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Thomas A. Fuchsluger, Daniel R. Saban, Andre Okanobo, Reza Dana; Anti-Apoptotic Gene Therapy to Promote Survival of Allogeneic Corneal Epithelial Grafts. Invest. Ophthalmol. Vis. Sci. 2011;52(14):2057.
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To study whether anti-apoptotic gene therapy to allogeneic corneal epithelial sheets in mice promotes graft survival. Engraftment of cultured allogeneic epithelium currently is a viable modality for patients bilaterally blind due to bilateral limbal stem cell deficiencies, e.g. after chemical burn. Despite full corticosteroid intervention, graft failure rate entailing apoptosis of donor cells is >50%. Gene transfer to cultured epithelial cell sheets in vitro before transplantation could improve the outcome in this most demanding ocular surface pathology .
Epithelial denuded syngeneic Balb/C corneal buttons were reconstituted with allo- or syngeneic epithelium, transplanted and compared to epithelial grafts transduced with the anti-apoptotic gene p35 (total n=40). Feasibility of gene transfer was determined by studying kinetics of IZsGreen expression. Epithelial integrity and graft survival were assessed by fluoresceine staining and corneal opacity. Allo-response was evaluated by delayed type hypersensitivity (DTH).
Transfer of IZsGreen in vitro was successfully achieved obtaining high expression rates. Denuded corneas or those reconstituted with allogeneic epithelium showed a significantly higher opacity rate and considerably prolonged periods for epithelial regeneration relative to syngeneic controls. Surprisingly, gene-therapeutically modulated allogeneic epithelial grafts showed a significantly lower opacity rate and significantly expedited restoration of the epithelial barrier function compared to allo-untreated epithelial grafts (p<0.01, respectively). In addition, DTH analysis consistently showed significantly reduced T-cell response in the treated group (p<0.01).
We herein established a novel model of allogeneic epithelial graft rejection. We were able to demonstrate that gene therapeutic modification of grafted cell sheets may be a viable option to increase allogeneneic epithelial graft survival. This sets the stage to further study gene therapeutic interventions of epithelial allo-grafts, and its impact on graft survival and immune response.
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