Microscopic Analysis of Oxidative Stress in Cultured Cells. II. Transmission Electron Microscopy

Abstract

The insulin-sensitizing agent troglitazone (Rezulin) has been shown to cause severe hepatotoxicity and liver failure. While the toxic mechanism is still unknown, a component may be oxidative stress. We have demonstrated that increased lipid peroxidation, decreased glutathione levels, and collapse in mitochondrial membrane potential occurred following exposure of cultured hepatocytes to troglitazone. These hallmarks of oxidative stress can manifest morphologically in several ways including damage to cellular membranes, mitochondrial abnormalities, and cell death. The purpose of this study was to assess morphologic changes caused by exposure to troglitazone or cumene hydroperoxide (CH), a compound known to induce oxidative stress.HepG2 and Novikoff hepatoma cells were grown on poly-L-lysine coated 6-well plates overnight, and exposed to 10, 25, 50 or 100 μM concentrations of troglitazone or CH in the growth media for 2 hours. Cells were fixed with 2.5% glutaraldehyde in phosphate buffer, harvested, and pelleted. Pellets were post-fixed in 1% osmium tetroxide, and processed to epoxy resin.