P-glycoprotein in HK-2 proximal tubule cell line

P-glycoprotein (PGP) is an efflux pump physiologically expressed in the api cal membrane of the proximal tubular cells. PGP may play a role in the elim ination of exogenous substances such as chemotherapeutic drugs, calcium cha nnel blockers and immunosuppressors. The involvement of renal PGP in the tr ansport of endogenous substrates is under investigation. HK-2 is an immorta lized proximal tubule cell fine from normal adult human kidney, reported to retain a phenotype indicative of a well-differentiated state. No data rega rding expression and/or activity of PGP in this cell line are available The aim of this study was to ascertain the usefulness of HK-2 cell line to inv estigate the properties and roles of PGP in proximal tubular cells. PGP exp ression in HK-2 cells was determined by immunoblotting analysis using the m onoclonal antibody C219. The activity of PGP was assessed by measuring the transport of the fluorescent probe Rhodamine 123 (R-123) in intact cell mon ostrates. The interactions of putative PGP modulators, including verapamil and cyclosporin A were also evaluated. Western blot revealed a C219 immunor eactive band of about 150 kDa consistent with the presence of PGP. HK-2 cel ls preloaded with R-123 rapidly effluxed the dye, the efflux being inhibite d by verapamil. Verapamil and, to a major extent cyclosporin A, significant ly increased R-123 intracellular accumulation. PGP immunoblottable amount w as increased when cells were cultured in the presence of either cyclosporin A or dexamethasone. The results suggest that the HK-2 cells, among the var ious differentiation features of proximal tubules, retain also the expressi on of a functional PGP in their membranes and that both PGP activity and ex pression may be modulated by drugs. Therefore, HK-2 line appears a suitable and promising tool for the study in vitro of renal transport processes dep endent on PGP.