Role of Neutrophils, MyD88-Mediated Neutrophil Recruitment, and Complement in Antibody-Mediated Defense against Pseudomonas aeruginosa Keratitis

Corneal infection due to Pseudomonas aeruginosa can result in devastating tissue destruction within a short period after the initiation of infection, leading to corneal scarification, opacification, and loss of vision.1–4 Numerous patients are at risk for such infections, notably those using extended-wear contact lenses,2,3,5,6 those undergoing various eye surgeries or treatments such as orthokeratology or keratoplasty,7–9 and individuals suffering eye trauma or ocular surface diseases.10 Therapy for such infections must be rapidly instituted after the onset of symptoms to minimize corneal damage, but currently recommended treatments are arduous, consisting of topical antibiotic applications every 5 minutes for 1 hour followed by additional drops every 15 minutes for 24 to 48 hours. More severe infections may require antibiotic injection into the eye itself. We have recently shown that a fully human monoclonal antibody (MAb F429) specific to the P. aeruginosa alginate surface polysaccharide was highly effective at reducing the infectious burden and corneal pathology in mice treated either prophylactically or therapeutically with the MAb.11 In vitro studies showed that MAb required both phagocytic polymorphonuclear neutrophils (PMN) and complement to mediate bacterial killing, but whether there was a similar requirement for these co-factors in vivo was not determined. Prior work has indicated that human MAbs to P. aeruginosa lipopolysaccharide had different requirements for complement in mediating opsonic killing and protecting against systemic infection, with IgM having an absolute requirement for complement while IgG and IgA did not.12 These studies did not address the role of PMN or other cellular factors. As the PMN-dominated inflammation in the cornea drives the pathology leading to loss of vision from microbial keratitis,13–17 we were interested in determining the contributions of locally-recruited PMN and complement to MAb F429-mediated reductions in bacterial levels and pathology during P. aeruginosa corneal infections. In particular, there was concern that inflammation-associated pathology might be exacerbated by MAb activation of complement and generation of complement split products chemotactic for PMN. Alternately, these may be critical co-factors for the beneficial effects of MAb F429 therapy. Thus, establishing the proper balance between antibody, PMN, and complement-mediated bacterial clearance while limiting inflammatory pathology is essential for optimizing outcomes in treatment of P. aeruginosa keratitis. In this study, we evaluated the activity of MAb F429 in reducing bacterial burdens and corneal pathology in MyD88 knock-out (KO) mice unable to recruit PMNs to tissue, systemically neutropenic mice, or complement depleted mice, after infection with two different P. aeruginosa strains, one representative of the ExoS+ invasive strains and the other of the cytotoxic ExoU+ strains.