Uric Acid Transport Research Articles

Advance in treatment of hyperuricemia by Chinese medicine based on uric acid transporterome

With the development of the quality of life, the morbidity of hyperuricemia is increasing year by year. At the same time, it appears that this disease attacks the young people currently. As the study of pathogenesis of hyperuricemia advanced, a series of uric acid transporters were found during this process. Meanwhile, the definition of transporterome was proposed. They were divided into three groups according to the functions: reabsorption proteins, excretion proteins and skeleton proteins. At moment, the drugs for hyperuricmia mainly include uric acid composition inhibitors and uric acid excretion promoters. Since the excretion of uric acid plays a leading role during the process of attack of hyperurecimia, it makes sense to explore Chinese medicines with clear mechanism targeting the transporterome. Therefore, this paper would focus on transporterome and summarize the mechanisms of Chinese medicines in treating hyperuricemia.

Hereditary Renal Hypouricemia Type 1 and Autosomal Dominant Polycystic Kidney Disease

BackgroundRenal hypouricemia (RHUC) is a heterogeneous inherited disorder characterized by impaired tubular uric acid (UA) transport with severe complications, such as acute kidney injury. Type 1 is caused by a mutation in the SLC22A12 gene (URAT1) and type 2 in the SLC2A9 gene (GLUT9). In this article, the authors present a coexpression functional characterization of variants responsible for RHUC type 1 in a Czech family with polycystic kidney disease (PKD). MethodsThe serum UA concentration in the proband was 1.1 mg/dL and was expressed as an increase in the fractional excretion of UA (43%). The URAT1 allelic variants were prepared and their urate uptake and subcellular localization were studied by Xenopus oocytes expression system. Linkage analysis using a panel of 4 CA-repeat flanking markers for chromosome 16 and a panel of 3 CA-repeat flanking markers for chromosome 4 was performed, which carry the PKD1 and PKD2 genes, respectively. ResultsCoexpression results showed that p.G366R and p.R477H suppressed urate transport by wild types. Colocalization studies showed an accumulation of URAT1 in the endoplasmic reticulum of the p.G366R variant and mainly retention of wild-type protein by variants p.G366R and p.R477H. ConclusionsThe findings suggest that not only loss-of-function mutation of URAT1 but also the dominant-negative effect cause RHUC through loss of UA absorption, partly due to protein misfolding caused by accumulation of URAT1 protein in the endoplasmic reticulum.

Expression and function of placental uric acid transporter GLUT9

Expression and function of placental uric acid transporter GLUT9

Acute Effects of Oral Tofisopam on Plasma Concentration and Urinary Excretion of Uric Acid and Oxypurinol "Preliminary Communication".

The effects of tofisopam, a GABA-receptor agonist, following oral administration (300mg) with and without allopurinol pretreatment on the plasma concentration and renal transport of uric acid and oxypurinol were investigated in 5 healthy subjects. Fractional and urinary excretions of uric acid were both significantly increased at 2-3 hours after tofisopam administration (559% and 459%, respectively), while plasma uric acid concentration was significantly decreased (36%) at 2.5 hours, suggesting that tofisopam affects uric acid metabolism via the tubular transport system. The hypouricemic effect of tofisopam was comparable to or greater than that of losartan and/or fenofibrate, which also have uric acid-lowering activity. In addition, with prior administration of allopurinol, the fractional and urinary excretions of oxypurinol were increased at 2-3 hours after tofisopam administration (51% and 33%, respectively), while the plasma oxypurinol concentration was significantly decreased at 1.5 and 2.5 hours (15% and 21%, respectively). Accordingly, tofisopam may be an attractive compound for treatment of hyperuricemia and/or gout, especially in patients complicated with autonomic dysfunction symptoms, though it is possible that the uric acid-lowering effect of oxypurinol is attenuated by tofisopam.

The reduced expression of proximal tubular transporters in acquired Fanconi syndrome with κ light chain deposition

In a case of acquired Fanconi syndrome associated with smoldering myeloma, we confirmed the deposition of protease-resistant κ light chain proteins in a proximal tubular injury and found the decreased expression of apical tubular transporters including sodium glucose co-transporter, sodium phosphate co-transporter, uric acid transporter 1, and a decrease of Na(+)/K(+)-ATPase in the basolateral membrane. The protease-resistant kappa light chain has a pathological role in the expression of tubular transporters in the proximal tubule and causes Fanconi syndrome associated with smoldering myeloma.

High frequency of SLC22A12 variants causing renal hypouricemia 1 in the Czech and Slovak Roma population; simple and rapid detection method by allele-specific polymerase chain reaction

Renal hypouricemia is a rare heterogeneous inherited disorder characterized by impaired tubular uric acid transport with severe complications, such as acute kidney injury. Type 1 and 2 are caused by loss-of-function mutations in the SLC22A12 and SLC2A9 gene, respectively. A cohort of 881 randomly chosen ethnic Roma from two regions in Eastern Slovakia and two regions in the Czech Republic participated. Genomic DNA was isolated from buccal swabs and/or from blood samples. The c.1245_1253del and c.1400C>T genotypes were determined using polymerase chain reaction with allele-specific primers in a multiplex arrangement and/or direct sequencing of exon 7 and 9. Allele frequencies and genotypes were tested for Hardy-Weinberg equilibrium using the Chi-square test. 25 subjects were heterozygous and three were homozygous for the c.1245_1253del, while 92 subjects were heterozygous and two were homozygous for the c.1400C>T. Moreover, two participants were compound heterozygotes. Frequencies of the c.1245_1253del and c.1400C>T variants were 1.87 and 5.56 %, respectively. Our finding confirms an uneven geographical and ethnic distribution of SLC22A12 mutant variants. We found that the c.1245_1253del and c.1400C>T variants were present in the Czech and Slovak Roma population at unexpectedly high frequencies. Renal hypouricemia should be kept in mind during differential diagnostic on Roma patients with low serum uric acid concentrations.

A network pharmacology approach to determine active ingredients and rationality of herb combinations of Modified-Simiaowan for treatment of gout

Ethnopharmacological relevanceModified Simiaowan (MSW) is a traditional Chinese medicine (TCM) formula and is widely used as a clinically medication formula for its efficiency in treating gouty diseases.To predict the active ingredients in MSW and uncover the rationality of herb combinations of MSW. Materials and methodsThree drug-target networks including the “candidate ingredient-target network” (cI-cT) that links the candidate ingredients and targets, the “core ingredient-target-pathway network” connecting core potential ingredients and targets through related pathways, and the “rationality of herb combinations of MSW network”, which was derived from the cI-cT network, were developed to dissect the active ingredients in MSW and relationship between ingredients in herb combinations and their targets for gouty diseases. On the other hand, herbal ingredients comparisons were also conducted based on six physicochemical properties to investigate whether the herbs in MSW are similar in chemicals. Moreover, HUVEC viability and expression levels of ICAM-1 induced by monosodium urate (MSU) crystals were assessed to determine the activities of potential ingredients in MSW. ResultsPredicted by the core ingredient-target-pathway network, we collected 30 core ingredients in MSW and 25 inflammatory cytokines and uric acid synthetase or transporters, which are effective for gouty treatment through some related pathways. Experimental results also confirmed that those core ingredients could significantly increase HUVEC viability and attenuate the expression of ICAM-1, which supported the effectiveness of MSW in treating gouty diseases. Moreover, heat-clearing and dampness-eliminating herbs in MSW have similar physicochemical properties, which stimulate all the inflammatory and uric acid-lowing targets respectively, while the core drug and basic prescription in MSW stimulate the major and almost all the core targets, respectively. ConclusionOur work successfully predicts the active ingredients in MSW and explains the cooperation between these ingredients and corresponding targets through related pathways for gouty diseases, and provides basis for an alternative approach to investigate the rationality of herb combinations of MSW on the network pharmacology level, which might be beneficial to drug development and applications.

Functional characterization of NAT/NCS2 proteins of Aspergillus brasiliensis reveals a genuine xanthine–uric acid transporter and an intrinsically misfolded polypeptide

The Nucleobase-Ascorbate Transporter (NAT) family includes members in nearly all domains of life. Functionally characterized NAT transporters from bacteria, fungi, plants and mammals are ion-coupled symporters specific for the uptake of purines, pyrimidines and related analogues. The characterized mammalian NATs are specific for the uptake of L-ascorbic acid. In this work we identify in silico a group of fungal putative transporters, named UapD-like proteins, which represent a novel NAT subfamily. To understand the function and specificity of UapD proteins, we cloned and functionally characterized the two Aspergillus brasiliensis NAT members (named AbUapC and AbUapD) by heterologous expression in Aspergillus nidulans. AbUapC represents canonical NATs (UapC or UapA), while AbUapD represents the new subfamily. AbUapC is a high-affinity, high-capacity, H+/xanthine–uric acid transporter, which can also recognize other purines with very low affinity. No apparent transport function could be detected for AbUapD. GFP-tagging showed that, unlike AbUapC which is localized in the plasma membrane, AbUapD is ER-retained and degraded in the vacuoles, a characteristic of misfolded proteins. Chimeric UapA/AbUapD molecules are also turned-over in the vacuole, suggesting that UapD includes intrinsic peptidic sequences leading to misfolding. The possible evolutionary implication of such conserved, but inactive proteins is discussed.

Direct radical scavenging activity of benzbromarone provides beneficial antioxidant properties for hyperuricemia treatment.

Uric acid exerts an important antioxidant effect against external oxidative stress under physiological conditions. However, uric acid itself can increase oxidative stress via reduced nicotinamide adenine dinucleotide phosphate (NADPH) oxidase activation in adipocytes and vascular cells. Uric acid transporter 1 is involved in the generation of this oxidative stress. Furthermore, uric acid locally activates the renin-angiotensin system, thus producing angiotensin II and subsequently increasing intracellular oxidative stress. Benzbromarone has been reported to suppress uric acid reabsorption via uric acid transporter 1 inhibition in renal tubular cells. In this study we evaluated the in vitro antioxidant effect of benzbromarone from several perspectives. First, the direct radical-trapping capacity of benzbromarone was measured by chemiluminescence assay and electron paramagnetic resonance spectroscopy. Second, the intracellular antioxidant activity of benzbromarone in hyperuricemia was evaluated using endothelial cells. In light of these results, benzbromarone is hypothesized directly to scavenge the superoxide anion radical. In addition, benzbromarone inhibited reactive oxygen species production that was induced by angiotensin II or uric acid in endothelial cells. These findings suggest that benzbromarone possesses the ability directly to scavenge radicals and may act as an antioxidant against uric acid and angiotensin II-induced oxidative stresses in endothelial cells at therapeutically achievable levels in blood.

Molecular Mechanisms Underlying Urate-Induced Enhancement of Kv1.5 Channel Expression in HL-1 Atrial Myocytes.

Hyperuricemia induces endothelial dysfunction, oxidative stress and inflammation, increasing cardiovascular morbidities. It also raises the incidence of atrial fibrillation; however, underlying mechanisms are unknown. The effects of urate on expression of Kv1.5 in cultured mouse atrial myocytes (HL-1 cells) using reverse transcriptase-PCR, immunoblots, flow cytometry and patch-clamp experiments were studied. Treatment with urate at 7 mg/dl for 24 h increased the Kv1.5 protein level, enhanced ultra-rapid delayed-rectifier K(+)channel currents and shortened action potential duration in HL-1 cells. HL-1 cells expressed the influx uric acid transporter (UAT), URATv1, and the efflux UATs, ABCG2 and MRP4. An inhibitor against URATv1, benzbromarone, abolished the urate effects, whereas an inhibitor against ABCG2, KO143, augmented them. Flow cytometry showed that urate induced an increase in reactive oxygen species, which was abolished by the antioxidant, N-acetylcysteine (NAC), and the NADPH-oxidase inhibitor, apocynin. Both NAC and apocynin abolished the enhancing effects of urate on Kv1.5 expression. A urate-induced increase in the Kv1.5 proteins was accompanied by phosphorylation of extracellular signal-regulated kinase (ERK), and was abolished by an ERK inhibitor, PD98059. NAC abolished phosphorylation of ERK by urate. Intracellular urate taken up by UATs enhanced Kv1.5 protein expression and function in HL-1 atrial myocytes, which could be attributable to ERK phosphorylation and oxidative stress derived from nicotinamide adenine dinucleotide phosphate (NADPH)-oxidase.

Depletion of Uric Acid Due to SLC22A12 (URAT1) Loss-of-Function Mutation Causes Endothelial Dysfunction in Hypouricemia.

Uric acid (UA) serves as an antioxidant in vascular endothelial cells. UA transporter 1 (URAT1) encoded by SLC22A12 is expressed in the kidney and vessels and its loss of function causes hypouricemia. The purpose of this study was to examine whether there is any endothelial dysfunction in patients with hypouricemia. Twenty-six patients with hypouricemia (<2.5 mg/dl) and 13 healthy control subjects were enrolled. Endothelial function was evaluated using flow-mediated dilation (FMD). mRNA of UA transporters expressed in cultured human umbilical endothelial cells (HUVEC) was detected on RT-PCR. There was a positive correlation between FMD and serum UA in the hypouricemia group. URAT1 loss-of-function mutations were found in the genome of 21 of 26 patients with hypouricemia, and not in the other 5. In the hypouricemia groups, serum UA in homozygous and compound heterozygous patients was significantly lower than in other groups, suggesting that severity of URAT1 dysfunction may influence the severity of hypouricemia. Thirteen of 16 hypouricemia subjects with homozygous and compound heterozygote mutations had SUA <0.8 mg/dl and their FMD was lower than in other groups. HUVEC do not express mRNA of URAT1, suggesting the null role of URAT1 in endothelial function. Depletion of UA due to SLC22A12/URAT1 loss-of-function mutations causes endothelial dysfunction in hypouricemia patients.

Effects of Xie-Zhuo-Chu-Bi-Fang on miR-34a and URAT1 and their relationship in hyperuricemic mice

Ethnopharmacological relevanceXie-Zhuo-Chu-Bi-Fang (XZCBF) is an empirical formula that was developed based on the principles of traditional Chinese medicine, for the therapeutic purpose of treating hyperuricemia. XZCBF has been clinically utilized in the Department of Traditional Chinese Medicine at General Hospital of Guangzhou Military Command of PLA for many years and has exhibited favorable efficacy. The aim of the study is to evaluate the effects of XZCBF on the expression of uric acid transporter 1 (URAT1) and miR-34a in hyperuricemic mice and to determine, the correlation between the two expression levels. Materials and methodsA hyperuricemic animal model was created by administering adenine and allantoxanic acid potassium salt to mice. The blood uric acid levels were measured in these model mice after treatment with XZCBF for 15 days. The potential targets of miR-34a were screened. The expression levels of miR-34a and URAT1 in the renal tissues collected from the model mice were determined by quantitative real-time polymerase chain reaction (qRT-PCR) analysis, and their correlation was further established by immunohistochemistry and in situ hybridization. ResultsThe uric acid levels in the model mice were significantly higher than those in the blank controls (P<0.05). These levels were significantly lower in the three groups receiving different doses of XZCBF (P<0.05), which was, in agreement with the downregulation of URAT1 and the upregulation of miR-34a in each group. The mRNA expression level of URAT1 was positively correlated with the concentration of uric acid but, negatively correlated with the expression level of miR-34a. ConclusionsThe ability of XZCBF to facilitate the excretion of uric acid and to lower its level in the model group was mediated by the upregulation of miR-34a and the inhibition of URAT1 mRNA expression, which suggests that XZCBF could be an option for the treatment of hyperuricemia in mice.

291: Gestational weight gain in twin pregnancies is not associated with discordance in birthweight or small for gestational age

291 Gestational weight gain in twin pregnancies is not associated with discordance in birthweight or small for gestational age Bahram Salmanian, Kathleen Antony, Diana Racusin, Kjersti Aagaard, Amirhoushang Shamshirsaz, Winston Campbell, Anthony Odibo, Alireza Shamshirsaz Baylor College of Medicine, Houston, TX, Washington University in St. Louis, St. Louis, MO, University of Connecticut, Storrs, CT, George Washington University, Washington, DC OBJECTIVE: Twins account for 3% of live births in the United States, and have a disproportionate burden of small-for-gestational age (SGA) and prematurity. Gestational weight gain guidelines (GWG) for women with twins are to gain significantly more weight than gravidae with singletons. The benefit of this increased GWG on birthweight has not been clearly demonstrated. We sought to apply published and original GWG guidelines for twins to assess association with growth discordance and SGA. STUDY DESIGN: This is a secondary analysis of a cohort of 600 gravidae carrying twins (1991-2011). GWG was categorized as insufficient, at goal, or excess using three different GWG guidelines: IOM guideline corrected for gestational age (GA), a validated gestational-week specific guideline (Guideline A), and a gestationalage specific guideline that we developed consisting of the IOM singleton GWG recommendation plus the contribution of an additional fetal-placental unit, including the estimated amniotic fluid weight, placental weight, and the “ideal” fetal weight (Guideline BK). The ideal fetal weight in this formula was calculated from coefficients for significant physiological variables affecting fetal growth, which were derived using backward stepwise multiple regression adjusting for chorionicity. The association between SGA <10th percentile defined using the twin chart versus traditional definitions of SGA were compared. Statistical analyses including adjusted OR, 95% confidence interval (CI), were performed. RESULTS: Using any of the three GWG models, GWG was not associated with growth discordance or SGA. CONCLUSION: GWG that is below the recommended IOM guidelines, below validated published guidelines, or our derived formula is not associated with growth discordance or SGA. The benefits of advising gravidae to gain significantly more weight than singletons do not appear to bear out in terms of twin growth. 292 Placental urate transport is GLUT9-mediated and unidirectional Benjamin Luscher, Camilla Marini, Xiao Huang, Christiane Albrecht, Matthias Hediger, Bruno Stieger, Daniel Surbek, Marc Baumann University Hospital of Bern, University of Bern, Departments of Obstetrics and Gynecology, Department of Clinical Research, Bern, Switzerland, University of Bern, Institute for Biochemestry and Molecular Medicine, Graduate School for Cellular and Biomedical Sciences (GCB), Bern, Switzerland, University of Bern, Institute for Biochemestry and Molecular Medicine, Bern, Switzerland, University Hospital of Zurich, Department of Clinical Pharmacology and Toxicology, Zurich, Switzerland OBJECTIVE: Pre-eclampsia is associated with increased maternal urate serum levels. Hyperuricemia might lead to long-term maternal cardiovascular risk and to fetal programming. The regulation of the placental urate transport system is not yet understood, but is likely to be predominantly regulated by GLUT-9 (glucose transporter 9), besides the multitransporter hOAT4 (human organic anion transporter). The aim of this study was to investigate the placental urate transport system and to characterize its transporter GLUT9. STUDY DESIGN: A transepithelial transport (Transwell ) model to assess urate transport activity was used. Electrophysiological techniques and radioactive ligand up-take assay were used to measure transport activity of GLUT9 overexpressed in Xenopus oocytes, and its activity under various conditions. We further analysed the placental GLUT9 expression using novel self-raised antibodies against human isoforms GLUT9a and -b. RESULTS: In the transwell system, urate is transported across the BeWo choriocarcinoma cell monolayer with 530 pmol/min. In the Xenopus oocytes system chloride replacing chloride with iodine resulted in a complete loss of urate transport. In radioactive up-take experiments iodine had no effect on urate transport. Both GLUT9 isoforms are present in the microvillous membrane (MVM) of the syncytiotrophoblast cells, but not in the basal membrane (BM). CONCLUSION: Our results for the first time show that urate transport in the placenta is unidirectional in the feto-maternal direction, because GLUT9 isoforms are expressed exclusively in the microvillous (apical) membrane of the trophoblast and because hOAT4 is known be localized on the basal membrane and to transport urate unidirectionally into trophoblast cells. Furthermore, GLUT9 uric acid transport activity is iodine-regulated by changing the mode of up-take from an electrogenic to an electroneutral transport. This may allow regulating urate transport by the change of an active to a passive transport. This study was funded by the Swiss National Fund NCCR grant “TransCure”

Uric acid promotes apoptosis in human proximal tubule cells by oxidative stress and the activation of NADPH oxidase NOX 4.

Mild hyperuricemia has been linked to the development and progression of tubulointerstitial renal damage. However the mechanisms by which uric acid may cause these effects are poorly explored. We investigated the effect of uric acid on apoptosis and the underlying mechanisms in a human proximal tubule cell line (HK-2). Increased uric acid concentration decreased tubule cell viability and increased apoptotic cells in a dose dependent manner (up to a 7-fold increase, p<0.0001). Uric acid up-regulated Bax (+60% with respect to Ctrl; p<0.05) and down regulated X-linked inhibitor of apoptosis protein. Apoptosis was blunted by Caspase-9 but not Caspase-8 inhibition. Uric acid induced changes in the mitochondrial membrane, elevations in reactive oxygen species and a pronounced up-regulation of NOX 4 mRNA and protein (p<0.05). In addition, both reactive oxygen species production and apoptosis was prevented by the NADPH oxidase inhibitor DPI as well as by Nox 4 knockdown. URAT 1 transport inhibition by probenecid and losartan and its knock down by specific siRNA, blunted apoptosis, suggesting a URAT 1 dependent cell death. In summary, our data show that uric acid increases the permissiveness of proximal tubule kidney cells to apoptosis by triggering a pathway involving NADPH oxidase signalling and URAT 1 transport. These results might explain the chronic tubulointerstitial damage observed in hyperuricaemic states and suggest that uric acid transport in tubular cells is necessary for urate-induced effects.

Regulatory effect of leonurus extracts on hyperuricemia in rats

In this study, SD rats were orally administrated with oteracil potassium (300 mg . kg-1 . d-1 ) to prepare the hyperuricemia model, and divided into normal, model, Allopurinol, LE high dosage, middle dosage and low dose (200, 100, 50 mg . kg-1 . d-1) groups. The rats were orally administrated with test drugs 1 hour later after being orally administrated with Oteracil potassium. After 7 days, serum uric acid, serum creatinine, uric acid and expression of relevant transporters in kidney were tested to study the regulatory effect of leonurus extracts on serum uric acid, renal function and relevant transporters in kidney of rats with hyperuricemia. Compared with the model group, the leonurus extract group could significantly down-regulate serum uric acid and creatinine levels of rats with hyperuricemia, and increase the urine uric acid level. Meanwhile, leonurus extracts could notably down-regulate the mRNA expressions of urate transporter 1 (URAT1) and glucose transporter 9 (GLUT9), up-regulate the mRNA expressions of organic cation transportanter (OCT) and Carnitine transporter (OCTN) and promote the excretion of uric acid of kidney.

Uric acid relates to dopamine transporter availability in Parkinson's disease.

Diagnosing Parkinson's disease (PD) and tracking its progression may require the combination of reliable biomarkers. Among them, both serum uric acid (UA) and dopamine transporter (DaT) binding deserve more investigations. We aimed to investigate the relationship between serum UA levels and DaT availability in newly diagnosed, drug-naïve PD patients, by means of semiquantitative [(123) I]FP-CIT-SPECT. We recruited 52 newly diagnosed, drug-naïve PD patients, and performed serum UA dosage and [(123) I]FP-CIT-SPECT. Pearson's correlation analysis showed that UA levels were significantly higher in patients with higher averaged, ipsilateral and contralateral DaT binding in caudate, putamen, and striatum. We showed, for the first time, by regional semiquantitative analysis of DaT binding in PD patients that UA levels significantly correlates with the severity of dopaminergic impairment in caudate, putamen, and striatum. This study broadens our knowledge on the importance of UA as a biomarker of PD.

Expression, purification, and structural insights for the human uric acid transporter, GLUT9, using the Xenopus laevis oocytes system.

The urate transporter, GLUT9, is responsible for the basolateral transport of urate in the proximal tubule of human kidneys and in the placenta, playing a central role in uric acid homeostasis. GLUT9 shares the least homology with other members of the glucose transporter family, especially with the glucose transporting members GLUT1-4 and is the only member of the GLUT family to transport urate. The recently published high-resolution structure of XylE, a bacterial D-xylose transporting homologue, yields new insights into the structural foundation of this GLUT family of proteins. While this represents a huge milestone, it is unclear if human GLUT9 can benefit from this advancement through subsequent structural based targeting and mutagenesis. Little progress has been made toward understanding the mechanism of GLUT9 since its discovery in 2000. Before work can begin on resolving the mechanisms of urate transport we must determine methods to express, purify and analyze hGLUT9 using a model system adept in expressing human membrane proteins. Here, we describe the surface expression, purification and isolation of monomeric protein, and functional analysis of recombinant hGLUT9 using the Xenopus laevis oocyte system. In addition, we generated a new homology-based high-resolution model of hGLUT9 from the XylE crystal structure and utilized our purified protein to generate a low-resolution single particle reconstruction. Interestingly, we demonstrate that the functional protein extracted from the Xenopus system fits well with the homology-based model allowing us to generate the predicted urate-binding pocket and pave a path for subsequent mutagenesis and structure-function studies.

Mendelian randomization provides no evidence for a causal role of serum urate in increasing serum triglyceride levels.

Triglycerides and their lipoprotein transport molecules are risk factors for heart disease. Observational studies have associated elevated levels of serum urate (SU) with triglycerides and risk of heart disease. However, owing to unmeasured confounding, observational studies do not provide insight into the causal relationship between SU and triglyceride. The aim of this study was to test for a causal role of SU in increasing triglyceride using Mendelian randomization that accounts for unmeasured confounding. Subjects were of European ancestry from the atherosclerosis risk in communities (n=5237) and Framingham heart (n=2971) studies. Mendelian randomization by the 2-stage least squares regression method was done with SU as the exposure, a uric acid transporter genetic risk score as instrumental variable, and triglyceride as the outcome. In ordinary linear regression, SU was significantly associated with triglyceride levels (β=2.69 mmol/L change in triglyceride per mmol/L increase in SU). However, Mendelian randomization-based estimation showed no evidence for a direct causal association of SU with triglyceride concentration-there was a nonsignificant 1.01 mmol/L decrease in triglyceride per mmol/L increase in SU attributable to the genetic risk score (P=0.21). The reverse analysis using a triglyceride genetic risk score provided evidence of a causal role for triglyceride in raising urate in men (P(Corrected)=0.018). These data provide no evidence for a causal role for SU in raising triglyceride levels, consistent with a previous Mendelian randomization report of no association between SU and ischemic heart disease.

Placental uric acid transporter GLUT9 is modulated by free iodine

PLACENTAL URIC ACID TRANSPORTER GLUT9 IS MODULATED BY FREE IODINE Objectives: Materno-fetal transplacental transport is crucial for the fetal well-being. The altered expression of placental transport proteins under specific pathophysiological conditions may affect the intrauterine environment. Pre-eclampsia is often associated with high maternal uric acid serum levels. The regulation of the placental uric transport system and its transporter glucose transporter (GLUT)-9 are not fully understood yet. The aim of this study was to investigate the placental urate transport and to characterize its transporter GLUT9. Methods: In this study we used a transepithelial transport (Transwell®) model to assess uric acid transport activity. Electrophysiological techniques and radioactive ligand up-take assays were used to measure transport activity of GLUT9 expressed in Xenopus oocytes. Results: In the Transwell/model uric acid is transported across the BeWo choriocarcinoma cell monolayer with 530 pmol/min at the linear stage. We could successfully over-express GLUT9 using the Xenopus laevis oocytes expression system. Chloride modulates the urate transport system: interestingly replacing chloride with iodine resulted in a complete loss of urate transport activity.We determined the IC50 of iodine at 30uM concentration. In radioactive up-take experiments iodinehad noeffect on uric acid transport. Conclusions: In vitro the “materno-fetal” transport of uric acid is slow. This indicates that in vivo the child is protected from short-term fluctuations of maternal uric acid serum concentrations. The different results regarding iodine-mediated regulation of GLUT9 transport activity between electrophysiological and radioactive ligand uptake experiments may suggest that iodine does not directly inhibit uric acid transport, but changes the mode of up-take from an electrogenic to an electroneutral transport. GLUT9 is not an uric acid uniporter, there are more ions involved in the transport. This may allow regulating uric acid transport by the change from an active to a passive transport.

Effects of partial pressure of oxygen on the modulation of syncytiotrophoblast potassium channels by reactive oxygen species

s / Placenta 35 (2014) A1eA112 A97 mouse knockout models of impaired sulfonation of structural molecules important for foetal development. Conclusion: Our novel mouse model is providing valuable insight into the critical role of placental sulphate transporters in meeting foetal nutrient demands during gestation. P2.115-N. PLACENTAL URIC ACID TRANSPORTER GLUT9 IS MODULATED BY FREE IODINE Benjamin P. Luscher , Michael Fine , Camilla Marini , Benjamin Clemencon , Christiane Albrecht , Matthias A. Hediger , Daniel V. Surbek , Marc U. Baumann a,b Department of Obstetrics and Gynecology, University Hospital of Bern, Bern, Switzerland; Department of Clinical Research, University of Bern, Bern, Switzerland; c Institute for Biochemistry and Molecular Medicine, University of Bern, Bern, Switzerland Objectives: Materno-fetal transplacental transport is crucial for the fetal well-being. The altered expression of placental transport proteins under specific pathophysiological conditions may affect the intrauterine environment. Pre-eclampsia is often associated with high maternal uric acid serum levels. The regulation of the placental uric transport system and its transporter glucose transporter (GLUT)-9 are not fully understood yet. The aim of this study was to investigate the placental urate transport and to characterize its transporter GLUT9. Methods: In this study we used a transepithelial transport (Transwell®) model to assess uric acid transport activity. Electrophysiological techniques and radioactive ligand up-take assays were used to measure transport activity of GLUT9 expressed in Xenopus oocytes. Results: In the Transwell/ model uric acid is transported across the BeWo choriocarcinoma cell monolayer with 530 pmol/min at the linear stage. We could successfully over-express GLUT9 using the Xenopus laevis oocytes expression system. Chloride modulates the urate transport system: interestingly replacing chloride with iodine resulted in a complete loss of urate transport activity. We determined the IC50 of iodine at 30uM concentration. In radioactiveup-takeexperiments iodinehadnoeffectonuric acid transport. Conclusions: In vitro the “materno-fetal” transport of uric acid is slow. This indicates that in vivo the child is protected from short-term fluctuations of maternal uric acid serum concentrations. The different results regarding iodine-mediated regulation of GLUT9 transport activity between electrophysiological and radioactive ligand uptake experiments may suggest that iodine does not directly inhibit uric acid transport, but changes the mode of up-take from an electrogenic to an electroneutral transport. GLUT9 is not an uric acid uniporter, there are more ions involved in the transport. This may allow regulating uric acid transport by the change from an active to a passive transport. P2.116. UPTAKE MECHANISM OF AN ESTRIOL PRECURSOR, 16a-HYDROXY DEHYDROEPIANDROSTERONE SULFATE, AT THE BASAL PLASMA MEMBRANE OF HUMAN TERM PLACENTA Masatoshi Tomi , Hiromi Eguchi , Tomohiro Nishimura , Tetsuo Maruyama , Emi Nakashima a Keio University Faculty of Pharmacy, Tokyo, Japan; Keio University School of Medicine, Tokyo, Japan Objectives: Estriol is mostly secreted from placental syncytiotrophoblasts during pregnancy. 16a-Hydroxy dehydroepiandrosterone sulfate (16a-OH DHEAS) is an estriol precursor, which is synthesized by fetal liver-specific CYP3A7 (16a-hydroxylase). To achieve placental estriol synthesis, syncytiotrophoblasts require organic anion transporter(s) for 16a-OH DHEAS uptake at the basal plasma membrane (BM) facing fetal circulation. The purpose of this study is to clarify the mechanism underlying 16a-OH DHEAS uptake at the BM of human placental syncytiotrophoblasts. Methods: The uptake of [3H]16a-OH DHEAS was characterized using basal-enriched plasma membrane vesicles (BMVs) derived from human term placenta. The expression of transporters in BMVs was determined by western blot analysis. Results:Proteinexpressionsof organic anion transporter (OAT)4 (SLC22A11) and organic anion transporting polypeptide 2B1 (SLCO2B1)were enriched in human placental BMVs compared with crude membrane fractions, which is consistent with immunohistochemical observations of the basal localization of these transporters in syncytiotrophoblasts. Uptake of [3H]16a-OH DHEAS by BMVs was sodiumand chloride-independent and was trans-stimulated by preloaded glutarate. The uptake of [3H]16a-OH DHEAS was significantly cis-inhibited byDHEAS, estrone sulfate, andbromosulfophthalein, but not by cyclosporin A, tetraethylammonium, p-aminohippuric acid, and cimetidine. These transport characteristics of [3H]16a-OHDHEAS are in good agreement with those in human OAT4-transfected COS7 cells. Conclusion: OAT4 expressed at the basal plasma membrane of the syncytiotrophoblast appears to play a predominant role in the placental uptake of 16a-OH DHEAS from the fetal circulation and contribute to estriol synthesis during pregnancy. P2.117. NANOPARTICLE INTERACTION WITH EARLY HUMAN PLACENTA: PLACENTAL BARRIER MATTERS? Liudmila Nikitina , Gottfried Dohr , Herbert Juch a,b a Institute of Cell Biology, Histology and Embryology, Medical University of Graz, Graz, Austria; b Institute of Human Genetic, Medical University of Graz, Graz,