Effect of adenoviral titer and instillation pressure on gene transfer efficiency to arterial and venous grafts ex-vivo

Abstract

Objective: Adenoviral-mediated gene transfer to arterial and venous grafts has potential in the treatment of a number of vascular diseases. Despite widespread use of these vectors to mediate gene transfer to blood vessel walls, the optimal transduction conditions for each type of vessel has yet to be determined. Our objective was to study the effect of adenoviral titer and instillation pressure on efficiency of gene transfer to arterial and venous grafts ex-vivo. Methods: Jugular vein and carotid artery segments of 8 cm were harvested from Yorkshire Cross pigs. Tissue culture media or different titers of an adenoviral vector encoding human placental alkaline phosphatase (hpAP) were instilled into venous and arterial grafts at 0 mm Hg or 80 to 100 mm Hg of pressure and bathed externally in the same solution at 37° C for 30 minutes. The grafts were rinsed, opened longitudinally, and incubated in culture media at 37° C for 48 hours. Grafts were fixed and stained for hpAP transgene expression to quantitate percent luminal transduction or homogenized for alkaline phosphatase (AP) activity to determine total transmural transduction. Results: For venous grafts, the percent luminal area stained for hpAP was greatest with 10 8 plaque-forming units/mL at 0 mm Hg (81% ± 7%) and decreased with increasing titers (53% ± 9% at 10 9 pfu/mL and 44% ± 11% at 5 × 10 9 pfu/mL; n = 7; P < .05). No increase in percent luminal area stain was achieved with an instillation pressure of 80 to 100 mm Hg at any viral titer. The inverse finding was observed in arterial grafts. For arterial grafts, the greatest percent luminal area stained was achieved with 5 × 10 9 pfu/mL at 80 to 100 mm Hg (76% ± 7%). An instillation pressure of 80 to 100 mm Hg increased the percent luminal area stained at 10 8 pfu/mL from 31% ± 9% to 66% ± 8% (n = 8; P = .01). For venous grafts, total AP activity peaked with 10 9 pfu/mL at 0 mm Hg and decreased with an instillation pressure of 80 to 100 mm Hg (30.6 ± 9.7 U/mg versus 10.9 ± 2.5 U/mg; n = 7; P < .01). However, for arterial grafts, total AP activity peaked with 5 × 10 9 pfu/mL (0 mm Hg) and increased with an instillation pressure of 80 to 100 mm Hg (32.8 ± 9.9 U/mg versus 63.4 ± 20.5 U/mg; n = 8; P < .05). Conclusion: High transduction efficiency can be achieved with adenoviral-mediated gene transfer of arterial and venous grafts. Gene transfer with the vascular graft's physiologic pressure conditions improved transduction efficiency for the artery (80 to 100 mm Hg) and vein (0 mm Hg). Comprehensive analysis of adenoviral transduction conditions is important to realize the full promise of adenoviral-mediated gene transfer. (J Vasc Surg 2002;36:263-70.)