EC:3.4.24.17 - FACTA Search

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)

Metalloproteases appear to play an important role in the pathophysiology of osteoarthritis (OA) and their expression is believed to be regulated by cytokines such as interleukin-1 (IL-1). Nuclear oncogene products are suggested as mediators through which IL-1 induces metalloprotease gene expression. Little data are available on the in vivo involvement of these agents in the pathophysiology of OA. This study examined by immunohistochemistry, using specific antibodies, the distribution of stromelysin, IL-1 alpha, IL-1 beta, and oncogene products (c-FOS, c-JUN, and c-MYC) in synovium and cartilage from normal and experimental canine models of OA. In the OA synovium, stromelysin and IL-1 were localized in the cytoplasm of superficial synovial lining cells, infiltrating mononuclear cells, and endothelial and smooth muscle cells of the blood vessels, whereas oncoproteins were detected predominantly in the synovial lining cells. Normal synovial membranes demonstrated low levels of specific staining in synovial lining cells with occasional staining of blood vessel cells for IL-1 alpha, IL-1 beta, and stromelysin. In OA cartilage, chondrocytes at the superficial and middle layers as well as in fibrillated areas were found to be involved in the synthesis of stromelysin, IL-1, and oncoproteins. Diffuse staining of stromelysin and IL-1 beta in OA cartilage matrix was also identified. In normal cartilage, only a few chondrocytes at the superficial layer showed a low level of antigens. These results demonstrate the in vivo concomitant cellular and/or matrical presence of stromelysin, IL-1, and oncogene proteins in tissues from experimentally induced OA with the most intense staining at the sites of cartilage erosion and synovial proliferation. These findings suggest that they may be involved in the pathophysiology of OA, and that the regulatory mechanisms involved in the expression of these proteins may be associated.
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PMID:Coordinate synthesis of stromelysin, interleukin-1, and oncogene proteins in experimental osteoarthritis. An immunohistochemical study. 842 68

Recombinant human fibroblast pro-MMP-3 (prostromelysin-1) expressed in Chinese hamster ovary cells and the zymogen from cultured human dermal fibroblasts have been purified by monoclonal antibody immunoaffinity chromatography, and the role of Ca2+ in proenzyme activation and thermostability of the low mass catalytic domain of MMP-3 has been investigated. In the presence of high Ca2+ (5.0 mM), the organomercurial aminophenylmercuric acetate (APMA) initiated the stepwise removal of both NH2- and COOH-terminal domains from both recombinant and dermal fibroblast proenzymes, resulting in the generation of a heterogeneous family of nonglycosylated low mass truncated active enzyme species beginning at Phe83. However, in the presence of low Ca2+ (0.1 mM), incubation of recombinant pro-MMP-3 with or without APMA did not result in formation of either the high or low mass forms of active MMP-3 but resulted in complete autolysis of both enzyme species. The concentration of Ca2+ required for optimal pro-MMP-3 activation and stability of the low mass catalytic domain was 2.0 mM. The low mass truncated enzyme species containing the catalytic domain were remarkably heat-stable (90 min at 60 degrees C) in high Ca2+ (5.0 mM) but rapidly autolyzed when heated at 60 degrees C in low Ca2+ (0.1 mM). The thermostability properties of MMP-3 appeared to be specific for Ca2+, since no other divalent metal ions tested were able to confer thermostability to the low mass catalytic domain of MMP-3. From homology to the thermostable bacterial metalloprotease, thermolysin, two putative Ca2+ binding sites were found in the catalytic domain of MMP-3 and several other members of the MMP gene family. These putative Ca2+ binding sites are postulated to play an important role in stabilizing active MMP-3 and other members of the matrix metalloprotease gene family by protecting them against autolysis.
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PMID:Recombinant Chinese hamster ovary cell matrix metalloprotease-3 (MMP-3, stromelysin-1). Role of calcium in promatrix metalloprotease-3 (pro-MMP-3, prostromelysin-1) activation and thermostability of the low mass catalytic domain of MMP-3. 844 Jul 30

This study explores novel aspects of the interaction between inflammatory mediators and extracellular matrix degradation. Here we have evaluated the effects of a T-cell cytokine interleukin-4 (IL-4) on the expression and activity of a metalloprotease, stromelysin, and its tissue inhibitor (TIMP-1) in human skin fibroblasts. IL-4 strongly decreased stromelysin mRNA levels and stromelysin-producing activity induced by IL-1 beta-treated and untreated cells. Under the same experimental conditions, TIMP-1 mRNA expression was slightly modified. Phorbol ester (PMA), a PKC activator, induced stromelysin gene expression, an effect enhanced by the addition of IL-1 beta. IL-4 was not able to decrease the PMA and PMA + IL1 beta effects. Calphostin, a specific PKC inhibitor, inhibited stromelysin mRNA expression induced by IL-1 beta. Forskolin, a PKA activator, did not modify mRNA levels and was not able to reduce the effect of IL-4 on IL-1 beta-induced stromelysin expression. These data suggest that in human dermal fibroblasts, activation of PKC abolishes the observed IL-4 effect on both basal and IL-1 beta-induced stromelysin gene expression. It therefore appears that lack of PKC activation is a prerequisite for the inhibitory effect of IL-4 in the system.
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PMID:Inhibition by Interleukin-4 of stromelysin expression in human skin fibroblasts: role of PKC. 861 84

Reactive oxygen species (ROS) are important second messengers for the induction of several genes in a variety of physiological and pathological conditions. Ultraviolet B (UVB) irradiation has recently been shown to generate lipid peroxidation products and hydroxyl radicals (HO.) with detrimental long term effects like cancer formation and premature aging of the skin. Here, we addressed the question of whether ferric/ferrous iron via the generation of ROS may mediate the UVB response, finally leading to connective tissue degradation, a hallmark in carcinogenesis and aging. Therefore, we studied the involvement of iron and ROS in the modulation of Jun N-terminal kinase 2 (JNK2) activity, c-jun and c-fos mRNA levels, key signaling steps in the transcriptional control of matrix-degrading metalloprotease (MMP)-1/interstitial collagenase and MMP-3/stromelysin-1 after UVB irradiation of human dermal fibroblasts in vitro. The iron-driven generation of lipid peroxides and hydroxyl radicals were identified as early events in the downstream signaling pathway of the UVB response leading to a 15-fold increase in JNK2 activity, a 3.5-fold increase in c-jun, to a 6-fold increase in MMP-1, and a 3.8-fold increase in MMP-3 mRNA levels, while virtually no alteration of c-fos mRNA levels were observed. Diminished generation of reactive oxygen species resulted in a significant reduction of JNK2 activity, c-jun, MMP-1, and MMP-3 mRNA levels after UVB irradiation compared with UVB-irradiated cells. Collectively, we have identified the iron-driven Fenton reaction and lipid peroxidation as possible central mechanisms underlying signal transduction of the UVB response.
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PMID:Central role of Ferrous/Ferric iron in the ultraviolet B irradiation-mediated signaling pathway leading to increased interstitial collagenase (matrix-degrading metalloprotease (MMP)-1) and stromelysin-1 (MMP-3) mRNA levels in cultured human dermal fibroblasts. 947 85

Exposure of rats to hypoxia causes pulmonary arterial remodeling, which is partly reversible after return to air. We hypothesized that degradation of excess collagen in remodeled pulmonary arteries in the posthypoxic period is mediated by endogenous matrix metalloproteinases (MMPs). Total proteolytic, collagenolytic, and gelatinolytic activities, levels of stromelysin-1 and tissue inhibitor of metalloprotease-1 (TIMP-1), and immunolocalization of stromelysin-1 in main pulmonary arteries were determined after exposure of rats to 10% O2 for 10 days followed by normoxia. We observed transient increases in total proteolytic, collagenolytic, and gelatinolytic activities and expression of approximately 72-, 68-, and 60-kDa gelatinases by zymography within 3 days of cessation of hypoxic exposure. The level of TIMP-1 increased as the stromelysin-1 level increased. Immunoreactive stromelysin-1 was localized predominantly in the luminal region of normal and hypertensive pulmonary arteries. These results are consistent with the notion that endogenous MMPs may mediate the breakdown of excess collagen in remodeled pulmonary arteries during the early posthypoxic period.
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PMID:Expression of matrix-degrading enzymes in pulmonary vascular remodeling in the rat. 970 Jan 2

Peptide libraries generated using phage display have been widely applied to proteolytic enzymes for substrate selection and optimization, but the reaction kinetics between the enzyme and substrate phage are not well understood. Using a quantitative ELISA assay to monitor the disappearance of substrate, we have been able to follow the course of reaction between stromelysin, a metalloprotease, and its substrate phage. We found that under the proteolytic conditions where the enzyme was present in nanomolar concentration or higher, in excess over the substrate, the proteolysis of substrate phage was a single exponential event and the observed rate linear with respect to enzyme concentration. The enzyme concentration dependence could be described by pseudo first-order kinetic equations. Our data suggest that substrate binding is slow relative to the subsequent hydrolysis step, implying that the phage display selection process enriches clones that have high binding affinity to the protease, and the selection may not discriminate those of different chemical reactivity toward the enzyme. Considering that multiple substrate molecules may be present on a single phage particle, we regard the substrate phage reaction kinetic model as empirical. The validity of the model was ascertained when we successfully applied it to determine the binding affinity of a competitive inhibitor of stromelysin.
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PMID:Reaction kinetics of protease with substrate phage. Kinetic model developed using stromelysin. 1115 96

The extracellular matrix metalloproteases (MMPs) secreted by various human tumor cells play a crucial role in tumor cell invasion and metastasis, but their expression in malignant mesothelioma (MM) cells has not been examined. In this study, we have investigated the spectrum of MMPs and tissue inhibitors of metalloproteases (TIMPs) produced by 8 MM cell lines. Using RT-PCR, we found that all investigated MM cell lines expressed genes encoding mRNA for MMP-1 (interstitial collagenase), MMP-2 (gelatinase A), MMP-3 (stromelysin-1), MMP-9 (gelatinase B) and TIMPs 1, 2 and 3. We also found that 6/8 MM cell lines expressed MMP-7 (matrilysin) and 3/8 MM cell lines expressed MMP-10 (stromelysin-2). MMP-11 (stromelysin-3) was not detected in any of the MM cell lines. Production of MMP-2 and MMP-9 was confirmed using gelatin zymography. In addition, all MM cell lines secreted a 66 kDa metalloprotease, while 3/8 MM cell lines secreted 46, 48, 51 and 63 kDa metalloproteases which specifically degraded the extracellular matrix components fibronectin, vitronectin and laminin. The 66 kDa protease was identified as MMP-3 by Western blot. Our results reveal a broad spectrum of MMPs and TIMPs produced by MM cells and indicate that different substrate specificities of MMPs may play a role in MM cell invasion.
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PMID:Expression and activity of matrix metalloproteases in human malignant mesothelioma cell lines. 1126 73

Studies of aggrecan proteolysis in human joints have implicated both the aggrecanase [ADAMTS, a disintegrin-like and metalloprotease (reprolysin-type) with thrombospondin type 1 motif] and matrix metalloproteinase (MMP) families. We have analysed the aggrecan core protein species present in vivo in both articular cartilage and synovial fluids from normal, acutely injured and osteoarthritic joints. Normal cartilage contains at least seven major G1 domain (the N-terminal globular domain of aggrecan)-bearing species, of which three (full-length core, G1-NITEGE(373) and G1-VDIPEN(341)) have been identified. The C-terminals of the others are unknown but digestion of fetal human aggrecan with MMP-3 and crude aggrecanase suggests that they are products of MMP-like activity in vivo. Normal synovial fluids contain at least 10 species, of which nine result from ADAMTS-dependent cleavage, and this cleavage occurs at all of the five known aggrecanase sites. Aggrecan fragments in the cartilage and synovial fluids of acutely injured joints are generally similar to normal, but all contain a markedly increased ratio of G1-NITEGE to G1-VDIPEN. Aggrecan from the cartilage of late-stage osteoarthritis patients is remarkably similar to normal, whereas the synovial fluid aggrecan is more fragmented than that from normal or injured knees. The analyses suggest that the role of the ADAMTS and these MMP-like activities in human cartilage are distinctly different. Excessive ADAMTS activity in vivo is destructive to cartilage matrix, since the bulk of the glycosaminoglycan (GAG)-bearing products are released from the tissue into the synovial fluid following cleavage of the Glu(373)-Ala(374) bond. In contrast, the MMP-like activity appears to be essentially non-destructive, since much of the GAG-bearing product is retained in the tissue following cleavages that are in the more C-terminal regions of the molecule.
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PMID:Analysis of aggrecan in human knee cartilage and synovial fluid indicates that aggrecanase (ADAMTS) activity is responsible for the catabolic turnover and loss of whole aggrecan whereas other protease activity is required for C-terminal processing in vivo. 1153 23

Intracellular signals generated by mechanical strain profoundly affect the metabolic function of osteoblast-like periodontal ligament (PDL) cells, which reside between the tooth and alveolar bone. In response to applied mechanical forces, PDL cells synthesize bone-resorptive cytokines to induce bone resorption at sites exposed to compressive forces and deposit bone at sites exposed to tensile forces in an environment primed for catabolic processes. The intracellular mechanisms that regulate this bone remodeling remain unclear. Here, in an in vitro model system, we show that tensile strain is a critical determinant of PDL-cell metabolic functions. Equibiaxial tensile strain (TENS), when applied at low magnitudes, acts as a potent antagonist of interleukin (IL)-1beta actions and suppresses transcriptional regulation of multiple proinflammatory genes. This is evidenced by the fact that TENS at low magnitude: (i) inhibits recombinant human (rh)IL-1beta-dependent induction of cyclooxygenase-2 (COX-2) mRNA expression and production of prostaglandin estradiol (PGE2); (ii) inhibits rhIL-1beta-dependent induction matrix metalloproteinase-1 (MMP-1) and MMP-3 synthesis by suppressing their mRNA expression; (iii) abrogates rhIL-1beta-induced suppression of tissue inhibitor of metalloprotease-II (TIMP-II) expression; and (iv) reverses IL-1beta-dependent suppression of osteocalcin and alkaline phosphatase synthesis. Nevertheless, these actions of TENS were observed only in the presence of IL-1beta, as TENS alone failed to affect any of the aforementioned responses. The present findings are the first to show that intracellular signals generated by low-magnitude mechanical strain interfere with one or more critical step(s) in the signal transduction cascade of rhIL-1beta upstream of mRNA expression, while concurrently promoting the expression of osteogenic proteins in PDL cells.
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PMID:Signaling by mechanical strain involves transcriptional regulation of proinflammatory genes in human periodontal ligament cells in vitro. 1193 44

Gelatinase B is a member of the matrix metalloproteinase family that efficiently cleaves gelatin, elastin, and types V and X collagen. To understand the contribution of the active site of the enzyme (amino acid residues 373-456) in these activities, we studied catalytic properties of a fusion protein consisting of maltose binding protein and the active site region of gelatinase B. We found that addition of the active site of gelatinase B, which corresponds to 12% of the total protein molecule, to maltose binding protein is sufficient to endow the protein with the ability to cleave the peptide substrates Mca-PLGL(Dpa)AR-NH(2) and DNP-PLGLWA-(D)-R-NH(2). The fusion protein hydrolyzed the Mca-PLGL(Dpa)AR-NH(2) peptide with the same efficiency as that of the stromelysin, k(cat)/K(m) approximately 1.07 x 10(6) M(-)(1) h(-)(1). The fusion protein, however, was not able to degrade the large substrate, gelatin. Inhibition of the activity of the protein by EDTA suggested that its activity was metal dependent. ESR analyses indicated that the fusion protein bound one molecule of Zn(2+). In addition, Z-Pro-Leu-Gly-hydroxamate and TIMP-1 inhibited the activity of the protein, suggesting that the structure of the active site of the fusion protein is similar to that of the other metalloproteinases. These data provide fundamental information about the structural elements required for transforming a protein to a metalloprotease.
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PMID:Identification of the active site of gelatinase B as the structural element sufficient for converting a protein to a metalloprotease. 1193 73

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