Efficient electrotransfection into canine muscle.
Two different types of electroporation protocols have been developed for efficient electrotransfer of plasmid DNA into skeletal muscle of experimental animals. At first, only low voltage electric pulses have been used, but lately, a combination of high and low voltage pulses has been suggested as more efficient. Up to date, in dogs, this type of electroporation protocol has never been used for muscle targeted plasmid DNA electrotransfection. In this study, we used two different DNA plasmids, one encoding green fluorescent protein and one encoding human interleukin-12. Five different electroporation protocols were evaluated. Three of them featured different combinations of high and low voltage pulses, and two were performed with delivery of low voltage pulses only. Our study shows that combination of 1 high voltage pulse (600 V/cm, 100 mus), followed by 4 low voltage pulses (80 V/cm, 100 ms, 1 Hz) yielded in the same transfection efficiency as the standard trains of low voltage pulses. However, this protocol is performed quicker and, thus, more suitable for potential use in clinical practice. In addition, it yielded in detectable systemic expression of human interleukin-12. Electrotransfer of either of the plasmids was associated with only mild and transitory local side effects, without clinically detectable systemic side effects. The results indicate that electrotransfection is a feasible, effective, and safe method for muscle targeted gene therapy in dogs, which could have potential for clinical applications in veterinary medicine of small animals.
Pavlin
D
D
University of Ljubljana, Veterinary Faculty Ljubljana, Gerbiceva 60, SI-1000 Ljubljana, Slovenia.
Tozon
N
N
Sersa
G
G
Pogacnik
A
A
Cemazar
M
M
D000818:Animals; D004285:Dogs; D018274:Electroporation / Q000379:methods*; D005260:Female; D006801:Humans; D007371:Interferon-gamma / Q000097:blood; D018664:Interleukin-12 / Q000097:blood; D008297:Male; D018482:Muscle, Skeletal / Q000378:metabolism*; D010957:Plasmids; D014162:Transfection / Q000379:methods*
D018664:Interleukin-12; D007371:Interferon-gamma