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. 2020 Jul 24;1(2):100068.
doi: 10.1016/j.xpro.2020.100068. eCollection 2020 Sep 18.

Protocol for Differentiation of Human iPSCs into Pulmonary Neuroendocrine Cells

Pooja Hor  1   2 , Justin K Ichida  2 , Zea Borok  1   3   4 , Amy L Ryan  1   2
Affiliations

Protocol for Differentiation of Human iPSCs into Pulmonary Neuroendocrine Cells

Pooja Hor et al. STAR Protoc. .

Abstract

Pulmonary neuroendocrine cells (PNECs) are sensory cells within the lung airway epithelia. Here, we provide a detailed protocol for generating induced PNECs (iPNECs) from human induced pluripotent stem cells (iPSCs). The cellular and molecular profile of iPNECs resembles primary human PNECs. Primary human PNECs are exceedingly rare, comprising only 1% of the adult lung. Therefore, a self-renewing source of patient-specific iPNECs facilitates the creation of reproducible human cellular models to study lung diseases characterized by PNEC dysfunction. For complete details on the use and execution of this protocol, please refer to Hor et al. (2020).

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Conflict of interest statement

J.K.I. is a co-founder of AcuraStem Incorporated. J.K.I. declares that he is bound by confidentiality agreements that prevent him from disclosing details of his financial interests in this work.

Figures

None
Graphical abstract
Figure 1
Figure 1
Effect of Plating Density on Differentiation Efficiency (A–C) Efficiency of definitive endoderm (DE) derivation at day 5 varies based on iPSC confluency before starting the differentiation. Representative immunofluorescence (IF) images of cells show expression of FOXA2 (green) and SOX17 (red). Nuclei are counterstained with DAPI (blue). Differentiation started with too few cells (<70% confluency) or too many cells results in poor (A) or low (B) DE differentiation. Differentiation started with optimal plating density of 70%–80% confluency at the time of starting the experiment results in best DE differentiation (C). Scale bars, 200 μm.
Figure 2
Figure 2
Generation of Lung Progenitors during iPNEC Differentiation from Human iPSCs (A–D) Cultures stained for (A) lung endoderm marker NKX2.1 (green), (B) distal progenitor marker SOX9 (green), (C) proximal airway progenitor marker SOX2 (green) and (D) tight junction marker ZO1 (green) at day 17 of differentiation. Nuclei are counterstained with DAPI (blue). Scale bars, 200 μm.
See this image and copyright information in PMC

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