Evaluation of the intracellular trafficking of siRNAs in A375 cells by confocal laser scanning microscopy  

利用共聚焦显微镜研究siRNA/CLDs复合物的胞内分布（英文）

作　　者：Yiping Diao Jing Sun Mengyi Yang Bo Xu Lihe Zhang Zhenjun Yang 刁怡萍;孙晶;杨梦依;徐波;张礼和;杨振军(北京大学医学部药学院天然药物及仿生药物国家重点实验室,北京100191)

机构地区：State Key Laboratory of Natural and Biomimetic Drugs, School of Pharmaceutical Sciences, Peking University Health Science Center, Beijing 100191, China

出　　处：《Journal of Chinese Pharmaceutical Sciences》2016年第12期859-868,共10页中国药学（英文版）

基　　金：Ministry of Science and Technology of China(Grant No.2012AA022501,2012CB720604);the National Natural Science Foundation of China(Grant No.20932001,81302626)

摘　　要：Investigation intracellular trafficking of siRNAs following their delivery to cells is of great interest to elucidate dynamics of siRNA in cytoplasm. In this study, we present a novel confocal laser scanning microscopy （CLSM） method to evaluate a novel delivery system of 3＇-peptide-siRNA therapeutic, which was named 3＇-pAs-siRNA/CLD. This method could not only calculate the content of the intracellular 3＇-peptide-siRNA, but also quantify its co-localization with cellular substructure. We observed that 3＇-pAs-siRNA/CLD, which provided the better antitumor capability, also had a better cell uptake, endosome escape and a longer retention time in A375. This novel strategy was proved to be efficient, quantified and visualized, thus making the dynamics research of siRNA in cytoplasm clear and simplified.研究siRNA在胞内的运输过程有助于阐明其在细胞质基质中的动力学性质及过程。本文报道了激光共聚焦扫描显微镜技术用于研究siRNA及其肽缀合物与胱氨酸骨架阳离子脂质体递送系统(siRNA/CLD)的胞内分布过程。发现siRNA在胞质内分布动力学与3′-单肽-siRNA缀合物/CLD递送系统的入胞途径紧密相关,其中具有较好抗肿瘤活性的3′-p As-siRNA/CLD,具有较快的细胞摄取及溶媒体逃逸过程,并能在A375细胞中潴留较长的时间。该方法高效、可定量并可视化,不仅能够测定siRNA在胞内的浓度,也能够定量分析其与细胞亚结构的共定位关系,进一步可系统研究siRNA及其缀合物在胞质内的动力学分布过程。

关 键 词：siRNA/CLD complex Endosomal escape Cytoplasmic distribution Confocal laser scanning microscopy 

分 类 号：R91[医药卫生—药学]