Studies of in vivo Gene Transfer by Electropulsation at the Low Skin Resistance Points (LSRPs)

Abstract

Electropulsation has been proved to be a powerful technique for DNA transfection in vitro. This study was undertaken to assess the feasibility and evaluate the potential of applying this approach for facilitating gene transfer in vivo. The pCMVβ plasmid, containing the Escherichia coli lacZ open reading frame under the cytomegalovirus early promoter, was complexed with poly-L-lysine (PLL) and injected directly into the livers of Wistar rats; electropulsation was then applied either at the low skin resistance points (LSRPs) of the skin, the non-LSRP, or directly at the liver by either a portable electrostimulator or an electropulsation unit. The locations of the LSRPs were detected by the unijunction transistor (UJT) relaxation oscillator. Seventy-two hours after gene transfer, the animals were sacrificed and the livers were excised, followed by examination of the transgene expression. The results obtained from X-gal staining, in situ hybridization, and Western blot analysis indicated that lacZ gene was expressed in the livers. Variations of the electropulsation parameters, including site of electropulsation, wave type, amplitude of the pulses, repetition rate, duty cycle, and duration of electropulsation treatment, do not reveal any statistically significant effects of differences of these variables on the transgene expression. Comparison of the results obtained from direct injection of PLL/pCMVβ (1:1) followed by electropulsation at the LSRPs with that of direct PLL/pCMVβ (1:1) injection, however, suggests that gene transfer by electropulsation achieved a higher transfection efficiency in vivo (p >. 05).