Development of chitosan nanoparticles for cell type specific plasmid dna delivery

Abstract

Chitosan is considered to be a good candidate for gene delivery since it is biocompatible, biodegradable and exhibits low toxicity against cells. Chitosans are cationic polymers with component of amino group and can self-assemble with plasmid into polyelectrolyte complexes. The chitosan-DNA nanoparticles (CNP-DNA complex) enter cells by endocytosis and are released from endosome into the nucleus, leading to transcription and translation of genes. The aim of this study was to prepare CNP-DNA complex using a complex coacervation process and to construct a cell-type specific DNA delivery system. The transfection efficiency of CNP-DNA complex with an polymer/DNA (N/P) ratios of 6:1 for human embryonic kidney cell line, 293T, was 55%. For murine macrophage cell line, Raw 264.7, the N/P ratio of CNP-DNA complex showed the highest transfection efficicey of 3.5%, in comparison to commercial transfection reagent, FUGENE HD, which yielded around 19.3% of transfection. The CNP-DNA complex particle size was within the range of 350–650 nm with a surface charge ranging from +6 to +11 mV at various N/P ratios, whereas the particle size of CNP-DNA complex was approximately of 125 nm as measured by scanning electron microscope. To modify chitosan with strepavidin, the chitosan was reacted with sodium meta-periodate and streptavidin hydrazide. The streptavidin CNP-DNA complex have a particle size of 290 nm with a surface charge ranging from -31 mV. The streptavidin CNP-DNA complex particle size was approximately of 134 nm and 350 nm by scanning electron microscope and transmission electron microscopy. Nevertheless the streptavidin CNP-DNA complex which was bound to F4/80 biotin, could mediate the expression of green fluorescent protein in RAW 264.7 cells at a noticeable level.