Progenitor Cells in Lung Development and Repair

肺发育和修复中的祖细胞

• 批准号: 7456820
• 负责人: BRIGID L.M. HOGAN
• 金额: $ 39万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1992
• 资助国家: 美国
• 起止时间: 1992-05-05 至 2013-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7456820
• 关键词: Address; Adult; Alleles; Alveolar Cell; Basal Cell; Biological Assay; Biological Models; Birth; Cell Separation; Cell physiology; Cells; Chromosome Mapping; Clara cell; Condition; Daughter; Defect; Development; Distal; Embryo; Embryonic Development; Epithelial; Epithelial Cells; Epithelium; Exposure to; Flow Cytometry; Functional RNA; Gene Expression; Gene Expression Profile; Genes; Genetic; Genetic Models; Genetic Recombination; Genetically Engineered Mouse; Homeostasis; Human; Individual; Injury; Lead; Link; Lung; Main Bronchus; Maintenance; MicroRNAs; Molecular; Multipotent Stem Cells; Mus; Naphthalene; Naphthalenes; Numbers; Organism; Pathway interactions; Physiologic pulse; Population; Protein Overexpression; Pseudostratified Epithelium; Pulse taking; Regulator Genes; Relative (related person); Risk; Role; Small RNA; Stem cells; Sulfur Dioxide; System; Tamoxifen; Testing; Therapeutic; Time; Trachea; Transgenic Organisms; Work; aptamer; direct application; genetic manipulation; improved; in vivo; loss of function; lung development; member; progenitor; repaired; research study; self-renewal; time use; tool; transcription factor

DESCRIPTION (provided by applicant): Relatively little is known about the molecular pathways defining the epithelial progenitor cells of the embryonic and adult lung and their role in lung development, steady state homeostasis and repair after damage caused by environmental and other agents. In this competitive renewal we will build on previous discoveries to obtain new information about basic mechanisms controlling the specification, proliferation, and differentiation of lung epithelial progenitor/stem cells, using the mouse as a model genetic organism. In Aim 1 we will test the hypothesis that the developmental potential of epithelial progenitor cells in the distal tips of the embryonic lung changes in an orderly way over time so that early born descendants give rise to cells in the proximal airways while late born daughters give rise to alveolar cells. For this aim we have already generated two new lines of genetically engineered mice, Id2-GFP and Id2-CreER. These will enable us to follow the developmental fate of distal tip cells in vivo, isolate the cells at different times using flow cytometry, analyze their transcriptome using gene arrays, and test the role of candidate regulatory genes using gain and loss of function experiments. In Aim 2 we will focus on the adult trachea and main bronchi which has a pseudostratified epithelium, as in a large proportion of the human airways. We will test the relative contribution of basal stem cells and secretory (Clara) cells in maintaining this region and in repairing it after injury by exposure to naphthalene or sulfur dioxide. This will be achieved using new K5-CreEr and Scgb1a1-CreEr alleles for fate mapping and gene manipulation. We will also elucidate basic mechanisms regulating the specification, self-renewal and differentiation of basal stem cells, focusing initially on the role of the transcription factor, Sox2. Finally, in a high risk Aim 3, we will test the hypothesis that small non-coding RNAs known as microRNAs can reversibly enhance the self-renewal potential of progenitor cells in the adult lung, either in the steady state or during repair. To do this we will initially inducibly express in basal and Clara cells the miR17-92 gene that encodes a conserved cluster of six microRNAs. We have already shown that this cluster enhances the proliferation of epithelial progenitors in the embryonic lung at the expense of their differentiation and we will now test this in adult progenitors. We will then test the activity of individual miR17-92 members or combinations of them. In the long term we will ask whether this activity can be obtained by direct application of double stranded small RNAs linked to cell-specific aptamers. PROJECT NARRATIVE. This work will increase our understanding of basic mechanisms regulating the embryonic development of the lung and the ability of the epithelial cells to undergo repair in the adult. In the long term the work may lead to potential therapies or predictive assays for pathological conditions related to defects in the supply, proliferation or differentiation of progenitor/stem cells in the human lung.

描述（由申请人提供）：关于定义胚胎和成人肺的上皮祖细胞的分子途径及其在肺发育、稳态稳态和环境和其他因素造成的损伤后的修复中的作用，人们知之甚少。在这次竞争性更新中，我们将在以前的发现的基础上，使用小鼠作为模型遗传生物，获得有关控制肺上皮祖细胞/干细胞的规范、增殖和分化的基本机制的新信息。在目标 1 中，我们将检验这样的假设：胚胎肺远端上皮祖细胞的发育潜力随着时间的推移以有序的方式变化，以便早出生的后代在近端气道中产生细胞，而晚出生的女儿则产生近端气道中的细胞到肺泡细胞。为了这个目标，我们已经培育了两个新的基因工程小鼠品系：Id2-GFP 和 Id2-CreER。这些将使我们能够跟踪体内远端细胞的发育命运，使用流式细胞术在不同时间分离细胞，使用基因阵列分析其转录组，并使用功能获得和丧失实验测试候选调控基因的作用。在目标 2 中，我们将重点关注成人气管和主支气管，它们具有假复层上皮，就像人类气道的大部分一样。我们将测试基底干细胞和分泌细胞（Clara）在维持该区域以及在暴露于萘或二氧化硫而受伤后修复该区域方面的相对贡献。这将通过使用新的 K5-CreEr 和 Scgb1a1-CreEr 等位基因进行命运图谱和基因操作来实现。我们还将阐明调节基底干细胞的规范、自我更新和分化的基本机制，首先关注转录因子 Sox2 的作用。最后，在高风险目标 3 中，我们将测试以下假设：被称为 microRNA 的小非编码 RNA 可以可逆地增强成体肺中祖细胞的自我更新潜力，无论是在稳定状态还是在修复过程中。为此，我们首先将在基底细胞和 Clara 细胞中诱导表达 miR17-92 基因，该基因编码六个保守的 microRNA 簇。我们已经表明，该簇增强了胚胎肺中上皮祖细胞的增殖，但代价是它们的分化，现在我们将在成年祖细胞中测试这一点。然后我们将测试单个 miR17-92 成员或它们组合的活性。从长远来看，我们会问是否可以通过直接应用与细胞特异性适体连接的双链小RNA来获得这种活性。项目叙述。这项工作将增加我们对调节肺胚胎发育的基本机制以及上皮细胞在成人中进行修复的能力的理解。从长远来看，这项工作可能会为与人肺中祖细胞/干细胞的供应、增殖或分化缺陷相关的病理状况带来潜在的治疗或预测分析。