Region-Dependent Modulation of Intestinal Permeability by Drug Efflux Transporters: In Vitro Studies in mdr1a(-/-) Mouse Intestine R. H. STEPHENS, J. TANIANIS-HUGHES, N. B. HIGGS, M. HUMPHREY, and G. WARHURST Gut Barrier Group, University of Manchester and Salford Hospitals Trust, Hope Hospital, Salford, (R.H.S., J.T.-H., N.B.H., G.W.), and Pharmaceutical Sciences, Pfizer Global Research & Development, Kent, United Kingdom (M.H.) Received July 9, 2002; accepted August 27, 2002 ABSTRACT Information on the extent to which xenobiotics interact with P-glycoprotein (PGP) during transit through the intestine is crucial in determining the influence of PGP on oral drug ab- sorption. We have recently described a novel use of isolated ileum from PGP-deficient mdr1a(-/-) mice to resolve PGP- and non-PGP-dependent drug efflux and provide a definitive measure of intrinsic drug permeability without recourse to in- hibitors (Stephens et al., 2002). The present study uses this approach to investigate the impact of PGP on intestinal per- meability of paclitaxel and digoxin in different regions of the mouse intestine (jejunum, ileum, and proximal and distal colon). Absorption of paclitaxel and digoxin in tissues from wild-type mice was low and showed little regional variation. In contrast, absorption of both drugs was markedly higher in mdr1a(-/-) intestine, although the increase was highly region-dependent, with the ileum and distal colon showing the greatest effect and much smaller changes in the jejunum and proximal colon. These effects were accompanied by the abolition of paclitaxel and digoxin secretion in mdr1a(-/-) mice, suggesting that regional variations in intestinal permeability are masked by differential PGP expression, confirmed by immunoblotting studies. Propranolol permeability, which is not influenced by PGP, showed similar regional variation in both wild-type and mdr1a(-/-) tissues, suggesting that differences are at the level of transcellular permeability. These data suggest that the ileum and the distal colon are regions of relatively high transcellular permeability for xenobiotics that are compensated by en- hanced expression of PGP. The xenobiotic transporter P-glycoprotein (PGP), the pro- tein product of the MDR1 gene, was initially associated with the development of multidrug resistance in tumor cells (Hunter and Hirst, 1997). It is now understood that this ABC protein forms an integral part of the intestinal barrier to- gether with several other polyspecific efflux transporters, including members of the multidrug resistance-related pro- tein (MRP) family and breast cancer resistance protein (BCRP) (Hunter and Hirst, 1997; Makhey et al., 1998; Gotoh et al., 2000; Jonker et al., 2000; Litman et al., 2001). The ability of PGP to interact with a broad range of natural and synthetic xenobiotics, drugs, and naturally occurring toxins or food constituents (Hunter and Hirst, 1997; Walle and Walle, 1999; Litman et al., 2001) strongly suggests that its ability to limit oral drug absorption is part of a broader protective role as a modulator of intestinal permeability. Indeed, growing awareness of the wider physiological context in which PGP resides has linked this and other efflux trans- port proteins to the regulation of electrolyte transport, apo- ptosis, and inflammation (Johnstone et al., 2000). Clearer definition of these physiological and pharmacolog- ical roles, especially those directly related to xenobiotic transport, requires a better understanding of the extent to which compounds interact with PGP as they move sequen- tially along the cephalocaudal axis of the gut. This will de- pend on how transporter activity varies along the gut and whether non-PGP transporters contribute to efflux. MRP and BCRP can interact with PGP substrates and appear to be expressed heterogeneously in the intestine (Jonker et al., 2000; Mottino et al., 2000; Litman et al., 2001; Maliepaard et al., 2001). Several other variables including membrane com- position and passive epithelial permeability may also be im- portant in determining which xenobiotics are transported effectively (Brasitus and Schachter, 1984; Fagerholm et al., 1997; Ungell et al., 1998). The way in which the activity of PGP and other efflux This work was supported by Pfizer Global Research and Development. Article, publication date, and citation information can be found at http://jpet.aspetjournals.org. DOI: 10.1124/jpet.102.041236. ABBREVIATIONS: PGP, P-glycoprotein; mdr1, multidrug resistance protein 1; MRP, multidrug resistance-associated protein; BCRP, breast cancer resistance protein; I SC , short-circuit current; R T , transepithelial electrical resistance; A-B, apical-to-basolateral; B-A, basolateral-to-apical; P app , apparent permeability; TBS-T, Tris-buffered saline/Tween 20; ANOVA, analysis of variance; WT, wild-type. 0022-3565/02/3033-1095–1101$7.00 THE JOURNAL OF PHARMACOLOGY AND EXPERIMENTAL THERAPEUTICS Vol. 303, No. 3 Copyright © 2002 by The American Society for Pharmacology and Experimental Therapeutics 41236/1025747 JPET 303:1095–1101, 2002 Printed in U.S.A. 1095 at ASPET Journals on February 23, 2016 jpet.aspetjournals.org Downloaded from