Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.24.17 (MMP-3)
 3,419 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We have compared the effects of a general matrix metalloproteinase (MMP) inhibitor (CT435) with those of a concentration-dependent specific gelatinase inhibitor (CT543; Ki < 20 nM) on bone resorption in vitro. The test systems consisted of measuring: (i) the release of 45Ca2+ from prelabelled mouse calvarial explants; (ii) the release of 45Ca2+ from prelabelled osteoid-free calvarial explants co-cultured with purified chicken osteoclasts; and (iii) lacunar resorption by isolated rat osteoclasts cultured on ivory slices. Both CT435 and CT543 dose-dependently inhibited the release of 45Ca2+ from neonatal calvarial bones stimulated by either parathyroid hormone or 1,25-dihydroxyvitamin D3. Moreover, CT543 produced a 40% inhibition at a concentration (10(-8) M) selective for the inhibition of human gelatinases A and B. CT435 (10(-5) M) and CT543 (10(-5) M) partially inhibited the release of 45Ca2+ from osteoid-free calvarial explants by chicken osteoclasts with a maximum of approximately 25% for unstimulated cultures, and approximately 36% for cultures stimulated by interleukin-1 alpha (IL-1 alpha; 10(-10) M). Neither inhibitor prevented lacunar resorption on ivory by unstimulated rat osteoclasts, but the compounds produced a partial reduction in both the number and total surface area of lacunae in IL-1 alpha-stimulated cultures, with maximal action at 10(-5) M. Neither of the inhibitors affected protein or DNA synthesis, nor the IL-1 alpha-stimulated secretion of the lysosomal enzyme beta-glucuronidase. Immunocytochemistry demonstrated that isolated rabbit osteoclasts constitutively expressed gelatinase A and synthesized gelatinase B, collagenase and stromelysin, as well as the tissue inhibitor of matrix metalloproteinases-1 (TIMP-1) following IL-1 alpha stimulation. These experiments have shown that in addition to collagenase, gelatinases A and B are likely to play a significant role in bone resorption. They further suggest that MMPs produced by osteoclasts are released into the sub-osteoclastic resorption zone where they participate in bone collagen degradation.
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PMID:The effects of selective inhibitors of matrix metalloproteinases (MMPs) on bone resorption and the identification of MMPs and TIMP-1 in isolated osteoclasts. 769 5

Scleroderma (systemic sclerosis: SSc) is an autoimmune disorder in which excessive extracellular matrix is deposited in skin and internal organs. Because of the importance of metalloproteinases in the turnover of connective tissue, in this study we have developed a novel procedure which utilises flow cytometry (FACS) to measure the production of stromelysin (MMP-3), gelatinase A (MMP-2), and the proteinase inhibitor TIMP-1, by SSc skin fibroblasts. In the presence of monensin, which prevents the secretion of these matrix proteins, there was a similar intracellular accumulation of gelatinase A in normal and SSc cells. However, whereas stromelysin levels also increased in the normal cells, no net synthesis could be detected in the SSc fibroblasts. In marked contrast, the synthesis of TIMP-1 was 50% greater in the SSc cells than in the normal fibroblasts. Our results thus show unequivocally, for the first time, that cells from SSc patients simultaneously produce less stromelysin but substantially higher amounts of TIMP-1 than do normal dermal fibroblasts, suggesting that abnormalities in the regulation of the matrix enzymes and their inhibitors play an important part in the molecular pathology of SSc.
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PMID:Excess matrix accumulation in scleroderma is caused partly by differential regulation of stromelysin and TIMP-1 synthesis. 770 50

Several members of the matrix metalloproteinase family have been reported to cleave aggrecan in the interglobular domain between Asn-341 and Phe-342. An antiserum was prepared against a peptide conjugate corresponding to the C-terminal sequence of the matrix metalloproteinase-generated aggrecan G1 fragment (Phe335-Val-Asp-Ile-Pro-Glu-Asn341). A quantitative radioimmunoassay, with a limit of detection of about 80 pM, was developed using this antiserum. This antiserum requires the free carboxyl group of the C-terminal asparagine for optimal recognition. If the C-terminal asparagine is excised from the sequence, replaced with closely related amino acids, or extended across the matrix metalloproteinase cleavage site, there is a 40-10,000-fold loss in detection. Using peptides cleaved from the N-terminus, it was determined that the antiserum requires the entire Phe-Val-Asp-Ile-Pro-Glu-Asn sequence for optimal recognition. The radioimmunoassay detects matrix metalloproteinase-generated G1 fragments with similar sensitivity to the Phe-Val-Asp-Ile-Pro-Glu-Asn peptide, but it does not recognize intact aggrecan. Immunoreactive aggrecan G1 fragments of molecular mass 50 kDa are generated by the matrix metalloproteinases stromelysin and gelatinase A. In contrast, under identical conditions, the closely related metalloproteinases, gelatinase B and collagenase, as well as cathepsin G, cathepsin B and human leucocyte elastase, did not generate a G1 fragment recognized by the antiserum. The anti-Phe-Val-Asp-Ile-Pro-Glu-Asn serum detects stromelysin-generated aggrecan G1 fragments from mouse, guinea pig, rabbit and human, indicating that the detection is not species-specific. This antiserum and radio-immunoassay should be useful for quantifying and characterizing matrix metalloproteinase-generated aggrecan G1 fragments in articular cartilage and synovial fluids from humans and various animal models of articular-cartilage destruction.
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PMID:Quantification of a matrix metalloproteinase-generated aggrecan G1 fragment using monospecific anti-peptide serum. 771 83

Matrix metalloproteinases share high protein sequence homology and have defined domain structures. Gelatinases have a unique 19 kDa fibronectin-like insert in the catalytic domain. A synthetic gene was made to express the catalytic domain of human gelatinase A (GCD), in which two polypeptide fragments of the catalytic domain were joined with deletion of the insert. The synthetic gene was highly expressed in Escherichia coli, and the 19 kDa GCD was purified to homogeneity after in vitro refolding. The GCD showed activity at a pH range of 5.5-9 in cleavage of the thiopeptolide Ac-Pro-Leu-Gly-thioester-Leu-Leu-Gly-OEt with optimal activity at neutral pH (Km = 134 microM and kcat = 16 s-1 at pH 7.0). The activity required both zinc and calcium ions, but high concentration of zinc ion showed inhibition. Several stromelysin catalytic domain inhibitors inhibited the GCD with similar specificity. The GCD cleaved the fluorogenic peptides Mca-Pro-Leu-Gly-Leu-Dpa-Ala-Arg-NH2 and Dnp-Pro-Leu-Gly-Leu-Trp-Ala-D-Arg-NH2 with catalytic efficiency close to full length human gelatinase A. The reconstructed GCD cleaves not only thiopeptolide and peptide substrates but also protein substrates such as gelatin. These results are consistent with the notion that gelatinases have the same structure for the catalytic domain as other matrix metalloproteinases like stromelysins and collagenases.
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PMID:Reconstructed 19 kDa catalytic domain of gelatinase A is an active proteinase. 771 75

Stromelysins are a group of proteases which degrade the extracellular matrix and activate other secreted proteases. Stromelysin (ST)-1 and ST-2 genes are induced by tumor promoters, oncogenes and growth factors, and have been involved in acquisition of the malignant phenotype. We show here that the thyroid hormone (T3) increases ST-1 and ST-2 expression in a non-transformed mouse mammary epithelial cell line (EpH4) in a way that is dependent on the level of thyroid receptor/c-erbA (TR alpha-1) expression. In agreement with this, T3 increases the secreted stromelysin activity and enhances the gelatinolytic activity of type IV collagenase. We have also demonstrated that T3 affects the epithelial polarity of EpH4 cells, diminishing the transepithelial electrical resistance of monolayers cultured on permeable filters, causing an abnormal distribution of polarization markers and the disruption of the organized 3-D structures formed by these cells in type I collagen gels. These results indicate that the ligand-activated TR alpha-1 plays an important role in regulating the morphogenetic and invasive capacities of mammary epithelial cells. Because the c-erbA locus is altered in several types of carcinoma, an altered or deregulated TR alpha-1 expression may also be important for breast cancer development and metastasis.
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PMID:Thyroid hormone regulates stromelysin expression, protease secretion and the morphogenetic potential of normal polarized mammary epithelial cells. 772 Jul 5

An Ets-related E1A-F has been characterized as an enhancer-binding protein for the adenovirus E1A gene. Here we show, in transient expression assays, that E1A-F can activate three different subclasses of the matrix metalloproteinase gene promoters. Expressions of the chloramphenicol acetyltransferase (CAT) reporter gene under the control of stromelysin, type I collagenase and 92 kD type IV collagenase promoters were increased approximately 10- to 20-fold by co-transfection with the E1A-F expression vector. Activation levels were as much high as those obtained by exogenous expression of AP-1 transcription factor. These results suggest that E1A-F positively regulates transcriptions from matrix metalloproteinase genes that are associated with invasion and metastasis of tumor cells.
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PMID:Ets-related protein E1A-F can activate three different matrix metalloproteinase gene promoters. 773

No measurable amounts of matrix metalloproteinases (MMPs) were produced by human breast adenocarcinoma cell lines MCF-7 and BT-20 in culture. When MCF-7 cells were co-cultured with human dermal fibroblasts enhanced production of precursors of MMP-1 (interstitial collagenase), MMP-2 (gelatinase A), MMP-3 (stromelysin 1) and tissue inhibitor of metalloproteinase type 1 (TIMP-1) was observed. Immunohistochemical studies indicated that these pro-MMPs originated primarily from the fibroblasts, suggesting that MCF-7 cells have a stimulatory effect on stromal cells to produce at least three pro-MMPs and TIMP-1. BT-20 cells also enhanced the production of pro-MMP-2 and TIMP-1 in the dermal fibroblasts, but not of pro-MMP-1 and pro-MMP-3. Normal mammary epithelial cells promoted only TIMP-1 production. To investigate further the stimulatory factors from MCF-7 cells, the conditioned medium and the cell membrane were prepared and examined. The cell membrane fraction enhanced the production of pro-MMP-1 and -3 and TIMP-1, but not of pro-MMP-2. The conditioned medium, on the other hand, augmented the production of all four proteins in the fibroblasts. These observations suggest that breast adenocarcinoma MCF-7 cells in culture produce both soluble and membrane-bound factor(s) which stimulate the production of pro-MMPs and TIMP-1 in neighbouring stromal cells, but the factor(s) released into the medium and that associated with cell membranes are probably different. Such communication between the normal and malignant cell types may, in part, assist the cancer cells to invade and metastasise.
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PMID:Co-culture of human breast adenocarcinoma MCF-7 cells and human dermal fibroblasts enhances the production of matrix metalloproteinases 1, 2 and 3 in fibroblasts. 773 96

Tissue inhibitor of metalloproteinases (TIMP)-2 forms a noncovalent complex with the precursor of matrix metalloproteinase 2 (proMMP-2, progelatinase A) through interaction of the C-terminal domain of each molecule. We have isolated the proMMP-2-TIMP-2 complex from the medium of human uterine cervical fibroblasts and investigated the processes involved in its activation by 4-aminophenylmercuric acetate (APMA). The treatment of the complex with APMA-activated proMMP-2 by disrupting the Cys73-Zn2+ interaction of the zymogen. This is triggered by perturbation of the proMMP-2 molecule, but not by the reaction of the SH group of Cys73 with APMA. The 'activated' proMMP-2 (proMMP-2*) formed a new complex with TIMP-2 by binding to the N-terminal inhibitory domain of the inhibitor without processing the propeptide. Thus the APMA-treated proMMP-2*-TIMP-2 complex exhibited no gelatinolytic activity. In the presence of a small amount of free MMP-2, however, proMMP-2* in the complex was converted into the 65 kDa MMP-2 by proteolytic attack of MMP-2, but the complex did not exhibit gelatinolytic activity. The gelatinolytic activity detected after APMA treatment was solely derived from the activation of free proMMP-2. The removal of the propeptide of the proMMP-2* bound to TIMP-2 was also observed by MMP-3 (stromelysin 1), but not by MMP-1 (interstitial collagenase). MMP-3 cleaved the Asn80-Tyr81 bond of proMMP-2*. On the other hand, when MMP-3 was incubated with the proMMP-2-TIMP-2 complex, it bound to TIMP-2 and rendered proMMP-2 readily activatable by APMA. These results indicate that the blockage of TIMP-2 of the complex with an active MMP is essential for the activation of proMMP-2 when it is complexed with TIMP-2.
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PMID:Steps involved in activation of the complex of pro-matrix metalloproteinase 2 (progelatinase A) and tissue inhibitor of metalloproteinases (TIMP)-2 by 4-aminophenylmercuric acetate. 777 54

Studies suggest that the interplay between matrix metalloproteinases (MMPs) and their inhibitors, tissue inhibitor of metalloproteinases (TIMPs), is an important mediator of tumour invasion and metastasis. Using immunohistochemistry, 40 specimens of colorectal cancer were examined for the presence of TIMP-1 and the MMPs, stromelysin, gelatinases A and B and interstitial collagenase. Neither enzyme nor TIMP-1 was detected in histologically normal mucosa. Within malignant tissue, stromelysin and gelatinase A were conspicuously absent in tumour cells but were immunolocalized to the extracellular matrix and for gelatinase A also to peritumoural fibroblast-like cells. Gelatinase B was confined to polymorphonuclear leucocytes. Interstitial collagenase was not identified. TIMP-1 was present in only three of the 40 tumours within the malignant stroma. These observations suggest that the mesenchymal elements of colorectal carcinomas, by acting as a source of MMPs and TIMPs, may modulate tumour invasion.
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PMID:The distribution of matrix metalloproteinases and tissue inhibitor of metalloproteinases in colorectal cancer. 778 Jun 9

Histological studies have previously demonstrated an association between mast-cell activation/degranulation and areas of connective-tissue lysis in vivo; in addition, mast-cell extracts have been shown to activate latent forms of collagenase and stromelysin. In the present study we have examined the potential roles of rat mast-cell proteinase (RMCP) I and RMCP II as activators of the precursors of matrix metalloproteinase (MMP)-1 (interstitial collagenase), MMP-2 (gelatinase A) and MMP-3 (stromelysin 1). Both RMCPs I and II activated proMMP-3 by converting the 57 kDa precursor into a 45 kDa polypeptide. The N-terminal amino acid of 45 kDa MMP-3 activated by RMCP II was identified as Phe83. By contrast, only RMCP II activated the 52 kDa proMMP-1 by converting it into a 41 kDa protein and generating the new N-termini, namely Gln80 and Val82. The collagenolytic activity which resulted from this cleavage was only 35% of the full activity, but this could not be augmented by subsequent treatment with MMP-3, the latter being a crucial enzyme for the generation of the fully active MMP-1 with Phe81 at the N-terminus, in conjunction with other serine proteinases. Thus RMCP II activates proMMP-1 via a mechanism different from that reported for the stepwise processing by combinations of other trypsin-like enzymes and MMP-3. ProMMP-2 (pro-gelatinase A) was not activated by either RMCP I or RMCP II, despite processing to smaller products.
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PMID:Activation of precursors for matrix metalloproteinases 1 (interstitial collagenase) and 3 (stromelysin) by rat mast-cell proteinases I and II. 782 45


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