Transplantation of Autologous Cultivated Conjunctival Epithelium on Amniotic Membrane for Total Limbal Stem Cell Deficiency in a Rabbit Model

Abstract

Abstract: : Purpose: To evaluate the feasibility of autologous cultivated conjunctival epithelium on amniotic membrane for total limbal stem cell deficiency.. Methods: Lamellar keratectomy to create total limbal stem cell deficiency was performed in the right eye of adult New Zealand white rabbit. Autologous conjunctival epithelium, obtained from the left eye, was cultivated by co–culture with 3T3 fibroblasts on a denuded human amniotic membrane for 3 to 4 weeks. Conjunctival epithelial sheets were examined histologically and immunohistochemically. At 3 to 4 weeks after the lamellar keratectomy, the conjunctivalized corneal surfaces were surgically removed and the autologous cultivated conjunctival epithelial sheets were transplanted (Conj–AM group, n=10). The no transplantation (n=6) and AM (rabbits with an amniotic membrane transplantation only, n=10) groups were the controls. Each group, observed for 2 months after surgery, was evaluated by the scoring systems of neovascularization and corneal opacity. Results: The cultivated conjunctival epithelial sheets formed 3–5 layers on denuded amniotic membrane in HE staining. Transmission electron microscopic examination revealed hemidesmosome, desmosome, and microvilli on the cultivated conjunctival epithelium. Averaged scores of corneal neovascularization and corneal opacity in the Conj–AM group were significantly suppressed as compared with those in the AM and no transplantation groups 2 months after transplantation. Immunohistochemical study showed cytokeratin 3 and 12 expressions on the conjunctival epithelial sheets in the Conj–AM group. Conclusions: Our findings indicate that transplantation of autologous cultivated conjunctival epithelial sheet can be an effective strategy for total limbal stem cell deficiency in a rabbit model.

Keywords: cornea: epithelium • conjunctiva • transplantation