P-glycoprotein (Pgp), the MDR-encoded membrane transporter, is physiologically expressed in normal tissues with excretory functions, including kidney proximal tubules. In a preliminary report we have shown that HK-2, an immortalized cell line from normal human proximal tubule, expresses a functional Pgp and may be considered a valuable model for in vitro investigations on the Pgp role(s) in human renal pathophysiology. The present investigation was designed to further characterize the properties of HK-2 Pgp by exploring its responsiveness to a variety of exogenous or endogenous modulators. HK-2 cells were cultured in Dulbecco's modified Eagle's medium/Ham's F-12 supplemented with 5% FCS in the absence or in the presence of modulators. Pgp mRNA expression was studied by RTPCR and the amount of Pgp was determined by Western blotting. Pgp activity was assessed by intracellular rhodamine-123 (R-123) accumulation. RT-PCR showed that HK-2 cells express MDR-1, but not MDR-3. Both MDR-1 Pgp and MDR-1 mRNA were significantly increased in cells cultured in the presence of cyclosporin A (CsA), 1,25(OH)(2)D-3, platelet activating factor, dexamethasone (Dex), or aldosterone. Verapamil (Vp), cimetidine, and trimethoprim did not affect HK-2 Pgp expression. Conversely, 2-acetylaminofluorene strongly downregulated Pgp expression. Vp, CsA, 1,25(OH)2D3 and Dex significantly increased R-123 intracellular retention, indicating the inhibition of Pgp-mediated transport. Drug-pretreated, Pgp-over-expressing cells showed increased Pgp activity and were less susceptible to toxic concentrations of CsA. MDR-1 Pgp in HK-2 appears to be responsive to many compounds, including classical Pgp inhibitors and putative physiological substrates, but some of its pharmacological properties are different from those described in other experimental, in particular nonhuman, cell models. (C) 2002 Elsevier Science (USA).
|Autori:||Romiti N; Tramonti G; Chieli E|
|Titolo:||Influence of different chemicals on MDR-1 P-glycoprotein expression and activity in the HK-2 proximal tubular cell line|
|Anno del prodotto:||2002|
|Digital Object Identifier (DOI):||10.1006/taap.2002.9461|
|Appare nelle tipologie:||1.1 Articolo in rivista|