scholarly journals Bacterial fibronectin-binding proteins and endothelial cell surface fibronectin mediate adherence of Staphylococcus aureus to resting human endothelial cells

Microbiology ◽  
1999 ◽  
Vol 145 (12) ◽  
pp. 3477-3486 ◽  
Author(s):  
Sharon J. Peacock ◽  
Timothy J. Foster ◽  
Brian J. Cameron ◽  
Anthony R. Berendt
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Surface ◽  
Binding Proteins ◽  
Human Endothelial Cells ◽  
Endothelial Cell Surface ◽  
Fibronectin Binding

CULTURED HUMAN ENDOTHELIAL CELLS BIND AND INTERNALIZE HIGH MOLECULAR WEIGHT KININOGEN

1987 ◽  
Author(s):  
Freek van Iwaarden ◽  
G Philip ◽  
de Groot ◽  
Bonno N Bouma
Keyword(s):  
Molecular Weight ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Surface ◽  
High Molecular Weight ◽  
Binding Sites ◽  
High Molecular Weight Kininogen ◽  
Human Endothelial Cells ◽  
Endothelial Cell Surface ◽  
Coagulation Pathway

The presence of High Molecular Weight kininogen (HMWK) was demonstrated in cultured human endothelial cells (EC) by immunofluorescence techniques. Using an enzyme linked immunosorbent assay a concentration of 58 ng HMWK/10 cells was determined. Immunoprecipitation studies performed with lysed metabolically labelled endothelial cells and mono-specific antisera directed against HMWK suggested that HMWK is not synthesized by the endothelial cells. Endothelial cells cultured in the presence of HMWK-depleted serum did not contain HMWK. This, suggests that endothelial cells can internalize HMWK. Using 125I-HMWK it was demonstrated that cultured endothelial cells bind HMWK in a time-dependent, specific and saturable.way. The cells were found to internalize 125I-HMWK, since I-HMWK was detected in solubilized endothelial cells after the cell bound 125I-HMWK had been eluted with dextran sulphate.The binding of I-HMWK required the presence of zinc ions. Optimal binding of 125I-HMWK was observed at 50 μM Zn++ . Calcium ions inhibited the Zn++ dependent binding of 125I-HMWK |25EC. In the presence of 3 mM CaCl2 the total binding of 125I-HMWK was significantly decreased, and a .concentration of 200 μM Zn++ was Required for the binding of 125I-HMWK to thecells. Higher,. Ca concentrations did not further decrease the binding of 125I-HMWK. Analysis of tl^e binding data by the ligand computer program indicated 3.2 x 10 binding sites per cell for HMWK with a Kd of 35 nM at 50 μM ZnCl2 and 1 mM CaCl2. Specify binding of HMWK did also occur at physiological plasma Zn++ concentrations. Half maximal binding was observed at HMWK concentrations of ± 105 nM at 10 μM ZnCl2 and 45 nM at 25 μM ZnCl2. The HMWK binding sites were saturatecT at HMWK concentrations of 130 nM with 1.6 x 10 molecules of HMWK bound per cell and at 80 nM with 2.8 x 10 molecules of HMWK bound per cell at 10 and 25 pM ZnCl2 respectively. These results suggest that at physiological zinc, calcium and HMWK concentrations the HMWK binding sites on the endothelial cell are saturated. The presence of HMWK on the endothelial cell surface may play a role in the initiation of the intrinsic coagulation pathway. M ZnCl2 and 45 nM at 25 μM ZnCl2. The HMWK binding sites were saturatecT at HMWK concentrations of 130 nM with 1.6 x 10 molecules of HMWK bound per cell and at 80 nM with 2.8 x 10 molecules of HMWK bound per cell at 10 and 25 μM ZnCl2 respectively. These results suggest that at physiological zinc, calcium and HMWK concentrations the HMWK binding sites on the endothelial cell are saturated. The presence of HMWK on the endothelial cell surface may play a role in the initiation of the intrinsic coagulation pathway. M ZnCl2 and 45 nM at 25 μM ZnCl2. The HMWK binding sites were saturatecT at HMWK concentrations of 130 nM with 1.6 x 10 molecules of HMWK bound per cell and at 80 nM with 2.8 x 10 molecules of HMWK bound per cell at 10 and 25 μM ZnCl2 respectively. These results suggest that at physiological zinc, calcium and HMWK concentrations the HMWK binding sites on the endothelial cell are saturated. The presence of HMWK on the endothelial cell surface may play a role in the initiation of the intrinsic coagulation pathway. M ZnCl2 and 45 nM at 25 μM ZnCl2. The HMWK binding sites were saturatecT at HMWK concentrations of 130 nM with 1.6 x 10 molecules of HMWK bound per cell and at 80 nM with 2.8 x 10 molecules of HMWK bound per cell at 10 and 25 μM ZnCl2 respectively. These results suggest that at physiological zinc, calcium and HMWK concentrations the HMWK binding sites on the endothelial cell are saturated. The presence of HMWK on the endothelial cell surface may play a role in the initiation of the intrinsic coagulation pathway.M ZnCl2 and 45 nM at 25 μM ZnCl2. The HMWK binding sites were saturatecT at HMWK concentrations of 130 nM with 1.6 x 16 molecules of HMWK bound per cell and at 80 nM with 2.8 x 106 molecules of HMWK bound per cell at 10 and 25 μM ZnCl2 respectively. These results suggest that at physiological zinc, calcium and HMWK concentrations the HMWK binding sites on the endothelial cell are saturated. The presence of HMWK on the endothelial cell surface may play a role in the initiation of the intrinsic coagulation pathway.

scholarly journals Shear Stress Prevents Fibronectin Binding Protein-Mediated Staphylococcus aureus Adhesion to Resting Endothelial Cells

2001 ◽  
Vol 69 (5) ◽  
pp. 3472-3475 ◽  
Author(s):  
Kesav Reddy ◽  
Julia M. Ross
Keyword(s):  
Staphylococcus Aureus ◽  
Endothelial Cells ◽  
Shear Stress ◽  
Endothelial Cell ◽  
Binding Proteins ◽  
Mechanical Forces ◽  
Cell Surfaces ◽  
Fibronectin Binding Protein ◽  
Fibronectin Binding

ABSTRACT Fibronectin binding proteins (FnBP) on the surface ofStaphylococcus aureus have previously been shown to mediate adherence of the organism to resting endothelial cells in static adhesion assays. However, in this study using well-defined flow assays, we demonstrate that physiologic levels of shear stress prevent FnBP-mediated adhesion of S. aureus 8325-4 to resting endothelial cells. This result suggests that mechanical forces present in vivo may influence the ability of staphylococci to bind endothelial cell surfaces.

scholarly journals Degradation of Sulphated Glycosaminoglycans at the Surface of Cultured Human Endothelial Cells by a Platelet Heparitinase

1977 ◽  
Author(s):  
C. Busch ◽  
B. Glimelius ◽  
Å Wastesson ◽  
B. Westermark
Keyword(s):  
Endothelial Cells ◽  
Cell Surface ◽  
Incubation Medium ◽  
Chondroitin Sulphate ◽  
Coagulation Factors ◽  
Platelet Lysate ◽  
Human Endothelial Cells ◽  
Endothelial Cell Surface ◽  
Sulphated Glycosaminoglycans

The non-thrombogenic property of the endothelial cell surface is a prerequisite for maintainance of blood circulation. The nature of this property is poorly understood. Recent advances in culturing techniques of endothelial cells in vitro may facilitate studies of the surface biochemistry. Human endothelial cells (EC) isolated from umbilical veins were shown to synthesize and secrete sulphated glycosaminoglycans (GAG). The recent finding of a platelet enzyme capable of degrading heparin sulphate (HS) raised the question:Can platelet lysate or a purified heparitinase detach and degrade endothelial HS? EC cultured in the presence of 35S-sulphate, produce 35S-labelled GAG which was isolated from the incubation medium from a cell associated trypsin labile pool and from a cellular pool not liberated by trypsin. After 48 hours of incorporation about 95% of 35S-GAG was found in the medium fraction, 5% in the trypsin fraction and negligible amounts in the cell fraction. In the trypsin pool (“surface fraction”) heparin sulphate comprised about 85%, while the remaining 15% consisted of chondroitin sulphate and/or dermatan sulphate. Incubation of 35S-labelled EC with platelet lysate or a partially purified preparation of the enzyme from the same source caused a marked release of cell-surface associated HS to the incubation medium as oligosaccharides. These effects could be ascribed to heparitinase activity and may alter the properties of the EC-surface and influence the interaction between these cells on one hand and blood cells or plasma components, e.g., coagulation factors on the other.

scholarly journals Dysfunction of annexin A2 contributes to hyperglycaemia-induced loss of human endothelial cell surface fibrinolytic activity

2013 ◽  
Vol 109 (06) ◽  
pp. 1070-1078 ◽  
Author(s):  
Zhanyang Yu ◽  
Xiang Fan ◽  
Ning Liu ◽  
Min Yan ◽  
Zhong Chen ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Surface ◽  
Protein Expression ◽  
Plasminogen Activator ◽  
Fibrinolytic Activity ◽  
Annexin A2 ◽  
Tissue Type ◽  
Endothelial Cell Surface ◽  
Pai 1

SummaryHyperglycaemia impairs fibrinolytic activity on the surface of endothelial cells, but the underlying mechanisms are not fully understood. In this study, we tested the hypothesis that hyperglycaemia causes dysfunction of the endothelial membrane protein annexin A2, thereby leading to an overall reduction of fibrinolytic activity. Hyperglycaemia for 7 days significantly reduced cell surface fibrinolytic activity in human brain microvascular endothelial cells (HBMEC). Hyperglycaemia also decreased tissue type plasminogen activator (t-PA), plasminogen, and annexin A2 mRNA and protein expression, while increasing plasminogen activator inhibitor-1 (PAI-1). No changes in p11 mRNA or protein expression were detected. Hyperglycaemia significantly increased AGE-modified forms of total cellular and membrane annexin A2. The hyperglycemia-associated reduction in fibrinolytic activity was fully restored upon incubation with recombinant annexin A2 (rA2), but not AGE-modified annexin A2 or exogenous t-PA. Hyperglycaemia decreased t-PA, upregulated PAI-1 and induced AGE-related disruption of annexin A2 function, all of which contributed to the overall reduction in endothelial cell surface fibrinolytic activity. Further investigations to elucidate the underlying molecular mechanisms and pathophysiological implications of A2 derivatisation might ultimately lead to a better understanding of mechanisms of impaired vascular fibrinolysis, and to development of new interventional strategies for the thrombotic vascular complications in diabetes.

Angiostatin and plasminogen share binding to endothelial cell surface actin

2005 ◽  
Vol 83 (1) ◽  
pp. 28-35 ◽  
Author(s):  
A K Dudani ◽  
M Ben-Tchavtchavadze ◽  
S Porter ◽  
E Tackaberry
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Surface ◽  
Dependent Manner ◽  
Proteolytic Fragment ◽  
Binding Motifs ◽  
Fluorescence Studies ◽  
Endothelial Cell Surface ◽  
Dose Dependent ◽  
Dependent Processes

Previous studies from this laboratory have demonstrated that plasminogen binds to endothelial cell surface-associated actin via its kringles in a dose-dependent and specific manner. The purpose of this study was to determine whether angiostatin, a proteolytic fragment of plasminogen, shares binding properties with plasminogen. Our results indicated that like plasminogen, angiostatin bound to actin in a time-, concentration-, and kringle-dependent manner. Furthermore, this binding was significantly inhibited by excess plasminogen, suggesting that both proteins shared binding motifs on the actin molecule. Fluorescence studies demonstrated that angiostatin bound to intact endothelial cells through its kringles, and this binding was also inhibited by plasminogen but not by unrelated proteins. Ligand blot analyses on endothelial cell lysates indicated that angiostatin interacted with a 42 kDa protein, which was identified as actin. Furthermore, an anti-actin antibody inhibited binding of angiostatin to endothelial cells by approximately 25%. These results suggest that angiostatin and plasminogen share binding to endothelial cell surface actin and, therefore, that angiostatin has the potential to inhibit plasmin-dependent processes such as cell migration–movement.Key words: plasminogen, angiostatin, endothelial cells, actin.

Endothelial adhesion molecules and their role in inflammation

1993 ◽  
Vol 71 (1) ◽  
pp. 76-87 ◽  
Author(s):  
C. Wayne Smith
Keyword(s):  
Endothelial Cells ◽  
Monoclonal Antibodies ◽  
Endothelial Cell ◽  
Cell Surface ◽  
Adhesion Molecules ◽  
Surrounding Tissue ◽  
Intercellular Adhesion Molecule ◽  
Intercellular Adhesion Molecule 1 ◽  
Endothelial Cell Surface

The emigration of leukocytes such as neutrophils into inflammatory sites requires adhesion to the endothelium of small venules. The initial adhesive event is margination characterized by rolling of neutrophils along the luminal surface of the endothelium. Each member of the selectin family of adhesion molecules has been shown to support neutrophil rolling under conditions of flow. E-selectin is synthesized by endothelial cells following cytokine stimulation, P-selectin is rapidly mobilized from Weibel–Palade bodies to the endothelial cell surface following stimulation with agents such as histamine, and L-selectin is constitutively expressed on the surface of leukocytes. Each selectin functions primarily as a lectin, recognizing carbohydrate structures on the leukocyte or endothelial cell surface. Once the marginated neutrophil forms a stationary adhesion with endothelial cells, it is stimulated by chemotactic factors to downregulate the selectin-based adhesion and upregulate adherence dependent on β2-integrins, principally CD11a/CD18 (LFA-1) and CD11b/CD18 (Mac-1). These adhesion molecules interact with intercellular adhesion molecule 1 (ICAM-1) and possibly other structures on the endothelial cell, and the leukocyte rapidly emigrates into surrounding tissue. Transendothelial migration in vitro is markedly inhibited by monoclonal antibodies against CD18 integrins or ICAM-1. Monoclonal antibodies against the selectins, CD18, CD11a, CD11b, and ICAM-1 have all been shown to significantly reduce the influx of neutrophils into sites of inflammation in various animal models.Key words: adhesion, integrins, selectins, leukocytes, endothelial cells.

Autoantibodies Against the Fibrinolytic Receptor, Annexin 2, in Antiphospholipid Syndrome.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 134-134
Author(s):  
Gabriela Cesarman ◽  
Nina P. Ríos-Luna ◽  
Arunkumar B. Deora ◽  
Maria del Carmen Cravioto ◽  
Donato Alarcón-Segovia ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Surface ◽  
Antiphospholipid Syndrome ◽  
Antiphospholipid Antibodies ◽  
Umbilical Vein ◽  
Healthy Individuals ◽  
Endothelial Cell Surface ◽  
Glycoprotein I ◽  
Annexin 2

Abstract The association of thrombosis, obstetric morbidity, and hemocytopenias with antiphospholipid antibodies is termed antiphospholipid syndrome. Annexin 2 is a profibrinolytic endothelial cell surface receptor that binds plasminogen, its tissue activator (tPA), and beta-2-glycoprotein-I, the main antigen for antiphospholipid antibodies. Here we evaluate annexin 2 as a target antigen in antiphospholipid syndrome. Serum samples from 434 individuals (206 patients with systemic lupus erythematosus without thrombosis, 62 with antiphospholipid syndrome, 21 with non-autoimmune thrombosis, and 145 healthy individuals) were analyzed by ELISA and immunoblotting for antiphospholipid and annexin 2 antibodies. IgG was purified and the effect of anti-annexin 2 antibodies on human umbilical vein endothelial cell activation and cell surface plasmin generation were evaluated. Anti-annexin 2 antibodies (titer >3SD) were significantly more prevalent in patients with antiphospholipid syndrome (22.6%, venous 17.5%, arterial 34.3% and mixed thrombosis 40.4%), than in healthy individuals (2.1%, p<0.001), patients with non-autoimmune thrombosis (0%, p=0.017) or patients with lupus without thrombosis (6.3%, p<0.001). Anti-annexin 2 antibodies enhanced the expression of tissue factor, a procoagulant protein, on endothelial cells (6.4 fold ± 0.13 SE), and blocked the fibrinolytic cofactor activity of purified placental annexin 2 in a tissue plasminogen activator-dependent plasmin generation assay (19 – 71%), independently of beta-2-glycoprotein-I. Similarly, cell surface plasmin generation on human umbilical vein endothelial cells was inhibited by 34–83%. We conclude that antibodies against the fibrinolytic receptor annexin 2 are significantly associated with thrombosis in antiphospholipid syndrome, and that anti-annexin 2 antibodies activate endothelial cells and inhibit endothelial cell surface-localized plasmin generation. We propose that these mechanisms contribute to the prothrombotic tendency in antiphospholipid syndrome.

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