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 been derived from the proximal tubule of kidney from transgenic mice harbouring the simian virus (SV40) large T and small t antigens placed under the control of the 5' regulatory sequence from the rat L-type pyruvate kinase (L-PK) gene. The cell lines (PKSV-PCT and PKSV-PR cells) were derived from early (PCT) and late (Pars Recta, PR) microdissected proximal tubules grown in D-glucose-enriched medium. In such conditions of culture, both cell lines exhibited L-PK transcripts, a stable expression of SV40-encoded nuclear large T antigen, a prolonged life span but failed to induce tumors when injected sub-cutaneously into athymic (nu-nu) mice. Confluent cells, grown on plastic support or porous filters, were organized as monolayers of polarized cuboid cells with well developed apical microvilli and formed domes. Both cell lines exhibited morphological features of proximal tubule cells with villin located in the apical brush-border and substantial amounts of hydrolase activity. By immunofluorescence studies using specific antibodies, aminopeptidase N appeared restricted to the apical microvillar domain, whereas the H2 histocompatibility antigen was distributed in the cytoplasm and lateral membranes. These results demonstrate that the proximal morphological phenotype has been fully preserved in these cultured cells derived from tissue-specific targeted oncogenesis in transgenic mice.
Cartier
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N
Institut Cochin de Génétique Moléculaire, Institut National de la Santé et de la Recherche Médicale (INSERM), Faculté Cochin, Paris, France.
Lacave
R
R
Vallet
V
V
Hagege
J
J
Hellio
R
R
Robine
S
S
Pringault
E
E
Cluzeaud
F
F
Briand
P
P
Kahn
A
A
D000626:Aminopeptidases / Q000032:analysis; D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics*; D018826:CD13 Antigens; D002352:Carrier Proteins / Q000032:analysis; D002454:Cell Differentiation; D002461:Cell Line, Transformed; D016764:Cell Polarity; D004847:Epithelial Cells; D004848:Epithelium / Q000648:ultrastructure; D005455:Fluorescent Antibody Technique; D007687:Kidney Tubules, Proximal / Q000737:chemistry / Q000166:cytology* / Q000648:ultrastructure; D051379:Mice; D008822:Mice, Transgenic; D008840:Microfilament Proteins / Q000032:analysis; D008871:Microvilli / Q000648:ultrastructure; D011770:Pyruvate Kinase / Q000235:genetics*; D012045:Regulatory Sequences, Nucleic Acid
D000952:Antigens, Polyomavirus Transforming; D002352:Carrier Proteins; D008840:Microfilament Proteins; C026615:villin; D011770:Pyruvate Kinase; D000626:Aminopeptidases; D018826:CD13 Antigens