癌变·畸变·突变 ›› 2017, Vol. 29 ›› Issue (3): 199-204.doi: 10.3969/j.issn.1004-616x.2017.03.008

• 论著 • 上一篇    下一篇

磁性纳米颗粒载ABCG2-siRNA逆转乳腺癌细胞对阿霉素耐药性的研究

朱婷1,2, 万丽丽2, 刘丽桃2, 阮海英2, 饶茜2, 谢妮2   

  1. 1.广州医科大学研究生院, 广东 广州 511436;
    2.深圳大学第一附属医院深圳市第二人民医院, 广东 深圳 518037
  • 收稿日期:2017-03-02 修回日期:2017-04-01 出版日期:2017-05-31 发布日期:2017-05-31
  • 通讯作者: 谢妮,E-mail:kejiaoke100@163.com E-mail:kejiaoke100@163.com
  • 作者简介:朱婷,E-mail:402087245@qq.com。
  • 基金资助:
    广东省自然科学基金(2016A030313029);广东省科技计划项目(2014A020212038);深圳市科技创新委基础研究项目(JCYJ20150 330102720122);深圳市科技创新委国际合作项目(GJHZ2016030 1163138685)

ABCG2-siRNA loaded magnetic nanoparticles enhanced therapeutic effect of adriamycin on multidrug resistant breast cancer cells

ZHU Ting1,2, WAN Lili2, LIU Litao2, RUAN Haiying2, RAO Xi2, XIE Ni2   

  1. 1. Graduate School of Guangzhou Medical University, Guangzhou 511436;
    2. Shenzhen Second People's Hospital, First Affiliated Hospital of Shenzhen University, Shenzhen 518037, Guangdong, China
  • Received:2017-03-02 Revised:2017-04-01 Online:2017-05-31 Published:2017-05-31

摘要: 目的:研究抑制乳腺癌耐药蛋白(ABCG2)表达对乳腺癌细胞耐药性的逆转作用,为乳腺癌基因靶向治疗提供新思路。方法:通过透射电镜和纳米粒度电位分析仪对磁性纳米颗粒(polyMAG)的粒径、电位进行表征,利用凝胶电泳阻滞实验检测polyMAG 与siRNA的结合情况,并用polyMAG携载不同浓度(5、10、20、50、100 nmol/L)ABCG2-siRNA,将其转染入乳腺癌细胞耐药株(MCF-7/ADR)后,采用CCK-8检测siRNA转染后细胞的存活率,荧光定量PCR和Western blotting分别检测ABCG2 mRNA和蛋白表达水平,Calcein-AM/PI染色观察细胞凋亡。结果:polyMAG粒径约100 nm,呈正电荷,表面电位29.6 mV。当polyMAG与ABCG2-siRNA的比为1:1和1:2时ABCG2-siRNA可完全结合。当20~100 nmol/L polyMAG-siRNA干扰ABCG2表达后,细胞存活率较未转染组明显降低(P < 0.05)。20 nmol/L polyMAG-siRNA转染组ABCG2 mRNA表达显著下调,约为未转染组的1/8(P < 0.05),且凋亡细胞较其他组明显增多;50 nmol/L polyMAG-siRNA转染后可明显干扰ABCG2蛋白表达(P < 0.05)。结论:polyMAG-siRNA能高效干扰ABCG2表达,增加MCF-7/ADR细胞对阿霉素敏感性,逆转细胞耐药。

关键词: 磁性纳米粒子, siRNA, 乳腺癌耐药蛋白, 多药耐药

Abstract: OBJECTIVE:To investigate the use of siRNA to silence the breast cancer resistance protein (BCRP/ABCG2) on overcoming multidrug resistance in breast cancer cells. METHODS:The morphologic,particle size,surface potential of magnetic nanoparticles (polyMAG) were characterized by transmission electron microscopy and by zeta potential analyzer,and the binding capacity of siRNA and polyMAG were investigated by gel retardation assay. We used polyMAG as a carrier to deliver several concentrations of siRNA (5,10,20,50,100 nmol/L) effectively into MCF-7/ADR cells. Quantitative real-time PCR (qPCR) and Western blotting were used to detect ABCG2 levels. CCK-8 and Calcein-AM/PI were stained and used to detect cell proliferation. RESULTS:The average size and zeta potentials of polyMAG was about 100 nm and 29.6 mv. Gel retardation assay confirmed that siRNA was well packaged into polyMAG under polyMAG/siRNA ratios 1:1 and 1:2. Cell viability of MCF-7/ADR was reduced while siRNA concentrations changed from 20 nmmol/L to 100 nmmol/L,compared with non-siRNA transfection (P < 0.05). Expression of ABCG2 mRNA was significantly down-regulated in the 20 nmol/L polyMAG-siRNA transfected group,which was about 1/8 of the un-transfected group (P < 0.05),and apoptotic cells were significantly increased compared with other groups. 50 nmol/L polyMAG-siRNA significantly interfered with the expression of ABCG2 protein (P < 0.05). CONCLUSION:polyMAG-siRNA effectively perturbed the ABCG2 expression and enhanced DOX susceptibility of MCF-7/ADR to overcome multidrug resistance in breast cancer cells.

Key words: magnetic nanoparticles, siRNA, breast cancer resistance protein, multidrug resistance

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