36 Airway epithelial cell integrity protects from cytotoxicity of Pseudomonas aeruginosa quorum-sensing signals

Objective To investigate the interactions between the P. aeruginosa N-3-oxo-dodecanoyl-L-homoserine lactone (C12) quorum-sensing molecule and human airway epithelial cell gap junctional intercellular communication (GJIC). Methods C12 degradation and its effects on cells were monitored in various human airway epithelial cell models grown under non-polarized and polarized conditions. Its concentration was further monitored in daily tracheal aspirates of colonized intubated patients. Results C12 rapidly altered epithelial integrity and decreased GJIC in non-polarized cells while other quorum-sensing molecules had no effect. C12 increased [Ca 2+ ] i by promoting calcium release from the ER and calcium influx. C12 cytotoxic effects were dependent on [Ca 2+ ] i and could be prevented by inhibitors of Src and Rho-associated protein kinases. In contrast, polarized airway cells grown on Transwell filters were protected from C12. In vivo during colonization of intubated patients, C12 did not accumulate, but paralleled bacterial densities. Interestingly, in vitro C12 degradation, a reaction catalyzed by intracellular paraoxonase 2 (PON2), was impaired in non-polarized cells whereas PON2 expression was increased during epithelial polarization. Finally, in a wound injury model of primary airway epithelial cells grown at the air-liquid interface, repeated exposure of C12 impairs airway epithelial cell repair. Conclusion The cytotoxicity of C12 on non-polarized epithelial cells, combined with its impaired degradation allowing its accumulation, provide an additional pathogenic mechanism for P. aeruginosa infections.