The Influence On The Expression Of SIRT1、PGC-1α From The Protective Effect Of Tangshenfang On Diabetic Liver Injury

Objective To study the protective effects and mechanism of Tangshenfang on the hepatic injury by observing the liver function,pathological changes and SIRT1,PGC-la and α-SMA expression changes in the liver tissue in diabetic rats,in order to provide new ideas and experimental basis to the prevention and treatment of diabetic liver injury.Methods 1.Type 2 diabetic models(T2DM)are made by the combination of 30mg/kg 1%Streptozocin(STZ)intravenous injection and high-carbohydrate and high-fat diet with 50 SPF male SD rats.Rats with FBG≥16.7mmol/L were considered as type 2 diabetic rats and fed with high-fat diet unceasingly.2.Type 2 diabetic models are randomly divided into diabetes mellitus(DM)model group,Tangshenfang low,meduim,high dose groups,valsartan group and healthy control group.3.Drug intervention:the valsartan group treated with valsartan mixed suspension;Tangshenfang low,medium,high dose groups were treated with concentrating agent of Tangshenfang in 6.5 g.kg-1.d-1,13g.kg-1.d-1 and 26g.kg-1.d-1;DM model group and healthy control group were treated with normal saline.All groups are treated by intragastric administration.4.After 12 weeks of drug intervention,serum biochemical indices such as FBG,TC,TG,LDL-C,HDL-C,ALT,AST are detected,FINS is tested by radioimmunoassay and HOMA-IR is counted,liver pathology changes are observed under light microscopy in HE and Masson staining,the expression of SIRT1,PGC-1α and a-SMA protein in liver are measured by immunohistochemistry technology,the expression of SIRT1 and PGC-la gene in liver are measured by fluorescence quantitative PCR technique,the content of SOD,MDA in liver tissue is measured by hydroxylamine method and thiobarbituric acid method.Results 1.Compared with healthy control group,rats in DM model group had the symptoms of being emaciated obviously,drinking more,urination more,slow response,drooping spirits,withered color etc.FBG,AST in serum increased apparently(P<0.05),FBG,TC,TG,LDL-C increased and HDL-C decreased apparently in serum(P<0.01),ALT,ALT,FINS,HOMA-IR in serum increased,MDA increased and SOD decreased apparently in liver tissue(P<0.05)in the rats of model group.After 12 weeks of Tangshenfang intervention,the general condition of rats are improved and the therapeutic effect is obvious.Compared with DM model group,FBG,TC,TG,LDL-C in serum decreased,HDL-C increased,ALT,AST decreased in serum.MDA decreased and SOD increased apparently in liver tissue(P<0.05).2.In the rats of DM model group,we found the obvious lipoid degeneratiaon of liver cells,companied by focal cell necrosis,inflammatory cell infiltration with HE staining and obvious fiber hyperplasia in intermediate zone and portal area with MASSON staining.After Tangshenfang intervention,most fat droplet vacuole reduced obviously or even dispeared in liver cells,inflammatory cell and collagenous fiber reduced distinctly.And the reduction was most significant and even disappeared in Tangshenfang high-dose group.3.Immunohistochemistry technology showed:Compared with the healthy control group,SIRT1 protein expression decreased,α-SMA,PGC-1α protein expression increased,hepatic stellate cell(HSC)was activated obviously in DM model group.Compared with the DM model group,SIRT1 protein expression increased,α-SMA,PGC-1α protein expression decreased,HSC expression decreased(P<0.05).4.Fluorescence quantitative PCR technique showed:compared with the healthy control group,SIRT1 gene expression decreased and PGC-1αgene expression increased in DM model group.After Tangshenfang intervention,compared with the DM model group,SIRT1 gene expression increased and PGC-1α gene expression decreased(P<0.05)in healthy control group.Conclusion 1.After 12 weeks of molding,we found fatty degeneration,focal necrosis,inflammatory cell infiltration,fibrous tissue proliferation in liver cells which indicated there was liver injury in diabetic rats and belonged to non-alcoholic fatty liver disease.After Tangshenfang intervention,pathological changes of liver tissues and liver function improved that meaned there was protective effect of Tangshenfang on diabetic liver injury.2.Biochemical criterion showed the increased blood glucose,blood fat,insulin resistance index and the related indicators of oxidative stress including decreased SOD and increased MDA in liver tissue,that indicated there were oxidative stress and insulin resistance in diabetic rats,confirming the pathophysiologic foundation of liver injury caused by diabetes.After Tangshenfang intervention,insulin resistance index,MDA decreased and SOD increased,indicating Tangshenfang might protect diabetic liver injury by intervening oxidative stress and insulin resistance.3.Decreased SIRT1 protein expression and increased PGC-1α,α-SMA protein expression indicated the pathogenesis of diabetic liver injury might be related with abnormal expression of the above protein.After Tangshenfang intervention,SIRT1 protein expression increased and PGC-1α,α-SMA protein expression decreased,manifesting Tangshenfang might protect diabetic liver injury by increasing SIRT1 protein expression and then activating PGC-1α to promote the metabolism of fat.