Estrogen receptor-alpha effects on right ventricular vascular density and angiogenesis in pulmonary hypertension

在退伍军人中，肺动脉高压(PH)和右室(RV)功能障碍是非常常见的。至.为止 80%患有慢性阻塞性肺疾病、肺纤维化、睡眠呼吸障碍或 左心功能不全(无论是收缩功能障碍还是舒张期功能障碍)都受到PH的影响。更好的房车功能和女性性行为联系在一起 对于提高PH患者的存活率，女性患者表现出比男性患者更好的RV功能。这 该提案建立在一个科学前提上，即尽管房车功能和女性性别是主要决定因素 对于在PH中存活的患者，没有RV特异性或性激素导向的治疗方法。内皮细胞(EC)功能障碍和 血管生成受损在RV衰竭的发展中起着主要作用，而在之前的研究中获得的数据 资助期显示，雌性性类固醇17β-雌二醇(E2)增加了RV的毛细血管密度 并刺激培养的心脏内皮细胞血管生成。这项提案的目标是确定新的和 E_2保护肺血管内皮细胞功能的治疗靶向机制。我们 提供证据证明E2通过其受体ERα发挥其RV EC保护作用，并建议一种新的 雌激素受体α激活骨形态发生蛋白受体2信号上调的机制 Apelin是一种有效的血管生成介质和EC生存因子，其在RV中的调节尚不清楚。 基于这些发现，我们现在提出了一个新的假设，即E2通过 ER、α和BMPR2依赖上调EC apelin。我们提出以下具体目标：1) 确定E2通过BMPR2依赖的EC apelin的增加增加RV的毛细血管密度，以及2) 目的：探讨ER-α在增加右室毛细血管密度中的作用。我们产生了一种新的ERα 这将使我们能够研究ERα在大鼠肺动脉结扎模型中的作用。 避免了之前在无RV衰竭的PH模型中进行的性激素信号转导研究的陷阱。 这些研究将得到对BMPR2缺陷大鼠和apelin缺陷小鼠的研究的补充。这些损失- 功能研究将伴随着我们询问ERα的治疗潜力的研究 激动剂、BMPR2激动剂和apelin受体激动剂。我们将补充这些活体研究 从RV衰竭的啮齿动物和代偿性(适应性)或慢性阻塞性肺疾病患者分离的RV ECs的实验 失代偿性(适应不良)的RV肥大。调查的终端将包括房车功能和结构 (通过压力-容积环和超声心动图)、运动能力(通过跑步机测量最大摄氧量 跑步)、右室毛细血管密度(使用无偏体视学和凝集素染色进行量化)、血管生成分析 (基底管形成和井间运移)、BMPR2和apelin信号通路以及介体 血管生成与EC存活和凋亡的关系。拟议的研究意义重大，因为它们将1)确定 ER-α作为RV功能的关键调节剂和2)建立一种新型的、具有治疗靶向性的E2-ERα- 右室内皮细胞中的BMPR2-apelin轴。拟议的研究具有创新性，因为它们将首次提供一种 E_2‘S RV和EC在PH中保护作用的分子基础。此外，它们还提供技术创新 通过使用新产生的ERα基因敲除大鼠模型，通过使用新的高选择性ERα激动剂 这将允许从失败的房车中机械地剖析ERα的S在EC中的角色，并通过使用公正的 体视学。在建议的研究完成后，我们将确定ERα是一种新的 房车内皮细胞中的自适应信号。这可能最终允许开发新的房车导向的，非 对患有PH和RV衰竭的女性和男性退伍军人进行激素治疗。

Pulmonary hypertension (PH) and right ventricular (RV) dysfunction are extremely common in veterans. Up to 80% of veterans with chronic obstructive pulmonary disease, pulmonary fibrosis, sleep disordered breathing or LV dysfunction (either systolic or diastolic) suffer from PH. Better RV function and female sex have been linked to improved survival in PH, and female patients exhibit better RV function than their male counterparts. This proposal builds on the scientific premise that even though RV function and female sex are major determinants of survival in PH, no RV-specific or sex steroid-directed therapies exist. Endothelial cell (EC) dysfunction and impaired angiogenesis play a major role in the development of RV failure, and data obtained in the previous funding period demonstrate that the female sex steroid 17β-estradiol (E2) increases capillary density in the RV and stimulates angiogenesis in cultured cardiac ECs. The goal of this proposal is to identify novel and therapeutically targetable mechanisms by which E2 exerts protective effects on RV EC function in PH. We provide evidence that E2 exerts its RV EC-protective effects via its receptor ERα, and suggest a new mechanism by which ERα activates bone morphogenetic protein receptor 2 (BMPR2) signaling to upregulate apelin, a potent angiogenesis mediator and EC survival factor, whose regulation in the RV is not yet known. Based on these findings, we now put forward the novel hypothesis that E2 improves RV function in PH by ERα- and BMPR2-dependent up-regulation of EC apelin. We propose the following specific aims: 1) To establish that E2 increases capillary density in the RV via BMPR2-dependent increases in EC apelin, and 2) To identify the contribution of ERα to increasing capillary density in the RV. We generated a novel ERα knockout rat that will enable us to study the role of ERα in the rat pulmonary artery banding model, thus avoiding the pitfalls of prior studies of sex hormone signaling performed in PH models without RV failure. These studies will be complemented by studies of BMPR2-deficient rats and apelin-deficient mice. These loss- of-function studies will be accompanied by studies in which we interrogate the therapeutic potential of ERα agonists, BMPR2 activators and apelin receptor agonists. We will complement these in vivo studies with experiments in RV ECs isolated from rodents with RV failure and from patients with compensated (adaptive) or decompensated (maladaptive) RV hypertrophy. Endpoints investigated will include RV function and structure (by pressure volume loops and echocardiography), exercise capacity (measured as VO2 max via treadmill running), RV capillary density (quantified using unbiased stereology and lectin staining), angiogenesis assays (matrigel tube formation and transwell migration), BMPR2 and apelin signaling pathways, as well as mediators of angiogenesis and EC survival and apoptosis. The proposed studies are significant, since they will 1) identify ERα as a critical modulator of RV function and 2) establish a novel and therapeutically targetable E2-ERα- BMPR2-apelin axis in RV ECs. The proposed studies are innovative, since they, for the first time, will provide a molecular basis for E2's RV- and EC-protective effects in PH. In addition, they provide technical innovation through use of a newly generated ERα knockout rat model, through use of a new highly selective ERα agonist that will allow for mechanistic dissection of ERα’s role in ECs from failing RVs, and through use of unbiased stereology. Upon completion of the proposed studies, we will have identified ERα as a novel mediator of adaptive signaling in RV ECs. This may ultimately allow for the development of new RV-directed, non- hormonal treatments for both female and male veterans with PH and RV failure.