Abstract
The secretion of angiogenic factors by vascular endothelial cells is one of the key mechanisms of angiogenesis. Here we report on the isolation of a new potent angiogenic factor, diuridine tetraphosphate (Up4U) from the secretome of human endothelial cells. The angiogenic effect of the endothelial secretome was partially reduced after incubation with alkaline phosphatase and abolished in the presence of suramin. In one fraction, purified to homogeneity by reversed phase and affinity chromatography, Up4U was identified by MALDI-LIFT-fragment-mass-spectrometry, enzymatic cleavage analysis and retention-time comparison. Beside a strong angiogenic effect on the yolk sac membrane and the developing rat embryo itself, Up4U increased the proliferation rate of endothelial cells and, in the presence of PDGF, of vascular smooth muscle cells. Up4U stimulated the migration rate of endothelial cells via P2Y2-receptors, increased the ability of endothelial cells to form capillary-like tubes and acts as a potent inducer of sprouting angiogenesis originating from gel-embedded EC spheroids. Endothelial cells released Up4U after stimulation with shear stress. Mean total plasma Up4U concentrations of healthy subjects (N=6) were sufficient to induce angiogenic and proliferative effects (1.34 ± 0.26 nmol L(-1)). In conclusion, Up4U is a novel strong human endothelium-derived angiogenic factor.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Adult
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Angiogenesis Inducing Agents / chemistry
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Angiogenesis Inducing Agents / metabolism*
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Angiogenesis Inducing Agents / pharmacology
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Animals
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Cell Movement / drug effects
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Chorioallantoic Membrane / drug effects
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Chorioallantoic Membrane / embryology
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Embryo, Mammalian / metabolism
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Endothelial Cells / cytology
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Endothelial Cells / drug effects
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Endothelial Cells / enzymology
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Endothelium, Vascular / cytology
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Endothelium, Vascular / drug effects
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Endothelium, Vascular / metabolism*
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Humans
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In Vitro Techniques
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Mitogen-Activated Protein Kinases / metabolism
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Molecular Weight
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Muscle, Smooth, Vascular / cytology
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Myocytes, Smooth Muscle / cytology
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Myocytes, Smooth Muscle / drug effects
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Myocytes, Smooth Muscle / metabolism
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Phosphorylation / drug effects
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Platelet-Derived Growth Factor / pharmacology
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Rats
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Rats, Wistar
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Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
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Uracil Nucleotides / chemistry
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Uracil Nucleotides / metabolism
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Uracil Nucleotides / pharmacology
Substances
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Angiogenesis Inducing Agents
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Platelet-Derived Growth Factor
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Uracil Nucleotides
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Mitogen-Activated Protein Kinases
Grants and funding
This study was supported by a grant from the German Research Foundation (DFG, Ja-972/11-1; JJ), the Sonnenfeld Foundation (JJ; VJ) and Federal Ministry of Education and Research (BMBF; 01GR0807 (JJ)). The author (VJ; JJ) was supported by grants FP7-HEALTH24 2009-2.4.5-2 to “SYSKID” (241544) and “HEALTH 2011.2.4.2-2″ to “Mascara” (278249) from the European Union. Eutox, FIS PS09/00447, ISCIII-RETIC REDinREN/RD12/0021/0001 and 0004. Comunidad de Madrid/S2010/BMD-2378(NMDS; AAO); FIS-Sara Borrell to MDSN, Programa Intensificación Actividad Investigadora (ISCIII/Agencia Laín-Entralgo/CM) (AAO). VJ received a grant from the Else-Kroener-foundation. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.