Protections And Mechanisms Of TRIM59 On The Development Of Non-alcoholic Fatty Liver Disease In Mice

Background and Objects:Non-alcoholic fatty liver disease（NAFLD）is a clinicopathological syndrome characterized by hepatocyte steatosis and lipid storage without a history of excessive drinking alcohols.NAFLD is not only causing liver disease,but also closely related to the high incidence of atherosclerotic cardiovascular diseases,metabolic syndromes and other diseases.In recent years,NAFLD has become an important cause of chronic liver disease due to sedentary lifestyle and western diets in developed countries such as Europe and the United States and wealthy areas of our country.However,there is no specific drug for the treatment of NAFLD.TRIM59 is a member of the TRIM family and plays an important role in tumors,but its role in NAFLD has not yet been reported.Our previous data revealed that TRIM59 is highly expressed in the liver and also plays an important role in the inflammatory response.In this study,we explored the role of TRIM59 in NAFLD and its related molecular mechanisms to provide new ideas for the prevention and treatment of NAFLD and new targets for drug development.Experimental Methods:1.Analysis of hepatic TRIM59 expression in NAFLD1)Establishing hepaticsteatosis models in mouse primary hepatocytes by different concentrations of oleic acid（OA）,2)Isolating and extracting the livers from wild-type mice fed ND and HFD for 18 weeks respectively,3)Collecting normal parts and fatty lesion parts of the livers from donor’s bodies;The total RNAs and proteins were extracted to detect the expression of TRIM59.2.Preparation and phenotypic analysis of mouse NAFLD models induced by high fat diet8 weeks old of TRIM59 knockdown(TRIM59^+/-)and control（WT）male mice were divided into four groups:1)ND-WT group,wild type（WT）mice were fed with normal diet（ND）,2)ND-TRIM59^+/-group,TRIM59 knockdown(TRIM59^+/-)mice were fed with normal diet,3)HFD-WT group,wild type（WT）mice were fed with high fat diet（HFD）,4)HFD-TRIM59^+/-group,TRIM59 knockdown(TRIM59^+/-)mice were fed with high fat diet,respectively,for 18 weeks.The body weights and blood glucoses were monitored weekly,the insulin tolerance test and glucose tolerance test were performed just before the end of the experiments.The serum,livers and white adipose tissues of each mice were collected.The total cholesterol（CHOL）,high-density lipoprotein（HDL）,low-density lipoprotein（LDL）,alanine aminotransferase（ALT）,aspartate aminotransferase（AST）and albumin（ALB）were detected in serum according to their related measurement kits.The white adipose tissues were weighed and fixed,and paraffin sections were prepared,and examined by H&E staining.Part of the liver tissues were fixed in 4%PFA,and prepared for cryosections to to analyze the accumulation of lipid droplets by H&E and oil red O.Moreover,the total RNAs and proteins were extracted to detect the expressions of genes involved in inflammation and lipid metabolism in liver tissue by q RT-PCR and Western blot;The intracellular triglyceride（TG）contents of liver tissue were also examined.3.Mechniasm of TRIM59 knockdown in development of NAFLD in miceAnalysis the signaling pathways involved in lipid metabolism in liver tissue by q RT-PCR and Western blot.Results:1.The expressions of TRIM59 were down-regulated in mouse primary hepatocytes under the stimulation of oleic acid,TRIM59 expressions were also significantly decreased in liver tissues from wild type NAFLD mice.Moreover,TRIM59 expressions were significantly decreased in fatty lesion parts comparing with normal parts of the autopsy livers from donors.These results suggest that TRIM59 may play an important role in the development of NAFLD.2.The results from intracellular triglyceride（TG）content,H&E staining,and Oil red staining showed that the lipid accumulations in liver were significantly increased in HFD-TRIM59^+/-mice comparing with HFD-WT mice.Knockdown of the TRIM59 gene significantly increases the weight of mice,but does not change the concentration of Blood glucose,GTT,ITT;increases the weight of White adipose tissue and the size of fat cells;increases the accumulation of lipids in the liver and also increases the blood lipid components in the serum;in addition,It also increases the production of inflammatory factors in the liver tissue.3.Knockdown of TRIM59 significantly increases the expression of CD36 and PPARγin liver tissue;overexpression of TRIM59 significantly reduces the expression of PPARγprotein,but does not change the level of its m RNA.Conclusions:1.TRIM59 expression was decreased in the liver of NAFLD models.2.TRIM59 knockdown can significantly promote non-alcoholic fatty liver induced by high-fat diet.The mechanism may be to promote CD36 expression by reducing the degradation of PPARγ,thereby promoting the absorption of lipids in the liver,and ultimately aggravating non-alcoholic fat the development of liver disease.