Abstract 36: Elevated Expression of Dual Oxidases (Duox) Under Ischemic Conditions Contributes to Oxidative Stress and Decreased Cell Viability.
Introduction: Redox imbalance during ischemia/reperfusion Injury in the brain following focal ischemic stroke is associated with impaired neurological functions. Following ischemia/reperfusion, there is a burst of oxidative stress/reactive oxygen species (ROS). ROS physiologically serve as signaling molecules but excessive amounts of ROS contribute to destruction of endothelial integrity, blood brain barrier disruption, and neuronal cell death. NADPH oxidases (NOX) are important sources of ROS during ischemic stroke. Dual oxidases (Duox) are new members of NOX family and are known for directly generating hydrogen peroxide/ROS. Role of Duox following brain ischemic injury has been little investigated.
Methods: Ischemia was induced by middle cerebral artery occlusion (MCAO) and reperfusion. In vitro ischemia was generated by oxygen-glucose deprivation (OGD). Expression was analysed by western blot analysis, RT-PCR, Immunofluorescence staining. ROS was measured by dichloro-dihydrofluorescein staining.
Results: Our preliminary studies for the first time have identified that expression of Duox are spatially and temporally regulated in ipsilateral hemisphere of spontaneously hyertensive rat brains following MCAO and reperfusion. Expression of Duox 1 was up-regulated about 2 folds after 3 days, and16 folds after 7 days of reperfusion. Its expression was further increased to about 30 folds at day 21 after reperfusion. The expression of Duox 1 was specifically up-regulated in activated astrocytes and VWF and CD31 positive endothelial cells indicating its presence in blood vessels. In support of these in vivo observations, Duox1 expression was significantly increased in astrocytes and endothelial cell cultures exposed to either ATP or OGD. Duox1 expression was associated with increased ROS production and decreased cell viability. However, pre-treatment with Duox1 specific siRNA decreased the ROS production and increased the cell viability by about 30 percent.
Summary: Up-regulation of Duox under ischemic/reperfusion conditions suggests that this enzyme potentially contributes to excessive ROS production. Further studies targeting these novel oxidases may open up a promising approach to reduce brain injury after stroke.
- © 2012 by American Heart Association, Inc.