饥饿诱导胶质瘤细胞自噬发生及Beclin-1表达的变化

目的 探讨饥饿诱导大鼠胶质瘤细胞C6自噬的发生及自噬相关蛋白Beclin-1的变化。方法 在培养大鼠胶质瘤细胞C6至对数生长期后，以无氨基酸培养液EBSS(Earle’s balanced salt)代替DMEM培养液培养细胞，培养1、3、6、12、18 h后收集细胞，采用蛋白质印迹分析和免疫荧光检测特异性标记自噬的微管相关蛋白1轻链3(microtubule-associated protein 1 light chain 3，LC-3)以及自噬相关蛋白Beclin-1，采用透射电镜观察自噬体。结果 饥饿处理1 h后胶质瘤细胞C6中LC-3表达开始增加，并随着饥饿时间延长而逐渐增加，12 h达到高峰后开始下降；免疫荧光下可见点状自噬体;透射电镜下观察到自噬体的形成。饥饿处理1 h后胶质瘤细胞C6中Beclin-1表达开始升高，3 h达到高峰后逐渐下降；免疫荧光下观察到Beclin-1表达增高。结论 饥饿可以诱导胶质瘤细胞C6发生自噬，Beclin-1与饥饿诱导的胶质瘤细胞C6的自噬相关，为进一步研究Beclin-1在胶质瘤细胞自噬中的作用机制打下基础。

Starvation-induced autophagy in glioma and changes in Beclin-1 expression

Objective To investigate the starvation-induced autophagy in rat C6 glioma cells and the changes of autophagy-related protein Beclin-1. Methods Rat C6 glioma cells of the logarithmic growth phase were cultured with amino acid-free EBSS medium instead of the DMEM medium. Cells were collected at 1 h, 3 h, 6 h, 12 h, and 18 h after EBSS culture. Western blotting analysis and immunofluorescence were used to detect the specific markers of autophagic microtubule-associated protein 1 light chain 3 (LC-3) and autophagy-related protein Beclin-1, and transmission electron microscope was used to observe the autophagic body. Results LC-3 expression was found in C6 glioma cells 1 h after starvation; the expression increased with the prolongation of starvation, and reached a peak 12 h later and then began to decline. Autophagic bodies were observed under transmission electron microscopy, and autophagosomes were found in immunofluorescence. Beclin-1 expression began to increase in C6 glioma cells 1 h after starvation, reached a peak at 3 h and then decreased gradually. Increased expression of Beclin-1 was also observed by immunofluorescence. Conclusion Starvation can induce autophagy in C6 glioma cells, which is correlated with the expression of autophagy-related protein Beclin-1. This study lays a foundation for studying the autophagy mechanism in glioma cells.