Growth factor dependence of the proliferation and survival of cultured lacrimal gland epithelial cells isolated from late-embryonic mice.

Epidermal growth factor (EGF) and hepatocyte growth factor (HGF) regulate the growth and morphogenesis of various exocrine glands with branched morphologies. Their roles in lacrimal gland (LG) development remain unknown, but fibroblast growth factor (FGF) 10 is crucial for early LG organogenesis. To clarify the roles of EGF, HGF, and FGF10 in LG development, LG epithelial cells were isolated from late-embryonic and neonatal mice; cultured; and treated with EGF, HGF, or FGF10 and their respective receptor tyrosine kinase (RTK) inhibitors AG1478, PHA665752, or SU5402. EGF and HGF increased the number of viable cells by enhancing DNA synthesis, FGF10 and SU5402 showed no such effect, and RTK inhibitors exhibited the opposite effect. EGF and HGF receptors were immunostained in cultured late-embryonic LG epithelial cells and terminal LG acini from late embryos and adult mice. HGF was detected in neonatal LG epithelial cell culture supernatants by western blotting. In the absence of EGF and HGF RTK inhibitors, growth factor addition increased the number of viable cells and suppressed cell death. However, when one RTK was inhibited and a growth factor targeting an intact RTK was added, the number of dead cells increased as the number of viable cells increased. No cells survived when both RTKs were inhibited. In explant cultures of LGs from embryos, AG1478 or PHA665752 decreased the number of Ki67-positive proliferating epithelial cells in terminal acini. Thus, EGF and HGF may function in a cooperative autocrine manner, supporting cell proliferation and survival during LG development in late-embryonic and neonatal mice.

Bacterial Translocation in Gastrointestinal Cancers and Cancer Treatment.

In recent years, there has been increasing evidence that gut microbiota is associated with the onset and exacerbation of various diseases, such as gastrointestinal cancer. For instance, it is well known that local inflammation of the intestinal tract in colorectal cancer that is caused by the increased number of Fusobacterium, due to changes in the intestinal bacterial flora, is involved in carcinogenesis. In contrast, gut bacteria or their products, pathogen-associated molecular patterns, not only cause intestinal inflammation but also invade the bloodstream through dysbiosis and gut barrier dysfunction, thereby leading to systemic inflammation, namely bacterial translocation. The involvement of bacterial translocation in the carcinogenesis of gastrointestinal cancers and their prognosis is increasingly being recognized. The Toll-like receptor signaling pathways plays an important role in the carcinogenesis of such cancers. In addition, bacterial translocation influences the treatment of cancers such as surgery and chemotherapy. In this review, we outline the concept of bacterial translocation, summarize the current knowledge on the relationship between gut bacteria and gastrointestinal cancer, and provide future perspectives of this field.

Ascorbic acid-2 glucoside mitigates intestinal damage during pelvic radiotherapy in a rat bladder tumor model.

PURPOSE: Ascorbic acid is a strong antioxidant and has potent radioprotective effects on radiation injuries. Ascorbic acid 2-glucoside (AA2G) is a stabilized derivative of ascorbic acid and rapidly hydrolyzed into ascorbic acid and glucose. Since there is the possibility that AA2G treatment interferes with the antitumor activity of radiotherapy, we investigated the effect of AA2G treatment during radiotherapy on acute radiation enteritis and antitumor activity of radiotherapy in rats. MATERIALS AND METHODS: AY-27 rat bladder tumor cells were used to induce bladder tumors in rats. Two weeks after inoculation rats received fractionated pelvic radiotherapy in eight fractions for 4 weeks totaling 40 Gy. During radiotherapy, one group of rats received per os AA2G (ascorbic acid: 250 mg/kg/day) and its bolus engulfment (ascorbic acid: 250 mg/kg) 8 h before each X-irradiation fraction. Seven days after the last X-irradiation, we studied histology, DNA double strand break (DSB) damage (by 53BP1 foci staining), and the M1/M2 macrophage response by immunohistochemistry of paraffin-fixed bladder and intestinal tissues. RESULTS: AA2G treatment reduced the intestinal damage (shortening of villi) but did not reduce antitumor effectiveness of radiotherapy against bladder tumors. Like the controls, AA2G-treated rats showed no residual tumor lesions in the bladder after X-irradiation. Both AA2G-treated and control groups showed similar persistent DSB damage (53BP1 foci) both in bladders and ilea seven days after radiotherapy. Radiotherapy tended to reduce CD163+ M2 macrophages, which are considered as an anti-inflammatory subtype favoring tissue repair, in the bladders. X-irradiation also reduced the occurrence of M2 macrophages in the ilea. AA2G treatment significantly increased CD163+/CD68+ macrophage ratio in the ilea of rats after pelvic irradiation in comparison to the sham irradiated control rats. AA2G treatment increased, albeit not significantly, the CD163+/CD68+ macrophage ratio in the irradiated bladders relative to the control irradiated rats. On the other hand, bladders and ilea of the irradiated rats with and without AA2G treatment showed similar frequencies of CD68+ macrophages. CONCLUSIONS: AA2G treatment mitigated radiation-induced intestinal damage without reducing antitumor activity after fractionated pelvic radiotherapy against bladder tumors in rats. The beneficial effect of AA2G treatment seems to promote a restoration of the M2 answer as well as tissue remodeling and wound healing. Similar residual DNA damage in bladders and ilea seven days post-irradiation is consistent with tumor control in both groups.

Modeling principles of protective thyroid blocking.

PURPOSE: In the case of a nuclear incident, the release of radioiodine must be expected. Radioiodine accumulates in the thyroid and by irradiation enhances the risk of cancer. Large doses of stable (non-radioactive) iodine may inhibit radioiodine accumulation and protect the thyroid ('thyroid blocking'). Protection is based on a competition at the active carrier site in the cellular membrane and an additional temporary inhibition of the organification of iodide (Wolff-Chaikoff effect). Alternatively, other agents like e.g. perchlorate that compete with iodide for the uptake into the thyrocytes may also confer thyroidal protection against radioiodine exposure.Biokinetic models for radioiodine mostly describe exchanges between compartments by first order kinetics. This leads to correct predictions only for low (radio)iodide concentrations. These models are not suited to describe the kinetics of iodine if administered at the dosages recommended for thyroid blocking and moreover does not permit to simulate either the protective competition mechanism at the membrane or the Wolff-Chaikoff effect. Models adapted for this purpose must be used. Such models may use a mathematical relation between the serum iodide concentration and a relative uptake suppression or a dependent rate constant determining total thyroidal radioiodine accumulation. Alternatively, the thyroidal uptake rate constant may be modeled as a function of the total iodine content of the gland relative to a saturation amount. Newer models integrate a carrier-mechanism described by Michalis-Menten kinetics in the membrane and in analogy to enzyme kinetics apply the rate law for monomolecular irreversible enzyme reactions with competing substrates to model the competition mechanism. An additional total iodide uptake block, independent on competition but limited in time, is used to simulate the Wolff-Chaikoff effect. CONCLUSION: The selection of the best model depends on the issue to be studied. Most models cannot quantify the relative contributions of the competition mechanism at the membrane and the Wolff-Chaikoff effect. This makes it impossible or exceedingly difficult to simulate prolonged radioiodine exposure and the effect of repetitive administrations of stable iodine. The newer thyroid blocking models with a separate modeling of competition and Wolff-Chaikoff effect allow better quantitative mechanistic insights and offer the possibility to simulate complex radioiodine exposure scenarios and various protective dosage schemes of stable iodine relatively easily. Moreover, they permit to study the protective effects of other competitors at the membrane carrier site, like e.g. perchlorate, and to draw conclusions on their protective efficacy in comparison to stable iodine.

Genetic epidemiological analysis of hypouricaemia from 4993 Japanese on non-functional variants of URAT1/SLC22A12 gene.

OBJECTIVES: Up to 0.3% of Japanese have hypouricaemia. Most cases appear to result from a hereditary disease, renal hypouricaemia (RHUC), which causes exercise-induced acute kidney injury and urolithiasis. However, to what extent RHUC accounts for hypouricaemia is not known. We therefore investigated its frequency and evaluated its risks by genotyping a general Japanese population. METHODS: A cohort of 4993 Japanese was examined by genotyping the non-functional variants R90H (rs121907896) and W258X (rs121907892) of URAT1/SLC22A12, the two most common causative variants of RHUC in Japanese. RESULTS: Participants' fractional excretion of uric acid and risk allele frequencies markedly increased at lower serum uric acid (SUA) levels. Ten participants (0.200%) had an SUA level ≤2.0 mg/dl and nine had R90H or W258X and were likely to have RHUC. Logistic regression analysis revealed these URAT1 variants to be significantly and independently associated with the risk of hypouricaemia and mild hypouricaemia (SUA ≤3.0 mg/dl) as well as sex, age and BMI, but these URAT1 variants were the only risks in the hypouricaemia population (SUA ≤2.0 mg/dl). W258X was only a risk in males with SUA ≤3.0 mg/dl. CONCLUSION: Our study accurately reveals the prevalence of RHUC and provides genetic evidence for its definition (SUA ≤2.0 mg/dl). We also show that individuals with SUA ≤3.0 mg/dl, especially males, are prone to RHUC. Our findings will help to promote a better epidemiological understanding of RHUC as well as more accurate diagnosis, especially in males with mild hypouricaemia.

Preoperative Fall Risk Assessment Score as a Prognostic Factor in Esophageal Cancer Patients after Esophagectomy.

The current study investigated the impact of preoperative fall risk assessment score (FRAS) on long-term prognoses in patients with esophageal cancer (EC). A total of 161 patients with EC who underwent curative surgery were classified into a high-risk (95, 41.0%) and low-risk (66, 41.0%) groups according to their FRAS. This study investigated the relationships between the FRAS and clinicopathological findings and prognoses. Accordingly, patients in the high-risk group were significantly older and had a significantly higher Charlson comorbidity index than those in the low-risk group. No significant difference was found in pathological findings between both groups. The high-risk group had significantly lower overall survival (OS) and relapse-free survival (RFS) rates than the low-risk group (p = 0.004 and 0.001, respectively). Multivariate analysis identified high FRAS as an independent prognostic factor for poor OS, with a hazard ratio of 1.75 (p = 0.033). Moreover, re-analysis of the data after excluding age as a category showed that the high-risk group had significantly worse OS (p = 0.004) and RFS (p = 0.003) than the low-risk group. The FRAS can, therefore, be considered a useful method for assessing frailty and a potential prognostic factor for EC.

Role of Microbial Infection-Induced Inflammation in the Development of Gastrointestinal Cancers.

There has been increasing evidence that a local inflammatory response stimulates tumor cells to acquire metastatic potential, and the concept of inflammatory oncotaxis has been spreading in recent years. However, the interaction between microbial inflammation and the development of gastrointestinal cancer is still unclear. This review summarizes the present knowledge on the role of microbial inflammation in the development of gastrointestinal cancers from the perspective of molecular biological findings. Chronic inflammation caused by bacterial infection is known to induce cancers as exemplified by Helicobacter pylori, which is associated with the development of gastric cancer via the activation of the TLR4 pathway by bacterial lipopolysaccharide followed by cancer growth through CagA-MET signaling. In addition, the development of inflammatory bowel diseases has been known to become a risk factor for colorectal cancers, where inflammation caused by certain bacterial infections plays a key role. It is also known that the cancer microenvironment is associated with cancer growth. Moreover, infectious complication after surgery for gastrointestinal cancers may promote tumor progression via the stimulation of pathogen-associated molecular patterns and various inflammatory mediators secreted by immunocytes. Further research on the link between microbial inflammation and cancer progression is needed to drive a paradigm shift in cancer treatment.

Porphyrin accumulation in humans with common dysfunctional variants of ABCG2, a porphyrin transporter: potential association with acquired photosensitivity.

Photosensitivity is a skin reaction disorder mediated by phototoxic and/or photoallergic mechanisms. The accumulation of porphyrins is generally considered to induce phototoxicity. ATP-binding cassette subfamily G member 2 (ABCG2) has been identified as a transporter of porphyrins and its common variants-p.Gln126Ter (rs72552713) and p.Gln141Lys (rs2231142)-reportedly decrease the function of porphyrin transport in vitro; however, the physiological importance of ABCG2 as a porphyrin transporter remains to be fully elucidated. We herein investigated whether ABCG2 dysfunction could lead to porphyrin accumulation and photosensitivity in Japanese subjects, and found it to be significantly correlated with erythrocyte protoporphyrin levels (P = 0.012). This appears to be the first clinical finding of ABCG2 dysfunction-associated protoporphyrin accumulation in humans. We divided the patients into a chronic actinic dermatosis (CAD) group and a non-CAD group. CAD was diagnosed based on the criteria of reduced minimal erythema doses to ultraviolet B (UVB) and/or ultraviolet A (UVA). The non-CAD group was composed of patients who exhibited normal reactions to UVB and UVA on phototesting, but had histories of recurrent erythema/papules on sun-exposed areas. Estimated ABCG2 function according to ABCG2 genotypes in the non-CAD group was significantly lower than in the general Japanese population (P = 0.045). In contrast, no difference was found in ABCG2 function between the CAD group and the general population, suggesting that ABCG2 dysfunction might be a genetic factor in non-CAD patients with clinical photosensitivity. In this context, genetic dysfunction of ABCG2 might be an overlooked pathological etiology of "photosensitivity of unknown cause."

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.

Helicobacter pylori-Mediated Immunity and Signaling Transduction in Gastric Cancer.

Helicobacter pylori infection is a leading cause of gastric cancer, which is the second-most common cancer-related death in the world. The chronic inflammatory environment in the gastric mucosal epithelia during H. pylori infection stimulates intracellular signaling pathways, namely inflammatory signals, which may lead to the promotion and progression of cancer cells. We herein report two important signal transduction pathways, the LPS-TLR4 and CagA-MET pathways. Upon H. pylori stimulation, lipopolysaccharide (LPS) binds to toll-like receptor 4 (TLR4) mainly on macrophages and gastric epithelial cells. This induces an inflammatory response in the gastric epithelia to upregulate transcription factors, such as NF-κB, AP-1, and IRFs, all of which contribute to the initiation and progression of gastric cancer cells. Compared with other bacterial LPSs, H. pylori LPS has a unique function of inhibiting the mononuclear cell (MNC)-based production of IL-12 and IFN-γ. While this mechanism reduces the degree of inflammatory reaction of immune cells, it also promotes the survival of gastric cancer cells. The HGF/SF-MET signaling plays a major role in promoting cellular proliferation, motility, migration, survival, and angiogenesis, all of which are essential factors for cancer progression. H. pylori infection may facilitate MET downstream signaling in gastric cancer cells through its CagA protein via phosphorylation-dependent and/or phosphorylation-independent pathways. Other signaling pathways involved in H. pylori infection include EGFR, FAK, and Wnt/ß-Catenin. These pathways function in the inflammatory process of gastric epithelial mucosa, as well as the progression of gastric cancer cells. Thus, H. pylori infection-mediated chronic inflammation plays an important role in the development and progression of gastric cancer.

A Potential Role of Adhesion Molecules on Lung Metastasis Enhanced by Local Inflammation.

BACKGROUND/AIM: Local and systemic inflammations are associated with negative long-term outcomes; however, their precise mechanism of action remains unclear. We previously demonstrated that hepatocyte growth factor (HGF)/c-Met signaling contributed to the enhancement of liver metastasis associated with peritonitis model. The aim of this study is to investigate the effect of local inflammation on the development of lung metastasis. MATERIALS AND METHODS: NL-17 cells were injected into BALB/c mice via the tail vein to produce a high potential model for lung metastasis. After injection of NL-17 cells, lipopolysaccharide (LPS) and live Pseudomonas aeruginosa, and phosphate-buffered saline were administered intratracheally to induce acute lung injury (ALI) and pneumonia, respectively. RESULTS: In both ALI and pneumonia mice, lung metastasis was significantly promoted compared to control mice. Concentrations of Interleukin-6, tumor necrosis factor-α, and HGF in the bronchoalveolar lavage fluid were significantly higher in ALI and pneumonia mice than in control mice. Neither administration of recombinant mouse HGF nor c-Met knockdown in NL-17 cells influenced the magnitude of lung metastasis. Yet stimulation with LPS increased the expression of α2 integrin, vascular cell-adhesion protein-1, and intercellular adhesion molecule-1 (ICAM-1) in the lung. Invasive activity of NL-17 cells was significantly up-regulated by LPS, but was suppressed by anti-ICAM-1 antibody. While LPS-stimulated NL-17 cells showed significantly promoted lung metastasis, E-selectin expression in the lungs of mice with ALI or pneumonia was significantly enhanced compared with control mice. CONCLUSION: Up-regulation of adhesion molecules, but not HGF/c-Met signaling, may contribute to the lung metastasis enhanced by local infection/inflammation.

A comparison of thyroidal protection by stable iodine or perchlorate in the case of acute or prolonged radioiodine exposure.

In the case of a nuclear power plant accident, repetitive/prolonged radioiodine release may occur. Radioiodine accumulates in the thyroid and by irradiation enhances the risk of cancer. Large doses of non-radioactive iodine may protect the thyroid by inhibiting radioiodine uptake into the gland (iodine blockade). Protection is based on a competition at the active carrier site in the cellular membrane and the Wolff-Chaikoff effect, the latter being, however, only transient (24-48 h). Perchlorate may alternatively provide protection by a carrier competition mechanism only. Perchlorate has, however, a stronger affinity to the carrier than iodide. Based on an established biokinetic-dosimetric model developed to study iodine blockade, and after its extension to describe perchlorate pharmacokinetics and the inhibition of iodine transport through the carrier, we computed the protective efficacies that can be achieved by stable iodine or perchlorate in the case of an acute or prolonged radioiodine exposure. In the case of acute radioiodine exposure, perchlorate is less potent than stable iodine considering its ED50. A dose of 100 mg stable iodine has roughly the same protective efficacy as 1000 mg perchlorate. For prolonged exposures, single doses of protective agents, whether stable iodine or perchlorate, offer substantially lower protection than after acute radioiodine exposure, and thus repetitive administrations seem necessary. In case of prolonged exposure, the higher affinity of perchlorate for the carrier in combination with the fading Wolff-Chaikoff effect of iodine confers perchlorate a higher protective efficacy compared to stable iodine. Taking into account the frequency and seriousness of adverse effects, iodine and perchlorate at equieffective dosages seem to be alternatives in case of short-term acute radioiodine exposure, whereas preference should be given to perchlorate in view of its higher protective efficacy in the case of longer lasting radioiodine exposures.

A Potential Mechanism of Tumor Progression during Systemic Infections Via the Hepatocyte Growth Factor (HGF)/c-Met Signaling Pathway.

BACKGROUND: Increasing evidence has demonstrated that postoperative infectious complications (PICs) after digestive surgery are significantly associated with negative long-term outcomes; however, precise mechanisms of how PICs affect the poor long-term survival remain unclear. Here, we focused on the hepatocyte growth factor (HGF)/c-Met signaling pathway as one of those mechanisms. Methods: In the clinical setting, serum HGF levels were measured in the patients with sepsis and those with PICs after undergoing esophagectomy. Using a liver metastasis mouse model with cecal ligation and puncture (CLP), expressions of HGF and the roles of the HGF/c-Met pathway in the progression of tumor cells were examined. Results: Serum HGF levels were very high in the patients with intra-abdominal infection on postoperative days (PODs) 1, 3, and 5; similarly, compared to the patients without PICs, those with PICs had significantly higher serum HGF levels on 1, 3, and 5 days after esophagectomy. The patients with PICs showed poorer overall survival than those without PICs, and the patients with high serum HGF levels on POD 3 showed poorer prognosis than those with low HGF levels. Similarly, at 24 and 72 h after operation, serum levels of HGF in CLP mice were significantly higher than those in sham-operated mice. Intraperitoneal injection of mouse recombinant HGF significantly promoted liver metastases in sham-operated mice on 14 days after surgery. Knocking down c-Met expression on NL17 tumor cells by RNAi technology significantly inhibited the promotion of CLP-induced liver metastases. Conclusions: Infections after surgery increased serum HGF levels in the clinical as well as experimental settings. Induction of high serum HGF levels by CLP promoted liver metastases in a murine liver metastasis model, suggesting the involvement of the HGF/c-Met signaling pathway in tumor promotion mechanisms. Thus, targeting the HGF/c-Met signaling pathway may be a promising approach for malignant tumors, particularly in the patients with PICs.

Dysfunctional ABCG2 gene polymorphisms are associated with serum uric acid levels and all-cause mortality in hemodialysis patients.

Dysfunctional variants of ATP-binding cassette transporter subfamily G member 2 (ABCG2), a urate transporter in the kidney and intestine, are the major causes of hyperuricemia and gout. A recent study found that ABCG2 is a major transporter of uremic toxins; however, few studies have investigated the relationship between ABCG2 gene polymorphisms and mortality. This prospective cohort study of 1214 hemodialysis patients investigated the association between serum uric acid levels and ABCG2 genotype and mortality. Genotyping of dysfunctional ABCG2 variants, Q126X (rs72552713) and Q141K (rs2231142), was performed using the patients' DNA. During the study period, 220 patients died. Lower serum uric acid levels were associated with higher mortality (hazard ratio [HR] 1.89, 95% confidence interval [CI] 1.14-3.10, P ≤ 0.001). ABCG2 dysfunction, estimated by genetic variants, had a significant positive association with serum uric acid levels (full function: 7.4 ± 1.2 mg/dl, 3/4 function: 7.9 ± 1.3 mg/dl, 1/2 function: 8.2 ± 1.4 mg/dl, ≤ 1/4 function: 8.7 ± 1.3 mg/dl, P ≤ 0.001). This association remained significant on multiple regression analysis. The Cox proportional hazard analysis indicated that the ABCG2 ≤ 1/4 function type was significantly associated with higher mortality (HR 6.66, 95% CI 2.49 to 17.8, P ≤ 0.001) than the other function types. These results showed that ABCG2 plays a physiologically important role in uric acid excretion, and that ABCG2 dysfunction is a risk factor for mortality in hemodialysis patients.

Development of New Biokinetic-Dosimetric Models for the Simulation of Iodine Blockade in the Case of Radioiodine Exposure in Man.

In the case of nuclear incidents, radioiodine may be liberated. After incorporation it accumulates in the thyroid and by internal irradiation enhances the risk of cancer occurrence. By administering a large dose of non-radioactive iodine the uptake of radioiodine into the gland can be inhibited ("iodine blockade"). Biokinetic models using first order kinetics are not suited to simulate iodine blockade, as the uptake into the gland is mediated by a saturable active transport. Therefore, we integrated an uptake mechanism described by a Michaelis-Menten kinetic into a simple ICRP biokinetic model. We moreover added a total uptake blocking mechanism representing the Wolff-Chaikoff effect becoming active when the gland is saturated with iodine. The validity of the model was ascertained by comparison with IMBA software. The competition of radioiodine and stable iodine at the membrane carrier site was modeled according to the rate law for monomolecular reactions for competing substrates. Our simulations show that competition for the uptake at the membrane carrier site accounts for about 60% and the saturation of the gland with iodine for over 35% of the total protective efficacy that exceeds 95%. Following acute radioiodine exposure, it is preferable to administer a single large dose of stable iodine. In the case of continuous radioiodine exposure, a single dose of stable iodine is less effective than after an acute exposure and splitting the total available dose and shortening the dosage intervals enhance efficacy. Model-based simulations may be a useful tool to develop antidote dosage schemes for uncommon emergencies.

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.