慢病毒介导微小RNA-20a过表达抑制胰腺癌细胞的增殖  

Inhibitory effects of ientivirus-mediated overexpression of microRNA-20a on proliferation of pan- creatic carcinoma cells

作　　者：燕海姣[1] 蒋敬庭[1] 刘文松[2] 王琦[1] 石红兵[1] 罗子俨[1] 吴昌平[1] 

机构地区：[1]苏州大学附属第三医院肿瘤生物诊疗中心常州市医学生物技术重点实验室,常州213003 [2]苏州大学附属第三医院肝胆外科

出　　处：《中华实验外科杂志》2012年第9期1771-1774,共4页Chinese Journal of Experimental Surgery

基　　金：国家自然科学基金资助项目（30972703、81171653）;江苏省卫生厅医学科技发展基金资助项目（17200932、P200933）;六大人才高峰第六批资金资助项目（BRA2010037）;常州市卫生局青年人才科技项目（QN201103）

摘　　要：目的观察慢病毒介导微小RNA（miRNA）-20a过表达对胰腺癌细胞增殖的影响，探讨胰腺癌基因治疗靶位。方法采用荧光原位杂交和实时定量逆转录-聚合酶链反应（RT—PCR）检测BxPC3、Panc-1和永生化的正常胰腺导管上皮细胞H6C7中miRNA-20a的表达。体外实验分析慢病毒介导miRNA-20a过表达对胰腺癌细胞株增殖能力的影响，流式细胞仪分析细胞周期分布。结果miRNA-20a在胰腺癌细胞中呈低表达（P〈0．01）；过表达miRNA-20a在体外可抑制胰腺癌细胞增殖（P〈0．01），生长曲线最大抑制效率分别是（59．21±0．31）％和（59．04±2．01）％；软琼脂克隆形成试验，增殖率分别下降到（45．21±1．72）％和（38．74±2．13）％。miRNA一20a使细胞周期阻滞于G0／G1期，分别为（62．3±4．5）％和（64．2±4．8）％（P〈0．05）。同时细胞周期蛋白D1（CyclinD1）表达水平下调，但其mRNA水平不变（P〉0．05）。结论miRNA-20a过表达能抑制胰腺癌细胞增殖，其机制可能是在转录后水平抑制CyclinD1蛋白的表达。Objective To investigate the＇e^ffect of microRNA （miRNA）-20a overexpression medi- ated by lentivirus on pancreatic carcinoma cell proliferation, and to find new target spot of gene therapy for pancreatic carcinoma. Methods In situ hybridization and quantitative real-time reverse transcription-poly- merase chain reaction （RT-PCR） was used to evaluate the expression of miRNA-20a in two pancreatic car- cinoma cell lines （ BxPC3 and Panc-1 ） and immortal human pancreatic duct epithelial cell line H6C7. The effect of lentivirus-mediated overexpression of miRNA-20a on pancreatic carcinoma cell proliferation in vitrowas analyzed, and the cell cycle distribution was measured by using flow cytometry. Results The ex- pression level of miRNA-20a was downregulated in pancreatic carcinoma cells as compared with that in nor- mal pancreatic cells. Stable overexpression of miRNA-20a significantly inhibited proliferation of pancreatic carcinoma cells BxPC-3 and Panc-1 in vitro （P 〈0. 01 ）. The maximum inhibition rate of growth curve was （59. 21±0. 31 ）% and （59. 04 ±2. 01 ）% respectively. In soft agar colon formation test, the percentage of proliferation was decreased to （45.21 ± 1.7 ） % and （ 38.74 ± 2. 10 ） % respectively. Cell cycle distri- bution analysis showed there were great changes in miRNA-20a overexpressing clones. Many cells were blocked in the G0/GI phase, and the proportion of BxPC3 and Pane-1 cells was （62. 3 ± 4. 5） % and （64. 2±4. 8）% respectively （P 〈 0.05）. Furthermore, it was found that Cyclin D1 protein expression was downregulated by overexpression of miRNA-20a without changing the Cyelin D1 mRNA level. Conclu- sion Overexpression of miRNA-20a significantly inhibited proliferation of pancreatic carcinoma cells, and the probable mechanism was the negatively regulation of Cyelin D1 protein expression at the post-transcrip- tional level.

关 键 词：胰腺癌 微小RNA 增殖 

分 类 号：R735[医药卫生—肿瘤]