Aging and Ovarian Stem Cell Niche Dysfunction

描述(申请人提供)：干细胞生物学最令人兴奋的领域之一与干细胞功能障碍在与衰老相关的器官功能恶化中发挥核心作用的可能性有关。最近的研究表明，卵原(产生卵母细胞的)干细胞存在于成年哺乳动物的卵巢中。OSCs的存在增加了一种可能性，即以含卵母细胞卵泡的丢失和耗尽为标志的卵巢衰老，可能同样涉及干细胞功能的进行性丧失。因此，现在重要的是确定OSCs在卵巢功能和衰老中的生理作用。我的长期目标是确定衰老过程如何对OSC功能产生负面影响，从而将OSCs作为模型来提供对基于干细胞的生物衰老机制的洞察。具体地说，我建议使用我们在过去三年中开发的新型自杀基因转基因(sg-tg)小鼠模型来研究这一点。这些小鼠模型的独特之处在于，可以使用自杀基因技术选择性地靶向和切除分化中的OSC子代细胞。在我们的初步数据中，我们表明，这些OSC分化途径的选择性中断会导致原始卵泡在基因上定义的可逆性丧失。这些发现支持哺乳动物成年期卵巢中卵母细胞储备的维持需要OSCs主动输入新的卵母细胞。这些数据也为未来卵巢生物学和疾病发病机制的研究奠定了坚实的基础，具有前所未有的可能性，包括了解女性生殖老化对卵巢功能的影响。本建议的具体目的是：1)利用我们培育的新的sg-TG小鼠，建立OSC在卵巢衰老中的生理作用，其中分化的OSC子代细胞可以被选择性地去除；2)评估胶质细胞源性神经营养因子(GDNF)和成纤维细胞生长因子(FGF9)分别作为刺激OSC增殖和抑制OSC减数分裂分化的可能的OSC生态位因子；3)表征OSC有丝分裂活性增加时期的OSC生态位，并检测血管内皮细胞在生态位功能中的参与；4)以卵巢体细胞为饲养细胞，建立改良的OSC培养条件，总结OSC-NICE的体外相互作用，并检测衰老对OSC活性的影响。最终，这些信息可能被用来开发新的、有针对性的疗法，在需要的时候通过刺激OSC活动来增加卵母细胞储备，从而挽救卵巢功能--例如，在卵巢早衰患者、高龄孕妇(推迟与年龄相关的卵巢衰竭和更年期)或女性癌症患者(抗癌治疗后挽救她们的卵巢功能和生育能力)中，所有这些情况都代表着公共卫生的相关性越来越大。

DESCRIPTION (provided by applicant): One of the most exciting areas of stem cell biology relates to the possibility that stem cell dysfunction plays central role in aging-related deterioration of organ function. Recent studies suggest that oogonial (oocyte- producing) stem cells (OSCs) exist in the adult mammalian ovary. Existence of OSCs raises the possibility that ovarian aging, marked by loss and exhaustion of oocyte-containing follicles, may similarly involve a progressive loss of stem cell function. Thus, it is now important to establish the physiological roles of OSCs in ovarian function and aging. My long-term goal is to determine how the aging process negatively affects OSC function, and thus use OSCs as a model to provide insight into stem cell-based mechanisms for organismal aging. Specifically, I propose to use novel suicide gene transgenic (sg-Tg) mouse models we have developed over the past three years to study this. These mouse models are unique in that differentiating OSC daughter cells can be selectively targeted and ablated using suicide gene technology. In our preliminary data, we show that selective disruption of these OSC differentiation pathways results in a genetically defined reversible loss of primordial follicles. These findings support that the maintenance of the oocyte reserve in mammalian ovaries during adulthood involves active input of new oocytes from OSCs. These data also lay a solid foundation for future studies of ovarian biology and disease pathogenesis with unprecedented possibilities, including an understanding of female reproductive aging that accounts for OSC contribution to ovarian function. The specific aims of this proposal are to: 1) establish the physiological roles of OSCs in ovarian aging using novel sg-Tg mice we have developed, in which differentiating OSC daughter cells can be selectively ablated; 2) evaluate glial cell-derived neurotrophic factor (GDNF) and fibroblast growth factor (FGF9) as possible OSC niche factors in stimulating OSC proliferation and suppressing OSC meiotic differentiation, respectively; 3) characterize OSC niches during periods of increased OSC mitotic activity and examine the participation of vascular endothelial cells in niche function; and, 4) develop improved OSC culture conditions by using ovarian somatic cells as feeder cells to recapitulate OSC-niche interaction ex vivo and examine the impact of aging on OSC activity. Ultimately, this information might be used to develop novel and targeted therapeutics that rescue ovarian function through increasing the oocyte reserve by stimulating OSC activity when it would be desirable - such as in patients with premature ovarian failure, in women of advanced maternal age (to postpone age- related ovarian failure and menopause) or in female cancer patients (to rescue their ovarian function and fertility after anti-cancer treatments) - all of these conditions represent increasing public health relevance.