背景:胎儿性别和胎盘发育影响妊娠结局和胎母健康,但是在人类妊娠中,对妊娠早期胎盘建立的关键时间点的研究不足。
方法:在绒毛膜绒毛取样的早期妊娠晚期(第10-14周)招募孕妇,产前诊断测试.收集并储存剩余胎盘组织,直到知道出生结果,然后从单例中分离DNA和RNA,正常核型妊娠导致活产。用IlluminaInfinium甲基化EPICBeadChip阵列测量DNA甲基化(n=56)。差异甲基化分析使用743,461个常染色体探针的广义线性模型比较了25名女性和31名男性。用RNA测序分析基因表达的性别差异(n=74)。使用线性回归进行整合分析,以关联51个重叠胎盘中的基因表达和DNA甲基化。
结果:甲基化分析鉴定出151种差异甲基化探针(DMPs)在错误发现率<0.05时具有显著性,包括89(59%)女性高甲基化。探头cg17612569(GABPA,ATP5J)是最显著的CpG位点,在男性中高度甲基化。有11个差异甲基化区域受胎儿性别影响,转录因子ZNF300和ZNF311在男性和女性中最显著的高甲基化,分别。RNA测序在错误发现率<0.05的情况下鉴定出152个明显的性别二态基因。151个DMPs与18个基因呈高甲基化方向下调(P<0.05)相关,包括2个显著错误发现率<0.05的基因(ZNF300和CUB和Sushi多域1,CSMD1)。这两个基因,以及具有序列相似性的家族228成员A(FAM228A),DNA甲基化与性二态基因表达显著相关,尽管FAM228ADNA甲基化性二态较少。与其他性别差异研究的比较发现,cg17612569在整个妊娠期间在胎盘和人类血液中直至成年期都是男性高甲基化的。
结论:总体而言,性别二态差异甲基化与妊娠早期胎盘中相关的差异基因表达较小,但仍有一些重要的基因可能通过甲基化调节,导致孕早期胎盘的差异。
胎儿性别和胎盘发育影响胎儿和母亲在整个妊娠期间的妊娠结局。包括流产的风险,早产,先兆子痫,和其他结果。表观遗传学,调节信号在DNA上的“叠加”影响DNA的读取方式,在怀孕早期,当发生影响怀孕其余部分的关键胎盘发育时,我们并没有很好地理解。这里,我们使用Cedars-Sinai患者在知情同意的情况下捐赠的剩余胎盘活检样本(n=56),以了解由于胎儿性别导致的孕早期人类胎盘DNA甲基化差异.在分析的743,461个站点中,我们在校正p值以减少假阳性(假发现率<0.05)后,确定了151个受胎儿性别显著影响的部位.我们还进行了分析以查看多个位点,并确定了基因组中的11个区域,这些区域由于胎儿性别而发生了显着的DNA甲基化变化。此外,因为DNA甲基化是DNA上的调节标记,通常会抑制基因表达,我们还比较了DNA甲基化的性别差异与胎盘RNA测序基因表达分析,使用的组织来自一个大部分重叠的患者组(n=74总测序,n=51重叠)。我们鉴定了18个基因,它们显示出显著的DNA甲基化差异和基因表达变化。最重要的基因是转录因子ZNF300,在男性中具有较高的DNA甲基化,在男性中具有较高的基因表达(因此在女性中具有较高的基因表达)。这项研究确定了一些性别差异,这些差异一直持续到怀孕后期,而其他性别差异是妊娠早期所独有的。
BACKGROUND: Fetal sex and placental development impact pregnancy outcomes and fetal-maternal health, but the critical timepoint of placenta establishment in first trimester is understudied in human pregnancies.
METHODS: Pregnant subjects were recruited in late first trimester (weeks 10-14) at time of chorionic villus sampling, a prenatal diagnostic test. Leftover placenta tissue was collected and stored until birth outcomes were known, then DNA and RNA were isolated from singleton, normal karyotype pregnancies resulting in live births. DNA methylation was measured with the Illumina Infinium MethylationEPIC BeadChip array (n = 56). Differential methylation analysis compared 25 females versus 31 males using a generalized linear model on 743,461 autosomal probes. Gene expression sex differences were analyzed with RNA-sequencing (n = 74). An integrated analysis was performed using linear regression to correlate gene expression and DNA methylation in 51 overlapping placentas.
RESULTS: Methylation analysis identified 151 differentially methylated probes (DMPs) significant at false discovery rate < 0.05, including 89 (59%) hypermethylated in females. Probe cg17612569 (GABPA, ATP5J) was the most significant CpG site, hypermethylated in males. There were 11 differentially methylated regions affected by fetal sex, with transcription factors ZNF300 and ZNF311 most significantly hypermethylated in males and females, respectively. RNA-sequencing identified 152 genes significantly sexually dimorphic at false discovery rate < 0.05. The 151 DMPs were associated with 18 genes with gene downregulation (P < 0.05) in the direction of hypermethylation, including 2 genes significant at false discovery rate < 0.05 (ZNF300 and CUB and Sushi multiple domains 1, CSMD1). Both genes, as well as Family With Sequence Similarity 228 Member A (FAM228A), showed significant correlation between DNA methylation and sexually dimorphic gene expression, though FAM228A DNA methylation was less sexually dimorphic. Comparison with other sex differences studies found that cg17612569 is male-hypermethylated across gestation in placenta and in human blood up to adulthood.
CONCLUSIONS: Overall, sex dimorphic differential methylation with associated differential gene expression in the first trimester placenta is small, but there remain significant genes that may be regulated through methylation leading to differences in the first trimester placenta.
Fetal sex and placenta development affect pregnancy outcomes for both the fetus and mother throughout pregnancy, including risk of miscarriages, preterm birth, preeclampsia, and other outcomes. Epigenetics, the “overlay” of regulatory signals on DNA which affects how DNA is read, is not well understood in early pregnancy when critical placenta developments are happening that affect the rest of pregnancy. Here, we use leftover placenta biopsy samples (n = 56) donated by Cedars-Sinai patients with informed consent to learn about first trimester human placenta DNA methylation differences due to fetal sex. Out of the total 743,461 sites analyzed, we identified 151 sites significantly affected by fetal sex after correcting p-values to reduce false positives (false discovery rate < 0.05). We also performed an analysis to look at multiple sites and identified 11 regions across the genome with significant DNA methylation changes due to fetal sex. Furthermore, because DNA methylation is a regulatory mark on DNA which typically dampens gene expression, we also compared the DNA methylation sex differences to placental RNA-sequencing gene expression analysis using the same tissue from a mostly overlapping patient group (n = 74 total sequenced, n = 51 overlap). We identify 18 genes which show both significant DNA methylation differences and gene expression changes. The most significant gene was transcription factor ZNF300 with higher DNA methylation in males and reduced gene expression in males (and thus higher gene expression in females). This study identifies some sex differences that continue until later pregnancy and others that are unique to first trimester.