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Harumasa Yokota, Subhadra P. Narayanan, Wenbo Zhang, Zhimin Xu, Tahira Lemtalsi, Taiji Nagaoka, Akitoshi Yoshida, Robert W. Caldwell, Ruth B. Caldwell; Deletion of NADPH Oxidase 2 Prevents Ganglion Cell Loss in the Retina after Ischemia Reperfusion Injury. Invest. Ophthalmol. Vis. Sci. 2011;52(14):6058.
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© ARVO (1962-2015); The Authors (2016-present)
Retinal ischemia causes not only neovascularization but also loss of ganglion cells. The aim of this study was to determine whether NOX2, one of the homologues of NADPH oxidase, plays a role in ganglion cell death during retinal ischemia.
Ischemia Reperfusion (I/R) injury was generated in wild type or NOX2 (-/-) mice by increasing the intraocular pressure to 110 mmHg for 40 minutes followed by reperfusion. The contralateral eyes were prepared as controls. The mice were sacrificed at 3h, 6h, 12h, 24h, 48h and 7 days after I/R. The whole mounted retinas were immunostained with NeuN antibody to detect ganglion cells 7 days after I/R. Ten non-overlapping confocal images were randomly taken around the mid-periphery at the ganglion cell layer level in each retina, and the ratio of the number of NeuN+ cells relative to that of the fellow eye was calculated. Western blotting was performed to examine the expression of molecules that initiate or promote apoptosis after I/R.
The expression of NOX2 was increased at 3h, reached maximum level at 6h and was almost normalized at 48h after I/R. While I/R resulted in a 60% reduction in the number of ganglion cells compared to the fellow eyes in WT, only 16% reduction in NeuN positive cells was observed in NOX2-/- retina showing a significant decrease (p<0.01) in retinal ganglion cell apoptosis due to NOX2 deletion. Phosphorylated NFkB was significantly increased after I/R in WT retina. In NOX2 (-/-) mice, the increase in p-NFkB was attenuated. In contrast indicators of the mitochondrial permeability-associated apoptotic pathway, Bcl-xL, Bcl-2 and cytochrome C, were unchanged after I/R in WT and NOX2-/- retinal samples.
Our data demonstrate that deletion of NOX2 can preserve retinal ganglion cells after I/R injury. The ganglion cell apoptosis caused by I/R is shown to be driven by NFkB dependent signaling mechanism, and the mitochondrial permeability-associated pathway is not involved in this process. These data suggest that inactivation of NOX2 can offer new therapeutic strategies for the prevention of ganglion cell loss as well as retinal inflammation in diabetic retinopathy.
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