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Objective:There is growing evidence that exposure to trauma prior to conception can affect offspring. The authors have reported that adult offspring of Holocaust survivors showed lower methylation ...

Background Epigenetic mechanisms may play a major role in the biological embedding of early-life stress (ELS). One proposed mechanism is that glucocorticoid (GC) release following ELS exposure induces long-lasting alterations in DNA methylation (DNAm) of important regulatory genes of the stress response. Here, we investigate the dynamics of GC-dependent methylation changes in key regulatory regions of the FKBP5 locus in which ELS-associated DNAm changes have been reported. Results We repeatedly measured DNAm in human peripheral blood samples from 2 independent cohorts exposed to the GC agonist dexamethasone (DEX) using a targeted bisulfite sequencing approach, complemented by data from Illumina 450K arrays. We detected differentially methylated CpGs in enhancers co-localizing with GC receptor binding sites after acute DEX treatment (1 h, 3 h, 6 h), which returned to baseline levels within 23 h. These changes withstood correction for immune cell count differences. While we observed main effects of sex, age, body mass index, smoking, and depression symptoms on FKBP5 methylation levels, only the functional FKBP5 SNP (rs1360780) moderated the dynamic changes following DEX. This genotype effect was observed in both cohorts and included sites previously shown to be associated with ELS. Conclusion Our study highlights that DNAm levels within regulatory regions of the FKBP5 locus show dynamic changes following a GC challenge and suggest that factors influencing the dynamics of this regulation may contribute to the previously reported alterations in DNAm associated with current and past ELS exposure. Electronic supplementary material The online version of this article (10.1186/s13148-019-0682-5) contains supplementary material, which is available to authorized users.

Lasting effects of adversity, such as exposure to childhood adversity (CA) on disease risk, may be embedded via epigenetic mechanisms but findings from human studies investigating the main effects of such exposure on epigenetic measures, including DNA methylation (DNAm), are inconsistent. Studies in perinatal tissues indicate that variability of DNAm at birth is best explained by the joint effects of genotype and prenatal environment. Here, we extend these analyses to postnatal stressors. We investigated the contribution of CA, cis genotype (G), and their additive (G + CA) and interactive (G × CA) effects to DNAm variability in blood or saliva from five independent cohorts with a total sample size of 1074 ranging in age from childhood to late adulthood. Of these, 541 were exposed to CA, which was assessed retrospectively using self-reports or verified through social services and registries. For the majority of sites (over 50%) in the adult cohorts, variability in DNAm was best explained by G + CA or G × CA but almost never by CA alone. Across ages and tissues, 1672 DNAm sites showed consistency of the best model in all five cohorts, with G × CA interactions explaining most variance. The consistent G × CA sites mapped to genes enriched in brain-specific transcripts and Gene Ontology terms related to development and synaptic function. Interaction of CA with genotypes showed the strongest contribution to DNAm variability, with stable effects across cohorts in functionally relevant genes. This underscores the importance of including genotype in studies investigating the impact of environmental factors on epigenetic marks.

Background: Maternal depression and anxiety during pregnancy may enhance fetal exposure to glucocorticoids (GCs) and harm neurodevelopment. We tested whether a novel cross-tissue polyepigenetic biomarker indicative of in utero exposure to GC is associated with mental and behavioral disorders and their severity in children, possibly mediating the associations between maternal prenatal depressive and anxiety symptoms and these child outcomes. Methods: Children (n = 814) from the Prediction and Prevention of Preeclampsia and Intrauterine Growth Restriction (PREDO) study were followed-up from birth to age 7.1–10.7 years. A weighted polyepigenetic GC exposure score was calculated based on the methylation profile of 24 CpGs from umbilical cord blood. Child diagnosis of mental and behavioral disorder (n = 99) and its severity, defined as the number of days the child had received treatment (all 99 had received outpatient treatment and 8 had been additionally in inpatient treatment) for mental or behavioral disorder as the primary diagnosis, came from the Care Register for Health Care. Mothers (n = 408) reported on child total behavior problems at child's age of 2.3–5.8 years and their own depressive and anxiety symptoms during pregnancy (n = 583). Results: The fetal polyepigenetic GC exposure score at birth was not associated with child hazard of mental and behavioral disorder (HR = 0.82, 95% CI 0.54; 1.24, p = 0.35) or total behavior problems (unstandardized beta = −0.10, 95% CI -0.31; 0.10, p = 0.33). However, for one standard deviation decrease in the polyepigenetic score, the child had spent 2.94 (95%CI 1.59; 5.45, p < 0.001) more days in inpatient or outpatient treatment with any mental and behavioral disorder as the primary diagnosis. Criteria for mediation tests were not met. Conclusions: These findings suggest that fetal polyepigenetic GC exposure score at birth was not associated with any mental or behavioral disorder diagnosis or mother-rated total behavior problems, but it may contribute to identifying children at birth who are at risk for more severe mental or behavioral disorders. publishedVersion

Epigenetic processes that regulate gene expression, such as DNA methylation (DNAm), have been linked to individual differences in physical aggression. Yet, it is currently unclear whether: (a) DNAm patterns in humans associate with physical aggression independently of other co-occurring psychiatric and behavioral symptoms; (b) whether these patterns are observable across multiple tissues; and (c) whether they may function as a causal versus noncausal biomarker of physical aggression. Here, we used a multisample, cross-tissue design to address these questions. First, we examined genome-wide DNAm patterns (buccal swabs; Illumina 450k) associated with engagement in physical fights in a sample of high-risk youth (n = 119; age = 16-24 years; 53% female). We identified one differentially methylated region in DRD4, which survived genome-wide correction, associated with physical aggression above and beyond co-occurring symptomatology (e.g., ADHD, substance use), and showed strong cross-tissue concordance with both blood and brain. Second, we found that DNAm sites within this region were also differentially methylated in an independent sample of young adults, between individuals with a history of chronic-high versus low physical aggression (peripheral T cells; ages 26-28). Finally, we ran a Mendelian randomization analysis using GWAS data from the EAGLE consortium to test for a causal association of DRD4 methylation with physical aggression. Only one genetic instrument was eligible for the analysis, and results provided no evidence for a causal association. Overall, our findings lend support for peripheral DRD4 methylation as a potential biomarker of physically aggressive behavior, with no evidence yet of a causal relationship.

Individuals who have experienced high levels of childhood stress are at increased risk for a wide range of behavioral problems that persist into adulthood, yet the neurobiological and molecular mechanisms underlying these associations remain poorly understood. Many of the difficulties observed in stress-exposed children involve problems with learning and inhibitory control. This experiment was designed to test individuals' ability to learn to inhibit responding during a laboratory task. To do so, we measured stress exposure among a community sample of school-aged children, and then followed these children for a decade. Those from the highest and lowest quintiles of childhood stress exposure were invited to return to our laboratory as young adults. At that time, we reassessed their life stress exposure, acquired functional magnetic resonance imaging data during an inhibitory control task, and assayed these individuals' levels of methylation in the FK506 binding protein 5 (FKBP5) gene. We found that individuals who experienced high levels of stress in childhood showed less differentiation in the dorsolateral prefrontal cortex between error and correct trials during inhibition. This effect was associated only with childhood stress exposure and not by current levels of stress in adulthood. In addition, FKBP5 methylation mediated the association between early life stress and inhibition-related prefrontal activity. These findings are discussed in terms of using multiple levels of analyses to understand the ways in which adversity in early development may affect adult behavioral adaptation.

Prenatal stress exposure is associated with risk for psychiatric disorders later in life. This may be mediated in part via enhanced exposure to glucocorticoids (GCs), which are known to impact neurogenesis. We aimed to identify molecular mediators of these effects, focusing on long-lasting epigenetic changes. In a human hippocampal progenitor cell (HPC) line, we assessed the short- and long-term effects of GC exposure during neurogenesis on messenger RNA (mRNA) expression and DNA methylation (DNAm) profiles. GC exposure induced changes in DNAm at 27,812 CpG dinucleotides and in the expression of 3,857 transcripts (false discovery rate [FDR] = 1.15). HPC expression and GC-affected DNAm profiles were enriched for changes observed during human fetal brain development. Differentially methylated sites (DMSs) with GC exposure clustered into 4 trajectories over HPC differentiation, with transient as well as long-lasting DNAm changes. Lasting DMSs mapped to distinct functional pathways and were selectively enriched for poised and bivalent enhancer marks. Lasting DMSs had little correlation with lasting expression changes but were associated with a significantly enhanced transcriptional response to a second acute GC challenge. A significant subset of lasting DMSs was also responsive to an acute GC challenge in peripheral blood. These tissue-overlapping DMSs were used to compute a polyepigenetic score that predicted exposure to conditions associated with altered prenatal GCs in newborn's cord blood DNA. Overall, our data suggest that early exposure to GCs can change the set point of future transcriptional responses to stress by inducing lasting DNAm changes. Such altered set points may relate to differential vulnerability to stress exposure later in life.

Dynamic adaptation to stressful life events requires the co-ordinated action of the central stress response, which is mediated by the hypothalamic-pituitary-adrenal (HPA) axis, to restore and maintain homeostasis. Excessive exposure to stress or traumatic life events, such as childhood maltreatment, has been linked to HPA axis dysfunction increasing the risk of developing stress-related psychopathologies such as major depressive disorder and post-traumatic-stress-disorder. Mounting evidence supports the notion that stressors throughout pre- and postnatal development as well as adulthood can induce neuroepigenetic regulation of gene expression within key nodes of the brain, which may in part mediate such HPA axis dysfunction. Neuroepigenetic mechanisms, particularly DNA methylation and small non-coding RNAs, are therefore considered to be molecular mechanisms by which stressful life events may perpetuate aberrant behavioral phenotypes associated with psychiatric disorders throughout one's life and even across generations. In this chapter we outline the progress made toward understanding the effects of stress-induced neuroepigenetic changes upon HPA axis function and highlight the need for novel research strategies to deepen our understanding of the establishment, maintenance and reversibility of neuroepigenetic regulation following stress to enable realization of potential novel therapeutic and preventative strategies for stress-related psychiatric disorders.

The development of mental disorders constitutes a complex phenomenon driven by unique social, psychological and biological factors such as genetics and epigenetics, throughout an individual's life course. Both environmental and genetic factors have an impact on mental health phenotypes and act simultaneously to induce changes in brain and behavior. Here, we describe and critically evaluate the current literature on gene-environment interactions and epigenetics on mental health by highlighting recent human and animal studies. We furthermore review some of the main ethical and social implications concerning gene-environment interactions and epigenetics and provide explanations and suggestions on how to move from statistical and epigenetic associations to biological and psychological explanations within a multi-disciplinary and integrative approach of understanding mental health.

Maternal behavior (MB) is observable across mammals and represents an important feature of environmental variation during early postnatal development. Oxytocin (OT) plays a crucial role in MB. Even prior to childbirth, pregnancy induces epigenetic and other downstream changes in the maternal OT-system, likely mediated by the actions of steroid hormones. However, little is known about the nature and consequences of epigenetic modifications in the maternal OT-encoding gene (OXT) during pregnancy. Our study aims to investigate temporal dynamics of OXT promoter DNA methylation (DNAm) throughout pregnancy in predicting MB in humans. In 107 mother-child dyads, maternal OXT DNAm was serially analyzed in whole blood in early, mid and late pregnancy. MB was coded based on standardized mother-child interactions at six months postpartum. After controlling for cellular heterogeneity, race/ethnicity, age, and socioeconomic status, OXT-promoter DNAm exhibited a dynamic profile during pregnancy (b = 0.026, t=−3.37, p < .001), with decreases in DNAm from early to mid-pregnancy and no further change until late pregnancy. Moreover, dynamic DNAm trajectories of the OXT-promoter region predicted MB (intrusiveness) at six months postpartum (b = 0.006, t = 2.0, p < 0.05), with 6% higher OXT DNAm in late pregnancy in intrusive compared to non-intrusive mothers. We here demonstrate that OXT promoter DNAm changes significantly throughout gestation in peripheral blood and that these changes are associated with variability in MB, providing a novel potential biomarker predicting postnatal MB.

Background The ability to accurately and efficiently measure DNA methylation is critical to advance the understanding of this epigenetic mechanism and its contribution to common diseases. Here, we present a highly accurate method to measure methylation using bisulfite sequencing (termed HAM-TBS). This novel method is able to assess DNA methylation in multiple samples with high accuracy in a cost-effective manner. We developed this assay for the FKBP5 locus, an important gene in the regulation of the stress system and previously linked to stress-related disorders, but the method is applicable to any locus of interest. Results HAM-TBS enables multiplexed analyses of up to 96 samples and regions spanning 10 kb using the Illumina MiSeq. It incorporates a triplicate bisulfite conversion step, pooled target enrichment via PCR, PCR-free library preparation and a minimum coverage of 1000×. TBS was able to resolve DNA methylation levels with a mean accuracy of 0.72%. Using this method, we designed and validated a targeted panel to specifically assess regulatory regions within the FKBP5 locus that are not covered in commercially available DNA methylation arrays. Conclusions HAM-TBS represents a highly accurate, medium-throughput sequencing approach for robust detection of DNA methylation changes in specific target regions. Electronic supplementary material The online version of this article (10.1186/s13072-018-0209-x) contains supplementary material, which is available to authorized users.

Dynamic adaptation to stressful life events requires the co-ordinated action of the central stress response, which is mediated by the hypothalamic-pituitary-adrenal (HPA) axis, to restore and maintain homeostasis. Excessive exposure to stress or traumatic life events, such as childhood maltreatment, has been linked to HPA axis dysfunction increasing the risk of developing stress-related psychopathologies such as major depressive disorder and post-traumatic-stress-disorder. Mounting evidence supports the notion that stressors throughout pre- and postnatal development as well as adulthood can induce neuroepigenetic regulation of gene expression within key nodes of the brain, which may in part mediate such HPA axis dysfunction. Neuroepigenetic mechanisms, particularly DNA methylation and small non-coding RNAs, are therefore considered to be molecular mechanisms by which stressful life events may perpetuate aberrant behavioral phenotypes associated with psychiatric disorders throughout one's life and even across generations. In this chapter we outline the progress made toward understanding the effects of stress-induced neuroepigenetic changes upon HPA axis function and highlight the need for novel research strategies to deepen our understanding of the establishment, maintenance and reversibility of neuroepigenetic regulation following stress to enable realization of potential novel therapeutic and preventative strategies for stress-related psychiatric disorders.

The development of mental disorders constitutes a complex phenomenon driven by unique social, psychological and biological factors such as genetics and epigenetics, throughout an individual's life course. Both environmental and genetic factors have an impact on mental health phenotypes and act simultaneously to induce changes in brain and behavior. Here, we describe and critically evaluate the current literature on gene-environment interactions and epigenetics on mental health by highlighting recent human and animal studies. We furthermore review some of the main ethical and social implications concerning gene-environment interactions and epigenetics and provide explanations and suggestions on how to move from statistical and epigenetic associations to biological and psychological explanations within a multi-disciplinary and integrative approach of understanding mental health.

Over the past decade, epigenetic analyses have made important contributions to our understanding of healthy development and a wide variety of adverse conditions such as cancer and psychopathology. There is increasing evidence that DNA methylation is a mechanism by which environmental factors influence gene transcription and, ultimately, phenotype. However, differentiating the effects of the environment from those of genetics on DNA methylation profiles remains a significant challenge. Monozygotic (MZ) twin study designs are unique in their ability to control for genetic differences because each pair of MZ twins shares essentially the same genetic sequence with the exception of a small number of de novo mutations and copy number variations. Thus, differences within twin pairs in gene expression and phenotype, including behavior, can be attributed in the majority of cases to environmental effects rather than genetic influence. In this article, we review the literature showing how MZ twin designs can be used to study basic epigenetic principles, contributing to understanding the role of early in utero and postnatal environmental factors on the development of psychopathology. We also highlight the importance of initiating longitudinal and experimental studies with MZ twins during pregnancy. This approach is especially important to identify: (1) critical time periods during which the early environment can impact brain and mental health development, and (2) the specific mechanisms through which early environmental effects may be mediated. These studies may inform the optimum timing and design for early preventive interventions aimed at reducing risk for psychopathology.

Posttraumatic stress disorder (PTSD) is a stress-related psychiatric disorder that is thought to emerge from complex interactions among traumatic events and multiple genetic factors. Epigenetic regulation lies at the heart of these interactions and mediates the lasting effects of the environment on gene regulation. An increasing body of evidence in human subjects with PTSD supports a role for epigenetic regulation of distinct genes and pathways in the pathogenesis of PTSD. The role of epigenetic regulation is further supported by studies examining fear conditioning in rodent models. Although this line of research offers an exciting outlook for future epigenetic research in PTSD, important limitations include the tissue specificity of epigenetic modifications, the phenomenologic definition of the disorder, and the challenge of translating molecular evidence across species. These limitations call for studies that combine data from postmortem human brain tissue and animal models, assess longitudinal epigenetic changes in living subjects, and examine dimensional phenotypes in addition to diagnoses. Moreover, examining the environmental, genetic, and epigenetic factors that promote resilience to trauma may lead to important advances in the field.

Exposure to early life stress (ELS), such as childhood abuse and neglect is a well established major risk factor for developing psychiatric and behavioral disorders later in life. Both prenatal and postnatal stressors have been shown to have a long-lasting impact on adult pathological states where the type and timing of the stressor are important factors to consider. There is a growing body of evidence suggesting that epigenetic mechanisms play a major role in the biological embedding of ELS. A number of studies now indicate that the epigenome is responsive to external environmental exposures, including the social environment, both during intra-uterine development and after birth. In this review, we summarize the evidence of long-lasting effects of ELS on mental health and behavior and highlight common and distinct epigenetic effects of stress exposure at different stages during development. These stages include postnatal stress, prenatal stress, i.e. in utero and stress occurring pre-conception, i.e. effects of stress exposure transmitted to the next generation. We also delineate the evidence for the possible molecular mechanisms involved in epigenetic programming by ELS and how these maybe distinct, according to the timing of the stress exposure.

BackgroundThe ability to accurately and efficiently measure DNA methylation is vital to advance the understanding of this mechanism and its contribution to common diseases. Here, we present a highly accurate method to measure methylation using bisulfite sequencing (termed HAM-TBS). This novel method is able to assess DNA methylation in multiple samples with high accuracy in a cost-effective manner. We developed this assay for the FKBP5 locus, an important gene in the regulation of the stress system and previously linked to stress-related disorders, but the method is applicable to any locus of interest.ResultsHAM-TBS enables multiplexing of up to 96 samples spanning a region of ~10 kb using the llumina MiSeq. It incorporates a triplicate bisulfite conversion step, pooled target enrichment via PCR, PCR-free library preparation and a minimum coverage of 1,000x. Furthermore, we designed and validated a targeted panel to specifically assess regulatory regions within the FKBP5 locus including the transcription start site, topologically associated domain boundaries, intergenic and proximal enhancers as well as glucocorticoid receptor and CTCF binding sites that are not covered in commercially available DNA methylation arrays.ConclusionsHAM-TBS represents a highly accurate, medium-throughput sequencing approach for robust detection of DNA methylation changes in specific target regions.

The first part of this chapter summarizes present knowledge on the development of human chronic physical aggression from early childhood onwards. The second part of the chapter summarizes the state of knowledge on the genetic, epigenetic, and environmental mechanisms that may be involved in the development of chronic physical aggression from conception onwards. The third part of the chapter describes examples of research on the prevention of chronic physical aggression that could advance both knowledge on the developmental GxE mechanisms involved in chronic physical aggression and knowledge on the effectiveness of preventive interventions targeting at-risk families from early pregnancy onwards.

Background Epigenetic modifications, in particular DNA methylation, play an important role in many biological processes in human health and disease. Understanding the cause of inter-individual differences in DNA methylation levels may help to identify novel molecular mechanisms contributing to a number of human diseases, including psychiatric diseases. Methods The aim of this study was to determine whether environmental risk factors for major depression (child abuse, socioeconomic status), genotype, or their interaction (GxE) best explain variablity in DNA methylation. We analyzed genome-wide DNA methylation (Illumina 450k arrays) from whole blood and genotype data of four independent cohorts (total N=1,.253). The cohorts consisted of adult individuals with diverse ethnic backgrounds, ages, and psychiatric disease status. Results We first identified the 15% most variably methylated (vm) CpGs in each cohort and observed that these vmCpGs had a higher correlation with brain methylation levels than expected by chance. Additionally, they were enriched in regulatory regions (including enhancer regions and specific, transcription factor binding sites). 40% (n=25.042) of the vmCpgGs overlapped between all 4 cohorts. We next analyzed, which factors explained most of the variance in methylation of these CpGs. While genotype alone explained most of the variance of about 20% of the vmCpGs, the majority of the vmCpGs were best explained by GxE (74%). E alone was almost never the model explaining most variance. We then investigated, how consistent the GxE effects on DNA methylation were across the 4 cohorts. We found that 30% of the vmCpGs were best explained by GxE in all four cohorts and 74% in at least three cohorts. The CpGs with GxE effects in all 4 cohorts also showed even stronger enrichment for enhancer regions than vmCpGs overall. Discussion Our study highlights the importance of considering both genetic and environmental data in interpreting epigenetic variation and suggests that integrating genotypes with epigenetic information could contribute to identifying functional genetic variants that moderate the impact of risk environments on the development of psychiatric disorders.

There is now convincing evidence from prospective and retrospective epidemiological studies that prenatal and early post-natal stressors have long term impacts on life span health and well-being. Unraveling the mechanisms by which early environmental stressors have an impact on DNA methylation and neurobehavioral development should provide the foundation for creating effective early protective interventions. We review the recent convergence of four research domains to explain the mechanisms leading to chronic physical aggression (behavior development, epigenetics, serotonin neurotransmission and immunology) and we discuss the next generation of studies that are needed to identify effective pre and post natal preventive interventions.

Early life is one of the most important and sensitive periods during the development of an individual. During this stage, the body and especially the brain are known to be greatly responsive to environmental cues, such as the early social environment. As a consequence, early life adverse social experiences in humans are associated with a wide range of health problems in adulthood. The broad range of phenotypes associated with early life stress (ELS) suggests a system-wide response of the organism, which is yet to be determined. In the last decade, increasing evidence suggests that epigenetic mechanisms underlie the effects of ELS on adult human health. However, there are critical challenges in delineating the direct effects of ELS on epigenetic profiles and phenotypes in human studies. It is impossible to randomize ELS and rare are the studies where complete information about past environmental insults is available, which would allow us to conclude on causality. Nonhuman primates offer several advantages in addressing these challenges. This chapter focuses on parental deprivation models in rhesus macaques which have been shown to produce an array of behavioral, physiological, and neurobiological deficits that parallel those identified in humans subjected to ELS. It describes the evidence for epigenetic alterations induced by differential rearing in this model and points out the differences between tissue-specific versus multi-tissue changes and outlines possible mechanisms for these to occur. In addition, it highlights the need for multi-omics longitudinal studies to better understand the epigenetic trajectories induced by ELS exposure and their impact on adult health.

Longitudinal epidemiological studies with birth cohorts have shown that physical aggression in humans does not appear suddenly in adolescence as commonly thought. In fact, physically aggressive behaviour is observed as early as 12 months after birth, its frequency peaks around 2–4 years of age and decreases in frequency until early adulthood. However, a minority of children (3–7%) maintain a high frequency of physical aggression from childhood to adolescence and develop serious social adjustment problems during adulthood. Genetic factors and early social experiences, as well as their interaction, have been shown to play an important role in the development of chronic aggressive behaviour. However, the biological mechanisms underlying these associations are just beginning to be uncovered. Recent evidence suggests that epigenetic mechanisms are responsive to adverse environments and could be involved in the development of chronic aggression. Using both gene candidate and genomic approaches, recent studies have identified epigenetic marks, such as DNA methylation alterations in genes involved in the stress response and the serotonin and immune systems to be partly responsible for the long-lasting effects of early adversity. Further longitudinal studies with biological, environmental and behavioural assessments from birth onwards are needed to elucidate the sequence of events that leads to these long-lasting epigenetic marks associated with early adversity and aggression.

A large body of evidence describes the long term impact of stress on a number of biological systems and with it associated adverse health outcomes. This article will discuss the epigenetic mechanisms of the embedding of these long term changes, the differences in these mechanisms depending on the type and timing of stress exposure, including transgenerational effects as well as differences in the mechanisms for tissue specific versus more global epigenetic changes. A mechanistic understanding of the long term epigenetic consequences of stress may allow novel, targeted intervention and prevention strategies for psychiatric and other stress-associated disorders.

5-Hydroxymethylcytosine (5hmC) is abundant in the brain, suggesting an important role in epigenetic control of neuronal functions. In this paper, we show that 5hmC and 5-methylcytosine (5mC) levels are coordinately distributed in gene promoters of the rhesus macaque prefrontal cortex. Although promoter hydroxymethylation and methylation are overall negatively correlated with expression, a subset of highly expressed genes involved in specific cerebral functions is associated with high levels of 5mC and 5hmC. These relationships were also observed in the mouse cortex. Furthermore, we found that early-life maternal deprivation is associated, in the adult monkey cortex, with DNA hydroxymethylation changes of promoters of genes related to neurological functions and psychological disorders. These results reveal that early social adversity triggers variations in brain DNA hydroxymethylation that could be detected in adulthood.

BACKGROUND: Chronic physical aggression (CPA) is characterized by frequent use of physical aggression from early childhood to adolescence. Observed in approximately 5% of males, CPA is associated with early childhood adverse environments and long-term negative consequences. Alterations in DNA methylation, a covalent modification of DNA that regulates genome function, have been associated with early childhood adversity. AIMS: To test the hypothesis that a trajectory of chronic physical aggression during childhood is associated with a distinct DNA methylation profile during adulthood. METHODS: We analyzed genome-wide promoter DNA methylation profiles of T cells from two groups of adult males assessed annually for frequency of physical aggression between 6 and 15 years of age: a group with CPA and a control group. Methylation profiles covering the promoter regions of 20 000 genes and 400 microRNAs were generated using MeDIP followed by hybridization to microarrays. RESULTS: In total, 448 distinct gene promoters were differentially methylated in CPA. Functionally, many of these genes have previously been shown to play a role in aggression and were enriched in biological pathways affected by behavior. Their locations in the genome tended to form clusters spanning millions of bases in the genome. CONCLUSIONS: This study provides evidence of clustered and genome-wide variation in promoter DNA methylation in young adults that associates with a history of chronic physical aggression from 6 to 15 years of age. However, longitudinal studies of methylation during early childhood will be necessary to determine if and how this methylation variation in T cells DNA plays a role in early development of chronic physical aggression. Language: en

Reported exposure to traumatic event is relatively common within the general population (40-90%), but only a fraction of individuals will develop posttraumatic stress disorder (PTSD). Indeed, the lifetime prevalence of PTSD is estimated to range between 7% and 12%. The factors influencing risk or resilience to PTSD after exposure to traumatic events are likely both environmental, such as type, timing, and extent of trauma, and genetic. Recently, epigenetic mechanisms have been implicated in mediating altered risk for PTSD as they can reflect both genetic and environmental influences. In this chapter, we describe the accumulating evidences for epigenetic factors in PTSD highlighting the importance of sensitive periods as well as methodological aspects such as tissue availabilities for such studies. We describe studies using a candidate gene approach focusing mainly on key players in the stress hormone regulation that show epigenetic alterations both in humans and in animal models for PTSD. We also summarize the results of epigenome-wide studies reporting associations with PTSD. For the above, we focus on one epigenetic mechanism, DNA methylation, as it is so far the best studied for this disorder. Finally, we describe how epigenetic mechanisms could be responsible for the long-lasting effects of gene-environment interactions observed in PTSD.

Background High frequency of physical aggression is the central feature of severe conduct disorder and is associated with a wide range of social, mental and physical health problems. We have previously tested the hypothesis that differential DNA methylation signatures in peripheral T cells are associated with a chronic aggression trajectory in males. Despite the fact that sex differences appear to play a pivotal role in determining the development, magnitude and frequency of aggression, most of previous studies focused on males, so little is known about female chronic physical aggression. We therefore tested here whether or not there is a signature of physical aggression in female DNA methylation and, if there is, how it relates to the signature observed in males. Methodology/Principal Findings Methylation profiles were created using the method of methylated DNA immunoprecipitation (MeDIP) followed by microarray hybridization and statistical and bioinformatic analyses on T cell DNA obtained from adult women who were found to be on a chronic physical aggression trajectory (CPA) between 6 and 12 years of age compared to women who followed a normal physical aggression trajectory. We confirmed the existence of a well-defined, genome-wide signature of DNA methylation associated with chronic physical aggression in the peripheral T cells of adult females that includes many of the genes similarly associated with physical aggression in the same cell types of adult males. Conclusions This study in a small number of women presents preliminary evidence for a genome-wide variation in promoter DNA methylation that associates with CPA in women that warrant larger studies for further verification. A significant proportion of these associations were previously observed in men with CPA supporting the hypothesis that the epigenetic signature of early life aggression in females is composed of a component specific to females and another common to both males and females.

Background Animal and human studies suggest that inflammation is associated with behavioral disorders including aggression. We have recently shown that physical aggression of boys during childhood is strongly associated with reduced plasma levels of cytokines IL-1α, IL-4, IL-6, IL-8 and IL-10, later in early adulthood. This study tests the hypothesis that there is an association between differential DNA methylation regions in cytokine genes in T cells and monocytes DNA in adult subjects and a trajectory of physical aggression from childhood to adolescence. Methodology/Principal Findings We compared the methylation profiles of the entire genomic loci encompassing the IL-1α, IL-6, IL-4, IL-10 and IL-8 and three of their regulatory transcription factors (TF) NFkB1, NFAT5 and STAT6 genes in adult males on a chronic physical aggression trajectory (CPA) and males with the same background who followed a normal physical aggression trajectory (control group) from childhood to adolescence. We used the method of methylated DNA immunoprecipitation with comprehensive cytokine gene loci and TF loci microarray hybridization, statistical analysis and false discovery rate correction. We found differentially methylated regions to associate with CPA in both the cytokine loci as well as in their transcription factors loci analyzed. Some of these differentially methylated regions were located in known regulatory regions whereas others, to our knowledge, were previously unknown as regulatory areas. However, using the ENCODE database, we were able to identify key regulatory elements in many of these regions that indicate that they might be involved in the regulation of cytokine expression. Conclusions We provide here the first evidence for an association between differential DNA methylation in cytokines and their regulators in T cells and monocytes and male physical aggression.

BACKGROUND: An increasing number of animal and human studies are indicating that inflammation is associated with behavioral disorders including aggression. This study investigates the association between chronic physical aggression during childhood and plasma cytokine levels in early adulthood. METHODOLOGY/PRINCIPAL FINDINGS: Two longitudinal studies were used to select males on a chronic physical aggression trajectory from childhood to adolescence (n = 7) and a control group from the same background (n = 25). Physical aggression was assessed yearly by teachers from childhood to adolescence and plasma levels of 10 inflammatory cytokines were assessed at age 26 and 28 years. Compared to the control group, males on a chronic physical aggression trajectory from childhood to adolescence had consistently lower plasma levels of five cytokines: lower pro-inflammatory interleukins IL-1α (T(28.7) = 3.48, P = 0.002) and IL-6 (T(26.9) = 3.76, P = 0.001), lower anti-inflammatory interleukin IL-4 (T(27.1) = 4.91, P = 0.00004) and IL-10 (T(29.8) = 2.84, P = 0.008) and lower chemokine IL-8 (T(26) = 3.69, P = 0.001). The plasma levels of four cytokines accurately predicted aggressive and control group membership for all subjects. CONCLUSIONS/SIGNIFICANCE: Physical aggression of boys during childhood is a strong predictor of reduced plasma levels of cytokines in early adulthood. The causal and physiological relations underlying this association should be further investigated since animal data suggest that some cytokines such as IL-6 and IL-1β play a causal role in aggression. Language: en

5-hydroxymethylcytosine (5hmC) is a recently re-discovered transient intermediate in the active demethylation pathway that also appears to play an independent role in modulating gene function. Epigenetic marks, particularly 5-methylcytosine, have been widely studied in relation to stress-related disorders given the long-lasting effect that stress has on these marks. 5hmC is a good candidate for involvement in the etiology of these disorders given its elevated concentration in mammalian neurons, its dynamic regulation during development of the central nervous system, and its high variability among individuals. Although we are unaware of any studies published to date examining 5 hmC profiles in human subjects who have developed a psychiatric disorder after a life stressor, there is emerging evidence from the animal literature that 5hmC profiles are altered in the context of fear-conditioning paradigms and stress exposure, suggesting a possible role for 5hmC in the biological underpinnings of stress-related disorders. In this review, the authors examine the available approaches for profiling 5hmC and describe their advantages and disadvantages as well as discuss the studies published thus far investigating 5hmC in the context of fear-related learning and stress exposure in animals. The authors also highlight the global versus locus-specific regulation of 5hmC in these studies. Finally, the limitations of the current studies and their implications are discussed.

Early-life adversity is associated with a broad scope of life-long health and behavioral disorders. Particularly critical is the role of the mother. A possible mechanism is that these effects are mediated by “epigenetic” mechanisms. Studies in rodents suggest a causal relationship between early-life adversity and changes in DNA methylation in several “candidate genes” in the brain. This study examines whether randomized differential rearing (maternal vs surrogate–peer rearing) of rhesus macaques is associated with differential methylation in early adulthood. The data presented here show that differential rearing leads to differential DNA methylation in both prefrontal cortex and T cells. These differentially methylated promoters tend to cluster by both chromosomal region and gene function. The broad impact of maternal rearing on DNA methylation in both the brain and T cells supports the hypothesis that the response to early-life adversity is system-wide and genome-wide and persists to adulthood. Our data also point to the feasibility of studying the impact of the social environment in peripheral T-cell DNA methylation.

DNA methylation and chromatin structure are two modes of epigenetic control of genome function. Although it is now well established that chromatin silencing could lead to DNA methylation, the relation between chromatin activation and DNA demethylation is unclear. It was generally believed that expression of methylated genes could only be restored by demethylating agents, such as 5-aza-deoxycytidine (5-azaCdR), and that inhibition of histone deacetylation by Trichostatin A (TSA) only activates transcription of unmethylated genes. In this report, we show that increase of histone acetylation by TSA was associated with a significant decrease in global methylation. This global demethylation occurs even when DNA replication is blocked by hydroxyurea, supporting a replication-independent-mechanism of demethylation. TSA also induces histone acetylation, demethylation and expression of the methylated E-CADHERIN and RARbeta2 genes. However, the genome-wide demethylation induced by TSA does not affect all methylated tumor suppressor genes equally suggesting that induction of acetylation and demethylation by TSA shows some gene selectivity. Taken together, our data provide evidence for a reversible crosstalk between histone acetylation and DNA demethylation, which has significant implications on the use of HDAC inhibitors as therapeutic agents.