The relationship between shape change and release reaction of lectin-stimulated platelets. Effects of cytochalasin B and prostaglandin E1

SummaryThe increase of the cytoplasmic Ca-concentration plays a central role in the initiation of platelet activation. Four kinds of movements of Ca-ions are presumed to occur during this process: a) Ca-ions liberated from membranes induce the rapid shape change, b) Vesicular organelles release Ca-ions into the cytoplasm which initiate the release reaction, c) The storage organelles called dense bodies, secrete their contents including Ca-ions to the outside during the release reaction, d) At the same time a rearrangement of the plasma membrane occurs, resulting in an increase in its permeability for Ca-ions as well as in an increase in the number of Ca-binding sites.Since most processes occurring during platelet activation are reversible, the platelet must be equipped with a mechanism which removes Ca-ions from the cytoplasm. A vesicular fraction obtained from homogenized platelets indeed accumulates Ca actively. This Ca- pump is stimulated by cyclic AMP and protein kinase; it may be involved in the recovery of platelets after activation.It becomes increasingly clear that the various manifestations of platelet activation are triggered by a rise in the cytoplasmic Ca2+-concentration. The evidence for this and possible mechanisms involved are discussed in some detail in the contributions by Detwiler et al. and by Gerrard and White to this symposium. In this article we shall discuss four different types of mobilization of Ca-ions which occur in the course of the activation of platelets. In addition, at least one transport step involved in the removal of Ca2+ must occur during relaxation of activated platelets.

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The relationship between Prostaglandin E1 applied area and flap survival rate

British Journal of Plastic Surgery ◽

10.1016/0007-1226(92)90211-f ◽

1992 ◽

Vol 45 (6) ◽

pp. 465-468 ◽

Cited By ~ 17

Author(s):

Yukimasa Sawada ◽

Takatoshi Yotsuyanagi ◽

Ichiro Hatayama ◽

Ken Sone

Keyword(s):

Survival Rate ◽

Prostaglandin E1 ◽

Flap Survival ◽

The Relationship

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ADP-induced inhibition of von Willebrand factor-mediated platelet agglutination

American Journal of Physiology-Legacy Content ◽

10.1152/ajplegacy.1976.230.5.1406 ◽

1976 ◽

Vol 230 (5) ◽

pp. 1406-1410 ◽

Cited By ~ 22

Author(s):

RA Grant ◽

MB Zucker ◽

J McPherson

Keyword(s):

Human Plasma ◽

Von Willebrand Factor ◽

Prostaglandin E1 ◽

Platelet Rich Plasma ◽

Shape Change ◽

Normal Human ◽

Von Willebrand ◽

Willebrand Factor ◽

Formalin Fixed ◽

Platelet Shape

Human plasma von Willebrand factor (vWF) plus the antibiotic ristocetin, or bovine or porcine vWF alone, agglutinates platelets in either normal human ethylenediaminetetraacetate (EDTA)-treated citrated platelet-rich plasma (PRP) or citrated PRP from patients with the congenital platelet defect thrombasthenia. The prior addition of 1-10 muM ADP, which causes platelet shape change but not aggregation under these conditions, inhibited vWF-mediated agglutination. Inhibition was prevented by 200 muM ATP. Addition of ADP caused prompt reversal of established vWF-mediated agglutination, which resumed when the ADP was enzymatically removed. EDTA-treated, Formalin-fixed, washed normal platelets also underwent vWF-mediated agglutination. ADP was inhibitory only when added before fixation. Epinephrine (40 muM), prostaglandin E1 (7 muM), or serotonin (2 muM) added before fixation caused slight to moderate inhibition but always less than ADP. Platelets from blood chilled before fixation were fully active. Platelets fixed in freshly prepared PRP did not agglutinate as well as those fixed after incubation of PRP, probably because centrifugation exposes the platelets to ADP. It concluded that ADP causes a reversible decrease in the accessibility of the membrane receptor to vWF.

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Platelet Shape Change Evoked by Selective Activation of P2X1 Purinoceptors with α,β-Methylene ATP

Thrombosis and Haemostasis ◽

10.1055/s-0037-1615684 ◽

2001 ◽

Vol 85 (02) ◽

pp. 303-308 ◽

Cited By ~ 100

Author(s):

Michael Rolf ◽

Charles Brearley ◽

Martyn Mahaut-Smith

Keyword(s):

Inhibitory Action ◽

Light Transmission ◽

Shape Change ◽

Second Messengers ◽

Receptor Activation ◽

Selective Activation ◽

The Relationship ◽

Shape Changes ◽

Platelet Shape

SummarySimultaneous measurements of [Ca2+]i and light transmission were used to examine the relationship between P2X1 receptor activation and functional platelet responses. The P2X1 agonist α,β-MeATP evoked a transient [Ca2+]i increase and a reversible decrease in light transmission; both responses required external Ca2+ and the nucleotidase apyrase. The transmission response was due to shape change only, verified by scanning electron microscopy and insensitivity to Reopro, a GPIIbIIIa antagonist. α,β-MeATP stimulated smaller shape changes than ADP, however P2X1 responses had a lifespan of <2 h following resuspension in saline and may be considerably larger in vivo. A peak [Ca2+]i increase of >50 nM was required for detectable shape change. Overlap of concentration-response relationships for α,β-MeATP-evoked [Ca2+]i and shape change suggests that other second messengers are not involved. Therefore, the physiological P2X1 agonist ATP can contribute to platelet activation, in contrast to its previously described inhibitory action at metabotropic platelet purinoceptors.

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The effect of aggregation and release on platelet prothrombin- converting activity

Blood ◽

10.1182/blood.v54.3.659.659 ◽

1979 ◽

Vol 54 (3) ◽

pp. 659-672 ◽

Cited By ~ 12

Author(s):

AC Cox ◽

P Inyangetor ◽

CT Esmon ◽

BN White

Keyword(s):

Platelet Aggregation ◽

Prostaglandin E1 ◽

Dense Body ◽

Shape Change ◽

Factor Xa ◽

Dense Bodies ◽

Activated Platelets ◽

Induced Aggregation ◽

Body Secretion ◽

Platelet Shape

Abstract Platelets provide a procoagulant activity for the conversion of prothrombin to thrombin during normal hemostatis. This activity designated as platelet prothrombin-converting activity (PPCA) was monitored as rate of thrombin production in a two-stage assay using gel- filtered bovine platelets, factor Xa, and prothrombin. Expression of PPCA was not associated with ADP-induced release or platelet shape change but was associated with aggregation. Release of the contents of dense bodies, measured by release of 14C-5-hydroxytryptamine, was not required for expression of PPCA during platelet aggregation. During the PPCA assay, 5-hydroxytrypamine was released, but only after onset of thrombin production. Furthermore, the release of 5-hydroxytryptamine was retarded during the assay by the addition of 2 mM theophylline and 100 nM prostaglandin E1 without a comparable reduction in PPCA. In addition, 125I-factor-Xa was bound in greater amounts to platelets (aspirin-treated) after ADP-induced aggregation (without detectable release) than to unactivated control platelets. Finally, the PPCA of the ADP-activated platelets was saturated with respect to factors Xa and Va at less than 1 nM concentrations, indicating that the aggregation induced by ADP leads to the exposure of specific procoagulant sites by some process other than dense body secretion.

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Part of the activating cross-linked immunoglobulin G is internalized by human platelets to sites not accessible for enzymatic digestion

Blood ◽

10.1182/blood.v67.1.12.12 ◽

1986 ◽

Vol 67 (1) ◽

pp. 12-18 ◽

Cited By ~ 14

Author(s):

MO Spycher ◽

UE Nydegger

Keyword(s):

Immunoglobulin G ◽

Prostaglandin E1 ◽

Cytochalasin B ◽

Enzymatic Digestion ◽

Human Platelets ◽

Serotonin Release ◽

Antimycin A ◽

Energy Dependent ◽

Increased Susceptibility ◽

Pronase Treatment

Abstract The differential uptake of tritium-labeled immunoglobulin G (IgG) cross- linked with bisdiazonium-benzidine (BDB) (3H-BDB-IgG) by washed, pooled human platelets to sites inaccessible to pronase digestion was tested. Up to 52% of the 3H-BDB-IgG associated with platelets at 37 degrees C resisted pronase treatment, whereas only 23% of the cross-linked IgG associated with platelets at 4 degrees C, or at 37 degrees C but in the presence of deoxyglucose/antimycin A, remained refractory to pronase. This effect was not due to platelet agglutination. Pronase resistance reached a maximum after a 60-minute incubation period at 37 degrees C. With increasing 3H-BDB-IgG input, both the total cross-linked IgG associated with platelets and the fraction resistant to pronase digestion approached saturation at 4 degrees C, but not at 37 degrees C. The proportion of 3H-BDB-IgG bound to platelets at 4 degrees C that was resistant to pronase treatment increased by 13% within five minutes of warming the platelets to 37 degrees C. Pretreatment of platelets with 10 mmol/L acetylsalicylic acid (or 10 mumol/L prostaglandin E1) prior to the addition of 3H-BDB-IgG led to a 74% (95%) inhibition of the 3H-BDB-IgG-induced 14C-serotonin release, but to only a 44% (49%) inhibition of pronase-digestible bound ligand. In contrast, pretreatment with 10 mumol/L cytochalasin B led to a mere 17% reduction of 14C-serotonin release, whereas acquisition of resistance to pronase digestion by the bound 3H-BDB-IgG was inhibited by 90%. Incubation of platelets at 37 degrees C with 3H-BDB-IgG and removal of unbound material prior to the addition of prostaglandin E1 or deoxyglucose/antimycin A had little effect on the susceptibility of platelet-associated 3H-BDB-IgG to pronase, whereas the addition of cytochalasin B to 3H-BDB-IgG-treated platelets resulted in greatly increased susceptibility of the platelet-associated ligand to pronase. Thus, after binding, 3H-BDB-IgG becomes transferred in an energy- dependent process to pronase-resistant cellular sites, most likely to the open canalicular system.

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Dibucaine-Activated Platelet Protease(S) Digests GPIb-EDTA Only Partially Inhibits

10.1055/s-0038-1652285 ◽

1981 ◽

Author(s):

Barry S Coller

Keyword(s):

Protease Activity ◽

Shape Change ◽

Initial Slope ◽

Short Term ◽

Release Reaction ◽

Calcium Dependent ◽

Total Inhibition ◽

Von Willebrand ◽

Willebrand Factor ◽

Ristocetin Cofactor

Local anesthetics have been reported to have several effects on platelets including: 1) inhibition of shape change, the release reaction and aggregation induced by several agents 2) dissolution of cytoskeletal structures and 3) activation of a protease activity which digests proteins of Mr 250,000 and 230,000. It was speculated that the last effect may be due to the Ca++-dependent protease previously described in platelets. I have recently observed that incubation of platelets with 1 mM dibucaine for brief periods of time (15-30 min) resulted in 50-100% enhancement of the initial slope of agglutination induced by either ristocetin or bovine von Willebrand factor (vWf) despite inhibiting the release reaction. If the platelets were formaldehyde-fixed after brief incubation, they also showed the enhancement (control 23 ± 3 U/min (mean ± SEM) and dibucaine 47 ± 6, p < 0.01, n = 7 for ristocetin and 36 ± 7 vs. 65 ± 5, p < 0.01, n= 6 for bovine vWf). Using these platelets, the ristocetin cofactor assay was more sensitive and yet equally specific. If, however, fresh platelets were incubated with dibucaine for > 1 hour, both ristocetin and bovine vWf-induced agglutination became progressively inhibited with almost total inhibition after several hours. This decrease in agglutination was associated with a progressive loss of a glycoprotein with the characteristics of GpIb (Mr 165,000 unreduced and 135,000 reduced) as judged by PAS-stained polyacrylamide gels. The presence of the platelet-impenetrable calcium chelator EDTA (10 mM) only partially prevented the decrease in agglutinability and loss of Gplb. I conclude that: 1. Short term incubation with dibucaine may improve the ristocetin cofactor assay. 2. A dibucaine-activated protease(s) appears to digest Gplb, probably accounting for the decreased agglutination with long incubations. 3. If operating from the outside of the platelet after leakage, at least some of the protease activity is not calcium dependent.

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Cdc42/Rac1-dependent activation of the p21-activated kinase (PAK) regulates human platelet lamellipodia spreading: implication of the cortical-actin binding protein cortactin

Blood ◽

10.1182/blood.v100.13.4462 ◽

2002 ◽

Vol 100 (13) ◽

pp. 4462-4469 ◽

Cited By ~ 113

Author(s):

Catherine Vidal ◽

Blandine Geny ◽

Josiane Melle ◽

Martine Jandrot-Perrus ◽

Michaëla Fontenay-Roupie

Keyword(s):

Shape Change ◽

Binding Protein ◽

Actin Binding ◽

Cortical Actin ◽

Toxin B ◽

Actin Reorganization ◽

Actin Binding Protein ◽

Direct Binding ◽

P21 Activated Kinase ◽

The Relationship

Platelet activation by thrombin or thrombin receptor-activating peptide (TRAP) results in extensive actin reorganization that leads to filopodia emission and lamellae spreading concomitantly with activation of the Rho family small G proteins, Cdc42 and Rac1. Evidence has been provided that direct binding of Cdc42-guanosine triphosphate (GTP) and Rac1-GTP to the N-terminal regulatory domain of the p21-activated kinase (PAK) stimulates PAK activation and actin reorganization. In the present study, we have investigated the relationship between shape change and PAK activation. We show that thrombin, TRAP, or monoclonal antibody (MoAb) anti-FcγRIIA IV.3 induces an activation of Cdc42 and Rac1. The GpVI ligand, convulxin (CVX), that forces platelets to lamellae spreading efficiently activates Rac1. Thrombin, TRAP, MoAb IV.3, and CVX stimulate autophosphorylation and kinase activity of PAK. Inhibition of Cdc42 and Rac1 with clostridial toxin B inhibits PAK activation and lamellae spreading. The cortical-actin binding protein, p80/85 cortactin, is constitutively associated with PAK in resting platelets and dissociates from PAK after thrombin stimulation. Inhibition of PAK autophosphorylation by toxin B prevents the dissociation of cortactin. These results suggest that Cdc42/Rac1-dependent activation of PAK may trigger early platelet shape change, at least in part through the regulation of cortactin binding to PAK.

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Sodium arachidonate can induce platelet shape change and aggregation which are independent of the release reaction

Science ◽

10.1126/science.1273582 ◽

1976 ◽

Vol 192 (4243) ◽

pp. 1011-1012 ◽

Cited By ~ 86

Author(s):

R. Kinlough-Rathbone ◽

H. Reimers ◽

J. Mustard ◽

M. Packham

Keyword(s):

Shape Change ◽

Release Reaction ◽

Platelet Shape

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