摘要
目的观察肿瘤相关钙信号传导蛋白-2（TROP-2）基因小干扰RNA（siRNA）对乳腺癌细胞黏附和侵袭力的影响。方法培养人乳腺癌Bcap-37、LCC1、MCF-7、MDA-MB-231、MDA-MB-435、MDA-MB-468及ZR75—1细胞株，以荧光实时定量聚合酶链反应（PCR）方法检测TROP-2基因mRNA表达；筛选出TROP-2表达最高者。采用TROP-2基因siRNA转染乳腺癌细胞，分别以荧光实时定量RT—PCR和免疫荧光方法观察TROP-2基因mRNA和蛋白水平，然后以噻唑蓝（MTT）比色法检测细胞黏附性，以Transwell方法检测癌细胞侵袭能力。结果人乳腺癌Bcap-37、LCCl、MCF-7、MDA-MB-231、MDA—MB435、MDA—MB468及ZR75-1细胞株TROP-2 mRNA分别是1．362±0．057、2．207±0．056、2．997±0．052、0．136±0．045、0．122±0．025、0．194±0．028和2．706±0．039，以MCF．7细胞最高；以TROP^siRNA转染乳腺癌MCF-7细胞后，癌细胞TROP-2基因mRNA和蛋白水平明显下降，且呈浓度依赖性；黏附实验结果显示，5、10、20nmol/L siRNA组黏附率分别为（52．9±2．5）％、（25．6±2．3）％、（12．8±2．2）％（P〈0．01）；Transwell实验结果显示，5、10和20nmoFLsiRNA组穿过滤膜的细胞分别为78±17、39±15、19±16，而对照组分别为136±25、139±21（P〈0．01）。结论TROP-2基因在乳腺癌细胞黏附和侵袭中发挥着重要作用；以siRNA转染乳腺癌细胞，可抑制乳腺癌细胞黏附和侵袭能力。
Objective To study the effects of tumor-associated calcium signal transducer-2 （TROP-2） gene small interfering RNA （siRNA） on adhesion and invasion of human breast cancer cells. Methods Real time PCR was used to detect the TROP-2 mRNA of seven human breast cancer cell lines： Bcap-37, LCC1, MCF-7, MDA-MB-231, MDA-MB-435, MDA-MB-468, and ZR75-1. The cell line with hifhest TROP-2 expression was transfected with different doses of TROP-2 siRNA. The expression of TROP-2 mRNA and protein was detected by real-time quantitative polymerase chain reaction （PCR） and immumoflurescence method. The cell adhesion was evaluated by methyl thiazol tetrazolium （MTT） assay, and invasion was exmined by boyden chamber, respectively. Results The TROP-2 mRNA in Bcap-37, LCC1, MCF-7, MDA-MB-231, MDA-MB-435, MDA-MB-468 and ZR75-1 cell lines was 1. 362 ±0. 057, 2. 207 ±0. 056, 2. 997 ±0. 052, 0. 136 ±0. 045, 0. 122 ±0. 025, 0. 194 ±0. 028 and 2. 706 ±0. 039 re- spectively, and MCF-7 showed the highest elevation of TROP-2 mRNA. The real-time quantitative PCR and immumoflurescence method revealed that the expression of TROP-2 mRNA and protein was reduced in a time- and dose-dependent manner （P 〈0. 01 ;P 〈0. 01 ）. The adhesive rate in siRNA groups （5, 10 and 20 nmol/L） was （52.9±2.5）%, （25.6±2.3）% and （12.8±2.2）% （P〈0.01）, respectively. The transwell results showed that the invasion cells were 78 ± 17, 39 ± 15, 19 ± 16, 136 ± 25 and 139 ± 21 in different groups （5, 10, 20 nmol/L siRNA, and controls）, respectively （P 〈0.01 ）. Conclusion TROP-2 gene might play an important role in adhesion and invasion of human breast cancer cells, siRNA targeted TROP-2 could effectively inhibit adhesion and invasion of human breast cancer cells.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2011年第10期1702-1704,共3页
Chinese Journal of Experimental Surgery
基金
镇江市分子内分泌重点文验室资助项目（SS2009012）
江苏省自然科学基金资助项目（BK2009205）
镇江市丰十会发展基金资助项目（SH2009014）
关键词
乳腺癌
肿瘤相关钙信号传导蛋白-2
RNA干扰
侵袭
Breast carcinoma
Tumor-associated calcium signal transducer-2
RNA interference
Invasion
