1. Dissecting the role of matrix metalloproteinases (MMP) and integrin alpha(v)beta3 in angiogenesis in vitro: absence of hemopexin C domain bioactivity, but membrane-Type 1-MMP and alpha(v)beta3 are critical.
- Author
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Nisato RE, Hosseini G, Sirrenberg C, Butler GS, Crabbe T, Docherty AJ, Wiesner M, Murphy G, Overall CM, Goodman SL, and Pepper MS
- Subjects
- Animals, Cattle, Cytokines pharmacology, Endothelial Cells enzymology, Endothelial Cells metabolism, Enzyme Activation, Fibroblast Growth Factor 2 pharmacology, Hemopexin pharmacology, Humans, Matrix Metalloproteinase 2 biosynthesis, Matrix Metalloproteinase 2 genetics, Matrix Metalloproteinase 2 pharmacology, Matrix Metalloproteinases, Membrane-Associated, Metalloendopeptidases antagonists & inhibitors, Metalloendopeptidases biosynthesis, Oligopeptides pharmacology, Protein Structure, Tertiary, RNA, Messenger biosynthesis, RNA, Messenger genetics, Vascular Endothelial Growth Factor A pharmacology, Hemopexin metabolism, Integrin alphaVbeta3 metabolism, Matrix Metalloproteinase 2 metabolism, Metalloendopeptidases metabolism, Neovascularization, Physiologic physiology
- Abstract
Matrix metalloproteinase (MMP)-2 and its hemopexin C domain autolytic fragment (also called PEX) have been proposed to be crucial for angiogenesis. Here, we have investigated the dependency of in vitro angiogenesis on MMP-mediated extracellular proteolysis and integrin alpha(v)beta3-mediated cell adhesion in a three-dimensional collagen I model. The hydroxamate-based synthetic inhibitors BB94, CT1399, and CT1847 inhibited endothelial cell invasion, as did neutralizing anti-membrane-type 1-MMP (MT1-MMP) antibodies and tissue inhibitor of MMP (TIMP)-2 and TIMP-3 but not TIMP-1. This confirmed the pivotal importance of MT1-MMP over other MMPs in this model. Invasion was also inhibited by a nonpeptidic antagonist of integrin alpha(v)beta3, EMD 361276. Although PEX strongly inhibited pro-MMP-2 activation, when contaminating lipopolysaccharide was neutralized, PEX neither affected angiogenesis nor bound integrin alpha(v)beta(3). Moreover, no specific binding of pro-MMP-2 to integrin alpha(v)beta3 was found, whereas only one out of four independently prepared enzymatically active MMP-2 preparations could bind integrin alpha(v)beta3 , and this in a PEX-independent manner. Likewise, integrin alpha(v)beta3 -expressing cells did not bind MMP-2-coated surfaces. Hence, these findings show that endothelial cell invasion of collagen I gels is MT1-MMP and alpha(v)beta3 - dependent but MMP-2 independent and does not support a role for PEX in alpha(v)beta3 integrin binding or in modulating angiogenesis in this system.
- Published
- 2005
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