Abstract
Herein, we report that VEGF-functionalized dextran (dexOx-VEGF) is comparatively superior to free VEGF in prolonging the phosphorylation of VEGF receptor 2 (VEGFR-2). Both dexOx-VEGF and free VEGF activate VEGFR-2, and the complexes are internalized into early endosomes (EEA1(+) vesicles) and then transported to lysosomes (Rab7(+) vesicles). However, after cell activation, dexOx-VEGF is preferentially colocalized in early endosomes where VEGF signaling is still active while free VEGF is preferentially transported to late endosomes or lysosomes. We further show that dexOx-VEGF after phosphorylation of VEGF receptor 2 induces an increase of intracellular Ca(2+) and activates VEGF downstream effectors such as Akt and extracellular signal-regulated kinase (ERK1/2) proteins. Under specific conditions, the activation level is different from the one observed for free VEGF, thus suggesting mechanistic differences, which is illustrated by cell migration and cord-like formation studies. DexOx-VEGF can be cross-linked with adipic acid dihydrazide to form a degradable gel, which in turn can be incorporated in a fibrin gel containing endothelial cells (ECs) to modulate their activity. We envision that these constructs might be beneficial to extend the pro-angiogenic activity of VEGF in ischemic tissues and to modulate the biological activity of vascular cells.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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Calcium / metabolism
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Cell Movement / drug effects
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Cell Survival / drug effects
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Cytoplasm / drug effects
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Cytoplasm / metabolism
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Dextrans / chemistry*
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Endocytosis / drug effects
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Endosomes / drug effects*
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Endosomes / metabolism
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Gene Expression / drug effects
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Human Umbilical Vein Endothelial Cells / cytology
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Human Umbilical Vein Endothelial Cells / drug effects*
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Human Umbilical Vein Endothelial Cells / metabolism
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Humans
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Hydrogels
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Lysosomes / drug effects*
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Lysosomes / metabolism
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Mitogen-Activated Protein Kinase 1 / genetics
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Mitogen-Activated Protein Kinase 1 / metabolism
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Mitogen-Activated Protein Kinase 3 / genetics
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Mitogen-Activated Protein Kinase 3 / metabolism
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Phosphorylation / drug effects
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Protein Isoforms / chemistry
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Protein Isoforms / metabolism
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Protein Isoforms / pharmacology
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Proto-Oncogene Proteins c-akt / genetics
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Proto-Oncogene Proteins c-akt / metabolism
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Signal Transduction / drug effects
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Vascular Endothelial Growth Factor A / chemistry*
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Vascular Endothelial Growth Factor A / metabolism
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Vascular Endothelial Growth Factor A / pharmacology
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Vascular Endothelial Growth Factor Receptor-2 / chemistry
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Vascular Endothelial Growth Factor Receptor-2 / metabolism
Substances
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Dextrans
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Hydrogels
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Protein Isoforms
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VEGFA protein, human
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Vascular Endothelial Growth Factor A
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Vascular Endothelial Growth Factor Receptor-2
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Proto-Oncogene Proteins c-akt
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MAPK1 protein, human
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Mitogen-Activated Protein Kinase 1
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Mitogen-Activated Protein Kinase 3
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Calcium