title: Nitric oxide activates PI3-K and MAPK signalling pathways in human and rat vascular smooth muscle cells: influence of insulin resistance and oxidative stress.
abstract: OBJECTIVE
  Vascular smooth muscle cells (VSMCs) from the animal model of insulin resistance obese Zucker rats (OZR) show impaired ability of nitric oxide (NO) to increase cGMP and of cGMP to activate its specific kinase PKG, these defects being attributable to oxidative stress. We aimed to investigate the intracellular signalling downstream PKG in human and rat VSMC, and to clarify whether it is modified by insulin resistance and oxidative stress.
  
  
  METHODS
  In aortic VSMC from humans, lean Zucker rats (LZR) and OZR, we measured by Western blots the activation induced by NO and cGMP of signalling molecules of PI3-K and MAPK pathways, with or without PKG inhibition, hydrogen peroxide and antioxidants. We explored the mechanism of the increased oxidative stress in VSMC from OZR by measuring superoxide anion concentrations (luminescence method) with or without inhibition of NADPH oxidase, xanthine oxidase, and mitochondrial electron transport chain complex and by measuring superoxide dismutase (SOD) expression (Western blot) and activity.
  
  
  RESULTS
  In VSMC from humans and LZR, the NO/cGMP/PKG pathway activates both PI3-K (Akt, mTOR) and MAPK (ERK-1/2, p38MAPK) signalling. This effect is attenuated in VSMC from OZR, in which the greater oxidative stress is mediated by NADPH oxidase and mitochondrial complex and by a reduced synthesis/activity of SOD. Impairment of the NO/cGMP/PKG signalling is reproduced in VSMC from LZR by hydrogen peroxide and reverted in VSMC from OZR by antioxidants.
  
  
  CONCLUSIONS
  In VSMC from an animal model of insulin resistance the NO/cGMP/PKG intracellular signalling is impaired due to an increased oxidative stress.
authors.affiliation: Internal Medicine and Metabolic Disease Unit, San Luigi Gonzaga Faculty of Medicine, Department of Clinical and Biological Sciences of the University of Turin, San Luigi Gonzaga Hospital, I-10043 Orbassano, Turin, Italy.
authors.forename: Gabriella
authors.initials: G
authors.lastname: Doronzo
authors.forename: Michela
authors.initials: M
authors.lastname: Viretto
authors.forename: Isabella
authors.initials: I
authors.lastname: Russo
authors.forename: Luigi
authors.initials: L
authors.lastname: Mattiello
authors.forename: Leonardo
authors.initials: L
authors.lastname: Di Martino
authors.forename: Franco
authors.initials: F
authors.lastname: Cavalot
authors.forename: Giovanni
authors.initials: G
authors.lastname: Anfossi
authors.forename: Mariella
authors.initials: M
authors.lastname: Trovati
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology; D015153:Blotting, Western; D006152:Cyclic GMP / Q000378:metabolism; D017869:Cyclic GMP-Dependent Protein Kinases / Q000378:metabolism; D045222:Electron Transport Chain Complex Proteins / Q000378:metabolism; D004789:Enzyme Activation; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006801:Humans; D007333:Insulin Resistance; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008297:Male; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000201:enzymology*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology*; D019255:NADPH Oxidases / Q000378:metabolism; D009569:Nitric Oxide / Q000378:metabolism*; D020030:Nitric Oxide Donors / Q000494:pharmacology; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000187:drug effects; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism*; D000081082:Phosphoinositide-3 Kinase Inhibitors; D010766:Phosphorylation; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D051381:Rats; D011924:Rats, Zucker; D038762:Ribosomal Protein S6 Kinases, 70-kDa / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D013997:Time Factors; D014969:Xanthine Oxidase / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D000975:Antioxidants; D045222:Electron Transport Chain Complex Proteins; D020030:Nitric Oxide Donors; D016877:Oxidants; D000081082:Phosphoinositide-3 Kinase Inhibitors; D047428:Protein Kinase Inhibitors; D013481:Superoxides; D009569:Nitric Oxide; D013482:Superoxide Dismutase; D014969:Xanthine Oxidase; D019255:NADPH Oxidases; D058539:Phosphatidylinositol 3-Kinase; D051057:Proto-Oncogene Proteins c-akt; D038762:Ribosomal Protein S6 Kinases, 70-kDa; D017869:Cyclic GMP-Dependent Protein Kinases; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D020928:Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D006152:Cyclic GMP