One Year Follow Up of Allograft Epithelial Corneal Cell Layers Transplantation in Rabbits

Abstract

Purpose:: To evaluate the efficacy of heterologous limbal cell sheet transplantation at one year of follow-up

Methods:: Limbal stem cell deficiency (LSCD) was induced in 13 New Zealand White rabbits by anterior keratectomy including the limbal area. A small 2.5 mm² limbal sample was obtained from healthy eyes of 6 rabbits. Limbal epithelial cells were cryopreserved. After thawing, cells were cultured on a fibroblast feeder layer grown on autologous clotted platelet poor plasma (PPP), and 21 days later stratified epithelial sheets were ready to be grafted. The heterologous epithelial grafts were sutured over the exposed corneal stroma. Control group included 3 rabbits that underwent anterior keratectomy and grafted with a sutured PPP membrane without cells. The animals received topical ciprofloxacine and dexametasone during 2 weeks after surgery. Clinical, histological and immunofluorescence results were assessed at one year.

Results:: Normal and clear corneas were seen at one year. No signs of limbal stem cells deficiency were observed. A full thickness continuous layer of stratified epithelial cells resembling corneal epithelium was seen in grafted corneas with cells. Neither clinical nor histological evidence of immune reaction was detected. Control animals showed a thin regenerative epithelium and a pronounced fibroblastic proliferation in two of them. Signs of limbal stem cells deficiency were noted in all of them. Expression of cytokeratins 19 and 3/12 on grafted corneas with cells were similar to that found in normal corneas. Control animals showed a stronger K19 staining than normal corneas.

Conclusions:: Allografts of bioengineered limbal epithelium promoted normal corneal re-epithelialization. Heterologous corneal epithelial cell transplantation may be a promising procedure for bilateral ocular surface diseases.

Keywords: transplantation • cornea: stroma and keratocytes • wound healing