Identification of Therapeutic Targets of Inflammatory Monocyte Recruitment to Modulate the Allogeneic Injury to Donor Cornea

Abstract

Citation: Lapp T, Zaher SS, Haas CT, et al. Identification of therapeutic targets of inflammatory monocyte recruitment to modulate the allogeneic injury to donor cornea. Invest Ophthalmol Vis Sci. 2015;56:7250-7259. DOI:10.1167/iovs.15-16941 PURPOSE. We sought to test the hypothesis that monocytes contribute to the immunopathogenesis of corneal allograft rejection and identify therapeutic targets to inhibit monocyte recruitment. METHODS.Monocytes and proinflammatory mediators within anterior chamber samples during corneal graft rejection were quantified by flow cytometry and multiplex protein assays. Lipopolysaccharide or IFN-c stimulation of monocyte-derived macrophages (MDMs) was used to generate inflammatory conditioned media (CoM). Corneal endothelial viability was tested by nuclear counting, connexin 43, and propidium iodide staining. Chemokine and chemokine receptor expression in monocytes and MDMs was assessed in microarray transcriptomic data. The role of chemokine pathways in monocyte migration across microvascular endothelium was tested in vitro by chemokine depletion or chemokine receptor inhibitors.RESULTS. Inflammatory monocytes were significantly enriched in anterior chamber samples within 1 week of the onset of symptoms of corneal graft rejection. The MDM inflammatory CoM was cytopathic to transformed human corneal endothelia. This effect was also evident in endothelium of excised human cornea and increased in the presence of monocytes. Gene expression microarrays identified monocyte chemokine receptors and cognate chemokines in MDM inflammatory responses, which were also enriched in anterior chamber samples. Depletion of selected chemokines in MDM inflammatory CoM had no effect on monocyte transmigration across an endothelial blood-eye barrier, but selective chemokine receptor inhibition reduced monocyte recruitment significantly.CONCLUSIONS. We propose a role for inflammatory monocytes in endothelial cytotoxicity in corneal graft rejection. Therefore, targeting monocyte recruitment offers a putative novel strategy to reduce donor endothelial cell injury in survival of human corneal allografts.

Elevated Cytokine Levels in the Aqueous Humor of Eyes With Bullous Keratopathy and Low Endothelial Cell Density

“…Eom et al 42 reported the elevation of IL-1 and IFN-c levels after argon LI induced apoptosis of endothelial cells in an animal model. We postulate that these results 17,41,42 involved a rapid decrease in the ECD in response to proinflammatory stimuli. In the present study, it is noteworthy that AqH cytokine levels were elevated, although we collected all samples during a clinically noninflammatory period.…”

“…39,40 It is probable that AqH cytokine levels are elevated in eyes with active inflammation, leading to ECD loss. Lapp et al 41 reported that proinflammatory mediators from monocytes are sufficient to induce the cell death of a human corneal endothelial cell line in vitro and death of primary human corneal endothelial cells in corneal buttons ex vivo. Eom et al 42 reported the elevation of IL-1 and IFN-c levels after argon LI induced apoptosis of endothelial cells in an animal model.…”

Iris Damage Is Associated With Elevated Cytokine Levels in Aqueous Humor

“…Previous laboratory studies have shown that inflammatory cytokines induce apoptosis in corneal endothelial cells. 17,35,36 We recently reported that severe preexisting iris damage was a risk factor for graft failure and rapid ECD loss after DSAEK. 12 Furthermore, we found that the inflammatory cytokine levels in AqH increased in eyes with BK and reduced ECD.…”

Longitudinal Changes to Tight Junction Expression and Endothelial Cell Integrity in a Mouse Model of Sterile Corneal Inflammation

“…33 A recent study on the pathologic capacity of activated monocyte-derived cells during allograft rejection provides evidence that macrophage-derived cytokines, including interferon-c, can directly induce corneal endothelial cell death in vitro. 34 Thus, it is possible that the persistence of activated leukocytes in our model of CpG-ODN-induced corneal inflammation contributes to the observed decline in endothelial cell density and the increased corneal thickness at 4 weeks post treatment.…”