Associations between anxious-depressed symptoms and cardiovascular risk factors in a longitudinal childhood study.

OBJECTIVE: To examine the influence of anxious/depressed scores on cardiovascular risk factors throughout childhood. METHODS: Data from the Western Australian Pregnancy Cohort (Raine) Study, a study of 2900 pregnancies recruited between 1989 and 1991, were used. Anxious-depressed scores (derived from the Childhood Behavior Checklist), body mass index (BMI) and blood pressure were measured at 5 (n=1681), 8 (n=1697), 10 (n=1575) and 14 (n=1386) years. At age 14 depressive symptom scores (Beck Depression Inventory for Youth), anxious-depressed scores (Youth Self-Report (YSR) and Teacher Report Form (TRF)) and fasting lipid, glucose and insulin were also available. Cross sectional and longitudinal analyses were conducted. RESULTS: At age 14, girls with higher anxious-depressed scores had higher BMI (p≤ 0.005) and homeostasis model assessment-estimated insulin resistance (p≤ 0.0001). This equated to a difference of 0.6 kg/m(2) and 0.3 units in predicted BMI and HOMA-IR respectively (top 5% vs. score of zero). Boys with higher anxious-depressed scores had lower systolic blood pressure trajectories (p=0.024). CONCLUSION: Depressive scores appear to have differing influences on BMI, homeostasis model assessment-estimated insulin resistance and systolic blood pressure in boys and girls. Paradoxically boys with higher anxious-depressed scores had lower blood pressure throughout childhood.

Separating the mechanism-based and off-target actions of cholesteryl ester transfer protein inhibitors with CETP gene polymorphisms.

BACKGROUND: Cholesteryl ester transfer protein (CETP) inhibitors raise high-density lipoprotein (HDL) cholesterol, but torcetrapib, the first-in-class inhibitor tested in a large outcome trial, caused an unexpected blood pressure elevation and increased cardiovascular events. Whether the hypertensive effect resulted from CETP inhibition or an off-target action of torcetrapib has been debated. We hypothesized that common single-nucleotide polymorphisms in the CETP gene could help distinguish mechanism-based from off-target actions of CETP inhibitors to inform on the validity of CETP as a therapeutic target. METHODS AND RESULTS: We compared the effect of CETP single-nucleotide polymorphisms and torcetrapib treatment on lipid fractions, blood pressure, and electrolytes in up to 67 687 individuals from genetic studies and 17 911 from randomized trials. CETP single-nucleotide polymorphisms and torcetrapib treatment reduced CETP activity and had a directionally concordant effect on 8 lipid and lipoprotein traits (total, low-density lipoprotein, and HDL cholesterol; HDL2; HDL3; apolipoproteins A-I and B; and triglycerides), with the genetic effect on HDL cholesterol (0.13 mmol/L, 95% confidence interval [CI] 0.11 to 0.14 mmol/L) being consistent with that expected of a 10-mg dose of torcetrapib (0.13 mmol/L, 95% CI 0.10 to 0.15). In trials, 60 mg of torcetrapib elevated systolic and diastolic blood pressure by 4.47 mm Hg (95% CI 4.10 to 4.84 mm Hg) and 2.08 mm Hg (95% CI 1.84 to 2.31 mm Hg), respectively. However, the effect of CETP single-nucleotide polymorphisms on systolic blood pressure (0.16 mm Hg, 95% CI -0.28 to 0.60 mm Hg) and diastolic blood pressure (-0.04 mm Hg, 95% CI -0.36 to 0.28 mm Hg) was null and significantly different from that expected of 10 mg of torcetrapib. CONCLUSIONS: Discordance in the effects of CETP single-nucleotide polymorphisms and torcetrapib treatment on blood pressure despite the concordant effects on lipids indicates the hypertensive action of torcetrapib is unlikely to be due to CETP inhibition or shared by chemically dissimilar CETP inhibitors. Genetic studies could find a place in drug-development programs as a new source of randomized evidence for drug-target validation in humans.

Association of PPARgamma allelic variation, osteoprotegerin and abdominal aortic aneurysm.

OBJECTIVE: We have previously demonstrated high concentrations of the glycoprotein osteoprotegerin (OPG) in biopsies of abdominal aortic aneurysm (AAA), and demonstrated that ligation of the nuclear receptor peroxisome proliferator-activated receptor gamma (PPARgamma) downregulates OPG in vitro and within a mouse model. The aims of this study were to assess the associations between circulating concentrations of OPG, polymorphisms of the gene encoding PPARgamma (PPARG), AAA presence and growth. DESIGN, PATIENTS AND MEASUREMENTS: Two genetic polymorphisms in PPARG were assessed in 4227 men, 699 of whom had an AAA. For 631 men, who had AAAs, maximum aortic diameter was monitored by yearly ultrasound for a median of 5 years. Plasma OPG was measured in 838 men, 318 of whom had an AAA. RESULTS: Plasma concentrations of OPG were independently associated with AAA (adjusted odds ratio 1.38, 95% CI 1.10-1.72). The PPARG c.1347C > T polymorphism was associated with plasma concentrations of OPG (beta 0.12, P < 0.01). The PPARG c.34G > C polymorphism was weakly associated with AAA (adjusted odds ratio 1.28, 95% CI 1.01-1.61). PPARG c.1347C > T was associated with increased AAA growth (recessive model, P = 0.03). CONCLUSIONS: Circulating concentrations of osteoprotegerin are associated with abdominal aortic aneurysm and with one peroxisome proliferator-activated receptor gamma gene polymorphism. Peroxisome proliferator-activated receptor gamma gene polymorphisms are weakly associated with abdominal aortic aneurysm presence and growth. Confirmation of these findings is required in other cohorts.

Analyses of associations with asthma in four asthma population samples from Canada and Australia.

Asthma, atopy, and related phenotypes are heterogeneous complex traits, with both genetic and environmental risk factors. Extensive research has been conducted and over hundred genes have been associated with asthma and atopy phenotypes, but many of these findings have failed to replicate in subsequent studies. To separate true associations from false positives, candidate genes need to be examined in large well-characterized samples, using standardized designs (genotyping, phenotyping and analysis). In an attempt to replicate previous associations we amalgamated the power and resources of four studies and genotyped 5,565 individuals to conduct a genetic association study of 93 previously associated candidate genes for asthma and related phenotypes using the same set of 861 single-nucleotide polymorphisms (SNPs), a common genotyping platform, and relatively harmonized phenotypes. We tested for association between SNPs and the dichotomous outcomes of asthma, atopy, atopic asthma, and airway hyperresponsiveness using a general allelic likelihood ratio test. No SNP in any gene reached significance levels that survived correction for all tested SNPs, phenotypes, and genes. Even after relaxing the usual stringent multiple testing corrections by performing a gene-based analysis (one gene at a time as if no other genes were typed) and by stratifying SNPs based on their prior evidence of association, no genes gave strong evidence of replication. There was weak evidence to implicate the following: IL13, IFNGR2, EDN1, and VDR in asthma; IL18, TBXA2R, IFNGR2, and VDR in atopy; TLR9, TBXA2R, VDR, NOD2, and STAT6 in airway hyperresponsiveness; TLR10, IFNGR2, STAT6, VDR, and NPSR1 in atopic asthma. Additionally we found an excess of SNPs with small effect sizes (OR < 1.4). The low rate of replication may be due to small effect size, differences in phenotypic definition, differential environmental effects, and/or genetic heterogeneity. To aid in future replication studies of asthma genes a comprehensive database was compiled and is available to the scientific community at http://genapha.icapture.ubc.ca/.

SimHap GUI: an intuitive graphical user interface for genetic association analysis.

BACKGROUND: Researchers wishing to conduct genetic association analysis involving single nucleotide polymorphisms (SNPs) or haplotypes are often confronted with the lack of user-friendly graphical analysis tools, requiring sophisticated statistical and informatics expertise to perform relatively straightforward tasks. Tools, such as the SimHap package for the R statistics language, provide the necessary statistical operations to conduct sophisticated genetic analysis, but lacks a graphical user interface that allows anyone but a professional statistician to effectively utilise the tool. RESULTS: We have developed SimHap GUI, a cross-platform integrated graphical analysis tool for conducting epidemiological, single SNP and haplotype-based association analysis. SimHap GUI features a novel workflow interface that guides the user through each logical step of the analysis process, making it accessible to both novice and advanced users. This tool provides a seamless interface to the SimHap R package, while providing enhanced functionality such as sophisticated data checking, automated data conversion, and real-time estimations of haplotype simulation progress. CONCLUSION: SimHap GUI provides a novel, easy-to-use, cross-platform solution for conducting a range of genetic and non-genetic association analyses. This provides a free alternative to commercial statistics packages that is specifically designed for genetic association analysis.

Association of PARL rs3732581 genetic variant with insulin levels, metabolic syndrome and coronary artery disease.

PARL (presenilin-associated rhomboid-like) is a mitochondrial protein involved in mitochondrial membrane remodelling, and maps to a quantitative trait locus (3q27) associated with metabolic traits. Recently the rs3732581 (Leu262Val) variant was found to be associated with increased levels of plasma insulin, a finding not replicated in a larger cohort. The aim of the current study was to investigate the associations between rs3732581 and levels of plasma insulin, metabolic syndrome (MetS) and its components, and cardiovascular disease. The CUPID population consisted of 556 subjects with angiographically proven CAD and the CUDAS cohort consisted of 1,109 randomly selected individuals from Perth, Western Australia. Samples were genotyped using mutation-specific PCR. No significant associations were observed between rs3732581 and levels of plasma insulin, glucose, BMI or MetS in either population. However, carriers of the minor allele had significantly lower mean intima-media thickness (IMT) [0.69 mm, 95% CI (0.69, 0.70 mm); P = 0.004], compared with major allele homozygotes [mean IMT = 0.71 mm, 95% CI (0.70, 0.72 mm)] in the CUDAS population. Further analysis using a recessive model showed homozygous carriers of the minor allele were predisposed to CAD [OR 1.55, 95% CI (1.11, 2.16); P = 0.01]. Despite the functional evidence for a role of PARL in regulating insulin levels, no association with rs3732581 was found in the current study. Additionally, there were no associations with glucose levels, BMI or MetS. There were significant effects of the variant on mean IMT and risk of CAD. A role for PARL in metabolic conditions cannot be excluded and more comprehensive genetic studies are warranted.

Association of Interleukin-1 gene polymorphisms with central obesity and metabolic syndrome in a coronary heart disease population.

The objective of this study was to determine whether single nucleotide polymorphisms (SNPs) in the Interleukin-1 (IL-1) gene family are associated with central obesity and metabolic syndrome in a coronary heart disease population. The IL-1 alpha C-889T (rs1800587) and IL-1 beta +3954 (rs1143634) SNPs were studied in a Western Australian coronary heart disease (CHD) population (N = 556). Subjects who were TT homozygous at either SNP had larger waist circumference (IL-1 alpha: 1.8 cm greater, P = 0.04; IL-1 beta: 4 cm greater, P = 0.0004) compared with major allele homozygotes. Individuals with two copies of the IL-1 alpha:IL-1 beta T:T haplotype had greater waist circumference (4.7 cm greater, P = 0.0001) compared to other haplotypes. There was a significant interaction between the IL-1 beta SNP and BMI level on waist circumference (P = 0.01). When the cohort was stratified by median BMI, TT carriers for IL-1 beta with above median BMI had greater waist circumference (6.1 cm greater, P = 0.007) compared to baseline carriers, whilst no significant association was seen in the below median group. Similarly, when the cohort was stratified by median fibrinogen level (IL-1 alpha interaction P = 0.01; IL-1 beta interaction P = 0.04), TT carriers for both SNPs in the above median fibrinogen group had greater waist circumference (IL-1 alpha 2.7 cm greater, P = 0.007; IL-1 beta 3.3 cm greater, P = 0.003) compared with major allele homozygotes. This association was not seen in the below median group. Also, we found a trend of increased metabolic syndrome for IL-1 beta TT homozygotes (P = 0.07). In conclusion, our findings suggest that in a CHD population IL-1 gene polymorphisms may be involved in increased central obesity, and the genetic influences are more evident among patients who have a higher level of obesity or inflammatory markers.

15-Lipoxygenase gene variants are associated with carotid plaque but not carotid intima-media thickness.

The major underlying cause of CHD is atherosclerosis, and oxidised LDL is known to play an important role in its development. We examined the role of three single nucleotide polymorphisms (SNPs) in the 15-lipoxygenase gene (ALOX15), in atherosclerosis. We genotyped three SNPs in the ALOX15 promoter in two Western Australian samples-1,111 community-based individuals and 556 with CHD. SNPs and haplotypes were tested for an association with carotid plaque, intima-media thickness and risk of CHD. The -611GG genotype was associated with increased likelihood of carotid plaque in CHD patients (OR = 4.01, 95%CI = 1.39-11.53, P = 0.005) and the C alleles of the G-220C and G-189C SNPs were associated with decreased likelihood of plaque among cases (OR = 0.66, 95%CI = 0.43-0.99, P = 0.05 and OR = 0.51, 95%CI = 0.34-0.78, P = 0.002 respectively). The GGG haplotype was associated with increased risk of carotid plaque in CHD patients (OR = 5.77, 95%CI = 1.82-18.29, P = 0.0007) and in community-based individuals under 53 years (OR = 4.15, 95%CI = 1.23-14.08, P = 0.02). No association was observed between ALOX15 SNPs or haplotypes and intima-media thickness. This study is novel as it is the first to examine the association between 15-lipoxygenase polymorphisms and atherosclerotic indicators. These findings suggest a possible role of ALOX15 polymorphisms in focal plaque formation.

Investigating the association between K198N coding polymorphism in EDN1 and hypertension, lipoprotein levels, the metabolic syndrome and cardiovascular disease.

Endothelin-1 is a potent vasoconstrictor in the body. Previous studies have identified associations between the coding polymorphism K198N and hypertension, systolic blood pressure and HDL levels. We sought to examine the evidence for these associations and, additionally, the association between K198N, insulin resistance, metabolic syndrome and coronary artery disease (CAD). We used generalised linear modelling to test K198N for association with hypertension and systolic blood pressure, lipid levels, insulin resistance scores and metabolic syndrome in a general cross-sectional community sample. Mean carotid intima media thickness and risk of carotid plaque were examined in the general population sample, and Gensini score was examined in a sample of patients with CAD. A case/control sample was used to examine the association of K198N with risk of CAD. There was no significant evidence for association between K198N and hypertension, systolic blood pressure, lipid levels, insulin resistance or metabolic syndrome in either population. The minor allele was marginally associated with increased mean IMT levels (P = 0.02) in the general population sample, although not with CAD in the case/control study or with the severity of disease in patients with CAD. In conclusion, we found no robust evidence for the associations between K198N and hypertension, systolic blood pressure or HDL levels seen in previous studies.

The apolipoprotein AII rs5082 variant is associated with reduced risk of coronary artery disease in an Australian male population.

Serum high density lipoprotein (HDL) levels are inversely related to the development of coronary artery disease (CAD). Apolipoproteins AI and AII are the major protein constituents of HDL particles. APOAI and APOAII genetic polymorphisms have been proposed to affect transcriptional efficiency of their respective genes, thereby altering serum lipid levels and influencing atherosclerotic disease risk. 556 subjects with angiographically proven CAD (>50% stenosis) and 1109 randomly selected individuals from metropolitan Perth, Western Australia, were included in an association study. APOAI -75G/A (rs670) and APOAII -256T/C (rs5082) polymorphisms were both found to be not associated with plasma HDL levels. In a case-control analysis of 484 male CAD patients and 498 male controls, individuals carrying the 'CC' genotype for the APOAII rs5082 polymorphism had significantly lower risk of CAD than the 'T' allele carriers (OR=0.57, 95% CI 0.39-0.84, p=0.004). The minor 'A' allele of the APOAI rs670 polymorphism was found to be not associated with CAD, contrary to previous reports. We conclude that the APOAII rs5082 polymorphism appears to be cardioprotective in this representative Caucasian Australian population.

IL18 haplotypes are associated with serum IL-18 concentrations in a population-based study and a cohort of individuals with premature coronary heart disease.

BACKGROUND: Interleukin (IL)-18 is a proinflammatory cytokine that has been implicated in several diseases, including atherosclerosis, and increased circulating IL-18 concentrations increase risk of future coronary heart disease (CHD). We evaluated the effect of common variation within the IL18 gene on concentrations of circulating IL-18. METHODS: We measured IL-18, by ELISA, in the population-based study group [Carotid Ultrasound Disease Assessment Study (CUDAS)] and a predominantly male cohort with premature cardiovascular disease [Carotid Ultrasound in Patients with Ischaemic Heart Disease (CUPID)]. Using a tagging single-nucleotide polymorphism (SNP) approach that captured >90% of genetic variation, we identified 4 common (>10%) haplotypes. RESULTS: A common SNP was associated with differences in IL-18 concentrations; in CUDAS individuals carrying 2 copies of the rare allele, concentrations were 13% higher than in those with no copies (P = 0.002). Haplotypes were also associated with significant differences in IL-18 concentrations in CUDAS and CUPID. Haplotype GTATA (frequency 23%) was associated with significantly lower IL-18 than others. In CUDAS, those carrying 2 copies had IL-18 concentrations 15% lower than those carrying no copies (P = 0.002); in CUPID, the difference was 22% (P = 0.004). These associations remained significant after adjustment for age, sex, hypertension, HDL cholesterol, waist-to-hip ratio, and alcohol consumption. Despite being associated with differences in IL-18 concentrations, the haplotypes did not occur at different frequencies in those with or without carotid atherosclerotic plaques. CONCLUSIONS: Variation within IL18 affects IL-18 concentrations in healthy and diseased individuals and thus may influence the pathophysiology of plaques at all stages of CHD progression.

Cholesteryl ester transfer protein gene haplotypes, plasma high-density lipoprotein levels and the risk of coronary heart disease.

High-density lipoprotein cholesterol (HDL-C) is a known inverse predictor of coronary heart disease (CHD) and is thus a potential therapeutic target. Cholesteryl ester transfer protein (CETP) is a key protein in HDL-C metabolism such that elevated CETP activity is associated with lower HDL-C. Currently available HDL-C raising drugs are relatively ineffective and evidence suggesting the role of CETP in HDL-C levels has promoted the development of CETP inhibitors as potential therapeutic agents for CHD. We investigated three SNPs in the CETP gene in two cross-sectional community-based populations (n = 1,574 and 1,109) and a population of 556 CHD patients to determine if reduced CETP activity due to genetic variations in the CETP gene would increase HDL-C levels and reduce the risk of CHD. CETP genotypes and haplotypes were tested for association with lipid levels, CETP activity and risk of CHD. Multivariate analysis showed the common AAB2 haplotype defined by the G-2708A, C-629A and TaqIB polymorphisms, was consistently associated with reduced CETP activity and increased HDL-C levels. A mean increase in HDL-C levels of 0.16-0.24 mmol/l was observed in individuals with two copies of the AAB2 haplotype relative to non AAB2 carriers across all three populations (P < 0.001). A case-control study of males indicated no association between single SNPs or haplotypes and the risk of CHD. These results suggest that raising HDL-C via CETP inhibition may not alter risk of CHD. Randomized control trials are needed to determine whether CETP inhibition will in reality reduce risk of CHD by raising HDL-C.

JLIN: a java based linkage disequilibrium plotter.

BACKGROUND: A great deal of effort and expense are being expended internationally in attempts to detect genetic polymorphisms contributing to susceptibility to complex human disease. Techniques such as Linkage Disequilibrium mapping are being increasingly used to examine and compare markers across increasingly large datasets. Visualisation techniques are becoming essential to analyse the ever-growing volume of data and results available with any given analysis. RESULTS: JLIN (Java LINkage disequilibrium plotter) is a software package designed for customisable, intuitive visualisation of Linkage Disequilibrium (LD) across all common computing platforms. Customisation allows the user to choose particular visualisations, statistical measures and measurement ranges. JLIN also allows the user to export images of the LD visualisation in several common document formats. CONCLUSION: JLIN allows the user to visually compare and contrast the results of a range of statistical measures on the input dataset(s). These measures include the commonly used D' and r2 statistics and empirical p-values. JLIN has a number of unique and novel features that improve on existing LD visualisation tools.

The effect of missing data on linkage disequilibrium mapping and haplotype association analysis in the GAW14 simulated datasets.

We used our newly developed linkage disequilibrium (LD) plotting software, JLIN, to plot linkage disequilibrium between pairs of single-nucleotide polymorphisms (SNPs) for three chromosomes of the Genetic Analysis Workshop 14 Aipotu simulated population to assess the effect of missing data on LD calculations. Our haplotype analysis program, SIMHAP, was used to assess the effect of missing data on haplotype-phenotype association. Genotype data was removed at random, at levels of 1%, 5%, and 10%, and the LD calculations and haplotype association results for these levels of missingness were compared to those for the complete dataset. It was concluded that ignoring individuals with missing data substantially affects the number of regions of LD detected which, in turn, could affect tagging SNPs chosen to generate haplotypes.

Genome-wide linkage and association mapping of disease genes with the GAW14 simulated datasets.

We combined the results of whole-genome linkage and association analyses to determine which markers were most strongly associated with Kofendrerd Personality Disorder. Using replicate 1 from the Genetic Analysis Workshop 14 Aipotu, Karangar, Danacaa, and New York City simulated populations, we determined that several markers showed significant linkage and association with disease status. We used both SNP and microsatellite markers to determine patterns and chromosomal regions of markers. Three consistently associated markers were C01R0050, C03R0280, and C10R0882. Using generalized linear mixed models, we modelled the effect of the three predefined phenotypic categories on disease status and concluded that the phenotypes defining the "anxiety-related" category best predicted the outcome.