ESTABLISHMENT OF RENAL PROXIMAL TUBULE CELL-LINES BY TARGETED ONCOGENESIS IN TRANSGENIC MICE USING THE L-PYRUVATE KINASE-SV40 (T) ANTIGEN HYBRID GENE

Targeted oncogenesis allowed us to obtain two cell lines which have be en derived from the proximal tubule of kidney from transgenic mice har bouring the simian virus (SV40) large T and small t antigens placed un der the control of the 5' regulatory sequence from the rat L-type pyru vate kinase (L-PK) gene. The cell lines (PKSV-PCT and PKSV-PR cells) w ere derived from early (PCT) and late (Pars Recta, PR) microdissected proximal tubules grown in D-glucose-enriched medium. In such condition s of culture, both cell lines exhibited L-PK transcripts, a stable exp ression of SV40-encoded nuclear large T antigen, a prolonged life span but failed to induce tumors when injected sub-cutaneously into athymi c (nu-nu) mice. Confluent cells, grown on plastic support or porous fi lters, were organized as monolayers of polarized cuboid cells with wel l developed apical microvilli and formed domes. Both cell lines exhibi ted morphological features of proximal tubule cells with villin locate d in the apical brush-border and subtantial amounts of hydrolase activ ity. By immunofluorescence studies using specific antibodies, aminopep tidase N appeared restricted to the apical microvillar domain, whereas the H2 histocompatibility antigen was distributed in the cytoplasm an d lateral membranes. These results demonstrate that the proximal morph ological phenotype has been fully preserved in these cultured cells de rived from tissue-specific targeted oncogenesis in transgenic mice.