| Background:Non-alcoholic fatty liver disease(NAFL)is a clinically common disease with no history of excessive drinking.It is a clinical pathological syndrome characterized by hepatic cell steatosis and lipid storage.With the progression of the disease,its pathological changes range from simple fatty liver,steatohepatitis,fatty liver fibrosis,liver cirrhosis,and even hepatocellular carcinoma.Micro RNA(micorRNA,miR)is a type of short-chain non-coding RNA.The miRNA is complementary to the 3 ’UTR of the mRNA molecule of the target gene,causing the degradation or translation inhibition of the mRNA and affecting the expression of downstream proteins.We found that miR-486 is highly expressed in the liver tissue of NAFL mouse,but the role of miR-486 in NAFL is not yet clear.In this study,miR-486 knockout mice and wild mice were fed with high-fat diet to simulate NAFL,and then exploring the impact of miR-486 on the progression of NAFL.Objective:To investigate the effect of micro RNA-486(miR-486)on the progression of non-alcoholic fatty liver in mice.Methods:The first part: the research in this part of the project firstly used C57BL/6J mice to be fed with high-fat diet to simulate NAFL.The experiment was divided into a control group(WT-NCD group)and an experimental group(WT-HFD group).The wildtype C57BL/6J mice were randomly assigned to 4cages per group,2 mice per cage,and 8 mice per group.The mice in the experimental group were fed with TP23400 high-fat model feed and the control group were fed with TP23402 low-fat and low-sugar control feed.Each mouse was fed with about 3g per day for 24 weeks and weighed once every 4 weeks.Then,after the experiment,the mice were sacrificed to record body weight and liver weight and collect their liver tissue and serum for further testing.After analysis and comparison,the cycle weight change,weight at the end of the experiment,liver weight,liver weight-to-body weight ratio,intrahepatic triglyceride content,and serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),and triglyceride(TG),cholesterol(CHO),tumor necrosis factor-α(TNF-α)were analyzed and compared.Liver tissue sections were stained with hematoxylin-eosin(HE)and saturated and oil red O.The effect of NAFL on the weight,liver and blood biochemistry of mice was evaluated at the same time.Then,the level of miR-486 in the liver was detected by real-time fluorescence quantitative polymerase chain reaction,and the relationship between miR-486 and the progression of NAFL was evaluated;Part two: after confirming the relationship between miR-486 and the progression of NAFL,miR-486 knockout mice was used to investigate the role of miR-486 in the progression of NAFL in this part of project.miR-486 knockout mice and wild type C57BL/6J mice were feed high-fat diet to simulate NAFL.The experiment was divided into miR-486 knockout control group(KO-NCD group),miR-486 knockout experiment group(KO-HFD group),wild control group(WT-NCD group)and wild experiment group(WT-HFD group).The miR-486 knockout mice and littermate mice were separately divided into 4 cages per group,2 mice per cage,and 8 mice per group.The mice in the experimental group were fed with TP23400 high-fat diet and the mice in the control group were fed with TP23402 low-fat and low-sugar control diet.Each mouse was fed with about 3g per day for 24 weeks and weighed once every 4 weeks.After the experiment,the mice were sacrificed to record body weight and liver weight and liver tissue and serum were collected for further testing.After analysis and comparison,the cycle weight change,weight at the end of the experiment,liver weight,liver weight-to-weight ratio,intrahepatic triglyceride content,and serum alanine aminotransferase(ALT),aspartate aminotransferase(AST),and triglyceride(TG),cholesterol(CHO),tumor necrosis factor-α(TNF-α)were analyzed and compared.Liver tissue sections were stained with hematoxylin eosin(HE)and oil red O staining;the protein levels of AMPK and ACC,a lipid metabolism-related molecule,were detected by Western blotting,and then the function of miR-486 in non-alcoholic fatty liver in mice was investigated.Results:The first part: the weight of mice in the WT-NCD group was significantly lower than that in the WT-HFD group.HE staining and saturated oil red O staining showed that liver fat in the WT-HFD group became the most severe,and fat in the WT-NCD group was significantly lighter.Compared with the WT-HFD group,serum ALT,AST and TNF-α in the WT-NCD group were significantly lower,and the differences were statistically significant;the level of miR-486 in the WT-HFD group was higher than that in the WT-NCD group,and the differences were statistical significance.Part two: the body weight of the KO-HFD group was significantly lower than that of the WT-HFD group.H&E staining and saturated oil red O staining showed that liver fat in the WT-HFD group was the most severe,and that in the KO-HFD group was significantly improved.Compared with the WT-HFD group,serum ALT,AST,and TNF-α in the KO-HFD group were significantly reduced,and the differences were statistically significant.Compared with the WT-HFD group,the AMPK mRNA level was higher in the KO-HFD group,while ACC was lower.And the difference was statistically significant.Conclusion:MiR-486 deficiency can alleviate non-alcoholic fatty liver disease in mice,and its mechanism may be involved in regulating lipid metabolism. |
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