Diets: Mice were fed either a high-fat diet (HFD; Research Diets #D12492) or a chow diet (Chow; LAB Diet #5010). Glucose dysmetabolism: Fasting glucose and insulin and glucose tolerance testing were quantified after an overnight fast. Liver: Hepatic triglycerides were quantified using Triglyceride Reagent and Triglyceride Standards (Pointe Scientific); serum alanine transaminase (ALT) levels were quantified using ALT Reagent and Catatrol I and II (Catachem); lipid peroxidation was quantified using 4-hydroxynonenal (4-HNE) enzyme-linked immunosorbent assay (ELISA) reagents (Cell Biolabs)—all according to the manufacturer's instructions.
Alanine transaminase (ALT) levels were quantified from 10 μL of mouse serum per sample in a 96 well flat-bottom plate (Costar) and a BioTek Synergy 2 Multi-Mode Microplate Reader with Gen5 ver. 2.00 software. Catatrol I and II (Catachem Inc.) were used as controls and were prepared according to manufacturer instructions. ALT buffer was prepared by mixing ALT Activator Reagent and ALT sample Diluent Reagent (Catachem Inc.) and 200 μL was subsequently added to each sample. Starting at time zero, absorbance was recorded at 340 nm and 37°C once per minute for 5 minutes and blank value was subtracted from all samples. The ALT concentration (U/L) was subsequently calculated using the equation ALT U/L = (OD/min x 205) / (6.22 x 0.2 x0.005), where OD = change in absorbance.