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:3.4.21.5 (thrombin)
33,306 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

1. The efficacy of oligotide, a single stranded polydeoxyribonucleotide complex, was examined in a feline model of myocardial ischaemia (MI: 90 min) and reperfusion (R: 270 min). Oligotide (15 mg kg-1 bolus) was administered intravenously 80 min after occlusion of the left anterior descending (LAD) coronary artery (i.e., 10 min prior to R) and continued for an additional 280 min (10 mg kg-1 h-1 infusion). 2. Oligotide-treated cats showed significantly smaller myocardial necroses and lower cardiac myeloperoxidase activities (significantly lower neutrophil infiltration) in the necrotic zone as compared to MI+R cats receiving only vehicle. 3. LAD coronary arteries isolated from MI+R cats exhibited a significant endothelial dysfunction (i.e., reduced endothelium-dependent relaxation), and significantly increased adherence of polymorphonuclear neutrophils (PMNs) ex vivo. However, oligotide significantly preserved endothelial function and attenuated PMN adherence in ischaemic LAD coronary arteries. 4. Oligotide attenuated P-selectin expression on thrombin-stimulated platelets as well as PMN adherence to thrombin-stimulated coronary endothelium. Immunohistochemical examination in vivo revealed that oligotide treatment also significantly inhibited coronary endothelial P-selectin expression after 90 min MI and 20 min R. 5. Oligotide exerted a significant cardioprotection in MI+R injury. The mechanism appears to be related to attenuation of PMN-endothelial interaction and eventual infiltration into the ischaemic myocardium.
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PMID:Cardioprotective actions of oligotide, a single stranded polydeoxyribonucleotide complex, in myocardial ischaemia and reperfusion injury. 888 89

P-selectin is rapidly translocated to the surface of endothelial cells and platelets following exposure to chemical mediators such as histamine, thrombin and complement factors. The Arthus reaction is caused by vascular injury which is initiated by the local deposition of the immune complex followed by the activation of complement and release of chemical mediators. In this report, the role of P-selectin in the early stage of reverse passive Arthus reaction in rat using monoclonal antibodies (mAbs) against rat P-selectin will be investigated. Intravenous administration of the mAb ARP2-4 significantly attenuated paw edema 1 h after challenging it with antigen by 31.5% (1 mg/kg) and 44.7% (3 mg/kg), respectively. Edema formation was also reduced by sulfatide (73.1%, 50 mg/kg) and inositol hexakisphosphate (InsP6) (72.9%, 30 mg/kg), which have been reported to block P-selectin-mediated neutrophil adhesion. Moreover, neutrophil accumulation into the inflammatory site in the Arthus reaction was inhibited by anti-P-selectin mAb. P-selectin expression was detected along vessel walls prior to neutrophil accumulation, as determined by immunohistochemical staining using the antibody. In addition, the expression of P-selectin mRNA was induced 4 h after deposition of immune complex. From these results, we concluded that P-selectin plays an important role in the pathogenesis of the Arthus reaction especially in the early stage by recruiting neutrophils into sites of inflammation.
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PMID:Role of P-selectin in the early stage of the Arthus reaction. 888 60

This report describes the effect of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF) on platelet production and platelet function in humans. Subjects with advanced solid tumors received PEG-rHuMGDF daily for up to 10 days. There was no increase in circulating platelet count at doses of 0.03 or 0.1 microgram/kg/d by day 12 of study. At doses of 0.3 and 1.0 microgram/kg/d there was a threefold median increase (maximum 10-fold) in platelet count by day 16. The platelets produced in vivo in response to PEG-rHuMGDF showed unchanged aggregation and adenosine triphosphate (ATP)-release responses in in vitro assays. Tests included aggregation and release of ATP in response to adenosine diphosphate (ADP) (10, 5, 2.5, and 1.25 mumol/L), collagen (2 micrograms/mL), thrombin-receptor agonist peptide (TRAP, 10 mumol/L) and ristocetin (1.5 mg/mL). Administration of aspirin to an individual with platelet count of 1,771 x 10(3)/L resulted in the typical aspirin-induced ablation of the normal aggregation and ATP-release response to stimulation with arachidonic acid (0.5 mg/mL), collagen, and ADP (2.5 and 1.25 mumol/L). There was no change in the expression of the platelet-surface activation marker CD62P (P-selectin) nor induction of the fibrinogen binding site on glycoprotein IIb/IIIa as reported by the monoclonal antibody, D3GP3. An elevation of reticulated platelets was evident after 3 days of treatment with PEG-rHuMGDF and preceded the increase in circulating platelet count by 5 to 8 days; this reflected the production of new platelets in response to PEG-rHuMGDF. At later time points, the mean platelet volume (MPV) decreased in a manner inversely proportional to the platelet count. Levels of plasma glycocalicin, a measure of platelet turnover, rose 3 days after the initial increase in the peripheral platelet count. The level of plasma glycocalicin was proportional to the total platelet mass, suggesting that platelets generated in response to PEG-rHuMGDF were not more actively destroyed. Thus, the administration of PEG-rHuMGDF, to humans, increased the circulating platelet count and resulted in fully functional platelets, which showed no detectable increase in reactivity nor alteration in activation status.
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PMID:Administration of pegylated recombinant human megakaryocyte growth and development factor to humans stimulates the production of functional platelets that show no evidence of in vivo activation. 889 92

Nitric oxide (NO) is known as a regulator of platelet function by its anti-adhesive, anti-aggregating, and disaggregating properties. We investigated the modulating effects of the NO-releasing compound SIN-1 (3-morpholino-sydnonimine) on platelet surface glycoprotein (GP) expression during stimulation with human alpha-thrombin. Analysis was performed with two-color flow cytometry using fluoresceine-isothiocyanate (FITC) and phycoerythrin-(PE)-conjugated monoclonal antibodies (MoAbs) directed against GPIb CD42b), GP IIb-IIIa (CD41), P-selectin (CD62P), and MoAb PAC-1 directed against activated GP IIb-IIIa. Preincubation of platelets with SIN-1 (IC50: 1 microM) significantly decreased expression of both total and activated GP IIb-IIIa, and P-selectin in platelets stimulated with thrombin (ED50: 0.05 U/ml), whereas thrombin-induced downregulation of GP Ib was not attenuated. P-selectin expression increased in thrombin-stimulated platelets over time; in contrast, activated GP-IIb-IIIa decreased after an initial peak, indicating that thrombin-induced GP IIb-IIIa activation is spontaneously reversible. SIN-1 reduced P-selectin expression only when added before or at the same time as thrombin, whereas conformationally changed GP-IIb-IIIa was significantly reversed at up to 60 minutes after stimulation by SIN-1. In conclusion, NO attenuates activation marker expression in a dose and time dependent manner. GP-IIb-IIIa is highly sensitive to NO which not only prevents receptor activation but also promotes reversal of activated GP IIb-IIIa complex.
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PMID:The effects of nitric oxide (NO) on platelet membrane receptor expression during activation with human alpha-thrombin. 889 51

Surface coverage with autogeneous endothelial cells is effective in reducing thrombogenicity of an artificial vascular graft, but procedure for obtaining the cells is invasive for patients. The purpose of this study was to establish cultures of human endothelial cells separated from a small piece of subcutaneous fat tissue. A piece of tissue weighing about 10 mg was obtained from subcutaneous fat using a biopsy needle, and treated with collagenase and dispase. Microvascular endothelial cells were selected and other types of cells contaminating the cultures were eliminated by scraping with a needle under a microscope. The yield of the cells was 8362 +/- 4264/10 mg of subcutaneous fat (n = 7). The cultures reached confluence in about 2 weeks. The cells were positive for von Willebrand factor, P-selectin, and uptake of acetylated low density lipoprotein. The cells produced 15.9 +/- 3.3 ng/mg cell protein/h of 6-ketoprostaglandin F1 alpha (n = 5) when stimulated with thrombin. Thrombin also stimulated the production of platelet-activating factor: 7653 +/- 4297 dpm/10(6) cells (n = 5). Endothelin-1 accumulation in the medium of unstimulated endothelial cells was 0.54 +/- 0.16 ng/mg cell protein/10 h (n = 8). As a preliminary experiment for graft seeding, the cells were also cultured on pieces of a gelatin-coated Dacron graft, and scanning electron microscopy revealed the surface coverage of the graft. We herein described about successful culture of human microvascular endothelial cells from subcutaneous fat tissue obtained using a biopsy needle. The cultured cells may be applicable to a seeded vascular graft.
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PMID:Surface coverage of vascular grafts with cultured human endothelial cells from subcutaneous fat tissue obtained with a biopsy needle. 890 4

The effects of endothelin-1 (ET-1) on P-selectin-mediated leukocyte endothelial interaction were examined in vitro. Adherence of autologous polymorphonuclear leukocytes (PMNs) to the endothelium was markedly enhanced by endothelial stimulation with either (2 U/mL) thrombin, (1 mumol/L) histamine, or (100 nmol/L) phorbol myristate acetate (PMA). In contrast, ET-1 alone (10 and 100 nmol/L) only slightly increased the number of adhering PMNs. The increased PMN adherence to thrombin- or histamine-stimulated endothelium, which was blocked by an anti-P-selectin monoclonal antibody, was also significantly attenuated by preincubation of coronary segments with (100 nmol/L) ET-1. We further investigated the mechanism of this anti-adherence action of ET-1 on thrombin-stimulated endothelial adhesiveness. Preincubation of coronary segments with a selective ETA receptor antagonist, BQ485 (1 mumol/L), had no effect on ET-1 inhibition of thrombin-induced PMN adherence. In contrast, preincubation with a selective ETB receptor antagonist, BQ788 (1 mumol/L) significantly reversed ET-1 inhibition of thrombin-induced PMN adherence, whereas the selective ETB receptor agonist BQ-3020 mimicked the inhibitory action of ET-1 on thrombin-induced PMN adherence. Furthermore, (100 mumol/L) N omega-nitro-L-arginine methyl ester (L-NAME), a nitric oxide synthase (NOS) inhibitor, significantly attenuated ET-1 inhibition of thrombin-stimulated PMN adherence. These results suggest that ET-1 may inhibit P-selectin-mediated leukocyte-endothelial interaction via ETB receptor stimulation and subsequent endothelial NO formation. This autocrine effect of ET-1 may be involved in pathophysiologic states such as early atherogenesis by preventing leukocyte-endothelial interaction in constricted blood vessels.
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PMID:Autocrine effects of endothelin-1 on leukocyte-endothelial interaction: stimulation of endothelin B receptor subtype reduces endothelial adhesiveness via a nitric oxide-dependent mechanism. 891 55

We examined the effects of nitric oxide (NO)/endothelium-derived relaxing factor (EDRF) on platelet surface glycoproteins (GP). As determined by flow cytometry, in both a washed platelet system and platelet-rich plasma, the EDRF congener (S-nitroso-N-acetylcysteine) markedly inhibited both the thrombin-induced and the (stable thromboxane A2 analogue) U-46619-induced upregulation of P-selectin (alpha-granule protein), CD63 (lysosomal protein), and the GPIIb-IIIa complex (fibrinogen receptor) but minimally inhibited downregulation of the GPIb-IX complex (von Willebrand factor receptor). The inhibitory effects of EDRF were markedly reduced in whole blood or by the addition of washed erythrocytes. Platelets in whole blood were still responsive to guanosine 3',5'-cyclic monophosphate (cGMP), as shown by complete inhibition of P-selectin upregulation by the stable analogue N6,2'-O dibutyryl cGMP. These data suggests that 1) cGMP negatively regulates the platelet surface expression of P-selectin, CD63, and the GPIIb-IIIa complex but not the platelet surface expression of the GPIb-IX complex and 2) hemoglobin within erythrocytes inhibits the effects of EDRF/NO on platelet surface glycoproteins.
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PMID:Effects of nitric oxide/EDRF on platelet surface glycoproteins. 892 69

Adhesion between platelets and polymorphonuclear leukocytes (PMN) is a key event in thrombosis and inflammation. Double color fluorescence-activated cell sorter (FACS) analysis was used to determine the extent and kinetics of adhesion of thrombin-activated platelets to resting or activated PMN when mixed cell populations were incubated in dynamic conditions. Activated platelets bound very rapidly to PMN. Mixed cell conjugates reached a maximum at 1 minute and were reversible within 10 minutes. Platelet/PMN adhesion required both Ca2+ and Mg2+ and was markedly increased by the presence of Mn2+. The latter made mixed cell conjugates stable up to 10 minutes. Adhesion of platelets required metabolic activity of PMN and was abolished by tyrosine kinase inhibitors. Furthermore, adhesion of platelets to PMN resulted in binding of a monoclonal antibody (MoAb 24) known as beta 2 integrins "activation reporter." When PMN were activated by exogenous stimuli, the adhesion of platelets was markedly increased: fMLP induced a rapid and transient effect, while PMA resulted in a slower, but stable, increase in mixed conjugates formation. The hypothesis that activated PMN beta 2 integrins are able to bind a counter-receptor on platelets was directly demonstrated by the increase of mixed cell conjugates following PMN treatment with KIM127 and KIM185, two anti-CD18 antibodies able to induce the active conformation of beta 2 integrins. Consistently, two other anti-CD18, as well as an anti-CD11b inhibitory antibody abolished platelet/PMN adhesion. PMN beta 2 integrin activation was not the only mechanism for activated platelet/PMN adhesion to occur: indeed, this phenomenon could also be inhibited by two anti-P-selectin antibodies. Resting platelets did not adhere to resting PMN, but markedly adhered to fMLP- or PMA-activated PMN. Resting platelet/fMLP-activated PMN adhesion was abolished by anti-CD18 antibodies, but not by anti-P-selectin antibodies. In conclusion, activated platelet/PMN interaction can be modeled as an adhesion cascade involving a P-selectin-dependent recognition step and a functional signal. The latter proceeds through tyrosine kinase activation and enables a beta 2 integrin-dependent adhesion to a not yet identified counter-receptor constitutively expressed on platelet surface.
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PMID:Platelet/polymorphonuclear leukocyte interaction in dynamic conditions: evidence of adhesion cascade and cross talk between P-selectin and the beta 2 integrin CD11b/CD18. 894 53

The role of GP Ib-IX complexes in platelet adhesion was discovered from studies on patients with the Bernard-Soulier Syndrome (BSS). In this inherited disorder, the bulk of the platelets are round and giant. Furthermore, the platelet count is decreased, often severely so. The relationship between these abnormalities and the deficiency of GP Ib-IX is not well understood. In normal discoid platelets, the bulk of GP Ib-IX is found on the plasma membrane. After platelet activation by thrombin, this distribution changes and the majority of GP Ib-IX complexes are located within the surface-connected canalicular system (SCCS). The platelets now possess pseudopods and are spheroid. Cytoskeletal modifications accompany these changes. We now report that platelets of a BSS variant with a qualitative defect of GP Ib show no translocation of GP Ib in response to thrombin, suggesting that the linkage with the cytoskeleton is impaired. Morphological studies of megakaryocytes (MK) from BSS patients show an altered maturation and an abnormal development of the membrane systems, implying a role for GP Ib-IX in megakaryocytopoiesis. In a case of Epstein syndrome, where giant platelets and thrombocytopenia are associated with deafness and renal dysfunction, platelets possess the bulk of GP Ib-IX inside the SCCS but without signs of platelet activation as assessed by P-selectin expression. This patient also shows an impaired megakaryocytopoiesis and an irregular development of the demarcation membranes within the MK. As collagen IV mutations are a feature of the related Alport syndrome, we hypothesize that defects in the link between extracellular matrix proteins, membrane receptors and the cytoskeleton could be a common cause of giant platelet syndromes.
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PMID:Giant platelets, megakaryocytes and the expression of glycoprotein Ib-IX complexes. 894 96

The adhesive characteristics of hematopoietic stem and progenitor cells may partly regulate their proliferation and differentiation and may be critical in the homing of transplanted stem cells. Using quantitative adhesion assays, we have examined the characteristics of activated platelet adhesion to CD34+ cells in human blood and to the KG1a cell line. Approximately 85-95% of CD34+ cells from both sources bound thrombin-activated platelets; like mature neutrophils, activated platelet binding was maximal within 2 minutes of coincubation. Activated platelet adhesion to CD34+ cells was completely inhibited by chelation of calcium or by preincubation with the G1 blocking monoclonal antibody (MoAb) to platelet P-selectin. Using MoAbs to P-selectin glycoprotein ligand-1 (PSGL-1), we demonstrated that PSGL-1 was present on the surface of CD34+ cells; preincubation of CD34+ cells with the PL1 blocking MoAb to PSGL-1 completely inhibited activated platelet adhesion to CD34+ cells. Furthermore, treatment of CD34+ cells with O-sialoglycoprotein endopeptidase, which destroyed the PL1 epitope of PSGL-1, also abolished activated platelet-CD34+ cell binding. By contrast, MoAb directed against control epitopes of PSGL-1 or endopeptidase-sensitive epitopes of the CD34 molecule had no effect on activated platelet adhesion to CD34+ cells. Unlike mature neutrophils that, when activated, decrease P-selectin-dependent platelet adhesion because of redistribution of PSGL-1, phorbol ester treatment of CD34+ cells had no effect on their ability to bind activated platelets or PSGL-1 MoAbs. This study identifies PSGL-1 on CD34+ cells as the ligand for platelet P-selectin and suggests functional differences between mature and precursor hematopoietic cells in the regulation of surface PSGL-1 expression.
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PMID:Characterization of the P-selectin ligand on human hematopoietic progenitors. 895 Feb 32


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