scholarly journals Binding of human xanthine oxidase to sulphated glycosaminoglycans on the endothelial-cell surface

1993 ◽  
Vol 289 (2) ◽  
pp. 523-527 ◽  
Author(s):  
T Adachi ◽  
T Fukushima ◽  
Y Usami ◽  
K Hirano
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Membrane ◽  
Xanthine Oxidase ◽  
Aortic Endothelial Cells ◽  
Endothelial Cell Surface ◽  
Arginine Residues ◽  
Sulphated Glycosaminoglycans ◽  
Porcine Aortic Endothelial Cells ◽  
Specific Reagent

Much evidence has suggested that the superoxide generated by xanthine oxidase (XOD) within the endothelial cell triggers characteristic free-radical-mediated tissue injuries. Although it has been reported that XOD exists not only in the cytoplasm, but also on the outside surface of the endothelial cell membrane, it is not clear how XOD localizes on the outside of the plasma membrane. Purified human xanthine oxidase (h-XOD) had an affinity for heparin-Sepharose. The binding was largely independent of the pH over the physiological range, whereas it tended to increase at lower pH and to decrease at higher pH. Exposure of h-XOD to the lysine-specific reagent trinitrobenzenesulphonic acid or the arginine-specific reagent phenylglyoxal caused it to lose its affinity for heparin-Sepharose. The binding of h-XOD to heparin is apparently of electrostatic nature, and both lysine and arginine residues are involved in the binding. h-XOD was found to bind to cultured porcine aortic endothelial cells, and this binding was inhibited by the addition of heparin or pretreatment of the cells with heparinase and/or heparitinase. Intravenous injection of heparin into two healthy persons led to a prompt increase in plasma h-XOD concentration. These results suggest that XOD localizes on the outside surface of endothelial cells by association with polysaccharide chains of heparin-like proteoglycans on the endothelial-cell membranes. Superoxide extracellularly generated by XOD may injure the source-endothelial-cell membrane and also attract and activate closely appositional neutrophils, which themselves actually cause progressive oxidative damage.

scholarly journals Histidine-rich glycoprotein does not interfere with interactions between antithrombin III and heparin-like compounds on vascular endothelial cells

Blood ◽  
1989 ◽  
Vol 73 (1) ◽  
pp. 191-193 ◽  
Author(s):  
K Shimada ◽  
A Kawamoto ◽  
K Matsubayashi ◽  
T Ozawa
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Antithrombin Iii ◽  
Vascular Endothelial Cells ◽  
Anticoagulant Activity ◽  
Molar Ratio ◽  
Vascular Endothelial ◽  
Aortic Endothelial Cells ◽  
Endothelial Cell Surface

Abstract The role of histidine-rich glycoprotein in controlling heparin-like compounds on the endothelial cell surface is still unclear. The effects of this heparin-neutralizing protein on the interaction between antithrombin III and cultured porcine aortic endothelial cells were examined. Displacement of 125I-labeled antithrombin III specifically bound to endothelial cells by unlabeled histidine-rich glycoprotein was much less potent than that by unlabeled antithrombin III. One hundred- fold molar excess of histidine-rich glycoprotein displaced specific 125I-antithrombin III binding only by 20%. Furthermore, the endothelial cell-mediated acceleration of thrombin inactivation by antithrombin III was diminished by protamine sulfate, but was not affected by histidine- rich glycoprotein even at a histidine-rich glycoprotein/antithrombin III molar ratio of approximately 7:1. These data indicate that histidine-rich glycoprotein does not interfere with the interaction of endothelial cell heparin-like compounds with antithrombin III. Thus, it may not play an important role in the modulation of anticoagulant activity of endothelial cells in vivo, suggesting that the commonly accepted view of the probable function of this protein is erroneous.

Comparative protection against oxyradicals by three flavonoids on cultured endothelial cells

1997 ◽  
Vol 75 (6) ◽  
pp. 717-720 ◽  
Author(s):  
Ling-Hua Zeng ◽  
Jun Wu ◽  
Beverly Fung ◽  
Jeffrey H Tong ◽  
Donald Mickle ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Oxygen Consumption ◽  
Xanthine Oxidase ◽  
Inhibitory Effect ◽  
Control Group ◽  
Aortic Endothelial Cells ◽  
Cytochrome C Reduction ◽  
Porcine Aortic Endothelial Cells ◽  
Aortic Endothelial

Oxygen-derived free radicals are known to injure the endothelium of aorta in diverse disorders. In this study we compared the cytoprotective effects of three flavonoids against oxyradical damage to porcine aortic endothelial cells in vitro. Cultured porcine aortic endothelial cells were exposed to oxyradicals generated by xanthine oxidase - hypoxanthine (XO-HP). The cytoprotective activities of morin, quercetin, and catechin on these systems were compared using established morphologic criteria. The results in the XO-HP system showed that morin at 0.125, 0.25, and 0.5 mM delayed cell necrosis to 27.4 ± 1.3, 46.8 ± 1.8, and longer than 70 min, respectively, compared with 12.0 ± 1.3 min in the control group. These degrees of protection were significantly stronger than those provided by quercetin and catechin at corresponding concentrations (p < 0.01). Morin and quercetin were moderate inhibitors of xanthine oxidase on the basis of the oxygen consumption rate, whereas catechin at the same concentrations had little inhibitory effect. The data from uric acid formation and cytochrome c reduction were consistent with the oxygen consumption measurement for the three flavonoids. Key words: flavonoids, oxyradicals, aortic endothelial cells.

scholarly journals Histidine-rich glycoprotein does not interfere with interactions between antithrombin III and heparin-like compounds on vascular endothelial cells

Blood ◽  
1989 ◽  
Vol 73 (1) ◽  
pp. 191-193
Author(s):  
K Shimada ◽  
A Kawamoto ◽  
K Matsubayashi ◽  
T Ozawa
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Antithrombin Iii ◽  
Vascular Endothelial Cells ◽  
Anticoagulant Activity ◽  
Molar Ratio ◽  
Vascular Endothelial ◽  
Aortic Endothelial Cells ◽  
Endothelial Cell Surface

The role of histidine-rich glycoprotein in controlling heparin-like compounds on the endothelial cell surface is still unclear. The effects of this heparin-neutralizing protein on the interaction between antithrombin III and cultured porcine aortic endothelial cells were examined. Displacement of 125I-labeled antithrombin III specifically bound to endothelial cells by unlabeled histidine-rich glycoprotein was much less potent than that by unlabeled antithrombin III. One hundred- fold molar excess of histidine-rich glycoprotein displaced specific 125I-antithrombin III binding only by 20%. Furthermore, the endothelial cell-mediated acceleration of thrombin inactivation by antithrombin III was diminished by protamine sulfate, but was not affected by histidine- rich glycoprotein even at a histidine-rich glycoprotein/antithrombin III molar ratio of approximately 7:1. These data indicate that histidine-rich glycoprotein does not interfere with the interaction of endothelial cell heparin-like compounds with antithrombin III. Thus, it may not play an important role in the modulation of anticoagulant activity of endothelial cells in vivo, suggesting that the commonly accepted view of the probable function of this protein is erroneous.

Convective Mass Transfer Effects on the Intracellular Calcium Response of Endothelial Cells

1992 ◽  
Vol 114 (3) ◽  
pp. 321-326 ◽  
Author(s):  
M. U. Nollert ◽  
L. V. McIntire
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Surface ◽  
Flow Rate ◽  
Parallel Plate ◽  
Cytosolic Calcium ◽  
Local Concentration ◽  
Aortic Endothelial Cells ◽  
Endothelial Cell Surface ◽  
Governing Differential Equation

Endothelial cells, which line the vasculature, respond to specific agonists such as adenosine triphosphate (ATP) by elevating cytosolic calcium levels and increasing production of the vasoactive compounds, prostacyclin and endothelial derived relaxing factor (EDRF). Endothelial cells express ecto-enzymes which metabolize ATP. If the activity of these enzymes is sufficiently high, then the concentration of ATP near the endothelial cell surface can be substantially lower than the bulk concentration. The ATP concentration is determined by a balance between the convection of fresh ATP from upstream and the degradation of ATP by the endothelial cells. In this report, we present a parallel plate flow system for measurement of cytosolic calcium levels ([Ca2+]i) of individual bovine aortic endothelial cells with the calcium sensitive fluorescent dye, fura-2. The cells respond to increases in the flow rate by increasing [Ca2+]i if there is ATP present in the perfusing buffer, but not in the absence of ATP. The amount of agonist in the perfusing fluid near the endothelial cell surface is estimated by solving the governing differential equation for the concentration profile of ATP in the parallel plate flow geometry. The solution indicates that one mechanism endothelial cells may use to detect changes in the flow rate is to respond to the change in the local concentration of agonist.

scholarly journals Identification of a heparin-binding protein using monoclonal antibodies that block heparin binding to porcine aortic endothelial cells

1995 ◽  
Vol 311 (2) ◽  
pp. 461-469 ◽  
Author(s):  
W A Patton ◽  
C A Granzow ◽  
L A Getts ◽  
S C Thomas ◽  
L M Zotter ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Monoclonal Antibodies ◽  
Endothelial Cell ◽  
Binding Protein ◽  
Heparan Sulphate ◽  
Heparin Binding ◽  
Aortic Endothelial Cells ◽  
Heparin Binding Protein ◽  
Porcine Aortic Endothelial Cells ◽  
Aortic Endothelial

The binding of heparin or heparan sulphate to a variety of cell types results in specific changes in cell function. Endothelial cells treated with heparin alter their synthesis of heparan sulphate proteoglycans and extracellular matrix proteins. In order to identify a putative endothelial cell heparin receptor that could be involved in heparin signalling, anti-(endothelial cell) monoclonal antibodies that significantly inhibit heparin binding to endothelial cells were prepared. Four of these antibodies were employed in affinity-chromatographic isolation of a heparin-binding protein from detergent-solubilized endothelial cells. The heparin-binding protein isolated from porcine aortic endothelial cells using four different monoclonal antibodies has an M(r) of 45,000 assessed by SDS/PAGE. The 45,000-M(r) heparin-binding polypeptide is isolated as a multimer. The antibody-isolated protein binds to heparin-affinity columns as does the pure 45,000-M(r) polypeptide, consistent with its identification as a putative endothelial heparin receptor.

scholarly journals Differential effects of hydrogen peroxide on indices of endothelial cell function.

1984 ◽  
Vol 159 (2) ◽  
pp. 592-603 ◽  
Author(s):  
A Ager ◽  
J L Gordon
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Xanthine Oxidase ◽  
Cell Function ◽  
Threshold Concentration ◽  
Endothelial Cell Function ◽  
Aortic Endothelial Cells ◽  
Prostaglandin Production ◽  
Chromium Release ◽  
Sublethal Doses

The responses of pig aortic endothelial cells to sublethal doses of potentially toxic stimuli were investigated by monitoring K+ efflux, prostaglandin production, and the release of cytoplasmic purines. Xanthine plus xanthine oxidase reversibly stimulated these three parameters of endothelial cell function at doses that were not cytotoxic, as measured by chromium release, adenine uptake, and vital dye exclusion. The effects of xanthine plus xanthine oxidase were inhibited by catalase but not by superoxide dismutase, suggesting that H2O2 was responsible. Reagent H2O2 also reversibly stimulated K+ efflux, prostaglandin production, and the release of purines. The threshold concentration of H2O2 for these effects was approximately 10 microM, which was at least 30-fold lower than that which caused cytotoxicity. In addition to the direct effect of H2O2 in stimulating prostaglandin production (PGI2 and PGE2), prior exposure of endothelial cells to lower doses of H2O2 (less than 0.1 microM) at high oxygen tension inhibited the subsequent stimulation of prostaglandin production by ATP, A23187, and H2O2 itself. We conclude that H2O2 has substantial effects on endothelial physiology at doses up to 3,000-fold lower than those which induce cytotoxicity.

scholarly journals Orai–vascular endothelial-cadherin signaling complex regulates high-glucose exposure-induced increased permeability of mouse aortic endothelial cells

2021 ◽  
Vol 9 (1) ◽  
pp. e002085
Author(s):  
Yuan Wei ◽  
Suwen Bai ◽  
YanHeng Yao ◽  
Wenxuan Hou ◽  
Junwei Zhu ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Cell Membrane ◽  
High Glucose ◽  
Expression Level ◽  
Vascular Endothelial ◽  
Aortic Endothelial Cells ◽  
Downstream Signaling ◽  
Signaling Complex ◽  
Aortic Endothelium ◽  
Aortic Endothelial

IntroductionDiabetes-associated endothelial barrier function impairment might be linked to disturbances in Ca2+ homeostasis. To study the role and molecular mechanism of Orais–vascular endothelial (VE)-cadherin signaling complex and its downstream signaling pathway in diabetic endothelial injury using mouse aortic endothelial cells (MAECs).Research design and methodsThe activity of store-operated Ca2+ entry (SOCE) was detected by calcium imaging after 7 days of high-glucose (HG) or normal-glucose (NG) exposure, the expression levels of Orais after HG treatment was detected by western blot analysis. The effect of HG exposure on the expression of phosphorylated (p)-VE-cadherin and VE-cadherin on cell membrane was observed by immunofluorescence assay. HG-induced transendothelial electrical resistance was examined in vitro after MAECs were cultured in HG medium. FD-20 permeability was tested in monolayer aortic endothelial cells through transwell permeability assay. The interactions between Orais and VE-cadherin were detected by co-immunoprecipitation and immunofluorescence technologies. Immunohistochemical experiment was used to detect the expression changes of Orais, VE-cadherin and p-VE-cadherin in aortic endothelium of mice with diabetes.Results(1) The expression levels of Orais and activity of SOCE were significantly increased in MAECs cultured in HG for 7 days. (2) In MAECs cultured in HG for 7 days, the ratio of p-VE-cadherin to VE-cadherin expressed on the cell membrane and the FD-20 permeability in monolayer endothelial cells increased, indicating that intercellular permeability increased. (3) Orais and VE-cadherin can interact and enhance the interaction ratio through HG stimulation. (4) In MAECs cultured with HG, the SOCE activator ATP enhanced the expression level of p-VE-cadherin, and the SOCE inhibitor BTP2 decreased the expression level of p-VE-cadherin. (5) Significantly increased expression of p-VE-cadherin and Orais in the aortic endothelium of mice with diabetes.ConclusionHG exposure stimulated increased expression of Orais in endothelial cells, and increased VE-cadherin phosphorylation through Orais–VE-cadherin complex and a series of downstream signaling pathways, resulting in disruption of endothelial cell junctions and initiation of atherosclerosis.

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