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Lookup NU author(s): Jamie Wright,
Professor Nicholas Simmons
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In order to determine the capacity and regulation of the breast cancer resistance protein (BCRP)-mediated transport in intact human intestinal epithelial monolayers (Caco-2) in which multiple ABC transporters are expressed, nitrofurantoin has been used as a selective transported substrate. Nitrofurantoin transepithelial secretion was confirmed in both human BCRP and mouse bcrp-transfected MDCKII epithelia, whereas no net transepithelial secretion was observed in native or human MDR1-MDCKII epithelia. Furthermore, nitrofurantoin transepithelial secretion by BCRP-MDCKII monolayers was inhibited by Ko143 (10 mu M), but not verapamil (100 mu M). In Caco-2 cells grown upon permeable supports, nitrofurantoin displayed a dose-dependent transepithelial secretion with an apparent Km = 69.41 +/- 22.3 mu M and Vmax = 14.03 +/- 2.27 nmol/(cm(2).h). Net nitrofurantoin transepithelial secretion by Caco-2 epithelia was inhibited 92% by 10 mu M Ko143. Regulation of expression and function of BCRP in Caco-2 epithelial monolayers was determined after 72-h pre-exposure of the monolayers to a number of potential inducing agents. Quantitative real-time PCR and Western blotting were used to correlate induction of BCRP transcript and protein levels with transport activity. 72-h pre-treatment with beta-napthoflavone and rosiglitazone up-regulates BCRP mRNA and protein expression and transport of nitrofurantoin. Ko143-sensitive transepithelial secretion of the bi-substrate (MDR1/BCRP) prazosin was also increased in the presence of rosiglitazone. We conclude that nitrofurantoin may be used to unambiguously measure BCRP-mediated fluxes in Caco-2 epithelial layers. Since dynamic regulation of BCRP expression and function is retained, the Caco-2 cell-line is useful as a screen for drug-drug and drug-diet interactions mediated by BCRP. (C) 2011 Elsevier B.V. All rights reserved.
Author(s): Wright JA, Haslam LS, Coleman T, Simmons NL
Publication type: Article
Publication status: Published
Journal: European Journal of Pharmacology
Print publication date: 10/10/2011
ISSN (print): 0014-2999
ISSN (electronic): 1879-0712
Publisher: Elsevier BV
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