摘要
目的观察葡聚糖磁性纳米颗粒(DMN)在外磁场协同作用下转染存活素(survivin)小干扰 RNA(siRNA)重组质粒对膀胱癌细胞 survivin 基因表达和凋亡的影响。方法利用化学共沉淀法制备 DMN 并作为基因载体,通过多聚赖氨酸静电吸附 siRNA-survivin 重组质粒,并在外磁场的协同作用下体外转染人膀胱癌 BIU-87细胞,分别采用四甲基偶氮唑蓝比色法、末端脱氧核苷酸转移酶介导的 dUTP 缺口末端标记技术法、逆转录聚合酶链反应和 Western Blotting 检测 BIU-87细胞的生长抑制率(IR)、凋亡指数(AI)、survivin mRNA 及其蛋白的相对表达水平。结果构建的 siRNA-DMN直径、有效粒径、比饱和磁化强度分别为10～12 nm、94.8 nm、0.19 emu/g。DMN 在外磁场的作用下转染 siRNA-survivin 入 BIU-87细胞后的 IR(39.60%)、AI(28.72%)均分别显著高于对照组和无磁场作用的 DMN 组(1.99%、3.75%和20.96%、16.71%)(P<0.05),survivin mRNA 及其蛋白相对表达水平均显著低于后两组。结论 DMN 在外磁场的协同作用下转染 siRNA-survivin 质粒入 BIU-87细胞后可有效抑制 survivin 基因表达并诱导细胞凋亡。
Objective To investigate the effects of the small interfering RNA plasmid-dextran magnetic nanoparticles （siRNA-DMN） combination with external magnetic fields on silencing survivin gene expression of bladder cancer cells and apoptosis when DMN used as gene carrier to transfer siRNA-survivin recombinant plasmid in vivo. Methods The siRNA-survivin recombinant plasmid specific targeted survivin was synthesized in previous experiment. DMN were prepared by chemical coprecipitation method and used as gene carrier. The siRNA-DMN were constructed by static electricity of polylysine and transferred into human bladder cancer BIU-87 cells with the help of external magnetic fields. The growth inhibiting rate （IR） of BIU-87 cells was observed by 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyl tetrazolium bromide＇s test and the apoptosis index （AI） was detected by transferase-mediated dUTP nick end labeling method. The relatively transcription levels of survivin mRNA and protein expression were respectively detected by semi-quantitive reverse transcription polymerase chain reaction and Western Blotting techniques. Results The diameter, effective diameter and saturation magnetization of DMN-siRNA were about 10-12 nm, 94.8 nm and 0. 19 emu/g, respectively. The IR （39. 60% ） and AI （28.72%） , the relative expression of survivin mRNA and protein of siRNA-DMN combination with external magnetic fields on BIU-87 cells were significantly higher and lower than those in the control group and single siRNA-DMN group, respectively （P〈0. 05）. Conclusions The siRNA-survivin plasmid-DMN combination with external magnetic fields could effectively inhibit survivin expression and induce BIU-87 cells apoptosis which provided experimental basis for the magnetic targeting gene therapy of bladder tumor.
出处
《中华外科杂志》
CAS
CSCD
北大核心
2006年第18期1248-1251,共4页
Chinese Journal of Surgery
基金
国家自然科学基金(30271300)
