title: Induction of human P-glycoprotein in Caco-2 cells: development of a highly sensitive assay system for P-glycoprotein-mediated drug transport.
abstract: The aim of this work is to develop a highly sensitive assay system for P-gp-mediated transport by using two methods, induction of P-gp and short-term culture of Caco-2 cells. To induce P-gp in Caco-2 cells, cells were cultured in vinblastine-containing medium. The mRNA level of P-gp was approximately 7-fold higher in Caco-2 cells cultured with vinblastine (P-gp-induced Caco-2 cells) than in control cells. Western blot analysis showed a significant increase in P-gp expression. After cell differentiation, the mRNA level of P-gp was downregulated, however, P-gp-induced Caco-2 cells still possessed a 5.6-fold higher mRNA level of P-gp compared to control cells. Polarized transport of substrate drugs was greater in the monolayer of P-gp-induced cells than in that of control cells. Moreover, we found that P-gp expression in Caco-2 cells could be further enhanced by applying the higher concentration of vinblastine. Transport activity of P-gp in Caco-2 cells cultured with higher concentration of vinblastine was markedly higher than that in P-gp-induced Caco-2 cells and was comparable with that in MDR1-MDCKII cells. In conclusion, this study provided a stable and highly sensitive in vitro assay system that can identify compounds that are subject to P-gp-mediated efflux.
authors.affiliation: Faculty of Pharmaceutical Sciences, Setsunan University, Hirakata, Osaka, Japan. 04d101sy@edu.setsunan.ac.jp
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Shirasaka
authors.forename: Masae
authors.initials: M
authors.lastname: Kawasaki
authors.forename: Toshiyasu
authors.initials: T
authors.lastname: Sakane
authors.forename: Hideaki
authors.initials: H
authors.lastname: Omatsu
authors.forename: Yuka
authors.initials: Y
authors.lastname: Moriya
authors.forename: Tsutomu
authors.initials: T
authors.lastname: Nakamura
authors.forename: Toshiyuki
authors.initials: T
authors.lastname: Sakaeda
authors.forename: Katsuhiko
authors.initials: K
authors.lastname: Okumura
authors.forename: Peter
authors.initials: P
authors.lastname: Langguth
authors.forename: Shinji
authors.initials: S
authors.lastname: Yamashita
mesh_terms: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics / Q000378:metabolism* / Q000502:physiology; D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000493:pharmacokinetics / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D015153:Blotting, Western; D018938:Caco-2 Cells; D002454:Cell Differentiation / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D048708:Cell Growth Processes / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D002460:Cell Line; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System / Q000235:genetics / Q000378:metabolism; D004077:Digoxin / Q000493:pharmacokinetics / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D008840:Microfilament Proteins / Q000235:genetics / Q000378:metabolism; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D000074011:Peptide Transporter 1; D004364:Pharmaceutical Preparations / Q000378:metabolism*; D011802:Quinidine / Q000493:pharmacokinetics / Q000494:pharmacology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D027981:Symporters / Q000235:genetics / Q000378:metabolism / Q000502:physiology; D014700:Verapamil / Q000493:pharmacokinetics / Q000494:pharmacology; D014747:Vinblastine / Q000493:pharmacokinetics / Q000494:pharmacology
chemical_list: D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000972:Antineoplastic Agents, Phytogenic; D051544:Cytochrome P-450 CYP3A; D003577:Cytochrome P-450 Enzyme System; D004077:Digoxin; D026901:Membrane Transport Proteins; D008840:Microfilament Proteins; D000089762:Multidrug Resistance-Associated Protein 2; D027425:ATP-Binding Cassette, Sub-Family C Proteins; D000074011:Peptide Transporter 1; D004364:Pharmaceutical Preparations; D011802:Quinidine; D012333:RNA, Messenger; D027981:Symporters; D014700:Verapamil; D014747:Vinblastine; C026615:villin; C510163:CYP3A4 protein, human; C421109:multidrug resistance-associated protein 1