Shear-Mediated Platelet Activation is Accompanied by Unique Alterations of Platelet Lipid Profile

Platelet activation by mechanical means such as shear stress, is a vital driver of thrombotic risk in implantable blood-contacting devices used in treatment of heart failure. Lipids are essential in platelets activation and have been studied following biochemical activation. However, little is known regarding lipid alterations occurring with mechanical-shear mediated platelet activation. Here, we determined if shear-activation of platelets induced lipidome changes that differ from those associated with biochemically-mediated platelet activation. We performed high-resolution lipidomic analysis on purified platelets from four healthy human donors. For each donor, we compared the lipidome of platelets that were non-activated or activated by shear, ADP, or thrombin treatment. We found that shear activation altered cell-associated lipids and led to the release of lipids into the extracellular environment. Shear-activated platelets released 21 phospholipids and sphingomyelins at levels statistically higher than platelets activated by biochemical stimulation. Many of the released phospholipids contained an arachidonic acid tail or were phosphatidylserine lipids, which have procoagulant properties. We conclude that shear-mediated activation of platelets alters the basal platelet lipidome. Further, these alterations differ and are unique in comparison to the lipidome of biochemically activated platelets. Our findings suggest that lipids released by shear-activated platelets may contribute to altered thrombosis in patients with implanted cardiovascular therapeutic devices.

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ULTRASTRUCTURAL INVESTIGATIONS ON THE MECHANISM OF “SHEAR-INDUCED PLATELET ACTIVATION”

10.1055/s-0038-1642845 ◽

1987 ◽

Author(s):

L J Wurzinger ◽

R Opitz ◽

H Schmid-Schönbein

Keyword(s):

Shear Stress ◽

Platelet Activation ◽

Platelet Rich Plasma ◽

Adenine Nucleotides ◽

Coagulation System ◽

Laminar Shear Stress ◽

Shear Forces ◽

Platelet Factor ◽

Membrane Defects ◽

Biochemical Activation

High shear forces are suspected to play a triggering role in the initiation of arterial thrombosis, by activating platelets and the coagulation system. In an earlier study a shear stress of 170 N/m2 acting for only 7 milliseconds (ms) on platelet rich plasma (PRP) was found to induce a significant increase in platelet factor 3 availability (Thromb. Haemost. 54: 381-386; 1985). To clarify the question whether platelets can be activated directly by mechanical forces in analogy to smooth muscle cells, electron micrographs of platelets subjected to laminar shear stress were analysed with morphometric methods. The level of activation of platelet suspensions was quantified by assessing 1) the elongation of platelet profiles giving a measure for the “flatness” of the discoid resting platelets, and 2) the centralization of granules.Exposure to a shear stress of 170 N/m2 for 113 ms leaves ca. 15 % of the platelets irreversibly damaged, featuring degenerative ballooning, with break-down of internal structure and cell membrane defects. The remaining 85 % appear typically activated with rounded shape, extension of pseudopods and centralization of granules. Addition of “ADP-scavengers” to the suspension medium totally changes the appearance of sheared platelets: still a comparable proportion of them has undergone irreversible degenerative changes, but the “surviving” population lacks ultrastructural signs of platelet activation. This is reflected in values of the morphometric parameters which are close to the level of unsheared control samples.It is therefore concluded that “shear-induced platelet activation” cannot be ascribed to a direct stimulating effect of shear forces, but rather to secondary biochemical activation by adenine nucleotides leaking from a small percentage of shear destroyed platelets. The latter process, however, requires a well stirred though undiluted environment, as it is provided in vortices and eddies.

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Soluble Platelet Release Factors as Biomarkers for Cardiovascular Disease

Frontiers in Cardiovascular Medicine ◽

10.3389/fcvm.2021.684920 ◽

2021 ◽

Vol 8 ◽

Author(s):

Gaukhar Baidildinova ◽

Magdolna Nagy ◽

Kerstin Jurk ◽

Philipp S. Wild ◽

Hugo ten Cate ◽

...

Keyword(s):

Platelet Activation ◽

Thrombus Formation ◽

Coagulation System ◽

Thrombotic Risk ◽

Activated Platelets ◽

Platelet Release ◽

Thrombotic Diseases ◽

Arterial Vascular Disease

Platelets are the main players in thrombotic diseases, where activated platelets not only mediate thrombus formation but also are involved in multiple interactions with vascular cells, inflammatory components, and the coagulation system. Although in vitro reactivity of platelets provides information on the function of circulating platelets, it is not a full reflection of the in vivo activation state, which may be relevant for thrombotic risk assessment in various disease conditions. Therefore, studying release markers of activated platelets in plasma is of interest. While this type of study has been done for decades, there are several new discoveries that highlight the need for a critical assessment of the available tests and indications for platelet release products. First, new insights have shown that platelets are not only prominent players in arterial vascular disease, but also in venous thromboembolism and atrial fibrillation. Second, knowledge of the platelet proteome has dramatically expanded over the past years, which contributed to an increasing array of tests for proteins released and shed from platelets upon activation. Identification of changes in the level of plasma biomarkers associated with upcoming thromboembolic events allows timely and individualized adjustment of the treatment strategy to prevent disease aggravation. Therefore, biomarkers of platelet activation may become a valuable instrument for acute event prognosis. In this narrative review based on a systematic search of the literature, we summarize the process of platelet activation and release products, discuss the clinical context in which platelet release products have been measured as well as the potential clinical relevance.

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Increased Thromboxane Biosynthesis in Essential Thrombocythemia

Thrombosis and Haemostasis ◽

10.1055/s-0038-1649916 ◽

1995 ◽

Vol 74 (05) ◽

pp. 1225-1230 ◽

Cited By ~ 71

Author(s):

Bianca Rocca ◽

Giovanni Ciabattoni ◽

Raffaele Tartaglione ◽

Sergio Cortelazzo ◽

Tiziano Barbui ◽

...

Keyword(s):

Platelet Count ◽

Platelet Activation ◽

Essential Thrombocythemia ◽

Therapeutic Strategy ◽

Low Dose ◽

Thrombotic Risk ◽

Dose Aspirin ◽

Gender And Age ◽

Low Dose Aspirin

SummaryIn order to investigate the in vivo thromboxane (TX) biosynthesis in essential thromboeythemia (ET), we measured the urinary exeretion of the major enzymatic metabolites of TXB2, 11-dehydro-TXB2 and 2,3-dinor-TXB2 in 40 ET patients as well as in 26 gender- and age-matched controls. Urinary 11-dehydro-TXB2 was significantly higher (p <0.001) in thrombocythemic patients (4,063 ± 3,408 pg/mg creatinine; mean ± SD) than in controls (504 ± 267 pg/mg creatinine), with 34 patients (85%) having 11-dehydro-TXB2 >2 SD above the control mean. Patients with platelet number <1,000 × 109/1 (n = 25) had significantly higher (p <0.05) 11 -dehydro-TXB2 excretion than patients with higher platelet count (4,765 ± 3,870 pg/mg creatinine, n = 25, versus 2,279 ± 1,874 pg/mg creatinine, n = 15). Average excretion values of patients aging >55 was significantly higher than in the younger group (4,784 ± 3,948 pg/mg creatinine, n = 24, versus 2,405 ± 1,885 pg/mg creatinine, n = 16, p <0.05). Low-dose aspirin (50 mg/d for 7 days) largely suppressed 11-dehydro-TXB2 excretion in 7 thrombocythemic patients, thus suggesting that platelets were the main source of enhanced TXA2 biosynthesis. The platelet count-corrected 11-dehydro-TXB2 excretion was positively correlated with age (r = 0.325, n = 40, p <0.05) and inversely correlated with platelet count (r = -0.381, n = 40, p <0.05). In addition 11 out of 13 (85%) patients having increased count-corrected 11-dehydro-TXB2 excretion, belonged to the subgroup with age >55 and platelet count <1,000 × 1099/1. We conclude that in essential thrombocythemia: 1) enhanced 11-dehydro-TXB2 excretion largely reflects platelet activation in vivo;2) age as well as platelet count appear to influence the determinants of platelet activation in this setting, and can help in assessing the thrombotic risk and therapeutic strategy in individual patients.

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A P-Selectin/CD62P Monoclonal Antibody (LYP-20), in Tandem with Flow Cytometry, Detects in vivo Activated Circulating Rat Platelets in Severe Vascular Trauma

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648952 ◽

1994 ◽

Vol 72 (05) ◽

pp. 745-749 ◽

Cited By ~ 7

Author(s):

Elza Chignier ◽

Maud Parise ◽

Lilian McGregor ◽

Caroline Delabre ◽

Sylvie Faucompret ◽

...

Keyword(s):

Monoclonal Antibody ◽

Flow Cytometry ◽

Platelet Activation ◽

Peripheral Blood ◽

Vascular Trauma ◽

Activated Platelets ◽

Group I ◽

Group Ii ◽

Rat Platelets

SummaryP-selectin, also known as CD62P, GMP140 or PADGEM, is present in platelet a-granules and endothelial cell Weibel-Palade bodies and is very rapidly expressed on the surface of these cells on activation. In this study, an anti P-selectin monoclonal antibody (LYP20) was used, in tandem with flow cytometry, to identify activated platelets at the site of induced vascular trauma or in peripheral blood. Moreover, electron microscopy was performed to characterize sites of vascular trauma and quantify the number of adhering platelets. The same induced vascular trauma was observed to result into animals responding in 2 different ways (Group I, Group II) following the degree of platelet activation. Five rats, out of 14 with induced vascular trauma, had more than half of their circulating platelets expressing P-selectin when drawn at the site of the trauma (67.4% ± 3.44) or in peripheral blood (78.5% ± 2.5) (Group I). In the remaining 9 animals a much smaller proportion of circulating platelets expressed P-selectin when assayed from trauma sites (18% ± 3.34) or in peripheral blood (18.0% ± 4.30) (Group II). Enhanced P-selectin expression by circulating platelets in Group I, compared to Group II, appears to be linked to the degree of activated platelets adhering at sites of trauma (171 ± 15 × 103 platelets versus 48 ± 31 × 103 platelets per mm2). In the 5 control animals, that were not operated on, platelets expressing P-selectin when drawn at the site of a mock trauma (7.0% ± 1.84) or in the peripheral blood (11.2% ± 3.30) showed little activation. In addition, no platelet adhesion was seen on the vascular bed of these animals. Results from this study show that analysis of P-selectin (CD62P) expression, in circulating platelets, is a valuable and rapid marker of platelet activation following severe vascular trauma induced in rats. However, activated platelets were not detected to the same extent in the peripheral blood of all animals having undergone vascular trauma. It is conceivable that platelets, depending on the degree of activation, may be actively sequestered in organs and prevented from circulating. Alternatively, P-selectin may be rapidly endocytosed, or not expressed, by activated circulating platelets depending on the type of agonists implicated in vivo activation.

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Immunological Evidence for Prothrombin in Human Platelets

Thrombosis and Haemostasis ◽

10.1055/s-0038-1661534 ◽

1986 ◽

Vol 55 (02) ◽

pp. 268-270

Author(s):

R J Alexander

Keyword(s):

Electrophoretic Mobility ◽

Platelet Activation ◽

Double Diffusion ◽

Human Platelets ◽

Secreted Proteins ◽

Platelet Surface ◽

Activated Platelets ◽

Similar Protein ◽

Diffusion Assay ◽

Supernatant Solution

SummaryAn attempt was made to isolate from plasma the platelet surface substrate for thrombin, glycoprotein V (GPV), because a GPV antigen was reported to be present in plasma (3). Plasma fractionation based on procedures for purification of GPV from platelets revealed a thrombin-sensitive protein with appropriate electrophoretic mobility. The protein was purified; an antiserum against it i) reacted with detergent-solubilized platelet proteins or secreted proteins in a double diffusion assay, ii) adsorbed a protein from the supernatant solution of activated platelets, and iii) inhibited thrombin-induced platelet activation, but the antiserum did not adsorb labeled GPV. The purified protein was immunochemically related to prothrombin rather than to GPV. Other antibodies against prothrombin were also able to adsorb a protein from platelets. It is concluded that 1) plasma does not contain appreciable amounts of GPV, and 2) platelets contain prothrombin or an immunochemically similar protein.

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Shear stress platelet activation test and “rebleeding” in patients undergoing cardiopulmonary bypass

Thrombosis Research ◽

10.1016/0049-3848(92)90037-b ◽

1992 ◽

Vol 68 (2) ◽

pp. 205-206 ◽

Cited By ~ 1

Author(s):

J. Butler

Keyword(s):

Shear Stress ◽

Cardiopulmonary Bypass ◽

Platelet Activation ◽

Activation Test

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Movement of Calcium Ions and their Role in the Activation of Platelets

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648654 ◽

1978 ◽

Vol 40 (02) ◽

pp. 212-218 ◽

Cited By ~ 60

Author(s):

P Massini ◽

R Käser-Glanzmann ◽

E F Lüscher

Keyword(s):

Plasma Membrane ◽

Platelet Activation ◽

Calcium Ions ◽

Binding Sites ◽

Shape Change ◽

Release Reaction ◽

Dense Bodies ◽

Activated Platelets ◽

Different Types ◽

Ca Ions

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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Multiscale Modeling of Flow Induced Thrombogenicity Using Dissipative Particle Dynamics and Molecular Dynamics

ASME 2013 2nd Global Congress on NanoEngineering for Medicine and Biology ◽

10.1115/nemb2013-93094 ◽

2013 ◽

Author(s):

Danny Bluestein ◽

João S. Soares ◽

Peng Zhang ◽

Chao Gao ◽

Seetha Pothapragada ◽

...

Keyword(s):

Molecular Dynamics ◽

Blood Flow ◽

Red Blood Cells ◽

Platelet Activation ◽

Blood Cells ◽

Dissipative Particle Dynamics ◽

Particle Dynamics ◽

Coagulation Cascade ◽

Shear Stresses ◽

Activated Platelets

The coagulation cascade of blood may be initiated by flow induced platelet activation, which prompts clot formation in prosthetic cardiovascular devices and arterial disease processes. While platelet activation may be induced by biochemical agonists, shear stresses arising from pathological flow patterns enhance the propensity of platelets to activate and initiate the intrinsic pathway of coagulation, leading to thrombosis. Upon activation platelets undergo complex biochemical and morphological changes: organelles are centralized, membrane glycoproteins undergo conformational changes, and adhesive pseudopods are extended. Activated platelets polymerize fibrinogen into a fibrin network that enmeshes red blood cells. Activated platelets also cross-talk and aggregate to form thrombi. Current numerical simulations to model this complex process mostly treat blood as a continuum and solve the Navier-Stokes equations governing blood flow, coupled with diffusion-convection-reaction equations. It requires various complex constitutive relations or simplifying assumptions, and is limited to μm level scales. However, molecular mechanisms governing platelet shape change upon activation and their effect on rheological properties can be in the nm level scales. To address this challenge, a multiscale approach which departs from continuum approaches, may offer an effective means to bridge the gap between macroscopic flow and cellular scales. Molecular dynamics (MD) and dissipative particle dynamics (DPD) methods have been employed in recent years to simulate complex processes at the molecular scales, and various viscous fluids at low-to-high Reynolds numbers at mesoscopic scales. Such particle methods possess important properties at the mesoscopic scale: complex fluids with heterogeneous particles can be modeled, allowing the simulation of processes which are otherwise very difficult to solve by continuum approaches. It is becoming a powerful tool for simulating complex blood flow, red blood cells interactions, and platelet-mediated thrombosis involving platelet activation, aggregation, and adhesion.

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Reduced NO-dependent arteriolar dilation during the development of cardiomyopathy

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.2000.278.2.h461 ◽

2000 ◽

Vol 278 (2) ◽

pp. H461-H468 ◽

Cited By ~ 24

Author(s):

Dong Sun ◽

An Huang ◽

Gong Zhao ◽

Robert Bernstein ◽

Paul Forfia ◽

...

Keyword(s):

Heart Failure ◽

Shear Stress ◽

No Production ◽

Basal Diameter ◽

Cardiac Decompensation ◽

Dependent Component ◽

Coronary Blood Vessels ◽

Arteriolar Diameter ◽

Coronary Arterioles

Our previous studies have suggested that there is reduced nitric oxide (NO) production in canine coronary blood vessels after the development of pacing-induced heart failure. The goal of these studies was to determine whether flow-induced NO-mediated dilation is altered in coronary arterioles during the development of heart failure. Subepicardial coronary arterioles (basal diameter 80 μm) were isolated from normal canine hearts, from hearts with dysfunction but no heart failure, and from hearts with severe cardiac decompensation. Arterioles were perfused at increasing flow or administered agonists with no flow in vitro. In arterioles from normal hearts, flow increased arteriolar diameter, with one-half of the response being NO dependent and one-half prostaglandin dependent. Shear stress-induced dilation was eliminated by removing the endothelium. Arterioles from normal hearts and hearts with dysfunction but no failure responded to increasing shear stress with dilation that reached a maximum at a shear stress of 20 dyn/cm2. In contrast, arterioles from failing hearts showed a reduced dilation, reaching only 55% of the dilation seen in vessels of normal hearts at a shear stress of 100 dyn/cm2. This remaining dilation was eliminated by indomethacin, suggesting that the NO-dependent component was absent in coronary microvessels after the development of heart failure. Similarly, agonist-induced NO-dependent coronary arteriolar dilation was markedly attenuated after the development of heart failure. After the development of severe dilated cardiomyopathy and heart failure, the NO-dependent component of both shear stress- and agonist-induced arteriolar dilation is reduced or entirely absent.

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