Cultured hepatocytes from adult Fischer 344 rats were transformed by virion or cloned simian virus 40 (SV40) DNA using the calcium phosphate method. Transformation by SV40 occurred in either serum-supplemented medium or chemically defined medium (CDM). The frequency was greatest in serum-supplemented medium but transformants did not remain differentiated. In contrast, SV40 transformants developed less frequently in CDM, but retained differentiated functions. The frequency of transformation was enhanced by treatments that stimulated cell proliferation, in particular supplementing CDM with epidermal growth factor. Hepatocytes transformed in CDM were epithelial in morphology, secreted albumin, transferrin, hemopexin, and expressed the enzyme glucose-6-phosphatase, all characteristics of normal liver. Transformants did not produce detectable levels of α-fetoprotein, a marker of fetal or abnormal liver. We conclude that (a) hepatocytes can be transformed by transfection with SV40 DNA; (b) the frequency of transformation is enhanced by stimulating DNA synthesis; and (c) the transformed cells retain specific functions of normal hepatocytes in situ. Using this system it will be possible to study transformation of hepatocytes by viral and cellular oncogenes and to determine their effects on hepatocellular differentiation.

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Supported in part by Grants CA 18450, CA 23931, and CA 09124 from the National Cancer Institute.

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