Mechanism of pyocyanin- and 1-hydroxyphenazine-induced lung neutrophilia in sheep airways

Pyocyanin (Pyo)and 1-hydroxyphenazine (1-HP) are extracellular products ofPseudomonas aeruginosa. To testwhether these products were capable of producing an inflammatoryresponse in the airways, combinations of Pyo and 1-HP at concentrationsof 10⁴ and10⁵ M were instilled intosheep airways, and indexes of inflammation were assessed bybronchoalveolar lavage (BAL) 24 h later. Challenge with the phenazinescaused a significant dose-dependent increase in the number of cells andneutrophils recovered by BAL. Control challenges produced no suchchanges. The lung neutrophilia was accompanied by an increasedconcentration of albumin in BAL. The increases in BAL neutrophils andalbumin could be blocked by treating the sheep with the 5-lipoxygenaseinhibitor zileuton. Neither 1-HP nor Pyo was chemotactic to neutrophilswhen tested in vitro, but when alveolar macrophages (AM) were culturedin vitro in the presence of both Pyo and 1-HP (1 µM), thesupernatants caused neutrophil chemotaxis. Analysis of AM culturesupernatants incubated with the combination of pigments showedsignificant increases in leukotrieneB₄ and interleukin-8, and blockingthese mediators separately or together reduced AM supernatant-inducedneutrophil chemotaxis. We conclude that local instillation of Pyo and1-HP can initiate an inflammatory response in the airways of sheep invivo. This effect can be explained, in part, by the release ofchemotactic factors produced by AM.