GENE EXPRESSION IN 3D FOLLICLES

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 这是路线图联盟(U54 RR02437，T.Woodruff，PI)的一个试点项目，题为“肿瘤生育联盟：为妇女保留生育能力”。体内卵泡发生的过程需要卵泡壁和卵丘细胞内的卵母细胞协调发育和成熟，以最终排卵和受精。在三维(3D)培养系统中生长小鼠卵泡的能力产生了能够受精和发育成活后代的成熟卵母细胞，这构成了我们目前对非人类灵长类动物的研究的基础。我们发现，猕猴卵泡在微囊化3D培养中可以存活、生长、形成窦腔并分泌激素。尽管将来自囊化3D培养的猕猴小腔卵泡暴露在成熟刺激下，大多数健康的卵母细胞仍未成熟。然而，两个卵母细胞成熟到中期II，一个通过胞质内单精子注射受精，并进行了早期胚胎卵裂。将猕猴卵泡的3D培养囊化到小胃窦阶段，可以单独分析每个卵泡室(体细胞、卵母细胞)的功能基因表达，并与体内发育到相同阶段/大小的卵泡进行比较。这些研究将评估被包裹的3D系统是否允许类似于体内的颗粒/膜细胞和卵丘-卵母细胞复合体的协调发展，如果不允许，将有助于确定需要在3D卵泡培养系统中进一步优化的细胞功能。这些知识对于优化3D卵泡培养技术转化为人体研究至关重要，这将有助于保留女性癌症幸存者的生育能力。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. This is a pilot project of the Roadmap Consortium (U54 RR02437, T. Woodruff, PI) entitled "The Oncofertility Consortium: Fertility Preservation for Women." The process of folliculogenesis in vivo requires the coordinated development and maturation of the follicle wall and the oocyte enclosed within cumulus cells for eventual ovulation and fertilization. The ability to grow mouse follicles in a three-dimensional (3D) culture system yielding mature oocytes capable of fertilization and development to live offspring formed the basis of our current studies in nonhuman primates. We found that macaque follicles can survive, grow, form an antrum and secrete hormones when grown in encapsulated 3D culture. Despite exposure of macaque small antral follicles derived from encapsulated 3D culture to a maturation stimulus, most of the healthy oocytes remained immature. However, two oocytes matured to metaphase II, one fertilized via intracytoplasmic sperm injection and underwent early embryonic cleavage. Encapsulated 3D culture of macaque follicles to the small antral stage allows the functional gene expression of each follicular compartment (somatic cells, oocytes) to be analyzed separately and compared to follicles that develop to an equivalent stage/size in vivo. These studies will assess whether the encapsulated 3D system allows coordinated development of granulosa/theca cells and cumulus-oocyte complexes similar to that in vivo, and if not, will help define cellular functions that require further optimization in the 3D follicle culture system. This knowledge is critical for optimizing the 3D follicle culture technology for translation to human studies which will contribute to fertility preservation for female cancer survivors.