RNA干扰ATF5基因下调NF-κB信号诱导脑胶质瘤细胞凋亡

Knock-down of ATF5 gene expression by RNA interference down-regulates NF-κB signaling to induce apoptosis in glioma cells

目的:探讨RNA干扰转录激活因子5(ATF5)基因表达对脑胶质瘤细胞活力、凋亡率及NF-κB信号通路的影响。方法:将设计合成的ATF5特异性siRNA(siATF5组)和阴性对照siRNA(NC组)分别转染人脑胶质瘤U251细胞,采用NF-κB信号通路抑制剂PDTC处理细胞,MTT法、流式细胞术及Western blot法分别检测细胞活力、凋亡率及ATF5、血管内皮生长因子(VEGF)、环氧化酶2(COX-2)、 p-p65、p-IκB和Bax的蛋白水平。结果:转染siATF5的U251细胞中ATF5的蛋白表达显著低于空白对照组(P<0.05)。与空白对照组比较,siATF5组细胞活力明显降低,凋亡率明显升高,VEGF、COX-2、 p-p65和p-IκB的蛋白水平明显降低,Bax的蛋白水平明显升高(P<0.05)。与siATF5组相比,siATF5和PDTC同时处理的U251细胞活力显著降低,凋亡率显著升高(P<0.05)。结论:下调ATF5基因表达可降低脑胶质瘤细胞的活力,诱导细胞凋亡,降低免疫抑制因子VEGF和COX-2表达,其机制可能与下调NF-κB信号通路有关。

AIM: To investigate the effect of activating transcription factor 5(ATF5) gene expression knock-down by RNA interference on the viability, apoptotic rate and NF-κB signaling in the glioma cells. METHODS: The U251 cells were transfected with ATF5 specific siRNA(siATF5 group) and negative control siRNA(NC group). NF-κB signaling pathway inhibitor PDTC was used to treat the cells. MTT assay, flow cytometry and Western blot were used to detect the cell viability, apoptotic rate and the protein levels of ATF5, vascular endothelial growth factor(VEGF), cyclooxygenase-2(COX-2), p-p65, p-IκB and Bax, respectively. RESULTS: The protein level of ATF5 in the U251 cells transfected with siATF5 was significantly lower than that in blank control group(P<0.05). Compared with blank control group, the cell viability was significantly decreased, the apoptotic rate was significantly increased, the protein levels of VEGF, COX-2, p-p65 and p-IκB were significantly decreased, and the protein expression of Bax was significantly increased in siATF5 group(P<0.05). The viability of U251 cells treated with siATF5 and PDTC was significantly lower than that in siATF5 group, and the apoptotic rate was higher than that in siATF5 group(P<0.05). CONCLUSION: Knock-down of ATF5 gene expression reduces the viability of glioma cells, induces apoptosis and reduces the protein levels of VEGF and COX-2. The mechanism may be related to the down-regulation of NF-κB signaling pathway.