Expression of intracellular fibrinogen on the surface of stimulated platelets

Courtois, Gilles; Ryckewaert, Jean Jacques; Woods, Virgil L.; Ginsberg, Mark H.; Plow, Edward F.; Marguerie, G

doi:10.1111/j.1432-1033.1986.tb09833.x

Cited by 35 publications

(14 citation statements)

References 26 publications

(5 reference statements)

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“…However, although inhibiting by more than 85% the binding of '251-fibrinogen to platelets, AP-2 does not prevent the surface expression of platelet fibrinogen upon thrombin stimulation (Legrand, C., unpublished results). A similar finding for platelet fibrinogen has also been observed by Courtois et al [43] using another antibody to GPIIb-IIIa complexes. Thus, these results do not preclude the possibility that thrombospondin is expressed through an interaction with fibrinogen bound to GPIIb-IIIa complexes at an early stage of the secretion process.…”

Section: Discussionsupporting

confidence: 87%

Use of a monoclonal antibody to measure the surface expression of thrombospondin following platelet activation

Legrand

Dubernard

Kieffer

et al. 1988

European Journal of Biochemistry

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The radiolabelled monoclonal antibody, 5G11, directed against native thrombospondin, has been used to assess the surface expression of secreted thrombospondin on human blood platelets. Emphasis has been placed on studying the role of fibrinogen in this process. Unstimulated platelets bound low amounts of 5G11 (about 2000 molecules/platelet). Binding increased 2‐fold and 5‐ 7‐fold after stimulation of platelets with ADP or thrombin (or ionophore A23187) respectively. Unstimulated platelets from patients deficient in α‐granule proteins (gray platelet syndrome) bound baseline levels of 5G11. However, binding was not increased after activation. Thrombospondin expression on thrombin‐stimulated normal platelets was for a large part divalent‐cation‐dependent and was not affected by AP‐2, a monoclonal antibody to GPIIb‐IIIa complexes. However, binding of 5G11 was some 50% lower when platelets were stimulated in the presence of Fab fragments of a polyclonal rabbit antibody to fibrinogen. This suggested either a direct binding of thrombospondin to surface‐bound fibrinogen or a steric inhibition due to a close proximity of the two proteins. The fact that binding of 5G11 was at the lower limit of the normal range to the stimulated platelets of an afibrinogenaemic patient specifically lacking detectable fibrinogen favoured the latter explanation. Thus, a major fibrinogen‐independent pathway for thrombospondin expression must exist.

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Section: Discussionsupporting

confidence: 87%

Use of a monoclonal antibody to measure the surface expression of thrombospondin following platelet activation

Legrand

Dubernard

Kieffer

et al. 1988

European Journal of Biochemistry

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“…Interestingly, the reduction in IAC-1 binding to ␣ 2 ␤ 1 on CVX-stimulated platelets in plasma where ␣ IIb ␤ 3 is blocked by aggrastat ( Figure 4A) was notably lower than the observed IAC-1 binding to ␣ 2 ␤ 1 on CVX-stimulated washed platelets ( Figure 1A). This suggests that autocrine fibrinogen, present in platelet ␣-granules (about 1 mg fibrinogen/10 10 platelets) and released upon platelet activation, 41 contributes to ␣ IIb ␤ 3 -mediated IAC-1 binding to washed platelets.…”

Section: Org Frommentioning

confidence: 99%

Activation of αIIbβ3 is a sufficient but also an imperative prerequisite for activation of α2β1 on platelets

Walle

Schoolmeester

Iserbyt

et al. 2006

Blood

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Platelet integrins ␣ 2 ␤ 1 and ␣ IIb ␤ 3 play critical roles in platelet adhesion and thrombus formation after vascular injury. On resting platelets, both integrins are in a low-affinity state. However, agonist stimulation results in conformational changes that enable ligand binding that can be detected with conformation dependent monoclonal antibodies (mAbs). By using such conformation-dependent mAbs, we could demonstrate that activation of integrin ␣ IIb ␤ 3 is not only sufficient, but also a prerequisite for ␣ 2 ␤ 1 activation. Compared with platelets in plasma, stimulation of washed platelets resulted in only a minor activation of ␣ 2 ␤ 1 , as detected with the activation-sensitive mAb IAC-1. Addition of fibrinogen to stimulated washed platelets greatly potentiated activation of this integrin. Also, treatment of ␣ IIb ␤ 3 with the ligand-mimetic peptide RGDS, resulting in outside-in signaling, led to a powerful ␣ 2 ␤ 1 activation, even in the absence of overall platelet activation, involving tyrosine kinase activity but no protein kinase C activation. The absolute necessity of ␣ IIb ␤ 3 for proper ␣ 2 ␤ 1 activation on platelets was demonstrated by using the ␣ IIb ␤ 3 antagonist aggrastat, which was able to completely abolish ␣ 2 ␤ 1 activation, both under static and flow conditions. In addition, analogous experiments with Glanzmann platelets lacking ␣ IIb ␤ 3 confirmed the indispensability of ␣ IIb IntroductionIntegrins are a large family of heterodimeric transmembrane receptors, each consisting of an ␣ and ␤ subunit, and are key effectors of cell growth, migration, differentiation, and survival. 1,2 Integrins possess the unique ability to signal across the plasma membrane in both directions, and since most integrins are not constitutively active, they are expressed on the cell surface as low-affinity receptors. When cells become activated, cytosolic proteins can bind to the cytoplasmic domains of integrins and as a consequence, the integrins are turned into their high-affinity state ("inside-out" signaling). In a process called "outside-in" signaling, ligand-binding of integrins then again activates intracellular pathways via their cytoplasmic domains, which are connected to the cytoskeleton and are associated with several intracellular signaling molecules. Important signaling modulators necessary for the generation of outside-in signals are members of the Src family protein tyrosine kinases, with c-Src as the initiating molecule due to its constitutive interaction with, for example, ␤ 3 integrins. 3 Such outside-in signals also result in conformational alterations of the integrin (also designated integrin activation), and subsequently these activated integrins can trigger another process of inside-out signaling. 4 This integrin activation upon conformational changes is often defined as an increase in integrin "affinity" for its ligand, and has been the topic of many studies. [4][5][6] In addition, cell activation also promotes clustering of integrins contributing to the "avidity" or "valency" regulation ...

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“…Platelet aggregation is provoked by the appearance on the cell surface of new proteins [2] and/or new phospholipids [3]. These factors come from plasma (coagulation factors [4], fibrinogen [5]) or from within the platelets (fibrinogen [6], different coagulation factors [7,8], phosphatidylserine [9]) depending on the organism. The present study demonstrates that a protein very similar to serum albumin is released from rabbit platelets under the influence of platelet activating factor (PAF-acether), with some evidence of a small contribution from cellular granules.…”

Section: Platelet-activating-factor-induced Serum-albumin Release Fromentioning

confidence: 99%

Platelet‐activating‐factor‐induced serum‐albumin release from rabbit platelets in the absence of calcium

Bette‐Bobillo¹,

Bienvenue²

1988

European Journal of Biochemistry

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Using a radioimmunoassay for rabbit-serum albumin, platelet-activating-factor-induced serum-albumin release by platelets was monitored under non-aggregating conditions. The four main results from this study are as follows. The ECS0 of the release was o f t h e same order of magnitude as the aggregation EC,, in the presence of calcium. The rclease took place within 2 min and was inhibited by BN 52021, which is a very specific inhibitor of the platelet-activating-factor-aggregating effect. Serum-albumin release was much greater than serotonin release.Many metabolic events occur after agonist binding to platelet membranes [I]. Besides secondary-messenger appearance in the cytosol, a fast and ATP-dependent actin polymerization causes the well known shape change of platelets. Platelet aggregation then occurs just after, or during, the release of the internal content of dense and a-granules, initiating the chain reaction activating new platelets.The components and properties of membranes are also modified. Platelet aggregation is provoked by the appearance on the cell surface of new proteins [2] and/or new phospholipids [3]. These factors come from plasma (coagulation factors [4], fibrinogen [5]) or from within the platelets (fibrinogen [6], different coagulation factors [7,8], phosphatidylserine [9]) depending on the organism. The present study demonstrates that a protein very similar to serum albumin is released from rabbit platelets under the influence of platelet activating factor (PAF-acether), with some evidence of a small contribution from cellular granules. MATERIALS A N D METHODS Radio-iodination of' rabbit-serum albuminRabbit-serum albumin (RSA, fraction V, Sigma) was radiolabeled to a specific activity of 76 Ci/mmol with "' I (New England Nuclear) by the chloramin-T method described by Greenwood et al. [lo]. The '2sI-labeled RSA (lz5I-RSA) preparation was subsequently diluted to 6 x 10' cpm/ml in NET buffer (150 mM NaC1, 5 m M EDTA, 50 mM Tris, and 0.02% sodium azide, pH 7.4) containing 1 mg/ml ovalbumin (Calbiochem-Behring ; NET-ovalbumin buffer). More than 97% of the radioactivity was acid-precipitable.

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Expression of intracellular fibrinogen on the surface of stimulated platelets

Cited by 35 publications

References 26 publications

Use of a monoclonal antibody to measure the surface expression of thrombospondin following platelet activation

Use of a monoclonal antibody to measure the surface expression of thrombospondin following platelet activation

Activation of αIIbβ3 is a sufficient but also an imperative prerequisite for activation of α2β1 on platelets

Platelet‐activating‐factor‐induced serum‐albumin release from rabbit platelets in the absence of calcium

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