| Objective:To explore the role of STING-IRF3 signaling pathway in the induction of diabetes mellitus with psoriatic skin lesions in mice induced by imiquimod.As well as the palmitic acid(PA)+ imiquimod induced human keratinocyte(HaCat)Cell inflammation.Methods:1.Animal experiments:(1)model of imiquimod-induced psoriatic lesions in mice: twenty-four 8-weekold male diabetic homozygous(db/db)and wild-type(wt/wt)mice with no specific pathogen(SPF)grade were randomly divided into four groups after 1 week of adaptive basal feeding.Group A(wt/wt + vaseline),group B(wt/wt + imiquimod),group C(db/db + vaseline),group D(db/db + imiquimod),6 in each group.Anesthetized mice and their back hair was removed,exposing a certain area of back skin;Among them,mice in groups B and D applied imiquimod cream(62.5mg/ piece)evenly on the back of the mice for 7 consecutive days,and mice in groups A and C applied the same amount of vaseline ointment for 7 consecutive days.Psoriasis lesion area and severity index(PASI)scores were performed daily during application.(2)skin pathomorphological observation: part of the fixed back skin tissues were embedded in paraffin and stained with HE,and the number of epidermal cell layers and the number of inflammatory cell infiltration were observed.(3)immunohistochemical staining: another part of the back skin tissue was fixed with paraffin embedded sections and immunohistochemical staining was conducted to observe the expression level of STING protein in the skin tissue.2.Cell experiment:(1)in vitro cultured Hacat cells were subcultured and divided into four groups:group A: cells were treated with culture medium only;Group B: treated with 1mg/m L imiquimod;Group C: cells were treated with palmitic acid containing 200umol/L.Group D cells were treated with imiquimod 1mgl/m L+200umol/L palmitic acid.(2)Western blot experiment: cell proteins were collected from the above four groups after cell culture for 6h,and the expression levels of STING,IRF3,P-IRF3 of the target proteins were detected by Western blot.Results:1.Animal experimental model:(1)visual observation of the skin: in group B,compared with group A,the back skin of mice presented obvious symptoms such as erythema,descination and skin thickening,and PASI score was significantly increased(P<0.05).Compared with group C,mice in group D showed obvious symptoms such as erythema,descination and skin thickening,and PASI score was significantly increased(P<0.05).Compared with group B,mice in group D showed obvious symptoms such as erythema,descination and skin thickening,and PASI score increased(P<0.05).(2)HE staining results showed that the skin tissue on the back of group B was significantly thicker,the number of cell layers was significantly increased,the keratinized layer was significantly increased,and the inflammatory cell infiltration was obvious(P<0.05).Compared with group C,the skin tissue on the back of group D was significantly thicker,the number of cell layers was significantly increased,the keratinized layer was significantly increased,and the inflammatory cell infiltration was obvious(P<0.05).Compared with group B,the skin tissue on the back of group D was significantly thicker,the number of cell layers was significantly increased,the keratinized layer was significantly increased,and the infiltration of inflammatory cells was more obvious(P<0.05).(3)skin immunohistochemical staining showed that the relative expression of STING protein in the dorsal skin of mice in group B was higher than that in group A(P<0.05).The relative expression of STING protein was increased in group D compared with group C(P<0.05).The relative expression of STING protein was increased in group D compared with group B(P<0.05).2.Cell experiment:(1)western blot analysis showed that the levels of STING and p-irf3 protein in group C were significantly higher than that in group A after 6h of intervention(P<0.05).The levels of STING and p-irf3 in group D were significantly higher than that in group A(P<0.05).The levels of STING and p-irf3 in group D were significantly higher than that in group C(P<0.05).Conclusions:(1)The psoriatic skin changes induced by imiquimod were more severe in obese diabetic mice than in non-obese diabetic mice.(2)Compared with non-obese diabetic mice,the expression of STING protein was significantly increased in the skin lesions induced by imiquimot-induced psoriasis in obese diabetic mice.(3)imiquimod combined with palmitic acid induced human keratinocytes cell STING,IRF3 and P-IRF3 expression significantly increased.(4)The STING-IRF3 pathway may play a role in the inflammatory response of diabetes mellitus with psoriasis. |
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