Department of Neurology, The First Affiliated Hospital of Soochow University, 188 Shizi Street, Suzhou, 215006, Jiangsu Province, China.
Department of Neurosurgery, The Affiliated Hospital of Qingdao University, 16 Jiangsu Road, Qingdao, 266003, Shandong Province, China.
Exp Neurol. 2019 Nov;321:113028. doi: 10.1016/j.expneurol.2019.113028. Epub 2019 Aug 1.
Owing to the continued high morbidity and high mortality rate after stroke, it is important to seek treatments other than conventional thrombolysis. Notch1 up-regulation participates in inflammatory responses after cerebral ischemia-reperfusion (I/R) injury, and it has been reported that Botch binds to and blocks Notch1 maturation. In this study, we investigated the role of Botch during cerebral (I/R) injury and explored its potential mechanisms. A middle-cerebral-artery occlusion/reperfusion (MCAO/R) model was established in adult male Sprague-Dawley rats in vivo, and cultured neurons and microglia were exposed to oxygen-glucose deprivation/reoxygenation (OGD/R) to mimic I/R injury in vitro. The results showed that protein levels of Botch and the Notch1 intracellular domain (NICD) were increased after MCAO/R. Furthermore, after overexpression of Botch, the generation of the activated form of Notch1, NICD, was decreased, while Botch knockdown or mutation led to an increase in NICD generation. As a result, Botch overexpression exhibited neuroprotective effects by significantly decreasing neurobehavioral phenotypes, improving infiltration of activated microglia, ameliorating inflammatory cytokine release, and inhibiting neuronal cell death. Conversely, Botch knockdown and mutation induced opposite effects. In addition, NICD was found to translocate to the mitochondria after OGD/R in neurons and microglia, which stimulated accumulation of reactive oxygen species in mitochondria and resulted in neuronal cell death and microglial activation. Botch overexpression inhibited the generation of NICD and decreased the translocation of NICD to the mitochondria, which inhibited neuronal cell death and ameliorated neuroinflammation. In conclusion, we found that Botch exerts neuroprotective effects via antagonizing the maturation of Notch1-induced neuronal injury and neuroinflammation, which may provide insights into novel therapeutic targets for the treatment of I/R injury.
由于中风后的高发病率和高死亡率，寻找除传统溶栓以外的治疗方法非常重要。Notch1 的上调参与了脑缺血再灌注（I/R）损伤后的炎症反应，据报道 Botch 与 Notch1 成熟结合并阻断其成熟。在本研究中，我们研究了 Botch 在脑（I/R）损伤中的作用，并探讨了其潜在机制。在体内建立成年雄性 Sprague-Dawley 大鼠大脑中动脉闭塞/再灌注（MCAO/R）模型，并在体外培养神经元和小胶质细胞进行氧葡萄糖剥夺/再氧合（OGD/R）模拟 I/R 损伤。结果表明，MCAO/R 后 Botch 和 Notch1 细胞内结构域（NICD）的蛋白水平增加。此外，Botch 过表达后 Notch1 激活形式的生成减少，而 Botch 敲低或突变导致 NICD 生成增加。因此，Botch 过表达通过显著降低神经行为表型、改善活化小胶质细胞浸润、改善炎症细胞因子释放和抑制神经元细胞死亡来发挥神经保护作用。相反，Botch 敲低和突变诱导了相反的作用。此外，在神经元和小胶质细胞中发现 NICD 在 OGD/R 后向线粒体易位，这刺激了线粒体中活性氧的积累，导致神经元细胞死亡和小胶质细胞激活。Botch 过表达抑制 NICD 的生成并减少 NICD 向线粒体的易位，从而抑制神经元细胞死亡和改善神经炎症。总之，我们发现 Botch 通过拮抗 Notch1 诱导的神经元损伤和神经炎症的成熟发挥神经保护作用，这可能为治疗 I/R 损伤提供新的治疗靶点。
