Cardiovascular Research Advance Access originally published online on November 5, 2008
Cardiovascular Research 2009 81(2):389-399; doi:10.1093/cvr/cvn304
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Role of neuronal NO synthase in regulating vascular superoxide levels and mitogen-activated protein kinase phosphorylation
1 Department of Physiology II, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521, Japan
2 Department of Pharmacology, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe Miki-cho, Kita-gun, Kagawa 761-0793, Japan
3 Division of Endocrinology and Metabolism, Department of Internal Medicine, Faculty of Medicine, Kagawa University, 1750-1 Ikenobe Miki-cho, Kita-gun, Kagawa 761-0793, Japan
* Corresponding author. Tel: +81 744 29 8829; fax: +81 744 23 4696. E-mail address: gxzhang{at}naramed-u.ac.jp
Aims: The present study is designed to investigate the role of neuronal nitric oxide synthase (nNOS) in the regulation of vascular mitogen-activated protein kinase (MAPK) activity under basal and angiotensin II (Ang II)-stimulated conditions.
Methods and results: Incubation with a potent nNOS inhibitor (L-VNIO) significantly increased superoxide (O2–) levels, with increased MAPK phosphorylation, in isolated aorta and vascular smooth muscle cells (VSMCs) from wild-type mice. Both increases were inhibited by the superoxide dismutase mimetic, tempol, but not by the peroxynitrite scavenger, FeTPPS. The levels of O2– and MAPK phosphorylation were higher in aorta from nNOS–/– mice than from wild-type mice. These parameters were suppressed by tempol and oxypurinal (a xanthine oxidase inhibitor). In isolated VSMCs or aorta from wild-type mice, Ang II stimulation markedly increased the levels of O2– and MAPK phosphorylation. L-VNIO significantly reduced Ang II-induced increases of these parameters. Apocynin, an NAD(P)H oxidase inhibitor, further inhibited Ang II-induced increases of these parameters compared with the L-VNIO-treated group. FeTPPS did not suppress the Ang II-induced increase of O2– levels, but markedly inhibited Ang II-induced MAPK phosphorylation. In contrast to the wild-type, in isolated aorta or VSMCs from nNOS–/– mice, Ang II failed to increase O2– levels and MAPK phosphorylation.
Conclusion: Under basal conditions, nNOS-derived NO acting as antioxidant reduces O2– accumulation and suppresses vascular MAPK phosphorylation. Under Ang II-stimulated conditions, NAD(P)H oxidase-derived O2–, inducing nNOS uncoupling, potentiates the Ang II-induced increase of O2– generation. The generated O2– may react with NO to form peroxynitrite (ONOO–). Both O2– and ONOO– participate in Ang II-induced activation of vascular MAPK.
KEYWORDS Neuronal nitric oxide synthase; Angiotensin II; Superoxide; Mitogen-activated protein kinase (MAPK)
Time for primary review: 32 days