Department of Neurology, The Hospital of Shunyi District Beijing, Beijing, 101300, China.
Department of General Surgery, The Second Xiangya Hospital of Central South University, Changsha, 410011, China.
Neurochem Res. 2020 Feb;45(2):404-417. doi: 10.1007/s11064-019-02931-0. Epub 2020 Jan 16.
The reperfusion after an acute ischemic stroke can lead to a secondary injury, which is ischemia-reperfusion (I/R) injury. During ischemia, the reactive oxygen species (ROS) is over-produced, mostly from nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX). Besides, miRNAs are also associated with neuronal death in ischemic stroke. MiR-124-5p is selectively expressed within central nervous system (CNS) and is predicted to bind to NOX2 directly. Herein, we successfully set up cerebral I/R injury model in rats through middle cerebral artery occlusion (MCAO) surgery. After 12 h or 24 h of refusion, the superoxide dismutase (SOD) activity was significantly inhibited, accompanied by NOX2 protein increase within MCAO rat infarct area. In vitro, oxygen-glucose deprivation/refusion (OGD/R) stimulation on PC-12 cells significantly increased NOX2 protein levels, ROS production, and the cell apoptosis, while a significant suppression on SOD activity; OGD/R stimulation-induced changes in PC-12 cells described above could be significantly attenuated by NOX2 silence. In vivo, miR-124 overexpression improved, whereas miR-124 inhibition aggravated I/R injury in MCAO rats. miR-124-5p directly bound to the CYBB 3'-untranslated region (UTR) to negatively regulate CYBB expression and NOX2 protein level. In vitro, miR-124 overexpression improved, while NOX2 overexpression aggravated OGD/R-induced cellular injuries; NOX2 overexpression significantly attenuated the effects of miR-124 overexpression. Besides, miR-124 overexpression significantly repressed NF-κB signaling activation and TNFα and IL-6 production through regulating NOX2. In conclusion, miR-124-5p/NOX2 axis modulates NOX-mediated ROS production, the inflammatory microenvironment, subsequently the apoptosis of neurons, finally affecting the cerebral I/R injury.
急性缺血性脑卒中再灌注后可导致继发性损伤，即缺血再灌注（I/R）损伤。在缺血期间，活性氧（ROS）过度产生，主要来自烟酰胺腺嘌呤二核苷酸磷酸（NADPH）氧化酶（NOX）。此外，miRNA 也与缺血性脑卒中的神经元死亡有关。miR-124-5p 选择性地在中枢神经系统（CNS）中表达，并被预测可直接与 NOX2 结合。在此，我们通过大脑中动脉闭塞（MCAO）手术成功地在大鼠中建立了脑 I/R 损伤模型。再灌注 12 或 24 小时后，超氧化物歧化酶（SOD）活性显著抑制，同时 MCAO 大鼠梗死区的 NOX2 蛋白增加。体外，氧葡萄糖剥夺/再灌注（OGD/R）刺激 PC-12 细胞显著增加了 NOX2 蛋白水平、ROS 产生和细胞凋亡，同时 SOD 活性显著降低；NOX2 沉默可显著减轻 OGD/R 刺激诱导的 PC-12 细胞的上述变化。体内，miR-124 的过表达改善了 MCAO 大鼠的 I/R 损伤，而 miR-124 的抑制则加重了损伤。miR-124-5p 直接与 CYBB 3'-非翻译区（UTR）结合，负调控 CYBB 表达和 NOX2 蛋白水平。体外，miR-124 的过表达改善了 OGD/R 诱导的细胞损伤，而 NOX2 的过表达则加重了损伤；NOX2 的过表达显著减弱了 miR-124 过表达的作用。此外，miR-124 的过表达通过调节 NOX2 显著抑制 NF-κB 信号激活和 TNFα 和 IL-6 的产生。综上所述，miR-124-5p/NOX2 轴调节 NOX 介导的 ROS 产生、炎症微环境，进而影响神经元凋亡，最终影响脑 I/R 损伤。
