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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 observe the effect of dextran magnetic nanoparticles (DMN) transfected with small interfering RNA (siRNA) recombinant plasmids on survivin gene expression and apoptosis in bladder cancer cells under the synergistic effect of external magnetic field. METHODS: DMN was prepared by chemical coprecipitation method and used as gene carrier. The recombinant plasmid siRNA-survivin was electrostatically adsorbed by polylysine and transfected into human bladder cancer cell line BIU-87 under synergistic effect of external magnetic field. The growth inhibition rate (IR) and apoptosis index (BIU) of BIU-87 cells were detected by the method of azithromycin blue, terminal deoxynucleotidyl transferase mediated dUTP nick end labeling, reverse transcription polymerase chain reaction and Western Blotting (AI), survivin mRNA and protein relative expression levels. Results The diameter of siRNA-DMN, effective particle size and specific saturation magnetization were 10-12 nm, 94.8 nm and 0.19 emu / g, respectively. The IR (39.60%) and AI (28.72%) of DMN transfected with siRNA-survivin into BIU-87 cells were significantly higher than those of control group and DMN group without magnetic field (1.99%, 3.75% and 20.96% and 16.71%, respectively) (P <0.05). The relative expression levels of survivin mRNA and protein were significantly lower than the latter two groups. Conclusion DMN transfected with siRNA-survivin plasmid into BIU-87 cells under the synergistic effect of external magnetic field can effectively inhibit the expression of survivin gene and induce apoptosis.