摘要
目的:探讨甲基苯丙胺(methamphetamine,METH)对SH-SY5Y细胞的毒性作用和对α-核突触蛋白(α-synuclein,α-SN)表达的影响,为研究α-SN在METH神经毒性机制中的作用奠定基础。方法:分别用浓度0、0.5、1.5、2.5、3.5、4.5 mmol.L-1的METH处理SH-SY5Y细胞24 h,倒置显微镜下观察细胞形态变化;透射电镜观察细胞超微结构;CCK-8法检测细胞存活率;流式细胞术分析细胞凋亡率;实时荧光定量PCR和Western Blot分别检测α-SN基因和蛋白水平的表达变化。结果:以0 mmol.L-1为对照组,0.5-4.5 mmol.L-1METH处理的SH-SY5Y细胞镜下可见胞体皱缩变圆,突起变短、断裂、消失。电镜显示,METH处理的细胞内可见包涵体和自噬小体。0.5 mmol.L-1处理组与对照组比较,细胞存活率无显著性差异(P=0.274),其他浓度处理组与对照组相比均有显著性差异(P<0.001)。细胞凋亡率随METH浓度增加而递增,与对照组相比均有显著性差异(P<0.001)。与对照组比较,0.5 mmol.L-1处理组α-SN-mRNA的表达量没有显著差异(P=0.936),其他浓度处理组α-SN-mRNA均显著性上升(P<0.001),且α-SN-mRNA表达量随METH浓度增加而逐步上升。α-SN蛋白的表达也随METH浓度增加而递增。结论:METH对SH-SY5Y细胞有毒性作用并可刺激细胞中α-SN基因和蛋白水平的高表达。细胞毒性作用和α-SN的表达均随METH浓度的增加而递增。
Objective：To investigate METH-induced toxic effects and the expression of α-synuclein in SH-SY5Y cells treated with METH.Methods：SH-SY5Y cells were treated by METH with the concentration of 0,0.5,1.5,2.5,3.5 or 4.5 mmol·L-1 for 24 h.The morphological changes and cell ultrastructure were observed by optical and electron microscopy,respectively.Cell viability and apoptosis rates were assessed by CCK-8 assay and flow cytometry,respectively.The expression levels of α-SN mRNA and protein were detected by quantitative real-time PCR and Western Blot respectively.Results：The SH-SY5Y cells exposed to METH were morphologically featured by shrinkage and dendrite disruption of cells.Autophagosome and inclusion body could be observed in cytoplasm.METH administration caused a dose-dependent reduction in the cell viability（P0.001）,resulting in an increased cell early apoptotic rate（P0.001）.The both of α-SN mRNA and protein expression increased with dose-dependent manner in response to METH treatment.Conclusion：Both the cytotoxicity and expression of α-SN in the SH-SY5Y cells are gradually enhanced with the increase of METH concentration.
出处
《中国药物依赖性杂志》
CAS
CSCD
2013年第3期176-181,共6页
Chinese Journal of Drug Dependence
基金
国家自然科学基金(81072506)
