Journal Article

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

William K. Kaufmann, Cynthia I. Behe, Vita M. Golubovskaya, Laura L. Byrd, Craig D. Albright, Kristen M. Borchet, Sharon C. Presnell, William B. Coleman, Joe W. Grisham and Gary J. Smith

in Carcinogenesis

Volume 22, issue 8, pages 1257-1269
Published in print August 2001 | ISSN: 0143-3334
Published online August 2001 | e-ISSN: 1460-2180 | DOI: http://dx.doi.org/10.1093/carcin/22.8.1257
Aberrant cell cycle checkpoint function in transformed hepatocytes and WB-F344 hepatic epithelial stem-like cells

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Cell cycle checkpoints are barriers to carcinogenesis as they function to maintain genomic integrity. Attenuation or ablation of checkpoint function may enhance tumor formation by permitting outgrowth of unstable cells with damaged DNA. To examine the function of cell cycle checkpoints in rat hepatocarcinogenesis, we analyzed the responses of the G 1 , G 2 and mitotic spindle assembly checkpoints in normal rat hepatocytes, hepatic epithelial stem-like cells (WB-F344) and transformed derivatives of both. Normal rat hepatocytes (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 measure of G 1 checkpoint function. Chemically and virally transformed hepatocyte 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 function comparable with NRH. Only one of four malignantly transformed WB-F344 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 induction of p21 Waf1 by IR, as well as aberrant function of the spindle assembly checkpoint. NRH displayed 93% inhibition of mitosis 2 h after 1 Gy IR as a quantitative measure of G 2 checkpoint function. All transformed hepatocyte and WB-F344 cell lines displayed significant attenuation of the G 2 checkpoint. Moreover, the parental WB-F344 line displayed significant age-related attenuation of G 2 checkpoint function. Abnormalities in the function of cell cycle checkpoints were detected in transformed hepatocytes and WB-F344 cells at stages of hepatocarcinogenesis preceding tumorigenicity, sustaining a hypothesis that aberrant checkpoint function contributes to carcinogenesis.

Keywords: BrdU , bromodeoxyuridine; FACS , fluorescence-activated cell sorter; FITC , fluorescein isothiocyanate; HBSS , Hank's balanced saline solution; IR , ionizing radiation; MEM , minimal essential medium; MPF , M-phase promoting factor; NHF , normal human fibroblasts; NRH , normal rat hepatocytes; PB , phenobarbital; PCR , polymerase chain reaction; SSCP , single-strand conformational polymorphism.

Journal Article.  11360 words.  Illustrated.

Subjects: Clinical Cytogenetics and Molecular Genetics

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