Abstract:

BACKGROUND: Nanoparticles formulated from biodegradable polymers such as mono methyl poly (ethylene glycol)-g-chitosan are being extensively investigated as non-viral gene delivery systems due to their sustained release characteristics, targeting and biocompatibility.
OBJECTIVE: To prepare mPEG-CS nanoparticles and to explore the feasibility of mPEG-CS as livin shRNA gene carrier, and to study the gene transfection efficiency to colorectal cancer cell HT-29 mediated mPEG-CS nanoparticles.
METHODS: mPEG-CS nanoparticles were prepared by ionic cross-linking method and mPEGylated -chitosan/ livin shRNA complex by Static adsorption. The pattern, size and Zeta potential of blank nanoparticles and mPEGylated-chitosan/ livin shRNA complex were detected by Zeta-size analyzer and transmission electron microscope (TEM). The encapsulation efficiency of gene-nano complex was measured. The protection of nanoparticles for gene was validated through Gel electrophoresis block experiment and DNase I enzyme digestion experiment. The transfection efficiency of livin shRNA was compared to that of CRC cell HT-29 mediated by mPEG-CS nanoparticles or not.
RESULTS AND CONCLUSION: mPEG-CS nanoparticles of the size about 60 nm were successfully prepared out, when nanoparticles /gene volume ratio was 3∶1, size of gene-nano complex was 100 nm and encapsulation efficiency was (94.32±0.35)%. The gel electrophoresis blocking test showed that nanoparticles could be effectively combined with the plasmid, and Dnase I test proved that the nanoparticles could protect the plasmid. Transfection efficiency mediated by mPEG-CS was higher than naked gene and working a longer time. As gene carrier, mPEG-CS nanoparticles can protective gene well, and it is able to transfect livin shRNA recombinant plasmid into colorectal cancer cells and express in a long time. The gene-nano complex overcomes the shortage of relatively short time of RNA interference in gene therapy for tumor.