| Atopic dermatitis is a common allergic,inflammatory,chronic and recurrent skin disease. In some AD patients after repeated use of topical GC, the effectiveness of GC is reduced, named “glucocorticoids tolerance” or “glucocorticoid resistance”. This phenomenon often occurs in the application of GC in other diseases such as asthma. The anti-inflammatory effect is produced through the activation of anti-inflammatory genes While GRα combined with GC being transferred to cell nucleus. GRβ acts as negative factor to inhibit the activity of GRα by specific binding with GRα, and reduces the anti-inflammatory effect.The colonization of staphylococcus aureus and superantigen(SEB) can be detected in local AD lesions. Currently, after SEB application to nasal or airway epithelial cells, “glucocorticoids tolerance” or “ glucocorticoid resistance” can be induced through several ways. It is confirmed that “glucocorticoid resistance” of rhinitis and asthma was caused by SEB, through upregulating GRβ. Whether the phenomenon that glucocorticoid resistance in skin disease is related with the upregulated expression of GRβ needs to be confirmed. H&E staining, immunohistochemistry, qRT-PCR and western blot were used to detect the influence of the expressions of GRs in BALB/c mice model, applied with SEB, dexamethasone(DEX) and SEB+DEX treatment.Objective:1. To construct AD animal model with DNCB.2. To explore the effect of SEB on the expression of GRα and GRβ in dermatitisMethod:Firstly, we summarized the experience of AD animal models, combining with the experimental requirements in animal model, and selected 1% DNCB, 2% DNCB. AD animal models were constructed respectively to observe and record physiological indicators of mice, dermatitis appearance, histopathology change to select the appropriate construction method of AD animal model for further study.Secondly, AD animal models were constructed by repeatedly stimulation of BALB/c mice back skin with 1% DNCB, then treated with 5μg SEB, 5μg SEB+0.03% DEX and 0.03% DEX respectively, taking the untreated mice model as the control. H&E staining was used to observe the pathological changes through counting the number of inflammatory cells in the dermis to reflect the dermatitis changes of different treated group. The qRT-PCR,Western blot and immunohistochemistry were chosen to detect the mRNA and protein expression level of GRα and GRβ.Results1. Applied with 1% DNCB as dermatitis-inducing agents,the mice skin mainly appears erythema, edema, scaling, rough and infiltration hypertrophy, without erosions, ulcers and scar. H&E staining showed that the cutaneous structure of AD model constructed with 1% DNCB was obvious, which is characterized by epidermal and dermal infiltration hypertrophy and infiltration of inflammatory cells in dermis, in line with the AD-like lesions and pathological changes.2. Histopathological changes: Compared with untreated AD mice, the number of the inflammatory cells treated with SEB was significantly increased, making inflammation worse. DEX played an anti-inflammatory effects, which was decreased by SEB. In addition, compared with untreated group, the expression level of GRβ was significantly increased in the dermatitis group treated with SEB(p<0.0001),and the expression was even significantly higher(P<0.05)in the SEB+DEX group.Conclusions1. The AD animal model is constructed stablely by repeated stimulation on BALB/c mice back skin with 1% DNCB, with the main characters of the dermatitis including erythema, edema, scaling, rough and epidermal hyperplasia. Histopathology shows obvious organizational structure and intercellular structured, which is suitable to observe the expression and distribution of target protein in tissues and cells by immunohistochemistry for further requirement.2. SEB upregulates the expression of GRβ in dermatitis, and plays an important role in topical GC “glucocorticoid resistance”. |
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