Contribution of Multidrug Resistance Protein 2 (MRP2/ABCC2) to the Renal Excretion of p-aminohippurate (PAH) and Identification of MRP4 (ABCC4) as a Novel PAH Transporter

Abstract

Abstract. p-Aminohippurate (PAH) is the classical substrate used in the characterization of organic anion transport in renal proximal tubular cells. Although basolateral transporters for PAH uptake from blood into the cell have been well characterized, there is still little knowledge on the apical urinary efflux transporters. The multidrug resistance protein 2 (MRP2/ ABCC2) is localized to the apical membrane and mediates ATP-dependent PAH transport, but its contribution to urinary PAH excretion is not known. In this report, we show that renal excretion of PAH in isolated perfused kidneys from wild-type and Mrp2-deficient (TR Ϫ ) rats is not significantly different. Uptake of [14 C]PAH in membrane vesicles expressing two different MRP2 clones isolated from Sf9 and MDCKII cells exhibited a low affinity for PAH (Sf9, 5 Ϯ 2 mM; MDCKII, 2.1 Ϯ 0.6 mM). Human MRP4 (ABCC4), which has recently been localized to the apical membrane, expressed in Sf9 cells had a much higher affinity for PAH (K m ϭ 160 Ϯ 50 M). Various inhibitors of MRP2-mediated PAH transport also inhibited MRP4. Probenecid stimulated MRP2 at low concentrations but had no effect on MRP4; but at high probenecid concentrations, both MRP2 and MRP4 were inhibited. Sulfinpyrazone only stimulated MRP2, but inhibited MRP4. Realtime PCR and Western blot analysis showed that renal cortical expression of MRP4 is approximately fivefold higher as compared with MRP2. MRP4 is a novel PAH transporter that has higher affinity for PAH and is expressed more highly in kidney than MRP2, and may therefore be more important in renal PAH excretion.

Chalcogenopyrylium Dyes as Differential Modulators of Organic Anion Transport by Multidrug Resistance Protein 1 (MRP1), MRP2, and MRP4

“…Previous studies have identified several organic anions that inhibit one MRP transporter while stimulating another, leading to the conclusion that some MRPs contain at least two interacting ligand binding sites (Bakos et al, 2000;Zelcer et al, 2003;Wittgen et al, 2011). For example, both sulfinpyrazone and probenicid inhibit MRP1 and MRP4 but stimulate MRP2 (Bakos et al, 2000;Ito et al, 2001a;Smeets et al, 2004;Huisman et al, 2005). Consequently, the opposite effects observed for CGP V-4 and V-6 on MRP1 (inhibitory) versus MRP2 (stimulatory) are in themselves not surprising.…”

Regulatory Pathways for ATP-binding Cassette Transport Proteins in Kidney Proximal Tubules

“…The wide range of drug substrates includes antibiotics, antivirals, and diuretics (8). In comparison to MRP2, MRP4 exhibits a higher expression in the kidney and a higher affinity for smaller organic anions (25). Polymorphisms in ABCC2 and ABCC4, leading to diminished MRP2 or MRP4 activity, respectively, are associated with increased sensitivity to nephrotoxicity or delayed graft function after kidney transplantation (26,27).…”

Multidrug Resistance–Associated Protein 4 Is Involved in the Urinary Excretion of Hydrochlorothiazide and Furosemide

“…Both transporters accept a variety of organic anions as substrates, such as p-aminohippurate (PAH), 17␤-estradiol-17␤-d-glucuronide, dehydroepiandrosterone sulfate (DHEAS), and methotrexate (18,(25)(26)(27). van Aubel et al (18) initially found that Mrp4 is localized on the BBM, and, subsequently, Mrp4 was suggested to play important roles in tubular secretion (18,25,28), although direct evidence of this still is lacking. For BCRP, an in vivo study using Bcrp knockout mice has demonstrated that BCRP is responsible for the urinary excretion of some organic anions, such as methotrexate and 6-hydroxy-5,7-dimethyl-2-methylamino-4-(3-pyridylmethyl) benzothiazole (E3040) sulfate (29,30).…”