Estrogen/GPR30 Modulation of Cardiac RAS Metabolism in Sex-Specific Hypertensive

项目3--项目摘要/摘要 高血压和左室舒张功能不全(LVDD)先于心力衰竭(HF)， 特别是在卵巢雌激素丢失的情况下(例如，更年期或卵巢早衰)。确实有 目前尚无有效的药物干预措施可延缓或逆转LVDD或随后的舒张期心衰。肾素- 血管紧张素系统(RAS)在高血压性心脏病的病理生理学中起着关键作用，但试验 提示血管紧张素转换酶抑制剂在接受高血压治疗的女性中可能不如男性有效， 高频。发现糜酶可以由Ang-(1-12)直接形成Ang II，而不依赖于ACE 为研究女性LVDD的发病机制开辟了新的方向。我们证明了慢性激活的 新的雌激素受体GPR30通过其激动剂G1减轻雌激素丢失对小鼠的不利影响 心脏LVDD表型和心脏糜酶表达减少，提示存在机制联系 在GPR30和当地RAS之间。我们的长期目标是了解凝乳酶/RAS的变化 雌激素丢失后的新陈代谢触发不适应途径，导致纤维化和左心室发育不良。的目标是 本项目旨在确定E2/GPR30限制肥大细胞糜酶作用的机制。 心肌细胞内Ang-(1-12)形成Ang II。我们假设这个表达式和 雌激素紊乱的高血压动物心脏中凝乳酶和Ang-(1-12)的功能较高 这些效应可以被雌激素(E2)替代和GPR30激活所逆转。 在强大的初步数据的指导下，我们将在三个目标上检验我们的假设：1)表征 自分泌/旁分泌酶途径导致心脏Ang II的形成、LVDD和心脏重构 雌激素丢失；2)确定雌激素是否激活GPR30，而不是经典雌激素的激活 受体ER？和ER？限制心脏糜酶/Ang-(1-12)代谢以保护左心室结构和 功能；以及3)确定E2/GPR30负性调节乳糜酶产生/释放的特定机制 肥大细胞和糜酶介导的心肌细胞Ang-(1-12)代谢/Ang II的产生。我们的 创新的全球系统生物学方法综合了(A)生理、生化、细胞和 分子方法学；(B)转基因啮齿动物和小鼠模型；(C)培养的肥大细胞和 (D)GPR30-、ER？和ER？基因使体外培养的细胞沉默。证实了我们的假设 将极大地促进我们对肥大细胞糜酶、Ang-(1-12)和Ang II表达的理解 和功能可能受E2通过GPR30调节，并提供了一种解释RAS抑制剂为什么 对雌激素后女性血管紧张素转换酶II介导的不良心脏重塑和左室舒张功能的阻断效果较差 损失。

Project 3 - Project Summary / Abstract Hypertension and left ventricular diastolic dysfunction (LVDD) precede heart failure (HF) in women, particularly with the loss of ovarian estrogens (e.g., with menopause or premature ovarian failure). There are no proven pharmacologic interventions that delay or reverse LVDD or subsequent diastolic HF. The renin- angiotensin system (RAS) plays a key role in the pathophysiology of hypertensive heart disease, but trials suggest ACE inhibitors may be less effective in women than men receiving treatment for hypertension and HF. The discovery that chymase can form Ang II directly from Ang-(1-12), independent of ACE, has opened up a new direction in research on the pathogenesis of LVDD in women. We showed that chronic activation of the novel estrogen receptor GPR30 by its agonist G1 mitigated the adverse effects of estrogen loss on the cardiac LVDD phenotype and reduced expression of cardiac chymase, suggesting a mechanistic link between GPR30 and the local RAS. Our long-term goal is to understand how changes in chymase/RAS metabolism after estrogen loss triggers maladaptive pathways leading to fibrosis and LVDD. The objective of this project is to determine the mechanism by which E2/GPR30 limits the action of mast cell chymase on intracellular Ang II formation from Ang-(1-12) in cardiomyocytes. We hypothesize that expression and function of chymase and Ang-(1-12) are higher in the hearts of hypertensive animals with disrupted estrogen activity, and that these effects can be reversed with estrogen (E2) replacement and GPR30 activation. Guided by strong preliminary data, we will test our hypothesis in three aims: 1) Characterize the autocrine/paracrine enzymatic pathway leading to cardiac Ang II formation, LVDD, and remodeling after estrogen loss; 2) Determine if E2 activation of GPR30, as opposed to activation of the classic estrogen receptors ER¿ and ER¿, limits cardiac chymase/Ang-(1-12) metabolism to preserve LV structure and function; and 3) Identify E2/GPR30-specific mechanisms that negatively regulate chymase production/release from mast cells and chymase-mediated Ang-(1-12) metabolism/Ang II production in cardiomyocytes. Our innovative global systems biology approach integrates the use of (a) physiological, biochemical, cellular, and molecular methodologies; (b) transgenic rodent and murine models; (c) cultured mast cells and cardiomyocytes; and (d) GPR30-, ER¿-, and ER¿-gene silenced cells in vitro. Confirmation of our hypothesis will significantly advance our understanding of how mast cell chymase, Ang-(1-12), and Ang II expression and function may be regulated by E2 via GPR30, and provide a mechanism that explains why RAS inhibitors are less effective in blocking Ang II-mediated adverse cardiac remodeling and LVDD in women after estrogen loss.