过表达miR-134b抑制CD133^+ U87胶质瘤干细胞增殖及侵袭  

Overexpressed miRNA-134b inhibits proliferation and invasion of CD133^+ U87glioma stem cells

作　　者：刘义锋[1] 张保朝[1] 温昌明[1] 闻公灵[1] 周国平[2] 张敬伟[3] 贺海发 汪宁[1] 李巍[5] LIU Yifeng ZHANG Baochao WEN Changming WEN Gongling ZHOU Guoping ZHANG Jingwei HE Haifa WANG Ning LI Wei(Department of Neurology Department of Neurosurgery Department of Medical Oncology Department of Pathology, Nanyang Central Hospital Affiliated to Zhengzhou University, Nanyang 473000 Department of Neurology, General Hospital of Shenyang Military Area Command, Shenyang 110016, China)

机构地区：[1]郑州大学附属南阳中心医院神经内科,河南南阳473000 [2]郑州大学附属南阳中心医院神经外科,河南南阳473000 [3]郑州大学附属南阳中心医院肿瘤内科,河南南阳473000 [4]郑州大学附属南阳中心医院病理科,河南南阳473000 [5]沈阳军区总医院神经内科,辽宁沈阳110016

出　　处：《细胞与分子免疫学杂志》2017年第5期637-642,共6页Chinese Journal of Cellular and Molecular Immunology

基　　金：国家自然科学基金(81401097)

摘　　要：目的探讨微小RNA-134b(miR-134b)在胶质瘤干细胞(GSC)中的作用及相关机制。方法实时定量PCR检测U87胶质瘤细胞系分离出的CD133^+ GSC和CD133^- GSC中miR-134b的表达量;包装慢病毒,构建过表达miR-134b的GSC,实时定量PCR检测其miR-134b、CD133、神经干细胞蛋白(nestin)、基质金属蛋白酶2(MMP-2)、MMP-9和MMP-12 mRNA表达水平;Western blot法检测过表达miR-134b的GSC中Notch2、MMP-2、MMP-9和MMP-12蛋白表达;Transwell^(TM)侵袭实验检测过表达miR-134b对GSC侵袭能力的影响;建立U87细胞裸鼠成瘤模型,皮下注射过表达miR-134b(实验组)及空载体(对照组)的CD133^+ GSC,定期测量肿瘤大小,70 d后处死裸鼠,测定肿瘤质量,以探讨过表达miR-134b对在体肿瘤生长的影响。结果实时定量PCR检测结果表明,与CD133^- GSC相比,CD133^+ GSC中miR-134b表达显著下调;通过慢病毒包装的方法成功构建过表达miR-134b的GSC,其miR-134b的表达显著高于空载体对照细胞,而MMP-2、MMP-9 mRNA及蛋白表达无显著变化,而MMP-12的mRNA及蛋白表达均显著下降;Transwell^(TM)侵袭实验结果表明,与对照组相比,过表达miR-134b抑制CD133^+ GSC的侵袭能力;实验组裸鼠体内肿瘤体积显著低于对照组,与对照组相比,过表达miR-134b的实验组肿瘤质量也降低了42%。结论过表达miR-134b抑制CD133^+ GSC的生长和侵袭。Objective To investigate the role of microRNA-134b （miR-134b） in the tumorigenesis of glioma stem cells （GSCs） and the possible molecular mechanism. Methods Real-time quantitative PCR （qRT-PCR） was used to evalate the expression of miR-134b in CD133^＋ and CD133^- U87 GSCs. A lentiviral vector overexpressing miR-134b in U87 GSCs was constructed, and the effect of miR-134b overexpression on matrix metalloproteinase-2 （MMP-2）, MMP-9 and MMP-12 expressions at both mRNA and protein levels were detected by qRT-PCR and Western blotting, respectively. TranswellTM assay was performed to determine the effect of miR-134b overexpression on GSCs invasion ability. Tumor xenograft models in nude mice were established to evaluate the effect of miR-134b overexpression on tumorgenesis in vivo. Results The qRT-PCR showed that, compared with CD133- cells, miR-134b was significantly down-regulated in CD133^＋ cells. Cell line over-expressing miR-134b was successfully established, and miR-134b was up-regulated significantly compared with empty vector control. Overexpression of miR-134b remarkably inhibited the invasion of U87 GSCs and the expression of MMP-12. However, overexpression of miR-134b did not affect MMP-2 and MMP-9 expressions, rniR-134b also suppressed U87 GSCs xenograft growth in vivo. Tumor volume in tumor xenograft model group was significantly lower than that in control group, and tumor weight decreased by 42% in the former group. Conclusion Overexpression of miR-134b inhibits the growth and invasion of CD133^＋ GSCs.

关 键 词：微小RNA-134b(miR-134b) 基质金属蛋白酶(MMP) U87胶质瘤 

分 类 号：R741[医药卫生—神经病学与精神病学] R392-33[医药卫生—临床医学]