Sulfate conjugating and transport functions of MDCK distal tubular cells.
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.
Ng
Kian Hong
KH
Department of Biochemistry, Faculty of Medicine, National University of Singapore, Kent Ridge, Singapore, Singapore.
Lim
Beng Gek
BG
Wong
Kim Ping
KP
D000818:Animals; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D019328:Buthionine Sulfoximine / Q000494:pharmacology; D006528:Carcinoma, Hepatocellular; D045744:Cell Line, Tumor; D016572:Cyclosporine / Q000494:pharmacology; D005978:Glutathione / Q000031:analogs & derivatives* / Q000494:pharmacology; D006247:Harmine / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D006801:Humans; D007166:Immunosuppressive Agents / Q000494:pharmacology; D007686:Kidney Tubules, Distal / Q000166:cytology / Q000378:metabolism*; D020024:Leukotriene Antagonists / Q000494:pharmacology; D008099:Liver / Q000166:cytology / Q000378:metabolism; D026901:Membrane Transport Proteins; D008727:Methotrexate / Q000494:pharmacology; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000378:metabolism; D027362:Organic Anion Transport Protein 1 / Q000378:metabolism; D011339:Probenecid / Q000494:pharmacology; D011422:Propionates / Q000494:pharmacology; D011804:Quinolines / Q000494:pharmacology; D011837:Radiation-Protective Agents / Q000494:pharmacology; D011838:Radiation-Sensitizing Agents / Q000494:pharmacology; D013431:Sulfates / Q000378:metabolism*; D014528:Uricosuric Agents / Q000494:pharmacology
D019328:Buthionine Sulfoximine; D016572:Cyclosporine; D005978:Glutathione; D006247:Harmine; D007166:Immunosuppressive Agents; D020024:Leukotriene Antagonists; D026901:Membrane Transport Proteins; D008727:Methotrexate; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D027362:Organic Anion Transport Protein 1; D011339:Probenecid; D011422:Propionates; D011804:Quinolines; D011837:Radiation-Protective Agents; D011838:Radiation-Sensitizing Agents; D013431:Sulfates; D014528:Uricosuric Agents; C000715087:ABCC2 protein, human; C104826:harmol sulfate; C059141:verlukast; C042431:S-ethyl glutathione