Mechanism of in vivo DNA transport into cells by electroporation: electrophoresis across the plasma membrane may not be involved.
BACKGROUND
Recently, in vivo gene transfer with electroporation (electro-gene transfer) has emerged as a leading technology for developing nonviral gene therapies and nucleic acid vaccines. The widely hypothesized mechanism is that electroporation induces structural defects in the membrane and provides an electrophoretic force to facilitate DNA crossing the permeabilized membrane. In this study, we have designed a device and experiments to test the hypothesis.


METHODS
In this study, we have designed a device that alternates the polarity of the applied electric field to elucidate the mechanism of in vivo electro-gene transfer. We also designed experiments to challenge the theory that the low-voltage (LV) pulses cannot permeabilize the membrane and are only involved in DNA electrophoresis, and answer the arguments that (1) the reversed polarity pulses can cause opposing sides of the cell membrane to become permeabilized and provide the electrophoresis for DNA entry; or (2) once DNA enters cytoplasmic/endosomal compartments after electroporation, it may bind to cellular entities and might not be reversibly extracted. Thus a gradual buildup of the DNA in the cell still seems quite possible even under the condition of the rapid reversal of polarity.


RESULTS
Our results indicate that electrophoresis does not play an important role in in vivo electro-gene transfer.


CONCLUSIONS
This study provides new insights into the mechanism of electro-gene transfer, and may allow the definition of newer and more efficient conditions for in vivo electroporation.
Liu
Feng
F
School of Pharmacy, University of North Carolina, Chapel Hill, NC 27599, USA. fliu@email.unc.edu
Heston
Steve
S
Shollenberger
Lisa M
LM
Sun
Bin
B
Mickle
Marlin
M
Lovell
Michael
M
Huang
Leaf
L
D000818:Animals; D002462:Cell Membrane / Q000502:physiology*; D003587:Cytomegalovirus / Q000235:genetics; D004247:DNA / Q000493:pharmacokinetics*; D004586:Electrophoresis; D018274:Electroporation; D018014:Gene Transfer Techniques; D049409:Luciferases, Firefly; D051379:Mice; D010539:Permeability; D010957:Plasmids
D004247:DNA; D049409:Luciferases, Firefly