NADPH oxidase 2 plays a role in experimental corneal neovascularization

Abstract

Corneal neovascularization, the growth of new blood vessels in the cornea, is a leading cause of vision impairment after corneal injury. Neovascularization typically occurs in response to corneal injury such as that caused by infection, physical trauma, chemical burns or in the setting of corneal transplant rejection. The NADPH oxidase enzyme complex is involved in cell signalling for wound-healing angiogenesis, but its role in corneal neovascularization has not been studied. We have now analysed the role of the Nox2 isoform of NADPH oxidase in corneal neovascularization in mice following chemical injury. C57BL/6 mice aged 8-14 weeks were cauterized with an applicator coated with 75% silver nitrate and 25% potassium nitrate for 8 s. Neovascularization extending radially from limbal vessels was observed in corneal whole-mounts from cauterized wild type mice and CD31+ vessels were identified in cauterized corneal sections at day 7. In contrast, in Nox2 knockout (Nox2 KO) mice vascular endothelial growth factor-A (Vegf-A), Flt1 mRNA expression, and the extent of corneal neovascularization were all markedly reduced compared with their wild type controls. The accumulation of Iba-1+ microglia and macrophages in the cornea was significantly less in Nox2 KO than in wild type mice. In conclusion, we have demonstrated that Nox2 is implicated in the inflammatory and neovascular response to corneal chemical injury in mice and clearly VEGF is a mediator of this effect. This work raises the possibility that therapies targeting Nox2 may have potential for suppressing corneal neovascularization and inflammation in humans.

Direct Activation of NADPH Oxidase 2 by 2-Deoxyribose-1-Phosphate Triggers Nuclear Factor Kappa B-Dependent Angiogenesis

“…We confirmed that NOX2 is also necessary for the proangiogenic effect of dRP in vivo . This is in line with two recent studies showing a role for this member of the NOX family in the stimulation of angiogenesis in vivo , although it should be emphasized that these reports, in contrast to the present study, did not provide clarity on the underlying molecular mechanisms ( 10 , 70 ).…”

Wound Healing After Alkali Burn Injury of the Cornea Involves Nox4-Type NADPH Oxidase

“…It is worth noting that the nitrate injury is associated with an induction of the proangiogenic VEGF-A but not profibrotic factor TGFβ1. 13 In contrast, NaOH injury stimulates the expression of both VEGF and TGFβ1 in the burnt corneas, consistent with recent findings by Gu et al 28 In this regard, both Nox2 and Nox4 appear to regulate the initial inflammation and neovascular responses in the corneas up to day 7 following chemical insults. The induction of TGFβ1 is ablated in burnt corneas from Nox4 KO mice, which actually show a lower degree of opacity and collagen accumulation in the burnt corneas when compared to the WT.…”

“…Interestingly, the details of these mechanisms of response to injury vary a little between different models of corneal injury. Chan et al 13 used acidic silver nitrate to inflict an injury to the corneal surface, and this results in an upregulation of Nox2 mRNA and a downregulation of the Nox4 gene over 7 days. In that study, Nox2 upregulation was found to be associated with inflammation and neovascularization in the burnt corneas.…”

“… 6 – 9 We and others have also shown key roles for Nox isoforms in retinal angiogenesis and neovascularization 10 – 12 and similar roles for NADPH oxidase 2 (Nox2) in wound healing and neovascularization following chemical burns in the cornea. 13 In this study, we sought to clarify the role of the Nox4 isoform in neovascularization and wound healing after an alkali burn injury, using Nox4 knockout mice. 14 …”

Preparation of arginine–glycine–aspartic acid-modified biopolymeric nanoparticles containing epigalloccatechin-3-gallate for targeting vascular endothelial cells to inhibit corneal neovascularization

“…Next, topical administration of 0.5% Alcaine for local anesthesia was performed, followed by pressing the tip of an applicator containing silver nitrate/potassium (Grafco) to the center of the cornea steadily for 8 seconds. 22 The excess of nitrate was eliminated by washing with 5 mL of saline solution. A burned cornea causing opacity on the eye was observed.…”