Biased signaling pathways in cumulus-oocyte complexes

在过去的几十年里，虽然牛奶产量在增加，但奶牛群的生育力却在下降。利用体外培养体系是迅速应对生育能力下降的一种方法。奶牛行业是加拿大最重要的农业食品部门之一，2012年140万头奶牛生产7900万百升牛奶，总农场收入为59亿美元，基础研究的重要需求对奶牛行业至关重要。该研究计划将研究卵母细胞体外成熟过程中减数分裂恢复的细胞内信号机制。卵母细胞减数分裂恢复是卵子受精的重要过程。改善卵母细胞质量的最佳时机出现在卵母细胞减数分裂恢复之前。卵丘细胞是已知的卵母细胞发育能力的调节剂。培养条件的改善，如通过抑制卵母细胞减数分裂恢复延长窗口，需要更好地理解所涉及的信号通路。众所周知，环腺苷一磷酸（cAMP）是卵泡细胞对促卵泡激素（FSH）反应的关键第二信使。环状AMP是哺乳动物卵母细胞减数分裂恢复的关键调节因子。合成和降解的产物调节细胞内cAMP浓度。除了cAMP合成外，磷酸二酯酶（PDE）的降解也是一种诱导细胞生理反应的强大调节剂。因此，咖啡因，一种非特异性PDE抑制剂，是我们日常生活中一杯咖啡的一部分。最引人注目的例子是伟哥®，一种特殊的PDE抑制剂，近二十年来一直用于治疗勃起功能障碍的男性。这些例子说明了PDE如何通过影响哺乳动物的关键生理事件来影响日常生活。在过去的几年里，我的实验室一直对PDE在不同生殖功能中的作用感兴趣。虽然PDE3A存在于牛卵母细胞中，但我们最近的发现之一是PDE8存在于牛卵母细胞和卵丘细胞中。我们的工作假设是，PDE8对cAMP的降解在卵巢卵泡细胞信号传导中起生理作用，作为综合信号细胞反应的一部分。因此，我们将研究PDE8在牛卵母细胞体外成熟过程中的作用。我们将分析FSH和表皮生长因子对体外卵母细胞减数分裂恢复的诱导作用，以及c型利钠肽的抑制作用。卵母细胞质量的提高将通过更好地理解卵母细胞在体外成熟的最初几个小时内减数分裂恢复之前发生的信号事件来实现。由于体外胚胎生产的改善是胚胎移植公司的当务之急，该项目将揭示使诊断和治疗工具的发展取得进展所需的知识。由于观察到奶牛群生育力的下降，这些进步对奶牛行业至关重要。该计划将评估基本信号，从而在短期内提供即时的有益应用。

During the last decades while milk production is increasing, fertility of the dairy cattle herd is decreasing. The utilization of in vitro culture system is one way to rapidly counter the decline of fertility. An important need of fundamental research is crucial for dairy cow industry that is one of the most important agri-food sectors in Canada, with a total farms receipts of $5.9 billion, 79 million hectolitres produced by 1.4 million of heads in 2012. The proposed research program will study intracellular signaling mechanisms involved in oocyte meiotic resumption during in vitro maturation. Oocyte meiotic resumption is an essential event allowing the egg to be fertilized. The optimal window to improve oocyte quality occurs before oocyte meiotic resumption. Cumulus cells are the known modulators of oocyte developmental competence. The improvement of culture conditions, such as prolongation of the window by inhibiting oocyte meiotic resumption requires a better understanding of the signalling pathways involved.It is well-known that cyclic adenosine monophosphate (cAMP) is a crucial second messenger in response of follicular cells to FSH (follicle-stimulating hormone). Cyclic AMP is a key regulator of oocyte meiotic resumption in mammals. The resultant of synthesis and degradation modulates intracellular cAMP concentration. Beyond cAMP synthesis, the degradation by phosphodiesterases (PDE) is a powerful modulator inducing physiological response of the cells. It is thus that caffeine, a non-specific PDE inhibitor, is part of our daily life with a cup of coffee. The most spectacular example relies on Viagra® a specific PDE inhibitor prescribed for almost two decades in men with erectile dysfunction. These examples are illustrating how PDE may impact the daily life by affecting crucial mammalian physiological events. Over the last years my laboratory has been interested in understanding the role of PDE in different reproductive functions. Although PDE3A is functionally present in bovine oocytes, one of our recent findings is that PDE8 was present in bovine oocyte and cumulus cells. Our working hypothesis is stating that cAMP degradation by PDE8 plays a physiological role in ovarian follicular cells signaling as part of an integrative signaling cellular response. We will thus investigate the role of PDE8 in bovine cumulus-oocytes complexes during in vitro maturation. This will be analyzed in response to induction of in vitro oocyte meiotic resumption by FSH and epidermal growth factor, and in response to inhibition by C-type natriuretic peptide. The improvement of oocyte quality will be achieved through a better understanding of the signalling events occurring prior to oocyte meiotic resumption during the first hours of in vitro maturation. Because the improvement of in vitro embryo production is a matter of urgency for embryo transfer companies, this project would reveal the knowledge required to enable progress in the development of diagnostic and therapeutic tools. Such advances are of prime importance to the dairy cow industry because of the observed decreases in dairy herd fertility. This program will assess fundamental signalling and thus provide immediate beneficial applications in the short-term.