Adenoviral-mediated gene transfer of a constitutively active form of the retinoblastoma gene product attenuates neointimal thickening in experimental vein grafts

Purpose: Inappropriate or excessive vascular smooth muscle cell proliferation leads to the development of occlusive lesions in up to 50% of vein grafts. The purpose of this study was to test the hypothesis that induced overexpression of a cytostatic nonphosphorylatable form of the retinoblastoma protein (ΔRb) would attenuate neointimal thickening in experimental vein grafts. Methods: A replication-deficient adenovirus vector that encoded a nonphosphorylatable, constitutively active form of ΔRB was constructed (AdΔRb) and contained an NH₂-terminal epitope tag from the influenza hemagglutinin molecule (HA). Forty-eight male New Zealand white rabbits underwent surgical exposure of the external jugular vein for transfection with either 3 x 10¹⁰ plaque-forming units/mL AdΔRb (n = 16), 3 x 10¹⁰ plaque-forming units/mL control adenovirus (AdBglII, n = 15), or vehicle (n = 17) for 10 minutes at 120 mm Hg. After vector exposure, the vein was excised and interposed end-to-end into the carotid circulation. After 5 days, 12 grafts (four from each group) were excised and assayed for genomic ΔRb DNA with the polymerase chain reaction or for hemagglutinin molecule expression and localization with immunohistochemistry. The remainder of the grafts (n = 36) were perfusion-fixed after 4 weeks, and 5 μm sections prepared for digital planimetric analysis. Results: Polymerase chain reaction results identified the ΔRb gene only in the grafts that were transfected with AdΔRb. Immunohistochemical analysis results revealed transgene expression in most of the endothelial cells and in many of the smooth muscle cells. After 4 weeks, the grafts that were exposed to AdΔRb exhibited a 22% reduction in neointimal thickness (vehicle, 77 ± 7 μm; AdBglII, 75 ± 5 μm; AdΔRb, 60 ± 5 μm; P = .05), and medial thickness, luminal diameter, and other parameters were unchanged (medial thickness: vehicle, 72 ± 10 μm; AdBglII, 85 ± 7 μm; AdΔRb, 69 ± 9 μm; P = NS; luminal diameter: vehicle, 4.5 ± 0.2 mm; AdBglII, 4.4 ± 0.2 mm; AdΔR, 4.7 ± 0.1 mm; P = NS). Conclusion: With this delivery system, adenoviral-mediated gene transfer is highly efficient and induced overexpression of ΔRb leads to a reduction in vein graft neointimal thickening.