Human iPS cell–derived alveolar epithelium repopulates lung extracellular matrix
Mahboobe Ghaedi, … , Eric S. White, Laura E. Niklason
Mahboobe Ghaedi, … , Eric S. White, Laura E. Niklason
Published November 1, 2013; First published October 25, 2013
Citation Information: J Clin Invest. 2013;123(11):4950-4962. https://doi.org/10.1172/JCI68793.
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Categories: Technical Advance Pulmonology

Human iPS cell–derived alveolar epithelium repopulates lung extracellular matrix

Abstract

The use of induced pluripotent stem cells (iPSCs) has been postulated to be the most effective strategy for developing patient-specific respiratory epithelial cells, which may be valuable for lung-related cell therapy and lung tissue engineering. We generated a relatively homogeneous population of alveolar epithelial type II (AETII) and type I (AETI) cells from human iPSCs that had phenotypic properties similar to those of mature human AETII and AETI cells. We used these cells to explore whether lung tissue can be regenerated in vitro. Consistent with an AETII phenotype, we found that up to 97% of cells were positive for surfactant protein C, 95% for mucin-1, 93% for surfactant protein B, and 89% for the epithelial marker CD54. Additionally, exposing induced AETII to a Wnt/β-catenin inhibitor (IWR-1) changed the iPSC-AETII–like phenotype to a predominantly AETI-like phenotype. We found that of induced AET1 cells, more than 90% were positive for type I markers, T1α, and caveolin-1. Acellular lung matrices were prepared from whole rat or human adult lungs treated with decellularization reagents, followed by seeding these matrices with alveolar cells derived from human iPSCs. Under appropriate culture conditions, these progenitor cells adhered to and proliferated within the 3D lung tissue scaffold and displayed markers of differentiated pulmonary epithelium.

Authors

Mahboobe Ghaedi, Elizabeth A. Calle, Julio J. Mendez, Ashley L. Gard, Jenna Balestrini, Adam Booth, Peter F. Bove, Liqiong Gui, Eric S. White, Laura E. Niklason

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Figure 5

iPSC-derived AETII recellularized 3D rat lung tissue scaffolds in a bioreactor.

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iPSC-derived AETII recellularized 3D rat lung tissue scaffolds in a bior...
(A) H&E staining of decellularized rat lung; (B and C) H&E staining of 3- and 7-day seeded rat lung with iPSC-derived AETII cells cultured in a bioreactor. Scale bars: 25 μm. (D–F) Immunofluorescent staining for pro-SPC in AETII seeded cells at day 3. (D) DAPI staining; (E) pro-SPC; (F) merge (arrows in F indicate cells positive for pro-SPC). (G–I) Immunostaining for NKX2.1 at day 7. (G) DAPI, (H) NKX2.1, (I) merge (arrows in 5I indicate cells positive for NKX2.1) (J–M) Caspase and PCNA immunostaining at day 7 (arrows indicates cells positive for PCNA in L and caspase in M). Scale bar: 25 μm. (N) Proliferation at day 7 compared with day 3. iPSC-AETII displayed a significantly increased fractional proliferation (P < 0.05) after 7 days when they were stained for PCNA (y axis, percentage proliferation based on the number of positive nuclei stained for PCNA) (O) Immunostaining of the few engrafted epithelial cells that acquired flattened morphology, positive for T1α, and negative for NKX2.1 at day 7. Scale bar: 63 μm. Arrows in O indicate cells positive for T1α. (P) Flow cytometry for SPC, T1α, CCSP, p63, and SOX2 before and after seeding into rat lung scaffold in bioreactor. The number of SPC-positive cells decreased during 7-day culture, while the number of cells positive for T1α increased from 9% to 31.2%. All differentiated cells from iPSCs were negative for CCSP, p63, and SOX2 before and after cell seeding.