Abstract
Tetrahydroxy bile acids become major biliary bile acids in Bsep(−/−) mice and Fxr(−/−) mice fed cholic acid; we characterized disposition of these novel bile acids that also occur in patients with cholestasis. We investigated mouse Mrp2 (mMrp2) and P-glycoprotein [(P-gp) mMdr1a]-mediated transport of a tetrahydroxy bile acid, 6α-OH-taurocholic acid (6α-OH-TC), and its biliary excretion in wild-type and Mrp2(−/−) mice in the presence or absence of N-(4-[2-(1,2,3,4-tetrahydro-6,7-dimethoxy-2-isoquinolinyl)ethyl]-phenyl)-9,10-dihydro-5-methoxy-9-oxo-4-acridine carboxamide (GF120918), a P-gp and breast cancer resistance protein inhibitor. 6α-OH-TC was rapidly excreted into bile of wild-type mice (78% recovery); coinfusion of GF120918 had no significant effect. In Mrp2(−/−) mice, biliary excretion was decreased (52% recovery) and coinfusion of GF120918 further decreased these values (34% recovery). In wild-type, but not Mrp2(−/−), mice, 6α-OH-TC increased bile flow 2.5-fold. Membrane vesicle transport studies of 6α-OH-TC (0.05–0.75 mM) yielded saturation kinetics with a higher apparent affinity for mMrp2 (Km = 0.13 mM) than for mMdr1a (Km = 0.33 mM); mBsep transported 6α-OH-TC with positive cooperativity (Hill slope = 2.1). Human multidrug resistance-associated protein (MRP) 2 and P-gp also transported 6α-OH-TC but with positive cooperativity (Hill slope = 3.6 and 1.6, respectively). After intraileal administration, the time course of 6α-OH-TC biliary recovery was similar to that of coinfused taurocholate, implying that 6α-OH-TC can undergo enterohepatic cycling. Thus, Mrp2 plays a key role in 6α-OH-TC biliary excretion, whereas P-glycoprotein plays a secondary role; Bsep likely mediates excretion of 6α-OH-TC in the absence of Mrp2 and P-gp. In Bsep(−/−) mice, efficient synthesis of tetrahydroxy bile acids that are Mrp2 and P-gp substrates can explain the noncholestatic phenotype.
Footnotes
This work was supported in part by the United States Public Health Service [Grant HD 58299] (to M.V.); and a Grant-in-Aid for Scientific Research from the Ministry of Education, Science, Sports, and Culture of Japan [Grant 19510223] (to T.I.) for 2009–2011 and Research Project of the Institute of Natural Sciences Nihon University for 2009–2010.
Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
doi:10.1124/dmd.110.033480.
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ABBREVIATIONS:
- ABC
- ATP-binding cassette
- BSEP/Bsep
- bile salt export pump
- MRP/Mrp
- multidrug resistance-associated protein
- P-gp
- P-glycoprotein
- Bcrp
- breast cancer resistance protein
- PFIC2
- type 2 progressive familial intrahepatic cholestasis
- 12α-OH-TβMC
- taurine-conjugated 12α-hydroxy β-muricholic acid
- Fxr
- farnesoid X receptor
- mMrp2
- mouse Mrp2
- 6α-OH-TC
- 6α-OH-taurocholic acid
- MDR1
- multidrug resistance 1
- [3H]6α-OH-TC
- taurine-conjugated 22, 23-3H-3α, 6α,7α,12α-tetrahydroxy-5β-cholan-24-oic acid
- HPLC
- high-performance liquid chromatography
- GF120918
- N-(4-[2-(1,2,3,4-tetrahydro-6,7-dimethoxy-2-isoquinolinyl)ethyl]-phenyl)-9,10-dihydro-5-methoxy-9-oxo-4-acridine carboxamide
- mP-gp
- mouse P-gp
- EV
- empty vector
- mBsep
- mouse Bsep
- hMRP
- human MRP
- hMDR1
- human MDR1
- TUDC
- tauroursodeoxycholate
- ANOVA
- analysis of variance.
- Received March 25, 2010.
- Accepted July 19, 2010.
- Copyright © 2010 by The American Society for Pharmacology and Experimental Therapeutics
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