Rapid chromatographic method to decipher distinct alterations in lipid classes in NAFLD/NASH

摘要

AIM: To establish a simple method to quantify lipid classes in liver diseases and to decipher the lipid profile in p62/IMP2-2/IGF2BP2-2 transgenic mice.METHODS: Liver-specific overexpression of the insulin-like growth factor 2 mRNA binding protein p62/IMP2-2/IGF2BP2-2 was used as a model for steatosis.Steatohepatitis was induced by feeding a methioninecholine deficient diet. Steatosis was assessed histologically. For thin layer chromatographic analysis, lipids were extracted from freeze-dried tissues by hexane/2-propanol, dried, redissolved, and chromatographically separated by a two-solvent system. Dilution series of lipid standards were chromatographed, detected, andquantified. The detection was performed by either2',7'-dichlorofluoresceine or a sulfuric acid/ethanol mixture.RESULTS: Histological analyses confirmed steatosis and steatohepatitis development. The extraction,chromatographic, and detection method showed high inter-assay reproducibility and allowed quantification of the different lipid classes. The analyses confirmed an increase of triglycerides and phosphatidylethanolamine and a decrease in phosphatidylcholine in the methionine-choline deficient diet. The method was used for the first time to asses the lipid classes induced in the p62-overexpressing mouse model and showed a significant increase in all detected lipid species with a prominent increase of triglycerides by 2-fold. Interestingly, the ratio of phosphatidylcholine to phosphatidylethanolamine was decreased, as previously suggested as a marker in the progression from steatosis to steatohepatitis.CONCLUSION: The thin layer chromatography analysis allows a reliable quantification of lipid classes and provides detailed insight into the lipogenic effect of p62.