Differential effects of NF-κB and p38 MAPK inhibitors and combinations thereof on TNF-α- and IL-1β-induced proinflammatory status of endothelial cells in vitro

2005 ◽  
Vol 289 (5) ◽  
pp. C1229-C1239 ◽  
Author(s):  
Joanna M. Kułdo ◽  
Johanna Westra ◽  
Sigridur A. Ásgeirsdóttir ◽  
Robbert J. Kok ◽  
Koen Oosterhuis ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
P38 Mapk ◽  
Umbilical Vein ◽  
Combined Treatment ◽  
Dominant Negative ◽  
Mrna Levels ◽  
Induced Expression ◽  
Inflammatory Genes ◽  
Tnf Α ◽  
Cox 2

Endothelial cells actively participate in inflammatory events by regulating leukocyte recruitment via the expression of inflammatory genes such as E-selectin, VCAM-1, ICAM-1, IL-6, IL-8, and cyclooxygenase (COX)-2. In this study we showed by real-time RT-PCR that activation of human umbilical vein endothelial cells (HUVEC) by TNF-α and IL-1β differentially affected the expression of these inflammatory genes. Combined treatment with TNF-α and IL-1β resulted in nonadditive, additive, and even synergistic induction of expression of VCAM-1, IL-8, and IL-6, respectively. Overexpression of dominant-negative inhibitor κB protein blocking NF-κB signaling confirmed a major role of this pathway in controlling both TNF-α- and IL-1β-induced expression of most of the genes studied. Although dexamethasone exerted limited effects at 1 μM, the thioredoxin inhibitor MOL-294, which regulates the redox state of NF-κB, mainly inhibited adhesion molecule expression. Its most pronounced effect was seen on VCAM-1 mRNA levels, especially in IL-1β-activated endothelium. One micromolar RWJ-67657, an inhibitor of p38 MAPK activity, diminished TNF-α- and IL-1β-induced expression of IL-6, IL-8, and E-selectin but had little effect on VCAM-1 and ICAM-1. Combined treatment of HUVEC with MOL-294 and RWJ-67657 resulted in significant blocking of the expression of E-selectin, IL-6, IL-8, and COX-2. The inhibitory effects were much stronger than those observed with single drug treatment. Application of combinations of drugs that affect multiple targets in activated endothelial cells may therefore be considered as a potential new therapeutic strategy to inhibit inflammatory disease activity.

A proteasome inhibitor reduces concurrent, sequential, and long-term IL-1β- and TNF-α-induced ECAM expression and adhesion

2003 ◽  
Vol 285 (4) ◽  
pp. C813-C822 ◽  
Author(s):  
Nilesh M. Dagia ◽  
Douglas J. Goetz
Keyword(s):  
Endothelial Cells ◽  
Cell Adhesion ◽  
Proteasome Inhibitor ◽  
Umbilical Vein ◽  
Hl60 Cell ◽  
Induced Expression ◽  
Tnf Α ◽  
Short Period

A promising approach for reducing aberrant leukocyte-endothelial adhesion during pathological inflammation is to inhibit endothelial cell adhesion molecule (ECAM) expression at the transcription level. Several compounds have been shown to decrease cytokine-induced upregulation of ECAMs primarily by modulating the activity of transcription factors [e.g., nuclear factor-κB (NF-κB)]. The majority of the in vitro studies have focused on the effect of transcription inhibitors on endothelial cells exposed to a single cytokine [primarily tumor necrosis factor-α (TNF-α)] for a relatively short period of time (primarily 4-6 h). However, in the in vivo setting, multiple cytokines [e.g., interleukin-1β (IL-1β) and TNF-α] may be present for extended periods of time. Thus we studied the effects of a transcription inhibitor, the proteasome inhibitor lactacystin, on ECAM expression and myeloid (HL60) cell adhesion to human umbilical vein endothelial cells (HUVEC) activated by concurrent, sequential, and long-term (24 h) treatment with IL-1β and TNF-α. We show, for the first time, that lactacystin inhibits 1) 4-h concurrent IL-1β- and TNF-α-induced expression of E-selectin, VCAM-1, ICAM-1, and HL60 cell adhesion to HUVEC; 2) 4-h TNF-α-induced expression of E-selectin, VCAM-1, and HL60 cell adhesion to HUVEC that have become desensitized to IL-1β activation; 3) 24-h TNF-α-induced expression of E-selectin and VCAM-1 but not ICAM-1; and 4) 24-h TNF-α-induced HL60 cell adhesion to HUVEC. Combined, our results demonstrate that a proteasome inhibitor can reduce concurrent, sequential, and long-term IL-1β- and TNF-α-induced ECAM expression and myeloid cell adhesion.

Red wine extract protects against oxidative-stress-induced endothelial senescence

2012 ◽  
Vol 123 (8) ◽  
pp. 499-507 ◽  
Author(s):  
Ilse P. G. Botden ◽  
Hisko Oeseburg ◽  
Matej Durik ◽  
Frank P. J. Leijten ◽  
Leonie C. Van Vark-Van Der Zee ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Endothelial Cells ◽  
Red Wine ◽  
Umbilical Vein ◽  
Critical Role ◽  
Sirtuin 1 ◽  
Mrna Levels ◽  
Nitric Oxide Synthase Inhibitor ◽  
Age Related ◽  
Cox 2

Red wine polyphenols may preserve endothelial function during aging. Endothelial cell senescence enhances age-related endothelial dysfunction. We investigated whether RWE (red wine extract) prevents oxidative-stress-induced senescence in HUVECs (human umbilical-vein endothelial cells). Senescence was induced by exposing HUVECs to tBHP (t-butylhydroperoxide), and quantified by senescence-associated β-galactosidase staining. RWE (0–50 μg/ml) concentration dependently decreased senescence by maximally 33±7.1%. RWE prevented the senescence-associated increase in p21 protein expression, inhibited tBHP-induced DNA damage of endothelial cells and induced relaxation of PCAs (porcine coronary arteries). Inhibition of SIRT1 (sirtuin 1) by sirtinol partially reversed the effect of RWE on tBHP-induced senescence, whereas both the NOS (nitric oxide synthase) inhibitor L-NMMA (NG-monomethyl-L-arginine) and the COX (cyclo-oxygenase) inhibitor indomethacin fully inhibited it. Furthermore, incubation of HUVECs with RWE increased eNOS (endothelial NOS) and COX-2 mRNA levels as well as phosphorylation of eNOS at Ser1177. RWE protects endothelial cells from tBHP-induced senescence. NO and COX-2, in addition to activation of SIRT1, play a critical role in the inhibition of senescence induction in human endothelial cells by RWE.

Increases in oxygen tension stimulate expression of ICAM-1 and VCAM-1 on human endothelial cells

1999 ◽  
Vol 276 (6) ◽  
pp. H2044-H2052 ◽  
Author(s):  
Carsten Willam ◽  
Ralf Schindler ◽  
Ulrich Frei ◽  
Kai-Uwe Eckardt
Keyword(s):  
Endothelial Cells ◽  
Adhesion Molecules ◽  
Adhesion Molecule ◽  
Umbilical Vein ◽  
Ischemia Reperfusion ◽  
White Blood Cells ◽  
Free Radical Scavengers ◽  
Intercellular Adhesion Molecule ◽  
Induced Expression ◽  
Tnf Α

Leukocyte infiltration plays a major role in ischemia-associated organ dysfunction and damage. A crucial step for extravasation of white blood cells is binding of leukocyte β-integrins to endothelial adhesion molecules intercellular adhesion molecule-1 (ICAM-1) and vascular adhesion molecule-1 (VCAM-1). To test for direct effects of oxygen on this process we studied ICAM-1 and VCAM-1 expression in human dermal microvascular and umbilical vein endothelial cells (EC) exposed to different oxygen tensions in the absence or presence of tumor necrosis factor-α (TNF-α). Hypoxia (95% N2-5% CO2) resulted in a downregulation of basal but not TNF-α-induced expression of ICAM-1 and VCAM-1. Subsequent rises in oxygen (21, 40, or 95% O2) led to marked increase of ICAM-1 and VCAM-1 cell surface and mRNA expression in both EC types, which after 16 h amounted to about one-third to one-half of maximal TNF-α-induced expression. This increase was greatest after 0.5-h hypoxia and was blunted with prolonged hypoxic preincubation. Exposure of cells preincubated under “normoxic” (21% O2) conditions to hyperoxia (40 or 95% O2) also enhanced expression of both adhesion molecules, but the increase was lower than in cells preexposed to hypoxia. The nitric oxide synthesis inhibitor NG-nitro-l-arginine methyl ester (l-NAME) enhanced ICAM-1 and VCAM-1 expression under basal and hypoxic conditions, but in the presence of l-NAME, levels in reoxygenated cells were not higher than basal levels. Moreover, the oxygen-induced rise could be mimicked by addition of H2O2to normoxic cells, and the oxygen-induced expression of VCAM-1 but not of ICAM-1 was inhibited by addition of the free radical scavengers superoxide dismutase, N-acetyl-l-cysteine, and pyrrolidinedithiocarbamate. These data indicate that an increase in oxygen availability stimulates ICAM-1 and VCAM-1 expression on micro- and macrovascular EC, which may contribute to adhesion and transmigration of different leukocyte populations in ischemia-reperfusion injuries.

Anti-Inflammatory Effects ofDanshenon Human Vascular Endothelial Cells in Culture

2013 ◽  
Vol 41 (05) ◽  
pp. 1065-1077 ◽  
Author(s):  
Christian Stumpf ◽  
Quiaoling Fan ◽  
Christian Hintermann ◽  
Dorette Raaz ◽  
Ingrid Kurfürst ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Adhesion Molecules ◽  
Vascular Endothelial Cells ◽  
Umbilical Vein ◽  
Cellular Adhesion ◽  
Tanshinone Iia ◽  
Protective Effects ◽  
Induced Expression ◽  
Tnf Α ◽  
Anti Inflammatory

Inflammation plays a crucial role in the pathophysiology of atherosclerosis. Besides cytokines, chemokines and cell adhesion molecules, CD40 and P-selectin play important roles as key regulators of the inflammatory process in atherosclerosis. Danshen (DS) is commonly used in traditional Chinese medicine for therapy of cardiovascular diseases such as coronary artery disease. The aim of the present study was to evaluate the protective effects of DS with respect to possible anti-inflammatory effects. Human umbilical vein endothelial cells as well as platelets were incubated with an extract of DS or one of its major ingredients salvianolic acid B (Sal B), tanshinone IIA (Tansh) and protocatechuic acid (Protoc) under tumor necrosis factor (TNF)-α or ADP stimulation. Expression of CD40 and cellular adhesion molecules (VCAM-1/ICAM-1) were assessed via flow cytometry. Levels of interleukin (IL)-6, IL-8, monocyte-chemoattractant-protein (MCP)-1 as well as soluble VCAM1 and ICAM-1 in the supernatants were examined via luminex based analysis. Treatment with DS attenuated TNF-α induced expression of CD40. Furthermore, the expression of VCAM-1 and ICAM-1 as well as the release of soluble VCAM-1 and ICAM-1 were downregulated. In the cell supernatants we also observed a significant reduction of IL-6, IL8 and MCP-1. DS and its major ingredients, Sal B and Protoc, significantly inhibited TNF-induced expression and release of adhesion molecules, cytokines and chemokines as well as ADP-induced expression of platelet P-selectin. Because of the key roles of inflammatory mediators in the etiology of atherosclerosis, this work provides useful insight in understanding the pharmacological efficacy of Chinese herbal medicine.

scholarly journals Superoxide, H2O2, and iron are required for TNF-α-induced MCP-1 gene expression in endothelial cells: role of Rac1 and NADPH oxidase

2004 ◽  
Vol 286 (3) ◽  
pp. H1001-H1007 ◽  
Author(s):  
Xi-Lin Chen ◽  
Qiang Zhang ◽  
Ruozhi Zhao ◽  
Russell M. Medford
Keyword(s):  
Gene Expression ◽  
Endothelial Cells ◽  
Nadph Oxidase ◽  
Gene Activation ◽  
Dominant Negative ◽  
Mrna Levels ◽  
Text Generation ◽  
Mrna Accumulation ◽  
Tnf Α ◽  
Diphenylene Iodonium

Reactive oxygen species (ROS) play an important but not yet fully defined role in the expression of inflammatory genes such as monocyte chemoattractant protein (MCP)-1. We used complementary molecular and biochemical approaches to explore the roles of specific ROS and their molecular linkage to inflammatory signaling in endothelial cells. Adenovirus-mediated expression of superoxide dismutase and catalase inhibited TNF-α-induced MCP-1 gene expression, suggesting important roles of superoxide ([Formula: see text]) and H2O2 in MCP-1 gene activation. In addition, the iron chelator 1,2-dimethyl-3-hydroxypyridin-4-one and the hydroxyl radical scavengers dimethylthiourea and dimethyl sulfoxide inhibited TNF-α-induced MCP-1 expression, suggesting important roles of iron and hydroxyl radicals in inflammatory signal activation. In contrast, scavenging of peroxynitrite with 5,10,15,20-tetrakis-(4-sulfonatophenyl)prophyrinato iron (III) chloride had no effect on TNF-α-induced MCP-1 expression. Inhibition of NADPH oxidase, the major oxidase responsible for [Formula: see text] generation, with diphenylene iodonium suppressed TNF-α-induced MCP-1 mRNA accumulation. Rac1 is an upstream signaling molecule for the activation of NADPH oxidase and [Formula: see text] generation. Expression of dominant negative N17Rac1 by adenovirus suppressed TNF-α-induced MCP-1 mRNA levels and MCP-1 protein secretion. Expression of N17Rac1 inhibited TNF-α-induced MCP-1 and NF-κB transcriptional activity. These data suggest that ROS such as superoxide and H2O2 derived from Rac1-activated NADPH oxidase mediate TNF-α-induced MCP-1 expression in endothelial cells.

Sphingosine kinase-1 mediates TNF-α-induced MCP-1 gene expression in endothelial cells: upregulation by oscillatory flow

2004 ◽  
Vol 287 (4) ◽  
pp. H1452-H1458 ◽  
Author(s):  
Xi-Lin Chen ◽  
Janice Y. Grey ◽  
Suzanne Thomas ◽  
Fei-Hua Qiu ◽  
Russell M. Medford ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Shear Stress ◽  
P38 Mapk ◽  
Protein Secretion ◽  
Oscillatory Flow ◽  
Fluid Shear Stress ◽  
Mrna Levels ◽  
Flow Shear ◽  
Tnf Α ◽  
Flow Shear Stress

Atherosclerosis is a focal inflammatory disease and preferentially occurs in areas of low fluid shear stress and oscillatory flow, whereas the risk of atherosclerosis is decreased in regions of high fluid shear stress and steady laminar flow. Sphingosine kinase-1 (SphK1) catalyzes the conversion of sphingosine to sphingosine-1 phosphate (S1P), a sphingolipid metabolite that plays important roles in angiogenesis, inflammation, and cell growth. In the present study, we demonstrated that exposure of human aortic endothelial cells to oscillatory flow (shear stress, ±5 dyn/cm2for 48 h) resulted in a marked increase in SphK1 mRNA levels compared with endothelial cells kept in static culture. In contrast, laminar flow (shear stress, 20 dyn/cm2for 48 h) decreased SphK1 mRNA levels. We further investigated the role of SphK1 in TNF-α-induced expression of inflammatory genes, such as monocyte chemoattractant protein-1 (MCP-1) and VCAM-1 by using small interfering RNA (siRNA) specifically for SphK1. Treatment of endothelial cells with SphK1 siRNA suppressed TNF-α-induced increase in MCP-1 mRNA levels, MCP-1 protein secretion, and activation of p38 MAPK. SphK1 siRNA also inhibited TNF-α-induced cell surface expression of VCAM-1, but not ICAM-1, protein. Exposure of endothelial cells to S1P led to an increase in MCP-1 protein secretion and MCP-1 mRNA levels and activation of NF-κB-mediated transcriptional activity. Treatment of endothelial cells with the p38 MAPK inhibitor SB-203580 suppressed S1P-induced MCP-1 protein secretion. These data suggest that SphK1 mediates TNF-α-induced MCP-1 gene expression through a p38 MAPK-dependent pathway and may participate in oscillatory flow-mediated proinflammatory signaling pathway in the vasculature.

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