S期激酶相关蛋白2基因沉默对人脑胶质瘤细胞U251生物学特性的影响  

Effect of S-phase kinase associated protein 2 gene silencing on the biological characteristics of human glioma U251 cells

作　　者：刘岳[1] 李彩丽[1] 曲新国[1] 吕伟[1] 林毅[1] 唐振刚[1] 

机构地区：[1]湖北医药学院附属人民医院神经外科,十堰442000

出　　处：《中华实验外科杂志》2017年第8期1274-1277,共4页Chinese Journal of Experimental Surgery

基　　金：湖北省教育厅科学研究计划（B2015171）

摘　　要：目的 观察S期激酶相关蛋白2（SKP2）基因沉默对人脑胶质瘤细胞U251生物学特性的影响,探讨其可能的机制.方法 体外培养U251细胞,分为3组,分别加入Ad-shSKP2（shSKP2组）、Ad-shNC（shNC组）及磷酸盐缓冲液（PBS,对照组）,采取细胞计数试剂盒（CCK-8）法检测细胞增殖的变化,流式细胞术检测细胞周期和凋亡的变化,划痕实验和Transwell侵袭实验观察细胞迁移和侵袭的变化,Western blot测定U251细胞B细胞淋巴瘤/白血病-2相关X蛋白（bax）、B细胞淋巴瘤/白血病-2（bcl-2）、基质金属蛋白酶（MMP）-9、MMP-2的蛋白表达的变化.结果 Ad-shSKP2转染24、48、72h后shSKP2组的细胞存活率分别是对照组的（87.17±2.60）%、（79.63±1.74）%和（72.81±1.78）%,而shNC组的细胞存活率无显著性变化.shSKP2组、shNC组及对照组的凋亡率分别为（18.27±2.44）%、（1.51±1.02）%和（1.11±1.20）%,G2/M期比例分别为（26.65±2.51）%、（14.46±2.58）%和（14.89±3.81）%.shNC组和shSKP2组的24h迁移距离分别为对照组的（93.45±16.77）%及（43.69±7.44）%,对照组、shNC组和shSKP2组24h通过Transwell小室的过膜细胞数（每200倍视野）分别为（87.50±5.66）、（79.78±8.03）、（38.24±6.64）个.Western blot显示,shSKP2组较对照组及shNC组bax表达上调,bcl-2、MMP-9、MMP-2蛋白表达下降.结论Ad-shSKP2可抑制U251细胞的体外增殖,可诱导U251细胞凋亡、引起G2/M期阻滞,减弱细胞的迁移及侵袭能力.Ad-shSKP2可使U251细胞bcl-2/bax比值的降低以及MMP-9和MMP-2蛋白的表达降低.Objective To observe the effect of S-phase kinase associated protein 2 （SKP2） gene silencing on the biological characteristics of human glioma U251 cells in vitro.Methods The U251 cells were treated with Ad-shSKP2 （shSKP2 group）,Ad-shNC （shNC group） and PBS （control group） respectively.The proliferation of U251 was evaluated by cell counting kit-8 （CCK-8） assay,cell cycle and apoptosis were examined by flow cytometry,the invasion and metastasis of U251 cells were examined by wound scratch assay and Transwell assay respectively,and the protein levels of B cell lymphoma/leukemia-2 associated X protein （bax）,B cell lymphoma/leukemia-2 （bcl-2）,matrix metalloproteinase （MMP）-9 and MMP-2 were detected by Western blotting.Results The cell viability of shSKP2 group was （87.17±2.60）%,（79.63±1.74）% and （72.81±1.78）% of that of control group respectively 24,48 and 72 h after Ad-shSKP2 transfection.The cell viability of the shNC group was not significantly different before and after transfection.The apoptosis rate of shSKP2 group,shNC group and control group was （18.27±2.44）%,（1.51±1.02）% and （1.11±1.20）%,and the ratio of G2/M phase was （26.65±2.51）%,（14.46±2.58）% and （14.89±3.81）%,respectively.The migration distances of shNC group and shSKP2 group were （93.45±16.77）% and （43.69±7.44）% of those of control group respectively.The number of transmembrane cells （per 200-fold field of view） passing through the Transwell chamber of the control group,shNC group and shSKP2 group was （87.50±5.66）%,（79.78±8.03）% and （38.24±6.64）% respectively.The expression of bax,bcl-2,MMP-9 and MMP-2 protein in shSKP2 group was down-regulated as compared with shNCK group and control group.ConclusionThe results in this study implied that SKP2 gene silencing inhibited the proliferation of U251 cells,induced cell apoptosis and G2/M phase arrest,and attenuated the ability of invasion and metastasis of U251 cells.SKP2 gene silencing can decrease the

关 键 词：神经胶质瘤 S期激酶相关蛋白2 脱噬作用 侵袭 迁移 

分 类 号：R739.41[医药卫生—肿瘤]