Rat hepatocytes isolated from alcohol-induced fatty liver have an increased sensitivity to anoxic injury

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Abstract
The aim of this study was to determine whether rat hepatocytes isolated from steatotic or nonsteatotic livers have different thresholds for injury due to anoxia-reoxygenation. Rats were fed ethanol or control diets for 8 weeks. Histology showed that more than 75% of the hepatocytes in alcohol-fed and less than 3% in control animals contained fatty vacuoles. The glycogen content was significantly reduced in steatotic livers. Isolated hepatocytes were cast in agarose gel threads and perfused with Krebs-Henseleit bicarbonate buffer. Cell viability was determined by Trypan Blue (TB) exclusion; cell injury was determined by lactate dehydrogenase (LDH) release; and superoxide anion (O2 ·−) was determined by lucigenin-enhanced chemiluminescence (LCL). During the pre-anoxic basal perfusion the following occurred: viability was 86% ± 1% and 85% ± 1%; LDH release was 16 ± 3 and 15 ± 3 mU/min; and LCL was 4 ± 1 and 5 ± 1 nA in steatotic and nonsteatotic hepatocytes, respectively. Cell viability decreased slightly under 4 hours of aerobic perfusion without differences between the two groups. In contrast, fatty hepatocytes died much faster than did control hepatocytes during anoxia; after 3 hours viability was 17% ± 8% vs. 60% ± 2% (P < .001), respectively. With reoxygenation following 2 hours of anoxia, the changes in viability, in LDH release, and in LCL were similar in both groups. These results indicate that in hepatocytes isolated from alcohol-fed rats when compared with control hepatocytes: (1) cell viability under aerobic conditions is not influenced; (2) anoxic injury is significantly increased; (3) a reduction in the hepatic glycogen stores, which may contribute to the enhanced sensitivity to anoxia, can be demonstrated; and (4) O2 ·− generation and cell injury occurring immediately after reoxygenation do not appear to be affected.

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