ZHANG Wenhua,LIANG Ye,ZHOU Quan,LI Tong,LI Mingyu, MIAO Yu,XU Wenhua,WANG Liping
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Chitosan is a natural cationic polysaccharide, which is often used for preparing biomedical materials because of its high biocompatibility. In this study, chitosan with different molecular weights was chosen to prepare chitosan nanoparticles (CSNPs) as gene vector by ionic cross-linking with tripolyphosphate (TPP). CSNPs were characterized in terms of particle size, zeta potential, polydispersity index (PDI) by the Zetasizer, and morphology was detected with transmission electron microscopy (TEM). Furthermore, the cytotoxicity and biocompatibility of CSNPs were correspondingly examined by MTT assay and the muscle slices of mice. The transfer effect on plasmids and siRNA was evaluated. Agarose gel electrophoresis method and spectrophotometry were used to detect the loading efficiency. The cell transfection efficiency of plasmids or siRNA loaded by CSNPs was analyzed by the fluorescence microscopy, laser scanning confocal microscopy or flow cytometry. The blood compatibility was analyzed by hemolytic experiment and full-automatic blood coagulator. Results showed that when CS(160 kDa) was 1 mg/mL with TTP(mass ratio was 10∶1), CSNPs were prepared successfully by ionic gelation method, which had smaller particle size (about 100 nm), stable dispersibility, high gene loading efficiency and spheroidal particles with morphological rules under TEM. Their cytotoxicity is at the 0-1 level, which is in line with the toxicity standard of biomedical materials. No obvious inflammatory reaction was found in the tissue section, showing good tissue-biocompatibility. CSNPs could transfer the plasmids or siRNA to cells with highloading rate. However, CSNPs might have much higher transfecting efficiency for siRNA than plasmids, which implied that CSNPs might be more suitable for being a safe and efficient vector for delivering siRNA than plasmids.The results of flow cytometry showed that the transfection rate was higher than that of commercial transfection reagents. Results also showed that CSNPs had good blood compatibility, so it can be applied as a safe and efficient siRNA delivery vector in cancer therapies.