Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.4.24.3 (collagenase)
18,340 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Several vertebrate collagenases have been reported to cleave type II collagen, leading to irreversible tissue destruction in osteoarthritis. We have investigated the action of MMP-1 and MMP-13 on type II collagen by use of neoepitope antibodies and N-terminal sequencing. Previous studies have suggested that the initial cleavage of type II collagen by MMP-13 is followed by a second cleavage, three amino acids carboxy-terminal to the primary cleavage site. We show here that this cleavage is also produced by APMA-activated MMP-1 in combination with MMP-3 (i.e. fully activated MMP-1). The use of a selective inhibitor of MMP-3 has shown that it is this enzyme, rather than interstitial collagenase which had been exposed to MMP-3, which makes the second cleavage. In addition we have identified, through N-terminal sequencing, a third cleavage site, three residues carboxy-terminal to the secondary site. Since MMP-2 is thought to be responsible for gelatinolytic action on type II collagen we have investigated the effect of MMP-2 after the initial helical cleavage made by either MMP-1 or MMP-13. A combination of MMPs-1, -2 and -3 results in both the second and third cleavage sites; adding MMP-2 to MMP-13 did not alter the cleavage pattern produced by MMP-13 on its own. We conclude that none of the three cleavage sites will provide information about the specific identity of the collagenolytic enzymes involved in collagen cleavage in situ. Staining of cartilage sections of osteoarthritis patients with the neoepitope antibodies revealed type II collagen degradation starting at or near the articular surface and extending into the mid and deep zones with increasing degeneration of the cartilage.
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PMID:Characterization of helical cleavages in type II collagen generated by matrixins. 948 Aug 69

Two groups of patients were studied, both in accordance with ACR criteria. First group (41 cases) suffering R.A. Second group (36 cases) suffering O.A. In both pathologies MMPs, ICAM and VCAM from synovial fluid and plasma were studied. Measurements were made with ELISA-sandwich in a Metrolab spectrophotometer at 410 nm for MMPs, and 491 nm for ICAM and VCAM. As control, samples of patients with noninflammatory muscle skeletal disorders or traumatic arthritis and healthy witness were used. Synovial concentration of MMPs in R.A. was 1402 +/- 76 ng/ml, a higher significant value (p < 0.0001) compared with osteoarthritis: 353 +/- 23 ng/ml. In the witness plasma, MMPs were not detected. Plasmatic and synovial levels of the adhesion molecules present different values in both pathologies and between them. Synovial ICAM level in R.A. (280 +/- 9.8 ng/ml) is significantly higher than in O.A. (163 +/- 10 ng/ml) (p < 0.001), but lower than the plasmatic ones (370 +/- 35 ng/ml) (p < 0.001). All these values are significantly higher than the normal plasma (121 +/- 6.5 ng/ml) (p < 0.01, p < 0.005, and p < 0.0001, respectively) VCAM increase regarding basal values (140 +/- 5.6 ng/ml) (p < 0.001) and in a similar proportion for both pathologies (R.A.: 186 +/- 9.3 ng/ml and O.A.: 207 +/- 14.3 ng/ml). Their plasmatic levels were higher (270 +/- 45 and 320 +/- 38 ng/ml) (p < 0.001) but without significative difference between them. There is correlation among MMPs, ICAM and VCAM variations. The variability can be explained by concomitance several evolutive steps. Each pathology shows a different grade of cellularity, inverted predominance in the relation TIMPs/ collagenase and different generator mechanisms of MMPs. Our findings reinforce the importance as diagnostic guide of adhesion molecules dosage, and possible therapeutic use of MMPs inhibitors and ICAM or VCAM antagonists en R.A. and related pathologies.
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PMID:Proteoglycan's activation by adhesion molecules and L metalloproteases in rheumatoid arthritis and osteoarthritis. 950 84

Cartilage, bone and the interstitial stroma, composed largely of the interstitial collagens, types I, II and III, are remodelled by three members of the metalloproteinase (MMP) family, collagenase-1 (MMP-1), collagenase-2 (MMP-8) and collagenase-3 (MMP-13). MMP-1 and MMP-13 may contribute directly to disease progression, since they are induced in patients with rheumatoid arthritis and osteoarthritis. The study of MMP-1 and MMP-13 gene regulation in models of arthritic disease has been problematic because mice and rats, which are typically used, only possess a homologue of MMP-13. Here we show that in contrast with mice and rats, rabbits possess distinct genes homologous to human MMP-1 and MMP-13. Furthermore, rabbit MMP-13 is expressed simultaneously with MMP-1 in chondrocytes and synovial fibroblasts in response to the cytokines interleukin-1 and tumour necrosis factor-alpha, or the phorbol ester PMA. The time course of MMP-13 induction is more rapid and transient than that of MMP-1, suggesting that distinct mechanisms regulate the expression of these two collagenases. We have cloned the rabbit MMP-13 gene from synovial fibroblasts and demonstrated that the rabbit gene shares greater homology with human MMP-13 than does the mouse interstitial collagenase. Together with the fact that mice and rats do not possess a homologue to human MMP-1, our data suggest that the rabbit provides an appropriate model for studying the roles of interstitial collagenases in connective-tissue diseases, such as rheumatoid arthritis and osteoarthritis.
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PMID:Cloning of the gene for interstitial collagenase-3 (matrix metalloproteinase-13) from rabbit synovial fibroblasts: differential expression with collagenase-1 (matrix metalloproteinase-1). 951 98

Osteoarthritis is the most common joint disease in humans. It is characterized by a gradual loss of extracellular matrix components of articular cartilage such as collagen and proteoglycan. Presently, however, emphasis is placed on enzymes exerting a strong influence on cartilage degradation. These enzymes include matrix metalloproteinases (MMP), their specific inhibitors (TIMP) and the plasminogen activator/inhibitor system. We applied monoclonal antibodies against MMP-1, -2, -3, -9 and their inhibitors TIMP-1/-2, as well as against urokinase-plasminogen activator u-PA and its inhibitor PAI to investigate their influence on articular cartilage degradation in patients with varusgonarthritis. We examined the cartilage of the lateral and medial compartments of 20 tibia plateaus, which can present with slight and severe cartilage degradations at the same time. In doing so, we tried to show whether or not immunohistological detection of enzymes could serve as a parameter for chondral degradation. The strongest immunoreaction for all enzymes was noted in the superficial layer of articular cartilage both medially and laterally. Between medial and lateral compartments, however, there were striking differences in the immunoreaction intensity of chondrocytes for MMP-1 and -3 as well as for TIMP-1 and u-PA. We noted that in cartilage with more advanced degradation, the immunoreaction for these enzymes was significantly higher in medial than in lateral compartments (p < 0.05). At the immunohistological level, a direct correlation between the grade of cartilage degradation and immunoreaction intensity was found. Our results corroborate the assumption that the expression of certain matrix-degradating enzymes serves as a parameter for the grade of cartilage degradation.
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PMID:Immunohistochemical analysis of several proteolytic enzymes as parameters of cartilage degradation. 958 19

Collagenase and gelatinase are matrix metalloproteinases (MMPs) which play an important role in tissue destruction in arthritic joints. Studies have demonstrated that tetracyclines can inhibit MMPs and prevent tissue destruction independent of their antimicrobial activity. The purpose of this pilot study is to assess the potential therapeutic role of Doxycycline in patients with advanced osteoarthritis of the temporomandibular joint (TMJ). This ongoing investigation includes patients with a diagnosis of osteoarthritis of the TMJ based on clinical and diagnostic imaging findings, symptoms (localized TMJ pain, limited mobility, dysfunction) for a minimum of 36 months, and failure of previous non-surgical and surgical modalities to alleviate the symptoms. A synovial fluid sample is collected by a saline injection and aspiration technique, followed by diagnostic arthroscopy. Patients are placed on Doxycycline 50 mg BID for three months and then undergo repeat diagnostic arthroscopy and synovial fluid collection. The samples are stored at -80 degrees C. Collagenase activity is determined by a combination of SDS-polyacrylamide gel electrophoresis and fluorography and calculated based on the percentage of collagen alpha chains that are degraded into alphaA breakdown products. Three patients have completed the three-month course of Doxycycline thus far, and 5 joints with osteoarthritis have been analyzed. All patients were female (mean age = 35, mean duration of symptoms = 132 months) and had undergone previous bilateral arthroscopies. One patient had undergone unilateral arthroplasty. The mean collagenase activity showed 55% collagen lysis prior to Doxycycline treatment and 19% after three months of therapy. The mean gelatinase activity was 28% prior to Doxycycline treatment and 7% after three months of therapy. The mean interincisal opening was 33 mm initially and 41 mm after three months of Doxycycline. Subjectively, two of the three patients reported significant improvement in their overall symptoms, which they had not experienced over the previous three years. One patient did not experience any change in symptoms, in spite of a marked reduction in collagenase activity from 86.4% to 9.6%. Because of the very small numbers of patients enrolled in this pilot study so far, no statistically significant differences could be appreciated. However, the dramatic reduction in collagenase activity in these patients, with a long history of TMJ symptoms from osteoarthritis, suggests the potential promising role of Doxycycline in the management of osteoarthritis, and further investigation is warranted.
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PMID:The potential role of doxycycline in the treatment of osteoarthritis of the temporomandibular joint. 997 22

Growth of and metalloproteinase production by fibroblast-like synoviocytes (FLSs) in patients with rheumatoid arthritis (RA) contribute to cartilage and bone destruction associated with development of the expanding inflammatory tissue referred to as pannus. Increased levels of extracellular matrix (ECM) proteins in the pannus suggest that intracellular signals generated through integrin receptors might control these processes. We developed a cell culture system permitting accurate assessment of the effect of cell adhesion to various ECM proteins on FLS phenotype. We show that FLS proliferation to platelet-derived growth factor requires a second signal provided by adhesion to an ECM protein. Fibronectin, vitronectin, collagen, or laminin could provide the second signal and was similarly required for the proliferation of FLSs from RA or osteoarthritis patients. Adhesion to fibronectin, collagen, or Arg-Gly-Asp peptide down-regulated collagenase expression. Primarily alphav integrin receptors mediated this down-regulation upon adhesion to fibronectin. Loss of cell adhesion and TNF-alpha stimulation synergistically increased collagenase expression. Increased collagenase expression upon nonadherence was mimicked by treatment with cytochalasin B, suggesting that the loss of cytoskeletal structure associated with a change in cell shape mediates increased collagenase in nonadherent cells. Thus, although increased fibronectin in the lining layer in RA might be expected to inhibit collagenase expression, the change in cell shape associated with this multilayer structure might actually lead to increased collagenase expression.
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PMID:Integrin engagement regulates proliferation and collagenase expression of rheumatoid synovial fibroblasts. 997 41

The possible mechanism of the chondroprotective effect of 6,7-dihydroxycoumarin (esculetin) was investigated using primary cultures of rabbit articular chondrocytes. Esculetin (EST) significantly suppressed the proteoglycan depletion and the release of pulse-labeled [35S]proteoglycan from the matrix layer of rabbit chondrocytes treated with recombinant human interleukin-1alpha. The matrix metalloproteinase inhibitor, 1,10-phenanthroline, also blocked the proteoglycan depletion and [35S]proteoglycan release. From these results, it is likely that recombinant human interleukin-1alpha-induced proteoglycan depletion is mediated by matrix metalloproteinases. Although esculetin did not directly inhibit collagenolytic activity in the culture media, it significantly suppressed the production of pro-matrix metalloproteinase-1/interstitial procollagenase and pro-matrix metalloproteinase-3/prostromelysin 1, accompanied by a decrease in the steady-state levels of their mRNAs. These results suggest that esculetin is a therapeutically effective candidate for inhibition of cartilage destruction in osteoarthritis and rheumatoid arthritis.
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PMID:Esculetin suppresses proteoglycan metabolism by inhibiting the production of matrix metalloproteinases in rabbit chondrocytes. 1033 6

Synovial fluid basic calcium phosphate (BCP) crystals are markers of severe joint degeneration in osteoarthritis. These crystals are mitogenic and induce protooncogene expression and matrix metalloproteinase (MMP) synthesis and secretion in human fibroblasts, effects that are specifically blocked by phosphocitrate (PC). We have recently determined that crystals transduce signals to the nucleus via the activation of the p42 and p44 mitogen-activated protein (MAP) kinases (Nair et al., 1997, J Biol Chem 272:18920-18925). Treatment of human fibroblasts (HF) with BCP induces phosphorylation of p42/44 MAPK, which is inhibited by PC in a dose-dependent manner. Blocking of p42/44 MAPK signal transduction with an inhibitor (PD98059) of MEK1, an upstream activator of MAPKs, reduces crystal-induced p42/44 MAPK activation and significantly inhibits crystal-induced cell proliferation. Based on these findings, we sought to determine the role of the p42/44 MAPK signal transduction pathway in crystal-induced expression of matrix MMPs. We demonstrate suppression of crystal-induced MMPs via the utilization of two different MEK inhibitors: PD98059 and the recently described U0126, a novel inhibitor of MEK1 and MEK2. Treatment of HF with PD98059 blocks the induction of crystal-stimulated collagenase 1 (MMP-1) and stromelysin (MMP-3) expression. PD98059 and PC reduced the level of crystal-induced MMP-1 and MMP-3 mRNA expression to that observed in nonstimulated cells. Likewise, PD98059 treatment of HF blocked the epidermal growth factor (EGF)- and crystal-induced increases in MMP-1 and MMP-3 protein expression and secretion as demonstrated by Western blotting and zymography. Treatment of HF with U0126 inhibits EGF-induced phosphorylation of p42/44 MAPK as well as crystal- and EGF-induced upregulation of MMP-1 mRNA. Additionally, we demonstrate that treatment of HF with BCP, EGF, or PD98059 does not significantly alter levels of gelatinase A (MMP-2) mRNA and protein expression.
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PMID:Basic calcium phosphate crystal induction of collagenase 1 and stromelysin expression is dependent on a p42/44 mitogen-activated protein kinase signal transduction pathway. 1039 91

The pseudojoint cavity formed in patients undergoing total hip arthroplasty (THA) is later remodeled to synovial membrane-like tissue, which produces pseudosynovial fluid. This pseudosynovium also is an important source of matrix metalloproteinases (MMPs). As it is widely speculated that synovial fluid MMPs may contribute to local tissue degradation in rheumatoid arthritis (RA) and osteoarthritis (OA), we hypothesize that locally produced MMPs are found in the pseudosynovial fluid, via which they have access to the implant-host interface, and that if they retain their proteolytic potential, they might contribute to aseptic loosening. Enzyme-linked immunosorbent assay (ELISA), immunoblotting, and zymography were used to analyze MMPs and tissue inhibitors of metalloproteinases (TIMPs) in synovial fluid in aseptic loosening, which was compared to RA and OA. Pseudosynovial THA fluid was characterized using low levels of MMP-1 but moderate levels of MMP-13 and MT1-MMP (MMP-14). Due to the lack of an appropriate assay, MMP-13 and MT1-MMP were not similarly assessed, but the immunoblotting indicated that they were in the 56 kD intermediate proteolytically processed forms. The MMP-9 level was intermediate between RA and OA. MMP-2 was on a significant level, but there were no differences among study groups. The THA group also was characterized using relatively high levels of TIMP-1 and TIMP-2. Accordingly, MMP-9 and MMP-2 were found to occur in the 92 kD and 72 kD proenzyme form, respectively, with full activity retained in all study groups. The data suggest that proMMP-2-TIMP-2 and proMMP-9-TIMP-1 complexes are formed in the pseudosynovial fluid due to the excess of TIMPs over MMPs in aseptic loosening of THA. TIMP-complexed MMPs are resistant to MMP-mediated proteolytic activation, which may explain their latency and proenzyme zymogen form. Thus, formation of stabilizing proMMP-TIMP complexes enable transportation of proMMPs far from their original site of production. Due to motion-associated cyclic changes of the intra-articular pressure, fluid-phase MMPs stabilized by TIMPs might be absorbed to implant surfaces and interface tissues and help to dissect the implant/cement-to-bone interface in situ. Consequently, they may contribute to local proteolytic/tissue destructive events and aseptic loosening.
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PMID:Matrix metalloproteinases and tissue inhibitors of metalloproteinases in joint fluid of the patients with loose artificial hip joints. 1039 73

Degradation of type II collagen is thought to be a key step in the destruction of articular cartilage in patients with rheumatoid arthritis or osteoarthritis. The aim of this study was to investigate whether type II collagen degradation is associated with cartilage destruction. Type II collagen degradation was studied in two murine arthritis models, zymosan-induced arthritis (ZIA), which develops reversible articular cartilage damage based on proteoglycan analysis, and antigen-induced arthritis (AIA), in which there is irreversible damage to the cartilage. Type II collagen degradation was assayed immunohistochemically using the COL2-3/4m antibody which recognizes denatured type II collagen, such as is produced by collagenase cleavage. In both models, degradation of type II collagen was observed in the non-calcified articular cartilage of arthritic but not of control knees. In the patella-femoral compartment, collagen denaturation started to increase on day 3 (ZIA) and day 7 (AIA) and remained high on day 14. In contrast, in the tibia-femoral compartment, type II collagen breakdown was not increased before 14 days in either model. By 28 days, collagen denaturation was strongly reduced in the patella-femoral compartment in the ZIA model, but persisted in the tibia-femoral compartment in both models. In conclusion, increased type II collagen degradation was found in articular cartilage of both ZIA and AIA animals. Since ZIA does not develop irreversible cartilage destruction, this indicates that cartilage may have the ability to withstand a limited degree of type II collagen degradation without developing irreversible damage.
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PMID:Denaturation of type II collagen in articular cartilage in experimental murine arthritis. Evidence for collagen degradation in both reversible and irreversible cartilage damage. 1041 4


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