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

• 批准号: 10419046
• 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
• 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/10419046
• 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; DNA Sequence Alteration; Data; Deposition; Development; Diffuse; Disease; Disease Progression; Dose; Drug Targeting; Early Intervention; Endothelial Cells; Endothelium; Epithelial; Epithelial Cells; 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 of activation protein; 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; base; 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

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.

项目摘要 特发性肺纤维化（IPF）是一种进行性瘢痕性肺病， 到呼吸衰竭仍然存在对有效疗法的大量未满足的需求。IPF被认为 由脆弱上皮的损伤驱动，对功能障碍敏感，或由遗传 突变或损伤的剂量或类型。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发挥 在扩大组织损伤中的作用，使平衡从修复转向纤维化。数据 出现肺内皮细胞异常的患者， 肺纤维化，尽管这些变化对纤维形成过程的原因/影响是 没有被很好地理解。IPF患者的肺部显示血管通透性增加 患者并预测死亡率。此外，最近的几项单细胞RNA测序研究 已经证明肺毛细血管的损失和支气管血管的增加 人口这些变化的全部机械影响尚未确定。 Rho相关激酶（ROCK）信号传导已被证明在几种细胞凋亡中起主要作用。 内皮细胞功能，包括细胞收缩性和随后的血管 磁导率我已经在初步数据中表明，在内皮细胞中ROCK 2特异性的缺失， 小鼠模型（EC ROCK 2 KO）中的细胞可预防肺纤维化的发生 由内注射博来霉素诱导。除了减少肺纤维化，这些 小鼠在博莱霉素治疗后的早期，纤维化之前， 发展起来的这一发现表明内皮细胞ROCK 2、血管功能和 肺纤维化，并暗示了早期干预的可能性，当 纤维化可以是可预防的或可逆的。 在目前的建议中，我建议探索EC ROCK 2 KO中的保护性表型 小鼠进一步通过在肺纤维化的重复博来霉素模型中使用这些小鼠， 已显示可产生进行性纤维化，与IPF更相似。我将描述 通过染色和渗透性，该重复博来霉素模型中的内皮变化 分析。我还将对有或没有ROCK 2的小鼠肺进行单细胞RNA测序， 肺纤维化的博来霉素重复模型。我将寻找机械联系， 内皮ROCK 2与血管生成、衰老、内皮细胞向间质细胞转化和 细胞凋亡，都是在纤维化的背景下。