Cytochrome P-450 Metabolites Mediate Norepinephrine-Induced Mitogenic Signaling

Abstract

Norepinephrine (NE) stimulates release of arachidonic acid (AA) from tissue lipids in blood vessels, which is metabolized via cyclooxygenase, lipoxygenase (LO), and cytochrome P-450 (CYP-450) pathways to biologically active products. Moreover, NE and AA have been shown to stimulate proliferation of vascular smooth muscle cells (VSMCs) of rat aorta. The purpose of this study was to determine the possible contribution of AA and its metabolites to NE-induced mitogenesis in VSMCs of rat aorta and the underlying mechanism of their actions. NE (0.1 to 10 μmol/L) increased DNA synthesis as measured by [ 3 H]thymidine incorporation in VSMCs, and this effect was attenuated by inhibitors of CYP-450 (17-octadecynoic acid, 5 μmol/L; 12-diabromododec-11-enoic acid, 10 μmol/L; and dibromo-dodecenyl-methylsulfimide, 10 μmol/L) and by the LO inhibitor (baicalein, 20 μmol/L), but not by the cyclooxygenase inhibitor (indomethacin, 5 μmol/L). CYP-450 and LO metabolites of AA, 20-hydroxyeicosatetraenoic acid (HETE) (0.1 to fimide, 10 μmol/L) and by the LO inhibitor (baicalein, 20 μmol/L), but not by the cyclooxygenase inhibitor (indomethacin, 5 μmol/L). CYP-450 and LO metabolites of AA, 20-hydroxyeicosatetraenoic acid (HETE) (0.1 to 0.5 μmol/L) and 12(S)-HETE, respectively, increased [ 3 H]thymidine incorporation in VSMCs. Both NE and 20-HETE increased mitogen activated protein (MAP) kinase activity as measured by the in-gel kinase assay. The inhibitor of MAP kinase kinase, PD-98059 (50 μmol/L), attenuated NE as well as 20-HETE induced [ 3 H]thymidine incorporation and MAP kinase activation in VSMCs. These data suggest that products of AA formed via CYP-450, most likely 20-HETE, and via LO mediate NE induced mitogenesis in VSMCs.