title: Alveolar epithelial differentiation of human induced pluripotent stem cells in a rotating bioreactor.
abstract: Traditional stem cell differentiation protocols make use of a variety of cytokines including growth factors (GFs) and inhibitors in an effort to provide appropriate signals for tissue specific differentiation. In this study, iPSC-derived type II pneumocytes (iPSC-ATII) as well as native isolated human type II pneumocytes (hATII) were differentiated toward a type I phenotype using a unique air-liquid interface (ALI) system that relies on a rotating apparatus that mimics in vivo respiratory conditions. A relatively homogenous population of alveolar type II-like cells from iPSC was first generated (iPSC-ATII cells), which had phenotypic properties similar to mature human alveolar type II cells. iPSC-ATII cells were then cultured in a specially designed rotating culture apparatus. The effectiveness of the ALI bioreactor was compared with the effectiveness of small molecule-based differentiation of type II pneumocytes toward type 1 pneumocytes. The dynamics of differentiation were examined by quantitative reverse transcriptase polymerase chain reaction (qRT-PCR), flow cytometry and immunocytochemistry. iPSC-ATII and hATII cells cultured in the ALI bioreactor had higher levels of type I markers, including aquaporin-5(AQ5), caveolin-1, and T1α, at both the RNA and protein levels as compared with the flask-grown iPSC-ATII and hATII that had been treated with small molecules to induce differentiation. In summary, this study demonstrates that a rotating bioreactor culture system that provides an air-liquid interface is a potent inducer of type I epithelial differentiation for both iPS-ATII cells and hATII cells, and provides a method for large-scale production of alveolar epithelium for tissue engineering and drug discovery.
authors.affiliation: Department of Anesthesiology, Yale University, New Haven, CT 06520, USA; Department of Biomedical Engineering, Yale University, New Haven, CT 06520, USA.
authors.forename: Mahboobe
authors.initials: M
authors.lastname: Ghaedi
authors.forename: Julio J
authors.initials: JJ
authors.lastname: Mendez
authors.forename: Peter F
authors.initials: PF
authors.lastname: Bove
authors.forename: Amogh
authors.initials: A
authors.lastname: Sivarapatna
authors.forename: Micha Sam B
authors.initials: MS
authors.lastname: Raredon
authors.forename: Laura E
authors.initials: LE
authors.lastname: Niklason
mesh_terms: D051402:Aquaporin 5 / Q000378:metabolism; D019149:Bioreactors; D051242:Caveolin 1 / Q000378:metabolism; D002454:Cell Differentiation; D002478:Cells, Cultured; D004847:Epithelial Cells / Q000166:cytology*; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology*; D008562:Membrane Glycoproteins; D008565:Membrane Proteins / Q000378:metabolism; D011650:Pulmonary Alveoli / Q000166:cytology*; D000098853:Podoplanin
chemical_list: D051402:Aquaporin 5; D051242:Caveolin 1; D008562:Membrane Glycoproteins; D008565:Membrane Proteins; C496036:AQP5 protein, human; C495540:CAV1 protein, human; C117468:PDPN protein, human; D000098853:Podoplanin
keywords: ALI; Air–liquid interface; Alveolar epithelium; Bioreactor; Induced pluripotent stem (iPS); air–liquid interface; hATI; hATII; human alveolar type I; human alveolar type II; iPSC-ATII; iPSC-derived alveolar type II