Effects of Ischemic Preconditioning and Bevacizumab on Apoptosis and Vascular Permeability Following Retinal Ischemia-Reperfusion Injury

PURPOSE. Using transient ischemia followed by reperfusion (IR) to model ischemic retinal disease, this study compares the effects of ischemic preconditioning (IPC) and therapies targeting vascular endothelial growth factor (VEGF) and tumor necrosis factor (TNF)-alpha on retinal apoptosis, vascular permeability, and mRNA expression. METHODS. Rats were subjected to 30 or 45 minutes of retinal ischemia followed by reperfusion for up to 48 hours. Neurodegeneration was quantified by caspase-3 (DEVDase) activity and by measuring nucleosomal DNA content (cell death ELISA). Vascular leakage was quantified by the Evans Blue dye method. A set of IR-responsive mRNAs was identified by whole-genome microarray and confirmed by RT-PCR analyses. VEGF protein was measured by Western blot analysis. IPC was accomplished with 10 minutes of ischemia 24 hours before IR. VEGF and TNF alpha signaling was inhibited by intravitreal injection of bevacizumab or etanercept, respectively. RESULTS. IR caused significant retinal cell apoptosis and vascular permeability after 4 and 48 hours. Whereas IR decreased VegfA mRNA, VEGF protein was significantly increased. IPC effectively inhibited neurodegeneration, bevacizumab effectively inhibited vascular permeability, and etanercept failed to affect either outcome. IPC significantly altered the IR responses of 15 of 33 IR-responsive mRNAs, whereas bevacizumab had no significant effect on these mRNAs. CONCLUSIONS. IR provides an acute model of ischemic retinopathy that includes neurodegeneration and VEGF-dependent vascular permeability and is amenable to rapid drug therapy testing. The distinct effects of IPC and bevacizumab demonstrate that the apoptotic and vascular responses to IR may be separated and that therapeutics targeting each pathologic endpoint may be warranted in treating ischemic retinal diseases. (Invest Ophthalmol Vis Sci. 2010;51:5920-5933) DOI:10.1167/iovs.10-5264