Human nasal inferior turbinate-derived airway organoids as an in vitro model of senescence-associated airway epithelial responses

Background: Cellular senescence in the airway epithelium has been implicated in impaired tissue repair, chronic inflammation, and age-related respiratory diseases. However, human upper airway models for examining senescence-associated epithelial responses remain limited.Methods: Human nasal inferior turbinate mucosa was characterized by histological analysis and immunofluorescence staining for ciliated, goblet, and basal cell markers. Epithelial cells were isolated, expanded in 2-dimensional culture, and embedded in Matrigel to generate three-dimensional airway organoids. Organoid growth, lumen formation, and epithelial lineage marker expression were assessed. Mature organoids were exposed to 10 μM etoposide for 48 hours, followed by 24 hours of recovery. Senescence-associated β-galactosidase (SA-β-Gal) staining intensity and the expression of senescence- and senescence-associated secretory phenotype (SASP)-related genes were subsequently evaluated.Results: Human nasal inferior turbinate tissue showed a pseudostratified respiratory epithelium composed of ciliated, goblet, and basal cells. Expanded epithelial cells expressed basal cell markers and generated airway organoids showing progressive growth, lumen formation, and markers of ciliated, goblet, and basal lineages. Etoposide treatment increased SA-β-Gal staining and altered the expression of senescence- and SASP-related genes, including CDKN1A (p21), CDKN2A (p16), TP53 (p53), GLB1, IL6, IL8, and IL1B. These changes were consistent with early senescence-associated responses.Conclusion: Human nasal inferior turbinate–derived airway organoids retained structural and epithelial lineage-associated features of the upper airway and developed an early senescence-associated response after etoposide treatment. This model offers a platform for investigating early senescence-associated responses in the airway epithelium. Longer recovery periods and proliferation assays are needed to determine whether stable senescence can be established.