【摘要】 目的探讨神经酰胺对胶质瘤细胞87-MG和U251自噬性死亡的作用及机制。方法采用MTT和流式细胞的方法检测不同浓度神经酰胺刺激87-MG和U251细胞后细胞存活和凋亡的改变;电镜和Western blotting技术检测自噬和JNK/c-Jun信号通路的改变,JNK药理性抑制剂SP600125特异性抑制JNK通路,观察其对神经酰胺诱导自噬死亡的影响。结果神经酰胺刺激24 h后,87-MG和U251存活呈现时间依赖性下降(P<0.05);相应的细胞死亡数目剂量依赖性升高(P<0.05),但凋亡性死亡比例较低;神经酰胺刺激后镜下观察到的自噬小体计数,LC3B/LC3A和Beclin-1的表达以及JNK/c-Jun的磷酸化程度都增加(P<0.05)。提前给予SP600125抑制JNK信号通路的活性后,可以阻断神经酰胺诱导的细胞自噬性死亡(P<0.05)。结论神经酰胺可诱导胶质瘤细胞87-MG和U251出现自噬性死亡,机制可能与JNK信号通路的活化有关。更多还原

【Abstract】 Objective To observe the autophagy of 87-MG and U251 glioma cells induced by ceramide and explore the possible mechanism.Methods The viability and apoptosis of 87-MG and U251 cells were detected by MTT assay and flow cytometry,respectively.Autophagic-related protein expressions of LC3 B /LC3 A and Beclin-1 were determined by Western blotting.The activation of JNK/c-Jun signaling pathway induced by ceramide with or without the treatment of JNK specific inhibitor SP600125 was also measured.Results 24 hours after treatment of ceramide,the growth of 87-MG and U251 cells was significantly inhibited time-dependently(P<0.05); and the number of autophagic cells increased dose-dependently(P<0.05).The levels of LC3B/LC3 A and Beclin-1 significantly increased after ceramide treatment(P<0.05).JNK signaling pathway was activated in the 87-MG and U251 cells and the phosphorylation of c-Jun also increased after ceramide treatment.This activation of autophagy could be reversed by the pre-treatment of SP600125.Conclusion Ceramide may induce autophagy in 87-MG and U251 glioma cells and the mechanism may be related to the activation of JNK/c-Jun signaling pathway.更多还原