Tumor necrosis factor induced oxidative stress in isolated mouse hepatocytes

Abstract

Tumor necrosis factor α (TNFα) is a macrophage-derived cytokine which participates in homeostatic tissue repair. It is also a potentially useful antitumor agent. Liver toxicity, however, limits TNFα's clinical utility and suggests that it may play a role in liver toxicity of various etiologies. To determine the direct effects of TNFα on hepatocytes, in the absence of infiltrating leukocytes and other inflammatory mediators, an isolated mouse hepatocyte model has been used in the present study. Hepatocytes exposed to recombinant human TNFα (1–10 μg/ ml) exhibited intracellular GSH depletion and GSSG efflux during the first 2 hr of exposure, but no cytotoxicity was observed. However, TNFα was toxic to hepatocytes pretreated with 1,3-bis(chloroethyl)-1-nitrosourea to inhibit GSSG-reductase activity. Furthermore, these cells exhibited a greater efflux of GSSG upon exposure to TNFα. TNFα also caused a marked decrease in cellular ATP concentrations, which occurred after initiation of effects on the glutathione pool. These findings indicate that high concentrations of TNFα induce an oxidant stress in isolated hepatocytes. The antioxidants mannitol and benzoate, as well as the iron chelator deferoxamine, reduced the extent of TNFα-induced oxidant effects in hepatocytes, which indicates that the oxidant stress may involve hydroxyl radical generation. Hepatocytes treated with ruthenium red or fructose were less susceptible to TNFα-induced ATP depletion, which suggests that mitochondrial calcium cycling may be involved in disruption to energy metabolism.