Search

Background and Aims: 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., Methods: 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., Results: 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., Conclusions: These findings indicate that Nod2 signalling may prevent pathological conversion of T helper cells for maintenance of tissue homeostasis., (© The Author(s) 2022. Published by Oxford University Press on behalf of European Crohn’s and Colitis Organisation. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

A decrease in the number of basophils in the peripheral blood, or basopenia, has been noted, reflecting the activity of chronic spontaneous urticaria (CSU). Infiltration of basophils into the skin has also been reported, but the mechanism of basopenia in CSU has not been clarified. The phenomenon of basopenia during the active phase of urticaria was confirmed, and basophil numbers increased following symptom improvement in 15 out of 17 patients treated with omalizumab and in 13 of 15 patients treated with antihistamines. Our examination by immunostaining also revealed basophil infiltration of the CSU lesions, as in previous reports, but since most of our patients were already taking oral steroids, it was not considered appropriate to examine the relationship between basophil numbers in tissue and peripheral blood. Then, we used mouse model of contact hypersensitivity with a single application of oxazolone, which is known to stimulate basophil infiltration, and investigated basophil counts in the skin, peripheral blood, and bone marrow. In this model, a decrease in peripheral blood basophil numbers was observed one day after challenge, but not after 2 days, reflecting supplementation from the bone marrow. Indeed, when cultured basophils expressing GFP were transplanted into the peripheral blood, GFP-positive basophil numbers in the peripheral blood remained low even after 2 days of challenge. Despite differences among species and models, these results suggest that one reason for the decrease of basophils in the peripheral blood in CSU may involve migration of circulating basophils into the skin., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Kishimoto, Ma, Takimoto-Ito, Nakashima, Otsuka, Walls, Tanizaki and Kambe.)

7-Met, a derivative of soybean isoflavone, is a natural flavonoid compound that has been reported to have multiple signaling pathways regulation effects. This study investigated the therapeutic effects of 7-Met on mice with atopic dermatitis induced by fluorescein isothiocyanate (FITC), or oxazolone (OXZ). 7-Met ameliorated FITC or OXZ-induced atopic dermatitis symptoms by decreasing ear thickness, spleen index, mast cell activation, neutrophil infiltration and serum IgE levels in female BALB/c mice. In FITC-induced atopic dermatitis mice, 7-Met reduced Th1 cytokines production and regulated Th1/Th2 balance by downregulating the secretion of thymic stromal lymphopoietin (TSLP) via inactivation of the NF-κB pathway. In OXZ-induced atopic dermatitis, 7-Met functioned through the reduction of Th17 cytokine production. Our study showed that 7-Methoxyisoflavone alleviated atopic dermatitis by regulating multiple signaling pathways and downregulating chemokine production., (© 2022. The Author(s).)

Background: Cutaneous bacterial dysbiosis is a characteristic hallmark of atopic dermatitis (AD), and it decisively influences the severity of the disease. Despite this, frequently used murine models of AD have not been characterized regarding the changes in skin microbiome communities., Objective: To analyse the skin microbiome of two frequently used murine models for AD for assessing their applicability in translational research., Methods: AD was induced in mice by topical application of calcipotriol or oxazolone. Following comparable elicitation of AD-like dermatitis, including IgE induction, the skin microbial communities were analysed and compared with human AD., Results: We detected critical differences in the microbiota composition of diseased skin. In contrast to calcipotriol treatment, application of oxazolone induced significant changes in the cutaneous microbiota and a drastic drop of bacterial richness. Furthermore, an expansion of Staphylococci, particularly S. xylosus, was observed in the oxazolone group, also displaying positive correlations with AD key markers including pH, TEWL, IL-4, TSLP and IL-33., Conclusions: In this article, we show that (a) the model of choice to investigate AD needs to be characterized for the cutaneous microbiota if applicable and (b) the oxazolone-mediated mixed Th1-Th2 immune response triggers microbiota-induced alterations which share similarities to dysbiosis in human AD and represents therefore a suitable model for translational research on AD if alterations of the microbiome are in the focus of the investigation., (© 2022 The Authors. Journal of the European Academy of Dermatology and Venereology published by John Wiley & Sons Ltd on behalf of European Academy of Dermatology and Venereology.)

Black soybean has been used in traditional medicine to treat inflammatory diseases, cancer, and diabetes and as a nutritional source since ancient times. We found that Korean black soybean cultivar A63 has more cyanidin-3- O -glucoside, (C3G), procyanidin B2 (PB2), and epicatechin (EPC) contents than other cultivars and has beneficial effects on cell viability and anti-oxidation. Given the higher concentration of anthocyanidins and their strong anti-oxidant activity, we predicted that A63 extract could relieve inflammatory disease symptoms, including those of atopic dermatitis (AD). Here, we evaluated the anti-AD activity of A63 extract in an oxazolone (OXA)-induced mouse model. A63 extract treatment significantly reduced epidermal thickness and inflammatory cell infiltration, downregulated the expression of AD gene markers, including Interleukin (IL)-4 and IL-5, and restored damaged skin barrier tissues. Furthermore, A63 extract influenced the activation of the signal transducer and activator of transcription (STAT) 3 and STAT6, extracellular regulatory kinase (ERK), and c-Jun N-terminal kinase (JNK) signaling pathways, which play a crucial role in the development of AD. Altogether, our results suggest that A63 can ameliorate AD-like skin inflammation by inhibiting inflammatory cytokine production and STAT3/6 and Mitogen-activated protein kinase (MAPK) signaling and restoring skin barrier function.

Background: Allergic dermatitis (AD) is a chronic inflammatory skin disease that a variety of immune cells are involved in the progression of AD. Among them, T cells are one of major players of AD pathogenesis. The V-domain Ig suppressor of T cell activation (VISTA) has been reported that it has a potential immunomodulatory for T cell response., Objective: This study aimed to investigate immunomodulatory of recombinant VISTA-Ig fusion protein in AD mice model., Methods: The model of AD was built with oxazolone (OXA) in BALB/c mice, then VISTA-Ig was used to treat AD by intraperitoneal (IP) injection. The ear thickness was measured by a digital thickness gauge. The ears tissues were collected and subjected to hematoxylin-eosin (H&E) and toluidine blue (TB) staining. The secretion levels of IL-4 and IgE in the serum were measured by enzyme linked immunosorbent assay (ELISA). The mRNA expression levels of inflammatory cytokines (IL-1β, IL-6, IL-12, and INF-γ) in ear tissues were measured using reverse transcription-quantitative polymerase chain reaction (RT-qPCR)., Results: Treatment with VISTA-Ig successfully alleviated the symptoms of AD, such as erythema, horny substance, and swelling. The infiltration of inflammatory cells was significantly reduced following VISTA-Ig therapy. The secretion levels of IL-4 and IgE in the serum were significantly attenuated following treatment with VISTA-Ig. Additionally, VISTA-Ig observably down-regulated inflammatory cytokines expression in ear tissues., Conclusions & Clinical Relevance: Taken together, our results showed that VISTA-Ig possessed the potential to be a novel immunomodulatory candidate drug against AD.

Allergic contact dermatitis (ACD) is a common allergic skin disease that affects individuals subjected to different antigen exposure conditions and significantly impacts the quality of life of those affected. Numerous studies have demonstrated that probiotics suppress inflammation through immunomodulatory effects. In this study, we aimed to evaluate the effect of the probiotic Bifidobacterium longum 5 1A as a preventive treatment for ACD using an oxazolone-induced murine model. We demonstrated that B. longum 5 1A exerted a prophylactic effect on oxazolone-induced ACD-like skin inflammation via reductions in ear and dermal thickness and leucocyte infiltration. The administration of inactivated B. longum 5 1A did not affect oxazolone-induced ACD-like skin inflammation, suggesting that the bacteria must be alive to be effective. Given that B. longum 5 1A is an acetate producer, we treated mice with acetate intraperitoneally, which also prevented ear and dermal thickening. Moreover, the tissue levels of the inflammatory cytokines and chemokines interleukin (IL)-10, IL-33, tumour necrosis factor-α, chemokine (C-C motif) ligand 2/monocyte chemoattractant protein-1 and chemokine (C-C motif) ligand 5/RANTES were significantly reduced after probiotic treatment, but only IL-33 and IL-10 were reduced when the mice were treated with acetate. These results show that B. longum 5 1A exerted a potential prophylactic effect on skin inflammation and that acetate represents one potential mechanism. However, other factors are likely involved since these two treatments do not yield the same results.

Atopic dermatitis (AD) is the most common inflammatory skin condition. Skin barrier dysfunction is of major importance in AD because it facilitates allergen sensitization and systemic allergic responses. Long regarded as a pro-apoptotic protease, emerging studies indicate granzyme B (GzmB) to have extracellular roles involving the proteolytic cleavage of extracellular matrix, cell adhesion proteins, and basement membrane proteins. Minimally expressed in normal skin, GzmB is elevated in AD and is positively correlated with disease severity and pruritus. We hypothesized that GzmB contributes to AD through extracellular protein cleavage. A causative role for GzmB was assessed in an oxazolone-induced murine model of dermatitis, comparing GzmB -/- mice with wild-type mice, showing significant reductions in inflammation, epidermal thickness, and lesion formation in GzmB -/- mice. Topical administration of a small-molecule GzmB inhibitor reduced disease severity compared with vehicle-treated controls. Mechanistically, GzmB impaired epithelial barrier function through E-cadherin and FLG cleavage. GzmB proteolytic activity contributes to impaired epidermal barrier function and represents a valid therapeutic target for AD., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Glucocorticoids (GCs) are potent anti-inflammatory drugs, the secretion of which is mediated and controlled by the hypothalamic-pituitary-adrenal axis. However, they are also secreted de novo by peripheral tissues for local use. Several tissues express 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1), including the skin. The inactive GC cortisone is converted by 11β-HSD1 to active GC cortisol, which is responsible for delayed wound healing during a systemic excess of GC. However, the role of 11β-HSD1 in inflammation is unclear. We assessed whether 11β-HSD1 affects the development of atopic dermatitis (AD) in vitro and in vivo. The expression of 11β-HSD1 in the epidermis of AD lesions was higher than that in the epidermis of healthy controls. Knockdown of 11β-HSD1 in human epidermal keratinocytes increased the production of thymic stromal lymphopoietin. In an oxazolone-induced mouse model of AD, localized inhibition of 11β-HSD1 aggravated the development of AD and increased serum cytokine levels associated with AD. Mice with whole-body knockout (KO) of 11β-HSD1 developed significantly worse AD upon induction by oxazolone. We propose that 11β-HSD1 is a major factor affecting AD pathophysiology via suppression of atopic inflammation due to the modulation of active GC in the skin.

Crohn's disease (CD) and ulcerative colitis (UC) actually had different pathological mechanisms, as the former was mainly induced by Th1 and Th17 response and the latter by Th2 response. Our previous study found that oxazolone-induced Th2-mediated colitis could not be attenuated by vitamin D supplementation. This study investigated the influence of intestinal vitamin D receptor (VDR) knockout on oxazolone-induced colitis and explored the possible immunological mechanism. Intestinal VDR knockout mice had milder oxazolone-induced colitis than wildtype controls, as demonstrated by less body weight decrease and faster recovery, more intact local structure, reduced cell apoptosis, and better preserved barrier function. Th2-mediated inflammation was significantly inhibited by VDR deficiency. Meanwhile, the percentage of invariant natural killer T (iNKT) cells did not increase as much in intestinal VDR knockout mice as in wild-type controls, nor did the iNKT cells develop normally as in the controls. Intestinal VDR knockout protected against oxazolone-induced colitis in mice by blocking Th2 cell response and reducing the function of intestinal iNKT cells. Vitamin D status had no influence on the severity of colitis. This study may explain the diverse outcomes after vitamin D supplementation in literature and add some clue to the targeted therapy of IBD.

Nintedanib, a receptor tyrosine kinase (RTK) inhibitor has been developed as therapeutics for idiopathic pulmonary fibrosis and non-small lung cancer. We found that the expression levels of RTK, especially VEGFR1 is increased in skin biopsies of dermatitis patients from multiple independent datasets. Moreover, VEGFR1 is highly expressed by infiltrated cells in dermis from oxazolone (OXA) treated mice. Interestingly, nintedanib alleviates dermatitis symptom in OXA-induced animal model. Especially, levels of epidermis thickness, infiltrated immune cells including mast cells and eosinophils were decreased from mice cotreated with nintedanib and OXA compared with OXA treated mice. Moreover, serum IgE and Th2 cytokines including IL-4 and IL-13 were decreased by nintedanib treatment. These results suggest an evidence that nintedanib alleviates animal model of dermatitis.

The function of regulatory immune cells in peripheral tissues is crucial to the onset and severity of various diseases. Interleukin-10 (IL-10)-producing regulatory B (IL-10 + B reg ) cells are known to suppress various inflammatory diseases. However, evidence for the mechanism by which IL-10 + B reg cells are generated and maintained is still very limited. Here, we found that IL-10 + B reg cells suppress the activation of IL-13-producing type 2 innate lymphoid cells (IL-13 + ILC2s) in an IL-10-dependent manner in mice with oxazolone-induced severe contact hypersensitivity (CHS). Mast cell (MC) IL-5 was important for maintaining the population of IL-10 + B reg cells in peripheral lymphoid tissues. Overall, these results uncover a previously unknown mechanism of MCs as a type of immunoregulatory cell and elucidate the cross-talk among MCs, IL-10 + B reg cells, and IL-13 + ILC2s in CHS.

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 + CD8 + regulatory T cells, while they did not differ in any inflammation parameters. No significant differences were found for intraepithelial lymphocytes or mesenteric lymphocytes at any time point after colitis induction. In conclusion, the priming did decrease local acute tissue inflammation of the colon in this commonly applied mouse model of T helper cell type 2-dominated model of inflammatory bowel disease.

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 low . Flow cytometry analysis of mesenteric lymph node cells showed that P2X7 activation (by ATP) triggered regulatory T cell death. In addition, such cells from P2X7 KO mice expressed more CD103, suggesting increased migration of regulatory T cells to the colon (relative to the WT). Our results show that the P2X7 has a key role during inflammation development in inflammatory bowel disease, by triggering the death and retention in the mesenteric lymph nodes of regulatory T cells that would otherwise promote immune system tolerance in the gut., (Copyright © 2017 Elsevier B.V. All rights reserved.)

The inflammasomes induce maturation of pro-interleukin-1β (IL-1β) and pro-IL-18. We investigated roles of the NLRP3 inflammasome in the pathogenesis of ulcerative colitis (UC). After induction of oxazolone-induced colitis, a mouse UC model, colonic tissues were assayed for inflammatory mediators. Histological studies were performed on inflamed colonic tissue from mice and UC patients. Histological severity of murine colitis peaked on day 1, accompanied by an increase in the expression of Th2 cytokines including IL-4 and IL-13. Oxazolone treatment stimulated maturation of pro-caspase-1 and pro-IL-1β, while it reduced IL-18 expression. Either exogenous IL-1β or IL-18 ameliorated the colitis with or without reduction in Th2 cytokine expression, respectively. Induction of colitis decreased MUC2 expression, which was reversed by administration of IL-18, but not IL-1β. Compared to wild-type mice, NLRP3 -/- mice exhibited higher sensitivity to oxazolone treatment with enhancement of Th2 cytokine expression and reduction of mature IL-1β and IL-18 production; this phenotype was rescued by exogenous IL-1β or IL-18. Immunofluorescent studies revealed positive correlation of NLRP3 expression with disease severity in UC patients, and localization of the inflammasome-associated molecules in macrophages. The NLRP3 inflammasome-derived IL-1β and IL-18 may play a protective role against UC through different mechanisms., Competing Interests: Toshio Watanabe, Tetsuya Tanigawa, Kazunari Tominaga and Yasuhiro Fujiwara – Faculty member of a course sponsored by Eisai Co., Ltd. The other authors have declared that no competing interests exist.

Allergic contact dermatitis is a chronic T cell-driven inflammatory skin disease that is caused by repeated exposure to contact allergens. Based on murine studies of acute contact hypersensitivity, mast cells (MCs) are believed to play a role in its pathogenesis. The role of MCs in chronic allergic contact dermatitis has not been investigated, in part because of the lack of murine models for chronic contact hypersensitivity. We developed and used a chronic contact hypersensitivity model in wild-type and MC-deficient mice and assessed skin inflammatory responses to identify and characterize the role of MCs in chronic allergic contact dermatitis. Ear swelling chronic contact hypersensitivity responses increased markedly, up to 4-fold, in MC-deficient Kit W-sh/W-sh (Sash) and MCPT5-Cre + iDTR + mice compared with wild-type mice. Local engraftment with MCs protected Sash mice from exacerbated ear swelling after repeated oxazolone challenge. Chronic contact hypersensitivity skin of Sash mice exhibited elevated levels of IFN-γ, IL-17α, and IL-23, as well as increased accumulation of Ag-specific IFN-γ-producing CD8 + tissue-resident memory T (T RM ) cells. The CD8 + T cell mitogen IL-15, which was increased in oxazolone-challenged skin of Sash mice during the accumulation of cutaneous T RM cells, was efficiently degraded by MCs in vitro. MCs protect from the exacerbated allergic skin inflammation induced by repeated allergen challenge, at least in part, via effects on CD8 + T RM cells. MCs may notably influence the course of chronic allergic contact dermatitis. A better understanding of their role and the underlying mechanisms may lead to better approaches for the treatment of this common, disabling, and costly condition., (Copyright © 2016 by The American Association of Immunologists, Inc.)

Conjugated linoleic acid (CLA) is one of the constituents of animal products with possible health benefits such as anti-carcinogenic and anti-obesity effects. In this study, we investigated the immunomodulatory effects of CLA using a mouse model of allergic dermatitis. Mice were orally administered either a CLA mixture containing equal amounts of 9c, 11 t-CLA and 10 t, 12c-CLA, or high linoleic acid safflower oil, and allergic dermatitis was induced on the ear by repeated topical applications of oxazolone. Oral administration of the CLA mixture but not the high linoleic safflower oil attenuated the symptoms of allergic dermatitis in both ear weights and clinical scores. This effect was associated with decreased levels of ear interleukin-4 (IL-4) and plasma immunoglobulin E. The immunomodulatory effects of the CLA isomers were compared by an in vitro cytokine production assay. The results showed that 9c, 11 t-CLA, the most predominant isomer in animal products, significantly inhibited IL-4 and interferon-γ production from mouse splenocytes with similar potency to 10 t, 12c-CLA. These findings suggest that CLA, a constituent of animal products, has a potentially beneficial effect for amelioration of allergic dermatitis., (© 2016 Japanese Society of Animal Science.)

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.

Inflammatory bowel diseases (IBDs) are chronic inflammatory gastrointestinal disorders caused by a dysregulated mucosal immune response and epithelial barrier disruption. Conventional treatment of IBD is currently limited to overcoming patient symptoms and is often associated with severe adverse effects from the drugs used. Modified Pulsatilla decoction has been used previously to treat ulcerative colitis (UC) in clinical practice in China, however, the underlying mechanism in the treatment of UC remains to be elucidated. In the present study, the efficiency and mechanisms of modified Pulsatilla decoction in the treatment of oxazolone‑induced colitis were investigated. Assessment of clinical colitis and histological examination found that the administration of modified Pulsatilla decoction attenuated the severity of oxazolone‑induced colitis in mice. Measurement of cytokine concentration, western blotting and reverse transcription‑quantitative polymerase chain reaction demonstrated modified Pulsatilla decoction treatment significantly reduced the secretion of pro‑inflammatory cytokines and restored alterations in tight junction proteins in the colon tissues. In addition, modified Pulsatilla decoction suppressed the activation of the nuclear factor‑κB signaling pathway. Thus, the findings of the present study demonstrated that modified Pulsatilla decoction offers an effective therapeutic approach for the treatment of IBD and revealed the underlying mechanisms of action offered by modified Pulsatilla decoction.

Atopic dermatitis (AD) is a common allergic skin disease, characterized by dryness, itchiness, thickening and inflammation of the skin. Infiltration of eosinophils into the dermal layer and presence of edema are typical characteristics in the skin biopsy of AD patients. Previous in vitro and clinical studies showed that the Pentaherbs formula (PHF) consisting of five traditional Chinese herbal medicines, Flos Lonicerae, Herba Menthae, Cortex Phellodendri, Cortex Moutan and Rhizoma Atractylodis at w/w ratio of 2:1:2:2:2 exhibited therapeutic potential in treating AD. In this study, an in vivo murine model with oxazolone (OXA)-mediated dermatitis was used to elucidate the efficacy of PHF. Active ingredients of PHF water extract were also identified and quantified, and their in vitro anti-inflammatory activities on pruritogenic cytokine IL-31- and alarmin IL-33-activated human eosinophils and dermal fibroblasts were evaluated. Ear swelling, epidermis thickening and eosinophils infiltration in epidermal and dermal layers, and the release of serum IL-12 of the murine OXA-mediated dermatitis were significantly reduced upon oral or topical treatment with PHF (all p < 0.05). Gallic acid, chlorogenic acid and berberine contents (w/w) in PHF were found to be 0.479%, 1.201% and 0.022%, respectively. Gallic acid and chlorogenic acid could suppress the release of pro-inflammatory cytokine IL-6 and chemokine CCL7 and CXCL8, respectively, in IL-31- and IL-33-treated eosinophils-dermal fibroblasts co-culture; while berberine could suppress the release of IL-6, CXCL8, CCL2 and CCL7 in the eosinophil culture and eosinophils-dermal fibroblasts co-culture (all p < 0.05). These findings suggest that PHF can ameliorate allergic inflammation and attenuate the activation of eosinophils.

Nerol is a natural monoterpene with antinociceptive and anti-inflammatory properties. Its possible beneficial effects in ulcerative colitis and its corresponding mechanism of action have not been determined to date. The aim of this study was to investigate whether nerol prevents the appearance of pathological markers and hyperalgesia in oxazolone-induced colitis, and protects against gastric damage produced by ethanol. The experimental design included groups of oxazolone-treated mice receiving nerol at 10-300 mg/kg, p.o., or a reference drug (sulfasalazine, 100 mg/kg, p.o.) compared to sham and untreated groups. Gastric damage was evaluated in the absolute ethanol-induced ulcer model in rats. Variables measured in animals with oxazolone-induced colitis included weight loss, stool consistency and macroscopic colon damage; mechanical nociception was determined by the use of von Frey filaments, whereas levels of inflammatory cytokines were assessed by enzyme-linked immunosorbent assay. Nerol (30-300 mg/kg, p.o.) prevented or significantly decreased the pathological alterations observed in the oxazolone- induced colitis model. It also showed antinociceptive effects and reduced the increased levels of inflammatory cytokines (IL-13 and TNF-α). Gastric damage was also prevented starting at 10 mg/kg, p.o. In conclusion, our results provide evidence for a beneficial effect of nerol after colitis induction involving tissue protection, antinociception and modulation of the immunological system, suggesting the therapeutic potential of this monoterpene as a novel alternative in controlling ulcerative colitis., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Murine models of intestinal inflammation have been widely used in biomedical research. Similarities in anatomy and physiology between such murine models and patients with inflammatory bowel diseases may allow a better understanding of the pathogenesis of Crohn's disease and ulcerative colitis. Additionally, models of intestinal inflammation may be used for the analysis of potentially new therapeutic agents. One key class of models consists of chemically induced inflammation models. Within this group, colitis induced by the haptenizing agent oxazolone is an important model that results in induction of acute or chronic inflammation of the large bowel. Here, we describe the induction and the analysis of this experimental colitis model.

Upon antigen/allergen recognition, epidermal Langerhans' cells (LC) are mobilized and migrate to the local lymph node where they play a major role in initiating or regulating immune responses. It had been proposed that all chemical allergens induce LC migration via common cytokine signals delivered by TNF-α and IL-1β. Here the dependence of LC migration on TNF-α following treatment of mice with various chemical allergens has been investigated. It was found that under standard conditions the allergens oxazolone, paraphenylene diamine, and trimellitic anhydride, in addition to the skin irritant sodium lauryl sulfate, were unable to trigger LC mobilization in the absence of TNF-α signalling. In contrast, two members of the dinitrohalobenezene family (2,4-dinitrochlorobenzene [DNCB] and 2,4-dinitrofluorobenzene [DNFB]) promoted LC migration independently of TNF-R2 (the sole TNF-α receptor expressed by LC) and TNF-α although the presence of IL-1β was still required. However, increasing doses of oxazolone overcame the requirement of TNF-α for LC mobilization, whereas lower doses of DNCB were still able to induce LC migration in a TNF-α-independent manner. These novel findings demonstrate unexpected heterogeneity among chemical allergens and furthermore that LC can be induced to migrate from the epidermis via different mechanisms that are either dependent or independent of TNF-α. Although the exact mechanisms with regard to the signals that activate LC have yet to be elucidated, these differences may translate into functional speciation that will likely impact on the extent and quality of allergic sensitization., (© 2014 John Wiley & Sons Ltd.)

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., (© 2014 British Society for Immunology.)

Interleukin-4 (IL-4) and IL-13 are critical drivers of immune activation and inflammation in ulcerative colitis, asthma and other diseases. Because these cytokines may have redundant function, dual targeting holds promise for achieving greater efficacy. We have recently described a bifunctional therapeutic targeting IL-4 and IL-13 developed on a novel protein scaffold, generated by combining specific binding domains in an optimal configuration using appropriate linker regions. In the current study, the bifunctional IL-4/IL-13 antagonist was evaluated in the murine oxazolone-induced colitis model, which produces disease with features of ulcerative colitis. The bifunctional IL-4/IL-13 antagonist reduced body weight loss throughout the 7-day course of the model, and ameliorated the increased colon weight and decreased colon length that accompany disease. Colon tissue gene expression was modulated in accordance with the treatment effect. Concentrations of serum amyloid P were elevated in proportion to disease severity, making it an effective biomarker. Serum concentrations of the bifunctional IL-4/IL-13 antagonist were inversely proportional to disease severity, colon tissue expression of pro-inflammatory genes, and serum amyloid P concentration. Taken together, these results define a panel of biomarkers signifying engagement of the IL-4/IL-13 pathway, confirm the T helper type 2 nature of disease in this model, and demonstrate the effectiveness of dual cytokine blockade., (© 2014 John Wiley & Sons Ltd.)

In inflammatory bowel disease (IBD), tissue damage is driven by an excessive immune response, poorly controlled by counter-regulatory mechanisms. SIRT1, a class III NAD+-dependent deacetylase, regulates negatively the expression of various proteins involved in the control of immune-inflammatory pathways, such as Stat3, Smad7, and NF-κB. Here we examined the expression, regulation, and function of SIRT1 in IBD. SIRT1 RNA and protein expression was less pronounced in whole biopsies and lamina propria mononuclear cells (LPMCs) of IBD patients in comparison with normal controls. SIRT1 expression was downregulated in control LPMC by tumor necrosis factor (TNF)-α and interleukin (IL)-21, and upregulated in IBD LPMC by neutralizing TNF-α and IL-21antibodies. Consistently, SIRT1 expression was increased in mucosal samples taken from IBD patients successfully treated with Infliximab. Treatment of IBD LPMC with Cay10591, a specific SIRT1 activator, reduced NF-κB activation and inhibited inflammatory cytokine synthesis, whereas Ex527, an inhibitor of SIRT1, increased interferon (IFN)-γ in control LPMC. SIRT1 was also reduced in mice with colitis induced by 2,4,6-trinitrobenzenesulphonic acid or oxazolone. Cay10591 prevented and cured experimental colitis whereas Ex527 exacerbated disease by modulating T cell-derived cytokine response. Data indicate that SIRT1 is downregulated in IBD patients and colitic mice and suggest that SIRT1 activation can help attenuate inflammatory signals in the gut.

Contact hypersensitivity (CHS) induced by topical application of haptens is a commonly used model to study dermal inflammatory responses in mice. Several recent studies have indicated that CHS-induced skin inflammation triggers lymphangiogenesis but may negatively impact the immune-function of lymphatic vessels, namely fluid drainage and dendritic cell (DC) migration to draining lymph nodes (dLNs). On the other hand, haptens have been shown to exert immune-stimulatory activity by inducing DC maturation. In this study we investigated how the presence of pre-established CHS-induced skin inflammation affects the induction of adaptive immunity in dLNs. Using a mouse model of oxazolone-induced skin inflammation we observed that lymphatic drainage was reduced and DC migration from skin to dLNs was partially compromised. At the same time, a significantly stronger adaptive immune response towards ovalbumin (OVA) was induced when immunization had occurred in CHS-inflamed skin as compared to uninflamed control skin. In fact, immunization with sterile OVA in CHS-inflamed skin evoked a delayed-type hypersensitivity (DTH) response comparable to the one induced by conventional immunization with OVA and adjuvant in uninflamed skin. Striking phenotypic and functional differences were observed when comparing DCs from LNs draining uninflamed or CHS-inflamed skin. DCs from LNs draining CHS-inflamed skin expressed higher levels of co-stimulatory molecules and MHC molecules, produced higher levels of the interleukin-12/23 p40 subunit (IL-12/23-p40) and more potently induced T cell activation in vitro. Immunization experiments revealed that blockade of IL-12/23-p40 during the priming phase partially reverted the CHS-induced enhancement of the adaptive immune response. Collectively, our findings indicate that CHS-induced skin inflammation generates an overall immune-stimulatory milieu, which outweighs the potentially suppressive effect of reduced lymphatic vessel function.

Atopic dermatitis (AD) is a refractory and recurrent inflammatory skin disease. Various factors including heredity, environmental agent, innate and acquired immunity, and skin barrier function participate in the pathogenesis of AD. T -helper (Th) 2-dominant immunological milieu has been suggested in the acute phase of AD. Antigen 85B (Ag85B) is a 30-kDa secretory protein well conserved in Mycobacterium species. Ag85B has strong Th1-type cytokine inducing activity, and is expected to ameliorate Th2 condition in allergic disease. To perform Ag85B function in vivo, effective and less invasive vaccination method is required. Recently, we have established a novel functional virus vector; recombinant human parainfluenza type 2 virus vector (rhPIV2): highly expressive, replication-deficient, and very low-pathogenic vector. In this study, we investigated the efficacy of rhPIV2 engineered to express Ag85B (rhPIV2/Ag85B) in a mouse AD model induced by repeated oxazolone (OX) challenge. Ear swelling, dermal cell infiltrations and serum IgE level were significantly suppressed in the rhPIV2/Ag85B treated mouse group accompanied with elevated IFN-γ and IL-10 mRNA expressions, and suppressed IL-4, TNF-α and MIP-2 mRNA expressions. The treated mice showed no clinical symptom of croup or systemic adverse reactions. The respiratory tract epithelium captured rhPIV2 effectively without remarkable cytotoxic effects. These results suggested that rhPIV2/Ag85B might be a potent therapeutic tool to control allergic disorders.

PGRN and its derived engineered protein, Atsttrin, were reported to antagonize TNFα and protect against inflammatory arthritis [Tang, W. et al. (2011) The growth factor progranulin binds to TNF receptors and is therapeutic against inflammatory arthritis in mice. Science 332 (6028) 478-484]. Here we found that PGRN level was also significantly elevated in skin inflammation. PGRN-/- mice exhibited more severe inflammation following induction of oxazolone (OXA). In contrast, recombinant Atsttrin protein effectively attenuated inflammation in mice dermatitis model. In addition, the protective role of PGRN and Atsttrin in dermatitis was probably due to their inhibition on NF-κB signaling. Collectively, PGRN, especially its derived engineered protein, Atsttrin, may represent a potential molecular target for prevention and treatment of inflammatory skin diseases., (Copyright © 2013 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.)

Patients suffering from ulcerative colitis (UC) exhibit chronic colonic inflammation caused by a dysregulated mucosal immune response and epithelial barrier disruption. Th2 cytokines, including IL-13, have been implicated in the pathogenesis of UC. IL-13 induces phosphorylation of STAT6, and we previously demonstrated increased epithelial p-STAT6 in children with UC. In this study, we investigated the role of STAT6 in oxazolone colitis, a murine model of UC, by inducing colitis in STAT6-deficient (STAT6(-/-)) and wild type (WT) mice. We observed increased epithelial cell, T cell, macrophage, and NKT cell STAT6 phosphorylation, as well as increased p-STAT6(+) IL-13-producing NKT cells, in colitic WT mice. Colitis was attenuated in STAT6(-/-) mice, with improvements in weight, colon length, and histopathology. There was decreased induction of the pore-forming tight junction protein claudin-2 in STAT6(-/-) mice. Similarly, short hairpin RNA STAT6 knockdown reduced claudin-2 induction and transepithelial resistance decrease in IL-13-treated human T84 cells. Tissue expression of IL-13, IFN-γ, IL-17, and IL-10 mRNA was similarly induced in WT and STAT6(-/-) colitic mice; however, we observed increased mRNA expression for the Th2-inducing cytokines IL-33 and thymic stromal lymphopoietin in WT mice with colitis, which was abrogated in STAT6(-/-) mice. Mesenteric lymph node cells from STAT6(-/-) mice with colitis exhibited reduced secretion of IL-4, IL-5, IL-13, and IFN-γ. IL-33 augmented mesenteric lymph node cell secretion of IL-5, IL-13, IL-6, and IFN-γ. These data implicate STAT6 in the pathogenesis of colitis in vivo with important roles in altering epithelial barrier function and regulating Th2-inducing cytokine production.

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.

A recent paper suggests that reduced exposure to germs results in the expansion of a cell type called natural killer T cells, which predisposes to colitis and asthma. Such a scenario could explain the Hygiene Hypothesis, which has been a puzzle for decades.

The recovery of skin barrier functions was investigated with pseudo-ceramide-based lipid microparticles. The microparticles were prepared by using a fluid bed technique where lipid components (a pseudo-ceramide, cholesterol and a fatty acid) were coated on a sugar seed, and a polymer was subsequently coated on the lipid microparticles. The microparticles contained large amount of pseudo-ceramide, and the pseudo-ceramide was in the form of lamellar structures mixed with other lipid components. In addition, the microparticles were stably dispersed in aqueous media or emulsion systems without any disruption of the microparticles' structures, thereby supplying sufficient amount of the pseudo-ceramide to skins for improving skin barrier functions such as preventing water loss. Such a role of the microparticles was proven by evaluating in vivo the efficacy of the lipid microparticles in reducing a trans-epidermal water loss (TEWL) of impaired murine skins. As a result, the novel pseudo-ceramide-based lipid microparticles for barrier recovery may potentially be applied in the field of dermatology, cosmetics and pharmaceuticals., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Among the various adverse effects of topical corticosteroids, impairment of the epidermal permeability barrier is well-known. Decreased synthesis of the epidermal lipid consequently leads to structural defects of the stratum corneum. Recently, the beneficial effects of physiologic lipid mixtures containing pseudoceramide on the impaired epidermal permeability barrier have been reported, which suggest that physiologic lipid mixtures may reduce the topical glucocorticoid-induced barrier impairment. In this study, the effect of a pseudoceramide-containing physiologic lipid mixture as a vehicle for a mid-potency topical glucocorticoid was evaluated in an oxazolone-induced atopic dermatitis-like murine model. The changes in transepidermal water loss, hydration and skin fold thickness were measured. Inflammatory cells in the dermis, including eosinophils, were counted and Staphylococcus aureus binding assay was performed. Immunohistochemical staining for inflammatory cytokines and antimicrobial peptides were also performed. The topical steroid in physiologic lipid mixture showed a significantly decreased infiltrate of inflammatory cells (p&#60;0.05) and a reduced number of adherent Staphylococcus aureus compared with the results of the topical steroid in polyethylene glycol/ethanol vehicle (p&#60;0.05). In conclusion, the pseudoceramide-containing physiologic lipid mixture as a vehicle for a topical steroid enhanced the anti-inflammatory effect of the topical steroid and accelerated the skin barrier function restoration.

Mitogen- and stress-activated protein kinase 1 and 2 (MSK1/2) are two kinases phosphorylated by both ERK1/2 and p38 MAPK. Recently, MSK1 and 2 have been reported to act as negative regulators of acute inflammation. In this study, we investigated the role of MSK1/2 in chronic skin inflammation using an oxazolone-induced allergic contact dermatitis model in MSK1/2 knockout mice and wild-type mice. MSK1/2 knockout mice were demonstrated to have significantly increased inflammation compared with wild-type mice. This was measured by an increased ear thickness, elevated infiltration of neutrophils in the skin and increased inflammatory histological changes. Furthermore, we found significantly elevated levels of the proinflammatory cytokines Tumor necrosis factor-α (TNF-α), IL-1β and IL-6 at both mRNA and protein levels in MSK1/2 knockout mice compared with wild-type mice after oxazolone treatment. In addition, the mRNA expression of the chemokine Thymus and activation regulated chemokine (TARC) was demonstrated to be significantly elevated in oxazolone-treated MSK1/2 knockout mice compared with wild-type mice. The increased expression of TARC was paralleled by increased infiltration of cells positive for the TARC receptor, CCR4, in the dermis of MSK1/2 knockout mice. Our results indicate that MSK1/2 are involved in the activation of feedback mechanisms that dampen oxazolone-induced skin inflammation., (© 2011 John Wiley & Sons A/S.)

Licochalcone, a constituent of licorice, has antitumor, antimicrobial, and anti-inflammatory effects. Recently, licochalcone E was isolated from the roots of Glycyrrhiza inflata and its biological functions are not fully examined. In this study, we investigated its ability to modulate production of IL-12p40, a common subunit of IL-12 and IL-23. Licochalcone E dose-dependently inhibited IL-12p40 production from lipopolysaccharide-stimulated RAW264.7 macrophage cells. The repressive effect was mapped to a region in the IL-12 gene promoter containing a binding site for NF-kappaB. Furthermore, licochalcone E decreased binding to the NF-kappaB site in RAW264.7 macrophage cells. Using a chronic allergic contact dermatitis model induced by repeated application of oxazolone, we showed that licochalcone E inhibited the increased IL-12p40 expression and ear thickness induced by oxazolone. Taken together, licochalcone E inhibits IL-12p40 production and has therapeutic potential to reduce skin inflammation., ((c) 2010 Elsevier B.V. All rights reserved.)

Cathepsin S (CATS) is a cysteine protease, well known for its role in MHC class II-mediated antigen presentation and extracellular matrix degradation. Disturbance of the expression or metabolism of this protease is a concomitant feature of several diseases. Given this importance we studied the localization and regulation of CATS expression in normal and pathological human/mouse skin. In normal human skin CATS-immunostaining is mainly present in the dermis and is localized in macrophages, Langerhans, T- and endothelial cells, but absent in keratinocytes. In all analyzed pathological skin biopsies, i.e. atopic dermatitis, actinic keratosis and psoriasis, CATS staining is strongly increased in the dermis. But only in psoriasis, CATS-immunostaining is also detectable in keratinocytes. We show that cocultivation with T-cells as well as treatment with cytokines can trigger expression and secretion of CATS, which is involved in MHC II processing in keratinocytes. Our data provide first evidence that CATS expression (i) is selectively induced in psoriatic keratinocytes, (ii) is triggered by T-cells and (iii) might be involved in keratinocytic MHC class II expression, the processing of the MHC class II-associated invariant chain and remodeling of the extracellular matrix. This paper expands our knowledge on the important role of keratinocytes in dermatological disease.

IL-1 is a prototypic inflammatory cytokine that has pathogenic roles in various skin disorders. Although Langerhans cells (LCs) have been reported to express IL-1beta mRNA upon application of contact sensitizers, it remains unclear whether other cell types produce IL-1beta in skin. Thus, we sought to directly identify IL-1beta-producing cells in living animals by construction of transgenic mice expressing DsRed fluorescence protein gene under the control of IL-1beta promoter. Little DsRed fluorescence signal was detected in skin under steady-state conditions. Striking increases in DsRed signal were observed after topical application of a contact sensitizer, oxazolone, which also induced markedly elevated IL-1beta mRNA and protein expression. DsRed signal was expressed primarily by CD45(+)/CD11b(+) myeloid leukocytes in both epidermal and dermal compartments and was detected only in small fractions of epidermal LCs. Interestingly, DsRed(+) cells emerged preferentially as clusters around hair follicles. Intravital confocal imaging experiments revealed highly motile potentials of DsRed(+) cells-they constantly crawled around hair follicles via amoeba-like movements with a mean velocity of 1.0+/-0.4 microm min(-1) (epidermis) or 2.7+/-1.4 microm min(-1) (dermis). The newly developed in vivo imaging system represents a useful tool for studying spatial regulation of IL-1beta production in skin.

Objective: To investigate the therapeutic effects of Bawei Xilei San (BWXLS), a compound traditional Chinese herbal medicine, on mice with oxazolone-induced colitis and to explore the mechanisms., Methods: Thirty-two BALB/c mice were randomly divided into 4 groups (8 for each): normal control group, untreated group, hydrocortisone group and BWXLS group. Except for the mice in the normal control group, all mice were intrarectally administered with 3.0% oxazolone to induce colitis. Then the mice in the normal control group and untreated group were administered with 0.9% carboxymethyl cellulose sodium solution. Mice in the BWXLS group were intrarectally administered with 0.2 mg/g BWXLS and hydrocortisone group with 0.02 mg/g respectively for 5 days. The body weight and stool consistency and occult or gross blood were recorded to calculate the disease activity index (DAI). The mice were sacrificed at the 6th day. The macroscopic and histological changes of the colon were evaluated. The expressions of Toll-like receptor 4 (TLR4), nuclear factor-kappaB (NF-kappaB), and epithelial tight junction protein occludin were assessed by immunohistochemical method. The level of tumor necrosis factor-alpha (TNF-alpha) in colonic mucosa was evaluated by enzyme-linked immunosorbent assay., Results: The DAI, and macroscopic and histological changes in the BWXLS group were improved as compared with those in the untreated group (P<0.05) but were similar to those in the hydrocortisone group. The expression of occludin was significantly increased (P<0.05) while the expressions of TLR4, NF-kappaB and TNF-alpha were significantly decreased in the BWXLS group as compared with the untreated group, and were similar to those in the hydrocortisone group (P>0.05)., Conclusion: Up-regulating the expression of occludin and down-regulating the expressions of TLR4 and NF-kappaB, and hence inhibiting TNF-alpha expression and improving the mucosa barrier function may be part of the mechanisms of BWXLS in treating oxazolone-induced colitis in mice.

Aim: To evaluate the therapeutic effects of a probiotic supplement (Clostridium butyricum, CGMCC0313) in a chemically-induced rat model of experimental colitis., Methods: An experimental ulcerative colitis model was established by rectal injection of oxazolone into the colon of 40 Wistar rats randomly divided into four groups. The positive control group was sacrificed 3 d after colitis onset. The remaining groups were fed daily with either 2 mL of C. butyricum (2.3 x 10(11) CFU/L), 2 mL of mesalamine (100 g/L), or 1 mL of sodium butyrate (50 mmol/L) for 21 d. The animals' body weight, behavior, and bowel movements were recorded weekly. After sacrifice, visual and microscopic observations of pathological changes of colon tissue were made, body weight and wet colon mass index were measured and recorded, and serum levels of interleukin-23 (IL-23) and TNF-alpha were measured using ELISA. Expression of calcitonin gene-related peptide in colon tissue was measured by RT-PCR. Finally, changes in rat intestinal microflora status were measured in all groups., Results: We found that treatment with C. butyricum lowered the serum levels of both IL-23 and tumor necrosis factor-alpha (TNF-alpha) with similar or even better efficiency than that of mesalamine or sodium butyrate. The rat intestinal flora appeared to recover more quickly in the group treated with C. butyricum than in the mesalamine and sodium butyrate groups. Finally, we found that the expression level of calcitonin gene related peptide was elevated in colon tissue in the sodium butyrate treated group but not in the C. butyricum or mesalamine treated groups, indicating a sensitization of colon following sodium butyrate treatment., Conclusion: In our experimental colitis model, treatment with C. butyricum CGMCC0313, a probiotic supplement, is at least as efficient as treatment with mesalamine.

Increase in the number of patients with atopic dermatitis (AD) has been recently reported. T helper (Th) cells that infiltrate AD skin lesions are Th2-type dominant; reduced exposure to environmental Th1-cytokine-inducing microbes is believed to contribute to the increased number of AD patients. Regulatory type immune responses have been also associated with the occurrence of AD. It has been reported that antigen 85B (Ag85B) purified from mycobacteria is a potent inducer of Th1-type immune response in mice as well as in humans. In this study, we have examined the effect of plasmid DNA encoding Ag85B derived from Mycobacterium kansasii on AD skin lesions induced by oxazolone (OX) application. Th2-cytokine mediated mouse AD model with immediate type response followed by a late phase reaction was developed by repeated applications of low-dose OX to sensitized mice. Mice were immunized with plasmid DNA encoding cDNA of Ag85B before OX sensitization or during repeated elicitation phase. Both therapies were associated with significant suppression of immediate type response, clinical appearance, dermal cell infiltration, reduced IL-4 production, and augmented IFN-gamma mRNA expression compared to placebo-treated mice. Additionally, increased number of Foxp3(+) regulatory T cells were observed in the skin sections in Ag85B treated mice. The results of this study suggest that Ag85B DNA vaccine is a potential therapy for Th2 type dermatitis.

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.

Background: An extract of the tropical fern Polypodium leucotomos (PL) administered orally to mice inhibits ultraviolet B (UVB) radiation-induced skin cancer formation. UVB-induced murine skin cancers occur, in part, because of UVB-induced immunosuppression. Thus, we examined whether PL inhibits UVB-suppression of the induction of contact hypersensitivity (CHS) locally or systemically., Methods: C57BL/6 mice received standard drinking water or water-containing PL. In the local model, mice were shaved on the dorsum and exposed to 3500 J/m(2) of UVB radiation daily for 4 days. Control mice were not irradiated. After the last irradiation they were sensitized to oxazolone topically at the irradiated site. To examine the ability of PL to inhibit systemic UVB-induced immunosuppression, mice were given 10,000 J/m(2) of UVB radiation once and immunized at a non-exposed site 3 days later. Six days after immunization (in both models), mice were challenged on the ears with oxazolone and 24/48 h ear swelling assessed., Results: PL in drinking water significantly reduced the inhibition of CHS observed with exposure to UVB radiation in both the local and systemic models., Conclusions: The ability of PL to inhibit UVB radiation-induced immune suppression may explain, in part, its ability to inhibit UVR-induced skin cancer induction in mice.

The sphingosine-1-phosphate analogue FTY720 is known to alter migration and homing of lymphocytes via sphingosine-1-phosphate receptors. However, several studies indicate that its mode of action is more complex and that FTY720 may also directly influence cytokine effector functions. Therefore, we studied the effect of FTY720 in T helper type (Th2)-mediated oxazolone-induced colitis in mice. Following rectal oxazolone instillation, Th2 cells producing IL-13 induce a progressive colitis resembling human ulcerative colitis. A rectal enema of oxazolone [90 mg/kg body weight] was applied to BALB/c mice. FTY720 was administered i.p. from day 0 to 3 or from day 3 to 5 following the instillation of the haptenating agent. Assessment of severity of colitis was performed daily. FTY720 plasma levels were detected using LC-MS/MS-analysis. Colon tissue was analyzed macroscopically and microscopically, myeloperoxidase activity as well as cytokine levels of lamina propria CD4(+) T-cells and T1/ST2 expression were determined. Treatment with FTY720 prominently reduced the clinical and histopathologic severity of oxazolone-induced colitis, abrogating body weight loss, diarrhea, and macroscopic and microscopic intestinal inflammation. The therapeutic effects of FTY720 were associated with a prominent reduction of the key effector Th2 cytokines IL-13, IL-4 and IL-5. Strikingly, FTY720 inhibited GATA3 and T1/ST2 expression which represent highly relevant markers for Th2 differentiation and Th2 effector function, respectively. Our data provide the first evidence that FTY720 exhibits beneficial prophylactic as well as therapeutic effects in Th2-mediated experimental colitis by directly affecting Th2 cytokine profiles probably by reducing T1/ST2, thus offering a new auspicious therapeutic instrument for the treatment of human ulcerative colitis.

Glycyrrhizin (18beta-glycyrrhetinic acid-3-O-beta-D-glucuronopyranosyl-(1 --> 2)-beta-D-glucuronide, GL) was transformed to 18beta-glycyrrhetinic acid-3-O-beta-D-glucuronide (GAMG) by Streptococcus LJ-22. The antiallergic activities of GL and GAMG was measured using a RBL cell assay system and contact hypersensitivity model mice. GAMG exhibited anti-allergic activity with IC50 values of 0.28 mM. GAMG, which is sweeter than GL, and 18beta-glycyrrhetinic acid, which is a GAMG metabolite by human intestinal bacteria, also inhibited the passive cutaneous anaphylaxis and skin contact inflammation. In conclusion, GAMG may be useful as a new sweet food additive and an anti-allergic agent.

Objective: 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., Methods: 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., Results: 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., Conclusion: 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.

To investigate the involvement of protease-activated receptor-2 (PAR-2) in allergic dermatitis, we generated PAR-2-deficient (PAR-2(-/-)) mice. Ear thickness, contact hypersensitivity (CH) induced by topical application of picryl chloride (PC) or oxazolone (Ox) after sensitization, and vascular permeability after ear passive cutaneous anaphylaxis (PCA) were compared between wild-type (WT) and PAR-2(-/-) mice. Ear thickness was almost the same in untreated WT and PAR-2(-/-) mice. Topical application of PC or Ox thickened the ears at 6, 24 and 48 h after challenge with a peak at 24 h in WT mice. In PAR-2(-/-) mice, the ear swelling induced by both PC and Ox was suppressed at every time point, and significant inhibition was found at 24 h in PC-induced CH and at 24 and 48 h in Ox-induced CH. Histopathological observation of the ears at 24 h after challenge revealed that PC- or Ox-induced ear edema and infiltration of inflammatory cells in WT mice were greatly attenuated in PAR-2(-/-) mice. The vascular permeability in the ears after PCA was not different between WT and PAR-2(-/-) mice. These results strongly suggest that PAR-2 plays a crucial role in type IV allergic dermatitis but not in type I allergic dermatitis.

P-selectin glycoprotein ligand 1 (PSGL-1) is a sialomucin expressed on leukocytes that mediates neutrophil rolling on the vascular endothelium. Here, the role of PSGL-1 in mediating lymphocyte migration was studied using mice lacking PSGL-1. In a contact hypersensitivity model, the infiltration of CD4(+) T lymphocytes into the inflamed skin was reduced in PSGL-1-deficient mice. In vitro-generated T helper (Th)1 cells from PSGL-1-deficient mice did not bind to P-selectin and migrated less efficiently into the inflamed skin than wild-type Th1 cells. To assess the role of PSGL-1 in P- or E-selectin-mediated migration of Th1 cells, the cells were injected into E- or P-selectin-deficient mice. PSGL-1-deficient Th1 cells did not migrate into the inflamed skin of E-selectin-deficient mice, indicating that PSGL-1 on Th1 cells is the sole ligand for P-selectin in vivo. In contrast, PSGL-1-deficient Th1 cells migrated into the inflamed skin of P-selectin-deficient mice, although less efficiently than wild-type Th1 cells. This E-selectin-mediated migration of PSGL-1-deficient or wild-type Th1 cells was not altered by injecting a blocking antibody to L-selectin. These data provide evidence that PSGL-1 on Th1 cells functions as one of the E-selectin ligands in vivo.

The role of interleukin-4 as a regulator of immune responses in the skin is investigated with regard to the outcome of contact hypersensitivity reaction in interleukin-4-deficient BALB/C mice. In previous studies conflicting results were obtained concerning the role of interleukin-4 in contact hypersensitivity reactions supporting either a proinflammatory or rather an inhibitory function of this cytokine. Interleukin-4 deficient BALB/C mice sensitized to 2,4-dinitrochlorobenzene showed after challenge a significant reduction in magnitude and duration of the contact hypersensitivity response in comparison with wild-type mice. This attenuation was accompanied by a significant reduction of edema and cellular infiltrates in the dermis and a lacking induction of IL-10 mRNA expression in skin. Also, adoptive transfer experiments revealed that BALB/C mice failed to exhibit contact hypersensitivity after injection of lymph node cells obtained from sensitized interleukin-4 deficient mice. To examine further the role of the contact allergen used to induce the contact hypersensitivity response, mice were also sensitized and challenged with Oxazolone. Here a similar magnitude and duration of contact hypersensitivity in both the interleukin-4 deficient mice and BALB/C control mice was observed. This indicates that the contact hypersensitivity response to 2,4-dinitrochlorobenzene and Oxazolone may partly evolve on different pathways being dependent and independent of interleukin-4. Our results clearly show that the complete loss of endogenous interleukin-4 expression in BALB/C mice is associated with an impaired manifestation of contact hypersensitivity response to 2,4-dinitrochlorobenzene, implying an important proinflammatory function of this cytokine.