Wound healing in the rabbit cornea after corneal collagen cross-linking with riboflavin and UVA.
OBJECTIVE
This study was undertaken to investigate the wound healing process of the first 6 weeks after photodynamic cross-linking treatment in the rabbit cornea, using the photosensitizer riboflavin and UVA.


METHODS
After removal of the central epithelium, the right corneas of 8 Chinchilla rabbits were cross-linked with a photosensitizing 0.1% riboflavin solution and UVA light (370 nm; irradiance, 3 mW/cm(2); dose, 5.4 J/cm(2)) for 30 minutes. Two animals were euthanized 3 days, 7 days, 4 weeks, and 6 weeks postoperatively. The corneas of the enucleated eyes were evaluated using 4-microm light microscopic sections with routine stains and avidin-biotin complex immunostaining with anti-alpha-smooth muscle actin.


RESULTS
By day 3 after treatment, complete apoptotic damage and loss of the endothelial cells and the stromal keratocytes were found in the irradiated area through the entire thickness of the stroma. There was marked stromal edema (850 +/- 66 vs. 332 +/- 43 microm in the untreated controls; P < 0.01). The epithelium was already closed again. At the margins of the lesion, there was a mild inflammatory reaction with scattered macrophages, lymphocytes, and neutrophils. By day 7, the endothelium was already intact again, and keratocyte repopulation of the posterior stroma was noted. By week 4, the keratocyte repopulation of the anterior stroma was observed with some acellular areas between. By week 6, the cytoarchitecture of the cornea seemed normal again. By weeks 4 and 6, alpha-actin-positive keratocytes were identified, especially in the periphery of the irradiated area.


CONCLUSIONS
After riboflavin/UVA cross-linking of rabbit cornea, a complete cell loss occurs in the irradiation area with an irradiance of 3 mW/cm(2). The cytotoxic damage is repaired by repopulation after approximately 4-6 weeks. A combination of cross-linking with other procedures such as the implantation of intracorneal rings should be performed only after a sufficient time interval of approximately 2 months, allowing cellular regeneration.
Wollensak
Gregor
G
Moscow Helmholtz Research Institute of Eye Diseases, Moscow, Russia. gwollens@hotmail.com
Iomdina
Elena
E
Dittert
Dag-Daniel
DD
Herbst
Hermann
H
D000199:Actins / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D003094:Collagen / Q000378:metabolism* / Q000528:radiation effects; D003315:Cornea / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D003319:Corneal Stroma / Q000473:pathology; D005260:Female; D005347:Fibroblasts / Q000473:pathology; D007124:Immunoenzyme Techniques; D010778:Photochemotherapy; D017319:Photosensitizing Agents / Q000494:pharmacology*; D011817:Rabbits; D012256:Riboflavin / Q000494:pharmacology*; D014466:Ultraviolet Rays; D014945:Wound Healing
D000199:Actins; D017319:Photosensitizing Agents; D003094:Collagen; D012256:Riboflavin