Molecular and functional expression of multidrug resistance-associated protein-1 in primary cultured rat alveolar epithelial cells.
Multidrug resistance-associated protein-1 (MRP1) is an integral membrane efflux protein that is implicated in multidrug resistance in cancer, but it is also expressed in normal tissues. The objective of this study was to determine the expression, localization and functional activity of MRP1 in primary cultured rat alveolar epithelial cells of types I- and II cell-like phenotypes. RT-PCR data showed 550-base pair fragments in both types I- and II-like pneumocytes that exhibited 99% identity to the rat MRP1 isoform. Significant levels of MRP1 protein were detected by western analysis of immunoprecipitates in both cell types, and immunofluorescence combined with confocal laser scanning microscopy indicated basolateral localization of MRP1. Indomethacin (0-100 microM) increased fluorescein basolateral-to-apical transport, and accumulation of fluorescein in the cells, in a dose-dependent manner. We therefore conclude that the MRP1 gene is present in primary cultured rat epithelial cells of both types I- and II-like phenotypes and its corresponding protein (MRP1) is localized in the basolateral membrane of these cells. Primary cultured monolayers of rat type II-like pneumocytes appear to be a useful tool for screening MRP1 substrates designed for pulmonary delivery/targeting.
Patel
Leena N
LN
Department of Pharmacology and Pharmaceutical Sciences, University of Southern California, Los Angeles, California 90089-9121, USA.
Uchiyama
Tomomi
T
Kim
Kwang-Jin
KJ
Borok
Zea
Z
Crandall
Edward D
ED
Shen
Wei-Chiang
WC
Lee
Vincent H L
VH
D000818:Animals; D015153:Blotting, Western; D018929:Cell Culture Techniques; D002462:Cell Membrane / Q000378:metabolism; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism*; D019793:Fluorescein / Q000378:metabolism; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes / Q000378:metabolism; D005838:Genotype; D007213:Indomethacin / Q000494:pharmacology; D008297:Male; D018613:Microscopy, Confocal; D027425:ATP-Binding Cassette, Sub-Family C Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D010641:Phenotype; D011650:Pulmonary Alveoli / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D020133:Reverse Transcriptase Polymerase Chain Reaction
D019793:Fluorescein; D005456:Fluorescent Dyes; D007213:Indomethacin; D027425:ATP-Binding Cassette, Sub-Family C Proteins; C421109:multidrug resistance-associated protein 1