A comparative study of in vitro air–liquid interface culture models of the human airway epithelium evaluating cellular heterogeneity and gene expression at single cell resolution

Rachel A. Prescott, Alec P. Pankow, Maren de Vries, Keaton M. Crosse, Roosheel S. Patel, Mark Alu, Cynthia Loomis, Victor Torres, Sergei Koralov, Ellie Ivanova, Meike Dittmann, Brad R. Rosenberg

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2 Scopus citations

Abstract

Background: The airway epithelium is composed of diverse cell types with specialized functions that mediate homeostasis and protect against respiratory pathogens. Human airway epithelial (HAE) cultures at air–liquid interface are a physiologically relevant in vitro model of this heterogeneous tissue and have enabled numerous studies of airway disease. HAE cultures are classically derived from primary epithelial cells, the relatively limited passage capacity of which can limit experimental methods and study designs. BCi-NS1.1, a previously described and widely used basal cell line engineered to express hTERT, exhibits extended passage lifespan while retaining the capacity for differentiation to HAE. However, gene expression and innate immune function in BCi-NS1.1-derived versus primary-derived HAE cultures have not been fully characterized. Methods: BCi-NS1.1-derived HAE cultures (n = 3 independent differentiations) and primary-derived HAE cultures (n = 3 distinct donors) were characterized by immunofluorescence and single cell RNA-Seq (scRNA-Seq). Innate immune functions were evaluated in response to interferon stimulation and to infection with viral and bacterial respiratory pathogens. Results: We confirm at high resolution that BCi-NS1.1- and primary-derived HAE cultures are largely similar in morphology, cell type composition, and overall gene expression patterns. While we observed cell-type specific expression differences of several interferon stimulated genes in BCi-NS1.1-derived HAE cultures, we did not observe significant differences in susceptibility to infection with influenza A virus and Staphylococcus aureus. Conclusions: Taken together, our results further support BCi-NS1.1-derived HAE cultures as a valuable tool for the study of airway infectious disease.

Original languageEnglish
Article number213
JournalRespiratory Research
Volume24
Issue number1
DOIs
StatePublished - Dec 2023

Keywords

  • Airway epithelium
  • Air–liquid interface epithelial culture
  • Innate immunity
  • Respiratory infection
  • Single cell transcriptomics

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