促红细胞生成素对MPP^+所致的PC12细胞线粒体氧化损伤的保护作用  

Protective Effects of Erythropoietin on PC12 Cells against MPP^+ Induced Oxidative Damage

作　　者：吴艳[1] 孙欣[1] 方鑫[1] 王芳[1] 黄露琪[1] 刘仁刚[2] 乔娴[1] 孙圣刚[1] 

机构地区：[1]华中科技大学同济医学院附属协和医院神经内科,武汉430022 [2]华中科技大学同济医学院基础医学院解剖学系,武汉430030

出　　处：《华中科技大学学报（医学版）》2010年第2期156-160,共5页Acta Medicinae Universitatis Scientiae et Technologiae Huazhong

基　　金：国家自然科学基金资助项目(No.30570627)

摘　　要：目的探讨促红细胞生成素(EPO)对1-甲基-4-苯基吡啶离子(MPP+)诱导PC12细胞变性损伤的保护作用及其机制。方法以MPP+损伤PC12细胞作为帕金森病(PD)细胞模型,采用四甲基偶氮唑盐(MTT)法检测细胞存活率,流式细胞术检测细胞凋亡率,罗丹明123(Rh123)染色检测细胞线粒体膜电位(ΔΨm),双氯荧光黄乙酸乙酯(DCF-DA)染色检测细胞内活性氧(ROS)的含量,免疫印迹法检测pAkt蛋白表达。结果①500μmol/L的MPP+可以导致PC12细胞的存活率下降,凋亡率增加,0.1~10 U/mL的EPO可以使PC12细胞的存活率增加,凋亡率降低,并在1 U/mL时达到最大保护效应;②MPP+导致ROS含量增加、ΔΨm降低;EPO可以抑制MPP+所致ROS和ΔΨm水平的变化;③EPO增加了Akt磷酸化水平,PI3K抑制剂LY294002拮抗了EPO的保护作用和对ROS产生的抑制作用。结论EPO对MPP+损伤PC12细胞具有保护作用,其机制是通过PI3K/Akt通路实现的。Objective To investigate whether erythropoietin（EPO） protects PC12 cells against MPP＋ induced oxidative damage.Methods PC12 cells treated by MPP＋ were used as the cell model of Parkinson＇s disease.Methyl thiazolyl tetrazolium（MTT） was used to measure the viability of the PC12 cells exposed to gradient concentration of EPO,and flow cytometry was used to assay the apoptosis rate of PC12 cells.The reactive oxygen species（ROS） level and the mitochondrial transmembrane potential in each group were determined by spectrofluorometer.The expression of pAkt in PC12 cells was detected by Western blot.Results ①Treatment of PC12 cells with MPP＋ reduced cell viability,which might be associated with the elevation in apoptosis rate.EPO significantly reversed these responses and had the maximum protective effect at 1 U/mL;②It was also found that EPO inhibited the formation of ROS and the disruption of mitochondrial transmembrane potential induced by MPP＋;③Akt,a substrate of PI3K,was activated by EPO.In addition,EPO failed to rescue cells from MPP＋ insult in the presence of the PI3K inhibitor,LY294002.Furthermore,the effect of EPO on ROS levels was also blocked by LY294002.Conclusion EPO affords significant neuroprotection against MPP＋ induced injury in PC12 cells via PI3K/Akt-mediated signaling pathway.

关 键 词：促红细胞生成素 1-甲基-4-苯基吡啶离子 PC12细胞 活性氧 线粒体膜电位 AKT 

分 类 号：R742.5[医药卫生—神经病学与精神病学]