Functional human iPSC-derived alveolar-like cells cultured in a miniaturized 96‑Transwell air–liquid interface model

Abstract

Abstract In order to circumvent the limited access and donor variability of human primary alveolar cells, directed differentiation of human pluripotent stem cells (hiPSCs) into alveolar-like cells, provides a promising tool for respiratory disease modeling and drug discovery assays. In this work, a unique, miniaturized 96-Transwell microplate system is described where hiPSC-derived alveolar-like cells were cultured at an air–liquid interface (ALI). To this end, hiPSCs were differentiated into lung epithelial progenitor cells (LPCs) and subsequently matured into a functional alveolar type 2 (AT2)-like epithelium with monolayer-like morphology. AT2-like cells cultured at the physiological ALI conditions displayed characteristics of AT2 cells with classical alveolar surfactant protein expressions and lamellar-body like structures. The integrity of the epithelial barriers between the AT2-like cells was confirmed by applying a custom-made device for 96-parallelized transepithelial electric resistance (TEER) measurements. In order to generate an IPF disease-like phenotype in vitro, the functional AT2-like cells were stimulated with cytokines and growth factors present in the alveolar tissue of IPF patients. The cytokines stimulated the secretion of pro-fibrotic biomarker proteins both on the mRNA (messenger ribonucleic acid) and protein level. Thus, the hiPSC-derived and cellular model system enables the recapitulation of certain IPF hallmarks, while paving the route towards a miniaturized medium throughput approach of pharmaceutical drug discovery.