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)

We previously suggested that periodontal pathogens might mediate connective tissue degradation in periodontal diseases through the ability of antigens from their cell walls to stimulate cytokine production by circulating mononuclear cells. Such cytokines would then induce metalloproteinase (MP) synthesis by resident gingival cells and thus initiate matrix degradation. In the present investigation human gingival fibroblasts (HGFs) were grown on [14C]-labelled type I collagen films and stimulated with either tumor necrosis factor (TNF) or interleukin-1 (IL-1) for 48 h. Collagenolysis occurred in a dose-dependent manner; the optimal dose for human rTNF alpha was 100 ng/ml and for rIL-1 alpha and rIL-1 beta, 1 ng/ml. Collagen degradation was accompanied by increased synthesis and release of the MPs collagenase, gelatinase and stromelysin, and there was a reduction in free TIMP (tissue inhibitor of metalloproteinases): collagenase and stromelysin were detected in both active and latent forms. Cytokine-stimulated collagenolysis was abolished by the addition of exogenous human rTIMP (5 units/ml). We also measured collagenase and TIMP by ELISAs which recognize all forms of collagenase (latent, active or complexed) and TIMP (free or complexed). These showed that while collagenase activity (0.6-1.2 microgram/ml) correlated with lysis, total TIMP levels remained unchanged at approximately 0.2 microgram/ml. These results demonstrate important roles for MPs and TIMP in regulating type I collagen degradation by HGFs, and support the hypothesis that connective tissue destruction during inflammatory diseases may be initiated, at least in part, by TNF and IL-1.
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PMID:Gingival fibroblasts degrade type I collagen films when stimulated with tumor necrosis factor and interleukin 1: evidence that breakdown is mediated by metalloproteinases. 255 Jun 4

Bovine aortic medial tissue and medial smooth muscle cells were demonstrated for the first time to synthesize a latent collagenase together with collagenase inhibitor in culture. Molecular weights of the latent collagenase and its inhibitor derived from aortic medial tissue explant were estimated to be about 52 K by gel filtration and 26.5 K by electrophoresis, respectively. Activated aortic collagenases cleaved type I collagen in solution into 3/4 (alpha A) and 1/4 (alpha B) length cleavage fragments and were inhibited by EDTA, o-phenanthroline, dithiothreitol, bovine serum, and highly purified dental pulp and aortic collagenase inhibitors. The aortic inhibitors showed inhibitory activity against all the animal collagenases tested, except for bacterial collagenase. Double-immunodiffusion analysis using a monospecific antiserum prepared against dental pulp inhibitor showed that the aortic inhibitors are immunologically identical to the pulp inhibitor. Using the same antiserum, we found immunoreactive collagenase inhibitor protein to be localized along the collagen fibers between elastic membranes in aortic medial tissue.
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PMID:Synthesis of latent collagenase and collagenase inhibitor by bovine aortic medial explants and cultured medial smooth muscle cells. 255 72

The atrophy of extracellular matrix is a common event during the aging of connective tissues. In this study, we tested the hypothesis that the altered ability of senescent cells to be modulated by serum growth factors correlated with a loss of regulation of collagenase synthesis. We examined the levels of immunoreactive procollagenase and collagenase inhibitor (the tissue inhibitor of metalloproteinases, TIMP) associated with young and senescent fibroblasts cultured in vitro. Young fibroblasts cultured in low (0.5%) concentrations of fetal bovine serum respond to increased (10%) serum by increasing levels of procollagenase and TIMP beginning 4.0 h after serum stimulation. In contrast, senescent fibroblasts constitutively produce relatively high levels of procollagenase even when cultured in low levels of serum and do not respond to serum stimulation by increasing procollagenase synthesis. In addition, senescent fibroblasts constitutively express a relatively small amount of TIMP which is not induced upon serum stimulation. This altered expression of collagenase and TIMP appears unique to the senescent phenotype and not merely a result of growth inhibition, since young cells growth arrested by density-dependent growth inhibition displayed a temporal pattern of procollagenase and TIMP expression upon serum stimulation similar to that of subconfluent young cultures. An assay of net collagenase activity revealed a greater than 20-fold elevation of activity in trypsin-activated extracts from senescent versus young fibroblasts when cultured in a low concentration of fetal bovine serum. These results demonstrate for the first time a direct correlation between alterations in the molecular pathways regulating connective tissue homeostasis and those of replicative senescence. The increased collagenolytic activity of senescent compared to young fibroblasts cultured in the presence of a low serum concentration suggests that aging fibroblasts may become increasingly fibroclastic causing many of the age-associated alterations in dermal collagen observed during aging in vivo.
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PMID:Replicative senescence of human skin fibroblasts correlates with a loss of regulation and overexpression of collagenase activity. 255 4

Two metalloproteinase inhibitors were purified from serum-free medium conditioned by bovine aortic endothelial cells. One of these inhibitors, with a molecular weight of 30,000-34,000 (reduced) is identified as tissue inhibitor of metalloproteinases; the second inhibitor has a molecular weight of 27,500 (reduced) and 20,400 (unreduced), is not recognized by an antiserum against bovine tissue inhibitor of metalloproteinases, appears unglycosylated, and has 51% identity with tissue inhibitor of metalloproteinases by NH2-terminal amino acid sequence analysis. This inhibitor has antiproteinase activities similar to those of tissue inhibitor of metalloproteinases, with inhibition of classical collagenase, type IV collagenase, and gelatinases but not trypsin, plasmin, or bacterial collagenase. Other properties shared with tissue inhibitor of metalloproteinases include trypsin sensitivity, acid and heat resistance, and inactivation by reduction-alkylation. The presence of these inhibitors in endothelial cells suggests that they may play important roles in protecting the integrity of the vascular basement membrane.
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PMID:Purification and characterization of two related but distinct metalloproteinase inhibitors secreted by bovine aortic endothelial cells. 255 3

The effect of various metalloproteinase-inhibiting compounds on collagen phagocytosis by fibroblasts was studied in cultured periosteal tissue. Evidence is presented indicating that neither anti-collagenase nor anti-stromelysin interfere with the uptake of collagen fibrils from the extracellular space and their intracellular digestion. Similar results were obtained with tissue inhibitor of metalloproteinases (TIMP). In the presence of the proteinase inhibitor leupeptin, a compound which strongly inhibits the intracellular degradation of phagocytosed collagen, a time-dependent increase in the amount of internalized collagen was found. This increase proved to be similar in explants treated as well as in those not treated with the metalloproteinase-inhibiting compounds. It is concluded that enzymes, such as collagenase and stromelysin, do not play a crucial role in the phagocytosis and intracellular digestion of collagen fibrils by fibroblasts. If these enzymes are involved it must be prior to these events. Based on the morphometric data the intralysosomal degradation time of collagen was calculated to be about 30 minutes. A comparison with findings in the literature on collagen metabolism in the periodontal ligament of the rat molar suggests that all collagen degraded may pass through the phagolysome pathway during physiological turnover and remodelling.
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PMID:Metalloproteinases are not involved in the phagocytosis of collagen fibrils by fibroblasts. 255 68

Mast cells have been implicated in the pathogenesis of the matrix degradation observed in the cartilaginous and osseous structures of the rheumatoid joint. We previously reported that human mast cell tryptase, a 134-kD granule-associated neutral protease, is present in rheumatoid synovium and can activate collagenase in crude culture medium in vitro. the present study attempts to depict the precise mechanism of this activation. To express full activation of latent collagenase, matrix metalloproteinase 3 (MMP-3) or stromelysin, can be activated by tryptase in a time and dose-dependent manner. Tryptase was not capable of generating active collagenase in the crude media from cultured rheumatoid synoviocytes depleted of proMMP-3 by immunoadsorption. In addition, the function of the tissue inhibitor of metalloproteinases (TIMP) was not altered by tryptase, and SDS-PAGE analysis revealed no degradation of TIMP by tryptase. The tryptase dependent activation of synoviocyte procollagenase thereby appears to be entirely dependent upon its ability to activate proMMP-3.
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PMID:Synovial procollagenase activation by human mast cell tryptase dependence upon matrix metalloproteinase 3 activation. 255 80

To understand the mechanisms regulating osteoid removal by osteoblasts, mouse calvarial osteoblasts were grown on 14C-labelled type I collagen films and stimulated with 1,25-dihydroxyvitamin D-3 (2.5.10(-8) M) for 48-72 h. In the presence of 5% non-inhibitory rabbit serum this resulted in a 2-3-fold increase in collagen degradation and a dramatic change in osteoblast morphology, when compared with untreated osteoblasts. Collagenolysis was accompanied by increased synthesis and release of latent collagenase, gelatinase and stromelysin and a concomitant decrease in their specific inhibitor, TIMP (tissue inhibitor of metalloproteinases). In serum-free medium, osteoblasts failed to degrade collagen, but their ability to lyse collagen could be restored by adding plasminogen (5 micrograms/ml) to the cultures. Plasminogen-dependent collagenolysis was inhibited by human recombinant TIMP (5 units/ml), demonstrating that plasmin, derived from plasminogen, activated latent collagenase and did not itself degrade collagen. Plasminogen activator production was confirmed by culturing osteoblasts on 125I-labelled fibrin plates. Comparison with urokinase-type and tissue-type plasminogen activator standards suggested that osteoblast plasminogen activator was predominantly cell-associated and likely to be of the urokinase type. Immunocytochemistry indicated that osteoblasts also constitutively produce plasminogen activator inhibitor-1. These findings provide evidence for the involvement of a plasminogen-plasmin-latent metalloproteinase activation cascade in type I collagen degradation by osteoblasts, and for its regulation by TIMP and plasminogen activator inhibitor-1.
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PMID:Type I collagen degradation by mouse calvarial osteoblasts stimulated with 1,25-dihydroxyvitamin D-3: evidence for a plasminogen-plasmin-metalloproteinase activation cascade. 255 72

Three forms of collagenase inhibitor were isolated; one bound to Con A-Sepharose and the other two did not. The Con A-bound (Mr 38,000) and the two unbound (Mr 50,000 and 22,000) inhibitors contained about 20, 15 and 65% of the total inhibitory activity, respectively. The bound and one of the unbound (Mr 22,000) inhibitors were fairly specific for mammalian collagenase; the other unbound inhibitor was rather non-specific and also inhibited trypsin and thermolysin, but not bacterial collagenase. All the inhibitors were heat stable (90 degrees C, 30 min) and unaffected by 4-aminophenylmercuric acetate, but were inactivated by reduction and alkylation.
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PMID:Separation and partial characterization of three forms of collagenase inhibitor from bovine gingiva. 255 85

On purification, human fibroblast collagenase breaks down into two major forms (Mr22,000 and Mr 27,000) and one minor form (Mr 25,000). The most likely mechanism is autolysis, although the presence of contaminating enzymes cannot be excluded. From N-terminal sequencing studies, the 22,000-Mr fragment contains the active site; differential binding to concanavalin A shows the 25,000-Mr fragment is a glycosylated form of the 22,000-Mr fragment. These low-Mr forms can be separated by Zn2+-chelate chromatography. An activity profile of this column, combined with data from substrate gels, indicates no activity against collagen in the 22,000-Mr and 25,000-Mr forms, but rather, activity casein and gelatin. The 27,000-Mr form has no activity. The 22,000/25,000-Mr form can act as an activator for collagenase in a similar way to that reported for stromelysin. The activity of the 22,000/25,000-Mr form is not inhibited by the tissue inhibitor of metalloproteinases (TIMP). The 27,000-Mr C-terminal part of the collagenase molecule therefore appears to be important in maintaining the substrate-specificity of the enzyme, and also plays a role in the binding of TIMP.
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PMID:Fragments of human fibroblast collagenase. Purification and characterization. 255 22

In the transition from proliferating to hypertrophic cell zones in the growth plate, there is an increased in chondrocyte cell volume and a corresponding decrease in collagen content to allow for cell enlargement. To substantiate our hypothesis that collagenase is responsible for these changes, growth plates from rats treated with bisphosphonate (HEBP) were compared histologically and biochemically with growth plates from normal and vitamin D and phosphate deficient (-VDP) rats. HEBP-treated rats developed an expanded hypertrophic cell zone (HCZ) characterized by the presence of two distinct populations of hypertrophic cells. The proximal hypertrophic cells were only 2-fold enlarged compared to the proliferating cells, whereas 1/6 of the distal hypertrophic cells were enlarged almost 5-fold and appeared morphologically identical with hypertrophic cells from normal and -VDP rats. The HEBP growth plates were divided into cross-sectional thirds and analyzed for active and latent collagenase. The juxta-metaphyseal (lower 1/3) cartilage contained 100% of the fully enlarged hypertrophic cells and appeared identical to those found in normal and -VDP growth plates, along with 81% of the active and 77% of the total collagenase. Collagenase and tissue inhibitor of metalloproteinases (TIMP) were measured in extracts of similarly divided tissues. The presence of true collagenas was confirmed by using [3H]-telopeptide-free collagen. TIMP levels were inversely related to the presence of active collagenase and cellular hypertrophy. Substantial levels of latent collagenase were found in the extracellular fluid at sites of active collagenolysis, but not in the fluid phase surrounding the 2-fold enlarged hypertrophic cells. It is postulated that increased amounts of active collagenase and insufficient levels of TIMP may account for the reduced collagen content seen in the lower HCZ of both -VDP and HEBP rickets. Unlike active collagenase, which remains localized by binding to collagen, latent enzyme is probably restricted in its mobility throughout the extracellular space by diffusion, itself, or the interstices of the extracellular matrix.
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PMID:Association of collagenase and tissue inhibitor of metalloproteinases (TIMP) with hypertrophic cell enlargement in the growth plate. 255 3


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