The role of chemokines in cumulus oocyte expansion (C-OE) and oocyte maturation

摘要 排卵是一个复杂的、类似炎症的过程，在这个过程中，完全发育的卵泡会破裂。 对周期中期促性腺激素激增的作用，释放卵丘-卵母细胞复合体(COC)进行传递 进入生殖道和可能的受精。在排卵前不久，黄体生成素(L H) 电涌诱导对生育至关重要的过程，包括卵丘-卵母细胞扩张(C-OE)和恢复 减数分裂。虽然已经确定了一些对这些事件重要的旁分泌作用因素，但 启动这种复杂过程的分子机制尚不完全清楚。有限 已发表的研究和PI的初步发现支持趋化因子调节事件的新作用 C-OE和卵母细胞成熟所必需的。啮齿动物研究表明，趋化因子信号调节 积云细胞外基质的组装，从而受精。PI的初步结果来自国内 CATS证明趋化因子MCP-1及其受体(CCR2)在卵母细胞和 小腔卵泡来源的COCs的卵丘细胞。在猴子的COC中也观察到了类似的结果 有腔卵泡和排卵前卵泡。此外，排卵前卵泡12中单核细胞趋化蛋白-1和CCR2基因表达增加 在排卵刺激后24小时和36小时，MCP-1恢复到hCG前水平。MCP-1蛋白含量 在hCG后12小时，卵泡液中的mRNA水平达到峰值，尤其是在 是在启动C-OE所必需的事件和恢复猕猴卵母细胞减数分裂之前 排卵前卵泡。基于这些发现，我们假设，与血管内皮细胞 趋化因子受体CCR2(如MCP-1)直接促进C-OE和/或卵母细胞 成熟。因此，为了验证这一假设，实验被设计成：1)评估促黄体生成素依赖 排卵前卵泡中趋化因子及其受体的调节，2)评估CCR2信号转导的作用 关于C-OE和/或卵母细胞成熟所必需的分子事件的调节，3)决定 COC中CCR2信号的抑制阻止了C-OE和/或卵母细胞的成熟。该项目将是 在猫模型中开发，它的优点是为理解提供了一个很好的替代品 人类的COC是生育所必需的；因为猫的卵母细胞与人类有几个共同的特征 卵母细胞。CAT还提供了一个独特而有价值的模型来研究分子过程。 排卵前卵泡和COC，由于每只动物提供3到7个排卵前卵泡， 在发情期自然选择处于“排卵准备”状态，等待促黄体生成素刺激，提供了一个窗口 样品可以在大约7天内采集。因此，这些新颖和创新的研究将有助于 增加了我们对受精所需事件的理解，从而导致了对小说的识别 避孕的非荷尔蒙目标以及确定不孕不育和/或良好的可能原因 卵母细胞质量的分子标记。

SUMMARY Ovulation is a complex, inflammation-like process whereby a fully-developed follicle ruptures in response to the actions of the mid-cycle gonadotropin surge, releasing the cumulus-oocyte complex (COC) for passage into the reproductive tract and possible fertilization. Shortly before ovulation, the luteinizing hormone (LH) surge induces processes critical for fertility, including cumulus-oocyte expansion (C-OE) and resumption of meiosis. While some of the paracrine-acting factors important for these events have been identified, the molecular mechanisms responsible for initiating such complex processes are not fully understood. Limited published studies and the PI's preliminary findings support a novel role for chemokine regulating events necessary for C-OE and oocyte maturation. Rodent studies suggest that chemokine signaling regulates the assembly of the cumulus extracellular matrix and thus fertilization. The PI's preliminary results from domestic cats demonstrated that the chemokine MCP-1 and its receptor (CCR2) are expressed in the oocyte and cumulus cells of COCs from small antral follicles. Similar results were observed in the monkey COC from small antral and preovulatory follicles. Moreover, MCP-1 and CCR2 mRNA increased in the preovulatory follicle 12 hr after an ovulatory stimulus with MCP-1 returning to pre-hCG levels at 24 and 36 hr. MCP-1 protein content in the follicular fluid paralleled the mRNA levels and peaked at 12 hr post hCG, notably the period of time that is just prior to the initiation of events necessary for C-OE and the resumption of oocyte meiosis in the rhesus preovulatory follicle. Based on these findings, we hypothesize that chemokines that interact with the chemokine receptor CCR2 (e.g. MCP-1) have a direct effect on promoting C-OE and/or oocyte maturation. Therefore, to test this hypothesis experiments are designed to: 1) Evaluate LH-dependent regulation of chemokines and their receptors in preovulatory follicles, 2) Assess the effects of CCR2 signaling on the regulation of molecular events necessary for C-OE and/or oocyte maturation, 3) Determine whether the inhibition of CCR2 signaling in the COC prevents C-OE and/or oocyte maturation. The project will be developed in a feline model, which has the advantage of providing an excellent surrogate for understanding events the human COC that are necessary for fertility; as cat oocytes share several characteristics with human oocytes. The cat also provides a unique and valuable model to study molecular processes within the preovulatory follicle and COC, due to the fact that each animal provides between 3 to 7 preovulatory follicles, naturally selected in an "ovulation-ready" state waiting for the LH stimulus during estrus, providing a window of approximately 7 days that sample can be collected. Therefore, these novel and innovative studies will aid in increasing our understanding of events required for fertilization, thereby leading to the identification of novel non-hormonal targets for contraception as well as identification of possible cause of infertility and/or good molecular markers for oocyte quality.