Sulfate conjugating and transport functions of MDCK distal tubular cells

Abstract

Background. Transfected Madin-Darby canine kidney (MDCK) cells (of distal tubular origin) have been used to study transport of organic anions. These cells have not been shown to possess sulfate-conjugating activity. Neither has transport activity been demonstrated in nontransfected MDCK cells. Methods. Polarized and monolayers of nontransfected MDCK type II cells were incubated with prototype substrates of phenolsulfotransferase (PST) and sodium sulfate in the absence or presence of known inhibitors of multidrug resistance protein (MRP): (3-3-(2-(7-chloro-2-quinionlinyl) ethenyl)phenyl)(3-dimethylamino-3-oxopropyl)thio)methyl)thio) propanoic acid (MK571), cyclosporin A (CsA), and probenecid. Effects of glutathione (GSH) and buthionine sulfoximine (BSO), potential modulators of the organic anion transporting protein/polypeptide (OATP) isoform, OATP1 were also examined. Sulfated conjugates were identified by high-performance liquid chromatography (HPLC)-radiometry or HPLC-fluorimetry. Results. Uptake, sulfate conjugation, and efflux of the sulfated conjugates of harmol, p-nitrophenol, N-acetyldopamine and acetaminophen were demonstrated. Activities in MDCK type II cells were higher than those in HepG2, human fetal liver, and Chang liver cells. A significant decrease in extracellular with a reciprocal increase in intracellular harmol sulfate was observed with MK571, CsA, and probenecid and with preloading of glutathione. Depletion of intracellular glutathione by BSO had the opposite effects. Conclusions. Normal (nontransfected) MDCK type II cells provide a suitable system for the study of the physiologic processes of uptake, sulfate conjugation, and transport of sulfated conjugates in kidney cells. Based on the action of specific inhibitors and modulators of MRP2 and OATP1, it was concluded that MRP2-like and OATP1-like transporters are possibly responsible for the transport of sulfated conjugates.