Isolated rat renal proximal tubular cells : a model for the study of drug-induced nephrotoxicity
Renal proximal tubular cells (&'62 90&'37) were isolated from the rat in high yield by collagenase digestion of renal cortical tissue, followed by isopycnic centrifugation in a Percoll density gradient. The cells were of high viability and their identity verified by morphology and PT specific enzyme activities. Cytochrome P450 and dependent monooxygenase activities were maintained and the rapid decline in reduced glutathione prevented by addition of glycine, glutamate and cystine to the buffers used. These parameters were also maintained in culture (24h). Cytochrome P450-dependent monooxygenase activity was induced in proximal tubular cells isolated after pretreatment of rats with 3-methylcholanthrene, particularly with ethoxyresorufin as substrate, indicating the presence of inducible P450c isoenzyme(s) in PT cells. In contrast, treatment with phenobarbitone showed no induction. Treatment in vivo with the renal toxins, gentamicin and cyclosporin A (CsA), resulted in altered glomerular function and renal parenchymal damage. Treatment with gentamicin in vivo and in vitro resulted in increased PAH uptake and decreased gamma glutamyl transferase activity, suggesting a primary effect of gentamicin on cell membranes. CsA treatment in vivo produced an effect on GSH, lipid peroxidation and succinate dehydrogenase activity in the isolated PT cells. On treatment in suspension, no significant effects were observed. Treatment with gentamicin in culture (24h) did not result in a significant effect on the cell parameters studied. In conclusion, PT cells can be isolated in high yield with good viability from the renal cortex. Cell function can be maintained both in suspension and in culture. Therefore, this system may be used as a model for the investigation of PT-specific drug-induced nephrotoxicity as well as renal cytochrome P450 isoenzyme composition and induction characteristics.