Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:3.4.21.5 (thrombin)
 33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Vitronectin (serum spreading factor), a major serum cell adhesion molecule, was compared with S-protein, the inhibitor of the C5-9 membrane attack complex. Data from the literature indicate that S-protein and vitronectin are alpha globulins with the same aminoterminal residues, amino acid compositions, and concentrations in normal plasma (150 to 250 micrograms/mL). Both proteins have been reported to interact with the thrombin-antithrombin complex. The cDNA sequences of vitronectin and S-protein were recently determined and found to be almost identical. In the present studies, rabbit-anti-S-protein and a monoclonal antibody to vitronectin both recognized 65,000- and 75,000-molecular weight (mol wt) polypeptides when plasma or serum proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to nitrocellulose paper. The 65,000 and 75,000-mol wt polypeptides bound more avidly from serum than plasma to monoclonal anti-vitronectin or heparin coupled to agarose. The presence or absence of the polypeptides constituted a major difference between the heparin-binding proteins of serum and plasma. When complement-activated serum and unactivated serum were separated by gel filtration, vitronectin coeluted with C9 in high-mol-wt fractions of activated serum but not unactivated serum. Purified S-protein was recognized by the monoclonal antibody to vitronectin and promoted spreading of human skin fibroblasts. Both vitronectin and S-protein were degraded by thrombin. On the basis of immunological and functional, as well as biochemical, properties, therefore, S-protein and vitronectin are the same.
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PMID:On the identity of vitronectin and S-protein: immunological crossreactivity and functional studies. 242 39

It is well known that granulocytes increase infarct size after reperfusion of the ischemic myocardium, and that monocytes promote atherogenesis. Those cells are also believed to play a contributory role in pathogenesis of coronary restenosis as response to arterial injury during balloon angioplasty. The adhesion of those leukocytes to the vascular endothelium is a prerequisite for their recruitment and accumulation in the lesion. Inflammatory mediators likely to occur under those conditions, e.g., histamine, thrombin, oxygen-derived free radicals (ODFR), interleukin (IL)-1, tumor necrosis factor (TNF)-alpha, and activated complement factors, induce in a distinct time course the (transient) expression of the leukocyte adhesion molecules P-selectin, E-selectin, intercellular adhesion molecule (ICAM)-1, and vascular cell adhesion molecule (VCAM)-1 on the endothelium. Only VCAM-1 is specific for monocytes; the others mediate the binding and subsequent extravasation of both monocytes and granulocytes. The response to the relevant inflammatory mediators, except for extracellularly produced ODFR, is coupled via specific receptors on the surface of the endothelium to specific signal transduction pathways and, except for P-selectin (early response), is directly dependent on protein synthesis (intermediate and late response). Protein kinase-C-induced phosphorylation of transcription factors is often shown to be involved. Protein synthesis is preceded by increased transcription of mRNA that is regulated in part by the transcription factor NF-kappa B. Indications have been obtained that intracellularly produced ODFR may be involved in the translocation of this transcription factor.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Leukocyte adhesion molecules on the vascular endothelium: their role in the pathogenesis of cardiovascular disease and the mechanisms underlying their expression. 752 71

Thrombospondin-1 is a glycoprotein that is released from platelet alpha-granules in response to thrombin stimulation and that is also a transient component of extracellular matrix in developing and repairing tissues. It is a 420 kDa homotrimer, each subunit of which consists of multiple structural domains. A variety of factors regulate thrombospondin-1 expression and the protein is degraded by both extracellular and intracellular routes. Thrombospondin-1 functions as a cell adhesion molecule and also modulates cell movement, cell proliferation, neurite outgrowth and angiogenesis. The molecular mechanisms underlying these activities are beginning to be examined. Medical interest in thrombospondin-1 centres on its roles in haemostasis and its effects on angiogenesis.
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PMID:Thrombospondin-1. 930

Although it has been reported that activated platelets can adhere to intact endothelium, the receptors involved have not been fully characterized. Also, it is not clear whether activated platelets bind primarily to matrix proteins at sites of endothelial cell denudation or directly to endothelial cells. Thus, this study was designed to further clarify the mechanisms of activated platelet adhesion to endothelium. Unstimulated human umbilical vein endothelial cell (HUVEC) monolayers were incubated with washed, stained, and thrombin-activated human platelets. To exclude matrix involvement, HUVEC were harvested mechanically and platelet binding was measured by flow cytometry. Before the adhesion assay, platelets or HUVEC were treated with different receptor antagonists. Whereas blockade of platelet beta1 integrins, GPIbalpha, GPIV, P-selectin, and platelet-endothelial cell adhesion molecule (PECAM)-1 did not reduce platelet adhesion to HUVEC, blockade of platelet GPIIbIIIa by antibodies or Arg-Gly-Asp (RGD) peptides markedly decreased adhesion. Moreover, when platelets were treated with blocking antibodies to GPIIbIIIa-binding adhesive proteins, including fibrinogen and fibronectin, and von Willebrand factor (vWF), platelet binding was also reduced markedly. Addition of fibrinogen, fibronectin, or vWF further increased platelet adhesion, indicating that both endogenous platelet-exposed and exogenous adhesive proteins can participate in the binding process. Evaluation of the HUVEC receptors revealed predominant involvement of intercellular adhesion molecule (ICAM)-1 and alphavbeta3 integrin. Blockade of these two receptors by antibodies decreased platelet binding significantly. Also, there was evidence that a component of platelet adhesion was mediated by endothelial GPIbalpha. Blockade of beta1 integrins, E-selectin, P-selectin, PECAM-1, vascular cell adhesion molecule (VCAM)-1 and different matrix proteins on HUVEC did not affect platelet adhesion. In conclusion, we show that activated platelet binding to HUVEC monolayers is mediated by a GPIIbIIIa-dependent bridging mechanism involving platelet-bound adhesive proteins and the endothelial cell receptors ICAM-1, alphavbeta3 integrin, and, to a lesser extent, GPIbalpha.
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PMID:Adhesion of activated platelets to endothelial cells: evidence for a GPIIbIIIa-dependent bridging mechanism and novel roles for endothelial intercellular adhesion molecule 1 (ICAM-1), alphavbeta3 integrin, and GPIbalpha. 944 13

C-CAM, a ubiquitously expressed cell adhesion molecule belonging to the carcinoembryonic antigen family, appears as two co-expressed isoforms, C-CAM-L and C-CAM-S, with different cytoplasmic domains, that can form homodimers in epithelial cells. In addition, C-CAM-L has been found in large molecular weight forms suggesting posttranslational, covalent modification. Here we have investigated the possibility that the cytoplasmic domain of C-CAM-L can act as a transglutaminase substrate. Glutathione S-transferase fusion proteins of the cytoplasmic domains of rat and mouse C-CAM-L as well as free cytoplasmic domains, released by thrombin cleavage from the fusion proteins, were converted into covalent dimers by tissue transglutaminase. These results demonstrate that the cytoplasmic domains of rat and mouse C-CAM-L are substrates for tissue transglutaminase, and lend support to the notion that higher molecular weight forms of C-CAM-L are formed by transglutaminase modification.
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PMID:The cell adhesion molecule C-CAM is a substrate for tissue transglutaminase. 954 Oct 24

P-selectin, an inducible cell adhesion molecule, mediates rolling of neutrophils on activated vascular endothelium. Because rolling is an early event of the inflammatory response, therapeutic applications of selectin antagonists have been of broad interest. There are, however, no truly satisfactory therapeutic candidates among known inhibitors. Consequently, we have used Systematic Evolution of Ligands by Exponential Enrichment (SELEX) technology, a process based on oligonucleotide combinatorial chemistry and in vitro selection, to develop aptamer antagonists of P-selectin. Equilibrium dissociation constants for aptamer/P-selectin binding range from 16 to 710 pM, a 10(5)-10(6)-fold improvement compared with the minimal carbohydrate ligand, sialyl Lewis X (sLeX). Aptamer binding is divalent cation dependent and, unlike sLeX, is specific for P-selectin. The selectivity for human P-selectin relative to human E-selectin or human L-selectin is 10(4)-10(5). In vitro, aptamers bind with subnanomolar affinities to P-selectin expressed on thrombin-activated platelets, inhibit the binding of P-selectin-IgG chimera to sLeX and to neutrophils, and block the binding activated platelets to neutrophils in flow cytometry and in hydrodynamic assays. Extrapolating from their in vitro characteristics, these novel P-selectin-specific antagonists may be suitable candidates for therapeutic development.
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PMID:Oligonucleotide inhibitors of P-selectin-dependent neutrophil-platelet adhesion. 974 65

The type IIA rat brain sodium channel is composed of three subunits: a large pore-forming alpha subunit and two smaller auxiliary subunits, beta1 and beta2. The beta subunits are single membrane-spanning glycoproteins with one Ig-like motif in their extracellular domains. The Ig motif of the beta2 subunit has close structural similarity to one of the six Ig motifs in the extracellular domain of the cell adhesion molecule contactin (also called F3 or F11), which binds to the extracellular matrix molecules tenascin-C and tenascin-R. We investigated the binding of the purified sodium channel and the extracellular domain of the beta2 subunit to tenascin-C and tenascin-R in vitro. Incubation of purified sodium channels on microtiter plates coated with tenascin-C revealed saturable and specific binding with an apparent Kd of approximately 15 nM. Glutathione S-transferase-tagged fusion proteins containing various segments of tenascin-C and tenascin-R were purified, digested with thrombin to remove the epitope tag, immobilized on microtiter dishes, and tested for their ability to bind purified sodium channel or the epitope-tagged extracellular domain of beta2 subunits. Both purified sodium channels and the extracellular domain of the beta2 subunit bound specifically to fibronectin type III repeats 1-2, A, B, and 6-8 of tenascin-C and fibronectin type III repeats 1-2 and 6-8 of tenascin-R but not to the epidermal growth factor-like domain or the fibrinogen-like domain of these molecules. The binding of neuronal sodium channels to extracellular matrix molecules such as tenascin-C and tenascin-R may play a crucial role in localizing sodium channels in high density at axon initial segments and nodes of Ranvier or in regulating the activity of immobilized sodium channels in these locations.
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PMID:Interaction of voltage-gated sodium channels with the extracellular matrix molecules tenascin-C and tenascin-R. 986 Oct 42

Activated platelets and endothelium surface express the cell adhesion molecule P-selectin (CD62P), which plays an important role in mediating interactions with leukocytes. Increased levels of a functional soluble form of P-selectin (sP-selectin) have been reported in several pathological states but it is not clear whether this circulating sP-selectin originates from platelets and/or endothelial cells. Here we describe the concurrent kinetics of intracellular storage, surface expression and release of platelet P-selectin induced by thrombin or the protein kinase C activator PMA. Platelet activation with submaximal concentrations of thrombin (0.1 U/ml) resulted in a rapid decrease of intracellular P-selectin. This decrease of intracellular P-selectin concurred with a gradual increase of surface expression and an initial increase of sP-selectin. Our results indicate that intracellular stores of P-selectin were only partly mobilized upon activation with submaximal concentrations of thrombin. A high concentration of thrombin (1.0 U/ml) induced a rapid and nearly total decrease of intracellular stores and a more pronounced, but transient, increase of surface expression. The release of P-selectin was fast and occurred during the initial activation phase. The NO donor SNAP inhibited both surface expression and release of platelet P-selectin in a similar manner. PMA (0.1-1.0 microM) mediated a more slow, gradual and sustained surface expression and release of P-selectin than thrombin. Thus, surface expression and release of platelet P-selectin show different kinetics depending on the mode of activation.
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PMID:Kinetics of platelet P-selectin mobilization: concurrent surface expression and release induced by thrombin or PMA, and inhibition by the NO donor SNAP. 986 63

Osteopontin is a secreted glycoprotein with adhesive and migratory functions. Cellular interactions with osteopontin are mediated through integrin receptors which recognize the RGD domain. Recently, CD44, a non-integrin, multifunctional adhesion molecule was identified as an osteopontin receptor. CD44 is a ubiquitous surface molecule that exists as a number of different isoforms, generated by alternative splicing. To analyze which forms of CD44 mediate binding to osteopontin, we used the standard form of CD44 as CD44-human immunoglobulin fusion proteins and several splice variants in enzyme-linked immunosorbant assays. Multiple preparations of osteopontin were used including native osteopontin derived from smooth muscle cells, human urinary osteopontin, full-length recombinant osteopontin, and two recombinant osteopontin fragments expected to be formed following thrombin cleavage. Our data show that although the CD44-hlg fusion proteins could interact with hyaluronic acid as expected, there was no interaction between CD44H, CD44E, CD44v3,v8-v10, or CD44v3 with osteopontin. These studies suggest that CD44-osteopontin interactions may not be common in vivo and may be limited to a specific CD44 isoform(s), and/or a particular modified form of osteopontin.
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PMID:CD44 is not an adhesive receptor for osteopontin. 1008 20

The distribution of a soluble form of a cell adhesion molecule, P-selectin, in human platelets and cultivated endothelial cells has been studied by enzyme-linked immunosorbent assay (ELISA). The concentration of soluble P-selectin in the blood plasma of healthy donors and patients with abnormal platelet count has also been determined. P-selectin was measured in the Triton X-100 lysate of platelets and endothelial cells (total P-selectin), in the 100,000g supernatant obtained after sedimentation of the membrane fraction from the homogenate of sonicated platelets and endothelial cells (intracellular soluble P-selectin), in the supernatant of activated and nonactivated platelets, and in the culture medium of endothelial cells. A soluble form of P-selectin which did not coprecipitate with the membrane fraction was detected in platelets and accounted for approximately 10% of the total P-selectin. Platelet activation by thrombin, ADP, or a thromboxane A2 analog resulted in the secretion of 30-50% of the intracellular soluble P-selectin. Measurements of P-selectin in endothelial cell culture revealed that endothelium from aorta contained about twofold more P-selectin than endothelium from umbilical vein. Intracellular soluble P-selectin was identified in both types of endothelial cells. In endothelial cells from the umbilical vein this form made up approximately 10% of the total P-selectin. Soluble P-selectin was also detected in the medium of cultivated endothelial cells, where its content correlated with the total cellular P-selectin. Concentration of P-selectin in blood plasma strongly correlated with the platelet count in the blood of healthy donors and patients with thrombocytosis and thrombocytopenia. These data indicate that platelets serve as one of the main source of plasma P-selectin. However, the presence of P-selectin in the plasma of patients with severe thrombocytopenia suggests that endothelium can also be involved in plasma P-selectin production. Thus, in vitro experiments as well as measurements of plasma P-selectin have shown that both platelets and endothelial cells can produce a soluble form of the protein. Platelet-derived soluble P-selectin and plasma P-selectin were shown to react with antibodies against the cytoplasmic domain of P-selectin. These data prove that at least part of soluble P-selectin is produced by synthesis employing special mRNA which lacks the sequence encoding the transmembrane domain, but not by the proteolytic shedding of the extracellular portion of membrane P-selectin.
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PMID:Production of soluble P-selectin by platelets and endothelial cells. 1061 41


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