Safari S, Zarrintan M H, Soleimani M, Dorkoosh F A, Akbari H, Larijani B, Rafiee Tehrani M
Department of Pharmaceutics, Faculty of Pharmacy, Tabriz University of Medical Sciences, Tabriz, Iran.
Adv Pharm Bull. 2012;2(1):7-16. doi: 10.5681/apb.2012.002. Epub 2012 Feb 15.
Non-viral vectors have been widely proposed as safer alternatives to viral vectors, and cationic polymers have gained increasing attention because they can form self-assembly with DNA. Chitosan is also considered to be a good candidate for gene delivery systems, since it is already known as a biocompatible, biodegradable, and low toxic material with high cationic potential. However, low solubility and transfection efficiency need to be overcome prior to clinical trial. In this work, we focus on alkyl modified chitosan which might be useful in DNA condensing and efficient gene delivery.
N, N- Diethyl N- Methyl (DEMC) and N- Triethyl Chitosan (TEC) were synthesized from chitosan polymer. In order to optimize the polymers for gene delivery, we used FITC-dextran (FD). Then the optimized polymer concentrations were used for gene delivery. Fluorescent microscope was used, in order to evaluate the polymers' efficiency for gene delivery to human embryonic kidney epithelial cells (HEK 293T).
This modification increased chitosan's positive charge, thus these chitosan derivatives spontaneously formed complexes with FD, green fluorescence protein plasmid DNA (pEGFP), red fluorescence protein plasmid DNA (pJred) and fluorescent labeled miRNA .RESULTS gained from fluorescent microscope showed that TEC and DEMC were able to transfer FD, DNA and miRNA (micro RNA) to HEK cell line.
We conclude that these chitosan derivatives present suitable characteristics to be used as non-viral gene delivery vectors to epithelial cells.
非病毒载体已被广泛提议作为病毒载体的更安全替代品，阳离子聚合物因其能与DNA形成自组装而受到越来越多的关注。壳聚糖也被认为是基因递送系统的良好候选者，因为它已被熟知是一种具有高阳离子潜力的生物相容性、可生物降解且低毒的材料。然而，在临床试验之前，需要克服其低溶解性和转染效率的问题。在这项工作中，我们专注于烷基修饰的壳聚糖，其可能在DNA凝聚和高效基因递送中有用。
从壳聚糖聚合物合成N，N - 二乙基N - 甲基（DEMC）和N - 三乙基壳聚糖（TEC）。为了优化用于基因递送的聚合物，我们使用了异硫氰酸荧光素 - 葡聚糖（FD）。然后将优化后的聚合物浓度用于基因递送。使用荧光显微镜来评估聚合物将基因递送至人胚肾上皮细胞（HEK 293T）的效率。
这种修饰增加了壳聚糖的正电荷，因此这些壳聚糖衍生物能自发地与FD、绿色荧光蛋白质粒DNA（pEGFP）、红色荧光蛋白质粒DNA（pJred）和荧光标记的微小RNA形成复合物。荧光显微镜获得的结果表明，TEC和DEMC能够将FD、DNA和微小RNA（miRNA）转移至HEK细胞系。
我们得出结论，这些壳聚糖衍生物具有适合用作上皮细胞非病毒基因递送载体的特性。
