Corneal Endothelial Cell Integrity in Precut Human Donor Corneas Enhanced by Autocrine Vasoactive Intestinal Peptide.

Abstract

Purpose:To demonstrate that vasoactive intestinal peptide (VIP), a corneal endothelial (CE) cell autocrine factor, maintains the integrity of corneal endothelium in human donor corneoscleral explants precut for endothelial keratoplasty.Methods:Twelve paired human donor corneoscleral explants used as control versus VIP-treated explants (10 nM, 30 minutes, 37°C) were shipped (4°C) to the Lions Eye Institute for Transplantation and Research for precutting (Moria CBM-ALTK Keratome), shipped back to the laboratory, and cultured in ciliary neurotrophic factor (CNTF, 0.83 nM, 37°C, 24 hours). Trephined endothelial discs (8–8.5 mm) were analyzed for differentiation markers (N-cadherin, CNTF receptor α subunit [CNTFRα], and connexin 43) by Western blot after a quarter of the discs from 4 paired explants were cut away and stained with alizarin red S for microscopic damage analysis. Two additional paired explants (6 days in culture) were stained for panoramic view of central CE damage.Results:VIP treatment increased N-cadherin and CNTFRα levels (mean ± SEM) to 1.38 ± 0.11-fold (P = 0.003) and 1.46 ± 0.22-fold (P = 0.03) of paired controls, respectively, whereas CE cell CNTF responsiveness in upregulation of connexin 43 increased to 2.02 ± 0.5 (mean ± SEM)-fold of the controls (P = 0.04). CE damage decreased from (mean ± SEM) 10.0% ± 1.2% to 1.6% ± 0.3% (P < 0.0001) and 9.1% ± 1.1% to 2.4% ± 1.0% (P = 0.0006). After 6 days in culture, the damage in whole CE discs decreased from 20.0% (control) to 5.5% (VIP treated).Conclusions:VIP treatment before precut enhanced the preservation of corneal endothelium.