Cyclic ADP ribose-mediated Ca2+ signaling in mediating endothelial nitric oxide production in bovine coronary arteries

2006 ◽  
Vol 290 (3) ◽  
pp. H1172-H1181 ◽  
Author(s):  
Guo Zhang ◽  
Eric G. Teggatz ◽  
Andrew Y. Zhang ◽  
Matthew J. Koeberl ◽  
Fan Yi ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Coronary Arteries ◽  
Ryanodine Receptors ◽  
No Production ◽  
Signaling Mechanism ◽  
Adp Ribosyl Cyclase ◽  
Cyclic Adp Ribose ◽  
Intracellular Ca ◽  
A Cell ◽  
Endothelial Nitric Oxide

The present study tested the hypothesis that cyclic ADP ribose (cADPR) serves as a novel second messenger to mediate intracellular Ca2+ mobilization in coronary arterial endothelial cells (CAECs) and thereby contributes to endothelium-dependent vasodilation. In isolated and perfused small bovine coronary arteries, bradykinin (BK)-induced concentration-dependent vasodilation was significantly attenuated by 8-bromo-cADPR (a cell-permeable cADPR antagonist), ryanodine (an antagonist of ryanodine receptors), or nicotinamide (an ADP-ribosyl cyclase inhibitor). By in situ simultaneously fluorescent monitoring, Ca2+ transient and nitric oxide (NO) levels in the intact coronary arterial endothelium preparation, 8-bromo-cADPR (30 μM), ryanodine (50 μM), and nicotinamide (6 mM) substantially attenuated BK (1 μM)-induced increase in intracellular [Ca2+] by 78%, 80%, and 74%, respectively, whereas these compounds significantly blocked BK-induced NO increase by about 80%, and inositol 1,4,5-trisphosphate receptor blockade with 2-aminethoxydiphenyl borate (50 μM) only blunted BK-induced Ca2+-NO signaling by about 30%. With the use of cADPR-cycling assay, it was found that inhibition of ADP-ribosyl cyclase by nicotinamide substantially blocked BK-induced intracellular cADPR production. Furthermore, HPLC analysis showed that the conversion rate of β-nicotinamide guanine dinucleotide into cyclic GDP ribose dramatically increased by stimulation with BK, which was blockable by nicotinamide. However, U-73122, a phospholipase C inhibitor, had no effect on this BK-induced increase in ADP-ribosyl cyclase activity for cADPR production. In conclusion, these results suggest that cADPR importantly contributes to BK- and A-23187-induced NO production and vasodilator response in coronary arteries through its Ca2+ signaling mechanism in CAECs.

Simultaneous in situ monitoring of intracellular Ca2+ and NO in endothelium of coronary arteries

2002 ◽  
Vol 283 (6) ◽  
pp. H2725-H2732 ◽  
Author(s):  
Fu-Xian Yi ◽  
Andrew Y. Zhang ◽  
William B. Campbell ◽  
Ai-Ping Zou ◽  
Cornelis van Breemen ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Coronary Arteries ◽  
High Speed ◽  
Tyrosine Phosphatase ◽  
In Situ Monitoring ◽  
No Production ◽  
Slow Increase ◽  
Intracellular Ca ◽  
Endothelial Nitric Oxide

We developed an in situ assay system to simultaneously monitor intracellular Ca2+concentration ([Ca2+]i, fura 2 as indicator) and nitric oxide (NO) levels [4,5-diaminofluorescein as probe] in the intact endothelium of small bovine coronary arteries by using a fluorescent microscopic imaging technique with high-speed wavelength switching. Bradykinin (BK; 1 μM) stimulated a rapid increase in [Ca2+]i followed by an increase in NO production in the endothelial cells. The protein tyrosine phosphatase inhibitor phenylarsine oxide (PAO; 10 μM) induced a gradual, small increase in [Ca2+]i and a slow increase in intracellular NO levels. Removal of extracellular Ca2+ and depletion of Ca2+ stores completely blocked BK-induced increase in NO production but had no effect on PAO-induced NO production. However, a further reduction of [Ca2+]i by application of BAPTA-AM or EGTA with ionomycin abolished the PAO-induced NO increase. These results indicate that a simultaneous monitoring of [Ca2+]i and intracellular NO production in the intact endothelium is a powerful tool to study Ca2+-dependent regulation of endothelial nitric oxide synthase, which provides the first direct evidence for a permissive role of Ca2+ in tyrosine phosphorylation-induced NO production.

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.

Recombinant Gene Transfer of Endothelial Nitric Oxide Synthase Augments Coronary Artery Relaxations During Hypoxia

Circulation ◽  
1999 ◽  
Vol 100 (suppl_2) ◽  
Author(s):  
David G. Cable ◽  
Vincent J. Pompili ◽  
Timothy O’Brien ◽  
Hartzell V. Schaff
Keyword(s):  
Nitric Oxide ◽  
Coronary Artery ◽  
Nitric Oxide Synthase ◽  
Gene Transfer ◽  
Coronary Arteries ◽  
Endothelial Nitric Oxide Synthase ◽  
No Production ◽  
Viral Control ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

Background —Coronary arteries respond to hypoxia with transient relaxations, which increases coronary blood flow, in part, by release of nitric oxide. We hypothesized that increased expression of nitric oxide synthase might further augment blood vessel relaxation during hypoxia. The present study examined the effect of adenovirus-mediated transfer of bovine endothelial nitric oxide synthase (eNOS) on hypoxia-induced transient relaxations in canine coronary arteries. Methods and Results —Paired segments of coronary arteries were exposed to vehicle (phosphate-buffered saline with albumin) or an adenovirus encoding either E coli β-galactosidase (Ad.CMVLacZ, viral control; 10 10 pfu/mL) or eNOS (Ad.CMVeNOS; 10 10 pfu/mL) for 2 hours at 37°C. Immunohistochemistry with a monoclonal antibody specific for eNOS documented both endothelial and adventitial expression in Ad.CMVeNOS arteries, whereas vehicle and viral controls demonstrated only constitutive expression. Levels of cGMP were increased 5-fold in Ad.CMVeNOS arteries compared with controls. In arteries exposed to Ad.CMVeNOS, maximum contraction to prostaglandin F 2α was reduced compared with viral controls, and this effect was eliminated by pretreatment with a competitive inhibitor of eNOS ( N G -monomethyl- l -arginine, 10 −3 mol/L). Hypoxia-induced transient relaxation (95% N 2 -5% CO 2 ) in Ad.CMVeNOS arteries (45.2±8.8%, n=6) was augmented compared with vehicle (26.3±6.0%) or viral (27.2±7.1%) controls. Conclusions —Adenovirus-mediated gene transfer of nitric oxide synthase reduces receptor-dependent contractions and augments hypoxia-induced relaxations in canine coronary arteries; this method of augmentation of NO production might be advantageous for reduction of coronary artery vasospasm.

Impairment of endothelial nitric oxide production by acute glucose overload

2001 ◽  
Vol 280 (1) ◽  
pp. E171-E178 ◽  
Author(s):  
Chiwaka Kimura ◽  
Masahiro Oike ◽  
Tetsuya Koyama ◽  
Yushi Ito
Keyword(s):  
Nitric Oxide ◽  
No Production ◽  
Nitric Oxide Production ◽  
Superoxide Anions ◽  
Aortic Endothelial Cells ◽  
Bovine Aortic Endothelial Cells ◽  
A 23187 ◽  
Intracellular Ca ◽  
Endothelial Nitric Oxide ◽  
Cellular Productivity

We examined the effects of acute glucose overload (pretreatment for 3 h with 23 mM d-glucose) on the cellular productivity of nitric oxide (NO) in bovine aortic endothelial cells (BAEC). We had previously reported (Kimura C, Oike M, and Ito Y. Circ Res, 82: 677–685, 1998) that glucose overload impairs Ca2+ mobilization due to an accumulation of superoxide anions (O2 −) in BAEC. In control cells, ATP induced an increase in NO production, assessed by diaminofluorescein 2 (DAF-2), an NO-sensitive fluorescent dye, mainly due to Ca2+ entry. In contrast, ATP-induced increase in DAF-2 fluorescence was impaired by glucose overload, which was restored by superoxide dismutase, but not by catalase or deferoxamine. Furthermore, pyrogallol, an O2 − donor, also attenuated ATP-induced increase in DAF-2 fluorescence. In contrast, a nonspecific intracellular Ca2+ concentration increase induced by the Ca2+ ionophore A-23187, which depletes the intracellular store sites, elevated DAF-2 fluorescence in both control and highd-glucose-treated cells in Ca2+-free solution. These results indicate that glucose overload impairs NO production by the O2 −-mediated attenuation of Ca2+entry.

Nitric oxide inhibits ADP-ribosyl cyclase through a cGMP-independent pathway in airway smooth muscle

2002 ◽  
Vol 283 (5) ◽  
pp. L1065-L1071 ◽  
Author(s):  
Thomas A. White ◽  
Timothy F. Walseth ◽  
Mathur S. Kannan
Keyword(s):  
Nitric Oxide ◽  
Smooth Muscle ◽  
Airway Smooth Muscle ◽  
Hydrolase Activity ◽  
Adp Ribosyl Cyclase ◽  
Cyclic Adp Ribose ◽  
Protein Sulfhydryl ◽  
Intracellular Ca ◽  
Protein Sulfhydryl Groups

There is evidence for a role of cyclic ADP-ribose (cADPR) in intracellular Ca2+ regulation in smooth muscle. cADPR is synthesized and degraded by ADP-ribosyl cyclase and cADPR hydrolase, respectively, by a bifunctional protein, CD38. Nitric oxide (NO) inhibits intracellular Ca2+mobilization in airway smooth muscle. The present study was designed to determine whether this inhibition is due to regulation of ADP-ribosyl cyclase and/or cADPR hydrolase activity. Sodium nitroprusside (SNP) and S-nitroso- N-acetylpenicillamine, NO donors, produced a concentration-dependent decrease in ADP-ribosyl cyclase, but not cADPR hydrolase, activity. The NO scavenger carboxy-PTIO prevented and reversed, and reduced glutathione prevented, the inhibition of ADP-ribosyl cyclase by SNP, suggesting S-nitrosylation by NO as a mechanism. N-ethylmaleimide, which covalently modifies protein sulfhydryl groups, making them incapable of nitrosylation, produced a marked inhibition of ADP-ribosyl cyclase, but not cADPR hydrolase, activity. SNP and N-ethylmaleimide significantly inhibited the ADP-ribosyl cyclase activity in recombinant human CD38 without affecting the cADPR hydrolase activity. These results provide a novel mechanism for differential regulation of CD38 by NO through a cGMP-independent pathway involving S-nitrosylation of thiols.

Neuregulin-1 Compensates for Endothelial Nitric Oxide Synthase Deficiency

2021 ◽  
Author(s):  
Hadis Shakeri ◽  
Jente R.A. Boen ◽  
Sofie De Moudt ◽  
Jhana O. Hendrickx ◽  
Arthur J.A. Leloup ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Cardiac Fibrosis ◽  
Separate Experiment ◽  
No Production ◽  
Ang Ii ◽  
Wild Type ◽  
Paracrine Factor ◽  
Renal Hypertrophy ◽  
Neuregulin 1 ◽  
Endothelial Nitric Oxide

Endothelial cells (ECs) secrete different paracrine signals that modulate the function of adjacent cells; two examples of these paracrine signals are nitric oxide (NO) and neuregulin-1 (NRG1), a cardioprotective growth factor. Currently, it is undetermined whether one paracrine factor can compensate for the loss of another. Herein, we hypothesized that NRG1 can compensate for endothelial NO synthase (eNOS) deficiency. Methods. We characterized eNOS null and wild type (WT) mice by cardiac ultrasound and histology and we determined circulating NRG1 levels. In a separate experiment, 8 groups of mice were divided into 4 groups of eNOS null mice and wild type (WT) mice; half of the mice received angiotensin II (Ang II) to induce a more severe phenotype. Mice were randomized to daily injections with NRG1 or vehicle for 28 days. Results. eNOS deficiency increased NRG1 plasma levels, indicating that ECs increase their NRG1 expression when NO production is deleted. eNOS deficiency also increased blood pressure, lowered heart rate, induced cardiac fibrosis, and affected diastolic function. In eNOS null mice, Ang II administration increased cardiac fibrosis, but also induced cardiac hypertrophy and renal fibrosis. NRG1 administration prevented the cardiac and renal hypertrophy and fibrosis caused by Ang II infusion and eNOS deficiency. Moreover, Nrg1 expression in the myocardium is shown to be regulated by miR-134. Conclusion. This study indicates that administration of endothelium-derived NRG1 can compensate for eNOS deficiency in the heart and kidneys.

scholarly journals Protein kinase Cδ regulates endothelial nitric oxide synthase expression via Akt activation and nitric oxide generation

2008 ◽  
Vol 294 (3) ◽  
pp. L582-L591 ◽  
Author(s):  
Neetu Sud ◽  
Stephen Wedgwood ◽  
Stephen M. Black
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Endothelial Nitric Oxide Synthase ◽  
Promoter Activity ◽  
Dominant Negative ◽  
No Production ◽  
Enos Expression ◽  
Akt Activation ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

In this study, we explore the roles of the delta isoform of PKC (PKCδ) in the regulation of endothelial nitric oxide synthase (eNOS) activity in pulmonary arterial endothelial cells isolated from fetal lambs (FPAECs). Pharmacological inhibition of PKCδ with either rottlerin or with the peptide, δV1-1, acutely attenuated NO production, and this was associated with a decrease in phosphorylation of eNOS at Ser1177 (S1177). The chronic effects of PKCδ inhibition using either rottlerin or the overexpression of a dominant negative PKCδ mutant included the downregulation of eNOS gene expression that was manifested by a decrease in both eNOS promoter activity and protein expression after 24 h of treatment. We also found that PKCδ inhibition blunted Akt activation as observed by a reduction in phosphorylated Akt at position Ser473. Thus, we conclude that PKCδ is actively involved in the activation of Akt. To determine the effect of Akt on eNOS signaling, we overexpressed a dominant negative mutant of Akt and determined its effect of NO generation, eNOS expression, and phosphorylation of eNOS at S1177. Our results demonstrated that Akt inhibition was associated with decreased NO production that correlated with reduced phosphorylation of eNOS at S1177, and decreased eNOS promoter activity. We next evaluated the effect of endogenously produced NO on eNOS expression by incubating FPAECs with the eNOS inhibitor 2-ethyl-2-thiopseudourea (ETU). ETU significantly inhibited NO production, eNOS promoter activity, and eNOS protein levels. Together, our data indicate involvement of PKCδ-mediated Akt activation and NO generation in maintaining eNOS expression.

Endothelin stimulates endothelial nitric oxide synthase expression in the thick ascending limb

2004 ◽  
Vol 287 (2) ◽  
pp. F231-F235 ◽  
Author(s):  
Marcela Herrera ◽  
Jeffrey L. Garvin
Keyword(s):  
Nitric Oxide ◽  
Nitric Oxide Synthase ◽  
Receptor Antagonist ◽  
Endothelial Nitric Oxide Synthase ◽  
No Production ◽  
Thick Ascending Limb ◽  
Enos Expression ◽  
No Release ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

Endothelin-1 (ET-1) acutely inhibits NaCl reabsorption by the thick ascending limb (THAL) by activating the ETB receptor, stimulating endothelial nitric oxide synthase (eNOS), and releasing nitric oxide (NO). In nonrenal tissue, chronic exposure to ET-1 stimulates eNOS expression via the ETB receptor and activation of phosphatidylinositol 3-kinase (PI3K). We hypothesized that ET-1 increases eNOS expression in the THAL by binding to ETB receptors and stimulating PI3K. In primary cultures of medullary THALs treated for 24 h, eNOS expression increased by 36 ± 18% with 0.01 nM ET-1, 123 ± 30% with 0.1 nM ( P < 0.05; n = 5), and 71 ± 30% with 1 nM, whereas 10 nM had no effect. BQ-788, a selective ETB receptor antagonist, completely blocked stimulation of eNOS expression caused by 0.1 nM ET-1 (12 ± 25 vs. 120 ± 40% for ET-1 alone; P < 0.05; n = 5). BQ-123, a selective ETA receptor antagonist, did not affect the increase in eNOS caused by 0.1 nM ET-1. Sarafotoxin c (S6c; 0.1 μM), a selective ETB receptor agonist, increased eNOS expression by 77 ± 30% ( P < 0.05; n = 6). Wortmannin (0.01 μM), a PI3K inhibitor, completely blocked the stimulatory effect of 0.1 μM S6c (77 ± 30 vs. −28 ± 9%; P < 0.05; n = 6). To test whether the increase in eNOS expression heightens activity, we measured NO release in response to simultaneous treatment with l-arginine, ionomycin, and clonidine using a NO-sensitive electrode. NO release by control cells was 337 ± 61 and 690 ± 126 pA in ET-1-treated cells ( P < 0.05; n = 5). Taken together, these data suggest that ET-1 stimulates THAL eNOS, activating ETB receptors and PI3K and thereby increasing NO production.

scholarly journals Nitric oxide function in atherosclerosis

1997 ◽  
Vol 6 (1) ◽  
pp. 3-21 ◽  
Author(s):  
K. E. Matthys ◽  
H. Bult
Keyword(s):  
Nitric Oxide ◽  
Leukocyte Adhesion ◽  
Atherosclerotic Lesion ◽  
Vascular Wall ◽  
Early Event ◽  
No Production ◽  
Oxygen Intermediates ◽  
Atherosclerotic Lesions ◽  
Oxide Synthase ◽  
Endothelial Nitric Oxide

Atherosclerosis is a chronic inflammatory process in the intima of conduit arteries, which disturbs the endothelium-dependent regulation of the vascular tone by the labile liposoluble radical nitric oxide (NO) formed by the constitutive endothelial nitric oxide synthase (eNOS). This defect predisposes to coronary vasospasm and cardiac ischaemia, with anginal pain as the typical clinical manifestation. It is now appreciated that endothelial dysfunction is an early event in atherogenesis and that it may also involve the microcirculation, in which atherosclerotic lesions do not develop. On the other hand, the inflammatory environment in atherosclerotic plaques may result in the expression of the inducible NO synthase (iNOS) isozyme. Whether the dysfunction in endothelial NO production is causal to, or the result of, atherosclerotic lesion formation is still highly debated. Most evidence supports the hypothesis that constitutive endothelial NO release protects against atherogenesis e.g. by preventing smooth muscle cell proliferation and leukocyte adhesion. Nitric oxide generated by the inducible isozyme may be beneficial by replacing the failing endothelial production but excessive release may damage the vascular wall cells, especially in combination with reactive oxygen intermediates.

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.

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