Role of NADPH Oxidase-4 in Human Endothelial Progenitor Cells

Abstract

Introduction: Endothelial progenitor cells (EPCs) display a unique ability to promote angiogenesis and restore endothelial function in injured blood vessels. NADPH oxidase 4 (NOX4)-derived hydrogen peroxide (H2O2) serves as a signaling molecule and promotes endothelial cell proliferation and migration as well as protecting against cell death. However, the role of NOX4 in EPC function is not completely understood.Methods: EPCs were isolated from human saphenous vein and mammary artery discarded during bypass surgery. NOX4 gene and protein expression in EPCs were measured by real time-PCR and Western blot analysis respectively. NOX4 gene expression was inhibited using an adenoviral vector expressing human NOX4 shRNA (Ad-NOX4i). H2O2 production was measured by Amplex red assay. EPC migration was evaluated using a transwell migration assay. EPC proliferation and viability were measured using trypan blue counts.Results: Inhibition of NOX4 using Ad-NOX4i reduced Nox4 gene and protein expression as well as H2O2 formation in EPCs. Inhibition of NOX4-derived H2O2 decreased both proliferation and migration of EPCs. Interestingly, pro-inflammatory cytokine tumor necrosis factor alpha (TNFα) decreased NOX4 expression and reduced survival of EPCs. However, the survival of EPCs was further diminished by TNF-α in NOX4-knockdown cells, suggesting that NOX4 has a protective role in EPCs.Conclusion: These findings suggest that NOX4-type NADPH oxidase is important for proliferation and migration functions of EPCs and protects against pro-inflammatory cytokine induced EPC death. These properties of NOX4 may facilitate the efficient function of EPCs which is vital for successful neovascularization.

Therapeutic Potential of Endothelial Colony-Forming Cells in Ischemic Disease: Strategies to Improve their Regenerative Efficacy

“…The transplantation of integrin β1-overexpressing ECFCs restored the local vascular network by physically engrafting within neovessels and rescued local blood flow [ 230 ]. A recent investigation focused on NADPH oxidase 4 (NOX4) [ 233 ], which is constitutively active and promotes H 2 O 2 production, thereby stimulating ECFC proliferation and migration [ 234 ]. NOX4 overexpression stimulated ECFC migration in vitro, albeit this effect was mediated by enhanced anion superoxide, rather than H 2 O 2 , production.…”

Inhibition of NADPH Oxidase Activation by Apocynin Rescues Seizure-Induced Reduction of Adult Hippocampal Neurogenesis

“…Although we have demonstrated that effects of NADPH oxidase activation in neuronal death after hypoglycemia [ 26 ], traumatic brain injury [ 22 ], and epilepsy [ 21 ], the efficacy of inhibition of NADPH oxidase activation by apocynin treatment in seizure-induced neurogenesis has not been demonstrated. We hypothesized that, if ROS are constantly produced after a seizure, the rate of newly proliferated neuronal survival is reduced as a function of the ROS production [ 29 , 30 , 31 ]. In the present study, we found that the survival rate and neurogenesis of hippocampal cells after seizure induction was increased via inhibition of NADPH oxidase.…”

Therapeutic Potential of Endothelial Colony Forming Cells in Ischemic Disease: Strategies to Improve Their Regenerative Efficacy

“…Transplantation of integrin β1-overexpressing ECFCs restored the local vascular network by physically engrafting within neovessels and rescued local blood flow [252]. A recent investigation focused on NADPH oxidase 4 (NOX4) [255], which is constitutively active and promotes H2O2 production, thereby stimulating ECFC proliferation and migration [256]. NOX4 overexpression stimulated ECFC migration in vitro, albeit this effect was mediated by enhanced anion superoxide, rather than H2O2, production.…”