慢病毒载体介导RNA干扰抑制Livin对人大肠癌HT-29细胞生物学行为的影响  

Inhibition of Livin Expression on Colon Cancer Line HT-29 Cells by Lentiviral Vector-Mediated RNA Interference and Its Impact on Biological Behaviors

作　　者：余娜莎[1] 何文静[2] 黎军和[2] 魏芹[2] 彭小东[2] 

机构地区：[1]江西省肿瘤医院内二科,南昌市330029 [2]南昌大学第一附属医院肿瘤科

出　　处：《中国肿瘤临床》2012年第15期1060-1064,共5页Chinese Journal of Clinical Oncology

基　　金：国家自然科学基金(编号:30960440)资助~~

摘　　要：目的:探讨慢病毒载体介导RNA干扰(RNAi)抑制大肠癌细胞系HT-29细胞Livin基因表达对其生物学行为的影响方法:设计、合成靶向Livin的siRNA,构建pGCSIL-GFP慢病毒载体,采用Real-time PCR和Western blot方法观察对人大肠癌HT-29细胞Livin mRNA及蛋白质表达的抑制;并通过台盼蓝拒染法、原位末端标记法(TUNEL)、流式细胞术及Matrigel穿膜侵袭实验检测细胞增殖、凋亡、细胞周期及细胞侵袭性的变化。结果:重组慢病毒Livin-RNAi-LV1能够有效抑制Livin的表达,使LivinmRNA和蛋白表达水平明显降低(P<0.01);Livin沉默能抑制HT-29细胞增殖(P<0.05),细胞凋亡增加(P<0.01),细胞周期重新分布,G_0/G_1期细胞比例升高(P<0.05),S期细胞比例降低(P<0.05),G_2/M期细胞比例降低(P<0.05) HT-29细胞体外侵袭能力也明显减弱(P<0.01)。结论:利用慢病毒载体介导RNA干扰技术沉默Livin基因的表达可以抑制大肠癌HT-29细胞增殖,促进细胞凋亡,使细胞侵袭性减弱。Objective： To investigate the inhibition of Livin expression on colon cancer cell line HT-29 by lentiviral vector-medi- ated RNA interference （ RNAi ） and its impact on biological behaviors. Method： Small-interference RNA （ siRNA ） molecules that tar- get Livin gene were designed, and a pGCSIL-GFP lentiviral vector was created. The inhibitory expression of Livin was detected using Real-time PCR and Western blot. Trypan blue staining was used to detect changes in proliferation. In situ end labeling （ TUNEL ） and flow cytometry were used to detect changes in apoptosis and cell cycle distribution, respectively. Matrigel invasion assay was used to detect changes in cell invasion ability. Results： Recombinant lentiviral Livin-RNAi-LV1 can efficiently inhibit Livin expression in mRNA and protein level （ P 〈 0.01 ）. Reduced Livin expression can inhibit cell proliferation （ P 〈 0.05 ）, promote cell apoptosis （ P 〈 0.01 ）, and re-distribute the cell cycle, leading to increased proportion of G0/G~ phase cells （ P 〈 0.05 ） and reduced S phase cells （ P 〈 0.05 ） and G2/M phase cells 〈 0.05 ）. In vitro invasive ability was significantly damaged （ P 〈 0.01 ）. Conclusion： The inhibition of Liv- in expression on HT-29 cell by Lentiviral vector-mediated RNA interference can inhibit proliferation, promote apoptosis, and reduce cell invasiveness; thus, it deserves further study.

关 键 词：慢病毒载体 RNA干扰 LIVIN 大肠癌 凋亡 

分 类 号：R735.34[医药卫生—肿瘤]