Role of mineralocorticoid receptor in primate ovarian function

描述(由申请人提供)：灵长类黄体的形成、功能和消亡是月经周期、不孕症和避孕干预的关键方面。申请人的实验室最近的研究表明，在非人类灵长类动物中存在卵巢内的矿物皮质激素系统。在进行控制性卵巢刺激的恒河猴中，排卵刺激以时间依赖的方式增加21-羟基酶(CYP21A2)mRNA的表达。CYP21A2将孕酮转化为有效的盐皮质激素受体(MR)激动剂11-脱氧皮质酮(DOC)，因此与观察到排卵hCG丸增加卵泡内DOC水平有关。为了证实DOC来自卵巢，在排卵刺激前分离颗粒细胞，并在hCG存在的情况下进行培养，导致DOC的培养液浓度显著增加。MR在黄体化颗粒细胞和黄体的细胞质中均有表达，直到黄体中期和中晚期，此时MR被转移到细胞核。重要的是，黄体化过程中MR的拮抗作用减弱了hCG诱导的孕酮，表明细胞质MR在体外颗粒细胞黄体化过程中起着关键作用。这个R21应用程序建议使用非人类灵长类动物模型来检验MR在黄体生命周期中至关重要的总体假设，并具有两个特定的目标。具体目标1将使用MR拮抗剂确定MR是否在黄体发育中起关键作用。预计体内MR的拮抗作用将阻止孕酮的临界上升，导致无排卵和黄体形成不完全。《特定目标2》将验证这样一种假设，即MR通过孕酮合成促进黄体功能，黄体内DOC水平与黄体内孕酮水平同步增加。在这两个目标中，MR的特定基因靶点将被阐明。这些研究将首次证明MR在任何物种的卵巢生理中的关键作用，并有望加深我们对黄体在生物医学相关灵长类动物模型中的发育、维持和死亡的理解。公共卫生相关性：与卵巢相关的病理具有很高的发病率和死亡率，这主要是由于难以发现，但也由于对卵巢基本过程的相当不完全的了解而缺乏治疗。这些疾病包括多囊卵巢综合征、卵巢癌和卵巢过度刺激综合征(OHSS)。此外，不孕症困扰着美国越来越多的夫妇(600万；疾病控制中心)；这种不孕不育的一部分可能与黄体期P分泌不足有关，尽管其根本原因尚不清楚。新的避孕方法仍然是一个问题，很大一部分北美妇女继续依赖于联合口服避孕药的衍生产品，而且看不到新的方法。在美国，衰老和生殖显然是一个越来越多的话题，更好地了解基本的卵巢过程将有助于理解女性的更年期转变。这些问题都支持这样一种观点，即卵巢生理学是一个相关的话题，具有深远的健康后果。总体而言，这些探索性研究应该揭示灵长类卵巢中一种新的信号类固醇途径，这可能对避孕发育、不孕和潜在的卵巢癌具有临床意义。

DESCRIPTION (provided by applicant): The formation, function, and demise of the primate corpus luteum are critical aspects of menstrual cyclicity, infertility, and contraceptive intervention. Recent work by the applicant's laboratory has demonstrated the presence of an intra-ovarian mineralocorticoid system in the non-human primate. In rhesus monkeys undergoing controlled ovarian stimulation, an ovulatory stimulus increases the expression of 21-hydroxylase (CYP21A2) mRNA in a time-dependent manner. CYP21A2 converts progesterone to the potent mineralocorticoid receptor (MR) agonist 11-deoxycorticosterone (DOC), and thus correlates with the observation that the ovulatory hCG bolus increases intra-follicular levels of DOC. In order to verify an ovarian origin for DOC, granulosa cells were isolated prior to an ovulatory stimulus and cultured in the presence of hCG, resulting in a dramatic increase in media concentrations of DOC. MR is expressed in the cytoplasm of both luteinizing granulosa cells as well as the corpus luteum until the mid to mid-late luteal phase, at which time MR is translocated to the nucleus. Importantly, antagonism of MR during luteinization attenuates hCG-induced progesterone, indicating that cytoplasmic MR has a key role in luteinization of granulosa cells in vitro. This R21 application proposes to test the overall hypothesis that MR is critical during the lifespan of the corpus luteum with two specific aims using the non-human primate model. Specific aim 1 will determine if MR has a critical role in the development of the corpus luteum using an MR antagonist. It is expected that antagonism of MR in vivo will prevent the critical rise in progesterone, leading to anovulation and incomplete luteal formation. Specific aim 2 will test the hypothesis that MR promotes luteal function through progesterone synthesis, and that intra-luteal levels of DOC increase in parallel with progesterone. In both aims, specific gene targets of MR will be elucidated. These studies will be the first to demonstrate a key role for MR in the ovarian physiology of any species, and are expected to further our understanding of corpus luteum development, maintenance, and demise in a biomedically relevant primate model. PUBLIC HEALTH RELEVANCE: Pathologies associated with the ovary carry a high morbidity and mortality, largely through difficulty in detection, but also from a lack of treatment stemming from a rather incomplete understanding of basic ovarian processes. These include, for example, PCOS, ovarian cancer, and ovarian hyperstimulation syndrome (OHSS). In addition, infertility plagues a growing number of couples in the United States (>6 million; Center for Disease Control); a proportion of this infertility can be linked to insufficiency of P production during the luteal phase, although the underlying causes of this are completely unknown. Novel contraception remains a problem, with a large preponderance of North American women continuing to rely on derivations of combined oral contraception and no new methods in sight. Aging and reproduction is clearly a growing topic in the United States, and a better knowledge of basic ovarian processes will facilitate understanding the menopausal transition in women. These issues all support the idea that ovarian physiology is a relevant topic with profound health outcomes. Overall, these exploratory studies should reveal a novel signaling steroid pathway in the primate ovary that could have clinical significance for contraceptive development, infertility, and potentially ovarian cancer.