Dylan R. Edwards

Transforming growth factor beta modulates the expression of collagenase and metalloproteinase inhibitor.

Exposure of quiescent MRC‐5 human fibroblasts to growth factors such as epidermal growth factor, basic fibroblast growth factor or embryonal carcinoma‐derived growth factor resulted in the induction of mRNA transcripts encoding the metalloproteinases collagenase and stromelysin and the specific metalloproteinase inhibitor TIMP, whilst expression of collagen and fibronectin was relatively unaffected. Exposure of quiescent cells to growth factors in the presence of transforming growth factor beta (TGF‐beta) resulted in inhibition of collagenase induction and a synergistic increase in TIMP expression. TGF‐beta alone did not significantly induce metalloproteinase or TIMP expression. These effects on mRNA transcripts were reflected in increased secretion of TIMP protein and collagenase activity. Nuclear run‐off analysis of growth factor‐induced transcription revealed that the TGF‐beta modulation of TIMP and collagenase expression was due to transcriptional mechanisms. The observations suggest that TGF‐beta exerts a selective effect on extracellular matrix deposition by modulating the action of other growth factors on metalloproteinase and TIMP expression.

Metalloproteinase inhibitors: Biological actions and therapeutic opportunities

Tissue inhibitors of metalloproteinases (TIMPs) are the major cellular inhibitors of the matrix metalloproteinase (MMP) sub-family, exhibiting varying efficacy against different members, as well as different tissue expression patterns and modes of regulation. Other proteins have modest inhibitory activity against some of the MMPs, including domains of netrins, the procollagen C-terminal proteinase enhancer (PCPE), the reversion-inducing cysteine-rich protein with Kazal motifs (RECK), and tissue factor pathway inhibitor (TFPI-2), but their physiological significance is not at all clear.α 2-Macroglobulin, thrombospondin-1 and thrombospondin-2 can bind to some MMPs and act as agents for their removal from the extracellular environment. In contrast, few effective inhibitors of other members of the metzincin family, the astacins or the distintegrin metalloproteinases, ADAMs have been identified. Many of these MMP inhibitors, including the TIMPs, possess other biological activities which may not be related to their inhibitory capacities. These need to be thoroughly characterized in order to allow informed development of MMP inhibitors as potential therapeutic agents. Over activity of MMPs has been implicated in many diseases, including those of the cardiovascular system, arthritis and cancer. The development of synthetic small molecule inhibitors has been actively pursued for some time, but the concept of the use of the natural inhibitors, such as the TIMPs, in gene based therapies is being assessed in animal models and should provide useful insights into the cell biology of degradative diseases.

Metalloproteinases in biology and pathology of the nervous system

Matrix metalloproteinases (MMPs) have been implicated in several diseases of the nervous system. Here we review the evidence that supports this idea and discuss the possible mechanisms of MMP action. We then consider some of the beneficial functions of MMPs during neural development and speculate on their roles in repair after brain injury. We also introduce a family of proteins known as ADAMs (a disintegrin and metalloproteinase), as some of the properties previously ascribed to MMPs are possibly the result of ADAM activity.

The ADAM metalloproteinases

Abstract The ADAMs (a disintegrin and metalloproteinase) are a fascinating family of transmembrane and secreted proteins with important roles in regulating cell phenotype via their effects on cell adhesion, migration, proteolysis and signalling. Though all ADAMs contain metalloproteinase domains, in humans only 13 of the 21 genes in the family encode functional proteases, indicating that at least for the other eight members, protein–protein interactions are critical aspects of their biological functions. The functional ADAM metalloproteinases are involved in “ectodomain shedding” of diverse growth factors, cytokines, receptors and adhesion molecules. The archetypal activity is shown by ADAM-17 (tumour necrosis factor-α convertase, TACE), which is the principal protease involved in the activation of pro-TNF-α, but whose sheddase functions cover a broad range of cell surface molecules. In particular, ADAM-17 is required for generation of the active forms of Epidermal Growth Factor Receptor (EGFR) ligands, and its function is essential for the development of epithelial tissues. Several other ADAMs have important sheddase functions in particular tissue contexts. Another major family member, ADAM-10, is a principal player in signalling via the Notch and Eph/ephrin pathways. For a growing number of substrates, foremost among them being Notch, cleavage by ADAM sheddases is essential for their subsequent “regulated intramembrane proteolysis” (RIP), which generates cleaved intracellular domains that translocate to the nucleus and regulate gene transcription. Several ADAMs play roles in spermatogenesis and sperm function, potentially by effecting maturation of sperm and their adhesion and migration in the uterus. Other non-catalytic ADAMs function in the CNS via effects on guidance mechanisms. The ADAM family are thus fundamental to many control processes in development and homeostasis, and unsurprisingly they are also linked to pathological states when their functions are dysregulated, including cancer, cardiovascular disease, asthma, Alzheimer’s disease. This review will provide an overview of current knowledge of the human ADAMs, discussing their structure, function, regulation and disease involvement.

Matrix metalloproteinases and diseases of the CNS

Matrix metalloproteinases (MMPs) are increasingly being implicated in the pathogenesis of several CNS diseases. In multiple sclerosis, MMPs could be responsible for the influx of inflammatory mononuclear cells into the CNS, contribute to myelin destruction and disrupt the integrity of the blood-brain barrier; in Alzheimers disease, MMPs might mediate the deposition of amyloid beta-proteins; and MMPs are known to contribute to the invasiveness of malignant glioma cells and might regulate their angiogenic capacity. Nonetheless, MMPs could also have beneficial roles in recovery from CNS injury.Therefore, both the identity of the MMP and its cellular origin could determine whether disease pathogenesis or regeneration occurs, and thus synthetic MMP inhibitors might be valuable for treating some CNS diseases.

The ADAMTS metalloproteinases

The ADAMTSs (a disintegrin and metalloproteinase with thrombospondin motifs) are a group of proteases that are found both in mammals and invertebrates. Since the prototype ADAMTS-1 was first described in 1997, there has been a rapidly expanding body of literature describing this gene family and the proteins they encode. The complete human family has 19 ADAMTS genes, together with three members of a newly identified subgroup, the ADAMTSL (ADAMTS-like) proteins, which have several domains in common with the ADAMTSs. The ADAMTSs are extracellular, multidomain enzymes whose known functions include: (i) collagen processing as procollagen N-proteinase; (ii) cleavage of the matrix proteoglycans aggrecan, versican and brevican; (iii) inhibition of angiogenesis; and (iv) blood coagulation homoeostasis as the von Willebrand factor cleaving protease. Roles in organogenesis, inflammation and fertility are also apparent. Recently, some ADAMTS genes have been found to show altered expression in arthritis and various cancers. This review highlights progress in understanding the structural organization and functional roles of the ADAMTSs in normal and pathological conditions.

The regulation of matrix metalloproteinases and their inhibitors

The matrix metalloproteinases (MMP) are a family of 23 enzymes in man. These enzymes were originally described as cleaving extracellular matrix (ECM) substrates with a predominant role in ECM homeostasis, but it is now clear that they have much wider functionality. Control over MMP and/or tissue inhibitor of metalloproteinases (TIMP) activity in vivo occurs at different levels and involves factors such as regulation of gene expression, activation of zymogens and inhibition of active enzymes by specific inhibitors. Whilst these enzymes and inhibitors have clear roles in physiological tissue turnover and homeostasis, if control of their expression or activity is lost, they contribute to a number of pathologies including e.g. cancer, arthritis and cardiovascular disease. The expression of many MMPs and TIMPs is regulated at the level of transcription by a variety of growth factors, cytokines and chemokines, though post-transcriptional pathways may contribute to this regulation in specific cases. The contribution of epigenetic modifications has also been uncovered in recent years. The promoter regions of many of these genes have been, at least partly, characterised including the role of identified single nucleotide polymorphisms. This article aims to review current knowledge across these gene families and use a bioinformatic approach to fill the gaps where no functional data are available.

Gelatinase-A (MMP-2), gelatinase-B (MMP-9) and membrane type matrix metalloproteinase-1 (MT1-MMP) are involved in different aspects of the pathophysiology of malignant gliomas.

SummaryMatrix metalloproteinases (MMPs) have been implicated as important factors in gliomas since they may both facilitate invasion into the surrounding brain and participate in neovascularization. We have tested the hypothesis that deregulated expression of gelatinase-A or B, or an activator of gelatinase-A, MT1-MMP, may contribute directly to human gliomas by quantifying the expression of these MMPs in 46 brain tumour specimens and seven control tissues. Quantitative RT-PCR and gelatin zymography showed that gelatinase-A in glioma specimens was higher than in normal tissue; these were significantly elevated in low grade gliomas and remained elevated in GBMs. Gelatinase-B transcript and activity levels were also higher than in normal brain and more strongly correlated with tumour grade. We did not see a close relationship between the levels of expression of MT1-MMP mRNA and amounts of activated gelatinase-A. In situ hybridization localized gelatinase-A and MT1-MMP transcripts to normal neuronal and glia, malignant glioma cells and blood vessels. In contrast, gelatinase-B showed a more restricted pattern of expression; it was strongly expressed in blood vessels at proliferating margins, as well as tumour cells in some cases. These data suggest that gelatinase-A, -B and MT1-MMP are important in the pathophysiology of human gliomas. The primary role of gelatinase-B may lie in remodelling associated with neovascularization, whereas gelatinase-A and MT1-MMP may be involved in both glial invasion and angiogenesis.

Increased gelatinase A (MMP-2) and gelatinase B (MMP-9) activities in human brain after focal ischemia

Matrix metalloproteinases (MMPs) are involved in remodelling extracellular matrix. Gelatinase B (MMP-9) is an inducible 92 kDa MMP expressed by neutrophils, microglia, and endothelial cells. Gelatinase A (MMP-2) is a 72 kDa MMP, constitutively expressed in brain. Elevated MMP activity has been linked to various pathologic conditions, and the therapeutic benefit of MMP inhibitors is under study in a few experimental models. Using gelatin zymography, we have compared activities of these MMPs in infarcted and matched non-infarcted cerebral tissue from eight subjects dying at intervals of less than 2 h to several years after a stroke. Gelatinase B activity was markedly elevated in the infarcted tissue at two days post-infarction, and remained elevated in cases dying months after the event. Increases in gelatinase A activity were subtle at 2-5 days; they were marked and significant in cases dying at 4 months and later. The findings indicate distinct temporal profiles of post-ischemic gelatinase activity in human brain, with earlier but equally persistent elevation in gelatinase B when compared to gelatinase A.

MicroRNAs and the hallmarks of cancer

It has become clear that particular microRNAs (miRNAs) function either as tumour suppressors or oncogenes, whose loss or overexpression, respectively, has diagnostic and prognostic significance. In several cases, miRNAs have been shown to affect target genes that are involved in the control of cell proliferation and apoptosis. However, malignant tumours display additional traits beyond the acquisition of enhanced growth potential and decreased cell death. Malignant disease is associated with altered tumour–host interactions leading to sustained angiogenesis and the ability to invade and metastasize. It is possible that miRNAs may act as master regulators of these aspects of tumour biology. Bioinformatic analysis of putative miRNA binding sites has indicated several novel potential gene targets of cancer-associated miRNAs that function in aspects of cell adhesion, neovascularization and tissue invasion. Among others, we speculate that miRNAs may find new roles in the regulation of E-cadherin, integrin αvβ3, hypoxia-inducible factor-1α, syndecan-1, lysyl oxidase, adamalysin metalloproteinase-17, tissue inhibitors of metalloproteinase-3, c-Met and CXCR-4 that underpin the tissue architectural changes associated with malignancy.