatment with ASHMI™ and did not re-occur following OVA re-challenge up to 8 weeks post-therapy. Decreased allergen-specific IgE and Th2 cytokine levels, and increased IFN-γ levels also persisted at least 8 weeks post-therapy. ASHMI™ effects were eliminated by the neutralization of IFN-γ, but not TGF-β, during therapy.. ASHMI™ induced long-lasting post-therapy tolerance to antigen-induced inflammation and AHR. IFN-γ is a critical factor in ASHMI™ effects.

Topics: Animals; Anti-Asthmatic Agents; Antibodies; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cells, Cultured; Disease Models, Animal; Drugs, Chinese Herbal; Female; Immunoglobulin E; Interferon-gamma; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Pulmonary Eosinophilia; Th2 Cells; Time Factors; Transforming Growth Factor beta

Airway remodeling and dysfunction are characteristic features of asthma thought to be caused by aberrant production of Th2 cytokines. Histamine H4 receptor (H4R) perturbation has previously been shown to modify acute inflammation and Th2 cytokine production in a murine model of asthma. We examined the ability of H4R antagonists to therapeutically modify the effects of Th2 cytokine production such as goblet cell hyperplasia (GCH), and collagen deposition in a sub-chronic model of asthma. In addition, effects on Th2 mediated lung dysfunction were also determined.. Mice were sensitized to ovalbumin (OVA) followed by repeated airway challenge with OVA. After inflammation was established mice were dosed with the H4R antagonist, JNJ 7777120, or anti-IL-13 antibody for comparison. Airway hyperreactivity (AHR) was measured, lungs lavaged and tissues collected for analysis.. Therapeutic H4R antagonism inhibited T cell infiltration in to the lung and decreased Th2 cytokines IL-13 and IL-5. IL-13 dependent remodeling parameters such as GCH and lung collagen were reduced. Intervention with H4R antagonist also improved measures of central and peripheral airway dysfunction.. These data demonstrate that therapeutic H4R antagonism can significantly ameliorate allergen induced, Th2 cytokine driven pathologies such as lung remodeling and airway dysfunction. The ability of H4R antagonists to affect these key manifestations of asthma suggests their potential as novel human therapeutics.

Topics: Airway Remodeling; Animals; Anti-Inflammatory Agents; Antibodies; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Collagen; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Goblet Cells; Histamine Antagonists; Hyperplasia; Indoles; Inflammation Mediators; Interleukin-13; Interleukin-5; Mice; Mice, Inbred BALB C; Ovalbumin; Piperazines; Receptors, G-Protein-Coupled; Receptors, Histamine; Receptors, Histamine H4; Th2 Cells

Montelukast and S-carbocysteine have been used in asthmatic patients as an anti-inflammatory or mucolytic agent respectively. S-carbocysteine also exhibits anti-inflammatory properties.. Ovalbumin (OVA) sensitized BALB/c mice were challenged with OVA for 3 days followed by single OVA re-challenge (secondary challenge) 2 weeks later. Forty-eight hours after secondary challenge, mice were assessed for airway hyperresponsiveness (AHR) and cell composition in bronchoalveolar lavage (BAL) fluid. Suboptimal doses of 10 mg.kg(-1) of S-carbocysteine by intraperitoneal injection (ip), 20 mg.kg(-1) of montelukast by gavage, the combination of S-carbocysteine and montelukast or 3 mg.kg(-1) of dexamethasone as a control were administered from 1 day before the secondary challenge to the last experimental day. Isolated lung cells were cultured with OVA and montelukast to determine the effects on cytokine production.. Treatment with S-carbocysteine or montelukast reduced both AHR and the numbers of eosinophils in BAL fluid. Neutralizing IFN-gamma abolished the effects of S-carbocysteine on these airway responses. Combination of the two drugs showed further decreases in both AHR and eosinophils in the BAL fluid. Goblet cell metaplasia and Th2-type cytokines, interleukin (IL)-4, IL-5 and IL-13, in BAL fluid were decreased with montelukast treatment. Conversely, S-carbocysteine increased Th1-type cytokines, IFN-gamma and IL-12 in BAL fluid.. The combination of two agents, montelukast and S-carbocysteine, demonstrated additive effects on AHR and airway inflammation in a secondary allergen model most likely through independent mechanisms of action.

Topics: Acetates; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carbocysteine; Cyclopropanes; Dexamethasone; Drug Synergism; Drug Therapy, Combination; Eosinophils; Female; Inflammation; Injections, Intraperitoneal; Mice; Mice, Inbred BALB C; Ovalbumin; Quinolines; Sulfides

CD27 is a costimulatory molecule of the TNFR family strongly expressed on activated CD4(+) and CD8(+) T lymphocytes. Binding with its ligand CD70, present on lymphocytes and DCs, leads to enhanced T cell activation and proliferation. Several other costimulatory molecules of the TNFR family like CD30, CD134 (OX40) or CD137 (4-1BB) have been shown to be critically involved in the development of asthma and/or respiratory tolerance. However, the role of CD27/CD70 signalling in these disease models has not been studied intensively. The aim of this study was to directly investigate the role of CD27 for the development of asthma and respiratory tolerance by comparative analysis of wild type (WT) and CD27(-/-) mice in the corresponding murine models. Ovalbumin (OVA)-sensitized and challenged CD27(-/-) mice developed comparably increased airway hyperreactivity (AHR), eosinophilic airway inflammation, mucus hypersecretion and elevated OVA-specific serum IgE levels in response to OVA sensitization as WT mice. In addition, Th2 cytokine production in spleen cell culture supernatants and proliferation of splenocytes after in vitro OVA restimulation was equally enhanced when derived from WT and CD27(-/-) mice. Furthermore, the absence of CD27 had no decisive impact on tolerance induction, so that WT and CD27(-/-) mice were comparably protected from asthma development by mucosal antigen application before sensitization. Our results suggest that CD27 costimulation is dispensable for a Th2 cell mediated allergic asthma response and respiratory tolerance induction in murine models.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Dendritic Cells; Disease Models, Animal; Humans; Immune Tolerance; Immunization; Immunoglobulin E; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Pulmonary Eosinophilia; T-Lymphocytes; Tumor Necrosis Factor Receptor Superfamily, Member 7

We evaluated if repeated stress modulates mucociliary clearance and inflammatory responses in airways of guinea pigs (GP) with chronic inflammation. The GP received seven exposures of ovalbumin or saline 0.9%. After 4th inhalation, animals were submitted to repeated forced swim stressor protocol (5x/week/2 weeks). After 7th inhalation, GP were anesthetized. We measured transepithelial potential difference, ciliary beat frequency, mucociliary transport, contact angle, cough transportability and serum cortisol levels. Lungs and adrenals were removed, weighed and analyzed by morphometry. Ovalbumin-exposed animals submitted to repeated stress had a reduction in mucociliary transport, and an increase on serum cortisol, adrenals weight, mucus wettability and adhesivity, positive acid mucus area and IL-4 positive cells in airway compared to non-stressed ovalbumin-exposed animals (p<0.05). There were no effects on eosinophilic recruitment and IL-13 positive cells. Repeated stress reduces mucociliary clearance due to mucus rheological-property alterations, increasing acid mucus and its wettability and adhesivity. These effects seem to be associated with IL-4 activation.

Topics: Adrenal Glands; Animals; Bronchial Hyperreactivity; Cilia; Disease Models, Animal; Epithalamus; Guinea Pigs; Hydrocortisone; Inflammation; Interleukin-3; Interleukin-4; Male; Mucociliary Clearance; Mucus; Organ Size; Ovalbumin; Stress, Psychological; Swimming

There is evidence that sex and sex hormones influence the severity of asthma. Airway and lung parenchyma remodeling and the relationship of ultrastructural changes to airway responsiveness and inflammation in male, female, and oophorectomized mice (OVX) were analyzed in experimental chronic allergic asthma. Seventy-two BALB/c mice were randomly divided into three groups (n=24/each): male, female, and OVX mice, whose ovaries were removed 7 days before the start of sensitization. Each group was further randomized to be sensitized and challenged with ovalbumin (OVA) or saline. Twenty-four hours after the last challenge, collagen fiber content in airways and lung parenchyma, the volume proportion of smooth muscle-specific actin in alveolar ducts and terminal bronchiole, the amount of matrix metalloproteinase (MMP)-2 and MMP-9, and the number of eosinophils and interleukin (IL)-4, IL-5, and transforming growth factor (TGF)-β levels in bronchoalveolar lavage fluid were higher in female than male OVA mice. The response of OVX mice was similar to that of males, except that IL-5 remained higher. Nevertheless, after OVA provocation, airway responsiveness to methacholine was higher in males compared with females and OVX mice. In conclusion, sex influenced the remodeling process, but the mechanisms responsible for airway hyperresponsiveness seemed to differ from those related to remodeling.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchodilator Agents; Chronic Disease; Collagen; Disease Models, Animal; Dose-Response Relationship, Drug; Extracellular Matrix; Female; Inflammation Mediators; Interleukin-4; Interleukin-5; Lung; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Ovariectomy; Pneumonia; Sex Factors; Time Factors; Transforming Growth Factor beta

Human rhinovirus is responsible for the majority of virus-induced asthma exacerbations. To determine the immunologic mechanisms underlying rhinovirus (RV)-induced asthma exacerbations, we combined mouse models of allergic airways disease and human rhinovirus infection. We inoculated OVA-sensitized and challenged BALB/c mice with rhinovirus serotype 1B, a minor group strain capable of infecting mouse cells. Compared with sham-infected, OVA-treated mice, virus-infected mice showed increased lung infiltration with neutrophils, eosinophils and macrophages, airway cholinergic hyperresponsiveness, and increased lung expression of cytokines including eotaxin-1/CCL11, IL-4, IL-13, and IFN-gamma. Administration of anti-eotaxin-1 attenuated rhinovirus-induced airway eosinophilia and responsiveness. Immunohistochemical analysis showed eotaxin-1 in the lung macrophages of virus-infected, OVA-treated mice, and confocal fluorescence microscopy revealed colocalization of rhinovirus, eotaxin-1, and IL-4 in CD68-positive cells. RV inoculation of lung macrophages from OVA-treated, but not PBS-treated, mice induced expression of eotaxin-1, IL-4, and IL-13 ex vivo. Macrophages from OVA-treated mice showed increased expression of arginase-1, Ym-1, Mgl-2, and IL-10, indicating a shift in macrophage activation status. Depletion of macrophages from OVA-sensitized and -challenged mice reduced eosinophilic inflammation and airways responsiveness following RV infection. We conclude that augmented airway eosinophilic inflammation and hyperresponsiveness in RV-infected mice with allergic airways disease is directed in part by eotaxin-1. Airway macrophages from mice with allergic airways disease demonstrate a change in activation state characterized in part by altered eotaxin and IL-4 production in response to RV infection. These data provide a new paradigm to explain RV-induced asthma exacerbations.

Topics: Animals; Antigens, CD; Antigens, Differentiation, Myelomonocytic; Bronchial Hyperreactivity; Chemokine CCL11; Cytokines; Eosinophils; HeLa Cells; Humans; Immunohistochemistry; Inflammation; Inflammation Mediators; Lung; Macrophages, Alveolar; Mice; Mice, Inbred BALB C; Microscopy, Confocal; Ovalbumin; Picornaviridae Infections; Reverse Transcriptase Polymerase Chain Reaction; Rhinovirus

The cause-and-effect relationship between airway smooth muscle (ASM) remodeling and airway hyperresponsiveness (AHR) following allergen challenge is not well established. Using a rat model of allergen-induced ASM remodeling we explored the relationship between the site of ASM remodeling and AHR. Brown Norway rats, sensitized and challenged (3 times at 5-day intervals) with ovalbumin, were intranasally administered 0.1 mg/kg budesonide 24 and 1 h before challenge. Airway responses to aerosolized methacholine were assessed 48 h or 1 wk after three challenges. Airways were stained and analyzed for total airway wall area, area of smooth muscle-specific α-actin, and goblet cell hyperplasia, and the constant-phase model was used to resolve the changes in respiratory system mechanics into large airway and peripheral lung responses. After three ovalbumin challenges, there was a significant increase in ASM area and in the total wall area in all sized airways as well as an increase in goblet cells in the central airways. Budesonide inhibited ASM growth and central airway goblet cell hyperplasia following ovalbumin challenges. Budesonide also inhibited small but not large airway total wall area. AHR was attributable to excessive responses of the small airways, whereas responsiveness of the large airways was unchanged. Budesonide did not inhibit AHR after repeated challenge. We conclude that ASM remodeling induced by repeated allergen challenges involves the entire bronchial tree, whereas AHR reflects alterations in the lung periphery. Prevention of ASM remodeling by corticosteroid does not abrogate AHR.

Topics: Airway Remodeling; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchodilator Agents; Budesonide; Cell Proliferation; Chemokines; Cytokines; Disease Models, Animal; Goblet Cells; Hyperplasia; Inflammation Mediators; Lung; Male; Muscle, Smooth; Ovalbumin; Rats; Rats, Inbred BN; Time Factors

This study investigated the effects of mycelial extract and culture filtrate of Cordyceps sphecocephala J201 on airway hyper-responsiveness, pulmonary immune cell infiltration, and Th2 cytokine expression in animal models of asthma. After Concanavalin A (Con A) activation of mouse primary spleen cells, the IL-4 and IL-13 cytokine expression were significantly decreased in the presence of the mycelial extract and culture filtrate of Cordyceps sphecocephala J201. The asthma model was induced by sensitization to ovalbumin by intraperitoneal (i.p.) injection treatment in mice. The Cordyceps sphecocephala J201 mycelial extract was injected in order to assess the effects of anti-asthmatic activity by comparing lung cell infiltration in ovalbumin-induced asthmatic mice. The results revealed that the increased IL-4, IL-13 and IL-25 expression were controlled by the mycelial extract and culture filtrate of Cordyceps sphecocephala J201, indicating that the extracts reduced the undesirable immune responses and/or cytokine expression exhibited in asthma.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Cell Line; Cordyceps; Cytokines; Disease Models, Animal; Lung; Male; Medicine, East Asian Traditional; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mycelium; Ovalbumin; Spleen

Resolvin E1 (RvE1) is an anti-inflammatory lipid mediator derived from the omega-3 fatty acid eicosapentaenoic acid (EPA), and strongly acts in the resolution of inflammation. We previously reported that RvE1 dampens airway inflammation and hyperresponsiveness in a murine model of asthma. In the present study, to elucidate the effects of RvE1 on the development of asthmatic airway inflammation, we investigated whether RvE1 acts on different phases of an OVA-sensitized and -challenged mouse model of asthma. RvE1 treatments at the time of either OVA sensitization or at the time of OVA challenge were investigated and compared with RvE1 treatments at the time of both OVA sensitization and challenge. After RvE1 was administered to mice intraperitoneally at the time of both OVA sensitization and challenge, there were decreases in airway eosinophil and lymphocyte recruitment, as well as a reduction in Th2 cytokine and airway hyperresponsiveness. RvE1 treatment at the time of either OVA sensitization or challenge also improved AHR and airway inflammation. Our results suggest that RvE1 acts on several phases of asthmatic inflammation and may have anti-inflammatory effects on various cell types.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eicosapentaenoic Acid; Female; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia

The mechanisms underlying the exaggerated distal airway inflammation and hyperresponsiveness that characterize acute exacerbations of asthma are largely unknown. Using BALB/c mouse experimental models, we demonstrated a potentially important role for alveolar macrophages (AM) in the development of an allergen-induced exacerbation of asthma. To induce features of airway inflammation and remodeling characteristic of mild chronic asthma, animals were systemically sensitized and exposed to low mass concentrations (≈3 mg/m(3)) of aerosolized ovalbumin for 30 minutes per day, 3 days per week, for 4 weeks. A subsequent single moderate-level challenge (≈30 mg/m(3)) was used to trigger an acute exacerbation. In chronically challenged animals, cytokine expression by AM was not increased, whereas after an acute exacerbation, AM exhibited significantly enhanced expression of proinflammatory cytokines, including interleukin (IL) 1β, IL-6, CXCL-1, and tumor necrosis factor α. In parallel, there was a marked increase in the expression of several cytokines by CD4(+) T-lymphocytes, notably the Th2 cytokines IL-4 and IL-13. Importantly, AM from an acute exacerbation stimulated the expression of Th2 cytokines when cocultured with CD4(+) cells from chronically challenged animals, and their ability to do so was significantly greater than AM from either chronically challenged or naïve controls. Stimulation was partly dependent on interactions involving CD80/86. We conclude that in an acute exacerbation of asthma, enhanced cytokine expression by AM may play a critical role in triggering increased expression of cytokines by pulmonary CD4(+) T-lymphocytes.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Chronic Disease; Cytokines; Enzyme-Linked Immunosorbent Assay; Female; Forced Expiratory Volume; Inflammation; Lung; Macrophages, Alveolar; Mice; Mice, Inbred BALB C; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Th2 Cells

We have recently reported that disruption of nuclear erythroid 2 P45-related factor 2 (Nrf2) enhances susceptibility to airway inflammatory responses induced by low-dose diesel exhaust particles (DEP) in mice. C57BL/6 Nrf2 knockout (Nrf2(-/-)) mice and wild-type (Nrf2(+/+)) mice were further exposed to low-dose DEP for 7h/day, 5 days/week, for a maximum of 8 weeks. After exposure to DEP for 5 weeks, allergic airway inflammation was generated in the mice by intraperitoneal sensitization with OVA followed by intranasal challenge. Nrf2(-/-) mice exposed to relatively low-dose DEP showed significantly increased percentage changes relative to the OVA alone group in terms of airway hyperresponsiveness (AHR) and inflammatory cells, levels of IL-5 and thymus and activation regulated chemokine (TARC) in bronchoalveolar lavage (BAL) fluid than did Nrf2(+/+) mice. Lung tissues of Nrf2(-/-) mice after DEP exposure showed inflammatory cell infiltrates, and increased PAS staining-positive mucus cell hyperplasia. In contrast, the percentage changes relative to the OVA group in the reduced glutathione (GSH)/oxidized glutathione (GSSG) ratio in whole blood was higher in Nrf2(+/+) mice than in Nrf2(-/-) mice. By using Nrf2(-/-) mice, it was shown for the first time that relatively low-dose DEP exposure induces oxidant stress, and that host anti-oxidant responses play a key role in the development of DEP-induced exacerbation of allergic airway inflammation.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Chemokines; Cytokines; Glutathione; Glutathione Disulfide; Immunoglobulin E; Immunoglobulin G; Inflammation; Leukocytes; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; NF-E2-Related Factor 2; Ovalbumin; Respiratory Hypersensitivity; Vehicle Emissions

Immunoglobulins (Igs) play important immunomodulatory effects on allergic asthma. Among these, IgG has been reported to regulate allergic inflammation in previous studies about immunotherapy and intravenous immunoglobulin therapy. In this study, to examine the immunomodulatory mechanisms of IgG and FcRs we evaluated the effects of intravenous (i.v.) rabbit IgG administration (IVIgG) on allergic airway inflammation and lung antigen-presenting cells (APCs) in a murine model of ovalbumin (OVA) sensitization and challenge. In OVA-challenged mice, IVIgG attenuated airway eosinophilia, airway hyperresponsiveness and goblet cell hyperplasia and also inhibited the local T helper type (Th) 2 cytokine levels. Additionally, IVIgG attenuated the proliferation of OVA-specific CD4(+) T cells transplanted into OVA-challenged mice. Ex vivo co-culture with OVA-specific CD4(+) cells and lung CD11c(+) APCs from mice with IVIgG revealed the attenuated transcription level of Th2 cytokines, suggesting an inhibitory effect of IVIgG on CD11c(+) APCs to induce Th2 response. Next, to analyse the effects on Fcγ receptor IIb and dendritic cells (DCs), asthmatic features in Fcγ receptor IIb-deficient mice were analysed. IVIgG failed to attenuate airway eosinophilia, airway inflammation and goblet cell hyperplasia. However, the lacking effects of IVIgG on airway eosinophilia in Fcγ receptor IIb deficiency were restored by i.v. transplantation of wild-type bone marrow-derived CD11c(+) DCs. These results demonstrate that IVIgG attenuates asthmatic features and the function of lung CD11c(+) DCs via Fcγ receptor IIb in allergic airway inflammation. Targeting Fc portions of IgG and Fcγ receptor IIb on CD11c(+) DCs in allergic asthma is a promising therapeutic strategy.

Topics: Adoptive Transfer; Animals; Antigen Presentation; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD11c Antigen; CD4-Positive T-Lymphocytes; Cell Count; Cell Differentiation; Cell Proliferation; Cytokines; Dendritic Cells; Disease Models, Animal; Eosinophils; Female; Immunoglobulin E; Immunoglobulin G; Immunoglobulins, Intravenous; Immunologic Factors; Inflammation; Mice; Mice, Inbred C57BL; Ovalbumin; Receptors, IgG; Specific Pathogen-Free Organisms; Th2 Cells

Inhaled bradykinin causes bronchoconstriction in asthmatic subjects but not nonasthmatics. To date, animal studies with inhaled bradykinin have been performed only in anesthetized guinea pigs and rats, where it causes bronchoconstriction through sensory nerve pathways. In the present study, airway function was recorded in conscious guinea pigs by whole-body plethysmography. Inhaled bradykinin (1 mM, 20 s) caused bronchoconstriction and influx of inflammatory cells to the lungs, but only when the enzymatic breakdown of bradykinin by angiotensin-converting enzyme and neutral endopeptidase was inhibited by captopril (1 mg/kg i.p.) and phosphoramidon (10 mM, 20-min inhalation), respectively. The bronchoconstriction and cell influx were antagonized by the B(2) kinin receptor antagonist 4-(S)-amino-5-(4-{4-[2,4-dichloro-3-(2,4-dimethyl-8-quinolyloxymethyl)phenylsulfonamido]-tetrahydro-2H-4-pyranylcarbonyl}piperazino)-5-oxopentyl](trimethyl)ammonium chloride hydrochloride (MEN16132) when given by inhalation (1 and 10 μM, 20 min) and are therefore mediated via B(2) kinin receptors. However, neither intraperitioneal MEN16132 nor the peptide B(2) antagonist icatibant, by inhalation, antagonized these bradykinin responses. Sensitization of guinea pigs with ovalbumin was not sufficient to induce airway hyperreactivity (AHR) to the bronchoconstriction by inhaled bradykinin. However, ovalbumin challenge of sensitized guinea pigs caused AHR to bradykinin and histamine. Infection of guinea pigs by nasal instillation of parainfluenza-3 virus produced AHR to inhaled histamine and lung influx of inflammatory cells. These responses were attenuated by the bradykinin B(2) receptor antagonist MEN16132 and H-(4-chloro)DPhe-2'(1-naphthylalanine)-(3-aminopropyl)guanidine (VA999024), an inhibitor of tissue kallikrein, the enzyme responsible for lung synthesis of bradykinin. These results suggest that bradykinin is involved in virus-induced inflammatory cell influx and AHR.

Topics: Animals; Bradykinin; Bradykinin B2 Receptor Antagonists; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Captopril; Cell Count; Glycopeptides; Guinea Pigs; Histamine; Male; Ornithine; Ovalbumin; Parainfluenza Virus 3, Human; Peptides; Plethysmography, Whole Body; Pneumonia; Protease Inhibitors; Respirovirus Infections; Sulfonamides; Tissue Kallikreins

The complement fragment C5a plays dual roles in the development of experimental allergic asthma. It protects from pulmonary allergy by a regulatory effect on dendritic cells during allergen sensitization, but is proallergic during the effector phase. C5a can bind to two distinct receptors (i.e., C5a receptor and C5a receptor-like 2 [C5L2]). The functional role of C5L2 in vivo remains enigmatic. In this study, we show in two models of OVA- and house dust mite (HDM)-induced experimental allergic asthma that C5L2-deficient mice are protected from the development of airway hyperresponsiveness, Th2 cytokine production, eosinophilic airway inflammation, serum IgE, or mucus production. Surprisingly, HDM-induced experimental asthma in C5L2-deficient mice was associated with increased pulmonary IL-17A production and increased airway neutrophil numbers. To directly assess the role of C5L2 on myeloid dendritic cells (mDCs) during allergen sensitization, we performed single or repeated adoptive transfers of C5L2-deficient mDCs into wild-type mice. HDM-pulsed C5L2-deficient mDCs induced strong Th2 cytokine production, which was associated with marked IFN-γ and IL-17A production, decreased eosinophil numbers, and reduced IgE production as compared with HDM-pulsed mDCs from wild-type mice. HDM stimulation of C5L2(-/-) mDCs in vitro resulted in production of Th17-promoting cytokine IL-23, which was absent in wild-type mDCs. Our findings suggest that C5L2 acts at the mDC/T cell interface to control the development of Th1 and Th17 cells in response to airway HDM exposure. Furthermore, it drives Th2 immune responses independent of mDCs, suggesting a complex role for C5L2 in the development of experimental allergic asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cell Communication; Cell Differentiation; Dendritic Cells; Disease Models, Animal; Dust; Inflammation Mediators; Interleukin-17; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Pyroglyphidae; Receptor, Anaphylatoxin C5a; Receptors, Chemokine; Th1 Cells

In most experimental model of asthma using guinea pigs, the animals are made to inhale an aerosolized antigen which passes through the nasal cavity. In the present study, we attempted to create an animal model of asthma showing a biphasic asthmatic response and airway hyperresponsiveness, in which the allergic responses are restricted to the lung. Guinea pigs were sensitized by the intratracheal instillation of ovalbumin (OVA)+Al(OH)₃ once a day for 7 d, and then intratracheally challenged with OVA 12 d after the last sensitization. The change in specific airway resistance (sRaw) and airway responsiveness to histamine were measured. Pranlukast (100 mg/kg), theophylline (50 mg/kg), and dexamethasone (10 mg/kg) were orally administered 18 and 2 h before the antigen challenge. The challenge caused a marked biphasic elevation of sRaw with peaks at 5 min and 4 h. At 24 h, airway hyperresponsiveness to histamine was observed. Pranlukast, theophylline, and dexamethasone suppressed the late asthmatic response and airway hyperresponsiveness. The early asthmatic response was inhibited by theophylline and dexamethasone. In conclusion, the intratracheal sensitization and challenge caused a biphasic asthmatic response and airway hyperresponsiveness in guinea pigs. This model may be useful for the evaluation of anti-asthma drugs.

Topics: Airway Resistance; Animals; Anti-Asthmatic Agents; Antigens; Asthma; Bronchial Hyperreactivity; Chromones; Dexamethasone; Disease Models, Animal; Guinea Pigs; Histamine; Lung; Male; Ovalbumin; Theophylline; Trachea

To explore the role of protein kinase C (PKC) in the augmented bronchial smooth muscle (BSM) contraction observed in the antigen-induced airway hyperresponsive (AHR) mice, the effects of a PKC activator, phorbol 12,13-dibutylate (PDBu), on BSM contraction were compared between the AHR and control mice. Actively sensitized mice were repeatedly challenged by antigen inhalation. Twenty-four hours after the final antigen challenge the isometrical contractions of the BSMs were measured. The BSM contraction induced by acetylcholine, but not high K(+) depolarization, was significantly augmented in the AHR mice. In BSMs of control mice, PDBu caused a significant increase in tension when the tissues were precontracted with high K(+), although PDBu itself had no effect on basal tone. The PDBu-mediated contraction was markedly augmented in BSMs of the AHR mice. These findings suggest that an increase in the PKC-mediated signaling is involved in the augmented contraction of BSMs in the antigen-induced AHR mice.

Topics: Acetylcholine; Animals; Bronchi; Bronchial Hyperreactivity; Carcinogens; Male; Mice; Mice, Inbred BALB C; Muscle Contraction; Muscle, Smooth; Ovalbumin; Phorbol 12,13-Dibutyrate; Protein Kinase C

Previous studies have suggested that the asthmatic responses of airway inflammation, remodeling, and hyperresponsiveness (AHR) are interrelated; in this study, we used exercise to examine the nature of this interrelationship. Mice were sensitized and challenged with ovalbumin (OVA); mice were then exercised via running on a motorized treadmill at a moderate intensity. Data indicate that, within the lungs of OVA-treated mice, exercise attenuated the production of inflammatory mediators, including chemokines KC, RANTES, and MCP-1 and IL-12p40/p80. Coordinately, OVA-treated and exercised mice displayed decreases in leukocyte infiltration, including eosinophils, as compared with sedentary controls. Results also show that a single bout of exercise significantly decreased phosphorylation of the NFkappaB p65 subunit, which regulates the gene expression of a wide variety of inflammatory mediators. In addition, OVA-treated and exercised mice exhibited decreases in the levels of Th2-derived cytokines IL-5 and IL-13 and the prostaglandin PGE(2), as compared with sedentary controls. In contrast, results show that a single bout of exercise had no effect on AHR in OVA-treated mice challenged with increasing doses of aerosolized methacholine (0-50 mg/ml) as compared with sedentary mice. Exercise also had no effect on epithelial cell hypertrophy, mucus production, or airway wall thickening in OVA-treated mice as compared with sedentary controls. These findings suggest that a single bout of aerobic exercise at a moderate intensity attenuates airway inflammation but not AHR or airway remodeling in OVA-treated mice. The implication of these findings for the interrelationship between airway inflammation, airway remodeling, and AHR is discussed.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Chemokines; Chemotactic Factors; Cytokines; Dinoprostone; Disease Models, Animal; Female; Humans; Inflammation; Leukocytes; Mice; Mice, Inbred BALB C; Ovalbumin; Physical Conditioning, Animal; Random Allocation; Respiratory Hypersensitivity; Th2 Cells; Transcription Factor RelA

The impact of cigarette smoke on allergic asthma remains controversial both clinically and experimentally. The objective of this study was to investigate, in a murine model, how cigarette smoke affects immune inflammatory processes elicited by a surrogate allergen. In our experimental design, mice were concurrently exposed to cigarette smoke and ovalbumin (OVA), an innocuous antigen that, unless introduced in the context of an adjuvant, induces inhalation tolerance. We show that cigarette smoke exposure has adjuvant properties, allowing for allergic mucosal sensitization to OVA. Specifically, concurrent exposure to cigarette smoke and OVA for 2 weeks led to airway eosinophilia and goblet cell hyperplasia. In vivo OVA recall challenge 1 month after the last smoke exposure showed that concurrent exposure to OVA and cigarette smoke induced antigen-specific memory. Robust eosinophilia and OVA-specific IgG1 and IgE characterized the ensuing inflammatory response. Mechanistically, allergic sensitization was, in part, granulocyte macrophage colony-stimulating factor (GM-CSF) dependent, as a significant reduction in BAL eosinophilia was observed in mice treated with an anti-GM-CSF antibody. Of note, continuous smoke exposure attenuated the OVA recall response; decreased airway eosinophilia was observed in mice continuously exposed to cigarette smoke compared with mice that ceased the smoke exposure protocol. In conclusion, we demonstrate experimentally that while cigarette smoke acts as an adjuvant allowing for allergic sensitization, it also attenuates the ensuing eosinophilic inflammatory response.

Topics: Adjuvants, Immunologic; Allergens; Animals; Antigens, CD; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dendritic Cells; Disease Models, Animal; Female; Humans; Immune Tolerance; Immunologic Memory; Inflammation; Mice; Mice, Inbred BALB C; Nicotiana; Ovalbumin; Smoke; T-Lymphocytes

Stat3, Socs3 and cytokines play an integral role in the coordination and persistence of inflammation. However, a clear understanding of the role played by the Stat3/IL-6 and Socs3 pathway in airway inflammation is lacking. We report the alteration in the status of expression and activation of Stat3 by ovalbumin (OVA), and establish its relationship with Socs3 and IL-6 in the lungs of mice with eosinophilic pulmonary inflammation and airway hyperresponsiveness.. Alterations in the expression of Stat3, Socs3 and IL-6 were determined in a murine model of asthma, where Balb/c mice were sensitized and challenged with OVA (OVA/OVA) and compared with control mice sensitized and challenged with saline (SAL) (SAL/SAL) mice. The OVA/OVA mice were characterized by a moderate increase in methacholine-induced specific airway resistance, the presence of 150 microg/ml of OVA-specific IgG and 8.93 microg/ml OVA-specific IgE antibody and elevated levels of eosinophils and Th2 cytokines (IL-4 and IL-5) in the bronchoalveolar lavage fluid. In contrast SAL/SAL mice had low eosinophils, IL-4 and IL-5 and no OVA-specific IgG and IgE antibodies in the BALF. Stat3 and Socs3 expression profiles were monitored in OVA/OVA and Stat3- and Socs3-silenced OVA/OVA mice. Furthermore, expression of IL-6 in Stat3- and Socs3-silenced mice and the exogenous effect of IL-6 on Stat3 were studied.. The results show that expression and activation of Stat3 mRNA and proteins are significantly low in lung of OVA/OVA mice in comparison to SAL/SAL mice following OVA challenge. An increased pool of Socs3 mRNA is observed in OVA/OVA mice with or without OVA challenge and in SAL/SAL mice 24 h after OVA challenge. Transient in vivo blocking of Socs3 gene by Socs3 siRNA restores the expression of IL-6 mRNA and protein in OVA/OVA mice, and nasal administration of recombinant IL-6 to OVA/OVA mice enhanced Stat3 mRNA expression.. Our data suggest that airway inflammation is associated with low expression of Stat3 and IL-6 and overexpression of Socs3 genes in a mouse model of asthma. Furthermore, IL-6 is under the influence of the Socs3 gene and may contribute to the negative regulation of Stat3 via IL-6 following a challenge with an allergen during the development of asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophils; Female; Gene Expression Regulation; Humans; Inflammation; Interleukin-6; Mice; Mice, Inbred BALB C; Ovalbumin; STAT3 Transcription Factor; Suppressor of Cytokine Signaling 3 Protein; Suppressor of Cytokine Signaling Proteins

The surfactant proteins SP-A and SP-D, components of the innate immune system, are involved in host defence.. We tested the hypothesis that ovalbumin (OVA) challenge leads to an upregulation of both proteins in alveolar epithelial type II cells (AEII) and Clara cells and to an enhanced uptake by macrophages.. After sensitization with OVA and heat-killed Bordetella pertussis challenge followed intratracheally with 0.5% OVA on day 13. One day after challenge lung tissue and bronchoalveolar lavage fluid (BALF) of sensitized NaCl- and OVA-challenged Brown Norway rats were compared with home cage controls using qRt-PCR, Western blot and immunohistochemistry.. After OVA challenge (1) eosinophils increased significantly in the BALF, (2) the total amount of SP-A and SP-D was significantly increased in lung tissue, (3) the amount of SP-A was significantly and the amount of SP-D was remarkably elevated in BALF, and (4) the levels of SP-A and SP-D mRNA in lung tissue were significantly elevated. Using quantitative immunohistochemistry, we found (5) significantly higher surface fractions of SP-A- and SP-D-labelled AEII, (6) no differences in the surface fractions of SP-A- and SP-D-labelled bronchial Clara cells, and (7) a significantly increased cell density of unlabelled and SP-A-labelled macrophages.. Thus, combining molecular biological and histological methods we suggest that after OVA challenge (1) AEII but not Clara cells show a significantly higher expression of SP-A and SP-D leading also to higher amounts of both SPs in BALF and (2) macrophages gather predominantly SP-A.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Inflammation; Lung; Male; Ovalbumin; Pulmonary Alveoli; Pulmonary Surfactant-Associated Protein A; Pulmonary Surfactant-Associated Protein D; Rats; Rats, Inbred BN; Respiratory Hypersensitivity; Up-Regulation

Arginase has been suggested to compete with nitric oxide synthase (NOS) for their common substrate, l-arginine. To study the mechanisms underlying this interaction, we compared arginase expression in isolated airways and the consequences of inhibiting arginase activity in vivo with NO production, lung inflammation, and lung function in both C57BL/6 and NOS2 knockout mice undergoing ovalbumin-induced airway inflammation, a mouse model of asthma. Arginases I and II were measured by western blot in isolated airways from sensitized C57BL/6 mice exposed to ovalbumin aerosol. Physiological and biochemical responses - inflammation, lung compliance, airway hyperreactivity, exhaled NO concentration, arginine concentration - were compared with the responses of NOS2 knockout mice. NOS2 knockout mice had increased total cells in lung lavage, decreased lung compliance, and increased airway hyperreactivity. Both arginase I and arginase II were constitutively expressed in the airways of normal C57BL/6 mice. Arginase I was up-regulated approximately 8-fold in the airways of C57BL/6 mice exposed to ovalbumin. Expression of both arginase isoforms were significantly upregulated in NOS2 knockout mice exposed to ovalbumin, with about 40- and 4-fold increases in arginases I and II, respectively. Arginine concentration in isolated airways was not significantly different in any of the groups studied. Inhibition of arginase by systemic treatment of C57BL/6 mice with a competitive inhibitor, Nomega-hydroxy-nor-l-arginine (nor-NOHA), significantly decreased the lung inflammatory response to ovalbumin in these animals. We conclude that NOS2 knockout mice are more sensitive to ovalbumin-induced airway inflammation and its sequelae than are C57BL/6 mice, as determined by increased total cells in lung lavage, decreased lung compliance, and increased airway hyperreactivity, and that these findings are strongly correlated with increased expression of both arginase isoforms in the airways of the NOS2 knockout mice exposed to ovalbumin.

Topics: Animals; Arginase; Arginine; Asthma; Breath Tests; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Enzyme Inhibitors; Lung; Lung Compliance; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide; Nitric Oxide Synthase Type II; Ovalbumin; Pneumonia; Up-Regulation

In this study, we have examined the role of phosphoinositide 3 kinase gamma (PI3Kgamma), a class Ib PI3K, in contributing to airway remodeling utilizing PI3Kgamma-deficient mice exposed to chronic allergen challenge. Wild-type (WT) mice sensitized to ovalbumin (OVA) and chronically challenged with OVA for 1 mo developed significantly increased levels of eosinophilic inflammation and airway remodeling. In contrast, PI3Kgamma-deficient mice challenged with OVA had significantly reduced numbers of bronchoalveolar lavage and peribronchial eosinophils compared with WT mice. There was no significant difference in the number of bone marrow or circulating peripheral blood eosinophils when comparing WT mice and PI3Kgamma-deficient mice, suggesting that trafficking of eosinophils into the lung was reduced in PI3Kgamma-deficient mice. PI3Kgamma-deficient and WT mice had similar levels of IL-5 and eotaxin-1. The reduced eosinophil recruitment to the airway in PI3Kgamma-deficient mice challenged with OVA was associated with significantly reduced numbers of TGF-beta1+ peribronchial cells, reduced numbers of pSmad 2/3+ airway epithelial cells, and pSmad 2/3+ peribronchial cells, as well as significantly reduced levels of peribronchial fibrosis (quantitated by trichrome staining and image analysis as well as by lung collagen levels). In addition, the area of peribronchial alpha-smooth muscle staining was significantly reduced in PI3Kgamma-deficient compared with WT mice. Overall, this study demonstrates an important role for PI3Kgamma in mediating allergen-induced eosinophilic airway inflammation and airway remodeling, suggesting that PI3Kgamma may be a novel therapeutic target in asthma.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Class Ib Phosphatidylinositol 3-Kinase; Eosinophils; Immunoblotting; Interleukin-5; Isoenzymes; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth; Ovalbumin; Phosphatidylinositol 3-Kinases; Pneumonia; Respiratory System; Smad2 Protein; Smad3 Protein; Transforming Growth Factor beta1

We previously reported a mouse model of bronchial asthma showing eosinophilic inflammation, but not airway hyperresponsiveness (AHR), after prolonged antigen exposure. This model showed an increase of IL-12 in the lung.. The aim of this study was to investigate the role of IL-12p40 in a murine asthma model with prolonged antigen exposures.. An ovalbumin (OVA)-induced asthma model was first established in wild-type (WT) and IL-12p40-deficient (IL-12p40(-/-)) mice. Both strains of mice were further exposed to either OVA (prolonged exposure group) or phosphate-buffered saline (positive control group) 3 days per week for 3 weeks. During week 4, both groups of mice were given a final challenge with OVA.. Prolonged antigen exposures resulted in marked suppression of airway eosinophilia in both WT and IL-12p40(-/-) mice. However, AHR persisted in IL-12p40(-/-) but not in WT mice. There were no significant differences of IL-5, IL-13 or IFN-gamma levels in bronchoalveolar lavage fluid between WT and IL-12p40(-/-) mice. The hydroxyproline content of the lung and peribronchial fibrosis were, however, significantly increased in IL-12p40(-/-) mice.. The results suggest that endogenous IL-12p40 is essential for inhibition of AHR and peribronchial fibrosis, but not eosinophilic inflammation, in a murine asthma model with prolonged antigen exposures.

Topics: Administration, Inhalation; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Down-Regulation; Eosinophils; Female; Immune Tolerance; Interleukin-12 Subunit p40; Leukocytes; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Respiratory Function Tests

Asthma is a disease characterized by variable and reversible airway obstruction and is associated with airway inflammation, airway remodelling (including goblet cell hyperplasia, increased collagen deposition and increased smooth muscle mass) and increased airway responsiveness. It is believed that airway inflammation plays a critical role in the development of airway remodelling, with IL-13 and TGF-beta1 pathways being strongly associated with the disease progression. Mouse models of asthma are capable of recapitulating some components of asthma and have been used to look at both IL-13 and TGF-beta1 pathways, which use STAT6 and SMAD2 signalling molecules, respectively.. Using brief and chronic models of allergen exposure, we utilized BALB/c and C57Bl/6 to explore the hypothesis that observed differences in responses to allergen between these mouse strains will involve fundamental differences in IL-13 and TGF-beta1 responses.. The following outcome measurements were performed: airway physiology, bronchoalveolar lavage cell counts/cytokine analysis, histology, immunoblots and gene expression assays.. We demonstrate in BALB/c mice an IL-13-dependent phosphorylation of STAT6, nuclear localized in inflammatory cells, which is associated with indices of airway remodelling and development of airway dysfunction. In BALB/c mice, phosphorylation of SMAD2 is delayed relative to STAT6 activation and also involves an IL-13-dependent mechanism. In contrast, despite an allergen-induced increase in IL-4, IL-13 and eosinophils, C57Bl/6 demonstrates a reduced and distinct pattern of phosphorylated STAT6, no SMAD2 phosphorylation changes and fail to develop indices of remodelling or changes in airway function.. The activation of signalling pathways and nuclear translocation of signalling molecules downstream of IL-13 and TGF-beta1 further support the central role of these molecules in the pathology and dysfunction in animal models of asthma. Activation of signalling pathways downstream from IL-13 and TGF-beta1 may be more relevant in disease progression than elevations in airway inflammation alone.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Humans; Interleukin-13; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Smad2 Protein; Species Specificity; STAT6 Transcription Factor; Transforming Growth Factor beta1

Asthma is a chronic inflammatory disease of the lung and its incidence has been increasing around the world. We previously reported that oral administration of a water-soluble extract prepared from Actinidia arguta, code-named PG102, could modulate the level of Th1 and Th2 cytokines and suppress the production of immunoglobulin E (IgE) in the ovalbumin (OVA)-immunized murine model as well as in the in vitro cell culture system, and furthermore could significantly improve dermatitis conditions in the NC/Nga murine model. These data suggested that PG102 might have therapeutic effects in a broad range of allergic diseases.. To assess the possible anti-allergic effects of PG102 in the OVA-induced murine asthma model.. The quality of PG102 was standardized, using its effects on the production of IgE, IL-5, and IL-13, in in vitro cell culture systems. To test effects on asthma, BALB/c mice were orally administrated with PG102, followed by OVA sensitization and challenge to induce asthmatic symptoms. Airway hyperresponsiveness (AHR), bronchoalveolar lavage fluid, serum, and lung tissue were analysed by using various methods.. PG102 could decrease the level of IgE, IL-5, and IL-13 in in vitro cell culture systems with IC(50) being 1.12-1.43 mg/mL. PG102 could ameliorate asthmatic symptoms, including AHR and eosinophilia in the lungs. Such improvement of asthmatic symptoms by PG102 was accompanied by the down-regulation of IL-5 and IgE. In PG102-treated mice, high level expression of heme oxygenase-1, a potent anti-inflammatory enzyme, was observed in alveolar inflammatory cells, while the mRNA levels of foxp3, TGF-beta1, and IL-10, important markers for regulatory T cells, were also up-regulated in the lung tissue.. PG102 may have potential as a safe and effective reagent for the prevention or treatment of asthma.

Topics: Actinidia; Animals; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophils; Female; Forkhead Transcription Factors; Fruit; Gene Expression; Heme Oxygenase-1; Humans; Immunoglobulin E; Interleukin-10; Lung; Mast Cells; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plant Extracts; Respiratory Function Tests; T-Lymphocytes; Transforming Growth Factor beta1

Asthma is a chronic respiratory disease, which needs a safer medication preferably in inhalation form. In view of this, we have evaluated the effect of inhaled carbenoxolone (CBX), a herbal-derived compound, on asthma in a mouse model.. Mice were sensitized and challenged with ovalbumin (OVA) to develop certain characteristic features of asthma such as airway hyperreactivity (AHR), airway eosinophilia, lung inflammation and mucus hypersecretion. To evaluate the effect of CBX on the above asthmatic features, CBX (2.5, 5 and 10 mg/ml, 3 ml) or vehicle (water) was given by inhalation. AHR was determined using whole-body plethysmography. Infiltration of eosinophils was estimated by microscopy. Lung inflammation and mucus hypersecretion were assessed using hematoxylin and eosin, and periodic acid-Schiff staining, respectively. Th-2 cytokines, IL-4 and IL-5 were measured in bronchoalveolar lavage (BAL) fluid and IgE in sera. To identify the possible mode of CBX action, we measured corticosterone levels in the BAL fluid and 5-lipoxygenase (5-LO) expression in the lungs.. CBX (5 mg/ml) inhalation markedly alleviated AHR (p = 0.0032) and reduced lung inflammation and mucus hypersecretion. Also, it prevented the increase in IL-4 (p = 0.0192), IL-5 (p = 0.0116) and eosinophils (p < 0.0005) in the BAL fluid, and OVA-specific IgE levels (p = 0.00061) in sera. 5-LO expression was also markedly reduced. However, corticosterone levels were not affected.. Inhaled CBX alleviates the asthmatic features in mice and could be a potent nebulized therapy in clinical asthma.

Topics: Administration, Inhalation; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carbenoxolone; Disease Models, Animal; Immunoglobulin E; Lung; Male; Mice; Mice, Inbred BALB C; Nebulizers and Vaporizers; Ovalbumin

The aim of the present work is focused on protective effects of an edible red alga, Laurencia undulata ethanolic (EtOH) extracts (LU) containing a large amount of polyphenols against OVA-induced murine allergic airway reactions using in vivo histological and cytokine assay. Mice sensitized and challenged with ovalbumin (OVA) showed typical asthmatic reactions as follows: an increase in the number of eosinophil in bronchoalveolar lavage fluid; a marked influx of inflammatory cells into the lung around blood vessels and airways, and airway luminal narrowing; the development of airway hyperresponsiveness; the detection of TNF-alpha and Th2 cytokines, such as IL-4 and IL-5 in the bronchoalveolar lavage (BAL) fluid; and detection of allergen-specific IgE in the serum. The successive intraperitoneal administration of LU before the last airway OVA-challenge resulted in a significant inhibition of all asthmatic reactions. These results suggest that L. undulata polyphenolic extracts possess therapeutic potential for combating bronchial asthma associated with allergic diseases.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Flavonoids; Laurencia; Lung; Mice; Mice, Inbred C57BL; Ovalbumin; Phenols; Plant Extracts; Polyphenols

The contribution of CD4 T cells and other CD4+ cells to lung inflammation and airway remodeling remains unclear during bouts of chronic exposure to airborne allergen. Previously, murine models have shown that CD4 T cells are required for initiation of acute inflammation and the remodeling process. However, it is unknown whether CD4 T cells or other CD4+ cells continue to be required for remodeling during ongoing allergen challenges after the development of acute eosinophilic lung inflammation. To test this, mice were sensitized and challenged with ovalbumin (OVA). After acute airway inflammation was established, a CD4 depleting antibody was administered for 4 wk during a period of chronic exposure to intranasal OVA, resulting in effective depletion of CD4+ cells from all organs, including the lung, lung-draining lymph nodes, and spleen. In these mice, levels of peribronchial inflammation, bronchoalveolar (BAL) eosinophils, and lung CD11c+, CD8+, and Siglec-F+CD11c- cells were significantly reduced. However, mucus metaplasia, peribronchial subepithelial fibrosis, and smooth muscle mass were not affected. Additionally, depletion of CD4+ cells before the last week of chronic allergen challenges also led to significant reductions in BAL eosinophils, peribronchial inflammation, and lung CD11c+, CD8+, and Siglec-F+CD11c- cells. These results show that CD4 T cells, and other CD4+ cells including subsets of dendritic cells, iNKT cells, and LTi cells, play a role in ongoing eosinophilic lung inflammation during periods of chronic allergen challenge, but are not required for progressive airway remodeling that develops after initial acute inflammation.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Chronic Disease; Class Ib Phosphatidylinositol 3-Kinase; Dendritic Cells; Flow Cytometry; Immunoblotting; Isoenzymes; Lung; Lymph Nodes; Mice; Mice, Inbred C57BL; Natural Killer T-Cells; Ovalbumin; Phosphatidylinositol 3-Kinases; Pulmonary Eosinophilia; Respiratory System

F(2)-isoprostanes have been associated with various forms of oxidant stress. The levels of F(2)-isoprostanes in a murine asthma model were studied both in situ and in vivo and further investigated whether the formation of F(2)-isoprostanes was associated with increased ovalbumin (OVA)-induced airway inflammation after a 17-day (OVA-17) or a 24-day (OVA-24) protocol. Bronchial reactivity was assessed by using a ventilator (FlexiVent). OVA-treated animals had higher lung resistance and lung compliance compared to control groups (P<0.001). 8-Iso-PGF(2)(alpha) levels in bronchoalveolar lavage (BAL) and 8-iso-PGF(2)(alpha) immunoreactivity in lung tissue were analyzed. OVA-17 mice showed a 2.5-fold increased level of 8-iso-PGF(2)(alpha) in BAL compared to PBS-17 mice (P=0.023). Lung tissue from OVA-24 mice had more intense 8-iso-PGF(2)(alpha) staining compared to OVA-17 mice. This study showed an accumulation of F(2)-isoprostanes in acute airway inflammation and a markedly increased tissue damage caused by oxidative stress in an ongoing inflammation.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; F2-Isoprostanes; Female; Humans; Inflammation; Mice; Mice, Inbred BALB C; Ovalbumin; Oxidative Stress

Increased tumour necrosis factor-alpha (TNF-alpha) is associated with airway hyperreactivity in antigen-challenged animals. In human asthmatics, TNF-alpha is increased and blocking it prevents airway hyperreactivity in some asthmatic patients. However, the mechanisms by which TNF-alpha mediates hyperreactivity are unknown. Airway hyperreactivity can be caused by dysfunction of neuronal M(2) muscarinic receptors that normally limit acetylcholine release from parasympathetic nerves. Here we test whether blocking TNF-alpha receptors with etanercept prevents M(2) receptor dysfunction and airway hyperreactivity in antigen-challenged guinea pigs.. Ovalbumin-sensitized guinea pigs were challenged by inhalation of antigen. Some animals received etanercept (3 mg kg(-1) i.p.) 3 h before challenge. 24 h after challenge, airway hyperreactivity and M(2) receptor function were tested. Inflammatory cells in bronchoalveolar lavage, blood and lung were counted. TNF-alpha and its receptors were detected by real-time RT-PCR and immunocytochemistry in parasympathetic nerves from humans and guinea pigs and in human neuroblastoma cells.. Antigen-challenged animals were hyperreactive to vagal stimulation and neuronal M(2) receptors were dysfunctional. Both M(2) receptor dysfunction and airway hyperreactivity were prevented by etanercept. Etanercept reduced eosinophils around airway nerves, and in blood, bronchoalveolar lavage and airway smooth muscle. Also, TNF-alpha decreased M(2) receptor mRNA in human and guinea pig parasympathetic neurons.. Tumour necrosis factor-alpha may contribute to M(2) receptor dysfunction and airway hyperreactivity directly by decreasing receptor expression and indirectly by promoting recruitment of eosinophils, containing major basic protein, an M(2) antagonist. This suggests that etanercept may be beneficial in treatment of allergic asthma.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cell Line, Tumor; Eosinophils; Etanercept; Guinea Pigs; Humans; Immunoglobulin G; Muscle, Smooth; Myocardium; Neurons; Ovalbumin; Parasympathetic Nervous System; Receptor, Muscarinic M2; Receptors, Tumor Necrosis Factor; Reverse Transcriptase Polymerase Chain Reaction; Trachea; Tumor Necrosis Factor-alpha

Dendritic cells (DCs) are considered to be the most efficient antigen-presenting cells. Intratracheal administration of allergen-pulsed bone marrow-derived dendritic cells (BMDCs) before allergen challenge induces airway hyperresponsiveness (AHR) and inflammation. Ovalbumin (OVA)-pulsed BMDCs from wild-type (WT) mice were transferred into naive WT, CD4(-/-), CD8(-/-), or IL-13(-/-) mice. Two days (short protocol) or 10 days (long protocol) after BMDC transfer, mice were challenged with 1% OVA for 3 days and assayed 2 days later. Transfer of OVA-primed BMDCs into BALB/c or C57BL/6 mice elicited AHR in both protocols. Airway eosinophilia, Th2 cytokines, or goblet cell metaplasia were increased in the long but not short protocol. Lung T cells from both protocols produced Th2 cytokines in response to OVA in vitro. Carboxyfluorescein diacetate succinimidylester-labeled BMDCs were observed in bronchoalveolar lavage (BAL) fluid and lung parenchyma at early time points, and were detected in draining lymph nodes 48 hours after transfer. CD8(-/-) mice developed AHR comparable to WT mice in the short protocol, but decreased levels of AHR, airway eosinophilia, Th2 cytokines in BAL fluid, and goblet cell metaplasia compared with WT mice in the long protocol. CD4(-/-) or IL-13(-/-) mice did not develop AHR or airway inflammation in either protocol. These data suggest that allergen-pulsed BMDCs initiate development of AHR that is dependent initially on CD4(+) T cells, and at later time periods on CD8+ T cells and IL-13. Thus, the timing of allergen challenge after transfer of allergen-pulsed BMDC affects the development of AHR and airway inflammation.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4 Antigens; CD4-Positive T-Lymphocytes; CD8 Antigens; CD8-Positive T-Lymphocytes; Cytokines; Dendritic Cells; Female; Goblet Cells; Inflammation; Interleukin-13; Lymphocyte Activation; Lymphocyte Subsets; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin

Endothelial lipase (EL) is a novel phospholipase that determines plasma high-density lipoprotein cholesterol (HDL-C) levels. We have investigated the role of HDL-C in lung allergic inflammation by using EL knockout (EL-KO) mice that are high in HDL-C. EL-KO and wild-type control mice were sensitized and challenged with ovalbumin to evoke eosinophilic inflammation in the lung. EL was expressed in epithelial cells, alveolar type II cells, and endothelial cells in the lung, and its expression was upregulated during inflammation. Concomitant with attenuated hyperresponsiveness of the airway smooth muscles, the number of eosinophils in bronchoalveolar lavage and the expression of VCAM-1 were lower in EL-KO mice than in control mice. HDL reduced cytokine-induced VCAM-1 expression in cultured endothelial cells. When plasma HDL levels were decreased to similar levels in both mouse groups by adenovirus-mediated overexpression of EL, however, eosinophil infiltration was still lower in EL-KO mice. In vitro adhesion assays revealed that EL expression on the cell surface promoted the interaction of eosinophils through the ligand-binding function of EL. In summary, targeted inactivation of EL attenuated allergic inflammation in the lung, and the protective effects in EL-KO mice were associated with high plasma HDL levels, downregulation of VCAM-1, and loss of the direct ligand-binding function of EL. Thus EL is a novel modulator of the progression of allergic asthma.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Adhesion; Cell Movement; Chlorocebus aethiops; Cholesterol, HDL; COS Cells; Endothelium; Eosinophils; Gene Targeting; Humans; Hypersensitivity; Lipase; Lung; Mice; Mice, Knockout; Ovalbumin; Pneumonia; Vascular Cell Adhesion Molecule-1

Although there have been numerous studies on the development of allergen-induced inflammation, the mechanisms leading to resolution of inflammation remain poorly understood. This represents an important consideration because failure to resolve allergen driven inflammation potentially leads to irreversible airway remodeling, characteristic of chronic asthma.. We investigated the resolution of allergic inflammation and identified the factors responsible.. BALB/c and C57BL/6 mice were sensitized to ovalbumin and challenged through the airways to induce allergic inflammation. Mice were analyzed at 24 hours and 7 days after the final challenge.. Airway hyperreactivity (AHR) and increased mucus production were present 7 days after the cessation of allergen challenge in BALB/c mice. Persisting AHR correlated with the continued presence of Th2 cells but not eosinophils in the lungs. The role of Th2 cells in maintaining AHR was confirmed using blocking antibodies against T1/ST2, IL-4, and IL-13 during the resolution period. Moreover, AHR in the "Th1 type" C57BL/6 mouse strain was resolved 1 week after allergen challenge, concomitant with clearance of Th2 cells from the lung. Expression of the T1/ST2 ligand, IL-33, also correlated with maintenance of AHR.. We have used blockade of Th2 function and strain differences to show for the first time that resolution of allergic inflammation and AHR may be dependent on the T1/ST2-IL-33 pathway and the presence of Th2 cells, suggesting they are necessary not only for the development of an allergic response but also for its maintenance.

Topics: Animals; Antibodies, Monoclonal; Bronchial Hyperreactivity; Chemokines; Eosinophils; Female; Immunoglobulin E; Inflammation; Interleukin-1 Receptor-Like 1 Protein; Interleukin-13; Interleukin-4; Leukocyte Count; Lung; Membrane Proteins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Receptors, Interleukin; Respiratory Hypersensitivity; Respiratory Mucosa; Th1 Cells; Th2 Cells

Interleukin-13 (IL-13) is one of the central mediators for development of airway hyperresponsiveness in asthma. The signal transducer and activation of transcription 6 (STAT6) is one of the major signal transducers activated by IL-13, and a possible involvement of IL-13/STAT6 pathway in the augmented bronchial smooth muscle (BSM) contraction has been suggested. In the present study, the effect of a novel STAT6 inhibitor, AS1517499, on the development of antigen-induced BSM hyperresponsiveness was investigated. In cultured human BSM cells, IL-13 (100 ng/ml) caused a phosphorylation of STAT6 and an up-regulation of RhoA, a monomeric GTPase responsible for Ca2+ sensitization of smooth muscle contraction: both events were inhibited by co-incubation with AS1517499 (100 nM). In BALB/c mice that were actively sensitized and repeatedly challenged with ovalbumin antigen, an increased IL-13 level in bronchoalveolar lavage fluids and a phosphorylation of STAT6 in bronchial tissues were observed after the last antigen challenge. These mice had an augmented BSM contractility to acetylcholine together with an up-regulation of RhoA in bronchial tissues. Intraperitoneal injections of AS1517499 (10 mg/kg) 1 hour before each ovalbumin exposure inhibited both the antigen-induced up-regulation of RhoA and BSM hyperresponsiveness, almost completely. A partial but significant inhibition of antigen-induced production of IL-13 was also found. These findings suggest that the inhibitory effects of STAT6 inhibitory agents, such as AS1517499, both on RhoA and IL-13 up-regulations might be useful for asthma treatment.

Topics: Animals; Anti-Asthmatic Agents; Antigens; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cells, Cultured; Disease Models, Animal; Dose-Response Relationship, Drug; Humans; Immunoglobulin E; Injections, Intraperitoneal; Interleukin-13; Male; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Phosphorylation; Pyrimidines; Recombinant Proteins; rho GTP-Binding Proteins; rhoA GTP-Binding Protein; STAT6 Transcription Factor; Time Factors

The interactions between airway responsiveness, structural remodelling and inflammation in allergic asthma remain poorly understood. Prolonged challenge with inhaled allergen is necessary to replicate many of the features of airway wall remodelling in mice. In both mice and humans, genetic differences can have a profound influence on allergy, inflammation, airway responsiveness and structural changes.. The aim of this study was to provide a comparative analysis of allergen-induced airway changes in sensitized BALB/c and C57BL/6 mice that were exposed to inhaled allergen for 2 ('acute'), 6 or 9 weeks ('chronic'). Inflammation, remodelling and responsiveness were analyzed.. Both strains developed a Th-2-driven airway inflammation with allergen-specific IgE, airway eosinophilia and goblet cell hyperplasia upon 2 weeks of allergen inhalation. This was accompanied by a significant increase in airway smooth muscle mass and hyperresponsiveness in BALB/c but not in C57BL/6 mice. However, airway eosinophilia was more pronounced in the C57BL/6 strain. Chronic allergen exposure (6 or 9 weeks) resulted in an increase in airway smooth muscle mass as well as subepithelial collagen and fibronectin deposition in both strains. The emergence of these structural changes paralleled the disappearance of inflammation in both C57BL/6 and BALB/c mice and loss of hyperresponsiveness in the BALB/c strain. TGF-beta(1 )was accordingly elevated in both strains.. Airway inflammation, remodelling and hyperresponsiveness are closely intertwined processes. Genetic background influences several aspects of the acute allergic phenotype. Chronic allergen exposure induces a marked airway remodelling that parallels a decreased inflammation, which was largely comparable between the two strains.

Topics: Acute Disease; Administration, Inhalation; Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chronic Disease; Disease Models, Animal; Eosinophilia; Eosinophils; Extracellular Matrix Proteins; Goblet Cells; Immunoglobulin E; Inflammation; Lung; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Muscle, Smooth; Ovalbumin; Transforming Growth Factor beta

Recent clinical trials, epidemiological studies and animal experiments have suggested that probiotics may help suppress the development of allergic responses.. To investigate whether the application of the probiotic Escherichia coli strain Nissle 1917 (EcN) protects mice from developing ovalbumin (OVA)-specific T helper-2 responses in the airways.. OVA-specific Th2 responses were induced by 2 intraperitoneal (i.p.) injections with OVA/alum followed by 1 intranasal (i.n.) challenge with OVA. EcN was given orally during the entire sensitization and challenge period, together with OVA/alum during the i.p. sensitizations, or i.n. before or during the airway challenge with OVA.. We found that when the bacteria were given together with OVA/alum airway eosinophilia, airway hyper-reactivity, goblet cell metaplasia and IL-5 levels in the bronchoalveolar lavage and mediastinal lymph node cell cultures were reduced. This effect was associated with increased numbers of IFN-gamma producing T helper-1 cells and IFN-gamma levels in the airways and strongly increased OVA-specific IgG(2a) titers in the serum. The suppressive effect on airway eosinophilia was dependent on IFN-gamma but not TLR-4. Applying EcN i.n. or orally did not reduce the development of allergen-specific Th2 responses.. Our results suggest that EcN can inhibit the development of allergic responses when the bacteria are present at the site of Th2 cell priming and that this immunomodulatory effect is due to a shift from Th2 to Th1 response. The data support the hypothesis that probiotics may help reduce allergic responses and that EcN may also be used as adjuvant therapy to induce allergen-specific Th1 responses.

Topics: Adjuvants, Immunologic; Administration, Intranasal; Administration, Oral; Allergens; Alum Compounds; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dendritic Cells; Eosinophilia; Escherichia coli; Female; Goblet Cells; Hypersensitivity; Interferon-gamma; Interleukin-5; Lipopolysaccharides; Mice; Mice, Inbred C57BL; Ovalbumin; Probiotics; Th1 Cells; Th2 Cells

This study aimed to investigate the effect of piperine on airway hyperresponsiveness, pulmonary eosinophilic infiltration, various immune cell phenotypes, Th2 cytokine production, immunoglobulin E and histamine production in a murine model of asthma.. Asthma was induced in Balb/c mice by ovalbumin sensitization and inhalation. Piperine (4.5 and 2.25 mg/kg) was orally administered 5 times a week for 8 weeks. At 1 day after the last ovalbumin exposure, airway hyperresponsiveness was determined and samples of bronchoalveolar lavage fluid, lung cells and serum were collected for further analysis.. Piperine-treated groups had suppressed eosinophil infiltration, allergic airway inflammation and airway hyperresponsiveness, and these occurred by suppression of the production of interleukin-4, interleukin-5, immunoglobulin E and histamine. Moreover, polymerase chain reaction products for thymus and activation regulated chemokine from lung cell RNA preparations were decreased in the piperine-treated group compared with control groups, although transforming growth factor-beta products were increased in the piperine-treated group.. The results suggest that the therapeutic mechanism by which piperine effectively treats asthma is based on a reduction of Th2 cytokines (interleukin-4, interleukin-5), eosinophil infiltration, and by marked reduction of thymus and activation regulated chemokine, eotaxin-2 and interleukin-13 mRNA expression (especially transcription of nuclear factor-kappaB dependent genes) in lung tissue, as well as reduced interleukin-4, interleukin-5 and eotaxin levels in bronchoalveolar lavage fluid, and histamine and ovalbumin-specific immunoglobulin E production in serum.

Topics: Alkaloids; Animals; Asthma; Benzodioxoles; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Eosinophils; Female; Histamine; Immunoglobulin E; Mice; Mice, Inbred BALB C; Ovalbumin; Piperidines; Polyunsaturated Alkamides; RNA, Messenger; T-Lymphocytes; Th2 Cells

Murine models of allergic lung disease have many similar traits to asthma in humans and can be used to investigate mechanisms of allergic sensitization and susceptibility factors associated with disease severity. The purpose of this study was to determine strain differences in allergic airway inflammation, immunoglobulin production, and changes in respiratory responses between systemic and mucosal sensitization routes in BALB/cJ, FVB/NJ, and C57BL/6J, and to provide correlations between immune and pathophysiological endpoints. After a single intranasal ovalbumin (OVA) challenge, all three strains of mice systemically sensitized with OVA and adjuvant exhibited higher airflow limitation than non-sensitized mice. No changes were seen in mice that were pre-sensitized via the nose with OVA. Systemic sensitization resulted in an elevated response to methacholine (MCH) in BALB/cJ and FVB/NJ mice and elevated total and OVA-specific IgE levels and pulmonary eosinophils in all three strains. The mucosal sensitization and challenge produced weaker responses in the same general pattern with the C57BL/6J strain producing less serum IgE, IL5, IL13, and eosinophils in lung fluid than the other two strains. The converse was found for IL6 where the C57BL/6J mice had more than twice the amount of this cytokine. The results show that the FVB/NJ and BALB/cJ mice are higher Th2-responders than the C57BL/6J mice and that the levels of pulmonary eosinophilia and cytokines did not fully track with MCH responsiveness. These differences illustrate the need to assess multiple endpoints to provide clearer associations between immune responses and type and severity of allergic lung disease.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Humans; Immunity, Mucosal; Immunoglobulin E; Immunoglobulin G; Lung; Lung Diseases; Mice; Ovalbumin; Species Specificity

Bronchial asthma is characterized by chronic airway inflammation caused by inflammatory cells. Phosphoinositide 3-kinases (PI3Ks) are known to play a prominent role in fundamental cellular responses of various inflammatory cells, including proliferation, differentiation, and cell migration. PI3Ks therefore are expected to have therapeutic potential for asthma. Although some investigations of the involvement between the pathogenesis of asthma and PI3K have been performed, it is unknown whether PI3Kgamma, a PI3K isoform, is involved in the pathogenesis of asthma.. We investigated the role of PI3Kgamma in allergen-induced allergic airway inflammation, airway hyperresponsiveness (AHR), and airway remodeling with PI3Kgamma-deficient mice.. After ovalbumin (OVA) sensitization, wild-type (WT) and PI3Kgamma-deficient mice were exposed to aerosolized OVA 3 days per week for 5 weeks.. In OVA-sensitized and OVA-challenged (OVA/OVA) PI3Kgamma-deficient mice, levels of airway inflammation, AHR, and airway remodeling were significantly decreased compared with those in OVA/OVA WT mice. On the other hand, no significant differences were detected in serum OVA-specific IgE and IgG1 levels and CD4/CD8 balance in bronchoalveolar lavage fluid between OVA/OVA WT mice and OVA/OVA PI3Kgamma-deficient mice. To determine in which phase of allergic responses PI3Kgamma plays a role, we transferred splenocytes from OVA-sensitized WT or PI3Kgamma-deficient mice to naive mice of either genotype. Similar increased levels of eosinophils were induced in both WT recipient mice but not in both PI3Kgamma-deficient recipient mice.. PI3Kgamma might be involved in allergic airway inflammation, AHR, and airway remodeling by regulating the challenge/effector phase of allergic responses.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Class Ib Phosphatidylinositol 3-Kinase; Cytokines; Female; Immunoglobulin E; Immunoglobulin G; Immunophenotyping; Isoenzymes; Mice; Mice, Inbred C57BL; Ovalbumin; Phosphatidylinositol 3-Kinases; T-Lymphocytes

Osteopontin (OPN) is a cytokine that is upregulated in epithelial cells and macrophages in the lungs of mice during chronic allergen challenge and airway remodeling and also in lungs of patients with asthma. However, it remains unclear whether OPN has an in vivo effect on lung remodeling in allergic asthma. Based on its ability to induce smooth muscle and fibroblast proliferation and migration we hypothesize that OPN regulates lung remodeling and also affects subsequent airway hyperresponsiveness (AHR).. Study the role of OPN in airway remodeling using OPN-knockout (KO) mice and a reversal approach administering recombinant mouse OPN (rOPN) in KO mice before challenge.. A chronic allergen-challenge model of airway remodeling with OPN KO mice, KO mice treated with rOPN, and human bronchial smooth muscle were used.. OPN deficiency protected mice against ova-induced AHR, which was associated with lower collagen and mucus production, gob-5 mRNA expression, submucosal cell area infiltration, and proliferation. Administration of rOPN to KO mice, just at the final five allergen challenges, exacerbated AHR and all the remodeling characteristics measured. In addition, rOPN increased the expression of IL-13 and pro-matrix metalloproteinase-9 in the lungs. Moreover, we demonstrated that rOPN induces proliferation of human BSM through binding to its alpha(v)beta3 integrin receptor and activation of PI3K/Akt downstream signaling pathway.. We conclude that OPN deficiency protects against remodeling and AHR. Thus our data reveal OPN as a novel therapeutic target for airway remodeling and associated AHR in chronic asthma.

Topics: Animals; Asthma; Blotting, Western; Bronchial Hyperreactivity; Cell Growth Processes; Disease Models, Animal; Extracellular Matrix; Goblet Cells; Humans; Interleukin-13; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocytes, Smooth Muscle; Osteopontin; Ovalbumin; Recombinant Proteins

T-bet(-/-) mice have been shown to have a profound deficiency in the ability to generate invariant NKT (iNKT) cells in the periphery due to a halt in terminal maturation, but despite this deficiency, T-bet(-/-) mice develop spontaneous airway hyperreactivity (AHR) and airway inflammation. Because in some situations the development of AHR requires the presence of iNKT cells, we sought to more clearly understand how AHR develops in T-bet(-/-) mice by examining T-bet(-/-) mice in several distinct mouse models of asthma, including spontaneous, OVA-induced and alpha-galactosylceramide (alpha-GalCer)-induced AHR. Surprisingly, we found that administration of alpha-GalCer, which very specifically activates iNKT cells, greatly increased the AHR response in the T-bet(-/-) mice. Moreover, in T-bet(-/-) mice, spontaneous AHR as well as AHR induced with OVA or alpha-GalCer were all eliminated by blocking CD1d, the restricting element of iNKT cells, using an anti-CD1d-blocking mAb. Although the number of the iNKT cells in T-bet(-/-) mice was reduced compared with that in wild-type mice, the remaining iNKT cells produced primarily IL-4 and IL-13, and only minimal amounts of IFN-gamma. We conclude therefore that the AHR that develops in T-bet(-/-) mice is dependent on the presence of iNKT cells, and that whereas T-bet(-/-) have reduced numbers of iNKT cells, these are sufficient for the development of AHR.

Topics: Animals; Antibodies, Blocking; Antigens, CD1d; Bronchial Hyperreactivity; Disease Models, Animal; Epitopes, T-Lymphocyte; Galactosylceramides; Lymphopenia; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Natural Killer T-Cells; Ovalbumin; T-Box Domain Proteins

Peroxynitrite has been shown to be crucially involved in airway hyperresponsiveness (AHR) after the late asthmatic reaction (LAR). Peroxynitrite production may result from simultaneous synthesis of nitric oxide (NO) and superoxide by inducible NO-synthase (iNOS) at low L-arginine concentrations. L-arginine availability to iNOS is regulated by its cellular uptake, which can be inhibited by eosinophil-derived polycations and by arginase, which competes with iNOS for the common substrate. Using a guinea pig model of allergic asthma, we investigated whether aberrant L-arginine homeostasis could underlie peroxynitrite-mediated AHR after the LAR. After the LAR, arginase activity in the airways and eosinophil peroxidase release from bronchoalveolar lavage cells were increased. These changes were associated with a 2.0-fold AHR to methacholine as measured in isolated perfused tracheal preparations. AHR was reduced by exogenous L-arginine administration. Moreover, both the arginase inhibitor N(omega)-hydroxy-nor-L-arginine (nor-NOHA) and the polycation antagonist heparin normalised airway responsiveness. These effects were reversed by the nitric oxide synthase inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME), indicating that both agents reduced AHR by restoring bronchodilating NO production. In conclusion, in allergen-challenged guinea pigs, the AHR after the LAR is caused by arginase- and polycation-induced attenuation of L-arginine availability to iNOS, which may switch the enzyme to simultaneous production of superoxide and NO, and, consequently, peroxynitrite.

Topics: Animals; Arginase; Arginine; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Guinea Pigs; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase Type II; Ovalbumin; Perfusion; Peroxynitrous Acid; Polyamines; Polyelectrolytes; Trachea

Topics: Animals; Antigens; Asthma; Bronchial Hyperreactivity; Cough; Dyspnea; Guinea Pigs; Histamine Antagonists; Inflammation; Male; Ovalbumin; Receptors, G-Protein-Coupled; Receptors, Histamine; Receptors, Histamine H4; Respiratory Mucosa

Eosinophil counts in the bronchoalveolar lavage fluid of wild-type (WT) mice increased after ovalbumin (OVA) challenge, a response that was diminished in comparably challenged low-expressing coagulation factor VII (FVII(tTA/tTA)) mice. Levels of T helper type 2 (Th2) cytokines, IL-4, IL-5, and IL-13, and eosinophil-attracting chemokines, eotaxin and RANTES, were also lower in the OVA-challenged FVII(tTA/tTA) mice. Eosinophils purified from low-FVII mice underwent apoptosis at a faster rate compared with WT eosinophils, and eosinophil migration in response to eotaxin was reduced in eosinophils obtained from FVII(tTA/tTA) mice. Airway hyperresponsiveness and mucous layer thickness were reduced in OVA-treated FVII(tTA/tTA) mice, and addition of exogenous coagulation factor X (FX) enhanced mucin production in human epithelial NCI-H292 cells. Correspondingly, incubation of FX with NCI-H292 cells resulted in activated (a) FX production, suggesting that the components required for FX activation were present on NCI-H292 cells. These results demonstrate that FVIIa functions in the asthmatic response to an allergen by stimulating lung eosinophilia, airway hyperresponsiveness, and mucin production, this latter effect through its ability to activate FX in conjunction with tissue factor.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cell Line; Cell Movement; Cell Survival; Cytokines; Eosinophils; Epithelial Cells; Factor VIIa; Female; Gene Expression Regulation; Inflammation; Inflammation Mediators; Lung; Mice; Mice, Inbred C57BL; Mucins; Ovalbumin; RNA, Messenger; Thromboplastin

Protein kinase C (PKC) is a complex family consisting of many types of isoenzymes, of which PKC-zeta, an atypical isoform, has been reportedly implicated in the regulation of apoptosis and NF-kappaB, as well as control of T-dependent responses. Based on the recent report that PKC-zeta controls TH2 response, the current study was aimed to evaluate PKC-zeta as a potential therapeutic target for asthma using a mouse model. Mouse allergic asthma was induced by repeated sensitization followed by intranasal challenge with OVA and PKC-zeta pseudosubstrate inhibitor (PPI) was intratracheally instilled before each OVA challenge. Airway hyperreactivity (AHR) was measured by beta-methacoline-induced airflow obstruction. Cellular and cytokine profile in bronchoalveolar lavage fluid (BALF) and level of serum IgE as well as cytokine production by draining lymph node cells were compared. AHR and numbers of eosinophils in BALF were significantly lowered by PPI, indicating that blocking of PKC-zeta activation alleviates asthmatic manifestations. Additionally, PPI instillation decreased IL-5 and IL-13 levels in BALF to approximately 20% of controls, but not IFN-gamma level. Instillation of PPI also caused a marked fall in the level of TNF-alpha, another NF-kappaB-dependent, proinflammatory cytokine. Serum OVA-specific IgE level and ex vivo IL-4, IL-5 and IL-13, but not IFN-gamma, production by peribronchial lymph node cells was also considerably lower in PPI-treated mice. In conclusion, blockade of PKC-zeta signals by intratracheal instillation of PPI alleviates allergen-specific TH2 response as well as asthmatic manifestations and hence PKC-zeta is a promising target for treatment of asthma.

Topics: Animals; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Proliferation; Disease Models, Animal; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Eosinophils; Immunoglobulin G; Interleukin-13; Interleukin-4; Interleukin-5; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Ovalbumin; Protein Kinase C; Pyrazoles; Pyrimidines; Tumor Necrosis Factor-alpha

Dendritic cell (DC) subsets display different functional roles in regulating immune responses and lead to various outcomes, including T(H)1 versus T(H)2 or regulatory versus immunologic responses. Administration of Fms-like tyrosine kinase 3 (Flt3) ligand prevents and reverses allergic airway inflammation and airway hyperresponsiveness in a mouse model. However, the underlying mechanisms are unclear.. We characterized and examined the role of lung DC subsets in the therapeutic effect of Flt3 ligand.. DCs were isolated from the lungs of ovalbumin (OVA)-sensitized and OVA-challenged mice treated with recombinant human Flt3 ligand. Two populations of CD11c+ cells labeled with fluorochrome-conjugated antibodies were sorted. The ability of the purified cells to stimulate T-cell proliferation and cytokine secretion patterns by different DC subsets was examined. Also, DCs were adoptively transferred in mice to examine their effect on pulmonary function.. Two DC populations, CD11c(high)CD11b(low) and CD11c(low)CD11b(high), were identified in the lungs of naive and OVA-sensitized and OVA-challenged mice with and without treatment with Flt3 ligand. The expression levels of CD8alpha, B220, CD19, F4/80, MHC II, CCR7, CD40, programmed death ligand 1, programmed death ligand 2, CD80, and CD86 were distinctly different between the 2 DC populations, which supports the notion that CD11c(high)CD11b(low) and CD11c(low)CD11b(high) DCs potentially have regulatory and immunogenic properties, respectively. Administration of Flt3 ligand increased the DCs with regulatory potential in the lungs of antigen-sensitized mice, and CD11c(high)CD11b(low) DCs acquired a maximum degree of regulatory capacity after Flt3 ligand treatment.. These data suggest that Flt3 ligand reverses airway hyperresponsiveness by regulating the function of lung DCs in a mouse model of allergic airway inflammation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; CD11b Antigen; CD11c Antigen; Cytokines; Dendritic Cells; Female; Immunophenotyping; Lung; Lymphocyte Activation; Membrane Proteins; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes; Th2 Cells

Existing asthma models develop tolerance when chronically exposed to the same allergen.. We sought to establish a chronic model that sustains features of asthma long after discontinuation of allergen exposure.. We immunized and exposed mice to a combination of single, double, or triple allergens (dust mite, ragweed, and Aspergillus species) intranasally for 8 weeks. Airway hyperreactivity (AHR) and morphologic features of asthma were studied 3 weeks after allergen exposure. Signaling effects of the allergens were studied on dendritic cells.. Sensitization and repeated exposure to a single allergen induced tolerance. Sensitization to double and especially triple allergens broke through tolerance and established AHR, eosinophilic inflammation, mast cell and smooth muscle hyperplasia, mucus production, and airway remodeling that persisted at least 3 weeks after allergen exposure. Mucosal exposure to triple allergens in the absence of an adjuvant was sufficient to induce chronic airway inflammation. Anti-IL-5 and anti-IL-13 antibodies inhibited inflammation and AHR in the acute asthma model but not in the chronic triple-allergen model. Multiple allergens produce a synergy in p38 mitogen-activated protein kinase signaling and maturation of dendritic cells, which provides heightened T-cell costimulation at a level that cannot be achieved with a single allergen.. Sensitivity to multiple allergens leads to chronic asthma in mice. Multiple allergens synergize in dendritic cell signaling and T-cell stimulation that allows escape from the single allergen-associated tolerance development.

Topics: Allergens; Ambrosia; Animals; Aspergillus; Asthma; Bronchial Hyperreactivity; Chemokines; Chronic Disease; Cytokines; Dendritic Cells; Disease Models, Animal; Extracellular Signal-Regulated MAP Kinases; Female; Immune Tolerance; Immunization; Immunoglobulin E; Mice; Mice, Inbred BALB C; Ovalbumin; Pyroglyphidae

Fluticasone affects airway bronchial hyperresponsiveness (BHR) and enhances bronchodilation and bronchoprotection induced by beta-adrenergic agonists. Interleukin 13 (IL-13), however, induces BHR.. To test the hypotheses that fluticasone inhibits BHR after either allergen sensitization or IL-13 administration and that fluticasone restores the bronchodilation and bronchoprotective effects of beta-agonists.. The BHR to methacholine induced by IL-13 or ovalbumin was determined in BALB/c mice, and the provocation concentration of methacholine that caused an increase in enhanced pause in expiration of 200% (PC200) was calculated. We compared this response to methacholine in control mice with the response after treatment with IL-13 receptor alpha 2-IgGFc fusion protein (IL-13R alpha 2) (an IL-13 blocker), fluticasone, albuterol, salmeterol, fluticasone-albuterol, and fluticasone-salmeterol.. IL-13R alpha 2 (PC200, 17.59) completely blocks the BHR-induced effects of IL-13 (PC200, 7.28; P < .005). After IL-13 therapy (PC200, 5.90; P < .005), 1 mg/mL of albuterol (PC200, 3.38; P = .33), fluticasone (PC200, 4.59; P = .40), or fluticasone plus 50 microg/mL of salmeterol (PC200, 5.59; P = .11) showed no significant bronchoprotection. In nonsensitized mice, fluticasone plus 0.25 microg/mL of salmeterol (PC200, 25.90; P < .005) showed significantly greater bronchoprotection than did salmeterol alone (PC200, 11.08; P = .26). Fluticasone plus 0.3 mg/mL of albuterol and fluticasone plus 1 mg/mL of albuterol were significantly more protective than was fluticasone or albuterol alone in ovalbumin-sensitized mice.. The protective effects of fluticasone, beta-agonists, and fluticasone plus beta-agonists are significantly less in IL-13-treated mice than in nonsensitized or ovalbumin-sensitized mice.

Topics: Adrenal Cortex Hormones; Adrenergic beta-Agonists; Albuterol; Androstadienes; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Bronchodilator Agents; Drug Combinations; Female; Fluticasone; Fluticasone-Salmeterol Drug Combination; Interleukin-13; Interleukin-13 Receptor alpha2 Subunit; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Salmeterol Xinafoate

Phosphodiesterase 4 selective inhibitor may prevent airway inflammation and remodeling.. The aim of this study was to investigate the effects of KF19514, a phosphodiesterase 4 and 1 dual inhibitor, on chronic airway inflammation and remodeling following chronic exposure to aerosolized antigen in mice.. Ovalbumin (OVA) was administered intraperitoneally to BALB/c mice on days 0 and 14, and the mice were then exposed to aerosolized OVA daily for 4 weeks. Twenty-four hours following the final inhalation, bronchial responsiveness to acetylcholine was measured, and histologic examination and hydroxyproline content of the lung were evaluated.. Bronchial responsiveness to acetylcholine, number of inflammatory cells and eosinophils in the lamina propria, thickness of epithelial and subepithelial collagen layers, and hydroxyproline content of the lung increased following chronic exposure to OVA for 7 weeks. KF19514 significantly prevented all of these changes.. Phosphodiesterase 4 and 1 inhibitors such as KF19514 may help prevent bronchial hyperresponsiveness and chronic asthma-induced airway remodeling.

Topics: Acetylcholine; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Cyclic Nucleotide Phosphodiesterases, Type 1; Female; Hydroxyproline; Inflammation; Lung; Mice; Mice, Inbred BALB C; Mucous Membrane; Naphthyridines; Ovalbumin; Phosphodiesterase 4 Inhibitors; Phosphodiesterase Inhibitors; Pulmonary Eosinophilia; Respiratory Mucosa; Specific Pathogen-Free Organisms; Trachea; Vaccination

Corneal allografts transplanted into hosts with allergic conjunctivitis experience an increased incidence and swifter tempo of immune rejection compared to corneal allografts transplanted to nonallergic hosts. Previous findings suggested that increased risk for rejection was not a local effect produced by an inflamed eye, but was due to perturbation of the systemic immune responses to alloantigens on the corneal allograft. We tested the hypothesis that another allergic disease, airway hyperreactivity (AHR), would also increase the risk for corneal allograft rejection. Induction of AHR with either ovalbumin (OVA) or short ragweed (SRW) extract prior to keratoplasty resulted in a steep increase in the speed and incidence of corneal allograft rejection. Delayed-type hypersensitivity (DTH) responses to corneal alloantigens were closely associated with corneal allograft rejection. However, the deleterious effect of AHR on corneal allograft survival was not reflected in a heightened magnitude of allospecific DTH, cytotoxic T lymphocyte and lymphoproliferative responses to the alloantigens on the corneal allograft. Unlike Th2-based immediate hypersensitivity, CD8+ T-cell-based contact hypersensitivity to oxazolone did not increase the risk for corneal allograft rejection. Thus, Th2-based allergic diseases significantly reduce the immune privilege of the corneal allograft and represent important risk factors for consideration in the atopic patient.

Topics: Animals; Asthma; Bronchial Hyperreactivity; CD8-Positive T-Lymphocytes; Conjunctivitis, Allergic; Corneal Transplantation; Disease Models, Animal; Female; Graft Rejection; Graft Survival; Isoantigens; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Risk Factors; T-Lymphocytes, Cytotoxic; Transplantation, Homologous

Definitive conclusions regarding the antiinflammatory effects of macrolide antibiotics for treatment of asthma are difficult to formulate since their beneficial effects may be related to their antimicrobial action. We hypothesized that azithromycin possesses distinct antiinflammatory properties and tested this assumption in a noninfectious mouse model of allergic asthma.. To induce allergic airway inflammation, 7-week-old BALB/cJ mice underwent intraperitoneal ovalbumin sensitization on days 0 and 7 followed by an intranasal challenge on day 14. Mice were treated with azithromycin or phosphate-buffered saline (PBS) solution on days 13 through 16. On day 17, airway inflammation was assessed by quantifying leukocytes in the airway, expression of multiple inflammatory mediators in the BAL fluid, and mucous cell metaplasia. In a separate set of experiments, azithromycin or PBS solution treatment were initiated after the ovalbumin challenge. Each experiment was repeated 3 times (a total of 9 to 11 mice in each group).. Compared to treatment with PBS solution, azithromycin attenuated the ovalbumin-dependent airway inflammation. We observed a decrease in total leukocytes in the lung tissue and BAL fluid. In addition, azithromycin attenuated the expression of cytokines (eg, interleukin [IL]-13 and IL-5) and chemokines (eg, CCL2, CCL3, and CCL4) in the BAL fluid and abrogated the extent of mucous cell metaplasia. Similar antiinflammatory effects were observed when azithromycin treatment was initiated after the ovalbumin challenge.. In this noninfectious mouse model of allergic asthma, azithromycin attenuated allergic airway inflammation. These findings demonstrate an antiinflammatory effect of azithromycin and suggest azithromycin may have beneficial effects in treating noninfectious airway inflammatory diseases, including asthma.

Topics: Animals; Asthma; Azithromycin; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Immunoglobulin E; Inflammation; Inflammation Mediators; Leukocyte Count; Lung; Mice; Mice, Inbred BALB C; Mucous Membrane; Ovalbumin; Random Allocation; Reference Values; Risk Factors; Sensitivity and Specificity

The most widely used protocol for the induction of experimental allergic airway inflammation in mice involves sensitization by intraperitoneal (i.p.) injections of the antigen ovalbumin (OVA) used in conjunction with the adjuvant aluminium hydroxide (alum). Although adjuvants are frequently used, there are questions regarding the necessity of alum for murine asthma studies due to the non-physiological nature of this chemical.. The objective of this study was to compare experimental asthma phenotypes between adjuvant and adjuvant-free protocols of murine allergic airway inflammation in an attempt to develop a standardized alternative to adjuvant use.. An adjuvant-free OVA model of experimental asthma was investigated in BALB/c mice using i.p. or subcutaneous (s.c.) sensitization routes. For the s.c. sensitization, beta-galactosidase (beta-gal) was also tested as an antigen. In addition, OVA adjuvant and adjuvant-free sensitization protocols were compared in BALB/c and C57BL/6 mice. Open-field testing was performed to assess the effect of alum on mouse behaviour.. Comparison of adjuvant vs. adjuvant-free and i.p. vs. s.c. protocols revealed that both adjuvant use and route of antigen application significantly influenced OVA-specific antibody production. Comparison of adjuvant and adjuvant-free protocols in this study clearly demonstrated the non-requirement of alum for the induction of acute allergic airway inflammation, as both protocols induce a similar disease phenotype. BALB/c mice were significantly more susceptible than C57BL/6 mice to sensitization. Using the improved s.c. adjuvant-free protocol, it was demonstrated that alternative antigens such as beta-gal can also be utilized. Behavioural studies indicated severe distress in mice treated with alum.. The OVA s.c. adjuvant-free protocol used in this study generates a phenotype comparable to the benchmark adjuvant protocol widely used in the literature. The adjuvant-free alternative avoids the added complication of non-physiological adjuvants that may interfere with asthma treatment or prevention strategies.

Topics: Adjuvants, Immunologic; Allergens; Aluminum Hydroxide; Animals; Asthma; beta-Galactosidase; Bronchial Hyperreactivity; Disease Models, Animal; Female; Injections, Intraperitoneal; Injections, Subcutaneous; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Phenotype; Sensitivity and Specificity; Skin Tests

To investigate whether challenge with increasing allergen doses could differently affect allergen-induced airway hyperresponsiveness (AHR) and inflammatory cell accumulation in mouse model of asthma, providing an experimental model to investigate their relationship.. AHR and accumulation of inflammatory cells in bronchoalveolar lavage fluid (BALF) and into the lungs were compared in ovalbumin-sensitized mice that were challenged intranasally with 2.5, 10, 25 or 100 microg of ovalbumin/mouse.. Both AHR and inflammatory cell accumulation were proportional to the ovalbumin dose used for challenge. However, in group challenged with 10 microg of ovalbumin airway inflammation was present, although allergen-induced AHR was not detected. Additional analysis indicated that neither mucous hyperproduction nor eosinophil degranulation could be correlated to presence of AHR in this model, whereas concentration of interleukin (IL)-13 in BALF was increased only in those groups in which AHR was present.. Altogether, intranasal challenge of mice with increasing allergen doses could serve as a suitable experimental system for investigation of mechanisms by which airway inflammation leads to allergen-induced AHR. Our initial findings are in line with previous reports that dissociate AHR from amount of eosinophil accumulation and imply the role of IL-13 in this process.

Topics: Administration, Intranasal; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophil Peroxidase; Eosinophils; Humans; Inflammation; Interleukin-13; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin

Although anticholinergic therapy inhibits bronchoconstriction in asthmatic patients and antigen-challenged animals, administration of atropine 1 h before antigen challenge significantly potentiates airway hyperreactivity and eosinophil activation measured 24 h later. This potentiation in airway hyperreactivity is related to increased eosinophil activation and is mediated at the level of the airway nerves. Since eosinophils produce nerve growth factor (NGF), which is known to play a role in antigen-induced airway hyperreactivity, we tested whether NGF mediates atropine-enhanced, antigen challenge-induced hyperreactivity. Antibody to NGF (Ab NGF) was administered to sensitized guinea pigs with and without atropine pretreatment (1 mg/kg iv) 1 h before challenge. At 24 h after challenge, animals were anesthetized, vagotomized, paralyzed, and ventilated. Electrical stimulation of both vagus nerves caused bronchoconstriction that was increased in challenged animals. Atropine pretreatment potentiated antigen challenge-induced hyperreactivity. Ab NGF did not affect eosinophils or inflammatory cells in any group, nor did it prevent hyperreactivity in challenged animals that were not pretreated with atropine. However, Ab NGF did prevent atropine-enhanced, antigen challenge-induced hyperreactivity and eosinophil activation (assessed by immunohistochemistry). This effect was specific to NGF, since animals given control IgG remained hyperreactive. These data suggest that anticholinergic therapy amplifies eosinophil interactions with airway nerves via NGF. Therefore, therapeutic strategies that target both eosinophil activation and NGF-mediated inflammatory processes in allergic asthma are likely to be beneficial.

Topics: Animals; Antigens; Asthma; Atropine; Blood Pressure; Bradycardia; Bronchial Hyperreactivity; Bronchoconstriction; Bronchodilator Agents; Electric Stimulation; Eosinophils; Female; Guinea Pigs; Heart Rate; Muscle, Smooth; Nerve Growth Factor; Ovalbumin; Parasympathetic Nervous System; Receptor, Muscarinic M2; Specific Pathogen-Free Organisms; Vagotomy; Vagus Nerve

It is widely accepted that atopic asthma depends on an allergic response in the airway, yet the immune mechanisms that underlie the development of airway hyperresponsiveness (AHR) are poorly understood. Mouse models of asthma have been developed to study the pathobiology of this disease, but there is considerable strain variation in the induction of allergic disease and AHR. The aim of this study was to compare the development of AHR in BALB/c, 129/Sv, and C57BL/6 mice after sensitization and challenge with ovalbumin (OVA). AHR to methacholine was measured using a modification of the forced oscillation technique in anesthetized, tracheostomized mice to distinguish between airway and parenchymal responses. Whereas all strains showed signs of allergic sensitization, BALB/c was the only strain to develop AHR, which was associated with the highest number of activated (CD69(+)) CD4(+) T cells in the airway wall and the highest levels of circulating OVA-specific IgG(1). AHR did not correlate with total or antigen-specific IgE. We assessed the relative contribution of CD4(+) T cells and specific IgG(1) to the development of AHR in BALB/c mice using adoptive transfer of OVA-specific CD4(+) T cells from DO11.10 mice. AHR developed in these mice in a progressive fashion following multiple OVA challenges. There was no evidence that antigen-specific antibody had a synergistic effect in this model, and we concluded that the number of antigen-specific T cells activated and recruited to the airway wall was crucial for development of AHR.

Topics: Airway Resistance; Animals; Antigens, CD; Antigens, Differentiation, T-Lymphocyte; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; CD4 Lymphocyte Count; CD4-Positive T-Lymphocytes; Disease Models, Animal; Female; Immunoglobulin E; Immunoglobulin G; Lectins, C-Type; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Specific Pathogen-Free Organisms

Methacholine (Mch) is routinely used to assess bronchial hyperreactivity; however, little is known about the differences in the lung response pattern between this provocation and that observed with ovalbumin (Ova) after allergic sensitization.. To compare (1) the central versus peripheral effects of Mch and Ova within the lung by combining measurements of airway and tissue mechanics with synchrotron radiation (SR) imaging, and (2) to assess the extent to which mechanical and imaging parameters are correlated.. We used the low-frequency forced oscillation technique and SR imaging in control (n = 12) and ovalbumin-sensitized (n = 13) rabbits, at baseline, during intravenous Mch infusion (2.5 microg/kg/min, 5.0 microg/kg/min, or 10.0 microg/kg/min), after recovery from Mch, and after intravenous Ova injection (2.0 mg). We compared intravenous Mch challenge with inhaled Mch (125 mg/ml, 90 s) in a separate group of control animals (n = 5).. Airway conductance and tissue elastance were measured by low-frequency forced oscillation technique. The central airway cross-sectional area, the ventilated alveolar area, and the heterogeneity of specific ventilation were quantified by SR imaging. Mch infusion induced constriction predominantly in the central airways, whereas Ova provocation affected mainly the peripheral airways, leading to severe ventilation heterogeneities in sensitized animals. Mch inhalation affected both conducting and peripheral airways. The correlations between airway conductance and central airway cross-sectional area (R = 0.71) and between tissue elastance and ventilated alveolar area (R = -0.72) were strong.. The pattern of lung response caused by intravenous Mch and Ova are fundamentally different. Although inhaled Mch induces a heterogeneous lung response similar to that observed with intravenous allergen, these similar patterns are due to different mechanisms.

Topics: Administration, Inhalation; Airway Resistance; Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstrictor Agents; Disease Models, Animal; Dose-Response Relationship, Drug; Image Processing, Computer-Assisted; Infusions, Intravenous; Lung; Lung Compliance; Methacholine Chloride; Oscillometry; Ovalbumin; Pulmonary Ventilation; Rabbits; Respiratory Mechanics; Synchrotrons; Tomography, X-Ray Computed

Tyrosine kinase signaling cascades play a critical role in the pathogenesis of allergic airway inflammation. Sunitinib, a multitargeted receptor tyrosine kinase inhibitor, has been reported to exert potent immunoregulatory, anti-inflammatory and anti-fibrosis effects. We investigated whether sunitinib could suppress the progression of airway inflammation, airway hyperresponsiveness (AHR), and airway remodeling in a murine model of chronic asthma.. Ovalbumin (OVA)-sensitized mice were chronically challenged with aerosolized OVA for 8 weeks. Some mice were intragastrically administered with sunitinib (40 mg/kg) daily during the period of OVA challenge. Twelve hours after the last OVA challenge, mice were evaluated for the development of airway inflammation, AHR and airway remodeling. The levels of total serum immunoglobulin E (IgE) and Th2 cytokines (interleukin (IL)-4 and IL-13) in bronchoalveolar lavage fluid (BALF) were measured by ELISA. The expression of phosphorylated c-kit protein in the lungs was detected by immunoprecipitation/Western blotting (IP/WB) analysis.. Sunitinib significantly inhibited eosinophilic airway inflammation, persistent AHR and airway remodeling in chronic experimental asthma. It reduced levels of total serum IgE and BALF Th2 cytokines and also lowered the expression of phosphorylated c-kit protein in remodelled airways.. Sunitinib may inhibit the development of airway inflammation, AHR and airway remodeling. It is potentially beneficial to the prevention or treatment of asthma.

Topics: Angiogenesis Inhibitors; Animals; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Female; Immunoglobulin E; Immunohistochemistry; Immunoprecipitation; In Vitro Techniques; Indoles; Inflammation; Interleukin-13; Interleukin-4; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Proto-Oncogene Proteins c-kit; Pyrroles; Sunitinib

Helper T (Th) cells are deeply involved in the pathophysiology of bronchial asthma, such as eosinophilic inflammation, bronchial hyperresponsiveness and remodeling. However, it is still unclear how Th cells contribute to airflow limitation, another cardinal feature of bronchial asthma.. Unprimed BALB/c mice were transferred with ovalbumin (OVA)-reactive Th clones. Pulmonary function was monitored using a Buxco BioSystem Plethysmograph before and after OVA challenge.. When Th-transferred mice were challenged with OVA, enhanced pause (P(enh)), an indicator of airflow limitation was significantly increased 6 and 24 h after challenge, while no response was observed 30 min after challenge. Neither bovine serum albumin, an irrelevant antigen, challenge on Th-transferred mice nor OVA challenge on Th-non-transferred mice caused airway responses.. Th cells conferred antigen-induced airflow limitation to unprimed mice.

Topics: Airway Obstruction; Animals; Antigens; Asthma; Bronchial Hyperreactivity; Clone Cells; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; T-Cell Antigen Receptor Specificity; T-Lymphocytes, Helper-Inducer; Time Factors

Animal models of asthma mimic major features of human disease. Since the genetic background of experimental animals might affect hyperresponsiveness and inflammation, we studied its potential influence and the mechanisms leading to differences in strains.. We applied a mouse model of allergic asthma to BALB/c and C57BL/6 mice.. BALB/c mice displayed greater levels of airway reactivity to methacholine than C57BL/6 mice. Moreover, BALB/c mice exhibited higher numbers of mast cells in lung tissue when compared to C57BL/6. On the contrary, eosinophil and neutrophil counts in bronchoalveolar lavage fluid (BALF) as well as peribronchial eosinophilia were greater in C57BL/6. IL (Interleukin)-4, IL-5, IL-13, and CCL11 levels measured in whole-lung extracts were higher in BALB/c, while, in sharp contrast, CCL11 and CCL5 levels were higher in BALF of C57BL/6 mice.. We observed phenotypic differences between C57BL/6 and BALB/c mice in an asthma model with different distributions of pro-inflammatory cytokines and inflammatory cells.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Humans; Immunoglobulin E; Inflammation; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin

The ability of corticosteroids to reduce airway inflammation and improve lung function is significantly reduced in asthmatics who are tobacco smokers compared with asthmatics who are nonsmokers. As not only high levels of tobacco smoke exposure in active smokers, but also significantly lower levels of tobacco smoke exposure from passive environmental tobacco smoke (ETS) exposure in nonsmokers can increase both asthma symptoms and the frequency of asthma exacerbations, we utilized a mouse model to determine whether corticosteroids can reduce levels of airway inflammation, airway remodeling, and airway hyperreactivity in mice exposed to the combination of chronic ETS and ovalbumin (OVA) allergen. Chronic ETS exposure alone did not induce increases in eosinophilic airway inflammation, airway remodeling, or airway hyperreactivity. Mice exposed to chronic OVA allergen had significantly increased levels of peribronchial fibrosis, increased thickening of the smooth muscle layer, increased mucus, and increased airway hyperreactivity, which was significantly enhanced by coexposure to the combination of chronic ETS and chronic OVA allergen. Administration of corticosteroids to mice exposed to chronic ETS and OVA allergen significantly reduced levels of eosinophilic airway inflammation, mucus production, peribronchial smooth muscle thickness, airway hyperreactivity, and the number of peribronchial TGF-beta1+ cells. Overall, this study demonstrates that corticosteroids can significantly reduce levels of eosinophilic inflammation, mucus expression, airway remodeling, and airway hyperreactivity in chronic ETS-exposed mice challenged with allergen.

Topics: Adrenal Cortex Hormones; Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophils; Female; Fibrosis; Mice; Mice, Inbred BALB C; Mucus; Muscle, Smooth; Ovalbumin; Pneumonia; Tobacco Smoke Pollution; Transforming Growth Factor beta1

Asthma is a dynamic disorder of airway inflammation and airway remodeling with an imbalance in T helper type 1 (Th(1))/Th(2) immune response. Increased Th(2) cytokines such as IL-4 and IL-13 induce arginase either directly or indirectly through transforming growth factor-beta(1) (TGF-beta(1)) and lead to subepithelial fibrosis, which is a crucial component of airway remodeling. Synthetic antimalarials have been reported to have immunomodulatory properties. Mepacrine is known for its reduction of airway inflammation in short-term allergen challenge model by reducing Th(2) cytokines and cysteinyl leukotrienes, which has an important role in the development of airway remodeling features. Therefore, we hypothesized that mepacrine may reduce airway remodeling. For this, extended subacute ovalbumin mice model of asthma was developed; these mice showed an increased expression of profibrotic mediators, subepithelial fibrosis, and goblet cell metaplasia along with airway inflammation, increased Th(2) cytokines, allergen-specific IgE, IgG(1), increased cytosolic PLA(2) (cPLA(2)), and airway hyperresponsiveness. Presence of intraepithelial eosinophils and significant TGF-beta(1) expression in subepithelial mesenchymal regions by repeated allergen exposures indicate that asthmatic mice of this study have developed human mimicking as well as late stages of asthma. However, mepacrine treatment decreased Th(2) cytokines and subepithelial fibrosis and alleviated asthma features. These reductions by mepacrine were associated with a decrease in levels and expression of TGF-beta(1) and the reduction in activity, expression of arginase in lung cytosol, and immunolocalization in inflammatory cells present in perivascular and peribronchial regions. These results suggest that mepacrine might reduce the development of subepithelial fibrosis by reducing the arginase and TGF-beta(1). These effects of mepacrine likely underlie its antiairway remodeling action in asthma.

Topics: Animals; Arginase; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Epithelial Cells; Fibrosis; Goblet Cells; Hydroxyeicosatetraenoic Acids; Inflammation; Lung; Metaplasia; Mice; Mice, Inbred BALB C; Ovalbumin; Phospholipases A2, Cytosolic; Quinacrine; Transforming Growth Factor beta1

Suppressing the abnormalities associated with asthma has been difficult to accomplish using immunotherapy or vaccination once the disease is established. The effector cells necessary for effective immunization/vaccination and immunotherapy of asthma are also not well understood. Therefore, we vaccinated allergen (OVA)-sensitized mice to determine whether therapeutic immunization could suppress airway hyperresponsiveness (AHR) and inflammation and to identify key immune effector cells and cytokines. Mice were immunized with a vaccine comprised of Ag and cationic liposome-DNA complexes (CLDC), a vaccine which has previously been shown to elicit strong CD4(+) and CD8(+) T cell responses and activation of Th1 immunity. We showed that immunization with the OVA-CLDC vaccine significantly suppressed AHR, eosinophilia, goblet cell metaplasia, and Th2 cytokine production. In contrast, immunization with CLDC alone suppressed eosinophilia and Th2 cytokine production, but failed to suppress AHR and goblet cell changes. Using adoptive transfer experiments, we found that suppression of AHR was mediated by Ag-specific CD8(+) T cells and was dependent on IFN-gamma production by the transferred T cells. Thus, we conclude that generation of strong, allergen-specific CD8(+) T cell responses by immunization may be capable of suppressing AHR and allergic airway inflammation, even in previously sensitized and challenged mice.

Topics: Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Cations; CD8-Positive T-Lymphocytes; Cytokines; Epitopes, T-Lymphocyte; Female; H-2 Antigens; Humans; Immunosuppressive Agents; Inflammation Mediators; Liposomes; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Vaccines, DNA

TNF receptor 1 can activate signaling pathways leading to the activation of NF-kappaB. A20, an NF-kappaB-inducible protein, negatively regulates these signaling pathways and acts as an anti-inflammatory mediator. Therefore, A20 is viewed as a potential therapeutic target for inflammatory disease. In this study, we examined the effect of A20 on an OVA-induced allergic airway inflammation model in mice. We used an adenovirus containing A20 cDNA (Ad-A20) that was delivered intratracheally before OVA challenge. Single administration of Ad-A20 reduced airway inflammatory cell recruitment and peribronchiolar inflammation and suppressed the production of various cytokines in bronchoalveolar fluid. In addition, Ad-A20 suppressed mucus production and prevented the development of airway hyperresponsiveness. The protective effect of Ad-A20 was mediated by the inhibition of the NF-kappaB signaling pathway. Taken together, our results suggest that the development of an immunoregulatory strategy based on A20 may have therapeutic potential for the treatment of allergic asthma.

Topics: Adenoviridae; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cysteine Endopeptidases; Cytokines; Genetic Vectors; Inflammation; Intracellular Signaling Peptides and Proteins; Mice; NF-kappa B; Ovalbumin; Signal Transduction; Tumor Necrosis Factor alpha-Induced Protein 3

Macrolide antibiotics, a class of potent antimicrobials, also possess immunomodulatory/anti-inflammatory properties. These properties are considered fundamental for the efficacy of macrolide antibiotics in the treatment of diffuse panbronchiolitis and cystic fibrosis. In patients with asthma, macrolide antibiotics have been reported to reduce airway hyperresponsiveness and improve pulmonary function. However, their beneficial actions in asthmatics possibly could be attributed to antimicrobial activity against atypical pathogens (e.g. Chlamydia pneumoniae), corticosteroid-sparing effect (inhibition of exogenous corticosteroid metabolism), and/or their anti-inflammatory/immunomodulatory effects. In order to investigate whether efficacy of macrolide antibiotics in asthma results from their immunomodulatory/anti-inflammatory activity, the influence of clarithromycin pretreatment (2 h before challenge) was examined on ovalbumin-induced airway hyperresponsiveness and airway inflammation in the mouse. Clarithromycin treatment (200 mg/kg intraperitoneally) decreased IL-4, IL-5, IL-13, CXCL2 and CCL2 concentrations in bronchoalveolar lavage fluid and markedly reduced inflammatory cell accumulation in bronchoalveolar lavage fluid and into the lungs, as revealed by histopathological examination. Furthermore, clarithromycin-induced reduction in inflammation was accompanied by normalization of airway hyperresponsiveness. In summary, in ovalbumin-induced mouse models, clarithromycin efficiently inhibited two important pathological characteristics of asthma, airway hyperresponsiveness and inflammation. These data suggest that the efficacy of clarithromycin, as well as of other macrolide antibiotics, in asthmatic patients could be attributed to their anti-inflammatory/immunomodulatory properties, and not only to their antimicrobial activity or exogenous corticosteroid-sparing effects.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Clarithromycin; Disease Models, Animal; Immunologic Factors; Inflammation; Lung; Male; Mice; Ovalbumin; Respiratory System

Airway hyperresponsiveness is a constant feature of asthma. The aim of the present study was to investigate airway hyperreactivity mediated by contractile and dilative receptors in an ovalbumin (OVA)-induced model of rat asthma.. Asthmatic E3 rats were prepared by intraperitoneal injection with OVA/aluminum hydroxide and then challenged with intranasal instillation of OVA-PBS two weeks later. The myograph method was used to measure the responses of constriction and dilatation in the trachea, main bronchi and lobar bronchi.. In asthmatic E3 rats, beta(2) adrenoceptor-mediated relaxation of airway smooth muscle pre-contracted with 5-HT was inhibited, and there were no obvious difference in relaxation compared with normal E3 rats. Contraction of lobar bronchi mediated by 5-HT and sarafotoxin 6c was more potent than in the trachea or main bronchi. Airway contractions mediated by the endothelin (ET)(A) receptor, ET(B) receptor and M(3) muscarinic receptor were augmented, and the augmented contraction was most obvious in lobar bronchi. The order of efficacy of contraction for lobar bronchi induced by agonists was ET-1, sarafotoxin 6c>ACh>5-HT. OX8 (an antibody against CD8(+) T cells) strongly shifted and OX35 (an antibody against CD4(+) T cells) modestly shifted isoprenaline-induced concentration-relaxation curves in a nonparallel fashion to the left with an increased R(max) in asthmatic rats and sarafotoxin 6c-induced concentration-contractile curves to the right with a decreased E(max).. The inhibition of airway relaxation and the augmentation of contraction mediated by receptors contribute to airway hyperresponsiveness and involve CD8(+) and CD4(+) T cells.Acta Pharmacologica Sinica (2009) 30: 965-972; doi: 10.1038/aps.2009.61.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchodilator Agents; Dose-Response Relationship, Drug; Humans; Isoproterenol; Muscle Contraction; Muscle Relaxation; Muscle, Smooth; Ovalbumin; Rats; Receptors, Cell Surface; Respiratory System; Serotonin; Vasoconstrictor Agents; Viper Venoms

We showed recently that IL-4 causes mitochondrial dysfunction in allergic asthma. IL-4 is also known to induce 12/15-lipoxygenase (12/15-LOX), a potent candidate molecule in asthma. Because vitamin E (Vit-E) reduces IL-4 and inhibits 12/15-LOX in vitro, here we tested the hypothesis that Vit-E may be effective in restoring key mitochondrial dysfunctions, thus alleviating asthma features in an experimental allergic murine model. Ovalbumin (OVA)-sensitized and challenged male BALB/c mice showed the characteristic features of asthma such as airway hyperresponsiveness (AHR), airway inflammation, and airway remodeling. In addition, these mice showed increase in the expression and metabolites of 12/15-LOX, reduction in the activity and expression of the third subunit of mitochondrial cytochrome-c oxidase, and increased cytochrome c in lung cytosol, which indicate that OVA sensitization and challenge causes mitochondrial dysfunction. Vit-E was administered orally to these mice, and 12/15-LOX expression, key mitochondrial functions, ultrastructural changes of mitochondria in bronchial epithelia, and asthmatic parameters were determined. Vit-E treatment reduced AHR, Th2 response including IL-4, IL-5, IL-13, and OVA-specific IgE, eotaxin, transforming growth factor-beta1, airway inflammation, expression and metabolites of 12/15-LOX in lung cytosol, lipid peroxidation, and nitric oxide metabolites in the lung, restored the activity and expression of the third subunit of cytochrome-c oxidase in lung mitochondria and bronchial epithelia, respectively, reduced the appearance of cytochrome c in lung cytosol, and also restored mitochondrial ultrastructural changes of bronchial epithelia. In summary, these findings show that Vit-E reduces key mitochondrial dysfunctions and alleviates asthmatic features.

Topics: 12-Hydroxy-5,8,10,14-eicosatetraenoic Acid; Airway Remodeling; Animals; Anti-Asthmatic Agents; Arachidonate 12-Lipoxygenase; Arachidonate 15-Lipoxygenase; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Cytochromes c; Disease Models, Animal; Electron Transport Complex IV; Goblet Cells; Hyperplasia; Hypersensitivity; Immunoglobulin E; Interleukin-13; Interleukin-4; Interleukin-5; Linoleic Acids; Lung; Male; Mice; Mice, Inbred BALB C; Mitochondria; Ovalbumin; Oxidative Stress; Pulmonary Fibrosis; Transforming Growth Factor beta1; Vitamin E

The effects of remodeling of airway smooth muscle (SM) by hyperplasia on airway SM contractility in vivo are poorly explored. The aim of this study was to investigate the relationship between allergen-induced airway SM hyperplasia and its contractile phenotype. Brown Norway rats were sensitized with ovalbumin (OVA) or saline on day 0 and then either OVA-challenged once on day 14 and killed 24 h later or OVA-challenged 3 times (on days 14, 19, and 24) and killed 2 or 7 days later. Changes in SM mass, expression of total myosin, SM myosin heavy chain fast isoform (SM-B) and myosin light chain kinase (MLCK), tracheal contractions ex vivo, and airway responsiveness to methacholine (MCh) in vivo were assessed. One day after a single OVA challenge, the number of SM cells positive for PCNA was greater than for control animals, whereas the SM mass, contractile phenotype, and tracheal contractility were unchanged. Two days after three challenges, SM mass and PCNA immunoreactive cells were increased (3- and 10-fold, respectively; P < 0.05), but airway responsiveness to MCh was unaffected. Lower expression in total myosin, SM-B, and MLCK was observed at the mRNA level (P < 0.05), and total myosin and MLCK expression were lower at the protein level (P < 0.05) after normalization for SM mass. Normalized tracheal SM force generation was also significantly lower 2 days after repeated challenges (P < 0.05). Seven days after repeated challenges, features of remodeling were restored toward control levels. Allergen-induced hyperplasia of SM cells was associated with a loss of contractile phenotype, which was offset by the increase in mass.

Topics: Allergens; Animals; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Disease Models, Animal; Male; Methacholine Chloride; Muscle, Smooth; Ovalbumin; Rats; Rats, Inbred BN; Receptor, Muscarinic M3; Respiratory System; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

This study was aimed to analyse the curative effects of Pinellia ternata, Citrus reticulata, and their combination on airway hyperresponsiveness (AHR) to inhaled methacholine, pulmonary eosinophilic infiltration, Th2 cytokine production, and IgE and histamine production in a murine model of asthma.. For this purpose, BALB/c mice were systemically sensitized to ovalbumin (OVA) followed intratracheally, intraperitoneally, and by aerosol allergen challenges for 12 weeks. We examined the development of pulmonary eosinophilic accumulation, control of Th2 cytokine, immunoglobulin E (IgE), and histamine productions in a murine model of asthma.. Our data suggest that the therapeutic mechanism by which Pinellia ternata, Citrus reticulata, and their combinational prescription effectively treats asthma is based on reductions of eosinophil infiltration, eotaxin receptor (CCR3), histamine, OVA-specific IgE productions in serum, and Th2 cytokines (IL-5, IL-13) by marked reductions of IL-5 and IL-13 mRNA expression in lung tissue.. These findings provide evidence that Pinellia ternata, Citrus reticulata, and their combination play a regulatory role in allergic inflammation and offer therapeutic approaches as novel CCR3 antagonists for treatment asthma. However, it is not clear whether pharmacological activities of prescription composed of two herbs are potentiated due to synergistic effect or additive effect.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Citrus; Disease Models, Animal; Drug Therapy, Combination; Enzyme-Linked Immunosorbent Assay; Female; Immunohistochemistry; Interleukin-13; Interleukin-4; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Pinellia; Plant Extracts; Polymerase Chain Reaction; Receptors, CCR3; Th2 Cells

Immunologic processes might contribute to the pathogenesis of pulmonary arterial hypertension (PAH), a fatal condition characterized by progressive pulmonary arterial remodeling, increased pulmonary vascular resistance, and right ventricular failure. Experimental allergen-driven lung inflammation evoked morphologic and functional vascular changes that resembled those observed in patients with PAH. Sphingosine kinase 1 (SphK1) is the main pulmonary contributor to sphingosine-1-phosphate (S1P) synthesis, a modulator of immune and vascular functions.. We sought to investigate the role of SphK1 in allergen-induced lung inflammation.. SphK1-deficient mice and C57Bl/6 littermates (wild-type [WT] animals) were subjected to acute or chronic allergen exposure.. After 4 weeks of systemic ovalbumin sensitization and local airway challenge, airway responsiveness increased less in SphK1(-/-) compared with WT mice, whereas pulmonary vascular responsiveness was greatly increased and did not differ between strains. Acute lung inflammation led to an increase in eosinophils and mRNA expression for S1P phosphatase 2 and S1P lyase in lungs of WT but not SphK1(-/-) mice. After repetitive allergen exposure for 8 weeks, airway responsiveness was not augmented in SphK1(-/-) or WT mice, but pulmonary vascular responsiveness was increased in both strains, with significantly higher vascular responsiveness in SphK1(-/-) mice compared with that seen in WT mice. Increased vascular responsiveness was accompanied by remodeling of the small and intra-acinar arteries.. : The data support a role for SphK1 and S1P in allergen-induced airway inflammation. However, SphK1 deficiency increased pulmonary vascular hyperresponsiveness, which is a component of PAH pathobiology. Moreover, we show for the first time the dissociation between inflammation-induced remodeling of the airways and pulmonary vasculature.

Topics: Acute Disease; Allergens; Animals; Bronchial Hyperreactivity; Chronic Disease; Cytokines; Hypertension, Pulmonary; Lung; Lysophospholipids; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Phosphotransferases (Alcohol Group Acceptor); Pulmonary Artery; RNA, Messenger; Sphingosine

Jagged1, a Notch ligand, and Notch have been implicated in Th2 differentiation, but their role in initiating IL-4 production and Th2 differentiation in vivo and the development of allergic airway responses has not been defined. In this study, we show that Jagged1 is up-regulated on bone marrow-derived dendritic cells (BMDCs) pulsed with allergen and that the transfer of these BMDCs before allergen challenge induces airway hyperresponsiveness (AHR) and eosinophilic airway inflammation. Treatment of CD4(+) T cells with a gamma-secretase inhibitor (GSI), which inhibits Notch signaling, resulted in decreased cytokine production when the cells were cocultured with allergen-pulsed, Jagged1-expressing BMDCs and, after the transfer of allergen-pulsed BMDCs, IL-4-deficient (IL-4(-/-)) recipients of GSI-treated naive CD4(+) T cells developed lower levels of AHR, reduced numbers of eosinophils, and lower Th2 cytokine levels when challenged with allergen. In vivo treatment of wild-type mice with Jagged1-Fc enhanced AHR and airway inflammation, whereas the transfer of BMDC transfected with Jagged1 small interfering RNA (siRNA) cells into WT or IL-4(-/-) mice before transfer of CD4(+) T cells resulted in decreased AHR, inflammation, and Th2 cytokines, indicating the critical role for Jagged1 expression on APCs. These data identify the essential role of the interactions between Notch on CD4(+) T cells and Jagged1 on APCs in the initiation of IL-4 production and Th2 differentiation for the development of AHR and allergic airway inflammation.

Topics: Animals; Bronchial Hyperreactivity; Calcium-Binding Proteins; CD4-Positive T-Lymphocytes; Cells, Cultured; Coculture Techniques; Dendritic Cells; Eosinophilia; Female; Inflammation Mediators; Intercellular Signaling Peptides and Proteins; Interleukin-4; Jagged-1 Protein; Lung; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Receptors, Notch; Serrate-Jagged Proteins; Th2 Cells

Oral cysteinyl-leukotriene (LT) receptor antagonists such as montelukast are used for reducing airway inflammation and exacerbations. However, inhaled therapy using LT receptor antagonists has not been studied. In the present study, the effect of inhaled montelukast was investigated on airway hyperresponsiveness measured by cysteinyl-LT induced bronchoconstriction in an animal model of asthma. Bronchoconstriction responses were induced by inhaled LTC4 and LTD4 (0.2 microg/ml each) or three doses of intravenous LTC4 and LTD4 (0.3, 1, 3 microg/kg) in ovalbumin (OVA)-sensitized Hartley male guinea-pigs. The response was measured by the change in peak pressure of airway opening (Pao). The effect of montelukast was evaluated by the comparison of bronchoconstriction responses between the groups of animals pre-treated with 15-min inhalation of 10mg/ml montelukast and saline. To evaluate the tissue injury which might be caused by montelukast inhalation, lung tissues were examined for the histology. The broncoconstriction responses induced by inhaled LTC4 and LTD4 were enhanced by OVA sensitization in the guinea-pigs. In sensitized animals, the significant increases in peak Pao were 18.5+/-2.1 cmH(2)O by LTC4 inhalation and 25.0+/-1.6 cmH(2)O by LTD4 inhalation on average. Prior treatment of inhaled montelukast potently suppressed the peak Pao increases induced by both inhaled and intravenous LTC4 and LTD4 (all P<0.01 vs. saline control). Moreover, the suppression of inhaled montelukast against LTD4-induced bronchoconstriction was observed for at least up to 24h. According to the histological examination, montelukast inhalation produced no injury to the lung tissue. Inhaled montelukast, a cysteinyl-LT receptor antagonist, was effective in inhibiting cysteinyl-LT-induced acute bronchoconstriction, and may have the potential for clinical use as a new asthma drug.

Topics: Acetates; Administration, Inhalation; Animals; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Cyclopropanes; Cysteine; Disease Models, Animal; Guinea Pigs; Immunologic Factors; Leukotriene Antagonists; Leukotriene C4; Leukotriene D4; Leukotrienes; Lung; Male; Ovalbumin; Quinolines; Sulfides

Tumor necrosis factor-alpha (TNF-alpha) is a proinflammatory cytokine that has been implicated in many aspects of the airway pathology in asthma. TNF-alpha blocking strategies are now being tried in asthma patients. This study investigated whether TNF-alpha blocking therapy inhibits airway inflammation and airway hyperresponsiveness (AHR) in a mouse model of asthma. We also evaluated the effect of TNF-alpha blocking therapy on cytokine production and adhesion molecule expression.. Ovalbumin (OVA) sensitized BALB/c female mice were exposed to intranasal OVA administration on days 31, 33, 35, and 37. Mice were treated intraperitoneally with soluble TNF-alpha receptor (sTNFR) during the OVA challenge.. There were statistically significant decreases in the numbers of total cell and eosinophil in bronchoalveolar lavage fluid (BALF) in the sTNFR treated group compared with the OVA group. However, sTNFR-treatment did not significantly decrease AHR. Anti-inflammatory effect of sTNFR was accompanied with reduction of T helper 2 cytokine levels including interleukin (IL)-4, IL-5 and IL-13 in BALF and vascular cell adhesion molecule 1 expression in lung tissue.. These results suggest that sTNFR treatment can suppress the airway inflammation via regulation of Th2 cytokine production and adhesion molecule expression in bronchial asthma.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Blotting, Western; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Enzyme-Linked Immunosorbent Assay; Female; Immunohistochemistry; Inflammation; Interleukin-13; Interleukin-4; Interleukin-5; Mice; Mice, Inbred BALB C; Ovalbumin; Tumor Necrosis Factor-alpha

Asthma is a chronic inflammatory disease affecting the airways. Increased levels of T cells are found in the lungs after the induction of an allergic-like inflammation in rats, and flow cytometry studies have shown that these levels are reduced in CD26-deficient rats. However, the precise anatomical sites where these newly recruited T cells appear primarily are unknown. Therefore, we quantified the distribution of T cells in lung parenchyma as well as in large, medium and small airways using immunohistochemical stainings combined with morphometric analyses. The number of T cells increased after the induction of an allergic-like inflammation. However, the differences between CD26-deficient and wild-type rats were not attributable to different cell numbers in the lung parenchyma, but the medium- and large-sized bronchi revealed significantly fewer T cells in CD26-deficient rats. These sites of T cell recruitment were screened further using immunohistochemistry and quantitative real-time polymerase chain reaction with regard to two hypotheses: (i) involvement of the nervous system or (ii) expression of chemokines with properties of a T cell attractor. No topographical association was found between nerves and T cells, but a differential transcription of chemokines was revealed in bronchi and parenchyma. Thus, the site-specific recruitment of T cells appears to be a process mediated by chemokines rather than nerve-T cell interactions. In conclusion, this is the first report showing a differential site-specific recruitment of T cells to the bronchi in a CD26-deficient rat substrain during an asthma-like inflammation.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemotaxis, Leukocyte; Dipeptidyl Peptidase 4; Immunoglobulin E; Immunohistochemistry; Lung; Lymphocyte Count; Male; Models, Animal; Ovalbumin; Rats; Rats, Inbred F344; Rats, Mutant Strains; Reverse Transcriptase Polymerase Chain Reaction; Statistics, Nonparametric; T-Lymphocytes

The anti-inflammatory, anti-allergic, antioxidant properties of flavonoids are known in the respiratory tract. We are interested in the role of Provinol during an allergic inflammation of the airway.. The aim of this study was to examine the influence of an acute administration of Provinol on tracheal smooth muscle reactivity in guinea pigs and to assess the involvement of nitric oxide in the mechanism of Provinol action.. This experiment was performed 14 days after the sensitization of animals by ovalbumin. In vivo, the specific airway conductance, as a tracheal smooth muscle reactivity parameter in response to bronchoconstrictor histamine, was evaluated after peroral administration of Provinol alone or together with L-NAME (N(omega)-nitro-L-arginine methyl ester). In vitro, Provinol alone or in combination with L-NAME were added into an organ baths before the supplement of direct bronchoconstrictor histamine, acetylcholine and the allergen ovalbumin in rising concentrations. The amplitude of the tracheal smooth muscle contraction, as a tracheal smooth muscle reactivity parameter in response to histamine, acetylcholine and ovalbumin was evaluated.. Our results showed that a Provinol has significant bronchodilatory activities both in vivo and in vitro.. Provinol alleviated the contraction of tracheal smooth muscle in guinea pigs sumin. Nitric oxide plays an important role in the mechanism of Provinol action (Fig. 2, Ref. 28n.(Fig. 2, Ref. 28).

Topics: Allergens; Animals; Bronchial Hyperreactivity; Flavonoids; Guinea Pigs; Muscle Contraction; Ovalbumin; Phenols; Polyphenols; Trachea; Wine

The aim of this study was to evaluate the effects of inhaled KP-496, a novel dual antagonist for cysteinyl leukotriene receptor 1 and thromboxane A(2) receptor, on the allergic asthmatic responses in guinea pigs.. Actively sensitized animals were repeatedly exposed to antigen, and KP-496 (0.01 and 0.1%) was inhaled for 5 min before every antigen exposure. After evaluating the effects of KP-496 on asthmatic responses, such as immediate and late asthmatic response (IAR and LAR) and airway hyperresponsiveness (AHR), histopathological analyses of the lungs of asthmatic animals were made.. KP-496 significantly inhibited both antigen-induced LAR and AHR to acetylcholine, and slightly inhibited antigen-induced IAR. Furthermore, histopathological analyses of the lungs of the asthmatic animals demonstrated the following: (1) KP-496 suppressed infiltration of eosinophils around airway smooth muscle, (2) KP-496 suppressed airway epithelial hypertrophy, and (3) KP-496 suppressed increased mucus production in the airway.. In addition to suppression of LAR and AHR, our findings demonstrated that KP-496 inhibits features of airway inflammation. Since these broad ameliorative effects of KP-496 on asthmatic pathology are thought to result from the inhibition of multiple chemical mediators, KP-496 will be a potent agent in the treatment of bronchial asthma.

Topics: Acetylcholine; Administration, Inhalation; Animals; Anti-Asthmatic Agents; Asthma; Benzoates; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Guinea Pigs; Lung; Male; Ovalbumin; Pneumonia; Receptors, Leukotriene; Receptors, Thromboxane A2, Prostaglandin H2; Thiazoles

Allergic asthma is characterized by airway hyperresponsiveness (AHR) and allergic inflammation of the airways, driven by allergen-specific Th2 cells. The asthma phenotypes and especially AHR are sensitive to the presence and activity of regulatory T (Treg) cells in the lung. Glucocorticoid-induced tumor necrosis factor receptor (GITR) is known to have a co-stimulatory function on effector CD4+ T cells, rendering these cells insensitive to Treg suppression. However, the effects of GITR signaling on polarized Th1 and Th2 cell effector functions are not well-established. We sought to evaluate the effect of GITR signaling on fully differentiated Th1 and Th2 cells and to determine the effects of GITR activation at the time of allergen provocation on AHR and airway inflammation in a Th2-driven mouse model of asthma.. CD4+CD25- cells were polarized in vitro into Th1 and Th2 effector cells, and re-stimulated in the presence of GITR agonistic antibodies to assess the effect on IFNgamma and IL-4 production. To evaluate the effects of GITR stimulation on AHR and allergic inflammation in a mouse asthma model, BALB/c mice were sensitized to OVA followed by airway challenges in the presence or absence of GITR agonist antibodies.. GITR engagement potentiated cytokine release from CD3/CD28-stimulated Th2 but not Th1 cells in vitro. In the mouse asthma model, GITR triggering at the time of challenge induced enhanced airway hyperresponsiveness, serum IgE and ex vivo Th2 cytokine release, but did not increase BAL eosinophilia.. GITR exerts a differential effect on cytokine release of fully differentiated Th1 and Th2 cells in vitro, potentiating Th2 but not Th1 cytokine production. This effect on Th2 effector functions was also observed in vivo in our mouse model of asthma, resulting in enhanced AHR, serum IgE responses and Th2 cytokine production. This is the first report showing the effects of GITR activation on cytokine production by polarized primary Th1 and Th2 populations and the relevance of this pathway for AHR in mouse models for asthma. Our data provides crucial information on the mode of action of the GITR signaling, a pathway which is currently being considered for therapeutic intervention.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstriction; CD28 Antigens; CD3 Complex; Cells, Cultured; Disease Models, Animal; Glucocorticoid-Induced TNFR-Related Protein; Humans; Immunoglobulin E; Interferon-gamma; Interleukin-2 Receptor alpha Subunit; Interleukins; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Pulmonary Eosinophilia; Rats; Receptors, Nerve Growth Factor; Receptors, Tumor Necrosis Factor; Recombinant Proteins; Signal Transduction; Th1 Cells; Th2 Cells

Formaldehyde (FA) is a common indoor air pollutant that can cause asthma in people experiencing long-term exposure. While FA and other man-made chemicals contribute to the stimulation of asthma in the general population, the underlying molecular pathogenesis of this relationship is not yet well understood.. To explore FA as an irritant for the onset of asthma and as an adjuvant for the induction of allergy.. In the present study, 40 Wistar rats in five experimental groups were exposed to: (i) saline; (ii) ovalbumin (OVA); (iii) OVA + FA at 417 ppb; (iv) OVA + FA at 2500 ppb; and (v) FA at 2500 ppb. Current and prior occupational exposure limits in China were established at 417 ppb and 2500 ppb, respectively. Gaseous FA was administrated to the animals for 6 h/day before and during OVA immunization or saline treatment. Measured outcomes included in situ lung function analysis, cytokine measurement, and histological changes in the rat lungs.. The airway reactivity, lung histological changes, pulmonary interleukin-4 secretion, and eosinophil infiltration in the OVA and FA exposed rats were significantly higher after gaseous FA exposures of 417 and 2500 ppb. While FA exposure alone did not induce significant structural changes to the airway, and the rate of inflammatory cell infiltration was the same as for the control group, pulmonary levels of interferon-gamma were significantly elevated in the exposed rats.. FA may be an irritant as well as serve as an adjuvant for the onset of asthma or asthma-like symptoms.

Topics: Air Pollutants; Air Pollution, Indoor; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Dose-Response Relationship, Drug; Formaldehyde; Gases; Inhalation Exposure; Interferon-gamma; Interleukin-4; Irritants; Male; Ovalbumin; Pneumonia; Pulmonary Eosinophilia; Rats; Rats, Wistar; Time Factors

Osteopontin (OPN) is a secreted phosphoglycoprotein with a wide range of functions, and is involved in various pathophysiological conditions. However, the role of OPN in IgE and Th2-associated allergic responses remains incompletely defined. The aim of this study was to elucidate the role of OPN in systemic allergen sensitization in mice. When compared with OPN(+/+) mice, significantly increased levels of OVA-induced IgE were found in OPN(-/-) mice. OPN(-/-) DC demonstrated an increased capacity to enhance Th2 cytokine production in CD4+ T cells from sensitized OPN(+/+) mice. Furthermore, significantly reduced levels of IL-12p70 expression were seen in LPS-stimulated OPN(-/-) DC as compared with the WT DC, and the reduction was reversible by the addition of recombinant OPN (rOPN). rOPN was able to suppress OVA-induced IL-13 production in the cultures of CD4 and OPN(-/-) DC, but this inhibitory activity was neutralized by the addition of anti-IL-12 Ab. In addition, administration of rOPN in vivo suppressed OVA-specific IgE production; however, this suppressive effect was abrogated in IL-12-deficient mice. These results indicate that DC-derived OPN plays a regulatory role in the development of systemic allergen sensitization, which is mediated, at least in part, through the production of endogenous IL-12.

Topics: Adoptive Transfer; Allergens; Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Dendritic Cells; Enzyme-Linked Immunosorbent Assay; Immunoglobulin E; Immunoglobulin G; Interferon-gamma; Interleukin-12; Interleukin-13; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Osteopontin; Ovalbumin; Recombinant Proteins

Th17 cells play key roles in mediating autoimmunity, inflammation and mucosal host defense against pathogens. To determine whether naturally occurring Treg (nTreg) limit Th17-mediated pulmonary inflammation, OVA-specific CD4+ Th17 cells and expanded CD4+CD25+Foxp3+ nTreg were cotransferred into BALB/c mice that were then exposed to OVA aerosols. Th17 cells, when transferred alone, accumulated in the lungs and posterior mediastinal LN and evoked a pronounced airway hyperreactivity and neutrophilic inflammation, characterized by B-cell recruitment and elevated IgA and IgM levels. Cotransfer of antigen-specific nTreg markedly reduced the Th17-induced pulmonary inflammation and associated neutrophilia, B-cell influx and polymeric Ig levels in the airways, but did not inhibit airway hyperreactivity. Moreover, the regulation appeared restricted to the site of mucosal inflammation, since transfer of nTreg did not affect the Th17 response developing in the lung draining LN, as evidenced by unaltered levels of IL-17 production and low numbers of Foxp3+ Treg. Our findings suggest a crucial role for Th17 cells in mediating airway B-cell influx and IgA response, and demonstrate that antigen-specific nTreg suppress Th17-mediated lung inflammation. These results provide new insights into how Th17 responses are limited and may facilitate development of novel approaches for controlling Th17-induced inflammation.

Topics: Adoptive Transfer; Animals; Antigens; B-Lymphocytes; Bronchial Hyperreactivity; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Forkhead Transcription Factors; Immunoglobulin A; Immunoglobulin M; Interferon-gamma; Interleukin-17; Lung; Mice; Mice, Inbred BALB C; Mice, Transgenic; Neutrophils; Ovalbumin; Pneumonia; T-Lymphocytes, Helper-Inducer; T-Lymphocytes, Regulatory

It is widely established that mast cells (MCs) have a harmful role in asthma, for example by secreting various proinflammatory substances stored within their secretory granule. However, in this study, we show that one of the substances stored within MC granule, chymase, in fact has a protective role in allergic airway inflammation, indicating that MCs may possess both harmful and protective activities in connection with this type of disease. Wild-type (WT) mice and mice lacking mouse MC protease 4 (mMCP-4), a chymase that is functionally homologous to human chymase, were sensitized and challenged with OVA, followed by the assessment of airway physiology and inflammatory parameters. Our results show that the airway hyperresponsiveness was significantly higher in mMCP-4(-/-) as compared with WT mice. Moreover, the degree of lung tissue inflammation was markedly higher in mice lacking mMCP-4 than in WT controls. Histological analysis revealed that OVA sensitization/challenge resulted in a marked increased in the thickness of the smooth muscle cell (SMC) layer and, notably, that the degree of SMC layer thickening was more pronounced in mMCP-4(-/-) animals than in WT controls, thus indicating that chymase may have an effect on airway SMCs. In support of this, mMCP-4-positive MCs were located in the close vicinity of the SMC layer, mainly in the upper airways, and mMCP-4 was shown to be the major chymase expressed in these MCs. Taken together, our results indicate that chymase present in the upper airways protects against allergic airway responses, possibly by regulating SMCs.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Hypersensitivity; Inflammation; Lung; Mast Cells; Mice; Mice, Inbred C57BL; Mice, Knockout; Myocytes, Smooth Muscle; Ovalbumin; Serine Endopeptidases

Andrographolide from traditional Chinese herbal medicines previously showed it possesses a strong anti-inflammatory activity. In present study, we investigated whether Andrographolide could inhibit allergen-induced airway inflammation and airways hyper-responsiveness and explored the mechanism of Andrographolide on allergen-induced airway inflammation and airways hyper-responsiveness. After sensitized and challenged by ovalbumin, the BALB/c mice were administered intraperitoneally with Andrographolide. Hyper-responsiveness was recorded. The lung tissues were assessed by histological examinations. NF-kappaB in lung was determined by immunofluorescence staining and Western blotting. Treatment of mice with Androqrapholide displayed lower Penh in response to asthma group mice. After treatment with Andrographolide, the extent of inflammation and cellular infiltration in the airway were reduced. Andrographolide interrupted NF-kappaB to express in cell nucleus. The level of NF-kappaB expression was inhibited by Andrographolide. The data indicate that Andrographolide from traditional Chinese herbal medicines could inhibit extensive infiltration of inflammatory cells in lung and decrease airway hyperreactivity. Andrographolide could inhibit NF-kappaB expression in lung and suppress NF-kappaB expressed in the nucleus of airway epithelial cells.

Topics: Allergens; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchioles; Cell Nucleus; Diterpenes; Down-Regulation; Drugs, Chinese Herbal; Epithelium; Mice; NF-kappa B; Ovalbumin; Pulmonary Alveoli

Airway inflammation and airway hyperresponsiveness are central issues in the pathogenesis of asthma. CD69 is a membrane molecule transiently expressed on activated lymphocytes, and its selective expression in inflammatory infiltrates suggests that it plays a role in the pathogenesis of inflammatory diseases. In CD69-deficient mice, OVA-induced eosinophilic airway inflammation, mucus hyperproduction, and airway hyperresponsiveness were attenuated. Cell transfer of Ag-primed wild-type but not CD69-deficient CD4 T cells restored the induction of allergic inflammation in CD69-deficient mice, indicating a critical role of CD69 expressed on CD4 T cells. Th2 responses induced by CD69-deficient CD4 T cells in the lung were attenuated, and the migration of CD4 T cells into the asthmatic lung was severely compromised. The expression of VCAM-1 was also substantially altered, suggesting the involvement of VCAM-1 in the CD69-dependent migration of Th2 cells into the asthmatic lung. Interestingly, the administration of anti-CD69 Ab inhibited the induction of the OVA-induced airway inflammation and hyperresponsiveness. This inhibitory effect induced by the CD69 mAb was observed even after the airway challenge with OVA. These results indicate that CD69 plays a crucial role in the pathogenesis of allergen-induced eosinophilic airway inflammation and hyperresponsiveness and that CD69 could be a possible therapeutic target for asthmatic patients.

Topics: Allergens; Animals; Antibodies, Monoclonal; Antigens, CD; Antigens, Differentiation, T-Lymphocyte; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophils; Inflammation Mediators; Lectins, C-Type; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Respiratory Hypersensitivity

Some studies have suggested that the polyphenolic compounds might reduce the occurrence of asthma symptoms. The aim of our experiments was to evaluate the effects of 21 days of the flavonoid Flavin7 administration on experimentally induced airway inflammation in ovalbumin-sensitized guinea pigs. We assessed tracheal smooth muscle reactivity by an in vitro muscle-strip method; changes in airway resistance by an in vivo plethysmographic method; histological picture of tracheal tissue; and the levels of interleukin 4 (IL-4), and interleukin 5 (IL-5) in bronchoalveolar lavage fluid (BALF). Histological investigation of tracheal tissue and the concentrations of the inflammatory cytokines IL-4 and IL-5 in BALF were used as indices of airway inflammation. Administration of Flavin7 caused a significant decrease of specific airway resistance after histamine nebulization and a decline in tracheal smooth muscle contraction amplitude in response to bronchoconstricting mediators. Flavin7 minimized the degree of inflammation estimated on the basis of eosinophil calculation and IL-4 and IL-5 concentrations. In conclusion, administration of Flavin7 showed bronchodilating and anti-inflammatory effects on allergen-induced airway inflammation.

Topics: Animals; Bronchial Hyperreactivity; Cytokines; Flavonoids; Guinea Pigs; Male; Ovalbumin; Stilbenes

To study the relation between Respiratory Syncytial Virus infection and asthma development by measuring airway responsiveness (AR) and M2R function.. Guinea pigs (n = 34) were randomly divided into 4 groups: Hep-2/NS group (group A, n = 9), RSV/NS group (group B, n =9), Hep-2/OVA group (group C, n = 8) and RSV/OVA group(group D, n = 8). On day 21 after infection we tested AR and M2R. Then counted eosinophils in BALF and observed pathological change.. Intraairway pressure(IP mmH20) of group B had no significant difference with group A(P > 0.01), and the extent of IP decrease also had no difference between groups A and B (P > 0. 05), but IP of C group were much higher than group A (P<0.05), with extent of IP decrease lower than group A (P < 0.05). And IP of group D were higher than group C (P < 0.01), with the extent of IP decrease much lower than group C (P < 0.05).. RSV infection could enhance OVA-induced M2R dysfunction, then develop AHR.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Female; Guinea Pigs; Male; Ovalbumin; Random Allocation; Receptor, Muscarinic M2; Respiratory Syncytial Virus Infections; Respiratory Syncytial Viruses

M2-type pyruvate kinase (M2PK) was found to interact directly with the 'ITAM' region of the gamma chain of the high-affinity IgE receptor (FcvarepsilonRI). Our hypothesis was that mast cell degranulation might require the FcvarepsilonRI-mediated inhibition of M2PK activity.. In rat basophilic leukaemia (RBL-2H3) cells, the effects of directly inhibiting M2PK or preventing the FcvarepsilonRI-mediated inhibition of M2PK (disinhibition) on degranulation was measured by hexosaminidase release. Effects of blocking the FcvarepsilonRI-mediated inhibition of M2PK was also assessed in vivo in a mouse model of allergen-induced airway hyper-responsiveness.. Activation of FcvarepsilonRI in RBL-2H3 cells caused the rapid phosphorylation of tyrosine residues in M2PK, associated with a decrease in M2PK enzymatic activity. There was an inverse correlation between M2PK activity and mast cell degranulation. FcvarepsilonRI-mediated inhibition of M2PK involved Src kinase, phosphatidylinositol 3-kinase, PKC and calcium. Direct inhibition of M2PK potentiated FcvarepsilonRI-mediated degranulation and prevention of the FcvarepsilonRI-mediated inhibition of M2PK attenuated mast cell degranulation. Transfection of RBL-2H3 cells with M1PK which prevents FcvarepsilonRI-induced inhibition of M2PK, markedly reduced their degranulation and exogenous M1PK (i.p.) inhibited ovalbumin-induced airway hyper-responsiveness in vivo.. We have identified a new control point and a novel biochemical pathway in the process of mast cell degranulation. Our study suggests that the FcvarepsilonRI-mediated inhibition of M2PK is a crucial step in responses to allergens. Moreover, the manipulation of glycolytic processes and intermediates could provide novel strategies for the treatment of allergic diseases.

Topics: Animals; Bronchial Hyperreactivity; Calcium; Cell Degranulation; Cell Line, Tumor; Disease Models, Animal; Down-Regulation; Enzyme Inhibitors; Female; Glycolysis; Hexosaminidases; Male; Mast Cells; Mice; Mice, Inbred C57BL; Ovalbumin; Phosphatidylinositol 3-Kinases; Phosphorylation; Protein Kinase C; Pyruvate Kinase; Rats; Receptors, IgE; Signal Transduction; src-Family Kinases; Transfection

Pyrrolidine dithiocarbamate (PDTC) is known to exert anti-tumor and anti-inflammatory effects. However, the effects of PDTC against airway inflammation and its underlying mechanisms have not been reported. In the present study, we examined the protective effects of PDTC in a murine model of asthma induced by ovalbumin. PDTC reduced the number of infiltrating inflammatory cells in concert with reduced eosinophil peroxidase (EPO) activity in bronchoalveolar lavage fluid. In parallel, PDTC decreased airway hyperresponsiveness in a dose dependent manner. All these effects were correlated with heme oxygenase-1 (HO-1) mRNA and protein induction, and reversed by ZnPP, a HO-1 inhibitor. In addition, PDTC reduced the secretion of Th(2) cytokines such as IL-4 and IL-5, whereas ZnPP blocked the inhibitory effects of PDTC on Th(2) cytokine secretion. These results suggest that PDTC protects against airway inflammation at least in part via HO-1 induction, and that inhibitory action on Th(2) cytokines may be associated with the protective mechanism of PDTC.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Enzyme Induction; Eosinophil Peroxidase; Heme Oxygenase-1; Interleukin-4; Interleukin-5; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Protoporphyrins; Pyrrolidines; Thiocarbamates

Dendritic cells (DC) are important APCs that control allergen-induced airway responses by interacting directly with T cells. Leukotriene B(4) (LTB(4)), interacting with its high-affinity receptor, LTB(4) receptor 1 (BLT1), is known to attract and activate leukocytes during inflammation. We have previously shown that BLT1 expression on Ag-primed T cells is required for the development of airway hyperresponsiveness (AHR; Miyahara et al. 2005. Am. J. Respir. Crit. Care Med. 172: 161-167). However, the role for the LTB(4)-BLT1 pathway in DC function in allergen-induced airway responses has not been defined. Bone marrow-derived DCs (BMDC) were generated. Naive BALB/c mice received OVA-pulsed BLT1-deficient (BLT1(-/-)) BMDCs or wild-type BMDCs intratracheally and were then challenged with OVA for 3 days. Airway responses were monitored 48 h after the last allergen challenge. BLT1(-/-) BMDCs showed normal maturation judged from surface expression of CD markers. Compared with recipients of wild-type BMDCs, mice that received BLT1(-/-) BMDCs developed significantly lower AHR to inhaled methacholine, lower goblet cell metaplasia, and eosinophilic infiltration in the airways and decreased levels of Th2 type cytokines in the bronchoalveolar lavage fluid. Migration of BLT1(-/-) BMDCs into peribronchial lymph nodes was significantly impaired compared with BLT1(+/+) BMDCs after intratracheal instillation. These data suggest that BLT1 expression on DCs is required for migration of DCs to regional lymph nodes as well as in the development of AHR and airway inflammation.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cytokines; Dendritic Cells; Female; Leukotriene A4; Lung; Lymph Nodes; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Receptors, Leukotriene B4; Th2 Cells

Adenosine is formed from extracellular purines by ecto-5'-nucleotidase (CD73) and is an essential player in allergic airway inflammation. The contribution of adenosine and other purines to electrolyte transport and mucociliary clearance was studied in airways of allergen challenged mice. No signs for allergen-induced inflammation were found in CD73-/- mice, and adenosine monophosphate (AMP) was unable to elicit airway Cl(-) secretion in these animals. Tracheas of ovalbumin (OVA)-treated BALB/c and CD73+/+ mice were hyperresponsive towards methacholine when assessed by Penh and direct optical measurement of contraction. In addition Cl(-) secretion activated by ATP and ADP was enhanced. These changes were not observed in CD73-/- mice. Expression of CFTR or CLCA was unchanged upon OVA treatment of CD73 mice, suggesting enhanced Cl(-) secretion due to upregulated purinergic pathways. Mucociliary clearance was determined by measuring particle transport in excised mouse tracheas and was strongly enhanced in OVA-challenged CD73+/+ mice, but remained unchanged in CD73-/- mice. While mucociliary clearance is activated by allergen exposure independent of functional ecto-5'-nucleotidase, airway inflammation is largely dependent on CD73. Thus, ecto-5'-nucleotidase may provide a novel target for therapeutic intervention, probably by local application of ecto-5'-nucleotidase inhibitors through inhalation.

Topics: 5'-Nucleotidase; Adenine Nucleotides; Adenosine; Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Bronchoconstrictor Agents; Chlorides; Cystic Fibrosis Transmembrane Conductance Regulator; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mucociliary Clearance; Ovalbumin; Trachea

Studies of human asthma and of animal models of allergic inflammation/asthma highlight a crucial role for T(H)2 cells in the pathogenesis of allergic asthma. Repressor of GATA (ROG) is a POZ (BTB) domain-containing Kruppel-type zinc finger family (or POK family) repressor. A repressive function to GATA3, a master transcription factor for T(H)2 cell differentiation, is indicated.. The aim of this study was to clarify the regulatory roles of ROG in the pathogenesis of T(H)2-driven allergic diseases, such as allergic asthma.. We examined allergic airway inflammation and airway hyperresponsiveness (AHR) in 3 different mouse models, which use either ROG-deficient (ROG(-/-)) mice, ROG transgenic mice, or adoptive transfer of cells.. In ROG(-/-) mice T(H)2 cell differentiation, T(H)2 responses, eosinophilic airway inflammation, and AHR were enhanced. In ROG transgenic mice the levels of eosinophilic airway inflammation and AHR were dramatically reduced. Furthermore, adoptive transfer of T(H)2 cells with increased or decreased levels of ROG expression into the asthmatic mice resulted in reduced or enhanced airway inflammation, respectively.. These results indicate that ROG regulates allergic airway inflammation and AHR in a negative manner, and thus ROG might represent another potential therapeutic target for the treatment of asthmatic patients.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Eosinophils; GATA Transcription Factors; GATA3 Transcription Factor; Inflammation; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Repressor Proteins; Th2 Cells

The decreased L-arginine bioavailability, the basic substrate for nitric oxide synthesis, can be one of the factors contributing to the airways hyperreactivity.. We investigated the effect of various inhibitors of the enzyme activities utilizing L-arginine in a guinea pig model of experimental ovalbumin-induced airway hyperreactivity.. We used the in vivo pre-treatment with non-specific inhibitor of NO synthase No-nitro-L-arginine metylester (L-NAME) and relatively specific inhibitor of inducible NO synthase--aminoguanidine. Inhibitors were administered in one-shot (on the 14th day, 30 minutes before the inhalation of ovalbumin) or in a long-time regime (during the whole period of sensibilization by ovalbumin--14 days). We administered the inhibitor of arginase Nomega-hydroxy-L-arginine (NOHA) to the tracheal and lung tissue smooth muscle strips from sensibilized animals.. We observed an increase in the tracheal smooth muscle response to histamine in animals that received an inhalation dose of L-NAME (40 mg/kg b.w.) or aminoguanidine (50 mg/kg b.w.) 30 minutes before the inhalation of ovalbumin but did not evoke any significant difference in the reactivity of lung tissue smooth muscle. Tracheal smooth muscle responded with enhanced contraction amplitude to histamine after chronic pre-treatment with L-NAME or aminoguanidine. The inhibition of arginase with NOHA in vitro decreased the tracheal and lung tissue smooth muscle reactivity to histamine.. The results suggest that NO-synthase isoforms as well as arginase are involved in the production of NO and in the control of bronchomotoric tonus (Fig. 4, Tab. 2, Ref. 31). Full Text (Free, PDF) www.bmj.sk.

Topics: Allergens; Animals; Arginase; Arginine; Bronchial Hyperreactivity; Enzyme Inhibitors; Guanidines; Guinea Pigs; In Vitro Techniques; Lung; Male; Muscle Contraction; Muscle, Smooth; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase Type I; Ovalbumin; Trachea

Non-selective cation influx through canonical transient receptor potential channels (TRPCs) is thought to be an important event leading to airway inflammation. TRPC6 is highly expressed in the lung, but its role in allergic processes is still poorly understood.. The purpose of this study was to evaluate the role of TRPC6 in airway hyperresponsiveness (AHR) and allergic inflammation of the lung.. Methacholine-induced AHR was assessed by head-out body plethysmography of wild type (WT) and TRPC6(-/-) mice. Experimental airway inflammation was induced by intraperitoneal ovalbumin (OVA) sensitization, followed by OVA aerosol challenges. Allergic inflammation and mucus production were analysed 24 h after the last allergen challenge.. Methacholine-induced AHR and agonist-induced contractility of tracheal rings were increased in TRPC6(-/-) mice compared with WT mice, most probably due to compensatory up-regulation of TRPC3 in airway smooth muscle cells. Most interestingly, when compared with WT mice, TRPC6(-/-) mice exhibited reduced allergic responses after allergen challenge as evidenced by a decrease in airway eosinophilia and blood IgE levels, as well as decreased levels of T-helper type 2 (Th2) cytokines (IL-5, IL-13) in the bronchoalveolar lavage. However, lung mucus production after allergen challenge was not altered by TRPC6 deficiency.. TRPC6 deficiency inhibits specific allergic immune responses, pointing to an important immunological function of this cation channel in Th2 cells, eosinophils, mast cells and B cells.

Topics: Animals; Bronchial Hyperreactivity; Cells, Cultured; Epithelial Cells; Guinea Pigs; Hypersensitivity; Immunoglobulin E; In Vitro Techniques; Interleukin-13; Interleukin-5; Leukocytes; Lung; Mice; Mice, Knockout; Mucus; Muscle Contraction; Muscle, Smooth; Ovalbumin; Pulmonary Eosinophilia; Trachea; TRPC Cation Channels; TRPC6 Cation Channel

Sublingual immunotherapy (SLIT) has been established in humans as a safe and efficacious treatment for type I respiratory allergies.. In this study, we compared three Toll-like receptor (TLR) 2 ligands (Pam3CSK4, Porphyromonas gingivalis lipopolysaccharide and lipoteichoic acid) as potential adjuvants for sublingual allergy vaccines.. These molecules were tested in co-cultures of adjuvant-pre-treated dendritic cells (DCs) with murine naïve CD4(+) T lymphocytes. Patterns of cytokine production, phenotype, proliferation and gene expression were analysed by ELISA, cytofluorometry and quantitative PCR, respectively. TLR2 ligands were subsequently tested in a model of SLIT in BALB/c mice sensitized with ovalbumin (OVA).. Among the three TLR2 ligands tested, the synthetic lipopeptide Pam3CSK4 is the most potent inducer of IL-12p35 and IL-10 gene expression in murine bone marrow-derived DCs, as well as in purified oral myeloid DCs. Only Pam3CSK4-treated DCs induce IFN-gamma and IL-10 secretion by naïve CD4(+) T cells. Sublingual administration of Pam3CSK4 together with the antigen in BALB/c mice sensitized to OVA decreases airway hyperresponsiveness as well as OVA-specific T-helper type 2 (Th2) responses in cervical lymph nodes dramatically.. Pam3CSK4 induces Th1/regulatory T cell responses, and as such, is a valid candidate adjuvant for sublingual allergy vaccines.

Topics: Adjuvants, Immunologic; Administration, Sublingual; Animals; Antigen Presentation; Asthma; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cytokines; Dendritic Cells; Desensitization, Immunologic; Gene Expression; Humans; Interferon-gamma; Interleukin-10; Lipopeptides; Lipopolysaccharides; Lymph Nodes; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Ovalbumin; Porphyromonas gingivalis; T-Lymphocytes, Helper-Inducer; Teichoic Acids; Toll-Like Receptor 2

TLR ligands and other allergen-nonspecific immunostimulatory molecules are ubiquitous in ambient air and have profound modulatory activities in animal models of allergic asthma. However, several of these molecules have been shown to promote exaggerated Th2-biased airway hypersensitivity responses (AHRs), whereas others attenuate the asthmatic phenotype. Therefore, it has proven difficult to extrapolate experimental results with purified molecules toward a more general understanding of the allergen-nonspecific immunomodulatory influence of living environments on the natural history of allergic asthma. These investigations determined how regular and intermittent airway exposures to an unpurified, but sterile house dust extract standard (HDEst) affected the OVA-specific AHR and immune status of previously Th2-sensitized mice. Low-dose daily and high-dose intermittent HDEst exposures modulated ongoing AHRs considerably, reducing eosinophil recruitment and methacholine responsiveness, while increasing neutrophilic inflammation. However, only daily airway delivery of low-dose HDEst attenuated OVA-specific Th2 cytokine production and Th2-biased AHRs to allergen challenge 1 mo later. Finally, whereas LPS mimicked many of the immunomodulatory characteristics of HDEst in this murine asthma model, daily airway HDEst delivery was highly effective in attenuating the AHR of OVA/alum-sensitized TLR4-deficient mice. Taken together, these investigations provide direct evidence that living environments contain allergen-nonspecific immunostimulatory molecules that influence the airway hypersensitivity status of allergen-sensitized mice by TLR4-dependent and independent mechanisms.

Topics: Adjuvants, Immunologic; Allergens; Animals; Bronchial Hyperreactivity; Dust; Female; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Respiratory Hypersensitivity; Respiratory Mucosa; Signal Transduction; Th2 Cells; Toll-Like Receptor 4

Chronic aspiration associated with gastro oesophageal reflux disease (GERD) is thought to play a substantial role in the development of asthma, the incidence of which is dramatically increasing in industrially developed countries. The majority of data examining the association between aspiration and asthma has been obtained from epidemiological studies, which show that between 50 and 90% of individuals with asthma experience some element of GERD. This study describes the effect of chronic aspiration on a model of experimentally induced airway hypersensitivity in Balb/c mice.. Four experimental groups were utilized: Aspiration/Asthma, Sham/Asthma, Aspiration/Sham and Sham/Sham. Mice were sensitized with aerosolized 1% ovalbumin on days 1 to 10 (sensitization phase), followed by repeated exposure on days 31 to 40 (challenge phase). Aspiration events occurred on days 1, 8,15, 22, 29, 36, 43 and 50. Animals were sacrificed on days 56 and 57.. Chronic aspiration of 10 microL of murine gastric fluid per week for eight weeks produced an injury pattern distinct from that of acute aspiration, with lung injury characterized by hyperplasia, neutrophil infiltration of the bronchioles and relative parenchymal sparing. Aspiration during induction of ovalbumin-induced airway hypersensitivity was associated with a trend toward decreased production of antiovalbumin IgG, antiovalbumin IgE, and total IgE. Further, aspiration induced a substantial and significant increase in antiovalbumin IgG1/IgG2a ratios, consistent with a shift toward a predominantly Th2 response.. These findings indicate that chronic aspiration has a profound effect on the nature of the immune response to aerosolized allergens in a model of experimentally induced airway hypersensitivity.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Immunoglobulin E; Lung; Mice; Ovalbumin

The liver X receptors (LXRalpha/beta) are orphan nuclear receptors that are expressed in a large number of cell types and have been shown to have anti-inflammatory properties. Nuclear receptors have previously proved to be amenable targets for small molecular mass pharmacological agents in asthma, and so the effect of an LXR ligand was assessed in models of allergic airway inflammation. LXR agonist, GW 3965, was profiled in rat and mouse models of allergic asthma. In the Brown Norway rats, GW 3965 (3-30 mg/kg) was unable to reduce the bronchoalveolar lavage eosinophilia associated with this model and had no impact on inflammatory biomarkers (eotaxin and IL-1beta). The compound did significantly stimulate ABCA-1 (ATP-binding cassette A1) mRNA expression, indicating that there was adequate exposure/LXR activation. In the mouse model, the LXR ligand surprisingly increased airway reactivity, an effect that was apparent in both the Ag and nonchallenged groups. This increase was not associated with a change in lung tissue inflammation or number of mucus-containing cells. There was, however, a marked increase in airway smooth muscle thickness in both treated groups. We demonstrated an increase in contractile response to exogenous methacholine in isolated airways taken from LXR agonist-treated animals compared with the relevant control tissue. We corroborated these findings in a human system by demonstrating increased proliferation of cultured airway smooth muscle. This phenomenon, if evidenced in man, would indicate that LXR ligands may directly increase airway reactivity, which could be detrimental, especially in patients with existing respiratory disease and with already compromised lung function.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Benzoates; Benzylamines; Bronchial Hyperreactivity; Cell Proliferation; DNA-Binding Proteins; Dose-Response Relationship, Immunologic; Humans; Liver X Receptors; Male; Mice; Mice, Inbred BALB C; Muscle, Smooth; Orphan Nuclear Receptors; Ovalbumin; Rats; Rats, Inbred BN; Receptors, Cytoplasmic and Nuclear; Up-Regulation

Levels of endothelins are particularly high in the lung, and there is evidence that these peptides are involved in asthma. Asthma is a chronic inflammatory disease associated with lymphocyte infiltration. In the present study, we used a murine model of asthma to investigate the role of endothelins in lymphocyte and eosinophil infiltration into the airway hyperreactivity and mucus secretion. Sensitized C57Bl/6 mice were treated with endothelin ETA receptor antagonist (BQ123) or endothelin ETB receptor antagonist (BQ788) 30 min before an antigen aerosol challenge. After 24 h, dose response curves to methacholine were performed in isolated lungs, FACS analysis of lymphocytes and eosinophil counts were performed in bronchoalveolar lavage fluid and mucus index was determined by histopathology. In sensitized and antigen-challenged mice there is a marked increase in the T CD4+, T CD8+, B220+, Tgammadelta+ and NK1.1+ lymphocyte subsets. Treatment with BQ123 further increased these cell populations. The number of eosinophils, airway hyperreactivity and mucus were all reduced by BQ123 treatment. The BQ 788 had no significant effect on the parameters analyzed. Treatment with BQ123 reduced the endothelin concentration in lung homogenates, suggesting that endothelins exert a positive feedback on their synthesis. We show here that in murine asthma the ETA receptor antagonist up-regulates lymphocyte infiltration and reduces eosinophils, hyperreactivity and mucus.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Endothelin A Receptor Antagonists; Eosinophils; Lung; Lymphocyte Subsets; Male; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mucus; Ovalbumin; Peptides, Cyclic

We recently demonstrated the pivotal role of the transcription factor (TF) activating TF 3 (ATF3) in dampening inflammation. We demonstrate that ATF3 also ameliorates allergen-induced airway inflammation and hyperresponsiveness in a mouse model of human asthma. ATF3 expression was increased in the lungs of mice challenged with ovalbumin allergen, and this was associated with its recruitment to the promoters of genes encoding Th2-associated cytokines. ATF3-deficient mice developed significantly increased airway hyperresponsiveness, pulmonary eosinophilia, and enhanced chemokine and Th2 cytokine responses in lung tissue and in lung-derived CD4(+) lymphocytes. Although several TFs have been associated with enhanced inflammatory responses in the lung, ATF3 attenuates the inflammatory responses associated with allergic airway disease.

Topics: Activating Transcription Factor 3; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Chemokines; Gene Expression Regulation; Humans; Interleukin-13; Interleukin-4; Interleukin-5; Lung; Mice; Mice, Knockout; Ovalbumin; Pneumonia; Promoter Regions, Genetic; Pulmonary Eosinophilia; Th2 Cells

Asthma is characterized as a chronic inflammatory disorder of the airways. Phosphodiesterases (PDE), which hydrolyze cAMP, are considered to play important roles in asthma. We previously reported that acetamide-45 could inhibit cAMP-PDE activity, and histamine- and methacholine-induced contractions of isolated guinea pig trachea. The purpose of this study is to determine whether this agent could suppress allergic-induced airway hyperresponsiveness (AHR) and airway inflammation in allergic mice.. A mouse model for asthma was used to investigate acetamide-45 on the airway lesions compared with glucocorticoids. The study was conducted on mice sensitized and challenged with ovalbumin and the whole body plethysmography was carried out to assess AHR. The bronchoalveolar lavage (BAL) histopathology was examined.. We found that acetamide-45 significantly inhibited the enhanced hyperresponsiveness and eosinophil recruitment in airways with elimination of cAMP-PDE activity in lung tissue. Elevated IL-4 and IL-5 in bronchoalveolar lavage fluid (BALF) in asthmatic mice were markedly decreased.. Our results indicate that the agent has a potential role in inflammatory disease.

Topics: Acetamides; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Hypersensitivity; Indoles; Inflammation; Interferon-gamma; Interleukin-4; Interleukin-5; Leukocytes; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Phosphodiesterase Inhibitors

Statins have been proposed as a novel treatment of respiratory diseases. To determine the beneficial effects of statins on the airway hyperresponsiveness, a characteristic feature of allergic bronchial asthma, the effect of systemic treatment with lovastatin on antigen-induced bronchial smooth muscle hyperresponsiveness was investigated in mice. Male BALB/c mice were sensitized and repeatedly challenged with ovalbumin antigen. Animals were also treated with lovastatin (4 mg/kg/day, i.p.) once a day prior to and during the antigen inhalation period. The bronchial smooth muscle responsiveness to acetylcholine, but not to high K(+)-depolarization, was markedly and significantly augmented in the repeatedly antigen challenged mice. The bronchial smooth muscle hyperresponsiveness to acetylcholine induced by antigen exposure was significantly attenuated by the systemic treatment with lovastatin. Thus, lovastatin might have therapeutic potential to ameliorate airway hyperresponsiveness in allergic bronchial asthma.

Topics: Acetylcholine; Animals; Antigens; Asthma; Atropine; Bronchial Hyperreactivity; Bronchodilator Agents; Cholinergic Agents; Disease Models, Animal; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Indomethacin; Lovastatin; Male; Mice; Mice, Inbred BALB C; Muscle Contraction; Muscle, Smooth; Ovalbumin

Airway hypersensitive reaction (AHR) is an animal model for asthma, which is caused or enhanced by environmental factors such as allergen exposure. However, the precise mechanisms that drive AHR remain unclear. We identified a novel subset of natural killer T (NKT) cells that expresses the interleukin 17 receptor B (IL-17RB) for IL-25 (also known as IL-17E) and is essential for the induction of AHR. IL-17RB is preferentially expressed on a fraction of CD4(+) NKT cells but not on other splenic leukocyte populations tested. IL-17RB(+) CD4(+) NKT cells produce predominantly IL-13 and Th2 chemokines upon stimulation with IL-25 in vitro. IL-17RB(+) NKT cells were detected in the lung, and depletion of IL-17RB(+) NKT cells by IL-17RB-specific monoclonal antibodies or NKT cell-deficient Jalpha18(-/-) mice failed to develop IL-25-dependent AHR. Cell transfer of IL-17RB(+) but not IL-17RB(-) NKT cells into Jalpha18(-/-) mice also successfully reconstituted AHR induction. These results strongly suggest that IL-17RB(+) CD4(+) NKT cells play a crucial role in the pathogenesis of asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Humans; Immunoglobulin J-Chains; Interleukins; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Natural Killer T-Cells; Ovalbumin; Phenotype; Receptors, Interleukin-17; T-Lymphocyte Subsets

The pharmacological profiles of antigen-induced immediate airway response (IAR) in rats are not fully understood. In this study, we established an ovalbumin (OVA)-induced IAR model using noninvasive measurement in rats, and evaluated the effects of commonly used and effective antiasthmatic drugs, i.e. ketotifen (antihistamine), pranlukast (anti-leukotriene C(4)/D(4)/E(4) (LT)), seratrodast (anti-thromboxane A(2) (TXA(2))), salbutamol (beta2-agonist), and prednisolone (steroid). The rat IAR model exhibited an optimal rapid airway response, and salbutamol inhalation completely suppressed the IAR. Ketotifen inhibited only the quick phase (QP; the reaction from 3 to 6 min after challenge), while pranlukast and seratrodast suppressed only the early phase (EP; the reaction from 6 to 30 min after challenge). Prednisolone inhibited both QP and EP. Further, continuous administration of compound 48/80, which depletes connective tissue mast cells (CTMC), partially inhibited QP but not EP. In conclusion, these findings suggest that the pharmacological profiles of noninvasive rat IAR are similar to those of asthmatic patients, and that rat IAR exhibits additional, immunological diverse characteristics, i.e. QP caused by the exocytosis of mediators in CTMCs and EP mediated by LT and TXA(2), which are produced by mucosal mast cells (MMCs) and possibly by other types of cells. This is the first report about the comprehensive pharmacological profiles of rodent IAR model, and these analyses of rat IAR model may help expand our understanding of the diverse mechanisms underlying human asthmatic diseases.

Topics: Administration, Inhalation; Adrenergic alpha-2 Receptor Agonists; Adrenergic beta-Agonists; Albuterol; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents, Non-Steroidal; Antigens; Benzoquinones; Bronchial Hyperreactivity; Chromones; Heptanoic Acids; Histamine H2 Antagonists; Hypersensitivity, Immediate; Ketotifen; Leukotriene Antagonists; Male; Ovalbumin; p-Methoxy-N-methylphenethylamine; Prednisolone; Rats; Rats, Sprague-Dawley; Thromboxane A2

Asthma is a complex disease characterized by airway hyperresponsiveness (AHR) and chronic airway inflammation. Epidemiologic studies have demonstrated that exposures to environmental factors such as ambient particulate matter (PM), a major air pollutant, contribute to increased asthma prevalence and exacerbations.. We investigated pathophysiologic responses to Baltimore, Maryland, ambient PM (median diameter, 1.78 mum) in a murine model of asthma and attempted to identify PM-specific genomic/molecular signatures.. We exposed ovalbumin (OVA)-sensitized A/J mice intratracheally to PM (20 mg/kg), and assayed both AHR and bronchoalveolar lavage (BAL) on days 1, 4, and 7 after PM exposure. Lung gene expression profiling was analyzed in OVA- and PM-challenged mice.. Consistent with this murine model of asthma, we observed significant increases in airway responsiveness in OVA-treated mice, with PM exposure inducing significant changes in AHR in both naive mice and OVA-induced asthmatic mice. PM evoked eosinophil and neutrophil infiltration into airways, elevated BAL protein content, and stimulated secretion of type 1 T helper (T(H)1) cytokines [interferon-gamma, interleukin-6 (IL-6), tumor necrosis factor-alpha] and T(H)2 cytokines (IL-4, IL-5, eotaxin) into murine airways. Furthermore, PM consistently induced expression of genes involved in innate immune responses, chemotaxis, and complement system pathways.. This study is consistent with emerging epidemiologic evidence and indicates that PM exposure evokes proinflammatory and allergic molecular signatures that may directly contribute to the asthma susceptibility in naive subjects and increased severity in affected asthmatics.

Topics: Air Pollutants; Animals; Bronchial Hyperreactivity; Genomics; Lung; Male; Mice; Ovalbumin; Particle Size

Kefiran is a major component of kefir which is a microbial symbiont mixture that produces jelly-like grains. This study aimed to evaluate the therapeutic availability of kefiran on the ovalbumin-induced asthma mouse model in which airway inflammation and airway hyper-responsiveness were found in the lung. BALB/c mice sensitized and challenged to ovalbumin were treated intra-gastrically with kefiran 1 hour before the ovalbumin challenge. Kefiran significantly suppressed ovalbumin-induced airway hyper-responsiveness (AHR) to inhaled methacholine. Administration of kefiran significantly inhibited the release of both eosinophils and other inflammatory cells into bronchoalveolar lavage (BAL) fluid and lung tissue which was measured by Diff-Quik. Interleukin-4 (IL-4) and interleukin-5 (IL-5) were also reduced to normal levels after administration of kefiran in BAL fluid. Histological studies demonstrate that kefiran substantially inhibited ovalbumin-induced eosinophilia in lung tissue by H&E staining and goblet cell hyperplasia in the airway by PAS staining. Taken above data, kefiran may be useful for the treatment of inflammation of lung tissue and airway hyper-responsiveness in a murine model and may have therapeutic potential for the treatment of allergic bronchial asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cytokines; Female; Interleukin-4; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Pneumonia; Polysaccharides; Th2 Cells

As the administration of many antitussive drugs is often associated with adverse effects, new alternatives are evaluated in experimental and clinical conditions. The aim of this study was to assess the influence of selective inhibitors of PDE3 (cilostazol) and PDE4 (citalopram) on cough and airway reactivity. The number of cough efforts, specific airway resistance, in vitro airway reactivity, and differential blood cells count were measured in healthy and in ovalbumin-sensitized guinea pigs before and after administration of cilostazol or citalopram (1 mg/kg). Cilostazol significantly suppressed citric acid induced cough only in healthy guinea pigs, whereas citalopram in both healthy and ovalbumin-sensitized animals. Both PDE inhibitors decreased in vivo and in vitro airway reactivity to histamine and the count of monocytes and neutrophils, confirming their anti-inflammatory potential. Administration of selective PDE3 and PDE4 inhibitors may influence cough and airway reactivity in the model of ovalbumin-sensitized guinea pigs.

Topics: Animals; Antigens; Antitussive Agents; Blood Cell Count; Bronchial Hyperreactivity; Bronchoconstriction; Cilostazol; Citalopram; Cough; Guinea Pigs; Histamine Release; Leukocyte Count; Male; Ovalbumin; Phosphodiesterase 3 Inhibitors; Phosphodiesterase 4 Inhibitors; Phosphodiesterase Inhibitors; Tetrazoles

Corticosteroids are known to inhibit bronchial hyperresponsiveness (BHR) and allergic inflammation but there is little information on its dose-dependence. We examined the effect of different doses of the glucocorticosteroid budesonide in an allergic model. Brown-Norway rats were sensitised to ovalbumin (OVA) and pretreated with an intra-gastric dose of budesonide (0.1, 1.0, or 10 mgkg(-1)). Exposure to OVA induced BHR, accumulation of eosinophils in the bronchoalveolar lavage (BAL) fluid and in the airways submucosa. Budesonide dose-dependently inhibited BAL fluid influx of lymphocytes, eosinophils and neutrophils, tissue eosinophils and lymphocytes and BHR. At 0.1 mgkg(-1), budesonide did not inhibit these parameters but at 1 mgkg(-1), BAL fluid eosinophils and T-cells, and submucosal T-cells were significantly reduced. At 10 mgkg(-1), budesonide suppressed BHR, BAL fluid inflammatory cells numbers and tissue eosinophilia. T-cell numbers were more related to BHR than eosinophil numbers. Budesonide inhibited both airway inflammation and BHR, but BAL fluid eosinophil cell counts may be dissociated from BHR.

Topics: Acetylcholine; Animals; Blood Cell Count; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Budesonide; Disease Models, Animal; Dose-Response Relationship, Drug; Glucocorticoids; Hypersensitivity; Male; Ovalbumin; Rats

Ecklonia cava (EC) is a brown alga that evidences radical scavenging activity, bactericidal activity, tyrosinase inhibitory activity, and protease inhibitory activity. However, its anti-allergic effects remain poorly understood. In the current study, we attempted to determine whether pretreatment with EC induces a significant inhibition of asthmatic reactions in a mouse asthma model. Mice sensitized and challenged with ovalbumin (OVA) evidenced typical asthmatic reactions, as follows: an increase in the number of eosinophils in bronchoalveolar lavage fluid; a marked influx of inflammatory cells into the lung around blood vessels and airways, and airway luminal narrowing; the development of airway hyperresponsiveness; the detection of tumor necrosis factor-alpha (TNF-alpha) and Th2 cytokines, including IL-4 and IL-5 in the bronchoalveolar lavage (BAL) fluid; and the detection of allergen-specific immunoglobulin E (IgE) in the serum. However, the administration of EC extract prior to the final airway OVA challenge resulted in a significant inhibition of all asthmatic reactions. We also demonstrated that EC extracts treatment resulted in significant reductions on matrix metalloproteinase-9 (MMP-9) and Suppressor of cytokine signaling-3 (SOCS-3) expression and a reduction in the increased eosinophil peroxidase (EPO) activity. The treatment of animals with EC extracts resulted in a significant reduction in the concentrations of the Th2 cytokine (IL-4 and IL-5) in the airways, without any concomitant increase in the concentration of Th1 cytokines. These findings indicate that EC extracts may prove useful as an adjuvant therapy for allergic airway reactions via the inhibition of the Th2 response. Accordingly, this study may provide evidence that EC extract performs a critical function in the amelioration of the pathogenetic process of asthma in mice.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cytokines; Eosinophil Peroxidase; Ethanol; Female; Immunoglobulin E; Matrix Metalloproteinase 3; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Phaeophyceae; Phytotherapy; Plant Extracts; Solvents; Suppressor of Cytokine Signaling 3 Protein; Suppressor of Cytokine Signaling Proteins

The alpha-2 adrenergic agonists clonidine and dexmedetomidine are used as an antihypertensive and a sedative, respectively. The aim of this study was to determine the effects of these agonists on ovalbumin-sensitized airway tone in guinea pigs.. The animals were divided into two groups: control and sensitized. The sensitized group received ovalbumin intraperitoneally and was boosted by exposure to aerosolized ovalbumin. The effects of the alpha-2 agonists were investigated by measuring (1) total lung resistance and (2) smooth muscle tension using a tracheal ring preparation.. In the control group, acetylcholine significantly increased total lung resistance in a dose-dependent manner. In the sensitized animals, total lung resistance was significantly higher (by 95%) at 6 mug kg-1 acetylcholine than that in the control group. Both clonidine and dexmedetomidine had a slight but significant inhibitory effect on the response curve of lung resistance at higher concentrations of carbachol, a potent muscarinic receptor agonist. Similar to the data obtained in the control group, both clonidine and dexmedetomidine significantly decreased total lung resistance and the inhibitory effects of these alpha-2 agonists on lung resistance were significantly distinguishable. Similar direct inhibitory effects of the alpha-2 agonists on carbachol-induced muscle contraction were observed in both the control and sensitized groups, the inhibitory effects in the sensitized group being significantly greater.. Both clonidine and dexmedetomidine can relax the airway even in the hyper-reactive state.

Topics: Acetylcholine; Adrenergic alpha-Agonists; Airway Obstruction; Animals; Bronchial Hyperreactivity; Carbachol; Clonidine; Dexmedetomidine; Guinea Pigs; Hypnotics and Sedatives; Lung; Male; Muscle, Smooth; Ovalbumin; Specific Pathogen-Free Organisms; Trachea

Previously, we found pulmonary gas trapping to be a rapid, simple and objective measure of methacholine-induced airway obstruction in naïve mice. In this study we extended that finding by using methacholine-induced pulmonary gas trapping to differentiate airway responses of ovalbumin-sensitized, ovalbumin-exposed (Positive Control) and ovalbumin-sensitized, sodium chloride-exposed (Negative Control) mice. Additionally, pulmonary gas trapping and enhanced pause were compared following methacholine exposure in sensitized and nonsensitized mice. Finally, we examined by nose-only inhalation the ability of the glucocorticosteroid budesonide and the peroxisome proliferator-activated receptor-gamma agonist ciglitazone to modify methacholine-induced airway responses in ovalbumin-sensitized mice. Positive Controls exhibited a 7.8-fold increase in sensitivity and a 2.4-fold enhancement in the maximal airway obstruction to methacholine versus Negative Controls. Following methacholine, individual Positive and Negative Control mouse enhanced pause values overlapped in 9 of 9 studies, whereas individual Positive and Negative Control mouse excised lung gas volume values overlapped in only 1 of 9 studies, and log[excised lung gas volume] correlated (P=0.023) with in vivo log[enhanced pause] in nonsensitized mice. Finally, budesonide (100.0 or 1000.0 microg/kg) reduced methacholine-mediated airway responses and eosinophils and neutrophils, whereas ciglitazone (1000.0 microg/kg) had no effect on methacholine-induced pulmonary gas trapping, but reduced eosinophils. In conclusion, pulmonary gas trapping is a more reproducible measure of methacholine-mediated airway responses in ovalbumin-sensitized mice than enhanced pause. Also, excised lung gas volume changes can be used to monitor drug interventions like budesonide. Finally, this study highlights the importance of running a positive comparator when examining novel treatments like ciglitazone.

Topics: Administration, Inhalation; Airway Obstruction; Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchodilator Agents; Budesonide; Disease Models, Animal; Dose-Response Relationship, Drug; Eosinophils; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; PPAR gamma; Thiazolidinediones

Asthma is a heterogeneous disease that has been increasing in incidence throughout western societies and cytokines, including proinflammatory tumour necrosis factor alpha (TNF-alpha), have been implicated in the pathogenesis of asthma. Anti-TNF-alpha therapies have been established successfully in the clinic for diseases such as rheumatoid arthritis and Crohn's disease. TNF-alpha-blocking strategies are now being trialled in asthma; however, their mode of action is poorly understood. Based on the observation that TNF-alpha induces lymph node hypertrophy we have attempted to investigate this as a mechanism of action of TNF-alpha in airway inflammation by employing two models of murine airway inflammation, that we have termed short and long models, representing severe and mild/moderate asthma, respectively. The models differ by their immunization schedules. In the short model, characterized by eosinophilic and neutrophilic airway inflammation the effect of TNF-alpha blockade was a reduction in draining lymph node (DLN) hypertrophy, eosinophilia, interleukin (IL)-5 production and immunoglobulin E (IgE) production. In the long model, characterized by eosinophilic inflammation, TNF-alpha blockade produced a reduction in DLN hypertrophy and IL-5 production but had limited effects on eosinophilia and IgE production. These results indicate that anti-TNF-alpha can suppress DLN hypertrophy and decrease airway inflammation. Further investigations showed that anti-TNF-alpha-induced inhibition of DLN hypertrophy cannot be explained by preventing l-selectin-dependent capture of lymphocytes into the DLN. Given that overall TNF blockade was able to suppress the short model (severe) more effectively than the long model (mild/moderate), the results suggest that TNF-alpha blocking therapies may be more effective in the treatment of severe asthma.

Topics: Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Eosinophilia; Etanercept; Flow Cytometry; Hypertrophy; Immunoglobulin G; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Models, Animal; Ovalbumin; Receptors, Tumor Necrosis Factor; T-Lymphocytes; Time; Tumor Necrosis Factor-alpha

Somatostatin released from activated capsaicin-sensitive afferents of the lung inhibits inflammation and related bronchial hyperreactivity presumably via somatostatin 4 receptors (sst(4)). The aim of this study was to examine the effects of TT-232, a heptapeptide sst(4)/sst(1) receptor agonist and J-2156, a high affinity sst(4) receptor-selective peptidomimetic agonist in airway inflammation models. Acute pneumonitis was evoked by intranasal lipopolysaccharide 24 h before measurement. Chronic inflammation was induced by ovalbumin inhalation on days 28, 29 and 30 after i.p. sensitization on days 1 and 14. Semiquantitative histopathological scoring was based on perivascular/peribronchial oedema, neutrophil/macrophage infiltration, goblet cell hyperplasia in the acute model and eosinophil infiltration, mucosal oedema, mucus production and epithelial cell damage in chronic inflammation. Myeloperoxidase activity of the lung was measured spectrophotometrically to quantify granulocyte accumulation and the broncoalveolar lavage fluid was analysed by flow cytometry. Carbachol-induced bronchoconstriction was assessed by unrestrained whole body plethysmography and its calculated indicator, enhanced pause (Penh) was determined. TT-232 and J-2156 induced similar inhibition on granulocyte recruitment and histopathological changes in both models, although macrophage infiltration in LPS-induced inflammation was unaltered by either compounds. Both agonists diminished inflammatory airway hyperresponsiveness. Since their single administration after the development of the inflammatory reactions also inhibited carbachol-induced bronchoconstriction, somatostatin sst(4) receptor activation on bronchial smooth muscle cells is likely to be involved in their anti-hyperreactivity effect. These results suggest that stable, somatostatin sst(4) receptor-selective agonists could be potential candidates for the development of a completely novel group of anti-inflammatory drugs for the treatment of airway inflammation and hyperresponsiveness.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Butanes; Carbachol; Cells, Cultured; Female; Interleukin-1beta; Lipopolysaccharides; Lung; Macrophages, Peritoneal; Membrane Proteins; Mice; Mice, Inbred BALB C; Naphthalenes; Ovalbumin; Peptides, Cyclic; Pneumonia, Bacterial; Receptors, Somatostatin; Respiratory System Agents; Somatostatin; Sulfones; Tetradecanoylphorbol Acetate; Time Factors

Airways hyperresponsiveness (AHR) is a hallmark feature of asthma, and can be caused by various disparate mechanisms. Mouse models of AHR have been useful for studying these mechanisms in isolation, but such models still typically do not exhibit the same degree of AHR as seen in severe human asthma. We hypothesized that more severe AHR in mice could be achieved by imbuing them with more than one mechanism of AHR.. We sought to determine if the airway wall thickening accompanying allergic inflammation and the exaggerated smooth muscle shortening induced by intratracheal cationic protein could act together to produce a severe form of AHR.. We used the forced oscillation technique to measure methacholine responsiveness in BALB/c mice that had been sensitized and challenged with ovalbumin followed by an intratracheal instillation of poly-l-lysine.. We found that both ovalbumin and poly-l-lysine treatment alone caused moderate levels of AHR. When the two treatments were combined, however, they synergized in terms of their effect on lung stiffness to an extent that could even be fatal, reflecting a significantly enhanced level of airway closure.. Our results suggest that mechanistic synergy between airway wall thickening and exaggerated smooth muscle shortening produces a more germane mouse model of asthma that may have particular relevance to the pathophysiology of the acute severe asthma exacerbation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Bronchoconstrictor Agents; Disease Models, Animal; Drug Synergism; Female; Mice; Muscle, Smooth; Ovalbumin; Polylysine

YM-58483/BTP-2, 4-methyl-4'-[3,5-bis(trifluoromethyl)-1H-pyrazol-1-yl]-1,2,3-thiadiazole-5-carboxanilide, blocks the store-operated Ca2+ entry (SOCE) that mediates the activation of non-excitable cells. This study investigated the pharmacological profile and therapeutic potential of YM-58483 as anti-asthma drug. YM-58483 inhibited DNP antigen-induced histamine release from and leukotrienes (LTs) production in IgE-primed RBL-2H3 cells, a rat basophilic leukemia cell line, with IC50 values of 460 and 310 nM, respectively. Prednisolone did not inhibit either of these responses. YM-58483 also inhibited phytohemagglutinin-P (PHA)-stimulated IL-5 and IL-13 production in human peripheral blood cells with IC50 values of 125 and 148 nM, respectively, which is approximately 5 times less potent than prednisolone. YM-58483 (30 mg/kg, p.o.) significantly suppressed ovalbumin (OVA)-induced bronchoconstriction in OVA-sensitized guinea pigs, whereas prednisolone did not. YM-58483 (3-30 mg/kg, p.o.) and prednisolone (100mg/kg, p.o.) both significantly and completely suppressed airway hyperresponsiveness (AHR) caused by OVA exposure. Since YM-58483 inhibits two major characteristic symptoms of bronchial asthma, namely bronchoconstriction and AHR via the suppression of inflammatory mediator and cytokine production, SOCE inhibition is a potential approach for treatment.

Topics: Anilides; Animals; Anti-Inflammatory Agents; Blood Cells; Bronchial Hyperreactivity; Bronchoconstriction; Calcium; Calcium Channel Blockers; Cell Line; Dinitrophenols; Dose-Response Relationship, Drug; Guinea Pigs; Histamine Release; Humans; In Vitro Techniques; Leukotrienes; Male; Muscle Contraction; Muscle, Smooth; Ovalbumin; Phytohemagglutinins; Prednisolone; Rats; Serum Albumin, Bovine; Thiadiazoles; Trachea

To clarify whether growth factors play critical roles in the development of airway hyperresponsiveness (AHR) and airway inflammation in the early stages of asthma, the relationship between growth factors and AHR and airway inflammation were analyzed in a mouse model of asthma.. Following ovalbumin (OVA) sensitization and challenge, airway function, inflammation, cytokine and growth factor levels were monitored.. AHR to inhaled methacholine increased at 6 h, peaked at 48 h, and remained elevated for 14 days. IL-4 and IL-5 levels in bronchoalveolar lavage (BAL) fluid were increased at 6 h, peaked at 24 h, but returned to baseline quickly. IL-13 levels increased up to 14 days, peaking at 48 h. Increases in BAL fluid transforming growth factor-beta(1) and platelet-derived growth factor were observed at 12 h, and remained elevated at 14 days. Nerve growth factor levels were increased at 24-28 days. BAL fluid hepatocyte growth factor (HGF) was detected at 12 h, peaked at 24 h, and returned to baseline by 72 h. c-Met/HGF receptor was detected in the airways at 6 h, before HGF in the BAL, and continued to be observed 96 h after the last OVA challenge.. These data identify a temporal association between growth factor production and Th2 cytokine production and the kinetics of AHR. Growth factors may play important roles in the development of allergic airway inflammation and AHR even in the early stages of asthma, before remodeling is initiated.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Female; Hepatocyte Growth Factor; Inflammation Mediators; Intercellular Signaling Peptides and Proteins; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Time Factors; Transforming Growth Factor beta

Asthma is characterised by an excessive airway narrowing in response to a variety of stimuli, called airway hyperresponsiveness (AHR). Previous comparisons between mouse strains have shown that increased velocity of airway narrowing correlates with baseline airway responsiveness. These data prompted the investigation into models of induced AHR to see whether airway narrowing dynamics correlated with in vivo responsiveness. In an attempt to reproduce some of the features of asthma, BALB/c mice were sensitised and subjected to either brief or chronic periods of allergen exposure. Brief exposure involved two challenges with intranasal chicken egg ovalbumin (OVA(in)). Chronic exposure involved six 2-day periods of OVA(in) challenges, each separated by 12 days. Control mice received intranasal saline challenges. Outcomes included videomicrometry of lung slices (magnitude and velocity of airway narrowing), in vivo respiratory physiology measurements and histological staining with morphometric analysis. Neither brief nor chronic allergen exposure resulted in greater airway narrowing and increased velocity compared with saline controls. Structural changes in the airway, such as goblet cell hyperplasia, subepithelial fibrosis and increased contractile tissue, were detected in mice chronically challenged with allergen. In conclusion, increased responsiveness to methacholine following allergen challenge may not be due to an intrinsic change to the smooth muscle per se, but rather to other changes in the lung, which ultimately manifest as an increase in respiratory resistance.

Topics: Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstrictor Agents; Disease Models, Animal; Image Processing, Computer-Assisted; Methacholine Chloride; Mice; Mice, Inbred BALB C; Microscopy, Video; Muscle, Smooth; Ovalbumin

Live BCG administered intranasally to mice inhibits the development of ovalbumin (OVA)-induced eosinophilia and airway hyperresponsiveness (AHR). It is unacceptable to treat human subjects intranasally with live BCG.. We investigated whether BCG killed by extended freeze-drying (EFD) and subcutaneously injected has a protective effect in murine and guinea pig models of allergic airway inflammation.. Mice were OVA sensitized (days 0 and 7), treated subcutaneously (day 14) with EFD and live or heat-killed BCG, and then OVA challenged (day 42). OVA-sensitized mice (days 0 and 7) were challenged (day 14) and EFD treated (day 18) before OVA rechallenge (day 46) to demonstrate the capacity of EFD to reverse the established lung inflammation. Guinea pigs were OVA sensitized (days 0 and 14), treated intradermally (day 35) with EFD, and OVA challenged (days 90-105).. In mice and guinea pigs EFD treatment reduced AHR. Among 3 BCG preparations, only EFD efficiently reduced AHR, eosinophilia, and the recruitment of dendritic cells to the lungs after OVA challenge. The protective effect of EFD is associated with production of the immunoregulatory cytokine IL-10. Moreover, EFD treatment did not induce toxic effects or delayed-type hypersensitivity to mycobacterial antigens; that is, it did not interfere with the diagnosis of tuberculosis.. EFD administered subcutaneously inhibits the development of allergic airway inflammation and prevents AHR without inducing delayed-type hypersensitivity and side effects associated with live or heat-killed BCG.

Topics: Animals; BCG Vaccine; Bronchial Hyperreactivity; Bronchitis; Dendritic Cells; Eosinophilia; Freeze Drying; Guinea Pigs; Hypersensitivity; Injections, Subcutaneous; Interleukin-10; Lung; Male; Mice; Ovalbumin; Pneumonia; Vaccines, Inactivated

The female hormone estrogen is an important factor in the regulation of airway function and inflammation, and sex differences in the prevalence of asthma are well described. Using an animal model, we determined how sex differences may underlie the development of altered airway function in response to allergen exposure. We compared sex differences in the development of airway hyperresponsiveness (AHR) after allergen exposure exclusively via the airways. Ovalbumin (OVA) was administered by nebulization on 10 consecutive days in BALB/c mice. After methacholine challenge, significant AHR developed in male mice but not in female mice. Ovariectomized female mice showed significant AHR after 10-day OVA inhalation. ICI182,780, an estrogen antagonist, similarly enhanced airway responsiveness even when administered 1 hour before assay. In contrast, 17beta-estradiol dose-dependently suppressed AHR in male mice. In all cases, airway responsiveness was inhibited by the administration of a neurokinin 1 receptor antagonist. These results demonstrate that sex differences in 10-day OVA-induced AHR are due to endogenous estrogen, which negatively regulates airway responsiveness in female mice. Cumulatively, the results suggest that endogenous estrogen may regulate the neurokinin 1-dependent prejunctional activation of airway smooth muscle in allergen-exposed mice.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Disease Models, Animal; Estradiol; Estrogen Receptor Modulators; Estrogens; Female; Fulvestrant; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Neurokinin A; Neurokinin-1 Receptor Antagonists; Ovalbumin; Sex Characteristics

Asthma is a chronic inflammatory disease that is characterized clinically by airway hyperresponsiveness (AHR) to bronchoconstricting agents. The physiological response of the asthmatic lung to inhaled allergen is often characterized by two distinct phases: an early-phase response (EPR) within the first hour following exposure that subsides and a late-phase response (LPR) that is more prolonged and may occur several hours later. Mouse models of asthma have become increasingly popular and should be designed to exhibit an EPR, LPR and AHR.. To determine whether a common model of asthma is capable of demonstrating an EPR, LPR and AHR.. BALB/c mice were sensitized to ovalbumin (OVA) and challenged with one or three OVA aerosols. Changes in lung mechanics in response to allergen inhalation were assessed using a modification of the low-frequency forced oscillation technique (LFOT). In order to assess AHR, changes in lung mechanics in response to aerosolized methacholine were assessed using LFOT. Inflammatory cell infiltration into the lung was measured via bronchoalveolar lavage (BAL). ELISAs were used to measure inflammatory cytokines in the BAL and levels of IgE in the serum.. An EPR was only detectable after three OVA aerosols in approximately half of the mice studied. There was no evidence of an LPR despite a clear increase in cellular infiltration 6 h post-allergen challenge. AHR was present after a single OVA aerosol but not after three OVA aerosols.. The lack of an LPR, limited EPR and the absence of a link between the LPR and AHR highlight the limitations of this mouse model as a complete model of the lung dysfunction associated with asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Humans; Immunoglobulin E; Immunoglobulin G; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Time Factors

Platelets are essential for pulmonary leukocyte recruitment, airway hyperresponsiveness, and bronchial remodeling in animals with allergic inflammation and can be found in bronchoalveolar lavage of sensitized animals. No studies, however, have explored the direct migration of platelets to lungs.. To assess whether platelets migrate into lung parenchyma in response to inhaled allergen in ovalbumin-sensitized mice; to assess the role of the FcepsilonRI receptor in this phenomenon; and to evaluate whether platelets from patients with asthma, or from sensitized mice, undergo chemotaxis in vitro in response to relevant antigens.. Ovalbumin-sensitized wild-type (WT) mice, or FcRgamma(-/-) mice lacking the FcepsilonRIgamma, were challenged with aerosolized allergen and lungs analyzed by platelet-specific immunohistochemistry. In some experiments, mice were depleted of platelets and cross-transfused with either WT or FcRgamma(-/-) platelets to assess the role of platelet FcRgamma(-/-). Chemotaxis of platelets from patients with asthma or from sensitized mice was studied in vitro.. Histology of lungs revealed isolated platelets, migrating out of vessels and localizing underneath the airways after allergen challenge in WT but not in FcRgamma(-/-) mice. Platelets from patients with asthma and from sensitized WT mice, but not from sensitized FcRgamma(-/-) mice, migrated in vitro toward the relevant allergen or an anti-IgE. Platelets from normal mice were found to express FcepsilonRIgamma and platelet-bound IgEs were increased in sensitized mice.. Platelets migrate extravascularly in response to a sensitizing allergen via a mechanism dependent on the interaction among allergen, allergen-specific IgE, and the FcepsilonRI, and this may allow them to participate directly in allergic tissue inflammation.

Topics: Allergens; Animals; Antibodies, Anti-Idiotypic; Asthma; Blood Platelets; Bronchial Hyperreactivity; Chemotaxis; Leukocytes; Lung; Male; Mice; Ovalbumin; Receptors, IgE

Epidemiological studies performed in developing as well as in western countries suggest that infection with Toxocara canis contributes to the development of atopic diseases.. To investigate the association between infection with this helminth and allergy, we examined the effect of T. canis infection on experimental allergic airway inflammation.. BALB/c mice were infected by oral administration with 500 embryonated T. canis eggs followed by ovalbumin (OVA) sensitization and challenge to induce allergic airway inflammation.. Infection with T. canis in combination with OVA treatment leads to exacerbation of pulmonary inflammation, eosinophilia, airway hyperresponsiveness, OVA specific and total IgE. Relative quantification of cytokine expression in the lungs of these mice showed increased expression of IL-4 compared with mice that were only T. canis infected or OVA treated. Increased expression of IL-5 and IL-10 was measured in the lungs of T. canis-infected or OVA-treated mice compared with controls; however, combining infection and OVA treatment did not significantly change the expression of these cytokines.. A previous infection with T. canis leads to exacerbation of experimental allergic airway inflammation. These results have important consequences for findings on the helminths-allergy association. Several factors, including parasite species, infection of definitive vs. accidental host, parasite load and timing of infection, may influence whether an infection with helminths protects one from or enhances allergic manifestations.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Inflammation; Lung; Lung Diseases, Parasitic; Mice; Mice, Inbred BALB C; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; Risk Factors; RNA, Messenger; Time Factors; Toxocara canis; Toxocariasis

We already demonstrated that adoptive transfer of alveolar macrophages (AMs) from non-allergic rats into AM-depleted allergic rats prevents airway hyperresponsiveness (AHR). We also showed that AMs from non-sensitized, but not from sensitized, allergy-prone rats can prevent AHR following allergen challenge in sensitized allergic animals, establishing the importance of rat immunological status on the modulation of AM functions and suggesting that an allergic lung environment alters AM functions.. We investigated how the activation of allergic AMs can be modulated to reinstitute them with their capacity to reduce AHR.. AMs from sensitized Brown Norway rats were cultured ex vivo for up to 18 h in culture media to deprogram them from the influence of the allergic lung before being reintroduced into the lung of AM-depleted sensitized recipient. AHR and cytokines in bronchoalveolar lavage (BAL) were measured following allergen challenge. AMs stimulated ex vivo with Bacillus Calmette-Guerin (BCG) were used as positive controls as BCG induces a T-helper type 1 activation in AMs.. AMs ex vivo cultured for 4-18 h reduced AHR to normal level. Interestingly, pro-allergic functions of AMs were dampened by 18 h culture and they reduced AHR even after spending 48 h in an allergic lung microenvironment. Furthermore, transfer of cultured AMs caused an increase in the levels of IFN-gamma and IL-12 in BAL when compared with their ovalbumin control. After 18 h of ex vivo culture, AMs expressed reduced levels of TNF, IL-1alpha, IL-6, and Arginase-2 mRNAs compared with freshly isolated AMs, suggesting that ex vivo culture exempted AMs from lung stimuli that affected their functions.. There is a significant crosstalk between lung microenvironment and AMs, affecting their functions. It is also the first report showing that sensitized AMs can be modulated ex vivo to reduce lung pro-allergic environment, opening the way to therapies targetting AMs.

Topics: Animals; Arginase; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Separation; Cells, Cultured; Cytokines; Hypersensitivity; Liposomes; Macrophages, Alveolar; Ovalbumin; Rats; Rats, Inbred BN; RNA, Messenger; Time Factors

The Th1/Th2 paradigm has become an important issue in the pathogenesis of asthma, characterized by normal Th1 and elevated Th2 cytokine expression. Vitamin A deficiency (VAD) can produce a Th1 bias, whereas high-level dietary vitamin A can promote a Th2 bias. We used the OVA exposure mouse model to determine the contributions of vitamin A-deficient, control (4IU/g), and high-level vitamin A (250-IU/g) diets to the development of allergic airway inflammation and hyperresponsiveness. VAD reduced serum IgE and IgG1 responses, pulmonary eosinophilia, and the levels of IL-4 and IL-5 in bronchoalveolar lavage specimens, whereas the 250-IU/g diet increased serum IgE. Also, VAD blocked pulmonary hyperresponsiveness following methacholine challenge while the 250-IU/g diet exacerbated pulmonary hyperresponsiveness. In conclusion, VAD diminished and high-level dietary vitamin A enhanced the development of experimental asthma in this model system. These data suggest that excessive intake of vitamin A may increase the risk or severity of asthma in industrialized countries whereas vitamin A deficiency continues to increase mortality from infectious diseases in developing countries.

Topics: Animals; Asthma; Body Weight; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Diet; Disease Models, Animal; Female; Immunoglobulin E; Immunoglobulin G; Interleukin-4; Interleukin-5; Liver; Lung; Macrophages; Male; Mice; Ovalbumin; Pulmonary Eosinophilia; Respiratory Hypersensitivity; Vitamin A; Vitamin A Deficiency

To investigate the effects of murine IL-12 (mIL-12) gene on the expression of CD44 and CD49d on pulmonary leukocytes.. BALB/c mice (n = 24) were sensitized to ovalbumin (OVA) by intraperitoneal injection and challenged with OVA aerosol for 7 consecutive days. mIL-12 plasmid was then intramuscularly administered in eight BALB/c mice every second day for three times during OVA challenge.. Administration of mIL-12 plasmid significantly reduced airway hyperreactivity, the expression of CD44 and CD49d on pulmonary leukocytes, pulmonary infiltration of inflammatory cells, Th2 cytokines production and goblet cell hyperplasia (P < 0.05). Importantly, the percentages of CD44+/CD49d+ leucocytes showed positive correlations with numbers of inflammatory cells, goblet cells hyperplasia and levels of IL-4 and IL-5 (P < 0.01).. Our study suggests that CD44 and CD49d might contribute to the preventive effects of mIL-12 gene in asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Genetic Therapy; Hyaluronan Receptors; Integrin alpha4; Interleukin-12; Interleukin-4; Interleukin-5; Leukocytes; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin

Allergic asthma is a chronic disease characterized by airway obstruction in response to allergen exposure. It results from an inappropriate T helper type 2 response to environmental airborne antigens and affects 300 million individuals. Its prevalence has increased markedly in recent decades, most probably as a result of changes in environmental factors. Exposure to environmental antigens during infancy is crucial to the development of asthma. Epidemiological studies on the relationship between breastfeeding and allergic diseases have reached conflicting results. Here, we have investigated whether the exposure of lactating mice to an airborne allergen affects asthma development in progeny. We found that airborne antigens were efficiently transferred from the mother to the neonate through milk and that tolerance induction did not require the transfer of immunoglobulins. Breastfeeding-induced tolerance relied on the presence of transforming growth factor (TGF)-beta during lactation, was mediated by regulatory CD4+ T lymphocytes and depended on TGF-beta signaling in T cells. In conclusion, breast milk-mediated transfer of an antigen to the neonate resulted in oral tolerance induction leading to antigen-specific protection from allergic airway disease. This study may pave the way for the design of new strategies to prevent the development of allergic diseases.

Topics: Animals; Antigens; Asthma; Bronchial Hyperreactivity; Female; Hypersensitivity; Immune Tolerance; Immunoglobulins; Lactation; Maternal Exposure; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Milk; Ovalbumin; Pneumonia; T-Lymphocytes, Regulatory

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Bronchoconstrictor Agents; Disease Models, Animal; Drug Synergism; Hypersensitivity; Ovalbumin; Polylysine

The peroxisome proliferator-activated receptors (PPARs) alpha, beta/delta, and gamma are ligand-activated transcription factors belonging to the nuclear receptor superfamily. In addition to their regulatory role on lipid and glucose metabolism, they exert anti-inflammatory properties. In skin both PPAR-alpha and PPAR-beta/delta regulate keratinocyte proliferation/differentiation and contribute to wound healing. The 3 PPAR isoforms are expressed by several cell types recruited into the dermis during inflammation.. We have investigated the role of PPAR-alpha in the regulation of atopic dermatitis (AD), a common skin inflammatory disease.. We chose a mouse model of inflammatory dermatosis with immunologic features of AD and used epicutaneous sensitization with ovalbumin in the absence of adjuvant, which mimics the human pathology.. On antigen sensitization, PPAR-alpha-deficient mice display increased epidermal thickening, dermal recruitment of inflammatory cells, lung inflammation, airway hyperresponsiveness, and IgE and IgG2a production compared with their wild-type counterparts. Increased inflammation was correlated to an enhancement of TH2 and, to a greater extent, TH1 responses and to increased skin expression of nuclear factor kappaB. Interestingly, PPAR-alpha expression was decreased in eczematous skin from patients with AD compared with skin from nonatopic donors, suggesting that defective PPAR-alpha expression might contribute to the pathology. Topical application of WY14643, a specific PPAR-alpha agonist, significantly decreased antigen-induced skin inflammation in the AD model.. PPAR-alpha acts as a negative regulator of skin inflammation in AD.

Topics: Administration, Cutaneous; Administration, Topical; Adult; Animals; Bronchial Hyperreactivity; Dermatitis, Atopic; Disease Models, Animal; Female; Humans; Immunoglobulin E; Immunoglobulin G; Inflammation Mediators; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; PPAR alpha; Pyrimidines

Exposure to ozone, which is a major component of air pollution, induces a form of asthma that occurs in the absence of adaptive immunity. Although ozone-induced asthma is characterized by airway neutrophilia, and not eosinophilia, it is nevertheless associated with airway hyperreactivity (AHR), which is a cardinal feature of asthma. Because AHR induced by allergens requires the presence of natural killer T (NKT) cells, we asked whether ozone-induced AHR had similar requirements. We found that repeated exposure of wild-type (WT) mice to ozone induced severe AHR associated with an increase in airway NKT cells, neutrophils, and macrophages. Surprisingly, NKT cell-deficient (CD1d(-/-) and Jalpha18(-/-)) mice failed to develop ozone-induced AHR. Further, treatment of WT mice with an anti-CD1d mAb blocked NKT cell activation and prevented ozone-induced AHR. Moreover, ozone-induced, but not allergen-induced, AHR was associated with NKT cells producing interleukin (IL)-17, and failed to occur in IL-17(-/-) mice nor in WT mice treated with anti-IL-17 mAb. Thus, ozone exposure induces AHR that requires the presence of NKT cells and IL-17 production. Because NKT cells are required for the development of two very disparate forms of AHR (ozone- and allergen-induced), our results strongly suggest that NKT cells mediate a unifying pathogenic mechanism for several distinct forms of asthma, and represent a unique target for effective asthma therapy.

Topics: Air Pollutants; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Interleukin-13; Interleukin-17; Interleukin-4; Killer Cells, Natural; Leukocyte Count; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Ozone; T-Lymphocytes

Interleukin-5 (IL-5) has been suggested to be involved in the development of airway hyper-responsiveness (AHR). Both clinical and experimental investigations have shown strong correlation between the presence of eosinophils and AHR. In this study, we used small interfering RNA (siRNA) as an approach to inhibiting the expression of IL-5 and reducing AHR. siRNAs targeting IL-5 were characterized in vitro, and siRNA-expressing lentiviruses were administered intratracheally to OVA-sensitized BALB/c mice. AHR, cytokine levels, serum levels of OVA-specific antibodies and infiltration of inflammatory cells were analyzed to investigate the effects of siRNA in an OVA-induced murine model of asthma. Lentivirus-delivered siRNA targeting IL-5 efficiently moderated the characteristics of asthma, including AHR, cellular infiltration of lung tissues, eotaxin levels in the bronchoalveolar lavage fluid and IL-5 mRNA levels in lungs in the mouse model of asthma. However, there was no effect on OVA-specific IgE level. These data demonstrate that siRNA delivered by the lentiviral system is an efficacious therapeutic strategy for asthma.

Topics: 5' Untranslated Regions; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Line, Tumor; Chemokine CCL11; Genetic Therapy; Interleukin-5; Lentivirus; Lung; Mice; Mice, Inbred BALB C; Models, Animal; Ovalbumin; Pulmonary Eosinophilia; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; RNA, Small Interfering

Mycobacterium bovis Bacillus Calmette-Guérin (BCG) vaccination has been shown to inhibit allergic airway inflammation in animal models, associated with the regulation of allergen-specific T-cell immunity. However, little is known about whether neonatal BCG treatment could inhibit allergic inflammation by regulating allergen-specific T-cell response in aged mice. This study was aimed to investigate the impact of neonatal BCG treatment on allergic asthma and possible mechanism(s) underlying the action of BCG in different ages of mice.. C57BL/6 neonates were vaccinated with BCG on days 1, 7 and 14, sensitized with ovalbumin (OVA) at 5 and 7 weeks of age, and then challenged with allergen at 9 or 45 weeks of age for early- or late-challenged asthma. Their airway inflammation and allergen-specific T-cell responses were characterized.. Following early-challenge, BCG vaccination inhibited airway hyper-responsiveness (AHR), infiltration of eosinophils and mucous overproduction (P < 0.05), and shifted OVA-specific predominant Th2- to Th1-type cytokine responses in both the bronchoalveolar lavage fluid and the splenocyte supernatants (P < 0.05). In late-challenged mice, neonatal BCG treatment attenuated AHR and eosinophilia (P < 0.05), but failed to modulate allergen-specific cytokine responses.. Our data suggest that neonatal BCG vaccination has a long-term effect on inhibiting AHR and eosinophilia, which is associated with the modulation of Th1/Th2 cytokine production in early-, but not in late-challenged mice. Thus, different mechanisms may mediate the long-term protective effect of BCG neonatal vaccination differently in younger adult and aged mice.

Topics: Animals; Animals, Newborn; Asthma; BCG Vaccine; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Freeze Drying; Humans; Hypersensitivity; Inflammation; Mice; Mice, Inbred C57BL; Mycobacterium bovis; Ovalbumin; T-Lymphocytes

Ozone (O(3)) exposure evokes asthma exacerbations by mechanisms that are poorly understood. We used a murine model to characterize the effects of O(3) on allergic airway inflammation and hyperresponsiveness and to identify factors that might contribute to the O(3)-induced exacerbation of asthma.. BALB/c mice were sensitized and challenged with Aspergillus fumigatus (Af). A group of sensitized and challenged mice was exposed to 3.0 ppm of O(3) for 2 h and studied 12 h later (96 h after Af challenge). Naive mice and mice exposed to O(3) alone were used as controls. Bronchoalveolar lavage (BAL) cellular and cytokine content, lung function [enhanced pause (P(enh))], isometric force generation by tracheal rings and gene and protein expression of Fas and FasL were assessed. Apoptosis of eosinophils was quantified by FACS.. In sensitized mice allergen challenge induced a significant increase of P(enh) and contractile force in tracheal rings that peaked 24 h after challenge and resolved by 96 h. O(3) inhalation induced an exacerbation of airway hyperresponsiveness accompanied by recurrence of neutrophils and enhancement of eosinophils 96 h after allergen challenge. The combination of allergen and O(3) exposure inhibited Fas and FasL gene and protein expression and eosinophil apoptosis and increased interleukin-5 (IL-5), granulocyte-macrophage-colony stimulating factor (GM-CSF) and G-CSF protein levels.. O(3) affects airway responsiveness of allergen-primed airways indirectly by increasing viability of eosinophils and eosinophil-mediated pathological changes.

Topics: Administration, Inhalation; Allergens; Animals; Apoptosis; Aspergillus fumigatus; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophilia; Fas Ligand Protein; Female; Gene Expression Regulation; In Vitro Techniques; Inflammation; Mice; Mice, Inbred BALB C; Ovalbumin; Oxidants, Photochemical; Ozone; Trachea

Allergic asthma is dependent on chemokine-mediated Th2 cell migration and Th2 cytokine secretion into the lungs. The inducible T cell tyrosine kinase Itk regulates the production of Th2 cytokines as well as migration in response to chemokine gradients. Mice lacking Itk are resistant to developing allergic asthma. However, the role of kinase activity of Itk in the development of this disease is unclear. In addition, whether distinct Itk-derived signals lead to T cell migration and secretion of Th2 cytokines is also unknown. Using transgenic mice specifically lacking Itk kinase activity, we show that active kinase signaling is required for control of Th2 responses and development of allergic asthma. Moreover, dominant suppression of kinase Itk activity led to normal Th2 responses, but significantly reduced chemokine-mediated migration, resulting in prevention of allergic asthma. These observations indicate that signals required for Th2 responses and migration are differentially sensitive to Itk activity. Manipulation of Itk's activity can thus provide a new strategy to treat allergic asthma by differentially affecting migration of T cells into the lungs, leaving Th2 responses intact.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Chemokines; Chemotaxis, Leukocyte; Cytokines; Inflammation Mediators; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Protein Structure, Tertiary; Protein-Tyrosine Kinases; Signal Transduction; Th2 Cells

Allergic bronchial asthma is a chronic inflammatory disease of the airways. The transcription factor GATA-3 was shown to play an important role in TH2 cell activation, but also in the regulation of other cell types involved in bronchial asthma including mast cells, eosinophils, and epithelial cells. DNAzymes represent a new class of antisense molecules that combines the specificity of DNA base pairing with an inherent RNA-cleaving enzymatic activity.. To develop a GATA-3 mRNA-specific DNAzyme and analyze its allergy-preventing activity in murine models of experimental allergic asthma.. The most active DNAzyme (termed gd21) was selected by in vitro cleavage assays. Allergic airway inflammation was assessed by inflammatory cell and cytokine analysis within bronchoalveolar lavage. Lung histology, including goblet cell hyperplasia and lung function, was analyzed using head-out body-plethysmography.. Intranasal administration of gd21 prevented airway inflammation and mucus production and inhibited development of airway hyperresponsiveness to methacholine in models of acute allergic airway inflammation. Similar effects were also detected in a model of chronic experimental asthma. Interestingly, gd21 was at least as effective as other antisense molecules, and off-target effects were not detected. Further experiments indicated that pulmonary surfactant may facilitate the cellular uptake of gd21 by acting as an endogenous transfectant.. These results indicate that topical application of the GATA-3-specific DNAzyme is a promising novel approach for the treatment of allergic bronchial asthma.

Topics: Acute Disease; Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Cell Line, Tumor; Chronic Disease; Disease Models, Animal; DNA, Antisense; DNA, Catalytic; Enzyme Activation; Female; GATA3 Transcription Factor; Inflammation Mediators; Mice; Mice, Inbred BALB C; Ovalbumin; RNA, Messenger; RNA, Small Interfering; Substrate Specificity

Recent epidemiologic studies have identified organophosphorus pesticides (OPs) as environmental factors potentially contributing to the increase in asthma prevalence over the last 25 years. In support of this hypothesis, we have demonstrated that environmentally relevant concentrations of OPs induce airway hyperreactivity in guinea pigs.. Sensitization to allergen is a significant contributing factor in asthma, and we have shown that sensitization changes virus-induced airway hyperreactivity from an eosinophil-independent mechanism to one mediated by eosinophils. Here, we determine whether sensitization similarly influences OP-induced airway hyperreactivity.. Nonsensitized and ovalbumin-sensitized guinea pigs were injected subcutaneously with the OP parathion (0.001-1.0 mg/kg). Twenty-four hours later, animals were anesthetized and ventilated, and bronchoconstriction was measured in response to either vagal stimulation or intravenous acetylcholine. Inflammatory cells and acetylcholinesterase activity were assessed in tissues collected immediately after physiologic measurements.. Ovalbumin sensitization decreased the threshold dose for parathion-induced airway hyperreactivity and exacerbated parathion effects on vagally induced bronchoconstriction. Pretreatment with antibody to interleukin (IL)-5 prevented parathion-induced hyperreactivity in sensitized but not in nonsensitized guinea pigs. Parathion did not increase the number of eosinophils in airways or the number of eosinophils associated with airway nerves nor did it alter eosinophil activation as assessed by major basic protein deposition.. Antigen sensitization increases vulnerability to parathion-induced airway hyperreactivity and changes the mechanism to one that is dependent on IL-5. Because sensitization to allergens is characteristic of 50% of the general population and 80% of asthmatics (including children), these findings have significant implications for OP risk assessment, intervention, and treatment strategies.

Topics: Acetylcholinesterase; Animals; Bradycardia; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Dose-Response Relationship, Drug; Electric Stimulation; Female; Guinea Pigs; Immunization; Interleukin-5; Ovalbumin; Parathion; Pesticides; Vagus Nerve

The angiopoietins (Ang) comprise a family of growth factors mainly known for their role in blood vessel formation and remodeling. The best-studied member, Ang-1, exhibits antiapoptotic and antiinflammatory effects. Although the involvement of Ang-1 in angiogenesis is well recognized, little information exists about its role in respiratory physiology and disease. On the basis of its ability to inhibit vascular permeability, adhesion molecule expression, and cytokine production, we hypothesized that Ang-1 administration might exert a protective role in asthma.. To determine changes in the expression of Ang and to assess the ability of Ang-1 to prevent the histologic, biochemical, and functional changes observed in an animal model of asthma.. To test our hypothesis, a model of allergic airway disease that develops after ovalbumin (OVA) sensitization and challenge was used.. Ang-1 expression was reduced at the mRNA and protein levels in lung tissue of mice sensitized and challenged with OVA, leading to reduced Tie2 phosphorylation. Intranasal Ang-1 treatment prevented the OVA-induced eosinophilic lung infiltration, attenuated the increase in IL-5 and IL-13, and reduced eotaxin and vascular cell adhesion molecule 1 expression. These antiinflammatory actions of Ang-1 coincided with higher levels of IkappaB and decreased nuclear factor-kappaB binding activity. More importantly, Ang-1 reversed the OVA-induced increase in tissue resistance and elastance, improving lung function.. We conclude that Ang-1 levels are decreased in asthma and that administration of Ang-1 might be of therapeutic value because it prevents the increased responsiveness of the airways to constrictors and ameliorates inflammation.

Topics: Airway Resistance; Angiopoietin-1; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Immunoglobulin E; Interleukin-5; Male; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Vascular Cell Adhesion Molecule-1

The present study is performed to investigate the inhibitory effects of Radix Adenophorae extract (RAE) on ovalbumin-induced asthma murine model. To study the anti-inflammatory and antiasthmatic effects of RAE, we examined the development of pulmonary eosinophilic inflammation and inhibitory effects of T cells in murine by RAE and cyclosporine A (CsA). We examined determination of airway hyperresponsiveness, flow cytometric analysis (FACS), enzyme-linked immunosorbent assay (ELISA), quantitative real time (PCR), hematoxylin-eosin staining, and Masson trichrome staining in lung tissue, lung weight, total cells, and eosinophil numbers in lung tissue. We demonstrated how RAE suppressed development on inflammation and decreased airway damage.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cyclosporine; Cytokines; Female; Humans; Immunoglobulin E; Immunosuppressive Agents; Lung; Mice; Mice, Inbred C57BL; Ovalbumin; Plant Extracts; Pulmonary Eosinophilia

It is unclear whether inhaled lidocaine is effective against airway hyperreactivity and inflammation in asthma. The aim of this study was to investigate the effects of inhaled lidocaine on airway hyperreactivity and inflammation. Airway reactivity to inhaled histamine, cellular composition of bronchoalveolar lavage (BAL) fluid, plasma substance P (SP), and isolated lung tissue were evaluated in ovalbumin (OVA)-sensitized guinea pigs 7 days after OVA challenge. The effects of inhaled lidocaine on this model were also evaluated. Treatment with lidocaine was administered in two fashions: as single inhalation or inhalation bid for 7 consecutive days, for comparison with a saline-inhaled control group. Airway hyperreactivity to histamine, increase in number of total cells and increased proportion of eosinophils in BAL fluid, and marked eosinophil infiltration in airway walls were noted even 7 days after OVA challenge in the control group. Plasma SP level was also significantly increased. Although treatment with single lidocaine inhalation did not affect airway hyperreactivity, continued inhalation (bid for 7 days) attenuated airway hyperreactivity. Continued, but not single, inhalation of lidocaine also suppressed infiltration of eosinophils in BAL fluid and in airway walls. In addition, plasma SP levels were significantly reduced by continued but not by single inhalation. It appears possible that lidocaine when inhaled suppresses eosinophilic inflammation of the airway and SP-induced neurogenic inflammation, leading to alleviation of airway hyperreactivity.

Topics: Administration, Inhalation; Anesthetics, Local; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Capsaicin; Cell Count; Cough; Eosinophils; Guinea Pigs; Histamine; Inflammation; Lidocaine; Lung; Male; Ovalbumin; Substance P

Leukotriene B4 (LTB4) is a potent inflammatory lipid mediator that binds to LTB4 receptor 1 (BLT1). Ligation of BLT1 by LTB4 plays an important role in the recruitment of effector memory CD8+ T cells into the airways of sensitized and challenged mice.. The effects of the corticosteroid dexamethasone (DEX) on BLT1-expressing effector memory CD8+ T cells and effector memory CD8+ T cell-mediated airway hyperresponsiveness (AHR) and allergic inflammation were determined.. Effector memory CD8+ T cells were generated from ovalbumin(257-264)-primed mononuclear cells from OT-1 mice in the presence of IL-2. In some cultures DEX was added. The effects of DEX on BLT1 expression, LTB4-induced Ca2+ influx, phosphorylation of extracellular signal-regulated kinase 1/2, chemotaxis, and effector memory CD8+ T cell-mediated AHR were examined.. DEX-treated effector memory CD8+ T cells showed significant increases in surface expression of BLT1, LTB4-induced intracellular Ca2+ influx, phosphorylation of extracellular signal-regulated kinase 1/2, and chemotaxis. Upregulation of BLT1 by DEX was accompanied by increased IL-2 receptor expression. Adoptive transfer of DEX-treated effector memory CD8+ T cells into ovalbumin-sensitized and ovalbumin-challenged CD8-/- mice resulted in significant increases in AHR, allergic inflammation, goblet cell metaplasia, and numbers of both CD8+ and CD4+ T cells in the bronchoalveolar lavage fluid and lungs.. Corticosteroids upregulate BLT1 on effector memory CD8+ T cells and related signaling pathways and potentiate allergic airway inflammation and AHR induced by these cells.

Topics: Adjuvants, Immunologic; Allergens; Animals; Bronchial Hyperreactivity; Calcium Signaling; CD8-Positive T-Lymphocytes; Cell Differentiation; Cells, Cultured; Dexamethasone; Egg Proteins; Immunologic Memory; Inflammation Mediators; MAP Kinase Signaling System; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Peptide Fragments; Phosphorylation; Receptors, Leukotriene B4; Up-Regulation

Allergic asthma is a complex immunologically mediated disease associated with increased oxidative stress and altered antioxidant defenses. It was hypothesized that alpha-tocopherol (alpha-T) decreases oxidative stress and therefore its absence may influence allergic inflammatory process, a pathobiology known to be accompanied by oxidative stress. Therefore, selected parameters of allergic asthma sensitization and inflammation were evaluated following ovalbumin sensitization and re-challenge of alpha-T transfer protein (TTP) knock-out mice (TTP(-/-)) that have greatly reduced lung alpha-T levels (e.g.<5%) compared to their litter mate controls (TTP(+/+)). Results showed that severe alpha-T deficiency result in a blunted lung expression of IL-5 mRNA and IL-5 protein and plasma IgE levels compared with TTP(+/+) mice following immune sensitization and rechallenge, although lung lavage eosinophil levels were comparable in both genomic strains. It is concluded that the initial stimulation of immune responses by the TTP(-/-) mice were generally blunted compared to the TTP(+/+) mice, thus diminishing some aspects of subsequent allergic inflammatory processes.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Carrier Proteins; Eosinophils; Gene Expression Regulation; Immunoglobulin E; Inflammation; Interleukin-5; Lung; Mice; Mice, Knockout; Ovalbumin; Oxidative Stress; Vitamin E Deficiency

Allergic inflammation is known to cause airway hyperresponsiveness in mice. However, it is not known whether inflammation affects the stiffness of the airway wall, which would alter the load against which the circumscribing smooth muscle shortens when activated. Accordingly, we measured the time course of airway resistance immediately following intravenous methacholine injection in acutely and chronically allergically inflamed mice. We estimated the effective stiffness of the airway wall in these animals by fitting to the airway resistance profiles a computational model of a dynamically narrowing airway embedded in elastic parenchyma. Effective airway wall stiffness was estimated from the model fit and was found not to change from control in either the acute or chronic inflammatory groups. However, the acutely inflamed mice were hyperresponsive compared with controls, which we interpret as reflecting increased delivery of methacholine to the airway smooth muscle through a leaky pulmonary endothelium. These results support the notion that acutely inflamed BALB/c mice represent an animal model of functionally normal airway smooth muscle in a transiently abnormal lung.

Topics: Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstrictor Agents; Computer Simulation; Disease Models, Animal; Elasticity; Female; Inflammation; Injections, Intravenous; Methacholine Chloride; Mice; Mice, Inbred BALB C; Models, Biological; Ovalbumin; Time Factors

Adoptive transfer of in vivo-primed CD8(+) T cells or in vitro-generated effector memory CD8(+) T (T(EFF)) cells restores airway hyperresponsiveness (AHR) and airway inflammation in CD8-deficient (CD8(-/-)) mice. Examining transcription levels, there was a strong induction of Notch1 in T(EFF) cells compared with central memory CD8(+) T cells. Treatment of T(EFF) cells with a gamma-secretase inhibitor (GSI) strongly inhibited Notch signaling in these cells, and after adoptive transfer, GSI-treated T(EFF) cells failed to restore AHR and airway inflammation in sensitized and challenged recipient CD8(-/-) mice, or to enhance these responses in recipient wild-type (WT) mice. These effects of GSI were also associated with increased expression of the Notch ligand Delta1 in T(EFF) cells. Treatment of sensitized and challenged WT mice with Delta1-Fc resulted in decreased AHR and airway inflammation accompanied by higher levels of interferon gamma in bronchoalveolar lavage fluid. These results demonstrate a role for Notch in skewing the T cell response from a T helper (Th)2 to a Th1 phenotype as a consequence of the inhibition of Notch receptor activation and the up-regulation of the Notch ligand Delta1. These data are the first to show a functional role for Notch in the challenge phase of CD8(+) T cell-mediated development of AHR and airway inflammation, and identify Delta1 as an important regulator of allergic airway inflammation.

Topics: Adoptive Transfer; Animals; Bronchial Hyperreactivity; CD8-Positive T-Lymphocytes; Cytokines; Immunologic Memory; Inflammation; Mice; Mice, Inbred C57BL; Mice, Transgenic; Oligonucleotide Array Sequence Analysis; Ovalbumin; Receptors, Antigen, T-Cell; Receptors, Notch; RNA

To evaluate the effect of erythromycin on bronchial hyperreactivity, inflammation, and T-cell related cytokine mRNA expression in rats sensitized to ovalbumin, three experimental groups of 10 brown Norway rats each were sensitized by breathing aerosolized ovalbumin. From day 1 to day 15, one group was given oral erythromycin 80 mg/kg/day, another group oral erythromycin 20mg/kg/day, and the third group oral saline only. A fourth control group of 10 rats breathed aerosolized saline. After sensitization, the three experimental groups were provoked by breathing ovalbumin, with the controls again breathing saline. The rats were then anesthetized and paralyzed, and pulmonary function tests were performed at baseline and after varying doses of acetylcholine. Bronchoalveolar lavage (BAL) fluid and lung tissues were examined for expression of mRNA for T-cell cytokines. Our results showed that erythromycin had no beneficial effects on pulmonary function and lung inflammation in the two erythromycin-treated experimental groups compared with the saline experimental group. Th2-related cytokines and their mRNA expression in the three experimental groups were higher than in controls but did not differ among the experimental groups. In conclusion, erythromycin does not prevent bronchial hyperreactivity or an inflammatory response in ovalbumin-sensitized rats.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chlamydophila; Cytokines; Dose-Response Relationship, Drug; Erythromycin; Gene Expression Regulation; Inflammation; Male; Mycoplasma; Ovalbumin; Plethysmography, Whole Body; Protein Synthesis Inhibitors; Rats; Rats, Inbred BN; T-Lymphocytes

Airway hyperresponsiveness (AHR) is a hallmark of bronchial asthma. Increased expression of smooth muscle contractile proteins or increased responsiveness of the contractile apparatus due to RhoA/Rho-kinase activation may contribute to AHR. BALB/c mice developed AHR following systemic sensitization by intraperitoneal injections of 20 microg ovalbumin (OVA) in presence of 2mg Al(OH)(3) on days 1 and 14, and airway challenge by 1% OVA-inhalation for 20 min each on days 28, 29 and 30. As assessed by Western blot, protein expression of RhoA, MLC (myosin light chain) and smMLCK (smooth muscle myosin light chain kinase) was increased in lungs of OVA/OVA-animals with AHR, as well as in lungs of OVA-sensitized and sham-challenged animals (OVA/PBS) without AHR, compared with lungs of PBS/PBS-animals. Pretreatment with the specific Rho-kinase inhibitor Y-27632 reduced MLC-phosphorylation and AHR. Contribution of Rho-kinase to bronchoconstriction was increased in lungs of OVA/OVA-animals compared with OVA/PBS- and PBS/PBS-animals, respectively. Furthermore, bronchoconstriction following MCh stimulation was significantly reduced after Y-27632 application. In conclusion, systemic allergen-sensitization increased pulmonary expression of proteins involved in smooth muscle contraction, which may contribute to development of AHR. However, this observation was independent from local allergen challenge, suggesting that additional cofactors may be required for the activation of Rho-kinase and thereby the induction of AHR. Rho-kinase may play an important role in murine AHR, and the bronchodilating action of Rho-kinase inhibition may offer a new therapeutic perspective in obstructive airway disease.

Topics: Amides; Animals; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchoconstriction; Disease Models, Animal; Enzyme Inhibitors; Female; Lung; Mice; Mice, Inbred BALB C; Muscle, Smooth; Myosin Light Chains; Myosin-Light-Chain Kinase; Ovalbumin; Perfusion; Phosphorylation; Pyridines; rho-Associated Kinases

The effect of ovalbumin (Ova) sensitization on pulmonary C-fiber sensitivity was investigated. Brown-Norway rats were sensitized by intraperitoneal injection of Ova followed by aerosolized Ova three times per week for 3 wk. Control rats received the vehicle. At the end of the third week, single-unit fiber activities (FA) of pulmonary C fibers were recorded in anesthetized, artificially ventilated rats. Our results showed the following: 1) Ova sensitization induced airway inflammation (infiltration of eosinophils and neutrophils) and airway hyperresponsiveness in rats; 2) baseline FA in sensitized rats was significantly higher than that in control ones; 3) similarly, the pulmonary C-fiber response to right atrial injection of capsaicin was markedly higher in sensitized rats, which were significantly amplified after the acute Ova inhalation challenge; and 4) similar patterns, but smaller magnitudes of the differences in C-fiber responses to adenosine and lung inflation, were also found between sensitized and control rats. In conclusion, Ova sensitization elevated the baseline FA and excitability of pulmonary C fibers, and the hypersensitivity was further potentiated after the acute Ova inhalation challenge in sensitized rats. Chronic allergic inflammatory reactions in the airway probably contributed to the sensitizing effect on these lung afferents.

Topics: Administration, Inhalation; Aerosols; Allergens; Animals; Blood Pressure; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Dose-Response Relationship, Drug; Heart Rate; Hypersensitivity; Inflammation; Lung; Male; Methacholine Chloride; Nerve Fibers, Unmyelinated; Neurons, Afferent; Ovalbumin; Rats; Rats, Inbred BN

Arginase gene expression in the lung has been linked to asthma both in clinical studies of human patients and in the well-studied mouse model of ovalbumin-induced airway inflammation. Arginase is thought to regulate NO levels in the lung by its ability to divert arginine, the substrate for nitric oxide synthases that produce citrulline and NO, into an alternative metabolic pathway producing ornithine and urea. In the present study arginase I and arginase II concentrations were measured in isolated microdissected airway preparations from sensitized Balb/c mice exposed to ovalbumin aerosol. We found that arginase II was constitutively expressed in the airways of normal mice, whereas arginase I was undetectable in normal airways, while its expression was increased in airways of mice exposed to ovalbumin. The expression of arginase I strongly correlated with the presence of lung inflammation, as quantified by differential cell counts in lung lavage, suggesting that most, or all, of the arginase I in lungs of mice exposed to ovalbumin is present in the inflammatory cells rather than in the airway epithelium. There was also a significant correlation between increased expression of arginase I in the isolated airways and decreased lung compliance. On the other hand, while we found arginase II expression to also be significantly increased in airways from mice exposed to ovalbumin as compared with normal airways, the relative increase was much less than that observed for arginase I, suggesting that there was a smaller contribution of inflammatory cells to the arginase II content of the airways in mice exposed to ovalbumin. There was no apparent correlation between the content of arginase in isolated airways and exhaled NO concentration in the expired air from mice exposed to ovalbumin. However, there was a correlation between exhaled NO concentration from mice exposed to ovalbumin and the lymphocyte content of the lung lavage. The concentration of arginine found in isolated airways from Balb/c mice exposed for 2 weeks to ovalbumin was about half of the value found in isolated microdissected airways from normal mice. Treatment of mice systemically with an arginase inhibitor significantly increased the amount of NO produced, as measured as the amount of nitrite+nitrate (NOx) in lung lavage supernatant prepared from mice exposed to ovalbumin. Our results are consistent with the hypothesis that the response of the lung to ovalbumin challenge includes an adaptive respon

Topics: Animals; Arginase; Arginine; Bronchial Hyperreactivity; Inflammation; Lung; Male; Mice; Mice, Inbred BALB C; Nitric Oxide; Ovalbumin

Asthma is a multifactorial respiratory disease. Though its incidence is increasing rapidly all over the world, the available therapeutic strategies are neither sufficient nor safe for long term use. Mepacrine, a known antimalarial drug, has been shown to possess antioxidant, anti-inflammatory, platelet anti-aggregant, and PLA2 inhibitory activities. However, its possible use in asthma has not been studied yet. The objective of this study was to investigate the anti-asthmatic property of mepacrine using a mouse model of asthma. To accomplish this, male BALB/c mice were sensitized and challenged with ovalbumin and treated with increasing concentrations of mepacrine. Airway hyperresponsiveness (AHR) to methacholine was assessed using unrestrained whole body plethysmography. Mepacrine (1 mg/kg) has shown marked attenuation of AHR. Cytokines such as IL-4, IL-5, IL-13 and IFN-gamma and OVA-specific IgE levels were measured in BAL (bronchoalveloar lavage) fluid and sera, respectively. Mepacrine effectively reduced the rise in IL-4, IL-5, IL-13, and OVA-specific IgE and restored IFN-gamma levels. Mepacrine also significantly prevented the increase of sPLA2 (secretory phospholipase A2) activity in BAL fluid supernatant and Cys-LT (cysteinyl leukotrienes) in lung tissue homogenates of asthmatic mice. In addition, mepacrine treatment reduced BAL fluid eosinophilia and signs of allergic airway inflammation such as perivascular and peribronchial distribution of inflammatory cells. These findings indicate that mepacrine reduces the asthmatic features in ovalbumin induced asthma by acting on PLA2-Cys-LT axis. Thus, it could be useful for the development of better asthma therapy.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cysteine; Cytokines; Disease Models, Animal; Eosinophils; Immunoglobulin E; Inflammation; Leukotrienes; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Phospholipases A2, Secretory; Quinacrine

Peroxiredoxin II (PrxII) is one of reactive oxygen species (ROS)-degrading enzyme. Here, we investigated the role of PrxII on toll-like receptor 4 (TLR4) and B-cell activating factor (BAFF) expression in ovalbumin (OVA)-induced mouse asthma. We used ROS-producing PrxII-/- mice of which cells up-regulate BAFF expression. As significant changes were detected in TLR4 mRNA with real-time quantitative RT-PCR analysis, TLR4 protein was decreased in PrxII-/- mouse splenocytes and peritoneal macrophages, compared to wild type cells. Airway hyper-responsiveness (AHR) was more severe in PrxII-/- mice than wild type mice, which was measured by the level of various parameters, number of eosinophils, IgE level, airway thickness, and mucous secretion. BAFF was detected in cells surrounding airways of OVA-induced mouse and it was highly augmented in PrxII-/- mice. BAFF promoter activity was also higher in PrxII-/- mouse embryonic fibroblast (MEF) than in wild type MEF. Collectively, results show that PrxII may have benefits in asthma through reducing ROS. It suggests that BAFF and TLR4 expressions are differentially regulated by PrxII and TLR4 protein level may not be crucial in OVA-induced asthma.

Topics: Animals; Asthma; B-Cell Activating Factor; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Fibroblasts; Macrophages, Peritoneal; Mice; Mice, Mutant Strains; Ovalbumin; Peroxiredoxins; Reactive Oxygen Species; Spleen; Toll-Like Receptor 4

T-bet and STAT4 play critical roles in helper T cell differentiation, especially for Th1 cells. However, it is still unknown about the relative importance and redundancy of T-bet and STAT4 for Th1 differentiation. It is also unknown about their independent role of T-bet and STAT4 in the regulation of allergic airway inflammation. In this study, we addressed these issues by comparing T-bet-deficient (T-bet(-/-)) mice, STAT4(-/-) mice, and T-bet- and STAT4-double-deficient (T-bet(-/-)STAT4(-/-)) mice on the same genetic background. Th1 differentiation was severely decreased in T-bet(-/-) mice and STAT4(-/-) mice as compared with that in wild-type mice, but Th1 differentiation was still observed in T-bet(-/-) mice and STAT4(-/-) mice. However, Th1 cells were hardly detected in T-bet(-/-)STAT4(-/-) mice. In contrast, the maintenance of Th17 cells was enhanced in T-bet(-/-) mice but was reduced in STAT4(-/-) mice and T-bet(-/-)STAT4(-/-) mice. In vivo, Ag-induced eosinophil and neutrophil recruitment into the airways was enhanced in T-bet(-/-) mice but was attenuated in STAT4(-/-) mice and T-bet(-/-)STAT4(-/-) mice. Ag-induced IL-17 production in the airways was also diminished in STAT4(-/-) mice and T-bet(-/-)STAT4(-/-) mice. These results indicate that STAT4 not only plays an indispensable role in T-bet-independent Th1 differentiation but also is involved in the maintenance of Th17 cells and the enhancement of allergic airway inflammation.

Topics: Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Differentiation; Cell Proliferation; Cytokines; Flow Cytometry; Hypersensitivity; Interleukin-17; Mice; Mice, Mutant Strains; Ovalbumin; Pneumonia; STAT4 Transcription Factor; T-Box Domain Proteins; T-Lymphocyte Subsets; Th1 Cells; Th2 Cells

Eosinophils have been implicated as playing a major role in allergic airway responses. However, the importance of these cells to the development of this disease has remained ambiguous despite many studies, partly because of lack of appropriate model systems. In this study, using transgenic murine models, we more clearly delineate a role for eosinophils in asthma. We report that, in contrast to results obtained on a BALB/c background, eosinophil-deficient C57BL/6 Delta dblGATA mice (eosinophil-null mice via the Delta DblGATA1 mutation) have reduced airway hyperresponsiveness, and cytokine production of interleukin (IL)-4, -5, and -13 in ovalbumin-induced allergic airway inflammation. This was caused by reduced T cell recruitment into the lung, as these mouse lungs had reduced expression of CCL7/MCP-3, CC11/eotaxin-1, and CCL24/eotaxin-2. Transferring eosinophils into these eosinophil-deficient mice and, more importantly, delivery of CCL11/eotaxin-1 into the lung during the development of this disease rescued lung T cell infiltration and airway inflammation when delivered together with allergen. These studies indicate that on the C57BL/6 background, eosinophils are integral to the development of airway allergic responses by modulating chemokine and/or cytokine production in the lung, leading to T cell recruitment.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Eosinophils; GATA1 Transcription Factor; Hypersensitivity; Inflammation; Interleukins; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Species Specificity; T-Lymphocytes

Eosinophils represent one of the main effector cell populations of allergic airway inflammation and allergic bronchial asthma. Their infiltration correlates with many characteristics of the disease, including airway hyperresponsiveness (AHR) and increased mucus production. CCR-3 is the principle chemokine receptor involved in eosinophil attraction into inflamed tissue. Therefore, antagonizing CCR-3 could be a novel promising approach toward asthma therapy. We investigated the effect of a low-molecular-weight CCR-3 antagonist on established airway inflammation in a chronic model of experimental bronchial asthma. For this purpose, BALB/c mice intraperitoneally sensitized with ovalbumin (OVA) were chronically challenged with OVA aerosol to induce chronic airway inflammation and airway remodeling. The effect of antagonizing CCR-3 on asthma pathology was examined in BAL and lung histology. Airway reactivity was assessed by head-out body plethysmography. Treatment with the CCR-3 antagonist resulted in a marked reduction of eosinophils in the bronchoalveolar lumen and in airway wall tissue, whereas infiltration of lymphocytes or macrophages remained unchanged. The reduction in eosinophil infiltration was accompanied by normalization of AHR and prevention of goblet cell hyperplasia, indicating reduced mucus production. Furthermore, antagonizing CCR-3 prevented airway remodeling as defined by subepithelial fibrosis and increased accumulation of myofibrocytes in the airway wall of chronically challenged mice. These data demonstrate that antagonism of CCR3 reduces eosinophil numbers, which is accompanied by diminution of asthma pathology in a mouse model of established chronic experimental asthma. Therefore, antagonizing CCR-3 represents a new approach toward a promising asthma therapy.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chronic Disease; Disease Models, Animal; Eosinophils; Female; Lung; Lymphocytes; Mice; Mice, Inbred BALB C; Ovalbumin; Receptors, CCR3; Receptors, Chemokine

The Th2 cytokines IL-4 and IL-13 mediate allergic pulmonary inflammation and airways hyperreactivity (AHR) in asthma models through signaling dependent upon the IL-4 receptor-alpha chain (IL-4Ralpha). IL-13 has been further implicated in the overproduction of mucus by the airway epithelium and in lung remodeling that commonly accompanies chronic inflammation. IL-4Ralpha-deficient mice are resistant to allergen-induced asthma, highlighting the therapeutic promise of selective molecular inhibitors of IL-4Ralpha. We designed a chemically modified IL-4Ralpha antisense oligonucleotide (IL-4Ralpha ASO) that specifically inhibits IL-4Ralpha protein expression in lung eosinophils, macrophages, dendritic cells, and airway epithelium after inhalation in allergen-challenged mice. Inhalation of IL-4Ralpha ASO attenuated allergen-induced AHR, suppressed airway eosinophilia and neutrophilia, and inhibited production of airway Th2 cytokines and chemokines in previously allergen-primed and -challenged mice. Histologic analysis of lungs from these animals demonstrated reduced goblet cell metaplasia and mucus staining that correlated with inhibition of Muc5AC gene expression in lung tissue. Therapeutic administration of inhaled IL-4Ralpha ASO in chronically allergen-challenged mice produced a spectrum of anti-inflammatory activity similar to that of systemically administered Dexamethasone with the added benefit of reduced airway neutrophilia. These data support the potential utility of a dual IL-4 and IL-13 oligonucleotide inhibitor in allergy/asthma, and suggest that local inhibition of IL-4Ralpha in the lung is sufficient to suppress allergen-induced pulmonary inflammation and AHR.

Topics: Administration, Inhalation; Aerosols; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Chemokines; Dendritic Cells; Disease Models, Animal; Eosinophils; Gene Expression Regulation; Goblet Cells; Inflammation; Lung; Macrophages, Alveolar; Male; Metaplasia; Mice; Mucins; Oligonucleotides, Antisense; Ovalbumin; Receptors, Cell Surface; Th2 Cells; Treatment Outcome

Eosinophils contribute to the early features of allergic lung inflammation through the generation and release of a plethora of mediators. Eosinophil peroxidase (EPO) is one of the eosinophil granule proteins involved in the early response, but its participation in airway remodeling is not established. The present study addressed this question comparing an EPO-deficient mouse strain (NZW) with BALB/c and C57Bl/c strains.. Mice were immunized with ovalbumin/alum, challenged twice with ovalbumin aerosol, and lung responses were measured at day 22 or 28. Collagen, mucus and eosinophils were determined in lung sections stained with picrosirius, periodic acid-Schiff or hematoxylin-eosin; transforming growth factor-beta and vascular endothelial growth factor were determined by ELISA, lipid bodies by enumeration in osmium-stained eosinophils, and airway reactivity to methacholine in isolated lung preparations.. NZW mice showed significantly less collagen around bronchi and blood vessels, less mucus and less eosinophils around bronchi. Eosinophil lipid body formation and airway hyperreactivity were comparable among strains. Levels of transforming growth factor-beta were also comparable; however, the NZW mice showed much higher levels of vascular endothelial growth factor, even under basal conditions.. In allergic lung inflammation, the combination of EPO deficiency and overexpression of VEGF found in NZW mice is associated with less collagen deposition, less mucus and reduced tissue eosinophilia. Eosinophil activation and airway hyperreactivity in NZW mice were similar to the other strains.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Collagen; Eosinophil Peroxidase; Eosinophilia; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Pirfenidone was administered to sensitized Brown Norway rats prior to a series of ovalbumin challenges. Airway hyperresponsiveness, inflammatory cell infiltration, mucin and collagen content, and the degree of epithelium and smooth muscle staining for TGF-beta were examined in control, sensitized, and sensitized/challenged rats fed a normal diet or pirfenidone diet. Pirfenidone had no effect on airway hyperresponsiveness, but reduced distal bronchiolar cell infiltration and proximal and distal mucin content. Statistical analysis showed that the control group and sensitized/challenged pirfenidone diet group TGF-beta staining intensity scores were not significantly different from isotype controls, but that the staining intensity scores for the sensitized/challenged normal diet group was significantly different from isotype controls. These results suggest that pirfenidone treatment is effective in reducing some of the components of acute inflammation induced by allergen challenge.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Collagen; Disease Models, Animal; Inflammation; Lung; Male; Mucins; Muscle, Smooth; Ovalbumin; Pyridones; Random Allocation; Rats; Rats, Inbred BN; Respiratory Mucosa; Transforming Growth Factor beta

Airway hyperresponsiveness is a critical feature of asthma. Substantial epidemiologic evidence supports a role for female sex hormones in modulating lung function and airway hyperresponsiveness in humans.. To examine the role of estrogen receptors in modulating lung function and airway responsiveness using estrogen receptor-deficient mice.. Lung function was assessed by a combination of whole-body barometric plethysmography, invasive measurement of airway resistance, and isometric force measurements in isolated bronchial rings. M2 muscarinic receptor expression was assessed by Western blotting, and function was assessed by electrical field stimulation of tracheas in the presence/absence of gallamine. Allergic airway disease was examined after ovalbumin sensitization and exposure.. Estrogen receptor-alpha knockout mice exhibit a variety of lung function abnormalities and have enhanced airway responsiveness to inhaled methacholine and serotonin under basal conditions. This is associated with reduced M2 muscarinic receptor expression and function in the lungs. Absence of estrogen receptor-alpha also leads to increased airway responsiveness without increased inflammation after allergen sensitization and challenge.. These data suggest that estrogen receptor-alpha is a critical regulator of airway hyperresponsiveness in mice.

Topics: Acetylcholine; Allergens; Animals; Bronchial Hyperreactivity; Cytokines; Electrophysiology; Estrogen Receptor alpha; Estrogens; Female; Inflammation; Lung; Methacholine Chloride; Mice; Mice, Knockout; Ovalbumin; Peripheral Nerves; Plethysmography; Receptor, Muscarinic M2; Respiratory Hypersensitivity; Serotonin; Trachea

The D6 chemokine receptor can bind and scavenge several chemokines, including the T-helper 2 (Th2)-associated chemokines CCL17 and CCL22. Although D6 is constitutively expressed in the lung, its pulmonary function is unknown.. This study tested whether D6 regulates pulmonary chemokine levels, inflammation, or airway responsiveness during allergen-induced airway disease.. D6-deficient and genetically matched C57BL/6 mice were sensitized and challenged with ovalbumin. ELISA and flow cytometry were used to measure levels of cytokines and leukocytes, respectively. Mechanical ventilation was used to measure airway reactivity.. The ability of D6 to diminish chemokine levels in the lung was chemokine concentration dependent. CCL17 and CCL22 were abundant in the airway, and their levels were attenuated by D6 when they were within a defined concentration range. By contrast, airway concentrations of CCL3, CCL5, and CCL11 were low and unaffected by D6. Allergen-challenged D6-deficient mice had more dendritic cells, T cells, and eosinophils in the lung parenchyma and more eosinophils in the airway than similarly challenged C57BL/6 mice. By contrast, D6-deficient mice had reduced airway responses to methacholine compared with C57BL/6 mice. Thus, D6 has opposing effects on inflammation and airway reactivity.. The ability of D6 to scavenge chemokines in the lung is dependent on chemokine concentration. The absence of D6 increases inflammation, but reduces airway reactivity. These findings suggest that inhibiting D6 function might be a novel means to attenuate airway responses in individuals with allergic asthma.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Chemokine Receptor D6; Chemokines, CC; Eosinophils; Inflammation; Leukocytes; Lung; Lung Diseases, Obstructive; Lymph Nodes; Methacholine Chloride; Mice; Mice, Knockout; Ovalbumin; Receptors, CCR10; Receptors, Chemokine; Respiratory Hypersensitivity; Transforming Growth Factor beta1

Antigen inhalation in patients with atopic asthma results in an early asthmatic response (EAR), accompanied by a late asthmatic response (LAR) in 60% of patients, airway hyperresponsiveness (AHR) and inflammatory cell infiltration to the lungs. An ideal animal model of asthma should, therefore, provide at least these 4 features consistently and reproducibly. The aim of this study was to optimise the ovalbumin (OA) sensitisation conditions, for achieving EAR, LAR, AHR and cell influx, in a guinea-pig model of asthma. Animals were sensitised with 10 micro g or 100 micro g OA, as either a single or booster (day 1 and day 5) injection. Airway responses to inhaled OA (10 micro g, 1 h) of actively sensitised, conscious guinea pigs were determined by whole body plethysmography as the change in specific airways conductance (sG(aw)) over a 12 h period and at 24 h. Bronchoconstriction by inhaled histamine (1 mM) was used to investigate AHR, and inflammatory cell influx was determined by bronchoalveolar lavage (BAL), both at 24 h post-challenge. A single sensitisation with 10 micro g OA did not reveal an EAR, LAR or AHR following exposure to OA. However, total and differential cell counts (eosinophils and macrophages) were significantly greater 24 h post-challenge, when compared to saline-challenged sensitised animals. The addition of a booster injection of 10 micro g revealed an EAR, but no LAR or AHR after ovalbumin inhalation. However, there was a significant cell influx. Sensitisation with 100 micro g OA (single and booster injections) revealed all four parameters of the asthmatic response (EAR, LAR, AHR and cell influx). The incorporation of the booster sensitisation injection resulted in a prolongation of the LAR, and the AHR was more pronounced and cell influx increased significantly, when compared to all other sensitisation protocols. Thus, sensitisation with 100 micro g OA (with a booster injection) can reveal an EAR, LAR, AHR and cell influx following inhalation exposure to OA (10 micro g). Cellular infiltration to the lung may be a poor marker of the asthmatic response, as a threshold level of cell influx (eosinophils) appears to be required in order to elicit the LAR and AHR. There was an association between the LAR and AHR.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Guinea Pigs; Histamine; Immunization; Macrophages; Male; Ovalbumin

The FcR common gamma-chain (FcRgamma) is an essential component of the receptors FcepsilonRI, FcgammaRI, and FcgammaRIII, which are expressed on many inflammatory cell types. The role of these receptors in the initiation or maintenance of allergic inflammation has not been well defined. FcRgamma-deficient (FcRgamma(-/-)) and control (wild-type (WT)) mice were sensitized and subsequently challenged with OVA. Following sensitization and challenge to OVA, FcRgamma-deficient (FcRgamma(-/-)) mice developed comparable levels of IgE and IgG1 as WT mice. However, numbers of eosinophils, levels of IL-5, IL-13, and eotaxin in bronchoalveolar lavage fluid, and mononuclear cell (MNC) proliferative responses to OVA were significantly reduced, as was airway hyperresponsiveness (AHR) to inhaled methacholine. Reconstitution of FcRgamma(-/-) mice with whole spleen MNC from WT mice before sensitization restored development of AHR and the numbers of eosinophils in bronchoalveolar lavage fluid; reconstitution after sensitization but before OVA challenge only partially restored these responses. These responses were also restored when FcRgamma(-/-) mice received T cell-depleted MNC, T and B cell-depleted MNC, or bone marrow-derived dendritic cells before sensitization from FcR(+/+) or FcgammaRIII-deficient but not FcRgamma(-/-) mice. The expression levels of FcgammaRIV on bone marrow-derived dendritic cells from FcR(+/+) mice were found to be low. These results demonstrate that expression of FcRgamma, most likely FcgammaRI, on APCs is important during the sensitization phase for the development of allergic airway inflammation and AHR.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Chemokine CCL11; Chemokines, CC; Cytokines; Dendritic Cells; Eosinophils; Immunoglobulin E; Immunoglobulin G; Lymphocytes; Methacholine Chloride; Mice; Mice, Mutant Strains; Ovalbumin; Pneumonia; Receptors, IgG; Respiratory Hypersensitivity

Airway inflammation and Th2 responses play central roles in allergic asthma. We have previously reported that Ganoderma tsugae supplementation could attenuate airway inflammation in the murine model. Since it remains unclear which part of the G. tsugae exerts this effect on allergic asthma in vivo, this study was meant to investigate if triterpenoid extracts have anti-inflammatory effects on airway responses and regulatory effects on Th2 responses.. BALB/c mice sensitized intraperitoneally and challenged with ovalbumin (OVA) were treated with either triterpenoid-rich extracts (TRE) of G. tsugae or prednisolone for 2 weeks. The effects of TRE on bronchial airway hyperresponsiveness (AHR), airway inflammation, serum antigen-specific antibody levels, and cytokine secretions from splenocytes were evaluated.. TRE supplementation significantly decreased AHR and reduced the total infiltrating leukocytes and eosinophils, as well as the levels of inflammatory mediators, such as interleukin (IL)-4, IL-5 and eotaxin in bronchoalveolar lavage fluid when compared with those of the control group. Lung histology also showed less cell recruitment and lung damage. TRE supplements suppressed IL-5 secretions from OVA-stimulated splenocytes, but did not affect the cell number of splenocytes, which was also reduced by prednisolone. Although OVA-specific immunoglobulin E levels were not significantly different among the groups, a lower level of OVA-specific immunoglobulin G1, another Th2-related antibody, was found in TRE and prednisolone treatment.. TRE of G. tsugae exert anti-inflammatory effects on airway responses and attenuate Th2 responses without the overall immunosuppression effects in allergic murine models of asthma.

Topics: Animals; Antibody Formation; Asthma; Bronchial Hyperreactivity; Drugs, Chinese Herbal; Female; Interleukin-5; Mice; Mice, Inbred BALB C; Ovalbumin; Prednisolone; Reishi; Spleen; Th2 Cells; Triterpenes

The relationship between airway inflammation and structural changes of airway remodeling, and their relative effects on airway function, are poorly understood. Remodeling is thought to result from chronic repetitive injury to the airway wall caused by airway inflammation; however, the mechanisms regulating remodeling changes have not been clearly defined. We examined the sequence of events in remodeling using three commonly used mouse models of allergic airways disease in which mice are exposed to nebulized ovalbumin for four consecutive days (acute), seven consecutive days (subacute), or three times a week for 6 wk (chronic). Surprisingly, we found that a very short period of exposure to ovalbumin was sufficient to elicit early changes of remodeling. Goblet cell hyperplasia and epithelial thickening were evident after just 4 d. In chronically challenged mice, these changes persisted and, in addition, subepithelial collagen deposition was significantly increased. This collagen deposition was associated with a failure to upregulate matrix metalloproteinase (MMP)-2, in conjunction with increased transforming growth factor-beta and MMP-9 expression. The relationship between inflammation, remodeling changes, and airway hyperresponsiveness (AHR) were examined. The acute and subacute models exhibited marked airway inflammation, whereas the chronic model had very modest inflammation. Conversely, airway fibrosis was only evident in the chronic model. AHR was present in all three models; however, it was significantly higher in the chronic model compared with the acute (P<0.05) and subacute (P<0.05) models. These data demonstrate that both airway inflammation and airway fibrosis may contribute to AHR, with airway fibrosis leading to the greatest increases in AHR.

Topics: Acute Disease; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Chronic Disease; Disease Models, Animal; Immunoglobulin E; Immunohistochemistry; Inflammation; Lung; Male; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Hypersensitivity; Transforming Growth Factor beta1

Prostaglandins (PGs) can enhance or suppress inflammation by acting on different receptors expressed by hematopoietic and nonhematopoietic cells. Prostaglandin D(2) binds to the D prostanoid (DP)1 and DP2 receptor and is seen as a critical mediator of asthma causing vasodilation, bronchoconstriction, and inflammatory cell influx. Here we show that inhalation of a selective DP1 agonist suppresses the cardinal features of asthma by targeting the function of lung dendritic cells (DCs). In mice treated with DP1 agonist or receiving DP1 agonist-treated DCs, there was an increase in Foxp3(+) CD4(+) regulatory T cells that suppressed inflammation in an interleukin 10-dependent way. These effects of DP1 agonist on DCs were mediated by cyclic AMP-dependent protein kinase A. We furthermore show that activation of DP1 by an endogenous ligand inhibits airway inflammation as chimeric mice with selective hematopoietic loss of DP1 had strongly enhanced airway inflammation and antigen-pulsed DCs lacking DP1 were better at inducing airway T helper 2 responses in the lung. Triggering DP1 on DCs is an important mechanism to induce regulatory T cells and to control the extent of airway inflammation. This pathway could be exploited to design novel treatments for asthma.

Topics: Alum Compounds; Animals; Asthma; Bronchial Hyperreactivity; Cell Differentiation; Cyclic AMP-Dependent Protein Kinases; Dendritic Cells; Interleukin-10; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Prostaglandin D2; Receptors, Prostaglandin; T-Lymphocytes, Regulatory

IL-13 is believed to be a central mediator of asthma, and TGF-beta1 is a key downstream mediator in the development of IL-13-mediated asthma phenotypes.. To evaluate the biological roles of basic fibroblast growth factor (FGF2) in phenotype expression in transgenic (TG) mice overexpressing lung-specific TGF-beta1, and the therapeutic effects of recombinant FGF2 in the development of asthma phenotypes.. To evaluate the roles of FGF2 in airway hyperresponsiveness (AHR) expression induced by high levels of TGF-beta1, TGF-beta1 TG (+) mice were bred with FGF2-deficient mice. To evaluate the therapeutic effects of recombinant FGF2 (rFGF2) in the development of asthma, mice were given 10 mug of rFGF2 subcutaneously once a day, 1 hour before the allergen challenge in an asthma mouse model. AHR was evaluated using noninvasive whole-body plethysmography, mucus production by diastase-resistant periodic acid Schiff (DPAS) staining, and lung inflammation using bronchoalveolar lavage (BAL) cellularity and lung histology.. AHR decreased in TGF-beta1 TG (+) mice and was accompanied by the upregulation of FGF2 mRNA expression in lung tissues, when compared with littermate wild-type control mice. Interestingly, AHR was enhanced markedly in TGF-beta1 (+) mice with homozygous FGF2 gene disruption. In an asthma mouse model, AHR, mucus production, and lung inflammation were inhibited markedly by rFGF2 treatment. This inhibition was accompanied by downregulation of the allergen-induced proliferation of T cells from regional lymph nodes.. FGF2 seems to be a key inhibitor in the development of AHR, and rFGF2 treatment constrains the development of asthma phenotypes.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Fibroblast Growth Factor 2; Inflammation Mediators; Injections, Subcutaneous; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Mucus; Ovalbumin; Recombinant Proteins; Transforming Growth Factor beta1

Pulmonary eosinophilia is one of the most consistent hallmarks of asthma. Infiltration of eosinophils into the lung in experimental asthma is dependent on the adhesion molecule vascular cell adhesion molecule-1 (VCAM-1) on endothelial cells. Ligation of VCAM-1 activates endothelial cell NADPH oxidase, which is required for VCAM-1-dependent leukocyte migration in vitro. To examine whether endothelial-derived NADPH oxidase modulates eosinophil recruitment in vivo, mice deficient in NADPH oxidase (CYBB mice) were irradiated and received wild-type hematopoietic cells to generate chimeric CYBB mice. In response to ovalbumin (OVA) challenge, the chimeric CYBB mice had increased numbers of eosinophils bound to the endothelium as well as reduced eosinophilia in the lung tissue and bronchoalveolar lavage. This occurred independent of changes in VCAM-1 expression, cytokine/chemokine levels (IL-5, IL-10, IL-13, IFNgamma, or eotaxin), or numbers of T cells, neutrophils, or mononuclear cells in the lavage fluids or lung tissue of OVA-challenged mice. Importantly, the OVA-challenged chimeric CYBB mice had reduced airway hyperresponsiveness (AHR). The AHR in OVA-challenged chimeric CYBB mice was restored by bypassing the endothelium with intratracheal administration of eosinophils. These data suggest that VCAM-1 induction of NADPH oxidase in the endothelium is necessary for the eosinophil recruitment during allergic inflammation. Moreover, these studies provide a basis for targeting VCAM-1-dependent signaling pathways in asthma therapies.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Chemokine CCL11; Chemokines, CC; Cytokines; Disease Models, Animal; Endothelium, Vascular; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; NADPH Oxidases; Ovalbumin; Pulmonary Eosinophilia; Vascular Cell Adhesion Molecule-1

Asthma is a chronic inflammatory disorder of the airways characterized by excess production of Th2 cytokines and eosinophil accumulation in the lungs. Fritillaria cirrhosa, Anemarrhena asphodeloides and Lee-Mo-Tang are well-known herbs used in oriental medicine for the treatment of asthma and bronchial inflammation. To clarify the anti-asthmatic effects of Fritillaria cirrhosa bulbus, Anemarrhena rhizoma and Lee-Mo-Tang, we examined the development of pulmonary eosinophilic accumulation, control of Th2 cytokine, immunoglobulin E (IgE) and histamine productions in a murine model of asthma. Eosinophil cell proliferation was performed by [(3)H]thymidine uptake, eosinophilic accumulation. Cell counts in bronchoalveolar lavage fluid were investigated by means of fluorescence activated cell sorter analysis and control of Th2 cytokine, IgE and histamine productions were investigated by RT-PCR and ELISA. Moreover, lung tissue was histologically analysed. The suppressive effects of Fritillaria cirrhosa bulbus, Anemarrhena rhizoma and Lee-Mo-Tang on eosinophil recruitment and airway inflammation were demonstrated throughout the reduction of eosinophil numbers. This result correlated with a marked reduction IL-5, IL-13 and IL-4 levels in the bronchoalveolar lavage fluid. Ovalbumin-specific IgE levels were also decreased in serum. Fritillaria cirrhosa bulbus, Anemarrhena rhizoma and Lee-Mo-Tang have deep inhibitory effects on airway inflammation by suppression of Th2 cytokines (IL-4, IL-5 and IL-13), IgE, histamine production, reduction eosinophilic accumulation and increase of interferon-gamma production.

Topics: Anemarrhena; Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Proliferation; Cyclosporine; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophilia; Flow Cytometry; Fritillaria; Herbal Medicine; Histamine; Immunoglobulin E; Interferon-gamma; Interleukins; Lung; Male; Mice; Ovalbumin; Phytotherapy; Plant Preparations; Reverse Transcriptase Polymerase Chain Reaction

We have recently demonstrated that naphthoquinone (NQ), one of extractable chemical compounds of diesel exhaust particles (DEP), enhances antigen-related airway inflammation with goblet cell hyperplasia in mice (Inoue et al. in Eur Respir J 209(2):259-267, 2007). Further, NQ has enhanced lung expressions of interleukin (IL)-4 and IL-5. However, the effects of NQ on other cardinal features of asthma have not been completely investigated. The aim of the present study was to evaluate the effects of NQ on airway responsiveness on the model. Vehicle, NQ, ovalbumin (OVA), or NQ + OVA was administered intratarcheally to ICR mice for 6 weeks. Twenty-four hours after the last instillation, lung histology, lung functions such as total respiratory system resistance (R) and Newtonian resistance (R (n)), and protein level of IL-13 and mRNA level for MUC5AC in the lung were examined. Repetitive exposure to NQ aggravated antigen-related lung inflammation. NQ alone enhanced R and R (n) as compared to vehicle without statistical significance. OVA alone or NQ plus OVA showed increases in R and R (n), which was prominent in NQ plus OVA (P < 0.05 vs. vehicle). Combined exposure to NQ and OVA elevated the levels of IL-13 and MUC5AC in the lung as compared with exposure to NQ or OVA alone. These results indicate that NQ can enhance airway hyperresponsiveness in the presence or absence of an antigen. Also, amplified lung expressions of IL-13 and MUC5AC might partly contribute to the deterioration of asthma features by NQ.

Topics: Animals; Antigens; Bronchial Hyperreactivity; Gene Expression Regulation; Interleukin-13; Lung; Male; Mice; Mice, Inbred ICR; Mucin 5AC; Mucins; Naphthoquinones; Ovalbumin; RNA, Messenger; Vehicle Emissions

Sphingosine-1-phosphate (S1P) has been shown to regulate numerous and diverse cell functions, including smooth muscle contraction. Here we assessed the role of S1P/Sphingosine kinase (SPK) pathway in the regulation of bronchial tone. Our objective was to determine, using an integrated pharmacologic and molecular approach, (1) the role of S1P as endogenous modulator of the bronchial tone, and (2) the linkage between S1P pathway and bronchial hyperresponsiveness. We evaluated S1P effects on isolated bronchi and whole lungs, harvested from Balb/c mice sensitized to ovalbumin (OVA) versus vehicle-treated mice, by measuring bronchial reactivity and lung resistance. We found that S1P administration on nonsensitized mouse bronchi does not cause any direct effect on bronchial tone, while a significant increase in Ach-induced contraction occurs after S1P challenge. Conversely, in OVA-sensitized mice S1P/SPK pathway triggers airway hyperesponsiveness. Indeed, S1P causes a dose-dependent contraction of isolated bronchi. Similarly, in the whole lung system S1P increased airway resistance only in OVA-sensitized mice. The action on bronchi of S1P is coupled to an enhanced expression of SPK(1) and SPK(2) as well as of S1P(2) and S1P(3) receptors. In these experiments the key role for S1P/SPK in hyperreactivity has been confirmed by pharmacologic modulation of SPKs. S1P/SPK pathway does not seem to play a major role in physiologic conditions, while it may become critical in pathologic conditions. These results open new windows for therapeutic strategies in diseases like asthma.

Topics: Acetylcholine; Animals; Bronchi; Bronchial Hyperreactivity; Cholinergic Agents; Dose-Response Relationship, Drug; Lysophospholipids; Mice; Mice, Inbred BALB C; Muscle Tonus; Muscle, Smooth; Ovalbumin; Phosphotransferases (Alcohol Group Acceptor); Receptors, Lysosphingolipid; Signal Transduction; Sphingosine

The role of macrophage migration inhibitory factor (MIF), a proinflammatory cytokine, was tested using a mouse asthma model.. One hundred and four male BALB/c mice were used in this study.. Mice were actively sensitized with an intraperitoneal injection of ovalbumin (OVA) and challenged with repeated nebulization of 1 w/v% OVA. Polyclonal anti-MIF antibody was intraperitoneally injected at 10 mg/kg during the antigen challenge period.. Bronchoalveolar lavage (BAL) was performed 8 h after the last challenge. Airway hyperresponsiveness to inhaled methacholine was measured 24 h after the last challenge.. Antigen challenge to immunized mice induced increase in inflammatory cells and concentration of Th2 cytokines in BAL fluid (BALF), and caused the development of airway hyperresponsiveness. Anti-MIF antibody significantly decreased the numbers of inflammatory cells including macrophages, eosinophils, lymphocytes and neutrophils in BALF from OVA-challenged mice. Prednisolone decreased the numbers of eosinophils, lymphocytes and neutrophils but not macrophages. Anti-MIF antibody reduced airway hyperresponsiveness. Anti-MIF antibody affected neither the cytokine levels in BALF nor the IgE levels in serum.. MIF was involved in the antigen-induced inflammatory cell accumulation in the lung and airway hyperresponsiveness without affecting immune responses.

Topics: Animals; Anti-Inflammatory Agents; Antibodies, Blocking; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Immunoglobulin E; Inflammation; Lipopolysaccharides; Macrophage Migration-Inhibitory Factors; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Plethysmography, Whole Body; Prednisolone; Respiratory Hypersensitivity; Shock, Septic

These studies were designed to assess the pharmacodynamic interaction between formoterol and beclomethasone dipropionate (BDP) in controlling the bronchoconstriction and inflammatory response induced by various challenges in guinea-pigs and rats. In anaesthetised guinea-pigs, superfusion of the formoterol/BDP combination into the tracheal lumen had significantly more effect than the single components in antagonising the bronchoconstricting and inflammatory responses to acetylcholine or ovalbumin in a standard model of airway hyper-responsiveness. After ovalbumin challenge, the combination completely protected animals from death at doses lower than those effective when given separately. The combination, at doses ineffective individually, even counteracted the development of lung oedema induced by sephadex in the rat. Finally, in tracheal strips from ovalbumin-sensitised guinea-pigs pre-treatment with BDP (30 mg kg(-1) i.m.) completely reversed the rightward shift of the formoterol dose-response curve due to beta(2)-receptor desensitisation. In conclusion, these results indicate that formoterol and BDP together induce a favourable pharmacodynamic interaction which can be considered more than additive, at least in these experimental settings.

Topics: Acetylcholine; Adrenergic beta-2 Receptor Agonists; Adrenergic beta-Agonists; Albuterol; Animals; Anti-Inflammatory Agents; Beclomethasone; Bronchial Hyperreactivity; Bronchoconstriction; Bronchodilator Agents; Dextrans; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Combinations; Drug Synergism; Ethanolamines; Formoterol Fumarate; Guinea Pigs; Inflammation; Male; Ovalbumin; Pulmonary Edema; Rats; Receptors, Adrenergic, beta-2; Trachea

TNF is thought to contribute to airway hyperreactivity (AHR) and airway inflammation in asthma. However, studies with TNF-deficient or TNF receptor-deficient mice have not produced a clear picture of the role of TNF in the AHR associated with allergic inflammation in the mouse.. We used a genetic approach to investigate the contributions of TNF to antigen-induced AHR and airway inflammation in mice on the C57BL/6 background.. We analyzed features of airway allergic inflammation, including antigen-induced AHR, in C57BL/6 wild-type and TNF(-/-) mice, using 2 different methods for sensitizing the mice to ovalbumin (OVA).. In mice sensitized to OVA administered with the adjuvant aluminum hydroxide (alum), which develop IgE-independent and mast cell-independent allergic inflammation and AHR, we found no significant differences in OVA-induced AHR in C57BL/6-TNF(-/-) versus wild-type mice. By contrast, in mice sensitized to OVA without alum, which develop allergic inflammation that is significantly mast cell-dependent, C57BL/6-TNF(-/-) mice exhibited significant reductions versus wild-type mice in OVA-induced AHR to methacholine; numbers of lymphocytes, neutrophils, and eosinophils in bronchoalveolar lavage fluid; levels of myeloperoxidase, eosinophil peroxidase, and the cytokines IL-4, IL-5, and IL-17 in lung tissue; and histologic evidence of pulmonary inflammation.. In pulmonary allergic inflammation induced in mice immunized with OVA without alum, TNF significantly contributes to several features of the response, including antigen-induced inflammation and AHR.. Our findings in mice support the hypothesis that TNF can promote the allergic inflammation and AHR associated with asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchitis; Cytokines; Disease Models, Animal; Immunoglobulin E; Lung; Mast Cells; Mice; Mice, Mutant Strains; Ovalbumin; Receptors, Tumor Necrosis Factor, Type I; Receptors, Tumor Necrosis Factor, Type II; Respiratory Hypersensitivity; Tumor Necrosis Factor-alpha

IL-10 affects dendritic cell (DC) function, but the effects on airway hyperresponsiveness (AHR) and inflammation are not defined.. We sought to determine the importance of IL-10 in regulating DC function in allergen-induced AHR and airway inflammation.. DCs were generated from bone marrow in the presence or absence of IL-10. In vivo IL-10-treated DCs from IL-10(+/+) and IL-10(-/-) donors pulsed with ovalbumin (OVA) were transferred to naive or sensitized mice before challenge. In recipient mice AHR, cytokine levels, cell composition of bronchoalveolar lavage (BAL) fluid, and lung histology were monitored.. In vitro, IL-10-treated DCs expressed lower levels of CD11c, CD80, and CD86; expressed lower levels of IL-12; and suppressed T(H)2 cytokine production. In vivo, after transfer of OVA-pulsed IL-10-treated DCs, naive mice did not have AHR, airway eosinophilia, T(H)2 cytokine increase in BAL fluid, or goblet cell metaplasia when challenged, and in sensitized and challenged mice IL-10-treated DCs suppressed these responses. Levels of IL-10 in BAL fluid and numbers of lung CD4(+)IL-10(+) T cells were increased in mice that received OVA-pulsed IL-10-treated DCs. Transfer of IL-10-treated DCs from IL-10-deficient mice were ineffective in suppressing the responses in sensitized and challenged mice.. These data demonstrate that IL-10-treated DCs are potent suppressors of the development of AHR, inflammation, and T(H)2 cytokine production; these regulatory functions are at least in part through the induction of endogenous (DC) production of IL-10.. Modification of DC function by IL-10 can attenuate lung allergic responses, including the development of AHR.

Topics: Adoptive Transfer; Animals; Bronchial Hyperreactivity; Cytokines; Dendritic Cells; Female; Flow Cytometry; Immunohistochemistry; Inflammation; Interleukin-10; Mice; Ovalbumin

Air pollution contributes to both exacerbation and development of bronchial asthma. Studies showed that coexposure to air pollution directly promotes sensitization to inhaled allergen in neonatal mice. The aim of this study was to investigate whether prenatal exposure to air pollution could also increase susceptibility to development of asthma in early life. Pregnant female BALB/c mice were exposed to aerosolized leachate of residual oil fly ash (ROFA, 50 mg/ml, 30 min) at 5, 3, and 1 d before delivery. Offspring were treated once at 3 d of age with ovalbumin (OVA, 5 mug) and alum (ip), an intentionally suboptimal dose for sensitization, exposed to aerosolized OVA (1%, 10 min) at 12-14 d or 32-35 d of age, and evaluated 2 d after the final exposure. The offspring of ROFA-exposed mothers (ROFA group) revealed increasing airway hyperresponsiveness (higher enhanced pause [Penh] to methacholine challenge) and elevated substantial numbers of eosinophils in the bronchoalveolar lavage flued (BALF). Histopathology revealed prominent inflammation in the lungs of ROFA group and showed increased allergen-specific IgE and IgG1 levels. Their cultured splenocytes showed an enhanced interleukin (IL)-4/interferon (IFN)-gamma cytokine, indicating Th2 skewed immunity. Data indicate that exposure of pregnant female mice to an air pollutant aerosol increased asthma susceptibility in their offspring.

Topics: Administration, Inhalation; Aerosols; Air Pollutants; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Carbon; Cells, Cultured; Coal Ash; Disease Susceptibility; Eosinophils; Female; Immunoglobulin E; Immunoglobulin G; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Particulate Matter; Pregnancy; Prenatal Exposure Delayed Effects; Respiratory Hypersensitivity; Spleen

These studies were conducted to investigate the role of dermal exposure to perfluorooctanoic acid (PFOA), a known immunosuppressant, on the hypersensitivity response to ovalbumin (OVA) in a murine model of asthma. PFOA has had widespread use as a carpet and fabric protectant. BALB/c mice were exposed dermally, on the dorsal surface of each ear, to concentrations of PFOA ranging from 0.01 to 1.5% (applied dose 0.25-50 mg/kg) for 4 days. In hypersensitivity studies, mice were also ip injected with 7.5 microg OVA and 2 mg alum on days 1 and 10 and in some studies challenged with 250 microg OVA by pharyngeal aspiration on days 17 and 26. Following exposure to PFOA, an increase in liver weights and a decrease in thymus and spleen weights and cellularities were observed. Similar immunomodulatory trends were demonstrated in mice coadministered PFOA and OVA. Compared to the OVA alone-exposed animals, an increase in total IgE was demonstrated when mice were coexposed to OVA and concentrations of PFOA ranging from 0.75 to 1.5%, while the OVA-specific IgE response peaked with 0.75% PFOA coexposure (p < or = 0.05). OVA-specific airway hyperreactivity was increased in the 1.0% PFOA coexposed group (p < or = 0.05), with an increased pleiotropic cell response characterized by eosinophilia and mucin production, in animals coexposed to concentrations of PFOA up to 1.0%, as compared to the OVA alone-exposed animals. In a murine model, PFOA was demonstrated to be immunotoxic following dermal exposure, with an enhancement of the hypersensitivity response to OVA, suggesting that PFOA exposure may augment the IgE response to environmental allergens.

Topics: Administration, Topical; Animals; Antigens; Asthma; Body Weight; Bronchial Hyperreactivity; Caprylates; Dose-Response Relationship, Drug; Enzyme-Linked Immunosorbent Assay; Female; Fluorocarbons; Hypersensitivity; Immunoglobulin E; Immunosuppressive Agents; Lung; Mice; Mice, Inbred BALB C; Organ Size; Ovalbumin; Phenotype; Respiratory Hypersensitivity

In asthmatic mice, dexamethasone (30.0 mg/kg) was administered orally once daily on Days 24-27. One hour after dexamethasone on Day 25-27, the mice were exposed to ovalbumin aerosols. Twenty-eight days after the initial ovalbumin immunization, we found that dexamethasone reduced methacholine-induced pulmonary gas trapping and inhibited bronchoalveolar lavage eosinophils and neutrophils. However, five days after the last dose of dexamethasone and last ovalbumin aerosol exposure in other asthmatic mice, the airway obstructive response to methacholine was exacerbated in dexamethasone-treated mice compared to vehicle-treated mice on Day 32. Further, eosinophils, but not neutrophils, were still inhibited after cessation of dexamethasone. Thus, discontinuing dexamethasone worsened methacholine-induced pulmonary gas trapping of asthmatic mice in the absence of eosinophilic airway inflammation.

Topics: Administration, Oral; Airway Resistance; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Dexamethasone; Disease Models, Animal; Drug Administration Schedule; Eosinophils; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Pulmonary Eosinophilia; Recurrence

Environmental tobacco smoke (ETS) can increase asthma symptoms and the frequency of asthma attacks. However, the contribution of ETS to airway remodeling in asthma is at present unknown. In this study, we have used a mouse model of allergen-induced airway remodeling to determine whether the combination of chronic exposure to ETS and chronic exposure to OVA allergen induces greater levels of airway remodeling than exposure to either chronic ETS or chronic OVA allergen alone. Mice exposed to chronic ETS alone did not develop significant eosinophilic airway inflammation, airway remodeling, or increased airway hyperreactivity to methacholine. In contrast, mice exposed to chronic OVA allergen had significantly increased levels of peribronchial fibrosis, increased thickening of the smooth muscle layer, increased mucus, and increased airway hyperreactivity which was significantly enhanced by coexposure to the combination of chronic ETS and chronic OVA allergen. Mice coexposed to chronic ETS and chronic OVA allergen had significantly increased levels of eotaxin-1 expression in airway epithelium which was associated with increased numbers of peribronchial eosinophils, as well as increased numbers of peribronchial cells expressing TGF-beta1. These studies suggest that chronic coexposure to ETS significantly increases levels of allergen-induced airway remodeling (in particular smooth muscle thickness) and airway responsiveness by up-regulating expression of chemokines such as eotaxin-1 in airway epithelium with resultant recruitment of cells expressing TGF-beta1 to the airway and enhanced airway remodeling.

Topics: Actins; Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Chemokine CCL11; Chemokines, CC; Collagen; Connective Tissue Growth Factor; Eosinophilia; Immediate-Early Proteins; Intercellular Signaling Peptides and Proteins; Interleukin-5; Mice; Mice, Inbred BALB C; Mucus; Muscle, Smooth; Ovalbumin; Tobacco Smoke Pollution; Transforming Growth Factor beta1

Allergic asthma is characterized by airway inflammation initiated by adaptive immune responses to aeroallergens. Recent data suggest that severe asthma may be a different form of asthma rather than an increase in asthma symptoms and that innate immune responses to LPS can modulate adaptive immune responses to allergens. In this study, we evaluated the hypothesis that airway exposure to different doses of LPS induces different form of asthma. Our study showed that neutrophilic inflammation and IFN-gamma expression were higher in induced sputum from severe asthma patients than from mild to moderate asthmatics. Animal experiments indicated that allergen sensitization with low-dose LPS (0.1 microg) induced type 2 asthma phenotypes, i.e., airway hyperresponsiveness, eosinophilic inflammation, and allergen-specific IgE up-regulation. In contrast, allergen sensitization with high-dose LPS (10 microg) induced asthma phenotypes, i.e., airway hyperresponsiveness and noneosinophilic inflammation that were not developed in IFN-gamma-deficient mice, but unaffected in the absence of IL-4. During the allergen sensitization period, TNF-alpha expression was found to be enhanced by both low- and high-dose LPS, whereas IL-12 expression was only enhanced by high-dose LPS. Interestingly, the asthma phenotypes induced by low-dose LPS, but not by high-dose LPS, were completely inhibited in TNF-alpha receptor-deficient mice, whereas the asthma phenotypes induced by high-dose LPS were abolished in the homozygous null mutation of the STAT4 gene. These findings suggest that airway exposure levels of LPS induces different forms of asthma that are type 1 and type 2 asthma phenotypes by high and low LPS levels, respectively.

Topics: Adult; Aged; Animals; Asthma; Bronchial Hyperreactivity; Female; Humans; Interferon-gamma; Interleukin-12; Lipopolysaccharides; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Middle Aged; Ovalbumin; Receptors, Tumor Necrosis Factor; RNA, Messenger; Signal Transduction; STAT4 Transcription Factor; Th1 Cells; Th2 Cells; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha

Microbial intestinal colonization in early in life is regarded to play a major role for the maturation of the immune system. Application of non-pathogenic probiotic bacteria during early infancy might protect from allergic disorders but underlying mechanisms have not been analysed so far.. The aim of the current study was to investigate the immune effects of oral application of probiotic bacteria on allergen-induced sensitization and development of airway inflammation and airway hyper-reactivity, cardinal features of bronchial asthma.. Newborn Balb/c mice received orally 10(9) CFU every second day either Lactobacillus rhamnosus GG or Bifidobacterium lactis (Bb-12) starting from birth for consecutive 8 weeks, during systemic sensitization (six intraperitoneal injections, days 29-40) and airway challenge (days 54-56) with ovalbumin.. The administration of either Bb-12 or LGG suppressed all aspects of the asthmatic phenotype: airway reactivity, antigen-specific immunoglobulin E production and pulmonary eosinophilia (mean: 137 vs. 17 and 13 cellsx10(3)/mL, respectively). Antigen-specific recall proliferation by spleen cells and T-helper type 2 cytokine production (IL-4, IL-5 and IL-10) by mesenteric lymph node cells also showed significant reduction, while TGF production remained unchanged. Oral LGG administration particularly suppressed allergen-induced proliferative responses and was associated with an increase in numbers of TGF-beta-secreting CD4+/CD3+ T cells in mesenteric lymph nodes (6.5, 16.7%) as well as nearly 2-fold up-regulation of Foxp3-expressing cells in peribronchial lymph nodes.. Neonatal application of probiotic bacteria inhibits subsequent allergic sensitization and airway disease in a murine model of asthma by induction of T regulatory cells associated with increased TGF-beta production.

Topics: Allergens; Animals; Asthma; Bifidobacterium; Bronchial Hyperreactivity; Cell Proliferation; Cytokines; Disease Models, Animal; Eosinophilia; Female; Forkhead Transcription Factors; Immunoglobulin E; Immunoglobulin G; Lacticaseibacillus rhamnosus; Lymph Nodes; Mice; Mice, Inbred BALB C; Ovalbumin; Probiotics; Reverse Transcriptase Polymerase Chain Reaction; Spleen; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Up-Regulation

A small GTPase, Rho, and its target molecule, Rho-kinase, play an important role in the cell functions, including contractility, chemotaxis, adhesion, and migration. It is generally considered that eosinophilic inflammation and hyper-reactivity to methacholine in airways are fundamental to the pathophysiology of bronchial asthma.. This study was designed to determine whether the Rho/Rho-kinase pathways are involved in the eosinophil recruitment and airway hyper-reactivity. We investigated inhibitory effects of fasudil, a specific inhibitor of Rho-kinase, on acute allergic inflammation in mice.. BALB/c mice were sensitized and challenged with ovalbumin (OVA). OVA-challenged mice were treated orally with fasudil (3, 10, 30 mg/kg) or saline before each OVA challenge. Total cell counts, differential cell counts, cytokines, and chemokines levels were measured in bronchoalveolar lavage (BAL), and lungs were examined histologically. Moreover, respiratory resistance in response to methacholine was measured.. When fasudil was administrated to OVA-challenged mice, increased cell numbers of total cells and eosinophils were significantly attenuated in a dose-dependent manner. However, inflammatory cells other than eosinophils were not affected by fasudil. Fasudil caused a dose-dependent inhibition in increased levels of IL-5, IL-13, and eotaxin in BAL fluid by OVA challenges. Histological analysis of the airways revealed that both infiltration of inflammatory cells and goblet cell hyperplasia were significantly suppressed in fasudil treatment. Furthermore, fasudil significantly suppressed the augmented responsiveness to methacholine induced by OVA challenges.. Oral administration of fasudil inhibits eosinophil recruitment, goblet cell hyperplasia and airway hyper-reactivity by allergen challenges. These effects of this agent may be mediated by suppressing a chemokine and cytokines related to the pathophysiology of bronchial asthma such as eotaxin, IL-5, and IL-13. Our findings provide evidence that inhibition of the Rho/Rho-kinase pathway may be beneficial for bronchial asthma.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokines, CC; Dose-Response Relationship, Drug; Drug Evaluation, Preclinical; Eosinophilia; Female; Interleukins; Intracellular Signaling Peptides and Proteins; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Protein Kinase Inhibitors; Protein Serine-Threonine Kinases; rho-Associated Kinases

The role of endothelin, PAF and thromboxane A2 in airway hyperreactivity (AHR) to carbachol induced by ovalbumin sensitization and challenge in Balb/c mice was investigated. Ovalbumin sensitization and challenge induced significant AHR to carbachol in actively sensitized and challenged mice. Treatment of these mice with the PAF antagonist CV-3988 (10 microg kg(-1), i.v.) completely abolished OVA-induced AHR to carbachol. Treatment of sensitized mice with the TxA2 antagonist L-654,664 (1 mg kg(-1), i.v.) partially blocked the induction of AHR in OVA-challenged mice. The intranasal administration of 50 pmol of the ET(A) receptor antagonist BQ-123 had no effect on the PIP but produced a significant reduction at the dose of 100 pmol. The intravenous administration of BQ-123 (100 pmol) reduced the PIP only at the highest doses of carbachol. The ET(B) receptor antagonist BQ-788 administered either via the intranasal or intravenous route had no effect on the PIP at the dose of 100 pmol. Naïve mice treated with either U-44069 (25 or 100 microg kg(-1), i.v.), endothelin-1 (100 pmol, intranasally) or the ET(B) receptor agonist IRL-1620 (100 pmol, intranasally) showed a marked increase in airway reactivity to carbachol. These results suggest an important role for endothelin, PAF and thromboxane A2 in AHR in mice actively sensitized and challenged with ovalbumin.

Topics: Animals; Antihypertensive Agents; Bronchial Hyperreactivity; Carbachol; Endothelin B Receptor Antagonists; Endothelin-1; Endothelins; Male; Mice; Mice, Inbred BALB C; Oligopeptides; Ovalbumin; Peptide Fragments; Phospholipid Ethers; Piperidines; Platelet Activating Factor; Platelet Aggregation Inhibitors; Prostaglandin Endoperoxides, Synthetic; Receptor, Endothelin B; Thromboxane A2

Enhanced expression of the suppressor of cytokine signalling (SOCS)-5 might be of therapeutic benefit for T-helper type 2 (Th2) dominant diseases, as its expression is reported to result in a reduction of Th2 differentiation in vitro due to the inhibition of IL-4 signalling.. To investigate the regulatory role of SOCS-5 in vivo, we explored the phenotype of an experimental asthma model developed in SOCS-5 transgenic (Tg) mice.. The SOCS-5 Tg mice or wild-type (WT) mice were sensitized and repeatedly challenged with ovalbumin (OVA). We examined bronchoalveolar lavage fluid (BALF), lung specimens, and airway hyperresponsiveness (AHR) to methacholine.. The production of IFN-gamma by CD4(+) T cells from unprimed SOCS-5 Tg mice was significantly increased in comparison with unprimed wild-type mice, indicating that SOCS-5 Tg mice have a Th1-polarizing condition under natural conditions. However, in an asthma model, significantly more eosinophils in the airways and higher levels of IL-5 and IL-13 in BALF were observed in the SOCS-5 Tg than the wild-type mice. AHR in the asthma model of SOCS-5 Tg was also more enhanced than that of wild-type mice. OVA-stimulated CD4(+) T cells from the primed SOCS-5 Tg mice produced significantly more IL-5 and IL-13 than CD4(+) T cells from wild-type mice.. Our results demonstrate that the overexpression of SOCS-5 does not inhibit Th2 response, but rather augments the phenotype of the asthma model in vivo. This finding throws into question the therapeutic utility of using enhancement of SOCS-5 expression for Th2-dominant disease.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Disease Models, Animal; Eosinophilia; Interferon-gamma; Lung; Mice; Mice, Inbred C57BL; Mice, Transgenic; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; Suppressor of Cytokine Signaling Proteins; Th2 Cells

Mast cells, IgE, and TNF, which have been implicated in human atopic asthma, contribute significantly to the allergic airway inflammation induced by ovalbumin (OVA) challenge in mice sensitized with OVA without alum. However, it is not clear to what extent mast cells represent a significant source of TNF in this mouse model.. We investigated the importance of mast cell-derived TNF in a mast cell-dependent model of OVA-induced airway hyperreactivity (AHR) and allergic airway inflammation.. Features of this model of airway inflammation were analyzed in C57BL/6J-wild-type mice, mast cell-deficient C57BL/6J-Kit(W-sh)(/W-sh) mice, and C57BL/6J Kit(W-sh/W-sh) mice that had been systemically engrafted with bone marrow-derived cultured mast cells from C57BL/6J-wild-type or C57BL/6J-TNF(-/-) mice.. Ovalbumin-induced AHR and airway inflammation were significantly reduced in mast cell-deficient Kit(W-sh/W-sh) mice versus wild-type mice. By contrast, Kit(W-sh/W-sh) mice that had been engrafted with wild-type but not with TNF(-/-) bone marrow-derived cultured mast cells exhibited responses very similar to those observed in wild-type mice. Mast cells and mast cell-derived TNF were not required for induction of OVA-specific memory T cells in the sensitization phase, but significantly enhanced lymphocyte recruitment and T(H)2 cytokine production in the challenge phase.. Mast cell-derived TNF contributes significantly to the pathogenesis of mast cell-dependent and IgE-dependent, OVA-induced allergic inflammation and AHR in mice, perhaps in part by enhancing lymphocyte recruitment and T(H)2 cytokine production.. Our findings in mice support the hypothesis that mast cell-derived TNF can promote allergic inflammation and AHR in asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Eosinophil Peroxidase; Lung; Mast Cells; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Peroxidase; Pneumonia; Th2 Cells; Tumor Necrosis Factor-alpha

Asthma is a common chronic inflammatory disease regulated by coordination of T-helper cell type 2 (Th2) cytokines and inflammatory signal molecules. Additionally, oxidative stress may play an important role in airway inflammation such as eosinophilia, mucus hypersecretion, and airway hyperresponsiveness (AHR). In the present report, we investigated whether anthocyanins would reduce airway inflammation in a mouse asthma model immunized and challenged with ovalbumin (OVA). OVA inhalation elicited inflammatory responses characterized by eosinophilia and increased lipid hydroperoxide (LPO) in bronchoalveolar lavage (BAL) fluid, enhanced pause (Penh), increased glycoprotein and proliferating cell nuclear antigen (PCNA) expressions in mucus hypersecretion, and an increased expression of various cytokines and cyclooxygenase (COX) 2 in lung tissues. All parameters were attenuated in a dose-dependant manner by the administration of anthocyanins. These results suggest that anthocyanins may attenuate the development of asthma by downregulating Th2 cytokines, proinflammatory cytokines, and COX-2. Our findings suggest that anthocyanins have positive contributions as a dietary supplement for the prevention of asthma.

Topics: Animals; Anthocyanins; Anti-Asthmatic Agents; Antioxidants; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cyclooxygenase 2; Immunohistochemistry; Interleukins; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Oxidative Stress; RNA, Messenger; Tumor Necrosis Factor-alpha

Intracellular signaling pathways that converge on Smad 3 are used by both TGF-beta and activin A, key cytokines implicated in the process of fibrogenesis. To determine the role of Smad 3 in allergen-induced airway remodeling, Smad 3-deficient and wild-type (WT) mice were sensitized to OVA and challenged by repetitive administration of OVA for 1 mo. Increased levels of activin A and increased numbers of peribronchial TGF-beta1(+) cells were detected in WT and Smad 3-deficient mice following repetitive OVA challenge. Smad 3-deficient mice challenged with OVA had significantly less peribronchial fibrosis (total lung collagen content and trichrome staining), reduced thickness of the peribronchial smooth muscle layer, and reduced epithelial mucus production compared with WT mice. As TGF-beta and Smad 3 signaling are hypothesized to mediate differentiation of fibroblasts to myofibroblasts in vivo, we determined the number of peribronchial myofibroblasts (Col-1(+) and alpha-smooth muscle actin(+)) as assessed by double-label immunofluorescence microscopy. Although the number of peribronchial myofibroblasts increased significantly in WT mice following OVA challenge, there was a significant reduction in the number of peribronchial myofibroblasts in OVA-challenged Smad 3-deficient mice. There was no difference in levels of eosinophilic airway inflammation or airway responsiveness in Smad 3-deficient compared with WT mice. These results suggest that Smad 3 signaling is required for allergen-induced airway remodeling, as well as allergen-induced accumulation of myofibroblasts in the airway. However, Smad 3 signaling does not contribute significantly to airway responsiveness.

Topics: Activins; Allergens; Animals; Azo Compounds; Bronchial Hyperreactivity; Cell Movement; Collagen; Eosine Yellowish-(YS); Fibroblasts; Lung; Methyl Green; Mice; Mice, Inbred C57BL; Mice, Knockout; Mucus; Muscle, Smooth; Ovalbumin; Pulmonary Eosinophilia; Respiratory Mucosa; Signal Transduction; Smad3 Protein

Sublingual immunotherapy is a noninvasive and efficacious treatment of type I respiratory allergies. A murine model of sublingual immunotherapy is needed to understand better the immune mechanisms involved in successful immunotherapy and to assess second-generation candidate vaccines.. Herein, we developed a therapeutic murine model of sublingual immunotherapy in which we document the value of mucoadhesive formulations to enhance treatment efficacy.. BALB/c mice were sublingually treated with soluble or formulated ovalbumin before or after sensitization with ovalbumin. Airways hyperresponsiveness and lung inflammation were assessed by whole-body plethysmography and lung histology, respectively. Humoral and cellular immune responses were monitored by ELISA and ELISPOT techniques.. Prophylactic sublingual administration of ovalbumin completely prevents airways hyperresponsiveness as well as IL-5 secretion and IgE induction. Therapeutic administration of ovalbumin as a solution via either the sublingual or oral route has a limited efficacy. In contrast, sublingual application of ovalbumin formulated with maltodextrin to enhance mucosal adhesion results in a major reduction of established airways hyperresponsiveness, lung inflammation, and IL-5 production in splenocytes. This mucoadhesive formulation significantly enhances ovalbumin-specific T-cell proliferation in cervical but not mesenteric lymph nodes, and IgA production in the lungs.. A mucoadhesive maltodextrin formulation of ovalbumin enhances priming of the local mucosal immune system and tolerance induction via the sublingual route.. Mucoadhesive formulations offer the opportunity to improve dramatically sublingual immunotherapy in human beings, most particularly by simplifying immunization schemes.

Topics: Administration, Sublingual; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchitis; Cell Proliferation; Female; Immunization; Immunoglobulin A; Immunoglobulin E; Immunotherapy; Interleukin-10; Interleukin-5; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Mouth Mucosa; Ovalbumin; Respiratory Mucosa; Spleen; T-Lymphocytes; Tissue Adhesives; Treatment Outcome

Based on epidemiological data, the hygiene hypothesis associates poor hygienic living conditions during childhood with a lower risk for the development of allergic diseases such as bronchial asthma. The role of viral infections, and especially of viral TLR ligands, within this context remains to be clarified. Viral TLR ligands involve dsRNA and ssRNA which are recognized by TLR-3 or TLR-7, respectively. In this study, we evaluated the impact of TLR-3 or TLR-7 activation on experimental asthma in mice. Systemic application of the synthetic TLR-3 or TLR-7 ligands polycytidylic-polyinosinic acid (p(I:C)) or R-848, respectively, during the sensitization phase prevented the production of OVA-specific IgE and IgG1 Abs and subsequently abolished all features of experimental asthma including airway hyperresponsiveness and allergic airway inflammation. Furthermore, administration of p(I:C) or R-848 to animals with already established primary allergic responses revealed a markedly reduced secondary response following allergen aerosol rechallenges. In contrast to wild-type animals, application of p(I:C) or R-848 to IL-12p35(-/-) mice had no effect on airway inflammation, goblet cell hyperplasia, and airway hyperresponsiveness. However, in the absence of IL-12, the numbers of eosinophils and lymphocytes in bronchoalveolar lavage fluids were still significantly reduced. These partial effects could also be abolished by neutralizing anti-IL-10 Abs in IL-12p35(-/-) mice. These data indicate that TLR-3 or TLR-7 activation by viral TLR ligands has both preventive as well as suppressive effects on experimental asthma which is mediated by the additive effects of IL-12 and IL-10.

Topics: Animals; Antibodies; Antiviral Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Female; Imidazoles; Immunoglobulin E; Immunoglobulin G; Immunosuppression Therapy; Interleukin-10; Interleukin-12; Interleukin-12 Subunit p35; Ligands; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Mutant Strains; Ovalbumin; Poly I-C; Toll-Like Receptor 3; Toll-Like Receptor 7

Repeated airway exposure to wood dust has been reported to cause adverse respiratory effects such as asthma and chronic bronchitis. In our recent study, we found that exposure of mice to oak dust induced more vigorous lung inflammation compared to birch dust exposure. In the present study, we assessed the immunomodulatory effects of repeated intranasal exposure to oak dust both in nonallergic and in ovalbumin-sensitized, allergic mice. Allergen-induced influx of eosinophils and lymphocytes was seen in the lungs of allergic mice. Oak dust exposure elicited infiltration of neutrophils, lymphocytes, and macrophages in nonallergic mice. Interestingly, oak dust-induced lung neutrophilia as well as oak dust-induced production of the proinflammatory cytokine TNF-alpha and chemokine CCL3 were significantly suppressed in allergic mice. On the other hand, allergen-induced expression of IL-13 mRNA and protein was significantly reduced in oak dust-exposed allergic mice. Finally, allergen-induced airway hyperreactivity to inhaled metacholine was significantly suppressed in oak dust-exposed allergic mice. The present results suggest that repeated airway exposure to oak dust can regulate pulmonary inflammation and airway responses depending on the immunological status of the animal.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Chemokine CCL3; Chemokine CCL4; Chemokines, CC; Disease Models, Animal; Dust; Female; Inhalation Exposure; Interleukin-13; Leukocytes; Lung; Macrophage Inflammatory Proteins; Macrophages; Methacholine Chloride; Mice; Mice, Inbred BALB C; Neutrophil Infiltration; Ovalbumin; Quercus; RNA, Messenger; Tumor Necrosis Factor-alpha; Wood

Although eosinophils play an essential role in allergic inflammation, their role has recently been under controversy. Epidemic studies suggest that hypereosinophilia induced by parasite infection could suppress subsequent Ag sensitization, although the mechanism has not been fully clarified. In this study, we investigated whether eosinophils could suppress the Ag-specific immune response in the airway. BALB/c mice were sensitized and airway challenged with OVA. Systemic hypereosinophilia was induced by delivery of an IL-5-producing plasmid. IL-5 gene delivery suppressed the Ag-specific proliferation and cytokine production of CD4+ T cells in the spleen. IL-5 gene delivery before OVA sensitization significantly suppressed airway eosinophilia and hyperresponsiveness provoked by subsequent OVA airway challenge, while delivery during the OVA challenge did not suppress them. This IL-5-induced immune suppression was abolished in eosinophil-ablated mice, suggesting an essential role of eosinophils. IL-5 treatment increased the production of TGF-beta1 in the spleen, and we demonstrated that the main cellular source of TGF-beta1 production was eosinophils, using eosinophil-ablated mice and depletion study. TGF-beta1, but not IL-5 itself, suppressed the Ag-specific immune response of CD4+ T cells in vitro. Furthermore, IL-5 treatment enhanced phosphorylation of Smad2 in CD4+ T cells. Finally, a TGF-beta type I receptor kinase inhibitor restored this IL-5-induced immune suppression both in vitro and in vivo. These results suggest that IL-5-induced hypereosinophilia could suppress sensitization to Ag via a TGF-beta-dependent mechanism, thus suppressed allergic airway inflammation. Therefore, hypereosinophilia could reveal an immunosuppressive effect in the early stage of Ag-induced immune response.

Topics: Animals; Antigens; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cells, Cultured; Eosinophils; Gene Transfer Techniques; Growth Inhibitors; Hypereosinophilic Syndrome; Immunosuppressive Agents; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Mice, Transgenic; Mucus; Ovalbumin; Plasmids; Spleen; Transforming Growth Factor beta

The incidence of asthma has increased the world over, and current therapies for the disease suffer from potential side-effects. This has created an opportunity to develop novel therapeutic approaches. Here, the anti-inflammatory activity of choline was investigated in a mouse model of allergic airway inflammation. Choline (1 mg.kg(-1)) was administered via oral gavage or intranasally before and after ovalbumin (OVA) challenge in sensitised mice. Airway hyperresponsiveness (AHR) to methacholine was measured in the mice by whole-body plethysmography. Type-2 T-helper cell cytokine and leukotriene levels were estimated in bronchoalveolar lavage fluid (BALF) and spleen culture supernatant by ELISA. Eosinophil peroxidase activity was also determined in the BALF supernatant. Choline treatment in sensitised mice before OVA challenge via oral/intranasal routes significantly inhibited eosinophilic airway inflammation and eosinophil peroxidase activity. It also reduced immunoglobulin E and G1 production and inhibited the release of type-2 T-helper cell cytokines and leukotrienes. However, the development of AHR was prevented effectively by intranasal choline treatment. Most importantly, choline treatment after OVA challenge by both routes could reverse established asthmatic conditions in mice by inhibiting AHR, eosinophilic airway inflammation and other inflammatory parameters. This study provides a new therapeutic approach for controlling as well as preventing asthma exacerbations.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Choline; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophil Peroxidase; Eosinophils; Female; Inflammation; Lipotropic Agents; Mice; Mice, Inbred BALB C; Ovalbumin; Spleen

Although respiratory viral infections in early childhood can enhance the development of airway allergen sensitization, the exact mechanisms of the effects of viral infections on the adaptive immune response to inhaled allergens are controversial.. We sought to evaluate the effects of double-stranded RNA (dsRNA) on airway sensitization to inhaled allergens.. Novel mouse models were created through simultaneous airway sensitization to an allergen and low or high doses of dsRNA. The mouse models were applied to Toll-like receptor 3-, IL-13-, IL-4-, signal transducer and activator of transcription (STAT) 6-, IFN-gamma-, and T-box expressed in T cells (T-bet)-deficient mice to evaluate underlying pathophysiologic mechanisms in the development of allergic lung inflammation.. We found that airway allergen sensitization with dsRNA induced lung inflammation that was not present in Toll-like receptor 3-deficient mice. Moreover, lung inflammation enhanced by low-dose dsRNA was impaired in IL-13-deficient mice, whereas lung inflammation by high-dose dsRNA was impaired in IFN-gamma-deficient mice. The models also demonstrated that low-dose dsRNA enhanced IL-4 expression during allergen sensitization and that inflammation enhanced by low-dose dsRNA was not present in IL-4- or STAT6-deficient mice. In contrast, the present study showed that high-dose dsRNA enhanced IFN-gamma expression during allergen sensitization and that the development of lung inflammation enhanced by high-dose dsRNA was impaired in T-bet-deficient mice.. These findings suggest that airway allergen exposure during respiratory viral infections might induce asthma induced by both T(H)1 and T(H)2 immune responses to inhaled allergens.. Targeting both T(H)1 and T(H)2 lung inflammation might be important in the treatment of virus-associated asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Interferon-gamma; Interleukin-13; Interleukin-4; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Poly I-C; RNA, Double-Stranded; Signal Transduction; STAT6 Transcription Factor; Th1 Cells; Th2 Cells; Toll-Like Receptor 3

Controversy still exists as to whether or not inhaled beta (2)-adrenoceptor agonists and corticosteroids act synergistically in vivo. In this study, we have used a murine model of lung inflammation to study the synergistic effect of an inhaled beta (2)-adrenoceptor agonist (formoterol) and an inhaled corticosteroid (mometasone).. Actively sensitized mice were challenged with aerosolized ovalbumin, once a day, for three consecutive days. Three days after the last of the three challenges, a final allergen challenge was given. Allergen-induced increase in Penh was measured 4 h after the last challenge. A day after the last challenge, increased airway sensitivity to aerosolized methacholine was demonstrated and this was concomitant with an influx of inflammatory cells in the bronchoalveolar lavage fluids.. Mometasone (0.1 to 3 mg kg(-1)) given intranasally either an hour before or after the last allergen challenge, dose-dependently inhibited all parameters. When given intranasally either one or three hours after the last allergen challenge, but not an hour before, formoterol (1.5 to 150 microg kg(-1)) also dose-dependently inhibited most of the parameters to different degree. A synergistic effect on the allergen-induced increase in Penh was demonstrated for mometasone and formoterol given in combination, an hour after the challenge, at the following doses: mometasone/formoterol (in microg kg(-1)) 1/10, 1/100, 5/10, and 5/100.. Our results support the hypothesis that when given as a fixed combination, inhaled corticosteroid and beta (2)-adrenoceptor agonist act synergistically in vivo.

Topics: Administration, Intranasal; Adrenergic beta-Agonists; Animals; Anti-Allergic Agents; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Combinations; Drug Synergism; Ethanolamines; Female; Formoterol Fumarate; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mometasone Furoate; Ovalbumin; Pregnadienediols; Respiratory Function Tests; Respiratory Hypersensitivity; Time Factors

Infection with influenza virus has been associated with seemingly opposing effects on the development of asthma. However, there are no data about the effects of mucosal vaccination with inactivated influenza on the inception of allergic asthma.. To assess the immunological effects of inhaled inactivated influenza vaccine, using two different types of flu vaccines, on the inception of allergic sensitization and allergen-mediated airway disease in a mouse model.. BALB/c mice were intranasally or intratracheally vaccinated with whole or split influenza virus vaccine (days -1 or -1, 27) before systemic sensitization with ovalbumin (OVA) (days 1, 14) and repeated airway allergen challenges (days 28-30). Allergen sensitization (IgE serum levels), airway inflammation (differential cells in bronchoalveolar lavage fluid) and airway hyper-reactivity (AHR) (in vivo lung function) were analysed.. The intranasal instillation of whole influenza vaccine before allergen sensitization significantly reduced the serum levels of total and OVA-specific IgE as well as allergen-induced AHR. Prevention was due to an allergen-specific shift from a predominant T helper (Th)2- towards a Th1-immune response. Application of split influenza vaccine did not show the same preventive effect.. Intranasal administration of inactivated whole influenza vaccine reduced subsequent allergen sensitization and prevented allergen-induced AHR. Our results show that the composition of the influenza vaccine has a major influence on subsequent development of allergen-induced sensitization and AHR, and suggest that mucosal inactivated whole influenza vaccination may represent a step towards the development of a preventive strategy for atopic asthma.

Topics: Administration, Intranasal; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Female; Influenza A virus; Influenza Vaccines; Mice; Mice, Inbred BALB C; Ovalbumin; Th1 Cells; Th2 Cells; Vaccines, Inactivated

Asthma is an inflammatory lung disease, in which CD1d-restricted natural killer T (NKT) cells play an important pathogenic role. Also, recent reports indicated that a cytokine, thymic stromal lymphopoietin (TSLP), is essential for the development of antigen-induced asthma. Here we examined the relationship between NKT cells and TSLP in a mouse model of asthma. NKT cells express TSLP receptor as well as IL-7 receptor alpha-chain. TSLP acts on NKT cells to preferentially increase their IL-13 production but not IFN-gamma and IL-4. In an allergen-induced asthma model, the development of airway hyperreactivity, a cardinal feature of asthma, was increased in TSLP transgenic mice, whereas this effect was not observed in TSLP transgenic mice lacking NKT cells. Interestingly, in the NKT cell-lacking TSLP transgenic mice, pulmonary eosinophilia and increase in IgE did not improve. Pulmonary lymphocytes from the NKT cell-lacking TSLP transgenic mice produced much less IL-13 upon CD3 stimulation than those from NKT cell-competent TSLP transgenic mice. These resultssuggest that, in allergen-induced asthma, TSLP acts on NKT cells to enhance airway hyperreactivity by upregulating their IL-13 production, whereas eosinophilia and IgE production are not influenced.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Proliferation; Cytokines; Immunoglobulin E; Immunoglobulins; Killer Cells, Natural; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Receptors, Cytokine; Recombinant Proteins; RNA, Messenger; Th2 Cells; Thymic Stromal Lymphopoietin

Population studies have suggested that chronic and intense helminth infections, in contrast to acute and mild helminth infections, might suppress allergic airway inflammation.. We sought to address the question of how the chronicity and intensity of helminth infections affect allergic airway inflammation in a well-defined experimental model.. C57/Bl6 mice were infected with Schistosoma mansoni, followed by sensitization and challenge with ovalbumin (OVA), and different stages and intensities of infection were studied. To this end, mice were analyzed at 8, 12, or 16 weeks, representing the acute, intermediate, or chronic phases of infection, respectively.. Lung lavage eosinophilia, peribronchial inflammation, and OVA-induced airway hyperresponsiveness were increased during acute infection but significantly decreased when infection progressed into chronicity. Decreases in lung lavage eosinophilia were parasite density-dependent. Similar levels of OVA-specific IgE were found during all phases of infection, whereas both OVA-specific and parasite-specific T(H)2 cytokine levels were significantly reduced during chronic infection. Inhibition of airway inflammation could be transferred to OVA-sensitized recipient mice by B cells and CD4(+) T cells from spleens of chronically, but not acutely, infected mice. This suppression was IL-10-dependent.. During chronic, but not acute, helminth infections, suppressive mechanisms are induced that regulate immune reactions to inhaled allergens. These data confirm human epidemiologic observations in a well-controlled animal model.. Characterization of chronic helminth infection-induced regulatory mechanisms will help in the development of future therapeutics to treat or prevent allergic disease.

Topics: Adoptive Transfer; Animals; Bronchial Hyperreactivity; Chronic Disease; Cytokines; Eosinophilia; Female; Immunoglobulin E; Interleukin-10; Lung; Mice; Mice, Inbred C57BL; Ovalbumin; Schistosomiasis mansoni

CD4+ T cells, particularly Th2 cells, play a pivotal role in allergic airway inflammation. However, the requirements for interactions between CD4+ and CD8+ T cells in airway allergic inflammation have not been delineated. Sensitized and challenged OT-1 mice in which CD8+ T cells expressing the transgene for the OVA(257-264) peptide (SIINFEKL) failed to develop airway hyperresponsiveness (AHR), airway eosinophilia, Th2 cytokine elevation, or goblet cell metaplasia. OT-1 mice that received naive CD4+IL-4+ T cells but not CD4+IL-4- T cells before sensitization developed all of these responses to the same degree as wild-type mice. Moreover, recipients of CD4+IL-4+ T cells developed significant increases in the number of CD8+IL-13+ T cells in the lung, whereas sensitized OT-1 mice that received primed CD4+ T cells just before challenge failed to develop these responses. Sensitized CD8-deficient mice that received CD8+ T cells from OT-1 mice that received naive CD4+ T cells before sensitization increased AHR and eosinophil numbers in bronchoalveolar lavage fluid when challenged with allergen. In contrast, sensitized CD8-deficient mice receiving CD8+ T cells from OT-1 mice without CD4+ T cells developed reduced AHR and eosinophil numbers in bronchoalveolar lavage fluid when challenged. These data suggest that interactions between CD4+ and CD8+ T cells, in part through IL-4 during the sensitization phase, are essential to the development of CD8+IL-13+ T cell-dependent AHR and airway allergic inflammation.

Topics: Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; CD4 Antigens; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Egg Proteins; Inflammation; Interleukin-4; Mice; Mice, Transgenic; Ovalbumin; Peptide Fragments; Transgenes

Asthma is characterized by increases in airway resistance, pulmonary remodeling, and lung inflammation. The cytokine transforming growth factor (TGF)-beta has been shown to have a central role in asthma pathogenesis and in mouse models of allergic airway disease.. To determine the contribution of TGF-beta to airway hyperresponsiveness (AHR), we examined the time course, source, and isoform specificity of TGF-beta production in an in vivo mouse asthma model. To then elucidate the function of TGF-beta in AHR, inflammation, and pulmonary fibrosis, we examined the effects of blocking TGF-beta signaling with neutralizing antibody.. Mice were sensitized and challenged with ovalbumin (OVA) to establish allergic airway disease. TGF-beta activity was neutralized by intranasal administration of monoclonal antibody.. TGF-beta1 protein levels were increased in OVA-challenged lungs versus naive controls, and airway epithelial cells were shown to be a likely source of TGF-beta1. In addition, TGF-beta1 levels were elevated in OVA-exposed IL-5-null mice, which fail to recruit eosinophils into the airways. Neutralization of TGF-beta1 with specific antibody had no significant effect on airway inflammation and eosinophilia, although anti-TGF-beta1 antibody enhanced OVA-induced AHR and suppressed pulmonary fibrosis.. These data show that TGF-beta1 is the main TGF-beta isoform produced after OVA challenge, with a likely cellular source being the airway epithelium. The effects of blocking TGF-beta1 signaling had differential effects on AHR, fibrosis, and inflammation. While TGF-beta neutralization may be beneficial to abrogating airway remodeling, it may be detrimental to lung function by increasing AHR.

Topics: Animals; Antibodies, Monoclonal; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Female; Immunologic Factors; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Transforming Growth Factor beta1

We previously reported that ovalbumin (OVA) and IL-18 nasally administered act on memory type T helper (Th)1 cells to induce airway hyperresponsiveness (AHR) and inflammation, which is characterized by peribronchial infiltration with neutrophils and eosinophils. Here, we report this administration also induces lung fibrosis in an IL-13-dependent manner. Th1 cells secrete several cytokines, including IFN-gamma and bronchogenic cytokine IL-13, when stimulated with antigen (Ag) and IL-18. However, IL-13 blockade failed to attenuate AHR, although this treatment inhibited eosinophilic infiltration. To understand the mechanism by which Th1 cells induce AHR after Ag plus IL-18 challenge, we established "passive" and "active" Th1 mice by transferring OVA-specific Th1 cells into naïve BALB/c mice or by immunizing naïve BALB/c mice with OVA/complete Freund's adjuvant, respectively. Administration of Ag and IL-18 induced both types of Th1 mice to develop AHR, airway inflammation, and lung fibrosis. Furthermore, this treatment induced deposition of periostin, a novel component of lung fibrosis. Neutralization of IL-13 or IFN-gamma during Ag plus IL-18 challenges inhibited the combination of eosinophilic infiltration, lung fibrosis, and periostin deposition or the combination of neutrophilic infiltration and AHR, respectively. We also found that coadministration of OVA and LPS into Th1 mice induced AHR and airway inflammation via endogenous IL-18. Thus, IL-18 becomes a key target molecule for the development of a therapeutic regimen for the treatment of Th1-cell-induced bronchial asthma.

Topics: Adjuvants, Immunologic; Administration, Inhalation; Animals; Bronchial Hyperreactivity; Cell Adhesion Molecules; Female; Fibrosis; Freund's Adjuvant; Interferon-gamma; Interleukin-13; Interleukin-18; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Ovalbumin; Specific Pathogen-Free Organisms; Th1 Cells

Allergen sensitization and allergic airway disease are likely to come about through the inhalation of Ag with immunostimulatory molecules. However, environmental pollutants, including nitrogen dioxide (NO2), may promote adaptive immune responses to innocuous Ags that are not by themselves immunostimulatory. We tested in C57BL/6 mice whether exposure to NO2, followed by inhalation of the innocuous protein Ag, OVA, would result in allergen sensitization and the subsequent development of allergic airway disease. Following challenge with aerosolized OVA alone, mice previously exposed via inhalation to NO2 and OVA developed eosinophilic inflammation and mucus cell metaplasia in the lungs, as well as OVA-specific IgE and IgG1, and Th2-type cytokine responses. One hour of exposure to 10 parts per million NO2 increased bronchoalveolar lavage fluid levels of total protein, lactate dehydrogenase activity, and heat shock protein 70; promoted the activation of NF-kappaB by airway epithelial cells; and stimulated the subsequent allergic response to Ag challenge. Furthermore, features of allergic airway disease were not induced in allergen-challenged TLR2-/- and MyD88-/- mice exposed to NO2 and aerosolized OVA during sensitization. These findings offer a mechanism whereby allergen sensitization and asthma may result under conditions of high ambient or endogenous NO2 levels.

Topics: Administration, Inhalation; Aerosols; Allergens; Animals; Bronchial Hyperreactivity; Eosinophilia; Immunologic Factors; Lung; Metaplasia; Mice; Mice, Inbred C57BL; Mice, Knockout; Mucus; Myeloid Differentiation Factor 88; Nitrogen Dioxide; Ovalbumin; Respiratory Hypersensitivity; Toll-Like Receptor 2

The aim of this study was to investigate the influence of stress and neuropeptides on airway responses in ovalbumin (OVA)-sensitized rats.. Three experimental conditions were employed: neonatal capsaicin treatment, foot shock stress and OVA sensitization. For neuropeptide depletion, male Wistar rats were neonatally treated with capsaicin (50 mg/kg) or with control solution 2 days after birth. Ninety days later, they were injected with OVA and aluminum hydroxide (ED0) or no injection. Thereafter, rats of the stressed groups were individually placed in a shuttle box where they received 50 mild escapable foot shocks/day; the stressful stimuli were repeated until ED14, when the animals received OVA aerosol. Pulmonary mechanic function was measured before and after OVA challenge in anesthetized and mechanically ventilated rats.. Data on ultrasonic vocalizations and corticosterone showed high levels of anxiety in stressed animals. As expected, a significant increment in airway elastance and resistance after the OVA challenge was found in sensitized rats compared to non-sensitized ones. Capsaicin treatment decreased the values of elastance in sensitized and non-stressed rats; however, after the OVA challenge, elastance was increased in stressed animals. No differences were found in the levels of resistance among sensitized and non-stressed rats; however, a reduced increment in resistance was verified in capsaicin-treated, stressed animals.. Our results suggest that neurokinin depletion and stress may affect smooth muscle tonus around the airways during an anaphylactic reaction. These data suggest that stress and neuropeptides play a significant role in pulmonary function in OVA-sensitized rats.

Topics: Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Corticosterone; Male; Muscle, Smooth; Neuropeptides; Ovalbumin; Rats; Rats, Wistar; Stress, Psychological

We investigated the suppressive effects of 3-O-methylquercetin 5,7,3',4'- O-tetraacetate (QMTA), a more-potent phosphodiesterase (PDE)3/4 inhibitor than quercetin 3-O-methyl ether (3-MQ), which has been reported to have the potential for treating asthma, against ovalbumin (OVA)-induced airway hyperresponsiveness (AHR). The IC50 value of QMTA for PDE3 was significantly less than that for PDE4. According to the Lineweaver-Burk analysis, QMTA (1-10 microM) competitively inhibited PDE3 and PDE4 activities. The Ki values were 0.9+/-0.3 (n=5) and 3.9+/-0.5 (n=5) microM, respectively, which significantly differed from each other, suggesting that QMTA has higher affinity for PDE3 than for PDE4. QMTA (3-10 microM) concentration-dependently relaxed the baseline level, and significantly inhibited cumulative OVA (10-100 microg/mL)-induced contractions in isolated sensitized guinea pig trachealis suggesting that QMTA has bronchodilator and inhibiting effects on mast cell degranulation. After the secondary challenge, the AHR was measured in unrestrained OVA-sensitized mice, with nebulized methacholine (MCh, 6.25-50 mg/mL), by barometric plethysmography using a whole-body plethysmograph. In the present results, QMTA (3-10 micromol/kg, I. P.) dose-dependently attenuated the enhanced pause (Penh) value induced by MCh (25-50 mg/mL). QMTA (3-10 micromol/kg, I. P.) also significantly inhibited total inflammatory cells, macrophages, neutrophils, lymphocytes, and eosinophils in BALF after determination of Penh values. It also significantly suppressed the release of interleukin (IL)-2, IL-4, IL-5, IFN-gamma, and TNF-alpha, with the exception that 3 micromol/kg QMTA did not suppress the releases of IL-5. QMTA even at 1 micromol/kg significantly inhibited eosinophils, IL-2, and TNF-alpha. In conclusion, our results strongly suggest that QMTA has greater potential than 3-MQ for the treatment of asthma.

Topics: Animals; Bronchial Hyperreactivity; Bronchodilator Agents; Female; Inhibitory Concentration 50; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; Plant Extracts; Quercetin; Rhamnus

Over recent decades, there has been a significant global increase in the prevalence of asthma, an inflammatory disease of the respiratory system. While ultraviolet radiation (UV) has been used successfully in the treatment of inflammatory conditions such as psoriasis, studies of UV-induced regulation of allergic respiratory responses have been rare, and have not analysed in vivo measurements of airway hyperresponsiveness (AHR) or the antigen specificity of the UV-induced effects.. To investigate the regulatory properties of erythemal ultraviolet B (UVB) irradiation of the skin and the induction of allergen-induced airway immunity in a murine asthma model, and to examine the mechanisms involved.. BALB/c mice were exposed to a single erythemal dose of UV 3 days before intraperitonial sensitization (day 0) and boost (day 14) with the antigen, ovalbumin (OVA). Airway-associated, asthma-like responses to aerosolized OVA at day 21 were analysed including (a) AHR measured in vivo, (b) OVA-specific proliferative responses and cytokine production by cells from the lung-draining lymph nodes (LDLN), and (c) inflammatory cells and cytokines in the bronchoalveolar lavage fluid. To determine UVB-induced mechanisms of regulation, LDLN cells from UVB irradiated, OVA-sensitized mice were adoptively transferred into naïve BALB/c mice that were subsequently sensitized and challenged with OVA, or a non-specific antigen.. UVB irradiation of skin significantly suppressed AHR to methacholine and OVA-specific responses in the LDLN and in the lung compartment. Reduced OVA-specific responses by LDLN cells from both UVB irradiated mice and mice that received 5 x 10(6) LDLN cells from UVB irradiated, but not from non-irradiated, OVA-sensitized mice suggested that UVB-induced regulatory cells are responsible for many of the asthma-reducing effects of dorsal UVB exposure.. UVB irradiation of skin suppresses AHR and cellular responses of the airways to respiratory allergens. Further, this study implicates UVB or its downstream mediators as a potential approach to reducing the severity of asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Immunity, Cellular; Immunization; Immunoglobulin E; Lymph Nodes; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Phenotype; Rats; Rats, Sprague-Dawley; Ultraviolet Rays; Ultraviolet Therapy

The effect of ageing on several pathologic features of allergic asthma (pulmonary inflammation, eosinophilia, mucus hypersecretion), and their relationship with airway hyperresponsiveness (AHR) is not well characterized.. To evaluate lung inflammation, mucus metaplasia and AHR in relationship with age in murine models of allergic asthma comparing young and older mice.. Young (6 weeks) and older (6, 12, 18 months) BALB/c mice were sensitized and challenged with ovalbumin (OVA). AHR and bronchoalveolar fluid (BALF), total inflammatory cell count and differential were measured. To evaluate mucus metaplasia, quantitative PCR for the major airway mucin-associated gene, MUC-5AC, from lung tissue was measured, and lung tissue sections stained with periodic acid-Schiff (PAS) for goblet-cell enumeration. Lung tissue cytokine gene expression was determined by quantitative PCR, and systemic cytokine protein levels by ELISA from spleen-cell cultures. Antigen-specific serum IgE was determined by ELISA.. AHR developed in both aged and young OVA-sensitized/challenged mice (OVA mice), and was more significantly increased in young OVA mice than in aged OVA mice. However, BALF eosinophil numbers were significantly higher, and lung histology showed greater inflammation in aged OVA mice than in young OVA mice. MUC-5AC expression and numbers of PAS+ staining bronchial epithelial cells were significantly increased in the aged OVA mice. All aged OVA mice had increased IL-5 and IFN-gamma mRNA expression in the lung and IL-5 and IFN-gamma protein levels from spleen cell cultures compared with young OVA mice. OVA-IgE was elevated to a greater extent in aged OVA mice.. Although pulmonary inflammation and mucus metaplasia after antigen sensitization/challenge occurred to a greater degree in older mice, the increase in AHR was significantly less compared with younger OVA mice. Antigen treatment produced a unique cytokine profile in older mice (elevated IFN-gamma and IL-5) compared with young mice (elevated IL-4 and IL-13). Thus, the airway response to inflammation is lessened in ageing animals, and may represent age-associated events leading to different phenotypes in response to antigen provocation.

Topics: Aging; Animals; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Enzyme-Linked Immunosorbent Assay; Eosinophilia; Female; Gene Expression; Interferon-gamma; Interleukin-13; Interleukin-4; Interleukin-5; Metaplasia; Mice; Mice, Inbred BALB C; Mucin 5AC; Mucins; Mucus; Ovalbumin; Pneumonia; Polymerase Chain Reaction; T-Lymphocytes

Kefir is a microbial symbiont mixture that produces jelly-like grains. As a widely used neutraceutical, however, the therapeutic applicability of kefir is not certain. In order to investigate the pharmacological effects of kefir, we used a mouse asthma model, in which airway inflammation and airway remodeling was produced by ovalbumin sensitization and challenge. BALB/c mice sensitized and challenged to ovalbumin, were treated with kefir (50mg/kg administered by intra-gastric mode) 1h before the ovalbumin challenge. Kefir significantly suppressed ovalbumin-induced airway hyper-responsiveness (AHR) to inhaled methacholine. Intra-gastric administration of kefir significantly inhibited the increase in the total inflammatory cell count induced by ovalbumin, and the eosinophil count in bronchoalveolar lavage fluid (BALF). Type 2 helper T cell (Th2) cytokines, such as interleukin-4 and interleukin-13, and total immunoglobulin E (Ig E) levels, were also reduced to normal levels in bronchoalveolar lavage fluid. Histological studies demonstrate that kefir substantially inhibited ovalbumin-induced eosinophilia in lung tissue and mucus hyper-secretion by goblet cells in the airway. Kefir displayed anti-inflammatory and anti-allergic effects in a mouse asthma model and may possess new therapeutic potential for the treatment of allergic bronchial asthma.

Topics: Administration, Oral; Animals; Asthma; Bronchial Hyperreactivity; Cultured Milk Products; Cytokines; Disease Models, Animal; Eosinophilia; Immunoglobulin E; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin

Brain stem noradrenergic cell groups mediating autonomic responses to stress project to airway-related vagal preganglionic neurons (AVPNs). In ferrets, their activation produces withdrawal of cholinergic outflow to the airways via release of norepinephrine and activation of alpha(2A)-adrenergic receptors (alpha(2A)-AR) expressed by AVPNs. In these studies, we examined the effects of allergen exposure of the airway (AE) with ovalbumin on noradrenergic transmission regulating the activity of AVPNs and, consequently, airway smooth muscle tone. Experiments were performed in vehicle control (Con) and AE ferrets. Microperfusion of an alpha(2A)-AR agonist (guanabenz) in close proximity to AVPNs elicited more pronounced effects in Con than AE ferrets, including a decrease in unit activity and reflexly evoked responses of putative AVPN neurons with a corresponding decrease in cholinergic outflow to the airways. Although no differences were found in the extent of noradrenergic innervation of the AVPNs, RT-PCR and Western blot studies demonstrated that AE and repeated exposure to antigen significantly reduced expression of alpha(2A)-ARs at message and protein levels. These findings indicate that, in an animal model of allergic asthma, sensitization and repeated challenges with a specific allergen diminish central inhibitory noradrenergic modulation of AVPNs, possibly via downregulation of alpha(2A)-AR expression by these neurons.

Topics: Action Potentials; Adrenergic alpha-Agonists; Adrenergic Fibers; Allergens; Animals; Asthma; Autonomic Fibers, Preganglionic; Brain Stem; Bronchial Hyperreactivity; Bronchoconstriction; Disease Models, Animal; Down-Regulation; Ferrets; Guanabenz; Male; Neural Inhibition; Norepinephrine; Ovalbumin; Receptors, Adrenergic, alpha-2; Research Design; Respiratory System; RNA, Messenger; Time Factors; Vagus Nerve

iNKT cells are required for the induction of airway hyperreactivity (AHR), a cardinal feature of asthma, but how iNKT cells traffic to the lungs to induce AHR has not been previously studied. Using several models of asthma, we demonstrated that iNKT cells required the chemokine receptor CCR4 for pulmonary localization and for the induction of AHR. In both allergen-induced and glycolipid-induced models of AHR, wild-type but not CCR4-/- mice developed AHR. Furthermore, adoptive transfer of wild-type but not CCR4-/- iNKT cells reconstituted AHR in iNKT cell-deficient mice. Moreover, we specifically tracked CCR4-/- vs wild-type iNKT cells in CCR4-/-:wild-type mixed BM chimeric mice in the resting state, and when AHR was induced by protein allergen or glycolipid. Using this unique model, we showed that both iNKT cells and conventional T cells required CCR4 for competitive localization into the bronchoalveolar lavage/airways compartment. These results establish for the first time that the pulmonary localization of iNKT cells critical for the induction of AHR requires CCR4 expression by iNKT cells.

Topics: Animals; Bronchial Hyperreactivity; Cell Movement; Chemokine CCL17; Chemokines, CC; Cytokines; Glycolipids; Mice; Mice, Knockout; Ovalbumin; Receptors, CCR4; Receptors, Chemokine; T-Lymphocytes, Regulatory

A subset of food-allergic patients does not only respond clinically with symptoms in the gastro-intestinal tract but also with asthmatic reactions.. The aim of this study was to analyse whether CD4+ T cells from mice with intestinal immediate-hypersensitivity reactions to food allergen are involved in the development of experimental asthma.. BALB/c mice were intraperitoneally sensitized to ovalbumin (OVA), followed by repeated intra-gastric (i.g.) OVA challenges. Control animals were either sham-sensitized or sham-challenged with phosphate-buffered saline (PBS). Duodenum, jejunum, ileum and colon were histologically examined. CD4+ T cells from mesenteric lymph nodes were transferred from various donor groups into recipient mice that received either OVA or PBS aerosol challenges. Recipients were analysed by measurements of lung function using head-out body-plethysmography and examination of broncho-alveolar lavage and lung histology.. The highest levels of OVA-specific IgE antibody levels were detected in OVA-sensitized and OVA-challenged mice. Throughout the lower intestinal tract, a marked infiltration with eosinophils was observed, and goblet cell numbers as well as goblet cell area were significantly increased. The villus/crypt ratio was decreased compared with controls. The transfer of CD4+ T cells from mesenteric lymph nodes of OVA-sensitized and OVA-challenged mice triggered airway hyperreactivity and eosinophilic airway inflammation in recipients aerosol challenged with OVA, but not with PBS.. We conclude that CD4+ T cells from mesenteric lymph nodes of mice with allergen-induced immediate-type hypersensitivity reactions in the gut are able to transfer the phenotype of experimental asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Female; Food Hypersensitivity; Hypersensitivity, Immediate; Immunoglobulin E; Intestines; Lymph Nodes; Mice; Mice, Inbred BALB C; Ovalbumin

Tobacco smoke (TS) exposure can induce airway hyperresponsiveness, especially in asthma. A feature of asthma is eosinophilia. We hypothesized that tobacco smoke exposure enhances eosinophil responsiveness in sensitized guinea pigs. Tobacco smoke-exposed, ovalbumin (OA)-sensitized guinea pigs were treated with TRFK-5 (1.0 mg/kg, intraperitoneal), an anti-interleukin (IL)-5 agent, or its vehicle. Guinea pigs were challenged with aerosols of OA, capsaicin, histamine, and methacholine. TRFK-5 attenuated airway responsiveness to OA but not to capsaicin, histamine, or methacholine. Bronchial alveolar lavage fluid analysis confirmed TRFK-5 attenuated airway eosinophilia in OA-treated guinea pigs. Therefore, airway responsiveness to OA is enhanced by eosinophils or IL-5 itself.

Topics: Animals; Antibodies, Monoclonal; Bronchial Hyperreactivity; Capsaicin; Cell Movement; Disease Models, Animal; Eosinophils; Guinea Pigs; Histamine; Interleukin-5; Male; Methacholine Chloride; Ovalbumin; Tobacco Smoke Pollution

Asthma is a common inflammatory disease of the airways. Current therapies alleviate symptoms but do not treat the disease. We aim to develop effective immunomodulatory therapies (IMTs) for asthma that target the underlying causes of disease based on Streptococcus pneumoniae (Spn). The effect of Spn IMT on the development of asthma [allergic airways disease (AAD)] was determined in mice. Killed Spn was administered before, during or after ovalbumin sensitization, and the subsequent development of AAD was assessed. IMT attenuated T cell cytokine production, goblet cell hyperplasia, airways hyperresponsiveness (AHR), and eosinophil numbers in the blood, bronchoalveolar lavage fluid and peribronchial tissue. This indicates the potential of Spn as an IMT for asthma.

Topics: Allergens; Animals; Antigens, Bacterial; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Hypersensitivity; Streptococcus pneumoniae; T-Lymphocytes

Tobacco smoke (TS) exposure exacerbates asthma and may induce airway hyperresponsiveness in asymptomatic individuals. We hypothesized that TS exposure is an adjuvant to airway responsiveness. Ovalbumin (OA) sensitized guinea pigs were TS or air exposed. At 30 exposure days OA airway responsiveness was demonstrable in OA-treated animals exposed to either TS or air. After 130 exposure days only TS-exposed guinea pigs demonstrated OA airway responsiveness. Capsaicin airway responsiveness developed in non-sensitized and OA-sensitized guinea pigs exposed to TS. Therefore TS-exposure acts as an adjuvant to antigenic and neurogenic airway responsiveness. Combined antigen and adjuvant avoidance may attenuate or reverse airway responsiveness.

Topics: Adjuvants, Immunologic; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Capsaicin; Guinea Pigs; Ovalbumin; Random Allocation; Tobacco Smoke Pollution

Nitric oxide (NO) levels are elevated in the exhaled breath of asthmatic patients and NO is considered as a biomarker of airway inflammation. However, the functions of NO in the airways are not completely understood. L-arginine, as the substrate of NO synthases, is the precursor of NO which stimulates guanylate cyclase and leads to the formation of cyclic GMP (cGMP). Sildenafil, a phosphodiestérase-5 (PDE-5) inhibitor, prevents the degradation of cGMP. In this study the effects of L-arginine and sildenafil treatment, alone or in combination, were evaluated in ovalbumin-sensitized BP2 mice. These effects concerning the airway responsiveness to inhaled methacholine (MCh) were evaluated by whole-body plethysmography (WBP), the inflammatory response evaluated by bronchoalveolar lavage fluid (BALF) analyses and lung tissue biopsies (eosinophilic inflammation associated with lung remodelling), and NO metabolite measurements (by Griess reaction) in BALF. Ovalbumin sensitization induced: (a) an inflammatory reaction with eosinophil and neutrophil influx in BALF and lung; and (b) an increased bronchial responsiveness to MCh. L-arginine treatment [50 mg/kg intraperitoneally (i.p.), for 7 days] increased the relative amount of eosinophils and neutrophils in BALF, had a tendency to increase the airway responsiveness to inhaled MCh and increased the NO metabolite level in BAL. Sildenafil treatment (20 mg/kg i.p. for 7 days) did not affect the airway responsiveness to MCh and had a lower effect compared with L-arginine on inflammatory reactions. The combination of the two treatments resulted in a dramatic enhancement of the airway responsiveness to inhaled MCh. The relative amount of eosinophils was increased and lung histology showed obvious worsened tissular lesions such as epithelial shedding and hypertrophy, hyperplasia of smooth muscle cells, and fibrosis. These findings are consistent with the notion that NO production plays a role in the development of airway inflammation and hyperresponsiveness of sensitized mice and highlighted the potential risk of the L-arginine dietary complement or PDE5 treatment in asthmatic patients.

Topics: Animals; Arginine; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cyclic GMP; Eosinophilia; Eosinophils; Lung; Male; Methacholine Chloride; Mice; Nitric Oxide; Ovalbumin; Phosphodiesterase 5 Inhibitors; Phosphodiesterase Inhibitors; Piperazines; Plethysmography, Whole Body; Purines; Sildenafil Citrate; Sulfones

Reactive oxygen species (ROS) play an important role in the pathogenesis of airway inflammation and hyperresponsiveness. Recent studies have demonstrated that antioxidants are able to reduce airway inflammation and hyperreactivity in animal models of allergic airway disease. A newly developed antioxidant, small molecular weight thiol compound, N-acetylcysteine amide (AD4) has been shown to increase cellular levels of glutathione and to attenuate oxidative stress related disorders such as Alzheimer's disease, Parkinson's disease, and multiple sclerosis. However, the effects of AD4 on allergic airway disease such as asthma are unknown. We used ovalbumin (OVA)-inhaled mice to evaluate the role of AD4 in allergic airway disease. In this study with OVA-inhaled mice, the increased ROS generation, the increased levels of Th2 cytokines and VEGF, the increased vascular permeability, the increased mucus production, and the increased airway resistance in the lungs were significantly reduced by the administration of AD4. We also found that the administration of AD4 decreased the increases of the NF-kappaB and hypoxia-inducible factor-1alpha (HIF-1alpha) levels in nuclear protein extracts of lung tissues after OVA inhalation. These results suggest that AD4 attenuates airway inflammation and hyperresponsiveness by regulating activation of NF-kappaB and HIF-1alpha as well as reducing ROS generation in allergic airway disease.

Topics: Acetylcysteine; Animals; Asthma; Bronchial Hyperreactivity; Hypoxia-Inducible Factor 1, alpha Subunit; Mice; NF-kappa B; Ovalbumin; Reactive Oxygen Species; Vascular Endothelial Growth Factor A

Asthma is a complex phenotype that involves multiple mechanisms, including adaptive and innate immunity as well as physiological and mechanical changes in the airways. A cardinal feature of asthma is airway hyperreactivity (AHR), a multifaceted reaction that can only be assessed in vivo. Mouse models of asthma replicate many of the features of human asthma, including AHR, which can be assessed using standard protocols. Examination of AHR in mice has provided important information about human asthma, primarily because the immunology of allergy is easily studied in mice, especially with the availability of reagents including genetically modified mice. In this unit we discuss the induction and measurement of AHR and the two most common methodologies: noninvasive measurement using a whole-body plethysmograph (WBP) and invasive measurement of lung resistance and dynamic compliance.

Topics: Airway Resistance; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Cell Count; Cell Proliferation; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Humans; Immunity, Cellular; Immunization; Lung Compliance; Methacholine Chloride; Mice; Ovalbumin; Plethysmography, Whole Body

In addition to being an air pollutant, NO2 is a potent inflammatory oxidant generated endogenously by myeloperoxidase and eosinophil peroxidase. In these studies, we sought to determine the effects of NO2 exposure on mice with ongoing allergic airway disease pathology. Mice were sensitized and challenged with the antigen ovalbumin (OVA) to generate airway inflammation and subsequently exposed to 5 or 25 ppm NO2 for 3 days or 5 days followed by a 20-day recovery period. Whereas 5 ppm NO2 elicited no pathological changes, inhalation of 25 ppm NO2 alone induced acute lung injury, which peaked after 3 days and was characterized by increases in protein, LDH, and neutrophils recovered by BAL, as well as lesions within terminal bronchioles. Importantly, 25 ppm NO2 was also sufficient to cause AHR in mice, a cardinal feature of asthma. The inflammatory changes were ameliorated after 5 days of inhalation and completely resolved after 20 days of recovery after the 5-day inhalation. In contrast, in mice immunized and challenged with OVA, inhalation of 25 ppm NO2 caused a marked augmentation of eosinophilic inflammation and terminal bronchiolar lesions, which extended significantly into the alveoli. Moreover, 20 days postcessation of the 5-day 25 ppm NO2 inhalation regimen, eosinophilic and neutrophilic inflammation, pulmonary lesions, and AHR were still present in mice immunized and challenged with OVA. Collectively, these observations suggest an important role for NO2 in airway pathologies associated with asthma, both in modulation of degree and duration of inflammatory response, as well as in induction of AHR.

Topics: Animals; Bronchi; Bronchial Hyperreactivity; Dose-Response Relationship, Drug; Hypersensitivity; Mice; Mice, Inbred C57BL; Nitrogen Dioxide; Ovalbumin; Oxidants, Photochemical; Pneumonia

Antibody-antigen interactions in the airway initiate inflammation in acute asthma exacerbations. This inflammatory response is characterized by the recruitment of granulocytes into the airways. In murine models of asthma, granulocyte recruitment into the lung contributes to the development of airway hyperresponsiveness (AHR), mucus production, and airway remodeling. Leukotriene B4 is a mediator released following antigen challenge that has chemotactic activity for granulocytes, mediated through its receptor, BLT1. We investigated the role of BLT1 in granulocyte recruitment following antigen challenge. Wild-type mice and BLT1-/- mice were sensitized and challenged with ovalbumin (OVA) to induce acute allergic airway inflammation. In addition, to explore the relevance to antibody-antigen interactions, we injected OVA bound to anti-OVA IgG1 or anti-OVA IgE intratracheally into naïve wild-type and BLT1-/- mice. Cell composition of the lungs, cytokine levels, histology, and AHR were determined. After sensitization and challenge with ovalbumin, there was significantly reduced neutrophil and eosinophil recruitment into the airways of BLT1-/- mice compared with wild-type animals after one or two daily antigen challenges, but this difference was not seen after three or four daily antigen challenges. Mucus production and AHR were not affected. Intratracheal injection of OVA bound to IgG1 or IgE induced neutrophil recruitment into the airways in wild-type mice but not in the BLT1-/- mice. We conclude that BLT1 mediates early recruitment of granulocytes into the airway in response to antigen-antibody interactions in a murine model of acute asthma.

Topics: Animals; Antigen-Antibody Reactions; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Chemotaxis, Leukocyte; Disease Models, Animal; Granulocytes; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Lung; Male; Mice; Mice, Knockout; Mucus; Neutrophils; Ovalbumin; Receptors, Leukotriene B4; Receptors, Purinergic P2

Soluble guanylyl cyclase (sGC) is an enzyme highly expressed in the lung that generates cGMP contributing to airway smooth muscle relaxation. To determine whether the bronchoconstriction observed in asthma is accompanied by changes in sGC expression, we used a well-established murine model of allergic asthma. Histological and biochemical analyses confirmed the presence of inflammation in the lungs of mice sensitized and challenged with ovalbumin (OVA). Moreover, mice sensitized and challenged with OVA exhibited airway hyperreactivity to methacholine inhalation. Steady-state mRNA levels for all sGC subunits (alpha1, alpha2, and beta1) were reduced in the lungs of mice with allergic asthma by 60-80%, as estimated by real-time PCR. These changes in mRNA were paralleled by changes at the protein level: alpha1, alpha2, and beta1 expression was reduced by 50-80% as determined by Western blotting. Reduced alpha1 and beta1 expression in bronchial smooth muscle cells was demonstrated by immunohistochemistry. To study if sGC inhibition mimics the airway hyperreactivity seen in asthma, we treated naïve mice with a selective sGC inhibitor. Indeed, in mice receiving ODQ the methacholine dose response was shifted to the left. We conclude that sGC expression is reduced in experimental asthma contributing to the observed airway hyperreactivity.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Enzyme Inhibitors; Guanylate Cyclase; Homeostasis; Hypersensitivity; Isoenzymes; Methacholine Chloride; Mice; Ovalbumin; Oxadiazoles; Quinoxalines; Receptors, Cytoplasmic and Nuclear; RNA, Messenger; Soluble Guanylyl Cyclase

There is conflicting information about the development and resolution of airway inflammation and airway hyperresponsiveness (AHR) after repeated airway exposure to allergen in sensitized mice.. Sensitized BALB/c and C57BL/6 mice were exposed to repeated allergen challenge on 3, 7, or 11 occasions. Airway function in response to inhaled methacholine was monitored; bronchoalveolar lavage fluid inflammatory cells were counted; and goblet cell metaplasia, peribronchial fibrosis, and smooth muscle hypertrophy were quantitated on tissue sections. Bone marrow-derived dendritic cells were generated after differentiation of bone marrow cells in the presence of growth factors.. Sensitization to ovalbumin (OVA) in alum, followed by three airway exposures to OVA, induced lung eosinophilia, goblet cell metaplasia, mild peribronchial fibrosis, and peribronchial smooth muscle hypertrophy; increased levels of interleukin (IL)-4, IL-5, IL-13, granulocyte-macrophage colony-stimulating factor, transforming growth factor-beta(1), eotaxin-1, RANTES (regulated on activation, normal T-cell expressed and secreted), and OVA-specific IgG1 and IgE; and resulted in AHR. After seven airway challenges, development of AHR was markedly decreased as was the production of IL-4, IL-5, and IL-13. Levels of IL-10 in both strains and the level of IL-12 in BALB/c mice increased. After 11 challenges, airway eosinophilia and peribronchial fibrosis further declined and the cytokine and chemokine profiles continued to change. At this time point, the number of myeloid dendritic cells and expression of CD80 and CD86 in lungs were decreased compared with three challenges. After 11 challenges, intratracheal instillation of bone marrow-derived dendritic cells restored AHR and airway eosinophilia.. These data suggest that repeated allergen exposure leads to progressive decreases in AHR and allergic inflammation, through decreases in myeloid dendritic cell numbers.

Topics: Allergens; Animals; Bone Marrow Cells; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Dendritic Cells; Disease Models, Animal; Female; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin

Airway responses induced by intranasal administration of mite allergen without adjuvant were studied in NC/Nga mice. A crude extract of Dermatophagoides farinae (Df) was administered for 5 consecutive days and a single intranasal challenge booster dose was given 1 week after the last sensitization. 24 h after the single challenge, the airway hyperresponsiveness (AHR) was measured and the bronchoalveolar lavage fluid (BALF) was analyzed for numbers of eosinophils and neutrophils, and both cytokine and chemokine levels. There were marked increases in number of eosinophils in the BALF, AHR, Th2 cytokines (IL-5 and IL-13), and chemokine (eotaxin-1 and eotaxin-2) levels in the BALF following Df exposure. C57BL/6N, A/J, BALB/c, and CBA/JN mouse strains were also exposed to Df crude extract, but all of the measured responses were strongest in NC/Nga mice. Furthermore, Df-exposed NC/Nga mice showed the goblet cell hyperplasia, pulmonary eosinophilic inflammation, and increases in both total serum IgE and Df-specific IgG1. After intranasal exposure of NC/Nga mice to crude extract of Dermatophagoides pteronyssinus, the BALF eosinophilia and AHR were similar to responses induced by Df. None of the study parameters were increased in response to intranasal exposure to ovalbumin. These data demonstrated that NC/Nga mice developed allergic asthma-like responses after intranasal exposure to mite allergens.

Topics: Acetylcholine; Administration, Intranasal; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Cytokines; Immunoglobulin E; Immunoglobulin G; Lung; Mice; Mice, Inbred A; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Inbred Strains; Mites; Nasal Mucosa; Ovalbumin

Recent epidemiological studies have shown that growing up on a traditional farm provides protection from the development of allergic disorders such as hay fever and allergic asthma. We present experimental evidence that substances providing protection from the development of allergic diseases can be extracted from dust collected in stables of animal farms.. Stable dust was collected from 30 randomly selected farms located in rural regions of the Alps (Austria, Germany and Switzerland). The dust was homogenised with glass beads and extracted with physiological sodium chloride solution. This extract was used to modulate immune response in a well established mouse model of allergic asthma.. Treatment of mice by inhalation of stable dust extract during sensitisation to ovalbumin inhibited the development of airway hyperresponsiveness and airway eosinophilia upon challenge, as well as the production of interleukin 5 by splenocytes and of antigen specific IgG(1) and IgE. Dust extract also suppressed the generation of human dendritic cells in vitro. The biological activity of the dust extract was not exclusively mediated by lipopolysaccharide.. Stable dust from animal farms contains strong immune modulating substances. These substances can interfere with the development of both cellular and humoral immunity against allergens, thus suppressing allergen sensitisation, airway inflammation, and airway hyperresponsiveness in a murine model of allergic asthma.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; Cytokines; Dendritic Cells; Dust; Female; Flow Cytometry; Immunization; Immunoglobulins; Inhalation Exposure; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Hypersensitivity; Spleen

We evaluated the role of Syk, using an inhibitor, on allergen-induced airway hyperresponsiveness (AHR) and airway inflammation in a system shown to be B cell- and mast cell-independent. Sensitization of BALB/c mice with ovalbumin (OVA) and alum after three consecutive OVA challenges resulted in AHR to inhaled methacholine and airway inflammation. The Syk inhibitor R406 (30 mg/kg, administered orally, twice daily) prevented the development of AHR, increases in eosinophils and lymphocytes and IL-13 levels in bronchoalveolar lavage (BAL) fluid, and goblet cell metaplasia when administered after sensitization and before challenge with OVA. Levels of IL-4, IL-5, and IFN-gamma in BAL fluid and allergen-specific antibody levels in serum were not affected by treatment. Because many of these responses may be influenced by dendritic cell function, we investigated the effect of R406 on bone marrow-derived dendritic cell (BMDC) function. Co-culture of BMDC with immune complexes of OVA and IgG anti-OVA together with OVA-sensitized spleen mononuclear cells resulted in increases in IL-13 production. IL-13 production was inhibited if the BMDCs were pretreated with the Syk inhibitor. Intratracheal transfer of immune complex-pulsed BMDCs (but not nonpulsed BMDCs) to naive mice before airway allergen challenge induced the development of AHR and increases in BAL eosinophils and lymphocytes. All of these responses were inhibited if the transferred BMDCs were pretreated with R406. These results demonstrate that Syk inhibition prevents allergen-induced AHR and airway inflammation after systemic sensitization and challenge, at least in part through alteration of DC function.

Topics: Allergens; Animals; B-Lymphocytes; Bone Marrow Cells; Bronchial Hyperreactivity; Cells, Cultured; Coculture Techniques; Cytokines; Dendritic Cells; Enzyme Activation; Female; Goblet Cells; Inflammation; Interleukin-13; Intracellular Signaling Peptides and Proteins; Mast Cells; Metaplasia; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Oxazines; Protein-Tyrosine Kinases; Pyridines; Respiratory System; Syk Kinase

It has been shown that the presence of certain helminth infections in humans, including schistosomes, may reduce the propensity to develop allergies in infected populations. Using a mouse model of schistosome worm vs worm + egg infection, our objective was to dissect the mechanisms underlying the inverse relationship between helminth infections and allergies. We have demonstrated that conventional Schistosoma mansoni egg-laying male and female worm infection of mice exacerbates airway hyperresponsiveness. In contrast, mice infected with only schistosome male worms, precluding egg production, were protected from OVA-induced airway hyperresponsiveness. Worm-infected mice developed a novel modified type 2 cytokine response in the lungs, with elevated allergen-specific IL-4 and IL-13 but reduced IL-5, and increased IL-10. Although schistosome worm-only infection is a laboratory model, these data illustrate the complexity of schistosome modulation of host immunity by the worm vs egg stages of this helminth, with the potential of infections to aggravate or suppress allergic pulmonary inflammation. Thus, infection of mice with a human parasitic worm can result in reduced airway inflammation in response to a model allergen.

Topics: Allergens; Animals; B-Lymphocytes; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Enzyme-Linked Immunosorbent Assay; Female; Flow Cytometry; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Ovum; Schistosoma mansoni; Schistosomiasis mansoni; Sex Factors

Eosinophils cluster around airway nerves in patients with fatal asthma and in antigen-challenged animals. Activated eosinophils release major basic protein, which blocks inhibitory M2 muscarinic receptors (M2Rs) on nerves, increasing acetylcholine release and potentiating vagally mediated bronchoconstriction. We tested whether GW701897B, an antagonist of CCR3 (the receptor for eotaxin as well as a group of eosinophil active chemokines), affected vagal reactivity and M2R function in ovalbumin-challenged guinea pigs. Sensitized animals were treated with the CCR3 antagonist before inhaling ovalbumin. Antigen-challenged animals were hyperresponsive to vagal stimulation, but those that received the CCR3 antagonist were not. M2R function was lost in antigen-challenged animals, but not in those that received the CCR3 antagonist. Although the CCR3 antagonist did not decrease the number of eosinophils in lung tissues as assessed histologically, CCR3 antagonist prevented antigen-induced clustering of eosinophils along the nerves. Immunostaining revealed eotaxin in airway nerves and in cultured airway parasympathetic neurons from both guinea pigs and humans. Both IL-4 and IL-13 increased expression of eotaxin in cultured airway parasympathetic neurons as well as in human neuroblastoma cells. Thus, signaling via CCR3 mediates eosinophil recruitment to airway nerves and may be a prerequisite to blockade of inhibitory M2Rs by eosinophil major basic protein.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokines, CC; Disease Models, Animal; Female; Guinea Pigs; Neurons; Ovalbumin; Parasympathetic Nervous System; Receptor, Muscarinic M2; Receptors, CCR3; Receptors, Chemokine; Receptors, Interleukin-4

Epidemiologic data suggest a link between nursing by asthmatic mothers and increased risk of allergy in babies. We sought to experimentally test the potential contribution of breast milk mediator(s) in a mouse model of maternal transmission of asthma risk by evaluating the effect of adoptive nursing on asthma susceptibility in the offspring. We measured airway hyperresponsiveness (AHR) and allergic airway inflammation (AI) after an intentionally suboptimal OVA Ag sensitization, tested the allergen independence of the maternal effect by using a second allergen, casein, for sensitization of the baby mice, and tested the potential role of cytokines by measuring their levels in breast milk. Offspring of asthmatic, but not normal, mothers showed AHR and AI, indicating a maternal transfer of asthma risk. After adoptive nursing, both groups (litters born to asthmatic mothers and nursed by normal mothers, and normal babies nursed by asthmatic mothers) showed AHR (enhanced pause after methacholine aerosol, 50 mg/ml, 3.7 +/- 0.7, 4.2 +/- 0.5, respectively, vs 1.1 +/- 0.1 normal controls, n = 25, p < 0.01) and AI, seen as eosinophilia on histology and bronchoalveolar lavage (40.7 +/- 4.5%, 28.7 +/- 3.7%, vs 1.0 +/- 0.5% normals, n = 25, p < 0.01) after OVA sensitization. Similar results using casein allergen were observed. Multiplex assays for cytokines (IFN-gamma, IL-2, IL-4, IL-5, TNF-alpha, and IL-13) in breast milk were negative. Breast milk is sufficient, but not necessary, to mediate allergen-independent maternal transmission of asthma risk to offspring.

Topics: Allergens; Animals; Animals, Newborn; Asthma; Base Sequence; Breast Feeding; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Female; Gene Expression; Humans; Immunity, Maternally-Acquired; Infant, Newborn; Mice; Mice, Inbred BALB C; Milk; Milk, Human; Ovalbumin; Pregnancy; Risk Factors; RNA, Messenger

We previously demonstrated that treatment of acute asthmatic rats with gene therapy using plasmid-encoding Galectin-3 (Gal-3) resulted in an improvement of cellular and functional respiratory parameters. The next question that we wanted to clarify was if in a chronic situation where the treated animal continues to inhale the Ag, does this procedure prevent the chronicity and the remodeling? Chronic inflammation was induced by intranasal administration of OVA over a period of 12 wk. In the treated group, the Gal-3 gene was introduced by intranasal instillation in 50 mul of plasmid-encoding Gal-3. Noninvasive airway responsiveness to methacholine was tested at different times. Cells were obtained by bronchoalveolar lavage and used for RNA extraction and cytometric studies. Eosinophils were counted in blood and bronchoalveolar lavage fluid. Real-time PCR was used to measure Gal-3 and cytokine mRNA expression in lung. Lungs were paraffined and histologic analyses were performed (H&E, periodic acid-Schiff, and Masson Trichrome stain). Our results showed that 12 wk after the first intranasal Ag instillation in chronically asthmatic mice, treatment with the Gal-3 gene led to an improvement in the eosinophil count and the normalization of hyperresponsiveness to methacholine. Concomitantly, this treatment resulted in an improvement in mucus secretion and subepithelial fibrosis in the chronically asthmatic mice, with a quantitatively measured reduction in lung collagen, a prominent feature of airway remodeling. Plasmid-encoding Gal-3 acts as a novel treatment for chronic asthma in mice producing nearly complete blockade of Ag responses with respect to eosinophil airway accumulation, airway hyperresponsiveness, and remodeling.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chronic Disease; Collagen; Cytokines; Disease Models, Animal; Eosinophilia; Galectin 3; Genetic Therapy; Humans; Immunoglobulin E; Immunoglobulin G; Leukocyte Count; Lung; Male; Mice; Mice, Inbred A; Ovalbumin

alpha(4)beta(1) and alpha(4)beta(7) integrins are preferentially expressed on eosinophils and mononuclear leukocytes and play critical roles in their recruitment to inflammatory sites. We investigated the effects of TR14035, a small molecule, alpha(4)beta(1)/alpha(4)beta(7) dual antagonist, in a rat model of allergic asthma. Actively sensitized rats were challenged with aerosol antigen or saline on day 21, and the responses evaluated 24 and 48-h later. TR14035 (3 mg kg(-1), p.o.) was given 1-h before and 4-h after antigen or saline challenge. Airway hyper-responsiveness to intravenous 5-hydroxytryptamine was suppressed in TR14035-treated rats. Eosinophil, mononuclear cell and neutrophil counts, and eosinophil peroxidase and protein content in the bronchoalveolar lavage fluid (BALF) were decreased in TR14035-treated rats. Histological study showed a marked reduction of lung inflammatory lesions by TR14035. At 24-h postchallenge, antigen-induced lung interleukin (IL)-5 mRNA upregulation was suppressed in TR14035-treated rats. By contrast, IL-4 levels in BALF were not significantly affected by TR14035 treatment. IL-4 selectively upregulates vascular cell adhesion molecule-1 (VCAM-1), which is the main endothelial ligand of alpha(4) integrins. Intravital microscopy within the rat mesenteric microcirculation showed that 24-h exposure to 1 microg per rat of IL-4 induced a significant increase in leukocyte rolling flux, adhesion and emigration. These responses were decreased by 48, 100 and 99%, respectively in animals treated with TR14035. In conclusion, TR14035, by acting on alpha(4)beta(1) and alpha(4)beta(7) integrins, is an orally active inhibitor of airway leukocyte recruitment and hyper-responsiveness in animal models with potential interest for the treatment of asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Gene Expression Regulation; Integrin alpha4beta1; Integrins; Interleukin-4; Interleukin-5; Leukocyte Rolling; Leukocytes; Lung; Male; Mesenteric Veins; Ovalbumin; Phenylalanine; Pneumonia; Rats; Rats, Inbred BN; RNA, Messenger; Serotonin

Eotaxin is a chemokine that has high potency and selectivity as a chemoattractant agent for eosinophils, signalling exclusively through the CCR3 receptor. Eotaxin is upregulated in the lungs within 3 h of antigen challenge, levels peak at 6 h in lung tissue and bronchoalveolar (BAL) fluid, and fall within 12 h of exposure. This study aimed to look at the effect(s) of eotaxin inhalation on airway function in guinea pigs, to determine if the expected inflammatory cell (eosinophil) infiltration could induce airway hyperreactivity (AHR) and a bronchoconstrictor response equivalent to the late asthmatic response (LAR) seen after antigen challenge. Animals were sensitised with 100 microg/ml OA with a dose on days 1 and 5. Airway responses to inhaled eotaxin (10 or 20 microg/ml) were determined by whole body plethysmography as the change in specific airway conductance (sGaw). Inhaled histamine (1mM) was used to investigate AHR, and cell influx was determined by BAL. Senitised animals exposed to 10 microg/ml eotaxin did not reveal a bronchoconstrictor response or AHR and cellular infiltration to the lungs was not evident 24 h after exposure. Both sensitised and non-sensitised animals exposed to 20 microg/ml eotaxin however revealed a significant bronchoconstrictor response 6h post-challenge, with reductions in sGaw of -27.0+/-6.6% and -32.3+/-6.8%, respectively. Both groups also displayed a bronchoconstrictor response to inhaled histamine 24h after exposure, indicating AHR, and a significant increase in both total and differential cell counts. Sensitised animals, however, revealed a significant increase in cell influx compared to non-sensitised animals. Nebulised eotaxin can reveal a LAR, AHR to inhaled histamine, and cellular infiltration to the lungs, possibly via the mobilisation of eosinophils from the bone marrow, and their subsequent recruitment to the airways.

Topics: Administration, Inhalation; Airway Resistance; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cell Count; Chemokine CCL11; Chemokines, CC; Data Interpretation, Statistical; Guinea Pigs; Histamine; Hypersensitivity; Inflammation; Male; Ovalbumin; Plethysmography, Whole Body; Respiratory Physiological Phenomena

Macrophage migration inhibitory factor (MIF) is a pro-inflammatory cytokine that reportedly counteracts the anti-inflammatory effect of endogenous glucocorticoids. There have only been a few reports that demonstrate a potential link between MIF and bronchial asthma. In an attempt to further clarify the precise role of MIF in asthma, the present authors examined the effect of anti-MIF antibody (Ab) on airway inflammation and airway hyperresponsiveness in an ovalbumin-immunised rat asthma model. Actively immunised Brown Norway rats received ovalbumin inhalation with or without treatment of anti-MIF Ab. The levels of MIF in bronchoalveolar lavage fluid were significantly elevated after the ovalbumin challenge. An immunohistochemical study revealed positive immunostaining for MIF in bronchial epithelium, even in nonsensitised rats, and the MIF staining in bronchial epithelium was enhanced after the ovalbumin challenge. Anti-MIF Ab significantly decreased the number of total cells, neutrophils and eosinophils in the bronchoalveolar lavage fluid of the ovalbumin-challenged rats, and also attenuated the ovalbumin-induced airway hyperresponsiveness to ovalbumin and methacholine. However, anti-MIF Ab did not affect the level of serum ovalbumin-specific IgE, suggesting that anti-MIF Ab did not suppress immunisation itself. The results indicate that macrophage migration inhibitory factor plays a crucial role in airway inflammation and airway hyperresponsiveness in asthma.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Enzyme-Linked Immunosorbent Assay; Eosinophils; Interleukin-13; Leukocyte Count; Macrophage Migration-Inhibitory Factors; Male; Methacholine Chloride; Ovalbumin; Rats; Rats, Inbred BN; Respiratory Mucosa

Mucosal tolerance can be induced by oral or nasal administration of soluble proteins and results in the suppression of cellular and/or humoral immune responses to the specific antigen.. To compare the effect of oral or nasal ovalbumin administration before, during or after immunization on the development of cellular and humoral immune responses by using a murine asthma model.. To induce lung allergic inflammation, animals were immunized twice with ovalbumin/aluminum hydroxide gel and challenged twice with ovalbumin. To induce tolerance, BALB/c mice received ovalbumin by the oral or nasal routes for 3 consecutive days. The ovalbumin administration was initiated before (day -7), during (day 0), or after immunization (day 7).. Airway eosinophilia, airway hyperreactivity, mucus hypersecretion, and cytokine production were suppressed when oral or nasal ovalbumin administration was initiated before immunization. Oral but not nasal ovalbumin exposure suppressed ovalbumin-specific nonanaphylactic IgG(1) antibodies, whereas both routes suppressed the production of anaphylactic IgG(1) and IgE antibodies. Mucosal ovalbumin administration at day 0 inhibited all T(H)2-mediated allergic parameters but not nonanaphylactic IgG(1) antibodies. Finally, ovalbumin exposure 7 days after immunization was still effective in suppressing lung allergy but not ovalbumin-specific anaphylactic IgG(1) and IgE antibodies.. We show that the effectiveness of mucosal tolerance depends on route and time and presents a hierarchical pattern of suppression in the following order: lung allergic responses > anaphylactic antibodies > ovalbumin-specific IgG(1).

Topics: Administration, Intranasal; Administration, Oral; Anaphylaxis; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Immune Tolerance; Immunization; Immunoglobulin E; Immunoglobulin G; Male; Mice; Mice, Inbred BALB C; Mucous Membrane; Ovalbumin; Pulmonary Eosinophilia; Time Factors

Actinidia polygama is one of the well known herb used in oriental medicine for treatment of anti-inflammatory and many allergic diseases. Anti-asthmatic effects of A. polygama in the development of OVA-induced eosinophilia and hyperresponsiveness in murine model of asthma have not been fully investigated in vivo. Cyclosporine A (CsA) has been shown to inhibit single allergen-induced allergic inflammation such as eosinophilic and lymphocytic infiltration and mRNA expression for interleukin (IL)-4 and IL-5. Asthma is a chronic inflammatory disease of the mucosa and is associated with excess production of Th2 cytokines and eosinophil influx in lung. To clarify the anti-inflammatory and anti-asthmatic effects of A. polygama and CsA, we examined the influence of A. polygama fructus extract (APF) and CsA on the development of pulmonary eosinophilic inflammation in murine model of asthma. Our results have shown that APF and CsA have profound inhibitory effects on the accumulation of eosinophills into airways, with the reduction of eosinophil and total lung leukocyte number by reducing IL-4, IL-5, IL-13 and IgE levels in the BALF. Moreover, APF decreased eosinophil CCR3 expression and CD11b expression in lung cells. These results indicate that APF has a deep inhibitory effect on airway inflammation and hyperresponsiveness in murine model of asthma and play a crucial role as an immunomodulator which possess anti-inflammatory and anti-asthmatic property by modulating the relationship between Th1/Th2 cytokine imbalance.

Topics: Actinidia; Animals; Anti-Asthmatic Agents; Antibodies; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cyclosporine; Enzyme-Linked Immunosorbent Assay; Eosinophilia; Flow Cytometry; Mice; Ovalbumin; Plant Extracts; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Serine Proteinase Inhibitors

P110delta phosphoinositide 3-kinase (PI3K) plays a pivotal role in the recruitment and activation of certain inflammatory cells. Recent findings revealed that the activity of p110delta also contributes to allergen-IgE-induced mast cell activation and vascular permeability. We investigated the role of p110delta in allergic airway inflammation and hyperresponsiveness using IC87114, a selective p110delta inhibitor, in a mouse asthma model. BALB/c mice were sensitized with OVA and, upon OVA aerosol challenge, developed airway eosinophilia, mucus hypersecretion, elevation in cytokine and chemokine levels, up-regulation of ICAM-1 and VCAM-1 expression, and airway hyperresponsiveness. Intratracheal administration of IC87114 significantly (P<0.05) attenuated OVA-induced influx into lungs of total leukocytes, eosinophils, neutrophils, and lymphocytes, as well as levels of IL-4, IL-5, IL-13, and RANTES in a dose-dependent manner. IC87114 also significantly (P<0.05) reduced the serum levels of total IgE and OVA-specific IgE and LTC(4) release into the airspace. Histological studies show that IC87114 inhibited OVA-induced lung tissue eosinophilia, airway mucus production, and inflammation score. In addition, IC87114 significantly (P<0.05) suppressed OVA-induced airway hyperresponsiveness to inhaled methacholine. Western blot analyses of whole lung tissue lysates shows that IC87114 markedly attenuated the OVA-induced increase in expression of IL-4, IL-5, IL-13, ICAM-1, VCAM-1, RANTES, and eotaxin. Furthermore, IC87114 treatment markedly attenuated OVA-induced serine phosphorylation of Akt, a downstream effector of PI3K signaling. Taken together, our findings implicate that inhibition of p110delta signaling pathway may have therapeutic potential for the treatment of allergic airway inflammation.

Topics: Adenine; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cell Adhesion Molecules; Chemokines; Chemotaxis, Leukocyte; Class I Phosphatidylinositol 3-Kinases; Cytokines; Eosinophilia; Female; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Models, Animal; Mucus; Ovalbumin; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Phosphorylation; Protein Processing, Post-Translational; Proto-Oncogene Proteins c-akt; Quinazolines

Complement component 5 (C5) has been described as either promoting or protecting against airway hyperresponsiveness (AHR) in experimental allergic asthma, suggesting pleomorphic effects of C5. Here we report that local pharmacological targeting of the C5a receptor (C5aR) prior to initial allergen sensitization in murine models of inhalation tolerance or allergic asthma resulted in either induction or marked enhancement of Th2-polarized immune responses, airway inflammation, and AHR. Importantly, C5aR-deficient mice exhibited a similar, increased allergic phenotype. Pulmonary allergen exposure in C5aR-targeted mice resulted in increased sensitization and accumulation of CD4+ CD69+ T cells associated with a marked increase in pulmonary myeloid, but not plasmacytoid, DC numbers. Pulmonary DCs from C5aR-targeted mice produced large amounts of CC chemokine ligand 17 (CCL17) and CCL22 ex vivo, suggesting a negative impact of C5aR signaling on pulmonary homing of Th2 cells. In contrast, C5aR targeting in sensitized mice led to suppressed airway inflammation and AHR but was still associated with enhanced production of Th2 effector cytokines. These data suggest a dual role for C5a in allergic asthma, i.e., protection from the development of maladaptive type 2 immune responses during allergen sensitization at the DC/T cell interface but enhancement of airway inflammation and AHR in an established inflammatory environment.

Topics: Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Dendritic Cells; Disease Models, Animal; Immunity, Innate; Inflammation Mediators; Lung; Mice; Mice, Inbred BALB C; Mice, Transgenic; Mucus; Ovalbumin; Receptor, Anaphylatoxin C5a; Signal Transduction; Th2 Cells

Matrix metalloproteinase 9 (MMP-9) and its tissue inhibitor of metalloproteinase 1 (TIMP-1) are hypothesized to play a role in the pathogenesis of airway remodeling in asthma.. We have used a mouse model of airway remodeling to determine the pattern of expression of MMP-9 and TIMP-1 in airway epithelium and peribronchial cells, and assess whether TIMP-1, an inhibitor of MMP-9, is expressed at the same sites in the airway. In addition, we have investigated whether immunostimulatory sequences (ISSs) of DNA modulate levels of expression of MMP-9, TIMP-1, and peribronchial fibrosis.. Levels of lung MMP-9 and TIMP-1 were assessed by zymography, ELISA, and immunohistochemistry.. Repetitive ovalbumin challenge induced a significant increase in levels of MMP-9, TIMP-1, and peribronchial collagen deposition. The pattern of expression of MMP-9 and TIMP-1 in the remodeled airway was significantly different. MMP-9 but not TIMP-1 was expressed in airway epithelium, whereas both MMP-9 and TIMP-1 were expressed in peribronchial inflammatory cells. ISS significantly reduced expression of MMP-9 in airway epithelium (which immunostained positive for Toll receptor 9), as well as in peribronchial inflammatory cells. In vitro studies demonstrated that ISS inhibited bone marrow macrophage generation of MMP-9.. Allergen-induced peribronchial fibrosis is associated with expression of MMP-9 and TIMP-1 at different anatomical sites in the remodeled airway. The ability of ISS to inhibit the expression of MMP-9 in airway epithelium (a site where its inhibitor TIMP-1 is not induced by allergen challenge) may be important in determining whether ISS contributes to reductions in airway remodeling by reducing levels of MMP-9.. Immunostimulatory sequences of DNA, which are being investigated as novel therapeutics in asthma, inhibit airway remodeling in mice as well as epithelial expression of MMP-9, an enzyme that degrades the extracellular matrix proteins surrounding the airway.

Topics: Adjuvants, Immunologic; Allergens; Animals; Bronchial Hyperreactivity; Collagen; Disease Models, Animal; DNA; Fibrosis; Immunohistochemistry; Matrix Metalloproteinase 9; Mice; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Ovalbumin; Respiratory Mucosa; Tissue Inhibitor of Metalloproteinase-1

Generalized underrepresentation of IFN-gamma has been implicated in the development of allergic asthma. However, the role of local IFN-gamma in the lung during the development of this disease has not been completely elucidated. We studied the influence of local pulmonary IFN-gamma expression on the development of allergen-induced lung inflammation. To restrict our analysis to IFN-gamma expression in the lung and to exclude influences of systemic IFN-gamma production, we generated a transgenic mouse line with a targeted deletion of the IFN-gamma gene and constitutive, lung-specific IFN-gamma expression (Clara cell 10 [CC10]-IFN-gamma-tg-IFN-gamma-KO mice), and compared allergen-induced airway inflammation in these mice with that of wild-type and IFN-gamma- KO mice on the C57BL/6 background. Cytokine quantification in lungs of mice with allergic airway inflammation revealed that pulmonary IFN-gamma expression increased expression of IL-5 and IL-13. Consistent with this observation, eosinophilia in bronchoalveolar lavage of CC10-IFN-gamma-tg-IFN-gamma-KO mice was profoundly increased, indicating that this critical component of asthma is enhanced by local IFN-gamma expression. In contrast, airway hyperresponsiveness and anti-ovalbumin-IgE serum levels were reduced by local IFN-gamma expression. Together, our results demonstrate pleiotropic action of constitutive IFN-gamma expression in the lung, and question the therapeutic value of IFN-gamma in allergic asthma. Local expression of IFN-gamma in the lung increases markers of allergic airway inflammation, but decreases airway hyperresponsiveness in a murine model of allergic-asthma.

Topics: Animals; Asthma; Biomarkers; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Crosses, Genetic; Disease Models, Animal; Eosinophilia; Immunoglobulin E; Inflammation; Interferon-gamma; Interleukin-13; Interleukin-5; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; RNA, Messenger

T cells expressing a type-2 T helper profile of cytokines (Th2 cells) have been demonstrated to play an important role in the initiation and progression of allergic asthma, and it is well known that Fas ligand (FasL) induces apoptosis when bound to its receptor, Fas. In the present study, we examined the possibility of modulating asthma manifestations by dendritic cells (DCs) genetically engineered to express FasL (DC-FasL), which could deliver a death signal to T cells in an antigen-specific manner. The delivery of DC-FasL into ovalbumin (OVA)-immunized allergic mice decreased the airway hyper-responsiveness (AHR). Moreover, we established a mouse model of airway inflammation by using an adoptive transfer of Th2 cells derived from ovalbumin T cell receptor transgenic mice to study the effect of DC-FasL on airway reactivity. The administration of DC-FasL in Th2-cell-induced allergic mice had significantly decreased AHR, airway inflammation, and IL-4, IL-5 and IL-13 production. Furthermore, the numbers of OVA-specific T cells were decreased in the lung of mice receiving DC-FasL. These results demonstrate that FasL-expressing dendritic cells might be applied for the modulation of allergic responses.

Topics: Adenoviridae; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cells, Cultured; Cytokines; Dendritic Cells; Disease Models, Animal; Fas Ligand Protein; Female; Immunization; Inflammation; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes; Th2 Cells; Transfection

CTLA-4 is a negative regulator of T-cell activation, and its inhibitory effects can be accomplished either by competition with CD28 or by transmitting negative signals through its intracellular domain. To utilize the cytoplasmic domain of CTLA-4 to suppress allergic inflammation, we fused it to a novel protein-transduction domain in the human transcriptional factor Hph-1. Transduction efficiency was verified in vitro and in vivo after ocular, intranasal and intradermal administration. After transduction into T cells, the Hph-1-ctCTLA-4 fusion protein inhibited the production of interleukin (IL)-2, and downregulated CD69 and CD25. Intranasal administration of Hph-1-ctCTLA-4 resulted in markedly reduced infiltration of inflammatory cells, secretion of T helper type 2 (T(H)2) cytokines, serum IgE levels and airway hyper-responsiveness in a mouse model of allergic airway inflammation. These results indicated that Hph-1-ctCTLA-4 constitutes an effective immunosuppressive protein drug for potential use in the treatment of allergic asthma, via nasal administration.

Topics: Administration, Intranasal; Animals; Antigens, CD; Antigens, Differentiation; Asthma; Bronchial Hyperreactivity; Carrier Proteins; CTLA-4 Antigen; Female; Humans; Immunoconjugates; Immunosuppressive Agents; Inflammation; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Polycomb Repressive Complex 1; Protein Structure, Tertiary; Recombinant Fusion Proteins; Transduction, Genetic

Airway hyperresponsiveness (AHR) is a hallmark of bronchial asthma. Important features of this exaggerated response to bronchoconstrictive stimuli have mostly been investigated in vivo in intact animals or in vitro in isolated tracheal or bronchial tissues. Both approaches have important advantages but also certain limitations. Therefore, the aim of our study was to develop an ex vivo model of isolated lungs from sensitized mice for the investigation of airway responsiveness (AR). BALB/c mice were sensitized by intraperitoneal ovalbumin (Ova) and subsequently challenged by Ova inhalation. In vivo AR was measured in unrestrained animals by whole body plethysmography after stimulation with aerosolized methacholine (MCh) with determination of enhanced pause (P(enh)). Twenty-four hours after each P(enh) measurement, airway resistance was continuously registered in isolated, perfused, and ventilated lungs on stimulation with inhaled or intravascular MCh or nebulized Ova. In a subset of experiments, in vivo AR was additionally measured in orotracheally intubated, spontaneously breathing mice 24 h after P(enh) measurement, and lungs were isolated further 24 h later. Isolated lungs of allergen-sensitized and -challenged mice showed increased AR after MCh inhalation or infusion as well as after specific provocation with aerosolized allergen. AR was increased on days 2 and 5 after Ova challenge and had returned to baseline on day 9. AHR in isolated lungs after aerosolized or intravascular MCh strongly correlated with in vivo AR. Pretreatment of isolated lungs with the beta(2)-agonist fenoterol diminished AR. In conclusion, this model provides new opportunities to investigate mechanisms of AHR as well as pharmacological interventions on an intact organ level.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Bronchodilator Agents; Female; Fenoterol; In Vitro Techniques; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Models, Biological; Ovalbumin; Time Factors

Patients with allergic asthma have symptoms of a predominant T(H)2 response, including airway eosinophilic inflammation and increased mucous production in the lungs. This accompanies increased airways responsiveness, which can be life threatening. Because T(H)2 cells and cytokines have been implicated in contributing to these symptoms, pathways that control the development of these cells or that regulate their cytokine production represent good targets for controlling this disease.. We have previously shown that mice lacking the tyrosine kinase inducible T-cell kinase (ITK) have drastically reduced airway inflammation in a model of allergic asthma. However, it was not clear whether this translated into reduced airways hyperresponsiveness. We have analyzed tracheal responsiveness and airways hyperresponsiveness of wild-type (WT) and ITK null mice during induction of experimental allergic asthma.. Experimental allergic asthma was induced in WT and ITK knockout mice. Tracheal responses to carbachol, acetylcholine, and potassium chloride were analyzed. Airways hyperresponsiveness to methacholine challenge was also analyzed in allergen-challenged mice, along with lung and bronchoalveolar lavage fluid T(H)2 cytokine message and protein.. ITK null mice have reduced tracheal responses to cholinergic challenge in vitro before as well as after allergen challenge. These mice also have reduced airways hyperresponsiveness in response to allergen challenge, which could be rescued by transferring WT splenocytes or purified WT CD4+ T cells. This reduced airways response was preferentially accompanied by reduced expression of T(H)2 cytokines in the lungs.. Our results indicate that the tyrosine kinase ITK and its function in T cells represent an attractive target for antiasthmatic drugs.. Modulating the expression or activity of ITK may be a novel strategy to block allergic airway inflammation.

Topics: Acetylcholine; Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Carbachol; CD4-Positive T-Lymphocytes; Cytokines; In Vitro Techniques; Lung; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Protein-Tyrosine Kinases; Th2 Cells; Trachea

Imiquimod is an imidazoquinoline, which class of compounds are known to have antiviral and antitumoural properties. In recent studies, it was shown that imiquimod modulates the T helper cell type Th1/Th2 response by inducing the production of Th1 cytokines like IFN-gamma, and by inhibiting the Th2 cytokines like interleukin (IL)-4. Several investigators have shown that T-bet and GATA-3 are master Th1 and Th2 regulatory transcription factors. This study investigated whether imiquimod treatment inhibited airway inflammation by modulating transcription factors T-bet and GATA-3.. Thirty-six male SD rats were randomly divided into a control group, an asthmatic group, and an imiquimod group, which was exposed to an aerosol of 0.15% imiquimod. Twenty-four hours after the last ovalbumin (OVA) challenge, airway responsiveness was measured and changes in airway histology were observed. The concentrations of IL-4, IL-5 and IFN-gamma in bronchoalveolar lavage fluid (BALF) and serum were measured by enzyme linked immunosorbent assay (ELISA). The mRNA expressions of IL-4, IL-5, IFN-gamma, T-bet and GATA-3 in lung and in CD4(+) T cells were determined by reverse transcription polymerase chain reaction (RT-PCR). The protein expressions of T-bet and GATA-3 were measured by Western blot.. It was demonstrated that imiquimod 1) attenuated OVA induced airway inflammation; 2) diminished the degree of airway hyperresponsiveness (AHR); 3) decreased the Th2 type cytokines and increased Th1 type cytokines mRNA and protein levels; 4) modulated the Th1/Th2 reaction by inhibiting GATA-3 production and increasing T-bet production.. Imiquimod treatment inhibits OVA induced airway inflammation by modulating key master switches GATA-3 and T-bet that result in committing T helper cells to a Th1 phenotype.

Topics: Administration, Inhalation; Aminoquinolines; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Cytokines; Eosinophils; GATA3 Transcription Factor; Gene Expression Regulation; Imiquimod; Lung; Male; Ovalbumin; Rats; Rats, Sprague-Dawley; RNA, Messenger; T-Box Domain Proteins; Transcription Factors

Vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) are mediators of airway inflammation and remodeling in asthma.. This study investigates a potential relationship between VEGF and MMP-9, and the mechanisms by which VEGF signaling regulates MMP-9 expression in asthma.. We evaluated whether levels of VEGF correlated with levels of MMP-9 in the sputum of asthma patients, and the effect of VEGF receptor inhibitors on MMP-9 expression in murine model of asthma.. We have found that levels of VEGF and MMP-9 are significantly higher in the sputum of patients with asthma than in healthy control subjects, and a significant correlation is found between the levels of VEGF and MMP-9. This study with the ovalbumin-induced model of asthma revealed the following typical pathophysiologic features of asthma in the lungs: increased numbers of inflammatory cells of the airways, airway hyperresponsiveness, increased vascular permeability, and increased levels of MMP-9 and VEGF. Administration of VEGF receptor inhibitors reduced the pathophysiologic signs of asthma and decreased the increased expression of MMP-9 after ovalbumin inhalation.. These results indicate that there is a close relationship between VEGF and MMP-9 expression and that inhibition of VEGF receptor down-regulates the expression of MMP-9. These findings suggest that VEGF signaling regulates MMP-9 expression and plays a critical role in initiation and maintenance of asthma.

Topics: Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cinnamates; Down-Regulation; Humans; Immunohistochemistry; Indoles; Interleukin-13; Interleukin-4; Interleukin-5; Matrix Metalloproteinase 9; Ovalbumin; Sputum; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2

Dendritic cells are the most powerful of the antigen-presenting cells and are known to play important roles in sensitization and inflammation in allergen-specific asthma. Various cytokines and chemokines are involved in the maturation and activation of dendritic cells. Among them is CC chemokine ligand (CCL)21, a key chemokine in the entry of naive T cells and antigen-stimulated dendritic cells into the T-cell zones of secondary lymphoid organs, which is a critical process in antigen-specific T-cell activation.. We studied the role of CCL21 in airway inflammation in asthma by using BALB/c-plt/plt (plt) mice, which possess genetic defects in expression of both CCL21 and CCL19.. Plt and control BALB/c mice were immunized with ovalbumin and alum 4 times and thereafter were subjected to a 2-week regimen of ovalbumin inhalation.. In plt mice, ovalbumin-specific IgE response was delayed compared with control BALB/c mice, but they had the same level of response after final immunization. Although airway inflammation and response to acetylcholine were significantly reduced compared with BALB/c mice, significant eosinophilic inflammation and hyperresponsiveness were also observed in plt mice after 2 weeks of inhalation. Four weeks after cessation of inhalation, airway inflammation and hyperresponsiveness in plt mice were greater than in BALB/c mice. At the time of resolution of airway inflammation, IL-10 production was enhanced in BALB/c mice but not in plt mice.. The chemokines CCL21 and CCL19 were critical for resolution of airway inflammation.. The findings about the chemokines for induction and resolution of inflammation are key to establishing a new strategy for asthma immunotherapy.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Chemokine CCL19; Chemokine CCL21; Chemokines, CC; Disease Models, Animal; Epitopes, T-Lymphocyte; Humans; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Inflammation; Inflammation Mediators; Lymph Nodes; Mice; Mice, Inbred BALB C; Mice, Mutant Strains; Ovalbumin; Receptors, CCR7; Receptors, Chemokine; Receptors, Interleukin-2

We investigated the therapeutic potential of a newly developed antifibrotic agent, pirfenidone, to regulate airway remodeling and the development of allergic airway inflammation and airway hyperresponsiveness after chronic allergen challenge. Administration of pirfenidone after sensitization but during the period of ovalbumin challenge significantly prevented the development of airway hyperresponsiveness and prevented eosinophil and lymphocyte accumulation in the airways. IL-4, IL-5, and IL-13 levels in bronchoalveolar lavage fluid and ovalbumin-specific serum IgE antibody levels were also significantly reduced. Treatment with pirfenidone significantly reduced transforming growth factor-beta1 and platelet-derived growth factor levels in bronchoalveolar lavage fluid. Pirfenidone reduced the expression of transforming growth factor-beta1, the development of goblet cell hyperplasia and subepithelial collagenization, and the increases in contractile elements in the lung. These data indicate that pirfenidone may play an important role in the treatment of asthma and has the potential reduce or prevent airway remodeling.

Topics: Allergens; Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Proliferation; Cytokines; Eosinophils; Goblet Cells; Hyperplasia; Immunoglobulin E; Interleukin-13; Interleukin-4; Interleukin-5; Leukocytes, Mononuclear; Lymphocytes; Mice; Mice, Inbred BALB C; Ovalbumin; Platelet-Derived Growth Factor; Pyridones; Transforming Growth Factor beta; Transforming Growth Factor beta1

An increasing prevalence of allergic diseases, such as atopic dermatitis, allergic rhinitis and bronchial asthma, has been noted worldwide. Allergic asthma strongly correlates with airway inflammation caused by the unregulated production of cytokines secreted by allergen-specific type-2 T helper (Th2) cells. This study aims to explore the therapeutic effect of the airway gene transfer of IL-12, IL-10 and TGF-beta on airway inflammation in a mouse model of allergic asthma.. BALB/c mice were sensitized to ovalbumin (OVA) by intraperitoneal injections with OVA and challenged by nebulized OVA. Different cytokine gene plasmids or non-coding vector plasmids were instilled daily into the trachea up to one day before the inhalatory OVA challenge phase.. Intratracheal administration of IL-10, IL-12 or TGF-beta can efficiently inhibit antigen-induced airway hyper-responsiveness and is able to largely significantly lower the number of eosinophils and neutrophils in bronchoalveolar lavage fluid of ovalbumin (OVA) sensitized and challenged mice during the effector phase. Furthermore, the effect of IL-10 plasmids is more remarkable than any other cytokine gene plasmid. On the other hand, local administration of IL-4 gene plasmids before antigen challenge can induce severe airway hyper-responsiveness (AHR) and airway eosinophilia.. Our data demonstrated that anti-inflammatory cytokines, particularly IL-10, have the therapeutic potential for the alleviation of airway inflammation in murine model of asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokines, CC; Disease Models, Animal; Female; Gene Transfer Techniques; Interleukin-10; Interleukin-12; Interleukin-4; Leukotriene B4; Mice; Mice, Inbred BALB C; Ovalbumin; Pulmonary Eosinophilia; Transforming Growth Factor beta

The precise role of each nitric oxide (NO) synthase (NOS) isoform in the pathobiology of asthma is not well established. Our objective was to investigate the contribution of constitutive NO synthase (cNOS) and inducible NOS (iNOS) isoforms to lung mechanics and inflammatory and remodeling responses in an experimental model of chronic allergic pulmonary inflammation. Guinea pigs were submitted to seven ovalbumin exposures with increasing doses (1 approximately 5 mg/ml) for 4 wk. The animals received either chronic L-NAME (N-nitro-L-arginine methyl ester, in drinking water) or 1,400 W (iNOS-specific inhibitor, intraperitoneal) treatments. At 72 h after the seventh inhalation of ovalbumin solution, animals were anesthetized, mechanically ventilated, exhaled NO was collected, and lung mechanical responses were evaluated before and after antigen challenge. Both L-NAME and 1,400 W treatments increased baseline resistance and decreased elastance of the respiratory system in nonsensitized animals. After challenge, L-NAME increased resistance of the respiratory system and collagen deposition on airways, and decreased peribronchial edema and mononuclear cell recruitment. Administration of 1,400 W reduced resistance of the respiratory system response, eosinophilic and mononuclear cell recruitment, and collagen and elastic fibers content in airways. L-NAME treatment reduced both iNOS- and neuronal NOS-positive eosinophils, and 1,400 W diminished only the number of eosinophils expressing iNOS. In this experimental model, inhibition of NOS-derived NO by L-NAME treatment amplifies bronchoconstriction and increases collagen deposition. However, blockage of only iNOS attenuates bronchoconstriction and inflammatory and remodeling processes.

Topics: Administration, Inhalation; Animals; Bronchial Hyperreactivity; Chronic Disease; Enzyme Inhibitors; Eosinophils; Guinea Pigs; Inhalation; Isoenzymes; Lung; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Ovalbumin; Pneumonia; Respiratory Mechanics; Time Factors; Trachea

The effects of nitric oxide (NO) on allergic inflammation are controversial. In particular, the role of endothelial nitric oxide synthase (eNOS) in asthma remains uncertain. In the present study, we examined the effects of overexpression of eNOS on allergic inflammation using eNOS transgenic (eNOS-Tg) mice, in which eNOS protein is overexpressed in the vascular endothelium and airway epithelium. We found that eNOS-Tg mice showed a reduction of the asthmatic response to allergen challenge. Eosinophilic accumulation in the airspaces, eosinophilic activity, and bronchial responsiveness to acetylcholine were significantly attenuated in eNOS-Tg mice, as compared with wild-type mice following ovalbumin sensitization/challenge, even though the levels of circulating eosinophils were comparable in the wild-type and eNOS-Tg mice. The concentrations of eotaxin in the bronchoalveolar lavage fluid were significantly less in eNOS-Tg mice than in the wild-type mice. In addition, immunohistochemical analysis showed that the expressions of both intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 on the pulmonary endothelium of eNOS-Tg mice was decreased compared with the controls. These results suggest that chronic eNOS overexpression contributes to the suppression of allergic inflammation by reducing the production of eotaxin in the airspaces and/or the expression of adhesion molecules in the vascular endothelium.

Topics: Acetylcholine; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokines, CC; Enzyme Inhibitors; Eosinophil Peroxidase; Intercellular Adhesion Molecule-1; Interferon-gamma; Interleukin-4; Leukocyte Count; Lung; Mice; Mice, Inbred C57BL; Mice, Transgenic; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase Type III; Ovalbumin; Pneumonia; Pulmonary Eosinophilia; Vascular Cell Adhesion Molecule-1

Propolis, which has been used widely in folk medicine, has been shown to exhibit various biological activities but its immunoregulatory and anti-inflammatory activities in intact animals have not been well studied. We investigated these activities of propolis using an ovalbumin-induced asthma animal model. Mice were immunized and sensitized by exposure to ovalbumin (OVA) antigen and administered with low- (65 mg/kg body weight) and high-dose (325 mg/kg body weight) propolis water extracts by tube feeding. The serum OVA-specific IgE titer and cytokine profiles in cultured splenocytes and bronchoalveolar lavage fluids (BALF) were analyzed. The number of eosinophils in BALF was counted. Here we demonstrate that propolis extracts can suppress the serum levels of OVA-specific IgE and IgG(1), and airway hyperresponsiveness (AHR) in OVA-sensitized mice. There are no significant differences in the concentration of eotaxin or the number of eosinophils in BALF among the four groups. However, the higher dose of propolis extracts decreases the level of IL-5 in BALF. The splenocytes from mice administered with propolis extracts (low- and high-dose groups) exhibit a strong inhibition of IL-10 secretion and up-regulation of IFN-gamma secretion in splenocytes stimulated with concanavalin A (ConA). In addition, cytokine (IFN-gamma, IL-6, and IL-10) secretion in OVA-stimulated splenocytes from the propolis groups was significantly lower than that in the control group. These results suggest that propolis extracts may be a potential novel therapeutic agent for asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokines, CC; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Immunoglobulin G; Leukocyte Count; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Propolis; Spleen

Death receptor-6 (DR6), a member of the death domain-containing TNFR superfamily, is highly expressed in lymphoid tissues and regulated upon lymphocyte activation. Targeted disruption of DR6 results in enhanced CD4(+) T cell proliferation and T helper 2 (Th2) differentiation in vitro, whereas the in vivo role of DR6 in regulating Th2 cell differentiation and effector function remains largely unknown. In the current study, we used a Th2-skewed allergic airway inflammation model induced by ovalbumin (OVA) sensitization and challenge to compare the inflammatory response in the lung of both wild type (WT) and DR6(-/-) mice. DR6(-/-) mice were protected from the development of airway inflammation as evidenced by attenuated eosinophil accumulation and reduced mucus-producing cells in the lining airways of allergen-challenged animals. Consistent with these observations, a profound reduction of Th2 cytokine production (IL-5 and IL-13) was detected in the bronchoalveolar lavage fluid (BAL). Furthermore, a significant increase in the frequency of IFN-gamma secreting cells was observed in the DR6(-/-) mouse lungs after OVA challenge, which may account for the reduced pulmonary Th2 cytokine production. These data point to a critical role of DR6 in regulating airway inflammation in the OVA-induced mouse model of asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Interferon-gamma; Mice; Mice, Knockout; Mucus; Ovalbumin; Pulmonary Eosinophilia; Receptors, Tumor Necrosis Factor; Th2 Cells

CARMA1 has been shown to be important for Ag-stimulated activation of NF-kappaB in lymphocytes in vitro and thus could be a novel therapeutic target in inflammatory diseases such as asthma. In the present study, we demonstrate that mice with deletion in the CARMA1 gene (CARMA1(-/-)) do not develop inflammation in a murine model of asthma. Compared with wild-type controls, CARMA1(-/-) mice did not develop airway eosinophilia, had no significant T cell recruitment into the airways, and had no evidence for T cell activation in the lung or draining lymph nodes. In addition, the CARMA1(-/-) mice had significantly decreased levels of IL-4, IL-5, and IL-13, did not produce IgE, and did not develop airway hyperresponsiveness or mucus cell hypertrophy. However, adoptive transfer of wild-type Th2 cells into CARMA1(-/-) mice restored eosinophilic airway inflammation, cytokine production, airway hyperresponsiveness, and mucus production. This is the first demonstration of an in vivo role for CARMA1 in a disease process. Furthermore, the data clearly show that CARMA1 is essential for the development of allergic airway inflammation through its role in T lymphocytes, and may provide a novel means to inhibit NF-kappaB for therapy in asthma.

Topics: Adoptive Transfer; Allergens; Animals; Apoptosis Regulatory Proteins; Asthma; Bronchial Hyperreactivity; CARD Signaling Adaptor Proteins; Cytokines; Disease Models, Animal; Guanylate Kinases; Immunoglobulin E; Immunophenotyping; Lung; Lymph Nodes; Lymphocyte Activation; Lymphocyte Count; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Respiratory Hypersensitivity; Th2 Cells

Asthma is one of the major public health problems worldwide and the morbidity and mortality of asthma has increased in the past two decades. Accumulating data suggest that unnecessary immune responses and inflammation should be suppressed to treat asthma. The purpose of this study is to investigate the anti-asthmatic effects of DA-9201, an ethanolic extract of black rice (Oryza sativa L. var japonica), on an ovalbumin (OVA)-induced mouse model of asthma. Balb/c mice immunized with OVA were administered with DA-9201 (30, 100 or 300 mg/kg, p.o.) or dexamethasone (3 mg/kg, p.o.) and challenged with 1% aerosolized OVA for 30 min. The effects on airway inflammation, airway hyperresponsiveness (AHR), antibody profiles and cytokines were evaluated. DA-9201 treatment significantly reduced the number of eosinophils in bronchoalveolar lavage fluid (BALF) and ameliorated the AHR. Lung histological features also showed that DA-9201 reduced airway inflammation. Furthermore, DA-9201 treatment decreased IFN-gamma as well as IL-4, IL-5 and IL-13 levels in the supernatant of cultured splenocytes, and suppressed the level of OVA-specific IgG, IgG2a, IgG1 and total IgE in plasma. DA-9201 showed anti-asthmatic effects by suppressing unnecessary immune responses, airway inflammation, eosinophilia, AHR and IgE level. These results suggest DA-9201 might be beneficial for the treatment of asthma.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Bronchoalveolar Lavage Fluid; Chromatography, High Pressure Liquid; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophils; Female; Immunoglobulins; Inflammation; Leukocyte Count; Mice; Mice, Inbred BALB C; Oryza; Ovalbumin; Plant Extracts; Spleen

Prostaglandins (PGs) are potent proinflammatory mediators generated through arachidonic acid metabolism by cyclooxygenase-1 and -2 (COX-1 and COX-2) in response to different stimuli and play an important role in modulating the inflammatory responses in a number of conditions, including allergic airway inflammation. Thymoquinone (TQ) is the main active constituent of the volatile oil extract of Nigella sativa seeds and has been reported to have anti-inflammatory properties. We examined the effect of TQ on the in vivo production of PGs and lung inflammation in a mouse model of allergic airway inflammation. Mice sensitized and challenged through the airways with ovalbumin (OVA) exhibited a significant increase in PGD2 and PGE2 production in the airways. The inflammatory response was characterized by an increase in the inflammatory cell numbers and Th2 cytokine levels in the bronchoalveolar lavage (BAL) fluid, lung airway eosinophilia and goblet cell hyperplasia, as well as the induction of COX-2 protein expression in the lung. Intraperitoneal injection of TQ for 5 days before the first OVA challenge attenuated airway inflammation as demonstrated by the significant decrease in Th2 cytokines, lung eosinophilia, and goblet cell hyperplasia. This attenuation of airway inflammation was concomitant to the inhibition of COX-2 protein expression and PGD2 production. However, TQ had a slight inhibitory effect on COX-1 expression and PGE2 production. These findings suggest that TQ has an anti-inflammatory effect during the allergic response in the lung through the inhibition of PGD2 synthesis and Th2-driven immune response.

Topics: Allergens; Animals; Anti-Inflammatory Agents; Benzoquinones; Bronchial Hyperreactivity; Cyclooxygenase 1; Cyclooxygenase 2; Cytokines; Dinoprostone; Disease Models, Animal; Female; Gene Expression Regulation; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Prostaglandin D2; Th2 Cells

Aqueous extract from the fruiting body of Cryptoporus volvatus has been reported to present anti-tumor, anti-allergy, anti-inflammation and immunomodulatory activities. However, the effect mechanisms of anti-allergy and anti-inflammation are poorly understood. The aim of study is to evaluate whether Cryptoporus polysaccharides (CP) extracted from fruiting body of Cryptoporus volvatus decrease the development of nasal symptoms, airway hyperresponsiveness (AHR) to methacholine (MCh) and the infiltration of eosinophils in nasal mucosa in rat model of allergic rhinitis, and investigate a possible action mechanism of CP by detecting the expression of eotaxin mRNA in nasal mucosa and lung tissues. Rats were immunized with ovalbumin and consecutive topical antigen instillation was performed. Repeated intranasal ovalbumin challenge caused rhinitis symptom, AHR to MCh, eosinophil infiltration and histological alterations into the nasal mucosa and increase of eotaxin mRNA expression in nasal mucosa and lung tissue were examined. Pretreatment with CP 3, 9 and 27 mg kg(-1) (ig) decreased the numbers of sneezing 27.4%, 38.4% and 44.3% and nasal rubbing 27.5%, 34.9% and 47.7% comparison with model group, respectively. CP caused a dose-related inhibition of MCh-induced AHR. CP 27 mg kg(-1) decreased the expression of eotaxin mRNA in the nasal mucosa by 35%. These results suggest CP can relieve the symptom, eosinophil infiltration and injury of tissue in nasal mucosa and lung tissue and AHR of allergic rhinitis in rats. Its action mechanism may be associated with the decrease of eotaxin mRNA expression.

Topics: Airway Resistance; Animals; Anti-Allergic Agents; Bronchial Hyperreactivity; Bronchial Provocation Tests; Chemokine CCL11; Chemokines, CC; Chemotaxis, Leukocyte; Disease Models, Animal; Eosinophils; Fruiting Bodies, Fungal; Gene Expression Regulation; Lung; Lung Compliance; Methacholine Chloride; Nasal Mucosa; Ovalbumin; Polyporaceae; Polysaccharides; Rats; Rats, Sprague-Dawley; Rhinitis, Allergic, Seasonal; RNA, Messenger

IL-10 is a potent anti-inflammatory cytokine, and IL-10-producing regulatory T cells are effective inhibitors of murine asthmatic responses. This study determined whether IL-10-dependent mechanisms mediated the local inhalational tolerance seen with chronic inhalational exposure to antigen.. Wildtype and IL-10(-/-) mice were sensitized with ovalbumin (OVA) and then challenged with daily OVA inhalations for 10 days or 6 weeks.. The 10-day animals developed allergic airway disease, characterized by BAL eosinophilia, histologic airway inflammation and mucus secretion, methacholine hyperresponsiveness, and OVA-specific IgE production. These changes were more pronounced in IL-10(-/-) mice. The 6-week IL-10(-/-) and wildtype animals both developed inhalational tolerance, with resolution of airway inflammation but persistence of OVA-specific IgE production.. IL-10 may have anti-inflammatory effects in the acute stage of murine allergic airways disease, but the cytokine does not mediate the development of local inhalational tolerance with chronic antigen exposure.

Topics: Acute Disease; Administration, Inhalation; Aerosols; Animals; Bronchial Hyperreactivity; Chronic Disease; Disease Models, Animal; Disease Progression; Female; Gene Expression Regulation, Developmental; Immune Tolerance; Immunoglobulin E; Inflammation; Interleukin-10; Leukocyte Count; Male; Mice; Mice, Knockout; Ovalbumin

Co-vaccination with cellular pertussis vaccine down-regulates allergic sensitization to diphtheria and tetanus antigens. Using a murine model, we investigated whether vaccination with diphtheria/tetanus toxoids, administered separately or simultaneously with the whole cell vaccine of Bordetella pertussis, inhibits subsequent allergen-induced immune and inflammatory responses.. BALB/c-mice were vaccinated intracutaneously with a combination of diphtheria and tetanus toxoids or a combination of diphtheria and tetanus toxoids with a whole cell vaccine of B. pertussis (three times, days -21 to -7) prior to systemic sensitization (days 1-14) and repeated airway challenges (days 28-30) with ovalbumin.. Compared with negative controls, systemic sensitization and airway allergen challenges induced high serum levels of allergen-specific IgE, predominant Th2-type cytokine production, airway inflammation and development of in vivo airway hyperreactivity. Vaccination with diphtheria and tetanus toxoids prior to sensitization suppressed IgE formation and development of eosinophilic airway inflammation. Co-vaccination with a whole cell pertussis vaccine inhibited allergen sensitization, airway inflammation and development of in vivo airway hyperreactivity. Prevention was due to an allergen-specific and general shift from a predominant Th2 towards a predominant Th1 immune response.. Vaccination with diphtheria and tetanus toxoids alone or in combination with whole cell pertussis vaccine prior to allergen sensitization prevented allergen-induced Th2 immune responses. Vaccine antigens may down-regulate allergic responses to a range of common allergens.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Proliferation; Cytokines; Diphtheria-Tetanus Vaccine; Diphtheria-Tetanus-Pertussis Vaccine; Disease Models, Animal; Female; Leukocytes, Mononuclear; Mice; Mice, Inbred BALB C; Ovalbumin; Spleen

We have developed an animal model to investigate the mechanisms underlying an acute exacerbation of chronic asthma. Sensitized BALB/c mice were exposed to aerosolized ovalbumin, either as chronic low-level challenge (mass concentration approximately 3 mg/m(3)) for 4 wk, a single moderate-level challenge (approximately 30 mg/m(3)), or chronic low-level followed by single moderate-level challenge (the acute exacerbation group). Compared with animals receiving chronic challenge alone, mice in the acute exacerbation group exhibited a more marked inflammatory response, with involvement of intrapulmonary airways and lung parenchyma, and increased numbers of lymphocytes and eosinophils in bronchoalveolar lavage fluid. They also developed airway hyperreactivity (AHR) to methacholine, demonstrable as increased transpulmonary resistance and decreased compliance. This pattern of AHR was absent in chronically challenged animals, but was also present in animals given single moderate-level challenge. However, compared with animals receiving a single moderate-level challenge, inflammation and AHR were induced more rapidly in the acute exacerbation group. Eosinophil-deficient GATA1 Deltadbl mice exhibited undiminished AHR in the acute exacerbation model. We conclude that in mice with pre-existing airway lesions resembling mild chronic asthma, exposure to a moderately high concentration of inhaled antigen induces features of an acute exacerbation. The inflammatory response involves distal airways and is associated with a distinct pattern of AHR, which develops independent of the enhanced eosinophilic inflammation.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Chronic Disease; Cytokines; Eosinophils; Female; GATA1 Transcription Factor; Humans; Inflammation; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin

An alteration in the balance between a T-helper type 2 cell (Th2) response and a Th1 response may predispose to the development of bronchial asthma. Interleukin-18 (IL-18) has an ability to promote both Th1 and Th2 responses, depending on the surrounding cytokine environment. Reactive oxygen species (ROS) play a crucial role in the pathogenesis of airway inflammation and hyperresponsiveness. Recent studies have demonstrated that antioxidants are able to reduce airway inflammation and hyperreactivity in animal models of asthma. In this study, we used a C57BL/6 mouse model of allergic asthma to examine the effects of antioxidants on the regulation of IL-18 expression. Our present study with ovalbumin-induced murine model of asthma revealed that ROS production in cells from bronchoalveolar lavage fluids was increased and that administration of L-2-oxothiazolidine-4-carboxylic acid or alpha-lipoic acid reduced the increased levels of ROS, the increased expression of IL-18 protein and mRNA, airway inflammation, and bronchial hyperresponsiveness. Our results also showed that antioxidants down-regulated a transcription factor, nuclear factor-kappaB (NF-kappaB), activity. These results indicate that antioxidants may reduce IL-18 expression in asthma by inhibiting the activity of NF-kappaB and suggest that ROS regulate the IL-18 expression.

Topics: Animals; Antioxidants; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Down-Regulation; Female; Glutathione; Glutathione Disulfide; Immunoglobulin E; Interleukin-18; Interleukins; Lung; Mice; Mice, Inbred C57BL; Ovalbumin; Reactive Oxygen Species; RNA, Messenger; Thioctic Acid; Transcription Factor RelA

Allergic airway disease can be refractory to anti-inflammatory treatment, whose cause is unclarified. Therefore, in the present experiment, we have tested the hypothesis that co-exposure to lipopolysacharide (Lps) and allergen results in glucocorticoid-resistant eosinophil airway inflammation and hyper-responsiveness (AHR). Ovalbumin (Ova)-sensitized BALB/c mice were primed with 10 microg intranasal Lps 24 h before the start of Ova challenges (20 min on 3 consecutive days). Dexamethasone (5 mg/kg/day) was given on the last 2 days of Ova challenges. AHR, cellular build-up, cytokine and nitrite concentrations of bronchoalveolar lavage fluid (BALF) and lung histology were examined. To assess the role of iNOS-derived NO in airway responsiveness, mice were treated with a selective inhibitor of this enzyme (1400W) 2 h before AHR measurements. More severe eosinophil inflammation and higher nitrite formation were found in Lps-primed than in non-primed allergized mice. After Lps priming, AHR and concentrations of T-helper type 2 cytokines in BALF were decreased, but still remained significantly higher than in controls. Eosinophil inflammation was partially, while nitrite production and AHR were observed to be largely dexamethasone resistant in Lps-primed allergized animals. 1400W effectively and rapidly diminished the AHR in Ova-sensitized and challenged mice, but failed to affect it after Lps priming plus allergization. In conclusion, Lps inhalation may exaggerate eosinophil inflammation and reduce responsiveness to anti-inflammatory treatment in allergic airway disease.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Dexamethasone; Drug Resistance; Female; Glucocorticoids; Imines; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Nitrates; Nitric Oxide Synthase Type II; Nitrites; Ovalbumin; Pulmonary Eosinophilia

Continuous exposure of sensitized mice to an innocuous antigen, such as OVA, does not lead to chronic airway eosinophilia, but induces antigen unresponsiveness and resolution of the inflammatory response. In this study we explored mechanisms underlying attenuation of the airway inflammatory response, assessed whether the phenomenon is strain-specific, and determined its consequences to airway physiology.. Mice were sensitized and exposed to OVA for two and four weeks. Analysis involved BAL, flow cytometry, adoptive transfer of OVA specific CD4 T cells, ex vivo cytokine expression and response to methacholine challenge.. Chronic exposure to antigen resulted in decreased eosinophilia in 5 different mouse strains. Likewise, numbers of lung CD4 T cells expressing activation and Th2 markers sharply declined following continuous OVA exposure. Transfer studies using OVA TcR transgenic cells revealed that the contraction of lung T cells included antigen-specific cells. Systemically, we observed a loss of Th2 memory effector function. Finally, we observed significantly attenuated airway hyper-responsiveness (AHR) in chronically exposed animals.. Attenuation of airway eosinophilia in response to chronic OVA exposure is independent of genetic background. Airway eosinophilia, but not systemic responses, correlates with and is predictive for airway hyperresponsiveness. Our study contributes to the understanding of immune regulatory processes controlling antigen-driven airway inflammatory responses.

Topics: Adoptive Transfer; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage; Bronchoconstrictor Agents; Cytokines; Eosinophilia; Female; Flow Cytometry; Immunologic Memory; Methacholine Chloride; Mice; Ovalbumin; T-Lymphocytes; Time Factors

Epidemiologic data indicate an increased incidence of asthma in the obese.. Because serum levels of the insulin-sensitizing and anti-inflammatory adipokine adiponectin are reduced in obese individuals, we sought to determine whether exogenous adiponectin can attenuate allergic airway responses.. We sensitized and challenged BALB/cJ mice with ovalbumin (OVA). Alzet micro-osmotic pumps were implanted in the mice to deliver continuous infusions of buffer or adiponectin (1.0 microg/g/d), which resulted in an approximate 60% increase in serum adiponectin levels. Two days later, mice were challenged with aerosolized saline or OVA once per day for 3 days. Mice were examined 24 hours after the last challenge.. OVA challenge increased airway responsiveness to intravenous methacholine, bronchoalveolar lavage fluid cells, and T(H)2 cytokine levels. Importantly, each of these responses to OVA was reduced in adiponectin- versus buffer-treated mice. OVA challenge caused a 30% reduction in serum adiponectin levels and a corresponding decrease in adipose tissue adiponectin mRNA expression. OVA challenge also decreased pulmonary mRNA expression of each of 3 proposed adiponectin-binding proteins, adiponectin receptor 1, adiponectin receptor 2, and T-cadherin.. Our results indicate that serum adiponectin is reduced during pulmonary allergic reactions and that adiponectin attenuates allergic airway inflammation and airway hyperresponsiveness in mice.. The data suggest that adiponectin might play a role in the relationship between obesity and asthma.

Topics: Adiponectin; Adipose Tissue; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cadherins; Female; Gene Expression Regulation; Immunoglobulin E; Leukocytes; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Receptors, Adiponectin; Receptors, Cell Surface; Recombinant Proteins; RNA, Messenger

Bronchial asthma is characterized by inflammation of the airways, which is usually accompanied by increased vascular permeability, resulting in plasma exudation. Vascular endothelial growth factor (VEGF) has been implicated in contributing to asthmatic tissue edema through its effect on vascular permeability. Many cellular responses of VEGF are regulated by the lipid products of phosphoinositide 3-kinase (PI3K). However, the effect of PI3K catalytic subunit p110delta on VEGF-mediated signaling is unknown. Recently, an isoform-specific small molecule inhibitor, IC87114, which is selective for p110delta catalytic activity, has been identified.. We have sought to investigate the role of PI3K-delta, more specifically in the increase of vascular permeability.. Female BALB/c mice were sensitized and challenged with ovalbumin. We have investigated the effect of IC87114 on airway inflammation, T(H)2 cytokines expression, airway hyperresponsiveness, plasma extravasation, hypoxia-inducible factor 1alpha expression, and VEGF expression in a murine model of asthma.. Our current study has revealed that IC87114 reduces antigen-induced airway infiltration of inflammatory cells, secretion of T(H)2 cytokines in lungs, airway hyperresponsiveness, and vascular permeability. Moreover, we have found that inhibition of p110delta reduces ovalbumin-induced upregulation of VEGF level.. These results suggest that PI3K-delta inhibitor attenuates antigen-induced airway inflammation and hyperresponsiveness by preventing vascular leakage in mice.. These findings provide a crucial molecular mechanism for the potential role of PI3K-delta in asthma and other airway inflammatory disorders.

Topics: Adenine; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Capillary Permeability; Cinnamates; Class I Phosphatidylinositol 3-Kinases; Cytokines; Disease Models, Animal; Enzyme Inhibitors; Female; Hypoxia-Inducible Factor 1, alpha Subunit; Indoles; Leukocyte Count; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Phosphoinositide-3 Kinase Inhibitors; Pneumonia; Quinazolines; Receptors, Vascular Endothelial Growth Factor; Vascular Endothelial Growth Factor A

Gastrointestinal allergy often precedes or coexists with respiratory allergy.. We hypothesized that established experimental gastrointestinal allergy would prime for the development of allergic respiratory responses.. BALB/c mice were sensitized with ovalbumin (OVA) in the presence of aluminum potassium sulfate and then subjected to intragastric saline or OVA challenges. After the development of allergen-induced gastrointestinal allergy, mice were intranasally exposed to either saline, OVA, or a neoaeroallergen house dust mite (HDM) extract. Airway inflammation (eg, bronchoalveolar lavage fluid cellularity, cytokine levels, and OVA-specific antibody levels) and airway responsiveness to methacholine exposure were assessed after intranasal allergen exposure.. A single intranasal exposure to OVA induced significantly more airway inflammation in intragastric OVA-challenged mice compared with that seen in intragastric saline-treated mice. Kinetic analysis revealed that the observed amplification of lung inflammation was sustained for up to 12 days after the last intragastric OVA challenge after resolution of blood eosinophilia. When mice with gastrointestinal allergy were repeatedly challenged with HDM in the respiratory tract, they experienced enhanced airway inflammation, including bronchoalveolar lavage fluid eosinophilia and increased IL-13 levels.. Taken together, our results demonstrate that OVA-induced gastrointestinal allergy enhances not only allergic airway responses to OVA but also to HDM, an unrelated aeroallergen.. Experimental gastrointestinal allergy primes for responses to allergens in the respiratory tract, enhancing antigen-specific antibody and T(H)2 cytokine production, airway inflammation, and airway hyperresponsiveness.

Topics: Allergens; Animals; Antigens, Dermatophagoides; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophilia; Female; Food Hypersensitivity; Immunoglobulin A; Immunoglobulin E; Immunoglobulin G; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Respiratory Hypersensitivity

We investigated the role of antioxidants in airway hyperresponsiveness to acetylcholine using young asthma model mice, which were sensitized and stimulated with ovalbumin.. The mice had been fed either a normal diet, an alpha-tocopherol-supplemented diet or a probucol-supplemented diet 14 days before the first sensitization. They were immunized with antigen at intervals of 12 days and, starting from 10 days after the second immunization, they were exposed to antigen 3 times every 4th day using an ultrasonic nebulizer. Twenty-four hours after the last antigen inhalation, airway responsiveness to acetylcholine was measured and bronchoalveolar lavage fluid (BALF) was collected. A blood and lung tissue study was also carried out.. Twenty-four hours after the last antigen challenge, both IL-4 and IL-5 in the BALF of alpha-tocopherol-supplemented mice were significantly decreased. The IL-5 level in probucol-supplemented mice was also decreased, but there was no difference in IL-4 levels. The serum IgE level was decreased in probucol-supplemented mice. Differential cell rates of the fluid revealed a significant decrease in eosinophils due to antioxidant supplementation. Airway hyperresponsiveness to acetylcholine was also repressed in antioxidant-supplemented mice. In histological sections of lung tissue, inflammatory cells and mucus secretion were markedly reduced in antioxidant-supplemented mice. We investigated the antioxidant effect on our model mice by examining 8-isoprostane in BALF and lung tissue, and acrolein in BALF; however, our experiment gave us no evidence of the antioxidant properties of either alpha-tocopherol or probucol contributing to the reduction of airway inflammation.. These findings indicate that alpha-tocopherol and probucol suppress allergic responses in asthma model mice, although these two drugs cause suppression in different ways that are unrelated to antioxidation.

Topics: Acrolein; Allergens; alpha-Tocopherol; Animals; Antioxidants; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Dietary Supplements; Dinoprost; Disease Models, Animal; Eosinophils; Female; Hypersensitivity; Immunoglobulin E; Interleukin-4; Interleukin-5; Mice; Mice, Inbred BALB C; Ovalbumin; Oxidative Stress; Probucol

Tachykinins-like substance P (SP) have been shown to play an important role in initiating and perpetuating airway inflammation. Furthermore, they are supposed to be released into tissues in response to stress.. The aim of this study was to investigate the effects of stress alone or in combination with allergic airway inflammation on SP expression in sensory neurons innervating the mouse airways.. Balb/c mice were systemically sensitized to ovalbumin (OVA), followed by allergen aerosol exposure, and compared with non-sensitized controls. Additionally, OVA-sensitized and -challenged and non-sensitized mice were exposed to sound stress. SP expression in airway-specific and overall vagal sensory neurons of the jugular and nodose ganglion complex was analysed using retrograde neuronal tracing in combination with immunohistochemistry. Preprotachykinin A (PPT-A) mRNA, the precursor for SP, was quantified in lung tissue by real-time PCR. Bronchoalveolar lavage (BAL) fluid was obtained, and cell numbers and differentiation were determined.. Stress and/or allergic airway inflammation significantly increased SP expression in retrograde-labelled vagal sensory neurons from the mouse lower airways compared with controls [stress: 15.7+/-0.8% (% of retrograde-labelled neurons, mean+/-SEM); allergen: 17.9+/-0.4%; allergen/stress: 13.1+/-0.7% vs. controls: 6.3+/-0.3%]. Similarly, SP expression increased in overall vagal sensory neurons identified by the neuronal marker protein gene product (PGP) 9.5 [stress: 9.3+/-0.6% (% of PGP 9.5-positive neurons, means+/-SEM); allergen: 12.5+/-0.4%; allergen/stress: 10.2+/-0.4% vs. controls: 5.1+/-0.3%]. Furthermore, stress significantly increased PPT-A mRNA expression in lung tissue from OVA-sensitized and -challenged animals, and immune cells were identified as an additional source of SP in the lung by immunohistochemistry. Associated with enhanced neuronal SP expression, a significantly higher number of leucocytes were found in the BAL following allergen exposure. Further, stress significantly increased allergen-induced airway inflammation identified by increased leucocyte numbers in BAL fluids.. The central event of sound stress leads to the stimulation of SP expression in airway-specific neurons. However, in sensitized stressed mice an additional local source of SP (probably inflammatory cells) might enhance allergic airway inflammation.

Topics: Allergens; Animals; Biomarkers; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Immunohistochemistry; Lung; Mice; Mice, Inbred BALB C; Models, Animal; Neural Pathways; Neurons, Afferent; Nodose Ganglion; Ovalbumin; Protein Precursors; Respiratory Hypersensitivity; Reverse Transcriptase Polymerase Chain Reaction; Stress, Psychological; Substance P; Tachykinins; Ubiquitin Thiolesterase

Chitosan, a polymer derived from chitin, has been used for nasal drug delivery because of its biocompatibility, biodegradability and bioadhesiveness. Theophylline is a drug that reduces the inflammatory effects of allergic asthma but is difficult to administer at an appropriate dosage without causing adverse side effects. It was hypothesized that adsorption of theophylline to chitosan nanoparticles modified by the addition of thiol groups would improve theophylline absorption by the bronchial epithelium and enhance its anti-inflammatory effects.. We sought to develop an improved drug-delivery matrix for theophylline based on thiolated chitosan, and to investigate whether thiolated chitosan nanoparticles (TCNs) can enhance theophylline's capacity to alleviate allergic asthma.. A mouse model of allergic asthma was used to test the effects of theophylline in vivo. BALB/c mice were sensitized to ovalbumin (OVA) and OVA-challenged to produce an inflammatory allergic condition. They were then treated intranasally with theophylline alone, chitosan nanoparticles alone or theophylline adsorbed to TCNs. The effects of theophylline on cellular infiltration in bronchoalveolar lavage (BAL) fluid, histopathology of lung sections, and apoptosis of lung cells were investigated to determine the effectiveness of TCNs as a drug-delivery vehicle for theophylline.. Theophylline alone exerts a moderate anti-inflammatory effect, as evidenced by the decrease in eosinophils in BAL fluid, the reduction of bronchial damage, inhibition of mucus hypersecretion and increased apoptosis of lung cells. The effects of theophylline were significantly enhanced when the drug was delivered by TCNs.. Intranasal delivery of theophylline complexed with TCNs augmented the anti-inflammatory effects of the drug compared to theophylline administered alone in a mouse model of allergic asthma. The beneficial effects of theophylline in treating asthma may be enhanced through the use of this novel drug delivery system.

Topics: Administration, Intranasal; Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Chitosan; Drug Delivery Systems; Mice; Mice, Inbred BALB C; Nanostructures; Ovalbumin; Theophylline; Thioglycolates

Spreading of sensitization with clinical manifestation of allergy is often observed in atopic individuals.. To investigate the effects of an established primary allergen sensitization on immune responses and airway inflammation/reactivity on secondary allergen sensitization and airway challenges in a murine model.. Balb/c mice were primarily sensitized intraperitoneally with ovalbumin or PBS, followed by systemic sensitization and airway challenges with latex extract as a secondary, unrelated allergen. Purely sham-sensitized animals were included as controls. In a second set of experiments, the primary and secondary allergens were switched.. Sensitization with ovalbumin before sensitization with latex resulted in increased production of total and latex-specific (Hev b 3-specific) IgE and IgG(1), and enhanced secretion of T(H)2-cytokines by spleen mononuclear cells cultured with mitogen compared with single latex-sensitized mice. Furthermore, airway challenges of double-sensitized mice (ovalbumin + latex) with latex caused a significant increase in airway reactivity compared with purely latex-sensitized and challenged animals. These effects were dependent on dosing and timing of the primary sensitization in relation to the secondary sensitization and independent of the primary allergen used.. Primary sensitization boosted systemic T(H)2 immune responses and enhanced the development of airway reactivity after sensitization and airway challenges with a secondary, unrelated allergen. This effect of consecutive priming was dependent on the strength of the primary sensitization but independent of the allergen used. The results explain the increased susceptibility toward sensitization spreading in atopic individuals.. Because sensitization spreading is facilitated by primary sensitization, early prevention measurements or immunotherapy should be considered at this stage of monosensitization.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cells, Cultured; Female; Immunization, Secondary; Latex; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Hypersensitivity; Th2 Cells

The neurotrophin nerve growth factor (NGF) has been implicated as a mediator in allergic asthma. Direct evidence that inhibition of NGF-induced activation of neurotrophin receptors leads to improvement of airway symptoms is lacking. We therefore studied the effects of inhibitors of NGF signal transduction on the development of airway hyper-responsiveness (AHR) and pulmonary inflammation in a guinea-pig model for allergic asthma.. Airway responsiveness to the contractile agonist histamine was measured in vivo in guinea-pigs that were sensitized and challenged with ovalbumin (OVA). Inflammatory cell influx and NGF levels were determined in bronchoalveolar lavage fluid (BALF). Substance P, a key mediator of inflammation, was measured in lung tissue by radioimmunoassay, while substance P immunoreactive neurons in nodose ganglia were measured by immunohistochemistry.. OVA challenge induced an AHR after 24 h in OVA-sensitized guinea-pigs. This coincided with an increase in the amount of NGF in BALF. Simultaneously, an increase in the percentage of substance P immunoreactive neurons in the nodose ganglia and an increase in the amount of substance P in lung tissue were found. We used tyrosine kinase inhibitors to block the signal transduction of the high-affinity NGF receptor, tyrosine kinase A (trkA). Treatment with the tyrosine kinase inhibitors (K252a or tyrphostin AG879) both inhibited the development of AHR, and prevented the increase in substance P in the nodose ganglia and lung tissue completely whereas both inhibitors had no effect on baseline airway resistance. Neither treatment with K252a or tyrphostin AG879 changed the influx of inflammatory cells in the BALF due to allergen challenge.. We conclude that substance P plays a role in the induction of AHR in our model for allergic asthma which is most likely mediated by NGF. As both tyrosine kinase inhibitors AG879 and K252a show a similar inhibitory effect on airway function after allergen challenge, although both tyrosine kinase inhibitors exhibit different non-specific inhibitory effects on targets other than trkA tyrosine kinases, it is likely that the induction of substance P derived from sensory nerves is mediated by NGF via its high-affinity receptor trkA.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carbazoles; Disease Models, Animal; Enzyme Inhibitors; Female; Guinea Pigs; Immunohistochemistry; Indole Alkaloids; Lung; Male; Nerve Growth Factor; Neurons; Nodose Ganglion; Ovalbumin; Receptor, trkA; Signal Transduction; Substance P; Tyrphostins

In conjunction with allergens, diesel exhaust particles act as an adjuvant to enhance IgE responses, inducing expression of cytokines/chemokines and adhesion molecules, and increasing airway hyper-responsiveness (AHR). As most studies were designed to expose animals to diesel exhaust throughout the periods of both sensitization and allergen challenge, it remains unclear whether diesel exhaust (DE) exposure exaggerates airway responses in asthmatic animals.. To study effects of exposure to low-dose DE on AHR and allergic airway inflammation in asthmatic mice.. BALB/c mice were sensitized by intraperitoneal injection of ovalbumin and challenged by intranasal administration with ovalbumin. They were exposed to low-dose DE for 7 h/day, 5 days/week, for up to 12 weeks. AHR to methacholine was evaluated by whole-body plethysmography as well as bronchoalveolar lavage cell analysis and cytokine gene expression in lungs.. Repeated exposure of asthmatic mice to low-dose DE resulted in increased AHR and gene expression of several pro-asthmatic cytokines/chemokines, but these effects rapidly subsided with continued exposure to DE.. Repeated exposure to low-dose DE after ovalbumin challenge exaggerates allergic responses in mice, but effects are not prolonged with continuous DE exposure.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Cytokines; Disease Models, Animal; Female; Inhalation Exposure; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; RNA, Messenger; Vehicle Emissions

Pharmacological inhibition or genetic disruption of cyclooxygenase (COX)-1 or COX-2 exacerbates the inflammatory and functional responses of the lung to environmentally relevant stimuli. To further examine the contribution of COX-derived eicosanoids to basal lung function and to allergic lung inflammation, transgenic (Tr) mice were generated in which overexpression of human COX-1 was targeted to airway epithelium. Although no differences in basal respiratory or lung mechanical parameters were observed, COX-1 Tr mice had increased bronchoalveolar lavage fluid PGE(2) content compared with wild-type littermates (23.0 +/- 3.6 vs 8.4 +/- 1.4 pg/ml; p < 0.05) and exhibited decreased airway responsiveness to inhaled methacholine. In an OVA-induced allergic airway inflammation model, comparable up-regulation of COX-2 protein was observed in the lungs of allergic wild-type and COX-1 Tr mice. Furthermore, no genotype differences were observed in allergic mice in total cell number, eosinophil content (70 vs 76% of total cells, respectively), and inflammatory cytokine content of bronchoalveolar lavage fluid, or in airway responsiveness to inhaled methacholine (p > 0.05). To eliminate the presumed confounding effects of COX-2 up-regulation, COX-1 Tr mice were bred into a COX-2 null background. In these mice, the presence of the COX-1 transgene did not alter allergen-induced inflammation but significantly attenuated allergen-induced airway hyperresponsiveness, coincident with reduced airway leukotriene levels. Collectively, these data indicate that COX-1 overexpression attenuates airway responsiveness under basal conditions but does not influence allergic airway inflammation.

Topics: Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cyclooxygenase 1; Cyclooxygenase 2; Dinoprostone; Humans; Hypersensitivity; Inflammation; Lung; Methacholine Chloride; Mice; Mice, Transgenic; Ovalbumin; Plethysmography, Whole Body; Respiratory Function Tests; Reverse Transcriptase Polymerase Chain Reaction; Transgenes

Extracellular cyclophilins have been well described as chemotactic factors for various leukocyte subsets. This chemotactic capacity is dependent upon interaction of cyclophilins with the cell surface signaling receptor CD147. Elevated levels of extracellular cyclophilins have been documented in several inflammatory diseases. We propose that extracellular cyclophilins, via interaction with CD147, may contribute to the recruitment of leukocytes from the periphery into tissues during inflammatory responses. In this study, we examined whether extracellular cyclophilin-CD147 interactions might influence leukocyte recruitment in the inflammatory disease allergic asthma. Using a mouse model of asthmatic inflammation, we show that 1) extracellular cyclophilins are elevated in the airways of asthmatic mice; 2) mouse eosinophils and CD4+ T cells express CD147, which is up-regulated on CD4+ T cells upon activation; 3) cyclophilins induce CD147-dependent chemotaxis of activated CD4+ T cells in vitro; 4) in vivo treatment with anti-CD147 mAb significantly reduces (by up to 50%) the accumulation of eosinophils and effector/memory CD4+ T lymphocytes, as well as Ag-specific Th2 cytokine secretion, in lung tissues; and 5) anti-CD147 treatment significantly reduces airway epithelial mucin production and bronchial hyperreactivity to methacholine challenge. These findings provide a novel mechanism whereby asthmatic lung inflammation may be reduced by targeting cyclophilin-CD147 interactions.

Topics: Animals; Antibodies, Monoclonal; Asthma; Basigin; Blotting, Western; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage; CD4-Positive T-Lymphocytes; Cyclophilins; Disease Models, Animal; Eosinophils; Extracellular Fluid; Female; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Mucins; Neutrophil Infiltration; Ovalbumin; Pneumonia; Pulmonary Eosinophilia

The authors hypothesized that sevoflurane had different inhibitory effects on hyperreactive airway smooth muscle contractility in different types of hyperreactive airway models.. The effects of sevoflurane on hyperreactive airways in ovalbumin-sensitized and chronic cigarette-smoking guinea pig models were investigated by measuring (1) total lung resistance, (2) smooth muscle tension and intracellular concentration of free Ca, (3) voltage-dependent Ca channel activity, and (4) cyclic adenosine monophosphate levels.. Ovalbumin and muscarinic airway hyperreactivity was seen in ovalbumin-sensitized animals. Enlarged alveolar ducts/alveoli and lesser muscarinic hyperreactivity were observed in chronic cigarette-smoke animals. Although sevoflurane inhibited the acetylcholine-induced increase in total lung resistance in the control and ovalbumin-sensitized models, the anesthetic had a smaller effect in the chronic cigarette-smoking model. Similarly, in the chronic cigarette-smoking model, sevoflurane had a smaller inhibitory effect on carbachol-induced muscle contraction and increase in intracellular concentration of free Ca. Sevoflurane also had a smaller inhibitory effect on voltage-dependent Ca channel activity in the chronic cigarette-smoking group than in the other two groups. The sevoflurane-induced increase in cyclic adenosine monophosphate that was seen in the control and ovalbumin-sensitized groups was significantly suppressed in the chronic cigarette-smoking group.. Although sevoflurane potently inhibited airway contractility in control and ovalbumin-sensitized models, the anesthetic had a smaller effect in a chronic cigarette-smoking model. The different inhibitory effects of sevoflurane on airway contractility depend, at least in part, on different effects on voltage-dependent Ca channel activity and cyclic adenosine monophosphate level.

Topics: Airway Resistance; Anesthetics, Inhalation; Animals; Bronchial Hyperreactivity; Calcium Channels; Cyclic AMP; Guinea Pigs; In Vitro Techniques; Lung Compliance; Male; Methyl Ethers; Muscle Contraction; Muscle, Smooth; Ovalbumin; Respiratory Hypersensitivity; Sevoflurane; Smoking; Trachea

Selective phosphodiesterases (PDE) inhibitors are the new group of antiasthmatic drugs, which integrate antiinflammatory activity with bronchoconstriction counteraction. Selective inhibitors of phosphodiesterase type 4 are used as alternative or assist drugs in treatment of respiratory system diseases. So far glucocorticosteroids remain the most efficient and widely used medicine in the treatment of asthma. However application of glucocorticosteroid is greatly limited because of numerous side effects, what induce to permanent search for new antiasthmatic drugs. Examination new substances are executed on animal models. Guinea pig model is widely used to research course of asthmatic reaction. This model is especially convenient on the ground of that: lung is major shock organ, airway respond to histamine, animals demonstrated early asthmatic reaction (EAR) and late asthmatic reaction (LAR), eosinophils flow in bronchoalveolar space during LAR. In ovalbumin (OA) sensitized guinea pigs hypersensitivity reaction breaks out as a result of OA provocation. Aims of our experiments, execute on guinea pig model were to determine the influence of rolipram (PDE 4 inhibitor) on modulation experimental asthmatic reaction and comparison activity of rolipram versus dexamethasone in attribution to chosen parameters of allergic reaction such as: lung resistance, influx of protein and inflammatory cells in airways, and mastocytes degranulation. Experiments were made on guinea pigs sensitized and provoked with ovalbumin The obtain data indicate that rolipram was effective in reduction the rise of lung resistance during EAR, restricted influx of eosinophils to bronchoalveolar space between 1,5 and 24 hours after provocation, and reduced increase of histamine concentration in bronchoalveolar lavage fluid (BALf). Rolipram had no influence on number of neutrophils present in BALf. Dexamethasone in double dose of 1,2mg/kg effectively bordered the growth of lung resistance during EAR, and broke influx of eosinophils and neutrophils to bronchoalveolar space.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Bronchodilator Agents; Dexamethasone; Disease Models, Animal; Guinea Pigs; Histamine Release; Ovalbumin; Phosphodiesterase Inhibitors; Rolipram

The mechanical properties and Ca2+ sensitivity of an organ-culture model derived from guinea pig airways have been examined. The cultured explants develop airway hyperresponsiveness to pharmacological agonists after 3-day culture, when compared with fresh and ovalbumin-sensitized tissues. The reactivity of cultured explants is dependent on the presence of the epithelium. They are also sensitive to glucocorticosteroid pretreatments, which neutralize the TNFalpha antibody and NF-kappaB inhibitor. Hence, specific immunostaining of NF-kappaB subunits (p65 and p50) was increased in the nuclear extract of cultured explants. In beta-escin-permeabilized preparations, step-increases in pCa revealed enhanced Ca2+ sensitivity of the contractile apparatus in cultured explants, which was prevented by epithelium removal. Pretreatments of cultured explants with neutralizing TNFalpha antibody and NF-kappaB inhibitor consistently reduced their Ca2+ sensitivity. These findings suggest that AHR developed in this organ culture model may be triggered by an inflammatory process mediated by the TNFalpha and NF-kappaB transcription factor, which results in an increased sensitivity to [Ca2+]i.

Topics: Animals; Anti-Inflammatory Agents; Antibodies, Blocking; Antigens; Blotting, Western; Bronchi; Bronchial Hyperreactivity; Calcium; Dose-Response Relationship, Drug; Electrophoresis, Polyacrylamide Gel; Epithelial Cells; Escin; Female; Glucocorticoids; Guinea Pigs; Isometric Contraction; Male; Microscopy, Fluorescence; NF-kappa B; Organ Culture Techniques; Ovalbumin; Subcellular Fractions; Tumor Necrosis Factor-alpha

Several studies showed that the guinea pig represents the animal of choice in the study of the asthma and more exactly in the study of the bronchial hyperreactivity.. In our model of asthma, guinea pigs were made sensitive with ovalbumine (OVA), a protein extracted from the white of egg, and provoked in a way repeated with aerosol challenge of OVA for the group OVA (1 challenge a day during six days). This group was compared with the group controls (C), animals injected with a salt solution (NaCl 0.9%) and receiving aerosol challenge of salt solution. The OVA group was subdivided into two groups: A studied group 6 hours after the aerosol challenge of OVA. A studied group 24 hours after the aerosol challenge of OVA.. We showed an increasing increase of airway hyperresponsiveness to increasing doses of histamine in all groups of animals. This increase was significantly more important 6 hours after the last aerosol challenge of OVA (early airway hyperreactivity, OVA-6 group, n = 8) that at 24 hours after the last aerosol challenge (late airway hyperreactivity, OVA-24 group, n = 8). We had also noted a modification of cellularity in bronchoalveolar lavage fluid with an increase of the total number of cells essentially by increase of the rate of eosinophilia in OVA-6 group (n = 6) compared with OVA-24 group (n = 6) and Control group (n = 6).. The model of bronchial hyperreactivity and modification of cellularity in guinea pig will allow us to envisage studies on the origin of differences of ability to react in the group OVA-6 and OVA-24 and to study the medicinal efficiency of plants used in Senegal in the treatment of the asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Guinea Pigs; Histamine; Histamine Agents; Male; Ovalbumin

The p38 mitogen-activated protein kinase (MAPK) plays a critical role in the activation of inflammatory cells. Therefore, we investigated the antiinflammatory effects of a respirable p38alpha MAPK antisense oligonucleotide (p38alpha-ASO) in a mouse asthma model. A potent and selective p38alpha-ASO was characterized in vitro. Inhalation of aerosolized p38alpha-ASO using an aerosol chamber dosing system produced measurable lung deposition of ASO and significant reduction of ovalbumin (OVA-)-induced increases in total cells, eosinophils, and interleukin 4 (IL-4), IL-5, and IL-13 levels in bronchoalveolar lavage fluid, and dose-dependent inhibition of airway hyperresponsiveness in allergen-challenged mice. Furthermore, inhaled p38alpha-ASO markedly inhibited OVA-induced lung tissue eosinophilia and airway mucus hypersecretion. Quantitative polymerase chain reaction analysis of bronchoalveolar lavage fluid cells and peribronchial lymph node cells showed that p38alpha-ASO significantly reduced p38alpha MAPK mRNA expression. Nose-only aerosol exposure of mice verified the p38alpha-ASO-induced inhibition of OVA-induced pulmonary eosinophilia, mucus hypersecretion, and airway hyperresponsiveness. None of the effects of the p38alpha-ASO were produced by a six-base mismatched control oligonucleotide. These findings demonstrate antisense pharmacodynamic activity in the airways after aerosol delivery and suggest that a p38alpha MAPK ASO approach may have therapeutic potential for asthma and other inflammatory lung diseases.

Topics: Administration, Inhalation; Aerosols; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Male; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinase 14; Mucus; Oligonucleotides, Antisense; Ovalbumin; Polymerase Chain Reaction; Pulmonary Eosinophilia

Airway remodeling, including subepithelial fibrosis, is a characteristic feature of asthma and likely contributes to the pathogenesis of airway hyperresponsiveness. We examined expression of genes related to airway wall fibrosis in a model of chronic allergen-induced airway dysfunction using laser capture microdissection and quantitative real-time PCR. BALB/c mice were sensitized and subjected to chronic ovalbumin exposure over a 12-wk period, after which they were rested and then harvested 2 and 8 wk after the last exposure. Chronic allergen-exposed mice had significantly increased indices of airway remodeling and airway hyperreactivity at all time points, although no difference in expression of fibrosis-related genes was found when mRNA extracted from whole lung was examined. In contrast, fibrosis-related gene expression was significantly upregulated in mRNA obtained from microdissected bronchial wall at 2 wk after chronic allergen exposure. In addition, when bronchial wall epithelium and smooth muscle were separately microdissected, gene expression of transforming growth factor-beta1 and plasminogen activating inhibitor-1 were significantly upregulated only in the airway epithelium. These data suggest that transforming growth factor-beta1 and other profibrotic mediators produced by airway wall, and specifically, airway epithelium, play an important role in the pathophysiology of airway remodeling.

Topics: Allergens; Animals; Biomarkers; Bronchi; Bronchial Hyperreactivity; Epithelium; Female; Fibrosis; Gene Expression Regulation; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Plasminogen Activator Inhibitor 1; RNA, Messenger; Transforming Growth Factor beta; Transforming Growth Factor beta1

In the present study we evaluated the role of neurokinins in the modulation of inducible nitric oxide synthase (iNOS) inflammatory cell expression in guinea pigs with chronic allergic airway inflammation. In addition, we studied the acute effects of nitric oxide inhibition on this response. Animals were anesthetized and pretreated with capsaicin (50 mg/kg sc) or vehicle 10 days before receiving aerosolized ovalbumin or normal saline twice weekly for 4 wk. Animals were then anesthetized, mechanically ventilated, given normal saline or N(G)-nitro-l-arginine methyl ester (l-NAME, 50 mg/kg ic), and challenged with ovalbumin. Prechallenge exhaled NO increased in ovalbumin-exposed guinea pigs (P < 0.05 compared with controls), and capsaicin reduced this response (P < 0.001). Compared with animals inhaled with normal saline, ovalbumin-exposed animals presented increases in respiratory system resistance and elastance and numbers of total mononuclear cells and eosinophils, including those expressing iNOS (P < 0.001). Capsaicin reduced all these responses (P < 0.05) except for iNOS expression in eosinophils. Treatment with l-NAME increased postantigen challenge elastance and restored both resistance and elastance previously attenuated by capsaicin treatment. Isolated l-NAME administration also reduced total eosinophils and mononuclear cells, as well as those cells expressing iNOS (P < 0.05 compared with ovalbumin alone). Because l-NAME treatment restored lung mechanical alterations previously attenuated by capsaicin, NO and neurokinins may interact in controlling airway tone. In this experimental model, NO and neurokinins modulate eosinophil and lymphocyte infiltration in the airways.

Topics: Administration, Inhalation; Aerosols; Airway Resistance; Animals; Bronchial Hyperreactivity; Capsaicin; Chronic Disease; Elasticity; Enzyme Inhibitors; Eosinophils; Guinea Pigs; Inflammation; Lung; Male; Neurokinin A; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Ovalbumin

Asthma is a complex inflammatory disorder involving obstruction, constriction, oedema, remodelling and hyperresponsiveness of the airways. These effects are induced by a raft of mediators, many of which exert their actions by stimulating specific G-protein-coupled receptors linked to a signal transduction pathway involving the monomeric GTPase; rho, and a downstream effector; rho kinase. The aim of this study was to determine whether administration of a selective inhibitor of rho kinase, Y-27632, attenuates airway inflammation, bronchoconstriction and hyperresponsiveness in a murine model of acute allergic inflammation. Intranasal administration of Y-27632 caused a dose-dependent inhibition in the number of eosinophils recovered from bronchoalveolar lavage fluid of ovalbumin-sensitised and challenged (allergic) mice. These inhibitory effects of intranasal Y-27632 on pulmonary eosinophilia were accompanied by a significant inhibition of the development of airways hyperresponsiveness in allergic mice. In additional studies, intranasal Y-27632 inhibited methacholine-induced increases in airways resistance in a time-dependent manner. In conclusion, these findings indicate that activation of rho kinase contributes to bronchoconstriction and eosinophil trafficking in murine models of acute allergic airway inflammation and to the development of airway hyperresponsiveness.

Topics: Administration, Intranasal; Aluminum Hydroxide; Amides; Animals; Australia; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Bronchoconstriction; Disease Models, Animal; Female; Hypersensitivity; Injections, Intraperitoneal; Intracellular Signaling Peptides and Proteins; Lung; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Protein Serine-Threonine Kinases; Pulmonary Eosinophilia; Pyridines; rho-Associated Kinases

Hepatocyte growth factor (HGF) is known to influence a number of cell types and their production of regulatory cytokines. We investigated the potential of recombinant HGF to regulate not only the development of allergic airway inflammation and airway hyperresponsiveness (AHR), but also airway remodeling in a murine model. Administration of exogenous HGF after sensitization but during ovalbumin challenge significantly prevented AHR, as well as eosinophil and lymphocyte accumulation in the airways; interleukin (IL)-4, IL-5, and IL-13 levels in bronchoalveolar lavage (BAL) fluid were also significantly reduced. Further, treatment with HGF significantly suppressed transforming growth factor-beta (TGF-beta), platelet-derived growth factor, and nerve growth factor levels in BAL fluid. The expression of TGF-beta, the development of goblet cell hyperplasia and subepithelial collagenization, and the increases in contractile elements in the lung were also reduced by recombinant HGF. Neutralization of endogenous HGF resulted in increased AHR as well as the number of eosinophils, levels of Th2 cytokines (IL-4, IL-5, and IL-13) and TGF-beta in BAL fluid. These data indicate that HGF may play an important role in the regulation of allergic airway inflammation, hyperresponsiveness, and remodeling.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Eosinophils; Female; Hepatocyte Growth Factor; Humans; Immunoglobulin E; Inflammation; Interferon-gamma; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Recombinant Proteins; Respiratory System; Transforming Growth Factor beta

It has recently been suggested that RhoA plays an important role in the enhancement of the Ca2+ sensitization of smooth muscle contraction. In the present study, a participation of RhoA-mediated Ca2+ sensitization in the augmented bronchial smooth muscle (BSM) contraction in a murine model of allergic asthma was examined.. Ovalbumin (OA)-sensitized BALB/c mice were repeatedly challenged with aerosolized OA and sacrificed 24 hours after the last antigen challenge. The contractility and RhoA protein expression of BSMs were measured by organ-bath technique and immunoblotting, respectively.. Repeated OA challenge to sensitized mice caused a BSM hyperresponsiveness to acetylcholine (ACh), but not to high K+-depolarization. In alpha-toxin-permeabilized BSMs, ACh induced a Ca2+ sensitization of contraction, which is sensitive to Clostridium botulinum C3 exoenzyme, indicating that RhoA is implicated in this Ca2+ sensitization. Interestingly, the ACh-induced, RhoA-mediated Ca2+ sensitization was significantly augmented in permeabilized BSMs of OA-challenged mice. Moreover, protein expression of RhoA was significantly increased in the hyperresponsive BSMs.. These findings suggest that the augmentation of Ca2+ sensitizing effect, probably via an up-regulation of RhoA protein, might be involved in the enhanced BSM contraction in antigen-induced airway hyperresponsiveness.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Calcium; Disease Models, Animal; Male; Mice; Mice, Inbred BALB C; Muscle Contraction; Muscle, Smooth; Ovalbumin; rhoA GTP-Binding Protein

In vivo models have demonstrated that interleukin-13 (IL-13) plays an important role in asthma; however, few studies have evaluated the effect of inhibition of IL-13 on established and persistent disease. In the present study, we have investigated the effect of a therapeutic dosing regimen with an anti-IL-13 monoclonal antibody (mAb) in a chronic mouse model of persistent asthma. BALB/c mice were sensitized to allergen [ovalbumin (OVA); on days 1 and 8] and challenged with OVA weekly from day 22. Anti-IL-13 mAb or vehicle dosing was initiated following two OVA challenges when disease was established. At this time, mice exhibited airway hyperresponsiveness (AHR), increased mucus production, inflammation, and initiation of subepithelial fibrosis compared with saline-challenged mice. Mice received four additional OVA challenges. Treatment with anti-IL-13 mAb inhibited AHR and prevented the further development of subepithelial fibrosis and progression of inflammation. Furthermore, mAb treatment reversed the mucus hyperplasia to basal levels. These effects were associated with an inhibition of cytokines, chemokines, and matrix metalloproteinase-9. These data demonstrate that neutralization of IL-13 can inhibit the progression of established disease in the presence of repeated allergen exposures.

Topics: Animals; Antibodies, Monoclonal; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Cytokines; Disease Progression; Enzyme-Linked Immunosorbent Assay; Female; Inflammation Mediators; Interleukin-13; Lung; Matrix Metalloproteinase 9; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Pulmonary Fibrosis

It is becoming increasingly evident that the compartmentalization of immune responses is governed, in part, by tissue-selective homing instructions imprinted during T cell differentiation. In the context of allergic diseases, the fact that "disease" primarily manifests in particular tissue sites, despite pervasive allergen exposure, supports this notion. However, whether the original site of Ag exposure distinctly privileges memory Th2 immune-inflammatory responses to the same site, while sparing remote tissue compartments, remains to be fully investigated. We examined whether skin-targeted delivery of plasmid DNA encoding OVA via gene-gun technology in mice could generate allergic sensitization and give rise to Th2 effector responses in the skin as well as in the lung upon subsequent Ag encounter. Our data show that cutaneous Ag priming induced OVA-specific serum IgE and IgG1, robust Th2-cytokine production, and late-phase cutaneous responses and systemic anaphylactic shock upon skin and systemic Ag recall, respectively. However, repeated respiratory exposure to aerosolized OVA failed to instigate airway inflammatory responses in cutaneous Ag-primed mice, but not in mice initially sensitized to OVA via the respiratory mucosa. Importantly, these contrasting airway memory responses correlated with the occurrence of Th2 differentiation events at anatomically separate sites: indeed cutaneous Ag priming resulted in Ag-specific proliferative responses and Th2 differentiation in skin-, but not thoracic-, draining lymph nodes. These data indicate that Ag exposure to the skin leads to Th2 differentiation within skin-draining lymph nodes and subsequent Th2 immunity that is selectively manifested in the skin.

Topics: Abdomen; Administration, Cutaneous; Aerosols; Animals; Antigens; Biolistics; Bronchial Hyperreactivity; Cell Proliferation; Ear; Epitopes, T-Lymphocyte; Female; Inflammation; Injections, Intravenous; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Mucosa; Skin; Th2 Cells

Although interleukin (IL)-10 is an immunoregulatory cytokine produced by various cells including T cells, its precise role in asthma remains uncertain. The aim of this study was to investigate the role of IL-10 in experimental asthma using ovalbumin (OVA)-sensitized mice.. Mice were challenged with OVA aerosol, and airway responsiveness and inflammation were measured. OVA-specific IL-10-producing CD4+ T cells were counted from lung cells collected by enzymatic digestion and stimulated ex vivo with OVA. The effects of an anti-IL-10 antibody on airway responsiveness and inflammation were also evaluated.. The OVA challenge caused airway hyperresponsiveness and eosinophilic inflammation. A significant increase in IL-10-producing CD4+ T cells was observed, mainly in the CD45RB(low) subset, for several days after the OVA challenge. Anti-IL-10 antibody treatment before the OVA challenge did not affect eosinophilic inflammation but significantly inhibited airway hyperresponsiveness 24 h after the OVA challenge. However, anti-IL-10 antibody treatment just before the last OVA challenge significantly attenuated the resolution of eosinophilic inflammation without affecting airway responsiveness 2 weeks after the OVA challenge.. Intrinsic IL-10 may have a distinct role in the early and late phases of asthmatic responses. In the early phase, IL-10 induces airway hyperresponsiveness, while in the late phase IL-10 contributes to the resolution of eosinophilic inflammation.

Topics: Allergens; Animals; Antibodies; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4 Lymphocyte Count; CD4-Positive T-Lymphocytes; Eosinophils; Immunization; Interleukin-10; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocyte Subsets; Time Factors

T(H)2-mediated allergic asthma is characterized by eosinophilia, mucus overproduction, and airway hyperresponsiveness (AHR). Although it is clear that T(H)2 cells and their cytokines play an important role in AHR, the roles of T(H)1 cells and neutrophils in AHR are controversial.. We sought to determine the roles of T(H)1 cells and neutrophils in AHR.. Ovalbumin-specific CD4(+) T cells were purified from DO11.10 mice, differentiated into T(H)1 cells, and injected into naive BALB/c, IL-4RalphaKO, or IL-8RKO mice. After ovalbumin antigen challenge, cytokine mRNA levels in lung samples, as well as inflammatory cell types and numbers in bronchoalveolar lavage fluid (BALF), were determined. AHR was assessed by measuring resistance in tracheostomized mice and enhanced pause in freely moving mice.. T(H)1 cells induced AHR as robust as T(H)2 cells. They also induced lung inflammation dominated by neutrophils. Neither AHR nor inflammation were reduced when T(H)1 cells were transferred into IL-4RalphaKO mice. When IL-8RKO mice were used as recipients of T(H)1 cells, neutrophilia was greatly reduced, but the AHR was as strong as that seen in wild-type mice. On the other hand, dexamethasone treatment had no effect on neutrophilia but has significantly reduced AHR. Reduction in AHR was accompanied by a reduction in the numbers of lymphocytes and macrophages in BALF.. T(H)1 cells can induce strong AHR independent of IL-4 and IL-13. The AHR is associated with the presence of lymphocytes and macrophages, but not neutrophils, in BALF. Our results point to a pathway whereby T(H)1 cells mediate AHR independent of neutrophilic inflammation.

Topics: Adoptive Transfer; Animals; Bronchial Hyperreactivity; Bronchitis; Cytokines; Dexamethasone; Glucocorticoids; Interleukin-13; Mice; Mice, Inbred BALB C; Mice, Knockout; Mucus; Neutrophils; Ovalbumin; Receptors, Interleukin-8A; RNA, Messenger; Severity of Illness Index; Th1 Cells; Th2 Cells

Gammadelta T cells suppress airway hyperresponsiveness (AHR) induced in allergen-challenged mice but it is not clear whether the suppression is allergen specific. The AHR-suppressive cells express TCR-Vgamma4. To test whether the suppressive function must be induced, we adoptively transferred purified Vgamma4(+) cells into gammadelta T cell-deficient and OVA-sensitized and -challenged recipients (B6.TCR-Vgamma4(-/-)/6(-/-)) and measured the effect on AHR. Vgamma4(+) gammadelta T cells isolated from naive donors were not AHR-suppressive, but Vgamma4(+) cells from OVA-stimulated donors suppressed AHR. Suppressive Vgamma4(+) cells could be isolated from lung and spleen. Their induction in the spleen required sensitization and challenge. In the lung, their function was induced by airway challenge alone. Induction of the suppressors was associated with their activation but it did not alter their ability to accumulate in the lung. Vgamma4(+) gammadelta T cells preferentially express Vdelta4 and -5 but their AHR-suppressive function was not dependent on these Vdeltas. Donor sensitization and challenge not only with OVA but also with two unrelated allergens (ragweed and BSA) induced Vgamma4(+) cells capable of suppressing AHR in the OVA-hyperresponsive recipients, but the process of sensitization and challenge alone (adjuvant and saline only) was not sufficient to induce suppressor function, and LPS as a component of the allergen was not essential. We conclude that AHR-suppressive Vgamma4(+) gammadelta T cells require induction. They are induced by allergen stimulation, but AHR suppression by these cells does not require their restimulation with the same allergen.

Topics: Adoptive Transfer; Allergens; Ambrosia; Animals; Antigens, Plant; Bronchial Hyperreactivity; Lymphocyte Activation; Mice; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Plant Proteins; Receptors, Antigen, T-Cell, gamma-delta; Serum Albumin, Bovine; T-Lymphocyte Subsets; T-Lymphocytes, Regulatory

Asthma is a worldwide disabling chronic inflammatory airway disease characterized by an intense eosinophilic inflammatory infiltrate on bronchial mucous membranes. Among the complementary therapeutic approaches to asthma, acupuncture has been widely used.. Here we used a rat pulmonary hypersensitivity experimental model that mimics human asthma in order to address whether electroacupuncture (EA) treatment could reduce the inflammatory process.. Experimental animals were divided in four groups: control (C), immobilized (I), sham-acupuncture (SA), and acupuncture (A). All rats were sensitized with heat-solidified hen egg white implant. Using clinical acupuncture points, EA treatment began 2 days after antigen priming and was repeated on alternate days for 2 weeks. Subsequently, animals were challenged by inhalation with aggregated ovalbumin and sacrificed 24 hours later when blood samples, bronchoalveolar lavage (BAL), and lungs were collected.. Histopathologic analyses showed that peribronchial and perivascular inflammatory cell infiltrates were significantly lower in group A compared to groups SA and I (shown to be similar to group C). Furthermore, BAL total cell count and percentage of polymorphonuclears (as well as the differential counts of neutrophils and eosinophils) were significantly reduced in group A compared to group I. Corsticosterone plasma levels were similar in all groups.. Taken together these results show that EA efficiently diminishes the bronchial immune-mediated inflammation induced in rats and that this effect is dependent on the choice of specific acupoints.

Topics: Analysis of Variance; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Electroacupuncture; Male; Ovalbumin; Rats; Rats, Wistar; Time Factors

The prevalence of asthma and allergic disease has increased in many countries, and there has been speculation that immunization promotes allergic sensitization. Bordetella pertussis infection exacerbates allergic asthmatic responses. We investigated whether acellular pertussis vaccine (Pa) enhanced or prevented B. pertussis-induced exacerbation of allergic asthma. Groups of mice were immunized with Pa, infected with B. pertussis, and/or sensitized to ovalbumin. Immunological, pathological, and physiological changes were measured to assess the impact of immunization on immune deviation and airway function. We demonstrate that immunization did not enhance ovalbumin-specific serum immunoglobulin E production. Histopathological examination revealed that immunization reduced the severity of airway pathology associated with sensitization in the context of infection and decreased bronchial hyperreactivity upon methacholine exposure of infected and sensitized mice. These data demonstrate unequivocally the benefit of Pa immunization to health and justify selection of Pa in mass vaccination protocols. In the absence of infection, the Pa used in this study enhanced the interleukin-10 (IL-10) and IL-13 responses and influenced airway hyperresponsiveness to sensitizing antigen; however, these data do not suggest that Pa contributes to childhood asthma overall. On the contrary, wild-type virulent B. pertussis is still circulating in most countries, and our data suggest that the major influence of Pa is to protect against the powerful exacerbation of asthma-like pathology induced by B. pertussis.

Topics: Animals; Asthma; Bordetella pertussis; Bronchial Hyperreactivity; Female; Interleukins; Mice; Mice, Inbred BALB C; Models, Animal; Ovalbumin; Pertussis Vaccine; Whooping Cough

Matrix metalloproteinase (MMP)-9 plays an important role in the pathogenesis of bronchial asthma. Neovastat, having significant antitumor and antimetastatic properties, is classified as a naturally occurring multifunctional antiangiogenic agent. We evaluated the therapeutic effect of Neovastat on airway inflammation in a mouse model of asthma. BALB/c mice were immunized subcutaneously with ovalbumin (OVA) on days 0, 7, 14, and 21 and challenged with inhaled OVA on days 26, 29, and 31. Neovastat was administrated by gavage (5 mg/kg body weight) three times with 12 h intervals, beginning 30 min before OVA inhalation. On day 32, mice were challenged with inhaled methacholine, and enhanced pause (Penh) was measured as an index of airway hyperresponsiveness. The severity of airway inflammation was determined by differential cell count of bronchoalveolar lavage (BAL) fluid. The MMP-9 concentration in BAL fluid samples was measured by ELISA, and MMP-9 activity was measured by zymography. The untreated asthma group showed an increased inflammatory cell count in BAL fluid and Penh value compared with the normal control group. Mice treated with Neovastat had significantly reduced Penh values and inflammatory cell counts in BAL fluid compared with untreated asthmatic mice. Furthermore, mice treated with Neovastat showed significantly reduced MMP-9 concentrations and activity in BAL fluid. These results demonstrate that Neovastat might have new therapeutic potential for airway asthmatic inflammation.

Topics: Airway Resistance; Angiogenesis Inhibitors; Animals; Asthma; Bronchial Hyperreactivity; Cartilage; Disease Models, Animal; Female; Immunization; Inflammation; Matrix Metalloproteinase Inhibitors; Mice; Mice, Inbred BALB C; Ovalbumin; Tissue Extracts; Tissue Inhibitor of Metalloproteinase-1

The effect of dietary supplementation with Vitamin E was studied in sensitized guinea pigs. After measurement of baseline airway reactivity and sensitization with ovalbumin, the animals were randomized into two groups: Group A, on a commercial feed and Group B, on dietary supplementation with oral Vitamin E (0.7 IU/kg). These were challenged with inhaled ovalbumin after 4 weeks. The following outcomes were studied: airway responses to ovalbumin inhalation, airway reactivity, sodium and calcium ion influx in isolated tracheal cells, Na+ K+ ATPase and Ca2+ ATPase activity in tracheal homogenate and plasma malonaldehyde. Sensitization increased airway reactivity in Group A but not in Group B. The tracheal cells of animals in Group B showed significantly lower rates of 45Ca and 22Na influx and lower activities of tracheal Na+ K+ ATPase and Ca2+ ATPase as compared to Group A. Plasma malonaldehyde was similar between two groups. We concluded that Vitamin E suppresses the increase in airway reactivity following sensitization and has membrane stabilizing actions.

Topics: Adenosine Triphosphatases; Animals; Bronchial Hyperreactivity; Calcium; Diet Therapy; Disease Models, Animal; Guinea Pigs; Histamine; Isotopes; Lipid Peroxidation; Male; Ovalbumin; Random Allocation; Time Factors; Trachea; Vitamin E

Recent studies in both human and rodents have indicated that in addition to CD4+ T cells, CD8+ T cells play an important role in allergic inflammation. We previously demonstrated that allergen-sensitized and -challenged CD8-deficient (CD8-/-) mice develop significantly lower airway hyperresponsiveness (AHR), eosinophilic inflammation, and IL-13 levels in bronchoalveolar lavage fluid compared with wild-type mice, and that all these responses were restored by adoptive transfer of in vivo-primed CD8+ T cells or in vitro-generated effector CD8+ T cells (T(EFF)). Recently, leukotriene B4 and its high affinity receptor, BLT1, have been shown to mediate in vitro-generated T(EFF) recruitment into inflamed tissues. In this study we investigated whether BLT1 is essential for the development of CD8+ T cell-mediated allergic AHR and inflammation. Adoptive transfer of in vivo-primed BLT1+/+, but not BLT1-/-, CD8+ T cells into sensitized and challenged CD8-/- mice restored AHR, eosinophilic inflammation, and IL-13 levels. Moreover, when adoptively transferred into sensitized CD8-/- mice, in vitro-generated BLT1+/+, but not BLT1-/-, T(EFF) accumulated in the lung and mediated these altered airway responses to allergen challenge. These data are the first to show both a functional and an essential role for BLT1 in allergen-mediated CD8+ T(EFF) recruitment into the lung and development of AHR and airway inflammation.

Topics: Adoptive Transfer; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD8 Antigens; CD8-Positive T-Lymphocytes; Cell Movement; Cytokines; Egg Proteins; Eosinophils; Female; Humans; In Vitro Techniques; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Peptide Fragments; Receptors, Leukotriene B4

Airway cyclooxygenase-2 (COX-2) is induced by cytokine-mediated inflammation such as occurs in asthma. However, the role of COX-2 in the pathophysiology of asthma is not fully understood.. Allergic inflammation, airway responsiveness to methacholine and mucous cell metaplasia after ovalbumin sensitization in the airways of COX-2 deficient (-/-) mice, COX-2 (+/+) mice and C57BL/6J mice treated with a selective COX-2 inhibitor, nimesulide were assessed. Histology, cell analysis, measurements of arachidonic acid metabolites and Th2 cytokine levels in bronchoalveolar lavage fluid (BALF), and measurement of serum IgE were performed.. Eosinophil infiltration into the airway wall, and the number of eosinophils in BALF were greater in sensitized COX-2 (-/-) mice than in sensitized COX-2 (+/+) mice. The levels of cysteinyl leukotrienes (LTC4/D4/E4), prostaglandin E2 (PGE2) and interleukin (IL)-13 as well as airway responsiveness did not differ in COX-2 (-/-) mice and COX-2 (+/+) mice. However, sensitized COX-2 (-/-) mice had higher LTC4/D4/E4 and lower PGE2 concentrations compared with non-sensitized COX-2 (-/-) mice. The number of PAS/alcian blue-positive airway epithelial cells and serum IgE were elevated in COX-2 (-/-) mice. Nimesulide-treated mice showed augmented eosinophilic inflammation, LTC4/D4/E4 concentrations and mucous cell metaplasia.. These data indicate that COX-2 deficiency augments allergic inflammation and mucous cell metaplasia.

Topics: Animals; Arachidonic Acid; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cyclooxygenase 2; Cyclooxygenase Inhibitors; Cysteine; Dinoprostone; Female; Immunoglobulin E; Inflammation; Interleukin-13; Leukotrienes; Lung; Male; Metaplasia; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Sulfonamides

The aim of this study was to investigate the number of Clara cells and the production and secretion of Clara cell 16 kDa protein (CC16) in a murine model of allergen-induced airway inflammation, as well as the effects of N-acetylcysteine (NAC) on CC16 and Clara cell numbers, in order to determine the mechanism of the anti-inflammatory effect of NAC.. BALB/c mice were divided into control, ovalbumin (OVA) and NAC groups. An allergen-induced airway inflammation model (OVA group) was established by sensitizing and challenging mice with OVA. NAC was administered as an oral treatment. The number of Clara cells and the production of CC16 were determined by immunohistochemistry. The CC16 levels in bronchoalveolar lavage fluid (BALF) were determined by Western blotting.. The proportion of Clara cells in terminal and respiratory bronchioles significantly decreased in the OVA group compared to the control group (P < 0.01). NAC treatment did not change the proportion of Clara cells in the OVA group (P > 0.05). CC16 production by Clara cells in the OVA groups was significantly lower than that of the control group (P < 0.01), but was elevated following NAC treatment (P < 0.05). The CC16 level in BALF of the OVA group was lower than that of the control group (P < 0.01), but was elevated by NAC treatment (P < 0.05). NAC reduced the total number of white cells and the percentage of eosinophils in BALF. Moreover, it inhibited airway inflammation.. The number of Clara cells and the production and secretion of CC16 were reduced in a murine model of allergen-induced airway inflammation. Antioxidants can enhance the expression of CC16, which might be a mechanism by which they suppress airway inflammation.

Topics: Acetylcysteine; Administration, Oral; Animals; Blotting, Western; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Disease Models, Animal; Female; Immunohistochemistry; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Uteroglobin

Epidemiologic studies suggest that tobacco smoke contributes to the prevalence and occurrence of exacerbations in asthma. The effect of active smoking in adolescents with atopy is poorly understood.. We developed an experimental model to investigate the influence of smoking on antigen-induced airway inflammation and airway responsiveness in mice that were previously sensitized.. Ovalbumin (OVA)-sensitized BALB/c mice were exposed to air or mainstream smoke (5 days/week) and to phosphate-buffered saline (PBS) or OVA aerosol (3 times/week) for 2 weeks (n = 8 for each group).. Airway responsiveness to intravenously injected carbachol was increased (p < 0.05) in smoke- and OVA-exposed mice compared with all other groups. There was an additive effect of smoke and OVA exposure on total cell numbers, macrophages, and dendritic cells in bronchoalveolar lavage fluid and on CD4+ and CD8+ T lymphocytes and dendritic cells in lung tissue (p < 0.05 compared with mice exposed to smoke and PBS and to mice exposed to air and OVA). Concurrent smoke and OVA exposure augmented OVA-specific IgE in serum compared with air and OVA exposure. In lavage fluid supernatant, eotaxin was increased in air- and OVA-exposed mice. The further increase observed in the group exposed to both OVA and cigarette smoke came close to formal significance (p = 0.06). Thymus- and activation-regulated chemokine was augmented in mice exposed to either smoke or OVA, without additional effect.. Our data indicate that acute concurrent exposure to allergen and mainstream cigarette smoke enhances airway inflammation and airway responsiveness in previously sensitized mice.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carbachol; Chemokines; Cytokines; Immunoglobulin E; Inflammation; Injections, Intravenous; Male; Mice; Mice, Inbred BALB C; Nicotiana; Ovalbumin; Respiratory Hypersensitivity; Smoke

IL-15 has been shown to accelerate and boost allergic sensitization in mice. Using a murine model of allergic sensitization to OVA, we present evidence that blocking endogenous IL-15 during the sensitization phase using a soluble IL-15Ralpha (sIL-15Ralpha) suppresses the induction of Ag-specific, Th2-differentiated T cells. This significantly reduces the production of OVA-specific IgE and IgG and prevents the induction of a pulmonary inflammation. Release of proinflammatory TNF-alpha, IL-1beta, IL-6, and IL-12 as well as that of Th2 cytokines IL-4, IL-5, and IL-13 into the bronchi are significantly reduced, resulting in suppressed recruitment of eosinophils and lymphocytes after allergen challenge. It is of clinical relevance that the airway hyper-responsiveness, a major symptom of human asthma bronchiale, is significantly reduced by sIL-15Ralpha treatment. Ex vivo analysis of the draining lymph nodes revealed reduced numbers of CD8, but not CD4, memory cells and the inability of T cells of sIL-15Ralpha-treated mice to proliferate and to produce Th2 cytokines after in vitro OVA restimulation. This phenomenon is not mediated by enhanced numbers of CD4(+)/CD25(+) T cells. These results show that IL-15 is important for the induction of allergen-specific, Th2-differentiated T cells and induction of allergic inflammation in vivo.

Topics: Allergens; Animals; Bronchi; Bronchial Hyperreactivity; CD8-Positive T-Lymphocytes; Cells, Cultured; Epitopes, T-Lymphocyte; Growth Inhibitors; Immunoglobulin E; Immunologic Memory; Immunosuppressive Agents; Inflammation; Interleukin-15; Mice; Mice, Inbred BALB C; Ovalbumin; Protein Subunits; Receptors, Interleukin-15; Receptors, Interleukin-2; Respiratory Hypersensitivity; Solubility; T-Lymphocyte Subsets; Th2 Cells

Leukotriene B4 (LTB4) is a rapidly synthesized, early leukocyte chemoattractant that signals via its cell surface receptor, leukotriene B4 receptor 1 (BLT1), to attract and activate leukocytes during inflammation. A role for the LTB4-BLT1 pathway in allergen-induced airway hyperresponsiveness and inflammation is not well defined.. To define the role of the LTB4 receptor (BLT1) in the development of airway inflammation and altered airway function.. BLT1-deficient (BLT1 -/-) mice and wild-type mice were sensitized to ovalbumin by intraperitoneal injection and then challenged with ovalbumin via the airways. Airway responsiveness to inhaled methacholine, bronchoalveolar lavage fluid cell composition and cytokine levels, and lung inflammation and goblet cell hyperplasia were assessed.. Compared with wild-type mice, BLT1 -/- mice developed significantly lower airway responsiveness to inhaled methacholine, lower goblet cell hyperplasia in the airways, and decreased interleukin (IL)-13 production both in vivo, in the bronchoalveolar lavage fluid, and in vitro, after antigen stimulation of lung cells in culture. Intracellular cytokine staining of lung cells revealed that bronchoalveolar lavage IL-13 levels and numbers of IL-13(+)/CD4+ and IL-13(+)/CD8+ T cells were also reduced in BLT1 -/- mice. Reconstitution of sensitized and challenged BLT1 -/- mice with allergen-sensitized BLT1 +/+ T cells fully restored the development of airway hyperresponsiveness. In contrast, transfer of naive T cells failed to do so.. These data suggest that BLT1 expression on primed T cells is required for the full development of airway hyperresponsiveness, which appears to be associated with IL-13 production in these cells.

Topics: Allergens; Animals; Antibody Formation; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; CD3 Complex; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cytokines; Female; Interleukin-13; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Receptors, Leukotriene B4; T-Lymphocytes

Mouse models of allergic airway disease have greatly contributed to our understanding of disease induction and pathogenesis. Although these models typically investigate responses to a single antigen or allergen, humans are frequently exposed to a myriad of allergens, each with distinct antigenic potential.. Given that airway exposure to ovalbumin (OVA), a prototypic innocuous antigen, induces inhalation tolerance, we wished to investigate how this response would be altered if OVA were encountered concurrently with a house dust mite extract (HDM), which we have recently shown is capable of eliciting a robust allergic airway inflammatory response that is mediated, at least in part, by granulocyte-macrophage colony-stimulating factor.. Balb/c mice were exposed daily to HDM (intranasally) followed immediately by exposure to aerosolized OVA for 5 weeks. To allow the inflammatory response elicited by HDM to subside fully, mice were then allowed to rest, unexposed, for 8 weeks, at which time they were rechallenged with aerosolized OVA for 3 consecutive days.. At this time, we observed a robust eosinophilic inflammatory response in the lung that was associated with an increase in bronchial hyperreactivity. Moreover, we documented significantly elevated serum levels of OVA-specific IgE and IgG(1) and increased production of the Th2 cytokines interleukin 4 (IL-4), IL-5, and IL-13 by splenocytes stimulated in vitro with OVA.. Our data demonstrate the potential of a potent allergen such as HDM to establish a lung microenvironment that fosters the development of allergic sensitization to otherwise weak or innocuous antigens, such as OVA.

Topics: Animals; Bronchial Hyperreactivity; Cytokines; Female; Immunoglobulin E; Immunoglobulin G; In Vitro Techniques; Inflammation; Mice; Mice, Inbred BALB C; Ovalbumin; Pyroglyphidae

Mechanisms that protect against asthma remain poorly understood. S-nitrosoglutathione (GSNO), an endogenous bronchodilator, is depleted from asthmatic airways, suggesting a protective role. We report that, following allergen challenge, wild-type mice exhibiting airway hyperresponsivity have increased airway levels of the enzyme GSNO reductase (GSNOR) and are depleted of lung S-nitrosothiols (SNOs). In contrast, mice with genetic deletion of GSNOR exhibit increases in lung SNOs and are protected from airway hyperresponsivity. Our results indicate that endogenous SNOs, governed by GSNOR, are critical regulators of airway responsivity and may provide new therapeutic approaches to asthma.

Topics: Adrenergic beta-Agonists; Alcohol Dehydrogenase; Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchodilator Agents; Female; Glutathione Reductase; Homeostasis; Imines; Isoproterenol; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitric Oxide; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Ovalbumin; S-Nitrosoglutathione; S-Nitrosothiols; Trachea

The effect of Bacillus Calmette-Guérin (BCG) treatment in allergic pulmonary reaction was studied in mice genetically selected accordingly to a High (H-IVA) or Low (L-IVA) antibody responsiveness. Mice were immunized with ovalbumin (OVA) or OVA plus BCG. Two days after nasal antigenic challenge, seric IgE and IgG1 anti-OVA, eosinophils in pulmonary tissue, inflammatory cells in bronchoalveolar lavage and the compliance and conductance of respiratory system were evaluated. H-IVA mice were found more susceptible than L-IVA, and BCG was able to inhibit simultaneously the production of IgE, the bronchopulmonary inflammation and bronchial hyperresponsiveness in these genetically selected mice.

Topics: Animals; BCG Vaccine; Bronchial Hyperreactivity; Enzyme-Linked Immunosorbent Assay; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Inflammation; Interferon-gamma; Leukocytes; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Passive Cutaneous Anaphylaxis; Rats; Rats, Inbred WF

Asthma is associated with increased expression of a typical array of genes involved in immune and inflammatory responses, including those encoding the prototypic Th2 cytokines interleukin (IL) 4, IL-5, and IL-13. Most of these genes contain binding sites for activator protein-1 (AP-1) within their promoter and are therefore believed to depend on AP-1 for their expression, suggesting that this transcription factor could be of particular importance in asthma pathophysiology.. To clarify the role of AP-1 in the effector phase of pulmonary allergy.. Ovalbumin (OVA)-sensitized mice were intratracheally given decoy oligodeoxyribonucleotides (ODNs) specifically directed to AP-1 or scrambled control ODNs before challenge with aerosolized OVA. Twenty-four hours after the last OVA challenge, airway hyperresponsiveness was measured and allergic airway inflammation was evaluated quantitatively. AP-1 decoys were localized using flow cytometry and immunohistochemistry. AP-1 activity in the lung was assessed using electrophoretic mobility shift assay.. Intratracheally delivered AP-1 decoys efficiently targeted airway immune cells, thus precluding AP-1 activation on OVA challenge. Decoy-mediated local inhibition of AP-1 resulted in significant attenuation of all the pathophysiologic features of experimental asthma-namely, eosinophilic airway inflammation, airway hyperresponsiveness, mucous cell hyperplasia, production of allergen-specific immunoglobulins, and synthesis of IL-4, IL-5, and IL-13. Scrambled control ODNs had no detectable effects.. Our results reveal a key role for AP-1 in the effector phase of pulmonary allergy and indicate that specific AP-1 inhibition in the airways may have therapeutic value in the control of established asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cell Survival; Eosinophils; Female; Hypersensitivity; Immune System; Mice; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Ovalbumin; Trachea; Transcription Factor AP-1

Liriope platyphylla is one of the well-known herb used in oriental medicine for treatment asthma and bronchial and lung inflammation. Anti-asthmatic effects of Liriope platyphylla in the development of OVA-induced airway inflammation and murine asthma model have not been fully investigated in vivo. Asthma is a chronic inflammatory disease of the mucosa and is associated with excess production of Th2 cytokines and eosinophil accumulation in lung. To clarify the anti-inflammatory and anti-asthmatic effects of Liriope platyphylla, we examined the influence of liriopis tuber (LRT) on the development of pulmonary eosinophilic inflammation in murine model of asthma. Our results have shown that LRT were demonstrated on the accumulation of eosinophills into airways, with reduction of eosinophil, total lung leukocytes numbers by reduction IL-5, IL-13, IL-4 and IgE levels in the BALF and serum. Moreover, LRT decreased eosinophil CCR3 expression and CD11b expression in lung cells. These results indicate that LRT has a deep inhibitory effects on airway inflammation and hyperresponsiveness in murine model of asthma and play an crucial role as a immunomodulator which possess anti-inflammatory and anti-asthmatic property by modulating the relationship between Th1/Th2 cytokine imbalance.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Eosinophils; Histamine; Immunoglobulin E; Lung; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Phytotherapy; Plant Extracts; Plants, Medicinal; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocytes, Regulatory

Asthma is a disease of chronic airway inflammation in which T helper (Th) 2 cells play a critical role. The molecular mechanisms controlling Th2 differentiation and function are of paramount importance in biology and immunology. PKCzeta has been implicated in the regulation of apoptosis and NF-kappaB, as well as in the control of T-dependent responses, although no defects were detected in naïve T cells from PKCzeta-/- mice. Here, we report that PKCzeta is critical for IL-4 signaling and Th2 differentiation. Thus, PKCzeta levels are increased during Th2 differentiation, but not Th1 differentiation, of CD4+ T cells, and the loss of PKCzeta impairs the secretion of Th2 cytokines in vitro and in vivo, as well as the nuclear translocation and tyrosine phosphorylation of Stat6 and Jak1 activation, essential downstream targets of IL-4 signaling. Moreover, PKCzeta-/- mice display dramatic inhibition of ovalbumin-induced allergic airway disease, strongly suggesting that PKCzeta can be a therapeutic target in asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cell Differentiation; Cytokines; Flow Cytometry; Fluorescent Antibody Technique; Inflammation; Interleukin-4; Isoenzymes; Mice; Mice, Knockout; Ovalbumin; Protein Kinase C; Signal Transduction; Th2 Cells

Nicotinic agonists, including 1,1-dimethyl-4-phenylpiperazinium (DMPP), have anti-inflammatory properties and in some instances smooth muscle relaxing effects. Since inflammation and airway smooth muscle contraction are two major components of asthma, the present authors investigated the effects of DMPP on airway inflammation and airway resistance in a mouse model of asthma. Mice were sensitised and challenged with ovalbumin (OVA) and treated either intraperitoneally or intranasally with DMPP. The effect of DMPP was tested on airway inflammation, airway resistance and on the increase of intracellular calcium in bronchial smooth muscle cells. DMPP given either during sensitisation, OVA challenges or throughout the protocol prevented lung inflammation and decreased the serum level of OVA specific immunoglobulin E. DMPP administration reduced the number of total cells, lymphocytes and eosinophils in the bronchoalveolar lavage (BAL) fluid. Intranasal DMPP administration was as effective as dexamethasone (DEXA) in reducing total cell count and eosinophil counts in BAL fluid. DMPP, but not DEXA, reduced tissue inflammation. Intranasal DMPP, given 10 min before the test, reduced airway responsiveness to metacholine. DMPP also reduced the increase in intracellular calcium in response to bradykinin. In conclusion, these results show that 1,1-dimethyl-4-phenylpiperazinium reduces lung inflammation and prevents airway hyperresponsiveness in the mouse model of asthma.

Topics: Airway Resistance; Analysis of Variance; Animals; Asthma; Biopsy, Needle; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dimethylphenylpiperazinium Iodide; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophils; Female; Immunoglobulin E; Immunohistochemistry; Mice; Mice, Inbred BALB C; Ovalbumin; Probability; Random Allocation; Reference Values

Splenic CD8alpha+ dendritic cells reportedly tolerize T cell responses by inducing Fas ligand-mediated apoptosis, suppressing IL-2 expression, or catabolizing T cell tryptophan reserves through expression of IDO. We report in this study that CD8alpha+, but not CD8alpha-, dendritic cells purified from the spleens of normal mice can tolerize the Th2 responses of cells from asthma phenotype mice through more than one mechanism. This tolerance could largely be reversed in vitro by anti-IL-10 or anti-TGFbeta Ab treatment. However, loss of direct dendritic cell-T cell contact also reduced tolerance, although to a lesser extent, as did adding the IDO inhibitor 1-methyltryptophan or an excess of free tryptophan to the cultures. Within 3 wk of reconstituting asthma phenotype mice with 1 x 10(5) OVA-pulsed CD8alpha+, but not CD8alpha-, dendritic cells, the mice experienced a reversal of airway hyperresponsiveness, eosinophilic airway responses, and pulmonary Th2 cytokine expression. This data indicates that CD8alpha+ dendritic cells can simultaneously use multiple mechanisms for tolerization of T cells and that, in vivo, they are capable of tolerizing a well-established disease complex such as allergic lung disease/asthma.

Topics: Allergens; Animals; Asthma; Biomarkers; Bronchial Hyperreactivity; CD8 Antigens; Cell Communication; Cells, Cultured; Coculture Techniques; Cytokines; Dendritic Cells; Disease Models, Animal; Eosinophilia; Eosinophils; Epitopes, T-Lymphocyte; Immune Tolerance; Immunoglobulin E; Immunoglobulin G; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells

(R)- and (S)-Enantiomers of albuterol likely exert differential effects in patients with asthma. The (R)-enantiomer binds to the beta2-adrenergic receptor with greater affinity than the (S)-enantiomer and is responsible for albuterol's bronchodilating activity. (S)-Albuterol augments bronchospasm and has proinflammatory actions.. The study aim was to determine whether the (S)-enantiomer, in contrast to the (R)-enantiomer, has adverse effects on allergic airway inflammation and hyperresponsiveness in a mouse asthma model.. Mice sensitized to ovalbumin (OVA) intraperitoneally on days 0 and 14 were challenged with OVA intranasally on days 14, 25, and 35. On day 36, 24 hours after the final allergen challenge, the effect of the (R)- and (S)-enantiomers of albuterol (1 mg x kg(-1) x d(-1) administered by means of a miniosmotic pump from days 13-36) on airway inflammation and hyperreactivity was determined.. In OVA-sensitized/OVA-challenged mice, (R)-albuterol significantly reduced the influx of eosinophils into the bronchoalveolar lavage fluid and airway tissue. (R)-Albuterol also significantly decreased airway goblet cell hyperplasia and mucus occlusion and levels of IL-4 in bronchoalveolar lavage fluid and OVA-specific IgE in plasma. Although (S)-albuterol significantly reduced airway eosinophil infiltration, goblet cell hyperplasia, and mucus occlusion, it increased airway edema and responsiveness to methacholine in OVA-sensitized/OVA-challenged mice. Allergen-induced airway edema and pulmonary mechanics were unaffected by (R)-albuterol.. Both (R)- and (S)-enantiomers of albuterol reduce airway eosinophil trafficking and mucus hypersecretion in a mouse model of asthma. However, (S)-albuterol increases allergen-induced airway edema and hyperresponsiveness. These adverse effects of the (S)-enantiomer on lung function might limit the clinical efficacy of racemic albuterol.

Topics: Adrenergic beta-Agonists; Albuterol; Animals; Apoptosis; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Female; Interleukin-13; Interleukin-4; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Stereoisomerism

The pathophysiology of infantile asthma may differ from that in older children or in adults, partly because of the different immune response depending upon maturation. In adult mice, the sensitizing dose of antigen is known to be critical to the polarized development of helper T cell subsets and allergic airway inflammation. We wanted to know the characteristics of allergic airway inflammation of infantile asthma by developing a murine model.. BALB/C mice at different stages of maturation (juvenile: 3 days after birth; adult: 8 weeks of age) were sensitized with 10 or 1,000 microg ovalbumin (OVA) or vehicle. The animals were then challenged with aerosolized OVA or saline once a day during 6 consecutive days. After the final challenge, bronchial hyperresponsiveness (BHR), bronchoalveolar lavage fluid (BALF), histological changes in the airways and immunological status were examined.. In both juvenile and adult animals, sensitization with 10 microg OVA induced the T helper 2 response (elevated IL-4 and decreased IFN-gamma levels). BHR, airway eosinophilia, the inflammatory cell infiltration, goblet cell metaplasia (GCM), and IgE antibody production were more prominent in animals given this dose than 1,000 microg OVA. Among these responses, GCM as well as BALF IL-4, and BHR were comparable between juvenile and adult animals, whereas other parameters were lower in juvenile animals, especially in those given 1,000 microg OVA.. GCM and, consequently, airway mucus hypersecretion may be an important component of allergic airway inflammation in infantile asthma.

Topics: Age Factors; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Eosinophilia; Goblet Cells; Immunoglobulin E; Inflammation; Lung; Metaplasia; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Th2 Cells

We have previously demonstrated that exposure to diesel exhaust particles (DEP) prior to ovalbumin (OVA) sensitization in rats reduced OVA-induced airway inflammation. In the present study, Brown Norway rats were first sensitized to OVA (42.3 +/- 5.7 mg/m3) for 30 min on days 1, 8, and 15, then exposed to filtered air or DEP (22.7 +/- 2.5 mg/m3) for 4 h/day on days 24-28, and challenged with OVA on day 29. Airway responsiveness was examined on day 30, and animals were sacrificed on day 31. Ovalbumin sensitization and challenge resulted in a significant infiltration of neutrophils, lymphocytes, and eosinophils into the lung, elevated presence of CD4+ and CD8+ T lymphocytes in lung draining lymph nodes, and increased production of serum OVA-specific immunoglobulin (Ig)E and IgG. Diesel exhaust particles pre-exposure augmented OVA-induced production of allergen-specific IgE and IgG and pulmonary inflammation characterized by marked increases in T lymphocytes and infiltration of eosinophils after OVA challenge, whereas DEP alone did not have these effects. Although OVA-sensitized rats showed modest response to methacholine challenge, it was the combined DEP and OVA exposure that produced significant airway hyperresponsiveness in this animal model. The effect of DEP pre-exposure on OVA-induced immune responses correlated with an interactive effect of DEP with OVA on increased production of reactive oxygen species (ROS) and nitric oxide (NO) by alveolar macrophages (AM) and alveolar type II (ATII) cells, NO levels in bronchoalveolar lavage fluid, the induction of inducible NO synthase expression in AM and ATII cells, and a depletion of total intracellular glutathione (GSH) in AM and lymphocytes. These results show that DEP pre-exposure exacerbates the allergic responses to the subsequent challenge with OVA in OVA-sensitized rats. This DEP effect may be, at least partially, attributed to the elevated generation of ROS in AM and ATII cells, a depletion of GSH in AM and lymphocytes, and an increase in AM and ATII cell production of NO.

Topics: Air Pollutants; Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Drug Synergism; Drug Therapy, Combination; Epithelial Cells; Inhalation Exposure; Leukocytes; Lymph Nodes; Macrophages, Alveolar; Male; Nitric Oxide; Ovalbumin; Pulmonary Alveoli; Rats; Rats, Inbred BN; Reactive Oxygen Species; T-Lymphocyte Subsets; Vehicle Emissions

The purpose of this study was to investigate the impact of mainstream cigarette smoke exposure (MTS) on allergic sensitization and the development of allergic inflammatory processes. Using two different experimental murine models of allergic airways inflammation, we present evidence that MTS increased cytokine production by splenocytes in response to OVA and ragweed challenge. Paradoxically, MTS exposure resulted in an overall attenuation of the immune inflammatory response, including a dramatic reduction in the number of eosinophils and activated (CD69+) and Th2-associated (T1ST2+) CD4 T lymphocytes in the lung. Although MTS did not impact circulating levels of OVA-specific IgE and IgG1, we observed a striking reduction in OVA-specific IgG2a production and significantly diminished airway hyperresponsiveness. MTS, therefore, plays a disparate role in the development of allergic responses, inducing a heightened state of allergen-specific sensitization, but dampening local immune inflammatory processes in the lung.

Topics: Allergens; Ambrosia; Animals; Antibody Formation; Bronchial Hyperreactivity; Cytokines; Female; Flow Cytometry; Hypersensitivity; Inflammation; Lung; Mice; Mice, Inbred BALB C; Nicotiana; Ovalbumin; Smoke; Tobacco Smoke Pollution

Exposure to diesel exhaust particles (DEP) during the sensitization process has been shown to increase antigen-specific IgE production and aggravate allergic airway inflammation in human and animal models. In this study, we evaluated the effect of short-term DEP exposure on ovalbumin (OVA)-mediated responses using a post-sensitization model. Brown Norway rats were first exposed to filtered air or DEP (20.6 +/- 2.7 mg/m3) for 4 h/day for five consecutive days. One day after the final air or DEP exposure (day 1), rats were sensitized with aerosolized OVA (40.5 +/- 6.3 mg/m3), and then again on days 8 and 15, challenged with OVA on day 29, and sacrificed on days 9 or 30, 24 h after the second OVA exposure or the final OVA challenge, respectively. Control animals received aerosolized saline instead of OVA. DEP were shown to elicit an adjuvant effect on the production of antigen-specific IgE and IgG on day 30. At both time points, no significant airway inflammatory responses and lung injury were found for DEP exposure alone. However, the OVA-induced inflammatory cell infiltration, acellular lactate dehydrogenase activity and albumin content in bronchoalveolar lavage (BAL) fluid, and numbers of T cells and their CD4+ and CD8+ subsets in lung-draining lymph nodes were markedly reduced by DEP on day 30 compared with the air-plus-OVA exposure group. The OVA-induced nitric oxide (NO) in the BAL fluid and production of NO, interleukin (IL)-10, and IL-12 by alveolar macrophages (AM) were also significantly lowered by DEP on day 30 as well as day 9. DEP or OVA alone decreased intracellular glutathione (GSH) in AM and lymphocytes on days 9 and 30. The combined DEP and OVA exposure resulted in further depletion of GSH in both cell types. These results show that short-term DEP exposure prior to sensitization had a delayed effect on enhancement of the sensitization in terms of allergen-specific IgE and IgG production, but caused an attenuation of the allergen-induced airway inflammatory responses.

Topics: Adjuvants, Immunologic; Air Pollutants; Allergens; Animals; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Glutathione; Immunoglobulin E; Inhalation Exposure; L-Lactate Dehydrogenase; Lymph Nodes; Macrophages, Alveolar; Male; Nitric Oxide; Ovalbumin; Rats; Rats, Inbred BN; T-Lymphocyte Subsets; Vehicle Emissions

To clarify the essential role of NKT cells in allergy, we investigated the contribution of NKT cells to the pathogenesis of eosinophilic airway inflammation using alpha-galactosylceramide (alpha-GalCer), a selective ligand for NKT cells. Although continuous administration of alpha-GalCer during ovalbumin (OVA) sensitization increased OVA-specific IgE levels and worsened eosinophil inflammation, a single administration of alpha-GalCer at the time of OVA challenge completely prevented eosinophilic infiltration in wild-type mice. This inhibitory effect of alpha-GalCer was associated with a decrease in airway hyperresponsiveness, an increase in IFN-gamma, and decreases in IL-4, IL-5 and IL-13 levels in the bronchoalveolar lavage fluids. Analysis of lung lymphocytes revealed that production of IFN-gamma increased in NK cells, but not in T or NKT cells, following alpha-GalCer administration. Induction of vascular cell adhesion molecule-1 in the lungs of wild-type mice was also significantly attenuated by treatment with alpha-GalCer. These effects of alpha-GalCer were abrogated in J alpha281-/- mice, which lack NKT cells, and in wild-type mice treated with anti-IFN-gamma Ab. Hence, our data indicate that alpha-GalCer suppresses allergen-induced eosinophilic airway inflammation, possibly by inducing a Th1 bias that results in inhibition of eosinophil adhesion to the lung vessels.

Topics: Animals; Blotting, Western; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Collagen Type I; Enzyme-Linked Immunosorbent Assay; Eosinophils; Flow Cytometry; Galactosylceramides; Inflammation; Interferon-gamma; Interleukin-13; Interleukin-4; Interleukin-5; Killer Cells, Natural; Lung; Mice; Mice, Mutant Strains; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocytes; Vascular Cell Adhesion Molecule-1

Allergic asthma is a multifaceted syndrome consisting of eosinophil-rich airway inflammation, bronchospasm, and airway hyper-responsiveness (AHR). Using a mouse model of allergic asthma, we previously reported that invariant NKT (iNKT) cells increase the severity of this disease. Herein, we demonstrate that a single i.v. injection of alpha-galactosylceramide (alpha-GalCer), 1 h before the first airway allergen challenge of OVA-sensitized mice, abrogates elicitation of AHR, airway eosinophilia, IL-4 and IL-5 production in bronchoalveolar lavage fluid, and specific anti-OVA IgE antibodies. Further, alpha-GalCer administered intranasally also strongly inhibited the major symptoms of asthma in sensitized and challenged mice. Alpha-GalCer treatment induces iNKT cell accumulation in the lungs, and shifts their cytokine profile from pro-asthmatic IL-4 to a protective IFN-gamma production. The role of IFN-gamma from iNKT cells in protection was shown by adoptive transfer of sorted iNKT cells from OVA-sensitized and alpha-GalCer-treated mice which protected immunized recipients from manifesting asthma by an IFN-gamma-dependent pathway. Our findings demonstrate for the first time that alpha-GalCer administered locally inhibits asthma symptoms, even in predisposed asthmatic mice, through an iNKT cell- and IFN-gamma-dependent pathway.

Topics: Administration, Intranasal; Adoptive Transfer; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Flow Cytometry; Galactosylceramides; Hypersensitivity; Injections, Intravenous; Interferon-alpha; Killer Cells, Natural; Male; Mice; Ovalbumin

S-carboxymethylcysteine (S-CMC) has been used as a mucoregulator in respiratory diseases. However, the mechanism of action of S-CMC on allergic airway inflammation has not yet been defined. In the present study, BALB/c mice were initially sensitised and challenged to ovalbumin (OVA) and, weeks later, re-challenged with OVA (secondary challenge). S-CMC (5-100 mg.kg-1) was administered from 2 days before the secondary challenge through to the day of assay. Mice developed airway hyperresponsiveness (AHR) 6 h after the secondary challenge and increased numbers of neutrophils were present in the bronchoalveolar lavage (BAL) fluid. At 72 h after secondary challenge, mice again developed AHR, but the BAL fluid contained large numbers of eosinophils. S-CMC treatment was found to reduce AHR and neutrophilia at 6 h, as well as eosinophilia and AHR at 72 h. These effects appeared to be dose dependent. Goblet cell hyperplasia, observed at 72 h, was reduced by S-CMC. In BAL fluid, increased levels of interferon-gamma, interleukin (IL)-12 and IL-10 and decreased levels of IL-5 and IL-13 were detected. In conclusion, the data indicate that S-carboxymethylcysteine is effective in reducing airway hyperresponsiveness and airway inflammation at two distinct phases of the response to the secondary allergen challenge in sensitised mice.

Topics: Allergens; Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Carbocysteine; Cytokines; Disease Models, Animal; Female; Mice; Mice, Inbred BALB C; Ovalbumin

Bronchial asthma is characterized by chronic airway inflammation and airway remodelling which occurs in both proximal and distal airways. These changes are associated with development of airway hyper-responsiveness and airflow limitation.. This study was aimed to analyse whether chronic inhalative allergen challenges in mice lead to morphological and physiological changes comparable with this phenotype.. For this purpose, BALB/c mice were systemically sensitized to ovalbumin (OVA) followed by aerosol allergen challenges on 2 consecutive days per week for 12 weeks.. In chronically challenged mice, tissue inflammation in proximal as well as distal airways was observed with a predominance of lymphocytes within the cellular infiltrate. In contrast, inflammation in the airway lumen decreased over time. These changes were associated by a shift in bronchoalveolar lavage-cytokine levels from IL-4, IL-5 and TNF-alpha production (during the acute phase) towards markedly increased levels of TGF-beta during the chronic phase. Goblet cell hyperplasia and subepithelial fibrosis occurred throughout the airway tree. In terms of lung function, chronically challenged mice developed persistent bronchial hyper-responsiveness and progressive airflow limitation. Six weeks after OVA aerosol discontinuation, airway inflammation still persisted although lung function was normalized.. These data indicate that our model of chronic aerosol allergen challenges leads to a phenotype of experimental asthma with participation of distal airways and persistence of inflammation thereby resembling many morphological and physiological aspects of human bronchial asthma.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chronic Disease; Cytokines; Disease Models, Animal; Disease Progression; Female; Mice; Mice, Inbred BALB C; Mucous Membrane; Ovalbumin; Transforming Growth Factor beta

Allergic asthma is mediated in part by unregulated Th2 inflammation in response to an allergen. Induction of peripheral tolerance by inoculation of Ags into the anterior chamber of the eye (ocular tolerance) before sensitization blocks Th2 responses. Thus, we proposed that induction of ocular tolerance to the allergen might modulate an ongoing allergen-induced Th2 pathogenesis in the lung. We initiated ocular tolerance in previously immunized mice in a classic mouse model of OVA-induced pulmonary allergic inflammation. In the model of ocular tolerance, the need for inoculation of Ag into the anterior chamber can be bypassed by i.v. inoculation of in vitro-generated tolerogenic (TGF-beta2-treated, Ag-pulsed) APC (tol-APC). We observed that with i.v. inoculation, such tolerogenic APC, but not control APC, reduced eosinophil and lymphocyte pulmonary infiltration in experimental mice. Similarly, production of Th2 cytokines (IL-4, -5, and -13), but not IFN-gamma, was reduced. Importantly, airway hyperresponsiveness and mucus production were significantly reduced after treatment with the tol-APC. We also show that in vitro suppression of IL-13 production from OVA-sensitized effector T cells was mediated by CD8+, not CD4+, T regulatory cells. Thus, i.v. inoculation of the tol-APC induced peripheral tolerance that suppressed Th2-mediated pathogenesis in the lungs of presensitized mice. The ability of the tol-APC to induce peripheral tolerance and suppress existing Th2 immune inflammation may lead to novel therapies for pulmonary allergic inflammation and its related pathology.

Topics: Adoptive Transfer; Animals; Antigen-Presenting Cells; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Gene Expression; Immune Tolerance; Injections, Intravenous; Lung; Mice; Oligonucleotide Array Sequence Analysis; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; T-Lymphocyte Subsets

In guinea pigs sensitized with ovalbumin the respiratory epithelium lost its ability to modulate the responses of airway smooth muscles to histaminergic stimuli. Incubation of bronchial segments with IL-5 potentiated the contractile responses of bronchial smooth muscles to histamine in both intact and sensitized animals. Incubation of bronchial segments with IL-5 receptors moderated contractile activity of segments from sensitized pigs, but not in the segments from intact controls.

Topics: Animals; Bronchi; Bronchial Hyperreactivity; Drug Synergism; Guinea Pigs; Histamine; In Vitro Techniques; Interleukin-5; Male; Muscle Contraction; Muscle, Smooth; Ovalbumin; Respiratory Mucosa

Increasing evidence suggests that macrophages (Mphi) play a crucial downregulatory role in the initiation and progression of allergic asthma. Recently, the current authors demonstrated that ovalbumin (OVA)-loaded Mphi (OVA-Mphi) suppress subsequent OVA-induced airway manifestations of asthma and that this effect could be potentiated upon selective activation. In the present study, the authors further delineated the underlying pathway by which Mphi exert this immunosuppressive effect. To examine the migration of OVA-Mphi, cells were labelled with 5'chloromethylfluorescein diacetate (CMFDA) and were administered (i.v.) into OVA-sensitised BALB/c mice. After 20 h, the relevant organs were dissected and analysed using fluorescent microscopy. Allergen-specificity was investigated by treating OVA-sensitised mice with keyhole limpet haemocyanin (KLH)-Mphi activated with immunostimulatory sequence oligodeoxynucleotide (ISS-ODN). By lengthening the period between treatment and challenge to 4 weeks it was examined whether OVA-Mphi exerted an immunosuppressive memory response. Strikingly, CMFDA-labelled Mphi were not trapped in the lungs, but migrated to the spleen. ISS-ODN-stimulated KLH-Mphi failed to suppress OVA-induced airway manifestations of asthma. Moreover, treatment with ISS-ODN-stimulated OVA-Mphi was still effective after lengthening the period between treatment and challenge. These data demonstrate that allergen-loaded macrophages can induce an indirect immunosuppressive response that is allergen-specific and long lasting, which are both hallmarks of a memory lymphocyte response.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Desensitization, Immunologic; Disease Models, Animal; Down-Regulation; Enzyme-Linked Immunosorbent Assay; Immunoglobulin E; Macrophages; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Probability; Respiratory Hypersensitivity; Sensitivity and Specificity; Statistics, Nonparametric

Airway hyperresponsiveness (AHR) is one of the most important characteristics of asthma. This study investigated the parameters, by which assess the airway responsiveness under tidal ventilation.. Female BALB/c mice were sensitized and challenged with ovalbumin (OVA) (group A), and part of them were treated with budesonide aerosol (group B). All the mice were anaesthetized and mechanically ventilated. The values of tidal volume (Vt), airway pressure (PA), airway flow (F), expiratory lung resistance (RL) and dynamic compliance of the thorax and lung (CT-L) were recorded by the AniRes2003 animal lung function system. In addition, the expiratory volume in the first 0.1 second after the start of expiration (EV0.1) was obtained according to the flow-volume (F-V) curve. The maximal or minimal values of EV0.1, RL and CT-L were documented after each dose of methacholine (MCH) and compared with values from negative control group (group C).. (1) When the dose of MCH reached 100 ng/g or 200 ng/g, the decrease of Vt in group A was much more significant than group C (P = 0.001, < 0.001 respectively), but not so between groups B and group C (P = 0.974, 0.362 respectively). (2) With the dose of 25, 50, 100 or 200 ng/g MCH, the decrease in percentage of EV0.1 in group A was much higher than group C (P = 0.012, 0.025, 0.001, 0.003 respectively), while that in group B showed no significant difference as compared with group C (P = 0.507, 0.896, 0.972, 0.785). (3) RL and CT-L: with the dose of 200 ng/g MCH, there was a statistically significant increase of RL in group A compared to group B or group C (P < 0.001, < 0.001 respectively), but no significant difference between groups B and C (P = 0.266). With doses of 100 ng/g and 200 ng/g MCH, there was a statistically significant decrease of CT-L in group A compared to group B (P = 0.001, = 0.001) and group C (P < 0.001, < 0.001 respectively), but no significant difference between groups B and C (P = 0.775, 0.310). (4) Histopathology: there were eosinophilic predominant peribronchial and perivascular inflammatory influx in murine lungs after OVA sensitizing and challenging, which could be counteracted by inhalation of budesonide in group B.. The decline in EV0.1 in response to MCH challenge correlated with simultaneous changes in Vt, RL and CT-L, but more sensitively than all the other parameters. The decline in EV0.1 and inflammation in murine lung could be significantly alleviated by inhalation of nebulized budesonide solution, which indicated that EV0.1 to MCH is a valid measure of AHR in mice.

Topics: Airway Resistance; Animals; Bronchial Hyperreactivity; Budesonide; Female; Lung Compliance; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin

Bromelain, a clinically used pineapple extract and natural product, has reported anti-inflammatory and immunomodulatory activities. The purpose of this study was to determine the effect of bromelain treatment in an ovalbumin (OVA)-induced murine model of allergic airway disease (AAD).. To establish AAD, mice were sensitized with intraperitoneal (i.p.) OVA/alum and challenged with daily OVA aerosols. Mice were treated i.p. with either saline, 2 or 6 mg/kg bromelain, twice daily for four consecutive days. Bronchoalveolar lavage leukocytes and cytokines, lung histology, airway hyperresponsiveness, and lymphocyte populations via flow cytometry were compared between groups.. Bromelain treatment of AAD mice resulted in reduced total BAL leukocytes, eosinophils, CD4+ and CD8+ T lymphocytes, CD4+/CD8+ T cell ratio, and IL-13.. Bromelain attenuated development of AAD while altering CD4+ to CD8+ T lymphocyte populations. The reduction in AAD outcomes suggests that bromelain may have similar effects in the treatment of human asthma and hypersensitivity disorders.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Bromelains; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Female; Flow Cytometry; Fluorescent Antibody Technique; Hyaluronan Receptors; Hypersensitivity; Mice; Mice, Inbred C57BL; Ovalbumin

Nuclear factor kappa B (NF-kappaB) regulates the expression of multiple cytokines, chemokines, and cell adhesion molecules that are involved in the pathogenesis of asthma. We investigated the anti-asthmatic effects and the mechanism of action of DA-9201, an extract of the black rice, in a mouse model of asthma. Mice immunized with ovalbumin (OVA) were administered with DA-9201 (30, 100 or 300 mg/kg) or dexamethasone (DEXA, 3 mg/kg) for 2 weeks and challenged with aerosolized OVA during the last 3 days. Anti-asthmatic effects were assessed by means of enhanced pauses, level of total IgE and Th2 cytokines in plasma or bronchoalveolar lavage fluid (BALF), the percentage of eosinophils in BALF, and histopathological examination. The expression of NF-kappaB in nuclear and cytoplasmic fraction and its DNA-binding activity in lung tissues were analyzed by means of Western blotting and electrophoretic gel mobility shift assay (EMSA), respectively. DA-9201 significantly reduced airway hyperresponsiveness (AHR), total IgE level in plasma and BALF, IL-4, IL-5, and IL-13 levels in BALF, and the percentage of eosinophils in BALF. Tissue inflammation was significantly improved by DA-9201 treatment. In addition, DA-9201 dramatically suppressed the expression of NF-kappaB and its DNA-binding activity. These results suggest that DA-9201 may be useful for the treatment of asthma and its efficacy is related to suppression of NF-kappaB pathway.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Down-Regulation; Eosinophilia; Ethanol; Female; Immunoglobulin E; Lung; Mice; Mice, Inbred BALB C; NF-kappa B; Oryza; Ovalbumin; Phytotherapy; Plant Extracts; Th2 Cells

The choice of BCG vaccine strain may play an important role in vaccination efficiency.. To investigate whether the suppressive effects of BCG on asthma depend on the strain of BCG.. Female BALB/c mice were injected intraperitoneally with 1 of 4 different strains of BCG (1 X 10(6) CFU): Pasteur F1173P2, Tokyo 172, Tice, and Connaught. Seven days later, the animals were sensitized by 2 injections of ovalbumin (20 microg) at 2-week intervals before being provoked with 1% ovalbumin aerosols on 3 successive days. Thereafter, the mice underwent a methacholine bronchial challenge and were killed to quantify the inflammatory cells and cytokines in bronchoalveolar lavage fluid and the supernatant of cultured splenocytes.. The eosinophil proportion in the bronchoalveolar lavage fluid was significantly lower and the concentration of interferon-gamma and the interferon-gamma-interleukin 5 (IL-5) ratio in the supernatant of cultured splenocytes were significantly higher in each of the BCG-treated groups (n=10 per group) than in the asthma control group (n=15). However, the methacholine sensitivity (P < .05) and IL-5 concentration (P < .01) in the supernatant of cultured splenocytes were significantly lower only in the group treated with the Tokyo strain of BCG. There was a significant positive correlation between IL-5 and IL-10 concentrations (r = 0.79; P < .001).. The 4 strains of BCG suppressed asthma to different degrees, but all strains induced a shift in the T(H)1/T(H)2 balance toward T(H)1 without increasing IL-10-related regulatory T-cell activity.

Topics: Animals; Asthma; BCG Vaccine; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Female; Humans; Immunoglobulin E; Mice; Mice, Inbred BALB C; Mycobacterium bovis; Ovalbumin; Respiratory Function Tests; Species Specificity; Specific Pathogen-Free Organisms; Vaccination

Phosphodiesterase (PDE)-5 degrades guanosine 3',5'cyclic monophosphate (cGMP) and its inhibitor sildenafil citrate (Viagra) treats erectile dysfunction by smooth muscle relaxation through elevated cGMP. Sildenafil was examined in two guinea pig models of airways disease: guinea pigs exposed to LPS or sensitized guinea pigs with atopy exposed to ovalbumen. Ovalbumen exposure caused early- and late-phase bronchoconstrictor responses, measured in conscious animals by whole-body plethysmography. Twenty-four hours after ovalbumen exposure there was airway hyperreactivity (AHR) to inhaled histamine and significantly elevated macrophages, eosinophils, and nitric oxide (NO) metabolites in bronchoalveolar lavage fluid. Sildenafil treatment (1 mg/kg, intraperitoneally) failed to affect the early and late responses but significantly reduced AHR, leukocyte influx, and elevated NO. LPS exposure (30 microg/ml) caused AHR to histamine at 1 hour and macrophage, eosinophil, and neutrophil influx at 24 hours with raised NO. Sildenafil pretreatment inhibited LPS-induced AHR, leukocyte influx, and NO generation. The effectiveness of sildenafil was not dependent on endogenous NO because inhibition of NO synthase with Nomega-nitro-L-arginine methyl ester did not prevent its action. Inhibition of PDE5 by sildenafil was confirmed by elevated S-nitroso-N-acetylpenicillamine-induced cGMP generation in isolated lungs. These antiinflammatory actions of sildenafil in guinea pig models suggest that PDE5 inhibitors may have potential in treating airways disease.

Topics: Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cyclic GMP; Eosinophils; Guinea Pigs; Histamine; In Vitro Techniques; Lipopolysaccharides; Lung; Macrophages; Male; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Ovalbumin; Phosphodiesterase Inhibitors; Piperazines; Purines; Sildenafil Citrate; Sulfones

Asthma is one of the most common diseases and is characterized by airway obstruction, airway inflammation, and increased airway responsiveness. Glucocorticoids are very effective in treatment, but their long-term use is associated with several side effects, so that new anti-inflammatory drugs are in development. Activated protein C (APC) is a serine protease with potent anti-inflammatory effects. This study evaluated the effect of inhaled APC on airway inflammation and hyperresponsiveness in a murine asthma model. Asthma was induced in BALB/c mice by exposure to chicken egg ovalbumin (OVA), and the effect of inhaled APC was assessed by administering prior to OVA exposure. Inhalation of APC significantly inhibited the expression of T helper 2 (Th2) cytokines, immunoglobulin E (IgE), eosinophilic inflammation, and hyperresponsiveness. APC also significantly suppressed the expression of Th2 cytokines and IgE from lymphocytes isolated from OVA-sensitized/challenged animals. In addition, binding of signal transducer and activator of transcription 6 (STAT6) and nuclear factor kappa B (NF-kappa B) oligonucleotides to lung nuclear proteins was significantly reduced in mice treated with inhaled APC. In brief, the exogenous supplementation of APC inhibits the immunologic and inflammatory responses induced by Th2 cytokines in a mouse model of asthma and may represent a novel anti-inflammatory treatment.

Topics: Animals; Antibodies; Antigens, CD; Asthma; Biomarkers; Blood Coagulation Factors; Bronchial Hyperreactivity; Cytokines; Dinoprostone; Disease Models, Animal; Dose-Response Relationship, Drug; Endothelial Protein C Receptor; Endothelins; Eosinophils; Female; Glycoproteins; HeLa Cells; Humans; Hypersensitivity; Immunoglobulin E; Mice; Mice, Inbred BALB C; NF-kappa B; Nitric Oxide; Ovalbumin; Protein C; Receptor, PAR-1; Receptors, Cell Surface; Specific Pathogen-Free Organisms; STAT6 Transcription Factor; Th2 Cells; Thrombin; Trans-Activators; U937 Cells

Immunostimulatory sequences of DNA (ISS) inhibit eosinophilic airway inflammation, Th2 responses, and airway hyperreactivity (AHR) in mouse models of acute ovalbumin (OVA)-induced airway inflammation. To determine whether ISS inhibits airway remodeling, we developed a mouse model of airway remodeling in which OVA-sensitized mice were repeatedly exposed to intranasal OVA administration for 1-6 mo. Mice chronically exposed to OVA developed sustained eosinophilic airway inflammation and sustained AHR to methacholine compared with control mice. In addition, the mice chronically exposed to OVA developed features of airway remodeling, including thickening of the peribronchial smooth muscle layer, peribronchial myofibroblast accumulation, expression of the profibrotic growth factor transforming growth factor-beta, and subepithelial collagen deposition (assessed by quantitation of the area of peribronchial trichrome staining using image analysis, and immunostaining with anti-collagen V antibodies). Administration of ISS systemically every other week significantly inhibited the development of AHR, eosinophilic inflammation, airway mucus production, and importantly, airway remodeling in mice chronically exposed to OVA for 3-6 mo. In addition, ISS significantly reduced bronchoalveolar lavage and lung levels of the profibrotic cytokine transforming growth factor-beta. These studies demonstrate that ISS prevents not only Th2-mediated airway inflammation in response to acute allergen challenge, but also airway remodeling associated with chronic allergen challenge.

Topics: Animals; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; DNA; Female; Humans; Inflammation; Interferon-gamma; Interleukin-5; Methacholine Chloride; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Respiratory System; Th2 Cells; Transforming Growth Factor beta

Sensitized mice acutely challenged with inhaled ovalbumin (OVA) develop allergic airway inflammation, characterized by OVA-specific IgE production, airway eosinophilia, increased pulmonary B and T lymphocytes, and airway hyperreactivity. In this study, a chronic exposure model was developed and two distinct patterns of response were observed. Discontinuous inhalational exposure to OVA (6 weeks) produced airway inflammation and hyperreactivity that were similar to acute (10 days) responses. Continuous inhalational exposure to OVA (6 or 11 weeks) resulted in attenuation of airway eosinophilia and hyperresponsiveness without reduction of OVA-specific IgE and IgG(1) levels. The inhibition of airway inflammation was dependent on continuous exposure to antigen, because continuously exposed mice with attenuated inflammatory responses redeveloped allergic airway disease if the OVA aerosols were interrupted and then restarted (11-week-discontinuous). Inhalational tolerance induced by continuous OVA exposure demonstrated bystander suppression of cockroach allergen-mediated airway eosinophilia. These findings may be attributed to changes in production of the anti-inflammatory cytokine interleukin-10 during continuous OVA aerosol exposure. The symptomatic and asymptomatic allergic responses in human asthmatics could be explained by similar variable or discontinuous exposures to aeroantigens.

Topics: Administration, Inhalation; Animals; Antigens; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Immune Tolerance; Immunoglobulin E; Immunoglobulin G; Male; Mice; Ovalbumin; T-Lymphocytes; Time Factors

Brain-derived neurotrophic factor (BDNF) is a candidate molecule for mediating functional neuronal changes in allergic bronchial asthma. Recently, enhanced production of BDNF during allergic airway inflammation caused by infiltrating T-cells and macrophages as well as by resident airway epithelial cells has been described. It was the aim of this study to investigate the effect of enhanced BDNF levels on lung function and airway inflammation in a mouse model of allergic inflammation. Ovalbumin-sensitised BALB/c mice were challenged in two consecutive allergen challenges. Prior to the challenge, the mice were treated with either anti-BDNF antibodies or isotype-matched control antibodies. Airway responsiveness to methacholine, capsaicin and electric field stimulation, as well as airway inflammation and chronic airway obstruction 1 week after the last allergen challenge were assessed. Anti-BDNF blocked enhanced reactivity in response to capsaicin, but not airway smooth muscle hyper-reactivity in vivo. Furthermore, persistent airway obstruction, as observed 1 week after the last allergen challenge, was to a large extent prevented by anti-BDNF treatment. In vitro, BDNF and anti-BDNF treatment had a profound effect on local neuronal hyper-reactivity, as shown by electric field stimulation experiments. In contrast, neither BDNF nor anti-BDNF treatment affected airway inflammation. Our data indicate that development of allergen-induced neuronal hyper-reactivity in mice is partially mediated by BDNF. British Journal of Pharmacology (2004) 141, 431-440. doi:10.1038/sj.bjp.0705638

Topics: Animals; Asthma; Brain-Derived Neurotrophic Factor; Bronchial Hyperreactivity; Female; Mice; Mice, Inbred BALB C; Neurons; Ovalbumin; Respiratory Function Tests

Little is known about effects of smoking on airway inflammation in asthma. We tested the hypothesis that smoking enhances established airway inflammation in a mouse model of allergic asthma. C57Bl/6j mice were sensitized to ovalbumin (OVA) and challenged with OVA (OVA-mice) or sham-sensitized to phosphate-buffered saline (PBS) and challenged with PBS aerosols (PBS-mice) for 7 wk. At 4 wk, mice were additionally exposed to air (nonsmoking controls) or mainstream smoke for 3 wk. Using whole body plethysmography, we found OVA-induced bronchoconstriction to be significantly inhibited in smoking OVA-mice as compared with nonsmoking OVA-mice (1 +/- 2% increase versus 22 +/- 6% increase in enhanced pause, respectively). Smoking did not change airway hyperresponsiveness (AHR) to methacholine in PBS-mice, yet significantly attenuated AHR in OVA-mice 24 h after OVA challenge as compared with nonsmoking mice. This was accompanied by reduced eosinophil numbers in lung lavage fluid and tissue of smoking OVA-mice compared with nonsmoking OVA-mice. In contrast to our hypothesis, short-term smoking reduced responsiveness to OVA and methacholine in OVA-mice and decreased airway inflammation when compared with nonsmoking mice. This effect of smoking may be different for long-term smoking, in which remodeling effects of smoking can be expected to interrelate with remodeling changes caused by asthmatic disease.

Topics: Administration, Inhalation; Air Pollutants; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cytokines; Disease Models, Animal; Inflammation; Lung; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Plethysmography, Whole Body; Smoke; T-Lymphocytes; Time Factors

The mechanism(s) of bradykinin-induced bronchoconstriction was investigated in the Brown Norway (BN) rat model of allergic asthma. Bronchoconstrictor responses to i.v. bradykinin in BN rats were maximally augmented 24 h following challenge with allergen and declined at later time points. Histological evaluation of the inflammatory status of the lungs after ovalbumin (OA) challenge showed a marked inflammatory response, which was maximal at 24 h and declined thereafter. However, pretreatment with budesonide did not inhibit the augmented bronchoconstrictor response to bradykinin 24 h after allergen challenge. The selective B1 receptor agonist, Lys-[desArg9]-BK had no bronchoconstrictor effects, whereas the selective B2 receptor antagonist, HOE 140, abolished the response to bradykinin in OA-challenged animals. The augmented response to bradykinin was not affected by methysergide, indomethacin, disodium cromoglycate, iralukast, the 5-lipoxygenase inhibitor, CGS8515, or the NK2 receptor antagonist, SR48968. It was, however, partially inhibited by atropine both in saline- and OA-challenged animals. Pretreatment with captopril and thiorphan markedly potentiated responses to bradykinin both in saline- and OA-challenged animals. Thus, augmentation of the bronchoconstrictor response to bradykinin occurs in actively sensitised BN rats 24 h after challenge with OA and is associated with marked pulmonary inflammation. The response is entirely B2 receptor mediated and approximately 50% of the response is cholinergic. However, mast cell activation, the products of the cyclooxygenase or 5-lipoxygenase pathways and tachykinins are not involved. Peptidase inhibition mimics the effect of allergen challenge on the bronchoconstrictor response to bradykinin and it remains possible that the mechanism of the augmented response to bradykinin following allergen challenge involves downregulation of peptidase activity as a consequence of the inflammatory response.

Topics: Adenosine; Adrenergic beta-1 Receptor Agonists; Adrenergic beta-2 Receptor Antagonists; Animals; Arachidonate 5-Lipoxygenase; Bradykinin; Bronchial Hyperreactivity; Bronchoconstriction; Bronchoconstrictor Agents; Disease Models, Animal; Endopeptidases; Lung; Male; Mast Cells; Methacholine Chloride; Ovalbumin; Rats; Rats, Inbred BN; Receptors, Muscarinic; Tachykinins

Cytokines play an integral role in the coordination and persistence of allergic inflammatory processes and therefore represent prime targets for novel therapies in diseases such as asthma. Multiple attempts to generate low-molecular-weight cytokine inhibitors have failed, and the main attention has focused on the development of biological agents such as neutralizing antibodies. The present work describes a simple and effective method to induce the production of therapeutic anti-cytokine autoantibodies by active immunization against a modified endogenous cytokine.. Balb/c mice were subcutaneously injected with AutoVac TNF106, a recombinant murine TNF-alpha molecule containing a foreign immunogenic T helper epitope, and the induction of neutralizing anti-TNF-alpha autoantibodies was analysed. These mice were then sensitized with ovalbumin (OVA), and the effect of neutralizing anti-TNF-alpha autoantibodies on the allergen-induced airway inflammation was analysed.. AutoVac TNF106-immunized mice developed high titres of neutralizing anti-TNF-alpha autoantibodies, which were maintained for at least 4 weeks after the last booster injection. Mice vaccinated with AutoVac TNF106 and further immunized against OVA showed diminished TNF-alpha levels in the bronchoalveolar lavage (BAL) fluid after OVA challenge. Moreover, pretreatment with AutoVac TNF106 resulted in significantly reduced numbers of eosinophils and neutrophils in BAL fluid in response to single or multiple allergen exposure.. The induction of anti-TNF-alpha autoantibody production by the AutoVac TNF106 technology not only confirmed the role of TNF-alpha in the induction of allergic inflammation but also offers a novel approach to block the activity of cytokines in order to treat allergic inflammatory conditions.

Topics: Animals; Autoantibodies; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cell Count; Cytokines; Enzyme-Linked Immunosorbent Assay; Female; Immunization; Immunoglobulin E; Immunotherapy; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Pulmonary Eosinophilia; Recombinant Proteins; Tumor Necrosis Factor-alpha

B7-H1 (PD-L1) and B7-DC (PD-L2) are the ligands for programmed death-1 (PD-1), which is a member of the CD28/CTLA-4 family and has been implicated in peripheral tolerance. We investigated the roles of B7-H1 and B7-DC in a murine OVA-induced allergic asthma model. B7-H1 was constitutively expressed on dendritic cells, macrophages, B cells, and T cells in the lungs of naive mice, and its expression could be dramatically increased after allergen challenge. In contrast, B7-DC expression was scarcely expressed on dendritic cells in naive mice, but was up-regulated after allergen challenge, although the up-regulation of B7-DC expression on macrophages was minimal. Treatment of mice with anti-B7-DC mAb at the time of allergen challenge, but not at the time of sensitization, significantly increased their airway hyper-reactivity and eosinophilia. Such treatment also resulted in the increased production of IL-5 and IL-13, and decreased IFN-gamma production in the lungs and draining lymph node cells. These changes were diminished when mice were depleted of IFN-gamma by anti-IFN-gamma mAb pretreatment. Interestingly, treatment with anti-B7-H1 or anti-PD-1 mAb did not significantly affect the asthmatic response. These results suggest a unique role for B7-DC in the regulation of asthmatic response through an IFN-gamma-dependent, but PD-1-independent, mechanism.

Topics: Allergens; Animals; Antibodies, Monoclonal; Antigens, Differentiation; Asthma; B7-1 Antigen; B7-H1 Antigen; Blood Proteins; Bronchial Hyperreactivity; Dendritic Cells; Disease Models, Animal; Interferon-gamma; Interleukin-13; Interleukin-5; Leukocytes; Lung; Lymph Nodes; Male; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Ovalbumin; Peptides; Programmed Cell Death 1 Ligand 2 Protein; Programmed Cell Death 1 Receptor; T-Lymphocyte Subsets; Up-Regulation

The role of Th2/CD4 T cells, which secrete IL-4, IL-5, and IL-13, in allergic disease is well established; however, the role of CD8(+) T cells (allergen-induced airway hyperresponsiveness (AHR) and inflammation) is less clear. This study was conducted to define the role of Ag-primed CD8(+) T cells in the development of these allergen-induced responses. CD8-deficient (CD8(-/-)) mice and wild-type mice were sensitized to OVA by i.p. injection and then challenged with OVA via the airways. Compared with wild-type mice, CD8(-/-) mice developed significantly lower airway responsiveness to inhaled methacholine and lung eosinophilia, and exhibited decreased IL-13 production both in vivo, in the bronchoalveolar lavage (BAL) fluid, and in vitro, following Ag stimulation of peribronchial lymph node (PBLN) cells in culture. Reconstitution of sensitized and challenged CD8(-/-) mice with allergen-sensitized CD8(+) T cells fully restored the development of AHR, BAL eosinophilia, and IL-13 levels in BAL and in culture supernatants from PBLN cells. In contrast, transfer of naive CD8(+) T cells or allergen-sensitized CD8(+) T cells from IL-13-deficient donor mice failed to do so. Intracellular cytokine staining of lung as well as PBLN T cells revealed that CD8(+) T cells were a source of IL-13. These data suggest that Ag-primed CD8(+) T cells are required for the full development of AHR and airway inflammation, which appears to be associated with IL-13 production from these primed T cells.

Topics: Administration, Inhalation; Adoptive Transfer; Animals; Antigens; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD8 Antigens; CD8-Positive T-Lymphocytes; Cell Division; Cells, Cultured; Cytokines; Female; Immunoglobulin E; Inflammation; Injections, Intraperitoneal; Interleukin-13; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin

Matrix metalloproteinases (MMPs) are a large family of endopeptidases that proteolytically degrade extracellular matrix. Many different cells produce MMP-9, and levels have been shown to be up-regulated in patients with allergic asthma. The aim of this study was to investigate the in vivo role of MMP-9 during allergen-induced airway inflammation. Acute allergic pulmonary eosinophilia was established in MMP-9 knockout (KO) and wild-type (WT) control mice by sensitization and challenge with OVA. Cell recruitment was significantly increased in both bronchoalveolar lavage (BAL) and lung tissue compartments in MMP-9 KO mice compared with WT mice. This heightened cell recruitment was primarily due to increased eosinophils and Th2 cells in the BAL and lung tissue of MMP-9 KO mice in comparison with WT controls. Moreover, levels of the Th2 cytokines, IL-4 and IL-13, and the chemokines eotaxin/CCL11 and macrophage-derived chemokine/CCL22 were substantially increased in MMP-9 KO mice compared with WT after OVA challenge. Resolution of eosinophilia was similar between MMP-9 KO and WT mice, but Th2 cells persisted in BAL and lungs of MMP-9 KO mice for longer than in WT mice. Our results indicate that MMP-9 is critically involved in the recruitment of eosinophils and Th2 cells to the lung following allergen challenge, and suggest that MMP-9 plays a role in the development of Th2 responses to allergen.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cell Movement; Cytokines; Eosinophils; Immunoglobulin E; Inflammation; Injections, Intraperitoneal; Lung; Matrix Metalloproteinase 9; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Th2 Cells

The NOD mouse has proved to be a relevant model of insulin-dependent diabetes mellitus, closely resembling the human disease. However, it is unknown whether this strain presents a general biastoward Th1-mediated autoimmunity or remains capable of mounting complete Th2-mediated responses. Here, we show that NOD mice have the capacity to develop a typical Th2-mediated disease, namely experimental allergic asthma. In contrast to what might have been expected, they even developed a stronger Th2-mediated pulmonary inflammatory response than BALB/c mice, a strain that shows a typical Th2 bias in this model. Thus, after allergen sensitization and intra-nasal challenge, the typical features of experimental asthma were exacerbated in NOD mice, including enhanced bronchopulmonary responsiveness, mucus production and eosinophilic inflammation in the lungs as well as specific IgE titers in serum. These hallmarks of allergic asthma were associated with increased IL-4, IL-5, IL-13 and eotaxin production in the lungs, as compared with BALB/c mice. Notwithstanding their quantitative and functional defect in NOD mice, CD1d-dependent NKT cells contribute to aggravate the disease, since in OVA-immunized CD1d(-/-) NOD mice, which are deficient in this particular T cell subset, airway eosinophilia was clearly diminished relative to NOD littermates. This is the first evidence that autoimmune diabetes-prone NOD mice can also give rise to enhanced Th2-mediated responses and might thus provide a useful model for the study of common genetic and cellular components, including NKT cells that contribute to both asthma and type 1 diabetes.

Topics: Animals; Antibodies, Monoclonal; Antigens, CD1; Antigens, CD1d; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokines, CC; Cytokines; Diabetes Mellitus, Type 1; Immunoglobulin E; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Mice, Inbred NOD; Mice, Knockout; Mucus; Ovalbumin; Pulmonary Eosinophilia; Th2 Cells

Increasing evidence suggests that alveolar macrophages (AM) are involved in asthma pathogenesis. To better understand the role that these cells play, we investigated the capacity of AM from allergy-resistant rat, Sprague Dawley (SD), to modulate airway hyperresponsiveness of allergy-susceptible rat, Brown Norway (BN). AM of ovalbumin (OVA)-sensitized BN rats were eliminated by intratracheal instillation of liposomes containing clodronate. AM from OVA-sensitized SD rats were transferred into AM-depleted BN rats 24 h before allergen challenge. Airway responsiveness to methacholine was measured the following day. Instillation of liposomes containing clodronate in BN rats eliminated 85% AM after 3 d compared with saline liposomes. Methacholine concentration needed to increase lung resistance by 200% (EC200RL) was significantly lower in OVA-challenged BN rats (27.9 +/- 2.8 mg/ml) compared with SD rats (63.9 +/- 8.6 mg/ml). However, when AM from SD rats were transferred into AM-depleted BN rats, airway responsiveness (64.0 +/- 11.3 mg/ml) was reduced to the level of naïve rats (54.4 +/- 3.7 mg/ml) in a dose-dependent manner. Interestingly, transfer of AM from BN rats into SD rats did not modulate airway responsiveness. To our knowledge, this is the first direct evidence showing that AM may protect against the development of airway hyperresponsiveness.

Topics: Adoptive Transfer; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Clodronic Acid; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Resistance; Genetic Predisposition to Disease; Immunoglobulin E; Immunoglobulin G; Liposomes; Macrophages, Alveolar; Male; Methacholine Chloride; Ovalbumin; Rats; Rats, Sprague-Dawley; Reaction Time

During the preclinical study of new therapeutic modality, we evaluate whether the treatment can reverse the established asthma phenotypes in animal model. However, few have reported on the long term persistence of asthma phenotypes upon re-challenge with allergen (secondary challenge) in animal model. We evaluated the persistence of asthma phenotypes by secondary challenge at different times in previously challenged murine asthma model. BALB/c mice sensitized by intraperitoneal injections of 20 micro g of ovalbumin and 1 mg of alum on days 1 and 14 were challenged initially by the inhalation of 1% ovalbumin for 30 min on days 21, 22, and 23. Each group of mice was rechallenged at 5, 7, 9, or 12 weeks after the initial challenge. Airway hyperresponsiveness, BAL fluid, airway histology and serum ovalbumin-specific IgE level were evaluated. Airway eosinophilia, airway inflammation and serum ovalbumin-specific IgE production persisted upon secondary allergen challenges at least 12 weeks after the initial challenge. However, airway hyperresponsiveness persisted only until mice were rechallenged 7 weeks after the initial challenge. Airway inflammation and allergen specific IgE production may persist longer than airway hyperresponsiveness in a mouse asthma model of secondary allergen challenge.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Bronchoalveolar Lavage Fluid; Female; Immunoglobulin E; Inflammation; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Phenotype; Respiratory Hypersensitivity; Respiratory System; Time Factors

Interleukin (IL)-18 was originally regarded to induce T helper cell (Th)1-related cytokines. In general, factors favoring interferon (IFN)-gamma production are believed to abolish allergic diseases. Thus, we tested the role of IL-18 in regulation of bronchial asthma. To avoid a background response of host-derived T cells, we administered memory type Th1 or Th2 cells into unsensitized mice and examined their role in induction of bronchial asthma. Administration of antigen (Ag) induced both airway inflammation and airway hyperresponsiveness (AHR) in mice receiving memory Th2 cells. In contrast, the same treatment induced only airway inflammation but not AHR in mice receiving memory Th1 cells. However, these mice developed striking AHR when they were coadministered with IL-18. Furthermore, mice having received IFN-gamma-expressing Th1 cells sorted from polarized Th1 cells developed severe airway inflammation and AHR after intranasal administration of Ag and IL-18. Thus, Th1 cells become harmful when they are stimulated with Ag and IL-18. Newly polarized Th1 cells and IFN-gamma-expressing Th1 cells, both of which express IL-18 receptor alpha chain strongly, produce IFN-gamma, IL-9, IL-13, granulocyte/macrophage colony-stimulating factor, tumor necrosis factor alpha, regulated on activation, normal T cell expressed and secreted, and macrophage inflammatory protein 1alpha upon stimulation with Ag, IL-2, and IL-18 in vitro. Thus, Ag and IL-18 stimulate memory Th1 cells to induce severe airway inflammation and AHR in the naive host.

Topics: Animals; Bronchial Hyperreactivity; Clone Cells; Disease Models, Animal; Female; Immunologic Memory; Inflammation; Interleukin-18; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes, Helper-Inducer; Th1 Cells; Th2 Cells

Allergic airway inflammation and hyperreactivity are modulated by gammadelta T cells, but different experimental parameters can influence the effects observed. For example, in sensitized C57BL/6 and BALB/c mice, transient depletion of all TCR-delta(+) cells just before airway challenge resulted in airway hyperresponsiveness (AHR), but caused hyporesponsiveness when initiated before i.p. sensitization. Vgamma4(+) gammadelta T cells strongly suppressed AHR; their depletion relieved suppression when initiated before challenge, but not before sensitization, and they suppressed AHR when transferred before challenge into sensitized TCR-Vgamma4(-/-)/6(-/-) mice. In contrast, Vgamma1(+) gammadelta T cells enhanced AHR and airway inflammation. In normal mice (C57BL/6 and BALB/c), enhancement of AHR was abrogated only when these cells were depleted before sensitization, but not before challenge, and with regard to airway inflammation, this effect was limited to C57BL/6 mice. However, Vgamma1(+) gammadelta T cells enhanced AHR when transferred before challenge into sensitized B6.TCR-delta(-/-) mice. In this study Vgamma1(+) cells also increased levels of Th2 cytokines in the airways and, to a lesser extent, lung eosinophil numbers. Thus, Vgamma4(+) cells suppress AHR, and Vgamma1(+) cells enhance AHR and airway inflammation under defined experimental conditions. These findings show how gammadelta T cells can be both inhibitors and enhancers of AHR and airway inflammation, and they provide further support for the hypothesis that TCR expression and function cosegregate in gammadelta T cells.

Topics: Adoptive Transfer; Animals; Antigens; Bronchial Hyperreactivity; Cytokines; Female; Inflammation; Lung; Lymphocyte Depletion; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocyte Subsets; Th2 Cells

Chronic inflammation in asthmatic airways can lead to characteristic airway smooth muscle (ASM) thickening and pathological changes within the airway wall.. We investigated the long-term effects of repeated allergen exposure.. Brown-Norway (BN) rats sensitized to ovalbumin (OVA) were exposed to OVA or saline aerosol every third day on six occasions and studied 24 h, 7 days and 35 days after the final exposure. We measured airway inflammation, ASM cell proliferation (by incorporation of bromodeoxyuridine; BrdU) and bronchial responsiveness to acetylcholine.. At 24 h, in OVA-exposed rats, we detected elevated OVA-specific serum IgE, increased numbers of macrophages, eosinophils, lymphocytes and neutrophils in the bronchoalveolar lavage (BAL) fluid and increased numbers of MBP+ (major basic protein) eosinophils and CD2+ T cells within the bronchial submucosa. This coincided with increased numbers of ASM cells expressing BrdU and with bronchial hyper-responsiveness (BHR). At 7 days, BHR was detected in OVA-exposed rats, coincident with increased numbers of macrophages and lymphocytes in BAL fluid together with increased numbers of CD2+ T cells within the bronchial submucosa. This coincided with increased numbers of ASM cells expressing BrdU. By day 35, the number of ASM cells expressing BrdU remained elevated in the absence of cellular infiltration and BHR.. Repeated OVA-challenge results in persistent ASM cell proliferation in the absence of bronchial inflammation and BHR, which lasts for at least 1 week following cessation of exposure.

Topics: Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD2 Antigens; Cell Division; Eosinophils; Immunoglobulin E; Leukocytes; Male; Muscle, Smooth; Ovalbumin; Rats; Rats, Inbred BN; T-Lymphocytes; Time Factors

Heme oxygenase (HO), the heme-degrading enzyme, has shown anti-inflammatory effects in several models of pulmonary diseases. HO is induced in airways during asthma; however, its functional role is unclear. Therefore, we evaluated the role of HO on airway inflammation [evaluated by bronchoalveolar lavage (BAL) cellularity and BAL levels of eotaxin, PGE(2), and proteins], mucus secretion (evaluated by analysis of MUC5AC gene expression and periodic acid-Schiff staining), oxidative stress (evaluated by quantification of 4-hydroxynonenal adducts and carbonylated protein levels in lung homogenates), and airway responsiveness to histamine in ovalbumin (OVA)-sensitized and multiple aerosol OVA or saline-challenged guinea pigs (6 challenges, once daily, OVA group and control group, respectively). Airway inflammation, mucus secretion, oxidative stress, and responsiveness were significantly increased in the OVA group compared with the control group. HO upregulation by repeated administrations of hemin (50 mg/kg i.p.) significantly decreased airway responsiveness in control animals and airway inflammation, mucus secretion, oxidative stress, and responsiveness in OVA animals. These effects were reversed by the concomitant administration of the HO inhibitor tin protoporphyrin-IX (50 micromol/kg i.p.). Repeated administrations of tin protoporphyrin-IX alone significantly increased airway responsiveness in control animals but did not modify airway inflammation, mucus secretion, oxidative stress, and responsiveness in OVA animals. These results suggest that upregulation of the HO pathway has a significant protective effect against airway inflammation, mucus hypersecretion, oxidative stress, and hyperresponsiveness in a model of allergic asthma in guinea pigs.

Topics: Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoconstriction; Enzyme Inhibitors; Guinea Pigs; Heme Oxygenase (Decyclizing); Hemin; Histamine; Metalloporphyrins; Ovalbumin; Oxidative Stress; Protoporphyrins; Up-Regulation

MX-68 is a newly synthesized antifolate, which is a derivative of methotrexate (MTX). In this paper, the effect of MX-68 on allergic airway responses in mice and guinea-pigs was studied. In the first experiment, antigen-induced airway inflammation and airway hyperresponsiveness (AHR) to acetylcholine in mice were examined and compared with the effects of classical antifolate methotrexate and prednisolone. Mice were sensitized with ovalbumin as an antigen and challenged with ovalbumin inhalation three times. After the last inhalation, AHR and airway inflammation were observed. An increase in Th2 cytokines (IL-4 and IL-5) and a decrease in a Th1 cytokine (IFN-gamma) in the bronchoalveolar lavage fluid (BALF), as well as an elevation of the immunoglobulin level in serum, were observed in sensitized mice. Oral administration of MX-68 had no effect on changes of body weight, but prednisolone reduced body weight during the experiment. The antigen-induced AHR and increases in the number of eosinophils and lymphocytes in BALF were significantly inhibited by MX-68. MX-68 interfered with the elevation of IL-4 and IL-5 levels in BALF, but had no effect on the decrease in IFN-gamma. Moreover, MX-68 significantly inhibited the elevation of serum IgE and IgG levels. In the guinea-pig model for bronchial asthma, biphasic increases in airway resistance (immediate asthmatic response, IAR, and late asthmatic response, LAR), as well as accumulated inflammatory cells in BALF, were observed after repeated antigen challenge. These asthmatic responses and inflammatory signs were significantly decreased by administration of MX-68. These results suggest that MX-68 obviously has an anti-inflammatory effect in an animal model of asthma and would be useful clinically for the treatment of bronchial asthma.

Topics: 2-Aminoadipic Acid; Acetylcholine; Administration, Inhalation; Allergens; Animals; Body Weight; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Dogs; Dose-Response Relationship, Drug; Guinea Pigs; Immunoglobulins; Injections, Intraperitoneal; Interferon-gamma; Interleukin-4; Interleukin-5; Male; Methotrexate; Mice; Mice, Inbred BALB C; Ovalbumin; Prednisolone

Bronchoconstrictor responses were measured in lungs isolated from spontaneously hypertensive (SHR) and normotensive rats, perfused via the airways. Lungs from SHRs were more responsive than lungs from normotensive rats to methacholine, 5-hydroxytryptamine (5-HT), arachidonic acid or prostaglandin H(2). The responses of SHR airways to methacholine or 5-HT were unaffected by pretreatment in vivo with an inhibitor of nitric oxide (NO) synthase, N(omega)-nitro-L-arginine methyl ester hydrochloride (L-NAME, 30 mg kg(-1)), although responses in normotensive airways to methacholine, but not to 5-HT, were enhanced. Antigen challenge of isolated lungs from actively sensitized rats elicited bronchoconstriction, not different between strains. Pretreatment with L-NAME increased the response to antigen challenge only in normotensive lungs. Compound 48/80 induced bronchoconstriction in lungs from either strain, equally. These responses to compound 48/80 were unaffected by L-NAME pretreatment. Thus, SHR airways lack relaxing factors and degranulation of mast cells in SHR lungs was not affected by endogenous NO.

Topics: Animals; Bronchial Hyperreactivity; Bronchoconstriction; Bronchoconstrictor Agents; Cell Degranulation; Dose-Response Relationship, Drug; Enzyme Inhibitors; Hypersensitivity; In Vitro Techniques; Lung; Male; Mast Cells; Methacholine Chloride; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Ovalbumin; p-Methoxy-N-methylphenethylamine; Rats; Rats, Inbred SHR; Rats, Wistar

Aerosolized monoclonal antibodies (mAbs) specific for T-cell receptors (TCR) were used to manipulate T-cell function in airways of ovalbumin (OVA)-sensitized and -challenged mice with airway hyperresponsiveness (AHR). The inhaled mAbs were found to be effective at low doses, had little or no systemic effect and specifically abrogated both effector and regulatory functions of the targeted T cells. Specific mAbs targeting alphabeta T cells suppressed and those targeting gammadelta T cells enhanced AHR. Moreover, specific mAbs directed against subsets of gammadelta T cells varied in their effect on AHR. Using this approach of targeting either alphabeta or gammadelta T cells reduced airway eosinophila, although the effect of mAbs specific for alphabeta T cells was stronger. The use of aerosolized anti-TCR mAbs may offer an effective approach for the treatment of airway inflammation and AHR.

Topics: Administration, Inhalation; Airway Resistance; Animals; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal; Antibody Specificity; Bronchial Hyperreactivity; Bronchitis; Disease Models, Animal; Dose-Response Relationship, Drug; Flow Cytometry; Lung; Lymphocyte Count; Lymphocyte Depletion; Methacholine Chloride; Mice; Mice, Inbred C57BL; Ovalbumin; Receptors, Antigen, T-Cell; Receptors, Antigen, T-Cell, gamma-delta; Respiratory Hypersensitivity; T-Lymphocyte Subsets

Flt3 ligand (Flt3-L) is a growth factor for dendritic cells and induces type 1 T cell responses. We recently reported that Flt3-L prevented OVA-induced allergic airway inflammation and suppressed late allergic response and airway hyper-responsiveness (AHR). In the present study we examined whether Flt3-L reversed allergic airway inflammation in an established model of asthma. BALB/c mice were sensitized and challenged with OVA, and AHR to methacholine was established. Then mice with AHR were randomized and treated with PBS or 6 microg of Flt3-L i.p. for 10 days. Pulmonary functions and AHR to methacholine were examined after rechallenge with OVA. Treatment with Flt3-L of presensitized mice significantly suppressed (p < 0.001) the late allergic response, AHR, bronchoalveolar lavage fluid total cellularity, absolute eosinophil counts, and inflammation in the lung tissue. There was a significant decrease in proinflammatory cytokines (TNF-alpha, IL-4, and IL-5) in bronchoalveolar lavage fluid, with a significant increase in serum IL-12 and a decrease in serum IL-5 levels. There was no significant effect of Flt3-L treatment on serum IL-4 and serum total IgE levels. Sensitization with OVA significantly increased CD11b(+)CD11c(+) cells in the lung, and this phenomenon was not significantly affected by Flt3-L treatment. These data suggest that Flt3-L can reverse allergic airway inflammation and associated changes in pulmonary functions in murine asthma model.

Topics: Administration, Inhalation; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Dendritic Cells; Disease Models, Animal; Female; Hypersensitivity, Delayed; Immunization, Secondary; Immunoglobulin E; Immunoglobulin G; Inflammation; Injections, Intraperitoneal; Leukocyte Count; Ligands; Lung; Membrane Proteins; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Hypersensitivity

Despite the controversy of bronchial responsiveness to beta2-agonist drugs in asthma, in a previous study we have shown increased responsiveness of asthmatic tracheobronchial tree to isoprenaline. Therefore, in the present study, tracheal responsiveness to isoprenaline and also beta-adrenergic receptor blockade were studied in sensitized guinea pigs. An experimental model of asthma was induced in guinea pigs by sensitization of animals with injection and inhalation of ovalbumin (OA). The responses of tracheal chains of sensitized and control animals to cumulative concentrations of isoprenaline (I) in the absence and presence of 10 nmol/l propranolol were measured, and the effective concentration of I causing 50% of maximum response (EC50 I) was obtained. The propranolol blockade (CR - 1) was calculated by: (post-propranolol EC50 I/EC50 I) - 1. Tracheal responses of sensitized and control animals to cumulative concentrations of methacholine (M) were also measured and EC50 M were obtained. The tracheal responses of sensitized guinea pig to isoprenaline was significantly higher than that of the control animals (EC50 I for sensitized and control animals were 0.24 +/- 0.04 and 0.58 +/- 0.07 micromol/l, respectively; p < 0.001). The beta-adrenergic receptor blockade by propranolol (CR - 1) was also significantly higher in sensitized guinea pigs than that of the control animals (p < 0.001). The results of this study indicate an increased tracheal response to beta-adrenergic-stimulating drug and enhancement of beta-adrenergic blockade by propranolol in the sensitized guinea pig.

Topics: Administration, Inhalation; Adrenergic beta-Agonists; Adrenergic beta-Antagonists; Aerosols; Animals; Bronchial Hyperreactivity; Disease Models, Animal; Guinea Pigs; Immunization; Injections, Intraperitoneal; Injections, Subcutaneous; Isoproterenol; Male; Methacholine Chloride; Ovalbumin; Propranolol; Trachea

Allergen exposure of sensitised lungs produces bronchial hyperresponsiveness in vivo associated with airway inflammation and remodelling. It is unclear if hyperresponsiveness is also present in airways in vitro under similar conditions of drug provocation as carried out in vivo, and at different times after allergen-challenge. This study records responsiveness of individual airway segments to acetylcholine (ACh) in sensitised bronchi after instillation of allergen (ovalbumin, OA). Airway histology and sensitivity and maximum effects to ACh were recorded 1, 24 and 72 h and 1 week after OA. OA-instilled airways exhibited eosinophilia and epithelial proliferation. Physiological recordings showed no change in maximum contractions of airway segments to acetylcholine placed in the airway lumen except at 24 h where they were reduced. In contrast maximum contractions to ACh to the airway adventitia were reduced at all times except 1 week, with the greatest change occurring at 24 h. There were no changes in airway sensitivity to either route of ACh in OA-instilled airways but the difference in sensitivity to adventitial and lumenal ACh was reduced. Results show that allergen does not produce hyperresponsiveness at the airway wall but it may alter an interaction between airway smooth muscle and other structural components of the airway.

Topics: Acetylcholine; Allergens; Animals; Bronchi; Bronchial Hyperreactivity; Cell Division; Eosinophils; Female; In Vitro Techniques; Instillation, Drug; Muscle, Smooth; Ovalbumin; Respiratory Mucosa; Swine

In certain models of allergic airway disease, mast cells facilitate the development of inflammation and airway hyper-responsiveness (AHR). To define the role of the high affinity IgE receptor (FcepsilonRI) in the development of AHR, mice with a disruption of the alpha subunit of the high affinity IgE receptor (FcepsilonRI(-/-)) were exposed on 10 consecutive days to nebulized OVA. Forty-eight hours after the last nebulization, airway responsiveness was monitored by the contractile response of tracheal smooth muscle to electrical field stimulation (EFS). After the 10-day OVA challenge protocol, wild-type mice demonstrated increased responsiveness to EFS, whereas similarly challenged FcepsilonRI(-/-) mice showed a low response to EFS, similar to nonexposed animals. Further, allergen-challenged FcepsilonRI(-/-) mice showed less airway inflammation, goblet cell hyperplasia, and lower levels of IL-13 in lung homogenates compared with the controls. IL-13-deficient mice failed to develop an increased response to EFS or goblet cell hyperplasia after the 10-day OVA challenge. We transferred bone marrow-derived mast cells from wild-type mice to FcepsilonRI(-/-) mice 1 day before initiating the challenge protocol. After the 10-day OVA challenge, recipient FcepsilonRI(-/-) mice demonstrated EFS-induced responses similar to those of challenged wild-type mice. Transferred mast cells could be detected in tracheal preparations. These results show that FcepsilonRI is important for the development of AHR after an aerosolized allergen sensitization protocol and that this effect is mediated through FcepsilonRI on mast cells and production of IL-13 in the lung.

Topics: Adjuvants, Immunologic; Adoptive Transfer; Aerosols; Allergens; Animals; Bone Marrow Transplantation; Bronchial Hyperreactivity; Electric Stimulation; Female; Goblet Cells; Hyperplasia; Immunoglobulin E; Immunoglobulin G; Inflammation; Interleukin-13; Leukopenia; Lung; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Mutant Strains; Mice, Transgenic; Nebulizers and Vaporizers; Ovalbumin; Receptors, IgE; Trachea

Toluene diisocyanate (TDI) is a leading cause of occupational asthma. Although considerable controversy remains regarding its pathogenesis, TDI-induced asthma is an inflammatory disease of the airways characterized by airway remodeling caused, at least in part, by an excess of extracellular matrix deposition in the airway wall. Matrix metalloproteinases (MMPs) are major proteolytic enzymes that are involved in extracellular matrix turnover because of their ability to cleave all proteins constituting extracellular matrix. Previous studies have reported that MMP-9 might play a role in chronic airway inflammation and remodeling in asthma.. An aim of the current study was to evaluate the effects of MMP-inhibiting antibiotic, doxycycline, and MMP inhibitors on hyperresponsiveness and inflammation of the airways in TDI-induced asthma.. We used a murine model for TDI-induced asthma to examine the effect of doxycycline or MMP inhibitors on bronchial inflammation and airway hyperresponsiveness.. The following typical pathophysiologic features are observed in the lungs of the mice: airway inflammation, airway hyperresponsiveness, and increased expression of MMP-9 mRNA and protein. Administration of doxycycline and MMP inhibitors reduced all of these pathophysiologic findings. In addition, the increased phosphorylated Akt but not Akt protein levels in lung tissues after TDI inhalation were significantly reduced by the administration of doxycycline and MMP inhibitors.. These findings suggest that doxycycline may reduce airway inflammation and hyperresponsiveness through phosphatidylinositol 3-kinase pathway in a murine model of TDI-induced asthma.

Topics: Animals; Anti-Bacterial Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Doxycycline; Female; Inflammation; Lung; Matrix Metalloproteinase 9; Matrix Metalloproteinase Inhibitors; Mice; Mice, Inbred BALB C; Ovalbumin; Protease Inhibitors; RNA, Messenger; Toluene 2,4-Diisocyanate

Mitogen-activated protein kinase (MAPK) signaling cascade plays a pivotal role in the activation of inflammatory cells. Recent findings revealed that the activity of p42/44 MAPK (also known as extracellular signal-regulated kinase (ERK)) in the lungs was significantly higher in asthmatic mice than in normal controls. We hypothesized that inhibition of ERK activity may have anti-inflammatory effects in allergic asthma. BALB/c mice were sensitized with OVA and, upon OVA aerosol challenge, developed airway eosinophilia, mucus hypersecretion, elevation in cytokine and chemokine levels, up-regulation of VCAM-1 expression, and airway hyperresponsiveness. Intraperitoneal administration of U0126, a specific MAPK/ERK kinase inhibitor, significantly (p < 0.05) inhibited OVA-induced increases in total cell counts, eosinophil counts, and IL-4, IL-5, IL-13, and eotaxin levels recovered in bronchoalveolar lavage fluid in a dose-dependent manner. U0126 also substantially (p < 0.05) reduced the serum levels of total IgE and OVA-specific IgE and IgG1. Histological studies show that U0126 dramatically inhibited OVA-induced lung tissue eosinophilia, airway mucus production, and expression of VCAM-1 in lung tissues. In addition, U0126 significantly (p < 0.05) suppressed OVA-induced airway hyperresponsiveness to inhaled methacholine in a dose-dependent manner. Western blot analysis of whole lung lysates shows that U0126 markedly attenuated OVA-induced tyrosine phosphorylation of ERK1/2. Taken together, our findings implicate that inhibition of ERK signaling pathway may have therapeutic potential for the treatment of allergic airway inflammation.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Butadienes; Cytokines; Disease Models, Animal; Enzyme Inhibitors; Eosinophils; Immunoglobulin G; Male; Mice; Mice, Inbred BALB C; Mitogen-Activated Protein Kinases; Mucus; Nitriles; Ovalbumin; Phosphorylation; Vascular Cell Adhesion Molecule-1

Efficacy of beta2-agonists (B2), anticholinergics, corticosteroids, anti-inflammatory drugs or antihistamines against methacholine, histamine, or allergen-induced lung constriction was tested in ovalbumin sensitized guinea pigs. Airway resistance (Raw) and parenchymal damping (G) and elastance (H) were determined from low-frequency forced oscillatory input impedance (ZL). ZL was measured under control conditions, during iv infusions of methacholine, and following iv boluses of histamine or ovalbumin. In decreasing potency, ipratropium, salmeterol, fluticasone or cromoglycate reduced the methacholine-induced increases in Raw and G. Only antihistamines had any effects on the histamine-induced increased in Raw, G, and H. The ovalbumin-induced increases in Raw, G and H in the control animals (120 +/- 90%, 201 +/- 126%, 86 +/- 71%) were markedly reduced by pretreatments with antihistamines (18 +/- 11%, 13 +/- 9%, 3+/- 3%) and cromoglycate (29 +/- 13%, 35 +/- 22%, 18 +/- 10%). Bronchoconstriction induced by muscarinic receptor stimulation is inhibited by anticholinergic, anti-inflammatory or beta2-agonist pretreatment; antihistaminic or anti-inflammatory premedication is beneficial if the release of histamine via an exposure to allergen is responsible for the fall in lung function.

Topics: Adrenergic Agonists; Adrenergic beta-2 Receptor Agonists; Albuterol; Allergens; Analysis of Variance; Androstadienes; Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchoconstriction; Bronchoconstrictor Agents; Bronchodilator Agents; Cholinergic Antagonists; Cromolyn Sodium; Dose-Response Relationship, Drug; Drug Interactions; Fluticasone; Guinea Pigs; Histamine; Histamine H1 Antagonists; Ipratropium; Male; Methacholine Chloride; Ovalbumin; Salmeterol Xinafoate

Interleukin (IL)-13 is regarded as being a central effector in the pathophysiology of airway hyperresponsiveness. We have described a mouse model in which chronic allergen exposure results in sustained airway hyperresponsiveness and aspects of airway remodeling, and here sought to demonstrate that this component of airway hyperresponsiveness is independent of biologically active IL-13. Sensitized mice were subjected to either brief or chronic periods of allergen exposure and studied 24 hours after brief or 4 weeks after chronic allergen inhalation. A soluble murine anti-IL-13 receptor fusion protein that specifically binds to and neutralizes IL-13 was given daily during the 4 days before the day of outcome measurements in both protocols. Outcome measurements included airway responses to intravenous methacholine, bronchoalveolar lavage fluid cell counts, and airway morphometry. Compared with the saline control, brief allergen challenge resulted in airway hyperresponsiveness, which was prevented by anti-IL-13 treatment. Chronic allergen challenge resulted in sustained airway hyperresponsiveness and indices of airway remodeling; IL-13 blockade failed to reverse this sustained airway hyperresponsiveness. These results confirm that IL-13 is critical for the development of airway hyperresponsiveness associated with brief allergen exposure, but is not necessary to maintain the sustained airway hyperresponsiveness associated with airway remodeling.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Female; Interleukin-13; Interleukin-13 Receptor alpha1 Subunit; Mice; Mice, Inbred BALB C; Ovalbumin; Receptors, Interleukin; Receptors, Interleukin-13

Histamine-1-receptor (H1R)-antagonists were shown to influence various immunological functions on different cell types and may thus be employed for immune-modulating strategies for the prevention of primary immune responses.. The aim of this study was to investigate the effects of an H1R-antagonist on allergen-induced sensitization, airway inflammation (AI) and airway hyper-reactivity (AHR) in a murine model.. BALB/c mice were systemically sensitized with ovalbumin (OVA) (six times, days 1-14) and challenged with aerosolized allergen (days 28-30). One day prior to the first and 2 h prior to every following sensitization, mice received either 1 or 0.01 microg of desloratadine (DL) or placebo per os.. Sensitization with OVA significantly increased specific and total IgE and IgG1 serum levels, as well as in vitro IL-5 and IL-4 production by spleen and peribronchial lymph node (PBLN) cells. Sensitized and challenged mice showed a marked eosinophilic infiltration in broncho-alveolar lavage fluids and lung tissues, and developed in vivo AHR to inhaled methacholine. Oral treatment with DL prior to OVA sensitization significantly decreased production of OVA-specific IgG1, as well as in vitro Th2-cytokine production by spleen and PBLN cells, compared with OVA-sensitized mice. Moreover, eosinophilic inflammation and development of in vivo AHR were significantly reduced in DL-treated mice, compared with sensitized controls.. Treatment with H1R-anatagonist prior to and during sensitization suppressed allergen-induced Th2 responses, as well as development of eosinophilic AI and AHR. This underscores an important immune modulating function of histamine, and implies a potential role of H1R-anatagonists in preventive strategies against allergic diseases.

Topics: Administration, Oral; Allergens; Animals; Bronchial Hyperreactivity; Cells, Cultured; Female; Histamine H1 Antagonists; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Interleukin-4; Interleukin-5; Loratadine; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Models, Animal; Ovalbumin; Spleen

Nerve growth factor (NGF) exerts an important functional impact on the pathogenesis of allergic diseases. Data obtained in animal models of allergic bronchial asthma indicate that NGF alters sensory nerve function and promotes allergic inflammation, bronchial hyper-reactivity, and airway obstruction.. To further delineate the effects of NGF on airway inflammation, we employed a transgenic (tg) animal model of allergic inflammation and asthma.. NGF-tg mice, which overexpress NGF in Clara cells of the airways, were compared with wild-type (wt) littermates regarding their ability to mount IgE-related airway inflammatory responses. Mice were sensitized intraperitoneally to ovalbumin (OVA) and locally challenged via the airways according to established protocols.. NGF-tg mice displayed enhanced levels of OVA-specific IgE antibody titres after repeated OVA aerosol exposure. In the airways, increased numbers of eosinophils were detected. These results were confirmed to be NGF specific, because similar results were obtained following local application of NGF into the airways of wt mice. The effect of NGF was partly mediated via neuropeptides, as treatment of OVA-sensitized NGF-tg mice with the dual neurokinin (NK) receptor NK-1/NK-2 antagonist partly prevented enhanced airway inflammation.. The present data indicate an important functional role of NGF in allergic airway inflammation and point to an involvement of tachykinins as mediators of NGF effects.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Eosinophils; Humans; Interleukin-5; Leukocyte Count; Lung; Mice; Mice, Transgenic; Models, Animal; Nerve Growth Factors; Ovalbumin; Peptides, Cyclic; Receptors, Tachykinin; Tachykinins

The purpose of this study was to examine the effects of NO(2), a major component of air pollution, on airway eosinophilic inflammation and bronchial hyperreactivity, using a mouse model of asthma.. BALB/c mice (eight mice per experimental group) were studied in a basic research laboratory at the University of Iowa.. Using a standard murine model of asthma, BALB/c mice were sensitized to ovalbumin (OVA) by intraperitoneal (IP) injections (days 1 and 7) and were challenged with aerosolized OVA (days 13 and 14). Some mice were exposed to NO(2) (2 ppm) in an exposure chamber for 24 h before undergoing OVA aerosol challenge. A control group was exposed to OVA alone.. The outcomes assessed included airway inflammation, bronchial hyperreactivity to inhaled methacholine, and goblet cell hyperplasia. We found that NO(2) exposure modestly increased airway neutrophilia but not airway eosinophilia in OVA-exposed mice. These mice exhibited epithelial damage and loss of epithelial mucin. Surprisingly, nonspecific bronchial hyperreactivity (ie, enhanced pause index) was not increased, although baseline smooth muscle tone was increased (p < 0.05) in the mice exposed to NO(2).. These data indicate that relatively short-term (24 h) exposure to NO(2) causes epithelial damage, reduced mucin expression, and increased tone of respiratory smooth muscle. Reduced mucin production may be a mechanism of injury following long-term exposure to inhaled NO(2). Despite enhancing epithelial damage in OVA-exposed mice, NO(2) exposure does not otherwise alter the expression of allergen-induced airway responses.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Female; Mice; Mice, Inbred BALB C; Nitrogen Dioxide; Ovalbumin; Oxidants, Photochemical; Respiratory Mucosa

Epidemiological evidence points to the inverse relationship between microbial exposure and the prevalence of allergic asthma and autoimmune diseases in Westernized countries. The molecular basis for this observation has not yet been completely delineated. Here we report that the administration of certain toll-like receptor (TLR) ligands, via the activation of innate immunity, induces high levels of indoleamine 2,3-dioxygenase (IDO), the rate-limiting enzyme of tryptophan catabolism in various organs. TLR9 ligand-induced pulmonary IDO activity inhibits Th2-driven experimental asthma. IDO activity expressed by resident lung cells rather than by pulmonary DCs suppressed lung inflammation and airway hyperreactivity. Our results provide a mechanistic insight into the various formulations of the hygiene hypothesis and underscore the notion that activation of innate immunity can inhibit adaptive Th cell responses.

Topics: Adoptive Transfer; Animals; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Dendritic Cells; Immunity, Innate; Indoleamine-Pyrrole 2,3,-Dioxygenase; Interferon-gamma; Interleukin-12; Ligands; Lung; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, SCID; Oligodeoxyribonucleotides; Ovalbumin; Receptors, Cell Surface; Spleen; Th2 Cells; Toll-Like Receptors; Tryptophan Oxygenase; Tumor Necrosis Factor-alpha

Allergic asthma is a complex syndrome characterized by airway obstruction, airway inflammation and airway hyper-responsiveness (AHR). Using a mouse model of allergen-induced AHR, we previously demonstrated that CD8-deficient mice develop significantly lower AHR, eosinophilic inflammation and interleukin (IL)-13 levels in bronchoalveolar lavage fluid compared with wild-type mice. These responses were restored by adoptive transfer of antigen-primed CD8(+) T cells. Previously, two distinct populations of antigen-experienced CD8(+) T cells, termed effector (T(EFF)) and central memory (T(CM)) cells, have been described. After adoptive transfer into CD8-deficient mice, T(EFF), but not T(CM), cells restored AHR, eosinophilic inflammation and IL-13 levels. T(EFF), but not T(CM), cells accumulated in the lungs, and intracellular cytokine staining showed that the transferred T(EFF) cells were a source of IL-13. These data suggest an important role for effector CD8(+) T cells in the development of AHR and airway inflammation, which may be associated with their Tc2-type cytokine production and their capacity to migrate into the lung.

Topics: Adoptive Transfer; Allergens; Alum Compounds; Analysis of Variance; Animals; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; CD8 Antigens; CD8-Positive T-Lymphocytes; Cells, Cultured; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Interleukin-13; Lung; Methacholine Chloride; Mice; Mice, Mutant Strains; Ovalbumin; T-Lymphocyte Subsets

Allergen-induced sensitization and airway disease are the results of adverse immune reactions against environmental antigens that may be prevented or inhibited by immune modifying strategies.. To investigate the effects of the novel immune response modifier resiquimod (R-848), from the family of imidazol-derivates, in a murine model of allergen-mediated Th2-immune responses and concomitant airway inflammation and airway hyper-reactivity.. BALB/c mice were systemically sensitized with ovalbumin (OVA) on days 1 and 14 and challenged with OVA aerosol on days 28 and 29. R-848 was applied intranasally to sensitized animals once prior to the first allergen airway challenge, on day 27.. A single application of R-848 significantly reduced numbers of eosinophils and lymphocytes in bronchoalveolar lavage fluid and inhibited mucus gland hyperplasia, compared with sensitized and challenged controls. Associated with the decrease in airway inflammation, single intranasal treatment with R-848 abolished the development of airway hyper-reactivity after allergen sensitization and airway challenges. Additionally, Th2-cytokine production in lung tissues from sensitized and R-848-treated animals was reduced, whereas IL-12 and IFN-gamma production was increased, compared with non-treated sensitized mice.. These data indicate that R-848 effectively inhibits allergen-induced airway inflammation and hyper-reactivity by modulation of increased Th2-immune responses.

Topics: Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Female; Imidazoles; Immunologic Factors; Interferon-gamma; Interleukin-12; Interleukin-4; Interleukin-5; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells

When wild-type BALB/c mice were transferred with OVA-specific Th2 cells followed by OVA inhalation, a severe eosinophilia, mucus hypersecretion and airway hyper-responsiveness (AHR) was induced in parallel with a marked elevation of IL-4, IL-5 and IL-13 levels in bronchoalveolar lavage fluid (BALF). However, neither eosinophilia, AHR nor mucus hypersecretion was induced in Th2 cell-transferred STAT6-/- mice. The failure of eosinophilia was not due to the defect of Th2 cytokine production in BALF of STAT6-/- mice transferred with Th2 cells, but because of the defect of STAT6-dependent eotaxin production. Indeed, intranasal administration of eotaxin reconstituted pulmonary eosinophilia but not AHR and mucus hypersecretion in OVA-inhalated STAT6-/- mice. These results initially provided direct evidence that STAT6-dependent eotaxin production is essential for pulmonary eosinophilia. We also dissociated the role of STAT6 for eosinophilia from that for AHR and mucus hypersecretion. Thus, STAT6 also plays a critical role at late phase of Th2-dependent allergy induction.

Topics: Administration, Intranasal; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Differentiation; Chemokine CCL11; Chemokines, CC; Eosinophilia; Hypersensitivity; Lung; Mice; Mucus; Ovalbumin; Recombinant Proteins; STAT6 Transcription Factor; Th2 Cells; Trans-Activators

Considerable variation exists in the protocols used to induce hyperresponsiveness in murine models of allergic sensitisation. We examined the effect of varying the number of antigen exposures at challenge on the development of methacholine responsiveness in systemically sensitised mice.. BALB/c mice were sensitised with ovalbumin (OVA), challenged with 1, 3 or 6 OVA aerosols. Lung function was measured using low frequency forced oscillations and partitioned into components representing the airways (Raw) and lung parenchyma (tissue damping (G) and tissue elastance (H)). Responsiveness to inhaled methacholine (MCh), inflammatory cell profile and circulating IgE were assessed 24 and 48 hours after challenge. The threshold dose of MCh required to elicit a detectable response (sensitivity) and response to 30 mg x mL(-1) (maximal response) were determined for each compartment.. Sensitivity; All three OVA protocols resulted in an increased sensitivity to MCh in Raw but not in G or H. These responses where present at 24 and 48 hrs, except 1 OVA aerosol in which changes had resolved by 48 hrs. Maximal response; 1 OVA aerosol increased maximal responses in Raw, G and H at 24 hrs, which was gone by 48 hrs. Three OVA aerosols increased responses in H at 48 hrs only. Six OVA challenges caused increases in Raw, G and H at both 24 and 48 hrs. Eosinophils increased with increasing antigen challenges. IgE was elevated by OVA sensitisation but not boosted by OVA aerosol challenge.. The pattern of eosinophilia, IgE and MCh responsiveness in mice was determined by antigen dose at challenge. In this study, increased sensitivity to MCh was confined to the airways whereas increases in maximal responses occurred in both the airway and parenchymal compartments. The presence of eosinophilia and IgE did not always coincide with increased responsiveness to inhaled MCh. These findings require further systematic study to determine whether different mechanisms underlie airway and parenchymal hyperresponsiveness post antigen challenge.

Topics: Animals; Antigens; Bronchi; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Disease Models, Animal; Female; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin

Oxidative stress plays an important role in the pathogenesis of bronchial asthma. An excess production of reactive oxygen species (ROS) and defective endogenous antioxidant defense mechanisms may be present in asthma. Reduced glutathione (GSH) is one of the most important reducing agents against oxidant free radicals. A reducing agent, L-2-oxothiazolidine-4-carboxylic acid (OTC), a prodrug of cysteine, increases intracellular GSH. We have used a mouse model for asthma to determine effects of OTC on allergen-induced bronchial inflammation and airway hyper-responsiveness. The administration of OTC reduced bronchial inflammation and airway hyper-responsiveness. ROS generation in bronchoalveolar lavage fluids was increased by ovalbumin (OVA) inhalation, but this increase was diminished by administration of OTC. The increased IL-4, IL-5, IL-13, and eosinophil cationic protein levels in lungs after OVA inhalation were significantly reduced by the administration of OTC. In addition, the increased expression of ICAM-1, VCAM-1, RANTES, and eotaxin in lungs after OVA inhalation was significantly reduced by the administration of OTC. We also showed that the increased NF-kappaB levels in nuclear protein extracts of lung tissues at 72 h after OVA inhalation were decreased by the administration of OTC. These findings suggest that OTC may reduce airway inflammation and hyper-responsiveness through regulation of NF-kappaB activity.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchitis; Cell Adhesion Molecules; Chemokines; Disease Models, Animal; Eosinophil Cationic Protein; Interleukins; Lung; Mice; NF-kappa B; Ovalbumin; Prodrugs; Pyrrolidonecarboxylic Acid; Reactive Oxygen Species; Thiazoles; Thiazolidines

Airway hyperresponsiveness, airway eosinophilia and increased IgE levels in serum are the important characteristic features of asthma. We evaluated the potential of para-Bromophenacyl bromide (PBPB), a known phospholipase A(2) inhibitor, on allergen-induced airway hyperresponsiveness in a mouse model. We sensitized and challenged mice with ovalbumin (OVA) to develop airway hyperresponsiveness as assessed by airway constriction and airway hyperreactivity (AHR) to methacholine (MCh) induced by allergen. The mice were orally treated with PBPB (0.1, 1 and 10 mg/kg) during or after OVA-sensitization and OVA-challenge to evaluate its protective or reversal effect on airway constriction and AHR to MCh. Determination of OVA-induced airway constriction and AHR to MCh were performed by measuring specific airway conductance (SGaw) using non-invasive dual-chamber whole body-plethysmography. We observed that PBPB (1 mg/kg) significantly reduced OVA-induced airway constriction and AHR to MCh (p<0.01). PBPB (1 mg/kg) treatment significantly inhibited PLA(2) activity in the BAL fluid. Cytokine analysis of the BAL fluid revealed that PBPB caused an increase in interferon-gamma (IFN-gamma) (p<0.02) and a decrease in interleukin-4 (IL-4) (p<0.05) and interleukin-5 (IL-5) (p<0.05) levels. The OVA-specific serum IgE levels (p<0.01) and the BAL eosinophils (p<0.001) were also reduced significantly. Thus, PBPB is capable of modulating allergen induced cytokine levels and serum IgE levels, and alleviating allergen induced airway hyperresponsiveness and eosinophils in mice. These data suggest that PBPB could be useful in the development of novel agents for the treatment of allergen induced airway hyperresponsiveness.

Topics: Acetophenones; Administration, Inhalation; Aerosols; Airway Obstruction; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cytokines; Disease Models, Animal; Drug Evaluation, Preclinical; Eosinophils; Immunization; Immunoglobulin E; India; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Phospholipases A

Knockout mice studies have revealed that NF-kappaB plays a critical role in Th2 cell differentiation and is therefore required for induction of allergic airway inflammation. However, the questions of whether NF-kappaB also plays a role in the effector phase of airway allergy and whether inhibiting NF-kappaB could have therapeutic value in the treatment of established asthma remain unanswered. To address these issues, we have assessed in OVA-sensitized wild-type mice the effects of selectively antagonizing NF-kappaB activity in the lungs during OVA challenge. Intratracheal administration of NF-kappaB decoy oligodeoxynucleotides to OVA-sensitized mice led to efficient nuclear transfection of airway immune cells, but not constitutive lung cells and draining lymph node cells, associated with abrogation of NF-kappaB activity in the airways upon OVA provocation. NF-kappaB inhibition was associated with strong attenuation of allergic lung inflammation, airway hyperresponsiveness, and local production of mucus, IL-5, IL-13, and eotaxin. IL-4 and OVA-specific IgE and IgG1 production was not reduced. This study demonstrates for the first time that activation of NF-kappaB in local immune cells is critically involved in the effector phase of allergic airway disease and that specific NF-kappaB inhibition in the lungs has therapeutic potential in the control of pulmonary allergy.

Topics: Allergens; Animals; Apoptosis; Asthma; Bronchial Hyperreactivity; Chemokine CCL11; Chemokines, CC; Female; Immunoglobulins; Interleukin-13; Interleukin-4; Interleukin-5; Intubation, Intratracheal; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Mucus; NF-kappa B; Oligodeoxyribonucleotides, Antisense; Ovalbumin; Respiratory Mucosa; Th2 Cells; Thorax

Although the critical role of cysteinyl leukotrienes (cysLTs) in the inflammation, especially eosinophilic lung inflammation, in asthma has been well documented, their role in the early stage of Ag-specific immune response has not been completely clarified. In the present study, with a mouse model of asthma and in vitro studies we demonstrated that cysLTs potentiated dendritic cell (DC) functions such as Ag-presenting capacity and cytokine production. The cysLT-1 receptor antagonist (LTRA) strongly suppressed the activation of these DC functions and led to inhibition of subsequent not only Th2, but also Th1, responses in the early stage of immune response. Moreover, treatment with LTRA during the early stage of the immune response potently suppressed the development of Ag inhalation-induced eosinophilic airway inflammation, mucus production, and airway hyper-reactivity in vivo. Treatment with LTRA significantly increased PGE(2) production in the lung, and treatment with the cyclooxygenase inhibitor indomethacin abolished LTRA's suppressive effect on DCs and deteriorated the Th2 and Th1 responses and airway inflammation. With in vitro studies, we also confirmed that cysLTs production by DCs increased with LPS stimulation, and that LTRA directly suppressed the alloantigen-presenting capacity of DCs. These results suggested that cysLTs potentiate DC functions both in vivo and in vitro, and that LTRA could be beneficial to suppress the initial immune response in many immune-mediated disorders beyond asthma.

Topics: Adjuvants, Immunologic; Animals; Anti-Inflammatory Agents, Non-Steroidal; Antigens; Arachidonate 5-Lipoxygenase; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cells, Cultured; Chromones; Cysteine; Dendritic Cells; Inflammation; Leukotriene Antagonists; Leukotrienes; Lung; Male; Membrane Proteins; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Receptors, Leukotriene; Th1 Cells

The transcription factor, nuclear factor (NF)-kappaB, which transactivates various genes for proinflammatory cytokines and many other immunoregulatory genes, plays an important role in the regulation of various inflammatory diseases including asthma. Its increased activation has been demonstrated in the lungs after allergen challenge and in airway epithelial cells and macrophages of asthmatic patients. In the present study, we investigated whether the pretreatment with p65 antisense results in a significant inhibition of asthmatic reactions in a mouse model. Mice sensitized and challenged with ovalbumin (OVA) showed typical asthmatic reactions as follows: (1) an increase in the number of eosinophil in bronchoalveolar lavage fluid; (2) a marked influx of inflammatory cells into the lung around blood vessels and airways, and airway luminal narrowing; (3) the development of airway hyperresponsiveness; (4) the detection of TNF-alpha and Th2 cytokines, such as IL-4 and IL-5 in the bronchoalveolar lavage (BAL) fluid; and (5) detection of allergen-specific IgE and IgG in the serum. Two successive intravenous administration of p65 antisense before the last airway OVA challenge resulted in a significant inhibition of all asthmatic reactions, whereas the p65 nonsense did not produce such effects. In addition, the p65 antisense inhibition of asthmatic reaction appears to be due to the initial suppression of an allergen-specific IgE response, inducing degranulation of mast cells through the cross-linking of allergen-specific IgE. This study may provide evidence that NF-kappaB plays a critical role in the pathogenesis of asthma in mice.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Blotting, Western; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Electrophoretic Mobility Shift Assay; Enzyme-Linked Immunosorbent Assay; Immunoglobulin E; Immunoglobulin G; Male; Mice; Mice, Inbred C57BL; NF-kappa B; Oligonucleotides, Antisense; Ovalbumin; Transcription Factor RelA

Inflammation of the asthmatic airway is usually accompanied by increased vascular permeability and plasma exudation. Cysteinyl leukotrienes (cysLTs) potently elicit increased vascular permeability in airways, leading to airway edema. Vascular endothelial growth factor (VEGF) is 1 of the most potent proangiogenic cytokines and also increases vascular permeability so that plasma proteins can leak into the extravascular space. However, the mechanisms by which cysLTs induce increased vascular permeability are not clearly understood.. An aim of the current study was to determine the role of the cysLTs, more specifically in the increase of vascular permeability.. We used a BALB/c mouse model of allergic asthma to examine effects of cysLT receptor antagonists on bronchial inflammation and airway hyperresponsiveness, more specifically on the increase of vascular permeability.. These mice develop the following typical pathophysiological features of asthma in the lungs: increased numbers of inflammatory cells of the airways, airway hyperresponsiveness, increased vascular permeability, and increased levels of VEGF. Administration of cysLT receptor antagonists markedly reduced plasma extravasation and VEGF levels in allergen-induced asthmatic lungs.. These results indicate that cysLT receptor antagonists modulate vascular permeability by reducing VEGF expression and suggest that cysLT receptor may regulate the VEGF expression.

Topics: Acetates; Animals; Bronchial Hyperreactivity; Capillary Permeability; Chromones; Cyclopropanes; Cytokines; Female; Indoles; Membrane Proteins; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Quinolines; Receptors, Leukotriene; Receptors, Vascular Endothelial Growth Factor; Sulfides; Transcription Factor RelA; Vascular Endothelial Growth Factor A

Airways display robust NF-kappaB activation and represent targets for anti-inflammatory asthma therapies, but the functional importance of NF-kappaB activation in airway epithelium remains enigmatic. Therefore, transgenic mice were created in which NF-kappaB activation is repressed specifically in airways (CC10-IkappaBalpha(SR) mice). In response to inhaled Ag, transgenic mice demonstrated significantly ameliorated inflammation, reduced levels of chemokines, T cell cytokines, mucus cell metaplasia, and circulating IgE compared with littermate controls. Despite these findings, Ag-driven airways hyperresponsiveness was not attenuated in CC10-IkappaBalpha(SR) mice. This study clearly demonstrates that airway epithelial NF-kappaB activation orchestrates Ag-induced inflammation and subsequent adaptive immune responses, but does not contribute to airways hyperresponsiveness, the cardinal feature that underlies asthma.

Topics: Allergens; Animals; Antibody Specificity; Bronchial Hyperreactivity; Chemokines; Cytokines; I-kappa B Proteins; Immunoglobulin E; Inflammation; Inflammation Mediators; Lung; Mice; Mice, Inbred BALB C; Mice, Transgenic; Mucus; NF-kappa B; NF-KappaB Inhibitor alpha; Ovalbumin; Respiratory Hypersensitivity; Respiratory Mucosa; T-Lymphocyte Subsets

There is currently considerable interest in the potential value of selective inhibitors of cyclic nucleotide phosphodiesterase 4 in the treatment of asthma. However, whether they influence eosinophilic airway inflammation-associated cough remains unclear. The objective of this study was to investigate the effects of selective phosphodiesterase 4 inhibitor SB207499 on cough response and airway inflammation in guinea pigs sensitized and challenged with ovalbumin.. Forty sensitized guinea pigs were randomly divided into four groups: control (n = 10), challenge (n = 10), SB207499 (n = 10) and aminophylline (n = 10), then challenged with aerosol of 1% ovalbumin or saline. Two hours later, animals were intraperitoneally injected with either saline, 25 mg/kg of SB207499 or aminophylline. At the 24th hour, the injection was repeated with 2.5 mg/kg and 25 mg/kg SB207499 or aminophylline, then cough response to inhaled capsaicin and airway responsiveness to methacholine inducing a 150% of the peak airway pressure to the baseline (PC150) was measured. Finally, total cell number and differentials in bronchoalveolar lavage fluid were analysed.. The cough frequency per 3 minutes and PC150 in the challenge group were (22 +/- 4) times/3 minutes and (198 +/- 54) microg/ml, which were significantly different from (6 +/- 2) times/3 minutes and (691 +/- 81) microg/ml in the control group (P < 0.05, respectively). The injection of 25 mg/kg SB207499 significantly inhibited the increased cough response and airway hyperresponsiveness, the cough frequency and PC150 in guinea pigs were (13 +/- 2) times/3 minutes and (680 +/- 81) microg/ml (P < 0.05), which differed significantly from (18 +/- 2) times/3 minutes and (400 +/- 86) microg/ml after the administration of the same dose of aminophylline (P < 0.05). The inhibition of SB207499 on cough response was dose-dependent. Similarly, SB207499 decreased the total cell number and percentage of eosinophils in bronchoalveolar lavage fluid to (2.1 +/- 0.5) x 10(6)/ml and (20 +/- 5)% respectively, which were significantly different from (3.2 +/- 0.5) x 10(6)/ml and (29 +/- 5)% in the aminophylline group (P < 0.05, respectively) or (4.2 +/- 0.7) x 10(6)/ml and (35 +/- 4)% in the challenge group (P < 0.05, respectively).. Phosphodiesterase 4 inhibitor may be more useful than aminophylline for cough associated with eosinophilic airway inflammation via inhibiting airway inflammation and airway hyperresponsiveness.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cough; Cyclic AMP; Cyclic Nucleotide Phosphodiesterases, Type 4; Cyclohexanecarboxylic Acids; Dose-Response Relationship, Drug; Guinea Pigs; Male; Nitriles; Ovalbumin; Phosphodiesterase Inhibitors

Allergic asthma is an inflammatory lung disease caused by a T(H)2-driven immune response. However, intranasal exposures to soluble antigen lead to mucosal tolerance, and the mechanism involved in generation of T(H)2 responses to inert inhaled allergens is unknown.. The aim of this study was to investigate whether CD1d-restricted natural killer T (NKT) cells can contribute to the induction of T(H)2-dependent allergic asthma in a mouse model.. To investigate the effect of NKT cells on the development of asthma, NKT cell ligand, alpha-galactosylceramide (alphaGC), was used with antigens. We intranasally sensitized Balb/c mice with various combinations of antigen and alphaGC for 3 consecutive days and challenged them 2 weeks later with an aerosol of ovalbumin. NKT cell-deficient or T(H) cell-deficient mice were immunized by administering ovalbumin and alphaGC together, and ovalbumin inhalation.. Only when immunized with ovalbumin plus alphaGC, airway hyperreactivity, airway eosinophils, elevated IgE level, and T(H)2-cytokine production were observed in Balb/c mice. Ovalbumin alone, alphaGC alone, or BSA plus alphaGC-immunized mice did not induce asthma. Studies in NKT cell-deficient, or CD4(+) T-cell-deficient mice intranasally exposed to ovalbumin plus alphaGC did not show the development of asthma. An increase of NKT cells in bronchoalveolar lavage was observed in the pathologic states.. These data demonstrate that NKT cells can play crucial roles in allergen sensitization and pathologic states in asthma. Furthermore, our new asthma model using alphaGC will be very useful to induce asthma and to dissect the role of NKT cells and other cells in asthma.

Topics: Administration, Intranasal; Allergens; Animals; Antigens, CD1; Antigens, CD1d; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Female; Galactosylceramides; Histocompatibility Antigens Class II; Killer Cells, Natural; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Ovalbumin

Galectin-3 is a member of a beta-galactoside-binding animal lectin family. Previous in vitro studies have demonstrated that galectin-3 is involved in a number of activities; however, the roles of this lectin in physiological and pathological processes in vivo remain to be elucidated. Herein, we show, in a murine model of ovalbumin (OVA)-induced asthma that 1) peribronchial inflammatory cells expressed large amounts of galectin-3; 2) bronchoalveolar lavage fluid from OVA-challenged mice contained significantly higher levels of galectin-3 compared to control mice; and 3) macrophages in bronchoalveolar lavage fluid were the major cell type that contained galectin-3. We investigated the role of galectin-3 in the allergic airway response by comparing galectin-3-deficient (gal3(-/-)) mice and wild-type (gal3(+/+)) mice. OVA-sensitized gal3(-/-) mice developed fewer eosinophils and lower goblet cell metaplasia, after airway OVA challenge compared to similarly treated gal3(+/+) mice. In addition, the OVA-sensitized gal3(-/-) mice developed significantly less airway hyperresponsiveness after airway OVA challenge compared to gal3(+/+) mice. Finally, gal3(-/-) mice developed a lower Th2 response, but a higher Th1 response, suggesting that galectin-3 regulates the Th1/Th2 response. We conclude that galectin-3 may play an important role in the pathogenesis of asthma and inhibitors of this lectin may prove useful for treatment of this disease.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cytokines; Disease Models, Animal; Eosinophils; Female; Galectin 3; Immunization; Immunoglobulin E; Macrophages; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; RNA, Messenger; Th1 Cells; Th2 Cells

Rhamnus nakaharai Hayata (Rhamnaceae) has been used as a folk medicine in Taiwan for treating constipation, inflammation, tumors, and asthma. 3-O-Methylquercetin (3-MQ), a main constituent of the plant, has been reported to inhibit total cAMP- and cGMP-phosphodiesterase (PDE) of guinea pig trachealis at low concentrations. 3-MQ has been also reported to more selectively inhibit PDE3 than PDE4 with a low K(m) value. Therefore we were interested in investigating its suppressive effects on ovalbumin (OVA)-induced airway hyperresponsiveness in vivo and in vitro. 3-MQ (3-30 micromol/kg, i. p.) significantly suppressed the enhanced pause (Penh) value induced by aerosolized methacholine (50 mg/mL) in sensitized mice after secondary allergen challenge. 3-MQ (3-30 micromol/kg, i. p.) also significantly suppressed total inflammatory cells, macrophages, neutrophils, and eosinophils, but not lymphocytes. In addition, 3-MQ (3 micromol/kg, i. p.) significantly decreased the secretion of TNF-alpha, and at the highest dose (30 micromol/kg, i. p.) even decreased the secretions of IL-4, IL-5, and TNF-alpha. 3-MQ (1-10 microM) as well as Ro 20-1724 (3-30 microM), a selective PDE4 inhibitor, significantly attenuated OVA (100 microg/mL)-induced contractions. 3-MQ (30 microM) as well as milrinone (1-10 microM), a selective PDE3 inhibitor, significantly enhanced baseline contractions in isolated guinea pig left and right atria. However, neither 3-MQ nor milrinone significantly affected baseline beating rate in the right atria. 3-MQ (3-30 micromol/kg, i. p.) did not significantly affect systolic pressure in conscious mice. In conclusion, 3-MQ has both anti-inflammatory and bronchodilating effects, and has the potential for use in the treatment of asthma at a dose without affecting blood pressure.

Topics: Animals; Bronchial Hyperreactivity; Bronchodilator Agents; Dose-Response Relationship, Drug; Female; Guinea Pigs; Injections, Intraperitoneal; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Phosphodiesterase Inhibitors; Phytotherapy; Quercetin; Rhamnus

The biochemical mechanisms underlying the development of sensitization-induced airway hyperresponsiveness (AHR) in asthma are poorly defined. Alterations in the regulation of intracellular calcium may play an important role in its pathogenesis. We carried out this study to see the effect of sensitization with ovalbumin on membrane ion fluxes and intracellular calcium in a guinea pig model.. Airway reactivity to inhaled histamine was measured initially and after sensitization with ovalbumin in 28 guineapigs. Intracellular calcium [Ca(2+)]i was measured in tracheal smooth muscle cells and peripheral leukocytes using fluorescent dye FURA 2AM. Calcium and sodium ion influx across the cell membrane was measured in leukocytes. Ouabain-sensitive Rubidium ((86)Rb) influx was measured in tracheal smooth muscles cells. The activities of Na(+), K(+) ATPase and Ca(2+) ATPase were measured in tracheal smooth muscle cells. Lipid peroxides were measured in plasma.. Airway responsiveness was significantly (P<0.001) increased after sensitization along with an increase in [Ca2+]i levels in leukocytes and tracheal smooth muscle cells, higher rates of (45)Ca and (22)Na influx in leukocytes and higher (86)Rb influx rates in tracheal smooth muscle cells, and increased levels of lipid peroxides in plasma.. In guineapig model of asthma sensitization to allergen increased the membrane permeability to calcium and sodium, and intracellular calcium levels. These alterations may play a role in the pathogenesis of airway hyper-responsiveness following sensitization.

Topics: Animals; Bronchial Hyperreactivity; Calcium; Cell Membrane; Fluorescent Dyes; Fura-2; Guinea Pigs; Histamine; Ions; Leukocytes; Lipid Peroxidation; Male; Myocytes, Smooth Muscle; Ovalbumin; Rubidium Radioisotopes; Sodium Radioisotopes; Sodium-Potassium-Exchanging ATPase; Trachea

To determine a change in airway smooth muscle contractility in a murine model of allergic asthma, the responsiveness of airway smooth muscles isolated from ovalbumin (OA)-sensitized and -challenged mice was compared with that from control animals.. Actively sensitized mice were repeatedly challenged by ovalbumin (OA) antigen inhalation. Twenty-four h after the last antigen challenge, tracheal and bronchial smooth muscle responsiveness to acetylcholine (ACh) and endothelin-1 (ET-1) were measured. Airway microvascular leakage and histochemistry were also determined as indices of airway inflammation.. Both the ACh and ET-1 responsiveness of bronchial, but not tracheal, smooth muscles were significantly augmented in OA-challenged mice, whereas no significant change in the expression levels of M2, M3 and ETB receptors was observed. The Evans blue dye extravasation in the main bronchial, but not tracheal, tissue of OA-challenged mice was significantly increased as compared with that of sensitized control animals. A marked inflammatory cells infiltration was also observed in bronchial but not tracheal tissues of OA-challenged mice.. Repeated antigen challenge to sensitized mice caused a hyperresponsiveness of bronchial, but not tracheal, smooth muscle accompanied with bronchial tissue inflammation.

Topics: Acetylcholine; Animals; Antigens; Asthma; Bronchi; Bronchial Hyperreactivity; Capillary Permeability; Disease Models, Animal; Endothelin-1; Immunization; Male; Mice; Mice, Inbred BALB C; Muscle Contraction; Muscle, Smooth; Ovalbumin; Receptor, Endothelin B; Receptor, Muscarinic M1; Receptor, Muscarinic M2; Trachea

In mice, intratracheal challenges with antigen (ovalbumin) or recombinant murine interleukin-13 (IL-13) induce lung inflammation, bronchial hyperreactivity (BHR), and mucus accumulation as independent events (Singer M, Lefort J, and Vargaftig BB. Am J Respir Cell Mol Biol 26: 74-84, 2002), largely mediated by leukotrienes (LT). We previously showed that LTC(4) was released 15 min after ovalbumin, and we show that it induces the expression of monocyte chemoattractant proteins 1 and 5 and KC in the lungs, as well as IL-13 mRNA. Instilled intratracheally, these chemokines induced BHR and mucus accumulation, which were inhibited by the 5-lipoxygenase inhibitor zileuton and by the cysteinyl-LT receptor antagonist MK-571, suggesting mediation by cysteinyl-LT. Because these chemokines also induced release of LT into the bronchoalveolar lavage fluid and IL-13 into the lungs, we hypothesize that LT- and chemokine-based loops for positive-feedback regulations cooperate to maintain and amplify BHR and lung mucus accumulation after allergic challenge and in situations where IL-13, LT, or chemokines are generated.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Hypersensitivity; Interleukin-13; Leukotriene C4; Leukotrienes; Lung; Male; Metaplasia; Mice; Mice, Inbred Strains; Mucins; Mucus; Ovalbumin; Recombinant Proteins; Respiratory Mucosa; RNA, Messenger

A strategy to deplete eosinophils from the lungs of ovalbumin (OVA)-sensitized/challenged mice was developed using antibody-mediated depletion. Concurrent administration [viz. the peritoneal cavity (systemic) and as an aerosol to the lung (local)] of a rat anti-mouse CCR3 monoclonal antibody resulted in the abolition of eosinophils from the lung such that the airway lumen was essentially devoid of eosinophils. Moreover, perivascular/peribronchial eosinophil numbers were reduced to levels indistinguishable from saline-challenged animals. This antibody-mediated depletion was not accompanied by effects on any other leukocyte population, including, but not limited to, T cells and mast cells/basophils. In addition, no effects were observed on other underlying allergic inflammatory responses in OVA-treated mice, including OVA-specific immunoglobulin production as well as T cell-dependent elaboration of Th2 cytokines. The ablation of virtually all pulmonary eosinophils in OVA-treated mice (i.e., without concurrent effects on T cell activities) resulted in a significant decrease in mucus accumulation and abolished allergen-induced airway hyperresponsiveness. These data demonstrate a direct causative relationship between allergen-mediated pulmonary pathologies and eosinophils.

Topics: Allergens; Animals; Antibodies, Monoclonal; Antibody Formation; Bronchial Hyperreactivity; Cell Count; Cytokines; Eosinophils; Goblet Cells; Immunoglobulins; Lung; Lymphocytes; Mice; Mice, Inbred C57BL; Mucus; Ovalbumin; Rats; Receptors, CCR3; Receptors, Chemokine; Th2 Cells

No current therapy is considered to be satisfactory for severe asthma, and alternative approaches are still required for what is a major unmet medical need. In this study, we compared the effect of a rapamycin derivative, SAR 943, with budesonide, using a murine model of lung inflammation and remodeling. Allergen challenge of ovalbumin-sensitized BALB/c mice induced an increase in the levels of interleukin-5 and interleukin-4; numbers of eosinophil, neutrophil, and lymphocyte; cellular fibronectin; lung epithelial cell proliferation and mucus hypersecretory phenotype; as well as hyperreactivity to methacholine. Both SAR 943 and budesonide, when given intranasally 1 hour before and 24 hours after the aerosol challenge, inhibited all of these parameters with a similar potency (effective dose 50% of 1 mg/kg). In primary cultured smooth muscle cells from human airways, SAR 943 dose dependently inhibited epidermal growth factor-induced proliferation but did not affect the basal cell proliferation. Neither the basal nor stimulated proliferation of a human bronchial epithelial cell line (16HBE14o-) was affected by SAR 943. In conclusion, SAR 943 is as effective as budesonide in inhibiting both lung inflammation and remodeling in a murine model of asthma. Hence, this class of compound could offer beneficial effects in patients with severe asthma.

Topics: Administration, Intranasal; Animals; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Budesonide; Cell Division; Cells, Cultured; Disease Models, Animal; Female; Immunohistochemistry; In Vitro Techniques; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Probability; Random Allocation; Reference Values; Sensitivity and Specificity; Sirolimus; Statistics, Nonparametric

In the airways, inhibitory M2 muscarinic receptors (M2Rs) on parasympathetic nerves limit acetylcholine release. Viral infection causes M2R dysfunction, which increases acetylcholine release and leads to airway hyperreactivity. In these studies we tested the role of CD8+ T cells in parainfluenza virus-induced hyperreactivity and M2R dysfunction in normal guinea pigs and in guinea pigs previously sensitized to ovalbumin. Depleting CD8+ T cells prevented virus-induced M2R dysfunction and hyperreactivity in sensitized animals, but not in nonsensitized animals. Sensitization increased the number of eosinophils in close relation to the airway nerves where, when activated, they release major basic protein, which binds to and blocks the M2Rs. Regardless of sensitization, viral infection decreased the number of visible tissue eosinophils, likely reflecting eosinophil degranulation via cytolysis. Depleting CD8+ T cells prevented this virus-induced eosinophil degranulation. In addition, an antiviral effect of sensitization, which we previously showed to be eosinophil mediated, was again seen. This was prevented by depletion of CD8+ Tcells. Thus, CD8+ T cells play a role in airway hyperreactivity and M2R dysfunction of sensitized virus-infected guinea pigs by mediating eosinophil degranulation near airway nerves. In contrast, CD8+ T cells are not necessary for virus-induced hyperreactivity and M2R dysfunction in nonsensitized guinea pigs.

Topics: Analysis of Variance; Animals; Asthma; Bronchial Hyperreactivity; CD8-Positive T-Lymphocytes; Cell Degranulation; Eosinophils; Female; Guinea Pigs; Immunization; Leukocyte Count; Ovalbumin; Paramyxoviridae Infections; Receptor, Muscarinic M2; Receptors, Muscarinic; Virus Diseases

Histamine has a variety of airway actions and is considered to be an important mediator in asthma. This study examined the role of endogenous histamine in allergic airway eosinophil recruitment and hyperresponsiveness using L-histidine decarboxylase gene knockout mice. Histamine levels of the airways in L-histidine decarboxylase knockout mice were largely diminished compared with wild-type mice. Inhalation challenge with ovalbumin (OVA) in OVA-sensitized wild-type mice caused eosinophil accumulation in the lung as well as airway hyperresponsiveness to methacholine 3 days after the challenge. The eosinophil recruitment was significantly reduced in the knockout mice. In the bone marrow, the proliferation of eosinophils was enhanced after OVA challenge in the wild-type mice; however, the proliferation was significantly reduced in the knockout mice. The induction of P-selectin in the lung after OVA challenge was also inhibited in the knockout mice. In contrast, airway hyperresponsiveness was not suppressed in the knockout mice. These results suggest that endogenous histamine is involved in the accumulation of eosinophils into the airways after allergic challenge, possibly acting in the bone marrow and producing P-selectin in the airways. Furthermore, allergen-induced airway hyperresponsiveness appeared to occur independently of airway eosinophilia in our present model.

Topics: Administration, Inhalation; Airway Resistance; Animals; Asthma; Bone Marrow Cells; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophilia; Eosinophils; Histamine; Histidine Decarboxylase; Immunoblotting; Interleukin-5; Leukocyte Count; Lung; Male; Methacholine Chloride; Mice; Mice, Knockout; Ovalbumin; Pulmonary Eosinophilia; Selectins; Time Factors

Allergic inflammation is mediated by Th2 cell-derived cytokines, including IL-4, IL-5, and IL-13, and down-regulated by IFN-gamma and IL-12. Tyk2 is a member of the Janus family of protein tyrosine kinases and is activated by a variety of cytokines: IFN-alphabeta, IL-6, IL-10, IL-12, and IL-13. In this study, we investigated the role of Tyk2 in the regulation of Ag-induced Th cell differentiation and Ag-induced allergic inflammation in the airways using Tyk2-deficient (Tyk2(-/-)) mice. When splenocytes were stimulated with antigenic peptide, IL-12-mediated Th1 cell differentiation was decreased, but IL-4-mediated Th2 cell differentiation was increased in Tyk2(-/-) mice. In vivo, Ag-specific IgE and IgG1 production was increased, but Ag-specific IgG2a production was decreased in Tyk2(-/-) mice as compared with those in control mice. In addition, Ag-induced eosinophil and CD4(+) T cell recruitment, as well as the production of Th2 cytokines in the airways, was increased in Tyk2(-/-) mice. Adoptive transfer experiments revealed that CD4(+) T cells were responsible for the enhanced Ag-induced eosinophil recruitment in Tyk2(-/-) mice. In contrast, although the level of IL-13 was increased in the airways of Tyk2(-/-) mice after Ag inhalation, the number of goblet cells, as well as Muc5ac mRNA expression, was decreased in Tyk2(-/-) mice. Together, these results indicate that Tyk2 plays a bilateral role in the regulation of allergic inflammation in the airways: Tyk2 plays a role in the down-regulation of Th2 cell-mediated Ab production and eosinophil recruitment in the airways by regulating Th1/Th2 balance toward Th1-type, while Tyk2 is necessary for the induction of IL-13-mediated goblet cell hyperplasia in the airways.

Topics: Administration, Inhalation; Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Differentiation; Cell Movement; Crosses, Genetic; Eosinophils; Epitopes, T-Lymphocyte; Goblet Cells; Hyperplasia; Immunoglobulin E; Inflammation; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Ovalbumin; Protein-Tyrosine Kinases; Proteins; Receptors, Antigen, T-Cell; Respiratory Hypersensitivity; Th2 Cells; TYK2 Kinase; Up-Regulation

Allergic asthma is characterized by airway hyperreactivity, inflammation, and a Th2-type cytokine profile favoring IgE production. Beneficial effects of TGF-beta and conflicting results regarding the role of Th1 cytokines have been reported from murine asthma models. In this study, we examined the T cell as a target cell of TGF-beta-mediated immune regulation in a mouse model of asthma. We demonstrate that impairment of TGF-beta signaling in T cells of transgenic mice expressing a dominant-negative TGF-beta type II receptor leads to a decrease in airway reactivity in a non-Ag-dependent model. Increased serum levels of IFN-gamma can be detected in these animals. In contrast, after injection of OVA adsorbed to alum and challenge with OVA aerosol, transgenic animals show an increased airway reactivity and inflammation compared with those of wild-type animals. IL-13 levels in bronchoalveolar lavage fluid and serum as well as the number of inducible NO synthase-expressing cells in lung infiltrates were increased in transgenic animals. These results demonstrate an important role for TGF-beta signaling in T cells in the regulation of airway responses and suggest that the beneficial effects observed for TGF-beta in airway hyperreactivity and inflammation may be due to its regulatory effects on T cells.

Topics: Administration, Inhalation; Aerosols; Alum Compounds; Animals; Antibody Specificity; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD2 Antigens; Cell Movement; Epitopes, T-Lymphocyte; Humans; Immunoglobulin E; Immunohistochemistry; Inflammation; Interferon-gamma; Interleukin-13; Lung; Mice; Mice, Inbred Strains; Mice, Transgenic; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Ovalbumin; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; T-Lymphocyte Subsets; Th1 Cells; Transforming Growth Factor beta

Asthma, a chronic inflammatory disease of the lung, is characterized by reversible airway obstruction and airway hyperresponsiveness (AHR), and is associated with increased production of IgE and Th2-type cytokines (IL-4, IL-5, and IL-13). Development of inflammation within the asthmatic lung depends on MHC class II-restricted Ag presentation, leading to stimulation of CD4(+) T cells and cytokine generation. Conventional MHC class II pathways require both MHC-associated invariant chain (Ii) and HLA-DM (H2-M in mice) chaperone activities, but alternative modes of Ag presentation may also promote in vivo immunity. In this study, we demonstrate that Ii(-/-) and H2-M(-/-) mice fail to develop lung inflammation or AHR following sensitization and challenge with OVA in a mouse model of allergic inflammation. To assess potentially distinct contributions by Ii chain isoforms to lung immunity, we also compared allergen-induced lung inflammation, eosinophilia, IgE production, and AHR in mice genetically altered to express either p31 Ii or p41 Ii isoform alone. Sole expression of either Ii isoform alone facilitates development of allergen-induced lung inflammation and eosinophilia. However, animals expressing only the p31 Ii isoform exhibit abrogated IgE and AHR responses as compared with p41 Ii mice in this model of allergen-induced lung inflammation, suggesting that realization of complete immunity within the lung requires expression of p41 Ii. These findings reveal a crucial role of Ii and H2-M in controlling the immune response within the lung, and suggest that p31 Ii and p41 Ii manifest nonredundant roles in development of immunity.

Topics: Adjuvants, Immunologic; Animals; Antigens, Differentiation, B-Lymphocyte; Bronchial Hyperreactivity; Cytokines; Histocompatibility Antigens Class II; Immunoglobulin E; Inflammation; Injections, Intraperitoneal; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Protein Isoforms; Pulmonary Eosinophilia

Bronchial asthma represents a severe chronic inflammatory disease with increasing prevalence. The pathogenesis is characterized by complex neuroimmune dysregulation. Although the immunopathogenesis of the disease has been extensively studied, the nature of neuronal dysfunction still remains poorly understood. Recent data indicate that neurotrophins contribute to airway inflammation, broncho-obstruction and airway hyperresponsiveness. Using an established murine model of allergic bronchial asthma, the contribution of the pan-neurotrophin receptor p75(NTR) was defined. This receptor is expressed both in normal and asthmatic lungs and airways. Analysis of p75(NTR-/-) mice, as well as in vivo blocking of p75(NTR), revealed that airway inflammation is to a large extent dependent upon functional receptor expression. Furthermore, neuronal hyperreactivity depends entirely on this receptor. Based on these data, a novel molecular pathway in the neuroimmune pathogenesis of bronchial asthma could be defined.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Immunohistochemistry; Inflammation; Mice; Mice, Inbred C57BL; Mice, Knockout; Nerve Growth Factors; Neurons; Ovalbumin; Receptor, Nerve Growth Factor; Receptors, Nerve Growth Factor; Respiratory System; Substance P

A nonpeptidyl small molecule antagonist, compound A, to nonactivated very late antigen-4 (VLA4) was examined in lung inflammation induced by a single dose of ovalbumin challenge. Compound A presented a good pharmacokinetic property, when given intratracheally, and the blood cells from such pharmacokinetic study showed good receptor occupancy of the compound for approximately 8 hours. Compound A was then tested in an ovalbumin-induced airway inflammation model by intranasal or intravenous route of administration. There was a dose-dependent inhibition of eosinophilia in the bronchiolar lavage fluid, when compound A was given intranasally but not when it was given intravenously. For comparison, antibody to VLA4 and another compound, BIO1211, which reacts only with activated VLA4, were examined in this system. Immunohistochemical analyses of the lung tissue substantiated the findings in the bronchiolar lavage fluid. Specific staining of the major basic protein of eosinophils showed peribronchiolar infiltration of eosinophils. Some of these eosinophils were also positive for nitrotyrosine, suggesting activation of eosinophils in the lung interstitium. There was deposition of major basic protein and nitrotyrosine at the base of the perivascular endothelium, indicative of degranulation of eosinophils in the area. After intranasal treatment with compound A, eosinophils in the lungs and their activation products were substantially decreased, documenting its effectiveness in inhibiting lung inflammation.

Topics: Administration, Inhalation; Animals; Biopsy, Needle; Bronchial Hyperreactivity; Disease Models, Animal; Dose-Response Relationship, Drug; Eosinophils; Female; Immunohistochemistry; Infusions, Intravenous; Integrin alpha4beta1; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia; Probability; Random Allocation; Reference Values; Sensitivity and Specificity

Maternal asthma is a risk factor for development of asthma in children, but mechanisms remain unclear. Offspring of asthmatic mother mice (sensitized and repeatedly exposed to OVA Ag) showed airway hyperresponsiveness and allergic pulmonary inflammation after an intentionally suboptimal OVA sensitization and exposure protocol that had little effect on normal offspring. Similar results were obtained when offspring of OVA-allergic mothers were exposed to an unrelated allergen, casein, indicating that the maternal effect is allergen independent and not transferred by OVA-specific Abs. Premating treatment with neutralizing anti-IL-4 Ab or reduction of maternal allergen exposure abrogated the maternal effect, showing a critical mechanistic role for IL-4 and suggesting an additional benefit of allergen avoidance.

Topics: Aerosols; Allergens; Animals; Animals, Newborn; Antibody Specificity; Asthma; Bronchial Hyperreactivity; Caseins; Disease Susceptibility; Epitopes; Female; Immunoglobulin E; Injections, Intraperitoneal; Male; Maternal Exposure; Mice; Mice, Inbred BALB C; Mothers; Ovalbumin; Respiratory Hypersensitivity

Airway mycoplasma infection may be associated with asthma pathophysiology. However, the direct effects of mycoplasma infection on asthma remain unknown. Using a murine allergic-asthma model, we evaluated the effects of different timing of airway Mycoplasma pneumoniae infection on bronchial hyperresponsiveness (BHR), lung inflammation, and the protein levels of Th1 (gamma interferon [IFN-gamma]) and Th2 (interleukin 4 [IL-4]) cytokines in bronchoalveolar lavage fluid. When mycoplasma infection occurred 3 days before allergen (ovalbumin) sensitization and challenge, the infection reduced the BHR and inflammatory-cell influx into the lung. This was accompanied by a significant induction of Th1 responses (increased IFN-gamma and decreased IL-4 production). Conversely, when mycoplasma infection occurred 2 days after allergen sensitization and challenge, the infection initially caused a temporary reduction of BHR and then increased BHR, lung inflammation, and IL-4 levels. Our data suggest that mycoplasma infection could modulate both physiological and immunological responses in the murine asthma model. Our animal models may also provide a new means to understand the role of infection in asthma pathogenesis and give evidence for the asthma hygiene hypothesis.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Interferon-gamma; Interleukin-4; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia, Mycoplasma

Airway function in health and disease as well as in response to bronchospastic stimuli (i.e., irritants, allergens, and inflammatory mediators) is controlled, in part, by cholinergic muscarinic receptor regulation of smooth muscle. In particular, the dependence of airway smooth muscle contraction/relaxation on heterotrimeric G protein-coupled receptor signaling suggests that these events underlie the responses regulating airway function. Galphaq-containing G proteins are proposed to be a prominent signaling pathway, and the availability of knockout mice deficient of this subunit has allowed for an investigation of its potential role in airway function. Airway responses in Galphaq-deficient mice (activities assessed by both tracheal tension and in vivo lung function measurements) were attenuated relative to wild-type controls. Moreover, ovalbumin sensitization/aerosol challenge of Galphaq-deficient mice also failed to elicit an allergen-induced increase in airway reactivity to methacholine. These findings indicate that cholinergic receptor-mediated responses are dependent on Galphaq-mediated signaling events and identify Galphaq as a potential target of preventative/intervening therapies for lung dysfunction.

Topics: Airway Resistance; Allergens; Animals; Bronchial Hyperreactivity; Bronchoconstrictor Agents; GTP-Binding Protein alpha Subunits, Gq-G11; Heterotrimeric GTP-Binding Proteins; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth; Ovalbumin; Signal Transduction; Trachea

We investigated the development of airway hyperreactivity (AHR) and inflammation in the lungs of nine genetically diverse inbred strains of mice [129/SvIm, A/J, BALB/cJ, BTBR+(T)/tf/tf, CAST/Ei, C3H/HeJ, C57BL/6J, DBA/2J, and FVB/NJ] after sensitization and challenge with ovalbumin (OVA). At 24, 48, and 72 h post-OVA exposure, the severity of AHR and eosinophilic inflammation of the mouse strains ranged from relatively unresponsive to responsive. The severity of the airway eosinophilia of some strains did not clearly correlate with the development of AHR. The temporal presence of T helper type 2 cytokines in lung lavage fluid also varied markedly among the strains. The levels of IL-4 and IL-13 were generally increased in the strains with the highest airway eosinophilia at 24 and 72 h postexposure, respectively; the levels of IL-5 were significantly increased in most of the strains with airway inflammation over the 72-h time period. The differences of physiological and biological responses among the inbred mouse strains after OVA sensitization and challenge support the hypothesis that genetic factors contribute, in part, to the development of allergen-induced airway disease.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Eosinophils; Hypersensitivity; Interleukin-13; Interleukin-4; Interleukin-5; Leukocyte Count; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Inbred DBA; Neutrophils; Ovalbumin; Species Specificity; Th2 Cells

Asthma and mouse models of allergic respiratory inflammation are invariably associated with a pulmonary eosinophilia; however, this association has remained correlative. In this report, a causative relationship between eosinophils and allergen-provoked pathologies was established using eosinophil adoptive transfer. Eosinophils were transferred directly into the lungs of either naive or OVA-treated IL-5(-/-) mice. This strategy resulted in a pulmonary eosinophilia equivalent to that observed in OVA-treated wild-type animals. A concomitant consequence of this eosinophil transfer was an increase in Th2 bronchoalveolar lavage cytokine levels and the restoration of intracellular epithelial mucus in OVA-treated IL-5(-/-) mice equivalent to OVA-treated wild-type levels. Moreover, the transfer also resulted in the development of airway hyperresponsiveness. These pulmonary changes did not occur when eosinophils were transferred into naive IL-5(-/-) mice, eliminating nonspecific consequences of the eosinophil transfer as a possible explanation. Significantly, administration of OVA-treated IL-5(-/-) mice with GK1.5 (anti-CD4) Abs abolished the increases in mucus accumulation and airway hyperresponsiveness following adoptive transfer of eosinophils. Thus, CD4(+) T cell-mediated inflammatory signals as well as signals derived from eosinophils are each necessary, yet alone insufficient, for the development of allergic pulmonary pathology. These data support an expanded view of T cell and eosinophil activities and suggest that eosinophil effector functions impinge directly on lung function.

Topics: Adoptive Transfer; Aerosols; Allergens; Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Disease Models, Animal; Eosinophils; Interleukin-5; Intubation, Intratracheal; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Mucus; Ovalbumin; Pulmonary Eosinophilia; Respiratory Hypersensitivity; Respiratory Mucosa

Asthma is characterized by acute and chronic airway inflammation, and the severity of the airway hyperreactivity correlates with the degree of inflammation. Many of the features of lung inflammation observed in human asthma are reproduced in OVA-sensitized/challenged mice. T lymphocytes, particularly Th2 cells, are critically involved in the genesis of the allergic response to inhaled Ag. In addition to antiapoptotic effects, broad-spectrum caspase inhibitors inhibit T cell activation in vitro. We investigated the effect of the broad-spectrum caspase inhibitor, N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (z-VAD-fmk), on airway inflammation in OVA-sensitized/challenged mice. OVA-sensitized mice treated with z-VAD-fmk immediately before allergen challenge showed marked reduction in inflammatory cell infiltration in the airways and pulmonary blood vessels, mucus production, and Th2 cytokine production. We hypothesized that the caspase inhibitor prevented T cell activation, resulting in the reduction of cytokine production and eosinophil infiltration. Treatment with z-VAD-fmk in vivo prevented subsequent T cell activation ex vivo. We propose that caspase inhibitors may offer a novel therapeutic approach to T cell-dependent inflammatory airway diseases.

Topics: Aerosols; Allergens; Amino Acid Chloromethyl Ketones; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Caspase Inhibitors; Cell Movement; Cysteine Proteinase Inhibitors; Disease Models, Animal; Inflammation; Interleukin-4; Interleukin-5; Intubation, Intratracheal; Leukocytes; Lung; Lymphocyte Activation; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes

(1) To clarify the involvement of Th2 responses in the development of allergen-induced airway remodelling, we investigated the effect of anti-CD4 monoclonal antibody (mAb) and anti-CD8 mAb, and the responses of IL-4 gene-knockout (KO) mice in a murine model of allergic asthma. (2) Mice were immunized twice by intraperitoneal injections of ovalbumin (OA), and exposed to aeroallergen (OA, 1% w v(-1)) for 3 weeks. Twenty-four hours after the final challenge, airway responsiveness to acetylcholine was measured, and bronchoalveolar lavage (BAL) and histological examinations were carried out. (3) Anti-CD4 mAb (1 mg kg(-1)) clearly inhibited allergen-induced increases in airway responsiveness to acetylcholine, the number of eosinophils in BAL fluid, serum OA-specific IgE levels, IL-13 and transforming growth factor-beta1 levels in BAL fluid, and amount of hydroxyproline in the lung by 100, 99, 100, 100, 84, and 60%, respectively. Furthermore, the antibody (1 mg kg(-1)) also attenuated allergen-induced goblet cell hyperplasia in the epithelium and subepithelial fibrosis by 72 and 83%, respectively. In contrast, anti-CD8 mAb (1 mg kg(-1)) showed no effect on each parameter. Furthermore, all these parameters were attenuated in IL-4KO mice by 57, 93, 100, 45, 84 and 60%, and also 72 and 83%, respectively. (4) These findings suggest that Th2 responses play a critical role for the development of allergen-induced airway remodelling, and that the inhibition of Th2 responses, e.g. using anti-CD4 mAb, is a therapeutic approach for the treatment of airway remodelling in asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; CD4 Antigens; CD8 Antigens; Female; Interleukin-4; Lung; Lymphocyte Depletion; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Rats; Spleen; Th1 Cells; Th2 Cells

The link between eosinophils and the development of airway hyperresponsiveness (AHR) in asthma is still controversial. This question was assessed in a murine model of asthma in which we performed a dose ranging study to establish whether the dose of steroid needed to inhibit the eosinophil infiltration correlated with that needed to block AHR.. The sensitised BALB/c mice were dosed with vehicle or dexamethasone (0.01-3 mg/kg) 2 hours before and 6 hours after each challenge (once daily for 6 days) and 2 hours before AHR determination by whole-body plethysmography. At 30 minutes after the AHR to aerosolised methacholine the mice were lavaged and differential white cell counts were determined. Challenging with antigen caused a significant increase in eosinophils in the bronchoalveolar lavage (BAL) fluid and lung tissue, and increased AHR.. Dexamethasone reduced BAL and lung tissue eosinophilia (ED50 values of 0.06 and 0.08 mg/kg, respectively), whereas a higher dose was needed to block AHR (ED50 of 0.32 mg/kg at 3 mg/ml methacholine. Dissociation was observed between the dose of steroid needed to affect AHR in comparison with eosinophilia and suggests that AHR is not a direct consequence of eosinophilia.. This novel pharmacological approach has revealed a clear dissociation between eosinophilia and AHR by using steroids that are the mainstay of asthma therapy. These data suggest that eosinophilia is not associated with AHR and questions the rationale that many pharmaceutical companies are adopting in developing low-molecular-mass compounds that target eosinophil activation/recruitment for the treatment of asthma.

Topics: Administration, Inhalation; Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Dexamethasone; Dose-Response Relationship, Drug; Eosinophilia; Immunization; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin

The prevalence of allergic diseases such as asthma has increased markedly over the past few decades. To evaluate the possible mutual influence of helminth infection and allergy, the combined effects of experimental allergic airway inflammation and infection with Strongyloides venezuelensis on various parasitological and inflammatory indices were evaluated in the rat. A challenge of immunized rats with aerosolized ovalbumin (OVA) resulted in eosinophilic inflammation that peaked 48 h after the challenge and was accompanied by airway hyperresponsiveness (AHR) to an intravenous acetylcholine challenge. S. venezuelensis infection concomitant with an OVA challenge of immunized rats resulted in prolonged pulmonary inflammation with increased eosinophil infiltration in bronchoalveolar lavage fluid but not in the lung tissue. These rats also showed a significant parasite burden reduction, especially during parasite migration through the lungs. However, the fecundity rates of worms that reached the intestine were similar in allergic and nonallergic animals. Despite airway inflammation, the increased responsiveness of the airways in the experimental asthma model was suppressed during parasite migration through the lungs (2 days). In contrast, parasite-induced AHR was unchanged 5 days after infection in immunized and challenged rats. In conclusion, infection with S. venezuelensis interfered with the onset of AHR following an antigen challenge of immunized rats. The ability of parasites to switch off functional airway responses is therapeutically relevant because we may learn from parasites how to modulate lung function and, hence, the AHR characteristic of asthmatic patients.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Eosinophils; Immunization; Immunoglobulin E; Interleukin-10; Lung; Male; Ovalbumin; Rats; Rats, Wistar; Strongyloidiasis

Most infections with respiratory viruses induce Th1 responses characterized by the generation of Th1 and CD8(+) T cells secreting IFN-gamma, which in turn have been shown to inhibit the development of Th2 cells. Therefore, it could be expected that respiratory viral infections mediate protection against asthma. However, the opposite seems to be true, because viral infections are often associated with the exacerbation of asthma. For this reason, we investigated what effect an influenza A (flu) virus infection has on the development of asthma. We found that flu infection 1, 3, 6, or 9 wk before allergen airway challenge resulted in a strong suppression of allergen-induced airway eosinophilia. This effect was associated with strongly reduced numbers of Th2 cells in the airways and was not observed in IFN-gamma- or IL-12 p35-deficient mice. Mice infected with flu virus and immunized with OVA showed decreased IL-5 and increased IFN-gamma, eotaxin/CC chemokine ligand (CCL)11, RANTES/CCL5, and monocyte chemoattractant protein-1/CCL2 levels in the bronchoalveolar lavage fluid, and increased airway hyperreactivity compared with OVA-immunized mice. These results suggest that the flu virus infection reduced airway eosinophilia by inducing Th1 responses, which lead to the inefficient recruitment of Th2 cells into the airways. However, OVA-specific IgE and IgG1 serum levels, blood eosinophilia, and goblet cell metaplasia in the lung were not reduced by the flu infection. Flu virus infection also directly induced AHR and goblet cell metaplasia. Taken together, our results show that flu virus infections can induce, exacerbate, and suppress features of asthmatic disease in mice.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cell Migration Inhibition; Cell Movement; Cells, Cultured; Chemokine CCL11; Chemokine CCL2; Chemokine CCL5; Chemokines, CC; Down-Regulation; Epitopes, T-Lymphocyte; Goblet Cells; Influenza A virus; Interferon-gamma; Interleukin-5; Lung; Lymphocyte Count; Lymphopenia; Metaplasia; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Nippostrongylus; Orthomyxoviridae Infections; Ovalbumin; Peptide Fragments; Pulmonary Eosinophilia; Strongylida Infections; Th2 Cells; Up-Regulation

One of the characteristic features of allergic asthma is recruitment of large numbers of inflammatory cells including eosinophils and Th2 lymphocytes to the lung. This influx of inflammatory cells is thought to be a controlled and coordinated process mediated by chemokines and their receptors. It is thought that distinct, differential expression of chemokine receptors allows selective migration of T cell subtypes in response to the chemokines that bind these receptors. Th2 cells preferentially express CCR8 and migrate selectively to its ligand, CC chemokine ligand (CCL)1. We studied the role of the CCR8 ligand, CCL1, in the specific recruitment of Th2 cells and eosinophils to the lung in a murine model of allergic airway disease. We have demonstrated for the first time that CCL1 is up-regulated in the lung following allergen challenge. Moreover, a neutralizing Ab to CCL1 reduced eosinophil migration to the lung, but had no effect on recruitment of Th2 cells following allergen challenge. In addition, there was no change in airway hyperresponsiveness or levels of Th2 cytokines. In a Th2 cell transfer system of pulmonary inflammation, anti-CCL1 also failed to affect recruitment of Th2 cells to the lung following allergen challenge. Significantly, intratracheal instillation of rCCL1 increased recruitment of eosinophils but not Th2 cells to the lung in allergen-sensitized and -challenged mice. In summary, our results indicate that CCL1 is important for the pulmonary recruitment of eosinophils, rather than allergen-specific Th2 cells, following allergen challenge.

Topics: Adoptive Transfer; Allergens; Animals; Bronchial Hyperreactivity; Chemokine CCL1; Chemokines, CC; Chemotaxis, Leukocyte; Cytokines; Female; Immune Sera; Injections, Intravenous; Intubation, Intratracheal; Ligands; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Plethysmography, Whole Body; Pulmonary Eosinophilia; Respiratory Hypersensitivity; T-Lymphocyte Subsets; Th2 Cells

Asthma is an inflammatory disease of the airways and the current focus in managing asthma is the control of inflammation. In this study, we attempted to investigate the anti-asthmatic potential of a plant derived natural compound, luteolin.. We used a murine model of airway hyperreactivity, which mimicked some of the characteristic features of asthma. Male BALB/c mice (8-9 weeks) were used for this study.. Mice (n = 6) were sensitized by intraperitoneal (i. p.) injection of 10 mg of ovalbumin (OVA) on days 0, 7 and 14 followed by aerosol inhalation (5% OVA) treatments daily beginning from day 19 to day 23. To study its preventive effect, luteolin (0.1, 1.0, and 10 mg/kg body weight; daily) was administered orally during the entire period (0 to 23 day) of sensitization. To study its curative effect, mice were first sensitized and then luteolin (1.0 mg/kg body weight daily) was given orally from day 26 to 32. The airway hyperreactivity, immunoglobulin E (IgE) in the sera, and cytokines (IFN-gamma, IL-4 and IL-5) in the bronchoalveolar lavage fluid (BALF) were measured.. Both during sensitization and after sensitization, luteolin, at a dose of 0.1 mg/kg body weight, significantly modulated OVA-induced airway bronchoconstriction and bronchial hyperreactivity (p < 0.05). Luteolin also reduced OVA-specific IgE levels in the sera, increased interferon gamma (IFN-gamma) levels and decreased the interleukin-4 (IL-4) and interleukin-5 (IL-5) levels in the BALF.. Our study showed that luteolin treatment during and after sensitization significantly attenuated the asthmatic features in experimental mice. Therefore, luteolin could be used either as a lead molecule to identify an effective antiasthma therapy or as a means to identify novel anti-asthma targets.

Topics: Airway Resistance; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Expectorants; Flavonoids; Immunoglobulin E; Interferon-gamma; Interleukin-4; Interleukin-5; Luteolin; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Muscarinic Agonists; Ovalbumin

Since oxidative stress contributes to the pathogenesis of asthma, this study addressed the question whether supplementing the endogenous antioxidant, glutathione (GSH), would alleviate features of allergic asthma in the mouse.. Ovalbumin-sensitized mice received aerosols of the GSH-donors, glutathione-ethyl ester (GSEt) or N-acetylcysteine, before or during respiratory allergen challenges, or during methacholine challenges given one day after the last allergen challenge. Lung GSH levels were measured shortly after allergen or methacholine challenge. In addition, the effect of GSH supplements on airway hyperresponsiveness and inflammatory cell numbers in the airway lumen was assessed.. GSEt decreased allergen-induced airway hyperresponsiveness when given in combination with methacholine. However, when given before or during allergen challenge, both GSH-donors failed to decrease the methacholine-induced airway contractility, change cell numbers in the airway lumen, or increase lung GSH levels. In addition, allergen challenges of sensitized mice did not decrease lung GSH levels.. In contrast to guinea pigs and humans, allergen challenges in mice does not lead to acute oxidative stress.

Topics: Acetylcysteine; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Expectorants; Glutathione; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Muscarinic Agonists; Ovalbumin; Sulfhydryl Compounds

To study the effects of chronic Ag deposition in the airway mucosa on CD4(+) T cell priming and subsequent airway disease, transgenic mice were generated that expressed OVA under the control of the surfactant protein C promoter. CD4 T cells from these mice were tolerant to OVA but this was overcome among spleen CD4 T cells by crossing to OVA-specific DO11.10 TCR-transgenic mice. Lungs from the double-transgenic mice developed lymphocytic infiltrates and modest mucus cell hyperplasia. Infiltrating cells were unaffected by the absence of either Rag-1 or Stat6, although the latter deficiency led to the disappearance of mucus. In the lung of double-transgenic mice, a large number of Ag-specific CD4 T cells expressed CD25 and functioned as regulatory T cells. The CD25(+) CD4 T cells suppressed proliferation of CD25(-) CD4 T cells in vitro and inhibited type 2 immune responses induced by aerosolized Ags in vivo. Despite their ability to suppress allergic type 2 immunity in the airways, however, CD25(+) CD4 regulatory T cells had no effect on the development of bronchial hyperreactivity.

Topics: Administration, Intranasal; Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Movement; Clone Cells; Crosses, Genetic; Down-Regulation; Epitopes, T-Lymphocyte; Homeodomain Proteins; Immune Tolerance; Immunophenotyping; Lung; Lymphocyte Subsets; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Pulmonary Surfactant-Associated Protein C; Receptors, Antigen, T-Cell; Receptors, Interleukin-2; STAT6 Transcription Factor; T-Lymphocyte Subsets; Th2 Cells; Trans-Activators; Transgenes

Tissue eosinophilia probably plays important roles in the pathophysiology of bronchial asthma and allergic disorders; however, this concept was challenged recently by controversial results in mouse models of bronchial asthma treated with anti-IL-5 Ab and the failure of anti-IL-5 therapy in humans. We have now used a unique model, IL-5 transgenic (TG) mice, to address a fundamental question: is airway eosinophilia beneficial or detrimental in the allergic response? After sensitization and challenge with OVA, IL-5 TG mice showed a marked airway eosinophilia. Surprisingly, these IL-5 TG mice showed lower airway reactivity to methacholine. Immunohistochemical analysis of the lungs revealed a marked peribronchial infiltration of eosinophils, but no eosinophil degranulation. In vitro, mouse eosinophils from peritoneal lavage fluids did not produce superoxide anion, but did produce an anti-inflammatory and fibrotic cytokine, TGF-beta 1. Indeed, the TGF-beta 1 levels in bronchoalveolar lavage fluid specimens from IL-5 TG mice directly correlated with airway eosinophilia (r = 0.755). Furthermore, anti-IL-5 treatment of IL-5 TG mice decreased both airway eosinophilia and TGF-beta 1 levels in bronchoalveolar lavage fluids and increased airway reactivity. Thus, in mice, marked eosinophilia prevents the development of airway hyper-reactivity during an allergic response. Overall, the roles of eosinophils in asthma and in animal models need to be addressed carefully for their potentially detrimental and beneficial effects.

Topics: Allergens; Animals; Antibodies, Monoclonal; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Humans; Inflammation; Interleukin-5; Male; Mice; Mice, Inbred C3H; Mice, Transgenic; Ovalbumin; Pulmonary Eosinophilia; Respiratory Hypersensitivity; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transgenes

In this study we have compared the therapeutic effect of the administration of immunostimulatory DNA sequences (ISS) with that of corticosteroids on the resolution of airway inflammation and airway hyperreactivity (AHR) in a mouse model. Mice which had already developed significant levels of eosinophilic airway inflammation 24 h after allergen challenge were then treated with either ISS or corticosteroids, and the effect on AHR and airway inflammation assessed 6 d later. ISS inhibited AHR as effectively as corticosteroids. Combination therapy with ISS and corticosteroids was more effective than monotherapy with either ISS or corticosteroids in inhibiting AHR. In ovalbumin-challenged mice, levels of bronchoalveolar lavage (BAL) eosinophils were significantly reduced with either ISS or corticosteroids. ISS induced significant levels of BAL interferon-gamma, whereas corticosteroids did not induce expression of BAL interferon-gamma. Both ISS and corticosteroids significantly reduced levels of interleukin-5 in BAL, as well as the number of Periodic Acid Schiff-positive airway epithelial cells. Corticosteroids, but not ISS, increased the number of eosinophils in regional mediastinal lymph nodes. Very few apoptotic peribronchial cells were noted following ovalbumin challenge as assessed by TUNEL assay. Corticosteroids, but not ISS, induced an increase in the small number of apoptotic peribronchial cells. The mechanism by which either ISS or corticosteroids inhibit AHR is likely to be mediated by distinct and shared cellular pathways. The combination of the shared and distinct anti-inflammatory pathways may account for the additive effect of ISS and corticosteroids on inhibiting AHR.

Topics: Administration, Inhalation; Adrenal Cortex Hormones; Animals; Anti-Inflammatory Agents; Apoptosis; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Dexamethasone; Disease Models, Animal; DNA; Drug Therapy, Combination; Eosinophils; Female; Inflammation; Injections, Intraperitoneal; Interferon-gamma; Interleukin-5; Lymph Nodes; Lymphocytes; Mice; Mice, Inbred BALB C; Ovalbumin; Pulmonary Eosinophilia; Respiratory Hypersensitivity; Treatment Outcome

Antigen induces murine bronchial hyperreactivity (BHR), inflammation, mucus accumulation, and airway remodeling. To investigate whether leukotrienes (LT) mediate the effects of antigen [ovalbumin (Ova)], we studied 5-lipoxygenase (5-LO) expression in immunized BP2 mice and blocked LT synthesis with the 5-LO inhibitor zileuton or antagonized their effects with receptor antagonists [cysteinyl leukotriene (Cys-LT)-ra MK-571, LY-171883; LTB4-ra PH-163]. Cys-LT content increased in the bronchoalveolar lavage fluid (BALF) as early as 15 min after the intratracheal instillation of Ova. Zileuton inhibited LT release in the BALF and eosinophil recruitment in the lungs, and dose dependently reduced BHR, mucus accumulation, and remodeling, as did the LT-ra. Thus LT, released just after antigen challenge, might constitute the first step in accounting for the effects of Ova. Because mucus accumulation is regulated via the EGF receptor (EGFR), which is also implicated in the effects of LT, we studied this pathway with AG-1478, an EGFR tyrosine kinase inhibitor given at 0.5, 4, and 20 mg/kg. AG-1478 inhibited BHR, inflammation, and lung remodeling induced by Ova or by molecules themselves generated by Ova, such as LT, IL-13, and monocyte chemoattractant protein-1, which promote identical effects, suggesting the involvement of the EGFR pathway in the asthma-like syndrome observed.

Topics: Animals; Arachidonate 5-Lipoxygenase; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Division; Chemokines; Collagen; Cytokines; Enzyme Inhibitors; Eosinophils; ErbB Receptors; Leukotrienes; Lung; Male; Mice; Mice, Inbred Strains; Mucus; Muscle, Smooth; Ovalbumin; Protein-Tyrosine Kinases; Quinazolines; RNA, Messenger; Tyrphostins

Alpha-melanocyte-stimulating hormone (alpha-MSH) is a neuropeptide controlling melanogenesis in pigmentary cells. In addition, its potent immunomodulatory and immunosuppressive activity has been recently described in cutaneous inflammatory disorders. Whether alpha-MSH is also produced in the lung and might play a role in the pathogenesis of inflammatory lung conditions, including allergic bronchial asthma, is unknown. Production and functional role of alpha-MSH were investigated in a murine model of allergic airway inflammation. alpha-MSH production was detected in bronchoalveolar lavage fluids. Although aerosol challenges stimulate alpha-MSH production in nonsensitized mice, this rapid and marked stimulation was absent in allergic animals. Treatment of allergic mice with alpha-MSH resulted in suppression of airway inflammation. These effects were mediated via IL-10 production, because IL-10 knockout mice were resistant to alpha-MSH treatment. This study provides evidence for a novel function of alpha-MSH linking neuroimmune functions in allergic airway inflammation.

Topics: Allergens; alpha-MSH; Animals; Antibody Formation; Bronchial Hyperreactivity; Cell Movement; Down-Regulation; Female; Inflammation Mediators; Injections, Intraperitoneal; Injections, Intravenous; Interleukin-10; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Respiratory Hypersensitivity

Airway hyperresponsiveness (AHR) is the main feature of allergic subjects/animals, and its underlying mechanism is not clear. We explored whether antigen-induced AHR is associated with cytokine generation, inflammatory cell infiltration, and/or remodeling of airway smooth muscle. Guinea pigs were divided into three groups: control-1, control-2, and ovalbumin (OA). Animals in the control-1 group were not sensitized, while those in the control-2 and the OA group were sensitized with OA. Forty to forty-two days after the initial sensitization or equivalent time, animals in the control-2 group inhaled saline aerosol and those in the OA group inhaled OA aerosol for 30 min. Twenty-four h after OA challenge or equivalent time, animals in each group were further divided into two subgroups: methacholine and hyperventilation. Functional tests were carried out before and after the methacholine or hyperventilation treatment. Immediately after the functional study, bronchoalveolar lavage fluid was collected for determination of inflammatory cells and tumor necrosis factor-alpha (TNF-alpha. The trachea was then removed to determine smooth muscle mass. In both the methacholine and hyperventilation subgroups, significantly more severe airway constriction was found in the OA group, indicating OA-induced AHR. Eosinophil accumulation increased in the control-2 group and this increase was further augmented in the OA group. In addition, TNF-alpha level and smooth muscle mass significantly increased in the OA group. These results suggest that OA challenge-induced AHR is associated with increases in TNF-alpha level, cellular infiltration, and airway smooth muscle mass.

Topics: Animals; Antigens; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Guinea Pigs; Muscle, Smooth; Ovalbumin; Tumor Necrosis Factor-alpha

Bakery workers may develop IgE-mediated allergy to liquid and aerosolized hen's egg proteins that are commonly used in the baking and confectionery industries.. We studied four bakery workers who had work-related allergic respiratory symptoms upon exposure to egg aerosols. The causative role of egg proteins in their respiratory symptoms was investigated by immunologic and specific inhalation challenge (SIC) tests.. Skin prick tests to egg white extract and to lysozyme gave positives responses in all the subjects, to ovalbumin in two, to ovomucoid in one and to egg yolk in two subjects. They were also sensitized to wheat, rye and barley flours. Specific IgE determinations to egg white were positive in all patients, to lysozyme in two, to ovalbumin in three, to ovomucoid in two and to egg yolk in two of them. Methacholine inhalation tests revealed bronchial hyperresponsiveness in all workers (PC20 < 16 mg/ml). SICs were performed with aqueous extracts of lysozyme (n = 4), ovalbumin (n = 2) and ovomucoid (n = 1), which elicited isolated early asthmatic reactions in all subjects. Double-blind, placebo-controlled, oral challenge tests with raw egg white were positive in three subjects.. These bakery workers had developed IgE-mediated occupational asthma to hen's egg white proteins.

Topics: Adult; Allergens; Antibody Specificity; Asthma; Biomarkers; Bronchial Hyperreactivity; Bronchial Provocation Tests; Egg Hypersensitivity; Egg Proteins; Flour; Food Industry; Forced Expiratory Volume; Humans; Immunoglobulin E; Inhalation Exposure; Male; Middle Aged; Muramidase; Occupational Exposure; Ovalbumin; Skin Tests

Asthma results from an intrapulmonary allergen-driven Th2 response and is characterized by intermittent airway obstruction, airway hyperreactivity, and airway inflammation. An inverse association between allergic asthma and microbial infections has been observed. Microbial infections could prevent allergic responses by inducing the secretion of the type 1 cytokines, IL-12 and IFN-gamma. In this study, we examined whether administration of bacterial LPS, a prototypic bacterial product that activates innate immune cells via the Toll-like receptor 4 (TLR4) could suppress early and late allergic responses in a murine model of asthma. We report that LPS administration suppresses the IgE-mediated and mast cell-dependent passive cutaneous anaphylaxis, pulmonary inflammation, airway eosinophilia, mucus production, and airway hyperactivity. The suppression of asthma-like responses was not due to Th1 shift as it persisted in IL-12(-/-) or IFN-gamma(-/-) mice. However, the suppressive effect of LPS was not observed in TLR4- or NO synthase 2-deficient mice. Our findings demonstrate, for the first time, that LPS suppresses Th2 responses in vivo via the TLR4-dependent pathway that triggers NO synthase 2 activity.

Topics: Administration, Inhalation; Allergens; Animals; Anti-Allergic Agents; Asthma; Bronchi; Bronchial Hyperreactivity; Cytokines; Enzyme Activation; Immunity, Innate; Inflammation; Injections, Intravenous; Interferon-gamma; Interleukin-12; Lipopolysaccharides; Lung; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Inbred C57BL; Mice, Knockout; Mucus; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Ovalbumin; Passive Cutaneous Anaphylaxis; Receptors, Cell Surface; Salmonella enterica; Signal Transduction; Th2 Cells; Toll-Like Receptor 4; Toll-Like Receptors

Histamine elicits many features of immediate hypersensitivity reactions. Recent evidence indicates that H1 receptors modulate immune responses to antigens. Desloratadine (DL), a new, long-acting, H1 receptor antagonist, has both a potent antihistaminic function and anti-inflammatory properties.. We sought to evaluate the effect of DL on allergic-airway responses in mice after inhalation of the naturally occurring aeroallergen Aspergillus fumigatus (Af ) and to examine the effects of DL on specific immune responses to a defined protein antigen with the use of an ovalbumin (OVA) model of asthma.. Mice were subjected either to repeated, intranasal application of Af extract or to intraperitoneal immunization with OVA, followed by inhalation challenge. DL or a control fluid was given daily throughout the sensitization process. Immunoglobulin E (IgE) levels, bronchoalveolar lavage-fluid cytokines and cytology, lung histology, and physiologic responses to methacholine were assessed in the allergen-treated mice. Anti-OVA IgE responses and OVA-driven T-cell cytokine production were examined.. Treatment with DL did not impair IgE production but did inhibit bronchial inflammation and bronchial hyperresponsiveness in both Af- and OVA-treated mice. This inhibition required that DL be administered concurrently with allergen sensitization, indicating that the attenuation of bronchial hyperresponsiveness and inflammation was not caused by anticholinergic receptor effects. OVA-responsive T cells from DL-treated mice exhibited depressed production of IL-4, IL-5, and IL-13 and normal amounts of interferon-gamma. The amounts of IL-5 and IL-13 were also diminished in the bronchoalveolar lavage fluid.. DL, given at the time of exposure to the allergen, inhibits T(H)2 responses, the induction of allergic pulmonary inflammation, and bronchial hyperresponsiveness. These results suggest that DL or similar agents given during times of antigen exposure might alter disease progression in patients with respiratory allergy.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Dose-Response Relationship, Drug; Female; Histamine H1 Antagonists, Non-Sedating; Immunoglobulin E; Inflammation; Loratadine; Mice; Mice, Inbred BALB C; Ovalbumin; Receptors, Histamine H1; T-Lymphocytes

Murine models of acute atopic asthma may be inadequate to study the effects of recurrent exposure to inhaled allergens, such as the epithelial changes seen in asthmatic patients. We developed a murine model in which chronic airway inflammation is maintained by repeated allergen [ovalbumin (OVA)] inhalation; using this model, we examined the response to mucosal administration of CpG DNA (oligonucleotides) and specific antigen immunotherapy. Mice repeatedly exposed to OVA developed significantly greater airway hyperresponsiveness and goblet cell hyperplasia, but not airway eosinophilia, compared with those exposed only twice. CpG-based immunotherapy significantly reversed both acute and chronic markers of inflammation as well as airway hyperresponsiveness. We further examined the effect of mucosal immunotherapy on the response to a second, unrelated antigen. Mice sensitized to both OVA and schistosome eggs, challenged with inhaled OVA, and then treated with OVA-directed immunotherapy demonstrated significant reduction of airway hyperresponsiveness and a moderate reduction in eosinophilia, after inhalation challenge with schistosome egg antigens. In this model, immunotherapy treatment reduced bronchoalveolar lavage (BAL) levels of Th2 cytokines (IL-4, IL-5, IL-13, and IL-10) without changing BAL IFN-gamma. Antigen recall responses of splenocytes from these mice demonstrated an antigen-specific (OVA) enhanced release of IL-10 from splenocytes of treated mice. These results suggest that CpG DNA may provide the basis for a novel form of immunotherapy of allergic asthma. Both antigen-specific and, to a lesser extent, antigen-nonspecific responses to mucosal administration of CpG DNA are seen.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Base Sequence; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Dinucleoside Phosphates; Disease Models, Animal; Female; Immunity, Mucosal; Immunotherapy; Mice; Mice, Inbred C57BL; Oligodeoxyribonucleotides; Ovalbumin

We found that continuous eosinophilic inflammation after repeated antigen instillation into the nose was observed only in A/J mice, not in three other strains. Histologic analysis of tissues from A/J mice revealed features typical of airway remodeling, i.e., airway wall thickening and increased collagen depositions were observed after 12 weeks' antigen exposure. Persistent airway hyperresponsiveness (AHR) was observed in chronically antigen-exposed A/J mice. Eosinophilic inflammation, collagen deposition, and airway wall thickening were all less marked in BALB/c mice than in A/J mice, and no AHR was observed in the former strain. In C57BL/6 and C3H/HeJ mice, eosinophilic inflammation, airway wall thickening, and AHR were not observed at all, although slightly increased collagen deposition was observed. Thus, we found that these changes were strain-dependent. On the other hand, in A/J mice inhalational antigen challenge after ovalbumin/alum immunization led only to a transient increase in eosinophils and to less airway wall thickening, indicating the importance of the protocol used. Use of A/J mice and giving antigen by instillation via the nose is to be recommended for studies of the mechanisms underlying asthma. In particular, useful qualitative and quantitative information relating to the structural and histologic changes in the lungs may be obtainable using this model.

Topics: Administration, Intranasal; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophils; Immunoglobulin E; Inflammation; Inhalation Exposure; Instillation, Drug; Lymphocytes; Mice; Mice, Inbred A; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Inbred C57BL; Ovalbumin; Pulmonary Eosinophilia; Time Factors

To investigate whether the inhibition of muscarinic M(2) receptors results in the enhancement of reflex bronchoconstriction under airway hyperresponsiveness, we evaluated the effects of muscarinic antagonists with or without M(2) antagonist activity on methacholine (MCh)- and SO(2)-induced airway responses in ovalbumin (OVA)-sensitized and -challenged mice. In this model, similar airway hyperresponsiveness to MCh (12 mg/ml) was observed on Days 31 and 37 (2.2-fold and 2.7-fold, respectively). However, airway hyperresponsiveness to SO(2) (0.05 l/min) on Day 37 was less than that on Day 31 (4.0- and 2.7-fold on Days 31 and 37), indicating reflex bronchoconstriction was enhanced on Day 31 in comparison to Day 37. Ipratropium (0.03 - 0.3 mg/ml, inhalation) and Compound A (0.1 - 3 mg/kg, p.o.) inhibited MCh-induced responses on Days 31 and 37. Although ipratropium (0.03 - 1 mg/ml) dose-dependently inhibited SO(2)-induced responses on Day 31, ipratropium at a dose of 0.1 mg/ml significantly increased SO(2)-induced responses on Day 37 (162.2% of the corresponding control). On the other hand, Compound A (0.03 - 0.3 mg/kg, p.o.) inhibited SO(2)-induced responses without any increases on Days 31 and 37. These results suggest that two different conditions of reflex bronchoconstriction are presented in this model: 1) SO(2)-induced responses are enhanced by dysfunctional M(2) receptors on Day 31; 2) the dysfunctional M(2) receptors are partially restored on Day 37. In addition, the inhibition of the restored M(2) receptors further enhance reflex bronchoconstriction.

Topics: Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Cholinergic Fibers; Female; Immunization; Methacholine Chloride; Mice; Mice, Inbred BALB C; Muscarinic Antagonists; Ovalbumin; Receptor, Muscarinic M2

Airway hyperreactivity (AHR), eosinophilic inflammation with a Th2-type cytokine profile, and specific Th2-mediated IgE production characterize allergic asthma. In this paper, we show that OVA-immunized Jalpha18(-/-) mice, which are exclusively deficient in the invariant Valpha14(+) (iValpha14), CD1d-restricted NKT cells, exhibit impaired AHR and airway eosinophilia, decreased IL-4 and IL-5 production in bronchoalveolar lavage fluid, and reduced OVA-specific IgE compared with wild-type (WT) littermates. Adoptive transfer of WT iValpha14 NKT cells fully reconstitutes the capacity of Jalpha18(-/-) mice to develop allergic asthma. Also, specific tetramer staining shows that OVA-immunized WT mice have activated (CD69(+)) iValpha14 NKT cells. Importantly, anti-CD1d mAb treatment blocked the ability of iValpha14 T cells to amplify eosinophil recruitment to airways, and both Th2 cytokine and IgE production following OVA challenge. In conclusion, these findings clearly demonstrate that iValpha14 NKT cells are required to participate in allergen-induced Th2 airway inflammation through a CD1d-dependent mechanism.

Topics: Administration, Intranasal; Adoptive Transfer; Allergens; Animals; Antigens, CD1; Antigens, CD1d; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Down-Regulation; Eosinophilia; Immunodominant Epitopes; Immunoglobulin E; Inflammation; Interleukin-4; Interleukin-5; Killer Cells, Natural; Lung; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Receptors, Antigen, T-Cell, alpha-beta; T-Lymphocyte Subsets; Th2 Cells

The Vgamma4(+) pulmonary subset of gammadelta T cells regulates innate airway responsiveness in the absence of alphabeta T cells. We now have examined the same subset in a model of allergic airway disease, OVA-sensitized and challenged mice that exhibit Th2 responses, pulmonary inflammation, and airway hyperreactivity (AHR). In sensitized mice, Vgamma4(+) cells preferentially increased in number following airway challenge. Depletion of Vgamma4(+) cells before the challenge substantially increased AHR in these mice, but had no effect on airway responsiveness in normal, nonchallenged mice. Depletion of Vgamma1(+) cells had no effect on AHR, and depletion of all TCR-delta(+) cells was no more effective than depletion of Vgamma4(+) cells alone. Adoptively transferred pulmonary lymphocytes containing Vgamma4(+) cells inhibited AHR, but lost this ability when Vgamma4(+) cells were depleted, indicating that these cells actively suppress AHR. Eosinophilic infiltration of the lung and airways, or goblet cell hyperplasia, was not affected by depletion of Vgamma4(+) cells, although cytokine-producing alphabeta T cells in the lung increased. These findings establish Vgamma4(+) gammadelta T cells as negative regulators of AHR and show that their regulatory effect bypasses much of the allergic inflammatory response coincident with AHR.

Topics: Adoptive Transfer; Animals; Bronchial Hyperreactivity; Cytokines; Female; Goblet Cells; Hyperplasia; Immunization; Injections, Intraperitoneal; Lymphocyte Count; Lymphocyte Depletion; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Pulmonary Alveoli; Pulmonary Eosinophilia; Receptors, Antigen, T-Cell, alpha-beta; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocyte Subsets; Up-Regulation

To study the effects of polysaccharides of cultured Cryptoporus volvatus(CVPS) on airway hyperresponsiveness of ovalbumin-sensitized rats and to evaluate their mechanisms.. Polysaccharides A, B (5mg/kg, 20mg/kg) and ketotifen(5mg/kg) or vehicle(same volume of saline) were administrated orally for 10 days in ovalbumin -sensitized rats, methacholine bronchial provocation tests were performed to determine airway hyperresponsiveness. Bronchoalveolar lavage fluid (BALF) and peritoneal lavage fluid were prepared after the animals were challenged by nebulized antigen. The differential white cell count in BALF,and the degranulated mast cell count as well as differential white cell count in peritoneal lavage fluid were performed.. Polysaccharides markedly inhibited the increased lung resistance and the decreased lung compliance induced by antigen challenge,significantly reduced total cell counts and absolute eosinophil counts in BALF(P<0.05); polysaccharides B was more effective than polysaccharides A. They also inhibited recruitment of inflammatory cells in peritoneal lavage fluid and inhibited the allergen-induced mast cell degranulation.. Polysaccharides of CVPS inhibit airway hyperresponsiveness by stabilizing mast cell membranes and reducing infiltration and chemotaxis of eosinophils and may be developed as a potential anti-asthmatic drug.

Topics: Animals; Anti-Asthmatic Agents; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Degranulation; Male; Mast Cells; Ovalbumin; Polyporaceae; Polysaccharides; Rats; Rats, Sprague-Dawley

Understanding the mechanisms of airway remodeling in chronic allergic conditions such as asthma is increasingly dependent on the use of animal models. Techniques for quantifying structural changes are required that are reproducible and responsive and that can be applied to different staining techniques in both human and animal airway tissues. Here, we characterize a morphometric technique to quantify changes in extracellular matrix and contractile tissue as two indices of airway remodeling in mice. Specific aims were to establish the optimum projection beneath the epithelium to assess remodeling changes and to determine whether such changes are reproducible within different areas of the lung. Finally, based on the variance within measurements, we calculated sample size requirements for research applications of this technique. BALB/c mice were sensitized to ovalbumin and studied after chronic allergen challenge. Lungs were formalin fixed and sectioned were then assayed for extracellular matrix or contractile tissue using morphometric/colorimetric techniques. In this model, the optimum projected distance to measure changes in extracellular matrix or contractile tissue was 20 micro m beneath the epithelium; projecting beyond this depth resulted in decreased ability to detect allergen-induced changes (signal) because of increased irrelevant staining of surrounding parenchymal tissue (noise). The technique was responsive, because an allergen-induced signal was detected in all airway sections and all lung regions studied (p < 0.05). The power of this analysis was such that allergen-induced changes can be reliably (>80% power) detected using 8 to 10 mice. This morphometric technique provides a valid and objective method to assess structural changes in the airways of mice after chronic allergen exposure.

Topics: Actins; Allergens; Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Chronic Disease; Disease Models, Animal; Extracellular Matrix; Female; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Pathology; Reproducibility of Results; Respiratory Mucosa; Sample Size; Specific Pathogen-Free Organisms

Sex hormones might affect the severity and evolution of bronchial asthma. From existing literature, there exists, however, no convincing evidence for either exacerbation or improvement of allergic symptoms by progesterone.. This study was aimed to explore the effect of exogenously administered progesterone in a mouse model of allergic asthma.. BALB/c mice were sensitized to ovalbumin (OVA) by intraperitoneal injections with OVA followed by chronic inhalation of nebulized OVA or physiologic saline (Sal). Medroxyprogesterone acetate or placebo was instilled daily into the oesophagus before and during the inhalatory OVA challenge phase.. Progesterone worsened allergic airway inflammation in OVA-challenged mice, as evidenced by enhanced bronchial responsiveness to inhaled metacholine and increased bronchial eosinophilia. Elevated airway eosinophilia corresponded with higher bronchial and systemic IL-5 levels in the progesterone group. The ratio of IL-4/IFN-gamma levels in bronchoalveolar lavage fluid and numbers of eosinophil colony-forming units in the bone marrow were also elevated in the latter group. Progesterone, however, did not influence allergen-specific IgE production, nor did it affect bronchial responses in Sal-challenged mice.. Our data show that exogenously administered progesterone aggravates the phenotype of eosinophilic airway inflammation in mice by enhancing systemic IL-5 production. Progesterone also increases bronchial hyper-reactivity.

Topics: Animals; Asthma; Bone Marrow; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Eosinophilia; Immunoglobulin E; Male; Medroxyprogesterone Acetate; Mice; Mice, Inbred BALB C; Ovalbumin

We have reported previously that HIV-TAT-dominant negative (dn) Ras inhibits eosinophil adhesion to ICAM-1 after activation by IL-5 and eotaxin. In this study, we evaluated the role of Ras in Ag-induced airway inflammation and hyperresponsiveness by i.p. administration into mice of dnRas, which was fused to an HIV-TAT protein transduction domain (TAT-dnRas). Uptake of TAT-dnRas (t(1/2) = 12 h) was demonstrated in leukocytes after i.p. administration. OVA-sensitization significantly increased eosinophil and lymphocyte numbers in bronchoalveolar lavage fluid 24 h after final challenge. Treatment of animals with 3-10 mg/kg TAT-dnRas blocked the migration of eosinophils from 464 +/- 91 x 10(3)/ml to 288 +/- 79 x 10(3)/ml with 3 mg/kg of TAT-dnRas (p < 0.05), and further decreased to 116 +/- 63 x 10(3)/ml after 10 mg/kg TAT-dnRas (p < 0.01). Histological examination demonstrated that inflammatory cell infiltration (largely eosinophils and mononuclear cells) and mucin production around the airways caused by OVA were blocked by TAT-dnRas. OVA challenge also caused airway hyperresponsiveness to methacholine, which was dose dependently blocked by treatment with TAT-dnRas. TAT-dnRas also blocked Ag-induced IL-4 and IL-5, but not IFN-gamma, production in lung tissue. Intranasal administration of IL-5 caused eosinophil migration into the airway lumen, which was attenuated by pretreatment with TAT-dnRas. By contrast, TAT-green fluorescent protein or dnRas lacking the TAT protein transduction domain did not block airway inflammation, cytokine production, or airway hyperresponsiveness. We conclude that Ras mediates Th2 cytokine production, airway inflammation, and airway hyperresponsiveness in immune-sensitized mice.

Topics: Administration, Intranasal; Animals; Antigens; Bronchial Hyperreactivity; Cell Migration Inhibition; Cytokines; Eosinophils; Gene Products, tat; HIV; Humans; Inflammation Mediators; Injections, Intraperitoneal; Interleukin-5; Kinetics; Lung; Lymphocytes; Male; Mice; Mice, Inbred C57BL; Mucus; Ovalbumin; ras Proteins; Recombinant Fusion Proteins; tat Gene Products, Human Immunodeficiency Virus; Th2 Cells; Transduction, Genetic

Few peribronchial mast cells are noted either in the lungs of naive mice or in the lungs of OVA-sensitized mice challenged acutely with OVA by inhalation. In this study, we demonstrate that OVA-sensitized mice exposed to repetitive OVA inhalation for 1-6 mo have a significant accumulation of peribronchial mast cells. This accumulation of peribronchial mast cells is associated with increased expression of the Th2 cell-derived mast cell growth factors, including IL-4 and IL-9, but not with the non-Th2 cell-derived mast cell growth factor, stem cell factor. Pretreating mice with immunostimulatory sequences (ISS) of DNA containing a CpG motif significantly inhibited the accumulation of peribronchial mast cells and the expression of IL-4 and IL-9. To determine whether mast cells express Toll-like receptor-9 (TLR-9; the receptor for ISS), TLR-9 expression by mouse bone marrow-derived mast cells (MBMMCs) was assessed by RT-PCR. MBMMCs strongly expressed TLR-9 and bound rhodamine-labeled ISS. However, incubation of MBMMCs with ISS in vitro neither inhibited MBMMC proliferation nor inhibited Ag/IgE-mediated MBMMC degranulation, but they did induce IL-6. Overall these studies demonstrate that mice exposed to repetitive OVA challenge, but not acute OVA challenge, have an accumulation of peribronchial mast cells and express increased levels of mast cell growth factors in the lung. Although mast cells express TLR-9, ISS does not directly inhibit mast cell proliferation in vitro, suggesting that ISS inhibits accumulation of peribronchial mast cells in vivo by indirect mechanism(s), which include inhibiting the lung expression of Th2 cell-derived mast cell growth factors.

Topics: Adjuvants, Immunologic; Administration, Intranasal; Allergens; Animals; Bone Marrow Cells; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Cell Aggregation; Cell Count; Cell Division; Chronic Disease; CpG Islands; Disease Models, Animal; DNA; DNA-Binding Proteins; Drug Administration Schedule; Female; Fluorescent Dyes; Growth Inhibitors; Immunoglobulin E; Inflammation; Injections, Subcutaneous; Interleukin-4; Interleukin-9; Lung; Mast Cells; Methacholine Chloride; Mice; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Ovalbumin; Receptors, Cell Surface; Rhodamines; Toll-Like Receptor 9

Chemokines play a key role in the recruitment of activated CD4(+) T cells and eosinophils into the lungs in animal models of airway inflammation. Inhibition of inflammation by N-terminally modified chemokines is well-documented in several models but is often reported with limited dose regimens. We have evaluated the effects of doses ranging from 10 ng to 100 micro g of two CC chemokine receptor antagonists, Met-RANTES/CC chemokine ligand 5 (CCL5) and aminooxypentane-RANTES/CCL5, in preventing inflammation in the OVA-sensitized murine model of human asthma. In the human system, aminooxypentane-RANTES/CCL5 is a full agonist of CCR5, but in the murine system neither variant is able to induce cellular recruitment. Both antagonists showed an inverse bell-shaped inhibition of cellular infiltration into the airways and mucus production in the lungs following allergen provocation. The loss of inhibition at higher doses did not appear to be due to partial agonist activity because neither variant showed activity in recruiting cells into the peritoneal cavity at these doses. Surprisingly, neither was able to bind to the major CCR expressed on eosinophils, CCR3. However, significant inhibition of eosinophil recruitment was observed. Both analogues retained high affinity binding for murine CCR1 and murine CCR5. Their ability to antagonize CCR1 and CCR5 but not CCR3 was confirmed by their ability to prevent RANTES/CCL5 and macrophage inflammatory protein-1beta/CCL4 recruitment in vitro and in vivo, while they had no effect on that induced by eotaxin/CCL11. These results suggest that CCR1 and/or CCR5 may be potential targets for asthma therapy.

Topics: Allergens; Animals; Binding, Competitive; Bronchial Hyperreactivity; Cell Line; Cell Migration Inhibition; Cell Movement; Chemokine CCL5; CHO Cells; Cricetinae; Down-Regulation; Female; Humans; Inflammation; Lung; Lymph Nodes; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Peritoneal Cavity; Protein Structure, Tertiary; Receptors, CCR3; Receptors, CCR5; Receptors, Chemokine; Recombinant Proteins

Respiratory syncytial virus (RSV) bronchiolitis in early life can lead to changes in airway function, but there are likely additional predisposing factors, such as prior allergen exposure, determining which children develop wheezing and asthma.. To define the effects of prior airway exposure to sensitizing allergen on the development of airway inflammation and hyperresponsiveness (AHR) to subsequent RSV infection.. BALB/c mice were exposed to ovalbumin or PBS exclusively through the airways and subsequently infected with RSV or sham-inoculated. AHR, lung inflammation, and the frequency of cytokine-producing T lymphocytes in the lung were determined.. In PBS-exposed mice, RSV infection induced AHR and an increased proportion of TH1-type (IFN-gamma and IL-12) cytokine-producing cells in the lungs. However, in mice previously exposed to ovalbumin through the airways and subsequently infected with RSV, the degree of AHR was significantly increased and was associated with an increased proportion of TH2 (IL-4, IL-5) cytokine-producing T lymphocytes. This response was also associated with an increased accumulation of eosinophils, neutrophils, and CD8+ T cells in the lungs.. These data suggest that prior airway exposure to allergen may predispose sensitized hosts to a greater degree of altered airway function upon subsequent respiratory viral infection.

Topics: Administration, Inhalation; Animals; Bronchial Hyperreactivity; Cytokines; Female; Lung; Lymphocytes; Mice; Mice, Inbred BALB C; Ovalbumin; Pneumonia, Viral; Respiratory Syncytial Virus Infections; Respiratory System

The aim of this study was to investigate the role of immunoglobulin E (IgE) in the late phase reaction (LPR) of murine experimental asthma. Our model consisted of an implant of DNP-conjugated, heat-coagulated hen's egg white (DNP-EWI), followed 14 days later by an intratracheal challenge with aggregated DNP-ovalbumin. Airway inflammation was analyzed 48 h after challenge and compared with a similarly immunized group of mice with highly suppressed humoral response due to anti-micro and anti-delta antibody treatment. Total number of cells in the bronchoalveolar lavage (BAL) (with predominance of eosinophils) and EPO activity in the lung homogenate were increased in the DNP-EWI-immunized group compared with immunosuppressed or nonimmunized mice. However, the cellular infiltration and EPO activity observed in the immunosuppressed group were still significantly above those obtained in the nonimmunized group, indicating that inhibition of antibody production did not completely prevent the inflammatory manifestations in BAL and lung. Airway hyperresponsiveness to methacoline was obtained in DNP-EWI-immunized mice, but the respiratory mechanical parameters returned to normal levels in the immunosuppressed group. When these mice were reconstituted with monoclonal anti-DNP antibodies, only IgE, but not IgG1, restored lung inflammation and decreased the conductance of the respiratory system, therefore, increasing hyperresponsiveness. These results indicate that antibodies are not essential for induction of LPR in the lung. However, IgE enhances pulmonary inflammation and hyperresponsiveness.

Topics: Animals; Antibody Formation; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Dinitrophenols; Egg White; Eosinophil Peroxidase; Eosinophils; Immunoglobulin E; Immunoglobulins; Inflammation; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Peroxidases

The most important pathological feature of asthma is airway inflammation, which results in airway hyper-responsiveness. We hypothesized that excessive oxidation is likely to contribute to airway inflammation in asthma. The aim of this study was to evaluate the effects of both acute exposure and a 30-day administration of ascorbic acid (AA), which has an antioxidant effect, on airway responsiveness in sensitized guinea pigs.. Guinea pigs sensitized to ovalbumin (OA), were given drinking water without AA (group 2) or with AA (group 3). The responses of tracheal chains of control animals (group 1) and both groups of sensitized guinea pigs (n = 10, for all groups) to cumulative concentrations of methacholine were measured, and the effective concentrations of methacholine causing 50% of maximum response (EC50 M) were obtained. The response of tracheal chains to 0.1% OA, relative to contraction obtained with 10 micro mol/L methacholine, was also measured. The tracheal responses to methacholine and OA were measured on tissues both incubated and not incubated with AA.. The tracheal responses of group 2 tissues were significantly greater than those of groups 1 and 3 to both OA and methacholine (P < 0.05). There were no significant differences in tracheal responses to OA and methacholine between groups 1 and 3. Acute incubation of tissues caused a reduction of tracheal response to methacholine in all groups, but this was only significantly differ-ent for group 3 (P < 0.05). Acute incubation of tissues did not change tracheal response to OA significantly.. These results showed that although short-term administration of AA had no major effect on tracheal responsiveness among sensitized animals, 30-day administration of AA could lead to a decrease in airway responsiveness of sensitized guinea pigs to both OA and methacholine.

Topics: Analysis of Variance; Animals; Antioxidants; Ascorbic Acid; Asthma; Bronchial Hyperreactivity; Drug Administration Schedule; Guinea Pigs; Male; Methacholine Chloride; Ovalbumin; Trachea

The pathogenesis of human asthma and the development of key features of pulmonary allergy in mouse models has been critically linked to IL-13. Analyses of the receptor components employed by IL-13 have shown that delivery of this cytokine to the airways of naive IL-4Ralpha gene targeted (IL-4Ralpha(-/-)) mice fails to induce disease, suggesting that this membrane protein is critical for transducing IL-13-mediated responses. The current study demonstrates that, in contrast to naive mice, T helper 2 bias, airways hyperreactivity (AHR) and tissue eosinophilia develop in Ovalbumin-sensitized IL-4Ralpha(-/-) mice and that these responses can be inhibited by the IL-13 antagonist sIL-13Ralpha2Fc. Therefore, antigen stimulation induces an IL-13-regulated response that is independent of IL-4Ralpha. To determine the role of IL-5 and eosinophils in the development of disease in antigen-exposed IL-4Ralpha(-/-) mice, pulmonary allergy was examined in mice deficient in both factors. IL-4Ralpha/IL-5(-/-) mice were significantly defective in their ability to produce IL-13 and failed to develop AHR, suggesting that IL-5 indirectly regulates AHR in allergic IL-4Ralpha(-/-) mice by an IL-13-dependent mechanism. Collectively, these results demonstrate that IL-13-dependent processes regulating the development of AHR and T helper bias persist in the in the lungs of allergic IL-4Ralpha(-/-) mice.

Topics: Animals; Bronchial Hyperreactivity; Eosinophilia; Interferon-gamma; Interleukin-13; Interleukin-13 Receptor alpha1 Subunit; Interleukin-5; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Protein Subunits; Receptors, Interleukin; Receptors, Interleukin-13; Receptors, Interleukin-4; Th1 Cells; Th2 Cells

Indoor formaldehyde (FA) might worsen allergies and be an underlying factor for the increasing incidence and severity of asthma; the exact mechanism, however, remains unclear.. The present study examined the effects of repeated exposure to FA on methacholine- and antigen-induced bronchoconstriction in guinea-pigs in vivo.. First, non-sensitized guinea-pigs were transnasally treated with 0.1 or 1.0% FA or saline three times a week for 6 weeks, and increasing concentrations of methacholine (50, 100, and 200 microg/mL) were inhaled at 5-min intervals. Second, guinea-pigs pre-treated with transnasal administration of FA or saline using the same protocol were passively sensitized with anti-ovalbumin (OA) serum 7 days before antigen challenge. Third, guinea-pigs were actively sensitized with OA and pre-treated with transnasal administration of FA or saline using the same protocol. The lateral pressure of the tracheal tube (Pao) was measured under anesthesia and artificial ventilation.. The antigen-induced increase in Pao in actively sensitized guinea-pigs was significantly potentiated by FA exposure in a dose-dependent manner. The dose-response curve of the methacholine-induced increase in Pao in non-sensitized guinea-pigs or of the antigen-induced increase in Pao in passively sensitized guinea-pigs was not altered by FA exposure. Transnasal administration of FA significantly increased the serum anti-OA homocytotropic antibody titre (IgG) as measured by the passive cutaneous anaphylaxis reaction in actively sensitized guinea-pigs.. The results suggest that repeated exposure to FA worsens allergic bronchoconstriction through enhancing antigen sensitization.

Topics: Allergens; Animals; Antibodies; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Fixatives; Formaldehyde; Guinea Pigs; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Male; Methacholine Chloride; Ovalbumin; Passive Cutaneous Anaphylaxis

IL-13 is a central mediator of allergen-induced airway hyperresponsiveness (AHR), but its role in respiratory syncytial virus (RSV)-induced AHR is not defined. The combination of allergen exposure and RSV infection is known to increase AHR and lung inflammation, but whether IL-13 regulates this increase is similarly not known.. Our objective was to determine the role of RSV infection and IL-13 on airway responsiveness and lung inflammation on sensitized and challenged mice.. Using a murine model of RSV infection and allergen exposure, we examined the role of IL-13 in the development of AHR and lung inflammation in IL-13 knockout mice, as well as using a potent IL-13 inhibitor (IL-13i). Mice were sensitized and challenged to allergen, and 6 days after the last challenge, they were infected with RSV. IL-13 was inhibited using an IL-13 receptor alpha(2)-human IgG fusion protein. AHR to inhaled methacholine was measured 6 days after infection, as was bronchoalveolar lavage fluid and lung inflammatory and cytokine responses.. RSV-induced AHR was unaffected by the IL-13i, despite prevention of goblet cell hyperplasia. Similar results were seen in IL-13-deficient mice. In sensitized and challenged mice, RSV infection significantly increased AHR, and after IL-13i treatment, AHR was significantly reduced, but to the levels seen in RSV-infected mice alone.. These results indicate that despite some similarities, the mechanisms leading to AHR induced by RSV are different from those that follow allergen sensitization and challenge. Because IL-13 inhibition is effective in preventing the increases in AHR and mucus production in sensitized and challenged mice infected with RSV, IL-13i could play an important role in preventing the consequences of viral infection in patients with allergic asthma.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Humans; Interleukin-13; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Syncytial Virus Infections; Respiratory Syncytial Virus, Human

Airway remodeling is one of the most important features of bronchial asthma. However, there are few studies that have used repeated antigen exposure in murine models. We designed a murine chronic antigen exposure model necessary for studying airway remodeling. Two different strains of mice, BALB/c mice and C57BL/6 mice, were sensitized and challenged for 3-7 weeks with ovalbumin (OVA). Bronchoalveolar lavage (BAL) and histology study were conducted in each phase. Morphometry was performed, and the epithelial area ratio (Ae ratio) and subepithelial area ratio (As ratio) were calculated. The Ae ratio and As ratio of BALB/c mice were significantly increased in sensitized mice compared with non-sensitized mice at 3 and 5 weeks, but not at 7 weeks. In C57BL/6 mice, the Ae ratio showed no significant changes, whereas the As ratio maintained high from 3 to 7 weeks. This thickening of the subepithelial layer consisted of collagen fibers with elastica van-Gieson (EVG) stain. Lymphocytes of the BAL showed a significant increase at 3 and 7 weeks in C57BL/6 mice, but not in BALB/c mice. A murine chronic OVA exposure model in C57BL/6 mice revealed subepithelial layer thickening consisting of collagen fibers and increased lymphocytes until 7 weeks. C57BL/6 mice are useful to elucidate the mechanism of airway remodeling.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Inhalation Exposure; Lung; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Models, Animal; Ovalbumin

Contact with immunomodulatory factors, such as LPS, in early infancy is associated with decreased allergen sensitization.. We sought to study the effects of systemic or airway exposure with LPS on the development of allergen sensitization, eosinophilic airway inflammation, and increased in vivo airway reactivity (AR) in a mouse model.. BALB/c mice were systemically sensitized with ovalbumin (OVA) plus adjuvant on days 1 and 14 and challenged through the airways with allergen on days 34 to 36. We performed measurement of OVA-specific IgE serum levels, in vitro T(H)2 cytokine production, differential cell counts in bronchoalveolar lavage fluids, and assessment of in vivo AR to inhaled methacholine by means of barometric whole-body plethysmography.. Systemic LPS administration before OVA sensitization reduced OVA-specific IgE serum levels (426 +/- 76 vs 880 +/- 104 U/mL, P <.01), T(H)2 cytokine production by splenic mononuclear cells (IL-4: 0.08 +/- 0.01 vs 0.17 +/- 0.01 ng/mL; IL-5: 1.98 +/- 0.52 vs 4.11 +/- 0.54 ng/mL; P <.01), and extent of airway eosinophilia (total cell counts: 93 vs 376 x 10(3)/mL; eosinophils: 23% vs 51%; P <.01) compared with that in OVA-sensitized mice. Local LPS administration to sensitized mice before airway allergen challenges particularly induced IFN-gamma production by peribronchial lymph node cells in vitro (1718 +/- 315 vs 483 +/- 103 ng/mL, P <.01) associated with reduced airway eosinophilia compared with that seen in OVA-sensitized mice. Development of increased AR was not affected by systemic or local LPS exposure. Inhibitory effects of LPS on allergen sensitization and eosinophilic airway inflammation were inhibited by administration of anti-IL-12 antibodies before LPS exposure.. These data indicate that local and systemic application of LPS modulates systemic and local T(H)1/T(H)2 immune responses in a distinct but similarly IL-12-dependent mode.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Eosinophilia; Female; Immunoglobulin E; Inflammation; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells

We have previously demonstrated that allergen inhalation induces expansion of bone marrow eosinophil progenitors in sensitized mice and subjects with asthma and that the inhaled corticosteroid, budesonide, reduced baseline but not allergen-induced increase in bone marrow eosinophil/basophil progenitors (EoB-CFU) in subjects with asthma. Here, we evaluated the effects of intranasal budesonide on allergen-induced increases in interleukin (IL)-5 and eotaxin in the airway and peripheral blood, expansion of bone marrow Eo-CFU and eosinophilia in bone marrow, peripheral blood and airway, as well as airway hyperresponsiveness, in ovalbumin (OVA)-sensitized mice. Budesonide treatment attenuated allergen-induced eosinophilia in bone marrow, peripheral blood, and airways as well as allergen-induced increases in bone marrow eosinophil progenitors but not allergen-induced increases in IL-5 or eotaxin 12 h following the second of two daily exposures to allergen; at later time points treatment was associated with attenuation of IL-5, eosinophilia, Eo-CFU, and airway hyperresponsiveness. These results suggest that a component of the mechanism by which corticosteroid treatment attenuates allergen-induced airway inflammation is through suppression of bone marrow eosinophilopoiesis, and that this is likely not mediated simply through the blocking of IL-5 production at the airway.

Topics: Administration, Intranasal; Airway Resistance; Allergens; Animals; Anti-Inflammatory Agents; Bone Marrow; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchodilator Agents; Budesonide; Chemokine CCL11; Chemokines, CC; Chemotactic Factors, Eosinophil; Eosinophils; Female; Glucocorticoids; Hematopoietic Stem Cells; Immunization; Interleukin-5; Leukocyte Count; Lung; Mice; Mice, Inbred BALB C; Ovalbumin

A catalytic antioxidant, AEOL 10113, was used in a murine model of asthma to test the hypothesis that oxidants contribute to the pathogenesis of asthma. Balb/c mice were immunized and challenged with ovalbumin. AEOL 10113 was administered to the mice by intratracheal instillation during ovalbumin challenges. Enhanced pause as an indicator of airway reactivity and differential cell count of lavage cells as an indicator of airway inflammation were assessed. Lung expressions of the adhesion molecules VCAM-1 and ICAM-1 were measured. We found that treatment of ovalbumin-challenged mice with AEOL 10113 drastically reduced the severity of airway inflammation as evidenced by the reduced numbers of eosinophils, neutrophils, and lymphocytes found in bronchoalveolar lavage fluid. Inhibition of ovalbumin-induced airway inflammation is associated with inhibited expression of VCAM-1, which is a key adhesion molecule responsible for the recruitment of inflammatory cells into the lungs of ovalbumin-challenged mice. In addition, treatment with AEOL 10113 reduced the magnitude of ovalbumin-induced airway hyperreactivity to methacholine. These results suggest that oxidative stress is an important factor in the pathogenesis of asthma and that a synthetic catalytic antioxidant could be effective in the treatment of asthma.

Topics: Animals; Antioxidants; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophils; Intercellular Adhesion Molecule-1; Lymphocytes; Male; Metalloporphyrins; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Vascular Cell Adhesion Molecule-1

The pulmonary effects of two environmentally relevant aldehydes were investigated in nonsensitized or ovalbumin (OA)-sensitized guineapigs (GPs). Four-week-old male Hartley GPs, weighing about 400 g, were intraperitoneally injected with 1 ml of an NaCl solution containing 100 microg OA and 100 mg Al(OH)(3). They were then exposed to either acetaldehyde (200 ppb) or benzaldehyde (500 ppb) for 4 wk (6 h/d, 5 d/wk). At the end of exposure, GPs were challenged with an OA aerosol (0.1% in NaCl) and pulmonary functions were measured. The day after, guinea pigs were anesthetized and several endpoints related to inflammatory and allergic responses were assessed in blood, whole-lung histology, and bronchoalveolar lavage (BAL). Sensitized nonexposed GPs showed bronchial hyperresponsiveness to OA and an increased number of eosinophils in blood and BAL, together with a rise in total protein and leukotrienes (LTB(4) and LTC(4)/D(4)/E(4)) in BAL. In nonsensitized GPs, exposure to acetaldehyde or benzaldehyde did not induce any change in the tested parameters, with the exception of irritation of the respiratory tract as detected by histology and an increased number of alveolar macrophages in animals exposed to acetaldehyde. In sensitized GPs, exposure to acetaldehyde induced a moderate irritation of the respiratory tract but no change in biological parameters linked to the inflammatory and allergic responses. In contrast, exposure to benzaldehyde induced a decrease both in OA-induced bronchoconstriction and in eosinophil and neutrophil numbers in BAL, an increase in the bronchodilatator mediator prostaglandin E(2) (PGE(2)), and a decrease in the bronchoconstrictor mediators LTC(4)/D(4)/E(4). Further investigations are needed to determine if the attenuated response observed in sensitized GPs exposed to benzaldehyde is due to an alteration of the mechanism of sensitization or to a more direct effect on various mechanisms of the allergic response.

Topics: Acetaldehyde; Administration, Inhalation; Air Pollutants; Animals; Benzaldehydes; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Guinea Pigs; Injections, Intraperitoneal; Lung; Male; Ovalbumin; Respiratory Function Tests; Respiratory Hypersensitivity

This report examines the effect of heat-killed Mycobacterium vaccae in a mouse model of allergic pulmonary inflammation. The s.c. administration of M. vaccae 3 wk before the immunization significantly reduced Ag-induced airway hyperreactivity and the increase in the numbers of eosinophils observed in the bronchoalveolar lavage fluid, blood, and bone marrow, even though no detectable changes in either cytokine (IL-4, IL-13, IL-5, and IFN-gamma) or total IgE levels were observed. Furthermore, transfer of splenocytes from OVA-immunized and M. vaccae-treated mice into recipient, OVA-immunized mice significantly reduced the allergen-induced eosinophilia by an IFN-gamma-independent mechanism, clearly indicating that the mechanism by which M. vaccae induces its inhibitory effect is not due to a redirection from a predominantly Th2 to a Th1-dominated immune response. The protective effect of M. vaccae on the allergen-induced eosinophilia lasted for at least 12 wk after its administration, and the treatment was also effective in presensitized mice. Moreover, the allergen specificity of the inhibitory effect could be demonstrated using a double-immunization protocol, where M. vaccae treatment before OVA immunization had no effect on the eosinophilic inflammation induced by later immunization and challenge with cockroach extract Ag. Taken together, these results clearly demonstrate that M. vaccae is effective in blocking allergic inflammation by a mechanism independent of IFN-gamma, induces long term and Ag-specific protection, and therefore has both prophylactic and therapeutic potential for the treatment of allergic diseases.

Topics: Adoptive Transfer; Animals; Antigens; Bacterial Vaccines; Bronchial Hyperreactivity; Chemokines; Cytokines; Eosinophilia; Eosinophils; Female; Hypersensitivity; Immunization; Immunoglobulin E; Immunoglobulin G; Inflammation; Mice; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Ovalbumin; Vaccines, Inactivated

Asthma is caused by T-helper cell 2 (Th2)-driven immune responses, but the immunological mechanisms that protect against asthma development are poorly understood. T-cell tolerance, induced by respiratory exposure to allergen, can inhibit the development of airway hyperreactivity (AHR), a cardinal feature of asthma, and we show here that regulatory T (T(R)) cells can mediate this protective effect. Mature pulmonary dendritic cells in the bronchial lymph nodes of mice exposed to respiratory allergen induced the development of T(R) cells, in a process that required T-cell costimulation via the inducible costimulator (ICOS-ICOS-ligand pathway. The T(R) cells produced IL-10, and had potent inhibitory activity; when adoptively transferred into sensitized mice, T(R) cells blocked the development of AHR. Both the development and the inhibitory function of regulatory cells were dependent on the presence of IL-10 and on ICOS-ICOS-ligand interactions. These studies demonstrate that T(R) cells and the ICOS-ICOS-ligand signaling pathway are critically involved in respiratory tolerance and in downregulating pulmonary inflammation in asthma.

Topics: Allergens; Animals; Antigens; Antigens, Differentiation, T-Lymphocyte; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Inducible T-Cell Co-Stimulator Ligand; Inducible T-Cell Co-Stimulator Protein; Interleukin-10; Ligands; Mice; Mice, Inbred BALB C; Mice, Transgenic; Ovalbumin; Proteins; T-Lymphocytes

Macrophages (Mphi) play an unique role in the activation and regulation of T cells through their ability to modulate specific costimulatory and cytokine signals. Here we investigated the immunomodulatory effects of allergen presentation by Mphi in a murine model of allergic asthma. Purified peritoneal Mphi were pulsed with ovalbumin (OVA) (OVA-Mphi), or the immunodominant epitope OVA(323-339) (OVA(323-339)-Mphi), and characterized for cell surface markers, cytokine production, and antigen-presenting capacity toward OVA(323-339)-specific DO11.10 T cells. Antigen-pulsed Mphi were injected (intravenously) in OVA-sensitized Balb/c mice that were repeatedly challenged with OVA or saline aerosol. Administration of OVA-Mphi inhibited airway eosinophilia and hyperresponsiveness to methacholine concomitant with a reduced interleukin (IL)-4 and IL-5 production by T cells upon OVA stimulation in vitro. Interestingly, OVA-induced IL-10 levels remained unchanged, whereas interferon-gamma could not be detected. In contrast to OVA-Mphi, OVA(323-339)-Mphi administration had no effects on these asthma manifestations. Additional in vitro studies demonstrated that OVA-Mphi, but not OVA(323-339)-Mphi, produced high levels of IL-10 upon interaction with the DO11.10 T cells. This IL-10 production by the OVA-Mphi was dependent on MHC-TCR and CD86-CD28, but not CD80-CD28 or CD40-CD154 interactions. Our data suggest that IL-10 production by allergen presenting Mphi plays a crucial role in successful immunotherapy.

Topics: Adjuvants, Immunologic; Animals; Antigen Presentation; Antigens; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Flow Cytometry; Interferon-gamma; Interleukin-10; Lung; Macrophages, Peritoneal; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Phenotype

We have studied murine models of asthma using FcepsilonRIalpha-chain-deficient (FcepsilonRIalpha(-/-)) mice to investigate the role of IgE-dependent mast cell activation in these models. When mice were either 1) immunized once with OVA in alum i.p. and then challenged with OVA intranasally, or 2) repeatedly immunized with OVA in the absence of adjuvant and subsequently challenged with nebulized OVA, FcepsilonRalpha(-/-) mice had significantly fewer eosinophils and lower IL-4 levels in their bronchoalveolar lavage fluid compared with wild-type mice. When mice were given anti-IL-5 antibody before OVA challenge in protocol 1, eosinophilic infiltration into the airways was significantly suppressed in both genotypes, but only FcepsilonRIalpha(-/-) mice showed significantly reduced airway hyperresponsiveness (AHR). In addition, when mice immunized and challenged with OVA also received a late OVA provocation at a higher concentration and were then exposed to methacholine, only wild-type mice developed a substantial increase in AHR. Since FcepsilonRI is expressed mainly on mast cells in mouse airways, we conclude that IgE-dependent activation of this cell type plays an important role in the development of allergic airway inflammation and AHR in mice. The models used may be of value for testing inhibitors of IgE or mast cells for development of therapeutic agents for human asthma.

Topics: Allergens; Animals; Antibodies; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Humans; Immunization; Immunoglobulin E; Interleukin-5; Mast Cells; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Receptors, IgE

The central role for Th2 cells in the development of Ag-induced airway hyperresponsiveness and eosinophilic inflammation is well documented. We have reported a crucial role for TCR-induced activation of the Ras/extracellular signal-regulated kinase mitogen-activated protein kinase cascade in Th2 cell differentiation. Here, we show that the development of both OVA-induced airway hyperresponsiveness and eosinophilic airway inflammation in a mouse asthma model are attenuated in transgenic mice by the overexpression of enzymatically inactive Ras molecules in T cells. In addition, reduced levels of IL-5 production and eosinophilic inflammation induced by nematode infection (Nippostrongylus brasiliensis or Heligmosomoides polygyrus) were detected. Thus, the level of Ras activation in T cells appears to determine Th2-dependent eosinophilic inflammation and Ag-induced airway hyperresponsiveness.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophilia; Gene Expression Regulation; Genes, ras; Inflammation; Interleukin-5; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Transgenic; Nematospiroides dubius; Nippostrongylus; Ovalbumin; Strongylida Infections; T-Lymphocytes; Th2 Cells

Interferon (IFN)-gamma reduces airway responses after allergen challenge in mice. The mechanisms of this effect are not clear. These studies investigate whether IFN-gamma can reverse prolonged airway responses after allergen challenge in IFN-gamma-deficient (IFN-gammaKO) mice. Sensitized mice (IFN-gammaKO and wild-type [WT]) were challenged with ovalbumin. Airway responsiveness, eosinophils in bronchoalveolar lavage fluid, and lung lymphocyte subsets (CD4(+) and CD8(+)) were measured 24 hours and 8 weeks after challenge. In further experiments, we treated IFN-gammaKO mice with recombinant IFN-gamma starting 4 weeks after the challenge for 1 week or 4 weeks. Airway responsiveness, bronchoalveolar lavage eosinophils, and lung CD4(+) cells were increased 8 weeks after challenge in IFN-gammaKO but not WT mice. IFN-gamma treatment returned lung CD4(+) cell numbers to values obtained in unchallenged mice. One week of IFN-gamma treatment also returned airway responsiveness to baseline levels; however, 4-week treatment with IFN-gamma failed to decrease airway responsiveness below levels observed in untreated animals. This suggests that IFN-gamma plays an essential role in reversing allergen-induced airway inflammation and hyperresponsiveness and that it may have dual actions on the latter. Observations that IFN-gamma reverses airway responses, even when administered after challenge, suggests that IFN-gamma treatment could control allergic disease, including asthma.

Topics: Airway Obstruction; Animals; Animals, Wild; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; CD4-CD8 Ratio; Disease Models, Animal; Drug Evaluation, Preclinical; Eosinophils; Female; Humans; Hypersensitivity; Inflammation; Interferon-gamma; Mice; Mice, Inbred BALB C; Ovalbumin

Exposure to endotoxin has been associated with an exacerbation of asthmatic responses in humans and animal models. However, recent evidence suggests that microbial exposure in early life may protect from the development of asthma and atopy. In this study, we sought to evaluate the effects of lipopolysaccaride (LPS) on airway function in developing mice. In addition, we evaluated the influence of LPS on subsequent allergen sensitization and challenge. Under light anesthesia, 2-3-week-old Balb/c mice received a single intranasal instillation of LPS or sterile physiologic saline. Measurements of airway function were obtained in unrestrained animals, using whole-body plethysmography. Airway responsiveness was expressed in terms of % enhanced pause (Penh) increase from baseline to aerosolized methacholine (Mch). In additional studies, we assessed the functional and cellular responses to ovalbumin sensitization and challenge following prior exposure to LPS. We found that exposure to LPS induced transient airway hyperresponsiveness to Mch. These functional changes were associated with the recruitment of neutrophils and lymphocytes into the bronchoalveolar lavage (BAL) fluid. Airway responsiveness after allergen sensitization and challenge was decreased by prior exposure to LPS. The analysis of BAL cells and cytokines (interferon-gamma and interleukin-4) did not reveal alterations in the overall Th1/Th2 balance. Our findings suggest that LPS leads to airway hyperresponsiveness in developing mice, and may protect against the development of allergen-driven airway dysfunction.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cell Count; Cytokines; Dose-Response Relationship, Drug; Endotoxins; Inflammation; Lipopolysaccharides; Lymphocytes; Methacholine Chloride; Mice; Mice, Inbred BALB C; Neutrophils; Ovalbumin; Reference Values

T(H)2 cells play a critical role in the pathogenesis of asthma, but the precise immunologic mechanisms that inhibit T(H)2 cell function in vivo are not well understood.. The purpose of our studies was to determine whether T cells producing IL-10 regulate the development of asthma.. We used gene therapy to generate ovalbumin-specific CD4 T-helper cells to express IL-10, and we examined their capacity to regulate allergen-induced airway hyperreactivity.. We demonstrated that the CD4 T-helper cells engineered to express IL-10 abolished airway hyperreactivity and airway eosinophilia in BALB/c mice sensitized and challenged with ovalbumin and in SCID mice reconstituted with ovalbumin-specific T(H)2 effector cells. The inhibitory effect of the IL-10-secreting T-helper cells was accompanied by the presence of increased quantities of IL-10 in the bronchoalveolar lavage fluid, was antigen-specific, and was reversed by neutralization of IL-10. Moreover, neutralization of IL-10 by administration of anti-IL-10 mAb in mice sensitized and challenged with ovalbumin seriously exacerbated airway hyperreactivity and airway inflammation.. Our results demonstrate that T cells secreting IL-10 in the respiratory mucosa can indeed regulate T(H)2-induced airway hyperreactivity and inflammation, and they strongly suggest that IL-10 plays an important inhibitory role in allergic asthma.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cell Line; Cytokines; Dose-Response Relationship, Drug; Genetic Therapy; Inflammation; Interleukin-10; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, SCID; Ovalbumin; Protein Engineering; Pulmonary Eosinophilia; T-Lymphocytes, Helper-Inducer; Th2 Cells; Transduction, Genetic

The role of nitric oxide (NO) in allergic inflammation and bronchial hyperresponsiveness is unclear. We studied a selective prodrug nitric oxide synthase (NOS)-2 inhibitor, L-N(6)-(1-iminoethyl)lysine 5-tetrazole amide (SC-51). In ovalbumin-sensitized and challenged rats, exhaled NO levels increased by 3 h following challenge (3.73 +/- 0.74 ppb; P < 0.05), peaking at 9 h (11.0 +/- 2.75; P < 0.01) compared to saline controls (1.87 +/- 0.26; P < 0.05 and 2.81 +/- 0.18; P < 0.01). Immunoreactive lung NOS2 expression was increased in ovalbumin-challenged rats compared with ovalbumin-sensitized, saline-challenged rats at 8 h post-challenge. SC-51 (10 mg/kg; p.o.) inhibited allergen-induced increase in exhaled NO levels to 1.3 +/- 0.17 ppb. SC-51 inhibited bronchial hyperresponsiveness in ovalbumin-sensitized and challenged rats (P < 0.05). In sensitized non-exposed rats, SC-51 increased bronchial responsiveness (P < 0.05). SC-51 reduced the allergen-induced increase in bronchoalveolar lavage neutrophils, but caused a nonsignificant reduction in bronchial mucosal eosinophil numbers. NO generated through NOS2 contributes to allergen-induced bronchial hyperresponsiveness but not to bronchial eosinophilia, indicating that these are independently expressed.

Topics: Acetylcholine; Airway Resistance; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Enzyme Inhibitors; Eosinophils; Homoarginine; Hypersensitivity; Immunohistochemistry; Inflammation; Isoenzymes; Lung; Male; Nitric Oxide; Nitric Oxide Synthase; Ovalbumin; Rats; Rats, Inbred BN

In a recent study, we found that TNF-alpha negatively regulates airway responsiveness through the activation of gammadelta T cells. The biological activities of TNF-alpha are mediated by two structurally related but functionally distinct receptors, p55 (TNFR1) and p75 (TNFR2), which are independently expressed on the cell surface. However, the relative importance of either TNFR in airway hyperresponsiveness (AHR) is unknown. To investigate the importance of these TNFRs in the development of allergen-induced AHR, p55-deficient and p75-deficient mice were sensitized to OVA by i.p. injection and subsequently challenged with OVA via the airways; airway responsiveness to inhaled methacholine was monitored. p75-deficient mice developed AHR to a similar degree as control mice. In contrast, p55-deficient mice, which were sensitized and challenged with OVA, failed to develop AHR. In p55-deficient mice, both the numbers of eosinophils and levels of IL-5 in bronchoalveolar lavage fluid were significantly lower than in sensitized/challenged control mice (p < 0.05). However, depletion of gammadelta T cells resulted in significant increases in AHR in the p55-deficient mice, whereas no significant effect of gammadelta T cell depletion was evident in the p75-deficient mice. These data indicate that, in the absence of TNFR1 (p55), where TNF-alpha uses the p75 pathway exclusively, the development of AHR is regulated by gammadelta T cells.

Topics: Animals; Antibodies, Monoclonal; Antigens, CD; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Female; Genetic Predisposition to Disease; Immunoglobulin E; Inflammation; Injections, Intraperitoneal; Injections, Intravenous; Lymphocyte Depletion; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Receptors, Antigen, T-Cell, alpha-beta; Receptors, Antigen, T-Cell, gamma-delta; Receptors, Tumor Necrosis Factor; Receptors, Tumor Necrosis Factor, Type I; Receptors, Tumor Necrosis Factor, Type II; T-Lymphocyte Subsets

Bronchial hyperresponsiveness and eosinophilia are major characteristics of asthma. Calcitonin gene-related peptide (CGRP) is a neuropeptide that has various biological actions. In the present study, we questioned whether CGRP might have pathophysiological roles in airway hyperresponsiveness and eosinophilia in asthma. To determine the exact roles of endogenous CGRP in vivo, we chose to study antigen-induced airway responses using CGRP gene-disrupted mice. After ovalbumin sensitization and antigen challenge, we assessed airway responsiveness and measured proinflammatory mediators. In the sensitized CGRP gene-disrupted mice, antigen-induced bronchial hyperresponsiveness was significantly attenuated compared with the sensitized wild-type mice. Antigen challenge induced eosinophil infiltration in bronchoalveolar lavage fluid, whereas no differences were observed between the wild-type and CGRP-mutant mice. Antigen-induced increases in cysteinyl leukotriene production in the lung were significantly reduced in the CGRP-disrupted mice. These findings suggest that CGRP could be involved in the antigen-induced airway hyperresponsiveness, but not eosinophil infiltration, in mice. The CGRP-mutant mice may provide appropriate models to study molecular mechanisms underlying CGRP-related diseases.

Topics: Animals; Antigens; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Calcitonin Gene-Related Peptide; Crosses, Genetic; Disease Models, Animal; DNA; Female; Genomic Library; Genotype; Immunohistochemistry; Leukocyte Count; Male; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin

Asthma is characterized by chronic airways inflammation, airway wall remodeling, and airway hyperresponsiveness (AHR). An increase in airway smooth muscle has been proposed to explain a major part of AHR in asthma. We have used unbiased stereological methods to determine whether airway smooth muscle hyperplasia and AHR occurred in sensitized, antigen-challenged Brown Norway (BN) rats. Ovalbumin (OA)-sensitized BN rats chronically exposed to OA aerosol displayed airway inflammation and a modest level of AHR to intravenously administered ACh 24 h after the last antigen challenge. However, these animals did not show an increase in smooth muscle cell (SMC) number in the left main bronchus, suggesting that short-lived inflammatory mechanisms caused the acute AHR. In contrast, 7 days after the last aerosol challenge, there was a modest increase in SMC number, but no AHR to ACh. Addition of FCS to the chronic OA challenge protocol had no effect on the degree of inflammation but resulted in a marked increase in both SMC number and a persistent (7-day) AHR. These results raise the possibility that increases in airway SMC number rather than, or in addition to, chronic inflammation contribute to the persistent AHR detected in this model.

Topics: Acetylcholine; Airway Resistance; Animals; Bronchi; Bronchial Hyperreactivity; Bronchitis; Cell Count; Hyperplasia; Hypersensitivity, Immediate; Male; Muscle, Smooth; Ovalbumin; Rats; Rats, Inbred BN

Although life-long immunity against pathogens is beneficial, immunological memory responses directed against allergens are potentially harmful. Because there is a paucity of information about Th2 memory cells in allergic disease, we established a model of allergic asthma in BALB/c mice to explore the generation and maintenance of Th2 memory. We induced disease without the use of adjuvants, thus avoiding Ag depots, and found that unlike allergic asthma in mice immunized with adjuvant, immunizing with soluble and aerosol OVA resulted in pathological lung lesions resembling human disease. To test memory responses we allowed mice with acute disease to recover and then re-exposed them to aerosol OVA a second time. Over 400 days later these mice developed OVA-dependent eosinophilic lung inflammation, airway hyperresponsiveness, mucus hypersecretion, and IgE. Over 1 year after recuperating from acute disease, mice had persistent lymphocytic lung infiltrates, Ag-specific production of IL-4 and IL-5 from spleen and lung cells in vitro, and elevated IgG1. Moreover, when recuperated mice were briefly aerosol challenged, we detected early expression of Th2 cytokine RNA in lungs. Taken together, these data demonstrate the presence of long-lived Th2 memory cells in spleen and lungs involved in the generation of allergic asthma upon Ag re-exposure.

Topics: Acute Disease; Aerosols; Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Female; Immunoglobulins; Immunologic Memory; Inflammation; Lung; Lymph Nodes; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Spleen; Th2 Cells; Time Factors

An oxidant/antioxidant imbalance is seen in the lungs of patients with asthma. This oxidative stress in asthmatic airways may lead to activation of redox-sensitive transcription factors, NF-kappaB and AP-1. We examined the effect of the small molecule inhibitor of redox-regulated NF-kappaB and AP-1 transcription, MOL 294 on airway inflammation and airway hyperreactivity (AHR) in a mouse model of asthma. MOL 294 is a potent nonpeptide inhibitor of NF-kappaB and AP-1 based upon a beta-strand template that binds to and inhibits the cellular redox protein thioredoxin. BALB/c mice after i.p. OVA sensitization (day 0) were challenged with intranasal OVA on days 14, 25, 26, and 27. MOL 294, administered intranasal on days 25-27, blocked the airway inflammatory response to OVA assessed 24 h after the last OVA challenge on day 28. MOL 294 reduced eosinophil, IL-13, and eotaxin levels in bronchoalveolar lavage fluid and airway tissue eosinophilia and mucus hypersecretion. MOL 294 also decreased AHR in vivo to methacholine. These results support redox-regulated transcription as a therapeutic target in asthma and demonstrate that selective inhibitors can reduce allergic airway inflammation and AHR.

Topics: Administration, Intranasal; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Movement; Chemokine CCL11; Chemokines, CC; Disease Models, Animal; Eosinophils; Female; Humans; Inflammation; Interleukin-13; Lung; Mice; Mice, Inbred BALB C; Mucus; NF-kappa B; Ovalbumin; Oxidation-Reduction; Pyridazines; Thioredoxins; Transcription Factor AP-1; Triazoles; Tumor Cells, Cultured

Microbial heat shock proteins (hsp) have been associated with the generation and induction of Th1-type immune responses. We tested the effects of treatment with five different microbial hsp (Mycobacterium leprae, Streptococcus pneumoniae, Helicobacter pylori, bacillus Calmette-Guérin, and Mycobacterium tuberculosis) in a murine model of allergic airway inflammation and airway hyperresponsiveness (AHR). Mice were sensitized to OVA by i.p. injection and then challenged by OVA inhalation. Hsp were administered to each group by i.p. injection before sensitization and challenge. Sensitized and challenged mice developed increased serum levels of OVA-specific IgE with significant airway eosinophilia and heightened responsiveness to methacholine when compared with nonsensitized animals. Administration of M. leprae hsp prevented both development of AHR as well as bronchoalveolar lavage fluid eosinophilia in a dose-dependent manner. Treatment with M. leprae hsp also resulted in suppression of IL-4 and IL-5 production in bronchoalveolar lavage fluid, while IL-10 and IFN-gamma production were increased. Furthermore, M. leprae hsp treatment significantly suppressed OVA-specific IgE production and goblet cell hyperplasia/mucin hyperproduction. In contrast, treatment with the other hsp failed to prevent changes in airway responsiveness, lung eosinophilia, or cytokine production. Depletion of gamma/delta T lymphocytes before sensitization and challenge abolished the effect of M. leprae hsp treatment on AHR. These results indicate selective and distinctive properties among the hsp, and that M. leprae hsp may have a potential therapeutic role in the treatment of allergic airway inflammation and altered airway function.

Topics: Animals; Bacterial Proteins; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Movement; Cytokines; Down-Regulation; Epitopes; Female; Goblet Cells; Heat-Shock Proteins; Hyperplasia; Immunoglobulin E; Inflammation; Interleukin-4; Interleukin-5; Lung; Lymph Nodes; Lymphocyte Depletion; Mice; Mucins; Mycobacterium leprae; Ovalbumin; Pulmonary Eosinophilia; Receptors, Antigen, T-Cell, gamma-delta; Respiratory Mucosa; T-Lymphocyte Subsets

Trypsin and mast cell tryptase can signal to epithelial cells, myocytes, and nerve fibers of the respiratory tract by cleaving proteinase-activated receptor 2 (PAR2). Since tryptase inhibitors are under development to treat asthma, a precise understanding of the contribution of PAR2 to airway inflammation is required. We examined the role of PAR2 in allergic inflammation of the airway by comparing OVA-sensitized and -challenged mice lacking or overexpressing PAR2. In wild-type mice, immunoreactive PAR2 was detected in airway epithelial cells and myocytes, and intranasal administration of a PAR2 agonist stimulated macrophage infiltration into bronchoalveolar lavage fluid. OVA challenge of immunized wild-type mice stimulated infiltration of leukocytes into bronchoalveolar lavage and induced airway hyperreactivity to inhaled methacholine. Compared with wild-type animals, eosinophil infiltration was inhibited by 73% in mice lacking PAR2 and increased by 88% in mice overexpressing PAR2. Similarly, compared with wild-type animals, airway hyperreactivity to inhaled methacholine (40 micro g/ml) was diminished 38% in mice lacking PAR2 and increased by 52% in mice overexpressing PAR2. PAR2 deletion also reduced IgE levels to OVA sensitization by 4-fold compared with those of wild-type animals. Thus, PAR2 contributes to the development of immunity and to allergic inflammation of the airway. Our results support the proposal that tryptase inhibitors and PAR2 antagonists may be useful therapies for inflammatory airway disease.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cell Line; Cell Movement; Crosses, Genetic; Female; Humans; Immunoglobulin E; Inflammation; Injections, Intraperitoneal; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Ovalbumin; Pulmonary Eosinophilia; Receptor, PAR-2; Receptors, Thrombin

Nitrogen dioxide (NO(2)) is a common indoor and outdoor air pollutant whose role in the induction of asthma is unclear. We investigated the effects of NO(2) on the development of asthma-like responses to allergenic challenge in BALB/c mice. Ovalbumin (OVA)-immunized mice were intranasally challenged with OVA or saline solution just before starting a 3 h exposure to 5 or 20 ppm NO(2) or air. Twenty parts per million of NO(2) induced a significant increase of bronchopulmonary hyperreactivity in OVA-challenged mice and of permeability according to the fibronectin content of the bronchoalveolar lavage fluid (BALF) 24 h after exposure, as compared with air or 5 ppm NO(2). Eosinophilia (cell counts in the BALF and eosinophil peroxidase of lung tissue) was detected at 24 and 72 h with similar levels for air and 20 ppm NO(2), whereas a marked reduction was unexpectedly observed for 5 ppm NO(2). At 24 h, interleukin-5 in the BALF was markedly reduced at 5 ppm compared with 20 ppm NO(2) and was also more intense for 20 ppm NO(2) than for the air group. In contrast to specific IgG1 titers, anti-OVA IgE titers and interleukin-4 in the BALF were not affected by NO(2) exposure. Irrespective of the concentration of NO(2), OVA-challenged mice did not develop late mucosal metaplasia compared with those exposed to OVA-air. These results indicate that a short exposure to NO(2) can exacerbate or inhibit some features of the development of allergic disease in mice and may depend on the concentration of pollutant.

Topics: Air Pollutants; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Fibronectins; Immunoglobulin E; Immunoglobulin G; Interleukin-4; Interleukin-5; Lung; Male; Mice; Mice, Inbred BALB C; Nitrogen Dioxide; Ovalbumin

The mechanisms underlying airway hyperresponsiveness remain unclear, although airway inflammation and remodeling are likely important contributing factors. We hypothesized that airway physiology would differ between mice subjected to brief or chronic allergen exposure, and that these differences would be associated with characteristic inflammatory markers and indices of airway remodeling. BALB/c mice were sensitized to ovalbumin and studied at several time points following brief or chronic allergen challenge protocols. By measuring airway responses to methacholine, we demonstrated increases in maximal inducible bronchoconstriction that persisted for 8 wk following either brief or chronic allergen challenge; we also observed increases in airway reactivity, although it was only in chronically challenged mice that these changes persisted beyond the resolution of allergen-induced inflammation. Using airway morphometry, we further demonstrated that increases in maximal bronchoconstriction were associated with increases in airway contractile tissue in both models, and that chronic, but not brief, allergen challenge resulted in subepithelial fibrosis. Our observations that different aspects of sustained airway dysfunction and remodeling persist beyond the resolution of acute inflammatory events support the concept that remodeling occurs as a consequence of allergic airway inflammation, and that these structural changes contribute independently to the persistence of airway hyperresponsiveness.

Topics: Actins; Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage; Bronchoconstriction; Eosinophils; Extracellular Matrix; Female; Inflammation; Interleukin-13; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mucins; Muscle, Smooth; Ovalbumin

The mechanisms of airway hyperresponsiveness (AHR) are still poorly understood. In this study we have established a model of persistent AHR and eosinophilia and evaluated the prophylactic vs. therapeutic effects of dexamethasone on these parameters. Mice were immunised with ovalbumin (OVA) on day 0 and challenged intranasally on days 10, 11, 12 and 13 with OVA or phosphate buffer saline (PBS). Airway responsiveness to methacholine, measured 24-h post multiple intranasal OVA challenges, was significantly increased compared to time matched PBS-controls (P<0.05). AHR could be detected for up to 14 days after the last OVA challenge although the magnitude of the AHR had diminished by day 14 compared to day 1. OVA challenge of mice induced a significant airway eosinophilia at 24h (P<0.05); this persisted for 2 weeks after the challenge. Prophylactic treatment with dexamethasone (1mg x kg (-1)) reduced the OVA induced AHR, eosinophilia and mucus cell hyperplasia/metaplasia measured 24h post challenge. Therapeutic treatment, with dexamethasone (2 mg x kg(-1)), significantly inhibited established airway eosinophilia, measured at 72 h post OVA challenge, only when treatment was initiated at 24h but not 48 h after challenge. In contrast, AHR measured at 72 h post OVA challenge was significantly reduced when treatment was started at either 24 or 48 h post challenge. Our data shows that the immunization and challenge procedures employed resulted in a persistent type of AHR. Prophylactic intervention with steroids almost completely inhibited its development; however therapeutic intervention only partially resolved AHR.

Topics: Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Dexamethasone; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Pulmonary Eosinophilia; Time Factors

Asthma is a chronic inflammatory disease of the lung resulting in airway obstruction. The airway inflammation of asthma is strongly linked to Th2 lymphocytes and their cytokines, particularly IL-4, IL-5, and IL-13, which regulate airway hyperresponsiveness, eosinophil activation, mucus production, and IgE secretion. Historically, complement was not thought to contribute to the pathogenesis of asthma. However, our previous reports have demonstrated that complement contributes to bronchial hyperreactivity, recruitment of airway eosinophils, IL-4 production, and IgE responses in a mouse model of pulmonary allergy. To define the complement activation fragments that mediate these effects, we assessed the role of the complement anaphylatoxin C3a in a mouse model of pulmonary allergy by challenging C3aR-deficient mice intranasally with a mixed Ag preparation of Aspergillus fumigatus cell culture filtrate and OVA. Analysis by plethysmography after challenge revealed an attenuation in airway hyperresponsiveness in C3aR-deficient mice relative to wild-type mice. C3aR-deficient mice also had an 88% decrease in airway eosinophils and a 59% reduction in lung IL-4-producing cells. Consistent with the reduced numbers of IL-4-producing cells, C3aR-deficient mice had diminished bronchoalveolar lavage levels of the Th2 cytokines, IL-5 and IL-13. C3aR knockout mice also exhibited decreases in IgE titers as well as reduced mucus production. Collectively, these data highlight the importance of complement activation, the C3a anaphylatoxin, and its receptor during Th2 development in this experimental model and implicate these molecules as possible therapeutic targets in diseases such as asthma.

Topics: Animals; Antigens, Fungal; Aspergillus fumigatus; Bronchial Hyperreactivity; Cell Movement; Complement C3a; Disease Models, Animal; Down-Regulation; Eosinophils; Immunoglobulin Isotypes; Inflammation; Injections, Intraperitoneal; Interferon-gamma; Interleukin-4; Lung; Membrane Proteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Mucins; Ovalbumin; Receptors, Complement; Respiratory Hypersensitivity; Th2 Cells

Atrial natriuretic peptide (ANP) is a bronchodilator; however, the short half-life of ANP in vivo limited its therapeutic utility to treat asthma. The efficacy of intranasally administered plasmid DNA-expressing ANP (pANP; amino acid 99-126; Acc. No. XM131840) on the prevention of allergen-induced airway hyperresponsiveness (AHR) was examined in this study by using a mouse model of asthma. Ovalbumin-sensitized mice were treated with pANP versus control plasmids, and AHR was monitored after ovalbumin challenge for 5 weeks on 10-day intervals starting 4 days after gene transfer. Mice administered pANP demonstrated significantly less AHR for 20 days after treatment. The results demonstrate that pANP gene transfer protects against AHR and might be useful in the treatment of asthma.

Topics: Administration, Intranasal; Animals; Asthma; Atrial Natriuretic Factor; Bronchial Hyperreactivity; Disease Models, Animal; Female; Gene Transfer Techniques; Genetic Therapy; Lung; Mice; Mice, Inbred BALB C; Ovalbumin

Toxic influence of high oxygen concentration on pulmonary function and structures has been known for many years. However, the influence of high oxygen concentration breathing on defensive respiratory reflexes is still not clear. In our previous experiments, we found an inhibitory effect of 100 % oxygen breathing on cough reflex intensity in healthy guinea pigs. The present study was designed to detect the effects of hyperoxia on cough reflex in guinea pigs with allergic airway inflammation. In the first phase of our experiment, the animals were sensitized with ovalbumin. Thirty-two sensitized animals were used in two separate experiments according to oxygen concentration breathing: 100 % or 50 % oxygen for 60 h continuously. In each experiment, one group of animals was exposed to hyperoxia, another to ambient air. The cough reflex was induced both by aerosol of citric acid before sensitization, then in sensitized animals at 24 h and 60 h of exposition to oxygen/air in awake animals, and by mechanical stimulation of airway mucosa in anesthetized animals just after the end of the experiment. In contrast to 50 % oxygen, 100 % oxygen breathing leads to significant decrease in chemically induced cough in guinea pigs with allergic inflammation. No significant changes were present in cough induced by mechanical stimulation of airways.

Topics: Animals; Bronchial Hyperreactivity; Citric Acid; Cough; Female; Guinea Pigs; Hyperoxia; Hypersensitivity; Ovalbumin; Oxygen; Reflex; Respiratory Mechanics

We investigated whether an expression of hemeoxygenase-1 (HO-1) within the lung tissue is related to allergic airway inflammation and HO-1 expression could influence airway hyperreactivity (AHR) and eosinophilia in C57BL/6 mice actively sensitized to ovalbumin. The number of HO-1 positive cells was increased in the subepithelium of the bronchi after OVA challenge and HO-1 was localized to alveolar macrophage.Zinc-protoporphyrin (Zn-PP), a competitive inhibitor of hemeoxygenase, by intraperitoneal injection clearly inhibited AHR, pulmonary eosinophilia and IL-5 and IL-13 in the lung tissue. These data indicate that the expression of HO-1 is increased within the lung tissue in allergic airway inflammation and the overexpression of HO-1 could enhanced allergic airway inflammation.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Enzyme Inhibitors; Heme Oxygenase (Decyclizing); Heme Oxygenase-1; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Ovalbumin; Protoporphyrins

IL-16 has been described as a natural soluble CD4-ligand with immunosuppressive effects in vitro. However, little is known about the effect of IL-16 on immune responses in vivo.. In the present study, we examined the effect of IL-16 administration in a murine model of allergic asthma. Next, we determined whether these effects were mediated by modulation of CD4+ T lymphocytes.. Intraperitoneal administration of IL-16 completely inhibits antigen-induced airway hyper-responsiveness and largely decreases the number of eosinophils in bronchoalveolar lavage fluid (> 90%) and airway tissue of ovalbumin-sensitized and challenged mice. Firstly, it appears that thoracic lymph node cells isolated from in vivo IL-16-treated ovalbumin-challenged animals produce less IL-4 (77%) and IL-5 (85%) upon antigenic re-stimulation, when compared to vehicle-treated mice. Secondly, pre-incubation of lymphocytes with IL-16 in vitro reduces antigen-induced proliferation (55%) and Th2-type cytokine production (IL-4; 56%, IL-5; 77%). Thirdly, the presence of IL-16 during priming cultures of TCR transgenic T cells (DO11.10), reduces IL-4 (33%) and IL-5 (35%), but not IL-10 and IFNgamma levels upon re-stimulation.. It can be concluded that IL-16 has potent immunosuppressive effects on a Th2dominated allergic airway response.

Topics: Animals; Antigens; Asthma; Bronchial Hyperreactivity; Cell Differentiation; Cytokines; Eosinophilia; Immunoglobulin E; Interferon-gamma; Interleukin-16; Lung; Male; Mice; Mice, Inbred BALB C; Models, Animal; Ovalbumin; Th2 Cells

To investigate bronchial responsiveness to acetylcholine in allergic airway inflammation of SD rats.. SD rats were immunized and challenged by chicken ovalbumin (OVA). Airway responsiveness, acetylcholine (Ach) provocation concentration needed to increase baseline airway resistance by 200% (PC(200)) were measured.. The value of baseline airway resistance in asthma group was significantly higher than that in control group (2.282 +/- 0.128 vs 3.193 +/- 0.239; P < 0.01). After multiple ovalbumin exposures, airway responsiveness to intravenous injection of acetylcholine decreased significantly (-LogPC(200): 4.006 +/- 0.554 vs 2.059 +/- 0.262; P < 0.01). Bronchial alveolar lavage fluid (BALF) and lung tissue specimen analysis indicated that airway allergic inflammation was present.. The study demonstrates a dissociation between the bronchoconstrictor response and bronchial hyper-responsiveness and indicates that multiple ovalbumin exposures induces persistent bronchoconstriction with airway hypo-responsiveness despite airway allergic inflammation.

Topics: Acetylcholine; Airway Resistance; Allergens; Animals; Bronchi; Bronchial Hyperreactivity; Bronchoconstriction; Lung; Male; Ovalbumin; Rats; Rats, Sprague-Dawley

Release of human lung mast cell tryptase may be important in the pathophysiology of asthma. We examined the effect of the reversible, nonelectrophilic tryptase inhibitor MOL 6131 on airway inflammation and hyper-reactivity in a murine model of asthma. MOL 6131 is a potent selective nonpeptide inhibitor of human lung mast cell tryptase based upon a beta-strand template (K(i) = 45 nM) that does not inhibit trypsin (K(i) = 1,061 nM), thrombin (K(i) = 23, 640 nM), or other serine proteases. BALB/c mice after i.p. OVA sensitization (day 0) were challenged intratracheally with OVA on days 8, 15, 18, and 21. MOL 6131, administered days 18-21, blocked the airway inflammatory response to OVA assessed 24 h after the last OVA challenge on day 22; intranasal delivery (10 mg/kg) had a greater anti-inflammatory effect than oral delivery (10 or 25 mg/kg) of MOL 6131. MOL 6131 reduced total cells and eosinophils in bronchoalveolar lavage fluid, airway tissue eosinophilia, goblet cell hyperplasia, mucus secretion, and peribronchial edema and also inhibited the release of IL-4 and IL-13 in bronchoalveolar lavage fluid. However, tryptase inhibition did not alter airway hyper-reactivity to methacholine in vivo. These results support tryptase as a therapeutic target in asthma and indicate that selective tryptase inhibitors can reduce allergic airway inflammation.

Topics: Animals; Asthma; Bridged Bicyclo Compounds, Heterocyclic; Bronchial Diseases; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Movement; Cytokines; Eosinophils; Humans; Inflammation; Lung; Mice; Mice, Inbred BALB C; Models, Molecular; Mucus; Ovalbumin; Piperidines; Pulmonary Edema; Pulmonary Eosinophilia; Serine Endopeptidases; Serine Proteinase Inhibitors; Tryptases; Vascular Cell Adhesion Molecule-1

CTLA-4 (CD152) expression is restricted to subsets of activated T lymphocytes and shares homology with CD28. CTLA-4 and CD28 molecules both bind to B7 molecules on antigen-presenting cells. Whereas CD28-B7 interaction enhances T cell activation, cytokine production and survival, CTLA-4 signaling down-regulates T cell responses. Here, we studied the involvement of CTLA-4 triggering in the pathogenesis of allergen-induced airway inflammation in mice. Anti-CTLA-4 mAb were injected during i.p. sensitization with ovalbumin (OVA). This treatment favored OVA-specific IgE production and augmented blood eosinophilia in BALB/c mice. In BALB/c mice, enhanced Th2 sensitization after anti-CTLA-4 mAb injections resulted in more severe airway inflammation, and increased airway hyperresponsiveness to metacholine, bronchial eosinophilia and IL-4 and IL-5 levels in broncho-alveolar lavage (BAL) fluid following repeated allergen inhalations. Importantly, aggravation of airway inflammation and enhancement of Th2 responses were accompanied by a significant reduction of pulmonary TGF-beta levels at protein level in BAL fluid as well as on mRNA level in inflamed lung tissue. In contrast to BALB/c mice, blockade of CTLA-4 did not alter IgE production nor the phenotype of airway inflammation or TGF-beta production in C57BL/6 mice. Our data suggest that CTLA-4 triggering represents an important regulatory mechanism for Th2 sensitization in genetically predisposed mice by modulating TGF-beta production.

Topics: Abatacept; Animals; Antibodies, Monoclonal; Antigens, CD; Antigens, Differentiation; Asthma; Bronchial Hyperreactivity; CTLA-4 Antigen; Disease Models, Animal; Eosinophilia; Immunization; Immunoconjugates; Immunoglobulin E; Lymphocyte Activation; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; RNA, Messenger; Signal Transduction; Species Specificity; T-Lymphocytes; Th2 Cells; Transforming Growth Factor beta

CCR3 is a chemokine receptor initially thought specific to eosinophils but subsequently identified on TH2 cell subsets, basophils, mast cells, neural tissue, and some epithelia. Because of the prominent role of these cells in allergic disease, including asthma, we generated mice deficient in CCR3 to determine its contribution in a model of allergic airway disease. Here we show that CCR3 is important for the basal trafficking of eosinophils to the intestinal mucosa but not the lung. In contrast, CCR3 disruption significantly curtails eosinophil recruitment to the lung after allergen challenge, with the majority of the eosinophils being arrested in the subendothelial space. Further, a role for CCR3 in mast cell homing has been identified; after sensitization and allergen challenge, we find increased numbers of intraepithelial mast cells in the trachea of knockout mice. Physiologically, we find that the net result of these complex cell fates after sensitization and allergen challenge is a paradoxical increase in airway responsiveness to cholinergic stimulation. These data underscore a more complex role for CCR3 in allergic disease than was anticipated.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Homeostasis; Inflammation; Lung; Mast Cells; Mice; Mice, Knockout; Ovalbumin; Receptors, CCR3; Receptors, Chemokine

Repeated aerosolized antigen challenges to brown Norway (BN) rats generate nonspecific airway hyperresponsiveness (AHR). On the other hand, some studies have demonstrated that repeated antigen challenge could attenuate antigen-specific AHR in BN rats. The authors questioned whether such dissociation in airway responses actually occurs when assessed in a single study in the same animals. The authors simultaneously measured AHR to methacholine and antigen-specific AHR in rats that were repeatedly exposed to aerosolized ovalbumin (OA) for 1 or 3 months after sensitization. Four days after the last challenge, airway responses to methacholine and OA, morphometry of the airways, the cell profile in bronchoalveolar lavage fluid, and cytokine messenger ribonucleic acid (mRNA) expression in the lungs were evaluated. The two types of AHR were modulated in opposite directions by repeated antigen challenges. The AHR to methacholine was significantly increased in the rats receiving antigen challenges compared with the control rats receiving saline challenges after sensitization; whereas, the antigen-specific AHR was significantly decreased. The number of alveolar macrophages in lavaged fluid and the expression of transforming growth factor-beta1 mRNA in lung tissue was significantly different between the antigen-challenged rats and the control rats. In conclusion, dissociation between nonspecific airway hyperresponsiveness and antigen-specific airway hyperresponsiveness in brown Norway rats after repeated antigen challenges was demonstrated. Sustained airway inflammation with macrophages and/or upregulation of transforming growth factor-beta1 messenger ribonucleic acid in the lung tissue may be responsible for this dissociation.

Topics: Aerosols; Animals; Antigens; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Male; Methacholine Chloride; Ovalbumin; Rats; Rats, Inbred BN; Specific Pathogen-Free Organisms

Pretreatment with mycobacterial Ags has been shown to be effective in preventing allergic airway inflammation from occurring in a mouse model. Because most asthmatics are treated after the development of asthma, it is crucial to determine whether mycobacterial Ags can reverse established allergic airway inflammation in the presensitized state. Our hypothesis, based upon our previous findings, is that mycobacteria treatment in presensitized mice will suppress the allergic airway inflammation with associated clinical correlates of established asthma, with the noted exception of factors associated with the early allergic response (EAR). BALB/c mice sensitized and challenged with OVA were evaluated for pulmonary functions during both the EAR and late allergic response, and airway hyperresponsiveness to methacholine. Following this, sensitized mice were randomized and treated with placebo or a single dose (1 x 10(5) CFUs) of bacillus Calmette-Guérin (BCG) or Mycobacterium vaccae via nasal or peritoneal injection. One week later, the mice were rechallenged with OVA and methacholine, followed by bronchoalveolar lavage (BAL) and tissue collection. Mice treated with intranasal BCG were most significantly protected from the late allergic response (p < 0.02), airway hypersensitivity (p < 0.001) and hyperreactivity (p < 0.05) to methacholine, BAL (p < 0.05) and peribronchial (p < 0.01) eosinophilia, and BAL fluid IL-5 levels (p < 0.01) as compared with vehicle-treated, sensitized controls. Intranasal M. vaccae treatment was less effective, suppressing airway hypersensitivity (p < 0.01) and BAL eosinophilia (p < 0.05). No changes were observed in the EAR, BAL fluid IL-4 levels, or serum total and Ag-specific IgE. These data suggest that mycobacterial Ags (BCG>>M. vaccae) are effective in attenuating allergic airway inflammation and associated changes in pulmonary functions in an allergen-presensitized state.

Topics: Administration, Intranasal; Airway Resistance; Animals; Antigens, Bacterial; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Female; Hypersensitivity, Immediate; Immunoglobulin E; Interferon-gamma; Interleukin-5; Kinetics; Leukocyte Count; Lung; Mice; Mice, Inbred BALB C; Mycobacterium; Mycobacterium bovis; Ovalbumin; Pulmonary Eosinophilia

The effects of silymarin on bronchoconstriction induced by antigen challenge and on post-antigen challenge hyperresponsiveness to substance P were evaluated in sensitized guinea-pigs. Silymarin significantly decreased the bronchoconstriction due to antigen administration in the early phase of the response. In contrast, the dose-response curve for substance P recorded 1 h after antigen challenge was not modified by pretreatment with silymarin. The influence of the flavonoid on hyperresponsiveness to histamine in propranolol- and PAF (platelet-activating factor)-treated animals was also assessed. Silymarin did not affect hyperresponsiveness to histamine induced by either propranolol or PAF although it had inhibitory activity on the bronchial contractile response to the autacoid. These results suggest that silymarin has a protective effect in the early phase of allergic asthma, an effect, which may be related to a negative influence of the flavonoid on bronchial responsiveness to histamine.

Topics: Animals; Bronchial Hyperreactivity; Bronchoconstriction; Guinea Pigs; Histamine; Male; Ovalbumin; Platelet Activating Factor; Propranolol; Protective Agents; Silymarin; Substance P; Time Factors

Evidence has shown that aminophylline has bronchoprotective, anti-inflammatory, and immunomodulatory effects. Our purpose was to evaluate the effect of different doses of aminophylline on the late-phase reaction, bronchial hyperresponsiveness (BHR) and T cell-related cytokine mRNA expression in brown Norway rats induced by ovalbumin (OA) sensitization.. Forty rats were equally divided into four groups. Groups I, II, and III animals were sensitized and subsequently provoked with OA. Aminophylline 25 mg/kg was given intraperitoneally to the group I animals and 5 mg/kg to group II animals. Group III animals received intraperitoneal normal saline. Group IV breathed aerosolized saline as a control. After OA provocation, the animals were anesthetized. Pulmonary function tests were performed at baseline and after varying doses of acetylcholine. Thereafter, bronchoalveolar lavage was performed and the lungs were examined histologically. Total RNA was extracted from lung tissue and reverse transcriptase-polymerase chain reaction was performed using primers for interleukin (IL)-2, IL-4, IL-5, IL-6, IL-10, interferon-gamma, inducible nitric oxide synthase, and beta-actin.. Group III had worse pulmonary function tests, more severe BHR, and more severe lung inflammation, higher IL-4 and IL-10 cytokine levels in bronchoalveolar lavage fluid, and higher IL-4, IL-5, IL-6, IL-10, tumor necrosis factor-alpha and inducible nitric oxide synthase mRNA expression than the other three groups. Expression of IL-2 and interferon-gamma was significantly reduced in group III.. Both low and high dose aminophylline are effective in preventing late-phase bronchoconstriction, BHR, and an inflammatory response. Aminopylline decreases T helper cell 2-related cytokine mRNA expression but increases T helper cell 1-related cytokines mRNA expression.

Topics: Allergens; Aminophylline; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Lung; Ovalbumin; Rats; Respiratory Function Tests; RNA, Messenger; Th2 Cells; Theophylline

We examined the effects of different immunomodulators administered topically on asthmatic responses in a rat model of asthma. Sensitised Brown-Norway rats were administered rapamycin, SAR943 (32-deoxorapamycin), IMM125 (a hydroxyethyl derivative of D-serine(8)-cyclosporine), and budesonide by intratracheal instillation 1 h prior to allergen challenge. Allergen exposure induced bronchial hyperresponsiveness, accumulation of inflammatory cells in bronchoalveolar lavage fluid, and also an increase in eosinophils and CD2+, CD4+ and CD8+ T cells in the airways. Interleukin-2, interleukin-4, interleukin-5, interleukin-10, and interferon-gamma mRNA expression was upregulated by allergen exposure. Budesonide abolished airway inflammation, suppressed the mRNA expression for interleukin-2, interleukin-4, and interleukin-5 (P<0.03), and bronchial hyperresponsiveness (P<0.05). IMM125 suppressed airway infiltration of eosinophils, and CD8+ T cells (P<0.02), and prevented the upregulated mRNA expression for interleukin-4, interleukin-5, and interferon-gamma (P<0.02). Rapamycin suppressed CD8+ T cell infiltration in airway submucosa (P<0.03), and mRNA expression for interleukin-2 (p<0.002), while SAR943 suppressed interleukin-2, interleukin-4, and interferon-gamma mRNA (P<0.05). IMM125, rapamycin and SAR943 did not alter airway submucosal CD2+ and CD4+ T cell infiltration, and bronchial hyperresponsiveness. CD8+ T cells, in contrast to CD4+ T cells, are more susceptible to the inhibition by IMM125 and rapamycin, which also caused greater suppression of Th1 compared to Th2 cytokine mRNA expression. In this acute model of allergic inflammation, differential modulation of Th1 and Th2 cytokines may determine the effects of various immunomodulators on airway inflammation and bronchial hyperresponsiveness.

Topics: Acetylcholine; Administration, Topical; Animals; Asthma; Bronchial Hyperreactivity; Bronchodilator Agents; Budesonide; Cyclosporins; Cytokines; Disease Models, Animal; Gene Expression Regulation; Immunosuppressive Agents; Inflammation; Male; Ovalbumin; Rats; Rats, Inbred BN; RNA, Messenger; Sirolimus; Specific Pathogen-Free Organisms; T-Lymphocytes; Vasodilator Agents

This study investigated the capacity of conjugated linoleic acids (CLA) to reduce ex vivo antigen-induced release of eicosanoids in a type I hypersensitivity model. Guinea pigs were fed a diet containing 0.25% safflower oil (control) or 0.25% CLA [43% trans (t)10, cis (c)12; 41% c9, t11/t9, c11 18:2] for 2 wk before and during sensitization to ovalbumin (OVA). Lungs, tracheas, and bladders were incubated in physiological saline solution (PSS) for 1 h (basal mediator release) and challenged with OVA (0.01 g/l PSS) for 1 h (mediator release in response to antigen). Eicosanoids were quantified by HPLC/tandem mass spectrometry or enzyme immunoassay. CLA feeding resulted in no change in basal release but decreased eicosanoid release from sensitized tissues in response to antigen challenge in the following manner: thromboxane B(2), 6-keto-prostaglandin (PG)F(1alpha), PGF(2alpha), PGD(2), PGE(2) by 57-75% in lung, 45-65% in trachea, and 38-60% in bladder; and leukotriene C(4)/D(4)/E(4) by 87, 90, and 50% in lung, trachea, and bladder, respectively. These data indicate that feeding CLA reduces lipid-derived inflammatory mediators produced by this type I hypersensitivity model.

Topics: 6-Ketoprostaglandin F1 alpha; Animals; Blotting, Western; Bronchial Hyperreactivity; Cyclooxygenase 2; Dinoprost; Dinoprostone; Eicosanoids; Female; Guinea Pigs; Hypersensitivity, Immediate; Isoenzymes; Linoleic Acid; Ovalbumin; Prostaglandin D2; Prostaglandin-Endoperoxide Synthases; Trachea; Urinary Bladder

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Chromosome Mapping; Eosinophils; Immunization; Immunoglobulin E; Mice; Mice, Congenic; Mice, Inbred BALB C; Ovalbumin

Airway obstruction, hyperresponsiveness, and the accumulation and persistence within the airways of inflammatory cells characterize asthma. Interleukin (IL)-3, granulocyte macrophage colony- stimulating factor (GM-CSF), and IL-5 are among several cytokines that have been shown to be increased in asthma and to contribute to atopic inflammation. They mediate their effect via receptors that have a common beta subunit (beta(c)). We hypothesized that blocking of this common beta(c) would impair the airway response to antigen. We report that an antisense (AS) phosphorothioate oligodeoxynucleotide (ODN) found to specifically inhibit transcription of the beta(c) in rat bone marrow cells also caused inhibition of beta(c) mRNA expression and of immunoreactive cells within the lungs of Brown Norway (BN) rats when injected intratracheally (p < 0.01). Inhibition of beta(c) significantly reduced (p < 0.01) experimentally induced eosinophilia in vivo in ovalbumin (OVA)-sensitized BN rats after antigen challenge. Furthermore, when compared with mismatch-treated rats, beta(c) AS-ODN caused inhibition of antigen-induced airway hyperresponsiveness to leukotriene D4. Taken together, our findings demonstrate that the common beta(c) of IL-3, IL-5, and GM-CSF receptors is involved in the eosinophil influx and airway hyperresponsiveness that follow OVA challenge and underscore the potential utility of a topical antisense approach targeting beta(c) for the treatment of asthma.

Topics: Animals; Antigens; Asthma; Bone Marrow Cells; Bronchial Hyperreactivity; Cell Count; Cells, Cultured; Dose-Response Relationship, Drug; Eosinophils; Immunization; Leukotriene D4; Lung; Male; Oligonucleotides, Antisense; Ovalbumin; Rats; Rats, Inbred BN; Receptors, Granulocyte-Macrophage Colony-Stimulating Factor; Receptors, Interleukin; Receptors, Interleukin-3; Receptors, Interleukin-5; RNA, Messenger; Transcription, Genetic

Type I diabetes is associated with a low incidence of asthma. We tested whether a decrease in sensory neuropeptide release is associated with an attenuated bronchoconstrictive response to field stimulation (FS; 100 stimuli, 20 V, 0.1 ms, 20 Hz) in streptozotocin (STZ)-induced diabetes. The organ fluid of the preparations were also tested for substance P, calcitonin gene-related peptide (CGRP), and somatostatin concentrations by RIA. Preparations were from either normal rats or those pretreated with 50 mg/kg STZ iv 8 wk before experiment. A group of STZ-treated animals was supplied with insulin delivery (4 IU/day sc) implants between 4 and 8 wk. A subgroup was formed to study the effect of capsaicin desensitization. The atropine-resistant contraction was attenuated by diabetes without capsaicin-sensitive relaxation response. Exogenous CGRP and substance P potentiated, whereas somatostatin inhibited (1 nM-10 microM) the FS-induced contractions in rings from either group. FS released somatostatin, CGRP, and substance P from 0.17 +/- 0.024, 0.15 +/- 0.022, and 1.65 +/- 0.093 to 0.58 +/- 0.032, 0.74 +/- 0.122, and 5.34 +/- 0.295 in preparations from normal, and from 0.19 +/- 0.016, 0.11 +/- 0.019, and 0.98 +/- 0.116 to 0.22 +/- 0.076, 0.34 +/- 0.099, and 1.84 +/- 0.316 fmol/mg wet wt in preparations from diabetic rats. Insulin supplementation restored neuropeptide release in rings from STZ-treated rats. The results show that the decreased FS-induced contractions occurred with a decrease in sensory neuropeptide release in STZ-diabetic rats.

Topics: Animals; Blood Glucose; Body Weight; Bronchial Hyperreactivity; Bronchoconstriction; Calcitonin Gene-Related Peptide; Capsaicin; Diabetes Mellitus, Experimental; Diabetes Mellitus, Type 1; Guinea Pigs; Insulin; Male; Neural Conduction; Neurons, Afferent; Ovalbumin; Rats; Rats, Wistar; Somatostatin; Substance P; Trachea

Asthma is an acute-on-chronic inflammatory disease of the airways, characterized by airflow obstruction and hyper-reactivity of the airways to a variety of stimuli. Chronic asthma is associated with remodeling of the airway wall, which may contribute to hyper-reactivity and fixed airflow obstruction. We used an improved mouse model of chronic asthma to investigate the role of CD4(+) T-lymphocytes in airway remodeling and hyper-reactivity. Animals functionally depleted of CD4(+) T-lymphocytes by repeated administration of a monoclonal antibody exhibited markedly decreased airway responsiveness. In addition, these mice had greatly diminished subepithelial fibrosis, epithelial thickening, and mucous cell hyperplasia/metaplasia. Chronic inflammation in the airway wall was moderately reduced, with a marked decrease in the accumulation of immunoglobulin-synthesizing plasma cells. However, intraepithelial accumulation of eosinophils was not significantly inhibited and airway epithelial expression of eotaxin was undiminished. This work provides the first experimental evidence that CD4(+) T-lymphocytes play a crucial role in the pathogenesis of the lesions of chronic asthma and lends support to the notion that functional inhibition of these cells may be an important therapeutic target.

Topics: Allergens; Animals; Antibodies, Monoclonal; Asthma; Bronchi; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Chronic Disease; Disease Models, Animal; Female; Image Processing, Computer-Assisted; Immunization; Immunoglobulin G; Immunohistochemistry; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Specific Pathogen-Free Organisms

Sensory neuropeptides are localized to airway nerves and endocrine cells in both human and animal species and may participate in the development of airway inflammation and hyperresponsiveness (AHR). We used a mouse model to identify the changes that occur in calcitonin gene-related peptide (CGRP) expression in the airways during development of allergic inflammation and to investigate the potential role of this neuropeptide in modulating AHR. In sensitized mice, allergen challenge induced eosinophilic airway inflammation and AHR and resulted in considerable depletion of CGRP in neuroepithelial bodies and submucosal nerve plexuses without altering the overall density of airway nerve fibers. This depletion was subsequent to the development of airway inflammation and was prevented by anti-very late antigen-4 and anti-interleukin-5 treatments, which blocked airway eosinophilia and abolished AHR. Administration of CGRP to sensitized and challenged mice resulted in the normalization of airway responsiveness to inhaled methacholine, an effect that was neutralized by the receptor antagonist CGRP(8-37). These data demonstrate that replacement of CGRP following its depletion in allergic mice can reverse the changes in airway responsiveness and suggest that CGRP may have potential for the treatment of allergic AHR.

Topics: Allergens; Animals; Antibodies; Bronchi; Bronchial Hyperreactivity; Calcitonin Gene-Related Peptide; Electric Stimulation; Eosinophils; Immunization; Immunohistochemistry; Inflammation; Integrin alpha4beta1; Integrins; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Muscle, Smooth; Ovalbumin; Receptors, Lymphocyte Homing; Respiratory Hypersensitivity

Nonselective cyclooxygenase (COX) inhibition during allergic sensitization with ovalbumin in a murine model leads to an increase in the Type 2 cytokines interleukin-5 (IL-5) and IL-13; however, the effect of selective COX-1 and COX-2 inhibitors on these cytokines is unknown. We found that COX-1 protein was constitutively expressed in lung tissue. Expression of COX-1 protein did not increase with ovalbumin sensitization, but expression of COX-2 protein did. Ovalbumin-sensitized mice treated with either selective COX-1 inhibitor SC58560 (OVA-COX-1 inhibitor) or selective COX-2 inhibitor SC58236 (OVA-COX-2 inhibitor) had significantly greater airway hyperresponsiveness (p < 0.05) and higher levels of IL-13 (p < 0.05) in lung supernatants than did untreated mice that were ovalbumin sensitized (OVA). Lung mRNA levels for the chemokine receptors CCR1 through CCR5 (expressed on eosinophils, basophils, lymphocytes, and dendritic cells) were increased in the OVA-COX-2 inhibitor and OVA-indomethacin groups. We conclude that in the BALB/c mouse, COX inhibition with either a COX-1 or COX-2 inhibitor during allergen sensitization augments production of IL-13 and increases airway hyperresponsiveness.

Topics: Animals; Arachidonic Acid; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cyclooxygenase 1; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Cyclooxygenase Inhibitors; Dinoprostone; Female; Immunization; Interleukins; Isoenzymes; Lung; Membrane Proteins; Methacholine Chloride; Mice; Mice, Inbred BALB C; Organic Chemicals; Ovalbumin; Prostaglandin-Endoperoxide Synthases; Pyrazoles; Receptors, Chemokine; Sulfonamides

Correcting Th1/Th2 imbalance with administration of IL-12 before and during antigen challenge holds therapeutic promise in asthma. However, the effects of IL-12 on the established asthmatic responses have not fully been examined.. We investigated whether IL-12 administered after antigen challenge could diminish airway hyper-reactivity (AHR) and eosinophilia in mice actively sensitized to ovalbumin. We also have investigated the ability of administered IL-12 to induce IL-18 receptor (IL-18R) expression that may lead possible synergic action of IL-12 with endogenous IL-18.. C57BL/6 mice immunized to ovalbumin (OVA) by intraperitoneal (i.p.) injection, were challenged three times with an aerosol of OVA every second day for 8 days. Recombinant IL-12 (500 ng) was intravenously administered on a single occasion 1 h after the final challenge of mice. Mice were analysed for effects of IL-12 on AHR, inflammatory cell infiltration and cytokine levels in lung tissue as well as serum immunoglobulin (Ig) E levels. Immunohistochemistry for IL-18R was performed using rat monoclonal antibody specific for murine IL-18Ralpha (IL-1 receptor related protein; IL-1Rrp).. An intravenous IL-12 administration diminished AHR, pulmonary eosinophilia and T lymphocyte infiltration, serum IgE, IL-4 and IL-13 in lung tissue. Expression of IL-18R was induced in the mononuclear cells in the lung of mice exposed to OVA. IL-12 administration enhanced the IL-18R expression compared with the control.. These data indicate that IL-12 can attenuate established antigen-induced AHR and inflammation. In this mechanism it would be interpreted as follows: IL-12 administration in OVA-challenged mice decreased IL-4 production and IgE production thereafter through direct effect on inhibiting the activation of established Th2 cells response and also combined effect with up-regulation of IL-18R expression by inflammatory cells in the lung.

Topics: Administration, Inhalation; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Immunoglobulin E; Interleukin-12; Interleukin-18 Receptor alpha Subunit; Lung; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Pulmonary Eosinophilia; Receptors, Interleukin; Receptors, Interleukin-18

Allergic asthma is an inflammatory disease of the airways characterized by eosinophilic inflammation and airway hyper-reactivity. Cytokines and chemokines specific for Th2-type inflammation predominate in asthma and in animal models of this disease. The role of Th1-type inflammatory mediators in asthma remains controversial. IFN-gamma-inducible protein 10 (IP-10; CXCL10) is an IFN-gamma-inducible chemokine that preferentially attracts activated Th1 lymphocytes. IP-10 is up-regulated in the airways of asthmatics, but its function in asthma is unclear. To investigate the role of IP-10 in allergic airway disease, we examined the expression of IP-10 in a murine model of asthma and the effects of overexpression and deletion of IP-10 in this model using IP-10-transgenic and IP-10-deficient mice. Our experiments demonstrate that IP-10 is up-regulated in the lung after allergen challenge. Mice that overexpress IP-10 in the lung exhibited significantly increased airway hyperreactivity, eosinophilia, IL-4 levels, and CD8(+) lymphocyte recruitment compared with wild-type controls. In addition, there was an increase in the percentage of IL-4-secreting T lymphocytes in the lungs of IP-10-transgenic mice. In contrast, mice deficient in IP-10 demonstrated the opposite results compared with wild-type controls, with a significant reduction in these measures of Th2-type allergic airway inflammation. Our results demonstrate that IP-10, a Th1-type chemokine, is up-regulated in allergic pulmonary inflammation and that this contributes to the airway hyperreactivity and Th2-type inflammation seen in this model of asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Chemokine CXCL10; Chemokine CXCL11; Chemokine CXCL9; Chemokines, CXC; Disease Models, Animal; Inflammation; Injections, Intraperitoneal; Intercellular Signaling Peptides and Proteins; Interferon-gamma; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Ovalbumin; Up-Regulation

CD8+T cells can suppress allergen-induced late airway responses (LARs) and airway inflammation.. To test the hypothesis that the suppression of LARs and airway eosinophilia by CD8+T cells is IFN-gamma mediated, we tested the effects of adoptively transferred CD8+T cells, in which IFN-gamma synthesis was inhibited by an antisense (AS) oligodeoxynucleotide (ODN), on the airway responses of a rat model of allergic asthma.. CD8+T cells were harvested from the cervical lymph nodes of ovalbumin (OVA)-sensitized Brown Norway rats for administration to other actively sensitized syngeneic rats. CD8+T cells (2 x 10(6)) were incubated for 6 hours with 2 micromol/L AS ODN or sense ODN and were injected intraperitoneally into recipients; inhibition of IFN-gamma expression in vitro by AS ODN was shown by means of flow cytometry. Two days later, rats were challenged with aerosolized OVA.. OVA-induced LAR and bronchoalveolar lavage (BAL) fluid eosinophilia were suppressed by sense ODN-treated CD8+T cells. IFN-gamma expression in BAL cells was elevated in these animals. IFN-gamma expression in BAL cells was at control levels in recipients of AS ODN-treated CD8+ cells, confirming the success of the AS treatment in vivo. BAL eosinophilia was also largely restored in the AS ODN treatment group. In contrast, the CD8+T cell-induced suppression of the LAR was not significantly affected by AS ODN pretreatment.. These results indicate that CD8+T cells inhibit airway eosinophilia through secretion of IFN-gamma but may suppress the LAR by means of other mechanisms.

Topics: Adoptive Transfer; Allergens; Animals; Bronchi; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; CD8-Positive T-Lymphocytes; Eosinophilia; Interferon-gamma; Male; Oligonucleotides, Antisense; Ovalbumin; Rats; Rats, Inbred BN; Time Factors

Activated T cells, through the release of specific cytokines (ie, IL-4, IL-5, and IL-13), regulate effector cell recruitment and function. In this way T cells orchestrate the inflammatory response, which leads to airway hyperresponsiveness (AHR), a cardinal feature of allergic asthma.. In the present study the direct role of T cells and, in particular, the importance of signal transducer and activator of transcription 6 (STAT6) in T cells was investigated in the development of AHR.. In a murine model of allergen-driven AHR, the effects of adoptive transfer of STAT6-containing (STAT6+/+) and STAT6-deficient (STAT6-/-) T cells from sensitized mice into allergen-challenged mice were tested.. Although greater in STAT6+/+ mice, both allergen-challenged STAT6+/+ and STAT6-/- mice had AHR after transfer of T cells from sensitized STAT6+/+ mice. In contrast, AHR did not develop in allergen-challenged STAT6-/- mice after transfer of T cells from sensitized STAT6-/- mice. Reconstitution of AHR after T-cell transfer was not associated with airway eosinophilia.. The data indicate that the STAT6 status of the donor mice is critical to the development of AHR. Although not critical for the development of AHR, the STAT6 status of the recipient mice might play a contributory-regulatory role in the AHR response. The results identify a STAT6-dependent T-cell pathway capable of modulating airway responsiveness, even in the absence of a significant airway eosinophilia.

Topics: Adoptive Transfer; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Eosinophilia; Female; Immunization; Male; Mice; Mice, Knockout; Ovalbumin; STAT6 Transcription Factor; T-Lymphocytes; Trans-Activators

1. A deficiency of constitutive nitric oxide synthase (cNOS)-derived nitric oxide (NO), due to reduced availability of L-arginine, importantly contributes to allergen-induced airway hyperresponsiveness (AHR) after the early asthmatic reaction (EAR). Since cNOS and arginase use L-arginine as a common substrate, we hypothesized that increased arginase activity is involved in the allergen-induced NO deficiency and AHR. 2. Using a guinea-pig model of allergic asthma, we addressed this hypothesis by examining the effects of the specific arginase inhibitor N(omega)-hydroxy-nor-L-arginine (nor-NOHA) on the responsiveness to methacholine of isolated perfused tracheae from unchallenged control animals and from animals 6 h after ovalbumin challenge. Arginase activity in these preparations was investigated by measuring the conversion of L-[14C]arginine to [14C]urea. 3. Airways from allergen-challenged animals showed a 2 fold (P<0.001) increase in responsiveness to intraluminal (IL) administration of methacholine compared to controls. A similar hyperresponsiveness (1.8 fold, P<0.01) was observed in control airways incubated with the NOS inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME, 0.1 mM, IL), while L-NAME had no further effect on the airways from challenged animals. 4. Remarkably, 5 microM nor-NOHA (IL) normalized the hyperresponsiveness of challenged airways to basal control (P<0.001), and this effect was fully reversed again by 0.1 mM L-NAME (P<0.05). Moreover, arginase activity in homogenates of the hyperresponsive airways was 3.5 fold (P<0.001) enhanced compared to controls. 5. The results indicate that enhanced arginase activity contributes to allergen-induced deficiency of cNOS-derived NO and AHR after the EAR, presumably by competition with cNOS for the common substrate, L-arginine. This is the first demonstration that arginase is involved in the pathophysiology of asthma.

Topics: Allergens; Animals; Arginase; Arginine; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Enzyme Inhibitors; Guinea Pigs; In Vitro Techniques; Methacholine Chloride; Models, Animal; NG-Nitroarginine Methyl Ester; Nitric Oxide; Nitric Oxide Synthase; Ovalbumin; Trachea

Smooth muscle contraction is one of the hallmarks of asthma. A recently developed pyridine derivative, Y-27632, a selective Rho kinase inhibitor, has been reported to inhibit the smooth muscle contraction of human and animal trachea in ex vivo systems but its effect in animal models of airway hyperresponsiveness (AHR) has not been examined. The purpose of this study was to evaluate the effect of Y-27632 in a murine model of allergic and virally induced AHR.. Baseline lung resistance and methacholine induced AHR were measured in mice sensitised to ovalbumin (OVA) and also in mice infected with respiratory syncytial virus (RSV) following ovalbumin sensitisation (OVA/RSV).. Time course and dose ranging experiments indicated that 30 mg/kg Y-27632 given by gavage 2 hours before methacholine challenge significantly reduced baseline lung resistance and prevented AHR in OVA sensitised mice. Y-27632 also suppressed AHR induced by the bronchospastic agent serotonin in OVA sensitised mice and prevented methacholine induced AHR in OVA/RSV mice.. These results suggest that the signalling pathway mediated through Rho kinase may have an important role in bronchial smooth muscle tone in allergen induced and virus induced AHR and should be considered as a novel target for asthma treatment.

Topics: Airway Resistance; Animals; Antihypertensive Agents; Asthma; Benzopyrans; Bronchial Hyperreactivity; Dose-Response Relationship, Drug; Female; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Syncytial Virus Infections

Airway hyperresponsiveness, airway inflammation, and reversible airway obstruction are physiological hallmarks of asthma. These responses are increasingly being studied in murine models of antigen exposure and challenge, using whole body plethysmography to noninvasively assess airway hyperresponsiveness. This approach infrequently has been correlated with indexes of airway hyperresponsiveness measured by invasive means. Furthermore, correlation with quantitative histological data for tissue infiltration by inflammatory and immune cells, particularly in the wall of airways, during daily airway challenge is lacking. To address these uncertainties, we used C57BL/6 mice that were immunized with ovalbumin or vehicle (saline) and sensitized to aerosolized ovalbumin or vehicle 8 days later. The mice were subsequently exposed to aerosolized ovalbumin or vehicle, respectively, on days 14-22. We assessed airway hyperresponsiveness to methacholine noninvasively on days 14, 15, 18, or 22; we studied the same mice 24 h later while they were anesthetized for invasive analyses of airway hyperresponsiveness. Plasma total IgE concentration was significantly higher in the ovalbumin-treated mice compared with the vehicle-treated mice, but this did not correlate with eosinophil number. Peak airway hyperresponsiveness measured by either approach correlated early during daily antigen challenge (days 14 and 15), but this correlation was lost later during subsequent daily antigen challenges (days 18 and 22). On days 14 and 15, peak airway hyperresponsiveness correlated with transmigration of neutrophils and macrophages, but not lymphocytes, in the peribronchovascular connective tissue sheaths. This extravascular accumulation was found to be focal by three-dimensional microscopy. We conclude that, although ovalbumin treatment changed lung function in mice, correlation between noninvasive and invasive measures of peak airway hyperresponsiveness was inconsistent.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Immunoglobulin E; Macrophages, Alveolar; Male; Mice; Mice, Inbred C57BL; Neutrophils; Ovalbumin; Plethysmography; Pulmonary Artery; Pulmonary Veins; Respiratory Mechanics; Specific Pathogen-Free Organisms

We carried out a time course study on the development of allergen-induced airway remodeling in a mouse model of allergic asthma. Moreover, we examined the effect of allergen avoidance on the established airway remodeling.. BALB/c mice were sensitized to ovalbumin (OA) with alum, and exposed daily for 3 weeks to aerosolized OA. At each designated point, bronchial responsiveness was measured, and bronchoalveolar lavage and histological examination were carried out.. The numbers of inflammatory leukocytes in the airways and the percentage of goblet cells in the epithelium, Th2 cytokine production, IgE production, collagen deposition beneath the basement membrane and bronchial responsiveness to acetylcholine were all markedly increased after repeated antigen challenge for 1-3 weeks. In contrast, after cessation of antigen exposure, goblet cell hyperplasia, inflammatory infiltrates and bronchial hyperresponsiveness were gradually attenuated and had almost resolved 4 weeks after cessation, but subepithelial fibrosis was still observed at this time point.. The present findings demonstrated that epithelial changes following repeated allergen challenge are rapidly induced and recover after the cessation of exposure, but subepithelial fibrosis has a late onset and relatively irreversible changes, and subepithelial fibrosis in contrast to goblet cells hyperplasia did not appear to contribute to bronchial hyperresponsiveness, at least, in this mouse model.

Topics: Acetylcholine; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Collagen; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Lung; Mice; Mice, Inbred BALB C; Ovalbumin

Th2 T cell immune-driven inflammation plays an important role in allergic asthma. We studied the effect of counterbalancing Th1 T cells in an asthma model in Brown Norway rats that favors Th2 responses. Rats received i.v. transfers of syngeneic allergen-specific Th1 or Th2 cells, 24 h before aerosol exposure to allergen, and were studied 18-24 h later. Adoptive transfer of OVA-specific Th2 cells, but not Th1 cells, and OVA, but not BSA exposure, induced bronchial hyperresponsiveness (BHR) to acetylcholine and eosinophilia in a cell number-dependent manner. Importantly, cotransfer of OVA-specific Th1 cells dose-dependently reversed BHR and bronchoalveolar lavage (BAL) eosinophilia, but not mucosal eosinophilia. OVA-specific Th1 cells transferred alone induced mucosal eosinophilia, but neither BHR nor BAL eosinophilia. Th1 suppression of BHR and BAL eosinophilia was allergen specific, since cotransfer of BSA-specific Th1 cells with the OVA-specific Th2 cells was not inhibitory when OVA aerosol alone was used, but was suppressive with OVA and BSA challenge. Furthermore, recipients of Th1 cells alone had increased gene expression for IFN-gamma in the lungs, while those receiving Th2 cells alone showed increased IL-4 mRNA. Importantly, induction of these Th2 cytokines was inhibited in recipients of combined Th1 and Th2 cells. Anti-IFN-gamma treatment attenuated the down-regulatory effect of Th1 cells. Allergen-specific Th1 cells down-regulate efferent Th2 cytokine-dependent BHR and BAL eosinophilia in an asthma model via mechanisms that depend on IFN-gamma. Therapy designed to control the efferent phase of established asthma by augmenting down-regulatory Th1 counterbalancing mechanisms should be effective.

Topics: Administration, Inhalation; Adoptive Transfer; Allergens; Animals; Antibodies, Monoclonal; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Line; Epitopes, T-Lymphocyte; Inflammation; Injections, Intravenous; Interferon-gamma; Interleukin-4; Lung; Male; Ovalbumin; Pulmonary Eosinophilia; Rats; Rats, Inbred BN; RNA, Messenger; Th1 Cells; Th2 Cells

Vaccination with naked DNA encoding a specific allergen has been shown previously to prevent, but not reverse, the development of allergen-induced airway hyperresponsiveness (AHR). To enhance the effectiveness of DNA vaccine therapies and make possible the treatment of established AHR, we developed a DNA vaccination plasmid containing OVA cDNA fused to IL-18 cDNA. Vaccination of naive mice either with this fusion DNA construct or with an OVA cDNA-containing plasmid protected the mice from the subsequent induction of AHR. Protection from AHR correlated with increased IFN-gamma production and reduced OVA-specific IgE production. The protection appeared to be mediated by IFN-gamma and CD8(+) cells because treatment of mice with neutralizing anti-IFN-gamma mAb or with depleting anti-CD8 mAb abolished the protective effect. Moreover, vaccination of mice with preexisting AHR with the OVA-IL-18 fusion DNA, but not with the OVA cDNA plasmid, reversed established AHR, reduced allergen-specific IL-4, and increased allergen-specific IFN-gamma production. Thus, combining IL-18 cDNA with OVA cDNA resulted in a vaccine construct that protected against the development of AHR, and that was unique among cDNA constructs in its capacity to reverse established AHR.

Topics: Allergens; Animals; Antibodies, Blocking; Asthma; Bronchial Hyperreactivity; CD8-Positive T-Lymphocytes; Cytokines; Disease Models, Animal; Epitopes; Genetic Vectors; Immunoglobulin E; Immunosuppressive Agents; Injections, Intramuscular; Interferon-gamma; Interleukin-18; Interleukin-4; Mice; Mice, Inbred BALB C; Ovalbumin; Recombinant Fusion Proteins; Vaccines, DNA

Dendritic cells (DC) are the primary APC responsible for the capture of allergens in the airways and the shuttling of processed allergens to the draining lymph nodes where Ag presentation and T cell activation take place. The mechanism of this Ag handling and presentation in asthma is poorly understood. In addition, the feasibility of asthma induction by DC priming has not been directly tested. In this report an asthma model using intratracheally (i.t.) injected splenic DC was used to address these issues. DC pulsed with a model Ag OVA or the major MHC class II-restricted OVA T epitope peptide OVA(323-339) and instilled i.t. primed mice to exhibit asthma-like diseases. With OVA as the Ag, mice exhibit airway hyperresponsiveness (AHR), lung eosinophilia and inflammation, and pulmonary goblet cell hyperplasia. In OVA(323-339)-immunized mice, AHR and goblet cell hyperplasia were noted, with little eosinophilia and parenchymal inflammation. The latter finding provides evidence for dissociating AHR from eosinophilia. In both cases mediastinal node hypertrophy occurred, and high levels of Th2 cytokines were produced by the lung and mediastinal lymph node cells (LNC). Interestingly, mediastinal LNC also produced high levels of Th1 cytokines. Lung cells produced much less Th1 cytokines than these LNC. These results demonstrate that DC when introduced i.t. are potent in inducing asthma-like diseases by recruiting lymphocytes to the lung-draining lymph nodes and by stimulating Th2 responses and also suggest that the lung environment strongly biases T cell responses to Th2.

Topics: Animals; Antibody Specificity; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Movement; Cells, Cultured; Cytokines; Dendritic Cells; Female; Goblet Cells; Hyperplasia; Immunodominant Epitopes; Inflammation; Intubation, Intratracheal; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Ovalbumin; Peptide Fragments; Pulmonary Eosinophilia; T-Lymphocytes; Th1 Cells; Th2 Cells

Tobacco smoke (TS) exposure induces airway hyperreactivity, particularly in sensitive individuals with asthma. However, the mechanism of this airway hyperreactivity is not well understood. To investigate the relative susceptibility of atopic and nonatopic individuals to TS-induced airway hyperreactivity, we exposed ovalbumin (OA)-sensitized and nonsensitized guinea pigs to TS exposure (5 mg/l air, 30-min exposure, 7 days/wk for 120-156 days). Two similar groups exposed to compressed air served as controls. Airway reactivity was assessed as an increase in enhanced pause (Penh) units using a plethysmograph that allowed free movement of the animals. After 90 days of exposure, airway reactivity increased in OA-TS guinea pigs challenged with capsaicin, bradykinin, and neurokinin A fragment 4--10 aerosols. In addition, substance P content increased in lung perfusate of OA-TS guinea pigs in response to acute TS challenge compared with that of the other groups. Airway hyperirritability was not enhanced by phosphoramidon but was attenuated by a cocktail of neurokinin antagonists, nor was airway hyperreactivity observed after either methacholine or histamine challenge in OA-TS guinea pigs. Chronic TS exposure enhanced neither airway reactivity to histamine or methacholine nor contractility of isolated tracheal rings. In conclusion, chronic TS exposure increased airway reactivity to capsaicin and bradykinin aerosol challenge, and OA-TS guinea pigs were most susceptible to airway dysfunction as the result of exposure to TS compared with the other groups. Increased airway reactivity to capsaicin suggests a mechanism involving neurogenic inflammation, such as increased activation of lung C fibers.

Topics: Administration, Inhalation; Airway Resistance; Animals; Bradykinin; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Capsaicin; Guinea Pigs; Histamine; Leukocyte Count; Lung; Male; Methacholine Chloride; Neurokinin A; Ovalbumin; Peptide Fragments; Smoking; Substance P; Wakefulness

The development of allergic asthma is influenced by both genetic and environmental factors. Epidemiologic data often show no clear relationship between the levels of allergen and clinical symptoms. Recent data suggest that bacterial LPS may be a risk factor related to asthma severity. Airborne LPS is typically present at levels that are insufficient to activate alveolar macrophages in the absence of the accessory molecule LPS binding protein (LBP). LBP levels are markedly elevated in bronchoalveolar lavage fluids obtained from asthmatic subjects compared with those in normal controls. We hypothesized that LBP present in the lung could augment the pulmonary inflammation and airway reactivity associated with allergic asthma by sensitizing alveolar macrophages to LPS or other bacterial products and triggering them to release proinflammatory mediators. We compared wild-type (WT) and LBP-deficient mice using a defined Ag immunization and aerosol challenge model of allergic asthma. Immunized LBP-deficient mice did not develop substantial Ag-induced airway reactivity, whereas WT mice developed marked bronchoconstriction following aerosol Ag sensitization and challenge with methacholine. Similarly, production of NO synthase 2 protein and the NO catabolite peroxynitrite was dramatically higher in the lungs of WT mice following challenge compared with that in LBP-deficient mice. Thus, NO production appears to correlate with airway reactivity. In contrast, both mice developed similar pulmonary inflammatory cell infiltrates and elevated mucin production. Thus, LBP appears to participate in the development of Ag-induced airway reactivity and peroxynitrite production, but does not seem to be required for the development of pulmonary inflammation.

Topics: Acute-Phase Proteins; Adjuvants, Immunologic; Aerosols; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Carrier Proteins; Disease Models, Animal; Immunoglobulin E; Immunoglobulin G; Inflammation; Injections, Intraperitoneal; Lipopolysaccharides; Lung; Membrane Glycoproteins; Mice; Mice, Inbred BALB C; Mice, Knockout; Nitrates; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Ovalbumin

Thymus- and activation-regulated chemokine (TARC; CCL17) is a lymphocyte-directed CC chemokine that specifically chemoattracts CC chemokine receptor 4-positive (CCR4(+)) Th2 cells. To establish the pathophysiological roles of TARC in vivo, we investigated here whether an mAb against TARC could inhibit the induction of asthmatic reaction in mice elicited by OVA. TARC was constitutively expressed in the lung and was up-regulated in allergic inflammation. The specific Ab against TARC attenuated OVA-induced airway eosinophilia and diminished the degree of airway hyperresponsiveness with a concomitant decrease in Th2 cytokine levels. Our results for the first time indicate that TARC is a pivotal chemokine for the development of Th2-dominated experimental allergen-induced asthma with eosinophilia and AHR. This study also represents the first success in controlling Th2 cytokine production in vivo by targeting a chemokine.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Antibody Specificity; Asthma; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Movement; Chemokine CCL17; Chemokines, CC; Cytokines; Disease Models, Animal; Immune Sera; Immunohistochemistry; Immunosuppressive Agents; Injections, Intraperitoneal; Lung; Lymphocyte Activation; Male; Mice; Mice, Inbred C57BL; Ovalbumin; Pulmonary Eosinophilia; RNA, Messenger; Th2 Cells; Thymus Gland

Asthma may result from excessive Th-2 response in children not previously exposed to Th-1-inducing infections. We tested the hypothesis that BCG vaccination in Th-2-susceptible newborn BP2 mice blocks allergic inflammation and bronchial hyperreactivity (BHR). Ten day-old BP2 mice received 10(5) CFU of BCG 1173P2 intranasally (IN), and 6, 10 or 14 weeks thereafter were sensitized with 100 microg ovalbumin (OVA) in aluminium hydroxide twice subcutaneously (SC) at 1 week interval, and challenged 1 week after the second sensitization with 10 microg OVA IN. Compared to OVA-challenged unvaccinated mice, those that received BCG 8 weeks before challenge developed intense bronchial inflammation, BHR, and high IgE titers. Inflammation involved T cells, macrophages, dendritic cells and was accompanied by increased levels of Interleukin-5 (IL-5) in the bronchoalveolar lavages (BAL). However, animals challenged 16 weeks after BCG vaccination did not develop BHR nor bronchial hypereosinophilia, and showed reduced IgE levels. Bronchial infiltration by immunocompetent cells was also significantly reduced. Increased levels of gamma-interferon (IFN-gamma) after in vitro stimulation of tracheo-bronchial lymph node cells accompanied this blockage, but levels of IL-5 remained high. We demonstrate that 16 weeks after vaccination with BCG in newborn BP2 mice which have a high Th-2 background, allergic inflammation and BHR were blocked, even though a clear Th-1 shift was not achieved.

Topics: Animals; Animals, Newborn; Asthma; BCG Vaccine; Bronchial Hyperreactivity; Child; Cytokines; Dose-Response Relationship, Immunologic; Humans; Hypersensitivity; Hypersensitivity, Delayed; Hypersensitivity, Immediate; Inflammation; Job Syndrome; Lung; Male; Mice; Mycobacterium bovis; Ovalbumin; Phenotype; Th1 Cells; Th2 Cells; Time Factors

The relationship between CD4(+) T cell-mediated airway eosinophilic inflammation and bronchial hyper-responsiveness (BHR) was investigated. Ovalbumin-reactive T(h)0 clones were adoptively transferred to unprimed BALB/c mice and then the mice were challenged by inhalation of the relevant antigen. Upon antigen provocation, infused T(h) clones infiltrated into the airways, followed by the accumulation and degranulation of eosinophils, goblet cell hyperplasia, edema and increase in bronchial responsiveness to acetylcholine. Transfer of several clones that differed in the levels of IL-5 production revealed that the magnitude of in vivo eosinophilia strongly correlated with the IL-5-producing capacity of the infused T(h) clones. Administration of anti-IL-5 mAb almost completely suppressed antigen-induced eosinophilic inflammation and BHR. Administration of anti-IL-4 mAb or anti-IFN-gamma mAb enhanced the eosinophilia and BHR, whereas anti-IL-2 mAb did not affect them. The number of accumulated eosinophils significantly correlated with the intensity of BHR. Our present results clearly demonstrated that CD4(+) T cells induced BHR as a result of eosinophilic inflammation. IL-5 totally regulated both responses.

Topics: Acetylcholine; Animals; Antibodies, Monoclonal; Bronchial Diseases; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Clone Cells; Cytokines; Eosinophil Peroxidase; Eosinophilia; Interferon-gamma; Interleukin-4; Interleukin-5; Leukocyte Count; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Peroxidases; Pulmonary Eosinophilia; Specific Pathogen-Free Organisms

Airway hyperresponsiveness (AHR) is one of the characteristic features of human asthma. The presence of AHR and the precise mechanisms immediately after establishment of sensitization in guinea pigs are unclear, although there are many reports showing allergen exposure that causes an increase in bronchial responsiveness associated with eosinophil influx into the airway in sensitized guinea pigs.. We investigated the inhibitory effects on AHR to histamine of ONO-1078, a leukotriene antagonist; indomethacin, a cyclooxygenase inhibitor; S-145, a thromboxane A(2) (TXA(2)) antagonist, and Y-24180, a platelet-activating factor (PAF) antagonist, to assess the involvement of chemical mediators in AHR employing ovalbumin (OA) sensitized guinea pig models.. Male Hartley guinea pigs were used. Each group comprised 4-7 animals. The animals were sensitized to OA, injecting intraperitoneally 30 mg of cyclophosphamide and 2,000 microg of OA together with 100 mg of aluminum hydroxide as the adjuvant. The guinea pigs were artificially ventilated via a cannula using a small-animal respirator after intraperitoneal anesthesia with pentobarbital sodium for tracheotomy. The pressure at the airway opening (PAO) was measured using a differential pressure transducer, and a differential pressure of peak PAO (peak DeltaPAO) at inspiratory phase as an overall index of bronchial response to bronchoactive agents was used. While being artificially ventilated, the animals were exposed to physiological saline solution containing various concentrations of histamine (4.9, 9.8, 20, 39, 78, and 156 microg/ml) by inhalation for 30 s at 3-min intervals. Determinations were made at 1 min after each inhalation. The chemical mediators were each (30 mg/kg of ONO-1078, 3 mg/kg of S-1452, and 1 mg/kg of Y-24180) administered orally to sensitized guinea pigs, and the airway response to histamine was assessed. Each group comprised 4-7 animals.. The airway response to histamine was significantly greater in the sensitized group than in the nonsensitized group at histamine concentrations of 36 (p < 0.05), 78, and 156 mg/ml (p < 0.01). Leukotrienes C(4) and D(4): 30 mg/kg of ONO-178 did not show any inhibitory effect on airway response to inhaled histamine. Cyclooxygenase: 5 mg/kg of indomethacin did not show any inhibitory effect on the airway response to inhaled histamine. TXA(2): the AHR to inhaled histamine at doses of 9.8, 39, 78, and 156 microg/ml was significantly inhibited by prior administration of 3 mg/kg of S-1452. PAF: the AHR to inhaled histamine at doses of 9.8, 39, and 78 microg/ml was significantly inhibited by prior administration of 1 mg/kg of Y-24180.. S-1452 (3 mg/kg) and Y-24180 (1 mg/kg) significantly inhibited AHR to histamine, while ONO-108 (30 mg/kg) and indomethacin (5 mg/kg) did not. The results suggest that TXA(2) and PAF are involved in AHR in OA-sensitized guinea pigs.

Topics: Airway Resistance; Animals; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Dose-Response Relationship, Drug; Guinea Pigs; Histamine; Indomethacin; Leukotriene C4; Leukotriene D4; Male; Ovalbumin; Platelet Activating Factor; Probability; Reference Values; Sensitivity and Specificity; Thromboxane A2

To evaluate the role of CCR2 in allergic asthma, mutant mice deficient in CCR2 (CCR2(-/-)) and intact mice were sensitized with i.p. OVA with alum on days 0 and 7, and challenged by inhalation with nebulization of either OVA or saline. Airway hyperreactivity, measured by the methacholine-provoked increase in enhanced pause, was significantly increased (p < 0.05) in OVA-challenged CCR2(-/-) mutant mice, compared with comparably challenged CCR2(+/+) mice. OVA-challenged CCR2(-/-) mutants also were also found to have enhanced bronchoalveolar lavage fluid eosinophilia, peribronchiolar cellular cuffing, and Ig subclass switching, with increase in OVA-specific IgG(1) and IgE. In addition, RNase protection assay revealed increased whole lung expression of IL-13 in OVA-challenged CCR2(-/-) mutants. Unexpectedly, serum monocyte chemotactic protein-1 levels were 8-fold higher in CCR2(-/-) mutants than in CCR2(+/+) mice sensitized to OVA, but OVA challenge had no additional effect on circulating monocyte chemotactic protein-1 in either genotype. Ag stimulation of lymphocytes isolated from OVA-sensitized CCR2 mutants revealed a significant increase (p < 0.05) in IL-5 production, which differed from OVA-stimulated lymphocytes from sensitized CCR2(+/+) mice. These experiments demonstrate an enhanced response in airway reactivity and in lung inflammation in CCR2(-/-) mutant mice compared with comparably sensitized and challenged CCR2(+/+) mice. These observations suggest that CC chemokines and their receptors are involved in immunomodulation of atopic asthma.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Chemokine CCL2; Disease Models, Animal; Eosinophilia; Immunoglobulin E; Immunoglobulin G; Injections, Intraperitoneal; Lung; Lymphocyte Activation; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; Ovalbumin; Receptors, CCR2; Receptors, Chemokine; Th2 Cells

The pathobiology of allergic asthma is being studied using murine models, most of which use systemic priming followed by pulmonary challenges with the immunizing antigen. In general, mice develop eosinophilic pulmonary inflammation, increased antigen-specific immunoglobulins, and airway hyperreactivity (AHR), all of which are dependent on antigen-specific T cell activation. To establish a model of allergic asthma, which did not require systemic priming, we exposed DO11.10 T cell receptor transgenic mice, which have an expanded repertoire of ovalbumin (OVA), peptide-specific T cells, to limited aerosols of OVA protein. DO11.10 +/- mice developed AHR in the absence of increases in total serum IgE, OVA-specific IgG, or eosinophilia. The AHR was accompanied by pulmonary recruitment of antigen-specific T cells with decreased expression of CD62L and CD45RB and increased expression of CD69, a phenotype indicative of T cell activation. Our results support recent hypotheses that T cells mediate AHR directly.

Topics: Administration, Intranasal; Aerosols; Airway Resistance; Animals; Bronchial Hyperreactivity; Cytokines; Eosinophilia; Female; Immunization; Immunization Schedule; Immunoglobulin E; Immunoglobulin G; Immunophenotyping; Lung; Lymphocyte Activation; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Models, Animal; Ovalbumin; Receptors, Antigen, T-Cell, alpha-beta; Specific Pathogen-Free Organisms

The closely related Th2 cytokines, IL-4 and IL-13, share many biological functions that are considered important in the development of allergic airway inflammation and airway hyperresponsiveness (AHR). The overlap of their functions results from the IL-4R alpha-chain forming an important functional signaling component of both the IL-4 and IL-13 receptors. Mutations in the C terminus region of the IL-4 protein produce IL-4 mutants that bind to the IL-4R alpha-chain with high affinity, but do not induce cellular responses. A murine IL-4 mutant (C118 deletion) protein (IL-4R antagonist) inhibited IL-4- and IL-13-induced STAT6 phosphorylation as well as IL-4- and IL-13-induced IgE production in vitro. Administration of murine IL-4R antagonist during allergen (OVA) challenge inhibited the development of allergic airway eosinophilia and AHR in mice previously sensitized with OVA. The inhibitory effect on airway eosinophilia and AHR was associated with reduced levels of IL-4, IL-5, and IL-13 in the bronchoalveolar lavage fluid as well as reduced serum levels of OVA-IGE: These observations demonstrate the therapeutic potential of IL-4 mutant protein receptor antagonists that inhibit both IL-4 and IL-13 in the treatment of allergic asthma.

Topics: Animals; Antibody Specificity; B-Lymphocyte Subsets; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cytokines; Down-Regulation; Eosinophils; Epithelium; Female; Humans; Immunoglobulin E; Immunophenotyping; Immunosuppressive Agents; Injections, Intraperitoneal; Injections, Subcutaneous; Interleukin-13; Interleukin-4; Leukocyte Count; Lung; Macrophages, Peritoneal; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Phosphorylation; Pulmonary Eosinophilia; Receptors, Interleukin-4; Recombinant Proteins; Spleen; STAT6 Transcription Factor; T-Lymphocyte Subsets; Trans-Activators

To test the effects of Bacillus Calmette-Guérin (BCG) on ovalbumin (OVA)-induced airway hyper-reactivity in guinea pigs, a total of 40 young guinea pigs was individually vaccinated subcutaneously with 0.2 mL of 2% OVA, 50 microg BCG, or a mixture of OVA and BCG (OVA+BCG). Airways were sensitized using nebulization with 1% OVA for 3 min once a week for two applications, followed by 2% OVA nebulized challenge for 3 min 1 week after the last application. Different concentrations of methacholine were used to detect airway hyperreactivities. At the third week, the guinea pigs were nebulized with either methacholine or OVA to test airway hyperreactivity. The OVA-vaccinated group presented with severe airway hyperresponsiveness after OVA and methacholine challenges; the BCG-vaccinated group showed mild airway hyperreactivity; and the OVA+BCG group showed the least amount of airway hyperreactivity. Lung histopathology in all groups, except the OVA+BCG-vaccinated group, showed severe thickening of the alveolar walls which became firmly fibrotic, and narrowing of the alveolar spaces was also noted. The guinea pigs in the OVA+BCG-vaccinated group had similar pulmonary morphology with that of naive guinea pigs, and had mild cell infiltration in the alveolar wall. The results of the skin biopsies at 6 h (2% OVA, 0.05 mL) and 36 h (20 microg PPD, 0.05 mL) after purified protein derivative (PPD) inoculation showed that infiltration of eosinophils and activation of CD4+ T-cells occurred in the OVA-vaccinated group. In the BCG-vaccinated groups, infiltration of CD4+ T-cells, CD8+ T-cells and macrophages occurred. OVA-specific IgG2 increased in the BCG-vaccinated groups after OVA-induced airway hyperreactivity occurred. The peripheral cell subpopulation showed that there was obviously increased activation of CD4+ and CD8+ T-cells in the OVA+BCG-vaccinated group. The phagocytic activity of macrophages also increased in both BCG- and OVA+BCG-vaccinated groups. The prevention of OVA-induced airway hyperreactivities using BCG vaccination in conjugation with OVA in these young guinea pigs indicated that it might be a good approach to avoid allergic reactions in humans.

Topics: Animals; Asthma; BCG Vaccine; Biopsy; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Disease Models, Animal; Flow Cytometry; Guinea Pigs; Immunoglobulin G; Immunohistochemistry; Lung; Lymphocyte Subsets; Male; Methacholine Chloride; Nebulizers and Vaporizers; Ovalbumin; Skin; Time Factors

The participation of mast cells in the induction of antigen-induced airway inflammation and bronchial hyperresponsiveness (BHR) to acetylcholine (ACh) was investigated using pharmacological agents and mast cell-deficient rats (Ws/Ws). A significant increase in the number of leukocytes in bronchoalveolar lavage fluid (BALF) and bronchial responsiveness to ACh were observed 24 h after antigen (ovalbumin) challenge in sensitized Brown-Norway (BN) rats. Disodium cromoglycate and terfenadine did not inhibit antigen-induced airway inflammation and BHR in sensitized BN rats. In contrast, cyclosporin A (CyA), FK-506 and prednisolone significantly inhibited antigen-induced airway inflammation and BHR in sensitized BN rats. In addition, disodium cromoglycate, terfenadine and prednisolone, but not CyA and FK-506, inhibited homologous passive cutaneous anaphylaxis in rats. Furthermore, a significant increase in the number of leukocytes in BALF and BHR was also observed in Ws/Ws rats 24 h after inhalation of antigen; however, the magnitude of BHR in Ws/Ws rats was lower than that in the congenic rats. These findings suggest that mast cells play a partial role in the development of antigen-induced BHR in rats and that the induction of BHR is barely suppressed by mast cell stabilizing agents.

Topics: Acetylcholine; Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cromolyn Sodium; Cyclosporine; Female; Immunosuppressive Agents; Inflammation; Leukocyte Count; Mast Cells; Ovalbumin; Passive Cutaneous Anaphylaxis; Prednisolone; Rats; Rats, Inbred BN; Rats, Wistar; Tacrolimus; Terfenadine

Antigen-specific CD4 T helper type 2 (Th2) cells play a pivotal role in the induction of allergic asthma, but the mechanisms regulating their recruitment into the airways are unknown. Signal transducer and activator of transcription factor (Stat)6 is a transcription factor essential for Th2 cell differentiation. Here we show that Stat6 also controls Th2 cell recruitment and effector function in allergic inflammation in vivo. To isolate the role of Stat6 in regulating Th2 cell trafficking and effector function from its role in Th2 cell differentiation, we used a murine model of asthma in which in vitro-differentiated Stat6(+/+) antigen-specific Th2 cells were adoptively transferred into naive Stat6(-/-) and Stat6(+/+) mice followed by aerosol antigen challenge. We found that all of the features of asthma, including Th2 cell accumulation, Th2 and eosinophil-active chemokine production, and airway eosinophilia, mucus production, and hyperresponsiveness seen in Stat6(+/+) mice, were dramatically absent in Stat6(-/)- mice that received Stat6(+/)+ antigen-specific Th2 cells. Our findings establish Stat6 as essential for Th2 cell trafficking and effector function and suggest that interruption of Stat6 signaling in resident cells of the lung is a novel approach to asthma therapy.

Topics: Animals; Antigens; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines; Cytokines; Eosinophils; Gene Expression Profiling; Goblet Cells; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Mice, Transgenic; Mucus; Ovalbumin; Signal Transduction; STAT6 Transcription Factor; Th2 Cells; Trans-Activators; Transcriptional Activation

We examined the role of IL-18 in preventing the development of and in reversing established allergen-induced airway inflammation and airway hyperreactivity (AHR), the cardinal features of asthma. IL-18, which potently induces IFN-gamma, was administered into the respiratory tract as cDNA in a replication-deficient adenovirus (Adv). Treatment of OVA-sensitized mice with the IL-18-expressing Adv reduced allergen-specific IL-4 production, airway eosinophilia, and mucus production, increased IFN-gamma production, and prevented the development of AHR. The effects of the IL-18 Adv treatment were dependent on the presence of IFN-gamma and IL-12. Moreover, administration of the IL-18 Adv to mice with established AHR greatly reduced AHR and IL-4 production and increased IFN-gamma production. These results demonstrate that IL-18, when administered by Adv into the respiratory tract, effectively reduces AHR and replaces an established Th2-biased immune response with a Th1-biased response.

Topics: Adenoviridae; Administration, Intranasal; Allergens; Animals; Antibodies, Blocking; Antibodies, Monoclonal; Bronchial Hyperreactivity; Cell Line; Cytokines; Genetic Vectors; HeLa Cells; Humans; Injections, Intraperitoneal; Interferon-gamma; Interleukin-12; Interleukin-18; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Th1 Cells; Th2 Cells; Transduction, Genetic

In antigen-challenged guinea pigs, airway hyperreactivity is due to recruitment of eosinophils to the airway nerves and dysfunction of M(2) muscarinic receptors. M(2) receptor dysfunction is caused by eosinophil major basic protein, which is an allosteric antagonist at the receptor. Because glucocorticoids inhibit airway hyperreactivity in humans and in animal models of asthma, we tested whether dexamethasone treatment (6 microg. kg(-)(1). d(-)(1) for 3 d, intraperitoneal) before antigen challenge prevents M(2) receptor dysfunction and airway hyperreactivity. Guinea pigs were sensitized to ovalbumin via intraperitoneal injections, and were challenged with ovalbumin via inhalation. Twenty-four hours later, hyperreactivity and M(2) receptor function were tested. Antigen-challenged animals were hyperreactive to vagal stimulation, and demonstrated loss of M(2) receptor function. Dexamethasone pretreatment prevented hyperreactivity and M(2) receptor dysfunction in antigen-challenged guinea pigs. Antigen challenge resulted in recruitment of eosinophils to the airways and to the airway nerves. Dexamethasone prevented recruitment of eosinophils to the airway nerves but did not affect total eosinophil influx into the airways. These results demonstrate that dexamethasone prevents antigen-induced hyperreactivity by protecting neuronal M(2) muscarinic receptors from antagonism by eosinophil major basic protein, and this protective mechanism appears to be by specifically inhibiting eosinophil recruitment to the airway nerves.

Topics: Analysis of Variance; Animals; Anti-Inflammatory Agents; Antigens; Balantidiasis; Biopsy; Bronchial Hyperreactivity; Dexamethasone; Disease Models, Animal; Drug Evaluation, Preclinical; Female; Glucocorticoids; Guinea Pigs; Immunization; Ovalbumin; Pulmonary Eosinophilia; Receptor, Muscarinic M2; Receptors, Muscarinic

Recent studies in transgenic mice have revealed that expression of a dominant negative form of the transcription factor GATA-3 in T cells can prevent T helper cell type 2 (Th2)-mediated allergic airway inflammation in mice. However, it remains unclear whether GATA-3 plays a role in the effector phase of allergic airway inflammation and whether antagonizing the expression and/or function of GATA-3 can be used for the therapy of allergic airway inflammation and hyperresponsiveness. Here, we analyzed the effects of locally antagonizing GATA-3 function in a murine model of asthma. We could suppress GATA-3 expression in interleukin (IL)-4-producing T cells in vitro and in vivo by an antisense phosphorothioate oligonucleotide overlapping the translation start site of GATA-3, whereas nonsense control oligonucleotides were virtually inactive. In a murine model of asthma associated with allergic pulmonary inflammation and hyperresponsiveness in ovalbumin (OVA)-sensitized mice, local intranasal administration of fluorescein isothiocyanate-labeled GATA-3 antisense oligonucleotides led to DNA uptake in lung cells associated with a reduction of intracellular GATA-3 expression. Such intrapulmonary blockade of GATA-3 expression caused an abrogation of signs of lung inflammation including infiltration of eosinophils and Th2 cytokine production. Furthermore, treatment with antisense but not nonsense oligonucleotides induced a significant reduction of airway hyperresponsiveness in OVA-sensitized mice to levels comparable to saline-treated control mice, as assessed by both enhanced pause (PenH) responses and pulmonary resistance determined by body plethysmography. These data indicate a critical role for GATA-3 in the effector phase of a murine asthma model and suggest that local delivery of GATA-3 antisense oligonucleotides may be a novel approach for the treatment of airway hyperresponsiveness such as in asthma. This approach has the potential advantage of suppressing the expression of various proinflammatory Th2 cytokines simultaneously rather than suppressing the activity of a single cytokine.

Topics: Animals; Asthma; Bronchial Hyperreactivity; DNA-Binding Proteins; Eosinophils; Female; GATA3 Transcription Factor; Interleukin-4; Interleukin-9; Lung; Mice; Mice, Inbred BALB C; Oligonucleotides, Antisense; Ovalbumin; Th2 Cells; Trans-Activators

Airway inflammation and airway hyperresponsiveness (AHR) are fundamental features of asthma. Migration of inflammatory cells from the circulation into the lungs is dependent on adhesion molecule interactions. The cell surface adhesion molecule L-selectin has been demonstrated to mediate leukocyte rolling on inflamed and noninflamed pulmonary endothelium. However, its role in the development of airway inflammation and AHR in asthma has not been examined.. We sought to characterize the role of L-selectin in the recruitment of inflammatory cells to the airway-lung and the development of AHR in a murine model of asthma.. An ovalbumin (OVA)-induced allergic airway disease model of asthma was applied to L-selectin-deficient (LKO) mice and C57BL/6 wild-type (WT) control mice. The development of airway inflammation was assessed by examining leukocyte influx into bronchoalveolar lavage (BAL) fluid and the lung. Total and differential BAL leukocyte counts were determined, and the immunophenotype of BAL lymphocytes was assessed by means of flow cytometry. The development of AHR was assessed by means of whole-body plethysmography.. Airway-lung inflammation was equivalent in LKO and WT mice sensitized-challenged with OVA, as measured by total and differential BAL cell counts and histologic analysis of lung tissue. Numbers of eosinophils, neutrophils, lymphocytes, and monocytes in BAL fluid were equivalent in LKO and WT mice. However, phenotypic analysis of BAL lymphocytes demonstrated significantly reduced CD3(+) populations and increased B220(+) populations in LKO compared with WT mice (P <.05). Remarkably, despite a fulminant inflammatory response in the airway-lung in LKO mice sensitized-challenged with OVA, AHR was completely abrogated.. L-selectin plays a crucial role in the development of AHR but not allergic inflammation in an animal model of asthma. L-selectin represents a potential target for novel asthma therapies specifically aimed at controlling AHR.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Flow Cytometry; Humans; Immunoglobulin E; Inflammation; L-Selectin; Leukocyte Count; Lung; Methacholine Chloride; Mice; Mice, Inbred C57BL; Ovalbumin

This study investigated whether immunostimulatory DNA sequences (ISS) induce a transient or sustained inhibition of Th2 responses to inhaled antigen. We sensitized mice with subcutaneous injections to develop a Th2 response to ovalbumin (ova) and then administered a dose of ISS prior to ova inhalation challenge. Mice were then rechallenged with ova by inhalation a second time at varying time points after the first ova inhalation (1 to 8 weeks later) to determine whether the ISS dose administered prior to the first ova inhalation protected against a subsequent second ova inhalation challenge. A single dose of ISS inhibited the Th2 response to the first inhalation of ova antigen, as well as 4 weeks later to the second inhalation of ova. However, ISS did not inhibit a Th2 response to the second inhalation of ova 8 weeks later. The reversible inhibition of Th2 responses at 8 weeks suggests the need for repeated ISS administration at monthly intervals.

Topics: Adjuvants, Immunologic; Animals; Asthma; Base Sequence; Bronchial Hyperreactivity; CpG Islands; Cytokines; Disease Models, Animal; DNA; Eosinophils; Female; Immunoglobulin E; Mice; Mice, Inbred BALB C; Ovalbumin; Th1 Cells; Th2 Cells

Allergic rhinitis is a risk factor for the development of asthma. About 80% of asthmatic patients also have rhinitis. However, the pattern of induction of allergic rhinitis and asthma remains unclear.. The purpose of this study was to investigate the development of upper airway inflammation in mice during the development of an asthma-like disease and after an acute allergen provocation.. BALB-c mice were sensitized intraperitoneally (i.p) to ovalbumin (OA, days 1-13) and were challenged with aerosols of either OA or saline on 8 consecutive days (days 33-40). In a second experiment, chronic exposure for 8 days was followed by 10 days of rest and then an acute nebulized allergen provocation was performed (day 50). Inflammatory parameters were investigated at different time-points.. Upper and lower eosinophilic airway inflammation were simultaneously induced in the course of repeated inhalations of nebulized OA, as shown by analyses of nasal and broncho-alveolar lavage fluids and histological sections of the nose and bronchi. Mice that developed bronchial hyper-responsiveness also had increased thickness of the nasal mucosa on magnetic resonance image (MRI) scans. When chronic exposure was followed by acute allergen provocation, the latter caused a systemic increase in IL-5 levels, with a concomitant rise in blood and airway eosinophils, primarily in the nose.. Simultaneous induction of eosinophilic inflammation in the nose and lungs was found in a mouse model of respiratory allergy. These findings support the viewpoint that upper and lower airway disease represent a continuum of inflammation involving one common airway and provide evidence for the concept of global airway inflammation after inhalation of allergen.

Topics: Aerosols; Allergens; Animals; Antibody Specificity; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophilia; Eosinophils; Follow-Up Studies; Immunization; Immunoglobulin E; Inhalation Exposure; Interleukin-5; Leukocyte Count; Lymphocyte Count; Magnetic Resonance Imaging; Male; Mice; Mice, Inbred BALB C; Nasal Lavage Fluid; Nasal Mucosa; Ovalbumin; Pilot Projects; Pneumonia; Radiography; Rhinitis

Different strains of mice exhibit different degrees of airway hyperresponsiveness after sensitization to and airway challenge with ovalbumin. Antibody responses in BALB/c mice far exceeded those in C57BL/6 mice; in contrast, although responsiveness to methacholine was much higher in the BALB/c mice, the number of eosinophils in the bronchoalveolar lavage fluid was higher in C57BL/6 animals. Sensitized and challenged BALB/c mice developed increases in lung resistance and decreases in dynamic compliance after methacholine or 5-hydroxytryptamine inhalation. C57BL/6 mice only exhibited significant levels of responsiveness when dynamic compliance was monitored in response to inhaled 5-hydroxytryptamine. Eosinophils accumulated in the peribronchial and peripheral lung tissue in BALB/c mice but were distributed diffusely in the peripheral lung tissue of C57BL/6 mice. Thus, in addition to differences in antibody responses and cholinergic agonist reactivity, differences in the responses of large and small airways may reflect the selective distribution of eosinophils in lung tissue.

Topics: Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Female; Immunization; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Reference Values; Species Specificity

Paradigms of eosinophil effector function in the lungs of asthma patients invariably depend on activities mediated by cationic proteins released from secondary granules during a process collectively referred to as degranulation. In this study, we generated knockout mice deficient for eosinophil peroxidase (EPO) to assess the role(s) of this abundant secondary granule protein in an OVA-challenge model. The loss of EPO had no effect on the development of OVA-induced pathologies in the mouse. The absence of phenotypic consequences in these knockout animals extended beyond pulmonary histopathologies and airway changes, as EPO-deficient animals also displayed OVA-induced airway hyperresponsiveness after provocation with methacholine. In addition, EPO-mediated oxidative damage of proteins (e.g., bromination of tyrosine residues) recovered in bronchoalveolar lavage from OVA-treated wild-type mice was <10% of the levels observed in bronchoalveolar lavage recovered from asthma patients. These data demonstrate that EPO activities are inconsequential to the development of allergic pulmonary pathologies in the mouse and suggest that degranulation of eosinophils recruited to the lung in this model does not occur at levels comparable to those observed in humans with asthma.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Cell Degranulation; Cell Movement; Crosses, Genetic; Cytoplasmic Granules; Disease Models, Animal; Eosinophil Peroxidase; Eosinophils; Injections, Intraperitoneal; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Oxidation-Reduction; Peroxidases; Proteins; Respiratory Hypersensitivity; Sequence Deletion

Asthma, a chronic inflammatory disease characterized by intermittent, reversible airflow obstruction and airway hyperresponsiveness (AHR), is classically characterized by an excess of Th2 cytokines (IL-13, IL-4) and depletion of Th1 cytokines (IFN-gamma, IL-12). Recent studies indicating an important role for Th1 immunity in the development of AHR with allergic inflammation suggest that Th1/Th2 balance may be important in determining the association of AHR with allergic inflammation. We hypothesized that administration of pentoxifylline (PTX), a phosphodiesterase inhibitor known to inhibit Th1 cytokine production, during allergen (OVA) sensitization and challenge would lead to attenuation of AHR in a murine model of allergic pulmonary inflammation. We found that PTX treatment led to attenuation of AHR when administered at the time of allergen sensitization without affecting other hallmarks of pulmonary allergic inflammation. Attenuation of AHR with PTX treatment was found in the presence of elevated bronchoalveolar lavage fluid levels of the Th2 cytokine IL-13 and decreased levels of the Th1 cytokine IFN-gamma. PTX treatment during allergen sensitization leads to a divergence of AHR and pulmonary inflammation following allergen challenge.

Topics: Aerosols; Allergens; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Drug Administration Schedule; Female; Injections, Intraperitoneal; Interphase; Lung; Lymphocyte Activation; Lymphocytes; Mice; Mice, Inbred BALB C; NF-kappa B; Ovalbumin; Pentoxifylline; Respiratory Hypersensitivity; Spleen; Thorax

A relationship between nonspecific bronchial hyperresponsiveness and allergic airway inflammation has been reported in children and in adults with asthma, but the relationship in infants with asthma is still unclear.. To evaluate the relationship between bronchial hyperresponsiveness and total serum IgE level throughout childhood. Bronchial reactivity to methacholine from the age of 1 to 16 years was studied by methacholine inhalation challenge using transcutaneous oxygen pressure (tcPO2) monitoring.. Two hundred one asthmatic children (boys:girls = 132:69; 7.3+/-4.0 years of age, mean +/- SD) were enrolled in this study. The tcPO2 was measured using a tcPO2 monitor. Serial doses of methacholine were doubled until a 10% decrease in tcPO2 from the baseline was reached. The cumulative dose of methacholine at the inflection point of tcPO2 was considered to represent the bronchial reactivity to methacholine.. There was no relationship between the cumulative dose of methacholine at the inflection point of tcPO2 and total serum IgE level in the group of children aged 1 to 4 years (P = 0.212), but significant correlations were found in the groups aged 5 to 10 years and 11 to 16 years (P = 0.044 and P = 0.014, respectively).. We conclude that there is an age-dependent relationship between bronchial reactivity to methacholine and the total serum IgE level and that inhaled allergens, which were more common allergens in older children, may have some effects on the degree of bronchial reactivity to methacholine in children with asthma.

Topics: Administration, Inhalation; Adolescent; Age Factors; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Child; Child, Preschool; Female; Food Hypersensitivity; Humans; Immunoglobulin E; Infant; Male; Methacholine Chloride; Milk; Ovalbumin; Oxygen; Partial Pressure; Radioallergosorbent Test

Ag-specific Th1 and Th2 cells have been demonstrated to play a critical role in the induction of allergic diseases. Here we have investigated the precise mechanisms of Th1-induced airway inflammation. Airway inflammation was induced in BALB/c mice by transfer of freshly induced OVA-specific Th1 or Th2 cells followed by OVA inhalation. In this model, both Th1 and Th2 cells induced airway inflammation. The former induced neutrophilia in airways, whereas the latter induced eosinophilia. Moreover, we found that Th1 cells induced more severe airway hyperresponsiveness (AHR) than Th2 cells. The eosinophilia induced by Th2 cell infusion was almost completely blocked by administration of anti-IL-5 mAb, but not anti-IL-4 mAb. In contrast, Th1-induced AHR and pulmonary neutrophilia were inhibited by the administration of anti-human IL-8R Ab, which blocks the function of mouse CXC chemokine(s). These findings reveal a critical role of mouse CXC chemokine(s) in Th1-dependent pulmonary neutrophilia and AHR.

Topics: Administration, Inhalation; Aerosols; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokines, CXC; Disease Models, Animal; Eosinophils; Inflammation; Leukocytosis; Lung; Mice; Mice, Inbred BALB C; Mice, Transgenic; Neutrophils; Ovalbumin; Peptide Fragments; Respiratory Hypersensitivity; Th1 Cells; Th2 Cells

1. In the present study, the roles of nitric oxide (NO) and superoxide anions (O2(-)) in allergen-induced airway hyperreactivity (AHR) after the late asthmatic reaction (LAR) were investigated ex vivo, by examining the effects of the NO synthase inhibitor N(omega)-nitro-L-arginine methyl ester (L-NAME) and superoxide dismutase (SOD) on the responsiveness to methacholine of isolated perfused guinea-pig tracheae from unchallenged (control) animals and from animals 24 h after ovalbumin challenge. 2. At 24 h after allergen challenge, the animals developed AHR in vivo, as indicated by a mean 2.63 +/- 0.54 fold (P < 0.05) increase in sensitivity to histamine inhalation. 3. Compared to unchallenged controls, tracheal preparations from the ovalbumin-challenged guinea-pigs displayed a significant 1.8 fold (P < 0.01) increase in the maximal response (E(max)) to methacholine, both after intraluminal (IL) and extraluminal (EL) administration of the agonist. No changes were observed in the sensitivity (pEC(50)) to the agonist. Consequently, the DeltapEC(50) (EL-IL), as a measure of epithelial integrity, was unchanged. 4. In the presence of L-NAME (100 microM, IL), tracheae from control guinea-pigs showed a 1.6 fold (P < 0.05) increase in the E(max) of IL methacholine. By contrast, the E(max) of IL methacholine was significantly decreased in the presence of 100 u ml(-1) EL SOD (54% of control, P < 0.01). 5. Remarkably, the increased responsiveness to IL methacholine at 24 h after allergen challenge was reversed by L-NAME to control (P < 0.01), and a similar effect was observed with SOD (P < 0.01). 6. The results indicate that both NO and O2(-) are involved in the tracheal hyperreactivity to methacholine after the LAR, possibly by promoting airway smooth muscle contraction through the formation of peroxynitrite.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Guinea Pigs; Histamine; Methacholine Chloride; NG-Nitroarginine Methyl Ester; Nitric Oxide; Ovalbumin; Perfusion; Respiration; Superoxide Dismutase; Superoxides; Trachea

Mac-1 (CD11b/CD18) is an important adhesion molecule involved in the migration of leukocytes, cell signaling, and subsequent secretory responses. Its precise role in eosinophil recruitment and activation in vivo is not entirely clear. We wished to directly examine the role of Mac-1 in eosinophil migration in a murine model of allergic pulmonary inflammation. Briefly, wild-type (C57Bl/6) and Mac-1-deficient/knockout (Mac-1 KO) mice were intraperitoneally sensitized with ovalbumin (OVA) and alum (AlOH) on Days 0 and 14, and intranasally challenged with OVA either once on Day 14 or five times on Days 14 and 25 through 28. Control animals were challenged with saline. Bronchial hyperresponsiveness was measured, bronchoalveolar lavage (BAL) fluid was collected, and lungs were harvested for histology 24 h after the last challenge. The data demonstrate that wild-type (WT) mice do not respond to one OVA challenge but do develop bronchial hyperreactivity and airway and tissue eosinophilia after five OVA challenges. Conversely, Mac-1 KO mice develop significant airway eosinophilia after one OVA challenge, and the degree of airway inflammation is comparable to that observed in allergic WT mice after five challenges. In Mac-1 KO mice, after five challenges, bronchial hyperreactivity and airway inflammation was significantly enhanced compared with their wild-type counterparts. Administration of an anti-Mac-1 antibody to WT mice, before each of five intranasal OVA challenges, significantly reduces the airway eosinophilia but has no effect on tissue eosinophilia or bronchial hyperresponsiveness. Intravenous injection of interleukin-5 induced a significant blood eosinophilia in both WT and Mac-1 KO mice. Intranasal eotaxin administration induced similar levels of eosinophil migration into the lung tissues and airways of both WT and Mac-1 KO mice. In conclusion, Mac-1-deficient mice develop enhanced eosinophilic inflammation in the lung in response to allergic antigen challenge.

Topics: Airway Resistance; Animals; Antigens; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophilia; Macrophage-1 Antigen; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Respiratory Hypersensitivity

1. The contribution of reactive nitrogen species to the development of airway hyperresponsiveness in a mouse model of allergic inflammation was investigated by the use of selective inhibitors of nitric oxide and superoxide formation. 2. Sensitized mice, repeatedly challenged with ovalbumin showed a significant (P<0.001, n=9) increase in airway responsiveness measured using whole body plethysmography. This hyperresponsiveness was accompanied by an influx of eosinophils into the airway lumen and increased levels of ovalbumin-specific serum IgE. 3. Treatment of mice with the iNOS inhibitor 1400 W or the NADPH-oxidase inhibitor apocynin did not significantly alter cellular influx into the airway lumen nor serum ovalbumin specific IgE. In contrast, apocynin as well as 1400 W inhibited ovalbumin-induced airway hyperresponsiveness (P<0.001 and P<0.05 respectively, n=9). Furthermore, the airways of allergen challenged animals showed clear 3-nitrotyrosine staining, which was mainly located in eosinophils. Remarkably, treatment with apocynin or 1400 W did not alter 3-nitrotyrosine staining. 4. These data suggest that the development of airway hyperresponsiveness during the airway inflammation upon ovalbumin challenge is dependent on the release of both superoxide and nitric oxide and is therefore likely to be dependent on reactive nitrogen species. This mechanism, however, is not reflected by 3-nitrotyrosine formation in the airways.

Topics: Acetophenones; Amidines; Animals; Antioxidants; Benzylamines; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Enzyme Inhibitors; Eosinophils; Hypersensitivity; Immunoglobulin E; Immunohistochemistry; Interferon-gamma; Interleukin-4; Interleukin-5; Lung; Male; Mice; Mice, Inbred BALB C; Neutrophils; Nitric Oxide Synthase; Ovalbumin; Specific Pathogen-Free Organisms; Tyrosine

Current therapeutic approaches to asthma have had limited impact on the clinical management and resolution of this disorder. By using a novel vaccine strategy targeting the inflammatory cytokine IL-5, we have ameliorated hallmark features of asthma in mouse models. Delivery of a DNA vaccine encoding murine IL-5 modified to contain a promiscuous foreign Th epitope bypasses B cell tolerance to IL-5 and induces neutralizing polyclonal anti-IL-5 Abs. Active vaccination against IL-5 reduces airways inflammation and prevents the development of eosinophilia, both hallmark features of asthma in animal models and humans. The reduced numbers of inflammatory T cells and eosinophils in the lung also result in a marked reduction of Th2 cytokine levels. Th-modified IL-5 DNA vaccination reduces the expression of IL-5 and IL-4 by approximately 50% in the airways of allergen-challenged mice. Most importantly, Th-modified IL-5 DNA vaccination restores normal bronchial hyperresponsiveness to beta-methacholine. Active vaccination against IL-5 reduces key pathological events associated with asthma, such as Th2 cytokine production, airways inflammation, and hyperresponsiveness, and thus represents a novel therapeutic approach for the treatment of asthma and other allergic conditions.

Topics: Animals; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Cells, Cultured; Cytokines; Immunoglobulins; Inflammation; Interleukin-5; Mice; Mice, Inbred C3H; Ovalbumin; Pulmonary Eosinophilia; Self Tolerance; Th2 Cells; Vaccines, DNA

Clara cell secretory protein (CCSP) is synthesized by nonciliated bronchiolar cells in the lung and modulates lung inflammation to infection. To determine the role of CCSP in the host response to allergic airway disease, CCSP-deficient [(-/-)] mice were immunized twice with ovalbumin (Ova) and challenged by Ova (2 or 5 mg/m(3)) aerosol. After 2, 3, and 5 days of Ova aerosol challenge (6 h/day), airway reactivity was increased in CCSP(-/-) mice compared with wild-type [CCSP(+/+)] mice. Neutrophils were markedly increased in the bronchoalveolar lavage fluid of CCSP(-/-) Ova mice, coinciding with increased myeloperoxidase activity and macrophage inflammatory protein-2 levels. Lung histopathology and inflammation were increased in CCSP(-/-) compared with wild-type mice after Ova challenge. Mucus production, as assessed by histological staining, was increased in the airway epithelium of CCSP(-/-) Ova mice compared with that in CCSP(+/+) Ova mice. These data suggest a role for CCSP in airway reactivity and the host response to allergic airway inflammation and provide further evidence for the role of the airway epithelium in regulating airway responses in allergic disease.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Chemokine CXCL2; Chemokines; Enzyme Inhibitors; Female; Histocytochemistry; Humans; Inflammation; Lung; Methacholine Chloride; Mice; Mice, Knockout; Neutrophils; Ovalbumin; Peroxidase; Proteins; Uteroglobin

1. We examined the effect of SP100030, a novel inhibitor of activator protein-1 (AP-1) and nuclear factor (NF)-kappa B transcription factors, in a rat model of asthma. 2. Sensitized Brown-Norway rats were treated with SP100030 (20 mg kg(-1) day(-1) for 3 days) intraperitoneally prior to allergen challenge. Allergen exposure of sensitized rats induced bronchial hyperresponsiveness (BHR), accumulation of inflammatory cells in bronchoalveolar lavage (BAL) fluid, and also an increase in eosinophils and CD2(+), CD4(+) and CD8(+) T-cells in the airways together with mRNA expression for IL-2, IL-4, IL-5, IL-10, and IFN-gamma. 3. Pre-treatment with SP100030 inhibited BAL lymphocyte influx (P<0.03), specifically reduced CD8(+) T-cell infiltration in the airway submucosa (P<0.03), and mRNA expression for IL-2, IL-5, and IL-10 (P<0.05). Neutrophil, eosinophil, and CD4(+) T-cells accumulation in the airways and BHR were not affected by SP100030. 4. Our results indicate that suppression of IL-2 and IL-5 mRNA expression may not necessarily lead to suppression of BHR. The expression of IL-5 mRNA may contribute to the airway accumulation of eosinophils, but does not correlate with the extent of eosinophilia. 5. The joint AP-1 and NF-kappa B inhibitor, SP100030, selectively inhibits CD8(+) T-cells, and mRNA expression of both Th1 and Th2 cytokines in vivo, but does not inhibit allergen-induced airway eosinophilia and BHR.

Topics: Allergens; Animals; Blotting, Western; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD2 Antigens; CD4 Antigens; CD8 Antigens; Cytokines; Eosinophils; Gene Expression Regulation; Immunohistochemistry; Immunosuppressive Agents; Interferon-gamma; Interleukin-10; Interleukin-2; Interleukin-4; Interleukin-5; Lung; Lymphocytes; Macrophages; Male; Neutrophils; NF-kappa B; Organic Chemicals; Ovalbumin; Proto-Oncogene Proteins c-jun; Pulmonary Eosinophilia; Rats; Rats, Inbred BN; Respiratory Mucosa; RNA, Messenger; Specific Pathogen-Free Organisms; Transcription Factor AP-1; Transcription Factors

Immunostimulatory DNA sequences (ISS) inhibit eosinophilic inflammation and airway hyperreactivity in mouse models of asthma. In vitro ISS activate natural killer (NK) cells to secrete IFN-gamma, and this cytokine is hypothesized to contribute to the antiallergic effect of ISS in vivo.. We investigated whether ISS activation of NK cells is important in mediating the reduction in airway hyperreactivity and the antieosinophilic effect of ISS in vivo.. We assessed whether ISS modulated the development of eosinophilic airway inflammation and airway hyperreactivity to methacholine in ovalbumin (OVA)-sensitized and OVA allergen-challenged mice pretreated with an antibody to deplete NK cells.. Mice sensitized and challenged with OVA had significant bronchoalveolar lavage and lung eosinophilia, as well as airway hyperresponsiveness. ISS induced significant inhibition of bronchoalveolar lavage and lung eosinophilia, as well as airway hyperresponsiveness, in OVA-sensitized mice pretreated before OVA challenge with an NK cell-depleting antibody (NK(-) mice), as well as in mice pretreated with a control non-NK cell-depleting antibody (NK(+) mice). The NK cell-depleting antibody inhibited ISS-induced IFN-gamma production by spleen cells.. These studies demonstrate that depletion of NK cells has no significant effect on ISS-mediated inhibition of airway eosinophilia and airway hyperresponsiveness in vivo, suggesting that non-NK cells and cytokines other than IFN-gamma derived from NK cells mediate the majority of the ISS-inhibitory effect on eosinophilic inflammation and airway hyperresponsiveness in vivo.

Topics: Adjuvants, Immunologic; Animals; Asthma; Bone Marrow Diseases; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Cells, Cultured; DNA; Eosinophilia; Female; Interferon-gamma; Killer Cells, Natural; Lymphocyte Depletion; Methacholine Chloride; Mice; Mice, Inbred C57BL; Ovalbumin; Pulmonary Eosinophilia

In a guinea pig model of allergic asthma, we investigated the effects of the selective phosphodiesterase inhibitors rolipram (phosphodiesterase 4-selective), Org 9935 (phosphodiesterase 3-selective) and Org 20241 (dual phosphodiesterase 4/phosphodiesterase 3-selective), administered by aerosol inhalation in approximately equipotent bronchodilatory doses, on allergen-induced early and late asthmatic reactions, airway hyperreactivity and airway inflammation. Using ovalbumin-sensitized non-challenged animals, different nebulizer concentrations of each inhibitor were tested for their protective effects against histamine-induced bronchoconstriction. Inhalation of 2.5 mM rolipram, 100 mM 4,5-dihydro-6-(5,6-dimethoxybenzo[b]thien-2-yl-5-methyl-3(2H)pyridazinone (Org 9935) and 10 and 100 mM N-hydroxy-4-(3,4-dimethoxyphenyl)-thiazole-2-carboximidamide HCl (Org 20241) provided a similar, 1.8-fold (P<0.01), 2.0-fold (P<0.05), and 1.8- and 1.9-fold (P<0.05) protection, respectively. The duration of these bronchoprotective effects were different, the rate of decline being faster with rolipram and the lower Org 20241 concentration than with Org 9935 and the higher concentration of Org 20241. All compounds strongly protected against the immediate allergen-induced bronchoconstriction and significantly (P<0.05) diminished the overall early asthmatic reaction from 0 to 6 h following allergen-provocation. The severity of the late asthmatic reaction was also significantly inhibited by rolipram (P<0.05) and Org 9935 (P<0.05). Allergen-induced airway hyperreactivity to inhaled histamine after the early reaction, at 6 h after ovalbumin challenge, was strongly reduced by rolipram (P<0.05) and completely prevented by the two other phosphodiesterase inhibitors; in addition, airway hyperreactivity after the late asthmatic reaction, at 24 h, was abolished in all treatment groups. Bronchoalveolar lavage performed at 24 h after allergen challenge revealed no inhibition of eosinophil infiltration in the rolipram-treated animals, whereas inhalation of Org 9935 and the higher-but not the lower-concentration of Org 20241 strongly reduced the influx of these cells. Eosinophil peroxidase activity in the lavage fluid tended to be diminished in all treatment groups but significance was not reached with the exception of the lower concentration of Org 20241. Infiltration of lymphocytes and macrophages was significantly inhibited by Org 9935 only (P<0.05 and P<0.01, respectively), whereas neutro

Topics: Administration, Inhalation; Allergens; Animals; Anti-Inflammatory Agents, Non-Steroidal; Asthma; Bronchial Hyperreactivity; Bronchoconstriction; Bronchodilator Agents; Disease Models, Animal; Dose-Response Relationship, Drug; Guinea Pigs; Histamine; Inflammation; Ovalbumin; Phosphodiesterase Inhibitors

Severe respiratory syncytial virus (RSV) infection has been hypothesized to be a risk factor for the development of allergy and asthma, but epidemiologic studies in humans have been inconclusive. By use of a well-characterized murine model of RSV infection and allergic sensitization with ovalbumin, the effect of a preceding severe RSV infection on the development of the pulmonary allergic inflammatory response and airway hyperresponsiveness (AHR) was tested. The impact of prior allergic sensitization on RSV-induced illness, as measured by weight loss, also was evaluated. RSV infection before allergic sensitization decreased allergen-induced AHR, production of interleukin-13 in lung tissue, and lung eosinophilia. In contrast, allergic sensitization before RSV infection increased AHR and decreased RSV-related weight loss and lung levels of interferon-gamma but did not alter viral clearance. These data provide evidence that RSV-associated AHR occurs in hosts with allergic responses and that allergic inflammation is diminished when preceded by RSV infection.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchoconstrictor Agents; Female; Hypersensitivity; Interferon-gamma; Interleukin-13; Lung; Lymphocyte Count; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Pulmonary Eosinophilia; Respiratory Syncytial Virus Infections; Respiratory Syncytial Viruses; Time Factors; Virus Replication; Weight Loss

Interleukin (IL)-9 is a T-cell-derived cytokine with pleiotropic activities on T helper 2 cells, B cells, and mast cells. IL-9 may therefore play an important role in the development of allergic pulmonary inflammatory diseases. In this study, an antimouse IL-9 (anti-mIL-9) antibody (Ab) was evaluated against pulmonary eosinophilia, histopathologic changes in lung tissues, serum immunoglobulin (Ig) E levels, and airway hyperresponsiveness (AHR) to methacholine in mice sensitized and challenged with ovalbumin (OVA). Additionally, steady-state levels of IL-4, IL-5, IL-13, and interferon-gamma messenger RNA (mRNA) in the lungs were measured. The anti-mIL-9 Ab (200 microg/mouse, intraperitoneally) was given as either four doses during the sensitization period or as a single dose before OVA challenge. Sensitized mice challenged with OVA displayed marked pulmonary eosinophilia, epithelial damage, and goblet cell hyperplasia. OVA challenge also increased mRNA levels of IL-4, IL-5, and IL-13 in the lungs. AHR was also increased twofold in sensitized, challenged mice. Treatment of sensitized, challenged mice with four doses of anti-mIL-9 Ab significantly reduced pulmonary eosinophilia, serum IgE levels, goblet cell hyperplasia, airway epithelial damage, and AHR, but had no effect on IL-4, IL-5, and IL-13 mRNA levels in the lungs. A single dose of the antibody was ineffective on all measures. These results indicate that an antibody to mIL-9 inhibits the development of allergic pulmonary inflammation and AHR in mice.

Topics: Animals; Antibodies, Monoclonal; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Eosinophils; Gene Expression; Goblet Cells; Hypersensitivity; Immunoglobulin E; Interferon-gamma; Interleukin-13; Interleukin-4; Interleukin-5; Interleukin-9; Male; Mice; Mice, Inbred Strains; Ovalbumin; Pneumonia; Respiratory Mucosa; RNA, Messenger

The ovalbumin (OVA)-sensitized guinea pig is often used as an animal model of asthma and airway hyperreactivity. A characteristic lesion of asthma is excessive production of mucin in the airways. Mechanistic studies of this lesion in guinea pigs have been limited due to lack of mucin gene probes for this species. The aim of the present study was to clone the cDNAs encoding two major airway mucins (Muc2 and Muc5ac) from the guinea pig, and investigate mucin gene expression in lungs of sensitized animals in response to antigen challenge. We isolated and sequenced two cDNA fragments coding for the sequences located within the carboxyl-terminal cysteine-rich region of guinea pig Muc2 and Muc5ac mucins. Comparison of cloned cDNAs with those from other species revealed high degrees of sequence identity and conservation of all cysteine residues in deduced primary sequences. Based on the resultant sequence information, we also designed oligonucleotide primers for specific detection of guinea-pig Muc2 and Muc5ac steady-state mRNA levels via reverse transcriptase/ polymerase chain reaction (RT-PCR). Levels of both Muc2 and Muc5ac mRNA in lungs of OVA-sensitized guinea pigs increased significantly by 30 min after an acute exposure to 0.3% OVA. In addition, levels of eotaxin mRNA also increased in these tissues, but the increases were not significant until 2 h after challenge. Correspondingly, the number of eosinophils in bronchoalveolar lavage fluid did not increase until 4 h postchallenge. Results of these studies suggest that the OVA-sensitized guinea pig responds to allergic challenge with enhanced expression of genes (e.g., eotaxin, Muc2, and Muc5ac) that likely play a role in increased airway inflammation and mucin overproduction, and enhanced mucin gene expression appears to occur before eosinophil infiltration.

Topics: Amino Acid Sequence; Animals; Antineoplastic Agents; Asthma; Base Sequence; Bronchial Hyperreactivity; Cells, Cultured; Chemokine CCL11; Chemokines, CC; Cloning, Molecular; Conserved Sequence; Disease Models, Animal; Eosinophils; Gene Expression; Guinea Pigs; Histamine; Hypersensitivity; Inflammation Mediators; Molecular Sequence Data; Mucin 5AC; Mucin-2; Mucins; Ovalbumin; Respiratory Mucosa; RNA, Messenger; Trachea; Tumor Necrosis Factor-alpha

1. Recently, much attention has been paid to the relationship between the nervous and immune systems. The present study was conducted to clarify the role of neurotrophin low affinity receptor (p75N) in allergic airway inflammation and hyper-responsiveness (AHR) in mice by employing p75N gene deficient mice. 2. Mice were immunized twice by intraperitoneal injections of ovalbumin (OA) at intervals of 12 days. OA was inhaled 10 days after the secondary immunization and repeated three times at 4 days interval. Twenty-four hours after the last inhalation, airway responsiveness to acetylcholine was measured and bronchoalveolar lavage fluid (BALF) was obtained for examining the number of inflammatory cells and the level of cytokines. Serum immunoglobulin was measured as a marker of systemic immune response before the final inhalation. 3. In wild-type mice, repeated antigen provocation resulted in airway eosinophilia, AHR and elevations in serum IgE and interleukin (IL)-4 and -5 in BALF. In p75N gene deficient mice, none of the above parameters was observed after antigen provocation. The antigen-induced production of interferon (IFN)-gamma and nerve growth factor (NGF) were not altered by depletion of p75N gene. 4. The present findings suggest that p75 gene deficiency disrupt an allergic airway inflammation and AHR in mice by interfering type 2 helper T (Th2) cell responses.

Topics: Acetylcholine; Animals; Antigens; Bronchial Hyperreactivity; Bronchitis; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cytokines; Dose-Response Relationship, Drug; Genotype; Immunoglobulin E; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Nerve Growth Factor; Ovalbumin; Receptor, Nerve Growth Factor; Receptors, Nerve Growth Factor; Vasodilator Agents

We investigated whether CTLA4-Ig can reverse established asthma manifestations in a novel murine model of ongoing disease. In BALB/c mice, sensitized to ovalbumin (OVA) without adjuvant, airway inflammation was induced by a first series of OVA aerosol challenges. Murine CTLA4-IgG was then administered, followed by a second series of OVA inhalations. In control-treated mice, two series of OVA challenges induced upregulation of OVA-specific IgE in serum, eosinophils in the bronchoalveolar lavage fluid (BALF), and IL-5 production by lung lymphocytes upon OVA restimulation in vitro, compared with saline-challenged mice. CTLA4-IgG significantly inhibited all of these parameters in OVA-challenged mice. Importantly, mCTLA4-IgG performed better than the gold-standard dexamethasone because this corticosteroid did not inhibit the upregulation of OVA-specific IgE in serum. In a more "severe" ongoing model, induced by sensitization to OVA emulsified in aluminum hydroxide, resulting in airway hyperresponsiveness to methacholine and stronger inflammatory responses, mCTLA4-IgG was less effective in that only the number of eosinophils in the BALF was reduced (P = 0.053), whereas dexamethasone inhibited both BALF eosinophilia and cytokine production by lung lymphocytes. Thus, CTLA4-Ig might be an effective alternative therapy in established allergic asthma, especially in situations of mild disease.

Topics: Abatacept; Aerosols; Allergens; Aluminum Hydroxide; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents; Antigens, CD; Antigens, Differentiation; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cells, Cultured; CTLA-4 Antigen; Cytokines; Dexamethasone; Disease Models, Animal; Emulsions; Eosinophils; Immunization; Immunoconjugates; Immunoglobulin E; Immunosuppressive Agents; Interleukin-5; Leukocyte Count; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Nasal Provocation Tests; Ovalbumin; Specific Pathogen-Free Organisms; T-Lymphocyte Subsets

The accumulation of eosinophils in the lung is a hallmark of asthma. In addition to cytokines such as IL-5 which are essential, chemokines have been implicated in the recruitment of eosinophils to the airway. In particular, eotaxin has been shown to be a selective and potent eosinophil chemoattractant, important in the pathogenesis of allergic disease. The goal of the present study was to define the role of eotaxin-1 in the development of allergen-induced eosinophilic airway inflammation and airway hyperresponsiveness (AHR) to inhaled methacholine (MCh).. Eotaxin-1-deficient mice were sensitized and exposed to a single challenge with allergen. Airway function and airway and tissue as well as peripheral blood and bone marrow eosinophilia were examined 18 and 48 h after the last challenge.. Following allergen sensitization and challenge, eotaxin-1-deficient mice developed levels of AHR to inhaled MCh at 18 and 48 h comparable to controls. Further, levels of bronchoalveolar lavage (BAL) and tissue eosinophilia at the same time points were comparable in the two strains of mice. Tissue eosinophilia, assessed by quantitating major basic protein staining cells, preceded BAL eosinophilia in a similar manner. Bone marrow and peripheral blood eosinophilia were unimpaired in deficient mice.. The results demonstrate that the major eotaxin, eotaxin-1 is not essential for the development of airway eosinophilia or AHR, implying that other chemokines, alone or in combination, can overcome this deficiency.

Topics: Allergens; Animals; Blood Cell Count; Bone Marrow; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokines, CC; Female; Leukocyte Count; Male; Methacholine Chloride; Mice; Ovalbumin; Pulmonary Eosinophilia

In naive rodents, repeated exposure to aerosolized antigen induces suppression of the Th2 response to the antigen. We hypothesized that more prolonged exposure of established asthma model to antigen aerosols may downregulate asthmatic phenotype.. After establishing an ovalbumin (OVA)-induced asthma model, mice were further exposed to OVA (prolonged exposure group) or phosphate-buffered saline (positive controls) 3 days per week for 6 weeks. During week 7, the mice of both groups were finally challenged with OVA.. Prolonged OVA exposure resulted in marked suppression of serum OVA-specific immunoglobulin E (IgE) antibody levels, eosinophilia of the airway, and airway hyperresponsiveness (AHR). However, airway remodeling characterized by goblet cell hyperplasia and airway fibrosis was observed to the same degree in both groups. These effects were accompanied by diminished production of Th2 cytokines such as interleukin-4 (IL-4), IL-5 and IL-13 in bronchoalveolar lavage fluid (BALF) and cultured supernatant of splenocytes. Furthermore, prolonged exposure markedly increased IL-12 levels in BALF.. Prolonged antigen exposure has inhibitory effects on eosinophilic inflammation, AHR and IgE response to antigen, but not on airway remodeling, presumably via inhibition of Th2 cytokines and increased IL-12 production in the lungs.

Topics: Animals; Antibody Specificity; Antigens; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Female; Germ-Free Life; Humans; Immune Tolerance; Immunoglobulin E; Inflammation; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory System

Exposure to diesel exhaust (DE) increased airway inflammatory responses and airway responsiveness to allergen challenge. To clarify the roles of T cells in DE exposure-induced early inflammation, we studied the effect of CD4 and CD8 cells on the effect DE might have on allergic inflammation by using monoclonal antibody-mediated cellular depletion assays. In the bronchoalveolar lavage (BAL) fluid, the numbers of inflammatory cells from 3 mg/m(3) DE-exposed and ovalbumin (OVA)-immunized mice markedly increased. Depletion of CD4(+) cells resulted in reduced accumulation of inflammatory cells. DE exposure to OVA-immunized mice significantly increased interleukin (IL)-1 beta production but decreased IL-12 production. DE exposure significantly enhanced production of the macrophage inflammatory proteins (MIP)-1 alpha and MIP-2, but not monocyte chemoattractant protein (MCP)-1 and regulated upon activation normal T cells expressed and secreted (RANTES). Treatment with anti-CD4 and anti-CD8 mAbs abrogated the adverse effect of DE exposure. In CLN cells from OVA + DE-exposed mice, CD45R/B220-, CD3-, CD4-, and CD8-positive cells were significantly increased, but the OVA-stimulated cytokine production remained at the same levels with OVA-immunized mice. These findings suggest that the induction of early inflammatory responses by DE exposure may initially be related to the modulated function of lymphocyte subpopulations.

Topics: Animals; Antibodies, Monoclonal; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Chemokine CCL2; Chemokine CCL4; Chemokine CCL5; Chemokine CXCL2; Chemokines; Disease Models, Animal; Immunization; Immunophenotyping; Inflammation; Inhalation Exposure; Interleukin-1; Interleukin-12; Lymphocyte Count; Macrophage Inflammatory Proteins; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Time Factors; Vehicle Emissions

Xiao-qing-long-tang (XQLT sho-seiru-to), a traditional Chinese medicine, has been used to treat patients with bronchial asthma in Oriental countries for several centuries. However, the therapeutic mechanisms of this Chinese medicine remain a matter of considerable debate. Therefore, a series of experiments using ovalbumin-sensitized guinea pigs was performed to elucidate the possible antiasthmatic effect of XQLT.. The effect of XQLT on ovalbumin-induced airway inflammation in a guinea pig model of allergic asthma was examined, and early and late asthmatic responses were measured in terms of airway resistance and extent of eosinophil infiltration. Furthermore, the bronchorelaxing effect of XQLT was measured in isolated guinea pig trachea.. XQLT significantly inhibited the antigen-induced immediate asthmatic response (IAR) and late asthmatic response (LAR) in actively sensitized guinea pigs. Cumulative administration of XQLT caused concentration-dependent relaxation of the carbachol-precontracted guinea pig trachea. The bronchorelaxing effect of XQLT was reversed by ICI-118551, a selective beta2-adrenoceptor antagonist. Furthermore, examination of bronchoalveolar lavage fluid (BALF) revealed that XQLT significantly suppressed the increase in eosinophils (24 h after antigen challenge) in the airway. In addition, XQLT significantly attenuated the increase in eosinophils at 1, 6, 24, 48, and 72 h after antigen challenge when it was administered once daily from the day of sensitization to the day of challenge. Histopathologic examination results showed that XQLT suppressed eosinophil infiltration into lung tissue.. These results demonstrate that the antiasthmatic effects of XQLT appear to be partly mediated by stimulation of beta2-adrenoceptors, leading to bronchorelaxation, and that XQLT inhibits the infiltration of eosinophils into the airway. Thus, XQLT may be useful for the prevention or treatment of asthma.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Cell Count; Disease Models, Animal; Drugs, Chinese Herbal; Eosinophils; Guinea Pigs; Immunization; Male; Ovalbumin; Time Factors; Trachea

Respiratory allergies represent a failure to generate nonpathogenic responses to innocuous foreign materials. Herein we assessed the role of the sensitizing dose of allergen in this response/nonresponse paradigm, sensitizing BALB/c mice with 5 ng-2 microg of OVA-alum and assessing their responses to repeated OVA aerosol challenge. Mice sensitized with < or = 25 ng of OVA-alum did not develop atopic antibodies, airway hyperresponsiveness (AHR), eosinophilia, or pulmonary Th2 responses, but the 25-ng group animals did develop significant IgA responses. The mice sensitized with 100 ng of OVA-alum developed AHR in the absence of detectable allergic disease, while the mice sensitized with 250 ng-2 microg of OVA/alum developed full-spectrum allergic disease (i.e., eosinophilia, IgE, IgG1, pulmonary Th2 cytokine responses, and AHR). These data indicate that limiting doses of allergen can differentially induce IgA or AHR in the absence of atopic disease in mice.

Topics: Allergens; Alum Compounds; Animals; Antibody Specificity; Bronchial Hyperreactivity; Cytokines; Disease Models, Animal; Dose-Response Relationship, Immunologic; Immunization; Immunoglobulin A; Immunoglobulin E; Immunoglobulin G; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Plethysmography, Whole Body; Pulmonary Eosinophilia; Respiratory Hypersensitivity; Th2 Cells

Tumor necrosis factor (TNF)-alpha is a potent cytokine with immunomodulatory, proinflammatory, and pathobiologic activities. Although TNF-alpha is thought to play a role in mediating airway inflammation and airway hyperresponsiveness (AHR), its function is not well defined. TNF-alpha-deficient mice and mice expressing TNF-alpha in their lungs because of a TNF-alpha transgene placed under the control of the surfactant protein (SP)-C promoter (SP-C/TNF-alpha-transgenic mice) were sensitized to ovalbumin (OVA) and subsequently challenged with OVA via the airways; airway function in response to inhaled methacholine was monitored. In the TNF-alpha-deficient mice, AHR was significantly increased over that in controls. In contrast, the transgenic mice failed to develop AHR. In addition, sensitized/ challenged TNF-alpha-deficient mice had significantly increased numbers of eosinophils and higher levels of interleukin (IL)-5 and IL-10 in their bronchoalveolar lavage fluid than were found for control mice. However, in SP-C/TNF-alpha-transgenic mice, both the numbers of eosinophils and levels of IL-5 and IL-10 were significantly lower than in sensitized/challenged transgene-negative mice. gammadelta T cells have been shown to be activated by TNF-alpha and to negatively regulate AHR. Depletion of gammadelta T cells in the TNF-alpha-transgenic mice in the present study increased AHR, whereas depletion of these cells had no significant effect in TNF-alpha-deficient mice. These data indicate that TNF-alpha can negatively modulate airway responsiveness, controlling airway function in allergen-induced AHR through the activation of gammadelta T cells.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Cytokines; Immunoglobulin E; Lung; Lymphocyte Activation; Methacholine Chloride; Mice; Mice, Transgenic; Ovalbumin; Proteolipids; Pulmonary Surfactants; Receptors, Antigen, T-Cell, gamma-delta; T-Lymphocytes; Tumor Necrosis Factor-alpha

To assess the effect of intratracheally administered IL-12 recombinant adenovirus (Adm IL-12) on ovalbumin (OVA) induced bronchial hyperresponsiveness in mouse model.. C57BL/6 mice were sensitized by ovalbumin. Adm IL-12 with the dose of 10(8)pfu/mouse was administered intratracheally before OVA challenge. IL-4, IL-5 and IFN-gamma in BALF, bronchial responsiveness, eosinophil count in peripheral blood and BALF, serum total IgE and specific IgE were measured.. There was IL-12 gene expression in the lung tissue after Adm IL-12 administration, concentration of IL-12 in sera and BALF were (540 +/- 60) U/ml and (4 700 +/- 800) U/ml respectively, but were unable to be detected in control group (P < 0.01). Compared with the control group, there were significantly lower levels of IL-4 [(3.5 +/- 2.0) ng/ml vs (85.0 +/- 25.0) ng/ml] (t = 27.97, P < 0.01) and IL-5 [(6.5 +/- 4.5) ng/ml vs (54.0 +/- 14.0) ng/ml], (t = 7.92, P < 0.01) in BALF, accompanying with a higher level of IFN-gamma [(690.0 +/- 32.0) ng/ml vs (12.5 +/- 3.2) ng/ml] (t = 51.6, P < 0.01), lower airway resistance [(360 +/- 30) cm H(2)O vs (810 +/- 50) cm H(2)O] (t = 18.9, P < 0.01) and lower eosinophil counts in both peripheral blood [(0.7 +/- 0.1)% vs (9.2 +/- 0.5)%] (t = 47.1, P < 0.01) and in BALF [(3.5 +/- 0.7) x 10(4)/ml vs (21.6 +/- 4.7) x 10(4)/ml)] (t = 9.33, P < 0.01). However, neither serum total IgE [(65 +/- 9) microgram/ml vs (67 +/- 10) microgram/ml], nor specific IgE [(32 +/- 8) microgram/ml vs (33 +/- 8) microgram/ml] showed significant difference (all P > 0.05).. Intratracheally administered Adm IL-12 inhibits ovalbumin induced airway hyperresponsiveness, which may be an effective approach in the management of antigen induced bronchial asthma.

Topics: Adenoviridae; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Eosinophils; Gene Expression; Genetic Therapy; Genetic Vectors; Humans; Immunoglobulin E; Injections; Interferon-gamma; Interleukin-12; Interleukin-4; Interleukin-5; Leukocyte Count; Mice; Mice, Inbred C57BL; Ovalbumin; Recombination, Genetic; Trachea; Tumor Cells, Cultured

Airway hyperresponsiveness (AHR) might be driven by mechanisms inherent to the airway wall, and/or by factors arising from outside the airways. A porcine model of allergen-induced AHR was utilized to investigate physiological responses in intact airways in vitro and their contribution to responsiveness in vivo. Responsiveness to acetylcholine (ACh) was measured in eight ovalbumin (OA)-sensitized/challenged pigs (tests) and eight saline-challenged controls. In vivo responsiveness to ACh was determined from pulmonary resistance (RL). In vitro responsiveness to ACh was determined from airway pressure in isovolumic bronchial segments, after exposure via the adventitial or the luminal surface. Test pigs had lung (255+/-26% increase in RL, p<0.0001) and skin responses to OA, and AHR to ACh (p<0.0001). In vitro, test bronchi were less sensitive than controls to ACh applied to the airway adventitia (negative log of the ACh concentration producing half the maximum response (pD2)=4.18 and 4.58 respectively, p<0.01), but not the lumen. Test bronchi had an increased amount of smooth muscle normalized for airway size versus controls (p<0.05). Maximum responses to lumenal ACh in vitro showed a weak positive correlation with maximum changes to ACh in vivo (r=0.599, p=0.05). This study concludes that the effect of antigen challenge on bronchial responsiveness varies with the route of exposure to acetylcholine. In vitro responses to lumenal acetylcholine are increased despite a possible reduction in responsiveness of airway smooth muscle. Responsiveness of the bronchial wall only partially explains responsiveness of the lungs in vivo.

Topics: Acetylcholine; Aerosols; Animals; Bronchi; Bronchial Hyperreactivity; Female; Immunization; In Vitro Techniques; Lung; Lung Compliance; Muscle, Smooth; Ovalbumin; Skin; Swine

Asthma is a respiratory disorder characterized by airway hyperreactivity (AHR) and inflammation and is associated with high serum IgE and overproduction of IL-4, IL-5, and IL-13 by allergen-specific Th2 cells. Our previous studies demonstrated that heat-killed Listeria monocytogenes (HKL) as an adjuvant in immunotherapy successfully reversed ongoing Ag-specific Th2-dominated responses toward Th1-dominated responses, but it was unclear if such immune modulation could reverse ongoing, established disease in target organs such as the lung. In this paper we show that a single dose of Ag plus HKL as adjuvant significantly reduced AHR in a murine model for asthma and reversed established AHR when given late after allergen sensitization. HKL as adjuvant also dramatically inhibited airway inflammation, eosinophilia, and mucus production, significantly reduced Ag-specific IgE and IL-4 production, and dramatically increased Ag-specific IFN-gamma synthesis. The inhibitory effect of HKL on AHR depended on the presence of IL-12 and CD8+ T cells and was associated with an increase of IL-18 mRNA expression. Thus, our results demonstrate that HKL as an adjuvant for immunotherapy mediates immune deviation from a pathological Th2-dominated response toward a protective immune response in peripheral lymphoid tissues and in the lungs and may be clinically effective in the treatment of patients with established asthma and allergic disease.

Topics: Adjuvants, Immunologic; Administration, Intranasal; Allergens; Animals; Antigens, Bacterial; Bacterial Vaccines; Bronchial Hyperreactivity; Cells, Cultured; Cytokines; Hot Temperature; Immunosuppressive Agents; Inflammation; Injections, Intradermal; Interleukin-18; Listeriosis; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; RNA, Messenger; Spleen; T-Lymphocytes; Th1 Cells; Vaccines, Inactivated

This study was carried out to determine if the bronchoconstrictive effect of cigarette smoke (CS) is enhanced when airway hyperresponsiveness is induced by ovalbumin (Ova) sensitization, and if so, whether an increase in endogenously released tachykinins is involved. The bronchoconstrictive effects of an acute CS inhalation challenge (15 ml; 50% concentration) were compared between guinea pigs sensitized with aerosolized Ova and matching control animals (receiving saline aerosol). In Ova-sensitized animals, there were marked increases in the numbers of eosinophils and neutrophils in the bronchoalveolar lavage fluid (BALF), which was accompanied by an elevated bronchomotor response to acetylcholine (ACh). The baseline lung resistance (RL) and dynamic pulmonary compliance (Cdyn) were not significantly different between the two groups; however, the same CS inhalation challenge evoked a significantly more intense bronchoconstriction in the Ova-sensitized group (control group: DeltaRL = 68 +/- 8%, DeltaCdyn = -26 +/- 6%; Ova group: DeltaRL = 425 +/- 76%; DeltaCdyn = -47 +/- 8%). The levels of substance P-like immunoreactivity (SP-LI) and calcitonin gene-related peptide-like immunoreactivity (CGRP-LI) measured in the bronchoalveolar lavage (BAL) collected after CS inhalation challenge were also significantly greater in Ova-sensitized animals than in control animals. Furthermore, pretreatment with SR-48968, a selective antagonist of neurokinin-2 (NK(2)) receptor, inhibited more than 85% of the enhanced bronchomotor responses to CS challenge, but did not significantly reduce the airway hyperresponsiveness to ACh in Ova-sensitized guinea pigs. These results show that Ova sensitization induces airway hyperresponsiveness to inhaled CS, and that the endogenous tachykinins evoked by CS-induced activation of lung C fibers play a primary role in this augmented response.

Topics: Acetylcholine; Airway Resistance; Animals; Benzamides; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstriction; Calcitonin Gene-Related Peptide; Disease Models, Animal; Eosinophils; Guinea Pigs; Leukocyte Count; Lung; Lung Compliance; Male; Neutrophils; Nicotiana; Ovalbumin; Piperidines; Plants, Toxic; Receptors, Neurokinin-2; Smoke

T helper 2 (Th2) cells play a critical role in the pathogenesis of asthma, but the precise immunological mechanisms that inhibit Th2 cell function in vivo are not well understood. Using gene therapy, we demonstrated that ovalbumin-specific (OVA-specific) Th cells engineered to express latent TGF-beta abolished airway hyperreactivity and airway inflammation induced by OVA-specific Th2 effector cells in SCID and BALB/c mice. These effects correlated with increased concentrations of active TGF-beta in the bronchoalveolar lavage (BAL) fluid, demonstrating that latent TGF-beta was activated in the inflammatory environment. In contrast, OVA-specific Th1 cells failed to inhibit airway hyperreactivity and inflammation in this system. The inhibitory effect of TGF-beta-secreting Th cells was antigen-specific and was reversed by neutralization of TGF-beta. Our results demonstrate that T cells secreting TGF-beta in the respiratory mucosa can indeed regulate Th2-induced airway hyperreactivity and inflammation and suggest that TGF-beta-producing T cells play an important regulatory role in asthma.

Topics: Allergens; Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Line; Eosinophilia; Genetic Therapy; Interferon-gamma; Mice; Mice, Inbred BALB C; Mice, SCID; Ovalbumin; Pneumonia; Transforming Growth Factor beta

In vitro and in vivo studies, in both animal models and human asthmatics, have implicated IL-4 as an important inflammatory mediator in asthma. In a murine asthma model, we examined the anti-inflammatory activities of soluble IL-4R (sIL-4R). In this model, mice sensitized to OVA by i.p. and intranasal (i.n.) routes are challenged with the allergen by i.n. administration. The OVA challenge elicits an eosinophil infiltration into the lungs, with widespread mucus occlusion of the airways, and results in bronchial hyperreactivity. sIL-4R (0.1-100 microgram) was administered by either i.n. or i.p. routes before OVA challenge in OVA-sensitized mice. Both blood and bronchoalveolar lavage fluid levels of sIL-4R were significantly elevated compared with controls by i.n. delivery of 100 microgram sIL-4R; i.p. delivery of 100 microgram sIL-4R only raised blood levels of sIL-4R. The i.n. administration of 100 microgram sIL-4R before allergen challenge significantly reduced late phase pulmonary inflammation, blocking airway eosinophil infiltration, VCAM-1 expression, and mucus hypersecretion. In contrast, i.p. delivery of 100 microgram sIL-4R inhibited only the influx of eosinophils into the lungs, but not airway mucus release. Furthermore, sIL-4R treatment by either i.n. or i.p. routes did not reduce airway hyperreactivity in response to methacholine challenge. Thus, elevating airway levels of sIL-4R through the administration of exogenous sIL-4R is effective in blocking the late phase pulmonary inflammation that occurs in this murine allergen-challenge asthma model. These results suggest that sIL-4R may have beneficial anti-inflammatory effects in asthmatic patients.

Topics: Administration, Intranasal; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Movement; Disease Models, Animal; Eosinophils; Female; Humans; Immunoglobulin E; Injections, Intraperitoneal; Leukocytes, Mononuclear; Lung; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Receptors, Interleukin-4; Solubility

The mechanisms regulating the selective migration and degranulation of eosinophils in the asthmatic lung and the subsequent development of airways hyperreactivity (AHR) have not been fully delineated. In this investigation, we have employed a novel transgene model to facilitate the dissection of the contributions of IL-5 and/or eotaxin to eosinophil function in the absence of complex tissue signals derived from the allergic lung. Gene transfer of IL-5 and/or eotaxin to the lungs of naive mice induced a pronounced and selective airways eosinophilia, but did not result in eosinophil degranulation or AHR. Airways eosinophilia occurred independently of the induction of a blood eosinophilia, but was markedly augmented by the coexpression of both cytokines and/or by the transient mobilization of eosinophils from the bone marrow by the administration of i.v. IL-5. However, for eosinophil degranulation and AHR to occur, the inhalation of Ag was required in association with IL-5 and eotaxin expression. Investigations in IL-5-deficient mice linked eosinophilia, and not solely IL-5 and eotaxin, with the induction of AHR. Furthermore, eosinophil degranulation and AHR were dependent on CD4+ T cells. Importantly, this investigation shows that IL-5 regulates eosinophilia within the lung as well as in the circulation and also amplifies eotaxin-induced chemotaxis in the airway compartment. Moreover, the interplay between these cytokines, CD4+ T cells, and factors generated by Ag inhalation provides fundamental signals for eosinophil degranulation and the induction of AHR.

Topics: Administration, Intranasal; Animals; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Cell Degranulation; Chemokine CCL11; Chemokines, CC; Chemotactic Factors, Eosinophil; Chemotaxis, Leukocyte; Choline; Cytokines; Eosinophilia; Eosinophils; Genetic Vectors; Injections, Intravenous; Interleukin-5; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Transgenic; Ovalbumin; Vaccinia virus

Tachykinins are involved in the development of bronchial inflammation and airway hyperresponsiveness (AHR); however, the role of the neurokinin-1 (NK(1)) receptor in acid-aerosol-induced bronchial impairment in asthmatic patients remains controversial.. To investigate the effects on the NK(1) receptor antagonist FK888 the neurokinin-2 (NK(2)) receptor antagonist SR48968 on sulfuric-acid (H(2)SO(4))-induced AHR in guinea pigs, specific airways resistance (sRaw) and airways responsiveness to methacholine (MCh) were measured before and after 6 h of exposure to H(2)SO(4) aerosol (pH 1.7, 82 mg/m(3)) in ovalbumin-sensitized guinea pigs.. Airway responsiveness to MCh significantly increased (p<0. 05) after the exposure, however sRaw did not. Treatment with FK888 significantly inhibited (p<0.05) H(2)SO(4)-induced AHR in a dose-dependent manner, as did SR48968.. These results suggest that not only NK(2) but also NK(1) receptors might have important roles in the development of acid-aerosol-induced AHR.

Topics: Aerosols; Airway Resistance; Animals; Benzamides; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoconstriction; Dipeptides; Guinea Pigs; Indoles; Male; Methacholine Chloride; Nebulizers and Vaporizers; Neurokinin-1 Receptor Antagonists; Ovalbumin; Piperidines; Receptors, Neurokinin-2; Sulfuric Acids

Mouse models have been established mirroring key features of human bronchial asthma including airway hyperresponsiveness (AHR). Acute airway obstruction in response to an allergen challenge, however, remains to be demonstrated in these models.. A mouse model of allergic lung inflammation was employed to analyze the development of specific (allergen-induced) and nonspecific (methacholine-induced) airway obstruction.. Mice were sensitized to ovalbumin (OVA) and challenged with OVA aerosol twice each week during four weeks. Changes in lung functions were determined by noninvasive head-out body plethysmography. The development of acute airway obstruction after OVA challenge and AHR after methacholine aerosol application were assessed by a decrease in the mid-expiratory flow rate (EF(50)).. Two airway challenges were sufficient to induce AHR (5.7 vs. 15 mg/ml methacholine). Further OVA challenges reduced the baseline EF(50) from 1.85 to 1.20 ml/s (4th week) and induced acute airway obstruction. The OVA-induced obstruction was maximal in the 4th week (EF(50) = 0.91 ml/s).. The development of acute airway obstruction in allergen-sensitized mice was demonstrated by means of head-out body plethysmography. In our model, AHR was observed before the development of airway obstruction.

Topics: Airway Obstruction; Airway Resistance; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Immunoglobulin E; Immunoglobulin G; Maximal Expiratory Flow Rate; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Plethysmography, Whole Body; Tidal Volume

Suplatast tosilate (IPD) is a newly developed 'anti-allergic' drug. It seems to be a unique compound because of its ability to suppress IgE but not IgG or IgM production in vivo and cytokine production from type 2 helper T cells (Th2) in vitro. However, information on its in vivo effect on an animal model of asthma is limited.. BALB/c mice sensitized to ovalbumin (3 times, 2-week interval) were challenged with ovalbumin by inhalation (50 mg/ml for 20 min, once a day for 6 days). In this study, we explored the influence of IPD on eosinophil infiltration into the airways, bronchial hyperresponsiveness (BHR) to methacholine, specific IgE antibody production, and cytokines in bronchoalveolar lavage fluid (BALF) using this murine model.. Treatment with IPD significantly reduced the number of total cells and eosinophils in BALF (around -40%) and almost completely inhibited the development of antigen-induced BHR. Histological findings confirmed the reduction of submucosal cell infiltration in the lung, and disclosed the marked inhibition of bronchial epithelial cell damage. Ovalbumin-specific IgE was slightly but significantly reduced. The levels of IL-4, IL-5 and IL-13 in BALF were significantly decreased in mice treated with the compound compared to those in untreated mice.. These results suggest that IPD is capable of inhibiting the production of Th2 cytokines, which inhibit eosinophil infiltration into the murine airway, IgE synthesis, and development of BHR, in a murine model of asthma.

Topics: Animals; Anti-Allergic Agents; Arylsulfonates; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Eosinophilia; Female; Immunoglobulin E; Immunoglobulin G; Lung; Methacholine Chloride; Mice; Ovalbumin; Sulfonium Compounds

Allergic asthma is caused by the aberrant expansion in the lung of T helper cells that produce type 2 (TH2) cytokines and is characterized by infiltration of eosinophils and bronchial hyperreactivity. This disease is often triggered by mast cells activated by immunoglobulin E (IgE)-mediated allergic challenge. Activated mast cells release various chemical mediators, including prostaglandin D2 (PGD2), whose role in allergic asthma has now been investigated by the generation of mice deficient in the PGD receptor (DP). Sensitization and aerosol challenge of the homozygous mutant (DP-/-) mice with ovalbumin (OVA) induced increases in the serum concentration of IgE similar to those in wild-type mice subjected to this model of asthma. However, the concentrations of TH2 cytokines and the extent of lymphocyte accumulation in the lung of OVA-challenged DP-/- mice were greatly reduced compared with those in wild-type animals. Moreover, DP-/- mice showed only marginal infiltration of eosinophils and failed to develop airway hyperreactivity. Thus, PGD2 functions as a mast cell-derived mediator to trigger asthmatic responses.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Crosses, Genetic; Female; Gene Targeting; Humans; Immunoglobulin E; Interferon-gamma; Interleukins; Lung; Lymphocytes; Male; Mast Cells; Mice; Mice, Inbred C57BL; Mucus; Ovalbumin; Prostaglandin D2; Receptors, Immunologic; Receptors, Prostaglandin; Respiratory Mucosa

Immunization of BALB/c mice with alum-adsorbed OVA, followed by three bronchoprovocations with aerosolized OVA, resulted in the development of airway hyperresponsiveness (AHR) and allergic inflammation in the lung accompanied by severe infiltration of eosinophils into airways. In this murine asthma model, administration of monoclonal anti-IL-5 Ab before each Ag challenge markedly inhibited airway eosinophilia, but the treatment did not affect the development of AHR. Immunization and aerosol challenges with OVA following the same protocol failed to induce AHR in the mast cell-deficient W/Wv mice, but induced AHR in their congenic littermates, i.e., WBB6F1 (+/+) mice. No significant difference was found between the W/Wv mice and +/+ mice with respect to the IgE and IgG1 anti-OVA Ab responses and to the airway eosinophilia after Ag provocations. It was also found that reconstitution of W/Wv mice with bone marrow-derived mast cells cultured from normal littermates restored the capacity of developing Ag-induced AHR, indicating that lack of mast cells was responsible for the failure of W/Wv mice to develop Ag-induced AHR under the experimental conditions. However, the OVA-immunized W/Wv mice developed AHR by increasing the frequency and Ag dose of bronchoprovocations. The results suggested that AHR could be developed by two distinct cellular mechanisms. One would go through mast cell activation and the other is IgE/mast cell independent but an eosinophil/IL-5-dependent mechanism.

Topics: Aerosols; Animals; Asthma; Bone Marrow Transplantation; Bronchial Hyperreactivity; Cells, Cultured; Disease Models, Animal; Male; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Mutant Strains; Nebulizers and Vaporizers; Ovalbumin

Recently, we reported that freely moving Flinders sensitive line rats (FSL, selectively bred for their cholinergic hyperresponsiveness) are more susceptible to allergen-induced airway hyperresponsiveness than their control counterparts-Flinders resistant line (FRL) rats. In this study the two Flinders lines were compared for responsiveness of excised tracheal and primary bronchial smooth muscle in vitro. FSL tissues were slightly but significantly more sensitive to cholinergic stimulation than FRL tissues (slightly lower EC(50) value for carbachol) but the FRL tissues were more responsive, exhibiting larger amplitude of response. Surprisingly, previous exposure to allergen challenge was accompanied by reduced in vitro responses to spasmogens in both rat lines. We conclude that FSL and FRL airways do not differ greatly with respect to sensitivity to cholinergic stimulation in vitro and that inflamed airways show reduced in vitro responses to spasmogens. The discrepancy between the in vivo and in vitro findings suggests that responsiveness of airway smooth muscle involves regulation from both proximal and distal sites.

Topics: Airway Resistance; Allergens; Animals; Bronchi; Bronchial Hyperreactivity; Bronchoconstriction; Bronchoconstrictor Agents; Carbachol; Cholinergic Agents; Electric Stimulation; Female; Muscle, Smooth; Ovalbumin; Parasympathetic Nervous System; Potassium Chloride; Rats; Rats, Inbred Strains; Respiratory Function Tests; Trachea

CD4(+) T cells are thought to play a major role in the initiation and perpetuation of T helper cell, type 2 (Th2)-like allergic airway inflammation. However, it is not clear whether activation of resident antigen-specific CD4(+) T cells is in itself sufficient to induce such a phenotype. Using ovalbumin (OVA)-specific alphabeta-T cell receptor transgenic Balb/c DO11.10 mice, we were able to test this hypothesis. Nonsensitized DO11.10 mice but not wild-type mice responded to a primary OVA aerosol with a rapid and impressive bronchoalveolar lavage (BAL) neutrophilia followed by a smaller but significant eosinophilia. Responses in DO11.10 mice were mediated by OVA-specific activation of CD4(+) T cells because in vivo depletion of CD4(+) but not CD8(+) T cells abrogated inflammatory cell influx. Cytokines measured in BAL fluid (BALF) after OVA aerosol exposure of DO11.10 mice were indicative of a T helper cell, type 1 (Th1)-like immune response. Further, neutralization of interferon gamma (IFN-gamma) with antibody enhanced eosinophil influx, suggesting that IFN-gamma production was limiting the development of a Th2 response. Despite this, an increased prevalence of cells staining for mucus was seen in the bronchial epithelium, a feature more commonly associated with a Th2-immune response. Unlike what was seen in OVA-sensitized wild-type mice, multiple OVA aerosol exposures of DO11.10 mice failed to induce airway hyperresponsiveness (AHR) to inhaled methacholine. In conclusion, in vivo stimulation of resident lung CD4(+) T cells with antigen caused lung inflammation with characteristics of both a Th1- and Th2-immune response but was insufficient to directly induce AHR.

Topics: Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Epitopes; Female; Interferon-gamma; Lung; Male; Mice; Mice, Inbred BALB C; Mice, Transgenic; Ovalbumin; Receptors, Antigen, T-Cell, alpha-beta; Respiratory Hypersensitivity; Th2 Cells

Human asthma is characterized by increased airway hyperreactivity to a variety of bronchoconstricting agents. Aberrant type 2 immune responses in the lung have been associated with airway hyperreactivity in both human asthma and in murine models of allergic airways disease. Despite their intrinsically elevated basal airway reactivity to smooth muscle constricting agents, A/J mice demonstrated no inherent inflammatory cell infiltration nor elevation of type 2 cytokines in the lung. Crossed bone marrow reconstitution experiments between A/J and MHC congenic B10.A mice revealed enhanced airway reactivity only in A/J recipients, irrespective of whether they had been reconstituted with A/J or B10. A hemopoietic cells. Further, A/J-derived bone marrow cells did not affect the reactivity of B10.A recipients. Although mice on RAG-deficient and IL-4-deficient backgrounds demonstrate substantial abrogation of allergen-induced airway hyperreactivity, these gene deletions had no impact on the elevated baseline reactivity when backcrossed onto A/J mice. Thus, in these mice, basal airway hyperreactivity is maintained independently of type 2 immunity induced by allergens.

Topics: Aerosols; Animals; B-Lymphocytes; Bone Marrow Cells; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Immunity, Cellular; Immunity, Innate; Interleukin-4; Lymphocyte Depletion; Mice; Mice, Congenic; Mice, Inbred A; Mice, Inbred C57BL; Nebulizers and Vaporizers; Ovalbumin; T-Lymphocytes

Although phosphodiesterase (PDE) 3 and 4 inhibitors have received much attention for the treatment of bronchial asthma, systemic adverse effects have also been reported.. The purpose of this study was to investigate the effect of inhaled olprinone, a newly developed PDE3 inhibitor, and KF19514, a PDE1 and 4 inhibitor, on antigen-induced airway reactions in guinea-pigs.. Fifteen minutes after inhalation of olprinone (0.1 or 1.0 mg/mL) and KF19514 (0.1 or 0.01 mg/mL), animals were given an antigen challenge. Bronchial hyper-responsiveness and bronchoalveolar lavage fluid cell analysis were performed 24 h after the antigen challenge.. Inhalation of olprinone and KF19514 caused a dose-related inhibition of antigen-induced bronchoconstriction. Antigen inhalation significantly increased bronchoconstrictor responses to methacholine, and airway accumulation of neutrophils and eosinophils, 24 h after the antigen challenge. These responses were dose-dependently prevented by KF19514, but not by olprinone.. The results indicate that inhaled PDE inhibitors might be useful for treatment of bronchial asthma.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Administration, Inhalation; Aerosols; Animals; Antigens; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchodilator Agents; Cyclic Nucleotide Phosphodiesterases, Type 1; Cyclic Nucleotide Phosphodiesterases, Type 3; Cyclic Nucleotide Phosphodiesterases, Type 4; Guinea Pigs; Imidazoles; Male; Methacholine Chloride; Naphthyridines; Ovalbumin; Phosphodiesterase Inhibitors; Pyridones

Chronic airway eosinophilia is associated with allergic asthma and is mediated in part by secretion of IL-5 from allergen-specific Th2 lymphocytes. IL-5 is a known maturation and antiapoptotic factor for eosinophils and stimulates release of nascent eosinophils from bone marrow into the peripheral circulation. An antisense oligonucleotide found to specifically inhibit IL-5 expression in vitro was observed to significantly reduce experimentally induced eosinophilia in vivo, in both the murine OVA lung challenge and allergic peritonitis models. Intravenous administration resulted in sequence-dependent inhibition of eosinophilia coincident with reduction of IL-5 protein levels, supporting an antisense mechanism of action. Potent suppression of lung eosinophilia was observed up to 17 days after cessation of oligonucleotide dosing, indicating achievement of prolonged protection with this strategy. Furthermore, sequence-specific, antisense oligonucleotide-mediated inhibition of Ag-mediated late phase airway hyperresponsiveness was also observed. These data underscore the potential utility of an antisense approach targeting IL-5 for the treatment of asthma and eosinophilic diseases.

Topics: Adjuvants, Immunologic; Animals; Antigens; Asthma; Bronchial Hyperreactivity; Disease Models, Animal; Eosinophilia; Gene Expression Regulation; Injections, Intraperitoneal; Injections, Intravenous; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Oligonucleotides, Antisense; Ovalbumin; RNA, Messenger; Time Factors; Tumor Cells, Cultured

Cytokines play an important role in modulating inflammatory responses and, as a result, airway tone. IL-10 is a regulatory cytokine that has been suggested for treatment of asthma because of its immunosuppressive and anti-inflammatory properties. In contrast to these suggestions, we demonstrate in a model of allergic sensitization that mice deficient in IL-10 (IL-10-/-) develop a pulmonary inflammatory response but fail to exhibit airway hyperresponsiveness in both in vitro and in vivo assessments of lung function. Reconstitution of these deficient mice with the IL-10 gene fully restores development of airway hyperresponsiveness comparable to control mice. These results identify an important role of IL-10, downstream of the inflammatory cascade, in regulating the tone of the airways after allergic sensitization and challenge.

Topics: Aerosols; Animals; Blood Proteins; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Electric Stimulation; Eosinophil Granule Proteins; Eosinophil Peroxidase; Eosinophilia; Female; Genetic Complementation Test; Genetic Therapy; Immunization; Inflammation; Interleukin-10; Leukotrienes; Lung; Male; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; Muscle, Smooth; Ovalbumin; Peroxidases; Respiratory Hypersensitivity; Ribonucleases; Specific Pathogen-Free Organisms; Trachea

Muscarinic cholinergic signaling plays an essential role in the control of the normal airway functions and in the development of pulmonary pathologies including asthma. In this paper we demonstrate that the airways of mice deficient in a cAMP-specific phosphodiesterase (PDE4D) are no longer responsive to cholinergic stimulation. Airway hyperreactivity that follows exposure to antigen was also abolished in PDE4D(-/-) mice, despite an apparently normal lung inflammatory infiltration. The loss of cholinergic responsiveness was specific to the airway, not observed in the heart, and was associated with a loss of signaling through muscarinic receptors with an inability to decrease cAMP accumulation. These findings demonstrate that the PDE4D gene plays an essential role in cAMP homeostasis and cholinergic stimulation of the airway, and in the development of hyperreactivity. In view of the therapeutic potentials of PDE4 inhibitors, our findings provide the rationale for novel strategies that target a single PDE isoenzyme.

Topics: 3',5'-Cyclic-AMP Phosphodiesterases; Animals; Asthma; Bronchial Hyperreactivity; Cyclic AMP; Cyclic Nucleotide Phosphodiesterases, Type 4; Isoenzymes; Lung; Mice; Mice, Inbred C57BL; Ovalbumin; Receptors, Muscarinic

The airway inflammation observed in asthma is orchestrated by activated Th-2 lymphocytes relevant for the induction of altered airway responsiveness. An increasing body of evidence is accumulating that not only the pro-inflammatory cytokines interleukin (IL)-4 and IL-5 but also the immunomodulating cytokines IL-12 and possibly IL-10 are crucial for regulating the allergic airway inflammation.. Since IL-10 is capable of downregulating a broad spectrum of pro-inflammatory cytokines, we wanted to address the role of endogenously produced IL-10 in vivo in allergic asthma.. Knockout (IL-10(-/-)) mice (C57BL/6-IL10tm1Cgn) and wild-type (WT) counterparts were immunized (day 0) and exposed (day 14-21) to ovalbumin (OVA). Airway inflammation and reactivity (AR), serum allergen-specific IgE responses and cytokine profiles in the bronchoalveolar lavage fluid (BALF) were studied.. The IL-10(-/-) mice had more eosinophilic airway inflammation but comparable levels of allergen-specific serum IgE compared to the WT mice after allergen challenge. The AR was comparably increased in the OVA challenged WT and IL-10(-/-) mice vs sham-exposed WT, but not vs sham-exposed IL-10(-/-)mice since these showed a higher baseline AR. IFN gamma, IL-4 and IL-13 were comparable and IL-5 was even lower in the BALF of the in IL-10(-/-) mice compared to the similarly exposed WT mice.. These results indicate that IL-10 plays an important and possibly direct role in the control of airway inflammation and responsiveness in an in vivo mouse model of allergy.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Carbachol; Cytokines; Disease Models, Animal; Germ-Free Life; Immunoglobulin E; Inflammation; Interleukin-10; Lung; Male; Mice; Mice, Inbred C57BL; Ovalbumin

The role of IL-5 receptor alpha chain (IL-5Ralpha) in the onset of bronchial hyperresponsiveness (BHR) to acetylcholine was investigated by testing IL-5Ralpha knockout (IL-5Ralpha KO) mice.. Mice were immunized with antigen at intervals of 12 days. Starting 10 days after the secondary immunization, mice were exposed to antigen three times every fourth day. Twenty-four hours after the last antigen challenge, bronchial responsiveness to acetylcholine was measured and bronchoalveolar lavage was carried out.. Twenty-four hours after the last antigen inhalation, total and differential cells counts of bronchoalveolar lavage revealed a significant increase in eosinophils and lymphocytes in ovalbumin-exposed wild-type mice. In IL-5Ralpha KO mice, there was little increase of eosinophils in bronchoalveolar lavage fluid (BALF). The production of IL-5 in BALF increased in both mice after repeated antigen challenge, and there was no significant difference between wild-type and IL-5Ralpha KO mice. Similar to the BAL study, histological sections of lung tissue from ovalbumin-exposed wild-type mice exhibited airway eosinophilic inflammation, which was attenuated by the deficiency of IL-5Ralpha chain. There was no significant difference in serum antigen-specific IgE levels between wild-type and IL-5Ralpha KO mice after immunization nor antigen inhalation. Repeated antigen provocation caused BHR to acetylcholine in wild-type mice. In contrast, no BHR was observed in IL-5Ralpha KO mice after repeated inhalation of antigen.. These findings indicate that IL-5Ralpha plays an important role in the development of antigen-induced airway eosinophilia and BHR in mice.

Topics: Acetylcholine; Allergens; Animals; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Eosinophils; Immunoglobulin E; Inflammation; Lymphocytes; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Receptors, Interleukin; Receptors, Interleukin-5

Prominent eosinophil infiltration takes place in asthmatic bronchi, and damages bronchial epithelial cells.. This study was designed to investigate whether induction of apoptosis in infiltrated cells in the airways is beneficial or harmful.. A/J mice, which are genetically predisposed to be hyperresponsive to acetylcholine, were immunized with ovalbumin (OA) and alum. Thereafter, they were subjected to a 2-week regimen of OA inhalation, during which they were also administered either hamster anti-mouse Fas monoclonal antibody or hamster IgG (sham control) intranasally. Pulmonary function was then analyzed using whole-body plethysmography.. Inhalation of OA increased both airway responsiveness to acetylcholine and infiltration of eosinophils. Administration of anti-Fas antibody induced apoptosis in the infiltrating eosinophils and abolished the increase in airway responsiveness to acetylcholine.. Induction of apoptosis in eosinophils infiltrating asthmatic bronchi has a beneficial effect on airway hyperresponsiveness.

Topics: Acetylcholine; Animals; Antibodies, Monoclonal; Apoptosis; Bronchi; Bronchial Hyperreactivity; Cricetinae; Eosinophils; fas Receptor; Mice; Ovalbumin

The effects of lipid mediator antagonists: the LTD4-receptor antagonist pranlukast, the TXA2-receptor antagonist seratrodast, and the novel dual LTD4- and TXA2-receptor antagonist YM158 (3-[(4-tert-butylthiazol-2-yl)methoxy]-5'-[3-(4-chlorobenzenesu lfonyl) propyl]-2'-(1H-tetrazol-5-ylmethoxy)benzanilide monosodium salt monohydrate) were investigated in animals exhibiting immediate asthmatic response (IAR), late asthmatic response (LAR) and airway hyper-responsiveness (AHR). Antigen-induced LAR and AfR are inhibited by orally administered pranlukast (30, 100 mg/kg) and seratrodast (3, 10 mg/kg). YM158 (30 mg/kg), orally administered before or after IAR induction, also inhibited both LAR and AHR. However, while the inhibitory effects of pranlukast and seratrodast on IAR were marginal, the effects of YM158 (3, 10, 30 mg/kg) were dose-dependent, probably due to its multiple sites of action. Additionally, orally administered YM158 (30 mg/kg) inhibited ozone-induced AHR in guinea pigs. Thus, an antagonist that inhibits several lipid mediators might exhibit greater efficacy in treating asthmatic responses than antagonists with a single site of action. Therefore, YM158 shows great promise as a drug that will be able to treat bronchial asthma and related disorders more potently than currently used single-pathway inhibitors.

Topics: Acetylcholine; Airway Resistance; Animals; Anti-Asthmatic Agents; Antigens; Asthma; Benzoquinones; Bronchial Hyperreactivity; Bronchial Provocation Tests; Chromones; Dose-Response Relationship, Immunologic; Guinea Pigs; Heptanoic Acids; Hypersensitivity, Delayed; Hypersensitivity, Immediate; Leukotriene Antagonists; Male; Membrane Proteins; Ovalbumin; Ozone; Receptors, Leukotriene; Receptors, Thromboxane; Tetrazoles; Thiazoles

Transforming growth factor (TGF)-beta has been implicated in immunosuppression. However, it remains obscure whether regulation of T cells by TGF-beta contributes to the immunosuppression in vivo. To address this issue, we developed transgenic mice expressing Smad7, an intracellular antagonist of TGF-beta/Smad signaling, selectively in mature T cells using a plasmid construct coding a promoter element (the distal lck promoter) that directs high expression in peripheral T cells. Peripheral T cells were not growth inhibited by TGF-beta in Smad7 transgenic mice. Although Smad7 transgenic mice did not spontaneously show a specific phenotype, antigen-induced airway inflammation and airway reactivity were enhanced in Smad7 transgenic mice associated with high production of both T helper cell type 1 (Th1) and Th2 cytokines. Thus, blockade of TGF-beta/Smad signaling in mature T cells by expression of Smad7 enhanced airway inflammation and airway reactivity, suggesting that regulation of T cells by TGF-beta was crucial for negative regulation of the inflammatory (immune) response. Our findings also implicated TGF-beta/Smad signaling in mature T cells as a regulatory component of allergic asthma.

Topics: Animals; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Cytokines; DNA-Binding Proteins; Lymphocyte Activation; Mice; Mice, Transgenic; Ovalbumin; Smad7 Protein; T-Lymphocytes; Trachea; Trans-Activators; Transforming Growth Factor beta

Cysteinyl leukotrienes are potent inflammatory molecules playing a major role in asthma. The involvement of these mediators in hypersensitivity in mice is not well known. This study aimed at elucidating their implication by using MK-571, a cysLT(1)receptor antagonist. Mice were sensitized with a suspension of ovalbumin (8 microg) adsorbed to alum (2 mg) and were challenged with an aerosolized ovalbumin solution (0.5%). Inflammatory cell infiltration in the bronchoalveolar lavage (mostly eosinophils) following antigen challenge was inhibited by dexamethasone (0.1, 1 and 5 mg kg(-1)s.c.) and MK-571 (1, 10, 100 mg kg(-1)i.v.) in a dose-dependent manner. Maximal inhibition was 95% with 5 mg kg(-1)dexamethasone and 90% with 100 mg kg(-1)MK-571. When injected together they showed an additive inhibitory effect on eosinophil infiltration. Bronchial hyperreactivity, measured by the increased pulmonary insufflation pressure to carbachol injections, was also inhibited dose-dependently by MK-571. The EC(50)values for carbachol were of 22.39+/-1.12 microg kg(-1)in sensitized and challenged animals that did not receive MK-571 and increased to 43.65+/-1.10, 50.12+/-1.15 and 83.18+/-1.16 microg kg(-1)in animals treated with 1, 10 and 100 mg kg(-1)MK-571 respectively. Lung microvascular leakage (as measured by Evans blue extravasation) induced by antigen bronchoprovocation was reduced by 22% after treatment with 10 mg kg(-1)MK-571. All these inhibitory effects of MK-571 suggest a role for leukotriene D(4)in this animal model of allergic asthma.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Capillary Permeability; Dexamethasone; Disease Models, Animal; Drug Therapy, Combination; Leukotriene Antagonists; Leukotriene D4; Male; Membrane Proteins; Mice; Mice, Inbred BALB C; Ovalbumin; Propionates; Pulmonary Eosinophilia; Quinolines; Receptors, Leukotriene

The effects of repeated ozone exposures on the development of immune responses toward ovalbumin (OVA) were investigated in BALB/c and C57BL/6 mice. Ozone exposures (180 to 500 microg/m(3); 4 h, three times/wk for 4 wk) were combined with a protocol of OVA-aerosol exposure (1% OVA). Immediate cutaneous hypersensitivity (ICHS) reactions and antibody titers were assessed in parallel to cytokine levels of bronchoalveolar lavage fluids. In BALB/c mice, ozone triggered a T-helper (Th)2-like response indicated by dose-dependent increases in total serum immunoglobulin (Ig) E (from 133 to 821 ng/ml), interleukin (IL)-4 (from 60 to 208 pg/ml), and IL-5 levels (from 43 to 356 pg/ml), and by the recruitment of eosinophils and lymphocytes into the airways. Ozone exposure (500 microg/m(3)) in parallel to OVA-aerosol exposure increased anti-OVA IgG(1) antibody titers by 80%, leukotrienes (C(4)/D(4)/E(4)) by 60%, and airway responsiveness (11.3 versus 7.2 mg/ml methacholine), and doubled the frequency of positive ICHS reactions. In C57BL/6 mice, only the combination of OVA and ozone exposure induced positive ICHS reactions, doubled anti-OVA IgG(1), and suppressed anti-OVA IgG(2a) (-64%) antibody titers. Ozone, therefore, shifted the immune responses to OVA toward a Th2-like pattern in both "IgE-high responder" (BALB/c) and "IgE-low responder" (C57BL/6) mice.

Topics: Administration, Inhalation; Animals; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Dose-Response Relationship, Drug; Female; Hypersensitivity, Immediate; Immunoglobulin E; Immunoglobulin G; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Ovalbumin; Oxidants, Photochemical; Ozone; Th2 Cells

Aeroallergens continuously enter the respiratory tract of atopic individuals and provoke the development of asthma characterized by airway hyperreactivity (AHR) and inflammation. By contrast, nonatopic individuals are exposed to the same aeroallergens, but airway inflammation does not develop. However, the mechanisms that prevent allergen-induced respiratory diseases in nonatopic subjects are poorly characterized.. In this study we compared the role of allergen-specific T-cell tolerance and immune deviation in conferring protection against the development of allergen-induced AHR.. We exposed mice to intranasal ovalbumin (OVA) to induce T-cell tolerance and examined its effects on the subsequent development of AHR and inflammation.. We demonstrated that exposure of mice to intranasal OVA resulted in peripheral CD4(+) T-cell unresponsiveness that very efficiently prevented not only the development of AHR but also greatly inhibited airway inflammation and OVA-specific IgE production. The induction of peripheral T-cell tolerance and protection against AHR were not dependent on the presence of IFN-gamma or IL-4. The development of AHR was also prevented by an OVA-specific T(H)1-biased immune response induced by inhalation of OVA in the presence of IL-12. However, the OVA-specific T(H)1 response was associated with a significant degree of pulmonary inflammation.. These results indicate that both allergen-specific T-cell tolerance and T(H)1-biased immune deviation prevent the development of AHR, but T(H)1 responses are associated with significantly greater inflammation in the lung than is associated with T-cell unresponsiveness. Therefore CD4(+) T-cell unresponsiveness critically regulates immune responses to aeroallergens and protects against the development of allergic disease and asthma.

Topics: Allergens; Animals; Bronchial Hyperreactivity; Immune Tolerance; Immunization; Inflammation; Injections, Intraperitoneal; Interferon-gamma; Interleukin-4; Mice; Mice, Inbred BALB C; Ovalbumin; Th1 Cells; Th2 Cells

As TEI-9874, 2-(4-(6-cyclohexyloxy-2-naphtyloxy)phenylacetamide)ben zoic acid reduces allergen-specific immunoglobulin E (IgE) production by human peripheral blood mononuclear cells in vitro, we evaluated its potency on an allergen-induced asthmatic model in Brown-Norway rats. Inhaled ovalbumin induced the immediate-phase asthmatic response, the late-phase asthmatic response, the infiltration of leukocytes into bronchoalveolar lavage fluid, and an increase of serum anti-ovalbumin IgE. These parameters were suppressed by the treatment with TEI-9874 (3, 10, and 30 mg/kg p.o.). The ovalbumin-induced airway hyperresponsiveness was prevented by TEI-9874 (30 mg/kg p.o.). Furthermore, the suppression of the immediate-phase asthmatic response and the late-phase asthmatic response by TEI-9874 was almost completely extinguished by the exogenous administration of rat anti-ovalbumin antiserum. These results indicate that the efficacy of TEI-9874 on the asthmatic response is mainly mediated by the suppression of allergen-specific IgE production and TEI-9874 appears to be a good candidate as therapy for IgE-mediated allergic asthma.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Asthma; Benzeneacetamides; Benzoates; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Enzyme-Linked Immunosorbent Assay; Immunoglobulin E; Immunoglobulin G; Indicators and Reagents; Methacholine Chloride; Naphthalenes; Ovalbumin; Rats; Rats, Inbred BN; Respiratory Mechanics

Our purpose was to evaluate the expression of Th-1 and Th-2 related cytokine mRNA and nuclear factor (NF) kB in the lung tissue of ovalbumin (OA) sensitized brown Norway rats (BNR). We also evaluated the correlation between bronchial hyperreactivity (BHR) and eosinophils with cytokine mRNA expression.. Eight BNR (weight range 250-350 g) were sensitized by inhaled OA (group I) with a 1-week interval between and then provoked with OA 1 week later. Pulmonary function tests (PFT) were performed at baseline and 24 h after acetylcholine challenge. Eight weight-matched normal BNR served as controls (group II). All animals were anesthetized, paralyzed with gallamine, and ventilated via tracheostomy. They were given varying doses of acetylcholine (25, 50, 75, 100 microg/kg) injected through a jugular venous catheter. Five seconds after acetylcholine injections, PFTs were performed, including a maximal forced expiratory maneuver (MFEM), airway opening pressure (P(ao)) at tidal breathing and total dynamic lung compliance (C(dyn)). Bronchoalveolar lavage (BAL) was then performed with 20 ml normal saline divided into two doses. Thereafter, the lungs were removed and examined histologically. Total RNA was extracted from lung tissue samples and reverse-transcriptase polymerase chain reaction was performed using primers for mRNA of IL-4, IL-5, IL-10, interferon-gamma (IFNr) and beta-actine.. Group I OA treated rats had typical airway obstruction on PFTs and airway inflammation on histological examination. Ratios of IL-4, IL-5, IL-10 and inducible nitric oxide synthase (iNOS) mRNA levels to beta-actine as measured by densitometry were significantly lower in controls than in OA-sensitized rats. The IFNr mRNA to beta-actin ratio was significantly reduced in OA-sensitized rats. Group I demonstrated a band shift when compared with group II in electromobility shift assay (EMSA) for NF-kB indicating increased activation of this transcription factor.. Th-2 related cytokine mRNA was increased but Th-1 related cytokine mRNA was decreased in OA-sensitized BNR. An increased level of Th-2 related cytokine mRNA correlated with decreased airflow and inflammatory changes. These results demonstrate the value of the BNR model for studying allergic asthma at the molecular level.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cytokines; Eosinophils; Lung; Male; Maximal Expiratory Flow Rate; NF-kappa B; Ovalbumin; Rats; RNA, Messenger; Th2 Cells

Allergen-induced asthma is characterized by chronic pulmonary inflammation, reversible bronchoconstriction, and airway hyperreactivity to provocative stimuli. Multiple CC-chemokines, which are produced by pulmonary tissue in response to local allergen challenge of asthmatic patients or experimentally sensitized rodents, chemoattract leukocytes from the circulation into the lung parenchyma and airway, and may also modify nonchemotactic function. To determine the therapeutic potential of local intrapulmonary CC-chemokine blockade to modify asthma, a recombinant poxvirus-derived viral CC-chemokine inhibitor protein (vCCI), which binds with high affinity to rodent and human CC-chemokines in vitro and neutralizes their biological activity, was administered by the intranasal route. Administration of vCCI to the respiratory tract resulted in dramatically improved pulmonary physiological function and decreased inflammation of the airway and the lung parenchyma. In contrast, vCCI had no significant effect on the circulating levels of total or allergen-specific IgE, allergen-specific cytokine production by peripheral lymph node T cells, or peritoneal inflammation after local allergen challenge, indicating that vCCI did not alter systemic Ag-specific immunity or chemoattraction at extrapulmonary sites. Together, these findings emphasize the importance of intrapulmonary CC-chemokines in the pathogenesis of asthma, and the therapeutic potential of generic and local CC-chemokine blockade for this and other chronic diseases in which CC-chemokines are locally produced.

Topics: Administration, Intranasal; Animals; Anti-Asthmatic Agents; Anti-Inflammatory Agents, Non-Steroidal; Bronchial Hyperreactivity; Chemokines, CC; Cowpox virus; Disease Models, Animal; Female; Humans; Immunity, Cellular; Mice; Mice, Inbred BALB C; Ovalbumin; Recombinant Fusion Proteins; Respiratory Hypersensitivity; T-Lymphocytes; Viral Proteins; Virulence Factors

Viral respiratory infections have been implicated in influencing allergen sensitization and the development of asthma, but their exact role remains controversial. Because respiratory exposure to Ag normally engenders T cell tolerance and prevents the development of airway hyperreactivity (AHR) and inflammation, we examined the effects of influenza A virus infection on tolerance induced by exposure to intranasal (i.n.) OVA and the subsequent development of AHR. We found that concurrent infection with influenza A abrogated tolerance induced by exposure to i.n. OVA, and instead led to the development of AHR accompanied by the production of OVA-specific IgE, IL-4, IL-5, IL-13, and IFN-gamma. When both IL-4 and IL-5 were neutralized in this system, AHR was still induced, suggesting that influenza-induced cytokines such as IL-13, or mechanisms unrelated to cytokines, might be responsible for the development of AHR. The length of time between influenza A infection and i.n. exposure to OVA was crucial, because mice exposed to i.n. OVA 15-30 days after viral inoculation developed neither AHR nor OVA-specific tolerance. These mice instead acquired Th1-biased OVA-specific immune responses associated with vigorous OVA-induced T cell proliferation, and reduced production of OVA-specific IgE. The protective effect of influenza A on AHR was dependent on IFN-gamma, because protection was abrogated with a neutralizing anti-IFN-gamma mAb. These results suggest that viral respiratory infection interferes with the development of respiratory allergen-induced tolerance, and that the time interval between viral infection and allergen exposure is critical in determining whether viral infection will enhance, or protect against, the development of respiratory allergen sensitization and AHR.

Topics: Administration, Intranasal; Allergens; Animals; Asthma; Bronchial Hyperreactivity; CD4-Positive T-Lymphocytes; Disease Models, Animal; Humans; Immune Tolerance; Influenza A virus; Influenza, Human; Interferon-gamma; Interleukin-4; Interleukin-5; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Respiratory Hypersensitivity; Time Factors

The development of airway hyperresponsiveness (AHR) is correlated with the infiltration into the lungs of activated eosinophils and T lymphocytes. In large part, influx of eosinophils into the lung is dependent on very late activating antigen-4 (VLA-4) expression. However, the kinetics of eosinophil recruitment and the development of AHR are not fully delineated. Airway function was monitored by changes in lung resistance (RL) and dynamic compliance (Cdyn) to methacholine (MCh) inhalation after anti-VLA-4. After ovalbumin (OVA) sensitization and airway challenge of BALB/c mice, AHR increased as did the number of lung inflammatory cells. Administration of anti-VLA-4 to sensitized mice 2 h before the first (of three) OVA airway challenges significantly prevented changes in RL. Moreover, injection of the antibody from 2 h before the first challenge to 42 h after the last challenge significantly prevented the increases in RL, as well as eosinophil and lymphocyte numbers in the bronchoalveolar lavage fluid (BALF); interleukin-5 (IL-5) and leukotriene concentrations in BALF were also significantly inhibited. Interestingly, treatment with anti-VLA-4 only prevented changes in Cdyn and goblet cell hyperplasia when administered 2 h before the first challenge. These studies demonstrate that the timing of anti-VLA-4 administration can selectively affect pathologic processes that contribute to altered airway function in the central and peripheral airways after allergen challenge.

Topics: Airway Resistance; Animals; Antibodies, Monoclonal; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchial Provocation Tests; Eosinophils; Female; Integrin alpha4beta1; Integrins; Lung; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Ovalbumin; Receptors, Lymphocyte Homing

Tumour necrosis factor-alpha (TNF-alpha) and interleukin 1beta (IL-1beta) have been implicated in the pathogenesis of asthma. The p38 kinase inhibitor, SB 203580 inhibits TNF-alpha and IL-1beta production in vitro and in vivo. In this study the effect of SB 203580 on allergen-induced airway TNF-alpha production and inflammatory cell recruitment was investigated in sensitized Brown Norway rats. The allergen-induced increase in bronchoalveolar lavage (BAL) TNF-alpha was inhibited by SB 203580 at every dose tested (10 - 100 mg kg(-1), p.o.). In contrast, neither ovalbumin-induced eosinophilia or neutrophilia were inhibited by SB 203580 (10 - 100 mg kg(-1), p.o.). In conclusion, SB 203580 inhibits BAL TNF-alpha production by 95% without inhibiting either antigen-induced airway eosinophilia or neutrophilia. This data suggests that either the residual TNF-alpha is sufficient to drive allergen-induced inflammatory cell recruitment into the lung or that TNF-alpha is not involved in allergen-induced inflammatory cell recruitment.

Topics: Administration, Oral; Animals; Anti-Inflammatory Agents; Bronchial Hyperreactivity; Bronchial Provocation Tests; Bronchoalveolar Lavage; Cell Movement; Disease Models, Animal; Enzyme Inhibitors; Female; Hypersensitivity; Imidazoles; Mitogen-Activated Protein Kinases; Ovalbumin; p38 Mitogen-Activated Protein Kinases; Pyridines; Rats; Tumor Necrosis Factor-alpha

Chronic Th2-dominated inflammation and exaggerated IL-6 production are characteristic features of the asthmatic airway. To understand the processes that are responsible for the chronicity of this response and the role(s) of IL-6 in the regulation of airway Th2 inflammation, we compared the responses induced by OVA in sensitized wild-type mice, IL-6 deficient (-/-) mice, and transgenic mice in which IL-6 was overexpressed in the airway (CC10-IL-6 mice). When compared with wild-type mice, IL-6-/- mice manifest exaggerated inflammation and eosinophilia, increased levels of IL-4, IL-5, and IL-13 protein and mRNA, exaggerated levels of eotaxin, JE/monocyte chemoattractant protein-1, macrophage inflammatory protein-1alpha and -2, and mRNA, increased bronchoalveolar lavage (BAL) TGF-beta1, and exaggerated airway responses to aerosolized methacholine. In contrast, CC10-IL-6 mice, on both C57BL/6 and BALB/c backgrounds, manifest diminished inflammation and eosinophilia, decreased levels of IL-4, IL-5, and IL-13 protein and mRNA, and decreased levels of bronchoalveolar lavage TGF-beta1. IL-6 also decreased the expression of endothelial VCAM-1 and airway responsiveness to methacholine in these animals. These alterations in the IL-6-/- and CC10-IL-6 mice were not associated with significant decreases or increases in the levels of IFN-gamma, respectively. These studies demonstrate that endogenous and exogenous IL-6 inhibit aeroallergen-induced Th2 inflammation and that this inhibition is not mediated by regulatory effects of IFN-gamma. IL-6 may be an important anti-inflammatory, counterregulatory, and healing cytokine in the airway.

Topics: Administration, Intranasal; Aerosols; Allergens; Animals; Bronchial Hyperreactivity; Chemokines; Cytokines; Gene Expression Regulation; Inflammation; Interleukin-6; Lung; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Inbred CBA; Mice, Knockout; Mice, Transgenic; Ovalbumin; Plethysmography, Whole Body; Pulmonary Eosinophilia; RNA, Messenger; Th2 Cells; Vascular Cell Adhesion Molecule-1

We demonstrated previously that CD81(-/-) mice have an impaired Th2 response. To determine whether this impairment affected allergen-induced airway hyperreactivity (AHR), CD81(-/-) BALB/c mice and CD81(+/+) littermates were sensitized i.p. and challenged intranasally with OVA. Although wild type developed severe AHR, CD81(-/-) mice showed normal airway reactivity and reduced airway inflammation. Nevertheless, OVA-specific T cell proliferation was similar in both groups of mice. Analysis of cytokines secreted by the responding CD81(-/-) T cells, particularly those derived from peribronchial draining lymph nodes, revealed a dramatic reduction in IL-4, IL-5, and IL-13 synthesis. The decrease in cytokine production was not due to an intrinsic T cell deficiency because naive CD81(-/-) T cells responded to polyclonal Th1 and Th2 stimulation with normal proliferation and cytokine production. Moreover, there was an increase in T cells and a decrease in B cells in peribronchial lymph nodes and in spleens of immunized CD81(-/-) mice compared with wild-type animals. Interestingly, OVA-specific Ig levels, including IgE, were similar in CD81(-/-) and CD81(+/+) mice. Thus, CD81 plays a role in the development of AHR not by influencing Ag-specific IgE production but by regulating local cytokine production.

Topics: Administration, Intranasal; Allergens; Animals; Antigens, CD; B-Lymphocytes; Bronchial Hyperreactivity; Cytokines; Down-Regulation; Eosinophilia; Epitopes, T-Lymphocyte; Immunoglobulin E; Immunologic Deficiency Syndromes; Inflammation; Injections, Intraperitoneal; Interphase; Lung; Lymph Nodes; Lymphocyte Activation; Lymphocyte Count; Lymphopenia; Membrane Proteins; Mice; Mice, Inbred BALB C; Mice, Knockout; Mucins; Ovalbumin; Species Specificity; Spleen; Tetraspanin 28; Th1 Cells; Th2 Cells

The relationship between eosinophils and the development of Ag-induced pulmonary pathologies, including airway hyper-responsiveness, was investigated using mice deficient for the secondary granule component, major basic protein-1 (mMBP-1). The loss of mMBP-1 had no effect on OVA-induced airway histopathologies or inflammatory cell recruitment. Lung function measurements of knockout mice demonstrated a generalized hyporeactivity to methacholine-induced airflow changes (relative to wild type); however, this baseline phenotype was observable only with methacholine; no relative airflow changes were observed in response to another nonspecific stimulus (serotonin). Moreover, OVA sensitization/aerosol challenge of wild-type and mMBP-1(-/-) mice resulted in identical dose-response changes to either methacholine or serotonin. Thus, the airway hyper-responsiveness in murine models of asthma occurs in the absence of mMBP-1.

Topics: Allergens; Animals; Antigens, Helminth; Asthma; Blood Proteins; Bronchial Hyperreactivity; Cell Movement; Cytoplasmic Granules; Disease Models, Animal; Eosinophil Granule Proteins; Eosinophils; Gene Deletion; Injections, Intraperitoneal; Lung; Mesocestoides; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Knockout; Microscopy, Electron; Ovalbumin; Ribonucleases

The kinetics of airway inflammation and remodeling processes following ovalbumin aerosol challenge in sensitized BALB/c mice was studied. Mice were exposed to either single or five ovalbumin challenges over 5 days. In both protocols, time-dependent increases in bronchoalveolar lavage (BAL) cellular fibronectin, neutrophils and eosinophils were observed. The kinetics of these events were similar in both protocols; however, the magnitude of the response was much greater following repeated challenges. BAL protein levels and lymphocyte numbers were increased only following repeated challenges, whereas interleukin (IL)-5 and IL-4 were increased in both protocols. Histological analysis revealed a time-dependent increase in epithelial cell proliferation and in mucus-producing epithelial cells. Proliferation of alveolar cells was observed only following repeated challenges. Airway hyperreactivity was observed in both protocols but was much greater following repeated challenges. Pretreatment with dexamethasone fully inhibited the inflammatory response and airway hyperreactivity but only partially inhibited the remodeling process. These data suggest that glucocorticoids, although potent anti-inflammatory agents, may not be potent in reducing the lung remodeling process associated with asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Bronchoconstrictor Agents; Cell Division; Dexamethasone; Disease Models, Animal; Eosinophils; Epithelial Cells; Fibronectins; Immunoglobulin E; Interleukin-4; Interleukin-5; Leukocyte Count; Male; Methacholine Chloride; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mucus; Ovalbumin; Plasma; Pulmonary Alveoli; Respiratory Mucosa; Species Specificity; Time Factors

Monocyte chemoattractant proteins-1 and -5 have been implicated as important mediators of allergic pulmonary inflammation in murine models of asthma. The only identified receptor for these two chemokines to date is the CCR2. To study the role of CCR2 in a murine model of Ag-induced asthma, we compared the pathologic and physiological responses of CCR2(-/-) mice with those of wild-type (WT) littermates following immunization and challenge with OVA. OVA-immunized/OVA-challenged (OVA/OVA) WT and CCR2(-/-) mice developed significant increases in total cells recovered by bronchoalveolar lavage (BAL) compared with their respective OVA-immunized/PBS-challenged (OVA/PBS) control groups. There were no significant differences in BAL cell counts and differentials (i.e., macrophages, PMNs, lymphocytes, and eosinophils) between OVA/OVA WT and CCR2(-/-) mice. Serologic evaluation revealed no significant difference in total IgE and OVA-specific IgE between OVA/OVA WT mice and CCR2(-/-) mice. Lung mRNA expression and BAL cytokine protein levels of IL-4, IL-5, and IFN-gamma were also similar in WT and CCR2(-/-) mice. Finally, OVA/OVA CCR2(-/-) mice developed increased airway hyper-responsiveness to a degree similar to that in WT mice. We conclude that following repeated airway challenges with Ag in sensitized mice, the development of Th2 responses (elevated IgE, pulmonary eosinophilia, and lung cytokine levels of IL-4 and IL5) and the development of airway hyper-responsiveness are not diminished by a deficiency in CCR2.

Topics: Animals; Antibody Specificity; Antigens; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemotaxis, Leukocyte; Cytokines; Eosinophil Peroxidase; Eosinophils; Immunoglobulin E; Injections, Intraperitoneal; Lung; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Peroxidases; Pulmonary Eosinophilia; Receptors, CCR2; Receptors, Chemokine; Ribonucleases; RNA, Messenger

Docosahexaenoic acid (DHA) found in fish oil is known to depress inflammation-related mediators. We investigated a novel delivery method of tridocosahexaenoyl-glycerol (DHA-TG).. BALB/c mice (6-8 weeks old) were primed intraperitoneally with ovalbumin (OVA) and Al(OH)(3) on days 0 and 7, and with aerosolized OVA on day 7. Primed mice were challenged by repeated exposure to aerosolized OVA on days 15-17. Just before each exposure to aerosolized OVA, the mice were also exposed to an aerosol of emulsified DHA-TG or soybean oil, or saline (days 7 and 15-17). Bronchial hyperresponsiveness (BHR) to methacholine was measured, and bronchoalveolar lavage fluid was obtained 24 h after the last challenge (day 18). Lungs were histologically examined.. Bronchoalveolar lavage fluid of saline-treated mice showed an increased cellularity with predominant eosinophils. Exposure to DHA-TG significantly reduced the total cell number and the eosinophil percentage in lavage fluid, whereas soybean oil did not.. DHA but not soybean oil exposure reduced BHR and cell infiltration to bronchovascular bundles. This type of DHA administration could be studied in clinical trials.

Topics: Aerosols; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Docosahexaenoic Acids; Emulsions; Eosinophils; Female; Humans; Lung; Mice; Mice, Inbred BALB C; Ovalbumin