Electromediated formation of DNA complexes with cell membranes and its consequences for gene delivery

• Published in: Biochimica et biophysica acta 2011-06, Vol.1808 (6), p.1538-1543
• Main Authors: Escoffre, Jean-Michel, Portet, Thomas, Favard, Cyril, Teissié, Justin, Dean, David S., Rols, Marie-Pierre
• Format: Article
• Language: English
• Subjects: Algorithms; Animals; Biotechnology; Cell Membrane - metabolism; Cellular Biology; Chemical Sciences; CHO Cells; Cricetinae; Cricetulus; DNA - genetics; DNA - metabolism; DNA transfer; Electroporation; Electroporation - methods; Fluorescence microscopy; Fluorescence Resonance Energy Transfer; Gene Transfer Techniques; Green Fluorescent Proteins - genetics; Green Fluorescent Proteins - metabolism; Life Sciences; Microscopy, Confocal; Modeling; Models, Genetic; Physics; Plasmids - genetics; Plasmids - metabolism; Time Factors
• ISSN: 0005-2736; 0006-3002; 1879-2642
• DOI: 10.1016/j.bbamem.2010.10.009

Electroporation is a physical method to induce the uptake of therapeutic drugs and DNA, by eukaryotic cells and tissues. The phenomena behind electro-mediated membrane permeabilization to plasmid DNA have been shown to be significantly more complex than those for small molecules. Small molecules cross the permeabilized membrane by diffusion whereas plasmid DNA first interacts with the electropermeabilized part of the cell surface, forming localized aggregates. The dynamics of this process is still poorly understood because direct observations have been limited to scales of the order of seconds. Here, cells are electropermeabilized in the presence of plasmid DNA and monitored with a temporal resolution of 2ms. This allows us to show that during the first pulse application, plasmid complexes, or aggregates, start to form at distinct sites on the cell membrane. FRAP measurements show that the positions of these sites are remarkably immobile during the application of further pluses. A theoretical model is proposed to explain the appearance of distinct interaction sites, the quantitative increase in DNA and also their immobility leading to a tentative explanation for the success of electro-mediated gene delivery. ► DNA/membrane interaction occurs during cell electrotransfection. ► This interaction is imaged at unprecedent time resolution (ms). ► Observations are consistent with our theoretical model. ► DNA aggregates are formed in the vicinity of or in the plasma membrane. ► Successive electric pulses always lead to accumulation of DNA exactly at the same location than after the first pulse.