title: Antioxidant effect of adrenomedullin on angiotensin II-induced reactive oxygen species generation in vascular smooth muscle cells.
abstract: Recent adrenomedullin (AM) gene-targeting studies have proposed a novel concept that AM plays a protective role against oxidative stress in vivo. The present study was undertaken to explore the underlying molecular mechanism of the putative antioxidant action of AM against angiotensin II (Ang II)induced reactive oxygen species (ROS) generation in rat vascular smooth muscle cells (VSMCs). Intracellular ROS levels were measured by dichlorofluoroscein fluorescence. Redox-sensitive c-Jun amino-terminal kinase (JNK) and ERK1/2 activation and gene expression induced by Ang II in VSMCs were also studied. AM dose-relatedly (10(-8)-10(-7) m) inhibited intracellular ROS generation stimulated by Ang II (10(-7) m), as mimicked by dibutyl-cAMP, the effect of which was inhibited by the pretreatment with N-(2-[p-bromocinnamylamino]ethyl)-5-isoquinolinesulfonamide hydrochloride, a protein kinase A inhibitor, and calcitonin gene-related peptide(8-37), an AM/calcitonin gene-related peptide receptor antagonist. Ang II induced JNK and ERK1/2 activation via a redox-sensitive manner, whereas AM inhibited JNK, but not ERK1/2, activation by Ang II. Furthermore, AM inhibited Ang II-induced redox-sensitive gene expression (plasminogen activator inhibitor-1 and monocyte chemoattractant protein-1) in the same manner as N-acetyl-l-cysteine, a potent antioxidant. AM also inhibited Ang II-induced up-regulation of Nox1, a critical membrane-bound component of reduced nicotinamide adenine dinucleotide phosphate oxidase in VSMCs, in the same degree as N-acetyl-l-cysteine. Our study demonstrates for the first time that AM directly inhibits intracellular ROS generation via an AM receptor-mediated and c-AMP-protein kinase A-dependent mechanism in VSMCs and that AM with its potent antioxidant action inhibits redox-sensitive JNK activation and gene expression induced by Ang II. These data suggest that AM plays a protective role as an endogenous antioxidant in Ang II-induced vascular injury.
authors.affiliation: Department of Clinical and Molecular Endocrinology, Tokyo Medical and Dental University Graduate School, 1-5-45 Yushima, Bunkyo-ku, Tokyo 113-8513, Japan. tyoshimoto.cme@tmd.ac.jp.
authors.forename: Takanobu
authors.initials: T
authors.lastname: Yoshimoto
authors.forename: Nozomi
authors.initials: N
authors.lastname: Fukai
authors.forename: Ryuji
authors.initials: R
authors.lastname: Sato
authors.forename: Toru
authors.initials: T
authors.lastname: Sugiyama
authors.forename: Naoko
authors.initials: N
authors.lastname: Ozawa
authors.forename: Masayoshi
authors.initials: M
authors.lastname: Shichiri
authors.forename: Yukio
authors.initials: Y
authors.lastname: Hirata
mesh_terms: D053607:Adrenomedullin; D000804:Angiotensin II / Q000494:pharmacology*; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001013:Aorta, Thoracic / Q000166:cytology; D002478:Cells, Cultured; D018932:Chemokine CCL2 / Q000235:genetics; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D048031:JNK Mitogen-Activated Protein Kinases; D008297:Male; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics; D000074624:NADPH Oxidase 1; D010084:Oxidation-Reduction; D010455:Peptides / Q000494:pharmacology*; D017395:Plasminogen Activator Inhibitor 1 / Q000235:genetics; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D058265:Receptors, Adrenomedullin; D018000:Receptors, Peptide / Q000378:metabolism; D014662:Vasoconstrictor Agents / Q000494:pharmacology*
chemical_list: D000975:Antioxidants; D018932:Chemokine CCL2; D010455:Peptides; D017395:Plasminogen Activator Inhibitor 1; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D058265:Receptors, Adrenomedullin; D018000:Receptors, Peptide; D014662:Vasoconstrictor Agents; D000804:Angiotensin II; D053607:Adrenomedullin; D009247:NADH, NADPH Oxidoreductases; D000074624:NADPH Oxidase 1; C000614712:NOX1 protein, rat; D017868:Cyclic AMP-Dependent Protein Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases