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Abstract Genome-wide association studies (GWAS) have become well-powered to detect loci associated with telomere length. However, no prior work has validated genes nominated by GWAS to examine their role in telomere length regulation. We conducted a multi-ancestry meta-analysis of 211,369 individuals and identified five novel association signals. Enrichment analyses of chromatin state and cell-type heritability suggested that blood/immune cells are the most relevant cell type to examine telomere length association signals. We validated specific GWAS associations by overexpressing KBTBD6 or POP5 and demonstrated that both lengthened telomeres. CRISPR/Cas9 deletion of the predicted causal regions in K562 blood cells reduced expression of these genes, demonstrating that these loci are related to transcriptional regulation of KBTBD6 and POP5. Our results demonstrate the utility of telomere length GWAS in the identification of telomere length regulation mechanisms and validate KBTBD6 and POP5 as genes affecting telomere length regulation.

Introduction: Educational attainment, widely used in epidemiologic studies as a surrogate for socioeconomic status, is a predictor of cardiovascular health outcomes.Methods: A two-stage genome-wide meta-analysis of low-density lipoprotein cholesterol (LDL), high-density lipoprotein cholesterol (HDL), and triglyceride (TG) levels was performed while accounting for gene-educational attainment interactions in up to 226,315 individuals from five population groups. We considered two educational attainment variables: “Some College” (yes/no, for any education beyond high school) and “Graduated College” (yes/no, for completing a 4-year college degree). Genome-wide significant (p < 5 × 10−8) and suggestive (p < 1 × 10−6) variants were identified in Stage 1 (in up to 108,784 individuals) through genome-wide analysis, and those variants were followed up in Stage 2 studies (in up to 117,531 individuals).Results: In combined analysis of Stages 1 and 2, we identified 18 novel lipid loci (nine for LDL, seven for HDL, and two for TG) by two degree-of-freedom (2 DF) joint tests of main and interaction effects. Four loci showed significant interaction with educational attainment. Two loci were significant only in cross-population analyses. Several loci include genes with known or suggested roles in adipose (FOXP1, MBOAT4, SKP2, STIM1, STX4), brain (BRI3, FILIP1, FOXP1, LINC00290, LMTK2, MBOAT4, MYO6, SENP6, SRGAP3, STIM1, TMEM167A, TMEM30A), and liver (BRI3, FOXP1) biology, highlighting the potential importance of brain-adipose-liver communication in the regulation of lipid metabolism. An investigation of the potential druggability of genes in identified loci resulted in five gene targets shown to interact with drugs approved by the Food and Drug Administration, including genes with roles in adipose and brain tissue.Discussion: Genome-wide interaction analysis of educational attainment identified novel lipid loci not previously detected by analyses limited to main genetic effects.

Summary: Background: Identifying novel epigenetic signatures associated with serum immunoglobulin E (IgE) may improve our understanding of molecular mechanisms underlying asthma and IgE-mediated diseases. Methods: We performed an epigenome-wide association study using whole blood from Framingham Heart Study (FHS; n = 3,471, 46% females) participants and validated results using the Childhood Asthma Management Program (CAMP; n = 674, 39% females) and the Genetic Epidemiology of Asthma in Costa Rica Study (CRA; n = 787, 41% females). Using the closest gene to each IgE-associated CpG, we highlighted biologically plausible pathways underlying IgE regulation and analyzed the transcription patterns linked to IgE-associated CpGs (expression quantitative trait methylation loci; eQTMs). Using prior UK Biobank summary data from genome-wide association studies of asthma and allergy, we performed Mendelian randomization (MR) for causal inference testing using the IgE-associated CpGs from FHS with methylation quantitative trait loci (mQTLs) as instrumental variables. Findings: We identified 490 statistically significant differentially methylated CpGs associated with IgE in FHS, of which 193 (39.3%) replicated in CAMP and CRA (FDR < 0.05). Gene ontology analysis revealed enrichment in pathways related to transcription factor binding, asthma, and other immunological processes. eQTM analysis identified 124 cis-eQTMs for 106 expressed genes (FDR < 0.05). MR in combination with drug-target analysis revealed CTSB and USP20 as putatively causal regulators of IgE levels (Bonferroni adjusted P

Although the common genetic variants contributing to blood lipid levels have been studied, the contribution of rare variants is less understood. Here, the authors perform a rare coding and noncoding variant association study of blood lipid levels using whole genome sequencing data.

Abstract Objective Obesity in pregnancy and gestational diabetes (GDM) increase cardiometabolic disease risk but are difficult to disentangle. This study aimed to test the hypothesis that 4–10 years after a pregnancy complicated by overweight/obesity and GDM (OB‐GDM), women and children would have greater adiposity and poorer cardiometabolic health than those with overweight/obesity (OB) or normal weight (NW) and no GDM during the index pregnancy. Methods In this cross‐sectional study, mother‐child dyads were stratified into three groups based on maternal health status during pregnancy (OB‐GDM = 67; OB = 76; NW = 76). Weight, height, waist and hip circumferences, and blood pressure were measured, along with fasting glucose, insulin, HbA1c, lipids, adipokines, and cytokines. Results Women in the OB and OB‐GDM groups had greater current adiposity and poorer cardiometabolic health outcomes than those in the NW group (p

This study of 23,000 non-diabetic individuals highlights loci associated with fasting glucose and fasting insulin in diverse cohorts with whole genome sequence data.

A GWAS of 355 lipid species in the Old Order Amish founder population reveals associations between Amish-enriched loci and several sphingolipids.

Summary: Anthropometric traits, measuring body size and shape, are highly heritable and significant clinical risk factors for cardiometabolic disorders. These traits have been extensively studied in genome-wide association studies (GWASs), with hundreds of genome-wide significant loci identified. We performed a whole-exome sequence analysis of the genetics of height, body mass index (BMI) and waist/hip ratio (WHR). We meta-analyzed single-variant and gene-based associations of whole-exome sequence variation with height, BMI, and WHR in up to 22,004 individuals, and we assessed replication of our findings in up to 16,418 individuals from 10 independent cohorts from Trans-Omics for Precision Medicine (TOPMed). We identified four trait associations with single-nucleotide variants (SNVs; two for height and two for BMI) and replicated the LECT2 gene association with height. Our expression quantitative trait locus (eQTL) analysis within previously reported GWAS loci implicated CEP63 and RFT1 as potential functional genes for known height loci. We further assessed enrichment of SNVs, which were monogenic or syndromic variants within loci associated with our three traits. This led to the significant enrichment results for height, whereas we observed no Bonferroni-corrected significance for all SNVs. With a sample size of ∼20,000 whole-exome sequences in our discovery dataset, our findings demonstrate the importance of genomic sequencing in genetic association studies, yet they also illustrate the challenges in identifying effects of rare genetic variants.

The influence of X chromosome genetic variation on blood lipids and coronary heart disease (CHD) is not well understood. Here, the authors analyse X chromosome sequencing data across 65,322 multi-ancestry individuals, identifying associations of the Xq23 locus with lipid changes and reduced risk of CHD and diabetes mellitus.

A substantial proportion of the adult United States population with type 2 diabetes (T2D) are undiagnosed, calling into question the comprehensiveness of current screening practices, which primarily rely on age, family history, and body mass index (BMI). We hypothesized that a polygenic score (PGS) may serve as a complementary tool to identify high-risk individuals. The T2D polygenic score maintained predictive utility after adjusting for family history and combining genetics with family history led to even more improved disease risk prediction. We observed that the PGS was meaningfully related to age of onset with implications for screening practices: there was a linear and statistically significant relationship between the PGS and T2D onset (−1.3 years per standard deviation of the PGS). Evaluation of U.S. Preventive Task Force and a simplified version of American Diabetes Association screening guidelines showed that addition of a screening criterion for those above the 90th percentile of the PGS provided a small increase the sensitivity of the screening algorithm. Among T2D-negative individuals, the T2D PGS was associated with prediabetes, where each standard deviation increase of the PGS was associated with a 23% increase in the odds of prediabetes diagnosis. Additionally, each standard deviation increase in the PGS corresponded to a 43% increase in the odds of incident T2D at one-year follow-up. Using complications and forms of clinical intervention (i.e., lifestyle modification, metformin treatment, or insulin treatment) as proxies for advanced illness we also found statistically significant associations between the T2D PGS and insulin treatment and diabetic neuropathy. Importantly, we were able to replicate many findings in a Hispanic/Latino cohort from our database, highlighting the value of the T2D PGS as a clinical tool for individuals with ancestry other than European. In this group, the T2D PGS provided additional disease risk information beyond that offered by traditional screening methodologies. The T2D PGS also had predictive value for the age of onset and for prediabetes among T2D-negative Hispanic/Latino participants. These findings strengthen the notion that a T2D PGS could play a role in the clinical setting across multiple ancestries, potentially improving T2D screening practices, risk stratification, and disease management.

The genetic basis of daytime napping and the directional effect of daytime napping on cardiometabolic health are unknown. Here, the authors perform a genome-wide association study on self-reported daytime napping in the UK Biobank and Mendelian randomization to explore causal associations.

Drugs targeting cardiovascular disease (CVD) can have negative consequences for liver function. Here, the authors combine genome wide analyses on 69,479 individuals to identify loss-of-function variants with beneficial effects on CVD-related traits without negative impacts on liver function.

Summary: Genetic studies on telomere length are important for understanding age-related diseases. Prior GWASs for leukocyte TL have been limited to European and Asian populations. Here, we report the first sequencing-based association study for TL across ancestrally diverse individuals (European, African, Asian, and Hispanic/Latino) from the NHLBI Trans-Omics for Precision Medicine (TOPMed) program. We used whole-genome sequencing (WGS) of whole blood for variant genotype calling and the bioinformatic estimation of telomere length in n = 109,122 individuals. We identified 59 sentinel variants (p < 5 × 10−9) in 36 loci associated with telomere length, including 20 newly associated loci (13 were replicated in external datasets). There was little evidence of effect size heterogeneity across populations. Fine-mapping at OBFC1 indicated that the independent signals colocalized with cell-type-specific eQTLs for OBFC1 (STN1). Using a multi-variant gene-based approach, we identified two genes newly implicated in telomere length, DCLRE1B (SNM1B) and PARN. In PheWAS, we demonstrated that our TL polygenic trait scores (PTSs) were associated with an increased risk of cancer-related phenotypes.

Abstract Background The lipoprotein insulin resistance (LPIR) score was shown to predict insulin resistance (IR) and type 2 diabetes (T2D) in healthy adults. However, the molecular basis underlying the LPIR utility for classification remains unclear. Objective To identify small molecule lipids associated with variation in the LPIR score, a weighted index of lipoproteins measured by nuclear magnetic resonance, in the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) study (n = 980). Methods Linear mixed effects models were used to test the association between the LPIR score and 413 lipid species and their principal component analysis-derived groups. Significant associations were tested for replication with homeostatic model assessment-IR (HOMA-IR), a phenotype correlated with the LPIR score (r = 0.48, p

Abstract Background The southeastern US is an epicenter for incident HIV in the US with high prevalence of human papillomavirus (HPV) co-infections. However, epidemiologies of HPV-associated clinical conditions (CC) among people living with HIV-1 infection (PLWH) are not fully known. Methods Electronic medical records (EMR) of PLWH attending one of the leading HIV clinics in the southeastern US between 2006 and 2018 were reviewed and analyzed. The retrospective study was nested within the University of Alabama at Birmingham HIV clinical cohort, which has electronically collected over 7000 PLWH’s clinical and sociobehavioral data since 1999. Incidence rates of HPV-related CC including anogenital warts, penile, anal, cervical, and vaginal/vulvar low- and high-grade squamous intraepithelial lesions (LSIL and HSIL) were estimated per 10,000 person years. Joinpoint regressions were performed to examine temporal changes in the trends of incident CC. All rates and trends were stratified by gender and race. Results Of the 4484 PLWH included in the study (3429 men, 1031 women, and 24 transgender), we observed 1038 patients with HPV-related CC. The median nadir CD4 count (cells/uL) was higher in the HPV-condition free group than the case groups (P

Sleep duration is associated with an adverse lipid profile. Here, the authors perform genome-wide gene-by-sleep interaction analysis and find 49 previously unreported lipid loci when considering short or long total sleep time.

Abstract Background Obesity, widely recognized as a serious health concern, is characterized by profoundly altered metabolism. However, the intermediate metabolites involved in this change remain largely unknown. Objective: We conducted targeted metabolomics profiling to identify moieties associated with adult obesity. Methods In this case-control study of Iranian adults, 200 obese patients were compared with 100 controls based on 104 metabolites profiled by a targeted metabolomic approach using liquid chromatography coupled to triple quadrupole mass spectrometry (LC-MS/MS). The analysis comprised acylcarnitines, diacyl-phosphatidylcholines (PCaa), acyl-alkyl-phosphatidylcholines (PCae), sphingomyelins (SM), lyso-phospholipids (LPC) and amino acids. We performed multivariable linear regression to identify metabolites associated with obesity, adjusting for age, sex, total energy intake, total physical activity, smoking, and alcohol consumption. The Bonferroni correction was used to adjust for multiple testing. Results A pattern of 19 metabolites was significantly associated with obesity. Branched chain amino acids, alanine, glutamic acid, proline, tyrosine LPCa C16:1, PCaa C32:1, PCaa C32:2 and PCaa C38:3 were positively, while serine, asparagine, LPCa C18:1, LPCa C18:2, LPCe C18:0, PCae C34:3, PCae C38:4 and PCae C40:6 were negatively associated with obesity (all p

Aim: Significant knowledge gaps exist regarding lipoprotein profiles in children with type 2 diabetes mellitus (T2DM). The primary objective was to analyze the type and nature of lipoprotein abnormalities present in children with T2DM and to identify determinants of adverse lipoprotein profiles. The secondary objective was to assess associations with elevated glycated hemoglobin (HbA1C), i.e., 8% had significantly higher total cholesterol (191.4 vs. 158.1 mg/dL, P = 0.0004), LDL-C (117.77 vs. 92.3 mg/dL, P = 0.002), apoB (99.5 vs. 80.9 mg/dL, P = 0.002), non-HDL-C (141.5 vs. 112.5, P = 0.002), and frequency of LDL pattern B (57% vs. 20%, P = 0.0008). Conclusion: HbA1C and BMI were associated with adverse lipoprotein profiles, and may represent two major modifiable cardiovascular risk factors in the pediatric T2DM population. Patients with an HbA1C higher than 8.0% had significantly worse atherogenic lipid profile, i.e., higher LDL-C, non-HDL-C, apoB and LDL pattern B, suggesting adequate glycemia may improve adverse lipoprotein profiles. Keywords: Type 2 diabetes mellitus, Lipoproteins, Cardiovascular risk, Dyslipidemia

GWAS have identified more than 500 genetic loci associated with blood lipid levels. Here, the authors report a genome-wide analysis of interactions between genetic markers and physical activity, and find that physical activity modifies the effects of four genetic loci on HDL or LDL cholesterol.

Abstract GAW20 provided participants with an opportunity to comprehensively examine genetic and epigenetic variation among related individuals in the context of drug treatment response. GAW20 used data from 188 families (N = 1105) participating in the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) study (clinicaltrials.gov identifier NCT00083369), which included CD4+ T-cell DNA methylation at 463,995 cytosine-phosphate-guanine (CpG) sites measured before and after a 3-week treatment with fenofibrate, single-nucleotide variation at 906,600 loci, metabolic syndrome components ascertained before and after the drug intervention, and relevant covariates. All GOLDN participants were of European descent, with an average age of 48 years. In addition, approximately half were women and approximately 40% met the diagnostic criteria for metabolic syndrome. Unique advantages of the GAW20data set included longitudinal (3 weeks apart) measurements of DNA methylation, the opportunity to explore the contributions of both genotype and DNA methylation to the interindividual variability in drug treatment response, and the familial relationships between study participants. The principal disadvantage of GAW20/GOLDN data was the spurious correlation between batch effects and fenofibrate effects on methylation, which arose because the pre- and posttreatment methylation data were generated and normalized separately, and any attempts to remove time-dependent technical artifacts would also remove biologically meaningful changes brought on by fenofibrate. Despite this limitation, the GAW20 data set offered informative, multilayered omics data collected in a large population-based study of common disease traits, which resulted in creative approaches to integration and analysis of inherited human variation.

Abstract Background Left ventricular (LV) hypertrophy affects up to 43% of African Americans (AAs). Antihypertensive treatment reduces LV mass (LVM). However, interindividual variation in LV traits in response to antihypertensive treatments exists. We hypothesized that genetic variants may modify the association of antihypertensive treatment class with LV traits measured by echocardiography. Methods We evaluated the main effects of the three most common antihypertensive treatments for AAs as well as the single nucleotide polymorphism (SNP)‐by‐drug interaction on LVM and relative wall thickness (RWT) in 2,068 participants across five community‐based cohorts. Treatments included thiazide diuretics (TDs), angiotensin converting enzyme inhibitors (ACE‐Is), and dihydropyridine calcium channel blockers (dCCBs) and were compared in a pairwise manner. We performed fixed effects inverse variance weighted meta‐analyses of main effects of drugs and 2.5 million SNP‐by‐drug interaction estimates. Results We observed that dCCBs versus TDs were associated with higher LVM after adjusting for covariates (p = 0.001). We report three SNPs at a single locus on chromosome 20 that modified the association between RWT and treatment when comparing dCCBs to ACE‐Is with consistent effects across cohorts (smallest p = 4.7 × 10−8, minor allele frequency range 0.09–0.12). This locus has been linked to LV hypertrophy in a previous study. A marginally significant locus in BICD1 (rs326641) was validated in an external population. Conclusions Our study identified one locus having genome‐wide significant SNP‐by‐drug interaction effect on RWT among dCCB users in comparison to ACE‐I users. Upon additional validation in future studies, our findings can enhance the precision of medical approaches in hypertension treatment.

Our understanding of genetic influences on the response of lipids to specific interventions is limited. In this study, we sought to elucidate effects of rare genetic variants on lipid response to a high-fat meal challenge and fenofibrate (FFB) therapy in the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) cohort using an exome-wide sequencing-based association study. Our results showed that the rare coding variants in ITGA7, SIPA1L2, and CEP72 are significantly associated with fasting LDL cholesterol response to FFB (P = 1.24E-07), triglyceride postprandial area under the increase (AUI) (P = 2.31E-06), and triglyceride postprandial AUI response to FFB (P = 1.88E-06), respectively. We sought to replicate the association for SIPA1L2 in the Heredity and Phenotype Intervention (HAPI) Heart Study, which included a high-fat meal challenge but not FFB treatment. The associated rare variants in GOLDN were not observed in the HAPI Heart study, and thus the gene-based result was not replicated. For functional validation, we found that gene transcript level of SIPA1L2 is associated with triglyceride postprandial AUI (P < 0.05) in GOLDN. Our study suggests unique genetic mechanisms contributing to the lipid response to the high-fat meal challenge and FFB therapy.

Abstract Background The high prevalence of obesity among US adults has resulted in significant increases in associated metabolic disorders such as diabetes, dyslipidemia, and high blood pressure. Together, these disorders constitute metabolic syndrome, a clinically defined condition highly prevalent among African-Americans. Identifying epigenetic alterations associated with metabolic syndrome may provide additional information regarding etiology beyond current evidence from genome-wide association studies. Methods Data on metabolic syndrome and DNA methylation was assessed on 614 African-Americans from the Hypertension Genetic Epidemiology Network (HyperGEN) study. Metabolic syndrome was defined using the joint harmonized criteria, and DNA methylation was assessed using the Illumina HumanMethylation450K Bead Chip assay on DNA extracted from buffy coat. Linear mixed effects regression models were used to examine the association between CpG methylation at > 450,000 CpG sites and metabolic syndrome adjusted for study covariates. Replication using DNA from a separate sample of 69 African-Americans, as well as meta-analysis combining both cohorts, was conducted. Results Two differentially methylated CpG sites in the IGF2BP1 gene on chromosome 17 (cg06638433; p value = 3.10 × 10− 7) and the ABCG1 gene on chromosome 21 (cg06500161; p value = 2.60 × 10− 8) were identified. Results for the ABCG1 gene remained statistically significant in the replication dataset and meta-analysis. Conclusion Metabolic syndrome was consistently associated with increased methylation in the ABCG1 gene in the discovery and replication datasets, a gene that encodes a protein in the ATP-binding cassette transporter family and is involved in intra- and extra-cellular signaling and lipid transport.

Abstract Background Recently, epigenetic age acceleration—or older epigenetic age in comparison to chronological age—has been robustly associated with mortality and various morbidities. However, accelerated epigenetic aging has not been widely investigated in relation to inflammatory or metabolic markers, including postprandial lipids. Methods We estimated measures of epigenetic age acceleration in 830 Caucasian participants from the Genetics Of Lipid Lowering Drugs and diet Network (GOLDN) considering two epigenetic age calculations based on differing sets of 5′-Cytosine-phosphate-guanine-3′ genomic site, derived from the Horvath and Hannum DNA methylation age calculators, respectively. GOLDN participants underwent a standardized high-fat meal challenge after fasting for at least 8 h followed by timed blood draws, the last being 6 h postmeal. We used adjusted linear mixed models to examine the association of the epigenetic age acceleration estimate with fasting and postprandial (0- and 6-h time points) low-density lipoprotein (LDL), high-density lipoprotein (HDL), and triglyceride (TG) levels as well as five fasting inflammatory markers plus adiponectin. Results Both DNA methylation age estimates were highly correlated with chronological age (r > 0.90). We found that the Horvath and Hannum measures of epigenetic age acceleration were moderately correlated (r = 0.50). The regression models revealed that the Horvath age acceleration measure exhibited marginal associations with increased postprandial HDL (p = 0.05), increased postprandial total cholesterol (p = 0.06), and decreased soluble interleukin 2 receptor subunit alpha (IL2sRα, p = 0.02). The Hannum measure of epigenetic age acceleration was inversely associated with fasting HDL (p = 0.02) and positively associated with postprandial TG (p = 0.02), interleukin-6 (IL6, p = 0.007), C-reactive protein (C-reactive protein, p = 0.0001), and tumor necrosis factor alpha (TNFα, p = 0.0001). Overall, the observed effect sizes were small and the association of the Hannum residual with inflammatory markers was attenuated by adjustment for estimated T cell type percentages. Conclusions Our study demonstrates that epigenetic age acceleration in blood relates to inflammatory biomarkers and certain lipid classes in Caucasian individuals of the GOLDN study. Future studies should consider epigenetic age acceleration in other tissues and extend the analysis to other ethnic groups.

DNA methylation (DNAm) is an important epigenetic process involved in the regulation of gene expression. While many studies have identified thousands of loci associated with age, few have differentiated between linear and non-linear DNAm trends with age. Non-linear trends could indicate early- or late-life gene regulatory processes. Using data from the Illumina 450K array on 336 human peripheral blood samples, we identified 21 CpG sites that associated with age (P

Background: Trimethylamine-N-oxide (TMAO), an atherogenic metabolite species, has emerged as a possible new risk factor for cardiovascular disease. Animal studies have shown that circulating TMAO levels are regulated by genetic and environmental factors. However, large-scale human studies have failed to replicate the observed genetic associations, and epigenetic factors such as DNA methylation have never been examined in relation to TMAO levels. Methods and results: We used data from the family-based Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) to investigate the heritable determinants of plasma TMAO in humans. TMAO was not associated with other plasma markers of cardiovascular disease, e.g. lipids or inflammatory cytokines. We first estimated TMAO heritability at 27%, indicating a moderate genetic influence. We used 1000 Genomes imputed data (n = 626) to estimate genome-wide associations with TMAO levels, adjusting for age, sex, family relationships, and study site. The genome-wide study yielded one significant hit at the genome-wide level, located in an intergenic region on chromosome 4. We subsequently quantified epigenome-wide DNA methylation using the Illumina Infinium array on CD4+ T-cells. We tested for association of methylation loci with circulating TMAO (n = 847), adjusting for age, sex, family relationships, and study site as the genome-wide study plus principal components capturing CD4+ T-cell purity. Upon adjusting for multiple testing, none of the epigenetic findings were statistically significant. Conclusions: Our findings contribute to the growing body of evidence suggesting that neither genetic nor epigenetic factors play a critical role in establishing circulating TMAO levels in humans.

Cluster of differentiation 36 (CD36) variants influence fasting lipids and risk of metabolic syndrome, but their impact on postprandial lipids, an independent risk factor for cardiovascular disease, is unclear. We determined the effects of SNPs within a ∼410 kb region encompassing CD36 and its proximal and distal promoters on chylomicron (CM) remnants and LDL particles at fasting and at 3.5 and 6 h following a high-fat meal (Genetics of Lipid Lowering Drugs and Diet Network study, n = 1,117). Five promoter variants associated with CMs, four with delayed TG clearance and five with LDL particle number. To assess mechanisms underlying the associations, we queried expression quantitative trait loci, DNA methylation, and ChIP-seq datasets for adipose and heart tissues that function in postprandial lipid clearance. Several SNPs that associated with higher serum lipids correlated with lower adipose and heart CD36 mRNA and aligned to active motifs for PPARγ, a major CD36 regulator. The SNPs also associated with DNA methylation sites that related to reduced CD36 mRNA and higher serum lipids, but mixed-model analyses indicated that the SNPs and methylation independently influence CD36 mRNA. The findings support contributions of CD36 SNPs that reduce adipose and heart CD36 RNA expression to inter-individual variability of postprandial lipid metabolism and document changes in CD36 DNA methylation that influence both CD36 expression and lipids.

Postprandial lipemia (PPL), the increased plasma TG concentration after consuming a high-fat meal, is an independent risk factor for CVD. Individual responses to a meal high in fat vary greatly, depending on genetic and lifestyle factors. However, only a few loci have been associated with TG-PPL response. Heritable epigenomic changes may be significant contributors to the unexplained inter-individual PPL variability. We conducted an epigenome-wide association study on 979 subjects with DNA methylation measured from CD4+ T cells, who were challenged with a high-fat meal as a part of the Genetics of Lipid Lowering Drugs and Diet Network study. Eight methylation sites encompassing five genes, LPP, CPT1A, APOA5, SREBF1, and ABCG1, were significantly associated with PPL response at an epigenome-wide level (P < 1.1 × 10−7), but no methylation site reached epigenome-wide significance after adjusting for baseline TG levels. Higher methylation at LPP, APOA5, SREBF1, and ABCG1, and lower methylation at CPT1A methylation were correlated with an increased TG-PPL response. These PPL-associated methylation sites, also correlated with fasting TG, account for a substantially greater amount of phenotypic variance (14.9%) in PPL and fasting TG (16.3%) when compared with the genetic contribution of loci identified by our previous genome-wide association study (4.5%). In summary, the epigenome is a large contributor to the variation in PPL, and this has the potential to be used to modulate PPL and reduce CVD.

Abstract GAW20 provided a platform for developing and evaluating statistical methods to analyze human lipid-related phenotypes, DNA methylation, and single-nucleotide markers in a study involving a pharmaceutical intervention. In this article, we present an overview of the data sets and the contributions analyzing these data. The data, donated by the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) investigators, included data from 188 families (N = 1105) which included genome-wide DNA methylation data before and after a 3-week treatment with fenofibrate, single-nucleotide polymorphisms, metabolic syndrome components before and after treatment, and a variety of covariates. The contributions from individual research groups were extensively discussed prior, during, and after the Workshop in groups based on discussion themes, before being submitted for publication.

Background: Associations of both common and rare genetic variants with fasting blood lipids have been extensively studied. However, most of the rare coding variants associated with lipids are population-specific, and exploration of genetic data from diverse population samples may enhance the identification of novel associations with rare variants.Results: We searched for novel coding genetic variants associated with fasting lipid levels in 894 samples from the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN) with exome-wide sequencing-based genotype data. In single variant tests, one variant (rs11171663 in ITGA7) was associated with fasting triglyceride levels (P = 7.66E-08), explaining approximately 3.2% of the total trait variance. In gene-based tests, we found statistically significant associations between ITGA7 (P = 1.77E-07) and SLCO2A1 (P = 7.18E-07) and triglycerides, as well as between POT1 (P = 3.00E-07) and low-density lipoprotein cholesterol. In another independent replication cohort consisting of 3,183 African American samples from Hypertension Genetic Epidemiology Network (HyperGEN) and the Genetic Epidemiology Network of Arteriopathy (GENOA), the top genes achieved P-values of 0.04 (ITGA7), 0.08 (SLCO2A1), and 0.02 (POT1). In GOLDN, gene transcript levels of ITGA7 and SLCO2A1 were associated with fasting triglycerides (P = 0.07 and P = 0.02), highlighting functional relevance of our findings.Conclusion: In this study, we present preliminary evidence of novel rare variant determinants of fasting lipids, and reveal potential underlying molecular mechanisms. Moreover, these results were replicated in an independent cohort. Our findings may inform novel biomarkers of disease risk and treatment targets.

BACKGROUND:The incidence of type 2 diabetes (T2DM) in children has increased dramatically. However, limited published information is known about the glycemic control and lipid outcomes in pediatric T2DM outside of clinical trials. OBJECTIVES:To determine the glycemic control and lipid measure outcomes at one and three- year follow-up in children with T2DM. METHODS:A retrospective electronic medical record review of children with T2DM at the Children's Hospital of Alabama over a 12-year period. RESULTS:There were 301 patients with a diagnosis of T2DM who had a 1-year follow-up visit, of which 184 also had a 3-year follow-up. Most patients (78%) received either insulin with metformin or insulin alone at diagnosis. At one year, 37% of the cohort achieved 'optimal glycemic control' (HbA1C ≤6.5%) and 58% of patients achieved durable glycemic control (HbA1C ≤8%). Optimal glycemic control was seen in 48 patients at 3 years. The patients treated with insulin (alone or in combination with metformin) tended to have higher HbA1C at diagnosis, but had improved lipid and glycemic outcomes at follow-up. The group treated with insulin along with metformin had significant improvements in non-HDL, HDL and TC/HDL ratios. The effects of insulin treatment on glycemic control at 3 years were not statistically significant. CONCLUSION:With the current modality of treatment, only a minority of patients achieve optimal glycemic control at 1 and 3 years of follow-up. Studies are warranted to further elucidate the optimal therapies in the management of pediatric T2DM.

Advancements in high-throughput technologies have made it feasible to study thousands of biological pathways simultaneously for a holistic assessment of health and disease risk via ‘omics platforms. A major challenge in ‘omics research revolves around the reproducibility of findings—a feat that hinges upon balancing false-positive associations with generalizability. Given the foundational role of reproducibility in scientific inference, replication and validation of ‘omics findings are cornerstones of this effort. In this narrative review, we define key terms relevant to replication and validation, present issues surrounding each concept with historical and contemporary examples from genomics (the most well-established and upstream ‘omics), discuss special issues and unique considerations for replication and validation in metabolomics (an emerging field and most downstream ‘omics for which best practices remain yet to be established), and make suggestions for future research leveraging multiple ‘omics datasets.

Among 29,701 Black and White participants aged 45 years and older in the Reasons for Geographic and Racial Difference in Stroke (REGARDS) study, allostatic load (AL) was defined as the sum score of established baseline risk-associated biomarkers for which participants exceeded a set cutoff point. Cox proportional hazard regression was utilized to determine the association of AL score with all-cause and cancer-specific mortality, with analyses stratified by body-mass index, age group, and race. At baseline, Blacks had a higher AL score compared with Whites (Black mean AL score: 2.42, SD: 1.50; White mean AL score: 1.99, SD: 1.39; p < 0.001). Over the follow-up period, there were 4622 all-cause and 1237 cancer-specific deaths observed. Every unit increase in baseline AL score was associated with a 24% higher risk of all-cause (HR: 1.24, 95% CI: 1.22, 1.27) and a 7% higher risk of cancer-specific mortality (HR: 1.07, 95% CI: 1.03, 1.12). The association of AL with overall- and cancer-specific mortality was similar among Blacks and Whites and across age-groups, however the risk of cancer-specific mortality was higher among normal BMI than overweight or obese participants. In conclusion, a higher baseline AL score was associated with increased risk of all-cause and cancer-specific mortality among both Black and White participants. Targeted interventions to patient groups with higher AL scores, regardless of race, may be beneficial as a strategy to reduce all-cause and cancer-specific mortality.

BACKGROUND:Odd-numbered chain saturated fatty acids (OCSFA) have been associated with potential health benefits. Although some OCSFA (e.g., C15:0 and C17:0) are found in meats and dairy products, sources and metabolism of C19:0 and C23:0 are relatively unknown, and the influence of non-dietary determinants, including genetic factors, on circulating levels of OCSFA is not established. OBJECTIVE:To elucidate the biological processes that influence circulating levels of OCSFA by investigating associations between genetic variation and OCSFA. DESIGN:We performed a meta-analysis of genome-wide association studies (GWAS) of plasma phospholipid/erythrocyte levels of C15:0, C17:0, C19:0, and C23:0 among 11,494 individuals of European descent. We also investigated relationships between specific single nucleotide polymorphisms (SNPs) in the lactase (LCT) gene, associated with adult-onset lactase intolerance, with circulating levels of dairy-derived OCSFA, and evaluated associations of candidate sphingolipid genes with C23:0 levels. RESULTS:We found no genome-wide significant evidence that common genetic variation is associated with circulating levels of C15:0 or C23:0. In two cohorts with available data, we identified one intronic SNP (rs13361131) in myosin X gene (MYO10) associated with C17:0 level (P = 1.37×10-8), and two intronic SNP (rs12874278 and rs17363566) in deleted in lymphocytic leukemia 1 (DLEU1) region associated with C19:0 level (P = 7.07×10-9). In contrast, when using a candidate-gene approach, we found evidence that three SNPs in LCT (rs11884924, rs16832067, and rs3816088) are associated with circulating C17:0 level (adjusted P = 4×10-2). In addition, nine SNPs in the ceramide synthase 4 (CERS4) region were associated with circulating C23:0 levels (adjusted P

BackgroundThe link between DNA methylation, obesity, and adiposity-related diseases in the general population remains uncertain.Methods and findingsWe conducted an association study of body mass index (BMI) and differential methylation for over 400,000 CpGs assayed by microarray in whole-blood-derived DNA from 3,743 participants in the Framingham Heart Study and the Lothian Birth Cohorts, with independent replication in three external cohorts of 4,055 participants. We examined variations in whole blood gene expression and conducted Mendelian randomization analyses to investigate the functional and clinical relevance of the findings. We identified novel and previously reported BMI-related differential methylation at 83 CpGs that replicated across cohorts; BMI-related differential methylation was associated with concurrent changes in the expression of genes in lipid metabolism pathways. Genetic instrumental variable analysis of alterations in methylation at one of the 83 replicated CpGs, cg11024682 (intronic to sterol regulatory element binding transcription factor 1 [SREBF1]), demonstrated links to BMI, adiposity-related traits, and coronary artery disease. Independent genetic instruments for expression of SREBF1 supported the findings linking methylation to adiposity and cardiometabolic disease. Methylation at a substantial proportion (16 of 83) of the identified loci was found to be secondary to differences in BMI. However, the cross-sectional nature of the data limits definitive causal determination.ConclusionsWe present robust associations of BMI with differential DNA methylation at numerous loci in blood cells. BMI-related DNA methylation and gene expression provide mechanistic insights into the relationship between DNA methylation, obesity, and adiposity-related diseases.

Lipoprotein subfractions help discriminate cardiometabolic disease risk. Genetic loci validated as associating with lipoprotein measures do not account for a large proportion of the individual variation in lipoprotein measures. We hypothesized that DNA methylation levels across the genome contribute to interindividual variation in lipoprotein measures. Using data from participants of the Genetics of Lipid Lowering Drugs and Diet Network (n = 663 for discovery and n = 331 for replication stages, respectively), we conducted the first systematic screen of the genome to determine associations between methylation status at ∼470,000 cytosine-guanine dinucleotide (CpG) sites in CD4+ T cells and 14 lipoprotein subfraction measures. We modeled associations between methylation at each CpG site and each lipoprotein measure separately using linear mixed models, adjusted for age, sex, study site, cell purity, and family structure. We identified two CpGs, both in the carnitine palmitoyltransferase-1A (CPT1A) gene, which reached significant levels of association with VLDL and LDL subfraction parameters in both discovery and replication phases (P < 1.1 × 10−7 in the discovery phase, P < .004 in the replication phase, and P < 1.1 × 10−12 in the full sample). CPT1A is regulated by PPARα, a ligand for drugs used to reduce CVD. Our associations between methylation in CPT1A and lipoprotein measures highlight the epigenetic role of this gene in metabolic dysfunction.

In this study, we conducted an epigenome-wide association study of metabolic syndrome (MetS) among 846 participants of European descent in the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN). DNA was isolated from CD4+ T cells and methylation at ~470,000 cytosine-phosphate-guanine dinucleotide (CpG) pairs was assayed using the Illumina Infinium HumanMethylation450 BeadChip. We modeled the percentage methylation at individual CpGs as a function of MetS using linear mixed models. A Bonferroni-corrected P-value of 1.1 x 10(-7) was considered significant. Methylation at two CpG sites in CPT1A on chromosome 11 was significantly associated with MetS (P for cg00574958 = 2.6x10(-14) and P for cg17058475 = 1.2x10(-9)). Significant associations were replicated in both European and African ancestry participants of the Bogalusa Heart Study. Our findings suggest that methylation in CPT1A is a promising epigenetic marker for MetS risk which could become useful as a treatment target in the future.

DNA methylation at CpG sites is both heritable and influenced by environment, but the relative contributions of each to DNA methylation levels are unclear. We conducted a heritability analysis of CpG methylation in human CD4+ cells across 975 individuals from 163 families in the Genetics of Lipid-lowering Drugs and Diet Network (GOLDN). Based on a broad-sense heritability (H2) value threshold of 0.4, we identified 20,575 highly heritable CpGs among the 174,445 most variable autosomal CpGs (SD > 0.02). Tests for associations of heritable CpGs with genotype at 2,145,360 SNPs using 717 of 975 individuals showed that ~74% were cis-meQTLs (< 1 Mb away from the CpG), 6% of CpGs exhibited trans-meQTL associations (>1 Mb away from the CpG or located on a different chromosome), and 20% of CpGs showed no strong significant associations with genotype (based on a p-value threshold of 1e-7). Genes proximal to the genotype independent heritable CpGs were enriched for functional terms related to regulation of T cell activation. These CpGs were also among those that distinguished T cells from other blood cell lineages. Compared to genes proximal to meQTL-associated heritable CpGs, genotype independent heritable CpGs were moderately enriched in the same genomic regions that escape erasure during primordial germ cell development and could carry potential for generational transmission.

Background Significant controversy exists regarding the best approach for nonculprit vessel revascularization in patients with multivessel coronary artery disease presenting with ST‐segment elevation myocardial infarction. We conducted a systematic investigation to pool data from current randomized controlled trials (RCTs) to assess optimal treatment strategies in this patient population. Methods and Results A comprehensive search of SCOPUS from inception through May 2015 was performed using predefined criteria. We compared efficacy and safety outcomes of different approaches by categorizing the studies into 3 groups: (1) complete revascularization (CR) versus culprit lesion revascularization (CL) at index hospitalization, (2) CR at index hospitalization versus staged revascularization (SR) of nonculprit vessels at a separate hospitalization, and (3) comparison of SR versus CL. Eight eligible RCTs met the inclusion criteria: (1) CR versus CL (6 RCTs, n=1727) (2) CR versus SR (3 RCTs, n=311), and (3) SR versus CL (1 RCT, n=149). We observed significantly lower rates of major adverse cardiovascular events, revascularization, and repeat percutaneous coronary interventions among patients treated with CR and SR compared with a CL approach (P

OBJECTIVES/SPECIFIC AIMS: The purpose of this study is to characterize children with nonalcoholic fatty liver disease (NAFLD) living in the Southeastern United States. METHODS/STUDY POPULATION: This retrospective electronic medical record chart review was conducted on a random sample of 206 children identified with NAFLD. Patients were included if they met the following criteria: confirmed NAFLD through either an ultrasound or liver biopsy or clinical suspicion of fatty liver disease alongside elevated alanine aminotransferase (ALT) in the absence of other etiologies causing elevated transaminases. Patients were excluded if they had hepatitis or other documented liver disease. Data collected at initial presentation included age, gender, ethnicity, height, weight, body mass index (BMI), BMI percentile, blood pressure, HbA1c, aspartate aminotransferase (AST), ALT, γ-glutamyl transferase (GGT), total cholesterol, total triglycerides, low-density lipoprotein, and high-density lipoprotein. Statistical analysis: for descriptive statistics, frequency counts and percentages alongside means, standard deviation, range, min/max values for the continuous variables were calculated. RESULTS/ANTICIPATED RESULTS: This study included 206 children diagnosed with NAFLD. Subjects were primarily male (n=136, 66%) and Caucasian (n=133, 66%), followed by Hispanic (n=42, 21%), Black (n=25, 12%), and Asian (n=2, 1%). Mean age at diagnosis was 12.3±3.5 years. Mean weight (lbs), height (in), and BMI (kg/m2) of subjects at diagnosis were 192±77 lbs, 61.7±6.6 in, 34.6±9.7 kg/m2, respectively. Patients had an average systolic blood pressure of 124±15.4 mmHg and diastolic blood pressure of 69.6±10.6 mmHg. Mean ALT was 91.8±67.2 U/L, AST was 61±38.8 U/L, and GGT was 55.1±64.6 U/L. Mean HbA1c was 5.8±1.4%, cholesterol was 176±36.3 mg/dL, triglycerides were 200±134 mg/dL, low-density lipoprotein was 107.6±32.1 mg/dL, and high-density lipoprotein was 39.9±8.4 mg/dL. DISCUSSION/SIGNIFICANCE OF IMPACT: In addition to having significantly elevated liver enzymes, children with NAFLD had several derangements in their metabolic profile, most notably high triglyceride levels and HbA1c values in the prediabetic range. Although lifestyle modification is the gold standard treatment for NAFLD, pharmacotherapy may need to be included to address metabolic syndrome.

Increased postprandial lipid (PPL) response to dietary fat intake is a heritable risk factor for cardiovascular disease (CVD). Variability in postprandial lipids results from the complex interplay of dietary and genetic factors. We hypothesized that detailed lipid profiles (eg, sterols and fatty acids) may help elucidate specific genetic and dietary pathways contributing to the PPL response.We used gas chromatography mass spectrometry to quantify the change in plasma concentration of 35 fatty acids and 11 sterols between fasting and 3.5 hours after the consumption of a high-fat meal (PPL challenge) among 40 participants from the GOLDN study. Correlations between sterols, fatty acids and clinical measures were calculated. Mixed linear regression was used to evaluate associations between lipidomic profiles and genomic markers including single nucleotide polymorphisms (SNPs) and methylation markers derived from the Affymetrix 6.0 array and the Illumina Methyl450 array, respectively. After the PPL challenge, fatty acids increased as well as sterols associated with cholesterol absorption, while sterols associated with cholesterol synthesis decreased. PPL saturated fatty acids strongly correlated with triglycerides, very low-density lipoprotein, and chylomicrons. Two SNPs (rs12247017 and rs12240292) in the sorbin and SH3 domain containing 1 (SORBS1) gene were associated with b-Sitosterol after correction for multiple testing (P≤4.5*10(-10)). SORBS1 has been linked to obesity and insulin signaling. No other markers reached the genome-wide significance threshold, yet several other biologically relevant loci are highlighted (eg, PRIC285, a co-activator of PPARa).Integration of lipidomic and genomic data has the potential to identify new biomarkers of CVD risk.

A recent large-scale meta-analysis of genome-wide studies has identified 95 loci, 59 of them novel, as statistically significant predictors of blood lipid traits; we tested whether the same loci explain the observed heterogeneity in response to lipid-lowering therapy with fenofibrate. Using data from the Genetics of Lipid Lowering Drugs and Diet Network (GOLDN, n = 861) we fit linear mixed models with the genetic markers as predictors and high-density lipoprotein (HDL) cholesterol, low-density lipoprotein (LDL) cholesterol, total cholesterol, and triglyceride concentrations as outcomes. For all four traits, we analyzed both baseline levels and changes in response to treatment with fenofibrate. For the markers that were significantly associated with fenofibrate response, we fit additional models evaluating potential epistatic interactions. All models were adjusted for age, sex, and study center as fixed effects, and pedigree as a random effect. Statistically significant associations were observed between the rs964184 polymorphism near APOA1 (P-value≤0.0001) and fenofibrate response for HDL and triglycerides. The association was replicated in the Pharmacogenetics of Hypertriglyceridemia in Hispanics study (HyperTG, n = 267). Suggestive associations with fenofibrate response were observed for markers in or near PDE3A, MOSC1, FLJ36070, CETP, the APOE-APOC1-APOC4-APOC2, and CILP2. Finally, we present strong evidence for epistasis (P-value for interaction = 0.0006 in GOLDN, 0.05 in HyperTG) between rs10401969 near CILP2 and rs4420638 in the APOE-APOC1-APOC4-APOC2 cluster with total cholesterol response to fenofibrate. In conclusion, we present evidence linking several novel and biologically relevant genetic polymorphisms to lipid lowering drug response, as well as suggesting novel gene-gene interactions in fenofibrate pharmacogenetics.

ObjectiveWe sought to identify rare variants influencing brain imaging phenotypes in the Framingham Heart Study by performing whole genome sequence association analyses within the Trans-Omics for Precision Medicine Program.MethodsWe performed association analyses of cerebral and hippocampal volumes and white matter hyperintensity (WMH) in up to 2,180 individuals by testing the association of rank-normalized residuals from mixed-effect linear regression models adjusted for sex, age, and total intracranial volume with individual variants while accounting for familial relatedness. We conducted gene-based tests for rare variants using (1) a sliding-window approach, (2) a selection of functional exonic variants, or (3) all variants.ResultsWe detected new loci in 1p21 for cerebral volume (minor allele frequency [MAF] 0.005, p = 10-8) and in 16q23 for hippocampal volume (MAF 0.05, p = 2.7 × 10-8). Previously identified associations in 12q24 for hippocampal volume (rs7294919, p = 4.4 × 10-4) and in 17q25 for WMH (rs7214628, p = 2.0 × 10-3) were confirmed. Gene-based tests detected associations (p ≤ 2.3 × 10-6) in new loci for cerebral (5q13, 8p12, 9q31, 13q12-q13, 15q24, 17q12, 19q13) and hippocampal volumes (2p12) and WMH (3q13, 4p15) including Alzheimer disease- (UNC5D) and Parkinson disease-associated genes (GBA). Pathway analyses evidenced enrichment of associated genes in immunity, inflammation, and Alzheimer disease and Parkinson disease pathways.ConclusionsWhole genome sequence-wide search reveals intriguing new loci associated with brain measures. Replication of novel loci is needed to confirm these findings.

Background: Hypertensive African Americans have a ~50% response rate to thiazide diuretic treatment. This contributes to a high prevalence of uncontrolled high blood pressure. Here, we examined the role of the mitochondrial genome on thiazide diuretic treatment response in hypertensive African Americans enrolled in a clinical trial. Methods: Participants from the Antihypertensive and Lipid Lowering Treatment to Prevent Heart Attack Trial (ALLHAT, n= 4279) were genotyped using the Illumina Infinium Multi-Ethnic Beadchip. Haplotype groups were called using HaploGrep. We used a linear regression analysis to examine the association between mitochondrial haplogroups (L, M, and N) and changes in blood pressure and fasting glucose over six months and two years, respectively. Results: The analysis revealed a null association between mitochondrial haplogroups M and N versus L for each of the outcomes. In subgroup analysis, the L subclades L1, L2, and L3/L4 (versus L0) were each inversely associated with fasting glucose response (p < 0.05). Conclusions: This discovery analysis suggests the mitochondrial genome has a small effect on fasting glucose response, but not that of blood pressure, to thiazide diuretic treatment in African Americans.

Background: Hypertensive African Americans have a ~50% response rate to thiazide diuretic treatment. This contributes to a high prevalence of uncontrolled high blood pressure. Here, we examined the role of the mitochondrial genome on thiazide diuretic treatment response in hypertensive African Americans enrolled in a clinical trial. Methods: Participants from the Antihypertensive and Lipid Lowering Treatment to Prevent Heart Attack Trial (ALLHAT, n= 4279) were genotyped using the Illumina Infinium Multi-Ethnic Beadchip. Haplotype groups were called using HaploGrep. We used a linear regression analysis to examine the association between mitochondrial haplogroups (L, M, and N) and changes in blood pressure and fasting glucose over six months and two years, respectively. Results: The analysis revealed a null association between mitochondrial haplogroups M and N versus L for each of the outcomes. In subgroup analysis, the L subclades L1, L2, and L3/L4 (versus L0) were each inversely associated with fasting glucose response (p < 0.05). Conclusions: This discovery analysis suggests the mitochondrial genome has a small effect on fasting glucose response, but not that of blood pressure, to thiazide diuretic treatment in African Americans.