The role of Pdgfra+ fibroblasts in lung fibrosis and alveolar homeostasis

Pdgfra成纤维细胞在肺纤维化和肺泡稳态中的作用

• 批准号: 8679394
• 负责人: Christina Eleanor Barkauskas
• 金额: $ 11.14万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-05 至 2019-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8679394
• 关键词: 3-Dimensional; Address; Advisory Committees; Alveolar; Area; Award; Behavior; Biological Assay; Bleomycin; Cell Culture Techniques; Cell Separation; Cells; Cellular biology; Cicatrix; Collagen; Data; Development; Differentiation and Growth; Disease; Drug Targeting; Educational Curriculum; Environment; Epithelial; Epithelial Cells; Extracellular Matrix; Fibroblasts; Fibrosis; Flow Cytometry; Fluorescence-Activated Cell Sorting; Future; Gases; Gene Expression Profiling; Genes; Genetic; Genomics; Goals; Growth; Hamman-Rich syndrome; Hand; Health; Homeostasis; Human; Human Genetics; Hyaluronan; Hyaluronic Acid; Immunohistochemistry; In Vitro; Individual; Injury; Knock-in Mouse; Knowledge; Laboratories; Link; LoxP-flanked allele; Lung; Lung Compliance; Lung diseases; Mediating; Mediator of activation protein; Mentors; Mentorship; Mesenchymal; Mesenchyme; Modeling; Mus; Mutation; Myofibroblast; Organoids; Pathogenesis; Pathologic; Physicians; Platelet-Derived Growth Factor alpha Receptor; Population; Positioning Attribute; Production; Proliferating; Pulmonary Fibrosis; Research; Research Project Grants; Respiratory Failure; Respiratory Mechanics; Role; Scientist; Signal Pathway; Signal Transduction; Smooth Muscle Actin Staining Method; Source; Stem cells; Stromal Cells; Structure; Supporting Cell; System; Techniques; Testing; Time; Training; Transplantation; Work; alveolar epithelium; alveolar homeostasis; base; cell type; combat; design; disease characteristic; effective therapy; experience; hyaluronan synthase 1; knowledge base; lung injury; mouse model; novel; novel strategies; precursor cell; public health relevance; repaired; research study; response; self-renewal; therapeutic target; tool

PROJECT SUMMARY/ABSTRACT Career development Goal / Plan: The goal of this project is to provide support and continued mentorship as I develop into an independent physician-scientist studying pulmonary fibrosis and the role of epithelial- mesenchymal interaction in the pathogenesis of the disease. The curriculum is specifically designed to strengthen my theoretical and practical knowledge base in murine models of lung fibrosis, genetic and phenotypic analysis, and human lung cell biology. This will be achieved through hands on laboratory experience, didactic training, presentation of data and results, interactions with experts in related fields, and mentorship from a dedicated advisory committee. As outlined in this proposal, I will have at least 75% of time protected for research. During this time, I will be able to gain practical expertise in a number of new techniques, including flow cytometry, fluorescence activated cell sorting (FACS), assessment of murine respiratory mechanics, and human lung cell isolation and culture. I will also build my knowledge base in relevant areas of genomics and gene expression analysis. I am well positioned to successfully complete the work outlined in the proposal given my experienced mentors and the rich and dynamic research environment in which I will perform the work. Research Project: Idiopathic pulmonary fibrosis (IPF) is a disease characterized by severe and progressive alveolar fibrosis that occurs most often in older individuals and that has no effective treatment short of lung transplant. Based upon human genetic studies revealing a link between mutations in genes specific to type 2 alveolar epithelial cells (AEC2s) and familial forms of pulmonary fibrosis, it is accepted that abnormalities in the alveolar epithelium are associated with disease development. We lack knowledge, however, of how these epithelial abnormalities trigger the exuberant response from the underlying mesenchyme that leads to significant and progressive scar formation. We also are unaware of which cells give rise to the pathologic myofibroblasts that are characteristic of the disease. I have identified a unique population of fibroblasts in the lung mesenchyme that are likely to be a source of myofibroblasts. These cells, which express PDGF receptor alpha (Pdgfra), lie in close proximity to AEC2s and, after bleomycin lung injury, proliferate, up-regulate myofibroblast markers, and express pro-fibrotic mediators such as hyaluronic acid synthase 2 (Has2). I have also shown that these cells support the clonal growth and differentiation of murine AEC2s in a 3-dimensional murine culture system, suggesting that they are critical components of the alveolar epithelial cell niche. I hypothesize that Pdgfra+ fibroblasts are a source of myofibroblasts following bleomycin lung injury, that these cells produce pro-fibrotic mediators such as hyaluronan (HA) and Has2 when triggered, and that these cells are critical components of the human AEC2 niche during homeostasis. I will test these hypotheses in the following specific aims: AIM 1: To determine if Pdgfra+ fibroblasts give rise to myofibroblasts after bleomycin lung injury. AIM 2: To test the hypothesis that Pdgfra+ fibroblasts are a major source of the pro-fibrotic mediators HA and Has2 in the development of bleomycin-induced pulmonary fibrosis. AIM 3: To test the hypothesis that primary human PDGFRa+ fibroblasts support the survival, proliferation, and differentiation of human AEC2s, thereby serving as components of the AEC2 niche.

项目总结/摘要 职业发展目标/计划：本项目的目标是提供支持和持续的指导，因为我 发展成为一个独立的医生，科学家研究肺纤维化和上皮细胞的作用， 间充质相互作用在疾病的发病机制。该课程是专门设计， 加强了我在肺纤维化小鼠模型、遗传和 表型分析和人类肺细胞生物学。这将通过实验室实践来实现 经验，教学培训，数据和结果的介绍，与相关领域专家的互动，以及 一个专门的咨询委员会的指导。正如这份提案中所概述的，我将有至少75%的时间 保护研究。在此期间，我将能够获得一些新的实用专业知识， 技术，包括流式细胞术、荧光激活细胞分选术（FACS）、评估小鼠免疫缺陷病毒（HCV）的能力。 呼吸力学和人肺细胞分离和培养。我还将建立我的知识基础， 基因组学和基因表达分析的相关领域。我有能力成功完成 我在建议书中概述的工作，考虑到我经验丰富的导师和丰富而充满活力的研究环境， 我来完成这项工作 研究项目：特发性肺纤维化（IPF）是一种以严重和进行性为特征的疾病， 肺泡纤维化最常发生在老年人中，除了肺部外没有有效的治疗方法。 移植基于人类遗传学研究揭示了2型糖尿病特异性基因突变之间的联系， 肺泡上皮细胞（AEC 2）和家族性肺纤维化，公认的是， 肺泡上皮细胞与疾病发展有关。然而，我们缺乏知识， 上皮细胞异常触发了下层间充质的旺盛反应， 显著和进行性瘢痕形成。我们也不知道是哪些细胞引起了病理性 肌成纤维细胞是疾病的特征。我发现了一个独特的成纤维细胞群， 肺间充质可能是肌成纤维细胞的来源。这些表达PDGF受体的细胞 Pdgfra α与AEC 2非常接近，在博来霉素肺损伤后， 肌成纤维细胞标志物，并表达促纤维化介质，如透明质酸合成酶2（Has 2）。我有 还表明这些细胞支持小鼠AEC 2的克隆生长和分化， 小鼠培养系统，表明它们是肺泡上皮细胞龛的关键组成部分。我 假设Pdgfra+成纤维细胞是博来霉素肺损伤后肌成纤维细胞来源， 细胞产生促纤维化介质，如透明质酸（HA）和Has 2，当触发时，这些细胞 是体内平衡过程中人类AEC 2生态位的关键组成部分。 目的1：确定Pdgfra+成纤维细胞是否能产生细胞因子。 博莱霉素肺损伤后肌成纤维细胞增多。目的2：验证Pdgfra+成纤维细胞是一种细胞因子的假说。 促纤维化介质HA和Has 2在博来霉素诱导的肺纤维化发展中的主要来源 纤维化目的3：为了检验原代人PDGFRa+成纤维细胞支持存活的假设， 增殖和分化，从而充当AEC 2小生境的组分。