The Role Of CCL20 In Ovulation: A Novel Mediator Of Ovarian Leukocyte Recruitment

描述(由申请人提供)：排卵前促黄体生成素高峰启动了一系列事件，最终导致卵泡壁的破裂和卵子在排卵过程中的释放。排卵涉及一个炎症过程，其特征是白细胞大量涌入卵巢。这种白细胞的流入对生育至关重要，因为在实验模型中，去除白细胞会阻止卵母细胞的释放。然而，白细胞募集到卵巢的潜在因素(S)尚不清楚。本提案将审查一种新确定的候选趋化因子配体20(CCL20)。我们的初步数据显示，在人类排卵前早期和晚期，CCL20的mRNA在卵泡的颗粒细胞和膜细胞中大量诱导。然而，除了我们的初步数据外，关于CCL20及其受体CCR6在卵巢中的表达、调节或功能的时间模式尚不清楚。目前的建议建立在这些挑衅性发现的基础上，并将检验以下总体假设：促黄体生成素激增刺激CCL20增加，而这种趋化因子的增加会导致白细胞的招募，从而影响排卵所必需的事件。这一假设将使用一个独特的模型来解决，在该模型中，将在hCG之前和之后的三个指定时间收集人类排卵周卵泡的颗粒和膜，以诱导排卵(早期、晚期和排卵后)。这些人类样本将与大鼠卵巢中CCL20和CCR6的变化进行比较，并使用具有良好特征的啮齿动物排卵模型进行对比。这将通过以下三个目标实现。目的1测定CCL20和CCR6系统在人和大鼠卵巢中的表达变化。这一目标也将决定CCL20和CCR6的表达如何通过黄体生成素诱导的主要排卵途径来调节。目的2利用已有的迁移和侵袭模型，探讨CCL20在白细胞重新聚集到卵巢中的作用。由于CCL20调节细胞分化的潜在作用，Aim 3将研究CCL20对卵巢细胞功能的作用，特别是颗粒细胞存活、分化和产生类固醇激素的能力。此外，将使用微阵列技术评估CCL20对卵巢细胞的影响。当前方案的主要优点在于使用特征良好的人类排卵前卵泡作为基础，以了解CCL20在人类排卵相关事件中的细胞表达、调节和影响，以及我们跨物种的综合比较方法来阐明这种趋化因子在排卵过程中所起的作用。关于CCL20及其在卵巢中的受体，目前尚不清楚。因此，建议的研究非常及时地阐明该趋化因子系统在卵泡破裂和卵母细胞释放的协调过程中所起的作用，而卵泡破裂和卵母细胞释放是正常卵巢生理的基本方面。

DESCRIPTION (provided by applicant): The preovulatory LH surge sets in motion a chain of events that culminate in breakdown of the ovarian follicle wall and release of the egg in a process known as ovulation. Ovulation involves an inflammatory process that is characterized by a massive influx of white blood cells, known as leukocytes, to the ovary. This influx of leukocytes is critical for fertility as removal of leukocytes in experimental models blocks oocyte release. However, the factor(s) underlying the recruitment of leukocytes to the ovary are unknown. The present proposal will examine a newly identified candidate, chemokine ligand 20 (CCL20). Our preliminary data shows that mRNA for CCL20 is massively induced in the human during the early and late preovulatory stage in the granulosa and theca cells from the ovarian follicle. Yet other than our preliminary data, nothing is known as to temporal patterns of expression, the regulation, or function of CCL20 and its receptor CCR6 in the ovary. The present proposal builds on these provocative findings and will test the overall hypothesis that the LH surge stimulates an increase in CCL20 and the increase in this chemokine results in recruitment of leukocytes that impact events necessary for ovulation. This hypothesis will be addressed using a unique model where the granulosa and theca from human periovulatory follicles will be collected prior to and at three designated times after hCG to induce ovulation (early, late and postovulatory). These human samples will be compared and contrasted to changes in CCL20 and CCR6 in the rat ovary using a well characterized rodent model for ovulation. This will be accomplished by the following three Aims. Aim 1 will determine the changes in the expression of the CCL20 and CCR6 system in both the human and rat ovary. This Aim will also determine how CCL20 and CCR6 expression is regulated via the major ovulatory pathways induced by LH. Aim 2 will explore the role of CCL20 in leukocyte recruitment into the ovary using well-characterized migration and invasion models. Due to the potential role of CCL20 to regulate cellular differentiation, Aim 3 will investigate the role of CCL20 on ovarian cell function, specifically the ability of granulosa cells to survive, differentiate, and make sterid hormones. Additionally, the impact of CCL20 on ovarian cells will be assessed using microarray technology. The major strengths of the current proposal lie in the use of well characterized human preovulatory follicles as a foundation to understand the cellular expression, regulation, and the impact of CCL20 on the events associated with ovulation in the human and, our integrated comparative approach across species to elucidate the role that this chemokine plays in the ovulatory process. Nothing is known about CCL20 and its receptor in the ovary. As such, the proposed studies are extremely timely to elucidate the role that this chemokine system plays in the coordinated processes of follicular rupture and oocyte release which are fundamental aspects of normal human ovarian physiology.