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.

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 Endothelin stimulates phosphatidylinositol hydrolysis and DNA synthesis in brain capillary endothelial cells

1990 ◽  
Vol 266 (2) ◽  
pp. 415-420 ◽  
Author(s):  
P Vigne ◽  
R Marsault ◽  
J P Breittmayer ◽  
C Frelin
Keyword(s):  
Endothelial Cells ◽  
Dna Synthesis ◽  
Cardiac Cells ◽  
Brain Capillary ◽  
Aortic Endothelial Cells ◽  
Brain Capillary Endothelial Cells ◽  
Important Target ◽  
Receptor Sites ◽  
Capillary Endothelial Cells

Endothelin-1 (ET-1) is a novel vasoconstricting and cardiotonic peptide that is synthesized by the vascular endothelium. Bovine aortic endothelial cells which secrete ET in vitro lack membrane receptor sites for the peptide. Endothelial cells from rat brain microvessels that do not secrete ET in vitro express large amounts of high-affinity receptors for 125I-labelled ET-1 (Kd 0.8 nM). The ET receptor is recognized by sarafotoxin S6b and the different ET peptides with the following order of potency: ET-1 (Kd 0.5 nM) approximately equal to ET-2 (Kd 0.7 nM) greater than sarafotoxin S6b (Kd 27 nM) greater than ET-3 (Kd 450 nM). This structure-activity relationship is different from those found in vascular smooth muscle cells, renal cells and cardiac cells. ET-1 stimulates DNA synthesis in brain capillary endothelial cells. It is more potent than basic fibroblast growth factor. The action of ET on endothelial cells from microvessels involves phosphatidylinositol hydrolysis and intracellular Ca2+ mobilization. These observations suggest that brain endothelial cells might be an important target for ET.

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.

The Effects of Shear Stress on Endothelial Cells at Hypothermic Temperatures

2002 ◽  
Author(s):  
John H. Slater ◽  
Shailendra Jain ◽  
Robin N. Coger ◽  
Charles Y. Lee
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
Endothelial Cell ◽  
Low Flow ◽  
Shear Stresses ◽  
Liver Sinusoids ◽  
Endothelial Cell Cultures ◽  
Shear Stress Response

Hypothermic machine perfusion preservation (MPP) has proven to be a successful technique for hypothermic kidney storage, however this technology has not successfully been applied to the liver. Recent research has indicated that the endothelial cells lining the liver sinusoids display rounding phenomena during MPP that is not fully understood. In order to gain a better understanding of endothelial cell shear stress response and the factors that induce rounding, a temperature-controlled micro-shear chamber has been designed and fabricated. The micro-shear chamber has been used to apply shear stresses, corresponding to those imposed during MPP, to rat liver primary endothelial cell cultures in order to form an understanding of how these stresses affect endothelial cell morphology. The chamber allows for the application of shear stresses ranging from 0.2 ± .01 dynes/cm2 to 2.3 ± 0.3 dynes/cm2, corresponding to what occurs during MPP.] Twenty-four hour in vitro experiments with shear stresses ranging from 0 to 1.49 dynes/cm2 at 4 °C were conducted in order to replicate in vivo conditions of the liver during hypothermic MPP. It has been demonstrated that endothelial cell rounding increases with increasing shear and can be prevented by utilizing low flow rates.

Angiogenic oligosaccharides of hyaluronan enhance the production of collagens by endothelial cells

1993 ◽  
Vol 105 (1) ◽  
pp. 213-218
Author(s):  
P. Rooney ◽  
M. Wang ◽  
P. Kumar ◽  
S. Kumar
Keyword(s):  
Endothelial Cells ◽  
Polyacrylamide Gel Electrophoresis ◽  
Chorioallantoic Membrane ◽  
Type I ◽  
Aortic Endothelial Cells ◽  
Type Viii ◽  
High Degree ◽  
Bovine Type

The present study demonstrates a relationship between angiogenic oligosaccharides of hyaluronan (HA) and the production of collagens during the process of angiogenesis in vivo and in vitro. The addition of angiogenic oligosaccharides of HA to the chorioallantoic membrane of the chick embryo induced a deposition of collagen fibrils. The treatment of sub-confluent cultures of bovine aortic endothelial cells with the same oligosaccharides (1 microgram/ml) increased the uptake of [3H]proline by approximately 60%. SDS-polyacrylamide gel electrophoresis of treated cultures demonstrated the enhanced synthesis of type I and type VIII collagens. The production of type VIII collagen was confirmed by western blotting and immunocytochemistry using antibodies to sheep and bovine type VIII collagen. Type VIII collagen is a short chain collagen that has a high degree of homology to cartilage-specific type X collagen. The biological functions of type VIII and type X collagens are unknown. We have suggested that the two collagens play a role in the process of angiogenesis.

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