title: 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). H2O2production 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 H2O2formation in EPCs. Inhibition of NOX4-derived H2O2decreased 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.
authors.affiliation: Centre for Eye Research Australia, Royal Victorian Eye and Ear HospitalEast Melbourne, VIC, Australia; Ophthalmology, University of Melbourne, Department of SurgeryEast Melbourne, VIC, Australia; Department of Pharmacology and Therapeutics, University of MelbourneMelbourne, VIC, Australia; Faculty of Applied Medical Sciences, King Abdulaziz UniversityJeddah, Saudi Arabia.
authors.forename: Nora Y
authors.initials: NY
authors.lastname: Hakami
authors.affiliation: Cardiology, Icahn School of Medicine at Mount Sinai Hospital New York, NY, USA.
authors.forename: Amaresh K
authors.initials: AK
authors.lastname: Ranjan
authors.affiliation: Diabetes and Islet Biology, NHMRC Clinical Trials Centre, University of Sydney Sydney, NSW, Australia.
authors.forename: Anandwardhan A
authors.initials: AA
authors.lastname: Hardikar
authors.affiliation: Centre for Eye Research Australia, Royal Victorian Eye and Ear HospitalEast Melbourne, VIC, Australia; Ophthalmology, University of Melbourne, Department of SurgeryEast Melbourne, VIC, Australia.
authors.forename: Greg J
authors.initials: GJ
authors.lastname: Dusting
authors.affiliation: Centre for Eye Research Australia, Royal Victorian Eye and Ear HospitalEast Melbourne, VIC, Australia; Ophthalmology, University of Melbourne, Department of SurgeryEast Melbourne, VIC, Australia.
authors.forename: Hitesh M
authors.initials: HM
authors.lastname: Peshavariya
keywords: H2O2; NOX4; ROS; TNFα; endothelial progenitor cells