Effects of ethanol and acetaldehyde on reactive oxygen species production in rat hepatic stellate cells

Background: Alcoholism is a common cause of cirrhosis. Hepatic stellate cells are the main source of collagen that ultimately leads to hepatic fibrosis and cirrhosis. Reactive oxygen species (ROS) enhance stellate cell activation and stimulate fibrogenesis. In this study, the acute effects of ethanol (ET) and acetaldehyde (AC) were determined on the production of ROS in isolated rat hepatic stellate cells. Methods: Rat stellate cells were isolated in situ by perfusion of the portal vein and cultured. Hydrogen peroxide (H ₂O₂) was determined by luminol-derived chemiluminescence (CL), while superoxide anion (O₂^•-) production was assessed by the fluorescent probe hydroethidine. Results: AC increased the formation of H₂O₂ and O₂^•-, and these effects were first detectable at AC concentrations of 5 and 10 μM, respectively, reaching a maximum at 50 to 75 μM. Reduction of glutathione (GSH) synthesis by 1-buthionine sulfoximide (BSO) or by GSH conjugation with dimethylmaleate (DEM) further enhanced the effects of AC on H₂O ₂ and O₂^•- formation, while N-acetylcysteine (NAC) decreased H₂O₂ and eliminated the enhanced generation of O₂^•- caused by AC. Raloxifene, which inhibits O₂^•- production by NAD(P)H oxidase, reduced the effects of AC on H₂O₂ and O₂ ^•- production. ET increased H₂O₂ or O ₂^•- only in the presence of BSO or DEM. Conclusions: This study shows that concentrations of AC, which occur in vivo after the ingestion of alcoholic beverages, result in the formation of ROS in rat hepatic stellate cells. The increases in ROS are known to activate stellate cells promoting fibrogenesis.