ABSTRACT
Noticeable modifications of in-serum transfection efficiency of dioctadecylamidoglycyl-spermine (DOGS)-DNA complexes are observed, depending on DNA condensation conditions. The structures of the complexes are studied, keeping in mind the variability of lipid polymorphism, by cryo-transmission electron microscopy and X-ray diffraction. By increasing both pH and ionic strength, well-organised lamellar structures with a period of 65 A replace supramicellar aggregates. A relationship between the structures and their in-vitro transfection activity is established. Efficiency in the presence of serum is maintained when a lamellar arrangement is involved.
Subject(s)
DNA/chemistry , Fetal Blood , Lipids/chemistry , Lipids/genetics , Transfection , 3T3 Cells , Animals , Cations , DNA/blood , DNA/ultrastructure , Ethanol , Glycine/analogs & derivatives , Glycine/chemistry , Lipids/blood , Mice , Microscopy, Electron , Sodium Chloride , Spermine/analogs & derivatives , Spermine/chemistry , X-Ray DiffractionABSTRACT
Gene therapy is based on the vectorization of genes to target cells and their subsequent expression. Cationic amphiphile-mediated delivery of plasmid DNA is the nonviral gene transfer method most often used. We examined the supramolecular structure of lipopolyamine/plasmid DNA complexes under various condensing conditions. Plasmid DNA complexation with lipopolyamine micelles whose mean diameter was 5 nm revealed three domains, depending on the lipopolyamine/plasmid DNA ratio. These domains respectively corresponded to negatively, neutrally, and positively charged complexes. Transmission electron microscopy and x-ray scattering experiments on complexes originating from these three domains showed that although their morphology depends on the lipopolyamine/plasmid DNA ratio, their particle structure consists of ordered domains characterized by even spacing of 80 A, irrespective of the lipid/DNA ratio. The most active lipopolyamine/DNA complexes for gene transfer were positively charged. They were characterized by fully condensed DNA inside spherical particles (diameter: 50 nm) sandwiched between lipid bilayers. These results show that supercoiled plasmid DNA is able to transform lipopolyamine micelles into a supramolecular organization characterized by ordered lamellar domains.
