Comparative Outcomes and Immune Mechanisms in Murine Endothelial Versus Penetrating Keratoplasty.

Abstract

PURPOSE: To compare graft survival and alloimmune responses in murine endothelial keratoplasty (EK) versus penetrating keratoplasty (PK) and to elucidate the immunological mechanisms that underlie the differential graft outcomes. METHODS: Allogeneic EK and PK were performed in BALB/c recipient mice using fully disparate C57BL/6 donors; syngeneic EK recipients served as controls. Graft clarity was monitored over 16 weeks by slitlamp biomicroscopy and scored using standardized opacity grading. Anterior segment optical coherence tomography (AS-OCT) was used to measure central corneal thickness. Graft survival was assessed using Kaplan-Meier analysis. Immunohistochemistry and confocal microscopy were performed to evaluate corneal endothelial cell (CEnC) integrity through ZO-1 staining. T-cell-mediated alloimmunity was assessed using intracellular IFN-&#x3b3; staining (flow cytometry) and ELISPOT assays targeting both direct and indirect antigen presentation pathways. RESULTS: PK allografts exhibited significantly higher corneal opacity and lower survival (50%) than allogeneic EK grafts (71.4%, P < 0.0001). AS-OCT showed that corneal edema was highest in rejected PK grafts at 4 weeks and in rejected EK grafts at 16 weeks, with EK displaying a more gradual increase in thickness. Flow cytometry revealed significantly greater frequencies of IFN-&#x3b3; + CD4 + T cells in PK recipients compared with EK recipients ( P < 0.001). ELISPOT assays demonstrated a more robust Th1 response in PK through both the direct and indirect sensitization pathways. Corneal endothelial cell (CEnC) density was significantly reduced in rejected EK and PK grafts compared with their respective accepted counterparts ( P < 0.01), whereas CEnC density was comparable between accepted EK and PK grafts. CONCLUSIONS: EK grafts exhibit higher graft survival rates and significantly reduced activation of host T-cell responses compared with PK grafts, which may be attributed to lower frequencies of graft-borne antigen presenting cells, thus resulting in a milder Th1-mediated immune response.