title: Thrombospondin-1-CD47 blockade and exogenous nitrite enhance ischemic tissue survival, blood flow and angiogenesis via coupled NO-cGMP pathway activation.
abstract: Tissue ischemia and ischemia-reperfusion (I/R) remain sources of cell and tissue death. Inability to restore blood flow and limit reperfusion injury represents a challenge in surgical tissue repair and transplantation. Nitric oxide (NO) is a central regulator of blood flow, reperfusion signaling and angiogenesis. De novo NO synthesis requires oxygen and is limited in ischemic vascular territories. Nitrite (NO(2-)) has been discovered to convert to NO via heme-based reduction during hypoxia, providing a NO synthase independent and oxygen-independent NO source. Furthermore, blockade of the matrix protein thrombospondin-1 (TSP1) or its receptor CD47 has been shown to promote downstream NO signaling via soluble guanylate cyclase (sGC) and cGMP-dependant kinase. We hypothesized that nitrite would provide an ischemic NO source that could be potentiated by TSP1-CD47 blockade enhancing ischemic tissue survival, blood flow and angiogenesis. Both low dose nitrite and direct blockade of TSP1-CD47 interaction using antibodies or gene silencing increased acute blood flow and late tissue survival in ischemic full thickness flaps. Nitrite and TSP1 blockade both enhanced in vitro and in vivo angiogenic responses. The nitrite effect could be abolished by inhibition of sGC and cGMP signaling. Potential therapeutic synergy was tested in a more severe ischemic flap model. We found that combined therapy with nitrite and TSP1-CD47 blockade enhanced flap perfusion, survival and angiogenesis to a greater extent than either agent alone, providing approximately 100% flap survival. These data provide a new therapeutic paradigm for hypoxic NO signaling through enhanced cGMP mediated by TSP1-CD47 blockade and nitrite delivery.
authors.affiliation: Vascular Medicine Institute of the University of Pittsburgh, Division of Pulmonary, Allergy and Critical Care, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261, USA. jsi5@pitt.edu
authors.forename: Jeff S
authors.initials: JS
authors.lastname: Isenberg
authors.forename: Sruti
authors.initials: S
authors.lastname: Shiva
authors.forename: Mark
authors.initials: M
authors.lastname: Gladwin
mesh_terms: D000704:Analysis of Variance; D000818:Animals; D001415:Back / Q000098:blood supply / Q000473:pathology; D051928:CD47 Antigen / Q000378:metabolism*; D006152:Cyclic GMP / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D020868:Gene Silencing; D007511:Ischemia / Q000188:drug therapy* / Q000378:metabolism* / Q000473:pathology; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D018919:Neovascularization, Physiologic / Q000187:drug effects; D009569:Nitric Oxide / Q000378:metabolism*; D009573:Nitrites / Q000494:pharmacology*; D010069:Oxadiazoles / Q000494:pharmacology; D011810:Quinoxalines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction; D013524:Surgical Flaps / Q000098:blood supply / Q000473:pathology; D019700:Thrombospondin 1 / Q000378:metabolism*; D014968:Xanthine Dehydrogenase / Q000378:metabolism
chemical_list: C095284:1H-(1,2,4)oxadiazolo(4,3-a)quinoxalin-1-one; D051928:CD47 Antigen; D004791:Enzyme Inhibitors; D009573:Nitrites; D010069:Oxadiazoles; D011810:Quinoxalines; D019700:Thrombospondin 1; D009569:Nitric Oxide; D014968:Xanthine Dehydrogenase; D006152:Cyclic GMP