Neutrophil MMP-9 proenzyme, unencumbered by TIMP-1, undergoes efficient activation in vivo and catalytically induces angiogenesis via a basic fibroblast growth factor (FGF-2)/FGFR-2 pathway.
The structural and catalytic requirements for neutrophil MMP-9 proenzyme (proMMP-9) to induce angiogenesis were investigated using a quantitative angiogenesis model based on grafting of collagen onplants onto the chorioallantoic membrane of chick embryos. Both physiological activation of neutrophil proMMP-9 and proteolytic activity of the generated MMP-9 enzyme were critically dependent on the tissue inhibitor of metalloproteinase (TIMP)-free status of the zymogen. The presence of an intact active site and hemopexin domain were required for full angiogenesis-inducing activity of the MMP-9 enzyme. Timed additions of TIMP-1 to the onplants containing TIMP-free neutrophil proMMP-9 indicated that in vivo activation of the zymogen occurred during the first 24 h after grafting. Within the onplant tissue, MMP-9 activation was accompanied by proteolytic modifications of fibrillar collagen and an influx of host proteins, the rate of which depended on the TIMP-free status of the zymogen. By quantifying the levels of host angiogenic factors, we demonstrated that basic fibroblast growth factor (FGF-2) was a major cytokine becoming bioavailable in the onplant tissue undergoing a neutrophil proMMP-9-mediated angiogenic switch. Inhibition of angiogenesis with specific function-blocking antibodies further indicated an involvement of a FGF-2/FGFR-2 pathway in neutrophil proMMP-9-induced angiogenesis. The enhanced angiogenesis catalyzed by neutrophil MMP-9 appears to evoke also a localized, low threshold level vascular endothelial growth factor (VEGF)/VEGFR-2 pathway, likely functioning in the formation and/or stabilization of blood vessels. That neutrophil proMMP-9, unencumbered by TIMP-1, directly mediates FGF-2-dependent angiogenesis was also demonstrated in our quantitative mouse angiogenesis model employing subcutaneous collagen implants, thus implicating the novel TIMP-free MMP-9/FGF-2/FGFR-2 pathway in proMMP-9-induced angiogenesis in a mammalian setting.
Ardi
Veronica C
VC
Department of Cell Biology, The Scripps Research Institute, La Jolla, California 92037, USA.
Van den Steen
Philippe E
PE
Opdenakker
Ghislain
G
Schweighofer
Bernhard
B
Deryugina
Elena I
EI
Quigley
James P
JP
D000818:Animals; D002642:Chick Embryo; D004792:Enzyme Precursors / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D016222:Fibroblast Growth Factor 2 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D020780:Matrix Metalloproteinase 9 / Q000235:genetics / Q000378:metabolism* / Q000494:pharmacology; D051379:Mice; D018919:Neovascularization, Physiologic / Q000187:drug effects / Q000502:physiology*; D009504:Neutrophils / Q000201:enzymology; D017434:Protein Structure, Tertiary / Q000502:physiology; D051497:Receptor, Fibroblast Growth Factor, Type 2 / Q000235:genetics / Q000378:metabolism*; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000235:genetics / Q000378:metabolism*
D004792:Enzyme Precursors; D019715:Tissue Inhibitor of Metalloproteinase-1; D016222:Fibroblast Growth Factor 2; C496353:FGFR2 protein, human; C496354:Fgfr2 protein, mouse; D051497:Receptor, Fibroblast Growth Factor, Type 2; C120487:pro-matrix metalloproteinase 9; D020780:Matrix Metalloproteinase 9