Abstract
The effect of the cholestatic estrogens ethynylestradiol (EE) and estradiol 17β-d-glucuronide (E2-17G) on expression and activity of intestinal multidrug resistant-associated protein 2 (Mrp2, Abcc2) was studied in rats. Expression and localization of Mrp2 were evaluated by Western blotting, real-time polymerase chain reaction, and confocal immunofluorescence microscopy. Mrp2 transport activity toward dinitrophenyl-S-glutathione (DNP-SG) was assessed in vitro in intestinal sacs. EE, administered subcutaneously at a 5 mg/kg b.wt. dose, for 5 consecutive days, produced a marked decrease in Mrp2 expression at post-transcriptional level, without affecting its normal localization at the apical membrane of the enterocyte. This effect was selective because expression of other ATP-binding cassette proteins such as breast cancer resistance protein and Mrp3 were not affected and that of multidrug resistance protein 1 was only minimally impaired. Consistent with down-regulation of expression of Mrp2, a significant impairment in serosal to mucosal transport of DNP-SG and in protection against absorption of this same compound were registered. Simultaneous administration of EE with spironolactone (200 μmol/kg b.wt./day for 3 days), an Mrp2 inducer, prevented these alterations, confirming down-regulation of expression of Mrp2 by EE as a major component of functional changes. Incorporation of E2-17G (30 μM) in the serosal medium of intestinal sacs decreased serosal to mucosal transport of DNP-SG, probably because of competitive inhibition, without affecting normal Mrp2 expression or localization. Our data indicate impairment of function of intestinal Mrp2 by both cholestatic estrogens, although through a different mechanism. This finding represents an aggravation of deteriorated hepatic Mrp2 function that could further increase bioavailability of specific xenobiotics after oral exposure.
Footnotes
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This work was supported in part by the Agencia Nacional de Promoción Científica y Tecnológica [Grant PICT No. 05-26306]; Consejo Nacional de Investigaciones Científicas y Técnicas [Grant PIP No. 6442]; and Universidad Nacional de Rosario, Argentina.
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.108.025643.
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ABBREVIATIONS: Mdr, multidrug resistance protein; Bcrp, breast cancer resistance protein; Mrp, multidrug resistance-associated protein; PHIP, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine; EE, ethynylestradiol; E2-17G, estradiol 17β-d-glucuronide; SL, spironolactone; CDNB, 1-chloro-2,4-dinitrobenzene; DNP-SG, dinitrophenyl-S-glutathione; ALP, alkaline phosphatase; BBM, brush border membrane; PCR, polymerase chain reaction; GST, glutathione S-transferase; ZO-1, zonula occludens 1; ABC, ATP-binding cassette; C, control.
- Received November 14, 2008.
- Accepted March 17, 2009.
- The American Society for Pharmacology and Experimental Therapeutics
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