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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。
To investigate the effect of cytoplasmic polyadenylation element binding protein 4 (CPEB4) on the apoptosis of glioma cells by regulating the NOTCH signaling pathway. Collected human glioma tissue and the corresponding para-tumor tissue, Western blotting detection of tissue CPEB4 protein levels. The human glioma U87 cells were transfected into U87 cells with CPEB4 siRNA and CPEB4 siRNA by cell transfection. At the same time, the cells of control group and control group Add transfection reagent only. Western blotting was used to detect the level of CPEB4 in cells. Flow cytometry detection of apoptosis. Western blotting was used to detect the intracellular domain of NICD1 and NOTCH2 receptors in NOTCH1 receptor, NICD2, the downstream target of NOTCH, HES1, cleaved cysteinyl aspartate specific proteinase 3 3, Cleaved Caspase-3) protein levels. Human glioma cells were treated with 20 μmol / L NOTCH signaling inhibitor S2188 for 48 hours, and the apoptosis and the protein levels of NICD1, NICD2, HES1 and Cleaved Caspase-3 were detected. The level of CPEB4 protein in human glioma tissues was significantly higher than that in the adjacent tissues (P <0.01). Compared with the control group, the levels of CPEB4, NICD1, NICD2, HES1 and Cleaved Caspase-3 protein in the siRNA control group did not change significantly (P> 0.05). The level of CPEB4 in CPEB4siRNA group was significantly lower than that in control group (P <0.01). The apoptosis rate and Cleaved Caspase-3 protein expression in CPEB4siRNA group were significantly higher than those in control group (P <0.01). The expression of NICD1, NICD2 and HES1 in CPEB4siRNA group was significantly lower than that in control group (P <0.01). The apoptosis of human glioma cells treated with NOTCH signaling pathway inhibitor was the same as CPEB4siRNA group. This shows that CPEB4 is upregulated in human glioma tissue and inhibition of CPEB4 expression can promote human glioma cell apoptosis, and its mechanism of action is related to NOTCH signaling pathway.