Optimisation of electrotransfer of plasmid into skeletal muscle by pretreatment with hyaluronidase – increased expression with reduced muscle damage

Long-term expression of full-length human dystrophin in transgenic mdx mice expressing internally deleted human dystrophins

“…30,31 Plasmid DNA is able to accommodate the 14 kb full-length cDNA and has been used to deliver the dystrophin gene into the muscles of mdx mice, 18,32,33 although with a low efficiency. Recently, the use of electroporation to introduce plasmids into muscle has increased gene transfer efficiency, 25 including delivery of full-length dystrophin into the skeletal muscle of mdx mice. 20,34,35 For viral-based delivery, adenoviral vectors have been engineered with deletion of all viral sequences, except for those required for packaging of the virus.…”

“…Mice were anaesthetized by inhalation of isofluorane, the DNA injected and an electrical field of 175 V/cm was applied in 10 20-ms square wave pulses at 1 Hz using a BTX ECM830 electroporator (VWR International, Lutterworth, UK) as described in McMahon et al 25 The left TA received a single dose of 50 mg of dystrophin plasmid DNA in 25 ml normal saline, injected percutaneously using a 27-gauge needle in a proximal to distal direction. The contralateral TA was either left uninjected or injected with the same quantity of CMV0 and served as a control for dystrophin-positive revertant fibres.…”

“…In the current study, plasmid DNA encoding full-length murine or human dystrophin driven by the CMV promoter was electroporated into the tibialis anterior (TA) muscles of mdx mice 2 h after treatment with hyaluronidase, as this procedure has been shown to achieve high transfection efficiencies in muscle. 20,25 We also investigated whether revertant fibres and/or expression of different dystrophin isoforms contribute to the tolerance to murine dystrophin. The role of the other smaller isoforms of dystrophin in inducing tolerance was examined in mdx3cv mice, a strain of the mdx mouse with a mutation in exon 65 and that does not produce any of the dystrophin isoforms.…”

Gene Therapy Progress and Prospects: Electroporation and other physical methods

“…One of the limitations of the electrotransfer is that there can be substantial damage associated with the procedure and that this can limit the efficiency of transfection. 13 A recent paper by Durieux et al 14 convincingly demonstrates that muscle damage is closely associated with the presence of the plasmid DNA during the electrotransfer and is made worse when the LacZ reporter gene is expressed. Modification of the magnitude and duration of the electrical pulses can ameliorate but not entirely prevent all the plasmid-associated damage following treatment of skeletal muscle.…”

Direct electrotransfer of hHGF gene into kidney ameliorates ischemic acute renal failure

“…To improve plasmid DNA diffusion, 25 U of bovine hyaluronidase (Sigma) in 60 ml saline was injected into the muscle 2 h before the administration of plasmid DNA and the electroporation. 43 Kidney electroporation. Left kidney and pedicle were surgically exposed with a midline abdominal incision and the renal artery was clamped proximal to the aorta.…”