CELL-LINES WITH HETEROGENEOUS PHENOTYPES RESULT FROM A SINGLE ISOLATION OF ALBUMIN-SV40 T-ANTIGEN TRANSGENIC RAT HEPATOCYTES

Several immortalized cell Lines were established from the livers of tw o transgenic rats expressing the simian virus protein large T antigen under the control of the albumin promoter. Hepatocytes from transgenic rats were isolated by a two step perfusion procedure from a normal-ap pearing liver and from a liver that contained a single neoplasm, Cells were also isolated from the dissociated liver neoplasm, After 5 weeks in culture, cell colonies were isolated and subcloned as individual c ell lines. Electron microscopy revealed that all cell lines had a high nuclear-to-cytoplasmic ratio compared with normal hepatocytes. The cy toplasm contained numerous organelles, including smooth and rough endo plasmic reticulum, Golgi, mitochondria, peroxisomes, and annulate lame llae, However, the lines exhibited a variety of different cell morphol ogies. All cell lines, including those derived from neoplastic cells, exhibited a similar doubling time of 26 hours and the ability to grow in soft agar, Northern blot analysis revealed that the cell lines diff erentially expressed hepatocyte markers, Large T antigen was expressed in the cultured cell lines at much higher levels than was observed in transgenic hepatocytes in vivo. This suggests that the viral protein is required to maintain cell viability in culture, In addition, the tu mor suppressor proteins, p53 and Rbp105, were detected in all cultured cell lines, In contrast, these same proteins were not detected by Wes tern blot in transgenic hepatocytes in vivo. All cell lines expressed the oncogene c-myc, yet growth factor-dependent and independent growth were observed, The data presented show for the first time the establi shment and characterization of a number of cell lines derived from hep atocytes isolated from an alb-SV40 large T-antigen transgenic rat, The se cell lines exhibited varied morphological and biochemical hepatocel lular characteristics in vitro, suggesting that the expression of the transgene in hepatocytes leads to considerable phenotypic diversity an alogous to that seen in hepatocellular carcinomas induced by chemical carcinogenesis.