Longitudinal DNA methylation profiling of the rectal mucosa identifies cell-specific signatures of disease status, severity and clinical outcomes in ulcerative colitis cell-specific DNA methylation signatures of UC.

BACKGROUND: In peripheral blood, DNA methylation (DNAm) patterns in inflammatory bowel disease patients reflect inflammatory status rather than disease status. Here, we examined DNAm in diseased rectal mucosa from ulcerative colitis (UC) patients, focusing on constituent cell types with the goal of identifying therapeutic targets for UC other than the immune system. We profiled DNAm of rectal mucosal biopsies of pediatric UC at diagnosis (n = 211) and non-IBD control (n = 85) patients and performed epigenome-wide association studies (EWAS) of specific cell types to understand DNAm changes in epithelial, immune and fibroblast cells across disease states, course, and clinical outcomes. We also examined longitudinal analysis on follow-up samples (n = 73), and comparisons were made among patients with clinical outcomes including those undergoing colectomy versus those who did not. Additionally, we included RNA-seq from the same subjects to assess the impact of CpG sites on the transcription of nearby genes during the disease course. RESULTS: At diagnosis, UC rectal mucosa exhibited a lower proportion of epithelial cells and fibroblasts, and higher proportion of immune cells, in conjunction with variation in the DNAm pattern. While treatment had significant effects on the methylation signature of immune cells, its effects on fibroblasts and epithelial cells were attenuated. Individuals who required colectomy exhibited cell composition and DNAm patterns at follow-up more similar to disease onset than patients who did not require colectomy. Combining these results with gene expression profiles, we identify CpG sites whose methylation patterns are most consistent with a contribution to poor disease outcomes and could thus be potential therapeutic targets. CONCLUSIONS: Cell-specific epigenetic changes in the rectal mucosa in UC are associated with disease severity and outcome. Current therapeutics may more effectively target the immune than the epithelial and fibroblast compartments.

Methylation quantitative trait loci are largely consistent across disease states in Crohn's disease.

Recently, we identified 1,189 CpG sites whose DNA methylation level in blood associated with Crohn's disease. Here, we examined associations between DNA methylation and genetic variants to identify methylation quantitative trait loci across disease states in (1) 402 blood samples from 164 newly diagnosed pediatric Crohn's disease patients taken at 2 time points (diagnosis and follow-up), and 74 non-inflammatory bowel disease controls, (2) 780 blood samples from a non-Crohn's disease adult population, and (3) 40 ileal biopsies (17 Crohn's disease cases and 23 non-inflammatory bowel disease controls) from group (1). Genome-wide DNAm profiling and genotyping were performed using the Illumina MethylationEPIC and Illumina Multi-Ethnic arrays. SNP-CpG associations were identified via linear models adjusted for age, sex, disease status, disease subtype, estimated cell proportions, and genotype-based principal components. In total, we observed 535,448 SNP-CpG associations between 287,881 SNPs and 12,843 CpG sites (P < 8.21 × 10-14). Associations were highly consistent across different ages, races, disease states, and tissue types, suggesting that the majority of these methylation quantitative trait loci participate in common gene regulation. However, genes near CpGs associated with inflammatory bowel disease SNPs were enriched for 18 KEGG pathways relevant to inflammatory bowel disease-linked immune function and inflammatory responses. We observed suggestive evidence for a small number of tissue-specific associations and disease-specific associations in ileum, though larger studies will be needed to confirm these results. Our study concludes that the vast majority of blood-derived methylation quantitative trait loci are common across individuals, though a subset may be involved in processes related to Crohn's disease. Independent cohort studies will be required to validate these findings.

Epigenetic prediction of 17ß-estradiol and relationship to trauma-related outcomes in women.

17ß-estradiol (E2) levels in women correlate with multiple neuropsychiatric symptoms, including those that are stress-related. Furthermore, prior work from our group has demonstrated that E2 status influences DNA methylation (DNAm) across the genome. We developed and validated a DNAm-based predictor of E2 (one of four naturally occurring estrogens) using a training set of 183 females and a test set of 79 females from the same traumatized cohort. We showed that predicted E2 levels were highly correlated with measured E2 concentrations in our testing set (r â€‹= â€‹0.75, p â€‹= â€‹1.8e-15). We further demonstrated that predicted E2 concentrations, in combination with measured values, negatively correlated with current post-traumatic stress disorder (PTSD) (ߠ​= â€‹-0.38, p â€‹= â€‹0.01) and major depressive disorder (MDD) diagnoses (ߠ​= â€‹-0.45, p â€‹= â€‹0.02), as well as a continuous measure of PTSD symptom severity (ߠ​= â€‹-2.3, p â€‹= â€‹0.007) in females. Finally, we tested our predictor in an independent data set (n â€‹= â€‹85) also comprised of recently traumatized female subjects to determine if the predictor would generalize to a different population than the one on which it was developed. We found that the correlation between predicted and actual E2 concentrations in the external validation data set was also high (r â€‹= â€‹0.48, p â€‹= â€‹3.0e-6). While further validation is warranted, a DNAm predictor of E2 concentrations will advance our understanding of hormone-epigenetic interactions. Furthermore, such a DNAm predictor may serve as an epigenetic proxy for E2 concentrations and thus provide an important biomarker to better evaluate the contribution of E2 to current and potentially future psychiatric symptoms in samples for which E2 is not measured.

Genome-wide DNA methylation differences and polychlorinated biphenyl (PCB) exposure in a US population.

Exposure to polychlorinated biphenyls (PCBs), an endocrine-disrupting compound, is ubiquitous despite decades-old bans on the manufacture and use of PCBs. Increased exposure to PCBs is associated with adverse health consequences throughout life, including type 2 diabetes and cancer. PCB exposure is also associated with alterations in epigenetic marks and gene transcription, which could lead to adverse health outcomes, but many of these are population-specific. To further investigate the association between PCB and epigenetic marks, DNA methylation was measured at 787,684 CpG sites in 641 peripheral blood samples from the Michigan Polybrominated Biphenyl (PBB) Registry. 1345 CpGs were associated with increased total PCB level after controlling for age, sex, and 24 surrogate variables (FDR < 0.05). These CpGs were enriched in active promoter and transcription associated regions (p < 0.05), and in regions around the binding sites for transcription factors involved in xenobiotic metabolism and immune function (FDR < 0.05). PCB exposure also associated with proportions of CD4T, NK, and granulocyte cell types, and with the neutrophil to lymphocyte ratio (NLR) (p < 0.05), and the estimated effect sizes of PCB on the epigenome were correlated with the effect sizes previously reported in an epigenome-wide study of C-reactive protein (r = 0.29; p = 2.22e-5), supporting previous studies on the association between PCB and immune dysfunction. These results indicate that PCB exposure is associated with differences in epigenetic marks in active regions of the genome, and future work should investigate whether these may mediate the association between PCB and health consequences.

PTSD is associated with increased DNA methylation across regions of HLA-DPB1 and SPATC1L.

Posttraumatic stress disorder (PTSD) is characterized by intrusive thoughts, avoidance, negative alterations in cognitions and mood, and arousal symptoms that adversely affect mental and physical health. Recent evidence links changes in DNA methylation of CpG cites to PTSD. Since clusters of proximal CpGs share similar methylation signatures, identification of PTSD-associated differentially methylated regions (DMRs) may elucidate the pathways defining differential risk and resilience of PTSD. Here we aimed to identify epigenetic differences associated with PTSD. DNA methylation data profiled from blood samples using the MethylationEPIC BeadChip were used to perform a DMR analysis in 187 PTSD cases and 367 trauma-exposed controls from the Grady Trauma Project (GTP). DMRs were assessed with R package bumphunter. We identified two regions that associate with PTSD after multiple test correction. These regions were in the gene body of HLA-DPB1 and in the promoter of SPATC1L. The DMR in HLA-DPB1 was associated with PTSD in an independent cohort. Both DMRs included CpGs whose methylation associated with nearby sequence variation (meQTL) and that associated with expression of their respective genes (eQTM). This study supports an emerging literature linking PTSD risk to genetic and epigenetic variation in the HLA region.

Epigenome-wide meta-analysis of PTSD across 10 military and civilian cohorts identifies methylation changes in AHRR.

Epigenetic differences may help to distinguish between PTSD cases and trauma-exposed controls. Here, we describe the results of the largest DNA methylation meta-analysis of PTSD to date. Ten cohorts, military and civilian, contribute blood-derived DNA methylation data from 1,896 PTSD cases and trauma-exposed controls. Four CpG sites within the aryl-hydrocarbon receptor repressor (AHRR) associate with PTSD after adjustment for multiple comparisons, with lower DNA methylation in PTSD cases relative to controls. Although AHRR methylation is known to associate with smoking, the AHRR association with PTSD is most pronounced in non-smokers, suggesting the result was independent of smoking status. Evaluation of metabolomics data reveals that AHRR methylation associated with kynurenine levels, which are lower among subjects with PTSD. This study supports epigenetic differences in those with PTSD and suggests a role for decreased kynurenine as a contributor to immune dysregulation in PTSD.

Sex-specific DNA methylation differences in people exposed to polybrominated biphenyl.

Aim: Michigan residents were exposed to polybrominated biphenyls (PBBs) when it was accidentally added to the food supply. Highly exposed individuals report sex-specific health problems, but the underlying biological mechanism behind these different health risks is not known. Materials and methods: DNA methylation in blood from 381 women and 277 men with PBB exposure was analyzed with the MethylationEPIC BeadChip. Results: 675 CpGs were associated with PBBs levels in males, while only 17 CpGs were associated in females (false discovery rate <0.05). No CpGs were associated in both sexes. These CpGs were enriched in different functional regions and transcription factor binding sites in each sex. Conclusion: Exposure to PBBs may have sex-specific effects on the epigenome that may underlie sex-specific adverse health outcomes.

Evaluating the impact of trauma and PTSD on epigenetic prediction of lifespan and neural integrity.

Post-traumatic stress disorder (PTSD) is a debilitating disorder that develops in some people following trauma exposure. Trauma and PTSD have been associated with accelerated cellular aging. This study evaluated the effect of trauma and PTSD on accelerated GrimAge, an epigenetic predictor of lifespan, in traumatized civilians. This study included 218 individuals with current PTSD, 427 trauma-exposed controls without any history of PTSD and 209 subjects with lifetime PTSD history who are not categorized as current PTSD cases. The Traumatic Events Inventory (TEI) and Clinician-Administered PTSD Scale (CAPS) were used to measure lifetime trauma burden and PTSD, respectively. DNA from whole blood was interrogated using the MethylationEPIC or HumanMethylation450 BeadChips. GrimAge estimates were calculated using the methylation age calculator. Cortical thickness of 69 female subjects was assessed by using T1-weighted structural MRI images. Associations between trauma exposure, PTSD, cortical thickness, and GrimAge acceleration were tested with multiple regression models. Lifetime trauma burden (p = 0.03), current PTSD (p = 0.02) and lifetime PTSD (p = 0.005) were associated with GrimAge acceleration, indicative of a shorter predicted lifespan. The association with lifetime PTSD was replicated in an independent cohort (p = 0.04). In the MRI sub sample, GrimAge acceleration also associated with cortical atrophy in the right lateral orbitofrontal cortex (padj = 0.03) and right posterior cingulate (padj = 0.04), brain areas associated with emotion-regulation and threat-regulation. Our findings suggest that lifetime trauma and PTSD may contribute to a higher epigenetic-based mortality risk. We also demonstrate a relationship between cortical atrophy in PTSD-relevant brain regions and shorter predicted lifespan.

Critical evaluation of copy number variant calling methods using DNA methylation.

Recent technological and methodological developments have enabled the use of array-based DNA methylation data to call copy number variants (CNVs). ChAMP, Conumee, and cnAnalysis450k are popular methods currently used to call CNVs using methylation data. However, so far, no studies have analyzed the reliability of these methods using real samples. Data from a cohort of individuals with genotype and DNA methylation data generated using the HumanMethylation450 and MethylationEPIC BeadChips were used to assess the consistency between the CNV calls generated by methylation and genotype data. We also took advantage of repeated measures of methylation data collected from the same individuals to compare the reliability of CNVs called by ChAMP, Conumee, and cnAnalysis450k for both the methylation arrays. ChAMP identified more CNVs than Conumee and cnAnalysis450k for both the arrays and, as a consequence, had a higher overlap (~62%) with the calls from the genotype data. However, all methods had relatively low reliability. For the MethylationEPIC array, Conumee had the highest reliability (57.6%), whereas for the HumanMethylation450 array, cnAnalysis450k had the highest reliability (43.0%). Overall, the MethylationEPIC array provided significant gains in reliability for CNV calling over the HumanMethylation450 array but not for overlap with CNVs called using genotype data.

Environmental exposure to polybrominated biphenyl (PBB) associates with an increased rate of biological aging.

Advanced age increases risk for cancer, cardiovascular disease, and all-cause mortality. However, people do not age at the same rate, and biological age (frequently measured through DNA methylation) can be older than chronological age. Environmental factors have been associated with the rate of biological aging, but it is not known whether persistent endocrine-disrupting compounds (EDCs) like polybrominated biphenyl (PBB) would associate with age acceleration. Three different epigenetic age acceleration measures (intrinsic, extrinsic, and phenotypic) were calculated from existing epigenetic data in whole blood from a population highly exposed to PBB (N=658). Association between serum PBB concentration and these measures was tested, controlling for sex, lipid levels, and estimated cell type proportions. Higher PBB levels associated with increased age acceleration (intrinsic: ß=0.24, 95%CI=0.01-0.46, p = 0.03; extrinsic: ß=0.39, 95%CI=0.12-0.65, p = 0.004; and phenotypic: ß=0.30, 95%CI=0.05-0.54, p = 0.01). Neither age when exposed to PBB nor sex statistically interacted with PBB to predict age acceleration, but, in stratified analyses, the association between PBB and age acceleration was only in people exposed before finishing puberty and in men. This suggests that EDCs can associate with the biological aging process, and further studies are warranted to investigate other environmental pollutants' effect on aging.

Association of HLA locus alleles with posttraumatic stress disorder.

BACKGROUND: Immune dysregulation has been widely observed in those with posttraumatic stress disorder (PTSD). An individual's immune response is shaped, in part, by the highly polymorphic Human Leukocyte Antigen (HLA) locus that is associated with major psychiatric disorders such as schizophrenia, major depression and bipolar disorder. The aim of the current study was to investigate the association between common HLA alleles and PTSD. METHODS: Genome-wide association data was used to predict alleles of 7 classical polymorphic HLA genes (A, B, C, DRB1, DQA1, DQB1, DPB1) in 403 lifetime PTSD cases and 369 trauma exposed controls of African ancestry. Association of HLA allelic variations with lifetime PTSD was analyzed using logistic regression, controlling for ancestry, sex and multiple comparisons. The effect of HLA alleles on gene expression was assessed by weighted correlation network analysis (WGCNA), using 353 subjects with available expression data. Enrichment analysis was performed using anRichment to identify associated pathways of each module. RESULTS: HLA-B*58:01 (p = 0.035), HLA-C*07:01 (p = 0.035), HLA-DQA1*01:01 (p = 0.003), HLA-DQB1*05:01 (p = 0.009) and HLA-DPB1*17:01 (p = 0.017) were more common in PTSD cases, while HLA-A*02:01 (p = 0.026), HLA-DQA1*05:05 (p = 0.011) and HLA-DRB1*11:01 (p < 0.001) were more frequent in controls. WGCNA was used to explore expression patterns of the PTSD related alleles. Gene expression modules of PTSD-related HLA alleles were enriched in various pathways, including pathways related to immune and neural activity. CONCLUSIONS: To the best of our knowledge, this is the first study to report an association of HLA alleles with PTSD. Altogether, our results support the link between the immune system, brain and PTSD.

Exposure to polybrominated biphenyl and stochastic epigenetic mutations: application of a novel epigenetic approach to environmental exposure in the Michigan polybrominated biphenyl registry.

Endocrine-disrupting compounds are associated with altered epigenetic regulation and adverse health outcomes, although inconsistent results suggest that people have varied responses to the same exposure. Interpersonal variation in response to environmental exposures is not identified using standard, population-based methods. However, methods that capture an individual's response, such as analyzing stochastic epigenetic mutations (SEMs), may capture currently missed effects of environmental exposure. To test whether polybrominated biphenyl (PBB) was associated with SEMs, DNA methylation was measured using Illumina's MethylationEPIC array in PBB-exposed individuals, and SEMs were identified. Association was tested using a linear regression with robust sandwich variance estimators, controlling for age, sex, lipids, and cell types. The number of SEMs was variable (range: 119-18,309), and positively associated with age (p = 1.23e-17), but not with sex (p = 0.97). PBBs and SEMs were only positively associated in people who were older when they were exposed (p = 0.02 vs. p = 0.91). Many subjects had SEMs enriched in biological pathways, particularly in pathways involved with xenobiotic metabolism and endocrine function. Higher number of SEMs was also associated with higher age acceleration (intrinsic: p = 1.70e-3; extrinsic: p = 3.59e-11), indicating that SEMs may be associated with age-related health problems. Finding an association between environmental contaminants and higher SEMs may provide insight into individual differences in response to environmental contaminants, as well as into the biological mechanism behind SEM formation. Furthermore, these results suggest that people may be particularly vulnerable to epigenetic dysregulation from environmental exposures as they age.

Blood-Derived DNA Methylation Signatures of Crohn's Disease and Severity of Intestinal Inflammation.

BACKGROUND & AIMS: Crohn's disease is a relapsing and remitting inflammatory disorder with a variable clinical course. Although most patients present with an inflammatory phenotype (B1), approximately 20% of patients rapidly progress to complicated disease, which includes stricturing (B2), within 5 years. We analyzed DNA methylation patterns in blood samples of pediatric patients with Crohn's disease at diagnosis and later time points to identify changes that associate with and might contribute to disease development and progression. METHODS: We obtained blood samples from 164 pediatric patients (1-17 years old) with Crohn's disease (B1 or B2) who participated in a North American study and were followed for 5 years. Participants without intestinal inflammation or symptoms served as controls (n = 74). DNA methylation patterns were analyzed in samples collected at time of diagnosis and 1-3 years later at approximately 850,000 sites. We used genetic association and the concept of Mendelian randomization to identify changes in DNA methylation patterns that might contribute to the development of or result from Crohn's disease. RESULTS: We identified 1189 5'-cytosine-phosphate-guanosine-3' (CpG) sites that were differentially methylated between patients with Crohn's disease (at diagnosis) and controls. Methylation changes at these sites correlated with plasma levels of C-reactive protein. A comparison of methylation profiles of DNA collected at diagnosis of Crohn's disease vs during the follow-up period showed that, during treatment, alterations identified in methylation profiles at the time of diagnosis of Crohn's disease more closely resembled patterns observed in controls, irrespective of disease progression to B2. We identified methylation changes at 3 CpG sites that might contribute to the development of Crohn's disease. Most CpG methylation changes associated with Crohn's disease disappeared with treatment of inflammation and might be a result of Crohn's disease. CONCLUSIONS: Methylation patterns observed in blood samples from patients with Crohn's disease accompany acute inflammation; with treatment, these change to resemble methylation patterns observed in patients without intestinal inflammation. These findings indicate that Crohn's disease-associated patterns of DNA methylation observed in blood samples are a result of the inflammatory features of the disease and are less likely to contribute to disease development or progression.

Exposure to polybrominated biphenyl (PBB) associates with genome-wide DNA methylation differences in peripheral blood.

In 1973, Michigan residents were exposed to polybrominated biphenyl (PBB) when it was accidentally added to farm animal feed. Highly exposed individuals and their children have experienced endocrine-related health problems, though the underlying mechanism behind these remains unknown. We investigated whether PBB exposure is associated with variation in DNA methylation in peripheral blood samples from 658 participants of the Michigan PBB registry using the MethylationEPIC BeadChip, as well as investigated what the potential function of the affected regions are and whether these epigenetic marks are known to associate with endocrine system pathways. After multiple test correction (FDR <0.05), 1890 CpG sites associated with total PBB levels. These CpGs were not enriched in any particular biological pathway, but were enriched in enhancer and insulator regions, and depleted in regions near the transcription start site or in CpG islands (p < 0.05). They were also more likely to be in ARNT and ESR2 transcription factor binding sites (p = 3.27e-23 and p = 1.62e-6, respectively), and there was significant overlap between CpGs associated with PBB and CpGs associated with estrogen (p < 2.2e-16). PBB-associated CpGs were also enriched for CpGs known to be associated with gene expression in blood (eQTMs) (p < 0.05). These eQTMs were enriched for pathways related to immune function and endocrine-related autoimmune disease (FDR <0.05). These results indicate that exposure to PBB is associated with differences in epigenetic marks that suggest that it is acting similarly to estrogen and is associated with dysregulated immune system pathways.

Characterization of gene expression changes over healthy term pregnancies.

During pregnancy, women experience numerous physiological changes but, to date, there is limited published data that characterize accompanying changes in gene expression over pregnancy. This study sought to characterize the complexity of the transcriptome over the course of pregnancy among women with healthy pregnancies. Subjects provided a venous blood sample during early (6-15 weeks) and late (22-33 weeks) pregnancy, which was used to isolate peripheral blood mononuclear cells prior to RNA extraction. Gene expression was examined for 63 women with uncomplicated, term deliveries. We evaluated the association between weeks gestation at sample collection and expression of each transcript. Of the 16,311 transcripts evaluated, 439 changed over pregnancy after a Bonferroni correction to account for multiple comparisons. Genes whose expression increased over pregnancy were associated with oxygen transport, the immune system, and host response to bacteria. Characterization of changes in the transcriptome over the course of healthy term pregnancies may enable the identification of genes whose expression predicts complications or adverse outcomes of pregnancy.

CD4+ and CD8+ T-Cell-Specific DNA Cytosine Methylation Differences Associated With Obesity.

OBJECTIVE: Lifestyle factors associated with obesity may alter epigenome-regulated gene expression. Most studies examining epigenetic changes in obesity have analyzed DNA 5´-methylcytosine (5mC) in whole blood, representing a weighted average of several distantly related and regulated leukocyte classes. To examine leukocyte-specific differences associated with obesity, a pilot study examining 5mC in three distinct leukocyte types isolated from peripheral blood of women with normal weight and obesity was conducted. METHODS: CD4+ T cells, CD8+ T cells, and CD16+ neutrophils were reiteratively isolated from blood, and 5mC levels were measured across >450,000 CG sites. RESULTS: Nineteen CG sites were differentially methylated between women with obesity and with normal weight in CD4+ cells, 16 CG sites in CD8+ cells, and 0 CG sites in CD16+ neutrophils (q < 0.05). There were no common differentially methylated sites between the T-cell types. The amount of visceral adipose tissue was strongly associated with the methylation level of 79 CG sites in CD4+ cells, including 4 CG sites in CLSTN1's promoter, which, this study shows, may regulate its expression. CONCLUSIONS: The methylomes of various leukocytes respond differently to obesity and levels of visceral adipose tissue. Highly significant differentially methylated sites in CD4+ and CD8+ cells in women with obesity that have apparent biological relevance to obesity were identified.

SLC9B1 methylation predicts fetal intolerance of labor.

Fetal intolerance of labor is a common indication for delivery by Caesarean section. Diagnosis is based on the presence of category III fetal heart rate tracing, which is an abnormal heart tracing associated with increased likelihood of fetal hypoxia and metabolic acidemia. This study analyzed data from 177 unique women who, during their prenatal visits (7-15 weeks and/or 24-32 weeks) to Atlanta area prenatal care clinics, consented to provide blood samples for DNA methylation (HumanMethylation450 BeadChip) and gene expression (Human HT-12 v4 Expression BeadChip) analyses. We focused on 57 women aged 18-36 (mean 25.4), who had DNA methylation data available from their second prenatal visit. DNA methylation patterns at CpG sites across the genome were interrogated for associations with fetal intolerance of labor. Four CpG sites (P value <8.9 × 10-9, FDR <0.05) in gene SLC9B1, a Na+/H+ exchanger, were associated with fetal intolerance of labor. DNA methylation and gene expression were negatively associated when examined longitudinally during pregnancy using a linear mixed-effects model. Positive predictive values of methylation of these four sites ranged from 0.80 to 0.89, while negative predictive values ranged from 0.91 to 0.92. The four CpG sites were also associated with fetal intolerance of labor in an independent cohort (the Johns Hopkins Prospective PPD cohort). Therefore, fetal intolerance of labor could be accurately predicted from maternal blood samples obtained between 24-32 weeks gestation. Fetal intolerance of labor may be accurately predicted from maternal blood samples obtained between 24-32 weeks gestation by assessing DNA methylation patterns of SLC9B1. The identification of pregnant women at elevated risk for fetal intolerance of labor may allow for the development of targeted treatments or management plans.

Methylation differences reveal heterogeneity in preterm pathophysiology: results from bipartite network analyses.

BACKGROUND: Recent studies have shown that epigenetic differences can increase the risk of spontaneous preterm birth (PTB). However, little is known about heterogeneity underlying such epigenetic differences, which could lead to hypotheses for biological pathways in specific patient subgroups, and corresponding targeted interventions critical for precision medicine. Using bipartite network analysis of fetal DNA methylation data we demonstrate a novel method for classification of PTB. METHODS: The data consisted of DNA methylation across the genome (HumanMethylation450 BeadChip) in cord blood from 50 African-American subjects consisting of 22 cases of early spontaneous PTB (24-34 weeks of gestation) and 28 controls (>39 weeks of gestation). These data were analyzed using a combination of (1) a supervised method to select the top 10 significant methylation sites, (2) unsupervised "subject-variable" bipartite networks to visualize and quantitatively analyze how those 10 methylation sites co-occurred across all the subjects, and across only the cases with the goal of analyzing subgroups and their underlying pathways, and (3) a simple linear regression to test whether there was an association between the total methylation in the cases, and gestational age. RESULTS: The bipartite network analysis of all subjects and significant methylation sites revealed statistically significant clustering consisting of an inverse symmetrical relationship in the methylation profiles between a case-enriched subgroup and a control-enriched subgroup: the former was predominantly hypermethylated across seven methylation sites, and hypomethylated across three methylation sites, whereas the latter was predominantly hypomethylated across the above seven methylation sites and hypermethylated across the three methylation sites. Furthermore, the analysis of only cases revealed one subgroup that was predominantly hypomethylated across seven methylation sites, and another subgroup that was hypomethylated across all methylation sites suggesting the presence of heterogeneity in PTB pathophysiology. Finally, the analysis found a strong inverse linear relationship between total methylation and gestational age suggesting that methylation differences could be used as predictive markers for gestational length. CONCLUSIONS: The results demonstrate that unsupervised bipartite networks helped to identify a complex but comprehensible data-driven hypotheses related to patient subgroups and inferences about their underlying pathways, and therefore were an effective complement to supervised approaches currently used.

Exposure to Violence Accelerates Epigenetic Aging in Children.

Epigenetic processes, including DNA methylation, change reliably with age across the lifespan, such that DNA methylation can be used as an "epigenetic clock". This epigenetic clock can be used to predict age and age acceleration, which occurs when methylation-based prediction of age exceeds chronological age and has been associated with increased mortality. In the current study we examined epigenetic age acceleration using saliva samples collected from children between ages 6-13 (N = 101). Children's exposure to neighborhood violence and heart rate during a stressful task were assessed. Age acceleration was associated with children's direct experience of violence (p = 0.004) and with decreased heart rate (p = 0.002). Children who were predicted to be older than their chronological age had twice as much violence exposure as other children and their heart rate was similar to that of adults. The results remained significant after controlling for demographic variables, such as sex, income and education. This is the first study to show the effects of direct violence exposure on epigenetic aging in children using salivary DNA. Although longitudinal studies are needed to determine whether accelerated epigenetic aging leads to adverse health outcomes later in life, these data point to DNA methylation during childhood as a putative biological mechanism.

Resilience and biomarkers of health risk in Black smokers and nonsmokers.

OBJECTIVES: Blacks are disproportionately affected by tobacco-related illnesses as well as traumatic events associated with psychiatric conditions and smoking. We examined the potential protective nature of resilience within this context, hypothesizing resilience differentially moderates the associations of traumatic experiences to depressive symptoms and to biomarkers of health risk among Black ever versus never smokers. METHOD: Measures of resilience, traumatic experiences, depressive symptoms, and biomarkers (interleukin-6 [IL-6], C-reactive protein [CRP], allostatic load) were obtained among 852 Blacks recruited from Grady Memorial Hospital in Atlanta. RESULTS: Ever smokers experienced more trauma (p < .001) and depressive symptoms (p = .01). Structural equation modeling indicated that, in ever smokers, childhood trauma was positively associated with depressive symptoms (p < .001); resilience was negatively associated with depressive symptoms (p = .01). Depressive symptoms were positively associated with IL-6 (p = .03), which was positively associated with allostatic load (p = .01). Adulthood trauma was associated with higher CRP levels (p = .03). In never smokers, childhood (p < .001) and adulthood trauma (p = .01) were associated with more depressive symptoms. Adulthood trauma was also associated with higher CRP levels (p < .001), which was positively associated with allostatic load (p < .001). Never smokers with higher resilience had a negative association between childhood trauma and depressive symptoms whereas those with lower resilience had a positive association between childhood trauma and depressive symptoms. Resilience was negatively associated with CRP levels (p < .001). CONCLUSIONS: Interventions targeting resilience may prevent smoking and adverse health outcomes. (PsycINFO Database Record