In vitro evaluation of synergistic effect of primary and tertiary amino groups in chitosan used as a non-viral gene carrier system

Abstract

The aim of this study is to investigate the synergistic effect of primary and tertiary amino groups of chitosan (Chi) molecule on the transfection efficiency of human embryonic kidney 293 (HEK293) cells. A non-viral gene carrier system was created by synthesizing a modified Chi with a tertiary amino group grafted to its methylol group (CH2OH) (Chi-tAm). The molecule was synthesized using 3-dimethylamino-1-propyl chloride hydrochloride (DAPC) and characterized using Fourier transform infrared spectroscopy (FTIR). The weight average molecular weight (Mw) and polydispersity index (PDIMw) were determined using gel permeation chromatography-size exclusion chromatography (GPC-SEC). The nanoparticles of Chi-tAm (nChi-tAm) were prepared by ionic gelation and were analyzed for particle size (nm), polydispersity (PDIn) and zeta potential (mV). The proton buffering capacity and gene complexing capacity of the nanoparticles were also determined. HEK293 cells were used to evaluate Chi-tAm cytotoxicity via MTT assay. Transfection efficiency and confluency of HEK293 cells after transfection with Chi-tAm were evaluated. All of the results were compared with those of non-modified Chi. The results showed that Chi-tAm could be used as a novel non-viral gene delivery system in the HEK293 cell line due to the unmodified primary amino groups and the grafted tertiary amino groups of Chi. It also allows many further reactions for the modification of Chi-tAm over primary amino groups in future studies.