MiR-206 Regulates Lipid Metabolism to Improve non-Alcoholic Fatty Liver Disease by The SIRT1/AMPK Pathway
Objective:To investigate the effect of miR-206 on the non alcoholic fatty liver(NAFLD)cell model induced by long chain free fatty acids(FFA)in HepG2 cells and its relationship with silent information regulatory factor 1(SIRT1)/AMPK pathway.Methods:HepG2 cells were treated with 1mM FFA for 24 hours to construct a NAFLD cell model.According to the different transfected substances,they were divided into 6 groups:Control group(without any treatment),FFA group(NAFLD model),FFA+mimics NC group(transfect mimics NC plasmid),FFA+miR-206 mimics group(transfect miR-206 mimics plasmid),FFA+Vector group(transfect Vector plasmid),and FFA+OE-SIRT1 group(transfect OE-SIRT1 plasmid).Oil red O staining was used to detect lipid accumulation in cells.En-zyme-linked immunosorbent assay(ELISA)was used to detect the content of aspartate aminotransferase(AST),alanine aminotransferase(ALT),and triglycerides(TG).Fluorescence quantitative polymerase chain reaction(qRT-PCR)was used to detect the expression of miR-206 and SIRT1 mRNA.Western blot was used to detect SIRT1,sterol regulatory element binding protein 1(SREBP1),fatty acid synthase(FAS),stearoyl coenzyme Al(SCD1),acetyl CoA carboxylase 1(ACC1),leukocyte differentiation antigen 36(CD36),AMP ac-tivated protein kinase(AMPK),and p-AMPK protein levels.TargetScan was used to predict the binding site of miR-206 to SIRT1.Dual luciferase reporter gene experiments was used to validate the targeting relationship between miR-206 and SIRT1.Results:Compared with the Control group,the FFA group showed a significant increase in AST,ALT,and TG content,an increase in TG content,a significant decrease in miR-206,SIRT1 protein and gene mRNA content,an increase in SREBP1,FAS,SCD1,ACC1,and CD36 protein content,and a decrease in p-AMPK/AMPK ratio(P＜0.01).Compared with the FFA+mimics NC group cells,the FFA+miR-206 mimics group cells showed a decrease in intracellular TG content,a decrease in SREBP1,FAS,SCD1,ACC1,and CD36 protein levels,an increase in SIRT1 protein and gene mRNA levels,and an increase in p-AMPK/AMPK ratio,with statistically significant differences(P＜0.01).Com-pared with the FFA+Vector group cells,the FFA+OE-SIRT1 group cells showed a decrease in TG content,a decrease in SREBP1,FAS,SCD1,ACC1,and CD36 protein levels,and an increase in p-AMPK/AMPK ratio(P＜0.01).There was a binding site between miR-206 and the SIRT1 wild-type.The cell luciferase activity in the SIRT1 WT+miR-206 mimics group was higher than that in the SIRT1 WT+mimics NC group(P＜0.01).Conclusion:Upregulation of miR-206 can alleviate NAFLD induced lipid metabolism in liver cells through the SIRT1/AMPK pathway.
Non alcoholic fatty liver diseaseMiR-206Silent Information Regulatory Factor 1
NETL
NSTL
万方数据
