Characterization and quantitation of morphological changes induced in bovine endothelial cells by interleukin-2 (IL-2)

Endothelial cells respond to endotoxins, arachidonic acid, and interferon-alpha by exhibiting a time- and dose-dependent production of prostacyclin which is accompanied by morphological changes in the endothelial cells. Recent studies showed that IL-2 also stimulates production of prostacyclin by endothelial cells. Therefore, the effect of interleukin-2 on the morphology of endothelial cells in vitro and ex vivo was investigated by LM, SEM and TEM. Quantitation of the cell changes in vitro was performed by image analysis and was correlated with time- and dosedependent production of prostacyclin.

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The d-Enantiomer Form of Indobufen Totally Accounts for the Anti-Cyclooxygenase and Antiplatelet Activity Ex Vivo and for the Increase in Bleeding Time by Indobufen in Man

Thrombosis and Haemostasis ◽

10.1055/s-0038-1648422 ◽

1992 ◽

Vol 67 (02) ◽

pp. 258-263 ◽

Cited By ~ 12

Author(s):

Raffaele De Caterina ◽

Rosa Sicari ◽

An Yan ◽

Walter Bernini ◽

Daniela Giannessi ◽

...

Keyword(s):

Arachidonic Acid ◽

Platelet Aggregation ◽

Plasma Levels ◽

Bleeding Time ◽

Ex Vivo ◽

Random Sequence ◽

Antiplatelet Agent ◽

Antiplatelet Drug ◽

Double Blind

SummaryIndobufen is an antiplatelet drug able to inhibit thromboxane production and cyclooxygenase-dependent platelet aggregation by a reversible inhibition of cyclooxygenase. Indobufen exists in two enantiomeric forms, of which only d-indobufen is active in vitro in inhibiting cyclooxygenase. In order to verify that also inhibition of platelet function is totally accounted for by d-indobufen, ten patients with proven coronary artery disease (8 male, 2 female, age, mean ± S.D., 58.7 ± 7.5 years) were given, in random sequence, both 100 mg d-indobufen and 200 mg dl-indobufen as single administrations in a double-blind crossover design study with a washout period between treatments of 72 h. In all patients thromboxane (TX) B2 generation after spontaneous clotting (at 0, 1, 2, 4, 6, 8, 12, 24 h), drug plasma levels (at the same times), platelet aggregation in response to ADP, adrenaline, arachidonic acid, collagen, PAF, and bleeding time (at 0, 2, 12 h) were evaluated after each treatment. Both treatments determined peak inhibition of TXB2 production at 2 h from administration, with no statistical difference between the two treatments (97 ±3% for both treatments). At 12 h inhibition was 87 ± 6% for d-indobufen and 88 ± 6% for dl-indobufen (p = NS). Inhibition of TXB2 production correlated significantly with plasma levels of the drugs. Maximum inhibitory effect on aggregation was seen in response to collagen 1.5 pg/ml (63 ± 44% for d-indobufen and 81 ± 22% for dl-indobufen) and arachidonic acid 0.5-2 mM (78 ± 34% for d-indobufen and 88 ± 24% for dl-indobufen) at 2 h after each administration. An effect of both treatments on platelet aggregation after 12 h was present only for adrenaline 2 μM (55 ± 41% for d-indobufen and 37 ± 54% for dl-indobufen), collagen 1.5 pg/ml (69 ± 30% for d-indobufen and 51 ± 61% for dl-indobufen), arachidonic acid 0.5-2 mM (56 ± 48% for d-indobufen and 35 ± 49% for dl-indobufen). The extent of inhibition of TX production and the extent of residual platelet aggregation were never significantly different between treatments. Bleeding time prolongation was similar in the two treatment groups without showing a pronounced and long lasting effect (from 7.0 ± 2.0 min to 10.0 ± 3.0 min at 2 h and 8.0 ± 2.0 min at 12 h for d-indobufen; from 6.0 ±1.0 min to 8.5 ± 2.0 min at 2 h and 8.0 ± 1.0 min at 12 h for dl-indobufen). These results demonstrate that the biological activity of dl-indobufen as an antiplatelet agent in vivo is totally accounted for by d-indobufen.

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Effects of Dipyrone on Prostaglandin Production by Human Platelets and Cultured Bovine Aortic Endothelial Cells

Thrombosis and Haemostasis ◽

10.1055/s-0038-1657338 ◽

1983 ◽

Vol 49 (02) ◽

pp. 132-137 ◽

Cited By ~ 16

Author(s):

A Eldor ◽

G Polliack ◽

I Vlodavsky ◽

M Levy

Keyword(s):

Endothelial Cells ◽

Arachidonic Acid ◽

Competitive Inhibition ◽

Inhibitory Effect ◽

Human Platelets ◽

Ionophore A23187 ◽

Aortic Endothelial Cells ◽

Bovine Aortic Endothelial Cells ◽

Aortic Endothelial

SummaryDipyrone and its metabolites 4-methylaminoantipyrine, 4-aminoantipyrine, 4-acetylaminoantipyrine and 4-formylaminoan- tipyrine inhibited the formation of thromboxane A2 (TXA2) during in vitro platelet aggregation induced by ADP, epinephrine, collagen, ionophore A23187 and arachidonic acid. Inhibition occurred after a short incubation (30–40 sec) and depended on the concentration of the drug or its metabolites and the aggregating agents. The minimal inhibitory concentration of dipyrone needed to completely block aggregation varied between individual donors, and related directly to the inherent capacity of their platelets to synthesize TXA2.Incubation of dipyrone with cultured bovine aortic endothelial cells resulted in a time and dose dependent inhibition of the release of prostacyclin (PGI2) into the culture medium. However, inhibition was abolished when the drug was removed from the culture, or when the cells were stimulated to produce PGI2 with either arachidonic acid or ionophore A23187.These results indicate that dipyrone exerts its inhibitory effect on prostaglandins synthesis by platelets or endothelial cells through a competitive inhibition of the cyclooxygenase system.

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Appraisal of Immunological Impacts of Melaleuca leucadendra Extract over Macrophage Performance in Vitro

International Journal of Veterinary Science ◽

10.37422/ijvs/20.051 ◽

2020 ◽

Keyword(s):

Nitric Oxide ◽

Interleukin 2 ◽

Dependent Manner ◽

Reduced Pressure ◽

Medical Plant ◽

Dose Dependent ◽

Level Production ◽

Melaleuca Leucadendra ◽

Phagocytic Killing

This trial research was performed to discuss the immune-influence of Melaleuca leucadendra ‘paper-bark tree’ dried leaves which is an important medical plant known in many regions in the world. The leaves were dissolved in a mixture of (ethanol + water) (3:1) mixture, then filtered, evaporated and dried under reduced pressure to obtain leaves extract. The macrophages of blood derived origin were provided from rats and mixed with three different leaves extracts doses in tissue culture plates and incubated then stained with fluorescent acridine orange and examined under fluorescent microscope to assess the phagocytic and killing potency. The wells contents were aspirated and assayed for nitric oxide and interleukin-2 levels. The results displayed an obvious increase in phagocytic, killing performance as well as nitric oxide and IL-2 level production than control in a dose dependent manner. The obtained results suggested the immune-stimulant impact of the paper-bark tree leaves.

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Antiangiogenic effects of phospholipase A2 Lys49 BnSP-7 from Bothrops pauloensis snake venom on endothelial cells: An in vitro and ex vivo approach

Toxicology in Vitro ◽

10.1016/j.tiv.2021.105099 ◽

2021 ◽

Vol 72 ◽

pp. 105099

Author(s):

Lorena Polloni ◽

Fernanda Van Petten Vasconcelos Azevedo ◽

Samuel Cota Teixeira ◽

Eloá Moura ◽

Tassia Rafaela Costa ◽

...

Keyword(s):

Endothelial Cells ◽

Phospholipase A2 ◽

Snake Venom ◽

Ex Vivo ◽

Ex Vivo Approach

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Neutrophil extracellular traps induce MCP-1 release from endothelial cells at the plaque rupture site in acute myocardial infarction

European Heart Journal ◽

10.1093/ehjci/ehaa946.3834 ◽

2020 ◽

Vol 41 (Supplement_2) ◽

Author(s):

A Ondracek ◽

T.M Hofbauer ◽

A Mangold ◽

T Scherz ◽

V Seidl ◽

...

Keyword(s):

Myocardial Infarction ◽

Acute Myocardial Infarction ◽

Endothelial Cells ◽

Plaque Rupture ◽

Ex Vivo ◽

Coronary Intervention ◽

Neutrophil Extracellular Traps ◽

Funding Source ◽

Extracellular Traps

Abstract Introduction Leukocyte-mediated inflammation is crucial in acute myocardial infarction (AMI). We recently observed that neutrophil extracellular traps (NETs) are increased at the culprit site, promoting activation and differentiation of fibrocytes, cells with mesenchymal and leukocytic properties. Fibrocyte migration is mediated by monocyte chemoattractant protein (MCP)-1 and C-C chemokine receptor type 2 (CCR2). We investigated the interplay between NETs, fibrocyte function, and MCP-1 in AMI. Methods Culprit site and femoral blood of AMI patients was drawn during percutaneous coronary intervention. We characterized CCR2 expression of fibrocytes by flow cytometry. MCP-1 and the NET marker citrullinated histone H3 (citH3) were measured by ELISA. Fibrocytes were treated in vitro with MCP-1. Human coronary arterial endothelial cells (hCAECs) were stimulated with isolated NETs, and MCP-1 was measured by ELISA and qPCR. The influence of MCP-1 on NET formation in vitro was assessed using isolated neutrophils. Results We have included 50 consecutive AMI patients into the study. NETs and concentrations of MCP-1 were increased at the CLS. NET stimulation of hCAECs induced MCP-1 on mRNA and protein level. Increasing MCP-1 gradient was associated with fibrocyte accumulation at the site of occlusion. In the presence of higher MCP-1 these fibrocytes expressed proportionally less CCR2 than peripheral fibrocytes. In vitro, MCP-1 dose-dependently decreased fibrocyte CCR2 and reduced ex vivo NET release of healthy donor neutrophils. Conclusions NETs induce endothelial MCP-1 release, presumably promoting a chemotactic gradient for leukocyte and fibrocyte migration. MCP-1 mediated inhibition of NET formation could point to a negative feedback loop. These data will shed light on vascular healing. Funding Acknowledgement Type of funding source: Public grant(s) – National budget only. Main funding source(s): Austrian Science Fund

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Interaction of Neutrophils with Endothelial Cells, Fibroblasts and Their Extracellular Matrices: Microscopic and Computerised Analysis

Alternatives to Laboratory Animals ◽

10.1177/026119298801600111 ◽

1988 ◽

Vol 16 (1) ◽

pp. 48-53

Author(s):

Marina Ziche ◽

Lucia Morbidelli ◽

Annalisa Rubino ◽

Piero Dolara ◽

Stefano Bianchi ◽

...

Keyword(s):

Endothelial Cells ◽

Image Analysis ◽

Vascular Endothelial Cells ◽

Extracellular Matrices ◽

Polymorphonuclear Neutrophil ◽

Initial Event ◽

Computerised Image Analysis ◽

Capillary Endothelial Cells ◽

Vitro Model

Polymorphonuclear neutrophil (PMN) interaction with vascular endothelial cells is the initial event in the migration of neutrophils through blood vessel walls before reaching inflammation sites in tissues. The interaction between fibroblasts and endothelial cells and their extracellular matrices might be modulated by the activation of neutrophils that occurs at inflammatory reaction sites. We have used an in vitro model to study PMN function, measuring the adhesion of human PMNs to capillary endothelial cells and fibroblasts grown in culture and to their extracellular matrices. The interaction was measured in basal conditions and in the presence of the chemotactic effector, formyl-methionyl-leucyl-phenylalanine (FMLP at the concentration of 10 7M). Adhesion was expressed by the number of adherent PMNs/mm2 on a histological specimen. Moreover, we have adapted a program for image analysis to quantify neutrophil adhesion. Three times more PMNs adhered to matrices than to monolayers, and adherence could be increased by the presence of 10-7M FMLP, except in the case of fibroblast monolayers. We found a good correlation between microscopic observation and computerised image analysis measuring PMN adhesiveness to extracellular matrices.

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Binding of low molecular weight heparin (Tinzaparin sodium) to bovine endothelial cells in vitro

Thrombosis Research ◽

10.1016/0049-3848(94)90067-1 ◽

1994 ◽

Vol 75 (2) ◽

pp. 185-194 ◽

Cited By ~ 2

Author(s):

Anni Larnkjær ◽

Per B. Østergaard ◽

Hans J. Flodgaard

Keyword(s):

Molecular Weight ◽

Endothelial Cells ◽

Low Molecular Weight Heparin ◽

Low Molecular Weight ◽

Tinzaparin Sodium ◽

Bovine Endothelial Cells

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Can Be miR-126-3p a Biomarker of Premature Aging? An Ex Vivo and In Vitro Study in Fabry Disease

Cells ◽

10.3390/cells10020356 ◽

2021 ◽

Vol 10 (2) ◽

pp. 356 ◽

Cited By ~ 1

Author(s):

Alessia Lo Curto ◽

Simona Taverna ◽

Maria Assunta Costa ◽

Rosa Passantino ◽

Giuseppa Augello ◽

...

Keyword(s):

Endothelial Cells ◽

Fabry Disease ◽

Healthy Subjects ◽

Aging Process ◽

Replicative Senescence ◽

Ex Vivo ◽

Umbilical Vein ◽

Quantitative Polymerase Chain Reaction ◽

Premature Aging

Fabry disease (FD) is a lysosomal storage disorder (LSD) characterized by lysosomal accumulation of glycosphingolipids in a wide variety of cytotypes, including endothelial cells (ECs). FD patients experience a significantly reduced life expectancy compared to the general population; therefore, the association with a premature aging process would be plausible. To assess this hypothesis, miR-126-3p, a senescence-associated microRNA (SA-miRNAs), was considered as an aging biomarker. The levels of miR-126-3p contained in small extracellular vesicles (sEVs), with about 130 nm of diameter, were measured in FD patients and healthy subjects divided into age classes, in vitro, in human umbilical vein endothelial cells (HUVECs) “young” and undergoing replicative senescence, through a quantitative polymerase chain reaction (qPCR) approach. We confirmed that, in vivo, circulating miR-126 levels physiologically increase with age. In vitro, miR-126 augments in HUVECs underwent replicative senescence. We observed that FD patients are characterized by higher miR-126-3p levels in sEVs, compared to age-matched healthy subjects. We also explored, in vitro, the effect on ECs of glycosphingolipids that are typically accumulated in FD patients. We observed that FD storage substances induced in HUVECs premature senescence and increased of miR-126-3p levels. This study reinforces the hypothesis that FD may aggravate the normal aging process.

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A tumor-promoting role for soluble TβRIII in glioblastoma

Molecular and Cellular Biochemistry ◽

10.1007/s11010-021-04128-y ◽

2021 ◽

Author(s):

Isabel Burghardt ◽

Judith Johanna Schroeder ◽

Tobias Weiss ◽

Dorothee Gramatzki ◽

Michael Weller

Keyword(s):

Endothelial Cells ◽

Ex Vivo ◽

Microvascular Endothelial Cells ◽

Smad2 Phosphorylation ◽

Cell Gene Expression ◽

Microvascular Endothelial ◽

Cell Gene ◽

Mouse Glioma

Abstract Purpose Members of the transforming growth factor (TGF)-β superfamily play a key role in the regulation of the malignant phenotype of glioblastoma by promoting invasiveness, angiogenesis, immunosuppression, and maintaining stem cell-like properties. Betaglycan, a TGF-β coreceptor also known as TGF-β receptor III (TβRIII), interacts with members of the TGF-β superfamily and acts as membrane-associated or shed molecule. Shed, soluble TβRIII (sTβRIII) is produced upon ectodomain cleavage of the membrane-bound form. Elucidating the role of TβRIII may improve our understanding of TGF-β pathway activity in glioblastoma Methods Protein levels of TβRIII were determined by immunohistochemical analyses and ex vivo single-cell gene expression profiling of glioblastoma tissue respectively. In vitro, TβRIII levels were assessed investigating long-term glioma cell lines (LTCs), cultured human brain-derived microvascular endothelial cells (hCMECs), glioblastoma-derived microvascular endothelial cells, and glioma-initiating cell lines (GICs). The impact of TβRIII on TGF-β signaling was investigated, and results were validated in a xenograft mouse glioma model Results Immunohistochemistry and ex vivo single-cell gene expression profiling of glioblastoma tissue showed that TβRIII was expressed in the tumor tissue, predominantly in the vascular compartment. We confirmed this pattern of TβRIII expression in vitro. Specifically, we detected sTβRIII in glioblastoma-derived microvascular endothelial cells. STβRIII facilitated TGF-β-induced Smad2 phosphorylation in vitro and overexpression of sTβRIII in a xenograft mouse glioma model led to increased levels of Smad2 phosphorylation, increased tumor volume, and decreased survival Conclusions These data shed light on the potential tumor-promoting role of extracellular shed TβRIII which may be released by glioblastoma endothelium with high sTβRIII levels.

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