NADPH Oxidase–Generated Reactive Oxygen Species Are Required for Stromal Cell–Derived Factor-1α–Stimulated Angiogenesis

Abstract

Objective— Reactive oxygen species (ROS) act as signaling molecules during angiogenesis; however, the mechanisms used for such signaling events remain unclear. Stromal cell–derived factor-1α (SDF-1α) is one of the most potent angiogenic chemokines. Here, we examined the role of ROS in the regulation of SDF-1α–dependent angiogenesis. Approach and Results— Bovine aortic endothelial cells were treated with SDF-1α, and intracellular ROS generation was monitored. SDF-1α treatment induced bovine aortic endothelial cell migration and ROS generation, with the majority of ROS generated by bovine aortic endothelial cells at the leading edge of the migratory cells. Antioxidants and nicotinamide adenine dinucleotide phosphate oxidase (NOX) inhibitors blocked SDF-1α–induced endothelial migration. Furthermore, knockdown of either NOX5 or p22phox (a requisite subunit for NOX1/2/4 activation) significantly impaired endothelial motility and tube formation, suggesting that multiple NOXs regulate SDF-1α–dependent angiogenesis. Our previous study demonstrated that c-Jun N-terminal kinase 3 activity is essential for SDF-1α–dependent angiogenesis. Here, we identified that NOX5 is the dominant NOX required for SDF-1α–induced c-Jun N-terminal kinase 3 activation and that NOX5 and MAP kinase phosphatase 7 (MKP7; the c-Jun N-terminal kinase 3 phosphatase) associate with one another but decrease this interaction on SDF-1α treatment. Furthermore, MKP7 activity was inhibited by SDF-1α, and this inhibition was relieved by NOX5 knockdown, indicating that NOX5 promotes c-Jun N-terminal kinase 3 activation by blocking MKP7 activity. Conclusions— We conclude that NOX is required for SDF-1α signaling and that intracellular redox balance is critical for SDF-1α–induced endothelial migration and angiogenesis.