Objective: To experimentally investigate direct intramuscular gene transfer of nanoparticles encoding vascular endothelial growth factor for the treatment of peripheral artery disease.
Methods: The human VEGF(165) cDNA was cloned into the eukaryotic expression vector PIRES2 under the control of cytomegalovirus promoter/enhancer. The recombinant gene was transferred into a rabbit model of chronic hindlimb ischemia by naked plasmid and nanoparticle respectively. Ischemia was induced in the hindlimb of New Zealand White rabbits by ligation of the distal external iliac artery and complete excision of the femoral artery and all its branches. At day 7 postoperation animals received VEGF(165) plasmid (10 intramuscular) or nanoparticle-VEGF(165) (8 intramuscular). With RT-PCR, immunohistochemistry analysis, and angiography, the expression and biological effects of VEGF(165) gene in experimental animals were investigated.
Results: Two weeks after initiation of therapy, angiography showed that the transfer of VEGF(165) gene stimulated the formation of focal neovessels and established collateral circulation. The adductor muscle of ischemic limbs was histologically examined at day 14. Capillary density was increased among VEGF(165)-transfected rabbits, especially Nano-VEGF(165)-treated animals (Naked VEGF(165) plasmid = 50.18 per mm(2), Nano-VEGF(165) = 81.22 per mm(2), Control = 29.54 per mm(2), P < 0.05). RT-PCR showed that the transcription and expression of VEGF(165) gene in experimental group were significantly higher than those of control groups.
Conclusions: Intramuscular administration of VEGF(165) induces collateral artery augmentation in the rabbit model of chronic limb ischemia. Nanoparticle can act as a vector to transfect specific gene and it will benefit gene transfer.