DNA Methylation Patterns In Blood Research Articles

Epigenetic Study of Cohort of Monozygotic Twins With Hypertrophic Cardiomyopathy Due to MYBPC3 (Cardiac Myosin-Binding Protein C).

Hypertrophic cardiomyopathy is an autosomal dominant cardiac disease. The mechanisms that determine its variable expressivity are poorly understood. Epigenetics could play a crucial role in bridging the gap between genotype and phenotype by orchestrating the interplay between the environment and the genome regulation. In this study we aimed to establish a possible correlation between the peripheral blood DNA methylation patterns and left ventricular hypertrophy severity in patients with hypertrophic cardiomyopathy, evaluating the potential impact of lifestyle variables and providing a biological context to the observed changes. Methylation data were obtained from peripheral blood samples (Infinium MethylationEPIC BeadChip arrays). We employed multiple pair-matched models to extract genomic positions whose methylation correlates with the degree of left ventricular hypertrophy in 3 monozygotic twin pairs carrying the same founder pathogenic variant (MYBPC3 p.Gly263Ter). This model enables the isolation of the environmental influence, beyond age, on DNA methylation changes by removing the genetic background. Our results revealed a more anxious personality among more severely affected individuals. We identified 56 differentially methylated positions that exhibited moderate, proportional changes in methylation associated with left ventricular hypertrophy. These differentially methylated positions were enriched in regions regulated by repressor histone marks and tended to cluster at genes involved in left ventricular hypertrophy development, such as HOXA5, TRPC3, UCN3, or PLSCR2, suggesting that changes in peripheral blood may reflect myocardial alterations. We present a unique pair-matched model, based on 3 monozygotic twin pairs carrying the same founder pathogenic variant and different phenotypes. This study provides further evidence of the pivotal role of epigenetics in hypertrophic cardiomyopathy variable expressivity.

Blood DNA methylomic signatures associated with CSF biomarkers of Alzheimer's disease in the EMIF-AD study.

We investigated blood DNA methylation patterns associated with 15 well-established cerebrospinal fluid (CSF) biomarkers of Alzheimer's disease (AD) pathophysiology, neuroinflammation, and neurodegeneration. We assessed DNA methylation in 885 blood samples from the European Medical Information Framework for Alzheimer's Disease (EMIF-AD) study using the EPIC array. We identified Bonferroni-significant differential methylation associated with CSF YKL-40 (five loci) and neurofilament light chain (NfL; seven loci) levels, with two of the loci associated with CSF YKL-40 levels correlating with plasma YKL-40 levels. A co-localization analysis showed shared genetic variants underlying YKL-40 DNA methylation and CSF protein levels, with evidence that DNA methylation mediates the association between genotype and protein levels. Weighted gene correlation network analysis identified two modules of co-methylated loci correlated with several amyloid measures and enriched in pathways associated with lipoproteins and development. We conducted the most comprehensive epigenome-wide association study (EWAS) of AD-relevant CSF biomarkers to date. Future work should explore the relationship between YKL-40 genotype, DNA methylation, and protein levels in the brain. Blood DNA methylation was assessed in the EMIF-AD MBD study. Epigenome-wide association studies (EWASs) were performed for 15 Alzheimer's disease (AD)-relevant cerebrospinal fluid (CSF) biomarker measures. Five Bonferroni-significant loci were associated with YKL-40 levels and seven with neurofilament light chain (NfL). DNA methylation in YKL-40 co-localized with previously reported genetic variation. DNA methylation potentially mediates the effect ofsingle-nucleotide polymorphisms (SNPs) in YKL-40 on CSF protein levels.

Capturing sex-specific and hypofertility-linked effects of assisted reproductive technologies on the cord blood DNA methylome

BackgroundChildren conceived through assisted reproduction are at an increased risk for growth and genomic imprinting disorders, often linked to DNA methylation defects. It has been suggested that assisted reproductive technology (ART) and underlying parental infertility can induce epigenetic instability, specifically interfering with DNA methylation reprogramming events during germ cell and preimplantation development. To date, human studies exploring the association between ART and DNA methylation defects have reported inconsistent or inconclusive results, likely due to population heterogeneity and the use of technologies with limited coverage of the epigenome. In our study, we explored the epigenetic risk of ART by comprehensively profiling the DNA methylome of 73 human cord blood samples of singleton pregnancies (n = 36 control group, n = 37 ART/hypofertile group) from a human prospective longitudinal birth cohort, the 3D (Design, Develop, Discover) Study, using a high-resolution sequencing-based custom capture panel that examines over 2.4 million autosomal CpGs in the genome.ResultsWe identified evidence of sex-specific effects of ART/hypofertility on cord blood DNA methylation patterns. Our genome-wide analyses identified ~ 46% more CpGs affected by ART/hypofertility in female than in male infant cord blood. We performed a detailed analysis of three imprinted genes which have been associated with altered DNA methylation following ART (KCNQ1OT1, H19/IGF2 and GNAS) and found that female infant cord blood was associated with DNA hypomethylation. When compared to less invasive procedures such as intrauterine insemination, more invasive ARTs (in vitro fertilization, intracytoplasmic sperm injection, embryo culture) resulted in more marked and distinct effects on the cord blood DNA methylome. In the in vitro group, we found a close to fourfold higher proportion of significantly enriched Gene Ontology terms involved in development than in the in vivo group.ConclusionsOur study highlights the ability of a sensitive, targeted, sequencing-based approach to uncover DNA methylation perturbations in cord blood associated with hypofertility and ART and influenced by offspring sex and ART technique invasiveness.

Blood DNA Methylation Patterns in Older Adults With Evolving Dementia.

Dementia and cognitive disorders are major aging-associated pathologies. The prevalence and severity of these conditions are influenced by both genetic and environmental factors. Reflecting this, epigenetic alterations have been associated with each of these processes, especially at the level of DNA methylation, and such changes may help explain the observed interindividual variability in the development of the 2 pathologies. However, the importance of epigenetic alterations in explaining their etiology is unclear because little is known about the timing of when they appear. Here, using Illumina MethylationEPIC arrays, we have longitudinally analyzed the peripheral blood methylomes of cognitively healthy older adults (>70 year), some of whom went on to develop dementia while others stayed healthy. We have characterized 34 individuals at the prediagnosis stage and at a 4-year follow-up in the postdiagnosis stage (total n = 68). Our results show multiple DNA methylation alterations linked to dementia status, particularly at the level of differentially methylated regions. These loci are associated with several dementia-related genes, including PON1, AP2A2, MAGI2, POT1, ITGAX, PACSIN1, SLC2A8, and EIF4E. We also provide validation of the previously reported epigenetic alteration of HOXB6 and PM20D1. Importantly, we show that most of these regions are already altered in the prediagnosis stage of individuals who go on to develop dementia. In conclusion, our observations suggest that dementia-associated epigenetic patterns that have specific biological features are already present before diagnosis, and thus may be important in the design of epigenetic biomarkers for disease detection based on peripheral tissues.

The Whole Blood DNA Methylation Patterns of Extrinsic Apoptotic Signaling Pathway Related Genes in Autoimmune Thyroiditis among Areas with Different Iodine Levels.

Autoimmune thyroiditis (AIT) has a complex etiology and the susceptibility to it is determined by a combination of genetic and environmental factors, although these are not yet fully understood. The present research aimed to explore the DNA methylation patterns in whole blood of extrinsic apoptotic signaling pathway related genes in AIT among areas with different iodine levels. We selected the iodine-fortification areas (IFA), iodine-adequate areas (IAA) and water-based iodine-excess areas (IEA) from Shandong Province of China as survey sites. Totally 176 AIT cases and 176 controls were included. MethylTargetTM and QT-PCR technology were used to detect candidate genes' DNA methylation levels and mRNA expression levels, respectively. We found that DAPK1 DNA methylation levels in AIT cases (especially in female) were significantly higher than controls (t=2.7715, P=0.0059; t=2.4638, P=0.0143 in female). There were differences in DAPK1(t=2.5384, P=0.0121), TNFSF8(t=2.1667, P=0.0334) and TNFAIP8(t=2.5672, P=0.0121) genes methylation between cases and controls with different water iodine levels. The mRNA expression of DAPK1(t=4.329, P<0.001) and TNFAIP8(t=3.775, P<0.001) in the cases were increased. We identified the differences in the DNA methylation status of the extrinsic apoptotic signaling pathway related genes between AIT and controls and in different iodine levels areas. The results were verified at the mRNA level. The environmental iodine may affect DNA methylation to some extent.

Longitudinal Changes in Epigenetic Age Acceleration in Aviremic Human Immunodeficiency Virus-Infected Recipients of Long-term Antiretroviral Treatment.

Human immunodeficiency virus (HIV) infection induces epigenetic age acceleration (EAA), but it remains unclear whether epigenetic aging continues to accelerate during successful antiretroviral therapy (ART) and prolonged virological suppression. We longitudinally analyzed 63 long-term aviremic HIV-infected adults. Using blood DNA methylation patterns, we calculated EAA measures based on 3 epigenetic clocks (Horvath's clock, PhenoAge, and GrimAge). We recorded the emergence of serious AIDS-related and non-AIDS-related events throughout the study to assess its association with EAA. All participants were on stable ART and were virologically suppressed. After 4 years of follow-up, PhenoAge-EAA and GrimAge-EAA showed no differences, whereas Horvath-EAA slightly decreased (median difference, -0.53 years; P = .015). Longitudinal changes in EAA measures were independent of changes in CD4 cell counts, the ART regimen, or other HIV-related factors. Nineteen percent of participants experienced a serious clinical event during the study. Horvath-EAA was significantly higher at baseline in participants with clinical events (P = .027). After adjusting for confounders, we found a trend toward an association of higher levels of all EAA measures at baseline with serious clinical events. Epigenetic aging did not accelerate in long-term aviremic HIV-infected adults after 4 years of successful ART. EAA measures deserve further study as potential tools for predicting clinical events.

Identification of epigenome-wide DNA methylation differences between carriers of APOE \u03b54 and APOE \u03b52 alleles

BackgroundThe apolipoprotein E (APOE) ε4 allele is the strongest genetic risk factor for late onset Alzheimer’s disease, whilst the ε2 allele confers protection. Previous studies report differential DNA methylation of APOE between ε4 and ε2 carriers, but associations with epigenome-wide methylation have not previously been characterised.MethodsUsing the EPIC array, we investigated epigenome-wide differences in whole blood DNA methylation patterns between Alzheimer’s disease-free APOE ε4 (n = 2469) and ε2 (n = 1118) carriers from the two largest single-cohort DNA methylation samples profiled to date. Using a discovery, replication and meta-analysis study design, methylation differences were identified using epigenome-wide association analysis and differentially methylated region (DMR) approaches. Results were explored using pathway and methylation quantitative trait loci (meQTL) analyses.ResultsWe obtained replicated evidence for DNA methylation differences in a ~ 169 kb region, which encompasses part of APOE and several upstream genes. Meta-analytic approaches identified DNA methylation differences outside of APOE: differentially methylated positions were identified in DHCR24, LDLR and ABCG1 (2.59 × 10−100 ≤ P ≤ 2.44 × 10−8) and DMRs were identified in SREBF2 and LDLR (1.63 × 10−4 ≤ P ≤ 3.01 × 10−2). Pathway and meQTL analyses implicated lipid-related processes and high-density lipoprotein cholesterol was identified as a partial mediator of the methylation differences in ABCG1 and DHCR24.ConclusionsAPOE ε4 vs. ε2 carrier status is associated with epigenome-wide methylation differences in the blood. The loci identified are located in trans as well as cis to APOE and implicate genes involved in lipid homeostasis.

Antibiotic Use During Pregnancy and the Maternal Epigenome

Background: Approximately 40% of women are prescribed an antibiotic during pregnancy. Though completed course of therapy is necessary for treatment of infections, antibiotic use has been associated with several adverse birth outcomes. Biological changes in the prenatal environment affected by antibiotic use are poorly understood. Specifically, examining maternal epigenetic patterns associated with antibiotic use can identify CpG sites that may provide insight into biological pathways associated with adverse birth outcomes.Objectives: To evaluate the association of antibiotic use, abstracted from medical records as participants who had ever been prescribed an antibiotic during pregnancy, and maternal peripheral blood DNA methylation patterns during pregnancy in the MADRES (Maternal and Developmental Risks from Environmental and Social Stressors) study, a pregnancy cohort of predominantly lower income Hispanic women in Los Angeles. A paired, longitudinal analysis was used to evaluate changes in methylation between the first and third trimester in women who had ever been prescribed an antibiotic during pregnancy. Exploration of discrete effects at the first and third trimester was also undertaken, as contraindication of antibiotic differs across trimesters of pregnancy.Methods: Antibiotic use, indicated by prescription of antibiotic, and its association with differentially methylated positions (DMPs) on the Illumina EPIC array were analyzed using linear mixed and linear regression models. Estimated blood cell type fractions and surrogate variables were used to control for both measured and unmeasured confounding variables in statistical analysis.Results: In the third trimester, 15 DMPs were identified (PFDR&lt;0.05) in relation to antibiotic use during pregnancy. Antibiotic use during pregnancy was not associated with first to third trimester methylation changes for any loci on the EPIC array.Conclusion: In the MADRES cohort, antibiotic use during pregnancy was not associated with changes in methylation during pregnancy, but was associated with differential methylation at 15 CpGs in third trimester peripheral blood samples.

Genome-wide identification of genes regulating DNA methylation using genetic anchors for causal inference

BackgroundDNA methylation is a key epigenetic modification in human development and disease, yet there is limited understanding of its highly coordinated regulation. Here, we identify 818 genes that affect DNA methylation patterns in blood using large-scale population genomics data.ResultsBy employing genetic instruments as causal anchors, we establish directed associations between gene expression and distant DNA methylation levels, while ensuring specificity of the associations by correcting for linkage disequilibrium and pleiotropy among neighboring genes. The identified genes are enriched for transcription factors, of which many consistently increased or decreased DNA methylation levels at multiple CpG sites. In addition, we show that a substantial number of transcription factors affected DNA methylation at their experimentally determined binding sites. We also observe genes encoding proteins with heterogenous functions that have widespread effects on DNA methylation, e.g., NFKBIE, CDCA7(L), and NLRC5, and for several examples, we suggest plausible mechanisms underlying their effect on DNA methylation.ConclusionWe report hundreds of genes that affect DNA methylation and provide key insights in the principles underlying epigenetic regulation.

On the Use of Blood Samples for Measuring DNA Methylation in Ecological Epigenetic Studies.

SynopsisThere is increasing interest in understanding the potential for epigenetic factors to contribute to phenotypic diversity in evolutionary biology. One well studied epigenetic mechanism is DNA methylation, the addition of a methyl group to cytosines, which have the potential to alter gene expression depending on the genomic region in which it takes place. Obtaining information about DNA methylation at genome-wide scale has become straightforward with the use of bisulfite treatment in combination with reduced representation or whole-genome sequencing. While it is well recognized that methylation is tissue specific, a frequent limitation for many studies is that sampling-specific tissues may require sacrificing individuals, something which is generally undesirable and sometimes impossible. Instead, information about DNA methylation patterns in the blood is frequently used as a proxy tissue. This can obviously be problematic if methylation patterns in the blood do not reflect that in the relevant tissue. Understanding how, or if, DNA methylation in blood reflect DNA methylation patterns in other tissues is therefore of utmost importance if we are to make inferences about how observed differences in methylation or temporal changes in methylation can contribute to phenotypic variation. The aim of this review is to examine what we know about the potential for using blood samples in ecological epigenetic studies. I briefly outline some methods by which we can measure DNA methylation before I examine studies that have compared DNA methylation patterns across different tissues and, finally, examine how useful blood samples may be for ecological studies of DNA methylation. Ecological epigenetic studies are in their infancy, but it is paramount for the field to move forward to have detailed information about tissue and time dependence relationships in methylation to gain insights into if blood DNA methylation patterns can be a reliable bioindicator for changes in methylation that generate phenotypic variation in ecologically important traits.

Abstract C047: Internal smoking dose is associated with specific blood DNA methylation patterns across race/ethnicity: The Multiethnic Cohort Study

Abstract Background: Lung cancer is the most common cancer in the U.S. and leading cause of cancer-related death. We demonstrated in the Multiethnic Cohort Study that for the same number of cigarettes smoked, Native Hawaiians and African Americans have the highest risk compared to whites, while Japanese Americans and Latinos are at lower risk of disease. We showed that internal smoking dose (as measured by total nicotine equivalents (TNE)) per cigarette differs across race/ethnicity; in part explaining why African Americans have a higher risk of disease and Japanese Americans have a lower risk. DNA methylation of CpG sites from cigarette smoking is one of the most common epigenetic modifications linked to lifestyle. Although many smoking-related DNA methylated CpG sites have been identified, these studies have been primarily conducted in populations of European ancestry. Moreover, the influence of internal smoking dose on the epigenome across populations has not been investigated. Here, we report on an epigenome-wide association study for urinary TNE, an optimal marker of internal smoking dose that is not biased by self-report and reflects the interindividual variation in nicotine metabolism. Methods: This study includes 1,996 current smokers at time of specimen collection, from five racial/ethnic groups: African American (n=364), European American (n=398), Japanese American (n=523), Native Hawaiian (n=311) and Latino (n=400). Genome-wide DNA methylation in blood leukocytes was measured using the Illumina MethEPIC BeadChip. Models were adjusted for age at biospecimen collection, genetic ancestry, sex, and cell-type. Results: TNE was associated with the differential methylation levels of 1,178 probes (Bonferoni corrected p&amp;lt;5 × 10-8). The top 5 overall significant associations were in AHRR (cg05575921; p=1.58 × 10^-131 and cg23576855; p=7.64 × 10^-72), 2q37.1 near ALPPL2 (cg21566642; p=6.29 × 10^-81), RARA (cg17739917; p=1.41 × 10^-74) and PRSS23 (cg14391737; p=1.42 × 10^-71). These had previously been associated with smoking status. By race/ethnicity, level of significance and the number of significantly differentially methylated probes differed. For instance, in African Americans the second most significantly associated probe was cg25845814, in ELMSAN1, (p=1.19 × 10^-10). This probe was also statistically significant in Japanese Americans (p=1.69 × 10^-13) and Latinos (p=2.93 × 10^-16), but not in European Americans (p=1.33 × 10^-6) and Native Hawaiians (p=1.48 × 10^-6). Cg25845814 has been found to regulate miRNA MIR1268A, which may modify hepatocellular carcinoma risk. European Americans had the greatest number of differentially methylated probes (n=68), followed by Latinos (n=63), Japanese Americans (n=63), African Americans (n=10) and Native Hawaiians (n=9). Conclusions: Smoking dose, as measured by TNE, may differentially impact DNA methylation of leukocytes by race/ethnicity. These differences may help to explain the population differences in smoking-related lung cancer risk. Citation Format: Sungshim L Park, Yesha Patel, Lenora W.M. Loo, Annette Lum-Jones, Maarit Tiirikainen, Sharon Murphy, Kimberly Siegmund, Daniel O. Stram, Loic Le Marchand. Internal smoking dose is associated with specific blood DNA methylation patterns across race/ethnicity: The Multiethnic Cohort Study [abstract]. In: Proceedings of the Twelfth AACR Conference on the Science of Cancer Health Disparities in Racial/Ethnic Minorities and the Medically Underserved; 2019 Sep 20-23; San Francisco, CA. Philadelphia (PA): AACR; Cancer Epidemiol Biomarkers Prev 2020;29(6 Suppl_2):Abstract nr C047.

A restricted spectrum of missense KMT2D variants cause a multiple malformations disorder distinct fromKabuki syndrome

PurposeTo investigate if specific exon 38 or 39 KMT2D missense variants (MVs) cause a condition distinct from Kabuki syndrome type 1 (KS1). MethodsMultiple individuals, with MVs in exons 38 or 39 of KMT2D that encode a highly conserved region of 54 amino acids flanked by Val3527 and Lys3583, were identified and phenotyped. Functional tests were performed to study their pathogenicity and understand the disease mechanism. ResultsThe consistent clinical features of the affected individuals, from seven unrelated families, included choanal atresia, athelia or hypoplastic nipples, branchial sinus abnormalities, neck pits, lacrimal duct anomalies, hearing loss, external ear malformations, and thyroid abnormalities. None of the individuals had intellectual disability. The frequency of clinical features, objective software-based facial analysis metrics, and genome-wide peripheral blood DNA methylation patterns in these patients were significantly different from that of KS1. Circular dichroism spectroscopy indicated that these MVs perturb KMT2D secondary structure through an increased disordered to ɑ-helical transition. ConclusionKMT2D MVs located in a specific region spanning exons 38 and 39 and affecting highly conserved residues cause a novel multiple malformations syndrome distinct from KS1. Unlike KMT2D haploinsufficiency in KS1, these MVs likely result in disease through a dominant negative mechanism.

15 LONGITUDINAL EVALUATION OF MUCOSAL DNA METHYLATION IN ULCERATIVE COLITIS SHOWS DISEASE-SPECIFIC CHANGES

Abstract Background Although inflammatory bowel disease (IBD) is heritable, the heritability is still largely unexplained despite substantial progress through GWAS and WGS. However, recent advances in epigenetics (gene environmental interactions), including DNA methylation (DNAm) analysis, has identified new disease-specific mechanisms controlling gene expression. We recently showed that IBD-associated blood DNAm patterns strongly correlated with inflammation (CRP as surrogate marker) and reverted to patterns seen in healthy controls following treatment, regardless of whether or not the underlying IBD was in remission, suggesting blood DNAm is more representative of the systemic inflammatory status rather than disease status (Gastroenterology, 2019, 156 (8): 2254–2265). Thus, we aimed to study DNAm patterns in rectal biopsies hypothesizing that DNAm in rectal biopsies will reflect disease status rather than systemic inflammatory status alone. Method Genome-wide DNA methylation was measured using the Illumina MethylationEPIC array on rectal biopsy DNA samples of 85 non-inflammatory, non-IBD controls and 215 newly diagnosed ulcerative colitis (UC) patients prior to therapy, along with 49 one-year follow-up UC cases (PROTECT cohort) to identify disease specific methylation patterns. Epigenome wide association studies (EWAS) were performed with UC and CRP as the outcomes in linear models adjusted for age, gender, race and five genotype-based principal components. Results At diagnosis, 2446 disease-specific CpG sites in rectal tissue (FDR&amp;lt;0.05) were identified. At baseline, the disease-associated DNAm signature in rectal tissue is distinct from our previous study from blood as only 15 CpGs were common between rectal tissue DNAm and blood DNAm. In contrast to what was observed in blood DNAm where the initial DNAm signature reverted back to control levels upon treatment, rectal tissue DNAm signatures remained persistent during follow-up (Figure 1). The majority of the disease-specific CpG sites identified in rectal biopsies showed a strong positive correlation with CRP. This evidence suggests that the treatment affects systemic measures of inflammation more strongly than disease tissues. Conclusion When studied longitudinally, the UC-specific DNA methylation patterns in rectal tissue are distinct from blood samples. In contrast to blood DNAm which normalizes after therapy during follow-up, rectal tissue DNAm changes persist after treatment in UC. This suggests the currently available therapies control the systemic inflammation effectively, but have less direct effect on the disease itself. Future therapies targeting disease-specific DNAm may be more effective in disease management and long-term remission.

Association of cord blood methylation with neonatal leptin: An epigenome wide association study.

BackgroundNeonatal adiposity is a risk factor for childhood obesity. Investigating contributors to neonatal adiposity is important for understanding early life obesity risk. Epigenetic changes of metabolic genes in cord blood may contribute to excessive neonatal adiposity and subsequent childhood obesity. This study aims to evaluate the association of cord blood DNA methylation patterns with anthropometric measures and cord blood leptin, a biomarker of neonatal adiposity.MethodsA cross-sectional study was performed on a multiethnic cohort of 114 full term neonates born to mothers without gestational diabetes at a university hospital. Cord blood was assayed for leptin and for epigenome-wide DNA methylation profiles via the Illumina 450K platform. Neonatal body composition was measured by air displacement plethysmography. Multivariable linear regression was used to analyze associations between individual CpG sites as well as differentially methylated regions in cord blood DNA with measures of newborn adiposity including anthropometrics (birth weight, fat mass and percent body fat) and cord blood leptin. False discovery rate was estimated to account for multiple comparisons.Results247 CpG sites as well as 18 differentially methylated gene regions were associated with cord blood leptin but no epigenetic changes were associated with birth weight, fat mass or percent body fat. Genes of interest identified in this study are DNAJA4, TFR2, SMAD3, PLAG1, FGF1, and HNF4A.ConclusionEpigenetic changes in cord blood DNA are associated with cord blood leptin levels, a measure of neonatal adiposity.

Altered Genome-Wide DNA Methylation in Peripheral Blood of South African Women with Gestational Diabetes Mellitus.

Increasing evidence implicate altered DNA methylation in the pathophysiology of gestational diabetes mellitus (GDM). This exploratory study probed the association between GDM and peripheral blood DNA methylation patterns in South African women. Genome-wide DNA methylation profiling was conducted in women with (n = 12) or without (n = 12) GDM using the Illumina Infinium HumanMethylationEPIC BeadChip array. Functional analysis of differentially methylated genes was conducted using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses. A total of 1046 CpG sites (associated with 939 genes) were differentially methylated between GDM and non-GDM groups. Enriched pathways included GDM-related pathways such as insulin resistance, glucose metabolism and inflammation. DNA methylation of the top five CpG loci showed distinct methylation patterns in GDM and non-GDM groups and was correlated with glucose concentrations. Of these, one CpG site mapped to the calmodulin-binding transcription activator 1 (CAMTA1) gene, which have been shown to regulate insulin production and secretion and may offer potential as an epigenetic biomarker in our population. Further validation using pyrosequencing and conducting longitudinal studies in large sample sizes and in different populations are required to investigate their candidacy as biomarkers of GDM.

24 - PREDICTING THE FUTURE DISEASE STATUS OF DEPRESSED PATIENTS FROM DNA METHYLATION PATTERNS IN BLOOD

Background Major Depressive Disorder (MDD) is a severe disorder with a lifetime prevalence of 15%. Recurrent or chronic MDD, which typically requires long-term treatment, occurs in the majority of patients. As previous efforts to predict recurrence/chronicity have met with limited success, novel avenues are required. In this study, we aimed to predict the future disease status of MDD patients from DNA methylation patterns in blood. Methods We assayed 28 million methylation sites in 581 MDD patients from the Netherlands Study of Depression and Anxiety (NESDA). A Machine Learning algorithm condensed all information into a single predictor labeled methylation risk score (MRS). The outcome was MDD disease status six years later. To evaluate the predictive power of our MRS, we obtained an unbiased estimate of the area under the curve (AUC) using k-fold cross validation. Results The AUC of the MRS was 0.724. We compared our MRS with predictions based on a set of five putative MDD biomarkers (e.g., assaying neurotrophic factors or inflammation), genome-wide genetic variant data, and 27 clinical, demographic or lifestyle variables (e.g., MDD symptom severity, childhood trauma, alcohol use). The MRS not only outperformed all these predictors but also seemed to capture their predictive power as the inclusion of any of these sets did not significantly increase the AUC of the MRS. Discussion The current study suggests a novel avenue for predicting future MDD status using DNA methylation patterns in blood. The predictive power of the MRS is comparable to the AUC of the Framingham Risk Score, one of the most widely used clinical tools to predict coronary heart disease. This AUC level can potentially support clinical decisions about treatment strategies by providing empirical information about the likelihood MDD is chronic or will recur in the future.

Interaction between prenatal pesticide exposure and a common polymorphism in the PON1 gene on DNA methylation in genes associated with cardio-metabolic disease risk\u2014an exploratory\xa0study

BackgroundPrenatal environmental conditions may influence disease risk in later life. We previously found a gene-environment interaction between the paraoxonase 1 (PON1) Q192R genotype and prenatal pesticide exposure leading to an adverse cardio-metabolic risk profile at school age. However, the molecular mechanisms involved have not yet been resolved. It was hypothesized that epigenetics might be involved. The aim of the present study was therefore to investigate whether DNA methylation patterns in blood cells were related to prenatal pesticide exposure level, PON1 Q192R genotype, and associated metabolic effects observed in the children.MethodsWhole blood DNA methylation patterns in 48 children (6–11 years of age), whose mothers were occupationally unexposed or exposed to pesticides early in pregnancy, were determined by Illumina 450 K methylation arrays.ResultsA specific methylation profile was observed in prenatally pesticide exposed children carrying the PON1 192R-allele. Differentially methylated genes were enriched in several neuroendocrine signaling pathways including dopamine-DARPP32 feedback (appetite, reward pathways), corticotrophin releasing hormone signaling, nNOS, neuregulin signaling, mTOR signaling, and type II diabetes mellitus signaling. Furthermore, we were able to identify possible candidate genes which mediated the associations between pesticide exposure and increased leptin level, body fat percentage, and difference in BMI Z score between birth and school age.ConclusionsDNA methylation may be an underlying mechanism explaining an adverse cardio-metabolic health profile in children carrying the PON1 192R-allele and prenatally exposed to pesticides.

Genome-wide identification of blood DNA methylation patterns associated with early-onset hepatocellular carcinoma development in hepatitis B carriers.

The etiology of early-onset hepatocellular carcinoma (HCC) among hepatitis B virus (HBV) carriers remains unclear. DNA methylation levels in peripheral leukocytes have been associated with different environmental exposures and immune or inflammatory response. We aimed to identify methylation signatures of peripheral leukocytes that could track hepatitis B progression to HCC, especially for early-onset HCC. We first performed an epigenome-wide association analysis on 48 matched case-control pairs in a nested case-control study within a 22-yr follow-up cohort of HBV carriers. Through this analysis we found that progression to early-onset HCC involved methylation variable positions across the genome, in which a substantial proportion displayed significant variation due to HBV viral load, chronic hepatitis status, and/or leukocyte subtype composition, and these associations were significantly enriched among genes in immune pathways. Methylation at probes cg00300879, cg06872964, and cg07080864, that are located within the proximal promoter of CNKSR1, IFI44L, and PENK, respectively, was validated by bisulfite pyrosequencing and findings were replicated in a case-sibling study of early-onset HCC (134 cases vs. 174 sibling controls). Furthermore, a high methylation score, constructed using the three probes, was predictive for the risk of early-onset HCC in two datasets (adjusted-odds ratios = 0.21-0.32, P ≤ 0.0206). This association was also observed for late-onset HCC (adjusted-odds ratio = 0.42-0.47, P ≤ 0.0194) in a nested case-control study (120 cases vs. 178 controls). In prospective analysis, change in the score was detected 5-9 yr before HCC onset. Blood-based methylation profiling provides new insights into the complexity of virus-host interaction underlying HBV-related HCC, holding promise for the disease risk management. © 2016 Wiley Periodicals, Inc.

DNA methylation changes of whole blood cells in response to active smoking exposure in adults: a systematic review of DNA methylation studies.

Active smoking is a major preventable public health problem and an established critical factor for epigenetic modification. In this systematic review, we identified 17 studies addressing the association of active smoking exposure with methylation modifications in blood DNA, including 14 recent epigenome-wide association studies (EWASs) and 3 gene-specific methylation studies (GSMSs) on the gene regions identified by EWASs. Overall, 1460 smoking-associated CpG sites were identified in the EWASs, of which 62 sites were detected in multiple (≥3) studies. The three most frequently reported CpG sites (genes) in whole blood samples were cg05575921 (AHRR), cg03636183 (F2RL3), and cg19859270 (GPR15), followed by other loci within intergenic regions 2q37.1 and 6p21.33. These significant smoking-related genes were further assessed by specific methylation assays in three GSMSs and reflected not only current but also lifetime or long-term exposure to active smoking. In conclusion, this review summarizes the evidences for the use of blood DNA methylation patterns as biomarkers of smoking exposure for research and clinical practice. In particular, it provides a reservoir for constructing a smoking exposure index score which could be used to more precisely quantify long-term smoking exposure and evaluate the risks of smoking-induced diseases.Electronic supplementary materialThe online version of this article (doi:10.1186/s13148-015-0148-3) contains supplementary material, which is available to authorized users.

Non-Steroidal Anti-Inflammatory Drug Use and Genomic DNA Methylation in Blood.

BackgroundNon-steroidal anti-inflammatory drug (NSAID) use is associated with decreased risk of some cancers. NSAID use modulates the epigenetic profile of normal colonic epithelium and may reduce risk of colon cancer through this pathway; however, the effect of NSAID use on the DNA methylation profile of other tissues including whole blood has not yet been examined.FindingsUsing the Sister Study cohort, we examined the association between NSAID usage and whole genome methylation patterns in blood DNA. Blood DNA methylation status across 27,589 CpG sites was evaluated for 871 women using the Illumina Infinium HumanMethylation27 Beadchip, and in a non-overlapping replication sample of 187 women at 485,512 CpG sites using the Infinium HumanMethylation450 Beadchip. We identified a number of CpG sites that were differentially methylated in regular, long-term users of NSAIDs in the discovery group, but none of these sites were statistically significant in our replication group.ConclusionsWe found no replicable methylation differences in blood related to NSAID usage. If NSAID use does effect blood DNA methylation patterns, differences are likely small.