Systematic evaluation and validation of reference and library selection methods for deconvolution of cord blood DNA methylation data

Background: Umbilical cord blood (UCB) is commonly used in epigenome-wide association studies of prenatal exposures. Accounting for cell type composition is critical in such studies as it reduces confounding due to the cell specificity of DNA methylation (DNAm). In the absence of cell sorting information, statistical methods can be applied to deconvolve heterogeneous cell mixtures. Among these methods, reference-based approaches leverage age-appropriate cell-specific DNAm profiles to estimate cellular composition. In UCB, four reference datasets comprising DNAm signatures profiled in purified cell populations have been published using the Illumina 450 K and EPIC arrays. These datasets are biologically and technically different, and currently, there is no consensus on how to best apply them. Here, we systematically evaluate and compare these datasets and provide recommendations for reference-based UCB deconvolution. Results: We first evaluated the four reference datasets to ascertain both the purity of the samples and the potential cell cross-contamination. We filtered samples and combined datasets to obtain a joint UCB reference. We selected deconvolution libraries using two different approaches: automatic selection using the top differentially methylated probes from the function pickCompProbes in minfi and a standardized library selected using the IDOL (Identifying Optimal Libraries) iterative algorithm. We compared the performance of each reference separately and in combination, using the two approaches for reference library selection, and validated the results in an independent cohort (Generation R Study, n = 191) with matched Fluorescence-Activated Cell Sorting measured cell counts. Strict filtering and combination of the references significantly improved the accuracy and efficiency of cell type estimates. Ultimately, the IDOL library outperformed the library from the automatic selection method implemented in pickCompProbes. Conclusion: These results have important implications for epigenetic studies in UCB as implementing this method will optimally reduce confounding due to cellular heterogeneity. This work provides guidelines for future referencebased UCB deconvolution and establishes a framework for combining reference datasets in other tissues

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 \u3c 1.15 × 10–7) in the discovery panel of European ancestry and replicated (P \u3c 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 \u3c 8.47 × 10–5), ten (17%) CpG sites were associated with a nearby genetic variant (P \u3c 2.50 × 10–3), and 51 (88%) were also associated with at least one related cardiometabolic entity (P \u3c 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

High pre-diagnosis inflammation-related risk score associated with decreased ovarian cancer survival

BACKGROUND: There is suggestive evidence that inflammation is related to ovarian cancer survival. However, more research is needed to identify inflammation-related factors that are associated with ovarian cancer survival and to determine their combined effects. METHODS: This analysis used pooled data on 8,147 women with invasive epithelial ovarian cancer from the Ovarian Cancer Association Consortium. Pre-diagnosis inflammatory-related exposures of interest included alcohol use, aspirin use, other nonsteroidal anti-inflammatory drug use, body mass index, environmental tobacco smoke exposure, history of pelvic inflammatory disease, polycystic ovarian syndrome, and endometriosis, menopausal hormone therapy use, physical inactivity, smoking status, and talc use. Using Cox proportional hazards (PH) models, the relationship between each exposure and survival was assessed in 50% of the data. A weighted inflammation-related risk score (IRRS) was developed and its association with survival was assessed using Cox PH models in the remaining 50% of the data. RESULTS: There was a statistically significant trend of increasing risk of death per quartile of the IRRS (HR=1.09, 95% CI 1.03-1.14). Women in the upper quartile of the IRRS had 31% higher death rate compared to the lowest quartile (95% CI 1.11-1.54). CONCLUSIONS: A higher pre-diagnosis IRRS was associated with increased mortality risk after an ovarian cancer diagnosis. Further investigation is warranted to evaluate whether post-diagnosis exposures are also associated with survival. IMPACT: Given that pre- and post-diagnosis exposures are often correlated and many are modifiable, our study results can ultimately motivate the development of behavioral recommendations to enhance survival among ovarian cancer patients

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 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

Meta-analysis of epigenome-wide association studies in newborns and children show widespread sex differences in blood DNA methylation

Publisher Copyright: © 2022 The AuthorsBackground: Among children, sex-specific differences in disease prevalence, age of onset, and susceptibility have been observed in health conditions including asthma, immune response, metabolic health, some pediatric and adult cancers, and psychiatric disorders. Epigenetic modifications such as DNA methylation may play a role in the sexual differences observed in diseases and other physiological traits. Methods: We performed a meta-analysis of the association of sex and cord blood DNA methylation at over 450,000 CpG sites in 8438 newborns from 17 cohorts participating in the Pregnancy And Childhood Epigenetics (PACE) Consortium. We also examined associations of child sex with DNA methylation in older children ages 5.5–10 years from 8 cohorts (n = 4268). Results: In newborn blood, sex was associated at Bonferroni level significance with differences in DNA methylation at 46,979 autosomal CpG sites (p < 1.3 × 10−7) after adjusting for white blood cell proportions and batch. Most of those sites had lower methylation levels in males than in females. Of the differentially methylated CpG sites identified in newborn blood, 68% (31,727) met look-up level significance (p < 1.1 × 10−6) in older children and had methylation differences in the same direction. Conclusions: This is a large-scale meta-analysis examining sex differences in DNA methylation in newborns and older children. Expanding upon previous studies, we replicated previous findings and identified additional autosomal sites with sex-specific differences in DNA methylation. Differentially methylated sites were enriched in genes involved in cancer, psychiatric disorders, and cardiovascular phenotypes.Peer reviewe

Meta-analysis of epigenome-wide association studies in neonates reveals widespread differential DNA methylation associated with birthweight

Birthweight is associated with health outcomes across the life course, DNA methylation may be an underlying mechanism. In this meta-analysis of epigenome-wide association studies of 8,825 neonates from 24 birth cohorts in the Pregnancy And Childhood Epigenetics Consortium, we find that DNA methylation in neonatal blood is associated with birthweight at 914 sites, with a difference in birthweight ranging from -183 to 178 grams per 10% increase in methylation (P-Bonferroni <1.06 x 10(-7)). In additional analyses in 7,278 participants,Peer reviewe

Epigenome-wide meta-analysis of prenatal maternal stressful life events and newborn DNA methylation.

This is the author accepted manuscript. The final version is available from Springer Nature via the DOI in this recordCode availability: The code used for this EWAS meta-analysis is available from the corresponding authors upon reasonable request.Prenatal maternal stressful life events are associated with adverse neurodevelopmental outcomes in offspring. Biological mechanisms underlying these associations are largely unknown, but DNA methylation likely plays a role. This meta-analysis included twelve non-overlapping cohorts from ten independent longitudinal studies (N = 5,496) within the international Pregnancy and Childhood Epigenetics consortium to examine maternal stressful life events during pregnancy and DNA methylation in cord blood. Children whose mothers reported higher levels of cumulative maternal stressful life events during pregnancy exhibited differential methylation of cg26579032 in ALKBH3. Stressor-specific domains of conflict with family/friends, abuse (physical, sexual, and emotional), and death of a close friend/relative were also associated with differential methylation of CpGs in APTX, MyD88, and both UHRF1 and SDCCAG8, respectively; these genes are implicated in neurodegeneration, immune and cellular functions, regulation of global methylation levels, metabolism, and schizophrenia risk. Thus, differences in DNA methylation at these loci may provide novel insights into potential mechanisms of neurodevelopment in offspring.Medical Research Council and Wellcome Trus