Molecular mechanism of hepatic carcinogenesis and hepatocyte apoptosis in the Rb transgenic mice

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We created the Rb transgenic mice in which the human Rb cDNA was controlled under the HNF-1 promoter/enhancer and got two lines. We determined that the A mouse had 11 copies of the transgene per haploid and the B mouse had 4 copies per haploid.We injected diethylnitrosamine into the abdominal cavity of the A mice, B mice and wild-type mice and then feed them with phenobarbital-containing water for 35 weeks. After sacrifice, the livers were examined. In the wild-type mice, several hepatocellular carcinomas were detected while no carcinomas were detected in the Rb transgenic mice, A and B mice. We extracted DNAs from the hepatocellular carcinoma in the wild-type mice and investigated the mutations of Rb,p53, Ras and Myc genes, but no mutations were detected. The number of nodules in the liver was largest in the wild-type mice, secondarily larger in the B mice and lesser in the A mice. This result indicates that Rb protein inhibits nodule formation in a dose-dependent manner.To find the molecules involved in resistance to fulminant hepatitis, firstly the cellular extracts from the livers were prepared and subjected to Western blotting. The results showed that the amounts of caspase 1, caspase 3,p53,E2F1-E2F5,Bcl-2,Bcl-XL,Bcl-XS, Bad and Bid proteins showed no differences between the wild-type and A mice. However, the Bax protein was decreased in the A mice and B mice compared to that in the wild-type mice. Second, we did 2-dimensional electrophoresis using the protein extracts from the livers in the A mice and wild-type mice. We found that 5 proteins appeared in the A mice, but not in the wild-type mice and 7 proteins disappeared in the A mice, but not in the wild-type mice.