Progenitor identification and SARS-CoV-2 infection in human distal lung organoids.

Salahudeen, Ameen A; Choi, Shannon S; Rustagi, Arjun; Zhu, Junjie; van Unen, Vincent; de la O, Sean M; Flynn, Ryan A; Margalef-Català , Mar; Santos, António J M; Ju, Jihang; Batish, Arpit; Usui, Tatsuya; Zheng, Grace X Y; Edwards, Caitlin E; Wagar, Lisa E; Luca, Vincent; Anchang, Benedict; Nagendran, Monica; Nguyen, Khanh; Hart, Daniel J; Terry, Jessica M; Belgrader, Phillip; Ziraldo, Solongo B; Mikkelsen, Tarjei S; Harbury, Pehr B; Glenn, Jeffrey S; Garcia, K Christopher; Davis, Mark M; Baric, Ralph S; Sabatti, Chiara; Amieva, Manuel R; Blish, Catherine A; Desai, Tushar J; Kuo, Calvin J

The distal lung contains terminal bronchioles and alveoli that facilitate gas exchange. Three-dimensional in vitro human distal lung culture systems would strongly facilitate investigation of pathologies including interstitial lung disease, cancer, and SARS-CoV-2-associated COVID-19 pneumonia. We generated long-term feeder-free, chemically defined culture of distal lung progenitors as organoids derived from single adult human alveolar epithelial type II (AT2) or KRT5+ basal cells. AT2 organoids exhibited AT1 transdifferentiation potential while basal cell organoids developed lumens lined by differentiated club and ciliated cells. Single cell analysis of basal organoid KRT5+ cells revealed a distinct ITGA6+ITGB4+ mitotic population whose proliferation further segregated to a TNFRSF12Ahi subfraction comprising ~10% of KRT5+ basal cells, residing in clusters within terminal bronchioles and exhibiting enriched clonogenic organoid growth activity. Distal lung organoids were created with apical-out polarity to display ACE2 on the exposed external surface, facilitating SARS-CoV-2 infection of AT2 and basal cultures and identifying club cells as a novel target population. This long-term, feeder-free organoid culture of human distal lung, coupled with single cell analysis, identifies unsuspected basal cell functional heterogeneity and establishes a facile in vitro organoid model for human distal lung infections including COVID-19-associated pneumonia.