Inducible Nitric Oxide Synthase Is Required in Alcohol-Induced Liver Injury: Studies with Knockout Mice

Abstract

Background & Aims: Oxidative stress contributes to early alcohol-induced liver injury, and superoxide (O2.-) production from NADPH oxidase plays a key role. However, the production of the free radical nitric oxide (NO.) by inducible nitric oxide synthase (iNOS) could also be involved. Methods: To test this hypothesis, iNOS knockout (B6.129P2-Nos2tm1 Lau) and wild-type mice were fed high-fat control or ethanol-containing diets for 4 weeks. Results: Mean body weight gains were not significantly different between treatment groups, and average urine ethanol concentrations were similar in wild-type and iNOS knockout mice. After 4 weeks, serum alanine aminotransferase (ALT) levels were increased significantly about 4-fold over control values (29 ± IU/L) by enteral ethanol (113 ± 20) in wild-type mice; this effect of ethanol was significantly blunted in iNOS knockout mice (50 ± 9). Similar protective effects against liver damage were observed if wild-type mice were treated with the iNOS inhibitor N-(3-aminomethyl)benzyl-acetamindine (1400W). Enteral ethanol also caused severe fatty accumulation, mild inflammation, and necrosis in the liver in wild-type mice but had no effect in iNOS knockout mice. The accumulation of 4-hydroxynonenal (lipid peroxidation) and 3-nitrotyrosine (reactive nitrogen species formation) protein adducts caused by alcohol was completely blocked in iNOS knockout mice. Conclusions: These data strongly support the hypothesis that iNOS is required for the pathogenesis of early alcohol-induced hepatitis by production of nitric oxide-derived pro-oxidants (e.g., peroxynitrite).