原代树突细胞基因递送系统的构建与应用

Construction and application of gene delivery systems for primary dendritic cells

目前,安全有效的基因递送系统应用于原代树突细胞(dendritic cell,DC)转染尚未见报道,本研究针对原代DC构建基于脂质体的基因递送系统,优化了制备方法以提高原代DC的转染效率。通过共孵育法、乙醇注入法、鱼精蛋白复合法分别制备含不同阳离子脂质的脂质体/siRNA复合物,并以粒径、电位、对基因药物siRNA的包载能力、安全性、稳定性、DC的摄取效率和基因沉默效率为考察指标,评价并筛选出具有高转染效率的基因递送系统。与市售制剂Lipo2000相比,利用共孵育法制备的赖氨酸谷氨酸双油醇酯(OA2)递送系统的DC摄取效率提高约35%,基因沉默效率提高约10倍,且细胞存活率比Lipo2000提高20%,具有良好的DC基因转染效率和体外安全性。本研究为原代DC提供了一种安全有效、制备简单的基因递送载体平台,具有良好的应用前景。

Nowadays,there is still no mature gene delivery system for safe and effective transfection on primary dendritic cells(DC).Herein,we constructed a liposome-based gene delivery system for primary DCs and optimized the preparation method to improve the transfection efficiency of siRNA on primary DCs.In this study,different methods,including co-incubation method,ethanol injection method,and protamine compound method,were used to prepare liposome/siRNA complexes based on different cationic lipids.Moreover,particle size,zeta potential,siRNA loading capacity,safety,stability,uptake efficiency and gene silencing efficiency of various liposome/siRNA complexes were detected to screen the optimal cationic lipid as well as its preparation method.We demonstrated that the OA2/siRNA delivery system prepared by the co-incubation method exhibited the best safety,uptake efficiency and gene silencing effect,compared to other siRNA delivery systems including the commercial Lipo2000.In summary,we provide a safe and effective gene delivery vector for primary DC cells through simple preparation method,which could also offer a gene delivery platform for other immune cells.