Abstract
Purpose::
Staphylococcus aureus is a common etiological agent of microbial keratitis, linked with non-surgical traumas to the eye and contact lens wear. These infections typically begin with colonization of the corneal epithelium and progression into underlying tissue. In vitro and in situ models suggest that adherence to the corneal epithelial is mediated by fibronectin- and collagen-binding proteins expressed on the bacterial surface. The ensuing necrosis is caused by lytic bacterial toxins and the host immune response. Interestingly, some lytic toxins can also modulate cellular behavior at sublethal concentrations by altering receptor processing and intracellular signaling events. Most of these toxins are modulated by global regulators Agr and Sar.Innate immunity at the ocular surface requires an intact and responsive corneal epithelium. This tissue acts as a barrier to infectious agents and secrets anti-microbial peptides, cytokines and chemokines following exposure to pathogenic organisms. Little is known regarding the effects of S. aureus toxins on corneal epithelium. The objective of this study is to determine if sublethal concentrations of Agr/Sar-regulated toxins derange the response of corneal epithelial cells to the presence of adherent S. aureus.
Methods::
S. aureus RN6390 and an isogenic Agr/Sar mutant ALC135 were incubated with confluent monolayers of human corneal epithelial cells (primary and Araki-Saki) and corneal/limbal epithelial cells (Rheinwald) in serum-free defined keratinocyte medium for 1 to 8 hrs. Co-cultures were assessed periodically for epithelial cell viability, hemolytic activity, and cytokine production.
Results::
Epithelial monolayers maintained ≥ 90% viability over 6hrs of co-culture with bacteria if the initial MOIs were ≤ 20. Within 4hrs, detected levels of hemolytic toxins were observed in RN6390 cultures. No hemolytic activity was found after 8hrs in the ALC135 cultures. Primary corneal epithelium secreted IL6, IL8, and IFNγ within 4hr of co-culture with S. aureus. Similar observations were made with transformed corneal epithelium and corneal-limbal epithelium cell lines. Comparable levels of cytokines and chemokines were noted in toxigenic and non-toxigenic S. aureus infections.
Conclusions::
Primary human corneal epithelial cells secret IL6, IL8, and IFNγ in response to adherent S. aureus. Sublethal concentrations of toxins do not appear to influence this immune response. It is unclear what role IFNγ originating from corneal epithelium plays in host immunity to S. aureus.
Keywords: Staphylococcus • keratitis • microbial pathogenesis: experimental studies