Interactions of urate transporter URAT1 in human kidney with uricosuric drugs.

AIM: Hyperuricaemia is a significant factor in a variety of diseases, including gout and cardiovascular diseases. The kidney plays a dominant role in maintaining plasma urate levels through the excretion process. Human renal urate transporter URAT1 is thought to be an essential molecule that mediates the reabsorption of urate on the apical side of the proximal tubule. In this study the pharmacological characteristics and clinical implications of URAT1 were elucidated. METHODS: Madin-Darby canine kidney (MDCK) cells stably expressing URAT1 (MDCK-URAT1) were established and examined the interactions of URAT1 with various drugs such as benzbromarone and its metabolites including 6-hydroxybenzbromarone, angiotensin-converting enzyme inhibitors, non-steroidal anti-inflammatory drugs and urate transport inhibitors including E3040 and probenecid. RESULTS: MDCK-URAT1 cells exhibited a time- and dose-dependent increase in urate uptake, with a Km value of 570.7 µmol/L. When an URAT1-green fluorescent protein fusion protein construct was expressed in MDCK cells, the protein was sorted mainly to the apical side of the membrane. The drugs except for captoril dose-dependently inhibited urate uptake mediated by URAT1, with half maximal inhibitory concentration (IC(50) ) values ranging 0.05-716 µmol/L. CONCLUSION: Comparing these IC(50) values with intratubular concentrations of unbound drugs in humans, it is thought that URAT1 is a target molecule of uricosuric drugs, including 6-hydroxybenzbromarone, probenecid, indomethacin and salicylate, to inhibit urate reabsorption in vivo. In addition, a cell line that stably expressing URAT1 could be a useful tool for the development of uricosuric drugs.

Ultrafiltration volume by once-weekly hemodialysis is a predictor of technique survival of combination therapy with peritoneal dialysis and hemodialysis.

Overhydration is a major cause of technique failure of peritoneal dialysis (PD). Hence, we investigated the impact of ultrafiltration (UF) volume by once-weekly hemodialysis (HD), excess volume beyond their dry weight, on technique survival of PD and HD combination therapy (PD+HD). Forty-six anuric PD+HD patients were divided into three groups according to baseline UF volume by HD: low-UF (

Interactions of human organic anion transporters with aristolochic acids.

Aristolochic acids (AAs), contained in Chinese herbal preparations, have been considered to induce nephropathy. In order to elucidate the molecular mechanisms of AA-induced nephrotoxicity, we have elucidated the interaction of human organic anion transporters (hOATs) with AAs using their stable cell lines. AA-I and AA-II inhibited organic anion uptake by hOAT1, hOAT3, and hOAT4 in dose-dependent manners. Treatment of hOAT3 with AA-I resulted in a significant reduction in viability compared with that of mock, which was rescued by the organic anion transport inhibitor probenecid. In conclusion, hOAT3-mediated AA-I uptake may be associated with the induction of nephrotoxicity.

Functional analysis of human organic cation transporter OCT3 (SLC22A3) polymorphisms.

We analyzed the functional properties of five single nucleotide polymorphisms (SNPs) in organic cation transporter OCT3 gene (SLC22A3) resulting in the amino acid changes with a transient expression system. Three SNPs (A116S, T400I, and A439V) exhibited reduced uptake of both [(3)H]histamine and [(3)H]MPP(+), although their protein expressions were detected in the plasma membrane of transfected cells. This study suggests that the OCT3 variants will contribute to inter-individual variations leading to the differences in cationic drug disposition as well as certain disease processes such as hypertension, allergic diseases, and neuropsychiatric diseases by the clearance of endogenous organic cations such as biogenic amines.

Bioimpedance Spectroscopy-Based Fluid Status in Combined Dialysis Compared With Hemodialysis and Peritoneal Dialysis: A Cross-Sectional Study.

Combination therapy with peritoneal dialysis and hemodialysis (PD+HD) is widely used in Japan for PD patients with decreased residual renal function. However, fluid status in PD+HD patients has not been well studied. In this cross-sectional study, we compared fluid status in 41 PD+HD patients with that in 103 HD and 92 PD patients using the bioimpedance spectroscopy. Extracellular water normalized to patient height (NECW, kg/m) was the highest in pre-HD (8.3 ± 1.6) followed by PD (7.9 ± 2.7), PD+HD (7.5 ± 2.5), and post-HD patients (6.9 ± 1.5) (P < 0.01). By multiple linear regression analysis, PD+HD was associated with a significantly lower NECW than pre-HD (ß = -0.8, P = 0.03) and similar to PD (ß = -0.5, P = 0.24) and post-HD (ß = 0.6, P = 0.08) after adjustment for age, sex, diabetic nephropathy, ischemic heart disease, dialysis period, and daily urine volume. There was no correlation between NECW and daily urine volume in all dialysis groups. Average daily fluid removal (a sum of urine volume and ultrafiltration volume by dialysis) was positively correlated with NECW in PD+HD and pre-HD, but not in PD and post-HD patients. Our results suggest that fluid status in PD+HD patients with decreased residual renal function is acceptable as compared with that in HD and PD patients.

Health-related quality of life on combination therapy with peritoneal dialysis and hemodialysis in comparison with hemodialysis and peritoneal dialysis: A cross-sectional study.

BACKGROUND: The health-related quality of life (HRQOL) of dialysis patients has not been well examined, especially in combination therapy with peritoneal dialysis and hemodialysis (PD+HD) patients. We compared the HRQOL of PD+HD patients with that of HD and PD patients. METHODS: A multicenter, cross-sectional study was conducted on 36 PD+HD, 103 HD, and 90 PD patients in Japan who completed the Kidney Disease Quality of Life Short Form 36, version 1.3. HRQOL scores were summarized into physical- (PCS), mental- (MCS), role/social- (RCS), and kidney disease component summaries (KDCS). RESULTS: Of the PD+HD patients, 31 (86%) transferred from PD and 5 (14%) transferred from HD. They had the longest dialysis vintage and the smallest urine volume. PCS, MCS, and KDCS HRQOL scores of PD+HD patients were comparable with those of HD and PD patients. However, the RCS score for PD+HD was significantly higher than that for HD (p = 0.020) and comparable with that for PD. PD+HD and PD were associated with significantly higher RCS scores than HD after adjusting for age, gender, diabetic nephropathy, dialysis vintage, ischemic heart disease, and peripheral arterial disease. CONCLUSIONS: For RCS, HRQOL in PD+HD patients was better than that in HD and comparable with that in PD patients, whereas the PCS, MCS, and KDCS HRQOL scores of PD+HD patients were comparable with those of HD and PD patients.

Hospitalization for Patients on Combination Therapy With Peritoneal Dialysis and Hemodialysis Compared With Hemodialysis.

INTRODUCTION: Combination therapy with peritoneal dialysis and hemodialysis (PD+HD) is widely used for PD patients with decreased residual kidney function in Japan; however, hospitalization for this combined dialysis has not been investigated so far. We compared the risk of hospitalization for PD+HD with that for HD. METHODS: A multicenter, prospective observational study was conducted on 42 PD+HD and 42 HD patients matched for age and diabetic nephropathy. The main outcome measure was the cumulative incidence of hospitalization for any cause assessed with the Kaplan-Meier method. Hospitalization rates (the number of admissions per 100 patient-years) associated with dialysis modality were also calculated. The impact of dialysis modality on time to hospitalization was analyzed using the Cox proportional hazard model. RESULTS: There was no significant difference between groups in terms of age, sex, dialysis vintage, diabetic nephropathy, and comorbidities. The cumulative incidence of hospitalization did not significantly differ between the groups (log-rank test, P = 0.36). Although total hospitalization rates were 66.0 in PD+HD and 59.2 in HD, hospitalization rates for the sum of PD-related infections (a composite of catheter-related infection and peritonitis) and vascular access troubles were 21.7 in PD+HD and 7.2 in HD. On univariate Cox proportional hazard analysis, dialysis modality had no significant impact on time to hospitalization. CONCLUSION: The risk of hospitalization was not significantly different between PD+HD and HD, although PD+HD patients had a higher risk of dialysis access-related complications than HD patients.

Role of human organic anion transporter 4 in the transport of ochratoxin A.

The purpose of this study was to investigate the characteristics of ochratoxin A (OTA) transport by multispecific human organic anion transporter 4 (hOAT4) using mouse proximal tubule cells stably transfected with hOAT4 (S(2) hOAT4). Immunohistochemical analysis revealed that hOAT4 protein was localized to the apical side of the proximal tubule. S(2) hOAT4 expressed hOAT4 protein in the apical side as well as basolateral side and the cells were cultured on the plastic dish for experiments. S(2) hOAT4 exhibited a time- and concentration-dependent, and a saturable increase in OTA uptake, with an apparent K(m) value of 22.9+/-2.44 microM. The OTA uptakes were inhibited by several substrates for the OATs. Probenecid, piroxicam, octanoate and citrinin inhibited OTA uptake by hOAT4 in a competitive manner (K(i)=44.4-336.4 microM), with the following order of potency: probenecid > piroxicam > octanoate >citrinin. The efflux of OTA by S(2) hOAT4 was higher than that by mock. Addition of OTA resulted in slight decrease in viability of S(2) hOAT4 compared with mock. These results indicate that hOAT4 mediates the high-affinity transport of OTA on the apical side of the proximal tubule, whereas the transport characteristics of OTA are distinct from those by basolateral OATs.

Interaction of human organic anion transporters with various cephalosporin antibiotics.

Cephalosporin antibiotics are thought to be excreted into the urine via organic anion transporters (OATs). The purpose of this study was to elucidate the interaction of human-OATs with various cephalosporin antibiotics, using proximal tubule cells stably expressing human-OAT1, human-OAT3 and human-OAT4. Human-OAT1 and human-OAT3 are localized to the basolateral side of the proximal tubule, whereas human-OAT4 is localized to the apical side. The cephalosporin antibiotics tested were cephalothin, cefoperazone, cefazolin, ceftriaxone, cephaloridine, cefotaxime, cefadroxil and cefamandole. All of these cephalosporin antibiotics significantly inhibited organic anion uptake mediated by human-OAT1, human-OAT3 and human-OAT4. Kinetic analysis revealed that these inhibitions were competitive. The inhibition constant (K(i)) values of cefoperazone, cefazolin, ceftriaxone and cephaloridine for human-OAT1 were much lower than those for human-OAT3 and human-OAT4, whereas the K(i) values of cephalothin and cefotaxime for human-OAT3 were much lower than those for human-OAT1 and human-OAT4. Human-OAT4 mediated the bidirectional transport of estrone sulfate, an optimal substrate for human-OAT4. These results suggest that human-OAT1, human-OAT3 and human-OAT4 interact with various cephalosporin antibiotics, and that human-OAT1 and human-OAT3 play a distinct role in the basolateral uptake of cephalosporin antibiotics. Since the K(i) value of cephaloridine for human-OAT4-mediated organic uptake was much higher than that for human-OAT1, the results indicate the possibility that human-OAT4 limits the efflux of cephaloridine, leading to the accumulation of cephaloridine and the induction of nephrotoxicity.

Interactions of human organic anion as well as cation transporters with indoxyl sulfate.

Various uremic toxicants including indoxyl sulfate exert a number of biological effects on uremic patients. In order to elucidate the molecular mechanisms for the pharmacokinetics of indoxyl sulfate in human, we examined the interactions of human organic anion transporters (human-OATs) and human organic cation transporters (human-OCTs) with indoxyl sulfate using stable transfectants. Indoxyl sulfate inhibited human-OAT1, human-OAT3 and human-OAT4, but not human-OAT2, human-OCT1 and human-OCT2. Kinetic analysis revealed that the K(i) values for human-OAT1, human-OAT3 and human-OAT4 were 22.7, 168.7 and 181.3 microM, respectively. Human-OAT1 and human-OAT3 mediated the uptake of indoxyl sulfate and human-OAT4 mediated not only the uptake but also the efflux of indoxyl sulfate. In conclusion, by comparing the K(i) values with the plasma concentration of unbound indoxyl sulfate, it was predicted that human-OAT1 and human-OAT3 mediate the transport of indoxyl sulfate in vivo. In addition, it was suggested that human-OAT1 and human-OAT3 are involved in the urinary excretion of indoxyl sulfate, the exacerbation of renal dysfunction and the induction of uremic encephalopathy by indoxyl sulfate.

Human organic cation transporter 3 mediates the transport of antiarrhythmic drugs.

Antiarrhythmic drugs have been considered to be transported by the organic cation transport system. The purpose of this study was to elucidate the molecular mechanism underlying the transport of antiarrhythmic drugs using cells from the second segment of the proximal tubule (S2) cells of mice expressing human-organic cation transporter 3 (S2 human-OCT3). The antiarrhythmic drugs tested were cibenzoline, disopyramide, lidocaine, mexiletine, phenytoin, pilsicanide, procainamide and quinidine. Human-OCT3 mediated a time- and dose-dependent uptake of quinidine and lidocaine, with Km values of 216 and 139 microM, respectively. Human-OCT3 also mediated the uptake of disopyramide and procainamide but not that of phenytoin. All antiarrhythmic drugs tested inhibited histamine uptake mediated by human-OCT3 in a dose-dependent manner. The IC50 values of antiarrhythmic drugs for human-OCT3 ranged between 0.75 and 656 microM. Kinetic analysis revealed that disopyramide, lidocaine, procainamide and quinidine inhibited histamine uptake mediated by human-OCT3 in a competitive manner. In conclusion, these results suggest that human-OCT3 mediates the transport of antiarrhythmic drugs, which may be the mechanism underlying the distribution and the elimination of these drugs.

Evidence for a role of human organic anion transporters in the muscular side effects of HMG-CoA reductase inhibitors.

The purpose of this study was to elucidate the role of human organic anion transporters (human OATs) in the induction of drug-induced skeletal muscle abnormalities. 3-Hydroxy-3-methylglutaryl-coenzyme A (HMG-CoA) reductase inhibitors have been clinically used for lowering plasma cholesterol levels, and are known to induce various forms of skeletal muscle abnormalities including myopathy and rhabdomyolysis. Immunohistochemical analysis revealed that human OAT1 and human OAT3 are localized in the cytoplasmic membrane of the human skeletal muscles. The activities of human OATs were measured using mouse cell lines from renal proximal tubules stably expressing human OATs. Human OAT3, but not human OAT1, mediates the transport of pravastatin. Fluvastatin inhibited organic anion uptake mediated by human OAT1 in a mixture of competitive and noncompetitive manner, whereas simvastatin and fluvastatin noncompetitively inhibited the organic anion uptake mediated by human OAT3. In conclusion, the organic anion transporters OAT1 and OAT3 are localized in the cytoplasmic membrane of human skeletal muscles. Pravastatin, simvasatin, and fluvasatin inhibit human OATs activity. These results suggest that muscle organic anion transporters play a role in the muscular side effects of HMG-CoA reductase inhibitors.

Interaction of human and rat organic anion transporter 2 with various cephalosporin antibiotics.

Cephalosporin antibiotics are thought to be excreted into the urine via organic anion transporters (OATs) and OAT can mediate nephrotoxicity by cephalosporins, particularly by cephaloridine. The purpose of this study was to elucidate the interaction of human-OAT2 and rat-OAT2 with cephalosporin antibiotics using proximal tubule cells stably expressing human-OAT2 and rat-OAT2. Human-OAT2 is localized to the basolateral side of the proximal tubule, whereas rat-OAT2 is localized to the apical side of the proximal tubule. Cephalosporins tested were cephalothin, cefoperazone, cefazolin, ceftriaxone, cephaloridine, cefotaxime, cefadroxil and cefamandole. These cephalosporins dose-dependently inhibited organic anion uptake mediated by human-OAT2 and rat-OAT2. There was no species difference observed for the effects of OAT2 with cephalosporins between human and rat transporters. Kinetic analysis revealed that the inhibitory effects for human-OAT2 were competitive. Cephaloridine significantly decreased the viability of cells stably expressing human-OAT2, human-OAT1, human-OAT3 and human-OAT4. The decreased viability of cells stably expressing human-OAT1, human-OAT3 and human-OAT4 but not human-OAT2 was reversed by probenecid. In conclusion, human-OAT2 interacts with cephalosporins, and thus, human-OAT2 may mediate the uptake of cephalosporins on the basolateral side of the proximal tubule. The interaction of human-OAT2 with cephalosporins was the weakest among the basolateral human-OATs tested. In addition, it is suggested that human-OATs mediate cephaloridine-induced nephrotoxicity.

Involvement of rat organic anion transporter 3 (rOAT3) in cephaloridine-induced nephrotoxicity: in comparison with rOAT1.

This study was performed to elucidate the possible involvement of organic anion transporter 3 (OAT3) in cephaloridine (CER)-induced nephrotoxicity and compare the substrate specificity between rOAT3 and rat OAT1 (rOAT1) for various cephalosporin antibiotics, using proximal tubule cells stably expressing rOAT3 (S2 rOAT3) and rOAT1 (S2 rOAT1). S2 rOAT3 exhibited a CER uptake and a higher susceptibility to CER cytotoxicity than did mock, which was recovered by probenecid. Various cephalosporin antibiotics significantly inhibited both estrone sulfate uptake in S2 rOAT3 and para-aminohippuric acid uptake in S2 rOAT1. The Ki values of CER, cefoperazone, cephalothin and cefazolin for rOAT3- and rOAT1-mediated organic anion transport ranged from 0.048 to 1.14 mM and from 0.48 to 1.32 mM, respectively. These results suggest that rOAT3, at least in part, mediates CER uptake and CER-induced nephrotoxicity as rOAT1. There was some difference of affinity between rOAT3 and rOAT1 for cephalosporin antibiotics.

Interactions of organic anion transporters and organic cation transporters with mycotoxins.

Mycotoxins are secondary metabolites of moulds that which exert adverse effects in humans and animals. It is known that direct cellular toxicity is often associated with increased cellular accumulation of toxic compounds, and membrane transport may be the first fundamental stage in the development of the cytotoxicity. To elucidate the entry pathway for mycotoxins into cells, we have investigated the interactions of human and rat organic anion transporters (hOATs/rOats) and human organic cation transporters (hOCTs) with mycotoxins using cells stably expressing hOATs/rOats/hOCTs. The mycotoxins tested were aflatoxin B1, alpha-zearalenol, citrinin, citrioveridine, cyclopiazonic acid, fumonisin B1, gliotoxin, patulin, penicillic acid, rubratoxin B, and zearalenone. These mycotoxins inhibited organic anion uptake mediated by hOAT1-4, and organic cation uptake mediated by hOCT1-2. By comparing the IC(50) values of mycotoxins for hOATs, it was found that hOAT1 and hOAT3 exhibited higher affinity interactions with mycotoxins than hOAT2 and hOAT4. There was no interspecies difference between humans and rats for the interactions of OATs with mycotoxins except that of OAT3 with rubratoxin B. Finally, we observed that hOAT1-4 and hOCT1-2 mediated the uptake of aflatoxin B1. In conclusion, hOATs and hOCTs interacted with various mycotoxins. Considering the localization of hOATs/rOats and hOCTs, it was suggested that these transporters were the possible entrance pathway for mycotoxins in kidney and liver, leading to the induction of adverse effects in humans and rats.

Functional characterization of rat organic anion transporter 5 (Slc22a19) at the apical membrane of renal proximal tubules.

A novel member of the organic anion transporter (OAT) family, Oat5 (Slc22a19), has been reported to transport a naturally occurring mycotoxin, ochratoxin A (OTA). However, neither its endogenous substrate and driving force nor physiological functions have been determined. Herein, we report the functional characterization of rat Oat5 (rOat5), as well as its intrarenal distribution and membrane localization. When expressed in Xenopus laevis oocytes, rOat5 mediated the transport of sulfate conjugates of steroids such as estrone-3-sulfate (E(1)S; K(m) = 18.9 +/- 3.9 microM) and dehydroepiandrosterone sulfate (K(m) = 2.3 +/- 0.2 microM) in a sodium-independent manner, in addition to OTA. The rOat5-mediated E(1)S transport was strongly inhibited by four-carbon (C4) dicarboxylate succinate and longer dicarboxylates (C7-C9). The uptake of [(3)H]E(1)S via rOat5 was significantly trans-stimulated by succinate, and the efflux of [(14)C]succinate was significantly trans-stimulated by E(1)S. A similar trans-stimulatory effect of preloaded succinate on E(1)S uptake was also detected in cells stably expressing rOat5 (S(2) rOat5). rOat5 interacted with chemically heterogenous anionic compounds. The rOat5-mediated E(1)S transport was inhibited by several sulfate conjugates, such as 4-methylumbelliferyl sulfate and beta-estradiol sulfate, but not by glucuronide conjugates. An immunohistochemical study showed that rOat5 was localized at the apical membrane of renal proximal tubules in the corticomedullary region. rOat5 mRNA was expressed in the late segments (S(2) and S(3)) of proximal tubules. These results indicate that rOat5 is renal organic anion/dicarboxylates exchanger and, under physiological conditions, may function as an apical reabsorptive pathway for organic anions in proximal tubules driven by an outward gradient of dicarboxylates.

Human organic anion transporters and human organic cation transporters mediate renal transport of prostaglandins.

Prostaglandin E(2) (PGE(2)) and prostaglandin F(2 alpha) (PGF(2 alpha)) have been used for the induction of labor and the termination of pregnancy. Renal excretion is shown to be an important pathway for the elimination of PGE(2) and PGF(2 alpha). The purpose of this study was to elucidate the molecular mechanism of renal PGE(2) and PGF(2 alpha) transport using cells stably expressing human organic anion transporter (hOAT) 1, hOAT2, hOAT3, and hOAT4, and human organic cation transporter (hOCT) 1 and hOCT2. A time- and dose-dependent increase in PGE(2) and PGF(2 alpha) uptake was observed in cells expressing hOAT1, hOAT2, hOAT3, hOAT4, hOCT1, and hOCT2. The K(m) values of PGE(2) uptake by hOAT1, hOAT2, hOAT3, hOAT4, hOCT1, and hOCT2 were 970, 713, 345, 154, 657, and 28.9 nM, respectively, whereas those of PGF(2 alpha) uptake by hOAT1, hOAT3, hOAT4, hOCT1, and hOCT2 were 575, 1092, 692, 477, and 334 nM, respectively. PGE(2) and PGF(2 alpha) significantly inhibited organic anion uptake by hOATs and organic cation uptake by hOCTs. In conclusion, considering the localization of these transporters, the results suggest that PGE(2) and PGF(2 alpha) transport in the basolateral membrane of the proximal tubule is mediated by multiple pathways including hOAT1, hOAT2, hOAT3, and hOCT2, whereas that in the apical side is mediated by hOAT4.

Urate transport via human PAH transporter hOAT1 and its gene structure.

BACKGROUND: We recently cloned the human organic anion transporter 1 (hOAT1) as a p-aminohippurate (PAH) transporter. Whether urate is transported by the PAH transporter in humans remains unclear. Familial juvenile gouty nephropathy (FJGN) is thought to develop as a result of an abnormality in the urate transporter. METHODS: To determine if hOAT1 transported urate, the cellular uptakes of PAH and urate were determined, as were the inhibition profiles of inorganic anions, and uricosuric and antiuricosuric agents using a mouse S2 cell line expressing hOAT1. The hOAT1 gene was cloned from a genomic library using full-length hOAT1-1 cDNA as a probe. The coding regions of the hOAT1 genes of two sisters with FJGN were sequenced. Also, immunohistochemical fluorescence analysis of hOAT1 in the kidney of the younger sister with FJGN was performed. RESULTS: The Km and Vmax values of urate transport via hOAT1 were 943 +/- 84 micromol/L and 1286 +/- 162 pmol/mg protein/min, respectively. The order of the IC50 of urate transport via hOAT1 was benzbromarone < probenecid < salicylate or pyrazine carboxylic acid. The 10.9 kb hOAT1 gene was found to be interrupted by nine introns. Mutations in the coding region of the hOAT1 gene from the two sisters with FJGN were undetectable. Immunohistochemical fluorescent staining showed that hOAT1 in the kidney of the younger sister was similar to that of control individuals. CONCLUSIONS: Our data show that hOAT1 transports urate, and the inhibition profiles of uricosuric and antiuricosuric agents are defined. hOAT1 is not responsible for FJGN in the two sisters examined in this study.

Human organic anion transporter 4 is a renal apical organic anion/dicarboxylate exchanger in the proximal tubules.

Human organic anion transporter OAT4 is expressed in the kidney and placenta and mediates high-affinity transport of estrone-3-sulfate (E1S). Because a previous study demonstrated no trans-stimulatory effects by E1S, 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 (S2 OAT4). OAT4-mediated E1S uptake was inhibited by glutarate (GA) (IC50:1.25 mM) and [14C]GA uptake via S2 OAT4 was significantly trans-stimulated by unlabeled GA (5 mM) (P<0.001). [3H]E1S uptake via S2 OAT4 was significantly trans-stimulated by preloaded GA (P<0.001) and its [14C]GA efflux was significantly trans-stimulated by unlabeled E1S in the medium (P<0.05). In addition, both the uptake and efflux of [14C]p-aminohippuric acid (PAH) and [14C]GA via S2 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.

Interactions of human organic anion transporters and human organic cation transporters with nonsteroidal anti-inflammatory drugs.

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.