A Preliminary Study On The Mechanism Of Lipid Metabolism Mediated By Acetyl-Coenzyme A Acyltransferase 2

No-nalcoholic fatty liver disease(NAFLD)refers to the accumulation of lipids in the liver caused by factors other than alcohol.The main pathological feature of NAFLD is diffuse hepatocyte steatosis.The main reason for this is the lipotoxicity caused by excessive free fatty acid(FFA)metabolism in the liver.The excess FFA in the liver is mainly stored in lipid droplets(LDs)in the form of triglycerides(TG).Therefore,the accumulation of TG and the metabolism of FFA are closely related to non-alcoholic fatty liver.Acetyl-Coenzyme A acyltransferase 2(ACAA2)also known as mitochondrial3-oxyacyl-Co A thiolytic enzyme,participates in fatty acid metabolism.However,the mechanism of the effect of ACAA2 on lipid metabolism is not clear.In this study,we first explored the effect of ACAA2 on lipid metabolism in mice liver by over-expression of ACAA2,in mice liver.The results showed that obvious lipid accumulation was found in the liver of mice after overexpression of ACAA2.And insulin resistance exists at the same time.However,the mechanism of the effect of ACAA2 on excessive fat accumulation in the liver of mice needs to be further studied.In this study,the effect of ACAA2 on liver lipid metabolism in mice was determined by overexpressing mice model.The effect of ACAA2 on fat metabolism was further confirmed by overexpression system in vitro.The molecular mechanism of ACAA2 promoting fat accumulation was further clarified by exploring the interaction between ACAA2 and ATGL.1.ACAA2 promotes fat accumulation in the liver.ACAA2 is located in mitochondria and plays a role in the last step of fatty acidβ-oxidation.We used adeno-associated virus to overexpress ACAA2 in mice liver.Through insulin tolerance and glucose tolerance test,we found that insulin sensitivity decreased in mice after overexpression of ACAA2.Through westernblot assay,we found that the phosphorylation of insulin receptor substrate IRS1 protein tyrosine decreased after overexpression of ACAA2,suggesting that the insulin signal pathway in the liver of mice was damaged.In addition,the contents of fat-related factors in liver and serum were detected,and Hype staining showed that ACAA2 promoted the accumulation of fat in the liver of mice.It is suggested that ACAA2 plays an important role in promoting fat accumulation in non-alcoholic fatty liver disease.2.ACAA2 promotes lipid accumulation by inhibiting ATGL.By overexpressing ACAA2 and ATGL,we found that ACAA2 and ATGL co-located in cells through co-immunoprecipitation and immunofluorescence experiments,and there was interaction between ATGL and ACAA2.The further results showed that the interaction domain between ACAA2 and ATGL was the carbon-terminal thiolytic domain of ACAA2 and the patatin-like phospholipase domain of ATGL.Through immunofluorescence experiment,it was found that the co-localization of ACAA2 and ATGL patatin-like phospholipase domain was different from that of ACAA2 thiolysis domain and ATGL patatin-like phospholipase domain.This difference in co-location suggests that the interaction between different regions of the two proteins may perform different functions.In addition,the effect of ACAA2 on the TG content of normal hepatocytes(L-02)was detected by overexpression in vitro and knockdown of ACAA2.The results showed that overexpression of ACAA2 inhibited the decomposition of TG,while knocking down ACAA2 promoted the decomposition of TG.These results show that ACAA2 inhibits the decomposition of TG by ATGL by interacting with ATGL,thus promoting the accumulation of fat in hepatocytes.3.Structural analysis of ACAA2 protein.ACAA2 is a member of the thiolytic enzyme family,which is located on mice chromosome 7 and consists of 397 amino acids.The crystal structure of mice ACAA2 was obtained by X-ray diffraction.ACAA2 crystal has 27 β-folds and 15 α-helices.In addition,through the GST-pulldown experiment,we found that there is a direct interaction between ACAA2 and ATGL.To sum up,ACAA2 is closely related to NAFLD.ACAA2 inhibits the decomposition of triglycerides by ATGL through direct interaction with ATGL.