Effect Of Staphylococcal Enterotoxin B Inhibiting Glucocorticoid Receptor Alpha Nuclear Translocation In Keratinocytes Of Allergic Contact Dermatitis

Background and ObjectiveTopical glucocorticoid(GC)as a treatment of various skin diseases has a long history.However,the GC curative effect attenuated after prolonged topical GC therapy in many patients with inflammatory skin disease.The phenomenon is known as GC "tachyphylaxis" or "acute tolerance".Knowledge of the molecular mechanisms responsible for GC-resistance or insensitivity is crucial for the improvement of effective therapies.Colonization of AD skin by S.aureus is increased in psoriasis,atopic dermatitis,which often need long-term GC therapy.Moreover,the exotoxins secreted by S.aureus are superantigens,such as SEB.They bind major histocompatibility complex(MHC)class-II molecules without any prior processing and specifically induce a polyclonal T lymphocyte activation(up to 20% of all T cells).Activation and proliferation of T cells induced by SEB can secrete great amounts of inflammatory cytokines.A number of studies found that SEB can induce systemic corticosteroid resistance,but whether it will induce topical glucocorticoid tolerance is still unknownOur recent study suggested that SEB may contribute to glucocorticoid resistance through increase of glucocorticoid receptor ?(GR?).But whether the mechanisms that SEB induce glucocorticoid insensitivity induced by decrease of the nuclear translocation of GR? are still unknown.In this work,allergic contact dermatitis model was established to observe whether SEB inhibit GR? nuclear translocation,and to investigate whether FKBP51 and FKBP52 participate in GR? nuclear translocation after SEB mediated,and whether tacrolimus reduce the effect of SEB impairing GR? nuclear translocation.Our study will put forward a new mechanism and a potential therapeutic target of GC-resistance or insensitivity induced by SEB.MethodsAllergic contact dermatitis model was established by 1% DNCB,and the changes in the number of inflammatory cells at the dermis were counted by HE staining.GR? nuclear translocation was performed by immunofluorescence staining and Western blot,respectively.Levels of cytokines including interleukine(IL)-2,IL-4,IL-13,and tumor necrosis factor-alpha(TNF-?)were observed by enzyme-linked immunosorbent assay(ELISA).Gene expression and protein levels of FKBP51 and FKBP52 was determined by RT-PCR and Western blot,respectively.This experiment was divided into two parts.The first part was that whether SEB will inhibit glucocorticoid receptor alpha nuclear translocation in keratinocytes of allergic contact dermatitis.The second part was that the mechanism of tacrolimus,FKBP51,or FKBP52 regulating SEB-mediated nuclear transport of the glucocorticoid receptor alpha in keratinocytes of allergic contact dermatitisResults1.Compared with untreated dermatitis group,the number of inflammatory cells were significantly increased in dermatitis group treated with SEB.Compared with dermatitis group treated with dexamethasone,the anti-inflammatory actions of dexamethasone were attenuated by SEB in dermatitis group treated with SEB+ dexamethasone.However,the effect of SEB could be antagonized by tacrolimus.2.Compared with untreated dermatitis group,the GR?(Cy3)nuclear:cytoplasmic ratio was significantly increased in dermatitis group treated with dexamethasone.Compared with dermatitis group treated with dexamethasone,the nuclear:cytoplasmic ratio of GR?(Cy3)was markedly decreased by SEB in dermatitis group treated with SEB+dexamethasone.However,the effect of SEB could be reversed by tacrolimus.3.Compared with untreated dermatitis group,the cytoplasmic localization of GR? was upregulated after dexamethasone treatment.Compared with dermatitis group treated with dexamethasone,SEB could increase the GR? residing in the cell cytoplasm,and decrease the GR? nuclear shuttling.And the impaired GR? nuclear shuttling could be partly reversed by tacrolimus.4.Compared with untreated dermatitis group,Levels of cytokines including interleukine(IL)-2,IL-4,IL-13,and tumor necrosis factor-alpha(TNF-?)were markedly decreased after dexamethasone treatment.Compared with dermatitis group treated with dexamethasone,SEB could increase levels of cytokines in dermatitis group treated with SEB+dexamethasone,this effect could be partly reversed by tacrolimus.5.Compared with normal control group,expression of FKBP51 was increased,and expression of FKBP52 was not significantly different in dermatitis group treated with dexamethasone.The result indicated that dexamethasone can induce expression of FKBP51 as a negative feedback mechanism in GR signaling.Similar phenomenon was also observed in dermatitis group treated with SEB and dermatitis group treated with SEB+ dexamethasone,suggesting that SEB upregulates the expression of FKBP51,which may contribute to decreased GR sensitivity.However,the increased FKBP51 could be inhibited after tacrolimus treatment,suggesting that the antagonistic effect of tacrolimus against SEB may partly dependent on downregulating the expression of FKBP51.Conclusion:1.SEB can attenuates the anti-inflammatory actions of dexamethasone in allergic contact dermatitis.2.SEB can cause steroid resistance by impairing dexamethasone-induced GR? nuclear translocation in keratinocytes of allergic contact dermatitis.3.SEB can impair GR? nuclear translocation through increasing of FKBP51 expression in keratinocytes of allergic contact dermatitis.4.Tacrolimus can restore glucocorticoid sensitivity in dermatitis group treated with SEB+dexamethasone,at least in part,by downregulating the expression of FKBP51 and increasing glucocorticoid-induced GR? nuclear translocation.