Overexpression of Human Endothelial Nitric Oxide Synthase in Rat Vascular Smooth Muscle Cells and in Balloon-Injured Carotid Artery

From the Departments of Surgery and Pathology (L.C., G.D., R.F., M.C., A.W.C.), University of Washington, Seattle, and Medizinische Universitätsklinik Würzburg (U.W.), Institut für Klinische Biochemie und Pathobiochemie, Würzburg, Germany.

Correspondence to Lihua Chen, Department of Surgery, Box 356410, University of Washington, Seattle, WA 98195. E-mail lihua{at}u.washington.edu

Abstract—Endothelial cells in normal blood vessels might prevent the unscheduled proliferation of smooth muscle cells (SMCs) by the expression of cell migration and growth inhibitors. NO, a potent vasodilator, generated by endothelium-specific constitutive NO synthase (ecNOS) might be such an inhibitor. To test this hypothesis, we overexpressed human ecNOS in syngeneic rat arterial SMCs using retrovirus-mediated gene transfer. Compared with SMCs transduced with vector alone (LXSN SMCs), DNA synthesis and cell proliferation were inhibited in the ecNOS-expressing SMCs (LCNSN SMCs). Basal and stimulated (by the calcium ionophore A23187) secretion of NO and intracellular cGMP were increased in LCNSN SMCs. N-Nitro-L-arginine (L-NA), an inhibitor of NO synthesis, enhanced the proliferation of LCNSN SMCs but had no effect on LXSN SMCs. LCNSN SMCs seeded onto the luminal surface of balloon-injured rat carotid arteries inhibited neointimal formation by 37% and induced marked dilatation (3-fold increase in vessel diameter) at 2 weeks compared with LXSN SMC–seeded arteries. Orally administered L-NA blocked these changes. Phosphorylation of vasodilator-stimulated phosphoprotein, which is regulated in part by NO, was elevated in LCNSN SMCs and in LCNSN SMC–seeded arteries. This study demonstrates that NO generation by ecNOS inhibits SMC proliferation in vitro and modulates vascular tone locally in vivo.

Key Words: nitric oxide • endothelial nitric oxide synthase • proliferation • vasodilatation • vasodilator-stimulated phosphoprotein

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