DataSheet_2_Neuroprotective Effects and Mechanisms of Zhenlong Xingnao Capsule in In Vivo and In Vitro Models of Hypoxia.docx

Zhenlong Xingnao Capsule (ZXC) is a Tibetan medicine used to treat ischemic stroke. In this study, we determined the in vitro and in vivo effects of ZXC on reactive oxygen species (ROS) in a mouse BV-2 microglial cell hypoxia-reoxygenation and rat middle cerebral artery occlusion infarction models. We aimed to clarify the role of ZXC in cerebral ischemia protection; reveal amino acid neurotransmitter changes in the frontal cortex after drug intervention; determine mRNA and protein expression changes in Bcl-2, Bax, caspase-3, P38, and nuclear factor (NF)-кB in the frontal cortex and changes in antioxidant indices in the brain; and elucidate the mechanisms underlying ZXC action. After hypoxia-reoxygenation, ROS levels were significantly increased in BV-2 cells, and their levels decreased after treatment with ZXC. ZXC had protective effects on ischemic/anoxic injury in vitro and in vivo by downregulating the expressions of caspase-3 and NF-кB mRNA during ischemia and reperfusion and that of p38 and caspase-3 during acute ischemia and reperfusion as well as the steady-state levels of excitatory amino acids/inhibitory amino acids and by improving the total antioxidant capacity and total superoxide dismutase activities during ischemia. These findings provide new molecular evidence for the mechanisms underlying ZXC action.

真龙醒脑胶囊（ZXC）为一种用于治疗缺血性脑卒中的藏药。本研究旨在探讨ZXC在体外和体内对小鼠BV-2小胶质细胞缺氧-复氧及大鼠大脑中动脉阻塞梗死模型中反应性氧种（ROS）的影响，以阐明ZXC在脑缺血保护中的作用；揭示药物干预后前额叶皮层中氨基酸神经递质的变化；确定前额叶皮层中Bcl-2、Bax、caspase-3、P38和核因子（NF)-кB的mRNA和蛋白表达变化，以及脑中抗氧化指数的变化；并阐明ZXC作用机制。缺氧-复氧处理后，BV-2细胞中的ROS水平显著升高，经ZXC治疗后其水平下降。ZXC通过在缺血和再灌注期间下调caspase-3和NF-кB mRNA的表达，以及在急性缺血和再灌注期间下调p38和caspase-3的表达，以及兴奋性氨基酸/抑制性氨基酸的稳态水平，并在缺血期间提高总抗氧化能力和总超氧化物歧化酶活性，从而在体外和体内对缺血/缺氧性损伤产生保护作用。这些发现为ZXC作用机制提供了新的分子证据。