title: Directed Induction of Functional Multi-ciliated Cells in Proximal Airway Epithelial Spheroids from Human Pluripotent Stem Cells.
abstract: Multi-ciliated airway cells (MCACs) play a role in mucociliary clearance of the lung. However, the efficient induction of functional MCACs from human pluripotent stem cells has not yet been reported. Using carboxypeptidase M (CPM) as a surface marker of NKX2-1(+)-ventralized anterior foregut endoderm cells (VAFECs), we report a three-dimensional differentiation protocol for generating proximal airway epithelial progenitor cell spheroids from CPM(+) VAFECs. These spheroids could be induced to generate MCACs and other airway lineage cells without alveolar epithelial cells. Furthermore, the directed induction of MCACs and of pulmonary neuroendocrine lineage cells was promoted by adding DAPT, a Notch pathway inhibitor. The induced MCACs demonstrated motile cilia with a "9 + 2" microtubule arrangement and dynein arms capable of beating and generating flow for mucociliary transport. This method is expected to be useful for future studies on human airway disease modeling and regenerative medicine.
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Satoshi
authors.initials: S
authors.lastname: Konishi
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan. Electronic address: a0009650@kuhp.kyoto-u.ac.jp.
authors.forename: Shimpei
authors.initials: S
authors.lastname: Gotoh
authors.affiliation: Laboratory of Biological Science, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Suita 565-0871, Japan.
authors.forename: Kazuhiro
authors.initials: K
authors.lastname: Tateishi
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Yuki
authors.initials: Y
authors.lastname: Yamamoto
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Korogi
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Tadao
authors.initials: T
authors.lastname: Nagasaki
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Hisako
authors.initials: H
authors.lastname: Matsumoto
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Shigeo
authors.initials: S
authors.lastname: Muro
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Toyohiro
authors.initials: T
authors.lastname: Hirai
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Isao
authors.initials: I
authors.lastname: Ito
authors.affiliation: Laboratory of Biological Science, Graduate School of Frontier Biosciences and Graduate School of Medicine, Osaka University, Suita 565-0871, Japan.
authors.forename: Sachiko
authors.initials: S
authors.lastname: Tsukita
authors.affiliation: Department of Respiratory Medicine, Graduate School of Medicine, Kyoto University, Kyoto 606-8507, Japan.
authors.forename: Michiaki
authors.initials: M
authors.lastname: Mishima
mesh_terms: D018929:Cell Culture Techniques; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D019070:Cell Lineage / Q000235:genetics; D002478:Cells, Cultured; D002923:Cilia; D004398:Dyneins / Q000378:metabolism; D004707:Endoderm / Q000166:cytology / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism / Q000648:ultrastructure; D058851:GPI-Linked Proteins / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D006801:Humans; D007422:Intestines / Q000166:cytology; D008168:Lung / Q000166:cytology*; D008666:Metalloendopeptidases / Q000378:metabolism; D008854:Microscopy, Electron; D008856:Microscopy, Fluorescence; D008870:Microtubules / Q000378:metabolism; D039904:Pluripotent Stem Cells / Q000166:cytology* / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D018874:Spheroids, Cellular / Q000166:cytology* / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D058851:GPI-Linked Proteins; C058479:carboxypeptidase M; D008666:Metalloendopeptidases; D004398:Dyneins