Control of serum protein production in hepatocyte hybridomas: immortalization and expression of normal hepatocyte genes.

Abstract

ABSTRACT"Hepatocyte hybridomas" have been isolated after fusion of adult hepatocytes and a-fetoprotein (AFP-producing mouse hepatoma cells. The yield of viable hybrid clones was low but could be increased by culturing the cells in the presence of insulin. On the basis of their chromosome constitution, the hybrids were classified into two groups characterized by either a single or a double set of mouse (hepatoma) chromosomes. The hybrids segregated rat chromosomes and thus constitute an excellent material for gene mapping studies in the rat.Most of the hepatocyte hybridomas retained the production of one or more rat serum proteins, indicating that the corresponding structural genes, contributed by the normal hepatocyte parent, have been immortalized and maintained in the active state after fusion. However, these hybrids do not produce rat AFP, although mouse AFP synthesis is maintained. This result strongly suggests that silent rat (hepatocyte) AFP genes coexist in hepatocyte hybridoma nuclei with active mouse (hepatoma) AFP genes. Finally, on the basis of certain proerties of these hepatocyte-hepatoma hybrids, we suggest that the nondividing state of the hepatocytes is actively controlled by a regulatory mechanism which prevents DNA synthesis or entry into mitosis or both. Hybrids formed between normal diploid cells and tumor cells of the same tissue origin (hybridomas) have proved useful for immortalizing certain functions expressed by normal differentiated cells, as shown by the numerous studies on the now classical "lymphocyte hybridomas" (1, 2). We have explored the possibility that differentiated liver functions could be immortalized by hybridization of normal hepatocytes with mouse hepatoma cells. In this communication, we report the successful isolation of interspecies "hepatocyte hybridomas" and show that these hybrids provide a tool for immortalizing normal liver functions.Hybridomas could also be valuable for analyzing the expression of differentiated functions subjected to developmental controls or to alterations associated with a malignant phenotype. This possibility has not yet been subjected to much investigation. Production of a-fetoprotein (AFP) is modulated during hepatic differentiation: AFP is produced actively by the fetal hepatocytes but only in trace amounts by the adult liver; and, in addition, it is synthesized by most hepatomas (3, 4). By fusing liver cells which differ in the expression of the AFP gene, it should be possible to explore the mechanisms controlling the expression of this gene. In previous studies, we have shown that hybrids formed between AFP-producing mouse hepatoma cells and non-AFP-producing rat hepatoma cells retain the capacity to secrete mouse AFP (5), unlike hybrids generated between AFP-producing mouse hepatoma cells and fibroblasts (6, 7). By fusing AFP-producing mouse hepatoma cells with adult ratThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U....

Differential activity of a tissue-specific extinguisher locus in hepatic and nonhepatic cells.

“…This raises the possibility that Tse-J is generally active in nonhepatic cells. Conversely, * intratypic hepatoma x hepatoma or hepatoma x hepatocyte hybrids continue to express liver functions; i.e., extinction does not occur in such hybrids (6,38). Accordingly, one might expect that Tse-J activity is not expressed in hepatic cells.…”

Neuronal traits of clonal cell lines derived by fusion of dorsal root ganglia neurons with neuroblastoma cells.

“…Thus, lymphocyte hybridomas continue to secrete antibodies, and liver and endothelial hybrids generate a-fetoprotein and factor VIII, respectively (7)(8)(9). Hybrids derived from fusion of sympathetic ganglia and neuroblastoma cells can synthesize dopamine (10).…”

Gene mapping in the rat by mouse-rat somatic cell hybridization: synteny of the albumin and α-fetoprotein genes and assignment to chromosome 14

“…The hybrids between mouse hepatoma cells (BWTG3) and rai hépatocytes were isolated as described previously (Szpirer et al, 1980). They were grown in DM AT medium (Eagle's medium, modified by Dulbecco) supplemented with 10% fetal calf scrum and I O'4 M hypoxanthine.…”

“…Immunodiffusion against anti-rat albumin was done as described previously (Szpirer et al 1980). The amount of rat albumin secreted by the hybrid cells was estimated by comparing serial dilutions of concentrated culture medium to standard solutions of purified rat albumin.…”