scholarly journals Platelet effects on tissue factor and fibrinolytic inhibition of cultured human fibroblasts and vascular cells

Blood ◽  
1982 ◽  
Vol 60 (1) ◽  
pp. 140-147 ◽  
Author(s):  
PE Smariga ◽  
JR Maynard
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Human Fibroblasts ◽  
Cellular Tissue ◽  
Cultured Human Fibroblasts ◽  
Platelet Concentration ◽  
The Media ◽  
Low Levels ◽  
Coagulation Pathway

Abstract Platelets stimulate tissue factor, the initiator of the extrinsic coagulation pathway, and increase fibrinolytic inhibition in fibroblasts grown in vitro. Cellular tissue factor increases an average of 2.8-fold over the control levels after a 6-hr incubation with platelets, and no activity is present in the media. Fibrinolytic inhibition is stimulated in both the fibroblasts and their media in the presence of platelets and accumulates throughout a 24-hr incubation. Neither leukocytes nor erythrocytes stimulate these changes. Both tissue factor and fibrinolytic inhibition increases are dependent on platelet concentration and are blocked by inhibitors of RNA or protein synthesis. Control smooth muscle cells have higher tissue factor and fibrinolytic inhibition than fibroblasts, but their response to the presence of platelets is similar. Confluent monolayers of endothelial cells have very low levels of tissue factor that are not altered by the presence of platelets. However, the ability of endothelial cells to inhibit fibrinolysis is enhanced by the presence of platelets. The fraction that stimulates tissue factor and fibrinolytic inhibition is distinct from platelet-derived growth factor and from the fraction that enhances leukocyte tissue factor. It is associated with an insoluble, nonmitogenic fraction that is not inactivated by phospholipase C, or diisopropylfluorophosphate, nor is it chloroform:methanol extractable. Platelets are a physiologic modulator for both cellular tissue factor and the fibrinolytic system in vitro.

scholarly journals Platelet effects on tissue factor and fibrinolytic inhibition of cultured human fibroblasts and vascular cells

Blood ◽  
1982 ◽  
Vol 60 (1) ◽  
pp. 140-147
Author(s):  
PE Smariga ◽  
JR Maynard
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Human Fibroblasts ◽  
Cellular Tissue ◽  
Cultured Human Fibroblasts ◽  
Platelet Concentration ◽  
The Media ◽  
Low Levels ◽  
Coagulation Pathway

Platelets stimulate tissue factor, the initiator of the extrinsic coagulation pathway, and increase fibrinolytic inhibition in fibroblasts grown in vitro. Cellular tissue factor increases an average of 2.8-fold over the control levels after a 6-hr incubation with platelets, and no activity is present in the media. Fibrinolytic inhibition is stimulated in both the fibroblasts and their media in the presence of platelets and accumulates throughout a 24-hr incubation. Neither leukocytes nor erythrocytes stimulate these changes. Both tissue factor and fibrinolytic inhibition increases are dependent on platelet concentration and are blocked by inhibitors of RNA or protein synthesis. Control smooth muscle cells have higher tissue factor and fibrinolytic inhibition than fibroblasts, but their response to the presence of platelets is similar. Confluent monolayers of endothelial cells have very low levels of tissue factor that are not altered by the presence of platelets. However, the ability of endothelial cells to inhibit fibrinolysis is enhanced by the presence of platelets. The fraction that stimulates tissue factor and fibrinolytic inhibition is distinct from platelet-derived growth factor and from the fraction that enhances leukocyte tissue factor. It is associated with an insoluble, nonmitogenic fraction that is not inactivated by phospholipase C, or diisopropylfluorophosphate, nor is it chloroform:methanol extractable. Platelets are a physiologic modulator for both cellular tissue factor and the fibrinolytic system in vitro.

scholarly journals Differential roles of factors IX and XI in murine placenta and hemostasis under conditions of low tissue factor

2020 ◽  
Vol 4 (1) ◽  
pp. 207-216 ◽  
Author(s):  
Steven P. Grover ◽  
Clare M. Schmedes ◽  
Alyson C. Auriemma ◽  
Emily Butler ◽  
Molly L. Parrish ◽  
...  
Keyword(s):  
Tissue Factor ◽  
Thrombin Generation ◽  
Critical Role ◽  
Wild Type ◽  
Genetic Studies ◽  
Acute Hemorrhage ◽  
Expected Frequency ◽  
Low Levels ◽  
Coagulation Pathway

Abstract The intrinsic tenase complex (FIXa-FVIIIa) of the intrinsic coagulation pathway and, to a lesser extent, thrombin-mediated activation of FXI, are necessary to amplify tissue factor (TF)-FVIIa–initiated thrombin generation. In this study, we determined the contribution of murine FIX and FXI to TF-dependent thrombin generation in vitro. We further investigated TF-dependent FIX activation in mice and the contribution of this pathway to hemostasis. Thrombin generation was decreased in FIX- but not in FXI-deficient mouse plasma. Furthermore, injection of TF increased levels of FIXa-antithrombin complexes in both wild-type and FXI−/− mice. Genetic studies were used to determine the effect of complete deficiencies of either FIX or FXI on the survival of mice expressing low levels of TF. Low-TF;FIX−/y male mice were born at the expected frequency, but none survived to wean. In contrast, low-TF;FXI−/− mice were generated at the expected frequency at wean and had a 6-month survival equivalent to that of low-TF mice. Surprisingly, a deficiency of FXI, but not FIX, exacerbated the size of blood pools in low-TF placentas and led to acute hemorrhage and death of some pregnant dams. Our data indicate that FIX, but not FXI, is essential for survival of low-TF mice after birth. This finding suggests that TF-FVIIa–mediated activation of FIX plays a critical role in murine hemostasis. In contrast, FXI deficiency, but not FIX deficiency, exacerbated blood pooling in low-TF placentas, indicating a tissue-specific requirement for FXI in the murine placenta under conditions of low TF.

scholarly journals Tissue Factor Expression in Bovine Endothelial Cells Induced by Pasteurella haemolytica Lipopolysaccharide and Interleukin-1

1994 ◽  
Vol 31 (1) ◽  
pp. 55-60 ◽  
Author(s):  
M. A. Breider ◽  
Z. Yang
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Interleukin 1 ◽  
Cellular Tissue ◽  
Pasteurella Haemolytica ◽  
Dependent Manner ◽  
Tissue Factor Expression ◽  
Vascular Thrombosis ◽  
Factor Expression

Pasteurella haemolytica in cattle produces fibrinohemorrhagic pleuropneumonia characterized by extensive pulmonary microvascular thrombosis and parenchymal necrosis. The purpose of this in vitro study was to determine if P. haemolytica lipopolysaccharide (LPS) promotes vascular thrombosis by inducing a procoagulant state in vascular endothelial cells. After treatment of confluent monolayers of bovine pulmonary artery endothelial cells with various concentrations of either P. haemolytica LPS or Escherichia coli LPS, the procoagulant activity of the endothelial cells was determined using a chromogenic assay dependent on cellular tissue factor expression. The LPS treatment induced significant increases in cellular tissue factor expression in a LPS concentration- and time-dependent manner. Highest levels of tissue factor were present at 22 hours after treatment, although high LPS concentrations induced moderate tissue factor levels at 5 hours after treatment. Interleukin-1 also induced tissue factor expression in endothelial cells and enhanced the LPS-induced effects. This interleukin-1 effect could be diminished by concurrent use of an interleukin-1 receptor antagonist. These results demonstrate that LPS and cytokine promotion of a procoagulant state in endothelial cells occurs in vitro. Similar mechanisms may play a role in P. haemolytica-mediated pulmonary vascular thrombosis.

scholarly journals Endotheliopathy in septic conditions: mechanistic insight into intravascular coagulation

Critical Care ◽  
2021 ◽  
Vol 25 (1) ◽  
Author(s):  
Takashi Ito ◽  
Midori Kakuuchi ◽  
Ikuro Maruyama
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Coagulation Factors ◽  
Intravascular Coagulation ◽  
Outer Leaflet ◽  
Tissue Factor Pathway ◽  
Recombinant Thrombomodulin ◽  
Coagulation Pathway ◽  
Insight Into

AbstractEndothelial cells play a key role in maintaining intravascular patency through their anticoagulant properties. They provide a favorable environment for plasma anticoagulant proteins, including antithrombin, tissue factor pathway inhibitor, and protein C. Under septic conditions, however, the anticoagulant properties of endothelial cells are compromised. Rather, activated/injured endothelial cells can provide a scaffold for intravascular coagulation. For example, the expression of tissue factor, an important initiator of the coagulation pathway, is induced on the surface of activated endothelial cells. Phosphatidylserine, a high-affinity scaffold for gamma-carboxyglutamate domain containing coagulation factors, including FII, FVII, FIX, and FX, is externalized to the outer leaflet of the plasma membrane of injured endothelial cells. Hemodilution decreases not only coagulation factors but also plasma anticoagulant proteins, resulting in unleashed activation of coagulation on the surface of activated/injured endothelial cells. The aberrant activation of coagulation can be suppressed in part by the supplementation of recombinant antithrombin and recombinant thrombomodulin. This review aims to overview the physiological and pathological functions of endothelial cells along with proof-of-concept in vitro studies. The pathophysiology of COVID-19-associated thrombosis is also discussed.

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 An endothelialized urothelial cell-seeded tubular graft for urethral replacement

2013 ◽  
Vol 7 (1-2) ◽  
pp. 4 ◽  
Author(s):  
Annie Imbeault ◽  
Geneviève Bernard ◽  
Alexandre Rousseau ◽  
Amélie Morissette ◽  
Stéphane Chabaud ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Cells ◽  
Human Fibroblasts ◽  
Surgical Techniques ◽  
Urethral Reconstruction ◽  
Full Thickness ◽  
Urothelial Cells ◽  
The Impact ◽  
Histology And Immunohistochemistry

Introduction: Many efforts are used to improve surgical techniques and graft materials for urethral reconstruction. We developed an endothelialized tubular structure for urethral reconstruction.Methods: Two tubular models were created in vitro. Human fibroblasts were cultured for 4 weeks to form fibroblast sheets. Then, endothelial cells (ECs) were seeded on the fibroblast sheets and wrapped around a tubular support to form a cylinder for the endothelialized tubular urethral model (ET). No ECs were added in the standard tubular model (T). After 21 days of maturation, urothelial cells were seeded into the lumen of both models. Constructs were placed under perfusion in a bioreactor for 1 week. At several times,histology and immunohistochemistry were performed on grafted nude mice to evaluate the impact of ECs on vascularization.Results: Both models produced an extracellular matrix, without exogenous material, and developed a pseudostratified urothelium. Seven days after the graft, mouse red blood cells were present only in the outer layers in T model, but in the full thickness of ET model. After 14 days, erythrocytes were present in both models, but in a greater proportion in ET model. At day 28, both models were well-vascularized, with capillary-like structures in the wholethickness of the tubes.Conclusion: Incorporating endothelial cells was associated with an earlier vascularization of the grafts, which could decrease the necrosis of the transplanted tissue. As those models can be elaborated with the patient’s cells, this tubular urethral graft would be unique in its autologous property.

scholarly journals Human umbilical vein endothelial cells secrete transcobalamin II

Blood ◽  
1990 ◽  
Vol 75 (1) ◽  
pp. 251-254 ◽  
Author(s):  
R Carmel ◽  
SM Neely ◽  
RB Jr Francis
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Human Fibroblasts ◽  
Surrounding Medium ◽  
Human Leukemia ◽  
Human Umbilical Vein ◽  
Intracellular Content ◽  
Umbilical Vein Endothelial Cells ◽  
Transcobalamin Ii

Abstract Transcobalamin II (TC II) is essential for cellular uptake of cobalamin. However, the origin of this transport protein is controversial and many organ sources have been suggested. We studied human umbilical vein endothelial cells cultured in vitro. The cells contained TC II (2.3 pmol/10(8) cells) and released progressively increasing amounts of the protein into the surrounding medium during the 3-day incubation period. This release exceeded the starting intracellular content of TC II. In contrast, endothelial cells did not contain or elaborate R binder, the other major circulating binding protein for cobalamin, Cycloheximide inhibited the elaboration of TC II, suggesting that the endothelial cells synthesize the protein. Thrombin, which stimulates tissue plasminogen activator release, did not enhance TC II release, and neither did endotoxin or mellitin. However, thrombin did appear to partially protect TC II release from inhibition by cycloheximide. Among other cells studied, human fibroblasts also released TC II into the incubation medium, while K562 human leukemia cells, ARH-77 and HS Sultan human plasma cell lines, and Raji strain lymphoblasts did not. The data suggest that endothelial cells are an important source of the metabolically crucial TC II.

scholarly journals Expression of tissue factor pathway inhibitor by cultured endothelial cells in response to inflammatory mediators

Blood ◽  
1992 ◽  
Vol 79 (12) ◽  
pp. 3219-3226 ◽  
Author(s):  
A Ameri ◽  
MN Kuppuswamy ◽  
S Basu ◽  
SP Bajaj
Keyword(s):  
Endothelial Cells ◽  
Blood Serum ◽  
Tissue Factor ◽  
Inflammatory Mediators ◽  
Whole Blood ◽  
Tissue Factor Pathway Inhibitor ◽  
Binding Motif ◽  
Cultured Endothelial Cells ◽  
The Media ◽  
Tissue Factor Pathway

Abstract We recently proposed that endothelium may represent the primary physiologic site of synthesis of the tissue factor pathway inhibitor (TFPI). In support of this conclusion, we have now found that the poly(A)+ RNAs obtained from rabbit and bovine lung tissues contain abundant amounts of TFPI messenger RNAs (mRNAs), whereas the poly(A)+ RNAs obtained from the liver of these animals contain less than 5% of that found in the lung tissues. Because inflammatory mediators are known to upregulate tissue factor (TF) expression by the endothelium, we have examined the effect of these agents on the TFPI expression by the cultured endothelial cells. When cultured human umbilical vein endothelial cells were stimulated (in 10% fetal bovine serum) with phorbol myristate acetate (PMA), endotoxin, interleukin-1, or tumor necrosis factor-alpha, the TF mRNA increased approximately 7- to 10- fold within 2 to 4 hours. Unstimulated cells constitutively expressed TFPI mRNA and its levels either did not change or increased slightly (up to 1.5-fold) upon stimulation with these inflammatory agents. TF mRNA abruptly declined to a negligible level and the TFPI mRNA returned essentially to the basal level at approximately 24 hours. The membrane- bound TF clotting activity of induced cells peaked between 4 and 8 hours, and finally declined. The cumulative TFPI activity secreted into the media was either unchanged or slightly higher in the induced cell cultures as compared with that present in the noninduced cultures. Endothelial cells were also cultured in 10% heat-inactivated human serum derived from plasma or whole blood. TFPI secreted into the media containing whole blood serum was consistently higher (approximately 1.5- fold at 8 hours) than that secreted into the media supplemented with serum obtained from plasma lacking the formed elements; these cells also expressed similarly increased levels of TFPI mRNA. Moreover, PMA- stimulated cells cultured in whole blood serum expressed modestly increased levels of TFPI mRNA (approximately 1.5-fold); supernatants from these cells also contained similarly increased TFPI activity. Cumulatively, our data indicate that, unlike thrombomodulin and fibrinolytic enzymes synthesized by the endothelial cells, TFPI synthesis is not downregulated and may be slightly upregulated during an inflammatory response. Inspection of the 5′ flanking region of the TFPI gene showed a conserved GATA-binding motif located approximately 400 bp upstream of the proposed transcription initiation site(s). This motif by binding to the GATA-2 transcriptional factor may keep the endothelium in an ‘on’ state for constitutive expression of TFPI.(ABSTRACT TRUNCATED AT 400 WORDS)

THE "LUPUS ANTICOAGULANT" INDUCES FUNCTIONAL CHANGES IN ENDOTHELIAL CELLS AND PLATELETS

1987 ◽  
Author(s):  
Anna E Schorer ◽  
Kathleen V Watson
Keyword(s):  
Endothelial Cells ◽  
Tissue Factor ◽  
Lupus Anticoagulant ◽  
Inhibitory Effect ◽  
Clinical Syndrome ◽  
Factor Vii ◽  
Factor X ◽  
Functional Changes ◽  
Thrombin Activation

The presence of the "lupus anticoagulant" (LA) predicts a clinical syndrome of excessive arterial, venous and microvascu-lar thrombosis. LA is an antibody which reacts with negatively charged phospholipid (PL) species in vitro. Since PL is involved in many aspects of the regulation of thrombosis, we postulated that LA might modify one or more of the membrane-(PL-dependent reactions of platelets and endothelial cells (EC). Blood samples from 20 patients with a history of thrombosis were tested for the presence of LA (kaolin PTT) and titres determined. LA-positive (LA+) sera and plasma were compared to LA-negative (LA−) samples from normal donors (n=6) or patients who had lupus but no clinical thrombosis (n=4). These specimens were tested in a panel of assays. The thrombin-stimulated release of prostacyclin (PG12) from cultured human EC was markedly reduced (52%±12.5 s.e.) by preincubation of the EC with LA+ sera (30 minutes). Purified LA+ IgG from one patient reproduced this effect. Thrombin induction of EC synthesis of the procoagulant, tissue factor-which is dissociable from prostaglandin metabolism-was also inhibited by LA+ sera. Normal platelets incubated in LA+ plasma became refractory to thrombin (1 unit/ml) but retained their responsiveness to epinephrine and ADP. The reduced responsiveness to thrombin was not due to altered (specific or total) binding of thrombin. The cleavage of Factor X by Factor VII requires PL as a co-factor for the EC procoagulant, tissue factor (TF). Unlike the inhibitory effect of LA on thrombin activation of EC and platelets, this distinct membrane-(PL-) dependent function was variably enhanced by LA+ sera. Brief (20 min) exposure of EC to LA+ sera increased TF co-catalysis of Factor VII cleavage of Factor X (measured by chromogenic Xa substrate, S-2222) by up to 10 fold (p<0.05, unpaired t test). This effect was not the result of EC disruption or changes in whole-cell TF content. These data suggest multiple, complex and heterogenous effects of LA, including impaired production of PG12, impaired EC modulation, and heightened ability of endogenous EC tissue factor to initiate coagulation. These (and perhaps other) membrane-dependent effects may contribute to the tendency of LA+ patients to develop clots.

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