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Background: Pinpointing genetic impacts on DNA methylation can improve our understanding of pathways that underlie gene regulation and disease risk., Results: We report heritability and methylation quantitative trait locus (meQTL) analysis at 724,499 CpGs profiled with the Illumina Infinium MethylationEPIC array in 2358 blood samples from three UK cohorts. Methylation levels at 34.2% of CpGs are affected by SNPs, and 98% of effects are cis-acting or within 1 Mbp of the tested CpG. Our results are consistent with meQTL analyses based on the former Illumina Infinium HumanMethylation450 array. Both SNPs and CpGs with meQTLs are overrepresented in enhancers, which have improved coverage on this platform compared to previous approaches. Co-localisation analyses across genetic effects on DNA methylation and 56 human traits identify 1520 co-localisations across 1325 unique CpGs and 34 phenotypes, including in disease-relevant genes, such as USP1 and DOCK7 (total cholesterol levels), and ICOSLG (inflammatory bowel disease). Enrichment analysis of meQTLs and integration with expression QTLs give insights into mechanisms underlying cis-meQTLs (e.g. through disruption of transcription factor binding sites for CTCF and SMC3) and trans-meQTLs (e.g. through regulating the expression of ACD and SENP7 which can modulate DNA methylation at distal sites)., Conclusions: Our findings improve the characterisation of the mechanisms underlying DNA methylation variability and are informative for prioritisation of GWAS variants for functional follow-ups. The MeQTL EPIC Database and viewer are available online at https://epicmeqtl.kcl.ac.uk ., (© 2023. The Author(s).)

DNA methylation variations are prevalent in human obesity but evidence of a causative role in disease pathogenesis is limited. Here, we combine epigenome-wide association and integrative genomics to investigate the impact of adipocyte DNA methylation variations in human obesity. We discover extensive DNA methylation changes that are robustly associated with obesity (N = 190 samples, 691 loci in subcutaneous and 173 loci in visceral adipocytes, P < 1 × 10-7). We connect obesity-associated methylation variations to transcriptomic changes at >500 target genes, and identify putative methylation-transcription factor interactions. Through Mendelian Randomisation, we infer causal effects of methylation on obesity and obesity-induced metabolic disturbances at 59 independent loci. Targeted methylation sequencing, CRISPR-activation and gene silencing in adipocytes, further identifies regional methylation variations, underlying regulatory elements and novel cellular metabolic effects. Our results indicate DNA methylation is an important determinant of human obesity and its metabolic complications, and reveal mechanisms through which altered methylation may impact adipocyte functions., (© 2023. The Author(s).)

Rationale: Methylation integrates factors present at birth and modifiable across the lifespan that can influence pulmonary function. Studies are limited in scope and replication. Objectives: To conduct large-scale epigenome-wide meta-analyses of blood DNA methylation and pulmonary function. Methods: Twelve cohorts analyzed associations of methylation at cytosine-phosphate-guanine probes (CpGs), using Illumina 450K or EPIC/850K arrays, with FEV 1 , FVC, and FEV 1 /FVC. We performed multiancestry epigenome-wide meta-analyses (total of 17,503 individuals; 14,761 European, 2,549 African, and 193 Hispanic/Latino ancestries) and interpreted results using integrative epigenomics. Measurements and Main Results: We identified 1,267 CpGs (1,042 genes) differentially methylated (false discovery rate, <0.025) in relation to FEV 1 , FVC, or FEV 1 /FVC, including 1,240 novel and 73 also related to chronic obstructive pulmonary disease (1,787 cases). We found 294 CpGs unique to European or African ancestry and 395 CpGs unique to never or ever smokers. The majority of significant CpGs correlated with nearby gene expression in blood. Findings were enriched in key regulatory elements for gene function, including accessible chromatin elements, in both blood and lung. Sixty-nine implicated genes are targets of investigational or approved drugs. One example novel gene highlighted by integrative epigenomic and druggable target analysis is TNFRSF4. Mendelian randomization and colocalization analyses suggest that epigenome-wide association study signals capture causal regulatory genomic loci. Conclusions: We identified numerous novel loci differentially methylated in relation to pulmonary function; few were detected in large genome-wide association studies. Integrative analyses highlight functional relevance and potential therapeutic targets. This comprehensive discovery of potentially modifiable, novel lung function loci expands knowledge gained from genetic studies, providing insights into lung pathogenesis.

Background: Ageing biomarkers can help us better understand how well-established socioeconomic position (SEP) disparities in ageing occur. A promising new set of DNAm methylation (DNAm)-based ageing biomarkers indicate through their age acceleration (AA) measures if biological ageing is slower or faster than chronological ageing. Few studies have investigated the association between SEP and DNAm AA., Methods: We used linear regression to examine the sex-adjusted relationships between childhood social class, adult social class, intergenerational social class change, education and adult household earnings with first (Horvath AA and Hannum AA) and second generation (PhenoAge AA and GrimAge AA) DNAm AA markers using data from the MRC National Survey of Health and Development., Results: In the first-generation biomarkers, there was little evidence of any associations with Horvath AA but associations of childhood social class and income with Hannum AA were observed. Strong associations were seen between greater disadvantage in childhood and adult SEP and greater AA in the second generation biomarkers. For example, those with fathers in an unskilled occupational social class in childhood had 3.6 years greater PhenoAge AA (95% CI 1.8 to 5.4) than those with fathers from a professional social class. Individuals without qualifications had higher AA compared with those with higher education (4.1 years greater GrimAge AA (95% CI 3.1 to 5.0))., Conclusion: Our findings highlight the importance of exposure to social disadvantage in childhood to the biological ageing process. The second generation clocks appear to be more sensitive to the accumulation of social disadvantage across the life course., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2021. Re-use permitted under CC BY. Published by BMJ.)

Characterizing genetic influences on DNA methylation (DNAm) provides an opportunity to understand mechanisms underpinning gene regulation and disease. In the present study, we describe results of DNAm quantitative trait locus (mQTL) analyses on 32,851 participants, identifying genetic variants associated with DNAm at 420,509 DNAm sites in blood. We present a database of >270,000 independent mQTLs, of which 8.5% comprise long-range (trans) associations. Identified mQTL associations explain 15-17% of the additive genetic variance of DNAm. We show that the genetic architecture of DNAm levels is highly polygenic. Using shared genetic control between distal DNAm sites, we constructed networks, identifying 405 discrete genomic communities enriched for genomic annotations and complex traits. Shared genetic variants are associated with both DNAm levels and complex diseases, but only in a minority of cases do these associations reflect causal relationships from DNAm to trait or vice versa, indicating a more complex genotype-phenotype map than previously anticipated., (© 2021. The Author(s), under exclusive licence to Springer Nature America, Inc.)

Background: In the first study of its kind, we examine the association between growth and development in early life and DNAm age biomarkers in mid-life., Methods: Participants were from the Medical Research Council National Survey of Health and Development (n = 1376). Four DNAm age acceleration (AgeAccel) biomarkers were measured when participants were aged 53 years: AgeAccelHannum; AgeAccelHorvath; AgeAccelLevine; and AgeAccelGrim. Exposure variables included: relative weight gain (standardised residuals from models of current weight z-score on current height, and previous weight and height z-scores); and linear growth (standardised residuals from models of current height z-score on previous height and weight z-scores) during infancy (0-2 years, weight gain only), early childhood (2-4 years), middle childhood (4-7 years) and late childhood to adolescence (7-15 years); age at menarche; and pubertal stage for men at 14-15 years. The relationship between relative weight gain and linear growth and AgeAccel was investigated using conditional growth models. We replicated analyses from the late childhood to adolescence period and pubertal timing among 240 participants from The National Child and Development Study (NCDS)., Results: A 1SD increase in relative weight gain in late childhood to adolescence was associated with 0.50 years (95% CI 0.20, 0.79) higher AgeAccelGrim. Although the CI includes the null, the estimate was similar in NCDS [0.57 years (95% CI - 0.01, 1.16)] There was no strong evidence that relative weight gain and linear growth in childhood was associated with any other AgeAccel biomarker. There was no relationship between pubertal timing in men and AgeAccel biomarkers. Women who reached menarche ≥ 12 years had 1.20 years (95% CI 0.15, 2.24) higher AgeAccelGrim on average than women who reached menarche < 12 years; however, this was not replicated in NCDS and was not statistically significant after Bonferroni correction., Conclusions: Our findings generally do not support an association between growth and AgeAccel biomarkers in mid-life. However, we found rapid weight gain during pubertal development, previously related to higher cardiovascular disease risk, to be associated with older AgeAccelGrim. Given this is an exploratory study, this finding requires replication., (© 2021. The Author(s).)

Objective: To investigate whether epigenetic profiles of mural granulosa cells (MGC) and leukocytes from women with diminished ovarian reserve (DOR) differ from those of women with normal or high ovarian reserve., Design: Prospectively collected material from a multicenter cohort of women undergoing fertility treatment., Setting: Private and university-based facilities for clinical services and research., Patient(s): One hundred and nineteen women of various ages and ovarian reserve status (antimüllerian hormone level) who provided blood samples and MGC., Intervention(s): None., Main Outcome Measure(s): Measures of epigenetic aging rates from whole-genome methylation array data: DNA methylation variability, age acceleration, DNA methylation telomere length estimator (DNAmTL), and accumulation of epimutations., Result(s): Comparison of DOR or high ovarian reserve samples to controls (normal ovarian reserve) showed differential methylation variability between DOR and normal samples at 4,199 CpGs in MGC, and 447 between high and normal (false-discovery rate < 0.05). Variable sites in MGC from DOR were enriched in regions marked with the repressive histone modification H3K27me3, and also included genes involved in folliculogenesis, such as insulin growth factor 2 (IGF2) and antimüllerian hormone (AMH). Regardless of ovarian reserve, very few signals were detected in leukocytes, and no overlaps with those in MGC were found. Furthermore, we found a higher number of epimutations in MGC from women with DOR (Kruskal-Wallis test, difference in mean = 3,485)., Conclusion(s): The somatic cells of human ovarian follicles have a distinctive epigenetic profile in women with DOR. A high frequency of epimutations suggests premature aging. Ovarian reserve status was not reflected in the leukocyte epigenetic profile., (Copyright © 2020. Published by Elsevier Inc.)

Advancing maternal age causes a progressive reduction in fertility. The decline in developmental competence of the oocyte with age is likely to be a consequence of multiple contributory factors. Loss of epigenetic quality of the oocyte could impair early developmental events or programme adverse outcomes in offspring that manifest only later in life. Here, we undertake joint profiling of the transcriptome and DNA methylome of individual oocytes from reproductively young and old mice undergoing natural ovulation. We find reduced complexity as well as increased variance in the transcriptome of oocytes from aged females. This transcriptome heterogeneity is reflected in the identification of discrete sub-populations. Oocytes with a transcriptome characteristic of immature chromatin configuration (NSN) clustered into two groups: one with reduced developmental competence, as indicated by lower expression of maternal effect genes, and one with a young-like transcriptome. Oocytes from older females had on average reduced CpG methylation, but the characteristic bimodal methylation landscape of the oocyte was preserved. Germline differentially methylated regions of imprinted genes were appropriately methylated irrespective of age. For the majority of differentially expressed transcripts, the absence of correlated methylation changes suggests a post-transcriptional basis for most age-related effects on the transcriptome. However, we did find differences in gene body methylation at which there were corresponding changes in gene expression, indicating age-related effects on transcription that translate into methylation differences. Interestingly, oocytes varied in expression and methylation of these genes, which could contribute to variable competence of oocytes or penetrance of maternal age-related phenotypes in offspring., (© 2020 The Authors. Aging Cell published by the Anatomical Society and John Wiley & Sons Ltd.)

DNA methylation has fundamental roles in gene programming and aging that may help predict mortality. However, no large-scale study has investigated whether site-specific DNA methylation predicts all-cause mortality. We used the Illumina-HumanMethylation450-BeadChip to identify blood DNA methylation sites associated with all-cause mortality for 12, 300 participants in 12 Cohorts of the Heart and Aging Research in Genetic Epidemiology (CHARGE) Consortium. Over an average 10-year follow-up, there were 2,561 deaths across the cohorts. Nine sites mapping to three intergenic and six gene-specific regions were associated with mortality ( P < 9.3x10 -7 ) independently of age and other mortality predictors. Six sites (cg14866069, cg23666362, cg20045320, cg07839457, cg07677157, cg09615688)-mapping respectively to BMPR1B, MIR1973, IFITM3, NLRC5 , and two intergenic regions-were associated with reduced mortality risk. The remaining three sites (cg17086398, cg12619262, cg18424841)-mapping respectively to SERINC2, CHST12 , and an intergenic region-were associated with increased mortality risk. DNA methylation at each site predicted 5%-15% of all deaths. We also assessed the causal association of those sites to age-related chronic diseases by using Mendelian randomization, identifying weak causal relationship between cg18424841 and cg09615688 with coronary heart disease. Of the nine sites, three (cg20045320, cg07839457, cg07677157) were associated with lower incidence of heart disease risk and two (cg20045320, cg07839457) with smoking and inflammation in prior CHARGE analyses. Methylation of cg20045320, cg07839457, and cg17086398 was associated with decreased expression of nearby genes ( IFITM3, IRF, NLRC5, MT1, MT2, MARCKSL1 ) linked to immune responses and cardiometabolic diseases. These sites may serve as useful clinical tools for mortality risk assessment and preventative care.

Background: DNA methylation (DNAm) age acceleration (AgeAccel) has been shown to be predictive of all-cause mortality but it is unclear what functional aspect(s) of aging it captures. We examine associations between four measures of AgeAccel in adults aged 45-87 years and physical and cognitive performance and their age-related decline., Methods: AgeAccelHannum, AgeAccelHorvath, AgeAccelPheno, and AgeAccelGrim were calculated in the Medical Research Council National Survey of Health and Development (NSHD), National Child Development Study (NCDS) and TwinsUK. Three measures of physical (grip strength, chair rise speed, and forced expiratory volume in one second [FEV1]) and two measures of cognitive (episodic memory and mental speed) performance were assessed., Results: AgeAccelPheno and AgeAccelGrim, but not AgeAccelHannum and AgeAccelHorvath were related to performance in random effects meta-analyses (n = 1,388-1,685). For example, a 1-year increase in AgeAccelPheno or AgeAccelGrim was associated with a 0.01 mL (95% confidence interval [CI]: 0.01, 0.02) or 0.03 mL (95% CI: 0.01, 0.05) lower mean FEV1 respectively. In NSHD, AgeAccelPheno and AgeAccelGrim at 53 years were associated with age-related decline in performance between 53 and 69 years as tested by linear mixed models (p < .05). In a subset of NSHD participants (n = 482), there was little evidence that change in any AgeAccel measure was associated with change in performance conditional on baseline performance., Conclusions: We found little evidence to support associations between the first generation of DNAm-based biomarkers of aging and age-related physical or cognitive performance in midlife to early old age. However, there was evidence that the second generation biomarkers, AgeAccelPheno and AgeAccelGrim, could act as makers of an individual's healthspan as proposed., (© The Author(s) 2019. Published by Oxford University Press on behalf of The Gerontological Society of America.)

Background: Genetic and environmental risk factors contribute to periodontal disease, but the underlying susceptibility pathways are not fully understood. Epigenetic mechanisms are malleable regulators of gene function that can change in response to genetic and environmental stimuli, thereby providing a potential mechanism for mediating risk effects in periodontitis. The aim of this study is to identify epigenetic changes across tissues that are associated with periodontal disease., Methods: Self-reported gingival bleeding and history of gum disease, or tooth mobility, were used as indicators of periodontal disease. DNA methylation profiles were generated using the Infinium HumanMethylation450 BeadChip in whole blood, buccal, and adipose tissue samples from predominantly older female twins (mean age 58) from the TwinsUK cohort. Epigenome-wide association scans (EWAS) of gingival bleeding and tooth mobility were conducted in whole blood in 528 and 492 twins, respectively. Subsequently, targeted candidate gene analysis at 28 genomic regions was carried out testing for phenotype-methylation associations in 41 (tooth mobility) and 43 (gingival bleeding) buccal, and 501 (tooth mobility) and 556 (gingival bleeding) adipose DNA samples., Results: Epigenome-wide analyses in blood identified one CpG-site (cg21245277 in ZNF804A) associated with gingival bleeding (FDR = 0.03, nominal p value = 7.17e-8) and 58 sites associated with tooth mobility (FDR < 0.05) with the top signals in IQCE and XKR6. Epigenetic variation at 28 candidate regions (247 CpG-sites) for chronic periodontitis showed an enrichment for association with periodontal traits, and signals in eight genes (VDR, IL6ST, TMCO6, IL1RN, CD44, IL1B, WHAMM, and CXCL1) were significant in both traits. The methylation-phenotype association signals validated in buccal samples, and a subset (25%) also validated in adipose tissue., Conclusions: Epigenome-wide analyses in adult female twins identified specific DNA methylation changes linked to self-reported periodontal disease. Future work will explore the environmental basis and functional impact of these results to infer potential for strategic personalized treatments and prevention of chronic periodontitis.

The lack of reliable measures of alcohol intake is a major obstacle to the diagnosis and treatment of alcohol-related diseases. Epigenetic modifications such as DNA methylation may provide novel biomarkers of alcohol use. To examine this possibility, we performed an epigenome-wide association study of methylation of cytosine-phosphate-guanine dinucleotide (CpG) sites in relation to alcohol intake in 13 population-based cohorts (n total =13 317; 54% women; mean age across cohorts 42-76 years) using whole blood (9643 European and 2423 African ancestries) or monocyte-derived DNA (588 European, 263 African and 400 Hispanic ancestry) samples. We performed meta-analysis and variable selection in whole-blood samples of people of European ancestry (n=6926) and identified 144 CpGs that provided substantial discrimination (area under the curve=0.90-0.99) for current heavy alcohol intake (⩾42 g per day in men and ⩾28 g per day in women) in four replication cohorts. The ancestry-stratified meta-analysis in whole blood identified 328 (9643 European ancestry samples) and 165 (2423 African ancestry samples) alcohol-related CpGs at Bonferroni-adjusted P<1 × 10 -7 . Analysis of the monocyte-derived DNA (n=1251) identified 62 alcohol-related CpGs at P<1 × 10 -7 . In whole-blood samples of people of European ancestry, we detected differential methylation in two neurotransmitter receptor genes, the γ-Aminobutyric acid-A receptor delta and γ-aminobutyric acid B receptor subunit 1; their differential methylation was associated with expression levels of a number of genes involved in immune function. In conclusion, we have identified a robust alcohol-related DNA methylation signature and shown the potential utility of DNA methylation as a clinically useful diagnostic test to detect current heavy alcohol consumption.

Study Question: Does women's age affect the DNA methylation (DNAm) profile differently in mural granulosa cells (MGCs) from other somatic cells?Summary Answer: Accumulation of epimutations by age and a higher number of age-related differentially methylated regions (DMR) in MGCs were found compared to leukocytes from the same woman, suggesting that the MGCs have a distinctive epigenetic profile.What Is Known Already: The mechanisms underlying the decline in women's fertility from the mid-30s remain to be fully elucidated. The DNAm age of many healthy tissues changes predictably with and follows chronological age, but DNAm age in some reproductive tissues has been shown to depart from chronological age (older: endometrium; younger: cumulus cells, spermatozoa).Study Design, Size, Duration: This study is a multicenter cohort study based on retrospective analysis of prospectively collected data and material derived from healthy women undergoing IVF or ICSI treatment following ovarian stimulation with antagonist protocol. One hundred and nineteen women were included from September 2016 to June 2018 from four clinics in Denmark and Sweden.Participants/materials, Setting, Methods: Blood samples were obtained from 118 healthy women with varying ovarian reserve status. MGCs were collected from 63 of the 119 women by isolation from pooled follicles immediately after oocyte retrieval. DNA from leukocytes and MGCs was extracted and analysed with a genome-wide methylation array. Data from the methylation array were processed using the ENmix package. Subsequently, DNAm age was calculated using established and tailored age predictors and DMRs were analysed with the DMRcate package.Main Results and Role Of Chance: Using established age predictors, DNAm age in MGCs was found to be considerable younger and constant (average: 2.7 years) compared to chronological age (average: 33.9 years). A Granulosa Cell clock able to predict the age of both MGCs (average: 32.4 years) and leukocytes (average: 38.8 years) was successfully developed. MGCs differed from leukocytes in having a higher number of epimutations (P = 0.003) but predicted telomere lengths unaffected by age (Pearson's correlation coefficient = -0.1, P = 0.47). DMRs associated with age (age-DMRs) were identified in MGCs (n = 335) and in leukocytes (n = 1) with a significant enrichment in MGCs for genes involved in RNA processing (45 genes, P = 3.96 × 10-08) and gene expression (152 genes, P = 2.3 × 10-06). The top age-DMRs included the metastable epiallele VTRNA2-1, the DNAm regulator ZFP57 and the anti-Müllerian hormone (AMH) gene. The apparent discordance between different epigenetic measures of age in MGCs suggests that they reflect difference stages in the MGC life cycle.Large Scale Data: N/A.Limitations, Reasons For Caution: No gene expression data were available to associate with the epigenetic findings. The MGCs are collected during ovarian stimulation, which may influence DNAm; however, no correlation between FSH dose and number of epimutations was found.Wider Implications Of the Findings: Our findings underline that the somatic compartment of the follicle follows a different methylation trajectory with age than other somatic cells. The higher number of epimutations and age-DMRs in MGCs suggest that their function is affected by age.Study Funding/competing Interest(s): This project is part of ReproUnion collaborative study, co-financed by the European Union, Interreg V ÖKS, the Danish National Research Foundation and the European Research Council. The authors declare no conflict of interest. [ABSTRACT FROM AUTHOR]