scholarly journals Chronic exposure to high fatty acids impedes receptor agonist-induced nitric oxide production and increments of cytosolic Ca2+ levels in endothelial cells

2011 ◽  
Vol 47 (3) ◽  
pp. 315-326 ◽  
Author(s):  
Yanxia Tang ◽  
GuoDong Li
Keyword(s):  
Nitric Oxide ◽  
Fatty Acids ◽  
Endothelial Cells ◽  
Chronic Exposure ◽  
Receptor Agonist ◽  
No Production ◽  
Dependent Manner ◽  
Receptor Affinity ◽  
Receptor Agonists ◽  
High Concentrations

Dyslipidemia is a common metabolic disorder in diabetes. Nitric oxide (NO) production from endothelium plays the primary role in endothelium-mediated vascular relaxation and other endothelial functions. Therefore, we investigated the effects of elevated free fatty acids (FFA) on the stimulation of NO production by phospholipase C (PLC)-activating receptor agonists (potent physiological endothelium-dependent vasodilators) and defined the possible alterations of signaling pathways implicated in this scenario. Exposure of bovine aortic endothelial cells (BAECs) to high concentrations of a mixture of fatty acids (oleate and palmitate) for 5 or 10 days significantly reduced NO production evoked by receptor agonists (bradykinin or ATP) in a time- and dose-dependent manner. Such defects were not associated with alterations of either endothelial NO synthase mass or inositol phospholipid contents but were probably due to reduced elevations of intracellular free Ca2+levels ([Ca2+]i) under these conditions. Exposure of BAECs to FFA significantly attenuated agonist-induced [Ca2+]iincreases by up to 54% in a dose- and time-dependent manner. Moreover, bradykinin receptor affinity on the cell surface was significantly decreased by high concentrations of FFA. The morphology of BAECs was altered after 10-day culture with high FFA. Co-culture with protein kinase C (PKC) inhibitors or antioxidants was able to reverse the impairments of receptor agonist-induced NO production and [Ca2+]irises as well as the alteration of receptor affinity in BAECs exposed to FFA. These data indicate that chronic exposure to high FFA reduces NO generation in endothelial cells probably by impairing PLC-mediated Ca2+signaling pathway through activation of PKC and excess generation of oxidants.

Adenosine enhances nitric oxide production by vascular endothelial cells

1995 ◽  
Vol 269 (2) ◽  
pp. C519-C523 ◽  
Author(s):  
J. M. Li ◽  
R. A. Fenton ◽  
B. S. Cutler ◽  
J. G. Dobson
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Vascular Tone ◽  
Vascular Endothelial Cells ◽  
Competitive Inhibitor ◽  
No Production ◽  
Dependent Manner ◽  
Bathing Medium ◽  
Adenosine Receptor Antagonist ◽  
Potent Vasodilator

Adenosine per se is a potent vasodilator of vascular smooth muscle. Endothelial cells modulate vascular tone via the release of nitric oxide (NO), which also elicits vasodilation. This study was undertaken to determine whether adenosine could directly stimulate endothelial cells to enhance NO production, which could subsequently reduce vascular tone. NO production was evaluated in porcine carotid artery endothelial cells (PCAEC) and human saphenous vein endothelial cells (HSVEC) seeded on multiwell plates, grown to confluence, and treated with adenosine for 1 h. The bathing medium was collected, and the NO production was determined as reflected by the formation of NO2- and NO3-. NO production by PCAEC was significantly increased by adenosine in a dose-dependent manner, whereas there was only an insignificant tendency for an increase by HSVEC. The addition of the NO synthase competitive inhibitor, NG-monomethyl-L-arginine (NMMA), or the adenosine receptor antagonist, theophylline, prevented the increase in NO production by adenosine. The results suggest that adenosine stimulates, by a receptor-mediated mechanism, the production of NO by arterial, but not by venous, endothelial cells.

scholarly journals Androgen Receptor-Dependent Activation of Endothelial Nitric Oxide Synthase in Vascular Endothelial Cells: Role of Phosphatidylinositol 3-Kinase/Akt Pathway

Endocrinology ◽  
2010 ◽  
Vol 151 (4) ◽  
pp. 1822-1828 ◽  
Author(s):  
Jing Yu ◽  
Masahiro Akishita ◽  
Masato Eto ◽  
Sumito Ogawa ◽  
Bo-Kyung Son ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Androgen Receptor ◽  
Pi3 Kinase ◽  
Maximal Response ◽  
No Production ◽  
Dependent Manner ◽  
Small Interference Rna ◽  
Enos Phosphorylation ◽  
Interference Rna

The mechanisms of testosterone-induced vasodilatation are not fully understood. This study investigated the effect of testosterone on nitric oxide (NO) synthesis and its molecular mechanism using human aortic endothelial cells (HAEC). Testosterone at physiological concentrations (1–100 nm) induced a rapid (15–30 min) increase in NO production, which was associated with phosphorylation and activation of endothelial NO synthase (eNOS). Then, the involvement of the androgen receptor (AR), which is abundantly expressed in HAEC, was examined. The effect of testosterone on eNOS activation and NO production were abolished by pretreatment with an AR antagonist nilutamide and by transfection with AR small interference RNA. In contrast, testosterone-induced eNOS phosphorylation was unchanged by pretreatment with an aromatase inhibitor or by transfection with ERα small interference RNA. 5α-Dihydrotestosterone, a nonaromatizable androgen, also stimulated eNOS phosphorylation. Next, the signaling cascade that leads to eNOS phosphorylation was explored. Testosterone stimulated rapid phosphorylation of Akt in a time- and dose-dependent manner, with maximal response at 15–60 min. The rapid phosphorylation of eNOS or NO production induced by testosterone was inhibited by Akt inhibitor SH-5 or by phosphatidylinositol (PI) 3-kinase inhibitor wortmannin. Co-immunoprecipitation assays revealed a testosterone-dependent interaction between AR and the p85α subunit of PI3-kinase. In conclusion, testosterone rapidly induces NO production via AR-dependent activation of eNOS in HAEC. Activation of PI3-kinase/Akt signaling and the direct interaction of AR with p85α are involved, at least in part, in eNOS phosphorylation.

Kininase I-type carboxypeptidases enhance nitric oxide production in endothelial cells by generating bradykinin B1receptor agonists

2003 ◽  
Vol 284 (6) ◽  
pp. H1959-H1968 ◽  
Author(s):  
Sakonwun Sangsree ◽  
Viktor Brovkovych ◽  
Richard D. Minshall ◽  
Randal A. Skidgel
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Interferon Γ ◽  
Linear Increase ◽  
Interleukin 1Β ◽  
No Production ◽  
Fourfold Increase ◽  
Receptor Agonists ◽  
Inflammatory Conditions ◽  
Golgi Region

Kininase I-type carboxypeptidases convert native kinin agonists for B2receptors into B1receptor agonists by specifically removing the COOH-terminal Arg residue. The membrane localization of carboxypeptidase M (CPM) and carboxypeptidase D (CPD) make them ideally situated to regulate kinin activity. Nitric oxide (NO) release from human lung microvascular endothelial cells (HLMVEC) was measured directly in real time with a porphyrinic microsensor. Bradykinin (1–100 nM) elicited a transient (5 min) peak of generation of NO that was blocked by the B2antagonist HOE 140, whereas B1agonist des-Arg10-kallidin caused a small linear increase in NO over 20 min. Treatment of HLMVEC with 5 ng/ml interleukin-1β and 200 U/ml interferon-γ for 16 h upregulated B1receptors as shown by an approximately fourfold increase in prolonged (>20 min) output of NO in response to des-Arg10-kallidin, which was blocked by the B1antagonist des-Arg10-Leu9-kallidin. B2receptor agonists bradykinin or kallidin also generated prolonged NO production in treated HLMVEC, which was significantly reduced by either a B1antagonist or carboxypeptidase inhibitor, and completely abolished with a combination of B1and B2receptor antagonists. Furthermore, CPM and CPD activities were increased about twofold in membrane fractions of HLMVEC treated with interleukin-1β and interferon-γ compared with control cells. Immunostaining localized CPD primarily in a perinuclear/Golgi region, whereas CPM was on the cell membrane. These data show that cellular kininase I-type carboxypeptidases can enhance kinin signaling and NO production by converting B2agonists to B1agonists, especially in inflammatory conditions.

Role of G proteins in shear stress-mediated nitric oxide production by endothelial cells

1994 ◽  
Vol 267 (3) ◽  
pp. C753-C758 ◽  
Author(s):  
M. J. Kuchan ◽  
H. Jo ◽  
J. A. Frangos
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
G Protein ◽  
G Proteins ◽  
No Production ◽  
Dependent Manner ◽  
Protein Activation ◽  
Synthase Inhibitor

Exposure of cultured endothelial cells to shear stress resulting from well-defined fluid flow stimulates the production of nitric oxide (NO). We have established that an initial burst in production is followed by sustained steady-state NO production. The signal transduction events leading to this stimulation are not well understood. In the present study, we examined the role of regulatory guanine nucleotide binding proteins (G proteins) in shear stress-mediated NO production. In endothelial cells not exposed to shear stress, AIF4-, a general activator of G proteins, markedly elevated the production of guanosine 3',5'-cyclic monophosphate (cGMP). Pretreatment with NO synthase inhibitor N omega-nitro-L-arginine completely blocked this stimulation. Incubation with guanosine 5'-O-(2-thiodiphosphate) (GDP beta S), a general G protein inhibitor, blocked the flow-mediated burst in cGMP production in a dose-dependent manner. Likewise, GDP beta S inhibited NOx (NO2 + NO3) production for the 1st h. However, inhibition was not detectable between 1 and 3 h. Pertussis toxin (PTx) had no effect on the shear response at any time point. The burst in NO production caused by a change in shear stress appears to be dependent on a PTx-refractory G protein. Sustained shear-mediated production is independent of G protein activation.

Modulation of the l-arginine/nitric oxide signalling pathway in vascular endothelial cells

2004 ◽  
Vol 71 ◽  
pp. 143-156 ◽  
Author(s):  
Amanda W. Wyatt ◽  
Joern R. Steinert ◽  
Giovanni E. Mann
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Amino Acid ◽  
Protein Kinase ◽  
Signalling Pathway ◽  
No Production ◽  
Dependent Manner ◽  
Amino Acid Transport System ◽  
Membrane Hyperpolarization ◽  
Cationic Amino Acid

Nitric oxide (NO) is synthesized from l-arginine, and in endothelial cells influx of l-arginine is mediated predominantly via Na+-independent cationic amino acid transporters. Constitutive, Ca2+-calmodulin-sensitive eNOS (endothelial nitric oxide synthase) metabolizes l-arginine to NO and l-citrulline. eNOS is present in membrane caveolae and the cytosol and requires tetrahydrobiopterin, NADPH, FAD and FMN as additional cofactors for its activity. Supply of l-arginine for NO synthesis appears to be derived from a membrane-associated compartment distinct from the bulk intracellular amino acid pool, e.g. near invaginations of the plasma membrane referred to as 'lipid rafts' or caveolae. Co-localization of eNOS and the cationic amino acid transport system y+ in caveolae in part explains the 'arginine paradox', related to the phenomenon that in certain disease states eNOS requires an extracellular supply of l-arginine despite having sufficient intracellular l-arginine concentrations. Vasoactive agonists normally elevate [Ca2+]i (intracellular calcium concentration) in endothelial cells, thus stimulating NO production, whereas fluid shear stress, 17ϐ-oestradiol and insulin cause phosphorylation of the serine/threonine protein kinase Akt/protein kinase B in a phosphoinositide 3-kinase-dependent manner and activation of eNOS at basal [Ca2+]i levels. Adenosine causes an acute activation of p42/p44 mitogen-activated protein kinase and NO release, with membrane hyperpolarization leading to increased system y+ activity in fetal endothelial cells. In addition to acute stimulatory actions of D-glucose and insulin on l-arginine transport and NO synthesis, gestational diabetes, intrauterine growth retardation and pre-eclampsia induce phenotypic changes in the fetal vasculature, resulting in alterations in the l-arginine/NO signalling pathway and regulation of [Ca2+]i. These alterations may have significant implications for long-term programming of the fetal cardiovascular system.

scholarly journals Nitric Oxide is Produced by Cowdria ruminantium-Infected Bovine Pulmonary Endothelial Cells In Vitro and Is Stimulated by Gamma Interferon

1998 ◽  
Vol 66 (5) ◽  
pp. 2115-2121 ◽  
Author(s):  
Mbithe Mutunga ◽  
Patricia M. Preston ◽  
Keith J. Sumption
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
No Synthase ◽  
No Production ◽  
Dependent Manner ◽  
Donor Molecule ◽  
No Donor ◽  
Pulmonary Endothelial Cells ◽  
Cowdria Ruminantium ◽  
Dose Dependent

ABSTRACT Nitric oxide (NO) is a labile inorganic free radical produced by NO synthase from the substrate l-arginine in various cells and tissues including endothelial cells. A substantial elevation of nitrite levels indicative of NO production occurred in cultures ofCowdria ruminantium-infected bovine pulmonary endothelial cells (BPEC) incubated in medium alone. Exposure of the infected cultures to recombinant bovine gamma interferon (BorIFN-γ) resulted in more rapid production of NO, reduced viability of C. ruminantium, and induction of endothelial cell death. Significant inhibition of NO production was noted after addition of the NO synthase inhibitor N-monomethyl-l-arginine (l-NMMA), indicating that the increase in production occurred via the inducible NO synthase pathway. Reduction in the infectivity of C. ruminantium elementary bodies (EBs) occurred in a dose-dependent manner after incubation with the NO donor moleculeS-nitroso-N-acetyl-dl-penicillamine (SNAP) prior to infection of endothelial cells. The level of infection in cultures maintained in SNAP was reduced in a dose-dependent manner with significant negative correlation between the final level of infection on day 7 and the level of SNAP (r = −0.96). It was established that pretreatment and cultivation of C. ruminantium EBs with the NO donor molecule SNAP reduced infectivity to cultures and viability of EBs with the implication that release of NO in vivo following infection of endothelial cells may have an effect upon the multiplication of the agent in the host animal and may be involved in the pathogenesis of heartwater through the effect of this molecule upon circulation.

Role of cytoskeleton in shear stress-induced endothelial nitric oxide production

1997 ◽  
Vol 273 (1) ◽  
pp. H347-H355 ◽  
Author(s):  
H. L. Knudsen ◽  
J. A. Frangos
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
Plasma Membrane ◽  
Cytochalasin D ◽  
Induced Response ◽  
No Production ◽  
Dependent Manner ◽  
Mechanochemical Transduction

To study the role of the cytoskeleton in mechanochemical transduction, human umbilical vein endothelial cells were exposed to cytoskeleton-disrupting or -stabilizing agents, and the flow-induced production of nitric oxide (NO) as monitored by intracellular levels of guanosine 3',5'-cyclic monophosphate (cGMP) was examined. A shear stress of 20 dyn/cm2 elevated cGMP levels approximately twofold relative to basal (stationary) levels (1.9 +/- 0.1 pmol cGMP in stationary controls; P < 0.01). Treatment with 1 microM phalloidin and 0.5 microM cytochalasin D did not significantly affect the flow-induced response (1.77 +/- 0.23 and 2.89 +/- 0.18 pmol cGMP in stationary controls, respectively), whereas disruption of microtubules with 0.5 microM colchicine significantly elevated the response (3.64 +/- 0.18 pmol cGMP in stationary controls; P < 0.01). The NO synthase inhibitor NG-amino-L-arginine abrogated all flow-induced elevations of cGMP, indicating that increased cGMP levels were mediated by NO. Cytoskeletal disruption with 0.2 microM cytochalasin D or 0.5 microM colchicine did not alter cGMP levels in response to 10 nM bradykinin. The role of the plasma membrane in mechanochemical transduction was examined by treatment with cholesteryl hemisuccinate, which attenuated the flow-induced response in a dose-dependent manner. In conclusion, the pathways of flow- and bradykinin-mediated NO production in endothelial cells did not require actin filament turnover or intact actin or microtubule cytoskeletons, and cholesterol, possibly by stiffening the plasma membrane, attenuated the flow response.

Shear Stress Effect on the Production of Nitric Oxide in Cultured Rat Aorta Endothelial Cells

2002 ◽  
Author(s):  
Jianfeng Ye ◽  
Baoguo Chen ◽  
Lisa X. Xu
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Shear Stress ◽  
Protein Tyrosine Kinase ◽  
Vascular Endothelial Cells ◽  
Free Calcium ◽  
Enos Gene ◽  
No Production ◽  
Stress Effect ◽  
Dependent Manner

Atherosclerotic lesions tend to develop in regions where there are separations from unidirectional laminar blood flow, typically near branches, bifurcations, regions of arterial narrowing, and curvatures in the arteries (1, 2). Obviously, homodynamic forces play a key role in atherosclerosis. Studies also indicate that vascular endothelium function disturbance, especially impairment of endothelium dependent vasodilation, is involved (3). Shear stress affects endothelial cells in many ways, such as cytoskeletal rearrangement, decrease of intracellular pH, release of PGI2 and some growth factors (PDGF, FGF, ECGF, TGF-b, etc), activation of IP3 and mitogen-activated protein kinases, and the significant increase in the production of nitric oxide (1,2,4,5). As an important function factor of vascular endothelial cells, nitric oxide (NO) is closely related to the endothelial dysfunction and atherosclerosis (6). Endothelial derived nitric oxide involves in many events in the vasculature, including vasodilation, inhibition of platelet aggregation, adhesion molecule expression, and vascular smooth muscle proliferation, which are directly or indirectly related to atherosclerosis. Endothelial cells release NO more potently in response to increased shear stress than to agonists that raise intracellular free calcium concentration [Ca2+]i. Studies have indicated that NO production increases with a calcium/CaM dependent manner in the first few minutes after exposed to shear stress, followed by a sustained NO production that occurs more than 30min which is Ca2+ independent (7). The activation of eNOS by shear stress, which modulated by Ca/CaM, G protein, tyrosine kinase phosphorylation and eNOS gene expression, is responsible for the increase of NO production (8). However, the contribution of extracellular calcium to the production of NO is somewhat contradictory.

Activated pericyte attenuates endothelial functions: nitric oxide – cGMP rescues activated pericyte-associated endothelial dysfunctions

2007 ◽  
Vol 85 (6) ◽  
pp. 709-720 ◽  
Author(s):  
Syamantak Majumder ◽  
K. P. Tamilarasan ◽  
Gopi Krishna Kolluru ◽  
Ajit Muley ◽  
C. Madhavan Nair ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Dysfunction ◽  
Phosphodiesterase Inhibitor ◽  
Egg Yolk ◽  
No Production ◽  
Dependent Manner ◽  
Endothelium Dysfunction ◽  
Vascular Bed

Hepatic stellate cells are liver-specific pericytes and exist in close proximity with endothelial cells. The activation of liver pericytes is intrinsic to liver pathogenesis, and leads to endothelial dysfunction, including the low bioavailability of nitric oxide (NO). However, the role of nitric oxide in pericyte–endothelium cross-talk has not yet been elucidated. This work examines the cellular mechanism of action of NO in pericyte-mediated endothelial dysfunction. We used in vitro coculture and conditioned medium systems to study the effects of activated liver pericytes on endothelial function, and an egg yolk vascular bed model was used to study the effects of activated pericytes on angiogenesis. This study also demonstrates that activated pericytes attenuate the migration, proliferation, permeability, and NO production of endothelial cells. Our results demonstrate that activated pericytes restrict angiogenesis in egg yolk vascular bed models, and NO supplementation recovers 70% of the inhibition. Our results also demonstrate that supplementation with NO, sildenafil citrate (phosphodiesterase inhibitor), and 8-bromo-cGMP (cGMP analog) partially recovers activated-pericyte-mediated endothelium dysfunction. We conclude that NO–cGMP alleviates activated-pericyte-associated endothelial dysfunction, including angiogenesis, in a cGMP-dependent manner.

scholarly journals Bradykinin stimulation of nitric oxide production is not sufficient for gamma-globin induction

2014 ◽  
Vol 142 (3-4) ◽  
pp. 189-196 ◽  
Author(s):  
Vladan Cokic ◽  
Tijana Suboticki ◽  
Bojana Beleslin-Cokic ◽  
Milos Diklic ◽  
Pavle Milenkovic ◽  
...  
Keyword(s):  
Gene Expression ◽  
Nitric Oxide ◽  
Endothelial Cells ◽  
Endothelial Cell ◽  
Globin Gene ◽  
No Production ◽  
Dependent Manner ◽  
Erythroid Progenitors ◽  
Globin Gene Expression ◽  
Stimulation Of

Introduction. Hydroxycarbamide, used in therapy of hemoglobinopathies, enhances nitric oxide (NO) production both in primary human umbilical vein endothelial cells (HUVECs) and human bone marrow endothelial cell line (TrHBMEC). Moreover, NO increases ?-globin and fetal hemoglobin levels in human erythroid progenitors. Objective. In order to find out whether simple physiologic stimulation of NO production by components of hematopoietic microenvironment can increase ?-globin gene expression, the effects of NO-inducer bradykinin were examined in endothelial cells. Methods. The study was performed in co-cultures of human erythroid progenitors, TrHBMEC and HUVECs by ozone-based chemiluminescent determination of NO and real-time quantitative RT-PCR. Results. In accordance with previous reports, the endogenous factor bradykinin increased endothelial cell production of NO in a dose- and time-dependent manner (0.1-0.6 ?M up to 30 minutes). This induction of NO in HUVECs and TrHBMEC by bradykinin was blocked by competitive inhibitors of NO synthase (NOS), demonstrating NOS-dependence. It has been shown that bradykinin significantly reduced endothelial NOS (eNOS) mRNA level and eNOS/?-actin ratio in HUVEC (by twofold). In addition, bradykinin failed to increase ?-globin mRNA expression in erythroid progenitors only, as well as in co-culture studies of erythroid progenitors with TrHBMEC and HUVEC after 24 hours of treatment. Furthermore, bradykinin did not induce ?/? globin ratio in erythroid progenitors in co-cultures with HUVEC. Conclusion. Bradykinin mediated eNOS activation leads to short time and low NO production in endothelial cells, insufficient to induce ?-globin gene expression. These results emphasized the significance of elevated and extended NO production in augmentation of ?-globin gene expression.

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