基于基因沉默的磁纳米介导体系的制备及性能研究

Preparation and Activity of Magnetic Nano-mediated System Based on Gene Silencing

目的:制备载基因的磁性纳米药物体系,并考察其生物特性和体外基因沉默效果。方法:采用化学共沉淀法制备Fe3O4磁纳米粒子,并进行包硅、氨基功能化,结合siIDO;红外光谱(FT-IR)、透射电镜(TEM)对载药磁纳米颗粒进行物理表征,并通过凝胶阻滞实验检测磁纳米粒子对siIDO的载药量;CCK8检测磁性纳米载体的细胞毒性,流式细胞仪检测磁纳米粒子-siIDO的转染效率,Q-PCR检测磁纳米粒子-siIDO的体外沉默效果。结果:磁纳米粒子呈球形,平均粒径10~20 nm,理想的磁性纳米粒子与siIDO质量比为1:1;CCK8检测结果显示,基因运载体系的浓度为5μg/m L时对细胞的活性没有影响;磁纳米粒子-siIDO对DC的转染效率为90%以上;Q-PCR检测磁纳米粒子-siIDO的体外沉默效果较好,使DC的IDO表达下调64.5%。结论:载基因的磁性纳米药物体系在肿瘤的免疫治疗领域具有应用潜力。

Objective:To prepare a gene carrying magnetic nanodrug system and to investigate its biological characteristics and gene silencing effect in vitro.Methods:Fe3O4 magnetic nanoparticles were prepared by chemical co-precipitation method,and functionalized with silicon and amino,combined with siIDO;The particles were characterized by infrared spectroscopy(FT-IR)and transmission electron microscopy(TEM),and the loading capacity of siIDO was detected by gel retardation test.CCK8 was used to detect the cytotoxicity of magnetic nanoparticle,flow cytometry was used to detect the transfection efficiency of magnetic nanoparticle-siIDO,and Q-PCR was used to detect the silencing effect of magnetic nanoparticle-siIDO in vitro.Results:The magnetic nanoparticles were spherical with an average particle size of 10~20 nm.The ideal magnetic nanoparticles and siIDO drug loading ratio was 1∶1.CCK8 showed that the gene delivery system concentration of 5μg/mL had no effect on cell activity.The transfection efficiency of magnetic nanoparticle siIDO for DC was more than 90%.Q-PCR showed a good silencing effect on magnetic nanoparticles-siIDO in vitro,and reduced the expression of IDO in DC by 64.5%.Conclusion:Magnetic nano-drug system carrying gene has potential application in the field of tumor immunotherapy.