Glycerol-3-Phosphate Dehydrogenase Activity Assay

Glycerol-3-phosphate dehydrogenase activity was measured by determining the decrease of dihydronicotinamide-adenine dinucleotide during glycerol-3-phosphate dehydrogenase (GPDH)-catalyzed reduction of dihydroxyacetone phosphate using a GPDH activity assay kit (Takara, Shiga, Japan). Briefly, differentiated adipocytes were rinsed with PBS, scraped into enzyme extraction buffer and sonicated. After centrifugation at 10,000 rpm for 5 min, GPDH activity in supernatants was determined spectrophotometrically at 340 nm with the GPDH assay kit, according to the manufacturer’s protocol. Protein concentration was measured using a BCA protein assay kit to correct the GPDH assay results. Data are expressed as a percentage of GPDH activity in GSPE-treated cells relative to that in control cells.