title: Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) Promotes Angiogenesis and Ischemia-Induced Neovascularization Via NADPH Oxidase 4 (NOX4) and Nitric Oxide-Dependent Mechanisms.
abstract: BACKGROUND
  Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) has the ability to inhibit angiogenesis by inducing endothelial cell death, as well as being able to promote pro-angiogenic activity in vitro. These seemingly opposite effects make its role in ischemic disease unclear. Using Trail(-/-) and wildtype mice, we sought to determine the role of TRAIL in angiogenesis and neovascularization following hindlimb ischemia.
  
  
  RESULTS
  Reduced vascularization assessed by real-time 3-dimensional Vevo ultrasound imaging and CD31 staining was evident in Trail(-/-) mice after ischemia, and associated with reduced capillary formation and increased apoptosis. Notably, adenoviral TRAIL administration significantly improved limb perfusion, capillary density, and vascular smooth-muscle cell content in both Trail(-/-) and wildtype mice. Fibroblast growth factor-2, a potent angiogenic factor, increased TRAIL expression in human microvascular endothelial cell-1, with fibroblast growth factor-2-mediated proliferation, migration, and tubule formation inhibited with TRAIL siRNA. Both fibroblast growth factor-2 and TRAIL significantly increased NADPH oxidase 4 (NOX4) expression. TRAIL-inducible angiogenic activity in vitro was inhibited with siRNAs targeting NOX4, and consistent with this, NOX4 mRNA was reduced in 3-day ischemic hindlimbs of Trail(-/-) mice. Furthermore, TRAIL-induced proliferation, migration, and tubule formation was blocked by scavenging H2O2, or by inhibiting nitric oxide synthase activity. Importantly, TRAIL-inducible endothelial nitric oxide synthase phosphorylation at Ser-1177 and intracellular human microvascular endothelial cell-1 cell nitric oxide levels were NOX4 dependent.
  
  
  CONCLUSIONS
  This is the first report demonstrating that TRAIL can promote angiogenesis following hindlimb ischemia in vivo. The angiogenic effect of TRAIL on human microvascular endothelial cell-1 cells is downstream of fibroblast growth factor-2, involving NOX4 and nitric oxide signaling. These data have significant therapeutic implications, such that TRAIL may improve the angiogenic response to ischemia and increase perfusion recovery in patients with cardiovascular disease and diabetes.
authors.affiliation: The Heart Research Institute, Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.) The University of Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.).
authors.forename: Belinda Ann
authors.initials: BA
authors.lastname: Di Bartolo
authors.affiliation: The Heart Research Institute, Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.) Centre for Vascular Research, University of New South Wales, Sydney, Australia (S.P.C., L.P.L., J.R., N.S.M.A., T.T., A.W.S.Y., S.R.T.) The University of Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.).
authors.forename: Siân Peta
authors.initials: SP
authors.lastname: Cartland
authors.affiliation: Centre for Vascular Research, University of New South Wales, Sydney, Australia (S.P.C., L.P.L., J.R., N.S.M.A., T.T., A.W.S.Y., S.R.T.).
authors.forename: Leonel
authors.initials: L
authors.lastname: Prado-Lourenco
authors.affiliation: Department of Urology, The University of Minnesota, Minneapolis, MN (T.S.G.).
authors.forename: Thomas Scott
authors.initials: TS
authors.lastname: Griffith
authors.affiliation: The Heart Research Institute, Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.) The University of Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.).
authors.forename: Carmine
authors.initials: C
authors.lastname: Gentile
authors.affiliation: Centre for Vascular Research, University of New South Wales, Sydney, Australia (S.P.C., L.P.L., J.R., N.S.M.A., T.T., A.W.S.Y., S.R.T.) School of Medical Sciences, University of New South Wales, Sydney, Australia (J.R., T.T., S.R.T.).
authors.forename: Jayant
authors.initials: J
authors.lastname: Ravindran
authors.affiliation: Centre for Vascular Research, University of New South Wales, Sydney, Australia (S.P.C., L.P.L., J.R., N.S.M.A., T.T., A.W.S.Y., S.R.T.).
authors.forename: Nor Saadah Muhammad
authors.initials: NS
authors.lastname: Azahri
authors.affiliation: Centre for Vascular Research, University of New South Wales, Sydney, Australia (S.P.C., L.P.L., J.R., N.S.M.A., T.T., A.W.S.Y., S.R.T.) School of Medical Sciences, University of New South Wales, Sydney, Australia (J.R., T.T., S.R.T.).
authors.forename: Thuan
authors.initials: T
authors.lastname: Thai
authors.affiliation: Centre for Vascular Research, University of New South Wales, Sydney, Australia (S.P.C., L.P.L., J.R., N.S.M.A., T.T., A.W.S.Y., S.R.T.).
authors.forename: Amanda Wing Shee
authors.initials: AW
authors.lastname: Yeung
authors.affiliation: Centre for Vascular Research, University of New South Wales, Sydney, Australia (S.P.C., L.P.L., J.R., N.S.M.A., T.T., A.W.S.Y., S.R.T.) School of Medical Sciences, University of New South Wales, Sydney, Australia (J.R., T.T., S.R.T.).
authors.forename: Shane Ross
authors.initials: SR
authors.lastname: Thomas
authors.affiliation: The Heart Research Institute, Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.) The University of Sydney, NSW, Australia (B.A.D.B., S.P.C., C.G., M.M.K.).
authors.forename: Mary Meltem
authors.initials: MM
authors.lastname: Kavurma
mesh_terms: D000818:Animals; D017209:Apoptosis; D015415:Biomarkers / Q000378:metabolism; D002196:Capillaries / Q000201:enzymology* / Q000473:pathology / Q000503:physiopathology; D002465:Cell Movement; D049109:Cell Proliferation; D002478:Cells, Cultured; D004195:Disease Models, Animal; D005260:Female; D005838:Genotype; D006614:Hindlimb; D006801:Humans; D007511:Ischemia / Q000000981:diagnostic imaging / Q000201:enzymology* / Q000235:genetics / Q000503:physiopathology; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D018482:Muscle, Skeletal / Q000098:blood supply* / Q000201:enzymology*; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases / Q000235:genetics / Q000378:metabolism*; D018919:Neovascularization, Physiologic; D009569:Nitric Oxide / Q000378:metabolism*; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D010641:Phenotype; D010766:Phosphorylation; D019408:Platelet Endothelial Cell Adhesion Molecule-1 / Q000378:metabolism; D034622:RNA Interference; D015398:Signal Transduction; D053221:TNF-Related Apoptosis-Inducing Ligand / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D018084:Ultrasonography, Interventional
chemical_list: D015415:Biomarkers; D019408:Platelet Endothelial Cell Adhesion Molecule-1; D053221:TNF-Related Apoptosis-Inducing Ligand; C505653:TNFSF10 protein, human; C505654:Tnfsf10 protein, mouse; D009569:Nitric Oxide; C496314:NOS3 protein, human; D052250:Nitric Oxide Synthase Type III; D000074663:NADPH Oxidase 4; D019255:NADPH Oxidases; C498910:NOX4 protein, human; C422943:Nox4 protein, mouse
keywords: angiogenesis; endothelial cell; gene expression; ischemia