The role of ciliary function in airway epithelial defense against Pseudomonas aeruginosa.

Respiratory cilia play a crucial role in clearing pathogens from the airway, and understanding how Pseudomonas aeruginosa (PA) impairs their function is essential for developing targeted therapies to enhance airway epithelial defense and hamper bacterial invasion. Thus, we investigated the mechanisms by which PA impairs respiratory cilia function and identified pharmacological interventions to restore cilia motility. We used mucociliary differentiated human airway organoids expressing motile cilia on the apical side, to model the initial stage of PA infection. We show that co-culturing of organoids with PA down-regulates the expression of genes associated with cilia formation, structure and function. Electron microscopy confirmed ciliary structural damage and membrane disruption in infected organoids and slowed ciliary beating frequency as quantified by slow-motion video recording. Reduction of cilia function resulted in increased bacterial cell invasion, and we identified the bacterial toxin Pyocyanin as one causative cilia damaging factor. We could further demonstrate, that the selective phosphodiesterase-4 inhibitor Roflumilast can maintain ciliary function and beating frequency upon infection of organoids with PA, thereby reducing bacterial invasion of epithelia in our model system. Our findings emphasize the critical role of motile cilia in innate host defense against PA infection, implying that treatment strategies aimed at restoring ciliary function can enhance epithelial robustness during bacterial invasion. Our human organoid infection model enables the evaluation of cellular and functional responses of the airway epithelium to respiratory pathogens and to test potential therapeutic strategies aimed at restoring ciliary function and improving infection control.