Abstract
Airway epithelial cells are of great interest for research on lung development, regeneration and disease modeling. This protocol describes how to generate cystic fibrosis (CF) transmembrane conductance regulator protein (CFTR)-expressing airway epithelial cells from human pluripotent stem cells (PSCs). The stepwise approach from PSC culture to differentiation into progenitors and then mature epithelia with apical CFTR activity is outlined. Human PSCs that were inefficient at endoderm differentiation using our previous lung differentiation protocol were able to generate substantial lung progenitor cell populations. Augmented CFTR activity can be observed in all cultures as early as at 35 d of differentiation, and full maturation of the cells in air-liquid interface cultures occurs in <5 weeks. This protocol can be used for drug discovery, tissue regeneration or disease modeling.
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Acknowledgements
We thank J.R. Riordan (University of North Carolina) for providing the monoclonal antibodies specific to CFTR (nos. 450, 596 and 660). This work was funded by the Canadian Institutes of Health Research (GPG-102171) to C.E.B. and J.R. A.P.W. was the recipient of the Cystic Fibrosis Canada postdoctoral fellowship. The CA1 hESC line was obtained from A. Nagy (Mount Sinai Hospital). H9 hESCs were obtained from The WiCell Research Institute. CF iPSC line GM00997 and GM04320 were obtained from J. Ellis (Hospital for Sick Children).
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A.P.W., J.R. and C.E.B. conceived the study and the experimental design. A.P.W. performed and analyzed the experiments and wrote the manuscript. J.G., S.C. and S.X. performed the experiments and helped in preparing manuscript figures. All authors edited and approved the final manuscript.
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Wong, A., Chin, S., Xia, S. et al. Efficient generation of functional CFTR-expressing airway epithelial cells from human pluripotent stem cells. Nat Protoc 10, 363–381 (2015). https://doi.org/10.1038/nprot.2015.021
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DOI: https://doi.org/10.1038/nprot.2015.021
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