Effect Of MiR-378b On Lipid Metabolism In Alcoholic Liver Disease

Objective: Alcoholic liver disease has become a major cause of morbidity and mortality all over the world,and the mechanisms underlying ALD still remain unclear.Recent studies highlight the involvement of Mi RNAs in regulating hepatic lipid metabolism.However,current information on the biological function of mi R-378 b inhepatic lipid metabolism is limited.The aim of this study was to clarify the role of mi R-378 b in alcohol-induced hepatic lipid accumulation and its underlying mechanism.Methods: In this study,the expression of mi R-378 b in L-02 cell was detected by Q-PCR after the model was established with ethanol in concentration of 200 m M for 48 h.In order to further explore whether mi R-378 b is involved in regulating alcohol-induced hepatic lipid accumulation,the target genes of mi R-378 b were predicted by bioinformatics methods,and confirmed by luciferase reporter assay and Western blot.Mi R-378 b mimics and inhibitors were transfected into L-02 cells by electroporation method.After transfection for 12 h,they were co-cultured in medium containing ethanol for 48 h.The triglyceride and cholesterol levels in the cells were detected with a kit.Alcoholic liver disease mouse model was established by feeding C57BL/6J mice a Lieber De Carli liquid diet containing5%(W/V)ethanol for 4 weeks.The expression of mi R-378 b in mouse liver was detected by q-PCR.The AA V-mi R-378b-up and AA V-mi R-378b-down adeno-associated virus vectors were injected into mice through the tail vein to construct upregulation and downregulation mi R-378 b mouse model.H&E staining was used to determine the degree of lesion and lipid accumulation in mouse liver tissues.Cholesterol and triglyceride levels in serum and liver of mice were measured by industrial kit and automatic biochemical analyzer.PCR and Western blot were used to detect the expression levels of lipid metabolism related factors in L-02 cells and mouse liver.Results:The results showed that expression of mi R-378 b was increased in alcohol-induced L-02 cells,and bioinformatics and luciferase reporter results showed that Ca MKK2 was the direct target of mi R-378 b.In addition,after transfection with mi R-378 b mimics,the expression of mi R-378 b was increased and the accumulation of TG and TC was increased in L-02 cells,while transfection with mi R-378 b inhibitor partially reversed the induction effect of alcohol on L-02 cells.After the injection of AA V-mi R-378b-up in mice,the expression of mi R-378 b in the liver of mice was increased,the levels of TG and TC in the serum and liver were increased,and the pathological injury of the liver of mice was further aggravated.While inhibition of mi R-378 b in vivo partially reversed the effects of alcohol in mice.In both L-02 cells and mice,Mir-378 b can influence the conduction of lipid metabolism pathway,and overexpression of mi R-378 b can significantly reduce the protein expression levels of Ca MKK2,P-AMPK/AMPK,CPT1α and PPARα,and significantly increase the protein expression levels of P-ACC /ACC,SREBP1 c and FASN.The inhibition of protein partially reversed the effects of ethanol on lipid metabolism-related signaling pathways in mice liver.Conclusions: Data suggest that mi R-378 b can regulate hepatic lipid metabolism by targeting Ca MKK2 in alcohol-induced L-02 cells and C57BL/6J mice model,and might serve as a potential therapeutic target for the treatment of ALD.