In vitro comparative evaluation of three CLD/siRNA nanoplexes prepared by different processes  

不同工艺制备的CLD/siRNA纳米粒体外性质差异性研究（英文）

作　　者：Chong Qiu Shihe Cui Jing Sun Jiancheng Wang 邱崇;崔仕贺;孙晶;王坚成(北京大学医学部药学院分子药剂学与新释药系统北京市重点实验室,北京100191)

机构地区：Beijing Key Laboratory of Molecular Pharmaceutics and New Drug Delivery Systems, School of Pharmaceutical Sciences, Peking University Health Science Center, Beijing 100191, China

出　　处：《Journal of Chinese Pharmaceutical Sciences》2016年第9期660-668,共9页中国药学（英文版）

基　　金：National Basic Research Program of China(973 program,Grant No.2013CB932501);the National Natural Science Foundation of China(Grant No.81273455,81473158 and 81573374);Programs of Ministry of Education of China(Grant No.NCET-11-0014 and BMU20110263)

摘　　要：In this study, a gemini-like cationic lipid （CLD） was used as the carrier to study the complexation features of CLD/ siRNA nanoplexes （CLD/siRNA NPs）. Three types of CLD/siRNA nanoplexes （named as AT NPs, HT NPs and MT NPs） were prepared by different processes： AT method （mixing siRNA solution with preformed CLD nanoparticles）, HT method （hydrating a CLD thin film with siRNA solution）, and MT method （dropping an ethanolic solution of CLD into siRNA solution under sonication）. The particle size, zeta potential, morphology, siRNA protection, cytotoxicity, cellular uptake, and targeted mRNA downregulation were studied. At the optimal N/P ratio of 10, the sizes of the three CLD/siRNA NPs were MT NPs （（222.3±19.1） nm）〉 HT NPs （（105.7±1.31） nm）〉AT NPs （（91.8±1.75） nm）. Different nanostructures were formed despite the fact that they were composed of the same components. Furthermore, the TEM images indicated that different morphologies were found in the three NPs, indicating that the nanoplexes were assembled by different mechanisms. Among the three NPs, the cell uptake capacity were as follows： AT NPs〉MT NPs〉HT NPs, whereas the silencing levels on epidermal growth factor receptor （EGFR） in HeLa cells were MT NPs〉AT NPs〉HT NPs. Based on the above results, we hypothesized that the different preparation processes resulted in nanostructures with varying biological effects. Therefore, we believe that structural optimization of siRNA nanoplexes is essential in achieving better siRNA encapsulation, protection, and gene silencing efficiency.在本研究中,选择一种双子型阳离子脂材(CLD)为递送载体,用于阐明CLD/siRNA纳米粒(CLD/siRNA NPs)的"复合物特性"。使用不同的工艺方法制备了三种类型的CLD/siRNA纳米粒(AT NPs,HT NPs和MT NPs):AT工艺(siRNA溶液和预先制备好的CLD空白纳米粒混合)、HT工艺(siRNA溶液水化CLD薄膜)和MT工艺(在涡旋作用下,CLD乙醇溶液滴入siRNA溶液),从粒径、电位、形貌、siRNA保护效果、细胞毒性、细胞摄取能力和靶向mRNA的下调作用等方面展开了一系列对比性评价研究。在优化N/P=10/1的条件下,使用不同方法制备的三种CLD/siRNA纳米粒的粒径大小及其顺序是:MT NPs((222.3±19.1)nm)>HT NPs((105.7±1.31)nm)>AT NPs((91.8±1.75)nm)。显然,由此推测,虽然它们具有相同的组成成分,但不同方法制备的三种纳米粒具有不同的纳米组装结构。此外,从TEM图片中可以看出,三种纳米粒具有显著不同的形貌,表明三种纳米粒的不同纳米结构可能源于不同的组装机制。在人宫颈癌Hela细胞中,三种纳米粒的摄取能力不同,顺序如下:AT NPs>MT NPs>HT NPs,而表皮生长因子受体的沉默水平的顺序如下:MT NPs>AT NPs>HT NPs。基于以上不一致的结果,可能的解释是不同制备工艺构建的不同纳米结构决定了相应的生物学效应。因此,我们认为si RNA纳米复合物的结构优化有助于获得更好的siRNA包载保护和基因沉默效果。

关 键 词：SIRNA Delivery NANOPARTICLES Preparation Cellular uptake 

分 类 号：R943[医药卫生—药剂学]