Formation and efflux of ATP-binding cassette transporter substrate 2,4-dinitrophenyl-S-glutathione from cultured human term placental villous tissue fragments.
Upon exposure to 1-chloro-2,4-dinitrobenzene (CDNB), the human placental tissue forms its glutathione conjugate 2,4-dinitrophenyl-S-glutathione (DNP-SG). The purpose of this study was to investigate the involvement of human placental ATP-binding cassette (ABC) transporters in the efflux of DNP-SG. Placental tissue samples were obtained from pregnant patients undergoing C-section deliveries following normal pregnancies; villous tissue was cultured in suspension, and DNP-SG formation and efflux upon exposure to 100 microM CDNB were measured by HPLC. DNP-SG efflux decreased by 69.1 (+/-11.3)%, 51.1 (+/-5.4)%, 56.7 (+/-8.3)% and 53.6 (+/-10.8)% (p < 0.05) in the presence of 5 mM sodium orthovanadate (ATPase inhibitor), 100 microM MK571 (MRP-inhibitor), 1 mM dipyridamole (BCRP/P-gp/MRP1-inhibitor) and 100 microM verapamil (P-gp/MRP1 inhibitor) respectively, without any change in DNP-SG formation, total tissue glutathione, GSH/GSSG ratio, tissue integrity or tissue viability. These data clearly established the role of ABC transporters in the human placental efflux of DNP-SG. To investigate the contribution of various ABC transporters toward DNP-SG transport, ATP-dependent transport of 3H-DNP-SG was determined in Sf9 membrane vesicles overexpressing P-gp, BCRP and the MRP proteins. MRP1-mediated DNP-SG transport was inhibited in the presence of sodium orthovanadate, MK571, dipyridamole and verapamil in the presence of glutathione. Furthermore, MRP1-mediated transport [K(t) = 11.3 +/- 1.3 microM and v(max) = 86.7 +/- 1.9 pmol/mg/min] was a high-affinity process compared to MRP2-mediated transport [K(t) = 168 +/- 7 microM and v(max) = 1367 +/- 18 pmol/mg/min]. The inhibition pattern and the kinetics of DNP-SG efflux in the placental villous tissue were consistent with MRP1-mediated DNP-SG efflux, suggesting a functional role and an apical localization for an MRP1-like transporter in the human placental syncytiotrophoblast.
Vaidya
Soniya S
SS
Departments of Pharmaceutics and Obstetrics & Gynecology, Virginia Commonwealth University, Medical College of Virginia Campus, Richmond, Virginia 23298-0533, USA.
Walsh
Scott W
SW
Gerk
Phillip M
PM
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters / Q000037:antagonists & inhibitors / Q000378:metabolism*; D000293:Adolescent; D000328:Adult; D001692:Biological Transport / Q000187:drug effects / Q000235:genetics; D015153:Blotting, Western; D004176:Dipyridamole / Q000494:pharmacology; D004976:Ethacrynic Acid / Q000494:pharmacology; D005260:Female; D005978:Glutathione / Q000031:analogs & derivatives* / Q000378:metabolism; D051549:Glutathione S-Transferase pi / Q000037:antagonists & inhibitors; D006801:Humans; D007700:Kinetics; D008875:Middle Aged; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D009363:Neoplasm Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism; D010920:Placenta / Q000187:drug effects / Q000378:metabolism*; D011247:Pregnancy; D011422:Propionates / Q000494:pharmacology; D011804:Quinolines / Q000494:pharmacology; D046509:Tissue Culture Techniques; D014700:Verapamil / Q000494:pharmacology; D055815:Young Adult
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000070997:ATP Binding Cassette Transporter, Subfamily G, Member 2; D018528:ATP-Binding Cassette Transporters; D004176:Dipyridamole; D004976:Ethacrynic Acid; D005978:Glutathione; D051549:Glutathione S-Transferase pi; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D009363:Neoplasm Proteins; D011422:Propionates; D011804:Quinolines; D014700:Verapamil; C000715087:ABCC2 protein, human; C116614:ABCG2 protein, human; C030190:S-(2,4-dinitrophenyl)glutathione; C059141:verlukast; C496556:GSTP1 protein, human; C421109:multidrug resistance-associated protein 1