Biochemical analyzer

Mice were anesthetized with isoflurane and blood collected via the orbital vascular plexus. The blood was centrifuged and the serum collected to detect the free fatty acids and low-density lipoprotein (LDL) using a fatty acid quantitation kit and LDL quantitation kit (Sigma), respectively, according to the manufacturer’s protocol as described previously [26 (link)]. The serum levels of glutamate oxaloacetate transaminase (GOT), glutamate pyruvate transaminase (GPT), total cholesterol (TC), high-density lipoprotein (HDL), and total TGs were measured using the biochemical analyzer (DRI-CHEM NX500, Fuji, Tokyo, Japan). The day before the end of the experiment, mice fasted for 16 h and were administered glucose by intraperitoneal injection to assay blood glucose levels using the biochemical analyzer (Fuji). Blood insulin was detected using the Insulin EIA Kit (Cayman, Ann Arbor, Michigan, USA) according to the manufacturer’s protocol. Liver glycogen was detected using the Glycogen Assay Kit (Cayman) according to the manufacturer’s protocol, and the glycogen levels were measured using a microplate reader (Multiskan FC) at an absorbance of 570 nm.

Mice in each group were anesthetized by isoflurane, and peripheral blood samples were obtained through eyeball blood collection. The levels of serum liver enzymes, including alanine aminotransferase (ALT), and aspartate aminotransferase (AST), were determined by a biochemical analyzer (Fuji, Tokyo, Japan).