Vascular AT2R expression and function during pregnancy

摘要 在怀孕期间，血管内皮细胞的扩张得到增强，从而降低血压，增加子宫血流量。 妊娠增强的血管扩张功能是通过增加血管紧张素2型的表达来实现的 子宫和全身血管内皮细胞的AT2R受体。在先兆子痫患者中，这种适应性 由于AT2R表达和活性下调而导致的内皮反应失败，与相关的内皮细胞 功能障碍，血管阻力增加，以及高血压。先兆子痫影响所有妊娠的5%-8%， 治疗仅限于松弛血管平滑肌的抗高血压药物。没有任何治疗方法可以治疗 直接解决内皮细胞故障。先兆子痫的原因尚不清楚，但特定于妊娠 AT2R在血管系统的上调可能是了解疾病起源的重要关键 以及母体器官功能的潜在原因。研究新奇口服液效果的机会 非肽成分21具有重要意义和应用价值。基于可靠的初步数据，我们认为 妊娠通过与配体激活的雌激素的差异结合选择性上调血管内皮细胞AT2R 受体(即ERβ与ERα)结合到AT2R启动子中的功能性雌激素反应元件(ERE)。故障： 妊娠期血管AT2R上调在血管内皮细胞功能障碍和 先兆子痫的血管收缩，因此，该系统的激活逆转了先兆子痫的血管 功能障碍。目标1将首先建立妊娠特异性的血管内皮细胞AT2R上调，然后 通过使用ER特异性激动剂、拮抗剂和α，确定ER、β或AT2R在上调AT2R中的机制作用 啮齿动物模型中的siRNA。AT2R转录机制将决定配体的差异结合- 人子宫动脉AT2R启动子ER和转录因子对ERE的激活作用 使用芯片和EMSA/SuperShift分析，然后使用报告分析来确定其 功能性。目标2将走向临床翻译相关性。我们将确定血管AT2R是否 参与子痫前期血管内皮细胞功能障碍，如果AT2R激活可挽救血管功能障碍。为了测试 AT2R介导的血管松弛通路的机制、EDHF、NO和PGI2通路将是 下定决心。内皮型一氧化氮合酶及其活性状态信号转导成分EDHF和PGI2的表达 途径以及硝酸盐/亚硝酸盐和PGI2的产生以及膜电位的变化-将 量过了。AIM 3将在体内验证AT2R效应。我们将使用两种妊娠期高血压模型来测试 两种选择性血管紧张素Ⅱ受体激动剂(CGp-42112肽和化合物21)对母体、胎盘和 与先兆子痫相关的胎儿异常。这些研究将提供怀孕的新信息 增加血管内皮细胞AT2R，这一过程的失败会导致子痫前期内皮功能障碍。这个 这些目标的积极翻译意义在于，它们评估了内皮的治疗效用- 靶向治疗在先兆子痫患者中可能是安全的。

ABSTRACT In pregnancy, endothelial vasodilation is enhanced to decrease blood pressure and increase uterine blood flow. Pregnancy-enhanced vasodilatory function is achieved through an increase in expression of angiotensin type 2 receptor (AT2R) in the endothelium of uterine and systemic vasculature. In preeclampsia, this adaptive endothelial response fails due to downregulation of AT2R expression and activity, with associated endothelial dysfunction, increased vascular resistance, and hypertension. Preeclampsia affects 5%–8% of all pregnancies, and treatment is limited to antihypertensives that relax vascular smooth muscle. There is no treatment to directly address endothelial failure. The cause of preeclampsia is not known, but the pregnancy-specific upregulation of AT2R in the vasculature may hold important keys to understanding the origins of the disease and the underlying causes of maternal organ function. The opportunity to study the effects of novel oral nonpeptide Compund21 is significant and valuable. Based on solid preliminary data, we propose that pregnancy selectively upregulates endothelial AT2R via differential binding of ligand-activated estrogen receptor (ie, ERβ vs ERα) to a functional estrogen response element (ERE) in the AT2R promoter. Failure of vascular AT2R upregulation during pregnancy plays a role in endothelial cell dysfunction and vasoconstriction in preeclampsia and, consequently, activation of this system reverses preeclamptic vascular dysfunction. Aim 1 will first establish pregnancy-specific upregulation of AT2R in the endothelium and then define the mechanistic role of ER α or β in upregulation of AT2R by using ER-specific agonists, antagonists, and siRNA in rodent models. AT2R transcription mechanisms will determine differential binding of ligand- activated ER and transcription factors to putative ERE in AT2R promoter in primary human uterine artery endothelial cells, using ChiP and EMSA/ supershift assays, and then use reporter assays to determine their functionality. Aim 2 will move towards clinical translational relevance. We will determine if vascular AT2R is involved in preeclamptic endothelial dysfunction and if AT2R activation rescues vascular dysfunction. To test the AT2R-mediated mechanisms, EDHF, NO, and PGI2 pathways of vascular relaxation pathways will be determined. Also, the expression of eNOS and its activity state—signaling components of EDHF and PGI2 pathways as well as nitrate/ nitrite and PGI2 production and changes in membrane potential—will be measured. Aim 3 will verify the AT2R effects in vivo. We will use 2 models of gestational hypertension to test the effect of 2 selective AT2R agonists (CGP-42112 peptide and Compound 21) on the maternal, placental, and fetal abnormalities associated with preeclampsia. These studies will provide new information of how pregnancy increases endothelial AT2R and that failure of this process leads to preeclamptic endothelial dysfunction. The positive translational significance of these aims is that they evaluate the therapeutic utility of an endothelium- targeted therapy which is potentially safe in preeclampsia subjects.