NFATc3 in chronic hypoxic pulmonary hypertension

NFATc3 在慢性缺氧性肺动脉高压中的作用

• 批准号: 7746406
• 负责人: Laura V Gonzalez Bosc
• 金额: $ 37.5万
• 依托单位: UNIVERSITY OF NEW MEXICO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-12-01 至 2012-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7746406
• 关键词: Abbreviations; Actins; Address; Agonist; Altitude; Animal Model; Asthma; Binding; Biology; Blood Vessels; Blood flow; Calcineurin; Calcium; Cattle; Cause of Death; Cell Differentiation process; Cessation of life; Chronic; Chronic Bronchitis; Collagen Diseases; Complement; Complex; Contractile Proteins; Cyclosporine; Cystic Fibrosis; Data; Development; Diagnosis; Disease; Down-Regulation; Endothelin-1; Event; Failure; Family; Fibroblasts; Figs - dietary; Gene Expression; Genes; Genetic; Genetic Programming; Genetic Transcription; Genomics; Goals; Heart Diseases; Heart failure; Hyperplasia; Hypertrophy; Hypoxia; Incidence; Ischemia; Knockout Mice; Lead; Left; Life; Link; Luciferases; Lung; Lung diseases; Maintenance; Measures; Medial; Mediating; Mediator of activation protein; Membrane Potentials; Messenger RNA; Methods; Molecular; Molecular Biology; Morbidity - disease rate; Mus; Muscle Contraction; Myosin Heavy Chains; National Heart, Lung, and Blood Institute; Nuclear; Pathology; Patients; Pattern; Phenotype; Physiological; Physiology; Plasmids; Play; Polycythemia; Potassium; Prevalence; Principal Investigator; Process; Progressive Disease; Protein Isoforms; Proteins; Pulmonary Emphysema; Pulmonary Hypertension; Pulmonary Vascular Resistance; Pulmonary artery structure; Quality of life; Regulation; Reporter; Reporting; Research; Resistance; Rheumatologic Disorder; Rho-associated kinase; Right Ventricular Hypertrophy; Rodent; Role; Side; Signal Pathway; Signal Transduction; Smooth Muscle; Smooth Muscle Myocytes; Suggestion; System; T-Cell Activation; Testing; Thick; Time; United States; Up-Regulation; Vascular Diseases; Vascular Smooth Muscle; Vascular remodeling; Vasoconstrictor Agents; Ventricular; alpha Actin; arterial remodeling; arteriole; base; design; improved; in vivo; inhibitor/antagonist; member; mortality; muscle hypertrophy; novel; novel therapeutic intervention; nuclear factors of activated T-cells; pressure; prevent; primary pulmonary hypertension; programs; promoter; protein expression; pulmonary arterial hypertension; research study; response; sudden cardiac death; transcription factor; transcription factor NF-AT c3; treatment strategy; vasoconstriction; voltage

DESCRIPTION (provided by applicant): Despite the origin of pulmonary arterial hypertension (PAH), pulmonary vascular resistance rises due to pulmonary vasoconstriction, arterial remodeling and polycythemia leading to right heart failure and death. Rodents exposed to chronic hypobaric hypoxia (CH) develop PAH. The complex process of developing PAH is driven, in part, by changes in gene expression. In PAH, smooth muscle intracellular Ca2+ is increased and endothelin 1 (ET-1) expression is up-regulated. Ca2+ regulates pulmonary arterial smooth muscle (PASMC) contraction and is linked to gene transcription through the nuclear factor of activated T cells (NFAT). NFATc3 isoform is specifically implicated in the development of the vasculature and maintenance of smooth muscle differentiate phenotype. The overall goal of this proposal is to determine the role of NFATc3 in the molecular mechanisms underlying the vascular changes associated with CH-PAH. The hypothesis is that CH activates NFATc3 in PASMC to mediate hypertrophy and enhance contractility of pulmonary arteries (PA) contributing to PAH. Specific Aim 1: To determine the role of NFATc3 in CH-induced PASMC hypertrophy and PAH. We will estimate PA pressure, measure mRNA and protein of the hypertrophic markers alpha-actin and myosin heavy chain in PA, NFATc3 binding to 1-actin and myosin heavy chain promoters, and determine structural changes of the pulmonary vasculature on wild type +/- calcineurin/NFAT inhibitor and NFATc3 knockout mice exposed to normoxia and hypobaric CH. Specific Aim 2: To establish the contribution of NFATc3 to CH-induced downregulation of Kv channel expression and increases in pulmonary vasoconstrictor reactivity. We will use the same animal models proposed in aim1 and determine mRNA and protein Kv isoforms; NFATc3 binding to KV 1.5 and 2.1 promoters and association to additional transcriptional regulators; PASMC membrane potential and agonist-induced vasoconstriction in isolated pressurized PA. Specific Aim 3: To determine the mechanisms by which CH increases NFATc3 transcriptional activity in PASMC. We will determine the mediators (ET-1, Ca2+, calcineurin and Rho-kinase) of CH-increased NFAT activity using NFAT-luciferase reporter mice and NFAT-luciferase crossed with NFATc3 KO mice. Findings from the proposed studies will provide novel information about the signaling mechanisms regulating changes in gene transcription in PAH. A better understanding of this mechanisms in PAH will lead to the development of novel therapeutic approaches to prevent and cure this debilitating disease. Project Narrative. In the United States it is estimated that 300 new cases of primary pulmonary hypertension are diagnosed each year but the true prevalence and incidence is unknown (NHLBI, Facts About Primary Pulmonary Hypertension). Secondary pulmonary hypertension is much more common because it is caused by a variety of obstructive pulmonary diseases and living at high-altitude, two conditions associated with chronic hypoxia. Sustained high pulmonary arterial resistance to blood flow causes an increase in the right ventricular (RV) filling pressure, which will eventually cause RV hypertrophy, ischemia, failure, and sudden cardiac death. The most common causes of death among patients with pulmonary arterial hypertension are related to progressive right-sided heart failure and sudden cardiac death events. Thus there is a need to understand the impact of chronic hypoxia on physiological responses at the cellular, molecular, and genomic levels in order to develop appropriate treatment strategies for the large group of patients. Our long-term research goal is to define the mechanisms whereby chronic hypoxia leads to the pathologies of pulmonary hypertension to allow more rational design of pharmacological approaches to improve the quality of life of patients suffering lung diseases associated with chronic hypoxia, and to decrease the rate of morbidity and mortality linked to this condition. The complex process of developing pulmonary hypertension is driven, in part, by changes in gene expression. The role of the calcium-regulated transcription factor NFATc3 in pulmonary arterial hypertension has not been previously addressed, highlighting the impact of and need for the proposed research. The proposed studies will define for the first time the regulators and targets of NFATc3 signaling in pulmonary arterial smooth muscle cells in the devastating pathological condition of pulmonary arterial hypertension. NFATc3 has been previously linked to vascular development, regulation of vascular smooth muscle cell differentiation, proliferation and contractility. Therefore, a better understanding of the molecular mechanisms that underlie the vascular remodeling and increased vasoconstriction in pulmonary hypertension is expected to lead to the development of novel therapeutic approaches to prevent and treat this disease. The most novel aspects of this proposal are its ability to examine NFATc3 regulation of pulmonary vascular function in a truly integrated fashion. The planned experiments will utilize our expertise in molecular biology, vascular biology and integrated systems physiology.

描述（由申请人提供）：尽管有肺动脉高压（PAH）的起源，但肺血管阻力由于肺血管收缩、动脉重塑和红细胞增多症而升高，导致右心衰竭和死亡。暴露于慢性低压缺氧 (CH) 的啮齿动物会患上 PAH。形成 PAH 的复杂过程部分是由基因表达的变化驱动的。在 PAH 中，平滑肌细胞内 Ca2+ 增加，内皮素 1 (ET-1) 表达上调。 Ca2+ 调节肺动脉平滑肌 (PASMC) 收缩，并通过活化 T 细胞核因子 (NFAT) 与基因转录相关。 NFATc3 亚型特别涉及脉管系统的发育和平滑肌分化表型的维持。该提案的总体目标是确定 NFATc3 在与 CH-PAH 相关的血管变化的分子机制中的作用。假设 CH 激活 PASMC 中的 NFATc3，介导肥厚并增强肺动脉 (PA) 的收缩性，从而导致 PAH。具体目标 1：确定 NFATc3 在 CH 诱导的 PASMC 肥大和 PAH 中的作用。我们将估计 PA 压力，测量 PA 中肥大标记物 α-肌动蛋白和肌球蛋白重链的 mRNA 和蛋白质，NFATc3 与 1-肌动蛋白和肌球蛋白重链启动子的结合，并确定野生型 +/- 上肺血管系统的结构变化钙调神经磷酸酶/NFAT 抑制剂和 NFATc3 敲除小鼠暴露于常氧和低压 CH。具体目标 2：确定 NFATc3 对 CH 诱导的 Kv 通道表达下调和肺血管收缩反应性增加的贡献。我们将使用与aim1中提出的相同的动物模型并确定mRNA和蛋白质Kv亚型； NFATc3 与 KV 1.5 和 2.1 启动子结合并与其他转录调节因子关联；隔离加压 PA 中 PASMC 膜电位和激动剂诱导的血管收缩。具体目标 3：确定 CH 增加 PASMC 中 NFATc3 转录活性的机制。我们将使用 NFAT-荧光素酶报告小鼠和与 NFATc3 KO 小鼠杂交的 NFAT-荧光素酶来确定 CH 增加的 NFAT 活性的介质（ET-1、Ca2+、钙调神经磷酸酶和 Rho-激酶）。拟议研究的结果将提供有关调节 PAH 基因转录变化的信号机制的新信息。更好地了解 PAH 的这种机制将有助于开发新的治疗方法来预防和治愈这种使人衰弱的疾病。项目叙述。在美国，估计每年诊断出 300 例新的原发性肺动脉高压病例，但真实的患病率和发病率尚不清楚（NHLBI，关于原发性肺动脉高压的事实）。继发性肺动脉高压更为常见，因为它是由多种阻塞性肺疾病和生活在高海拔地区引起的，这两种情况都与慢性缺氧有关。肺动脉对血流的持续高阻力导致右心室（RV）充盈压升高，最终导致右心室肥大、缺血、衰竭和心源性猝死。肺动脉高压患者最常见的死亡原因与进行性右心衰竭和心源性猝死事件有关。因此，需要了解慢性缺氧对细胞、分子和基因组水平上生理反应的影响，以便为大量患者制定适当的治疗策略。我们的长期研究目标是明确慢性缺氧导致肺动脉高压的病理机制，以便更合理地设计药理学方法，以改善患有慢性缺氧相关肺部疾病的患者的生活质量，并降低其发生率。与这种情况相关的发病率和死亡率。肺动脉高压的复杂过程部分是由基因表达的变化驱动的。钙调节转录因子 NFATc3 在肺动脉高压中的作用此前尚未得到解决，突出了拟议研究的影响和必要性。拟议的研究将首次确定肺动脉高压破坏性病理条件下肺动脉平滑肌细胞中 NFATc3 信号传导的调节因子和靶标。 NFATc3 先前已被认为与血管发育、血管平滑肌细胞分化、增殖和收缩性的调节有关。因此，更好地了解肺动脉高压中血管重塑和血管收缩增加的分子机制有望导致开发预防和治疗这种疾病的新治疗方法。该提案最新颖的方面是它能够以真正集成的方式检查 NFATc3 对肺血管功能的调节。计划中的实验将利用我们在分子生物学、血管生物学和综合系统生理学方面的专业知识。