Human pulmonary alveolar type II cells (AT2s) are facultative progenitors of the distal lung epithelium and secrete surfactant, a protein-lipid mixture that lowers alveolar surface tension and prevents alveolar collapse during expiration. AT2s exist at a physiological interface between inhaled air and pulmonary vasculature, and are among the pulmonary cell types that are directly exposed to inhaled environmental stimuli such as respiratory viruses and cigarette smoke. Primary human AT2s can be cultured in vitro in three-dimensional organoids known as alveolospheres, but are difficult to culture in the physiologically relevant air-liquid interface (ALI) format due to their tendency to lose their AT2 phenotype and senesce in culture. Human induced pluripotent stem cells (iPSCs) can be directed to differentiate to iPSC-derived AT2s (iAT2s) in three-dimensional spheres, where they transcriptomically resemble primary human fetal lung. Here we report the successful adaptation of iAT2s to ALI culture, which promotes their maturation and permits exposure to inhaled environmental stimuli. We transcriptomically profile iAT2s cultured at ALI and find that they upregulate key markers of AT2 maturation as they downregulate cell cycle-associated transcripts. We then evaluate the extent of iAT2 maturation at ALI within the developmental context by transcriptomic comparison to cultured and freshly isolated primary AT2s. We find that iAT2s cultured at ALI are more similar to primary AT2s than iAT2s cultured as spheres, and that the main differences between iAT2s at ALI and primary AT2s are due to primary AT2s’ response to immune stimuli. We then test the capacity of iAT2s to respond to immune stimuli and serve as useful in vitro model system for human respiratory viral infections by infecting iAT2s at ALI with SARS-CoV-2, the virus that causes Coronavirus Disease 2019 (COVID-19). We find that iAT2s are permissive to SARS-CoV-2 infection, mount an epithelial-intrinsic interferon and inflammatory response to infection, and can serve as an in vitro platform for testing antiviral therapeutics. Finally, we demonstrate that iAT2s at ALI also have utility as a system for modeling the response to cigarette smoke and electronic cigarette vapor, enabling the direct comparison of these two common inhaled noxious stimuli. Overall, we describe a novel disease modeling platform that enables future exploration of gene-environment interactions unique to inhaled exposures of the alveolar epithelium.
| Identifer | oai:union.ndltd.org:bu.edu/oai:open.bu.edu:2144/43763 |
| Date | 01 February 2022 |
| Creators | Abo, Kristine M. |
| Contributors | Wilson, Andrew |
| Source Sets | Boston University |
| Language | en_US |
| Detected Language | English |
| Type | Thesis/Dissertation |
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