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DNA methylation of imprinted genes in Mexican–American newborn children with prenatal phthalate exposure
Epigenomics
Abstract
Aim:
Imprinted genes exhibit expression in a parent-of-origin-dependent manner and are critical for child development. Recent limited evidence suggests that prenatal exposure to phthalates, ubiquitous endocrine disruptors, can affect their epigenetic dysregulation.
Materials & methods:
We quantified DNA methylation of nine imprinted gene differentially methylated regions by pyrosequencing in 296 cord blood DNA samples in a Mexican-American cohort. Fetal exposure was estimated by phthalate metabolite concentrations in maternal urine samples during pregnancy.
Results:
Several differentially methylated regions of imprinted genes were associated with high molecular weight phthalates. The most consistent, positive, and false discovery rate significant associations were observed for MEG3.
Conclusion:
Phthalate exposure in utero may affect methylation status of imprinted genes in newborn children.
... Some studies in humans have demonstrated that developmental phthalate exposure is connected with an altered epigenome in early life, and scientists have subsequently suggested that these chemicals can have a significant impact on the epigenome and associated long-term health (LaRocca et al., 2014;Wang I.-J. et al., 2015;Goodrich et al., 2016;Huen et al., 2016;Solomon et al., 2017;Chen et al., 2018;Huang et al., 2018;Montrose et al., 2018;Tindula et al., 2018;Miura et al., 2021). Most of these studies, however, utilized targeted techniques to examine how methylation in a small number of genes and genetic elements is associated with phthalate exposure. ...
... March 2022 | Volume 13 | Article 793278 decade, mounting evidence has linked exposure to classically used phthalates with a number of significant human health effects. There is a growing body of evidence suggesting that prenatal phthalate exposure is connected with epigenetic effects in developing infants (LaRocca et al., 2014;Wang IJ. et al., 2015;Goodrich et al., 2016;Huen et al., 2016;Solomon et al., 2017;Chen et al., 2018;Huang et al., 2018;Montrose et al., 2018;Tindula et al., 2018;Miura et al., 2021). The main analysis in this manuscript combined data from two different newborn sample types (blood spots and cord blood) to examine DNA methylation across the genome in relation to maternal exposure to metabolites of six different commonly used phthalates and one nonspecific metabolite (ΣDEHP, DEP, ΣDiNP, DiBP, DnBP, BBzP, and MCPP). ...
... Enriched pathways presently echoed these proposed mechanisms, with altered DNA methylation in pathways related to TNF-β signaling (BBzP in females), androgen signaling (ΣDEHP in females), and mitochondrial functions (ΣDEHP in males; DnBP in females). There were not, however, differences in either the methylation of CpG sites or the regulatory pathways related to any key imprinted genes that have previously been reported to be differentially methylated, including IGF2, H19, and MEG3 (LaRocca et al., 2014;Goodrich et al., 2016;Montrose et al., 2018;Tindula et al., 2018), which may be due to differences in EPIC array CpGs and specific CpGs that have been examined in pyrosequencing assays. Pathways were also quite different between infant sex models, once again highlighting the importance of sexstratified models when it comes to these chemicals. ...
Phthalates are a diverse group of chemicals used in consumer products. Because they are so widespread, exposure to these compounds is nearly unavoidable. Recently, growing scientific consensus has suggested that phthalates produce health effects in developing infants and children. These effects may be mediated through mechanisms related to the epigenome, the constellation of mitotically heritable chemical marks and small compounds that guide transcription and translation. The present study examined the relationship between prenatal, first-trimester exposure of seven phthalates and epigenetics in two pregnancy cohorts (n = 262) to investigate sex-specific alterations in infant blood DNA methylation at birth (cord blood or neonatal blood spots). Prenatal exposure to several phthalates was suggestive of association with altered DNA methylation at 4 loci in males (all related to ΣDEHP) and 4 loci in females (1 related to ΣDiNP; 2 related to BBzP; and 1 related to MCPP) at a cutoff of q < 0.2. Additionally, a subset of dyads (n = 79) was used to interrogate the relationships between two compounds increasingly used as substitutions for common phthalates (ΣDINCH and ΣDEHTP) and cord blood DNA methylation. ΣDINCH, but not ΣDEHTP, was suggestive of association with DNA methylation (q < 0.2). Together, these results demonstrate that prenatal exposure to both classically used phthalate metabolites and their newer alternatives is associated with sex-specific infant DNA methylation. Research and regulatory actions regarding this chemical class should consider the developmental health effects of these compounds and aim to avoid regrettable substitution scenarios in the present and future.
... Prior EWAS and candidate gene studies in human populations have examined the associations between maternal urinary phthalate concentrations during pregnancy and DNAm in the placenta (LaRocca et al., 2014;Zhao et al., 2016Zhao et al., , 2015 and umbilical cord blood Huang et al., 2018;Huen et al., 2016;Miura et al., 2021;Solomon et al., 2017;Tindula et al., 2018). In placental tissue, maternal urinary concentrations of DEHP and associated metabolites have been associated with DNAm of growth-related genes (e.g., IGF2, AHRR) (LaRocca et al., 2014;Zhao et al., 2016). ...
... A Chinese cohort study reported that maternal mono-benzyl phthalate (MBzP) exposure was associated with cord blood methylation patterns in transposable elements (i.e., LINE-1, Alu) in a sample of 53 female infants . The Centre for the Health Assessment of Mothers and Children of Silanas (CHAMACOS), a larger study involving 296-355 mother-child pairs of Mexican-Americans, reported that mono-ethyl phthalate (MEP) was associated with cord blood DNAm in LINE-1 and Alu (Huen et al., 2016), and that HMWP metabolites were associated with differentially methylated regions for genes involved with endocrine function (i.e., CNPY1), inflammatory response (i.e., IRAK4, ESM1), growth (i.e., MEG3), and male reproductive function (e.g., TESC, PRDM8) (Solomon et al., 2017;Tindula et al., 2018). Recent work on 203 mother-child pairs from the Hokkaido Study on Environment and Children's Health found that prenatal exposure to mono-2-ethylhexyl phthalate (MEHP) was related to cord blood DNAm in two CpGs that may be related to fetal growth (Miura et al., 2021). ...
... Specifically, DNAm alterations were examined in venous buffy coat blood from 74 infants and in BECs from 78 infants. Analyses involved the examination of 67 a priori candidate CpGs and/or associated genes identified in previous studies that investigated associations between prenatal phthalate exposure and DNAm variation in placental tissue and umbilical cord blood Huang et al., 2018;Huen et al., 2016;LaRocca et al., 2014;Miura et al., 2021;Solomon et al., 2017;Tindula et al., 2018;Zhao et al., 2016), and an exploratory association study that examined the remaining variable CpGs. Post-hoc analyses investigated cross-tissue effects, biological effect differences for individual phthalate metabolites, genomic features, comparison of peripheral and brain DNAm, and potential confounders (e.g., prenatal exposure to other EDCs such as bisphenol A (BPA)). ...
Background:
Prenatal exposure to phthalates has been associated with adverse health and neurodevelopmental outcomes. DNA methylation (DNAm) alterations may be a mechanism underlying these effects, but prior investigations of prenatal exposure to phthalates and neonatal DNAm profiles are limited to placental tissue and umbilical cord blood.
Objective:
Conduct an epigenome-wide association study (EWAS) of the associations between prenatal exposure to phthalates and DNAm in two accessible infant tissues, venous buffy coat blood and buccal epithelial cells (BECs).
Methods:
Participants included 152 maternal-infant pairs from the Alberta Pregnancy Outcomes and Nutrition (APrON) study. Maternal second trimester urine samples were analyzed for nine phthalate metabolites. Blood (n = 74) or BECs (n = 78) were collected from 3-month-old infants and profiled for DNAm using the Infinium HumanMethylation450 (450K) BeadChip. Robust linear regressions were used to investigate the associations between high (HMWPs) and low molecular weight phthalates (LMWPs) and change in methylation levels at variable Cytosine-phosphate-Guanine (CpG) sites in infant tissues, as well as the sensitivity of associations to potential confounders.
Results:
One candidate CpG in gene RNF39 reported by a previous study examining prenatal exposure to phthalates and cord blood DNAm was replicated. The EWAS identified 12 high-confidence CpGs in blood and another 12 in BECs associated with HMWPs and/or LMWPs. Prenatal exposure to bisphenol A (BPA) associated with two of the CpGs associated with HMWPs in BECs.
Discussion:
Prenatal exposure to phthalates was associated with DNAm variation at CpGs annotated to genes associated with endocrine hormone activity (i.e., SLCO4A1, TPO), immune pathways and DNA damage (i.e., RASGEF1B, KAZN, HLA-A, MYO18A, DIP2C, C1or109), and neurodevelopment (i.e., AMPH, NOTCH3, DNAJC5). Future studies that characterize the stability of these associations in larger samples, multiple cohorts, across tissues, and investigate the potential associations between these biomarkers and relevant health and neurodevelopmental outcomes are needed.
... Indeed, epigenetics has been recently explored in a few studies in placental models: Meruvu et al. (2016aMeruvu et al. ( , 2016b have reported a downregulation of ROS-responsive miRNAs expression in HTR-8/Svneo cells treated with MEHP; Kang et al. (2011) described an alteration in the expression of imprinted genes in placentas from mice under treatment with DEHP; LaRocca et al. (2014) described an association between an increased deviation of allele-specific expression of H19 and DEHP metabolites in urine; Zhao et al. (2015) found a negative association between urinary levels of DEHP metabolites and DNA methylation in LINE-1 sequences; Tindula et al. (2018) reported an association between DEHP in utero exposure and an alteration in DNA methylation status of the imprinted gene MEG3 in placental tissue; and Machtinger et al. (2018) found a correlation between DEHP exposure and increased levels of several lncRNAs in the placenta (Kang et al., 2011;LaRocca et al., 2014;Machtinger et al., 2018;Meruvu et al., 2016aMeruvu et al., , 2016bTindula et al., 2018;Zhao et al., 2015). These studies reflect the relevance of deciphering the effects of the exposure to DEHP and MEHP, which may deregulate very important epigenetic processes as genomic imprinting and silencing of repetitive elements in the genome that in turn are fundamental to development and health. ...
... Indeed, epigenetics has been recently explored in a few studies in placental models: Meruvu et al. (2016aMeruvu et al. ( , 2016b have reported a downregulation of ROS-responsive miRNAs expression in HTR-8/Svneo cells treated with MEHP; Kang et al. (2011) described an alteration in the expression of imprinted genes in placentas from mice under treatment with DEHP; LaRocca et al. (2014) described an association between an increased deviation of allele-specific expression of H19 and DEHP metabolites in urine; Zhao et al. (2015) found a negative association between urinary levels of DEHP metabolites and DNA methylation in LINE-1 sequences; Tindula et al. (2018) reported an association between DEHP in utero exposure and an alteration in DNA methylation status of the imprinted gene MEG3 in placental tissue; and Machtinger et al. (2018) found a correlation between DEHP exposure and increased levels of several lncRNAs in the placenta (Kang et al., 2011;LaRocca et al., 2014;Machtinger et al., 2018;Meruvu et al., 2016aMeruvu et al., , 2016bTindula et al., 2018;Zhao et al., 2015). These studies reflect the relevance of deciphering the effects of the exposure to DEHP and MEHP, which may deregulate very important epigenetic processes as genomic imprinting and silencing of repetitive elements in the genome that in turn are fundamental to development and health. ...
Di(2-ethylhexyl) phthalate (DEHP) is a chemical widely distributed in the environment as is extensively used in the plastic industry. DEHP is considered an endocrine disruptor chemical (EDC) and humans are inevitably and unintentionally exposed to this EDC through several sources including food, beverages, cosmetics, medical devices, among others. DEHP exposure has been associated and may be involved in the development of various pathologies; importantly, pregnant women are a particular risk group considering that endocrine alterations during gestation may impact fetal programming leading to the development of several chronic diseases in adulthood. Recent studies have indicated that exposure to DEHP and its metabolite Mono(2-ethylhexyl) phthalate (MEHP) may impair placental development and function, which in turn would have a negative impact on fetal growth.
Studies performed in several trophoblastic and placental models have shown the negative impact of DEHP and MEHP in key processes related to placental development such as implantation, differentiation, invasion and angiogenesis. In addition, many alterations in placental functions like hormone signaling, metabolism, transfer of nutrients, immunomodulation and oxidative stress response have been reported. Moreover, clinical-epidemiological evidence supports the association between DEHP exposure and adverse pregnancy outcomes and pathologies. In this review, we aim to summarize for the first time current knowledge about the impact of DEHP and MEHP exposure on placental development and pathophysiology, as well as the mechanisms involved. We also remark the importance of exploring DEHP and MEHP effects in different trophoblast cell populations and discuss new perspectives regarding this topic.
... There is growing evidence that environmentally-induced epigenetic perturbations, especially during susceptible periods of development such as gestation, can persist throughout life. Exposure to phthalates both in utero and in childhood has been associated with DNA methylation at specific genes, including imprinted genes such as H19, and repetitive elements (20)(21)(22). For example, phthalate exposures both in utero and later in development have been shown to correlate with DNA methylation of H19 and HSD11B2 in peri-adolescent children (20). ...
... Biological mechanisms such as epigenetic alteration may underlie susceptibility to effects on adiposity from phthalate exposures at key developmental windows. While evidence exists for the influence of phthalates, especially gestational exposure, on the epigenome (20)(21)(22)36), this is the first study to examine DNA methylation as a mediator between exposure and adiposity. The H19 imprint control region (ICR) and promoter of HSD11B2 were interrogated due to the functions of these genes in growth regulation as well as previous associations in the ELEMENT cohort between phthalate exposures and DNA methylation at these genes (20). ...
Phthalates are a class of endocrine disrupting chemicals with near ubiquitous exposure to populations around the world. Phthalates have been associated with children's adiposity in previous studies, though discrepancies exist across studies that may be due to timing of exposure or outcome assessment and population differences (i.e., genetics, other confounders). DNA methylation, an epigenetic modification involved in gene regulation, may mediate the effects of early life phthalate exposures on health outcomes. This study aims to evaluate the mediating effect of DNA methylation at growth-related genes on the association between phthalate exposure and repeat measures of adiposity (BMI-for-age z-score, waist circumference, and skinfolds thickness) in Mexican children. Urinary phthalate metabolite concentrations were quantified in mothers at each of the three trimesters of pregnancy and in children at the first peri-adolescent study visit. Blood leukocyte DNA methylation at H19 and HSD11B2 was quantified during the first peri-adolescent visit, and adiposity was measured at the first visit and again ~3 years later among participants (n = 109 boys, 114 girls) from the Early Life Exposure in Mexico to Environmental Toxicants (ELEMENT) project. Associations between phthalates or DNA methylation and repeat outcome measures were assessed separately in boys and girls using generalized estimating equation models including covariates (urinary specific gravity, maternal education, and child's age). Sobel tests were used to assess DNA methylation as a mediator in models adjusting for the same covariates. Associations between phthalates and adiposity varied by phthalate and timing of exposure. Early gestation MBP, MIBP, and MBzP were associated with adiposity among girls. For example, among girls first trimester maternal urine concentrations of MIBP were associated with increases in skinfold thickness, BMI-for-age, and waist circumference (p < 0.01). Second trimester and adolescent MBzP were associated with adiposity among boys in opposite directions. In girls, H19 methylation was positively associated with skinfold thickness. No significant mediation of phthalate exposure on adiposity by DNA methylation of H19 or HSD11B2 was observed (Sobel p > 0.05). However, the mediation analysis was underpowered to detect small to medium effect sizes, and the role of DNA methylation as a mediator between phthalates and outcomes merits further study.
... Ena od njih je poročala o spremenjenem metilomu posteljice ob izpostavljenosti KMES (ftalatom) (79). Več raziskav je povezalo izpostavljenost KMES s spremembami metilacije DNA izbranih genov v posteljici (80) in popkovnični krvi (81)(82)(83). Geni v teh spremenjenih področjih so povezani z vnetnim odzivom, kancerogenezo, endokrinim sistemom, moško plodnostjo (84)(85)(86) in andogenim odgovorom (86). ...
Ljudje smo vsakodnevno izpostavljeni različnim kemičnim motilcem endokrinega sistema (KMES). Izpostavljenost KMES je povezana s številnimi motnjami in boleznimi reproduktivnega sistema, vendar njihov vpliv še ni natančno pojasnjen. Opisanih je več različnih mehanizmov preko katerih KMES lahko motijo delovanje organizma. Reproduktivno zdravje je odvisno od pravilnega prenatalnega razvoja jajčnikov, ki je ključen za njihovo pravilno delovanje. Ugotovitve epidemioloških študij in študij na živalih kažejo, da prenatalna izpostavljenost KMES lahko povzroči različne bolezni reproduktivnega sistema pozneje v življenju, kar povzema hipoteza sindroma ovarijske disgeneze. Eden od vzročnih mehanizmov delovanja KMES za pojav sindroma ovarijske disgeneze naj bi bile tudi epigenetske spremembe. V članku, ki je pregled literature na tem področju, je predstavljena vloga epigenetskih procesov (DNA metilacija, ne-kodirajoča RNA, histonske modifikacije) v toksičnem delovanju KMES na reproduktivni sistem.
... In this regard, urinary MEOHP and MEHHP levels have reported a negative correlation with IGF2 DNA methylation, especially when fetal growth restriction takes place [125]. Phthalate metabolites also seem to modify the methylation of Maternally Expressed 3 (MEG3) gene, which is tightly linked to early growth, metabolism, and tumorigenesis, according to a longitudinal birth cohort study carried out by Tindula et al. [126]. The most relevant effects of phthalates on pregnancy and possible associated complications are shown in Figure 3. ...
Endocrine disrupting chemicals (EDCs) are exogenous substances widely disseminated both in the environment and in daily-life products which can interfere with the regulation and function of the endocrine system. These substances have gradually entered the food chain, being frequently found in human blood and urine samples. This becomes a particularly serious issue when they reach vulnerable populations such as pregnant women, whose hormones are more unstable and vulnerable to EDCs. The proper formation and activity of the placenta, and therefore embryonic development, may get seriously affected by the presence of these chemicals, augmenting the risk of several pregnancy complications, including intrauterine growth restriction, preterm birth, preeclampsia, and gestational diabetes mellitus, among others. Additionally, some of them also exert a detrimental impact on fertility, thus hindering the reproductive process from the beginning. In several cases, EDCs even induce cross-generational effects, inherited by future generations through epigenetic mechanisms. These are the reasons why a proper understanding of the reproductive and gestational alterations derived from these substances is needed, along with efforts to establish regulations and preventive measures in order to avoid exposition (especially during this particular stage of life).
... Manhattan plot of associations between a mixture of phthalates and bisphenols during first (A), second (B) and third (C) trimester with DNA methylation at birth among girls. In all Manhattan plots, the x-axis represents the autosomal chromosomes, the y-axis represents the −log 10 of the p value and the dots represent CpGs analyses using pyrosequencing [19,44,45]. Different methods have also been used to model chemical exposure, including exposure in the 25th percentile versus the 75th percentile, comparing subjects above and below the 75th percentile, or including the chemicals separately as continuous variables [17, 20-24, 27, 29, 44, 46]. ...
Background
Phthalates and bisphenols are non-persistent endocrine disrupting chemicals that are ubiquitously present in our environment and may have long-lasting health effects following fetal exposure. A potential mechanism underlying these exposure–outcome relationships is differential DNA methylation. Our objective was to examine the associations of maternal phthalate and bisphenol concentrations during pregnancy with DNA methylation in cord blood using a chemical mixtures approach.
Methods
This study was embedded in a prospective birth cohort study in the Netherlands and included 306 participants. We measured urine phthalates and bisphenols concentrations in the first, second and third trimester. Cord blood DNA methylation in their children was processed using the Illumina Infinium HumanMethylation450 BeadChip using an epigenome-wide association approach. Using quantile g-computation, we examined the association of increasing all mixture components by one quartile with cord blood DNA methylation.
Results
We did not find evidence for statistically significant associations of a maternal mixture of phthalates and bisphenols during any of the trimesters of pregnancy with DNA methylation in cord blood (all p values > 4.01 * 10–8). However, we identified one suggestive association (p value < 1.0 * 10–6) of the first trimester maternal mixture of phthalates and bisphenols and three suggestive associations of the second trimester maternal mixture of phthalates and bisphenols with DNA methylation in cord blood.
Conclusions
Although we did not identify genome-wide significant results, we identified some suggestive associations of exposure to a maternal mixture of phthalates and bisphenols in the first and second trimester with DNA methylation in cord blood that need further exploration in larger study samples.
... 49 Phthalates were also found to disrupt oocyte development and maturation and have been linked to differentially methylated regions of genes associated with metabolic processes and early growth and tumorigenesis. 72,73 Although the effects of EDCs are often more pronounced when the exposure occurs early in life, epigenetic changes following phthalate exposure are not exclusive to exposure during early development. Differentially methylated thyroid receptor-interacting protein 6 (TRIP6) gene promoters were found in peripubertal children exposed to phthalates. ...
To understand the effects of endocrine-disrupting chemicals (EDCs), the mechanism(s) by which EDCs exert their harmful effects on humans and their offspring needs careful examination and clarification. Epigenetic modification, including DNA methylation, expression of aberrant microRNA (miRNA), and histone modification, is one mechanism assumed to be a primary pathway leading to the untoward effects of endocrine disruptors. However, it remains unclear whether such epigenetic changes caused by EDCs are truly predicting adverse outcomes. Therefore, it is important to understand the relationship between epigenetic changes and various endocrine endpoints or markers. This paper highlights the possibility that certain chemicals (Cd, As, Pb, bisphenol A, phthalate, polychlorinated biphenyls) reported having ED properties may adversely affect the epigenome. Electronic database sources PubMed, SCOPUS, JSTOR, and the Google Scholar web browser were used to search the literature. The search was based on keywords from existing theories and basic knowledge of endocrine disorders and epigenetic effects, well-known EDCs, and previous search results. Unclear and often conflicting results regarding the effects of EDCs indicate the need for further research to support better risk assessments and management of these chemicals.
... As for tissues other than placenta, a few studies investigated associations between maternal urinary concentrations of DEHP metabolites and DNA methylation in cord blood. A study carried on the CHAMACOS cohort and focusing on ten candidate imprinted genes reported positive associations between MECPP, MEHHP, MEOHP, and ΣDEHP and DNA methylation averaged across seven CpGs of maternally expressed 3 (MEG3) gene (n = 296, Tindula et al. 2018). No effect was reported for the MEG3 expression. ...
Background
Exposure to phthalates during pregnancy may alter DNA methylation in the placenta, a crucial organ for the growth and development of the fetus.
Objectives
We studied associations between urinary concentrations of phthalate biomarkers during pregnancy and placental DNA methylation.
Methods
We measured concentrations of 11 phthalate metabolites in maternal spot urine samples collected between 22 and 29 gestational weeks in 202 pregnant women. We analyzed DNA methylation levels in placental tissue (fetal side) collected at delivery. We first investigated changes in global DNA methylation of repetitive elements Alu and LINE-1. We then performed an adjusted epigenome-wide association study using IlluminaHM450 BeadChips and identified differentially methylated regions (DMRs) associated with phthalate exposure.
Results
Monobenzyl phthalate concentration was inversely associated with placental methylation of Alu repeats. Moreover, all phthalate biomarkers except for monocarboxy-iso-octyl phthalate and mono(2-ethyl-5-hydroxyhexyl) phthalate were associated with at least one DMR. All but three DMRs showed increased DNA methylation with increased phthalate exposure. The largest identified DMR (22 CpGs) was positively associated with monocarboxy-iso-nonyl phthalate and encompassed heat shock proteins (HSPA1A, HSPA1L). The remaining DMRs encompassed transcription factors and nucleotide exchange factors, among other genes.
Conclusions
This is the first description of genome-wide modifications of placental DNA methylation in association with pregnancy exposure to phthalates. Our results suggest epigenetic mechanisms by which exposure to these compounds could affect fetal development. Of interest, four identified DMRs had been previously associated with maternal smoking, which may suggest particular sensitivity of these genomic regions to the effect of environmental contaminants.
... Lastly, studies investigating other EDCs showed gene-specific or global DNAm changes after prenatal exposure to EDCs. For instance, significant associations were found between phthalates and gene-specific DNAm changes in Mexican-American newborns [39] as well as between bisphenol A (BPA) and gene-specific DNAm changes in the Michigan Mother-Infant Pairs birth cohort [40]. Also, prenatal exposure to phthalates revealed significant associations with epigenome-wide DNAm marks in placental tissues [41]. ...
Prenatal exposure to endocrine disrupting chemicals can interfere with development, and has been associated with social-cognitive functioning and adverse health outcomes later in life. Exposure-associated changes of DNA methylation (DNAm) patterns have been suggested as a possible mediator of this relationship. This study investigated whether prenatal low-dose exposure to polychlorinated biphenyls (PCBs) and polychlorinated dibenzo-p-dioxins and dibenzofurans (PCDD/Fs) is associated with altered DNAm patterns across the genome in a Western urban-industrial population. In 142 mother-infant pairs from the Duisburg Birth Cohort Study, PCBs and PCDD/Fs levels were quantified from maternal blood during late pregnancy and associated with DNAm levels in cord blood using the Illumina EPIC beadchip. The epigenome-wide association studies (EWAS) identified 32 significantly differentially methylated positions (DMPs) and eight differentially methylated regions (DMRs) associated with six congeners of PCB and PCDD in females or males (FDRs < 0.05). DMPs and DMRs mapped to genes involved in neurodevelopment, gene regulation, and immune functioning. Weighted gene correlation network analysis (WGCNA) showed 31 co-methylated modules (FDRs < 0.05) associated with one congener of PCDF levels in females. Results of both analytical strategies indicate that prenatal exposure to PCBs and PCDD/Fs is associated with altered DNAm of genes involved in neurodevelopment, gene expression and immune functioning. DNAm and gene expression levels of several of these genes were previously associated with EDC exposure in rodent models. Follow-up studies will clarify whether these epigenetic changes might contribute to the origin for adverse mental and health outcomes.
... Tindula et al. examined the association of prenatal phthalate exposure and imprinted gene DNA methylation profiles in cord blood of newborn children. In newborns of the Center for the Health Assessment of Mothers and Children of Salinas (CHAMACOS) study, prenatal exposure to several DEHP metabolites was positively associated with DNA methylation at the MEG3 DMR [111]. Given that previous birth cohorts only investigated 2-3 imprinted genes, one of the strengths of this study was the evaluation of DNA methylation profiles of multiple imprinted genes in newborns prenatally exposed to phthalates [112,113]. ...
Genomic imprinting is an epigenetic mechanism that results in monoallelic, parent-of-origin-specific expression of a small number of genes. Imprinted genes play a crucial role in mammalian development as their dysregulation result in an increased risk of human diseases. DNA methylation, which undergoes dynamic changes early in development, is one of the epigenetic marks regulating imprinted gene expression patterns during early development. Thus, environmental insults, including endocrine disrupting chemicals during critical periods of fetal development, can alter DNA methylation patterns, leading to inappropriate developmental gene expression and disease risk. Here, we summarize the current literature on the impacts of in utero exposure to endocrine disrupting chemicals on genomic imprinting and metabolism in humans and rodents. We evaluate how early-life environmental exposures are a potential risk factor for adult metabolic diseases. We also introduce our mouse model of phthalate exposure. Finally, we describe the potential of genomic imprinting to serve as an environmental sensor during early development and as a novel biomarker for postnatal health outcomes.
... The impacts of phthalates on human beings vary from gene expression to physiological changes. High molecular weight phthalates exposure is found to cause methylation status of imprinted genes, which could be directly related to androgen response, estrogen response, protein secretion, and spermatogenesis [22,23]. Human epidemiological studies have shown a significant association between phthalates exposures and adverse reproductive outcomes in both women and men, for instance, type II diabetes and insulin resistance, overweight/obesity, allergy, asthma [24]. ...
Phthalates are a series of widely used chemicals that demonstrate to be endocrine disruptors and are detrimental to human health. Phthalates can be found in most products that have contact with plastics during producing, packaging, or delivering. Despite the short half-lives in tissues, chronic exposure to phthalates will adversely influence the endocrine system and functioning of multiple organs, which has negative long-term impacts on the success of pregnancy, child growth and development, and reproductive systems in both young children and adolescents. Several countries have established restrictions and regulations on some types of phthalates; however, we think that more countries should establish constraints or substitute measures for phthalates to reduce health risks. This article aims to summarize the adverse impacts of phthalates on human health, analyze the toxicity mechanism, assess the risks, and finally provide feasible strategies to reduce exposure of the public to phthalates.
... Previous publications suggest that exposure to DEHP and other endocrine disruptors can profoundly alter epigenetic marks in imprinted genes (53,55,56); however, the effect of perinatal DEHP in blood and liver specifically remains unknown. Here, we compared the methylation level of mouse imprinted genes at CpG islands, shores, and shelves, and discovered that the effect of DEHP on imprinted genes is both sex and tissue specific ( Fig. 5A and B). ...
Di(2-ethylhexyl) phthalate (DEHP) is a type of phthalate plasticizer found in a variety of consumer products and poses a public health concern due to its metabolic and endocrine disruption activities. Dysregulation of epigenetic modifications, including DNA methylation, has been shown to be an important mechanism for the pathogenic effects of prenatal exposures, including phthalates. In this study, we used an established mouse model to study the effect of perinatal DEHP exposure on the DNA methylation profile in liver (a primary target tissue of DEHP) and blood (a common surrogate tissue) of both juvenile and adult mice. Despite exposure ceasing at 3 weeks of age (PND21), we identified thousands of sex-specific differential DNA methylation events in 5-month old mice, more than identified at PND21, both in blood and liver. Only a small number of these differentially methylated cytosines (DMCs) overlapped between the time points, or between tissues (i.e. liver and blood), indicating blood may not be an appropriate surrogate tissue to estimate the effects of DEHP exposure on liver DNA methylation. We detected sex-specific DMCs common between 3-week and 5-month samples, pointing to specific DNA methylation alterations that are consistent between weanling and adult mice. In summary, this is the first study to assess the genome-wide DNA methylation profiles in liver and blood at two different aged cohorts in response to perinatal DEHP exposure. Our findings cast light on the implications of using surrogate tissue instead of target tissue in human population-based studies and identify epigenetic biomarkers for DEHP exposure.
... In humans, recent data highlight the significant effect the prenatal exposure to phthalates and their consequence on DNA methylation. Phthalate exposure in utero may affect the methylation pattern status of imprinted genes in newborn children, in particular for the following genes: a first group involved in androgen response MEG3 [39], PA2G4, HMGCR, and XRCC6, all involved in androgen response [40], and a second group of genes involved in early birth delivery, such as Alu and LINE1 [41]. The long-lasting effect of the epigenetic changes due to phthalates may lead to promote tumor progressions [42,43] and infertility [44]. ...
Phthalates, as other endocrine disrupting chemicals (EDCs), may alter the homeostasis and the action of hormones and signaling molecules, causing adverse health outcomes. This is true especially for infants, who are both more exposed and sensitive to their effects. Phthalates are particularly harmful when the exposure occurs during certain critical temporal windows of the development, such as the prenatal and the early postnatal phases. Phthalates may also interfere with the neuroendocrine systems (e.g., thyroid hormone signaling or metabolism), causing disruption of neuronal differentiation and maturation, increasing the risk of behavioral and cognitive disorders (ADHD and autistic behaviors, reduced mental, psychomotor, and IQ development, and emotional problems). Despite more studies being needed to better understand the role of these substances, plenty of evidence suggests the impact of phthalates on the neuroendocrine system development and function. This review aims to update the knowledge on the neuroendocrine consequences of neonatal and perinatal exposure to phthalates.
... Prenatal and early postnatal life represent the most critical windows for development impairment and, as the placental barrier is not completely impermeable to the passage of harmful substances, environmental exposure can affect and permanently reprogram normal physiological responses [19]. Significantly, fetal growth restriction and premature birth have been associated with EDC exposure and with health consequences later in life (increased risk of short stature, metabolic syndrome, high blood pressure, obesity etc.) [20 && , [21][22][23][24]. Moreover, placenta should be considered both as a filter for the passage of EDCs, and an endocrine organ itself [25]. ...
Purpose of review:
Health status is the result of complex interaction between individual factors, general environmental factors and specific factors as nutrition or the presence of chemicals. Aim of this review is to point out the more recent knowledge covering the role of the endocrine disrupting chemical (EDC) on pediatric population wellbeing.
Recent findings:
Prenatal, postnatal life and puberty are the three main temporal windows of susceptibility when EDCs may act. The mechanism is independent from dose or duration of exposition, sex, age or combination of chemicals and may also be transgenerational, affecting both growth and pubertal timing. A window of susceptibility for breast cancer has been detected. Thyroid gland is influenced by environmental chemicals, both in utero and during childhood. Alteration in Thyrotropin stimulating hormone (TSH) levels and neurodevelopmental impairment have been demonstrate. It has been detected a pro-obesogenic action of specific chemicals, impairing also glucose homeostasis during childhood.
Summary:
With a multidisciplinary approach and the use of big data platforms, an attempt has to be made to verify biological variations related to a disease, and how much the risk is influenced by the presence of the endocrine disruptors. This may help the future generation to better interpret uncommunicable diseases.
... They specifically focused on fetal phthalates metabolites' exposure during pregnancy, assessed on maternal urines, and their association with imprinted genes DNA methylation quantified on cord blood samples at term. Interestingly, this investigation demonstrated a significant positive association between phthalate metabolites concentration and the methylation of Maternally Expressed 3 (MEG3) gene, known for its role in early growth, tumorigenesis and metabolic processes [84]. ...
Endocrine-disrupting chemicals (EDCs) are exogenous substances able to mimic or to interfere with the endocrine system, thus altering key biological processes such as organ development, reproduction, immunity, metabolism and behavior. High concentrations of EDCs are found in several everyday products including plastic bottles and food containers and they could be easily absorbed by dietary intake. In recent years, considerable interest has been raised regarding the biological effects of EDCs, particularly Bisphenol A (BPA) and phthalates, on human pregnancy and fetal development. Several evidence obtained on in vitro and animal models as well as by epidemiologic and population studies strongly indicated that endocrine disruptors could negatively impact fetal and placental health by interfering with the embryonic developing epigenome, thus establishing disease paths into adulthood. Moreover, EDCs could cause and/or contribute to the onset of severe gestational conditions as Preeclampsia (PE), Fetal Growth Restriction (FGR) and gestational diabetes in pregnancy, as well as obesity, diabetes and cardiovascular complications in reproductive age. Therefore, despite contrasting data being present in the literature, endocrine disruptors must be considered as a therapeutic target. Future actions aimed at reducing or eliminating EDC exposure during the perinatal period are mandatory to guarantee pregnancy success and preserve fetal and adult health.
... Fecundity appeared to be inconclusive also and warrants further investigations in human reproduction. Prenatal exposure to endocrine disruptors has been found to be associated with various gynecological diseases including breast cancer, eclampsia, and infertility [135] with epigenetic changes hypothesized [172,173], which deserve evaluations for the phthalate investigation. ...
Phthalate esters (PAE) are widely used plasticizers and solvents that are added to many consumer products used in our daily life. PAEs are not chemically bound to the polymer, and therefore are easily released or migrate into the environment including air, drink, foodstuffs, or furniture. Humans can be exposed to PAE through inhalation, ingestion, dermal absorption, or contact with medical devices. Approximately 70% of the oral dose is excreted in urine. Urinary monoesters are the major urinary metabolites of PAEs and commonly used as internal exposure indexes. Infants and children are more prone to expose to certain PAE than adolescents and adults due to their hand-to-mouth behavior. Adverse effects of PAE exposure have been observed in human studies, and verified in animal experiments. Prenatal PAE exposure is related to decreased levels of testosterone (TT), free TT, progesterone, triiodothyronine, and thyroxine in children. PAE exposure is also associated with developing allergic disease and obesity, decreasing intelligence quotient scores, and affecting psychological behaviors and renal function in children. Higher PAE exposure is also associated with endometriosis, leiomyoma, spontaneous abortion, fertility, and breast cancer in women and semen quality in men. More data are necessary for cancers of the breast, endometrial tissue, ovary, and/or prostate to understand the potential risk related to sex hormone sensitive neoplasms. In summary, phthalates exposure was found to be associated with various adverse effects related to altered functions of systems including the endocrine, immune, nervous, and reproduction, particularly at critical development windows during fetal, fast growing, and pubertal stages. Future research is directed to multiple generation approach in humans and/or testing animals, and considerations of psychological parameters (i.e., stress) to provide a further wide observational window for the conclusion.
... Phthalates are a family of phthalic acid diesters that are commonly found in consumer products, resulting in ubiquitous exposure in the USA. Limited but growing evidence in human studies indicate that phthalate exposure is associated with DNA methylation changes of imprinted genes in cord blood and placenta [47]. Animal studies demonstrate associations between phthalates and global and site-specific methylation [48]; however, studies were performed in mixed cell populations, and changes in DNA methylation were usually modest. ...
Type 2 diabetes prevalence is increasing dramatically across the globe, imposing a tremendous toll on individuals and healthcare systems. Reversing these trends requires comprehensive approaches to address both classical and emerging diabetes risk factors. Recently, environmental toxicants acting as endocrine-disrupting chemicals (EDCs) have emerged as novel metabolic disease risk factors. EDCs implicated in diabetes pathogenesis include various inorganic and organic molecules of both natural and synthetic origin, including arsenic, bisphenol A, phthalates, polychlorinated biphenyls and organochlorine pesticides. Indeed, evidence implicates EDC exposures across the lifespan in metabolic dysfunction; moreover, specific developmental windows exhibit enhanced sensitivity to EDC-induced metabolic disruption, with potential impacts across generations. Importantly, differential exposures to diabetogenic EDCs likely also contribute to racial/ethnic and economic disparities. Despite these emerging links, clinical practice guidelines fail to address this underappreciated diabetes risk factor. Comprehensive approaches to stem the tide of diabetes must include efforts to address its environmental drivers.
... Previous studies have demonstrated that prenatal exposure to maternal stress and anxiety was negatively associated with methylation level in the DMR of IGF2 [34,35]. On contrary, in Mexican-American newborn children, prenatal phthalate and estradiol exposures increased methylation of the IGF2 DMR [36,37]. Nutrition supply during pregnancy also influences DNA methylation in the IGF2 gene. ...
Objective:
To evaluate the association of early-life exposure to the Chinese Great Famine (1959-1961) with DNA methylation in IGF2 and its subsequent influence on blood lipid levels in late adulthood among participants of the Genomic Research of the Chinese Famine (GRECF) study.
Methods:
The GRECF study recruited 790 participants born between 1956 and 1964 from 2 neighbor provinces, Anhui and Jiangxi, in China through a multistage, clustered, random sampling. The current study included a random sample of 188 GRECF participants. IGF2 differential methylation region (DMR) is an intragenic DMR located upstream of the imprinted promoters of IGF2 exon 3. DNA methylation were quantified at 8 cytosine-phosphate-guanine dinucleotides (CpG) sites at the IGF2 DMR (chr11p15.5) using the Sequenom EpiTYPER method and the MassARRAY system. Multivariate linear regressions were used to evaluate pairwise associations among famine severity, DNA methylation in the IGF2 gene, and lipid levels. We controlled for age and sex in the base model and additionally controlled for education, smoking, and drinking status in the fully adjusted model. Mediation analysis was applied to assess the mediation effect of DNA methylation at the IGF2 gene on the association between early-life exposure to severe famine and adult lipid levels.
Results:
Exposure to severe famine was associated with elevated methylation at CpG1 (chr11: 2126041, build 36) of the IGF2 DMR (β = 0.07; P = 0.0008) and total cholesterol (β = 0.72; P = 1.09 × 10-7). After adjustment for age and sex, each unit increase in methylation of the CpG1 site was associated with 1.09-unit increase in total cholesterol (P = 0.03). After further adjustment for all covariates, these associations were still significant (Pfamine-CpG1 = 0.002, Pfamine-total cholesterol = 1.28 × 10-6, and PCpG1-total cholesterol = 0.05).
Conclusion:
Increased methylation level in the IGF2 gene was associated with early-life exposure to severe famine, and this change was also positively associated with total cholesterol in late adulthood.
... DNA methylation patterns are retained following somatic cell division [10]. In the Center for the Health Assessment of Mothers and Children of Salinas (CHAMACOS) cohort and as part of the Pregnancy and Childhood Epigenetics consortium, we have explored the relationship of infant DNA methylation with early life environmental exposure of mothers to chemicals during pregnancy, including endocrine disrupting chemicals [11][12][13] and smoking [14]. Since the epigenome undergoes remodeling and rapid cell division in the prenatal period [8], this life stage is considered particularly sensitive to environmental insults and can have implications for disease trajectories [15]. ...
Lipids play a role in many biological functions and the newly emerging field of lipidomics aims to characterize the varying classes of lipid molecules present in biological specimens. Animal models have shown associations between maternal dietary supplementation with fatty acids during pregnancy and epigenetic changes in their offspring, demonstrating a mechanism through which prenatal environment can affect outcomes in children; however, data on maternal lipid metabolite levels during pregnancy and newborn DNA methylation in humans are sparse. In this study, we assessed the relationship of maternal lipid metabolites measured in the blood from pregnant women with newborn DNA methylation profiles in the Center for the Health Assessment of Mothers and Children of Salinas cohort. Targeted metabolomics was performed by selected reaction monitoring liquid chromatography and triple quadrupole mass spectrometry to measure 92 metabolites in plasma samples of pregnant women at ∼26 weeks gestation. DNA methylation was assessed using the Infinium HumanMethylation 450K BeadChip adjusting for cord blood cell composition. We uncovered numerous false discovery rate significant associations between maternal metabolite levels, particularly phospholipid and lysolipid metabolites, and newborn methylation. The majority of the observed relationships were negative, suggesting that higher lipid metabolites during pregnancy are associated with lower methylation levels at genes related to fetal development. These results further elucidate the complex relationship between early life exposures, maternal lipid metabolites, and infant epigenetic status.
... Our pathway analysis also suggests a novel relationship with nucleic acid metabolism, an association in line with our previous report linking phthalate exposure and differential DNA methylation in the same cohort (12,73,74). The correlation of several nucleotides and nucleosides in plasma and urine with higher exposures suggests that phthalates may influence degradation, salvage, and elimination pathways of nucleic acid metabolism. ...
Phthalates are known endocrine disruptors and found in almost all people with several associated adverse health outcomes reported in humans and animal models. Limited data are available on the relationship between exposure to endocrine disrupting chemicals and the human metabolome. We examined the relationship of metabolomic profiles in plasma and urine of 115 pregnant women with eleven urine phthalate metabolites measured at 26 weeks of gestation to identify potential biomarkers and relevant pathways. Targeted metabolomics was performed by selected reaction monitoring liquid chromatography and triple quadrupole mass spectrometry to measure 415 metabolites in plasma and 151 metabolites in urine samples. We have chosen metabolites with the best defined peaks for more detailed analysis (138 in plasma and 40 in urine). Relationship between urine phthalate metabolites and concurrent metabolomic markers in plasma and urine suggested potential involvement of diverse pathways including lipid, steroid, and nucleic acid metabolism and enhanced inflammatory response. Most of the correlations were positive for both urine and plasma, and further confirmed by regression and PCA analysis. However, after the FDR adjustment for multiple comparisons, only 9 urine associations remained statistically significant (q-values 0.0001–0.0451), including Nicotinamide mononucleotide, Cysteine T2, Cystine, and L-Aspartic acid. Additionally, we found negative associations of maternal pre-pregnancy body mass index (BMI) with more than 20 metabolomic markers related to lipid and amino-acid metabolism and inflammation pathways in plasma (p = 0.01–0.0004), while Mevalonic acid was positively associated (p = 0.009). Nicotinic acid, the only significant metabolite in urine, had a positive association with maternal BMI (p = 0.002). In summary, when evaluated in the context of metabolic pathways, the findings suggest enhanced lipid biogenesis, inflammation and altered nucleic acid metabolism in association with higher phthalate levels. These results provide new insights into the relationship between phthalates, common in most human populations, and metabolomics, a novel approach to exposure and health biomonitoring.
Humans are ubiquitously exposed to environmental endocrine disrupting chemicals such as phthalates. Phthalates can migrate out of products and enter the human body through ingestion, inhalation, or dermal application, can have potential estrogenic/antiestrogenic and/or androgenic/antiandrogenic activity, and are involved in many diseases. As a female reproductive organ that is regulated by hormones such as estrogen, progesterone and androgen, the uterus can develop several disorders such as leiomyoma, endometriosis and abnormal bleeding. In this review, we summarize the hormone-like activities of phthalates, in vitro studies of endometrial cells exposed to phthalates, epigenetic modifications in the uterus induced by phthalate exposure, and associations between phthalate exposure and uterine disorders such as leiomyoma and endometriosis. Moreover, we also discuss the current research gaps in understanding the relationship between phthalate exposure and uterine disorders.
This overview discusses the role of imprinting in the development of an organism, and how exposure to environmental chemicals during fetal development leads to the physiological and biochemical changes that can have adverse lifelong effects on the health of the offspring. There has been a recent upsurge in the use of chemical products in everyday life. These chemicals include industrial byproducts, pesticides, dietary supplements, and pharmaceutical products. They mimic the natural estrogens and bind to estradiol receptors. Consequently, they reduce the number of receptors available for ligand binding. This leads to a faulty signaling in the neuroendocrine system during the critical developmental process of ‘imprinting’. Imprinting causes structural and organizational differentiation in male and female reproductive organs, sexual behavior, bone mineral density, and the metabolism of exogenous and endogenous chemical substances. Several studies conducted on animal models and epidemiological studies provide profound evidence that altered imprinting causes various developmental and reproductive abnormalities and other diseases in humans. Altered metabolism can be measured by various endpoints such as the profile of cytochrome P-450 enzymes (CYP450’s), xenobiotic metabolite levels, and DNA adducts. The importance of imprinting in the potentiation or attenuation of toxic chemicals is discussed.
Exposure to phthalates has been shown to impede the human endocrine system, resulting in deleterious effects on pregnant women and their children. Phthalates modify DNA methylation patterns in infant cord blood. We examined the association between prenatal phthalate exposure and DNA methylation patterns in cord blood in a Korean birth cohort. Phthalate levels were measured in 274 maternal urine samples obtained during late pregnancy and 102 neonatal urine samples obtained at birth, and DNA methylation levels were measured in cord blood samples. For each infant in the cohort, associations between CpG methylation and both maternal and neonate phthalate levels were analyzed using linear mixed models. The results were combined with those from a meta-analysis of the levels of phthalates in maternal and neonatal urine samples, which were also analyzed for MEOHP, MEHHP, MnBP, and DEHP. This meta-analysis revealed significant associations between the methylation levels of CpG sites near the CHN2 and CUL3 genes, which were also associated with MEOHP and MnBP in neonatal urine. When the data were stratified by the sex of the infant, MnBP concentration was found to be associated with one CpG site near the OR2A2 and MEGF11 genes in female infants. In contrast, the concentrations of the three maternal phthalates showed no significant association with CpG site methylation. Furthermore, the data identified distinct differentially methylated regions in maternal and neonatal urine samples following exposure to phthalates. The CpGs with methylation levels that were positively associated with phthalate levels (particularly MEOHP and MnBP) were found to be enriched genes and related pathways. These results indicate that prenatal phthalate exposure is significantly associated with DNA methylation at multiple CpG sites. These alterations in DNA methylation may serve as biomarkers of maternal exposure to phthalates in infants and are potential candidates for investigating the mechanisms by which phthalates impact maternal and neonatal health.
The indiscriminate use of phthalate-containing products in daily life can adversely affect pregnant women and their children. Phthalate can modify DNA methylation in the cord blood of infants. Therefore, we examined the association between prenatal phthalate exposure and cord blood DNA methylation in a Korean birth cohort. Phthalate levels in maternal blood during late pregnancy and cord blood were measured and DNA methylation of cord blood was measured using the Illumina HumanMethylationEPIC BeadChip kit. The association between CpG methylation and phthalate levels was analyzed using the ‘limma’ package in R, adjusting for infant sex, maternal body mass index, current maternal smoking status, and estimated leukocyte composition. We used data from 274 samples for estimating mono-(2-ethyl-5-oxohexyl) phthalate (MEOHP) and mono-(2-ethyl-5-hydroxyhxyl) phthalate (MEHHP) levels and 273 samples for estimating mono-n-butyl phthalate (MnBP) levels to determine maternal phthalate concentrations during late pregnancy. Additionally, 102 samples were analyzed for all three types of phthalates in the cord blood. The meta-analysis revealed significant associations between the CpG sites near the CHN2 and CUL3 genes and cord blood MEOHP and MnBP concentrations, respectively. However, the three maternal phthalate concentrations during late pregnancy showed no significant association with CpG sites. In conclusion, prenatal phthalate exposure is significantly associated with DNA methylation at several CpG sites.
Purpose of Review
There is interest in evaluating the developmental origins of health and disease (DOHaD) which emphasizes the role of prenatal and early-life environments on non-communicable health outcomes throughout the life course. The ability to rigorously assess and identify early-life risk factors for later health outcomes, including those with childhood onset, in large population samples is often limited due to measurement challenges such as impractical costs associated with prospective studies with a long follow-up duration, short half-lives for some environmental toxicants, and lack of biomarkers that capture inter-individual differences in biologic response to external environments.
Recent Findings
Epigenomic patterns, and DNA methylation in particular, have emerged as a potential objective biomarker to address some of these study design and exposure measurement challenges. In this article, we summarize the literature to date on epigenetic changes associated with specific prenatal and early-life exposure domains as well as exposure mixtures in human observational studies and their biomarker potential. Additionally, we highlight evidence for other types of epigenetic patterns to serve as exposure biomarkers.
Summary
Evidence strongly supports epigenomic biomarkers of exposure that are detectable across the lifespan and across a range of exposure domains. Current and future areas of research in this field seek to expand these lines of evidence to other environmental exposures, to determine their specificity, and to develop predictive algorithms and methylation scores that can be used to evaluate early-life risk factors for health outcomes across the life span.
Increasing evidence has associated the exposure of endocrine-disrupting chemicals (EDCs) with the cardiovascular (CV) system. This exposure is particularly problematic in a sensitive window of development, pregnancy. Pregnancy exposome can affect the overall health of the pregnancy by dramatic changes in vascular physiology and endocrine activity, increasing maternal susceptibility. Moreover, fetoplacental vascular function is generally altered, increasing the risk of developing pregnancy complications (including cardiovascular diseases, CVD) and predisposing the foetus to adverse health risks later in life. Thus, our review summarizes the existing literature on exposures to EDCs during pregnancy and adverse maternal health outcomes, focusing on the human placenta, vein, and umbilical artery associated with pregnancy complications. The purpose of this review is to highlight the role of fetoplacental vasculature as a model for the study of human cardiovascular endocrine disruption. Therefore, we emphasize that the placenta, together with the umbilical arteries and veins, allows a better characterization of the pregnant woman's exposome. Consequently, it contributes to the protection of the mother and foetus against CV disorders in life.
Purpose of Review
Endocrine-disrupting chemical (EDC) exposure during pregnancy is linked to adverse maternal and child health outcomes that are racially/ethnically disparate. Personal care products (PCP) are one source of EDCs where differences in racial/ethnic patterns of use exist. We assessed the literature for racial/ethnic disparities in pregnancy and prenatal PCP chemical exposures.
Recent Findings
Only 3 studies explicitly examined racial/ethnic disparities in pregnancy and prenatal exposure to PCP-associated EDCs. Fifty-three articles from 12 cohorts presented EDC concentrations stratified by race/ethnicity or among homogenous US minority populations. Studies reported on phthalates and phenols. Higher phthalate metabolites and paraben concentrations were observed for pregnant non-Hispanic Black and Hispanic women. Higher concentrations of benzophenone-3 were observed in non-Hispanic White women; results were inconsistent for triclosan.
Summary
This review highlights need for future research examining pregnancy and prenatal PCP-associated EDCs disparities to understand and reduce racial/ethnic disparities in maternal and child health.
Ovary plays an important role in the female reproductive system. The maintenance and regulation of ovarian function are affected by various physical and chemical factors. With the development of industrialization, environmental pollutants have caused great harm to public health. Phthalates, as a class of endocrine-disrupting chemicals (EDCs), are synthesized and used in large quantities as plasticizers due to their chemical properties. They are easily released into environment because of their noncovalent interactions with substances, causing human exposure and possibly impairing ovary. In recent years, more and more attention has been paid to the role of epigenetics in the occurrence and development of diseases. And it is urgent to study the role of methylation, gene imprinting, miRNA, and other epigenetic mechanisms in reproductive toxicology.
Prenatal exposure to phthalates negatively affects the offspring's health. In particular, epigenetic alterations, such as DNA methylation, may connect phthalate exposure with health outcomes. Here, we evaluated the association of di-2-ethylhexyl phthalate (DEHP) exposure in utero with cord blood epigenome-wide DNA methylation in 203 mother-child pairs enrolled in the Hokkaido Study on Environment and Children's Health, using the Illumina HumanMethylation450 BeadChip. Epigenome-wide association analysis demonstrated the predominant positive associations between the levels of the primary metabolite of DEHP, mono(2-ethylhexyl) phthalate (MEHP), in maternal blood and DNA methylation levels in cord blood. The genes annotated to the CpGs positively associated with MEHP levels were enriched for pathways related to metabolism, the endocrine system, and signal transduction. Among them, methylation levels of CpGs involved in metabolism were inversely associated with the offspring's ponderal index (PI). Further, clustering and mediation analyses suggested that multiple increased methylation changes may jointly mediate the association of DEHP exposure in utero with the offspring's PI at birth. Although further studies are required to assess the impact of these changes, this study suggests that differential DNA methylation may link phthalate exposure in utero to fetal growth and further imply that DNA methylation has predictive value for the offspring's obesity.
DNA methylation (DNAm) plays a significant role in deleterious health effects inflicted by fine particulate matter (PM2.5) on the human body. Recent, studies have reported that DNAm of imprinted control regions (ICRs) in imprinted genes may be a sensitive biomarker of environmental exposure. Less is known about specific biomarkers of imprinted genes after PM2.5 exposure. The relationship between PM2.5 and its chemical constituents and DNAm of ICRs in imprinted genes after short-term exposure was investigated to determine specific human biomarkers of its adverse health effects. A panel study was carried out in healthy young people in Guangzhou, China. Mixed-effects models were used to evaluate the influence of PM2.5 and its constituent exposure on DNAm while controlling for potential confounders. There was no significant correlation between DNAm and personal PM2.5 exposure mass. DNAm changes in eight ICRs (L3MBTL1, NNAT, PEG10, GNAS Ex1A, MCTS2, SNURF/SNRPN, IGF2R, and RB1) and a non-imprinted gene (CYP1B1) were significantly associated with PM2.5 constituents. Compared to non-imprinted genes, imprinted gene methylation was more susceptible to interference with PM2.5 constituent exposure. Among those genes, L3MBTL1 was the most sensitive to personal PM2.5 constituent exposure. Moreover, transition metals derived from traffic sources (Cd, Fe, Mn, and Ni) significantly influenced DNAm of the imprinted genes, suggesting the importance of more targeted measures to reduce toxic constituents. Bioinformatics analysis indicated that imprinted genes (RB1) may be correlated with pathways and diseases (non-small cell lung cancer, glioma, and bladder cancer). The present study suggests that screening the imprinted gene for DNAm can be used as a sensitive biomarker of PM2.5 exposure. The results will provide data for prevention of PM2.5 exposure and a novel perspective on potential mechanisms on an epigenetic level.
Background:
It has been estimated that a substantial portion of chronic and noncommunicable diseases can be caused or exacerbated by exposure to environmental chemicals. Multiple lines of evidence indicate that early life exposure to environmental chemicals at relatively low concentrations could have lasting effects on individual and population health. Although the potential adverse effects of environmental chemicals are known to the scientific community, regulatory agencies, and the public, little is known about the mechanistic basis by which these chemicals can induce long-term or transgenerational effects. To address this question, epigenetic mechanisms have emerged as the potential link between genetic and environmental factors of health and disease.
Objectives:
We present an overview of epigenetic regulation and a summary of reported evidence of environmental toxicants as epigenetic disruptors. We also discuss the advantages and challenges of using epigenetic biomarkers as an indicator of toxicant exposure, using measures that can be taken to improve risk assessment, and our perspectives on the future role of epigenetics in toxicology.
Discussion:
Until recently, efforts to apply epigenomic data in toxicology and risk assessment were restricted by an incomplete understanding of epigenomic variability across tissue types and populations. This is poised to change with the development of new tools and concerted efforts by researchers across disciplines that have led to a better understanding of epigenetic mechanisms and comprehensive maps of epigenomic variation. With the foundations now in place, we foresee that unprecedented advancements will take place in the field in the coming years. https://doi.org/10.1289/EHP6104.
Background:
Characterization of the epigenome is a primary interest for children's environmental health researchers studying the environmental influences on human populations, particularly those studying the role of pregnancy and early-life exposures on later-in-life health outcomes.
Objectives:
Our objective was to consider the state of the science in environmental epigenetics research and to focus on DNA methylation and the collective observations of many studies being conducted within the Children's Environmental Health and Disease Prevention Research Centers, as they relate to the Developmental Origins of Health and Disease (DOHaD) hypothesis.
Methods:
We address the current laboratory and statistical tools available for epigenetic analyses, discuss methods for validation and interpretation of findings, particularly when magnitudes of effect are small, question the functional relevance of findings, and discuss the future for environmental epigenetics research.
Discussion:
A common finding in environmental epigenetic studies is the small-magnitude epigenetic effect sizes that result from such exposures. Although it is reasonable and necessary that we question the relevance of such small effects, we present examples in which small effects persist and have been replicated across populations and across time. We encourage a critical discourse on the interpretation of such small changes and further research on their functional relevance for children's health.
Conclusion:
The dynamic nature of the epigenome will require an emphasis on future longitudinal studies in which the epigenome is profiled over time, over changing environmental exposures, and over generations to better understand the multiple ways in which the epigenome may respond to environmental stimuli.
Strong evidence implicates maternal phthalate exposure during pregnancy in contributing to adverse birth outcomes. Recent research suggests these effects might be mediated through the improper regulation of DNA methylation in offspring tissue. In this study, we examined associations between prenatal phthalate exposure and DNA methylation in human placenta. We recruited 181 mother-newborn pairs (80 fetal growth restriction newborns, 101 normal newborns) in Wenzhou, China and measured third trimester urinary phthalate metabolite concentrations and placental DNA methylation levels of IGF2 and AHRR. We found urinary concentrations of mono (2-ethyl-5- hydroxyhexyl) phthalate (MEHHP), and mono (2-ethyl-5-oxohexyl) phthalate (MEOHP) were significantly inversely associated with placental IGF2 DNA methylation. The associations were much more evident in fetal growth restriction (FGR) newborns than those in normal newborns. These findings suggest that changes in placental DNA methylation might be part of the underlying biological pathway between prenatal phthalate exposure and adverse fetal growth.
Epigenetic perturbations induced by environmental exposures at susceptible lifestages contribute to disease development. Even so, the influence of early life and ongoing exposures on the adolescent epigenome is rarely examined. We examined the association of exposure biomarkers for lead (Pb), bisphenol A (BPA), and nine phthalates metabolites with blood leukocyte DNA methylation at LINE-1 repetitive elements and environmentally responsive genes (IGF2, H19, and HSD11B2) in peri-adolescents. Participants (n = 247) from the Early Life Exposures in Mexico to Environmental Toxicants (ELEMENT) birth cohorts were followed-up once between the ages of 8 and 14 years, and concurrent exposures were measured in biospecimen collected at that time (blood Pb, urinary BPA, and phthalate metabolites). Prenatal and childhood exposures to Pb were previously approximated using maternal and child samples. BPA and phthalate metabolites were measured in third trimester maternal urine samples. Significant associations (P < 0.05) were observed between DNA methylation and exposure biomarkers that were gene and biomarker specific. For example, Pb was only associated with LINE-1 hypomethylation during pregnancy (P = 0.04), while early childhood Pb was instead associated with H19 hypermethylation (P = 0.04). Concurrent urinary mono (2-ethylhexyl) phthalate (MEHP) was associated with HSD11B2 hypermethylation (P = 0.005). Sex-specific associations, particularly among males, were also observed. In addition to single exposure models, principal component analysis was employed to examine exposure mixtures. This method largely corroborated the findings of the single exposure models. This study along with others in the field suggests that environment-epigenetic relationships vary by chemical, exposure timing, and sex.
Background
Epigenetic reprogramming in mammalian gametes resets methylation marks that regulate monoallelic expression of imprinted genes. In males, this involves erasure of the maternal methylation marks and establishment of paternal-specific methylation to appropriately guide normal development. The degree to which exogenous factors influence the fidelity of methylation reprogramming is unknown. We previously found an association between paternal obesity and altered DNA methylation in umbilical cord blood, suggesting that the father’s endocrine, nutritional, or lifestyle status could potentiate intergenerational heritable epigenetic abnormalities. In these analyses, we examine the relationship between male overweight/obesity and DNA methylation status of imprinted gene regulatory regions in the gametes. Methods
Linear regression models were used to compare sperm DNA methylation percentages, quantified by bisulfite pyrosequencing, at 12 differentially methylated regions (DMRs) from 23 overweight/obese and 44 normal weight men. Our study population included 69 volunteers from The Influence of the Environment on Gametic Epigenetic Reprogramming (TIEGER) study, based in NC, USA. ResultsAfter adjusting for age and fertility patient status, semen from overweight or obese men had significantly lower methylation percentages at the MEG3 (β = −1.99; SE = 0.84; p = 0.02), NDN (β = −1.10; SE = 0.47; p = 0.02), SNRPN (β = −0.65; SE = 0.27; p = 0.02), and SGCE/PEG10 (β = −2.5; SE = 1.01; p = 0.01) DMRs. Our data further suggest a slight increase in DNA methylation at the MEG3-IG DMR (β = +1.22; SE = 0.59; p = 0.04) and H19 DMR (β = +1.37; SE = 0.62; p = 0.03) in sperm of overweight/obese men. Conclusions
Our data support that male overweight/obesity status is traceable in the sperm epigenome. Further research is needed to understand the effect of such changes and the point of origin of DNA methylation differences between lean and overweight/obese men. Together with our earlier reports on paternal obesity and epigenetic shifts in the offspring, our studies set the groundwork for future studies investigating male gametic methylation aberrations due to paternal lifestyle factors such as obesity.
Compelling evidence suggests that maternal mental health in pregnancy can influence fetal development. The imprinted genes, insulin-like growth factor 2 (IGF2) and H19, are involved in fetal growth and each is regulated by DNA methylation. This study aimed to determine the association between maternal mental well-being during pregnancy and differentially methylated regions (DMRs) of IGF2 (DMR0) and the IGF2/H19 imprinting control region (ICR) in newborn offspring. Maternal depression, anxiety and perceived stress were assessed at 28 weeks of pregnancy in the Barwon Infant Study (n=576). DNA methylation was measured in purified cord blood mononuclear cells using the Sequenom MassArray Platform. Maternal anxiety was associated with a decrease in average ICR methylation (Δ=-2.23%; 95% CI=-3.68 to -0.77%), and across all six of the individual CpG units in anxious compared with non-anxious groups. Birth weight and sex modified the association between prenatal anxiety and infant methylation. When stratified into lower (⩽3530 g) and higher (>3530 g) birth weight groups using the median birth weight, there was a stronger association between anxiety and ICR methylation in the lower birth weight group (Δ=-3.89%; 95% CI=-6.06 to -1.72%), with no association in the higher birth weight group. When stratified by infant sex, there was a stronger association in female infants (Δ=-3.70%; 95% CI=-5.90 to -1.51%) and no association in males. All the linear regression models were adjusted for maternal age, smoking and folate intake. These findings show that maternal anxiety in pregnancy is associated with decreased IGF2/H19 ICR DNA methylation in progeny at birth, particularly in female, low birth weight neonates. ICR methylation may help link poor maternal mental health and adverse birth outcomes, but further investigation is needed.
Epigenome-wide association studies of disease widely use DNA methylation measured in blood as a surrogate tissue. Cell proportions can vary between people and confound associations of exposure or outcome. An adequate reference panel for estimating cell proportions from adult whole blood for DNA methylation studies is available, but an analogous cord blood cell reference panel is not yet available. Cord blood has unique cell types and the epigenetic signatures of standard cell types may not be consistent throughout the life course. Using magnetic bead sorting, we isolated cord blood cell types (nucleated red blood cells, granulocytes, monocytes, natural killer cells, B cells, CD4(+)T cells, and CD8(+)T cells) from 17 live births at Johns Hopkins Hospital. We confirmed enrichment of the cell types using fluorescence assisted cell sorting and ran DNA from the separated cell types on the Illumina Infinium HumanMethylation450 BeadChip array. After filtering, the final analysis was on 104 samples at 429,794 probes. We compared cell type specific signatures in cord to each other and methylation at 49.2% of CpG sites on the array differed by cell type (F-test P<10(-8)). Differences between nucleated red blood cells and the remainder of the cell types were most pronounced (36.9% of CpG sites at P<10(-8)) and 99.5% of these sites were hypomethylated relative to the other cell types. We also compared the mean-centered sorted cord profiles to the available adult reference panel and observed high correlation between the overlapping cell types for granulocytes and monocytes (both r=0.74), and poor correlation for CD8(+)T cells and NK cells (both r=0.08). We further provide an algorithm for estimating cell proportions in cord blood using the newly developed cord reference panel, which estimates biologically plausible cell proportions in whole cord blood samples.
People are exposed to phthalates through their wide use as plasticizers and in personal care products. Many phthalates are endocrine disruptors and have been associated with adverse health outcomes. However, knowledge gaps exist in understanding the molecular mechanisms associated with the effects of exposure in early and late pregnancy. In this study, we examined the relationship of eleven urinary phthalate metabolites with isoprostane, an established marker of oxidative stress, among pregnant Mexican-American women from an agricultural cohort. Isoprostane levels were on average 20% higher at 26 weeks than at 13 weeks of pregnancy. Urinary phthalate metabolite concentrations suggested relatively consistent phthalate exposures over pregnancy. The relationship between phthalate metabolite concentrations and isoprostane levels was significant for the sum of di-2-ethylhexyl phthalate and the sum of high molecular weight metabolites with the exception of monobenzyl phthalate, which was not associated with oxidative stress at either time point. In contrast, low molecular weight metabolite concentrations were not associated with isoprostane at 13 weeks, but this relationship became stronger later in pregnancy (p-value = 0.009 for the sum of low molecular weight metabolites). Our findings suggest that prenatal exposure to phthalates may influence oxidative stress, which is consistent with their relationship with obesity and other adverse health outcomes.
Prenatal exposure to lead (Pb) is known to decrease fetal growth; but its effects on postnatal growth and mechanistic insights linking Pb to growth are not clearly defined. Genomically imprinted genes are powerful regulators of growth and energy utilization, and may be particularly vulnerable to environmental Pb exposure. Because imprinting is established early and maintained via DNA methylation, we hypothesized that prenatal Pb exposure alters DNA methylation of imprinted genes resulting in lower birth weight and rapid growth. Pb was measured by inductively coupled plasma mass spectrometry (ICP-MS) in peripheral blood of 321 women of the Newborn Epigenetic STudy (NEST) obtained at gestation ∼12 weeks. Linear and logistic regression models were used to evaluate associations between maternal Pb levels, methylation of differentially methylated regions (DMRs) regulating H19, MEG3, PEG3, and PLAGL1, measured by pyrosequencing, birth weight, and weight-for-height z score gains between birth and age 1 year, ages 1–2 years, and 2–3 years. Children born to women with Pb levels in the upper tertile had higher methylation of the regulatory region of the MEG3 DMR imprinted domain (β = 1.57, SE = 0.82, P = 0.06). Pb levels were also associated with lower birth weight (β = −0.41, SE = 0.15, P = 0.01) and rapid gains in adiposity (OR = 12.32, 95% CI = 1.25–121.30, P = 0.03) by age 2–3 years. These data provide early human evidence for Pb associations with hypermethylation at the MEG3 DMR regulatory region and rapid adiposity gain – a risk factor for childhood obesity and cardiometabolic diseases in adulthood.
A biological mechanism by which exposure to environmental contaminants results in gene-specific CpG methylation patterning is currently unknown. We hypothesize that gene-specific CpG methylation is related to environmentally perturbed transcription factor occupancy. To test this hypothesis, a database of 396 genes with altered CpG methylation either in cord blood leukocytes or placental tissue was compiled from 14 studies representing assessments of six environmental contaminants. Subsequently, an in silico approach was used to identify transcription factor binding sites enriched among the genes with altered CpG methylation in relationship to the suite of environmental contaminants. For each study, the sequences of the promoter regions (representing −1000 to +500 bp from the transcription start site) of all genes with altered CpG methylation were analyzed for enrichment of transcription factor binding sites. Binding sites for a total of 56 unique transcription factors were identified to be enriched within the promoter regions of the genes. Binding sites for the Kidney-Enriched Krupple-like Factor 15, a known responder to endogenous stress, were enriched (P < 0.001–0.041) among the genes with altered CpG methylation associated for five of the six environmental contaminants. These data support the transcription factor occupancy theory as a potential mechanism underlying environmentally-induced gene-specific CpG methylation.
Background:
Bisphenol A (BPA) and phthalates are ubiquitous non-persistent endocrine disrupting chemicals whose relation with infant birth size is not clearly understood.
Methods:
We examined associations between maternal and paternal preconception urinary concentrations of total BPA and 14 phthalate metabolites and birth size for 233 infants. Multiple linear regression models were used to estimate parental quartiles of BPA and phthalates in relation to birth weight, length, head circumference, and ponderal index with separate models run for each parent adjusting for age, smoking, body mass index, education, alcohol, parity, and creatinine. Models also included an interaction term for each chemical and infant sex and were further adjusted to include the other partner's chemical concentrations.
Results:
In maternal models adjusted for partner's exposure and covariates, reductions in birth weight (range: 178-215 g; p < 0.05) were observed for the 2(nd) quartile of maternal monomethyl phthalate, mono-[(2-carboxymethyl) hexyl] phthalate and mono-n-octyl phthalate when compared with the 1(st) quartiles. The 3(rd) quartile of monoethylhexyl phthalate (mEHP) was also associated with a 200.16 g (95 % CI: -386.90, -13.42) reduction. Similar reductions in birth weight were observed for the 2(nd) quartile of paternal mEHP (β = -191.93 g; 95 % CI: -381.61, -2.25). Additionally, select maternal urinary metabolites were associated with decreased head circumference, birth length and gestational age. However, paternal concentrations were generally associated with increased birth length and gestational age.
Conclusions:
We observed some suggestion that preconception maternal and paternal urinary concentration of BPA and specific phthalate metabolites may be associated with smaller birth size and increased gestational age, though the findings appeared to be parent and chemical specific.
Cadmium (Cd) is a ubiquitous and environmentally persistent toxic metal that has been implicated in neurotoxicity, carcinogenesis and obesity and essential metals including zinc (Zn) and iron (Fe) may alter these outcomes. However mechanisms underlying these relationships remain limited.
We examined whether maternal Cd levels during early pregnancy were associated with offspring DNA methylation at regulatory sequences of genomically imprinted genes and weight at birth, and whether Fe and Zn altered these associations. Cd, Fe and Zn were measured in maternal blood of 319 women ≤12 weeks gestation. Offspring umbilical cord blood leukocyte DNA methylation at regulatory differentially methylated regions (DMRs) of 8 imprinted genes was measured using bisulfite pyrosequencing. Regression models were used to examine the relationships among Cd, Fe, Zn, and DMR methylation and birth weight.
Elevated maternal blood Cd levels were associated with lower birth weight (p = 0.03). Higher maternal blood Cd levels were also associated with lower offspring methylation at the PEG3 DMR in females (β = 0.55, se = 0.17, p = 0.05), and at the MEG3 DMR in males (β = 0.72, se = 0.3, p = 0.08), however the latter association was not statistically significant. Associations between Cd and PEG3 and PLAGL1 DNA methylation were stronger in infants born to women with low concentrations of Fe (p < 0.05).
Our data suggest the association between pre-natal Cd and offspring DNA methylation at regulatory sequences of imprinted genes may be sex- and gene-specific. Essential metals such as Zn may mitigate DNA methylation response to Cd exposure. Larger studies are required.
The importance of imprinted genes in regulating feto-placental development has been long established. However, a comprehensive assessment of the role of placental imprinted gene expression on fetal growth has yet to be conducted. In this study, we examined the association between the placental expression of 108 established and putative imprinted genes and birth weight in 677 term pregnancies, oversampled for small for gestational age (SGA) and large for gestational age (LGA) infants. Using adjusted multinomial regression analyses, a two-fold increase in the expression of 9 imprinted genes were positively associated with LGA status (BLCAP (Odds ratio=3.78, [95% Confidence interval: 1.83, 7.82]), DLK1 (1.63, [1.27, 2.09]), H19 (2.79, [1.77, 4.42]), IGF2 (1.43, [1.31, 2.40]), MEG3 (1.42, [1.19, 1.71]), MEST (4.78, [2.64, 8.65]), NNAT (1.40, [1.05, 1.86]), NDN (2.52, [1.72, 3.68], PLAGL1 (1.85, [1.40, 2.44])). For SGA status, a two-fold increase of MEST expression was associated with decreased risk (0.31, [0.17, 0.58]) while a two-fold increase of NNAT expression was associated with increased risk (1.52, [1.1, 2.1]). Following a factor analysis, all genes significantly associated with SGA or LGA status loaded onto 2 of the 8 gene-sets underlying the variability in the dataset. Our comprehensive placental profiling of imprinted genes in a large birth cohort supports the importance of these genes for fetal growth. Given that abnormal birth weight is implicated in numerous diseases and developmental abnormalities, the expression pattern of placental imprinted genes has the potential to be developed as a novel biomarker for postnatal health outcomes.
Lead exposure during early development causes neurodevelopmental disorders by unknown mechanisms. Epidemiologic studies have focused recently on determining associations between lead exposure and global DNA methylation; however, such approaches preclude the identification of loci that may alter human disease risk.
The objective of this study was to determine if maternal, postnatal and early childhood lead exposure alter the differentially methylated regions (DMRs) that control the monoallelic expression of imprinted genes involved in metabolism, growth and development.
Questionnaire data and serial blood lead levels were obtained from 105 participants (64 females, 41 males) of the Cincinnati Lead Study from birth to 78 months. During adulthood, peripheral blood DNA was used to quantify CpG methylation in peripheral blood leukocytes at DMRs of 22 human imprinted genes using Sequenom EpiTYPER assays. Statistical analyses were conducted using linear regression.
Mean blood lead concentration from birth to 78 months was associated with a significant decrease in PEG3 DMR methylation, (β=-0.0014, 95% CI:-0.0023, -0.0005, p=0.002), stronger in males, (β=-0.0024, 95% CI:-0.0038, -0.0009, p=0.003) than females (β=-0.0009, 95% CI:-0.0020, 0.0003, p=0.1). Elevated mean childhood blood lead concentration was also associated with a significant decrease in IGF2/H19 (β=-0.0013, 95% CI:-0.0023, -0.0003, p=0.01) DMR methylation, but primarily in females, (β=-0.0017, 95% CI:-0.0029, -0.0006, p=0.005) than males, (β=-0.0004, 95% CI:-0.0023, 0.0015, p=0.7). Elevated blood lead concentration during the neonatal period was associated with higher PLAGL1/HYMAI DMR methylation regardless of sex, (β=0.0075, 95% CI:0.0018, 0.0132, p=0.01). The magnitude of associations between cumulative lead exposure and CpG methylation remained unaltered from 30 to 78 months.
Our findings provide evidence for early childhood lead exposure resulting in sex-dependent and gene-specific DNA methylation differences in the DMRs of PEG3, IGF2/H19 and PLAGL1/HYMAI in adulthood.
BACKGROUND: Human evidence on the effects of early life phthalate exposure on obesity and cardiovascular disease risks, reported by experimental studies, is limited to a few cross-sectional studies.
OBJECTIVES: We evaluated the associations between prenatal phthalate exposure and childhood growth and blood pressure in a Spanish birth cohort study.
METHODS: We assessed exposure using the average of two phthalate metabolite spot-urine concentrations collected from the mothers in the first and third pregnancy trimesters (creatinine-adjusted, n=391). Study outcomes were the difference in age- and sex- specific Z-scores for weight between birth and 6 months of age; and repeated age- and sex-specific Z-scores for body mass index (BMI) at 1, 4 and 7 years; waist-to-height ratio at 4 and 7 years; and age- and height-specific Z-scores for systolic and diastolic blood pressure at 4 and 7 years.
RESULTS: The sum of 5 high molecular weight phthalate metabolites (ΣHMWPm) was associated with lower weight Z-score difference between birth and 6 months (β per doubling of exposure=-0.41; 95%CI: -0.75, -0.06) and BMI Z-scores at later ages in boys (β=-0.28; 95% CI:-0.60, 0.03) and with higher weight Z-score difference (β=0.24; 95% CI:-0.16, 0.65) and BMI Z-scores in girls (β=0.30; 95% CI:-0.04, 0.64) (P for sex interaction=0.01 and 0.05, respectively). The sum of 3 low molecular weight phthalates (ΣLMWPm) was not significantly associated with any of the growth outcomes. ΣHMWPm and ΣLMWPm were associated with lower systolic blood pressure Z-scores in girls but not in boys.
CONCLUSIONS: This study suggests that prenatal phthalate exposure may be associated with postnatal growth and blood pressure in a sex-specific manner. Inconsistencies with previous cross-sectional findings highlight the necessity for evaluating phthalate health effects in prospective studies.
ABSTRACT DNA methylation data assayed using pyrosequencing techniques are increasingly being used in human cohort studies to investigate associations between epigenetic modifications at candidate genes and exposures to environmental toxicants and to examine environmentally-induced epigenetic alterations as a mechanism underlying observed toxicant-health outcome associations. For instance, in utero lead (Pb) exposure is a neurodevelopmental toxicant of global concern that has also been linked to altered growth in human epidemiological cohorts; a potential mechanism of this association is through alteration of DNA methylation (e.g., at growth-related genes). However, because the associations between toxicants and DNA methylation might be weak, using appropriate quality control and statistical methods is important to increase reliability and power of such studies. Using a simulation study, we compared potential approaches to estimate toxicant-DNA methylation associations that varied by how methylation data were analyzed (repeated measures vs. averaging all CpG sites) and by method to adjust for batch effects (batch controls vs random effects). We demonstrate that correcting for batch effects using plate controls yields unbiased associations, and that explicitly modeling the CpG site-specific variances and correlations among CpG sites increases statistical power. Using the recommended approaches, we examined the association between DNA methylation (in LINE-1 and growth related genes IGF2, H19 and HSD11B2) and three biomarkers of Pb exposure (Pb concentrations in umbilical cord blood, maternal tibia, and maternal patella), among mother-infant pairs of the Early Life Exposures in Mexico to Environmental Toxicants (ELEMENT) cohort (n = 247). Those with 10 μg/g higher patella Pb had, on average, 0.61% higher IGF2 methylation (p = 0.05). Sex-specific trends between Pb and DNA methylation (p<0.1) were observed among girls including a 0.23% increase in HSD11B2 methylation with 10 μg/g higher patella Pb.
Epigenetic regulation of imprinted genes is regarded as a highly plausible explanation for linking dietary exposures in early life with the onset of diseases during childhood and adulthood. We sought to test whether prenatal dietary supplementation with docosahexaenoic acid (DHA) during pregnancy may modulate epigenetic states at birth. This study was based on a randomized intervention trial conducted in Mexican pregnant women supplemented daily with 400 mg of DHA or a placebo from gestation week 18-22 to parturition. We applied quantitative profiling of DNA methylation states at IGF2 promoter 3 (IGF2 P3), IGF2 differentially methylated region (DMR), and H19 DMR in cord blood mononuclear cells of the DHA-supplemented group (n=131) and the control group (n=130). In stratified analyses, DNA methylation levels in IGF2 P3 were significantly higher in the DHA group than the control group in preterm infants (p=0.04). We also observed a positive association between DNA methylation levels and maternal body mass index (BMI); IGF2 DMR methylation was higher in the DHA group than the control group in infants of overweight mothers (p=0.03). In addition, at H19 DMR, methylation levels were significantly lower in the DHA group than the control group in infants of normal weight mothers (p=0.01). Finally, methylation levels at IGF2/H19 imprinted regions were associated with maternal BMI. These findings suggest that epigenetic mechanisms may be modulated by DHA, with potential impacts on child growth and development.
Background: Studies suggest that phthalate exposures may adversely affect child respiratory health.
Objectives: We evaluated associations between asthma diagnosed in children between 5 and 11 years of age and prenatal exposures to butylbenzyl phthalate (BBzP), di-n-butyl phthalate (DnBP), di(2-ethylhexyl) phthalate (DEHP), and diethyl phthalate (DEP).
Methods: Phthalate metabolites were measured in spot urine collected from 300 pregnant inner-city women. Children were examined by an allergist or pulmonologist based on the first parental report of wheeze, other respiratory symptoms, and/or use of asthma rescue/controller medication in the preceding 12 months on repeat follow-up questionnaires. Standardized diagnostic criteria were used to classify these children as either having or not having current asthma at the time of the physician examination. Children without any report of wheeze or the other asthma-like symptoms were classified as nonasthmatics at the time of the last negative questionnaire. Modified Poisson regression analyses were used to estimate relative risks (RR) controlling for specific gravity and potential confounders.
Results: Of 300 children, 154 (51%) were examined by a physician because of reports of wheeze, other asthma-like symptoms, and/or medication use; 94 were diagnosed with current asthma and 60 without current asthma. The remaining 146 children were classified as nonasthmatic. Compared with levels in nonasthmatics, prenatal metabolites of BBzP and DnBP were associated with a history of asthma-like symptoms (p < 0.05) and with the diagnosis of current asthma: RR = 1.17 (95% CI: 1.01, 1.35) and RR = 1.25 (95% CI: 1.04, 1.51) per natural log-unit increase, respectively. Risk of current asthma was > 70% higher among children with maternal prenatal BBzP and DnBP metabolite concentrations in the third versus the first tertile.
Conclusion: Prenatal exposure to BBzP and DnBP may increase the risk of asthma among inner-city children. However, because this is the first such finding, results require replication.
Citation: Whyatt RM, Perzanowski MS, Just AC, Rundle AG, Donohue KM, Calafat AM, Hoepner LA, Perera FP, Miller RL. 2014. Asthma in inner-city children at 5–11 years of age and prenatal exposure to phthalates: the Columbia Center for Children’s Environmental Health Cohort. Environ Health Perspect 122:1141–1146; http://dx.doi.org/10.1289/ehp.1307670
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.
Diabetes mellitus represents a group of complex metabolic diseases that result in impaired glucose homeostasis, which includes destruction of β-cells or the failure of these insulin-secreting cells to compensate for increased metabolic demand. Despite a strong interest in characterizing the transcriptome of the different human islet cell types to understand the molecular basis of diabetes, very little attention has been paid to the role of long non-coding RNAs (lncRNAs) and their contribution to this disease. Here we summarize the growing evidence for the potential role of these lncRNAs in β-cell function and dysregulation in diabetes, with a focus on imprinted genomic loci.
Epigenetic mechanisms are proposed to link maternal concentrations of methyl group donor nutrients with the risk of low birth weight. However, empirical data are lacking. We have examined the association between folate and birth weight and assessed the mediating role of DNA methylation at nine differentially methylated regions (DMRs) of genomically imprinted genes in these associations. Compared with newborns of women with folate levels in the lowest quartile, birth weight was higher in those newborns of mothers in the second (β = 143.2, se = 63.2, P = 0.02), third (β = 117.3, se = 64.0, P = 0.07), and fourth quartile (β = 133.9, se = 65.2, P = 0.04), consistent with a threshold effect. This pattern of association did not vary by race/ethnicity but was more apparent in newborns of non-obese women. DNA methylation at the PLAGL1, SGCE, DLK1/MEG3 and IGF2/H19 DMRs was associated with maternal folate levels and also birth weight, suggestive of threshold effects. A role for methylation in the mediation of the association between maternal folate levels and birth weight was significant only for the MEG3 DMR (P<0.05). While the small sample size and partial scope of examined DMRs limit our conclusions, our data suggest that, with respect to birth weight, no additional benefits may be derived from increased maternal folate concentrations, especially in non-obese women. These data also support epigenetic plasticity as a key mechanistic response to folate availability during early fetal development.
Background: Phthalates are ubiquitous environmental contaminants. Because of potential adverse effects on human health, butylbenzyl phthalate (BBzP; metabolite = monobenzyl phthalate (MBzP)), di-n-butyl phthalate (DnBP; metabolite = mono-n-butyl phthalate (MnBP)), and di(2-ethylhexyl) phthalate (DEHP) are being replaced by substitutes including other phthalates, however little is known about consequent trends in population-level exposures.
Objective: To examine temporal trends in urinary concentrations of phthalate metabolites in the US general population, and whether trends vary by socio-demographic characteristics.
Methods: We combined data on 11 phthalate metabolites for 11071 participants from five cycles of the National Health and Nutrition Examination Survey (2001-2010). Percent changes and least square geometric means (LSGMs) were calculated from multivariate regression models.
Results: LSGM concentrations of monoethyl phthalate, MnBP, MBzP, and ∑DEHP metabolites decreased between 2001-2002 and 2009-2010 (percent change (95% CI): -42% (-49, -34); -17% (-23, -9); -32% (-39, -23) and -37% (-46, -26), respectively). In contrast, LSGM concentrations of monoisobutyl phthalate, mono(3-carboxypropyl) phthalate (MCPP), monocarboxyoctyl phthalate, and monocarboxynonyl phthalate (MCNP) increased over the study period (percent change (95% CI): 206% (178, 236); 25% (8, 45); 149% (102, 207); and 15% (1, 30), respectively). Trends varied by subpopulations for certain phthalates. For example, LSGM concentrations of ∑DEHP metabolites, MCPP, and MCNP were higher in children than adults but the gap between groups narrowed over time (pinteraction < 0.01).
Conclusions: US population exposure to phthalates has changed in the last decade. Data gaps make it difficult to explain trends but legislative activity and advocacy campaigns by non-governmental organizations may play a role.
Background:Several epidemiologic studies have demonstrated associations between periconceptional environmental exposures and health status of the offspring in later life. Although these environmentally related effects have been attributed to epigenetic changes, such as DNA methylation shifts at imprinted genes, little is known about the potential effects of maternal and paternal preconceptional overnutrition or obesity.Objective:We examined parental preconceptional obesity in relation to DNA methylation profiles at multiple human imprinted genes important in normal growth and development: MEG3, MEST, PEG3, PLAGL1, SGCE/PEG10, and NNAT.Methods:We measured methylation percentages at the DMRs by bisulfite pyrosequencing in DNA extracted from umbilical cord blood leukocytes of 92 newborns. Preconceptional obesity, defined as BMI 30 kg/m(2), was ascertained through standardized questionnaires.Results:After adjusting for potential confounders and cluster effects, paternal obesity was significantly associated with lower methylation levels at the MEST (β=-2.57; s.e.=0.95; P=0.008), PEG3 (-1.71; SE=0.61; P=0.005), and NNAT (-3.59; SE=1.76; P=0.04) DMRs. Changes related to maternal obesity were detected at other loci: β-coefficient was +2.58 (s.e.=1.00; P=0.01) at PLAGL1 DMR, and -3.42 (s.e.=1.69; P=0.04) at the MEG3 DMR.Conclusion:We found altered methylation outcomes at multiple imprint regulatory regions in children born to obese parents, compared to children born to non-obese parents. In spite of the small sample size, our data suggest a preconceptional influence of parental life-style or overnutrition on the reprogramming of imprint marks during gametogenesis. More specifically, the significant and independent association between paternal obesity and the offspring's methylation status suggests the susceptibility of the developing sperm for environmental insults. The acquired imprint instability may be carried on to the next generation and increase the risk for chronic diseases in adulthood.International Journal of Obesity accepted article preview online, 25 October 2013; doi:10.1038/ijo.2013.193.
Analysis of epigenetic mechanisms, particularly DNA methylation, is of increasing interest for epidemiologic studies examining disease etiology and impacts of environmental exposures. The Infinium HumanMethylation450 BeadChip® (450K), which interrogates over 480,000 CpG sites and is relatively cost effective, has become a popular tool to characterize the DNA methylome. For large-scale studies, minimizing technical variability and potential bias is paramount. The goal of this paper was to evaluate the performance of several existing and novel color channel normalizations designed to reduce technical variability and batch effects in 450K analysis from a large population study. Comparative assessment of 10 normalization procedures included the GenomeStudio® Illumina procedure, the lumi smooth quantile approach, and the newly proposed All Sample Mean Normalization (ASMN). We also examined the performance of normalizations in combination with correction for the two types of Infinium chemistry utilized on the 450K array. We observed that the performance of the GenomeStudio® normalization procedure was highly variable and dependent on the quality of the first sample analyzed in an experiment, which is used as a reference in this procedure. While the lumi normalization was able to decrease batch variability, it increased variation among technical replicates, potentially reducing biologically meaningful findings. The proposed ASMN procedure performed consistently well, both at reducing batch effects and improving replicate comparability. In summary, the ASMN procedure can improve existing color channel normalization, especially for large epidemiologic studies, and can be successfully implemented to enhance a 450K DNA methylation data pipeline.
Objectives:
Low birth weight (LBW) has been associated with common adult-onset chronic diseases, including obesity, cardiovascular disease, type II diabetes and some cancers. The etiology of LBW is multi-factorial. However, recent evidence suggests exposure to antibiotics may also increase the risk of LBW. The mechanisms underlying this association are unknown, although epigenetic mechanisms are hypothesized. In this study, we evaluated the association between maternal antibiotic use and LBW and examined the potential role of altered DNA methylation that controls growth regulatory imprinted genes in these associations.
Methods:
Between 2009-2011, 397 pregnant women were enrolled and followed until delivery. Prenatal antibiotic use was ascertained through maternal self-report. Imprinted genes methylation levels were measured at differentially methylated regions (DMRs) using bisulfite pyrosequencing. Generalized linear models were used to examine associations among antibiotic use, birth weight and DMR methylation fractions.
Results:
After adjusting for infant gender, race/ethnicity, maternal body mass index, delivery route, gestational weight gain, gestational age at delivery, folic acid intake, physical activity, maternal smoking and parity, antibiotic use during pregnancy was associated with 138 g lower birth weight compared with non-antibiotic use (β-coefficient=-132.99, s.e.=50.70, P=0.008). These associations were strongest in newborns of women who reported antibiotic use other than penicillins (β-coefficient=-135.57, s.e.=57.38, P=0.02). Methylation at five DMRs, IGF2 (P=0.05), H19 (P=0.15), PLAGL1 (P=0.01), MEG3 (P=0.006) and PEG3 (P=0.08), was associated with maternal antibiotic use; among these, only methylation at the PLAGL1 DMR was also associated with birth weight.
Conclusion:
We report an inverse association between in utero exposure to antibiotics and lower infant birth weight and provide the first empirical evidence supporting imprinted gene plasticity in these associations.
Cytology-based screening for invasive cervical cancer (ICC) lacks sensitivity and specificity to discriminate between cervical intraepithelial neoplasia (CIN) likely to persist or progress from cases likely to resolve. Genome-wide approaches have been used to identify DNA methylation marks associated with CIN persistence or progression. However, associations between DNA methylation marks and CIN or ICC remain weak and inconsistent. Between 2008–2009, we conducted a hospital-based, case-control study among 213 Tanzania women with CIN 1/2/3 or ICC. We collected questionnaire data, biopsies, peripheral blood, cervical scrapes, Human papillomavirus (HPV) and HIV-1 infection status. We assessed PEG3 methylation status by bisulfite pyrosequencing. Multinomial logistic regression was used to estimate odds ratios (OR) and confidence intervals (CI 95%) for associations between PEG3 methylation status and CIN or ICC. After adjusting for age, gravidity, hormonal contraceptive use and HPV infection, a 5% increase in PEG3 DNA methylation was associated with increased risk for ICC (OR = 1.6; 95% CI 1.2–2.1). HPV infection was associated with a higher risk of CIN1-3 (OR = 15.7; 95% CI 5.7–48.6) and ICC (OR = 29.5, 95% CI 6.3–38.4). Infection with high risk HPV was correlated with mean PEG3 differentially methylated regions (DMRs) methylation (r = 0.34 p<0.0001), while the correlation with low risk HPV infection was weaker (r = 0.16 p = 0.047). Although small sample size limits inference, these data support that PEG3 methylation status has potential as a molecular target for inclusion in CIN screening to improve prediction of progression.
Several phthalates, particularly diethyl phthalate (DEP) and di-n-butyl phthalate, can be used in personal care products (PCPs) to fix fragrance and hold color. We investigated associations between women's reported use of PCPs within the 24 h before urine collection and concentrations of several urinary phthalate metabolites. Between 2002 and 2005, 337 women provided spot urine samples and answered questions regarding their use of 13 PCPs at a follow-up visit 3-36 months after pregnancy. We examined associations between urinary concentrations of several phthalate metabolites and use of PCPs using linear regression. Use of individual PCPs ranged from 7% (nail polish) to 91% (deodorant). After adjusting for age, education, and urinary creatinine, women reporting use of perfume had 2.92 times higher (95% CI: 2.20-3.89) concentration of monoethyl phthalate (MEP; the primary metabolite of DEP) than other women. Other PCPs that were significantly associated with MEP concentrations included: hair spray, nail polish, and deodorant. MEP concentrations increased with the number of PCPs used. PCP use was widespread in this group of recently pregnant women. Women's use of PCPs, particularly of perfumes and fragranced products, was positively associated with urinary concentration of multiple phthalate metabolites.Journal of Exposure Science and Environmental Epidemiology advance online publication, 21 November 2012; doi:10.1038/jes.2012.105.
Quantitative measurements of environmental factors greatly improve the quality of epidemiologic studies but can pose challenges because of the presence of upper or lower detection limits or inter- fering compounds, which do not allow for precise measured values. We consider the regression of an environmental measurement (dependent variable) on several covariates (independent variables). Various strategies are commonly employed to impute values for interval-measured data, including assignment of one-half the detection limit to nondetected values or of "fill-in" values randomly selected from an appropriate distribution. On the basis of a limited simulation study, we found that the former approach can be biased unless the percentage of measurements below detection limits is small (5-10%). The fill-in approach generally produces unbiased parameter estimates but may pro- duce biased variance estimates and thereby distort inference when 30% or more of the data are below detection limits. Truncated data methods (e.g., Tobit regression) and multiple imputation offer two unbiased approaches for analyzing measurement data with detection limits. If interest resides solely on regression parameters, then Tobit regression can be used. If individualized values for measurements below detection limits are needed for additional analysis, such as relative risk regression or graphical display, then multiple imputation produces unbiased estimates and nominal confidence intervals unless the proportion of missing data is extreme. We illustrate various
Epigenetic plasticity in relation to in utero exposures may mechanistically explain observed differences in the likelihood of developing common complex diseases including hypertension, diabetes and cardiovascular disease through the cumulative effects of subtle alterations in gene expression. Imprinted genes are essential mediators of growth and development and are characterized by differentially methylated regulatory regions (DMRs) that carry parental allele-specific methylation profiles. This theoretical 50% level of methylation provides a baseline from which endogenously- or exogenously-induced deviations in methylation can be detected. We quantified DNA methylation at imprinted gene DMRs in a large panel of human conceptal tissues, in matched buccal cell specimens collected at birth and at one year of age, and in the major cell fractions of umbilical cord blood to assess the stability of methylation at these regions. DNA methylation was measured using validated pyrosequencing assays at seven DMRs regulating the IGF2/H19, DLK1/MEG3, MEST, NNAT and SGCE/PEG10 imprinted domains. DMR methylation did not significantly differ for the H19, MEST and SGCE/PEG10 DMRs across all conceptal tissues analyzed (ANOVA p>0.10). Methylation differences at several DMRs were observed in tissues from brain (IGF2 and MEG3-IG DMRs), liver (IGF2 and MEG3 DMRs) and placenta (both DLK1/MEG3 DMRs and NNAT DMR). In most infants, methylation profiles in buccal cells at birth and at one year of age were comparable, as was methylation in the major cell fractions of umbilical cord blood. Several infants showed temporal deviations in methylation at multiple DMRs. Similarity of inter-individual and intra-individual methylation at some, but not all of the DMRs analyzed supports the possibility that methylation of these regions can serve as useful biosensors of exposure.
Background
There has been a long-standing need in biomedical research for a method that quantifies the normally mixed composition of leukocytes beyond what is possible by simple histological or flow cytometric assessments. The latter is restricted by the labile nature of protein epitopes, requirements for cell processing, and timely cell analysis. In a diverse array of diseases and following numerous immune-toxic exposures, leukocyte composition will critically inform the underlying immuno-biology to most chronic medical conditions. Emerging research demonstrates that DNA methylation is responsible for cellular differentiation, and when measured in whole peripheral blood, serves to distinguish cancer cases from controls.
Results
Here we present a method, similar to regression calibration, for inferring changes in the distribution of white blood cells between different subpopulations (e.g. cases and controls) using DNA methylation signatures, in combination with a previously obtained external validation set consisting of signatures from purified leukocyte samples. We validate the fundamental idea in a cell mixture reconstruction experiment, then demonstrate our method on DNA methylation data sets from several studies, including data from a Head and Neck Squamous Cell Carcinoma (HNSCC) study and an ovarian cancer study. Our method produces results consistent with prior biological findings, thereby validating the approach.
Conclusions
Our method, in combination with an appropriate external validation set, promises new opportunities for large-scale immunological studies of both disease states and noxious exposures.
In many normal tissues, proliferation rates decline postnatally, causing somatic growth to slow. Previous evidence suggests that this decline is due, in part, to decline in the expression of growth-promoting imprinted genes including Mest, Plagl1, Peg3, Dlk1, and Igf2. Embryonal cancers are composed of cells that maintain embryonic characteristics and proliferate rapidly in childhood. We hypothesized that the abnormal persistent rapid proliferation in embryonal cancers occurs in part because of abnormal persistent high expression of growth-promoting imprinted genes.
Analysis of microarray data showed elevated expression of MEST, PLAGL1, PEG3, DLK1, and IGF2 in various embryonal cancers, especially rhabdomyosarcoma, as compared to nonembryonal cancers and normal tissues. Similarly, mRNA expression, assessed by real-time PCR, of MEST, PEG3, and IGF2 in rhabdomyosarcoma cell lines was increased as compared to nonembryonal cancer cell lines. Furthermore, siRNA-mediated knockdown of MEST, PLAGL1, PEG3, and IGF2 expression inhibited proliferation in Rh30 rhabdomyosarcoma cells.
These findings suggest that the normal postnatal downregulation of growth-promoting imprinted genes fails to occur in some embryonal cancers, particularly rhabdomyosarcoma, and contributes to the persistent rapid proliferation of rhabdomyosarcoma cells and, more generally, that failure of the mechanisms responsible for normal somatic growth deceleration can promote tumorigenesis.
Epigenetic changes such as DNA methylation may be a molecular mechanism through which environmental exposures affect health. Phthalates are known endocrine disruptors with ubiquitous exposures in the general population including pregnant women, and they have been linked with a number of adverse health outcomes. We examined the association between in utero phthalate exposure and altered patterns of cord blood DNA methylation in 336 Mexican-American newborns. Concentrations of 11 phthalate metabolites were analyzed in maternal urine samples collected at 13 and 26 weeks gestation as a measure of fetal exposure. DNA methylation was assessed using the Infinium HumanMethylation 450K BeadChip adjusting for cord blood cell composition. To identify differentially methylated regions (DMRs) that may be more informative than individual CpG sites, we used two different approaches, DMRcate and comb-p. Regional assessment by both methods identified 27 distinct DMRs, the majority of which were in relation to multiple phthalate metabolites. Most of the significant DMRs (67%) were observed for later pregnancy (26 weeks gestation). Further, 51% of the significant DMRs were associated with the di-(2-ethylhexyl) phthalate metabolites. Five individual CpG sites were associated with phthalate metabolite concentrations after multiple comparisons adjustment (FDR), all showing hypermethylation. Genes with DMRs were involved in inflammatory response (IRAK4 and ESM1), cancer (BRCA1 and LASP1), endocrine function (CNPY1), and male fertility (IFT140, TESC, and PRDM8). These results on differential DNA methylation in newborns with prenatal phthalate exposure provide new insights and targets to explore mechanism of adverse effects of phthalates on human health. Environ. Mol. Mutagen., 2017. © 2017 Wiley Periodicals, Inc.
Background:
Although experiments in animals suggest phthalates may have obesogenic effects, studies of prenatal exposure in children show inconsistent results.
Methods:
We measured urinary concentrations of 11 phthalate metabolites collected twice during pregnancy from mothers in the Center for the Health Assessment of Mothers and Children of Salinas (CHAMACOS) cohort study (N=345). Height, weight, waist circumference, and percent body fat were assessed in their children between 5 and 12 years of age. We used generalized estimating equations to examine associations at each age and tested for interaction by sex.
Results:
Metabolites of diethyl phthalate (DEP), di-n-butyl phthalate (DBP), butyl benzyl phthalate, and di(2-ethylhexyl) phthalate (DEHP) were positively associated with body mass index z-score, waist circumference z-score, and percent body fat at multiple ages. At age 12, we observed increased odds of being overweight/obese with each doubling of prenatal concentrations of DEP (OR=1.3; 95% CI: 1.1, 1.4), DBP (1.2; 1.0, 1.4), and DEHP (1.3; 1.0, 1.6) metabolites. Results were similar in boys and girls except for DBP metabolites and the non-specific metabolite mono-(3-carboxypropyl) phthalate, which showed positive associations only in boys.
Conclusions:
In utero exposure to certain phthalates is associated with increased BMI and risk of overweight/obesity in childhood.Pediatric Research accepted article preview online, 20 April 2017. doi:10.1038/pr.2017.112.
Background:
Being small for gestational age (SGA), a foetal growth abnormality, has a long-lasting impact on childhood health. Its aetiology and underlying mechanisms are not well understood. Underlying epigenetic changes of imprinted genes have emerged as a potential pathological pathway because they may be associated with growth, including SGA. As a common methyl donor, folic acid (FA) is essential for DNA methylation, synthesis and repair, and FA supplementation is widely recommended for women planning pregnancy. The present study aimed to investigate the inter-relationships among methylation levels of two imprinted genes [H19 differentially methylated regions (DMRs) and MEST DMRs], maternal FA supplementation and SGA.
Methods:
We conducted a case-control study. Umbilical cord blood was taken from 39 SGA infants and 49 controls whose birth weights are appropriate for gestational age (AGA). DNA methylation levels of H19 and MEST DMRs were determined by an analysis of mass array quantitative methylation.
Results:
Statistically significantly higher methylation levels were observed at sites 7.8, 9 and 17.18 of H19 (P = 0.030, 0.016 and 0.050, respectively) in the SGA infants compared to the AGA group. In addition, the association was stronger in male births where the mothers took FA around conception at six H19 sites (P = 0.004, 0.005, 0.048, 0.002, 0.021 and 0.005, respectively).
Conclusions:
Methylation levels at H19 DMRs were higher in SGA infants compared to AGA controls. It appears that the association may be influenced by maternal peri-conception FA supplementation and also be sex-specific.
Phthalates are frequently used in personal care products and plasticizers and phthalate exposure is ubiquitous in the US population. Exposure to phthalates during critical periods in utero has been associated with a variety of adverse health outcomes but the biological mechanisms linking these exposures with disease are not well characterized. In this study, we examined the relationship of in utero phthalate exposure with repetitive element DNA methylation, an epigenetic marker of genome instability, in children from the longitudinal birth cohort CHAMACOS. Methylation of Alu and long interspersed nucleotide elements (LINE-1) was determined using pyrosequencing of bisulfite-treated DNA isolated from whole blood samples collected from newborns and 9 year old children (n=355). Concentrations of eleven phthalate metabolites were measured in urine collected from pregnant mothers at 13 and 26 weeks gestation. We found a consistent inverse association between prenatal concentrations of monoethyl phthalate, the most frequently detected urinary metabolite, with cord blood methylation of Alu repeats (β(95%CI): -0.14 (-0.28,0.00) and -0.16 (-0.31, -0.02)) for early and late pregnancy, respectively, and a similar but weaker association with LINE-1 methylation. Additionally, increases in urinary concentrations of di-(2-ethylhexyl) phthalate metabolites during late pregnancy were associated with lower levels of methylation of Alu repeats in 9 year old blood (significant p-values ranged from 0.003 to 0.03). Our findings suggest that prenatal exposure to some phthalates may influence differences in repetitive element methylation, highlighting epigenetics as a plausible biological mechanism through which phthalates may affect health.
Three recent genome-wide studies in mice and humans have produced the most definitive map to date of genomic imprinting (gene expression that depends on parental origin) by incorporating multiple tissue types and developmental stages. Here, we explore the results of these studies in light of the kinship theory of genomic imprinting, which predicts that imprinting evolves due to differential genetic relatedness between maternal and paternal relatives. The studies produce a list of imprinted genes with around 120-180 in mice and ∼100 in humans. The studies agree on broad patterns across mice and humans including the complex patterns of imprinted expression at loci like Igf2 and Grb10. We discuss how the kinship theory provides a powerful framework for hypotheses that can explain these patterns. Finally, since imprinting is rare in the genome despite predictions from the kinship theory that it might be common, we discuss evolutionary factors that could favor biallelic expression.
Prenatal exposure to endocrine disrupting chemicals may affect fetal development through disruption of hormonal actions and epigenetic modifications, potentially predisposing individuals to later on-set health risks, such as obesity. The objective of this study was to determine associations between biological exposure markers of various endocrine disrupting chemicals and birth weight in a newly established, prospective mother-child cohort in the Netherlands. Birth weight (n = 91) was obtained from birth records, and exposure to dichlorodiphenyldichloroethylene (DDE), three di-2-ethylhexyl phthalate (DEHP) metabolites, polychlorinated biphenyl-153, perfluorooctanesulfonic acid (PFOS), and perfluorooctanoic acid (PFOA) was determined in cord plasma. For DDE, exposure was also measured in breast milk. Linear regression analysis was used to determine associations between compounds and birth weight, which were stratified for gender and adjusted for a priori defined covariates. Increased exposure to DDE was associated with lower birth weight in boys (>95.89 ng L(-1), -325.9 g, 95% CI -634.26 to -17.56), whereas in girls a tendency towards a higher birth weight was observed. Lower birth weights for boys were also observed for high exposure to MECPP, and to a certain extent also for PFOA. MEHHP and PFOS exposure on the other hand were associated with higher birth weights in boys. In girls no effects were observed for these compounds. It can be concluded that prenatal exposure to DDE, perfluorinated alkyl acids, and phthalates was associated with changes in birth weight in this population. Associations were gender specific, and appeared to be non-linear. Since the population was relatively small, results should be interpreted with caution.
Confounding by cellular heterogeneity has become a major concern for epigenome-wide association studies (EWAS) in peripheral blood samples from population and clinical studies. Adjusting for white blood cell percentage estimates produced by the minfi implementation of the Houseman algorithm (minfi) during statistical analysis is now an established method to account for this bias in adults. However, minfi has not been benchmarked against white blood cell counts in children that may differ substantially from the reference dataset used in its estimation. We compared estimates of white blood cell type percentages produced by two methods, minfi and differential cell count (DCC), in a birth cohort at two time points (birth and 12 years of age). We found that both minfi and DCC had similar trends as children aged, and neither count method differed by sex among newborns (P > 0.10). However, minfi estimates did not correlate well with DCC in samples from newborns (ρ = −0.05 for granulocytes; ρ = −0.03 for lymphocytes). In older children, correlation improved substantially (ρ = 0.77 for granulocytes; ρ = 0.75 for lymphocytes), likely due to increasing similarity with minfi's adult reference data as children aged. Our findings suggest that the minfi method may provide suitable estimates of white blood cell composition for samples from adults and older children, but may not currently be appropriate for EWAS involving newborns or young children. Environ. Mol. Mutagen., 2015. © 2015 Wiley Periodicals, Inc.
Increasing evidence indicates that long noncoding RNAs (lncRNAs) are involved in diverse biological process. Mouse maternal expressed gene 3 (Meg3), is an imprinted gene and essential for development. Here, we explored the relationship between Meg3 and the function of mouse beta cells in vitro and vivo. Real-time PCR analyses revealed Meg3 was more abundantly expressed in Balb/c mouse islets than exocrine glands. Moreover, the expression of Meg3 in islets was decreased in T1DM (NOD female mice) and T2DM (db/db mice) models. Meg3 expression was modulated dynamically by glucose in Min6 cells and isolated mouse islets. The function role of Meg3 was investigated in Min6 cells and normal mouse by knockdown of Meg3 using small interfering RNA. After suppression of Meg3 expression in vitro, insulin synthesis and secretion were impaired and the rate of beta cells apoptosis was increased. Moreover, knockdown of Meg3 in vivo led to the impaired glucose tolerance and decreased insulin secretion, consisted with the reduction of insulin positive cells areas by immunochemistry assays. Notably, islets from Meg3 interference groups showed significant decrease of Pdx-1 and MafA expression in mRNA and protein levels. These results indicate that Meg3 may function as a new regulator of maintaining beta cells identity via affecting insulin production and cell apoptosis. This article is protected by copyright. All rights reserved.
This article is protected by copyright. All rights reserved.
Exposure to environmental toxicants during fetal development alters gene expression and promotes disease later in life. Di-(2-ethylhexyl) phthalate (DEHP) is a plasticizer widely used for the manufacturing of consumer products. Exposure to DEHP has been associated with obesity, asthma, and low testosterone levels. In utero exposure of pregnant dams to DEHP from gestational day 14 until birth resulted in reduced levels of serum testosterone and aldosterone in the adult male offspring. Since DEHP is rapidly cleared from the body, the effects observed in the adult are likely epigenetic in origin. Under the same experimental conditions, we used reduced-representation bisulfite sequencing to assess changes in DNA methylation. We identified hot spots of DNA methylation changes primarily within CpG islands followed by shelf regions of the genome known to control regional gene expression. We also identified epigenomic areas responsive to exposure to environmental levels of DEHP and found the chromosomal region that houses genes controlling immune responsiveness to be a primary target of DEHP. These data suggest that DEHP phthalate exposure early in life induces epigenetic changes that may be linked to altered gene expression and function in the adult.
Prenatal exposure to inorganic arsenic (iAs) is detrimental to the health of newborns and increases the risk of disease development later in life. Here we examined a subset of newborn cord blood leukocyte samples collected from mothers enrolled in the Biomarkers of Exposure to ARsenic (BEAR) pregnancy cohort in Gómez Palacio, Mexico who were exposed to a range of drinking water arsenic concentrations (0.456-236 μg/L). Changes in iAs-associated DNA 5-methyl cytosine methylation were assessed across 424,935 CpG sites representing 18,761 genes and were compared to corresponding expression levels and birth outcomes. In the context of arsenic exposure, a total of 2,705 genes were identified with iAs-associated differences in DNA methylation. Site-specific analyses identified DNA methylation changes that were most predictive of gene expression levels. Specifically, CpG methylation within CpG islands positioned within the first exon and 200bp upstream of the transcription start site yielded the most significant association with gene expression levels. A set of 16 genes was identified with correlated iAs-associated changes in DNA methylation and mRNA expression and all were highly enriched for binding sites of the early growth response (EGR) and CCCTC-binding factor (CTCF) transcription factors. Furthermore, DNA methylation levels of seven of these genes were associated with differences in birth outcomes including gestational age, placental weight and head circumference. These data highlight the complex interplay between DNA methylation and functional changes in gene expression and health outcomes and underscore the need for functional analyses coupled to epigenetic assessments.
Genomic imprinting leads to parent-of-origin specific gene expression and is determined by epigenetic modification of genes. The paternally expressed gene insulin-like growth-factor 2 (IGF2) is located about ~100kb from the maternally expressed non-coding gene H19 on human chromosome 11, and both genes play major roles in embryonic and placental growth. Given adverse gestational environments can influence DNA methylation patterns in extra-embryonic tissues, we hypothesized that prenatal exposure to endocrine disrupting chemicals (EDCs) alters H19 and IGF2 methylation in placenta. Our study was restricted to a total of 196 women co-enrolled in the Predictors of Preeclampsia Study and the Harvard Epigenetic Birth Cohort. First trimester urine concentrations of 8 phenols and 11 phthalate metabolites were measured and used to characterize EDC exposure profiles. We assessed methylation of differentially methylated regions (DMRs) by pyrosequencing of H19, IGF2DMR0, and IGF2DMR2 and correlated values with phenol and phthalate metabolites. We also assessed overall expression and allele-specific expression of H19 and IGF2. We found several significant associations between DNA methylation and additive biomarker measurements. A significant decrease in H19 methylation was associated with high levels of the sum (Σ) of phthalate metabolites and metabolites of low molecular weight (LMW) phthalates. Σphthalate and LMW phthalate concentrations were inversely associated with IGF2DMR0 methylation values. Variation in methylation was not associated with changes in allele-specific expression. However increased deviation of allele-specific expression of H19 was associated with Σdi(2-ethylhexyl) phthalate metabolites and high molecular weight phthalates. Neither methylation nor expression of these imprinted regions had a significant impact on birth length or birth weight. Overall, our study provides new insight into an epigenetic mechanism that occurs following EDC exposure.
Genomic imprinting affects a subset of genes in mammals and results in a monoallelic, parental-specific expression pattern. Most of these genes are located in clusters that are regulated through the use of insulators or long noncoding RNAs (lncRNAs). To distinguish the parental alleles, imprinted genes are epigenetically marked in gametes at imprinting control elements through the use of DNA methylation at the very least. Imprinted gene expression is subsequently conferred through lncRNAs, histone modifications, insulators, and higher-order chromatin structure. Such imprints are maintained after fertilization through these mechanisms despite extensive reprogramming of the mammalian genome. Genomic imprinting is an excellent model for understanding mammalian epigenetic regulation.
Type 2 diabetes mellitus (T2DM) is a complex disease characterized by the inability of the insulin-producing β cells in the endocrine pancreas to overcome insulin resistance in peripheral tissues. To determine if microRNAs are involved in the pathogenesis of human T2DM, we sequenced the small RNAs of human islets from diabetic and nondiabetic organ donors. We identified a cluster of microRNAs in an imprinted locus on human chromosome 14q32 that is highly and specifically expressed in human β cells and dramatically downregulated in islets from T2DM organ donors. The downregulation of this locus strongly correlates with hypermethylation of its promoter. Using HITS-CLIP for the essential RISC-component Argonaute, we identified disease-relevant targets of the chromosome 14q32 microRNAs, such as IAPP and TP53INP1, that cause increased β cell apoptosis upon overexpression in human islets. Our results support a role for microRNAs and their epigenetic control by DNA methylation in the pathogenesis of T2DM.
Importance:
Preterm birth is a leading cause of neonatal mortality, with a variety of contributing causes and risk factors. Environmental exposures represent a group of understudied, but potentially important, factors. Phthalate diesters are used extensively in a variety of consumer products worldwide. Consequently, exposure in pregnant women is highly prevalent.
Objective:
To assess the relationship between phthalate exposure during pregnancy and preterm birth.
Design, setting, and participants:
This nested case-control study was conducted at Brigham and Women's Hospital, Boston, Massachusetts. Women were recruited for a prospective observational cohort study from 2006-2008. Each provided demographic data, biological samples, and information about birth outcomes. From within this group, we selected 130 cases of preterm birth and 352 randomly assigned control participants, and we analyzed urine samples from up to 3 time points during pregnancy for levels of phthalate metabolites.
Exposure:
Phthalate exposure during pregnancy.
Main outcomes and measures:
We examined associations between average levels of phthalate exposure during pregnancy and preterm birth, defined as fewer than 37 weeks of completed gestation, as well as spontaneous preterm birth, defined as preterm preceded by spontaneous preterm labor or preterm premature rupture of the membranes (n = 57).
Results:
Geometric means of the di-2-ethylhexyl phthalate (DEHP) metabolites mono-(2-ethyl)-hexyl phthalate (MEHP) and mono-(2-ethyl-5-carboxypentyl) phthalate (MECPP), as well as mono-n-butyl phthalate (MBP), were significantly higher in cases compared with control participants. In adjusted models, MEHP, MECPP, and Σ DEHP metabolites were associated with significantly increased odds of preterm birth. When spontaneous preterm births were examined alone, MEHP, mono-(2-ethyl-5-oxohexyl) phthalate, MECPP, Σ DEHP, MBP, and mono-(3-carboxypropyl) phthalate metabolite levels were all associated with significantly elevated odds of prematurity.
Conclusions and relevance:
Women exposed to phthalates during pregnancy have significantly increased odds of delivering preterm. Steps should be taken to decrease maternal exposure to phthalates during pregnancy.
Objective:
Phthalates and bisphenol A (BPA) are ubiquitous environmental toxicants, present in high concentrations in numerous consumer products. We hypothesized that maternal exposure to phthalates and BPA in pregnancy is associated with shortened gestation.
Methods:
Urinary phthalate and BPA metabolites from 72 pregnant women were measured at the last obstetric clinic visit prior to delivery. Using linear regression models, we estimated the change in gestational age associated with each interquartile range (IQR) increase in phthalate and BPA metabolite concentration.
Results:
IQR increases in urinary mono(2-ethyl-5-hydroxyhexyl) phthalate (MEHHP) and BPA concentrations were associated with 4.2 and 1.1 d decreases in gestation, respectively. When stratified by gender, these alterations were found only in male infants.
Conclusions:
We conclude that MEHHP and BPA (free + glucuronide) are associated with reductions in gestation, with effects observed only in males. Our findings are consistent with the idea that these agents induce gender-specific alterations in signaling via PPAR-γ transcription factor, androgen precursors and/or inflammatory mediators during the initiation of labor.
Epigenetic phenomena in animals and plants are mediated by DNA methylation and stable chromatin modifications. There has been considerable interest in whether environmental factors modulate the establishment and maintenance of epigenetic modifications, and could thereby influence gene expression and phenotype. Chemical pollutants, dietary components, temperature changes and other external stresses can indeed have long-lasting effects on development, metabolism and health, sometimes even in subsequent generations. Although the underlying mechanisms remain largely unknown, particularly in humans, mechanistic insights are emerging from experimental model systems. These have implications for structuring future research and understanding disease and development.