Regulation of Primordial Follicle Formation and Oocyte Survival

 描述(由申请人提供)：尽管原始生殖细胞对物种的延续很重要，但对原始生殖细胞分化为有功能的卵母细胞知之甚少。原始生殖细胞在小鼠胚胎发育期间开始分化为卵母细胞。在出生之前，卵母细胞发育在称为生殖系囊的簇中，这是脊椎动物和无脊椎动物卵母细胞发育的一个保守阶段。在胎儿晚期和新生儿早期发育期间，小鼠生殖细胞囊分裂成单个卵母细胞，这些卵母细胞被颗粒前细胞包围，形成原始卵泡。在这个包囊破裂的过程中，每个包囊中的一部分细胞死亡，只有初始数量的三分之一的卵母细胞存活下来，形成原始卵泡。目前，控制卵囊破裂、卵母细胞凋亡和卵泡组装的机制尚不清楚。长期目标是了解调节囊泡破裂和程序性细胞死亡的分子和细胞机制，以建立小鼠卵巢中的原始卵泡池。本研究的目的是确定KIT信号通路和BCL2凋亡通路在调节囊泡破裂和卵母细胞数量中的作用。这项研究的中心假设是，囊泡破裂和生殖细胞死亡是由受体酪氨酸激酶KIT的信号和bcl2促凋亡和抗凋亡蛋白的平衡控制的。我们实验室最近的研究表明，KIT信号可能起着重要作用。此外，我们还提供了初步的数据，表明BCL2家族蛋白在调节囊泡破裂和相关的程序性细胞死亡中也是重要的。该提案探讨了KIT信号和程序化细胞死亡调节因子在囊泡破裂和卵母细胞存活中的分子和细胞方面的作用。本研究的具体目的是：1)阐明KIT信号在囊泡破裂和相关的卵母细胞丢失中的分子和细胞机制；2)确定参与调控卵母细胞存活的bcl2家族成员。这些目标将通过免疫细胞化学、共聚焦显微镜、Western blotting、卵巢器官培养、药物抑制剂、siRNA技术和遗传学等技术实现。在目前的应用中提出的研究具有重要意义，因为它将通过阐明囊泡破裂和相关的卵母细胞丢失的调节机制来增强我们目前的知识。这笔赠款的结果将有助于改进卵巢生物学和治疗导致女性不孕的疾病(如原发性卵巢功能不全)的研究工作。

 DESCRIPTION (provided by applicant): Despite its importance to the continuation of species, the differentiation of primordial germ cells into functional oocytes is poorly understood. Primordial germ cells begin to differentiate into oocytes during embryonic development in the mouse. Prior to birth, the oocytes develop in clusters called germline cysts, a conserved phase of oocyte development in both vertebrates and invertebrates. During late fetal and early neonatal development, mouse germ cell cysts break apart into single oocytes that become surrounded by pre-granulosa cells to form primordial follicles. During this process of cyst breakdown, a subset of cells in each cyst die with only a third of the initial number of oocytes surviving to form primordial follicles. The mechanisms that control cyst breakdown, oocyte apoptosis and follicle assembly are currently unknown. The long-term goal is to understand molecular and cellular mechanisms that regulate cyst breakdown and programmed cell death to establish the primordial follicle pool in the mouse ovary. The objective of this proposal is to determine the role of two pathways, the KIT signaling pathway and the BCL2 apoptotic pathway in modulating cyst breakdown and oocyte numbers. The central hypothesis of the proposed research is that cyst breakdown and germ cell death are controlled by signaling through the receptor tyrosine kinase, KIT and by a balance of BCL2 pro- and anti-apoptotic proteins. Recent work from our laboratory suggests KIT signaling may play an important role. In addition, we provide preliminary data demonstrating that BCL2 family proteins are also important for the regulation of cyst breakdown and associated programmed cell death. This proposal explores the molecular and cellular aspects of KIT signaling and programmed cell death regulators in cyst breakdown and oocyte survival. The specific aims of this research are to: 1) elucidate the molecular and cellular mechanisms of KIT signaling in cyst breakdown and associated oocyte loss; and 2) identify BCL2 family members involved in regulating oocyte survival. These goals will be achieved through techniques including immunocytochemistry, confocal microscopy, Western blotting, ovary organ culture, pharmacological inhibitors, siRNA technology and genetics. Research proposed in the current application is significant because it will enhance our current knowledge by elucidating the mechanisms by which cyst breakdown and associated oocyte loss are regulated. Results obtained in this grant will help improve research efforts in ovarian biology and in treatment of conditions causing female infertility such as primary ovarian insufficiency.