Generation of expandable multipotential distal lung progenitors from human pluripotent stem cells that model idiopathic pulmonary fibrosis [scRNA-Seq]. Generation of expandable multipotential distal lung progenitors from human pluripotent stem cells that model idiopathic pulmonary fibrosis [scRNA-Seq]

Human lungs contain unique cell populations in distal respiratory airways and terminal bronchioles (RA/TRB) that accumulate in patients with lung injury and idiopathic pulmonary fibrosis (IPF), a lethal lung disease. As these populations are absent in rodents, deeper understanding requires a human in vitro model. Here we convert human pluripotent stem cells into expandable spheres, called induced respiratory airway progenitors (iRAPs), consisting of ~99% RA/TRB-associated cell types. One hPSC can give rise to 1010 iRAP cells. We differentiate iRAPs through a stage consistent with transitional AT2 like cells into a population corresponding to mature AT1 cells with 95% purity. iRAPs with deletion of HPS1, which causes pulmonary fibrosis in humans, replicate the aberrant differentiation and recruitment of profibrotic fibroblasts observed in IPF, indicating that intrinsic dysfunction of RA/TRB-associated alveolar progenitors contributes to HPS1-related IPF. iRAPs may provide a system suitable for IPF drug discovery and validation. Overall design: We performed single cell RNA sequencing on iRAPs, iAT1 and transitional type 2 alveolar epithelial. The samples were dissociated to single cells and resuspended at a concentration of 5000 cells/ul for sequencing.

人类肺部远端呼吸道与终末细支气管（respiratory airways and terminal bronchioles, RA/TRB）中存在独特的细胞群，这类细胞群会在肺损伤患者以及致死性肺部疾病特发性肺纤维化（idiopathic pulmonary fibrosis, IPF）患者体内异常蓄积。由于啮齿类动物体内不存在此类细胞群，对其开展深入研究需依托人类体外模型。
本研究将人类多能干细胞（human pluripotent stem cells, hPSCs）诱导为可扩增的细胞球，命名为诱导型呼吸道气道祖细胞（induced respiratory airway progenitors, iRAPs），其中约99%为RA/TRB相关细胞类型。单个人类多能干细胞可产生10^10个iRAP细胞。
我们可将iRAPs分化至与过渡态AT2样细胞一致的阶段，进而获得纯度达95%的成熟AT1细胞群。
当iRAPs中HPS1基因发生缺失（该基因在人类中可引发肺纤维化）时，其异常分化以及特发性肺纤维化中观察到的促纤维化成纤维细胞募积现象将得到重现，这表明RA/TRB相关肺泡祖细胞的内在功能异常参与了HPS1相关特发性肺纤维化的发生。iRAPs有望成为适用于特发性肺纤维化药物研发与验证的研究体系。
实验整体设计：我们对iRAPs、iAT1以及过渡态2型肺泡上皮细胞开展了单细胞RNA测序。样本被解离为单细胞后，以5000个细胞/微升的浓度重悬，用于后续测序。