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:3.4.21.7 (plasmin)
9,023 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The capacity of human monocytoid cell lines and peripheral blood monocytes to modulate their expression of plasminogen receptors has been assessed. After PMA stimulation, THP-1 or U937 monocytoid cells were separated into adherent and nonadherent populations. Plasminogen bound to adherent cells with similar capacity and affinity as to nonstimulated cells. In contrast, the nonadherent cells bound plasminogen with 5-17-fold higher capacity (without a change in affinity). This increase was selective as urokinase bound with similar affinity and capacity to the adherent and nonadherent populations. Upregulation of plasminogen receptors on the nonadherent monocytoid cells was rapid, detectable within 30 min, and reversible, adhesion of the nonadherent cells resulted in a sixfold decrease in plasminogen binding within 90 min. The increase in plasminogen binding to the nonadherent cells was associated with a marked increase in their capacity to generate plasmin activity from cell-bound plasminogen. PMA stimulation of human peripheral blood monocytes increased their expression of plasminogen receptors by two- to fourfold. This increase was observed in both adherent and nonadherent monocytes. Freshly isolated monocytes maximally bound 5.0 x 10(5) plasminogen molecules per cell, whereas monocytes cultured for 18 h or more maximally bound 1.7 x 10(7) molecules per cell, a 30-fold difference in receptor number. These results indicate that both monocytes and monocytoid cell lines can rapidly and markedly regulate their expression of plasminogen binding sites. As enhanced plasminogen binding is correlated with an increased capacity to generate plasmin, an enzyme with broad substrate recognition, modulation of plasminogen receptors may have profound functional consequences.
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PMID:Regulation of plasminogen receptor expression on human monocytes and monocytoid cell lines. 217 Apr 26

Mononuclear phagocytes regulate the generation of plasmin by secreting urokinase-type plasminogen activator (uPA) and plasminogen activator inhibitor-2 (PAI-2). We investigated the production of plasminogen activator (PA) and PA inhibitor by the human monocytic leukemia cell line, THP-1. Similar to U937 monoblast-like cells and peripheral blood monocytes (PBM), THP-1 cells produce a PA that is specifically neutralized by anti-uPA antibody and comigrates with human high molecular mass uPA (54 kDa) on casein-plasminogen zymogaphy. PA activity could be dissociated from intact THP-1 cells by brief treatment with a weak acid-glycine buffer, indicating that the uPA is secreted and bound to receptors on the plasma membrane. Regulation of uPA proceeds normally in THP-1 cells, with cell-associated PA activity increasing from 77 +/- 20 to 163 +/- 26 and 325 +/- 30 mPU/10(6) cells in response to PMA and LPS, respectively; parallel increases in steady state levels of uPA mRNA were observed. In contrast to normal expression of uPA activity, functional PAI-2 could not be demonstrated in either the conditioned media or cell lysates of THP-1 under basal or stimulated conditions. Both U937 and PBM secrete low levels of PA inhibitor activity that increase substantially in response to stimulation with PMA and LPS. Immunoreactive PAI-2, measured by ELISA, was undetectable in THP-1 lysates or conditioned medium, but was consistently present in U937 and PBM, paralleling the presence of PA inhibitor activity. THP-1 cells express low levels of an abnormally sized mRNA for PAI-2 and demonstrate a regulatory defect whereby steady state levels of PAI-2 mRNA are markedly reduced upon stimulation with PMA or LPS. By contrast, U937 and PBM respond to identical stimulation with increases in PAI-2 mRNA. We conclude that THP-1 cells express a structurally abnormal species of PAI-2 mRNA, with complete loss of inhibitory activity as well as altered function of PMA- and LPS-responsive regulatory elements.
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PMID:The THP-1 cell line is a urokinase-secreting mononuclear phagocyte with a novel defect in the production of plasminogen activator inhibitor-2. 230 45

The production of interleukin (IL 1) by normal human peripheral blood monocytes purified by Ficoll-Hypaque density sedimentation, Percoll-gradient sedimentation, and plastic adherence can be detected as early as 30 min intracellularly, and extracellularly within 1 hr after stimulation with lipopolysaccharide (LPS). Production of mRNA coding for the isoelectric point 7.0 species of IL 1 was also detected as early as 1 hr after LPS stimulation and reached a maximum level at 6 hr. Cell-associated IL 1 activity could be extracted with CHAPS detergent from every cell fraction (i.e., membranes, cytosol, and particulates), but was present mainly (greater than 95%) in the cytosol of LPS-activated monocytes and the myelomonocytic cell line, THP-1. The apparent m.w. of IL 1 activity on high pressure liquid chromatography gel filtration in every cell fraction was approximately 23,000 daltons, with a minor peak at 31,000 daltons, whereas the IL 1 activity in the culture supernatants was 17,000 daltons. Western blotting analysis of LPS-stimulated monocyte extracts showed two forms of IL 1 corresponding to 31,000 daltons and 25,000 daltons. Exposure of viable cells to trypsin and plasmin released biologically active 23,000 dalton IL 1 only from IL 1-producing cells such as activated monocytes and IL 1-producing Ebstein-Barr virus B lymphocyte cell lines. Consequently, biologically active IL 1 is presumably exposed on the outer surface of cell membranes. Furthermore, IL 1 release by human monocytes in plasminogen-depleted fetal calf serum was considerably decreased. Conversely, supplementation of plasminogen-depleted serum with purified plasminogen restored the IL 1 production, suggesting that plasmin or plasmin-like factors may be involved in the regulation of the release of IL 1 from IL 1-producing cells. In conclusion, the results suggest that IL 1 is rapidly produced, is pooled in the cytosol, and in part is processed by enzymes, is transferred to the plasma membranes, and is then released from the cells. Tissue plasminogen activator and serum enzymes such as plasmin may therefore be involved in the release of IL 1 from IL 1-producing cells.
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PMID:Intracellular localization of human monocyte associated interleukin 1 (IL 1) activity and release of biologically active IL 1 from monocytes by trypsin and plasmin. 242 Aug 74

The activation of plasminogen by macrophage is regulated by their expression of receptors for urokinase and plasmin(ogen). In these studies we have examined plasmin(ogen) binding to adherent human THP-1 macrophage. Plasmin bound to the THP-1 cells in a time- and dose-dependent manner (Kd 15.8 +/- 6.2 nM; Bmax 1.4 +/- 0.3 x 10(6)/cell). The lysine analog epsilon-aminocaproic acid competitively inhibited plasmin binding. The fraction of membrane-bound plasmin, however, became increasingly resistant to displacement with epsilon-aminocaproic acid. Over a 24-h period, membrane-bound plasmin activity fell 80% despite the presence of catalytically active plasmin in the incubation media. The loss of receptor-bound plasmin activity was not due to proteolytic alterations of its receptor since 125I-Lys-plasminogen bound to THP-1 cells pretreated with plasmin with similar affinity as to untreated cells. Following a 24-h incubation of 125I-Lys-plasminogen or 125I-plasmin with THP-1 cells, several degradative fragments were apparent in their conditioned media. The smaller degradative fragments (28 and 36 kDa) lacked cell binding activity and were demonstrated to be active by casein-zymography. A 48-kDa fragment bound to cells in a lysine-dependent manner but was not active. In contrast, phenylmethylsulfonyl fluoride-inactivated 125I-plasmin retained its binding activity over 24 h, and degradative fragments were not present in the conditioned media. The binding of 125I-Lys-plasmin(ogen) to THP-1 cells was also examined in the presence of excess alpha 2 plasmin inhibitor. Despite the absence of fluid-phase plasmin activity, membrane-bound 125I-Lys-plasmin(ogen) decreased over 24 h. At 24 h a radiolabeled 48-kDa fragment was observed in the conditioned media and together with 125I-Lys-plasmin(ogen) was bound to cells. Unlike 125I-Lys-plasmin, the 48-kDa fragment did not form a complex with alpha 2 plasmin inhibitor. Thus, autoproteolysis of receptor-bound plasmin results in fragments with truncated physiologic properties that possess either cell binding or catalytic activities. We propose that autoproteolysis is a mechanism for regulating membrane-bound plasmin activity.
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PMID:Regulation of macrophage receptor-bound plasmin by autoproteolysis. 752 19

Receptors for urokinase (uPA) and plasminogen provide a mechanism to direct the cellular activation of plasminogen. The regulation of these receptors is important for several macrophage functions. In these studies, the effect of transforming growth factor-beta 1 (TGF-beta 1) on uPA, uPA receptor, and plasminogen receptor expression by human THP-1 macrophage was examined. TGF-beta 1 induction of uPA expression by THP-1 cells was differentiation dependent. Suspension and adherent cultures expressed similar constitutive levels of uPA. Exposure of adherent cells to TGF-beta 1 led to a dose- and time-dependent increase in uPA activity which was paralleled by an increase in uPA antigen and uPA mRNA. In contrast, uPA expression by suspension cultures was unresponsive to TGF-beta 1. The differential response exhibited by suspension and adherent THP-1 cells may reflect differences in their expression of TGF-beta 1 receptors, since when assayed by crosslinking techniques, suspension cells primarily expressed a 65 kDa receptor; whereas, the adherent cells expressed 65 and 100 kDa receptors. TGF-beta 1-induced alterations in uPA receptor expression by adherent THP-1 cells were examined by quantitating membrane-bound uPA activity. Membrane-bound uPA activity increased three-fold when cells were incubated with TGF-beta 1. The increase in membrane-uPA activity expressed by TGF-beta 1-treated cells was not due to increased uPA receptor occupancy since incubation of either control or TGF-beta 1 primed cells with exogenous uPA did not lead to an increase in membrane-bound uPA activity. Furthermore, immunoreactive uPA receptor was increased in TGF-beta 1-treated cells. Following incubation with plasminogen, membrane-bound plasmin activity increased three-fold in TGF-beta 1-treated cells. However, no change in immunoreactive membrane-bound plasmin(ogen) was observed. In addition, binding of 125I-Lys-plasminogen to THP-1 cells was not affected by TGF-beta 1 treatment. We conclude that TGF-beta 1 stimulates membrane-bound plasmin activity, without affecting plasminogen receptor expression, through the up-regulation of uPA and the uPA receptor expression.
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PMID:THP-1 macrophage membrane-bound plasmin activity is up-regulated by transforming growth factor-beta 1 via increased expression of urokinase and the urokinase receptor. 762 80

Fibrin(ogen) (FGN) is important for hemostasis and wound healing and is cleared from sites of injury primarily by the plasminogen activator system. However, there is emerging evidence in plasminogen activator-deficient transgenic mice that nonplasmin pathways may be important in fibrin(ogen)olysis, as well. Given the proximity of FGN and monocytes within the occlusive thrombus at sites of vascular injury, we considered the possibility that monocytes may play an ancillary role in the degradation and clearance of fibrin. We found that monocytes possess an alternative fibrinolytic pathway that uses the integrin Mac-1, which directly binds and internalizes FGN, resulting in its lysosomal degradation. At 4 degrees C, FGN binds to U937 monocytoid cells in a specific and saturable manner with a kd of 1.8 mumol/L. Binding requires adenosine diphosphate stimulation and is calcium-dependent. At 37 degrees C, FGN and fibrin monomer (FM) are internalized and degraded at rates of 0.37 +/- 0.13 and 0.55 +/- 0.03 microgram/10(6) cells/h by U937 cells, 1.38 +/- 0.02 and 1.20 +/- 0.30 microgram/10(6) cells/h by THP-1 cells, and 2.10 +/- 0.20 and 2.52 +/- 0.18 micrograms/10(6) cells/h by human peripheral blood mononuclear cells, respectively. The serine protease inhibitors, PPACK and aprotinin, and the specific elastase inhibitor, AAPVCK, do not significantly inhibit degradation. However, degradation is inhibited by chloroquine, suggesting that a lysosomal pathway is involved. Factor X, a competitive ligand with FGN for the Mac-1 receptor, also blocks degradation, as does a monoclonal antibody to the alpha-subunit of Mac-1. Autoradiography of radioiodinated, internalized FGN shows that FGN proteolysis by the pathway produces a unique degradation pattern distinct from that observed with plasmin. In a fibrin clot lysis assay, Mac-1-mediated fibrinolysis contributed significantly to total fibrinolysis. In summary, FGN is internalized and degraded by activated human monocytoid cells via Mac-1 in the absence of plasmin, thereby providing an alternative fibrinolytic pathway. Thus, in addition to the function of cell adhesion, integrins may also act as receptors that mediate the internalization and degradation of bound ligands.
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PMID:Fibrin(ogen) is internalized and degraded by activated human monocytoid cells via Mac-1 (CD11b/CD18): a nonplasmin fibrinolytic pathway. 840 Feb 91

This study examined the effects of endogenous urokinase (uPA) on lipopolysaccharide (LPS)-stimulated tumor necrosis factor alpha (TNF-alpha) secretion in THP-1 mononuclear phagocytes. Anti-uPA monoclonal antibody (mAb) suppressed LPS-driven TNF-alpha secretion by 61.6 +/- 5.9% (P<.001), and PAI-1, a uPA inhibitor, suppressed it to 53.1 +/- 8.2% of the control value (P<.001). Up-regulation of TNF-alpha mRNA was suppressed in parallel with secreted TNF-alpha protein. TNF-alpha secretion was unaffected by depleting plasminogen or by aprotinin, a plasmin inhibitor. When endogenous uPA was displaced from the cell, exogenous high-molecular-weight (intact) uPA augmented LPS-driven TNF-alpha secretion. By contrast, a uPA fragment containing the catalytic domain was inhibitory, and the uPA receptor-binding domain had no effect. We conclude that endogenous uPA amplifies TNF-alpha neosynthesis of LPS-stimulated THP-1 mononuclear phagocytes. The effect requires intact uPA and is independent of plasmin activity. This represents a novel mechanism by which a mononuclear phagocyte-derived protease contributes to generating proinflammatory signals.
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PMID:Endogenously produced urokinase amplifies tumor necrosis factor-alpha secretion by THP-1 mononuclear phagocytes. 860 4

Plasminogen binding sites function to arm cell surfaces with the proteolytic activity of plasmin, critical for degradation of extracellular matrices. We have assessed the effects of adhesion of the representative monocytoid cell lines, THP-1 and U937, to purified extracellular matrix proteins on their expression of plasminogen receptors. After adhesion to immobilized fibronectin, adherent and nonadherent subpopulations of cells were separated. Plasminogen binding to the nonadherent population of cells increased 3-fold, whereas binding to the adherent population decreased by 60%. These changes were due to differences in the plasminogen binding capacities of the cells, while the affinities of the cells for plasminogen were unchanged. The up-regulation of receptor expression in the nonadherent cell population was: 1) induced rapidly and reversibly, 2) independent of new protein synthesis, 3) required an interaction between adherent and nonadherent cell populations, and 4) associated with an enhanced ability of the cells to promote plasminogen activation and to degrade fibronectin. Other immobilized adhesive proteins, laminin and vitronectin, also supported up-regulation of plasminogen receptors in the nonadherent cells. Carboxypeptidase B treatment eliminated the increment in the plasminogen binding capacity of the nonadherent cells, suggesting that the increase in binding was due to exposure of new carboxyl-terminal lysyl residues on the cell surfaces. Furthermore, both the adherence of the cells and up-regulation of plasminogen binding sites was abolished by beta1-integrin monoclonal antibodies. These results suggest that proteins found in extracellular matrices have the capacity to modulate the expression of plasminogen binding sites, thus regulating local proteolysis and cell migration.
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PMID:Regulation of plasminogen receptor expression on monocytoid cells by beta1-integrin-dependent cellular adherence to extracellular matrix proteins. 879 2

Plasminogen and tPA bind to a common set of binding sites on nucleated cells. To assess the functional consequences of cellular binding, we have measured the kinetic changes induced by plasminogen activation by tPA on cell surfaces. These studies were carried out with U937 and THP-1 monocytoid cells, with Raji, Nalm6 and Molt4 lymphoid cells and with peripheral blood monocytes and neutrophils. The interactions of plasminogen and tPA with cells induced an increase in the rate of plasmin generation which depended upon the cell concentration. With saturating amounts of U937 monocytoid cells (1.25 x 10(5)/ml) the rate of plasmin generation was 0.39 nM.s-1 versus 0.07 and 0.09 nM.s-1 without cells or without tPA, respectively. The catalytic efficiency of Glu- or Lys-plasminogen activation by tPA increased by 7.2- and 24.2-fold, respectively. These changes were induced by a 72-242-fold reduction in the Km of these interactions which was in the range of 0.3-0.9 microM. These values are below the plasminogen concentration in plasma (1-2 microM). Moreover, we provide new data indicating that 1) only a specific subset of plasminogen binding sites, i.e. molecules exposing carboxyl terminal lysines on the cell surface, promotes plasminogen activation on cells; 2) the first four kringles of plasminogen and the finger of tPA are critical for enhanced plasmin generation on cell surfaces; 3) the simultaneous co-localization of tPA with plasminogen on cell surfaces is required for enhanced plasminogen activation; 4) modulation of plasminogen/tPA receptor expression induces concomitant modulation of the stimulatory effects of cells on plasminogen activation and 5) in a direct comparison, the mechanism by which cells and fibrin fragments accelerate plasminogen activation are similar but not identical. These data suggest that modulation of plasminogen/tPA binding sites permits local and efficient generation of plasmin on cell surfaces.
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PMID:Characterization of cellular binding sites and interactive regions within reactants required for enhancement of plasminogen activation by tPA on the surface of leukocytic cells. 890 99

Ceruloplasmin is a 132-kDa glycoprotein abundant in human plasma. It has multiple in vitro activities, including copper transport, lipid pro- and antioxidant activity, and oxidation of ferrous ion and aromatic amines; however, its physiologic role is uncertain. Although ceruloplasmin is synthesized primarily by the liver in adult humans, production by cells of monocytic origin has been reported. We here show that IFN-gamma is a potent inducer of ceruloplasmin synthesis by monocytic cells. Activation of human monoblastic leukemia U937 cells with IFN-gamma increased the production of ceruloplasmin by at least 20-fold. The identity of the protein was confirmed by plasmin fingerprinting. IFN-gamma also increased ceruloplasmin mRNA. Induction followed a 2- to 4-h lag and was partially blocked by cycloheximide, indicating a requirement for newly synthesized factors. Ceruloplasmin induction in monocytic cells was agonist specific, as IL-1, IL-4, IL-6, IFN-alpha, IFN-beta, TNF-alpha, and LPS were completely ineffective. The induction was also cell type specific, as IFN-gamma did not induce ceruloplasmin synthesis in endothelial or smooth muscle cells. In contrast, IFN-gamma was stimulatory in other monocytic cells, including THP-1 cells and human peripheral blood monocytes, and also in HepG2 cells. Ceruloplasmin secreted by IFN-gamma-stimulated U937 cells had ferroxidase activity and was, in fact, the only secreted protein with this activity. Monocytic cell-derived ceruloplasmin may contribute to defense responses via its ferroxidase activity, which may drive iron homeostasis in a direction unfavorable to invasive organisms.
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PMID:Induction of ceruloplasmin synthesis by IFN-gamma in human monocytic cells. 925 59


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