HEPTOCYTE PROLIFERATION AND AFLATOXIN METABOLISM

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The hypothesis that continued proliferation of hepatocytes is associated with an transgenic immature phenotype in regard to metabolism of aflatoxin (AFB1) and development of hepatocellular carcinoma (HCC) will be analyzed in p53 null and p53ser246 mutant transgenic, and HBV mice. World-wide HCC is arguably the tumor that causes the highest cancer mortality. HCCs occurring in high-risk areas of the world are associated with continued proliferation of liver cells in response to liver injury (HBV and HCV), and with aflatoxin (AFB1) exposure. AFB1/HBV associated HCCs have a high frequency of mutations in the tumor suppressor gene p53 at codon 249 (p53ser249). Recently we reported that hepatocytes of p53 null mice continue to proliferate into adulthood and do not become polyploid with aging. In addition, hepatocytes of p53ser246 transgenic mice (equivalent to p53ser249 of humans) maintain a high number of cells in G1. P53 null and p53ser246 mice also have increased susceptibility to AFB1 hepatocarcinogenesis when the carcinogen is administered during the first week of life, and HBV transgenic mice are susceptible to AFB1 carcinogenesis, even when the carcinogen is administered at one month of age when normal mice of the same strain are resistant. We wish to test the hypothesis that the changes in status of cells in the livers of these mice reflects a less-differentiated phenotype associated with increase production of carcinogenic metabolites and increased risk of HCC development in adult mice. In specific aim 1, the metabolism of AFB1 at different ages of normal and transgenic mice will be examined by AFB1 epoxide and adduct formation, glutathione-s-transferase activity and p450 isoenzyme distribution. In addition, oval cells or small hepatocytes will be isolated from cocaine-treated mice and from neonatal and adult transgenic livers to determine if there is distinctive metabolism of AFB1 in these cells. The effect of proliferation of "mature" hepatocytes will be determined using microsomes from partially hepatectomized livers, and metabolism of AFB1 by cultured liver cells at different stages of differentiation will be examined. In Specific Aim 2, AFB1 will be administered after one month (when normal mice are not susceptible to carcinogenesis) to p53+/- hemizygous mice, to p53-/- null mice, to p53ser246 hemizygous and homozygous transgenic mice, and to F1 mice of p53ser246 mice cross-bred to p53 null mice and the development of HCC determined. the effect Of HBV associated injury and AFB I hepatocarcinogensis in these mice. In Specific Aim 3, the effect of HBV associated injury and AFB1 hepatocarcinogenesis in these mice will be examined.