Sustained inhibition of experimental neointimal hyperplasia with a genetically modified herpes simplex virus

Objective: Reported herein is a potential strategy for sustained smooth muscle cell (SMC) inhibition with a virulence-attenuated herpes simplex virus (HSV). Experiments were conducted in vitro to demonstrate selective SMC cytotoxicity and in vivo to demonstrate reduced neointimal hyperplasia (NIH) in a clinically relevant animal model. Methods: In vitro: Cultured human umbilical artery smooth muscle cells (UASMC) and venous endothelial cells (HUVEC) were exposed to varying multiplicities of infection (MOI) of a γ₁34. 5-deleted HSV-1 virus (R849). Cell survival was assessed at 48 and 72 hours with a colorimetric MTT viability assay. In vivo: New Zealand White rabbit external jugular veins (n = 21) were exposed to R849 (2.5 × 10⁶ pfu/mL) or culture medium at 110 to 120 mm Hg for 10 minutes, then fashioned as vein patches on carotid arteries. Carotid arteries were ligated distally to decrease blood flow and stimulate a hyperplastic response (ultra-low shear stress model). After 2, 4, 12, and 24 weeks, patched segments were perfusion-fixed with glutaraldehyde and morphometrically examined for NIH formation. Results: In vitro: At 48 hours, R849 exhibited preferential cytotoxicity to UASMC compared with HUVEC, with 11% ± 10% of UASMCs and 49% ± 8% of HUVECs surviving after infection with MOI = 25 (P < .05). Higher MOI resulted in poor survival of both cell lines. In vivo: Blood flow was similarly reduced in all animals both at surgery (0.9 ± 0.1 mL/min vs 1.6 ± 0.3 mL/min) and at harvest (2.7 ± 0.4 mL/min vs 2.5 ± 0.5 mL/min). R849-infected patches exhibited markedly less NIH than control patches did at 2 weeks (162 ± 14 μm vs 49 ± 6 μm; P < .05), 4 weeks (190 ± 27 μm vs 67 ± 8 μm; P < .05), and 12 weeks (233 ± 18 μm 113 ± 2 μm; P < .05). Conclusion: The virulence-attenuated HSV strain R849 demonstrates selective cytotoxicity for SMC and is capable of sustained inhibition of NIH in an experimental model of vein graft failure.