Expression of glutathione-dependent enzymes and cytochrome P450s in freshly isolated and primary cultures of proximal tubular cells from human kidney

The expression of glutathione (GSH)-dependent enzymes and cytochrome P450 ( P450) proteins in freshly isolated proximal tubular cells from human kidney (hPT), and the effect of primary culture on these enzymes, were determined . Freshly isolated hPT cells had relatively high activities of gamma-glutam yltransferase, gamma-glutamylcysteine synthetase, glutathione S-transferase (GST), glutathione disulfide reductase, and GSH peroxidase. Cytochrome P45 0 4A11 was detected in freshly isolated hPT cells, whereas CYP2E1 was not. Freshly isolated hPT cells also expressed GSTA, GSTP, and GSTT but not GSTM . Primary cultures of hPT cells maintained their epithelial-like nature and diploid status, based on measurements of morphology, cytokeratin expressio n, and flow cytometric analysis. hPT cells retained GSH-dependent enzyme ac tivities during primary culture, whereas cells that had undergone subsequen t passage exhibited a loss of activities of most GSH-dependent enzymes and no longer expressed P450s or GSTs. CYP4A11 expression in primary cultures o f hPT cells was significantly increased after treatment for 48 h with eithe r ethanol (50 mM) or dexamethasone (7 nM). GSTA, GSTP, and GSTT contents, a lthough still detectable, were decreased compared with those of freshly iso lated hPT cells. Our data show that hPT cells express enzymes involved in x enobiotic disposition, and that they thus provide a model suitable for stud ies of human renal drug metabolism. Furthermore, primary cultures of hPT ce lls may afford the opportunity to study factors regulating P450 enzyme expr ession in human kidney.