Lycii radicis cortex inhibits glucocorticoid­induced bone loss by downregulating Runx2 and BMP­2 expression.

Lycii radicis cortex (LRC) has been used to regulate high blood pressure, body temperature, pain and bone disorders in East Asia. Glucocorticoids (GCs), also known as steroids, are potent immunity regulators widely used in the treatment of inflammatory diseases. However, despite their effectiveness, GC usage is strictly controlled due to severe side­effects, such as osteoporosis. However, further research is required as to date, at least to the best of our knowledge, there is no appropriate model to overcome secondary osteoporosis as a side­effect of GC use. Thus, the aim of the present study was to establish an experimental model of osteoporosis induced by GC. Furthermore, the present study aimed to establish the research methodology for medical evaluations of the effectiveness and side­effects of GCs. A secondary osteoporosis animal model was established, and the animals were divided into two groups as follows: The allergic contact dermatitis (ACD)­induced group and the non­ACD­induced group. In the ACD­induced group, a GC topical application group was compared with a GC subcutaneous injection group. The results revealed that the presence of ACD affected the induction of GC­mediated osteoporosis. Therefore, the group exhibiting induced ACD that was treated with a topical application of GC was selected for examining the side­effects of GCs. The effects of LRC on secondary osteoporosis were confirmed in vivo and in vitro. The results indicated that LRC regulated dexamethasone­induced osteoblast apoptotic markers, including caspase­6, caspase­9, X­linked inhibitor of apoptosis, apoptosis inhibitor 1 and apoptosis inhibitor 2, and increased the expression of osteoblast differentiation­related genes, such as Runt­related transcription factor 2 and bone morphogenetic protein 2 in the MC3T3E­1 cell line. LRC also significantly reduced GC­induced osteoporosis and exerted anti­inflammatory effects in vivo. In addition, LRC inhibited the reduction of calbindin­D28k in the kidney. Overall, the results of the present study suggest that the use of LRC alleviates GC­induced secondary osteoporosis.

Immunosuppressive effect of hispidulin in allergic contact dermatitis.

BACKGROUND: Long-term use of most immunosuppressants to treat allergic contact dermatitis (ACD) generates unavoidable severe side effects, warranting discovery or development of new immunosuppressants with good efficacy and low toxicity is urgently needed to treat this condition. Hispidulin, a flavonoid compound that can be delivered topically due to its favorable skin penetrability properties, has recently been reported to possess anti-inflammatory and immunosuppressive properties. However, no studies have investigated the effect of hispidulin on Th1 cell activities in an ACD setting. METHODS: A contact hypersensitivity (CHS) mouse model was designed to simulate human ACD. The immunosuppressive effect of hispidulin was investigated via ear thickness, histologic changes (i.e., edema and spongiosis), and interferon-gamma (IFN-γ) gene expression in 1-fluoro-2,4-dinitrobenzene (DNFB)-sensitized mice. Cytotoxicity, total number of CD4+ T cells, and percentage of IFN-γ-producing CD4+ T cells were also investigated in vitro using isolated CD4+ T cells from murine spleens. RESULTS: Topically applied hispidulin effectively inhibited ear swelling (as measured by reduction in ear thickness), and reduced spongiosis, IFN-γ gene expression, and the number of infiltrated immune cells. The inhibitory effect of hispidulin was observed within 6 h after the challenge, and the observed effects were similar to those effectuated after dexamethasone administration. Hispidulin at a concentration up to 50 µM also suppressed IFN-γ-producing CD4+ T cells in a dose-dependent manner without inducing cell death, and without a change in total frequencies of CD4+ T cells among different concentration groups. CONCLUSION: The results of this study, therefore, suggest hispidulin as a novel compound for the treatment of ACD via the suppression of IFN-γ production in Th1 cells.

Electroacupuncture inhibits mast cell degranulation via cannabinoid CB2 receptors in a rat model of allergic contact dermatitis.

OBJECTIVE: Cannabinoid CB2 receptors (CB2Rs) are mainly present on immune cells including mast cells, which participate in 2,4-dinitrofluorobenzene (DNFB)-induced allergic contact dermatitis (ACD). In this study, we aimed to investigate whether inhibition of mast cell degranulation was involved in the anti-ACD effect of electroacupuncture (EA) at ST36 via CB2R. METHODS: Sprague-Dawley rats were sensitised and challenged with DNFB following EA stimulation for 1 week. Ear swelling, serum IgE levels, local cytokine production and mast cell infiltration were evaluated. Additionally, rat peritoneal mast cells (RPMCs) were isolated and cultured for detection of CB2R expression, mitogen-activated protein kinase (MAPK) signalling activation and mast cell degranulation (including ß-hexosaminidase and histamine release) in the presence or absence of CB2R antagonists. RESULTS: EA treatment inhibited ear swelling, suppressed IgE and cytokine production, decreased the number of mast cells and curbed mast cell degranulation, which was associated with the inhibition of p38 phosphorylation in DNFB-induced ACD. Importantly, EA enhanced the expression of CB2R mRNA and protein in the RPMCs. CB2R antagonist AM630 but not CB1R antagonist AM251 effectively reversed the suppressive effect of EA on p38 activation, mast cell infiltration and degranulation. CONCLUSION: These findings provide more evidence to support the hypothesis that EA promotes CB2R expression in mast cells, which is followed by inhibition of the p38 MAPK pathway, potentially resulting in the anti-ACD effect of EA. This suggests that EA at ST36 may be an effective candidate therapy for treating inflammatory skin diseases such as ACD.

Calycosin alleviates allergic contact dermatitis by repairing epithelial tight junctions via down-regulating HIF-1α.

Calycosin, a bioactive component derived from Astragali Radix (AR; Huang Qi), has been shown to have an effect of anti-allergic dermatitis with unknown mechanism. This study aims to investigate the mechanism of calycosin related to tight junctions (TJs) and HIF-1α both in FITC-induced mice allergic contact dermatitis and in IL-1ß stimulated HaCaT keratinocytes. Th2 cytokines (IL-4, IL-5 and IL-13) were detected by ELISA. The epithelial TJ proteins (occludin, CLDN1 and ZO-1), initiative key cytokines (TSLP and IL-33) and HIF-1α were assessed by Western blot, real-time PCR, immunohistochemistry or immunofluorescence. Herein, we have demonstrated that allergic inflammation and the Th2 cytokines in ACD mice were reduced significantly by calycosin treatment. Meanwhile, calycosin obviously decreased the expression of HIF-1α and repaired TJs both in vivo and in vitro. In HaCaT keratinocytes, we noted that IL-1ß induced the deterioration of TJs, as well as the increased levels of TSLP and IL-33, which could be reversed by silencing HIF-1α. In addition, administration of 2-methoxyestradiolin (2-ME), a HIF-1α inhibitor,significantly repaired the TJs and alleviated the allergic inflammation in vivo. Furthermore, TJs were destroyed by DMOG or by overexpressing HIF-1α in HaCaT keratinocytes, and simultaneously, calycosin down-regulated the expression of HIF-1α and repaired the TJs in this process. These results revealed that calycosin may act as a potential anti-allergy and barrier-repair agent via regulating HIF-1α in AD and suggested that HIF-1α and TJs might be possible therapy targets for allergic dermatitis.

GPR91 deficiency exacerbates allergic contact dermatitis while reducing arthritic disease in mice.

BACKGROUND: Succinate, in addition to its role as an intermediary of the citric acid cycle, acts as an alarmin, initiating and propagating danger signals resulting from tissue injury or inflammatory stimuli. The contribution of this immune sensing pathway to the development of allergic and inflammatory responses is unknown. METHODS: Ear thickness of wild-type (wt) and Sucnr1-deficient (Sucnr1-/- ) mice, sensitized and challenged with oxazolone, was used as a criterion to assess the relevance of SUCNR1/GPR91 expression mediating allergic contact dermatitis (ACD). Results obtained in this system were contrasted with data generated using passive cutaneous anaphylaxis, ovalbumin-induced asthma and arthritis models. RESULTS: We found augmented ACD reactions in Sucnr1-/- mice. This observation correlated with increased mast cell activation in vitro and in vivo. However, exacerbated mast cell activation in Sucnr1-/- mice did not contribute to the enhancement of asthma or arthritis and seemed to be due to alterations during mast cell development as augmented mast cell responses could be recapitulated in wt mast cells differentiated in the absence of succinate. CONCLUSIONS: A deficiency in succinate sensing during mast cell development confers these cells with a hyperactive phenotype. Such a phenomenon does not translate into exacerbation of asthma or mast cell-dependent arthritis. On the contrary, the fact that Sucnr1-/- mice developed reduced arthritic disease, using two different in vivo models, indicates that GPR91 antagonists may have therapeutic potential for the treatment of allergic and autoimmune diseases.

Genetic endowment of proinflammatory cytokine tumor necrosis factor-α (-) 308 G > A polymorphism to Parthenium hysterophorus-induced airborne contact dermatitis in an Indian cohort.

BACKGROUND: Parthenium dermatitis is a common airborne allergic health problem that induces a cell-mediated hypersensitivity immune response involving activated T lymphocytes, which culminates in injury to the skin. The disease is manifested as itchy erythematous papules and plaques and primarily affects the exposed areas and flexures. This study aimed to identify the role of tumor necrosis factor (TNF)-α (-) 308 G>A polymorphism in the pathogenesis of parthenium dermatitis. MATERIALS AND METHODS: A total of 120 subjects, including 60 patients exclusively diagnosed for parthenium dermatitis and 60 healthy individuals, were included in the study. The genotyping of the TNF-α (-) 308 G>A region was carried out by the amplification refractory mutational system. RESULTS: In the present study, we demonstrated that polymorphism of the TNF-α (-) 308 position (A and/or G) was not statistically significant, and there was no difference in the distribution of any alleles of this locus in cases and controls. CONCLUSION: The present study suggests that there is a lack of association of potent proinflammatory cytokine TNF-α (-) 308 G>A polymorphism in parthenium dermatitis in the Indian cohort. It interprets genetically endowed transcriptional capacity due to this particular single nucleotide polymorphism but does not support the prevalence of high serum levels of TNF-α in parthenium-induced skin allergic inflammation.

Modulation of cutaneous inflammation by angiotensin-converting enzyme.

Cutaneous neurogenic inflammation is a complex biological response of the host immune system to noxious stimuli. Present evidence suggests that zinc metalloproteases may play an important role in the regulation of neurogenic inflammation by controlling the local availability of neuropeptides, such as substance P (SP), that are capable of initiating or amplifying cutaneous inflammation after release from sensory nerves. To address the hypothesis that the dipeptidyl carboxypeptidase angiotensin-converting enzyme (ACE) is capable of modulating skin inflammation, we have analyzed murine allergic contact dermatitis (ACD) and irritant contact dermatitis (ICD) using wild-type C57BL/6J (ACE(+/+)) or genetically engineered mice with a heterozygous deletion of somatic ACE (ACE(+/-)). In 2,4-dinitro-1-fluorobenzene-sensitized ACE(+/-) mice, ACD was significantly augmented in comparison to ACE(+/+) controls as determined by the degree of ear swelling after exposure to hapten. Likewise, systemic treatment of ACE(+/+) mice with the ACE inhibitor captopril before sensitization or elicitation of ACD significantly augmented the ACD response. In contrast, local damage and neuropeptide depletion of sensory nerves following capsaicin, injection of a bradykinin B(2), or a SP receptor antagonist before sensitization significantly inhibited the augmented effector phase of ACD in mice with functionally absent ACE. However, in contrast to ACD, the response to the irritant croton oil was not significantly altered in ACE(+/-) compared with ACE(+/+) mice. Thus, ACE by degrading bradykinin and SP significantly controls cutaneous inflammatory responses to allergens but not to irritants, which may explain the frequently observed exacerbation of inflammatory skin disease in patients under medication with ACE inhibitors.

Neutral endopeptidase terminates substance P-induced inflammation in allergic contact dermatitis.

Sensory nerve-derived neuropeptides such as substance P demonstrate a number of proinflammatory bioactivities, but less is known about their role in inflammatory skin disease. The cell surface metalloprotease neutral endopeptidase (NEP) is the principal proteolytic substance P-degrading enzyme. This study tests the hypothesis that the absence of NEP results in dysregulated inflammatory skin responses. The effector phase of allergic contact dermatitis (ACD) responses was examined in NEP(-/-) knockout and NEP(+/+) wild-type mice and compared with the irritant contact dermatitis response in these animals. NEP was found to be normally immunolocalized in epidermal keratinocytes and dermal blood vessels. The ACD ear swelling response was 2.5-fold higher in animals lacking NEP and was accompanied by a significant increase in plasma extravasation and infiltration of inflammatory leukocytes. The augmented ACD response in NEP(-/-) animals was abrogated by either administration of a neurokinin receptor 1 antagonist or by repeated pretreatment with topical capsaicin. Similar to NEP(-/-) mice, the acute inhibition of NEP in NEP(+/+) animals resulted in an augmented ACD response. In contrast to the ACD responses, little differences were observed in the irritant contact dermatitis response of NEP(-/-) compared with NEP(+/+) animals after epicutaneous application of the skin irritants croton oil or SDS. Thus, these results indicate that NEP and cutaneous neuropeptides have a significant role in the pathogenesis of ACD.

Quantitative analysis of T-cell receptor beta variable-gene usage in cutaneous late-phase reactions: implications for T-lymphocyte recruitment in cutaneous inflammation.

To determine if functionally distinct T-lymphocyte (T cell) subsets accumulate in late-phase immunoglobulin E-mediated reactions (LPR), we quantitatively analyzed the immunophenotype and the T-cell receptor beta variable-gene (Vbeta) repertoire of T cells in cutaneous LPR. Peripheral blood and skin biopsies were obtained 6 or 24 h after sensitive subjects were challenged with intradermal injections of grass pollen allergen (Ag) and control (C) solution. The frequency of cells expressing CD3, CD4, CD8, CD45RO, and CD25/mm2 was determined by immunohistochemistry in nine subjects. Vbeta usage was assessed by reverse transcription-PCR in five of nine subjects. A significantly greater frequency of CD3(+) and CD45RO+ (memory) T cells was detected in Ag sites than in C sites at 24 h after challenge but not at 6 h. The frequency of activated (CD25(+)) and helper (CD4(+)) T cells appeared to be increased in Ag sites as well, though not significantly. Vbeta6 was the most commonly expressed Vbeta detected in Ag sites, but it was also detected in accompanying C sites. Vbeta2 was the most commonly expressed Vbeta detected in C sites. Sequence analysis in one case revealed Vbeta expression in a 6-h Ag site to be essentially polyclonal. Our findings suggest that memory T cells with Vbeta expression similar to that in normal skin accumulate in developing cutaneous LPR. The limited usage of Vbeta suggests a preferential recruitment or retention of reactive T cells from an endogenous subset of skin-homing T cells with its own skewed Vbeta repertoire.