Alteration of Arterial Vasomotor Function in Vitro by Gene Transfer with a Replication-Deficient Adenovirus

Abstract

Gene transfer technologies offer great potential both to investigate and alter the course of vessel wall pathophysiology. Replication-deficient adenovirus vectors appear particu larly useful for the transfection of blood vessels, because of their ability to accommodate large cDNA inserts and to rapidly and efficiently infect mammalian endothelial and smooth muscle cells. The potential effects of transfection with a replication-deficient adenovirus on vasomotor function have not, however, been described. This study reports gene transfer experiments with a replication-deficient adenovirus containing a cytome galovirus promotor and a nuclear localizing variant of the bacterial β-galactosidase gene. Excised carotid artery segments from 6 rabbits were divided into four segments and infected in pairs for thirty minutes with four viral titers (control [0], 2.5×10⁹, 5×10⁹, and 1×10¹⁰ pfu/mL) at 37°C in serum-free culture media (M199). Expression of β-galac tosidase and in vitro vasomotor function were determined after seventy-two hours' incu bation. Isometric tension studies were performed to examine the response of the vessel segments to the contractile agonist norepinephrine. β-galactosidase was expressed in all vessel segments exposed to the adenovirus. There was a significant difference in the sensitivity to norepinephrine (P < 0.04) of the segments treated with the highest and lowest viral titers (mean ±sem, -log₁₀ [EC₅₀] of 5.98 ±0.09, 5.77 ±0.11, 5.68 ±0.14, and 5.52 ±0.13 for the control, 2.5×10⁹, 5×10⁹, and 1×10¹⁰ plaque-forming units (pfu)/mL groups respectively). Replication-deficient adenovirus vectors can rapidly and efficiently infect rabbit carotid vessels, but there is, however, a dose-dependent alteration of in vitro vessel vasomotor function that may be mediated by a cytotoxic effect of the viral vector. Although further in vivo work is required to verify these results, this effect needs to be taken into account in studies involving adenoviral gene transfer.

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