Pseudomonas aeruginosa-induced lung epithelial production of IL-8 contributes to PMN sub-mucosa recruitment but not trans-epithelial migration in vitro.

Abstract

Abstract Background Numerous lung associated diseased states such as acute pneumonia and cystic fibrosis (CF) are associated with increased concentrations of IL8 and infiltration of neutrophils (PMNs) in the airspace. IL8 production by lung epithelial cells (ECs) is stimulated by Pseudomonas aeruginosa infection, an opportunistic pathogen involved in acute pneumonia, and lung infection of CF patients. The exact role IL8 plays in migration of circulating PMNs into the infected lung and airspace is still uncertain. Objective Assess the role of IL8 in mediating PMN migration in response to P. aeruginosa infected lung epithelium. Methods CRISPR/Cas9 was used to delete the IL8 gene from human lung ECs, which were grown as polarized monolayers in a co-culture transwell model. Basolateral to apical PMN migration to PAO1 infection was assessed. A novel double transwell model was developed to observe PMN submucosa migration in response to PAO1 infection in the presence and absence of IL8 lung EC production. Results PAO1 induced PMN trans-epithelial (TE) migration was not impacted in the absence of IL8 expression. When PMNs were placed in a separate transwell suspended 5 mm above the basolateral side of the infected epithelium, the IL8 gradient produced was sufficient to cause relevant migration of PMNs to the basolateral side of the infected epithelium. The level of migrated PMNs in the double transwell system with tissues devoid of the IL8 gene was equivalent to the presence of PMNs which responded to un-infected epithelial tissues. Conclusion These findings further support that IL8 is integral in PMN recruitment, and suggests that IL8 is most important towards recruiting PMNs to the submucosa during P. aeruginosa PAO1 infection of lung epithelium.