Bacterial keratitis is a leading cause of blindness and visual impairment in the developing and industrialized world. Trauma, ocular surface disorders, inappropriate contact lens wear and suture abscess post corneal surgery are among the causative factors that lead to breakdown of corneal defense mechanisms and facilitate access of bacteria and bacterial products into the deeper epithelial and stromal layers. Bacterial products such as lipopolysaccharide (LPS), peptidoglycan (PGN) and unmethylated bacterial DNA (CpG DNA) then activate specific Toll-like receptors (TLR) on resident corneal cells. TLRs initiate a cascade of intracellular signaling events resulting in NFkB translocation into the nucleus, and transcription of proinflammatory and chemotactic cytokines. LPS from most Gram negative bacteria activate TLR4, PGN from gram positive bacterial activate TLR2, and CpG DNA activates TLR9. Activation of specific TLRs induces distinct patterns of cytokine production, despite many shared characteristics of TLR signaling. Experiments outlined in this proposal will utilize TLR and MyD88 deficient mice to identify shared and distinct effects of LPS, PGN and CpG DNA on on cytokine and chemokine production by resident corneal epithelial cells and keratocytes, on expression of vascular cell adhesion molecules on limbal vessels and on infiltrating neutrophils and macrophages. Proposed experiments will utilize bone marrow chimeras to delineate the role of TLRs on infiltrating cells from that of resident corneal cells in mediating bacterial keratitis, which will be measured quantitatively by in vivo scanning confocal microscopy. We anticipate that results of these studies will elucidate how signals from TLRs and cytokines integrate in regulating the development and severity of bacterial keratitis, and will lead to the rational design of novel therapeutic agents.
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