绞股蓝总苷对谷氨酸诱导的胎鼠大脑皮层神经元氧化性损伤保护机制的研究

Protection of gypenoside on glutamate-induced oxidation in the cultured cortical neurons of embryonic rats

目的：探讨绞股蓝总苷（gypenosides，GP）对谷氨酸（glutamic acid，Glu）诱导的胎鼠大脑皮层神经元损伤的保护作用，并从信号通路角度探讨其作用机制。方法：以体外培养的13-14d胎鼠大脑皮层神经元为研究对象，建立谷氨酸损伤和GP保护模型。采用MTT法测定细胞的存活率；荧光染料Ho33342和碘化丙碇（PI）荧光双染法鉴定细胞死亡方式；硝酸还原法测定细胞培养上清液中NO含量；生化法检测H2O2含量；流式细胞仪检测线粒体膜电位变化，分析GP对谷氨酸氧化性神经损伤的保护机制。结果：谷氨酸可导致神经细胞发生凋亡和坏死两种死亡方式；GP（200μ／ml）可明显抑制谷氨酸引起的NO、H2O2含量的升高，有效地防止线粒体膜电位的下降，从而提高细胞存活率。结论：GP可明显拮抗谷氨酸介导的氧化性神经损伤，提高细胞的存活率，其机制可能与提高神经细胞内抗氧化酶活性，清除氧自由基对线粒体膜的损伤有关。

Objective： To explore the neuroprotective effects of gypenoside（GP） on glutamate-induced cytotoxicity to cortical neurons and investigate its signaling pathway. Methods： The oxidative neurotoxicity model was constructed in primary cortical neurons isolated from Wistar rat E13 - 14 embryos following a high dose glutamate to the neurons directly. GP was added before the damage. The viability of cells after various treatments was detected bv MTr method. Ho33342 and PI double labeled method was used to analyse cellular death pathways. The contents of NO and H2O2 were detected. The changes of mitochondria membrane potential of neurons were analyzed by flow cytometry. Results： Glutamate triggered two cellular death pathways： necrosis and apoptosis. The cells, exposed toglutamate （2 mmol/L for 12hours）, showed charateristic change of damage, which could be relieved by GP （200 μg/ml） with cell survival rate increment. GP could block the increase of NO and H2O2, and the decrease of mitochondria memrane potential induced by glutamate. Consequently, the survival rate of neurons was greatly increased. Conclusion： Gypenoside can prevent the neurons from the damage of glutamate oxidative neurotoxicity and raise the neuron survival rate, which may be due to cleaning of the free radicals and improving of the mitochondrial function.