Effect of Mg ions on efficiency of gene electrotransfer and on cell electropermeabilization.
Gene electrotransfer is a promising nonviral method that enables DNA to be transferred into living cells with electric pulses. However, there are many parameters that determine gene electrotransfer efficiency. One of the steps involved in gene electrotransfer is interaction of DNA with the cell membrane. Divalent cations in the electroporative media can influence the anchoring of DNA to the cell membrane and by that gene electrotransfer efficiency. Here we report the effect of different concentrations of Mg2+ on electropermeabilization for small molecule (propidium iodide), gene electrotransfer and viability of the cells. We also used TOTO-1 dye to visualize DNA-cell membrane interaction for different [Mg]. For this purpose, we used different electroporative media with increasing [Mg]. Our study shows that higher [Mg] lead to higher electropermeabilization for propidium iodide and higher viability, while causing lower gene electrotransfer efficiency. Because we observed higher TOTO-1 labeled DNA at cell surface when using higher [Mg], we suggest that Mg2+ ions can bind DNA at cell surface at such strength that cannot pass into the cell during application of electric pulses, which can lead to lower gene transfection. There may also be other mechanisms involved, since there are many steps of gene electrotransfer on which Mg2+ ions can have an effect on. Our results also imply that membrane permeability changes are not sufficient for an efficient gene electrotransfer.
Haberl
Sasa
S
University of Ljubljana, Faculty of Electrical Engineering, Trzaska 25, SI-1000 Ljubljana, Slovenia.
Miklavcic
Damijan
D
Pavlin
Mojca
M
D000818:Animals; D016466:CHO Cells; D002413:Cations, Divalent / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects*; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D004396:Coloring Agents / Q000737:chemistry; D006224:Cricetinae; D003412:Cricetulus; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D018274:Electroporation / Q000379:methods*; D018014:Gene Transfer Techniques; D008274:Magnesium / Q000737:chemistry / Q000494:pharmacology*; D008856:Microscopy, Fluorescence; D011419:Propidium / Q000737:chemistry; D013844:Thiazoles / Q000737:chemistry
D002413:Cations, Divalent; D004396:Coloring Agents; C086329:TOTO-1 thiazole orange dye; D013844:Thiazoles; D011419:Propidium; D004247:DNA; D008274:Magnesium