A simple multi-well stretching device to induce inflammatory responses of vascular endothelial cells

Objective TGR5, a membrane-bound, G-protein-coupled receptor for bile acids, is known to be involved in regulation of energy homeostasis and inflammation. However, little is known about the function of TGR5 in vascular endothelial cells. In the present study, we examined whether TGR5 agonism represents anti-inflammatory effects in vascular endothelial cells focusing on nitric oxide (NO) production. Methods and Results In human umbilical vein endothelial cells (HUVECs), treatment with taurolithocholic acid (TLCA), which has the highest affinity to TGR5 among various bile acids, significantly reduced tumor necrosis factor (TNF)-α-induced vascular cell adhesion molecule (VCAM)-1 protein expression and adhesion of human monocytes, U937. These effects were abrogated by a NO synthase (NOS) inhibitor, N G -Monomethyl-L-arginine (L-NMMA). In bovine aortic endothelial cells (BAECs), treatment with TLCA as well as lithocholic acid, which also has high affinity to TGR5, significantly increased the NO production. In contrast, deoxycholic acid and chenodeoxycholic acid, which possess low affinity to TGR5, did not affect the NO production. Gene depletion of TGR5 by siRNA transfection abolished TLCA-induced NO production in BAECs. TLCA-induced NO production was also observed in HUVECs measured as intracellular cGMP accumulation. We next investigated the signal pathways responsible for the TLCA-induced NO production in endothelial cells. Treatment with TLCA increased endothelial NOS (eNOS) ser1177 phosphorylation in HUVECs. This response was accompanied by increased Akt ser473 phosphorylation and intracellular Ca 2+ ([Ca 2+ ] i ). Treatment with phosphoinositide 3-kinase (PI3K) inhibitor, LY294002, or blockade of calcium channel with La 3+ , significantly decreased TLCA-induced eNOS ser1177 phosphorylation and subsequent NO production. Conclusion These results indicate that TGR5 agonism can mediate anti-inflammatory responses by suppressing VCAM-1 expression and monocytes adhesion to endothelial cells. This function is dependent on NO production via Akt activation and [Ca 2+ ] i increase.

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Interaction between vascular endothelial cells and vascular intimal spindle-shaped cells in vitro

Journal of Cell Science ◽

10.1242/jcs.60.1.89 ◽

1983 ◽

Vol 60 (1) ◽

pp. 89-102

Author(s):

D de Bono ◽

C. Green

Keyword(s):

Extracellular Matrix ◽

Endothelial Cells ◽

Vascular Endothelial Cells ◽

Three Dimensional ◽

Vascular Endothelial ◽

Pure Cultures ◽

Species Specific ◽

Endothelial Growth Factor

The interactions between human or bovine vascular endothelial cells and fibroblast-like vascular intimal spindle-shaped cells have been studied in vitro, using species-specific antibodies to identify the different components in mixed cultures. Pure cultures of endothelial cells grow as uniform, nonoverlapping monolayers, but this growth pattern is lost after the addition of spindle cells, probably because the extracellular matrix secreted by the latter causes the endothelial cells to modify the way they are attached to the substrate. The result is a network of tubular aggregates of endothelial cells in a three-dimensional ‘polylayer’ of spindle-shaped cells. On the other hand, endothelial cells added to growth-inhibited cultures of spindle-shaped cells will grow in sheets over the surface of the culture. Human endothelial cells grown in contact with spindle-shaped cells have a reduced requirement for a brain-derived endothelial growth factor. The interactions of endothelial cells and other connective tissue cells in vitro may be relevant to the mechanisms of endothelial growth and blood vessel formation in vivo, and emphasize the potential importance of extracellular matrix in controlling endothelial cell behaviour.

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Induction of Apoptotic Cell Death in Vascular Endothelial Cells Cultured in Three-Dimensional Collagen Lattice

Experimental Cell Research ◽

10.1006/excr.1999.4422 ◽

1999 ◽

Vol 248 (2) ◽

pp. 498-508 ◽

Cited By ~ 41

Author(s):

Masafumi Kuzuya ◽

Shosuke Satake ◽

Miguel A. Ramos ◽

Shigeru Kanda ◽

Teruhiko Koike ◽

...

Keyword(s):

Endothelial Cells ◽

Cell Death ◽

Vascular Endothelial Cells ◽

Apoptotic Cell ◽

Three Dimensional ◽

Apoptotic Cell Death ◽

Vascular Endothelial ◽

Collagen Lattice ◽

Cells Cultured

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Novel control of cAMP-regulated transcription in vascular endothelial cells

Biochemical Society Transactions ◽

10.1042/bst20110606 ◽

2012 ◽

Vol 40 (1) ◽

pp. 1-5 ◽

Cited By ~ 9

Author(s):

Gillian R. Milne ◽

Timothy M. Palmer ◽

Stephen J. Yarwood

Keyword(s):

Endothelial Cells ◽

Molecular Mechanisms ◽

Inflammatory Responses ◽

Vascular Endothelial Cells ◽

Inflammatory Diseases ◽

Circulatory System ◽

Vascular Endothelial ◽

Protective Mechanisms ◽

Enhancer Binding Protein ◽

Developed World

Chronic inflammatory diseases, such as atherosclerosis, are a major cause of death and disability in the developed world. In this respect, although cholesterol obviously plays a predominant role in atherosclerosis, targeting inflammation at lesion sites may be just as important. Indeed, elevated IL-6 (interleukin 6) levels are as strongly associated with coronary heart disease as increased cholesterol. We have been investigating novel cAMP-regulated pathways that combat the action of pro-inflammatory cytokines, such as IL-6 and leptin, in the VECs (vascular endothelial cells) of the circulatory system. In this respect, we have begun to unravel new molecular mechanisms by which the cAMP/Epac1 (exchange protein directly activated by cAMP 1)/Rap1 pathway can initiate a rigorous programme of protective anti-inflammatory responses in VECs. Central to this is the coupling of cAMP elevation to the mobilization of two C/EBP (CCAAT/enhancer-binding protein) family transcription factors, resulting in the induction of the SOCS3 (suppressor of cytokine signalling 3) gene, which attenuates pro-inflammatory cytokine signalling in VECs. These novel ‘protective’ mechanisms of cAMP action will inform the development of the next generation of pharmaceuticals specifically designed to combat endothelial inflammation associated with cardiovascular disease.

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Inorganic polyphosphate elicits pro-inflammatory responses through activation of the mammalian target of rapamycin complexes 1 and 2 in vascular endothelial cells

Journal of Thrombosis and Haemostasis ◽

10.1111/jth.12899 ◽

2015 ◽

Vol 13 (5) ◽

pp. 860-871 ◽

Cited By ~ 35

Author(s):

S. M. Hassanian ◽

P. Dinarvand ◽

S. A. Smith ◽

A. R. Rezaie

Keyword(s):

Endothelial Cells ◽

Inflammatory Responses ◽

Vascular Endothelial Cells ◽

Mammalian Target Of Rapamycin ◽

Target Of Rapamycin ◽

Vascular Endothelial ◽

Inorganic Polyphosphate

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Mediators released from LPS-challenged lungs induce inflammatory responses in liver vascular endothelial cells and neutrophilic leukocytes

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00278.2009 ◽

2009 ◽

Vol 297 (6) ◽

pp. G1066-G1076 ◽

Cited By ~ 20

Author(s):

N. Markovic ◽

L. A. McCaig ◽

J. Stephen ◽

S. Mizuguchi ◽

R. A. W. Veldhuizen ◽

...

Keyword(s):

Endothelial Cells ◽

Inflammatory Response ◽

Inflammatory Responses ◽

Vascular Endothelial Cells ◽

Vascular Endothelial ◽

Ros Production ◽

Proinflammatory Mediators ◽

Primary Mouse ◽

Mouse Lungs

The systemic inflammatory response plays an important role in the progression of acute lung injury (ALI) to multiple organ dysfunction syndrome (MODS). However, the role of lung-derived inflammatory mediators in induction of the inflammatory response in remote organs is poorly understood. To address the above, we investigated the effects of lung inflammation on induction of inflammatory response(s) in the liver in vitro. Inflammation in mouse lungs was induced by intranasal administration of lipopolysaccharide (LPS; 1 mg/ml) followed by mechanical ventilation using the isolated perfused mouse lung method to obtain and characterize lung perfusate from the pulmonary circulation. LPS administration to mouse lungs resulted in an increased release of inflammation-relevant cytokines and chemokines into the perfusate (Luminex assay) compared with the saline-controls. Subsequently, primary mouse liver vascular endothelial cells (LVEC) or mouse polymorphonuclear leukocytes (PMN) in vitro were stimulated with the perfusate obtained from saline- or LPS-challenged lungs and assessed for various inflammation-relevant end points. The obtained results indicate that stimulation of LVEC with perfusate obtained from LPS-challenged lungs results in 1) reactive oxygen species (ROS) production; 2) activation of NF-κB; and 3) expression of E-selectin, ICAM-1, and VCAM-1 and a subsequent increase in PMN rolling and adhesion to LVEC. In addition, perfusate from LPS-challenged lung induced activation of PMN with respect to increased ROS production and upregulation of cell surface levels of adhesion molecules MAC-1 and VLA-4. Heat-inactivation of the perfusate obtained from LPS-challenged lungs was very effective in suppressing increased proadhesive phenotype (i.e., E-selectin and ICAM-1 expression) in LVEC, whereas targeted inhibition (immunoneutralization) of TNF-α and/or IL-6 in LPS-lung perfusate had no effect. Taken together, these findings indicate that multiple proinflammatory mediators (proteinaceous in nature) released from inflamed lungs act synergistically to induce systemic activation of circulating PMN and promote inflammatory responses in liver vascular endothelial cells.

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