Previously, we developed a novel siRNA transfer method to the liver by sequential intravenous injection of poly-L-glutamic acid (PGA) and cationic liposome/siRNA complex (cationic lipoplex). In this study, we examined...Previously, we developed a novel siRNA transfer method to the liver by sequential intravenous injection of poly-L-glutamic acid (PGA) and cationic liposome/siRNA complex (cationic lipoplex). In this study, we examined the effects of the charge ratio (+/-) of cationic liposome/siRNA, molecular weight of PGA and cationic lipid of cationic liposome on the biodistribution of siRNA after sequential injection of PGA plus cationic lipoplex. When 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP)/cholesterol (Chol) lipoplex was intravenously injected into mice, the accumulation of siRNA was mainly observed in the lungs. In contrast, when DOTAP/Chol lipoplex was intravenously injected at 1 min after intravenous injection of PGA, siRNA was largely accumulated in the liver. The charge ratio (+/-) of DOTAP/Chol liposome/siRNA did not affect the biodistribution of siRNA after sequential injection. As regards the molecular weight of PGA, the accumulation of siRNA was observed mainly in the liver after the sequential injection of PGA of 20.5, 38, 64 or 200 kDa plus DOTAP/Chol lipoplex. Furthermore, to examine the effect of cationic lipid of cationic liposome on the biodistribution of siRNA, we prepared other cationic liposomes composed of 1,2-di-O-octadecenyl-3-trimethylammonium propane chloride (DOTMA)/Chol, dimethyldioctade-cylammonium bromide (DDAB)/Chol and O,O’-ditetradecanoyl-N-(α-trimethylammonioacetyl)di-ethanolamine chloride (DC-6-14)/Chol. For the cationic liposomes, the accumulation of siRNA was observed mainly in the liver when their cationic lipoplexes were sequentially injected after injection of PGA into mice. From these findings, sequential injection of PGA plus cationic lipoplex could deliver siRNA efficiently into the liver regardless of the charge ratio (+/-) of lipoplex, lengths of PGA and cationic lipid of liposome.展开更多
To increase the in vivo stability of polycation gene carriers,a pH-sensitive shielding system,γ-benzyl L-glutamate-co-glutamate acid polymer(PGA(60)(60 refers to the molar ratio of glutamate acid in the polymer)),was...To increase the in vivo stability of polycation gene carriers,a pH-sensitive shielding system,γ-benzyl L-glutamate-co-glutamate acid polymer(PGA(60)(60 refers to the molar ratio of glutamate acid in the polymer)),was synthesized and characterized.PGA(60) showed pH sensitivity at about pH 6.0.PGA(60) shielded the positive charge of DNA/PEI(1:1) complexes.Gel retardation assay showed that no DNA-strand exchange with PGA(60) occurred after PGA(60) was added to DNA/PEI complexes at different proportions.MTT cytotoxicity tests demonstrated that neither PGA(60) nor DNA/PEI/PGA(60) ternary complexes had cytotoxicity at the test concentration.The transfection efficiency was improved when the positive charge was partly shielded by PGA(60).Because of the charge repulsion between the surface of cells and ternary complex particles,there was almost no transfection efficiency when the zeta potential of ternary complexes turned to negative.Because of the suitable pH sensitive range,PGA(60) may be a potential shielding system for polycation gene carriers to be used in vivo.展开更多
文摘Previously, we developed a novel siRNA transfer method to the liver by sequential intravenous injection of poly-L-glutamic acid (PGA) and cationic liposome/siRNA complex (cationic lipoplex). In this study, we examined the effects of the charge ratio (+/-) of cationic liposome/siRNA, molecular weight of PGA and cationic lipid of cationic liposome on the biodistribution of siRNA after sequential injection of PGA plus cationic lipoplex. When 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP)/cholesterol (Chol) lipoplex was intravenously injected into mice, the accumulation of siRNA was mainly observed in the lungs. In contrast, when DOTAP/Chol lipoplex was intravenously injected at 1 min after intravenous injection of PGA, siRNA was largely accumulated in the liver. The charge ratio (+/-) of DOTAP/Chol liposome/siRNA did not affect the biodistribution of siRNA after sequential injection. As regards the molecular weight of PGA, the accumulation of siRNA was observed mainly in the liver after the sequential injection of PGA of 20.5, 38, 64 or 200 kDa plus DOTAP/Chol lipoplex. Furthermore, to examine the effect of cationic lipid of cationic liposome on the biodistribution of siRNA, we prepared other cationic liposomes composed of 1,2-di-O-octadecenyl-3-trimethylammonium propane chloride (DOTMA)/Chol, dimethyldioctade-cylammonium bromide (DDAB)/Chol and O,O’-ditetradecanoyl-N-(α-trimethylammonioacetyl)di-ethanolamine chloride (DC-6-14)/Chol. For the cationic liposomes, the accumulation of siRNA was observed mainly in the liver when their cationic lipoplexes were sequentially injected after injection of PGA into mice. From these findings, sequential injection of PGA plus cationic lipoplex could deliver siRNA efficiently into the liver regardless of the charge ratio (+/-) of lipoplex, lengths of PGA and cationic lipid of liposome.
基金supported by the National Natural Science Foundation of China (Grant Nos 20604028,50873102,50733003,A3 Foresight Program 20621140369)Ministry of Science and Technology of China (Grant No 2007DFR5020)
文摘To increase the in vivo stability of polycation gene carriers,a pH-sensitive shielding system,γ-benzyl L-glutamate-co-glutamate acid polymer(PGA(60)(60 refers to the molar ratio of glutamate acid in the polymer)),was synthesized and characterized.PGA(60) showed pH sensitivity at about pH 6.0.PGA(60) shielded the positive charge of DNA/PEI(1:1) complexes.Gel retardation assay showed that no DNA-strand exchange with PGA(60) occurred after PGA(60) was added to DNA/PEI complexes at different proportions.MTT cytotoxicity tests demonstrated that neither PGA(60) nor DNA/PEI/PGA(60) ternary complexes had cytotoxicity at the test concentration.The transfection efficiency was improved when the positive charge was partly shielded by PGA(60).Because of the charge repulsion between the surface of cells and ternary complex particles,there was almost no transfection efficiency when the zeta potential of ternary complexes turned to negative.Because of the suitable pH sensitive range,PGA(60) may be a potential shielding system for polycation gene carriers to be used in vivo.
