Genome-wide identification of DNA methylation QTLs in whole blood highlights pathways for cardiovascular disease.

Identifying methylation quantitative trait loci (meQTLs) and integrating them with disease-associated variants from genome-wide association studies (GWAS) may illuminate functional mechanisms underlying genetic variant-disease associations. Here, we perform GWAS of >415 thousand CpG methylation sites in whole blood from 4170 individuals and map 4.7 million cis- and 630 thousand trans-meQTL variants targeting >120 thousand CpGs. Independent replication is performed in 1347 participants from two studies. By linking cis-meQTL variants with GWAS results for cardiovascular disease (CVD) traits, we identify 92 putatively causal CpGs for CVD traits by Mendelian randomization analysis. Further integrating gene expression data reveals evidence of cis CpG-transcript pairs causally linked to CVD. In addition, we identify 22 trans-meQTL hotspots each targeting more than 30 CpGs and find that trans-meQTL hotspots appear to act in cis on expression of nearby transcriptional regulatory genes. Our findings provide a powerful meQTL resource and shed light on DNA methylation involvement in human diseases.

Blood Leukocyte DNA Methylation Predicts Risk of Future Myocardial Infarction and Coronary Heart Disease.

BACKGROUND: DNA methylation is implicated in coronary heart disease (CHD), but current evidence is based on small, cross-sectional studies. We examined blood DNA methylation in relation to incident CHD across multiple prospective cohorts. METHODS: Nine population-based cohorts from the United States and Europe profiled epigenome-wide blood leukocyte DNA methylation using the Illumina Infinium 450k microarray, and prospectively ascertained CHD events including coronary insufficiency/unstable angina, recognized myocardial infarction, coronary revascularization, and coronary death. Cohorts conducted race-specific analyses adjusted for age, sex, smoking, education, body mass index, blood cell type proportions, and technical variables. We conducted fixed-effect meta-analyses across cohorts. RESULTS: Among 11 461 individuals (mean age 64 years, 67% women, 35% African American) free of CHD at baseline, 1895 developed CHD during a mean follow-up of 11.2 years. Methylation levels at 52 CpG (cytosine-phosphate-guanine) sites were associated with incident CHD or myocardial infarction (false discovery rate<0.05). These CpGs map to genes with key roles in calcium regulation (ATP2B2, CASR, GUCA1B, HPCAL1), and genes identified in genome- and epigenome-wide studies of serum calcium (CASR), serum calcium-related risk of CHD (CASR), coronary artery calcified plaque (PTPRN2), and kidney function (CDH23, HPCAL1), among others. Mendelian randomization analyses supported a causal effect of DNA methylation on incident CHD; these CpGs map to active regulatory regions proximal to long non-coding RNA transcripts. CONCLUSION: Methylation of blood-derived DNA is associated with risk of future CHD across diverse populations and may serve as an informative tool for gaining further insight on the development of CHD.

Cognition level and change in cognition during adolescence are associated with cognition in midlife.

PURPOSE: Cognitive development during adolescence affects health long term. We investigated whether level of and change in language-based cognition during adolescence are associated with cognitive performance in midlife. METHODS: Participants were enrolled in the Child Health and Development Study and followed during midlife (47-52 years). Adolescent cognition was measured with the Peabody Picture Vocabulary Test at ages 9-11 years (PPVT-9) and 15-17 years (PPVT-15). We examined PPVT-9, as well as a PPVT change score (derived using the standardized regression-based method) in relation to midlife cognition measures of Wechsler Test of Adult Reading, Verbal Fluency, and Digit Symbol tests. Linear regression models were adjusted for childhood socioeconomic status, age, sex, race, and midlife marital status, education, and occupational score. RESULTS: In 357 participants (52.1% female, 25.6% African American), both PPVT-9 (ß [95% confidence interval [CI] = 0.26 [0.18, 0.34]) and PPVT change score (ß [95% CI] = 2.03 [1.27, 2.80]) were associated with Wechsler Test of Adult Reading at midlife. PPVT-9 was associated with midlife Verbal Fluency (ß [95% CI] = 0.18 [0.10, 0.25]), whereas PPVT change score was not (ß [95% CI] = -0.01 [-0.68, 0.67]). Neither PPVT-9 nor PPVT change score was associated with midlife Digit Symbol. CONCLUSIONS: Both level of and change in language-based cognition during adolescence were associated with midlife vocabulary and language function, even after controlling for midlife occupation and education.

Mendelian Randomization and the Environmental Epigenetics of Health: a Systematic Review.

PURPOSE OF REVIEW: Epigenetic modifications are environmentally responsive and may play a mechanistic role in the development of disease. Mendelian randomization uses genetic variation to assess the causal effect of modifiable exposures on health outcomes. We conducted a systematic review of Mendelian randomization studies evaluating the causal role of DNA methylation (DNAm) changes on the development of health states, emphasizing on studies that formally evaluate exposure-DNAm, in addition to DNAm-outcome, causal associations. RECENT FINDINGS: We identified 15 articles, 4 of them including an environmental determinant of DNAm, including self-reported tobacco smoke exposure, in utero tobacco smoke exposure, measured vitamin B12, and glycemia. Selected articles suggest a causal association of DNAm with some cardiometabolic endpoints. DNAm seemed to partly explain the association of postnatal and prenatal exposure to tobacco smoke and vitamin B12 with inflammation biomarkers, birth weight, and cognitive outcomes, respectively. However, the current evidence is not sufficient to infer causality. Additional Mendelian randomization studies from large epidemiologic samples are needed to support the causal role of environmental factors as determinants of health-related epigenetic modifications.

Early life disadvantage and adult adiposity: tests of sensitive periods during childhood and behavioural mediation in adulthood.

BACKGROUND: Early exposure to socioeconomic disadvantage is associated with obesity. Here we investigated how early, and conducted mediation analyses to identify behavioural factors in adulthood that could explain why. METHODS: Among 931 participants in the New England Family Study, we investigated the associations of family socioeconomic disadvantage measured before birth and at age 7 years with the following measures of adiposity in mid-adulthood (mean age = 44.4 years): body mass index (BMI), waist circumference and, among 400 participants, body composition from dual-energy X-ray absorption scans. RESULTS: In linear regressions adjusting for age, sex, race and childhood BMI Z-score, participants in the highest tertile of socioeconomic disadvantage at birth had 2.6 additional BMI units in adulthood [95% confidence interval (CI) = 1.26, 3.96], 5.62 cm waist circumference (95% CI = 2.69, 8.55), 0.73 kg of android fat mass (95% CI = 0.25, 1.21), and 7.65 higher Fat Mass Index (95% CI = 2.22, 13.09). Conditional on disadvantage at birth, socioeconomic disadvantage at age 7 years was not associated with adult adiposity. In mediation analyses, 10-20% of these associations were explained by educational attainment and 5-10% were explained by depressive symptoms. CONCLUSIONS: Infancy may be a sensitive period for exposure to socioeconomic disadvantage, as exposure in the earliest years of life confers a larger risk for overall and central adiposity in mid-adulthood than exposure during childhood. Intervention on these two adult risk factors for adiposity would, if all model assumptions were satisfied, only remediate up to one-fifth of the excess adult adiposity among individuals born into socioeconomically disadvantaged households.

Analysis of repeated leukocyte DNA methylation assessments reveals persistent epigenetic alterations after an incident myocardial infarction.

BACKGROUND: Most research into myocardial infarctions (MIs) have focused on preventative efforts. For survivors, the occurrence of an MI represents a major clinical event that can have long-lasting consequences. There has been little to no research into the molecular changes that can occur as a result of an incident MI. Here, we use three cohorts to identify epigenetic changes that are indicative of an incident MI and their association with gene expression and metabolomics. RESULTS: Using paired samples from the KORA cohort, we screened for DNA methylation loci (CpGs) whose change in methylation is potentially indicative of the occurrence of an incident MI between the baseline and follow-up exams. We used paired samples from the NAS cohort to identify 11 CpGs which were predictive in an independent cohort. After removing two CpGs associated with medication usage, we were left with an "epigenetic fingerprint" of MI composed of nine CpGs. We tested this fingerprint in the InCHIANTI cohort where it moderately discriminated incident MI occurrence (AUC = 0.61, P = 6.5 × 10-3). Returning to KORA, we associated the epigenetic fingerprint loci with cis-gene expression and integrated it into a gene expression-metabolomic network, which revealed links between the epigenetic fingerprint CpGs and branched chain amino acid (BCAA) metabolism. CONCLUSIONS: There are significant changes in DNA methylation after an incident MI. Nine of these CpGs show consistent changes in multiple cohorts, significantly discriminate MI in independent cohorts, and were independent of medication usage. Integration with gene expression and metabolomics data indicates a link between MI-associated epigenetic changes and BCAA metabolism.

DNA methylation in blood as a mediator of the association of mid-childhood body mass index with cardio-metabolic risk score in early adolescence.

Obesity is associated with higher cardio-metabolic risk even in childhood and adolescence; whether this association is mediated by epigenetic mechanisms remains unclear. We examined the extent to which mid-childhood body mass index (BMI) z-score (median age 7.7 years) was associated with cardio-metabolic risk score in early adolescence (median age 12.9 years) via mid-childhood DNA methylation among 265 children in the Project Viva. We measured DNA methylation in leukocytes using the Infinium Human Methylation450K BeadChip. We assessed mediation CpG-by-CpG using epigenome-wide association analyses, high-dimensional mediation analysis, and natural effect models. We observed mediation by mid-childhood DNA methylation at 6 CpGs for the association between mid-childhood BMI z-score and cardio-metabolic risk score in early adolescence in the high-dimensional mediation analysis (accounting for 10% of the total effect) and in the natural effect model (ß = 0.04, P = 3.2e-2, accounting for 13% of the total effect). The natural direct effect of BMI z-score on cardio-metabolic risk score was still evident (ß = 0.27, P = 1.1e-25). We also observed mediation by mid-childhood DNA methylation at 5 CpGs that was in the opposite direction from the total effect (natural effect model: ß = -0.04, P = 2.0e-2). Mediation in different directions implies a complex role of DNA methylation in the association between BMI and cardio-metabolic risk and needs further investigation. Future studies with larger sample size and greater variability in cardio-metabolic risk will further help elucidate the role of DNA methylation for cardio-metabolic risk.

Residential Proximity to Major Roadways at Birth, DNA Methylation at Birth and Midchildhood, and Childhood Cognitive Test Scores: Project Viva(Massachusetts, USA).

BACKGROUND: Epigenetic variability is hypothesized as a regulatory pathway through which prenatal exposures may influence child development and health. OBJECTIVE: We sought to examine the associations of residential proximity to roadways at birth and epigenome-wide DNA methylation. We also assessed associations of differential methylation with child cognitive outcomes. METHODS: We estimated residential proximity to roadways at birth using a geographic information system (GIS) and cord blood methylation using Illumina's HumanMethylation450-array in 482 mother-child pairs in Project Viva. We identified individual CpGs associated with residential-proximity-to-roadways at birth using robust linear regression [[Formula: see text]]. We also estimated association between proximity-to-roadways at birth and methylation of the same sites in blood samples collected at age 7-11 y ([Formula: see text]). We ran the same analyses in the Generation R Study for replication ([Formula: see text]). In Project Viva, we investigated associations of differential methylation at birth with midchildhood cognition using linear regression. RESULTS: Living closer to major roadways at birth was associated with higher cord blood (and-more weakly-midchildhood blood) methylation of four sites in LAMB2. For each halving of residential-proximity-to-major-roadways, we observed a 0.82% increase in DNA methylation at cg05654765 [95% confidence interval (CI): (0.54%, 1.10%)], 0.88% at cg14099457 [95% CI: (0.56%, 1.19%)], 0.19% at cg03732535 [95% CI: (0.11%, 0.28)], and 1.08% at cg02954987 [95% CI: (0.65%, 1.51%)]. Higher cord blood methylation of these sites was associated with lower midchildhood nonverbal cognitive scores. Our results did not replicate in the Generation R Study. CONCLUSIONS: Our discovery results must be interpreted with caution, given that they were not replicated in a separate cohort. However, living close to major roadways at birth was associated with cord blood methylation of sites in LAMB2-a gene known to be linked to axonal development-in our U.S. cohort. Higher methylation of these sites associated with lower nonverbal cognitive scores at age 7-11 y in the same children. https://doi.org/10.1289/EHP2034.

Sex-specific epigenetic mediators between early life social disadvantage and adulthood BMI.

AIM: The objective of this study was to identify potential epigenetic mediating pathways linking early life social disadvantage (ELSD) to adulthood BMI. METHODS: Sex-specific epigenome-wide two-stage mediation analyses were conducted in blood and adipose tissue, and mediation estimates were obtained using cross-product mediation analysis. Pathway analyses were conducted using GREAT software (Bejerano Lab, CA, USA). RESULTS: Candidate mediation CpG sites were identified in adipose tissue, but not blood, and were sex-specific. Significant mediation sites in females included CpG loci in genes: PKHG1, BCAR3, ADAM5P, PIEZO1, FGFRL1, FASN and DPP9, among others. Pathway analyses revealed evidence of enrichment for processes associated with TFG-ß signaling and immunologic signatures. In males, significant mediation loci included sites in MAP3K5 and RPTOR, which have previously been associated with adipogenesis, inflammation and insulin resistance. CONCLUSION: Our findings provide supportive evidence for the mediating role of epigenetic mechanisms in the effect of early life social disadvantage on adulthood BMI.

Association of Methylation Signals With Incident Coronary Heart Disease in an Epigenome-Wide Assessment of Circulating Tumor Necrosis Factor α.

Importance: Tumor necrosis factor α (TNF-α) is a proinflammatory cytokine with manifold consequences for mammalian pathophysiology, including cardiovascular disease. A deeper understanding of TNF-α biology may enhance treatment precision. Objective: To conduct an epigenome-wide analysis of blood-derived DNA methylation and TNF-α levels and to assess the clinical relevance of findings. Design, Setting, and Participants: This meta-analysis assessed epigenome-wide associations in circulating TNF-α concentrations from 5 cohort studies and 1 interventional trial, with replication in 3 additional cohort studies. Follow-up analyses investigated associations of identified methylation loci with gene expression and incident coronary heart disease; this meta-analysis included 11 461 participants who experienced 1895 coronary events. Exposures: Circulating TNF-α concentration. Main Outcomes and Measures: DNA methylation at approximately 450 000 loci, neighboring DNA sequence variation, gene expression, and incident coronary heart disease. Results: The discovery cohort included 4794 participants, and the replication study included 816 participants (overall mean [SD] age, 60.7 [8.5] years). In the discovery stage, circulating TNF-α levels were associated with methylation of 7 cytosine-phosphate-guanine (CpG) sites, 3 of which were located in or near DTX3L-PARP9 at cg00959259 (ß [SE] = -0.01 [0.003]; P = 7.36 × 10-8), cg08122652 (ß [SE] = -0.008 [0.002]; P = 2.24 × 10-7), and cg22930808(ß [SE] = -0.01 [0.002]; P = 6.92 × 10-8); NLRC5 at cg16411857 (ß [SE] = -0.01 [0.002]; P = 2.14 × 10-13) and cg07839457 (ß [SE] = -0.02 [0.003]; P = 6.31 × 10-10); or ABO, at cg13683939 (ß [SE] = 0.04 [0.008]; P = 1.42 × 10-7) and cg24267699 (ß [SE] = -0.009 [0.002]; P = 1.67 × 10-7), after accounting for multiple testing. Of these, negative associations between TNF-α concentration and methylation of 2 loci in NLRC5 and 1 in DTX3L-14 PARP9 were replicated. Replicated TNF-α-linked CpG sites were associated with 9% to 19% decreased risk of incident coronary heart disease per 10% higher methylation per CpG site (cg16411857: hazard ratio [HR], 0.86; 95% CI, 0.78-1.95; P = .003; cg07839457: HR, 0.89; 95% CI, 0.80-0.94; P = 3.1 × 10-5; cg00959259: HR, 0.91; 95% CI, 0.84-0.97; P = .002; cg08122652: HR, 0.81; 95% CI, 0.74-0.89; P = 2.0 × 10-5). Conclusions and Relevance: We identified and replicated novel epigenetic correlates of circulating TNF-α concentration in blood samples and linked these loci to coronary heart disease risk, opening opportunities for validation and therapeutic applications.

Maternal BMI at the start of pregnancy and offspring epigenome-wide DNA methylation: findings from the pregnancy and childhood epigenetics (PACE) consortium.

Pre-pregnancy maternal obesity is associated with adverse offspring outcomes at birth and later in life. Individual studies have shown that epigenetic modifications such as DNA methylation could contribute. Within the Pregnancy and Childhood Epigenetics (PACE) Consortium, we meta-analysed the association between pre-pregnancy maternal BMI and methylation at over 450,000 sites in newborn blood DNA, across 19 cohorts (9,340 mother-newborn pairs). We attempted to infer causality by comparing the effects of maternal versus paternal BMI and incorporating genetic variation. In four additional cohorts (1,817 mother-child pairs), we meta-analysed the association between maternal BMI at the start of pregnancy and blood methylation in adolescents. In newborns, maternal BMI was associated with small (<0.2% per BMI unit (1 kg/m2), P < 1.06 × 10-7) methylation variation at 9,044 sites throughout the genome. Adjustment for estimated cell proportions greatly attenuated the number of significant CpGs to 104, including 86 sites common to the unadjusted model. At 72/86 sites, the direction of the association was the same in newborns and adolescents, suggesting persistence of signals. However, we found evidence for acausal intrauterine effect of maternal BMI on newborn methylation at just 8/86 sites. In conclusion, this well-powered analysis identified robust associations between maternal adiposity and variations in newborn blood DNA methylation, but these small effects may be better explained by genetic or lifestyle factors than a causal intrauterine mechanism. This highlights the need for large-scale collaborative approaches and the application of causal inference techniques in epigenetic epidemiology.

Exposure to Low Levels of Lead in Utero and Umbilical Cord Blood DNA Methylation in Project Viva: An Epigenome-Wide Association Study.

BACKGROUND: Early-life exposure to lead is associated with deficits in neurodevelopment and with hematopoietic system toxicity. DNA methylation may be one of the underlying mechanisms for the adverse effects of prenatal lead on the offspring, but epigenome-wide methylation data for low levels of prenatal lead exposure are lacking. OBJECTIVES: We investigated the association between prenatal maternal lead exposure and epigenome-wide DNA methylation in umbilical cord blood nucleated cells in Project Viva, a prospective U.S.-based prebirth cohort with relatively low levels of lead exposure. METHODS: Among 268 mother-infant pairs, we measured lead concentrations in red blood cells (RBC) from prenatal maternal blood samples, and using HumanMethylation450 Bead Chips, we measured genome-wide methylation levels at 482,397 CpG loci in umbilical cord blood and retained 394,460 loci after quality control. After adjustment for batch effects, cell types, and covariates, we used robust linear regression models to examine associations of prenatal lead exposure with DNA methylation in cord blood at epigenome-wide significance level [false discovery rate (FDR)<0.05]. RESULTS: The mean [standard deviation (SD)] maternal RBC lead level was 1.22 (0.63) µg/dL. CpG cg10773601 showed an epigenome-wide significant negative association with prenatal lead exposure (-1.4% per doubling increase in lead exposure; p=2.3×10-7) and was annotated to C-Type Lectin Domain Family 11, Member A (CLEC11A), which functions as a growth factor for primitive hematopoietic progenitor cells. In sex-specific analyses, we identified more CpGs with FDR<0.05 among female infants (n=38) than among male infants (n=2). One CpG (cg24637308), which showed a strong negative association with prenatal lead exposure among female infants (-4.3% per doubling increase in lead exposure; p=1.1×10-06), was annotated to Dynein Heavy Chain Domain 1 gene (DNHD1) which is highly expressed in human brain. Interestingly, there were strong correlations between blood and brain methylation for CpG (cg24637308) based on another independent set of samples with a high proportion of female participants. CONCLUSION: Prenatal low-level lead exposure was associated with newborn DNA methylation, particularly in female infants. https://doi.org/10.1289/EHP1246.

Persistent DNA methylation changes associated with prenatal mercury exposure and cognitive performance during childhood.

Prenatal exposure to mercury, a known neurotoxic metal, is associated with lower cognitive performance during childhood. Disruption of fetal epigenetic programming could explain mercury's neurodevelopmental effects. We screened for epigenome-wide methylation differences associated with maternal prenatal blood mercury levels in 321 cord blood DNA samples and examined the persistence of these alterations during early (n = 75; 2.9-4.9 years) and mid-childhood (n = 291; 6.7-10.5 years). Among males, prenatal mercury levels were associated with lower regional cord blood DNA methylation at the Paraoxonase 1 gene (PON1) that persisted in early childhood and was attenuated in mid-childhood blood. Cord blood methylation at the PON1 locus predicted lower cognitive test scores measured during early childhood. Methylation at the PON1 locus was associated with PON1 expression in an independent set of cord blood samples. The observed persistent epigenetic disruption of the PON1 gene may modulate mercury toxicity in humans and might serve as a biomarker of exposure and disease susceptibility.

DNA methylation signatures of chronic low-grade inflammation are associated with complex diseases.

BACKGROUND: Chronic low-grade inflammation reflects a subclinical immune response implicated in the pathogenesis of complex diseases. Identifying genetic loci where DNA methylation is associated with chronic low-grade inflammation may reveal novel pathways or therapeutic targets for inflammation. RESULTS: We performed a meta-analysis of epigenome-wide association studies (EWAS) of serum C-reactive protein (CRP), which is a sensitive marker of low-grade inflammation, in a large European population (n = 8863) and trans-ethnic replication in African Americans (n = 4111). We found differential methylation at 218 CpG sites to be associated with CRP (P < 1.15 × 10-7) in the discovery panel of European ancestry and replicated (P < 2.29 × 10-4) 58 CpG sites (45 unique loci) among African Americans. To further characterize the molecular and clinical relevance of the findings, we examined the association with gene expression, genetic sequence variants, and clinical outcomes. DNA methylation at nine (16%) CpG sites was associated with whole blood gene expression in cis (P < 8.47 × 10-5), ten (17%) CpG sites were associated with a nearby genetic variant (P < 2.50 × 10-3), and 51 (88%) were also associated with at least one related cardiometabolic entity (P < 9.58 × 10-5). An additive weighted score of replicated CpG sites accounted for up to 6% inter-individual variation (R2) of age-adjusted and sex-adjusted CRP, independent of known CRP-related genetic variants. CONCLUSION: We have completed an EWAS of chronic low-grade inflammation and identified many novel genetic loci underlying inflammation that may serve as targets for the development of novel therapeutic interventions for inflammation.

Birth weight-for-gestational age is associated with DNA methylation at birth and in childhood.

BACKGROUND: Both higher and lower fetal growth are associated with cardio-metabolic health later in life, suggesting that prenatal developmental programming determines long-term cardiovascular disease risk. Epigenetic mechanisms, which orchestrate fetal growth and development, may offer insight on the early programming of health and disease. We investigated whether birth weight-for-gestational is associated with DNA methylation at birth and mid-childhood, measured via the Infinium 450K array. METHODS/RESULTS: Participants were from Project Viva, a pre-birth cohort of pregnant women and their children in Eastern Massachusetts. After exclusion of participants with maternal type 1 or 2 diabetes and gestational age <34 weeks, we used DNA methylation assays from 476 venous umbilical cord blood samples and a subset of 235 who additionally had peripheral blood samples available in mid-childhood (age 7-10 years). Among 392,918 CpG sites analyzed, birth weight-for-gestational age z-score was associated with cord blood DNA methylation at 34 CpGs (false discovery rate P < 0.05), after adjusting for maternal age, race/ethnicity, education, smoking, parity, delivery mode, pre-pregnancy BMI, gestational diabetes status, child sex, and estimated cord blood cell proportions based on a cord blood reference panel. Two of these CpGs were previously reported in epigenome-wide analyses of birth weight, and several other CpGs map to genes relevant to fetal growth and development. Namely, higher birth weight-for-gestational age was associated with higher methylation at four CpGs at the PBX1 locus (e.g., ß (95% CI) for lead signal at cg06750897 = 1.9 (1.2, 2.6)), which encodes a transcription factor that regulates embryonic development. Birth weight-for-gestational age was also associated with mid-childhood blood DNA methylation at four of the 34 CpGs identified in cord blood analyses, including sites at the PBX1 locus described. CONCLUSIONS: We identified CpG sites where birth weight-for-gestational age was associated with DNA methylation at birth, and for a subset of these sites, birth weight-for-gestational age was also associated with DNA methylation at mid-childhood.

Epigenetic Mediators Between Childhood Socioeconomic Disadvantage and Mid-Life Body Mass Index: The New England Family Study.

OBJECTIVE: Childhood socioeconomic disadvantage is associated with adulthood obesity risk; however, epigenetic mechanisms are poorly understood. This work's objective was to evaluate whether associations of childhood socioeconomic disadvantage with adulthood body mass index (BMI) are mediated by DNA methylation. METHODS: Participants were 141 men and women from the New England Family Study, prospectively followed prenatally through a mean age of 47 years. Epigenomewide DNA methylation was evaluated in peripheral blood and adipose tissue obtained at adulthood, using the Infinium HumanMethylation450K BeadChip. Childhood socioeconomic status (SES) at age 7 years was assessed directly from parents' reports. Offspring adiposity was directly assessed using BMI at a mean age of 47 years. Associations of SES, DNA methylation, and BMI were estimated using least square estimators. Statistical mediation analyses were performed using joint significance test and bootstrapping. RESULTS: Of CpG sites significant at the 25% false discovery rate level in epigenomewide methylation BMI analyses, 91 sites in men and 71 sites in women were additionally significant for SES-methylation associations (p < .001) in adipose tissue. Many involved genes biologically relevant for development of obesity, including fatty acid synthase, transmembrane protein 88, signal transducer and activator of transcription 3, and neuritin 1. There was no evidence of epigenetic mediation in peripheral blood leukocytes. CONCLUSIONS: DNA methylation at specific genes may be mediators of associations between childhood socioeconomic disadvantage and mid-life BMI in adipose tissue. Findings motivate continued efforts to study if and how childhood socioeconomic disadvantage is biologically embedded at the level of the epigenome in regions etiologically relevant for adiposity.

The Role of DNA Methylation in Cardiovascular Risk and Disease: Methodological Aspects, Study Design, and Data Analysis for Epidemiological Studies.

Epidemiological studies have demonstrated that genetic, environmental, behavioral, and clinical factors contribute to cardiovascular disease development. How these risk factors interact at the cellular level to cause cardiovascular disease is not well known. Epigenetic epidemiology enables researchers to explore critical links between genomic coding, modifiable exposures, and manifestation of disease phenotype. One epigenetic link, DNA methylation, is potentially an important mechanism underlying these associations. In the past decade, there has been a significant increase in the number of epidemiological studies investigating cardiovascular risk factors and outcomes in relation to DNA methylation, but many gaps remain in our understanding of the underlying cause and biological implications. In this review, we provide a brief overview of the biology and mechanisms of DNA methylation and its role in cardiovascular disease. In addition, we summarize the current evidence base in epigenetic epidemiology studies relevant to cardiovascular health and disease and discuss the limitations, challenges, and future directions of the field. Finally, we provide guidelines for well-designed epigenetic epidemiology studies, with particular focus on methodological aspects, study design, and analytical challenges.

Cardiovascular Health and Incident Cardiovascular Disease and Cancer: The Women's Health Initiative.

INTRODUCTION: The American Heart Association's "Simple 7" offers a practical public health conceptualization of cardiovascular health (CVH). CVH predicts incident cardiovascular disease (CVD) in younger populations, but has not been studied in a large, diverse population of aging postmenopausal women. The extent to which CVH predicts cancer in postmenopausal women is unknown. METHODS: Multivariable Cox regression estimated hazard ratios and 95% CIs for the association between CVH and incident CVD, any cancer, and cancer subtypes (lung, colorectal, and breast) among 161,809 Women's Health Initiative observational study and clinical trial participants followed from 1993 through 2010. Data were analyzed in 2013. CVH score was characterized as the number (0 [worst] to 7 [best]) of the American Heart Association's ideal CVH behaviors and factors at baseline: smoking, BMI, physical activity, diet, total cholesterol, blood pressure, and fasting glucose. RESULTS: Median follow-up was approximately 13 years. Fewer minorities and less educated women achieved ideal CVH, a common benchmark. In adjusted models, compared with women with the highest (best) CVH scores, those with the lowest (worst) CVH scores had nearly seven times the hazard of incident CVD (6.83, 95% CI=5.83, 8.00) and 52% greater risk of incident cancer (1.52, 95% CI=1.35, 1.72). Ideal CVH was most strongly inversely associated with lung cancer, then colorectal cancer, and then breast cancer. CONCLUSIONS: Lower ideal CVH is more common among minority and less educated postmenopausal women and predicts increased risk of CVD and cancer in this population, emphasizing the importance of prevention efforts among vulnerable older adults.

Adiposity is associated with DNA methylation profile in adipose tissue.

BACKGROUND: Adiposity is a risk factor for type 2 diabetes and cardiovascular disease, suggesting an important role for adipose tissue in the development of these conditions. The epigenetic underpinnings of adiposity are not well understood, and studies of DNA methylation in relation to adiposity have rarely focused on target adipose tissue. Objectives were to evaluate whether genome-wide DNA methylation profiles in subcutaneous adipose tissue and peripheral blood leukocytes are associated with measures of adiposity, including central fat mass, body fat distribution and body mass index. METHODS: Participants were 106 men and women (mean age 47 years) from the New England Family Study. DNA methylation was evaluated using the Infinium HumanMethylation450K BeadChip. Adiposity phenotypes included dual-energy X-ray absorptiometry-assessed android fat mass, android:gynoid fat ratio and trunk:limb fat ratio, as well as body mass index. RESULTS: Adipose tissue genome-wide DNA methylation profiles were associated with all four adiposity phenotypes, after adjusting for race, sex and current smoking (omnibus p-values <0.001). After further adjustment for adipose cell-mixture effects, associations with android fat mass, android:gynoid fat ratio, and trunk:limb fat ratio remained. In gene-specific analyses, adiposity phenotypes were associated with adipose tissue DNA methylation in several genes that are biologically relevant to the development of adiposity, such as AOC3, LIPE, SOD3, AQP7 and CETP. Blood DNA methylation profiles were not associated with adiposity, before or after adjustment for blood leukocyte cell mixture effects. CONCLUSION: Findings show that DNA methylation patterns in adipose tissue are associated with adiposity.

Healthy lifestyle and decreasing risk of heart failure in women: the Women's Health Initiative observational study.

BACKGROUND: The impact of a healthy lifestyle on risk of heart failure (HF) is not well known. OBJECTIVES: The objectives of this study were to evaluate the effect of a combination of lifestyle factors on incident HF and to further investigate whether weighting each lifestyle factor has additional impact. METHODS: Participants were 84,537 post-menopausal women from the WHI (Women's Health Initiative) observational study, free of self-reported HF at baseline. A healthy lifestyle score (HL score) was created wherein women received 1 point for each healthy criterion met: high-scoring Alternative Healthy Eating Index, physically active, healthy body mass index, and currently not smoking. A weighted score (wHL score) was also created in which each lifestyle factor was weighted according to its independent magnitude of effect on HF. The incidence of hospitalized HF was determined by trained adjudicators using standardized methodology. RESULTS: There were 1,826 HF cases over a mean follow-up of 11 years. HL score was strongly associated with risk of HF (multivariable-adjusted hazard ratio [HR] [95% confidence interval (CI)] 0.49 [95% CI: 0.38 to 0.62], 0.36 [95% CI: 0.28 to 0.46], 0.24 [95% CI: 0.19 to 0.31], and 0.23 [95% CI: 0.17 to 0.30] for HL score of 1, 2, 3, and 4 vs. 0, respectively). The HL score and wHL score were similarly associated with HF risk (HR: 0.46 [95% CI: 0.41 to 0.52] for HL score; HR: 0.48 [95% CI: 0.42 to 0.55] for wHL score, comparing the highest tertile to the lowest). The HL score was also strongly associated with HF risk among women without antecedent coronary heart disease, diabetes, or hypertension. CONCLUSIONS: An increasingly healthy lifestyle was associated with decreasing HF risk among post-menopausal women, even in the absence of antecedent coronary heart disease, hypertension, and diabetes. Weighting the lifestyle factors had minimal impact.