Caffeine citrate increases ciliary beat frequency in human respiratory epithelial cells

BACKGROUND: Caffeine citrate is the gold-standard pharmacological treatment for apnea of prematurity, improving survival and long-term respiratory outcomes in preterm infants. Beyond its established central nervous system effects, emerging evidence suggests broader benefits in reducing extubation failure, lowering bronchopulmonary dysplasia rates, and decreasing the need for mechanical ventilation-but its direct actions on the respiratory epithelium remain largely unknown. Caffeine is a methylxanthine with multiple molecular targets, including adenosine receptor antagonism, phosphodiesterase inhibition, and stimulation of calcium (Ca²⁺) release from intracellular stores-mechanisms also linked to the regulation of ciliary beat frequency (CBF), a critical determinant of mucociliary clearance. Since impaired CBF is a hallmark of several chronic airway diseases, understanding whether caffeine citrate can directly modulate this function could broaden its therapeutic relevance. We hypothesized that caffeine citrate directly enhances CBF in human respiratory epithelial cells (hRECs), thereby supporting airway defense mechanisms in health and disease. RESULTS: ) in both healthy and CF-derived cells, indicating a robust and reproducible stimulatory effect. Mechanistic experiments identified ryanodine receptor (RYR)-mediated Ca²⁺ efflux from intracellular stores as the key pathway, as pharmacological blockade of RYR abolished the caffeine-induced CBF enhancement. CONCLUSIONS: This study reveals a novel pharmacological effect of caffeine citrate-direct stimulation of CBF in primary hRECs through RYR-dependent Ca²⁺ signaling. By improving mucociliary clearance, caffeine citrate may offer peripheral airway benefits in addition to its central actions for apnea prevention. Given its favorable pharmacokinetic profile, long half-life, and good tolerability compared to other methylxanthines, caffeine citrate warrants further evaluation as a dual-action therapy for respiratory diseases with compromised clearance, including CF and bronchopulmonary dysplasia. These findings expand the mechanistic understanding of caffeine's actions and highlight new translational opportunities in respiratory care.