Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) Promotes Angiogenesis and Ischemia-Induced Neovascularization Via NADPH Oxidase 4 (NOX4) and Nitric Oxide-Dependent Mechanisms

Belinda Ann Di Bartolo; Siân Peta Cartland; Leonel Prado-Lourenco; Thomas Scott Griffith; Carmine Gentile; Jayant Ravindran; Nor Saadah Muhammad Azahri; Thuan Thai; Amanda Wing Shee Yeung; Shane Ross Thomas; Mary Meltem Kavurma

doi:10.1161/JAHA.115.002527

Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand (TRAIL) Promotes Angiogenesis and Ischemia-Induced Neovascularization Via NADPH Oxidase 4 (NOX4) and Nitric Oxide-Dependent Mechanisms

J Am Heart Assoc. 2015 Nov 16;4(11):e002527. doi: 10.1161/JAHA.115.002527.

Affiliations

¹ 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.).
² 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.).
³ 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.).
⁴ Department of Urology, The University of Minnesota, Minneapolis, MN (T.S.G.).
⁵ 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.).

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.

Methods and 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.

Keywords: angiogenesis; endothelial cell; gene expression; ischemia.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Animals
Apoptosis
Biomarkers / metabolism
Capillaries / enzymology*
Capillaries / pathology
Capillaries / physiopathology
Cell Movement
Cell Proliferation
Cells, Cultured
Disease Models, Animal
Female
Genotype
Hindlimb
Humans
Ischemia / diagnostic imaging
Ischemia / enzymology*
Ischemia / genetics
Ischemia / physiopathology
Mice, Inbred C57BL
Mice, Knockout
Muscle, Skeletal / blood supply*
Muscle, Skeletal / enzymology*
NADPH Oxidase 4
NADPH Oxidases / genetics
NADPH Oxidases / metabolism*
Neovascularization, Physiologic*
Nitric Oxide / metabolism*
Nitric Oxide Synthase Type III / metabolism
Phenotype
Phosphorylation
Platelet Endothelial Cell Adhesion Molecule-1 / metabolism
RNA Interference
Signal Transduction
TNF-Related Apoptosis-Inducing Ligand / deficiency
TNF-Related Apoptosis-Inducing Ligand / genetics
TNF-Related Apoptosis-Inducing Ligand / metabolism*
Time Factors
Transfection
Ultrasonography, Interventional

Substances

Biomarkers
Platelet Endothelial Cell Adhesion Molecule-1
TNF-Related Apoptosis-Inducing Ligand
TNFSF10 protein, human
Tnfsf10 protein, mouse
Nitric Oxide
NOS3 protein, human
Nitric Oxide Synthase Type III
NADPH Oxidase 4
NADPH Oxidases
NOX4 protein, human
Nox4 protein, mouse