Role of the PAR4 Thrombin Receptor in Stabilizing Platelet-platelet Aggregates as Revealed by a Patient with Hermansky-Pudlak Syndrome

2002 ◽  
Vol 87 (04) ◽  
pp. 722-727 ◽  
Author(s):  
Lidija Covic ◽  
Christopher Singh ◽  
Hedy Smith ◽  
Athan Kuliopulos
Keyword(s):  
Thrombin Receptor ◽  
Aggregate Formation ◽  
Dense Granules ◽  
Normal Individuals ◽  
Platelet Aggregate ◽  
Adp Release ◽  
Platelet Aggregates ◽  
Adp Receptor ◽  
Hermansky Pudlak Syndrome

SummaryIndividuals with Hermansky-Pudlak Syndrome (HPS) lack platelet dense granules and have no ADP-autocrine response. Despite these platelet deficiencies, HPS patients exhibit a surprisingly mild bleeding phenotype. We hypothesize that activation of the PAR4 thrombin receptor compensates for the lack of an ADP-autocrine response by the P2Y12 ADP receptor in individuals with HPS. Here, we determine that PAR4 activation by thrombin occurs well after ADP release from dense granules in normal individuals. However, the signal from PAR4 stabilizes platelet-platelet aggregate formation in the absence of P2Y12 activation by ADP. Thus, the strong signal emanating from PAR4 during platelet aggregation would provide an explanation for the mild bleeding diathesis of HPS.

scholarly journals DDAVP enhances platelet adherence and platelet aggregate growth on human artery subendothelium

Blood ◽  
1984 ◽  
Vol 64 (1) ◽  
pp. 229-236 ◽  
Author(s):  
KS Sakariassen ◽  
M Cattaneo ◽  
A v.d. Berg ◽  
ZM Ruggeri ◽  
PM Mannucci ◽  
...  
Keyword(s):  
Bleeding Time ◽  
Close Association ◽  
Aggregate Formation ◽  
Human Artery ◽  
Primary Hemostasis ◽  
Platelet Adherence ◽  
Normal Individuals ◽  
Before And After ◽  
Platelet Aggregate ◽  
Aggregate Growth

Abstract The effect of intravenous 1-deamino (8-D-arginine)vasopressin (DDAVP) administration on platelet interaction with human artery subendothelium was investigated with flowing blood from five normal individuals and 12 patients with von Willebrand's disease (vWD). Three of the patients were diagnosed as vWD subtype I, four as subtype IIa, and five as subtype IIb. DDAVP administration to normals enhanced platelet adherence, in parallel with increasing plasma levels of factor VIII- related antigen ( FVIIIR :Ag) and ristocetin cofactor activity ( FVIIIR :RCF). Platelet aggregate formation was transiently increased within 90 minutes. Platelet adherence in patient blood before DDAVP infusion was subnormal. In patients with subtype I, administration of DDAVP normalized the bleeding time, enhanced the platelet adherence, and transiently improved the platelet aggregate formation. The platelet adherence was more corrected than would have been expected on the basis of the FVIIIR :Ag and FVIIIR :RCF levels. In patients with subtype IIa, infusion of DDAVP increased the FVIIIR :Ag levels approximately threefold, without affecting the FVIIIR :RCF levels, and in only two of four patients was a transiently enhanced platelet adherence with a corresponding shortening of the bleeding time observed. In patients with subtype IIb, administration of DDAVP increased the FVIIIR :Ag levels about threefold and the FVIIIR :RCF levels five to tenfold, but decreased the platelet adherence significantly. The bleeding time values were not normalized. A close association between the bleeding time values and corresponding platelet adherence values before and after DDAVP infusion was observed. Normalization of the bleeding time was paralleled with normalization of platelet adherence. We conclude that DDAVP improves the primary hemostasis by causing enhanced FVIII- vWF-mediated platelet adherence. DDAVP has little or no effect on the bleeding time in patients with subtype IIa and subtype IIb, because the platelet adherence is not normalized.

Continuous signaling via PI3K isoforms β and γ is required for platelet ADP receptor function in dynamic thrombus stabilization

Blood ◽  
2006 ◽  
Vol 108 (9) ◽  
pp. 3045-3052 ◽  
Author(s):  
Judith M. E. M. Cosemans ◽  
Imke C. A. Munnix ◽  
Reinhard Wetzker ◽  
Regine Heller ◽  
Shaun P. Jackson ◽  
...  
Keyword(s):  
Thrombus Formation ◽  
G Protein Coupled Receptors ◽  
Platelet Aggregate ◽  
Platelet Aggregates ◽  
Adp Receptor ◽  
Phosphoinositide 3 Kinase ◽  
Novel Concept ◽  
Induced Aggregation ◽  
G Protein Coupled ◽  
Intrinsic Dynamic

Abstract Signaling from collagen and G protein–coupled receptors leads to platelet adhesion and subsequent thrombus formation. Paracrine agonists such as ADP, thromboxane, and Gas6 are required for platelet aggregate formation. We hypothesized that thrombi are intrinsically unstable structures and that their stabilization requires persistent paracrine activity and continuous signaling, maintaining integrin αIIbβ3 activation. Here, we studied the disassembly of human and murine thrombi formed on collagen under high shear conditions. Platelet aggregates rapidly disintegrated (1) in the absence of fibrinogen-containing plasma; (2) by blocking or inhibiting αIIbβ3; (3) by blocking P2Y12 receptors; (4) by suppression of phosphoinositide 3-kinase (PI3K) β. In murine blood, absence of PI3Kγ led to formation of unstable thrombi, leading to dissociation of multiplatelet aggregates. In addition, blocking PI3Kβ delayed initial thrombus formation and reduced individual platelet-platelet contact. Similarly without flow, agonist-induced aggregation was reversed by late suppression of P2Y12 or PI3K isoforms, resulting in single platelets that had inactivated αIIbβ3 and no longer bound fibrinogen. Together, the data indicate that continuous outside-in signaling via P2Y12 and both PI3Kβ and PI3Kγ isoforms is required for perpetuated αIIbβ3 activation and maintenance of a platelet aggregate. This novel concept of intrinsic, dynamic thrombus instability gives possibilities for the use of antiplatelet therapy.

scholarly journals Intercellular calcium communication regulates platelet aggregation and thrombus growth

2003 ◽  
Vol 160 (7) ◽  
pp. 1151-1161 ◽  
Author(s):  
Warwick S. Nesbitt ◽  
Simon Giuliano ◽  
Suhasini Kulkarni ◽  
Sacha M. Dopheide ◽  
Ian S. Harper ◽  
...  
Keyword(s):  
Platelet Adhesion ◽  
Purinergic Receptor ◽  
Cytosolic Calcium ◽  
Calcium Flux ◽  
Aggregate Formation ◽  
Signaling Mechanism ◽  
Activated Platelets ◽  
Platelet Aggregate ◽  
Platelet Aggregates

The ability of platelets to form stable adhesion contacts with other activated platelets (platelet cohesion or aggregation) at sites of vascular injury is essential for hemostasis and thrombosis. In this study, we have examined the mechanisms regulating cytosolic calcium flux during the development of platelet–platelet adhesion contacts under the influence of flow. An examination of platelet calcium flux during platelet aggregate formation in vitro demonstrated a key role for intercellular calcium communication (ICC) in regulating the recruitment of translocating platelets into developing aggregates. We demonstrate that ICC is primarily mediated by a signaling mechanism operating between integrin αIIbβ3 and the recently cloned ADP purinergic receptor P2Y12. Furthermore, we demonstrate that the efficiency by which calcium signals are propagated within platelet aggregates plays an important role in dictating the rate and extent of thrombus growth.

scholarly journals DDAVP enhances platelet adherence and platelet aggregate growth on human artery subendothelium

Blood ◽  
1984 ◽  
Vol 64 (1) ◽  
pp. 229-236 ◽  
Author(s):  
KS Sakariassen ◽  
M Cattaneo ◽  
A v.d. Berg ◽  
ZM Ruggeri ◽  
PM Mannucci ◽  
...  
Keyword(s):  
Bleeding Time ◽  
Close Association ◽  
Aggregate Formation ◽  
Human Artery ◽  
Primary Hemostasis ◽  
Platelet Adherence ◽  
Normal Individuals ◽  
Before And After ◽  
Platelet Aggregate ◽  
Aggregate Growth

The effect of intravenous 1-deamino (8-D-arginine)vasopressin (DDAVP) administration on platelet interaction with human artery subendothelium was investigated with flowing blood from five normal individuals and 12 patients with von Willebrand's disease (vWD). Three of the patients were diagnosed as vWD subtype I, four as subtype IIa, and five as subtype IIb. DDAVP administration to normals enhanced platelet adherence, in parallel with increasing plasma levels of factor VIII- related antigen ( FVIIIR :Ag) and ristocetin cofactor activity ( FVIIIR :RCF). Platelet aggregate formation was transiently increased within 90 minutes. Platelet adherence in patient blood before DDAVP infusion was subnormal. In patients with subtype I, administration of DDAVP normalized the bleeding time, enhanced the platelet adherence, and transiently improved the platelet aggregate formation. The platelet adherence was more corrected than would have been expected on the basis of the FVIIIR :Ag and FVIIIR :RCF levels. In patients with subtype IIa, infusion of DDAVP increased the FVIIIR :Ag levels approximately threefold, without affecting the FVIIIR :RCF levels, and in only two of four patients was a transiently enhanced platelet adherence with a corresponding shortening of the bleeding time observed. In patients with subtype IIb, administration of DDAVP increased the FVIIIR :Ag levels about threefold and the FVIIIR :RCF levels five to tenfold, but decreased the platelet adherence significantly. The bleeding time values were not normalized. A close association between the bleeding time values and corresponding platelet adherence values before and after DDAVP infusion was observed. Normalization of the bleeding time was paralleled with normalization of platelet adherence. We conclude that DDAVP improves the primary hemostasis by causing enhanced FVIII- vWF-mediated platelet adherence. DDAVP has little or no effect on the bleeding time in patients with subtype IIa and subtype IIb, because the platelet adherence is not normalized.

Glycoprotein VI is not a Functional Platelet Receptor for Fibrin Formed in Plasma or Blood

2020 ◽  
Vol 120 (06) ◽  
pp. 977-993 ◽  
Author(s):  
Danmei Zhang ◽  
Mariam Ebrahim ◽  
Kristin Adler ◽  
Xavier Blanchet ◽  
Janina Jamasbi ◽  
...  
Keyword(s):  
Platelet Adhesion ◽  
Aggregate Formation ◽  
Glycoprotein Vi ◽  
Platelet Receptor ◽  
Platelet Aggregate ◽  
Flowing Blood ◽  
Coated Surfaces ◽  
Long Fibers ◽  
Collagen Receptor

AbstractGlycoprotein VI (GPVI), a platelet collagen receptor, is crucial in mediating atherothrombosis. Besides collagen, injured plaques expose tissue factor (TF) that triggers fibrin formation. Previous studies reported that GPVI also is a platelet receptor for fibrinogen and fibrin. We studied the effect of anti-GPVI antibodies and inhibitors of GPVI signaling kinases (Syk and Btk) on platelet adhesion and aggregate formation onto immobilized fibrinogen and different types of fibrin under arterial flow conditions. Fibrin was prepared from isolated fibrinogen (“pure fibrin”), recombinant fibrinogen (“recombinant fibrin”), or generated more physiologically from endogenous fibrinogen in plasma (“plasma fibrin”) or by exposing TF-coated surfaces to flowing blood (“blood fibrin”). Inhibition of GPVI and Syk did not inhibit platelet adhesion and aggregate formation onto fibrinogen. In contrast anti-GPVI antibodies, inhibitors of Syk and Btk and the anti-GPIb antibody 6B4 inhibited platelet aggregate formation onto pure and recombinant fibrin. However, inhibition of GPVI and GPVI signaling did not significantly reduce platelet coverage of plasma fibrin and blood fibrin. Plasma fibrin contained many proteins incorporated during clot formation. Advanced optical imaging revealed plasma fibrin as a spongiform cushion with thicker, knotty, and long fibers and little activation of adhering platelets. Albumin intercalated in plasma fibrin fibers left only little space for platelet attachment. Pure fibrin was different showing a dense mesh of thin fibers with strongly activated platelets. We conclude that fibrin formed in plasma and blood contains plasma proteins shielding GPVI-activating epitopes. Our findings do not support a role of GPVI for platelet activation by physiologic fibrin.

Circulating Platelet Aggregates in Normal Pregnancy and Rpeeclampsia

1979 ◽  
Author(s):  
I. Rákóczi ◽  
F. Tallián ◽  
I. Cseh ◽  
I. Gáti
Keyword(s):  
Pregnant Women ◽  
Platelet Activation ◽  
Normal Pregnancy ◽  
Third Trimester ◽  
The Third ◽  
Platelet Aggregate ◽  
Circulating Platelet Aggregates ◽  
Platelet Aggregates ◽  
The Mean

Circulating platelet aggregates have been observed in various thromboembolic states. It is known that severe preeclamsia is associated with features of intravascular coagulation. To evaluate the role of platelets in this disorder we have determined circulating platelet aggregates in 10 patients with severe preeclampsia, in 30 patients in the third trimester of uncomplicated pregnancies and in 35 healthy nonpregnant volunteers. Platelet aggregate ratio /P.A.R./ was measured by a modification of a method described by Wu and Hoak, The mean P.A.R. of severe preeclamptic patients /0.732 ± 0,063 SEM/ was significantly lower than that of the uncomplicated pregnant women /0,860 ± 0,052 SEM/ and of the nonpregnant volunteers /0.880 ± 0,061 SEM/.The results indicate that severe preeclamptic patients have increased levels of circulating platelet aggregates and platelet activation is a feature of preeclampsia.

Effect of Dipyridamole on Spontaneous Platelet Aggregation in Whole Blood Decreases with the Time After Venepuncture: Evidence for the Role of ADP

1987 ◽  
Vol 58 (02) ◽  
pp. 744-748 ◽  
Author(s):  
A R Saniabadi ◽  
G D O Lowe ◽  
J C Barbenel ◽  
C D Forbes
Keyword(s):  
Platelet Aggregation ◽  
Correlation Coefficient ◽  
Whole Blood ◽  
Blood Platelet ◽  
Inhibitory Effect ◽  
Time Interval ◽  
Adp Receptor ◽  
Spontaneous Platelet Aggregation

SummarySpontaneous platelet aggregation (SPA) was studied in human whole blood at 3, 5, 10, 20, 30, 40 and 60 minutes after venepuncture. Using a whole blood platelet counter, SPA was quantified by measuring the fall in single platelet count upon rollermixing aliquots of citrated blood at 37° C. The extent of SPA increased with the time after venepuncture, with a correlation coefficient of 0.819. The inhibitory effect of dipyridamole (Dipy) on SPA was studied: (a) 10 μM at each time interval; (b) 0.5-100 μM at 3 and 30 minutes and (c) 15 μM in combination with 100 μM adenosine, 8 μM 2-chloroadenosine (2ClAd, an ADP receptor blocker) and 50 μM aspirin. There was a rapid decrease in the inhibitory effect of Dipy with the time after venepuncture; the correlation coefficient was -0.533. At all the concentrations studied, Dipy was more effective at 3 minutes than at 30 minutes after venepuncture. A combination of Dipy with adenosine, 2ClAd or aspirin was a more effective inhibitor of SPA than either drug alone. However, when 15 μM Dipy and 10 μM Ad were added together, the inhibitory effect of Dipy was not increased significantly, suggesting that Dipy inhibits platelet aggregation independent of Ad. The increase in SPA with the time after venepuncture was abolished when blood was taken directly into the anticoagulant containing 5 μM 2ClAd. It is suggested that ADP released from the red blood cells is responsible for the increased platelet aggregability with the time after venepuncture and makes a serious contribution to the artifacts of in vitro platelet function studies.

In Vivo Effect of Suloctidil as an Antiplatelet Agent

1979 ◽  
Vol 41 (03) ◽  
pp. 465-474 ◽  
Author(s):  
Marcia R Stelzer ◽  
Thomas S Burns ◽  
Robert N Saunders
Keyword(s):  
Ex Vivo ◽  
Antiplatelet Agent ◽  
Ratio Method ◽  
Platelet Aggregate ◽  
Permanent Effect ◽  
Platelet Aggregates ◽  
5 Ht Uptake ◽  
High Doses

SummaryThe relationship between the effects of suloctidil in vivo as an antiplatelet agent and in vitro as a modifier of platelet serotonin (5-HT) parameters was investigated. Suloctidil was found to be effective in reducing platelet aggregates formation in the retired breeder rat as determined using the platelet aggregate ratio method (PAR) with an ED50 of 16.1 mg/kg 24 hours post administration. In contrast to the hypothesis that 5-HT depletion is involved in the anti-aggregatory mechanism of suloctidil, no correlation was found between platelet 5- HT content and this antiplatelet activity. Reduction of platelet 5-HT content required multiple injections of high doses (100 mg/kg/day) of suloctidil. Suloctidil administration for 8 days at 100 mg/kg/day, which lowered platelet 5-HT content by 50%, resulted in no permanent effect on ex vivo platelet 5-HT uptake or thrombin-induced release, nor alteration in the plasma 5-HT level. However, these platelets exhibited a short-lived, significant increase in percent leakage of 5-HT after 30 minutes of incubation. Therefore, suloctidil treatment at high doses may with time result in platelet 5-HT depletion, however this effect is probably not related to the primary anti-aggregatory activity of the drug.

Different Abilities of Thrombin Receptor Activating Peptide and Thrombin to Induce Platelet Calcium Rise and Full Release Reaction

1995 ◽  
Vol 74 (05) ◽  
pp. 1323-1328 ◽  
Author(s):  
Dominique Lasne ◽  
José Donato ◽  
Hervé Falet ◽  
Francine Rendu
Keyword(s):  
Essential Role ◽  
Calcium Influx ◽  
Dense Granule ◽  
Receptor Interaction ◽  
Thrombin Receptor ◽  
Release Reaction ◽  
External Calcium ◽  
Maximum Rise ◽  
Granule Release

SummarySynthetic peptides (TRAP or Thrombin Receptor Activating Peptide) corresponding to at least the first five aminoacids of the new N-terminal tail generated after thrombin proteolysis of its receptor are effective to mimic thrombin. We have studied two different TRAPs (SFLLR, and SFLLRN) in their effectiveness to induce the different platelet responses in comparison with thrombin. Using Indo-1/AM- labelled platelets, the maximum rise in cytoplasmic ionized calcium was lower with TRAPs than with thrombin. At threshold concentrations allowing maximal aggregation (50 μM SFLLR, 5 μM SFLLRN and 1 nM thrombin) the TRAPs-induced release reaction was about the same level as with thrombin, except when external calcium was removed by addition of 1 mM EDTA. In these conditions, the dense granule release induced by TRAPs was reduced by over 60%, that of lysosome release by 75%, compared to only 15% of reduction in the presence of thrombin. Thus calcium influx was more important for TRAPs-induced release than for thrombin-induced release. At strong concentrations giving maximal aggregation and release in the absence of secondary mediators (by pretreatment with ADP scavengers plus aspirin), SFLLRN mobilized less calcium, with a fast return towards the basal level and induced smaller lysosome release than did thrombin. The results further demonstrate the essential role of external calcium in triggering sustained and full platelet responses, and emphasize the major difference between TRAP and thrombin in mobilizing [Ca2+]j. Thus, apart from the proteolysis of the seven transmembrane receptor, another thrombin binding site or thrombin receptor interaction is required to obtain full and complete responses.

Increased Plasma Fibrinogen and Platelet-Aggregates in Type II Hyperlipoproteinaemia

1979 ◽  
Vol 42 (05) ◽  
pp. 1503-1507 ◽  
Author(s):  
G D O Lowe ◽  
Maureen M Drummond ◽  
Jane L H C Third ◽  
W F Bremner ◽  
C D Forbes ◽  
...  
Keyword(s):  
Plasma Lipids ◽  
Young Patients ◽  
Adult Life ◽  
Arterial Disease ◽  
Arterial Injury ◽  
Plasma Fibrinogen ◽  
Type Ii ◽  
Familial Type ◽  
Platelet Aggregate ◽  
Platelet Aggregates

SummaryPlasma fibrinogen and platelet-aggregates (method of Wu and Hoak) were measured in 21 patients with familial Type II hyperlipoproteinaemia and 21 matched control subjects. Patients with hyperlipoproteinaemia had increased levels of fibrinogen and platelet- aggregates (p<0.01). Young patients with hyperlipoproteinaemia had prematurely high fibrinogen levels, and the normal rise in fibrinogen during adult life was abolished. There were no statistically significant correlations within the patient group between fibrinogen, platelet-aggregates, and plasma lipids. High fibrinogen and platelet-aggregate levels may play a part in the development of the premature arterial disease associated with Type II hyperlipoproteinaemia, or may be markers of arterial injury.

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