Experimental Research Of Wenyanghuoxue Tongluo Recipe On Inhibition Of Skin Fibrosis In Bleomycin-induced Scleroderma Mice

Objective:To observe the effect of Wenyanghuoxue Tongluo Recipe(WTR)on Scleroderma,we observed the WTR on influence of Smad-4,COX-2,VEGF expression in skin tissue of bleomycin-induced Scleroderma(BIS)mouse model.Methods:Bleomycin 0.1 ml(200ug/ml)was injected in back skin of female BALB/c mice,1 time daily for 4 weeks,for successfully preparing BIS mice model.The skin macro-damage and micro-pathological changes were observed to verify the characteristics of BIS model.The BIS mice were randomly divided as no medicine-feed group,WTR-feed group,Prednisone(Pred)-feed group,WTR+Pred-feed group,as well as normal mice group without bleomycin injection as negative control.11 mice were enrolled in each group.After intragastric administration of medicine for 4 weeks,the back skin of mice was fixed by 10%formalin.The pathological changes in the skin tissue of mice were evaluated with HE staining,and detected the expression of Smad-4 in skin tissue by using immunohistochemical method and Western Blotting.The level of Cyclooxygenase-2(COX-2)and Vascular Endothelial Growth Factor(VEGF)in the skin was also detected by Western Blotting.Results:Injection of bleomycin successfully induced BIS mice model.HE staining shows that the skin tissue in model group was obviously thickening,collagen fiber hyperplasia,compared with negative group.The pathological changes in the skin tissues of three groups treated with WTR,Pred,or WTR+Pred,were all improved significantly(P<0.05).The expression level of Smad-4,COX-2,VEGF protein in BIS mice model skin significantly increased than that in negative control mice(P<0.01).And compared with model group,the level of these proteins in the skin of three groups treated with WTR,Pred,or WTR+Pred,were all decreased significantly(P<0.05).Conclusion:The WTR can ameliorate pathological changes of BIS mice skin.Smad-4,COX-2,VEGF might be involved in the pathological course of BIS mice,and the efficacy of WTR on inhibition of skin fibrosis in BIS mice may be related to the down-regulation of Smad-4,COX-2,VEGF protein expression in skin tissue.