Mdecular Mechanism of Oogenesis-Assisted Reproductive Technology and genomic imprinting-

为了解辅助生殖技术对人类早期发育过程中表观遗传学变化的影响，本研究以母体血浆和脐带血中游离DNA和RNA为主要研究对象，首先对3例父源性限制性胎盘嵌合体（父源性三体）进行CA重复序列分析。在所有三个病例中，在母体血浆中检测到父系传递的额外等位基因，但在胎儿中未检测到，这表明无细胞DNA来自胎盘（Masuzaki J Med Genetics 2004）。在阴道分娩的病例中，脐带血中母体DNA的检出率显著高于剖宫产病例，表明分娩增加了脐带血中母体DNA的污染（Masuzaki et al. Clin Chemi 2004）。采用真实的实时定量RT-PCR方法，检测孕妇血浆中两种mRNA随孕周的变化。然后，我们应用这项技术来评估胎盘状态的情况下，前置胎盘percreta（临床化学，出版）。在这个项目（15591761），我们增加了三个新的发现，我们的知识。以下三个新的发现表明，从胎盘表达的印迹基因是一个目标，以评估表观遗传状态。1)2）母体血浆中游离DNA来源于胎盘，2）存在双向输血，且至少受分娩的影响，3）母体血浆中胎盘mRNA可能是了解胎盘状态的一个靶点，从而为了解辅助生殖技术如何影响人类早期发育过程中的表观遗传变化提供了线索。

To know how assisted reproductive technology affects on the epigenetic change during early human development, we aimed at the cell free DNA and RNA in maternal plasma and cord blood as the key tool to resolve this question.First, three cases with paternal confined placental mosaicism (trisomy is from paternal origin) were analyzed by using CA repeats markers. In all three cases, paternal transmitted extra-alleles were detected in maternal plasma, but not in fetus, suggesting that cell free DNA is from placenta (Masuzaki J Med Genetics 2004).Also, we detected maternal DNA in cord blood using 8 highly polymorphic markers in Japanese. Detection rate of maternal DNA in cord blood was significantly higher in cases with vaginal delivery than in cases with Cesarean Section, indicating that labor increases maternal DNA contamination in cord blood (Masuzaki et al. Clin Chemi 2004).Second, hCG and hPL genes were selected as genes expressed from placenta. By using real time quantitative RT-PCR, changing concentration of both mRNA in maternal plasma was measured as the progress of gestational age. And then, we applied this technique to the evaluation of placental status in a case with placenta previa-percreta (Clin Chemi, in press).In this project(15591761), we add three novel findings to our knowledge. Three novel findings are below and suggested that imprinting gene expressed from placenta is a target to evaluate the epigenetic status. 1)Cell free DNA in maternal plasma is from placenta, 2)Bidirectional blood transfusion exists and this is affected by labor at least, and 3)there is the possibility that placental mRNA in maternal plasma is a target to know the placental status.Therefore, we got a clue to know how assisted reproductive technology affects on the epigenetic change during early human development.