Role of nuclear progesterone receptors in the control of human parturition

描述（由申请方提供）：早产是新生儿发病率和死亡率的主要原因，预防早产的治疗方法的开发需要清楚地了解将子宫肌层从松弛状态转变为分娩状态的激素相互作用。在这种情况下，孕激素是一个主要的参与者，因为它促进子宫肌层松弛的大部分怀孕，它的撤退启动劳动，和孕激素预防降低早产的发生率在一个子集的妇女。拟议的研究通过确定孕酮如何通过其与受体PR-A和PR-B的相互作用来影响人类子宫肌层细胞中的基因表达，从而检查孕酮如何发挥这些作用。我们发表的数据表明，孕酮通过PR-B促进子宫肌层松弛，分娩是由子宫肌层细胞表达PR-A增加引起的，PR-A通过抑制PR-B的转录活性引起功能性孕酮戒断。为了验证这一 假设我们开发了人子宫肌层细胞系以检测PR-A和PR-B如何影响孕酮的基因组作用。我们发现孕酮具有抗炎作用，并抑制PR-B优势细胞中促炎基因的表达，但在PR-A优势细胞中具有相反的作用。这种作用似乎是通过PR对促炎基因启动子的直接作用和核因子-<$B（NF <$B）活性的调节介导的;特别是通过I <$B <$（NF <$B的主要抑制因子）表达的增加。这些发现很重要，因为分娩被认为是子宫肌层炎症的后遗症。因此，我们认为孕酮通过PR-B抑制NF-κ B介导的促炎基因表达促进子宫肌层松弛，而通过PR-A介导的促炎基因表达上调促进分娩。为了验证这一假设，我们将：1）确定孕酮如何通过其与PR-A和PR-B的相互作用影响人子宫肌层细胞中前列腺素（PG）-内过氧化物合酶2（PTGS 2;选择为原型促分娩/促炎基因）和I B的基础和马槟榔碱诱导的表达（特异性目的1）。我们将描述PR-A和PR-B在PTGS 2和I B启动子处相互作用的机制，以及它们如何调节NF-B活性。我们的研究中出现的另一个重要概念是，分娩相关的子宫肌层细胞PR-A表达增加，这是人类分娩的关键事件，由PGF 2？上调。具体目标2将是确定细胞内信号通路和顺式/反式调节因子，通过这些通路PGF 2？影响人子宫肌层细胞中PR-A的表达。PGF 2 <$通过其受体影响PR-A表达的信号通路和PR-A启动子中的顺式元件以及作用于PGF 2 <$诱导PR-A表达的那些元件的转录因子将被鉴定。这两个目标的实验将在人子宫肌层细胞系中进行，包括我们的表达PR-A和PR-B以响应独立诱导剂的细胞系，以及子宫肌层外植体和足月子宫肌层的原代肌细胞培养物。这项研究将促进对孕酮如何控制人类妊娠和分娩的理解，并提供开发/改善基于孕酮的治疗以预防早产所需的知识。 公共卫生相关性：早产是一个主要的社会经济问题，影响10-15%的怀孕，并造成70-80%的新生儿死亡和发病率。尽管新生儿护理的进步提高了存活率，但由于早产而引起的混淆问题深刻地影响了早产儿未来的健康。显然，胎儿发育的最后阶段最好在子宫环境中完成，而不是在新生儿重症监护室。为此，我们必须预防和/或抑制早产。然而，目前抑制早产的疗法通常无效。为了解决这个问题，我们必须填补知识空白，限制我们的能力，以开发有效的治疗早产。为此，我们的目标是阐明在人类怀孕期间控制子宫收缩力的激素相互作用。在这种情况下，类固醇激素孕酮是至关重要的。该建议解决了核孕酮受体PR-A和PR-B介导人子宫肌层细胞中孕酮作用的分子机制。对这一过程的清楚理解可能会揭示抑制早产和预防早产的新的治疗靶点。

DESCRIPTION (provided by applicant): The development of therapies to prevent preterm birth, which is the major cause of neonatal morbidity and mortality, requires a clear understanding of the hormonal interactions that transform the myometrium from the relaxed to the laboring state. In this context, progesterone is a major player because it promotes myometrial relaxation for most of pregnancy, its withdrawal initiates labor, and progestin prophylaxis decreases the incidence of preterm birth in a subset of women. The proposed research examines how progesterone exerts these actions by determining how, via its interaction with its receptors, PR-A and PR-B, it affects gene expression in human myometrial cells. Our published data suggest that progesterone promotes myometrial relaxation via PR-B and that labor is initiated by increased myometrial cell expression of PR-A, which causes functional progesterone withdrawal by inhibiting the transcriptional activity of PR-B. To test this hypothesis we developed a human myometrial cell line to examine how PR-A and PR-B affect genomic actions of progesterone. We found that progesterone was anti-inflammatory and inhibited the expression of pro- inflammatory genes in PR-B-dominant cells but had the opposite effect in PR-A-dominant cells. The effects appeared to be mediated by direct actions of the PRs at the promoters of pro-inflammatory genes and by modulation of nuclear factor -¿B (NF¿B) activity; especially by increased expression of I¿B¿, a major repressor if NF¿B. These findings are important because labor is thought to be sequelae of myometrial inflammation. Thus we propose that progesterone via PR-B promotes myometrial relaxation by inhibiting NF-¿B-mediated pro-inflammatory gene expression, whereas it promotes labor via PR-A-mediated up-regulation of pro- inflammatory gene expression. To test this hypothesis we will: 1) determine how progesterone via its interaction with PR-A and PR-B affects basal and cytokine-induced expression of prostaglandin (PG) - endoperoxide synthase 2 (PTGS2; chosen as a prototypical pro-labor/pro-inflammatory gene) and I¿B¿ in human myometrial cells (Specific Aim 1). We will characterize the mechanism by which PR-A and PR-B interact at the PTGS2 and I¿B¿ promoters, and how they may modulate NF-¿B activity. Another important concept to arise from our research is that the parturition-associated increase in myometrial cell PR-A expression, which is a key event in human parturition, is up-regulated by PGF2¿. Specific Aim 2 wil be to identify the intracellular signaling pathways and cis/trans regulators through which PGF2¿ affects PR-A expression in human myometrial cells. The signaling pathways through which PGF2¿ via its receptor affects PR-A expression and the cis elements in the PR-A promoter and the transcription factors acting on those elements through which PGF2¿ induces PR-A expression will be identified. Experiments for both aims will be performed in human myometrial cell lines, including our line that expresses PR-A and PR-B in response to independent inducers, and myometrial explant and primary myocyte cultures of term myometrium. The research will advance understanding of how progesterone controls human pregnancy and parturition and provide knowledge needed to develop/improve progestin-based therapies to prevent preterm birth. PUBLIC HEALTH RELEVANCE: Preterm birth is a major socioeconomic problem that affects 10-15% of pregnancies and causes 70-80% of neonatal mortality and morbidity. Despite advances in neonatal care that have improved survival outcomes, the confounding problems due to preterm birth profoundly affect a preterm infant's future health. Clearly, the final phase o fetal development is best achieved in the womb environment rather than in the neonatal intensive care unit. To this end, we must prevent and/or suppress preterm labor. However, current therapies to suppress preterm labor are generally ineffective. To solve this problem we must fill the knowledge-gaps that limit our capacity to develop effective treatments for preterm labor. To this end, our goal is to elucidate the hormonal interactions that control uterine contractility during human pregnancy. In this context the steroid hormone progesterone is critical. This proposal addresses the molecular mechanisms by which the nuclear progesterone receptors PR-A and PR-B mediate progesterone actions in human myometrial cells. A clear understanding of this process may reveal novel therapeutic targets for the suppression of preterm labor and the prevention of preterm birth.