miR-17-92基因簇增强前列腺癌DU145细胞的迁移、侵袭能力及对顺铂的耐药性  

miR-17-92 cluster increases the migration and invasion abilities of DU145 prostate cancer cells and enhances the cisplatin resistance

作　　者：陈昊[1] 周鹏[1] 徐晶晶[1] 周珺[1] 国风[1] 

机构地区：[1]苏州大学附属第一医院中心实验室,江苏苏州215007

出　　处：《中国癌症杂志》2017年第2期95-101,共7页China Oncology

基　　金：国家自然科学基金资助项目(81172433,81400154);江苏省自然科学基金资助项目(BK20151211)

摘　　要：背景与目的:mi R-17-92基因簇与多种疾病的发生密切相关,其在肺癌、肝癌、胃癌和前列腺癌等多种肿瘤细胞中均高表达。本研究利用慢病毒包装系统建立稳定高表达mi R-17-92基因簇的DU145细胞株,探讨mi R-17-92基因簇对前列腺癌DU145细胞的迁移、侵袭能力及对顺铂耐药性的影响。方法:构建高表达mi R-17-92基因簇的表达载体,转染DU145细胞株,同时转染空载体作为对照,并用实时荧光定量聚合酶链反应(real-time fluorescent quantitative polymerase chain reaction,RTFQ-PCR)进行鉴定。用x CELLigence系统监测细胞的迁移、侵袭能力及顺铂处理后的生长情况;通过划痕实验观察细胞的迁移情况;采用蛋白[质]印迹法(Western blot)、凝胶酶谱实验和RTFQ-PCR检测相关蛋白质和基因的表达以探讨mi R-17-92增强DU145细胞的迁移、侵袭能力及对顺铂耐药性的相关机制。结果:DU145-mi R-17-92细胞迁移速率和侵袭能力高于DU145-control细胞(P<0.01)。DU145-mi R-17-92细胞中整合素β1的蛋白质表达水平和基质金属蛋白酶-9(matrix metalloprotein-9,MMP-9)的活性显著高于DU145-control细胞。顺铂处理后,DU145-mi R-17-92细胞的生长速度自12 h起快于DU145-control细胞并呈顺铂耐药性(P<0.01)。细胞外调节蛋白激酶1/2(extracellular regulated protein kinases,ERK1/2)在DU145-mi R-17-92细胞中呈现持续高水平磷酸化,顺铂处理后,其磷酸化水平无明显变化。DU145-mi R-17-92细胞中切除修复互补交叉基因1(excision repair cross complementing1,ERCC1)的m RNA和蛋白质表达水平显著高于DU145-control细胞。结论:高表达mi R-17-92增强了DU145细胞的迁移、侵袭能力,其机制与整合素β1的表达上调及MMP-9活性增强有关。此外,高表达mi R-17-92增强了DU145细胞对顺铂的耐药性,该过程与ERK1/2的磷酸化水平增加和ERCC1的表达水平上调相关。Background and purpose： miR-17-92 gene cluster overexpression has been observed in various cancers, such as lung cancer, liver cancer, gastric cancer and prostate cancer. In this study, we established the stable cell line overexpressing miR-17-92 to explore the influence of miR-17-92 on the migration, invasion abilities and cisplatin resistance of the prostate cancer DU145 cells. Methods： miR-17-92 overexpression vectors were constructed. DU145 cells were infected with the viral supernatants produced by Phoenix A packaging system. Real-time fluorescent quanti- tative polymerase chain reaction （RTFQ-PCR） was conducted to detect the expression level of miR-17-92 in the cells. The migration and invasion abilities were measured by a real-time xCELLigence system. The scratch healing assay was carried out to investigate the migration abilities. The expression of integrin 131 was detected by Western blot, and the activities of matrix metalloprotein-2 （MMP-2） and matrix metalloprotein-9 （MMP-9） were measured by gelatin zymography experiment. The cell growth of the two cell lines after the treatment of cisplatin was detected by a real-time xCELLigence system. The mRNA expression of ERCC1 was measured by RTFQ-PCR. Western blot was conducted to investigate the protein expressions ofERCC1, ERK1/2 and pERK1/2. Results： DU145-miR-17-92 cells migrated faster than DU145-control cells during the 24 h continuous monitoring （P〈0.01）. The scratch healing assay indicated that DU145-miR-17-92 cells migrated from the edge towards the scratch center faster than DU145-control cells. DU145- miR-17-92 cells invaded through matrigel markedly faster than DU145-control cells （P〈0.01）. The protein expression level of integrin 131 and the MMP-9 activities in DU145-miR-17-92 cells were increased than those in DU145-control cells. After the treatment of cisplatin, DU 145-miR- 17-92 cells grew faster than DU 145-control cells, presenting cisplatin resistance （P〈0.01）. The phosphorylation of ERK1/2 in DU145-miR-1

关 键 词：miR-17-92 前列腺肿瘤 DUl45 侵袭 迁移 顺铂 

分 类 号：R737.25[医药卫生—肿瘤]