中华胃肠外科杂志
中華胃腸外科雜誌
중화위장외과잡지
CHINESE JOURNAL OF GASTROINTESTINAL SURGERY
2012年
11期
1182-1186
,共5页
贺孝文%陶峰%骆珊珊%余小科
賀孝文%陶峰%駱珊珊%餘小科
하효문%도봉%락산산%여소과
结肠肿瘤%RNA干扰%增殖%凋亡
結腸腫瘤%RNA榦擾%增殖%凋亡
결장종류%RNA간우%증식%조망
Colonic neoplasms%RNA interference%Proliferation%Apoptosis
目的 探讨RNA干扰技术沉默人NOB1基因对人结肠癌RKO细胞增殖和凋亡的影响.方法 设计靶向NOB1基因的小干扰RNA(siRNA),构建NOB1基因特异性的RNA干扰慢病毒载体,感染RKO细胞,并设置阴性对照组.实时定量PCR和Western blot分别检测两组细胞NOB1 mRNA及蛋白的表达情况;Cellomics ArrayScan VTI HCS高内涵分析仪检测感染细胞的增殖和克隆形成能力;流式细胞术检测细胞周期及凋亡的变化;裸鼠成瘤实验检测NOB1-siRNA对肿瘤的抑制作用.结果 NOB1-shRNA慢病毒感染RKO细胞3d后,与阴性对照组比较,NOB1 mRNA和蛋白的表达明显降低,克隆形成数减少,而细胞凋亡数增加(均P＜0.05).裸鼠成瘤实验结果显示,NOB1-shRNA感染组裸鼠移植瘤体积为(405±102)mm3,小于阴性对照组的(870±165)mm3(P＜0.05).结论 通过RNA干扰方式沉默NOB1基因的表达,有望作为抑制结肠癌发展的有效手段.
目的 探討RNA榦擾技術沉默人NOB1基因對人結腸癌RKO細胞增殖和凋亡的影響.方法 設計靶嚮NOB1基因的小榦擾RNA(siRNA),構建NOB1基因特異性的RNA榦擾慢病毒載體,感染RKO細胞,併設置陰性對照組.實時定量PCR和Western blot分彆檢測兩組細胞NOB1 mRNA及蛋白的錶達情況;Cellomics ArrayScan VTI HCS高內涵分析儀檢測感染細胞的增殖和剋隆形成能力;流式細胞術檢測細胞週期及凋亡的變化;裸鼠成瘤實驗檢測NOB1-siRNA對腫瘤的抑製作用.結果 NOB1-shRNA慢病毒感染RKO細胞3d後,與陰性對照組比較,NOB1 mRNA和蛋白的錶達明顯降低,剋隆形成數減少,而細胞凋亡數增加(均P＜0.05).裸鼠成瘤實驗結果顯示,NOB1-shRNA感染組裸鼠移植瘤體積為(405±102)mm3,小于陰性對照組的(870±165)mm3(P＜0.05).結論 通過RNA榦擾方式沉默NOB1基因的錶達,有望作為抑製結腸癌髮展的有效手段.
목적 탐토RNA간우기술침묵인NOB1기인대인결장암RKO세포증식화조망적영향.방법 설계파향NOB1기인적소간우RNA(siRNA),구건NOB1기인특이성적RNA간우만병독재체,감염RKO세포,병설치음성대조조.실시정량PCR화Western blot분별검측량조세포NOB1 mRNA급단백적표체정황;Cellomics ArrayScan VTI HCS고내함분석의검측감염세포적증식화극륭형성능력;류식세포술검측세포주기급조망적변화;라서성류실험검측NOB1-siRNA대종류적억제작용.결과 NOB1-shRNA만병독감염RKO세포3d후,여음성대조조비교,NOB1 mRNA화단백적표체명현강저,극륭형성수감소,이세포조망수증가(균P＜0.05).라서성류실험결과현시,NOB1-shRNA감염조라서이식류체적위(405±102)mm3,소우음성대조조적(870±165)mm3(P＜0.05).결론 통과RNA간우방식침묵NOB1기인적표체,유망작위억제결장암발전적유효수단.
Objective To investigate the effect of NOB1 gene on proliferation and apoptosis of human colon cancer cell line RKO by RNA interference.Methods Small interference RNA(siRNA)targeting NOB1 gene was cloned into lentivirus vector.Then the lentivirus particles expressing NOB1 short harpin RNA(shRNA)were infected into RKO cells.Real-time PCR and Western blot were performed to examine the expression of NOB1 in lentivirus infected cells.The Thermo Scientific Cellomics ArrayScan VTI HCS Reader was used to test the proliferation and colony-formation of RKO cells,and flow cytometry assay was performed to detect cell cycle and apoptosis.Xenograft tumor was established by injection of RKO cells into nude mice,then NOB1-shRNA was injected into the tumor and tumor volume was detected.Results Compared to negative controls,the expression levels of NOB1 mRNA and protein were both significantly down-regulated,the proliferation and colony-forming capacity of RKO cells were significantly inhibited,and cell apoptosis was increased after 3 days of NOB1-shRNA lentivirus infection(all P＜0.05).The tumor volume was significantly smaller in NOB1-shRNA group than that in Scr-shRNA group[(405±102)mm3 vs.(870±165)mm3,P＜0.05].Conclusion Silencing NOB1 gene by RNA interference may provide an inhibitive effect on human colon cancer development.
