Therapeutic ECM Resorption in Cellular Systems and Precision Cut Lung Slices.

细胞系统中的治疗性 ECM 吸收和精密切割肺切片。

• 批准号: 10318078
• 负责人: Daniel J. Tschumperlin
• 金额: $ 61.97万
• 依托单位: MAYO CLINIC ROCHESTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-12-15 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10318078
• 关键词: Address; Age; Alveolar; Animal Model; Appearance; Architecture; Biological; Biological Assay; Biological Models; Biological Response Modifier Therapy; Cell Communication; Cells; Cellular Assay; Characteristics; Chemicals; Cicatrix; Collagen; Cyclic AMP; Data; Deposition; Disease; Environment; Epigenetic Process; Extracellular Matrix; Failure; Fibrillar Collagen; Fibroblasts; Fibrosis; Gases; Goals; Human; Impairment; In Vitro; Individual; Lung; Mediating; Mediator of activation protein; Metabolic; Modeling; Molecular; Natural regeneration; Organoids; Participant; Pathogenesis; Pathway interactions; Patients; Pharmacology; Physiological; Process; Pulmonary Fibrosis; Research; Signal Transduction; Slice; Stimulus; Structure of parenchyma of lung; System; Testing; Therapeutic; Therapeutic Intervention; Tissues; Variant; cell type; crosslink; fibrotic lung; high throughput screening; idiopathic pulmonary fibrosis; insight; lung repair; macrophage; method development; miniaturize; novel; repaired; therapeutic evaluation; tissue repair; tool; ventilation

Project Summary Idiopathic Pulmonary Fibrosis (IPF) is characterized by progressive replacement of functional alveolar gas exchange tissue with collagen rich scar. The accumulation, crosslinking and stiffening of this matrix are defining features of the disease, and definitive barriers to effective repair or regeneration. Emerging evidence indicates that fibrotic scar remains highly resorbable under appropriate conditions], a process that appears to be impaired or absent in humans with IPF. The conditions and pathways that promote fibroblasts (and other cell types) to resorb collagen rich scar are largely unknown. Development of methods and approaches to address this critical gap in understanding is the focus of this U01 proposal. We propose to develop model systems in which ECM deposition and resorption can be efficiently studied in both primary cultured lung fibroblasts as well as precision cut lung slices (PCLS). Our preliminary data show that under appropriate stimuli, IPF-derived human lung fibroblasts can be prompted to degrade and resorb fibrillar collagen. We hypothesize that under appropriate stimuli fibroblasts can be stimulated to not only resorb collagen rich ECM in vitro, but also resorb scar-associated ECM in the lungs from patients with IPF. We propose to develop and leverage novel in vitro tools to test this hypothesis, with the goal of identifying biological pathways and therapeutic interventions that mediate physiologic collagen resorption. We propose to pursue these goals through two aims. In the first aim we will develop culture systems allowing us to identify the signals that promote ECM resorption by lung fibroblasts and delineate the molecular mechanisms of collagen resorption. We will validate these assays for high-throughput discovery and perform a focused screen as proof of concept of the value of this approach. We will also compare the innate ECM deposition and degradation characteristics of IPF and control fibroblasts and fibroblast subsets. In the second aim we will develop ex vivo lung tissue assays to test therapeutic modulation and mechanisms of clearance of scar-associated IPF ECM. We will characterize baseline and stimulus evoked collagenolytic activity in control and IPF lung tissue, and define the association of this activity with lung cell types and histopathological appearance of the tissue. We will also test candidate hits identified in aim 1 for their capacity to increase targeted fibrillar collagen degradation in the native IPF ECM environment. Together the proposed studies will establish robust models of ECM deposition and resorption in primary human IPF fibroblasts and ex vivo lung slices. This platform will open new avenues for identifying signals and mechanisms that shift fibroblasts in IPF toward a matrix resorbing state, generating new opportunities to develop advanced therapeutics for IPF.

项目摘要 特发性肺纤维化（IPF）的特征是逐步替代功能性肺泡气体 用富含胶原蛋白的疤痕交换组织。该矩阵的积累，交联和僵硬是 定义疾病的特征，以及有效修复或再生的确定性障碍。新兴证据 表明在适当条件下纤维化疤痕仍然高度吸收] IPF的人类在人类中受到损害或不存在。促进成纤维细胞的条件和途径（以及其他 细胞类型）以吸收富含胶原蛋白的疤痕在很大程度上是未知的。开发方法和方法 在理解方面解决这个关键差距是该U01提案的重点。我们建议开发模型 可以在两个原发性培养的肺中有效研究ECM沉积和吸收的系统 成纤维细胞以及精确切割的肺切片（PCL）。我们的初步数据表明，在适当的情况下 可以提示刺激，IPF衍生的人肺成纤维细胞降解和吸收纤维胶原蛋白。我们 假设在适当的刺激下可以刺激成纤维细胞，不仅可以吸收富含胶原蛋白的ECM 体外，但还来自IPF患者的肺中与疤痕相关的ECM。我们建议开发和 利用新颖的体外工具来检验这一假设，目的是确定生物学途径和 介导生理胶原吸收的治疗干预措施。我们建议追求这些目标 通过两个目标。在第一个目的中，我们将开发文化系统，使我们能够确定信号 通过肺成纤维细胞促进ECM吸收，并描述胶原蛋白吸收的分子机制。 我们将验证这些测定法以进行高通量发现，并执行集中屏幕作为概念证明 这种方法的价值。我们还将比较先天的ECM沉积和降解特征 IPF和控制成纤维细胞和成纤维细胞子集的。在第二个目的中，我们将开发离体肺组织 测试与疤痕相关的IPF ECM清除的治疗调节和机制的测定。我们将 表征基线和刺激在对照和IPF肺组织中引起的胶原式活性，并定义 这种活性与组织的肺细胞类型和组织病理学外观的关联。我们还将测试 AIM 1中确定的候选命中率是因为它们增加了目标纤维胶原蛋白降解的能力 本地IPF ECM环境。拟议的研究将共同​​建立ECM沉积的强大模型 并在原代人IPF成纤维细胞和离体肺切片中吸收。该平台将开放新途径 用于识别将IPF中的成纤维细胞转移到矩阵解析状态的信号和机制，生成 为IPF开发高级治疗剂的新机会。