项目名称： 核移植诱导体细胞重编程中供核细胞分化状态的影响

项目编号： No.30871430

项目类型： 面上项目

立项/批准年度： 2009

项目学科： 电工技术

项目作者： 李劲松

作者单位： 中国科学院上海生命科学研究院

项目金额： 33万元

中文摘要： 核移植诱导体细胞重编程具有重要的应用前景，然而，克隆胚胎发育率低、重编程机制不清楚。本研究中，利用不同的供体细胞研究重编程，共发表研究论文6篇（通讯及共同通讯作者论文5篇），撰写应邀综述1篇。已取得的主要前沿性研究进展：揭示母源因子 Tet3 参与了受精后雄原核及核移植后体细胞核移植的主动去甲基化过程，Tet3 的缺失会导致胚胎发育的异常（Nature, 2011）,这表明动物克隆和自然受精过程很可能采用了同样的重编程机制，为提高动物克隆效率带来了新的理论依据；确证克隆囊胚的滋养外胚层细胞的异常是克隆胚胎发育失败的关键原因，通过修复异常使克隆动物出生率提高了 6 倍(Cell Stem Cell, 2011)，这一成果为核移植的基础研究提供了重要信息；发现核移植技术具有比 iPS 技术更好的重编程能力(J Mol Cell Biol, 2011)；证明两细胞卵裂球具有重编程雄性配子细胞的能力(Cell Res, 2011)；发现体细胞核能在未去核卵母细胞中高效重编程(Cell Res, 2010)。这些研究成果有利于核移植诱导体细胞重编程方法的发展和完善。

中文关键词： 核移植;克隆胚胎;体细胞重编程;供体细胞

英文摘要： Nuclear transfer (NT) has offerred exciting promises in basic and applied research, however, the developmental efficiency of cloned embryos remain disppointingly low and the mechanism of reprogramming induced by NT is not clear. In this project, we studied NT using different donor cells and made the following progresses. The first technology breakthrough in my lab was the establishment of a simple and highly efficient reprogramming procedure , in which, intact MII oocytes are used as recipient instead of enucleated oocytes in mouse somatic NT. In the same time, my lab also demonstrated that the remodeling ability persists in the blastomeres of two-cell embryos . We found that two-cell blastomeres can partially remodel mature sperm, but can completely remodel round spermatids. More recently, my lab had made another major strive to demonstrate that defects in trophoblast cell lineage account for the impaired in vivo development of cloned embryos generated by somatic nuclear transfer . In this study, we separated the ICMs from cloned blastocysts and aggregated the cloned ICM with two fertilization-derived (FD) tetraploid (4N) embryos. We found that the full-term development of cloned ICMs was dramatically improved after the trophoblast cells in the cloned blastocysts were replaced by cells from tetraploid embryos, thus providing direct evidence that defects in trophoblast cell lineage underlie the low success rate of somatic NT. Furthermore, we, cooperated with Dr. Guoliang Xu's lab, identifed that Tet 3 is involved in DNA active-demethylaiton in male pronucleus and pseudo-pronucleus in oocyte respectively formed from sperm and injected somatic nucleus . These results suggest that natural fertilization and somatic reprogramming induced by NT may share same molecular mechanisms. In a long-term project, my lab would like to investigate the mechanisms involved in NT and iPS technology by comparing the reprogrammed cells and reprogramming processes in different strategies. In a preliminary study, we established a comparative system, in which donor cells with genetically identical background were used for different reprogramming strategies, and found that reprogramming method per se determines the pluripotency of resulting cells . My future research will prococeed to the next stage, a more exciting stage in which I focus primarily on applying the system to reveal important reprogramming factors and to establish high-quality reprogramming strategies.

英文关键词： Nuclear transfer; cloned embryos; somatic reprogramming; donor cells