title: 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.
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.
authors.affiliation: Department of Morphology, University Medical Center, Geneva, Switzerland.
authors.forename: Riccardo E
authors.initials: RE
authors.lastname: Nisato
authors.forename: Ghamartaj
authors.initials: G
authors.lastname: Hosseini
authors.forename: Christian
authors.initials: C
authors.lastname: Sirrenberg
authors.forename: Georgina S
authors.initials: GS
authors.lastname: Butler
authors.forename: Thomas
authors.initials: T
authors.lastname: Crabbe
authors.forename: Andrew J P
authors.initials: AJ
authors.lastname: Docherty
authors.forename: Matthias
authors.initials: M
authors.lastname: Wiesner
authors.forename: Gillian
authors.initials: G
authors.lastname: Murphy
authors.forename: Christopher M
authors.initials: CM
authors.lastname: Overall
authors.forename: Simon L
authors.initials: SL
authors.lastname: Goodman
authors.forename: Michael S
authors.initials: MS
authors.lastname: Pepper
mesh_terms: D000818:Animals; D002417:Cattle; D016207:Cytokines / Q000494:pharmacology; D042783:Endothelial Cells / Q000201:enzymology / Q000378:metabolism; D004789:Enzyme Activation; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D006466:Hemopexin / Q000378:metabolism* / Q000494:pharmacology; D006801:Humans; D039302:Integrin alphaVbeta3 / Q000378:metabolism*; D020778:Matrix Metalloproteinase 2 / Q000096:biosynthesis / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000096:biosynthesis / Q000378:metabolism*; D018919:Neovascularization, Physiologic / Q000502:physiology*; D009842:Oligopeptides / Q000494:pharmacology; D017434:Protein Structure, Tertiary; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D042461:Vascular Endothelial Growth Factor A / Q000494:pharmacology
chemical_list: D016207:Cytokines; D039302:Integrin alphaVbeta3; D009842:Oligopeptides; D012333:RNA, Messenger; D042461:Vascular Endothelial Growth Factor A; D016222:Fibroblast Growth Factor 2; C047981:arginyl-glycyl-aspartic acid; D006466:Hemopexin; D053504:Matrix Metalloproteinases, Membrane-Associated; D008666:Metalloendopeptidases; D020778:Matrix Metalloproteinase 2