The Study On The Mechanism Of Asiaticoside On Cellular Immunity And Fibrosis In Systemic Sclerosis

Objective: To investigate the effect and mechanism of asiaticoside(AS)on cellular immunity and fibrosis in systemic sclerosis.Methods: in vivo experiment: the SSC model mice were established by subcutaneous injection of bleomycin for 3 weeks.The mice were treated with penicillamine and AS at low,medium and high concentrations for 4 weeks.The body weight of each group of mice was weighed,and the central skin of the back was fixed and paraffin embedded.Peripheral blood of mice was taken and serum was separated.The levels of IL-6,IL-17 and IL-21 in serum were detected by ELISA kit.In vitro experiment: human fibroblasts were treated with AS of different concentrations,and the expression of VEGF and CTGF protein was detected by Western blot;Peripheral blood samples from patients with SSc were collected and Th17 cells were isolated by density gradient centrifugation and co-cultured with stimulants.Groups were incubated with STAT3 inhibitors and low,medium and high concentrations of AS,for 48 hours.The number of Th17 cells was detected by flow cytometry to determine the proliferation rate of Th17 cells.Th17 cells from peripheral blood of patients with SSc were selected and co-cultured with stimulants.after48 hours of culture with inhibitor Stattic and low,middle and high concentration of AS,the m RNA expression of JAK2 and STAT3 was detected by real-time fluorescence quantitative PCR,and the expression of JAK2 and STAT3 protein was detected by cellular immunofluorescence technique.Results: The life of SSC model mice is basically normal,but the spirit is depressed,the weight is reduced,the back skin HE staining has obvious fibrosis performance,which indicates the establishment of SSC model is successful;low,medium and high concentrations of penicillamine and AS can inhibit the skinfibrosis of mice;the content of IL-6,IL-17 and IL-21 in peripheral blood of mice is detected by ELISA,the results showed that AS could inhibit the increase of these three kinds of interleukin in mice.The number of human fibroblasts decreased and the number of cells shortened under the action of AS The results of blot showed that AS inhibited the expression of VEGF and CTGF in fibroblasts;the results of flow cytometry showed that AS could inhibit the proliferation of Th17 cells in a concentration dependent manner;the m RNA of JAK2 and STAT3 detected by real-time fluorescence quantitative PCR showed that the transcription of these two proteins was inhibited;the results of cell immunofluorescence showed that the expression of JAK2 and STAT3 / p-STAT3 was inhibited To inhibit.Conclusions: AS,especially the high concentration group(20%),significantly changed the weight loss of SSc model mice,alleviated skin fibrosis,inhibited the proliferation of Th17 cells and the increase of three closely related interleukin in systemic sclerosis,inhibited fibrinogen,and decreased the transcriptional and expression levels of JAK2 and STAT3.This suggests that AS can reduce the degree of inflammation and fibrosis in SSc,and JAK2/STAT3 pathway may be an important pathway of inflammatory response in SSc.