Department of Integrative Medicine, Zhongshan Hospital Fudan University, Shanghai, China.
Laboratory of Neurology, Institute of Integrative Medicine, Fudan University, Shanghai, China.
Phytother Res. 2019 Sep;33(9):2329-2336. doi: 10.1002/ptr.6414. Epub 2019 Jun 26.
Bilobalide (BB), a constituent of the Ginkgo biloba extract, is a neuroprotective agent with multiple mechanisms of action. To further explore the potential therapeutic effects of BB in stroke, we investigated its effects on primary astrocytes using the oxygen and glucose deprivation-reoxygenation (OGD-R) model. Cell viability was measured by lactate dehydrogenase release assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Cell death was measured by annexin 5 conjgated with fluorescein isothiocyanate (V-FITC) assay, and reactive oxygen species (ROS) production was measured by 2',7'-Dichlorodihydrofluorescein Diacetate (DCFH-DA) probe. Manganese superoxide dismutase (MnSOD) expression was measured by western blot and immunofluorescence. Mitochondrial membrane potential was monitored using JC-1 staining. Our results show that OGD-R downregulated MnSOD and impaired mitochondrial function, which further enhanced ROS production in primary astrocytes. As a result, cell viability was compromised, and cell death increased. BB treatment protected astrocytes from those injuries mainly by restoring MnSOD level as MnSOD inhibitor abolished the effects of BB. In conclusion, we demonstrated that OGD-R induced astrocytic injury, but BB increased the expression of MnSOD, the ROS scavenger, to reverse the exacerbated astrocytic injury.
银杏叶提取物的成分白果内酯（BB）是一种具有多种作用机制的神经保护剂。为了进一步探讨 BB 在中风治疗中的潜在疗效，我们利用氧葡萄糖剥夺-复氧（OGD-R）模型研究了其对原代星形胶质细胞的作用。通过乳酸脱氢酶释放试验和 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐（MTT）比色法测定细胞活力。通过膜联蛋白 V-异硫氰酸荧光素（V-FITC）法测定细胞死亡，通过 2',7'-二氯二氢荧光素二乙酸酯（DCFH-DA）探针测定活性氧（ROS）的产生。通过蛋白质印迹法和免疫荧光法测定锰超氧化物歧化酶（MnSOD）的表达。通过 JC-1 染色监测线粒体膜电位。我们的结果表明，OGD-R 下调了 MnSOD 并损害了线粒体功能，这进一步增加了原代星形胶质细胞中的 ROS 产生。结果，细胞活力受损，细胞死亡增加。BB 治疗主要通过恢复 MnSOD 水平来保护星形胶质细胞免受这些损伤，因为 MnSOD 抑制剂消除了 BB 的作用。总之，我们证明了 OGD-R 诱导星形胶质细胞损伤，但 BB 增加了 ROS 清除剂 MnSOD 的表达，从而逆转了加剧的星形胶质细胞损伤。
