Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: UNIPROT:P43026 (lipopolysaccharide)
 62,215 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Matrilysin (PUMP, MMP-7) is a member of the metalloprotease gene family, whose constituents are responsible for the remodeling of extracellular matrix. The matrilysin protein is a 28-kDa zymogen possessing catalytic activities against a broad range of extracellular matrix substrates including proteoglycans, gelatin, fibronectin, laminin, and elastin. To gain insights into the biological expression of matrilysin in human cell types, we generated a monospecific, polyclonal antibody against a 16-amino acid sequence derived from its catalytic domain, a region which lacked significant homology with other matrix metalloenzymes. We found this antibody capable of precipitating a 28-kDa protein from the conditioned media of human bone marrow-derived promonocytes and human peripheral blood monocytes cultivated in vitro. Promonocyte matrilysin was rapidly converted to a 19-kDa form by organomercurial activation. While matrilysin was constitutively synthesized by bone marrow-derived promonocytes, its secretion was markedly up-regulated by the mononuclear phagocyte activator, lipopolysaccharide. Furthermore, despite its expression in monocyte precursors, blood monocytes, and monocyte-derived macrophages, matrilysin was not synthesized by human alveolar macrophages under any tested condition. In situ hybridization studies with matrilysin cRNA confirmed the presence of specific mRNA in both human promonocytes and monocytes. Moreover, a marked increase in hybridizable mRNA was observed with lipopolysaccharide treatment suggesting that matrilysin synthesis is pretranslationally regulated. In summary, this represents the first report documenting constitutive and regulated synthesis of matrilysin by a normal human cell type and suggests that matrilysin is expressed as a significant secreted product of mononuclear phagocytes at an intermediate stage of cellular differentiation.
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PMID:The matrix metalloprotease matrilysin (PUMP) is expressed in developing human mononuclear phagocytes. 137 84

We investigated the effects of a number of stimulatory agents on the production of both cell-associated and extracellular elastase-type enzymes on human monocyte-macrophages in vitro and of the modulation of such effects by modification of cellular cholesterol content. The stimulatory agents included phorbol myristate acetate (PMA) and the inflammatory mediators, lipopolysaccharide (LPS), opsonized zymosan (OZ), and platelet activating factor (PAF). Using the synthetic substrate, N-succinyl-trialanyl-paranitroanilide (SANA), we detected cell-associated elastase-like activity in monocyte-derived macrophages. Such activity increased markedly with cell maturation over the period from 5 to 15 days of adherence culture. While PAF (10 micrograms/ml) and LPS (10 micrograms/ml) were without effect on cell-associated elastase-like activity in macrophages, PMA (100 ng/ml) and OZ (1 mg/ml) markedly stimulated such activity in cells cultured for 15 days. Furthermore, a fivefold increase in the cell-associated elastase-like activity of macrophages occurred upon cholesterol loading of the cells with acetylated low density lipoprotein (AcLDL). By contrast, this activity was markedly diminished upon depletion of cellular cholesterol content after incubation with high density lipoprotein (HDL3). Latent elastinolytic activity in the culture medium was detected by use of a radioactive substrate, insoluble 3H-elastin, after initial tryptic treatment of the medium. Such latent elastase activity was secreted only by activated macrophages; the relative potency of stimulation was: PMA greater than LPS = PAF greater than OZ. Increase in cellular cholesterol content alone markedly enhanced the secretion of elastase (from undetectable levels to 28 ng of 3H-elastin degraded/hr/micrograms DNA). In all cases, both the cell-associated and secreted latent elastinolytic activities were due to metalloproteases, in view of their 90% inhibition by 2 mM EDTA. Cholesterol-loaded macrophages, which displayed an approximately 40-fold increase in total cholesterol content as compared to control cells, remained sensitive to the action of activators of OZ and PMA, while LPS and PAF exerted only weak effects. Our data indicate that cellular cholesterol content and inflammatory mediators are effective stimulants of the production and secretion of elastase-type enzymes by human monocyte-macrophages. Among these factors, cellular cholesterol content, OZ, PAF, and LPS may represent factors of relevance to the inflammatory role of the macrophage in atherogenesis and more specifically to the alteration of elastin structure in the extracellular matrix of the vessel wall.
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PMID:Expression of elastase activity by human monocyte-macrophages is modulated by cellular cholesterol content, inflammatory mediators, and phorbol myristate acetate. 231 58

Matrix metalloproteinases (MMPs) and elastase are proteolytic enzymes specifically directed against extracellular matrix (ECM) components. They are secreted by inflammatory cells and may consequently contribute to the lesions of the ECM observed during acute pulmonary edema. We therefore evaluated the MMP and elastase activities, which are secreted by cultured alveolar macrophages (AMACs) and polymorphonuclear neutrophils (PMNs) and present in the bronchoalveolar lavage (BAL) fluid in a guinea pig model of acute lung injury induced by intratracheal instillation of lipopolysaccharide (LPS). The control group was given 0.9% NaCl. 24 h after instillation, a BAL was performed, the BAL fluid was separated from the cells by centrifugation, and AMACs and PMNs were separately cultured for 24 h. In BAL fluid from LPS-treated guinea pigs, we found 1) an increase in free gelatinase activity, tested on [3H]gelatin (0.7 +/- 0.2 micrograms.200 microliters BAL fluid-1.48 h-1 vs. 0.2 +/- 0.1 in controls, P < 0.05), and 2) increased total gelatinase activities, as assessed by zymography. The molecular masses of the major gelatinase species found in BAL fluid by zymography were 92 and 68 kDa. The 92-kDa gelatinase was secreted by both AMACs and PMNs, as demonstrated by zymography of their respective culture media. When tested on [3H]elastin, the elastase activity of BAL fluid of LPS-treated animals exhibited no increase, but when tested on a synthetic peptidic substrate [N-succinyl-(L-alanine)3-p-nitro anilide (SLAPN)], increased elastase-like activity was observed (from 17 +/- 4 nmol of SLAPN.200 microliters BAL fluid-1.24 h-1 in control group to 34 +/- 8 in LPS group, P < 0.05). This increase was attributable to the activity of a metalloendopeptidase that was inhibited by the metal chelator EDTA but not by the specific tissue inhibitor of MMPs.
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PMID:Matrix metalloproteinase and elastase activities in LPS-induced acute lung injury in guinea pigs. 816 90

The horse leucocyte elastase inhibitor (HLEI), present in neutrophils, monocytes and bone marrow cells, is apparently a cytoplasmic protein which is not released from cells even in response to stimulation with lipopolysaccharide, phorbol ester, tumour necrosis factor alpha, interleukin-1 or elastin degradation products. Although no expression of the inhibitor was detected in neutrophils, both monocytes and bone marrow cells were efficient in its synthesis. Using a new expression vector pREST5d, recombinant inhibitor was produced in a large quantity in a soluble form, with a yield of 88 mg per 10 litres of E. coli culture. A two-step purification procedure, consisting of ion-exchange chromatography and gel filtration, yielded 36 mg of the recombinant inhibitor of a purity higher than 95%, as judged by SDS/PAGE. The recombinant protein had physicochemical and kinetic properties indistinguishable from those of the natural one, including irreversible elastase inhibition with an association rate constant kass > 10(7) M-1s-1. Both proteins were eliminated from rat circulation at the same ratio, and within the first 20 min 70% of the protein was removed. Such a short half-life in the circulation suggests that local delivery of HLEI directly to lungs in the form of aerosol could be a more efficient therapeutic approach than its intravenous injection.
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PMID:Biosynthesis and distribution of leucocyte elastase inhibitor. Production of recombinant inhibitor. 892 32

The regulation of the expression of mouse macrophage elastase (MME) was investigated using the murine tumor cell line P388D1. The effects of three factors were studied: a phorbol ester (4beta-phorbol 12-myristate 13-acetate, PMA), an endotoxin (lipopolysaccharide, LPS) and a corticosteroid (dexamethasone). Both in situ hybridization and northern blot analysis showed that P388D1 cells constitutively express the MME gene. Quantification of the MME mRNA by northern blot analysis showed that only PMA and dexamethasone significantly regulate MME gene expression in a time-dependent and dose-dependent manner. After PMA treatment, the MME mRNA level was maximal between 4 h and 9 h (medium-term response), and the mean amplitude of the response to a concentration of 100 nM was 2.5-fold (P<0.01). LPS did not induce any significant change in MME mRNA level even when 1% serum was added to the cultures. Following dexamethasone treatment, the MME mRNA level was minimal between 21 h and 33 h (long-term response), and the mean amplitude of the response to a concentration of 100 nM was 0.49-fold (P < 0.05). Using actinomycin D, it appeared that the inhibition of RNA synthesis reduces the ulterior stimulating effect of PMA from 184% to 121%, and that MME mRNA has a half-life longer than 8 h, which is not diminished by dexamethasone. These results strongly suggest that the two factors modify MME mRNA level by stimulating (PMA) or inhibiting (dexamethasone) the transcription of the gene, rather than by modifying the transcript stability. Analysis of the cell-conditioned media by elastin zymography showed the MME as a lysis band in the 22-kDa region, the intensity of which varied with the treatments. The MME secretion is stimulated by PMA, inhibited by dexamethasone and does not show any variation after LPS treatment.
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PMID:Metalloelastase expression in a mouse macrophage cell line--regulation by 4beta-phorbol 12-myristate 13-acetate, lipopolysaccharide and dexamethasone. 926 1

FR134043, disodium(Z,1S,15S,8S,24S,27R,29S,34S,37R)-29-ben zyl-21-ethylidene-27-hydroxy-15-isobutyrylamino-34-isopropyl-31,37 -dimethyl-10,16,19,22,30,32,35,38-octaoxo-36-oxa-9,11,17,20,23,28, 31,33-octaazatetracyclo[16.13.6.1(24),(28).0(3),(8)]octatricont a-3,5,7-trien-5,6-diyl disulfate, is a water-soluble inhibitor of human neutrophil elastase with a molecular mass of 1166.15 Da. FR134043 demonstrated a characteristic competitive inhibition of human neutrophil elastase with a Ki of 8 nM. In studies using synthetic substrates, FR134043 inhibited both neutrophil elastase activity and porcine pancreatic elastase activity with IC50 values of 35 nM and 49 nM respectively. FR134043 also inhibited hydrolysis of bovine neck ligament elastin by human neutrophil elastase with an IC50 value of 210 nM. In in vivo experiments, FR134043 protected animals against human neutrophil elastase (50 microg/animal)-induced lung hemorrhage in hamsters with an ED50 value of 3.1 microg/animal for intratracheal administration and 5.0 mg/kg for intravenous administration. Subcutaneous treatment with FR134043 significantly suppressed human neutrophil elastase (20 microg/paw)-induced paw edema in mice with an ED50 value of 3.3 mg/kg when evaluated 4 h after elastase injection. The potency of FR134043 given intratracheally to protect against porcine pancreatic elastase (100 microg/animal)-induced emphysema in hamsters was relatively low (Quasi-static lung compliance; ED50 = 1590 microg/animal) compared to that in acute animal models. FR134043 (10 mg/kg per h i.v. infusion) significantly improved lipopolysaccharide (0.25 mg/kg per h i.v. infusion)-induced thrombocytopenia and some coagulation parameters in rats. These results suggest that systemic administration of FR134043 would be advantageous over intratracheal administration of FR134043 for the treatment of adult respiratory distress syndrome, septic shock and pulmonary emphysema and other pathophysiologic conditions in which elastases are thought to be involved.
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PMID:Biochemical and pharmacological characterization of FR134043, a novel elastase inhibitor. 959 30

Evidence presented in the accompanying article (Gibbs, D. F., T. P. Shanley, R. L. Warner, H. S. Murphy, J. Varani, and K. J. Johnson. 1999. Role of matrix metalloproteinases in models of macrophage-dependent acute lung injury: evidence for alveolar macrophage as source of proteinases. Am. J. Respir. Cell Mol. Biol. 20:1145-1154) implicates alveolar macrophage matrix metalloproteinases (MMPs) in two models of acute lung inflammation in the rat. As a prerequisite to understanding which specific MMPs might be involved in the injury and how they might function, it was necessary to know the spectrum of enzymes present. To this end, alveolar macrophages were obtained from normal rat lungs by bronchoalveolar lavage, placed in culture with and without various agonists, and assessed by a variety of techniques for MMPs. The identification process involved characterization by gelatin, beta-casein, and kappa-elastin zymography, with confirmation of identity by Western blot/immunoprecipitation. Message levels of detected MMPs were assessed by Northern blot. Rat alveolar macrophages were found to produce a low constitutive level of MMP-2 (72-kD gelatinase A) that was only modestly upregulated following stimulation with phorbol myristate acetate, bacterial lipopolysaccharide, or immunoglobulin A-containing immune complexes. Although control cells were found to produce little or no MMP-9 (92-kD gelatinase B) or MMP-12 (metalloelastase), both enzymes were markedly upregulated upon stimulation. In the same stimulated macrophages there was little activity against type I collagen (associated with MMP-13 [collagenase-3] on the basis of Western blotting), no activity suggestive of stromelysin or matrilysin, and no measurable secretion of the serine proteinases, elastase and cathepsin G. These data demonstrate the ability of rat alveolar macrophages to elaborate certain MMPs under proinflammatory conditions, consistent with their possible involvement in the progression of acute inflammation.
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PMID:Characterization of matrix metalloproteinases produced by rat alveolar macrophages. 1034 Sep 32

Destruction of lung elastin is critical for development of emphysema associated with chronic obstructive pulmonary disease (COPD). Lung macrophages release elastolytic enzymes, including matrix metalloproteinase (MMP)-9, along with tissue inhibitors of MMP (TIMP). We examined the production and activity of macrophage-derived MMP-9 and TIMP-1 from alveolar macrophages (AM) from smokers with COPD, healthy smokers (HS), and nonsmokers (NS). AM were stimulated with either lipopolysaccharide (LPS), interleukin (IL)-1 beta, or cigarette smoke-conditioned culture medium (CSM). AM from patients with COPD released greater amounts of MMP-9 with greater enzymatic activity than HS and NS. In contrast, AM from NS released more TIMP-1 than cells from HS and subjects with COPD. LPS and IL-1 beta caused a dose-dependent increase in MMP-9 release and activity, together with increased levels of TIMP-1. Dexamethasone prevented the increase in MMP-9 release, and increased TIMP-1 release. CSM increased MMP-9 and TIMP-1 release from AM of all groups. Dexamethasone decreased CSM-stimulated MMP-9 release, but had no effect on MMP-9 activity This study suggests that macrophages might be important in the development of COPD because these cells exhibit increased levels of elastolytic activity.
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PMID:Release and activity of matrix metalloproteinase-9 and tissue inhibitor of metalloproteinase-1 by alveolar macrophages from patients with chronic obstructive pulmonary disease. 1197 Sep 13

Secretion of proteases is critical for degradation of the extracellular matrix during an inflammatory response. Cathepsin (Cat) S and L are the major elastinolytic cysteine proteases in mouse macrophages. A 65 amino acid segment of the p41 splice variant (p41(65aa)) of major histocompatibility complex class II-associated invariant chain (Ii) binds to the active site of CatL and permits the maintenance of a pool of mature enzyme in endosomal compartments of macro-phages and dendritic cells (DCs). Here we show that interaction of p41(65aa) with mature CatL allows extracellular accumulation of the active enzyme. We detected mature CatL as a complex with p41(65aa) in culture supernatants from antigen-presenting cells (APCs). Extracellular accumulation of mature CatL is up-regulated by inflammatory stimuli as observed in interferon (IFN)-gamma-treated macrophages and lipopolysaccharide (LPS)-activated DCs. Despite the neutral pH of the extracellular milieu, released CatL associated with p41(65aa) is catalytically active as demonstrated by active site labeling and elastin degradation assays. We propose that p41(65aa) stabilizes CatL in the extracellular environment and induces a local increase in the concentration of matrix-degrading enzymes during inflammation. Through its interaction with CatL, Ii may therefore control the migratory response of APCs and/or the recruitment of effectors of the inflammatory response.
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PMID:Invariant chain controls the activity of extracellular cathepsin L. 1241 35

Arg-gingipain (Rgp) and Lys-gingipain (Kgp) are Porphyromonas gingivalis cysteine proteinases implicated as major virulence factors in pathologies of periodontitis. We purified a 660-kDa cell-associated gingipain complex existing as a homodimer of two catalytically active monomers which comprises their catalytic and adhesin domains. Electron microscopy revealed that the complex was composed of a globular particle with a 10-nm external diameter possessing one or two electron-dense hole-like structures. Two-dimensional gel electrophoresis and immunoblot analyses revealed the association of lipopolysaccharide (LPS) with the catalytic domains and a hemagglutinin domain, Hgp44, of Rgp and Kgp in the complex. The complex significantly degraded human type I collagen and elastin and strongly disrupted viability of human gingival fibroblasts and umbilical vein endotherial cells with an efficiency which was higher than that of the monomeric gingipains. The native complex produced only a small amount of nitrogen dioxide, tumor necrosis factor alpha, and interleukin-6 by macrophages, whereas the heat-denatured complex resulted in increased production. Inhibition of the proteolytic activities of the gingipain complex did not up-regulate the cytokine production, indicating that the functional domains in LPS are structurally masked by the complex proteins. These results indicate the importance of the complex in evasion of host defense mechanisms as well as in host tissue breakdown.
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PMID:A functional virulence complex composed of gingipains, adhesins, and lipopolysaccharide shows high affinity to host cells and matrix proteins and escapes recognition by host immune systems. 1566 30


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