Role of endothelial Twist1 in vascular remodeling in pulmonary hypertension

内皮细胞 Twist1 在肺动脉高压血管重塑中的作用

基本信息

批准号：
10089472
负责人：
TADANORI MAMMOTO
金额：
$ 30.41万
依托单位：
MEDICAL COLLEGE OF WISCONSIN
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-02-01 至 2023-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10089472
关键词：
Apoptosis Attenuated BMP2 gene Behavior Blood Vessels Cells Conditioned Culture Media Data Development Distal Endothelial Cells Endothelium Exposure to Fibrin Functional disorder Gel Gene Mutation Goals Heritability Human Hypoxia Implant In Vitro Induced Mutation Knock-out Knockout Mice Label Lead Life Lung Mediating Modeling Mus Muscle Mutation Myofibroblast Organ Paracrine Communication Pathogenesis Pathologic Pathologic Neovascularization Patients Pericytes Phosphorylation Platelet-Derived Growth Factor B Play Proto-Oncogene Proteins c-akt Pulmonary Heart Disease Pulmonary Hypertension Pulmonary artery structure Role Severities Signal Transduction Smooth Muscle Actin Staining Method Smooth Muscle Myocytes Structure Structure of parenchyma of lung System Thymidine Phosphorylase Vascular Diseases Vascular remodeling Wild Type Mouse arteriole behavior in vitro bone morphogenetic protein receptors cadherin 5 cell behavior implantation knock-down migration normoxia novel novel strategies overexpression paracrine pulmonary arterial hypertension recruit symptomatic improvement targeted treatment transcription factor

项目摘要

Project Summary Pulmonary hypertension (PH) is a multifactorial life threatening pulmonary vascular disease characterized by aberrant muscularization of the normally non-muscularized distal pulmonary arterioles (PAs). Dysfunction of PA endothelial cells (ECs) plays key roles in accumulation of pulmonary artery smooth muscle cells (PASMCs) to distal PAs. Expression of the transcription factor Twist1 is upregulated in the lungs of PH patients. However, the role of endothelial Twist1 in pathogenesis of PH has not been fully understood. The goal of this proposal is to explore the mechanism by which endothelial Twist1 controls accumulation of PASMCs to distal PAs. Our new preliminary data demonstrate that Twist1 overexpression increases the levels of platelet-derived growth factor B (PDGFB) in human pulmonary arterial endothelial (HPAE) cells and induces proliferation and migration of PASMCs. Hypoxia increases Twist1 Ser42 phosphorylation and PDGFB expression in HPAE cells and conditioned medium from these cells stimulates migration of PASMCs, while Twist1 knockdown in ECs inhibits the effects. When we implanted fibrin gel supplemented with fluorescently labeled ECs on the mouse lung, Twist1 overexpression in these ECs increases PDGFB expression and promotes the recruitment of α-smooth muscle actin (SMA)-positive cells in the gel. Hypoxia induces accumulation of αSMA-positive cells to blood vessels formed in the gel, while Twist1 knockdown in ECs attenuates the effects. Hypoxia also stimulates accumulation of αSMA-positive cells to small PAs in the mouse lungs, which is suppressed in Tie2-specific Twist1 knockout mice. The levels of Twist1 and PDGFB are higher in lung ECs isolated from mice with type-2 bone morphogenetic protein receptor (Bmpr2) mutation that influences the severity of human PH. The Bmpr2 mutation also stimulates accumulation of αSMA-positive cells to ECs in the gel implanted on the hypoxia- treated mouse lungs. We hypothesize that endothelial Twist1 mediates hypoxia-induced SMC accumulation to distal PAs through PDGFB paracrine signaling. In Aim 1, we will investigate whether endothelial Twist1 or its phosphorylation controls PASMC behaviors through PDGFB paracrine signaling in vitro. In Aim 2, we will determine whether endothelial Twist1 mediates hypoxia-induced SMC accumulation to blood vessels in the mouse lung using the mouse lung fibrin gel implantation system. We will also explore whether endothelial Twist1 mediates hypoxia-induced vascular remodeling using inducible VE-cadherin-specific Twist1 knockout mice. In Aim 3, we will investigate whether endothelial Twist1 mediates Bmpr2 mutations-induced vascular remodeling in ECs under normoxia or hypoxia in vitro and in the gel implantation model. Our focus on exploring the role of endothelial Twist1 in hypoxia-induced SMC accumulation to distal PAs using the mouse lung gel implantation system is unique and relevant advances, which will further our understanding of the pathogenesis of PH.

项目概要肺动脉高压（PH）是一种威胁生命的多因素肺血管疾病，其特征是正常情况下非肌肉化的远端肺小动脉（PA）的异常肌肉化。 PA 内皮细胞 (EC) 在肺动脉平滑肌细胞 (PASMC) 积累中发挥关键作用然而，PH 患者肺部转录因子 Twist1 的表达上调。内皮 Twist1 在 PH 发病机制中的作用尚未完全了解。该提案的目标是。探索内皮 Twist1 控制 PASMC 向远端 PA 积累的机制。新的初步数据表明 Twist1 过度表达可增加血小板衍生生长的水平人肺动脉内皮 (HPAE) 细胞中的 B 因子 (PDGFB) 并诱导增殖和迁移缺氧会增加 HPAE 细胞中的 Twist1 Ser42 磷酸化和 PDGFB 表达来自这些细胞的条件培养基刺激 PASMC 的迁移，而 EC 中的 Twist1 敲除则抑制当我们将补充有荧光标记 EC 的纤维蛋白凝胶植入小鼠肺部时，这些 EC 中 Twist1 的过表达会增加 PDGFB 的表达并促进 α-smooth 的募集凝胶中的肌肉肌动蛋白 (SMA) 阳性细胞会因缺氧而导致 αSMA 阳性细胞积聚到血液中。凝胶中形成血管，而 EC 中的 Twist1 敲除也会减弱缺氧的影响。 αSMA 阳性细胞在小鼠肺部的小 PA 中积累，这种情况在 Tie2 特异性中受到抑制 Twist1 敲除小鼠从 2 型小鼠中分离出的肺 EC 中 Twist1 和 PDGFB 的水平较高。骨形态发生蛋白受体 (Bmpr2) 突变影响人类 PH 的严重程度。突变还刺激αSMA阳性细胞在植入缺氧细胞的凝胶中积累到EC中。我们发现经过处理的内皮 Twist1 介导缺氧诱导的 SMC 积累通过 PDGFB 旁分泌信号传导的远端 PA 在目标 1 中，我们将研究内皮 Twist1 或其是否参与。在体外，磷酸化通过 PDGFB 旁分泌信号控制 PASMC 行为。确定内皮 Twist1 是否介导缺氧诱导的 SMC 在血管中的积累我们还将探索使用小鼠肺纤维蛋白凝胶植入系统的小鼠肺。 Twist1 通过诱导 VE-钙粘蛋白特异性 Twist1 敲除介导缺氧诱导的血管重塑在目标 3 中，我们将研究内皮 Twist1 是否介导 Bmpr2 突变诱导的血管。我们重点探索常氧或缺氧体外 EC 的重塑以及凝胶植入模型。使用小鼠肺凝胶观察内皮 Twist1 在缺氧诱导的 SMC 向远端 PA 聚集中的作用植入系统的独特性和相关进展，这将进一步我们对发病机制的理解 PH值。