title: Thrombospondin-1 regulates blood flow via CD47 receptor-mediated activation of NADPH oxidase 1.
abstract: OBJECTIVE
  Although the matricellular protein thrombospondin-1 (TSP1) is highly expressed in the vessel wall in response to injury, its pathophysiological role in the development of vascular disease is poorly understood. This study was designed to test the hypothesis that TSP1 stimulates reactive oxygen species production in vascular smooth muscle cells and induces vascular dysfunction by promoting oxidative stress.
  
  
  RESULTS
  Nanomolar concentrations of TSP1 found in human vascular disease robustly stimulated superoxide (O(2)(•-)) levels in vascular smooth muscle cells at both cellular and tissue level as measured by cytochrome c and electron paramagnetic resonance. A peptide mimicking the C terminus of TSP1 known to specifically bind CD47 recapitulated this response. Transcriptional knockdown of CD47 and a monoclonal inhibitory CD47 antibody abrogated TSP1-triggered O(2)(•-) in vitro and ex vivo. TSP1 treatment of vascular smooth muscle cells activated phospholipase C and protein kinase C, resulting in phosphorylation of the NADPH oxidase organizer subunit p47(phox) and subsequent Nox1 activation, leading to impairment of arterial vasodilatation ex vivo. Further, we observed that blockade of CD47 and NADPH oxidase 1 gene silencing in vivo in rats improves TSP1-induced impairment of tissue blood flow after ischemia reperfusion.
  
  
  CONCLUSIONS
  Our data suggest a highly regulated process of reactive oxygen species stimulation and blood flow regulation promoted through a direct TSP1/CD47-mediated activation of Nox1. This is the first report, to our knowledge, of a matricellular protein acting as a ligand for NADPH oxidase activation and through specific engagement of integrin-associated protein CD47.
authors.affiliation: Vascular Medicine Institute, Department of Pharmacology and Chemical Biology, University of Pittsburgh, PA 15261, USA.
authors.forename: Gábor
authors.initials: G
authors.lastname: Csányi
authors.forename: Mingyi
authors.initials: M
authors.lastname: Yao
authors.forename: Andrés I
authors.initials: AI
authors.lastname: Rodríguez
authors.forename: Imad
authors.initials: I
authors.lastname: Al Ghouleh
authors.forename: Maryam
authors.initials: M
authors.lastname: Sharifi-Sanjani
authors.forename: Giovanna
authors.initials: G
authors.lastname: Frazziano
authors.forename: Xiaojun
authors.initials: X
authors.lastname: Huang
authors.forename: Eric E
authors.initials: EE
authors.lastname: Kelley
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Isenberg
authors.forename: Patrick J
authors.initials: PJ
authors.lastname: Pagano
mesh_terms: D000818:Animals; D051928:CD47 Antigen / Q000235:genetics / Q000502:physiology*; D020868:Gene Silencing; D008297:Male; D051379:Mice; D009131:Muscle, Smooth, Vascular / Q000187:drug effects / Q000502:physiology*; D009247:NADH, NADPH Oxidoreductases / Q000235:genetics / Q000502:physiology*; D000074624:NADPH Oxidase 1; D018384:Oxidative Stress / Q000187:drug effects / Q000502:physiology; D011493:Protein Kinase C / Q000502:physiology; D051381:Rats; D012039:Regional Blood Flow / Q000187:drug effects / Q000502:physiology*; D015398:Signal Transduction / Q000502:physiology; D013482:Superoxide Dismutase / Q000502:physiology; D000072105:Superoxide Dismutase-1; D019700:Thrombospondin 1 / Q000494:pharmacology / Q000502:physiology*; D010738:Type C Phospholipases / Q000502:physiology
chemical_list: D051928:CD47 Antigen; C000606290:SOD1 protein, human; D019700:Thrombospondin 1; C000606291:Sod1 protein, mouse; C000606292:Sod1 protein, rat; D013482:Superoxide Dismutase; D000072105:Superoxide Dismutase-1; D009247:NADH, NADPH Oxidoreductases; D000074624:NADPH Oxidase 1; C000614711:NOX1 protein, mouse; C000614712:NOX1 protein, rat; D011493:Protein Kinase C; D010738:Type C Phospholipases