The multidrug resistance protein Mrp2 is an ATP-binding cassette (ABC) transporter mainly expressed in liver, kidney, and intestine. One of the physiological roles of Mrp2 is to transport bilirubin glucuronides from the liver into the bile. Current in vivo models to study Mrp2 are the transporter-deficient and Eisai hyperbilirubinemic rat strains. Previous reports showed hyperbilirubinemia and induction of Mrp3 in the hepatocyte sinusoidal membrane in the mutant rats. In addition, differences in liver cytochrome P450 and UGT1a levels between wild-type and mutant rats were detected. To study whether these compensatory mechanisms were specific to rats, we characterized Mrp2(-/-) mice. Functional absence of Mrp2 in the knockout mice was demonstrated by showing increased levels of bilirubin and bilirubin glucuronides in serum and urine, a reduction in biliary excretion of bilirubin glucuronides and total glutathione, and a reduction in the biliary excretion of the Mrp2 substrate dibromosulfophthalein. To identify possible compensatory mechanisms in Mrp2(-/-) mice, the expression levels of 98 phase I, phase II, and transporter genes were compared in liver, kidney, and intestine of male and female Mrp2(-/-) and control mice. Unlike in Mrp2 mutant rats, no induction of Mrp3 in Mrp2(-/-) mice was detected. However, Mrp4 mRNA and protein in liver and kidney were increased approximately 6- and 2-fold, respectively. Phenotypic analysis of major cytochrome P450-mediated activities in liver microsomes did not show differences between wild-type and Mrp2(-/-) mice. In conclusion, Mrp2(-/-) mice are a new valuable tool to study the role of Mrp2 in drug disposition.
"Breeder pairs of Mrp2−/− mice  were obtained from Taconic (Germantown, NY) and were mated in our animal care facility. Friend Virus B (FVB) mice, which were used as control mice, were obtained from Charles River Laboratories (Wilmington, MA) and mated in our facility. "
[Show abstract][Hide abstract] ABSTRACT: Multidrug resistance-associated proteins (MRP) 2 and 4 are localized in proximal tubular epithelial cells and participate in the renal elimination of xenobiotics. MRP2 has also been implicated in the renal and hepatic elimination of mercury. The current study tested the hypothesis that MRP2 and MRP4 are involved in renal and hepatic handling of inorganic mercury (Hg(2+)). We examined the disposition of Hg(2+) in Mrp2(-/-) mice and assessed the transport of mercuric conjugates in inside-out membrane vesicles containing human MRP4. Since MRP2 has been shown to utilize glutathione (GSH) for transport of select substrates, we examined renal concentrations of GSH and cysteine and the expression of glutamate cysteine ligase (GCL) in Mrp2(-/-) and FVB mice. The effect of Hg(2+) exposure on renal GSH levels was also assessed in these mice. Our data suggest that MRP2, but not MRP4, is involved in proximal tubular export of Hg(2+). In addition, GSH levels are greater in Mrp2(-/-) mice and exposure to Hg(2+) reduced renal levels of GSH. Expression of GCL was also altered in Mrp2(-/-) mice under normal conditions and following exposure to HgCl2. This study provides important novel data regarding the transport of Hg(2+) and the effect of Hg(2+) exposure on GSH levels.
PLoS ONE 09/2013; 8(9):e73559. DOI:10.1371/journal.pone.0073559 · 3.23 Impact Factor
"MRP2, MRP3 and MRP4 are involved in transport of bile acids and/or bilirubin in the liver and kidney , . MRP2 is localized at the canalicular membrane of hepatocytes and MRP2-null mice show increased serum conjugated bilirubin , , indicating that MRP2 plays a role in biliary excretion of conjugated bilirubin. MRP3 and MRP4 are expressed at the basolateral hepatocyte membrane . "
[Show abstract][Hide abstract] ABSTRACT: The vitamin D receptor (VDR) mediates the physiological and pharmacological actions of 1α,25-dihydroxyvitamin D(3) in bone and calcium metabolism, cellular growth and differentiation, and immunity. VDR also responds to secondary bile acids and belongs to the NR1I subfamily of the nuclear receptor superfamily, which regulates expression of xenobiotic metabolism genes. When compared to knockout mouse investigations of the other NR1I nuclear receptors, pregnane X receptor and constitutive androstane receptor, an understanding of the role of VDR in xenobiotic metabolism remains limited. We examined the effect of VDR deletion in a mouse model of cholestasis. We performed bile duct ligation (BDL) on VDR-null mice and compared blood biochemistry, mRNA expression of genes involved in bile acid and bilirubin metabolism, cytokine production, and expression of inflammatory regulators with those of wild-type mice. VDR-null mice had elevated plasma conjugated bilirubin levels three days after BDL compared with wild-type mice. Urine bilirubin levels and renal mRNA and/or protein expression of multidrug resistance-associated proteins 2 and 4 were decreased in VDR-null mice, suggesting impaired excretion of conjugated bilirubin into urine. While VDR-null kidney showed mRNA expression of interleukin-6 (IL-6) after BDL and VDR-null macrophages had higher IL-6 protein levels after lipopolysaccharide stimulation, the induction of intestinal Il6 mRNA expression and plasma IL-6 protein levels after BDL was impaired in VDR-null mice. Immunoblotting analysis showed that expression of an immune regulator, IκBα, was elevated in the jejunum of VDR-null mice, a possible mechanism for the attenuated induction of Il6 expression in the intestine after BDL. Increased expression of IκBα may be a consequence of compensatory mechanisms for VDR deletion. These results reveal a role of VDR in bilirubin clearance during cholestasis. VDR is also suggested to contribute to tissue-selective immune regulation.
PLoS ONE 12/2012; 7(12):e51664. DOI:10.1371/journal.pone.0051664 · 3.23 Impact Factor
"Depending on the background strain (FVB versus C57BL/6) in Abcc2(Ϫ/Ϫ) mice, Abcc3 and Abcc4 expression in kidney and liver ranged from no different than WT to 2-fold elevation (Chu et al., 2006; Vlaming et al., 2006). Similar expression data are not available in the Abcc3(Ϫ/Ϫ) strain, but given that this group showed the greatest compensatory increase in clearance at the 50 mg/kg dose level, it suggests altered transporter expression underlies the higher elimination capacity. "
[Show abstract][Hide abstract] ABSTRACT: Methotrexate (MTX) is a substrate for numerous human ATP-binding cassette (ABC) efflux transporters, yet the impact of these transporters on MTX pharmacokinetics (PK) over a large dose range has not been examined. To investigate the effects of two transporters-ABC subfamily C member 2 (Abcc2; multidrug resistance protein 2) and ABC subfamily C member 3 (Abcc3; multidrug resistance protein 3)-involved in MTX hepatobiliary disposition in vivo, MTX plasma, urine, and feces concentrations were analyzed after 10, 50, and 200 mg/kg i.v. doses to groups of wild type (WT), Abcc2(-/-), and Abcc3(-/-) mice. The absence of Abcc2 caused a decrease in total clearance of MTX relative to WT mice at all dose levels yet was accompanied by compensatory increases in renal excretion and metabolism to 7-hydroxymethotrexate (7OH-MTX). In Abcc3(-/-) mice, total clearance was elevated at the two lower dose levels and was attributed to stimulation of biliary excretion and confirmed by elevated fecal excretion; however, at the high 200 mg/kg dose, clearance was severely retarded and could be attributed to hepatotoxicity because conversion to 7OH-MTX was diminished. The findings confirmed that both Abcc2 and Abcc3 significantly influenced the PK properties of MTX, and depending on the MTX dose and strain, alternate elimination pathways were elicited and saturable.
Drug metabolism and disposition: the biological fate of chemicals 08/2011; 39(11):2155-61. DOI:10.1124/dmd.111.041228 · 3.25 Impact Factor
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