| Budget Amount *help |
¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1993: ¥500,000 (Direct Cost: ¥500,000)
Fiscal Year 1992: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Research Abstract |
The liver cells from the 18-week-old human embryo were immortalized by transfection with the simian virus 40 (SV40) T-antigen gene, and a cell line OUMS-22 was established. On the other hand, the cells from normal adult human liver were also transfected with SV40 T-antigen gene, but failed to become an immortal cell line. OUMS-22 cells were SV-40 T-antigen-positive, epithelial-like, and immunoreactive against an anti-keratin 18 monoclonal antibody. The staining pattern of cytokeratin in the cells was similar to that in the differentiated human hepatocellular carcinoma cell lines but not to that in the human cholangiocellular carcinoma cell lines. OUMS-22 cells expressed neither alpha-fetoprotein nor albumin mRNAs. However, OUMS-22 cells were sensitive to cytotoxicity of aflatoxin B1 and benzo[a]pyrene, whereas human embryo lung fibroblasts were insensitive to the cyntotoxicity of these carcinogens. These findings suggest that OUMS-22 cells may arise from undifferentiated liver stem cells or from hepatocytes that lost their ability to express the liver-specific functions prior to immortalization. The expression of glutathione S-transferase pi mRNA highly increased in OUMS-22 cells with their immortalization. Karyotypic analysis showed that numerical and structural aberrations of the chromosomes were profound in OUMS-22 cells. OUMS-22 cells could grow in soft agar, but they were not tumorigenic when transplanted into nude mice. Then, OUMS-22 cells were transfected with the cloned activated c-Ha-ras oncogene containing a point mutation within codon 61. However, the activated c-Ha-ras transfected cells were not tumorigenic. Even after further transfection with the mutant p53 gene, which has a conversion of CGT into CAT at codon 273 in the exon 8, the cells failed to become malignant. On the other hand, the cloning efficiency of OUMS-22 cells was significantly increased by treatment with a combination of aflatoxin B1 and phenobarbital, but the cells formed no tumors whe
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