| Objective:Shugan granule mainly prevents and treats the alcoholic liver disease (ALD),in particular the alcohol of liver fibrosis(LF).In order to explore whether shugan granule can reverse the LF,shugan granule were used to the cultured rat hepatic stellate cells(HSC),to find out its impact on the expression of HSCα-smooth muscle actin,tumor necrosis factor-α,transforming growth factor-β1,to explored the impact and related mechanisms of the proliferation and activation of HSC by shugan granule,and to provide further theoretical basis for shugan granule,s clinical application. Methods:HSC were cultured in RPMI1640 medium which contain 10% premium fetal bovine serum in 37℃,5%CO2 incubator.The impact of shugan granule on cell proliferation were detected in different concentration (0.01mg/ml,0.02mg/ml,0.04mg/ml,0.08mg/ml) with MTT.In accordance with the test results,the experiment was divided into the blank control group,shugan granule 0.01mg/ml group,shugan granule 0.02mg/ml group and shugan granule 0.04mg/ml group.HSC was intervented in vitro for 24h,then the morphological changes were detected,α-SMA expression was investigated with Immunohistochemical method,TNF-αexpression with radioimmunoassay, TGF-β1 expression with fluorescence quantitative PCR.Results:1.ODs in 4 concentrations of Shugan granule group were less than ODs in control group,0.01mg/ml group had no significant difference with control group(P>0.05),other 3 groups had significant difference with control group(P < 0.05).2.The morphohological changes of cells were observed in four groups by microscopic.In control group,cell's bodys were flat and extended into a membranous flim,which showed myofibroblast changes;in shugan granule 0.01mg/ml group the cell bodys were slightly inhibited and the shape looked like stellate;in shugan granule 0.02mg/ml group the shape of cell body looked like between shugan granule 0.01mg/ml group and 0.04mg/ml group;in shugan granule 0.04mg/ml group the cell body were inhibited significantly and looked like spindle similar to quiescent form.Each group were no karyopyknosis or apoptosis.3.α-SMA espression was detected with Immunohistochemical method:α-SMA expressed in each group,in control group the positive rate was(62.83±3.42)%,in shugan granule 0.01mg/ml group was(47.31±2.74)%, in shugan granule 0.02mg/ml group was(37.79±3.01)%,in shugan granule 0.04mg/ml group was(25.74±2.83)%,among control group and three shugan granule intervention group there had a significant difference(P < 0.01).4.TNF-αexpression was detected with radioimmunoassay:the mean values of TNF-α'expression in control group,shugan granule 0.01mg/ml group,0.02mg/ml group and 0.04mg/ml group were:(2.44±0.54)ng/ml,(2.13±0.35)ng/ml,(1.92±0.33)ng/ml and(1.37±0.35)ng/ml,there was no significant difference between control group and shugan granule 0.01mg/ml group(P>0.05),while among shugan granule 0.02mg/ml,0.04mg/ml group and control group the differences were statistically siganificant(P<0.05).5.TGF-β1 espression was detected with fluorescence quantitative PCR:the numerical of 2-△△Ct of control group and each drug intervention group were:(1.00±0.09),(0.71±0.08), (0.55±0.07),(0.44±0.03),and there were statistical significance among them(P<0.01).Conclusion:1.Shugan granule may decrease the proliferation of HSCs to prevent and cure hepatic fibrosis in a concentration dependent manner.2.Shugan granule can inhibit the HSC activation and the expression of HSCα-SMA.3.Shugan granule can inhibit the expression of HSC TNF-α.4.Shugan granule can inhibit the expression of HSC TGF-β1.
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