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PubMed1981

Antibodies against platelet membrane glycoproteins. I. Crossed immunoelectrophoresis studies with antibodies that inhibit ristocetin-induced platelet aggregation.

Platelet membrane glycoproteins have been isolated by lectin-affinity chromatography and antibodies prepared against them. Platelets that have lost glycocalicin no longer respond to ristocetin-human VIIIR:WF, bovine VIIIR:WF, or to glycocalicin or glycoproteins Ia and Ib antibodies but are still agglutinated by glycoproteins IIb and IIIa antibodies. Glycoproteins Ia and Ib and glycocalicin antibodies, IgG and Fab' fragments, inhibited ristocetin-human VIIIR:WF-induced aggregation of fixed, washed platelets and of platelets in plasma while glycoproteins IIb and IIIa antibodies were without effect. Cross immunoelectrophoretic studies showed that glycocalicin was present on whole platelets in only trace amounts. Glycocalicin antibodies, however, recognized a slower migrating component. Platelets incubated in an EDTA-free medium no longer respond to ristocetin-human VIIIR:WF. Membranes isolated from such platelets contained glycocalicin which cross-reacted with a remnant of the slower migrating component. Glycoproteins Ia and Ib antibodies gave more complex patterns but it was possible to identify the slower moving component recognized by the glycocalicin antibodies. These results show that glycocalicin is not normally found as such on whole platelets but is present as a precursor which is most likely glycoprotein Ib. On degradation of this precursor, glycocalicin is released from the membrane and VIIIR:WF-receptor activity is lost.

Animals↗
PubMed1986

Alloantibody-induced platelet serotonin release is blocked by antibody to the platelet PLA1 antigen.

Anti-platelet antibodies are known to activate the platelet release mechanism, usually measured by [14C]serotonin secretion. We have made immunoglobulin preparations of antibodies to HLA antigens, blood group A and platelet-specific PlA1 antigen on the platelet surface and have defined the optimal conditions for [14C]serotonin secretion by washed platelets. For optimal [14C]serotonin release, platelets were washed in apyrase and suspended in Tyrode-albumin with Ca++ and Mg++ and fibrinogen. Anti-PlA1 at high concentration did not cause [14C]serotonin release and inhibited serotonin release by other alloantibodies to antigens on the platelet surface. This was shown to be a result of the inhibition of fibrinogen binding to the platelet by bound anti-PlA1.

ABO Blood-Group System↗
PubMed2006

Differences in platelet growth factor release and leucocyte kinetics during autologous platelet gel formation.

Three commercial systems for whole blood separation were compared to obtain the buffy coat composed of platelet-rich plasma (BC-PRP) and leucocytes . These samples of the buffy coat were used to make a platelet gel (PG), which was used to measure platelet growth factor (PGF) release, to perform a white blood cell (WBC) count and to measure myeloperoxidase (MPO) release from WBCs. Aliquots of whole blood obtained from ten volunteers were distributed either to a blood cell separator (The Electa Cell-Separator, E-CS) or to a tabletop centrifuge (Gravitational Platelet Sequestration System, GPS) to prepare the BC-PRP. The third system combines the BC-PRP production by E-CS with a micro porous filter (Autologous Growth Factor filter, AGF) to enrich for the BC-PRP. Autologous thrombin was used to activate the BC-PRP and to prepare the PG and subsequently to degranulate the platelet concentrate. Platelet-derived growth factor-AB and transforming growth factor-beta1 were present in high levels after thrombin activation of the E-CS or GPS prepared samples. However, the AGF prepared samples released their growth factors before thrombin activation. The WBCs were significantly increased with each of the three systems. Contrary to the AGF, no leucocyte degranulation occurred with the E-CS or GPS prepared samples, based upon the low MPO concentrations in the BC-PRP. The three types of apparatus had different harvesting capacities for collecting the enriched platelets and the release of high concentrations of PGF. When the E-CS and GPS, but not the AGF, were used, low levels of MPO were maintained in the PG, which potentially contributes to antimicrobial properties of platelet gel at the site of application.

Adult↗
PubMed2005

Frequencies of maternal platelet alloantibodies and autoantibodies in suspected fetal/neonatal alloimmune thrombocytopenia, with emphasis on human platelet antigen-15 alloimmunization.

BACKGROUND AND OBJECTIVES: Serological evaluation of maternal sera for platelet antibodies in suspected fetal/neonatal alloimmune thrombocytopenia (FNAITP) discloses in only approximately 30% of individuals a platelet-specific antibody. Transfusion-induced alloimmunization against human platelet antigen-15 (HPA-15) has been reported to be about as common as against HPA-5, the second most common platelet antibody. Thus, anti-HPA-15 may also contribute significantly to yet-unclear cases of FNAITP. MATERIALS AND METHODS: In this retrospective analysis, we provide data on maternal platelet antibodies from 309 mothers who delivered an offspring with suspected FNAITP. RESULTS: Genotyping maternal and paternal samples (together n = 573) revealed a gene frequency of 0.496 for HPA-15a and a gene frequency of 0.504 for HPA-15b. HPA-15 antibodies were detected in 2% of all samples. Anti-HPA-15a and -15b were detected in two and three samples, respectively. One serum reacted equally with HPA-15a and -15b platelets. The most frequent platelet-specific antibodies were anti-HPA-1a (22%), but anti-HPA-5b (8.4%) were more frequent than anti-HPA-15. In addition, panreactive (5.5%) or autoreactive (5.2%) anti-GPIIb/IIIa or anti-GPIb/IX were detectable in maternal samples. CONCLUSIONS: These data indicate that HPA-15 alloimmunization needs only to be considered in subjects with suspected FNAITP if no other platelet-specific antibody is detectable. The presence of panreactive or autoreactive antibodies should also be considered in neonatal thrombocytopenia.

Antigens, CD↗
PubMed1991

Stimulation of non-selective cation channels providing Ca2+ influx into platelets by platelet-activating factor and other aggregation inducers.

To elucidate the mechanism of the receptor-stimulated Ca2+ entry into human platelets, the influence of Ca(2+)-mobilizing agonists on plasma membrane potential (Em) has been studied. Em changes were registered using potentiometric probe 3,3'-dipropyl-2,2'-thiadicarbocyanine iodide. The agonist effect on Em varied from hyperpolarization to slight and slow rise. On the contrary, after loading of platelets with intracellular Ca2+ indicator quin2, platelet-activating factor (PAF), thrombin, vasopressin, ADP and thromboxane-A2-mimetic U46619 cause substantial transient membrane depolarization. Similar effects were observed after platelet loading with other Ca2+ chelators fura-2 and indo-1. Agonist-induced depolarization considerably reduced if quin2-loaded platelets were suspended in isoosmotic choline-containing medium. Using Ba2+ as a substitute of Ca2+, we have demonstrated that in choline-containing medium PAF-induced Ba2+ entry into platelets results in membrane depolarization. Dependence on Ba2+ concentration and depolarization kinetics correlates with the dose dependence and kinetics of Ba2+ entry detected by quin2 fluorescence. The agonists also stimulate considerable Na+, Li+ and Cs+ inward currents into platelets. Na(+)-dependent depolarization is 2-5-fold suppressed by extracellular Ca2+ [median inhibitory concentration (IC50) approximately 0.3 mM]. Ni2+ and Cd2+ at similar concentrations block Ca2+ entry and agonist-induced Na2+ current (IC50 for both cations approximately 50 microM). Agonist-induced depolarization is blocked by the adenylate cyclase stimulator prostaglandin E1 and the protein kinase C stimulator phorbol ester. It is concluded that agonists stimulate Ca2+ entry into human platelets via receptor-operated channels which are not strictly selective toward divalent cations and are permeable to Na+, Li+ and Cs+.

Aminoquinolines↗
PubMed1990

Endothelin-1 inhibits platelet aggregation in vivo: a study with 111indium-labelled platelets.

1. A non-invasive technique for the scintigraphic determination of 111indium-labelled platelet aggregation stimulated with submaximal doses of adenosine diphosphate (ADP, 56 micrograms kg-1 i.v.), collagen (100 micrograms kg-1 i.v.), platelet-activating factor (PAF, 0.1 microgram kg-1 i.v.) or thrombin (18 iu kg-1 i.v.) was used to investigate the platelet-inhibitory effects of endothelin 1 (ET-1) in anaesthetized rabbits in vivo. 2. ET-1 (1 nmol kg-1 i.v.) inhibited ADP-stimulated platelet aggregation in vivo; a maximum inhibition of 78% of the control value was reached at 3 min, with 45% inhibition at 15 min, and a return to control values at 30 min after injection of the peptide. 3. ET-1 (1 nmol kg-1 i.v.) inhibited in vivo platelet aggregation in response to collagen or PAF by 86% and 52%, respectively, but had no effect on thrombin-induced platelet aggregation. 4. Indomethacin (5 mg kg-1 i.v.) abolished the ET-1-induced inhibition of ADP-stimulated platelet aggregation and significantly potentiated and prolonged the pressor response brought about by ET-1. 5. In conclusion, the data demonstrate that ET-1 potently inhibits platelet aggregation in the anaesthetized rabbit in vivo by releasing a hypotensive and anti-aggregatory cyclo-oxygenase product, presumably prostacyclin, into the circulation.

Adenosine Diphosphate↗
PubMed2004

Ingestion of quercetin inhibits platelet aggregation and essential components of the collagen-stimulated platelet activation pathway in humans.

BACKGROUND: Quercetin, a flavonoid present in the human diet, which is found in high levels in onions, apples, tea and wine, has been shown previously to inhibit platelet aggregation and signaling in vitro. Consequently, it has been proposed that quercetin may contribute to the protective effects against cardiovascular disease of a diet rich in fruit and vegetables. OBJECTIVES: A pilot human dietary intervention study was designed to investigate the relationship between the ingestion of dietary quercetin and platelet function. METHODS: Human subjects ingested either 150 mg or 300 mg quercetin-4'-O-beta-D-glucoside supplement to determine the systemic availability of quercetin. Platelets were isolated from subjects to analyse collagen-stimulated cell signaling and aggregation. RESULTS: Plasma quercetin concentrations peaked at 4.66 microm (+/- 0.77) and 9.72 microm (+/- 1.38) 30 min after ingestion of 150-mg and 300-mg doses of quercetin-4'-O-beta-D-glucoside, respectively, demonstrating that quercetin was bioavailable, with plasma concentrations attained in the range known to affect platelet function in vitro. Platelet aggregation was inhibited 30 and 120 min after ingestion of both doses of quercetin-4'-O-beta-D-glucoside. Correspondingly, collagen-stimulated tyrosine phosphorylation of total platelet proteins was inhibited. This was accompanied by reduced tyrosine phosphorylation of the tyrosine kinase Syk and phospholipase Cgamma2, components of the platelet glycoprotein VI collagen receptor signaling pathway. CONCLUSIONS: This study provides new evidence of the relatively high systemic availability of quercetin in the form of quercetin-4'-O-beta-D-glucoside by supplementation, and implicates quercetin as a dietary inhibitor of platelet cell signaling and thrombus formation.

Blood Platelets↗
PubMed1990

Platelet calmodulin correlates with platelet turnover.

We measured the calmodulin content in platelets in 13 normal persons and in 62 patients with hematological diseases. The level of platelet calmodulin was higher in patients with idiopathic thrombocytopenic purpura (ITP), systemic lupus erythematosus, myeloproliferative disorders, acute leukemia in a recovery phase, aplastic anemia, thrombosis and hypersplenism as compared to the controls. Among the patients with ITP, calmodulin was lower in responders than in nonresponders and those at the initial diagnosis. We also measured the volume, life-span and aggregation of the platelets and demonstrated a significant relationship between the calmodulin level and the platelet volume, and a negative relationship between the calmodulin level and platelet life-span, there was no correlation between the calmodulin level and platelet aggregation. We thus conclude that platelet calmodulin is inversely correlated with platelet turnover.

Biomarkers↗
PubMed2006

Reduced blood platelet sensitivity to aspirin in coronary artery disease: are dyslipidaemia and inflammatory states possible factors predisposing to sub-optimal platelet response to aspirin?

The study was designed to assess blood platelet sensitivity to acetylsalicylic acid and its associations with dyslipidaemia and inflammation in coronary artery disease patients. Platelet non-responsiveness to aspirin is associated with an increased risk of serious cardiovascular events. Several environmental and hereditary factors are reportedly involved in sub-optimal acetylsalicylic acid response. Forty-five coronary artery disease patients and 45 non-coronary artery disease controls received acetylsalicylic acid at a daily dose of 75-150 mg. Controls were examined twice: on the day of entering the study and 10 days later. Urinary 11-dehydrothromboxane B2 was assessed as the marker of platelet thromboxane generation. Aggregation was studied in platelet-rich plasma using turbidimetric aggregometry with collagen and arachidonic acid. Fifty to seventy percent of coronary artery disease patients showed an extent of collagen-induced aggregation above the upper quartile of the reference range compared with 8-15% in controls (P<0.003). For arachidonic acid-activated aggregation these proportions were 45-50% in coronary artery disease versus 7% in controls (P<0.007). In coronary artery disease patients, the acetylsalicylic acid-mediated platelet inhibition positively correlated with increased triglycerides (in arachidonic acid-stimulated platelets, r=0.30, P=0.0018), total cholesterol (r=0.33, P<0.0001 in coll and arachidonic acid-activated platelets) and elevated serum C-reactive protein (CRP) (r=0.27, P=0.0024). In coronary artery disease patients urine 11-dehydrothromboxane B2 concentrations were significantly increased compared to controls after 10 day acetylsalicylic acid intake (563; 313-728 pg/mg creatinine versus 321; 246-488 pg/mg creatinine, P=0.04). The incidence of suboptimal acetylsalicylic acid response incidence was more common in patients with coronary artery disease. Acetylsalicylic acid inhibition of blood platelet reactivity and thromboxane generation was less effective in these patients. Dyslipidaemia and chronic inflammatory states may promote suboptimal acetylsalicylic acid response in coronary artery disease patients.

Anti-Inflammatory Agents, Non-Steroidal↗
PubMed2002

Comparative analysis of various platelet glycoprotein IIb/IIIa antagonists on shear-induced platelet activation and adhesion.

Platelet accretion into arterial thrombus in stenotic arterial vessels involves shear-induced platelet activation and adhesion. The Cone and Plate(let) Analyzer (CPA) is designed to simulate such conditions in vitro under a rotating high shear rate in whole blood. In the present study, we evaluated various experimental conditions (including aspirin, temperature, and calcium concentration) and investigated the effects of small molecules along with peptide glycoprotein IIb/IIIa antagonists on platelet adhesion using the CPA system. Concentration-dependent effect of glycoprotein IIb/IIIa antagonists on shear-induced platelet adhesion showed marked differences in potencies: IC50 = 34, 35, 91, 438, and 606 nM for DPC802 (a specific glycoprotein IIb/IIIa antagonist), roxifiban, sibrafiban, lotrafiban, and orbofiban (free acid forms), respectively, and IC50 values of 43, 430, and 5781 nM for abciximab, tirofiban, and eptifibatide, respectively. Parallel study was also conducted to evaluate the effect of glycoprotein IIb/IIIa inhibitors using optical aggregometry. The potency of fibans in blocking shear-induced platelet adhesion correlated well with their binding affinity to the resting and activated glycoprotein IIb/IIIa receptors, as well as their "off-rates". Nevertheless, none of these fibans was able to effectively block shear-induced platelet adhesion at targeted clinical dosing regimens except for abciximab. These data suggest that glycoprotein IIb/IIIa antagonists that show similar efficacy in the inhibition of platelet aggregation in a static in vitro assay may differ substantially in a shear-based system of platelet adhesion. The clinical significance of this phenomenon awaits further investigation.

Abciximab↗
PubMed1994

Pertussis toxin activates platelets through an interaction with platelet glycoprotein Ib.

Platelets present a unique model to study the B-oligomer effects of pertussis toxin because they become activated in response to the B oligomer but are not susceptible to ADP-ribosylation by the holotoxin. In these studies, the B oligomer of pertussis toxin caused concentration-dependent platelet activation, as determined by increases in intracellular calcium concentration, dense granule secretion, and platelet aggregation. Stirring was required for pertussis toxin to increase intracellular calcium. A monoclonal antibody against platelet glycoprotein Ib abolished increases in intracellular calcium concentration and increased the latency and reduced the slope of the aggregation response elicited by the B oligomer. Pertussis toxin also evoked [14C]serotonin release from platelets, and this effect was inhibited, though not eliminated, by an antibody against platelet glycoprotein Ib. Binding of pertussis toxin to glycoprotein Ib was observed after nonreducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis. These data suggest that the B oligomer of pertussis toxin induces platelet activation mediated, at least in part, by an interaction with platelet glycoprotein Ib.

Blood Platelets↗
PubMed1995

Decreased platelet function in aortic valve stenosis: high shear platelet activation then inactivation.

OBJECTIVE: To elucidate the mechanism of the bleeding tendency observed in patients with aortic valve stenosis. DESIGN: A prospective study of high and low shear platelet function tests in vitro in normal controls compared with that in patients with severe aortic valve stenosis with a mean (SD) systolic gradient by Doppler of 75 (18) mm Hg before and at least 4 months after aortic valve replacement. SETTING: District general hospital. RESULTS: The patients showed reduced retention in the high shear platelet function tests. (a) Platelet retention in the filter test was 53.6 (12.6)% in patients with aortic valve stenosis and 84.8 (9.6)% in the controls (P < 0.001). (b) Retention in the glass bead column test was 49.8 (19.2) in the patients and 87.4 (8.7) in the controls (P < 0.001). (c) The standard bleeding time was longer in the patients (P < 0.06). Results of the high shear tests (a, b, and c) after aortic valve replacement were within the normal range. The platelet count was low but within the normal range before surgery and increased postoperatively (P < 0.01). There were no differences in the results of standard clotting tests, plasma and intraplatelet von Willebrand's factor, or in 15 platelet aggregation tests using five agonists between patients with aortic valve stenosis and controls. CONCLUSIONS: The high shear haemodynamics of aortic valve stenosis modify platelet function in vivo predisposing to a bleeding tendency. This abnormality of platelet function is detectable only in vitro using high shear tests. The abnormal function is reversed by aortic valve replacement. High shear forces in vitro activate and then inactivate platelets. By the same mechanisms aortic valve stenosis seems to lead to high shear damage in vivo, resulting in a clinically important bleeding tendency in some patients.

Aged↗
PubMed1991

Platelet-targeted fibrinolysis enhances clot lysis and inhibits platelet aggregation.

BACKGROUND: Although plasminogen activator therapy has been shown to reduce mortality in patients with severe myocardial infarction, several problems fuel the search for more potent and specific thrombolytic agents. METHODS AND RESULTS: To explore the effect of plasminogen activator targeting to platelets, we covalently linked urokinase that had been modified with N-succinimidyl-3-(2-pyridyldithio)propionate to the Fab' of a monoclonal antibody (7E3) that selectively binds to platelet membrane glycoprotein (GP) IIb/IIIa. In an assay measuring (as reflected by plasmin generation) a plasminogen activator's ability to bind GP IIb/IIIa immobilized on plastic, urokinase-7E3 Fab' produced 31-fold more plasmin than did urokinase (p = 0.0001). The addition of solubilized GP IIb/IIIa blocked this enhancement of plasmin generation, indicating that binding was impaired. Plasmin generation reflecting binding to immobilized intact platelets was 2.4-fold greater for urokinase-7E3 Fab' than for unconjugated urokinase (p = 0.002). In a plasma clot lysis assay, urokinase-7E3 Fab' was at least 25-fold more potent than either urokinase alone or a mixture of urokinase and 7E3 (Fab')2 (p less than 0.009), and potency could be related to platelet concentration in the clot. Ex vivo, ADP-induced platelet aggregation was inhibited by a urokinase-7E3 IgG conjugate at a concentration of 8 nM, whereas a mixture of urokinase and 7E3 (Fab')2 in equimolar amounts required 60 nM and urokinase alone required 1 microM to achieve the same effect. CONCLUSIONS: Therefore, the targeting of urokinase to the GP IIb/IIIa platelet receptor both accelerates clot lysis (when platelets are associated with a fibrin clot) and inhibits platelet aggregation.

Antigens↗
PubMed1999

Platelet microparticles promote platelet interaction with subendothelial matrix in a glycoprotein IIb/IIIa-dependent mechanism.

BACKGROUND: Platelets, on activation, release vesicular particles called platelet microparticles. Despite their procoagulant activity, their functional role in platelet-vessel wall interactions is not known. METHODS AND RESULTS: We examined the binding of microparticles to vessel wall components in vitro and in vivo. Microparticles bound to fibrinogen-, fibronectin-, and collagen-coated surfaces. Compared with activated platelets, we observed minimal binding of microparticles to vitronectin and von Willebrand factor. The glycoprotein IIb/IIIa (GP IIb/IIIa) inhibitors abciximab and eptifibatide (Integrilin) inhibited the binding to fibrinogen and fibronectin but had minimal effect on binding to collagen. Furthermore, monoclonal antibodies to GP Ib or anionic phospholipid-binding proteins (beta2-glycoprotein I or annexin V) had no effect in these interactions. Microparticles did not bind to monolayers of resting or stimulated human umbilical vein endothelial cells (HUVECs), even in the presence of fibrinogen or von Willebrand factor. However, under similar conditions, microparticles bound to extracellular matrix produced by cultured HUVECs. Abciximab inhibited this interaction by approximately 50%. In a rabbit model of arterial endothelial injury, the infusion of 51Cr-labeled microparticles resulted in a 3- to 5-fold increase of microparticle adhesion to the injured site compared with the uninjured site (P<0.05%). Furthermore, activated platelets bound to surface-immobilized microparticles in a GP IIb/IIIa-dependent mechanism. This binding increased in the presence of fibrinogen by approximately 30%. CONCLUSIONS: Platelet microparticles bind to subendothelial matrix in vitro and in vivo and can act as a substrate for further platelet binding. This interaction may play a significant role in platelet adhesion to the site of endothelial injury.

Adult↗
PubMed1984

Retention of the glycoprotein IIb-IIIa complex in the isolated platelet cytoskeleton. Effects of separable assembly of platelet pseudopodal and contractile cytoskeletons.

To investigate the association of the putative platelet fibrinogen receptor (glycoprotein IIb-III(a) with the cytoskeleton, 125I-surface labeled human platelets washed by gel-filtration were activated under conditions which allow selective assembly of the platelet cytoskeleton. The four conditions were activation with arachidonate or phorbol 12-myristate 13-acetate (PMA) with and without pretreatment with cytochalasin E. Activation with arachidonate generates a complete cytoskeletal core (pseudopodal and contractile elements) while PMA activation forms only an actin plus actin-binding protein pseudopodal core. Pretreatment with cytochalasin E leads to actomyosin contractile core formation if arachidonate activated, and essentially blocks cytoskeletal development if PMA activated. Cytoskeletal cores from arachidonate or PMA-activated platelets retained 26 (+/- 3%) of the total 125I-IIIa. Pretreatment with cytochalasin E followed by arachidonate or PMA activation reduced the 125I-IIIa retention to near control levels (unactivated platelets: 4 +/- 2%). The role of aggregation vs. receptor occupancy in the retention of IIb-IIIa was assessed by activation of platelets with arachidonate in the presence of fibrinogen fragment D (0.6-12 mg/ml). Aggregation was blocked by increasing concentrations of fragment D reagent while cytoskeletal assembly was not altered. The IIIa retention correlated with extent of aggregation with maximal retention corresponding to full aggregation. To determine if cytoskeletal development is necessary for the expression of the fibrinogen binding site, binding studies were performed with unlabeled platelets and 125I-fibrinogen. The mean number of binding sites and the mean dissociation constant were not significantly different among the four activation conditions. Although the development of a platelet cytoskeletal core is not required for the expression of the fibrinogen binding site, the retention of the glycoprotein IIb-IIIa complex is dependent on fibrinogen-supported aggregation as well as the formation of the pseudopodal cytoskeleton.

Arachidonic Acid↗
PubMed1991

von Willebrand factor binding to platelet GpIb initiates signals for platelet activation.

The hypothesis that von Willebrand factor (vWF) binding to platelet membrane glycoprotein Ib (GpIb) initiates intracellular pathways of platelet activation was studied. We measured the biochemical responses of intact human platelets treated with ristocetin plus vWF multimers purified from human cryoprecipitate. vWF plus ristocetin causes the breakdown of phosphatidylinositol 4,5-bisphosphate, the production of phosphatidic acid (PA), the activation of protein kinase C (PKC), increase of ionized cytoplasmic calcium ([Ca2+]i), and the synthesis of thromboxane A2. PA production, PKC activation, and the rise of [Ca2+]i stimulated by the ristocetin-induced binding of vWF multimers to platelets are inhibited by an anti-GpIb monoclonal antibody, but are unaffected by anti-GpIIb-IIIa monoclonal antibodies. Indomethacin also inhibits these responses without impairing platelet aggregation induced by vWF plus ristocetin. These results indicate that vWF binding to platelets initiates specific intraplatelet signaling pathways. The mechanism by which this occurs involves an arachidonic acid metabolite-dependent activation of phospholipase C after vWF binding to platelet membrane GpIb. This signal then causes PKC activation and increases of [Ca2+]i, which promote platelet secretion and potentiate aggregation.

Calcium↗
PubMed2002

Platelet endothelial cell adhesion molecule-1 signaling inhibits the activation of human platelets.

Platelet endothelial cell adhesion molecule-1 (PECAM-1/CD31) is a 130-kd transmembrane glycoprotein and a member of the growing family of receptors with immunoreceptor tyrosine-based inhibitory motifs (ITIMs). PECAM-1 is expressed on platelets, certain T cells, monocytes, neutrophils, and vascular endothelial cells and is involved in a range of cellular processes, though the role of PECAM-1 in platelets is unclear. Cross-linking of PECAM-1 results in phosphorylation of the ITIM allowing the recruitment of signaling proteins that bind by way of Src-homology domain 2 interactions. Proteins that have been implicated in the negative regulation of cellular activation by ITIM-bearing receptors include the tyrosine phosphatases SHP-1 and SHP-2. Tyrosine phosphorylation of immunoreceptor tyrosine-based activatory motif (ITAM)-bearing receptors such as the collagen receptor GPVI-Fc receptor gamma-chain complex on platelets leads to activation. Increasing evidence suggests that ITIM- and ITAM-containing receptors may act antagonistically when expressed on the same cell. In this study it is demonstrated that cross-linking PECAM-1 inhibits the aggregation and secretion of platelets in response to collagen and the GPVI-selective agonist convulxin. In these experiments thrombin-mediated platelet aggregation and secretion were also reduced, albeit to a lesser degree than for collagen, suggesting that PECAM-1 function may not be restricted to the inhibition of ITAM-containing receptor pathways. PECAM-1 activation also inhibited platelet protein tyrosine phosphorylation stimulated by convulxin and thrombin; this was accompanied by inhibition of the mobilization of calcium from intracellular stores. These data suggest that PECAM-1 may play a role in the regulation of platelet function in vivo.

Antibodies↗
PubMed2006

Specificities of platelet autoantibodies and platelet activation in lupus anticoagulant patients: a relation to their history of thromboembolic disease.

Lupus anticoagulants (LA) prolong in vitro phospholipid-dependent coagulation tests, but are associated with thromboembolic disease (TE). However, a subgroup of individuals with LA has no TE, and it is therefore desirable to distinguish those at risk for TE from those without. Whether platelets have a primary role in the development of TE is not clear yet. We determined platelet autoantibodies to identify a specific platelet target which is associated with platelet activation in 97 patients with a long history of detectable LA, 65 patients with TE (LA/TE+), and 32 individuals without TE (LA/TE-). Thrombocytopenia was more common in the LA/TE- than in the LA/TE+ group (P < 0.05). Both groups had platelet antibodies, but the frequency of antibodies was lower in LA/TE+ than LA/TE- patients (P < 0.01), who had higher antibody titres against glycoprotein IIb/IIIa and glycoprotein Ib/IX (P < 0.05). Also, their platelets were more activated, as determined by PAC-1 binding (P < 0.01). These differences were also noted if patients with arterial thrombosis were evaluated separately. These findings in LA/TE- individuals were similar to those in patients with chronic autoimmune thrombocytopenia. However, there was no autoantibody target identifiable to distinguish between LA/TE- from LA-TE+ individuals. We therefore conclude that the presence of platelet antibodies, even if associated with platelet activation, is not sufficient to dispose LA patients to thromboembolic disease.

Adolescent↗