scholarly journals Bivalirudin Inhibits Thrombin-Mediated Tissue Factor Expression in Human Endothelial Cells

2017 ◽  
Vol 2 ◽  
Author(s):  
Plinio Cirillo ◽  
Grazia Pellegrino ◽  
Stefano Conte ◽  
Giovanni Cimmino ◽  
Giusi Barra ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Coagulation Factors ◽  
Direct Thrombin Inhibitor ◽  
Procoagulant Activity ◽  
Thrombin Inhibitor ◽  
Pathophysiological Mechanism ◽  
Font Size ◽  
Mrna Transcription ◽  
Expression Activity

<p><span style="font-family: Times;"><span style="font-size: medium;">Thrombosis is the main pathophysiological mechanism in Acute Coronary Syndromes (ACS), and involves the activation of platelets and of Tissue Factor (TF)-dependent extrinsic coagulation pathway. TF-mRNA and antigen are detectable in the adventitia of normal vessels. On the contrary, little TF immunoreactivity is measurable in the smooth muscle cells of uninjured vessels and unperturbed endothelial cells, being in contact with circulating blood, usually do not express TF activity. However, several stimuli are able to induce TF in endothelial cells, including thrombin. Thus in an acute "scenario", thrombin might be responsible for creating a prothombotic milieau. Bivalirudin (BIVA) is a synthetic, reversible direct thrombin inhibitor actually considered a valuable alternative to heparins in patients who need anticoagulation in the setting of ACS and percutaneous coronary intervention to avoid acute thrombotic events. In the present study we have investigated whether BIVA, by inhibiting thrombin, might have effects on TF expression and procoagulant activity in endothelial cells. Human Umbilical Endothelial Cells (HUVEC) were stimulated with thrombin or with the activated coagulation factors FVIIa/FXa for 2 hrs to evaluate TF-mRNA transcription by real-time PCR and for 6 hrs to measure TF expression/activity  </span><span style="font-size: medium;">on cell surface by FACs analysis and procoagulant activity. In additional experiments HUVEC were </span><span style="font-size: medium;"> </span><span style="font-size: medium;">pre-treated with BIVA for 1 hr before being stimulated and processed as above. Thrombin induced TF-mRNA transcription as well TF expression/activity on HUVEC shifting them to a procoagulant phenotype. On the contrary, the activated coagulation factors FVIIa/FXa did not affect TF expression/activity, indicating that thrombin plays a pivotal role in mediating this phenomenon. BIVA was able to prevent these thrombin deleterious effects. Data of the present study, although in vitro, suggest that BIVA, in the context of ACS, might significantly reduce thrombogenicity not only by acting as direct thrombin inhibitor but through its effects on TF expression/activity too.</span><span style="font-size: medium;">    </span></span></p>

Desmopressin (DDAVP) Enhances Platelet Adhesion to the Extracellular Matrix of Cultured Human Endothelial Cells through Increased Expression of Tissue Factor

1997 ◽  
Vol 77 (05) ◽  
pp. 0975-0980 ◽  
Author(s):  
Angel Gálvez ◽  
Goretti Gómez-Ortiz ◽  
Maribel Díaz-Ricart ◽  
Ginés Escolar ◽  
Rogelio González-Sarmiento ◽  
...  
Keyword(s):  
Extracellular Matrix ◽  
Endothelial Cells ◽  
Tissue Factor ◽  
Platelet Adhesion ◽  
Umbilical Vein ◽  
Procoagulant Activity ◽  
Experimental Conditions ◽  
Chromogenic Assay

SummaryThe effect of desmopressin (DDAVP) on thrombogenicity, expression of tissue factor and procoagulant activity (PCA) of extracellular matrix (ECM) generated by human umbilical vein endothelial cells cultures (HUVEC), was studied under different experimental conditions. HUVEC were incubated with DDAVP (1, 5 and 30 ng/ml) and then detached from their ECM. The reactivity towards platelets of this ECM was tested in a perfusion system. Coverslips covered with DD A VP-treated ECMs were inserted in a parallel-plate chamber and exposed to normal blood anticoagulated with low molecular weight heparin (Fragmin®, 20 U/ml). Perfusions were run for 5 min at a shear rate of 800 s1. Deposition of platelets on ECMs was significantly increased with respect to control ECMs when DDAVP was used at 5 and 30 ng/ml (p <0.05 and p <0.01 respectively). The increase in platelet deposition was prevented by incubation of ECMs with an antibody against human tissue factor prior to perfusion. Immunofluorescence studies positively detected tissue factor antigen on DDAVP derived ECMs. A chromogenic assay performed under standardized conditions revealed a statistically significant increase in the procoagulant activity of the ECMs produced by ECs incubated with 30 ng/ml DDAVP (p <0.01 vs. control samples). Northern blot analysis revealed increased levels of tissue factor mRNA in extracts from ECs exposed to DDAVP. Our data indicate that DDAVP in vitro enhances platelet adhesion to the ECMs through increased expression of tissue factor. A similar increase in the expression of tissue factor might contribute to the in vivo hemostatic effect of DDAVP.

scholarly journals Immunologic Identification of Tissue Factor (Thromboplastin) Synthesized by Cultured Fibroblasts

Blood ◽  
1973 ◽  
Vol 41 (5) ◽  
pp. 671-678 ◽  
Author(s):  
Leo R. Zacharski ◽  
Leon W. Hoyer ◽  
O. Ross McIntyre
Keyword(s):  
Factor Viii ◽  
Tissue Factor ◽  
Coagulation Factors ◽  
Procoagulant Activity ◽  
Factor Vii ◽  
Cultured Fibroblasts ◽  
Procoagulant Effect ◽  
Tissue Thromboplastin ◽  
Thromboplastin Factor

Abstract Immunologic methods were employed in an attempt to identify a potent procoagulant present in homogenates of human skin fibroblasts cultured in vitro. The activity of this procoagulant was restricted to the early stages of coagulation and was heretofore considered to be due to tissue factor (tissue thromboplastin, factor III) either alone or in combination with one or more of the first-stage coagulation factors (VIII, IX, XI, XII). The present studies demonstrated that procoagulant activity was not diminished by incubation with anti-VIII or anti-IX antibodies of human origin or with anti-VIII antibody of rabbit origin. Furthermore, cell culture homogenates failed to bind antifactor VIII antibody and did not contain an inhibitor of the reaction between factor VIII and its antibody. By contrast, procoagulant activity was obliterated by an antibody to tissue factor protein regardless of whether plasmas deficient in factor VIII, IX, XI, or XII were used in the assay system. The antitissue factor antibody failed to block the procoagulant effect after tissue factor had complexed factor VII. The procoagulant, therefore, consisted entirely of tissue factor.

scholarly journals Association of Tissue Factor Pathway Inhibitor With Human Umbilical Vein Endothelial Cells

Blood ◽  
1997 ◽  
Vol 90 (9) ◽  
pp. 3568-3578 ◽  
Author(s):  
John-Bjarne Hansen ◽  
Randi Olsen ◽  
Paul Webster
Keyword(s):  
Endothelial Cells ◽  
Cell Surface ◽  
Tissue Factor ◽  
Tissue Factor Pathway Inhibitor ◽  
Umbilical Vein ◽  
Coagulation Factors ◽  
Coagulation System ◽  
Human Umbilical Vein ◽  
Tissue Factor Pathway ◽  
Umbilical Vein Endothelial Cells

AbstractTissue factor pathway inhibitor (TFPI) is a serine protease inhibitor of the extrinsic coagulation system, synthesized in endothelial cells, which has recently been shown to play an important role in the regulation of activated coagulation factors at the endothelial cell surface. In the present study we investigated the subcellular localization and metabolism of TFPI in human umbilical vein endothelial cells (HUVEC). Immunocytochemical labeling of HUVEC with anti-TFPI showed specific labeling associated with the cell surface and with many intracellular organelles including the Golgi complex. Further characterization of these organelles was performed by colocalizing the anti-TFPI with 3-(2,4-dinitroanilino)′-amino-N-methyldipropylamine (DAMP; to demonstrate low pH), mannose phosphate receptor (endosomes), and LAMP 1 (late endocytic compartments). TFPI also colocalized with antibodies to the human transferrin receptor, a marker for early endocytic, recycling compartment. Endogenous TFPI colocalized with biotin in intracellular vesicles during endocytosis after biotinylation of the cell surface, which indicated that TFPI was being co-internalized with the surface biotin. The binding of exogenously added 125I-TFPI increased linearly to HUVEC over the concentration range of 0 to 32 nmol/L without saturation, the binding was not affected by up to a thousand-fold molar excess of unlabeled TFPI, and heparin inhibited the binding dose dependently. An intact C-terminal domain was important for the interaction between TFPI and the cell surface of HUVEC, because less than 10% of a C-terminal truncated form of TFPI (TFPI1-161 ) was bound after addition of equimolar concentrations of full-length TFPI. Exogenously added 125I-TFPI was not degraded in HUVEC during 4 hours at 37°C. The presence of TFPI in endocytic and recycling compartments support the hypothesis that endogenous, membrane-anchored TFPI could be internalized for subsequent recycling back to the cell surface.

CD40 Engagement on Endothelial Cells Promotes Tissue Factor-dependent Procoagulant Activity

1998 ◽  
Vol 79 (05) ◽  
pp. 1025-1028 ◽  
Author(s):  
Ling Zhou ◽  
Patrick Stordeur ◽  
Aurore de Lavareille ◽  
Kris Thielemans ◽  
Paul Capel ◽  
...  
Keyword(s):  
T Cells ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Tissue Factor ◽  
T Cell Activation ◽  
Cell Activation ◽  
Umbilical Vein ◽  
Procoagulant Activity ◽  
Factor Vii ◽  
The Impact

SummaryThe CD40 molecule expressed on endothelial cells has been shown to transduce activation signals resulting in upregulation of adhesion molecules. Herein, we studied the impact of CD40 engagement on the induction of tissue factor (TF)-dependent procoagulant activity (PCA) at the surface of human umbilical vein endothelial cells (HUVECs). First, we found that co-incubation of HUVECs with 3T6 fibroblasts transfected with the CD40L gene (3T6-CD40L) resulted in a clear induction of PCA which was not observed with control untransfected fibroblasts. The specificity of this finding was established by inhibition experiments using monoclonal antibodies (mAbs) blocking CD40 or CD40L. PCA induced by CD40 ligation was TF-related as it was not observed in factor VII-deficient plasma and was associated with the accumulation of TF mRNA. To investigate the role of CD40/CD40L interactions in the induction of endothelial cell PCA by lymphocytes, interferon (IFN)-γ-stimulated EC were incubated with T cells in the absence or presence of anti-CD40 or anti-CD40L mAb. The 60-70% inhibition of PCA induced by these mAbs but not their isotype-matched control indicated that the CD40 pathway is involved in the induction of PCA resulting from interactions between activated HUVECs and T cells. We conclude that activation signals elicited by CD40 engagement on endothelial cells result in the induction of TF-dependent PCA. The CD40/CD40L pathway might therefore be involved in the development of prothrombic states during diseases associated with endothelial cell and T cell activation.

A simplified and low-cost one-stage chromogenic assay for tissue factor dependent procoagulant activity of endothelial cells

1995 ◽  
Vol 80 (6) ◽  
pp. 527-534 ◽  
Author(s):  
Claire Pouplard ◽  
Pascale Reverdiau-Moalic ◽  
Régis Piquemal ◽  
Hervé Watier ◽  
Yvon Lebranchu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Low Cost ◽  
Procoagulant Activity ◽  
One Stage ◽  
Chromogenic Assay

Abstract 713: S 18886 Inhibits Thromboxane A 2 Receptor-dependent Tissue Factor Expression, Procoagulant Activity And NADP(H) Oxidase Activation In Stimulated Endothelial Cells

Circulation ◽  
2007 ◽  
Vol 116 (suppl_16) ◽  
Author(s):  
Serena Del Turco ◽  
Giuseppina Basta ◽  
Guido Lazzerini ◽  
Laurent Chancharme ◽  
Laurence Lerond ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Receptor Antagonist ◽  
Tissue Factor ◽  
Molecular Mechanisms ◽  
Umbilical Vein ◽  
Factor Xa ◽  
Cell Types ◽  
Procoagulant Activity ◽  
Tp Receptor ◽  
Tnf Α

Background Tissue factor (TF) expression and surface exposure are key events in thrombosis, likely contributing to clinical events in vascular disease. Thromboxane (TX)A 2 , an unstable metabolite of arachidonic acid released from vaious cell types, is known for its pro-aggregating and vasoconstrictor properties. Cellular effects of TXA 2 are effected through the TP (TX-prostaglandin endoperoxide) receptor, also expressed in endothelial cells (EC). The TP receptor antagonist S 18886 (Terutroban) demonstrated antithrombotic and antiatherogenic effects in activated EC. As the underlying molecular mechanisms are largely unexplored, we studied the effects of TP agonism and of antagonism on TF expression and procoagulant activity in human umbilical vein endothelial cells (HUVEC), and signal transduction pathways involved. Methods and Results HUVEC ± 30 min pretreatment with the TP antagonist S 18886 were stimulated with the TP receptor agonist U 46619 or TNF-α for 6 hours. TF total expression and surface exposure were assessed by enzyme immunoassays, and TF-dependent procoagulant activity by the generation of Factor Xa. HUVEC exposed to U 46619 featured a concentration-dependent increase in TF total expression and surface exposure. These were associated with enhanced procoagulant activity. S 18886 (1 μmol/L) significantly reduced U 46619 (1 μM)-induced TF expression (−20% ± 7%, P<0.05) and procoagulant activity (−32% ± 11%, P<0.05). Interestingly, S 18886 (1 μmol/L) prevented the increase of TF expression after TNF-α (20 ng/mL) stimulation (−25% ± 9%, P<0.05). Both U 46619- and TNF-α-induced TF expression were mediated by the increase of intracellular reactive oxygen species (ROS), and this was inhibited by S 18886 (−44% ± 6% and −24% ± 5% P<0.05, respectively). S 18886 decreased the membrane association of p47-phox component of NADP(H) oxidase, accounting for the reduced production of ROS. Conclusions Our results show that endothelial TP receptor mediates TF expression, surface exposure and activity stimulated both by TP agonists and by TNF-α. This occurs through NADP(H) oxidase activation and the consequent generation of ROS. These procoagulant and oxidant pathways are inhibited by the TP receptor antagonist S 18886.

The Role of Fcγ Receptor Signaling, Tissue Factor Expression and Monocyte-Platelet Cross-Talk in the Initiation of Thrombosis by Immune Complexes.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 2167-2167
Author(s):  
Peter Casasanto ◽  
Michael P. Reilly ◽  
Ming-Lin Liu ◽  
Kevin Jon Williams ◽  
Steven E. McKenzie
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Immune Complex ◽  
Cross Talk ◽  
Immune Complexes ◽  
Thrombus Formation ◽  
Procoagulant Activity ◽  
Monocytic Cell ◽  
Platelet Factor ◽  
Activated Platelets

Abstract Thrombus formation in response to physical disruption of the vascular endothelium is an essential response to vessel injury. In contrast, thrombus formation is pathologic when the endothelium is physically intact but blood and endothelial cells are activated by inflammation. Thrombosis secondary to immune complexes is a major cause of morbidity and mortality in hospitalized patients. We recently generated and characterized the first transgenic mouse model of heparin-induced thrombocytopenia/thrombosis (HIT/T) to recapitulate the salient features of the disease and confirmed that complexes of heparin and platelet factor 4, antibodies to the complex, and FcγRIIa-dependent platelet activation are both necessary and sufficient to model the disease in vivo. It is also likely that immune complex activation of monocytes and endothelial cells occurs in HIT/T. However, the interaction between activated blood and endothelial cells and tissue factor positive microparticles (TF+-MP) that may result in thrombin generation is not clear. Recent studies (del Conde et al., Blood 2005) showed that phosphatidylserine and PSGL-1 on the surface of monocyte-derived TF+-MP enables their fusion with activated platelets. Collagen-activated platelets incubated with TF+-MP were reported to cause increased TF-VIIa procoagulant activity (PCA) compared to non-activated platelets. We hypothesized that platelets activated by HIT/T immune complexes would also result in increased TF-PCA when incubated with monocyte-derived TF+-MP. To test this hypothesis we generated TF+-MP from THP-1 cells, a human monocytic cell line, stimulated with LPS (6 hr) and A23187 (subsequent 15 min). TF+-MPs were co-incubated with untreated or agonist-treated platelets. The HIT/T immune complex was prepared by incubating optimal ratios of heparin and recombinant human PF4 with KKO, a mouse monoclonal anti-heparin-PF4 antibody. Other agonists included anti-CD9 (producing a particulate immune complex) and collagen. Using a chromogenic assay of Xa generation we found that TF+-MPs were necessary to detect TF-PCA. PCA increased by 12–25% when TF+-MPs were incubated with platelets stimulated by collagen or anti-CD9 as compared to untreated platelets. When TF+-MPs were incubated with platelets stimulated by the HIT/T immune complex, there was a 2-fold increase in the PCA. The increase in TF-PCA was observed to be proportional to the concentration of microparticles added. The results suggest an important role in platelet-monocyte cross-talk in initiating and increasing TF procoagulant activity upon immune complex stimulation.

Anesthesia in a Patient on Anticoagulant Dabigatran Presenting for Emergency Surgery

2018 ◽  
pp. 315-319
Author(s):  
Indu Kapoor ◽  
Charu Mahajan ◽  
Hemanshu Prabhakar
Keyword(s):  
Creatinine Clearance ◽  
Factor Xa ◽  
Coagulation Factors ◽  
Direct Thrombin Inhibitor ◽  
Thrombin Inhibitor ◽  
Detailed Knowledge ◽  
Safe Alternative ◽  
Routine Monitoring ◽  
Life Threatening ◽  
Improved Outcome

Dabigatran is an anticoagulant which acts by directly inhibiting the specific coagulation factors. It is used as a safe alternative to warfarin. This agent provides multiple benefits over warfarin which includes fixed dosing, lower incidence of hemorrhagic stroke, and no need of routine monitoring. Dabigatranetexilate (prodrug) is a direct thrombin inhibitor whereas rivaroxaban, edoxaban, and apixaban act by inhibiting factor Xa. Dabigatran is excreted through kidney (80%), so the dosage should be adjusted according to the creatinine clearance value. Life-threatening bleeding caused by dabigatran can be managed with its specific antidote idarucizumab to reverse its action. Factor 3 or 4 prothrombin concentrate complex may still be required in patients who bleed even after idarucizumab. Detailed knowledge about this medication can contribute to decrease in morbidity and improved outcome.

Sign in / Sign up

Export Citation Format

Share Document