The increasing diversity in both substrates and functions of matrix metalloproteinases (MMPs) makes these enzymes central regulators in the complex tumor ecosystem composed of cancer cells and their microenvironment. In the majority of cancers, membrane-associated and extracellular proteases are mainly produced by host cells including inflammatory cells, endothelial cells, pericytes and fibroblasts. Recent data based on in vitro and in vivo studies have demonstrated the relevance of these enzymes in multiple processes controlling cancer growth, angiogenesis and metastatic dissemination. This review will present the emerging MMP-related features of cancer cells and host cells.

Previous studies have shown that the production of gelatinase (MMP-2 and MMP-9) belonging to the matrix metalloproteinase family is increased in human choroidal neovascularization (CNV) occurring during the exudative most aggressive form of age-related macular degeneration (AMD). To more precisely delineate the respective roles of MMP-2 and MMP-9 in choroidal neo-angiogenesis, we investigated their expression and activities in the course of laser-induced murine choroidal neovascularization,. This model was applied to single (MMP-9 KO, MMP-2 KO) or double (MMP-2,9 KO) -deficient mice and to their corresponding wildtype (WT) controls. PRINCIPAL FINDINGS MMP-2 and MMP-9 are produced in lesions associated with choroidal neovascularizationGelatin zymography analysis of ocular posterior segments demonstrated that both MMP-2 and MMP-9 were increasingly produced during the early stages of CNV formation, with the appearance of active forms of MMP-2. In situ zymography revealed a predominant gelatinase activity in the CNV area. Different regulation of MMP-2 and MMP-9 expressionRT-PCR evaluation showed that MMP-9 expression was up-regulated during early phases of CNV formation whereas MMP-2 was constitutively expressed without any transcriptional modulation. MT1-MMP mRNA was concomitantly up-regulated, suggesting that the presence of active MMP2 forms was due to the expression and activity of its activator. Severe inhibition of choroidal neovascularization in MMP-2,9 double-deficient miceFluorescein angiography performed before death (Fig. 1F) showed a significant reduction (PϽ0.001) in the number of leaking spots (corresponding to newly formed immature microvessels with leakage of fluorescein) in MMP-2,9 double KO mice compared with MMP-2-or MMP-9-deficient mice. This was associated with a strong inhibition of neovascular progression estimated on day 14 after induction by immunostaining with anti-PECAM antibodies in combined KO mice (Fig. 1D) as compared with each single-deficient mouse (Fig. 1B, C) or WT (Fig. 1A). Choroidal lesion associated with neovascularization was then quantified by determining the B/C ratio between total lesion thickness (B, maximal height lesion measured from the bottom of the choroid to the top of the neovascular area) to the thickness of adjacent normal choroid (C). A significant reduction of the B/C ratio was observed in MMP-9 (33%), MMP-2 (44%), and MMP-2,9 (56%) -deficient mice vs. their corresponding WT (PϽ0.001, Fig. 1E). Fibrinogen/fibrin accumulation in double MMP-2,9-deficient animalsConsistent with the modulation of fibrinolysis by matrix metalloproteinase system, we observed by immunohistochemical staining that fibrinogen/fibrin accumulated in double MMP-2,9-deficient animals. In contrast, similar fibrinogen/fibrin deposits were found in WT and single gene-deficient mice. These findings suggest that the absence of both gelatinases impaired fibrinolytic activity in choroidal neovascular membrane.1 To read the full text of this article, go to http://www.fasebj. org/cgi

The matrix metalloproteinases (MMPs) play a key role in normal and pathological angiogenesis by mediating extracellular matrix degradation and/or controlling the biological activity of growth factors, chemokines, and/or cytokines. Specific functions of individual MMPs as anti-or proangiogenic mediators remain to be elucidated. In the present study, we assessed the impact of single or combined MMP deficiencies in in vivo and in vitro models of angiogenesis (malignant keratinocyte transplantation and the aortic ring assay, respectively). MMP-9 was predominantly expressed by neutrophils in tumor transplants, whereas MMP-2 and MMP-3 were stromal. Neither the single deficiency of MMP-2, MMP-3, or MMP-9, nor the combined absence of MMP-9 and MMP-3 did impair tumor invasion and vascularization in vivo. However, there was a striking cooperative effect in double MMP-2:MMP-9-deficient mice as demonstrated by the absence of tumor vascularization and invasion. In contrast, the combined lack of MMP-2 and MMP-9 did not impair the in vitro capillary outgrowth from aortic rings. These results point to the importance of a cross talk between several host cells for the in vivo tumor promoting and angiogenic effects of MMP-2 and MMP-9. Our data demonstrate for the first time in an experimental model that MMP-2 and MMP-9 cooperate in promoting the in vivo invasive and angiogenic phenotype of malignant keratinocytes. Keywordsangiogenesis; tumor invasion; proteolysis; gelatinases; stromal MMP Matrix metalloproteinases (MMPs) are a family of structurally related zinc-and calciumdependent endopeptidases that can degrade extracellular matrix (ECM) components (1,2).

Plasminogen activator inhibitor type 1 (PAI-1) plays a key role in tumor progression and is believed to control proteolytic activity and cell migration during angiogenesis. We report here that host PAI-1, at physiological concentration, promotes in vivo tumor invasion and angiogenesis. In sharp contrast, inhibition of tumor vascularization was observed when PAI-1 was produced at supraphysiologic levels, either by host cells (transgenic mice overexpressing PAI-1) or by tumor cells (after transfection with murine PAI-1 cDNA). This study provides for the first time in vivo evidence for a dosedependent effect of PAI-1 on tumor angiogenesis. Of great interest is the finding that PAI-1 produced by tumor cells, even at high concentration, did not overcome the absence of PAI-1 in the host, emphasizing the importance of the cellular source of PAI-1.

New blood formation or angiogenesis has become a key target in therapeutic strategies aimed at inhibiting tumor growth and other diseases associated with neovascularization. Angiogenesis is associated with important extracellular remodeling involving different proteolytic systems among which the plasminogen system plays an essential role. It belongs to the large serine proteinase family and can act directly or indirectly by activating matrix metalloproteinases or by liberating growth factors and cytokines sequestered within the extracellular matrix. Migration of endothelial cells is associated with significant upregulation of proteolysis and conversely, immunoneutralization or chemical inhibition of the system reduces angiogenesis in vitro. On the other hand genetically altered mice developed normally without overt vascular anomalies indicating the possibility of compensation by other proteases in vivo. Nevertheless, they have in some experimental settings revealed unanticipated roles for previously characterized proteinases or their inhibitors. In this review, the complex mechanisms of action of the serine proteases in pathological angiogenesis are summarized alongside possible therapeutic applications.