中华普通外科杂志
中華普通外科雜誌
중화보통외과잡지
CHINESE JOURNAL OF GENERAL SURGERY
2011年
10期
856-859
,共4页
唐志强%姚磊%金光鑫%吴德全
唐誌彊%姚磊%金光鑫%吳德全
당지강%요뢰%금광흠%오덕전
癌,肝细胞%RNA,小分子干扰%动物实验%微小RNA-21
癌,肝細胞%RNA,小分子榦擾%動物實驗%微小RNA-21
암,간세포%RNA,소분자간우%동물실험%미소RNA-21
Carcinoma,hepatocellular%RNA,small interfering%Animal experimentation%MicroRNA-21
目的 通过构建慢病毒介导( lentivirus,LV)的microRNA-21-siRNA,探讨下调microRNA-21对肝癌细胞系HepG2生物学行为的影响.方法 针对目标序列合成特异性siRNA干扰片段并克隆入慢病毒载体pGCSIL-GFP,将重组microRNA-21-RNAi-LV转染肝细胞癌HepG2.体外设为干扰组(microRNA-21 -siRNA,SI组)、阴性对照组(negative control,NC组)和正常组(normal,N组)；分别采用聚合酶链反应方法检测microRNA-21目的基因的表达的情况；用噻唑蓝比色法检测增殖情况、transwell小室迁移实验检测迁移情况、Hochest33258凋亡实验检测凋亡情况.体内实验设为SI组和NC组,观察反义microRNA-21对裸鼠移植瘤生长的影响.结果 (1)micmRNA-21 -RNAi-LV可显著降低microRNA-21的表达.(2) MTT实验96h后SI组增殖能力显著低于NC组和N组(P=0.0031,P＜0.05).(3)迁移能力减弱(P =0.0004,P＜0.05).(4)SI组细胞凋亡明显增加,促凋亡基因caspase3基因表达上调(P =0.0002,P＜0.05).(5)各组裸鼠皮下肿瘤生长比较差异无统计学意义(P=0.0624,P＞0.05).结论 microRNA-21 -siRNA通过抑制microRNA-21的表达抑制细胞增殖,促进细胞凋亡,降低其迁移能力.
目的 通過構建慢病毒介導( lentivirus,LV)的microRNA-21-siRNA,探討下調microRNA-21對肝癌細胞繫HepG2生物學行為的影響.方法 針對目標序列閤成特異性siRNA榦擾片段併剋隆入慢病毒載體pGCSIL-GFP,將重組microRNA-21-RNAi-LV轉染肝細胞癌HepG2.體外設為榦擾組(microRNA-21 -siRNA,SI組)、陰性對照組(negative control,NC組)和正常組(normal,N組)；分彆採用聚閤酶鏈反應方法檢測microRNA-21目的基因的錶達的情況；用噻唑藍比色法檢測增殖情況、transwell小室遷移實驗檢測遷移情況、Hochest33258凋亡實驗檢測凋亡情況.體內實驗設為SI組和NC組,觀察反義microRNA-21對裸鼠移植瘤生長的影響.結果 (1)micmRNA-21 -RNAi-LV可顯著降低microRNA-21的錶達.(2) MTT實驗96h後SI組增殖能力顯著低于NC組和N組(P=0.0031,P＜0.05).(3)遷移能力減弱(P =0.0004,P＜0.05).(4)SI組細胞凋亡明顯增加,促凋亡基因caspase3基因錶達上調(P =0.0002,P＜0.05).(5)各組裸鼠皮下腫瘤生長比較差異無統計學意義(P=0.0624,P＞0.05).結論 microRNA-21 -siRNA通過抑製microRNA-21的錶達抑製細胞增殖,促進細胞凋亡,降低其遷移能力.
목적 통과구건만병독개도( lentivirus,LV)적microRNA-21-siRNA,탐토하조microRNA-21대간암세포계HepG2생물학행위적영향.방법 침대목표서렬합성특이성siRNA간우편단병극륭입만병독재체pGCSIL-GFP,장중조microRNA-21-RNAi-LV전염간세포암HepG2.체외설위간우조(microRNA-21 -siRNA,SI조)、음성대조조(negative control,NC조)화정상조(normal,N조)；분별채용취합매련반응방법검측microRNA-21목적기인적표체적정황；용새서람비색법검측증식정황、transwell소실천이실험검측천이정황、Hochest33258조망실험검측조망정황.체내실험설위SI조화NC조,관찰반의microRNA-21대라서이식류생장적영향.결과 (1)micmRNA-21 -RNAi-LV가현저강저microRNA-21적표체.(2) MTT실험96h후SI조증식능력현저저우NC조화N조(P=0.0031,P＜0.05).(3)천이능력감약(P =0.0004,P＜0.05).(4)SI조세포조망명현증가,촉조망기인caspase3기인표체상조(P =0.0002,P＜0.05).(5)각조라서피하종류생장비교차이무통계학의의(P=0.0624,P＞0.05).결론 microRNA-21 -siRNA통과억제microRNA-21적표체억제세포증식,촉진세포조망,강저기천이능력.
Objective To study the effect of lentivirus-mediated microRNA-21 RNAi on biological behaviors in human hepatic cancer cells.Methods MicroRNA-21 specific siRNA gene was synthesized and cloned into the recombinant lentiviral vector,pGCSIL-GFP.HepG2 cells were infected by microRNA-21 siRNA recombinant lentivirus (miR-21-siRNA-Ⅳ).The HepG2 cells were devided into SI group,NC group and N group in vitro.The expression of the targets of miR-21 was detected by RT-PCR.Cell growth was analyzed by MTT assay.The invasion was dectected by tmnswell method.Apoptosis was detected by Hoechst33258.BALB/c nude mice were randomly divided into SI group and NC group.The growth of transplant tumors in BALB/c nude mice were observed.Results ( 1 ) The expression level of miR-21 was inhibited significantly by miR-21-siRNA-lⅣ.(2) The proliferation of HepG2 was also markedly suppressed in MTT at the 96 h point ( P =0.0031,P ＜ 0.05 ).(3) The number of cells that migrated through the chamber of SI group decreased ( P =0.0004,P ＜ 0.05 ).(4) The cell apopotosis in SI group increased markedly.In addition,the caspase 3 mRNA significantly increased ( P =0.0002,P ＜ 0.05 ).( 5 ) Tumor growth curve was not statistically different between groups ( P =0.0002,P ＜ 0.05 ).Conclusions MicroRNA-21 specific siRNA suppresses the proliferation and migration of HepG2 cells and induces tumor cell apoptosis inhibiting the growth of transplanted tumor in Balb/c nude mice.
