胞质多聚腺苷酸化元件结合蛋白4调控NOTCH信号通路对脑胶质瘤细胞凋亡的影响

The effect of CPEB4 on the apoptosis of brain glioma cells through NOTCH signaling pathway

探讨胞质多聚腺苷酸化元件结合蛋白4(cytoplasmic polyadenylation element binding protein 4,CPEB4)调控NOTCH信号通路对脑胶质瘤细胞凋亡的影响。收集人脑胶质瘤组织及对应的瘤旁组织,Western blotting检测组织中CPEB4蛋白水平。以人脑胶质瘤细胞U87为研究对象,通过细胞转染的方法将CPEB4小干扰RNA(CPEB4siRNA)和对照小干扰RNA(siRNA control)转染至U87细胞中,同时设置对照组,对照组细胞中只加入转染试剂。Western blotting检测细胞中CPEB4水平。流式细胞仪检测细胞凋亡情况。Western blotting检测NOTCH1受体胞内段基因NICD1、NOTCH2受体胞内段基因NICD2、NOTCH通路下游靶基因HES1、活化的含半胱氨酸的天冬氨酸蛋白水解酶3(cleaved cysteinyl aspartate specific proteinase 3,Cleaved Caspase-3)蛋白水平。人脑胶质瘤细胞经20μmol/L的NOTCH信号通路抑制剂S2188作用48h后,检测细胞凋亡及NICD1、NICD2、HES1、Cleaved Caspase-3蛋白水平。人脑胶质瘤组织中CPEB4蛋白水平明显高于瘤旁组织(P<0.01)。siRNA control组细胞中CPEB4、NICD1、NICD2、HES1、Cleaved Caspase-3蛋白水平和细胞凋亡率与对照组相比没有明显变化(P>0.05)。CPEB4siRNA组细胞中CPEB4水平明显低于对照组(P<0.01)。CPEB4siRNA组细胞凋亡率和Cleaved Caspase-3蛋白表达水平明显高于对照组(P<0.01)。CPEB4siRNA组细胞中NICD1、NICD2、HES1蛋白表达水平明显低于对照组(P<0.01)。NOTCH信号通路抑制剂作用后的人脑胶质瘤细胞凋亡情况同CPEB4siRNA组一样。由此可知CPEB4在人脑胶质瘤组织中表达上调,抑制CPEB4的表达能够促进人脑胶质瘤细胞凋亡,其作用机制与NOTCH信号通路有关。

We aimed to investigate the effect of CPEB4 on the apoptosis of brain glioma cells through NOTCH signaling pathway.CPEB4 protein was detected by Western blotting in the human brain glioma tissues and the corresponding adjacent tumor tissues.Human glioma cells U87 served as the research object.CPEB4 siRNA and siRNA control were transfected into U87 cells by cell transfection,while the control group was only added to the transfection reagent.The levels of CPEB4 in cells were detected by Western blotting.Cell apoptosis was detected by flow cytometry.Western blotting was used to determine the levels of NICD1,NICD2,HES1 and Cleaved Caspase-3.Human glioma cells were treated with 20μmol/L of NOTCH signaling pathway inhibitor S2188 for 48 h,and NICD1,NICD2,Cleaved Caspase-3,HES1 protein levels and the cell apoptosis were measured.The results showed that the levels of CPEB4 protein in human glioma tissues were significantly higher than those in the adjacent tumor tissues（P 0.01）.CPEB4,NICD1,NICD2,HES1,Cleaved Caspase-3 protein and apoptosis rate in siRNA control group had no significant change compared with the control group（P 0.05）.The level of CPEB4 in CPEB4 siRNA group was significantly lower than that in control group（P 0.01）.The cell apoptosis rate and the expression level of Cleaved Caspase-3 in CPEB4 siRNA group were significantly higher than those in control group（P 0.01）.The expression levels of NICD1,NICD2 and HES1 in CPEB4 siRNA group were significantly lower than those in control group（P 0.01）.The apoptosis of human glioma cells after blockage of NOTCH signaling pathway by inhibitor was consistent with the CPEB4 siRNA group.CPEB4 expression was up-regulated in human glioma tissues.Thus,inhibiting the expression of CPEB4 can promote the apoptosis of human glioma cells,and the mechanism is related to the NOTCH signaling pathway.