title: Nitric oxide regulates angiogenesis through a functional switch involving thrombospondin-1.
abstract: Nitric oxide (NO) donors have been shown to stimulate and inhibit the proliferation, migration, and differentiation of endothelial cells in vitro and angiogenesis in vivo. Recently, we have shown distinct thresholds for NO to regulate p53-Ser-15P, phosphorylated extracellular signal-regulated kinase (pERK), and hypoxia inducible factor 1alpha in tumor cells. Because these signaling pathways also promote the growth and survival of endothelial cells, we examined their roles in angiogenic responses of venous endothelial cells and vascular outgrowth of muscle explants elicited by NO. An additional protein involved in the regulation of angiogenesis is thrombospondin-1 (TSP1), a matricellular glycoprotein known to influence adhesion, migration, and proliferation of endothelial cells. Here we demonstrate a triphasic regulation of TSP1 mediated by a slow and prolonged release of NO that depends on ERK phosphorylation. Under conditions of 5% serum, a 24-h exposure of NO donor (0.1-1,000 microM) mediated a triphasic response in the expression of TSP1 protein: decreasing at 0.1 microM, rebounding at 100 microM, and decreasing again at 1,000 microM. Under the same conditions, we observed a dose-dependent increase in P53 phosphorylation and inverse biphasic responses of pERK and mitogen-activated protein kinase phosphatase-1. Both the growth-stimulating activity of low-dose NO for endothelial cells and suppression of TSP1 expression were ERK-dependent. Conversely, exogenous TSP1 suppressed NO-mediated pERK. These results suggest that dose-dependent positive- and negative-feedback loops exist between NO and TSP1. Limiting TSP1 expression by positive feedback through the ERK mitogen-activated protein kinase pathway may facilitate switching to a proangiogenic state at low doses of NO.
authors.affiliation: Radiation Biology Branch and Laboratory of Pathology, National Cancer Institute, National Institutes of Health, Bethesda, MD 20892, USA. ridnourl@mail.nih.gov
authors.forename: Lisa A
authors.initials: LA
authors.lastname: Ridnour
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Isenberg
authors.forename: Michael G
authors.initials: MG
authors.lastname: Espey
authors.forename: Douglas D
authors.initials: DD
authors.lastname: Thomas
authors.forename: David D
authors.initials: DD
authors.lastname: Roberts
authors.forename: David A
authors.initials: DA
authors.lastname: Wink
mesh_terms: D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D054638:Dual Specificity Phosphatase 1; D004730:Endothelium, Vascular / Q000166:cytology / Q000502:physiology*; D025461:Feedback, Physiological; D005786:Gene Expression Regulation / Q000187:drug effects*; D006801:Humans; D017874:Immediate-Early Proteins / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D018919:Neovascularization, Physiologic; D009569:Nitric Oxide / Q000494:pharmacology*; D010369:Pectoralis Muscles / Q000166:cytology; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010766:Phosphorylation; D054645:Protein Phosphatase 1; D017027:Protein Tyrosine Phosphatases / Q000378:metabolism; D019700:Thrombospondin 1 / Q000235:genetics* / Q000502:physiology; D016159:Tumor Suppressor Protein p53 / Q000378:metabolism; D014471:Umbilical Veins / Q000166:cytology
chemical_list: D018797:Cell Cycle Proteins; D017874:Immediate-Early Proteins; D019700:Thrombospondin 1; D016159:Tumor Suppressor Protein p53; D009569:Nitric Oxide; D048052:Mitogen-Activated Protein Kinase 3; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1; C517019:DUSP1 protein, human; D054638:Dual Specificity Phosphatase 1; C517020:Dusp1 protein, mouse; D017027:Protein Tyrosine Phosphatases