The DNA methylation landscape of the human oxytocin receptor gene (OXTR): data-driven clusters and their relation to gene expression and childhood adversity.

The oxytocin receptor gene (OXTR) is of interest when investigating the effects of early adversity on DNA methylation. However, there is heterogeneity regarding the selection of the most promising CpG sites to target for analyses. The goal of this study was to determine functionally relevant clusters of CpG sites within the OXTR CpG island in 113 mother-infant dyads, with 58 of the mothers reporting childhood maltreatment (CM). OXTR DNA methylation was analyzed in peripheral/umbilical blood mononuclear cells. Different complexity reduction approaches were used to reduce the 188 CpG sites into clusters of co-methylated sites. Furthermore, associations between OXTR DNA methylation (cluster- and site-specific level) and OXTR gene expression and CM were investigated in mothers. Results showed that, first, CpG sections differed strongly regarding their statistical utility for research of individual differences in DNA methylation. Second, cluster analyses and Partial Least Squares (PLS) suggested two clusters consisting of intron1/exon2 and the protein-coding region of exon3, respectively, as most strongly associated with outcome measures. Third, cross-validated PLS regression explained 7% of variance in CM, with low cross-validated variance explained for the prediction of gene expression. Fourth, substantial mother-child correspondence was observed in correlation patterns within the identified clusters, but only modest correspondence outside these clusters. This study makes an important contribution to the mapping of the DNA methylation landscape of the OXTR CpG island by highlighting clusters of CpG sites that show desirable statistical properties and predictive value. We provide a Companion Web Application to facilitate the choice of CpG sites.

Investigating mitochondrial bioenergetics in peripheral blood mononuclear cells of women with childhood maltreatment from post-parturition period to one-year follow-up.

BACKGROUND: Childhood maltreatment (CM) exerts various long-lasting psychological and biological changes in affected individuals, with inflammation being an interconnecting element. Besides chronic low-grade inflammation, CM might also affect the energy production of cells by altering the function and density of mitochondria, i.e. the body's main energy suppliers. Here, we compared mitochondrial respiration and density in intact peripheral blood mononuclear cells (PBMC), from women with and without CM between two time points, i.e. at the highly inflammatory phase within 1 week after parturition (t0) and again after 1 year (t2). METHODS: CM exposure was assessed with the Childhood Trauma Questionnaire. Whole blood was collected from n = 52 healthy women within the study 'My Childhood - Your Childhood' at both time points to isolate and cryopreserve PBMC. Thawed PBMC were used to measure mitochondrial respiration and density by high-resolution respirometry followed by spectrophotometric analyses of citrate-synthase activity. RESULTS: Over time, quantitative respiratory parameters increased, while qualitative flux control ratios decreased, independently of CM. Women with CM showed higher mitochondrial respiration and density at t0, but not at t2. We found significant CM group × time interaction effects for ATP-turnover-related respiration and mitochondrial density. CONCLUSIONS: This is the first study to longitudinally investigate mitochondrial bioenergetics in postpartum women with and without CM. Our results indicate that CM-related mitochondrial alterations reflect allostatic load, probably due to higher inflammatory states during parturition, which normalize later. However, later inflammatory states might moderate the vulnerability for a second-hit on the level of mitochondrial bioenergetics, at least in immune cells.

A history of childhood maltreatment is associated with altered DNA methylation levels of DNA methyltransferase 1 in maternal but not neonatal mononuclear immune cells.

Childhood maltreatment (CM) is associated with alterations in DNA methylation (DNAm) especially in stress response genes. Due to the higher risk of overall health complications of individuals with a parental history of CM, intergenerational transmission of CM-associated DNAm changes has been investigated but remains unclear. In this study, we investigated if different severities of CM have any influence on the DNAm of DNA methyltransferase 1 (DNMT1), an important enzyme of the DNAm machinery, in immune and buccal cells of mother-newborn dyads. DNAm was assessed by mass spectrometry using immune cell DNA from mothers (N = 117) and their newborns (N = 113), and buccal cell DNA of mother-newborn dyads (N = 68 each). Mothers with a history of CM had lower mean methylation of DNMT1 in immune cells compared to the mothers without a CM history. CM status only influenced maternal DNMT1 gene expression when at least moderate CM was reported. Buccal cell DNAm was not associated with CM status. Maternal history of CM was not linked to any alterations in DNMT1 mean DNAm in any of the cell types studied in newborns. We conclude that the CM-associated alterations in DNMT1 DNAm might point to allostatic load and can be physiologically relevant, especially in individuals with more severe CM experiences, resulting in an activated DNA methylation machinery that might influence stress response genes. Our lack of significant findings in buccal cells shows the tissue-specific effects of CM on DNAm. In our sample with low to moderate maternal CM history, there was no intergenerational transmission of DNMT1 DNAm in newborns.

Reactivity of the Oxytocinergic and Neuroendocrine System Following the Adult Attachment Projective Picture System in Men of Recent Fatherhood: Results from an Exploratory Pilot Study with a Cross-Sectional Design.

The attachment representation (AR) of individuals affects emotional and cognitive information processes and is considered an important modulating factor of neuroendocrine stress responses. The neuropeptide oxytocin is studied as one biomolecular component underpinning this modulation. A validated procedure used in attachment-related research is the Adult Attachment Projective Picture System (AAP). To date, only a limited number of studies investigated oxytocin and neuroendocrine reactivity in the context of an attachment-related stimulus similar to the APP. In this pilot study, N = 26 men of recent fatherhood were exposed to the AAP to classify AR and to investigate salivary changes in oxytocin, cortisol and dehydroepiandrosterone (DHEA) after AAP stimulation. We observed increased oxytocin levels in response to AAP exposure, and this increase was more pronounced in fathers with unresolved/disorganized AR. No significant changes in cortisol and DHEA concentrations were observed in response to AAP administration. Interestingly, differences in basal cortisol levels (before the AAP) also depended on AR classification. Here, the group of men with unresolved/disorganized AR showed higher levels of cortisol compared to fathers with organized AR. To conclude, the finding of increased salivary oxytocin levels in response to the AAP further indicates its validity as an instrument to stimulate the attachment system.

Characterization of the effects of age and childhood maltreatment on ELOVL2 DNA methylation.

DNA methylation of the elongation of very long chain fatty acids protein 2 (ELOVL2) was suggested as a biomarker of biological aging, while childhood maltreatment (CM) has been associated with accelerated biological aging. We investigated the association of age and CM experiences with ELOVL2 methylation in peripheral blood mononuclear cells (PBMC). Furthermore, we investigated ELOVL2 methylation in the umbilical cord blood mononuclear cells (UBMC) of newborns of mothers with and without CM. PBMC and UBMC were isolated from 113 mother-newborn dyads and genomic DNA was extracted. Mothers with and without CM experiences were recruited directly postpartum. Mass array spectrometry and pyrosequencing were used for methylation analyses of ELOVL2 intron 1, and exon 1 and 5' end, respectively. ELOVL2 5' end and intron 1 methylation increased with higher age but were not associated with CM experiences. On the contrary, overall ELOVL2 exon 1 methylation increased with higher CM, but these changes were minimal and did not increase with age. Maternal CM experiences and neonatal methylation of ELOVL2 intron 1 or exon 1 were not significantly correlated. Our study suggests region-specific effects of chronological age and experienced CM on ELOVL2 methylation and shows that the epigenetic biomarker for age within the ELOVL2 gene does not show accelerated biological aging years after CM exposure.

Associations between childhood maltreatment and DNA methylation of the oxytocin receptor gene in immune cells of mother-newborn dyads.

The neuropeptide oxytocin (OXT) and its receptor (OXTR) modulate interpersonal relationships, particularly mother-child interactions. DNA methylation (DNAm) changes of the OXTR gene were observed in individuals who experienced Childhood Maltreatment (CM). A modulatory role of single nucleotide polymorphisms (SNP) within OXTR in association with CM on the regulation of OXTR was also postulated. Whether these CM-induced epigenetic alterations are biologically inherited by the offspring remains unknown. We thus investigated possible intergenerational effects of maternal CM exposure on DNAm and OXTR gene expression, additionally accounting for the possible influence of three SNP: rs53576 and rs2254298 (OXTR gene), and rs2740210 (OXT gene). We used the Childhood Trauma Questionnaire to classify mothers into individuals with (CM+) or without CM (CM-). Maternal peripheral immune cells were isolated from venous blood (N = 117) and fetal immune cells from the umbilical cord (N = 113) after parturition. DNA methylation was assessed using MassARRAY. Taqman assays were performed for genotyping and gene expression analyses. Among mothers, CM was not associated with OXTR mean methylation or gene expression. However, four CpG sites showed different methylation levels in CM- compared to CM+. In mothers, the OXTR rs53576 and OXT rs2740210 allelic variations interacted with CM load on the OXTR mean methylation. Maternal and newborns' mean methylation of OXTR were positively associated within CM- dyads, but not in CM+ dyads. We show gene×environment interactions on the epigenetic regulation of the oxytocinergic signaling and show the intergenerational comparability of the OXTR DNAm might be altered in infants of CM+ mothers.

Childhood maltreatment is associated with changes in mitochondrial bioenergetics in maternal, but not in neonatal immune cells.

Childhood maltreatment (CM) comprises experiences of abuse and neglect during childhood. CM causes psychological as well as biological alterations in affected individuals. In humans, it is hardly explored whether these CM consequences can be transmitted directly on a biological level to the next generation. Here, we investigated the associations between maternal CM and mitochondrial bioenergetics (mitochondrial respiration and intracellular mitochondrial density) in immune cells of mothers and compared them with those of their newborns. In n = 102 healthy mother-newborn dyads, maternal peripheral blood mononuclear cells and neonatal umbilical cord blood mononuclear cells were collected and cryopreserved shortly after parturition to measure mitochondrial respiration and intracellular mitochondrial density with high-resolution respirometry and spectrophotometric analyses, respectively. Maternal CM was assessed with the Childhood Trauma Questionnaire Maternal and neonatal mitochondrial bioenergetics were quantitatively comparable and positively correlated. Female newborns showed higher mitochondrial respiration compared to male newborns. Maternal CM load was significantly and positively associated with mitochondrial respiration and density in mothers, but not with mitochondrial respiration in newborns. Although maternal and neonatal mitochondrial bioenergetics were positively correlated, maternal CM only had a small effect on mitochondrial density in newborns, which was not significant in this study after adjustment for multiple comparisons. The biological relevance of our finding and its consequences for child development need further investigation in future larger studies. This study reports data on mitochondrial bioenergetics of healthy mother-newborn dyads with varying degrees of CM.

The effects of childhood maltreatment on epigenetic regulation of stress-response associated genes: an intergenerational approach.

While biological alterations associated with childhood maltreatment (CM) have been found in affected individuals, it remains unknown to what degree these alterations are biologically transmitted to the next generation. We investigated intergenerational effects of maternal CM on DNA methylation and gene expression in N = 113 mother-infant dyads shortly after parturition, additionally accounting for the role of the FKBP5 rs1360780 genotype. Using mass array spectrometry, we assessed the DNA methylation of selected stress-response-associated genes (FK506 binding protein 51 [FKBP5], glucocorticoid receptor [NR3C1], corticotropin-releasing hormone receptor 1 [CRHR1]) in isolated immune cells from maternal blood and neonatal umbilical cord blood. In mothers, CM was associated with decreased levels of DNA methylation of FKBP5 and CRHR1 and increased NR3C1 methylation, but not with changes in gene expression profiles. Rs1360780 moderated the FKBP5 epigenetic CM-associated regulation profiles in a gene × environment interaction. In newborns, we found no evidence for any intergenerational transmission of CM-related methylation profiles for any of the investigated epigenetic sites. These findings support the hypothesis of a long-lasting impact of CM on the biological epigenetic regulation of stress-response mediators and suggest for the first time that these specific epigenetic patterns might not be directly transmitted to the next generation.

Intergenerational gene × environment interaction of FKBP5 and childhood maltreatment on hair steroids.

BACKGROUND: The inconsistency in results of cortisol alterations after childhood maltreatment (CM) might arise due to the fact that no study so far considered the effects of environmental factors such as maltreatment load and genetic factors such as the influence of FKBP5 genotype on stress hormone regulation. This study analyzed the interaction between the single nucleotide polymorphism rs1360780 within the FKBP5 gene and the severity of maternal CM experiences (maltreatment load) on hair steroid levels of mother-infant-dyads. METHODS: Hair samples of N = 474 mothers and N = 331 newborns were collected < 1 week after parturition enabling a retrospective assessment of cortisol, cortisone, and dehydroepiandrosterone (DHEA) using mass spectrometry. The sum score of the Childhood Trauma Questionnaire operationalized the maternal maltreatment load. DNA from whole blood or buccal cells was used for FKBP5 genotyping. RESULTS: The higher the maltreatment load, the higher maternal hair cortisol and cortisone levels in T allele carriers of FKBP5 rs1360780 were observed. Hair cortisol and DHEA levels of newborns with the T allele were reduced with an increasing maternal maltreatment load, while there was an increase of hair cortisol and DHEA in newborns homozygous for the C allele. CONCLUSIONS: This study is the very first uncovering a gene (FKBP5) × environment (maltreatment load) interaction on hair steroids in mothers and their offspring, indicating an intergenerational transmission of hypothalamic-pituitary-adrenal axis alterations. These results may help to explain the inconsistency in previous findings on steroid hormone alterations after chronic and traumatic stress and should be considered in future studies.

Epigenetic Alterations Associated with War Trauma and Childhood Maltreatment.

Survivors of war trauma or childhood maltreatment are at increased risk for trauma-spectrum disorders such as post-traumatic stress disorder (PTSD). In addition, traumatic stress has been associated with alterations in the neuroendocrine and the immune system, enhancing the risk for physical diseases. Traumatic experiences might even affect psychological as well as biological parameters in the next generation, i.e. traumatic stress might have transgenerational effects. This article outlines how epigenetic processes, which represent a pivotal biological mechanism for dynamic adaptation to environmental challenges, might contribute to the explanation of the long-lasting and transgenerational effects of trauma. In particular, epigenetic alterations in genes regulating the hypothalamus-pituitary-adrenal axis as well as the immune system have been observed in survivors of childhood and adult trauma. These changes could result in enduring alterations of the stress response as well as the physical health risk. Furthermore, the effects of parental trauma could be transmitted to the next generation by parental distress and the pre- and postnatal environment, as well as by epigenetic marks transmitted via the germline. While epigenetic research has a high potential of advancing our understanding of the consequences of trauma, the findings have to be interpreted with caution, as epigenetics only represent one piece of a complex puzzle of interacting biological and environmental factors. Copyright © 2015 John Wiley & Sons, Ltd.