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:3.4.21.4 (trypsin)
42,187 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

GMP-140 (P-selectin), a 140-kDa granular membrane glycoprotein localized to the alpha granules of platelets and the Weibel-Palade bodies of endothelial cells, is thought to play an important role in adhesive interactions predominantly between granulocytes, platelets and vascular endothelial cells during inflammation. Although GMP-140 binds to granulocytes, its binding to lymphocytes has not been demonstrated. Using genetically engineered IgG C gamma 1 fusion protein of the extracellular domains of GMP-140, we demonstrate that GMP-140 binds to chronically antigen (Ag)-stimulated CD4+ T cells. Freshly isolated CD4+ T cells did not bind GMP-140, but priming and subsequent stimulation with alloantigen induced and gradually increased expression of GMP-140-reactive structures on their surface. T cells isolated from rheumatoid synovial fluids also exhibited strong binding to GMP-140. The binding of GMP-140 to primed T cells is not influenced by preactivation with phorbol 12-myristate 13-acetate, is almost completely abolished by pretreatment of T cells with neuraminidase or trypsin, and is also strongly inhibited by EDTA, the soluble sulfated glycans dextran sulfate, fucoidan, and heparin, but not by chondroitin sulfates. In spite of its strong binding to Ag-primed T cells, GMP-140 did not modulate the proliferative responses of these cells to various stimuli. However, GMP-140 in conjunction with anti-T cell receptor alpha beta monoclonal antibodies augmented the production of granulocyte-macrophage colony-stimulating factor GM-CSF and inhibited the production of interleukin-8 by Ag-primed T cells without influencing their tumor necrosis factor-alpha production. These results suggest that GMP-140 binds to chronically stimulated CD4+ T cells and differentially modulates their production of proinflammatory cytokines. The ability of Ag-primed T cells to bind GMP-140 may facilitate interactions with activated platelets and endothelial cells affecting the course of inflammation.
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PMID:GMP-140 (P-selectin/CD62) binds to chronically stimulated but not resting CD4+ T lymphocytes and regulates their production of proinflammatory cytokines. 137 17

L-Selectin is a lectin-like receptor on lymphocytes which mediates their attachment to high endothelial venules (HEV) within lymph nodes. Previous work has identified HEV-associated endothelial ligands for L-selectin as sialylated, fucosylated and sulphated glycoproteins of approximately 50 kDa and approximately 90 kDa (Sgp50 and Sgp90). The interaction of L-selectin with these ligands is carbohydrate directed, reflecting the involvement of its amino-terminal, calcium-type lectin domain. It has been reported, and we have confirmed, that anti-Ly22 blocks the adhesive function of L-selectin without reducing its binding to a carbohydrate- based ligand PPME (phosphomannan monoester core from Hansenula hostii). The epitope for this monoclonal antibody depends on the epidermal growth factor (EGF) domain of L-selectin. We demonstrate that anti-Ly22 inhibits the interaction of L-selectin with both of the Sgps, thus establishing that the interaction of L-selectin with HEV can be accounted for by the Sgps. Furthermore, the interaction of trypsin fragments of Sgp50 with L-selectin is inhibitable both by an antibody that maps to the lectin domain and by anti-Ly22. These findings raise the possibility that anti-Ly22 is affecting the function of the lectin domain of L-selectin rather than directly antagonizing the EGF domain. Toward a further characterization of L-selectin's carbohydrate specificity, we show that Sgp50 is partially inactivated by the linkage-specific Newcastle Disease virus sialidase (alpha 2,3 linkage). We additionally demonstrate that a sialyl Lewis x-related tetrasaccharide can interact with L-selectin, as has also been demonstrated for E-selectin and P-selectin.
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PMID:Further characterization of the interaction between L-selectin and its endothelial ligands. 138 20

Activated platelets and stimulated endothelial cells express P-selectin, an integral membrane protein receptor that binds monocytes and neutrophils. P-selectin mediates adhesion to glycoproteins with carbohydrate structures containing sialyl-Lewis X. Since many carcinoma cells also express these carbohydrate structures and are known to interact with platelets, we asked whether P-selectin may mediate this interaction. Both small cell lung cancer and neuroblastoma cell lines bound to activated platelets, and this interaction was blocked with inhibitory anti-P-selectin antibodies and by pretreatment of these cancer cells with neuraminidase or trypsin. Platelet binding to the small cell lung cancer cells was not inhibited with anti-GP IIb-IIIa antibody or Arg-Gly-Asp-Ser peptide. Pretreatment of the neuroblastoma cells with inhibitors of N-linked carbohydrate biosynthesis had little effect on binding to P-selectin, indicating that relevant carbohydrate ligand(s) may be O-linked. In addition, lipospheres containing P-selectin specifically bound to cryostat sections derived from a small cell lung tumor and two neuroblastoma tumors, but not to sections of normal lung. These observations demonstrate that P-selectin mediates binding of platelets to small cell lung cancer and to neuroblastoma and suggest a possible role for this lectin in metastasis.
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PMID:P-selectin mediates adhesion of platelets to neuroblastoma and small cell lung cancer. 768 63

The stimulated release of von Willebrand factor (vWF) from endothelial cells by secretagogues such as thrombin is associated with the translocation of Weibel-Palade bodies to the cell membrane and the surface expression of P-selectin (also known as GMP 140, PADGEM and CD 62). P-selectin, which is stored in Weibel-Palade bodies, is a neutrophil and monocyte adhesion molecule important in the initiation of inflammation. We have developed a simple assay for the detection of P-selectin on endothelial cells using indirect immunofluorescence and flow cytometry and have confirmed that this is temporally related to vWF release. The assay has been used to demonstrate that IL-1 does not cause Weibel-Palade body degranulation but that trypsin does. This has implications for the use of passaged endothelial cells in the study of vWF release and the assay has numerous possible applications in study of mechanisms of stimulated vWF release.
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PMID:von Willebrand factor release and P-selectin expression is stimulated by thrombin and trypsin but not IL-1 in cultured human endothelial cells. 769 90

Sequestration of Plasmodium falciparum infected erythrocytes in the cerebral circulation is strongly implicated in the pathogenesis of cerebral malaria. From previous studies it was postulated that genes essential for cytoadherence were located on the right arm of chromosome 9 as P. falciparum isolates with a deletion in this region lost the capacity to cytoadhere in vitro and no longer expressed Plasmodium falciparum erythrocyte membrane protein-1 (PfEMP-1) on the surface of the infected cells. We have selected a P. falciparum isolate from Papua New Guinea for high levels of cytoadherence to human umbilical vein endothelial cells (HUVECs) and have shown that the cloned parasite has several novel properties related to cytoadherence. The cloned parasite adheres to HUVECs, does not bind to melanoma cells, and expresses a surface molecule with most of the properties of PfEMP-1, despite a deletion in the right arm of chromosome 9. Interestingly, the surface expressed PfEMP-1 in this strain is resistant to trypsin treatment and infected cells continue to cytoadhere after trypsin digestion at a concentration of 100 micrograms ml-1. The receptor on HUVECs for the cloned parasite lines is a molecule different from any previously described, as parasitized cells do not adhere to soluble intercellular adhesion molecule 1, thrombospondin, vascular cell adhesion molecule 1, E-selectin or P-selectin, nor to CD36. Our work, taken together with the results from previous studies, suggest that the ability of parasites to cytoadhere is encoded in at least two distinct genomic locations in the parasite, and the diversity of receptor-ligand interaction is greater than previously described.
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PMID:A Plasmodium falciparum isolate with a chromosome 9 deletion expresses a trypsin-resistant cytoadherence molecule. 783 80

A mouse homolog of P-selectin glycoprotein ligand-1 (PSGL-1), a P-selectin receptor on myeloid cells, has been cloned using the human cDNA sequence to probe a cDNA library prepared from the mouse WEHI-3 monocytic cell line and a genomic DNA library prepared from 129/SvJ mouse tissue. The gene flanking the entire open reading frame of 397 amino acids is composed of a single exon. Mouse and human PSGL-1 show an overall similarity of 67% and an identity of 50% and contain a similar domain organization. However, there are 10 threonine/serine-rich decameric repeats in mouse PSGL-1 as compared with 15 threonine-rich repeats in human PSGL-1. When the mouse PSGL-1 cDNA is coexpressed with an alpha 1,3/1,4 fucosyltransferase cDNA in COS cells, a functional protein is expressed on the COS cell surface mediating binding to human P-selectin. The mouse PSGL-1 gene, Selpl, was mapped to a position on mouse chromosome 5 (Chr 5). Northern blot analyses of mouse tissues showed moderate expression of a PSGL-1 mRNA species in most tissues including heart, kidney, liver, muscle, ovary, and stomach and high levels of expression in blood, bone marrow, brain, adipose tissue, spleen, and thymus. Whereas certain mouse myeloid cell lines including PU5-1.8, WEHI-3B, and 32DC13 express high levels of PSGL-1 mRNA, only WEHI-3B and 32DC13 bind to P-selectin; this interaction is blocked by anti-PSGL-1 antibody. WEHI-3B cells bind significantly better to P-selectin than to E-selectin. Although comparable P-selectin binding is observed in 32DC13 cells, these cells bind better to E-selectin. Binding of 32DC13 cells to E-selectin is not blocked by anti-PSGL-1 antibody. Treatment of WEHI-3B cells with trypsin or neuraminidase abolished their ability to interact with P-selectin. These results indicate that mouse PSGL-1 has structural and functional homology to human PSGL-1 but is characterized by differences in the composition and number of the decameric repeats. PSGL-1 on mouse myeloid cells is critical for high-affinity binding to P-selectin but not E-selectin.
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PMID:Mouse P-selectin glycoprotein ligand-1: molecular cloning, chromosomal localization, and expression of a functional P-selectin receptor. 863 76

The human pancreatic tumor cell line SUIT-2 was derived from a metastatic lesion in the liver of a patient with pancreatic adenocarcinoma. SUIT-2 and clonal cell lines derived from it show spontaneous metastasis to lung and regional lymph nodes from s.c. nude mouse xenografts and were found to express P-selectin mRNA and protein. Surface expression of P-selectin protein was increased by exposure of the pancreatic tumor cells to thrombin, oxygen radicals, and trypsin, suggesting that common cellular mechanisms for regulating P-selectin surface expression exist among platelets, endothelial cells, and these pancreatic tumor cells. The finding that P-selectin is expressed by metastatic pancreatic tumor cells demonstrates that the range of cell types that express these adhesion molecules is broader than believed previously.
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PMID:P-selectin expression in a metastatic pancreatic tumor cell line (SUIT-2). 906 94

Plasmodium falciparum-infected erythrocytes (IRBC) roll on the adhesion molecule P-selectin in vitro under flow conditions that approximate the shear stress in capillary and postcapillary venules in which cytoadherence occurs in vivo. The pathological significance of this adhesive interaction is currently unknown. In this study, we further investigated the molecular interactions between IRBC and P-selectin by using a laminar flow system that allowed for the direct visualization of IRBC-substratum interactions. The results showed that the IRBC-P-selectin interaction was Ca2+-dependent and involved the lectin domain of P-selectin and a sialic acid residue on IRBC. The sialylated P-selectin ligand was trypsin-sensitive, which suggests that it could be part of the parasite antigen PfEMP1 that interacts with CD36 and intercellular adhesion molecule-1 (ICAM-1), but different from a trypsin-resistant IRBC ligand that adheres selectively to chondroitin sulfate A. Studies on the rolling and adhesion of IRBC on activated platelets that express both CD36 and P-selectin showed that inhibition of rolling on P-selectin reduced the adhesion of some clinical parasite isolates to CD36, whereas other parasite isolates appeared to interact directly with CD36. Thus, cytoadherence under physiological flow conditions may be mediated by multiple IRBC ligands that interact with different adhesion molecules in a cooperative fashion. These findings underscore the complexity of the interactions betweeen IRBC and vascular endothelium.
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PMID:Characterization of Plasmodium falciparum-infected erythrocyte and P-selectin interaction under flow conditions. 961 80

Short-term exposure to ozone at peak ambient levels induces neutrophil influx and impairs lung function in healthy humans. In order to investigate the mechanisms contributing to neutrophil recruitment and to examine the role of T-cells in the acute inflammatory response, we exposed 12 healthy humans to 0.2 parts per million (ppm) of ozone and filtered air on two separate occasions for 2 h with intermittent periods of rest and exercise (minute ventilation = 30 L x min(-1)). Fibreoptic bronchoscopy was performed 6 h after the end of exposures. Total protein, tryptase, histamine, myeloperoxidase, interleukin (IL)-8 and growth-related oncogene-alpha (Gro-alpha) were measured and total and differential cell counts were performed in bronchoalveolar lavage (BAL) fluid. Flow cytometry was performed on BAL cells to study total T-cells, T-cell receptors (alphabeta and gammadelta), T-cell subsets (CD4+ and CD8+ cells) and activated T-cell subsets (CD25+). Using immunohistochemistry, neutrophils, mast cells, total T-cell numbers, T-cell subsets, CD25+ T-cells and leukocyte endothelial adhesion molecules including P-selectin, E-selectin, intercellular adhesion molecule (ICAM)-1 and vascular adhesion molecule (VCAM)-1 were quantified in the bronchial biopsies. Paired samples were available from nine subjects. Following ozone exposure there was a threefold increase in the proportion of polymorphonuclear neutrophils (PMNs) (p=0.07) and epithelial cells (p=0.05) in BAL fluid. This was accompanied by increased concentrations of IL-8 (p=0.01), Gro-alpha (p=0.05) and total protein (p=0.058). A significant positive correlation was demonstrated between the two chemokines and proportion of PMNs in BAL fluid. After ozone exposure there was a significant decrease in the CD4/CD8 ratio (p=0.05) and the proportion of activated CD4+ (p=0.01) and CD8+ T-cells (p=0.04). However, no significant changes were demonstrable in any of the inflammatory markers studied in the biopsies. Short-term exposure of healthy humans to 0.2 ppm ozone induced a neutrophil influx in peripheral airways at 6 h post exposure, but no apparent inflammatory response in proximal airways. This response seems to be mediated at least in part by interleukin-8 and growth-related oncogene-alpha.
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PMID:Effects of 0.2 ppm ozone on biomarkers of inflammation in bronchoalveolar lavage fluid and bronchial mucosa of healthy subjects. 965 69

P-selectin glycoprotein ligand-1 (PSGL-1) is a disulfide-bonded, homodimeric mucin ( approximately 250 kDa) on leukocytes that binds to P-selectin on platelets and endothelial cells during the initial steps in inflammation. Because it has been proposed that only covalently dimerized PSGL-1 can bind P-selectin, we investigated the factors controlling dimerization of PSGL-1 and re-examined whether covalent dimers are required for binding its P-selectin. Recombinant forms of PSGL-1 were created in which the single extracellular Cys (Cys(320)) was replaced with either Ser (C320S-PSGL-1) or Ala (C320A-PSGL-1). Both recombinants migrated as monomeric species of approximately 120 kDa under both nonreducing and reducing conditions on SDS-polyacrylamide gel electrophoresis. P-selectin bound similarly to cells expressing either wild type or mutated forms of PSGL-1 in both flow cytometric and rolling adhesion assays. Unexpectedly, chemical cross-linking studies revealed that both C320S- and C320A-PSGL-1 noncovalently associate in the plasma membrane and cross-linking generates dimeric species. Chimeric recombinants of PSGL-1 in which the transmembrane domain in PSGL-1 was replaced with the transmembrane domain of CD43 (CD43TMD-PSGL-1) could not be chemically cross-linked, suggesting that residues within the transmembrane domain of PSGL-1 are required for noncovalent association. Cells expressing CD43TMD-PSGL-1 bound P-selectin. To further address the ability of P-selectin to bind monomeric derivatives of PSGL-1, intact HL-60 cells were trypsin-treated, which generated a soluble approximately 25-kDa NH(2)-terminal fragment of PSGL-1 that bound to immobilized P-selectin. Because N-glycosylation of PSGL-1 hinders trypsin cleavage, a recombinant form of PSGL-1 was generated in which all three potential N-glycosylation sites were mutated (DeltaN-PSGL-1). Cells expressing DeltaN-PSGL-1 bound P-selectin, and trypsin treatment of the cells generated NH(2)-terminal monomeric fragments (<10 kDa) of PSGL-1 that bound to P-selectin. These results demonstrate that Cys(320)-dependent dimerization of PSGL-1 is not required for binding to P-selectin and that a small monomeric fragment of PSGL-1 is sufficient for P-selectin recognition.
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PMID:Noncovalent association of P-selectin glycoprotein ligand-1 and minimal determinants for binding to P-selectin. 1071 99


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