title: Carbon monoxide releasing molecule induces endothelial nitric oxide synthase activation through a calcium and phosphatidylinositol 3-kinase/Akt mechanism.
abstract: The production of nitric oxide (NO) by endothelial NO synthase (eNOS) plays a major role in maintaining vascular homeostasis. This study elucidated the potential role of carbon monoxide (CO)-releasing molecules (CORMs) in NO production and explored the underlying mechanisms in endothelial cells. We observed that 25μM CORM-2 could increase NO production and stimulate an increase in the intracellular Ca2+level. Furthermore, ethylene glycol-bis(β-aminoethyl ether)-N,N,N',N'-tetra acetic acid caused CORM-2-induced NO production, which was abolished by 1,2-bis(2-aminophenoxy) ethane-N,N,N',N'-tetraacetic acid tetraacetoxy-methyl ester (BAPTA-AM), indicating that intracellular Ca2+release plays a major role in eNOS activation. The inhibition of the IP3 receptor diminished the CORM-2-induced intracellular Ca2+increase and NO production. Furthermore, CORM-2 induced eNOS Ser1179phosphorylation and eNOS dimerization, but it did not alter eNOS expression. CORM-2 (25μM) also prolonged Akt phosphorylation, lasting for at least 12h. Pretreatment with phosphatidylinositol 3-kinase inhibitors (wortmannin or LY294002) inhibited the increases in NO production and phosphorylation but did not affect eNOS dimerization. CORM-2-induced eNOS Ser1179phosphorylation was intracellularly calcium-dependent, because pretreatment with an intracellular Ca2+chelator (BAPTA-AM) inhibited this process. Although CORM-2 increases intracellular reactive oxygen species (ROS), pretreatment with antioxidant enzyme catalase and N-acetyl-cysteine did not abolish the CORM-2-induced eNOS activity or phosphorylation, signifying that ROS is not involved in this activity. Hence, CORM-2 enhances eNOS activation through intracellular calcium release, Akt phosphorylation, and eNOS dimerization.
authors.affiliation: Department of Microbiology, Immunology and Biopharmaceuticals, National Chiayi University, Chiayi 600, Taiwan; Department of Ophthalmology, Chiayi Christian Hospital, Chiayi, Taiwan.
authors.forename: Po-Min
authors.initials: PM
authors.lastname: Yang
authors.affiliation: Department of Microbiology, Immunology and Biopharmaceuticals, National Chiayi University, Chiayi 600, Taiwan.
authors.forename: Yu-Ting
authors.initials: YT
authors.lastname: Huang
authors.affiliation: Department of Microbiology, Immunology and Biopharmaceuticals, National Chiayi University, Chiayi 600, Taiwan.
authors.forename: Yu-Qi
authors.initials: YQ
authors.lastname: Zhang
authors.affiliation: Department of Microbiology, Immunology and Biopharmaceuticals, National Chiayi University, Chiayi 600, Taiwan.
authors.forename: Chia-Wen
authors.initials: CW
authors.lastname: Hsieh
authors.affiliation: Department of Microbiology, Immunology and Biopharmaceuticals, National Chiayi University, Chiayi 600, Taiwan. Electronic address: bswung@mail.ncyu.edu.tw.
authors.forename: Being-Sun
authors.initials: BS
authors.lastname: Wung
mesh_terms: D000730:Androstadienes / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002248:Carbon Monoxide / Q000378:metabolism*; D002417:Cattle; D002867:Chromones / Q000494:pharmacology; D004533:Egtazic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D042783:Endothelial Cells / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism; D009025:Morpholines / Q000494:pharmacology; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism*; D009942:Organometallic Compounds / Q000494:pharmacology*; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D013997:Time Factors; D000077191:Wortmannin
chemical_list: D000730:Androstadienes; D002867:Chromones; D009025:Morpholines; D009942:Organometallic Compounds; D017382:Reactive Oxygen Species; C447082:tricarbonyldichlororuthenium (II) dimer; C070379:1,2-bis(2-aminophenoxy)ethane N,N,N',N'-tetraacetic acid acetoxymethyl ester; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D004533:Egtazic Acid; D002248:Carbon Monoxide; D052250:Nitric Oxide Synthase Type III; D058539:Phosphatidylinositol 3-Kinase; D051057:Proto-Oncogene Proteins c-akt; D002118:Calcium; D000077191:Wortmannin
keywords: Akt; Calcium; Carbon monoxide; Endothelial nitric oxide synthase