蜕膜间质细胞旁分泌的EGF通过诱导上皮间质转化促进滋养细胞侵袭和迁移的研究

The trophoblasts invading into the maternal endometrium is the key point to establish the process of pregnancy and sustainable embryo development, which is also one of the hot research area of reproductive medicine. Our previous studies found that, compared to the endometrial stromal cells, decidual stromal cells(DSC) promoted trophoblast invasion and migration. Further studies demonstrated that thophoblastic epithelial mesenchymal transition (EMT)may involve in the process and EGF may be the active cytokines secreted by DSC in regulation of trophoblastic EMT. This project is planned to investigate the effect of DSC on trophoblast invasion and migration induced by EMT and the effect of EGF secreted by DSC on trophoblast EMT and its intracellular pathways. This results will help to further understand the molecular dialogue between DSC and trophoblasts, especially the mechanism of DSC inducing EMT throught secretion of EGF to promote trophoblast invasion and migration, which is of great significance to completely illustrate the cell and molecular activity at the maternal-fetal interface during the process of embryo implantation and maintenance of pregnancy.

胚胎植入子宫内膜是妊娠建立的初始过程和胚胎持续发育的关键，也是生殖医学研究的热点。在前期工作中我们发现与子宫内膜间质细胞相比，早孕期蜕膜间质细胞条件培养基能显著促进滋养细胞的侵袭和迁移。进一步的结果还提示滋养细胞上皮间质转化（EMT）可能参与此过程，而EGF可能是蜕膜间质细胞分泌的参与调节滋养细胞EMT转化的活性细胞因子。本项目通过对蜕膜间质细胞诱导滋养细胞EMT转化对细胞侵袭和迁移的影响及旁分泌EGF调节细胞内通路激活影响滋养细胞EMT的胞内分子调节过程及机制的研究，进一步了解滋养细胞与蜕膜细胞之间的分子对话，尤其是蜕膜间质细胞通过分泌EGF诱导滋养细胞发生EMT转化的作用机制，对完整阐述胚胎植入和妊娠维持过程中母胎界面细胞和分子生物学活动具有重要意义。

胚胎植入子宫内膜是妊娠建立的初始过程和胚胎持续发育的关键。本项目在前期工作的基础上，对蜕膜间质细胞诱导滋养细胞EMT转化对细胞侵袭和迁移的影响及旁分泌EGF调节细胞内通路激活影响滋养细胞EMT的胞内分子调节过程及机制进行研究。我们在体外分离培养子宫内膜间质细胞和蜕膜间质细胞的原代培养细胞系，发现蜕膜细胞条件培养基DSC-CM和内膜细胞条件培养基EMC-CM显著增强滋养细胞的迁移能力，而DSC-CM培养基处理后，滋养细胞迁移能力的增加更为明显。DSC-CM处理滋养细胞 JEG3 后，细胞由连接紧密的卵圆形转变为分布松散的梭形。免疫荧光结果显示：JAR细胞系cytokeratin表达阳性，CM或EGF处理后其表达下降；HTR8细胞系vimentin表达阳性，CM或EGF处理后其表达升高，这提示蜕膜间质细胞条件培养基和EGF在促进滋养细胞系上皮间质转化中起重要作用。由于蜕膜间质细胞在体外传代培养过程中发生分化，其蜕膜化Marker，如IGFBP1、PRL表达降低。为了更直观地研究内膜发生蜕膜化改变，观察其旁分泌作用对滋养层细胞的影响，我们建立了体外蜕膜化培养体系，并发现Deci EMC-CM可能通过激活Erk通路介导EMT marker的改变。外源性EGF处理滋养层细胞Jeg-3后，Erk信号通路被激活，EMT 上皮marker E-cadherin表达下降、vimentin表达上升，EMT转录因子Snail表达上升，提示EGF能促进滋养细胞发生EMT，其作用对与体外蜕膜化内膜细胞条件培养基对滋养细胞的作用相似，提示蜕膜化的内膜间质细胞可能通过旁分泌的作用激活EGF下游的ERK通路，从而增加滋养细胞的迁移与EMT的发生。