A time course of NADPH-oxidase up-regulation and endothelial nitric oxide synthase activation in the hippocampus following neurotrauma

• Published in: Free radical biology & medicine 2014-12, Vol.77, p.21-29
• Main Authors: Ansari, Mubeen A., Roberts, Kelly N., Scheff, Stephen W.
• Format: Article
• Language: English
• Subjects: Animals; Brain Injuries - enzymology; Cell Membrane - enzymology; Enzyme Activation; Free radicals; Hippocampus; Male; NADH, NADPH Oxidoreductases - metabolism; NADPH Oxidases - metabolism; NADPH-oxidase; Nitric Oxide - metabolism; Nitric Oxide Synthase Type III - metabolism; Phosphoproteins - metabolism; Protein Subunits - metabolism; Protein Transport; Rats, Sprague-Dawley; Secondary injury cascades; Superoxides - metabolism; Traumatic brain injury; Up-Regulation
• ISSN: 0891-5849; 1873-4596
• DOI: 10.1016/j.freeradbiomed.2014.08.025

Nicotinamide adenine dinucleotide phosphate oxidase (NADPH-oxidase; NOX) is a complex enzyme responsible for increased levels of reactive oxygen species (ROS), superoxide (O2•−). NOX-derived O2•− is a key player in oxidative stress and inflammation-mediated multiple secondary injury cascades (SIC) following traumatic brain injury (TBI). The O2•− reacts with nitric oxide (NO), produces various reactive nitrogen species (RNS), and contributes to apoptotic cell death. Following a unilateral cortical contusion, young adult rats were killed at various times postinjury (1, 3, 6, 12, 24, 48, 72, and 96h). Fresh tissue from the hippocampus was analyzed for NOX activity, and level of O2•−. In addition we evaluated the translocation of cytosolic NOX proteins (p67Phox, p47Phox, and p40Phox) to the membrane, along with total NO and the activation (phosphorylation) of endothelial nitric oxide synthase (p-eNOS). Results show that both enzymes and levels of O2•− and NO have time-dependent injury effects in the hippocampus. Translocation of cytosolic NOX proteins into membrane, NOX activity, and O2•− were also increased in a time-dependent fashion. Both NOX activity and O2•− were increased at 6h. Levels of p-eNOS increased within 1h, with significant elevation of NO at 12h post-TBI. Levels of NO failed to show a significant association with p-eNOS, but did associate with O2•−. NOX up-regulation strongly associated with both the levels of O2•− and the total NO. The initial 12h post-TBI are very important as a possible window of opportunity to interrupt SIC. It may be important to selectively target the translocation of cytosolic subunits for the modulation of NOX function. •NADPH-oxidase (NOX) is up-regulated by 6h in the hippocampus following TBI.•Up-regulation of NOX and derived O2•− are associated with the increased NO after TBI.•Following TBI, up-regulation of eNOS in the hippocampus occurred early after 1h.•Activation of eNOS is not driving an increase in NO in the hippocampus following TBI.