CISPLATIN-INDUCED TOXICITY IN IMMORTALIZED RENAL-CELL LINES ESTABLISHED FROM TRANSGENIC MICE HARBORING TEMPERATURE-SENSITIVE SV40 LARGE T-ANTIGEN GENE

We established renal cell lines from definite nephron segments which w ere microdissected from kidneys of transgenic C57BL/6 mice, harboring the large T-antigen gene of temperature-sensitive mutant simian virus 40, pSVtsA58(ori-). Cell culture was under a humidified atmosphere of 5% CO2 in air, on collagen-coated dishes, and in RITC80-7 medium with 5% fetal bovine serum, 10 mu g/ml transferrin, 1 mu g/ml insulin, 10 n g/ml recombinant human EGF penicillin and streptomycin. Cell line whic h kept contact inhibition character was established from each segment. Cells derived from distal tubule, cortical and outer medullary collec ting duct possessed their cyclic AMP response to arginine-vasopressin, like their original nephron segment. On the other hand, cells derived from terminal proximal tubules (S3 segment) formed a cobblestone-like confluent monolayer, and did not respond to arginine-vasopressin like their fresh segments. Since cisplatin, a well-known nephrotoxic subst ance, damages proximal tubules (especially S3) rather than collecting ducts, we assayed cell number, protein content: and ATP content of cul tured S3 cells at various times after addition of 0.2 mM cisplatin. De crease of cell number, total protein content and total ATP content of culture cells occurred after 10 h incubation with 0.2 mM cisplatin. Th e 50% lethal dose (LD(50)) of cisplatin in S3 cells was 4 x 10(-5) M a fter 20 h incubation and 8.5 x 10(-6) M after 40 h incubation. Outer m edullary collecting duct (OMCD) cells were damaged 30% maximally after 20 h incubation with cisplatin, and LD(50) in them became 2.5 x 10(-5 ) M after 40 h incubation. We could show that the LD(50) of cisplatin in the OMCD cell line was three times higher than that in the S3 cell line. Thus, these cell lines are the first in the kidney to definite t he segmental origin and to maintain some differentiated unique functio ns. They are valuable for studies on intrarenal site-specific actions and possible mechanisms of action of pharmacological and toxic substan ces.