Quantitative Transmission Disequilibrium Test Research Articles

Estimating the Genetic Variance of Five Lipid-Relevant Genes for Determining the Plasma Lipid Profiles

Background and Objectives: The plasma lipid levels play crucial roles in the development of atherosclerotic diseases. We estimated the genetic variance of the lipid levels according to the contributions of the single nucleotide polymorphisms (SNPs) and haplotypes in 5 candidate genes. Subjects and Methods: We selected SNPs in the ATP binding cassette A1 (ABCA1) gene, the apolipoprotien A5 (APOA5), apolipoprotien E (APOE) gene, the cholesterol ester transfer protein (CETP) gene and the hepatic triglyceride lipase (LIPC) gene in 383 individuals from 100 Korean families. The genotype was determined by Orchid’s SNP-IT TM technology. The association analysis of the quantitative traits was performed using the quantitative transmission disequilibrium test. Results: A component analysis of the phenotypic variance explained 24.7% of the genetic variance on the total cholesterol, 26.4% of the genetic variance of the high density lipoprotein (HDL)-cholesterol, 11% of the genetic variance of the triglycerides, 35.6% of the genetic variance of the low density lipoprotein (LDL)-cholesterol and 18.9% of the genetic variance of the LDL-C/HDL-C, respectively. The association of the SNPs in the candidate genes explained a major fraction of the genetic phenotypic variance in the LDL-C/HDL-C ratio, but not in the other lipid profiles. The association with SNPs explained 38.5% of the variance for the total cholesterol, 32.2% of the variance for HDLcholesterol and 29.5% of the variance for LDL-cholesterol relative to the polygenic background. An analysis of the contribution of each gene to the genetic variance showed that ABCA1, APOE, CETP and LIPC influenced the variations in total cholesterol, LDL-cholesterol and LDL-C/HDL-C. The variation in HDL-cholesterol was influenced by ABCA1, APOA5 and APOE. Conclusion: We identified that the genetic variance for the total cholesterol, HDL-cholesterol and LDL-cholesterol, and the LDL-C/HDL-C ratio was significantly influenced by the genetic polymorphisms in 5 candidate genes in the Korean population. Further studies are necessary to identify other genes that can explain a major fraction of the genetic variance for the lipid levels. (Korean Circ J 2008;38:197-204)

Genetic Determinants of Susceptibility to Cytarabine Arabinoside.

Cytarabine arabinoside (ara-C) is an essential component of treatment for acute myeloid leukemia (AML) and resistance to this agent is a main reason for treatment failure. To identify genetic factors that are important in susceptibility to the cytotoxic effects of ara-C, we utilized an unbiased whole-genome approach using lymphoblastoid cell lines that are part of the International HapMap Project. We exposed 85 cell lines derived from Utah residents with ancestry from northern and western Europe (CEU) and 89 cell lines derived from individuals of Yoruba in Ibadan, Nigeria (YRI) to a range of concentrations of ara-C (1 – 80 μM) for 72 hours. We used a short-term proliferation assay, alamarBlue®, to measure cell growth inhibition at each concentration of ara-C. In addition, we used the Affymetrix Human Exon 1.0 ST array to determine gene expression of each cell line. We utilized publicly available HapMap (release #22) single nucleotide polymorphisms (SNPs) to perform genome wide association after filtering to remove Mendelian errors and SNPs with mean allele frequencies <5%, leaving approximately 2 million SNPs. We then performed a 3-step analysis to identify genetic signatures associated with gene expression important in susceptibility to ara-C cytotoxicity. The first step involved a Quantitative Transmission Disequilibrium Test (QTDT) association analysis between SNP genotype and percent survival at each concentration of ara-C. Using the SNPs identified in this initial step, we performed a second QTDT analysis between genotype and gene expression of approximately 17,000 transcripts from the exon array to identify SNPs that were associated with the expression of any genes. Finally, we performed a linear regression analysis between the expression of genes identified in this second step and percent survival to identify a subset of these genes whose expression is correlated with susceptibility to ara-C. Using a multivariable backward elimination approach we also determined how much variation in ara-C percent survival is explained by the SNPs identified in step 2. The mean (range) percent survival in the CEU compared to the YRI population was 53.3 (20.8 – 82.8) vs. 47.3 (25.2 – 73.7) at 5μM (p = .0018), 46.8 (18.7–73.2) vs. 39.9 (22.2 – 72.4) at 10μM (p = 0.0001), 40.3 (20.3 – 68.2)vs. 32.8 (17.5 – 53.9) at 40μM (p < 0.0001), and 37.3 (19.1 – 61.3) vs. 30.4 (15.8 – 50.0) (p < 0.0001) at 80μM ara-C. Among the 85 CEU cell lines we identified 177 SNPs for which genotype significantly associated with percent survival at 1μM, using an FDR cutoff of 0.05. Eleven of these 177 SNPs were significantly associated with the expression of a total of 24 genes and all were located trans to the associated gene. The expression of 23 of these genes significantly correlated with percent survival. Of the 11 SNPs that were associated with percent survival and gene expression, 4 SNPs explained 61.2% of the variation in susceptibility to ara-C cytotoxicity among the CEU cell lines. By using an cell-based model and a whole-genome approach that integrates genetic variation, expression variation and sensitivity to drug, we were able to identify a genetic signature for susceptibility to the cytotoxic effects of ara-C. We can now concentrate on the candidate genes identified from this analysis for further functional validation studies.

Association between myostatin gene polymorphisms and peak BMD variation in Chinese nuclear families

We identified 17 polymorphisms in myostatin by sequencing, and three informative single nucleotide polymorphisms (SNPs) were selected for further observation for their association with peak BMD of women in 401 Chinese nuclear families. Our results suggest that genetic polymorphisms in myostatin likely play a role in attainment of peak BMD in Chinese women. Myostatin is a TGF-beta family member that is a negative regulator of skeletal muscle growth. We identified SNPs in myostatin by direct sequencing. Furthermore, using a quantitative transmission disequilibrium test (QTDT). we tested and further test whether SNPs were associated with peak bone mineral density (BMD) variation at the spines and hips of 401 Chinese nuclear families. We identified 17 polymorphisms in myostatin by sequencing. Next, we selected three informative SNPs for further observation of an association with peak BMD of premenopausal women in 401 Chinese nuclear families. Using QTDT for the within-family association, we found significant association between rs2293284 and total hip, femoral neck, and trochanter BMD (all p < 0.05), while rs7570532 was associated with total hip and trochanter BMD (p = 0.034 and p = 0.035, respectively). The within-family association was significant between BMI and +2278G > A (p = 0.022). Subsequent permutations were in agreement with these significant within-family association results. Moreover, analyses of the haplotypes confer further evidence for association of rs2293284 and rs7570532 with hip peak BMD variation. These results suggest, for the first time, the genetic polymorphisms in myostatin likely play a role in attainment of peak BMD in Chinese women.

Case–control and within-family tests for association between 5HTTLPR and conduct problems in a longitudinal adolescent sample

Several recent studies have reported an association between the serotonin transporter 5HTTLPR (s-allele) and aggression; however, non-replications have also been reported. Inconsistencies may be explained by gene-environment interactions. Using a large general population sample, we sought to test for an association between 5HTTLPR and conduct problems, and to explore for a possible 5HTTLPR by maltreatment interaction. Using Caucasian adolescents from the genetic-pairs sample of the National Longitudinal Study of Adolescent Health (n=1,736), we tested for an association between 5HTTLPR and both a categorical and, separately, a continuous measure of conduct problems using regression analyses while controlling for sex, family effects, and age. We then tested for an association between 5HTTLPR and conduct problems using the within-family test Quantitative Transmission Disequilibrium Test. Analyses were repeated for a measure of adolescence-limited delinquency. Results did not support an association between 5HTTLPR and conduct problems or delinquency. The Quantitative Transmission Disequilibrium Test analyses, which account for population stratification, were nonsignificant (F=0.17; P=0.68); introducing maltreatment as a covariate into the model did not affect this association (F=0.17; P=0.68). No association was seen between 5HTTLPR and a measure of adolescence-limited delinquency (F=0.54; P=0.46). Using two methods in a large general population sample we did not find a significant association between 5HTTLPR and conduct problems. A gene by maltreatment interaction was not supported.

AHSG gene polymorphisms are associated with bone mineral density in Caucasian nuclear families

To investigate the role of alpha2-HS glycoprotein (AHSG) gene on bone mineral density (BMD) variation. A total of 665 subjects from 157 Caucasian nuclear families were genotyped at the AHSG NlaIII, SacI sites. The association and linkage between the single SNP markers and haplotypes constructed by two markers in this gene and BMDs at the spine and hip were determined by using quantitative transmission disequilibrium test (QTDT). Significant within-family associations were obtained for spine BMD at both of studied markers (P = 0.036 and 0.005 at the NlaIII and SacI sites, respectively). Significant (P = 0.008 at the NlaIII locus) (P = 0.004 at the SacI locus) total associations at spine BMD were detected. Haplotype analyses confirmed those within-family and total association. These data suggest the polymorphisms in the AHSG gene may have effects on BMD variation in Caucasian population.

Mapping Genes that Contribute to Daunorubicin-Induced Cytotoxicity

Daunorubicin is an anthracycline antibiotic agent used in the treatment of hematopoietic malignancies. Toxicities associated with this agent include myelosuppression and cardiotoxicity; however, the genes or genetic determinants that contribute to these toxicities are unknown. We present an unbiased genome-wide approach that incorporates heritability, whole-genome linkage analysis, and linkage-directed association to uncover genetic variants contributing to the sensitivity to daunorubicin-induced cytotoxicity. Cell growth inhibition in 324 Centre d' Etude du Polymorphisme Humain lymphoblastoid cell lines (24 pedigrees) was evaluated following treatment with daunorubicin for 72 h. Heritability analysis showed a significant genetic component contributing to the cytotoxic phenotypes (h2 = 0.18-0.63 at 0.0125, 0.025, 0.05, 0.1, 0.2, and 1.0 mumol/L daunorubicin and at the IC50, the dose required to inhibit 50% cell growth). Whole-genome linkage scans at all drug concentrations and IC50 uncovered 11 regions with moderate peak LOD scores (> 1.5), including 4q28.2 to 4q32.3 with a maximum LOD score of 3.18. The quantitative transmission disequilibrium tests were done using 31,312 high-frequency single-nucleotide polymorphisms (SNP) located in the 1 LOD confidence interval of these 11 regions. Thirty genes were identified as significantly associated with daunorubicin-induced cytotoxicity (P < or = 2.0 x 10(-4), false discovery rate < or = 0.1). Pathway and functional gene ontology analysis showed that these genes were overrepresented in the phosphatidylinositol signaling system, axon guidance pathway, and GPI-anchored proteins family. Our findings suggest that a proportion of susceptibility to daunorubicin-induced cytotoxicity may be controlled by genetic determinants and that analysis using linkage-directed association studies with dense SNP markers can be used to identify the genetic variants contributing to cytotoxicity.

Allelic variation in GAD1 (GAD67) is associated with schizophrenia and influences cortical function and gene expression

Cortical GABAergic dysfunction has been implicated as a key component of the pathophysiology of schizophrenia and decreased expression of the gamma-aminobutyric acid (GABA) synthetic enzyme glutamic acid decarboxylase 67 (GAD(67)), encoded by GAD1, is found in schizophrenic post-mortem brain. We report evidence of distorted transmission of single-nucleotide polymorphism (SNP) alleles in two independent schizophrenia family-based samples. In both samples, allelic association was dependent on the gender of the affected offspring, and in the Clinical Brain Disorders Branch/National Institute of Mental Health (CBDB/NIMH) sample it was also dependent on catechol-O-methyltransferase (COMT) Val158Met genotype. Quantitative transmission disequilibrium test analyses revealed that variation in GAD1 influenced multiple domains of cognition, including declarative memory, attention and working memory. A 5' flanking SNP affecting cognition in the families was also associated in unrelated healthy individuals with inefficient BOLD functional magnetic resonance imaging activation of dorsal prefrontal cortex (PFC) during a working memory task, a physiologic phenotype associated with schizophrenia and altered cortical inhibition. In addition, a SNP in the 5' untranslated (and predicted promoter) region that also influenced cognition was associated with decreased expression of GAD1 mRNA in the PFC of schizophrenic brain. Finally, we observed evidence of statistical epistasis between two SNPs in COMT and SNPs in GAD1, suggesting a potential biological synergism leading to increased risk. These coincident results implicate GAD1 in the etiology of schizophrenia and suggest that the mechanism involves altered cortical GABA inhibitory activity, perhaps modulated by dopaminergic function.

Association and linkage analysis of COL1A1 and AHSG gene polymorphisms with femoral neck bone geometric parameters in both Caucasian and Chinese nuclear families

To simultaneously investigate the contribution of the alpha 1 chain of collagen type 1 (COL1A1) and alpha2-HS-glycoprotein (AHSG) genes to the variation of bone geometric parameters in both Caucasians and Chinese. Six hundred and five Caucasian individuals from 157 nuclear families and 1228 Chinese subjects from 400 nuclear families were genotyped at the AHSG-SacI, COL1A1- PCOL2 and Sp1 polymorphisms using polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP). 5 FN bone geometric parameters were calculated based on bone mineral density and bone area of femoral neck (FN) measured by dual energy X-ray absorptiometry. Population stratification, total family association, within-family association, and linkage tests were performed by the quantitative transmission disequilibrium test program. The t-test showed the significant differences of all bone geometric phenotypes (except ED) between Caucasians and Chinese in the offspring using both unadjusted and adjusted (by age, height, weight, and gender) data. In Caucasians, we found significant within-family association results between the COL1A1-Sp1 polymorphism (rs1800012) and cross sectional area (CSA), cortical thickness (CT), endocortical diameter (ED), buckling ratio (BR) (P=0.018, 0.002, 0.023, and 0.001, respectively); the COL1A1-Sp1 polymorphism also detected significant linkage with BR (P=0.039). In the population of China, the within-family associations between the COL1A1-PCOL2 polymorphism (rs1107946) and CT, BR were significant (P=0.012 and 0.008, respectively). Furthermore, evidence of linkage were observed between the AHSG-SacI polymorphism (rs4918) and CT, BR (P=0.042 and 0.014, respectively) in Caucasians, but not in Chinese. Our results suggest that the COL1A1 gene may have significantly association with bone geometry in both Caucasians and Chinese, and the AHSG gene may be linked to bone geometry in Caucasians, but not in Chinese. This study represents our first efforts on investigating the importance of the COL1A1 and AHSG genes on bone geometry in both Caucasians and Chinese.

Association between SNP and haplotypes in PPARGC1 and adiponectin genes and bone mineral density in Chinese nuclear families

To assess the contribution of single nucleotide polymorphisms (SNP) and haplotypes in the peroxisome proliferators-activated receptor-gamma co-activator-1 (PPARGC1) and adiponectin genes to normal bone mineral density (BMD) variation in healthy Chinese women and men. We performed population-based (ANOVA) and family-based (quantitative trait locus transmission disequilibrium test) association studies of PPARGC1 and adiponectin genes. SNP in the 2 genes were genotyped. BMD was measured using dual-energy X-ray absorptiometry in the lumbar spine and hip in 401 nuclear families with a total of 1260 subjects, including 458 premenopausal women, 20-40 years of age; 401 postmenopausal women (mothers), 43-74 years of age; and 401 men (fathers), 49-76 years of age. Significant within-family association was found between the Thr394Thr polymorphism in the PPGAGC1 gene and peak BMD in the femoral neck (P=0.026). Subsequent permutations were in agreement with this significant within-family association result (P=0.016), but Thr394Thr SNP only accounted for 0.7% of the variation in femoral neck peak BMD. However, no significant within-family association was detected between each SNP in the adiponectin gene and peak BMD. Although no significant association was found between BMD and SNP in the PPARGC1 and adiponectin genes in both men and postmenopausal women, haplotype 2 (T-T) in the adiponectin gene was associated with lumbar spine BMD in postmenopausal women (P=0.019). Our findings suggest that Thr394Thr SNP in the PPARGC1 gene was associated with peak BMD in the femoral neck in Chinese women. Confirmation of our results is needed in other populations and with more functional markers within and flanking the PPARGC1 or adiponectin genes region.

Evidence for epistasis between SLC6A4 and ITGB3 in autism etiology and in the determination of platelet serotonin levels

Autism is a neurodevelopmental disorder of unclear etiology. The consistent finding of platelet hyperserotonemia in a proportion of patients and its heritability within affected families suggest that genes involved in the serotonin system play a role in this disorder. The role in autism etiology of seven candidate genes in the serotonin metabolic and neurotransmission pathways and mapping to autism linkage regions (SLC6A4, HTR1A, HTR1D, HTR2A, HTR5A, TPH1 and ITGB3) was analyzed in a sample of 186 nuclear families. The impact of interactions among these genes in autism was assessed using the multifactor-dimensionality reduction (MDR) method in 186 patients and 181 controls. We further evaluated whether the effect of specific gene variants or gene interactions associated with autism etiology might be mediated by their influence on serotonin levels, using the quantitative transmission disequilibrium test (QTDT) and the restricted partition method (RPM), in a sample of 109 autistic children. We report a significant main effect of the HTR5A gene in autism (P = 0.0088), and a significant three-locus model comprising a synergistic interaction between the ITGB3 and SLC6A4 genes with an additive effect of HTR5A (P < 0.0010). In addition to the previously reported contribution of SLC6A4, we found significant associations of ITGB3 haplotypes with serotonin level distribution (P = 0.0163). The most significant models contributing to serotonin distribution were found for interactions between TPH1 rs4537731 and SLC6A4 haplotypes (P = 0.002) and between HTR1D rs6300 and SLC6A4 haplotypes (P = 0.013). In addition to the significant independent effects, evidence for interaction between SLC6A4 and ITGB3 markers was also found. The overall results implicate SLC6A4 and ITGB3 gene interactions in autism etiology and in serotonin level determination, providing evidence for a common underlying genetic mechanism and a molecular explanation for the association of platelet hyperserotonemia with autism.

The chemokine (C-C-motif) receptor 3 (CCR3) gene is linked and associated with age at menarche in Caucasian females

Chemokine (C-C-motif) receptor 3 (CCR3), playing an important role in endometrium related metabolic pathways, may influence the onset of menarche. To test linkage and/or association between CCR3 polymorphisms with the variation of age at menarche (AAM) in Caucasian females, we recruited a sample of 1,048 females from 354 Caucasian nuclear families and genotyped 16 SNPs spanning the entire CCR3 gene. Linkage disequilibrium and haplotype blocks were inferred by Haploview. Both single-SNP markers and haplotypes were tested for linkage and/or association with AAM using QTDT (quantitative transmission disequilibrium test). We also tested associations between CCR3 polymorphisms and AAM in a selected random sample of daughters using ANOVA (analysis of variance). We identified two haplotype blocks. Only block two showed significant results. After correction for multiple testing, significant total associations of SNP7, SNP9 with AAM were detected (P = 0.009 and 0.006, respectively). We also detected significant within-family association of SNP9 (P = 0.01). SNP14 was linked to AAM (P = 0.02) at the nominal level. In addition, there was evidence of significant total association and nominal significant linkage (P = 0.008 and 0.03, respectively) with AAM for the haplotype AGA reconstructed by SNP7, SNP9 and SNP13. ANOVA confirmed the results by QTDT. For the first time we reported that CCR3 is linked and associated with AAM variation in Caucasian women. However, further studies are necessary to substantiate our conclusions.

Neuregulin 1 (8p12) and childhood-onset schizophrenia: susceptibility haplotypes for diagnosis and brain developmental trajectories

Childhood-onset schizophrenia (COS), defined as onset of psychosis by the age of 12, is a rare and malignant form of the illness, which may have more salient genetic influence. Since the initial report of association between neuregulin 1 (NRG1) and schizophrenia in 2002, numerous independent replications have been reported. In the current study, we genotyped 56 markers (54 single-nucleotide polymorphisms (SNPs) and two microsatellites) spanning the NRG1 locus on 78 COS patients and their parents. We used family-based association analysis for both diagnostic (extended transmission disequilibrium test) and quantitative phenotypes (quantitative transmission disequilibrium test) and mixed-model regression. Most subjects had prospective anatomic brain magnetic resonance imaging (MRI) scans at 2-year intervals. Further, we genotyped a sample of 165 healthy controls in the MRI study to examine genetic risk effects on normal brain development. Individual markers showed overtransmission of alleles to affecteds (P=0.009-0.05). Further, several novel four-marker haplotypes demonstrated significant transmission distortion. There was no evidence of epistasis with SNPs in erbB4. The risk allele (0) at 420M9-1395 was associated with poorer premorbid social functioning. Further, possession of the risk allele was associated with different trajectories of change in lobar volumes. In the COS group, risk allele carriers had greater total gray and white matter volume in childhood and a steeper rate of subsequent decline in volume into adolescence. By contrast, in healthy children, possession of the risk allele was associated with different trajectories in gray matter only and was confined to frontotemporal regions, reflecting epistatic or other illness-specific effects mediating NRG1 influence on brain development in COS. This replication further documents the role of NRG1 in the abnormal brain development in schizophrenia. This is the first demonstration of a disease-specific pattern of gene action in schizophrenia.

The Human Calcium-Sensing Receptor and Interleukin-6 Genes are Associated with Bone Mineral Density in Chinese

Calcium sensing receptor (CASR) is a central factor involved in calcium metabolism. Interleukin-6 (IL-6) is a pleiotropic cytokine that plays an important role in osteoclast differentiation. Thus, both CASR and IL-6 are important in bone and mineral metabolism and are prominent candidate genes for osteoporosis. The study aimed to test association and/or linkage between the CASR and IL-6 genes with bone mineral density (BMD) variation in a Chinese population. A cytosine-adenine ( CA) n repeat polymorphism in the CASR gene and the IL-6 gene was genotyped, respectively, in 1 263 subjects from 402 Chinese nuclear families. Employing tests implemented in the program QTDT (quantitative transmission disequilibrium tests), a significant total association of the CASR (CA) 12 allele ( P = 0.006) and (CA) 18 allele ( P = 0.02) with BMD at the femoral neck was found. For the IL-6 gene, significant within-family associations were found between the ( CA) 14 allele and BMD at the total hip ( P = 0.021), the femoral neck ( P = 0.041), and the intertrochanteric region ( P = 0.029). A significant linkage was also observed between IL-6 CA repeat polymorphism and BMD at the spine ( P = 0.001). The results suggest that the CASR gene and the IL-6 gene may have effects on BMD variation in Chinese.

Linkage and Association Between CA Repeat Polymorphism of the TNFR2 Gene and Obesity Phenotypes in Two Independent Caucasian Populations

Previously, our group has reported a suggestive linkage evidence of 1p36 with body mass index (BMI) ( LOD = 2.09). The tumor necrosis factor receptor 2 ( TNFR2) at 1p36 is an excellent positional and functional candidate gene for obesity. In this study, we have investigated the linkage and association between the TNFR2 gene and obesity phenotypes in two large independent samples, using the quantitative transmission disequilibrium tests (QTDT). The first group was made up of 1 836 individuals from 79 multi-generation pedigrees. The second group was a randomly ascertained set of 636 individuals from 157 US Caucasian nuclear families. Obesity phenotypes tested include BMI, fat mass, and percentage fat mass (PFM). A significant result ( P = 0.0056) was observed for linkage with BMI in the sample of the multigenerational pedigrees. Our data support the TNFR2 gene as a quantitative trait locus (QTL) underlying BMI variation in the Caucasian populations.

A Quantitative Trait Locus for Body Fat on Chromosome 1q43 in French Canadians: Linkage and Association Studies

To explore a quantitative trait locus (QTL) on human chromosome 1q affecting BMI, adiposity, and fat-free mass phenotypes in the Quebec Family Study cohort. Non-parametric sibpair and variance component linkage analyses and family-based association studies were performed with a dense set of chromosome 1q43 microsatellites and single-nucleotide polymorphism markers in 885 adult individuals. Linkage was observed between marker D1S184 and BMI (p = 0.0004) and with body fat mass or percentage body fat (p < or = 0.0003), but no linkage was detected with fat-free mass. Furthermore, significant linkages (p < 0.0001) were achieved with subsamples of sibpairs at both ends of phenotype distributions. Association studies with quantitative transmission disequilibrium tests refined the linkage to a region overlapping the regulator of G-protein signaling 7 (RGS7) gene and extending to immediate upstream gene loci. The present study indicates that the QTL on chromosome 1q43 specifically affects total adiposity and provides a genetic mapping framework for the dissection of this adiposity locus.

24-h ambulatory blood pressure is linked to chromosome 18q21–22 and genetic variation of NEDD4L associates with cross-sectional and longitudinal blood pressure in Swedes

Numerous linkage studies have indicated chromosome 18q21-22 as a locus of importance for blood pressure regulation. This locus harbors the neural precursor cell expressed developmentally downregulated 4-like (NEDD4L) gene, which is instrumental for the regulation of the amiloride-sensitive epithelial sodium channel (ENaC). In a linkage study of 16 markers (including two single nucleotide polymorphism markers located within the NEDD4L gene) on chromosome 18 between 70-104 cM and ambulatory blood pressure (ABP), in 118 families, the strongest evidence of linkage was found for 24 h and day-time systolic ABP at the NEDD4L locus (82.25 cM) (P=0.0014). In a large population sample (n=4001), we subsequently showed that a NEDD4L gene variant (rs4149601), which by alternative splicing leads to varying expression of a functionally crucial C2 domain, was associated with diastolic blood pressure (DBP) (P=0.03) and DBP progression over time (P=0.04). A genotype combination of the rs4149601 and an intronic NEDD4L marker (rs2288774) was associated with systolic blood pressure (SBP) (P=0.01), DBP (P=0.04), and progression of both SBP (P=0.03) and DBP (P=0.05) over time. A quantitative transmission disequilibrium test in the family material of the rs4149601 supported this NEDD4L variant as being at least partially causative of the linkage result. In conclusion, our findings suggest that the chromosome 18 linkage peak at 82.25 cM is explained by genetic NEDD4L variation affecting cross-sectional and longitudinal blood pressure, possibly as a consequence of altered NEDD4L interaction with ENaC.

Effect of the BDNF V166M polymorphism on working memory in healthy adolescents

Brain-derived neurotrophic factor (BDNF) may play a role in modulating memory function and there is growing evidence that the BDNF V166M polymorphism may influence episodic memory in humans. However, previous association studies examining this polymorphism and working memory are inconsistent. The current study examined this association in a large sample of adolescent twin-pairs and siblings (785 individuals from 439 families). A range of measures (event-related potential, general performance and reaction time) was obtained from a delayed-response working-memory task and total association was examined using the quantitative transmission disequilibrium tests (QTDT) program. Analyses had approximately 93-97% power (alpha= 0.05) to detect an association accounting for as little as 2% of the variance in the phenotypes examined. Results indicated that the BDNF V166M polymorphism is not associated with variation in working memory in healthy adolescents.

NOS2A and the modulating effect of cigarette smoking in Parkinson's disease

Inducible nitric oxide synthase, a protein product of NOS2A, generates nitric oxide as a defense mechanism, but excessive levels threaten cellular survival. NOS2A is a candidate gene for Parkinson's disease (PD) that potentially interacts with cigarette smoking. We examined NOS2A for association with PD risk and age at onset (AAO) and for interaction with smoking. We genotyped 13 NOS2A single nucleotide polymorphisms (SNPs) in 466 singleton families and in a validation set of 286 multiplex families. We tested allelic and haplotypic association using the association in the presence of linkage test, genotypic associations using the genotype pedigree disequilibrium test, AAO effects using the quantitative transmission disequilibrium test, and interactions using generalized estimating equations. Among the pooled earliest onset families, rs2255929 and rs1060826 generated significant allelic (p = 0.000059 and 0.0062, respectively) and genotypic (p = 0.0039 and 0.0014, respectively) associations with risk and AAO (p = 0.00070 and 0.0073, respectively); the two-SNP haplotype generated even stronger association with PD (p = 0.000013). Significant interactions with smoking (p = 0.0015 for rs 2255929 and p < 0.0001 for rs 1060826) were detected in a subset of the families; smoking was inversely associated with PD among risk allele noncarriers, but significance diminished among carriers. Our findings support NOS2A as a genetic risk factor in PD, potentially by influencing AAO and by modifying the inverse association between PD and smoking.

Association analyses of CYP19 gene polymorphisms with height variation in a large sample of Caucasian nuclear families

Human height is a complex trait regulated by multiple genetic and environmental factors. CYP19 (cytochrome P450 19) encodes aromatase, which catalyses the rate-limiting step in the conversion of androgens to estrogens. Deleterious mutations in CYP19 can result in estrogen deficiency that will influence adult height to certain extent. In the present study, we aimed to test the associations between the CYP19 gene polymorphisms with adult height variation, using family-based association methods, such as QTDT (quantitative transmission disequilibrium test) and FBAT (family-based association test) in 1,873 subjects from 405 Caucasian nuclear families. We found one SNP (rs730154) significantly associated with height by both QTDT (P=0.0030) and FBAT (P=0.0016) analyses. Haplotype analyses corroborated our single-marker results by showing that the haplotypes in block 4 containing rs730154 were significantly associated with height variation. We thus concluded that CYP19 could be one of the genetic factors influencing adult height in Caucasians. Further studies are required to identify the causal functional variants responsible for Caucasian height within the CYP19 gene.

Assessment of linkage and association of 13 genetic loci with bone mineral density

Bone mineral density (BMD), an important risk factor for osteoporosis, is a complex trait likely affected by multiple genes. The linkage and/or association of 13 polymorphic loci of seven candidate genes (estrogen receptor alpha [ERalpha] and beta [ERbeta], calcium-sensing receptor, vitamin D receptor, collagen type 1alpha1, low-density lipoprotein [LDL] receptor-related protein 5 [LRPS], and transforming growth factor beta1) were evaluated in 177 southern Chinese pedigrees of 674 subjects, with each pedigree identified through a proband having a BMD Z score of -1.28 or less at the hip or spine. A suggestive linkage was detected between the IVS1-351A/G polymorphism of ERalpha and spine BMD, and between the 1082G/A, 1730G/A, and D14S1026 polymorphisms of ERbeta and BMD at both spine and hip. The quantitative transmission disequilibrium test (QTDT) detected total family association between 1730G/A of ERbeta and BMD at spine and hip; between D14S1026 of ERbeta and hip BMD; and between the 266A/G and 2220C/T polymorphisms of LRP5 and hip BMD. Similar total family associations were detected when only the females were analyzed. In addition, the IVS1-397T/C polymorphism of ERalpha was associated with spine BMD, and the 266A/G and 2220C/T polymorphisms of LRP5 were associated with femoral neck BMD in the females. A within-family association was detected with the IVS1-397T/C polymorphism of ERalpha, and the 266A/G and 2220C/T polymorphisms of LRP5 in the females. The effect of each polymorphism on BMD variance ranged from 1% to 4%. In conclusion, ERalpha, ERbeta and LRP5 are important candidate genes determining BMD variation, especially in females.