Control by osmotic pressure of voltage-induced permeabilization and gene transfer in mammalian cells.
Cells can be transiently permeabilized by a membrane potential difference increase induced by the application of high electric pulses. This was shown to be under the control of the pulsing buffer osmotic pressure, when short pulses were applied. In this paper, the effects of buffer osmotic pressure during electric treatment and during the following 10 min were investigated in Chinese hamster ovary cells subjected to long (ms) square wave pulses, a condition needed to mediate gene transfer. No effect on cell permeabilization for a small molecule such as propidium iodide was observed. The use of a hypoosmolar buffer during pulsation allows more efficient loading of cells with beta-galactosidase, a tetrameric protein, but no effect of the postpulse buffer osmolarity was observed. The resulting expression of plasmid coding for beta-galactosidase was strongly controlled by buffer osmolarity during as well as after the pulse. The results, tentatively explained in terms of the effect of osmotic pressure on cell swelling, membrane organization, and interaction between molecules and membrane, support the existence of key steps in plasmid-membrane interaction in the mechanism of cell electrically mediated gene transfer.
Golzio
M
M
Institut de Pharmacologie et de Biologie Structurale du CNRS UPR 9062, Toulouse, France.
Mora
M P
MP
Raynaud
C
C
Delteil
C
C
Teissié
J
J
Rols
M P
MP
D000818:Animals; D016466:CHO Cells; D002463:Cell Membrane Permeability; D002470:Cell Survival; D006224:Cricetinae; D018274:Electroporation / Q000379:methods; D018014:Gene Transfer Techniques; D008564:Membrane Potentials / Q000502:physiology; D009997:Osmotic Pressure; D010957:Plasmids; D011419:Propidium / Q000493:pharmacokinetics; D011994:Recombinant Proteins / Q000096:biosynthesis; D014162:Transfection / Q000379:methods; D001616:beta-Galactosidase / Q000096:biosynthesis
D011994:Recombinant Proteins; D011419:Propidium; D001616:beta-Galactosidase