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: 8845610
• 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/8845610
• 关键词: 3-Dimensional; Address; Advisory Committees; Alveolar; Area; Award; Behavior; Biological Assay; Bleomycin; Cell Culture Techniques; Cell Separation; Cells; Cellular biology; Characteristics; 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; Genetic study; 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; effective therapy; experience; hyaluronan synthase 1; knowledge base; lung injury; mouse model; novel; novel strategies; precursor cell; repaired; research study; response; self-renewal; therapeutic target; tool

DESCRIPTION (provided by applicant): 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.

描述(由申请人提供)：这个项目的目标是提供支持和持续的指导，因为我发展成为一名独立的内科医生-科学家，研究肺纤维化和上皮-间充质相互作用在疾病发病机制中的作用。这门课程是专门为加强我在小鼠肺纤维化模型、遗传和表型分析以及人类肺细胞生物学方面的理论和实践知识而设计的。这将通过实验室实践经验、教学培训、数据和结果的展示、与相关领域专家的互动以及来自专门咨询委员会的指导来实现。正如这项提案所概述的那样，我将至少有75%的时间被保护用于研究。在这段时间里，我将能够在许多方面获得实用的专业知识 新技术，包括流式细胞术、荧光激活细胞分选(FACS)、小鼠呼吸力学评估以及人肺细胞分离和培养。我还将建立我在基因组学和基因表达分析相关领域的知识基础。考虑到我的经验丰富的导师和丰富而充满活力的研究环境，我将成功完成提案中概述的工作。研究项目：特发性肺纤维化(IPF)是一种以严重的进行性肺泡纤维化为特征的疾病，最常发生在老年人，除肺移植外没有有效的治疗方法。根据人类遗传学研究发现，肺泡上皮细胞(AEC2)特异性基因突变与家族性肺纤维化之间存在联系，肺泡上皮细胞异常与疾病的发展有关。然而，我们缺乏关于这些上皮异常如何触发潜在间充质的活跃反应从而导致显著和进行性瘢痕形成的知识。我们也不知道是哪些细胞产生了这种疾病特有的病理性肌成纤维细胞。我已经在肺间充质中鉴定出一种独特的成纤维细胞群，它可能是肌成纤维细胞的来源。这些细胞表达PDGF受体α(PDGFRA)，靠近AEC2，在博莱霉素肺损伤后增殖，上调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(Has2)。我还证明了这些细胞在三维小鼠培养系统中支持小鼠AEC2的克隆生长和分化，表明它们是肺泡上皮细胞生态位的关键组成部分。我假设PDGFRA+成纤维细胞是博莱霉素肺损伤后肌成纤维细胞的来源，这些细胞在被触发时产生促纤维化介质，如透明质酸(HA)和Has2，这些细胞是人体AEC2生态位在动态平衡期间的关键组成部分。我将在以下几个具体目标上验证这些假说：目的1：确定博莱霉素肺损伤后PDGFRA+成纤维细胞是否产生肌成纤维细胞。目的：验证PDGFRA+成纤维细胞是博莱霉素性肺纤维化形成过程中促纤维化介质HA和Has2的主要来源。目的：验证原代人PDGFRA+成纤维细胞支持人AEC2细胞存活、增殖和分化的假说，从而成为AEC2生态位的组成部分。