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Updated: May 29, 2026
A Mouse Model of Retinal Ischemia-Reperfusion Injury Through Elevation of Intraocular Pressure
Published on: July 14, 2016
Neuroprotection from retinal ischemia/reperfusion injury by NOX2 NADPH oxidase deletion
Harumasa Yokota1, Subhadra P Narayanan, Wenbo Zhang
1Vascular Biology Center, Georgia Health Sciences University, Augusta, Georgia 30912-2500, USA.
Purpose:
The aim of this study was to determine whether NOX2, one of the homologs of NADPH oxidase, plays a role in neuronal cell death during retinal ischemia.
Methods:
Ischemia reperfusion (I/R) injury was generated in C57/BL6 and NOX2(-/-) mice by increasing the intraocular pressure (IOP) to 110 mm Hg for 40 minutes followed by reperfusion. Quantitative PCR and Western blot analysis were performed to measure NOX2 expression. Reactive oxygen species (ROS) formation was assessed by dihydroethidium imaging of superoxide formation and Western blot analysis for tyrosine nitration. TUNEL assay was performed to determine cell death at 3 days after I/R. Survival of neurons within the ganglion cell layer (GCL) was assessed at 7 days after I/R by confocal morphometric imaging of retinal wholemounts immunostained with NeuN antibody. Activation of mitogen-activated protein kinases and nuclear factor κB (NF-κΒ) was measured by Western blot analysis.
Results:
NOX2 mRNA and protein and ROS were significantly increased in wild-type I/R retinas. This effect was associated with a 60% decrease in the number of GCL neurons and a 10-fold increase in TUNEL-positive cells compared with the fellow sham control eyes. Phosphorylation of ERK and NF-κB was significantly increased in wild-type I/R retinas. Each of these effects was markedly attenuated in the NOX2(-/-) retina (P < 0.01).
Conclusions:
These data demonstrate that the deletion of NOX2 can reduce I/R-induced cell death and preserve retinal GCL neurons after I/R injury. The neuronal cell injury caused by I/R is associated with the activation of ERK and NF-κB signaling mechanisms.
Insights
NOX2 (NADPH oxidase homolog) contributes to neuronal cell death in retinal ischemia. Deleting NOX2 protects retinal ganglion cells from ischemia-reperfusion injury and reduces cell death.
Area of Science:
- Ophthalmology
- Neuroscience
- Molecular Biology
Background:
- Retinal ischemia-reperfusion (I/R) injury causes significant neuronal cell death.
- The role of NADPH oxidase homolog 2 (NOX2) in this process is not fully understood.
Purpose of the Study:
- To investigate the involvement of NOX2 in neuronal cell death during retinal ischemia.
- To determine if NOX2 deficiency protects against I/R-induced retinal injury.
Main Methods:
- Ischemia-reperfusion injury was induced in wild-type and NOX2 knockout mice.
- NOX2 expression, reactive oxygen species (ROS) formation, and neuronal cell death were assessed.
- Activation of ERK and NF-κB signaling pathways was measured.
Main Results:
- NOX2 expression and ROS levels were significantly elevated in wild-type retinas post-I/R.
- This correlated with a 60% decrease in retinal ganglion cell (GCL) neurons and increased cell death.
- NOX2 deficiency markedly attenuated these effects and preserved GCL neurons.
Conclusions:
- NOX2 plays a critical role in I/R-induced neuronal cell death in the retina.
- NOX2 deletion protects retinal ganglion cells by reducing cell death and preserving neuronal survival.
- ERK and NF-κB signaling are implicated in NOX2-mediated I/R injury.
