Multidrug resistance-associated protein-1 functional activity in Calu-3 cells

The purpose of this work was to determine whether the in vitro bronchiolar epithelial cell model, Calu-3, possesses efflux pump activity by the multid rug resistance-associated protein-1 (MRP1). Reverse transcription-polymeras e chain reaction demonstrated MRP1 gene expression in Calu-3 cells. Indirec t fluorescence studies showed a basolateral membrane localization of MRP1 c ompared with P-glycoprotein (Pgp) that was found on the apical side of thes e cells. An increase in the rate of accumulation of the MRP1 substrate calc ein was observed following treatment with the organic anion/MRP1 inhibitor indomethacin, the Pgp inhibitors cyclosporin A (CsA) and vinblastine, as we ll as conditions of energy depletion. Total calcein efflux was significantl y decreased with the MRP1 inhibitors probenecid and indomethacin, while tot al efflux was unchanged following treatment with CsA. In the latter case, h owever, intracellular calcein levels postefflux were significantly greater. Probenecid and indomethacin increased calcein net secretion 2.4- and 3.5-f old, respectively. The efflux of etoposide, a known substrate for both Pgp and MRP1, was shown to be mainly Pgp-mediated by using the multidrug-resist ant inhibitors quinidine (mixed Pgp/MRP1), CsA (Pgp), and MK571 (MRP1). Tog ether, these data suggest that Calu-3 cells possess MRP1 functional activit y that is subordinate to Pgp efflux. We present here kinetic analysis of ca lcein eff lux from Calu-3 cells to support our findings.