皮肤毛囊中不同Niche干细胞向原始生殖细胞及卵母细胞分化潜能的差异分析

Gamete differentiation from stem cells is a difficult but attractive topic in the field of reproduction and development. The use of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) is limited in clinical application due to their material sources, tumorigenicity, ethics and immune rejection. In comparison, adult stem cells within hair follicles are promising stem cell resources for gamete differentiation. To select the suitable stem cells for gamete differentiation and elucidate the mechanisms of oocyte differentiation, this program aims to analyze the differences of germ cell differentiation from skin hair follicle stem cells derived from different niches, and select the suitable niche stem cells for gamete differentiation, in order to obtain high quality of primordial germ cells. At the meantime, we will investigate the necessity of epiblast-like cells (EpiLCs) formation during differentiation from hair follicle stem cells to primordial germ cells, and elucidate the specialized process of primordial germ cell formation in vitro including the regulatory mechanisms of epigenetic modifications. Furthermore, this program will demonstrate the mechanisms underlying meiosis initiation during oocyte differentiation from in vitro derived primordial germ cells, and promote the acquisition of functional eggs via in vivo ectopic development. The scientific issues involved in this project will theoretically enrich our knowledge on early gametogenesis. In practice, oocyte differentiation from hair follicle stem cells may provide a new strategy for future treatment of infertility.

干细胞分化配子是生殖发育研究领域的难点和热点。胚胎干细胞和诱导多能性干细胞在临床应用上由于材料来源、致瘤性、伦理和免疫排斥等问题限制了其应用。相比较而言，皮肤毛囊结构中的干细胞却是一种非常有前景的向配子分化的干细胞类型。为了筛选适宜向配子分化的干细胞，并探索干细胞向卵母细胞分化的诱导机制，本项目拟分析皮肤毛囊不同Niche来源干细胞向生殖细胞分化的差异，甄选较适宜Niche的干细胞用于诱导分化，以期获得高质量的原始生殖细胞；探讨毛囊干细胞向原始生殖细胞分化中外胚层细胞形成的必要性，阐述体外形成的原始生殖细胞的特化过程及其表观调节机制；分析体外来源的原始生殖细胞向卵母细胞分化中减数分裂启动的诱导调控机制，借助体内异位发育促进功能卵子的获得。本项目拟解决的关键科学问题在理论上将丰富我们对早期配子发生机制的认识；在实践中，毛囊干细胞分化卵母细胞可能会为今后的不孕不育治疗提供一种新的策略和途径。

干细胞分化配子是生殖发育研究领域的难点和热点。胚胎干细胞和诱导多能性干细胞在临床应用上由于材料来源、致瘤性、伦理和免疫排斥等问题限制了其应用。相比较而言，皮肤来源干细胞（SDSCs）却是一种非常有前景的向配子分化的干细胞类型。为了筛选适宜向配子分化的干细胞，并探索干细胞向卵母细胞分化的诱导机制，本项目建立了SDSCs体外培养体系，利用10×RNA-seq技术构建了新生鼠皮肤组织及SDSCs的单细胞转录组图谱，甄选皮肤组织中具有生殖细胞诱导潜力的最佳干细胞类型，即毛乳头细胞来源的干细胞。继而，系统研究了毛乳头细胞来源的干细胞的体外增殖、向生殖细胞样细胞诱导分化及减数分裂进入等发育过程。在此基础上，进一步研究了毛囊干细胞（HFSCs）体外增殖、拟胚体形成、向外胚层样细胞（EpiLCs）和原始生殖细胞样细胞（PGCLCs）的体外诱导分化，建立了HFSCs向卵母细胞样细胞（OLCs）诱导分化的技术平台，相关成果获2项国家发明专利授权，发表论文12篇，培养研究生11名、青年教师3名，获省级科研成果奖3项，项目负责人被评选为山东省泰山学者特聘专家。综上，本项目顺利完成研究计划，明确了皮肤中最适宜向生殖细胞分化的干细胞类型，并阐述了HFSCs分化OLCs过程中的EpiLCs形成和PGCLCs特化的调节机制，丰富了我们对卵母细胞早期发生机制的认识，为今后不孕不育的自体干细胞治疗提供一种新的策略和技术支撑。