The in vitro multilineage differentiation and maturation of lung and airway cells from human pluripotent stem cell–derived lung progenitors in 3D
- 1 March 2021
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature Protocols
- Vol. 16 (4), 1802-1829
- https://doi.org/10.1038/s41596-020-00476-z
Abstract
Lung and airway epithelial cells generated in vitro from human pluripotent stem cells (hPSCs) have applications in regenerative medicine, modeling of lung disease, drug screening and studies of human lung development. Here, we describe a strategy for directed differentiation of hPSCs into mature lung and airway epithelial cells obtained through maturation of NKX2.1+ hPSC-derived lung progenitors in a 3D matrix of collagen I in the absence of glycogen synthase kinase 3 inhibition. This protocol is an extension of our previously published protocol on the directed differentiation of lung and airway epithelium from hPSCs that modifies the technique and offers additional applications. This protocol is conducted in defined media conditions, has a duration of 50–80 d, does not require reporter lines and results in cultures containing mature alveolar type II and I cells as well as airway basal, ciliated, club and neuroendocrine cells. We also present a flow cytometry strategy to assess maturation in the cultures. Several of these populations, including mature NGFR+ basal cells, can be prospectively isolated by cell sorting and expanded for further investigation.Funding Information
- U.S. Department of Health & Human Services | NIH | National Heart, Lung, and Blood Institute (1U01HL134760, 1R01HL142727)
This publication has 51 references indexed in Scilit:
- Type 2 alveolar cells are stem cells in adult lungJCI Insight, 2013
- Stem cells of the respiratory system: From identification to differentiation into functional epitheliumBioEssays, 2012
- Pathogenesis of acute respiratory illness caused by human parainfluenza virusesCurrent Opinion in Virology, 2012
- Generation of Multipotent Lung and Airway Progenitors from Mouse ESCs and Patient-Specific Cystic Fibrosis iPSCsCell Stem Cell, 2012
- Evidence for type II cells as cells of origin of K-Ras–induced distal lung adenocarcinomaProceedings of the National Academy of Sciences of the United States of America, 2012
- HTII-280, a Biomarker Specific to the Apical Plasma Membrane of Human Lung Alveolar Type II CellsJournal of Histochemistry & Cytochemistry, 2010
- Alveolar Surfactant Homeostasis and the Pathogenesis of Pulmonary DiseaseAnnual Review of Medicine, 2010
- Basal cells as stem cells of the mouse trachea and human airway epitheliumProceedings of the National Academy of Sciences of the United States of America, 2009
- MUC1 MucinAmerican Journal of Respiratory Cell and Molecular Biology, 2008
- T1α, a lung type I cell differentiation gene, is required for normal lung cell proliferation and alveolus formation at birthDevelopmental Biology, 2003