Gene transfer by cationic surfactants is essentially limited by the trapping of the surfactant/DNA complexes onto the cell membrane: a fluorescence investigation

The interaction between complexes of plasmid DNA with cetyltrimethylammoniu m bromide (CTAB) and L929 fibroblasts was first examined using confocal mic roscopy. The complexes labeled with the DNA intercalator, YOYO-1, were foun d to be trapped onto the external face of the plasma membrane; a feature th at may constitute a major limiting step in transfection. Moreover, since no cytotoxic effect appeared in these conditions, we further inferred that th e CTAB molecules remained bound to the DNA. The interaction of the complexe s with the membranes was best modeled with neutral vesicles. From anisotrop y thermotropic curves of DPHpPC-labeled vesicles and fluorescence resonance energy transfer measurements between these vesicles and YOYO-labeled compl exes, we evidenced that the binding of the complexes to the vesicle surface opened the micelle-like domains and unwound DNA. However, DNA was not rele ased but remained stably bound via electrostatic interactions to the CTAB m olecules incorporated in the external liposome leaflet. Consequently, the l arge diameter of the unwound plasmid DNA is likely the major factor that pr ecludes its internalization into the cells by endocytosis. In contrast, ani onic vesicles that mimic the cytoplasmic facing monolayer of the plasma mem brane rapidly released DNA from the complex. This may explain the previousl y reported high transfection efficiency of DNA complexed with liposomes com posed of neutral lipids and cationic surfactants, since the latter may dest abilize the endosomal membrane and induce the release of DNA in the cytopla sm. (C) 2000 Elsevier Science B.V. All rights reserved.