scholarly journals Enhancement of thrombogenesis by plasma fibronectin cross-linked to fibrin and assembled in platelet thrombi

Blood ◽  
2006 ◽  
Vol 107 (9) ◽  
pp. 3555-3563 ◽  
Author(s):  
Jaehyung Cho ◽  
Deane F. Mosher
Keyword(s):  
Ex Vivo ◽  
Thrombus Formation ◽  
Flow Chamber ◽  
Plasma Fibronectin ◽  
Increased Risk ◽  
Platelet Thrombus ◽  
Platelet Thrombi ◽  
Artery Disease ◽  
Fibronectin Deposition ◽  
The Impact

To learn how plasma fibronectin stabilizes platelet-rich thrombi in injured mesenteric arterioles of mice, we studied the impact of plasma fibronectin on platelet thrombus formation ex vivo in a parallel flow chamber. Thrombi were greater on surfaces coated with fibrin cross-linked to fibronectin by activated factor XIII than on surfaces coated with fibrin lacking cross-linked fibronectin or with fibronectin alone. Platelet thrombi were even greater when plasma fibronectin was perfused with platelets, resulting in deposition of the perfused fibronectin in platelet thrombi. The effect of perfused fibronectin on thrombogenesis was lost if fibronectin deposition was blocked by coperfusion with the N-terminal 70-kDa fragment of fibronectin or a peptide based on the functional upstream domain of protein F1 of Streptococcus pyogenes. Increases in thrombus formation were dependent on a platelet activator such as lysophosphatidic acid, amount of fibronectin cross-linked to fibrin, and concentration of fibronectin in the perfusate. The dependency of fibronectin concentration extended into the range of fibronectin concentrations associated with increased risk of coronary artery disease. At such concentrations, the 2 mechanisms for insolubilization of plasma fibronectin—cross-linking to fibrin and assembly by adherent and aggregating platelets—synergize to result in many-fold enhancement of platelet thrombus formation.

scholarly journals Impact of established cardiovascular disease on 10-year death after coronary revascularization for complex coronary artery disease

2021 ◽  
Author(s):  
Rutao Wang ◽  
Scot Garg ◽  
Chao Gao ◽  
Hideyuki Kawashima ◽  
Masafumi Ono ◽  
...  
Keyword(s):  
Cardiovascular Disease ◽  
Coronary Artery Disease ◽  
Coronary Artery ◽  
Coronary Revascularization ◽  
Long Term Outcomes ◽  
Vital Status ◽  
Increased Risk ◽  
Artery Disease ◽  
The Impact

Abstract Aims To investigate the impact of established cardiovascular disease (CVD) on 10-year all-cause death following coronary revascularization in patients with complex coronary artery disease (CAD). Methods The SYNTAXES study assessed vital status out to 10 years of patients with complex CAD enrolled in the SYNTAX trial. The relative efficacy of PCI versus CABG in terms of 10-year all-cause death was assessed according to co-existing CVD. Results Established CVD status was recorded in 1771 (98.3%) patients, of whom 827 (46.7%) had established CVD. Compared to those without CVD, patients with CVD had a significantly higher risk of 10-year all-cause death (31.4% vs. 21.7%; adjusted HR: 1.40; 95% CI 1.08–1.80, p = 0.010). In patients with CVD, PCI had a non-significant numerically higher risk of 10-year all-cause death compared with CABG (35.9% vs. 27.2%; adjusted HR: 1.14; 95% CI 0.83–1.58, p = 0.412). The relative treatment effects of PCI versus CABG on 10-year all-cause death in patients with complex CAD were similar irrespective of the presence of CVD (p-interaction = 0.986). Only those patients with CVD in ≥ 2 territories had a higher risk of 10-year all-cause death (adjusted HR: 2.99, 95% CI 2.11–4.23, p < 0.001) compared to those without CVD. Conclusions The presence of CVD involving more than one territory was associated with a significantly increased risk of 10-year all-cause death, which was non-significantly higher in complex CAD patients treated with PCI compared with CABG. Acceptable long-term outcomes were observed, suggesting that patients with established CVD should not be precluded from undergoing invasive angiography or revascularization. Trial registration SYNTAX: ClinicalTrials.gov reference: NCT00114972. SYNTAX Extended Survival: ClinicalTrials.gov reference: NCT03417050. Graphic abstract

scholarly journals Ex vivo culture of Fancc-/- stem/progenitor cells predisposes cells to undergo apoptosis, and surviving stem/progenitor cells display cytogenetic abnormalities and an increased risk of malignancy

Blood ◽  
2005 ◽  
Vol 105 (9) ◽  
pp. 3465-3471 ◽  
Author(s):  
Xiaxin Li ◽  
Michelle M. Le Beau ◽  
Samantha Ciccone ◽  
Feng-Chun Yang ◽  
Brian Freie ◽  
...  
Keyword(s):  
Progenitor Cells ◽  
Genome Stability ◽  
Bone Marrow Cells ◽  
Ex Vivo ◽  
Acquired Resistance ◽  
Intrinsic Defects ◽  
Cytogenetic Abnormalities ◽  
Increased Risk ◽  
Ex Vivo Culture ◽  
The Impact

AbstractCurrent strategies for genetic therapy using Moloney retroviruses require ex vivo manipulation of hematopoietic cells to facilitate stable integration of the transgene. While many studies have evaluated the impact of ex vivo culture on normal murine and human stem/progenitor cells, the cellular consequences of ex vivo manipulation of stem cells with intrinsic defects in genome stability are incompletely understood. Here we show that ex vivo culture of Fancc-/- bone marrow cells results in a time-dependent increase in apoptosis of primitive Fancc-/- progenitor cells in conditions that promote the proliferation of wild-type stem/progenitor cells. Further, recipients reconstituted with the surviving Fancc-/- cells have a high incidence of cytogenetic abnormalities and myeloid malignancies that are associated with an acquired resistance to tumor necrosis factor α (TNF-α). Collectively, these data indicate that the intrinsic defects in the genomic stability of Fancc-/- stem/progenitor cells provide a selective pressure for cells that are resistant to apoptosis and have a propensity for the evolution to clonal hematopoiesis and malignancy. These studies could have implications for the design of genetic therapies for treatment of Fanconi anemia and potentially other genetic diseases with intrinsic defects in genome stability.

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.

Platelets, circulating tissue factor, and fibrin colocalize in ex vivo thrombi: real-time fluorescence images of thrombus formation and propagation under defined flow conditions

Blood ◽  
2002 ◽  
Vol 100 (8) ◽  
pp. 2787-2792 ◽  
Author(s):  
Viji Balasubramanian ◽  
Eric Grabowski ◽  
Alessandra Bini ◽  
Yale Nemerson
Keyword(s):  
Real Time ◽  
Tissue Factor ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
High Sensitivity ◽  
Flow Chamber ◽  
Flow Conditions ◽  
Coated Glass ◽  
Relative Contribution ◽  
Fluorescent Markers

Although it is generally accepted that the initial event in coagulation and intravascular thrombus formation is the exposure of tissue factor (TF) to blood, there is still little agreement about the mechanisms of thrombus propagation and the identities of the molecular species participating in this process. In this study, we characterized the thrombotic process in real-time and under defined flow conditions to determine the relative contribution and spatial distribution of 3 components of the thrombi: circulating or blood-borne TF (cTF), fibrin, and platelets. For this purpose, we used high-sensitivity, multicolor immunofluorescence microscopy coupled with a laminar flow chamber. Freshly drawn blood, labeled with mepacrine (marker for platelets and white cells), anti-hTF1Alexa.568 (marker for tissue factor), and anti-T2G1Cy­5 (marker for fibrin) was perfused over collagen-coated glass slides at wall shear rates of 100 and 650 s−1. A motorized filter cube selector facilitated imaging every 5 seconds at 1 of 3 different wavelengths, corresponding to optimal wavelengths for the 3 markers above. Real-time video recordings obtained during each of 10 discrete experiments show rapid deposition of platelets and fibrin onto collagen-coated glass. Overlay images of fluorescent markers corresponding to platelets, fibrin, and cTF clearly demonstrate colocalization of these 3 components in growing thrombi. These data further support our earlier observations that, in addition to TF present in the vessel wall, there is a pool of TF in circulating blood that contributes to the propagation of thrombosis at a site of vascular injury.

Abstract 54: ML355 in Prevention of Thrombosis in vivo with Minimal Effects on Hemostasis

2017 ◽  
Vol 37 (suppl_1) ◽  
Author(s):  
Reheman Adili ◽  
Katherine Mast ◽  
Michael Holinstat
Keyword(s):  
Ex Vivo ◽  
Platelet Reactivity ◽  
Thrombus Formation ◽  
Flow Chamber ◽  
Human Platelets ◽  
Mass Spectrometry Analysis ◽  
Vessel Occlusion ◽  
Spectrometry Analysis ◽  
Thrombosis Model

12-lipoxygenase (12-LOX) has been demonstrated to regulate platelet function, hemostasis, and thrombosis ex vivo , supporting a key role for 12-LOX in regulation of in vivo thrombosis. While pharmacologically targeting 12-LOX in vivo has been a challenge to date, the recent development of the 12-LOX selective inhibitor, ML355, as an effective antiplatelet therapeutic in vivo was assessed. ML355 potently inhibited thrombin and other agonist-induced platelet aggregation ex vivo in washed human platelets and inhibited downstream oxylipin production of platelet 12-LOX as confirmed by Mass spectrometry analysis. Ex vivo flow chamber assays confirmed that human platelet adhesion and thrombus formation at arterial shear over collagen was attenuated in human whole blood treated with ML355 to a greater extent compared to aspirin. In vivo , PK assessment of ML355 showed reasonable 12-LOX plasma levels 12 hours following administration of ML355. FeCl 3 -induced injury of the mesenteric arterioles resulted in less stable thrombi in 12-LOX -/- mice and ML355-treated WT mice resulting in impairment of vessel occlusion. Additionally, ML355 dose-dependently inhibited laser-induced thrombus formation in the cremaster arteriole thrombosis model in WT, but not in 12-LOX -/- mice. Importantly, hemostatic plug formation and bleeding following treatment with ML355 were not affected in response to laser ablation on the saphenous vein or in a cremaster microvasculature laser-induced rupture model. Our data strongly supports 12-LOX as a key determinant of platelet reactivity in vivo and inhibition of platelet 12-LOX with ML355 may represent a new class of antiplatelet therapeutics.

A Double-Blind Randomized Comparison of Combined Aspirin and Ticlopidine Therapy Versus Aspirin or Ticlopidine Alone on Experimental Arterial Thrombogenesis in Humans

Blood ◽  
1998 ◽  
Vol 92 (5) ◽  
pp. 1518-1525 ◽  
Author(s):  
Jean-Pierre Bossavy ◽  
Claire Thalamas ◽  
Luc Sagnard ◽  
André Barret ◽  
Kjell Sakariassen ◽  
...  
Keyword(s):  
Ex Vivo ◽  
Thrombus Formation ◽  
Randomized Study ◽  
Combined Treatment ◽  
Antithrombotic Effect ◽  
Ex Vivo Model ◽  
Double Blind ◽  
Thrombotic Complications ◽  
Platelet Thrombus ◽  
American Society

Abstract No randomized study comparing the effect of combined ticlopidine and aspirin therapy versus each drug alone in reducing poststenting thrombotic complications has been performed. To compare these three antiplatelet regimens versus placebo, we conducted a double-blind randomized study using an ex vivo model of thrombosis. Sixteen healthy male volunteers were assigned to receive for 8 days the following four regimens separated by a 1-month period: aspirin 325 mg/d, ticlopidine 500 mg/d, aspirin 325 mg/d + ticlopidine 500 mg/d, and placebo. At the end of each treatment period, native nonanticoagulated blood was drawn directly from an antecubital vein over collagen- or tissue factor (TF)-coated coverslips positioned in a parallel-plate perfusion chamber at an arterial wall shear rate (2,600 s−1 ) for 3 minutes. Thrombus, which formed on collagen in volunteers treated by placebo, were rich in platelets and poor in fibrin. As compared with placebo, aspirin and ticlopidine alone reduced platelet thrombus formation by only 29% and 15%, respectively (P &gt; .2). In contrast, platelet thrombus formation was blocked by more than 90% in volunteers treated by aspirin + ticlopidine (P &lt; .01v placebo or each treatment alone). Furthermore, the effect of the drug combination therapy was significantly larger than the sum of the two active treatments (P &lt; .05). Thrombus, which formed on TF-coated coverslips in volunteers treated by placebo, were rich in fibrin and platelets. Neither of the three antiplatelet treatments significantly inhibited fibrin deposition and platelet thrombus formation on this surface (P &gt; .2). Thus, the present study shows that combined aspirin and ticlopidine therapy dramatically potentiates the antithrombotic effect of each drug alone, but that the antithrombotic effect of the combined treatment depends on the nature of the thrombogenic surface. © 1998 by The American Society of Hematology.

scholarly journals Severe platelet dysfunction in NHL patients receiving ibrutinib is absent in patients receiving acalabrutinib

2017 ◽  
Vol 1 (26) ◽  
pp. 2610-2623 ◽  
Author(s):  
Alexander P. Bye ◽  
Amanda J. Unsworth ◽  
Michael J. Desborough ◽  
Catherine A. T. Hildyard ◽  
Niamh Appleby ◽  
...  
Keyword(s):  
Platelet Function ◽  
Ex Vivo ◽  
Thrombus Formation ◽  
Critical Role ◽  
Platelet Dysfunction ◽  
Non Hodgkin Lymphoma ◽  
Healthy Donors ◽  
Increased Risk ◽  
Btk Inhibitor ◽  
Size And Morphology

Abstract The Bruton tyrosine kinase (Btk) inhibitor ibrutinib induces platelet dysfunction and causes increased risk of bleeding. Off-target inhibition of Tec is believed to contribute to platelet dysfunction and other side effects of ibrutinib. The second-generation Btk inhibitor acalabrutinib was developed with improved specificity for Btk over Tec. We investigated platelet function in patients with non-Hodgkin lymphoma (NHL) receiving ibrutinib or acalabrutinib by aggregometry and by measuring thrombus formation on collagen under arterial shear. Both patient groups had similarly dysfunctional aggregation responses to collagen and collagen-related peptide, and comparison with mechanistic experiments in which platelets from healthy donors were treated with the Btk inhibitors suggested that both drugs inhibit platelet Btk and Tec at physiological concentrations. Only ibrutinib caused dysfunctional thrombus formation, whereas size and morphology of thrombi following acalabrutinib treatment were of normal size and morphology. We found that ibrutinib but not acalabrutinib inhibited Src family kinases, which have a critical role in platelet adhesion to collagen that is likely to underpin unstable thrombus formation observed in ibrutinib patients. We found that platelet function was enhanced by increasing levels of von Willebrand factor (VWF) and factor VIII (FVIII) ex vivo by addition of intermediate purity FVIII (Haemate P) to blood from patients, resulting in consistently larger thrombi. We conclude that acalabrutinib avoids major platelet dysfunction associated with ibrutinib therapy, and platelet function may be enhanced in patients with B-cell NHL by increasing plasma VWF and FVIII.

scholarly journals Heterozygous ABCG5 Gene Deficiency and Risk of Coronary Artery Disease

2020 ◽  
Vol 13 (5) ◽  
pp. 417-423 ◽  
Author(s):  
Akihiro Nomura ◽  
Connor A. Emdin ◽  
Hong Hee Won ◽  
Gina M. Peloso ◽  
Pradeep Natarajan ◽  
...  
Keyword(s):  
Coronary Artery Disease ◽  
Coronary Artery ◽  
Large Scale ◽  
Carrier Status ◽  
Loss Of Function ◽  
Increased Risk ◽  
Genetic Deficiency ◽  
Heterozygous Carriers ◽  
Artery Disease ◽  
The Impact

Background: Familial sitosterolemia is a rare Mendelian disorder characterized by hyperabsorption and decreased biliary excretion of dietary sterols. Affected individuals typically have complete genetic deficiency—homozygous loss-of-function (LoF) variants—in the ABCG5 or ABCG8 genes and have substantially elevated plasma sitosterol and LDL (low-density lipoprotein) cholesterol (LDL-C) levels. The impact of partial genetic deficiency of ABCG5 or ABCG8 —as occurs in heterozygous carriers of LoF variants—on LDL-C and risk of coronary artery disease (CAD) has remained uncertain. Methods: We first recruited 9 sitosterolemia families, identified causative LoF variants in ABCG5 or ABCG8 , and evaluated the associations of these ABCG5 or ABCG8 LoF variants with plasma phytosterols and lipid levels. We next assessed for LoF variants in ABCG5 or ABCG8 in CAD cases (n=29 321) versus controls (n=357 326). We tested the association of rare LoF variants in ABCG5 or ABCG8 with blood lipids and risk for CAD. Rare LoF variants were defined as protein-truncating variants with minor allele frequency <0.1% in ABCG5 or ABCG8 . Results: In sitosterolemia families, 7 pedigrees harbored causative LoF variants in ABCG5 and 2 pedigrees in ABCG8 . Homozygous LoF variants in either ABCG5 or ABCG8 led to marked elevations in sitosterol and LDL-C. Of those sitosterolemia families, heterozygous carriers of ABCG5 LoF variants exhibited increased sitosterol and LDL-C levels compared with noncarriers. Within large-scale CAD case-control cohorts, prevalence of rare LoF variants in ABCG5 and in ABCG8 was ≈0.1% each. ABCG5 heterozygous LoF variant carriers had significantly elevated LDL-C levels (25 mg/dL [95% CI, 14–35]; P =1.1×10 −6 ) and were at 2-fold increased risk of CAD (odds ratio, 2.06 [95% CI, 1.27–3.35]; P =0.004). By contrast, ABCG8 heterozygous LoF carrier status was not associated with increased LDL-C or risk of CAD. Conclusions: Although familial sitosterolemia is traditionally considered as a recessive disorder, we observed that heterozygous carriers of an LoF variant in ABCG5 had significantly increased sitosterol and LDL-C levels and a 2-fold increase in risk of CAD.

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.

scholarly journals A novel nucleotide-based thrombin inhibitor inhibits clot-bound thrombin and reduces arterial platelet thrombus formation

Blood ◽  
1994 ◽  
Vol 83 (3) ◽  
pp. 677-682 ◽  
Author(s):  
WX Li ◽  
AV Kaplan ◽  
GW Grant ◽  
JJ Toole ◽  
LL Leung
Keyword(s):  
Ex Vivo ◽  
Platelet Rich Plasma ◽  
Thrombus Formation ◽  
Thrombin Inhibitors ◽  
Thrombin Inhibitor ◽  
Shear Rates ◽  
High Shear Rates ◽  
Dependent Inhibition ◽  
Platelet Thrombus

A novel thrombin inhibitor based on single-stranded (ss) deoxynucleotides with the sequence GGTTGGTGTGGTTGG (thrombin aptamer) has been recently discovered. In this study, we tested its efficacy in inhibiting clot-bound thrombin activity and platelet thrombus formation in an ex vivo whole artery angioplasty model. The thrombin aptamer showed a specific dose-dependent inhibition of thrombin-induced platelet aggregation (0.5 U/mL) in human platelet-rich plasma, with an IC50 of approximately 70 to 80 nmol/L. In an in vitro clot-bound thrombin assay system, heparin, used at clinically relevant concentrations of 0.2 U/mL and 0.4 U/mL, was ineffective in inhibiting clot-bound thrombin (6.5% and 34.9% inhibition at 0.2 U/mL and 0.4 U/mL, respectively). In contrast, the thrombin aptamer at an equivalent anticoagulant concentration inhibited clot-bound thrombin (79.7% inhibition). In an ex vivo whole artery angioplasty model, the thrombin aptamer markedly suppressed the generation of fibrinopeptide A (FPA), whereas heparin at 2 U/mL was ineffective. Compared with a scrambled ssDNA control, the thrombin aptamer reduced platelet deposition by 34.5% +/- 5% (mean +/- SEM, n = 4, P = .09) at low shear rates (approximately 200 s-1) and 61.3% +/- 11% (mean +/- SEM, n = 4, P = .05) at high shear rates (approximately 850 s-1). Thrombin aptamers based on ssDNA molecules represent a new class of thrombin inhibitors with potent anticoagulant and antithrombotic properties.

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