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)

A novel polypeptide with a molecular weight of 17 kd (17k protein) was identified in bone marrow cell cultures. The synthesis of 17k protein is elevated in cell cultures maintained under Dexter conditions, which support myelopoiesis. The predominance of macrophages in the stromal layer of these cultures and the observation that a mouse myelomonocytic cell line P388D1 is capable of synthesizing large amounts of 17k protein led us to the study of its synthesis by bone marrow macrophages. Metabolic labeling with [14C]proline and partial amino acid analysis of 17k protein demonstrated that the polypeptide contains relatively high amounts of proline and is also sensitive to degradation with bacterial collagenase. However, no hydroxyproline is detectable in 17k protein, and it is extensively degraded with bacterial collagenase. However, no hydroxyproline is detectable in 17k protein, and it is extensively degraded by proteolysis with pepsin, using conditions under which collagen triple helices are resistant to degradation, suggesting that collagen-like structures are not contained in 17k protein. This polypeptide is found predominantly in the cellular layers of bone marrow macrophage cultures. Incorporation of [14C]proline into 17k protein is diminished by increasing concentrations of colony-stimulating factor 1 (CSF-1). The 17k protein may be involved in macrophage proliferation because its synthesis is inhibited by CSF-1, which is required for the maintenance of bone marrow macrophages in vitro.
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PMID:A 17-kd polypeptide, sensitive to bacterial collagenase, is synthesized by bone marrow macrophages. 216 40

The effects of a broad array of cytokines, individually and in combination, were determined on separate functions (proliferation, collagenase production, and granulocyte macrophage colony-stimulating factor [GM-CSF] production) and phenotype (expression of class II MHC antigens) of cultured fibroblast-like RA synoviocytes. The following recombinant cytokines were used: IL-1 beta, IL-2, IL-3, IL-4, IFN-gamma, tumor necrosis factor (TNF)-alpha, GM-CSF, and macrophage colony-stimulating factor (M-CSF). Only IFN-gamma induced HLA-DR (but not HLA-DQ) expression. TNF-alpha inhibited IFN-gamma-mediated HLA-DR expression (46.7 +/- 4.1% inhibition) and HLA-DR mRNA accumulation. This inhibitory effect was also observed in osteoarthritis synoviocytes. Only TNF-alpha and IL-1 increased synoviocyte proliferation (stimulation index 3.60 +/- 1.03 and 2.31 +/- 0.46, respectively). IFN-gamma (but none of the other cytokines) inhibited TNF-alpha-induced proliferation (70 +/- 14% inhibition) without affecting the activity of IL-1. Only IL-1 beta and TNF-alpha induced collagenase production (from less than 0.10 U/ml to 1.10 +/- 0.15 and 0.72 +/- 0.24, respectively). IFN-gamma decreased TNF-alpha-mediated collagenase production (69 +/- 19% inhibition) and GM-CSF production but had no effect on the action of IL-1. These data demonstrate mutual antagonism between IFN-gamma and TNF-alpha on fibroblast-like synoviocytes and suggest a novel homeostatic control mechanism that might be defective in RA where very little IFN-gamma is produced.
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PMID:Cytokines in chronic inflammatory arthritis. V. Mutual antagonism between interferon-gamma and tumor necrosis factor-alpha on HLA-DR expression, proliferation, collagenase production, and granulocyte macrophage colony-stimulating factor production by rheumatoid arthritis synoviocytes. 217 6

Degradation of the atherosclerotic plaque extracellular matrix could destabilize the lesion, rendering it more prone to rupture. Both macrophages and vascular smooth muscle cells (SMCs) are potential sources of matrix metalloproteinases (MMPs), secreted enzymes that can digest vascular matrix. We explored interactions between human vascular SMCs and human monocytes that result in the secretion of interstitial collagenase (MMP-1) and stromelysin (MMP-3). Monocytes alone or those treated with SMC-conditioned media did not secrete these metalloproteinases as detectable by Western blot analysis. SMCs increased secretion of both MMP-1 and MMP-3 greater than 20-fold when cocultured with monocytes or when treated with monocyte-conditioned media. Addition of macrophage colony stimulating factor (< or = 1000 U/mL) to cocultures of monocytes and SMCs did not affect metalloproteinase secretion. Recombinant interleukin (IL)-1 receptor antagonist inhibited MMP-1 and MMP-3 induction in SMC cultures treated with monocyte-conditioned media (94% and 96% reduction, respectively), while a neutralizing antibody to tumor necrosis factor-alpha had no significant effect on metalloproteinase secretion. In contrast to the induction by monocyte-conditioned media of MMP-1 and MMP-3 secretion by SMCs, monocyte-conditioned media did not increase secretion of 72-kD gelatinase (MMP-2). Thus, monocytes induce MMP-1 and MMP-3 secretion by vascular SMCs through an IL-1-dependent mechanism. This response of SMCs to a defined macrophage product may contribute to plaque destabilization by mononuclear phagocytes in the lesion.
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PMID:Human vascular smooth muscle cell-monocyte interactions and metalloproteinase secretion in culture. 748 54

Nitric oxide is a highly reactive mediator released in the liver by hepatocytes, Kupffer cells and endothelial cells during endotoxin-induced inflammation. In this study we determined whether Ito cells also produce nitric oxide after exposure to endotoxin. For induction of endotoxemia, rats were injected intravenously with Escherichia coli lipopolysaccharide (2.5 mg/kg). Ito cells were isolated from the animals 48 hr later by means of in situ perfusion of the liver with protease and collagenase followed by purification on an arabinogalactan gradient. Ito cells from untreated and endotoxemic rats were found to produce low levels of nitric oxide in response to interferon-gamma. In both cell types, this response depended on L-arginine and was blocked by NG-monomethyl-L-arginine, a specific nitric oxide synthase inhibitor. Cells from rats treated with endotoxin produced significantly more nitric oxide than did cells from untreated animals; this was due, at least in part, to increased expression of protein for an inducible form of nitric oxide synthase. These cells also responded to stimulation with lipopolysaccharide in vitro, as well as the combination of interferon-gamma and lipopolysaccharide, which was synergistic in stimulating nitric oxide production. Tumor necrosis factor-alpha and macrophage colony-stimulating factor were also found to stimulate nitric oxide production by Ito cells from endotoxemic rats. In addition, in these cells, tumor necrosis factor-alpha synergized with interferon-gamma in inducing nitric oxide production. The combination of interferon-gamma and lipopolysaccharide was also found to inhibit Ito cell DNA synthesis, as measured on the basis of [3H]-thymidine uptake.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Induction of hepatic Ito cell nitric oxide production after acute endotoxemia. 752 4

Biochemical and molecular studies of osteoclasts generally require cells in a reasonable degree of purity. The chicken has been extremely useful in this regard, as abundant avian osteoclasts can be generated in vitro entirely from pure populations of marrow macrophage precursors. Propagation of murine osteoclasts is, in contrast, far less efficient, demanding the presence of stromal cells. The aims of this study were to develop a method by which murine osteoclasts generated in culture, can be effectively enriched while maintaining viability and, to explore the mechanisms by which stromal cells promote murine osteoclast generation and survival. We find that 10(6) fractionated murine marrow cells enriched, for marrow-residing colony-forming units (CFU-cs), yield 3000-4000 tartrate-resistant acid phosphatase (TRAP)-expressing multinucleated giant cells when cultured for 12 days with ST-2 stromal cells. These cells are osteoclasts as evidenced by their ability to "pit" bone slices, resorb radiolabeled bone particles, and generate cyclic AMP in response to calcitonin. Treatment of these generated osteoclast cultures with bacterial collagenase for 2 hours at 37 degrees selectively removes virtually all ST-2 cells, yielding a > 60% pure population of TRAP and calcitonin receptor-expressing cells, 90% of which are viable. These cells continue to respond to calcitonin and survive for 24 hours in the absence of ST-2 cells. We also found that murine osteoclast generation depends upon contact of osteoclast precursors with viable ST-2 cells. Furthermore, the stromal cells secrete macrophage colony-stimulating factor (CSF-1), and the anti-CSF-1 antibody 5A1 inhibits murine osteoclastogenesis.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Enrichment of generated murine osteoclasts. 753 41

Treatment of rats with bacterially derived lipopolysaccharide (LPS), a condition that mimics acute endotoxemia, results in a significant increase in the number of endothelial cells and macrophages in the liver. This is correlated with the release of proinflammatory and cytotoxic mediators that induce liver damage. In the present studies, we analyzed the effects of various inflammatory mediators released during the pathogenesis of hepatic injury on proliferation of liver nonparenchymal cells. To induce acute endotoxemia female Sprague-Dawley rats were injected intravenously with 5 mg/kg LPS. Endothelial cells and macrophages were isolated 48 h later by combined collagenase and pronase perfusion of the liver followed by centrifugal elutriation. Interleukin-1 alpha (IL-1 alpha), interleukin-6 (IL-6), and tumor necrosis factor alpha (TNF-alpha) had no effect on proliferation of either endothelial cells or macrophages. In contrast, whereas interleukin-1 beta (IL-1 beta) inhibited the proliferation of endothelial cells from untreated rats, this cytokine stimulated the growth of cells from endotoxemic rats. The colony-stimulating factors, granulocyte-macrophage colony-stimulating factor (GM-CSF) and macrophage colony-stimulating factor (M-CSF), also markedly enhanced the proliferation of endothelial cells, as well as macrophages from endotoxemic rats. Macrophages from endotoxemic rats were more sensitive to the colony-stimulating factors than cells from untreated rats. In contrast, the inflammatory mediators LPS and interferon-gamma (IFN-gamma) inhibited endothelial cell and macrophage growth, an effect that was partially blocked in endothelial cells by the nitric oxide synthase inhibitor NG-monomethyl-L-arginine (L-NMMA). This suggests that growth inhibition in these cells is mediated, in part, by nitric oxide. Interestingly, in both endothelial cells and macrophages from endotoxemic rats, GM-CSF, M-CSF, and IL-1 beta synergized with LPS and IFN-gamma to induce nitric oxide production. This was correlated with a further inhibition of proliferation that was partially reversed by L-NMMA in endothelial cells but not macrophages. Taken together these data demonstrate that endothelial cell and macrophage proliferation in the liver is controlled by a variety of mediators released during endotoxemia; however, the mechanisms regulating growth in the two cell types are distinct.
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PMID:Regulation of hepatic endothelial cell and macrophage proliferation and nitric oxide production by GM-CSF, M-CSF, and IL-1 beta following acute endotoxemia. 814 21

The antigen-presenting cell system in the gastrointestinal tract, one of three main sites (skin and lung being the others) of primary antigen contact, is poorly understood. Our study focused on dendritic cells (DC) as possible candidates for antigen uptake, processing and presentation in mucosal inductive sites, such as Peyer's patches (PP). To investigate the morphology, immunophenotype and stimulatory activity of intestinal DC, a procedure was developed to obtain a cell population by using collagenase digestion of PP, density centrifugation and cell sorting on the basis of CD11c expression. The resultant low-density cell fraction consisted of a nonadherent cell population expressing different intensities of CD11c that could at least be characterized by typical DC morphology (e.g. abundant cytoplasma with veil-like cytoplasmatic dendrites, irregularly shaped nuclei, multivesicular and multilamellar bodies), constitutive levels of surface MHC class II, the presence of macrophage-specific markers, such as F4/80, Mac-I and Fc receptors, respectively, on subpopulations of CD11c+ sorted cells and expression of adhesion and co-stimulatory receptors like ICAM-1 and CD44. The capability of this low-density CD11c+ fraction to stimulate T cell responses was demonstrated in primary allogeneic mixed-lymphocyte reactions (MLR). Herein, we show that the freshly isolated CD11c+ cells showed weak accessory function, but develop this capacity following short-term culture in vitro in the presence of granulocyte/macrophage colony-stimulating factor. Although the nature and functional capacity of the isolated CD11c+ needs further clarification, these preliminary results describing phenotype and accessory function provide some evidence that these cells isolated from the PP may be immature forms of DC and play a crucial role as antigen-presenting cells with important implications for understanding the complex network regulating intestinal antigen uptake, processing and presentation.
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PMID:Phenotypic and functional characterization of CD11c+ dendritic cell population in mouse Peyer's patches. 876 24

1. We recently demonstrated the presence of phospholipase C-coupled bradykinin (BK) B2-receptors in human primary and SV40 virus-immortalized corneal epithelial (CEPI) cells. 2. The aims of the present studies were to demonstrate the specific binding of [3H]-BK to CEPI cell membranes and to study its pharmacological characteristics. In addition, we wished to study the functional coupling of the BK receptors to various physiological and pathological mechanisms in the CEPI cells, including phosphoinositide (PI) turnover, intracellular Ca2+-mobilization ([Ca2+]i), cell proliferation (via [3H]-thymidine incorporation), and the release of various cytokines, collagenase-1 (matrix metalloproteinase-1) and prostaglandin E2 (PGE2). 3. Specific [3H]-BK binding comprised 83 +/- 2% of the total binding, and was of high affinity (Kd = 1.66 +/- 0.52 nM, n = 5), saturable (Bmax = 640 +/- 154 fmol g(-1) wet weight) and reversible. Competition studies yielded the following affinity values for BK and a number of BK-related peptides: Hoe-140 (D-Arg-[Hyp3,Thi5,D-Tic7,Oic8]BK; icatibant): Ki = 0.17 +/- 0.07 nM; BK: Ki = 1.0 +/- 0.11 nM; [Tyr8]-BK: Ki = 12.9 +/- 2.3 nM; [des-Arg9]-BK: Ki > 9,200 nM (all n = 3-5)). 4. BK potently stimulated PI turnover (EC50 = 2.3 +/- 0.3 nM; n = 7) and [Ca2+]i mobilization (EC50 = 8-20 nM) in CEPI cells and both responses were inhibited in a concentration-dependent manner by 100 nM-10 microM Hoe-140, a selective B2-receptor antagonist, and also inhibited by the selective phospholipase C (PLC) inhibitor, U73122 (1-(6-((17beta-3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1 H-pyrrole-2,5-dione) (IC50 = 3.0 +/- 1.6 microM). BK-induced [Ca2+]i mobilization was reduced by about 30% in the presence of 4 mM EGTA, but was not significantly affected by 100 nM nifedipine. 5. BK (0.1 nM-10 microM) significantly (P<0.05-0.001) stimulated [3H]-thymidine incorporation into CEPI cellular DNA. However, while interleukin-1alpha (IL-1alpha; 10 ng ml(-1)) potently stimulated the release of IL-6, IL-8 and granulocyte macrophage colony-stimulating factor from CEPI cells, BK (0.1 nM-10 microM) was without effect. 6. Whilst phorbol-12-myristate-13-acetate (PMA; 3 microg ml(-1)) and 10% foetal bovine serum (positive control agents) significantly stimulated the release of both MMP-1 and PGE2 from CEPI cells, BK (0.1 nM-10 microM) was without any significant effect under these conditions. 7. In conclusion, these data indicate that the CEPI cells express high-affinity [3H]-BK binding sites representing B2-subtype BK receptors coupled to PI turnover and [Ca2+]i mobilization which appear to stimulate [3H]-thymidine incorporation into cellular DNA. In contrast, BK failed to elicit the release of PGE2, various cytokines and MMP-1 from CEPI cells. These results suggest that BK may have a potential role in corneal epithelium wound healing by stimulating cell proliferation.
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PMID:Effects of bradykinin on signal transduction, cell proliferation, and cytokine, prostaglandin E2 and collagenase-1 release from human corneal epithelial cells. 955 96

Osteoclast-like multinucleated cells (OCLs) were prepared on collagen gels in a coculture system of mouse bone marrow cells and osteoblasts, and purified by collagenase and a subsequent pronase treatment. More than 80% of the purified OCLs were found to undergo apoptotic cell death by 48 h during the culture in a culture medium containing 10% fetal bovine serum (FBS). Withdrawal of FBS from the culture medium accelerated the cell death, which induced more than 80% of OCLs to undergo apoptotic cell death by as early as 18 h. Two peptide inhibitors of caspases (interleukin-1beta-converting enzyme family proteases), benzyloxycarbonyl-Val-Ala-Asp (OMe)-fluoromethyl ketone (Z-VAD-FMK) and benzyloxycarbonyl-Asp-Glu-Val-Asp (OMe)-fluoromethyl ketone (Z-DEVD-FMK), extended the survival time of OCLs in the presence and absence of 10% FBS, but the effect was rather limited in the absence of FBS. Because interleukin-1alpha (IL-1alpha) and the macrophage colony stimulating factor (M-CSF) are known to promote the survival of osteoclasts, we examined the effect of the peptide inhibitors and these cytokines. Combinations of the peptide inhibitors and IL-1alpha, or the peptide inhibitors and M-CSF, were more effective than the inhibitors alone. When endogenous caspase activities of OCLs were analyzed using fluorescence peptide substrates, the activities, in particular, caspase-3 (CPP32)-like activity, were markedly increased in OCLs by the withdrawal of FBS from the culture medium. IL-1alpha and M-CSF suppressed the activation of the caspases. In addition, western blot analysis revealed that the expression of Bcl-2, which inhibits the activation of caspases, was very weak or even negligible in OCLs. Taken together, these results suggest that the caspases are involved in the regulation of survival and apoptotic cell death of osteoclasts.
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PMID:Caspases (interleukin-1beta-converting enzyme family proteases) are involved in the regulation of the survival of osteoclasts. 966 28

We recently reported on the successful generation of immortalized (CEPI-17-CL4) cells from primary human corneal epithelial (P-CEPI) cells which exhibited phenotypic, immunohistochemical and metabolic characteristics akin to the P-CEPI cells. The aims of the present studies were to investigate the ligand binding and functional coupling of the histamine receptors to various biochemical and physiological systems in the P-CEPI and CEPI-17-CL4 cells and to relate these findings to the normal and/or pathophysiological role of histamine on the human ocular surface. Specific [3H]-pyrilamine binding to CEPI-17-CL4 cell homogenates comprised >93% of the total binding and represented interaction with an apparent single population of high affinity (Kd=3.76+/-0.78 nM; n=4) and saturable (Bmax = 1582+/-161 fmol g(-1) tissue) number of histamine-1 (H1) receptor binding sites on CEPI-17-CL4 cell homogenates. The H1-receptor selective antagonists, pyrilamine (Ki=3.6+/-0.84 nM, n=4) and triprolidine (Ki = 7.7+/-2.6 nM, n=3), potently displaced [3H]-pyrilamine binding, while the H2- and H3-receptor selective antagonists, ranitidine and clobenpropit, were weak inhibitors (K(i)s>13 microM). Histamine induced phosphoinositide (PI) hydrolysis 2.7-4.4 fold above basal levels and with a potency of 14.9+/-4.9 microM (n=9) and 4.7+/-0.2 microM (n=9) in P-CEPI and CEPI-17-CL4 cells, respectively. Histamine-induced PI turnover was antagonized by H1-receptor selective antagonist, triprolidine, with a potency (Ki) of 3.2+/-0.66 nM (n=10) and 3.03+/-0.8 nM (n=4) in P-CEPI and CEPI-17-CL4 cells, respectively, but weakly effected by 10 microM cimetidine and clobenpropit, H2- and H3-receptor antagonists. The PI turnover response was attenuated by pre-treatment of the cells with the selective phospholipase C inhibitor, U73122 (1-(6-((17beta-3-methoxyestra- 1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione) (IC50=4.8+/-2.4 microM, n = 3). Histamine stimulated intracellular Ca2+ ([Ca2+]i) mobilization in CEPI-17-CL4 cells with a potency of 6.3+/-1.5 microM (n=4). The histamine-induced [Ca2+]i mobilization was reduced by about 28% following pre-incubation of the cells with 4 mM EGTA. While triprolidine completely inhibited histamine-induced [Ca2+]i mobilization, it did not influence the bradykinin-induced [Ca2+]i mobilization response. Histamine (EC50s = 1.28-2.77 microM, n=3-4) concentration-dependently stimulated the release of interleukin-6 (IL-6), IL-8 and granulocyte macrophage colony-stimulating factor, but it did not significantly alter release of tumour necrosis factor-alpha, PGE2 or collagenase-1 (matrix metalloproteinase-1; MMP-1) from CEPI cells. However, IL-1 (10 ng ml(-1)), foetal bovine serum (10%) and phorbol-12-myristate-13-acetate (3 microg ml(-1)) were effective positive control secretagogues of all the cytokines, PGE2 and MMP-1, respectively, from these cells. It is concluded that the CEPI cells express H1-histamine receptors which are positively coupled to PI turnover and [Ca2+]i mobilization which may be directly or indirectly responsible for the release of various cytokines from these cells at physiologically and/or pathologically relevant concentrations.
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PMID:Pharmacology of [3H]-pyrilamine binding and of the histamine-induced inositol phosphates generation, intracellular Ca2+ -mobilization and cytokine release from human corneal epithelial cells. 986 65


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