The Relevant Research Of MiR-378b In Hepatic Insulin Resistance Induced By Alcohol

Objective: Insulin resistance has been implicated in alcoholic liver disease(ALD).It has been reported that micro RNA(miRNA)play a major role in the production,secretion,and function of insulin.The purpose of this study was to investigate the role of miR-378 b in alcohol induced hepatic insulin resistance and its underlying mechanism.Methods: Human hepatocyte L-02 cells were induced with 200 m M ethanol for 48 h,and miR-378 b expression was detected by Q-PCR.miR-378 b mimics and miR-378 b inhibitor were transferred into L-02 cells by electroporation method.After 12 h of transfection,they were co-cultured in medium containing ethanol for 48 h.Glycogen levels in L-02 cells and glucose contents in culture medium were detected by commercial kits.ALD in vivo was induced by feeding a Lieber-De Carli liquid diet containing 5%(w/v)alcohol for 4 weeks in C57BL/6J mice.The expression of miR-378 b in liver was detected by Q-PCR.In addition,in order to construct the model of upregulation and downregulation miR-378 b in vivo,the AAV-miR-378b-up and AAV-miR-378b-down vector were injected into mice through tail vein.H&E staining was used to detect the degree of pathological changes in mouse liver.Insulin and glucose levels in serum and TG contents in liver were detected by using commercial kits and automatic biochemical analyzer.Glucose tolerance test and insulin tolerance test were used to observe the tolerance of mice to glucose.Western blot and immunocoprecipitation were used to detect the expression of related genes in the insulin signaling pathway of L-02 cells and mouse liver.In order to further explore the molecular mechanism of miR-378 b regulating hepatic insulin resistance,bioinformatics was used to predicte the target gene of miR-378 b,and double luciferase reporter gene experiment and Western blot were used to further verity.Results: The expression of miR-378 b in ethanol induced L-02 cells was significantly increased.After transfection with miR-378 b mimics,the expression level of miR-378 b was increased,the synthesis of glycogen was decreased and the glucose level in culture medium was increased in L-02 cells.However,the effects of miR-378 b mimics on L-02 cells were reversed after transfection with miR-378 b inhibitor.After the AAV-miR-378b-up was injected into mice,the expression level of miR-378 b in liver was increased,and the content of TG in serum and liver were increased,and the liver pathological damage in mice was aggravated.The AAV-miR-378b-down by tail vein injection also reversed the effects of AAV-miR-378b-up in vivo.miR-378 b could affect the insulin signaling pathway.Overexpression miR-378 b significantly reduced the protein expression levels of IR,p-IR,p-IRS1 and p-IRS2,as well as reduced the binding of p110? and p85? and decreased protein expression of p-Akt1 and p-Akt2 in L-02 cells and mice.However,loss of miR-378 b reversed the effects of overexpression miR-378 b on the insulin signaling pathway.The results of bioinformatics prediction and dual luciferase reporter gene showed that IR and p110? are indeed direct targets of miR-378 b.Conclusions: In ethanol-induced L-02 cells and C57BL/6J mice models,miR-378 b can regulate insulin sensitivity by targeting IR and p110?,suggesting that miR-378 b may be a potential target for hepatic insulin resistance,providing new insights for clinical effective diagnosis and treatment of hepatic insulin resistance.