Modulation of hydrogen peroxide induced injury to corneal endothelium by virus mediated catalase gene transfer.
OBJECTIVE
To examine the effect of catalase gene transfer on survival of corneal endothelial cells (EC) following challenge with hydrogen peroxide (H(2)O(2)) in an ex vivo model of oxidative stress.


METHODS
A recombinant adenovirus vector (AdCL) was used to transfer human catalase cDNA into EC of whole thickness rabbit corneas ex vivo. The resulting catalase protein concentration was measured in corneal lysates by ELISA; catalase functional activity in lysates was determined using a H(2)O(2) activity assay. To examine the morphological effects of catalase gene transfer in modulation of H(2)O(2) induced injury, transduced corneas were maintained in ex vivo culture and challenged with H(2)O(2). Laser scanning confocal microscopy was used to image EC injury. Cell density, cell morphology, and ratios of viable to necrotic cells were determined.


RESULTS
Following incubation with AdCL, catalase expression reached maximum at 5-7 days. Corneas transduced with AdCL showed increased EC cell survival following challenge with H(2)O(2) on day 3 when compared to null vector control or mock infected corneas.


CONCLUSIONS
Ex vivo catalase gene transfer can protect EC from death mediated by H(2)O(2). This gene based approach to the protection of corneal endothelium from oxidative stress may have application in prevention of EC loss in pathological conditions in which H(2)O(2) is involved and in ex vivo donor corneal storage before transplantation.
Department of Pathology, Institute of Ophthalmology, University College London, UK.
T
T
Hudde
R M
RM
Comer
M T
MT
Kinsella
L
L
Buttery
P J
PJ
Luthert
J M
JM
Polak
A J T
AJ
George
D F P
DF
Larkin
D000256:Adenoviridae / Q000235:genetics; D000818:Animals; D002374:Catalase / Q000235:genetics* / Q000378:metabolism; D003315:Cornea / Q000187:drug effects*; D004727:Endothelium / Q000166:cytology; D015870:Gene Expression; D018014:Gene Transfer Techniques; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D018384:Oxidative Stress / Q000235:genetics; D011817:Rabbits; D014161:Transduction, Genetic
D006861:Hydrogen Peroxide; D002374:Catalase