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The aryl hydrocarbon receptor (AHR), a major chemical sensor, is thought to play a role in various biological contexts, including cell cycle regulation and tumorigenesis. However, its regulatory mechanisms remain unclear. We propose herein a novel mechanism through which AHR promotes tumorigenesis by targeting expression of the oncogene intestine-specific homeobox (ISX) in hepatocellular carcinoma (HCC). Compared to paired tumor-adjacent tissues and non-HCC tumors, HCCs exhibited an increased and hierarchical pattern of AHR expression. Patients exhibiting high AHR expression had a significantly shorter survival duration, compared to those with low and medium expression. Functionally, AHR was found to target the newly discovered proto-oncogene, ISX, resulting in the increased expression of this gene and its downstream targets, CCND1 and E2F1. Ablation of AHR or ISX in hepatoma cells suppressed cell growth, whereas overexpression promoted cell proliferation and led to enhanced tumorigenic activity in vitro and in vivo. These results provide evidence to support a critical role for the AHR/ISX axis in HCC tumorigenesis and suggest its potential utility as a new therapeutic and prognostic target for HCC.

Peripheral artery disease (PAD) is a manifestation of systemic atherosclerosis and conveys a significant health burden globally. Critical limb ischaemia encompasses the most severe consequence of PAD. Our previous studies indicate that microRNA let‐7g prevents atherosclerosis and improves endothelial functions. This study aimed to investigate whether and how let‐7g therapy may improve blood flow to ischaemic limbs. The present study shows that let‐7g has multiple pro‐angiogenic effects on mouse ischaemic limb model and could be a potential therapeutic agent for PAD. Mice receiving intramuscular injection of let‐7g had more neovascularization, better local perfusion and increased recruitment of endothelial progenitor cells after hindlimb ischaemia. The therapeutic effects of let‐7g's on angiogenesis are mediated by multiple regulatory machinery. First, let‐7g increased expression of vascular endothelial growth factor‐A (VEGF‐A) and VEGF receptor‐2 (VEGFR‐2) through targeting their upstream regulators HIF‐3α and TP53. In addition, let‐7g affected the splicing factor SC35 which subsequently enhanced the alternative splicing of VEGF‐A from the anti‐angiogenic isoform VEGF‐A165b towards the pro‐angiogenic isoform VEGF‐A164a. The pleiotropic effects of let‐7g on angiogenesis imply that let‐7g may possess a therapeutic potential in ischaemic diseases.

The intestine-specific homeobox transcription factor intestine-specific homeobox (ISX) is an IL6-inducible proto-oncogene implicated in the development of hepatocellular carcinoma, but its mechanistic contributions to this process are undefined. In this study, we provide evidence that ISX mediates a positive feedback loop integrating inflammation, tryptophan catabolism, and immune suppression. We found that ISX-mediated IL6-induced expression of the tryptophan catabolic enzymes Indoleamine 2,3-dioxygenase 1 (IDO1) and tryptophan 2,3-dioxygenase in hepatocellular carcinoma cells, resulting in an ISX-dependent increase in the tryptophan catabolite kynurenine and its receptor aryl hydrocarbon receptor (AHR). Activation of this kynurenine/AHR signaling axis acted through a positive feedback mechanism to increase ISX expression and enhance cellular proliferation and tumorigenic potential. RNAi-mediated attenuation of ISX or AHR reversed these effects. In an IDO1-dependent manner, ectopic expression of ISX induced expression of genes encoding the critical immune modulators CD86 (B7-2) and programmed death ligand-1 (PD-L1), through which ISX conferred a significant suppressive effect on the CD8

The level of global DNA methylation may be related to the cerebrovascular disease. The methylation level of Long Interspersed Nucleotide Element 1 (LINE-1) can represent the global methylation level. We investigated the association between the methylation levels of LINE-1 and ischemic stroke in Chinese. Two hundred and eighty patients of ischemic stroke and 280 age- and sex-matched controls were enrolled. The mean percentage of three CpG sites of LINE-1 was calculated for each subject and was used for analysis. Twenty four samples were re-checked for reproducibility of the methylation assay. Multivariate regression model was used to estimate the odds ratio of stroke risk for one percent change of methylation. Sex-specific analysis was also conducted. Thirty-two cases and 11 controls did not pass the methylation assay criteria, and were excluded from further analysis. The intra-class correlation has a coefficient of 0.97 for the methylation assay. The stroke cases had a lower methylation level than controls (p=0.002), especially male subjects (p=0.001). Sex-specific analysis showed that a decrease of 1% methylation level in men could increase a stroke risk by 1.2-fold after adjusting for other risk factors. LINE-1 methylation levels did not have a significant association with stroke in women. The present study shows that a lower level of LINE-1 methylation was associated with a higher risk for ischemic stroke in men, but methylation level in women did not affect the stroke risk. Our finding in Chinese is consistent with a previous result based on elderly white men.

Objective Single nucleotide polymorphisms (SNPs) located at microRNA (miRNA) binding sites (miR-SNPs) can affect the expression of genes. This study aimed to identify the miR-SNPs associated with atherosclerosis and stroke. Methods Patients with ischemic stroke ( n = 657) and stroke- and myocardial infarction-free volunteers ( n = 1571) were enrolled. The carotid intima-media thickness (IMT) was measured in the control participants. Seventy-nine stroke susceptibility genes were initially selected and 13 genes were predicted to have miR-SNPs at their 3′ untranslated regions (3′UTR). The miRNA arrays were used to further identify potential miR-SNPs. The miR-SNP rs3735590 at the paraoxonase 1 (PON1) gene was finally selected and its associations with stroke and carotid IMT were evaluated. The 3′UTR reporter and SNP functional assays were then performed to validate the results. Results Compared with CC genotype, patients with CT or TT genotype at rs3735590 had lower risk of ischemic stroke (OR = 0.72, p = 0.036; OR = 0.83, p = 0.077, respectively). Among the healthy participants, the CT or TT genotype was associated with thinner IMT in the internal carotid arteries in comparison with CC genotype ( β = −0.76, p = 0.003; β = −0.022, p = 0.452, respectively). Our findings suggested that the minor allele T had a protective effect on atherosclerosis. Results from 3′UTR reporter assays showed that PON1 is a direct target gene of miR-616. In plasmid constructs carrying the risk allele C at rs3735590, miR-616 inhibited the genetic expression of PON1. However, substitution of C by T at rs3735590 reduced the miR-616 binding affinity, leading to overexpression of the PON1 gene. Conclusion Our study is the first to show that the miR-SNP at PON1 could affect genetic expression and is associated with an elevated risk for ischemic stroke and subclinical atherosclerosis.

Hepatitis C virus (HCV) can replicate in peripheral blood mononuclear cells (PBMCs), which can produce interferon to defend against virus infection. We hypothesized that dynamic gene expression in PBMCs might impact the treatment efficacy of peginterferon/ribavirin in HCV patients. PBMCs were collected at baseline, 1st week and 4th week of treatment from 27 chronic HCV-1 patients with 48-week peginterferon/ribavirin therapy (screening dataset n = 7; validation dataset n = 20). A sustained virologic response (SVR) was defined as undetectable HCV RNA throughout the 24 weeks after end-of-treatment. A complete early virologic response (cEVR) was defined as negative HCV RNA at treatment week 12. Forty-three differentially expressed genes identified by Affymetrix microarray were validated by quantitative polymerase chain reaction. Thirteen genes at week 1 and 24 genes at week 4 were upregulated in the SVR group compared with the non-SVR group. We selected 8 target genes (RSAD2, LOC26010, HERC5, HERC6, IFI44, SERPING1, IFITM3, and DDX60) at week 1 as the major components of the predictive model. This predictive model reliably stratified the responders and non-responders at week 1 (AUC = 0.89, p = 0.007 for SVR; AUC = 0.95, p = 0.003 for cEVR), especially among patients carrying the IL28B rs8099917 TT genotype (AUC = 0.89, p = 0.02 for SVR; AUC = 1.0, p = 0.008 for cEVR). The performance of this predictive model was superior to traditional predictors, including the rapid virologic response, viral load and IL28B genotype.

Histone deacetylase 8 (HDAC8), a unique member of class I histone deacetylases, shows remarkable correlation with advanced disease stage and multiple malignant tumors However, little is known about the contribution of HDAC8 to the tumorigenesis of hepatocellular carcinoma (HCC). The present study investigated the expression of HDAC8 regulated by the aryl hydrocarbon receptor (AHR) in HCC cell lines and tissues, and the roles of HDAC8 overexpression in cell proliferation, including potentially underlying mechanisms. We assessed the correlation between the clinic-pathological parameters and the expression of AHR and HDAC8. Further, we analyzed the AHR siRNA transfection and HDAC8 inhibitors to explore the functions of HDAC8 in HCC progression in vitro and in vivo. In a panel of 289 HCC patients, HDAC8 was shown to be highly correlated with AHR expression at both mRNA and protein levels. HCC patients with high AHR expression showed a shorter survival time than that with low AHR expression. We then found that the expression of both AHR and HDAC8 was significantly upregulated in both HCC cell lines and tumor tissues compared to human normal hepatocytes and matched non-tumor tissues. Furthermore, HDAC8 inhibition remarkably inhibited hepatoma cell proliferation and transformation activity via upregulation of RB1 in vitro and in vivo. Our data revealed an important role of the AHR-HDAC8 axis in promoting HCC tumorigenesis, thus identifying HDAC8 as a potential therapeutic target for HCC treatment.

Background Gallstone is a complex disease caused by multiple environmental and genetic factors. One of these is mucin glycoproteins. This case–control study aimed to investigate the association between single nucleotide polymorphisms (SNPs) at the MUC1-4 genes and gallstone. Methods The study included 475 cases and 941 controls. Eight tagging SNPs were selected: one at MUC1, two at MUC2, and five at MUC4. There was no available tagging SNP at MUC3. Genetic effects were initially evaluated by multivariate logistic regression. The combined effects from multiple genes were further evaluated, as well as the sex-specific effect. Permutation was used to correct for multiple testing. Results The genotypes were all in Hardy–Weinberg equilibrium. SNP rs7396030 at MUC2 yielded a p value of 0.03. Further sex-specific analysis showed significance solely with male subjects ( p = 0.005). Similarly, SNP rs4072037 at MUC1 was only significant ( p = 0.035) in males. The permutation empirical p values were 0.005 for rs7396030 and 0.02 for rs4072037. For males, the combined genetic effect yielded an OR of 4.68 ( p = 0.0008). Conclusions The SNPs at MUC1 and MUC2 are significantly associated with gallstone in men but not in women. These genes can work jointly to further increase susceptibility to gallstone in a Chinese population.

To identify candidate genes in relation to plasma lipid levels in Caribbean Hispanics.A total of 114 single nucleotide polymorphisms (SNPs) at 17 lipid-related genes were genotyped in 477 Caribbean Hispanics from the Northern Manhattan Study (NOMAS). Analyses for each SNP and haplotype were performed to evaluate the associations with four lipid traits: high- and low-density lipoprotein cholesterol (HDL-C, LDL-C), triglyceride (TG) and total cholesterol (TC).We identified 19 SNPs at 10 genes that were significantly related to lipids (p0.01), including nine involved in the reverse cholesterol transport pathway, and one involved in bile acid synthesis. Three genes, namely the apolipoprotein A5, apolipoprotein B and cytochrome p450 polypeptide 7A1 genes, accounted for the largest proportion of variation in HDL-C/TG, TC and LDL-C respectively.The cumulative effects of multiple genetic variants led to a substantially better prediction of inter-individual variations in lipid levels.

// Shen-Nien Wang 1, 2, * , Li-Ting Wang 2, * , Ding-Ping Sun 3, 4, * , Chee-Yin Chai 5 , Edward Hsi 6 , Hsing-Tao Kuo 7, 8 , Kazunari K. Yokoyama 2, 9, 10, 11, 12, 13 , Shih-Hsien Hsu 2, 13 1 Division of Hepatobiliary Surgery, Department of Surgery, Faculty of Medicine, Kaohsiung Medical University Hospital, Kaohsiung, Taiwan 2 Graduate Institute of Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan 3 Division of General Surgery, Department of Surgery, Chi-Mei Medical Center, Tainan, Taiwan 4 Department of Food Science and Technology, Chia Nan University of Pharmacy and Science, Tainan, Taiwan 5 Department of Pathology, Faculty of Medicine, College of Medicine, Kaohsiung, Taiwan 6 Department of Genome Medicine, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan 7 Department of Internal Medicine, Division of Hepatogastroenterology, Chi-Mei Medical Center, Tainan, Taiwan 8 Department of Senior Citizen Service Management, Chia Nan University of Pharmacy & Science, Tainan, Taiwan 9 Research Center for Stem Cell Research, Kaohsiung Medical University, Kaohsiung, Taiwan 10 Center for Environmental Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan 11 Department of Molecular Preventive Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan 12 Faculty of Science and Engineering, Tokushima Bunri University, Sanuki, Japan 13 Center of Infectious Disease and Cancer Research, Kaohsiung Medical University, Kaohsiung, Taiwan * These authors have contributed equally to this work Correspondence to: Shih-Hsien Hsu, e-mail: jackhsu@kmu.edu.tw Kazunari K. Yokoyama, e-mail: kazu@kmu.edu.tw Hsing-Tao Kuo, e-mail: kuohsu2003@yahoo.com.tw Keywords: cyclin D1 , DP1, E2F1 , hepatocellular carcinoma (HCC), ISX Received: October 05, 2015 Accepted: April 16, 2016 Published: May 09, 2016 ABSTRACT Intestine-specific homeobox ( ISX ), a newly identified proto-oncogene, is involved in cell proliferation and progression of hepatocellular carcinoma (HCC). However, the underlying mechanisms linking gene expression and tumor formation remain unclear. In this study, we found that ISX transcriptionally activated E2F transcription factor 1 ( E2F1 ) and associated oncogenic activity by directly binding to the E2 site of its promoter. Forced expression of ISX increased the expression of and phosphorylated the serine residue at position 332 of E2F1 , which may be translocated into the nucleus to form the E2F1 –DP-1 complex, suggesting that the promotion of oncogenic activities of the ISX–E2F1 axis plays a critical role in hepatoma cells. Coexpression of ISX and E2F1 significantly promoted p53 and RB-mediated cell proliferation and anti-apoptosis, and repressed apoptosis and autophagy. In contrast, short hairpin RNAi-mediated attenuation of ISX and E2F1 decreased cell proliferation and malignant transformation, respectively, in hepatoma cells in vitro and in vivo . The mRNA expression of E2F1 and ISX in 238 paired specimens from human HCC patients, and the adjacent, normal tissues exhibited a tumor-specific expression pattern which was highly correlated with disease pathogenesis, patient survival time, progression stage, and poor prognosis. Therefore, our results indicate that E2F1 is an important downstream gene of ISX in hepatoma progression.

The phosphodiesterase 4D (PDE4D) gene has been reported as a risk gene for ischemic stroke. The vascular factors are between the hypothesized etiologies of sudden sensorineural hearing loss (SSNHL), and this genetic effect might be attributed for its role in SSNHL. We hypothesized that genetic variants of the PDE4D gene are associated with susceptibility to SSNHL. We conducted a case-control study with 362 SSNHL cases and 209 controls. Three single nucleotide polymorphisms (SNPs) were selected. The genotypes were determined using TaqMan technology. Hardy-Weinberg equilibrium (HWE) was tested for each SNP, and genetic effects were evaluated according to three inheritance modes. We carried out sex-specific analysis to analyze the overall data. All three SNPs were in HWE. When subjects were stratified by sex, the genetic effect was only evident in females but not in males. The TT genotype of rs702553 exhibited an adjusted odds ratio (OR) of 3.83 (95 % confidence interval = 1.46-11.18) (p = 0.006) in female SSNHL. The TT genotype of SNP rs702553 was associated with female SSNHL under the recessive model (p = 0.004, OR 3.70). In multivariate logistic regression analysis, TT genotype of rs702553 was significantly associated with female SSNHL (p = 0.0043, OR 3.70). These results suggest that PDE4D gene polymorphisms influence the susceptibility for the development of SSNHL in the southern Taiwanese female population.

Background Estrogen is involved in neuron plasticity and can promote neuronal survival in stroke. Its actions are mostly exerted via estrogen receptor alpha (ERα). Previous animal studies have shown that ERα is upregulated by DNA demethylation following ischemic injury. This study investigated the methylation levels in the ERα promoter in the peripheral blood of ischemic stroke patients. Methods The study included 201 ischemic stroke patients, and 217 age- and sex-comparable healthy controls. The quantitative methylation level in the 14 CpG sites of the ERα promoter was measured by pyrosequencing in each participant. Multivariate regression model was used to adjust for stroke traditional risk factors. Stroke subtypes and sex-specific analysis were also conducted. Results The results demonstrated that the stroke cases had a lower ERα methylation level than controls in all 14 CpG sites, and site13 and site14 had significant adjusted p-values of 0.035 and 0.026, respectively. Stroke subtypes analysis showed that large-artery atherosclerosis and cardio-embolic subtypes had significantly lower methylation levels than the healthy controls at CpG site5, site9, site12, site13 and site14 with adjusted p = 0.039, 0.009, 0.025, 0.046 and 0.027 respectively. However, the methylation level for the patients with small vessel subtype was not significant. We combined the methylation data from the above five sites for further sex-specific analysis. The results showed that the significant association only existed in women (adjusted p = 0.011), but not in men (adjusted p = 0.300). Conclusions Female stroke cases have lower ERα methylation levels than those in the controls, especially in large-artery and cardio-embolic stroke subtypes. The study implies that women suffering from ischemic stroke of specific subtype may undergo different protective mechanisms to reduce the brain injury.

The present study aims to discover single nucleotide polymorphisms (SNPs) at the apelin gene (APLN) in relation to arterial stiffness, and to explore its molecular mechanisms. A two-step genetic association study was conducted using 799 and 937 subjects in the screening and validation data, respectively. Four tagging SNPs of APLN were tested. SNP rs3115757 was significantly associated with stiffness parameters (β, Ep and PWV) in women, but not in men. The function of rs3115757 was tagged by rs3115758 which is located in miR-765 binding site in the 3′ untranslated region of APLN. The reporter assay confirmed that different alleles of rs3115758 interfered with miR-765 binding and then modified APLN expression. Over-expression of miR-765 in endothelial cells decreased mRNA and protein levels of APLN, which further inhibited the phosphorylation of eNOS and ERK/Akt/AMPK signaling. Collectively, our data showed that rs3115758 accounts for the susceptibility of arterial stiffening through miR-765-induced APLN repression.

Background.The interval between intra-arterial infusion chemotherapy (IAIC) and surgery was investigated in terms of its effects on survival in patients with locally advanced oral squamous cell carcinoma (OSCC).Methods.This retrospective study analyzed 126 patients who had completed treatment modalities for stage IV OSCC. All patients were followed up for 3 years. Kaplan-Meier and Cox regression methods were used to determine how survival was affected by general factors, primary tumor volume, TNM stage, and duration of neoadjuvant chemotherapy.Results.In 126 patients treated for locally advanced OSCC by preoperative induction IAIC using methotrexate, multivariate analysis of relevant prognostic factors showed that an IAIC duration longer than 90 days was significantly associated with poor prognosis (hazard ratio, 1.77;P=0.0259).Conclusions.Duration of IAIC is a critical factor in the effectiveness of multimodal treatment for locally advanced OSCC. Limiting the induction course to 90 days improves overall survival.

To evaluate the expression and prognostic value of two autophagy-related (Atg) proteins, Beclin-1 and Atg5, in human oral squamous cell carcinoma (OSCC) and to correlate findings with clinical outcomes.Immunohistochemistry for Beclin-1 and Atg5 was assessed in tumor specimens from 90 patients with OSCC. Immunopositivity was semi-quantitatively scored and receiver-operating characteristic curve analysis was used to determine the cut-off positivity score.55 (61.1%) and 52 (57.8%) cases showed positive Beclin-1 and Atg5 staining, respectively. 40 tumors (44.4%) were positive for both Beclin-1 and Atg5 expression and 23 cases (25.6%) showed absence of both proteins. Beclin-1 expression significantly correlated with tumor grade (p=0.008) and lymph node metastasis (p=0.009). The expression of Atg5 was associated with tumor grade (p=0.016), advanced clinical stage (p0.001), large tumor size (p=0.002), and lymph node metastasis (p0.001). A significant difference in 3-year OS (p=0.050) and TTR (p=0.049) between the patients with Beclin-1 expression and those not showing Beclin-1 expression was found whereas the difference did not reach a statistical significance for Atg5 expression. 3-year OS and TTR differed significantly between patients with dual expression and those with double-negative expression (p=0.022 and p=0.026, respectively).Dual expression of tumor Beclin-1 and Atg5 expression may be an adverse prognostic indicator for OSCC.

Backgrounds Chronic hepatitis C virus (HCV) infection has been associated with induction of microRNAs (miRNAs) in peripheral blood mononuclear cells (PBMC). We aimed to evaluate the role of PBMC–miRNAs in the treatment outcome to antiviral therapy for HCV genotype 1 (HCV-1) patients. Methods Treatment-naive chronic HCV-1 patients, including 13 in screening phase and 48 in validation phase, were treated with 48 weeks of peginterferon/ribavirin. The primary end-point was the achievement of a sustained virological response (SVR, HCV RNA undetectable during 24 weeks post-treatment follow-up). Expression profiling of PBMC–miRNAs was performed by quantitative PCR-based array in typical responders and null-responders. Then candidate PBMC–miRNAs were validated by quantitative PCR in an independent validation set. Results PBMC–miR-125b was significantly predictive of an SVR, with expression levels of 5.28-fold lower in sustained responders versus null-responders (p = 0.0163). In multivariate analysis, PBMC–miR-125b was significantly associated with the achievement of SVR (per 2-fold decrease, odds ratio/95% confidence interval (OR/CI): 2.07/1.14-6.31) independent of sex, age and interleukin-28B genotype. In patients who did not achieve a rapid virological response (RVR, undetectable HCV RNA at treatment week 4), PBMC–miR-125b was the only predictive factor of an SVR (per 2-fold decrease, OR/CI: 2.07/1.14-6.31). However, the circulating and hepatic miR-125b did not show significant difference between responders and non-responders. Conclusions PBMC–miR-125b expression levels were inversely related to the achievement of an SVR in HCV-1 patients, independent of interleukin-28B genotype, and was the single predictor of SVR in non-RVR patients.

Purpose We studied the relationship between high myopia and the MMP3 and TIMP1 genes. Design Case-control study. Methods We enrolled 150 high myopic individuals (≤ −6 diopters) and 262 controls (≥ −1.5 diopters) initially, and another 216 cases and 474 controls were enrolled for genotyping promising polymorphisms for replication. Thirteen polymorphisms were genotyped in the initial data. Logistic regression was used to test for genetic effects. Subset analyses were performed according to the education level and family history. Results There was no significant association between any polymorphism and high myopia in the initial data. Among the highly educated subjects, the 5A/6A polymorphism at the MMP3 gene suggested a potential relationship with high myopia, and it was genotyped in the follow-up samples. However, this polymorphism failed to show any significant results in the overall subjects. Conclusions The two genes may not play a crucial role for high myopia in young Taiwanese men.

ATG9A is an integral membrane protein required for autophagosome formation and a membrane carrier in the autophagy pathways. The present study was designed to investigate the expression of ATG9A in oral squamous cell carcinoma (OSCC). Clinically annotated tumor specimens from 90 patients with OSCC were subjected to immunohistochemistry using an antibody against ATG9A and immunoreactivity was scored using an immunoreactivity score (IRS). Scores were compared with clinical and pathologic data to assess association with outcome. Overexpression of ATG9A was defined as an IRS of ≥9 by receiver operating characteristics curve analysis and was identified in 25 (28 %) of 90 cases. ATG9A overexpression was associated with disease recurrence and overall survival (OS) in both univariate (p = 0.030 and 0.025, respectively) and multivariate (p = 0.026 and 0.038, respectively) Cox analyses. Kaplan–Meier plots also showed that patients with ATG9A overexpression had shorter 3-year OS (p = 0.017) and time to recurrence (p = 0.021) than those with low ATG9A expression. These results suggest that the presence of ATG9A in the cytoplasm of tumor cells may be an independent biomarker for disease recurrence and survival in patients with OSCC.

Immune dysregulation may be involved in the development of endometriosis. The anti-inflammatory cytokine IL-10 plays an important role in eliminating unwanted cells and cellular debris in a silent way. We investigated the modulatory role of IL-10 in the development of endometriosis. We observed that the serum level of IL-10 in patients with endometriosis was significantly higher than that in healthy subjects or in control subjects with other gynecological disease. Monocyte-derived dendritic cells acquired from male donors and subsequently conditioned with serum from women with endometriosis exhibited a tolerogenic phenotype, including increased IL-10 production, lower IL-12 secretion, and down-regulation of CD86 and HLA-DR molecules. Depletion of IL-10 activity in a C57BL/6 mouse model of surgically induced endometriosis significantly decreased the size of endometrial lesions. In contrast, IL-10 administration promoted the growth of endometrial lesions in this model. In addition, infiltrated plasmacytoid dendritic cells were the primary IL-10–secreting immune cells in endometrial lesions. Our findings suggest that IL-10 may suppress immunity against endometrial implants, contributing to development of endometriosis.

Purpose Fibroblast growth factor-2 (FGF2) has been implied in the development of myopia according to previous studies investigating FGF2 in the sclera and retinal pigment epithelium. This study measured retinal FGF2 gene expression in an animal model and also tested for the association between single nucleotide polymorphisms (SNPs) in FGF2 and high myopia. Methods The guinea pigs were assigned to 2 groups: form deprivation myopia (FDM) for two weeks and normal control (free of form deprivation). Biometric measurement was performed and FGF2 expression levels were compared among the FDM eyes, the fellow eyes of the FDM group and the normal eyes in retina. We also enrolled 1,064 cases (≤-6.0 D) and 1,001 controls (≥-1.5 D) from a Chinese population residing in Taiwan. Six tagging SNPs were genotyped to test for an association between genotypes and high myopia. Results The FDM eyes had the most prominent changes of refraction and axial length. Compared with the mRNA levels of FGF2 in the normal eyes, the FDM eyes had the highest levels of mRNA (p=0.0004) followed by the fellow eyes (p=0.002). The FDM and normal eyes became more myopic compared with the fellow eyes, but the fellow eyes became more hyperopic (p=0.004) in the end of the experiment which may be due to its relatively short axial length when compared with normal eyes (p=0.05). The SNP genotypes were all in Hardy–Weinberg equilibrium. However, none of the SNPs were significantly associated with high myopia (all p values >0.1). Conclusions We identified a significant change of FGF2 expression in the FDM eyes but FGF2 genetic variants are unlikely to influence susceptibility to myopia. There may be a systemic effect to influence gene expression and refraction on the fellow eyes, which may perturb emmetropization in the fellow eyes. Our data also suggest using normal eyes rather than the fellow eyes as the control eyes when study the form deprivation myopia.

Background: Genetic variations on chromosome 9p21 have been found to be associated with peripheral artery disease (PAD), but have not been investigated in Asians. The ankle-brachial index (ABI) is a widely-used measurement for PAD. We previously reported the BRAP gene is associated with the ABI, so the aim of the present study was to test whether the ANRIL gene on 9p21 is associated with the ABI and to test an interaction between BRAP and ANRIL in a Chinese population. Methods and Results: A total of 745 subjects with a family history of myocardial infarction or stroke were enrolled. The multiplicative and additive effects of 2 significant single-nucleotide polymorphisms (SNPs) were evaluated by multivariable regression analysis. SNP rs2383207 on ANRIL was most significantly associated with lower ABI. Similar to our previous findings, SNP rs11066001 on BRAP was associated with lower ABI. A dose-response relationship between ABI values and the number of risk alleles from the 2 significant SNPs was observed in both men and women (adjusted P=0.004 for men, 0.008 for women). The combined genetic effect on ABI was stronger in smokers than in non-smokers. Conclusions:ANRIL on 9p21 and BRAP were both associated with ABI in a Taiwanese population. An additive effect between variants of these 2 genes was found. The finding of a potential gene-gene interaction and gene-environment interaction is interesting, but needs further investigation. (Circ J 2012; 76: 446-452)

Aim Matrix metalloproteinase (MMP)-8 is a protease that degrades numerous extracellular molecules and has been implicated in the pathogenesis of periodontitis. Polymorphism in the MMP-8 could affect the susceptibility to disease. Our aim was to evaluate the association between periodontitis and MMP-8 -799 C>T polymorphism. Material and methods Genomic DNA was obtained from 361 chronic periodontitis patients (CP), 96 aggressive periodontitis patients (AgP), and 106 periodontally healthy controls (HC). MMP-8 -799 C>T polymorphism was determined using the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Results The frequencies of genotypes in diseased groups were similar but were significantly different from those in the HC. Multivariate logistic regression analysis with adjustment for age, gender and smoking indicated that increased risks of AgP and CP were associated with the -799 T allele (in AgP, adjusted OR = 1.99, p = 0.04; in CP, adjusted OR = 1.87, p = 0.007). To avoid the confounded effect of smoking on MMP-8 polymorphism to periodontitis, the analysis was conducted on non-smokers and the associations were significant. Conclusions These results suggested that non-smoking Taiwanese with the MMP-8 -799 T allele were associated with the risks of both CP and AgP. Further studies in other ethnic populations are necessary.

Both interleukin-28B genetic variants and on-treatment virological responses are factors predictive of treatment outcome in hepatitis C virus genotype 1 (HCV-1) patients. We aimed to compare the clinical significance of the two factors.Rs8099917 genotype and on-treatment responses were determined in 182 HCV-1 patients with 48-week peginterferon/ribavirin.Comparing to patients with rs8099917 TG/GG genotype, those with TT genotype had significantly higher rapid virological response (RVR, 46.2% vs. 19.2%, P=0.01) and sustained virological response (SVR, 85.3% vs. 42.3%, P0.001) rates. Logistic regression analysis revealed that the strongest factor predictive of a RVR was the carriage of rs8099917 TT genotype (odds ratio/95% confidence intervals [OR/CI]: 4.25/1.39-13.01). The most important factor predictive of an SVR was the attainment of a RVR (OR/CI: 57.22/6.23-525.37), followed by the carriage of rs8099917 TT genotype (OR/CI: 10.06/3.12-32.44). However, while on-treatment factors were taken into account, the cEVR was the most important determinant to an SVR (OR/CI:54.98/9.07-333.38), whereas the influence of rs8099917 genotype became non-significant in non-RVR patients.Rs8099917 TT genotype is significantly independently predictive of on-treatment virological responses, which were the major determinants of an SVR, in Asian HCV-1 patients.

Background Gallstone disease (GSD) is a common disease that can be caused by environmental influences, common genetic factors and their interactions. Mucin glycoproteins may be one important factor for GSD. We conducted a case–control study to investigate the relationship between the mucin-like protocadherin (MUPCDH) gene polymorphisms and GSD. Methods The study included 452 GSD cases and 491 healthy controls who had no evidence of gallstones by ultrasound examination. Two common tagging single nucleotide polymorphism (SNP) rs3758650 and rs7932167, and four non-synonymous SNPs rs34362213, rs2740375, rs7108757 and rs2740379 were genotyped. The genetic effects were evaluated using the multivariate regression model. Results The genotypes of these SNPs were all in Hardy–Weinberg equilibrium. Three non-synonymous SNPs (rs34362213, rs7108757 and rs2740379) were monomorphic. The single SNP analysis showed two SNPs (rs7932167 and rs2740375) were not associated with GSD and only SNP rs3758650 had the association of the presence of GSD with an odds ratio (OR) of 1.59 (adjusted P = 0.013) for the AG genotype and 5.82 (adjusted P = 0.007) for the AA genotype when compared with the reference GG genotype. The haplotype analysis of the three polymorphic SNPs showed GCA was significant for GSD (adjusted p = 0.001) with an odds ratio (OR) of 1.41 when compared to other haplotypes. Conclusions The MUPCDH genetic polymorphism rs3758650 was considered a genetic marker to predict symptomatic GSD subjects. It may be of importance for GSD patients with the risk SNPs to be frequently checked because they may develop symptomatic GSD.

Coxsackievirus A4 outbreaks occurred in Taiwan in 2004 and 2006. The spatiotemporal transmission of this error-prone RNA virus involves a continuous interaction between rapid sequence variation and natural selection. To elucidate the molecular characteristics of CV-A4 and the spatiotemporal dynamic changes in CV-A4 transmission, worldwide sequences of the 3' VP1 region (420 nt) obtained from GenBank were analyzed together with sequences isolated in Taiwan from 2002 to 2009. Sequences were characterized in terms of recombination, variability, and selection. Phylogenetic trees were constructed using neighbor-joining, maximum likelihood and Monte Carlo Markov Chain methods. Spatiotemporal dynamics of CV-A4 transmission were further estimated by a Bayesian statistical inference framework. No recombination was detected in the 420 nt region. The estimated evolution rate of CV-A4 was 8.65 × 10(-3) substitutions/site/year, and a purifying selection (d(N)/d(S)=0.032) was noted over the 3' VP1 region. All trees had similar topology: two genotypes (GI and GII), each including two subgenotypes (A and B), with the prototype and a Kenyan strain in separate branches. The results revealed that the virus first appeared in USA in 1950. Since 1998, it has evolved into the Kenya, GI-A (Asia) and GII-A (Asia and Europe) strains. Since 2004, GI-B and GII-B have evolved continuously and have remained prevalent. The co-existence of several positive selection lineages of GI-B in 2006 indicates that the subgenotype might have survived lineage extinction. This study revealed rapid lineage turnover of CV-A4 and the replacement of previously circulating strains by a new dominant variant. Therefore, continuous surveillance for further CV-A4 transmission is essential.

We previously showed an association between the exon1 +49 A/G single nucleotide polymorphism (SNP) and the relapse of Graves' disease (GD). The G allele was associated with early relapse.In this follow-up study, we sought to replicate the result by genotyping nine additional polymorphisms and recruiting another 60 GD patients.The GD patients were divided into three groups: recurred within 9 months, between 10-36 months, and more than 36 months. There were 65 patients with early recurrence, 55 with medium recurrence, and 88 with late recurrence. Although several SNPs were associated with recurrence, the most significant marker was still exon1 +49 A/G. Separate analysis of the genotypes for the 60 newly enrolled patients indicated that our present study was not biased by the previous samples. Once exon1 +49 A/G was included in the model to predict recurrence, other markers would not add more predictive information. Haplotype analysis did not show an additional value once exon1 +49 A/G was compulsorily included.Multivariate logistic regression analysis showed that GG genotype of exon1 +49 A/G SNP had an adjusted odds ratio of 2.2 (95% confidence interval, 1.1-4.4) compared with the combined group of GA plus AA. Other significant predictors were large goiter size at the end of the treatment and positive TSH-binding inhibitory Ig at the end of the treatment.This follow-up study confirms the usefulness of the exon1 +49 A/G SNP of the cytotoxic T lymphocyte-associated molecule-4 gene in predicting recurrence after cessation of treatment. There is no additional power by including other polymorphisms to predict recurrence.

Ankylosing spondylitis (AS) is a chronic inflammation of the sacroiliac joints, spine and peripheral joints. The aetiology of ankylosing spondylitis is still unclear. Previous studies have indicated that genetics factors such as human leukocyte antigen HLA-B27 associates to AS susceptibility. We carried out a case-control study to determine whether the genetic polymorphisms of ORAI1 gene, a major component of store-operated calcium channels that involved the regulation of immune system, is a susceptibility factor to AS in a Taiwanese population. We enrolled 361 AS patients fulfilled the modified New York criteria and 379 controls from community. Five tagging single nucleotides polymorphisms (tSNPs) at ORAI1 were selected from the data of Han Chinese population in HapMap project. Clinical statuses of AS were assessed by the Bath Ankylosing Spondylitis Disease Activity Index (BASDAI), Bath Ankylosing Spondylitis Functional Index (BASFI), and Bath Ankylosing Spondylitis Global Index (BAS-G). Our results indicated that subjects carrying the minor allele homozygote (CC) of the promoter SNP rs12313273 or TT homozygote of the SNP rs7135617 had an increased risk of HLA-B27 positive AS. The minor allele C of 3′UTR SNP rs712853 exerted a protective effect to HLA-B27 positive AS. Furthermore, the rs12313273/rs7135617 pairwise allele analysis found that C-G (OR 1.69, 95% CI 1.27, 2.25; p = 0.0003) and T-T (OR 1.75, 95% CI 1.36, 2.27; p

Kawasaki disease (KD) is characterized by systemic vasculitis with unknown etiology. Previous studies from Japan indicated that a gene polymorphism of ITPKC (rs28493229) is responsible for susceptibility to KD. We collected DNA samples from 1,531 Taiwanese subjects (341 KD patients and 1,190 controls) for genotyping ITPKC. In this study, no significant association was noted for the ITPKC polymorphism (rs28493229) between the controls and KD patients, although the CC genotype was overrepresented. We further combined our data with previously published case/control KD studies in the Taiwanese population and performed a meta-analysis. A significant association between rs28493229 and KD was found (Odds Ratio:1.36, 95% Confidence Interval 1.12-1.66). Importantly, a significant association was obtained between rs28493229 and KD patients with aneurysm formation (P = 0.001, under the recessive model). Taken together, our results indicated that C-allele of ITPKC SNP rs28493229 is associated with the susceptibility and aneurysm formation in KD patients in a Taiwanese population.