母血浆非性别依赖胎儿游离DNA对早期预测子痫前期价值的研究

Preeclampsia is one of the major causes of maternal and perinatal morbidity and mortality. Effective screening for early detection of pregnancies that are destined to experience preeclampsia is important to estimate their proximity to onset of the clinical disease, and would permit medical care providers to plan a more detailed course of action. Currently, a wide variety of markers have been proposed to predict preeclampsia, however, most of them do not have highly enough sensitivity and specificity. Several studies show that the quantitative changes of cell-free fetal DNA (cffDNA) in maternal plasma, such as elevated levels of cffDNA during first trimester of pregnancies and slow clearance of cffDNA after deliveries, can be used as an effective indicator for impending preeclampsia. We propose to continuously measure the concentrations of cffDNA from 40 normal pregnancies and 20 cases with preeclampsia using the next-generation sequencing (NGS) platform. From NGS data, we could accurately demonstrate the changing patterns of cffDNA concentration levels in different gestational age from the plasma of normal pregnancies and preeclampsia patients. Also, we hope to examine the cffDNA genomic coverage and gene content differences between the case and control samples using the NGS data, which may lead to the further understanding of the cffDNA metabolism and the possible function impact of cffDNA and preeclampsia in the molecular level.

子痫前期是最严重的妊娠并发症之一，是导致孕产妇和围产儿死亡的最主要原因之一。对子痫前期早期预测并早期干预可减少或避免不良母婴结局。目前子痫前期的预测方法多种，但检测的特异性及灵敏度均不是很高。研究发现，子痫前期孕妇血浆胎儿游离DNA(cffDNA)在早孕期含量增加及产后清除率降低，提示母血中cffDNA浓度可用来预测子痫前期的发生及预后。本研究连续采集40例正常孕妇和20例子痫前期患者不同孕周外周血，通过第二代测序平台建立正常妊娠和子痫前期孕妇外周血cffDNA含量动态变化曲线。通过生物信息学分析，寻找cffDNA在子痫前期患者外周血浆浓度上升过程中在人类全基因组的分布规律，寻找潜在的cffDNA富集区域，探索可能与子痫前期发病相关的基因特异性。通过对比子痫前期患者产后血浆中留存cffDNA的基因位置，研究cffDNA清除速率降低的可能分子通路机制，评价cffDNA对子痫前期预测价值。

子痫前期作为严重的产科并发症，可导致胎儿生长受限、医源性早产等很多异常。目前对子痫前期尚缺乏有效的早期诊断和干预措施，且其病因和发病机制尚不明确。为评价孕妇外周血胎儿游离DNA（cffDNA）含量的变化情况对子痫前期的发病的预测价值，我们首先利用收集的正常孕妇血浆样本采用覆盖全基因组的高频SNP位点进行cSMART建库测序，开发建立了基于高通量测序但不依赖胎儿性别的精准cffDNA含量测定技术平台，该技术可保证cffDNA含量这一重要指标在无创产前诊断方面的精确应用。对收集的不同孕周、围产期的正常孕妇和子痫前期孕妇的血浆样本，采用本项目开发的基于SNP-cSMART测序的cffDNA含量测定技术对比分析了正常孕妇和子痫前期孕妇的cffDNA含量随孕周的变化曲线，结果暗示cffDNA含量指标目前可能并不适用于预测子痫前期的发生和发展。为了进一步探索子痫前期发病的分子机制以及寻找可能的早期诊断依据，对孕妇胎盘的DNA甲基化水平进行了研究，发现子痫前期孕妇的胚胎DNA较正常孕妇呈低甲基化变化。子痫前期胚胎DNA甲基化水平的变化为指导该疾病的早期诊断和治疗在分子、生化代谢方面提供了可能的方向。