靶向抑制KIAA1199基因的表达对人结肠癌细胞SW620生长的影响  

The effects of target inhibiting KIAA1199 on the growth of human colorectal cancer cell line SW620

作　　者：董晴晴[1] 许慈[1] 蔡懿婷[1] 王碧君[1] 戴强[1] 朱黎明[1] 

机构地区：[1]上海交通大学医学院附属第九人民医院消化科,上海201999

出　　处：《热带医学杂志》2017年第4期456-460,494,F0003,共7页Journal of Tropical Medicine

基　　金：上海交通大学医学院基金项目(14XJ10066);上海市宝山区科委科研基金项目(13-E-2)

摘　　要：目的探讨靶向沉默KIAA1199基因的表达对人结肠癌SW620细胞生长能力的影响。方法运用RNA干扰(shRNA)技术构建携带靶向抑制KIAA1199分子的慢病毒载体LV3-KIAA1199-shRNA并感染人结肠癌细胞株SW620为实验组(shKIAA1199),携带无义序列的对照慢病毒载体LV3-NC-shRNA为阴性对照组(shNC),未转染的细胞为空白对照组(MOCK)。72 h后用荧光显微镜观察慢病毒的感染效率,RT-PCR法验证慢病毒感染后KIAA1199 mRNA在MOCK、shNC和shKIAA1199各组细胞中的表达水平,并用嘌呤霉素筛选KIAA1199静默和阴性对照的稳转细胞。用MTT法检测MOCK、shNC和shKIAA1199各组细胞活力,流式细胞仪分别检测三组细胞的凋亡率和细胞周期的变化。结果携带靶向干扰KIAA1199基因的慢病毒感染人结肠癌细胞株SW620后,能有效下调KIAA1199在细胞内的表达水平并成功筛选KIAA1199静默的SW620稳转细胞;与MOCK和shNC组比较,shKIAA1199组细胞活力减少(P<0.05),细胞凋亡率明显升高(P<0.05),处于细胞周期中G1期的细胞数量比例明显下降(P<0.05),G2/M期细胞比例增加(P<0.05)。结论下调KIAA1199基因的表达可抑制SW620细胞的增殖,促进该细胞的凋亡,使该肿瘤细胞的生长周期受到阻滞。Objective To study the effect of KIAA1199 silencing on the growth of human colorectal cancer cell line SW620 in vitro. Methods RNAi targeting by KIAA1199 was transfected into lentivirus carrier 3（LV3-KIAA1199-shRNA）and then the transfected virus carrier was used to infect human colorectal cancer cell line SW620 for 72 hours（shKIAA1199group）. Other two groups including uninfected cells（MOCK group）and lenti-virus carrier 3 containing nonsense shRNA sequence（LV3-NC-shRNA）infected cells（shNC group）. The efficiency of infection was observed by fluorescence microscope.The mRNA expression of KIAA1199 was determined by reverse transcriptase polymerase chain reaction（RT-PCR）. The stable cells of KIAA1199 silencing expression and NC by lentivirus-mediated RNA interference were selected by puromycin.The cell viability was measured by MTT assay;cell apoptosis and cell cycle were determined by flow cytometry in MOCK,shNC and shKIAA1199 groups. Results Compared with MOCK and shNC groups,the mRNA expression level of KIAA1199 was downregulated significantly in shKIAA1199 group in SW620 cells after LV3-KIAA1199-shRNA lentivirus vector infection for 72 hours. The stable cells of KIAA1199 silencing expression and NC were selected. The cell viability was decreased（P〈0.05）and cell apoptosis was significantly increased（P〈0.05）in shKIAA1199 group compared with shNC and MOCK group in SW620 cells. The cell percentage of G1 phase was reduced（P〈0.05）and the cell percentage of G2/M phase was elevated（P〈0.05） in shKIAA1199 group compared with MOCK and shNC group. Conclusions Silencing KIAA1199 expression could effectively inhibit the proliferation of colorectal cancer cells and induce cell apoptosis in SW620 cells. Inhibition the expression of KIAA1199 also induced cell cycle blockage in G2/M phase.

关 键 词：KIAA1199 RNA干扰 结肠肿瘤 增殖 凋亡 

分 类 号：R735.35[医药卫生—肿瘤]