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REVIEW
Maternal DNA Methylation During Pregnancy: a Review
Jagyashila Das
1
&Arindam Maitra
1
Received: 29 June 2020 / Accepted: 29 December 2020
#Society for Reproductive Investigation 2021
Abstract
Multiple environmental, behavioral, and hereditary factors affect pregnancy. Recent studies suggest that epigenetic modifica-
tions, such as DNA methylation (DNAm), affect both maternal and fetal health during the period of gestation. Some of the
pregnancy-related risk factors can influence maternal DNAm, thus predisposing both the mother and the neonate to clinical
adversities with long-lasting consequences. DNAm alterations in the promoter and enhancer regions modulate gene expression
changes which play vital physiological role. In this review, we have discussed the recent advances in our understanding of
maternal DNA methylation changes during pregnancy and its associated complications such as gestational diabetes and anemia,
adverse pregnancy outcomes like preterm birth, and preeclampsia. We have also highlighted some major gaps and limitations in
the area which if addressed might improve our understanding of pregnancy and its associated adverse clinical conditions,
ultimately leading to healthy pregnancies and reduction of public health burden.
Keywords Maternal .DNA methylation .Pregnancy .Pregnancy-related anemia .Gestational diabetes .Preterm birth .
Preeclampsia
Introduction
Pregnancy involves complex dynamic cascades of physiolog-
ical processes in the mother and the fetus. Both genetic and
environmental factors modulate pregnancy [1]. According to
the Developmental Origins of Health and Disease (DOHaD)
theory, which was based on epidemiological observations,
early-life environment, i.e., of the fetus, determines the health
trajectories in later life [2]. Women, who are malnourished
and/or underweight during pregnancy, are more likely to have
inadequate gestational weight gain leading to adverse impact
on fetal development [3–5]. On the other hand, maternal obe-
sity, high weight gain during pregnancy and gestational dia-
betes leads to increased risk of cancer, type 2 diabetes and
obesity in the offspring during adulthood [6,7].
Adverse birth outcomes have heterogeneous underlying
etiologies. Most adverse outcomes have familial aggregation
and have genetic underpinnings. Although multiple genome-
wide association studies (GWAS) have been undertaken to
identify genetic variants in adverse pregnancy outcomes, most
of these studies do not explain a substantial proportion of the
risk [8–11]. Epidemiologic observations suggest a role for
other factors like maternal obesity, stress, socioeconomic sta-
tus (SES), environmental conditions [12]. Studies also suggest
impact of early-life stress during the developmental stages on
adverse pregnancy outcomes [13]. These factors modulate the
cellular functioning primarily via DNA methylation (DNAm),
non-coding RNA, and/or histone modifications, of which
DNAm is the most widely studied phenomenon [14–16].
Studies have only recently been initiated on pregnancy, birth,
and child health to investigate the impact of such factors on
the epigenome during gestation [17–20].
The covalently bound methyl groups on 5′cytosine sites
are not easily lost during routine DNA extraction, which ren-
ders DNAm as a reliable epigenomic marker which can be
utilized in large investigations [21]. Some DNAm marks are
inherited from parents, while others are acquired during the
lifetime of the individual [22,23]. The inherited marks can
persist for multiple generations, thus behave similar to DNA
mutations [24]. They may contribute to the “missing heritabil-
ity”of complex traits that are otherwise not explained by
DNA mutations alone [25]. On the other hand, dynamic
DNAm changes regulate the variable expression of the ge-
nome in diverse maternal tissues like the uterus, breasts, etc.
and in response to environmental factors from nulligravid to
postpartum state [26]. Orchestrated switching of DNAm
*Arindam Maitra
am1@nibmg.ac.in
1
National Institute of Biomedical Genomics, Kalyani, West
Bengal 741251, India
Reproductive Sciences
https://doi.org/10.1007/s43032-020-00456-4
marks across specific windows of gestation is key to healthy
pregnancy [2]. Recent evidences suggest that besides individ-
ual genome, maternal lifestyle shapes the trajectory of preg-
nancy with lasting impact onthe child [10]. One such example
is the Hunger Winter Families study, where it was found that
the adults who were prenatally exposed to famine, harbor
hypomethylation patterns in maternally imprinted insulin-
like growth factor II (IGF2) gene, a key factor in human
growth and development [27–29]. Hence, studies on
epigenomics of pregnancy may reveal important information
on the interactions between the maternal as well as the fetal
genome and the environment, which may ultimately lead to
reduction in occurrences of adverse pregnancy outcomes [30].
DNAm plays a crucial role in gene expression regulation,
normal cellular function, and embryonic development [31].
Understanding DNAm changes during pregnancy would help
to explain the underlying biological mechanisms important for
physiological alterations and adaptations that are required for
fetal development, as well as to prepare the mother for child-
birth and postnatal period. In this review we have focused on
discussing the information available on DNAm in women
during gestation and its relationship to adverse clinical condi-
tions associated with pregnancy.
DNA Methylation in Normal Pregnancy
After conception, a pregnant woman undergoes substantial
physiological and anatomical changes throughout pregnancy
to nurture the developing fetus. This is followed by invasion
of the fetal trophoblast cells by epithelial to mesenchymal
transition (EMT) and migration within the endometrium, pro-
moting angiogenesis and establishing an exchange of nutri-
ents, gases and waste between fetus and mother [32,33].
Muller et al. compared methylation status of healthy controls
with breast cancer patients and pregnant women [34]. They
showed that methylation profiles of candidate genes (CDH1,
PTGS2,APC,RASSF1A) behaved similarly in both the preg-
nant group and the advanced cancer group. These genes show
similar DNAm pattern in trophoblasts, to that of invading
tumor, which is also a characteristic of progressive cancer
[35]. However, age is known to effect methylation status
and this study was limited by the lack of matching of age of
participants [36].
Natural killer cells and macrophages facilitate implantation
and placentation [37]. During this process and subsequently,
maternal leukocytes come into contact with the placental tro-
phoblast cells of paternal origin. Immune tolerance is induced
during pregnancy in order to avoid rejection of the
semiallogenic fetus [38,39]. Hence the first and early second
trimester of pregnancy is considered to be the inflammatory
phase [40,41]. Leukocytes play a pivotal role in proliferation,
invasion, inflammation, and immune tolerance, critical for
maternal adaptation for normal placental and fetal develop-
ment [42]. The modifications of the immune system altogether
make the uterus an immunity “hotspot.”Agenome-wide
DNAm study on maternal leukocytes found evidences of
global hypomethylation in pregnant state compared to two
non-pregnant states, i.e., before pregnancy and 6 months post-
partum. The study identified eight hypomethylated genes,
which are related to immunity (IL1R2,HPR,TREML1), ga-
metogenesis (SPAG4,CCIN), and a few housekeeping genes
(PC,NDFUS2)[43]. Involvement of interleukin-1 (IL1)fam-
ily of genes has been previously implicated in the establish-
ment and progression of pregnancy [44]. Hypomethylation of
haptoglobin-related protein (HPR) gene might lead to in-
creased HPR expression during normal pregnancy [45,46].
The study showed that transient methylation in maternal leu-
kocyte DNA during pregnancy might result in
immunotolerant adaptation, which is lost soon after delivery.
A study on maternal venous white blood cells (WBCs)
identified changes in methylation profiles of the LINE-1 ele-
ments (long interspersed nucleotide elements 1), which are
typically heavily methylated in normal conditions, but are
hypomethylated during cellular stress [47]. The study sug-
gested that hypermethylation of these elements in early preg-
nancy may lead to PTB [48]. Whether these alterations are
specific for pregnancy remains undetermined as data on
DNAm status of the LINE1 elements in women before and
after pregnancy is unreported.
In a recent longitudinal study, Gruzieva et al. compared
global epigenomic alterations between two non-pregnant (be-
fore conception and 2–4 days after delivery) states, and
through period of gestation (10–14 weeks, 26–28 weeks into
pregnancy) [49]. The per-pregnancy state was used as the
baseline for methylation levels, for comparison with later
stages of pregnancy. An overall decrease in methylation of
178 CpGs among the 196 significant sites during pregnancy
was observed. There was an overlap of about 47% of signif-
icantly associated probes among the discovery and replication
cohort, with most probes exhibiting the same direction of
change of DNAm level. Some of the pathways identified were
those which control various metabolic processes such as insu-
lin receptor signaling, mammary gland fat development, and
adipose tissue development in the body. However, this study
on peripheral blood from pregnant women did not account for
the contribution of circulating fetal cells in the detected meth-
ylation patterns, which are expected to affect the findings [50].
It was observed that SERPINB5 (maspin) promoter was
completely methylated in maternal blood, whereas it was
unmethylated in the placenta in case of normal pregnancy
outcomes [51]. In another study, RASSF1A gene was found
to be hypomethylated in maternal blood throughout gestation,
while it was hypermethylated in placenta [52]. In a study on
assessment of intergenerational impact of childhood maltreat-
ment (CM) on DNAm changes in peripheral blood
Reprod. Sci.
mononuclear cells (PBMCs) from mothers and neonates [53],
researchers identified adaptive immune cell specific DNAm
alteration in stress-response related genes like FKBP5,
CRHR1,andNR3C1 (involved in cortisol signaling), which
were however not transmitted from mothers with CM to neo-
nates directly after birth. These DNAm differences of the
same genes between the mother and the fetus might thereby
enable identification of fetal-specific markers for non-invasive
prenatal diagnosis.
Most methylation studies conducted so far are mostly
based on candidate genes in fetal (placental, cord blood) tis-
sues, emphasizing on disorders and co-morbidities of preg-
nancy, such as preeclampsia, in vitro fertilization, and gesta-
tional diabetes [54–58]. However, identifying the genome-
wide differential methylation patterns in women during nor-
mal or healthy pregnancies might result in improved under-
standing of the genes and corresponding pathways that are
important for pregnancy. This might help us eventually to
improve our understanding of human pregnancies.
Pregnancy-Related Complications
and Maternal DNA Methylation
Gestational Diabetes
Altered secretion of hormones, such as cortisol, human pla-
cental lactogen insulin, estrogen and progesterone leads to
glucose-insulin imbalance during pregnancy, which results
in development of gestational diabetes (GD) [59,60]. In ad-
dition to these, a multitude of factors like ethnicity, polycystic
ovary syndrome, hypertension, etc. also contribute towards
GD in approximately 50% of the cases [61,62]. Most studies
of DNAm in GD have been undertaken in fetal tissues like the
cord blood and placenta [58,63]. Enquobahrie et al., in their
study on multigravidae women who have suffered from GD in
at least any one of the pregnancies, identified genes that were
hypermethylated (SEP11,ZAR1,DDR1) and those that were
hypomethylated (NDUFC,HAPLN3,HHLA3,RHOG)inma-
ternal peripheral blood mononuclear cells (PBMCs), as com-
pared to non-GD multigravidae women [64]. Interestingly,
one of the hypomethylated genes, NDUFC1, encodes for a
protein belonging to the complex I of the electron transport
chain located in the mitochondrial membrane [65]. It is known
that mitochondrial function in tissues like liver, muscles, and
pancreatic beta cells is critical for cellular metabolism and
modulation of oxidative activity and nutrient load [66,67].
However, roles of most of the other genes in pregnancy are
yet to be delineated.
Kang et al. compared global DNAm patterns of GD affect-
ed pregnant women and their children, with that of healthy
mother-offspring pairs, in blood samples drawn upon admis-
sion before delivery, and cord blood within short time interval
after delivery, in a cohort in Taiwan. The study identified 151
differentially methylated genes in GD mothers [68].
Association between GD and differentially methylated genes
belonging to carbohydrate and lipid metabolism pathways
have been previously reported [69]. This study also found
genes SLC22A4,ADRA1A,CACNB2,andSERPINE1 which
belong to the lipid metabolism pathways, to be differentially
methylated in GD mothers. Of these, ADRA1A has been re-
ported to be also involved in carbohydrate metabolism [70].
Alteration in DNAm of genes belonging to the GD related
metabolic pathways such as the Janus kinase (JAK)/mitogen
activated protein kinase (MAPK) pathways were also identi-
fied, which have been traditionally associated with endo-
toxins, inflammatory cytokines, and environmental stress
[68].
GD has been associated with macrosomia, large for gesta-
tional age, perinatal mortality, preeclampsia etc. and hence
there is a growing need to address it in the early stages of
pregnancy [71]. In an attempt to find clinical biomarkers in
women with GD, Wu et al., identified genome-wide DNAm
alterations in maternal peripheral whole blood collected from
pregnant women 12–16 weeks into pregnancy, prior to diag-
nosis of gestational diabetes mellitus (GDM). They identified
two differentially methylated genes (HOOK2,RDH1) which
were also reported in other studies conducted on placenta and
cord blood [72,73]. HOOK2 codes for a linker protein that
mediates binding to organelles and is responsible for morpho-
genesis of cilia and endocytosis [74]. RDH12 encodes a retinal
reductase, which plays a role in the metabolism of short-chain
aldehydes [75]. Five additional CpG loci in COPS8,PIK3R5,
HAAO,CCDC124,andC5orf34 genes were also identified,
although the CpG locations in the genomic context were not
mentioned [76]. These genes have the potential to be used as
biomarkers once validated in larger validation cohorts.
Pregnancy-Related Anemia
Globally, 41.8% of pregnant women are anemic [77]. The mid
trimester is marked by rapid feto-placental growth and devel-
opment, with increased blood volume but lowered viscosity
and reduced hemoglobin concentration in the mother, typical-
ly causing anemia [78–80]. There is a greater requirement of
folate during pregnancy, which is likely to be related to the
fetal growth and development [81]. It is known that folate-
mediated 1-carbon metabolism is essential for the high-
fidelity synthesis of DNA, maintenance of DNAm and regu-
lation of chromatin structure [82]. Knight et al., in 2018, in-
vestigated the association between one carbon metabolism
and DNAm changes during pregnancy in maternal peripheral
blood collected at < 12 weeks of pregnancy and at delivery
[83]. Out of the 993 CpG sites whose methylation levels in
maternal peripheral blood were found to be altered during
pregnancy, one site was found to be negatively correlated with
Reprod. Sci.
dimethylglycine (DMG) level in blood [83]. DMG is one of
the metabolites of the one carbon metabolism. Two CpGs
in PSMB7 and B4GALT5 genes were found to be associat-
ed with DMG concentrations in cord blood. Most of the
other CpG sites did not belong to any gene. This study
identified genome-wide significant metabolite-
methylation associations. However, whether DNAm was
regulated by metabolites or metabolites regulated DNAm
warrants further evaluation.
Maternal DNA Methylation in Adverse
Pregnancy Outcomes
Preterm Birth
Preterm birth (PTB), i.e., birth before 37 complete weeks of
gestation, is globally the leading cause of neonatal morbidity
[84,85]. Studies suggest that nutritional deficiencies, stress,
other factors such as cigarette smoking, maternal infection
(bacterial vaginosis/intra-amniotic infection), and race in-
crease the risk of PTB. Since some of these factors are envi-
ronmental in nature, it is likely that epigenetic mechanisms
such as DNAm may be involved in modulation of this risk
[86]. Multiple studies have been undertaken on PTB, primar-
ily in Caucasians and African Americans [87–89]. A study on
African-American women from the Boston Birth Cohort iden-
tified association of altered DNAm of two CpG sites with
spontaneous PTB (sPTB), in maternal WBCs collected 24–
72 h after delivery [90]. These CpG sites, one each in the
promoter regions of CYTIP (cytohesin 1 interacting protein)
and LINC0114 (long non-coding RNA) genes, were
hypomethylated in mothers who delivered early sPTB (24–
33.6/7 weeks) compared to those who delivered at term (39–
42 weeks). CYTIP is involved in leukocyte trafficking, T cell
receptor mediated signaling and labor, events which are
known to play key roles in pregnancy [91–94]. LINC0114 is
known to mediate cell differentiation and immune response,
which are important for pregnancy [95–97]. These associa-
tions with sPTB were observed only in maternal blood but
not in cord blood.
In another study on an US based African-American cohort
which analyzed genome-wide DNAm in maternal leukocytes,
two CpG sites were found to be significantly associated with
early PTB (24.1–34.0 weeks) [98]. Both of these sites
belonged to the regulatory-associated protein of MTOR
(RPTOR), component of a signaling pathway that regulates
cell growth in response to nutrient and insulin levels [99,
100]. This pathway has been previously implicated in
myometrial proliferation during pregnancy [101]. The study
also identified 5171 CpGs whose fetal methylation status
could be predicted from maternal signatures on those sites
(98.8% in the same direction). In an earlier study on a multi-
generational Caucasian cohort, about 75% of these sites were
found to be either single nucleotide polymorphisms (SNPs) or
methylation quantitative trait loci (meQTL) of multiple heri-
table traits, which suggested that most of these heritable CpG
sites might be under genetic influence [102].
Previously, genetic variants in matrix metalloprotease 1
gene (MMP1) were found to be associated with preterm pre-
mature rupture of the membranes (pPROM) [103]. In 2008, a
study reported association of promoter hypomethylation of a
SNP in MMP1 in fibroblast cells from the amnion of mothers
with pPROM, leading to sPTB [104].Thecombinedgenetic
and DNAm factors might determine MMP1 expression and
influence the association with pPROM. Such combinatorial
studies would help to identify birth outcome specific bio-
markers in pregnant women.
PTB is a complex phenotype of heterogeneous etiologies
[105]. In a recent study based on multi-omics approach in-
volving GWAS, RNA-Seq and Epigenome-wide association
(EWAS) using maternal blood collected after delivery,
Knijnenberg et al. compared full term birth (FTB, ≥37 to <
42 weeks) with very early PTB (VEPTB, < 28 weeks) and
early PTB (< 34 weeks) [106]. They found association of
RAB31 and RBPJ genes with PTB using all three platforms.
RAB31 is a member of the RAS oncogene family coding for a
small GTPase-binding protein, upregulated at full term com-
pared to midway through gestation which also has been im-
plicated in a GWAS on placental abruption [107,108]. RBPJ
is hypermethylated in VEPTB compared to FTB and is a
transcriptional regulator in the Notch signaling pathway
[109]. In addition to such studies, combining longitudinal epi-
genetic data with genetic data might help establish the causal
relationship of associated genes.
Investigation of temporal changes in DNAm coupled with
transcriptomic alterations in pregnant women, along with pre-
cise measurement of POG using ultrasound and metabolites,
might help to improve our understanding of PTB and hence
reduce health burdens associated with PTB outcomes.
Preeclampsia
Preeclampsia (PE) is a major cause of PTB and maternal and
infant mortality, characterized by hypertension and protein-
uria after 20 weeks of healthy pregnancy [110,111]. PE can
also lead to intrauterine growth restriction (IUGR) of fetus,
leading to PTB [112]. PE is associated with oxidative stress
and obesity, both of which are known to influence DNAm
[113–116]. Substantial research has been conducted on the
epigenetics of PE, primarily on the placenta and cell-free fetal
(cff) DNA in maternal blood [51,117–120]. Chim et al. ob-
served 5.7-fold higher abundance of unmethylated MASPIN
in maternal plasma from mothers with preeclampsia compared
to those with normal pregnancy outcome [51]. Tsui et al.
found elevated level of hypermethylated fraction of
Reprod. Sci.
RASSF1A gene in maternal plasma derived from cff DNA [120].
The abundance of cff DNA in maternal plasma of PE women are
most likely due to placental apoptosis, a characteristic of PE; or
due to reduced clearance of cff DNA from maternal system
[121–123]. RASSF1A is primarily a tumor suppressor, mostly
found to be inactivated by promoter hypermethylation in many
tumor types [124]. It is also found to be hypermethylated in
placenta in complicated pregnancies [125]. In a study conducted
on maternal peripheral blood, placenta, and umbilical cord blood,
promoter hypermethylation of soluble-cytoplasmic COMT (s-
COMT) was detected, whereas, it was found hypomethylated
in PE-specific placenta [56]. Promoter hypomethylation in the
placenta may indicate facilitated interaction of transcription factor
GATA-binding protein 2 (GATA2) and E1A-binding protein
p300 (EP300) with the S-COMT promoter which play a role in
placental development and the homeostasis of placental oxygen
tension [126]. The above tissue-specific methylation changes
might serve as biomarkers of early prediction of PE in pregnant
women.
In 2012, Mousa et al. reported hypomethylation of throm-
boxane synthase gene (TBXAS1) in systemic omental blood
vessels of mothers with PE [127]. TBXAS1 codes for an en-
zyme that catalyzes isomerization of prostaglandin H2 into
thromboxane [128]. Elevated thromboxane and decreased
prostacyclin acts as markers of PE [129,130].
Hypomethylation of TBXAS1 may contribute to the pathogen-
esis of PE and may increase maternal risk of cardiovascular
diseases later in life.
Anderson et al. 2013, identified 133 hypermethylated CpG
sites corresponding to 71 genes, out of 207 differentially meth-
ylated CpGs from peripheral WBCs during first trimester of
preeclamptic mothers, compared to normotensive (normal
blood pressure) mothers [131]. Seven of these belonged to the
pleckstrin homology domain, which although small in size, but
are present in a large variety of signaling proteins, serving as
lipid binding domains, and protease inhibitors. The remaining
74 CpGs from 38 genes were hypomethylated, and associated
with cellular metabolism, endothelin signaling, T cell activa-
tion, insulin signaling, progesterone-related oocyte maturation,
immune injury, phospholipase C-epsilon, G protein signaling,
encephalin release, and metabotropic glutamate receptor group
1 pathway. The same study identified thirteen differentially
methylated CpG sites as novel putative biomarkers of early
detection of PE (in the first trimester) in mothers [131]. These
genes play important roles in pregnancy, miscarriage, implan-
tation, and immune tolerance. KH homology domain and
stathmin family of genes play important roles in pregnancy
and implantation [132]. E2 class of ubiquitin conjugating en-
zyme has previously been implicated in early miscarriage,
CD80 in maternal immune tolerance to the fetus, RAP1A pro-
tein in GDM [133–135].
Longitudinal characterization of the PE phenotype (early-
onset PE, or late-onset PE) and its epigenomic underpinnings
is an area yet to be investigated in depth. Studies on identifi-
cation of epigenetic predisposition for PE in the maternal sys-
tem might enable early medical intervention during pregnancy
to reduce incidence of PE.
Limitations and Future Directions
We have summarized the studies in Table 1. The genes and
associated CpG sites along with their location (if discussed in
the study) are provided in Supplementary Table 1.Moststud-
ies on pregnancy were aimed to capture differences between
normal and extreme phenotypes. However, causal association
of genes with the reported outcomes are not always conclu-
sive. There is a paucity of understanding of the DNAm chang-
es that occur during the sequential events in pregnancy.
Although intrauterine growth restriction (IUGR) is an impor-
tant adverse condition of pregnancy, there has been no study
conducted on the maternal DNA methylation changes in
IUGR till date. However, multiple studies have implicated
maternal factors like hypertension, maternal substance abuse,
age, GDM, and anemia that can lead to this outcome [112,
136].
Multiple epidemiological and genetic risk factors are associ-
ated with pregnancy and its outcomes. Environmental, behav-
ioral, nutritional, and psycho-social conditions also modulate
pregnancy [137]. Hence, there is a burgeoning need to study
the genetic and epigenetic factors like DNAm from minimally
invasive maternal tissue like whole blood, which could help
monitor feto-maternal conditions during pregnancy and subse-
quently the health of the neonate. Blood provides a unique
potential window into the health and phenotype of the individ-
ual and hence longitudinal epigenetic profiling of peripheral
blood DNA from pregnant women during pregnancy and after
delivery might provide improved insights into the physiological
changes taking place. This might serve to identify potential
predictive biomarkers of pregnancy outcomes [138].
Studies have improved our understanding of early-life
stressors on women, especially during pregnancy.
Epigenome-wide studies across mother-offspring dyads are
required in the future to find the impact of maternal early-
life stress induced specific DNAm marks on pregnancy and
neonatal health. This approach would illuminate the impact of
early-lifepsycho-social factors on pregnancy and how it might
prospectively modulate the potential transmission of such
stressors on neonates.
There is a general limitation of most studies based on
whole blood DNAm. Whole blood is composed of multiple
cell types, and each cell type contributes to CpG locus specific
DNA methylation signal. Research on DNAm in whole
blood conducted until date has not yet fully addressed the
issue of cellular heterogeneity. Although various methods ex-
ist for adjusting for cellular heterogeneity, they commonly use
Reprod. Sci.
Table 1 Summary of the maternal DNA methylation studies discussed in the review
Study design Approach Phenotype (sample size) Time points Tissue Cell types Main findings References
Case-control Candidate
gene
Healthy pregnancy (32),
eclampsia, PE, HELLP
syndrome (17), healthy
controls (10), primary
breast cancer (26),
metastasized advance
breast cancer (10)
10–15 weeks Maternal sera - CDH1,PTGS2,APC,
RASSF1A behave
similar in pregnant
and cancer groups
Muller, 2004 (32)
Healthy pregnancy (20),
PE (10)
At delivery Maternal peripheral
blood, placenta
- Hypermethylated RASSF1A
were 4.3-fold higher in
maternal plasma of PE
subjects than in controls
Tsui, 2007 (116)
PPROM (284), healthy
pregnancy (361)
At delivery Amnion Fibroblast promoter hypomethylation
of MMP1 in amnion of
pPROM patients lead
to sPTB
Wang, 2008 (100)
Childhood maltreatment
(CM+) mother (58):
infant (55), CM mother
(59): infant(58)
After delivery Maternal peripheral
blood, umbilical
blood mononuclear
cells
Peripheral blood
mononuclear
cells (PBMC)
FKBP5,CRHR1:CM+
mothers show less
methylation than CM
mothers, NR3C1:CM+
mothers show more
methylation than CM
mothers
Ramo-Fernández (53)
Genome
wide
Healthy pregnancy (6),
gestational diabetes (6)
~16 weeks Maternal peripheral
blood
Peripheral blood
mononuclear
cells (PBMC)
Hypomethylated (NDUFC1,
HAPLN3,HHLA3,RHOG),
hypermethylated (SEPT11,
ZAR1,DDR)
Enquobahrie,
2015 (61)
Healthy pregnancy (8),
PE (8)
1st trimester,
3rd trimester
Maternal peripheral blood
Maternal plasma, placenta
- methylated MASPIN in
maternal plasma
Chim, 2005 (49)
Healthy pregnancy (14),
nulligravid (14)
7–15 weeks, 6–8
months postpartum
Maternal peripheral blood Leukocytes IL1R2,HPR,TREML1,
SPAG4,CCIN,PC,
NDUFS2,AGAP4/CTGLF1
hypomethylated in
pregnant state
White, 2012 (41)
Healthy pregnant women
(5), PE cases (7)
At delivery Maternal omental arteries - 65 genes were identified,
which showed reduced
methylation in preeclampsia
Mousa, 2012 (123)
Nulliparous (55), PE (6) 1st trimester, At
delivery
Maternal peripheral
blood, placental
chorionic tissue
White blood
cells
12 genes with significant
differences in mean
methylation in maternal
white blood cells,
putative biomarkers
Anderson, 2014 (127
GDM (11), healthy
pregnancy (11)
12–16 weeks Maternal peripheral
blood
-COPS8,PIK3R5,HAAO,
CCDC124,C5orf34 as
potential clinical
biomarkers
Wu, 2018 (73)
Reprod. Sci.
Table 1 (continued)
Study design Approach Phenotype (sample size) Time points Tissue Cell types Main findings References
Preterm delivery (16),
term delivery (24)
Time of admission
for labor
Maternal peripheral b
lood
Leukocytes 5171 CpGs maternal
methylation associated
with fetal methylation,
98.8% of which occurred
in the same direction
Parets, 2015 (94)
Discovery set: early sPTB
delivery (150), term
delivery (150); validation
set: (mother-child dyads)
early sPTBs (7 pairs), late
sPTBs (34 pairs), medically
indicated PTBs (14 pairs),
term birth (55 pairs)
24–72 h after delivery Maternal peripheral
blood, cord blood
White blood
cells
CYTIP,LINC00114
methylation lower in
mothers with early
sPTBs than TB
Hong, 2018 (86)
Healthy mother-child (8 pairs),
GDM mother-child (8 pairs)
Before delivery,
minutes after delivery
Maternal peripheral
blood, cord blood
- Differential methylation
SLC22A4,ADRA1A,
CACNB2,SERPINE1
Kang, 2017 (67)
Healthy pregnancy (21),
PE (16)
At delivery Maternal plasma,
placenta
- Placenta-specific s-COMT
promoter, a potential
marker for early
prediction of PE in
maternal plasma
Zhao, 2010 (53)
Observation,
cross-species
comparison
Candidate
gene
Healthy pregnancy (5),
pregnant mice, rhesus
monkeys
1st trimester, 3rd
trimester, at delivery
Maternal peripheral
blood, Placenta
-RASSF1A
hypermethylated in
placenta, unmethylated
in maternal blood cells
Chiu, 2007 (121)
Cross-sectional Candidate gene Healthy pregnancy (1160) 1st trimester, 2nd
trimester, At delivery
Maternal venous
blood
Venous cord blood
White blood
cells
LINE-1 elements
hypermethylation
Burris, 2012 (46)
Genome wide Healthy pregnancy (24) <12 weeks, at delivery Maternal peripheral
blood, cord blood
-PSMB7,B4GALT5
associated with DMG
concentrations in
cord blood
Knight, 2018 (79)
Longitudinal Genome wide Non-pregnant state, pregnant
state, postpartum (21)
10–14 weeks,
26–28 weeks,
2–4daysafter
delivery
Maternal peripheral
blood
- 91% of 196 significant
probes showed decrease
in methylation levels
across pregnancy.
Gruzieva, 2019 (47)
Multi-omics
integrative
study
Genome wide PTB (270 family trios), term
birth (521 family trios),
VEPTB, pPROM, PE,
uterine anomalies and
endometriosis, cervix
related, and idiopathic
PTB
1–4daysafter
delivery
Whole blood - RAB31 and RBPJ had
significant associations
found for all three data
types (WGS, DNA
methylation, and
mRNA expression)
Knijnenburg,
2019 (102)
Reprod. Sci.
a reference methylation data, based on which the sample cell
proportions are estimated [139]. Other bioinformatic tools for
reference-free cell type estimation mostly rely on prior infor-
mation on cell type-specific marks of specific cells from
existing database and infer cell proportion based on this infor-
mation [140,141]. These methods do not fully account for the
cell type diversity of study samples under question. DNAm-
based deconvolution of selected cell type-specific differential-
ly methylated regions (DMRs) from sorted and purified cell
populations of individual subjects are yet to be investigated.
Moreover, during pregnancy, maternal blood also contains
fetal genetic materials, and thus there remains a necessity to
account for both to exclusively parse the maternal component.
Studies have been conducted to investigate genetic as well as
other epigenetic processes such as histone modifications, non-
coding RNA, etc., on pregnancy and adverse pregnancy out-
comes [142–144]. Integration of such approaches with DNAm
studies would enable holistic understanding of pregnancy phe-
notype at the cellular and systemic level. Analysis of
biospecimens collected prior to conception, during gestation
and after delivery will help to identify unique DNAm changes
occurring specifically due to pregnancy. Prospective cohorts
should therefore be used to capture dynamic epigenetic modifi-
cations during pregnancy [145]. Since pregnancy-related compli-
cations owe its etiology to various factors such as gene-gene,
gene-environment interaction, etc. a multidimensional approach
towards pregnancy can address the effect-size of epistatic as well
as pleiotropic effects of genes [146]. A multi-omics approach
should help unravel the complex events of pregnancy which
leads to diverse outcomes. Such an approach could lead to re-
purposing of existing as well as discovery of novel therapeutic
agents, for clinical intervention in case of adversity. Designing a
prospective study cohort which captures relevant clinical, SES,
and demographic data would help to develop a holistic under-
standing of pregnancy. Cohorts like that of the Group for
Advanced Research on Birth Outcomes—DBT India Initiative
(GARBH-Ini) from India have taken into account the clinical,
SES, genomics, epigenomics, metagenomics, microbial and pro-
teomic correlates of pregnancy for integrated analysis [147].
Such longitudinal cohort studies can provide platforms which
can facilitate improved understanding of the pregnancy and its
outcomes. They may be better poised to address the epigenomics
of pregnancy, primarily DNAm landscape in mothers through
multiple time points during pregnancy and at postpartum, than
cross-sectional studies. Such strategy of following up mothers
would help identify pregnancy specific alterations, as well as
provide predictive markers for maternal health.
Conclusion
We have reviewed studies conducted on changes in maternal
DNA methylation during pregnancy and its associated
adverse conditions. Exploration of epigenomic changes can
provide improved understanding of the dynamic biological
processes that take place during pregnancy. This would allow
clinicians to identify women at increased risk for adverse
pregnancy outcomes and develop precise, individualized
risk-specific interventions.
Supplementary Information The online version contains supplementary
material available at https://doi.org/10.1007/s43032-020-00456-4.
Acknowledgments We thank Professor Partha Pratim Majumder for his
mentorship, advice and suggestions.
Funding JD is supported by the Research Fellowship (NET) of the
University Grants Commission (UGC), India.
Compliance with Ethical Standards
Conflict of Interest The authors declare that they have no conflict of
interest.
Ethical Consents This is a reviewarticle and does not involve collection
of human biospecimens or analysis. The review is based on published
information. Hence no ethical consent is required.
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