scholarly journals Global Thrombosis Test: Occlusion is attributable to shear-induced platelet thrombus formation.

TH Open ◽  
2021 ◽  
Author(s):  
Diana Adrienne Gorog ◽  
J. Yamamoto
Keyword(s):  
Platelet Aggregation ◽  
Experimental Approach ◽  
Thrombus Formation ◽  
Published Data ◽  
High Shear ◽  
Coagulation Time ◽  
P2y12 Inhibitors ◽  
Large Platelet ◽  
Platelet Thrombus ◽  
Platelet Aggregates

Herein, we set out a rebuttal to the publication by Claveria and co-workers published in TH Open this month entitled “Global Thrombosis Test: Occlusion by Coagulation or SIPA?” We strongly believe that the conclusions of their paper, suggesting that occlusion (OT) in the Global Thrombosis Test (GTT) is due to coagulation, rather than shear-induced platelet thrombus formation, is incorrect and the evidence and arguments they present are fundamentally flawed, with major errors both in the experimental approach and in the interpretations of the results. The evidence which they demonstrate, shows that occlusion in the GTT is, in fact, caused by high shear induced platelet thrombus formations. We set out herein the evidence for that, based on histology of the thrombus from the GTT in earlier work using electron microscopy showing large platelet aggregates, the very brief timescale of OT in the GTT compared to coagulation time and the sensitivity of the OT in the GTT to the effects of heparin, t-PA and P2Y12 inhibitors. In addition, we revisit the known pathomechanism of high shear-mediated platelet aggregation to underpin our rationale and show that the modifications to the instrument proposed by Claveria and co-authors would render the technique unphysiological. We highlight several methodological concerns and apparent misinterpreted of the data obtained. We present evidence predominantly from the authors’ own data, together with our earlier published data and evidence from the literature, showing that occlusion in the GTT occurs do to shear-induced platelet aggregation.

scholarly journals Distinct dietary alpha-linolenic acid-dependent shifts in the fecal microbiome composition suppresses aging-associated inflammatory responses and thrombus formation

2020 ◽  
Vol 41 (Supplement_2) ◽  
Author(s):  
S.S Saeedi Saravi ◽  
N.R Bonetti ◽  
G.G Camici ◽  
T.F Luscher ◽  
J.H Beer
Keyword(s):  
Platelet Aggregation ◽  
Shear Flow ◽  
Plasma Levels ◽  
Inflammatory Responses ◽  
Thrombus Formation ◽  
Funding Source ◽  
High Shear ◽  
Fecal Microbiome ◽  
Microbiome Composition

Abstract Background Aging is associated with alterations in the fecal microbiome composition. The microbiota-derived trimethylamine-N-oxide (TMAO) correlates with arterial thrombotic events, e.g. myocardial infarction and stroke, the leading causes of mortality worldwide. The omega-3 fatty acid (n-3 FA) α-linolenic acid (ALA) has been shown to be protective against thrombosis and associated pathologies. Therefore, we hypothesized that long-term dietary ALA supplementation protects against the aging-associated microbiome dysbiosis, and reduces inflammatory and thrombotic responses. Methods 24 week-old male C57BL/6 mice were fed either a high ALA (7.3g%) or low ALA (0.03g%) diet for 12 months. We examined the compositional changes of fecal microbiota of the animals treated with high vs. low ALA via 16S rRNA gene sequencing. The plasma levels of TMAO and its precursors choline and betaine, and LPS were measured by ELISA. Additionally, the platelet aggregation in response to thrombin, and thrombus formation on collagen under high-shear flow conditions of 3000/sec (to mimic blood flow in stenosed arteries) were investigated. Results Genomic analyses showed that the abundance of Phylum Proteobacteria and the family of desulfovibrio were reduced 71.72% and 51.73% in the aged high ALA-treated mice (p<0.01 and p<0.001, resp.) that may result in decrease in TAMO production and the subsequent inflammatory responses. However, microbial diversity of Bacteroidetes or Fermicutes and Bacteroidetes/Fermicutes ratio did not demonstrate a significant change between high vs. low ALA groups. Interestingly, the dietary intake of high ALA increased the abundance of Lachnospiraceae (p<0.01) that may exert anti-inflammatory effects. Importantly, high ALA significantly decreased the plasma levels of TMAO (p<0.01) and its precursor choline (P<0.05), but not betaine. The pro-inflammatory cytokine TNF-α showed a significant reduction (p<0.05), whereas plasma IL-1β did not change significantly following high ALA supplementation. An increased thrombus formation on collagen under high-shear flow (36.34%, p<0.01) and thrombin-induced platelet aggregation (31.31%, p<0.05) were found in the aged mice. Conclusion These studies demonstrate that an ALA-rich diet induces beneficial bacterial shifts in the aging-associated fecal microbiome that may lead to the suppression of inflammatory and thrombotic responses. Hence, long-term dietary ALA supplementation may be exploited as a nutritional antithrombotic strategy in the aging. Microbiome-Thrombosis-Aging Funding Acknowledgement Type of funding source: Foundation. Main funding source(s): Swiss National Science Foundation (SNSF)

scholarly journals Antithrombotic effect of a monoclonal antibody to the platelet glycoprotein IIb/IIIa receptor in an experimental animal model

Blood ◽  
1986 ◽  
Vol 68 (3) ◽  
pp. 783-786 ◽  
Author(s):  
BS Coller ◽  
JD Folts ◽  
LE Scudder ◽  
SR Smith
Keyword(s):  
Monoclonal Antibody ◽  
Animal Model ◽  
Platelet Aggregation ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Platelet Glycoprotein ◽  
Antithrombotic Effect ◽  
Antithrombotic Agent ◽  
Antithrombotic Effects ◽  
Platelet Thrombus

A murine monoclonal antibody directed at the platelet glycoprotein IIb/IIIa complex, which blocks platelet aggregation ex vivo, was tested for its antithrombotic effects in an established animal model of acute platelet thrombus formation in partially stenosed arteries. Infusion of 0.7 to 0.8 mg/kg of the F(ab')2 fragment of the antibody completely blocked new thrombus formation despite multiple provocations, making it the most potent antithrombotic agent tested in this model.

scholarly journals Haemodynamic flow conditions at the initiation of high-shear platelet aggregation: a combined in vitro and cellular in silico study

2020 ◽  
Vol 11 (1) ◽  
pp. 20190126 ◽  
Author(s):  
B. J. M. van Rooij ◽  
G. Závodszky ◽  
A. G. Hoekstra ◽  
D. N. Ku
Keyword(s):  
Platelet Aggregation ◽  
In Silico ◽  
Platelet Rich Plasma ◽  
High Shear ◽  
Shear Rates ◽  
Flow Simulations ◽  
In Silico Study ◽  
Platelet Aggregates

The influence of the flow environment on platelet aggregation is not fully understood in high-shear thrombosis. The objective of this study is to investigate the role of a high shear rate in initial platelet aggregation. The haemodynamic conditions in a microfluidic device are studied using cell-based blood flow simulations. The results are compared with in vitro platelet aggregation experiments performed with porcine whole blood (WB) and platelet-rich-plasma (PRP). We studied whether the cell-depleted layer in combination with high shear and high platelet flux can account for the distribution of platelet aggregates. High platelet fluxes at the wall were found in silico . In WB, the platelet flux was about twice as high as in PRP. Additionally, initial platelet aggregation and occlusion were observed in vitro in the stenotic region. In PRP, the position of the occlusive thrombus was located more downstream than in WB. Furthermore, the shear rates and stresses in cell-based and continuum simulations were studied. We found that a continuum simulation is a good approximation for PRP. For WB, it cannot predict the correct values near the wall.

scholarly journals Platelet-derived microparticles associate with fibrin during thrombosis

Blood ◽  
1996 ◽  
Vol 87 (11) ◽  
pp. 4651-4663 ◽  
Author(s):  
P Siljander ◽  
O Carpen ◽  
R Lassila
Keyword(s):  
Type I Collagen ◽  
Thrombus Formation ◽  
Flow Chamber ◽  
Type I ◽  
Binding Studies ◽  
Vitro Binding ◽  
Large Platelet ◽  
Platelet Aggregates ◽  
Thrombin Antithrombin Iii Complex

Platelet-derived microparticles (MP) are reported to express both pro- and anticoagulant activities. Nevertheless, their functional significance has remained unresolved. The present study monitored the generation and fate of MP in an experimental model of thrombosis with costimulation of platelets by collagen and thrombin. When minimally anticoagulated (0.5 micromol/L PPACK) blood was perfused over immobilized fibrillar type I collagen in a flow chamber at a low shear rate (300 s(-1)), endogenous thrombin was generated, as evidenced by thrombin-antithrombin III complex. In contrast to full anticoagulation 150 micromol/L PPACK) and the absence of collagen, large platelet aggregates and fibrin ensued during perfusions over collagen in the presence of thrombin. In these thrombi, MP, defined as GPIIbIIIa- and P- selectin-positive vesicles (<1 micron), were found to align fibrin in immunofluorescence and scanning immunoelectron microscopy. Moreover, in sections of embolectomized thromboemboli from patients GPIIbIIIa- and P- selectin-positive material compatible with MP was detected in a fibrin strand-like pattern. In vitro binding studies showed that MP bound to fibrin and acted there as procoagulants. In summary, we show that MP generated during thrombus formation associate with local fibrin. This adhesive function fibrin could imply a sustained modulatory role for MP in evolving thrombi.

scholarly journals Biomechanical thrombosis: the dark side of force and dawn of mechano-medicine

2019 ◽  
Vol 5 (2) ◽  
pp. 185-197 ◽  
Author(s):  
Yunfeng Chen ◽  
Lining Arnold Ju
Keyword(s):  
Platelet Aggregation ◽  
Blood Clotting ◽  
Thrombus Formation ◽  
Dark Side ◽  
Atherosclerotic Plaques ◽  
Excessive Bleeding ◽  
Glycoprotein Vi ◽  
Device Implantation ◽  
Platelet Binding ◽  
Platelet Thrombus

Arterial thrombosis is in part contributed by excessive platelet aggregation, which can lead to blood clotting and subsequent heart attack and stroke. Platelets are sensitive to the haemodynamic environment. Rapid haemodynamcis and disturbed blood flow, which occur in vessels with growing thrombi and atherosclerotic plaques or is caused by medical device implantation and intervention, promotes platelet aggregation and thrombus formation. In such situations, conventional antiplatelet drugs often have suboptimal efficacy and a serious side effect of excessive bleeding. Investigating the mechanisms of platelet biomechanical activation provides insights distinct from the classic views of agonist-stimulated platelet thrombus formation. In this work, we review the recent discoveries underlying haemodynamic force-reinforced platelet binding and mechanosensing primarily mediated by three platelet receptors: glycoprotein Ib (GPIb), glycoprotein IIb/IIIa (GPIIb/IIIa) and glycoprotein VI (GPVI), and their implications for development of antithrombotic ‘mechano-medicine’ .

Endotoxin enhances microvascular thrombosis in mouse cremaster venules via a TLR4-dependent, neutrophil-independent mechanism

2006 ◽  
Vol 290 (4) ◽  
pp. H1671-H1679 ◽  
Author(s):  
Rolando E. Rumbaut ◽  
Ricardo V. Bellera ◽  
Jaspreet K. Randhawa ◽  
Corie N. Shrimpton ◽  
Swapan K. Dasgupta ◽  
...  
Keyword(s):  
Platelet Aggregation ◽  
Platelet Adhesion ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Mouse Strains ◽  
Microvascular Endothelium ◽  
Microvascular Thrombosis ◽  
Platelet Thrombus ◽  
Adhesive Interactions

Endotoxemia promotes adhesive interactions between platelets and microvascular endothelium in vivo. We sought to determine whether endotoxin (lipopolysaccharide, LPS) modified platelet thrombus formation in mouse cremaster venules and whether Toll-like receptor 4 (TLR4) and neutrophils were involved in the response. Intravital videomicroscopy was performed in the cremaster microcirculation of pentobarbital-anesthetized mice; venular platelet thrombi were induced with a light/dye endothelial injury model. C57BL/6 mice treated with Escherichia coli endotoxin had enhanced rates of venular platelet thrombus formation: the time to microvessel occlusion was reduced by ∼50% ( P < 0.005) compared with saline-treated animals. Enhanced microvascular thrombosis was evident as early as 2 h after LPS administration. LPS had no effect on thrombosis in either of two mouse strains with altered TLR4 signaling (C57BL/10ScNJ or C3H/HeJ), whereas it enhanced thrombosis in the control strains (C57BL/10J and C3H/HeN). LPS also enhanced platelet adhesion to endothelium in the absence of light/dye injury. Platelet adhesion, but not enhanced thrombosis, was inhibited by depletion of circulating neutrophils. LPS failed to enhance platelet aggregation ex vivo and did not influence platelet P-selectin expression, a marker of platelet activation. These findings support the notion that endotoxemia promotes platelet thrombus formation independent of neutrophils and without enhancement of platelet aggregation, via a TLR4-dependent mechanism.

Different Effects of Aspirin, Dipyridamole and UD-CG 115 on Platelet Activation in a Model of Vascular Injury: Studies with Extracellular Matrix Covered with Endothelial Cells

1986 ◽  
Vol 56 (03) ◽  
pp. 333-339 ◽  
Author(s):  
A Eldor ◽  
I Vlodavsky ◽  
Z Fuks ◽  
T H Muller ◽  
W G Eisert
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Platelet Aggregation ◽  
Cyclic Amp ◽  
Platelet Activation ◽  
Platelet Rich Plasma ◽  
Thromboxane A2 ◽  
Phase Microscopy ◽  
Large Platelet ◽  
Platelet Aggregates

SummaryCultured endothelial cells produce an extracellular matrix (ECM) which activates platelets, similarly to deendothelialized vascular segments. Platelet-rich plasma (PRP) was incubated with endothelial cells cultures seeded in various densities on ECM. The interaction of the platelets with this artifical intima was evaluated by phase microscopy and by thromboxane A2 (TXA2) and prostacyclin (PGI2) measurement. Large platelet aggregates were formed on exposed ECM. Platelets aggregation but not adhesion on the ECM was markedly inhibited by the presence of endothelial cells. Pretreatment of the endothelial cells with 0.1 mM aspirin reduced their PGI2 synthesis and was associated with platelet aggregation on the ECM. 10 μM dipyridamole markedly inhibited platelet activation by ECM when the drug was added to citrated whole blood before PRP preparation. UD-CG 115 which elevates cyclic AMP in cardiac muscle, inhibited platelet aggregation and TXA2 production induced by ECM, in the presence as well as in the absence of endothelial cells, without any effect on endothelial PGI2 production.

The Formation of Fibrin Around Large Platelet Aggregates (PA’s) in Heparinized Platelet Plasma (PRP)

1979 ◽  
Author(s):  
L.J. Wurzinger ◽  
P. Blasberg ◽  
H. Vehr
Keyword(s):  
Platelet Aggregation ◽  
Local Formation ◽  
Shear Field ◽  
Heparin Concentration ◽  
Fibrin Formation ◽  
Platelet Protein ◽  
Fibrin Network ◽  
Large Platelet ◽  
Platelet Aggregates

In concavities on the surface of large PA’s (107-108 μm3) which were produced “spontaneously” or upon addition of ADP 10-6 M in a homogenous shear field in a Couette-type aggregometer (40 s-1), we observed the formation of a fibrin network within 5 mins after sampling. This fibrin network, which could be identified with fluorescent antibodies, formed despite anticoagulation with 7.5 II heparin/ml. The formation of this fibrin network was less pronounced and delayed when the heparin concentration was raised to 20 U/mL 50 U/ml completely suppressed the formation of the network. Concomitantly we found masses of degranulated and ballooned platelets around PA’s with fibrin formation. As we suspected secreted’PF 4 (Antiheparinfactor) being responsible for this phenomenon, we analysed the β-thromboglobulin (β-TC)in heparinized PRP, which is secreted along with PF 4.We found the formation of fibrin not so much correlated to the extent of platelet aggregation in the whole sample, but to the release of β-TG. The samples wherein fibrin was formed, were characterized by β-TG levels in plasma that amounted to up 5% of total platelet protein. In our view this explains the fact, that despite heparinization during extracoporeal circulation the local formation of fibrin-consolidated white thrombi and emboli is observed.

Abstract 33: Platelet 12-lipoxygenase is a Key Regulator of Platelet Reactivity and Thrombus Formation in vivo

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Reheman Adili ◽  
Katherine Mast ◽  
Theodore R Holman ◽  
Michael Holinstat
Keyword(s):  
Platelet Aggregation ◽  
Ex Vivo ◽  
Platelet Reactivity ◽  
Thrombus Formation ◽  
High Shear ◽  
Vessel Occlusion ◽  
12 Lipoxygenase ◽  
Induced Aggregation

Background: Platelet reactivity is required to maintain hemostasis, however high platelet reactivity leads to thrombus formation, myocardial infarction, and stroke. Platelet 12-lipoxygenase (12-LOX) has been demonstrated by our lab and others to regulate agonist-mediated platelet reactivity suggesting a role for 12-LOX in regulation of in vivo thrombosis. The ability to target 12-LOX in vivo has not been established to date. Therefore, we sought to determine if 12-LOX regulates platelet reactivity and thrombus formation in vivo using the selective 12-LOX inhibitor ML355 to determine whether platelet 12-LOX is an effective target for anti-platelet therapeutics. Methods: ML355 effects on human platelet function was assessed in vitro by platelet aggregometry, ex vivo by perfusion chamber, and in vivo by thrombus formation and vessel occlusion in small and large vessels in 12-LOX -/- , WT mice, and mice treated with ML355 via intravital microscopy using the FeCl 3 and laser injury models. Results: In in vitro platelet aggregation, ML355 dose-dependently inhibited agonist-induced aggregation. In ex vivo flow chamber assays, platelet adhesion and thrombus formation on collagen-coated surfaces at high shear was attenuated in both mouse and human whole blood after incubation with ML355. Further, platelet aggregation and thrombus growth in 12-LOX -/- mice were impaired in both laser and FeCl 3 -induced mesenteric, carotid artery and cremaster arteriole thrombosis models. Thrombi in 12-LOX -/- mice were unstable and frequently formed emboli, which resulted in impaired vessel occlusion or reopening. Additionally, thrombus formation and vessel occlusion was impaired in ML355 treated WT mice. Conclusions: The 12-LOX inhibitor ML355 inhibits platelet aggregation induced by a number of platelet agonists. Ex vivo high shear conditions in both mice and human was attenuated in the presence of ML355. Thrombus formation and vessel occlusion were impaired in mice deficient in 12-LOX. Finally, ML355 attenuates thrombus formation and prevents vessel occlusion in vivo . Our data strongly indicates 12-LOX is an important determinant of platelet reactivity and inhibition of platelet 12-LOX may represent a new target for anti-platelet therapeutics.

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