Nox2 Deficiency Enhances Revascularization in Ischemic Retinopathy

Abstract

Purpose: : Oxidative stress plays a critical role in retinal vascular diseases, including ischemic retinopathy. NOX2 NADPH oxidase is a major source of reactive oxygen species in the vasculature. We have found that NOX2 is upregulated in the retina during oxygen induced retinopathy (OIR). The goal of this study was to determine the potential role of NOX2 in physiological revascularization in OIR.

Methods: : OIR model was induced by exposing C57BL/6J wild type (WT) or NOX2 knockout (NOX2-ko) mice to 70% oxygen from postnatal day (P)7 to P12 followed by return to room air. The retinal vasculature was examined in retinal wholemounts after labeling with isolectin-B4 in combination with markers for specific cell types. Vascular dropout area was measured on digital images using NIH imageJ software.

Results: : At P12, there was no difference in extent of vaso-obliteration between the NOX2-ko and WT mice (32.2% vs. 32.7%). At P17 the NOX2-ko mice showed significantly greater revascularization than WT. The capillary-free area in NOX2-ko retinas was decreased by 26.1% as compared with WT (P<0.001). In addition, the NOX2-ko retinas at both P12 and P17 showed a dramatic increase in a population of round, lectin-positive cells that were not incorporated into the vasculature, but were closely associated with the sprouting filipodia of endothelial tip cells. Studies using specific cell markers demonstrated that this cell subset was not endothelial progenitor cells since they were negative for CD34. In contrast, they were positive for markers of leukocytes (CD45) and monocyte/microgila (Iba1). Given that the morphology of these cells was different from that of the resident microglia, these results indicate that a subset of pro-angiogenic retinal monocytes/microglia is increased by NOX2 deficiency.

Conclusions: : Lack of NOX2 NADPH oxidase enhances recruitment of a subset of pro-angiogenic monocyte/microglial cells and increases retinal revascularization in ischemic retinopathy. This result indicates NOX2 is a potential candidate for improving revascularization in ischemic retinopathy.

Keywords: retinal neovascularization • signal transduction • microglia