Human induced pluripotent stem cell-derived alveolar type 2 cells mature at air-liquid interface and respond to airborne stimuli

Abstract

Human alveolar type II cells (AT2s) are progenitors of the alveolar epithelium and are among the pulmonary cells that are directly exposed to inhaled stimuli. Primary human AT2s can be cultured in three-dimensional alveolospheres, but are difficult to culture in the physiologically relevant air-liquid interface (ALI) format. Human induced pluripotent stem cells (iPSCs) can be directed to differentiate to iPSC-derived AT2s (iAT2s) in alveolospheres, where they transcriptomically resemble fetal lung. Here we report the successful adaptation of iAT2s to ALI culture, which promotes their maturation and permits exposure to inhaled stimuli. We transcriptomically profile iAT2s cultured at ALI and find that they mature as they downregulate cell cycle-associated transcripts. We then evaluate the extent of iAT2 maturation at ALI within the developmental context by comparison to primary AT2s. We find that iAT2s at ALI are more similar to primary AT2s than iAT2s cultured as spheres, and that differences are driven by primary AT2s’ response to immune stimuli. We then test the capacity of iAT2s to respond to immune stimuli by infecting with SARS-CoV-2. We find that iAT2s mount an epithelial-intrinsic interferon and inflammatory response to SARS-CoV-2 infection, and can serve as a platform for testing antiviral therapeutics. Finally, we demonstrate that iAT2s at ALI respond to cigarette smoke and electronic cigarette vapor, enabling the direct comparison of these common inhaled stimuli. Overall, we describe a novel disease modeling platform that will enable exploration of gene-environment interactions unique to inhaled exposures of the alveolar epithelium.