中华实验外科杂志
中華實驗外科雜誌
중화실험외과잡지
CHINESE JOURNAL OF EXPERIMENTAL SURGERY
2013年
4期
667-670
,共4页
胡凡果%只向成%史玉荣%牛瑞芳%刘彤
鬍凡果%隻嚮成%史玉榮%牛瑞芳%劉彤
호범과%지향성%사옥영%우서방%류동
乳腺癌%端粒酶%RNA 干扰%增殖
乳腺癌%耑粒酶%RNA 榦擾%增殖
유선암%단립매%RNA 간우%증식
Breast carcinoma%Telomerase%RNA interference%Proliferation
目的 观察针对人端粒酶逆转录酶(hTERT)的短发夹RNA (shRNA)质粒表达载体对乳腺癌T47D细胞端粒酶活性及增殖能力的影响.方法 将针对hTERT的5种shRNA质粒表达载 体并转染到乳腺癌T47D细胞,采用逆转录-聚合酶链反应(RT-PCR)及Western blot法检测hTERT在mRNA和蛋白水平上的表达；抗酒石酸酸性磷酸酶-酶联免疫吸附试验(TRAP-ELISA)法检测细胞的端粒酶活性变化；噻唑蓝(MTT)法绘制细胞生长曲线,流式细胞仪结合碘化丙锭(PI)染色检测细胞周期分布.结果 同对照组T47D细胞比较,小干扰RNA(siRNA)1组和阴性对照siRNA2-N组hTERT基因在mRNA和蛋白水平的表达以及细胞端粒酶活性的变化差异无统计学意义(P＞0.05),而siRNA2组分别下降57.1％、57.0％和57.0(P＜0.01),siRNA3组分别下降53.7％、53.4％和56.5％ (P＜0.01),siRNA4组分别下降70.0％、70.3％和81.9％ (P＜0.01)；siRNA1、siRNA2-N组细胞生长速度及各期细胞比例差异无统计学意义(P＞0.05),而siRNA2、siRNA3和siRNA4组细胞自24h起就出现生长速度明显降低(P＜0.01),G0/G1期细胞比例增高(P＜0.01)和S期细胞比例降低(P＜0.01).结论 针对hTERT的shRNA可抑制肿瘤细胞的增殖.
目的 觀察針對人耑粒酶逆轉錄酶(hTERT)的短髮夾RNA (shRNA)質粒錶達載體對乳腺癌T47D細胞耑粒酶活性及增殖能力的影響.方法 將針對hTERT的5種shRNA質粒錶達載 體併轉染到乳腺癌T47D細胞,採用逆轉錄-聚閤酶鏈反應(RT-PCR)及Western blot法檢測hTERT在mRNA和蛋白水平上的錶達；抗酒石痠痠性燐痠酶-酶聯免疫吸附試驗(TRAP-ELISA)法檢測細胞的耑粒酶活性變化；噻唑藍(MTT)法繪製細胞生長麯線,流式細胞儀結閤碘化丙錠(PI)染色檢測細胞週期分佈.結果 同對照組T47D細胞比較,小榦擾RNA(siRNA)1組和陰性對照siRNA2-N組hTERT基因在mRNA和蛋白水平的錶達以及細胞耑粒酶活性的變化差異無統計學意義(P＞0.05),而siRNA2組分彆下降57.1％、57.0％和57.0(P＜0.01),siRNA3組分彆下降53.7％、53.4％和56.5％ (P＜0.01),siRNA4組分彆下降70.0％、70.3％和81.9％ (P＜0.01)；siRNA1、siRNA2-N組細胞生長速度及各期細胞比例差異無統計學意義(P＞0.05),而siRNA2、siRNA3和siRNA4組細胞自24h起就齣現生長速度明顯降低(P＜0.01),G0/G1期細胞比例增高(P＜0.01)和S期細胞比例降低(P＜0.01).結論 針對hTERT的shRNA可抑製腫瘤細胞的增殖.
목적 관찰침대인단립매역전록매(hTERT)적단발협RNA (shRNA)질립표체재체대유선암T47D세포단립매활성급증식능력적영향.방법 장침대hTERT적5충shRNA질립표체재 체병전염도유선암T47D세포,채용역전록-취합매련반응(RT-PCR)급Western blot법검측hTERT재mRNA화단백수평상적표체；항주석산산성린산매-매련면역흡부시험(TRAP-ELISA)법검측세포적단립매활성변화；새서람(MTT)법회제세포생장곡선,류식세포의결합전화병정(PI)염색검측세포주기분포.결과 동대조조T47D세포비교,소간우RNA(siRNA)1조화음성대조siRNA2-N조hTERT기인재mRNA화단백수평적표체이급세포단립매활성적변화차이무통계학의의(P＞0.05),이siRNA2조분별하강57.1％、57.0％화57.0(P＜0.01),siRNA3조분별하강53.7％、53.4％화56.5％ (P＜0.01),siRNA4조분별하강70.0％、70.3％화81.9％ (P＜0.01)；siRNA1、siRNA2-N조세포생장속도급각기세포비례차이무통계학의의(P＞0.05),이siRNA2、siRNA3화siRNA4조세포자24h기취출현생장속도명현강저(P＜0.01),G0/G1기세포비례증고(P＜0.01)화S기세포비례강저(P＜0.01).결론 침대hTERT적shRNA가억제종류세포적증식.
Objective To observe the effects of human telomerase reverse transcriptase (hTERT)directed short hairpin RNA (shRNA)-expressing plasmid on telomerase activity and proliferation in breast cancer cell T47D.Methods hTERT-directed shRNA-expressing plasmid was transfected into T47D cells with liposome.Reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting were used to detect the expression of hTERT mRNA and protein.The telomerase activity was examined by using tartrate resistant acid phosphatase (TRAP)-enzyme linked immunosorbent assay (ELISA).MTT assay was used to draw cell growth line,and cell cycles were detected by using flow cytometry.Results Compared with T47D control group,the expression levels of hTERT mRNA and protein,and telomerase activity in small interfering RNA (siRNA)1 group and siRNA2-N group were decreased with no statistical significance (P＞0.01),while those in siRNA2 group were decreased by 57.1％,57.0％ and 57.0％ (P＜0.01),those in siRNA3 group were decreased by 53.7％,53.4％ and 56.5％ (P ＜0.01),and those in siRNA4 group were decreased by 70.0％,70.3％ and 81.9％ (P ＜0.01),respectively.The cell growth rate and cell cycles in siRNA1 and siRNA2-N groups changed with no statistical significance (P ＞ 0.05),while the cell growth rate in siRNA2,siRNA3 and siRNA4 group was decreased at the first 24 h (P ＜ 0.01),and the cell cycles changed significantly (P ＜ 0.01).Conclusion The hTERT-targeted shRNA-expressing vector can inhibit the proliferation of cancer cells.
