MRP2, a human conjugate export pump, is present and transports fluo 3 intoapical vacuoles of Hep G2 cells

The multidrug resistance protein 2 (MRP2, symbol ABCC2) transports anionic conjugates and certain amphiphilic anions across the apical membrane of pol arized cells, Human hepatoma Hep G2 cells retain hepatic polarity and form apical vacuoles into which cholephilic substances are secreted. Immunofluor escence microscopy showed that human MRP2 was expressed in the apical vacuo le membrane of polarized Hep G2 cells, whereas the isoform MRP3 was localiz ed to the lateral membrane, Expression of both MRP2 and MRP3 was confirmed by immunoblotting and reverse transcription PCR. Flue 3 secretion into the apical vacuoles was inhibited by cyclosporin A but not by selective inhibit ors of multidrug resistance 1 P-glycoprotein. In addition, carboxyfluoresce in, rhodamine 123, and the fluorescent bile salt derivatives ursodeoxycholy l-(N epsilon-nitrobenzoxadiazolyl)-lysine and cholylglycylamido-fluorescein were secreted into the apical vacuoles; the latter two probably via the bi le salt export pump. We conclude that MRP2 mediates flue 3 secretion into t he apical vacuoles of polarized Hep G2 cells. Thus the function of human MR P2 and the action of inhibitors can be analyzed by the secretion of fluores cent anions such as fluo 3.