Adenovirus-mediated gene transfer to human lens epithelial cells in organ culture
L M Carrington1, T Southgate, L A Saxby
1Department of Ophthalmology, University of Manchester, Manchester, UK.
Purpose:
To assess the feasibility of using recombinant adenovirus vectors to transduce the human lens epithelial cells (LECs) involved in posterior capsule opacification (PCO).
Setting:
Department of Ophthalmology and Molecular Medicine Unit, University of Manchester, Manchester, United Kingdom.
Methods:
Seventeen human lens capsules were maintained in organ culture to allow LECs to proliferate onto the posterior capsule. Partly covered and completely covered capsules were infected with a recombinant adenovirus vector RAd35, encoding for the marker gene beta-galactosidase at plaque-forming units per milliliter (pfu/mL) ranging from 10(7) to 10(10) for up to 48 hours. Assessment of infection and transduction of the marker gene were achieved by calculating the percentage of cells exhibiting X-gal staining both macroscopically and microscopically.
Results:
Staining appeared to be dependent on virus dose, with most intense staining at doses of 10(8) and 10(9) pfu/mL with decreased staining at higher and lower viral doses. Microscopic assessment demonstrated that all cells expressed beta-galactosidase when infected with 10(9) pfu, 84% at 10(8) pfu, and 45% at 10(7) pfu. At 10(10) pfu, some cytotoxicity was observed.
Conclusions:
These results indicate that recombinant adenoviruses can be used to transfer genes to the LECs involved in PCO. The transfer of cytotoxic genes after cataract surgery may be considered a preventive measure for PCO.
