Derivation of Epithelial-Only Airway Organoids from Human Pluripotent Stem Cells.
Derivation of Epithelial-Only Airway Organoids from Human Pluripotent Stem Cells.
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DOI：
10.1002/cpsc.51
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发表时间：
2018-05-01
影响因子：
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通讯作者：
Kotton, Darrell N
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文献类型：
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作者：
McCauley, Katherine B;Hawkins, Finn;Kotton, Darrell N
从人类多能干细胞（PSCs）高效生成功能性气道上皮类器官的新方案，将是朝着肺部疾病的有效疾病建模、药物筛选和基于细胞的疗法迈出的重要一步。本单元描述了一种利用特定阶段的信号通路调控方法，通过关键的发育中间体将细胞分化为近端气道上皮。细胞经定型内胚层（DE）定向分化为前肠前部，然后特化为NKX2 - 1⁺肺上皮祖细胞。这些肺祖细胞使用细胞表面标记物分选进行纯化，并在特定的培养条件下重新接种，以形成三维的、仅含上皮的气道类器官。这种使用无血清、特定培养基的定向分化方法还包括评估DE诱导的方案、NKX2 - 1特化效率和富集的细胞内流式细胞术分析，以及气道类器官的表征和扩增方法。综上所述，这代表了一种从人类PSCs生成可扩增的气道类器官的高效且可重复的方法，可用于众多下游应用。© 2018由约翰威立父子公司出版
New protocols to efficiently generate functional airway epithelial organoids from human pluripotent stem cells (PSCs) would represent a major advance towards effective disease modeling, drug screening and cell based therapies for lung disorders. This unit describes an approach using stage-specific signaling pathway manipulation to differentiate cells to proximal airway epithelium via key developmental intermediates. Cells are directed via definitive endoderm (DE) to anterior foregut, and then specified to NKX2-1+ lung epithelial progenitors. These lung progenitors are purified using cell surface marker sorting and replated in defined culture conditions to form three-dimensional, epithelial-only airway organoids. This directed differentiation approach using serum-free, defined media also includes protocols for evaluation of DE induction, intracellular FACS analysis of NKX2-1 specification efficiency and enrichment, and approaches for characterization and expansion of airway organoids. Taken together, this represents an efficient and reproducible approach to generate expandable airway organoids from human PSCs for use in numerous downstream applications. © 2018 by John Wiley & Sons, Inc.