Gene Transfection Ability Of Cationic Liposomes And Their Environmental Effects

Gene therapy is expected to be an effective method to treat cancer and other diseases. The success of gene therapy highly depends on the development of effective and secure delivery vectors. Cationic liposome has advantages of safety, easy production, much lower immunotoxicity and effectiveness in gene delivery. Therefore, much attention has been paid to cationic liposome. Research has shown that the complex vector can be used with good prospect. New lipid molecules were synthesized continuously. Research about relationship between chemical structure with higher transfection efficiency and intracellular pathway of liposome would have significance in lipid synthesis and modification.Four types of cationic lipids-quaternary ammonium based-surfactants gene delivery vectors were used to prepare cationic liposomes. They were characterized by TEM, dynamic light scattering and gel retarding. The average particle sizes of liposomes were changed from100to300nm. The liposomes could bind DNA efficiently. The transfection efficiency of liposome/pDNA complexes was investigated by using GFP and luciferase as marker gene, which showed higher transfection efficiency in Hela and Hep-2cells. Structure-activity relationship research showed that lipid of gemini headgroup and12carbon hydrocarbon tails of C3lipid used for gene delivery obtained higher transfection efficiency than monovalent C2and multivalent C4lipids.The novel type of stable ternary complexes was formed by mixing low molecular weight chitosan with cationic liposome/pDNA lipoplex via non-covalent conjugation for the efficient delivery of plasmid DNA. They were characterized by atomic force microscopy (AFM), gel retarding. The complexes could bind pDNA effectively and retard it completely. Lipopolyplexes were in uncompacted spheroids, short rod-and irregular lump. Uncompacted complex morphology would increase gene delivery ability. Liposome/pDNA and chitosan/pDNA weight ratio affected transfection efficiency. Chitosan could decrease cell toxicity of liposomes. Chitosan enhanced gene delivery of cationic liposome effectively with low cell toxicity and new ternary complex would be used as novel non-vial vector.Liposomes C3were marked by NDB-PE, which could enter cell through membrane efficiently. The intracellular trafficking and mechanism of ternary complexes were examined by confocal laser scanning microscopy. Rapid pDNA delivery to the nucleus was obtained after4h transfection by chitosan enhanced cationic liposome. Cationic lipids used in this paper belong to quaternary ammonium compounds (QACs). Most uses of QACs led to their release into wastewater treatment systems or the environment and were harm to organisms in water body. However, little information was available on the ecological properties of lipid-based surfactants. Four series and twelve compounds were used to evaluate their ecological properties. Biological toxicity and biodegradation ability were studied in this paper. Based on environmental considerations, high efficiency and low toxicity molecular design would obtain support form these data.Four series of cationic liposomes and twelve compounds were used to evaluate Hela and MCF-7cell toxicity. Lower cell toxicity was obtained and cell viability was higher than75%. Chitosan could decrease cell toxicity of liposomes. Acute toxicity tests on freshwater algae Scenedusmus obliquus and as well as on saltwater algae Undaria pinnatifida gametophates and luminescent bacteria were carried out to assess the aquatic toxicity of the cationic surfactant based lipid. IC50value of Scenedusmus sp were below2mg/L at48h and72h except for C2-14(72h, IC50was2.69mg/L). Higher concentration lipids could inhibit photosynthesis and POD activity, thus decrease algae survival ability. On Undaria pinnatifida gametophates, lower concentration lipids could stimulate gametophate growth and higher concentration lipids inhibit its growth. C3-12obtained the highest toxicity at48and72h with IC501.59mg/L and8.90mg/L, whereas C2-14obtained the lowest toxicity at48and72h with IC501.85mg/L and16.01mg/L. About luminescent bacteria, Lower dose of lipids increased the growth of bacteria and higher dose inhibited luminescence. Study of relationship between structure and activity showed that for the same head group and similar molecular structure, the toxicity became decreased with longer as the chain length increased. Lipids of multivalent headgroup had higher toxicity than that of monovalent headgroup. The toxicity mainly belonged to charge effects.QACs were persistent in the environment. Biodegradation of lipid vector would be imperative in reducing surfactants concentration in urban wastewater close to the back-ground levels. Ultimate biodegradation was examined by activated sludge method. TOC results showed that the (?)ltimate biodegradation of C3-12, Cl-12, C2-12, and C4-12with the same carbon tails were69.0%,61.1%,73.7%and68.7%. The biodegradation efficiency decreased with hydrophobic chain length increased of the same lipid headgroup.New synthesized lipid molecules had higher gene delivery ability. Novel chitosan/liposome/pDNA ternary complexes significantly improved gene delivery ability. Both types of non-viral gene vector had potential ability used for gene therapy. The study of lipid structure and activity of gene delivery, toxicity, and biodegradation allowed the development of lipid design for high efficiency, low toxicity and high biodegradation efficiency and provided theory basis and reference for lipid synthesis.