title: Transport of fluorescein methotrexate by multidrug resistance-associated protein 3 in IEC-6 cells.
abstract: The transport characteristics of fluorescein methotrexate (F-MTX) were studied by using the rat intestinal crypt cell line IEC-6. Enhanced accumulation of F-MTX at 4 degrees C suggests the existence of an active efflux system. MK-571, an inhibitor of the multidrug resistance-associated protein/ATP binding cassette C (MRP/ABCC) family, also enhanced the accumulation of F-MTX. Transcellular transport of F-MTX from the apical to the basolateral compartment was 2.5 times higher than the opposite direction. This vectorial transport was also reduced by MK-571, indicating the presence of Mrp-type transporter(s) on the basolateral membrane. Mrp3 mRNA was readily detectable, and the protein was localized on the basolateral membrane. Uptake of FMTX into membrane vesicles from IEC-6 cells and Spodoptera frugiperda-9 cells expressing rat Mrp3 were both ATP dependent and saturable as a function of the F-MTX concentration. Similar Km values (11.0 +/- 1.8 and 4.5 +/- 1.1 microM) and inhibition profiles by MK-571, estradiol-17beta-d-glucuronide, and taurocholate for the ATP-dependent transport of F-MTX into these vesicles were obtained. These findings suggest that the efflux of F-MTX is mediated by Mrp3 on the basolateral membrane of IEC-6 cells.
authors.affiliation: Department of Biopharmaceutics, Graduate School of Pharmaceutical Sciences, Chiba University, Chiba 263-8522, Japan.
authors.forename: Tiesong
authors.initials: T
authors.lastname: Li
authors.forename: Kousei
authors.initials: K
authors.lastname: Ito
authors.forename: Toshiharu
authors.initials: T
authors.lastname: Horie
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000378:metabolism*; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D018528:ATP-Binding Cassette Transporters / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D002460:Cell Line; D005455:Fluorescent Antibody Technique; D007413:Intestinal Mucosa / Q000166:cytology / Q000378:metabolism*; D008727:Methotrexate / Q000031:analogs & derivatives* / Q000493:pharmacokinetics*; D051381:Rats; D018411:Spodoptera
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D018528:ATP-Binding Cassette Transporters; C036539:fluoresceinated methotrexate; C095122:multidrug resistance protein 3; D008727:Methotrexate