Substantial anti-gout effect conferred by common and rare dysfunctional variants of URAT1/SLC22A12.

OBJECTIVES: Gout, caused by chronic elevation of serum uric acid levels, is the commonest form of inflammatory arthritis. The causative effect of common and rare variants of ATP-binding cassette transporter G2 (ABCG2/BCRP) on gout risk has been studied, but little attention has been paid to the effect of common (rs121907892, p.W258X) and rare variants of urate transporter 1 (URAT1/SLC22A12) on gout, despite dysfunctional variants of URAT1 having been identified as pathophysiological causes of renal hypouricaemia. METHODS: To address this important but overlooked issue, we investigated the effects of these URAT1 variants on gout susceptibility, using targeted exon sequencing on 480 clinically defined gout cases and 480 controls of Japanese males in combination with a series of functional analyses of newly identified URAT1 variants. RESULTS: Our results show that both common and rare dysfunctional variants of URAT1 markedly decrease the risk of gout (OR 0.0338, reciprocal OR 29.6, P = 7.66 × 10-8). Interestingly, we also found that the URAT1-related protective effect on gout eclipsed the ABCG2-related causative effect (OR 2.30-3.32). Our findings reveal only one dysfunctional variant of URAT1 to have a substantial anti-gout effect, even in the presence of causative variants of ABCG2, a 'gout gene'. CONCLUSION: Our findings provide a better understanding of gout/hyperuricaemia and its aetiology that is highly relevant to personalized health care. The substantial anti-gout effect of common and rare variants of URAT1 identified in the present study support the genetic concept of a 'Common Disease, Multiple Common and Rare Variant' model.

Thromboxane A2 receptor antagonist (ONO-8809) attenuates renal disorders caused by salt overload in stroke-prone spontaneously hypertensive rats.

Epidemiological and clinical studies have demonstrated that excessive salt intake causes severe hypertension and exacerbates organ derangement, such as in chronic kidney disease (CKD). In this study, we focused on evaluating the histological and gene expression effects in the kidneys of stroke-prone spontaneously hypertensive rats (SHRSP) with a high salt intake and the thromboxane A2 / prostaglandin H2 receptor (TPR) blocker ONO-8809. Six-week-old SHRSPs were divided into three groups and were fed normal chow containing 0.4% NaCl, 2.0%NaCl or 2.0%NaCl + ONO-8809 (0.6 mg/kg p.o. daily). Histological analyses with immunohistochemistry and a gene expression assay with a DNA kidney microarray were performed after 8 weeks. The following changes were observed in SHRSPs with the high salt intake. Glomerular sclerotic changes were remarkably observed in the juxtamedullary cortex areas. The ED1, monocyte chemoattractant protein-1 (MCP-1), nitrotyrosine and hypoxia inducible factor 1α (HIF-1α) staining areas were increased in the glomeruli and interstitial portion of the kidneys. The genes Tbxa2r (that encodes TPR), Prcp and Car7 were significantly underexpressed in the kidneys. The plasma 8-isoprostane level was significantly elevated and was attenuated with the ONO-8809 treatment. Thromboxane A2 (TXA2 ) and oxidative stress exaggerated renal dysfunction in the salt-loaded SHRSPs, and ONO-8809 as a TPR blocker suppressed these changes. Therefore, ONO-8809 is a candidate drug to prevent CKD in hypertensive patients when CKD is associated with a high salt intake.

Subtype-specific gout susceptibility loci and enrichment of selection pressure on ABCG2 and ALDH2 identified by subtype genome-wide meta-analyses of clinically defined gout patients.

OBJECTIVES: Genome-wide meta-analyses of clinically defined gout were performed to identify subtype-specific susceptibility loci. Evaluation using selection pressure analysis with these loci was also conducted to investigate genetic risks characteristic of the Japanese population over the last 2000-3000 years. METHODS: Two genome-wide association studies (GWASs) of 3053 clinically defined gout cases and 4554 controls from Japanese males were performed using the Japonica Array and Illumina Array platforms. About 7.2 million single-nucleotide polymorphisms were meta-analysed after imputation. Patients were then divided into four clinical subtypes (the renal underexcretion type, renal overload type, combined type and normal type), and meta-analyses were conducted in the same manner. Selection pressure analyses using singleton density score were also performed on each subtype. RESULTS: In addition to the eight loci we reported previously, two novel loci, PIBF1 and ACSM2B, were identified at a genome-wide significance level (p<5.0×10-8) from a GWAS meta-analysis of all gout patients, and other two novel intergenic loci, CD2-PTGFRN and SLC28A3-NTRK2, from normal type gout patients. Subtype-dependent patterns of Manhattan plots were observed with subtype GWASs of gout patients, indicating that these subtype-specific loci suggest differences in pathophysiology along patients' gout subtypes. Selection pressure analysis revealed significant enrichment of selection pressure on ABCG2 in addition to ALDH2 loci for all subtypes except for normal type gout. CONCLUSIONS: Our findings on subtype GWAS meta-analyses and selection pressure analysis of gout will assist elucidation of the subtype-dependent molecular targets and evolutionary involvement among genotype, phenotype and subtype-specific tailor-made medicine/prevention of gout and hyperuricaemia.

Genome-wide association study revealed novel loci which aggravate asymptomatic hyperuricaemia into gout.

OBJECTIVE: The first ever genome-wide association study (GWAS) of clinically defined gout cases and asymptomatic hyperuricaemia (AHUA) controls was performed to identify novel gout loci that aggravate AHUA into gout. METHODS: We carried out a GWAS of 945 clinically defined gout cases and 1003 AHUA controls followed by 2 replication studies. In total, 2860 gout cases and 3149 AHUA controls (all Japanese men) were analysed. We also compared the ORs for each locus in the present GWAS (gout vs AHUA) with those in the previous GWAS (gout vs normouricaemia). RESULTS: This new approach enabled us to identify two novel gout loci (rs7927466 of CNTN5 and rs9952962 of MIR302F) and one suggestive locus (rs12980365 of ZNF724) at the genome-wide significance level (p<5.0×10-8). The present study also identified the loci of ABCG2, ALDH2 and SLC2A9. One of them, rs671 of ALDH2, was identified as a gout locus by GWAS for the first time. Comparing ORs for each locus in the present versus the previous GWAS revealed three 'gout vs AHUA GWAS'-specific loci (CNTN5, MIR302F and ZNF724) to be clearly associated with mechanisms of gout development which distinctly differ from the known gout risk loci that basically elevate serum uric acid level. CONCLUSIONS: This meta-analysis is the first to reveal the loci associated with crystal-induced inflammation, the last step in gout development that aggravates AHUA into gout. Our findings should help to elucidate the molecular mechanisms of gout development and assist the prevention of gout attacks in high-risk AHUA individuals.

Common variant of BCAS3 is associated with gout risk in Japanese population: the first replication study after gout GWAS in Han Chinese.

BACKGROUND: Gout is a common disease resulting from hyperuricemia which causes acute arthritis. A recent genome-wide association study (GWAS) of gout identified three new loci for gout in Han Chinese: regulatory factor X3 (RFX3), potassium voltage-gated channel subfamily Q member 1 (KCNQ1), and breast carcinoma amplified sequence 3 (BCAS3). The lack of any replication studies of these three loci using other population groups prompted us to perform a replication study with Japanese clinically defined gout cases and controls. METHODS: We genotyped the variants of RFX3 (rs12236871), KCNQ1 (rs179785) and BCAS3 (rs11653176) in 723 Japanese clinically defined gout cases and 913 controls by TaqMan method. rs179785 of KCNQ1 is also evaluated by direct sequencing because of difficulties of its genotyping by TaqMan method. RESULTS: Although the variants of RFX3 and BCAS3 were clearly genotyped by TaqMan method, rs179785 of KCNQ1 was not, because rs179785 (A/G) of KCNQ1 is located at the last nucleotide ("A") of the 12-bp deletion variant (rs200562977) of KCNQ1. Therefore, rs179785 and rs200562977 of KCNQ1 were genotyped by direct sequencing in all samples. Moreover, by direct sequencing with the same primers, we were able to evaluate the genotypes of rs179784 of KCNQ1 which shows strong linkage disequilibrium with rs179785 (D' = 1.0 and r 2 = 0.99). rs11653176, a common variant of BCAS3, showed a significant association with gout (P = 1.66 × 10- 3; odds ratio [OR] = 0.80); the direction of effect was the same as that seen in the previous Han Chinese GWAS. Two variants of KCNQ1 (rs179785 and rs179784) had a nominally significant association (P = 0.043 and 0.044; OR = 0.85 and 0.86, respectively), but did not pass the significance threshold for multiple hypothesis testing using the Bonferroni correction. On the other hand, rs200562977 of KCNQ1 and rs12236871 of RFX3 did not show any significant association with gout. CONCLUSION: BCAS3 is a coactivator of estrogen receptor alpha, and the influence of estrogen to serum uric acid level is well known. Our present replication study, as did the previous gout GWAS, demonstrated the common variant of BCAS3 to be associated with gout susceptibility.

GWAS of clinically defined gout and subtypes identifies multiple susceptibility loci that include urate transporter genes.

OBJECTIVE: A genome-wide association study (GWAS) of gout and its subtypes was performed to identify novel gout loci, including those that are subtype-specific. METHODS: Putative causal association signals from a GWAS of 945 clinically defined gout cases and 1213 controls from Japanese males were replicated with 1396 cases and 1268 controls using a custom chip of 1961 single nucleotide polymorphisms (SNPs). We also first conducted GWASs of gout subtypes. Replication with Caucasian and New Zealand Polynesian samples was done to further validate the loci identified in this study. RESULTS: In addition to the five loci we reported previously, further susceptibility loci were identified at a genome-wide significance level (p<5.0×10-8): urate transporter genes (SLC22A12 and SLC17A1) and HIST1H2BF-HIST1H4E for all gout cases, and NIPAL1 and FAM35A for the renal underexcretion gout subtype. While NIPAL1 encodes a magnesium transporter, functional analysis did not detect urate transport via NIPAL1, suggesting an indirect association with urate handling. Localisation analysis in the human kidney revealed expression of NIPAL1 and FAM35A mainly in the distal tubules, which suggests the involvement of the distal nephron in urate handling in humans. Clinically ascertained male patients with gout and controls of Caucasian and Polynesian ancestries were also genotyped, and FAM35A was associated with gout in all cases. A meta-analysis of the three populations revealed FAM35A to be associated with gout at a genome-wide level of significance (p meta =3.58×10-8). CONCLUSIONS: Our findings including novel gout risk loci provide further understanding of the molecular pathogenesis of gout and lead to a novel concept for the therapeutic target of gout/hyperuricaemia.

Mechanisms of thiazide-induced hypertension treatment: insights from gene expression and histological analysis in malignant stroke-prone spontaneously hypertensive rats.

OBJECTIVE: Diuretics, including thiazides and thiazide-like drugs, are commonly recommended for treating hypertension, though their precise mechanism of action is not fully understood. This study aimed to investigate the pharmacological effects of trichloromethiazide (TCM) in malignant stroke-prone spontaneously hypertensive rats (M-SHRSP). METHODS: M-SHRSPs were treated with varying doses of TCM. Prognosis, histological changes, and mRNA expression related to hypertension and stroke were assessed. RESULTS: The high-dose TCM group (3%) exhibited significantly lower SBP compared with the untreated group, whereas the low-dose group (0.3%) did not show a significant reduction in SBP. The survival rate was 54% in the low-dose group, whereas all rats in the high-dose group survived without experiencing a stroke by 16 weeks of age. Organ weights in both TCM-treated groups were lower than those in the control group, without severe histological abnormalities, including stroke and sclerosis. Plasma levels of thiobarbituric acid-reactive substances (TBARS) were significantly reduced in both TCM-treated groups. Additionally, 20 genes related to tissue protection, repair, proliferation, maintenance, and function were significantly expressed. CONCLUSION: TCM administration in M-SHRSPs significantly modulated the expression of 20 genes associated with tissue protection and maintenance, and reduced plasma TBARS levels. These findings suggest that TCM, a thiazide diuretic, may protect against tissue impairment in hypertension by modulating gene expression and exhibiting antioxidant activity.

Coincidence of acral peeling skin syndrome and Nagashima-type palmoplantar keratosis in a Japanese pedigree with acral skin peeling.

Acral peeling skin syndrome (APSS; MIM 609796) is a rare genodermatosis characterized by painless focal cutaneous exfoliation of the dorsal hands and feet, typically displaying autosomal recessive inheritance. While cases associated with a founder mutation in TGM5 are relatively common in European Caucasian populations, no APSS cases have been reported from Japan or other East Asian countries. In contrast, Nagashima-type palmoplantar keratosis (NPPK; MIM 615598), caused by variants in SERPINB7, is relatively common in East Asia due to founder mutations. We describe a 27-year-old Japanese woman with spontaneous focal cutaneous exfoliation of the dorsal hand following prolonged glove use, indicative of APSS. Histopathological examination revealed a cleft between the stratum corneum and stratum granulosum and within the horny layer of the epidermis, supporting this diagnosis. However, her mother and maternal uncle exhibited similar symptoms, and there was no reported consanguinity in the patient's parents or grandparents, prompting suspicion of an autosomal dominant genodermatosis. Whole-genome sequencing (WGS) revealed compound heterozygous variants in TGM5 (c.1037G>A and c.684 + 1G>A) as suspected causative variants in the patient, leading to an APSS diagnosis, the first reported in East Asia. On the other hand, her mother and maternal uncle were diagnosed with NPPK due to compound heterozygous pathogenic variants in SERPINB7 (c.796C>T and c.455-1G>A). This case highlights the complexity of diagnosing skin disorders when multiple genodermatoses with similar phenotypes exist within a pedigree. Comprehensive genetic analyses, such as whole-exome sequencing and WGS, are invaluable for identifying causative variants in such complex cases.