血管内皮生长因子mRNA干扰提高胶质母细胞瘤U251细胞对放化疗敏感性的研究  

Knockdown of vascular endothelial cell growth factor and sensitivity of U251 cells to radiotherapy and chemotherapy

作　　者：俞洋[1] 宗祥云[1] 冯建国[1] 吴斌[1] 

机构地区：[1]浙江省肿瘤医院脑外科,杭州310022

出　　处：《中华实验外科杂志》2013年第4期718-721,共4页Chinese Journal of Experimental Surgery

摘　　要：目的探讨血管内皮生长因子（VEGF） mRNA干扰对胶质母细胞瘤的治疗意义。方-法应用RNA干扰技术，对胶质母细胞瘤细胞株U251进行了 VEGF mRNA干扰、辅以6 Gy的放射剂量照射、化疗处理。在干扰前后用流式细胞技术、噻唑蓝（MTT）比色法及倒置显微镜观察等技术对U251细胞株的细胞周期、凋亡率、抑制率、细胞形态进行检测。结果胶质母细胞瘤U251细胞在转染VEGF小干扰RNA（siRNA）后，逆转录-聚合酶链反应（HT-PCR）检测发现VEGF mRNA的表达显著下降，均数下调大于60% ;流式细胞技术显示，U251转染VEGF siRNA后细胞出现G0-G1阻滞，G2 ＋M期细胞数目减少，转染细胞出现不同程度凋亡;MTT比色法可见，转染前不同浓度的紫杉醇对U251细胞有一定的抑制，作用48 h半数抑制剂量（IC50）=28. 1g/L0干扰后，紫杉醇在不同浓度对U251细胞的抑制率都有大幅度的提高，IC50=0. 02 g/L;集落形成抑制实验中，单药物组与单放疗组在克隆形成率上差异无统计学意义（p〉0.05） ,U251细胞转染VEGF siRNA后，明显抑制肿瘤细胞的集落形成，并对紫杉醇及放射治疗具有明显的协同作用;细胞学形态变化观察发现，转染后实验组的细胞形态明显差于未转染组细胞。结论VEGF基因干扰可以抑制U251细胞的增殖、促进其凋亡，并能提高U251细胞对放化疗的敏感性。脂质体紫杉醇在细胞实验中表现出较好的抑制肿瘤作用及对VEGF基因干扰和放疗的协同作用。Objective To explore the effect of vascular endothelial growth factor （ VEGF） mRNAinterference on glioblatoma. Methods After knockdown of VEGF mRNA expression using VEGF shorthairpin RNA （ shRNA）,glioma U251 cell were treated with chemotherapy or radiotherapy or chemotherapyplus radiotherapy or without any treatment. The changes in cell cycle, apoptosis rate,cell colony-formationand cell morphology were observed. Results After knockdown of VEGF mRNA expression using VEGFshRNA, VEGF mRNA expression levels in glioma U251 cells were inhibited significantly （ P 〈 0. 001）,The flow cytometry revealed that both control cells and VEGF shRNA-transfected cells exhibited G0-Gj ar-rest, and reduced number of cells in the G2 and M phases. The apoptosis rate of VEGF shRNA-transfectedcells was increased as compared with the control cells. It was found that various concentrations of paciltaxelreduced U251 cell viability 50% inhibitory dose （ IC50 ） = 28. 1 mg/L, and VEGF knockdown significantlysensitized U251 cells to paclitaxel （IC50 = 0. 02 mg/L） . Groups with drug treatment alone and the radiother-apy alone showed no significant difference （P 〉 0. 05 ）. After VEGF knockdown,colony formation in paclita-xel and radiation treated U251 cells was significantly reduced （ 17. 57% to 6. 33% 〈 0. 01 ）. Under themicroscopy, it was found VEGF shRNA-transfected cells showed worse cellular morphology than non-trans-fected cells. Conclusion The interference of VEGF gene expression can inhibit the proliferation of U251cells, promote apoptosis of U251 cells, and increase the sensitivity of U251 cells to chemotherapy and radio-therapy. Liposomal paclitaxel can enhance drug delivery in vivo,although the VEGF gene interference andthe practical application of liposomal paclitaxel remains to be tested and explored in future studies.

关 键 词：胶质母细胞瘤 血管内皮生长因子 RNA干扰 放射治疗 化学治疗 

分 类 号：R73[医药卫生—肿瘤]