Chunghyuldan activates NOS mRNA expression and suppresses VCAM-1 mRNA expression in human endothelial cells

2005 ◽  
Vol 83 (12) ◽  
pp. 1101-1108 ◽  
Author(s):  
Seong-Uk Park ◽  
Woo-Sang Jung ◽  
Sang-Kwan Moon ◽  
Chang-Nam Ko ◽  
Ki-Ho Cho ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Umbilical Vein ◽  
Stroke Recurrence ◽  
No Production ◽  
Clinical Effects ◽  
Dependent Manner ◽  
Human Endothelial Cells ◽  
Synthesis Inhibitor ◽  
Anti Inflammatory

Chunghyuldan (CHD), a combinatorial drug that has antihyperlipidemic and anti-inflammatory activities, has been shown to improve arterial stiffness and inhibit stroke recurrence in clinical study. To understand the molecular basis of CHD's clinical effects, we explored its effect on cell proliferation and expression of nitric oxide synthase (NOS) and vascular cell adhesion molecule (VCAM-1) in human umbilical vein endothelial cells (HUVECs). Cell number counting and [3H]thymidine incorporation assay demonstrated that nontoxic doses of CHD have an inhibitory effect on DNA synthesis and suppress cell cycle progression of HUVECs. CHD treatment led to a marked induction of NO production through up-regulation of NOS mRNA expression in a dose- and time-dependent manner, whereas it suppressed VCAM-1 expression. CHD inhibition of VCAM-1 expression was totally blocked by pretreatment with the NO synthesis inhibitor L-NMMA, whereas pretreatment with the NO donor DETA-NO further decreased VCAM-1 level in CHD-treated HUVECs, indicating that VCAM-1 regulation by CHD is mediated through increased NO synthesis by CHD. In addition, TNF-α-mediated VCAM-1 activation was substantially impeded by CHD treatment. Collectively, our data suggest that anti-inflammatory or anti-hyperlipidemic effects of CHD might be associated with its ability to activate NO production and suppress VCAM-1 expression in human endothelial cells.

scholarly journals Endothelial AMP-Activated Kinase α1 Phosphorylates eNOS on Thr495 and Decreases Endothelial NO Formation

2018 ◽  
Vol 19 (9) ◽  
pp. 2753 ◽  
Author(s):  
Nina Zippel ◽  
Annemarieke Loot ◽  
Heike Stingl ◽  
Voahanginirina Randriamboavonjy ◽  
Ingrid Fleming ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Vascular Reactivity ◽  
Umbilical Vein ◽  
No Production ◽  
Dependent Manner ◽  
Calmodulin Binding ◽  
Umbilical Vein Endothelial Cells ◽  
Endothelial Nitric Oxide ◽  
Specific Deletion

AMP-activated protein kinase (AMPK) is frequently reported to phosphorylate Ser1177 of the endothelial nitric-oxide synthase (eNOS), and therefore, is linked with a relaxing effect. However, previous studies failed to consistently demonstrate a major role for AMPK on eNOS-dependent relaxation. As AMPK also phosphorylates eNOS on the inhibitory Thr495 site, this study aimed to determine the role of AMPKα1 and α2 subunits in the regulation of NO-mediated vascular relaxation. Vascular reactivity to phenylephrine and acetylcholine was assessed in aortic and carotid artery segments from mice with global (AMPKα−/−) or endothelial-specific deletion (AMPKαΔEC) of the AMPKα subunits. In control and AMPKα1-depleted human umbilical vein endothelial cells, eNOS phosphorylation on Ser1177 and Thr495 was assessed after AMPK activation with thiopental or ionomycin. Global deletion of the AMPKα1 or α2 subunit in mice did not affect vascular reactivity. The endothelial-specific deletion of the AMPKα1 subunit attenuated phenylephrine-mediated contraction in an eNOS- and endothelium-dependent manner. In in vitro studies, activation of AMPK did not alter the phosphorylation of eNOS on Ser1177, but increased its phosphorylation on Thr495. Depletion of AMPKα1 in cultured human endothelial cells decreased Thr495 phosphorylation without affecting Ser1177 phosphorylation. The results of this study indicate that AMPKα1 targets the inhibitory phosphorylation Thr495 site in the calmodulin-binding domain of eNOS to attenuate basal NO production and phenylephrine-induced vasoconstriction.

scholarly journals Bavachalcone Enhances RORαExpression, Controls Bmal1 Circadian Transcription, and Depresses Cellular Senescence in Human Endothelial Cells

2015 ◽  
Vol 2015 ◽  
pp. 1-11 ◽  
Author(s):  
Yanqi Dang ◽  
Shuang Ling ◽  
Jing Ma ◽  
Rongzhen Ni ◽  
Jin-Wen Xu
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Replicative Senescence ◽  
Orphan Receptor ◽  
Luciferase Reporter ◽  
Dependent Manner ◽  
Orphan Nuclear Receptor ◽  
Human Endothelial Cells ◽  
Peripheral Tissues ◽  
Natural Medicine

The circadian clock regulates many aspects of (patho)physiology in the central nervous system and in the peripheral tissues. RAR-related orphan receptorα(RORα), an orphan nuclear receptor, is involved in circadian rhythm regulation, including regulation of cardiovascular function. Bavachalcone, a prenylchalcone, is a major bioactive chalcone isolated fromPsoralea corylifolia. This natural ingredient activated RORα1 luciferase reporter activity on drug screening. In addition, bavachalcone induced RORα1 expression in mRNA and protein levels in a dose-dependent manner and enhanced the circadian amplitude of Bmal1 mRNA expression after serum shock. Moreover, bavachalcone suppressed senescence in human endothelial cells and mRNA expression ofp16ink4a(a marker of replicative senescence) and IL-1α(a proinflammatory cytokine of the senescence-associated secretory phenotype). These inhibitory effects were partially reversed by the RORαinhibitor VPR-66. Our results demonstrate that bavachalcone, as a natural medicine ingredient, has a pharmacological function in regulating RORα1.

scholarly journals Epigallocatechin gallate inhibits endothelial exocytosis

2008 ◽  
Vol 389 (7) ◽  
Author(s):  
Munekazu Yamakuchi ◽  
Clare Bao ◽  
Marcella Ferlito ◽  
Charles J. Lowenstein
Keyword(s):  
Endothelial Cells ◽  
Green Tea ◽  
Umbilical Vein ◽  
Vascular Inflammation ◽  
Epigallocatechin Gallate ◽  
Initial Step ◽  
No Production ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Akt Phosphorylation

Abstract Consumption of green tea is associated with a decrease in cardiovascular mortality. The beneficial health effects of green tea are attributed in part to polyphenols, organic compounds found in tea that lower blood pressure, reduce body fat, decrease LDL cholesterol, and inhibit inflammation. We hypothesized that epigallocatechin gallate (EGCG), the most abundant polyphenol in tea, inhibits endothelial exocytosis, the initial step in leukocyte trafficking and vascular inflammation. To test this hypothesis, we treated human umbilical-vein endothelial cells with EGCG and other polyphenols, and then measured endothelial exocytosis. We found that EGCG decreases endothelial exocytosis in a concentration-dependent manner, with the effects most prominent after 4 h of treatment. Other catechin polyphenols had no effect on endothelial cells. By inhibiting endothelial exocytosis, EGCG decreases leukocyte adherence to endothelial cells. In searching for the mechanism by which EGCG affects endothelial cells, we found that EGCG increases Akt phosphorylation, eNOS phosphorylation, and nitric oxide (NO) production. NOS inhibition revealed that NO mediates the anti-inflammatory effects of EGCG. Our data suggest that polyphenols can decrease vascular inflammation by increasing the synthesis of NO, which blocks endothelial exocytosis.

Decrease of tetrahydrobiopterin and NO generation in endothelial cells exposed to simulated diving

2019 ◽  
pp. 159-169
Author(s):  
Ronja Hesthammer ◽  
◽  
Torunn Eide ◽  
Eimar Thorsen ◽  
Asbjørn M. Svardal ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Decompression Sickness ◽  
Synergistic Effects ◽  
Umbilical Vein ◽  
Bubble Formation ◽  
No Production ◽  
Dependent Manner ◽  
Hyperoxic Exposure ◽  
Umbilical Vein Endothelial Cells ◽  
Dose Dependent

Purpose: Nitric oxide (NO) has been shown to protect against bubble formation and the risk of decompression sickness. We hypothesize that oxidation of tetrahydrobiopterin (BH4) leads to a decreased production of NO during simulated diving. Methods: Human umbilical vein endothelial cells (HUVEC) were exposed to hyperoxia or simulated diving for 24 hours. The levels of biopterins (BH4, BH2 and B) were determined by LC-MS/MS, and the production of NO by monitoring the conversion of L-arginine to L-citrulline. Results: Exposure to hyperoxia decreased BH4 in a dose-dependent manner; by 48 ± 15% following exposure to 40 kPa O2 (P < 0.001 vs. control at 20 kPa O2), and 70 ± 16% following exposure to 60 kPa O2. Exposure to 40 kPa O2 decreased NO production by 25 ± 9%, but there was no further decrease when increasing oxygen exposure to 60 kPa (25 ± 10%). No additional effects of simulated diving were observed, indicating no additive or synergistic effects of hyperbaria and hyperoxia on the BH4 level or NO generation. Conclusion: NO generation in intact human endothelial cells was decreased by simulated diving, as well as by hyperoxic exposure, while BH4 levels seem to be affected only by hyperoxia. Hence, the results suggest that BH4 is not the sole determinant of NO generation in HUVEC.

Inflammation-induced up-regulation of TLR2 expression in human endothelial cells is independent of differential methylation in the TLR2 promoter CpG island

2011 ◽  
Vol 18 (1) ◽  
pp. 112-123 ◽  
Author(s):  
Britta Diesel ◽  
Nadège Ripoche ◽  
Rebecca T Risch ◽  
Sascha Tierling ◽  
Jörn Walter ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Cpg Island ◽  
Umbilical Vein ◽  
Methylation Pattern ◽  
Human Endothelial Cells ◽  
Tlr2 Expression ◽  
Inflammatory Conditions ◽  
Promoter Dna ◽  
Umbilical Vein Endothelial Cells

Toll-like receptors play an important role in endothelial inflammation; however, little is known on the mechanisms regulating their expression. Differential promoter DNA methylation is an increasingly recognized mechanism that determines a switch between gene silencing and gene transcription. We hypothesized that epigenetic mechanisms are involved in the regulation of endothelial TLR2 expression because of the localization of the TLR2 promoter on a CpG-island. Resting human umbilical vein endothelial cells (HUVECs) displayed rather low TLR2 mRNA expression, while a strong expression increase occurred under inflammatory conditions. We examined the TLR2 promoter methylation pattern in resting HUVECs and compared it to cells treated either with the inflammatory cytokine TNF-α or the DNA-demethylating agent 5-azacytidine. DNA bisulfite conversion was followed by either genomic sequencing or single nucleotide primer extension (SNuPE) HPLC. Results of both techniques showed a low- or non-methylated TLR2 promoter in resting HUVECs and no alteration of the methylation pattern under inflammatory conditions. Whereas 5-azacytidine significantly increased the mRNA expression of the epigenetically regulated gene H19, TLR2 expression was not affected. Taken together, employing different methodological approaches, our data show no implication of methylation pattern changes in inflammatory induction of TLR2 expression in human endothelial cells.

Momordica charantia Inhibits Inflammatory Responses in Murine Macrophages via Suppression of TAK1

2018 ◽  
Vol 46 (02) ◽  
pp. 435-452 ◽  
Author(s):  
Woo Seok Yang ◽  
Eunju Yang ◽  
Min-Jeong Kim ◽  
Deok Jeong ◽  
Deok Hyo Yoon ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Molecular Mechanisms ◽  
Transforming Growth Factor ◽  
Momordica Charantia ◽  
Luciferase Reporter ◽  
No Production ◽  
Dependent Manner ◽  
Raw264.7 Macrophages ◽  
Anti Inflammatory ◽  
Dose Dependent

Momordica charantia known as bitter melon is a representative medicinal plant reported to exhibit numerous pharmacological activities such as antibacterial, antidiabetic, anti-inflammatory, anti-oxidant, antitumor, and hypoglycemic actions. Although this plant has high ethnopharmacological value for treating inflammatory diseases, the molecular mechanisms by which it inhibits the inflammatory response are not fully understood. In this study, we aim to identify the anti-inflammatory mechanism of this plant. To this end, we studied the effects of its methanol extract (Mc-ME) on lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages. Specifically, we evaluated nitric oxide (NO) production, mRNA expression of inflammatory genes, luciferase reporter gene activity, and putative molecular targets. Mc-ME blocked NO production in a dose-dependent manner in RAW264.7 cells; importantly, no cytotoxicity was observed. Moreover, the mRNA expression levels of inducible NO synthase (iNOS) and cyclooxygenase (COX)-2 were decreased by Mc-ME treatment in a dose-dependent manner. Luciferase assays and nuclear lysate immunoblotting analyses strongly indicated that Mc-ME decreases the levels of p65 [a nuclear factor (NF)-[Formula: see text]B subunit] and c-Fos [an activator protein (AP)-1 subunit]. Whole lysate immunoblotting assays, luciferase assays, and overexpression experiments suggested that transforming growth factor [Formula: see text]-activated kinase 1 (TAK1) is targeted by Mc-ME, thereby suppressing NF-[Formula: see text]B and AP-1 activity via downregulation of extracellular signal-regulated kinases (ERKs) and AKT. These results strongly suggest that Mc-ME exerts its anti-inflammatory activity by reducing the action of TAK1, which also affects the activation of NF-[Formula: see text]B and AP-1.

scholarly journals Orosomucoid has a cAMP-dependent effect on human endothelial cells and inhibits the action of histamine

2000 ◽  
Vol 278 (5) ◽  
pp. H1725-H1731 ◽  
Author(s):  
Jenny Sörensson ◽  
Maria Ohlson ◽  
Anna Björnson ◽  
Börje Haraldsson
Keyword(s):  
Endothelial Cells ◽  
Inflammatory Mediator ◽  
Umbilical Vein ◽  
Primary Cultures ◽  
Microvascular Endothelial Cells ◽  
Capillary Barrier ◽  
Human Endothelial Cells ◽  
Anti Inflammatory ◽  
Umbilical Vein Endothelial Cells ◽  
Dose Dependent

The plasma protein orosomucoid (α1-acid glycoprotein) has previously been shown to constitute a critical component of the capillary barrier. The protein has also been suggested to act as an anti-inflammatory mediator in a diversity of experimental situations. Recently we reported that orosomucoid is synthesized by the microvascular endothelial cells per se. In the present study, the effects of orosomucoid on primary cultures of human umbilical vein endothelial cells (HUVEC) were studied using the Cytosensor microphysiometer. We found that 1) orosomucoid (0.01 g/l) increased the metabolic activity of HUVEC as reflected by the increased acidification rate of +14 ± 1%; 2) pretreatment with 0.5 mM 8-bromo-cAMP for 20 min markedly and reversibly inhibited the effect of orosomucoid, whereas 8-bromo-cGMP did not; 3) histamine elicited a dose-dependent response that was abolished by pretreatment with either cAMP or cGMP; and finally, 4) pretreatment of HUVEC for 6 min with orosomucoid (0.01 g/l) inhibited the action of histamine. In summary, this is the first report demonstrating that orosomucoid affects human endothelial cells and that it does so by using cAMP as a second messenger. This provides an explanation for previous findings of anti-inflammatory effects of the protein and shows that orosomucoid affects the endothelium during both normal and pathophysiological conditions.

scholarly journals Requirement for NF-κB in Transcriptional Activation of Monocyte Chemotactic Protein 1 by Chlamydia pneumoniae in Human Endothelial Cells

2000 ◽  
Vol 68 (7) ◽  
pp. 4282-4288 ◽  
Author(s):  
Robert E. Molestina ◽  
Richard D. Miller ◽  
Alex B. Lentsch ◽  
Julio A. Ramirez ◽  
James T. Summersgill
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Transcriptional Activation ◽  
Chlamydia Pneumoniae ◽  
Nuclear Translocation ◽  
Umbilical Vein ◽  
Specific Inhibitor ◽  
Human Endothelial Cells ◽  
Monocyte Chemotactic Protein ◽  
Chemotactic Protein

ABSTRACT Infection with Chlamydia pneumoniae, a causative agent of acute and chronic respiratory diseases, has recently been implicated as a potential risk factor in atherosclerosis. Atherosclerotic lesions are characterized by monocyte infiltration, which may be regulated by the chemokine monocyte chemotactic protein 1 (MCP-1). We have previously shown that C. pneumoniae infection stimulates MCP-1 production in human endothelial cells, an event which may be specific to this species of Chlamydia, sinceChlamydia trachomatis infection fails to induce this response. To examine the underlying mechanisms by which C. pneumoniae infection induces MCP-1 production in endothelial cells, the present study investigated the role of transcription factor NF-κB in MCP-1 mRNA expression. Human umbilical vein endothelial cells (HUVEC) were infected with the coronary isolate C. pneumoniae A-03 or with C. trachomatis L2, and MCP-1 mRNA expression was assessed after different periods of infection by reverse transcription-PCR. Expression of MCP-1 mRNA in C. pneumoniae-infected HUVEC was significantly elevated as early as 1 h postinfection and increased dramatically by 12 and 24 h compared to baseline controls. Nuclear translocation of NF-κB occurred by 30 min of infection, as determined by electrophoretic mobility shift assays and immunofluorescence staining. Treatment ofC. pneumoniae-infected HUVEC with parthenolide, a specific inhibitor of NF-κB activation, suppressed MCP-1 mRNA expression. In contrast, infection with C. trachomatis L2 did not induce MCP-1 mRNA in infected HUVEC and failed to activate NF-κB. Results from this study demonstrate a requirement for NF-κB activation in stimulation of MCP-1 gene expression by C. pneumoniae in human endothelial cells. Furthermore, the data suggest that, within the genus Chlamydia, functionally distinct signaling pathways leading to NF-κB activation are utilized by C. pneumoniaein endothelial cells during infection.

Purification, molecular cloning and mechanism of action of graminelysin I, a snake-venom-derived metalloproteinase that induces apoptosis of human endothelial cells

2001 ◽  
Vol 357 (3) ◽  
pp. 719-728 ◽  
Author(s):  
Wen Bin WU ◽  
Shin C. CHANG ◽  
Ming-Yi LIAU ◽  
Tur-Fu HUANG
Keyword(s):  
Endothelial Cells ◽  
Umbilical Vein ◽  
Matrix Proteins ◽  
Dependent Manner ◽  
Single Chain ◽  
Human Endothelial Cells ◽  
Putative Precursor ◽  
Induced Apoptosis ◽  
Umbilical Vein Endothelial Cells ◽  
Dose Dependent

Apoptosis, a programmed, physiological mode of cell death, is important in tissue homoeostasis. Here we report that a new metalloproteinase, graminelysin I, purified from Trimeresurus gramineus venom, induced apoptosis of human endothelial cells as examined by electrophoresis and flow cytometry. Graminelysin I contains only a metalloproteinase domain. It is a single-chain proteinase with a molecular mass of 27020Da. cDNA sequence analysis revealed that the disintegrin-like and cysteine-rich domains of the putative precursor protein of graminelysin I are likely to be processed post-translationally, producing the proteinase domain (graminelysin I). Graminelysin I cleaved the α chain of fibrinogen preferentially and cleaved the β chain either on longer incubation or at higher concentration. Graminelysin I inhibited the adhesion of human umbilical-vein endothelial cells (HUVECs) to immobilized fibrinogen and induced HUVECs detachment in a dose-dependent manner. These effects on HUVECs were abolished when graminelysin I was pretreated with EDTA. However, graminelysin I did not inhibit the adhesion of HUVECs to immobilized collagen. HUVECs were susceptible to death after treatment with graminelysin I when they were cultured on immobilized fibrinogen. In contrast, HUVECs were rather resistant to treatment with graminelysin I if they were cultured on immobilized collagen. Furthermore, graminelysin I induced apoptosis of HUVECs in a dose-dependent manner. Similarly, its apoptosis-inducing activity was blocked if it was treated with EDTA. These results suggest that the catalytic activity of graminelysin I on matrix proteins contributes to its apoptosis-inducing activity.

scholarly journals Repeated and long-term treatment with physiological concentrations of resveratrol promotes NO production in vascular endothelial cells

2011 ◽  
Vol 107 (6) ◽  
pp. 774-780 ◽  
Author(s):  
Satoru Takahashi ◽  
Yukiko Nakashima
Keyword(s):  
Endothelial Cells ◽  
Mrna Expression ◽  
Term Treatment ◽  
No Synthase ◽  
No Production ◽  
Dependent Manner ◽  
Long Term Treatment ◽  
Enos Mrna ◽  
Enos Protein

In the present study, we examined the effect of repeated and long-term treatment with resveratrol on NO production in endothelial cells as a model of routine wine consumption. Repeated treatment with resveratrol for 5 d resulted in an increase in endothelial NO synthase (eNOS) protein content and NO production in human umbilical vein endothelial cell (HUVEC) in a concentration-dependent manner. A significant increase in functional eNOS protein content was observed with resveratrol, even at 50 nm. In contrast, eNOS phosphorylation was not stimulated and inducible NO synthase (iNOS) was not detected after resveratrol treatment. Both eNOS protein and mRNA expression were promoted by 50 nm-resveratrol in a time-dependent manner. Increased eNOS mRNA expression in response to resveratrol was not decreased by an oestrogen receptor (ER) antagonist ICI182780, a PPARα inhibitor MK886 or a sirtuin inhibitor Salermide. However, a combination of ICI182780 and MK886 significantly inhibited resveratrol-induced eNOS mRNA expression. Salermide had no effect even in the presence of ICI182780 or MK886. These results demonstrate that resveratrol within the physiological range increases eNOS mRNA and protein expression through ER and PPARα activation, thereby promoting NO production in endothelial cells. eNOS induction might result from the accumulative effect of nanomolar concentrations of resveratrol. The present study results can account in part for the observation that cardiovascular benefits of red wine are experienced with routine consumption, but not with acute consumption.

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