Roland Cassingena

Mineralocorticoid receptors in SV40-transformed tubule cell lines derived from rabbit kidney

The presence of mineralocorticoid (MR) and glucocorticoid (GR) receptors was investigated in two renal tubular cell lines, derived from primary cultures of isolated rabbit kidney cortical cells infected with the wild-type SV40 virus, which exhibit thick ascending limb (RC.SV2) and collecting tubule (RC.SV3) phenotypes (Vandewalle et al. J. Cell. Physiol. 141, 1989, 203-221). MR and GR were quantified, in cell monolayers and cell cytosolic fractions, with [3H]aldosterone, [3H]dexamethasone and [3H]RU486, an antiglucocorticoid with no affinity for MR. Cytosolic receptors from RC.SV2 and RC.SV3 cells labeled with [3H]aldosterone, [3H]dexamethasone or [3H]RU486 sedimented at approximately 8 S in a 15-40% glycerol gradient. All steroids displaced bound [3H]dexamethasone to the same extent, suggesting that dexamethasone bound to both MR and GR: under the conditions of assay, [3H]aldosterone binds exclusively to MR, and [3H]RU486 to GR. In both RC.SV2 and RC.SV3 cells, [3H]aldosterone bound to one class of high affinity sites (Kd 0.14-0.8 nM; Nmax 8 to 22 fmol/mg protein). In both cell lines, the number of high affinity binding sites for [3H]dexamethasone ranged from 9 to 18 fmol/mg protein with an affinity of 0.5-1.3 nM. Compared to renal cortex, the most striking observation was a marked decrease in [3H]dexamethasone binding in primary cultures and SV40-transformed cells. These results indicate that MR and GR are expressed in two established mammalian kidney tubular cell lines providing new models of cultured renal cells for studies on the physiological effects of corticosteroid hormones.

Establishment of Permanent Renal Tubule Cell Lines by Infection With the Wild-Type and a Thermosensitive Mutant of the Simian Virus 40

This study summarizes the properties of rabbit renal tubule cell lines transformed by the simian virus 40 (SV40). By infection with the wild type SV40, it was possible to establish three different cell lines exhibiting the main functions of proximal (RC.SV1), thick ascending limb-distal (RC.SV2), and collecting (RC.SV3) tubules. Another cell line infected with a thermosensitive mutant of SV40 (RC.SVtsA58) was highly sensitive to arginine vasopressin when cells were cultured at the restrictive temperature of 39.5 degrees C, suggesting they derived from principal cells of the collecting tubule.

Accelerated malignant conversion of human HBL-100 cells by the v-Ki-ras oncogene.

The human epithelial HBL-100 cell line harbors SV40 genetic information and has an unlimited growth potential. Despite displaying properties characteristic of transformation since its early in vitro passages, it is capable of producing progressively growing tumors in nude mice only after long-term culture. This is a reproducible phenomenon and apparently not the consequence of a selection of preexisting malignant cells. Superinfection of early passage nontumorigenic HBL-100 cells with Kirsten murine sarcoma virus, which contains a Ki-ras oncogene having undergone multiple activating events, induces morphologic alterations and rapidly converts the cells to neoplastic cells, further supporting the hypothesis of multistep carcinogenesis. The HBL-100 cell line might be useful in defining the oncogenes representative of different families, which are able to complement SV40 in this system.

Activation of c-Ki-ras coexists with c-myc amplification in cells from a nude mouse tumor induced by the human breast carcinoma cell line SW 613-S.

In vitro transfection experiments have shown that cooperation between two different oncogenes can confer a fully malignant phenotype to primary rodent cells. We have previously reported that SW 613‐Tul cells, derived from a tumor induced in a nude mouse by the human breast carcinoma cell line SW 613‐S, showed a 30‐fold amplification of the c‐ myc gene. In the present work, we show that these cells also harbor an activated c‐Ki‐ras gene capable of inducing the formation of foci upon transfection of NIH 3T3 cells with SW 613‐Tul genomic DNA. Our results suggest that both the c‐myc and c‐Ki‐ras oncogenes, activated by two different mechanisms, may cooperate in the full expression of the tumorigenic phenotype of SW 613‐Tul cells.

Linkage relationship between TK and SV40 T-antigen genes in the SV40-transformed WI98VaD human cell line☆

Abstract The transfer of isolated metaphase chromosomes from SV40-transformed WI 98 VaD human cells (SV40 T-Ag + ), to thymidine kinase deficient (TK − ) 3T3 mouse cells or hypoxanthine phosphoribosyltransferase deficient (HPRT − ) Chinese hamster cells (CHK), suggest that: in the WI 98 VaD cells SV40 is inserted at least in chromosome(s) 17, the carrier of the TK gene and that, in such cells, there is no linkage relationship between HPRT and SV40 T-Ag genes.

The Induction of SV40-Transformed Chinese Hamster and Mouse Kidney Cells by Mitomycin C

The induction by mitomycin C (MC) of SV40 production in three lines of semi- or nonpermissive SV40-transformed cells is described. The production of virions by a line of semipermissive, virus-shedding Chinese hamster kidney cells is increased 1,000-fold by MC treatment; the proportion of infectious centers and V-antigen-containing cells is increased to a lesser degree, suggesting that both the burst per cell and the number of virus-producing cells are increased. MC treatment induces the production of small amounts of infectious DNA and virions in one line of nonpermissive mouse kidney cells, and infectious DNA apparently alone in another.

Viral Immortalization and Phenotypic Characterization of a Rat Hepatocyte Cell Line

Viral immortalization (SV40) of adult rat hepatocytes was performed in controlled culture conditions (isolation method, collagen matrix, defined culture medium, short-time culture before virus infecti

Methionine dependence of rat hepatocarcinoma cells in culture: relationship with tumourigenicity and oncogene expression.

The culture in methionine-deprived medium of a rat hepatoma cell line (LF) significantly decreases: (1) the growth in vitro of these cells; (2) their tumourigenicity; and (3) the expression of a panel of oncogenes. When the LF cells are replaced in a medium containing methionine, they maintain a diminished in vitro growth and tumourigenic capacity, despite an overexpression of the same oncogenes. These data point out the determinant role of a nutritional factor, methionine, in tumour growth.