In vivo electroporation enhances plasmid-based gene transfer of basic fibroblast growth factor for the treatment of ischemic limb.
BACKGROUND
Angiogenic therapy for ischemic tissues using angiogenic growth factors has been reported on an experimental and a clinical level. Electroporation enhances the efficiency of plasmid-based gene transfer in a variety of tissues. The purpose of this study was to evaluate the angiogenic effects of plasmid-based gene transfer using basic fibroblast growth factor (bFGF) in combination with electroporation.


METHODS
The transfection efficiency of in vivo electroporation in rabbit skeletal muscles was evaluated using pCAccluc+ encoding luciferase. To evaluate the angiogenic effects of bFGF gene in ischemic limb, we constructed a plasmid, pCAcchbFGFcs23, containing human bFGF cDNA fused with the secretory signal sequence of interleukin (IL)-2. Then, 500 microg of pCAcchbFGFcs23 or pCAZ3 (control plasmid) was injected into the ischemic thigh muscles in a rabbit model of hind limb ischemia with in vivo electroporation (bFGF-E(+) group and LacZ-E(+) group). Other sets of animals were injected with pCAcchbFGFcs23 (bFGF-E(-) group) or pCAZ3 (LacZ-E(-) group) without electroporation. Then 28 days later, calf blood pressure ratio, angiographic score, in vivo blood flow, and capillary density in the ischemic limb were measured.


RESULTS
Gene transfer efficiency increased markedly with the increase in voltage up to 100 V. Regarding angiogenic responses, calf blood pressure ratio, in vivo blood flow, and capillary density only in the bFGF-E(+) group were significantly higher than those in LacZ-E(-) group. Angiographic scores in the bFGF-E(+) and bFGF-E(-) groups were significantly higher than that in the LacZ-E(-) group.


CONCLUSIONS
These data suggest that in vivo electroporation enhances bFGF gene transfer for the treatment of ischemic limb muscles.
Nishikage
Seiji
S
Department of Vascular Regeneration, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.
Koyama
Hiroyuki
H
Miyata
Tetsuro
T
Ishii
Shigeyuki
S
Hamada
Hirohumi
H
Shigematsu
Hiroshi
H
D000818:Animals; D018274:Electroporation / Q000379:methods*; D016222:Fibroblast Growth Factor 2 / Q000096:biosynthesis / Q000235:genetics*; D018014:Gene Transfer Techniques; D006614:Hindlimb / Q000098:blood supply*; D006801:Humans; D007511:Ischemia / Q000628:therapy*; D008297:Male; D023421:Models, Animal; D018919:Neovascularization, Physiologic / Q000235:genetics / Q000502:physiology; D010957:Plasmids; D011817:Rabbits
D016222:Fibroblast Growth Factor 2