Maternal DNA Methylation Research Articles

Methylation for making a placenta

DNA Methylation The trophoblast cells of a developing embryo give rise to extra-embryonic tissues that support embryonic growth, such as the placenta. The epigenetic modification DNA methylation is critical for the regulation of embryonic development. Branco et al. show that maternal DNA methylation also plays a role in trophoblast development. DAN methylation is required for the regulation of genes involved in cell adhesion and migration, processes required for proper placental formation that are perturbed in placental disorders such as pre-eclampsia. Dev. Cell 10.1016/j.devcel.2015.12.027 (2015).

Maternal DNA Methylation of TH17 Cytokine Genes in Second Half of Pregnancy Changes with Parity

Maternal DNA Methylation of TH17 Cytokine Genes in Second Half of Pregnancy Changes with Parity

Pervasive polymorphic imprinted methylation in the human placenta.

The maternal and paternal copies of the genome are both required for mammalian development, and this is primarily due to imprinted genes, those that are monoallelically expressed based on parent-of-origin. Typically, this pattern of expression is regulated by differentially methylated regions (DMRs) that are established in the germline and maintained after fertilization. There are a large number of germline DMRs that have not yet been associated with imprinting, and their function in development is unknown. In this study, we developed a genome-wide approach to identify novel imprinted DMRs in the human placenta and investigated the dynamics of these imprinted DMRs during development in somatic and extraembryonic tissues. DNA methylation was evaluated using the Illumina HumanMethylation450 array in 134 human tissue samples, publicly available reduced representation bisulfite sequencing in the human embryo and germ cells, and targeted bisulfite sequencing in term placentas. Forty-three known and 101 novel imprinted DMRs were identified in the human placenta by comparing methylation between diandric and digynic triploid conceptions in addition to female and male gametes. Seventy-two novel DMRs showed a pattern consistent with placental-specific imprinting, and this monoallelic methylation was entirely maternal in origin. Strikingly, these DMRs exhibited polymorphic imprinted methylation between placental samples. These data suggest that imprinting in human development is far more extensive and dynamic than previously reported and that the placenta preferentially maintains maternal germline-derived DNA methylation.

Maternal DNA Methylation Regulates Early Trophoblast Development

SummaryCritical roles for DNA methylation in embryonic development are well established, but less is known about its roles during trophoblast development, the extraembryonic lineage that gives rise to the placenta. We dissected the role of DNA methylation in trophoblast development by performing mRNA and DNA methylation profiling of Dnmt3a/3b mutants. We find that oocyte-derived methylation plays a major role in regulating trophoblast development but that imprinting of the key placental regulator Ascl2 is only partially responsible for these effects. We have identified several methylation-regulated genes associated with trophoblast differentiation that are involved in cell adhesion and migration, potentially affecting trophoblast invasion. Specifically, trophoblast-specific DNA methylation is linked to the silencing of Scml2, a Polycomb Repressive Complex 1 protein that drives loss of cell adhesion in methylation-deficient trophoblast. Our results reveal that maternal DNA methylation controls multiple differentiation-related and physiological processes in trophoblast via both imprinting-dependent and -independent mechanisms.

Whole-genome fetal and maternal DNA methylation analysis using MeDIP-NGS for the identification of differentially methylated regions.

DNA methylation is an epigenetic marker that has been shown to vary significantly across different tissues. Taking advantage of the methylation differences between placenta-derived cell-free DNA and maternal blood, several groups employed different approaches for the discovery of fetal-specific biomarkers. The aim of this study was to analyse whole-genome fetal and maternal methylomes in order to identify and confirm the presence of differentially methylated regions (DMRs). We have initially utilized methylated DNA immunoprecipitation (MeDIP) and next-generation sequencing (NGS) to identify genome-wide DMRs between chorionic villus sampling (CVS) and female non-pregnant plasma (PL) and peripheral blood (WBF) samples. Next, using specific criteria, 331 fetal-specific DMRs were selected and confirmed in eight CVS, eight WBF and eight PL samples by combining MeDIP and in-solution targeted enrichment followed by NGS. Results showed higher enrichment in CVS samples as compared to both WBF and PL samples, confirming the distinct methylation levels between fetal and maternal DNA for the selected DMRs. We have successfully implemented a novel approach for the discovery and confirmation of a significant number of fetal-specific DMRs by combining for the first time MeDIP and in-solution targeted enrichment followed by NGS. The implementation of this double-enrichment approach is highly efficient and enables the detailed analysis of multiple DMRs by targeted NGS. Also, this is, to our knowledge, the first reported application of MeDIP on plasma samples, which leverages the implementation of our enrichment methodology in the detection of fetal abnormalities in maternal plasma.

Length of paternal lifespan is manifested in the DNA methylome of their nonagenarian progeny.

The heritability of lifespan is 20-30%, but only a few genes associated with longevity have been identified. To explain this discrepancy, the inheritance of epigenetic features, such as DNA methylation, have been proposed to contribute to the heritability of lifespan.We investigated whether parental lifespan is associated with DNA methylation profile in nonagenarians. A regression model, adjusted for differences in blood cell proportions, identified 659 CpG sites where the level of methylation was associated with paternal lifespan. However, no association was observed between maternal lifespan and DNA methylation. The 659 CpG sites associated with paternal lifespan were enriched outside of CpG islands and were located in genes associated with development and morphogenesis, as well as cell signaling. The largest difference in the level of methylation between the progeny of the shortest-lived and longest-lived fathers was identified for CpG sites mapping to CXXC5. In addition, the level of methylation in three Notch-genes (NOTCH1, NOTCH3 and NOTCH4) was also associated with paternal lifespan.There are implications for the inheritance of acquired traits via epigenetic mechanisms in mammals. Here we describe DNA methylation features that are associated with paternal lifespan, and we speculate that the identified CpG sites may represent intergenerational epigenetic inheritance.

Genetic and epigenetic transgenerational implications related to omega-3 fatty acids. Part II: maternal FADS2 rs174575 genotype and DNA methylation predict toddler cognitive performance

Maternal transfer of fatty acids is important to fetal brain development. The prenatal environment may differentially affect the substrates supporting declarative memory abilities, as the level of fatty acids transferred across the placenta may be affected by the maternal fatty acid desaturase 2 (FADS2) rs174575 single nucleotide polymorphism. In this study, we hypothesized that toddler and maternal rs174575 genotype and FADS2 promoter methylation would be related to the toddlers’ declarative memory performance. Seventy-one 16-month-old toddlers participated in an imitation paradigm designed to test immediate and long-term declarative memory abilities. FADS2 rs174575 genotype was determined and FADS2 promoter methylation was quantified from blood by bisulfite pyrosequencing for the toddlers and their natural mothers. Toddlers of GG mothers at the FADS2 rs174575 single nucleotide polymorphism did not perform as well on memory assessments as toddlers of CC or CG mothers when controlling for plasma α-linolenic acid and child genotype. Toddler methylation status was related to immediate memory performance, whereas maternal methylation status was related to delayed memory performance. Thus, prenatal experience and maternal FADS2 status have a pervasive, long-lasting influence on the brain development of the offspring, but as the postnatal environment becomes more primary, the offsprings’ own biology begins to have an effect.

Genetic and epigenetic transgenerational implications related to omega-3 fatty acids. Part I: maternal FADS2 genotype and DNA methylation correlate with polyunsaturated fatty acid status in toddlers: an exploratory analysis

Polyunsaturated fatty acid metabolism in toddlers is regulated by a complex network of interacting factors. The contribution of maternal genetic and epigenetic makeup to this milieu is not well understood. In a cohort of mothers and toddlers 16 months of age (n = 65 mother-child pairs), we investigated the association between maternal genetic and epigenetic fatty acid desaturase 2 (FADS2) profiles and toddlers' n-6 and n-3 fatty acid metabolism. FADS2 rs174575 variation and DNA methylation status were interrogated in mothers and toddlers, as well as food intake and plasma fatty acid concentrations in toddlers. A multivariate fit model indicated that maternal rs174575 genotype, combined with DNA methylation, can predict α-linolenic acid plasma concentration in all toddlers and arachidonic acid concentrations in boys. Arachidonic acid intake was predictive for its plasma concentration in girls, whereas intake of 3 major n-3 species (eicosapentaenoic, docosapentaenoic, and docosahexaenoic acids) were predictive for their plasma concentrations in boys. FADS2 genotype and DNA methylation in toddlers were not related to plasma concentrations or food intakes, except for CpG8 methylation. Maternal FADS2 methylation was a predictor for the boys' α-linolenic acid intakes. This exploratory study suggests that maternal FADS2 genetic and epigenetic status could be related to toddlers' polyunsaturated fatty acid metabolism.

Early Pregnancy Maternal Blood DNA Methylation in Repeat Pregnancies and Change in Gestational Diabetes Mellitus Status—A Pilot Study.

Repeat pregnancies with different perinatal outcomes minimize underlying maternal genetic diversity and provide unique opportunities to investigate nongenetic risk factors and epigenetic mechanisms of pregnancy complications. We investigated gestational diabetes mellitus (GDM)-related differential DNA methylation in early pregnancy peripheral blood samples collected from women who had a change in GDM status in repeat pregnancies. Six study participants were randomly selected from among women who had 2 consecutive pregnancies, only 1 of which was complicated by GDM (case pregnancy) and the other was not (control pregnancy). Epigenome-wide DNA methylation was profiled using Illumina HumanMethylation 27 BeadChips. Differential Identification using Mixture Ensemble and false discovery rate (<10%) cutoffs were used to identify differentially methylated targets between the 2 pregnancies of each participant. Overall, 27 target sites, 17 hypomethylated (fold change [FC] range: 0.77-0.99) and 10 hypermethylated (FC range: 1.01-1.09), were differentially methylated between GDM and control pregnancies among 5 or more study participants. Novel genes were related to identified hypomethylated (such as NDUFC1, HAPLN3, HHLA3, and RHOG) or hypermethylated sites (such as SEP11, ZAR1, and DDR). Genes related to identified sites participated in cell morphology, cellular assembly, cellular organization, cellular compromise, and cell cycle. Our findings support early pregnancy peripheral blood DNA methylation differences in repeat pregnancies with change in GDM status. Similar, larger, and repeat pregnancy studies can enhance biomarker discovery and mechanistic studies of GDM.

Altered LINE-1 Methylation in Mothers of Children with Down Syndrome.

Down syndrome (DS, also known as trisomy 21) most often results from chromosomal nondisjunction during oogenesis. Numerous studies sustain a causal link between global DNA hypomethylation and genetic instability. It has been suggested that DNA hypomethylation might affect the structure and dynamics of chromatin regions that are critical for chromosome stability and segregation, thus favouring chromosomal nondisjunction during meiosis. Maternal global DNA hypomethylation has not yet been analyzed as a potential risk factor for chromosome 21 nondisjunction. This study aimed to asses the risk for DS in association with maternal global DNA methylation and the impact of endogenous and exogenous factors that reportedly influence DNA methylation status. Global DNA methylation was analyzed in peripheral blood lymphocytes by quantifying LINE-1 methylation using the MethyLight method. Levels of global DNA methylation were significantly lower among mothers of children with maternally derived trisomy 21 than among control mothers (P = 0.000). The combination of MTHFR C677T genotype and diet significantly influenced global DNA methylation (R2 = 4.5%, P = 0.046). The lowest values of global DNA methylation were observed in mothers with MTHFR 677 CT+TT genotype and low dietary folate. Although our findings revealed an association between maternal global DNA hypomethylation and trisomy 21 of maternal origin, further progress and final conclusions regarding the role of global DNA methylation and the occurrence of trisomy 21 are facing major challenges.

Maternal prenatal depressive symptoms predict infant NR3C1 1F and BDNF IV DNA methylation

Prenatal maternal psychological distress increases risk for adverse infant outcomes. However, the biological mechanisms underlying this association remain unclear. Prenatal stress can impact fetal epigenetic regulation that could underlie changes in infant stress responses. It has been suggested that maternal glucocorticoids may mediate this epigenetic effect. We examined this hypothesis by determining the impact of maternal cortisol and depressive symptoms during pregnancy on infant NR3C1 and BDNF DNA methylation. Fifty-seven pregnant women were recruited during the second or third trimester. Participants self-reported depressive symptoms and salivary cortisol samples were collected diurnally and in response to a stressor. Buccal swabs for DNA extraction and DNA methylation analysis were collected from each infant at 2 months of age, and mothers were assessed for postnatal depressive symptoms. Prenatal depressive symptoms significantly predicted increased NR3C1 1F DNA methylation in male infants (β = 2.147, P = 0.044). Prenatal depressive symptoms also significantly predicted decreased BDNF IV DNA methylation in both male and female infants (β = −3.244, P = 0.013). No measure of maternal cortisol during pregnancy predicted infant NR3C1 1F or BDNF promoter IV DNA methylation. Our findings highlight the susceptibility of males to changes in NR3C1 DNA methylation and present novel evidence for altered BDNF IV DNA methylation in response to maternal depression during pregnancy. The lack of association between maternal cortisol and infant DNA methylation suggests that effects of maternal depression may not be mediated directly by glucocorticoids. Future studies should consider other potential mediating mechanisms in the link between maternal mood and infant outcomes.

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Objectives Preeclampsia is a pregnancy-associated complex condition associated with inflammation, oxidative stress and angiogenic imbalance. Stress sensitive transcription factors nuclear factor-erythroid 2-like 1 (Nrf1) and nuclear factor-erythroid 2-like 2 (Nrf2) are involved in regulation of angiogenic, inflammatory and oxidative stress pathways. Recent evidence suggests a link between Nrf2 and angiogenic factor balance in preeclampsia though the degree to which maternal and placental DNA methylation contributes to disruption of Nrf pathways among women with preeclampsia is unknown. Our central hypothesis is that distinct patterns of DNA methylation in Nrf pathway and angiogenic genes contribute to altered angiogenic balance in preeclampsia. Methods Prospectively, we collected blood in each of three trimesters of pregnancy, maternal white blood cells in the first trimester of pregnancy and placental tissue at delivery from nulliparous women. We analyzed DNA methylation at individual CpG dinucleotides (Illumina Infinium) in Nrf and angiogenic (VEGFA, VEGFB, VEGC, FLT1, KDR, NRF1, PlGF) genes in maternal peripheral blood cells and placenta and circulating angiogenic factors (Millipore Bioplex) from women who developed preeclampsia and normotensive controls ( n = 6/group). Differences in average methylation beta scores (change in beta score >0.2 indicated significant gain of methylation; >−0.2 indicated significant loss of methylation) determined significant differences between groups. Results DNA methylation patterns in Nrf and angiogenic genes in maternal white blood cells and placenta were not significantly different in cases/controls. Among women with preeclampsia, PlGF was significantly lower across pregnancy and sFLT1 was significantly higher in the third trimester among women with preeclampsia. Conclusions DNA methylation in Nrf and angiogenic genes was not associated with altered angiogenic balance in women who developed late onset preeclampsia. Differences in DNA methylation may occur later in pregnancy when circulating factors differed between groups. Disclosures C.M. Anderson: None. J.L. Ralph: None. J.E. Ohm: None.

Associations between Maternal Circulating DNA Methylation during Pregnancy and Exposures to Environmental Metals in the National Children’s Study

Associations between Maternal Circulating DNA Methylation during Pregnancy and Exposures to Environmental Metals in the National Children’s StudyAbstract Number:2816 Kelly Bakulski*, Jason Feinberg, Shannon Brown, Laura Caulfield, Lynn Goldman, Andrew Feinberg, and Dani Fallin Kelly Bakulski* Johns Hopkins University, United States, E-mail Address: [email protected] Search for more papers by this author , Jason Feinberg Johns Hopkins University, United States, E-mail Address: [email protected] Search for more papers by this author , Shannon Brown Johns Hopkins University, United States, E-mail Address: [email protected] Search for more papers by this author , Laura Caulfield Johns Hopkins University, United States, E-mail Address: [email protected] Search for more papers by this author , Lynn Goldman George Washington University, United States, E-mail Address: [email protected] Search for more papers by this author , Andrew Feinberg Johns Hopkins University, United States, E-mail Address: [email protected] Search for more papers by this author , and Dani Fallin Johns Hopkins University, United States, E-mail Address: [email protected] Search for more papers by this author AbstractBackground: Early life environments can influence later life health outcomes and DNA methylation may represent a persistent mark of these conditions. The relationships between maternal metals exposures in early pregnancy and subsequent DNA methylation measurements in moms and babies are poorly understood.Methods: Blood and urine metals exposures were measured in 138 pregnant women enrolled in the National Children’s Study Vanguard sites during trimester one. Repeated measures of buffy coat genome-wide DNA methylation were assessed on the Illumina 450K array during trimesters one and three as well as from the cord blood in a subset of babies (n=81). We tested for region and site specific associations between DNA methylation and metal exposures, adjusting for cell type composition and other potential confounding factors.Results: In our study population, mean blood cadmium exposure was 0.27 µg/L (95% CI: 0.11-0.62), mean blood manganese was 11.7 µg/L (95% CI: 7.2-19.7), mean blood lead was 0.52 µg/dL (95% CI: 0.18-1.14), mean blood selenium was 178.7 µg/L (95% CI: 143.1-210.3), and mean total blood mercury was 0.99 µg/L (95% CI: 0.11-3.3). Fifteen CpG sites were associated with cadmium exposure in trimester one at a genome-wide significance level of p-value<10-7 and ten of those sites replicated in terms of magnitude and direction in trimester three. Similarly, fifteen sites reached genome-wide significance in trimester one for mercury and fourteen of those replicated in trimester three.Conclusions: We observed maternal DNA methylation differences with trimester one cadmium and mercury exposure levels that persisted to trimester three DNA methylation in a cohort of normal births. The identified regions may act as biomarkers of exposure in cohorts lacking prenatal exposure information. Cord blood methylation and urinary metals exposures will be discussed.

Prenatal exposure to maternal cigarette smoking and DNA methylation: epigenome-wide association in a discovery sample of adolescents and replication in an independent cohort at birth through 17 years of age.

Background: Prenatal exposure to maternal cigarette smoking (prenatal smoke exposure) had been associated with altered DNA methylation (DNAm) at birth.Objective: We examined whether such alterations are present from birth through adolescence.Methods: We used the Infinium HumanMethylation450K BeadChip to search across 473,395 CpGs for differential DNAm associated with prenatal smoke exposure during adolescence in a discovery cohort (n = 132) and at birth, during childhood, and during adolescence in a replication cohort (n = 447).Results: In the discovery cohort, we found five CpGs in MYO1G (top-ranking CpG: cg12803068, p = 3.3 × 10–11) and CNTNAP2 (cg25949550, p = 4.0 × 10–9) to be differentially methylated between exposed and nonexposed individuals during adolescence. The CpGs in MYO1G and CNTNAP2 were associated, respectively, with higher and lower DNAm in exposed versus nonexposed adolescents. The same CpGs were differentially methylated at birth, during childhood, and during adolescence in the replication cohort. In both cohorts and at all developmental time points, the differential DNAm was in the same direction and of a similar magnitude, and was not altered appreciably by adjustment for current smoking by the participants or their parents. In addition, four of the five EWAS (epigenome-wide association study)–significant CpGs in the adolescent discovery cohort were also among the top sites of differential methylation in a previous birth cohort, and differential methylation of CpGs in CYP1A1, AHRR, and GFI1 observed in that study was also evident in our discovery cohort.Conclusions: Our findings suggest that modifications of DNAm associated with prenatal maternal smoking may persist in exposed offspring for many years—at least until adolescence.Citation: Lee KW, Richmond R, Hu P, French L, Shin J, Bourdon C, Reischl E, Waldenberger M, Zeilinger S, Gaunt T, McArdle W, Ring S, Woodward G, Bouchard L, Gaudet D, Davey Smith G, Relton C, Paus T, Pausova Z. 2015. Prenatal exposure to maternal cigarette smoking and DNA methylation: epigenome-wide association in a discovery sample of adolescents and replication in an independent cohort at birth through 17 years of age. Environ Health Perspect 123:193–199; http://dx.doi.org/10.1289/ehp.1408614

Human in vitro oocyte maturation is not associated with increased imprinting error rates at LIT1, SNRPN, PEG3 and GTL2

Does in vitro maturation (IVM) of cumulus-enclosed germinal vesicle (GV) stage oocytes retrieved from small antral follicles in minimally stimulated cycles without an ovulatory hCG dose induce imprinting errors at LIT1, SNRPN, PEG3 and GTL2 in human oocytes? There is no significant increase in imprinting mutations at LIT1, SNRPN, PEG3 and GTL2 after IVM of cumulus-enclosed GV oocytes from small antral follicles in minimally stimulated cycles without hCG priming. Animal models have generally demonstrated correct methylation imprint establishment for in vitro grown and matured oocytes. For human IVM, well-designed studies allowing conclusions on imprint establishment are currently not available. Immature oocyte-cumulus complexes from 2 to 9 mm follicles were retrieved in polycystic ovary syndrome (PCOS) subjects in minimally stimulated cycles without hCG priming and matured in vitro. In vivo grown oocytes were retrieved after conventional ovarian stimulation for IVF/ICSI or after ovulation induction. Imprinting error rates at three maternally methylated (LIT1, SNRPN and PEG3) and one paternally methylated (GTL2) imprinted genes were compared in 71 in vitro and 38 in vivo matured oocytes. The limiting dilution bisulfite sequencing technique was applied, allowing increased sensitivity based on multiplex PCR for the imprinted genes and the inclusion of non-imprinted marker genes for cumulus cell DNA contamination. In vitro as well as in vivo matured oocytes showed only a few abnormal alleles, consistent with epimutations. The abnormalities were more frequent in immature than in mature oocytes for both groups, although no significant difference was reached. There was no statistically significant increase in imprinting errors in IVM oocytes. This single cell methylation analysis was restricted to a number of well-selected imprinted genes. Genome-wide methylation analysis of single human oocytes is currently not possible. IVM is a patient-friendly alternative to conventional ovarian stimulation in PCOS patients and is associated with reduced gonadotrophin costs and avoidance of OHSS. The results of this study show for the first time that optimized human IVM procedures have no significant effects on the establishment of maternal DNA methylation patterns at LIT1, SNRPN, PEG3 and GTL2. This study was supported by research funds from Agentschap voor Innovatie door Wetenschap en Technologie (IWT-TBM 110680), Wetenschappelijk Fonds Willy Gepts (WFWG 2011) and German Research Foundation (HA 1374/12-2). There are no competing interests.

Maternal smoking and DNA methylation in newborns: in utero effect or epigenetic inheritance?

Maternal smoking in pregnancy is associated with adverse health outcomes in children, including cancers; underlying mechanisms may include epigenetic modifications. Using Illumina's 450K array, we previously identified differential DNA methylation related to maternal smoking during pregnancy at 26 CpG sites (CpGs) in 10 genes in newborn cord bloods from the Norwegian Mother and Child Cohort Study (MoBa). Whether these methylation signals in newborns reflect in utero exposure only or possibly epigenetic inheritance of smoking-related modifications is unclear. We therefore evaluated the impact of the timing of mother's smoking (before or during pregnancy using cotinine measured at 18 weeks gestation), the father's smoking before conception, and the grandmother's smoking during her pregnancy with the mother on methylation at these 26 CpGs in 1,042 MoBa newborns. We used robust linear regression, adjusting for covariates, applying Bonferroni correction. The strongest and only statistically significant associations were observed for sustained smoking by the mother during pregnancy through at least gestational week 18 (P < 1.6 × 10(-5) for all 26 CpGs). We observed no statistically significant differential methylation due to smoking by the mother before pregnancy or that ceased by week 18, father's smoking before conception, or grandmother's smoking while pregnant with the mother. Differential methylation at these CpGs in newborns seems to reflect sustained in utero exposure rather than epigenetic inheritance. Smoking cessation in early pregnancy may negate effects on methylation. Analyses of maternal smoking during pregnancy and offspring health outcomes, including cancer, limited to ever smoking might miss true associations. Cancer Epidemiol Biomarkers Prev; 23(6); 1007-17. ©2014 AACR.

Litter and sex effects on maternal behavior and DNA methylation of the Nr3c1 exon 17 promoter gene in hippocampus and cerebellum

Early life events can alter gene expression through DNA methylation. The methylation status of the exon 17 promoter of the glucocorticoid receptor (Nr3c1 gene) in hippocampus associates with frequency of pup licking. Much of this work was conducted with male rats. Because dams more frequently lick male pups, this may contribute to sex differences in phenotypes through DNA methylation. Modifying litter gender composition (LGC), in which offspring of single-sex litters are compared to mixed-sex litters, alters maternal behavior. Previously, we demonstrated that LGC and sex affected pup licking times as well as anxiety and hippocampal DNA methylation of the Nr3c1 exon 17 promoter gene in adolescence. Now, we expand upon this work by examining effects in cerebellum and measuring mRNA levels. We also re-assessed DNA methylation in hippocampus using pyrosequencing and re-analyzed pup licking with the more commonly used frequency measure. Litters, culled to 8 pups on postnatal day 1 (PN1), were assigned to one of three conditions: all male (n=10), all female (n=12), or half of each sex (n=20). Licking was rated on PN4, 7, and 10. On PN35, hippocampal and cerebellar samples were obtained. Single-sex males were licked the least and mixed-sex males, the most. Hippocampal Nr3c1 mRNA levels were lowest in mixed females with no LGC or Sex effects in DNA methylation. Cerebellar DNA methylation levels were lowest in mixed males with no effect on mRNA levels. Maternal pup licking associated with DNA methylation of the Nr3c1 exon 17 promoter gene in cerebellum and with hippocampal mRNA.

Association of homocysteine with global DNA methylation in vegetarian Indian pregnant women and neonatal birth anthropometrics

Objective: The present study was designed to evaluate if plasma maternal folate, vitamin B-12 and homocysteine levels had an effect on maternal global DNA methylation and neonatal anthropometrics in Indian pregnant women.Methods: A total of 49 participants having completed ≥36 weeks of pregnancy were enrolled in the study. Estimation of folate was by Ion capture assay, vitamin B-12 by microparticle enzyme immunoassay, total homocysteine by fluorescence polarization immunoassay and global DNA methylation using Cayman’s DNA methylation enzyme immunoassay (EIA) kit.Results: Folate and vitamin B-12 were inversely correlated to homocysteine in pregnant women consuming vegetarian and non-vegetarian diet. No difference in global DNA methylation was found between the vegetarian and non-vegetarian pregnant women. Folate and vitamin B-12 did not show association with global DNA methylation, however plasma total homocysteine of the vegetarian group showed significant correlation to global DNA methylation (r2 = 0.49, p = 0.011). Plasma total homocysteine was inversely related to tricep skinfold (r2 = −0.484, p = 0.01) and chest circumference (r2 = −0.104, p = 0.04) of neonates in vegetarian group.Conclusion: Moderate vitamin B-12 deficiency in vegetarian pregnant women might be the cause of hyperhomocystinemia, hypermethylation when compared to vitamin B-12 sufficient non-vegetarian group.

Maternal stress, preterm birth, and DNA methylation at imprint regulatory sequences in humans.

In infants exposed to maternal stress in utero, phenotypic plasticity through epigenetic events may mechanistically explain increased risk of preterm birth (PTB), which confers increased risk for neurodevelopmental disorders, cardiovascular disease, and cancers in adulthood. We examined associations between prenatal maternal stress and PTB, evaluating the role of DNA methylation at imprint regulatory regions. We enrolled women from prenatal clinics in Durham, NC. Stress was measured in 537 women at 12 weeks of gestation using the Perceived Stress Scale. DNA methylation at differentially methylated regions (DMRs) associated with H19, IGF2, MEG3, MEST, SGCE/PEG10, PEG3, NNAT, and PLAGL1 was measured from peripheral and cord blood using bisulfite pyrosequencing in a sub-sample of 79 mother–infant pairs. We examined associations between PTB and stress and evaluated differences in DNA methylation at each DMR by stress. Maternal stress was not associated with PTB (OR = 0.98; 95% CI, 0.40–2.40; P = 0.96), after adjustment for maternal body mass index (BMI), income, and raised blood pressure. However, elevated stress was associated with higher infant DNA methylation at the MEST DMR (2.8% difference, P < 0.01) after adjusting for PTB. Maternal stress may be associated with epigenetic changes at MEST, a gene relevant to maternal care and obesity. Reduced prenatal stress may support the epigenomic profile of a healthy infant.

158: Genome wide maternal leukocyte DNA methylation analysis performed longitudinally throughout normal pregnancy

Pregnancy is associated with significant change in leukocyte composition/function in the establishment/adaptation to pregnancy and parturition. We characterized methylation profiles longitudinally in a group of healthy pregnant women.