TRPA1 Enhances The2,4-dinirtochlorobenzene-induced Atopic Dermatitis Skin Inflammation

Objectives:To explore the regulatory role of TRPA1 in DNCB-induced AD mouse model,and to further investigate whether TRPA1 antagonist HC-030031 has a certain therapeutic effect on DNCB-induced AD mice.Methods:（1）DNCB-induced AD model was established in wild male C57BL/6 mice,and the expression levels of TRPA1 mRNA and protein in backlesionsweredetectedbyqRT-PCR,westernblotand immunohistochemistry.（2）In order to further study the role and mechanism of TRPA1 in AD,DNCB-induced AD model was established again,which was divided into three groups:TRPA1^-/-+DNCB group,WT+DNCB group and vehicle group.The dynamic changes of dermatitis score,ear thickness and pruritus score were observed.The pathological changes were assessed by HE staining.The changes of serum total IgE level were compared by ELISA.The infiltration degree of mast cells was assessed by toluidine blue staining.The expression of cytokines in skin lesions was detected by qRT-PCR.The infiltration of macrophages was detected by WB and immunofluorescence.（3）In order to explore whether TRPA1antagonistHC-030031hastherapeuticeffecton AD,DNCB-induced AD model was established again,which was divided into three groups:HC-030031+DNCB group,WT+DNCB group and Vehicle group.The severity of skin lesions,ear thickness and pruritus score in mice were observed.After the model was established,skin lesions of back and ear were taken to detect the pathological changes.Results:（1）Skin lesions and HE staining showed that DNCB-induced AD model was successfully established in C57BL/6 mice,and the expression of TRPA1 was increased in back lesions.（2）The dermatitis score,ear thickness and pruritus score in TRPA1^-/-group were significantly lower than those in WT group（P<0.05）.HE staining showed that the epidermal proliferation and inflammatory cell infiltration in TRPA1^-/-group were less than those in WT group,and the dermal mast cell infiltration in TRPA1^-/-group was less than that in WT group,but there was no significant difference in serum total IgE level between TRPA1^-/-group and WT group.The results of qRT-PCR showed that the levels of IL-1β,TNF-α,IL-6 and Th2 cytokines（IL-4,IL-13）in the TRPA1^-/-group were lower than those in WT mice but the levels of Th1 cytokines INF-γwas no significantly difference.It was also found that TRPA1^-/-alleviated the infiltration of macrophages in the lesions.（3）After treatment with HC-030031,the severity of skin lesions,ear thickness and pruritus score in mice were lower than those of WT group,and the degree of epidermal hyperplasia and inflammatory cell infiltration was reduced.Conclusions:In DNCB-induced AD mice,TRPA1 deficiency not only alleviates pruritus,but also plays an immunoprotective role by reducing mast cell infiltration,proinflammation,Th2 inflammatory factors and macrophage infiltration.Moreover,TRPA1 antagonist HC-030031 has a certain therapeutic effect on DNCB-induced AD mice.