Gene transfer to ovine corneal endothelium

Purpose: Modification of a donor cornea by gene therapy has potential to mo dulate irreversible rejection, the major cause of corneal graft failure. Th e sheep is a useful model for the human in this respect, as ovine endotheli al cells are amitotic. The aim of the study was to investigate the ability of various non-viral and viral agents to transfer a reporter gene to ovine corneal endothelium. Methods: The non-viral agents Transfectin-10, Transfectin-20, Transfectin-5 0, SuperFect, Effectene and CLONfectin were used to deliver the reporter ge ne, Escherichia coli lacZ, to ovine corneal endothelium in vitro. A Herpes simplex virus-1 and an adenoviral vector each encoding E. coli lacZ were si milarly tested. Infected corneas were organ-cultured for up to 7 days in vi tro to allow transfection efficiency, duration of gene expression and toxic ity attributable to each vector to be compared. Results: Scattered single or clusters of endothelial cells expressing the r eporter gene were observed after transfection with CLONfectin, Transfectin- 10, Transfectin-20 and Transfectin-50. SuperFect and Effectene were virtual ly in-effective. At best, the absolute number of infected cells per endothe lial monolayer after 3 or 7 days of organ culture was estimated as < 0.01%. The Herpes simplex virus-1 vector also failed to transduce ovine corneal e ndothelium efficiently. In contrast, transfection rates of up to 70% of end othelial cells were observed with the adenoviral vector. Conclusion: Non-viral vectors and Herpes simplex virus-1 are unlikely to be suitable for gene therapy of corneal endothelium, because the efficiency o f transfection is low compared with the rates achieved with adenoviral vect ors.