芍药苷对H_2O_2诱导SH-SY5Y神经细胞损伤的保护作用  

Protective effects of paeoniflorin against hydrogen peroxide-induced cell injury in SH-SY5Y nerve cells

作　　者：郭春燕[1] 罗强[2] 孙黎[2] 张丹参[1,3] 张纪平[1] 

机构地区：[1]河北北方学院药学系,河北张家口075000 [2]河北北方学院生命科学研究中心,河北张家口075000 [3]河北科技大学,河北石家庄050018

出　　处：《中草药》2013年第20期2864-2871,共8页Chinese Traditional and Herbal Drugs

基　　金：国家自然科学基金资助项目(81274005);河北省教育厅资助项目(2007302)

摘　　要：目的研究芍药苷对过氧化氢（H202）诱导人神经瘤母细胞（SH-SY5Y）凋亡的保护作用及其机制。方法制备H202诱导的氧化应激损伤模型。相差显微镜观察SH-SY5Y细胞形态的变化；CCK-8法测定细胞存活率；Hoechst33258染色法观察细胞凋亡；碘化丙啶染色、流式细胞仪检测细胞周期；2’，7’-二氯荧光素二乙酸盐（DCFH．DA）法检测细胞内活性氧（ROS）；INT显色反应法测定乳酸脱氢酶（LDH）的量；ELISA法检测8-羟基脱氧鸟苷（8-OHdG）的量；caspase-3催化特异性底物反应检测caspase-3活性。结果与对照组相比，200μmol／LH202作用SH-SY5Y细胞24h，使细胞存活力显著下降（P〈0．01），细胞凋亡率上升（P〈0．01），增殖指数（PI）降低（P〈0．05），8-OHdG的量上升（P〈0．01），LDH释放量和细胞内ROS量增加（P〈0．01），caspase-3活性增强（P〈0．01）。与H202损伤组相比，芍药苷终浓度为20-40gmol／L，可浓度相关地减轻H202诱导的sH—SY5Y细胞上述指标的变化（P〈O．05）；芍药苷10gmol／L组细胞PI、LDH和8-OHdG量未有明显改观，但能显著减轻H202诱导的SH-SY5Y细胞毒性、ROS和caspase-3活性（P〈0．05）。结论芍药苷对H2O2诱导SH—SY5Y细胞损伤具有显著保护作用，该作用可能与其清除ROS、减轻DNA氧化损伤、抑制细胞凋亡的caspase通路激活和调节细胞周期有关。Objective To investigate the neuroprotective effects ofpaeoniflorin （PF） against cell death induced by hydrogen peroxide （H2O2） in human neuroblastoma cells and its mechanisms. Methods H2O2 was used to induce SH-SYSY cell damages, and the cell survival rate was detected by CCK-8 assay; the cell morphologic changes were observed by inverted optical microscope; the apoptosis was tested using Hoechst 33258 staining; flow cytometer （FCM） and propidium iodide staining were used to analyze the apoptosis and cell cycle alteration; reactive oxygen species （ROS） production was determined by 2＇, 7＇-dichlorodihydrofluorescein diacetate （DCFH-DA） fluorescence; lactic dehydrogenase （LDH） release was detected by reaction of diaphorase and INT; 8-OHdG production was determined by enzyme-linked immunosorbent assay （ELISA）; caspase-3 activity was determined by caspase-3 catalyzing the specific substrate. Results Compared with control group, after the treatment with H2O2 （200 μmol/L） for 24 h, the viability and proliferation index of CH-SY5Y cells were significantly decreased （P 〈 0.01, 0.05）, the apoptosis rate and content of 8-OHdG were increased （P 〈 0.01）, the LDH resease and ROS production were increased （P 〈 0.01）; the activity of caspase 3 was increased （P 〈0.01）. Compared with H202 injury group, PF （20--40 μmol/L） significantly ameliorated the changes in SH-SY5Y cells induced by H2O2 in concentration-related manner （P 〈 0.05）. PF （10 μmol/L） did not significantly change the above indexes except the cell viability, ROS, and caspase-3 activity induced by H2O2 （P 〈 0.05）. Conclusion PF has the significant protective effect against the H2O2-induced cell injury, which may be related to eliminatinging ROS, alleviating DNA oxidative damage, regulation of cell cycle, and inhibition of apoptosis of caspase pathway activation.

关 键 词：芍药苷 SH—SY5Y细胞 过氧化氢 细胞凋亡 细胞周期 氧化应激 caspase通路 

分 类 号：R282.710.5[医药卫生—中药学] R971[医药卫生—中医学]