孤雌胚胎干细胞获得四倍体补偿能力的机制研究

Parthenogenetic embryonic stem cells (pESCs) derived from chemical activated oocytes are one kind of multiple pluripotent, abundant and novel stem cells, which show great values in regenerative medicine research and gametes carrying gene imprinting regulation research. However, pESCs had limited differentiation potential in some specific lineages due to bi-maternal imprinting comparing to zygote-derived ESCs, which made them can not develop to term by tetraploid complementation. To figure out the mechanism underlying limited plripotency of pESCs is quite important to push forward application of stem cells and better understand mechanism of development of stem cells. In our study, we introduce single allele H19 and Gtl2 knock-out in pESCs to advance pluripotency of them. This project will prove that pESCs can support tetraploid complementation development with some gene modification and clarify the mechanism behind this phenotype, which would supply theory support and animal models for epigenetic regulation research and application of pESCs.

从卵母细胞孤雌激活胚胎建立的孤雌胚胎干细胞是一种既具有较高的分化潜能，又具有来源方便、资源充足等优势的新型多能干细胞，因此在再生医学应用中具有重要价值，同时也是研究配子携带的基因印记修饰的重要工具。然而，研究显示相比受精卵来源的胚胎干细胞，孤雌胚胎干细胞向特定胚层的发育能力有限，更无法独立发育成一个完整个体(四倍体补偿能力)。解析孤雌胚胎干细胞发育受限机制，拓展其发育能力，对孤雌胚胎干细胞的转化应用和干细胞发育机制解析均有重要意义。本研究在基因组水平上对孤雌胚胎干细胞中的两个重要印记基因H19和Gtl2进行敲除，预期获得具有四倍体补偿能力的孤雌胚胎干细胞，并在分子生物学水平上阐释其多能性提高的分子机制，从而为印记基因功能调控研究和推动孤雌干细胞的应用提供理论基础和实验模型。

研究显示相比，孤雌胚胎干细胞与受精卵来源的胚胎干细胞相比，其向特定胚层的发育能力有限，更无法独立发育成一个完整个体(四倍体补偿能力)。解析孤雌胚胎干细胞发育受限机制，拓展其发育能力，对孤雌胚胎干细胞的转化应用和干细胞发育机制解析均有重要意义。本研究在基因组水平上对孤雌胚胎干细胞中的两个重要印记基因H19和Gtl2进行了敲除，预期获得具有四倍体补偿能力的孤雌胚胎干细胞，并在分子生物学水平上阐释其多能性提高的分子机制，从而为印记基因功能调控研究和推动孤雌干细胞的应用提供理论基础和实验模型。目前取得的成果为1. 获得了小鼠孤雌胚胎干细胞；2. 构建了印记基因H19的同源重组敲除质粒；3. 获得了H19-/-基因型的pESCs细胞系。