scholarly journals β-Cyclodextrin Inhibits Monocytic Adhesion to Endothelial Cells through Nitric Oxide-Mediated Depletion of Cell Adhesion Molecules

Molecules ◽  
2020 ◽  
Vol 25 (16) ◽  
pp. 3575
Author(s):  
Sujeong Jang ◽  
Seongsoo Lee ◽  
Heonyong Park
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Cell Adhesion ◽  
No Production ◽  
Aortic Endothelial Cells ◽  
Enos Phosphorylation ◽  
Inflammatory Processes ◽  
Cell Adhesion Molecule 1 ◽  
Endothelial Nitric Oxide ◽  
Inflammatory Effect

Cyclodextrins (CDs) are used as drug delivery agents. In this study, we examined whether CDs have an inflammatory effect on endothelial cells. First, we found that β-CD promoted cell proliferation in bovine aortic endothelial cells and elevated nitric oxide (NO) production through dephosphorylation of threonine-495 (T-495) in endothelial nitric oxide synthetase (eNOS). Dephosphorylation of T-495 is known to activate eNOS. Phosphorylation of T-495 was found to be catalyzed by protein kinase Cε (PKCε). We then found that β-CD inhibits binding of PKCε to diacylglycerol (DAG) via formation of a β-CD-DAG complex, indicating that β-CD inactivates PKCε. Furthermore, β-CD controls activation of PKCε by reducing the recruitment of PKCε into the plasma membrane. Finally, β-CD inhibits expression of intercellular and vascular cell adhesion molecule-1 by increasing NO via control of PKCε/eNOS and suppression of THP-1 cell adhesion to endothelial cells. These findings imply that β-CD plays an important role in anti-inflammatory processes.

scholarly journals Cadmium reduces nitric oxide production by impairing phosphorylation of endothelial nitric oxide synthase

2008 ◽  
Vol 86 (1) ◽  
pp. 1-10 ◽  
Author(s):  
Syamantak Majumder ◽  
Ajit Muley ◽  
Gopi Krishna Kolluru ◽  
Samir Saurabh ◽  
K. P. Tamilarasan ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Function ◽  
Egg Yolk ◽  
Cellular Migration ◽  
No Production ◽  
Enos Phosphorylation ◽  
Low Levels ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

Cadmium (Cd) perturbs vascular health and interferes with endothelial function. However, the effects of exposing endothelial cells to low doses of Cd on the production of nitric oxide (NO) are largely unknown. The objective of the present study was to evaluate these effects by using low levels of CdCl2 concentrations, ranging from 10 to 1000 nmol/L. Cd perturbations in endothelial function were studied by employing wound-healing and MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assays. The results suggest that a CdCl2 concentration of 100 nmol/L maximally attenuated NO production, cellular migration, and energy metabolism in endothelial cells. An egg yolk angiogenesis model was employed to study the effect of Cd exposure on angiogenesis. The results demonstrate that NO supplementation restored Cd-attenuated angiogenesis. Immunofluorescence, Western blot, and immuno-detection studies showed that low levels of Cd inhibit NO production in endothelial cells by blocking eNOS phosphorylation, which is possibly linked to processes involving endothelial function and dysfunction, including angiogenesis.

scholarly journals Phytoestrogen Genistein Up-Regulates Endothelial Nitric Oxide Synthase Expression Via Activation of cAMP Response Element-Binding Protein in Human Aortic Endothelial Cells

Endocrinology ◽  
2012 ◽  
Vol 153 (7) ◽  
pp. 3190-3198 ◽  
Author(s):  
Hongwei Si ◽  
Jie Yu ◽  
Hongling Jiang ◽  
Hazel Lum ◽  
Dongmin Liu
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Nitric Oxide Synthase ◽  
No Production ◽  
Enos Expression ◽  
Camp Response Element ◽  
Aortic Endothelial Cells ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide ◽  
Human Aortic Endothelial Cells

We previously reported that genistein, a phytoestrogen, up-regulates endothelial nitric oxide synthase (eNOS) and prevents hypertension in rats that are independent of estrogen signaling machinery. However, how genistein regulates eNOS expression is unknown. In the present study, we show that genistein enhanced eNOS expression and NO synthesis in primary human aortic endothelial cells. Inhibition of extracellular signal regulated kinase, phosphoinositol-3 kinase, or protein kinase C did not affect genistein-enhanced eNOS expression and NO synthesis. However, chemical inhibition of protein kinase A (PKA) or adenoviral transfer of the specific endogenous PKA inhibitor gene completely abolished PKA activity and genistein-stimulated eNOS expression and NO production. Accordingly, genistein induced PKA activity and subsequent phosphorylation of cAMP response element (CRE)-binding protein (CREB) at Ser133. Suppression of CREB by small interfering RNA transfection abolished genistein-enhanced eNOS expression and NO production. Consistently, deletion of the CRE site within human eNOS promoter eliminated genistein-stimulated eNOS promoter activity. These findings provide the first evidence to our knowledge that genistein may play a beneficial role in vascular function through targeting the PKA/CREB/eNOS/NO signaling pathway.

Tumor necrosis factor-α reduces argininosuccinate synthase expression and nitric oxide production in aortic endothelial cells

2007 ◽  
Vol 293 (2) ◽  
pp. H1115-H1121 ◽  
Author(s):  
Bonnie L. Goodwin ◽  
Laura C. Pendleton ◽  
Monique M. Levy ◽  
Larry P. Solomonson ◽  
Duane C. Eichler
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Elevated Serum ◽  
Proximal Promoter ◽  
No Production ◽  
Aortic Endothelial Cells ◽  
Tnf Α ◽  
Argininosuccinate Synthase ◽  
Endothelial Nitric Oxide ◽  
Aortic Endothelial

Endothelial dysfunction associated with elevated serum levels of TNF-α observed in diabetes, obesity, and congenital heart disease results, in part, from the impaired production of endothelial nitric oxide (NO). Cellular NO production depends absolutely on the availability of arginine, substrate of endothelial nitric oxide synthase (eNOS). In this report, evidence is provided demonstrating that treatment with TNF-α (10 ng/ml) suppresses not only eNOS expression but also the availability of arginine via the coordinate suppression of argininosuccinate synthase (AS) expression in aortic endothelial cells. Western blot and real-time RT-PCR demonstrated a significant and dose-dependent reduction of AS protein and mRNA when treated with TNF-α with a corresponding decrease in NO production. Reporter gene analysis demonstrated that TNF-α suppresses the AS proximal promoter, and EMSA analysis showed reduced binding to three essential Sp1 elements. Inhibitor studies suggested that the repression of AS expression by TNF-α may be mediated, in part, via the NF-κB signaling pathway. These findings demonstrate that TNF-α coordinately downregulates eNOS and AS expression, resulting in a severely impaired citrulline-NO cycle. The downregulation of AS by TNF-α is an added insult to endothelial function because of its important role in NO production and in endothelial viability.

scholarly journals Role of local production of endothelium-derived nitric oxide on cGMP signaling and S-nitrosylation

2010 ◽  
Vol 298 (1) ◽  
pp. H112-H118 ◽  
Author(s):  
Jin Qian ◽  
Qian Zhang ◽  
Jarrod E. Church ◽  
David W. Stepp ◽  
Radu D. Rudic ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Vascular Function ◽  
Soluble Guanylate Cyclase ◽  
No Production ◽  
Local Synthesis ◽  
Intracellular Location ◽  
Aortic Endothelial Cells ◽  
Cgmp Signaling ◽  
Endothelial Nitric Oxide

Nitric oxide (NO), synthesized by endothelial nitric oxide synthase (eNOS), exerts control over vascular function via two distinct mechanisms, the activation of soluble guanylate cyclase (sGC)/cGMP-dependent signaling or through S-nitrosylation of proteins with reactive thiols ( S-nitrosylation). Previous studies in cultured endothelial cells revealed that eNOS targeted to the plasma membrane (PM) releases greater amounts of NO compared with Golgi tethered eNOS. However, the significance of eNOS localization to sGC-dependent or -independent signaling is not known. Here we show that PM-targeted eNOS, when expressed in human aortic endothelial cells (HAEC) and isolated blood vessels, increases sGC/cGMP signaling to a greater extent than Golgi-localized eNOS. The ability of local NO production to influence sGC-independent mechanisms was also tested by monitoring the secretion of Von Willebrand factor (vWF), which is tonically inhibited by the S-nitrosylation of N-ethylmaleimide sensitive factor (NSF). In eNOS “knockdown” HAECs, vWF secretion was attenuated to a greater degree by PM eNOS compared with a Golgi-restricted eNOS. Moreover, the PM-targeted eNOS induced greater S-nitrosylation of NSF vs. Golgi eNOS. To distinguish between the amount of NO generated and the intracellular location of synthesis, we expressed Golgi and PM-targeted calcium-insensitive forms of eNOS in HAEC. These constructs, which generate equal amounts of NO regardless of location, produced equivalent increases in cGMP in bioassays and equal inhibition of vWF secretion. We conclude that the greater functional effects of PM eNOS are due to the increased amount of NO produced rather than effects derived from the local synthesis of NO.

scholarly journals Rho GTPase/Rho Kinase Negatively Regulates Endothelial Nitric Oxide Synthase Phosphorylation through the Inhibition of Protein Kinase B/Akt in Human Endothelial Cells

2002 ◽  
Vol 22 (24) ◽  
pp. 8467-8477 ◽  
Author(s):  
Xiu-Fen Ming ◽  
Hema Viswambharan ◽  
Christine Barandier ◽  
Jean Ruffieux ◽  
Kozo Kaibuchi ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Nitric Oxide Synthase ◽  
Rho Gtpase ◽  
Protein Kinase B ◽  
Rho Kinase ◽  
Enos Expression ◽  
Enos Phosphorylation ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

ABSTRACT Endothelial nitric oxide synthase (eNOS) is an important regulator of cardiovascular homeostasis by production of nitric oxide (NO) from vascular endothelial cells. It can be activated by protein kinase B (PKB)/Akt via phosphorylation at Ser-1177. We are interested in the role of Rho GTPase/Rho kinase (ROCK) pathway in regulation of eNOS expression and activation. Using adenovirus-mediated gene transfer in human umbilical vein endothelial cells (HUVECs), we show here that both active RhoA and ROCK not only downregulate eNOS gene expression as reported previously but also inhibit eNOS phosphorylation at Ser-1177 and cellular NO production with concomitant suppression of PKB activation. Moreover, coexpression of a constitutive active form of PKB restores the phosphorylation but not gene expression of eNOS in the presence of active RhoA. Furthermore, we show that thrombin inhibits eNOS phosphorylation, as well as expression via Rho/ROCK pathway. Expression of the active PKB reverses eNOS phosphorylation but has no effect on downregulation of eNOS expression induced by thrombin. Taken together, these data demonstrate that Rho/ROCK pathway negatively regulates eNOS phosphorylation through inhibition of PKB, whereas it downregulates eNOS expression independent of PKB.

Effects of homocysteine on endothelial nitric oxide production

2000 ◽  
Vol 279 (4) ◽  
pp. F671-F678 ◽  
Author(s):  
Xiaohui Zhang ◽  
Hong Li ◽  
Haoli Jin ◽  
Zachary Ebin ◽  
Sergey Brodsky ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Redox State ◽  
Calcium Ionophore ◽  
No Production ◽  
Cellular Redox ◽  
A Cell ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide ◽  
Peroxynitrite Scavengers

Hyperhomocysteinemia (HHCy) is an independent and graded cardiovascular risk factor. HHCy is prevalent in patients with chronic renal failure, contributing to the increased mortality rate. Controversy exists as to the effects of HHCy on nitric oxide (NO) production: it has been shown that HHCy both increases and suppresses it. We addressed this problem by using amperometric electrochemical NO detection with a porphyrinic microelectrode to study responses of endothelial cells incubated with homocysteine (Hcy) to the stimulation with bradykinin, calcium ionophore, or l-arginine. Twenty-four-hour preincubation with Hcy (10, 20, and 50 μM) resulted in a gradual decline in responsiveness of endothelial cells to the above stimuli. Hcy did not affect the expression of endothelial nitric oxide synthase (eNOS), but it stimulated formation of superoxide anions, as judged by fluorescence of dichlorofluorescein, and peroxynitrite, as detected by using immunoprecipitation and immunoblotting of proteins modified by tyrosine nitration. Hcy did not directly affect the ability of recombinant eNOS to generate NO, but oxidation of sulfhydryl groups in eNOS reduced its NO-generating activity. Addition of 5-methyltetrahydrofolate restored NO responses to all agonists tested but affected neither the expression of the enzyme nor formation of nitrotyrosine-modified proteins. In addition, a scavenger of peroxynitrite or a cell-permeant superoxide dismutase mimetic reversed the Hcy-induced suppression of NO production by endothelial cells. In conclusion, electrochemical detection of NO release from cultured endothelial cells demonstrated that concentrations of Hcy >20 μM produce a significant indirect suppression of eNOS activity without any discernible effects on its expression. Folates, superoxide ions, and peroxynitrite scavengers restore the NO-generating activity to eNOS, collectively suggesting that cellular redox state plays an important role in HCy-suppressed NO-generating function of this enzyme.

scholarly journals Cilostazol Induces eNOS and TM Expression via Activation with Sirtuin 1/Krüppel-like Factor 2 Pathway in Endothelial Cells

2021 ◽  
Vol 22 (19) ◽  
pp. 10287
Author(s):  
Chih-Hsien Wu ◽  
Yi-Lin Chiu ◽  
Chung-Yueh Hsieh ◽  
Guo-Shiang Tsung ◽  
Lian-Shan Wu ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Nitric Oxide Synthase ◽  
Umbilical Vein ◽  
Sirtuin 1 ◽  
No Production ◽  
Beneficial Effects ◽  
Umbilical Vein Endothelial Cells ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

Cilostazol was suggested to be beneficial to retard in-stent atherosclerosis and prevent stent thrombosis. However, the mechanisms responsible for the beneficial effects of cilostazol are not fully understood. In this study, we attempted to verify the mechanism of the antithrombotic effect of cilostazol. Human umbilical vein endothelial cells (HUVECs) were cultured with various concentrations of cilostazol to verify its impact on endothelial cells. KLF2, silent information regulator transcript-1 (SIRT1), endothelial nitric oxide synthase (eNOS), and endothelial thrombomodulin (TM) expression levels were examined. We found cilostazol significantly activated KLF2 expression and KLF2-related endothelial function, including eNOS activation, Nitric oxide (NO) production, and TM secretion. The activation was regulated by SIRT1, which was also stimulated by cilostazol. These findings suggest that cilostazol may be capable of an antithrombotic and vasculoprotective effect in endothelial cells.

Laminar Flow on Endothelial Cells Suppresses eNOS O-GlcNAcylation to Promote eNOS Activity

2021 ◽  
Author(s):  
Sarah Basehore ◽  
Samantha Bohlman ◽  
Callie Weber ◽  
Swathi Swaminathan ◽  
Yuji Zhang ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
Laminar Flow ◽  
No Production ◽  
Disturbed Flow ◽  
Enos Activity ◽  
Enos Phosphorylation ◽  
Steady Laminar ◽  
In Cells

Rationale: In diabetic animals as well as high glucose cell culture conditions, endothelial nitric oxide synthase (eNOS) is heavily O-GlcNAcylated, which inhibits its phosphorylation and nitric oxide (NO) production. It is unknown, however, whether varied blood flow conditions, which affect eNOS phosphorylation, modulate eNOS activity via O-GlcNAcylation-dependent mechanisms. Objective: The goal of this study was to test if steady laminar flow, but not oscillating disturbed flow, decreases eNOS O-GlcNAcylation, thereby elevating eNOS phosphorylation and NO production. Methods and Results: Human umbilical vein endothelial cells (HUVEC) were exposed to either laminar flow (20 dynes/cm2 shear stress) or oscillating disturbed flow (4{plus minus}6 dynes/cm2 shear stress) for 24 hours in a cone-and-plate device. eNOS O-GlcNAcylation was almost completely abolished in cells exposed to steady laminar but not oscillating disturbed flow. Interestingly, there was no change in protein level or activity of key O-GlcNAcylation enzymes (OGT, OGA, or GFAT). Instead, metabolomics data suggest that steady laminar flow decreases glycolysis and hexosamine biosynthetic pathway (HBP) activity, thereby reducing UDP-GlcNAc pool size and consequent O-GlcNAcylation. Inhibition of glycolysis via 2-deoxy-2-glucose (2-DG) in cells exposed to disturbed flow efficiently decreased eNOS O-GlcNAcylation, thereby increasing eNOS phosphorylation and NO production. Finally, we detected significantly higher O-GlcNAcylated proteins in endothelium of the inner aortic arch in mice, suggesting that disturbed flow increases protein O-GlcNAcylation in vivo. Conclusions: Our data demonstrate that steady laminar but not oscillating disturbed flow decreases eNOS O-GlcNAcylation by limiting glycolysis and UDP-GlcNAc substrate availability, thus enhancing eNOS phosphorylation and NO production. This research shows for the first time that O-GlcNAcylation is regulated by mechanical stimuli, relates flow-induced glycolytic reductions to macrovascular disease, and highlights targeting HBP metabolic enzymes in endothelial cells as a novel therapeutic strategy to restore eNOS activity and prevent EC dysfunction in cardiovascular disease.

scholarly journals (−)-Epicatechin induces calcium and translocation independent eNOS activation in arterial endothelial cells

2011 ◽  
Vol 300 (4) ◽  
pp. C880-C887 ◽  
Author(s):  
Israel Ramirez-Sanchez ◽  
Lisandro Maya ◽  
Guillermo Ceballos ◽  
Francisco Villarreal
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Cardiovascular Effects ◽  
Mesenteric Arteries ◽  
No Production ◽  
Human Coronary Artery ◽  
Calmodulin Binding ◽  
Pathway Activation ◽  
Functional Relevance ◽  
Endothelial Nitric Oxide

The consumption of cacao-derived (i.e., cocoa) products provides beneficial cardiovascular effects in healthy subjects as well as individuals with endothelial dysfunction such as smokers, diabetics, and postmenopausal women. The vascular actions of cocoa are related to enhanced nitric oxide (NO) production. These actions can be reproduced by the administration of the cacao flavanol (−)-epicatechin (EPI). To further understand the mechanisms behind the vascular action of EPI, we investigated the effects of Ca2+ depletion on endothelial nitric oxide (NO) synthase (eNOS) activation/phosphorylation and translocation. Human coronary artery endothelial cells were treated with EPI or with bradykinin (BK), a well-known Ca2+-dependent eNOS activator. Results demonstrate that both EPI and BK induce increases in intracellular calcium and NO levels. However, under Ca2+-free conditions, EPI (but not BK) is still capable of inducing NO production through eNOS phosphorylation at serine 615, 633, and 1177. Interestingly, EPI-induced translocation of eNOS from the plasmalemma was abolished upon Ca2+ depletion. Thus, under Ca2+-free conditions, EPI can stimulate NO synthesis independent of calmodulin binding to eNOS and of its translocation into the cytoplasm. We also examined the effect of EPI on the NO/cGMP/vasodilator-stimulated phosphoprotein (VASP) pathway activation in isolated Ca2+-deprived canine mesenteric arteries. Results demonstrate that under these conditions, EPI induces the activation of this vasorelaxation-related pathway and that this effect is inhibited by pretreatment with nitro-l-arginine methyl ester, suggesting a functional relevance for this phenomenon.

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