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胚性幹細胞の生殖細胞分化と核の初期化

生殖细胞分化和胚胎干细胞的核重编程

基本信息

批准号：
18051020
负责人：
野瀬俊明
金额：
$ 21.95万
依托单位：
Mitsubishi Kagaku Institute of Life Sciences
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research on Priority Areas
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

项目摘要

1.ES細胞の長期分化培養から選別した増殖性生殖細胞は,生体内の精子幹細胞に特異的な遺伝子発現を示すほか,分化条件下で卵形成や精細胞形質の発現を示す。この培養分画からさらに精子幹細胞を精製し,幹細胞特性を固定する培養条件の検討を進めた。ES細胞由来Vasa発現細胞にはc-kit発現において陽陰両方の細胞が混在する。このうち,より未分化性を反映するc-kit陰性の細胞を分画して幹細胞培地を用いた継続培養を行うことによって,Vasa陽性Oct4陰性の培養株が得られた。この細胞株は,生殖幹細胞の指標となるPlzf蛋白発現が確認された。さらに,この幹細胞様細胞のin vitro精子形成分化の可能性を検討した結果,培地から幹細胞維持因子を除去することによって,数日間の短い期間に様々な減数分裂形質さらに精子細胞形質の発現が誘起されることが判明した。2.受精時およびクローン核移植時に発現亢進されることが判明したVasa遺伝子は,その発現制御と初期化現象との間に密接な関連性が推定された。DNA(CpG)メチル化修飾の解析からは,Vasaプロモーター域はin vivo,in vitroの生殖細胞分化に共通して,特有のメチル化修飾とそれに続く脱メチル化のプロセスを経過することが判明した。また,ES細胞からのin vitro分化系によって均一なVasa発現細胞の調製が得られたことから,Vasa発現の有無によるクロマチン修飾変動の解析が可能となった。その解析結果はH3-K79の2,3メチル化の他,幾つかの修飾がVasaプロモーター活性に相関していることを示した。

In the culture of long-term differentiation of 1.ES cells, gonadal reproductive cells were selected, the spermatozoa of spermatozoa were specific for differentiation, and the spermatozoa were formed under the condition of differentiation. The sperm, the cell, the sperm, the cell, the cell The origin of the ES cells Vasa shows that the cells are mixed with each other in the c-kit. "undifferentiated" reflects "c-kit", "cell", "cell culture", "cell culture", "Vasa", "undifferentiated", "undifferentiated" and "undifferentiated". The expression of Plzf protein was confirmed by cell line and reproductive cell. The possibility of spermatogenesis and differentiation of in vitro cells were analyzed. The cell maintenance factors of culture cells were removed. Within a few days, the cells were divided into two short periods, and the sperm cells were divided into two groups. two。 During the insemination test, during the nuclear transfer, it was found that the Vasa was detected, and the system was designed to make the system more sensitive to the presumption of connectivity. DNA (CpG) chemical modification is used to analyze the differentiation of reproductive cells in the domain of Vasa. The reproductive cells of in vivo,in vitro are differentiated in a common way, which is specific to the differentiation of reproductive cells. In the in vitro differentiation system of ES, the whole system of differentiation has been detected in the same Vasa. In addition, the Vasa has shown that it is possible to improve the performance of the system. The results of H3-K79 analysis showed that he was changed in 3 minutes, and the activity of the Vasa was modified.