Nerve growth factor induces endothelial cell invasion and cord formation by promoting matrix metalloproteinase-2 expression through the phosphatidylinositol 3-kinase/Akt signaling pathway and AP-2 transcription factor

J Biol Chem. 2007 Oct 19;282(42):30485-96. doi: 10.1074/jbc.M701081200. Epub 2007 Jul 31.

Abstract

Nerve growth factor (NGF) is a well characterized neurotrophic agonist in the nervous system that triggers angiogenesis. In this study, we investigated the signaling mechanisms involved in NGF-induced angiogenesis. NGF stimulated endothelial cell invasion and cord formation on Matrigel in vitro but had marginal effect on proliferation and migration of these cells. NGF stimulated matrix metalloproteinase (MMP)-2 mRNA expression and protein secretion in human umbilical vein endothelial cells. Using synthetic and endogenous inhibitors of MMP-2 and MMP-2 small interfering RNA suppressed NGF-induced invasion and cord formation. We demonstrated that NGF-induced MMP-2 secretion, invasion, and cord formation are regulated via activation of the NGF receptor, TrkA, phosphatidylinositol 3-kinase (PI3K), and Akt using various pharmacological inhibitors. Specifically, NGF enhanced TrkA phosphorylation, PI3K activity, and Akt phosphorylation. Introduction of NGF-neutralizing antibodies, dominant-negative Akt, or wild-type PTEN effectively inhibited NGF-induced MMP-2 secretion and cord formation. Deletion and site-directed mutagenesis analysis of the MMP-2 promoter demonstrated that the AP-2-binding site is critical for NGF-induced MMP-2 promoter activity. NGF increased the DNA binding activity of AP-2, which was suppressed by inhibitors of TrkA and PI3K. Furthermore, transfection of AP-2 small interfering RNA effectively blocked NGF-induced MMP-2 secretion and cord formation. Finally, NGF promoted neovessel formation in Matrigel plugs in vivo, which was significantly inhibited by K252a and LY294002, but it failed to promote angiogenesis using MMP-2 knock-out mice. Our data collectively suggest that NGF stimulates endothelial cell invasion and cord formation by augmenting MMP-2 via the PI3K/Akt signaling pathway and AP-2 transcription factor, which may be responsible for triggering angiogenesis.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Movement / drug effects*
  • Cell Movement / physiology
  • Cells, Cultured
  • Chromones / pharmacology
  • Endothelial Cells / enzymology*
  • Enzyme Activation / drug effects
  • Enzyme Activation / physiology
  • Enzyme Inhibitors / pharmacology
  • Gene Deletion
  • Humans
  • Matrix Metalloproteinase 2 / genetics
  • Matrix Metalloproteinase 2 / metabolism*
  • Matrix Metalloproteinase Inhibitors
  • Mice
  • Mice, Knockout
  • Morpholines / pharmacology
  • Mutagenesis, Site-Directed
  • Neovascularization, Physiologic / drug effects
  • Neovascularization, Physiologic / physiology
  • Nerve Growth Factor / pharmacology*
  • Nitriles / pharmacology
  • Phosphatidylinositol 3-Kinases / metabolism*
  • Phosphoinositide-3 Kinase Inhibitors
  • Protein Binding / drug effects
  • Protein Binding / physiology
  • Proto-Oncogene Proteins c-akt / antagonists & inhibitors
  • Proto-Oncogene Proteins c-akt / metabolism*
  • Pyrrolidines / pharmacology
  • RNA, Small Interfering / pharmacology
  • Receptor, trkA / antagonists & inhibitors
  • Receptor, trkA / metabolism
  • Signal Transduction / drug effects
  • Signal Transduction / physiology
  • Transcription Factor AP-2 / metabolism*
  • Umbilical Veins / cytology
  • Umbilical Veins / enzymology*

Substances

  • (S)-1-(4-methyl-1-(2-pyrimidinyl)-4-piperidylamino)acetyl-2-pyrrolidinecarbonitrile
  • Chromones
  • Enzyme Inhibitors
  • Matrix Metalloproteinase Inhibitors
  • Morpholines
  • Nitriles
  • Phosphoinositide-3 Kinase Inhibitors
  • Pyrrolidines
  • RNA, Small Interfering
  • Transcription Factor AP-2
  • 2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one
  • Nerve Growth Factor
  • Receptor, trkA
  • Proto-Oncogene Proteins c-akt
  • MMP2 protein, human
  • Matrix Metalloproteinase 2
  • Mmp2 protein, mouse