Tissue Inhibitor of Metalloproteinase-3 (TIMP-3) Suppresses VEGF Receptor-2 Mediated Signaling Independent of Metalloproteinase Inhibition

Abstract

Abstract: : Purpose: Tissue Inhibitor of Metalloproteinase-3 (TIMP-3) is a potent anti-angiogenic factor that is produced by the retinal pigment epithelium (RPE). In the retina, vascular endothelial growth factor (VEGF), an angiogenic factor secreted by the RPE induces endothelial cell migration, proliferation and vascular leakage. This study investigated the mechanism by which TIMP-3 inhibits VEGF mediated signaling via VEGF receptor-2. Methods: We have introduced TIMP-3 into porcine aortic endothelial cells expressing VEGFR-2. In vitro, VEGF induced proliferation and migration of endothelial cells were analyzed by Coulter particle counting and modified Boyden chamber assays respectively. VEGF-induced autophosphorylation of VEGFR-2 and activation of p44/p42 MAP kinase was evaluated by western blot analysis. Results: TIMP-3 overexpression in endothelial cells resulted in an inhibition of the proliferative and migratory response of these cells to VEGF. VEGF-induced autophosphorylation of VEGFR-2 and p44/p42 MAP kinase activation were also suppressed. These effects are specific for TIMP-3 as TIMP-1, TIMP-2 and synthetic matrix metalloproteinase (MMP) inhibitors demonstrated no effect. Conclusion:Our data indicate that TIMP-3 inhibits VEGF-mediated angiogenesis by interfering with the VEGF receptor-2 signaling pathway and that this effect is independent of its metalloproteinase inhibitory activity.

Keywords: 483 neovascularization • 580 signal transduction