葡聚糖磁性纳米颗粒外加磁场对人树突状细胞基因转染效率的研究

Effect of Dextran-coated Magnetic Iron Oxide Nanoparticles in the Magnetic Field on Transfection Efficiency of Target Gene into Human Dendritic Cells

目的研究葡聚糖磁性纳米颗粒（the dextran coated magnetic iron oxide nanoparticles，DMN）在外加钕一铁一硼稀土固定磁场的作用下对人树突状细胞转染效率以及安全性的影响。方法先通过磁力计对DMN进行分析；再将修饰有多聚赖氨酸（Poly-L—Lysine，PLL）的DMN携带绿色荧光蛋白pEGFP—Cl质粒报告基因，在钕-铁-硼稀土周定强磁场的作用下，体外转染人树突状细胞，用荧光显微镜直接观察和流式细胞仪检测来评价外加磁场对DMN作为人树突状细胞转染载体效率的影响；在转染后采用MTT比色法测定在磁场干预下的DMN对人树突状细胞增殖和功能的影响以了解其细胞毒性。结果DMN的核心直径〈30nm，具有明硅的超顺磁性，比饱和磁化强度也明显高于相同Fe3O4含量的普通磁块；DMN作为基因载体在外加磁场作用下，转染12h即可将报告基因转染至人树突状细胞内并成功表达，在荧光显微镜下可观察到绿色荧光细胞，24h转染率可达到最高（约为27％），转染效率较未加磁场组提高了2～4倍。而且转染后的人树突状细胞增殖活性及功能未因DMN外加磁场及其作用时间的长短而受到影响。结论超顺磁性的DMN在外加磁场作用下可以明显、安全、有效地提高对人树突状细胞的转染效率。

Objective To evaluate transfection efficiency and safety of dextran-coated magnetic iron oxide nanoparticles （DMN） as gene carrier to deliver target gene into human dendritic cells （HDC） in the Nd-Fe-B permanent magnetic field. Methods The dextran-eoated magnetic iron oxide nanopartieles were first characterized by vibrating sample magnetometer signal processor, and then modified with Poly-L-Lysine （PLL） as a gene carrier containing pEGFP-C1 report plasmid. The modified DMN cartier was transfected into human dendritic cells （HDC） in the magnetic field using Nd-Fe-B permanent magnet. Transfection efficiency of DMN was evaluated by detecting GFP intensity under fluorescence microscope magnetic field on HDC cells was assessed by and flow cytometer. The cytotoxicity of DMN in the use of MTT proliferation assay. Results The dextrancoated magnetic iron oxide nanoparticles showed superparamagnetic characteristics at the size of core diameters smaller than 30nm, with significantly higher saturation and magnetization than that of common magnetism containing equal amount of Fe304 12h after transfection of the GFP reporter delivering DMN vector in the magnetic field, GFP was successfully expressed in HDC cells, achieving 27% transfection efficiency after 24h, indicative for 2-4 fold increase of transfection efficiency by the DMN. magnetic field, compared to non-magnetic transfection. MTT assay showed that proliferation and function of HDC cells after transfection were not affected by the DMN in the magnetic field or the time course of DMN contact. Conclusion Superparamagnetic dextran-coated magnetic iron oxide nanoparticles in the magnetic field provide a safe and efficient transfection vector to enhance delivery of target gene into HDC cells in vitro.