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Conditional system of controllable fertility through targeting implantation

定向着床可控生育的条件体系

基本信息

批准号：
17390453
负责人：
MARUYAMA Tetsuo
金额：
$ 10.05万
依托单位：
Keio University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2005
资助国家：
日本
起止时间：
2005 至 2006
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-17390453/
关键词：
implantation conditional knockout transgenic mouse doxycycline コンデイショナルノックアウト

项目摘要

The purpose of this study was to develop a conditional animal model whose fertility could be controlled through internal and external modification of implantation-associated genes. We employed two different approaches : 1) generation of genetically engineered mice (OPG-creTG) whose endometrial genes could be modulated spatially and temporally by doxycycline (DOX)- driven Cre-loxP system, and 2) introduction of DNA harboring the gene of interest into the uterine cavity by ultrasound exposure in the presence of microbubbles (sonoporation). Cre recombinase was successfully induced in the presence of DOX in the uterine tissue excised from OPG-creTG. Furthermore, OPG-creTG were crossed with the Rosa26R reporter mouse, which possessed a floxed repressor element associated with a lacZ transgene, in order to validate the functional efficacy of the conditionally expressed Cre. The results demonstrated that excision of the floxed element could be achieved specifically and conditionally in the endometrium, in particular, endometrial glandular cells; however, the conditions for maximal and reproducible induction of Cre remained to be improved and optimized. We found that genes could be introduced into the endometrium, in particular, luminal and glandular epithelium, much more efficiently by sonoporation than by liposome-based transfection. Thus, our animal model may be applicable as a conditional system of controllable fertility through targeting implantation, i.e., interaction between embryo and surface epithelium of the endometrium.

本研究的目的是建立一种条件动物模型，通过对着床相关基因的内部和外部修饰来控制其生育能力。我们采用了两种不同的方法：1)建立基因工程小鼠(OPG-creTG)，其子宫内膜基因可以通过多西环素(DOX)驱动的CRE-loxP系统在空间和时间上进行调节，以及2)在微泡存在的情况下，通过超声照射将携带有目的基因的DNA导入宫腔。在DOX存在下，OPG-CreTG切除的子宫组织中成功地诱导了Cre重组酶。此外，将OPG-creTG与Rosa26R报告小鼠杂交，以验证有条件表达的Cre的功能效果。结果表明，在子宫内膜，特别是子宫内膜腺细胞中，可以特异性和有条件地切除Cre，但最大限度和可重复性地诱导Cre的条件仍有待改进和优化。我们发现，基因可以通过超声导入子宫内膜，特别是腔上皮和腺上皮，比脂质体转染法更有效。因此，我们的动物模型可能适用于通过靶向植入，即胚胎与子宫内膜表面上皮之间的相互作用，来实现可控生育的条件系统。