Vascular endothelial growth factor-A gene electrotransfer promotes angiogenesis in a porcine model of cardiac ischemia

Abstract

This study aimed to assess safety and therapeutic potential of gene electrotransfer (GET) as a method for delivery of plasmid encoding vascular endothelial growth factor A (VEGF-A) to ischemic myocardium in a porcine model. Myocardial ischemia was induced by surgically occluding the left anterior descending coronary artery in swine. GET following plasmid encoding VEGF-A injection was performed at four sites in the ischemic region. Control groups either received injections of the plasmid without electrotransfer or injections of the saline vehicle. Animals were monitored for 7 weeks and the hearts were evaluated for angiogenesis, myocardial infarct size and left ventricular contractility. Arteriograms suggest growth of new arteries as early as 2 weeks after treatment in electrotransfer animals. There is a significant reduction of infarct area and left ventricular contractility is improved in GET-treated group compared with controls. There was no significant difference in mortality of animals treated with GET of plasmid encoding VEGF-A from the control groups. Gene delivery of plasmid encoding VEGF-A to ischemic myocardium in a porcine model can be accomplished safely with potential for myocardial repair and regeneration.

Figure 1. Permanent ischemia was induced via LAD occlusion

A) TIMI flow index drops significantly after occlusion, with no significant difference in TIMI between treatment groups (Two-way ANOVA and Tukey’s multiple comparisons test, alpha=0.05, P = <0.0001). B) SPY based measurement of ischemic surface area of the left ventricle indicates consistent occlusion between treatment groups, with no significant difference in ischemic area (One-way ANOVA, alpha = 0.05, P=0.6388). C) SPY based measurement of drop in perfusion in the ischemic area of the left ventricle indicates a consistent reduction in perfusion, with GET group receiving a slightly larger drop in perfusion than the IO group (One-way ANOVA, Tukey’s multiple comparisons test alpha=0.05, P=0.0012).

Figure 2. GET of VEGF-A has no deleterious effects on survival or heart performance over 7 weeks post MI and gene delivery

A) Kaplan-Meier Survival curve with no significant difference between the groups (Logrank test for trend, P=0.4639), B) Ejection fraction (P=0.9240), C) Cardiac Output (P=0.5603), D) Fractional Shortening(P=0.3775), E) Left ventricular end systolic volume (P=0.5603), F) Left Ventricular end diastolic volume(P=0.92), with no significant difference between treatment groups based on echocardiographic analysis (Two-way ANOVA and Tukey’s multiple comparison test, alpha=0.05)

Figure 3. GET of pVEGF-A induces angiogenesis in ischemic myocardium at treatment sites

A) Arteriograms of GET and IO animals, before occlusion, immediately after occlusion and 7 weeks after occlusion, with new arteries visible in the GET heart. *LAD occlusion site, red arrows point to new vessels. B) The proportion of hearts with new arteries visible via arteriogram 2 (P=0.152313) and 7 weeks (P=0.243917) after treatment is higher in hearts treated with GET of pVEGF-A (Chi-square test). C) vWF staining for blood vessels at a treatment site of GET of pVEGF-A seven weeks after MI and treatment. D) Blood vessel density at treatment sites seven weeks after MI and treatment is not significantly different between treatment groups on a microscopic level (One-way ANOVA, alpha=0.05, P=0.9704).

Figure 4. Myocardial infarct size is reduced at GET of pVEGF treatment sites 7 weeks post treatment

A) Infarct area to ischemic area ratio is significantly lower in GET treated left ventricles than in Sham control group, (One-way ANOVA, and Tukey’s multiple comparison test, alpha=0.05, p=0.0349), B) Total myocardial infarct size including untreated septum and untreated right ventricle, with no significant differences between the groups (One-way ANOVA, alpha=0.05, P=0.2028).