靶向FOXP3基因的短发夹RNA对人胶质瘤细胞增殖和凋亡的影响  

Effects of short hairpin RNA targeting FOXP3 gene on proliferation and apoptosis of glioma cells

作　　者：豆玉超 张飚[2] 王雷波 杜春发 徐新女[4] 张述生 尚超 王琼 冯学泉[4] 王金环 

机构地区：[1]天津医科大学研究生院,300070 [2]天津市环湖医院检验科 [3]天津市神经外科研究所天津市脑血管与神经变性病重点实验室 [4]天津市第一中心医院

出　　处：《中华神经外科杂志》2015年第3期289-293,共5页Chinese Journal of Neurosurgery

基　　金：国家自然科学基金（81101911）,天津市应用基础及前沿技术研究计划（11JCYBJC12100）;天津市抗癌重大专项计划（12ZDZSY17700）;天津市卫生局科技计划项目（11KG115）;国家临床重点专科建设专项基金资助（2011-327）

摘　　要：目的 探讨叉头状螺旋转录因子3基因（Forkhead box P3,FOXP3）的短发夹RNA（shRNA）对人胶质瘤细胞系U87和LN229增殖能力及凋亡的影响.方法 用脂质体法将4个含有靶向FOXP3的shRNA序列（shRNA1 ～4）和1个含有shRNA无意义随机阴性对照序列（neg-shRNA）的质粒转染入人胶质瘤U87细胞和LN229细胞;应用Western blot检测转染后FOXP3蛋白的表达情况筛选出最有效的一个shRNA干扰质粒;应用流式细胞术、CCK-8法检测转染后U87细胞和LN229细胞的凋亡和增殖活性的变化.结果 转染72 h后,Western blot结果显示空白对照组、shRNA-1、shRNA-2、shRNA-3、shRNA-4组和neg-shRNA组中FOXP3蛋白的表达受抑制最明显的是shRNA-1组.CCK-8结果显示转染72 h后,shRNA-1转染组U87细胞和LN229细胞增殖活性分别为（122.00±4.32）％、（118.36±2.49）％,与neg-shRNA组（98.55±4.34）％、（99.87±2.17）％和空白对照组100％相比均显著升高（P＜0.05）;流式细胞术结果显示转染72 h后,shRNA-1转染的U87细胞和LN229细胞凋亡率分别为（7.03±3.36）％和（9.40±2.51）％,neg-shRNA转染U87细胞和LN229细胞的凋亡率分别为（17.70±4.39）％和（22.63±1.86）％,在U87和LN229细胞中空白对照组凋亡率分别为（16.57±2.30）％和（21.67±1.93）％,两种细胞的凋亡率在shRNA-1组与neg-shRNA组和空白对照组比较差异均有统计学意义（P＜0.05）.结论 靶向FOXP3基因的shRNA能够有效抑制该基因表达,抑制胶质瘤细胞的凋亡,促进细胞增殖.Objective To investigate the effects of short hairpin RNA （shRNA） targeting Forkhead box P3 （FOXP3） gene on the proliferation and apoptosis of human glioma cells U87 and LN229.Methods 4 plasmids containing different sequences of shRNA targeting FOXP3 gene and 1 plasmids containing random sequences were transfected into human glioma U87 cells and LN229 cells mediated by Lipofectamine 2000.The expression of FOXP3 protein was detected by Western blot to screen the most effective shRNA plasmid.The cell proliferation and apoptosis were detected by CCK-8 assay and flow cytometry,respectively.Results At 72 h after transfection,the expression of FOXP3 protein was most obviously suppressed by shRNA-1 sequence compared with other shRNA sequences.The CCK-8 assay results showed that the proliferation rates of U87 cell and LN229 cell tranfected with shRNA-1 plasmid were （122.00 ±4.32）% and （118.36 ±2.49）% at 72 h,respectively,which were significantly higher than that transfected with neg-shRNA plasmid （neg-shRNA group） and the non-transfection group （control group） （P 〈0.05）.At 72 h after transfection,the apoptosis rates of U87 cell and LN229 cell in shRNA-1 group were （7.03 ± 3.36）% and （9.40 ± 2.51）%,respectively,which were decreased significantly compared with those of the neg-shRNA group （17.70 ± 4.39）% and （22.63 ± 1.86）%,and the control group （16.57 ±2.30）% and（21.67 ± 1.93）% （P 〈 0.05）.Conclusions The shRNA targeting FOXP3 gene could efficiently inhibit the gene expression and hence it could significantly inhibit the apoptosis of glioma cells and promote cell proliferation.

关 键 词：胶质瘤 叉头状螺旋转录因子3 凋亡 增殖 

分 类 号：R739.41[医药卫生—肿瘤]