Human iPSC-derived ovarian follicles as a model of female reproduction

人类 iPSC 衍生的卵泡作为女性生殖模型

• 批准号: 10731755
• 负责人: Merrick De Forest Pierson Smela
• 金额: $ 4.13万
• 依托单位: HARVARD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10731755
• 关键词: American; Angelman Syndrome; Area; Biological Models; Cell Line; Cell physiology; Cells; ChIP-seq; Characteristics; Chromosome abnormality; Church; Coculture Techniques; DNA methyltransferase inhibition; Development; Developmental Process; Disease; Down Syndrome; Embryonic Development; Engineering; Epigenetic Process; Estradiol; FOXL2 gene; Female; Female infertility; Fertility; Fetus; Gene Expression Profiling; Generations; Genetic; Genetic Transcription; Germ Cells; Goals; Growth; Growth Factor; Human; Human Development; Impairment; In Vitro; Knock-out; Meiosis; Methods; Modeling; Mus; Oocytes; Oogenesis; Oogonia; Organism; Organoids; Ovarian; Ovarian Follicle; Ovarian Granulosa Cell; Ovary; Paracrine Communication; Phenotype; Pluripotent Stem Cells; Prevention; Process; Production; Publishing; Regulation; Reporter; Repression; Reproduction; Research; Route; Signal Transduction; Somatic Cell; Specific qualifier value; Structure; Structure of primordial sex cell; System; Techniques; Transgenic Organisms; Validation; Woman; bisulfite sequencing; cell motility; cell type; combinatorial; computerized tools; developmental disease; differentiation protocol; directed differentiation; embryonic stem cell; epigenetic profiling; epigenome; experimental study; extracellular; fetal; granulosa cell; human model; improved; in vitro Model; induced pluripotent stem cell; infertility treatment; insight; interest; mouse model; reconstitution; reproductive; reproductive development; response; screening; sex development disorder; single-cell RNA sequencing; stem cell differentiation; stem cells; transcription factor; transcriptomics; transmission process

PROJECT SUMMARY The central process of female reproduction is the formation of oocytes within the developing ovary, known as oogenesis. This process is crucial for the formation of healthy oocytes and proper transmission of genetic and epigenetic information to begin embryonic development. Abnormalities in ovarian development and oogenesis are a leading cause of female infertility and disorders of sexual development, and furthermore are the cause of many developmental disorders in the subsequent generation, such as Down syndrome and Angelman syndrome. However, relatively little is known about the genetic regulation of human ovarian development. This is in contrast to other organisms such as the mouse, where transgenic and knockout lines, and a short reproductive cycle, have allowed much research in this area. An in vitro organoid model of human ovarian development would help fill this gap, and enable an improved understanding of human ovarian development that could lead to treatments for infertility and prevention of developmental disorders. Ovarian development involves interactions between primordial germ cells (PGCs) and somatic cells (granulosa cells). The granulosa cells enclose the PGCs within ovarian follicles, and support their differentiation into oogonia, progression through meiosis, and development as oocytes. Therefore, both lineages will be required to model this process in vitro. Existing methods allow differentiation of induced pluripotent stem cells (iPSCs) into PGC-like cells, but these cells are in an immature state, retaining epigenetic characteristics of iPSCs. For an in vitro model of oogenesis to be successful, improved methods must be developed to generate mature germline cells and granulosa cells from iPSCs. Reprogramming of cellular identity by expression of transcription factors (TFs) is a powerful technique that can allow both reprogramming of somatic cells into iPSCs, and directed differentiation of iPSCs to specific cell types. The Church lab has recently developed computational tools to predict TFs that specify cell identity, as well as screening methods for combinatorial TF expression to find sets that can differentiate iPSCs to a cell type of interest. The currently proposed research will identify TFs that can promote maturation of PGC- like cells and produce granulosa cells from iPSCs. Results will be evaluated by single-cell transcriptomic and epigenetic profiling, and by functional validation of key phenotypes. This research will provide an improved understanding of the genetic regulation of ovarian development, leading to an in vitro model of human oogenesis.

项目概要 女性生殖的核心过程是在发育中的卵巢内形成卵母细胞，称为 卵子发生。这个过程对于健康卵母细胞的形成以及遗传和基因的正确传递至关重要。 表观遗传信息开始胚胎发育。卵巢发育和卵子发生异常 是导致女性不孕和性发育障碍的主要原因，也是导致 下一代的许多发育障碍，例如唐氏综合症和天使综合症。 然而，人们对人类卵巢发育的遗传调控知之甚少。这是在 与其他生物体（例如小鼠）相比，转基因和敲除系以及短 生殖周期，使得这方面的研究得以进行。人卵巢体外类器官模型 开发将有助于填补这一空白，并提高对人类卵巢的了解 可能导致治疗不孕症和预防发育障碍的发展。 卵巢发育涉及原始生殖细胞 (PGC) 和体细胞（颗粒细胞）之间的相互作用 细胞）。颗粒细胞将 PGC 包围在卵泡内，并支持其分化为 卵原细胞、通过减数分裂的进展以及卵母细胞的发育。因此，两个血统都需要 在体外模拟这个​​过程。现有方法可分化诱导多能干细胞 (iPSC) 转化为PGC样细胞，但这些细胞处于未成熟状态，保留了iPSC的表观遗传特征。为了 卵子发生的体外模型要成功，必须开发改进的方法来产生成熟的卵子 来自 iPSC 的生殖细胞和颗粒细胞。 通过转录因子 (TF) 的表达对细胞身份进行重编程是一种强大的技术，可以 允许将体细胞重编程为 iPSC，并将 iPSC 定向分化为特定细胞 类型。 Church 实验室最近开发了计算工具来预测指定细胞身份的 TF，如 以及组合 TF 表达的筛选方法，以找到可将 iPSC 分化为细胞的组 兴趣类型。目前提出的研究将确定能够促进 PGC 成熟的 TF 类似细胞并从 iPSC 产生颗粒细胞。结果将通过单细胞评估 转录组和表观遗传分析，以及关键表型的功能验证。这项研究将 更好地了解卵巢发育的遗传调控，从而产生体外 人类卵子发生模型。