The urate-anion exchanger URAT1 is a member of the organic anion transporter (OAT) family that regulates blood urate level in humans and is targeted by uricosuric and antiuricosuric agents (Enomoto, A., Kimura, H., Chairoungdua, A., Shigeta, Y., Jutabha, P., Cha, S. H., Hosoyamada, M., Takeda, M., Sekine, T., Igarashi, T., Matsuo, H., Kikuchi, Y., Oda, T., Ichida, K., Hosoya, T., Shimotaka, K., Niwa, T., Kanai, Y., and Endou, H. (2002) Nature 417, 447-452). URAT1 is expressed only in the kidney, where it is thought to participate in tubular urate reabsorption. We found that the multivalent PDZ (PSD-95, Drosophila discs-large protein, Zonula occludens protein 1) domain-containing protein, PDZK1 interacts with URAT1 in a yeast two-hybrid screen. Such an interaction requires the PDZ motif of URAT1 in its extreme intracellular C-terminal region and the first, second, and fourth PDZ domains of PDZK1 as identified by yeast two-hybrid assay, in vitro binding assay and surface plasmon resonance analysis (K D ‫؍‬ 1.97-514 nM). Coimmunoprecipitation studies revealed that the wildtype URAT1, but not its mutant lacking the PDZ-motif, directly interacts with PDZK1. Colocalization of URAT1 and PDZK1 was observed at the apical membrane of renal proximal tubular cells. The association of URAT1 with PDZK1 enhanced urate transport activities in HEK293 cells (1.4-fold), and the deletion of the URAT1 C-terminal PDZ motif abolished this effect. The augmentation of the transport activity was accompanied by a significant increase in the V max of urate transport via URAT1 and was associated with the increased surface expression level of URAT1 protein from HEK293 cells stably expressing URAT1 transfected with PDZK1. Taken together, the present study indicates the novel role of PDZK1 in regulating the functional activity of URAT1-mediated urate transport in the apical membrane of renal proximal tubules.Urate is the major inert end product of purine degradation in humans and higher primates in contrast to most other mammals because of the genetic silencing of hepatic oxidative enzyme uricase (1, 2). The kidney plays a dominant role in urate elimination; it excretes ϳ70% of the daily urate production. Urate exists primarily as a weak acid at physiological pH (pK a 5.75), and most of it is dissociated in blood and is freely filtered through the glomerulus. Thus, urate enters the proximal tubule in its anionic form, but it hardly permeates the tubular cells in the absence of facilitated mechanisms owing to its hydrophilicity. The transport mechanisms for urate are localized in the proximal tubule. In humans, urate is almost completely reabsorbed, which results in the excretion of ϳ10% of its filtered load. The absence of uricase and the presence of an effective renal urate reabsorption system contribute to higher blood urate levels in humans. Therefore, it was postulated that defects in tubular urate transport cause hypouricemia and decreased renal urate clearance leads to hyperuricemia in most hyperuricemic patients (3).Recently, we have id...

Abstract. Human organic anion transporter OAT4 is expressed in the kidney and placenta and mediates high-affinity transport of estrone-3-sulfate (E 1 S). Because a previous study demonstrated no trans-stimulatory effects by E 1 S, the mode of organic anion transport via OAT4 remains still unclear. In the present study, we examined the driving force of OAT4 using mouse proximal tubular cells stably expressing OAT4 (S 2 OAT4). OAT4-mediated E 1 S uptake was inhibited by glutarate (GA) (IC 50 : 1.25 mM) and [14 C]GA uptake via S 2 OAT4 was significantly trans-stimulated by unlabeled GA (5 mM) (P<0.001).[3 H]E 1 S uptake via S 2 OAT4 was significantly trans-stimulated by preloaded GA (P<0.001) and its [ 14 C]GA efflux was significantly trans-stimulated by unlabeled E 1 S in the medium (P<0.05). In additon, both the uptake and efflux of [14 C]p-aminohippuric acid (PAH) and [14 C]GA via S 2 OAT4 were significantly trans-stimulated by unlabeled GA or PAH. The immunoreactivities of OAT4 were observed in the apical membrane of proximal tubules along with those of basolateral organic anion / dicarboxylate exchangers such as hOAT1 and hOAT3 in the same tubular population. These results indicate that OAT4 is an apical organic anion / dicarboxylate exchanger and mainly functions as an apical pathway for the reabsorption of some organic anions in renal proximal tubules driven by an outwardly directed dicarboxylate gradient.

The purpose of this study was to elucidate the interactions of human organic anion transporters (hOATs) and human organic cation transporters (hOCTs) with nonsteroidal anti-inflammatory drugs (NSAIDs) using cells stably expressing hOATs and hOCTs. NSAIDs tested were acetaminophen, acetylsalicylate, salicylate, diclofenac, ibuprofen, indomethacin, ketoprofen, mefenamic acid, naproxen, piroxicam, phenacetin, and sulindac. These NSAIDs inhibited organic anion uptake mediated by hOAT1, hOAT2, hOAT3, and hOAT4. By comparing the IC 50 values of NSAIDs for hOATs, it was found that hOAT1 and hOAT3 exhibited higher affinity interactions with NSAIDs than did hOAT2 and hOAT4. HOAT1, hOAT2, hOAT3, and hOAT4 mediated the uptake of either ibuprofen, indomethacin, ketoprofen, or salicylate, but not acetylsalicylate. Although organic cation uptake mediated by hOCT1 and hOCT2 was also inhibited by some NSAIDs, hOCT1 and hOCT2 did not mediate the uptake of NSAIDs. In conclusion, hOATs and hOCTs interacted with various NSAIDs, whereas hOATs but not hOCTs mediated the transport of some of these NSAIDs. Considering the localization of hOATs, it was suggested that the interactions of hOATs with NSAIDs are associated with the pharmacokinetics and the induction of adverse reactions of NSAIDs.Nonsteroidal anti-inflammatory drugs (NSAIDs) have been widely used for their anti-inflammatory and analgesic properties. The indications of NSAIDs are broadening from rheumatic diseases and various pain states, such as cancer pain, and biliary and colic pain, to include possibly Alzheimer's disease and colon cancer prevention (Day et al., 2000). Table 1 shows the chemical structures of NSAIDs tested in the current study. Although all of these NSAIDs are weak organic acids, they are grouped in several classes based on their chemical structures. Although the chemical diversity yields a broad range of pharmacokinetic characteristics (Frust and Munster, 2000), they have some general properties in common. NSAIDs have been shown to induce various forms of adverse drug reactions including adverse gastrointestinal effects (Day et al., 2000), renal dysfunction and nephrotoxicity (Day et al., 2000), liver damage (Zimmerman, 1981;Wood et al., 1985;Purcell et al., 1991;Day et al., 2000), adverse neurological effects (Hoppman et al., 1991;Day et al., 2000), and rhabdomyolysis (Ross and Hoppel, 1987;Leventhal et al., 1989;Delrio et al., 1996).The secretion of numerous organic anions and cations, including endogenous metabolites, drugs, and xenobiotics, is an important physiological function of the renal proximal tubule. The process of secreting organic anions and cations through the proximal tubule cells is achieved via unidirectional transcellular transport involving the uptake of organic anions and cations into the cells from the blood across the basolateral membrane, followed by extrusion across the brush-border membrane into the proximal tubule fluid (Pritchard and Miller, 1993). Recently, cDNAs encoding the human organic anion transporter (hOAT) fa...

No abstract

No abstract