transforming-growth-factor-beta and Asthma

Structural changes involving tissue remodelling and fibrosis are major features of many pulmonary diseases, including asthma, chronic obstructive pulmonary disease (COPD) and idiopathic pulmonary fibrosis (IPF). Abnormal deposition of extracellular matrix (ECM) proteins is a key factor in the development of tissue remodelling that results in symptoms and impaired lung function in these diseases. Tissue remodelling in the lungs is complex and differs between compartments. Some pathways are common but tissue remodelling around the airways and in the parenchyma have different morphologies. Hence it is critical to evaluate both common fibrotic pathways and those that are specific to different compartments; thereby expanding the understanding of the pathogenesis of fibrosis and remodelling in the airways and parenchyma in asthma, COPD and IPF with a view to developing therapeutic strategies for each. Here we review the current understanding of remodelling features and underlying mechanisms in these major respiratory diseases. The differences and similarities of remodelling are used to highlight potential common therapeutic targets and strategies. One central pathway in remodelling processes involves transforming growth factor (TGF)-β induced fibroblast activation and myofibroblast differentiation that increases ECM production. The current treatments and clinical trials targeting remodelling are described, as well as potential future directions. These endeavours are indicative of the renewed effort and optimism for drug discovery targeting tissue remodelling and fibrosis.

Topics: Airway Remodeling; Asthma; Calcium-Binding Proteins; Extracellular Matrix; Fibroblasts; Fibrosis; Glycoproteins; Humans; Idiopathic Pulmonary Fibrosis; Lung Diseases; Matrix Metalloproteinases; Pulmonary Disease, Chronic Obstructive; Transforming Growth Factor beta

Topics: Animals; Asthma; Chronic Disease; Humans; Multigene Family; Rhinitis, Allergic; Signal Transduction; Sinusitis; Transforming Growth Factor beta

The study of developmental trajectories is where epigenetics truly shines. The "epi" in epigenetics captures the fact that although epigenetic processes also preside over the maintenance and termination of gene expression, the unfolding and remodeling of chromatin architecture are especially critical to prepare genes for regulated transcription. These properties imply being on a path, a trajectory to events that will occur later thanks to epigenetic programming. Thus epigenetics is about timed and timely events. In this article we discuss epigenetic and genetic evidence from several independent studies of asthma, chronic obstructive pulmonary disease, and lung function, which converge to highlight a potential role of the TGF-β gene pathway in these processes. These results raise the possibility that at least in a subset of subjects, these conditions might be functionally connected in ways that need to be further defined but that likely reflect the uniquely pleiotropic nature of TGF-β pathway genes, particularly their ability to control both lung development and immune responses essential for regulation and inflammation. Further characterization of this pathway in longitudinally phenotyped populations might unmask novel trajectories to lung disease that begin in utero and unfold into old age.

Topics: Adult; Animals; Asthma; Child; Epigenesis, Genetic; Genetic Pleiotropy; Humans; Pulmonary Disease, Chronic Obstructive; Signal Transduction; Transforming Growth Factor beta

Asthma is the most common chronic pulmonary disease in China and is characterized by airway inflammation and episodic airflow obstruction. The aim of this meta-analysis was to evaluate the relation of two transforming growth factor-β1 (TGF-β1) polymorphisms with asthma risk in Chinese population.. PubMed, Springer, EMBASE, MEDLINE, CNKI (China National Knowledge Infrastructure), and Wanfang databases were used to search and retrieve relevant eligible case-control studies published through December 2017. The odds ratios (OR) and 95% confidence intervals (CI) were used to evaluate the effect.. A total of 2040 asthma patients and 1952 controls from 12 studies were analyzed. Two polymorphic sites of TGF-β1 gene were identified: -509C/T and +869T/C. We found that the -509C/T polymorphism was associated with increased asthma risk under the heterozygous model (CT vs. CC: OR = 1.40, 95% CI = 1.03-1.90, p = 0.03) and the dominant model (TT+CT vs. CC: OR = 1.41, 95% CI = 1.05-1.90, p = 0.02). Subgroup analyses by age suggested that -509C/T variant was associated with childhood asthma. Analysis of disease severity indicated that this variant was associated with both mild-to-moderate asthma and severe asthma. However, the +869T/C polymorphism was not associated with asthma susceptibility in subgroup analysis by age or disease severity.. This study demonstrated that the -509C/T polymorphism of the TGF-β1 gene might be a risk factor for asthma in the Chinese population, especially in Chinese children. Further large-scale case-control studies are still required.

Topics: Asian People; Asthma; Case-Control Studies; China; Female; Genetic Association Studies; Genetic Predisposition to Disease; Genotype; Humans; Male; Odds Ratio; Polymorphism, Genetic; Risk Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1

Transforming growth factor (TGF)-β is an evolutionarily conserved pleiotropic factor that regulates a myriad of biological processes including development, tissue regeneration, immune responses, and tumorigenesis. TGF-β is necessary for lung organogenesis and homeostasis as evidenced by genetically engineered mouse models. TGF-β is crucial for epithelial-mesenchymal interactions during lung branching morphogenesis and alveolarization. Expression and activation of the three TGF-β ligand isoforms in the lungs are temporally and spatially regulated by multiple mechanisms. The lungs are structurally exposed to extrinsic stimuli and pathogens, and are susceptible to inflammation, allergic reactions, and carcinogenesis. Upregulation of TGF-β ligands is observed in major pulmonary diseases, including pulmonary fibrosis, emphysema, bronchial asthma, and lung cancer. TGF-β regulates multiple cellular processes such as growth suppression of epithelial cells, alveolar epithelial cell differentiation, fibroblast activation, and extracellular matrix organization. These effects are closely associated with tissue remodeling in pulmonary fibrosis and emphysema. TGF-β is also central to T cell homeostasis and is deeply involved in asthmatic airway inflammation. TGF-β is the most potent inducer of epithelial-mesenchymal transition in non-small cell lung cancer cells and is pivotal to the development of tumor-promoting microenvironment in the lung cancer tissue. This review summarizes and integrates the current knowledge of TGF-β signaling relevant to lung health and disease.

Topics: Animals; Asthma; Humans; Lung; Lung Diseases; Lung Neoplasms; Pulmonary Emphysema; Pulmonary Fibrosis; Signal Transduction; Transforming Growth Factor beta

Asthma is a chronic heterogeneous illness of the lower airway with an inflammatory basis, developing from hyperresponsiveness and bronchial obstruction. One of the more unfavourable processes occurring in the airway are the long-term changes of the respiratory tract known as remodelling, resulting in complete irreversible obstruction. Bone morphogenetic protein (BMP) is a member of the Transforming Growth Factor beta (TGF-b) superfamily, which regulates processes in embryonic and post-embryonic development. The role played by BMP is regulation of degradation and remodelling of the extracellular matrix, which is one of the elements involved in the reconstruction of the structure of the bronchi in severe asthma. This paper presents the antagonistic properties of BMP against TGF-b, anti-inflammatory and counteracting fibrosis in the respiratory tract. The current state of knowledge indicates that this group of cytokines are potential new markers of remodelling in severe asthma, and further studies on their therapeutic value are necessary.

Topics: Asthma; Bone Morphogenetic Protein 7; Bone Morphogenetic Proteins; Bronchi; Extracellular Matrix; Humans; Transforming Growth Factor beta; Transforming Growth Factor beta1

Asthma is a chronic inflammatory heterogeneous disease of the lower respiratory tract characterised by the occurrence of bronchial hyper-responsiveness and paroxysmal, changeable bronchial obstruction. Transforming growth factor-beta (TGF-b) is one of the cytokines involved in mediating airway inflammation and remodelling. The level of TGF-b1 gene expression correlates with severity of symptoms. Alterations in the main SMAD signal transmission, overexpression of TGF-b genes and changes in the transcriptome cause excessive secretion of TGF-b and its increased expression in target cells, which clinically induces a moderate-severe or severe course of asthma as well as an earlier and faster disease progression. Knowledge of these processes allows clinicians to assess immune responses in patients, which affects adequate disease control and prevention of remodelling.

Topics: Airway Remodeling; Asthma; Gene Expression; Humans; Signal Transduction; Smad Proteins; Transcription, Genetic; Transforming Growth Factor beta

The complement mediators are the major effectors of the immune balance, which operates at the interface between the innate and adaptive immunity, and is vital for many immunoregulatory functions. Activation of the complement cascade through the classical, alternative or lectin pathways thus generating opsonins like C3b and C5b, anaphylatoxins C3a and C5a, chemotaxin, and inflammatory mediators, which leads to cellular death. Complement mediators that accelerate the airway remodeling are not well defined; however, an uncontrolled Th2-driven adaptive immune response has been linked to the major pathophysiologic features of asthma, including bronchoconstriction, airway hyperresponsiveness, and airway inflammation. The mechanisms leading to complement mediated airway tissue remodeling, and the effect of therapy on preventing and/or reversing it are not clearly understood. This review highlights complement-mediated inflammation, and the mechanism through it triggers the airway tissue injury and remodeling in the airway epithelium that could serve as potential targets for developing a new drug to rescue the asthma patients.

Topics: Airway Remodeling; Anaphylatoxins; Animals; Asthma; Chemotactic Factors; Complement Activation; Complement C3a; Complement C5a; Humans; Immunity, Innate; Inflammation; Inflammation Mediators; Interleukin-13; Opsonin Proteins; Th2 Cells; Transforming Growth Factor beta

Asthma markedly diminishes quality of life due to limited activity, absences from work or school and hospitalizations. Patients with severe asthma which are not controlled despite taking effective therapy are most in need of new treatment approaches. IL-13 was demonstrated as 'central mediator of allergic asthma'.. IL-13 has been implicated in the pathogenesis of asthma, idiopathic pulmonary fibrosis and COPD. IL-13 levels in the sputum and bronchial biopsy samples remain elevated in severe asthma despite the use of inhaled and systemic corticosteroids. Thus, IL-13 is a mediator involved in corticosteroid resistance. Periostin enhances profibrotic TGF-β signaling in subepithelial fibrosis associated with asthma. IL-13 induces bronchial epithelial cells to secrete periostin. Periostin may be a biomarker for Th2 induced airway inflammation. Lebrikizumab is a monoclonal antibody against IL-13. Lebrikizumab improved lung function in asthmatics who were symptomatic despite treatment with long acting beta agonist and inhaled corticosteroids and provided benefit in the treatment of severe uncontrolled asthma.. Lebrikizumab block IL-13 signaling through the IL-13Rα1/IL-4Rα receptor. There was a larger reduction in FENO in the high periostin subgroup than in the low periostin subgroup (34.4 vs 4.3%). Serum CCL17, CCL13 and total IgE levels decreased in the lebrikizumab group.

Topics: Adrenal Cortex Hormones; Animals; Antibodies, Monoclonal; Asthma; Biological Products; Biomarkers; Bronchi; Cell Adhesion Molecules; Eosinophils; Humans; Hypersensitivity; Inflammation; Interleukin-13; Interleukin-33; Interleukin-4; Interleukins; Lung; Nitric Oxide; Quality of Life; Transforming Growth Factor beta

Asthma is a chronic inflammatory airway disease involving complex interplay between resident and infiltrative cells, which in turn are regulated by a wide range of host mediators. Identifying useful biomarkers correlating with clinical symptoms and degree of airway obstruction remain important to effective future asthma treatments. Transforming growth factor β (TGF-β) is a major mediator involved in pro-inflammatory responses and fibrotic tissue remodeling within the asthmatic lung. Its role however, as a therapeutic target remains controversial. The aim of this review is to highlight its role in severe asthma including interactions with adaptive T-helper cells, cytokines and differentiation through regulatory T-cells. Associations between TGF-β and eosinophils will be addressed and the effects of genetic polymorphisms of the TGF-β1 gene explored in the context of asthma. We highlight TGF-β1 as a potential future therapeutic target in severe asthma including its importance in identifying emerging clinical phenotypes in asthmatic subjects who may be suitable for individualized therapy through TGF-β modulation.

Topics: Asthma; Cytokines; Eosinophils; Humans; Polymorphism, Genetic; T-Lymphocytes, Regulatory; Th2 Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1

The JmjC protein Mina is an important immune response regulator. Classical forward genetics first discovered its immune role in 2009 in connection with the development of T helper 2 (Th2) cells. This prompted investigation into Mina's role in the two best-studied contexts where Th2 responses are essential: atopic asthma and helminth expulsion. In work focused on a mouse model of atopic asthma, Mina deficiency was found to ameliorate airway hyper-resistance and pulmonary inflammation. And, in a case-control study genetic variation at the human MINA locus was found to be associated with the development of childhood atopic asthma. Although the underlying cellular and molecular mechanism of Mina's involvement in pulmonary inflammation remains unknown, our recent work on parasitic helminth expulsion suggests the possibility that, rather than T cells, epithelial cells responding to TGFβ may play the dominant role. Here we review the growing body of literature on the emerging Mina pathway in T cells and epithelial cells and attempt to set these into a broader context.

Topics: Animals; Asthma; Dioxygenases; Disease Models, Animal; Genetic Loci; Genetic Variation; Helminthiasis; Histone Demethylases; Humans; Inflammation; Mice; Neoplasm Proteins; Nuclear Proteins; Th2 Cells; Transforming Growth Factor beta

The airway smooth muscle (ASM) plays an important role in the pathophysiology of asthma and chronic obstructive pulmonary disease (COPD). ASM cells express a wide range of receptors involved in contraction, growth, matrix protein production and the secretion of cytokines and chemokines. Transforming growth factor beta (TGF-β) is one of the major players in determining the structural and functional abnormalities of the ASM in asthma and COPD. It is increasingly evident that TGF-β functions as a master switch, controlling a network of intracellular and autocrine signaling loops that effect ASM phenotype and function. In this review, the various elements that participate in non-canonical TGF-β signaling, including MAPK, PI3K, WNT/β-catenin, and Ca(2+), are discussed, focusing on their effect on ASM phenotype and function. In addition, new aspects of ASM biology and their possible association with non-canonical TGF-β signaling will be discussed.

Topics: Animals; Asthma; Chemokines; Cytokines; Humans; Muscle, Smooth; Myocytes, Smooth Muscle; Phenotype; Pulmonary Disease, Chronic Obstructive; Signal Transduction; Transforming Growth Factor beta

Fibrosis is one of the key pathological features of airway remodeling in asthma. In the normal airway the amount of collagen and other extracellular matrix components is kept in equilibrium by regulation of synthesis and degradation. In asthma this homeostasis is disrupted due to genetic and environmental factors. In the airways of patients with the disease there is increased extracellular matrix deposition, particularly in the reticular basement membrane region, lamina propria and submucosa. Fibrosis is important as it can occur early in the pathogenesis of asthma, be associated with severity and resistant to therapy. In this review we will discuss current knowledge of relaxin and other key regulators of fibrosis in the airway including TGFβ, Smad2/3 and matrix metalloproteinases. As fibrosis is not directly targeted or effectively treated by current asthma drugs including corticosteroids, characterization of airway fibrosis and how it is regulated will be essential for the development of novel therapies for asthma.

Topics: Airway Remodeling; Asthma; Collagen; Extracellular Matrix; Humans; Matrix Metalloproteinase Inhibitors; Matrix Metalloproteinases; Pulmonary Fibrosis; Relaxin; Respiratory System; Smad2 Protein; Smad3 Protein; Tissue Inhibitor of Metalloproteinases; Transforming Growth Factor beta

TGF-β is one of the main mediators involved in tissue remodeling in the asthmatic lung. This profibrotic cytokine is produced by a number of cells, including macrophages, epithelial cells, fibroblasts, and eosinophils. High expression of TGF-β in patients with asthma was reported by many investigators. However, controversy remains whether the concentration of TGF-β correlates with disease severity. TGF-β is believed to play an important role in most of the cellular biological processes leading to airway remodeling. It was shown to be involved in epithelial changes, subepithelial fibrosis, airway smooth muscle remodeling, and microvascular changes. Here, sources of TGF-β, as well as its role in the development of airway remodeling, will be reviewed. Therapeutic strategies that modulate TGF-β will also be discussed.

Topics: Airway Remodeling; Animals; Asthma; Eosinophils; Goblet Cells; Humans; Inflammation Mediators; Lung; Microcirculation; Mucus; Muscle, Smooth; Pulmonary Circulation; Pulmonary Fibrosis; Respiratory Mucosa; Signal Transduction; Transforming Growth Factor beta

Activin-A is a pleiotropic cytokine that belongs to the TGF-β superfamily and plays an important role in fundamental biological processes, such as development and tissue repair. Growing evidence proposes a crucial role for activin-A in immune-mediated responses and associated diseases, with both enhancing and suppressive effects depending on the cell type, the cytokine micromilieu and the context of the response. Several recent studies have demonstrated a striking increase in activin-A expression in experimental models of asthma, as well as, in the asthmatic airway in humans. Importantly, a strong immunoregulatory role for activin-A in allergic airway disease, with suppression of T helper (Th) type 2 cell-driven allergic responses and protection against the development of cardinal features of the asthmatic phenotype was revealed by in vivo functional studies. Activin-A-mediated immunosuppression is associated with induction of functional allergen-specific regulatory T cells. In human asthma, although activin-A levels are increased in the airway epithelium and submucosal cells, the expression of its signalling components is markedly decreased, pointing to decreased regulation. Nevertheless, a rapid activation of the activin-A signalling pathway is observed in the airway of individuals with asthma following inhalational allergen challenge, suggestive of an inherent protective mechanism to control disease. In support, in vitro studies using human airway epithelial cells have demonstrated that endogenous activin-A suppresses the release of inflammatory mediators, while it induces epithelial repair. Collectively, compelling evidence suggests that activin-A orchestrates the regulation of key events involved in the pathogenesis of allergic asthma. The critical role of activin-A in allergic airway responses places this cytokine as an exciting new therapeutic target for asthma.

Topics: Activins; Airway Remodeling; Animals; Asthma; Cytokines; Humans; Inflammation; Signal Transduction; Transforming Growth Factor beta

Chronic rhinosinusitis (CRS) is a heterogeneous group of inflammatory diseases of the nasal and paranasal cavities either accompanied by polyp formation (CRSwNP) or without polyps (CRSsNP). CRSsNP and CRSwNP are prevalent medical conditions associated with substantial impaired quality of life, reduced workplace productivity, and serious medical treatment costs. Despite recent research evidence that contributes to further unveiling the pathophysiology of these chronic airway conditions, the cause remains poorly understood and appears to be multifactorial. A diverse spectrum of alterations involving histopathology, inflammatory cell and T-cell patterns, remodeling parameters (eg, TGF-β), eicosanoid and IgE production, microorganisms, and epithelial barrier malfunctions is reported in the search to describe the pathogenesis of this heterogeneous group of upper airway diseases. Furthermore, novel evidence indicates considerable heterogeneity within the CRSwNP subgroup determining the risk of comorbid asthma. The characterization of specific disease subgroups is a challenging scientific and clinical task of utmost importance in the development of diagnostic tools and application of individualized treatments. This review focuses on recent evidence that sheds new light on our current knowledge regarding the inflammatory process of CRS to further unravel its pathogenesis.

Topics: Animals; Asthma; Chronic Disease; Eicosanoids; Humans; Immunoglobulin E; Inflammation; Rhinitis; Risk Factors; Sinusitis; T-Lymphocytes; Transforming Growth Factor beta

Fibrogenic lung reactions occur as a common phenotype shared among disorders of heterogeneous etiologies. Even with a common etiology, the extent and pattern of fibrosis vary greatly among individuals, even within families, suggesting complex gene-environment interactions. The search for mechanisms shared among all fibrotic lung diseases would represent a major advance in the identification of therapeutic targets that could have a broad impact on lung health. Although it is difficult to grasp all of the complexities of the varied cell types and cytokine networks involved in lung fibrogenic responses, and to predict the biologic responses to the overexpression or deficiency of individual cytokines, a large body of evidence converges on a single common theme: the central importance of the transforming growth factor beta (TGF-beta) pathway. Therapies that act upstream or downstream of TGF-beta activation have the therapeutic potential to treat all fibrogenic responses in the lung.

Topics: Asthma; Emphysema; Fibroblasts; Humans; Lung Diseases; Pulmonary Disease, Chronic Obstructive; Pulmonary Fibrosis; Transforming Growth Factor beta

That regulatory T cells (Tregs) have a crucial role in controlling allergic diseases such as asthma is now undisputed. The cytokines most commonly implicated in Treg-mediated suppression of allergic asthma are transforming growth factor-beta (TGF-beta) and interleukin (IL)-10). In addition to naturally occurring Tregs, adaptive Tregs, induced in response to foreign antigens, have been shown in recent studies. The concept of inducible/adaptive Tregs (iTregs) has considerable significance in preventing asthma if generated early enough in life. This is because cytokines such as IL-4 and IL-6 inhibit Foxp3 induction in naive CD4+ T cells and therefore de novo generation of Tregs can be expected to be less efficient when it is concomitant with effector cell development in response to an allergen. However, if iTregs can be induced, the process of infectious tolerance would facilitate expansion of the iTreg pool as suggested in the recent literature. It is tempting to speculate that there is a window of opportunity in early life in the context of a relatively immature immune system that is permissive for the generation of iTregs specific to a spectrum of allergens that would regulate asthma for lifelong. The focus of this review is the relevance of nTregs and iTregs in controlling asthma from early life into adulthood, the mechanisms underlying Treg function, and the prospects for using our current concepts to harness the full potential of Tregs to limit disease development and progression.

Topics: Allergens; Animals; Asthma; Forkhead Transcription Factors; Humans; Immune Tolerance; Interleukin-10; Interleukin-4; Interleukin-6; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Asthma is characterized by T helper cell 2 (Th2) type inflammation, leading to airway hyperresponsiveness and tissue remodeling. Th2 cell-driven inflammation is likely to represent an abnormal response to harmless airborne particles. These reactions are normally suppressed by regulatory T cells, which maintain airway tolerance. The anti-inflammatory cytokine IL-10 is likely to play a central role. The role of the cytokine transforming growth factor beta (TGF-beta) is more complex, with evidence for immune suppression and remodeling in the airways. In asthmatic individuals there is a breakdown in these regulatory mechanisms. There is emerging evidence that early life events, including exposure to allergen and infections, are critical in programming effective regulatory pathways to maintain pulmonary homeostasis. In this review we examine the clinical and experimental evidence for T regulatory cell function in the lung and discuss the events that might influence the functioning of these cells. Ultimately, the ability to enhance regulatory function in affected individuals may represent an effective treatment for asthma.

Topics: Animals; Asthma; Homeostasis; Humans; Immunity, Mucosal; Interleukin-10; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Asthma, chronic obstructive pulmonary disorder (COPD), and cystic fibrosis (CF), chronic diseases of the airways, are characterized by symptoms such as inflammation of the lung tissue, mucus hypersecretion, constriction of the airways, and excessive fibrosis of airway tissue. Transforming growth factor (TGF)-beta, a cytokine that affects many different cell processes, has an important role in the lungs of patients with some of these chronic airway diseases, especially with respect to airway remodeling. Eosinophils can be activated by and are a major source of TGF-beta in asthma. The action of TGF-beta also shows associations with other cell types, such as T cells and neutrophils, which are involved in the pathogenesis of asthma. TGF-beta can perpetuate the pathogenesis of COPD and CF, as well, through its induction of inflammation via release from and action on different cells. The intracellular signaling induced by TGF-beta in various cell types has been elucidated and may point to mechanisms of action by TGF-beta on different structural or immune cells in these airway diseases. Some possible treatments, especially that prevent the deleterious airway changes induced by the action of either eosinophils or TGF-beta in asthma, have been investigated. TGF-beta-induced signaling pathways, especially those in different cell types in asthma, COPD, or CF, may provide potential therapeutic targets for the treatment of some of the most devastating airway diseases.

Topics: Animals; Asthma; Cystic Fibrosis; Eosinophils; Humans; Neutrophils; Pulmonary Disease, Chronic Obstructive; T-Lymphocytes; Transforming Growth Factor beta

Airway remodeling is a characteristic feature of allergic asthma that is now thought to contribute to airway dysfunction and, ultimately, to clinical symptoms. A prevalent hypothesis holds that eosinophil-derived transforming growth factor-beta (TGF-beta) is a predominant underlying mechanism driving the development of remodeling and thus, represent promising targets for therapeutic intervention. This notion is supported by in vivo evidence from loss of function experiments conducted in animal models employing the surrogate allergen ovalbumin (OVA), and by indirect evidence from studies in human asthmatics. However, it is important to note that various studies in OVA systems have reported disconnects between eosinophils, TGF-beta and allergic remodeling. Moreover, recent investigations in a mouse model induced by respiratory exposure to a house dust mite extract have shown that remodeling can develop independently of TGF-beta. These findings challenge the above hypothesis and suggest that the mechanisms governing remodeling may be context specific. In addition to TGF-beta and eosinophils, several other factors have been implicated in the development of airway remodeling. Among these, interleukin (IL)-13 may be of particular importance given its role in type-2 immunity and in the tissue repair/fibrotic response. This review will appraise the evidence pertaining to the roles of TGF-beta, eosinophils and IL-13 in allergic remodeling, and will suggest that identifying robust targets for therapeutic intervention might benefit from a reconsideration of our approach to understanding remodeling.

Topics: Allergens; Animals; Anti-Allergic Agents; Asthma; Eosinophils; Humans; Interleukin-13; Respiratory Hypersensitivity; Transforming Growth Factor beta

Although histologic features of airway remodeling have been well characterized in asthma, the immunologic and inflammatory mechanisms that drive progression of asthma to remodeling are still incompletely understood. Conceptually, airway remodeling may be a result of persistent inflammation and/or aberrant tissue repair mechanisms. It is likely that several immune and inflammatory cell types and mediators are involved in mediating airway remodeling. In addition, different features of airway remodeling are likely mediated by different inflammatory pathways. Several important candidate mediators of remodeling have been identified, including TGF-beta and T(H)2 cytokines (including IL-5 and IL-13), as well as vascular endothelial growth factor, a disintegrin and metalloproteinase 33, and matrix metalloproteinase 9. Mouse models of airway remodeling have provided important insight into potential mechanisms by which TGF-beta activation of the Smad-2/3 signaling pathway may contribute to airway remodeling. Human studies have demonstrated that anti-IL-5 reduces levels of airway eosinophils expressing TGF-beta, as well as levels of airway remodeling as assessed by bronchial biopsies. Further such studies confirming these observations, as well as alternate studies targeting additional individual cell types, cytokines, and mediators, are needed in human subjects with asthma to determine the role of candidate mediators of inflammation on the development and progression of airway remodeling.

Topics: ADAM Proteins; Animals; Asthma; Disease Progression; Eosinophils; Humans; Inflammation; Lung; Matrix Metalloproteinase 9; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Both structural and inflammatory cells are capable of secreting transforming growth factor (TGF)-beta and expressing TGF-beta receptors. TGF-beta can induce multiple cellular responses including differentiation, apoptosis, survival and proliferation, and has been implicated in the development of several pathogenic conditions including cancer and asthma. Elevated levels of TGF-beta have been reported in the asthmatic airway. TGF-beta binds to its receptor complex and activates multiple pathways involving proteins such as Sma and Mad homologues, phosphatidylinositol-3 kinase and the mitogen-activated protein kinases, leading to the transcription of several genes. Cell type, cellular condition, and microenvironment, all play a role in determining which pathway is activated, which, in turn, is an indication of which gene is to be transcribed. TGF-beta has been shown to induce apoptosis in airway epithelial cells. A possible role for TGF-beta in the regulation of epithelial cell adhesion properties has also been reported. Enhancement of goblet cell proliferation by TGF-beta suggests a role in mucus hyper-secretion. Elevated levels of TGF-beta correlate with subepithelial fibrosis. TGF-beta induces proliferation of fibroblast cells and their differentiation into myofibroblasts and extracellular matrix (ECM) protein synthesis during the development of subepithelial fibrosis. TGF-beta also induces proliferation and survival of and ECM secretion in airway smooth muscle cells (ASMCs), suggesting a possible cause of increased thickness of airway tissues. TGF-beta also induces the production and release of vascular endothelial cell growth factor and plasminogen activator inhibitor, contributing to the vascular remodeling in the asthmatic airway. Blocking TGF-beta activity inhibits epithelial shedding, mucus hyper-secretion, angiogenesis, ASMC hypertrophy and hyperplasia in an asthmatic mouse model. Reduction of TGF-beta production and control of TGF-beta effects would be beneficial in the development of therapeutic intervention for airway remodeling in chronic asthma.

Topics: Animals; Asthma; Humans; Respiratory System; Signal Transduction; Transforming Growth Factor beta

Airway remodelling refers to changes in the airway structure and includes subepithelial fibrosis, increased smooth muscle mass, submucosal gland enlargement, neovascularisation and epithelial alterations. Remodelling is observed in response to chronic injury and is seen not only in asthma but in all airway diseases. Remodelling is associated with more severe airflow obstruction and airway hyperresponsiveness in asthma; however, the clinical significance of this is still a matter of debate. Research should be pursued to better understand the accurate implication of airway remodelling in disease and its therapeutic modulation. To allow research in this field, accurate and standardised methods should be utilised to measure airway alterations in disease and following therapy. The standard detection of structural alterations is through direct analyses of airway tissues obtained during a post mortem, surgically or by flexible bronchoscopy. To avoid invasive techniques, other tools have been developed to indirectly measure remodelling, including induced sputum, bronchoalveolar lavage fluid, blood and urine analyses, physiological and radiological assessments, as well as in vitro techniques. Although of great interest, the exact significance of airway remodelling measurements gained through such indirect techniques is uncertain and further research is needed. Despite their invasive nature, direct methods should be favoured to adequately measure airway remodelling in disease and its modulation by therapy.

Topics: Actins; Airway Resistance; Asthma; Biopsy; Bronchoscopy; Collagen Type III; Fibrosis; Humans; Matrix Metalloproteinases; Muscle, Smooth; Neovascularization, Pathologic; Pulmonary Disease, Chronic Obstructive; Research; Respiratory Mucosa; Transforming Growth Factor beta

IL-6 plays a pivotal role in immune responses and certain oncologic conditions. The intense investigation of its biological activity and function led to the discovery of two different IL-6-driven signalling pathways. Binding to the membrane-bound IL-6 receptor (mIL-6R, CD126) causes the recruitment of two gp130 co-receptor molecules (CD130) and the activation of intracellular signalling cascades via gp130. Although this classical pathway is mainly limited to hepatocytes, neutrophils, monocytes/macrophages and certain other leukocyte populations, which express IL-6R on their surface, an alternative mechanism has also been described. Proteolytic cleavage of the mIL-6R protein or translation from alternatively spliced mRNA leads to the generation of a soluble form of the IL-6R (sIL-6R), which is likewise able to bind to IL-6. The resulting IL-6/sIL-6R complex is also capable of binding to gp130 and inducing intracellular signalling. Through this so-called 'trans-signalling' mechanism, IL-6 is able to stimulate cells that lack an endogenous mIL-6R. High levels of IL-6 and sIL-6R have been reported in several chronic inflammatory and autoimmune diseases as well as in cancer. Preclinical animal disease models have provided strong evidence that specific blockade of IL-6-regulated signalling pathways represents a promising approach for the therapy of these diseases. An optimised variant of the recently described fusion protein sgp30Fc is now heading towards its clinical evaluation.

Topics: Alternative Splicing; Animals; Anti-Inflammatory Agents; Antineoplastic Agents; Arthritis; Arthritis, Rheumatoid; Asthma; Autoimmune Diseases; Cell Line; Clinical Trials as Topic; Colitis; Colonic Neoplasms; Cytokine Receptor gp130; Disease Models, Animal; Drug Delivery Systems; Drug Evaluation, Preclinical; Humans; Inflammation; Interleukin-6; Leukocytes; Male; Mice; Neoplasms; Receptors, Interleukin-6; Recombinant Fusion Proteins; Signal Transduction; Solubility; Transforming Growth Factor beta

Asthma is a chronic inflammatory disorder of the airways; therefore, establishing a simple monitoring system of airway inflammation would be useful for asthma management. Exhaled breath condensate (EBC), which is formed by breathing with a cooling system, is a non-invasive biomarker for airway inflammation. We have shown that nine kinds of molecules; namely, IL-4, IL-8, IL-17, TNF-alpha, RANTES, IP-10, TGF-beta, MIP-1alpha, and MIP-1beta, are over-expressed in asthmatic airways compared with those of healthy subjects using EBC analysis. In addition, the levels of some over-expressed molecules and lung physiologic indices are correlated. RANTES expression was significantly correlated with the degree of airflow limitation. Further, both TNF-alpha and TGF-beta values were significantly correlated with the degree of airway responsiveness and variability of peak expiratory flow. Additionally, the comparison of molecular properties between EBC and saliva indicated that saliva contamination is not a dominant contributor to the assessment of inflammatory molecules in EBC. Analysis of inflammatory molecules using EBC should be useful for monitoring the asthmatic airway condition, and might be a promising approach to assess the efficacy of pharmacologic interventions and to investigate the pathophysiology of asthma.

Topics: Asthma; Breath Tests; Chemokine CCL5; Cytokines; Humans; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Airway remodeling in asthma is defined by several structural changes including epithelial cell mucus metaplasia, an increase in peribronchial smooth muscle mass, subepithelial fibrosis, and angiogenesis. Cytokines, chemokines, and growth factors released from inflammatory and structural cells in the airway are considered to play a pivotal role in the development of remodeling. Studies of allergen induced airway remodeling in transgenic mice suggest an important role for TGF-beta, VEGF, Th2 cytokines (IL-5, IL-9, IL-13), and epithelial derived NF-kappaB regulated chemokines in airway remodeling. Although studies of bronchial biopsies from human asthmatics also demonstrate expression of TGF-beta, VEGF, IL-5, IL-9, IL-13, and NF-kappaB regulated chemokines, further human intervention studies are required in which individual cytokines or chemokines are neutralized to define their role in airway remodeling.

Topics: Animals; Asthma; CD4-Positive T-Lymphocytes; Chemokines; Eosinophils; Humans; Interleukins; Metalloproteases; Th2 Cells; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Asthma is increasing in prevalence in the developing world, affecting approximately 10% of the world's population. It is characterised by chronic lung inflammation and airway remodelling associated with wheezing, shortness of breath, acute bronchial hyperresponsiveness to a variety of innocuous stimuli and a more rapid decline in lung function over time. Airway remodelling, involving proliferation and differentiation of mesenchymal cells, particularly myofibroblasts and smooth muscle cells, is generally refractory to corticosteroids and makes a major contribution to disease chronicity. Transforming growth factor-beta is a potent profibrogenic factor whose expression is increased in the asthmatic airways and is a prime candidate for the initiation and persistence of airway remodelling in asthma. This review highlights the role of transforming growth factor-beta in the asthmatic lung, incorporating biosynthesis, signalling pathways and functional outcome. In vivo, however, it is the balance between transforming growth factor-beta and other growth factors, such as epidermal growth factor, which will determine the extent of fibrosis in the airways. A fuller comprehension of the actions of transforming growth factor-beta, and its interaction with other signalling pathways, such as the epidermal growth factor receptor signalling cascade, may enable development of therapies that control airway remodelling where there is an unmet clinical need.

Topics: Asthma; Chronic Disease; Epidermal Growth Factor; Epithelial Cells; Extracellular Matrix; Humans; Microscopy, Electron; Signal Transduction; Transforming Growth Factor beta

Allergic diseases are caused by the induction of T helper (Th)2 cells and IgE responses specific for common environmental antigens (allergens) in susceptible individuals. There is increasing interest in the role of both naturally occurring and induced regulatory T cell (Treg) populations in preventing these inappropriate immune responses and the underlying sensitisation to allergens. Current evidence suggests that Tregs may actively prevent Th2 responses to allergens occurring in healthy non-atopic individuals and that their function may be impaired in allergic patients. Evidence that existing therapies may act by modulating Treg function is reviewed. Future research aims to understand the mechanisms involved in the generation and function of allergen-specific Tregs. A primary aim is to promote the development of optimised therapeutic regimens with the capacity to provide long-lasting, allergen-specific, inhibitory mechanisms at the time and site of allergen challenge.

Topics: Allergens; Animals; Asthma; Humans; Hypersensitivity; Hypersensitivity, Immediate; Immunotherapy; Inflammation; Interleukin-10; Models, Biological; T-Lymphocytes; T-Lymphocytes, Regulatory; Th2 Cells; Transforming Growth Factor beta

Asthma is a chronic disease of the airways affecting around 10% of the population. The majority of cases are well controlled with current therapies, however in approximately 20% of severe asthmatics the available therapeutic strategies are inadequate. Structural changes in the asthmatic airway, including an increase in smooth muscle mass and an increased deposition of extracellular matrix proteins, which correlate with airway hyperresponsivenes, reduced lung function and an increase in fibroblast/myofibroblast numbers, are not specifically targetted by current therapeutic agents and therefore represent an area of unmet need. The mechanisms involved in the development of airway remodelling are incompletely understood but are thought to involve one or more isoforms of transforming growth factor-beta (TGF-beta). The TGF-betas are pleiotropic mediators which have important roles in the regulation of inflammation, cell growth, differentiation and wound healing. All three mammalian isoforms of TGF-beta are present in the airways and at least TGF-beta1 and TGF-beta2 have been shown to be increased in asthmatic airways and cells, together with evidence of increased TGF-beta signalling. In addition, evidence from animal models suggests that airway remodelling may be prevented or reversed using agents which target TGF-beta. Therefore modulation of TGF-betas or their activity represent a potential therapeutic target for asthma. This review focuses on the current knowledge of TGF-beta1-3, their their role in normal and asthmatic airways, as well as the potential for modulating the TGF-betas and their effects as a therapeutic approach to asthma.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Fibroblasts; Fibrosis; Humans; Myocytes, Smooth Muscle; Protein Isoforms; Receptors, Transforming Growth Factor beta; Signal Transduction; Transforming Growth Factor beta

Naturally occurring CD4+CD25+ regulatory T cells mediate immune suppression to limit immunopathogenesis associated with chronic inflammation, persistent infections and autoimmune diseases. Their mode of suppression is contact-dependent, antigen-nonspecific and involves a nonredundant contribution from the cytokine transforming growth factor (TGF)-beta. Not only can TGF-beta mediate cell-cell suppression between the regulatory T cells and CD4+CD25- or CD8+ T cells, but new evidence also reveals its role in the conversion of CD4+CD25- T cells, together with TCR antigen stimulation, into the regulatory phenotype. Elemental to this conversion process is induction of expression of the forkhead transcription factor, Foxp3. This context-dependent coercion of naive CD4+ T cells into a powerful subset of regulatory cells provides a window into potential manipulation of these cells to orchestrate therapeutic intervention in diseases characterized by inadequate suppression, as well as a promising means of controlling pathologic situations in which excessive suppression dominates.

Topics: Animals; Arthritis, Rheumatoid; Asthma; Autoimmune Diseases; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Clonal Anergy; Disease Models, Animal; Forkhead Transcription Factors; Gene Expression Regulation; Humans; Immune Tolerance; Immunotherapy, Adoptive; Inflammation; Lupus Erythematosus, Systemic; Mice; Mice, Knockout; Receptors, Interleukin-2; Signal Transduction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

1. Severe persistent asthma is accompanied by structural changes in the airway, referred to as remodelling. The mechanisms driving airway remodelling are poorly understood. 2. Transforming growth factor (TGF)-beta is increased in the airways of patients with asthma. Many of the effects of TGF-beta are mediated by connective tissue growth factor (CTGF). 3. Overexpression of CTGF is linked to many fibrotic diseases, but its exact role in airway remodelling is unknown. 4. Connective tissue growth factor mediates cell adhesion, migration, proliferation, survival, extracellular matrix synthesis and has a role in angiogenesis. 5. Current asthma therapies do not inhibit CTGF induction. 6. Understanding the mechanisms underlying the role of CTGF in airway remodelling may lead to new therapeutic strategies for asthma.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Connective Tissue Growth Factor; Extracellular Matrix; Extracellular Matrix Proteins; Gene Expression; Humans; Immediate-Early Proteins; Integrins; Intercellular Signaling Peptides and Proteins; Myocytes, Smooth Muscle; Receptors, Cell Surface; Signal Transduction; Transforming Growth Factor beta

Over the years, the role of the eosinophil in asthma and allergic processes has been disputed. Recent human experiments using a humanised monoclonal antibody to interleukin-5 (IL-5), and animal studies involving specific IL-5 gene deletion, indicates that eosinophils might control downstream repair and remodelling processes. Eosinophils are a rich source of fibrogenic factors, particularly transforming growth factor-beta (TGF-beta), the latent form of which is activated by epithelial-cell expression of the intergin alpha(v)beta(6). The emerging role for the eosinophil in airway remodelling might be important in future anti-asthma strategies. However, more effective eosinophil-depleting agents than anti-IL-5 are required before the definitive role of this cell type in asthma airway pathophysiology can be established.

Topics: Animals; Antibodies, Monoclonal; Asthma; Eosinophils; Humans; Interleukin-5; Lung; Signal Transduction; Transforming Growth Factor beta

Interleukin 5 (IL-5) is a key cytokine in the regulation of eosinophilia and eosinophil activation in humans. Monoclonal antibodies to anti-IL-5 have become available for use in clinical studies in humans. This article discusses the rationale for the use of anti-IL-5 therapy in asthma and hypereosinophilic syndrome and summarizes the available clinical data on the use of anti-IL-5 to treat these disorders.

Topics: Asthma; Eosinophils; Extracellular Matrix Proteins; Humans; Hypereosinophilic Syndrome; Interleukin-5; RNA, Messenger; Transforming Growth Factor beta; Transforming Growth Factor beta1

Topics: Airway Obstruction; Airway Resistance; Animals; Antibodies; Asthma; CpG Islands; Humans; Immunotherapy; Interleukin-5; Respiratory System; Transforming Growth Factor beta

Topics: Adjuvants, Immunologic; Asthma; Collagen Type I; Drug Synergism; Fetus; Fibroblasts; Gels; Humans; In Vitro Techniques; Interleukin-4; Lung; Lung Injury; Prenatal Injuries; Transforming Growth Factor beta; Transforming Growth Factor beta1; Wound Healing

Transforming growth factor-beta (TGF-beta) is an important fibrogenic and immunomodulatory factor that may play a role in the structural changes observed in the asthmatic airways. In vitro as well as in vivo studies have evidenced a dual role for TGF-beta: it can either function as a pro- or anti-inflammatory cytokine on inflammatory cells, participating into the initiation and resultion of inflammatory and immune responses in the airways. TGF-beta is also involved in the remodelling of the airway wall, and has in particular been related to the subepithelial fibrosis. TGF-beta is produced in the airways by inflammatory cells infiltrated in the bronchial mucosa, as well as by structural cells of the airway wall including fibroblasts, epithelial, endothelial and smooth muscle cells. By releasing TGF-beta, these different cell types may then participate into the increased levels of TGF-beta observed in bronchoalveolar lavage fluid from asthmatic patients. Taken together, these results suggest that TGF-beta may play a role in inflammation in asthma. However, as its role is dual in the modulation of inflammation, further studies are needed to elucidate the precise role of TGF-beta in the airways.

Topics: Animals; Asthma; Bronchi; Humans; Receptors, Transforming Growth Factor beta; Transforming Growth Factor beta

The development of asthmatic inflammation involves a complex array of cytokines that promote the recruitment and activation of a number of different immune cells. While factors involved in initiating and establishing inflammation are well characterized, the process by which this pro-inflammatory cascade is regulated is less well understood. The identification and characterization of immunomodulatory cytokines in asthma has been a difficult proposition. Many of the putative regulatory factors have pleiotropic bioactivities and have been characterized as pro-inflammatory in association with certain pathologic conditions. This chapter addresses the potential role of several endogenous factors which appear to attenuate asthmatic inflammation. Understanding the integration of these factors into the regulation of the inflammatory process will likely result in novel therapeutic approaches.

Topics: Animals; Asthma; Binding Sites; Cytokines; Humans; Inflammation; Interferon-gamma; Interleukin-10; Interleukin-16; Mice; Models, Biological; Surface-Active Agents; T-Lymphocytes; Transforming Growth Factor beta

A distinctive feature of asthma is the presence of a fibrotic response characterized by excess extracellular matrix deposition and proliferation of myofibroblasts, in the airway wall. This review discusses recent advances in the understanding of this phenomenon at the pathological and molecular levels, and its likely importance in the pathophysiology and symptomatology of the disease. Particular interest has centered on the potent fibrogenic molecule, transforming growth factor-beta (TGF-beta), and the evidence to link this molecule to airway fibrosis is discussed. Finally, the ability of existing anti-asthma treatments to influence the fibrotic response and the need to develop novel therapeutic strategies are considered.

Topics: Anti-Asthmatic Agents; Asthma; Bronchi; Fibroblasts; Fibrosis; Humans; Transforming Growth Factor beta

Asthma is a chronic inflammatory disease of the airways, with associated repair processes. Both inflammatory and repair processes appear to be strictly related, and can lead to several histopathological alterations of the bronchial mucosa, such as the shedding of epithelium and increased thickness of the basement membrane. The integrity as well as the alterations of the bronchial structure are the consequence of several biological events, such as cell proliferation and death, cell activation and inhibition, and extracellular matrix (ECM) production and degradation. These events are critically regulated by polypeptides called growth factors (GFs), which are able, functioning in an autocrine and paracrine fashion, to affect and modulate cell functions and ECM turnover. Although the importance of GFs has been widely demonstrated in other pulmonary conditions, such as lung fibrotic diseases, their possible involvement in the pathogenesis of inflammatory and postinflammatory processes in asthma is still not completely clear. The aim of the present review was to discuss the biological evidence concerning the role of several growth factors, such as transforming growth factor-beta (TGF-beta), epidermal growth factor (EGF), granulocyte/macrophage colony-stimulating factor (GM-CSF), platelet-derived growth factor (PDGF) and endothelin, in asthma and chronic bronchitis.

Topics: Asthma; Bronchi; Endothelial Growth Factors; Granulocyte-Macrophage Colony-Stimulating Factor; Growth Substances; Humans; Hyperplasia; Inflammation; Lung; Platelet-Derived Growth Factor; Respiratory System; Transforming Growth Factor beta

Asthma is known as one of the most common chronic inflammatory diseases characterized by recurrent obstruction and inflammation of the airways. Probiotics are defined as a group of beneficial living microorganisms that are beneficial in many disorders, including allergies. The aim of this study was to investigate the probiotic supplement effects on improvement of clinical asthma symptom and changes in the pattern of Th17-related inflammatory cytokines in asthmatic patients.. This was a randomized controlled clinical trial with parallel, double-blind groups. Forty patients with asthma were enrolled and received 1 capsule/day of a probiotic supplement for 8 weeks. Respiratory function tests; and the level of IL-6, IL-17, IL-21 and TGF-β were evaluated at the baseline and end of intervention.. The results showed that the level of IL-6 and IL-17 in patients after receiving probiotics was reduced and expression of TGF-β was increased as compared to the baseline. Also, the expression of IL-17 and IL-21 in the probiotic group was significantly lower than the placebo group at the end of the intervention. In addition, an improvement in pulmonary function tests and clinical symptoms was observed after receiving probiotics.. Eight-weeks treatment with a probiotic supplementation suggests that it may effect on Th17 cells-associated IL-6, IL-17 and TGF-β; and Forced Expiratory Volume in 1 s and Forced Vital Capacity. Taken together, these results suggest that probiotics may have the ability to affect neutrophilic asthma and they can possibly be used besides common treatments for patients with neutrophilic asthma.

Topics: Asthma; Cytokines; Double-Blind Method; Humans; Interleukin-17; Interleukin-6; Probiotics; Th17 Cells; Transforming Growth Factor beta

Airway inflammation characterized as asthma is one of the most common chronic diseases in the world. The aim of this study was to evaluate the possible effect of inspiratory muscle training on inflammation markers and oxidative stress levels in childhood asthma. A total of 105 children (age range 8-17 years), including 70 asthmatics and 35 healthy children, participated in the study. The 70 asthma patients were randomly assigned to the inspiratory muscle training (IMT) group (n = 35) and control group (n = 35), and healthy children were assigned to the healthy group (n = 35). The IMT group was treated with the threshold IMT device for 7 days/6 weeks at 30% of maximum inspiratory pressure. Respiratory muscle strength was evaluated with a mouth pressure measuring device, and respiratory function was evaluated with a spirometer. In addition, CRP, periostin, TGF-β, and oxidative stress levels were analyzed. The evaluation was performed only once in the healthy group and twice (at the beginning and end of 6 weeks) in asthma patients. In the study, there were significant differences between asthma patients and the healthy group in terms of MIP and MEP values, respiratory function, oxidative stress level, periostin, and TGF-β. Post-treatment, differences were observed in the oxidative stress level, periostin, and TGF-β of the IMT group (p < .05).. After 6 weeks of training, IMT positively contributed to reducing the inflammation level and oxidative stress. This suggests that IMT should be used as an alternative therapy to reduce inflammation and oxidative stress. (Trial Registration: The clinical trial protocol number is NCT05296707).. • It is known that adjunctive therapies given in addition to pharmacological treatment contribute to improving symptom control and quality of life in individuals with asthma.. • There are no studies about the effect of respiratory physiotherapy on biomarkers in asthmatic children. The sub-mechanism of improvement in individuals has not been elucidated. • In this context, inspiratory muscle training has a positive effect on inflammation and oxidative stress levels in children with asthma and IMT should be used as an alternative treatment for childhood asthma.

Topics: Adolescent; Asthma; Breathing Exercises; Child; Humans; Muscle Strength; Oxidative Stress; Quality of Life; Respiratory Muscles; Transforming Growth Factor beta

Topics: Adrenal Cortex Hormones; Asthma; CD4-Positive T-Lymphocytes; Cholecalciferol; Dendritic Cells; Gene Expression; Humans; Interferon-gamma; Interleukins; Monocytes; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Vitamin D Deficiency

Chronic obstructive pulmonary disease (COPD) and bronchial asthma (BA) are 2 severe respiratory disorders with different predominated immunopathologies. There are several "novel molecules" from different families that are proposed as part of the etiopathogenesis of COPD and BA. Proteinase-activated receptor 2 (PAR-2), thymic stromal lymphoprotein (TSLP), interleukin-4 and its receptor (CD124), Yin-Yang 1 (YY1), and transforming growth factor beta (TGF-β) have been previously shown to be involved in the pathophysiology of both these diseases. We investigated PAR-2, TSLP, CD124 (interleukin-4R), TGF-β, and YY1 immunohistochemical expression in endobronchial and transbronchial biopsies from 22 BA patients and 20 COPD patients. Immunostaining for the above-mentioned antigens was quantified using a modified semiquantitative scoring system and statistically evaluated. The values of TGF-β in the epithelial cells (P=0.0007) and TGF-β in the submucosa (P=0.0075) were higher in the BA samples, whereas values of CD124 (P=0.0015) and TSLP (P=0.0106) were higher in the COPD samples. No statistically significant differences between the groups were recorded for PAR-2 and YY1. Airway inflammatory reaction diversity in BA and COPD seems to be disease specific; however, there are also shared mechanisms involved in the pathophysiology of both diseases.

Topics: Adult; Aged; Aged, 80 and over; Asthma; Biopsy; Bronchi; Cytokines; Female; Gene Expression Regulation, Neoplastic; Humans; Interleukin-4 Receptor alpha Subunit; Male; Middle Aged; Pulmonary Disease, Chronic Obstructive; Receptor, PAR-2; Thymic Stromal Lymphopoietin; Transforming Growth Factor beta

Sublingual allergen-specific immunotherapy is gaining popularity for treatment of allergic diseases, but underlying immunological mechanisms are unresolved.. To perform detailed immunological investigation of sublingual house dust mite (HDM) immunotherapy.. A 12-month randomized double-blind placebo-controlled study of sublingual HDM immunotherapy in 30 HDM-allergic subjects was performed, with 1-year open extension in 9 patients on active treatment. HDM-stimulated blood mononuclear cells were analyzed for proliferation, cytokines, and regulatory T cells (Tregs) by flow cytometry and ELISA. Effects of blocking transforming growth factor (TGF)-beta and IL-10 on proliferation were determined. Treg suppressor function and allergen-specific antibody levels were measured. Clinical efficacy was assessed by symptom, medication, and Juniper quality-of-life scores.. Allergen-induced CD4(+) T-cell division and IL-5 production were significantly decreased after 6- and 12-months' active treatment but not placebo. sTGF-betaRII blocked immunotherapy-induced suppression of allergen-specific T-cell proliferation, maximal at 6 months. Decreased allergen-specific CD4(+) T-cell proliferation and increased IL-10 secretion and serum Der p 2-specific IgG(4) were maximal at 24 months' active treatment. Treg (CD4(+)CD25(+)CD127(lo)/Foxp3(+)) function was demonstrated by suppression of allergen-specific effector T-cell (CD4(+)CD25(-)CD127(hi)) proliferation and cytokine production. Clinical efficacy of immunotherapy was supported by significantly decreased rhinitis symptom score, total asthma score, and Juniper quality-of-life score.. This study establishes the novel finding that TGF-beta mediates the immunological suppression seen early in clinically effective sublingual HDM immunotherapy in addition to an increase in Tregs with suppressor function. Clinical trial registered with www.clinicaltrials.gov (NCT00250263).

Topics: Administration, Sublingual; Adolescent; Adult; Aged; Allergens; Animals; Asthma; Double-Blind Method; Female; Humans; Immunotherapy; Interleukin-5; Male; Middle Aged; Pyroglyphidae; Respiratory Hypersensitivity; Rhinitis, Allergic, Perennial; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

The possibility of additional strategies to enhance the effectiveness of specific immunotherapy (SIT) is highly attractive.. The aim of our study was to assess the influence of oral corticosteroids and oral corticosteroids combined with vitamin D(3) on the early clinical and immunological effects of SIT.. It was a randomized, double-blind, placebo-controlled trial conducted in 54 asthmatic children allergic to house dust mites. Intervention was based on receiving a single dose of oral steroid, with or without vitamin D(3), or placebo only on the day of the build-up phase of SIT.. After 12 months of SIT, the median daily inhaled corticosteroid (ICS) dose, which controls the symptoms of asthma, was reduced by 25% in the steroid group. However, a 50% reduction of the median daily ICS dose was observed in the control group. The clinical effects of SIT were not affected in the steroid+D(3) group. Concomitantly, we found that intervention with prednisone significantly impaired the induction of T regulatory lymphocytes. Importantly, the clinical and immunological effects of SIT were not affected by intervention with steroids administered with vitamin D(3).. Our study failed to show a beneficial effect of oral corticosteroids on allergen-specific immunotherapy. We observed that the combined administration of a corticosteroid drug and allergen extract suppressed the early clinical and immunological effects of SIT and that vitamin D(3) prevented this 'adverse' influence of steroids.

Topics: Adrenal Cortex Hormones; Animals; Antigens, Dermatophagoides; Arthropod Proteins; Asthma; Calcifediol; Child; Cholecalciferol; Combined Modality Therapy; Dermatophagoides farinae; Dermatophagoides pteronyssinus; Desensitization, Immunologic; Double-Blind Method; Female; Forced Expiratory Volume; Forkhead Transcription Factors; Gene Expression; Humans; Interleukin-10; Interleukin-13; Interleukin-5; Leukocytes, Mononuclear; Male; Prednisone; Steroids; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Treatment Outcome

High-altitude climate therapy is a well-established therapeutic option, which improves clinical symptoms in asthma. However, little is known about the underlying immunological mechanisms. The study investigates the influence of high-altitude climate therapy on airway inflammation and cellular components of specific and unspecific immune response. Exhaled NO significantly decreased within 3 weeks of therapy in patients with allergic and intrinsic, moderate and severe asthma. Interleukin-10 (IL-10)-secreting peripheral blood mononuclear cells (PBMC) increased within 3 weeks of therapy in six of 11 patients, whereas transforming growth factor-beta(1)-secreting PBMC remained stable. Furthermore, monocyte activation, assessed by CD80 expression significantly decreased during therapy. The frequency of CRTH2-expressing T cells decreased, while regulatory T cells (T(reg)) remained stable. FOXP3 and GATA-3 mRNA expression in CD4(+) T cells did not change, while interferon-gamma and IL-13 mRNA expression decreased in eight of 10 patients. The current data demonstrate that high-altitude climate therapy reduces local airway inflammation. Furthermore, monocytes switch towards a tolerogenic phenotype under high-altitude climate therapy. The T(reg)/Th2 ratio increases; however, because of the absence of antigens/allergens, no de novo differentiation of Th2 nor T(reg) cells is observed. The high-altitude climate therapy therefore may form the immunological basis for the endogenous control of allergen-driven diseases.

Topics: Adult; Altitude; Antigen-Presenting Cells; Asthma; Bronchitis; Climate; GATA3 Transcription Factor; Humans; Interferon-gamma; Interleukin-10; Interleukin-13; L-Selectin; Lymphocyte Activation; Middle Aged; Nitric Oxide; Receptors, Immunologic; Receptors, Prostaglandin; T-Lymphocytes, Regulatory; Transcription Factors; Transforming Growth Factor beta

We have shown previously that inflammation in asthma is not restricted to central airways but can also be demonstrated in peripheral airways. It is not clear whether inflammation of the peripheral airways is associated with structural changes and whether this remodeling process can be modulated by deposition of inhaled corticosteroids (ICSs).. To compare remodeling in peripheral and central airways and to investigate the effects of hydrofluoroalkane (HFA)-ICS on remodeling at these sites.. Transbronchial and endobronchial biopsies were obtained from 12 patients with mild to moderate asthma before and after a 6-week course of HFA-ICS (flunisolide). Total collagen deposition, expression of collagen III, TGF-beta, and alpha-smooth muscle actin were examined by using Van Gieson staining and immunocytochemistry, respectively.. Total collagen occupied 37.7% of the wall area of peripheral airways, compared with 54.5% of the wall area of central airways (P = .04). There was no significant difference in central versus peripheral airways for collagen III or alpha-smooth muscle actin immunoreactivity and in the number of TGF-beta(+) cells in the submucosa. The only significant effect of HFA-flunisolide was a decrease in alpha-smooth muscle actin area in peripheral airways (13.4% vs 4.6%; P = .01) that correlated with the percentage increase in forced expiratory flow at 25% to 75% of vital capacity (r(s) = -1.00; P = .00).. Our data show that there is a considerable degree of airway remodeling in peripheral airways in patients with asthma and confirm the inability of ICS to modulate collagen deposition and TGF-beta expression. Treatment with HFA-flunisolide is associated with a significant decrease in the expression of alpha-smooth muscle actin in peripheral airways, which correlated with improvement in peripheral airway function.

Topics: Actins; Adult; Anti-Asthmatic Agents; Asthma; Bronchi; Collagen; Drug Combinations; Female; Fluocinolone Acetonide; Humans; Male; Muscle, Smooth; Respiratory Function Tests; Severity of Illness Index; Transforming Growth Factor beta; Treatment Outcome

Important features of airway remodeling in asthma include the formation of subepithelial fibrosis and increased deposition of types I and III collagen. TGF-beta, IL-11, and IL-17 are profibrotic cytokines involved in the formation of subepithelial fibrosis and are increased in patients with asthma, particularly in those with severe disease.. The purpose of this study was to investigate the effect of corticosteroids on the expression of these profibrotic cytokines and on extracellular matrix deposition.. We used immunocytochemistry to measure the expression of TGF-beta, IL-11, IL-17, and collagen types I and III in the airways of patients with mild asthma (n = 9), patients with moderate-to-severe asthma (n = 10), and control subjects without asthma (n = 6). Baseline bronchial biopsy specimens were obtained in all groups. In addition, repeat biopsies were obtained in the patients with moderate-to-severe asthma after a 2-week course of oral corticosteroids.. TGF-beta expression was significantly higher in all groups with asthma, and it did not decrease after treatment with oral corticosteroids. Levels of IL-11 and IL-17 were increased in patients with moderate-to-severe asthma compared with patients with mild asthma and normal controls (P <.05). The expression of these cytokines decreased with oral corticosteroids in the moderate-to-severe group to levels that were comparable to those seen in the patients with mild asthma and in the normal controls (P <.005). Expression of types I and III collagens was higher in the patients with moderate-to-severe asthma than in the patients with mild asthma and the controls (P <.05; P <.001). Treatment with corticosteroids did not decrease the expression of types I and III collagens.. These results confirm the association of increased levels of TGF-beta, IL-11, IL-17, and types I and III collagens with severe disease and suggest that the failure of cortico-steroids to decrease collagen deposition might be due to persistently elevated TGF-beta expression.

Topics: Administration, Oral; Adult; Asthma; Bronchi; Collagen Type I; Collagen Type III; Cytokines; Female; Fibrillar Collagens; Fibrosis; Glucocorticoids; Humans; Immunohistochemistry; Interleukin-11; Interleukin-17; Male; Methylprednisolone; Transforming Growth Factor beta

Eosinophil-derived TGF-beta has been implicated in remodeling events in asthma. We hypothesized that reduction of bronchial mucosal eosinophils with anti-IL-5 would reduce markers of airway remodeling. Bronchial biopsies were obtained before and after three infusions of a humanized, anti-IL-5 monoclonal antibody (mepolizumab) in 24 atopic asthmatics in a randomized, double-blind, placebo-controlled study. The thickness and density of tenascin, lumican, and procollagen III in the reticular basement membrane (RBM) were quantified immunohistochemically by confocal microscopy. Expression of TGF-beta1 mRNA by airway eosinophils was assessed by in situ hybridization, and TGF-beta1 protein was measured in bronchoalveolar lavage (BAL) fluid by ELISA. At baseline, airway eosinophil infiltration and ECM protein deposition was increased in the RBM of asthmatics compared with nonasthmatic controls. Treating asthmatics with anti-IL-5 antibody, which specifically decreased airway eosinophil numbers, significantly reduced the expression of tenascin, lumican, and procollagen III in the bronchial mucosal RBM when compared with placebo. In addition, anti-IL-5 treatment was associated with a significant reduction in the numbers and percentage of airway eosinophils expressing mRNA for TGF-beta1 and the concentration of TGF-beta1 in BAL fluid. Therefore eosinophils may contribute to tissue remodeling processes in asthma by regulating the deposition of ECM proteins.

Topics: Antibodies, Monoclonal; Antibodies, Monoclonal, Humanized; Asthma; Basement Membrane; Bronchi; Chondroitin Sulfate Proteoglycans; Collagen Type III; Double-Blind Method; Eosinophils; Extracellular Matrix Proteins; Humans; Immunohistochemistry; Interleukin-5; Keratan Sulfate; Lumican; RNA, Messenger; Tenascin; Transforming Growth Factor beta; Transforming Growth Factor beta1

Topics: Antigens, CD; Asthma; Dendritic Cells; GPI-Linked Proteins; Humans; Hypersensitivity; Inflammation; Neoplasm Proteins; Transforming Growth Factor beta

Topics: Asthma; Child; Humans; Integrin beta4; Mycoplasma pneumoniae; Pneumonia, Mycoplasma; Transforming Growth Factor beta

Transforming growth factor-beta TGF-β-induced epithelial-mesenchymal transition (EMT) in bronchial epithelial cells contributes to airway wall remodeling in asthma. This study aims to explore the role of amygdalin, an active ingredient in bitter almonds, in TGF-β-induced EMT in bronchial epithelial cells and to elucidate the possible mechanisms underlying its biological effects.. An asthmatic mouse model was established through ovalbumin induction. Primary mouse bronchial epithelial cells and a human bronchial epithelial cell line were incubated with transforming growth factor-beta (TGF-β) to induce EMT, whose phenotype of cells was evaluated by the expressions of EMT markers [alpha-smooth muscle actin (α-SMA), vimentin, and fibronectin] and cell migration capacity. A co-immunoprecipitation assay was performed to assess the ubiquitination of heparanase (HPSE).. In asthmatic model mice, amygdalin treatment relieved airway wall remodeling and decreased expressions of EMT markers (α-SMA and vimentin). In TGF-β-treated bronchial epithelial cells, amygdalin treatment decreased the mRNA and protein levels of EMT markers (α-SMA, vimentin, and fibronectin) without impairing cell viability. Through the Swiss Target Prediction database, HPSE was screened as a candidate downstream target for amygdalin. HPSE overexpression further promoted TGF-β-induced EMT while the HPSE inhibitor suppressed TGF-β-induced EMT in bronchial epithelial cells. In addition, HPSE overexpression reversed the inhibitory effect of amygdalin on TGF-β-induced EMT in bronchial epithelial cells. The following mechanism exploration revealed that amygdalin downregulated HPSE expression by enhancing ubiquitination.. Our study showed that amygdalin inhibited TGF-β-induced EMT in bronchial epithelial cells and found that the anti-EMT activity of amygdalin might be related to its regulatory effect on HPSE expression.

Topics: Amygdalin; Animals; Asthma; Epithelial Cells; Epithelial-Mesenchymal Transition; Fibronectins; Humans; Mice; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transforming Growth Factors; Vimentin

Asthma is a chronic airway disease. Allergic reactions and T helper (h)2 immune response play a key role in asthma occurrence. Cell therapy can control inflammation and remodeling responses in allergic asthma, and cytokines can change this effect. Therefore, in this study, the effect of treated cell therapy with IL-2 to control allergic asthma was studied. Bone marrow cells were extracted and co-cultured with IL-2 and the cells were used via intra-tracheal administration in allergic asthma mice. Levels of IL-4, IL-5, IL-13, Leukotriene B4 and C4, and remodeling factors were measured. At least, a histopathology test of lung tissue was done. Type2 cytokines, leukotrienes, remodeling factors, mucus secretion, goblet cell hyperplasia, peri-bronchial and peri-vascular inflammation were significantly (p˂0.05) decreased by treating with bone marrow-derived mononuclear cells (BMDMCs) and IL-2-BMDMCs. Treatment with IL-2-BMDMCs could significantly decrease IL-13, transforming growth factor (TGF)-β, HP levels, and mucus secretion (p˂0.05) compared to BMDMCs treatment. In this study, BMDMCs and IL-2-BMDMCs therapy could decrease inflammation, allergic, and remodeling factors in allergic asthma. Cell therapy with BMDMCs had a strong and notable effect on the control of allergic asthma pathophysiology when co-cultured and used with IL-2.

Topics: Animals; Asthma; Bone Marrow; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Hypersensitivity; Inflammation; Interleukin-13; Interleukin-2; Leukocytes, Mononuclear; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Transforming Growth Factor beta

Allergic asthma is a chronic inflammatory illness of the respiratory system characterized by an increase in the number of inflammatory cells in the airways and trouble breathing. Mesenchymal stem cells (MSCs) have the potential to be used in inflammatory diseases as a cellular immunosuppressive treatment. They express calcitriol receptors and communicate with other immunocytes, which increases their anti-inflammatory activity. This study aimed to determine the effects of calcitriol-treated MSC treatment on allergic asthma pathways in a mouse model.. To generate a mouse model of asthma, the mice were sensitized intraperitoneally with ovalbumin (OVA) and aluminum hydroxide emulsion and then challenged intra-nasally with OVA. On day 14, experimental mice received tail vein injections of calcitriol-treated MSCs in PBS prior to allergen exposure. The cytokines assays including IL-4, 10, 12, 17, TGF-β and IFN-γ, splenocytes proliferation, and histological examination of lungs samples were performed. The mice were sensitized with OVA and the response to dexamethasone treatment was compared.. Calcitriol-treated MSCs significantly increased the levels of IL-12, TGF-β, and IFN-γ compared to non-treated MSCs groups. Moreover, calcitriol-treated and non-treated MSCs significantly decreased IL-4 and IL-17 compared to asthmatic groups. The results of the histopathological examination showed that calcitriol-treated MSCs reduced the accumulation of inflammatory cells and bronchial wall thickening in comparison with the asthma group.. Using the allergic asthma model, we were able to show that calcitriol-treated MSCs had an inhibitory impact on airway inflammation. Our findings suggest that the injection of calcitrioltreated MSCs may be a viable treatment option for allergic asthma.

Topics: Animals; Asthma; Calcitriol; Cytokines; Disease Models, Animal; Immunomodulation; Interleukin-4; Lung; Mesenchymal Stem Cells; Mice; Ovalbumin; Transforming Growth Factor beta

Asthma is a complex respiratory disease, which is controlled by genetic and environmental factors. Type 2-dominant immune response is responsible for asthma. Decorin (Dcn) and stem cells have modulatory effect on immune system and may control tissue remodeling and asthma pathophysiology. In this study, immunomodulatory effect of transduced induced pluripotent stem cells (iPSCs) with expression of

Topics: Animals; Asthma; Cytokines; Decorin; Disease Models, Animal; Immunoglobulin E; Induced Pluripotent Stem Cells; Interleukin-13; Interleukin-33; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Transforming Growth Factor beta

The airway wall remodeling observed in asthma is associated with subepithelial fibrosis and enhanced activation of human bronchial fibroblasts (HBFs) in the fibroblast to myofibroblast transition (FMT), induced mainly by transforming growth factor-β (TGF-β). The relationships between asthma severity, obesity, and hyperlipidemia suggest the involvement of peroxisome proliferator-activated receptors (PPARs) in the remodeling of asthmatic bronchi. In this study, we investigated the effect of PPARδ ligands (GW501516 as an agonist, and GSK0660 as an antagonist) on the FMT potential of HBFs derived from asthmatic patients cultured in vitro. This report shows, for the first time, the inhibitory effect of a PPARδ agonist on the number of myofibroblasts and the expression of myofibroblast-related markers-α-smooth muscle actin, collagen 1, tenascin C, and connexin 43-in asthma-related TGF-β-treated HBF populations. We suggest that actin cytoskeleton reorganization and Smad2 transcriptional activity altered by GW501516 lead to the attenuation of the FMT in HBF populations derived from asthmatics. In conclusion, our data demonstrate that a PPARδ agonist stimulates antifibrotic effects in an in vitro model of bronchial subepithelial fibrosis. This suggests its potential role in the development of a possible novel therapeutic approach for the treatment of subepithelial fibrosis during asthma.

Topics: Asthma; Bronchi; Cells, Cultured; Fibroblasts; Fibrosis; Humans; Myofibroblasts; PPAR delta; Transforming Growth Factor beta; Transforming Growth Factor beta1

The primary underlying defect in cystic fibrosis (CF) is disrupted ion transport in epithelia throughout the body. It is unclear if symptoms such as airway hyperreactivity (AHR) and increased airway smooth muscle (ASM) volume in people with CF are due to inherent abnormalities in smooth muscle or are secondary to epithelial dysfunction. Transforming Growth Factor beta 1 (TGFβ) is an established genetic modifier of CF lung disease and a known driver of abnormal ASM function. Prior studies have demonstrated that CF mice develop greater AHR, goblet cell hyperplasia, and ASM hypertrophy after pulmonary TGFβ exposure. However, the mechanism driving these abnormalities in CF lung disease, specifically the contribution of CFTR loss in ASM, was unknown.. In this study, mice with smooth muscle-specific loss of CFTR function (Cftr. Cftr. These results demonstrate a direct smooth muscle contribution to CF airway obstruction mediated by TGFβ. Dysfunction in non-epithelial tissues should be considered in the development of CF therapeutics, including potential genetic therapies.

Topics: Animals; Asthma; Cystic Fibrosis; Cystic Fibrosis Transmembrane Conductance Regulator; Hyperplasia; Mice; Muscle, Smooth; Transforming Growth Factor beta

Dual-specificity phosphatase 8 (DUSP8) is a MAPK phosphatase that dephosphorylates and inactivates the kinase JNK. DUSP8 is highly expressed in T cells; however, the in vivo role of DUSP8 in T cells remains unclear. Using T cell-specific Dusp8 conditional KO (T-Dusp8 cKO) mice, mass spectrometry analysis, ChIP-Seq, and immune analysis, we found that DUSP8 interacted with Pur-α, stimulated interleukin-9 (IL-9) gene expression, and promoted Th9 differentiation. Mechanistically, DUSP8 dephosphorylated the transcriptional repressor Pur-α upon TGF-β signaling, leading to the nuclear export of Pur-α and subsequent IL-9 transcriptional activation. Furthermore, Il-9 mRNA levels were induced in Pur-α-deficient T cells. In addition, T-Dusp8-cKO mice displayed reduction of IL-9 and Th9-mediated immune responses in the allergic asthma model. Reduction of Il-9 mRNA levels in T cells and allergic responses of T-Dusp8-cKO mice was reversed by Pur-α knockout. Remarkably, DUSP8 protein levels and the DUSP8-Pur-α interaction were indeed increased in the cytoplasm of T cells from people with asthma and patients with atopic dermatitis. Collectively, DUSP8 induces TGF-β-stimulated IL-9 transcription and Th9-induced allergic responses by inhibiting the nuclear translocation of the transcriptional repressor Pur-α. DUSP8 may be a T-cell biomarker and therapeutic target for asthma and atopic dermatitis.

Topics: Active Transport, Cell Nucleus; Animals; Asthma; Dermatitis, Atopic; Dual-Specificity Phosphatases; Humans; Hypersensitivity; Inflammation; Interleukin-9; Mice; RNA, Messenger; Transcription Factors; Transforming Growth Factor beta

Childhood asthma is the most universal chronic disease, with significant cases reported. Despite the current progress in treatment, prognosis remains poor and the existing drugs cause serious side effects. This investigation explored the mechanisms and use of miR-335-5p on childhood asthma therapy. MiR-335-5p and ATG5 expression was analyzed in clinical plasma samples through RT-qPCR. Airway smooth muscle cells (ASMCs) were cultured, and transfected with miR-335-5p mimic, miR-335-5p inhibitor, and pcDNA3.1-ATG5, or co-transfected with miR-335-5p mimic + pcDNA3.1-ATG5. Asthma cell models were constructed through TGF-β1, and animal models through ovalbumin (OVA). Monocyte-macrophage infiltration in bronchoalveolar lavage fluid (BALF) was determined by May-Grunwald-Giemsa staining, and collagen in lung tissue was assessed via Masson staining. Relationship between miR-335-5p and ATG5 was detected by dual-luciferase assay. Cell proliferation was detected by MTT assay. MiR-335-5p and ATG5 RNA expression was determined by RT-qPCR. Collagen I, collagen III, α-SMA, ATG5, LC3I/II, Beclin-1, and p62 protein expression levels in ASMCs were detected by western blot. MiR-335-5p expression was low, but ATG5 expression was high in childhood asthma. Versus OVA+ mimic NC group, the number of eosinophil and collagen in OVA+ miR-335-5p mimic group were reduced. In contrast to TGF-β1 + mimic NC group, TGF-β1 + miR-335-5p mimic group reduced inflammatory, airway fibrosis, and autophagy in ASMCs. ATG5 was miR-335-5p target. Overexpressing ATG5 significantly reversed the inhibitory effects of miR-335-5p on inflammatory response, fibrosis, and autophagy in ASMCs. Overall, the study concludes that MiR-335-5p alleviate inflammatory response, airway fibrosis, and autophagy in childhood asthma through targeted regulation of ATG5.

Topics: Animals; Asthma; Autophagy; Autophagy-Related Protein 5; Bronchoalveolar Lavage Fluid; Cell Proliferation; Cells, Cultured; Child; Disease Models, Animal; Female; Humans; Male; MicroRNAs; Myocytes, Smooth Muscle; Ovalbumin; Respiratory System; Signal Transduction; Transforming Growth Factor beta

Airway remodelling is a critical feature of chronic lung diseases. Epithelial-mesenchymal transition (EMT) represents an important source of myofibroblasts, contributing to airway remodelling. Here, we investigated the sphingosine-1-phosphate (S1P) role in EMT and its involvement in asthma-related airway dysfunction.. A549 cells were used to assess the S1P effect on EMT and its interaction with TGF-β signalling. To assess the S1P role in vivo and its impact on lung function, two experimental models of asthma were used by exposing BALB/c mice to subcutaneous administration of either S1P or ovalbumin (OVA).. Following incubation with TGF-β or S1P, A549 acquire a fibroblast-like morphology associated with an increase of mesenchymal markers and down-regulation of the epithelial. These effects are reversed by treatment with the TGF-β receptor antagonist LY2109761. Systemic administration of S1P to BALB/c mice induces asthma-like disease characterized by mucous cell metaplasia and increased levels of TGF-β, IL-33 and FGF-2 within the lung. The bronchi harvested from S1P-treated mice display bronchial hyperresponsiveness associated with overexpression of the mesenchymal and fibrosis markers and reduction of the epithelial.The S1P-induced switch from the epithelial toward the mesenchymal pattern correlates to a significant increase of lung resistance and fibroblast activation. TGF-β blockade, in S1P-treated mice, abrogates these effects. Finally, inhibition of sphingosine kinases by SK1-II in OVA-sensitized mice, abrogates EMT, pulmonary TGF-β up-regulation, fibroblasts recruitment and airway hyperresponsiveness.. Targeting S1P/TGF-β axis may hold promise as a feasible therapeutic target to control airway dysfunction in asthma.

Topics: Airway Remodeling; Animals; Asthma; Epithelial Cells; Epithelial-Mesenchymal Transition; Lysophospholipids; Mice; Mice, Inbred BALB C; Ovalbumin; Sphingosine; Transforming Growth Factor beta; Transforming Growth Factor beta1

Airway smooth muscle (ASM) cells are fundamental to asthma pathogenesis, influencing bronchoconstriction, airway hyperresponsiveness and airway remodelling. The extracellular matrix (ECM) can influence tissue remodelling pathways; however, to date no study has investigated the effect of ASM ECM stiffness and cross-linking on the development of asthmatic airway remodelling. We hypothesised that transforming growth factor-β (TGF-β) activation by ASM cells is influenced by ECM in asthma and sought to investigate the mechanisms involved.. This study combines. We found that asthmatic ASM cells activated more TGF-β basally than nonasthmatic controls and that diseased cell-derived ECM influences levels of TGF-β activated. Our data demonstrate that the ECM cross-linking enzyme LOXL2 is increased in asthmatic ASM cells and in bronchial biopsies. Crucially, we show that LOXL2 inhibition reduces ECM stiffness and TGF-β activation. These data are the first to highlight a role for LOXL2 in the development of asthmatic airway remodelling and suggest that LOXL2 inhibition warrants further investigation as a potential therapy to reduce remodelling of the airways in severe asthma.

Topics: Airway Remodeling; Amino Acid Oxidoreductases; Animals; Asthma; Mice; Muscle, Smooth; Protein-Lysine 6-Oxidase; Transforming Growth Factor beta

Obesity is a common comorbid condition in adult asthmatics and known as a feature of asthma severity. However, the molecular mechanism under obesity-induced inflammation has not yet been fully understood.. Considering the essential role of hydrophobic surfactant protein B (SP-B) in lung function, SP-B was targeted to examine its involvement in the development of obesity-induced airway inflammation in asthmatics.. The aim was to examine an alteration in circulating SP-B according to obesity in adult asthmatics, 129 asthmatics were enrolled and classified into 3 groups (obese, overweight and normal-weight groups) according to body mass index (BMI). Circulating SP-B levels were determined by enzyme-linked immunosorbent assay. Four single nucleotide polymorphisms of SFTPB gene were genotyped. Serum ceramide levels were measured by liquid chromatography-tandem mass spectrometry.. Significantly lower serum SP-B levels were noted in the obese group than in the overweight or normal-weight group (p = .002). The serum SP-B level was significantly correlated with serum levels of C18:0 ceramide and transforming growth factor beta 1 as well as BMI (r = -0.200; r = -0.215; r = -0.332, p < .050 for all). An inverse correlation was noted between serum SP-B and fractional exhaled nitric oxide levels in female asthmatics (r = -0.287, p = .009). Genetic predisposition of the SFTPB gene at 9306 A>G to the obese and overweight groups was noted.. Obesity altered ceramide metabolism leading to pulmonary surfactant dysfunction and impaired resolution of airway inflammation, finally contributing to the phenotypes of obese asthmatics.

Topics: Asthma; Ceramides; Female; Humans; Inflammation; Obesity; Overweight; Surface-Active Agents; Transforming Growth Factor beta

Airway inflammation and airway hyperresponsiveness (AHR) are pivotal characteristics of equine asthma. Lipopolysaccharide (LPS) may have a central role in modulating airway inflammation and dysfunction. Therefore, the aim of this study was to match the inflammatory and contractile profile in LPS-challenged equine isolated bronchi to identify molecular targets potentially suitable to counteract AHR in asthmatic horses.. Equine isolated bronchi were incubated overnight with LPS (0.1-100 ng/ml). The contractile response to electrical field stimulation (EFS) and the levels of cytokines, chemokines, and neurokinin A (NKA) were quantified. The role of capsaicin sensitive-sensory nerves, neurokinin-2 (NK. LPS 1 ng/ml elicited AHR to EFS (+238.17 ± 25.20% P < 0.001 vs. control). LPS significantly (P < 0.05 vs. control) increased the levels of IL-4 (+36.08 ± 1.62%), IL-5 (+38.60 ± 3.58%), IL-6 (+33.79 ± 2.59%), IL-13 (+40.91 ± 1.93%), IL-1β (+1650.16 ± 71.16%), IL-33 (+88.14 ± 8.93%), TGF-β (22.29 ± 1.03%), TNF-α (+56.13 ± 4.61%), CXCL-8 (+98.49 ± 17.70%), EOTAXIN (+32.26 ± 2.27%), MCP-1 (+49.63 ± 4.59%), RANTES (+36.38 ± 2.24%), and NKA (+112.81 ± 6.42%). Capsaicin sensitive-sensory nerves, NK. Targeting IL-6 with specific antibody may represent an effective strategy to treat equine asthma, especially in those animals suffering from severe forms of this disease.

Topics: Animals; Asthma; Bronchi; Capsaicin; Horses; Inflammation; Interleukin-33; Interleukin-6; Lipopolysaccharides; Neurokinin A; Transforming Growth Factor beta

The impaired production of extracellular matrix (ECM) proteins by airway smooth muscle cells (ASMC) and pulmonary fibroblasts (PF) is a part of airway remodeling in asthma. This process might be influenced by eosinophils that migrate to the airway and abundantly secrete various cytokines, including TGF-β. We aimed to investigate the effect of asthmatic eosinophils on the gene expression of ECM proteins in ASMC and PF. A total of 34 study subjects were recruited: 14 with allergic asthma (AA), 9 with severe non-allergic eosinophilic asthma (SNEA), and 11 healthy subjects (HS). All AA patients underwent bronchial allergen challenge with

Topics: ADAM Proteins; Allergens; Animals; Asthma; Collagen; Decorin; Dermatophagoides pteronyssinus; Eosinophils; Extracellular Matrix; Extracellular Matrix Proteins; Fibroblasts; Fibronectins; Gene Expression; Humans; Lung; Myocytes, Smooth Muscle; Tissue Inhibitor of Metalloproteinase-2; Transforming Growth Factor beta

Asthma is characterized by persistent inflammation, airway hypersensitivity and remodelling. Bone Morphogenetic Proteins belong to the Transforming Growth Factor Superfamily and have a similar signalling transduction pathway and common co-mediating protein. However, the BMPs role in the remodelling remains unclear; they appear to be involved in the airway inflammation and fibrogenesis process.. 60 patients with asthma and 48 healthy volunteers were recruited for the study. Blood samples were collected before, 1 hour, 24 and 48 hours after the allergen or the methacholine challenge test. Evaluation of BMP-4 and BMP-7 serum concentration and expression was performed using ELISA and real time PCR methods, respectively.. Statistically significant differences in BMP-7 concentration between healthy controls and asthmatics before the chal-lenge were noted. We found two statistically significant correlations: between the basal BMP-4 concentration and the FEV1(L) raw value and FEV1/FVC(%) index. We did not observe significant changes in the gene expression of BMP-4 and BMP-7 in different time points.. Observed differences in BMP-7 concentration between asthmatic and healthy groups and correlations between BMP-4 concentration and some lung function test values may indicate the role of the BMPs in the etiopathogenesis of asthma. The unique characteristic of our study is the evaluation of BMPs serum levels, not in the bronchial epithelium.

Topics: Asthma; Bone Morphogenetic Protein 4; Bone Morphogenetic Protein 7; Bone Morphogenetic Proteins; Humans; Inflammation; Smad Proteins; Transforming Growth Factor beta

Type 2-high asthma is a chronic inflammatory disease of the airways which is increasingly prevalent in countries where helminth parasite infections are rare, and characterized by T helper 2 (Th2)-dependent accumulation of eosinophils in the lungs. Regulatory cytokines such as TGF-β can restrain inflammatory reactions, dampen allergic Th2 responses, and control eosinophil activation. The murine helminth parasite Heligmosomoides polygyrus releases a TGF-β mimic (Hp-TGM) that replicates the biological and functional properties of TGF-β despite bearing no structural similarity to the mammalian protein. Here, we investigated if Hp-TGM could alleviate allergic airway inflammation in mice exposed to Alternaria alternata allergen, house dust mite (HDM) extract or alum-adjuvanted ovalbumin protein (OVA). Intranasal administration of Hp-TGM during Alternaria exposure sharply reduced airway and lung tissue eosinophilia along with bronchoalveolar lavage fluid IL-5 and lung IL-33 cytokine levels at 24 h. The protective effect of Hp-TGM on airway eosinophilia was also obtained in the longer T-cell mediated models of HDM or OVA sensitisation with significant inhibition of eotaxin-1, IL-4 and IL-13 responses depending on the model and time-point. Hp-TGM was also protective when administered parenterally either when given at the time of allergic sensitisation or during airway allergen challenge. This project has taken the first steps in identifying the role of Hp-TGM in allergic asthma and highlighted its ability to control lung inflammation and allergic pathology. Future research will investigate the mode of action of Hp-TGM against airway allergic eosinophilia, and further explore its potential to be developed as a biotherapeutic in allergic asthma.

Topics: Allergens; Animals; Asthma; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Cytokines; Eosinophilia; Helminths; Interleukin-13; Interleukin-33; Interleukin-4; Interleukin-5; Lung; Mammals; Mice; Mice, Inbred BALB C; Ovalbumin; Transforming Growth Factor beta

Airway fibrosis is one of the pathological characteristics of severe asthma. Transforming growth factor (TGF)-β has been known to promote epithelial-mesenchymal transition formation and to play a role in the progression of tissue fibrosis. Cellular communication network factor 2 (CCN2) and fibronectin (FN) are well-known markers of EMT and fibrosis. However, whether AREG is involved in TGF-β-induced CCN2 and FN expression in human lung epithelial cells is unknown.. AREG and FN were analyzed by immunofluorescence staining on ovalbumin-challenged mice. CCN2 and FN expression were evaluated in human lung epithelial (A459) cells following TGF or AREG treatment for the indicated times. Secreted AREG from A549 cells was detected by ELISA. Cell migration was observed by a wound healing assay. Chromatin immunoprecipitation was used to detect the c-Jun binding to the CCN2 promoter.. AREG and FN expression colocalized in lung tissues from mice with ovalbumin-induced asthma by immunofluorescence staining. Moreover, TGF-β caused the release of AREG from A549 cells into the medium. Smad3 siRNA down-regulated AREG expression. AREG also stimulated CCN2 and FN expression, JNK and c-Jun phosphorylation, and cell migration in A549 cells. AREG small interfering (si) RNA inhibited TGF-β-induced expression of CCN2, FN, and cell migration. Furthermore, AREG-induced CCN2 and FN expression were inhibited by EGFR siRNA, a JNK inhibitor (SP600125), and an activator protein-1 (AP-1) inhibitor (curcumin). EGFR siRNA attenuated AREG-induced JNK and c-Jun phosphorylation. Moreover, SP600125 downregulated AREG-induced c-Jun phosphorylation.. These results suggested that AREG mediates the TGF-β-induced EMT in human lung epithelial cells through EGFR/JNK/AP-1 activation. Understanding the role of AREG in the EMT could foster the development of therapeutic strategies for airway remodeling in severe asthma.

Topics: Amphiregulin; Animals; Asthma; Epithelial Cells; Epithelial-Mesenchymal Transition; ErbB Receptors; Fibronectins; Fibrosis; Humans; Lung; Mice; Ovalbumin; RNA, Small Interfering; Transcription Factor AP-1; Transforming Growth Factor beta; Transforming Growth Factor beta1

Asthma is a chronic airway disease whose exacerbations are often triggered by rhinovirus infection. TGF-β1 induces rhinovirus replication in infected cells. Moreover, TGF-β1 is a pleiotropic mediator that is produced by many immune cells in the latent, inactive form bound to the latency-associated peptide (LAP) and to the transmembrane protein glycoprotein A repetitions predominant (GARP). In this study we wanted to investigate the effect of rhinovirus infection on the TGF-β secretion and the downstream signaling via TGF-βRI/RII in peripheral blood mononuclear cells from control and asthmatic patients after rhinovirus infection ex vivo. Here, we found a significant upregulation of TGF-βRII in untouched PBMCs of asthmatics as well as a suppression of TGF-β release in the rhinovirus-infected PBMC condition. Moreover, consistent with an effect of TGF-β on Tregs, PBMCs infected with RV induced Tregs, and TGF-βRII directly correlated with RV1b mRNA. Finally, we found via flow cytometry that NK cells expressed less GARP surface-bound TGF-β, while cytokine-producing NK

Topics: Asthma; Enterovirus Infections; Glycoproteins; Humans; Killer Cells, Natural; Leukocytes, Mononuclear; Membrane Proteins; Rhinovirus; Transforming Growth Factor beta; Transforming Growth Factor beta1

Accumulating evidence has implicated the potential of natural compounds in treatment of asthma. Bixin is a natural food coloring isolated from the seeds of Bixa Orellana, which possesses anti-tumor, anti-inflammatory and antioxidative properties. Nevertheless, its therapeutic effect in asthma has not been elucidated. Our present study demonstrated that administration of Bixin suppressed allergic airway inflammation and reversed glucocorticoids resistance, as well as alleviated airway remodeling and airway hyperresponsiveness (AHR) in asthmatic mice. In vitro studies showed that Bixin treatment could inhibit the development of epithelial-mesenchymal transition (EMT) mediated by transforming growth factor beta (TGF-β) signaling. Importantly, Bixin antagonized activation of phosphatidylinositol 3-kinase/protein kinase B (PI3K/Akt) pathway both in vitro and in vivo. Above all, our findings reveal that Bixin functions as a potent antagonist of PI3K/Akt signaling to protect against allergic asthma, highlighting a novel strategy for asthma treatment based on natural products.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Asthma; Bixaceae; Carotenoids; Disease Models, Animal; Female; Humans; Mice; Mice, Inbred BALB C; Oncogene Protein v-akt; Ovalbumin; Phosphatidylinositol 3-Kinases; Respiratory Hypersensitivity; Signal Transduction; Transforming Growth Factor beta

Murine mast cells (MCs) contain two lineages: inducible bone marrow-derived mucosal MCs (MMCs) and constitutive embryonic-derived connective tissue MCs (CTMCs). Here, we use RNA sequencing, flow cytometry, and genetic deletion in two allergic lung inflammation models to define these two lineages. We found that inducible MCs, marked by β7 integrin expression, are highly distinct from airway CTMCs at rest and during inflammation and unaffected by targeted CTMC deletion. β7High MCs expand and mature during lung inflammation as part of a TGF-β-inducible transcriptional program that includes the MMC-associated proteases Mcpt1 and Mcpt2, the basophil-associated protease Mcpt8, granule components, and the epithelial-binding αE integrin. In vitro studies using bone marrow-derived MCs (BMMCs) identified a requirement for SCF in this this TGF-β-mediated development and found that epithelial cells directly elicit TGF-β-dependent BMMC up-regulation of mMCP-1 and αE integrin. Thus, our findings characterize the expansion of a distinct inducible MC subset in C57BL/6 mice and highlight the potential for epithelium to direct MMC development.

Topics: Animals; Asthma; Bone Marrow Cells; Cell Lineage; Integrin beta Chains; Mast Cells; Mice; Mice, Transgenic; Respiratory Mucosa; Transforming Growth Factor beta; Tryptases

The use of probiotics has been broadly popularized due to positive effects in the attenuation of aberrant immune responses such as asthma. Allergic asthma is a chronic respiratory disease characterized by airway inflammation and remodelling.. This study was aimed to evaluate the effect of oral administration of Lactococcus lactis NZ9000 on asthmatic airway inflammation and lung tissue remodelling in rats and its relation to the maintenance of an adequate intestinal barrier.. Wistar rats were ovalbumin (OVA) sensitized and challenged and orally treated with L. lactis. Lung inflammatory infiltrates and cytokines were measured, and remodelling was evaluated. Serum OVA-specific immunoglobulin (Ig) E levels were assessed. We also evaluated changes on intestinal environment and on systemic immune response.. L. lactis diminished the infiltration of proinflammatory leucocytes, mainly eosinophils, in the bronchoalveolar compartment, decreased lung IL-4 and IL-5 expression, and reduced the level of serum allergen-specific IgE. Furthermore, L. lactis prevented eosinophil influx, collagen deposition, and goblet cell hyperplasia in lung tissue. In the intestine, L. lactis-treated asthmatic rats increased Peyer's patch and goblet cell quantity and mRNA expression of IgA, MUC-2, and claudin. Additionally, intestinal morphological alterations were normalized by L. lactis administration. Splenocyte proliferative response to OVA was abolished, and serum levels of transforming growth factor (TGF)-β were increased by L. lactis treatment.. These findings suggest that L. lactis is a potential candidate for asthma prevention, and the effect is mediated by the improvement of intestinal barrier function and systemic TGF-β production.

Topics: Airway Remodeling; Animals; Asthma; Cytokines; Disease Models, Animal; Immunoglobulin E; Inflammation Mediators; Intestinal Mucosa; Lactococcus lactis; Leukocytes; Ovalbumin; Probiotics; Rats; Transforming Growth Factor beta

The airway epithelium is continuously exposed to a variety of pollutants and allergens, thanks to both natural and manmade environmental pollution. With numerous protective mechanisms, the airway epithelium protects the lungs. DNA repair mechanism is one such protective response and its failure could lead to the accumulation of DNA mutations. Our lab had earlier demonstrated the dysfunctional mitochondria in airway epithelium of the asthmatic mice lungs. Here, we show that Ku70 modulation by the administration of Ku70 plasmid attenuates asthma features and reduces mitochondrial dysfunction in the lungs of allergen exposed mice. Ku70 is a key DNA repair protein with diverse roles including VDJ recombination, telomere maintenance, and maintenance of cell homeostasis. Recently, we found a reduction in Ku70 expression in asthmatic airway epithelium, and this was associated with mitochondrial dysfunction in asthmatic condition. In this study, we have shown that Ku70 over-expression in asthmatic mice attenuated airway hyperresponsiveness, airway inflammation, sub-epithelial fibrosis along with reduction in TGF-β with no effect in IL-13 levels and goblet cell metaplasia. Ku70 over-expression in asthmatic mice reduced 8-isoprostane, a marker of oxidative stress, and restored the mitochondrial function in asthmatic mice. We further found these roles of Ku70 to be independent of DNA damage as Ku70 overexpressed mice did not show any reduction in DNA tail, an index of DNA damage. Thus, our findings indicate that Ku70 can attenuate crucial features of asthma along with the restoration of mitochondrial function. This implies that Ku70 could be a therapeutic target for asthma without affecting DNA repair function.

Topics: Animals; Asthma; Bronchoalveolar Lavage; Disease Models, Animal; Genetic Vectors; Injections, Intravenous; Ku Autoantigen; Lung; Male; Mice; Mitochondria; Ovalbumin; Plasmids; Transforming Growth Factor beta

There is growing inclination towards developing bioactive molecule-based strategies for the management of allergic airway inflammation associated respiratory diseases. Vitex negundo Linn., also known as Nirgundi, is one such medicinal plant enriched with phytochemicals and used for inflammatory and respiratory disorders including asthma in traditional system of medicine. Preliminary studies have claimed anti-tussive and bronchodilator potential of V. negundo Linn. However, its attributes as well as molecular mechanism (s) in modulation of asthma mediated by allergic inflammation are yet to be delineated scientifically.. Present study attempted to assess the effectiveness of Vitex negundo leaf extract (VNLE) in mitigation of allergen induced inflammation associated asthmatic lung damage with emphasis to delineate its molecular mechanism (s).. Allergic lung inflammation was established in Balb/c mice using Ovalbumin-lipopolysaccharide (OVA-LPS). Several allergic inflammatory parameters, histopathological changes, alveolar macrophage activation and signalling pathways were assessed to examine protective effects of VNLE. UHPLC-DAD-QTOF-ESI-IMS was used to characterize VLNE.. VNLE administration effectively attenuated LPS-induced oxi-inflammatory stress in macrophages suggesting its anti-inflammatory potential. Further, VNLE showed protective effect in mitigating asthmatic lung damage as evident by reversal of pathological changes including inflammatory cell influx, congestion, fibrosis, bronchial thickness and alveolar collapse observed in allergen group. VNLE suppressed expressions of inflammatory Th1/Th2 cytokines, chemokines, endopeptidases (MMPs), oxidative effector enzyme (iNOS), adhesion molecules, IL-4/IFN-γ release with simultaneous enhancement in levels of IL-10, IFN-γ, MUC3 and tight junction proteins. Subsequent mechanistic investigation revealed that OVA-LPS concomitantly enhanced phosphorylation of NF-κB, PI3K, Akt and p38MAPKs and downregulated AMPK which was categorically counteracted by VNLE treatment. VNLE also suppressed OVA-LPS induced fibrosis, apoptosis, autophagy and gap junction proteins which were affirmed by reduction in TGF-β, Smad2/3/4, Caspase9/3, Bax, LC3A/B, connexin 50, connexin 43 and enhancement in Bcl2 expression. Additionally, suppression of alveolar macrophage activation, inflammatory cells in blood and elevation of splenic CD8+T cells was demonstrated. UHPLC-DAD-QTOF-ESI-IMS revealed presence of iridoids glycoside and phenolics which might contribute these findings.. These findings confer protective effect of VNLE in attenuation of allergic lung inflammation and suggest that it could be considered as valuable medicinal source for developing safe natural therapeutics for mitigation of allergic inflammation during asthma.

Topics: AMP-Activated Protein Kinases; Animals; Anti-Inflammatory Agents; Asthma; Caspases; Disease Models, Animal; Inflammation; Lipopolysaccharides; Lung Injury; Macrophage Activation; Mice, Inbred BALB C; Microtubule-Associated Proteins; NF-kappa B; Ovalbumin; p38 Mitogen-Activated Protein Kinases; Phosphatidylinositol 3-Kinases; Plant Extracts; Proto-Oncogene Proteins c-akt; Proto-Oncogene Proteins c-bcl-2; Signal Transduction; Smad Proteins; Transforming Growth Factor beta; Vitex

Asthma is a heterogeneous disease characterized by multiples respiratory symptoms; this is a polygenic entity that involves a complex interaction of environmental factors and inherent to the individual. To understand the development of asthma, some phenotypes have been proposed.. This work's purpose was to explore different molecules related to asthma development and to define each phenotype's specific characteristics.. 96 adult patients diagnosed with asthma before any treatment were enrolled in the protocol. Spirometric parameters, circulating leukocytes, serum IgE, body mass index, exhaled nitric oxide (FENO), and leukotrienes (LTB4) in urine were determined in each patient. The presence of asthma phenotypes proposed by the Global Initiative for Asthma (GINA) were explored: A) Allergic asthma, B) Non-allergic asthma, C) Late-onset asthma, D) Asthma with persistent airflow limitation, and E) Asthma with overweight and obesity.. In the cohort analyzed, we found four of phenotypes proposed by GINA; however, these phenotypes overlapped, due to this, 4 groups were integrated with allergic, non-allergic and obese patients, which were the main phenotypes. The main overlap was that of patients not-obese allergic, and was characterized by earlier onset, elevated levels of IgE, LTB4 and inflammasome related cytokines. Non-allergic patients had a significant association between interleukin (IL)-18 and IL-18 binding protein (BP) with narrow ratio between these cytokines. Finally, LTB4 had remarkable capacity to discriminate between allergic and not allergic patients.. Asthmatic phenotypes exist as interrelated characteristics and not as discrete entities. High levels of leukotrienes and IgE are hallmarks in the allergic phenotype of asthma.

Topics: Adult; Age of Onset; Asthma; Biomarkers; Cytokines; Eosinophils; Female; Humans; Hypersensitivity; Immunoglobulin E; Inflammasomes; Inflammation Mediators; Intercellular Signaling Peptides and Proteins; Interleukin-18; Interleukin-8; Leukotrienes; Male; Middle Aged; Overweight; Phenotype; Transforming Growth Factor beta

Neutrophilic asthmatics (NA) have less response to inhaled corticosteroids. We aimed to find out the predictor of treatment response in NA.. Asthmatics (n = 115) and healthy controls (n = 28) underwent clinical assessment during 6-month follow-up with standardized therapy. Asthmatics were categorized by sputum differential cell count. The mRNA expressions were measured by RT-qPCR for sputum cytokines (IFN-γ, IL-1β, IL-27, FOXP3, IL-17A, and IL-5). The protein of IL-1β in sputum supernatant was detected by ELISA. Reticular basement membranes (RBM) were measured in the biopsy samples. The role and signaling pathways of IL-1β mediating the epithelial-mesenchymal transition (EMT) process were explored through A549 cells.. NA had increased baseline sputum cell IL-1β expression compared to eosinophilic asthmatics (EA). After follow-up, NA had less improvement in FEV. IL-1β level in baseline sputum predicts the poor lung function improvement in NA. The potential mechanism may be related to IL-1β augmenting TGF-β1-induced steroid-resistant EMT through MAPK signaling pathways.. This study was approved by the Ethics Committee of Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology (IRB ID: 20150406).

Topics: A549 Cells; Adult; Asthma; Epithelial-Mesenchymal Transition; Female; Follow-Up Studies; Humans; Interleukin-1beta; Male; Middle Aged; Neutrophils; Respiratory Mucosa; Transforming Growth Factor beta; Young Adult

Airway remodeling happens in childhood asthma, in parallel with, but not necessarily subsequent to, airway inflammation. The differentiation of airway epithelial cells into myofibroblasts via epithelial-mesenchymal-transition (EMT) is one of the mechanisms underlying airway remodeling. This study aimed at identifying novel molecules involved in pediatric asthma-associated airway remodeling. Asthma model was established by challenging C57BL/6 mouse pups with ovalbumin (OVA). We found that the expression of Cadherin 11 (CDH11), a type II cadherin, was increased by OVA treatments in the airway epithelium. Our earlier microarray data suggested miRNA-451a-5p (miRNA-451a) as a potential regulator of CDH11. In contrast to CDH11, miRNA-451a expression decreased in the asthmatic lung. MiRNA-451a was then packaged into a lentivirus vector and systematically given to the asthmatic pups. Our data indicated that OVA-induced infiltration of inflammatory cells, including eosnophils, neutrophils, macrophages and lymphocytes, was reduced by miRNA-451a over-expression. EMT was initiated in asthmatic mice as demonstrated by increased alpha-smooth muscle actin (α-SMA) positive cells present in airway epithelium, which was inhibited by miRNA-451a. CDH11 elevation in vivo was also inhibited by miRNA-451a. Dual-Luciferase analysis further showed CDH11 as a novel valid target of miRNA-451a. Additionally, in vitro, EMT was triggered in human 16HBE airway epithelial cells by pro-fibrotic transforming growth factor β (TGF-β). Corresponding to the anti-EMT effects observed in vivo, miRNA-451a also inhibited TGF-β-induced collagen deposition in cultured airway epithelial cells by targeting in CDH11. In summary, our study demonstrates that the deregulated miRNA-451a-CDH11 axis contributes to airway remodeling in childhood asthma.

Topics: Airway Remodeling; Allergens; Animals; Animals, Newborn; Asthma; Cadherins; Cells, Cultured; Disease Models, Animal; Humans; Hypersensitivity; Male; Mice; Mice, Inbred C57BL; MicroRNAs; Ovalbumin; Respiratory Mucosa; Signal Transduction; Transforming Growth Factor beta

Allergic asthma (AA) is characterized as a Th2-biased airway inflammation that can develop lung inflammation and remodeling of the respiratory tract.

Topics: Animals; Asthma; Cells, Cultured; Disease Models, Animal; Disease Resistance; Goblet Cells; Humans; Hyperplasia; Hypersensitivity; Interleukin-6; Mice; Mice, Knockout; Pneumonia; Pneumonia, Pneumococcal; Respiratory Mucosa; RNA, Small Interfering; Signal Transduction; Streptococcus pneumoniae; Tight Junctions; Transforming Growth Factor beta

Microalgal exopolysaccharides (EPSs) are given great attention due to their potential biotechnology applications. Purified C. vulgaris EPS was subjected to compositional and sugar linkage analyses, and partial acid hydrolysis. Hydrolysate separation by gel chromatography afforded oligosaccharide fractions. Both, EPS and oligomers were studied by NMR spectroscopy. Data suggest very complex highly branched α-L-arabino-α-L-rhamno-α,β-D-galactan structure. Backbone repeating unit is formed by →2)-α-L-Rha (1 → 3)-α-L-Rha(1 → sequence, highly branched by long 1,6-linked α-D-Galp side chains, further branched at C2, C3 or C4 by α-L-Araf, α-D-Galf and β-D-Galf residues. α-L-Araf form longer 1,2-linked chains branched at C3, C4 or C5. Galf residues are localized as terminal units predominantly in the β configuration, while α-D-Galp and α-L-Araf may be partially O-methylated. Ex vivo biological assays showed increased interleukin-12 (IL-12) and interferon-gamma (INF-γ) levels corresponding to transforming growth factor beta (TGF-β) decrease in guinea pig model experimental asthma. These facts point to the anti-remodelling effect of Chlorella EPS and suggest its possible application in the treatment of asthma and chronic obstructive pulmonary disorder.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Budesonide; Chlorella vulgaris; Chromatography, Gel; Disease Models, Animal; Galactans; Guinea Pigs; Hydrolysis; Interferon-gamma; Interleukin-12; Magnetic Resonance Spectroscopy; Oligosaccharides; Ovalbumin; Transforming Growth Factor beta

Asthma is characterized by airway hyperresponsiveness and allergic inflammation, detrimentally affecting the patients' quality of life. The development of new drugs for the treatment of asthma is warranted to alleviate these issues. Recent studies have demonstrated that sirtuin2 (SIRT2) aggravates asthmatic inflammation by up-regulation of T-helper type 2 responses and macrophage polarization. However, effects of SIRT2 on mast cell activation remain obscure. In this study, we investigated the effects of AGK2, an inhibitor for SIRT2, on mast cell-mediated allergic airway inflammation. Pre-treatment with AGK2 inhibited degranulation of mast cells by suppressing the FcεRI signaling pathway and intracellular calcium influx. The expression of pro-inflammatory cytokines, such as tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-4, IL-5, IL-6, and IL-8, was inhibited via regulation of transcription factors such as NF-κB and NRF2. These effects of AGK2 were verified in passive cutaneous anaphylaxis and acute lung injury animal models. AGK2 attenuated Evans blue pigmentation by inhibiting mast cell activation and lung barrier dysfunction by inhibiting inflammatory responses in these animal models. In the ovalbumin (OVA)-induced allergic airway inflammation murine model, AGK2 alleviated allergic asthma symptoms such as lung histological changes (immune cell and mast cell infiltration, collagen deposition, and α-smooth muscle actin expression) and serum immunoglobulins (Ig) levels (IgE, OVA-specific IgE, IgG1, and IgG2a). Moreover, AGK2 reduced the levels of pro-inflammatory cytokines (TNF-α, IL-1β, IL-4, IL-5, and IL-6) and inflammatory mediators (myeloperoxidase, eosinophil peroxidase, and tumor growth factor-α) in the bronchoalveolar lavage fluid and lung tissues. In addition, the anti-fibrotic effects of AGK2 were verified using lung epithelial cells and TGF-β/Smad reporter stable cells. In conclusion, our findings suggest that SIRT2 plays a role in mast cell-mediated airway inflammatory disease. Therefore, AGK2 is a good potential candidate for treating allergic asthma and lung inflammation.

Topics: A549 Cells; Airway Remodeling; Animals; Anti-Asthmatic Agents; Asthma; Cell Degranulation; Cytokines; Disease Models, Animal; Female; Fibrosis; Furans; Histamine Release; Histone Deacetylase Inhibitors; Humans; Inflammation Mediators; Lung; Male; Mast Cells; Mice, Inbred BALB C; Mice, Inbred ICR; Passive Cutaneous Anaphylaxis; Quinolines; Rats, Sprague-Dawley; Receptors, IgE; Signal Transduction; Sirtuin 2; Transforming Growth Factor beta

Bronchoconstriction was recently shown to cause airway remodeling and induce allergic airway inflammation in asthma. However, the mechanisms how mechanical stress via bronchoconstriction could induce airway inflammation and remodeling remain unclear.. We investigated the effect of bronchoconstriction induced by methacholine inhalation in a murine model of asthma.. BALB/c female mice were sensitized and challenged with ovalbumin (OVA), followed by treatment with methacholine by a nebulizer twice a day for 7 days. Twenty-four hours after the last methacholine treatment, the bronchoalveolar lavage fluid (BALF) and lung tissues were collected. The BALF was analyzed for total and differential cell counts and cytokine levels. The lung tissues were analyzed for goblet cell metaplasia, thickness of the smooth muscle, and lung fibrosis. The expression of cytokines in the lung was also examined.. OVA sensitization and challenge induced infiltration of total cells, macrophages, and eosinophils in the BALF along with goblet cell metaplasia and increased airway smooth muscle hypertrophy. Seven days after the last OVA challenge, untreated mice achieved reduction in airway inflammation, while methacholine maintained the number of BALF total cells, macrophages, and eosinophils. The percentage of goblet cells and the thickness of airway smooth muscle were also maintained by methacholine. Moreover, the treatment of methacholine induced the expression of transforming growth factor (TGF)-β in the lung. This result indicates that the production of TGF-β is involved in induction of airway remodeling caused by bronchoconstriction with methacholine.. Repeated bronchoconstriction caused by methacholine inhalation elicited allergic airway inflammation and airway remodeling.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Bronchoconstriction; Disease Models, Animal; Eosinophils; Female; Humans; Lung; Macrophages; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Transforming Growth Factor beta

Topics: Animals; Asthma; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cell Differentiation; Female; Forkhead Transcription Factors; Immunoglobulin E; Indoleamine-Pyrrole 2,3,-Dioxygenase; Interleukin-17; Interleukin-4; Interleukin-6; Lung; Mice; Mice, Inbred BALB C; Nasal Provocation Tests; Random Allocation; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta; Tryptophan

Asthma is a common obstructive airway disease characterized by inflammation and remodeling with a progressive decline in lung function. Fangxiao Formula (FXF) is an herbal medicine that has achieved significant clinical benefits toward asthma patients, but the relevant mechanism has not yet been clarified. The aim of this study was to determine the inhibitory effects of FXF on airway inflammation and remodeling, and investigate the activities of TGF‑β/Smads signaling pathway in the rat asthma model. Rats were sensitized by ovalbumin (OVA) for six weeks to establish the asthma experimental model. OVA-challenged animals were randomly divided into 5 groups and received different concentrations of FXF or dexamethasone. The animals in blank control group received saline only. Lung tissues were collected and analyzed for determining the inflammatory cells infiltration, HE and PAS staining, airway wall thickness and collagen deposition. The productions of inflammatory cytokine productions were analyzed by ELISA in the bronchoalveolar lavage (BAL) fluid. Immunohistochemical analysis was performed to measure the expression of α-SMA and PCNA in lung tissue after the treatment of FXF. The levels of TGF-β were assessed by both immunohistology and western blotting, and the expression of p-Smad2/3 proteins were determined by western blotting analysis. Our results indicated that FXF attenuated the infiltration of inflammatory cells, decreased the production of Th2 cytokines and simultaneously increased the levels of Th1 cytokine in the asthma rat model. In addition, FXF reduced allergen-induced increased airway wall thickness, goblet cell hyperplasia and collagen deposition. Furthermore, the expression levels of TGF-β and p-Smad3 were obviously reduced after the treatment of FXF. These results indicate that FXF alleviates airway inflammation and remodeling by restoring the balance of Th1/Th2 cytokines and the TGF-β/Smad-3 pathway, therefor providing potential therapeutic approach for asthmatic patients.

Topics: Airway Remodeling; Animals; Asthma; Bronchoalveolar Lavage Fluid; Collagen; Cytokines; Disease Models, Animal; Drugs, Chinese Herbal; Hyperplasia; Inflammation; Lung; Male; Proliferating Cell Nuclear Antigen; Rats, Sprague-Dawley; Signal Transduction; Smad3 Protein; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

Airway remodeling is an important pathological feature of asthma. Excessive deposition of extracellular matrix (e.g., collagen) secreted from fibroblasts is a major factor contributing to airway remodeling. Currently, the mechanism by which collagen continues to be oversynthesized in the airway remains unclear. In this study, we investigated the role of the microRNA-21 (miR-21) and TGFβ/Smad signaling pathway in human bronchial fibroblasts (HBFs), and explored the regulatory mechanism of airway remodeling.. HBFs were cultured in vitro and treated with the transforming growth factor β (TGFβ), receptor inhibitor (SB431542), and TGFβ1. miR-21 and Smad7 overexpressing lentiviruses, as well as an miR-21 interfering lentivirus were constructed and transfected into HBFs. Western blotting was used to determine the expression of airway remodeling-related proteins and proteins in the TGFβ/Smad signaling pathway. miR-21 expression was measured by quantitative real-time PCR.. The high expression of miR-21 induced by TGFβ1 was reduced following the treatment with the SB431542 in HBFs. Smad7 overexpression inhibited the elevated expression of the COL I protein induced by miR-21 overexpression in HBFs. Inhibiting miR-21 expression upregulated the level of Smad7 protein, thus reducing the expression of airway remodeling-related proteins induced by TGFβ1 stimulation in HBFs.. TGFβ1 can induce miR-21 expression in HBFs through the TGFβ/Smad signaling pathway to promote airway remodeling. miR-21 downregulates Smad7, activates the TGFβ/Smad signaling pathway, and promotes airway remodeling. Mutual regulation between miR-21 and the TGFβ/Smad signaling pathway in HBFs promotes airway remodeling.

Topics: Airway Remodeling; Analysis of Variance; Asthma; Blotting, Western; Cells, Cultured; Cohort Studies; Female; Fibroblasts; Gene Expression Regulation; Humans; Male; MicroRNAs; Real-Time Polymerase Chain Reaction; Reference Values; Signal Transduction; Smad7 Protein; Transforming Growth Factor beta

The effects of bone marrow-derived mesenchymal stem cells (BMSCs) and simvastatin combination therapy on serum total and specific IgE levels and lung IL-13 and TGF-β levels in sensitized mouse were examined.. Serum total and specific IgE levels as well as lung IL-13 and TGF-β levels were significantly increased in S group compared to control group (P < 0.001 for all cases). Treatment with BMSCs, simvastatin and their combination significantly decreased serum total and specific IgE levels (P < 0.05 to P < 0.01). However, IL-13 and TGF-β levels were significantly decreased by BMSCs and BMSC + simvastatin combination therapy (P < 0.05 for all cases). The effect of simvastatin and BMSCs combination therapy on serum specific IgE levels as well as lung IL-13 and TGF-β levels were significantly higher than the effect of BMSCs and simvastatin alone (P < 0.001 for IL-13 and P < 0.01 for other cases).. Simvastatin and BMSCs combination therapy affects serum IgE as well as lung IL-13 and TGFβ levels more than BMSC therapy and simvastatin therapy alone which may be due to increased BMSCs migration into the lung tissue.

Topics: Animals; Asthma; Bone Marrow; Bronchoalveolar Lavage; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Immunoglobulin E; Interleukin-13; Lung; Male; Mesenchymal Stem Cells; Mice; Mice, Inbred BALB C; Ovalbumin; Simvastatin; Transforming Growth Factor beta

Th1, Th2, Th17, and Treg cells and their cytokine gene expressions in splenocytes of control mice, ovalbumin sensitized (S), and S treated with dexamethasone and carvacrol during a sensitization period were examined. Th2 and Th17 population as well as the gene expression of IL-4, IL-17, and TGF-β were increased, but Th1, Th1/Th2 ratio, the gene expression of IFN-γ and FOXP3 as well as the IFN-γ/IL-4 ratio were decreased in S compared with control group ( P < 0.001 for all cases). Carvacrol treatment caused significant reduction of Th2 and Th17 population as well as gene expression of IL-4, IL-17, and TGF-β but increase in Treg cells, Th1/Th2 ratio, gene expressions of FOXP3, IFN-γ, and IFN-γ/IL-4 ratio ( P < 0.05 to P < 0.001). The population of Th1, Th2, Th17 cells as well as the gene expression of IL-4, IL-17, and TGF-β were significantly decreased, but only Treg was increased in the dexamethasone treatment group ( P < 0.05 to P < 0.001). Carvacrol treatment during the sensitization period showed a more specific effect on Th1/Th2 imbalance in sensitized mice than dexamethasone, which may indicate the therapeutic potentials of carvacrol in disorders associated with Th1/Th2 imbalance such as asthma.

Topics: Animals; Asthma; Cymenes; Cytokines; Dexamethasone; Gene Expression; Gene Expression Regulation; Humans; Interferon-gamma; Interleukin-17; Interleukin-4; Mice; Ovalbumin; T-Lymphocytes, Regulatory; Th1 Cells; Th17 Cells; Th2 Cells; Transforming Growth Factor beta

Thymic stromal lymphopoietin (TSLP) mediates immune reaction in patients with asthma. Matrix metalloproteinase (MMP), connective tissue growth factor (CTGF), and transforming growth factor-β (TGF-β) are inflammatory mediators whose responses to the anti-TSLP antibody are unknown. This study examined the effect of an anti-TSLP antibody on MMP, CTGF, TGF-β, and airway structural changes in airway remodeling in asthma.. Mice were randomly divided into phosphate-buffered-saline-challenged (PBS), ovalbumin-challenged (OVA), and ovalbumin-challenged with anti-TSLP antibody (OVA + anti-TSLP) groups. Airway responsiveness and serum ovalbumin-specific immunoglobulin E were measured. Differential cell counts and MMP-2 and MMP-9 were evaluated in bronchoalveolar lavage fluid (BALF). Airway structural changes were quantified using morphometric analysis and presentation by immunohistochemistry staining. Lung CTGF, TGF-β, and TSLP were analyzed using western blot.. Airway responsiveness was significantly lower in OVA + anti-TSLP and PBS groups than in OVA group. Airway structural changes exhibited less smooth muscle thickness in OVA + anti-TSLP and PBS groups than in OVA group. MMP-2 and MMP-9 in BALF and CTGF, TGF-β, and TSLP in lungs significantly decreased in OVA + anti-TSLP and PBS groups compared with OVA group.. Anti-TSLP antibody exerts the preventive effect of decreasing airway structural changes through reduction of MMP, TGF-β, and CTGF in airway remodeling of asthma.

Topics: Airway Remodeling; Animals; Antibodies, Monoclonal; Asthma; Connective Tissue Growth Factor; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Inflammation; Lung; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Matrix Metalloproteinases; Methacholine Chloride; Mice; Mice, Inbred BALB C; Ovalbumin; Plethysmography; Thymic Stromal Lymphopoietin; Transforming Growth Factor beta

Tissue remodeling of subepithelial mesenchymal cells is a major pathologic condition of chronic obstructive pulmonary disease and asthma. Fibroblasts contribute to fibrotic events and inflammation in both airway diseases. Recent mechanistic studies established a link between mitochondrial dysfunction or aberrant biogenesis leading to tissue remodeling of the airway wall in asthma. Protein arginine methyltransferase-1 (PRMT1) participated in airway wall remodeling in pulmonary inflammation. This study investigated the mechanism by which PRMT1 regulates mitochondrial mass in primary human airway wall fibroblasts. Fibroblasts from control or asthma patients were stimulated with TGF-β for up to 48 h, and the signaling pathways controlling PRMT1 expression and mitochondrial mass were analyzed. PRMT1 activity was suppressed by the pan-PRMT inhibitor AMI-1. The SMAD2/3 pathway was blocked by SB203580 and C/EBPβ by small interference RNA treatment. The data obtained from unstimulated cells showed a significantly higher basal expression of PRMT1 and mitochondrial markers in asthmatic compared with control fibroblasts. In all cells, TGF-β significantly increased the expression of PRMT1 through SMAD2/3 and C/EBPβ. Subsequently, PRMT1 upregulated the expression of the mitochondria regulators PGC-1α and heat shock protein 60. Both the inhibition of the SAMD2/3 pathway or PRMT1 attenuated TGF-β-induced mitochondrial mass and C/EBPβ and α-SMA expression. These findings suggest that the signaling sequence controlling mitochondria in primary human lung fibroblasts is as follows: TGF-β→SMAD2/3→C/EBPβ→PRMT1→PGC-1α. Therefore, PRMT1 and C/EBPβ present a novel therapeutic and diagnostic target for airway wall remodeling in chronic lung diseases.

Topics: Airway Remodeling; Asthma; Cells, Cultured; Chaperonin 60; Fibroblasts; Humans; Imidazoles; Mitochondria; Naphthalenesulfonates; Peroxisome Proliferator-Activated Receptor Gamma Coactivator 1-alpha; Protein-Arginine N-Methyltransferases; Pyridines; Repressor Proteins; RNA, Small Interfering; Signal Transduction; Smad2 Protein; Smad3 Protein; Steroid Isomerases; Transforming Growth Factor beta; Up-Regulation; Urea

Natural killer T cells (NKT cells) and regulatory T cells (Treg cells) are two important immune regulatory cells which both play critical roles in asthma. Our previous experiments revealed that activation of Treg cells suppressed NKT cells in asthma. However, the possible regulatory effects and the mechanisms linking Treg cells and NKT cells remain poorly understood. The current study was designed to further investigate the regulatory effect and its possible mechanisms of Treg cells on NKT cells function, especially the distribution of NKT cells. Regulatory T cell (Treg), responder T cell (Teff) and Natural killer T cell (NKT) were isolated and purified. After Lentivirus carrying CD39 (Le-CD39) was transfected into Treg cells, the immune phenotype of Treg cells was changed and the suppressive effect of Treg cells on Teff cells with an activation of Treg cells was enhanced, marking with a high expression level of interleukin 10 (IL-10) and transforming growth factor β (TGF-β). Up-regulation of CD39 expression led to lower ATP level in cell culture supernatant. To further explore its function in asthma, we introduced an ovalbumin (OVA)-induced mice model of asthma. And the data showed that up-regulation of CD39 remarkably alleviated OVA-induced hallmarks of the asthma and increased NKT cells in the spleen and peripheral blood; however, decreased NKT cells in the lung. Furthermore, up-regulation of CD39 decreased the levels of interleukin 4 (IL-4) and interferon γ (IFN-γ) in the lung of OVA-treated mice. Our results strongly suggest that Treg cells could be activated by CD39 signal transduction and then affected the distribution of NKT cells in the OVA-induced mice model of asthma.

Topics: Adenosine Triphosphate; Animals; Antigens, CD; Apyrase; Asthma; Cells, Cultured; Disease Models, Animal; Gene Expression; Interleukin-10; Male; Mice, Inbred BALB C; Natural Killer T-Cells; Ovalbumin; Signal Transduction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Up-Regulation

Topics: ADAM Proteins; Allergens; Animals; Asthma; Chlorocebus aethiops; COS Cells; Epithelial Cells; Lung; Mice, Transgenic; Protein Domains; Pyroglyphidae; Transforming Growth Factor beta

We previously reported that neuroimmune semaphorin (Sema) 4A regulates the severity of experimental allergic asthma and increases regulatory T (Treg) cell numbers in vivo; however, the mechanisms of Sema4A action remain unknown. It was also reported that Sema4A controls murine Treg cell function and survival acting through neuropilin 1 (NRP-1) receptor. To clarify Sema4A action on human T cells, we employed T cell lines (HuT78 and HuT102), human PBMCs, and CD4

Topics: Antibodies; Asthma; Cell Line; Cell Proliferation; Cell Survival; Forkhead Transcription Factors; Humans; Interleukin-2; Interleukin-2 Receptor alpha Subunit; Nerve Tissue Proteins; Neuropilin-1; Phenotype; Receptors, Cell Surface; Semaphorins; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Topics: Animals; Asthma; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Coinfection; Disease Models, Animal; Disease Susceptibility; Humans; Influenza A virus; Influenza, Human; Interferon-gamma; Mice, Inbred BALB C; Mice, Inbred C57BL; Pneumococcal Infections; Streptococcus pneumoniae; Survival Analysis; Transforming Growth Factor beta

CD4

Topics: Animals; Antibodies, Monoclonal; Asthma; Biomarkers; Cytokines; Disease Models, Animal; Female; Gene Expression; Immunomodulation; Immunosuppressive Agents; Interleukin-10; Lymphocyte Count; Mice; Schistosomiasis japonica; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Omalizumab is an anti-immunoglobulin E (IgE) monoclonal antibody used in the treatment of severe asthma. Its therapeutic efficacy is primarily attributed to reduction of serum-free IgE and in the expression of high-affinity IgE receptor, fc epsilon RI. However, its effect on the low-affinity IgE receptor fc epsilon RII/CD23 in vivo has not been evaluated.. To determine whether CD23 plays a role in the inflammatory process in severe uncontrolled asthma and whether anti-IgE therapy modulates fc epsilon RII/CD23 expression in these patients.. We evaluated the expression of IgE receptors fc epsilon RI, fc epsilon RII/CD23, and soluble CD23 (sCD23), and the activation state of peripheral blood monocytes (tumor necrosis factor alpha, interleukin (IL) 1-beta, transforming growth factor (TGF) beta expression) in the patients with severe asthma before and after 24 weeks of omalizumab treatment and in the healthy controls. Cytokine expression of monocytes in response to different stimulation (IL-4, IL-4 plus IgE, IL-4 plus IgE plus anti-IgE, and IL-4 plus IgE plus anti-IgE plus anti-CD23 for 72 hours) was determined by enzyme-linked immunosorbent assay.. Treatment with omalizumab (for 24 weeks) improved disease control and pulmonary function (forced expiratory volume in the first second of expiration, 64.5 versus 74%; p = 0.021). Mean ± SE expression of fc epsilon RI on monocytes was higher in the patients with asthma versus the controls (45.7 ± 12.2% versus 18.6 ± 5.8%; p = 0.04) and was reduced after omalizumab treatment (45.7 ± 12.2% versus 15.6 ± 4.4%; p = 0.027). Mean ± SE TGF-beta levels in supernatants from monocytes were reduced in the patients treated with omalizumab (211 ± 6 pg/mL versus 184 ± 9 pg/mL; p = 0.036).. Modulation of the low affinity IgE receptor CD23 in severe asthma is complex, and sCD23 may inversely reflect disease activity. Treatment with omalizumab was associated with reduced monocyte activation.

Topics: Anti-Asthmatic Agents; Asthma; Case-Control Studies; Cytokines; Humans; Monocytes; Omalizumab; Receptors, IgE; Transforming Growth Factor beta; Treatment Outcome

Rhinovirus (RV) can exacerbate allergen-driven asthma. However, it has been suggested that serial infections with RV may also lead to asthma-like features in childhood without prior allergen exposure.. We sought to test the effects of RV infection in the absence of allergen challenge on lung tissue remodeling and to understand whether RV induced factors in common with allergen that promote remodeling.. We infected C57BL/6 mice multiple times with RV in the absence or presence of allergen to assess airway remodeling. We used knockout mice and blocking reagents to determine the participation of LIGHT (TNFSF14), as well as IL-1β and TGF-β, each previously shown to contribute to lung remodeling driven by allergen.. Recurrent RV infection without allergen challenge induced an increase in peribronchial smooth muscle mass and subepithelial fibrosis. Rhinovirus (RV) induced LIGHT expression in mouse lungs after infection, and alveolar epithelial cells and neutrophils were found to be potential sources of LIGHT. Accordingly, LIGHT-deficient mice, or mice where LIGHT was neutralized, displayed reduced smooth muscle mass and lung fibrosis. Recurrent RV infection also exacerbated the airway remodeling response to house dust mite allergen, and this was significantly reduced in LIGHT-deficient mice. Furthermore, neutralizing IL-1β or TGF-β also limited subepithelial fibrosis and/or smooth muscle thickness induced by RV.. Rhinovirus can promote airway remodeling in the absence of allergen through upregulating common factors that also contribute to allergen-associated airway remodeling.

Topics: Airway Remodeling; Allergens; Animals; Asthma; Biomarkers; Biopsy; Bronchoalveolar Lavage; Collagen; Disease Models, Animal; Interleukin-1beta; Mice; Mice, Knockout; Muscle, Smooth; Picornaviridae Infections; Recurrence; Rhinovirus; Transforming Growth Factor beta; Tumor Necrosis Factor Ligand Superfamily Member 14

Asthma has been associated with increased collagen deposition in both conducting airways and alveolar parenchyma. Mast cells (MCs) are key effector cells in asthma and have the ability to affect collagen synthesis. However, the link between clinical control and changes in bronchial and alveolar MC phenotypes and specific collagens in controlled and uncontrolled asthma remains unknown.. To investigate MC phenotypes in correlation with deposition of specific collagen subtypes in patients with controlled and uncontrolled asthma as well as to healthy controls.. In the alveolar parenchyma, the total number of MCs, as well as the number of FcεRI. Patients with uncontrolled atopic asthma have an altered pro-fibrotic MC

Topics: Adult; Asthma; Collagen Type VI; Female; Humans; Male; Mast Cells; Phenotype; Pulmonary Alveoli; Receptors, IgE; Transforming Growth Factor beta

Topics: Airway Obstruction; Asthma; Humans; Myocytes, Smooth Muscle; Smad3 Protein; Transforming Growth Factor beta; Transforming Growth Factor beta1; Transforming Growth Factors

While calcitriol can inhibit airway remodeling in asthmatic mice, the mechanism remains unclear. The purpose of this study was to explore the mechanism of action of calcitriol on airway remodeling in asthma and its interaction with budesonide.. A mouse model of asthma was established by allergic sensitization and challenge with ovalbumin. The mice were treated with budesonide, calcitriol, or budesonide plus calcitriol. The expression of airway remodeling-related proteins, transforming growth factor. Monotherapy with budesonide or calcitriol inhibited the high expression of collagen type I protein and upregulated the low expression of Smad7 in asthmatic mice. There was a synergistic interaction between budesonide and calcitriol in combined treatment. The expression of miR-21 in the combined treatment group was significantly lower than that in the calcitriol treatment group. VDR expression in the combined treatment group was significantly higher than that of the calcitriol treatment group.. Budesonide and calcitriol have a synergistic effect on airway remodeling in asthmatic mice.

Topics: Airway Remodeling; Animals; Anti-Inflammatory Agents; Asthma; Budesonide; Calcitriol; Calcium-Regulating Hormones and Agents; Disease Models, Animal; Drug Evaluation, Preclinical; Drug Synergism; Female; Mice, Inbred BALB C; MicroRNAs; Ovalbumin; Random Allocation; Receptors, Calcitriol; Receptors, Glucocorticoid; Retinoid X Receptors; Smad Proteins; Transforming Growth Factor beta

Asthma is a chronic inflammatory disease of the airway. Repeated inflammatory injury and tissue repair can lead to airway remodeling. The airway epithelial mesenchymal transformation (EMT) plays an important role in airway remodeling of asthma. Various cytokines and signaling pathways, such as transforming growth factor β (TGF-β), nuclear factor-kappa B (NF-κB) and bromodomain-containing protein 4 (BRD4), are involved in the molecular regulation of EMT.. 哮喘是一种慢性气道炎症性疾病，反复的炎症损伤与组织修复可导致气道重塑。气道上皮间质转化(epithelial mesenchymal transition，EMT)在哮喘的气道重塑中发挥了重要作用。转化生长因子β，核因子‐κB和溴结构域蛋白4等多种细胞因子及信号通路参与了EMT的分子调控。.

Topics: Airway Remodeling; Asthma; Cell Cycle Proteins; Epithelial-Mesenchymal Transition; Humans; NF-kappa B; Nuclear Proteins; Transcription Factors; Transforming Growth Factor beta

Asthma is a heterogeneous disease characterized by chronic airway inflammation resulting from airway hyper-responsiveness to diverse stimuli. In this study, we investigated whether microRNA-142 (miR-142) expression affects proliferation and apoptosis in airway smooth muscle cells (ASMCs) during airway remodeling in asthmatic rats.. Thirty six Wistar rats were randomly classified into a control group and an model group. miR-142 mimics and inhibitors were constructed, and ASMCs were transfected using liposomes according to the following groups: blank, negative control (NC), miR-142 mimics, miR-142 inhibitors, si-TGF-β and miR-142 inhibitors + si-TGF-β. We verified that miR-142 targets TGF-β using a dual-luciferase reporter assay. The expression levels of miR-142, TGF-β, EGFR and apoptosis signaling pathway-related genes were determined using RT-qPCR and western blotting. Changes in cell proliferation, cell cycle progression and apoptosis were analyzed using MTT assays and flow cytometry.. Rats with asthma had higher expression levels of EGFR and Akt and lower miR-142 levels. miR-142 was negatively correlated with TGF-β expression. In ASMCs, the expression of TGF-β, EGFR, Akt, phosphorylated-Akt (p-Akt), Bcl-2 and Bcl-xl and the rate of early apoptosis were decreased while expression of Bax and p21 and the proliferation rate were elevated with the upregulation of miR-142. The opposite results were observed with the downregulation of miR-142. Finally, the proliferative rate was decreased while the apoptosis rate was increased and expression levels of EGFR, Akt, p-Akt, Bcl-2 and Bcl-xl were reduced while Bax and p21 were elevated in the ASMCs transfected with miR-142 inhibitors and si-TGF-β.. The results of our study suggest that miR-142 inhibits proliferation and promotes apoptosis in ASMCs during airway remodeling in asthmatic rats by inhibiting TGF-β expression via a mechanism involving the EGFR signaling pathway.

Topics: Airway Remodeling; Animals; Apoptosis; Asthma; Cell Proliferation; ErbB Receptors; Male; MicroRNAs; Myocytes, Smooth Muscle; Rats; Rats, Wistar; Signal Transduction; Transforming Growth Factor beta

Nicotine, an nAChR agonist, shows prominent anti-inflammatory properties, and some studies have illustrated its suppressive effects on inflammation. Here, we have examined whether nicotine as a medicine may have beneficial effects on the treatment of asthma in a mouse model of allergic asthma. BALB/c mice were sensitized with OVA and alum. Two weeks later, the mice received nicotine with concentrations of 1 and 10 mg/kg three times every other day. After 10 days, the mice were challenged with OVA (5%) using an ultrasonic nebulizer and died the next day. Our results showed that the administration of nicotine reduced lung-tissue inflammation, the number of eosinophils in bronchoalveolar fluid, allergen-specific IgE and IL-4 production, while it increased the TGF-β/IL-4 ratio and the number of Treg cells. Our results showed that nicotine applies its suppressive effects in a dose-dependent manner: administration of 10 mg/kg of nicotine showed more suppressive effects than 1 mg/kg. Such data suggested that nicotine might be a good candidate to be used as a medicine in the treatment of allergic asthma by decreasing allergic inflammation severity and potentiating Treg cells proliferation against the allergen.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Asthma; Bronchoalveolar Lavage Fluid; Eosinophils; Immunoglobulin E; Interleukin-4; Male; Mice, Inbred BALB C; Nicotine; Ovalbumin; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

The activation of human bronchial fibroblasts by transforming growth factor-β₁ (TGF-β₁) leads to the formation of highly contractile myofibroblasts in the process of the fibroblast⁻myofibroblast transition (FMT). This process is crucial for subepithelial fibrosis and bronchial wall remodeling in asthma. However, this process evades current therapeutic asthma treatment strategies. Since our previous studies showed the attenuation of the TGF-β₁-induced FMT in response to lipid-lowering agents (e.g., statins), we were interested to see whether a corresponding effect could be obtained upon administration of hypolipidemic agents. In this study, we investigated the effect of fenofibrate on FMT efficiency in populations of bronchial fibroblasts derived from asthmatic patients. Fenofibrate exerted a dose-dependent inhibitory effect on the FMT, even though it did not efficiently affect the expression of α-smooth muscle actin (α-SMA; marker of myofibroblasts); however, it considerably reduced its incorporation into stress fibers through connexin 43 regulation. This effect was accompanied by disturbances in the actin cytoskeleton architecture, impairments in the maturation of focal adhesions, and the fenofibrate-induced deactivation of TGF-β₁/Smad2/3 signaling. These data suggest that fenofibrate interferes with myofibroblastic differentiation during asthma-related subepithelial fibrosis. The data indicate the potential application of fenofibrate in the therapy and prevention of bronchial remodeling during the asthmatic process.

Topics: Asthma; Cell Proliferation; Cell Survival; Cells, Cultured; Connexin 43; Fenofibrate; Fibroblasts; Humans; Myofibroblasts; Signal Transduction; Smad2 Protein; Smad3 Protein; Transforming Growth Factor beta

Chronic respiratory conditions continue to plague millions of people worldwide. We aimed to elucidate the detailed mechanisms of microRNA-485 (miR-485) in airway smooth muscle cell (ASMC) proliferation and apoptosis in chronic asthmatic mice.. A mouse model of chronic asthma was established. Ovalbumin was used to induce chronic asthma in the mice. The levels of transforming growth factor β (TGF-β), interleukin (IL)-4, IL-5, IL-13 and IL-17 in bronchoalveolar lavage fluid in mice were measured by enzyme-linked immunoassays (ELISAs). ASMCs were transfected with miR-485 mimic, miR-485 inhibitor and siRNA-Smurf2. The reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blot analyses were applied to detect the mRNA and protein levels of Smurf2, α-SMA, TGF-β1 and decapentaplegic homolog (Smads). The MTT assay was utilized for cell proliferation, while flow cytometry was conducted to assess cell cycle distribution and apoptosis.. Lower expression of miR-485 and higher expression levels of TGF-β1, IL-4, IL-5, IL-13 and IL-17 were detected in mice with chronic asthma. Smurf2 was identified as the target gene of miR-485. Upregulation of miR-485 mimic and downregulation of Smurf2 decreased expression levels of Smurf2, α-SMA, TGF-β1 and Smad3, inhibited cell proliferation and increased apoptosis, while contrary results were observed in ASMCs transfected with miR-485 inhibitor.. Overexpressed miR-485 inhibits cell proliferation and promotes apoptosis of ASMCs through the Smurf2-mediated TGF-β/Smads signaling pathway in mice with chronic asthma.

Topics: 3' Untranslated Regions; Animals; Antagomirs; Apoptosis; Asthma; Cell Proliferation; Chronic Disease; Cytokines; Female; G1 Phase Cell Cycle Checkpoints; Mice; Mice, Inbred BALB C; MicroRNAs; Myocytes, Smooth Muscle; RNA Interference; RNA, Small Interfering; Signal Transduction; Smad Proteins; Transforming Growth Factor beta; Ubiquitin-Protein Ligases

Bacillus Calmette-Guerin (BCG) is a potent agent for the prevention of tuberculosis. Current studies have regarded BCG as an immunomodulator. However, there is little information on whether it can be used to inhibit airway inflammation and airway remodeling caused by asthma. Therefore, in this study, we investigate the role of epithelial-mesenchymal transition (EMT) in airway inflammation and airway remodeling as well as the possible therapeutic mechanism of BCG for the treatment of asthma. Wistar rats were sensitized and challenged by ovalbumin for 2 weeks or 8 weeks. BCG was subcutaneously administered daily before every ovalbumin challenge to determine its therapeutic effects. The 2 weeks model group showed extensive eosinophilia, chronic inflammatory responses, bronchial wall thickening, airway epithelium damage, increased levels of transforming growth factor β 1 (TGF-β

Topics: Actins; Airway Remodeling; Animals; Asthma; BCG Vaccine; Bronchi; Bronchoalveolar Lavage Fluid; Cadherins; Disease Models, Animal; Epithelial-Mesenchymal Transition; Fibronectins; Goblet Cells; Humans; Inflammation; Ovalbumin; Rats; Rats, Wistar; Transforming Growth Factor beta; Transforming Growth Factor beta1

Asthma is characterized by chronic airway inflammation, leading to structura1 changes in the airway, collectively termed airway remodeling. Airway remodeling is thought to contribute to airway hyper responsiveness and irreversible airflow limitation. The combination of Herba Epimedii (HE) and Fructus Ligustri Lucidi (FLL) decoction and the systemic administration of glucocorticoids (GC) had a synergistic inhibitory action on airway inflammation in the asthmatic model rats. However, the effects of the combination on airway remodeling have not been studied and compared. In the present study, we investigated the effects of the co-administration of combined extracts of HE and FLL with inhaled GC (budesonide) on airway remodeling in the rat asthmatic model induced by ovalbumin (OVA).. Male Sprague-Dawley rats were sensitized to intraperitoneal OVA followed by repetitive OVA challenge for 7 weeks. Treatments included extracts of HE and FLL (Extracts for short, 100 mg/kg by gastric perfusion), budesonide (1 mg budesonide suspension in 50 ml sterile physiological saline, 3 rats in an ultrasonic nebulizer by nebulized inhabation with a flow of 1.6 ml/min for 30 min), and co-administration of extracts of HE and FLL with budesonide (Co-administration for short) for 4 weeks. Lung histomorphometry and bronchoalveolar lavage fluid (BALF) cell count were assessed 24 h after the final OVA challenge. Levels of interleukin (IL)-4, IL-5 and IgE were measured by ELISA. Expressions of Collagen I and Collagen III were tested by immunohistology. Expressions of transforming growth factor (TGF) -β1, TGF-β2 and Smads mRNA were measured by quantitative real-time PCR.. Extracts, budesonide and Co-administration significantly reduced allergen-induced increases in the serum levels of IL-4, IL-5 and IgE, the number of eosinophils in BALF, goblet cell hyperplasia, Collagen III integral optical density (IOD) and the mRNA expression of TGF-β2 and Smad2. Extracts and Co-administration could depress the IOD level of Collagen I and the positive area of Collagen I and Collagen III. Budesonide and Co-administration significantly alleviated the thickening of airway wall. Only Co-administration significantly decreased collagen deposition according to the morphometry of Masson's-stained lung sections, the thickening of airway smooth muscle layer, the number of lymphocytes in BALF and the mRNA expression of TGF-β1 and Smad3, and this was associated with a significant increase in levels of Smad7 mRNA.. The findings suggested that the combination of budesonide and the herbal extracts had a better synergistic effect on airway remodeling in OVA-reduced asthma rats than the single use of budesonide.

Topics: Airway Remodeling; Animals; Anti-Inflammatory Agents; Asthma; Budesonide; Collagen; Drug Synergism; Drugs, Chinese Herbal; Epimedium; Immunoglobulin E; Interleukins; Leukocytes; Ligustrum; Lung; Male; Muscle, Smooth; Ovalbumin; Phytotherapy; Rats, Sprague-Dawley; Real-Time Polymerase Chain Reaction; RNA, Messenger; Smad Proteins; Transforming Growth Factor beta

Helminths and their products are considered to possess therapeutic capability to control or even prevent immune-mediated diseases. Studies suggest that helminths induce a systemic immunomodulatory network, including regulatory T cells and anti-inflammatory interleukin-10 (IL-10), which might play a key role in the protection against the allergic phenotype. Thus, helminthic therapy is becoming of a major interest, and several researchers are enthusiastically tended to explore its role in allergic diseases.. Peripheral blood mononuclear cells (PBMCs) from 25 asthmatic and 25 healthy human were collected. After isolation of PBMCs, they were stimulated with excretory/secretory (ES) antigen of M. marshalli, in incubator with 5% CO. Results showed that the IFN-γ expression was significantly decreased in culture condition with ES Ag of M. marshalli in healthy and asthmatic patients (P < 0.05). Similar data were obtained for IL-4 expression in both healthy individuals (<0.006) and asthmatic patients (<0.001). The increment of regulatory cytokines IL-10 and TGF-β expression was considerably increased in our investigation.. To our knowledge, this is the first report to document the suppressive effect that the M. marshalli ES product has on PBMSC cell culture of asthmatic patients. The present study provides new insights into understanding the immune modulation governed by parasite-derived products and the development of new asthma treatment strategies.

Topics: Adult; Animals; Antigens, Helminth; Asthma; Cytokines; Down-Regulation; Female; Humans; Interferon-gamma; Interleukin-10; Interleukin-4; Leukocytes, Mononuclear; Male; Transforming Growth Factor beta; Trichostrongyloidea; Young Adult

IL-22- and IL-17-producing T cells have important roles in allergic diseases. MicroRNAs (miRNAs) are posttranscriptional regulators of gene expression and modulate numerous biological processes. Little is known about the functions of miRNAs in IL-22/IL-17-producing T cells.. IL-22- and IL-17-positive T cells were sorted from human peripheral blood mononuclear cells (PBMCs) by intracellular staining and dual-secretion assay. miRNA expression profiles were detected with TaqMan array microfluidic cards. T cells were transfected with miRNA mimics. Gene expression was analyzed using RT-qPCR and/or enzyme-linked immunosorbent assay in T-cell subsets and PBMCs from patients with asthma and atopic dermatitis.. The increased expression of miR-323-3p and noncoding RNA nc886 and reduced expression of miR-93, miR-181a, miR-26a, and miR-874 were detected in IL-22-producing T cells. The pathway analysis of the putative targets suggested that these differentially expressed miRNAs could impact the proliferation, differentiation, and effector functions of T cells. Further analyses showed the highest expression for miR-323-3p in IL-22- and IL-17-double-positive T cells and its capacity to suppress multiple genes from the transforming growth factor-β pathway and the production of IL-22 in T cells. An increased expression of miR-323-3p in PBMCs from patients with asthma and reverse correlation between miR-323-3p levels and IL-22 production in PBMCs cultured in T-cell growth conditions was observed.. Our data suggest that miR-323-3p acts in a negative feedback loop to control the production of IL-22 in IL-22/IL-17-producing T cells and might thus impact the T-cell responses in asthma.

Topics: Adult; Asthma; Base Pairing; Cluster Analysis; Gene Expression Profiling; Gene Expression Regulation; Humans; Interleukin-17; Interleukin-22; Interleukins; MicroRNAs; Middle Aged; RNA, Messenger; Signal Transduction; STAT3 Transcription Factor; T-Lymphocyte Subsets; Transforming Growth Factor beta; Young Adult

Asthma is a chronic disease characterized by airway inflammation, obstruction and hyperresponsiveness. Severe asthma affects a small proportion of subjects but results in most of the morbidity, costs and mortality associated with the disease. Studies have suggested that Schistosoma mansoni infection reduces the severity of asthma and prevent atopy.. We evaluated the ability of S. mansoni antigens, Sm29 and Sm29TSP-2 to modulate lymphocyte activation status in response to the allergen of the mite Dermatophagoides pteronyssinus (Der p1) in cell cultures of individuals with asthma.. Thirty four patients were enrolled in this study: seventeen patients with severe asthma (SA group), seventeen patients with mild asthma (MA group) and six controls with no asthma. Peripheral blood mononuclear cells (PBMC) were obtained and stimulated with Sm29 and Sm29TSP-2 in the presence or absence of Der p1. The expression of surface markers and cytokines on lymphocytes was evaluated by flow cytometry and the levels of IL-10 in the culture supernatant were determined by ELISA.. The addition of Sm29 and Sm29TSP-2 antigens to PBMC cultures from both groups of subjects with asthma stimulated with Der p1 reduced the frequency of CD4. These results suggest that the addition of Sm29 and Sm29TSP-2 to the cells cultures from subjects with asthma reduced cell activation markers and altered the cytokine production pattern in a way that can potentialy control the inflammatory response associated with asthma.

Topics: Adult; Animals; Antigens, Helminth; Asthma; Case-Control Studies; CD4-Positive T-Lymphocytes; Cytokines; Female; Humans; Interleukin-10; Interleukin-13; Interleukin-5; Leukocytes, Mononuclear; Lymphocyte Activation; Male; Middle Aged; Schistosoma mansoni; Transforming Growth Factor beta

Asthma and chronic obstructive pulmonary disease (COPD) exacerbations are commonly associated with respiratory syncytial virus (RSV), rhinovirus (RV) and influenza A virus (IAV) infection. The ensuing airway inflammation is resistant to the anti-inflammatory actions of glucocorticoids (GCs). Viral infection elicits transforming growth factor-β (TGF-β) activity, a growth factor we have previously shown to impair GC action in human airway epithelial cells through the activation of activin-like kinase 5 (ALK5), the type 1 receptor of TGF-β. In the current study, we examine the contribution of TGF-β activity to the GC-resistance caused by viral infection. We demonstrate that viral infection of human bronchial epithelial cells with RSV, RV or IAV impairs GC anti-inflammatory action. Poly(I:C), a synthetic analog of double-stranded RNA, also impairs GC activity. Both viral infection and poly(I:C) increase TGF-β expression and activity. Importantly, the GC impairment was attenuated by the selective ALK5 (TGFβRI) inhibitor, SB431542 and prevented by the therapeutic agent, tranilast, which reduced TGF-β activity associated with viral infection. This study shows for the first time that viral-induced glucocorticoid-insensitivity is partially mediated by activation of endogenous TGF-β.

Topics: Anti-Inflammatory Agents; Antiviral Agents; Asthma; Benzamides; Cell Line; Dioxoles; Drug Resistance, Viral; Enzyme Activation; Epithelial Cells; Glucocorticoids; Humans; Influenza A virus; Influenza, Human; ortho-Aminobenzoates; Picornaviridae Infections; Poly I-C; Protein Serine-Threonine Kinases; Pulmonary Disease, Chronic Obstructive; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Respiratory Mucosa; Respiratory Syncytial Virus Infections; Respiratory Syncytial Viruses; Rhinovirus; Transforming Growth Factor beta

In a previous study, we demonstrated that intravenous administration of adipose tissue stem cells (ASCs) could significantly reduce allergic symptoms and suppress eosinophilic inflammation.. To evaluate the secretome of ASCs, we administrated culture supernatant of ASCs (ASC sup, which contains the ASC secretome) and uncultured fresh medium (con sup) into a mouse model of allergic airway inflammation. Subsequently we observed the mice for signs of inflammation and investigated Th1-, Th2-, and T. We found that ASC sup could ameliorate allergic airway inflammation in this model; the value of airway hyperresponsiveness, and the occurrence of inflammatory cell infiltration in the lung, as well as the number of eosinophils, and goblet cells in the lung epithelium were all significantly decreased by ASC sup treatment. In addition, ASC sup treatment significantly decreased the levels of IL-4, IL-5, and IL-13 in the bronchial alveolar lavage fluid and in culture medium of lung-draining lymph node cells of the animal model of acute asthma. We detected numerous CTLA-4 and Foxp3-expressing cells in the lung after ASC sup treatment. ASC sup was found to have a higher concentration of IL-10 and TGF-β compared to con sup.. Stem cells have powerful potential for therapeutic functions in various diseases, but they also have many drawbacks. In this study, we found strong immunosuppressive ability of ASC sup in an allergic airway mouse model. It may be possible to use ASC sup for treatment of many immunological diseases in the near future.

Topics: Adipose Tissue; Animals; Asthma; Bronchoalveolar Lavage Fluid; Culture Media, Conditioned; Disease Models, Animal; Eosinophils; Female; Immunoglobulins; Interleukins; Lung; Lymph Nodes; Mice; Mice, Inbred C57BL; Ovalbumin; Stem Cells; T-Lymphocytes, Regulatory; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

Topics: Animals; Asthma; Disease Models, Animal; Host-Pathogen Interactions; Humans; Leukocytes, Mononuclear; Mice; Picornaviridae Infections; Rhinovirus; Transforming Growth Factor beta; Viral Load

Bronchial asthma is a common chronic airway inflammatory disease. MicroRNAs (miRNAs) play an important role in the pathogenesis of asthma. We have previously shown that miR-20b can inhibit airway inflammation in asthmatic mice, but the exact mechanism is unknown. In the present study, we show that administration of nasal drops containing miR-20b induced an increase in the percentage of Gr1

Topics: Administration, Intranasal; Animals; Asthma; Bone Marrow; CD11b Antigen; Cell Count; Chickens; Chronic Disease; Lung; Mice, Inbred BALB C; MicroRNAs; Myeloid-Derived Suppressor Cells; Spleen; Transforming Growth Factor beta

Upregulation of WISP1 has been demonstrated in lung remodeling. Moreover, it has been recently found that some signaling components of WNT pathway can activate GSK3β signaling to mediate remodeling of airway smooth muscle (ASM) in asthma. Therefore, we hypothesized that WISP1, a signaling molecule downstream of the WNT signaling pathway, is involved in PI3K/GSK3β signaling to mediate ASM remodeling in asthma. Our results showed that WISP1 depletion partly suppressed OVA-induced ASM hypertrophy in vivo. In vitro, WISP1 could induce hBSMC hypertrophy and proliferation, accompanied by upregulation of levels of PI3K, p-Akt, p-GSK3β, and its own expression. TGF-β treatment could increase expression of PI3K, p-Akt, p-GSK3β, and WISP1. SH-5 treatment could partly suppress TGF-β-induced hypertrophy and proliferation of hBSMC, and depress expression of p-GSK3β and WISP1. In conclusion, WISP1 may be a potential inducer of ASM proliferation and hypertrophy in asthma. The pro-remodeling effect of WISP1 is likely due to be involved in PI3K-GSK3β-dependent noncanonical TGF-β signaling.

Topics: Airway Remodeling; Animals; Asthma; Bronchi; CCN Intercellular Signaling Proteins; Cell Line; Cell Movement; Cell Proliferation; Glycogen Synthase Kinase 3 beta; Humans; Hyperplasia; Hypertrophy; Male; Myocytes, Smooth Muscle; Ovalbumin; Phosphatidylinositol 3-Kinases; Proto-Oncogene Proteins; Proto-Oncogene Proteins c-akt; Rats; Rats, Sprague-Dawley; Signal Transduction; Transforming Growth Factor beta

Bacillus Calmette-Guerin (BCG) has been shown to have therapeutic effects on asthma through CD4+CD25+ regulatory T cells (Tregs). We sought to assess pretreatment with inactivated BCG on CD4+CD25+ Tregs and its functional and structural effects in rat asthma model. The rat asthma model was established using ovalbumin (OVA) sensitization and challenge. Ten rats were pretreated with BCG prior to OVA and received continued BCG injections during OVA challenge (BCG+OVA group), 10 rats were treated with OVA alone (OVA group), and 10 rats were treated with saline (control group). After 9 weeks, histamine dihydrochloride effect on airway resistance was measured. Number of CD4+CD25+ Tregs was measured by flow cytometry, expression of Foxp3 and CTLA-4 mRNA was measured, and serum TGF-β levels were determined. Differential cell count in bronchoalveolar lavage fluid (BALF) was determined, and lung tissue was processed and stained with hematoxylin and eosin, Masson's trichrome, and alcine blue and periodic acid Schiff's reaction to evaluate inflammatory cell infiltration, collagen deposition, and presence of goblet cells, respectively. BCG treatment led to an increase in CD4+CD25+ Tregs, as well as an increase in Foxp3 and CTLA-4 expression and serum TGF-β levels. In addition, we observed a decrease in histamine dihydrochloride-induced airway resistance, a decrease in inflammatory leukocytes in BALF, and a decrease in airway remodeling indicators in BCG+OVA-treated rats compared with OVA-treated rats. Intradermally injected inactivated BCG has the potential to improve airway inflammation, airway resistance, and airway remodeling through a mechanism that may involve CD4+CD25+ Tregs.

Topics: Airway Resistance; Animals; Asthma; BCG Vaccine; Disease Models, Animal; Lung; Rats; Rats, Sprague-Dawley; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

We investigated the effects of pan-class I PI3K inhibitor, ZSTK474 on vascular remodeling using a murine model of allergic vasculitis with eosinophil infiltration.. C57BL/6 mice were sensitized with OVA. The positive controls were exposed to aerosolized OVA daily for 7 days. The other group of mice were administered ZSTK474 (30 mg/kg, p.o. daily) in parallel with daily exposure to aerosolized OVA for 7 days. On the 3rd and 7th day, bronchoalveolar lavage (BAL) was performed and the lungs were excised for pathological analysis. Cell differentials were determined and the concentrations of IL-4, IL-5, IL-13 and TGF-βin BAL fluid were measured.. The total cell numbers and eosinophil numbers in BALF were greatly reduced in the ZSTK474-treated group on the 3rd and 7th day after exposure to OVA. The numbers of total white blood cells and eosinophils in the peripheral blood were significantly reduced in the ZSTK474-treated group on the 3rd and 7th day after exposure to OVA. The concentrations of IL-4, IL-5, and IL-13 in BAL fluids were also reduced significantly on the 3rd day in the ZSTK474-treated group. The concentrations of TGF-β in BAL fluids were also reduced significantly on the 3rd and 7th day in the ZSTK474-treated group. The pathological scores reduced significantly in the ZSTK474-treated group compared to the control group.. The PI3K inhibitor, ZSTK474 suppressed pulmonary vascular remodeling in the murine model of allergic vasculitis with eosinophil infiltration. PI3K signal transduction may have a critical role in the immunological process that induces allergic vasculitis.

Topics: Animals; Asthma; Disease Models, Animal; Eosinophils; Female; Hypersensitivity; Inflammation; Interleukin-13; Interleukin-4; Interleukin-5; Leukocyte Count; Lung; Mice; Mice, Inbred C57BL; Ovalbumin; Phosphoinositide-3 Kinase Inhibitors; Transforming Growth Factor beta; Triazines; Vascular Remodeling; Vasculitis

Children with asthma have low vitamin D levels; however, we do not know whether low vitamin D is associated with impaired T-regulatory (Treg) cell population or high IgE receptors (CD23 and CD21) on B cells.. To examine Treg cell function, CD23 and CD21 receptors, vitamin D, and vitamin D-regulating enzymes in children with asthma.. Sixty children (2-6 years old) with asthma and 60 age-matched healthy children were selected as study participants. After collecting demographic and clinical data, blood samples were collected. Treg cells and CD23/CD21 expressions were evaluated by flow cytometry, 25-hydroxyvitamin D3 (25[OH]D3) was quantified by high-performance liquid chromatography, and cytokines and total IgE were estimated by enzyme-linked immunosorbent assay. Messenger RNA (mRNA) expressions of FOXP3, CD23, CD21, vitamin D receptors, and vitamin D-regulating enzymes were assessed by reverse transcription-polymerase chain reaction.. The 25(OH)D3 concentrations and proportion of Treg cells were lower (P < .05) among children with asthma. In contrast, the proportions of B cells with CD23 and CD21 expression were higher (P < .05) in patients with asthma compared with controls. Interleukin 10 and transforming growth factor β were also altered in asthma. The mRNA expression of CD23, CD21, and vitamin D receptors was up-regulated, whereas mRNA expression of vitamin D-regulating enzymes, CYP2R1, CYP27B1, and vitamin D binding protein (except CYP24A1) were up-regulated among children with asthma.. The current study found impaired Treg cell population and high numbers of B cells with IgE receptors (CD23 and CD21) and altered regulatory cytokines in children with asthma, suggesting impaired immune regulation.

Topics: Asthma; B-Lymphocytes; Calcifediol; Child; Child, Preschool; Female; Humans; Interleukin-10; Male; Receptors, Complement 3d; Receptors, IgE; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Experimental studies have reported that aerobic exercise after asthma induction reduces lung inflammation and remodeling. Nevertheless, no experimental study has analyzed whether regular/moderate aerobic training before the induction of allergic asthma may prevent these inflammatory and remodeling processes. For this purpose, BALB/c mice (n = 96) were assigned into non-trained and trained groups. Trained animals ran on a motorized treadmill at moderate intensity, 30 min/day, 3 times/week, for 8 weeks, and were further randomized into subgroups to undergo ovalbumin sensitization and challenge or receive saline using the same protocol. Aerobic training continued until the last challenge. Twenty-four hours after challenge, compared to non-trained animals, trained mice exhibited: (a) increased systolic output and left ventricular mass on echocardiography; (b) improved lung mechanics; (c) decreased smooth muscle actin expression and collagen fiber content in airways and lung parenchyma; (d) decreased transforming growth factor (TGF)-β levels in bronchoalveolar lavage fluid (BALF) and blood; (e) increased interferon (IFN)-γ in BALF and interleukin (IL)-10 in blood; and (f) decreased IL-4 and IL-13 in BALF. In conclusion, regular/moderate aerobic training prior to allergic asthma induction reduced inflammation and remodeling, perhaps through increased IL-10 and IFN-γ in tandem with decreased Th2 cytokines.

Topics: Airway Remodeling; Animals; Asthma; Bronchoalveolar Lavage Fluid; Cytokines; Immunohistochemistry; Inflammation; Interferon-gamma; Interleukin-10; Interleukin-13; Interleukin-4; Lung; Male; Mice; Mice, Inbred BALB C; Microscopy, Electron, Transmission; Ovalbumin; Physical Conditioning, Animal; Pneumonia; Transforming Growth Factor beta

Topics: Airway Remodeling; Asthma; Cell Adhesion Molecules; Dexamethasone; Eosinophils; Epithelial Cells; Fibroblasts; Humans; Hypersensitivity; Inflammation; Interleukin-5; Macrophage-1 Antigen; Th2 Cells; Transforming Growth Factor beta; Up-Regulation; Vascular Cell Adhesion Molecule-1

Asthma is a common worldwide health burden, the prevalence of which is increasing. Recently, the biologically active form of vitamin D3, 1,25-dihydroxyvitamin D3, has been reported to have a protective role in murine asthma; however, the molecular mechanisms by which vitamin D3 attenuates asthma‑associated airway injury remain elusive. In the present study, BALB/c mice were sensitized to ovalbumin (OVA) and were administered 100 ng 1,25-dihydroxyvitamin D3 (intraperitoneal injection) 30 min prior to each airway challenge. The inflammatory responses were measured by ELISA, airway damage was analyzed by hematoxylin and eosin staining, airway remodeling was analyzed by Masson staining and periodic acid‑Schiff staining, markers of oxidative stress were measured by commercial kits, and the expression levels of α‑smooth muscle actin (α-SMA) and the activity of the NF‑E2‑related factor 2 (Nrf2)/heme oxygenase‑1 (HO‑1) and the transforming growth factor‑β (TGF‑β)/Smad signaling pathways were measured by immunohistochemistry and western blotting. The results demonstrated that OVA‑induced airway inflammation and immunoglobulin E overexpression were significantly reduced by vitamin D3 treatment. In addition, treatment with vitamin D3 decreased α‑SMA expression, collagen deposition and goblet cell hyperplasia, and inhibited TGF‑β/Smad signaling in the asthmatic airway. The upregulated levels of malondialdehyde, and the reduced activities of superoxide dismutase and glutathione in OVA‑challenged mice were also markedly restored following vitamin D3 treatment. Furthermore, treatment with vitamin D3 enhanced activation of the Nrf2/HO‑1 pathway in the airways of asthmatic mice. In conclusion, these findings suggest that vitamin D3 may protect airways from asthmatic damage via the suppression of TGF‑β/Smad signaling and activation of the Nrf2/HO‑1 pathway; however, these protective effects were shown to be accompanied by hypercalcemia.

Topics: Airway Remodeling; Animals; Asthma; Biomarkers; Bronchoalveolar Lavage Fluid; Cholecalciferol; Disease Models, Animal; Female; Heme Oxygenase-1; Immunoglobulin E; Mice; NF-E2-Related Factor 2; Oxidative Stress; Protective Agents; Signal Transduction; Smad Proteins; Transforming Growth Factor beta

Airway remodeling includes lung structural changes that have a role in the irreversibility of pulmonary dysfunction shown in chronic bronchial asthmatics. The current experiment investigated the effect of the mitochondrial antioxidant, tiron in comparison with dexamethasone (DEXA) on airway remodeling in chronic asthma. Sensitized BALB/c mice were challenged with ovalbumin (OVA) aerosol for 8weeks, OVA sensitized-challenged mice were treated with either DEXA or tiron, respectively. After that, lung tissue and bronchoaveolar lavage fluid (BALF) were used for measurement of different biological markers. Lungs were examined for histopathological changes and immunohistochemistry. Upon comparing with vehicle treated animals, trion or DEXA treatment significantly reduced eosinophils, lymphocytes, neutrophils and macrophages count in the BALF. Both drugs significantly alleviated chronic OVA-induced oxidative stress as illustrated by decreased pulmonary malondialdenhyde (MDA) and increased glutathione (GSH) and superoxide dismutase (SOD) levels. Asthmatic mice exhibited elevated levels of NOx, IL-13 and TGF-β1 that were reduced by DEXA and tiron. Histopathological changes and increased immunoreactivity of nuclear factor-Kappa B (NF-κ B) in OVA-challenged mice were minimized by tiron and DEXA treatment. In conclusion, in this model of chronic asthma DEXA and tiron ameliorated airway remodeling and inflammation in experimental chronic asthma with no difference between the effect of tiron and DEXA. Tiron has a potential role as adjuvant treatment in chronic asthma.

Topics: 1,2-Dihydroxybenzene-3,5-Disulfonic Acid Disodium Salt; Airway Remodeling; Animals; Asthma; Dexamethasone; Eosinophils; Female; Inflammation; Lung; Lymphocytes; Macrophages; Mice; Mice, Inbred BALB C; Neutrophils; NF-kappa B; Oxidative Stress; Superoxide Dismutase; Transforming Growth Factor beta

Asthma is a chronic respiratory disease characterized by reversible airway obstruction with persistent airway inflammation and airway remodelling, which is associated with increased airway smooth muscle (ASM) mass. Roxithromycin (RXM) has been widely used in asthma treatment; however, its mechanism of action is poorly understood. Vascular endothelial growth factor (VEGF) has been implicated in inflammatory and airway blood vessel remodelling in patients with asthma, and shown to promote ASM cell proliferation. Here, we investigated the effect of RXM on VEGF-induced ASM cell proliferation and attempted to elucidate the underlying mechanisms of action. We tested the effect of RXM on proliferation and cell cycle progression, as well as on the expression of phospho-VEGF receptor 2 (VEGFR2), phospho-extracellular signal-regulated kinase 1/2 (ERK1/2), phospho-Akt, and caveolin-1 in VEGF-stimulated ASM cells. RXM inhibited VEGF-induced ASM cell proliferation and induced cell cycle arrest. Additionally, VEGF-induced ASM cell proliferation was suppressed by inhibiting the activity of ERK1/2, but not that of Akt. Furthermore, RXM treatment inhibits VEGF-induced activation of VEGFR2 and ERK and downregulation of caveolin-1 in a dose-dependent manner. RXM also inhibited TGF-β-induced VEGF secretion by ASM cells and BEAS-2B cells. Collectively, our findings suggest that RXM inhibits VEGF-induced ASM cell proliferation by suppression of VEGFR2 and ERK1/2 activation and caveolin-1 down-regulation, which may be involved in airway remodelling. Further elucidation of the mechanisms underlying these observations should enable the development of treatments for smooth muscle hyperplasia-associated diseases of the airway such as asthma.

Topics: Asthma; Caveolin 1; Cell Cycle Checkpoints; Cell Proliferation; Extracellular Signal-Regulated MAP Kinases; G1 Phase; Humans; Lung; Myocytes, Smooth Muscle; Phosphorylation; Roxithromycin; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factor Receptor-2

The biochemical mechanisms through which eosinophils contribute to asthma pathogenesis are unclear. Here we show eosinophil peroxidase (EPO), an abundant granule protein released by activated eosinophils, contributes to characteristic asthma-related phenotypes through oxidative posttranslational modification (PTM) of proteins in asthmatic airways through a process called carbamylation. Using a combination of studies we now show EPO uses plasma levels of the pseudohalide thiocyanate (SCN

Topics: A549 Cells; Animals; Asthma; Citrulline; Eosinophil Peroxidase; Eosinophils; Humans; Interferon-gamma; Interleukin-13; Mice; Protein Processing, Post-Translational; Th2 Cells; Transforming Growth Factor beta

Although indoleamine 2,3-dioxygenase (IDO)-mediated immune suppression of mesenchymal stem cells (MSCs) has been revealed in septic and tumor microenvironments, the role of IDO in suppressing allergic airway inflammation by MSCs is not well documented. We evaluated the effects of adipose-derived stem cells (ASCs) on allergic inflammation in IDO-knockout (KO) asthmatic mice or asthmatic mice treated with ASCs derived from IDO-KO mice.. ASCs were injected intravenously in wild-type (WT) and IDO-KO asthmatic mice. Furthermore, asthmatic mice were injected with ASCs derived from IDO-KO mice. We investigated the immunomodulatory effects of ASCs between WT and IDO-KO mice or IDO-KO ASCs in asthmatic mice. In asthmatic mice, ASCs significantly reduced airway hyperresponsiveness, the number of total inflammatory cells and eosinophils in bronchoalveolar lavage fluid (BALF), eosinophilic inflammation, goblet hyperplasia, and serum concentrations of total and allergen-specific IgE and IgG1. ASCs significantly inhibited Th2 cytokines, such as interleukin (IL)-4, IL-5, and IL-13, and enhanced Th1 cytokine (interferon-γ) and regulatory cytokines (IL-10, TGF-β) in BALF and lung draining lymph nodes (LLNs). ASCs led to significant increases in regulatory T-cells (Tregs) and IL-10+ T cell populations in LLNs. However, the immunosuppressive effects of ASCs did not significantly differ between WT and IDO-KO mice. Moreover, ASCs derived from IDO-KO mice showed immunosuppressive effects in allergic airway inflammation.. IDO did not play a pivotal role in the suppression of allergic airway inflammation through ASCs, suggesting that it is not the major regulator responsible for suppressing allergic airway inflammation.

Topics: Adipose Tissue; Animals; Asthma; Bronchoalveolar Lavage Fluid; Cell- and Tissue-Based Therapy; Cells, Cultured; Eosinophils; Female; Goblet Cells; Hyperplasia; Immunoglobulin E; Immunoglobulin G; Indoleamine-Pyrrole 2,3,-Dioxygenase; Inflammation; Interferon-gamma; Interleukin-10; Interleukin-13; Interleukin-4; Interleukin-5; Lymphocyte Count; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Methacholine Chloride; Mice; Mice, Inbred C57BL; Mice, Knockout; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

The reparative and immunoregulatory properties of mesenchymal stromal cells (MSCs) have made them attractive candidates for cellular therapy. However, the underlying mechanism of the effects of transplanted MSCs on allergic asthma remains elusive. Here, we show that administration of MSCs isolated from human bone marrow provoked a pronounced polarization in alveolar macrophages to M2 subtypes, rather than induced an increase in the total macrophage number, and efficiently inhibited hallmark features of asthma, including airway hyperresponsiveness and eosinophilic accumulation. Moreover, transforming growth factor beta (TGF-β) signaling pathway appeared to mediate the effects of MSCs on macrophage polarization and subsequently the inhibition of hallmark features of asthma. Inhibition of TGF-β signaling was sufficient to inhibit the macrophage polarization in response to MSCs and consequently reserved the inhibitory effects of macrophage polarization on hallmark features of asthma. Collectively, our data demonstrate that human MSCs have immunosuppressive activity on asthma, which is mediated by TGF-β-signaling-dependent alveolar macrophage polarization.

Topics: Animals; Asthma; Bone Marrow Cells; Bone Marrow Transplantation; Cell Differentiation; Cell Polarity; Cells, Cultured; Eosinophils; Humans; Immunosuppressive Agents; Macrophages, Alveolar; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred NOD; Mice, SCID; Ovalbumin; Signal Transduction; Transforming Growth Factor beta; Transplantation, Heterologous

Asthma is a disease of acute and chronic inflammation in which cytokines play a critical role in orchestrating the allergic inflammatory response. IL-13 and transforming growth factor (TGF)-β promote fibrotic airway remodeling, a major contributor to disease severity. Improved understanding is needed, because current therapies are inadequate for suppressing development of airway fibrosis. IL-13 is known to stimulate respiratory epithelial cells to produce TGF-β, but the mechanism through which this occurs is unknown. Here, we tested the hypothesis that reactive oxygen species (ROS) are a critical signaling intermediary between IL-13 or allergen stimulation and TGF-β-dependent airway remodeling. We used cultured human bronchial epithelial cells and an in vivo mouse model of allergic asthma to map a pathway where allergens enhanced mitochondrial ROS, which is an essential upstream signal for TGF-β activation and enhanced collagen production and deposition in airway fibroblasts. We show that mitochondria in airway epithelium are an essential source of ROS that activate TGF-β expression and activity. TGF-β from airway epithelium stimulates collagen expression in fibroblasts, contributing to an early fibrotic response to allergen exposure in cultured human airway cells and in ovalbumin-challenged mice. Treatment with the mitochondrial-targeted antioxidant, (2-(2,2,6,6-Tetramethylpiperidin-1-oxyl-4-ylamino)-2-oxoethyl)triphenylphosphonium chloride (mitoTEMPO), significantly attenuated mitochondrial ROS, TGF-β, and collagen deposition in OVA-challenged mice and in cultured human epithelial cells. Our findings suggest that mitochondria are a critical source of ROS for promoting TGF-β activity that contributes to airway remodeling in allergic asthma. Mitochondrial-targeted antioxidants may be a novel approach for future asthma therapies.

Topics: Animals; Antioxidants; Asthma; Cells, Cultured; Collagen; Disease Models, Animal; Humans; Interleukin-13; Mice; Mice, Transgenic; Mitochondria; Organophosphorus Compounds; Piperidines; Reactive Oxygen Species; Transforming Growth Factor beta

The aim of this study was to investigate if the aerobic training (AT) reverses airway remodeling (AR) in an asthma model. BALB/c were divided into four groups: control (unsensitized and untrained); ovalbumin (OVA: sensitized and untrained); AT (unsensitized and trained) and OVA + AT. Allergic inflammation was induced with intraperitoneal and OVA inhalation. AT (low intensity; 5×/week; 60 min/session) was performed at 7, 15, and 30 days. Leukocyte counting in the bronchoalveolar lavage fluid; the expression of IL-5, eotaxin, RANTES, intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1); AR features (airway smooth muscle, epithelium thickness, collagen and elastic fibers, mucus production); and AR inducers (transforming growing factor-beta, osteopontin, vascular endothelial growth factor). OVA induced an increase in leukocyte airway migration and increased AR features (P < 0.05). After 7 days, AT reversed the OVA-induced eosinophil and macrophage airway migration, the expression of IL-5, eotaxin, RANTES, ICAM-1, VCAM-1, and all AR inducers. However, total reversion of the AR features and inducers and airway inflammation occurred only after 15 days of AT compared with the OVA groups (P < 0.05) and the effects were maintained until the 30th day. AT reverses AR after 15 days and this effect is preceded by the inhibition of leukocyte migration and occurs simultaneously with the reduction in the expression of inflammatory mediators and AR inducers.

Topics: Airway Remodeling; Animals; Asthma; Bronchi; Bronchoalveolar Lavage Fluid; Cell Movement; Chemokine CCL5; Chemokines, CC; Chronic Disease; Collagen; Disease Models, Animal; Elastic Tissue; Eosinophils; Intercellular Adhesion Molecule-1; Interleukin-5; Leukocytes; Macrophages, Alveolar; Mice; Mice, Inbred BALB C; Mucus; Muscle, Smooth; Osteopontin; Ovalbumin; Physical Conditioning, Animal; Respiratory Mucosa; Transforming Growth Factor beta; Vascular Cell Adhesion Molecule-1; Vascular Endothelial Growth Factor A

Airway smooth muscle (ASM) mass is increased in asthma, and ASM cells from patients with asthma are hyperproliferative and release more IL-6 and CXCL8. The BET (bromo- and extra-terminal) family of proteins (Brd2, Brd3, and Brd4) govern the assembly of histone acetylation-dependent chromatin complexes. We have examined whether they modulate proliferation and cytokine expression in asthmatic ASM cells by studying the effect of BET bromodomain mimics JQ1/SGCBD01 and I-BET762. ASM cells from healthy individuals and nonsevere and severe asthmatics were pretreated with JQ1/SGCBD01 and I-BET762 prior to stimulation with FCS and TGF-β. Proliferation was measured by BrdU incorporation. IL-6 and CXCL8 release was measured by ELISA, and mRNA expression was measured by quantitative RT-PCR. ChIP using a specific anti-Brd4 antibody and PCR primers directed against the transcriptional start site of IL-6 and CXCL8 gene promoters was performed. Neither JQ1/SGCBD01 nor I-BET762 had any effect on ASM cell viability. JQ1/SGCBD01 and I-BET762 inhibited FCS+TGF-β-induced ASM cell proliferation and IL-6 and CXCL8 release in healthy individuals (≥ 30 nM) and in nonsevere and severe asthma patients (≥100 nM), with the latter requiring higher concentrations of these mimics. JQ1/SGCBD01 reduced Brd4 binding to IL8 and IL6 promoters induced by FCS+TGF-β. Mimics of BET bromodomains inhibit aberrant ASM cell proliferation and inflammation with lesser efficiency in those from asthmatic patients. They may be effective in reducing airway remodeling in asthma.

Topics: Asthma; Cell Proliferation; Cells, Cultured; Cytokines; Gene Knockdown Techniques; Humans; RNA, Messenger; Trachea; Transforming Growth Factor beta

We sought to modulate mucosal immune responses using neonatal vaccination to avert the development of allergic airways disease (AAD). Pulmonary pathology in AAD is driven by T helper (TH)2 cytokines, in particular interleukin (IL)4 and IL13, the expression and actions of which are regulated by the transcription factor STAT6. We developed a peptide homolog of STAT6, STAT6-IP. Neonatal mice given, intranasally, STAT6-IP, in an effort to modulate de novo airways immune responses, developed tolerance following subsequent allergen sensitization, with either ovalbumin or ragweed allergens, as demonstrated by reduced TH2 cytokines and specific immunoglobulin (Ig)E and the significant increases in the latency-associated peptide (LAP)(+) T-regulatory (Treg) cell subset and expression of transforming growth factor (TGF)-β. This regulatory phenotype was transferrable by CD4(+) T cells or CD11c(+) dendritic cells (DCs) derived from STAT6-IP-vaccinated mice. Anti-TGF-β treatment during allergen sensitization, however, re-established the pro-inflammatory TH2 response. Thus, neonatal STAT6-IP vaccination induces prospective TGF-β-dependent tolerance to allergen and constitutes a novel highly effective immunomodulatory allergy prevention strategy.

Topics: Adoptive Transfer; Allergens; Animals; Animals, Newborn; Asthma; Cell Separation; Desensitization, Immunologic; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Hypersensitivity; Immune Tolerance; Mice; Mice, Inbred BALB C; Real-Time Polymerase Chain Reaction; STAT6 Transcription Factor; Transforming Growth Factor beta; Vaccines, Subunit

Protein arginine methyltransferase (PRMT)1, methylating both histones and key cellular proteins, has emerged as a key regulator of various cellular processes. This study aimed to identify the mechanism that regulates PRMT1 in chronic Ag-induced pulmonary inflammation (AIPI) in the E3 rat asthma model. E3 rats were challenged with OVA for 1 or 8 wk to induce acute or chronic AIPI. Expression of mRNAs was detected by real-time quantitative PCR. PRMT1, TGF-β, COX2, and vascular endothelial growth factor protein expression in lung tissues was determined by immunohistochemistry staining and Western blotting. In the in vitro study, IL-4-stimulated lung epithelial cell (A549) medium (ISEM) with or without anti-TGF-β Ab was applied to human fibroblasts from lung (HFL1). The proliferation of HFL1 was determined by MTT. AMI-1 (pan-PRMT inhibitor) was administered intranasally to chronic AIPI rats to determine PRMT effects on asthmatic parameters. In lung tissue sections, PRMT1 expression was significantly upregulated, mainly in epithelial cells, in acute AIPI lungs, whereas it was significantly upregulated mainly in fibroblasts in chronic AIPI lungs. The in vitro study revealed that ISEM elevates PRMT1, COX2, and vascular endothelial growth factor expressions, and it promoted fibroblast proliferation. The application of anti-TGF-β Ab suppressed COX2 upregulation by ISEM. AMI-1 inhibited the expression of COX2 in TGF-β-stimulated cells. In the in vivo experiment, AMI-1 administered to AIPI rats reduced COX2 production and humoral immune response, and it abrogated mucus secretion and collagen generation. These findings suggested that TGF-β-induced PRMT1 expression participates in fibroblast proliferation and chronic airway inflammation in AIPI.

Topics: Acute Disease; Animals; Antibodies; Asthma; Cell Proliferation; Chronic Disease; Culture Media, Conditioned; Cyclooxygenase 2; Enzyme Inhibitors; Epithelial Cells; Fibroblasts; Gene Expression Regulation; Humans; Interleukin-4; Lung; Naphthalenesulfonates; Ovalbumin; Pneumonia; Protein-Arginine N-Methyltransferases; Rats; Signal Transduction; Transforming Growth Factor beta; Urea; Vascular Endothelial Growth Factor A

Despite major advances in our understanding of TGF-β signaling in multiple cell types, little is known about the direct target genes of this pathway in human eosinophils. These cells constitute the major inflammatory component present in the sputum and lung of active asthmatics and their numbers correlate well with disease severity. During the transition from acute to chronic asthma, TGF-β levels rise several fold in the lung which drives fibroblasts to produce extracellular matrix (ECM) and participate in airway and parenchymal remodeling. In this report, we use purified blood eosinophils from healthy donors and analyze baseline and TGF-β responsive genes by RNA Seq, and demonstrate that eosinophils (PBE) express 7981 protein-coding genes of which 178 genes are up-regulated and 199 genes are down-regulated by TGF-β. While 18 target genes have been previously associated with asthma and eosinophilic disorders, the vast majority have been implicated in cell death and survival, differentiation, and cellular function. Ingenuity pathway analysis revealed that 126 canonical pathways are activated by TGF-β including iNOS, TREM1, p53, IL-8 and IL-10 signaling. As TGF-β is an important cytokine for eosinophil function and survival, and pulmonary inflammation and fibrosis, our results represent a significant step toward the identification of novel TGF-β responsive genes and provide a potential therapeutic opportunity by selectively targeting relevant genes and pathways.

Topics: Asthma; Computational Biology; Eosinophils; Gene Expression Profiling; Gene Expression Regulation; High-Throughput Nucleotide Sequencing; Humans; Molecular Sequence Annotation; Signal Transduction; Transcriptome; Transforming Growth Factor beta

Pingchuan Formula (PCF) is a traditional Chinese recipe. PCF improves chronic airway inflammation by correcting the imbalance of T-helper cell ratio. The purpose of this study was to investigate the effect of PCF on pathological changes in the lungs of asthmatic mice in terms of Treg/Th17 balance.. A bronchial asthma BALB/c mouse model was established using the ovalbumin excitation method. Distilled water (for MDL group) and drugs (for DEX or PCF group) were administered by gavage immediately after the first excitation. Mice were sacrificed after 7 and 28 d treatment. Lung tissues and bronchoalveolar lavage fluid were collected and lung pathological changes were observed after hematoxylin and eosin staining. Differential cell counts, concentrations of interleukins-6, -17, -23 and TGF-β in bronchoalveolar lavage fluid were determined by enzyme-linked immunosorbent assay. Expression of transcriptional factors Foxp3 and RORγt was determined by immunohistochemistry and immunoblot.. An asthma model was successfully established. After 7 or 28 d treatment, lung pathological changes were improved and concentration of interleukins-6, -17, -23 and TGF-β in bronchoalveolar lavage fluid significantly decreased in the PCF group. RORγt expression in lung tissue was decreased in the PCF group, while Foxp3 expression increased (all P values<0.05 compared with the MDL group). There was no significant difference between the PCF and DEX group except that mice in the PCF group lost less bodyweight.. Treatment with PCF downregulates RORγt, elevates Foxp3 expression, reduces interleukins-6, -17, -23 and TGF-β in bronchoalveolar lavage fluid, thus restoring Th17/Treg balance, improving airway inflammation and reducing asthma symptoms.

Topics: Animals; Asthma; Bronchoalveolar Lavage Fluid; Disease Models, Animal; Drugs, Chinese Herbal; Enzyme-Linked Immunosorbent Assay; Inflammation; Lung; Magnoliopsida; Male; Mice; Mice, Inbred BALB C; Ovalbumin; Phytotherapy; T-Lymphocytes, Regulatory; Th17 Cells; Transforming Growth Factor beta

Exposure to cockroach allergen leads to allergic sensitization and increased risk of developing asthma. Aryl hydrocarbon receptor (AhR), a receptor for many common environmental contaminants, can sense not only environmental pollutants but also microbial insults. Mesenchymal stem cells (MSCs) are multipotent progenitor cells with the capacity to modulate immune responses. In this study, we investigated whether AhR can sense cockroach allergens and modulate allergen-induced lung inflammation through MSCs. We found that cockroach allergen-treated AhR-deficient (AhR(-/-)) mice showed exacerbation of lung inflammation when compared with wild-type (WT) mice. In contrast, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), an AhR agonist, significantly suppressed allergen-induced mouse lung inflammation. MSCs were significantly reduced in cockroach allergen-challenged AhR(-/-) mice as compared with WT mice, but increased in cockroach allergen-challenged WT mice when treated with TCDD. Moreover, MSCs express AhR, and AhR signaling can be activated by cockroach allergen with increased expression of its downstream genes cyp1a1 and cyp1b1. Furthermore, we tracked the migration of i.v.-injected GFP(+) MSCs and found that cockroach allergen-challenged AhR(-/-) mice displayed less migration of MSCs to the lungs compared with WT. The AhR-mediated MSC migration was further verified by an in vitro Transwell migration assay. Epithelial conditioned medium prepared from cockroach extract-challenged epithelial cells significantly induced MSC migration, which was further enhanced by TCDD. The administration of MSCs significantly attenuated cockroach allergen-induced inflammation, which was abolished by TGF-β1-neutralizing Ab. These results suggest that AhR plays an important role in protecting lungs from allergen-induced inflammation by modulating MSC recruitment and their immune-suppressive activity.

Topics: Allergens; Animals; Antibodies, Blocking; Asthma; Cell Movement; Cells, Cultured; Cockroaches; Culture Media, Conditioned; Cytochrome P-450 CYP1A1; Cytochrome P-450 CYP1B1; Epithelial Cells; Hypersensitivity; Immunization; Insect Proteins; Mesenchymal Stem Cells; Mice; Mice, Knockout; Pneumonia; Polychlorinated Dibenzodioxins; Receptors, Aryl Hydrocarbon; Transforming Growth Factor beta

Airway remodelling is a feature of asthma that contributes to loss of lung function. One of the central components of airway remodelling is subepithelial fibrosis. Interleukin (IL)-13 is a key T-helper 2 cytokine and is believed to be the central mediator of allergic asthma including remodelling, but the mechanism driving the latter has not been elucidated in human asthma. We hypothesised that IL-13 stimulates collagen type-1 production by the airway fibroblast in a matrix metalloproteinase (MMP)- and transforming growth factor (TGF)-β1-dependent manner in human asthma as compared to healthy controls. Fibroblasts were cultured from endobronchial biopsies in 14 subjects with mild asthma and 13 normal controls that underwent bronchoscopy. Airway fibroblasts were treated with various mediators including IL-13 and specific MMP-inhibitors. IL-13 significantly stimulated collagen type-1 production in asthma compared to normal controls. Inhibitors of MMP-2 significantly attenuated collagen production in asthma but had no effect in normal controls. IL-13 significantly increased total and active forms of TGF-β1, and this activation was blocked using an MMP-2 inhibitor. IL-13 activated endogenous MMP-2 in asthma patients as compared to normal controls. In an ex vivo model, IL-13 potentiates airway remodelling through a mechanism involving TGF-β1 and MMP-2. These effects provide insights into the mechanism involved in IL-13-directed airway remodelling in asthma.

Topics: Actins; Adult; Asthma; Biopsy; Bronchi; Collagen Type I; Extracellular Matrix Proteins; Female; Fibroblasts; Gene Expression Regulation; Humans; Interleukin-13; Male; Matrix Metalloproteinase 2; Respiratory Function Tests; Transforming Growth Factor beta

Increased airway smooth muscle (ASM) mass is a feature of asthmatic airways, and could result from augmented proliferation. We determined whether proliferation and IL-6 release are abnormal in ASM cells (ASMCs) from patients with severe asthma, and whether these features could be mediated by microRNA-221 and microRNA-222, through modulation of the cyclin-dependent kinase inhibitors, p21(WAF1) and p27(kip1). ASMCs cultured from bronchial biopsies of healthy subjects and patients with nonsevere or severe asthma were studied. Proliferation was measured by the incorporation of bromodeoxyuridine and IL-6 by ELISA. FCS and transforming growth factor (TGF)-β caused greater proliferation and IL-6 release in patients with severe compared with nonsevere asthma and normal subjects. FCS + TGF-β inhibited p21(WAF1) and p27(kip1) expression, and increased microRNA-221 (miR-221) expression in ASMCs from individuals with severe asthma. miR-221, and not miR-222, mimics the increased proliferation and IL-6 release induced by FCS + TGF in healthy ASM, whereas in patients with severe asthma, the inhibition of miR-221, but not miR-222, inhibited proliferation and IL-6 release. miR-221 inhibition led to the increased expression of FCS + TGF-β-induced p21(WAF1) and p27(kip1). Dexamethasone suppressed proliferation in healthy subjects, but not in subjects with asthma. IL-6 was less suppressible by dexamethasone in patients with nonsevere and severe asthma, compared with healthy subjects. miR-221 did not influence the effects of dexamethasone. ASM from patients with severe asthma shows greater proliferation and IL-6 release than in patients with nonsevere asthma, but both groups show corticosteroid insensitivity. miR-221 regulates p21(WAF1) and p27(kip1) expression levels. Furthermore, miR-221 regulates the hyperproliferation and IL-6 release of ASMCs from patients with severe asthma, but does not regulate corticosteroid insensitivity.

Topics: Adult; Asthma; Bronchi; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p21; Cyclin-Dependent Kinase Inhibitor p27; Female; Humans; Interleukin-6; Male; MicroRNAs; Muscle, Smooth; Myocytes, Smooth Muscle; Transforming Growth Factor beta

Asthma is a chronic airway disorder characterized by recurrent attacks of breathlessness and wheezing, affecting 300 million people around the world (available at: www.who.int). To date, genetic factors associated with asthma susceptibility have been unable to explain the full etiology of asthma. Recent studies have demonstrated that the epigenetic disruption of gene expression plays an equally important role in the development of asthma through interaction with our environment. We sensitized 6-week-old C57BL/6J mice with house-dust-mite (HDM) extracts intraperitoneally followed by 5 weeks of exposure to HDM challenges (three times a week) intratracheally. HDM-exposed mice showed an increase in airway hyper-responsiveness (AHR) and inflammation together with structural remodeling of the airways. We applied methylated DNA immunoprecipitation-next generation sequencing (MeDIP-seq) for profiling of DNA methylation changes in the lungs in response to HDM. We observed about 20 million reads by a single-run of massive parallel sequencing. We performed bioinformatics and pathway analysis on the raw sequencing data to identify differentially methylated candidate genes in HDM-exposed mice. Specifically, we have revealed that the transforming growth factor beta signaling pathway is epigenetically modulated by chronic exposure to HDM. Here, we demonstrated that a specific allergen may play a role in AHR through an epigenetic mechanism by disrupting the expression of genes in lungs that might be involved in airway inflammation and remodeling. Our findings provide new insights into the potential mechanisms by which environmental allergens induce allergic asthma and such insights may assist in the development of novel preventive and therapeutic options for this debilitative disease.

Topics: Animals; Asthma; Bronchoalveolar Lavage Fluid; Cells, Cultured; Computational Biology; DNA Methylation; Epigenesis, Genetic; Gene Expression Profiling; High-Throughput Nucleotide Sequencing; Hypersensitivity; Immunoprecipitation; Inflammation; Lung; Male; Mice; Mice, Inbred C57BL; Pyroglyphidae; Signal Transduction; Trachea; Transforming Growth Factor beta

Airway exposure levels of lipopolysaccharide (LPS) are known to determine type I versus type II helper T cell induced experimental asthma. While low doses of LPS derive Th2 inflammatory responses, high (and/or intermediate) LPS levels induce Th1- or Th17-dominant responses. The present paper develops a mathematical model of the phenotypic switches among three Th phenotypes (Th1, Th2, and Th17) in response to various LPS levels. In the present work, we simplify the complex network of the interactions between cells and regulatory molecules. The model describes the nonlinear cross-talks between the IL-4/Th2 activities and a key regulatory molecule, transforming growth factor β (TGF-β), in response to high, intermediate, and low levels of LPS. The model characterizes development of three phenotypes (Th1, Th2, and Th17) and predicts the onset of a new phenotype, Th17, under the tight control of TGF-β. Analysis of the model illustrates the mono-, bi-, and oneway-switches in the key regulatory parameter sets in the absence or presence of time delays. The model also predicts coexistence of those phenotypes and Th1- or Th2-dominant immune responses in a spatial domain under various biochemical and bio-mechanical conditions in the microenvironment.

Topics: Asthma; Computer Simulation; Humans; Lipopolysaccharides; Models, Immunological; Phenotype; Th1 Cells; Th17 Cells; Th2 Cells; Transforming Growth Factor beta

Glucocorticoid-induced TNFR family-related protein (GITR)-mediated activation of JNK was shown to regulate the suppressive activity of CD4(+)CD25(+) naturally occurring T regulatory cells (nTregs) in wild-type (WT) hosts. In this study, CD4(+)CD25(+) T cells were shown to be capable of becoming pathogenic effector cells in sensitized and challenged CD8(-/-) recipient mice. Only GITR-expressing CD4(+)CD25(+) T cells, but neither GITR knocked-in CD4(+)CD25(-) T cells nor GITR-silenced CD4(+)CD25(+) T cells, enhanced development of lung allergic responses. Inhibition of JNK in WT nTregs or nTregs from GITR(-/-)and JNK2(-/-) mice failed to enhance lung allergic responses in sensitized and challenged CD8(-/-) recipient mice. The failure to enhance responses was associated with increased bronchoalveolar lavage fluid levels of IL-10 and TGF-β and decreased levels of IL-5, IL-6, and IL-13. In contrast, nTregs from JNK1(-/-) mice, similar to WT nTregs, were fully effective in enhancing responses. Thus, GITR stimulation of nTregs and signaling through JNK2, but not JNK1, triggered the loss of regulatory function while concomitantly gaining pathogenic CD4(+) T effector cell function responsible for exacerbating asthma-like immunopathology.

Topics: Animals; Asthma; Glucocorticoid-Induced TNFR-Related Protein; Interleukin-10; Lung; MAP Kinase Signaling System; Mice; Mice, Knockout; Mitogen-Activated Protein Kinase 8; Mitogen-Activated Protein Kinase 9; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

MicroRNAs are short non-coding single stranded RNAs that regulate gene expression. While much is known about the effects of individual microRNAs, there is now growing evidence that they can work in co-operative networks. MicroRNAs are known to be dysregulated in many diseases and affect pathways involved in the pathology. We investigated dysregulation of microRNA networks using asthma as the disease model. Asthma is a chronic inflammatory disease of the airways characterized by bronchial hyperresponsiveness and airway remodelling. The airway epithelium is a major contributor to asthma pathology and has been shown to produce an excess of inflammatory and pro-remodelling cytokines such as TGF-β, IL-6 and IL-8 as well as deficient amounts of anti-viral interferons. After performing microRNA arrays, we found that microRNAs -18a, -27a, -128 and -155 are down-regulated in asthmatic bronchial epithelial cells, compared to cells from healthy donors. Interestingly, these microRNAs are predicted in silico to target several components of the TGF-β, IL-6, IL-8 and interferons pathways. Manipulation of the levels of individual microRNAs in bronchial epithelial cells did not have an effect on any of these pathways. Importantly, knock-down of the network of microRNAs miR-18a, -27a, -128 and -155 led to a significant increase of IL-8 and IL-6 expression. Interestingly, despite strong in silico predictions, down-regulation of the pool of microRNAs did not have an effect on the TGF-β and Interferon pathways. In conclusion, using both bioinformatics and experimental tools we found a highly relevant potential role for microRNA dysregulation in the control of IL-6 and IL-8 expression in asthma. Our results suggest that microRNAs may have different roles depending on the presence of other microRNAs. Thus, interpretation of in silico analysis of microRNA function should be confirmed experimentally in the relevant cellular context taking into account interactions with other microRNAs when studying disease.

Topics: Asthma; Base Sequence; Bronchi; Down-Regulation; Epithelial Cells; Gene Expression Regulation; Gene Regulatory Networks; Humans; Inflammation; Interleukin-6; MicroRNAs; Molecular Sequence Data; RNA, Messenger; Signal Transduction; Transforming Growth Factor beta

Sensitization to specific olive pollen-allergens (Ole e 2 and 10) has been correlated with a clinical pattern of asthma. This study analyzes the association between several polymorphims of TNFA (G-308A, C-857T, and C-1031T), IL10 (C-571A and A-1117G), and TGFB (C-509-T) and these sensitizations. These polymorphisms were genotyped by allelic discrimination, in olive pollen-allergic patients (phenotyped for specific Ole e 2 and 10 sensitizations) and healthy controls. Levels of serum-soluble cytokines were correlated with specific genotypes and clinical phenotypes. The results showed that heterozygous TGFB C-509T genotype, besides having the lowest sera TGF- levels, was significantly increased in olive pollen-allergic patients compared with controls. According specific sensitizations, CC genotype of IL10 C-571A could be a protective factor for Ole e 2 sensitization and mainly for asthmatic Ole e 2 sensitized patients compared with asthmatic non-Ole e 2 sensitized patients (OR: 0.26, P = 0.008). In contrast, heterozygous CA genotype was increased in Ole e 2 asthmatic subjects compared to asthmatic non-Ole e 2 sensitized patients. Lastly, heterozygous TNFA G-308A genotype was associated with Ole e 10 sensitization (OR: 2.5, P = 0.04). In conclusion, these results suggest a role of TGF-β1 in olive-pollen sensitization and TNF-α and IL-10 genotypes in the asthma induced by specific olive-pollen allergens.

Topics: Adult; Antigens, Plant; Asthma; Female; Genetic Association Studies; Genetic Predisposition to Disease; Genotype; Humans; Immunization; Immunoglobulin E; Interleukin-10; Male; Middle Aged; Olea; Plant Proteins; Polymorphism, Genetic; Rhinitis, Allergic, Seasonal; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Young Adult

Allergic asthma is an inflammatory disease of the airways in which epithelial cells, dendritic cells and innate lymphoid cells are increasingly implicated. Recent findings suggest that apoptotic cells are phagocytosed by airway epithelial cells in a Rac1-dependent manner and this leads to dampening of innate and adaptive immunity to allergens.

Topics: Allergens; Apoptosis; Asthma; Dendritic Cells; Epithelial Cells; Humans; Interleukin-10; Interleukin-33; Interleukins; Phagocytes; rac1 GTP-Binding Protein; Th2 Cells; Transforming Growth Factor beta

Neutrophilic airway inflammation associated with multiple allergens has been related to steroid-resistant asthma. However, most animal models use only one allergen, which cannot simulate asthma closely as seen in patients. To determine the mechanism of inflammatory process involved in this severe condition, BALB/c mice were repetitively challenged with the pooled extract of dust mite, ragweed, and Aspergillus species (DRA). We found that DRA increased interleukin (IL)-10 and TGF-β levels and neutrophil recruitment in bronchial alveolar lavage fluid. We also found that although dexamethasone suppressed the release of these two cytokines, mast cells recruitment, and mucus hypersecretion, it actually increased neutrophil infiltration and the level of keratinocyte-derived chemokine (mKC), a functional homolog of human IL-8. Treatment of human lung alveolar A549 cells with Der p1, an extract of house dust mite Dermatophagoides pteronyssinus, increased the expression of IL-8 and activity of NF-κB. The elevated IL-8 level was suppressed by BAY11-7082, a selective NF-κB inhibitor, but not by dexamethasone. These results suggest that increased IL-8 (mKC) levels may be involved in steroid-resistant neutrophilic airway inflammation through an NF-κB-dependent pathway.

Topics: Animals; Antigens, Plant; Aspergillus; Asthma; Bronchoalveolar Lavage Fluid; Cell Line, Tumor; Dermatophagoides pteronyssinus; Dexamethasone; Female; Humans; Inflammation; Interleukin-10; Interleukin-8; Mice; Mice, Inbred BALB C; Neutrophil Infiltration; NF-kappa B; Plant Extracts; Transforming Growth Factor beta

Subepithelial fibrosis is one of the most critical structural changes affecting bronchial airway function during asthma. Eosinophils have been shown to contribute to the production of pro-fibrotic cytokines, TGF-β and IL-11, however, the mechanism regulating this process is not fully understood.. In this report, we investigated whether cytokines associated with inflammation during asthma may induce eosinophils to produce pro-fibrotic cytokines.. Eosinophils were isolated from peripheral blood of 10 asthmatics and 10 normal control subjects. Eosinophils were stimulated with Th1, Th2 and Th17 cytokines and the production of TGF-β and IL-11 was determined using real time PCR and ELISA assays.. The basal expression levels of eosinophil derived TGF-β and IL-11 cytokines were comparable between asthmatic and healthy individuals. Stimulating eosinophils with Th1 and Th2 cytokines did not induce expression of pro-fibrotic cytokines. However, stimulating eosinophils with Th17 cytokines resulted in the enhancement of TGF-β and IL-11 expression in asthmatic but not healthy individuals. This effect of IL-17 on eosinophils was dependent on p38 MAPK activation as inhibiting the phosphorylation of p38 MAPK, but not other kinases, inhibited IL-17 induced pro-fibrotic cytokine release.. Th17 cytokines might contribute to airway fibrosis during asthma by enhancing production of eosinophil derived pro-fibrotic cytokines. Preventing the release of pro-fibrotic cytokines by blocking the effect of Th17 cytokines on eosinophils may prove to be beneficial in controlling fibrosis for disorders with IL-17 driven inflammation such as allergic and autoimmune diseases.

Topics: Adult; Asthma; Bronchi; Cytokines; Enzyme-Linked Immunosorbent Assay; Eosinophils; Female; Fibrosis; Humans; Inflammation; Interleukin-11; Interleukin-17; Male; Polymerase Chain Reaction; Th2 Cells; Transforming Growth Factor beta

People living in damp buildings are typically exposed to spore and mycelial fragments of the fungi that grow on damp building materials. There is experimental evidence that this exposure to triple-helical (1, 3)-β-D glucan and low molecular weight toxins may be associated with non-atopic asthma observed in damp and moldy buildings. However, the mechanisms underlying this response are only partially resolved. Using the pure (1, 3)-β-D glucan, curdlan, and the murine macrophage cell line, RAW 264.7, there were two objectives of this study. The first was to determine whether signal transduction pathways activating asthma-associated cell signaling pathways were stimulated using mouse transduction Pathway Finder(®) arrays and quantitative real-time (QRT) PCR. The second objective was to evaluate the dose and temporal responses associated with transcriptional changes in asthma-associated cytokines, the signal transduction receptor gene Dectin-1, and various transcription factor genes related to the induction of asthma using customized RT-PCR-based arrays. Compared to controls, the 10(-7) M curdlan treatment induced significant changes in gene transcription predominately in the NFkB, TGF-β, p53, JAK/STAT, P13/AKT, phospholipase C, and stress signaling pathways. The 10(-8) M curdlan treatment mainly induced NFkB and TGF-β pathways. Compared to controls, curdlan exposures also induced significant dose- and time-dependent changes in the gene translations. We found that that curdlan as a non-allergenic potentiator modulates a network of transduction signaling pathways not only associated with TH-1, TH-2, and TH-3 cell responses including asthma potentiation, but a variety of other cell responses in RAW 264.7 cells. These results help provide mechanistic basis for some of the phenotypic changes associated with asthma that have been observed in in vitro, in vivo, and human studies and open up a hypothesis-building process that could explain the rise of non-atopic asthma associated with fungi.

Topics: Animals; Asthma; beta-Glucans; Cell Line; Cytokines; Dose-Response Relationship, Drug; Lectins, C-Type; Macrophages; Mice; NF-kappa B; Real-Time Polymerase Chain Reaction; Signal Transduction; Time Factors; Transcription, Genetic; Transforming Growth Factor beta

Airway tolerance is the usual outcome of inhalation of harmless antigens. Although T cell deletion and anergy are likely components of tolerogenic mechanisms in the lung, increasing evidence indicates that antigen-specific regulatory T cells (inducible Treg cells [iTreg cells]) that express Foxp3 are also critical. Several lung antigen-presenting cells have been suggested to contribute to tolerance, including alveolar macrophages (MØs), classical dendritic cells (DCs), and plasmacytoid DCs, but whether these possess the attributes required to directly promote the development of Foxp3(+) iTreg cells is unclear. Here, we show that lung-resident tissue MØs coexpress TGF-β and retinal dehydrogenases (RALDH1 and RALDH 2) under steady-state conditions and that their sampling of harmless airborne antigen and presentation to antigen-specific CD4 T cells resulted in the generation of Foxp3(+) Treg cells. Treg cell induction in this model depended on both TGF-β and retinoic acid. Transfer of the antigen-pulsed tissue MØs into the airways correspondingly prevented the development of asthmatic lung inflammation upon subsequent challenge with antigen. Moreover, exposure of lung tissue MØs to allergens suppressed their ability to generate iTreg cells coincident with blocking airway tolerance. Suppression of Treg cell generation required proteases and TLR-mediated signals. Therefore, lung-resident tissue MØs have regulatory functions, and strategies to target these cells might hold promise for prevention or treatment of allergic asthma.

Topics: Allergens; Animals; Antineoplastic Agents; Asthma; Female; Immune Tolerance; Lung; Macrophages, Alveolar; Mice; Mice, Knockout; Signal Transduction; T-Lymphocytes, Regulatory; Toll-Like Receptors; Transforming Growth Factor beta; Tretinoin

The induction of regulatory T cells (Tregs) to suppress aberrant inflammation and immunity has potential as a therapeutic strategy for asthma. Recently, we identified key immunoregulatory components of Streptococcus pneumoniae, type 3 polysaccharide and pneumolysoid (T+P), which suppress allergic airways disease (AAD) in mouse models of asthma. To elucidate the mechanisms of suppression, we have now performed a thorough examination of the role of Tregs. BALB/c mice were sensitized to OVA (day 0) i.p. and challenged intranasal (12-15 d later) to induce AAD. T+P was administered intratracheally at the time of sensitization in three doses (0, 12, and 24 h). T+P treatment induced an early (36 h-4 d) expansion of Tregs in the mediastinal lymph nodes, and later (12-16 d) increases in these cells in the lungs, compared with untreated allergic controls. Anti-CD25 treatment showed that Treg-priming events involving CD25, CCR7, IL-2, and TGF-β were required for the suppression of AAD. During AAD, T+P-induced Tregs in the lungs displayed a highly suppressive phenotype and had an increased functional capacity. T+P also blocked the induction of IL-6 to prevent the Th17 response, attenuated the expression of the costimulatory molecule CD86 on myeloid dendritic cells (DCs), and reduced the number of DCs carrying OVA in the lung and mediastinal lymph nodes. Therefore, bacterial components (T+P) drive the differentiation of highly suppressive Tregs, which suppress the Th2 response, prevent the Th17 response and disable the DC response resulting in the effective suppression of AAD.

Topics: Animals; Asthma; B7-2 Antigen; Cells, Cultured; Dendritic Cells; Female; Inflammation; Interleukin-2; Interleukin-2 Receptor alpha Subunit; Interleukin-6; Lung; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Ovalbumin; Polysaccharides, Bacterial; Receptors, CCR7; Streptococcus pneumoniae; T-Lymphocytes, Regulatory; Th17 Cells; Th2 Cells; Transforming Growth Factor beta

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

The respiratory mucosa is a major coordinator of the inflammatory response in chronic airway diseases, including asthma and chronic obstructive pulmonary disease (COPD). Signals produced by the chronic inflammatory process induce epithelial mesenchymal transition (EMT) that dramatically alters the epithelial cell phenotype. The effects of EMT on epigenetic reprogramming and the activation of transcriptional networks are known, its effects on the innate inflammatory response are underexplored. We used a multiplex gene expression profiling platform to investigate the perturbations of the innate pathways induced by TGF β in a primary airway epithelial cell model of EMT. EMT had dramatic effects on the induction of the innate pathway and the coupling interval of the canonical and noncanonical NF- κ B pathways. Simulation experiments demonstrate that rapid, coordinated cap-independent translation of TRAF-1 and NF- κ B2 is required to reduce the noncanonical pathway coupling interval. Experiments using amantadine confirmed the prediction that TRAF-1 and NF- κ B2/p100 production is mediated by an IRES-dependent mechanism. These data indicate that the epigenetic changes produced by EMT induce dynamic state changes of the innate signaling pathway. Further applications of systems approaches will provide understanding of this complex phenotype through deterministic modeling and multidimensional (genomic and proteomic) profiling.

Topics: Asthma; Epithelial-Mesenchymal Transition; Gene Expression Profiling; Humans; Immunity, Innate; Inflammation; NF-kappa B; Proteomics; Pulmonary Disease, Chronic Obstructive; Respiratory Mucosa; Signal Transduction; TNF Receptor-Associated Factor 1; Transforming Growth Factor beta

Airway inflammation is an important characteristic of asthma and has been associated with airway remodelling and bronchial hyperreactivity. The mucosal microenvironment composed of structural cells and highly specialised extracellular matrix is able to amplify and promote inflammation. This microenvironment leads to the development and maintenance of a specific adaptive response characterized by Th2 and Th17. Bronchial fibroblasts produce multiple mediators that may play a role in maintaining and amplifying this response in asthma.. To investigate the role of bronchial fibroblasts obtained from asthmatic subjects and healthy controls in regulating Th17 response by creating a local micro-environment that promotes this response in the airways.. Human bronchial fibroblasts and CD4(+)T cells were isolated from atopic asthmatics and non-atopic healthy controls. CD4(+)T were co-cultured with bronchial fibroblasts of asthmatic subjects and healthy controls. RORc gene expression was detected by qPCR. Phosphorylated STAT-3 and RORγt were evaluated by western blots. Th17 phenotype was measured by flow cytometry. IL-22, IL17, IL-6 TGF-β and IL1-β were assessed by qPCR and ELISA.. Co-culture of CD4(+)T cells with bronchial fibroblasts significantly stimulated RORc expression and induced a significant increase in Th17 cells as characterized by the percentage of IL-17(+)/CCR6(+) staining in asthmatic conditions. IL-17 and IL-22 were increased in both normal and asthmatic conditions with a significantly higher amount in asthmatics compared to controls. IL-6, IL-1β, TGF-β and IL-23 were significantly elevated in fibroblasts from asthmatic subjects upon co-culture with CD4(+)T cells. IL-23 stimulates IL-6 and IL-1β expression by bronchial fibroblasts.. Interaction between bronchial fibroblasts and T cells seems to promote specifically Th17 cells profile in asthma. These results suggest that cellular interaction particularly between T cells and fibroblasts may play a pivotal role in the regulation of the inflammatory response in asthma.

Topics: Adult; Asthma; Bronchi; Case-Control Studies; CD4-Positive T-Lymphocytes; Cellular Microenvironment; Coculture Techniques; Female; Fibroblasts; Gene Expression Regulation; Humans; Interleukin-17; Interleukin-1beta; Interleukin-22; Interleukin-6; Interleukins; Male; Middle Aged; Nuclear Receptor Subfamily 1, Group F, Member 3; Phosphorylation; Primary Cell Culture; Receptors, CCR6; Signal Transduction; STAT3 Transcription Factor; Transforming Growth Factor beta

Current pharmacotherapy is highly effective in the clinical management of the majority of patients with stable asthma, however severe asthma remains inadequately treated. Prevention of airway remodelling is a major unmet clinical need in the management of patients with chronic severe asthma and other inflammatory lung diseases. Accumulating evidence convincingly demonstrates that activin A, a member of the transforming growth factor (TGF)-β superfamily, is a key driver of airway inflammation, but its role in chronic asthmatic airway remodelling is ill-defined. Follistatin, an endogenously produced protein, binds activin A with high affinity and inhibits its bioactivity. The aim of this study was to test the potential of follistatin as a therapeutic agent to inhibit airway remodelling in an experimental model of chronic allergic airway inflammation.. BALB/c mice were systemically sensitised with ovalbumin (OVA), and challenged with OVA intranasally three times a week for 10 weeks. Follistatin was instilled intranasally during allergen challenge.. Chronic allergen challenge induced mucus hypersecretion and subepithelial collagen deposition which persisted after cessation of challenge. Intranasal follistatin (0.05, 0.5, 5 µg) inhibited the airway remodelling and dose-dependently decreased airway activin A and TGF-β1, and allergen-specific T helper 2 cytokine production in the lung-draining lymph nodes. Follistatin also impaired the loss of TGF-β1 and activin RIB immunostaining in airway epithelium which occurred following chronic allergen challenge.. These data demonstrate that follistatin attenuates asthmatic airway remodelling. Our findings point to the potential of follistatin as a therapeutic for prevention of airway remodelling in asthma and other inflammatory lung diseases.

Topics: Activins; Administration, Intranasal; Airway Remodeling; Analysis of Variance; Animals; Asthma; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Follistatin; Immunohistochemistry; Interleukin-13; Interleukin-4; Interleukin-5; Mice; Mice, Inbred BALB C; Ovalbumin; Random Allocation; Reference Values; Sensitivity and Specificity; Transforming Growth Factor beta

The number of people with asthma has dramatically increased over the past few decades and the cost of care is more than $11·3 billion per year. The use of steroids is the major treatment to control asthma symptoms, but the side effects are often devastating. Seeking new drugs or new strategies to reduce the dose of steroid taken has always been an important task. A bovine whey protein extract (WPE), which is enriched in transforming growth factor-β (TGF-β), has been demonstrated to have the potential for reducing symptoms associated with mild-to-moderate T-helper cell type 1-mediated psoriasis in human subjects. However, whether WPE also has potential for inhibiting T-helper cell type 2 (Th2)-mediated disease remains unclear. In the present study, using a murine asthma model, we found that sensitised mice fed WPE daily, before they were challenged, resulted in reducing airway inflammation, serum ovalbumin-specific IgE, Th2-related cytokine production and airway hyperresponsiveness. Increase in the regulatory T cell (Treg) population in vitro and in vivo was observed when treated with WPE. According to the results from the TGF-β-blocking antibody study, we suggest that TGF-β is the main component that endows WPE with the potential to reduce the generation of Treg. Thus, the present data suggest that WPE has the potential to alleviate the symptoms of asthma by inducing the generation of Treg. Therefore, regular administration of WPE might be potentially beneficial for patients with asthma.

Topics: Animals; Anti-Inflammatory Agents; Asthma; Cattle; Cytokines; Disease Models, Animal; Female; Immunoglobulin E; Inflammation; Mice; Mice, Inbred BALB C; Milk Proteins; Ovalbumin; T-Lymphocytes, Regulatory; Th2 Cells; Transforming Growth Factor beta; Whey Proteins

In asthma, basic fibroblast growth factor (FGF-2) plays an important (patho)physiological role. This study examines the effects of FGF-2 on the transforming growth factor-β (TGF-β)-stimulated differentiation of airway smooth muscle (ASM) cells in vitro. The differentiation of human ASM cells after incubation with TGF-β (100 pM) and/or FGF-2 (300 pM) for 48 hours was assessed by increases in contractile protein expression, actin-cytoskeleton reorganization, enhancements in cell stiffness, and collagen remodeling. FGF-2 inhibited TGF-β-stimulated increases in transgelin (SM22) and calponin gene expression (n = 15, P < 0.01) in an extracellular signal-regulated kinase 1/2 (ERK1/2) signal transduction-dependent manner. The abundance of ordered α-smooth muscle actin (α-SMA) filaments formed in the presence of TGF-β were also reduced by FGF-2, as was the ratio of F-actin to G-actin (n = 8, P < 0.01). Furthermore, FGF-2 attenuated TGF-β-stimulated increases in ASM cell stiffness and the ASM-mediated contraction of lattices, composed of collagen fibrils (n = 5, P < 0.01). However, the TGF-β-stimulated production of IL-6 was not influenced by FGF-2 (n = 4, P > 0.05), suggesting that FGF-2 antagonism is selective for the regulation of ASM cell contractile protein expression, organization, and function. Another mitogen, thrombin (0.3 U ml(-1)), exerted no effect on TGF-β-regulated contractile protein expression (n = 8, P > 0.05), α-SMA organization, or the ratio of F-actin to G-actin (n = 4, P > 0.05), suggesting that the inhibitory effect of FGF-2 is dissociated from its mitogenic actions. The addition of FGF-2, 24 hours after TGF-β treatment, still reduced contractile protein expression, even when the TGF-β-receptor kinase inhibitor, SB431542 (10 μM), was added 1 hour before FGF-2. We conclude that the ASM cell differentiation promoted by TGF-β is antagonized by FGF-2. A better understanding of the mechanism of action for FGF-2 is necessary to develop a strategy for therapeutic exploitation in the treatment of asthma.

Topics: Actins; Asthma; Calcium-Binding Proteins; Calponins; Cell Differentiation; Cells, Cultured; Collagen; Contractile Proteins; ets-Domain Protein Elk-1; Fibroblast Growth Factor 2; Humans; Interleukin-6; MAP Kinase Signaling System; Microfilament Proteins; Muscle Proteins; Muscle, Smooth; Myocytes, Smooth Muscle; Respiratory System; Serum Response Factor; Signal Transduction; Transforming Growth Factor beta

Asthma is a multifactor inflammatory disorder, and its management requires understanding of its various pathogenesis and control mechanisms. Cytokines and other inflammatory mediators are important factors in asthma pathophysiology. In this study, we evaluated the role of cytokine polymorphisms in the asthma susceptibility, progress, control, and lung functions. IL-4-C590T polymorphism by PCR-RFLP method, IFN-γ T+874A, TNF-α-A308G, IL-6 G-174C and TGF-β T+869C variants by ARMS-PCR method and IgE serum level by ELISA technique were determined in 81 asthmatic patients and 124 normal subjects. Asthma diagnosis, treatment and control levels were considered using standard schemes and criteria. TNF-α-308GA genotype was more frequent in asthmatics (P = 0.025, OR 3.352), and polymorphisms between different asthma control levels (P > 0.05) were not different. IFN-γ+874AT genotype had a positive correlation with the familial history of asthma (P = 0.034, OR 2.688). IL-6-174C allele (P = 0.045), TNF-α-308GG genotype (P = 0.002) and TNF-α-308G allele (P = 0.004) showed reduced values, and TNF-α-308GA genotype (P = 0.002) increased FEF25-75 value in asthmatics. IFN-γ+874AA genotype caused a decrease in FVC factor (P = 0.045). This study showed that TNF-α-308GA is a risk factor for asthma, but cytokine gene variants do not affect asthma control and IgE serum levels. Variants producing lower levels of IL-6, TNF-α and IFN-γ are associated with reduced pulmonary capacities. To achieve an appropriate schema for asthma management, further studies with consideration of different aspects in a larger group of patients would be more elucidative.

Topics: Adult; Analysis of Variance; Asthma; Cytokines; Enzyme-Linked Immunosorbent Assay; Female; Genetic Predisposition to Disease; Humans; Interferon-gamma; Interleukin-4; Interleukin-6; Male; Maximal Midexpiratory Flow Rate; Middle Aged; Polymerase Chain Reaction; Polymorphism, Restriction Fragment Length; Polymorphism, Single Nucleotide; Risk Factors; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

The pathophysiology of asthma involves allergic inflammation and remodelling in the airway and airway hyperresponsiveness (AHR) to cholinergic stimuli, but many details of the specific underlying cellular and molecular mechanisms remain unknown. Periostin is a matricellular protein with roles in tissue repair following injury in both the skin and heart. It has recently been shown to be up-regulated in the airway epithelium of asthmatics and to increase active TGF-β. Though one might expect periostin to play a deleterious role in asthma pathogenesis, to date its biological role in the airway is unknown.. To determine the effect of periostin deficiency on airway responses to inhaled allergen.. In vivo measures of airway responsiveness, inflammation, and remodelling were made in periostin deficient mice and wild-type controls following repeated intranasal challenge with Aspergillus fumigatus antigen. In vitro studies of the effects of epithelial cell-derived periostin on murine T cells were also performed.. Surprisingly, compared with wild-type controls, periostin deficient mice developed increased AHR and serum IgE levels following allergen challenge without differences in two outcomes of airway remodelling (mucus metaplasia and peribronchial fibrosis). These changes were associated with decreased expression of TGF-β1 and Foxp3 in the lungs of periostin deficient mice. Airway epithelial cell-derived periostin-induced conversion of CD4(+) CD25(-) cells into CD25(+) , Foxp3(+) T cells in vitro in a TGF-β dependent manner.. Allergen-induced increases in serum IgE and bronchial hyperresponsiveness are exaggerated in periostin deficient mice challenged with inhaled aeroallergen. The mechanism of periostin's effect as a brake on allergen-induced responses may involve augmentation of TGF-β-induced T regulatory cell differentiation.

Topics: Airway Remodeling; Animals; Antigens, Fungal; Aspergillus fumigatus; Asthma; Bronchial Hyperreactivity; Cell Adhesion Molecules; Disease Models, Animal; Hypersensitivity; Immunoglobulin E; Inflammation; Lung; Mice; Mice, Inbred C57BL; Transforming Growth Factor beta

A skewed T-helper (T(h))1/T(h)2 immune response is considered to be the major cause of allergic disorders. Overproduction of T(h)2 cytokines, which promote recruitment and activation of mast cells and eosinophils, plays a key part in the pathogenesis of allergic asthma. The mechanisms by which omalizumab is effective in asthma treatment are not yet fully understood. A 16-year-old girl who was experiencing frequent asthma attacks in spite of daily administration of budesonide (640 μg) and montelukast (10mg) was given omalizumab (375 mg) at intervals of 2 weeks, to prevent a visit to the emergency room. Plasma levels of T(h)1 cytokines [interferon (IFN)-γ and interleukin (IL)-12p70], T(h)2 cytokines (IL-4 and IL-13), other proinflammatory and regulatory cytokines [IL-6, IL-10, IL-17, tumor necrosis factor (TNF)-α, and transforming growth factor (TGF)-β], chemokines [monocyte chemotactic protein (MCP)-1, chemokine ligand (CCL)7, and CCL17], and soluble Fas ligand (sFasL) were measured before treatment and after treatment for 8 weeks. She showed a good clinical response to omalizumab: her lung function parameters improved and the use of β2-agonist decreased. No emergency room visits were required after omalizumab treatment for 8 weeks. Plasma levels of sFasL and TGF-β showed obvious increases after omalizumab therapy. IL-12p70 levels were decreased as compared to the corresponding baseline levels. These findings suggest that the effects of omalizumab in asthma treatment are not restricted to the regulation of the skewed T(h)1/T(h)2 cytokine immune response, and sFasL-mediated apoptosis and regulatory T-cell (Treg)-mediated TGF-β seem to have important roles in the therapeutic effects of omalizumab.

Topics: Acetates; Adolescent; Antibodies, Anti-Idiotypic; Antibodies, Monoclonal, Humanized; Apoptosis; Asthma; Budesonide; Cyclopropanes; Fas Ligand Protein; Female; Humans; Immunologic Factors; Lung; Omalizumab; Quinolines; Respiratory Function Tests; Sulfides; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Treatment Outcome

Patients with allergic bronchial asthma can develop various types of asthmatic response to bronchial challenge with allergens, such as immediate (IAR), late (LAR) or delayed (DYAR) response, due to different immunological mechanisms.. The DYAR, beginning within 26- 32 h after the challenge, reaching its maximum between 32 and 48 h and resolving within 56 h (p < 0.001), differs from IAR and LAR regarding the clinical features, diagnostic and immunological parameters.. The repeated DYAR (p < 0.001) in 28 patients was supplemented with recording of cytokine concentrations in the serum before and up to 72 h after the bronchial challenge by means of enzyme-linked immunoassay.. The DYAR was accompanied by a significant increase in the serum concentration (p < 0.05) of IL-2 at 24, 36 and 48 h; IL-10 at 12, 24, 36 and 48 h; IL-18 at 12 and 24 h; IFN-γ at 24, 36, 48 and 56 h; G-CSF at 1 and 12 h; TNF-α at 12, 36 and 48 h and TGF-β at 12 and 36 h, and a significant decrease in the concentration (p < 0.05) of IL-7 at 36 and 48 h and IL-12p70 at 12 h, as compared both with the prechallenge and with PBS control values. No significant changes in the serum cytokines were recorded during the PBS controls (p > 0.2).. These results support the evidence for an active involvement of the Th(1) cells, neutrophils, monocytes, macrophages, epithelial cells and endothelial cells, upon cooperation of other cell types, in the immunological mechanism(s) underlying the DYAR.

Topics: Adult; Allergens; Asthma; Bronchial Provocation Tests; Case-Control Studies; Cytokines; Female; Granulocyte Colony-Stimulating Factor; Humans; Interferon-gamma; Interleukins; Male; Middle Aged; Time Factors; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

In a biphasic, ovalbumin (OVA)-induced murine asthma model where allergic airway disease is followed by resolution and the development of local inhalational tolerance (LIT), transforming growth factor (TGF)-β-expressing CD5(+) B cells were selectively expanded locally in hilar lymph nodes (HLN) of LIT mice. LIT HLN CD5(+) B cells, but not LIT HLN CD5(-) B cells, induced expression of Foxp3 in CD4(+)CD25(-) T cells in vitro. These CD5(+) regulatory B cells (Breg) and CD4(+)Foxp3(+) T cells demonstrated similar increases in expression of chemokine receptors (CXCR4 and CXCR5) and co-localized in HLN B cell zones of LIT mice. The adoptive transfer of LIT HLN CD5(+) B cells, but not LIT HLN CD5(-) B cells, increased the number of CD4(+)Foxp3(+) T cells in the lung and inhibited airway eosinophilia in this OVA model. Thus, Breg in HLNs of LIT mice reside in a CD5(+) TGF-β-producing subpopulation and co-localize with CD4(+)Foxp3(+) T cells.

Topics: Adoptive Transfer; Animals; Asthma; B-Lymphocytes, Regulatory; CD4 Antigens; CD5 Antigens; Cell Proliferation; Disease Models, Animal; Eosinophilia; Female; Forkhead Transcription Factors; Gene Expression; Immune Tolerance; Lung; Lymph Nodes; Lymphocyte Count; Mice; Ovalbumin; Receptors, CXCR4; Receptors, CXCR5; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Both nature and induced regulatory T (Treg) lymphocytes are potent regulators of autoimmune and allergic disorders. Defects in endogenous Treg cells have been reported in patients with allergic asthma, suggesting that disrupted Treg cell-mediated immunological regulation may play an important role in airway allergic inflammation. In order to determine whether adoptive transfer of induced Treg cells generated in vitro can be used as an effective therapeutic approach to suppress airway allergic inflammation, exogenously induced Treg cells were infused into ovalbumin-sensitized mice prior to or during intranasal ovalbumin challenge. The results showed that adoptive transfer of induced Treg cells prior to allergen challenge markedly reduced airway hyperresponsiveness, eosinophil recruitment, mucus hyper-production, airway remodeling, and IgE levels. This effect was associated with increase of Treg cells (CD4(+)FoxP3(+)) and decrease of dendritic cells in the draining lymph nodes, and with reduction of Th1, Th2, and Th17 cell response as compared to the controls. Moreover, adoptive transfer of induced Treg cells during allergen challenge also effectively attenuate airway inflammation and improve airway function, which are comparable to those by natural Treg cell infusion. Therefore, adoptive transfer of in vitro induced Treg cells may be a promising therapeutic approach to prevent and treat severe asthma.

Topics: Adoptive Transfer; Airway Remodeling; Animals; Asthma; CD4 Lymphocyte Count; Cells, Cultured; Cytokines; Dendritic Cells; Female; Forkhead Transcription Factors; Lung; Lymph Nodes; Mice; Mice, Inbred C57BL; Ovalbumin; Spleen; T-Lymphocytes, Helper-Inducer; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Lung fibrosis can affect the parenchyma and the airways, classically giving rise to idiopathic pulmonary fibrosis (IPF) in the parenchyma or airway remodeling in asthma and chronic obstructive pulmonary disease. TGF-β activation has been implicated in the fibrosis of both IPF and airway remodeling. However, the mechanisms of TGF-β activation appear to differ depending on the cellular and anatomical compartments, with implications on disease pathogenesis. Although it appears that epithelial cell activation of TGF-β by the αvβ6 integrin is central in IPF, mesenchymal activation of TGF-β by the αvβ5 and αvβ8 integrins appears to predominate in airway remodeling. Interestingly, the mechanism of TGF-β by the integrins αvβ6 and αvβ5 is shared, relying on cytoskeletal changes, whereas activation of TGF-β by the αvβ8 integrin is distinct, relying on proteolytic cleavage of the latency-associated peptide of TGF-β by matrix metalloproteinase 14. This article describes the mechanisms through which epithelial cells activate TGF-β by the αvβ6 integrin and mesenchymal cells activate TGF-β by the αvβ5 integrin, and highlights their roles in lung fibrosis.

Topics: Airway Remodeling; Asthma; Epithelial Cells; Epithelial-Mesenchymal Transition; Extracellular Matrix; Humans; Integrin alpha Chains; Integrin alphaV; Mesoderm; Pulmonary Alveoli; Pulmonary Fibrosis; Transforming Growth Factor beta

Several epidemiologic studies have shown that growing up in a farming environment is associated with a decreased risk of allergies. A factor that correlates strongly with this effect is the early ingestion of unheated cow's milk. Although, to date, no controlled studies on raw milk consumption have been performed to formally demonstrate this effect, several factors in bovine milk have been described that might explain how raw cow's milk consumption can decrease the risk of allergies. In addition, increasing knowledge on the immunologically active factors in breast milk have also contributed to our understanding of the effects of bovine milk in infants because many of the factors in bovine milk are expected to have functional effects in human subjects as well. Here we review these factors and their mechanisms of action and compare their presence in bovine milk and breast milk. A better understanding of these factors, as well as how to retain them, might ultimately lead to the development of mildly processed milk and infant nutrition products that could become a part of preventive strategies to reduce the incidence of allergic disease.

Topics: Animals; Asthma; Breast Feeding; Cattle; Humans; Milk Hypersensitivity; Milk Proteins; Transforming Growth Factor beta

Allergic asthma is an inflammatory disorder characterized by infiltration of the airway wall with inflammatory cells driven mostly by activation of Th2-lymphocytes, eosinophils and mast cells. There is a link between increased allergy and a reduction of some infections in Western countries. Epidemiological data also show that respiratory allergy is less frequent in people exposed to orofecal and foodborne microbes such as Toxoplasma gondii. We previously showed that both acute and chronic parasite T. gondii infection substantially blocked development of airway inflammation in adult BALB/c mice. Based on the high levels of IFN-γ along with the reduction of Th2 phenotype, we hypothesized that the protective effect might be related to the strong Th1 immune response elicited against the parasite. However, other mechanisms could also be implicated. The possibility that regulatory T cells inhibit allergic diseases has received growing support from both animal and human studies. Here we investigated the cellular mechanisms involved in T. gondii induced protection against allergy. Our results show for the first time that thoracic lymph node cells from mice sensitized during chronic T. gondii infection have suppressor activity. Suppression was detected both in vitro, on allergen specific T cell proliferation and in vivo, on allergic lung inflammation after adoptive transference from infected/sensitized mice to previously sensitized animals. This ability was found to be contact-independent and correlated with high levels of TGF-β and CD4(+)FoxP3(+) cells.

Topics: Animals; Asthma; Bronchoalveolar Lavage; CD4 Antigens; Forkhead Transcription Factors; Hypersensitivity; Inflammation; Interferon-gamma; Lymph Nodes; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Mice, Knockout; Respiratory Hypersensitivity; Th2 Cells; Toxoplasma; Toxoplasmosis; Transforming Growth Factor beta

(±)-Praeruptorin A (PA) is a pair of coumarin enantiomers isolated from the root of Peucedanum praeruptorum Dunn (PPD), a common Chinese herbal medicine for the treatment of asthma. Considering its anti-inflammatory, anti-contractile and anti-hyperplasia activities, the effects of PA on airway inflammation and airway remodeling were investigated using a murine model of chronic asthma. Ovalbumin-sensitized BALB/c mice were challenged with ovalbumin to induce asthma every other day on eight successive weeks. PA was administered intragastrically before every ovalbumin challenge. Airway responsiveness was evaluated by a lung function analysis system 48 h after the last ovalbumin challenge. The total and differential leukocytes in bronchoalveolar lavage fluid (BALF) were counted using a hemocytometer and Diff-Quick-stained smears. Lung tissue samples were used for hematoxylin and eosin, periodic acid Schiff, Masson's trichrome and α-SMA immunohistochemistry staining. Levels of cytokines in BALF, immunoglobulin (Ig) E in serum as well as expression of TGF-β1 and Smad proteins in lung tissue were measured by enzyme-linked immunosorbent assay, immunohistochemistry or western blot analysis. Compared with the model group, PA suppressed airway inflammation, airway hyperresponsive and remodeling, reduced levels of IL-4 and IL-13 in BALF, and IgE in serum, inhibited expression of TGF-β1 and pSmad2/3, up-regulated the expression of Smad7 in lung tissue, and also increased the levels of INF-γ in BALF. These results suggested that PA significantly suppressed airway inflammation and airway remodeling induced by ovalbumin challenge, and is a potential candidate for the treatment of asthma.

Topics: Airway Remodeling; Animals; Anti-Asthmatic Agents; Asthma; Budesonide; Cell Line; Coumarins; Dose-Response Relationship, Drug; Female; Gene Expression Regulation; Immunoglobulin E; Inflammation; Lung; Macrophages; Mice; Mice, Inbred BALB C; Molecular Structure; Smad Proteins; Transforming Growth Factor beta

Allergic asthma is showed an increase in Th2-cytokine and IgE levels and an accumulation activation of Th2 cells, eosinophils and mast cells. However, recent studies focused on cell-based mechanisms for the pathogenesis of allergic asthma. Objectives. In this study, we compare the anti-IgE treatment modality in the dynamics of immune system cytokine levels in severe persistent asthma (SPA) patients who had no other any allergic disease, newly diagnosed allergic asthma patients and healthy volunteers.. The study population consisted of 14 SPA patients, 14 newly diagnosed allergic asthma patients and 14 healthy volunteers included as controls. Cytokine levels were measured. Total and specific IgE levels of anti-IgE monoclonal antibody treated patients, serum high-sensitivity C-reactive protein (hsCRP) levels, FEV1/FVC rates and asthma control test (ACT) were measured for the clinical follow-up.. We observed that SPA patients presented increasing levels of IL-8, IL-10, TGF-β and GCSF during the anti-IgE treatment in period of sampling times at 4 months and 18 months. However this increase was not correlated neither with serum hsCRP levels nor FEV1/FVC rates.. Our study gives a different perspective for the SPA and anti-IgE immunotherapy efficacy at the cell cytokine-linked step.

Topics: Adolescent; Adult; Antibodies, Anti-Idiotypic; Asthma; C-Reactive Protein; Cytokines; Female; Forced Expiratory Volume; Granulocyte Colony-Stimulating Factor; Humans; Interleukin-10; Interleukin-8; Male; Middle Aged; Transforming Growth Factor beta

Plasmin, the activated protease product of plasminogen, is involved in collagen remodeling, and is strongly implicated in asthma pathophysiology by recent genome-wide association studies. This study examines plasminogen "activation" by airway smooth muscle cells, and its regulation in a fibrotic environment created by culture on type I collagen and incubation with transforming growth factor (TGF)-β. Urokinase plasminogen activator (uPA) activity was detected in the supernatants of human airway smooth muscle cell cultures maintained in serum-free conditions. Incubation with plasminogen (1.5-50.0 μg/ml, 24 h) increased plasmin activity in a concentration-dependent manner (P < 0.001). uPA activity was higher in cultures maintained on fibrillar type I collagen substrata than in those on plastic, as was plasmin activity after incubation with plasminogen (20 μg/ml). Pretreatment with TGF-β (100 pM) for 18 hours inhibited plasminogen activation by airway smooth muscle cells maintained on plastic, but not on collagen. TGF-β stimulated an increase in the level of uPA mRNA in airway smooth muscle cells grown on collagen, but not on plastic. Reducing the levels of β1-integrin collagen receptor, using interference RNA, attenuated plasmin formation by airway smooth muscle cells grown on collagen, and restored the inhibitory effect of TGF-β. This study shows that airway smooth muscle activation of plasminogen by uPA is accelerated in a collagen-rich environment in which the inhibitory effect of TGF-β is attenuated in association with greater uPA expression induced via β1-integrin signaling. These findings suggest that the plasminogen-activation system involving uPA has the potential to contribute to airway wall remodeling in asthma.

Topics: Asthma; Caseins; Collagen Type I; Fibrinolysin; Gene Expression Regulation; Humans; Integrin beta1; Models, Biological; Muscle, Smooth; Plasminogen; RNA, Messenger; Signal Transduction; Time Factors; Transforming Growth Factor beta; Urokinase-Type Plasminogen Activator

Intranasal administration of high amount of allergen was shown to induce tolerance and to reverse the allergic phenotype. However, mechanisms of tolerance induction via the mucosal route are still unclear.. To characterize the therapeutic effects of intranasal application of ovalbumin (OVA) in a mouse model of bronchial inflammation as well as the cellular and molecular mechanisms leading to protection upon re-exposure to allergen.. After induction of bronchial inflammation, mice were treated intranasally with OVA and re-exposed to OVA aerosols 10 days later. Bronchoalveolar lavage fluid (BALF), T cell proliferation and cytokine secretion were examined. The respective role of CD4(+)CD25(+) and CD4(+)CD25(-) T cells in the induction of tolerance was analysed.. Intranasal treatment with OVA drastically reduced inflammatory cell recruitment into BALF and bronchial hyperresponsiveness upon re-exposure to allergen. Both OVA- specific-proliferation of T cells, T(h)1 and T(h)2 cytokine production from lung and bronchial lymph nodes were inhibited. Transfer of CD4(+)CD25(-) T cells, which strongly expressed membrane-bound transforming growth factor β (mTGFβ), from tolerized mice protected asthmatic recipient mice from subsequent aerosol challenges. The presence of CD4(+)CD25(+)(Foxp3(+)) T cells during the process of tolerization was indispensable to CD4(+)CD25(-) T cells to acquire regulatory properties. Whereas the presence of IL-10 appeared dispensable in this model, the suppression of CD4(+)CD25(-)mTGFβ(+) T cells in transfer experiments significantly impaired the down-regulation of airways inflammation.. Nasal application of OVA in established asthma led to the induction of CD4(+)CD25(-)mTGFβ(+) T cells with regulatory properties, able to confer protection upon allergen re-exposure.

Topics: Administration, Intranasal; Allergens; Animals; Asthma; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Cell Proliferation; Cytokines; Desensitization, Immunologic; Drug Evaluation, Preclinical; Immune Tolerance; Interleukin-2 Receptor alpha Subunit; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes, Regulatory; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

S100A8 is implicated in the pathogenesis of inflammatory diseases. S100A8 is upregulated in macrophages by Toll-like receptors (TLR)-3, 4, and 9 agonists in an IL-10-dependent manner, and by corticosteroids in vitro and in vivo, and scavenges oxidants generated by activated phagocytes. Because if its elevated expression in various lung disorders, we asked whether S100A8 was protective in allergic inflammation. S100A8, but not Cys(41)-Ala S100A8, in which the single reactive Cys residue was replaced by Ala, reduced mast cell (MC) degranulation and production of particular cytokines (IL-6, IL-4, and granulocyte macrophage colony-stimulating factor) in response to IgE-crosslinking in vitro, likely by inhibiting intracellular reactive oxygen species production, thereby reducing downstream linker for activation of T cells and extracellular signal regulated kinase/mitogen-activated protein kinase phosphorylation. In lungs of mice with acute asthma, S100A8, but not Cys(41)-Ala S100A8, reduced MC degranulation, production of eosinophil chemoattractants (IL-5, eotaxin, and monocyte chemoattractant protein-1), and eosinophil infiltration. Suppression of IL-6 and IL-13 could have contributed to reduced mucus production seen in lungs of S100A8-treated mice. IgE production was unaffected. In asthma, there is an imbalance of anti-oxidant systems that are generally protective. Our results strongly support a protective role for S100A8 in allergic inflammation by modulating MC activation and eosinophil recruitment, and by scavenging oxidants generated by activated leukocytes, in processes reliant on its thiol-scavenging capacity.

Topics: Animals; Apoptosis; Asthma; Calgranulin A; Cell Movement; Cells, Cultured; Chemokine CCL2; Eosinophils; Female; Granulocyte-Macrophage Colony-Stimulating Factor; Interleukin-10; Interleukin-13; Interleukin-4; Interleukin-5; Interleukin-6; Mast Cells; Mice; Mice, Inbred BALB C; Phosphorylation; Polymerase Chain Reaction; Reactive Oxygen Species; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Although a similar prevalence of smoking is evident among patients with asthma and the general population, little is known about the impact of cigarette smoke on the immune inflammatory processes elicited by common environmental allergens. We investigated the impact of exposure to cigarette smoke on house dust mite (HDM)-induced allergic airway inflammation and its consequences for tissue remodeling and lung physiology in mice. BALB/c mice received intranasal HDMs daily, 5 days per week, for 3 weeks to establish chronic airway inflammation. Subsequently, mice were concurrently exposed to HDMs plus cigarette smoke, 5 days per week, for 2 weeks (HDMs + smoke). We observed significantly attenuated eosinophilia in the bronchoalveolar lavage of mice exposed to HDMs + smoke, compared with animals exposed only to HDMs. A similar activation of CD4 T cells and expression of IL-5, IL-13, and transforming growth factor-β was observed between HDM-treated and HDM + smoke-treated animals. Consistent with an effect on eosinophil trafficking, HDMs + smoke exposure attenuated the HDM-induced expression of eotaxin-1 and vascular cell adhesion molecule-1, whereas the survival of eosinophils and the numbers of blood eosinophils were not affected. Exposure to cigarette smoke also reduced the activation of B cells and the concentrations of serum IgE. Although the production of mucus decreased, collagen deposition significantly increased in animals exposed to HDMs + smoke, compared with animals exposed only to HDMs. Although airway resistance was unaffected, tissue resistance was significantly decreased in mice exposed to HDMs + smoke. Our findings demonstrate that cigarette smoke affects eosinophil migration without affecting airway resistance or modifying Th2 cell adaptive immunity in a murine model of HDM-induced asthma.

Topics: Airway Remodeling; Airway Resistance; Allergens; Animals; Asthma; B-Lymphocytes; Bronchial Hyperreactivity; Chemokine CCL11; Dendritic Cells; Disease Models, Animal; Female; Interleukin-13; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Pulmonary Eosinophilia; Pyroglyphidae; Smoking; T-Lymphocytes; Time Factors; Transforming Growth Factor beta; Vascular Cell Adhesion Molecule-1

The evolution of allergic asthma is tightly controlled by effector and regulatory cells, as well as cytokines such as IL-10 and/or TGF-β, and it is widely acknowledged that environmental exposure to allergens and infectious agents can influence these processes. In this context, the recognition of pathogen-associated motifs, which trigger TLR activation pathways, plays a critical role with important consequences for disease progression and outcome. We addressed the question whether the TLR7 ligand resiquimod (R848), which has been shown to be protective in several experimental allergic asthma protocols, can also suppress typical asthma symptoms once the disease is established. To this end, we used an OVA-induced experimental model of murine allergic asthma in which R848 was injected after a series of challenges with aerosolized OVA. We found that the treatment attenuated allergic symptoms through a mechanism that required Tregs, as assessed by the expansion of this population in the lungs of mice having received R848, and the loss of R848-mediated suppression of allergic responses after in vivo Treg depletion. IL-10 provided only a minor contribution to this suppressive effect that was largely mediated through a TGF-β-dependent pathway, a finding that opens new therapeutic opportunities for the pharmacological targeting of Tregs.

Topics: Animals; Asthma; Disease Models, Animal; Imidazoles; Interleukin-10; Lung; Male; Membrane Glycoproteins; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; T-Lymphocytes, Regulatory; Toll-Like Receptor 7; Transforming Growth Factor beta

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

Individuals with chronic asthma show a progressive decline in lung function that is thought to be due to structural remodeling of the airways characterized by subepithelial fibrosis and smooth muscle hyperplasia. Here we show that the tumor necrosis factor (TNF) family member LIGHT is expressed on lung inflammatory cells after allergen exposure. Pharmacological inhibition of LIGHT using a fusion protein between the IgG Fc domain and lymphotoxin β receptor (LTβR) reduces lung fibrosis, smooth muscle hyperplasia and airway hyperresponsiveness in mouse models of chronic asthma, despite having little effect on airway eosinophilia. LIGHT-deficient mice also show a similar impairment in fibrosis and smooth muscle accumulation. Blockade of LIGHT suppresses expression of lung transforming growth factor-β (TGF-β) and interleukin-13 (IL-13), cytokines implicated in remodeling in humans, whereas exogenous administration of LIGHT to the airways induces fibrosis and smooth muscle hyperplasia, Thus, LIGHT may be targeted to prevent asthma-related airway remodeling.

Topics: Airway Remodeling; Animals; Asthma; Disease Models, Animal; Humans; Inflammation Mediators; Interleukin-13; Lung; Lymphotoxin alpha1, beta2 Heterotrimer; Mice; Mice, Knockout; Signal Transduction; Transforming Growth Factor beta; Tumor Necrosis Factor Ligand Superfamily Member 14

To investigate the mechanisms of enhanced airway deposition of subepithelial collagen in asthma and its sensitivity to drug therapy with combination of an inhaled glucocorticosteroid (GC) and a long-acting β(2)-agonist (LABA), a cell model system involving bronchial fibroblasts derived from biopsies from patients with stable mild-to-moderate asthma has been used. To mimic unstable conditions and severe asthma, fibroblasts were stimulated ex vivo with TGFβ1. Primary fibroblasts established from central bronchial biopsies from 8 asthmatic patients were incubated for 24 h with 0.4% serum or TGFβ1 (10 ng/ml) with/without the GC budesonide (BUD; 10 nM) and/or the LABA formoterol (FORM; 0.1 nM). Procollagen peptide I (PICP), metalloproteinase (MMP)-1 and tissue inhibitor of MMPs (TIMP-1) were determined in culture media using ELISA while the activity of MMP-2, -3, -9 by zymography. Metabolically labeled proteoglycans, biglycan and decorin, associated with collagen fibrillation/deposition, were separated using chromatography and SDS-PAGE. The levels of PICP and biglycan were increased 2-fold by TGFβ1 (p < 0.05). The BUD and FORM combination reduced the PICP increase by 58% (p < 0.01) and the biglycan by 36% (p < 0.05) while each drug alone had no effect. Decorin levels were reduced by TGFβ1 in fibroblasts of most patients; BUD alone and BUD and FORM completely counteracted this decrease. MMPs and TIMP-1 were not affected by TGFβ1 or the drugs. These results suggest that BUD and FORM combination therapy, without affecting metalloproteolytic balance, has a potential to counteract enhanced collagen production by bronchial fibroblasts in asthma and to normalize the production of small proteoglycans which may affect collagen fibrillation and deposition.

Topics: Administration, Inhalation; Adult; Asthma; Bronchi; Bronchodilator Agents; Budesonide; Chromatography; Collagen; Electrophoresis, Polyacrylamide Gel; Ethanolamines; Female; Fibroblasts; Formoterol Fumarate; Humans; Male; Middle Aged; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta

Vascular remodeling is an important feature in asthma pathophysiology. Although investigations suggested that nitric oxide (NO) is involved in lung remodeling, little evidence established the role of inducible NO synthase (iNOS) isoform in bronchial vascular remodeling. The authors investigated if iNOS contribute to bronchial vascular remodeling induced by chronic allergic pulmonary inflammation. Guinea pigs were submitted to ovalbumin exposures with increasing doses (1∼5 mg/mL) for 4 weeks. Animals received 1400W (iNOS-specific inhibitor) treatment for 4 days beginning at 7th inhalation. Seventy-two hours after the 7th inhalation, animals were anesthetized, mechanical ventilated, exhaled NO was collected, and lungs were removed and submitted to picrosirius and resorcin-fuchsin stains and to immunohistochemistry for matrix metalloproteinase-9 (MMP-9), tissue inhibitor of metalloproteinase-1 (TIMP-1), and transforming growth factor-β (TGF-β). Collagen and elastic fiber deposition as well as MMP-9, TIMP-1, and TGF-β expression were increase in bronchial vascular wall in ovalbumin-exposed animals. The iNOS inhibition reduced all parameters studied. In this model, iNOS inhibition reduced the bronchial vascular extracellular remodeling, particularly controlling the collagen and elastic fibers deposition in pulmonary vessels. This effect can be associated to a reduction on TGF-β and on metalloproteinase-9/TIMP-1 vascular expression. It reveals new therapeutic strategies and some possible mechanism related to specific iNOS inhibition to control vascular remodeling.

Topics: Administration, Inhalation; Amidines; Animals; Asthma; Benzylamines; Blood Vessels; Bronchi; Collagen; Elastic Tissue; Enzyme Inhibitors; Extracellular Matrix; Guinea Pigs; Male; Matrix Metalloproteinase 9; Nitric Oxide; Nitric Oxide Synthase Type II; Ovalbumin; Pneumonia; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta

To explore the effects of serum of asthmatic patients, dexamethasone, interleukin-4 (IL-4), interferon-gamma (IFN-γ) and transforming growth factor-β (TGF-β) on the expression of interleukin-22 receptor 1 (IL-22R1) mRNA and protein in HASMCs in vitro.. IL-22R1 mRNA and protein expressions in HASMCs treated with different stimulating agents were measured by real-time PCR and Western blotting, respectively.. IL-22R1 mRNA and protein expressions in HASMCs were significantly increased after stimulation by serum from asthmatic patients, but decreased after co-stimulation with dexamethasone. IL-22R1 mRNA and protein expressions in the cells both increased after stimulation by IL-4, IFN-γ and TGF-β.. IL-22R1 in HASMCs might be involved in the pathogenesis of asthma, and the therapeutic effect of dexamethasone on asthma is mediated, at least partially, by IL-22R1. The effects of IFN-γ, IL-4, and TGF-β on asthma may also be attributed to their actions on HASMCs.

Topics: Asthma; Cell Line; Humans; Interferon-gamma; Interleukin-4; Myocytes, Smooth Muscle; Receptors, Interleukin; RNA, Messenger; Transforming Growth Factor beta

Aberrant airway smooth muscle cell (ASMC) function and overexpression of transforming growth factor (TGF)-β, which modulates ASMC proliferative and inflammatory function and induces oxidant release, are features of asthma. Nuclear factor E2-related factor 2 (Nrf2) activates antioxidant genes conferring protection against oxidative stress.. To determine the role of Nrf2 in ASMCs and its modulation by TGF-β, and compare Nrf2 activity in ASMCs from subjects with severe and nonsevere asthma and healthy subjects.. ASMCs were cultured from airways of subjects without asthma, and from airway biopsies from patients with severe and nonsevere asthma. We studied Nrf2 activation on antioxidant gene expression and proliferation, the effect of TGF-β on Nrf2 transcriptional activity, and the impact of Nrf2 activation on TGF-β–mediated proliferation and IL-6 release. Nrf2–antioxidant response elements binding and Nrf2-dependent antioxidant gene expression was determined in asthmatic ASMCs.. Activation of Nrf2 led to up-regulation of the antioxidant genes heme oxygenase (HO)-1, NAD(P)H:quinone oxidoreductase, and manganese superoxide dismutase, and a reduction in proliferation. TGF-β reduced Nrf2-mediated antioxidant gene transcription through induction of activating transcription factor-3 expression. Nrf2 activation attenuated TGF-β–mediated reduction in HO-1,ASMC proliferation, and IL-6 release. Nrf2–antioxidant response elements binding was reduced in ASMCs from patients with severe asthma compared with ASMCs from patients with nonsevere asthma and normal subjects. HO-1 expression was reduced in ASMCs from patients with both nonsevere and severe asthma compared with healthy subjects.. Nrf2 regulates antioxidant responses and proliferation in ASMCs and is inactivated by TGF-β. Nrf2 reduction may underlie compromised antioxidant protection and aberrant ASM function in asthma.

Topics: Adenoviridae; Antioxidants; Asthma; Cell Proliferation; Gene Expression; Gene Expression Regulation; Genetic Vectors; Humans; Isothiocyanates; Muscle, Smooth; NF-E2 Transcription Factor; RNA, Small Interfering; Sulfoxides; Thiocyanates; Transfection; Transforming Growth Factor beta

Severe asthma is associated with airway remodeling, characterized by structural changes including increased smooth muscle mass and matrix deposition in the airway, leading to deteriorating lung function. TGF-β is a pleiotropic cytokine leading to increased synthesis of matrix molecules by human airway smooth muscle (HASM) cells and is implicated in asthmatic airway remodeling. TGF-β is synthesized as a latent complex, sequestered in the extracellular matrix, and requires activation for functionality. Activation of latent TGF-β is the rate-limiting step in its bioavailability. This study investigated the effect of the contraction agonists LPA and methacholine on TGF-β activation by HASM cells and its role in the development of asthmatic airway remodeling. The data presented show that LPA and methacholine induced TGF-β activation by HASM cells via the integrin αvβ5. Our findings highlight the importance of the β5 cytoplasmic domain because a polymorphism in the β5 subunit rendered the integrin unable to activate TGF-β. To our knowledge, this is the first description of a biologically relevant integrin that is unable to activate TGF-β. These data demonstrate that murine airway smooth muscle cells express αvβ5 integrins and activate TGF-β. Finally, these data show that inhibition, or genetic loss, of αvβ5 reduces allergen-induced increases in airway smooth muscle thickness in two models of asthma. These data highlight a mechanism of TGF-β activation in asthma and support the hypothesis that bronchoconstriction promotes airway remodeling via integrin mediated TGF-β activation.

Topics: Airway Remodeling; Animals; Asthma; Blotting, Western; Cell Line; Cell Separation; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Humans; Immunohistochemistry; Immunoprecipitation; Mice; Myocytes, Smooth Muscle; Real-Time Polymerase Chain Reaction; Receptors, Vitronectin; Respiratory System; Reverse Transcriptase Polymerase Chain Reaction; Transfection; Transforming Growth Factor beta

Activin A is a member of the TGF-beta superfamily and plays a role in allergic inflammation and asthma pathogenesis. Recent evidence suggests that activin A regulates proinflammatory cytokine production and is regulated by inflammatory mediators. In a murine model of acute allergic airway inflammation, we observed previously that increased activin A concentrations in bronchoalveolar lavage (BAL) fluid coincide with Th2 cytokine production in lung-draining lymph nodes and pronounced mucus metaplasia in bronchial epithelium. We therefore hypothesized that IL-13, the key cytokine for mucus production, regulates activin A secretion into BAL fluid in experimental asthma. IL-13 increased BAL fluid activin A concentrations in naive mice and dose dependently induced activin A secretion from cultured human airway epithelium. A key role for IL-13 in the secretion of activin A into the BAL fluid during allergic airway inflammation was confirmed in IL-13-deficient mice. Eosinophils were not involved in this response because there was no difference in BAL fluid activin A concentrations between wild-type and eosinophil-deficient mice. Our data highlight an important role for IL-13 in the regulation of activin A intraepithelially and in BAL fluid in naive mice and during allergic airway inflammation. Given the immunomodulatory and fibrogenic effects of activin A, our findings suggest an important role for IL-13 regulation of activin A in asthma pathogenesis.

Topics: Activin Receptors; Activins; Animals; Asthma; Bronchoalveolar Lavage Fluid; Cells, Cultured; Disease Models, Animal; Eosinophils; Epithelial Cells; Female; Humans; Inhibin-beta Subunits; Interleukin-13; Interleukin-5; Metaplasia; Mice; Mice, Inbred BALB C; Mice, Knockout; Ovalbumin; Pneumonia; Recombinant Proteins; Respiratory Mucosa; Signal Transduction; Time Factors; Transforming Growth Factor beta

We examined the contribution of p70 ribosomal S6 kinase (p70S6K) to airway smooth muscle hypertrophy, a structural change found in asthma. In human airway smooth muscle cells, transforming growth factor (TGF)-beta, endothelin-1, and cardiotrophin-1 each induced phosphorylation of p70S6K and ribosomal protein S6 while increasing cell size, total protein synthesis, and relative protein abundance of alpha-smooth muscle actin and SM22. Transfection of myocytes with siRNA against either p70S6K or S6, or infection with retrovirus encoding a kinase-dead p70S6K, reduced cell size and protein synthesis but had no effect on contractile protein expression per mg total protein. Infection with a retrovirus encoding a constitutively active, rapamycin-resistant (RR) p70S6K increased cell size but not contractile protein expression. siRNA against S6 decreased cell size in myocytes expressing RR p70S6K. Finally, TGF-beta treatment, but not RR p70S6K expression, increased KCl-induced fractional shortening. Together, these data suggest that p70S6K activation is both required and sufficient for airway smooth muscle cell size enlargement but not contractile protein expression. Further, ribosomal protein S6 is required for p70S6K-mediated cell enlargement. Finally, we have shown for the first time in a functional cell system that p70S6K-mediated myocyte enlargement alone, without preferential contractile protein expression, is insufficient for increased cell shortening.

Topics: Airway Remodeling; Animals; Asthma; Cell Enlargement; Cells, Cultured; Contractile Proteins; Cytokines; Disease Models, Animal; Endothelin-1; Enzyme Activation; Humans; Hypertrophy; Lung; Mice; Mice, Inbred BALB C; Microfilament Proteins; Muscle Contraction; Muscle Proteins; Muscle, Smooth; Mutation; Myocytes, Smooth Muscle; Ovalbumin; Phenotype; Phosphorylation; Potassium Chloride; Ribosomal Protein S6; Ribosomal Protein S6 Kinases, 70-kDa; RNA Interference; Transduction, Genetic; Transforming Growth Factor beta

We have reported previously that moderate intensity aerobic exercise training attenuates airway inflammation in a murine asthma model. Recent studies implicate regulatory T (Treg) cells in decreasing asthma-related airway inflammation; as such, the current study examined the effect of exercise on Treg cell function in a murine asthma model. Mice were sensitized with ovalbumin (OVA) prior to the start of exercise training at a moderate intensity 3x/week for 4weeks; exercise was performed as treadmill running (13.5m/min, 0% grade). Mice were OVA challenged repeatedly throughout the exercise protocol. At protocol completion, mice were analyzed for changes in the number and suppressive function of CD4(+)CD25(+)Foxp3(+) cells isolated from lungs, mediastinal lymph nodes, and spleens. Results show that exercise increased significantly the number of Foxp3(+) cells within the lungs and mediastinal lymph nodes, but not the spleens, of OVA-treated mice as compared with sedentary controls. Exercise also enhanced the suppression function of CD4(+)CD25(+)Foxp3(+) Treg cells derived from OVA-treated mice as compared with sedentary controls. Specifically, Treg cells from exercised, OVA-treated mice more effectively suppressed CD4(+)CD25(-) cell proliferation and Th2 cytokine production in vitro. Enhanced suppression was associated with increased protein levels of TGF-beta and lesser amounts of IL-10 and IL-17; however, blocking TGF-beta had no effect on suppressive functions. These data demonstrate that exercise-mediated increases in Treg cell function may play a role in the attenuation of airway inflammation. Further, these results indicate that moderate intensity aerobic exercise training may alter the Treg cell function within the asthmatic airway.

Topics: Aerobiosis; Animals; Asthma; Bronchoalveolar Lavage Fluid; CD4 Antigens; Cell Count; Female; Flow Cytometry; Forkhead Transcription Factors; Interleukin-10; Interleukin-17; Interleukin-2 Receptor alpha Subunit; Lung; Lymph Nodes; Mice; Mice, Inbred BALB C; Ovalbumin; Physical Conditioning, Animal; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

As passive environmental tobacco smoke (ETS) exposure in nonsmokers can increase both asthma symptoms and the frequency of asthma exacerbations, we utilized a mouse model, in which ovalbumin (OVA) + ETS induce significantly increased levels of eosinophilic airway inflammation and remodeling compared to either stimulus alone, to determine whether a Toll-like receptor-9 (TLR-9) agonist could reduce levels of airway inflammation, airway remodeling and airway hyperreactivity (AHR).. Mice treated with or without a TLR-9 agonist were sensitized to OVA and challenged with OVA + ETS for 1 month. AHR to methacholine was assessed in intubated and ventilated mice. Lung Th2 cytokines and TGF-beta(1) were measured by ELISA. Lungs were processed for histology and immunohistology to quantify eosinophils, mucus, peribronchial fibrosis and smooth muscle changes using image analysis.. Administration of a TLR-9 agonist to mice coexposed to chronic ETS and chronic OVA allergen significantly reduced levels of eosinophilic airway inflammation, mucus production, peribronchial fibrosis, the thickness of the peribronchial smooth muscle layer, and AHR. The reduced airway remodeling in mice treated with the TLR-9 agonist was associated with significantly reduced numbers of peribronchial MBP+ and peribronchial TGF-beta(1)+ cells, and with significantly reduced levels of lung Th2 cytokines [interleukin-5 and interleukin-13] and TGF-beta(1).. These studies demonstrate that TLR-9-based therapies inhibit airway inflammation, remodeling and AHR in mice coexposed to ETS and allergen who exhibit enhanced airway inflammation and remodeling.

Topics: Air Pollutants; Airway Remodeling; Allergens; Animals; Asthma; Cell Movement; Eosinophils; Interleukin-13; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Oligodeoxyribonucleotides; Ovalbumin; Pulmonary Fibrosis; Smoking; Toll-Like Receptor 9; Transforming Growth Factor beta

In the present study the effects of bradykinin receptor antagonists were investigated in a murine model of asthma using BALB/c mice immunized with ovalbumin/alum and challenged twice with aerosolized ovalbumin. Twenty four hours later eosinophil proliferation in the bone marrow, activation (lipid bodies formation), migration to lung parenchyma and airways and the contents of the pro-angiogenic and pro-fibrotic cytokines TGF-beta and VEGF were determined. The antagonists of the constitutive B(2) (HOE 140) and inducible B(1) (R954) receptors were administered intraperitoneally 30min before each challenge. In sensitized mice, the antigen challenge induced eosinophil proliferation in the bone marrow, their migration into the lungs and increased the number of lipid bodies in these cells. These events were reduced by treatment of the mice with the B(1) receptor antagonist. The B(2) antagonist increased the number of eosinophils and lipid bodies in the airways without affecting eosinophil counts in the other compartments. After challenge the airway levels of VEGF and TGF-beta significantly increased and the B(1) receptor antagonist caused a further increase. By immunohistochemistry techniques TGF-beta was found to be expressed in the muscular layer of small blood vessels and VEGF in bronchial epithelial cells. The B(1) receptors were expressed in the endothelial cells. These results showed that in a murine model of asthma the B(1) receptor antagonist has an inhibitory effect on eosinophils in selected compartments and increases the production of cytokines involved in tissue repair. It remains to be determined whether this effects of the B(1) antagonist would modify the progression of the allergic inflammation towards resolution or rather towards fibrosis.

Topics: Analysis of Variance; Animals; Asthma; Bradykinin B1 Receptor Antagonists; Bradykinin B2 Receptor Antagonists; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Cell Count; Cell Movement; Cell Proliferation; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Eosinophils; Immunohistochemistry; Lung; Mice; Receptor, Bradykinin B1; Receptor, Bradykinin B2; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Once the umbilical cord has been cut, immunologists have often looked at the neonate as an entity that develops on its own. For years, breast milk was considered mainly as a source of nutrients for the developing child. The extensive observations that breastfeeding affords protection toward infectious diseases and could reduce by more than the half the mortality rate because of common infections have added another key role to breastfeeding. This protection relies in great part on the passive transfer through breast milk of high amounts of microbe-specific immunoglobulins that compensate for the deficiency of immunoglobulins synthesis during the first year of life. Here, we will present and discuss our data showing how breast milk can actively shape the immune response of the progeny, particularly in the context of allergic disease. Indeed, our data obtained in a mouse model suggest that the protection attributed to breastfeeding toward asthma development might rely on immune tolerance induction. For this to occur, the mother mice needed to be exposed to the allergen by aerosol or oral route during the lactation period, which resulted into the transfer of the allergen to breast milk. The presence of the allergen together with transforming growth factor-beta in breast milk was necessary and sufficient to induce the development of regulatory T lymphocytes in the progeny and their protection from asthma development. If confirmed in human beings, this study may suggest new strategies for asthma prevention such as deliberate exposure of mother to allergens during breastfeeding and qualitative modification of artificial milks.

Topics: Allergens; Animals; Animals, Suckling; Asthma; Female; Immune Tolerance; Immunity, Maternally-Acquired; Lactation; Maternal Exposure; Mice; Milk; Ovalbumin; Signal Transduction; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Eotaxin plays a central role in the development of allergic disease, including atopic dermatitis, asthma, and nasal allergy. Interleukin (IL)-4 induces eotaxin production in normal human dermal fibroblasts. On the other hands, Transforming growth factor-beta (TGF-beta), a multifunctional regulatory cytokine, affects many biological functions, including fibroblast growth and differentiation and Th2 cytokine regulation. In this study, we investigated the effect of TGF-beta on IL-4-induced eotaxin production by normal human fibroblasts, as well as the effect of suplatast tosilate, an antiallergic drug that selectively inhibits Th2 cytokine production. Dermal fibroblast treatment with IL-4 and TGF-beta for 24 h increased eotaxin production and expression of eotaxin mRNA, as measured by enzyme-linked immunosorbent assay (ELISA) and reverse-transcriptase polymerase chain reaction (RT-PCR), respectively. TGF-beta synergistically up-regulated eotaxin production and eotaxin mRNA expression when stimulated with IL-4. Suplatast tosilate dose-dependently inhibited eotaxin production induced by IL-4 or IL-4 plus TGF-beta. These results suggest that TGF-beta may regulate skin allergic inflammation by up-regulating eotaxin production in dermal fibroblasts. Suplatast tosilate might suppress this inflammation by inhibiting eotaxin production.

Topics: Anti-Allergic Agents; Arylsulfonates; Asthma; Cells, Cultured; Chemokine CCL11; Chemotactic Factors, Eosinophil; Dose-Response Relationship, Drug; Drug Synergism; Enzyme-Linked Immunosorbent Assay; Fibroblasts; Gene Expression Regulation; Humans; Interleukin-4; Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Skin; Sulfonium Compounds; Transforming Growth Factor beta; Up-Regulation

Chemokines ligands of CCR3 including eotaxin/CC chemokine ligand 11 (CCL11) may contribute to the pathogenesis of asthma. These chemokines and a growth factor (TGF-beta) may be involved in the process of airway remodelling.. We analysed the effects of TGF-beta on the expression of CCR3 ligands in human airway smooth muscle (HASM) cells and investigated the mechanisms.. HASM cells were cultured and treated with TGF-beta and Th2 cytokines IL-4 or IL-13. Expression of mRNA was analysed by real-time PCR. Secretion of CCL11 into the culture medium was analysed by ELISA. Transcriptional regulation of CCL11 was analysed by luciferase assay using CCL11 promoter-luciferase reporter plasmids.. IL-4 or IL-13 significantly up-regulated the expression of mRNAs for CCL11 and CCL26. TGF-beta alone did not increase the expression of chemokine mRNAs, but enhanced the induction of only CCL11 by IL-4 or IL-13 among CCR3 ligands. Activity of the CCL11 promoter was stimulated by IL-4, and this activity was enhanced by TGF-beta. Activation by IL-4 or IL-4 plus TGF-beta was lost by mutation of the binding site for signal transducers and activators of transcription-6 (STAT6) in the promoter. Cooperative activation by IL-4 and TGF-beta was inhibited by mutation of the binding site for nuclear factor-kappaB (NF-kappaB) in the promoter. Pretreatment with an inhibitor of NF-kappaB and glucocorticoid fluticasone propionate significantly inhibited the expression of CCL11 mRNA induced by IL-4 plus TGF-beta, indicating the importance of NF-kappaB in the cooperative activation of CCL11 transcription by TGF-beta and IL-4.. These results indicate that Th2 cytokines and TGF-beta may contribute to the pathogenesis of asthma by stimulating expression of CCL11. The transcription factors STAT6 and NF-kappaB may play pivotal roles in this process.

Topics: Asthma; Binding Sites; Cell Line; Chemokine CCL11; Gene Expression Regulation; Humans; Interleukin-13; Interleukin-4; Muscle, Smooth; NF-kappa B; Promoter Regions, Genetic; Protein Binding; Recombinant Proteins; Th2 Cells; Transcription, Genetic; Transforming Growth Factor beta; Up-Regulation

To observe the expression of CD4(+) CD25(+) Foxp3(+) Treg, IL-10 and TGF-beta in lymph and blood, as well as the effect of dexamethasone on it and investigate the relationship between them.. Lymph and blood samples of 0 h, 24 h, 48 h after the last challenge were collected from the rat model of asthma. The percentage of CD4(+) CD25(+) Foxp3(+) Treg were detected by flow cytometer (FCM), while the levels of IL-10 and TGF-beta were determined by ELISA.. The percentage of CD4(+) CD25(+) Foxp3(+) Treg in the blood and the levels of IL-10 and TGF-beta in the plasma in cases of asthma group were significantly lower than that of the control and therapy group at different time points (P<0.05); The percentage of CD4(+) CD25(+) Foxp3(+) Treg and the level of IL-10 in lymph were significantly higher than that of blood (P<0.05), however, the level of TGF-beta in lymph were significantly lower than that of blood (P<0.05); There were no significant differences which the percentage of CD4(+) CD25(+) Foxp3(+) Treg in the blood and the levels of IL-10 and TGF-beta in the plasma between control group and therapy group (P>0.05).. There is a disbalance both quantity and function of CD4(+) CD25(+) Foxp3(+) in the lymph of bronchial asthmatic rat, and the percentage of CD4(+) CD25(+) Foxp3(+) Treg in lymph are significantly higher than that of blood, which dexamethasone may effectively increase the percentage of CD4(+) CD25(+) Foxp3(+) Treg in bronchial asthmatic rat.

Topics: Animals; Asthma; CD4-Positive T-Lymphocytes; Cells, Cultured; Dexamethasone; Disease Models, Animal; Female; Humans; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Lymph; Male; Rats; Rats, Sprague-Dawley; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Bone marrow stromal cells [BMSCs; also known as mesenchymal stem cells (MSCs)] effectively suppress inflammatory responses in acute graft-versus-host disease in humans and in a number of disease models in mice. Many of the studies concluded that BMSC-driven immunomodulation is mediated by the suppression of proinflammatory Th1 responses while rebalancing the Th1/Th2 ratio toward Th2. In this study, using a ragweed induced mouse asthma model, we studied if BMSCs could be beneficial in an allergic, Th2-dominant environment. When BMSCs were injected i.v. at the time of the antigen challenge, they protected the animals from the majority of asthma-specific pathological changes, including inhibition of eosinophil infiltration and excess mucus production in the lung, decreased levels of Th2 cytokines (IL-4, IL-5, and IL-13) in bronchial lavage, and lowered serum levels of Th2 immunoglobulins (IgG1 and IgE). To explore the mechanism of the effect we used BMSCs isolated from a variety of knockout mice, performed in vivo blocking of cytokines and studied the effect of asthmatic serum and bronchoalveolar lavage from ragweed challenged animals on the BMSCs in vitro. Our results suggest that IL-4 and/or IL-13 activate the STAT6 pathway in the BMSCs resulting in an increase of their TGF-beta production, which seems to mediate the beneficial effect, either alone, or together with regulatory T cells, some of which might be recruited by the BMSCs. These data suggest that, in addition to focusing on graft-versus-host disease and autoimmune diseases, allergic conditions--specifically therapy resistant asthma--might also be a likely target of the recently discovered cellular therapy approach using BMSCs.

Topics: Ambrosia; Animals; Asthma; Bronchoalveolar Lavage Fluid; Cytokines; Disease Models, Animal; Humans; Immunosuppression Therapy; In Vitro Techniques; Lung; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Th2 Cells; Transforming Growth Factor beta; Transplantation, Homologous; Transplantation, Isogeneic

The TGF-beta family of mediators are thought to play important roles in the regulation of inflammation and airway remodelling in asthma. All three mammalian isoforms of TGF-beta, TGF-beta(1-3), are expressed in the airways and TGF-beta(1) and -beta(2) are increased in asthma. However, there is little information on the specific roles of individual TGF-beta isoforms. In this study we assess the roles of TGF-beta(1) and TGF-beta(2) in the regulation of allergen-induced airway inflammation and remodelling associated with asthma, using a validated murine model of ovalbumin sensitization and challenge, and isoform specific TGF-beta neutralising antibodies. Antibodies to both isoforms inhibited TGF-beta mediated Smad signalling. Anti-TGF-beta(1) and anti-TGF-beta(2) inhibited ovalbumin-induced sub-epithelial collagen deposition but anti-TGF-beta(1) also specifically regulated airway and fibroblast decorin deposition by TGF-beta(1). Neither antibody affected the allergen-induced increase in sub-epithelial fibroblast-like cells. Anti- TGF-beta(1) also specifically inhibited ovalbumin-induced increases in monocyte/macrophage recruitment. Whereas, both TGF-beta(1) and TGF-beta(2) were involved in regulating allergen-induced increases in eosinophil and lymphocyte numbers. These data show that TGF-beta(1) and TGF-beta(2) exhibit a combination of specific and shared roles in the regulation of allergen-induced airway inflammation and remodelling. They also provide evidence in support of the potential for therapeutic regulation of specific subsets of cells and extracellular matrix proteins associated with inflammation and remodelling in airway diseases such as asthma and COPD, as well as other fibroproliferative diseases.

Topics: Animals; Asthma; Bronchoalveolar Lavage Fluid; Collagen; Eosinophils; Extracellular Matrix; Image Processing, Computer-Assisted; Immunohistochemistry; Inflammation; Lung; Lymphocytes; Mice; Mice, Inbred C57BL; Protein Isoforms; Transforming Growth Factor beta

Upregulation of transforming growth factor-beta (TGF-beta) signaling is interrelated with the development of airway remodeling. In this study, we examined the role of two E3 ubiquitin ligases, Arkadia and Smurf2, which are critically required for TGF-beta signaling in airway remodeling. Rats were immunized with ovalbumin (OVA) and then challenged with an OVA aerosol. In in vitro experiments, normal human bronchial epithelial cells were stimulated with TGF-beta(1) with or without the preincubation of Arkadia/Smurf2 small interfering RNA (siRNA) or lactacystin (an inhibitor of proteasomal degradation). In the lungs of OVA-treated rats, a large number of inflammatory cells were present near the airways. An increased subepithelial collagen deposition was associated with high expression levels of Smad7, SnoN and Ski mRNAs, Arkadia, Smurf2, and TGF-beta type I receptor (TbetaRI), but low expression levels of Smad7, SnoN and Ski proteins. Smad7, SnoN and Ski interacted with both Arkadia and Smurf2 while TbetaRI only interacted with Smurf2 but not with Arkadia. In in vitro experiments, the inhibitory effect of TGF-beta(1) on the expression of Smad7, SnoN and Ski was reversed by Arkadia siRNA and lactacystin, whereas the stimulatory effect of TGF-beta(1) on the expression of TbetaRI protein and Smad7/SnoN/Ski mRNAs was not affected. In contrast, Smurf2 siRNA did not influence the effects of TGF-beta(1) on the expression of the above proteins. Our results suggest that Arkadia may contribute to the pathogenesis of airway remodeling through enhancing TGF-beta signaling by inducing the reduction of Smad7, SnoN and Ski proteins in OVA-sensitized and -challenged rats.

Topics: Animals; Asthma; Cells, Cultured; Epithelial Cells; Humans; Lung; Male; Nerve Tissue Proteins; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; Rats; Rats, Sprague-Dawley; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; RNA, Small Interfering; Smad7 Protein; Transcription Factors; Transforming Growth Factor beta; Ubiquitin-Protein Ligases

TGF-beta has been postulated to play an important role in the maintenance of epithelial homeostasis and the development of epithelium-derived cancers. However, most of previous studies are mainly focused on the function of TGF-beta in immune cells to the development of allergic asthma and how TGF-beta signaling in airway epithelium itself in allergic inflammation is largely unknown. Furthermore, the in vivo TGF-beta function specifically in the airway epithelium during lung cancer development has been largely elusive.. To evaluate the in vivo contribution of TGF-beta signaling in lung epithelium to the development of allergic disease and lung cancer, we generated a transgenic mouse model with Smad7, an intracellular inhibitor of TGF-beta signaling, constitutively expressed in mouse airway Clara cells using a mouse CC10 promoter. The mice were subjected to the development of OVA-induced allergic asthma and urethane-induced lung cancer. The Smad7 transgenic animals significantly protected from OVA-induced asthma, with reduced airway inflammation, airway mucus production, extracellular matrix deposition, and production of OVA-specific IgE. Further analysis of cytokine profiles in lung homogenates revealed that the Th2 cytokines including IL-4, IL-5 and IL-13, as well as other cytokines including IL-17, IL-1, IL-6, IP10, G-CSF, and GM-CSF were significantly reduced in the transgenic mice upon OVA induction. In contrast, the Smad7 transgenic animals had an increased incidence of lung carcinogenesis when subjected to urethane treatment.. These studies, therefore, demonstrate for the first time the in vivo function of TGF-beta signaling specifically in airway epithelium during the development of allergic asthma and lung cancer.

Topics: Animals; Asthma; Cytokines; Disease Models, Animal; Genetic Therapy; Inflammation; Lung Neoplasms; Mice; Mice, Transgenic; Ovalbumin; Respiratory Mucosa; Signal Transduction; Smad7 Protein; Th2 Cells; Transforming Growth Factor beta; Urethane

Genetic and epigenetic programming of T helper (Th) cell subsets during their polarization from naive Th cells establishes long-lived memory Th cells that stably maintain their lineage signatures. However, whether memory Th cells can be redifferentiated into another Th lineage is unclear. In this study, we show that Ag-specific memory Th cells were redifferentiated into Foxp3(+) T cells by TGF-beta when stimulated in the presence of all-trans retinoic acid and rapamycin. The "converted" Foxp3(+) T cells that were derived from Th2 memory cells down-regulated GATA-3 and IRF4 and produced little IL-4, IL-5, and IL-13. Instead, the converted Foxp3(+) T cells suppressed the proliferation and cytokine production of Th2 memory cells. More importantly, the converted Foxp3(+) T cells efficiently accumulated in the airways and significantly suppressed Th2 memory cell-mediated airway hyperreactivity, eosinophilia, and allergen-specific IgE production. Our findings reveal the plasticity of Th2 memory cells and provide a strategy for adoptive immunotherapy for the treatment of allergic diseases.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Cytokines; Eosinophils; Epitopes; Female; Forkhead Transcription Factors; GATA3 Transcription Factor; Immunologic Memory; Inflammation; Mice; Mice, Inbred BALB C; Neutralization Tests; Sirolimus; T-Lymphocytes, Regulatory; Th2 Cells; Transforming Growth Factor beta; Tretinoin

Bronchial epithelium is considered a key player in coordinating airway wall remodelling. The function of epithelial cells can be modulated by the underlying fibroblasts through autocrine and paracrine mechanisms.. To investigate the effect of phenotypic changes in bronchial fibroblasts from asthmatic subjects on epithelial cell proliferation.. Epithelial cells and fibroblasts derived from bronchial biopsies of asthmatic and healthy controls were cultured in an engineered model. Proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium-bromid (MTT). Epidermal growth factor receptor (EGFR), cyclin-dependent kinase inhibitors p21 and p27 were measured by western blots. Total and active forms of transforming growth factor (TGF)-β₁ were measured using ELISA and bioassay. TGF-β was inhibited using a recombinant TGF-β soluble receptor II protein.. Proliferation of epithelial cells from asthmatics (AE) is increased when cells were cultured with fibroblasts from normal controls (NF). Fibroblasts from asthmatics (AF) significantly decreased the proliferation of epithelial cells from healthy subjects (NE). Activation of p21, p27, EGFR and TGF-β₁ reflects the proliferation data by decreasing in AE cultured with NF and increasing in NE cultured with AF. Neutralization of TGF-β increased proliferation of epithelial cells cultured in the asthmatic model.. Fibroblasts from asthmatic subjects regulate epithelial cell prolifearation, and TGF-β signalling may represent one of the pathway involved in these interactions.

Topics: Asthma; Blotting, Western; Bronchi; Cell Communication; Cell Proliferation; Cells, Cultured; Coculture Techniques; Cyclin-Dependent Kinase Inhibitor Proteins; Enzyme-Linked Immunosorbent Assay; Epithelial Cells; ErbB Receptors; Fibroblasts; Humans; Microscopy, Electron, Scanning; Respiratory Mucosa; Reverse Transcriptase Polymerase Chain Reaction; Signal Transduction; Tissue Engineering; Transforming Growth Factor beta

Asthma is characterized by airway remodeling associated with an increase in the deposition of ECM proteins such as type I collagen. These components are mainly produced by fibroblasts. Inhaled corticosteroids are considered the cornerstone of asthma therapy. Despite substantial evidence as to the anti-inflammatory action of corticosteroids, their effect on controlling ECM protein deposition in the airways is not completely understood. This study determined the effect of dexamethasone (Dex) on collagen production by bronchial fibroblasts derived from asthmatic and healthy subjects. Expression of procollagen mRNA in fibroblasts from asthmatics and normal controls was determined by quantitative PCR. Regulation of the procollagen-alpha(1)I promoter was evaluated by transient transfections. Transforming growth factor-beta (TGF-beta) protein expression was determined by ELISA. Protein expression of glucocorticoid receptor (GR) and interaction with activator protein-1 (AP-1), a collagen regulatory transcription factor, was assessed by Western blots, coimmunoprecipitations, and EMSA. AP-1 overexpression was performed by transient transfection using c-Fos/c-Jun expression plasmids. Dex significantly downregulated procollagen production and promoter activity in normal fibroblasts but had no effect on asthmatic fibroblasts. AP-1 and GR interaction increased after Dex stimulation in asthmatic fibroblasts. AP-1 overexpression in control fibroblasts abrogated collagen gene response to Dex. These results show that Dex failed to reduce collagen production in fibroblasts from asthmatic subjects. This impaired response may be related to AP-1 overexpression in these cells.

Topics: Adult; Airway Remodeling; Asthma; Bronchi; Collagen Type I; Collagen Type I, alpha 1 Chain; Dexamethasone; Fibroblasts; Gene Expression; Humans; Promoter Regions, Genetic; Receptors, Glucocorticoid; Transcription Factor AP-1; Transfection; Transforming Growth Factor beta

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

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

Periostin is considered to be a matricellular protein with expression typically confined to cells of mesenchymal origin. Here, by using in situ hybridization, we show that periostin is specifically up-regulated in bronchial epithelial cells of asthmatic subjects, and in vitro, we show that periostin protein is basally secreted by airway epithelial cells in response to IL-13 to influence epithelial cell function, epithelial-mesenchymal interactions, and extracellular matrix organization. In primary human bronchial epithelial cells stimulated with periostin and epithelial cells overexpressing periostin, we reveal a function for periostin in stimulating the TGF-beta signaling pathway in a mechanism involving matrix metalloproteinases 2 and 9. Furthermore, conditioned medium from the epithelial cells overexpressing periostin caused TGF-beta-dependent secretion of type 1 collagen by airway fibroblasts. In addition, mixing recombinant periostin with type 1 collagen in solution caused a dramatic increase in the elastic modulus of the collagen gel, indicating that periostin alters collagen fibrillogenesis or cross-linking and leads to stiffening of the matrix. Epithelial cell-derived periostin in asthma has roles in TGF-beta activation and collagen gel elasticity in asthma.

Topics: Asthma; Bronchi; Cell Adhesion Molecules; Cells, Cultured; Collagen Type I; Elasticity; Epithelial Cells; Gels; Humans; Matrix Metalloproteinases; Transforming Growth Factor beta

The T cell-driven airway inflammation in chronic asthma is uninhibited and sustained. We examined the resistance of T cells from asthmatic patients against suppression by TGF-β, IL-10 and glucocorticoids and explored its signaling mechanism. CD4(+)CD25(-) T cells from allergic asthmatic subjects demonstrated increased TCR-stimulated proliferation as compared with healthy and chronic obstructive pulmonary disease controls. This proliferation was resistant to inhibition by TGF-β, IL-10, and dexamethasone and to anergy induction. CD4 T cells from asthmatic patients, but not chronic obstructive pulmonary disease, allergic rhinitis, and healthy subjects, showed increased expression of MEK1, heightened phosphorylation of ERK1/2, and increased levels of c-Fos. IL-2 and IL-4 stimulated the expression of MEK1 and c-Fos and induced T cell resistance. The inhibition of MEK1 reversed, whereas induced expression of c-Fos and JunB promoted T cell resistance against TGF-β- and IL-10-mediated suppression. We have uncovered an IL-2- and IL-4-driven MEK1 induction mechanism that results in heightened ERK1/2 activation in asthmatic T cells and make them resistant to certain inhibitory mechanisms.

Topics: Adult; Aged; Asthma; CD4-Positive T-Lymphocytes; Cell Proliferation; Cell Separation; Clonal Anergy; Enzyme-Linked Immunosorbent Assay; Flow Cytometry; Fluorescent Antibody Technique; Gene Expression; Gene Expression Regulation; Humans; Interleukin-10; Interleukin-2; Interleukin-4; Lymphocyte Activation; MAP Kinase Kinase 1; Middle Aged; Signal Transduction; Transforming Growth Factor beta; Young Adult

Topics: Animals; Anti-Asthmatic Agents; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Drugs, Chinese Herbal; Humans; Immunoglobulin E; Interferon-gamma; Lung; Pneumonia; Pulmonary Eosinophilia; Tea; Th2 Cells; Transforming Growth Factor beta

Airway remodeling is observed in the airways of patients with asthma, and differentiation of fibroblasts to myofibroblasts plays a critical role in the progress of airway remodeling. Viral infection induces not only the disease development and exacerbations but also airway remodeling. The aim of this study was to evaluate whether the activation of Toll-like receptor 3 (TLR3) can affect the differentiation of fibroblasts to myofibroblasts and the extracellular matrix (ECM) protein production. Human fetal lung fibroblasts (HFL-1) and adult lung fibroblasts were treated with a synthetic double-stranded RNA, polyinosine-polycytidylic acid (poly[I:C]) and the expression of alpha-smooth muscle actin (alpha-SMA), a marker of myofibroblast differentiation, was evaluated. The release of transforming growth factor-beta(1) (TGF-beta(1)) and ECM protein production were assessed. The effect of anti-TGF-beta antibody on the alpha-SMA and ECM production was also assessed. Poly(I:C) significantly augmented the alpha-SMA expression (P < 0.01) and release of TGF-beta(1) (P < 0.01) compared with control. Bafilomycin, an inhibitor of TLR3 signaling, diminished poly(I:C)-augmented TGF-beta(1) release. Anti-TGF-beta(1) antibody inhibited the poly(I:C)-augmented alpha-SMA expression. Poly(I:C) enhanced translocation of nuclear factor-kB (NF-kappaB) and interferon regulatory factor-3 (IRF-3) into the nucleus. Poly(I:C)-augmented TGF-beta(1) release was almost completely blocked by NF-kappaB inhibitors, but not by silencing IRF-3. The production of fibronectin and collagen I expression were significantly increased by poly(I:C) (P < 0.01) and they were inhibited by anti-TGF-beta antibody. These results suggest that activation of TLR3 can affect the differentiation to myofibroblasts and enhance ECM production via the NF-kappaB-TGF-beta(1)-dependent pathway.

Topics: Asthma; Cell Differentiation; Collagen; Fibroblasts; Fibronectins; Gene Silencing; Humans; Microscopy, Fluorescence; Models, Biological; Muscles; NF-kappa B; Poly I-C; Signal Transduction; Toll-Like Receptor 3; Transforming Growth Factor beta

Increased airway smooth muscle (ASM) mass, a characteristic finding in asthma, may be caused by hyperplasia or hypertrophy. Cell growth requires increased translation of contractile apparatus mRNA, which is controlled, in part, by glycogen synthase kinase (GSK)-3beta, a constitutively active kinase that inhibits eukaryotic initiation factor-2 activity and binding of methionyl tRNA to the ribosome. Phosphorylation of GSK-3beta inactivates it, enhancing translation. We sought to quantify the contributions of hyperplasia and hypertrophy to increased ASM mass in ovalbumin (OVA)-sensitized and -challenged BALB/c mice and the role of GSK-3beta in this process. Immunofluorescent probes, confocal microscopy, and stereological methods were used to analyze the number and volume of cells expressing alpha-smooth muscle actin and phospho-Ser(9) GSK-3beta (pGSK). OVA treatment caused a 3-fold increase in ASM fractional unit volume or volume density (Vv) (PBS, 0.006 +/- 0.0003; OVA, 0.014 +/- 0.001), a 1.5-fold increase in ASM number per unit volume (Nv), and a 59% increase in volume per cell (Vv/Nv) (PBS, 824 +/- 76 microm(3); OVA, 1,310 +/- 183 mum(3)). In OVA-treated mice, there was a 12-fold increase in the Vv of pGSK (+) ASM, a 5-fold increase in the Nv of pGSK (+) ASM, and a 1.6-fold increase in Vv/Nv. Lung homogenates from OVA-treated mice showed increased GSK-3beta phosphorylation and lower GSK-3beta activity. Both hyperplasia and hypertrophy are responsible for increased ASM mass in OVA-treated mice. Phosphorylation and inactivation of GSK-3beta are associated with ASM hypertrophy, suggesting that this kinase may play a role in asthmatic airway remodeling.

Topics: Actins; Animals; Asthma; Cell Size; Flow Cytometry; Fluorescent Antibody Technique; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Hyperplasia; Hypertrophy; Immunoblotting; Immunoprecipitation; Lung; Mice; Mice, Inbred BALB C; Microscopy, Confocal; Microscopy, Fluorescence; Muscle, Smooth; Ovalbumin; Phosphorylation; Pneumonia; Respiratory System; Transforming Growth Factor beta

Cytokine-mediated interactions among inflammatory cells may contribute to pathogenesis of allergic asthma. To understand the role of soluble interleukin-10 (IL-10) and transforming growth factor-beta (TGF-beta) on the disease activity and regulation in asthma, changes in serum concentrations of IL-10 and TGF-beta elaborated by activated T-lymphocyte before and after prednisolone therapy with clinical improvement were determined. Circulating levels of IL-10 and TGF-beta in sera from 16 normal control subjects and in sera from 22 allergic asthmatic children with acute exacerbation and in stable condition were respectively detected by commercially available enzyme-linked immunosorbent assay kits. The mean concentrations of serum IL-10 in asthmatics with acute exacerbation (6.77 +/- 4.08 pg/mL) and during stable condition (5.14 +/- 1.17 pg/mL) were lower than that in control subjects (7.15 +/- 4.72 pg/mL). However, the difference was not statistically significant among these three study groups. The mean concentration of serum TGF-beta in stable asthmatics (40.73 +/- 15.95 pg/mL) was significantly higher than that in asthmatics with acute exacerbation (27.64 +/- 3.66 pg/mL; p < 0.05) and that in healthy control group (28.77 +/- 8.35 pg/mL; p < 0.05), while there was no statistical difference between the latter two groups. This study provides further evidence that serum TGF-beta, rather than IL-10, may play a role in regulation of disease activity and serve as an indicator for clinical control of allergic asthmatics.

Topics: Adolescent; Anti-Inflammatory Agents; Asthma; Biomarkers; Child; Child, Preschool; Enzyme-Linked Immunosorbent Assay; Humans; Interleukin-10; Prednisolone; Time Factors; Transforming Growth Factor beta

Allergic asthma is a chronic inflammatory disease mediated by T helper (Th)2 cell immune responses. Currently, immunotherapies based on both immune deviation and immune suppression, including the development of recombinant mycobacteria as immunoregulatory vaccines, are attractive treatment strategies for asthma. In our previous studies, we created a genetically recombinant form of bacille Calmette-Guerin (rBCG) that expressed Der p2 of house dust mites and established that it induced a shift from a Th2 response to a Th1 response in naive mice. However, it is unclear whether rBCG could suppress allergic airway inflammation in a mouse model. In this article we report that rBCG dramatically inhibited airway inflammation, eosinophilia, mucus production and mast cell degranulation in allergic mice. Analysis of interferon-gamma (IFN-gamma) and interleukin-4 (IL-4) levels in bronchoalveolar lavage fluid (BALF) and lung tissue revealed that the suppression was associated with a shift from a Th2 response to a Th1 response. At the same time, rBCG induced a CD4(+) CD25(+) Foxp3(+) T-cell subtype that could suppress the proliferation of Th2 effector cells in vitro in an antigen-specific manner. Moreover, suppression of CD4(+) CD25(+) T cells could be adoptively transferred. Thus, our results demonstrate that rBCG induces both generic and specific immune responses. The generic immune response is associated with a shift from a Th2 to a Th1 cytokine response, whereas the specific immune response against Der p2 appears to be related to the expansion of transforming growth factor-beta (TGF-beta)-producing CD4(+) CD25(+) Foxp3(+) regulatory T cells. rBCG can suppress asthmatic airway inflammation through both immune deviation and immune suppression and may be a feasible, efficient immunotherapy for asthma.

Topics: Adoptive Transfer; Animals; Antigens, Dermatophagoides; Arthropod Proteins; Asthma; BCG Vaccine; Bronchoalveolar Lavage Fluid; Cell Movement; Cytokines; Disease Models, Animal; Eosinophilia; Female; Immunotherapy, Active; Interferon-gamma; Interleukin-4; Lung; Mice; Mice, Inbred C57BL; Pyroglyphidae; Recombinant Fusion Proteins; T-Lymphocytes, Regulatory; Th1 Cells; Th2 Cells; Transforming Growth Factor beta

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

In the airways of asthmatic patients, activated fibroblasts account for an excessive matrix production including proteoglycans (PGs). Transforming growth factor-beta (TGFbeta), metalloproteinases (MMPs) and their tissue inhibitors (TIMPs) play key roles in matrix turnover. It is unclear whether asthma therapy with combination of inhaled glucocorticoids and long-acting beta(2)-agonists affects metalloproteolytic equilibrium and by that counteracts airway fibrosis. The effects of the glucocorticoid, budesonide, and the long-acting beta(2)-agonist, formoterol, on the PG production and the activity of PGs' main regulators: MMP-3, MMP-9, MMP-2 and TIMP-1 were investigated in human lung fibroblasts (HFL-1) treated for 24h with TGFbeta1 (10 ng/ml) without/with budesonide (10(-9) to 10(-6)M) and/or formoterol (10(-11) to 10(-6)M). TGFbeta1 significantly increased production of PGs and TIMP-1, and the activity of MMP-3, MMP-9 and MMP-2. Concurrent budesonide/formoterol combination counteracted the enhanced: PG and TIMP-1 production, MMP-9 activity and MMP-9/TIMP-1 ratio, whereas MMP-2 and MMP-3 were not affected and so their ratios to TIMP-1 were significantly increased. Budesonide or formoterol alone achieved equal effects as budesonide/formoterol on MMP-9 and MMP-9/TIMP-1 ratio but had no effects on TIMP-1, MMP-2 or MMP-3. In the formoterol absence, higher budesonide concentrations were required to reduce the PG production, whereas formoterol alone had no effects. These results suggest that the budesonide/formoterol combination enhanced metalloproteolytic activity of human lung fibroblasts via a synergistic decrease of TIMP-1, and that this mechanism may be involved in the synergistic inhibition of the TGFbeta1-induced PG production. This implies that budesonide/formoterol combination therapy can counteract excessive matrix production and thus pathological airway fibrotic remodeling in asthma.

Topics: Asthma; Bronchodilator Agents; Budesonide; Cells, Cultured; Ethanolamines; Fibroblasts; Formoterol Fumarate; Humans; Lung; Matrix Metalloproteinase 2; Matrix Metalloproteinase 3; Matrix Metalloproteinase 9; Matrix Metalloproteinases; Proteoglycans; Respiratory System; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta

Chronic asthma is characterized by ongoing recruitment of inflammatory cells and airway hyperresponsiveness leading to structural airway remodeling. Although alpha 4 beta 1 and beta2 integrins regulate leukocyte migration in inflammatory diseases and play decisive roles in acute asthma, their role has not been explored under the chronic asthma setting. To extend our earlier studies with alpha 4(Delta/Delta) and beta2(-/-) mice, which showed that both alpha 4 and beta2 integrins have nonredundant regulatory roles in acute ovalbumin (OVA)-induced asthma, we explored to what extent these molecular pathways control development of structural airway remodeling in chronic asthma.. Control, alpha 4(Delta/Delta), and beta2(-/-) mouse groups, sensitized by intraperitoneal OVA as allergen, received intratracheal OVA periodically over days 8 to 55 to induce a chronic asthma phenotype. Post-OVA assessment of inflammation and pulmonary function (airway hyperresponsiveness), together with airway modeling measured by goblet cell metaplasia, collagen content of lung, and transforming growth factor beta1 expression in lung homogenates, were evaluated.. In contrast to control and beta2(-/-) mice, alpha 4(Delta/Delta) mice failed to develop and maintain the composite chronic asthma phenotype evaluated as mentioned and subepithelial collagen content was comparable to baseline. These data indicate that beta2 integrins, although required for inflammatory migration in acute asthma, are dispensable for structural remodeling in chronic asthma.. alpha 4 integrins appear to have a regulatory role in directing transforming growth factor beta-induced collagen deposition and structural alterations in lung architecture likely through interactions of Th2 cells, eosinophils, or mast cells with endothelium, resident airway cells, and/or extracellular matrix.

Topics: Animals; Asthma; CD18 Antigens; Chemotaxis, Leukocyte; Chronic Disease; Collagen; Inflammation; Integrin beta4; Lung; Mice; Mice, Knockout; Models, Genetic; Ovalbumin; Phenotype; Respiratory Function Tests; Transforming Growth Factor beta

Treatment of patients with allergic asthma using low doses of peptides containing T cell epitopes from Fel d 1, the major cat allergen, reduces allergic sensitization and improves surrogate markers of disease. Here, we demonstrate a key immunological mechanism, linked epitope suppression, associated with this therapeutic effect. Treatment with selected epitopes from a single allergen resulted in suppression of responses to other ("linked") epitopes within the same molecule. This phenomenon was induced after peptide immunotherapy in human asthmatic subjects and in a novel HLA-DR1 transgenic mouse model of asthma. Tracking of allergen-specific T cells using DR1 tetramers determined that suppression was associated with the induction of interleukin (IL)-10(+) T cells that were more abundant than T cells specific for the single-treatment peptide and was reversed by anti-IL-10 receptor administration. Resolution of airway pathophysiology in this model was associated with reduced recruitment, proliferation, and effector function of allergen-specific Th2 cells. Our results provide, for the first time, in vivo evidence of linked epitope suppression and IL-10 induction in both human allergic disease and a mouse model designed to closely mimic peptide therapy in humans.

Topics: Allergens; Animals; Asthma; Bronchial Hyperreactivity; Cats; Desensitization, Immunologic; Disease Models, Animal; Double-Blind Method; Epitopes; Forkhead Transcription Factors; Genes, MHC Class II; Glycoproteins; HLA-DR1 Antigen; Humans; Immune Tolerance; Interleukin-10; Lung; Mice; Mice, Inbred C57BL; Mice, Transgenic; Peptides; Placebos; Randomized Controlled Trials as Topic; Receptors, Interleukin-10; Th2 Cells; Transforming Growth Factor beta

We try to find out the influence of traditional Chinese Medicine Astragali-Cordyceps Mixtura (ACM) on TGF-beta/Smad signal pathway in the lung of asthma airway remodeling.. Mice were sensitized and challenged by OVA to establish a model of asthma. To assess the effects of ACM on the mice, animals of the ACM groups were treated with ACM. Data were achieved by using techniques as follow: counting cell number of BALF, assaying the amount of collagen deposition by Masson's staining, performing RT-PCR and immunohistochemistry for mRNA and protein expression of TGF-beta1, Smad3 and Smad7.. The depositions of collagen in airway wall greatly increased at the model group compared with that of the normal group. In contrast, these decreased at the ACM groups. As compared with the control group, TGF-beta1 expression also decreased at both mRNA and protein level at the ACM-M group versus increased both at the model group. Whereas, Smad7 significantly decreased only at the model group and partly restored at the ACM-M group.. ACM greatly improves the symptoms of asthma airway remodeling by inhibiting the expression of TGF-beta1 and upregulating the amount of Smad7.

Topics: Animals; Asthma; Astragalus Plant; Base Sequence; Cordyceps; DNA Primers; Lung; Male; Mice; Mice, Inbred C57BL; Plant Extracts; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; Smad3 Protein; Smad7 Protein; Transforming Growth Factor beta

Both transforming growth factor (TGF)-beta(1) and activin-A have been implicated in airway remodeling in asthma, but the modulation of their specific signaling pathways after disease activation remains undefined.. To define the expression kinetics of TGF-beta(1), activin-A ligands, and follistatin (a natural activin inhibitor), their type I and type II receptors (activin-like kinase[ALK]-1, ALK-5, ALK-4, TbetaRII, and ActRIIA/RIIB) and activation of signaling (via phosphorylated (p) Smad2), in the asthmatic airway after allergen challenge.. Immunohistochemistry was performed on bronchial biopsies from 15 mild atopic patients with asthma (median age, 25 years; median FEV(1)% predicted, 97%) at baseline and 24 hours after allergen inhalation. Functional effects of activin-A were evaluated by using cultured normal human bronchial epithelial (NHBE) cells.. pSmad2(+) epithelial cells increased at 24 hours (P = .03), and pSmad2 was detected in submucosal cells. No modulation of activin-A, follistatin, or TGF-beta(1) expression was demonstrated. Activin receptor(+) cells increased after allergen challenge: ALK-4 in epithelium (P = .04) and submucosa (P = .04), and ActRIIA in epithelium (P = .01). The TGF-beta receptor ALK-5 expression was minimal in the submucosa at baseline and after challenge and was downregulated in the epithelium after challenge (P = .02), whereas ALK-1 and TbetaRII expression in the submucosa increased after allergen challenge (P = .03 and P = .004, respectively). ALK-1 and ALK-4 expression by T cells was increased after allergen challenge. Activin-A induced NHBE cell proliferation, was produced by NHBE cells in response to TNF-alpha, and downregulated TNF-alpha and IL-13-induced chemokine production by NHBE cells.. Both TGF-beta and activin signaling pathways are activated on allergen provocation in asthma. Activin-A may contribute to resolution of inflammation.

Topics: Activin Receptors, Type I; Activin Receptors, Type II; Activins; Adult; Allergens; Asthma; Bronchial Provocation Tests; Cell Proliferation; Cells, Cultured; Cytokines; Epithelial Cells; Follistatin; Humans; Interleukin-13; Middle Aged; Receptors, Transforming Growth Factor beta; Signal Transduction; Smad2 Protein; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

IL-33, a new member of the IL-1 cytokine family, promotes Th2 inflammation, but evidence on the implications of this cytokine in asthma is lacking. IL-33 would be mainly expressed by structural cells, but whether proinflammatory cytokines modulate its expression in airway smooth muscle cells (ASMC) is unknown. Endobronchial biopsies were obtained from adults with mild (n = 8), moderate (n = 8), severe (n = 9), asthma and from control subjects (n = 5). Immunocytochemistry, laser-capture microdissection, reverse transcriptase, and real-time quantitative PCR were used for determining IL-33 expression in the lung tissues. ASMC isolated from resected lung specimens were cultured with proinflammatory cytokines and with dexamethasone. IL-33 expression by ASMC was determined by PCR, ELISA, and Western blotting. Higher levels of IL-33 transcripts are detected in biopsies from asthmatic compared with control subjects, and especially in subjects with severe asthma. ASMC show IL-33 expression at both protein and mRNA levels. IL-33 and TNF-alpha transcript levels correlate in the lung tissues, and TNF-alpha up-regulates IL-33 expression by cultured ASMC in a time- and dose-dependent manner. IFN-gamma also increases IL-33 expression and shows synergistic effect with TNF-alpha. Dexamethasone fails to abolish TNF-alpha-induced IL-33 up-regulation. IL-33 expression increases in bronchial biopsies from subjects with asthma compared with controls, as well as subjects with asthma severity. ASMC are a source of the IL-33 cytokine. Our data propose IL-33 as a novel inflammatory marker of severe and refractory asthma.

Topics: Adult; Aged; Aged, 80 and over; Anti-Inflammatory Agents; Antiviral Agents; Asthma; Biomarkers; Cells, Cultured; Dexamethasone; Drug Synergism; Female; Gene Expression; Humans; Interferon-gamma; Interleukin-13; Interleukin-33; Interleukin-4; Interleukins; Lung; Male; Middle Aged; Myocytes, Smooth Muscle; Plicamycin; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha; Up-Regulation

While recent studies have shown that patients with COPD and patients with asthma exhibit evidence of airway and systemic inflammation, markers of systemic inflammation have not been compared between the two diseases. To evaluate circulating inflammatory markers, blood was sampled from 111 patients with COPD, 75 control subjects and 46 asthmatic patients (some of whom were smokers). Measurements of WCC, serum levels of fibrinogen, high-sensitivity c-reactive protein (hs-CRP), interleukin-8 (IL-8), interleukin-6 (IL-6), tumour necrosis factor-alpha (TNF-alpha), transforming growth factor (TGF)-beta1, tissue inhibitors of metalloproteinase (TIMP)-1, neutrophil elastase and alphal-antitrypsin (alpha1-AT) were done. Serum TNF-alpha, IL-6, and TIMP-1 concentrations were significantly higher in patients with stable COPD and patients with asthma than in control patients. Serum alpha1-AT levels were significantly higher in COPD patients than in asthmatic patients and control subjects and serum TGF-beta1 levels were higher in asthma patients than in COPD patients. Smoking status had no effect on markers in COPD and asthmatic patients. Although COPD and asthma share common markers of systemic inflammation, serum levels of TGF-beta1 and alpha1-AT may reflect differences between the diseases.

Topics: Aged; alpha 1-Antitrypsin; Asthma; Biomarkers; Humans; Inflammation; Male; Middle Aged; Pulmonary Disease, Chronic Obstructive; Transforming Growth Factor beta

CD4(+)CD25(+) regulatory T cells can inhibit excessive T-cell responses in vivo. We have previously demonstrated that prophylactic administration of CD4(+)CD25(+) regulatory T cells suppresses the development of acute allergen-induced airway inflammation in vivo.. We sought to determine the effect of therapeutic transfer of CD4(+)CD25(+) regulatory T cells on established pulmonary inflammation and the subsequent development of airway remodeling.. CD4(+)CD25(+) cells were transferred after the onset of allergic inflammation, and airway challenges were continued to induce chronic inflammation and airway remodeling.. Administration of CD4(+)CD25(+) regulatory T cells reduced established lung eosinophilia, T(H)2 infiltration, and expression of IL-5, IL-13, and TGF-beta. Moreover, subsequent mucus hypersecretion and peribronchial collagen deposition were reduced after prolonged challenge. In contrast, transfer of CD4(+)CD25(+) regulatory T cells had no effect on established airway hyperreactivity either 7 days or 4 weeks after transfer.. In this study we demonstrate for the first time that therapeutic transfer of CD4(+)CD25(+) regulatory T cells can resolve features of chronic allergen-induced inflammation and prevent development of airway remodeling.

Topics: Adoptive Transfer; Allergens; Animals; Asthma; Cell Separation; Collagen; Eosinophils; Female; Immunoglobulin E; Inflammation; Interleukin-13; Interleukin-5; Lung; Mice; Mice, Inbred BALB C; Mucus; Ovalbumin; Respiratory Hypersensitivity; T-Lymphocytes, Regulatory; Th2 Cells; Transforming Growth Factor beta

Asthma is a chronic inflammatory disorder caused by T-cell-mediated inflammation within airways. No antigen-specific treatment has been available. Using an OVA-induced murine asthma model, we find that co-immunization of an OVA epitope peptide with a DNA vaccine encoding the same epitope is able to prevent this experimental asthma as evidenced in the marked reduction of infiltrations of eosinophils and lymphocytes into the site of the allergen challenge. We demonstrate that the prevention of experimental asthma was directly related to the induction of a population of OVA-specific T-regulatory cells (Treg) exhibiting a CD4(+)CD25(-)FoxP3(+) phenotype and expressing IL-10, TGF-beta and IFN-gamma following the co-immunization. Blockade of IL-10 and TGF-beta of the Treg by anti-IL-10 and TGF-beta antibodies is partially able to reverse the suppression in vitro and in vivo, which caused the recurrence of the inflammation. Furthermore, adoptive transfer of the induced Treg is also able to suppress the OVA-induced asthma. To our knowledge, the combination of peptide with its cognate DNA vaccine protect experimental asthma via the induced epitope-specific Treg has not been previously reported and such strategy may lead to a novel immunotherapy against asthma in humans.

Topics: Adoptive Transfer; Allergens; Animals; Asthma; CD4-Positive T-Lymphocytes; Disease Models, Animal; Epitopes; Female; Interferon-gamma; Interleukin-10; Mice; Mice, Inbred BALB C; Ovalbumin; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Vaccination; Vaccines, DNA

Asthma is a complex respiratory disease, characterized by airway inflammation and reversible airway obstruction. Both genetic and environmental factors are important in the development and expression of the disease. In order to analyze the genetic profile of Th1 and Th2 cytokines in Iranian asthmatic patients, this study was performed. The allele and genotype frequencies of a number of polymorphic genes coding for IL-10, TGF-beta, IL-2, IL-12, and IFN-gamma were investigated in 60 patients with asthma in comparison with 140 controls. The most frequent genotypes in our patients were IL-10 GA at position-1082 (p = 0.001), IL-10 CT at position -819 (p = 0.001), IL-10 CA at position -592 (p = 0.0001), IL-12 CA at position -1188 (p = 0.003), TGF-beta CG at codon 25 (p = 0.002), IL-2 GT at position -330 (p = 0.004). In contrast, the frequencies of the genotypes IL-10 AA at position -1082 (p = 0.0001) and GG at position -1082 (p = 0.01), IL-10 CC at position -819 (p = 0.001) and TT at position -819 (p = 0.01), TGF-beta TT at codon 10 (p = 0.001), TGF-beta GG at codon 25 (p = 0.005), IL-12 AA at position -1188 (p = 0.004), IL-2 TT at position -330 (p = 0.01) were significantly lower in the patient group. The most frequent haplotypes in the patients were IL-10 GCC (p = 0.008) and ATA (p = 0.0001) at position -1082, -819, -592, and TGF-beta CC (p = 0.036) at codon 10 and codon 25. In contrast, the frequencies of the IL-10 ACC (p = 0.001), TGF-beta TG (p = 0.024), and IL-2 TT (p = 0.001) and GT (p= 0.0001) in the patients were significantly lower than controls. Considering the high frequency of presence of IL-10 ATA haplotype and the IL-2 GT genotype, it seems that the production of IL-10 and IL-2 in the asthmatic patients could be lower than normal subjects.

Topics: Asthma; Cytokines; Female; Gene Frequency; Genotype; Humans; Interferon-gamma; Interleukin-10; Interleukin-12; Interleukin-2; Iran; Male; Polymerase Chain Reaction; Polymorphism, Genetic; Transforming Growth Factor beta

Periostin is an extracellular matrix protein that has been primarily studied in the context of the heart, where it has been shown to promote cardiac repair and remodeling. In this study, we focused on the role of periostin in an allergic eosinophilic inflammatory disease (eosinophilic esophagitis (EE)) known to involve extensive tissue remodeling. Periostin was indeed markedly overexpressed (35-fold) in the esophagus of EE patients, particularly in the papillae, compared with control individuals. Periostin expression was downstream from transforming growth factor-beta and interleukin-13, as these cytokines were elevated in EE esophageal samples and markedly induced periostin production by primary esophageal fibroblasts (107- and 295-fold, respectively, at 10 ng ml(-1)). A functional role for periostin in eliciting esophageal eosinophilia was demonstrated, as periostin-null mice had a specific defect in allergen-induced eosinophil recruitment to the lungs and esophagus (66 and 72% decrease, respectively). Mechanistic analyses revealed that periostin increased (5.8-fold) eosinophil adhesion to fibronectin. As such, these findings extend the involvement of periostin to esophagitis and uncover a novel role for periostin in directly regulating leukocyte (eosinophil) accumulation in T helper type 2-associated mucosal inflammation in both mice and humans.

Topics: Animals; Asthma; Cell Adhesion; Cell Adhesion Molecules; Dermatitis, Atopic; Eosinophils; Esophagitis; Esophagus; Fibroblasts; Humans; Hypersensitivity; Interleukin-13; Lung; Mice; Mice, Knockout; Pulmonary Eosinophilia; Rhinitis; Transforming Growth Factor beta

Epithelial damage is an important pathophysiologic feature of asthma. Bronchial epithelium damage results in release of growth factors such as transforming growth factor (TGF)-beta(1) that may affect epithelial cell proliferation. The objective of our study is to evaluate the importance of TGF-beta(1) in regulating epithelial cell repair in asthma. We evaluated the effect of TGF-beta(1) on epidermal growth factor (EGF)-induced proliferation and downstream signaling in epithelial cells obtained from subjects with asthma compared with cells from healthy subjects. Cell proliferation was evaluated by bromodeoxyuridine incorporation. EGF receptor (EGFR), mitogen-activated protein kinase, TGF-beta receptors, Smads, Smad anchor for receptor activation (SARA), and cyclin-dependant kinase inhibitors were evaluated by Western blot. TGF-beta(1) and receptor expression were measured by RT-PCR and by enzyme-linked immunosorbent assay. Proliferation of epithelial cells at baseline and after EGF stimulation was significantly reduced in cells derived from subjects with asthma compared with cells obtained from healthy control subjects. EGF-induced ERK1/2 phosphorylation was reduced in epithelial cells from subjects with asthma compared with cells from healthy control subjects. This was paralleled with a reduced EGFR phosphorylation. Addition of TGF-beta(1) significantly decreased EGF-induced cell proliferation. TGF-beta(1) production was higher in asthmatic epithelial cells compared with normal cells. This was supported by a high expression of pSmad 3 and SARA in cells derived from individuals with asthma compared with normal subjects. Cycline-dependent kinase inhibitors were highly expressed in asthmatic compared with normal cells. Inhibition of TGF-beta(1) signaling in asthmatic epithelial cells restored EGFR, ERK1/2 phosphorylation, and cell proliferation induced by EGF. Our results suggest that TGF-beta restrains EGFR phosphorylation and downstream signaling in bronchial epithelial cells.

Topics: Asthma; Base Sequence; Blotting, Western; Case-Control Studies; Cell Proliferation; DNA Primers; Enzyme-Linked Immunosorbent Assay; Epidermal Growth Factor; Epithelial Cells; Humans; MAP Kinase Signaling System; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta

Abnormal epithelial repair to damage participates in airway remodeling in asthma by the paracrine regulation of mesenchymal cell functions. Retinoids control epithelial functions through nuclear retinoic acid receptor (RAR) and retinoid X receptor (RXR) activation, yet their expression and contribution to epithelial repair and to airway remodeling in asthma are unknown. We determined the plasma levels of retinol and the immunohistochemical expression of retinoid receptors in damaged and repaired bronchial epithelium from 9 control subjects, 10 subjects with intermittent asthma, 8 subjects with mild-to-moderate asthma, and 8 subjects with severe asthma. In addition, the effect of the retinoid receptor ligands, all-trans-retinoic acid, and 9-cis retinoic acid, on the synthesis of 38 factors potentially involved in epithelial repair and in airway remodeling was determined in human cultured airway epithelial cells and correlated with cell migration and proliferation. Circulating retinol was similar in the three patient groups. In contrast, the epithelial expression of RARgamma, RXRalpha, and RXRgamma was greater in subjects with severe asthma, as compared with patients with milder disease and to control subjects. Retinoid receptor expression correlated positively with the proportion of morphologically intact epithelium. In vitro, retinoids up-regulated the expression of the transcripts encoding transforming growth factor (TGF)-beta1, metalloproteinase-9, beta1-integrin, and hepatocyte growth factor receptor, and promoted wound repair and chemokinesis of human airway epithelial cells without altering proliferation. Cell treatment with an anti-TGF-beta1 monoclonal antibody partially reduced retinoid-induced effects. Persistent interaction between retinoids and some of their receptors, which are overexpressed by the bronchial epithelium of individuals with severe asthma, may contribute to an abnormal repair and to airway remodeling, partly through TGF-beta1 production.

Topics: Asthma; Bronchi; Case-Control Studies; Cell Movement; Cell Proliferation; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Epithelial Cells; Gene Expression Regulation; Humans; Immunohistochemistry; Integrin beta1; Ligands; Matrix Metalloproteinase 9; Nasal Mucosa; Proto-Oncogene Proteins c-met; Receptors, Retinoic Acid; Regression Analysis; RNA, Messenger; Severity of Illness Index; Transforming Growth Factor beta; Tretinoin; Vitamin A; Wound Healing

Asthma can progress to subepithelial airway fibrosis, mediated in large part by transforming growth factor-beta (TGF-beta). The scaffolding protein caveolin-1 (cav1) can inhibit the activity of TGF-beta, perhaps by forming membrane invaginations that enfold TGF-beta receptors. The study goals were 1) to evaluate how allergen challenge affects lung expression of cav1 and the density of caveolae in vivo 2) to determine whether reduced cav1 expression is mediated by interleukin (IL)-4 and 3) to measure the effects of decreased expression of cav1 on TGF-beta signaling. C57BL/6J, IL-4-deficient mice, and cav1-deficient mice, sensitized by intraperitoneal injections of phosphate-buffered saline or ovalbumin (OVA) at days 0 and 12, received intranasal phosphate-buffered saline or OVA challenges at days 24, 26, and 28. Additionally, another group of C57BL/6J mice received IL-4 by intratracheal instillation for 7 days. We confirmed that the OVA-allergen challenge increased eosinophilia and T-helper type 2-related cytokine levels (IL-4, IL-5, and IL-13) in bronchoalveolar lavage. Allergen challenge reduced lung cav1 mRNA abundance by 40%, cav1 protein by 30%, and the number of lung fibroblast caveolae by 50%. Administration of IL-4 in vivo also substantially decreased cav1 expression. In contrast, the allergen challenge did not decrease cav1 expression in IL-4-deficient mice. The reduced expression of cav1 was associated with activation of TGF-beta signaling that was further enhanced in OVA-sensitized and challenged cav1-deficient mice. This study demonstrates a previously unknown modulation of TGF-beta signaling by IL-4, via cav1, suggesting novel therapeutic targets for controlling the effects of TGF-beta and thereby ameliorating pathological airway remodeling.

Topics: Allergens; Animals; Asthma; Bronchoalveolar Lavage; Caveolin 1; Down-Regulation; Interleukins; Mice; Mice, Knockout; Pulmonary Fibrosis; Signal Transduction; Th2 Cells; Time Factors; Transforming Growth Factor beta

It is now believed that both chronic airway inflammation and remodeling contribute significantly to airway dysfunction and clinical symptoms in allergic asthma. Transforming growth factor (TGF)-beta is a powerful regulator of both the tissue repair and inflammatory responses, and numerous experimental and clinical studies suggest that it may play an integral role in the pathogenesis of asthma.. We investigated the role of TGF-beta in the regulation of allergic airway inflammation and remodeling using a mouse model of house dust mite (HDM)-induced chronic allergic airway disease.. We have previously shown that intranasal administration of an HDM extract (5 d/wk for 5 wk) elicits robust Th2-polarized airway inflammation and remodeling that is associated with increased airway hyperreactivity. Here, Balb/c mice were similarly exposed to HDM and concurrently treated with a pan-specific TGF-beta neutralizing antibody.. We observed that anti-TGF-beta treatment in the context of either continuous or intermittent HDM exposure had no effect on the development of HDM-induced airway remodeling. To further confirm these findings, we also subjected SMAD3 knockout mice to 5 weeks of HDM and observed that knockout mice developed airway remodeling to the same extent as HDM-exposed littermate controls. Notably, TGF-beta neutralization exacerbated the eosinophilic infiltrate and led to increased airway hyperreactivity.. Collectively, these data suggest that TGF-beta regulates HDM-induced chronic airway inflammation but not remodeling, and furthermore, caution against the use of therapeutic strategies aimed at interfering with TGF-beta activity in the treatment of this disease.

Topics: Animals; Asthma; Bronchial Hyperreactivity; Bronchial Provocation Tests; Eosinophils; Female; Hypersensitivity; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Pyroglyphidae; Smad3 Protein; Transforming Growth Factor beta

Despite increasing recognition of bone morphogenetic protein (BMP) signaling in tissue remodeling, the expression pattern of ligands and signaling pathways remain undefined in the asthmatic airway.. To determine expression of BMP ligands (BMP-2, BMP-4, and BMP-7) and type I and type II receptors (ALK-2, ALK-3, ALK-6, and BMPRII) as well as evidence for activation of BMP signaling via detection of phosphorylated Smad1/5 (pSmad1/5) expression in asthmatic airways at baseline (compared with nonasthmatic controls), and after allergen challenge.. Bronchial biopsies were obtained from 6 nonasthmatic control volunteers, and 15 atopic patients with asthma (median age, 25 yr; median FEV(1)% predicted, 97%) at baseline, then at 24 hours and 7 days after allergen challenge. Expression of BMP ligands, receptors, and signaling was analyzed using immunohistochemistry.. BMP ligand expression did not differ between asthmatic and control airways at baseline. Compared with the normal airway, there was significant down-regulation of ALK-2 (P = 0.001), ALK-6 (P = 0.0009), and BMPRII (P = 0.009) expression in asthma. Allergen challenge was associated with marked and sustained up-regulation of BMP-7 in airway epithelium (P = 0.017) and infiltrating inflammatory cells (P = 0.071) (predominantly in eosinophils, but also CD4(+) T cells, mast cells, and macrophages). Up-regulation of pSmad1/5 expression (P = 0.031), ALK-2 (P = 0.002), and ALK-6 (P < 0.001) was observed indicating active signaling.. BMP receptor expression is down-regulated in the asthmatic airway, which may impede repair responses. Allergen provocation increases expression of the regulatory ligand BMP-7, activates BMP signaling, and increases receptor expression, all of which may contribute to repair and control of inflammation.

Topics: Activin Receptors, Type I; Adult; Asthma; Biopsy; Bone Morphogenetic Protein 7; Bone Morphogenetic Protein Receptors, Type I; Bone Morphogenetic Protein Receptors, Type II; Bone Morphogenetic Proteins; Bronchi; Bronchial Provocation Tests; Case-Control Studies; Down-Regulation; Epithelial Cells; Female; Humans; Male; Middle Aged; Signal Transduction; Smad Proteins; Transforming Growth Factor beta; Up-Regulation

Topics: Asthma; Bronchi; Cell Proliferation; Cells, Cultured; Drug Design; ErbB Receptors; Fibroblasts; Heparin-binding EGF-like Growth Factor; Humans; Intercellular Signaling Peptides and Proteins; Mitogen-Activated Protein Kinase 3; Phosphatidylinositol 3-Kinases; Signal Transduction; Transforming Growth Factor beta

1alpha,25-Dihydroxyvitamin D(3) (1,25(OH)(2)D(3)), a potent inhibitor of NF-kappaB expression, can prevent the maturation of dendritic cells in vitro leading to tolerogenic dendritic cells with increased potential to induce regulatory T cells. Herein, we investigated whether the combination of allergen immunotherapy with 1,25(OH)(2)D(3) potentiates the suppressive effects of immunotherapy and whether the immunoregulatory cytokines IL-10 and TGF-beta are involved in the effector phase. OVA-sensitized and challenged BALB/c mice displayed airway hyperresponsiveness (AHR) and increased serum OVA-specific IgE levels, bronchoalveolar lavage eosinophilia, and Th2 cytokine levels. In this model, the dose response of allergen immunotherapy 10 days before OVA inhalation challenge shows strong suppression of asthma manifestations at 1 mg of OVA, but partial suppression of bronchoalveolar lavage eosinophilia, IgE up-regulation, and no reduction of AHR at 100 microg. Interestingly, coadministration of 10 ng of 1,25(OH)(2)D(3) with 100 microg of OVA immunotherapy significantly inhibited AHR and potentiated the reduction of serum OVA-specific IgE levels, airway eosinophilia, and Th2-related cytokines concomitant with increased IL-10 levels in lung tissues and TGF-beta and OVA-specific IgA levels in serum. Similar effects on suboptimal immunotherapy were observed by inhibition of the NF-kappaB pathway using the selective IkappaB kinase 2 inhibitor PS-1145. The suppressive effects of this combined immunotherapy were partially reversed by treatment with mAb to either IL-10R or TGF-beta before OVA inhalation challenge but completely abrogated when both Abs were given. These data demonstrate that 1,25(OH)(2)D(3) potentiates the efficacy of immunotherapy and that the regulatory cytokines IL-10 and TGF-beta play a crucial role in the effector phase of this mouse model.

Topics: Animals; Asthma; Cytokines; Desensitization, Immunologic; Disease Models, Animal; Immunoglobulin E; Immunoglobulin G; Interleukin-10; Lung; Male; Mice; Mice, Inbred BALB C; NF-kappaB-Inducing Kinase; Ovalbumin; Protein Serine-Threonine Kinases; Pulmonary Eosinophilia; Transforming Growth Factor beta; Vitamin D

A disintegrin and metalloprotease (ADAM)-33 is a susceptibility gene for asthma and chronic obstructive pulmonary disease whose function remains unknown.. Because asthmatic bronchoalveolar lavage fluid contains high levels of soluble ADAM33 (sADAM33), which includes the catalytic domain, we postulated that its release from cell membranes might play functional roles in airway remodeling by promoting angiogenesis.. The proangiogenic activity of the highly purified catalytic domain of ADAM33 or a catalytically inactive mutant was studied in vitro (Matrigel assay), ex vivo (human embryonic/fetal lung explants) and in vivo (chorioallantoic membrane assay). The regulation of sADAM33 release from cells overexpressing full-length ADAM33 and its biological activity were characterized.. We show that the purified catalytic domain of ADAM33, but not its inactive mutant, causes rapid induction of endothelial cell differentiation in vitro, and neovascularization ex vivo and in vivo. We also show that TGF-beta(2) enhances sADAM33 release from cells overexpressing full-length ADAM33 and that this truncated form is biologically active.. The discovery that sADAM33 promotes angiogenesis defines it as a tissue remodeling gene with potential to affect airflow obstruction and lung function independently of inflammation. As TGF-beta(2) enhances sADAM33 release, environmental factors that cause epithelial damage may synergize with ADAM33 in asthma pathogenesis, resulting in a disease-related gain of function. This highlights the potential for interplay between genetic and environmental factors in this complex disease.

Topics: ADAM Proteins; Asthma; Catalytic Domain; Cell Differentiation; Endothelial Cells; Humans; Lung; Neovascularization, Pathologic; Transforming Growth Factor beta

The mechanisms underlying exacerbation of asthma induced by respiratory syncytial virus (RSV) infection have been extensively studied in human and animal models. However, most of these studies focused on acute inflammation and little is known of its long-term consequences on remodelling of the airway tissue.. The aim of the study was to use a murine model of prolonged allergen-induced airway inflammation to investigate the effect of RSV infection on allergic airway inflammation and tissue remodelling.. We subjected mice to RSV infection before or during the chronic phase of airway challenges with OVA and compared parameters of airway inflammation and remodelling at the end-point of the prolonged allergen-induced airway inflammation protocol.. RSV infection did not affect the severity of airway inflammation in any of the groups studied. However, RSV infection provoked airway remodelling in non-sensitized, allergen-challenged mice that did not otherwise develop any of the features of allergic airways disease. Increased collagen synthesis in the lung and thickening of the bronchial basal membrane was observed in non-sensitized allergen-challenged mice only after prior RSV infection. In addition, fibroblast growth factor (FGF)-2 but not TGF-beta(1) was increased in this group following RSV infection.. Our data show for the first time that RSV infection can prime the lung of mice that are not previously systemically sensitized, to develop airway remodelling in response to allergen upon sole exposure via the airways. Moreover, our results implicate RSV-induced FGF-2 in the remodelling process in vivo.

Topics: Alum Compounds; Analysis of Variance; Animals; Asthma; Bronchoalveolar Lavage Fluid; Collagen; Cytokines; Extracellular Matrix; Female; Fibroblast Growth Factor 2; Immunohistochemistry; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Syncytial Virus Infections; Respiratory Syncytial Viruses; Statistics, Nonparametric; Transforming Growth Factor beta

Churg-Strauss syndrome (CSS) is a rare systemic necrotizing vasculitis that develops in some asthma patients. However, it remains unknown what types of asthma patients can develop this condition. In addition, CSS patients experience eosinophilic pneumonia (EP) before the onset, but some asthma patients do not develop CSS following EP. Few studies have addressed the immunological differences in both cases. Additionally, it is well known that abnormality or failure of immunological tolerance by regulatory T cells (Treg) directly triggers the onset of autoimmune diseases. To elucidate the involvement of Treg in the difference in the pathogenesis of patients with and without CSS, the frequency of naturally occurring Treg (nTreg) and Treg which dominantly produce IL-10 and TGF-beta (Tr1) was measured in peripheral blood.. Mononuclear leukocytes were obtained from patients with asthma, asthma accompanying EP and CSS. The cells were stimulated with PMA and ionomycin in the presence of brefeldin A to generate and accumulate cytokines. Cytokines in the cells were detected after fixation and permeabilization of these cells. Tr1 cells were evaluated as CD4+CD25+ T cells dominantly producing IL-10 and TGF-beta, and nTreg were also evaluated as CD4+CD25+ T cells expressing FOXP3, a master transcriptional factor.. There was a significantly greater number of nTreg in the peripheral blood of EP patients in comparison to both patients with asthma and whole CSS. However, no significant difference was observed regarding the number of nTreg when patients with CSS were divided into inactive and active stage. In contrast, the detection frequency of Tr1 remarkably decreased in active CSS, especially at the time of onset of CSS, in comparison with asthma, EP and inactive CSS, while there was no difference between EP and inactive CSS. Additionally, the ability of CD4+CD25- T cells (responder T cells) to generate IL-2 when stimulated with PMA and ionomycin sharply decreased in active CSS.. In the patient who does not develop CSS even after repeat EP, the frequency of Tr1 and the ability of responder T cells to generate IL-2 do not decrease, but show a remarkable decrease in the EP patient who develops CSS. These findings strongly suggest that Tr1 is responsible for maintaining immunological tolerance in peripheral blood, while also inhibiting the onset of CSS from EP.

Topics: Adult; Aged; Aged, 80 and over; Asthma; Cell Count; Churg-Strauss Syndrome; Female; Humans; Immunologic Tests; Interleukin-10; Interleukin-2; Male; Middle Aged; Pulmonary Eosinophilia; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

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

There is a paradoxical finding that eosinophils are frequently accumulated at the sites of allergic inflammation where transforming growth factor (TGF)-beta, a negative regulator of eosinophil survival, is upregulated; however, eosinophil accumulation is persistent. We thus hypothesized that eosinophils might have aberrant TGF-beta signaling and be unresponsive to TGF-beta. To test the hypothesis, we examined the expression and function of Smad proteins, which are central mediators for TGF-beta signaling, in human eosinophils.. Eosinophils were isolated from the peripheral blood of normal donors, and the expression and activation of endogenous Smad proteins were examined by reverse transcription polymerase chain reaction and Western blotting. The Smad function in the transcription of the major TGF-beta target gene Smad7 was investigated using a dominant negative form of Smad3. The effect of TGF-beta on eosinophil survival was then evaluated by a cell viability assay using normal and asthmatic eosinophils.. Human eosinophils expressed mRNAs and proteins of TGF-beta typeI and type II receptors, Smad2, Smad3 and Smad4. TGF-beta induced the phosphorylation of Smad2 in eosinophils, which was blocked by SB431542, an inhibitor of TGF-beta type I receptor kinase. A dominant negative Smad3 protein suppressed TGF-beta-induced Smad7 mRNA expression in eosinophils. Finally, TGF-beta prevented granulocyte macrophage colony-stimulating factor- or interferon-gamma-mediated survival of eosinophils obtained from asthmatic patients as well as normal subjects.. Human eosinophils have an intact Smad signaling pathway leading to a major TGF-beta target gene expression. Thus, eosinophils might become resistant to TGF-beta only in in vivo circumstances.

Topics: Activin Receptors, Type I; Asthma; Cell Survival; Cells, Cultured; Eosinophils; Gene Expression Regulation; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Interferon-gamma; Protein Serine-Threonine Kinases; Receptor, Transforming Growth Factor-beta Type I; Receptor, Transforming Growth Factor-beta Type II; Receptors, Transforming Growth Factor beta; RNA, Messenger; Smad2 Protein; Smad3 Protein; Smad4 Protein; Smad7 Protein; Transforming Growth Factor beta

Airway wall remodelling and inflammation are features of chronic asthma. Transforming growth factor beta (TGF-beta) has been implicated in these processes.. To determine the effect of allergen challenge on airway inflammation and remodelling and whether TGF-beta isoforms and the Smad signalling pathways are involved.. Thirteen patients with atopic asthma underwent inhalational challenge with 0.9% saline, followed by allergen 3-4 weeks later. After both challenges, fibreoptic bronchoscopy was undertaken to obtain bronchial biopsies and tissue samples were processed for immunohistochemistry and examined by microscopy.. Forced expiratory volume in 1 s (FEV(1)) fell after allergen challenge (mean (SE) -28.1 (0.9)% at 30 min with a late response at 7 hours (-23.0 (1.2)%). Allergen challenge caused an increase in neutrophils and eosinophils in the bronchial mucosa compared with saline. Sub-basement membrane (SBM) thickness did not change after allergen, but tenascin deposition in SBM was increased. Intranuclear (activated) Smad 2/3 and Smad 4 detected by immunohistochemistry were increased after allergen challenge in epithelial and subepithelial cells of bronchial biopsies. No inhibitory Smad (Smad 7) protein was detected. TGF-beta isoforms 1, 2 and 3 were expressed predominantly in bronchial epithelium after saline and allergen challenges, but only TGF-beta(2) expression was increased after allergen. Double immunostaining showed an increase in TGF-beta(2) positive eosinophils and neutrophils but not in TGF-beta(1) positive eosinophils and neutrophils after allergen challenge.. TGF-beta(2) may contribute to the remodelling changes in allergic asthma following single allergen exposure.

Topics: Adult; Allergens; Asthma; Biopsy; Bronchi; Bronchitis; Bronchoscopy; Chronic Disease; Eosinophils; Fiber Optic Technology; Forced Expiratory Volume; Humans; Neutrophils; Respiratory Mucosa; Transforming Growth Factor beta

Our purpose was to determine whether numbers of CD4(+)CD25(+) T [T regulatory (T(reg))] cells and mRNA expression of functional molecules of T(reg) are related to airway allergy and disease severity in 51 paediatric patients with allergic rhinitis or bronchial asthma and 47 healthy controls. Surface markers were evaluated with flow cytometry, and mRNA was determined with real-time polymerase chain reaction. Children with allergic disease had fewer CD4(+)CD25(+) T cells (8 x 49% +/- 2 x 41% versus 9 x 58% +/- 2 x 43%, P<0 x 05) and CD4(+)CD25(hi) T cells (1 x 32% +/- 0 x 68% versus 1 x 70% +/- 0 x 68%, P<0 x 01) than control subjects. Numbers of CD4(+)CD25(+) and CD4(+)CD25(hi) T lymphocytes were higher in children with persistent allergic rhinitis and/or moderate-severe bronchial asthma than in those with respective milder disease. The number of T(reg) cells was correlated positively with total immunoglobulin E level. The mRNA expression of forkhead box P3 (FoxP3) was increased in moderate-severe versus mild asthma (2 x 93 +/- 0 x 38 versus 1 x 60 +/- 0 x 31, P< 0 x 01). Patients with moderate-severe bronchial asthma also had increased mRNA expression of interleukin (IL)-10 compared with patients with mild asthma (15 x 24 +/- 4 x 07 versus 3 x 77 +/- 2 x 18, P<0 x 01). The suppressive function of T(reg) cells from patients with more severe asthma was competent in vitro. On average, decreased numbers of T(reg) cells in children with allergic airway disease might represent a defect of the T(reg) population. With increased expression of FoxP3 and IL-10 in T(reg) from patients with relatively severe allergic disease, adaptive and functional T(reg) might be generated in response to aggravated atopy and disease severity.

Topics: Antigens, CD; Antigens, Differentiation; Asthma; Cell Proliferation; Cells, Cultured; Child; Child, Preschool; CTLA-4 Antigen; Female; Forkhead Transcription Factors; Gene Expression; Glucocorticoid-Induced TNFR-Related Protein; Humans; Immune Tolerance; Immunoglobulin E; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Lymphocyte Count; Male; Polymerase Chain Reaction; Receptors, Nerve Growth Factor; Receptors, Tumor Necrosis Factor; Rhinitis; RNA, Messenger; Severity of Illness Index; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

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

Cumulative evidence suggests the up-regulation of interleukin (IL)-10 and T-regulatory (Treg) cells is implicated in anti-inflammatory effect of heme oxygenase-1 (HO-1). Thus, we postulated that induction of HO-1 could augment IL-10 and transforming growth factor (TGF)-beta production and foxp3+CD4+CD25+ Treg cell function, thereby leading to attenuation of airway inflammation. In this study, CD4+CD25+ Treg cells isolated from mouse spleen were either transfected with a HO-1 expression vector (pcDNA3HO-1) or treated with a HO-1 inducer (hemin). Up-regulation of HO-1 enhanced foxp3 expression and IL-10 secretion in the Treg cells in vitro. Next, BALB/c, C57/B6.129, and IL-10-deficient B6.129P2-Il10tm1Cgn/J mice were challenged by ovalbumin to induce airway inflammation. Consistent with in vitro findings, hemin treatment resulted in induction of HO-1 and foxp3 and production of IL-10 and membrane-bound TGF-beta1 in vivo. This was further correlated with decrease of ovalbumin-specific immunoglobulin E level and eosinophil infiltration in bronchial alveolar lavage fluid from the asthmatic mice. Furthermore, hemin significantly enhanced the biological activity of CD4+CD25+ Treg cells. This protective effect was specifically blocked by Sn-protoporphyrin, a HO-1 enzymatic inhibitor. Finally, hemin failed to up-regulate the function of CD4+CD25+ Treg cells from IL-10-deficient mice. Our study indicates that HO-1 exerts its protective effect on asthma through a mechanism mediated by foxp3+CD4+CD25+ Treg cells, IL-10, and membrane-bound TGF-beta1.

Topics: Animals; Asthma; CD4 Antigens; Female; Forkhead Transcription Factors; Heme Oxygenase-1; Hemin; Immunoglobulin E; Inflammation; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Lung; Metalloporphyrins; Mice; Mice, Inbred Strains; Ovalbumin; Protoporphyrins; Spleen; T-Lymphocytes, Regulatory; Transcription, Genetic; Transforming Growth Factor beta

Although animal models with ovalbumin have been used to study chronic asthma, there are difficulties in inducing recurrence as well as in maintaining chronic inflammation in this system. Using a murine model of house dust mite (HDM)-induced bronchial asthma, we examined the airway remodeling process in response to the chronic exposure to HDM. During the seventh and twelfth weeks of study, HDM were inhaled through the nose for three consecutive days and airway responsiveness was measured. Twenty-four hours later, bronchoalveolar lavage and histological examination were performed. The degree of overproduction of mucus, subepithelial fibrosis, and the thickness of the peribronchial smooth muscle in the experimental group was clearly increased compared to the control group. In addition, HDM-exposed mice demonstrated severe airway hyperreactivity to methacholine. In the bronchoalveolar lavage fluid, the number of total cells and eosinophils was increased; during the twelfth week, the number of neutrophils increased in the experimental group. With regard to changes in cytokines, the concentrations of IL-4, IL- 13, and transforming growth factor-beta (TGF-beta) were increased in the experimental group. The data suggest that eosinophils, IL-4, IL-13, and TGF-beta might play an important role in the airway remodeling process and that neutrophils may be involved with increased exposure time.

Topics: Animals; Asthma; Eosinophils; Female; Immunoglobulin E; Immunoglobulin G; Inflammation; Interleukin-13; Interleukin-4; Lung; Mice; Mice, Inbred BALB C; Pyroglyphidae; Transforming Growth Factor beta

Airway remodeling describes the structural changes that occur in the asthmatic airway that include airway smooth muscle hyperplasia, increases in vascularity due to angiogenesis, and thickening of the basement membrane. Our aim in this study was to examine the effect of transforming growth factor-beta on the release of connective tissue growth factor and vascular endothelial growth factor from human airway smooth muscle cells derived from asthmatic and nonasthmatic patients. In addition we studied the immunohistochemical localization of these cytokines in the extracellular matrix after stimulating bronchial rings with transforming growth factor-beta. Connective tissue growth factor and vascular endothelial growth factor were released from both cell types and colocalized in the surrounding extracellular matrix. Prostaglandin E2 inhibited the increase in connective tissue growth factor mRNA but augmented the release of vascular endothelial growth factor. Matrix metalloproteinase-2 decreased the amount of connective tissue growth factor and vascular endothelial growth factor, but not fibronectin deposited in the extracellular matrix. This report provides the first evidence that connective tissue growth factor may anchor vascular endothelial growth factor to the extracellular matrix and that this deposition is decreased by matrix metalloproteinase-2 and prostaglandin E2. This relationship has the potential to contribute to the changes that constitute airway remodeling, therefore providing a novel focus for therapeutic intervention in asthma.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Asthma; Bronchi; Case-Control Studies; Collagen; Connective Tissue Growth Factor; Dinoprostone; Extracellular Matrix; Female; Fibronectins; Humans; Immediate-Early Proteins; Immunoprecipitation; In Vitro Techniques; Intercellular Signaling Peptides and Proteins; Male; Matrix Metalloproteinase 2; Middle Aged; Muscle, Smooth; Recombinant Proteins; Tissue Distribution; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

Transforming growth factor (TGF)-beta and connective tissue growth factor may be implicated in extracellular matrix protein deposition in asthma. We have recently reported that TGF-beta increased connective tissue growth factor expression in airway smooth muscle cells isolated from patients with asthma. In this study, we examined fibronectin and collagen production and signal transduction pathways after stimulation with TGF-beta and connective tissue growth factor. In both asthmatic and nonasthmatic airway smooth muscle cells, TGF-beta and connective tissue growth factor led to the production of fibronectin and collagen I. Fibronectin and collagen expression was extracellular regulated kinase-dependent in both cell types but phosphoinositide-3 kinase-dependent only in asthmatic airway smooth muscle cells. p38 was implicated in fibronectin but not collagen expression in both cell types. TGF-beta induction of fibronectin and collagen was in part mediated by an autocrine action of connective tissue growth factor. Phosphorylation of SMAD-2 may represent an additional pathway because this was increased in asthmatic cells. Our results suggest that these two cytokines may be important in the deposition of extracellular matrix proteins and that the signal transduction pathways may be different in asthmatic and nonasthmatic cells.

Topics: Adolescent; Adult; Aged; Asthma; Cells, Cultured; Collagen; Connective Tissue Growth Factor; Extracellular Matrix Proteins; Female; Fibronectins; Humans; Immediate-Early Proteins; Intercellular Signaling Peptides and Proteins; Male; Middle Aged; Myocytes, Smooth Muscle; Signal Transduction; Transforming Growth Factor beta

The C-509T polymorphism of TGF-beta(1) gene has been associated with asthma and atopy in white populations.. We investigated the association between asthma and previously identified polymorphisms at C-509T and T869C of the TGF-beta(1) gene among 250 Chinese patients with asthma and 308 healthy controls in Hong Kong.. Genotyping was performed on peripheral blood genomic DNA by using PCR-RFLP.. There were no differences in the frequencies of genotypes and alleles between patients and controls. The C-509T and T869C polymorphisms were in tight linkage disequilibrium (P < .01). Among atopic subjects, significant differences were found in genotype and allele frequencies for T869C polymorphism between patients and controls (P = .014 and P = .019, respectively), and individuals bearing the CC genotype were associated with increased risk for the development of asthma (odds ratio, 2.58; 95% CI, 1.17-5.66; P = .018) after adjusting for age, sex, and smoking status. Individuals with asthma bearing the CT genotype of the C-509T polymorphism had significantly increased risk for severe airflow obstruction compared with individuals who had mild obstruction (odds ratio, 4.00; 95% CI, 1.06-15.08; P = .035).. Our results indicate that the polymorphisms at C-509T and T869C of the TGF-beta(1) gene are associated with asthma susceptibility in atopic subjects of the Hong Kong Chinese population, and the C-509T polymorphism may play a role in the pathogenesis of airflow obstruction.

Topics: Adult; Aged; Asthma; Female; Gene Frequency; Genetic Predisposition to Disease; Humans; Logistic Models; Male; Middle Aged; Polymorphism, Genetic; Transforming Growth Factor beta; Transforming Growth Factor beta1

Allergic asthma typically shows activated, allergen-specific CD4(+) T cells in the early phase and airway remodeling in the late phase of the disease. Although TGF-beta plays a crucial role in airway remodeling, it is only marginally induced in CD4(+) T cells in the early allergen-dependent activation of the immune system.. To elucidate the transition between early- and late-phase events, we investigated the role of activin A, a close family member of TGF-beta.. Activin A and TGF-beta(1) levels were measured systemically in the serum and in CD4(+) T cells of asthmatic patients, as well as locally in the lung.. Activin A serum levels were increased in patients with severe asthma compared with levels in patients with moderate asthma and healthy control subjects, whereas all patients showed significantly increased TGF-beta(1) serum levels independent of disease severity. In T cells only patients with moderate asthma showed increased activin A mRNA expression, whereas TGF-beta(1) expression was equal to that seen in healthy subjects. Accordingly, ovalbumin sensitization in a mouse model of allergic asthma could induce activin A mRNA expression, but not TGF-beta(1) expression, in the lung. Immunohistochemistry of mice and human specimens revealed an abundant expression of activin A by infiltrating lymphocytes and structural cells of the lung. Although TGF-beta(1) more potently enhanced proliferation and Smad 2/3-dependent reporter genes in fibroblasts, activin A was capable of inducing TGF-beta(1) and vice versa.. Activin A provides a link between acute allergen-specific T-cell responses and chronic TGF-beta(1)-mediated airway remodeling in asthma.

Topics: Activins; Animals; Asthma; CD4-Positive T-Lymphocytes; Humans; Immunohistochemistry; Inhibin-beta Subunits; Lymphocyte Activation; Mice; Mice, Inbred BALB C; RNA, Messenger; Transforming Growth Factor beta; Transforming Growth Factor beta1

Airway remodeling in asthma is the result of increased expression of connective tissue proteins, airway smooth muscle cell (ASMC) hyperplasia and hypertrophy. TGF-beta1 has been found to increase ASMC proliferation. The activation of mitogen-activated protein kinases (MAPKs), p38, ERK, and JNK, is critical to the signal transduction associated with cell proliferation. In the present study, we determined the role of phosphorylated MAPKs in TGF-beta1 induced ASMC proliferation.. Confluent and growth-arrested bovine ASMCs were treated with TGF-beta1. Proliferation was measured by [3H]-thymidine incorporation and cell counting. Expressions of phosphorylated p38, ERK1/2, and JNK were determined by Western analysis.. In a concentration-dependent manner, TGF-beta1 increased [3H]-thymidine incorporation and cell number of ASMCs. TGF-beta1 also enhanced serum-induced ASMC proliferation. Although ASMCs cultured with TGF-beta1 had a significant increase in phosphorylated p38, ERK1/2, and JNK, the maximal phosphorylation of each MAPK had a varied onset after incubation with TGF-beta1. TGF-beta1 induced DNA synthesis was inhibited by SB 203580 or PD 98059, selective inhibitors of p38 and MAP kinase kinase (MEK), respectively. Antibodies against EGF, FGF-2, IGF-I, and PDGF did not inhibit the TGF-beta1 induced DNA synthesis.. Our data indicate that ASMCs proliferate in response to TGF-beta1, which is mediated by phosphorylation of p38 and ERK1/2. These findings suggest that TGF-beta1 which is expressed in airways of asthmatics may contribute to irreversible airway remodeling by enhancing ASMC proliferation.

Topics: Animals; Antibodies; Asthma; Blood Proteins; Cattle; Cell Division; Cells, Cultured; Drug Synergism; Epidermal Growth Factor; Extracellular Signal-Regulated MAP Kinases; Fibroblast Growth Factor 2; Insulin-Like Growth Factor I; JNK Mitogen-Activated Protein Kinases; MAP Kinase Signaling System; Myocytes, Smooth Muscle; p38 Mitogen-Activated Protein Kinases; Phosphorylation; Platelet-Derived Growth Factor; Trachea; Transforming Growth Factor beta; Transforming Growth Factor beta1

Activated fibroblasts, which have previously been obtained from bronchoalveolar lavage fluid (BALF), are proposed to be important cells in the fibrotic processes of asthma and scleroderma (SSc). We have studied the motility for BALF derived fibroblasts in patients with SSc that may explain the presence of these cells in the airway lumen. Furthermore, we have compared phenotypic alterations in activated fibroblasts from BALF and bronchial biopsies from patients with mild asthma and SSc that may account for the distinct fibrotic responses.. Fibroblasts were cultured from BALF and bronchial biopsies from patients with mild asthma and SSc. The motility was studied using a cell migration assay. Western Blotting was used to study the expression of alpha-smooth muscle actin (alpha-SMA), ED-A fibronectin, and serine arginine splicing factor 20 (SRp20). The protein expression pattern was analyzed to reveal potential biomarkers using two-dimensional electrophoresis (2-DE) and sequencing dual matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-TOF). The Mann-Whitney method was used to calculate statistical significance.. Increased migration and levels of ED-A fibronectin were observed in BALF fibroblasts from both groups of patients, supported by increased expression of RhoA, Rac1, and the splicing factor SRp20. However, these observations were exclusively accompanied by increased expression of alpha-SMA in patients with mild asthma. Compared to BALF fibroblasts in mild asthma, fibroblasts in SSc displayed a differential protein expression pattern of cytoskeletal- and scavenger proteins. These identified proteins facilitate cell migration, oxidative stress, and the excessive deposition of extracellular matrix observed in patients with SSc.. This study demonstrates a possible origin for fibroblasts in the airway lumen in patients with SSc and important differences between fibroblast phenotypes in mild asthma and SSc. The findings may explain the distinct fibrotic processes and highlight the motile BALF fibroblast as a potential target cell in these disorders.

Topics: Adult; Aged; Asthma; Biopsy; Bronchoalveolar Lavage Fluid; Cell Movement; Female; Fibroblasts; Fibronectins; GTP Phosphohydrolases; Humans; Male; Middle Aged; Myocytes, Smooth Muscle; Phenotype; Proteome; RNA-Binding Proteins; Scleroderma, Systemic; Serine-Arginine Splicing Factors; Transforming Growth Factor beta

Transforming growth factor beta (TGF beta) upregulates a number of smooth muscle specific genes in (myo)fibroblasts. As asthma is characterised by an increase in airway smooth muscle, we postulated that TGFbeta(2) favours differentiation of asthmatic (myo)fibroblasts towards a smooth muscle phenotype.. Primary fibroblasts were grown from bronchial biopsy specimens from normal (n = 6) and asthmatic (n = 7) donors and treated with TGF beta2 to induce myofibroblast differentiation. The most stable genes for normalisation were identified using RT-qPCR and the geNorm software applied to a panel of 12 "housekeeping" genes. Expression of alpha-smooth muscle actin (alpha SMA), heavy chain myosin (HCM), calponin 1 (CPN 1), desmin, and gamma-actin were measured by RT-qPCR. Protein expression was assessed by immunocytochemistry and western blotting.. Phospholipase A2 and ubiquitin C were identified as the most stably expressed and practically useful genes for normalisation of gene expression during myofibroblast differentiation. TGF beta2 induced mRNA expression for all five smooth muscle related transcripts; alpha SMA, HCM and CPN 1 protein were also increased but desmin protein was not detectable. Although there was no difference in basal expression, HCM, CPN 1 and desmin were induced to a significantly greater extent in asthmatic fibroblasts than in those from normal controls (p = 0.041 and 0.011, respectively).. Although TGF beta2 induced the transcription of several smooth muscle related genes, not all were translated into protein. Thus, while TGF beta2 is unable to induce a bona fide smooth muscle cell phenotype, it may "prime" (myo)fibroblasts for further differentiation, especially if the cells are derived from asthmatic airways.

Topics: Adult; Asthma; Bronchi; Cell Differentiation; Cells, Cultured; Female; Fibroblasts; Humans; Immunohistochemistry; Male; Muscle Cells; Muscle, Smooth; Phospholipases A; Phospholipases A2; Reverse Transcriptase Polymerase Chain Reaction; RNA; Transforming Growth Factor beta; Transforming Growth Factor beta2; Ubiquitin C; Up-Regulation

Regulatory T cells, which stimulate or inhibit the effector functions of distinct T cell subsets, are critical in the control of the immune response. We investigated the effect of TGF-beta and IL-10 on T cell subsets according to the Th1/Th2 immune status. Sixty-two patients with asthma and 38 patients with pulmonary tuberculosis were included. Allergy skin tests, tuberculin tests, and chest radiography were performed. The levels of circulating IL-4, IFN-gamma, TGF-beta1, and IL-10 were measured using ELISA. The level of TGF-beta1 was higher in patients with asthma than in those with tuberculosis, but the IL-10 levels were the same between the asthma and tuberculosis groups. Atopy was unrelated to the tuberculin response. The IFN-gamma level was correlated with the IL-10 level, and the level of IL-4 was unrelated to the IL-10 or TGF-beta1 level. The level of IL-10 was higher in the negative tuberculin reactors than in the positive tuberculin reactors among patients with asthma, and TGF-beta1 was higher in the positive tuberculin reactors than in the negative tuberculin reactors among patients with tuberculosis. These results demonstrate that the regulatory effects of circulating TGF-beta and IL-10 on T cell cytokines may be different between Th2-type asthma and Th1 tuberculosis.

Topics: Adult; Asthma; Cytokines; Female; Humans; Interferon-gamma; Interleukin-10; Interleukin-4; Male; Respiratory Function Tests; Skin Tests; Th1 Cells; Th2 Cells; Transforming Growth Factor beta; Tuberculin Test; Tuberculosis

IL-17E is a new TH2 cytokine that promotes airway eosinophilia in mice. IL-17E proinflammatory activity has been proposed to involve induction of cytokine and chemokine production. Recruitment of inflammatory cells may be mediated by tissue-resident cells.. This study aimed to evaluate whether fibroblasts represent a target of IL-17E for the production of eosinophil active mediators in the lung.. Expression of IL-17B receptor (IL-17BR), a receptor for IL-17E, was evaluated by immunofluorescent staining, Western blot, and real-time PCR in human primary lung fibroblasts. Mediator production was analyzed by using real-time PCR and ELISA after stimulation of fibroblasts with IL-17E alone or in combination with TNF-alpha and TGF-beta1. Expression of IL-17E and of eosinophil major basic protein was evaluated by immunohistochemistry in bronchial biopsies from subjects with asthma.. Human primary lung fibroblasts constitutively expressed IL-17BR. IL-17BR mRNA levels were increased in cells stimulated with TNF-alpha and decreased with TGF-beta1. IL-17E slightly upregulated CC chemokine ligand (CCL)-5, CCL-11, GM-CSF, and CXC chemokine ligand (CXCL)-8 mRNA in fibroblasts. Moreover, IL-17E and TNF-alpha synergistically induced GM-CSF and CXCL-8 mRNA. IL-17E also potentiated the upregulation of CXCL-8 transcripts observed with TGF-beta1. In contrast, TGF-beta1 decreased IL-17E-induced CCL-11 mRNA. The capacity of IL-17E to enhance GM-CSF and CXCL-8 responses to TNF-alpha was accompanied by production and secretion of both proteins by lung fibroblasts. Finally, IL-17E was detected in asthma in eosinophil-infiltrated bronchial submucosa.. IL-17E may contribute to the induction and maintenance of eosinophilic inflammation in the airways by acting on lung fibroblasts. This study supports a role for IL-17E in asthma pathophysiology.

Topics: Asthma; Biopsy; Bronchi; Cells, Cultured; Chemokine CCL11; Chemokine CCL5; Chemokines, CC; Chemokines, CXC; Chemotactic Factors, Eosinophil; Chemotaxis, Leukocyte; Cytokines; Eosinophil Major Basic Protein; Eosinophils; Fibroblasts; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Inflammation; Interleukin-17; Lung; Receptors, Interleukin; Receptors, Interleukin-17; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha; Up-Regulation

Allergic asthma is a TH2 cell-driven immunological disease, characterized by eosinophilic inflammation. The cytotoxic agent cyclophosphamide paradoxically augments several immune responses.. We studied the proposal that cyclophosphamide may aggravate airway inflammation in allergic mice, and these features might result from the loss of regulatory T cells.. BALB/c mice were immunized with ovalbumin on days 0 and 14 and challenged with aerosolized ovalbumin from days 21 to 27. Some mice also received cyclophosphamide on days -2 and 12.. In the lungs of cyclophosphamide-treated animals, pronounced worsening of inflammatory features was noted, including increased eosinophil infiltration, epithelial thickness, mucus occlusion, and eosinophil numbers in bronchoalveolar lavage fluid. There was also increased total and ovalbumin-specific serum IgE, increased IL-4 and IL-5 secretion by peritracheal lymph node cells, and reduced lung mRNA expression of IL-10 and TGF-beta in animals treated with cyclophosphamide. The expression of FoxP3, a marker of regulatory T cells, was significantly reduced in lymphoid organs after the second injection of cyclophosphamide, and in the lung tissue after allergen challenge in cyclophosphamide-treated mice. Lung IL-10+CD4+ T cells and cytotoxic T lymphocyte-associated antigen 4+CD4+ T cells were reduced after allergen challenge in cyclophosphamide-treated mice.. Cyclophosphamide worsened features of allergic pulmonary inflammation in this model, in association with increased production of IgE and TH2 cytokines. The reduced expression of FoxP3 and immunosuppressive cytokines by cyclophosphamide is consistent with the possibility that toxicity to regulatory T cells may contribute to the increased inflammation.

Topics: Adjuvants, Immunologic; Allergens; Animals; Asthma; Cyclophosphamide; Cytokines; Disease Models, Animal; Eosinophils; Forkhead Transcription Factors; Immunoglobulin E; Interleukin-10; Lung; Mice; Mice, Inbred BALB C; Ovalbumin; Th2 Cells; Transforming Growth Factor beta

Excessive deposition of extracellular matrix occurs in proximal airways of individuals with asthma, but fibrosis in distal lung has not been observed. Whether differing fibrotic capacities of fibroblasts from these two regions contribute to this variability is unknown.. We compared morphologic and functional characteristics of fibroblasts isolated from proximal airways and distal lung parenchyma to determine phenotypic differences.. Concurrent proximal airway and distal lung biopsies were obtained by bronchoscopy from subjects with asthma to isolate airway and distal lung fibroblasts, respectively. The following characteristics were compared: morphology, proliferation, alpha-smooth muscle actin expression, and synthesis of procollagen type I and eotaxin-1.. Airway fibroblasts (AFs) are morphologically distinct from distal lung fibroblasts (DLFs): they are larger (2.3-fold greater surface area vs. matched DLFs; p = 0.02), stellate in appearance, and with more cytoplasmic projections compared with the spindle-shaped DLFs. AFs synthesized more procollagen type I than did DLFs at baseline (twofold higher; p = 0.003) and after transforming growth factor-beta stimulation (1.4-fold higher; p = 0.02). Similarly, AFs produced more eotaxin-1 than did DLFs at baseline (2.5-fold higher; p = 0.004) and after interleukin-13 stimulation (13-fold higher; p = 0.0001). In contrast, DLFs proliferate more than AFs with serum stimulation (about sixfold greater; p = 0.03). Unstimulated DLFs also expressed more alpha-smooth muscle actin than did corresponding AFs (p = 0.006).. These studies suggest that at least two phenotypes of fibroblast exist in the lung. These phenotypic differences may partially explain the variable responses to injury and repair between proximal airways and distal lung/parenchyma in asthma and other respiratory diseases.

Topics: Actins; Adult; Asthma; Bronchi; Cell Proliferation; Chemokine CCL11; Chemokines, CC; Collagen Type I; Extracellular Matrix; Female; Fibroblasts; Humans; Lung; Male; Middle Aged; Phenotype; Transforming Growth Factor beta

Mast cell microlocalisation within the airway smooth muscle (ASM) bundle is an important determinant of the asthmatic phenotype. We hypothesised that mast cells migrate towards ASM in response to ASM derived chemokines.. Primary ASM cultures from subjects with and without asthma were stimulated with interleukin (IL)-1beta, IL-4, and IL-13 alone and in combination. Mast cell chemotaxis towards these ASM supernatants was investigated, and the chemotaxins mediating migration by using specific blocking antibodies for stem cell factor (SCF) and the chemokine receptors CCR3, CXCR1, 3 and 4 as well as the Gi inhibitor pertussis toxin and the tyrosine kinase inhibitor genistein were defined. The concentrations of CCL11, CXCL8, CXCL10, TGF-beta, and SCF in the supernatants were measured and the effect of non-asthmatic ASM supernatants on the mast cell chemotactic activity of asthmatic ASM was examined.. Human lung mast cells and HMC-1 cells migrated towards Th2 stimulated ASM from asthmatics but not non-asthmatics. Mast cell migration was mediated through the combined activation of CCR3 and CXCR1. CCL11 and CXCL8 expression by ASM increased markedly after stimulation, but was similar in those with and without asthma. ASM supernatants from non-asthmatics inhibited mast cell migration towards the asthmatic ASM supernatant.. Th2 stimulated ASM from asthmatics is chemotactic for mast cells. Non-asthmatic ASM releases a mediator or mediators that inhibit mast cell migration towards stimulated asthmatic ASM. Specifically targeting mast cell migration into the ASM bundle may provide a novel treatment for asthma.

Topics: Asthma; Bronchi; Cell Line; Chemokines; Chemotaxis; Genistein; Humans; Interleukins; Mast Cells; Muscle, Smooth; Pertussis Toxin; Receptors, Chemokine; Stem Cell Factor; Th2 Cells; Transforming Growth Factor beta

To investigate the role of CD4(+)CD25(+) regulatory T cells (Tr cells) in the pathogenesis of asthma in children.. Peripheral blood samples were collected from 20 pediatric patients with asthma, 10 male and 10 female, aged 7 (3-12), and 20 healthy children, 10 male and 10 female, aged 6.5 (2-11). Lymphocytes were isolated. Flow cytometry was used to examine the percentages of CD4(+)CD25(+) regulatory T cells, IL-10 secreting CD4(+)CD25(+) regulatory T cells (CD4(+)CD25(+)-IL-10), and transforming growth factor (TGF)-beta secreting CD4(+)CD25(+) regulatory T cells (CD4(+)CD25(+)-TGF-beta). RT-PCR and real-time PCR were used to detect the mRNA expression of suppressor of cytokine signal 1 (SOSC1) and Foxp3.. The percentages of CD4(+)CD25(+) regulatory T cells of the asthma children was 6.51% +/- 1.94%, significantly lower than that of the healthy children (11.96% +/- 2.30%, P < 0.01); the percentage of CD4(+)CD25(+)-IL-10 of the asthma children was 1.46% +/- 0.35%, significantly lower than that of the healthy children (5.65% +/- 1.70%, P < 0.01); and the percentage of CD4(+)CD25(+)-TGF-beta of the asthma children was 1.24% +/- 0.21%, significantly lower than that t of the healthy children (4.23% +/- 1.65%, P < 0.01). The Foxp3 mRNA expression of the asthma children was 0.12 +/- 0.05, significantly lower than that of the healthy children (1.71 +/- 0.58, P < 0.01); and the SOCS1 mRNA expression of the asthma children was 0.38 +/- 0.19, significantly lower than that of the healthy children (1.51 +/- 0.41, P < 0.01).. The decrease of CD4(+)CD25(+) regulatory T cells may be involved in the pathogenesis of asthma. The decreased mRNA expression of Foxp3 and SOCS1 may be associated with the aberrant development of CD4(+)CD25(+) regulatory T cells.

Topics: Asthma; Child; Child, Preschool; Female; Flow Cytometry; Forkhead Transcription Factors; Humans; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Male; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Suppressor of Cytokine Signaling 1 Protein; Suppressor of Cytokine Signaling Proteins; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Airway remodeling is an important feature of chronic asthma that causes irreversible airflow obstruction. Although asthma is considered to be a Th2 disease, the role of T-bet and GATA-3, the key transcription factors for differentiation toward Th1 and Th2 cells, in the pathogenesis of airway remodeling is poorly understood.. We therefore examined the effects of GATA-3 or T-bet induction of Th1/Th2 bias on the development of airway remodeling in mice.. The development of airway remodeling after repeated allergen challenges was analyzed using transgenic mice overexpressing either GATA-3 or T-bet.. The degrees of subepithelial fibrosis and airway smooth muscle hyperplasia after repeated allergen exposure were significantly enhanced in mice overexpressing GATA-3, compared with wild-type mice. Allergen-induced goblet cell hyperplasia and mucus hypersecretion were significantly lower in mice overexpressing T-bet than in wild-type mice. Eosinophilic airway inflammation increased in mice overexpressing GATA-3, but decreased in mice overexpressing T-bet after repeated allergen exposure. Cytokine analysis revealed that the Th1/Th2 cytokine balance shifted to Th2 in lung homogenates and lung T cells of mice overexpressing GATA-3, whereas this balance shifted to Th1 in those of mice overexpressing T-bet after allergen exposure. Lung transforming growth factor-beta and eotaxin levels were associated with the degree of subepithelial fibrosis and eosinophilic airway inflammation, respectively.. Overall, the results indicate that development of airway remodeling is regulated by the lung Th1/Th2 bias induced by GATA-3 and T-bet.

Topics: Animals; Asthma; Chemokine CCL11; Chemokines, CC; Disease Models, Animal; Eosinophils; Fibrosis; GATA3 Transcription Factor; Goblet Cells; Hyperplasia; Hypertrophy; Immunoglobulins; Interferon-gamma; Interleukin-4; Lung; Mice; Mice, Transgenic; Mucins; Muscle, Smooth; T-Box Domain Proteins; Th1 Cells; Th2 Cells; Transcription Factors; Transforming Growth Factor beta

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

Transforming growth factor (TGF)-beta is a multifunctional regulator of cell growth and differentiation with both pro- and anti-inflammatory properties. We used an inhibitor of TGF-beta receptor I (TGF-betaRI) kinase, SD-208 (2,4-disubstituted pteridine, a ATP-competitive inhibitor of TGF-betaRI kinase), to determine the role of TGF-beta in airway allergic inflammation and remodeling. Brown-Norway rats sensitized and repeatedly exposed to ovalbumin (OVA) aerosol challenge were orally administered SD-208 twice daily, before each of six OVA exposures to determine the preventive effects, or only before each of the last three of six OVA exposures to investigate its reversal effects. SD-208 (60 mg/kg) reversed bronchial hyperresponsiveness (BHR) induced by repeated allergen exposure, but it did not prevent it. SD-208 prevented changes in serum total and OVA-specific IgE, but it did not reverse them. SD-208 had both a preventive and reversal effect on airway inflammation as measured by major basic protein-positive eosinophils and CD2(+) T-cell counts in mucosal airways, cell proliferation measured by 5-bromo-2'-deoxyuridine expression in airway smooth muscle (ASM) cells and epithelial cells, and goblet cell hyperplasia induced by repeated allergen challenges. There was a significant decrease in intracellular Smad2/3 expression. SD-208 did not significantly decrease the increased ASM thickness induced by allergen exposure. These findings support a proinflammatory and proremodeling role for TGF-beta in allergic airway inflammation. Inhibition of TGF-betaRI kinase activities by SD-208 may be a useful approach to the reversal of BHR and to the prevention and reversal of inflammatory and remodeling features of chronic asthma.

Topics: Acetylcholine; Activin Receptors, Type I; Animals; Asthma; Bromodeoxyuridine; Bronchi; Chronic Disease; Dose-Response Relationship, Drug; Female; Immunoglobulin E; Muscle, Smooth; Ovalbumin; Protein Serine-Threonine Kinases; Pteridines; Rats; Rats, Inbred BN; Receptor, Transforming Growth Factor-beta Type I; Receptors, Transforming Growth Factor beta; Smad2 Protein; Smad3 Protein; Transforming Growth Factor beta

The role of bacterial enterotoxins like Staphylococcus aureus enterotoxin B (SEB) in allergic asthma remains unknown. We used a mouse model of airway allergy to study the effects of nasal or bronchial contact with SEB on bronchial allergic inflammation.. The features of allergic asthma were induced in ovalbumin (OVA)-sensitized mice (days 1-13) by repeated exposures to nebulized OVA (days 33-37). Nasal or bronchial application of SEB was performed on three occasions (days 33-35-37), and the effects on bronchial inflammation, IgE titres and expression levels of mRNA for T helper type 2 cytokines and other inflammatory mediators were evaluated.. Both nasal and bronchial SEB enhanced the allergen-induced bronchial inflammation, as reflected by more eosinophilic inflammation in the airway lumen and in bronchial tissue. Aggravation of experimental asthma correlated with higher expression of mRNA for IL-5, IL-4, IFN-gamma, IL-12 p40, eotaxin-1 and TGF-beta in bronchi. In addition, nasal SEB elevated concentrations of IL-4, IL-5 and IFN-gamma in serum and bronchial SEB increased titres of OVA-specific and total IgE in serum.. Our data illustrate the potential of both nasal as well as bronchial SEB to aggravate several features of allergic asthma in a mouse model.

Topics: Acute Disease; Animals; Antigens, Bacterial; Asthma; Bronchi; Chemokine CCL11; Chemokines, CC; Cytokines; Enterotoxins; Eosinophilia; Immunoglobulin E; Interferon-gamma; Interleukin-12; Interleukin-4; Interleukin-5; Male; Mice; Mice, Inbred BALB C; Models, Animal; Nasal Mucosa; Ovalbumin; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Staphylococcal Infections; Th2 Cells; Transforming Growth Factor beta

Topics: Adult; Analysis of Variance; Asthma; Cell Differentiation; Cells, Cultured; Chemokine CCL5; Cytokines; Dexamethasone; Female; Humans; Interleukin-10; Interleukin-8; Male; Middle Aged; Sputum; Transforming Growth Factor beta; Transforming Growth Factor beta1

Heme oxygenase-1 (HO-1) has anti-inflammatory effects in asthma. CD4+CD25(high) regulatory T cells (Treg) are a potent immunoregulator that suppresses the immune response. We studied the effects of HO-1-mediated CD4+CD25(high) Treg on suppression of allergic airway inflammation by comparing mice treated with hemin, OVA, Sn-protoporphyrin (SnPP), and hemin plus SnPP. Airway responsiveness, airway eosinophil infiltration, the level of OVA-specific IgE, and the numbers of cells in general and eosinophils in particular in bronchial alveolar lavage fluid were lower in the hemin group than in the OVA, SnPP, and hemin plus SnPP groups. The expressions of HO-1 mRNA and protein in the lung were increased by repeated administrations of hemin and SnPP. However, the activity of HO-1 was highest in hemin mice. The percentage and suppressive function of CD4+CD25(high) Treg and the expression of Foxp3 mRNA were obviously enhanced after treatment with hemin. This increase was diminished by the administration of SnPP. The concentration of serum IL-10 was higher in the hemin group than in the other groups, whereas the level of serum TGF-beta did not significantly differ across groups. Furthermore, the ratio of IFN-gamma/IL-4 mRNA in the lung was higher in hemin-treated mice than in OVA and SnPP mice. The suppressive capacity of CD4+CD25(high) Treg was not enhanced in the IL-10-deficient mice treated with hemin. In conclusion, our experiments in the animal model demonstrated that HO-1 has anti-inflammatory effects, probably via enhancement of the secretion of IL-10 and promotion of the percentage of CD4+CD25(high) Treg.

Topics: Animals; Asthma; CD4 Antigens; Disease Models, Animal; Eosinophils; Female; Forkhead Transcription Factors; Heme Oxygenase-1; Hemin; Hypersensitivity; Immunoglobulin E; Inflammation; Interferon-gamma; Interleukin-10; Interleukin-2 Receptor alpha Subunit; Interleukin-4; Lung; Metalloporphyrins; Mice; Mice, Inbred BALB C; Ovalbumin; Protoporphyrins; T-Lymphocytes, Regulatory; Transforming Growth Factor beta

Allergic asthma is characterized by airway hyperreactivity (AHR), eosinophilic airway inflammation and elevated serum IgE levels. T-helper 2 (Th2) cells play a critical role in the pathogenesis of asthma, but the immunological mechanisms that inhibit Th2 cell function in vivo are not well understood. Conflicting results regarding the protective role of Th1 cytokines and TGF-beta in asthma have been reported. To further investigate the role of TGF-beta(1 )in asthma, we examined mice heterozygous for deletion of the TGF-beta(1) gene (TGF-beta(1) (+/-) mice) in a murine asthma model. While TGF-beta(1) (+/-) mice seem phenotypically normal, they express only about 30% of wild type TGF-beta(1) protein levels as shown before. The reduced expression of TGF-beta(1) is accompanied by a strikingly increased eosinophilic inflammation and mucus secretion in response to ovalbumin (OVA) sensitization. Moreover, TGF-beta(1) (+/-) mice develop significantly enhanced Th2-cytokine levels, decreased IFN-gamma production and increased levels of OVA-specific IgE in serum. In contrast, AHR in response to methacholine is not altered significantly. Our data demonstrate that reduced expression of TGF-beta(1) exacerbates pathology in an experimental asthma model and support the view that the elevated levels of TGF-beta(1) in asthmatic airways might be, at least in part, a result of anti-inflammatory compensation by this cytokine.

Topics: Animals; Asthma; Cytokines; Disease Models, Animal; Female; Heterozygote; Immunization; Immunoglobulins; Inflammation; Interferon-gamma; Lung; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Mucus; Ovalbumin; Respiratory Hypersensitivity; Th2 Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1

Airway eosinophilia and thickened subepithelial basement membrane have previously been reported to increase with increases in TGF-beta expression. However, little is known regarding the expression of specific TGF-beta isoforms (TGF-beta1, TGF-beta2, and TGF-beta3) in asthma, despite recent evidence suggesting that isoforms may have differing biologic activities.. This study examined airway tissue expression of the 3 TGF-beta isoforms and several downstream pathway elements in 48 patients with severe asthma with or without persistent eosinophilia, 14 patients with mild asthma, and 21 normal subjects.. Immunochemistry/immunofluorescence, quantitative real-time PCR and enzyme immunoassay were used to evaluate the 3 TGF-beta isoforms, their receptors, collagen I deposition, connective tissue growth factor expression, and tissue inhibitor of metalloproteinases 1 levels.. Of the isoforms, only TGF-beta2 was different among the groups and increased in severe asthma (overall P < .0001). The increase was due to severe asthma tissue eosinophils which, unlike eosinophils in other groups, expressed high amounts of TGF-beta2. Subjects with severe asthma also had the thickest subbasement membrane and highest tissue inhibitor of metalloproteinases 1 levels. In contrast, TGF-beta receptor 1 and connective tissue growth factor were both consistently downregulated in asthma, regardless of severity.. TGF-beta2, expressed mainly by eosinophils, is the predominant isoform expressed in severe asthma, and is associated with increased profibrotic responses. Decreased expression of TGF-beta receptor 1 and connective tissue growth factor in all asthma severity groups suggests a degree of activation of the TGF-beta pathway in airway tissue of all asthmatic compared with normal airways.

Topics: Adult; Asthma; Bronchi; Connective Tissue; Eosinophilia; Eosinophils; Female; Growth Substances; Humans; Immunohistochemistry; Male; Middle Aged; Muscle, Smooth; Protein Isoforms; Receptors, Transforming Growth Factor beta; RNA, Messenger; Transforming Growth Factor beta; Transforming Growth Factor beta2

Asthma is a complex disorder of the airways of the lungs. TGF-beta1 plays a key role in airway remodeling and asthma by having both proinflammatory and anti-inflammatory activities, making TGFbeta1 an important candidate gene to study.. To investigate the association of TGFbeta1 gene polymorphisms with asthma.. A case-control study was designed for identifying polymorphisms and haplotypes associated with asthma and associated phenotypes. We have verified our results in 2 independent cohorts collected from northern (number of patients, 187; number of controls, 187) and western India (number of patients, 209; number of controls, 190). We measured the serum TGF-beta1 levels of selected individuals and correlated them with genotypes and haplotypes.. A novel (CT)n(CA)m repeat polymorphism (BV209662) 24.9 kb upstream of TGFbeta1 was identified. A significant association was seen at the level of alleles and genotypes with asthma in the 2 cohorts studied independently (P < .05). Interestingly, a novel 3-locus haplotype, 23_G_T, was found to be significantly associated with asthma (P = .00001 in cohorts A and B) as well as with higher serum TGF-beta1 level (P = .01). On the other hand, a novel haplotype, 22_G_C, was negatively associated with asthma (P = .00001 for cohorts A and B) and with lower serum TGF-beta1 level (P = .0019).. This is the first study identifying novel risk and protective haplotypes--23_G_T and 22_G_C, respectively--in the TGFbeta1 gene that are associated with asthma. We also demonstrate the functional significance of these haplotypes with serum TGF-beta1 levels. These results would be valuable in elucidating the role of TGF-beta1 in asthma pathogenesis.

Topics: Adult; Asthma; Case-Control Studies; Female; Gene Frequency; Haplotypes; Humans; India; Male; Molecular Sequence Data; Polymerase Chain Reaction; Polymorphism, Single Nucleotide; Transforming Growth Factor beta

Airway inflammation and remodelling are important pathophysiologic features of chronic asthma. Although current steroid use demonstrates anti-inflammatory activity, there are limited effects on the structural changes in the lung tissue.. We have used a mouse model of prolonged allergen challenge that exhibits many of the salient features of airway remodelling in order to investigate the anti-remodelling effects of Budesonide.. Treatment was administered therapeutically, with dosing starting after the onset of established eosinophilic airway inflammation and hyper-reactivity.. Budesonide administration reduced airway hyper-reactivity and leukocyte infiltration in association with a decrease in production of the Th2 mediators, IL-4, IL-13 and eotaxin-1. A reduction in peribronchiolar collagen deposition and mucus production was observed. Moreover, our data show for the first time that, Budesonide treatment regulated active transforming growth factor (TGF)-beta signalling with a reduction in the expression of pSmad 2 and the concomitant up-regulation of Smad 7 in lung tissue sections.. Therefore, we have determined that administration of Budesonide modulates the progression of airway remodelling following prolonged allergen challenge via regulation of inflammation and active TGF-beta signalling.

Topics: Allergens; Animals; Anti-Asthmatic Agents; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Budesonide; Chemokine CCL11; Chemokines, CC; Collagen; Extracellular Matrix; Female; Image Processing, Computer-Assisted; Interleukin-13; Interleukin-4; Lymphocyte Activation; Mice; Mice, Inbred BALB C; Models, Animal; Mucus; Signal Transduction; Time Factors; Transforming Growth Factor beta

CD4 + lymphocytes play a key role in asthma pathogenesis, but much remains unknown about the genetic mechanisms that affect disease severity. In this study we sought to investigate global patterns of gene expression in CD4 + lymphocytes isolated from subjects with severe asthma through the use of microarray technology. CD4 + lymphocytes were separated from peripheral blood, total RNA was purified, and biotinylated complementary RNA was prepared and hybridized to Affymetrix HU133 chips (Affymetrix, Santa Clara, Calif). Using the robust multi-chip average procedure, we compared the messenger RNA expression profiles of more than 33,000 genes of CD4 + lymphocytes in subjects with severe ( n = 5) and mild ( n = 5) asthma. Forty genes had 2-fold mean expression differences or greater. Thirty-seven genes were up-regulated, including transforming growth factor-beta and those involved in T-cell activation, proliferation, and cytoskeletal changes. Three genes were down-regulated, including the T-cell-receptor delta locus. This study demonstrates a method by which CD4 + lymphocytes can be extracted from blood for the purpose of microarray analysis. Furthermore, we show that T-lymphocytes from the peripheral blood of subjects with severe and mild asthma differ in their gene-expression profiles, supporting the view that asthma is a systemic disease. These differentially expressed genes identify potential molecular targets for preventive and therapeutic options for severe asthma.

Topics: Adult; Asthma; CD4-Positive T-Lymphocytes; Female; Gene Expression; Gene Expression Profiling; Gene Expression Regulation; Humans; Lymphocyte Activation; Male; Oligonucleotide Array Sequence Analysis; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor beta

Mucosal tolerance prevents pathological reactions against environmental and food antigens, and its failure results in exacerbated inflammation typical of allergies and asthma. One of the proposed mechanisms of oral tolerance is the induction of Tregs. Using a mouse model of hyper-IgE and asthma, we found that oral tolerance could be effectively induced in the absence of naturally occurring thymus-derived Tregs. Oral antigen administration prior to i.p. immunization prevented effector/memory Th2 cell development, germinal center formation, class switching to IgE, and lung inflammation. Oral exposure to antigen induced development of antigen-specific CD4CD25Foxp3CD45RB cells that were anergic and displayed suppressive activity in vivo and in vitro. Oral tolerance to the Th2 allergic response was in large part dependent on TGF-beta and independent of IL-10. Interestingly, Tregs were also induced by single i.p. immunization with antigen and adjuvant. However, unlike oral administration of antigen, which induced Tregs but not effector T cells, i.p. immunization led to the simultaneous induction of Tregs and effector Th2 cells displaying the same antigen specificity.

Topics: Administration, Oral; Animals; Antigens; Asthma; Base Sequence; CD4-Positive T-Lymphocytes; Disease Models, Animal; DNA, Complementary; Immune Tolerance; Immunity, Mucosal; Injections, Intraperitoneal; Job Syndrome; Lung; Mice; Mice, Inbred BALB C; Mice, Knockout; Neutralization Tests; Ovalbumin; T-Lymphocyte Subsets; T-Lymphocytes; Transforming Growth Factor beta

The chromosomal region 19q13 has been found in linkage to allergic diseases in several genome-wide linkage screens. One candidate gene within this region is the gene coding for TGF-beta1. Transforming growth factor (TGF)-beta acts as an anti-inflammatory cytokine suppressing allergic inflammation and hyper-reactivity. However, in ongoing inflammation of the lungs it can induce fibrosis and airway remodelling as seen in chronic asthma. Several polymorphisms within TGF-beta1 have been identified and one, -C509T, has been shown to be in association with elevated immunoglobulin E levels and severe bronchial asthma in different populations. However, other studies failed to confirm the association. The present study investigated two polymorphisms within the gene coding for TGF-beta1, -C509T and G915C, and for their potential association with bronchial asthma in Caucasian children. Genotyping of these polymorphisms was performed by means of restriction fragment length polymorphisms in a population of 231 asthmatic children and a control population of 269 individuals. Statistical analyses made use of the Armitage's trend test. In addition haplotypes were calculated by arlequin. None of the two polymorphisms showed association with bronchial asthma. They were found to be in linkage disequilibrium. We conclude from our data that TGF-beta1 is unlikely to represent a major gene in the development of bronchial asthma in the Caucasian population.

Topics: Adolescent; Asthma; Child; Child, Preschool; Germany; Humans; Polymorphism, Genetic; Transforming Growth Factor beta; White People

Eosinophils may play an important role in the pathogenesis of airway remodeling in asthma through the production of various fibrogenic cytokines such as transforming growth factor-beta1 (TGF-beta1). Cysteinyl leukotrienes are also suggested to be involved in remodeling with their potential to induce proliferation of airway smooth muscle cells. Since massive eosinophil infiltration and the release of cysteinyl leukotrienes in airway secretions are often seen in asthma, we hypothesized that cysteinyl leukotrienes may be involved in airway remodeling through induction of TGF-beta1 from eosinophils. Peripheral blood eosinophils were cultured with leukotriene D(4) (LTD(4)) and/or interleukin-5 (IL-5) or granulocyte colony-stimulating factor (GM-CSF) for 16 h and gene expression of TGF-beta1 was quantified with real-time PCR. A combination of LTD(4) and IL-5 or LTD(4) and GM-CSF synergistically induced TGF-beta1 expression in eosinophils although stimulation with single factor, LTD(4), IL-5 or GM-CSF did not induce the gene expression. LTD(4) also induced significant gene expression in eosinophils cultured in an intercellular adhesion molecule-1-coated plate. The results suggested that CysLTs stimulate eosinophils to induce TGF-beta1 production in allergic inflammation where IL-5 and GM-CSF are abundant and may be involved in the pathogenesis of airway remodeling.

Topics: Asthma; Eosinophils; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Interleukin-5; Leukotriene D4; Reverse Transcriptase Polymerase Chain Reaction; Transforming Growth Factor beta; Transforming Growth Factor beta1; Up-Regulation

Mice with a targeted deletion of the T-bet gene exhibit spontaneous airway hyperresponsiveness (AHR), airway inflammation, enhanced recovery of T(h)2 cytokines from bronchoalveolar lavage fluid, sub-epithelial collagen deposition and myofibroblast transformation. Here we analyze the mechanisms responsible for the chronic airway remodeling observed in these mice. CD4+ T cells isolated from the lung of T-bet-deficient mice were spontaneously activated CD44(high)CD69(high) memory T cells, with a typical T(h)2 cytokine profile. Neutralization of IL-13 but not IL-4 resulted in amelioration of AHR in airways of mice lacking T-bet. IL-13 blockade also led to reduced eosinophilia and decreased vimentin, transforming growth factor beta (TGF-beta) and alpha smooth muscle actin (alphaSMA) levels. T-bet(-/-) lung fibroblasts proliferated very rapidly and released increased amounts of TGF-beta. Interestingly, neutralization of TGF-beta ameliorated aspects of the chronic airway remodeling phenotype but did not reduce AHR. These data highlight a T-bet-directed function for IL-13 in controlling lung remodeling that is both dependent on and independent of its interaction with TGF-beta in the asthmatic airway.

Topics: Actins; Animals; Asthma; CD4-Positive T-Lymphocytes; Cells, Cultured; Cytokines; DNA-Binding Proteins; Fibroblasts; Immunologic Memory; Interleukin-13; Interleukin-4; Lung; Lymphocyte Activation; Mice; Mice, Inbred C57BL; Mice, Knockout; Smad3 Protein; Smad7 Protein; T-Box Domain Proteins; Trans-Activators; Transcription Factors; Transforming Growth Factor beta; Vimentin

Lymphangioleiomyomatosis (LAM) is associated with abnormal airway smooth muscle that leads to the characteristic pathology of lung nodule formation and destruction of lung tissue. The current authors have previously identified abnormal behaviour of airway smooth muscle cells from patients with asthma. In this study, cells and tissue sections derived from patients with LAM (n=7), asthma (n=8), and nonasthmatic controls (n=9) were compared. The presence of the antigen human melanosome (HM)B-45 was investigated, along with the proliferation and release of extracellular matrix proteins, release of endogenous prostaglandin E2 (PGE2), vascular endothelial growth factor and connective tissue growth factor, and the expression of integrins. Positive HMB-45 staining was found in all LAM patients and no controls. Proliferation of LAM cells was not different from control cells nor was its inhibition by beta-agonists, corticosteroids, rapamycin or PGE2. However, endogenous PGE2 levels were markedly decreased in LAM cells, and this was associated with decreased expression of the inducible form of cyclooxygenase (COX-2). The increased levels of connective tissue growth factor seen in asthma cells were not observed in LAM. Elastin mRNA in response to transforming growth factor-beta stimulation was markedly lower in LAM cells than either asthma or control cells. In conclusion, lymphangioleiomyomatosis cells exhibit abnormal properties in vitro that may contribute to pathophysiology and symptomatology in patients with lymphangioleiomyomatosis.

Topics: Adolescent; Adult; Aged; Antigens, Neoplasm; Asthma; Cells, Cultured; Connective Tissue Growth Factor; Cyclooxygenase 2; Dinoprostone; Extracellular Matrix Proteins; Female; Growth Substances; Humans; Immediate-Early Proteins; Integrins; Intercellular Signaling Peptides and Proteins; Lung; Lymphangioleiomyomatosis; Male; Melanoma-Specific Antigens; Middle Aged; Myocytes, Smooth Muscle; Neoplasm Proteins; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A

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

Topics: Animals; Asthma; Bacterial Proteins; Disease Models, Animal; Interferon-gamma; Interleukin-10; Interleukin-12; Lipoproteins; Mice; Mice, Inbred BALB C; Toll-Like Receptor 2; Transforming Growth Factor beta

Transforming growth factor-beta1 (TGF-beta1) is increased in the lungs of individuals with asthma and may modulate airway inflammation and remodeling. Some genetic studies have found that a C-to-T single-nucleotide polymorphism (C-509T) in the TGF-beta1 gene promoter may be associated with altered gene expression and asthma phenotype. To build on these data, we performed a case-control association study at this locus involving 527 subjects with asthma and 170 control subjects without asthma. All individuals were white. Genotyping at 49 unlinked polymorphisms indicated that a subset of case subjects and all control subjects were well matched and without evidence of population stratification. Logistic regression was used to model the effects of age, sex, and genotype on case-control status. The diagnosis of asthma was positively associated with the T allele and TT genotype under a codominant model (odds ratio, 2.98; 95% confidence interval, 1.45 to 6.25; p = 0.003). Total serum IgE, eosinophil count, and FEV1% predicted levels were not associated with this polymorphism. Furthermore, we show that the C-509T polymorphism alters TGF-beta1 promoter-reporter activity and promoter interactions with the transcription factor Yin Yang 1. We conclude that the T allele of C-509T is associated with the diagnosis of asthma and may enhance TGF-beta1 gene transcription.

Topics: Adult; Asthma; Case-Control Studies; DNA-Binding Proteins; Erythroid-Specific DNA-Binding Factors; Female; Gene Frequency; Genotype; Humans; Male; Polymorphism, Single Nucleotide; Promoter Regions, Genetic; Transcription Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1; YY1 Transcription Factor

In addition to its central role in hemostasis, thrombin may play a role in inflammation and remodeling. To investigate the contribution of thrombin to allergic airway inflammation in asthma, we used an enzymatic assay to determine thrombin activity in bronchoalveolar lavage fluid obtained from 19 subjects with atopic asthma before (Day 0) and 48 hours after (Day 2) segmental bronchoprovocation with antigen. Thrombin activity increased from 0 (0, 2.9) on Day 1 to 41.1 (0.3, 75.6) U x 10(-3)/ml on Day 2 (p = 0.002) and correlated with total protein levels in lavage fluid on Day 2 (r = 0.885, p < 0.001). After antigen challenge, thrombin activity also showed significant correlations with interleukin-5 (r = 0.66, p = 0.002), transforming growth factor beta1 (r = 0.70, p < 0.001), fibronectin (r = 0.85, p < 0.001) and tissue factor (r = 0.55, p = 0.03) levels in lavage fluid. Furthermore, Day 2, but not Day 0 lavage fluid, induced proliferation of human airway fibroblasts. This mitogenic effect was significantly reduced with hirudin, a specific thrombin inhibitor. Taken together, our findings suggest that allergen-driven airway inflammation in asthma is associated with enhanced potential for fibroblast proliferation that is related, at least in part, to increased thrombin activity. We propose that enhanced thrombin activity provides a potential link between allergic inflammation and initiation of airway remodeling.

Topics: Adult; Allergens; Asthma; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Cohort Studies; Enzyme-Linked Immunosorbent Assay; Female; Fibronectins; Humans; Inflammation; Inflammation Mediators; Interleukin-5; Male; Middle Aged; Probability; Prospective Studies; Sensitivity and Specificity; Statistics, Nonparametric; Thrombin; Thromboplastin; Transforming Growth Factor beta

Eosinophils accumulate in high numbers in the lungs of asthmatic patients. These cells have the ability to induce tissue damage, a capacity that relates to their traditional role in host defense against parasitic worms. On the other hand, eosinophils produce growth factors associated with tissue repair and remodeling, notably TGF-beta1. The relationship of these activities to lung dysfunction in asthma is highly controversial, but recent observations in humans and in animal models add spice to the debate.

Topics: Animals; Asthma; Eosinophils; Humans; Interleukin-5; Transforming Growth Factor beta; Transforming Growth Factor beta1

Asthma is a chronic inflammatory disease characterized by variable bronchial obstruction, hyperresponsiveness, and by tissue damage known as airway remodeling. In the present study we demonstrate that interleukin (IL)-5 plays an obligatory role in the airway remodeling observed in experimental asthma. BALB/c mice sensitized by intraperitoneal injections of ovalbumin and exposed daily to aerosol of ovalbumin for up to 3 wk, develop eosinophilic infiltration of the bronchi and subepithelial and peribronchial fibrosis. The lesions are associated with increased amounts of hydroxyproline in the lungs and elevated levels of eosinophils and transforming growth factor (TGF)-beta1 in the bronchoalveolar lavage fluid. After 1 wk of allergen challenge, TGF-beta is mainly produced by eosinophils accumulated in the peribronchial and perivascular lesions. At a later stage of the disease, the main source of TGF-beta is myofibroblasts, identified by alpha-smooth muscle actin mAb. We show that all these lesions, including fibrosis, are abolished in sensitized and allergen-exposed IL-5 receptor-null mice, whereas they are markedly accentuated in IL-5 transgenic animals. More importantly, treatment of wild-type mice with neutralizing anti-IL-5 antibody, administered before each allergen challenge, almost completely prevented subepithelial and peribronchial fibrosis. These findings demonstrated that eosinophils are involved in allergen-induced subepithelial and peribronchial fibrosis probably by producing a fibrogenic factor, TGF-beta1.

Topics: Actins; Allergens; Animals; Antibodies; Asthma; Bronchi; Chemotaxis, Leukocyte; Collagen; Disease Models, Animal; Disease Progression; Eosinophils; Female; Fibroblasts; Hydroxyproline; Interleukin-5; Mice; Mice, Knockout; Ovalbumin; Pulmonary Fibrosis; Receptors, Interleukin; Receptors, Interleukin-5; Transforming Growth Factor beta; Transforming Growth Factor beta1; Up-Regulation

Asthmatic airway remodelling is characterized by myofibroblast hyperplasia and subbasement membrane collagen deposition. We hypothesized that cytokines and growth factors implicated in asthmatic airway remodelling are increased in bronchoalveolar lavage (BAL) fluid of asthmatics after segmental allergen challenge (SAC), and that these growth factors and cytokines increase alpha-smooth muscle actin (alpha-SMA) and collagen III synthesis by human lung fibroblasts (HLFs).. Transforming growth factor (TGF)-beta1, TGF-beta2, IL-4 and IL-13 levels were measured in BAL fluid from 10 asthmatics and 9 non-asthmatic controls at baseline and then 1 day, 1 week and 2 weeks after SAC. Confluent cultures of HLFs were stimulated by exogenous addition of TGF-beta1, TGF-beta2, IL-4 or IL-13 (concentration range 0.01-10 ng/mL) over 48 h. Collagen III was measured in culture supernates and alpha-SMA in cell lysates by Western blot.. At baseline, there was no difference in BAL fluid concentrations of TGF-beta1, IL-4 and IL-13 between asthmatics and controls; however, non-asthmatics had higher concentrations of total TGF-beta2. In asthmatics, BAL fluid concentrations of all four factors increased significantly 1 day after SAC. TGF-beta1, TGF-beta2 and IL-13 concentrations returned to baseline by 1 week after SAC, but BAL fluid IL-4 concentration remained elevated for at least 2 weeks. TGF-beta1, TGF-beta2 and IL-4 significantly increased alpha-SMA in fibroblasts, but only IL-4 caused corresponding increases in collagen III synthesis. IL-13 had no direct effects on collagen III synthesis and alpha-SMA expression.. Because IL-4 caused a dose-dependent increase in alpha-SMA and collagen III synthesis, it may be an important cytokine mediating asthmatic airway remodelling. TGF-beta1 and TGF-beta2 may also play a role in airway remodelling by stimulating phenotypic change of fibroblasts to myofibroblasts. Additionally, collagen III synthesis appears to be independent of myofibroblast phenotype and is apparently regulated by different growth factors and cytokines.

Topics: Actins; Adult; Allergens; Analysis of Variance; Asthma; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Case-Control Studies; Cells, Cultured; Collagen Type III; Dose-Response Relationship, Drug; Female; Fibroblasts; Humans; Interleukin-13; Interleukin-4; Lung; Male; Muscle, Smooth; Transforming Growth Factor beta

Inflammatory infiltrates, airway hyper-responsiveness, goblet cell hyperplasia and subepithelial thickening are characteristic of chronic asthma. Current animal models of allergen-induced airway inflammation generally concentrate on the acute inflammation following allergen exposure and fail to mimic all of these features.. The aim of this study was to use a murine model of prolonged allergen-induced airway inflammation in order to characterize the cells and molecules involved in the ensuing airway remodelling. Moreover, we investigated whether remodelling persists in the absence of continued allergen challenge.. Acute pulmonary eosinophilia and airways hyper-reactivity were induced after six serial allergen challenges in sensitized mice (acute phase). Mice were subsequently challenged three times a week with ovalbumin (OVA) (chronic phase) up to day 55. To investigate the persistence of pathology, one group of mice were left for another 4 weeks without further allergen challenge (day 80).. The extended OVA challenge protocol caused significant airway remodelling, which was absent in the acute phase. Specifically, remodelling was characterized by deposition of collagen as well as airway smooth muscle and goblet cell hyperplasia. Importantly, these airway changes, together with tissue eosinophilia were sustained in the absence of further allergen challenge. Examination of cytokines revealed a dramatic up-regulation of IL-4 and tumour growth factor-beta1 during the chronic phase. Interestingly, while IL-4 levels were significantly increased during the chronic phase, levels of IL-13 fell. Levels of the Th1-associated cytokine IFN-gamma also increased during the chronic phase.. In conclusion, we have demonstrated that prolonged allergen challenge results in persistent airway wall remodelling.

Topics: Acute Disease; Allergens; Animals; Asthma; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Chronic Disease; Collagen; Female; Goblet Cells; Hyperplasia; Interferon-gamma; Interleukin-13; Interleukin-4; Mice; Mice, Inbred BALB C; Models, Animal; Muscle, Smooth; Ovalbumin; Pulmonary Eosinophilia; Respiratory System; Time Factors; Transforming Growth Factor beta

Growth factors produced by airway epithelial cells may be important in the pathogenesis of subepithelial fibrosis, a distinctive lesion of chronic human asthma.. To examine the relationship between the development of subepithelial fibrosis and the expression of transforming growth factor-beta 1 (TGF-beta 1) and ligands for the epidermal growth factor receptor.. BALB/c mice sensitized to ovalbumin were chronically challenged by inhalation of low levels of antigen, leading to development of subepithelial fibrosis and other changes of airway wall remodelling. Growth factor expression was assessed by immunohistochemistry and enzyme immunoassay.. Allergic sensitization directly correlated with airway epithelial expression of both the cleaved, potentially biologically active form of TGF-beta 1 and of amphiregulin in response to allergen challenge. Accumulation of TGF-beta 1 was related to remodelling of the airway wall in chronic asthma, whereas expression of amphiregulin did not exhibit a similar relationship. Production of epithelial cell-derived TGF-beta 1 appeared to be regulated by IL-13, while both IL-13 and CD4(+) T cells regulated accumulation of TGF-beta 1. In contrast to results reported in high-level exposure models of airway fibrosis, eosinophils did not appear to be a significant source of TGF-beta 1.. Airway epithelial cell-derived TGF-beta 1 has a potentially crucial role in the development of airway wall remodelling in asthma. Immunological mechanisms may regulate the release and accumulation of TGF-beta 1.

Topics: Allergens; Amphiregulin; Animals; Asthma; Bronchoalveolar Lavage Fluid; CD4-Positive T-Lymphocytes; Chronic Disease; Disease Models, Animal; EGF Family of Proteins; Epidermal Growth Factor; Epithelial Cells; Female; Fibrosis; Glycoproteins; Immunoenzyme Techniques; Intercellular Signaling Peptides and Proteins; Interleukin-13; Ligands; Mice; Mice, Inbred BALB C; Ovalbumin; Respiratory Mucosa; Trachea; Transforming Growth Factor beta; Transforming Growth Factor beta1

Atopic disorders have been associated with a Th-2 cytokine predominance. This study investigated Th1- and Th2-related gene expression in asthmatics, atopics and healthy individuals.. We compared Th1- and Th2-related in vivo-signals using gene expression arrays in 18 atopic asthmatics, 8 atopic non-asthmatic and 14 healthy control subjects. Purified mRNA from peripheral blood mononuclear cells was reverse-transcribed and hybridised to cDNA membranes. Group differences were assessed after standardisation with Mann-Whitney U-test.. Atopic individuals had upregulated lymphotoxin-alpha and downregulated IFNGR1. On the other hand, they had particularly high IL-4, IL-5 and IL4R levels, together with significantly upregulated IL10. Asthmatic individuals had normal Th1-gene expression, but an upregulation og Th-2 genes. Atopic individuals had high, asthmatic individuals excessively high IL12RB1-levels. No Th-2 gene was downregulated in both atopic phenotypes. The expression of IL6R correlated with the daily dose of inhaled corticosteroids.. Atopic individuals had a down regulation of key TH1- and Th2-genes, resulting in a balanced upregulation of Th-specific genes. In contrast, asthmatic subjects had normal Th1-gene expression but a constant upregulation of Th2-specific genes, leading to Th2-predominance.

Topics: Adolescent; Adult; Aged; Asthma; Down-Regulation; Female; Humans; Hypersensitivity; Interleukins; Male; Middle Aged; Oligonucleotide Array Sequence Analysis; Receptors, Cytokine; Th1 Cells; Th2 Cells; Transforming Growth Factor beta; Up-Regulation

The development and expression of allergic rhinitis and asthma may be influenced by the elaboration of specific cytokines. Cytokine genotypes moderate illness severity in a variety of inflammatory disorders. Cytokine genotyping was performed on 124 infants (85% white, 57% male) to determine whether specific cytokine genotypes are associated with a parental history of allergic rhinitis and/or asthma. DNA was extracted from buccal brushings and assayed for tumor necrosis factor alpha (TNF-alpha), interferon gamma (IFN-gamma), interleukin (IL)-6, IL-10, and transforming growth factor (TGF)-beta1 genotypes using polymerase chain reaction-sequence specific primer technology. Outcomes consisted of parental history of allergy and asthma, and results were evaluated by logistic regression. TNF-alpha and TGF-beta genotypes were related to maternal and/or paternal history of allergic rhinitis and asthma, respectively. The frequencies of the genotype associated with high production of TNF-alpha were 41% versus 18% in infants with and without a parental history of allergic rhinitis, respectively (p < 0.01). The frequencies of the genotype associated with low production of TGF-beta1 were 14% versus 1% in infants with and without a parental history of asthma, respectively (p < 0.01). There were no associations between IFN-gamma, IL-6, and IL-10 genotypes and any of the outcome parameters. These results suggest a role for TNF-alpha and TGF-beta1 genotypes in the pathogenesis of allergic rhinitis and asthma, respectively. If confirmed by future studies, cytokine genotyping may be a useful tool for identifying at-risk infants who may benefit from the selective use of preventative and/or early intervention treatments for these disorders.

Topics: Asthma; DNA Fingerprinting; Female; Genetic Predisposition to Disease; Genotype; Humans; Infant; Male; Polymorphism, Genetic; Rhinitis; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Allergic asthma is associated with persistent functional and structural changes in the airways and involves many different cell types. Peroxisome proliferator-activated receptor gamma (PPAR-gamma), a member of the nuclear hormone receptor superfamily, is predominantly expressed in adipose tissue and plays a major role in regulating adipocyte differentiation and glucose metabolism. Recently, PPAR-gamma has been shown to play an important role in the control of inflammatory responses, including within the lung, acting on both immune and nonimmune cells.. Our aim was to assess the anti-inflammatory potential of a PPAR-gamma agonist locally delivered by means of nebulization.. We used a mouse model of asthma induced by sensitization and airway challenge with ovalbumin. Ciglitazone, a PPAR-gamma agonist, was administered by means of nebulization alone at the time of antigen challenge or by means of gavage and nebulization. Treatments with both ciglitazone and GW9662, a specific antagonist, were also performed to verify that ciglitazone's effects were mediated through PPAR-gamma activation.. Our results show that PPAR-gamma is mainly expressed in airway epithelium on antigen sensitization. Treatment with ciglitazone reduced PPAR-gamma levels in the lung, whereas combined treatment with GW9662 abrogated this inhibition. Importantly, nebulization with ciglitazone decreased airway hyperresponsiveness, basement membrane thickness, mucus production, collagen deposition, and TGF-beta synthesis. A significant correlation was also found between airway hyperresponsiveness, basement membrane thickness, and TGF-beta levels.. These results demonstrate that inhaled agonistic ligands of PPAR-gamma might have new therapeutic potential for airway asthmatic inflammation.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Female; Immunohistochemistry; Lung; Mice; Mice, Inbred BALB C; Nebulizers and Vaporizers; Ovalbumin; Receptors, Cytoplasmic and Nuclear; Thiazolidinediones; Trachea; Transcription Factors; Transforming Growth Factor beta

Repeated attacks of bronchial asthma lead to different degrees of airway remodeling, the mechanism of which is not yet clear. Some evidences indicate that it is related to the excessive expression of some growth promotion factors. Angiotensin II is a polypeptide that may be involved in airway remodeling. To evaluate its role in airway remodeling in asthma, we observed the effects of an angiotensin II type 1 receptor antagonist (valsartan) on the expression of collagen III, collagen V, and transforming growth factor beta1 (TGF-beta1) mRNA and protein in the airway walls of sensitized rats.. Forty Wistar rats were randomly divided into 5 groups: control group, sensitized group, and valsartan groups 1, 2, and 3. The rats in the sensitized group and in valsartan groups 1, 2, and 3 were sensitized and challenged with ovalbumin. Rats in control group were sensitized and challenged with 0.9% NaCl. Rats from valsartan groups 1, 2, and 3 were drenched with valsartan (10 microg, 20 microg, or 30 microg, respectively) at the time of the ovalbumin challenges. The expression of collagen III, collagen V, and TGF-beta1 protein were detected using immunohistochemical method in combination with image analysis methods. The expression of TGF-beta1 mRNA was detected by in situ hybridization.. The expression in the airways of collagen III and collagen V was significantly higher in rats from the sensitized group (7.73 +/- 0.81, 1.34 +/- 0.28) and from valsartan groups 1, 2, and 3 (5.73 +/- 0.64, 1.13 +/- 0.15; 4.96 +/- 0.51, 0.98 +/- 0.08; 4.43 +/- 0.35, 0.93 +/- 0.06, respectively) than those in the control group (2.65 +/- 0.38, 0.67 +/- 0.08, P < 0.05). In addition, collagen levels were significantly lower in valsartan groups 1, 2, and 3 than those from the sensitized group (P < 0.05). The expression of TGF-beta1 mRNA and protein in the airways was significantly higher in rats from the sensitized group (20.49% +/- 3.46%, 29.73% +/- 3.25%) and from valsartan groups 1, 2, and 3 (16.47% +/- 1.94%, 19.41% +/- 1.87%; 14.38% +/- 1.58%, 18.29% +/- 1.43%; 12.96% +/- 1.73%, 18.63% +/- 1.11%, respectively) than that from the control group (7.84% +/- 1.61%, 5.63% +/- 1.07%, P < 0.05). TGF-beta1 mRNA and protein levels were significantly lower in valsartan groups 1, 2, and 3 than that in the sensitized group (P < 0.05).. Angiotensin II receptor antagonist valsartan can suppress synthesis of collagen III and collagen V by downregulating TGF-beta1 mRNA and protein expression. Valsartan can decrease airway remodeling and could play a role in asthma therapy.

Topics: Angiotensin Receptor Antagonists; Animals; Asthma; Bronchi; Collagen Type III; Collagen Type V; Immunization; Male; Ovalbumin; Random Allocation; Rats; Rats, Wistar; RNA, Messenger; Tetrazoles; Transforming Growth Factor beta; Valine; Valsartan

Immunostimulatory DNA-based vaccines can prevent the induction of CD4(+) type 2 T helper (Th2) cell-mediated airway inflammation in experimental models, when administered before or at the time of allergen exposure. Here we demonstrate their efficacy in limiting the progression of an established response to chronic pulmonary inflammation and airway remodelling on subsequent allergen challenge. Mice exhibiting Th2-mediated airway inflammation induced following sensitization and challenge with group 1 allergen derived from Dermatophagoides pteronyssinus group species (Der p 1), a major allergen of house dust mite, were treated with pDNA vaccines. Their airways were rechallenged and the extent of inflammation assessed. In plasma DNA (pDNA)-vaccinated mice, infiltration of inflammatory cells, goblet cell hyperplasia and mucus production were reduced and subepithelial fibrosis attenuated. The reduction in eosinophil numbers correlated with a fall in levels of the profibrotic mediator transforming growth factor (TGF)-beta1 in bronchoalveolar lavage (BAL) and lung tissue. In addition to lung epithelial cells and resident alveolar macrophages, infiltrating eosinophils, the principle inflammatory cells recruited following allergen exposure, were a major source of TGF-beta1. Protection, conferred irrespective of the specificity of the pDNA construct, did not correlate with a sustained increase in systemic interferon (IFN)-gamma production but in a reduction in levels of the Th2 pro-inflammatory cytokines. Notably, there was a reduction in levels of interleukin (IL)-5 and IL-13 produced by systemic Der p 1 reactive CD4(+) Th2 cells on in vitro stimulation as well as in IL-4 and IL-5 levels in BAL fluid. These data suggest that suppression of CD4(+) Th2-mediated inflammation and eosinophilia were sufficient to attenuate progression towards airway remodelling. Immunostimulatory DNA may therefore have a therapeutic application in treatment of established allergic asthma in patients.

Topics: Allergens; Animals; Antigens, Dermatophagoides; Arthropod Proteins; Asthma; Bronchoalveolar Lavage Fluid; Chronic Disease; Cysteine Endopeptidases; Disease Progression; Immunization; Immunoglobulin E; Immunoglobulin G; Interleukin-4; Interleukin-5; Mice; Mice, Inbred C57BL; Pulmonary Eosinophilia; Respiratory Hypersensitivity; Th2 Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1; Vaccines, DNA

Corticosteroids play an important role in inflammation and remodelling of airways and are considered an important therapeutic target in asthma. Inflammation in asthma is characterized by a dysregulation of eosinophil apoptosis and of markers of airways remodelling. We evaluated the ability of flunisolide to inhibit in vitro the release of metalloproteinases-9 (MMP-9), tissue inhibitor metalloproteinases-1 (TIMP-1), transforming growth factor (TGF-beta) and fibronectin by sputum cells (SC) as well as to induce sputum eosinophil apoptosis.. The SC, isolated from induced sputum samples of 12 mild-to-moderate asthmatics, were cultured for 24 h in the presence or absence of flunisolide (1, 10 and 100 microM). The release of mediators was assessed by enzyme-linked immunosorbent assay (ELISA) whereas apoptosis was studied by TUNEL technique.. Flunisolide (10 microM) significantly reduced MMP-9 and TIMP-1 (P = 0.0011 and P < 0.0001 respectively) and increased MMP-9/TIMP-1 molar ratio (P = 0.004). In addition, flunisolide decreased TGF-beta and fibronectin release by SC (P = 0.006; and P < 0.0001 respectively) and increased eosinophil apoptosis (P < 0.001).. These results demonstrate that flunisolide may play an important role in the inhibition of airway inflammation and remodelling, by promoting the resolution of eosinophilic inflammation and by inhibiting the release of MMP-9, TIMP-1, TGF-beta and fibronectin.

Topics: Adult; Apoptosis; Asthma; Fibronectins; Fluocinolone Acetonide; Humans; Matrix Metalloproteinase 9; Middle Aged; Sputum; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta; Transforming Growth Factor beta1

Features of chronic asthma include airway hyperresponsiveness, inflammatory infiltrates, and structural changes in the airways, termed remodeling. The contribution of eosinophils, cells associated with asthma and allergy, remains to be established. We show that in mice with a total ablation of the eosinophil lineage, increases in airway hyperresponsiveness and mucus secretion were similar to those observed in wild-type mice, but eosinophil-deficient mice were significantly protected from peribronchiolar collagen deposition and increases in airway smooth muscle. These data suggest that eosinophils contribute substantially to airway remodeling but are not obligatory for allergen-induced lung dysfunction, and support an important role for eosinophil-targeted therapies in chronic asthma.

Topics: Animals; Asthma; Bronchi; Cell Division; Collagen; Eosinophils; Interleukins; Leukocyte Count; Lung; Mice; Mice, Inbred BALB C; Mucus; Muscle, Smooth; Respiratory Function Tests; Respiratory Hypersensitivity; Th2 Cells; Transforming Growth Factor beta; Transforming Growth Factor beta1

To study the relation between Interleukin-4 (IL-4), IL-5, IL-13, transforming growth factor-beta(2) (TGF-beta(2)) and airway remodeling and to investigate the effects of Montelukast (MK) on airway inflammation and airway remodeling of asthma.. Twenty female BALB/c mice were randomly divided into a remodeling group and a treatment group (MK group), with 10 BALB/c mice in each group. The mice were sensitized by ovalbumin (OVA), and only the MK group was treated with MK (15 mg/kg). The number of total cells and eosinophils in bronchoalveolar lavage fluid (BALF) were counted. Light and electronic microscope were used to detect the pathologic histology and morphologic change. In situ hybridization and reverse transcription-polymerase chain reaction (RT-PCR) were used to measure IL-4, L-5, IL-13, and TGF-beta(2) mRNAs in the lung.. The numbers of total cells and eosinophils in BALF of the remodeling group were (5.4 +/- 1.1) x 10(5)/ml and 2.32 +/- 0.20, while those of the treatment group were (3.9 +/- 1.6) x 10(5)/ml and 1.64 +/- 0.32, respectively, the difference being significant (P < 0.01). Histological and electronic microscopic examination showed extensive airway inflammation, notably accumulation of significant numbers of eosinophils and lymphocytes in the remodeling group. Other features including prominent proliferation of airway epithelial cells protruded like fingers, increased thickness of smooth muscle, hyperplasia of connective tissue, goblet cell hyperplasia and a marked increase in airway mucus secretion with mucus plugging and extensive collagen deposition around the airways were also noted in the remodeling group. In the treatment group, the inflammation was significantly decreased, with decreased production of mucus, decreased collagen and granule of mucus around airway, less proliferation of airway epithelium, smooth muscle hypertrophy and airway spasm. In situ hybridization showed that the expression of IL-13 mRNA and TGF-beta(2) mRNA in the lung of the remodeling group were 24 +/- 7 and 17 +/- 5 respectively, while those of the treatment group were 17 +/- 4 and 10 +/- 3. RT-PCR results showed that the absorbance of IL-4 mRNA and IL-5 mRNA in the lung of the remodeling group were 0.91 and 0.96, while those of the treatment group were 0.22 and 0.35; the differences between the groups were all significant (all P < 0.01).. MK could effectively inhibit airway remodeling, which suggests a possible role of cysteinyl leukotrienes in the pathogenesis of chronic allergic inflammation with fibrosis.

Topics: Acetates; Animals; Asthma; Bronchi; Cyclopropanes; Disease Models, Animal; Eosinophils; Female; Interleukins; Leukotriene Antagonists; Mice; Mice, Inbred BALB C; Quinolines; Random Allocation; RNA, Messenger; Sulfides; Transforming Growth Factor beta; Transforming Growth Factor beta2

The transforming growth factor (TGF)-beta family is important for tissue repair in pathological conditions including asthma. However, little is known about the impact of either TGF-beta1 or TGF-beta2 on asthmatic airway epithelial mucin expression. We evaluated bronchial epithelial TGF-beta1 and TGF-beta2 expression and their effects on mucin expression, and the role of TGF-beta1 or TGF-beta2 in interleukin (IL)-13-induced mucin expression. Epithelial TGF-beta1, TGF-beta2, and mucin expression were evaluated in endobronchial biopsies from asthmatics and normal subjects. The effects of TGF-beta1 or TGF-beta2 on mucin MUC5AC protein and mRNA expression, and the impact of IL-13 on epithelial TGF-beta1, TGF-beta2, and MUC5AC were determined in cultured bronchial epithelial cells from endobronchial brushings of both subject groups. In biopsy tissue, epithelial TGF-beta2 expression levels were higher than TGF-beta1 in both asthmatics and normals. TGF-beta2, but not TGF-beta1, was increased in asthmatics compared with normals, and significantly correlated with mucin expression. TGF-beta2, but not TGF-beta1, increased mucin expression in cultured epithelial cells from both subject groups. IL-13 increased the release of TGF-beta2, but not TGF-beta1, from epithelial cells. A neutralizing TGF-beta2 antibody partially inhibited IL-13-induced mucin expression. These data suggest that TGF-beta2 production by asthmatic bronchial epithelial cells may increase airway mucin expression. IL-13-induced mucin expression may occur in part through TGF-beta2 up-regulation.

Topics: Adult; Asthma; Biopsy; Cells, Cultured; Female; Humans; Interleukin-13; Male; Middle Aged; Mucins; Respiratory Mucosa; Transforming Growth Factor beta

Asthma is associated with recruitment of eosinophils, accumulation of chronic inflammatory cells in the airway walls, subepithelial fibrosis and other structural changes of airway wall remodelling. The role of ongoing exposure to allergens in their pathogenesis remains unclear.. To examine whether changes of inflammation and remodelling were reversible following cessation of antigenic challenge in a mouse model of chronic asthma.. BALB/c mice sensitized to ovalbumin (OVA) were chronically challenged by inhalation of a low mass concentration of antigen for 8 weeks, leading to development of acute-on-chronic airway inflammation, subepithelial fibrosis and other changes of airway wall remodelling. Epithelial injury was assessed by immunohistochemistry, while inflammation and remodelling were quantified by appropriate histomorphometric techniques. Regression of lesions was assessed in animals examined at 1, 2 and 4 weeks after exposure to OVA ceased.. We did not find evidence of airway epithelial injury in this model of low-level chronic inhalational exposure to antigen. Persistence of the recruitment of eosinophils and chronic inflammatory cells in the airway walls was dependent on continuing antigenic challenge, as was persistence of mucous cell hyperplasia/metaplasia. Subepithelial fibrosis and epithelial hypertrophy exhibited delayed reversibility following cessation of exposure to antigen, possibly related to matrix-associated accumulation of transforming growth factor-beta(1).. In chronic asthma, low-level antigenic challenge may be required to maintain the inflammatory response in the airway wall, but airway remodelling may persist in its absence.

Topics: Administration, Inhalation; Allergens; Animals; Asthma; Chronic Disease; Disease Models, Animal; Eosinophils; Epithelial Cells; Female; Fibrosis; Hyperplasia; Mice; Mice, Inbred BALB C; Ovalbumin; Remission, Spontaneous; Respiratory Mucosa; Trachea; Transforming Growth Factor beta; Transforming Growth Factor beta1

Topics: Asthma; Humans; Inflammation; Matrix Metalloproteinase 9; Neutrophil Activation; Neutrophils; Transforming Growth Factor beta

T regulatory (T reg ) cells are characterized by expression of suppressive cytokines and the transcription factor FOXP3. They play a key role in balancing immune responses and maintain peripheral tolerance against antigens and allergens. The loss of peripheral tolerance against allergens causes diseases that can be therapeutically controlled with glucocorticoids.. The present study investigates whether glucocorticoids affect the activity of T reg cells on the basis of FOXP3 and cytokine expression.. CD4 + T cells from healthy donors and glucocorticoid-treated asthmatic patients were isolated, and expression of FOXP3, along with IL-10 and TGF-beta1, was determined. The effect of glucocorticoids on T reg cells was measured in vivo before and after GC treatment and in in vitro cultures.. FOXP3 mRNA expression was significantly increased in asthmatic patients receiving inhaled glucocorticoid treatment, systemic glucocorticoid treatment, or both. FOXP3 tightly correlated with IL10 mRNA expression. No correlation of FOXP3 mRNA expression was observed in relation to a (GT)n microsatellite promoter polymorphism on chromosome Xp11.23 or total IgE level. The frequency of CD25 + memory CD4 + T cells and transient FOXP3 mRNA expression by CD4 + T cells significantly increased after systemic glucocorticoid treatment, whereas TGFB1 expression did not change. Furthermore, glucocorticoids induced IL10 and FOXP3 expression in short-term and long-term cultures in vitro.. These findings demonstrate that glucocorticoid treatment is not only immunosuppressive and anti-inflammatory but also promotes or initiates differentiation toward T R 1 cells by a FOXP3-dependent mechanism. Strategies that convert transient glucocorticoid-induced T reg activity into a stable phenotype might improve allergy and asthma therapy.

Topics: Adult; Asthma; CD4-Positive T-Lymphocytes; Cell Differentiation; DNA-Binding Proteins; Forkhead Transcription Factors; Gene Expression Regulation; Glucocorticoids; Humans; Immunoglobulin E; Interleukin-10; Microsatellite Repeats; Middle Aged; Promoter Regions, Genetic; RNA, Messenger; T-Lymphocytes, Regulatory; Transforming Growth Factor beta; Transforming Growth Factor beta1; Up-Regulation

To observe the effect and mechanism of ligustrazine on the airway remodeling.. Thirty-two SD rats were randomly divided into 4 groups: the normal group (A), the model group (B), the ligustrazine low-dose group (C, 40 mg/kg) and the ligustrazine high-dose group (D, 80 mg/kg), with 8 rats in each group. The chronic asthmatic model was established by repeated inhalation of ovalbumin. The changes of collagen and transforming growth factor-beta(1) (TGF-beta(1)) contents in the airway wall, the thickness of smooth muscle and basement membrane, inner and outer diameter were measured by the computerized image analysis system.. The thickness of smooth muscle and basement membrane were (11.3 +/- 1.3, 11.3 +/- 1.7) microm in D group, (19.7 +/- 1.8, 19.8 +/- 1.6) microm in B group, the difference being significant (P < 0.01), as compared with A group [(10.6 +/- 1.2) microm, (9.8 +/- 1.6) microm] and C group [(11.6 +/- 0.9) microm, (12.3 +/- 1.8) microm], the difference being not significant (all P > 0.05). The difference in the ratio of inner diameter to outer diameter was significant between D group (0.77 +/- 0.06) and B group (0.63 +/- 0.05), P < 0.01. The contents of collagen type III and TGF-beta(1) were (21 +/- 5, 26 +/- 5) in D group, (55 +/- 7, 69 +/- 14) in B group, the difference being significant (P < 0.01). The differences were also significant when C group [32 +/- 8, 38 +/- 10] was compared with D group (P < 0.05) and B group (P < 0.01). The contents of collagen type I showed no difference among the 4 groups (A group: 34 +/- 13, B group: 44 +/- 8, C group: 36 +/- 8, D group: 39 +/- 8; all P > 0.05). A close correlation between TGF-beta(1) and collagen type III was demonstrated (r = 0.844 2, P < 0.01).. Ligustrazine might suppress airway remodeling by decreasing the expression of TGF-beta(1) and reducing deposition of collagen.

Topics: Animals; Anti-Asthmatic Agents; Asthma; Collagen Type III; Disease Models, Animal; Drugs, Chinese Herbal; Immunohistochemistry; Male; Muscle, Smooth, Vascular; Ovalbumin; Pyrazines; Rats; Rats, Sprague-Dawley; Transforming Growth Factor beta

Because the asthmatic bronchial epithelium is characterized by widespread damage, we postulated that this is associated with expression of cell cycle inhibitors that control proliferation. Using bronchial biopsies, the epithelium was the major site of expression of the cyclin-dependent kinase inhibitor, p21(waf). Immunostaining usually occurred in the cytoplasm of columnar cells; however, in severe asthma, nuclear staining was also evident in the proliferative, basal cell compartment. p21(waf) expression was significantly higher in asthmatic versus nonasthmatic epithelium and was unaffected by corticosteroid treatment; proliferating cell nuclear antigen was not significantly different in any group. p21(waf), but not p27(kip1), mRNA and protein were induced by treatment of bronchial epithelial cells in vitro with transforming growth factor (TGF)-beta or H2O2, but not by dexamethasone, which induced p57(kip2). TGF-beta and dexamethasone inhibited epidermal growth factor (EGF)-induced DNA synthesis, whereas low concentrations of H2O2 synergized with EGF; at higher doses, growth inhibition and induction of apoptosis occurred. TGF-beta caused p21(waf) to become nuclear, suggesting interaction with the replicative machinery; however, in oxidant-stressed cells, p21(waf) was predominantly cytoplasmic, where it has been linked to cell survival. We conclude that p21(waf) overexpression in asthma influences cell proliferation and survival. This may cause abnormal repair responses that contribute to airway inflammation and remodeling.

Topics: Asthma; Blotting, Western; Bronchi; Cells, Cultured; Cyclin-Dependent Kinase Inhibitor p21; Cyclins; Dexamethasone; DNA Replication; Epidermal Growth Factor; Epithelial Cells; Flow Cytometry; Humans; Hydrogen Peroxide; Immunohistochemistry; Proliferating Cell Nuclear Antigen; RNA, Messenger; Transforming Growth Factor beta

There is strong evidence to implicate transforming growth factor-beta in the remodeling that occurs in asthma, as levels are increased in bronchial lavage fluid and gene expression is increased in bronchial tissue. Transforming growth factor-beta is also known to increase the release of collagen from airway smooth muscle. Here we identify for the first time a possible mechanism for the effects of transforming growth factor-beta. Transforming growth factor-beta specifically induces mRNA and protein for connective tissue growth factor in airway smooth muscle, and moreover, we report that the connective tissue growth factor response is greater in airway smooth muscle cultured from patients with asthma compared with patients without asthma. This occurs at both the level of mRNA (37.53 +/- 11.62- and 13.59 +/- 3.12-fold increase at 24 hours compared with time 0, respectively, p < 0.02) and protein production (67.57 +/- 27.80- and 3.58 +/- 0.6-fold increase at 24 hours compared with time 0, respectively, p < 0.03). The differential connective tissue growth factor response to transforming growth factor-beta in asthmatic airway smooth muscle identifies a potential role for connective tissue growth factor in the remodeling that is characteristic of severe persistent asthma.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Asthma; Blotting, Western; Bronchi; Cells, Cultured; Connective Tissue Growth Factor; Female; Humans; Immediate-Early Proteins; Immunohistochemistry; Intercellular Signaling Peptides and Proteins; Male; Middle Aged; Muscle, Smooth; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Transforming Growth Factor beta; Transforming Growth Factor beta1

Transforming growth factor-beta (TGF-beta) and endothelin (ET) are found in elevated amounts in the airways of individuals with asthma. The cellular source of these peptides and their role in mediating the airway fibrosis of chronic asthma are unknown. In response to mechanical stresses similar to those occurring in vivo during airway constriction, bronchial epithelial cells increase the steady-state level of mRNA for both ET-1 and ET-2, followed by increased release of ET protein. Mechanical stress also enhances release of TGF-beta2 from a preformed cell-associated pool. TGF-beta2 and ET act individually and, more importantly, synergistically to promote fibrotic protein synthesis in reporter fibroblasts. To confirm the role of these intermediates in stress-induced fibrosis, conditioned medium from mechanically stressed bronchial epithelial cells was shown to elicit fibrotic protein synthesis in reporter fibroblasts; this effect was significantly inhibited by combined treatment with ET receptor antagonists and a neutralizing antibody to TGF-beta2. These data are consistent with a primary pathogenic role for mechanical stress-induced release of both TGF-beta2 and ET in the subepithelial fibrosis that characterizes chronic asthma.

Topics: Asthma; Bronchi; Cells, Cultured; Culture Media, Conditioned; Endothelin-1; Endothelin-2; Epithelial Cells; Fibrosis; Humans; Protein Biosynthesis; RNA, Messenger; Stress, Mechanical; Transforming Growth Factor beta; Transforming Growth Factor beta2

Eosinophil and mast cell infiltrations are consistent findings in nasal polyp tissue. Previous studies have shown that matrix metalloproteinases (MMPs) may be involved in eosinophil infiltration in airway mucosa of asthmatic patients, and that transforming growth factor-beta1 (TGF-beta1) induces extracellular matrix deposition in nasal polyp tissue. The aim of this study was to evaluate the role of MMPs and tissue-inhibitor of metalloproteinase-1 (TIMP-1) in association with TGF-beta1, eosinophils and mast cell activation in nasal polyp tissue. Nasal polyp tissues from 20 patients who underwent polypectomies were collected and prepared into tissue homogenate. Eosinophil cationic protein (ECP) and tryptase levels were measured by CAP system (Pharmacia, Sweden). MMP-2, MMP-9, TIMP-1 and TGF-beta1 levels were measured by enzyme-liked immunosorbent assay. MMP-2 was the predominant form of MMPs, followed by MMP-9 and TIMP-1. There were significant correlations between ECP, and MMP-9, MMP-2, TGF-beta1 and tryptase, but not with TIMP-1. Significant correlations were noted between tryptase, and MMP-2, MMP-9, and TGF-beta1, but not with TIMP-1. Close correlations were noted between TGF-beta1, and MMP-9 and MMP-2, but not with TIMP-1. MMP-2, MMP-9, and TGF-beta1 may contribute to eosinophil and mast cell migrations into nasal polyp tissue.

Topics: Adult; Asthma; Blood Proteins; Chemotaxis, Leukocyte; Eosinophil Granule Proteins; Eosinophilia; Eosinophils; Female; Humans; Male; Mast Cells; Matrix Metalloproteinase 2; Matrix Metalloproteinase 9; Middle Aged; Nasal Polyps; Rhinitis; Ribonucleases; Serine Endopeptidases; Tissue Inhibitor of Metalloproteinase-1; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tryptases

Mycoplasma pneumoniae infection exacerbates asthma in children and may play a role in the pathogenesis of chronic asthma. Because the airway epithelium is a preferential site for M. pneumoniae infection and a major source of the chemokine regulated on activation, normal T cells expressed and secreted (RANTES) and transforming growth factor (TGF)-beta1, we postulated that this microorganism may contribute to the disease by inducing these mediators through direct interaction with airway epithelial cells. We investigated the effects of M. pneumoniae on RANTES and TGF-beta1 production in primary cultures of normal human bronchial epithelial (NHBE) cells and small airway epithelial (SAEC) cells. Both cell types were permissive to M. pneumoniae infection in vitro, but their responses were different. TGF-beta1 was induced at higher levels in NHBE than in SAEC cultures, whereas RANTES was induced in SAEC cultures but not in NHBE cultures. These effects were attenuated by erythromycin and dexamethasone. In vitro adherence assays further indicated that the effects of erythromycin were mediated through its antimicrobial action, resulting in diminished adherence of the pathogen, whereas the effects of dexamethasone did not appear to be by inhibition of adherence. These results suggest that M. pneumoniae infection may contribute to the pathogenesis of chronic asthma at different levels of the airways, by inducing TGF-beta1 in large airways and the chemokine RANTES in small airways.

Topics: Asthma; Bronchi; Cell Adhesion; Cell Line; Cells, Cultured; Chemokine CCL5; Cytokines; Dexamethasone; DNA Primers; Dose-Response Relationship, Drug; Endotoxins; Epithelial Cells; Erythromycin; Glucocorticoids; Humans; Kinetics; Mycoplasma; Mycoplasma pneumoniae; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Time Factors; Transforming Growth Factor beta; Transforming Growth Factor beta1

Oral administration of allergens can induce immune tolerance to specific allergens in rodents and hence might be a possibility to prevent and treat allergic diseases in human subjects. However, the gastrointestinal tract of mice is different from that of human subjects. The absorption of specific antigens and subsequent antigen presentation to intestinal T cells is different in both species, making it difficult to extrapolate results.. We investigated primary oral tolerance to ovalbumin (OVA) in an IgE high-responder dog model, which is more predictive for human allergic diseases than corresponding rodent models.. Oral tolerance was induced by means of a 28-day treatment with OVA dissolved in cow's milk.. We observed reduced OVA-specific IgE and IgG production in response to ensuing subcutaneous challenges. Allergic conjunctivitis induced by means of ocular and airway provocation was significantly reduced in tolerized animals compared with that seen in nontolerized control animals. In addition, eosinophilia and neutrophilia in bronchoalveolar lavage fluid and bronchoconstriction after airway allergen challenge were significantly suppressed in tolerized animals. Cytokine analysis by means of real-time PCR on bronchoalveloar fluid cells after allergen challenge revealed a high-level expression of IL-10 and transforming growth factor beta, predominantly in the CD14(+) population.. Feeding infant beagles with OVA for 4 weeks is sufficient to prevent hallmark manifestations of asthma and allergy in adult life. The mechanism of oral tolerance involved an increased expression of IL-10 and transforming growth factor beta cytokines.

Topics: Administration, Oral; Animals; Asthma; Conjunctivitis; Dogs; Hypersensitivity; Immune Tolerance; Immunoglobulin E; Immunoglobulin G; Interleukin-10; Ovalbumin; RNA, Messenger; Transforming Growth Factor beta

Topics: Allergens; Animals; Asthma; Disease Models, Animal; Mast Cells; Mice; Pulmonary Fibrosis; Transforming Growth Factor beta

T cells are key regulators of immunologic disease parameters. However, their contribution to the process of tissue remodeling is ill defined. In the present study, we investigated gene expression of allergy-characteristic, IL-4-rich T cell cDNAs to monitor expression of genes that might participate in the pathogenesis of allergic diseases.. cDNAs of freshly isolated and restimulated CD4+ T cells from patients with allergic asthma (AA) or atopic dermatitis (AD) and healthy subjects were analyzed on Nylon membrane-based DNA arrays. Three patients were selected for an allergy-characteristic T cell phenotype with high IL-4 expression (AA) or IL-13 expression (AD).. Several gene families such as the TGF-beta family, chemokines and chemokine receptors were found to be upregulated. Matrix metalloproteinases and their inhibitors were also found to be expressed in an enhanced manner. Furthermore, factors regulating tissue turnover such as fibroblast growth factors and neurotrophic as well as vasoactive factors were found be expressed at a higher level in allergic patient compared to healthy donors.. The present study reveals and confirms genes relevant for allergy and highlights an approach to applying a DNA array technique for diagnostic discrimination of allergic diseases.

Topics: Asthma; CD4-Positive T-Lymphocytes; Cytokines; Dermatitis, Atopic; Endothelin-3; Gene Expression Regulation; Humans; Interleukin-13; Interleukin-4; Matrix Metalloproteinases; Oligonucleotide Array Sequence Analysis; Polymerase Chain Reaction; Receptors, Chemokine; RNA; Tissue Inhibitor of Metalloproteinases; Transforming Growth Factor beta

Asthmatic patients have higher numbers of mast cells in the smooth muscle layer of airways than normal subjects. Human airway smooth muscle cells (HASMCs) are a source of various cytokines including transforming growth factor beta1 (TGF-beta1), which is chemotactic for mast cells. We have thus examined the potential for interaction between HASMCs and mast cells and have investigated, in particular, the hypothesis that after stimulation, HASMCs can induce mast cell chemotaxis through the production of cytokines. Supernatants of HASMCs treated with the major mast cell product tryptase had increased chemotactic activity for the HMC-1 mast cell line. The effect depended on an intact catalytic site for tryptase and could be induced by a peptide agonist for protease activated receptor 2. Chemotactic activity was related to the synthesis of TGF-beta1 by HASMCs and, to a lesser extent, to stem cell factor. The number of mast cells within the smooth muscle layer of asthmatic patients was closely related to TGF-beta1 expression by smooth muscle. HASMCs may thus be able to stimulate the accumulation of mast cells, and these cells may, in turn, stimulate the secretion of chemotactic factors by HASMCs.

Topics: Asthma; Cell Communication; Cell Line; Cells, Cultured; Chemotaxis; Cytokines; Humans; Mast Cells; Models, Biological; Muscle, Smooth; Respiratory System; Serine Endopeptidases; Stem Cell Factor; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tryptases

Increased transforming growth factor beta1 (TGF-beta1) levels have been reported in bronchoalveolar lavage fluid and bronchial biopsy specimens from asthmatic patients. However, systemic TGF-beta1 levels have not been reported in asthma.. To evaluate the levels of plasma TGF-beta1 in asthmatic patients and matched, healthy controls to determine the associations with atopic status, disease severity, and duration.. Asthmatic patients and healthy controls were recruited prospectively from a university hospital outpatient department between January 2001 and May 2002. Plasma TGF-beta1 and serum IgE levels were estimated using established methods. Patients were classified as atopic or nonatopic based on the presence or absence of serum specific IgE directed to common allergens.. Of the 56 patients recruited for the study, 32 were atopic and 24 nonatopic. The median value of plasma TGF-beta1 was significantly higher in nonatopic asthmatic patients (2.5 ng/mL) compared with controls (1.5 ng/mL, P = .002) and atopic asthmatic patients (1.4 ng/mL, P = .008). The median absolute neutrophil count in the nonatopic asthmatic patients (4.0 x 10(9)/L) was significantly higher compared with atopic asthmatic patients (3.0 x 10(9)/L) and healthy controls (3.5 x 10(9)/L) (P = .01 and P = .04). There was no significant correlation between the duration or severity of asthma and plasma TGF-beta1 levels. The distribution of moderate-persistent asthma cases was similar in atopic and nonatopic groups.. Compared with atopic asthmatic patients and healthy controls, patients with nonatopic asthma have elevated plasma TGF-beta1 levels and leukocytosis. These data suggest that nonatopic asthmatic patients exhibit an altered inflammatory response, perhaps to a respiratory infection.

Topics: Administration, Inhalation; Adolescent; Adrenal Cortex Hormones; Adult; Androstadienes; Anti-Inflammatory Agents; Asthma; Biomarkers; Budesonide; Female; Fluticasone; Humans; Immunoglobulin E; Leukocyte Count; Male; Middle Aged; Neutrophils; Prospective Studies; Severity of Illness Index; Statistics as Topic; Transforming Growth Factor beta; Transforming Growth Factor beta1; Treatment Outcome

The role of transforming growth factor beta1 (TGF beta1) in airway remodeling in asthma and chronic obstructive pulmonary disease (COPD) has not been fully described. To evaluate the possible pathogenetic role of TGF beta1 in asthma and COPD, immunohistochemical expression of TGF beta1 was described in bronchial biopsies from patients with asthma and COPD compared with healthy individuals. Twelve subjects with asthma, 13 subjects with COPD, and 10 healthy individuals enrolled in the study. Bronchial biopsies were stained with hematoxylin and eosin and anti-TGF beta1 antibody. As a result, immunoreactive TGF beta1 was mainly localized in association with connective tissue in all groups. The staining intensity was not statistically different among the groups in bronchial epithelium, whereas it was significantly higher in the group of asthma in the submucosa. Because there is evidence showing a significant increase of staining intensity in the submucosa from asthmatics but not from subjects with COPD, we may conclude that TGF beta1 may play a significant role in pathogenesis of asthma but not in COPD.

Topics: Adult; Asthma; Biopsy; Bronchi; Humans; Middle Aged; Pulmonary Disease, Chronic Obstructive; Transforming Growth Factor beta; Transforming Growth Factor beta1

Transforming growth factor beta (TGF-beta) has been suggested to play an important role in the development of airway remodeling in asthma; this suggestion is based on evidence that expression levels of TGF-beta are correlated with unique parameters of airway remodeling, such as thickness of basement membrane. However, the relevant studies were inconclusive because they were unable to demonstrate active signaling mediated by the cytokine in the airways of asthmatic individuals.. We sought to determine whether TGF-beta signaling was active in the airways of asthmatic subjects and, if so, whether it was correlated with clinicopathologic features associated with the development of airway remodeling in asthma.. We examined the phosphorylation status of Smad2 in bronchial biopsy samples obtained from 40 asthmatic subjects as a marker of active TGF-beta signaling, and we studied its correlation with basement membrane thickness.. Expression levels of phosphorylated Smad2 in bronchial biopsy specimens from asthmatic subjects were higher than those in specimens from normal subjects, and they were correlated with basement membrane thickness in asthma.. The findings provide evidence that TGF-beta signaling was active in asthmatic airways and that the activity was associated with the development of airway remodeling in asthma.

Topics: Adult; Asthma; Bronchi; DNA-Binding Proteins; Humans; Phosphorylation; Signal Transduction; Smad2 Protein; Trans-Activators; Transforming Growth Factor beta; Transforming Growth Factor beta1

Chronic diseases may involve an "innate" response followed by an adaptive immune response, of a Th1 or Th2 variety. Little is known regarding the interactions of these responses. We hypothesized that TGF-beta1 (innate response factor associated with wound repair) in combination with IL-13 (Th2 factor) might augment inflammatory processes associated with asthma. Airway fibroblasts were cultured from asthmatic subjects and normal controls. These fibroblasts were exposed to TGF-beta1 and IL-13 alone or in combination, and eotaxin-1 expression and production were evaluated. At 48 h, eotaxin-1 production was markedly increased with the combination of TGF-beta1 and IL-13 (p < 0.0001) compared with either stimulus alone. mRNA increased slightly at 1 h with IL-13 or TGF-beta1 plus IL13, peaked, and became significantly increased over IL-13 alone at 24 h. Protein was measurable from 6 h with IL-13 and TGF-beta1 plus IL-13, but greater levels were measured over time with the combination. Actinomycin ablated the increase in mRNA and protein seen with IL-13 alone and with TGF-beta1 plus IL-13. Cycloheximide blocked the increase in mRNA at 6 h in both conditions, but also blocked the increase at 24 h with TGF-beta1 plus IL-13. STAT-6 was rapidly activated with both IL-13 and the combination, without difference. Finally, eotaxin-1-positive fibroblasts were identified in severe asthma biopsies in greater numbers than in normals. These results support the concept that interactions of innate and adaptive immune systems may be important in promoting the tissue eosinophilia of asthma, particularly in those with more severe disease.

Topics: Adjuvants, Immunologic; Asthma; Blotting, Northern; Bronchi; Cells, Cultured; Chemokine CCL11; Chemokines, CC; Chemotactic Factors, Eosinophil; Cycloheximide; Dactinomycin; Dose-Response Relationship, Immunologic; Drug Synergism; Fibroblasts; Humans; Interleukin-13; Interleukin-8; Lung; Polymerase Chain Reaction; RNA, Messenger; Signal Transduction; STAT6 Transcription Factor; Trans-Activators; Transforming Growth Factor beta; Transforming Growth Factor beta1; Up-Regulation

We have previously demonstrated that CpG oligodeoxynucleotides (CpG-ODNs) protect against eosinophilia and airway hyperresponsiveness in murine models of allergen-induced asthma. Acute inflammation is hypothesized to induce chronic airway responses, but no previous studies have evaluated the effects of CpG-ODNs on allergen-induced airway remodeling. Because remodeling is thought to be responsible for many of the long-term adverse effects on asthmatic patients, we evaluated whether CpG-ODNs might similarly prevent these changes using a murine model of recurrent allergen exposure.. The purpose of this study was to evaluate the effect of CpG-ODNs on chronic inflammatory changes and airway remodeling by using a murine model of chronic allergen-induced asthma.. C57BL/6 mice were sensitized to ovalbumin (OVA) and subsequently exposed to nebulized OVA by means of inhalation 3 times weekly for 6 weeks. Some mice received CpG-ODNs by means of intraperitoneal injection at the time of sensitization. At the end of the exposure period, mice were evaluated for the development of airway inflammation, airway hyperresponsiveness, and airway remodeling.. OVA-sensitized mice exposed to recurrent airway challenge with OVA have chronic inflammation, persistent airway hyperresponsiveness, and evidence of airway remodeling, including subepithelial collagen deposition and goblet cell hyperplasia-metaplasia. These changes are significantly reduced in mice treated with CpG-ODNs. Interestingly, mice treated with CpG-ODNs exhibit increased levels of bronchoalveolar lavage transforming growth factor beta, suggesting that regulatory T cells might be responsible for some of these protective effects.. CpG-ODNs are effective not only in preventing acute inflammation but also appear to reduce markers of airway remodeling that develop after chronic allergen exposure.

Topics: Animals; Asthma; Bronchi; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chronic Disease; Collagen; Disease Models, Animal; Eosinophilia; Female; Fibrosis; Immunoglobulin E; Mice; Mice, Inbred C57BL; Oligodeoxyribonucleotides; Transforming Growth Factor beta

Smad7 is an intracellular antagonist of transforming growth factor beta (TGF-beta) signaling, which could determine the intensity or duration of the TGF-beta signal. Because TGF-beta has been implicated in the development of airway remodeling in asthma on the basis of its strong capacity to induce extracellular matrix production, it is possible that Smad7 also plays some roles in the regulation of the process.. We sought to determine the relationships between Smad7 expression in bronchial biopsy samples from asthmatic subjects and clinicopathologic features.. Bronchial biopsy specimens were obtained from 40 asthmatic subjects and 6 healthy control subjects. Expression levels of Smad7 on a histologic section were estimated by immunohistochemical staining. In addition, the roles of Smad7 in TGF-beta-mediated transcriptional responses were studied by in vitro studies.. Smad7 immunoreactivity was detected mainly in bronchial epithelial cells in control and asthmatic subjects. Interestingly, asthmatic subjects exhibited less Smad7 immunoreactivity in bronchial epithelial cells than normal subjects. Expression levels of Smad7 in bronchial epithelial cells were inversely correlated with basement membrane thickness and airway hyperresponsiveness in asthmatic subjects. In addition, abrogation of endogenous Smad7 expression through use of an antisense oligonucleotide enhanced transcriptional responses to TGF-beta, whereas overexpression of Smad7 inhibited TGF-beta-induced plasminogen activator inhibitor 1production in a human bronchial epithelial cell line, BEAS2B cells.. These findings suggest that Smad7 is a key molecule that defines the susceptibility of bronchial epithelial cells to TGF-beta action and that regulation of Smad7 expression in bronchial epithelial cells might be related to the development of airway remodeling and airway hyperresponsiveness in asthma.

Topics: Adult; Asthma; Basement Membrane; Bronchi; Bronchial Hyperreactivity; DNA-Binding Proteins; Epithelial Cells; Female; Humans; Immunohistochemistry; Male; Middle Aged; Plasminogen Activator Inhibitor 1; Smad7 Protein; Trans-Activators; Transcription, Genetic; Transforming Growth Factor beta

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

Signal transducer and activator of transcription 6 (Stat6) is critical for Th2-mediated responses during allergic airway disease. To investigate the role of Stat6 in fungus-induced airway hyperresponsiveness and remodeling, Stat6-deficient (Stat6-/-) and Stat6-wildtype (Stat6+/+) mice were sensitized to Aspergillus fumigatus and airway disease was subsequently assessed in both groups at days 21, 30, 38, and 44 after an intratracheal challenge with live A. fumigatus conidia. At all times after conidia, histological analysis revealed an absence of goblet cell hyperplasia and markedly diminished peribronchial inflammation in Stat6-/- mice in contrast to Stat6+/+ mice. Airway hyperresponsiveness and peribronchial fibrosis in Stat6-/- mice were significantly reduced at day 21 after conidia compared with Stat6+/+ mice, but both groups exhibited significant, similar increases in these parameters at all subsequent times after conidia. In separate experiments, IL-13-responsive cells in Stat6-/- mice were targeted via the daily intranasal administration of 200 ng of IL-13-PE38QQR (IL13-PE), comprised of human IL-13 and a derivative of Pseudomonas exotoxin, from days 38 to 44 after the conidia challenge. IL13-PE treatment abolished airway hyperresponsiveness, but not peribronchial fibrosis in Stat6-/- mice. Taken together, these data demonstrate that the chronic development of airway hyperresponsiveness during fungal asthma is IL-13-dependent but Stat6-independent.

Topics: Animals; Aspergillosis, Allergic Bronchopulmonary; Aspergillus fumigatus; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Chemokine CCL11; Chemokine CCL2; Chemokine CCL5; Chemokines, CC; Chronic Disease; Collagen; Humans; Immunoglobulin E; Interleukin-13; Interleukin-4; Leukocytes; Lung; Mice; Mice, Inbred BALB C; Mice, Transgenic; Pulmonary Fibrosis; STAT6 Transcription Factor; T-Lymphocytes; Trans-Activators; Transforming Growth Factor beta

Expression microarrays are a powerful tool that could provide new information about the molecular pathways regulating common lung diseases. To exemplify how this tool can be useful, selected examples of informative experiments are reviewed. In studies relevant to asthma, the cytokine interleukin-13 has been shown to produce many of the phenotypic features of this disease, but the cellular targets in the airways and the molecular pathways activated are largely unknown. We have used microarrays to begin to dissect the different transcriptional responses of primary lung cells to this cytokine. In experiments designed to identify global transcriptional programs responsible for regulating lung inflammation and pulmonary fibrosis, we performed microarray experiments on lung tissue from wild-type mice and mice lacking a member of the integrin family know to be involved in activation of latent transforming growth factor (TGF)-beta. In addition to identifying distinct cluster of genes involved in each of these processes, these studies led to the identification of novel pathways by which TGF-beta can regulate acute lung injury and emphysema. Together, these examples demonstrate how careful application and thorough analysis of expression microarrays can facilitate the discovery of novel molecular targets for intervening in common lung diseases.

Topics: Animals; Antigens, Neoplasm; Asthma; Gene Expression; Genes, Regulator; Genetic Predisposition to Disease; Humans; In Vitro Techniques; Integrins; Interleukin-13; Lung; Matrix Metalloproteinase 12; Matrix Metalloproteinase 7; Metalloendopeptidases; Mice; Mice, Knockout; Oligonucleotide Array Sequence Analysis; Pulmonary Emphysema; Pulmonary Fibrosis; Respiratory Distress Syndrome; Transcriptional Activation; Transforming Growth Factor beta

Asthma is characterized by chronic inflammation of the airway wall with the presence of activated T helper 2 (Th2) lymphocytes. The current study assessed the ability of Th2 cytokines to modulate fibroblast-mediated contraction of collagen gels to determine if Th2 cytokines could contribute to tissue remodeling by altering mesenchymal cell contraction. Human fetal lung fibroblasts, human adult bronchial fibroblasts and human airway smooth muscle cells were cast into native type I collagen gels and allowed to contract in the presence or absence of IL (interleukin)-4, IL-5, IL-10, or IL-13. IL-4 and IL-13 but not IL-5 and IL-10 augmented collagen gel contraction in a concentration-dependent manner. Neither IL-4 nor IL-13 altered fibroblast production of transforming growth factor-beta or fibronectin. Both, however, decreased fibroblast prostaglandin (PG) E(2) release. Decreased PGE(2) release was associated with a decreased expression of cyclooxygenase 1 and 2 protein and mRNA. Indomethacin completely inhibited PGE(2) release and also augmented contraction. IL-4 and IL-13, however, added together with indomethacin further augmented contraction suggesting both a PGE-dependent and a PGE-independent effect. These findings suggest that IL-4 and IL-13 may modulate airway tissue remodeling and, therefore, could play a role in the altered airway connective tissue which characterizes asthma.

Topics: Animals; Asthma; Cells, Cultured; Collagen Type I; Cyclooxygenase 1; Cyclooxygenase 2; Dinoprostone; Fetus; Fibroblasts; Fibronectins; Fibrosis; Gels; Gene Expression Regulation, Enzymologic; Humans; Interleukins; Isoenzymes; Lung; Membrane Proteins; Mesoderm; Muscle, Smooth; Prostaglandin-Endoperoxide Synthases; Rats; RNA, Messenger; Th2 Cells; Transforming Growth Factor beta

Collagen type I is one of the major deposits in thickening of the reticular basement membrane of asthma.. In this study, we assessed turnover of collagen type I in asthma by measuring procollagen type I C-terminal peptide (PICP) and collagen type I C-terminal telopeptide (ICTP) in induced sputum.. PICP but not ICTP was found to be significantly higher in asthma subjects than in normal volunteers (P < 0.05). In asthma, PICP was inversely correlated with %FEV(1.0) (r = -0.539), and its levels significantly increased upon exacerbation (P < 0.05), indicating that collagen synthesis increases during asthma exacerbation. Additionally, PICP was found to significantly correlate with eosinophil counts in sputum (r = 0.539), indicating that eosinophils stimulate collagen turnover. Because eosinophils can produce TGF-beta, a potent stimulator of collagen synthesis, we immunocytochemically examined TGF-beta-positive cells in sputum. TGF-beta-positive cells significantly correlated with eosinophil counts (r = 0.811) and PICP (r = 0.569), suggesting that TGF-beta released from eosinophils is involved in collagen synthesis.. The results of the present study suggest that collagen synthesis is stimulated in asthmatic airways by eosinophils through TGF-beta, while collagen degradation is not, and that PICP in sputum can act as a new marker for airway inflammation in asthma.

Topics: Asthma; Biomarkers; Collagen Type I; Enzyme-Linked Immunosorbent Assay; Eosinophils; Female; Forced Expiratory Volume; Humans; Immunochemistry; Leukocyte Count; Male; Middle Aged; Peptide Fragments; Peptides; Procollagen; Serum Albumin; Sputum; Statistics as Topic; Transforming Growth Factor beta

To investigate the role of alveolar macrophages in airway remodeling in asthma.. Sixteen Wistar rats were divided into two groups: control group and asthmatic group. The expressions of platelet-derived growth factor-b (PDGF-b) and transforming growth factor-beta. (TGF-beta 1) in the alveolar macrophages of allergic rats were detected by immunohistochemistry and in situ hybridization.. The expression of TGF-beta 1 in alveolar macrophages in allergic group was increased in both protein level (P < 0.05) and TGF-beta 1 mRNA(P < 0.01); but the expression of PDGF-b had no difference between the two groups.. The alveolar macrophages may involve in the airway remodeling of asthma.

Topics: Animals; Asthma; Female; Macrophages, Alveolar; Male; Proto-Oncogene Proteins c-sis; Random Allocation; Rats; Rats, Wistar; RNA, Messenger; Transforming Growth Factor beta

Transforming growth factor beta1 (TGF-beta1) is produced by many types of cells that are activated in the asthmatic response. Recent studies have highlighted this cytokine as an important negative regulator in an experimental model of asthma. Although the role of TGF-beta1 in human asthma remains obscure, data derived from animal models have encouraged the further investigation of such suppression mechanisms in order to develop novel therapies for asthma.

Topics: Animals; Asthma; Disease Models, Animal; Humans; Mice; T-Lymphocytes; Transforming Growth Factor beta; Transforming Growth Factor beta1

In severe or chronic asthma, there is an increase in airway smooth muscle cell (ASMC) mass as well as an increase in connective tissue proteins in the smooth muscle layer of airways. Transforming growth factor-beta (TGF-beta) exists in three isoforms in mammals and is a potent regulator of connective tissue protein synthesis. Using immunohistochemistry, we had previously demonstrated that ASMCs contain large quantities of TGF-beta1-3. In this study, we demonstrate that bovine ASMC-derived TGF-beta associates with the TGF-beta latency binding protein-1 (LTBP-1) expressed by the same cells. The TGF-beta associated with LTBP-1 localizes TGF-beta extracellularly. Furthermore, plasmin, a serine protease, regulates the secretion of a biologically active form of TGF-beta by ASMCs as well as the release of extracellular TGF-beta. The biologically active TGF-beta released by plasmin induces ASMCs to synthesize collagen I in an autocrine manner. The autocrine induction of collagen expression by ASMCs may contribute to the irreversible fibrosis and remodeling seen in the airways of some asthmatics.

Topics: Animals; Aprotinin; Asthma; Autocrine Communication; Biological Assay; Carrier Proteins; Cattle; Cells, Cultured; Collagen; Extracellular Space; Fibrinolysin; Fibrosis; Intracellular Signaling Peptides and Proteins; Latent TGF-beta Binding Proteins; Mink; Muscle, Smooth; Procollagen; Serine Proteinase Inhibitors; Trachea; Transforming Growth Factor beta

Topics: Asthma; Humans; Phenotype; Transforming Growth Factor beta; Transforming Growth Factor beta1

Eosinophilic bronchitis is an essential component of bronchial asthma, and eosinophils play an important role. We studied the effect of eosinophils on cell surface plasmin generation by bronchial epithelial cells, because plasmin is thought to be involved in bronchial tissue repair/remodeling by means of fibrinolysis and the activation of proteases such as matrix metalloproteases. Plasmin was generated from exogenous plasminogen on the cell surface of cultured bronchial epithelial cells, NCI-H292. Transforming growth factor beta (TGF-beta) treatment resulted in reduced cell surface plasmin generation and a large increase in plasminogen activator inhibitor-type 1 (PAI-1) antigen production in NCI-H292 cells, whereas no conspicuous effects were observed with IL-1 beta and TNF alpha treatment (regulators in pulmonary epithelial cells). On the other hand, this cell surface plasmin generation was reduced by co-incubation with Eol-1, an eosinophil cell line. The addition of TGF-beta antisense and anti-TGF-beta antibodies attenuated this adverse effect of Eol-1 cell co-incubation. These data suggest that eosinophils play an inhibitory role on cell surface plasmin generation by bronchial epithelial cells by means of the up-regulation of PAI-1 expression induced by TGF-beta. Therefore, the accumulation of eosinophils in bronchial walls is thought to be involved in bronchial tissue repair/remodeling in asthma through this protease network.

Topics: Asthma; Cells, Cultured; Eosinophils; Epithelial Cells; Fibrinolysin; Humans; Oligonucleotides, Antisense; Plasminogen Activator Inhibitor 1; Transforming Growth Factor beta; Up-Regulation

Interleukin (IL)-4 and IL-13 are key proinflammatory cytokines in asthma. Studies in transgenic mice show that both cytokines cause inflammation, but only IL-13 causes subepithelial fibrosis, a characteristic feature of asthma. We compared the in vitro profibrogenic effects of IL-4 and IL-13 using bronchial fibroblasts from asthmatic subjects. In the presence of transforming growth factor (TGF)-beta the cells transformed into contractile myofibroblasts and expressed alpha-smooth muscle actin and procollagen I. IL-4 and IL-13 also stimulated proliferation, but were relatively ineffective in promoting myofibroblast transformation. TGF-beta was more potent than the cytokines in stimulating release of endothelin-1 and vascular endothelial growth factor, whereas IL-4 and IL-13 were more potent stimuli for eotaxin release. Although neither IL-4 nor IL-13 induced profibrotic responses, both cytokines caused a corticosteroid-insensitive stimulation of TGF-beta2 release from primary bronchial epithelial cells. These data indicate that epithelial activation by IL-13 or IL-4 plays a critical role in initiating remodeling through release of TGF-beta2. TGF-beta2 then activates the underlying myofibroblasts to secrete matrix proteins and smooth muscle and vascular mitogens to propagate remodeling changes into the submucosa. In contrast, direct activation of submucosal fibroblasts by IL-4 and IL-13 has a proinflammatory effect via eotaxin release and recruitment of eosinophils into the airways.

Topics: Actins; Adult; Asthma; Bronchi; Cell Division; Cells, Cultured; Chemokine CCL11; Chemokines, CC; Collagen Type I; Culture Media, Conditioned; Cytokines; Endothelial Growth Factors; Endothelin-1; Epithelial Cells; Fibroblasts; Gene Expression Regulation; Humans; Hypersensitivity, Immediate; Interleukin-13; Interleukin-4; Lymphokines; Protein Isoforms; Transforming Growth Factor beta; Vascular Endothelial Growth Factor A; Vascular Endothelial Growth Factors

Topics: Adult; Asthma; Female; Humans; Male; Polymorphism, Genetic; Transforming Growth Factor beta; White People

Transforming growth factor beta1 (TGFbeta1) is a multifunctional cytokine involved in pro- and anti-inflammatory pathways and is expressed in several cell types. Subepithelial fibrosis is one of the principle features of airway remodelling in asthma and is increased in severe patients. TGFbeta1 is implicated in fibrosis, including the deposition of extracellular matrix proteins. TGFbeta1 mRNA levels in eosinophils are increased in severe asthmatics relative to mild asthmatics. Therefore, TGFbeta1 is a promising candidate gene for contributing to asthma severity. Four polymorphisms located in the promoter region and signal peptide (C-509T, 72insC, T869C and G915C) were genotyped in groups of severe asthmatic, mild asthmatic or control individuals defined by steroid usage and pulmonary function. Significant differences ( P=0.016) were found between the groups for the genotype frequencies at C-509T, attributable mainly to a greater relative frequency of homozygosity for the -509T allele in the severe group compared to the mild and control groups. Individuals homozygous for -509T were also homozygous at the other variant sites for the 72C, 869C and 915G alleles (haplotype 1). The T allele creates a putative YY1 transcription factor binding site, but binding between YY1 and the DNA sequence of the T allele was not detected in vitro. In this study, we show that the -509T variant on haplotype 1 is the most informative marker of the TGFbeta1 contribution to asthma severity.

Topics: Adrenal Cortex Hormones; Adult; Alleles; Asthma; Female; Gene Frequency; Haplotypes; Humans; Male; Middle Aged; Point Mutation; Polymorphism, Genetic; Promoter Regions, Genetic; Protein Sorting Signals; Respiratory Function Tests; Severity of Illness Index; Transforming Growth Factor beta; Transforming Growth Factor beta1

We examined the effect of airway inflammation on airway remodeling and bronchial responsiveness in a mouse model of allergic asthma.. BALB/c mice were sensitized to ovalbumin (OA), and exposed to aerosolized OA (0.01, 0.1 and 1%). Twenty-four hours after the final antigen challenge, bronchial responsiveness was measured, and bronchoalveolar lavage (BAL) and histological examinations were carried out.. Repeated antigen exposure induced airway inflammation, IgE/IgG1 responses, epithelial changes, collagen deposition in the lungs, subepithelial fibrosis associated with increases in the amount of transforming growth factor (TGF)-beta1 in BAL fluid (BALF), and bronchial hyperresponsiveness to acetylcholine. The number of eosinophils in BALF was significantly correlated with TGF-beta1 production in BALF and the amount of hydroxyproline. Furthermore, significant correlations were found between these fibrogenic parameters and the bronchial responsiveness.. These findings demonstrated that in this murine model airway eosinophilic inflammation is responsible for the development of airway remodeling as well as bronchial hyperresponsiveness in allergic bronchial asthma.

Topics: Acetylcholine; Allergens; Animals; Asthma; Bronchial Hyperreactivity; Bronchoalveolar Lavage Fluid; Collagen; Cytokines; Epithelial Cells; Female; Hydroxyproline; Hypersensitivity; Immunoglobulin E; Immunoglobulin G; Immunoglobulins; Lung; Mice; Mice, Inbred BALB C; Respiratory System; Transforming Growth Factor beta

In asthma, treatment with inhaled corticosteroids reduces chronic peribronchial inflammation and restores the balance within macrophage subpopulations. This study investigates whether corticosteroids can regulate monocyte differentiation in vitro and thereby influence the balance of functionally distinct macrophages. Graded doses of fluticasone propionate (FP) were added to cultures of normal peripheral blood monocytes in the presence or absence of IL-4. Cells were harvested after 7 days' culture. Double immunofluorescence studies were performed on cytospins of differentiated macrophages using the MoAbs RFD1 and RFD7 to distinguish inductive and suppressive macrophages by their respective phenotypes. Macrophage function was determined by quantifying allostimulation in a mixed leucocyte reaction and by measuring tumour necrosis factor-alpha (TNF-alpha) production. FP reduced the number of mature cells with a D1+ antigen-presenting phenotype and up-regulated the development of cells with the D1/D7+ and D7+ phenotypes. Functionally, this was associated with reduced stimulation of T cell proliferation in a mixed leucocyte reaction (MLR). Fluticasone also reversed the increase in both D1+ expression and TNF-alpha production induced by IL-4. The effect of FP persisted for 24 h after removal of FP from the culture medium. These results suggest that FP treatment of asthmatics may have a direct beneficial effect by normalizing the macrophage subset imbalance that contributes to the chronic peribronchial inflammation present in this condition.

Topics: Adrenal Cortex Hormones; Androstadienes; Anti-Asthmatic Agents; Asthma; Cell Differentiation; Fluticasone; Humans; In Vitro Techniques; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; Macrophages; Phenotype; T-Lymphocytes; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

Mast cells are versatile effector cells of primary importance in asthma and airway inflammation. During inflammation mast cells accumulate in the bronchial epithelium. The mechanism for this increase in mast cell number has not been defined.. The aim of this study was to examine the presence of mast cell chemotactic activity in bronchoalveolar lavage (BAL) fluid taken before and at the end of 2 pollen seasons from patients with allergic asthma.. To measure mast cell chemotactic activity, we used a modified Boyden chamber and the human mast cell line HMC-1 or in vitro-developed mast cells as responder cells.. A total of 27 patients were investigated, of which 8 exhibited mast cell chemotactic activity in their BAL fluid collected before season. A significant increase in the activity was found in 18 of 27 BAL fluids sampled at the end of the pollen season. No difference was found between patients treated with immunotherapy or placebo. The presence of stem cell factor could be detected in all BAL fluids analyzed. Blocking antibodies against stem cell factor or transforming growth factor-beta partially blocked the activity in some of the BAL fluids. Treatment of the responder cells with pertussis toxin reduced the migratory activity in 13 of 14 BAL fluids collected during pollen season.. This study demonstrates the presence of mast cell chemotactic activity in BAL fluids from patients with allergic asthma, with a significant increase in activity during pollen season. The major part of this activity consisted of factors mediating their effect through G(i)-protein coupled receptors. This activity may be responsible for the mast cell accumulation in the intraepithelial layer seen in allergic asthmatic patients.

Topics: Asthma; Bronchoalveolar Lavage Fluid; Cell Line; Cell Movement; Chemotaxis; Enzyme Inhibitors; Genistein; Humans; Indoles; Maleimides; Mast Cells; Pollen; Protein Kinase Inhibitors; Protein-Tyrosine Kinases; Seasons; Stem Cell Factor; Transforming Growth Factor beta

Epithelial damage and airway remodeling are consistent features of bronchial asthma and are correlated with disease chronicity, severity, and bronchial hyperreactivity. To examine the mechanisms that control bronchial epithelial repair, we investigated expression of the epidermal growth factor receptor (c-erbB1, EGFR) in asthmatic bronchial mucosa and studied repair responses in vitro. In biopsies from asthmatic subjects, areas of epithelial damage were frequently observed and exhibited strong EGFR immunostaining. EGFR expression was also high in morphologically intact asthmatic epithelium. Using image analysis, EGFR immunoreactivity (% of total epithelial area, median (range) was found to increase from 9.4 (4.1-20.4) in normal subjects (n=10) to 18.4 (9.3-28.9) in mild asthmatics (P<0.01, n=13) and 25.4 (15.4-31.8) in severe asthmatics (P<0.00, n=5). Epithelial EGFR immunoreactivity remained elevated in patients treated with corticosteroids and was positively correlated with subepithelial reticular membrane thickening. Using 16HBE 14o- bronchial epithelial cells, we found that EGF accelerated repair of scrape-wounded monolayers and that the EGFR-selective inhibitor, tyrphostin AG1478, inhibited both EGF-stimulated and basal wound closure whereas dexamethasone was without effect. Intrinsic activation of the EGFR was confirmed by analysis of tyrosine phosphorylated proteins, which revealed a rapid, damage-induced phosphorylation of the EGFR, irrespective of the presence of exogenous EGF. To assess the relationship between EGFR-mediated repair and tissue remodeling, release of the profibrogenic mediator TGF-beta2 was also measured. Scrape wounding increased release of TGF-beta2 from epithelial monolayers and EGF had no additional stimulatory effect. However, when repair was retarded with AG1478, the amount of TGF-beta2 increased significantly. These data indicate that the EGFR may play an important role in bronchial epithelial repair in asthma and that impairment of this function may augment airway remodeling.

Topics: Adult; Asthma; Bronchi; Case-Control Studies; Cell Line; Epithelium; ErbB Receptors; Female; Humans; Immunohistochemistry; Male; Middle Aged; Models, Biological; Mucous Membrane; Phosphorylation; Transforming Growth Factor beta; Wound Healing

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

To investigate whether extracellular matrix glycosaminoglycan hyaluronan (HA) modulates eosinophil activation and transforming growth factor (TGF)-beta production by eosinophils, human peripheral blood eosinophils (purity > 99%) from 12 patients with mild to moderate asthma or six healthy subjects were isolated and incubated with increasing concentrations of low molecular weight (mol wt) HA ( approximately 0.2 x 10(6) D) or high mol wt HA (3.0 to approximately 5.8 x 10(6) D). We found that the low mol wt HA has a pronounced effect on eosinophil survival in both patients with asthma and healthy subjects in a dose-dependent fashion on Days 2 and 4. Whereas the high mol wt HA had a smaller effect on eosinophil survival than did the low mol wt HA. The HA-mediated eosinophil survival was partially but significantly inhibited ( approximately 50% inhibition) by a blocking monoclonal antibody for CD44, a specific receptor of HA, and largely inhibited by an anti-granulocyte macrophage colony-stimulating factor (GM-CSF) neutralizing antibody but not by an anti-interleukin (IL)-3 or anti-IL-5 neutralizing antibody. In addition, the low mol wt HA increased GM-CSF messenger RNA (mRNA) expression and protein secretion by eosinophils in a dose-dependent fashion, suggesting that the HA-mediated eosinophil survival is due mainly to induction of GM-CSF release through partial CD44 signaling. Furthermore, we demonstrated that the low mol wt HA results in morphologic changes in eosinophils such as transforming from a round to a spindle shape and in homotypic aggregation, upregulates intercellular adhesion molecule-1 expression, and increases TGF-beta mRNA expression and protein secretion by eosinophils. These observations suggest previously unforeseen interactions between eosinophils and low mol wt extracellular matrix and, thus, novel pathways by which eosinophils may contribute to the regulation of airway inflammation and airway remodeling.

Topics: Asthma; Base Sequence; Cell Survival; DNA Primers; Eosinophils; Histocompatibility Antigens Class II; Humans; Hyaluronic Acid; Intercellular Adhesion Molecule-1; Molecular Weight; Transforming Growth Factor beta

Adhesion molecules are involved in inflammatory and repair processes of the bronchial epithelium. ICAM-1 is mainly involved in inflammatory reactions, whereas integrins, such as alpha3beta1, are mainly involved in repair processes.. Using bronchial biopsies from 10 asthmatics and eight controls, we first evaluated by immunohistochemistry expression of alpha3beta1 and ICAM-1 in intact and damaged epithelium. Then, using the human pulmonary epithelial cell line WI-26 VA, we studied, by flow-cytometry, the modulation of ICAM-1 and alpha3beta1 expression, and, by ELISA, the release of fibronectin by proinflammatory cytokines, such as IL-5, and anti-inflammatory cytokines, such as IL-4, TGF-beta, and EGF.. alpha3beta1 expression was slightly higher in asthma than in controls, as well as in damaged epithelium than in undamaged epithelium. ICAM-1 expression was higher in asthma than in controls, and similarly distributed in intact or damaged epithelium. In vitro, alpha3beta1 was significantly increased by TGF-beta, EGF, and IL-4, and significantly decreased by IL-5. Fibronectin release was significantly increased by TGF-beta and IL-4, unchanged by EGF, and slightly but significantly decreased by IL-5. ICAM-1 expression was significantly decreased by TGF-beta and IL-4, unchanged by EGF, and significantly increased by IL-5.. These differences in adhesion molecule expression and fibronectin release may be important in epithelial cell inflammation and repair.

Topics: Adolescent; Adult; Aged; Asthma; Biopsy; Bronchi; Cell Line; Cytokines; Enzyme-Linked Immunosorbent Assay; Epidermal Growth Factor; Epithelial Cells; Fibronectins; Flow Cytometry; Humans; Inflammation; Integrin alpha3beta1; Integrins; Intercellular Adhesion Molecule-1; Interleukin-4; Middle Aged; Respiratory Mucosa; Transforming Growth Factor beta

Airway remodeling may play an important role in asthma pathophysiology. Transforming growth factor beta (TGF-beta) has a critical role in the remodeling process. Although cellular sources for TGF-beta have been previously investigated in asthma airways, the expression, release, or both of TGF-beta from asthmatic airways and blood neutrophils has not been reported.. The current study evaluated the TGF-beta protein and messenger (m)RNA expression by airway and peripheral blood neutrophils in asthmatic and normal subjects.. TGF-beta protein expression by airway and peripheral blood neutrophils was detected by using immunocytochemistry. TGF-beta protein levels in blood neutrophil supernatant were measured by using an enzyme immunoassay. TGF-beta mRNA expression was evaluated by using reverse transcription-PCR.. Higher numbers of TGF-beta(+) cells and neutrophils were found in airway tissue of asthmatic (n = 15) compared with normal subjects (n = 10). Although neutrophils in both asthmatic and normal airway tissue expressed TGF-beta protein and the percentage of neutrophils expressing TGF-beta was similar between the two groups, the total number of TGF-beta(+) neutrophils was higher in the asthmatic subjects (P =.01). Peripheral blood neutrophils from asthmatic (n = 5) and normal subjects (n = 7) also expressed TGF-beta protein and mRNA. Blood neutrophils from asthmatic subjects spontaneously released significantly higher levels of TGF-beta than those from normal subjects (P =.007).. These data suggest that airway and blood neutrophils from both asthmatic and normal subjects can express and release TGF-beta. Higher levels of TGF-beta expression-release from asthmatic neutrophils indicate that neutrophils may be involved in the airway remodeling process of asthmatic subjects.

Topics: Adult; Asthma; Biopsy; Bronchi; Female; Humans; Male; Microscopy, Confocal; Neutrophils; Pancreatic Elastase; Respiratory System; RNA, Messenger; Transforming Growth Factor beta

Topics: Animals; Asthma; Dexamethasone; Disease Models, Animal; Platelet-Derived Growth Factor; Rats; Rats, Wistar; Transforming Growth Factor beta

Bronchial subepithelial fibrosis is an histological characteristic of asthma. Cytokines and other mediators, such as PDGF-BB, TGF-beta1 and ET-1 found in the asthmatic submucosa can potentially activate a repair process that leads to fibroblast proliferation and collagen synthesis. The mechanisms of modulation of the repair process leading to extracellular matrix deposition are still to be documented. In this study, we assessed the in vitro proliferation and collagen synthesis of bronchial fibroblasts isolated from normal and asthmatic subjects in response to ET-1, platelet-derived growth factor (PDGF)-BB and transforming growth factor (TGF)-beta1 alone or in combination, in the presence or absence of dexamethasone. The combination of ET-1 with one of the other two growth factors, or the triple combination, significantly increased DNA synthesis and collagen production of bronchial fibroblasts isolated from both normal and asthmatic subjects, but the same growth factors used separately had no significant effect on the same parameters. These results suggest that the simultaneous presence of ET-1, PDGF-BB and TGF-beta1 in both normal and asthmatic subjects is necessary to activate bronchial fibroblast proliferation and collagen synthesis. As these mediators are present in the submucosa of the asthmatic bronchi, they could be responsible, at least in part, for the accumulation of collagen in the mucosa.

Topics: Analysis of Variance; Asthma; Becaplermin; Bronchi; Case-Control Studies; Cell Division; Cells, Cultured; Dexamethasone; DNA; Endothelin-1; Fibroblasts; Glucocorticoids; Humans; Platelet-Derived Growth Factor; Procollagen; Proto-Oncogene Proteins c-sis; Transforming Growth Factor beta

Asthma is characterized histologically by a bronchial subepithelial fibrosis. Cytokines and other mediators released in the asthmatic chronic inflammatory microenvironment can activate the repair process that leads to fibroblast proliferation and collagen synthesis. To our knowledge, there are no data regarding the effect of a chronic inflammatory microenvironment on the phenotype of human bronchial fibroblasts. In the present study, we address this issue by comparing bronchial fibroblasts isolated from normal and asthmatic subjects in terms of: (a) proliferation over cell passage; (b) in vitro lifespan; (c) proliferative response to transforming growth factor-beta 1, platelet-derived growth factor-BB, dexamethasone, and retinoic acid; and (d) base-line synthesis of procollagens I and III. Bronchial fibroblasts from asthmatic subjects demonstrated lower DNA synthesis with cell passage than bronchial fibroblasts from normals. The in vitro lifespan of asthmatic bronchial fibroblasts was lower than in those from normal subjects and was significantly correlated with airway responsiveness. Platelet-derived growth factor-BB and dexamethasone increased 3H-thymidine incorporation in asthmatic bronchial fibroblasts without having any significant effect on normal fibroblast proliferation. Transforming growth factor-beta 1 and retinoic acid had no significant effect on bronchial fibroblast proliferation. Base-line procollagens I and III synthesis measurements showed no differences between normal and asthmatic fibroblasts. Taken together, these results indicate that the chronic inflammatory microenvironment found in asthma can modulate some aspects of bronchial fibroblast phenotype.

Topics: Adult; Asthma; Basement Membrane; Becaplermin; Bronchi; Cell Division; Cells, Cultured; Dexamethasone; Fibroblasts; Glucocorticoids; Humans; Middle Aged; Phenotype; Platelet-Derived Growth Factor; Procollagen; Proto-Oncogene Proteins c-sis; Reference Values; Transforming Growth Factor beta; Tretinoin

Bronchial asthma is characterised by airway structural changes, including mucosal inflammatory infiltration and subepithelial collagen deposition, that may represent the morphological basis for the chronicity of the disease. The relationship between airway wall thickness and growth factors in asthma has not been elucidated.. Bronchial biopsy specimens were obtained from 21 asthmatic patients and eight healthy subjects and the basement membrane thickness was measured by light microscopy and electron microscopy. At the same time the numbers of eosinophils and fibroblasts were assessed and the expression of transforming growth factor beta 1 (TGF-beta 1), platelet derived growth factor (PDGF), and insulin like growth factor (IGF) I in the bronchial mucosa was examined by immunostaining. The relationship between the degree of thickening of the subepithelial layer and both the clinical data and pulmonary function were also investigated.. The basement membrane of the asthmatic patients was thicker than that of the healthy controls (median 8.09 versus 4.02 microns). Electron microscopic examination of the basement membrane revealed thickening of the subepithelial lamina reticularis; this thickening significantly correlated with the number of fibroblasts in the submucosa in the asthmatic subjects (rs = 0.88) but not in the controls (rs = 0.70). There was a significantly higher number of eosinophils in the airways of the asthmatic subjects than in the healthy subjects (EG1 + cells: 52.0 versus 2.0/mm2, EG2 + cells: 56.0 versus 1.5/mm2). The expression of each growth factor in the bronchial mucosa was similar in asthmatic and healthy subjects (TGF-beta 1: 18.0% versus 16.0%, PDGF: 37.0% versus 32.5%, IGF-I: 15.0% versus 8.0%). A weak but statistically significant correlation was found between the number of fibroblasts and the expression of TGF-beta 1 in asthmatic subjects (rs = 0.50). There was a significant correlation between the thickness of the subepithelial layer in asthmatic subjects and the attack score (rs = 0.58) and a significant inverse correlation between the subepithelial collagen thickness in asthmatic subjects and airway hypersensitivity (rs = -0.65).. These findings indicate that the thickening of the subepithelial layer in bronchial asthma is due to an increase in fibroblasts, and that the thickness of the subepithelial collagen appears to be linked to an increase in bronchial responsiveness and exacerbation of clinical manifestations.

Topics: Adolescent; Adult; Asthma; Basement Membrane; Bronchi; Cell Count; Eosinophils; Female; Fiber Optic Technology; Fibroblasts; Growth Substances; Humans; Insulin-Like Growth Factor I; Male; Microscopy, Electron; Middle Aged; Mucous Membrane; Platelet-Derived Growth Factor; Transforming Growth Factor beta

The cysteinyl leukotrienes (CysLTs) mimic many of the features of asthma and are implicated in its pathophysiology. Little, however, is known about the effects of the CysLTs on airways remodeling. In this study the effects of leukotriene D4 (LTD4) on human airway smooth muscle (HASM) cell proliferation and expression of extracellular matrix proteins were investigated. LTD4 (0.1-10 microM) alone had no effect on DNA synthesis in HASM. LTD4, however, markedly augmented proliferation induced by the mitogen, epidermal growth factor (EGF, 1 ng/ml). The potentiating effect of LTD4 (1 microM) on EGF-induced DNA synthesis was abolished by pranlukast (1 microM) or pobilukast (30 microM), but unaffected by zafirlukast (1 microM). In contrast, pranlukast (pKB = 6.9), pobilukast (pKB = 7.0), and zafirlukast (pKB = 6.5) had equivalent potencies for inhibition of LTD4-induced contraction in human bronchus. LTD4 (0.1 or 10 microM) did not increase the total messenger RNA expression of the extracellular matrix proteins (pro-alpha[I] type I or alpha1[IV] type IV collagen), elastin, biglycan, decorin, and fibronectin, and did not influence tumor growth factor-beta (10 ng/ml)-induced effects on the expression of these proteins in HASM cells. These data indicate that LTD4 augments growth factor-induced HASM proliferation but does not alter the expression of various extracellular matrix components. The observed differences in sensitivity to the antagonists suggests that the former phenomenon may be mediated by a CysLT receptor distinct from that which mediates LTD4-induced HASM contraction. Collectively, these results provide preliminary evidence that CysLTs may play a role in airways remodeling in asthma.

Topics: Asthma; Carbachol; Cells, Cultured; Chromones; Dicarboxylic Acids; DNA Replication; Epidermal Growth Factor; Extracellular Matrix Proteins; Gene Expression Regulation; Humans; Indoles; Leukotriene Antagonists; Leukotriene D4; Membrane Proteins; Muscle Contraction; Muscle, Smooth, Vascular; Phenylcarbamates; Receptors, Leukotriene; RNA, Messenger; Sulfonamides; Tosyl Compounds; Trachea; Transforming Growth Factor beta

IL-10 can modulate the differentiation of normal monocytes to macrophages, increasing the proportion of maturing cells with a phenotype consistent with T cell suppressive activity. Analysis of the immunopathology in endobronchial biopsies from asthmatic subjects has revealed significantly reduced proportions of suppressive macrophage populations associated with chronic T-cell mediated inflammation.. This study investigates whether the altered homeostasis within the lung macrophage populations in asthma is reflected in aberrant differentiation of peripheral blood monocytes and whether this differentiation may be influenced by IL-10.. Monocytes from 14 normal individuals and 14 atopic asthmatics were grown in culture for 7 days in the presence or absence of IL-10, added on day 5. Double immunofluoresence studies were performed on cytospins of the differentiated macrophages using the monoclonal antibodies RFD1 and RFD7 to distinguish inductive and suppressive macrophages by their respective phenotypes. HLADR expression was quantified using the monoclonal antibody RFDR1. Macrophage function was determined by quantifying allostimulation in a mixed leucocyte reaction and by measuring TNFalpha and TGFbeta production.. With no cytokine addition the proportion of maturing macrophages with a suppressive phenotype (D1+D7+) at day 7 was lower in the asthmatic samples (18%) compared with normals (25%). IL-10 increased the proportion of suppressive cells in cultures of both asthmatic and normal monocytes with the increase in the asthmatic subjects (94% increase) being significantly greater than that in normal subjects (32% increase) (P<0.01). Asthmatic monocytes had a greater effect in stimulating MLR than normals (P < 0.05) but the addition of IL-10 reduced T cell proliferation in an MLR to a equivalent level in both groups.. These results suggest that a fundamental problem may exist in the differentiation of monocytes in asthma which may be reversed by IL-10.

Topics: Adolescent; Adult; Asthma; Cell Differentiation; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; HLA-DR Antigens; Homeostasis; Humans; Immunophenotyping; Interleukin-10; Lymphocyte Culture Test, Mixed; Macrophages, Alveolar; Middle Aged; Monocytes; Transforming Growth Factor beta; Tumor Necrosis Factor-alpha

1. Airway remodelling occurs in asthma and involves an increase in airway smooth muscle mass through cell proliferation and hypertrophy. Increased eosinophil density in the airways is a feature of asthma. Eosinophils exhibiting activation in the airways of asthmatics also exhibit increased expression of transforming growth factor beta (TGF-beta1). We have examined the capacity of TGF-beta1 and epidermal growth factor (EGF) to influence airway smooth muscle division and the effect of heparin on TGF-beta1. EGF and serum-induced smooth muscle DNA synthesis in confluent airway smooth muscle cells (ASMC) as an indication of entry into S phase preceding mitogenesis. 2. ASMC were obtained from cell populations growing out from explanted bovine trachealis muscle sections. Cell division was monitored in sparse plated cells by direct cell counting following nuclear staining. Cell DNA synthesis in confluent cells was monitored by uptake of [3H]-thymidine. 3. TGF-beta1 (100 microM) inhibited FBS (10%)-induced smooth muscle division in sparsely plated cells (40%). TGF-beta1 (100 pM) increased cell DNA synthesis (200%) in confluent cells in the presence of bovine serum albumin (BSA, 0.25%). EGF (0.7 nM) also increased airway smooth muscle DNA synthesis (69%) in the presence of BSA (0.25%). The facilitatory effect of TGF-beta1 was observed between 1-100 pM, while that of EGF was observed between 20 200 pM. 4. Heparin inhibited serum and TGF-beta1-induced DNA synthesis in confluent ASMC (55%), consistent with our previous observation of inhibition of division in sparsely populated ASMC (Kilfeather et al., 1995a). This action of heparin was observed between concentrations of 1-100 microg ml(-1). Heparin did not inhibit DNA synthesis in response to EGF. An anti-mitogenic effect of heparin was also observed against responses to combined exposure to TGF-beta1 and EGF. 5. There was a clear inhibitory effect of heparin in absolute terms against serum-induced division in cells plated at 10, 20 and 45 x 10(3) cells cm(-2). The inhibitory effect of heparin was also observed at a plating density of 45,000 cells cm(-2) when responses to serum were expressed as fold-stimulation of basal DNA synthesis. 6. These findings demonstrate a potential role of TGF-beta1, EGF and heparin-related molecules in regulation of airway smooth muscle division.

Topics: Animals; Asthma; Blood Physiological Phenomena; Cattle; Cell Division; Cells, Cultured; Dose-Response Relationship, Drug; Epidermal Growth Factor; Heparin; Muscle, Smooth; Nucleic Acid Synthesis Inhibitors; Trachea; Transforming Growth Factor beta

The mechanisms responsible for persistence of T lymphocytes at the sites of allergic inflammation are not completely understood. Activated T cells, usually expressing Fas on their surface, undergo activation-induced apoptotic death, thus limiting the dangerous consequences of a persistent immune reaction. We have previously shown that pulmonary T lymphocytes from untreated asthmatic subjects do not express surface Fas receptors nor do they contain Fas mRNA, yet they display normal levels of Fas ligand. This is not an inherited defect and is confined to mucosal T cells. To gain insights into the mechanism responsible for these findings, we performed a set of experiments with both purified Dermatophagoides pteronyssinus allergen and recombinant human cytokines: interleukin 2 (IL-2), IL-4, IL-5, transforming growth factor beta1, interferon gamma, and granulocyte-macrophage colony-stimulating factor (GM-CSF). In vitro exposure of purified CD4(+) lymphocytes to allergen yielded only transient up-regulation of surface Fas but did not influence susceptibility to Fas-mediated cell death. T-helper type 2 cytokines (IL-4, IL-5, and GM-CSF) had a dose-dependent and specific inhibitory effect on Fas mRNA, suggesting a new fundamental biological role in the survival of inflammatory cells during allergen exposure.

Topics: Adolescent; Adult; Antigens, Dermatophagoides; Asthma; CD4-Positive T-Lymphocytes; Cell Death; Child; Cytokines; Down-Regulation; fas Receptor; Female; Glycoproteins; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Interferon-gamma; Interleukins; Lymphocyte Activation; Male; Receptors, Tumor Necrosis Factor; Th2 Cells; Transforming Growth Factor beta

Chronic airway inflammation and remodeling, including fibrosis, have been proposed as important contributors to asthma pathophysiology. Previous studies of airway fibrosis have been performed mainly in mild and moderate asthmatics at the subepithelial "basement membrane" (SBM) level. The current study was designed to evaluate the large airway SBM thickness and submucosal collagen deposition, as measured by three different collagen staining methods, in endobronchial biopsies from 17 severe, nine moderate, and seven mild asthmatics, as well as eight normal control subjects. Tissue eosinophils and transforming growth factor-beta (TGF-beta) immunoreactivity were also examined. There were no statistically significant differences in the SBM thickness, submucosal collagen deposition, eosinophil numbers, or TGF-beta positive cells among the three groups of asthmatics and the normal control subjects. It was only when examining all asthmatics (n = 33) together, that a modestly thickened SBM (p = 0.04), as evaluated by collagen type III immunostaining, was observed as compared with normal control subjects. Despite this difference, no significant differences were found in the amount of submucosal collagen deposition and the number of eosinophils or TGF-beta expressing cells when comparing total asthmatics and normal control subjects. Additionally, no significant correlations were found between collagen deposition and eosinophil count, TGF-beta expression level, FEV1, or duration of asthma. These results suggest that although increased collagen deposition in the SBM at the large airway level is a characteristic of asthma, it may not explain the differences in severity of asthma.

Topics: Adult; Asthma; Azo Compounds; Basement Membrane; Biopsy; Bronchi; Bronchitis; Bronchoscopy; Collagen; Coloring Agents; Eosinophils; Female; Fibrosis; Forced Expiratory Volume; Humans; Leukocyte Count; Lissamine Green Dyes; Male; Middle Aged; Mucous Membrane; Time Factors; Transforming Growth Factor beta

Interleukin-10 (IL-10) and transforming growth factor beta (TGF-beta) are inhibitory for B and T cells, IgE production, and mast cell proliferation, and they induce apoptosis in eosinophils. These cytokines are therefore candidate genes which could contribute to the development of asthma or allergies. We investigated the hypothesis that polymorphic nucleotides within the IL-10 and TGF-beta gene promoters would link to the expression of allergies and asthma. DNA taken from families with an asthmatic proband was examined for base exchanges by single-stranded conformational polymorphism (SSCP). We demonstrated the presence of a polymorphism in the promoter region of the IL-10 gene and four in the TGF-beta gene promoters (3 in TGF-beta1 and 1 in TGF-beta2). The IL-10 gene polymorphism was a C-to-A exchange 571 base pairs upstream from the translation start site and was present between consensus binding sequences for Sp1 and elevated total serum. This polymorphism was associated with elevated total serum IgE in subjects heterozygotic or homozygotic for this base exchange (p < 0.009). The base exchange at -509 (from the transcription initiation site) in the TGF-beta promoter also linked to elevated total IgE (p < 0.01). This polymorphism represented a C-to-T base exchange which induced a YY1 consensus sequence and is present in a region of the promoter associated with negative transcription regulation.

Topics: Adenine; Apoptosis; Asthma; B-Lymphocytes; Base Pairing; Base Sequence; Cell Division; Child; Child, Preschool; Consensus Sequence; Cytosine; Eosinophils; Gene Expression Regulation; Humans; Hypersensitivity; Immunoglobulin E; Interleukin-10; Mast Cells; Polymorphism, Genetic; Polymorphism, Single-Stranded Conformational; Promoter Regions, Genetic; Protein Biosynthesis; Sp1 Transcription Factor; T-Lymphocytes; Thymine; Transforming Growth Factor beta

Airway wall remodelling is an established pathological feature of asthma but its causes are not well understood. One cytokine of potential relevance is transforming growth factor-beta1 (TGF-beta 1). The immunolocalization of TGF-beta 1 and of its small binding proteoglycan decorin have been examined in the airways of normal subjects and atopic asthmatics. Bronchial biopsy specimens were obtained by fibreoptic bronchoscopy, processed into glycolmethacrylate resin, and stained immunohistochemically using specific antibodies. Immunoreactive TGF-beta 1 was principally localized extracellularly in association with subepithelial connective tissue. Some staining of bronchial epithelial cells was also evident, but otherwise there was little intracellular staining. The overall pattern of immunohistochemical staining was indistinguishable in biopsy specimens from asthmatic and control subjects. Comparison of adjacent sections demonstrated the co-localization of immunoreactivity for TGF-beta 1 and decorin in the mucosa. It is concluded that immunoreactive TGF-beta 1 in human airways is principally extracellular and that matrix-associated TGF-beta 1 is likely to be bound at least in part to decorin. This interaction may provide a reservoir of TGF-beta 1 that can be released in an active form in response to appropriate stimuli.

Topics: Adult; Asthma; Biopsy; Bronchi; Decorin; Extracellular Matrix Proteins; Female; Humans; Immunoenzyme Techniques; Male; Middle Aged; Proteoglycans; Transforming Growth Factor beta

Asthma is characterized by alterations of the bronchial epithelium associated with inflammatory cell infiltrates and sub-epithelial fibrosis. Transforming Growth Factor-beta (TGF-beta) is an anti-inflammatory and fibrosing cytokine normally present in bronchial epithelial cells and also potentially produced by inflammatory cells. Thus, TGF-beta could play a role in the asthmatic process, and its expression could be modified in asthmatic airways.. To test this latter hypothesis, we studied the bronchial distribution of TGF-beta in asthmatic patients.. TGF-beta 1, 2, 3 distribution was studied by immunohistochemistry in bronchial biopsies from 12 asthmatic patients and 10 non-asthmatic subjects.. Bronchial epithelial cells from asthmatics were negative or faintly positive while a bright staining was detected in these from non-asthmatics (P < 0.01). In both groups, when inflammatory cells were present beneath the basement membrane, they were stained by the anti-TGF-beta antibody.. This study shows an altered compartimentalization of TGF-beta in asthma. (a) TGF-beta is scarse in asthmatic bronchial epithelial cells, which could favour the perennization of the bronchial inflammation, and (b) TGF-beta is present in inflammatory cells beneath the basement membrane, where it could be involved in the frequent sub-epithelial fibrosis.

Topics: Adult; Aged; Asthma; Basement Membrane; Bronchi; Case-Control Studies; Epithelium; Humans; Immunity, Mucosal; Immunohistochemistry; Middle Aged; Tissue Distribution; Transforming Growth Factor beta

We assessed whether transforming growth factor-beta (TGF-beta), a fibrogenic growth factor, may be involved in remodeling of asthma and chronic bronchitis; its expression was compared with that of epidermal growth factor (EGF) and granulocyte macrophage colony-stimulating factor (GM-CSF) in bronchial mucosal biopsies from 13 normal subjects, 24 asthmatics, and 19 patients with chronic bronchitis. TGF-beta immunoreactivity was highly increased in epithelium and submucosa of those with bronchitis and to a lesser extent in asthmatics. By comparison, with normal subjects, EGF immunoreactivity was significantly increased in the epithelium of bronchitic subjects and submucosa of asthmatics, and, GM-CSF immunoreactivity was increased in both epithelial and submucosal cells of asthmatics and to a lesser extent in submucosa of bronchitics. A significant correlation was found between the number of epithelial or submucosal cells expressing TGF-beta in both asthma and chronic bronchitis and basement membrane thickness and fibroblast number. No such correlation was found for EGF or GM-CSF. in situ hybridization for TGF-beta 1 mRNA confirmed the results obtained by immunohistochemistry. By combining in situ hybridization and immunohistochemistry, it was found that eosinophils and fibroblasts were synthetizing TGF-beta in asthma and bronchitis. These data suggest that TGF-beta, but not EGF or GM-CSF, is involved in airways remodeling in asthma and chronic bronchitis.

Topics: Adolescent; Adult; Aged; Asthma; Biopsy; Bronchi; Bronchitis; Chronic Disease; Epidermal Growth Factor; Epithelium; Granulocyte-Macrophage Colony-Stimulating Factor; Humans; Immunohistochemistry; In Situ Hybridization; Middle Aged; Mucous Membrane; Transforming Growth Factor beta

Airway wall remodeling is an established pathological feature in asthma. Its causes are not well understood, but one mediator of potential relevance is transforming growth factor-beta 1 (TGF-beta 1). We have measured levels of immunoreactive TGF-beta 1 in bronchoalveolar lavage (BAL) fluid from clinically stable atopic asthmatics and healthy control subjects. We have also examined the influence of allergen exposure on TGF-beta 1 release in the airways using a segmental bronchoprovocation model, with BAL performed at two time points following endobronchial allergen and sham saline challenges. Basal concentrations of TGF-beta 1 were significantly higher in asthmatics than control subjects (median 8.0 versus 5.5 pg/ml, p = 0.027). Following segmental bronchoprovocation, concentrations of TGF-beta 1 at the allergen- and saline-challenged sites were not significantly different after 10 min, (31.3 versus 25.0 pg/ml, p = 0.78), but after 24 h there were significantly higher TGF-beta 1 concentrations at the allergen-challenged sites (46.0 versus 21.5 pg/ml, p = 0.017). We conclude that basal TGF-beta 1 levels in the airways are elevated in atopic asthma and that these levels increase further in response to allergen exposure. These findings are consistent with the hypothesis that TGF-beta 1 is implicated in airway wall remodeling in asthma.

Topics: Adult; Allergens; Asthma; Biopsy; Bronchi; Bronchial Provocation Tests; Bronchoalveolar Lavage Fluid; Bronchoscopy; Female; Fiber Optic Technology; Humans; Hypersensitivity, Immediate; Male; Statistics, Nonparametric; Time Factors; Transforming Growth Factor beta

The histopathology of bronchial asthma is associated with structural changes within the airways, including subepithelial fibrosis, as well as chronic eosinophilic inflammation. The mechanisms responsible for this tissue remodeling, and in particular the role of inflammatory cells, remain to be established. Transforming growth factor-beta (TGF-beta) is a potent profibrotic cytokine which may contribute to the thickening of the reticular lamina by the deposition of collagen fibers. To investigate the molecular mechanisms underlying these structural changes, we have investigated the expression of TGF-beta1 mRNA and immunoreactivity within the bronchial mucosa of mild to severe asthmatic individuals and normal control subjects using the techniques of in situ hybridization and immunocytochemistry. As eosinophils are prominent within the asthmatic airway and are known to synthesize pro-inflammatory cytokines, the presence of TGF-beta1 mRNA and immunoreactive protein in eosinophils was also examined. Asthmatic individuals exhibited a greater expression of TGF-beta1 mRNA and immunoreactivity in the airways submucosa than normal control subjects (P < 0.05), and these increases were directly related to the severity of the disorder. The extent of airways fibrosis, as detected histochemically, was also increased in asthmatics compared with normal control subjects (P < 0.005). In asthmatic subjects, the presence of subepithelial fibrosis was associated with the severity of the disease and correlated with the decline in forced expiratory volume in 1 s (r2 = 0.78; P < 0.05). Within the asthmatic airways, EG2-positive eosinophils represented the major source of TGF-beta1 mRNA and immunoreactivity. These results provide evidence that TGF-beta1 may play a role in the fibrotic changes occurring within asthmatic airways and that activated eosinophils are a major source of this cytokine.

Topics: Adult; Asthma; Bronchi; Eosinophils; Female; Gene Expression; Histocytochemistry; Humans; In Situ Hybridization; Male; Pulmonary Fibrosis; RNA, Messenger; Transforming Growth Factor beta

We have shown that interleukin-5 (IL-5) and granulocyte-macrophage colony-stimulating factor (GM-CSF) are present in sputum from patients experiencing acute asthma attacks, by eosinophil survival assay. The viability of guinea-pig eosinophils was significantly increased in the presence of such sputum extracts after 3 days' culture, and it was inhibited by the addition of anti-IL-5 and anti-GM-CSF antibodies. However, the contribution of IL-5 to the increase in eosinophil viability was less than expected from the values of IL-5 measured by enzyme-linked immunosorbent assay (ELISA). Therefore, we speculated that something in sputum inhibited the function of IL-5.. Transforming growth factor-beta (TGF-beta) was the only cytokine we tested that inhibited the prolongation of survival of guinea-pig eosinophils induced by IL-5. The objective of this study is to detect TGF-beta in the same sputum.. Guinea-pig eosinophils were cultured with or without anti-TGF-beta antibody in the presence of sputum extracts, and the eosinophil viability was counted after 3 days. Measurement of TGF-beta 1 in sputum was performed by ELISA.. Eosinophil viabilities with and without anti-TGF-beta antibody were 79.7 +/- 2.9% and 69.0 +/- 2.7%, respectively, and the difference between them was statistically significant (P < 0.05, n = 9). The concentration of TGF-beta 1 in the sputum was 21.7 +/- 3.3 ng/mL (n = 9).. These observations suggest that TGF-beta is present in sputum from patients with bronchial asthma.

Topics: Adult; Aged; Animals; Antibodies, Monoclonal; Asthma; Cell Survival; Cells, Cultured; Enzyme-Linked Immunosorbent Assay; Eosinophils; Female; Guinea Pigs; Humans; Interleukin-5; Male; Middle Aged; Sputum; Transforming Growth Factor beta

The increase in thickness of bronchial walls by such structural changes as subepithelial fibrosis contributes to the severity and chronicity of asthma by amplifying airway narrowing. However, the pathogenesis of this structural alteration is not known. Transforming growth factor beta 1 (TGF beta 1) is known to have biologic activities relevant to the cellular and molecular events in subepithelial fibrosis, such as the deposition of collagen I and III and the increase of myofibroblasts beneath the epithelial basement membrane. Therefore, we examined TGF beta 1 gene expression in bronchial biopsy tissues from five severe asthmatics, five mild asthmatics, and five normal subjects using in situ hybridization combined with histochemical staining. Cells expressing TGF beta 1 mRNA were detected in tissues from four normal subjects, one mild asthmatic, and five severe asthmatics. The density of positive cells in severe asthmatic tissues (52.1 +/- 22.7, mean +/- SD/mm2) was significantly greater than that in mild asthmatic tissues (1.0 +/- 1.9/mm2, P < 0.01) or normal tissues (10.5 +/- 10.6/mm2, P < 0.02). The density in mild asthmatic tissues was not significantly different from that in normal tissues. The vast majority of positive cells in severe (99.1 +/- 1.7%) and mild (100%) asthmatic tissues were identified as eosinophils. In contrast, eosinophils constituted a small portion of positive cells (20.8 +/- 21.6%) in normal tissues. These results indicated that TGF beta 1 mRNA was overexpressed in severe asthmatics and that the main source of the mRNA was eosinophils, suggesting that eosinophils play an important role in the pathogenesis not only of inflammation but also of structural changes, such as subepithelial fibrosis, in asthmatic airways.

Topics: Adult; Asthma; Bronchi; Eosinophils; Female; Gene Expression; Histocytochemistry; HL-60 Cells; Humans; In Situ Hybridization; Inflammation; Male; Middle Aged; RNA, Messenger; Transforming Growth Factor beta

Asthma and chronic bronchitis are associated with airway remodelling, and airway macrophages are present in bronchial inflammation. TGF-beta and fibronectin released by alveolar macrophages possess a fibrogenic potency. The potential role of alveolar macrophages in airway remodelling was studied in asthma and chronic bronchitis by the release of TGF-beta and fibronectin. Alveolar macrophages were isolated by bronchoalveolar lavage in 14 control subjects, 14 asthmatics and 14 chronic bronchitics. The spontaneous and lipopolysaccharide (LPS)- or concanavalin A (Con A)-induced release of TGF-beta and fibronectin was measured by ELISA. Alveolar macrophages from chronic bronchitics spontaneously release greater amounts of TGF-beta and fibronectin than those from asthmatic and control subjects. Alveolar macrophages from asthmatics release greater amounts of TGF-beta and fibronectin than those from control subjects. The spontaneous release of TGF-beta is significantly correlated with that of fibronectin. Fibronectin release was significantly reduced after LPS stimulation, and TGF-beta release was significantly increased after LPS stimulation, except in chronic bronchitis patients. Con A increased the release of TGF-beta in cells from normal subjects. This study suggests that activated macrophages play a role in airway remodelling in chronic bronchitis and to a lesser extent in asthma.

Topics: Adult; Asthma; Bronchitis; Chronic Disease; Concanavalin A; Female; Fibronectins; Humans; Lipopolysaccharides; Macrophages, Alveolar; Male; Middle Aged; Transforming Growth Factor beta

T lymphocytes expressing the alpha E beta 7 integrin are localized and selectively retained in mucosal tissues. To investigate a potential relationship between alpha E beta 7 expression and pulmonary inflammation, the distribution of alpha E beta 7-bearing CD4+ and CD8+ T cells in peripheral blood and bronchoalveolar lavage (BAL) fluids obtained from patients with allergic asthma, sarcoidosis, hypersensitivity pneumonitis, and idiopathic pulmonary fibrosis (IPF) was determined. In contrast to the distribution in peripheral blood, BAL fluid from these patients contained high number of cells expressing alpha E beta 7 with markedly different expression patterns on CD4 or CD8 cells as well as among the various diseases. Despite similar numbers of activated CD4 cells, alpha E beta 7+CD4+ T cells ranged from 15% in asthmatics to 70% in IPF. In contrast, even in normal individuals, 60% to 90% of BAL fluid CD8+ T cells express alpha E beta 7, suggesting differential induction mechanisms on CD4 and CD8 cells. In vitro experiments revealed that a substantial proportion of peripheral blood CD+ T cells express alpha E beta 7 after stimulation with anti-CD3 antibodies, and up to 80% positive cells were found after the addition of TGF-beta. In contrast, less than 10% of CD4 cells express this particular integrin after in vitro stimulation, and the presence of TGF-beta only increased the number to 30%. Supernatants from in vitro-activated BAL cells as well as concentrated BAL fluid from patients with high alpha E beta 7 expression had no further enhancing effect. However, crosslinking of alpha 4 beta 1-, but not beta 2-integrins, significantly increased the number of alpha E beta 7 expressing CD4+ and CD8+ T cells, even in the absence of TGF-beta. These data indicate that in addition to TGF-beta, the interaction of particular T-cell subsets with specific endothelial cell and extracellular matrix proteins may upregulate alpha E beta 7 integrin expression and thereby contribute to the selective accumulation of these cells in inflammatory lung diseases.

Topics: Adult; Alveolitis, Extrinsic Allergic; Antibodies; Asthma; Bronchoalveolar Lavage Fluid; Cells, Cultured; Cross-Linking Reagents; Eosinophils; Female; HLA-DR Antigens; Humans; Integrin alpha4beta1; Integrins; Leukocyte Common Antigens; Lung Diseases, Interstitial; Lymphocyte Activation; Lymphocyte Subsets; Macrophages, Alveolar; Male; Middle Aged; Neutrophils; Pulmonary Fibrosis; Receptors, Interleukin-2; Receptors, Lymphocyte Homing; Sarcoidosis; T-Lymphocytes; Transforming Growth Factor beta

Topics: Animals; Asthma; Bronchial Provocation Tests; Cells, Cultured; Histamine; In Vitro Techniques; Inflammation; Interleukin-1; Interleukin-11; Interleukin-4; Lung; Mice; Mice, Inbred BALB C; Respiratory Tract Infections; Transforming Growth Factor beta; Virus Diseases

Asthmatic airways have a characteristic deposition of connective tissue under the epithelial basement membrane, but the mediators involved in this alteration are unknown. Several authors have postulated that transforming growth factor beta 1 (TGF-beta 1) could be overexpressed in asthmatic airways.. Lung samples from 16 asthmatic patients, six patients with chronic obstructive pulmonary disease (COPD), and six non-obstructed smokers were analysed. RNA was extracted from these tissues to measure expression of TGF-beta 1 by Northern blot analysis using a cDNA probe for TGF-beta 1. The level of expression was quantitated by densitometry using glyceraldehyde 3-phosphate dehydrogenase mRNA as a control. TGF-beta 1 was localised to specific cell types in these lungs by immunohistochemical analysis using polyclonal antibodies specific for intracellular and extracellular TGF-beta 1.. The 2.5 kb TGF-beta 1 mRNA was seen in all 18 samples analysed by Northern blotting and densitometric analysis showed no difference between the asthmatic group (mean (SD) 108% (43%)), the group with COPD (122% (33%)), and the non-obstructed group (100% (49%)). The TGF-beta 1 precursor was immunolocalised throughout the airway wall including the epithelium and in alveolar macrophages. The mature TGF-beta 1 was localised primarily within the connective tissue of the airway wall. These patterns of expression of both forms of TGF-beta 1 were similar in lungs from asthmatic patients, those with COPD, and controls.. While TGF-beta 1 mRNA and protein are abundantly expressed in human lungs, there is no clear difference in expression between the airways of asthmatic subjects and those of smokers with and without COPD.

Topics: Adolescent; Adult; Aged; Aged, 80 and over; Asthma; Blotting, Northern; Child; Child, Preschool; DNA Probes; Female; Gene Expression; Humans; Immunohistochemistry; Lung; Lung Diseases, Obstructive; Male; Middle Aged; RNA, Messenger; Smoking; Transforming Growth Factor beta