超顺磁性葡聚糖氧化铁纳米颗粒的制备及其作为基因载体的可行性研究

背景与目的：磁性纳米颗粒作为基因载体在肿瘤基因治疗中的应用得到了迅速发展。为了能获得驱动目的基因高效稳定表达、安全无害、靶向性高、简便的新型非病毒型基因导入和治疗系统,本研究探讨超顺磁性葡聚糖氧化铁纳米颗粒(superparamapnetic dextran iron oxide nanoparticles,SDION)的制备及其作为体外基因载体的可行性.方法：采用化学共沉淀法制作SDION,通过丙烯葡聚糖凝胶S-300HR色谱和离心法分离SDION,用透射电镜、粒度分析仪和磁力计对SDION进行分析。以绿色荧光蛋白(GFP-C2)质粒为靶基因,通过氧化还原法构建SDION-GFP-C2复合物,用紫外分光光度计和琼脂糖凝胶电泳检测两者的结合率。以脂质体转染作为对照,荧光显微镜分别观察SDION和脂质体体外转染GFP-C2入膀胱癌细胞BIU-87的转染效率。结果：SDION直径在3～8nm之间,有效粒径为59．2nm,比饱和磁化强度为0．23emu／g。分别经10mmol／L的高碘酸钠氧化、0．5mol／L的硼氢化钠还原作用后的sDION和GFP的结合比例最大,sDION对GFPDNA的转染效率为45％左右,明显高于脂质体的转染效率(30％左右)。结论：SDION可通过氧化还原反应与GFP质粒相连,在体外可将GFP基因成功转染入人膀胱癌BIU-87细胞。

Preparation and feasibility of superparamagnetic dextran iron oxide nanoparticles as gene carrier

BACKGROUND &OBJECTIVE: Application of magnetic nano-particles as gene carrier in gene therapy of tumor has developed quickly. To obtain a new typ e non-viral gene introduction and therapy system,which is convenient,and can dr ive target gene to express highly and stably,this study was designed to explore the preparation of superparamagnetic dextran iron oxide nanoparticles(SDION),and the feasibility of SDION used as gene carrier in vitro. METHODS: SDION were pre pared by chemical co-precipitation,separated by gel filtration chromatography o n Sephacryl S-300HR,and centrifugation techniques,characterized by transmission electron microscopy,laser scattering system,and vibrating sample magnetometer s ignal processor. The green fluorescent protein-C2 (GFP-C2) plasmid was used as target gene. SDION-GFP-C2 compounds were synthesized by oxidation-reduction reaction. The connection rate of SDION and GFP-C2 was analyzed by agarose elect rophoresis,and evaluated by measuring concentration of GFP in the supernatant af ter centrifugation. Liposome transfection was used as control,the efficiencies o f SDION and liposome in transferring GFP gene into human bladder cancer BIU-87 cells were evaluated under fluorescence microscope in vitro. RESULTS: The diamet er of SDION ranged from 3 nm to 8 nm,the effective diameter was 59.2 nm,and the saturation magnetization was 0.23 emu/g. After oxidized by sodium periodate of 1 0 mmol/L,and deoxidized by sodium hydride boron of 0.5 mol/L,SDION could connect with GFP in maximum degree,the transfection efficiency of SDION as gene carrier was about 45%,even higher than that of liposome (about 30%). CONCLUSION: SDIO N could connect with GFP plasmid by oxidation-reduction reaction,and success to transfer GFP gene into human bladder cancer BIU-87 cells in vitro.