Aberrant cell cycle checkpoint function in transformed hepatocytes and WB-F344 hepatic epithelial stem-like cells

Cell cycle checkpoints are barriers to carcinogenesis as they function to m aintain genomic integrity. Attenuation or ablation of checkpoint function m ay enhance tumor formation by permitting outgrowth of unstable cells with d amaged DNA. To examine the function of cell cycle checkpoints in rat hepato carcinogenesis, we analyzed the responses of the G(1), G(2) and mitotic spi ndle assembly checkpoints in normal rat hepatocytes, hepatic epithelial ste m-like cells (WB-F344) and transformed derivatives of both. Normal rat hepa tocytes (NRH) displayed a 73% reduction in the fraction of nuclei in early S-phase 6-8 h following 8 Gy of ionizing radiation (IR) as a quantitative m easure of G(1) checkpoint function. Chemically and virally transformed hepa tocyte lines displayed significant attenuation of G(1) checkpoint function, ranging from partial to complete ablation. WB-F344 rat hepatic epithelial cell lines at low, mid and high passage levels expressed G(1) checkpoint fu nction comparable with NRH. Only one of four malignantly transformed WB-F34 4 cell lines displayed significant attenuation of G(1) checkpoint function. Attenuation of G(1) checkpoint function in transformed hepatocytes and WB- F344 cells was associated with alterations in p53, ablated/attenuated induc tion of p21(Waf1) by IR, as well as aberrant function of the spindle assemb ly checkpoint. NRH displayed 93 % inhibition of mitosis 2 h after 1 Gy IR a s a quantitative measure of G(2) checkpoint function. All transformed hepat ocyte and WB-F344 cell lines displayed significant attenuation of the G(2) checkpoint. Moreover, the parental WB-F344 line displayed significant age-r elated attenuation of G(2) checkpoint function. Abnormalities in the functi on of cell cycle checkpoints were detected in transformed hepatocytes and W B-F344 cells at stages of hepatocarcinogenesis preceding tumorigenicity, su staining a hypothesis that aberrant checkpoint function contributes to carc inogenesis.