Marjolein M van Vliet, Ruben G Boers, Joachim B Boers, Olivier J M Schäffers, Lotte E van der Meeren, Régine P M Steegers-Theunissen, Joost Gribnau, Sam Schoenmakers
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We therefore aim to investigate the feasibility of genome-wide methylation profiling of first trimester maternal cfDNA using MeD-seq, by identifying placental-specific methylation marks in cfDNA.</p><p><strong>Methods: </strong>We collected cfDNA from nonpregnant controls (female n = 6, male n = 12) and pregnant women (n = 10), first trimester placentas (n = 10), and paired preconceptional and first trimester buffy coats (total n = 20). Differentially methylated regions (DMRs) were identified between pregnant and nonpregnant women. We investigated placental-specific markers in maternal cfDNA, including RASSF1 promoter and Y-chromosomal methylation, and studied overlap with placental and buffy coat DNA methylation.</p><p><strong>Results: </strong>We identified 436 DMRs between cfDNA from pregnant and nonpregnant women, which were validated using male cfDNA. RASSF1 promoter methylation was higher in maternal cfDNA (fold change 2.87, unpaired t-test p < .0001). Differential methylation of Y-chromosomal sequences could determine fetal sex. DMRs in maternal cfDNA showed large overlap with DNA methylation of these regions in placentas and buffy coats. Sixteen DMRs in maternal cfDNA were specifically found only in placentas. These novel potential placental-specific DMRs were more prominent than RASSF1.</p><p><strong>Conclusions: </strong>MeD-seq can detect (novel) genome-wide placental DNA methylation marks and determine fetal sex in maternal cfDNA. Our results indicate a placental and immune-cell contribution to the pregnancy-specific cfDNA methylation signature. 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引用次数: 0
摘要
背景：胎盘源性无细胞DNA（cfDNA）为远程研究（外）遗传胎盘编程提供了独特的机会，但研究cfDNA甲基组的研究很少，而且通常在技术上具有挑战性。甲基化DNA测序（MeD-seq）与低浓度的cfDNA有很好的兼容性，而且具有很高的全基因组覆盖率。因此，我们旨在研究利用 MeD-seq 对妊娠头三个月的母体 cfDNA 进行全基因组甲基化分析的可行性，方法是识别 cfDNA 中胎盘特异性的甲基化标记：我们收集了非妊娠对照组（女性 n = 6，男性 n = 12）和孕妇（n = 10）的 cfDNA、头三个月胎盘（n = 10）以及配对的孕前和头三个月水包衣（共 n = 20）。确定了孕妇和非孕妇之间的差异甲基化区域（DMRs）。我们研究了母体 cfDNA 中的胎盘特异性标记，包括 RASSF1 启动子和 Y 染色体甲基化，并研究了与胎盘和水包衣 DNA 甲基化的重叠情况：我们在孕妇和非孕妇的cfDNA中发现了436个DMRs，并用男性cfDNA进行了验证。母体 cfDNA 中 RASSF1 启动子甲基化程度较高（折叠变化 2.87，非配对 t 检验 p 结论：MeD-seq 可以检测（新的）妊娠和非妊娠妇女中的 RASSF1 启动子甲基化：MeD-seq 可检测（新的）全基因组胎盘 DNA 甲基化标记，并确定母体 cfDNA 中的胎儿性别。我们的研究结果表明，胎盘和免疫细胞对妊娠特异性 cfDNA 甲基化特征有贡献。这项研究支持未来对母体 cfDNA 甲基化的研究。
Genome-wide methylation profiling of maternal cell-free DNA using methylated DNA sequencing (MeD-seq) indicates a placental and immune-cell signature.
Background: Placental-originated cell-free DNA (cfDNA) provides unique opportunities to study (epi)genetic placental programming remotely, but studies investigating the cfDNA methylome are scarce and usually technologically challenging. Methylated DNA sequencing (MeD-seq) is well compatible with low cfDNA concentrations and has a high genome-wide coverage. We therefore aim to investigate the feasibility of genome-wide methylation profiling of first trimester maternal cfDNA using MeD-seq, by identifying placental-specific methylation marks in cfDNA.
Methods: We collected cfDNA from nonpregnant controls (female n = 6, male n = 12) and pregnant women (n = 10), first trimester placentas (n = 10), and paired preconceptional and first trimester buffy coats (total n = 20). Differentially methylated regions (DMRs) were identified between pregnant and nonpregnant women. We investigated placental-specific markers in maternal cfDNA, including RASSF1 promoter and Y-chromosomal methylation, and studied overlap with placental and buffy coat DNA methylation.
Results: We identified 436 DMRs between cfDNA from pregnant and nonpregnant women, which were validated using male cfDNA. RASSF1 promoter methylation was higher in maternal cfDNA (fold change 2.87, unpaired t-test p < .0001). Differential methylation of Y-chromosomal sequences could determine fetal sex. DMRs in maternal cfDNA showed large overlap with DNA methylation of these regions in placentas and buffy coats. Sixteen DMRs in maternal cfDNA were specifically found only in placentas. These novel potential placental-specific DMRs were more prominent than RASSF1.
Conclusions: MeD-seq can detect (novel) genome-wide placental DNA methylation marks and determine fetal sex in maternal cfDNA. Our results indicate a placental and immune-cell contribution to the pregnancy-specific cfDNA methylation signature. This study supports future research into maternal cfDNA methylation.
期刊介绍：
EJCI considers any original contribution from the most sophisticated basic molecular sciences to applied clinical and translational research and evidence-based medicine across a broad range of subspecialties. The EJCI publishes reports of high-quality research that pertain to the genetic, molecular, cellular, or physiological basis of human biology and disease, as well as research that addresses prevalence, diagnosis, course, treatment, and prevention of disease. We are primarily interested in studies directly pertinent to humans, but submission of robust in vitro and animal work is also encouraged. Interdisciplinary work and research using innovative methods and combinations of laboratory, clinical, and epidemiological methodologies and techniques is of great interest to the journal. Several categories of manuscripts (for detailed description see below) are considered: editorials, original articles (also including randomized clinical trials, systematic reviews and meta-analyses), reviews (narrative reviews), opinion articles (including debates, perspectives and commentaries); and letters to the Editor.
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