Methods for the amplification and identification of airway stem cells

描述（由申请人提供）：慢性肺部疾病具有上皮修复缺陷的共同特征，导致组织稳态的丧失和肺功能的进一步恶化。本提案的目标是提高我们在健康和疾病中调节组织干细胞的能力，并制定通过刺激原位驻留细胞或通过开发基于细胞的治疗策略来增强上皮再生的策略。我们最近开发并鉴定了通过条件表达n端截断稳定形式的¿-catenin蛋白扩增导气管内的干细胞的小鼠。本应用程序的目标是通过条件表达N¿-catenin来开发一种小鼠模型，以实现干细胞表型的可逆体内稳定。该模型将用于揭示上皮细胞分化的关键阶段，在发育和成人肺易受稳定¿-catenin的影响。目的1将建立小鼠，允许在发育和成年肺中表达CCSP的细胞中有条件地调节稳定的-catenin。该模型将用于确定稳定的-catenin在小鼠细支气管上皮细胞中阻止分化特性的可逆性。目的2将确定稳定的¿-catenin的分化调节作用是否仅限于干细胞，还是可以更广泛地应用于调节其他祖细胞类型（Clara细胞）的分化潜力。这项应用的结果将决定旨在稳定¿-catenin的基因干预（以及外推的药物干预）是否可以用作气道干细胞在体内扩增的工具。这些实验是进一步发展针对内源性肺组织干细胞的基于细胞的治疗干预的必要前提。公共卫生相关性：肺气道上皮细胞在健康中发挥着重要的功能，而这些功能在肺部疾病中丢失。潜在的缺陷是上皮修复或重建正常功能的能力。本课题将重点研究气道损伤后上皮修复的新方法。目的是利用调节上皮细胞数量的信号通路来修复气道上皮。这些研究将为开发能够促进慢性肺部疾病患者的上皮修复和肺功能恢复的药物提供新的信息。

DESCRIPTION (provided by applicant): Chronic lung diseases share the common feature of epithelial repair defects that contribute to loss of tissue homeostasis and further deterioration of lung function. Goals of this proposal are to improve our ability regulate tissue stem cells in health and disease, and to develop strategies to enhance epithelial regeneration either through stimulation of resident cells in situ or through development of cell-based therapeutic strategies. We have recently developed and characterized mice in which stem-like cells within conducting airways are amplified through conditional expression of an N-terminally truncated stabilized form of the ¿-catenin protein. Goals of this application are to develop a mouse model for reversible in vivo stabilization of the stem cell phenotype through conditional expression of ?N¿-catenin. This model will be used to reveal critical stages of epithelial cell differentiation in the developing and adult lung that are susceptible to the effects of stabilized ¿-catenin. Aim 1 will establish mice allowing conditional regulation of stabilized ¿-catenin within CCSP- expressing cells of the developing and adult lung. This model will be used to determine the reversibility of differentiation-arresting properties of stabilized ¿-catenin in mouse bronchiolar epithelial cells. Aim 2 will determine whether the differentiation modulating effects of stabilized ¿-catenin are limited to stem cells or can be more broadly applied to modulate the differentiation potential of other progenitor cell types (Clara cells). Results from this application will determine whether genetic interventions (and by extrapolation, pharmacologic interventions) aimed at stabilizing ¿-catenin can be used as a tool for amplification of airway stem cells in vivo. These experiments are a necessary prerequisite for the further development of therapeutic interventions aimed at cell-based therapies targeted towards endogenous lung tissue stem cells. PUBLIC HEALTH RELEVANCE: Epithelial cells that line airways of the lung fulfill critical functions in health that are lost in the setting of lung disease. The underlying defect is the capacity of the epithelium to repair or to re-establish normal function. Research to be conducted in this proposal will focus on the development of new methods to repair the epithelium of airways following injury. Goals are to take advantage of signaling pathways that regulate the number of epithelial cells capable of repairing the airway epithelium. These studies will provide new information for the development of drugs capable of enhancing epithelial repair and restoration of lung function to patients with chronic lung disease.