MircoRNA-214对吉非替尼敏感及耐药细胞增殖和凋亡的影响  

The Effect of microRNA-214 on Growth and Apoptosis of Gefitinib Sensitive and Resistant Cells

作　　者：姜涵毅[1] 钱丹雯 王永生[1] 苗立云[1] 蔡后荣[1] 

机构地区：[1]南京大学医学院附属鼓楼医院,江苏南京210008 [2]南京市红十字医院,江苏南京210001

出　　处：《中国肿瘤》2015年第9期785-791,共7页China Cancer

基　　金：国家自然科学基金(81301882);中央高校青年教师苗圃项目(20620140118)

摘　　要：[目的]研究mirco RNA-214(miR-214)对PC9肺腺癌及吉非替尼耐药细胞(PC9/GR)增殖和凋亡的影响。[方法]在PC9细胞中转染miR-214模拟物及PC9/GR中转染miR-214抑制剂,使用定量逆转录PCR(q RT-PCR)检测其表达。MTT检测细胞转染miR-214模拟物或其抑制剂后的存活及增殖。在PC9细胞中顺时转染miR-214模拟物以检测上调miR-214对PC9细胞耐药性的影响,在PC9/GR细胞中顺时转染miR-214抑制物以检测下调miR-214对PC9/GR细胞耐药性的影响,并用流式细胞仪检测细胞的凋亡。Western blotting检测PTEN在PC9和PC9/GR细胞中的表达。构建PTEN 3’-UTR荧光素酶报告质粒验证miR-214的靶基因;建立异种移植模型检测miR-214抑制物对肺癌移植瘤的影响。[结果]PC9细胞中miR-214低表达,上调miR-214的表达后PC9细胞对吉非替尼的敏感性降低,并且抵抗吉非替尼诱导的凋亡;而在PC9/GR细胞中低表达miR-214后,下调miR-214增加PC9/GR细胞对吉非替尼的敏感性,并且可以增强吉非替尼诱导的凋亡作用。荧光素酶报告载体实验证实PTEN是miR-214在细胞内的靶基因。动物异种移植模型表明miR-214抑制物可以增强PC9/GR对吉非替尼的敏感性。[结论]Mi R-214可能通过靶基因PTEN调控吉非替尼的获得性耐药。[Purpose] To investigate the effect of micro RNA-214 on the growth and apoptosis of Gefitinib sensitive and resistant cell lines. [Methods] q RT-PCR was performed to detect the expression of mi R-214 in PC9 and PC9/GR after transfection. MTT was used to analyze the survival and growth of cells. mi R-214 mimic was simultaneously trasfected into PC9 cells and the cells response to mi R-214 was analyzed;mi R-214 inhibitor was transfected into PC9/GR and then investigate the cells’ sensitivity to Gefitinib. The apoptosis was detected by flow cytometry and the PTEN expression was detected by Western-blot. Dual luciferase system was establised to demonstrate the downstream target of mi R-214 in tumor cells. Xenografte mice models were established to test the mi R-214 impact in vivo. [Results] The result showed that in Gefitinib sensitive cell line-PC9,mi R-214 was down-regulated,and if mi R-214 was up-regulated,PC9 became resistant to apoptosis induced by Gefitinib. On contrast,mi R-214 was found up-regulated in PC9/GR（Gefitinib resistant cell line）,if the mi R-214 was knock down,PC9/GR became more sensitive to Gefitinib.Dual luciferase report system demonstrated that PTEN was the downstream effector of mi R-214 in tumor cells. Moreover,the xenograft models demonstrated that mi R-214 inhibitor was able to induce the sensitivity of tumor cells to Gefitinib. [Conclusion] mi R-214 might regulate the Gefitinib acquired resistance via PTEN.

关 键 词：miR-214 非小细胞肺癌 吉非替尼 获得性耐药 PTEN 

分 类 号：R734.2[医药卫生—肿瘤]