沉默VEGF对人鼻咽癌放射抗拒细胞株CNE-2R侵袭迁移的影响

Effect of silencing VEGF on the invasion and migration of radioresistant human nasopharyngeal carcinoma cell line CNE-2R

目的探讨沉默血管内皮生长因子(vascular endothelial growth factor,VEGF)对人鼻咽癌放射抗拒细胞株CNE-2R迁移和侵袭能力的影响。方法制备含VEGF-shRNA的慢病毒载体(VEGF-LV1、VEGF-LV2、VEGF-LV3)和对照shRNA重组慢病毒(VEGF-NC),并转染人鼻咽癌放射抗拒细胞株CNE-2R,建立稳定沉默VEGF细胞(CNE-LV1、CNE-LV2和CNE-LV3)和阴性对照细胞(CNE-NC)。采用RT-qPCR和Western blot法检测细胞转染效果,划痕实验和Transwell小室实验检测VEGF沉默后细胞的迁移和侵袭能力。结果成功构建VEGF-shRNA重组慢病毒并建立了稳定沉默VEGF细胞CNE-LV3。划痕实验和Transwell小室迁移实验显示,与空白对照组和CNE-NC组相比,CNE-LV3组细胞迁移率明显降低[(50.17±1.76)%vs(37.97±1.56)%,P=0.001;(50.63±1.52)%vs(37.97±1.56)%,P=0.001],穿膜细胞数亦明显下降[(95.3±3.8)个vs(41.3±1.5)个,P<0.001;(94.3±4.0)个vs(41.3±1.5)个,P<0.001];Transwell小室侵袭实验结果亦显示,CNE-LV3组细胞穿膜细胞数较空白对照组和CNE-NC组明显下降[(46.3±2.1)个vs(105.3±1.5)个,P<1.001;(46.3±2.1)个vs(93.3±2.5)个,P<0.001]。结论沉默VEGF表达可抑制人鼻咽癌放射抗拒细胞株CNE-2R的迁移和侵袭能力。

Objective To investigate the effect of silencing the gene of VEGF on the migration and invasion of radioresistant human nasopharyngeal carcinoma cell line CNE-2R.Methods Three groups of lentiviral vectors containing VEGF-shRNA(VEGF-LV1,VEGF-LV2,VEGF-LV3)and control shRNA recombinant lentivirus(VEGF-NC)were prepared.Cells of radio resistant human nasopharyngeal carcinoma CNE-2R were transfected to establish a cell line stably silencing VEGF expression(CNE-LV1,CNE-LV2,and CNE-LV3),and a negative control group(NE-NC)was established.RT-qPCR and Western blot were used to detect the infection effect of each group.Scratch test and Transwell chamber assay were used to detect cell migration and invasion after VEGF silencing.Results VEGF-shRNA recombinant lentivirus were successfully constructed and the stable subcellular cell line(CNE-LV3)with VEGF silencing was established. The scratch test showed that the cell migration rate of the CNE-LV3 group was significantly lower than that of the blank control group and the CNE-NC group[(37.97±1.56)% vs (50.17±1.76)%,P=0.001;(37.97±1.56)% vs (50.63±1.52)%,P=0.001]. Transwell chamber migration assay showed that the number of transmembrane cells in the CNE-LV3 group was significantly decreased compared with the blank control group and CNE-NC group(41.3±1.5 vs 95.3±3.8,P<0.001;41.3±1.5 vs 94.3±4.0,P<0.001). And Transwell chamber invasion assay also showed that the number of transmembrane cells in the CNE-LV3 group was significantly decreased compared with the blank control group and CNE-NC group(46.3±2.1 vs 105.3±1.5,P<0.001;46.3±2.1 vs 93.3±2.5,P<0.001). Conclusion Silencing VEGF expression can inhibit the migration and invasion of radioresistant human nasopharyngeal carcinoma cell line CNE-2R.