Alcohol & Alcoholism Vol. 39, No. 4, pp. 290-296, 2004
Alcohol & Alcoholism Vol. 39, No. 4 © Medical Council on Alcohol 2004; all rights reserved
DUAL EFFECT OF ETHANOL ON CELL DEATH IN PRIMARY CULTURE OF HUMAN AND RAT HEPATOCYTES
Unidad Clínica Aparato Digestivo, Hospital Universitario Reina Sofía, Córdoba, Spain
* Author to whom correspondence should be addressed at: Unidad de Investigación, Unidad Clínica Aparato Digestivo, Hospital Universitario Reina Sofía, Av. Menendez Pidal s/n, E-14004 Córdoba, Spain. Tel.: +34 957 011070; Fax: +34 957 010452; E-mail: jordi.muntane.exts{at}juntadeandalucia.es
(Received 16 September 2003; in revised form 28 February 2004; accepted 9 March 2004)
Aims: In-vivo and in-vitro studies have shown that ethanol induces hepatocyte damage. The aim of the present study was to evaluate the effect of a broad range of ethanol concentrations on apoptosis and necrosis in primary culture of human and rat hepatocytes. Methods: Human and rat hepatocytes were isolated from human hepatectomies and male Wistar rats (200–250 g) using the classical collagenase perfusion method. After stabilization of cell culture, ethanol (0–10 mmol/l) was administered and the parameters were measured 24 h after ethanol addition. Apoptosis was studied by DNA fragmentation, iodide propidium–DNA staining, caspase-3 activity and annexin V binding in hepatocytes. Necrosis was evaluated by lactate dehydrogenase (LDH) release. Malondialdehyde (MDA) and GSH/GSSG were used as parameters of oxidative stress. Results: Ethanol enhanced dose-dependently all the parameters associated with apoptosis in human and rat hepatocytes. Low or high ethanol concentrations induced an opposite action against cell necrosis in cultured hepatocytes. Low concentrations of ethanol (1–2 mmol/l) reduced LDH release from human and rat hepatocytes. However, the highest ethanol concentration (10 mmol/l) induced a sharp increase in cell necrosis. The effect of ethanol on cell necrosis was related to lipid peroxidation in hepatocytes. Conclusions: Ethanol differentially regulates apoptosis or necrosis in cultured hepatocytes. Although ethanol exerted a dose-dependent induction of apoptosis, low ethanol concentrations were able to reduce basal lipid peroxidation and necrosis in hepatocytes. The highest ethanol concentration (10 mmol/l) induced apoptosis and necrosis in human and rat cultured hepatocytes.