scholarly journals Vasoactive amines directly modify endothelial cells to affect polymorphonuclear leukocyte diapedesis in vitro

Blood ◽  
1987 ◽  
Vol 69 (6) ◽  
pp. 1563-1569
Author(s):  
J Doukas ◽  
D Shepro ◽  
HB Hechtman
Keyword(s):  
Endothelial Cells ◽  
Basement Membrane ◽  
Polymorphonuclear Leukocyte ◽  
Cytochalasin B ◽  
Membrane Surface ◽  
Stress Fiber ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells

Bovine aortic endothelial cells were cultured on the basement membrane surface of amnionic membrane and used as a substrate for polymorphonuclear leukocyte (PMN) diapedesis in vitro. Norepinephrine (NE), serotonin (5HT), or phalloidin treatment of the endothelial cells (ECs) reduces, whereas histamine or cytochalasin B increases, the number of PMNs migrating across the ECs and amnionic membrane. In contrast, amine treatment of PMNs or acellular amnionic membrane does not alter PMN diapedesis or chemotaxis. The NE and histamine effects are blocked by appropriate receptor antagonists, but the 5HT effect is not. All the agents' effects are also reversible. Qualitatively similar effects on EC permeability to Evan's blue-labeled albumin occur with all agents; however, PMN adhesion to ECs is not affected. Previously, we reported that NE and 5HT increase stress fiber numbers and decrease EC permeability to macromolecules in vitro, whereas histamine has the opposite effects, and that NE and 5HT eliminate the erythrocyte extravasation associated with thrombocytopenia in vivo. In this study, we propose that these vasoactive amines also alter PMN diapedesis in vitro through a direct effect on the EC, in part due to alterations in the EC cytoskeleton.

scholarly journals Vasoactive amines directly modify endothelial cells to affect polymorphonuclear leukocyte diapedesis in vitro

Blood ◽  
1987 ◽  
Vol 69 (6) ◽  
pp. 1563-1569 ◽  
Author(s):  
J Doukas ◽  
D Shepro ◽  
HB Hechtman
Keyword(s):  
Endothelial Cells ◽  
Basement Membrane ◽  
Polymorphonuclear Leukocyte ◽  
Cytochalasin B ◽  
Membrane Surface ◽  
Stress Fiber ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells

Abstract Bovine aortic endothelial cells were cultured on the basement membrane surface of amnionic membrane and used as a substrate for polymorphonuclear leukocyte (PMN) diapedesis in vitro. Norepinephrine (NE), serotonin (5HT), or phalloidin treatment of the endothelial cells (ECs) reduces, whereas histamine or cytochalasin B increases, the number of PMNs migrating across the ECs and amnionic membrane. In contrast, amine treatment of PMNs or acellular amnionic membrane does not alter PMN diapedesis or chemotaxis. The NE and histamine effects are blocked by appropriate receptor antagonists, but the 5HT effect is not. All the agents' effects are also reversible. Qualitatively similar effects on EC permeability to Evan's blue-labeled albumin occur with all agents; however, PMN adhesion to ECs is not affected. Previously, we reported that NE and 5HT increase stress fiber numbers and decrease EC permeability to macromolecules in vitro, whereas histamine has the opposite effects, and that NE and 5HT eliminate the erythrocyte extravasation associated with thrombocytopenia in vivo. In this study, we propose that these vasoactive amines also alter PMN diapedesis in vitro through a direct effect on the EC, in part due to alterations in the EC cytoskeleton.

scholarly journals Capillary endothelial cell cultures: phenotypic modulation by matrix components.

1983 ◽  
Vol 97 (1) ◽  
pp. 153-165 ◽  
Author(s):  
J A Madri ◽  
S K Williams
Keyword(s):  
Endothelial Cells ◽  
Basement Membrane ◽  
Membrane Surface ◽  
Phenotypic Expression ◽  
Aortic Endothelial Cells ◽  
Capillary Endothelial Cells ◽  
Endothelial Cell Cultures ◽  
Cell Densities

Capillary endothelial cells of rat epididymal fat pad were isolated and cultured in media conditioned by bovine aortic endothelial cells and substrata consisting of interstitial or basement membrane collagens. When these cells were grown on interstitial collagens they underwent proliferation, formed a continuous cell layer and, if cultured for long periods of time, formed occasional tubelike structures. In contrast, when these cells were grown on basement membrane collagens, they did not proliferate but did aggregate and form tubelike structures at early culture times. In addition, cells grown on basement membrane substrata expressed more basement membrane constituents as compared with cells grown on interstitial matrices when assayed by immunoperoxidase methods and quantitated by enzyme-linked immunosorbent inhibition assays. Furthermore, when cells were grown on either side of washed, acellular amnionic membranes their phenotypes were markedly different. On the basement membrane surface they adhered, spread, and formed tubelike structures but did not migrate through the basement membrane. In contrast, when seeded on the stromal surface, these cells were observed to proliferate and migrate into the stromal aspect of the amnion and ultimately formed tubelike structures at high cell densities at longer culture periods (21 d). Thus, connective tissue components play important roles in regulating the phenotypic expression of capillary endothelial cells in vitro, and similar roles of the collagenous components of the extracellular matrix may exist in vivo following injury and during angiogenesis. Furthermore, the culture systems outlined here may be of use in the further study of differentiated, organized capillary endothelial cells in culture.

In vivo and in vitro evidence for ACh-stimulated l-arginine uptake

2004 ◽  
Vol 287 (1) ◽  
pp. H395-H400 ◽  
Author(s):  
Melinda M. Parnell ◽  
Jaye P. F. Chin-Dusting ◽  
Jennifer Starr ◽  
David M. Kaye
Keyword(s):  
Endothelial Cells ◽  
Arterial Infusion ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Arginine Uptake ◽  
Human Forearm ◽  
Few Data ◽  
Forearm Circulation

Whereas l-arginine is clearly recognized as the precursor for nitric oxide synthesis, and its entry into endothelial cells via system y+ transport is established, few data exist regarding the acute regulation of this transport process. We specifically investigated the effect of ACh and isoprenaline (Iso) on l-arginine uptake in the human forearm and in cultured bovine aortic endothelial cells (BAEC). Sixteen healthy males were studied. During a steady-state intra-arterial infusion of [3H]l-arginine (100 nCi/min), the effects of ACh (9.25 and 37 μg/min), Iso (25–50 and 200 μg/min), and sodium nitroprusside (SNP) (1–2 and 8 μg/min) on forearm plasma flow (FPF), l-[3H]arginine uptake, and l-[3H]citrulline release were determined. In parallel experiments, the effects of ACh, Iso, and SNP on l-[3H]arginine uptake were studied in BAEC. l-Arginine uptake was inversely related to FPF ( r = −0.50; P < 0.005). At a similar FPF (ACh 56.82 ± 9.25, Iso 58.49 ± 5.56, SNP 57.92 ± 4.96 ml/min; P = ns), intra-arterial ACh significantly increased forearm uptake of l-[3H]arginine (54,655 ± 8,018 dpm/min), compared with that observed with either Iso (40,517.23 ± 6,841 dpm/min; P = 0.01) or SNP (36,816 ± 4,650 dpm/min; P = 0.011). This was associated with increased ACh-induced l-[3H]citrulline release compared with Iso and SNP ( P = 0.046). Similarly, in BAEC, ACh significantly increased l-[3H]arginine uptake compared with control, Iso, or SNP (ACh 12.0 × 107 ± 1.83 × 107 vs. control 6.67 × 107 ± 1.16 × 107 vs. Iso 7.35 × 107 ± 1.63 × 107 vs. SNP 6.01 × 107 ± 1.11 × 107 fmol·min−1·mg−1 at 300 μmol/l l-arginine; P = 0.043). Taken together, these data indicate that ACh stimulates l-arginine uptake in cultured endothelial cells and in human forearm circulation, indicating the potential for acute modulation of endothelial l-arginine uptake.

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

1983 ◽  
Vol 49 (02) ◽  
pp. 132-137 ◽  
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.

scholarly journals Urea transporters are distributed in endothelial cells and mediate inhibition of l-arginine transport

2002 ◽  
Vol 283 (3) ◽  
pp. F578-F582 ◽  
Author(s):  
Laszlo Wagner ◽  
Janet D. Klein ◽  
Jeff M. Sands ◽  
Chris Baylis
Keyword(s):  
Endothelial Cells ◽  
Cell Types ◽  
Inhibitory Effect ◽  
Rat Aorta ◽  
Wide Distribution ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
Elevated Blood Urea Nitrogen ◽  
Aortic Endothelial

Our laboratory previously reported that uremic levels of urea inhibitl-arginine (l-Arg) transport into endothelial cells. The present study further investigated this effect. We measuredl-Arg transport in cultured bovine aortic endothelial cells with normal or high urea (25 mM). The urea transport inhibitor phloretin abolished the inhibitory effect of urea on l-Arg transport, suggesting a role for urea transporters (UTs). We screened bovine aortic endothelial cells and several other endothelial cell types for the presence of UTs by using Western blot analysis. UT-B was present in all endothelial cells, irrespective of species or location of derivation, whereas UT-A distribution was variable and sparse. UT-B was also abundant in rat aorta, mesenteric blood vessels, and spinotrapezius muscle, whereas UT-A distribution was, again, variable and sparse. Chronic elevation of urea had variable, inconsistent effects on UT abundance. This study showed that urea must enter endothelial cells, probably by UT-B, to inhibit l-Arg transport. In view of the wide distribution of UT-B in rat vasculature, elevated blood urea nitrogen may lead to endothelial l-Arg deficiency in vivo.

scholarly journals Perturbations in the fibrinolytic pathway abolish cyst formation but not capillary-like organization of cultured murine endothelial cells

Blood ◽  
1994 ◽  
Vol 83 (11) ◽  
pp. 3206-3217 ◽  
Author(s):  
N Dubois-Stringfellow ◽  
A Jonczyk ◽  
VL Bautch
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Basement Membrane ◽  
Organizational Behavior ◽  
Cell Lines ◽  
Primary Cultures ◽  
Cyst Formation ◽  
Fibrin Gels

Abstract Fibrinolytic activity and its relation to morphogenesis was investigated in several transformed murine endothelial cell lines and primary cultures of endothelial cells. Two in vitro systems, fibrin gels and Matrigel (Collaborative Research, Bedford, MA), were used. Fibrin gels model a fibrin-rich extracellular matrix that frequently supports neovascularization in vivo, and Matrigel models the basement membrane surrounding quiescent endothelial cells in vivo. The transformed endothelial cell lines have higher levels of plasminogen activator (PA) mRNA than primary cultures of endothelial cells, and an increased PA-mediated proteolytic activity was correlated with formation of cysts in fibrin gels. Addition of neutralizing anti- urokinase antibodies, plasminogen depletion, or addition of a plasmin inhibitor prevented cyst formation. Addition of plasminogen restored the ability to form cysts in the plasminogen-depleted system. Normal endothelial cells organized into capillary-like structures in fibrin gels regardless of manipulations affecting the fibrinolytic pathway. In Matrigel, both transformed and primary cultures of endothelial cells rapidly formed a capillary-like network that was not affected by plasminogen depletion or addition of plasmin inhibitors. Thus, elements of the fibrinolytic pathway necessary for cyst formation are not critical in capillary-like structure formation on a reconstituted basement membrane. These results suggest that plasmin is essential for hemangioma formation but is not critical to the organizational behavior of normal endothelial cells.

Endopeptidases 3.4.24.15 and 24.16 in endothelial cells: potential role in vasoactive peptide metabolism

2003 ◽  
Vol 284 (6) ◽  
pp. H1978-H1984 ◽  
Author(s):  
M. Ursula Norman ◽  
Shane B. Reeve ◽  
Vincent Dive ◽  
A. Ian Smith ◽  
Rebecca A. Lew
Keyword(s):  
Endothelial Cells ◽  
High Performance ◽  
Vascular Endothelial Cells ◽  
Umbilical Vein ◽  
Hybrid Cell ◽  
Fluorescent Substrate ◽  
Intact Cells ◽  
Aortic Endothelial Cells

The closely related metalloendopeptidases EC 3.4.24.15 (EP24.15; thimet oligopeptidase) and 24.16 (EP24.16; neurolysin) cleave a number of vasoactive peptides such as bradykinin and neurotensin in vitro. We have previously shown that hypotensive responses to bradykinin are potentiated by an inhibitor of EP24.15 and EP24.16 (26), suggesting a role for one or both enzymes in bradykinin metabolism in vivo. In this study, we have used selective inhibitors that can distinguish between EP24.15 and EP24.16 to determine their activity in cultured endothelial cells (the transformed human umbilical vein endothelial hybrid cell line EA.hy926 or ovine aortic endothelial cells). Endopeptidase activity was assessed using a specific quenched fluorescent substrate [7-methoxycoumarin-4-acetyl-Pro-Leu-Gly-d-Lys(2,4-dinitrophenyl)], as well as the peptide substrates bradykinin and neurotensin (assessed by high-performance liquid chromatography with mass spectroscopic detection). Our results indicate that both peptidases are present in endothelial cells; however, EP24.16 contributes significantly more to substrate cleavage by both cytosolic and membrane preparations, as well as intact cells, than EP24.15. These findings, when coupled with previous observations in vivo, suggest that EP24.16 activity in vascular endothelial cells may play an important role in the degradation of bradykinin and/or other peptides in the circulation.

scholarly journals The tight junctions protein Claudin-5 limits endothelial cell motility

2020 ◽  
pp. jcs.248237
Author(s):  
Zhenguo Yang ◽  
Shuilong Wu ◽  
Federica Fontana ◽  
Yanyu Li ◽  
Wei Xiao ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Endothelial Cell ◽  
Cell Motility ◽  
Dorsal Aorta ◽  
Aortic Endothelial Cells ◽  
Differential Adhesion ◽  
Adhesive Forces

Steinberg's differential adhesion hypothesis suggests that adhesive mechanisms are important for sorting of cells and tissues during morphogenesis (Steinberg, 2007). During zebrafish vasculogenesis, endothelial cells sort into arterial and venous vessel beds but it is unknown whether this involves adhesive mechanisms. Claudins are tight junction proteins regulating the permeability of epithelial and endothelial tissue barriers. Previously, the roles of Claudins during organ development have exclusively been related to their canonical functions in determining paracellular permeability. Here, we use atomic force microscopy to quantify Claudin-5-dependent adhesion and find that this strongly contributes to the adhesive forces between arterial endothelial cells. Based on genetic manipulations, we reveal a non-canonical role of Claudin-5a during zebrafish vasculogenesis, which involves the regulation of adhesive forces between adjacent dorsal aortic endothelial cells. In vitro and in vivo studies demonstrate that loss of Claudin-5 results in increased motility of dorsal aorta endothelial cells and in a failure of the dorsal aorta to lumenize. Our findings uncover a novel role of Claudin-5 in limiting arterial endothelial cell motility, which goes beyond its traditional sealing function during embryonic development.

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