PURPOSE: P-Glycoprotein (Pgp) plays an important role in drug disposition and excretion in various tissues such as the brain, intestine, and kidney. Moreover, we have demonstrated that Pgp is expressed on the brush-border membranes of trophoblast cells in the placenta and restricts drug transfer from the maternal circulation to the fetus. However, the transport kinetics of physiologically expressed Pgp has scarcely been investigated. METHODS: In this study, we assessed the functional kinetics of transport mediated by Pgp that is physiologically expressed in normal tissue by using human placental brush-border membrane vesicles (BBMVs). Digoxin and vinblastine were used as typical substrates of Pgp. RESULTS: The uptakes of [3H]digoxin and [3H]vinblastine into BBMVs were significantly increased in the presence of an ATP-regenerating system. The ATP-dependent uptakes of [3H]digoxin and [3H]vinblastine into BBMVs exhibited saturable kinetics. The Michaelis constants (Kt values) were 2.65 +/- 1.80 microM and 21.9 +/- 3.37 microM, respectively. In the presence of a Pgp inhibitor such as verapamil, cyclosporine A, or progesterone, the ATP-dependent uptakes of [3H]digoxin and [3H]vinblastine into BBMVs were significantly reduced. Anti-Pgp monoclonal antibody C219 completely inhibited the uptake of [3H]digoxin. CONCLUSIONS: The transport kinetics of [3H]digoxin and [3H]vinblastine by physiologically expressed Pgp were successfully evaluated by using BBMVs prepared from normal human placenta. The present method enabled us to evaluate the function of physiologically expressed Pgp and is superior to the use of cultured transfectants in terms of the yield of vesicles. The present method may also be applicable to investigating the influence of various factors such as the genotype of the MDR1 gene or various pathophysiologic states of neonates on the function of Pgp.
PURPOSE:P-Glycoprotein (Pgp) plays an important role in drug disposition and excretion in various tissues such as the brain, intestine, and kidney. Moreover, we have demonstrated that Pgp is expressed on the brush-border membranes of trophoblast cells in the placenta and restricts drug transfer from the maternal circulation to the fetus. However, the transport kinetics of physiologically expressed Pgp has scarcely been investigated. METHODS: In this study, we assessed the functional kinetics of transport mediated by Pgp that is physiologically expressed in normal tissue by using human placental brush-border membrane vesicles (BBMVs). Digoxin and vinblastine were used as typical substrates of Pgp. RESULTS: The uptakes of [3H]digoxin and [3H]vinblastine into BBMVs were significantly increased in the presence of an ATP-regenerating system. The ATP-dependent uptakes of [3H]digoxin and [3H]vinblastine into BBMVs exhibited saturable kinetics. The Michaelis constants (Kt values) were 2.65 +/- 1.80 microM and 21.9 +/- 3.37 microM, respectively. In the presence of a Pgp inhibitor such as verapamil, cyclosporine A, or progesterone, the ATP-dependent uptakes of [3H]digoxin and [3H]vinblastine into BBMVs were significantly reduced. Anti-Pgp monoclonal antibody C219 completely inhibited the uptake of [3H]digoxin. CONCLUSIONS: The transport kinetics of [3H]digoxin and [3H]vinblastine by physiologically expressed Pgp were successfully evaluated by using BBMVs prepared from normal human placenta. The present method enabled us to evaluate the function of physiologically expressed Pgp and is superior to the use of cultured transfectants in terms of the yield of vesicles. The present method may also be applicable to investigating the influence of various factors such as the genotype of the MDR1 gene or various pathophysiologic states of neonates on the function of Pgp.
Authors: Y Nakamura; S Ikeda; T Furukawa; T Sumizawa; A Tani; S Akiyama; Y Nagata Journal: Biochem Biophys Res Commun Date: 1997-06-27 Impact factor: 3.575
Authors: M V St-Pierre; M A Serrano; R I Macias; U Dubs; M Hoechli; U Lauper; P J Meier; J J Marin Journal: Am J Physiol Regul Integr Comp Physiol Date: 2000-10 Impact factor: 3.619
Authors: Y Tanigawara; N Okamura; M Hirai; M Yasuhara; K Ueda; N Kioka; T Komano; R Hori Journal: J Pharmacol Exp Ther Date: 1992-11 Impact factor: 4.030
Authors: P Borst; A H Schinkel; J J Smit; E Wagenaar; L Van Deemter; A J Smith; E W Eijdems; F Baas; G J Zaman Journal: Pharmacol Ther Date: 1993-11 Impact factor: 12.310
Authors: F G Russel; P E van der Linden; W G Vermeulen; M Heijn; C H van Os; C A van Ginneken Journal: Biochem Pharmacol Date: 1988-07-01 Impact factor: 5.858
Authors: Sarah J Hemauer; Svetlana L Patrikeeva; Tatiana N Nanovskaya; Gary D V Hankins; Mahmoud S Ahmed Journal: Am J Obstet Gynecol Date: 2010-04 Impact factor: 8.661
Authors: Sarah J Hemauer; Svetlana L Patrikeeva; Xiaoming Wang; Doaa R Abdelrahman; Gary D V Hankins; Mahmoud S Ahmed; Tatiana N Nanovskaya Journal: Biochem Pharmacol Date: 2010-06-23 Impact factor: 5.858
Authors: Anand A Joshi; Soniya S Vaidya; Marie V St-Pierre; Andrei M Mikheev; Kelly E Desino; Abner N Nyandege; Kenneth L Audus; Jashvant D Unadkat; Phillip M Gerk Journal: Pharm Res Date: 2016-09-19 Impact factor: 4.200
Authors: Tatiana N Nanovskaya; Svetlana L Patrikeeva; Jonathan Paul; Maged M Costantine; Gary D V Hankins; Mahmoud S Ahmed Journal: Am J Obstet Gynecol Date: 2013-10 Impact factor: 8.661
Authors: Sarah J Hemauer; Svetlana L Patrikeeva; Tatiana N Nanovskaya; Gary D V Hankins; Mahmoud S Ahmed Journal: Biochem Pharmacol Date: 2009-07-08 Impact factor: 5.858
Authors: Sarah J Hemauer; Tatiana N Nanovskaya; Sherif Z Abdel-Rahman; Svetlana L Patrikeeva; Gary D V Hankins; Mahmoud S Ahmed Journal: Biochem Pharmacol Date: 2009-11-06 Impact factor: 5.858
Authors: Tatiana Nanovskaya; Svetlana Patrikeeva; Ying Zhan; Valentina Fokina; Gary D V Hankins; Mahmoud S Ahmed Journal: Am J Obstet Gynecol Date: 2012-08-04 Impact factor: 8.661