Immortalization of normal liver functions in cell culture: Rat hepatocyte‐hepatoma cell hybrids expressing ornithine carbamoyltransferase activity

Abstract

Normal rat hepatocytes have been fused with highly differentiated rat hepatoma cells. Some of the hybrids express a physiologically significant level of activity of the urea cycle enzyme ornithine carbamoyltransferase (OCT), a liver-specific function not found in the hepatoma cells. These hybrids have 10% of the adult rat liver OCT specific activity, incorporate 3H-ornithine into protein arginine, and can be selectively grown in arginine-free medium supplemented with ornithine. Somatic cell hybridization of normal differentiated cells with highly differentiated neoplastic cells of the same tissue type may be useful as a general method for obtaining permanent cell lines with new tissue-specific phenotypes.

Developmentally regulated mRNAs in mouse liver.

“…cifically how individual abundant liver mRNAs are regulated during development, it may be necessary to study their expression in vitro. This should be possible because many laboratories have succeeded in maintaining cultured hepatocytes that have retained their differentiated functions and hormonal inducibility (23)(24)(25)(26).…”

Recent advances in 2D and 3D in vitro systems using primary hepatocytes, alternative hepatocyte sources and non-parenchymal liver cells and their use in investigating mechanisms of hepatotoxicity, cell signaling and ADME

“…A successful immortalization of normal hepatocytes from different species was achieved using different strategies including cell transformation with virus genes or oncogenes (i.e. simian virus 40 large T antigen, c-myc, cH-ras) (Osanai et al 1997, reviewed by Castell et al 2006), hybrid cells obtained by the fusion of hepatocytes and immortalized cell lines (Widman et al 1979; Cassio et al 1991) or the generation of hepatic cell lines from transgenic animals (reviewed by Castell et al 2006). Immortalized hepatic cell lines have been established from livers of transgenic mouse and rat expressing the SV-40 large T antigen under the control of the hepatic L-pyruvate kinase (Courjault-Gautier et al 1997) or albumin (Bulera et al 1997) promoter, respectively; or from animals over-expressing growth factors (TGF-α or human growth hormone), early growth signals (constitutively active met-protooncogene) or truncated growth suppressor genes (p53-knockout mice) (reviewed by Castell et al 2006).…”

Expression of ornithine carbamoyltransferase gene in rat hepatoma-derived cell lines, H4-II-E and R-Y121B

“…Therefore, this level may be the maximum level attainable by culturing the cells in the medium which lacks arginine but is supplemented with ornithine. Widman et al (1979) observed that rat hepatocyte-hepatoma cell hybrids had 6-10% of the OCT activity of rat liver extract. Delers et al (1984) also observed that variant cells obtained from H4-II-EC3 possessed not more than 7.5% of the OCT activity of normal hepatocytes when they were cultured in a medium in which arginine was replaced by ornithine.…”

“…Therefore, it may be possible to reveal what kind of factor(s) is necessary for the expression of the OCT gene by examining R-Y121B cells and H4-II-E cells. Widman et al (1979) and Farmer and Goss (1991) found that the cell fusion between the hepatoma cells and normal hepatocytes was necessary for the activation of the silent OCT gene in the hepatoma cells. Farmer and Goss (1991) postulated that some activator for the OTC expression was lacking in the hepatoma cells.…”