Genome-Wide Analysis of Attention Deficit Hyperactivity Disorder in Norway

Background

Attention deficit hyperactivity disorder (ADHD) is a highly heritable neuropsychiatric condition, but it has been difficult to identify genes underlying this disorder. This study aimed to explore genetics of ADHD in an ethnically homogeneous Norwegian population by means of a genome-wide association (GWA) analysis followed by examination of candidate loci.

Materials and Methods

Participants were recruited through Norwegian medical and birth registries as well as the general population. Presence of ADHD was defined according to DSM-IV criteria. Genotyping was performed using Illumina Human OmniExpress-12v1 microarrays. Statistical analyses were divided into several steps: (1) genome-wide association in the form of logistic regression in PLINK and follow-up pathway analyses performed in DAPPLE and INRICH softwares, (2) SNP-heritability calculated using genome-wide complex trait analysis (GCTA) tool, (3) gene-based association tests carried out in JAG software, and (4) evaluation of previously reported genome-wide signals and candidate genes of ADHD.

Results

In total, 1.358 individuals (478 cases and 880 controls) and 598.384 autosomal SNPs were subjected to GWA analysis. No single polymorphism reached genome-wide significance. The strongest signal was observed at rs9949006 in the ENSG00000263745 gene (OR=1.51, 95% CI 1.28–1.79, p=1.38E-06). Pathway analyses of the top SNPs implicated genes involved in the regulation of gene expression, cell adhesion and inflammation. Among previously identified ADHD candidate genes, prominent association signals were observed for SLC9A9 (rs1393072, OR=1.46, 95% CI = 1.21–1.77, p=9.95E-05) and TPH2 (rs17110690, OR = 1.38, 95% CI = 1.14–1.66, p=8.31E-04).

Conclusion

This study confirms the complexity and heterogeneity of ADHD etiology. Taken together with previous findings, our results point to a spectrum of biological mechanisms underlying the symptoms of ADHD, providing targets for further genetic exploration of this complex disorder.     

Association Study of Gene LPP in Women with Polycystic Ovary Syndrome

Background

Previous genome-wide association study (GWAS) of polycystic ovary syndrome (PCOS) in Han Chinese population has found that SNPs in LPP gene were nominally significant in PCOS patients (P around 10E-05). Replication of the GWAS was applied to further confirm the relationship between LPP gene and PCOS.

Methods

Three polymorphisms of LPP gene (rs715790, rs4449306, rs6782041) were selected and replicated in additional 1132 PCOS cases and 1142 controls. Genotyping of LPP gene was carried out by Taqman-MGB method.

Results

In rs715790, the allele frequency is significantly different between the PCOS group and the control group. Meta-analysis showed that the allele frequencies of the three SNPs rs715790 (P_meta = 1.89E-05, OR = 1.23), rs4449306 (P_meta = 3.0E-04, OR = 1.10), rs6782041 (P_meta = 2.0E-04, OR = 1.09), were significantly different between PCOS cases and controls.

Conclusions

Our results suggest that LPP gene might be a novel candidate for PCOS.     

A genome-wide association study for racing performances in Thoroughbreds clarifies a candidate region near the MSTN gene

Using 1400 microsatellites, a genome-wide association study (GWAS) was performed to identify genomic regions associated with lifetime earnings and performance ranks, as determined by the Japan Racing Association (JRA). The minimum heritability (h(2) ) was estimated at 7-8% based on the quantitative trait model, suggesting that the racing performance is heritable. Following GWAS with microsatellites, fine mapping led to identification of three SNPs on ECA18, namely, g.65809482T>C (P=1.05E-18), g.65868604G>T (P=6.47E-17), and g.66539967A>G (P=3.35E-14) associated with these performance measures. The haplotype of these SNPs, together with a recently published nearby SNP, g.66493737C>T (P=9.06E-16) in strong linkage disequilibrium, also showed a very clear association with the performance (P<1E-05). The candidate genomic region contained eight genes annotated by ENSEMBL, including the myostatin gene (MSTN). These findings suggest the presence of a gene affecting the racing performance in Thoroughbred racehorses in this region on ECA18.     

Gene-based analysis of regionally enriched cortical genes in GWAS data sets of cognitive traits and psychiatric disorders

Background

Despite its estimated high heritability, the genetic architecture leading to differences in cognitive performance remains poorly understood. Different cortical regions play important roles in normal cognitive functioning and impairment. Recently, we reported on sets of regionally enriched genes in three different cortical areas (frontomedial, temporal and occipital cortices) of the adult rat brain. It has been suggested that genes preferentially, or specifically, expressed in one region or organ reflect functional specialisation. Employing a gene-based approach to the analysis, we used the regionally enriched cortical genes to mine a genome-wide association study (GWAS) of the Norwegian Cognitive NeuroGenetics (NCNG) sample of healthy adults for association to nine psychometric tests measures. In addition, we explored GWAS data sets for the serious psychiatric disorders schizophrenia (SCZ) (n = 3 samples) and bipolar affective disorder (BP) (n = 3 samples), to which cognitive impairment is linked.

Principal Findings

At the single gene level, the temporal cortex enriched gene RAR-related orphan receptor B (RORB) showed the strongest overall association, namely to a test of verbal intelligence (Vocabulary, P = 7.7E-04). We also applied gene set enrichment analysis (GSEA) to test the candidate genes, as gene sets, for enrichment of association signal in the NCNG GWAS and in GWASs of BP and of SCZ. We found that genes differentially expressed in the temporal cortex showed a significant enrichment of association signal in a test measure of non-verbal intelligence (Reasoning) in the NCNG sample.

Conclusion

Our gene-based approach suggests that RORB could be involved in verbal intelligence differences, while the genes enriched in the temporal cortex might be important to intellectual functions as measured by a test of reasoning in the healthy population. These findings warrant further replication in independent samples on cognitive traits.     

Pathway analysis of genome-wide association study for bone mineral density

The aim of this study was to identify the candidate causal single nucleotide polymorphisms (SNPs) and candidate causal mechanisms that contribute to bone mineral density (BMD) and to generate a SNP to gene to pathway hypothesis using an analytical pathway-based approach. We used hip BMD GWAS data of the genotypes of 301,019 SNPs in 5,715 Europeans. ICSNPathway (identify candidate causal SNPs and pathways) analysis was applied to the BMD GWAS dataset. The first stage involved the pre-selection of candidate causal SNPs by linkage disequilibrium analysis and the functional SNP annotation of the most significant SNPs found. The second stage involved the annotation of biological mechanisms for the pre-selected candidate causal SNPs using improved-gene set enrichment analysis. ICSNPathway analysis identified seven candidate SNPs, eight candidate pathways, and seven hypothetical biological mechanisms. Eight pathways are as follows; gamma-hexachlorocyclohexane degradation (nominal p-value < 0.001, false discovery rate (FDR) <0.001), regulation of the smoothened signaling pathway (nominal p-value < 0.001, FDR = 0.016), TACI and BCMA stimulation of B cell immune response (nominal p-value < 0.001, FDR = 0.021), endonuclease activity (nominal p-value = 0.001, FDR = 0,026), regulation of defense response to virus (nominal p-value = 0.001, FDR = 0.028), serine_type_endopeptidase_inhibitor_activity (nominal p-value = 0.001, FDR = 0.044), endoribonuclease activity (nominal p-value = 0.002, FDR = 0.045), and myeloid leukocyte differentiation (nominal p-value = 0.001, FDR = 0.050). The most significant causal pathway was gamma-hexachlorocyclohexane degradation. CYP3A5, PON2, PON3, CMBL, PON1, ALPL, CYP3A43, CYP3A7, ACP6, ACPP, and ALPI (p < 0.05) are involved in the pathway of gamma-hexachlorocyclohexane degradation. Further examination of the gene contents revealed that DBR1, DICER1, EXO1, FEN1, POP1, POP4, RPP30, and RPP38 were involved in 2 of the 8 pathways (p < 0.05). By applying ICSNPathway analysis to BMD GWAS data, we identified seven candidate SNPs and eight pathways involving gamma-hexachlorocyclohexane degradation, which may contribute to low BMD.     

In Search for the Genetic Basis of Quality of Life in Healthy Swedish Women—A GWAS Study Using the iCOGS Custom Genotyping Array

Background

Quality of life (QoL) is increasingly measured in both research and clinical practice. QoL-assessments are built on a long, empirically-based, and stringent approach. There is ample evidence that QoL is, in part, heritable. We therefore performed a GWAS relating genetic variation to QoL in healthy females.

Methods

In 5,142 healthy females, background characteristics (e.g. demographic, clinical, lifestyle and psychological factors) and QoL by means of the EORTC QLQ-C30 were measured. Moreover, women were genotyped using a custom array including ~210,000 single nucleotide polymorphisms (SNPs). Initially, SNPs were related to each QoL-domain, by means of partially adjusted (controlling for age and population stratification) and fully adjusted (controlling for age, population stratification, and background characteristics) regression analyses. Additionally, gene-based analyses were performed relating the combined effect of SNPs within each gene to QoL using the statistical software package VEGAS.

Results

None of the associations between QoL and genetic variation (i.e. individual SNPs and genes) reached the bonferroni corrected significance level.

Conclusion

Reasons for a lack of association between genetic markers and QoL could be low variation in QoL-scores; selecting genetic markers not tagging QoL; or that the genetic effect that impacts one’s QoL is mediated through biological pathways rather than the effect of single SNPs or genes. Therefore, we opt for a pathway-based or system biology approach as a complementary and powerful approach to analyze the combined effect of genes and their biological implications in future studies focusing on QoL-issues.     

Identification of single nucleotide polymorphisms in the TNFRSF17 gene and their association with gastrointestinal disorders

TNFRSF17 is preferentially expressed in mature B lymphocytes, and may be important for the development of B cells. TNFRSF17 is selected as a candidate susceptibility gene to IBD pathogenesis by our cDNA microarray analysis, and we showed the specific expression of TNFRSF17 in resting and activated CD19⁺ cells obtained from human blood. We identified four SNPs (g-1729G>A, g.2295T>C, g.2445G>A and g.2493G>A) and one variation site (g.894delT) in the TNFRSF17 gene using direct sequencing analysis. In addition, the association of the genotype and allelic frequencies of these SNPs was studied in healthy controls and in patients with ulcerative colitis (UC) or irritable bowel syndrome (IBS). Although, the genotype and allelic frequencies of these SNPs, in the UC and IBS patients, were not significantly different from those in the healthy controls, the distribution of the AAG, GGA, AGG and AAA haplotypes, of the SNPs (g.-1729G>A, g.2445G> A and g.2493G>A) associated with the TNFRSF17 gene, in the UC patients, were notably different from those of the healthy controls (P = 0.002, 0.002, 4.7E-4 and 3.3E-6, respectively). Moreover, the frequencies of the AAG, AGG, GAG and GAA haplotypes were significantly different in the IBS patients compared to the healthy controls (P = 4.2E-5, 4.4E-17, 1.8E-22 and 1.6E-10, respectively). These results suggest that the haplotypes of the TNFRSF17 polymorphisms might be associated with UC and IBS susceptibility.     

Meta-analysis of two Chinese populations identifies an autoimmune disease risk allele in 22q11.21 as associated with systemic lupus erythematosus

IntroductionSystemic lupus erythematosus (SLE) is a heterogeneous disease with a diverse spectrum of clinical symptoms, ranging from skin rash to end-organ damage. 22q11.21 has been identified as a susceptibility region for several autoimmune diseases, including SLE. However, detailed information for SLE association and the underlying functional mechanism(s) is still lacking.MethodsThrough meta-analysis of two genome-wide association studies (GWAS) on Han Chinese populations, comprising a total of 1,659 cases and 3,398 controls matched geographically, we closely examined the 22q11.21 region, especially on the reported single-nucleotide polymorphisms (SNPs) associated with different autoimmune diseases and their relationships. We further replicated the most significant associations of SNPs with SLE using 2,612 cases and 2,323 controls of Asian ancestry.ResultsAll reported SNPs in the 22q11.21 region with different autoimmune diseases were examined using the two GWAS data and meta-analysis results, and supportive evidence of association with SLE was found (meta-analysis: P_meta ≤ 7.27E-05), which might require further investigation. SNP rs2298428 was identified as the most significant SNP associated with SLE in this region (P_meta =2.70E-09). It showed independent effects through both stepwise and conditional logistic regression, and there is no evidence of other independent association signals for SLE in this region. The association of rs2298428 was further replicated in three cohorts from Hong Kong, Anhui and Thailand comprising a total of 2,612 cases and 2,323 controls (joint analysis of GWAS and replication result: P_all =1.31E-11, odds ratio =1.23). SNP rs2298428 was shown to be an expression quantitative locus for UBE2L3 gene in different cell types, with the risk allele (T) being correlated with higher expression of UBE2L3. This is consistent with earlier reports on higher expression of UBE2L3 in patients with SLE.ConclusionsAssociation with distinct autoimmune diseases highlights the significance of this region in autoreactive responses and potentially shared functional mechanisms in these diseases.

Electronic supplementary material

The online version of this article (doi:10.1186/s13075-015-0577-6) contains supplementary material, which is available to authorized users.     

Treatment response to low-dose ketamine infusion for treatment-resistant depression: A gene-based genome-wide association study

BackgroundsEvidence suggested the crucial roles of brain-derived neurotrophic factor (BDNF) and glutamate system functioning in the antidepressant mechanisms of low-dose ketamine infusion in treatment-resistant depression (TRD).Methods65 patients with TRD were genotyped for 684,616 single nucleotide polymorphisms (SNPs). Twelve ketamine-related genes were selected for the gene-based genome-wide association study on the antidepressant effect of ketamine infusion and the resulting serum ketamine and norketamine levels.ResultsSpecific SNPs and whole genes involved in BDNF–TrkB signaling (i.e., rs2049048 in BDNF and rs10217777 in NTRK2) and the glutamatergic and GABAergic systems (i.e., rs16966731 in GRIN2A) were associated with the rapid (within 240 min) and persistent (up to 2 weeks) antidepressant effect of low-dose ketamine infusion and with serum ketamine and norketamine levels.DiscussionOur findings confirmed the predictive roles of BDNF–TrkB signaling and glutamatergic and GABAergic systems in the underlying mechanisms of low-dose ketamine infusion for TRD treatment.     

Genetic factors of reaction time performance: DRD4 7‐repeat allele associated with slower responses

Twin studies indicate substantial inherited components in cognitive abilities. One of the most extensively studied candidate genes of cognitive functioning is the dopamine D4 receptor gene (DRD4), which has been suggested to be related to attentional disorders. Based on reaction time data of 245 Caucasians participating in different cognitive tasks, slower responses characterized the group with the 7‐repeat allele. This effect was present in both sexes and was not because of fatigue. To our knowledge, this is the first report on significant association (P = 0.0001) between the DRD4 variable number of tandem repeat (VNTR) polymorphism and response latencies in a non‐clinical adult sample. Other studied dopaminergic polymorphisms did not show an association with reaction time. These results illustrate that speed‐of‐performance measures derived from multiple reaction time tasks using standardization procedures could be promising tools to detect unique genetic effects in the background of cognitive abilities.     

Comprehensive SNP Scan of DNA Repair and DNA Damage Response Genes Reveal Multiple Susceptibility Loci Conferring Risk to Tobacco Associated Leukoplakia and Oral Cancer

Polymorphic variants of DNA repair and damage response genes play major role in carcinogenesis. These variants are suspected as predisposition factors to Oral Squamous Cell Carcinoma (OSCC). For identification of susceptible variants affecting OSCC development in Indian population, the “maximally informative” method of SNP selection from HapMap data to non-HapMap populations was applied. Three hundred twenty-five SNPs from 11 key genes involved in double strand break repair, mismatch repair and DNA damage response pathways were genotyped on a total of 373 OSCC, 253 leukoplakia and 535 unrelated control individuals. The significantly associated SNPs were validated in an additional cohort of 144 OSCC patients and 160 controls. The rs12515548 of MSH3 showed significant association with OSCC both in the discovery and validation phases (discovery P-value: 1.43E-05, replication P-value: 4.84E-03). Two SNPs (rs12360870 of MRE11A, P-value: 2.37E-07 and rs7003908 of PRKDC, P-value: 7.99E-05) were found to be significantly associated only with leukoplakia. Stratification of subjects based on amount of tobacco consumption identified SNPs that were associated with either high or low tobacco exposed group. The study reveals a synergism between associated SNPs and lifestyle factors in predisposition to OSCC and leukoplakia.     

Identifying consensus disease pathways in Parkinson's disease using an integrative systems biology approach

Parkinson's disease (PD) has had six genome-wide association studies (GWAS) conducted as well as several gene expression studies. However, only variants in MAPT and SNCA have been consistently replicated. To improve the utility of these approaches, we applied pathway analyses integrating both GWAS and gene expression. The top 5000 SNPs (p<0.01) from a joint analysis of three existing PD GWAS were identified and each assigned to a gene. For gene expression, rather than the traditional comparison of one anatomical region between sets of patients and controls, we identified differentially expressed genes between adjacent Braak regions in each individual and adjusted using average control expression profiles. Over-represented pathways were calculated using a hyper-geometric statistical comparison. An integrated, systems meta-analysis of the over-represented pathways combined the expression and GWAS results using a Fisher's combined probability test. Four of the top seven pathways from each approach were identical. The top three pathways in the meta-analysis, with their corrected p-values, were axonal guidance (p = 2.8E-07), focal adhesion (p = 7.7E-06) and calcium signaling (p = 2.9E-05). These results support that a systems biology (pathway) approach will provide additional insight into the genetic etiology of PD and that these pathways have both biological and statistical support to be important in PD.     

Heroin addiction in African Americans: a hypothesis-driven association study

Heroin addiction is a chronic complex disease with a substantial genetic contribution. This study was designed to identify gene variants associated with heroin addiction in African Americans. The emphasis was on genes involved in reward modulation, behavioral control, cognitive function, signal transduction and stress response. We have performed a case–control association analysis by screening with 1350 variants of 130 genes. The sample consisted of 202 former severe heroin addicts in methadone treatment and 167 healthy controls with no history of drug abuse. Single nucleotide polymorphism (SNP), haplotype and multi-SNP genotype pattern analyses were performed. Seventeen SNPs showed point-wise significant association with heroin addiction (nominal P < 0.01). These SNPs are from genes encoding several receptors: adrenergic ( ADRA1A ), arginine vasopressin ( AVPR1A ), cholinergic ( CHRM2 ), dopamine (DRD1 ), GABA-A ( GABRB3 ), glutamate ( GRIN2A ) and serotonin ( HTR3A ) as well as alcohol dehydrogenase ( ADH7 ), glutamic acid decarboxylase ( GAD1 and GAD2 ), the nucleoside transporter ( SLC29A1 ) and diazepam-binding inhibitor ( DBI ). The most significant result of the analyses was obtained for the GRIN2A haplotype G-A-T (rs4587976-rs1071502-rs1366076) with protective effect ( P _uncorrected = 9.6E- 05, P _corrected = 0.058). This study corroborates several reported associations with alcohol and drug addiction as well as other related disorders and extends the list of variants that may affect the development of heroin addiction. Further studies will be necessary to replicate these associations and to elucidate the roles of these variants in drug addiction vulnerability.     

Modeling of exposure to mercury in different environmental media over a 30-year period: A case study of Shimen reservoir,northern Taiwan

Abstract

Mercury is affected by the movement mechanisms in the environmental media and is normally present in dry and wet depositions and surface and water vapor, among other things. The rapid growth of mercury-related industries in the past two decades reflects the result of its increased use in water sources such as in the Shimen reservoir, northern Taiwan. Consequently, residents living nearby are exposed to mercury almost every day. In light of the effects of continued exposure to the deleterious properties of mercury, this study provides modeling results of the atmosphere, soil, and freshwater over a 30-year period (2016–2046). The associated influences in the media and mercury contamination during this period will be determined via sensitivity analysis. Finally, the results of this study facilitate the assessment of potential health hazards associated with mercury inhalation and the ingestion of MeHg-contaminated fish. The mean daily dose (mg/kg) and hazard quotient (HQ) in the children and adult were 3.52E-13 (HQ = 4.10E-09) and 1.19E-13 (HQ = 1.39E-09) for Hg inhalation and 6.38E-05 (HQ = 6.38E-01) and 4.47E-05 (HQ = 4.47E-01) for ingestion of MeHg+-contaminated fish.     

Pathway analysis of a genome-wide association study in schizophrenia

Objective

The aim of this study was to identify the candidate single nucleotide polymorphisms (SNPs) and candidate mechanisms that contribute to schizophrenia susceptibility and to generate a SNP to gene to pathway hypothesis using an analytical pathway-based approach.

Methods

We used schizophrenia GWAS data of the genotypes of 660,259 SNPs in 1378 controls and 1351 cases of European descent after quality control filtering. ICSNPathway (Identify candidate Causal SNPs and Pathways) analysis was applied to the schizophrenia GWAS dataset. The first stage involved the pre-selection of candidate SNPs by linkage disequilibrium analysis and the functional SNP annotation of the most significant SNPs found. The second stage involved the annotation of biological mechanisms for the pre-selected candidate SNPs using improved-gene set enrichment analysis.

Results

ICSNPathway analysis identified fifteen candidate SNPs, ten candidate pathways, and nine hypothetical biological mechanisms. The most strongly associated potential pathways were as follows. First, rs1644731 and rs1644730 to RDH8 to estrogen biosynthetic process (p < 0.001, FDR < 0.001). The genes involved in this pathway are RDH8 and HSD3B1 (p < 0.05). All-trans-retinol dehydrogenase (RDH8) is a visual cycle enzyme that reduces all-trans-retinal to all-trans-retinol in the presence of NADPH. The chemical reactions and pathways involved result in the formation of estrogens, which are C18 steroid hormones that can stimulate the development of female sexual characteristics. Second, rs1146031 to ACVR1 to mesoderm formation and activin binding (p < 0.001, FDR = 0.032, 0.034). Two of 15 candidate genes are known genes associated with schizophrenia: KCNQ2 and APOL2. One of the 10 candidate pathways, estrogen biosynthetic process, is known to be associated with schizophrenia (p < 0.001, FDR < 0.001). However, 13 of candidate genes (RDH8, ACVR1, PSMD9, KCNAB1, SLC17A3, ARCN1, COG7, STAB2, LRPAP1, STAB1, CXCL16, COL4A4, EXOSC3) and 9 of candidate pathways were novel.

Conclusion

By applying ICSNPathway analysis to schizophrenia GWAS data, we identified candidate SNPs, genes like KCNQ2 and APOL2 and pathways involving the estrogen biosynthetic process may contribute to schizophrenia susceptibility. Further analyses are needed to validate the results of this analysis.     

Novel insights into systemic autoimmune rheumatic diseases using shared molecular signatures and an integrative analysis

We undertook this study to identify DNA methylation signatures of three systemic autoimmune rheumatic diseases (SARDs), namely rheumatoid arthritis, systemic lupus erythematosus, and systemic sclerosis, compared to healthy controls. Using a careful design to minimize confounding, we restricted our study to subjects with incident disease and performed our analyses on purified CD4⁺ T cells, key effector cells in SARD. We identified differentially methylated (using the Illumina Infinium HumanMethylation450 BeadChip array) and expressed (using the Illumina TruSeq stranded RNA-seq protocol) sites between cases and controls, and investigated the biological significance of this SARD signature using gene annotation databases. We recruited 13 seropositive rheumatoid arthritis, 19 systemic sclerosis, 12 systemic lupus erythematosus subjects, and 8 healthy controls. We identified 33 genes that were both differentially methylated and expressed (26 over- and 7 under-expressed) in SARD cases versus controls. The most highly overexpressed gene was CD1C (log fold change in expression = 1.85, adjusted P value = 0.009). In functional analysis (Ingenuity Pathway Analysis), the top network identified was lipid metabolism, molecular transport, small molecule biochemistry. The top canonical pathways included the mitochondrial L-carnitine shuttle pathway (P = 5E-03) and PTEN signaling (P = 8E-03). The top upstream regulator was HNF4A (P = 3E-05). This novel SARD signature contributes to ongoing work to further our understanding of the molecular mechanisms underlying SARD and provides novel targets of interest.     

A scan of chromosome 10 identifies a novel locus showing strong association with late-onset Alzheimer disease

Strong evidence of linkage to late-onset Alzheimer disease (LOAD) has been observed on chromosome 10, which implicates a wide region and at least one disease-susceptibility locus. Although significant associations with several biological candidate genes on chromosome 10 have been reported, these findings have not been consistently replicated, and they remain controversial. We performed a chromosome 10–specific association study with 1,412 gene-based single-nucleotide polymorphisms (SNPs), to identify susceptibility genes for developing LOAD. The scan included SNPs in 677 of 1,270 known or predicted genes; each gene contained one or more markers, about half (48%) of which represented putative functional mutations. In general, the initial testing was performed in a white case-control sample from the St. Louis area, with 419 LOAD cases and 377 age-matched controls. Markers that showed significant association in the exploratory analysis were followed up in several other white case-control sample sets to confirm the initial association. Of the 1,397 markers tested in the exploratory sample, 69 reached significance (P<.05). Five of these markers replicated at P<.05 in the validation sample sets. One marker, rs498055, located in a gene homologous to RPS3A (LOC439999), was significantly associated with Alzheimer disease in four of six case-control series, with an allelic P value of .0001 for a meta-analysis of all six samples. One of the case-control samples with significant association to rs498055 was derived from the linkage sample (P=.0165). These results indicate that variants in the RPS3A homologue are associated with LOAD and implicate this gene, adjacent genes, or other functional variants (e.g., noncoding RNAs) in the pathogenesis of this disorder.     

Clinical Utility of a Coronary Heart Disease Risk Prediction Gene Score in UK Healthy Middle Aged Men and in the Pakistani Population

BackgroundNumerous risk prediction algorithms based on conventional risk factors for Coronary Heart Disease (CHD) are available but provide only modest discrimination. The inclusion of genetic information may improve clinical utility.MethodsWe tested the use of two gene scores (GS) in the prospective second Northwick Park Heart Study (NPHSII) of 2775 healthy UK men (284 cases), and Pakistani case-control studies from Islamabad/Rawalpindi (321 cases/228 controls) and Lahore (414 cases/219 controls). The 19-SNP GS included SNPs in loci identified by GWAS and candidate gene studies, while the 13-SNP GS only included SNPs in loci identified by the CARDIoGRAMplusC4D consortium.ResultsIn NPHSII, the mean of both gene scores was higher in those who went on to develop CHD over 13.5 years of follow-up (19-SNP p=0.01, 13-SNP p=7x10^-3). In combination with the Framingham algorithm the GSs appeared to show improvement in discrimination (increase in area under the ROC curve, 19-SNP p=0.48, 13-SNP p=0.82) and risk classification (net reclassification improvement (NRI), 19-SNP p=0.28, 13-SNP p=0.42) compared to the Framingham algorithm alone, but these were not statistically significant. When considering only individuals who moved up a risk category with inclusion of the GS, the improvement in risk classification was statistically significant (19-SNP p=0.01, 13-SNP p=0.04). In the Pakistani samples, risk allele frequencies were significantly lower compared to NPHSII for 13/19 SNPs. In the Islamabad study, the mean gene score was higher in cases than controls only for the 13-SNP GS (2.24 v 2.34, p=0.04). There was no association with CHD and either score in the Lahore study.ConclusionThe performance of both GSs showed potential clinical utility in European men but much less utility in subjects from Pakistan, suggesting that a different set of risk loci or SNPs may be required for risk prediction in the South Asian population.     

Polymorphisms in the 3′ UTR in the neurocalcin δ gene affect mRNA stability, and confer susceptibility to diabetic nephropathy

Using a large-scale genotyping analysis of gene-based single nucleotide polymorphisms (SNPs) in Japanese type 2 diabetic patients, we have identified a gene encoding neurocalcin δ (NCALD) as a candidate for a susceptibility gene to diabetic nephropathy; the landmark SNP was found in the 3′ UTR of NCALD (rs1131863: exon 4 +1340 A vs. G, P = 0.00004, odds ratio = 1.59, 95% CI 1.27–1.98). We also discovered two other SNPs in exon 4 of this gene (+999 T/A, +1307 A/G) that showed absolute linkage disequilibrium to the landmark SNP. Subsequent in vitro functional analysis revealed that synthetic mRNA corresponding to the disease susceptible haplotype (exon 4 +1340 G, +1307 G, +999 A) was degraded faster than mRNA corresponding to the major haplotype (exon 4 +1340 A, +1307 A, +999 T), and allelic mRNA expression of the disease susceptibility allele was significantly lower than that of the major allele in normal kidney tissues. In an experiment using a short interfering RNA targeting NCALD, we found that silencing of the NCALD led to a considerable enhancement of cell migration, accompanied by a significant reduction in E-cadherin expression, and by an elevation of α smooth muscle actin expression in cultured renal proximal tubular epithelial cells. We also identified the association of the landmark SNP with the progression of diabetic nephropathy in a 8-year prospective study (A vs. G, P = 0.03, odds ratio = 1.91, 95% CI 1.07–3.42). These results suggest that the NCALD gene is a likely candidate for conferring susceptibility to diabetic nephropathy. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.