Reversal of Ovarian Aging in Mice Through AAV-mediated Oocyte Reprogramming in vivo

项目摘要 衰老是一个复杂的、多因素的生物学过程，所有活着的有机体都有这个过程。在卵巢中，它表现为 正常生殖功能的逐渐下降，当卵母细胞的数量达到绝经期时 跌破一个门槛。更年期之前，从35年代中期开始，更年期显著增加 因年龄增长而导致减数分裂错误、非整倍体和不良的卵母细胞功能障碍的发生率 胚胎发育。这种细胞功能障碍的临床表现为不孕、辅助生殖失败。 技术(ART)、(复发性)妊娠丢失和出生缺陷。目前还没有有效的治疗方法 改善高龄育龄妇女的卵母细胞功能和新的方法策略是 这是急需的。参与卵母细胞减数分裂机制的许多因素都与年龄有关- 相关的卵母细胞功能障碍。然而，卵巢老化的根本原因以及为什么雌性生殖系 相对于体细胞而言，经历加速衰老的过程仍不清楚。随时间变化的累积 细胞损伤被广泛认为是衰老的基础，有证据表明衰老的速度得到了控制， 至少在某种程度上，通过进化中保守的遗传途径。最近的研究表明，在躯体中 细胞表观遗传格局的侵蚀(表观基因组漂移)导致的细胞功能障碍可能主要是 对衰老负有责任。有趣的是，这些研究表明，通过异位表达进行的部分重编程 一组有限的多能因子可以恢复年轻的表观遗传模式并逆转衰老的迹象 体细胞。然而，部分重编程对减数分裂细胞的细胞特性的影响，表现出 独特的基因调控网络和维持功能的多能性，是未知的。 这项建议研究了卵巢老化的全球机制，挑战了它的不可逆转性，并开发了一种 为未来开发延缓或逆转卵巢衰老的治疗方法提供了新的途径。我们将聘请一名 转基因小鼠模型检验卵丘细胞-卵母细胞部分重编程的假说 复合体可以逆转卵巢衰老，恢复生殖功能。我们将使用整合的多元组学来 明确甲基化时钟和表观遗传漂移在卵巢衰老中的作用，以及非卵母细胞是否 由于年龄增长，卵巢导致卵母细胞功能障碍。我们将调查部分重新编程是否可以 逆转卵母细胞-卵丘复合体中与年龄相关的转录和表观遗传学变化，使其更年轻 在不改变卵母细胞的细胞特性的情况下，恢复体内的生育能力。然后我们将研究 这种方法的翻译潜力使用一种新型的AAV递送系统，具有跨越血液的能力- 卵泡屏障和体内靶向卵母细胞和颗粒细胞。这项工作将增加我们对 卵巢衰老的生物学研究，为表观遗传药物的开发提供新的分子靶点 逆转人类生殖卵巢功能下降，减少流产、反复妊娠 与高龄生育有关的丢失和出生缺陷。

Project Summary Aging is a complex multifactorial biological process shared by all living organisms. In the ovary, it manifests as a gradual decline of normal reproductive function which culminates in menopause when the number of oocytes falls below a threshold. Menopause is preceded, from the mid-30s onward, by a dramatically increased incidence of oocyte dysfunction due to aging which results in meiotic segregation errors, aneuploidy and poor embryo development. This cellular dysfunction clinically presents as infertility, failure of assisted reproduction technologies (ART), (recurrent) pregnancy loss and birth defects. There are currently no effective treatments to improve oocyte function in women with advanced reproductive age and novel methodological strategies are critically needed. Many factors involved in the meiotic machinery in oocytes have been implicated in age- related oocyte dysfunction. However, the underlying cause of ovarian aging and why the female germline undergoes accelerated aging relative to somatic cells remains unknown. Time-dependent accumulation of cellular damage is widely considered to underlie aging and there is evidence that the rate of aging is controlled, at least to some extent, by genetic pathways conserved in evolution. Recent studies indicate that in somatic cells an erosion of the epigenetic landscape (epigenomic drift) resulting cellular dysfunction may be primarily responsible for aging. Interestingly, these studies show that ‘partial reprogramming’ through ectopic expression of a limited set of pluripotency factors can restore youthful epigenetic patterns and reverse signs of aging in somatic cells. However, the effect of partial reprogramming on the cell identity of meiotic cells, which exhibit unique gene regulatory networks and maintain features of pluripotency, is unknown. This proposal examines the global mechanism of ovarian aging, challenges its irreversibility and develops a novel pathway for the future development of treatments to slow or reverse ovarian aging. We will employ a transgenic mouse model to examine the hypothesis that partial reprogramming of the cumulus cell-oocyte complex can reverse ovarian aging and restore reproductive function. We will use integrative multi-omics to define the role of the methylation clock and epigenetic drift in ovarian aging and whether non-oocyte cells of the ovary contribute to oocyte dysfunction due to aging. We will investigate whether partial reprogramming can reverse age-related transcriptional and epigenetic changes in the oocyte-cumulus-complex to more youthful patterns and restore fertility in vivo without altering the cellular identity of oocytes. We will then examine the translational potential of this approach using a novel AAV delivery system with the ability to cross the blood- follicle barrier and target oocytes and granulosa cells in vivo. This work will increase our understanding of the biology of ovarian aging and provide novel molecular targets for the development of epigenetic drugs that reverse the decline in human reproductive ovarian function and reduce the miscarriages, recurrent pregnancy loss and birth defects associated with advanced reproductive age.