ACUTE AND CHRONIC ETHANOL INCREASES REACTIVE OXYGEN SPECIES GENERATION AND DECREASES VIABILITY IN FRESH, ISOLATED RAT HEPATOCYTES

Although reactive oxygen species (ROS) have been implicated in the eti ology of alcohol-induced liver disease, neither their relative contrib ution to cell death nor the cellular mechanisms mediating their format ion are known. The purpose of this study was to test the hypothesis th at acute and chronic ethanol exposure enhances the mitochondrial gener ation of ROS in fresh, isolated hepatocytes, Acute ethanol exposure st imulated ROS production, increased the cellular NADH/NAD(+) ratio, and decreased hepatocyte viability slightly which was prevented by pretre atment with 4-methylpyrazole (4-MP), an inhibitor of alcohol dehydroge nase. Similarly, xylitol, an NADH-generating compound, enhanced hepato cyte ROS production and decreased viability Incubation with pyruvate, an NADH-oxidizing compound, and cyanamide, an inhibitor of aldehyde de hydrogenase, significantly decreased ROS levels in acute ethanol-treat ed hepatocytes. Chronic ethanol consumption produced a sixfold increas e in hepatocyte ROS production compared with levels measured in contro ls. Hepatocytes from ethanol-fed rats were less viable compared with c ontrols, e.g., viability was 68% +/- 2% (ethanol) versus 83% +/- 1% (c ontrol) after 60 minutes of incubation. Antimycin A increased ROS prod uction and decreased cell viability; however, the toxic effect of anti mycin A was more pronounced in ethanol-fed hepatocytes. These results suggest that acute and chronic ethanol exposure exacerbates mitochondr ial ROS production, contributing to cell death.