Effects of chronic alcohol and excessive iron intake on mitochondrial DNA damage in the rat liver

Abstract

In this study, we investigated the effects of chronic alcohol and excessive iron intake on mitochondrial DNA (mtDNA) damage and the progression of alcoholic liver injury in rats. Methods: Twenty-four Sprague-Dawley male rats were divided into four groups (Control, EtOH, Fe, and EtOH + Fe), and fed either control or ethanol (36% of total calories) liquid diet with or without 0.6% carbonyl iron for eight weeks. Serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) activities, liver malondialdehyde concentrations were measured by colorimetric assays. Liver histopathology was examined by Hematoxylin-eosin staining of the fixed liver tissues. The integrity of the hepatic mtDNA and nuclear DNA was measured by long-range PCR. The gene expression levels of cytochrome c oxidase subunit 1 (Cox1) and NADH dehydrogenase subunit 4 (Nd4) were examined by real-time PCR. Results: Serum ALT and AST activities were significantly higher in the EtOH+Fe group, as compared to the Control group. Similarly, among four groups, liver histology showed the most severe lipid accumulation, inflammation, and necrosis in the EtOH + Fe group. PCR amplification of near-full-length (15.9 kb) mtDNA showed more than 50% loss of full-length product in the liver of the EtOH + Fe group, whereas amounts of PCR products of a nuclear DNA were unaffected. In addition, the changes in the mtDNA integrity showed correlation with reductions in the mRNA levels of mitochondrial gene Cox1 and Nd4. Conclusion: Our data suggested that the liver injury associated with excessive iron and alcohol intake involved mtDNA damage and corresponding mitochondrial dysfunction.