Corneal Endothelial Cell Integrity in Precut Human Donor Corneas Enhanced by Autocrine Vasoactive Intestinal Peptide.
OBJECTIVE
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.
Departments of *Ophthalmology and Visual Sciences; †Physiology, University of Maryland, Baltimore, MD; and ‡Lions Eye Institute for Transplantation and Research, Tampa, FL.
Shay-Whey M
SM
Koh
Timothy
T
Coll
Dante
D
Gloria
Nicholas
N
Sprehe
D000328:Adult; D000368:Aged; D015703:Antigens, CD / Q000378:metabolism; D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism; D002470:Cell Survival / Q000502:physiology*; D020934:Ciliary Neurotrophic Factor / Q000494:pharmacology; D053739:Ciliary Neurotrophic Factor Receptor alpha Subunit / Q000378:metabolism; D018031:Connexin 43 / Q000378:metabolism; D057111:Descemet Stripping Endothelial Keratoplasty; D004728:Endothelium, Corneal / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D005260:Female; D006801:Humans; D008297:Male; D008875:Middle Aged; D018696:Neuroprotective Agents / Q000627:therapeutic use*; D009924:Organ Culture Techniques; D009926:Organ Preservation; D019188:Organ Preservation Solutions; D014019:Tissue Donors; D020858:Tissue and Organ Harvesting; D014660:Vasoactive Intestinal Peptide / Q000627:therapeutic use*
D015703:Antigens, CD; C050361:CDH2 protein, human; D015820:Cadherins; D020934:Ciliary Neurotrophic Factor; D053739:Ciliary Neurotrophic Factor Receptor alpha Subunit; D018031:Connexin 43; C500216:GJA1 protein, human; D018696:Neuroprotective Agents; D019188:Organ Preservation Solutions; D014660:Vasoactive Intestinal Peptide