Xenotransplantation: a new paradigm for human meiosis

描述(申请人提供)：女性的生育能力是多种分子途径的产物，这些途径始于胚胎发育，包括生殖细胞进入减数分裂，以及导致原始卵泡形成的卵巢结构重组。女性对这些事件知之甚少，因为很难获得孕周较晚的胎儿卵巢样本。了解这些事件的重要性通过观察到减数分裂在妇女中非常容易出错，70%的妊娠损失是由于母体减数分裂I错误造成的。虽然在减数分裂前期对人胎儿卵母细胞进行了一些有限的分析，但很难研究前期I的后期以及在出生前后进入二倍体停滞状态，这是触发发育中卵巢内重要结构变化的过程。为了克服这些困难，R21的应用试图建立一种新的异种移植系统来分析人类减数分裂和卵巢发育。在目标1中，我们将建立人胎儿卵巢和不同免疫缺陷小鼠宿主的异种移植方法学，以优化异种移植的建立条件。将通过一系列免疫组织化学和细胞遗传学标准评估异种移植的细胞活性和发育。在目标2中，我们将通过前期I期研究异种移植的人类卵母细胞的进展，利用我们在哺乳动物减数分裂方面的重要专业知识来分析同源染色体的相互作用、重组动力学和交叉形成。在目标3中，我们将研究人类卵母细胞中二倍体停滞的开始，以确定在间质重组和原始卵泡形成过程中的分子/细胞事件。这一模型为研究卵巢发育提供了一个新的范式，这是一个知之甚少的过程，但其中的缺陷可能导致女性观察到的高非整倍体率，与其他哺乳动物相比，卵巢早衰和其他影响女性的生育问题。

DESCRIPTION (provided by applicant): Fertility in women is a product of multiple molecular pathways that start during embryogenesis and involve the entry of germ cells into meiosis, together with the structural reorganization of the ovary that results in primordial follicle formation. These events are poorly understood in women because of the difficulty in obtaining fetal ovarian samples of later gestational ages. The importance of understanding these events is underscored by the observation that meiosis is highly error-prone in women, with 70% of pregnancy losses being due to maternal meiosis I errors. While some limited analysis has been performed on human fetal oocytes during early meiotic prophase I, it has been hard to examine later stages of prophase I and the entry into dictyate arrest which occurs at around the time of birth and which triggers the important structural changes within the developing ovary. To overcome these difficulties, this R21 application seeks to establish a novel xenotransplantation system for analyzing human meiosis and ovarian development. In aim 1, we will establish the xenografting methodology using human fetal ovaries and different immunocompromised mouse hosts, to optimize the conditions for xenograft establishment. Xenografts will be assessed for cell viability and development by an array of immunohistochemical and cytogenetic criteria. In aim 2, we will examine the progression of xenotransplanted human oocytes through prophase I, taking advantage of our significant expertise in mammalian meiosis to analyze homologous chromosome interactions, recombination dynamics and crossover formation. In aim 3, we will examine the onset of dictyate arrest in human oocytes to define the molecular/cellular events during stromal reorganization and primordial follicle formation. This model provides a new paradigm for studying ovarian development, a poorly understood process, but defects in which may contribute to the high aneuploidy rates observed in women compared to other mammals, premature ovarian failure and other fertility issues affecting women.