Endothelium-driven signaling network in the development of pulmonary hypertension

肺动脉高压发生过程中内皮驱动的信号网络

• 批准号: 10646507
• 负责人: Hua Linda Cai
• 金额: $ 52.15万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10646507
• 关键词: Animal Disease Models; Animals; Arteries; Attenuated; Biological Availability; Blood Vessels; Candidate Disease Gene; Cellular biology; Collagen; Coupling; Data; Deposition; Development; Dihydrofolate Reductase; Disease; Disease Progression; Ear; Electron Spin Resonance Spectroscopy; Endothelial Cells; Endothelium; Enzymes; Event; GTP Cyclohydrolase I; Gene Expression Profile; Gene Expression Regulation; Gene Transfer Techniques; Genes; Human; Hypertension; Hypoxia; Interruption; Knock-out; Knockout Mice; Lesion; LoxP-flanked allele; Lung; Medial; Mediating; Modeling; Molecular; Mus; NADPH Oxidase; NG-Nitroarginine Methyl Ester; NOS3 gene; Nitric Oxide; Oxidation-Reduction; Pathogenesis; Pathologic; Pathologic Processes; Patients; Phenotype; Physiological; Prevention; Production; Protein Isoforms; Pulmonary Heart Disease; Pulmonary Hypertension; Pulmonology; Rodent; Role; Signal Transduction; Spectrophotometry; Superoxides; Systolic Pressure; Therapeutic; Thick; Time; Transgenic Mice; Vascular remodeling; Vascular resistance; Ventricular; attenuation; cardiopulmonary system; cofactor; conditional knockout; design; efficacy evaluation; endoplasmic reticulum stress; endothelial dysfunction; genetic strain; human disease; hypoxia-induced pulmonary hypertension; inhibitor; innovation; mitochondrial dysfunction; molecular phenotype; mortality; mouse model; novel; novel therapeutics; overexpression; preservation; prevent; pulmonary arterial hypertension; pulmonary arterial pressure; right ventricular failure; screening; small molecule; small molecule inhibitor; tetrahydrobiopterin; transcriptome sequencing; translational study; vasoconstriction

ABSTRACT ______ The central focus of application is to establish a novel pulmonary hypertension (PH) model in mice by specifically targeting the endothelium, and reveal a novel endothelium-driven signaling network of uncoupled eNOS-ER stress- mitochondrial dysfunction axis in the pathogenesis of PH, targeting of which would result in novel therapeutics for PH. Pulmonary hypertension is a severe human disease characterized by intensive remodeling of small arteries in the lung, resulting in vasoconstriction, elevated vascular resistance and pulmonary arterial pressure, and eventually right heart failure. In preliminary studies, we have generated a novel PH model by directly targeting eNOS to provoke eNOS uncoupling (DAHP to inhibit GTPCHI) and eNOS uncoupling-dependent endothelial dysfunction. Of note, endothelial dysfunction is one of the earliest events and may the initiating step of idiopathic pulmonary artery hypertension (IPAH). Importantly, DAHP-treated mice developed robust PH phenotypes of increased mean pulmonary artery pressure (mPAP) and right ventricular systolic pressure (RVSP), accompanied by extensive vascular remodel characterized by typical human like vascular lesions of medial thickness, neointimal formation, and plexiform features. RNA-sequencing (RNA- seq) data indicated that, comparing to human patients with PH, the DAHP model had more overlappingly and substantially regulated genes vs. the hypoxia model (217 vs. 92). In preliminary studies we have also revealed new molecular mechanisms mediating PH development downstream of uncoupled eNOS, involving ER stress and mitochondrial dysfunction. Additionally, since reversal of eNOS cofactor tetrahydrobiopterin salvage enzyme dihydrofolate reductase (DHFR) deficiency downstream of NADPH oxidase (NOX) activation is robustly effective in preserving eNOS coupling activity, novel genetic strains specifically targeting NOX isoforms and DHFR will be examined for efficacies modulating PH phenotypes (16 novel and unique strains, most of which made in house). In Aim 1 we aim to establish a novel human like murine model of PH by fully characterizing phenotypes of DAHP-treated mice, and by comparing its gene regulation profile to that of human patients with PH and of Sugen5416/Hypoxia (SuHx)-treated mice. Also to further examine roles in PH development of novel candidate genes identified by RNA-seq analyses. In Aim 2, we will examine novel endothelium- driven signaling network of uncoupled eNOS-ER stress-mitochondrial dysfunction axis in the pathogenesis of PH using DAHP, hypoxia and Su/Hx models of PH. In Aim 3, we will examine whether strategies targeting endothelial DHFR, such as endothelium-specific transgenesis of DHFR, or knockout of NOX isoforms specifically in the endothelium to preserve DHFR function, would be of novel therapeutic potential for PH. We will also examine whether global and conditional knockout of DHFR, or endothelium-specific overexpression of NOX isoforms, leads to PH development and exaggerated PH. Effects on PH development of novel DHFR activators, and small molecule inhibitors for NOX, will also be examined. These studies are highly significant and translational in potentially identifying novel therapeutic options for the treatment and/or prevention of PH, namely via attenuation of NOX isoform activation to preserve endothelial DHFR function and eNOS coupling activity to shut down ER stress and mitochondrial dysfunction, or via direct activation of DHFR.

抽象的 ______ 应用的主要重点是通过专门在小鼠中建立新型的肺动脉高压（pH）模型 靶向内皮，并揭示一个新型的内皮驱动的信号网络 线粒体功能障碍轴在pH的发病机理中，其靶向将导致pH的新型疗法。 肺动脉高压是一种严重的人类疾病，其特征是对肺部小动脉进行密集的重塑， 导致血管收缩，血管耐药性和肺动脉压的升高，最终右心 失败。在初步研究中，我们通过直接靶向eNOS引起eNOS产生了一种新型的pH模型 解偶联（DAHP以抑制GTPCHI）和ENOS解开依赖性内皮功能障碍。值得注意的是，内皮 功能障碍是最早的事件之一，可能是特发性肺动脉高压（IPAH）的起始步骤。 重要的是，DAHP处理的小鼠发展了平均肺动脉压（MPAP）增加的强大pH表型 和右心收缩压（RVSP），伴随着广泛的血管重塑，其特征是 像内侧厚度，新内膜形成和丛状特征的血管病变一样。 RNA - 序列（RNA- seq）数据表明，与人类pH患者相比，DAHP模型的模型更加严重，实质上 调节基因与缺氧模型（217 vs. 92）。在初步研究中，我们还揭示了新的分子 未耦合eNOS下游介导pH发育的机制，涉及ER应力和线粒体 功能障碍。此外，由于eNOS辅助因子四氢无生翅目酶的逆转酶二氢叶酸还原酶还原酶 （DHFR）NADPH氧化酶（NOX）激活下游的缺陷在保持eNOS耦合方面非常有效 活性，专门针对NOX同工型和DHFR的新型遗传菌株将检查是否调节pH值 表型（16种新颖而独特的菌株，其中大部分是在室内制造的）。在目标1中，我们旨在建立一个像 通过充分表征DAHP处理的小鼠的表型的鼠模型，并通过比较其基因调节 pH和SUGEN5416/缺氧（SUHX）治疗的小鼠的人类患者的特征。也进一步研究 通过RNA-Seq分析鉴定的新型候选基因的pH发展。在AIM 2中，我们将检查新型的内皮 - 使用pH发病机理中未耦合的eNOS-ER应力 - 处方功能障碍轴的驱动信号网络使用 pH的DAHP，缺氧和SU/HX模型。在AIM 3中，我们将研究针对内皮DHFR的策略是否存在 作为DHFR的内皮特异性转基因或在内皮中特别保留的NOX同工型的敲除 DHFR功能将具有pH的新型治疗潜力。我们还将研究全球和有条件 DHFR的敲除或NOX同工型的内皮特异性过表达，导致pH发育并夸大 ph。还将检查对新型DHFR激活剂的pH发育以及NOX的小分子抑制剂的影响。 这些研究在潜在地识别治疗的新型治疗选择方面非常重要，并且转化 和/或预防pH，即通过衰减NOX同工型激活来保留内皮DHFR功能和 eNOS耦合活性以关闭ER应力和线粒体功能障碍，或通过DHFR的直接激活。

项目成果

NADPH oxidases and oxidase crosstalk in cardiovascular diseases: novel therapeutic targets.

• DOI: 10.1038/s41569-019-0260-8
• 发表时间: 2020-03
• 期刊: Nature reviews. Cardiology
• 作者: Zhang Y;Murugesan P;Huang K;Cai H
• 通讯作者: Cai H