oxazolone and Inflammation

T helper type 2 (Th2)-characterized inflammatory responses are highly dynamic processes initiated by epithelial cell damage resulting in remodelling of the tissue architecture to prevent further harm caused by a dysfunctional epithelial barrier or migrating parasites. This process is a temporal and spatial response which requires communication between immobile cells such as epithelial, endothelial, fibroblast and muscle cells and the highly mobile cells of the innate and adaptive immunity. It is further characterized by a high cellular plasticity that enables the cells to adapt to a specific inflammatory milieu. Incipiently, this milieu is shaped by cytokines released from epithelial cells, which stimulate Th2, innate lymphoid and invariant natural killer (NK) T cells to secrete Th2 cytokines and to activate dendritic cells which results in the further differentiation of Th2 cells. This milieu promotes wound-healing processes which are beneficial in parasitic infections or toxin exposure but account for increasingly dysfunctional vital organs, such as the lung in the case of asthma and the colon in ulcerative colitis. A better understanding of the dynamics underlying relapses and remissions might lead ultimately to improved therapeutics for chronic inflammatory diseases adapted to individual needs and to different phases of the inflammation.

Topics: Animals; Cellular Microenvironment; Chronic Disease; Disease Models, Animal; Humans; Inflammation; Neoplasms; Oxazolone; Th2 Cells

NOD2 has emerged as a critical player in the induction of both Th1 and Th2 responses for potentiation and polarisation of antigen-dependent immunity. Loss-of-function mutations in the NOD2-encoding gene and deregulation of its downstream signalling pathway have been linked to Crohn's disease. Although it is well documented that NOD2 is capable of sensing bacterial muramyl dipeptide, it remains counter-intuitive to link development of overt intestinal inflammation to a loss of bacterial-induced inflammatory response. We hypothesised that a T helper bias could also contribute to an autoimmune-like colitis different from inflammation that is fully fledged by Th1 type cells.. An oedematous bowel wall with a mixed Th1/Th2 response was induced in mice by intrarectal instillation of the haptenating agent oxazolone. Survival and clinical scoring were evaluated. At several time points after instillation, colonic damage was assessed by macroscopic and microscopic observations. To evaluate the involvement of NOD2 in immunochemical phenomena, quantitative polymerase chain reaction [PCR] and flow cytometry analysis were performed. Bone marrow chimera experimentation allowed us to evaluate the role of haematopoietic/non-hematopoietic NOD2-expressing cells.. Herein, we identified a key regulatory circuit whereby NOD2-mediated sensing of a muramyl dipeptide [MDP] by radio-resistant cells improves colitis with a mixed Th1/Th2 response that is induced by oxazolone. Genetic ablation of either Nod2 or Ripk2 precipitated oxazolone colitis that is predominantly linked to a lack of interferon-gamma. Bone marrow chimera experiments revealed that inactivation of Nod2 signalling in non-haematopoietic cells is causing a biased M1-M2 polarisation of macrophages and a decreased frequency of splenic regulatory T cells that correlates with an impaired activation of CD4 + T cells within mesenteric lymph nodes. Mechanistically, mice were protected from oxazolone-induced colitis upon administration of MDP in an interleukin-1- and interleukin-23-dependent manner.. These findings indicate that Nod2 signalling may prevent pathological conversion of T helper cells for maintenance of tissue homeostasis.

Topics: Acetylmuramyl-Alanyl-Isoglutamine; Animals; Colitis; Inflammation; Mice; Nod2 Signaling Adaptor Protein; Oxazolone; Signal Transduction

Dermal regulatory T cells (Tregs) are essential for maintenance of skin homeostasis and control of skin inflammatory responses. In mice, Tregs in the skin are characterized by high expression of CD103, the αE integrin. Evidence indicates that CD103 promotes Treg retention within the skin, although the mechanism underlying this effect is unknown. The main ligand of CD103, E-cadherin, is predominantly expressed by cells in the epidermis. However, because Tregs are predominantly located within the dermis, the nature of the interactions between E-cadherin and CD103-expressing Tregs is unclear. In this study, we used multiphoton intravital microscopy to examine the contribution of CD103 to Treg behavior in resting and inflamed skin of mice undergoing oxazolone-induced contact hypersensitivity. Inhibition of CD103 in uninflamed skin did not alter Treg behavior, whereas 48 h after inducing contact hypersensitivity by oxazolone challenge, CD103 inhibition increased Treg migration. This coincided with E-cadherin upregulation on infiltrating myeloid leukocytes in the dermis. Using CD11c-enhanced yellow fluorescent protein (EYFP) × Foxp3-GFP dual-reporter mice, inhibition of CD103 was found to reduce Treg interactions with dermal dendritic cells. CD103 inhibition also resulted in increased recruitment of effector CD4+ T cells and IFN-γ expression in challenged skin and resulted in reduced glucocorticoid-induced TNFR-related protein expression on Tregs. These results demonstrate that CD103 controls intradermal Treg migration, but only at later stages in the inflammatory response, when E-cadherin expression in the dermis is increased, and provide evidence that CD103-mediated interactions between Tregs and dermal dendritic cells support regulation of skin inflammation.

Topics: Animals; Cadherins; Dermatitis, Contact; Inflammation; Integrin alpha Chains; Mice; Oxazolone; T-Lymphocytes, Regulatory

Topics: Animals; Cytokines; Dermatitis, Atopic; Disease Models, Animal; Glycine max; Inflammation; Mice; Mice, Inbred BALB C; Oxazolone; Plant Extracts; Skin

Clinical challenges in inflammatory bowel diseases require microscopic in vivo evaluation of inflammation. Here, label-free imaging holds great potential, and recently, our group demonstrated the advantage of using in vivo multiphoton endomicroscopy for longitudinal animal studies. This article extends our previous work by in-depth analysis of label-free tissue features in common colitis models quantified by the multiphoton colitis score (MCS).. Fresh mucosal tissues were evaluated from acute and chronic dextran sulfate sodium (DSS), TNBS, oxazolone, and transfer colitis. Label-free imaging was performed by using second harmonic generation and natural autofluorescence. Morphological changes in mucosal crypts, collagen fibers, and cellularity in the stroma were analyzed and graded.. Our approach discriminated between healthy (mean MCS = 2.5) and inflamed tissue (mean MCS > 5) in all models, and the MCS was validated by hematoxylin and eosin scoring of the same samples (85.2% agreement). Moreover, specific characteristics of each phenotype were identified. While TNBS, oxazolone, and transfer colitis showed high cellularity in stroma, epithelial damage seemed specific for chronic, acute DSS and transfer colitis. Crypt deformations were mostly observed in acute DSS.. Quantification of label-free imaging is promising for in vivo endoscopy. In the future, this could be valuable for monitoring of inflammatory pathways in murine models, which is highly relevant for the development of new inflammatory bowel disease therapeutics.

Topics: Animals; Colitis; Dextran Sulfate; Disease Models, Animal; Inflammation; Inflammatory Bowel Diseases; Mice; Oxazolone

The functions of basophils have not been elucidated until recently because of their rarity. However, with recent developments in basophil-specific antibodies and basophil-deficient animals, the roles of basophils in various diseases related to chronic inflammation have been clarified. In this study, we aimed to investigate the roles of basophils in human ulcerative colitis (UC) and oxazolone (OXA) colitis using genetically engineered Mcpt8. Immunohistochemical staining of human colon specimens was performed to examine the involvement of basophils in the pathogenesis of UC. We examined the correlation between the number of infiltrating basophils and the UC endoscopic index of severity (UCEIS), Mayo score, and Matts score. We also examined the correlation between eosinophil count and basophil infiltration. In murine experiments, we examined whether basophil infiltration was involved in OXA-induced colitis and whether basophil depletion improved inflammation in Mcpt8. Colonic basophil infiltration was significantly increased in patients with UC. There were significant correlations between UCEIS, Mayo score, Matts score, and the number of infiltrating basophils. In murine OXA-induced colitis, a significant increase in basophil infiltration was observed. When basophils were depleted by diphtheria toxin in Mcpt8. Basophil infiltration correlated with endoscopic, clinical, and pathological scores in human UC independently of eosinophil infiltration, and depletion of basophils ameliorated mucosal inflammation in murine OXA-induced colitis, collectively suggesting that basophils exert a proinflammatory role in chronic intestinal inflammation such as UC.

Topics: Animals; Basophils; Colitis; Colitis, Ulcerative; Humans; Inflammation; Intestines; Mice; Oxazolone

Contact allergy is a common skin allergy, which can be studied utilising contact hypersensitivity (CHS) animal model. However, it is not clear, whether CHS is a suitable model to investigate skin microbiota interactions. We characterised the effect of contact dermatitis on the skin microbiota and studied the biological effects of oxazolone (OXA) -induced inflammation on skin thickness, immune cell numbers and changes of the microbiota in CHS mouse model (n = 72) for 28 days. Through 16S rRNA gene sequencing we defined the composition of bacterial communities and associations of bacteria with inflammation. We observed that the vehicle solution of acetone and olive oil induced bacterial community changes on day 1, and OXA-induced changes were observed mainly on day 7. Many of the notably enriched bacteria present in the OXA-challenged positive group represented the genus Faecalibaculum which were most likely derived from the cage environment. Additionally, skin inflammation correlated negatively with Streptococcus, which is considered a native skin bacterium, and positively with Muribacter muris, which is typical in oral environment. Skin inflammation favoured colonisation of cage-derived faecal bacteria, and additionally mouse grooming transferred oral bacteria on the skin. Due to the observed changes, we conclude that CHS model could be used for certain skin microbiome-related research set-ups. However, since vehicle exposure can alter the skin microbiome as such, future studies should include considerations such as careful control sampling and statistical tests to account for potential confounding factors.

Topics: Acetone; Animals; Bacteria; Dermatitis, Allergic Contact; Disease Models, Animal; Inflammation; Mice; Microbiota; Olive Oil; Oxazolone; RNA, Ribosomal, 16S

Dictamnine is an active pharmaceutical ingredient in Dictamnus dasycarpus, a Chinese herbal medicine widely used for the treatment of skin inflammations such as atopic dermatitis (AD). Oxazolone has been demonstrated to induce significant skin inflammation and produce inflammatory cytokine expression identical to that of AD. An in vitro HaCaT inflammation model treated with dictamnine, which efficiently scavenged the reactive oxygen species (ROS) and mitochondrial ROS (mROS), and it reduced interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) expression, NLRP3 inflammasome activation, and NF-κB expression. To explore the anti-inflammatory mechanism of dictamnine and enhance sustained drug release and penetration into epidermal structures in a dermatitis mouse model, we prepared PLGA-nanocarrier-encapsulated dictamnine (Dic-PLGA-NC) in a specifically designed bioreactor, namely an ultrasound composite streams-impinging mixer (U-SiM). Mouse dermatitis model was treated with Dic-PLGA-NC medication, spleens were collected to evaluate body weight ratio, and skin was retrieved for histological examination and two-photon microscopy. The data demonstrate that Dic-PLGA-NC efficiently penetrated the dermal layer, making it superior to naked dictamnine; moreover, it ameliorated the dermatitis symptoms and inflammatory cytokine expression in vivo. Dic-PLGA-NC produced using the U-SiM bioreactor could be used in new manufacturing processes for drugs to treat AD.

Topics: Animals; Cytokines; Dermatitis, Atopic; Disease Models, Animal; Inflammation; Mice; NF-kappa B; Oxazolone; Quinolines; Skin; Tumor Necrosis Factor-alpha

Ulcerative colitis (UC) is a chronic inflammatory bowel disease that is associated with high sphingosine kinase 1(SPHK1) expression in the colon, however its role in pathogenesis of UC is not clearly understood so, the aim of the present study was to clarify the role of SPHK1 and investigate whether the anti-inflammatory effects of metformin in UC is mediated by Sphingosine kinase 1/sphingosine 1 phosphate (S1P) signaling pathway.. Colitis was induced in adult male wistar rats by intra rectal administration of oxazolone in the fifth and seventh days from initial presensitization. Oxazolone treated rats were divided into untreated oxazolone group, metformin and mesalazine treated groups both in a dose of 100 mg/kg/day orally for 21 days. Along with these groups normal control and saline groups were used .Colitis was assessed by colon length, disease activity index (DAI) and histological examination of colontissue. Plasma samples were used to measure S1P.SPHK1 activity, signal transducer and activator of transcription -3(STAT-3), interleukin-6 (IL-6), nitric oxide (NO), myeloperoxidase activity (MPO), reduced glutathione (GSH) and tissue expression of intracellular cell adhesion molecule -1(ICAM-1) and caspase-3 genes were measured in tissue.. Metformin successfully attenuated oxazolone colitis by increasing colon length, decreasing DAI and improved colon histologic picture. Metformin also induced a significant decrease in Plasma SIP, SPHK1 activity, inflammatory, oxidative stress markers, ICAM-1 and Caspase-3 genes expression compared to oxazolone group.. It is revealed that metformin alleviated inflammation and underlying mechanism may result from inhibition of SPHK1/S1P signaling pathway.

Topics: Animals; Colitis, Ulcerative; Colon; Inflammation; Lysophospholipids; Male; Metformin; Oxazolone; Phosphotransferases (Alcohol Group Acceptor); Rats; Rats, Wistar; Signal Transduction; Sphingosine

Atopic dermatitis (AD) is a chronic inflammatory disease of the skin, characterized by dryness and more or less severe itching. The etiology of AD is complex and has not been fully clarified, involving genetic susceptibility, immunological abnormalities, epidermal barrier dysfunction, and environmental factors. Xyloglucan (XG) and pea protein (PP) are two compounds of natural origin characterized by the ability to create a physical barrier that protects mucosae membranes, reducing inflammation. The aim of the present study was to evaluate the potential beneficial effects of XG + PP in both a mouse model of AD and

Topics: Animals; Cell Degranulation; Cytokines; Dermatitis, Atopic; Disease Models, Animal; Erythema; Female; Filaggrin Proteins; Glucans; Inflammation; Intermediate Filament Proteins; Mast Cells; Mice; Nitric Oxide Synthase Type II; Occludin; Oxazolone; Pea Proteins; Skin; Staphylococcal Infections; Staphylococcus aureus; Tight Junctions; Xylans

Acute orbital inflammation can lead to irreversible vision loss in serious cases. Treatment thus far has been limited to systemic steroids or surgical decompression of the orbit. An animal model that mimics the characteristic features of acute orbital inflammation as found in thyroid eye disease can be used to explore novel treatment modalities.. We developed a murine model of orbital inflammation by injecting oxazolone into the mouse orbit. The mice underwent magnetic resonance imaging (MRI) and were euthanized at various time points for histologic examination. Immunofluorescence studies of specific inflammatory cells and cytokine arrays were performed.. We found clinical and radiographic congruity between the murine model and human disease. After 72 hours, sensitized mice exhibited periorbital dermatitis and inflammation in the eyelids of the injected side. By one week, increased proptosis in the injected eye with significant eyelid edema was appreciated. By four weeks, inflammation and proptosis were decreased. At all three time points, the mice demonstrated exophthalmos and periorbital edema. Histopathologically, populations of inflammatory cells including T cells, macrophages, and neutrophils shared similarities with patient samples in thyroid eye disease. Proteomic changes in the levels of inflammatory and angiogenic markers correlated to the expected angiogenic, inflammatory, and fibrotic responses observed in patients with thyroid eye disease.. A murine model of orbital inflammation created using oxazolone recapitulates some of the clinical features of thyroid eye disease and potentially other nonspecific orbital inflammation, typified by inflammatory cell infiltration, orbital tissue expansion and remodeling, and subsequent fibrosis.. This animal model could serve as a viable platform with which to understand the underlying mechanisms of acute orbital inflammation and to investigate potential new, targeted treatments.

Topics: Animals; Disease Models, Animal; Graves Ophthalmopathy; Humans; Inflammation; Mice; Oxazolone; Proteomics

Allergic contact dermatitis (ACD) is a common skin disorder affecting an estimated 15-20% of the general population. The mouse model of ACD is contact hypersensitivity (CHS), which consists of two phases: induction and elicitation. Although neutrophils are required for both CHS disease phases their mechanisms of action are poorly understood. Neutrophils release myeloperoxidase (MPO) that through oxidation of biomolecules leads to cellular damage.. This study investigated mechanisms whereby MPO contributes to CHS pathogenesis.. CHS was induced in mice using oxazolone (OX) as the initiating hapten applied to the skin. After 7 days, CHS was elicited by application of OX to the ear and disease severity was measured by ear thickness and vascular permeability in the ear. The role of MPO in the two phases of CHS was determined utilizing MPO-deficient mice and a specific MPO inhibitor.. During the CHS induction phase MPO-deficiency lead to a reduction in IL-1β production in the skin and a subsequent reduction in migratory dendritic cells (DC) and effector T cells in the draining lymph node. During the elicitation phase, inhibition of MPO significantly reduced both ear swelling and vascular permeability.. MPO plays dual roles in CHS pathogenesis. In the initiation phase MPO promotes IL-1β production in the skin and activation of migratory DC that promote effector T cell priming. In the elicitation phase MPO drives vascular permeability contributing to inflammation. These results indicate that MPO it could be a potential therapeutic target for the treatment of ACD in humans.

Topics: Animals; Cell Movement; Dendritic Cells; Dermatitis, Allergic Contact; Dermatitis, Contact; Haptens; Inflammation; Interleukin-1beta; Lymph Nodes; Mice; Mice, Inbred C57BL; Neutrophils; Oxazolone; Peroxidase; Skin; T-Lymphocytes

The interleukin 7 receptor alpha chain (IL-7Rα) is predominately expressed by lymphocytes, and activation by its ligand IL-7 supports the development and maintenance of T cells and boosts T-cell mediated immunity. We recently reported that lymphatic endothelial cells (LECs) in dermal lymphatics also express IL-7 and its receptor chains (IL-7Rα and CD132) and that IL-7 supports lymphatic drainage. This suggested that activation of IL-7Rα signaling in lymphatics could exert inflammation-resolving activity, by promoting the clearance of excess tissue fluid. Here we investigated how the potentially opposing effects of IL-7Rα signaling in immune cells and in the lymphatic vasculature would affect the development and progression of psoriasis-like skin inflammation. We found that during acute and chronic skin inflammation mice with an endothelial-specific deletion of IL-7Rα (IL-7Rα

Topics: Animals; Antibodies, Neutralizing; CD4-Positive T-Lymphocytes; Disease Models, Animal; Endothelial Cells; Female; Gene Expression Regulation; Humans; Imiquimod; Inflammation; Interleukin-7; Lymph Nodes; Lymphatic Vessels; Male; Mice; Mice, Inbred C57BL; Organ Specificity; Oxazolone; Psoriasis; Receptors, Interleukin-7; Signal Transduction; Skin; Tetradecanoylphorbol Acetate

Topics: Animals; Biomarkers; Colitis, Ulcerative; Colon; Cytokines; Diterpenes; Inflammation; Male; Oxazolone; Peroxidase; Rats, Sprague-Dawley; Receptors, Interleukin-4; Signal Transduction; STAT6 Transcription Factor; Transcription Factor RelA

Contact dermatitis and psoriasis are skin disorders caused by immune dysregulation, yet much remains unknown about their underlying mechanisms. Ghrelin, a recently discovered novel peptide and potential endogenous anti-inflammatory factor expressed in the epidermis, is involved in skin repair and disease. In this study, we investigated the expression pattern and therapeutic effect of ghrelin in both contact dermatitis and psoriasis mouse models induced by oxazolone (OXA) and imiquimod (IMQ), respectively, and in TNF-α-stimulated RAW264.7 macrophages, NHEKs and skin fibroblasts. Ghrelin expression was reduced in both the OXA-induced contact dermatitis and IMQ-induced psoriasis mouse models. Furthermore, treatment with ghrelin attenuated skin inflammation in both the contact dermatitis and psoriasis mouse models. Mice administered PBS after OXA- or IMQ-induced model generation exhibited typical skin inflammation, whereas ghrelin treatment in these mouse models substantially decreased the dermatitis phenotype. In addition, exogenous ghrelin attenuated the inflammatory reaction induced by TNF-α in RAW264.7 cells. Moreover, ghrelin administration limited activation of NF-κB signaling. In summary, ghrelin may represent a potential molecular target for the prevention and treatment of inflammatory skin diseases, including contact dermatitis and psoriasis.

Topics: Animals; Dermatitis, Contact; Disease Models, Animal; Fibroblasts; Gene Expression Regulation; Ghrelin; Humans; Imiquimod; Immune System Diseases; Inflammation; Mice; NF-kappa B; Oxazolone; Psoriasis; RAW 264.7 Cells; Signal Transduction; Skin; Tumor Necrosis Factor-alpha

Inflammatory priming of immune cells in early life may optimize the response to a subsequent inflammatory challenge later in life. To prime the immune cells in the gut in vivo through a short inflammatory insult, we administered a low dose of dextran sulfate sodium (DSS) to 5-weeks-old BALB/c mice in the drinking water. We hypothesized that DSS-primed mice would show decreased inflammation and difference in immunological profiling, when subjected to presensitizing and oxazolone-induced colitis by rectal instillation at 9 weeks compared to non-DSS-primed control mice. In fact, this low-dose DSS priming apparently decreased the acute inflammation, as colitis scores along with IFNγ, IL-1ß, and IL-4 were significantly decreased with the same tendency for IL-5, TNFα, and IL-2 on day 3 post-induction compared to control mice. On day 7, both DSS-primed and control mice had significantly higher numbers of FoxP3

Topics: Adjuvants, Immunologic; Animals; Colitis; Colon; Dextran Sulfate; Inflammation; Inflammatory Bowel Diseases; Lymphocytes; Mice; Oxazolone; T-Lymphocytes, Regulatory

Oxazolone-induced colitis has been frequently used in literature as a model of IBD, but insights into the underlying immune response and pathological features are surprisingly still very limited. Vagus nerve stimulation (VNS) has proven to be effective in innate and Th1/Th17 predominant inflammatory models, including pre-clinical models of colitis, however to what extent VNS is also effective in ameliorating Th2-driven colitis remains to be studied. In the present study, we therefore further characterized the immune response in oxazolone-induced colitis and investigated the potential therapeutic effect of VNS.. Colitis was induced in Balb/c mice by cutaneous sensitization with 3% oxazolone followed by intracolonic administration of 1% oxazolone 7 days later. To evaluate the effect of VNS on the development of Th2-driven colitis, VNS and sham-treated mice were challenged with 1% oxazolone.. Intracolonic oxazolone administration resulted in a severe destruction of the colonic mucosa and a rapid drop in body temperature leading to a 65% mortality rate at day 5. Severe infiltration of neutrophils and monocytes was detected 6h after oxazolone administration which was associated with a Th2-type inflammatory response. VNS significantly improved survival rate which correlated with decreased levels of HMGB1 and reduced colonic (il6 and cxcl1 mRNA) and serum cytokine levels (IL-6, TNFα and CXCL1) compared to sham treated mice.. Oxazolone-induced colitis rather represents a model of sepsis and, at best, may resemble a severe type of ulcerative colitis, associated with early and severe mucosal damage and a high mortality rate. VNS reduces colonic inflammation and improves survival in this model, supporting the anti-inflammatory properties of VNS, even in an aggressive model as oxazolone-induced colitis.

Topics: Animals; Colitis; Cytokines; Disease Models, Animal; Female; Hypothermia; Inflammation; Intestinal Mucosa; Mice, Inbred BALB C; Natural Killer T-Cells; Oxazolone; Survival Analysis; Vagus Nerve Stimulation

T cells are the most abundant cell type found in afferent lymph, but their migration through lymphatic vessels (LVs) remains poorly understood. Performing intravital microscopy in the murine skin, we imaged T cell migration through afferent LVs in vivo. T cells entered into and actively migrated within lymphatic capillaries but were passively transported in contractile collecting vessels. Intralymphatic T cell number and motility were increased during contact-hypersensitivity-induced inflammation and dependent on ICAM-1/LFA-1 interactions. In vitro, blockade of endothelial cell-expressed ICAM-1 reduced T cell adhesion, crawling, and transmigration across lymphatic endothelium and decreased T cell advancement from capillaries into lymphatic collectors in skin explants. In vivo, T cell migration to draining lymph nodes was significantly reduced upon ICAM-1 or LFA-1 blockade. Our findings indicate that T cell migration through LVs occurs in distinct steps and reveal a key role for ICAM-1/LFA-1 interactions in this process.

Topics: Animals; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Adhesion; Cell Movement; Flow Cytometry; Inflammation; Intercellular Adhesion Molecule-1; Interferon-gamma; Killer Cells, Natural; Lymph Nodes; Lymphatic Vessels; Lymphocyte Function-Associated Antigen-1; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microscopy, Confocal; Oxazolone; Skin; T-Lymphocytes; Time-Lapse Imaging; Tumor Necrosis Factor-alpha

This study aimed to investigate whether mangiferin played a protective role in a well-established dermatitis mouse model and tumor necrosis factor alpha (TNF-α)-induced RAW264.7 macrophages. Contact dermatitis is an inflammatory skin disease in the clinic, while its underlying mechanism still remains to be elucidated. Mangiferin, 1,3,6,7-tetrahydroxyxanthone-C2-β-d-glucoside (C-glucosyl xanthone), a natural antioxidant that was reported to inhibit inflammatory reactions, has been recently proved to be a potential therapy for inflammation. As a result, the oxazolone-induced dermatitis mice models were established to explore whether mangiferin has an anti-inflammatory role in vivo. The phosphate-buffered saline treatment groups showed emblematic skin inflammation, whereas the administration of mangiferin obviously inhibited dermatitis in the mice models. Furthermore, exogenous mangiferin alleviated the inflammatory reaction in TNF-α-induced macrophages by suppressing the production of inflammation- and oxidative stress-associated molecules. Also, mangiferin treatment repressed the activation of nuclear factor-kappaB signaling pathway. To sum up, mangiferin could provide a new target for the therapy and prevention of skin inflammation.

Topics: Animals; Anti-Inflammatory Agents; Dermatitis, Contact; Disease Models, Animal; Humans; Inflammation; Macrophages; Male; Mice; Mice, Inbred C57BL; Oxazolone; RAW 264.7 Cells; Signal Transduction; Skin; Tumor Necrosis Factor-alpha; Xanthones

P2X7 receptor activation contributes to inflammation development in different pathologies. We previously reported that the P2X7 receptor is over-expressed in the gut mucosa of patients with inflammatory bowel disease, and that P2X7 inhibition protects against chemically induced colitis. Here, we investigated in detail the role of the P2X7 receptor in inflammatory bowel disease development, by treating P2X7 knockout (KO) and WT mice with two different (and established) colitis inductors. P2X7 KO mice were protected against gut inflammation induced by 2,4,6-trinitrobenzenesulfonic acid or oxazolone, with no weight loss or gut histological alterations after treatment. P2X7 receptor knockout induced regulatory T cell accumulation in the colon, as evaluated by qRT-PCR for FoxP3 expression and immunostaining for CD90/CD45RB

Topics: Animals; Colitis; Female; Immunity, Mucosal; Inflammation; Inflammatory Bowel Diseases; Intestinal Mucosa; Mice; Mice, Knockout; Oxazolone; Receptors, Purinergic P2X7; T-Lymphocytes; Trinitrobenzenesulfonic Acid

Anti-oxidant coenzyme Q10 (Co-Q10) is commonly used in clinic. Recently, Co-Q10 was reported to antagonize TNF-α-induced inflammation and play a protective role in various inflammatory conditions. However, its role in dermatitis is unknown. Herein, RAW264.7 macrophage cell line was cultured with stimulation of TNF-α, and administration of Co-Q10 alleviated TNF-α-mediated inflammatory reaction in vitro. Furthermore, oxazolone-induced dermatitis mice model was established, and treatment of Co-Q10 markedly attenuated dermatitis phenotype in this mice model. Moreover, the protective role of Co-Q10 in vitro and in dermatitis was probably due to its repression on NF-κB signaling. Collectively, Co-Q10 may represent a potential molecular target for prevention and treatment of inflammatory skin diseases.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Cell Line; Dermatitis, Contact; Disease Models, Animal; Inflammation; Interleukin-1beta; Interleukin-6; Macrophages; Male; Mice; Mice, Inbred C57BL; NF-kappa B; Oxazolone; Skin; Tumor Necrosis Factor-alpha; Ubiquinone

The inflammatory response is one of the first defenses our body has to fight against potential endangerments. It plays a critical role in host defense, clearing and slowing the infection in the case of microbial invasion. During an inflammatory response, a variety of cytokines are produced by cells and trigger or enhance the specific inflammation response. TNFα, one of these factors, plays a crucial role in many immune and inflammatory processes, such as proliferation, apoptosis, necrosis, and cell survival. It acts in orchestrating the cytokine cascade and the major regulator of inflammatory cytokine production. Abnormality of TNFα signaling leads to many diseases, including rheumatoid arthritis, psoriasis, Crohn's disease, atherosclerosis, and cancer. Due to the importance of TNFα, regulating TNFα activity is a key to treat the related diseases. There is a long history of using medicinal herbs to treat diseases related to inflammation. We searched for an ingredient that has the ability to inhibit TNFα, we examined AO herbal extract, containing 10 individual herbs and most of these herbs have anti-inflammatory activity within humans. We have tested the anti-inflammatory ability of AO herbal extract on mice. Furthermore, we used macrophage cell from young mice and found that AO extract has the ability to reduce the inflammation by inhibiting TNFα level.

Topics: Animals; Anti-Inflammatory Agents; Cells, Cultured; Female; Inflammation; Inflammation Mediators; Macrophages, Peritoneal; Mice, Inbred BALB C; Oxazolone; Phytotherapy; Plant Extracts; Plants, Medicinal; Signal Transduction; Tumor Necrosis Factor-alpha

Carrageenan is a traditional ingredient that has been widely used in the food industry. In the present study, we propose a hypothesis that carrageenan is a conditional inflammatory agent. When the intestinal tract is in an "unhealthy" state such as that during bacterial infection or acute inflammation, carrageenan can synergistically enhance the inflammatory response.. BALB/C mice received κ-carrageenan via intragastric administration prior to the induction of oxazolone colitis. Weight changes, survival rate, histologic change, secretion of inflammatory cytokines, ratio of regulatory T cells (Tregs) in peripheral blood, and expression of genes and proteins involved in inflammation and cell proliferation in the colonic mucosa were examined.. Intragastric administration of κ-carrageenan to BALB/c mice prior to the induction of oxazolone colitis resulted in an aggravation of body weight loss, a decrease in the survival ratio, aggravation of colonic inflammation, and decrease in the ratio of CD4 + CD25+/CD4+. The secretion of interleukin-4 (IL-4), interleukin-10 (IL-10), tumor necrosis factor-α (TNF-α), and interleukin-6 (IL-6) also significantly increased after κ-carrageenan administration. κ-Carrageenan, together with oxazolone, suppressed the expression of forkhead box p3 (FOXp3) and increased the expression of Toll-like receptor 4 (TLR4), Nuclear factor-κB (NF-κB), and proliferating cell nuclear antigen in the colonic mucosa. These results were confirmed by qRT-PCR and western blot analyses at the molecular and protein levels, respectively.. κ-Carrageenan aggravated oxazolone-induced intestinal inflammation in BALB/c mice. This effect is associated with an activation of the TLR4-NF-κB pathway, a decreased ratio of Tregs, and the induction of Th2-dependent immune responses.

Topics: Adjuvants, Immunologic; Animals; Blotting, Western; Carrageenan; Cell Proliferation; Colitis; Colon; Cytokines; Drug Synergism; Forkhead Transcription Factors; Inflammation; Interleukin-10; Interleukin-4; Interleukin-6; Intestinal Mucosa; Mice; Mice, Inbred BALB C; NF-kappa B; Oxazolone; Proliferating Cell Nuclear Antigen; Real-Time Polymerase Chain Reaction; Reverse Transcriptase Polymerase Chain Reaction; Toll-Like Receptor 4; Tumor Necrosis Factor-alpha

Crohn's disease (CD) and ulcerative colitis (UC) have different immunological mechanisms, while both of them are potential targets of vitamin D treatment. In this study, we have tried to address the role of vitamin D in CD and UC using two mouse models. Mice of C57B6L were given vitamin D before the induction of colitis. Our results showed that vitamin D attenuated 2,4,6-trinitrobenzene sulphonic acid (TNBS)-induced colitis but not oxazolone-induced colitis. Vitamin D could preserve the local histology, alleviate inflammation, suppress apoptosis, maintain tight junction function and decrease permeability. Interestingly, it has more of an effect on local structure preservation and inflammation inhibition in CD than in UC mice. Vitamin D blocked the increase of helper T-cell type 1 (Th1)- and helper T-cell type 17 (Th17)-related cytokines in TNBS-induced colitis. But the increase of helper T-cell type 2 (Th2)- and regulatory T cells (Treg)-related cytokines was augmented at the same time in oxazolone-induced colitis which counteracted each other. Our study helps elucidate the differential protective effects of vitamin D on CD and UC patients, as reported in literature.

Topics: Animals; Apoptosis; Colitis, Ulcerative; Crohn Disease; Cytokines; Disease Models, Animal; Female; Inflammation; Intestinal Mucosa; Male; Mice; Mice, Inbred C57BL; Oxazolone; T-Lymphocytes, Regulatory; Th1 Cells; Th17 Cells; Th2 Cells; Trinitrobenzenesulfonic Acid; Vitamin D

Skin inflammation plays a central role in the pathophysiology and symptoms of diverse chronic skin diseases including atopic dermatitis (AD). In this study, we examined if caffeic acid phenethyl ester (CAPE), a skin-permeable bioactive compound from propolis, was protective against skin inflammation using in vitro cell system and in vivo animal disease models. CAPE suppressed TNF-α-induced NF-κB activation and expression of inflammatory cytokines in human keratinocytes (HaCaT). The potency and efficacy of CAPE were superior to those of a non-phenethyl derivative, caffeic acid. Consistently, topical treatment of CAPE (0.5 %) attenuated 12-O-tetradecanoylphorbol-13-acetate(TPA)-induced skin inflammation on mouse ear as CAPE reduced ear swelling and histologic inflammation scores. CAPE suppressed increased expression of pro-inflammatory molecules such as TNF-α, cyclooxygenase-2 and inducible NO synthase in TPA-stimulated skin. TPA-induced phosphorylation of IκB and ERK was blocked by CAPE suggesting that protective effects of CAPE on skin inflammation is attributed to inhibition of NF-κB activation. Most importantly, in an oxazolone-induced chronic dermatitis model, topical application of CAPE (0.5 and 1 %) was effective in alleviating AD-like symptoms such as increases of trans-epidermal water loss, skin thickening and serum IgE as well as histologic inflammation assessment. Collectively, our results propose CAPE as a promising candidate for a novel topical drug for skin inflammatory diseases.

Topics: Acute Disease; Animals; Caffeic Acids; Cell Line; Chronic Disease; Cyclooxygenase 2; Dermatitis, Atopic; Disease Models, Animal; Humans; Immunoglobulin E; Inflammation; Keratinocytes; Mice; Mice, Inbred Strains; NF-kappa B; Oxazolone; Phenylethyl Alcohol; Propolis; Skin; Tetradecanoylphorbol Acetate; Tumor Necrosis Factor-alpha; Water Loss, Insensible

Bcl-3 is an atypical member of the IκB family. Bcl-3 functions as a cofactor of p50/NF-κB1 or p52/NF-κB2 homodimers in nuclei, where it modulates NF-κB-regulated transcription in a context-dependent way. Bcl-3 has tumorigenic potential, is critical in host defense of pathogens, and has been reported to ameliorate or exacerbate inflammation, depending on disease model. However, cell-specific functions of Bcl-3 remain largely unknown. Here, we explored the role of Bcl-3 in a contact hypersensitivity (CHS) mouse model, which depends on the interplay between keratinocytes and immune cells. Bcl-3-deficient mice exhibited an exacerbated and prolonged CHS response to oxazolone. Increased inflammation correlated with higher production of chemokines CXCL2, CXCL9, and CXCL10, and consequently increased recruitment of neutrophils and CD8(+) T cells. BM chimera experiments indicated that the ability of Bcl-3 to reduce the CHS response depended on Bcl-3 activity in radioresistant cells. Specific ablation of Bcl-3 in keratinocytes resulted in increased production of CXCL9 and CXCL10 and sustained recruitment of specifically CD8(+) T cells. These findings identify Bcl-3 as a critical player during the later stage of the CHS reaction to limit inflammation via actions in radioresistant cells, including keratinocytes.

Topics: Animals; B-Cell Lymphoma 3 Protein; CD8-Positive T-Lymphocytes; Chemokine CXCL10; Chemokine CXCL2; Chemokine CXCL9; Dermatitis, Allergic Contact; Inflammation; Inflammation Mediators; Keratinocytes; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; NF-kappa B p50 Subunit; NF-kappa B p52 Subunit; Oxazolone; Proto-Oncogene Proteins; Radiation Tolerance; Transcription Factors; Transcription, Genetic

Leukotriene B4 (LTB4 ) is a lipid mediator that is rapidly generated in inflammatory sites, and its functional receptor, BLT1, is mostly expressed on immune cells. Contact dermatitis is a common inflammatory skin disease characterized by skin oedema and abundant inflammatory infiltrates, primarily including neutrophils and CD8(+) T cells. The role of the LTB4 -BLT1 axis in contact dermatitis remains largely unknown. In this study, we found up-regulated gene expression of 5-lipoxygenase and leukotriene A4 hydrolase, two critical enzymes for LTB4 synthesis, BLT1 and elevated LTB4 levels in skin lesions of oxazolone (OXA)-induced contact dermatitis. BLT1 deficiency or blockade of LTB4 and BLT1 by the antagonists, bestatin and U-75302, respectively, in the elicitation phase caused significant decreases in ear swelling and skin-infiltrating neutrophils and CD8(+) T cells, which was accompanied by significantly reduced skin expression of CXCL1, CXCL2, interferon-γ and interleukin-1β. Furthermore, neutrophil depletion during the elicitation phase of OXA-induced contact dermatitis also caused significant decreases in ear swelling and CD8(+) T-cell infiltration accompanied by significantly decreased LTB4 synthesis and gene expression of CXCL2, interferon-γ and interleukin-1β. Importantly, subcutaneous injection of exogenous LTB4 restored the skin infiltration of CD8(+) T cells in neutrophil-depleted mice following OXA challenge. Collectively, our results demonstrate that the LTB4 -BLT1 axis contributes to OXA-induced contact dermatitis by mediating skin recruitment of neutrophils, which are a major source of LTB4 that sequentially direct CD8(+) T-cell homing to OXA-challenged skin. Hence, LTB4 and BLT1 could be potential therapeutic targets for the treatment of contact dermatitis.

Topics: Animals; Arachidonate 5-Lipoxygenase; CD8-Positive T-Lymphocytes; Chemokine CXCL1; Chemokine CXCL2; Dermatitis, Contact; Epoxide Hydrolases; Fatty Alcohols; Female; Glycols; Inflammation; Interferon-gamma; Interleukin-1beta; Leucine; Leukotriene B4; Mice; Mice, Inbred C57BL; Mice, Knockout; Neutrophils; Oxazolone; Receptors, Leukotriene B4; Skin

Topical pseudoceramides are successfully used in skin barrier repair therapy for atopic dermatitis (AD) and demonstrated to reduce the adverse effects of topical glucocorticoids (GC). However, the molecular mechanisms involved are not fully understood. We investigated whether PC-9S (myristoyl/palmitoyloxostearamide/arachamide MEA, Neopharm, Daejeon, Korea), one of the synthetic pseudoceramides, could stimulate peroxisome proliferator-activated receptor (PPAR)α expression in a hapten [oxazolone (oxa)]-induced AD murine model (oxa-AD mice) and subsequently improved permeability barrier, reduced inflammation, and increased antimicrobial peptides (AMPs) expression. Normal hairless mice and oxa-AD mice were topically treated twice daily with either PC-9S-containing physiologic lipid mixture (PLM), vehicle (PLM), or PPARα agonist for 4 days. Topical PC-9S significantly increased PPARα expression in mouse epidermis in vivo and in oxa-AD mice skin comparable with PPARα agonist. Topical PC-9S-containing PLM significantly reduced basal trans-epidermal water loss (TEWL), surface pH, and mast cell infiltrates and prevented the decline of AMPs expression in oxa-AD mice, which were abrogated by PPARα antagonist. Then, oxa-AD mice were treated with super-potent topical GC twice daily for 4 days with or without PC-9S co-applications. Co-treatment with PC-9S-containing PLM suppressed GC-induced increase in basal TEWL, epidermal thinning, reduced loricrin expression, and impaired barrier recovery and these effects were attenuated by PPARα antagonist. Collectively, our findings suggest that pseudoceramide PC-9S-induced stimulation of PPARα expression provides a new mechanism by which pseudoceramides show anti-inflammatory property, improve the permeability and antimicrobial barrier function, and prevent the negative effects of topical GC.

Topics: Administration, Topical; Animals; Anti-Inflammatory Agents; Arachidonic Acids; Ceramides; Dermatitis, Atopic; Female; Glucocorticoids; Inflammation; Membrane Proteins; Mice; Mice, Hairless; Oxazolone; PPAR alpha; Skin; Stearic Acids; Tight Junctions

The activation and infiltration of polymorphonuclear neutrophils (PMN) are critical key steps in inflammation. PMN-mediated inflammation is limited by anti-inflammatory and pro-resolving mechanisms, including specialized pro-resolving lipid mediators (SPM). We examined the effects of 15-epi-LXA4 on inflammation and the biosynthesis of pro-inflammatory mediators, such as prostaglandins, leukotriene B4 and various hydroxyeicosatetraenoic acids and SPM, in an oxazolone (OXA)-induced hypersensitivity model for dermal inflammation. 15-epi-LXA4 (100 μM, 5 μL subcutaneously injected) significantly (P < 0.05) reduced inflammation in skin, 24 hours after the OXA challenge, as compared to skin treated with vehicle. No significant influence on the biosynthesis of prostaglandins or leukotriene B4 was observed, whereas the level of 15S-hydroxy-eicosatetraenoic acid was significantly (P < 0.05) lower in the skin areas treated with 15-epi-LXA4. In spite of the use of a fully validated analytical procedure, no SPM were detected in the biological samples. To investigate the reason for the lack of analytical signal, we tried to mimic the production of SPM (lipoxins, resolvins, maresin and protectin) by injecting them subcutaneously into the skin of mice and studying the in vivo availability and distribution of the compounds. All analytes showed very little lateral distribution in skin tissue and their levels were markedly decreased (> 95%) 2 hours after injection. However, docosahexaenoic acid derivatives were biologically more stable than SPM derived from arachidonic acid or eicosapentaenoic acid.

Topics: Animals; Calibration; Chromatography, Liquid; Cyclooxygenase 2; Dermis; Hydroxyeicosatetraenoic Acids; Inflammation; Inflammation Mediators; Injections, Subcutaneous; Leukotriene B4; Lipids; Lipoxins; Luminescent Measurements; Mass Spectrometry; Mice, Inbred C57BL; Oxazolone; Peptidylprolyl Isomerase; Reproducibility of Results; Ribosomal Proteins

The mechanisms by which mucosal homeostasis is maintained are of central importance to inflammatory bowel disease. Critical to these processes is the intestinal epithelial cell (IEC), which regulates immune responses at the interface between the commensal microbiota and the host. CD1d presents self and microbial lipid antigens to natural killer T (NKT) cells, which are involved in the pathogenesis of colitis in animal models and human inflammatory bowel disease. As CD1d crosslinking on model IECs results in the production of the important regulatory cytokine interleukin (IL)-10 (ref. 9), decreased epithelial CD1d expression--as observed in inflammatory bowel disease--may contribute substantially to intestinal inflammation. Here we show in mice that whereas bone-marrow-derived CD1d signals contribute to NKT-cell-mediated intestinal inflammation, engagement of epithelial CD1d elicits protective effects through the activation of STAT3 and STAT3-dependent transcription of IL-10, heat shock protein 110 (HSP110; also known as HSP105), and CD1d itself. All of these epithelial elements are critically involved in controlling CD1d-mediated intestinal inflammation. This is demonstrated by severe NKT-cell-mediated colitis upon IEC-specific deletion of IL-10, CD1d, and its critical regulator microsomal triglyceride transfer protein (MTP), as well as deletion of HSP110 in the radioresistant compartment. Our studies thus uncover a novel pathway of IEC-dependent regulation of mucosal homeostasis and highlight a critical role of IL-10 in the intestinal epithelium, with broad implications for diseases such as inflammatory bowel disease.

Topics: Animals; Antigens, CD1d; Carrier Proteins; Colitis; Disease Models, Animal; Epithelial Cells; Female; HSP110 Heat-Shock Proteins; Humans; Immunity, Mucosal; Inflammation; Inflammatory Bowel Diseases; Interleukin-10; Intestinal Mucosa; Male; Mice; Natural Killer T-Cells; Oxazolone; STAT3 Transcription Factor

Allergic contact dermatitis is a common skin disease associated with inflammation and persistent pruritus. Transient receptor potential (TRP) ion channels in skin-innervating sensory neurons mediate acute inflammatory and pruritic responses following exogenous stimulation and may contribute to allergic responses. Genetic ablation or pharmacological inhibition of TRPA1, but not TRPV1, inhibited skin edema, keratinocyte hyperplasia, nerve growth, leukocyte infiltration, and antihistamine-resistant scratching behavior in mice exposed to the haptens, oxazolone and urushiol, the contact allergen of poison ivy. Hapten-challenged skin of TRPA1-deficient mice contained diminished levels of inflammatory cytokines, nerve growth factor, and endogenous pruritogens, such as substance P (SP) and serotonin. TRPA1-deficient sensory neurons were defective in SP signaling, and SP-induced scratching behavior was abolished in Trpa1(-/-) mice. SP receptor antagonists, such as aprepitant inhibited both hapten-induced cutaneous inflammation and scratching behavior. These findings support a central role for TRPA1 and SP in the integration of immune and neuronal mechanisms leading to chronic inflammatory responses and pruritus associated with contact dermatitis.

Topics: Animals; Dermatitis, Allergic Contact; Female; Inflammation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Oxazolone; Transient Receptor Potential Channels; TRPA1 Cation Channel

Serious infections of the head and neck cause lymphedema that can lead to airway compromise and oropharyngeal obstruction. Lymphangiogenesis occurs in the head and neck during infection and after immunization. The goal of this project was to develop tools to image lymphatic vessels in living animals and to be able to isolate individual lymphatic endothelial cells in order to quantify changes in single cells caused by inflammation.. The ProxTom transgenic red-fluorescent reporter mouse was developed specifically for the purpose of imaging lymphatic vessels in vivo. Prox1 is a transcription factor that is necessary for lymphangiogenesis in development and for the maintenance of lymphatics in adulthood. Mice were immunized and their lymphatic vessels in lymph nodes were imaged in vivo. Individual lymphatic endothelial cells were isolated by means of their fluorescence.. The ProxTom transgene has the red-fluorescent reporter td-Tomato under the control of Prox1 regulatory elements. tdTomato was faithfully expressed in lymphatic vessels coincident with endogenous Prox1 expression. We show lymphangiogenesis in vivo after immunization and demonstrate a method for the isolation of lymphatic endothelial cells by their tdTomato red-fluorescence.. The faithful expression of the red-fluorescent reporter in the lymphatic vessels of ProxTom means that these mice have proven utility for in vivo study of lymphatic vessels in the immune response. ProxTom has been made available for distribution from the Jackson Laboratory: http://jaxmice.jax.org/strain/018128.html .

Topics: Animals; Endothelial Cells; Flow Cytometry; Head; Homeodomain Proteins; Immunization; Inflammation; Luminescent Proteins; Lymph Nodes; Lymphangiogenesis; Lymphatic Vessels; Mice; Mice, Inbred C57BL; Mice, Transgenic; Microscopy, Fluorescence; Neck; Oxazolone; Promoter Regions, Genetic; Red Fluorescent Protein; Tumor Suppressor Proteins

Annexin A1 (AnxA1) is recognized as an endogenous anti-inflammatory molecule. However, its effects on the adaptive immune response and, in particular, on T cells remain unclear. In this study, we investigated the actions of AnxA1 in three distinct models of T cell-mediated inflammation. In contact hypersensitivity, collagen-induced arthritis, and inflammation induced by OT-II TCR transgenic T cells responding to OVA, AnxA1 deficiency significantly increased Ag-induced T cell proliferation and the resultant level of inflammation. In the contact hypersensitivity model, this was associated with increased adhesion of CD4(+) T cells, CD8(+) T cells, and neutrophils in the dermal microvasculature, as well as increased T cell expression of RORγt and IL-17A. In collagen-induced arthritis, deficiency of endogenous AnxA1 increased susceptibility to arthritis and Ag-specific T cell activation. Deficiency of AnxA1 also increased OVA-induced cutaneous delayed-type hypersensitivity and IFN-γ and IL-17 release. Transfer experiments using CD4(+) T cells from AnxA1(-/-) mice demonstrated that the absence of AnxA1 solely in T cells resulted in increased inflammatory responses in wild-type recipients. Similarly, experiments using AnxA1(-/-) OT-II CD4(+) T cells demonstrated that the absence of AnxA1 in T cells was sufficient to induce increased Ag-specific CD4(+) T cell proliferation in vivo, augment T cell production of IFN-γ, IL-17, TNF, and IL-6, and increase Akt, ERK, and p38 activation. Together, these findings indicate that T cell-expressed AnxA1 functions to attenuate T cell-driven inflammatory responses via T cell-intrinsic effects on intracellular signaling, proliferation, and Th1/Th17 cytokine release.

Topics: Animals; Annexin A1; Arthritis, Experimental; CD4-Positive T-Lymphocytes; CD8-Positive T-Lymphocytes; Cell Adhesion; Dermatitis, Allergic Contact; Enzyme Activation; Gene Expression Regulation; Hypersensitivity, Delayed; Inflammation; Interleukin-17; Lymphocyte Activation; Lymphokines; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Neutrophils; Nuclear Receptor Subfamily 1, Group F, Member 3; Ovalbumin; Oxazolone; Peptide Fragments; Signal Transduction; Specific Pathogen-Free Organisms

The effect of a single time exposure of SLS to the buccal mucosa of mice was compared to one application of the hapten OXA (oxazolone), evaluated by routine histology, immunohistochemistry and ELISA quantifications of cytokines. The SLS concentrations (2%, 4% and 8%) resulted in epithelial surface necrosis at 1-6 h, after 2-6 h accumulation of intra-epithelial neutrophils and at 24 h the main inflammatory cells were mononuclear. Increased concentrations of SLS gave more severe damage. CD4(+) T cells were found at 6 h and increased slightly up to 24 h and were most frequently seen at the lowest SLS dose. The CD8(+) T cells were kept at a low number during the whole 24 h observation period, but increased proportionally to the CD4(+) T cells. One application of 1% OXA did not raise the number of cells of either phenotype (2-24 h). Neither IL-2 nor IFN-γ demonstrated increased levels during the week of observation at any concentration of SLS, contrary to one application of OXA which caused increased IL-2 levels both at the local application site and in the regional and distant lymph nodes. Regardless of SLS concentration, a minor increase in regional lymph node weight was observed 8-12 h after substance application, quickly to subside whilst one OXA application gave a maximal weight increase at 48-72 h. We conclude that oral mucosa irritant SLS reactions gave early surface necrosis and neutrophil infiltrations and later mononuclear cell infiltrations dominated by CD4(+) T cells. The cytokines IL-2 and IFN-γ and lymphocyte proliferation in the regional lymph nodes was not observed after SLS application, contrary to hapten application.

Topics: Animals; Enzyme-Linked Immunosorbent Assay; Immunoenzyme Techniques; Inflammation; Interleukin-2; Mice; Mouth Mucosa; Necrosis; Oxazolone; Sodium Dodecyl Sulfate; T-Lymphocytes; Tumor Necrosis Factor-alpha

To investigate the role of cannabinoid receptor-2 (CB2) in allergic inflammation in CB2 knockout (CB2-KO) mice.. The swelling reaction of the pinna to various stimuli was compared between CB2-KO and wild-type (WT) mice in terms of edema and acanthosis.. Ear swelling induced by repeated application of 2,4-dinitrofluorobenzene in CB2-KO mice was significantly decreased compared with that in WT mice. In an ovalbumin model, pinna edema was significantly suppressed in CB2-KO mice in comparison with that in WT mice. The contribution of CB2 to edema was investigated in a more extreme dermatitis model using oxazolone. Delayed-type hypersensitivity reactions in this model were also suppressed in CB2-KO mice. In each of these three different allergic dermatitis models, there was a significant decrease in edema and acanthosis in CB2-KO mice compared with WT mice.. These results clearly demonstrate that CB2 and its endogenous ligands participate not only in the acute, edematous phase of allergic dermatitis, but also in the chronic irreversible acanthosis reaction.

Topics: Animals; Dermatitis; Disease Models, Animal; Edema; Hypersensitivity, Delayed; Inflammation; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Ovalbumin; Oxazolone; Receptor, Cannabinoid, CB2

Interleukin-25 (IL-25) is a potent activator of type-2 immune responses. Mucosal inflammation in ulcerative colitis is driven by type-2 cytokines. We have previously shown that a neutralizing anti-IL-25 antibody abrogated airways hyperreactivity in an experimental model of lung allergy. Therefore, we asked whether blocking IL-25 via neutralizing antibodies against the ligand or its receptor IL-17BR could protect against inflammation in an oxazolone-induced mouse model of colitis.. Neutralizing antibodies to IL-25 or IL-17BR were administered to mice with oxazolone-induced colitis, a model of ulcerative colitis. The disease onset was evaluated by weight loss and degree of colon ulceration. Also, lamina propria and mesenteric lymph node (MLN) infiltrates were assessed for mucosal inflammation and cultured in vitro to determine cytokine production.. We found that in oxazolone colitis IL-25 production derives from intestinal epithelial cells and that IL-17BR(+) IL-13-producing natural killer T (NKT) cells and nuocytes drive the intestinal inflammation. Blocking IL-25 signalling considerably improved the clinical aspects of the disease, including weight loss and colon ulceration, and resulted in fewer nuocytes and NKT cells infiltrating the mucosa. The improved pathology correlated with a decrease in IL-13 production by lamina propria cells, a decrease in the production of other type-2 cytokines by MLN cells, and a decrease in blood eosinophilia and IgE.. IL-25 plays a pro-inflammatory role in the oxazolone colitis model, and neutralizing antibodies to IL-25 or IL-17BR can slow the ongoing inflammation in this disease. Because this model mimics aspects of human ulcerative colitis, these antibodies may represent potential therapeutics for reducing gut inflammation in patients.

Topics: Animals; Antibodies, Neutralizing; Colitis, Ulcerative; Disease Models, Animal; Female; Inflammation; Interleukin-13; Interleukin-17; Intestinal Mucosa; Mice; Mice, Inbred BALB C; Natural Killer T-Cells; Oxazolone; Receptors, Interleukin-17; Signal Transduction

Irritant contact dermatitis (ICD) is an inflammatory skin disease triggered by exposure to a chemical that is toxic or irritating to the skin. A major characteristic of chronic ICD is an inflammatory dry-skin condition with associated itching. Although glucosylceramide (GlcCer) is known to improve the skin barrier function, its mechanism of action is unknown.. Using a mouse model of oxazolone-induced chronic ICD, this study investigated the effects of oral administration of GlcCer on inflammatory dry skin.. Chronic ICD was induced by repeated application of oxazolone in mice. GlcCer was orally administered once daily throughout the elicitation phase. The beneficial efficacy of GlcCer on cutaneous inflammation was evaluated by assessing ear thickness, lymph node weight, histological findings, and mRNA expression of pro-inflammatory cytokines such as IL-1β and IL-6. Additionally, parameters of the itch-associated response, including scratching behavior, water content of the skin, and aquaporin-3 levels in the lesional ear, were measured.. Oral GlcCer administration significantly suppressed mRNA expression of the pro-inflammatory cytokines IL-1β and IL-6. GlcCer also suppressed ear swelling, lymph node weight gains, and infiltration of leukocytes and mast cells in ICD mice. In oxazolone-induced ICD mice, GlcCer significantly inhibited irritant-related scratching behavior and dehydration of the stratum corneum, and decreased aquaporin-3 expression.. Our results indicate that GlcCer suppressed inflammation not only by inhibiting cytokine production but also by repairing the skin barrier function, suggesting a potential beneficial role for GlcCer in the improvement of chronic ICD.

Topics: Administration, Oral; Animals; Aquaporin 3; Dermatitis, Contact; Dermatitis, Irritant; Ear; Glucosylceramides; Inflammation; Interleukin-1beta; Interleukin-6; Lymph Nodes; Male; Mice; Mice, Inbred BALB C; Models, Biological; Oxazolone; RNA, Messenger; Skin

Secondary lymphedema in humans is a common consequence of axillary lymph node dissection (ALND) to treat breast cancer. Remarkably, secondary lymphedema generally first appears following a delay of over a year and can be triggered suddenly by an inflammatory insult. However, it remains unclear why the apparently functional lymphatic system is unable to accommodate an inflammatory trigger. To provide mechanistic insight into the delayed and rapid secondary lymphedema initiation, we compared the ability of the ALND-recovered rat foreleg lymphatic system to prevent edema during an inflammatory challenge with that of the uninjured lymphatic system. At 73 days postsurgery, the forelegs of ALND(-)- and ALND(+)-sensitized rats were exposed to the proinflammatory agent oxazolone, which was found to reduce fluid drainage and increase skin thickness in both ALND(-) and ALND(+) forelegs (P < 0.05). However, drainage in the ALND-recovered forelegs was more severely impaired than ALND(-) forelegs, as visualized by indocyanine green lymphography and quantified by interstitial transport of fluid marker (P < 0.05). Although both ALND(+) and ALND(-) forelegs experienced significant inflammation-induced edema with the oxazolone exposure (P < 0.05), the peak tissue swelling in the ALND(+) group was significantly greater than that of the ALND(-) forelegs (arm area peaked at ∼13.4 vs. ∼5.7% swelling, respectively, P < 0.005; wrist diameter peaked at 9.7 vs. 2.2% swelling, respectively, P < 0.005). The findings demonstrate that outward recovery from ALND in the rat foreleg masks an ensuing chronic and latent lymphatic insufficiency, which reduces the ability of the foreleg lymphatic system to prevent edema during an acute inflammatory process.

Topics: Acute Disease; Animals; Female; Forelimb; Inflammation; Lymph Node Excision; Lymphatic System; Lymphedema; Lymphography; Models, Animal; Oxazolone; Rats; Rats, Sprague-Dawley

The prevention and treatment of diabetic complications are considered to be the most important for the general care of diabetic patients. We have been conducting pre-clinical animal experiments related to diabetes using kangen-karyu, a Chinese prescription, to examine its therapeutic potential. In the present study, we further studied the anti-diabetic mechanism of kangen-karyu, especially on the regulation of hyperglycemia-induced hepatic oxidative stress and inflammation in db/db mice. Kangen-karyu (100 or 200 mg/kg body weight/day, per os (p.o.) was administered every day for 18 weeks to db/db mice, and its effect was compared with vehicle-treated db/db and m/m mice. The administration of kangen-karyu decreased the elevated serum and hepatic glucose concentration in db/db mice. The elevated expressions of p22(phox) and Nox-4 proteins (reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits) were significantly decreased after kangen-karyu treatments. The oxidative stress-related markers in hepatic tissue (reactive oxygen species, reduced glutathione/oxidized glutathione ratio, and thiobarbituric acid-reactive substance) were also significantly ameliorated by the kangen-karyu treatments. The db/db mice exhibited the up-regulation of nuclear factor-κBp65, cyclooxygenase-2, and inducible nitric oxide synthase levels in the liver; however, kangen-karyu treatment significantly reduced those expressions. Taking these into consideration, our findings support the therapeutic evidence for kangen-karyu ameliorating the development of diabetic hepatic damages via regulating oxidative stress and inflammation.

Topics: Animals; Anti-Inflammatory Agents; Antioxidants; Biomarkers; Cyclooxygenase Inhibitors; Diabetes Complications; Drugs, Chinese Herbal; Glucose; Hyperglycemia; Inflammation; Inflammation Mediators; Liver; Liver Diseases; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; NADPH Oxidase 4; NADPH Oxidases; NF-kappa B; Nitric Oxide Synthase Type II; Oxazoles; Oxidative Stress; Phytotherapy

Patients with prolonged ulcerative colitis (UC) frequently develop colorectal adenocarcinoma for reasons that are not fully clear. To analyze inflammation-associated colonic tumorigenesis, we developed a chronic form of oxazolone-induced colitis in mice that, similar to UC, was distinguished by the presence of IL-13-producing NKT cells. In this model, the induction of tumors using azoxymethane was accompanied by the coappearance of F4/80+CD11b(high)Gr1(low) M2 macrophages, cells that undergo polarization by IL-13 and are absent in tumors that lack high level IL-13 production. Importantly, this subset of macrophages was a source of tumor-promoting factors, including IL-6. Similar to dextran sodium sulfate-induced colitis, F4/80+CD11b(high)Gr1(intermediate) macrophages were present in the mouse model of chronic oxazolone-induced colitis and may influence tumor development through production of TGF-β1, a cytokine that inhibits tumor immunosurveillance. Finally, while robust chronic oxazolone-induced colitis developed in myeloid differentiation primary response gene 88-deficient (Myd88-/-) mice, these mice did not support tumor development. The inhibition of tumor development in Myd88-/- mice correlated with cessation of IL-6 and TGF-β1 production by M2 and F4/80+CD11b(high)Gr1(intermediate) macrophages, respectively, and was reversed by exogenous IL-6. These data show that an UC-like inflammation may facilitate tumor development by providing a milieu favoring development of MyD88-dependent tumor-supporting macrophages.

Topics: Animals; Antigens, Ly; CD11b Antigen; CD8-Positive T-Lymphocytes; Colitis; Colitis, Ulcerative; Inflammation; Interleukin-13; Interleukin-6; Macrophages; Mice; Mice, Inbred BALB C; Mice, Transgenic; Myeloid Differentiation Factor 88; Oxazolone; Signal Transduction; Transforming Growth Factor beta1

Skin protects the body from the environment and is an important component of the innate and adaptive immune systems. Atopic dermatitis and contact dermatitis are among the most frequent inflammatory skin diseases and are both determined by multigenic predisposition, environmental factors, and aberrant immune response. Peptidoglycan Recognition Proteins (Pglyrps) are expressed in the skin and we report here that they modulate sensitivity to experimentally-induced atopic dermatitis and contact dermatitis. Pglyrp3(-/-) and Pglyrp4(-/-) mice (but not Pglyrp2(-/-) mice) develop more severe oxazolone-induced atopic dermatitis than wild type (WT) mice. The common mechanism underlying this increased sensitivity of Pglyrp3(-/-) and Pglyrp4(-/-) mice to atopic dermatitis is reduced recruitment of Treg cells to the skin and enhanced production and activation Th17 cells in Pglyrp3(-/-) and Pglyrp4(-/-) mice, which results in more severe inflammation and keratinocyte proliferation. This mechanism is supported by decreased inflammation in Pglyrp3(-/-) mice following in vivo induction of Treg cells by vitamin D or after neutralization of IL-17. By contrast, Pglyrp1(-/-) mice develop less severe oxazolone-induced atopic dermatitis and also oxazolone-induced contact dermatitis than WT mice. Thus, Pglyrp3 and Pglyrp4 limit over-activation of Th17 cells by promoting accumulation of Treg cells at the site of chronic inflammation, which protects the skin from exaggerated inflammatory response to cell activators and allergens, whereas Pglyrp1 has an opposite pro-inflammatory effect in the skin.

Topics: Allergens; Animals; Blotting, Western; Carrier Proteins; Dermatitis, Atopic; Disease Models, Animal; Enzyme-Linked Immunosorbent Assay; Female; Flow Cytometry; Immune System; Inflammation; Interleukin-17; Male; Mice; Mice, Inbred BALB C; Mice, Knockout; Oxazolone; Real-Time Polymerase Chain Reaction; RNA, Messenger; T-Lymphocytes, Regulatory

To trigger an effective immune response, antigen and antigen-presenting cells travel to the lymph nodes via collecting lymphatic vessels. However, our understanding of the regulation of collecting lymphatic vessel function and lymph transport is limited. To dissect the molecular control of lymphatic function, we developed a unique mouse model that allows intravital imaging of autonomous lymphatic vessel contraction. Using this method, we demonstrated that endothelial nitric oxide synthase (eNOS) in lymphatic endothelial cells is required for robust lymphatic contractions under physiological conditions. By contrast, under inflammatory conditions, inducible NOS (iNOS)-expressing CD11b(+)Gr-1(+) cells attenuate lymphatic contraction. This inhibition of lymphatic contraction was associated with a reduction in the response to antigen in a model of immune-induced multiple sclerosis. These results suggest the suppression of lymphatic function by the CD11b(+)Gr-1(+) cells as a potential mechanism of self-protection from autoreactive responses during on-going inflammation. The central role for nitric oxide also suggests that other diseases such as cancer and infection may also mediate lymphatic contraction and thus immune response. Our unique method allows the study of lymphatic function and its molecular regulation during inflammation, lymphedema, and lymphatic metastasis.

Topics: Animals; Bone Marrow Cells; CD11b Antigen; Immune System; Immunosuppression Therapy; Inflammation; Kinetics; Lymphatic Metastasis; Lymphatic System; Lymphatic Vessels; Mice; Mice, Inbred C57BL; Microscopy; Nitric Oxide Synthase Type III; Oxazolone; Skin

Glucocorticoids (GC) are highly potent anti-inflammatory agents frequently administered in clinical medicine. However, even for the most potent GC dexamethasone, only modest effects have been observed in several murine studies. Here we demonstrate that intraperitoneal administration of dexamethasone displays no anti-inflammatory activity in two different mouse models. Low doses of topically applied dexamethasone entirely prevented ear swelling in a contact hypersensitivity model in BALB/c mice, while intraperitoneally injected dexamethasone had no effect on disease progression. Moreover, subcutaneously administered dexamethasone completely inhibited lipopolysaccharide (LPS)-mediated lethality in C57BL/6 mice. In contrast, even ultra-high doses of intraperitoneally injected dexamethasone could not prevent endotoxin-induced death. In conclusion, these results demonstrate that intraperitoneal application of dexamethasone is ineffective in these models of inflammation, which has broad implications for mouse models evaluating the in vivo efficiency of GCs.

Topics: Animals; Anti-Inflammatory Agents; Dermatitis, Contact; Dexamethasone; Disease Models, Animal; Disease Progression; Drug Administration Routes; Endotoxemia; Immunity; Inflammation; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Oxazolone

It is widely accepted that cationic antimicrobial peptides possess potent microbicidal properties. Recent studies show that in addition to their antimicrobial action, these peptides can exhibit anti-inflammatory activity. The purpose of this chapter is to describe in vivo ear inflammation models that can be used for evaluating the anti-inflammatory activity of antimicrobial peptides. The models are based on different mechanisms of inflammation development and include irritant dermatitis (a model induced by a single application of 12-o-tetradecanoylphorbol acetate [TPA]) and allergic dermatitis, or delayed type hypersensitivity reaction (a model induced by repetitive application of oxazolone).

Topics: Animals; Anti-Inflammatory Agents; Antimicrobial Cationic Peptides; Dermatitis, Allergic Contact; Dermatitis, Irritant; Disease Models, Animal; Female; Humans; Inflammation; Mice; Oxazolone; Tetradecanoylphorbol Acetate

We have shown that somatostatin released from activated capsaicin-sensitive nociceptive nerve endings during inflammatory processes elicits systemic anti-inflammatory and analgesic effects. With the help of somatostatin receptor subtype 4 gene-deleted mice (sst(4)(-/-)), we provide here several lines of evidence that this receptor has a protective role in a variety of inflammatory disease models; several symptoms are more severe in the sst(4) knockout animals than in their wild-type counterparts. Acute carrageenan-induced paw edema and mechanical hyperalgesia, inflammatory pain in the early phase of adjuvant-evoked chronic arthritis, and oxazolone-induced delayed-type hypersensitivity reaction in the skin are much greater in mice lacking the sst(4) receptor. Airway inflammation and consequent bronchial hyperreactivity elicited by intranasal lipopolysaccharide administration are also markedly enhanced in sst(4) knockouts, including increased perivascular/peribronchial edema, neutrophil/macrophage infiltration, mucus-producing goblet cell hyperplasia, myeloperoxidase activity, and IL-1beta, TNF-alpha, and IFN-gamma expression in the inflamed lung. It is concluded that during these inflammatory conditions the released somatostatin has pronounced counterregulatory effects through sst(4) receptor activation. Thus, this receptor is a promising novel target for developing anti-inflammatory, analgesic, and anti-asthmatic drugs.

Topics: Animals; Bronchial Hyperreactivity; Dermatitis, Allergic Contact; Female; Hyperalgesia; Inflammation; Lipopolysaccharides; Male; Mice; Mice, Inbred C57BL; Mice, Knockout; Oxazolone; Receptors, Somatostatin

Oatmeal has been used for centuries as a soothing agent to relieve itch and irritation associated with various xerotic dermatoses; however few studies have sought to identify the active phytochemical(s) in oat that mediate this anti-inflammatory activity. Avenanthramides are phenolic compounds present in oats at approximately 300 parts per million (ppm) and have been reported to exhibit anti-oxidant activity in various cell-types. In the current study we investigated whether these compounds exert anti-inflammatory activity in the skin. We found that avenanthramides at concentrations as low as 1 parts per billion inhibited the degradation of inhibitor of nuclear factor kappa B-alpha (IkappaB-alpha) in keratinocytes which correlated with decreased phosphorylation of p65 subunit of nuclear factor kappa B (NF-kappaB). Furthermore, cells treated with avenanthramides showed a significant inhibition of tumor necrosis factor-alpha (TNF-alpha) induced NF-kappaB luciferase activity and subsequent reduction of interleukin-8 (IL-8) release. Additionally, topical application of 1-3 ppm avenanthramides mitigated inflammation in murine models of contact hypersensitivity and neurogenic inflammation and reduced pruritogen-induced scratching in a murine itch model. Taken together these results demonstrate that avenanthramides are potent anti-inflammatory agents that appear to mediate the anti-irritant effects of oats.

Topics: Animals; Avena; Cells, Cultured; Dermatitis, Contact; Disease Models, Animal; Diterpenes; Flavonoids; Humans; Inflammation; Interleukin-8; Keratinocytes; Mice; Mice, Inbred ICR; NF-kappa B; ortho-Aminobenzoates; Oxazolone; Phenols; Phytotherapy; Polyphenols; Pruritus; Signal Transduction

The hyaluronan receptor LYVE-1 is expressed abundantly on the surfaces of lymphatic vessels and lymph node sinus endothelial cells from early development, where it has been suggested to function both in cell adhesion/transmigration and as a scavenger for hyaluronan turnover. To investigate the physiological role(s) of LYVE-1, we generated mice in which the gene for the receptor was inactivated by replacement with a beta-galactosidase reporter. LYVE-1(-/-) mice displayed an apparently normal phenotype, with no obvious alteration in lymphatic vessel ultrastructure or function and no apparent change in secondary lymphoid tissue structure or cellularity. In addition, the levels of hyaluronan in tissue and blood were unchanged. LYVE-1(-/-) mice also displayed normal trafficking of cutaneous CD11c(+) dendritic cells to draining lymph nodes via afferent lymphatics and normal resolution of oxazolone-induced skin inflammation. Finally, LYVE-1(-/-) mice supported normal growth of transplanted B16F10 melanomas and Lewis lung carcinomas. These results indicate that LYVE-1 is not obligatory for normal lymphatic development and function and suggest either the existence of compensatory receptors or a role more specific than that previously envisaged.

Topics: Animals; beta-Galactosidase; Carcinoma, Lewis Lung; CD11c Antigen; Cell Movement; Dendritic Cells; Dermatitis, Contact; Glycoproteins; Hyaluronic Acid; Inflammation; Lymph Nodes; Lymphatic Vessels; Melanoma; Membrane Transport Proteins; Mice; Mice, Knockout; Neoplasm Transplantation; Oxazolone

CD47 on the surface of T cells was shown in vitro to mediate either T cell activation or, in the presence of high amounts of thrombospondin (TSP), T cell apoptosis. We report here that CD47-deficient mice, as well as TSP-1 or TSP-2-deficient mice, sustain oxazolone-induced inflammation for more than four days, whereas wild-type mice reduce the inflammation within 48 h. We observe that prolonged inflammation in CD47-, TSP-1-, or TSP-2-deficient mice is accompanied by a local deficiency of T cell apoptosis. Finally, we show that upon activation normal T cells increase the expression of the proapoptotic Bcl-2 family member BNIP3 (Bcl-2/adenovirus E1B 19-kDa interacting protein) and undergo CD47-mediated apoptosis. This finding is consistent with our previous demonstration of a physical interaction between BNIP3 and CD47 that inhibits BNIP3 degradation by the proteasome, sensitizing T cells to CD47-induced apoptosis. Overall, these results reveal an important role in vivo for this new CD47/BNIP3 pathway in limiting inflammation by controlling the number of activated T cells.

Topics: Animals; Apoptosis; CD47 Antigen; Dermatitis; Inflammation; Membrane Proteins; Mice; Mice, Mutant Strains; Mitochondrial Proteins; Oxazolone; Proteasome Endopeptidase Complex; T-Lymphocytes; Thrombospondin 1; Thrombospondins

Chronic contact hypersensitivity (CH) models induced by repeated hapten exposure exhibit chronic dermatitis and immunological abnormalities resembling atopic dermatitis. To assess the contribution of endothelial selectins (P- and E-selectins) to cutaneous chronic inflammation, chronic CH responses were assessed in mice lacking P- or E-selectin. Elicitation with oxazolone on the ears of P-selectin(-/-) mice 7 days after the sensitization induced a typical delayed-type hypersensitivity response similar to that found in wild-type mice. By contrast, a significant increase in ear swelling was observed in E-selectin(-/-) mice 36 to 48 hours after first elicitation. E-selectin(-/-) mice showed augmented P-selectin up-regulation, and administration of anti-P-selectin monoclonal antibody significantly inhibited the enhanced ear response, suggesting that the enhanced ear-swelling response in E-selectin(-/-) mice resulted from compensatory increase in P-selectin expression. In the late phase of chronic CH, acceleration of ear swelling was significantly reduced in both E- and P-selectin(-/-) mice relative to wild-type littermates. Thus, the loss of P- or E-selectin suppressed inflammatory responses during the chronic phase of the chronic models, whereas early-phase inflammatory responses were exacerbated by E-selectin blockade. Collectively, P- and E-selectins cooperatively regulate CH response, although their roles may be different depending on the phase of the reaction.

Topics: Adjuvants, Immunologic; Animals; Chronic Disease; Dermatitis, Atopic; Dermatitis, Contact; E-Selectin; Immunohistochemistry; Inflammation; Mice; Mice, Inbred C57BL; Oxazolone; P-Selectin; Reverse Transcriptase Polymerase Chain Reaction

Interleukin (IL)-13 is a major inducer of fibrosis in many chronic infectious and autoimmune diseases. In studies of the mechanisms underlying such induction, we found that IL-13 induces transforming growth factor (TGF)-beta(1) in macrophages through a two-stage process involving, first, the induction of a receptor formerly considered to function only as a decoy receptor, IL-13Ralpha(2). Such induction requires IL-13 (or IL-4) and tumor necrosis factor (TNF)-alpha. Second, it involves IL-13 signaling through IL-13Ralpha(2) to activate an AP-1 variant containing c-jun and Fra-2, which then activates the TGFB1 promoter. In vivo, we found that prevention of IL-13Ralpha(2) expression reduced production of TGF-beta(1) in oxazolone-induced colitis and that prevention of IL-13Ralpha(2) expression, Il13ra2 gene silencing or blockade of IL-13Ralpha(2) signaling led to marked downregulation of TGF-beta(1) production and collagen deposition in bleomycin-induced lung fibrosis. These data suggest that IL-13Ralpha(2) signaling during prolonged inflammation is an important therapeutic target for the prevention of TGF-beta(1)-mediated fibrosis.

Topics: Animals; Bleomycin; Blotting, Western; Cell Lineage; Colitis; Collagen; Cytokines; Down-Regulation; Enzyme-Linked Immunosorbent Assay; Etanercept; Fibrosis; Flow Cytometry; Gene Silencing; Genetic Vectors; Humans; Immunoglobulin G; Inflammation; Interleukin-13; Interleukin-13 Receptor alpha1 Subunit; Luciferases; Lung; Macrophages; Mice; Mice, Inbred C57BL; Monocytes; NF-kappa B; Oxazolone; Promoter Regions, Genetic; Receptors, Interleukin; Receptors, Interleukin-13; Receptors, Tumor Necrosis Factor; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; Signal Transduction; Time Factors; Transcription Factor AP-1; Transforming Growth Factor beta; Transforming Growth Factor beta1; Tumor Necrosis Factor-alpha; Up-Regulation

The possible involvement of 2-arachidonoylglycerol (2-AG), an endogenous ligand for the cannabinoid receptors (CB1 and CB2), in contact dermatitis in mouse ear was investigated. We found that the level of 2-AG was markedly elevated in the ear following a challenge with oxazolone in sensitized mice. Of note, the swelling following the challenge was suppressed by either the administration of SR144528, a CB2 receptor antagonist, immediately after sensitization, or the administration of SR144528 upon the challenge. The effect of AM251, a CB1 receptor antagonist, was marginal in either case. It seems apparent, therefore, that the CB2 receptor and its endogenous ligand 2-AG are closely involved in both the sensitization phase and the elicitation phase of oxazolone-induced contact dermatitis. In line with this, we found that Langerhans cells (MHC class II(+)) contain a substantial amount of CB2 receptor mRNA, whereas keratinocytes (MHC class II(-)) do not. We also obtained evidence that the expression of mRNAs for proinflammatory cytokines following a challenge with oxazolone was markedly suppressed by treatment with SR144528. We next examined whether the CB2 receptor and 2-AG participate in chronic contact dermatitis accompanied by the infiltration of tissues by eosinophils. The amount of 2-AG in mouse ear dramatically increased following repeated challenge with oxazolone. Importantly, treatment with SR144528 attenuated both the recruitment of eosinophils and ear swelling in chronic contact dermatitis induced by repeated challenge with oxazolone. These results strongly suggest that the CB2 receptor and 2-AG play important stimulative roles in the sensitization, elicitation, and exacerbation of allergic inflammation.

Topics: Animals; Arachidonic Acids; Dermatitis, Contact; Ear; Endocannabinoids; Glycerides; Inflammation; Keratinocytes; Langerhans Cells; Ligands; Male; Mice; Mice, Inbred ICR; Oxazolone; Receptor, Cannabinoid, CB2; RNA, Messenger

Human semicarbazide-sensitive amine oxidase (SSAO) or vascular adhesion protein-1 (VAP-1) is a copper-containing amine oxidase (AOC3, EC 1.4.3.6) that has both enzymatic and adhesive function. SSAO catalyzes the oxidative deamination of primary amines, resulting in the formation of the corresponding aldehyde and release of hydrogen peroxide and ammonia. Membrane-bound SSAO is an inflammation-inducible endothelial cell adhesion molecule that mediates the interaction between leukocytes and activated endothelial cells in inflamed vessels. Both the direct adhesive and enzymatic functions seem to be involved in the adhesion cascade. LJP 1207 [N'-(2-phenyl-allyl)-hydrazine hydrochloride] is a potent (human SSAO IC(50) = 17 nM), selective, and orally available SSAO inhibitor that blocks both the enzymatic and adhesion functions of SSAO/VAP-1. In a mouse model of ulcerative colitis, LJP 1207 significantly reduces mortality, loss of body weight, and colonic cytokine levels. Quantitative histopathological assessment of colitis activity in this model showed a highly significant suppression of inflammation, injury, and ulceration scores in the animals treated with the SSAO/VAP-1 inhibitor. LJP 1207 also reduced serum levels of tumor necrosis factor-alpha and interleukin 6 in lipopolysaccharide (LPS)-challenged mice and prolonged survival post-LPS-induced endotoxemia. Therapeutic and prophylactic administration of LJP 1207 in the rat carrageenan footpad model also markedly inhibited swelling and inflammation. Overall, the data suggest that small molecule SSAO/VAP-1 inhibitors may provide clinical benefit in the treatment of acute and chronic inflammatory diseases.

Topics: Amine Oxidase (Copper-Containing); Animals; Anti-Inflammatory Agents; Carrageenan; Cell Adhesion; Cloning, Molecular; Colitis; Cyclooxygenase 2; Cytokines; Edema; Endothelial Cells; Endotoxemia; Female; Hydrazines; Inflammation; Male; Mice; Mice, Inbred Strains; Monoamine Oxidase Inhibitors; Oxazolone; Rats; Rats, Sprague-Dawley; Reverse Transcriptase Polymerase Chain Reaction

Activation of colonic proteinase activated receptor-1 (PAR1) provokes colonic inflammation and increases mucosal permeability in mice. The mechanism of inflammation is not neurogenic like in the paw of rats but depends on PAR1-mediated activation monocytic cells. PAR1 activation in the colon increases the release of lymphocyte T helper-1 (TH1) cytokines. Moreover, PAR1 expression is increased in biopsies from patients with inflammatory bowel disease, and its activation during TH1-mediated colitis in mice increases all of the hallmarks of inflammation.. This study aimed to characterize the effects of PAR1 activation in oxazolone-mediated colitis, involving a TH2 cytokine profile.. Intracolonic administration of oxazolone increased myeloperoxidase activity, damage score, and interleukin (IL)-4, IL-10, tumor necrosis factor alpha, and IL-1beta mRNA expression but lowered interferon-gamma mRNA expression, indicating colonic inflammation of a TH2 profile. The concurrent intracolonic administration of a PAR1 agonist in oxazolone-treated mice inhibited colitis, resulting in a reduction of myeloperoxidase activity, damage score, and inflammatory cytokine mRNA expression. Using PAR1-deficient mice, we confirmed that the anti-inflammatory effects of PAR1 agonists were mediated by PAR1. Moreover, in PAR1-deficient mice or in mice treated with a PAR1 antagonist, oxazolone-induced colitis was exacerbated, showing an endogenous modulatory role for PAR1 in this TH2 cytokine profile of colitis.. Thus, as opposed to a previously shown proinflammatory role for PAR1 in a TH1 cytokine-mediated colitis, our new data show anti-inflammatory role for PAR1 activation in the setting of TH2 cytokine colitis model.

Topics: Animals; Colitis; Cytokines; Inflammation; Intestinal Mucosa; Mice; Mice, Inbred C57BL; Oxazolone; Receptor, PAR-1; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Th1 Cells

Carcinoembryonic antigen-related cellular adhesion molecule 1 (CEACAM1) is a cell surface molecule that has been proposed to negatively regulate T cell function. We have shown that CEACAM1 is associated with specific regulation of T helper cell (Th)1 pathways, T-bet-mediated Th1 cytokine signaling, and Th1-mediated immunopathology in vivo. Mice treated with anti-mouse CEACAM1-specific monoclonal antibody (mAb) CC1 during the effector phase exhibited a reduced severity of trinitrobenzene sulfonic acid colitis in association with decreased interferon (IFN)-gamma production. Although oxazolone colitis has been reported as Th2 mediated, mice treated with the CC1 mAb or a CEACAM1-Fc chimeric protein exhibited a reduced severity of colitis in association with a significant reduction of IFN-gamma and T-bet activation, whereas signal transducer and activator of antigen 4 activation was unaffected. Both interleukin-4 and IFN-gamma gene-deficient mice exhibited less severe colitis induction by oxazolone. Direct ligation of T cells in vitro with the murine hepatitis virus spike protein, a natural ligand for the N-domain of CEACAM1, inhibited the differentiation of naive cells into Th1 but not Th2 cells and activation of Th1 but not Th2 cytokine production. These results indicate that CEACAM1 isoforms are a novel class of activation-induced cell surface molecules on T cells that function in the specific regulation of Th1-mediated inflammation such as that associated with inflammatory bowel disease.

Topics: Animals; Antibodies, Monoclonal; Carcinoembryonic Antigen; Colitis; Disease Models, Animal; Female; Immunoglobulin Fc Fragments; Inflammation; Interferon-gamma; Interleukin-1; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Mice, Knockout; Oxazolone; Recombinant Fusion Proteins; T-Lymphocytes, Helper-Inducer; Th1 Cells

Contact sensitivity (CS) is an inflammatory disorder characterized by early and late phases of leukocyte recruitment. We used a noninvasive intravital microscopy technique allowing for the direct visualization of leukocyte rolling and adhesion on blood vessel endothelium. By blocking specific adhesion molecules, we elucidated the molecular mechanisms mediating early leukocyte recruitment to be E- and P-selectin and demonstrated that leukocyte recruitment in the late phase had a different adhesive profile (mainly alpha(4)-integrin). Complete blockade of E- and P-selectin within the first 2 h of leukocyte-endothelial cell interactions (but not later) eliminated selectin-independent leukocyte recruitment at 24 h. Despite the predominance of neutrophils in the early phase, specific elimination of CD4(+) lymphocytes in the early phase eliminated the late response. CD4(+) lymphocytes homed to skin via E- and P-selectin within the early phase and induced the late phase response. Addition of these same CD4(+) lymphocytes 2 h after antigen challenge was too late for these cells to home to the skin and induce late phase responses. Our data clearly demonstrate that the antigen-challenged microenvironment is only accessible to CD4(+) lymphocytes for the first 2 h, and that this process is essential for the subsequent recruitment of other leukocyte populations in late phase responses.

Topics: Adoptive Transfer; Animals; CD4-Positive T-Lymphocytes; Cell Adhesion; Chemotaxis, Leukocyte; Dermatitis, Contact; E-Selectin; Hypersensitivity, Delayed; Inflammation; Leukocytes; Lymphocyte Transfusion; Male; Mice; Mice, Inbred C57BL; Oxazolone; P-Selectin; Receptors, Lymphocyte Homing; Selectins; Skin; Time Factors

Animal models are useful for studying disease, but there is a shortage of suitable models of ulcerative colitis. The aim of the present study was to set up an oxazolone-induced murine colitis model and use it to research the pathogenesis of inflammatory bowel disease.. BALB/c mice were presensitized by painting the skin with 0.2 mL 3% oxazolone in 100% ethanol on days 0 and 1 followed by intrarectal administration of 0.15 mL 1% oxazolone in 50% ethanol on day 7. The disease activity index (DAI), histological changes of the colon, myeloperoxidase (MPO) activity and production of cytokines (TNF-alpha, IL-4, IFN-gamma) by the mucosa were evaluated.. There were obvious changes in the DAI, histology and MPO activity, and the production of interleukin-4 was markedly increased compared with the concentrations of TNF-alpha and IFN-gamma, which remained normal, in the lesions.. Oxazolone colitis is Th2-mediated and has similar histologic features and distribution of inflammation to ulcerative colitis (UC), which has important implications for the use of this model in the study of the pathogenesis and treatment of UC.

Topics: Adjuvants, Immunologic; Animals; Colitis, Ulcerative; Cytokines; Disease Models, Animal; Female; Humans; Inflammation; Intestinal Mucosa; Mice; Mice, Inbred BALB C; Oxazolone; Peroxidase

The cellular immune response depends on the delivery of lymphocytes from the lymph node to the peripheral site of antigenic challenge. During their passage through the inflammatory microcirculaton, the migratory cells can become transiently immobilized or "trapped" in small caliber vessels. In this report, we used intravital microscopy and temporal area mapping to define the dynamic deformation of efferent lymph-derived mononuclear cells trapped in the systemic inflammatory microcirculation. Mononuclear cells obtained from the efferent lymph draining the oxazolone-stimulated microcirculation were labeled with fluorescent dye and reinjected into the feeding arterial circulation. Intravital video microscopy observed thousands of cells passing through the microcirculation; 35 cells were "trapped" in the oxazolone-stimulated microcirculation. Temporal area maps of the trapped cells demonstrated dramatic slowing and deformation. The cells were trapped in the microcirculation for a median of 8.90 sec (range 5-17 sec) prior to returning to the flow stream. During this period, the cells showed sustained movement associated with both antegrade locomotion (mean cell velocity = 7.92 microm/sec; range 1.16-14.23 microm/sec) and dynamic elongation (median cell length = 73.8 microm; range 58-144 microm). In contrast, efferent lymph-derived cells passing unimpeded through the microcirculation demonstrated rapid velocity (median velocity = 216 microm/sec) and spherical geometry (median diameter = 14.6 microm). Further, the membrane surface area of the "trapped" cells, calculated based on digital image morphometry and corrosion cast scanning electron microscopy, suggested that the fractional excess membrane of the cells in the efferent lymph was significantly greater than previous estimates of membrane excess. These data indicate that transient immobilization of efferent lymph-derived mononuclear cells in the systemic inflammatory microcirculation is rare. When "trapping" does occur, the shape changes and sustained cell movement facilitated by excess cell membrane may contribute to the return of the "trapped cells" into the flow stream.

Topics: Adjuvants, Immunologic; Animals; Cell Membrane; Cell Size; Chemotaxis, Leukocyte; Fluorescent Antibody Technique; Inflammation; Lymph Nodes; Lymphocytes; Microcirculation; Microscopy, Electron, Scanning; Monocytes; Oxazolone; Sheep

Angiogenesis and enhanced microvascular permeability are hallmarks of a large number of inflammatory diseases. Although up-regulation of proangiogenic factors such as vascular endothelial growth factor and interleukin-8 have been previously reported in inflamed tissue, the biologic role of endogenous inhibitors of angiogenesis in inflammation has remained unclear. To investigate the biologic role of the potent angiogenesis inhibitor thrombospondin-2 (TSP-2) in the control of cutaneous inflammation, delayed-type hypersensitivity reactions were elicited in the ear skin of wild-type and TSP-2-deficient mice by topical sensitization and challenge with oxazolone. Cutaneous TSP-2 expression was up-regulated in the inflamed skin of wild-type mice, predominantly in dermal fibroblasts and microvessels. Lack of TSP-2 resulted in a significantly enhanced inflammatory response with increased angiogenesis, edema formation, and inflammatory infiltration. Ear swelling and inflammation persisted for more than 2 weeks in TSP-2-deficient mice, as compared with 1 week in wild-type mice. Although baseline vascular permeability was unchanged, significantly enhanced microvascular leakage was found in the inflamed skin of TSP-2-deficient mice. Moreover, the fraction of rolling leukocytes was significantly increased in the untreated skin of TSP-2-deficient mice. These results reveal an important role of TSP-2 in limiting the extent and the duration of edema formation, angiogenesis, and inflammatory cell infiltration during acute and chronic inflammation.

Topics: Animals; Capillary Leak Syndrome; Capillary Permeability; Chemotaxis, Leukocyte; Dermatitis, Allergic Contact; Ear; Edema; Flow Cytometry; Image Processing, Computer-Assisted; Inflammation; Leukocyte Count; Male; Mice; Mice, Knockout; Neovascularization, Pathologic; Oxazolone; Thrombospondins

The systemic immune response is a dynamic process involving the trafficking of lymphocytes from the Ag-stimulated lymph node to the peripheral tissue. Studies in sheep have demonstrated several phases of cell output in the efferent lymph after Ag stimulation. When skin contact sensitizers are used as Ag, the efferent lymph cell output peaks approximately 96 h after Ag stimulation and is temporally associated with the recruitment of cells into the skin. To investigate the relative contribution of this high-output phase of efferent lymphocytes to lymphocytic inflammation in the skin, we used a common contact sensitizer 2-phenyl-4-ethoxymethylene-5-oxazolone (oxazolone) to stimulate the skin and draining prescapular lymph node of adult sheep. The efferent lymph ducts draining the Ag-stimulated and contralateral control lymph nodes were cannulated throughout the experimental period. The lymphocytes leaving the lymph nodes during the 72-h period before maximum infiltration were differentially labeled with fluorescent tracers, reinjected into the arterial circulation, and tracked to the site of Ag stimulation. Quantitative tissue cytometry of the skin at the conclusion of the injection period (96 h after Ag stimulation) demonstrated more migratory cells derived from the Ag-stimulated lymph node than the contralateral control (median 18.5 vs 15.5 per field; p < 0.05). However, when corrected for total cell output of the lymph node, the Ag-stimulated migratory cells were 3.8-fold more prevalent in the skin than the contralateral control cells. These results suggest that the in situ immune response generally mirrors the frequency of recruitable lymphocytes in the peripheral blood.

Topics: Adjuvants, Immunologic; Administration, Cutaneous; Animals; Antigens; Cell Movement; Ear; Fluorescent Dyes; Inflammation; Lymph; Lymph Nodes; Lymphatic System; Lymphocyte Count; Lymphocytes; Oxazolone; Sheep; Skin; Stochastic Processes

To elucidate if the planar chiral paracyclophane moiety conveys pharmacological activity to arylacetic acid analogs in two animal models.. Female NMRI mice (6 mice/group); female Wistar rats (8 rats/group); thrombocytes from human blood.. The enantiomers of [2.2]paracyclophaneacetic acid were applied locally (10(-7) and 10(-6) mol/ear) and orally (10-100 mg/kg).. (a) Phorbol myristyl acetate model of acute inflammation of the inner auricle. (b) Oxazolone model of allergic contact dermatitis. (c) Carrageenan model of acute inflammation. (d) Inhibition of cyclooxygenase-1 and 12-lipoxygenase (in vitro).. (a) PMA model: pR-(-)-[2.2]paracyclophaneacetic acid (10(-6) mmol/ear): 58% inhibition after 24 h (p < 0.05). (b) Oxazolone model: pR-(-)-[2.2]paracyclophaneacetic acid (10(-6) mmol/ear): 42% inhibition after 24 h (p < 0.05). (c) Carrageenan model: pR-(-)-[2.2]paracyclophaneacetic acid (10 mg/kg): 31.4% inhibition (paw volume 0.48 +/- 0.13 ml). (d) Cyclooxygenase-1 and 12-lipoxygenase: no inhibition at concentrations up to 10 microM.. The easily accessible [2.2]paracyclophane moiety should find its use in medicinal chemistry as it is a pharmacophoric substituent with the interesting feature of planar chirality.

Topics: Acetates; Animals; Anti-Inflammatory Agents, Non-Steroidal; Carrageenan; Edema; Female; Humans; Inflammation; Mice; Molecular Conformation; Oxazolone; Polycyclic Compounds; Rats; Rats, Wistar; Stereoisomerism; Structure-Activity Relationship; Tetradecanoylphorbol Acetate

1. Small, N- to C-terminal cyclized peptides containing the leucyl-aspartyl-valine (LDV) motif from fibronectin connecting segment-1 (CS-1) have been investigated for their effects on the adhesion of human T-lymphoblastic leukaemia cells (MOLT-4) to human plasma fibronectin in vitro mediated by the integrin Very Late Antigen (VLA)-4 (alpha4beta1, CD49d/CD29). 2. Cyclo(-isoleucyl-leucyl-aspartyl-valyl-aminohexanoyl-) (c(ILDV-NH(CH2)5CO)) was approximately 5 fold more potent (IC50 3.6+/-0.44 microM) than the 25-amino acid linear CS-1 peptide. Cyclic peptides containing two more or one less methylene groups had similar potency to c(ILDV-NH(CH2)5CO) while a compound containing three less methylene groups, c(ILDV-NH(CH2)2CO), was inactive at 100 microM. 3. c(ILDV-NH(CH2)5CO) had little effect on cell adhesion mediated by two other integrins, VLA-5 (alpha5,beta1, CD49e/CD29) (K562 cell adhesion to fibronectin) or Leukocyte Function Associated molecule-1 (LFA-1, alphabeta2, CD11a/CD18) (U937 cell adhesion to Chinese hamster ovary cells transfected with intercellular adhesion molecule-1) at concentrations up to 300 microM. 4. c(ILDV-NH(CH2)5CO) inhibited ovalbumin delayed-type hypersensitivity or oxazolone contact hypersensitivity in Balb/c mice when dosed continuously from subcutaneous osmotic mini-pumps (0.1-10 mg kg(-1) day(-1)). Maximum inhibition (approximately 40%) was similar to that caused by the monoclonal antibody PS/2 (7.5 mg kg(-1) i.v.) directed against the alpha4 integrin subunit. 5. c(ILDV-NH(CH2)5CO) also inhibited oxazolone contact hypersensitivity when dosed intravenously 20 h after oxazolone challenge (1-10 mg kg(-1)). Ear swelling was reduced at 3 h and 4 h but not at 1 h and 2 h post-dose (10 mg kg(-1)). 6. Small molecule VLA-4 inhibitors derived from c(ILDV-NH(CH2)5CO) may be useful as anti-inflammatory agents.

Topics: Amino Acid Sequence; Animals; Anti-Inflammatory Agents, Non-Steroidal; Cell Adhesion; CHO Cells; Cricetinae; Dermatitis, Contact; Female; Fibronectins; Humans; Hypersensitivity, Delayed; Inflammation; Integrin alpha4beta1; Integrins; Intercellular Adhesion Molecule-1; Intercellular Signaling Peptides and Proteins; Leukemia, Erythroblastic, Acute; Leukemia, T-Cell; Mice; Mice, Inbred BALB C; Molecular Sequence Data; Ovalbumin; Oxazolone; Peptides; Rats; Receptors, Lymphocyte Homing; T-Lymphocytes; Transfection

In this study we describe oxazolone colitis, a new form of experimental colitis. This model is induced in SJL/J mice by the rectal instillation of the haptenating agent, oxazolone, and is characterized by a rapidly developing colitis confined to the distal half of the colon; it consists of a mixed neutrophil/lymphocyte infiltration limited to the superficial layer of the mucosa which is associated with ulceration. Oxazolone colitis is a T helper cell type 2 (Th2)-mediated process since stimulated T cells from lesional tissue produce markedly increased amounts of interleukin (IL)-4 and IL-5; in addition, anti-IL-4 administration leads to a striking amelioration of disease, whereas anti-IL-12 administration either has no effect or exacerbates disease. Finally, this proinflammatory Th2 cytokine response is counterbalanced by a massive transforming growth factor-beta (TGF-beta) response which limits both the extent and duration of disease: lesional (distal) T cells manifest a 20-30-fold increase in TGF-beta production, whereas nonlesional (proximal) T cells manifest an even greater 40-50-fold increase. In addition, anti-TGF-beta administration leads to more severe inflammation which now involves the entire colon. The histologic features and distribution of oxazolone colitis have characteristics that resemble ulcerative colitis (UC) and thus sharply distinguish this model from most other models, which usually resemble Crohn's disease. This feature of oxazolone colitis as well as its cytokine profile have important implications to the pathogenesis and treatment of UC.

Topics: Administration, Rectal; Animals; Antibodies; Colitis; Colitis, Ulcerative; Colon; Cytokines; Disease Models, Animal; Histocytochemistry; Humans; Inflammation; Interleukin-4; Interleukins; Mice; Mice, Inbred Strains; Oxazolone; Th2 Cells

Transcription factor interferon regulatory factor-1 (IRF-1) is implicated in regulating class I MHC expression in vitro. We investigated the in vivo relationship between IRF-1 and MHC expression in kidney and other nonlymphoid organs, assessing MHC expression in mice with disrupted IRF-1 genes (IRF-1 KO) compared with mice with intact IRF-1 genes (WT). In kidneys of IRF-1 KO mice, basal class I expression was decreased, particularly on arterial endothelium, but basal class II expression was unchanged. The induction of both class I and class II expression by injected rIFN-gamma was reduced in IRF-1 KOs, compared with WT mice. Similarly, stimuli that induce endogenous IFN-gamma production (LPS or oxazolone) massively increased MHC expression in kidneys of WT mice, with little increase in IRF-1 KO mice. Impaired class II induction by rIFN-gamma in IRF-1 KO mice probably reflects the role of IRF-1 in regulating class II transactivator (CIITA) expression: rIFN-gamma induced CIITA mRNA less in kidneys of IRF-1 KO mice than in WT mice. In organs of WT mice, IRF-1 mRNA was expressed in the basal state, and rIFN-gamma treatment increased IRF-1 mRNA before the induction of class I or CIITA mRNA. Treatment of WT mice with cycloheximide plus rIFN-gamma superinduced IRF-1 mRNA expression, but partially inhibited CIITA mRNA expression, indicating that IRF-1 mRNA induction is not dependent on new protein synthesis, unlike CIITA. Thus, in vivo, IRF-1 plays a major role in basal and induced class I expression and in induction of class II by IFN-gamma, probably via CIITA induction.

Topics: Animals; Cycloheximide; DNA-Binding Proteins; Gene Expression Regulation; Genes, MHC Class I; Genes, MHC Class II; Inflammation; Interferon Regulatory Factor-1; Interferon-gamma; Lipopolysaccharides; Mice; Mice, Inbred BALB C; Mice, Knockout; Nuclear Proteins; Oxazolone; Phosphoproteins; Protein Synthesis Inhibitors; Recombinant Proteins; RNA, Messenger; Time Factors; Tissue Distribution; Trans-Activators

Allergic contact dermatitis differs from most other immune reactions by its strict dose dependence during the elicitation phase. Moreover, almost all known contact allergens can also induce dose-dependent irritative dermatitis and in general only elicit allergic contact dermatitis in sensitized individuals when applied within a narrow dose range. Therefore, we hypothesized that elicitation of contact hypersensitivity (CHS) may require two signals, antigen-specific effector cell activation and a non-antigen-specific proinflammatory signal, both of which are provided by application of a sufficient dose of hapten. To dissociate these putative two signals, oxazolone-sensitized mice were ear challenged with a dose of the specific hapten which was too low to elicit CHS. At the same time, an unrelated hapten was applied in a conventional concentration to the same skin site. Whereas neither treatment alone elicited a significant CHS response, application of both compounds together resulted in a strong CHS response that was indistinguishable from that elicited by the full dose of the specific hapten. Upon coadministration of the irrelevant hapten, allergic contact dermatitis could be elicited even when the dose of the specific hapten was further reduced by a factor of 10(3). In contrast, a dose reduction of the irrelevant hapten by a factor of two resulted in the loss of the CRS response. These data indicate that non-antigen-specific effects of epicutaneously applied haptens significantly contribute to the elicitation of CHS responses and that the capacity of the hapten to evoke this proinflammatory stimulus rather than its antigenicity is responsible for the strict concentration dependence.

Topics: Administration, Cutaneous; Animals; Dermatitis, Allergic Contact; Dose-Response Relationship, Drug; Female; Haptens; Immunization; Inflammation; Irritants; Male; Mice; Mice, Inbred BALB C; Mice, Inbred C3H; Oxazolone; Picryl Chloride

The inflammatory response at sites of contact hypersensitivity induced by oxazolone was examined in the ears of P-selectin-deficient and wild-type mice. Accumulation of CD4+ T lymphocytes, monocytes, and neutrophils was reduced significantly in the mutant mice, as well as mast cell degranulation. In contrast, there was no significant difference in vascular permeability or edema between the two genotypes. The results demonstrate a role for P-selectin in recruitment of CD4+ T lymphocytes and show that P-selectin plays a role in long-term inflammation as well as in acute responses.

Topics: Animals; Dermatitis, Contact; Female; Inflammation; Mast Cells; Mice; Mice, Inbred C57BL; Mice, Inbred Strains; Mice, Mutant Strains; Neutrophils; Oxazolone; P-Selectin; Platelet Membrane Glycoproteins; Skin

L-selectin, a cell surface adhesion molecule that is expressed by most leukocytes, mediates leukocyte rolling along vascular endothelium at sites of inflammation. The contribution of L-selectin to leukocyte migration in models of chronic inflammation was assessed by using mice that lack cell surface L-selectin expression. Significant inhibition of neutrophil (56-62%), lymphocyte (70-75%), and monocyte (72-78%) migration into an inflamed peritoneum was observed 24 and 48 h after administration of thioglycollate, an inflammatory stimulus. L-selectin-deficient mice were also significantly impaired in delayed-type hypersensitivity reactions. Footpad swelling in response to sheep red blood cell challenge was reduced 75% in L-selectin-deficient mice compared with wild-type mice. Ear swelling in a model of contact hypersensitivity induced by oxazolone challenge was also reduced by 69% compared to wild-type mice. Consistent with L-selectin-mediating leukocyte migration into diverse vascular beds during inflammation, L-selectin-deficient mice were significantly resistant to death resulting from lipopolysaccharide (LPS)-induced toxic shock. LPS administration resulted in a 90% mortality rate in control mice after 24 h, while there was a 90% survival rate in L-selectin-deficient mice. These results demonstrate that L-selectin plays a prominent role in leukocyte homing to nonlymphoid tissues during inflammation and that blocking this process can be beneficial during pathological inflammatory responses.

Topics: Animals; Cell Adhesion; Cell Adhesion Molecules; Crosses, Genetic; Dermatitis, Contact; Endothelium, Vascular; Inflammation; L-Selectin; Leukocytes; Lymphocytes; Macrophages; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Neutrophils; Oxazolone

Topics: Administration, Cutaneous; Administration, Sublingual; Animals; Antigen Presentation; Dendritic Cells; Dermatitis, Contact; Ear, External; Hindlimb; Immunotherapy, Adoptive; Inflammation; Langerhans Cells; Lymph Nodes; Mice; Mice, Inbred BALB C; Mouth Mucosa; Organ Specificity; Oxazolone; Picryl Chloride; Skin

Tumor facilitating factor is a cell surface glycoprotein produced by B16 melanoma that has been found to reduce the lethal inoculum for B16. Tumor facilitating factor induces macrophage spreading in vitro, reduces macrophage chemotaxis in vivo, and depresses lymphocyte mitogenesis in vitro.. It is assumed that the immune modifying effects are responsible for tumor facilitation. As tumors may be poor immunogens or inducers of inflammation, studies were conducted to determine whether tumor facilitating factor alters the inflammatory cascade of cells found in infiltrates of delayed type hypersensitivity.. Freeze-thawed B16 cells, used as the source of TFF, caused a suppression of delayed type hypersensitivity measured as ear swelling in the mouse. When culture supernatant was substituted for freeze-thawed cells as a source of TFF and injected at different time points of the delayed type hypersensitivity response, the greater suppression was with tumor facilitating factor injections at 24 hours pre-elicitation only (82%), and 24 hours both presensitization and pre-elicitation (89%). Immunohistological staining demonstrated that tumor facilitating factor decreases ear thickness and cellular infiltrates, specifically Mac-1 staining cells, to a site of delayed hypersensitivity. Peritoneal cell analysis confirmed these findings.. These data are consistent with the hypothesis that tumor facilitating factor alters immune functions including macrophage and lymphocyte mobility and recruitment to a target site, thereby allowing for facilitation of tumor growth.

Topics: Animals; Dinitrofluorobenzene; Dose-Response Relationship, Immunologic; Female; Freezing; Growth Substances; Hypersensitivity, Delayed; Inflammation; Kinetics; Melanoma, Experimental; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Oxazolone; Peritoneum; Tumor Cells, Cultured

Methyl 20 beta-dihydroprednisolonate, a new local antiinflammatory steroid, and its ester hydrolysis product, 20 beta-dihydroprednisolonic acid, were evaluated for potential immunosuppressive actions as determined by the splenic plaque-forming cell response assay. The parent compound, prednisolone, was used as a comparative agent. All three corticosteroid agents, when administered intraperitoneally or subcutaneously by separate injections given at the same time as antigen, demonstrated significant dose-related suppression of this immunologic parameter. In a croton oil-induced inflammation test (ear challenge in mice), methyl 20 beta-dihydroprednisolonate demonstrated significant antiinflammatory effects, but only in the ear to which it was applied. The comparative drug, prednisolone, significantly reduced inflammation in the ear to which it was applied and in the contralateral ear as well. The methyl ester derivative demonstrated effective antiinflammatory activity when applied topically in the oxazolone delayed-type hypersensitivity test (ear challenge in mice). The results seen in the plaque-forming cell response are in contrast to the generally noted lack of systemic glucocorticoid actions of the steroid acid ester derivatives reported in other studies.

Topics: Animals; Dose-Response Relationship, Drug; Edema; Female; Hemolytic Plaque Technique; Hypersensitivity, Delayed; Immunosuppressive Agents; Inflammation; Mice; Oxazolone; Prednisolone; Spleen

Various models of delayed hypersensitivity (DH) were used in mice: contact hypersensitivity reactions to picryl chloride and oxazolone and reactions to methylated bovine serum albumin (MBSA) and sheep red blood cells (SRBC). Drugs of different classes were tested in these models by systemic treatment around the challenge period: non-steroidal anti-inflammatory drugs (cyclooxygenase inhibitors, and inhibitors of both cyclooxygenase and lipoxygenase); glucocorticoids and immunosuppressants (cyclosporin A. CsA; cyclophosphamide, Cy; methotrexate, Mtx; azathioprine, Aza). These compounds were also studied and compared for their effects on the 3-h and 24-h phase of the carrageenin mouse-paw edema (in which inflammation is maximal after 24 h). Non-steroidal anti-inflammatory drugs (including double inhibitors of cyclooxygenase and lipoxygenase) had little or no effect on DH models, except indometacin. Glucocorticoids inhibited all immune reactions except that to MBSA. Of the immunosuppressants, CsA reduced all the DH reactions while Aza mainly reduced the reaction to SRBC; Cy and Mtx were mainly active on SBRC and MBSA inflammations. On another hand CsA, Cy and Mtx were inactive on the 3-h phase but decreased the 24-h phase of carrageenin edema. At doses active on the DH models and on carrageenin inflammation, Cy induced a lasting blood leukopenia, but CsA and Mtx did not. This combination of tests in the mouse seems to be of interest to demonstrate any action on DH and any anti-inflammatory effect and to suggest whether these activities are related to a possible leukopenic effect.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Carrageenan; Dermatitis, Contact; Edema; Glucocorticoids; Hypersensitivity, Delayed; Immunosuppressive Agents; Inflammation; Kinetics; Leukocyte Count; Male; Mice; Oxazolone; Picryl Chloride; Serum Albumin, Bovine; Sheep

Oxazolone-induced delayed hypersensitivity in mice produced swelling with concomitant increased tissue levels of leukotrienes and prostaglandins. Pharmacological agents were coapplied topically with oxazolone at the time of challenge in an attempt to modulate the immune-based inflammation. Dexamethasone inhibited both swelling and increases in eicosanoid levels. Indomethacin reduced prostaglandin levels but failed to inhibit swelling or reduce leukotriene levels. L-651,896 (2,3-dihydro-6-[3-(2-hydroxymethyl)phenyl-2-propenyl]-5-benzofuranol), a 5-lipoxygenase inhibitor, reduced leukotriene levels but did not reduce swelling or prostaglandin levels. A combination of indomethacin and L-651,896 reduced eicosanoid levels but did not reduce swelling. These data suggested that the reduction in tissue levels of 5-lipoxygenase or cyclooxygenase oxygenation products of arachidonic acid either singularly or together did not result in the concomitant reduction of the inflammation associated with oxazolone-induced delayed hypersensitivity.

Topics: Animals; Cyclooxygenase Inhibitors; Dexamethasone; Dinoprostone; Female; Hypersensitivity, Delayed; Inflammation; Leukotriene B4; Lipoxygenase Inhibitors; Mice; Oxazoles; Oxazolone; Prostaglandins E; SRS-A

In previous studies, the alpha 2-adrenoceptor agonist clonidine has been shown to suppress the wheal and flare reaction in guinea pigs sensitized to ovalbumin. This phenomenon has been further studied with special reference to effects on the dermal inflammatory cell infiltrate and mast cells. Clonidine lessens the degranulation of mast cells seen in control untreated immediate hypersensitivity reactions. Less neutrophils and eosinophils arrive to the treated reactions. Basophils and mononuclear cells (chiefly lymphocytes) which characterize the late phase of the wheal and flare reaction were not influenced by clonidine. Clonidine had a possible minimal effect on allergic contact (delayed hypersensitivity) reactions. The toxic contact reaction to croton oil (nonspecific cutaneous inflammation) was not affected.

Topics: Animals; Cell Movement; Clonidine; Croton Oil; Dermatitis, Contact; Drug Hypersensitivity; Female; Granulocytes; Guinea Pigs; Histamine; Hypersensitivity, Delayed; Hypersensitivity, Immediate; Inflammation; Mast Cells; Ovalbumin; Oxazolone; Skin; Skin Tests

The percutaneous administration of in vitro haptenated epidermal cells (EC) has become established as a procedure to produce contact sensitivity (CS) in experimental animals for routine use. The cells have also been found to elicit a significant delayed-type skin reaction by intradermal test in the animals sensitized by painting the skin with the hapten. The fate of 2,4-dinitrophenylated (DNP) isogeneic epidermal cell suspensions (EC) injected intradermally was investigated histologically in intact or 2,4-dinitrochlorobenzene (DNCB)-sensitized strain 13 guinea pigs to study the role of the cells in CS. DNP-EC were found to proliferate actively in the dermis and formed EC nests with central keratinization and then elicited inflammatory reaction associated with necrosis of the epidermal structures 7 days after injection in the intact animals. DNP-EC injected intradermally into the animals which had received and reacted against DNCB underwent a suppression of EC proliferation. These findings are discussed in relation to the role of the haptenated EC in CS.

Topics: Animals; Dinitrobenzenes; Dinitrochlorobenzene; Epidermis; Guinea Pigs; Haptens; Hypersensitivity, Delayed; Inflammation; Injections, Intradermal; Male; Oxazolone; Skin Diseases

Picryl chloride applied to the ears of Swiss mice induced a clearcut primary irritation inflammation (maximal after 3 to 6 hr) and after contact sensitization performed 7 days before a delayed hypersensitivity reaction. Oxazolone produced only a weak primary irritation reaction. After contact sensitization in the same conditions as above, oxazolone induced an immune response that was already substantial 3 to 6 hr after the challenge and generally reached a maximum after 24 hr. We tried to alter these four types of inflammation (the primary irritation and delayed hypersensitivity to picryl chloride, and the 6-hr and 24-hr phases of hypersensitivity to oxazolone) by various types of compounds administered cutaneously or sytemically. Mepyramine, methysergide, cimetidine, disodium cromoglycate, phenylbutazone, and acetylsalicylic acid reduced to varying degrees after cutaneous application the primary irritation and the delayed hypersensitivity inflammation induced by picryl chloride. Methysergide was the only one of these drugs that on topical application clearly reduced the immune response to oxazolone (decrease in the 6-hr phase). After systemic administration, these same drugs had no effect on the four types of reaction. Both the corticosteroids tested (hydrocortisone acetate and desonide) reduced all the inflammations to various degrees and were always more active (particularly desonide) when applied topically than when administered systemically. On the other hand indomethacin, which inhibited all types of inflammation, was more active when administered systemically. Study of the kinetics and trials of pharmacological modulation of the various reactions induced by picryl chloride and oxazolone in Swiss mice provided evidence of differences in behavior between the two agents.

Topics: Adrenal Cortex Hormones; Animals; Anti-Inflammatory Agents; Dermatitis, Contact; Hypersensitivity, Delayed; Inflammation; Irritants; Kinetics; Male; Mice; Oxazoles; Oxazolone; Picryl Chloride

The topical effects of N0164 (a phenyl phosphonate derivative which is a partially selective antagonist of prostagladin E2), indomethacin and triamcinolone acetonide have been shown to reduce the erythema and ear weight gain from inflammation induced by experimental contact allergic eczema. Oxazolone sensitized Swiss Webster mice were used, ear erythema and ear weights being used as a measure of the anti-inflammatory response to the drugs. N0164 was also shown to have systemic anti-inflammatory activity after intraperitoneal injection.

Topics: Administration, Topical; Animals; Anti-Inflammatory Agents; Dermatitis, Contact; Erythema; Indomethacin; Inflammation; Injections, Intraperitoneal; Mice; Organophosphonates; Oxazolone; Prostaglandin Antagonists; Triamcinolone Acetonide

Topics: Animals; Capillary Permeability; Cell Movement; Female; Inflammation; Male; Mice; Mice, Inbred BALB C; Neutrophils; Oxazoles; Oxazolone; Reserpine; Serotonin