Identifying mechanistic effects on growth of single and combined genetic alterations in mouse hepatocytes

My research objective was to further define how specific molecular alterations affect hepatocyte growth homeostasis. Interactions between constitutively-activated Hras with c-myc, TGFalpha or a heterozygous p53 allele were assessed using standard transgenic mouse methodologies. The combination of activated H-ras with altered p53 status resulted in a reduced time to development of endstage disease compared to the liver targeted expression of H-ras alone. However, the addition of c-myc or TGFalpha did not result in differences in the liver weight as a percentage of body weight when compared to the liver-targeted expression of activated H-ras alone. Using a novel hepatocyte transplantation assay, H-ras/myc and H-ras/TGFalpha increased the growth of foci during the growth permissive phase of hepatic repopulation, with activated H-ras alone and in combination with altered p53 status being sufficient to induce continued focus growth in a growth restrictive environment. Furthermore, I generated and characterized transgenic mice with inducible hepatic expression of mutant beta-catenin using the same approaches noted above. Initiating expression of mutant beta-catenin in utero or in adult liver resulted in the rapid development of hepatomegaly and was associated with altered ammonia metabolism. Mutant beta-catenin did not increase growth of foci during either growth permissive or restrictive phases, but was associated with development of liver tumors at the time of endstage disease development in transplant recipients. Finally, I found the presence p53 heterozygosity did not appreciably affect lesion latency or phenotype associated with myc/TGFalpha-induced liver tumors in transgenic mice. These studies provide additional insight regarding the mechanistic effects of specific genetic alterations alone or in combination during hepatocarcinogenesis. H-ras or beta-catenin alterations are frequently found in mouse liver tumors and are almost exclusive of one another. My studies indicate these alterations act via distinct mechanisms in hepatocarcinogenesis: H-ras by enabling hepatocytes to grow in a cell autonomous fashion, and beta-catenin by altering hepatocellular function which may produce a microenvironment conducive to generating additional genetic alterations. The lack of effect associated with p53 heterozygosity on the development of myc/TGFalpha-induced liver tumors is striking, given previous reports of increased levels of genomic instability associated with these tumors.