Role of Endothelial Anoctamin-1 in Pulmonary Arterial Hypertension

内皮 Anoctamin-1 在肺动脉高压中的作用

• 批准号: 10200883
• 负责人: Gaurav Choudhary
• 金额: $ 40.54万
• 依托单位: OCEAN STATE RESEARCH INSTITUTE, INC.
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2023-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10200883
• 关键词: Animal Model; Animals; Apoptosis; Apoptotic; Attenuated; Blood Vessels; Calcium; Cell Line; Cell Proliferation; Cell Proliferation Regulation; Cell membrane; Cells; Chloride Channels; Chlorides; Conflict (Psychology); DNA; DNA Methylation; Disease; Dissociation; Endothelial Cells; Endothelium; Epidermal Growth Factor Receptor; Epigenetic Process; Foundations; Growth; In Vitro; Inhalation; Ion Channel; Lesion; Lung; Malignant Neoplasms; Medial; Mediating; Methylation; Mitochondria; Morbidity - disease rate; OPA1 gene; Optic Atrophy; Outcome; Pathology; Patients; Pharmacology; Phenotype; Pre-Clinical Model; Proteins; Pulmonary Hypertension; Pulmonary artery structure; Rattus; Regulatory Element; Resistance; Role; Signal Pathway; Signal Transduction; Site; Testing; Therapeutic; Tissues; Vascular remodeling; cell motility; design; effective therapy; improved; in vitro testing; in vivo; methylation pattern; mortality; novel; novel therapeutic intervention; overexpression; preclinical study; pressure; prevent; pulmonary arterial hypertension; response; small molecule; treatment strategy; vasoconstriction

ABSTRACT Pulmonary arterial hypertension (PAH) is associated with significant morbidity and mortality. PAH pathology includes vasoconstriction, medial and adventitial remodeling, and microvascular endothelial cell (EC) proliferation leading to vaso-occlusive plexiform lesions. EC in PAH are both apoptosis resistant and hyperproliferative. There are no effective treatments for the severe vascular remodeling observed with PAH, and therapeutic strategies that target the dysfunctional EC and plexiform lesions are urgently needed. We have discovered that calcium activated chloride channel, Ano1, is localized to both the EC plasma membrane (pl-Ano1) and mitochondria (mito-Ano1). Ano1 expression is upregulated in EC in settings of PAH and associated with a hyperproliferative and apoptosis resistant phenotype. However, unregulated activation of this channel results in apoptosis of hyperproliferative apoptosis-resistant ECs. Our overall objective in this proposal is to delineate the mechanisms underlying these seemingly conflicting observations to identify therapeutic opportunities to decrease EC proliferation and target hyperproliferative ECs for apoptosis, thereby improving pulmonary vascular remodeling in PAH. We hypothesize that epigenetically regulated increased expression of Ano1 results in EC proliferation and apoptosis resistance, yet unregulated opening of Ano1 in the context of overexpression is detrimental and can potentially be exploited to promote apoptosis of hyperproliferative ECs in PAH. In Aim 1, we will determine the mechanism of increased Ano1 expression in PAH EC with a focus on DNA methylation. We will use complementary in vitro and in vivo approaches to design novel constructs to selectively alter the methylation pattern of Ano1 regulatory elements in lung EC and test if this strategy prevents or treats PAH. In Aim 2, we will determine the effect of increased pl- and mito- Ano1 expression on EC proliferation and apoptosis resistance and elucidate the underlying mechanisms. We will investigate the signaling pathways in vitro and then use selective inhibition of Ano1 expression in lung EC in vivo to determine its effect on preventing or treating PAH. In Aim 3, we will determine the mechanism of the effect of pl- and mito-Ano-1 opening on EC apoptosis in vitro and test if opening of Ano1 by a small molecule delivered by inhalation attenuates PAH. As a result of these studies we expect to delineate the mechanism of Ano1-mediated EC proliferation and apoptosis resistance in settings of PAH as well as develop novel therapeutic strategy targeting Ano1 to improve pulmonary vascular remodeling and PAH.

抽象的 肺动脉高压（PAH）与明显的发病率和死亡率有关。 PAH病理学 包括血管收缩，内侧和外膜重塑以及微血管内皮细胞（EC） 增殖导致血管囊化的丛状病变。 PAH中的EC均具有抗凋亡性和 超增量。对于用PAH观察到的严重血管重塑没有有效的治疗方法 迫切需要针对功能失调的EC和神经损伤的治疗策略。我们 已经发现钙活化的氯化物通道ANO1位于EC质膜上 （PL-ANO1）和线粒体（mito-ano1）。在PAH和 与耐药性表型相关。但是，不受监管的激活 通道导致抗凋亡抗凋亡的ECS的细胞凋亡。我们的整体目标 提案是描述这些看似相互矛盾的观察的机制，以识别 减少EC增殖并靶向凋亡的靶向超增殖的治疗机会，从而 改善PAH中的肺血管重塑。我们假设表观遗传调节增加 ANO1的表达会导致EC增殖和凋亡耐药性，但在ANO1中的开放不受监管 过表达的背景是有害的，有可能被利用以促进 PAH中的超增量EC。在AIM 1中，我们将确定在ANO1表达增加的机制 PAH EC以DNA甲基化为重点。我们将使用互补的体外和体内方法 设计新颖的结构，以选择性地改变肺EC和 测试此策略是否阻止或治疗PAH。在AIM 2中，我们将确定PL和Mito-增加的影响 ANO1在EC增殖和凋亡耐药性上的表达并阐明了潜在的机制。我们 将在体外研究信号通路，然后在肺EC中使用ANO1表达的选择性抑制 在体内确定其对预防或治疗PAH的影响。在AIM 3中，我们将确定 PL-和MITO-ANO-1开放对体外EC凋亡的影响，并测试小分子开放ANO1是否打开 通过吸入递送减弱PAH。由于这些研究，我们期望描述 ANO1介导的EC增殖和PAH环境中的凋亡耐药性以及发展新颖 针对ANO1的治疗策略以改善肺血管重塑和PAH。