1Department of Pharmacology, School of Medicine, National Yang-Ming University, Taipei, Taiwan
2National Research Institute of Chinese Medicine, Taipei, Taiwan
3Division of Neurovascular Disease, Neurological Institute, Taipei Veterans General Hospital and School of Medicine, National Yang-Ming University, Taipei, Taiwan
4Institute of Biomedical Sciences, National Chung-Hsing University, Taichung, Taiwan
5Department of Psychiatry, Taipei Veterans General Hospital, Taipei, Taiwan
6Department of Chinese Martial Arts, Chinese Culture University, Taipei, Taiwan
7Department of Nursing, College of Medicine and Nursing, Hungkuang University, Taichung, Taiwan
This study aimed to explore the mechanisms by which andrographolide protects against hypoxia-induced oxidative/nitrosative brain injury provoked by cerebral ischemic/reperfusion (CI/R) injury in mice. Hypoxia in vitro was modeled using oxygen-glucose deprivation (OGD) followed by reoxygenation of BV-2 microglial cells. Our results showed that treatment of mice that have undergone CI/R injury with andrographolide (10–100 µg/kg, i. v.) at 1 h after hypoxia ameliorated CI/R-induced oxidative/nitrosative stress, brain infarction, and neurological deficits in the mice, and enhanced their survival rate. CI/R induced a remarkable production in the mouse brains of reactive oxygen species (ROS) and a significant increase in protein nitrosylation; this primarily resulted from enhanced expression of NADPH oxidase 2 (NOX2), inducible nitric oxide synthase (iNOS), and the infiltration of CD11b cells due to activation of nuclear factor-kappa B (NF-κB) and hypoxia-inducible factor 1-alpha (HIF-1α). All these changes were significantly diminished by andrographolide. In BV-2 cells, OGD induced ROS and nitric oxide production by upregulating NOX2 and iNOS via the phosphatidylinositol-3-kinase (PI3K)/AKT-dependent NF-κB and HIF-1α pathways, and these changes were suppressed by andrographolide and LY294002. Our results indicate that andrographolide reduces NOX2 and iNOS expression possibly by impairing PI3K/AKT-dependent NF-κB and HIF-1α activation. This compromises microglial activation, which then, in turn, mediates andrographolide's protective effect in the CI/R mice.
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