Nitrosative stress and corneal transplant endothelial cell death during acute graft rejection.
BACKGROUND
Nitrosative stress takes place in endothelial cells (EC) during corneal acute graft rejection. The purpose of this study was to evaluate the potential role of peroxynitrite on corneal EC death.


METHODS
The effect of peroxynitrite was evaluated in vivo. Fifty, 250, and 500 microM in 1.5 microL of the natural or denatured peroxynitrite in 50 microM NaOH, 50 microM NaOH alone, or balanced salt solution were injected into the anterior chamber of rat eyes (n=3/group). Corneal toxic signs after injection were assessed by slit-lamp, in vivo confocal imaging, pachymetry, and EC count. The effect of peroxynitrite was also evaluated on nitrotyrosine and leucocyte elastase inhibitor/LDNase II immunohistochemistry. Human corneas were incubated with peroxynitrite and the effect on EC viability was evaluated. A specific inducible nitric oxide synthase inhibitor (iNOS) was administered systemically in rats undergoing allogeneic corneal graft rejection and the effect on EC was evaluated by EC count.


RESULTS
Rat eyes receiving as little as 50 microM peroxynitrite showed a specific dose-dependent toxicity on EC. We observed an intense nitrotyrosine staining of human and rat EC exposed to peroxynitrite associated with leucocyte elastase inhibitor nuclear translocation, a noncaspase dependent apoptosis reaction. Specific inhibition of iNOS generation prevented EC death and enhanced EC survival of the grafted corneas. However, inhibition of iNOS did not have a significant influence on the incidence of graft rejection.


CONCLUSIONS
Nitrosative stress during acute corneal graft rejection in rat eyes induces a noncaspase dependent apoptotic death in EC. Inhibition of nitric oxide production during the corneal graft rejection has protective effects on the corneal EC survival.
Bourges
Jean-Louis
JL
INSERM, UMRS, Team 17, Physiopathology of Ocular Diseases, Therapeutic Innovations, Paris, France.
Torriglia
Alicia
A
Valamanesh
Fatemeh
F
Benezra
David
D
Renard
Gilles
G
Behar-Cohen
Francine F
FF
D000818:Animals; D017209:Apoptosis / Q000187:drug effects / Q000502:physiology*; D002470:Cell Survival / Q000187:drug effects / Q000502:physiology; D016039:Corneal Transplantation / Q000473:pathology* / Q000502:physiology; D004728:Endothelium, Corneal / Q000166:cytology / Q000187:drug effects / Q000473:pathology*; D006084:Graft Rejection / Q000473:pathology* / Q000503:physiopathology; D006801:Humans; D007097:Imines / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000037:antagonists & inhibitors; D015538:Nitrosation; D030421:Peroxynitrous Acid / Q000494:pharmacology / Q000502:physiology*; D051381:Rats; D011914:Rats, Inbred BN; D011917:Rats, Inbred Lew
D007097:Imines; C496401:N-((3-(aminomethyl)phenyl)methyl)ethanimidamide; D030421:Peroxynitrous Acid; D009569:Nitric Oxide; D052247:Nitric Oxide Synthase Type II