Apoptosis in the endothelium of human corneas for transplantation

PURPOSE: To determine whether endothelial cell loss of human corneas stored in organ culture before transplantation is due to apoptosis. METHODS: The corneal endothelium of human corneas, stored in organ culture at 34 degrees C for varying periods of time, were analyzed for the presence of apoptotic cells using the TdT-mediated dUTP nick-end labeling (TUNEL) technique. Corneal endothelial cell apoptosis was confirmed by Hoechst staining and immunolabeling with anti-caspase 3 active antibody. RESULTS: Apoptotic cells were identified in the corneal endothelium of human organ cultured corneas: their number and distribution demonstrated a close correlation with corneal folding and overall quality of the corneal endothelium. TUNEL-positive labeling of cells was confirmed as apoptotic by the presence of morphologic nuclear alterations identified by Hoechst staining and the presence of immunostaining for caspase-3 activity. Corneal endothelial cell apoptosis was independent of cause of donor death, death to enucleation time, and death to culture times. CONCLUSIONS: Corneal endothelial cell apoptosis appears to determine the suitability of a cornea for transplantation.