人子宫内膜上皮细胞来源iPS细胞定向分化用于子宫内膜再生的效应和机制研究

Damage caused by mechanical force or infection to the basalis layer would disturb the self-repair of the endometrium, and subsequently cause fibrosis, scar formation, menstrual disorder, uterine adherence and infertility, which could hardly be cured. Tissue engineering is a promising new medical treatment.on the repair of damaged endometrium, however there is a lack of seed cells. Thus, this project aims at directed differentiation of induced pluripotent stem cells (iPS) into human endometrial epithelial progenitor stem cells and their application in the repair of mice model of endometrial injury. By using iPS technology, autologous somatic cells could be reprogrammed and subsequently re-differentiated into endometrial progenitor cells. Combining the endometrial progenitor cells as seed cells with silk fibroin microspheres as tissue engineering scaffolds so as to coat vascular endothelial growth factor (VEGF) and guide the growth and differentiation of seed cells, tissue engineering treatment on the repair of endometrial damage would be investigated. By conducting this project, developmental model of endometrial epithelia would be setup, and systematic investigation of the molecular regulatory network during the development of the endometrial epithelia could be viable, autologous cell based tissue engineering treatment on the repair of endometrial damage could also be developed.

由机械或者感染等原因引起的子宫内膜基底层损伤，将导致子宫内膜纤维化、斑痕化、宫腔粘连及不孕，治疗困难。利用组织工程技术修复受损子宫内膜是一种非常具有前景的新手段，然而种子细胞来源困难，因此本课题将人子宫内膜上皮细胞来源诱导多能干细胞（iPS）细胞定向分化用于子宫内膜再生的效应和机制研究。利用iPS技术将自体子宫内膜上皮细胞重编程后再定向诱导分化成为人子宫内膜上皮前体/干细胞作为种子细胞，研究子宫内膜上皮干细胞的分化条件和机制，并用丝素蛋白微球作为组织工程支架包被血管内皮生长因子(VEGF)作为载体，用于体内研究修复子宫内膜损伤的效应。本项目发现将建立子宫内膜上皮细胞发育过程各个不同阶段模型，揭示子宫内膜分化各阶段的基因表达调控机制，获得基于自体来源种子细胞组织工程技术修复受损子宫内膜新方法。

子宫在精子迁移、胚胎着床及胎儿给养等过程中起关键作用，而流产、刮宫、宫腔镜手术或感染等原因一旦损伤子宫内膜基底层，将导致上皮、基质细胞再生障碍，内膜难以自我修复，进而可引发妇科疾病，导致胚胎着床率下降、流产率上升、甚至不孕不育。目前尚无有效的防治手段。基于支架材料、生物活性物质和种子细胞的组织工程手段是一种非常具有应用前景的新型医疗手段，本项目利用当前防治方法的优势，探寻理想的、基于发育生物学的组织工程种子细胞及新型生物支架材料，研发修复受损子宫的新技术，以期用于子宫的修复再生研究。本研究首先通过化学分子、共培养分步诱导，将iPS细胞经中内胚层（mixl1,brachyury）细胞，中间中胚层（lim1, pax2,osr1）细胞，最后向人子宫上皮细胞（GdA阳性）诱导分化。随后通过单细胞分析手段，系统解析了子宫上皮细胞发育、分化过程中ALDH1A1阳性的上皮前体干细胞亚群，揭示了信号通路、表观、代谢及转录因子的时序性分化调控分子机制。最后构建大鼠子宫机械损伤模型，并以新生鼠子宫内膜细胞作为种子细胞，以脱细胞基质，丝素/纤维素膜为支架材料，复合SDF1a生长因子，发现可显著促进大鼠子宫内膜层及血管的再生，提高受孕胚胎的数量，实现子宫的功能性再生修复。本研究建立了iPS细胞向子宫内膜上皮细胞的分阶段发育、分化模型，揭示了发育、分化早期子宫内膜上皮前体干细胞亚群及分化调控分子机制，开发了基于发育来源种子细胞及新型生物材料的组织工程修复受损子宫新技术。