| Budget Amount *help |
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 2002: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 2001: ¥1,100,000 (Direct Cost: ¥1,100,000)
Fiscal Year 2000: ¥1,800,000 (Direct Cost: ¥1,800,000)
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| Research Abstract |
Background : Chronic alcohol consumption depresses ATP synthesis and induces mitochondrial DNA (Mt-DNA) deletion in the liver. ATP content in cells may play a critical role in inducing cell death, apoptosis or necrosis. However, it is not known which type of cell death occurs in alcoholic liver disease. In this study, the deletions of hepatic Mt-DNA including ATPase (ATP synthase) encoding region and hepatic ATP content in rats treated with ethanol were determined to elucidate the relationship between Mt-DNA deletion and ATP synthesis. Single stranded DNA (ss-DNA) of hepatocytes was also stained immunohistochemically to determine the relationship between hepatic ATP content and apoptosis.
Methods : Sixteen male Wistar rats were fed with a 36% ethanol-containing liquid diet (AL-group) or control diet without ethanol (C-group) for 5 weeks. Hepatic ATP content was measured in the extract of fresh liver tissue. To detect apoptosis of hepatocytes, ss-DNA in liver sections was stained immunoh
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istochemically. Ss-DNA positive cells per 10,000 hepatocytes of each liver tissue section were calculated. Using frozen liver sections, the deletions of Mt-DNA encoding complexes I (ND4 + ND5), IV (CO I, II and III) and V (ATPase 8/6) were determined by polymerase chain reaction (PCR).
Results : Fatty change was observed in more than one third of lobules in AL-group, but not in C-group. Hepatic ATP content in AL-group was 0.44 ± 0.10 μmol/g liver which was significantly lower than that in C-group, 0.84 ± 0.31 μmol/g liver (P<0.005). On the other hand, no deletion of Mt-DNA encoding complexes I, IV and V was detected in either AL- or C-group. Ss-DNA staining was clearly observed in the nuclei of hepatocytes in both groups. The number of ss-DNA positive hepatocytes in AL-group was 5.6 ± 1.8/10,000 hepatocytes which was significantly less than that in C-group, 20.6 ± 4.8/10,000 hepatocytes (P<0.0001). There was a positive correlation between hepatic ATP content and the number of ss-DNA positive cells (r = 0.674, P<0.005).
Conclusion : The results suggest that mitochondrial function, at least in part ATP level, was depressed before the damage of Mt-DNA by chronic ethanol consumption. Chronic ethanol consumption may not be responsible for the apoptosis of hepatocytes. Less
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