Conditional system of controllable fertility through targeting implantation

本研究的目的是建立一种能够通过植入相关基因的内外修饰来控制其生育能力的条件动物模型。我们采用了两种不同的方法：1)产生基因工程小鼠（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；2)在微泡存在的情况下，通过超声暴露将含有感兴趣基因的DNA引入子宫腔（超声穿孔）。在DOX存在的情况下，成功地在OPG-creTG切除的子宫组织中诱导了Cre重组酶。此外，为了验证条件表达的Cre的功能有效性，将OPG-creTG与Rosa26R报告小鼠进行杂交，Rosa26R报告小鼠具有与lacZ转基因相关的floxed阻遏元件。结果表明，在子宫内膜，特别是子宫内膜腺细胞中，可以特异性地、有条件地切除含氟元素；然而，最大限度和可重复性地诱导Cre的条件仍有待改进和优化。我们发现基因可以被引入子宫内膜，特别是腔上皮和腺上皮，比脂质体转染更有效。因此，我们的动物模型可能适用于通过靶向着床（即胚胎与子宫内膜表面上皮相互作用）实现可控生育的条件系统。

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.