Abstract
KDR/FIk-1 tyrosine kinase, one of the two VEGF receptors induces mitogenesis and differentiation of vascular endothelial cells. We have previously reported that a major target molecule of KDR/Flk-1 kinase is PLC-gamma, and that VEGF induces activation of MAP kinase, mainly mediated by protein kinase C (PKC) in the NIH3T3 cells overexpressing KDR/FIk-1 (Takahashi and Shibuya, 1997). However, the signal transduction initiated from VEGF in endothelial cells remains to be elucidated. In primary sinusoidal endothelial cells which showed strictly VEGF-dependent growth, we found that VEGF stimulated the activation of Raf-1-MEK-MAP kinase cascade. To our surprise, an important regulator, Ras was not efficiently activated to a significant level in response to VEGF. Consistent with this, dominant-negative Ras did not block the VEGF-induced phosphorylation of MAP kinase. On the other hand, PKC-specific inhibitors severely reduced VEGF-dependent phosphorylation of MEK, activation of MAP kinase and subsequent DNA synthesis. A potent PI3 kinase inhibitor, Wortmannin, could not inhibit either of them. These results suggest that in primary endothelial cells, VEGF-induced activation of Raf-MEK-MAP kinase and DNA synthesis are mainly mediated by PKC-dependent pathway, much more than by Ras-dependent or PI3 kinase-dependent pathway.
Publication types
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Research Support, Non-U.S. Gov't
MeSH terms
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3T3 Cells / drug effects
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Adenoviridae / genetics
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Adenoviridae / physiology
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Androstadienes / pharmacology
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Animals
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Calcium-Calmodulin-Dependent Protein Kinases / physiology*
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Carbazoles / pharmacology
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Chromones / pharmacology
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DNA Replication / drug effects*
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Endothelial Growth Factors / pharmacology*
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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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Enzyme Activation
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Enzyme Inhibitors / pharmacology
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Flavonoids / pharmacology
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Genes, ras
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Genetic Vectors / genetics
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Genetic Vectors / physiology
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Indoles / pharmacology
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Lymphokines / pharmacology*
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MAP Kinase Kinase Kinase 1*
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Maleimides / pharmacology
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Mice
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Morpholines / pharmacology
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Phosphatidylinositol 3-Kinases / physiology
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Phosphoinositide-3 Kinase Inhibitors
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Protein Isoforms / physiology
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Protein Kinase C / physiology*
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Protein Serine-Threonine Kinases / antagonists & inhibitors
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Protein Serine-Threonine Kinases / physiology*
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Proto-Oncogene Proteins c-raf / physiology*
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Proto-Oncogene Proteins p21(ras) / physiology
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Rats
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Receptor Protein-Tyrosine Kinases / physiology
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Receptors, Growth Factor / physiology
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Receptors, Vascular Endothelial Growth Factor
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Signal Transduction / drug effects*
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Tetradecanoylphorbol Acetate / pharmacology
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Transfection
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Vascular Endothelial Growth Factor A
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Vascular Endothelial Growth Factors
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Wortmannin
Substances
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Androstadienes
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Carbazoles
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Chromones
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Endothelial Growth Factors
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Enzyme Inhibitors
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Flavonoids
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Indoles
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Lymphokines
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Maleimides
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Morpholines
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Phosphoinositide-3 Kinase Inhibitors
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Protein Isoforms
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Receptors, Growth Factor
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Vascular Endothelial Growth Factor A
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Vascular Endothelial Growth Factors
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Go 6976
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2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
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Receptor Protein-Tyrosine Kinases
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Receptors, Vascular Endothelial Growth Factor
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Protein Serine-Threonine Kinases
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Proto-Oncogene Proteins c-raf
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Protein Kinase C
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Calcium-Calmodulin-Dependent Protein Kinases
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MAP Kinase Kinase Kinase 1
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Map3k1 protein, mouse
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Proto-Oncogene Proteins p21(ras)
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bisindolylmaleimide I
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Tetradecanoylphorbol Acetate
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2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one
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Wortmannin