The Function Of ATP Binding Cassette Transporter A6 In Fatty Acid Oxidation

Objectives::ATP binding cassette(ABC)transporters is a transmembrane protein.It can mediate transmembrane transport of lipid and other molecules via ATP.ABCA6 is a member of the ABCA subfamily,it shows ubiquitous expression in tissue.Highest expression was found in the liver,In the recent finding,the expression of ABCA6 mRNA was increased during the differentiation of macrophages,and decreased significantly after Ac-LDL stimulation.It suggests that ABCA6 may be involved in maintaining cholesterol homeostasis of cells,but the specific biological function is not clear.Our research group found that fasting can increase the expression of ABCA6 in the liver of mice.By immunohistochemical examination,we found that ABCA6 was distributed in the cytoplasm of hepatocytes and concentrated on one side of the nucleus,which was probably located in the trans-Golgi apparatus.In this study,we observed the effect of ABCA6 gene deletion on the metabolism of mice during fasting and high fat diet(HFD),and investigated the role of ABCA6 in hepatic metabolism.Method:Through the TSE animal metabolism measurement and analysis system,the activities,respiratory quotient,oxygen consumption and other metabolic indicators of ABCA6 KO mice and WT mice during feeding and fasting state were monitored.Therefore,to further investigate the role of ABCA6 during feeding and fasting 8h,12h state,plasma glucose,serum ketone bodies and free fatty acid levels of ABCA6 KO mice and WT mice and hepatic glycogen content was measured.After feeding and fasting 8h,mRNA levels of fatty acid oxidation related genes were detected by realtime-PCR.Next,Giving the ABCA6 KO and WT mice a high-fat diet(HFD)for 6 weeks,Body weight were measured every week.After 6 weeks,epididymal adipose tissue and renal adipose tissue were collected.Observed the changes in the body and adipose tissue weight.To further study the glucose metabolism and energy consumption of ABCA6 KO mice after 6 weeks HFD,several examination were carried to assess the glucose tolerance,insulin resistance and pyruvate metabolic tolerance.Plasma was collected after 6 weeks HFD,respectively,to detect the level of biochemical parameters,including plasma glucose,total cholesterol(TCH),triglycerides(TG),ketone bodies,alanine aminotransferase(ALT)and aspartate aminotransferase(AST).Liver tissue of ABCA6 KO and WT mice was collected after 6 weeks HFD,hematoxylin and eosin-stained and oil red O stained were performed to observe mice hepatocytes ultrastructure and liver fat deposition.We examined mRNA levels of gene responsible for fatty acid oxidation,lipogenesis and gluconeogenesis.Moreover,After 6 weeks HFD,fatty acid composition in the liver sample of ABCA6-KO and WT mice were measured by HPLC.Results:In the feeding state,compare with WT mice,the CO2 produced volume,O2 consumption volume,temperature,heat production were much lower in ABCA6 KO mice;In light,respiratory quotient of ABCA6 KO mice was significantly higher than WT mice.But,In the night there was no difference between two groups;We also found that the food intakes of ABCA6 KO mice was much more than WT mice;Spontaneous activities of two groups were not significantly difference.In the fasting state,The respiratory quotient of KO ABCA6 mice was significantly higher than that of WT mice within 0 to 6h after fasting,but after that there was no significant difference between WT and ABCA6 KO mice.ABCA6 KO mice caused an obvious decrease of liver glycogen content in the fed state.Fasting early,compared with WT mice,ABCA6 KO plasma glucose and serum ketone was markedly decreased.But after the prolonging of fasting,There was no metabolic changes between ABCA6 and WT mice.In the feeding,The hepatic mRNAs of CD36,ACBP,involved in fatty acid transport,were significantly increased in ABCA6 KO mice compared to WT mice;The hepatic mRNAs of HSD17BS,TLA,involved in peroxisomal fatty acid beta oxidation,were significantly increased;The hepatic mRNAs of CYP2E1,involved in microsomal fatty acid Omega oxidation,was increased;No difference was found in the hepatic mRNA levels of CPT1,LOAD and PDK4,both being involved in mitochondrial fatty acid beta oxidation;besides,Gluconeogenesis genes such as PCK and PGC1?,G6Pase and lipid synthesis genes FAS were not obvious changed.After fasting 8h,and compared to WT mice,The hepatic mRNAs of CYP2E1,CYP4A10,involved in microsomal Omega oxidation,were significantly increased in ABCA6 KO mice.The hepatic mRNAs of ACC2,involved in inhibiting mitochondrial beta oxidation,were significantly decreased.besides,The hepatic mRNAs of FAS expression was decreased.After 6 weeks high fat diet,KO ABCA6 mice have a higher weight gain rate,epididymal adipose tissue and renal adipose tissue weight was significantly higher than that WT mice.The lipid accumulation was much more in the liver of ABCA6 KO mice fed the high-fat diet was examined by H&E staining and oil red O staining.Total cholesterol(TCH),triglycerides(TG),ketone bodies,alanine aminotransferase(ALT)and aspartate aminotransferase(AST)had no significant difference.But plasma glucose were significantly increased and the sensitivity to insulin was decreased in ABCA6 KO mice,compared with WT mice,.After 6 weeks high fat diet,the hepatic mRNA levels of MGAT,involved in lipid synthesis,was significantly increased;the hepatic mRNA levels of PGC1 was decreased,fatty acid oxidation related genes such as UCP2,PDK4,CPT1,PPARawere decreased.But there was no significant difference in liver fatty acid composition between the two groups.Conclusion:ABCA6 KO mice showed fatty acid oxidation disorders When the food was abundant and the fatty acid oxidation level was low,the ABCA6 KO mice hepatic cells were compensated by increasing the fatty acid transport,enhancing the fatty acids oxidation of the microsomal and peroxidase in the liver.ABCA6 KO mice showed disorder in changing glucose oxidation to fatty acid oxidation under starvation or fasting state,fatty acid oxidation were enhanced.With the prolonging of fasting,Fatty acid oxidation disorders caused by ABCA6 KO mice can be fully compensated.After 6 weeks high fat diet,the final body weights of ABCA6 KO mice fed the high-fat diet(HFD)were significantly higher than WT mice,liver lipid accumulation is more obvious and the sensitivity of insulin is decreased in ABCA6 KO mice.Our research suggests that ABCA6 transporters may be involved in the transport of fatty acid oxidation intermediately,which play an important role in the process of rapid enhancement of fatty acid oxidation.