Histologic and Transcriptional Profiling of Endothelial Cells During Progressive Pulmonary Fibrosis

进行性肺纤维化期间内皮细胞的组织学和转录谱

基本信息

批准号：
10616601
负责人：
Rachel S Knipe
金额：
$ 8.33万
依托单位：
MASSACHUSETTS GENERAL HOSPITAL
依托单位国家：
美国
项目类别：
财政年份：
2022
资助国家：
美国
起止时间：
2022-05-01 至 2024-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10616601
关键词：
Abnormal Endothelial Cell Actins Apoptosis Attenuated Biological Assay Bleomycin Blood Vessels Blood capillaries Capillary Endothelial Cell Cell Aging Cell Communication Cell physiology Cells Clinical Clinical Trials Cytoskeleton DNA Sequence Alteration Data Deposition Development Diffuse Disease Disease Progression Dose Drug Targeting Early Intervention Endothelial Cells Endothelium Epithelial Cells Epithelium Equilibrium Extracellular Matrix Fibroblasts Fibrosis Functional disorder Gene Expression Gene Expression Profile Gene Expression Profiling Genetic Genetic Transcription Goals Histologic Homeostasis Human Imaging Techniques Immunofluorescence Immunologic Injections Injury Knockout Mice Knowledge Lead Link Lung Lung diseases Lung fibrogenesis Mediating Mediator Mesenchymal Modeling Mus Myofibroblast Myosin ATPase Normal tissue morphology Pathogenesis Pathologic Patients Permeability Phenotype Play Population Process Pulmonary Fibrosis Research Respiratory Failure Rho-associated kinase Role Signal Transduction Stains Structure of parenchyma of lung Therapeutic Intervention Time Vascular Permeabilities angiogenesis cell injury density effective therapy endothelial dysfunction experimental study fibrogenesis fibrotic lung idiopathic pulmonary fibrosis improved insight lung development lung injury migration molecular imaging mortality mouse model new therapeutic target novel prevent programs pulmonary function pulmonary vascular permeability repaired senescence sildenafil single-cell RNA sequencing tissue injury tissue repair tool trafficking transcriptome wound healing

项目摘要

PROJECT SUMMARY Idiopathic Pulmonary Fibrosis (IPF) is a progressive scarring lung disease that very often leads to respiratory failure. There remains a large unmet need for effective therapies. IPF is thought to be driven by injury to a vulnerable epithelium, sensitive to dysfunction either by genetic mutations or by the dose or type of injury. Epithelial dysfunction and senescence lead to diffuse tissue injury and activation of myofibroblasts. Dysregulated wound repair mechanisms may play a role in amplification of the tissue injury, tipping the balance from repair to fibrosis. Data is emerging that there are abnormalities in the pulmonary endothelium of patients who develop pulmonary fibrosis, although the cause/effect of these changes on the process of fibrogenesis is not well understood. Increased vascular permeability has been shown in the lungs of IPF patients and predicts mortality. In addition, several recent single-cell RNA sequencing studies have demonstrated loss of pulmonary capillaries and an increase in a bronchial vessel population. The full mechanistic implications of these changes have not been defined. Rho-associated kinase (ROCK) signaling has been shown to play a major role in several endothelial cell functions, including cell contractility and subsequent alterations in vascular permeability. I have shown in preliminary data that loss of ROCK2 specifically in endothelial cells in a mouse model (EC ROCK2 KO) prevents the development of pulmonary fibrosis induced by the delivery of intratracheal bleomycin. In addition to less pulmonary fibrosis, these mice had less vascular permeability in the early time period after bleomycin, before fibrosis develops. This finding suggests a link between endothelial ROCK2, vascular function, and pulmonary fibrosis, and hints at the potential for an opportunity for early intervention, when fibrosis may be preventable or reversible. In this current proposal, I propose to explore the protective phenotype in EC ROCK2 KO mice further by using these mice in a repetitive bleomycin model of pulmonary fibrosis, which has been shown to produce progressive fibrosis and is more similar to IPF. I will characterize the endothelial changes in this repetitive bleomycin model through staining and permeability assays. I will also perform single-cell RNA sequencing of mouse lungs with or without ROCK2, in the repetitive bleomycin model of pulmonary fibrosis. I will look for mechanistic links between endothelial ROCK2 and angiogenesis, senescence, endothelial to mesenchymal transition and apoptosis, all in the context of fibrogenesis.

项目概要特发性肺纤维化 (IPF) 是一种进行性疤痕性肺部疾病，通常会导致直至呼吸衰竭。对有效疗法的需求仍有大量未得到满足。 IPF被认为由脆弱上皮损伤驱动，对遗传性功能障碍敏感突变或伤害的剂量或类型。上皮功能障碍和衰老导致弥漫性组织损伤和肌成纤维细胞活化。失调的伤口修复机制可能会发挥作用放大组织损伤的作用，使修复和纤维化之间的平衡发生倾斜。数据是出现这种情况的患者的肺内皮存在异常肺纤维化，尽管这些变化对纤维发生过程的原因/影响是不太理解。 IPF 肺部血管通透性增加患者并预测死亡率。此外，最近的几项单细胞 RNA 测序研究已证明肺毛细血管减少和支气管血管增加人口。这些变化的完整机制影响尚未确定。 Rho 相关激酶 (ROCK) 信号传导已被证明在多种疾病中发挥重要作用内皮细胞功能，包括细胞收缩性和随后的血管变化渗透性。我在初步数据中表明，ROCK2 的缺失特别是在内皮细胞中小鼠模型中的细胞（EC ROCK2 KO）可预防肺纤维化的发展由气管内输送博来霉素诱导。除了减少肺纤维化之外，这些在博来霉素治疗后的早期、纤维化之前，小鼠的血管通透性较低发展。这一发现表明内皮 ROCK2、血管功能和肺纤维化，并暗示有可能进行早期干预，当纤维化是可以预防或可逆转的。在当前的提案中，我建议探索 EC ROCK2 KO 的保护表型通过在重复的博来霉素肺纤维化模型中使用这些小鼠，进一步对小鼠进行了研究，已被证明会产生进行性纤维化，并且与 IPF 更相似。我将表征通过染色和渗透性观察重复博莱霉素模型中的内皮变化化验。我还将对有或没有 ROCK2 的小鼠肺部进行单细胞 RNA 测序，在肺纤维化的重复博来霉素模型中。我将寻找之间的机械联系内皮 ROCK2 与血管生成、衰老、内皮间质转化和细胞凋亡，所有这些都发生在纤维发生的背景下。