Interrogation of individual cells to identify progenitor cells and their niches

询问单个细胞以识别祖细胞及其生态位

• 批准号: 8114057
• 负责人: MARK A KRASNOW
• 金额: $ 101.61万
• 依托单位: STANFORD UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2016-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8114057
• 关键词: Animal Model; Cells; Cellular biology; Color; Complex; Development; Disease; Drosophila genus; Gene Expression; Genetic; Genomics; Human; In Situ Hybridization; In Vitro; Individual; Libraries; Lung; Lung diseases; Maintenance; Mesenchyme; Methods; Microfluidics; Microscopic; Microscopy; Molecular Profiling; Mouse Strains; Mus; Pathway interactions; Pattern; Population; Procedures; Resolution; Signal Pathway; Signal Transduction; Signaling Molecule; Stem cells; Structure of parenchyma of lung; Techniques; Tissues; Transgenic Mice; in vivo; lung development; molecular marker; prevent; progenitor; research study; response; three dimensional structure; tool

DESCRIPTION (provided by applicant): Identifying and characterizing lung progenitor cells and their signaling niches is crucial for understanding how a healthy lung is built and maintained, how alteration of development and maintenance pathways cause or contribute to lung disease, and how disease can be prevented and damaged lung tissue restored or replaced. Identifying and characterizing lung progenitor cells and their niches has been hampered by the complex three-dimensional structure of the lung and the lack of tools to mark, follow the fate, and manipulate gene expression in individual lung cells in vivo. Here we propose to develop such tools, by adapting for use in mouse lung the systematic genetic approaches and single cell resolution genetic tools ("clonal analysis") that have been used over the past decade to elucidate progenitor and stem cells and their signaling niches in the model organism Drosophila. To facilitate this, we will develop a new microscopy procedure (multidimensional microscopic molecular profiling) that allows the levels of dozens of molecular markers to be quantified at subcellular resolution on a single tissue section, the equivalent of multi-dimensional FACS analysis for intact tissues. We combine these in vivo approaches with a high throughput in vitro approach that takes advantage of recent advances in genomics and microfluidics to characterize individual lung progenitor cells and their developmental responses to the signals identified in the in vivo experiments. This combined approach is general and applicable to progenitor cells throughout the lung and other mouse tissues, although we focus on the poorly characterized progenitor cells in lung mesenchyme.

描述（由申请人提供）： 识别和表征肺祖细胞及其信号传导生态位对于了解健康的肺是如何构建和维持的、发育和维持途径的改变如何导致或促成肺疾病以及如何预防疾病和恢复或替换受损的肺组织至关重要。由于肺复杂的三维结构以及缺乏在体内标记、追踪命运和操纵单个肺细胞基因表达的工具，识别和表征肺祖细胞及其生态位一直受到阻碍。在这里，我们建议开发这样的工具，通过适应在小鼠肺中使用系统遗传方法和单细胞分辨率遗传工具（“克隆分析”），这些工具在过去十年中被用来阐明模式生物果蝇中的祖细胞和干细胞及其信号传导生态位。为了实现这一目标，我们将开发一种新的显微镜程序（多维显微分子分析），可以在单个组织切片上以亚细胞分辨率对数十种分子标记物的水平进行定量，相当于对完整组织进行多维 FACS 分析。我们将这些体内方法与高通量体外方法相结合，利用基因组学和微流体学的最新进展来表征个体肺祖细胞及其对体内实验中确定的信号的发育反应。这种组合方法是通用的，适用于整个肺和其他小鼠组织的祖细胞，尽管我们关注的是肺间充质中特征不明确的祖细胞。