Ethanol potentiates hypoxic liver injury: role of hepatocyte Na(+) overload

Centrilobular hypoxia has been suggested to contribute to hepatic damage caused by alcohol intoxication. However, the mechanisms involved are still poorly understood. We have investigated whether alterations of Na(+) homeostasis might account for ethanol-mediated increase in hepatocyte sensitivity to hypoxia. Addition of ethanol (100 mmol/l) to isolated rat hepatocytes incubated under nitrogen atmosphere greatly stimulated cell death. An increase in intracellular Na(+) levels preceded cell killing and Na(+) levels in hepatocytes exposed to the combination of ethanol and hypoxia were almost twice those in hypoxic cells without ethanol. Na(+) increase was also observed in hepatocytes incubated with ethanol in oxygenated buffer. Ethanol addition significantly lowered hepatocyte pH. Inhibiting ethanol and acetaldehyde oxidation with, respectively, 4-methylpyrazole and cyanamide prevented this effect. 4-methylpyrazole, cyanamide as well as hepatocyte incubation in a HCO(3)(-)-free buffer or in the presence of Na(+)/H(+) exchanger blocker 5-(N,N-dimethyl)-amiloride also reduced Na(+) influx in ethanol-treated hepatocytes. 4-methylpyrazole and cyanamide similarly prevented ethanol-stimulated Na(+) accumulation and hepatocyte killing during hypoxia. Moreover, ethanol-induced Na(+) influx caused cytotoxicity in hepatocytes pre-treated with Na(+), K(+)-ATPase inhibitor ouabain. Also in this condition 4-methylpyrazole and 5-(N,N-dimethyl)-amiloride decreased cell killing. These results indicate that ethanol can promotes cytotoxicity in hypoxic hepatocytes by enhancing Na(+) accumulation.