Study Of Cationic Lipids With Polyhydroxyl For Gene Delivery

Gene therapy is a new and revolutionary method for treatment of immunodeficiency ill, tumour and so on. Its curative effect is confirmed by clinic, but high efficiency and low cytotoxicity gene delivery is the key factor to develop the technology. Application of viral vector is limited prodigiously because of their security problem. Non-viral vector has the excellence of low cytotoxicity, non-immunogenicity, without the size of genetic fragments limit, easy preparation and so on. So research is focused on high transfection efficiency and low cytotoxicity non-viral vector.In this work, cationic lipids with polyhydroxyl were designed and synthesized with carbamate-based tertiary amine as precursor. Intermediates and target products were confirmed by MS,1H NMR and 13C NMR, and their purity is satisfied with transfection.The values of critical micelle concentration (cmc), hydrophile-lipophile balance number (HLB), critical array parameters (P) of cationic lipids were researched. The same type of cationic lipids whose carbon chains were 12 showed smallerγcmc and cmc (lipids 6a and 6c), bigger HLB. Liposomes 5a,5b,5c,5d,6a,6b,6c and 6d were prepared using the cationic lipid and the liposome states were studied by TEM and Nano Particle Size and Potential Analyzer.These cationic liposomes had the ability of delaying DNA from agarose gel electrophoresis experiment, so they could combine with DNA and lipoplexes were formed. Size and Zeta potential were tested by ZETASIZER nanoseries Nano-ZS90 and pGFP-N2 was delivered into hela,7721, Hep-2 and Hepa1-6 cells with liposomes as gene delivery. It can be seen from transfection efficiency and cytotoxicity experiments that transfection efficiency of liposomes 5b and 5c for Hela was as good as that of liposome 5a and it was closed to lipo2000, but transfection efficiency of 5d was lower. Transfection efficiency of liposomes 6a,6b,6c and 6d for Hela and 7721 was nearly zero, and it was low for Hepa1-6, but their transfection efficiency was high for Hep-2. Transfection efficiency of 5b and 5c was higher than that of lipo2000 and 5a. Cytotoxicity of liposomes 5b,5c and 5d was low for Hela cells, and it was closed to that of Lipo2000 and 5a. Cytotoxicity of liposomes 6a,6b and 6c was lower than that of lipo2000 for Hepa1-6 cells.Selectivity of cationic lipids with galactose for different cells was confirmed from initial transfection experiment. Hepa1-6 was worse than Hep-2 but better than Hela and 7721 whose transfection efficiency were nearly zero. Targeting selectivity of liposomes 6a,6b,6c and 6d for hepatocyte need be confirmed by more experiments.