A Comparison of Different Approaches to Quantify Nitric Oxide Release from NO-Releasing Materials in Relevant Biological Media

The development of solid materials that deliver nitric oxide (NO) are of interest for several therapeutic applications. Nevertheless, due to NO’s reactive nature, rapid diffusion and short half-life, reporting their NO delivery characteristics is rather complex. The full knowledge of this parameter is fundamental to discuss the therapeutic utility of these materials, and thus, the NO quantification strategy must be carefully considered according to the NO-releasing scaffold type, to the expected NO-releasing amounts and to the medium of quantification. In this work, we explore and discuss three different ways of quantifying the release of NO in different biological fluids: haemoglobin assay, Griess assay and NO electrochemical detection. For these measurements, different porous materials, namely zeolites and titanosilicates were used as models for NO-releasing platforms. The oxyhaemoglobin assay offers great sensitivity (nanomolar levels), but it is only possible to monitor the NO release while oxyhaemoglobin is not fully converted. On the other hand, Griess assay has low sensitivity in complex biological media, namely in blood, and interferences with media make NO measurements questionable. Nevertheless, this method can measure micromolar amounts of NO and may be useful for an initial screening for long-term release performance. The electrochemical sensor enabled real-time measurements in a variety of biological settings. However, measured NO is critically low in oxygenated and complex media, giving transient signals, which makes long-term quantification impossible. Despite the disadvantages of each method, the combination of all the results provided a more comprehensive NO release profile for these materials, which will help to determine which formulations are most promising for specific therapeutic applications. This study highlights the importance of using appropriate NO quantification tools to provide accurate reports.

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Difference in Endothelium-Derived Hyperpolarizing Factor–Mediated Hyperpolarization and Nitric Oxide Release Between Human Internal Mammary Artery and Saphenous Vein

Circulation ◽

10.1161/circ.102.suppl_3.iii-296 ◽

2000 ◽

Vol 102 (suppl_3) ◽

Author(s):

Zhi-Gang Liu ◽

Zhi-Dong Ge ◽

Guo-Wei He

Keyword(s):

Nitric Oxide ◽

Saphenous Vein ◽

Internal Mammary Artery ◽

Resting Membrane Potential ◽

Nitric Oxide Release ◽

No Release ◽

Basal Release ◽

Internal Mammary ◽

Sensitive Electrode

Background —The greater nitric oxide (NO) release that occurs in the internal mammary artery (IMA) when compared with the saphenous vein (SV) has been suggested by more endothelium-dependent relaxation in the IMA or measured by bioassay; however, no direct measurement of NO- or endothelium-derived hyperpolarizing factor (EDHF)–mediated hyperpolarization has been reported. The present study measured such hyperpolarization, as well as NO release, in these vessels. Methods and Results —IMA (n=46) and SV (n=61) segments taken from patients undergoing coronary surgery were studied in the organ chamber. Hyperpolarization (by intracellular glass microelectrode) and NO release (by NO-sensitive electrode) in response to acetylcholine and bradykinin, with and without incubation with N G -nitro- l -arginine, indomethacin, and oxyhemoglobin, were measured. The resting membrane potential of the smooth muscle cells from the IMA (58±0.8 mV; n=15) was higher than that in those from the SV (−62±0.9 mV; n=23; P =0.0001). The EDHF-mediated hyperpolarization induced by acetylcholine (10 −5 mol/L: −9.4±1.5 mV in IMA, n=10, versus −4.5±1.0 mV in SV, n=17; P <0.01) and bradykinin (10 −7 mol/L: −10.9±1.5 mV in IMA, n=8, versus −5.1±0.5 mV in SV, n=8; P <0.01) and the basal release of NO (16.8±1.6 nmol/L in IMA, n=13, versus 9.9±2.8 nmol/L in SV, n=13; P <0.001) were significantly greater in the IMA than in the SV. The duration of acetylcholine- and bradykinin-induced NO release was longer in the IMA than in the SV. Conclusions —The basal release of NO and EDHF-mediated hyperpolarization were significantly greater in the IMA than in the SV. In addition, the duration of the stimulated release of NO was longer in the IMA than in the SV. These differences may contribute to the superior long-term patency of IMA grafts.

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Retracted Article: Light-triggered nitric oxide release and targeted fluorescence imaging in tumor cells developed from folic acid-graft-carboxymethyl chitosan nanospheres

RSC Advances ◽

10.1039/c4ra03034f ◽

2014 ◽

Vol 4 (57) ◽

pp. 30129-30136 ◽

Cited By ~ 18

Author(s):

Rijun Gui ◽

Ajun Wan ◽

Yalei Zhang ◽

Huili Li ◽

Tingting Zhao

Keyword(s):

Nitric Oxide ◽

Folic Acid ◽

Tumor Cells ◽

Fluorescence Imaging ◽

Carboxymethyl Chitosan ◽

Nitric Oxide Release ◽

No Release ◽

Retracted Article ◽

Potential Applications

This article reported the synthesis of CMC–FA–RBS(CQD) nanospheres and studied their potential applications for NO release and fluorescence imaging.

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Nitric oxide release by N-(2-chloroethyl)-N-nitrosoureas: a rarely discussed mechanistic path towards their anticancer activity

RSC Advances ◽

10.1039/c4ra11137k ◽

2015 ◽

Vol 5 (3) ◽

pp. 2137-2146

Author(s):

Amrita Sarkar ◽

Subhendu Karmakar ◽

Sudipta Bhattacharyya ◽

Kallol Purkait ◽

Arindam Mukherjee

Keyword(s):

Nitric Oxide ◽

Anticancer Activity ◽

Nitric Oxide Release ◽

No Release

Our work shows that NO release is a feasible pathway of action for aromatic and heterocyclic N-(2-chloroethyl)-N-nitrosoureas and faster NO release may not lead to higher cytotoxicity.

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Long-term administration of L-arginine improves nitric oxide release from kidney in deoxycorticosterone acetate-salt hypertensive rats.

Hypertension ◽

10.1161/01.hyp.23.6.752 ◽

1994 ◽

Vol 23 (6_pt_1) ◽

pp. 752-756 ◽

Cited By ~ 44

Author(s):

H Hayakawa ◽

Y Hirata ◽

E Suzuki ◽

K Kimura ◽

K Kikuchi ◽

...

Keyword(s):

Nitric Oxide ◽

Hypertensive Rats ◽

Deoxycorticosterone Acetate ◽

Nitric Oxide Release ◽

Acetate Salt ◽

Term Administration

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Asenapine modulates nitric oxide release and calcium movements in cardyomyoblasts

European Psychiatry ◽

10.1016/j.eurpsy.2016.01.2041 ◽

2016 ◽

Vol 33 (S1) ◽

pp. S553-S553

Author(s):

P. Zeppegno ◽

C. Gramaglia ◽

E. Gattoni ◽

S. Gili ◽

E. Gambaro ◽

...

Keyword(s):

Nitric Oxide ◽

Hydrogen Peroxide ◽

Clinical Relevance ◽

Nitric Oxide Release ◽

No Release ◽

Intracellular Ca ◽

Specific Receptors ◽

Competing Interest ◽

Dopaminergic Antagonists ◽

Serotoninergic Receptors

ObjectiveTo examine the effects of asenapine on NO release and Ca2+ transients in H9C2, which were either subjected to peroxidation or not.Materials and methodsH9C2 were treated with asenapine alone or in presence of intracellular kinases blockers, serotoninergic and dopaminergic antagonists, and voltage Ca2+ channels inhibitors. Experiments were also performed in H9C2 treated with hydrogen peroxide. NO release and intracellular Ca2+ were measured through specific probes.ResultsIn H9C2, asenapine differently modulated NO release and Ca2+ movements depending on the peroxidative condition. The Ca2+ pool mobilized by asenapine mainly originated from the extracellular space and was slightly affected by thapsigargin. Moreover, the effects of asenapine were reduced or prevented by kinases blockers, dopaminergic and serotoninergic receptors inhibitors and voltage Ca2+ channels blockers.ConclusionsOn the basis of our findings we can conclude that asenapine by interacting with its specific receptors, exerts dual effects on NO release and Ca2+ homeostasis in H9C2; this would be of particular clinical relevance, when considering their role in cardiac function modulation.Disclosure of interestThe authors have not supplied their declaration of competing interest.

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Direct measurement of nitric oxide release from vascular endothelial cells

Journal of Applied Physiology ◽

10.1152/jappl.1996.81.2.774 ◽

1996 ◽

Vol 81 (2) ◽

pp. 774-779 ◽

Cited By ~ 57

Author(s):

J. P. Guo ◽

T. Murohara ◽

M. Buerke ◽

R. Scalia ◽

A. M. Lefer

Keyword(s):

Nitric Oxide ◽

Endothelial Cells ◽

Methyl Ester ◽

Vascular Endothelial Cells ◽

Calmodulin Antagonist ◽

Nitric Oxide Release ◽

A 23187 ◽

No Release ◽

Basal Release ◽

Isolated Rat

A nitric oxide (NO)-selective electrode was used to directly measure NO release from isolated rat aortic endothelium and cultured rat aortic endothelial cells (RAECs). Basal release of NO was significantly attenuated by a NO synthase inhibitor NG-nitro-L-arginine methyl ester (1 mM) to 42 +/- 14 pmol/1 x 10(5) cells (P < 0.01). The basal release of NO was also significantly inhibited by a calmodulin antagonist W-7 at 15 microM (P < 0.01). L-Arginine (1 mM), significantly stimulated NO release (P < 0.05 vs. control basal release). Stimulation of cultured RAECs with two endothelium-dependent vasodilators, acetylcholine (100 nM) and A-23187 (1 microM), significantly increased NO release [574 +/- 112 pmol/1 x 10(5) cells (n = 5) and 658 +/- 119 pmol/1 x 10(5) cells (n = 5) in acetylcholine- and A-23187-stimulated RAECs, respectively]. Basal release of NO was also detectable in isolated rat aortic rings with intact endothelium. NO release was significantly attenuated by NG-nitro-L-arginine methyl ester and augmented by human superoxide dismutase. These data indicate the physiological usefulness of the amperometric measurement of NO employing a NO-specific electrode in biological systems.

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Functionalized graphene-based biomimetic microsensor interfacing with living cells to sensitively monitor nitric oxide release

Chemical Science ◽

10.1039/c4sc03123g ◽

2015 ◽

Vol 6 (3) ◽

pp. 1853-1858 ◽

Cited By ~ 43

Author(s):

Yan-Ling Liu ◽

Xue-Ying Wang ◽

Jia-Quan Xu ◽

Chong Xiao ◽

Yan-Hong Liu ◽

...

Keyword(s):

Nitric Oxide ◽

Real Time ◽

Living Cells ◽

Functionalized Graphene ◽

Nitric Oxide Release ◽

No Release

We present a biomimetic and reusable microsensor with sub-nanomolar sensitivity by elaboratly functionalizing graphene for monitoring NO release in real-time.

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cAMP-Dependent Long-Term Potentiation of Nitric Oxide Release from Cerebellar Parallel Fibers in Rats

Journal of Neuroscience ◽

10.1523/jneurosci.18-21-08551.1998 ◽

1998 ◽

Vol 18 (21) ◽

pp. 8551-8558 ◽

Cited By ~ 40

Author(s):

Shinji Kimura ◽

Seiji Uchiyama ◽

Hideaki E. Takahashi ◽

Katsuei Shibuki

Keyword(s):

Nitric Oxide ◽

Long Term Potentiation ◽

Parallel Fibers ◽

Nitric Oxide Release ◽

Term Potentiation

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Reduced perivascular Po2 increases nitric oxide release from endothelial cells

AJP Heart and Circulatory Physiology ◽

10.1152/ajpheart.00759.2002 ◽

2003 ◽

Vol 285 (2) ◽

pp. H507-H515 ◽

Cited By ~ 40

Author(s):

G. P. Nase ◽

J. Tuttle ◽

H. G. Bohlen

Keyword(s):

Nitric Oxide ◽

Endothelial Cells ◽

Parenchymal Cell ◽

Oxygen Availability ◽

No Production ◽

Main Site ◽

Nitric Oxide Release ◽

No Release ◽

Parenchymal Cells

Many studies have suggested that endothelial cells can act as “oxygen sensors” to large reductions in oxygen availability by increasing nitric oxide (NO) production. This study determined whether small reductions in oxygen availability enhanced NO production from in vivo intestinal arterioles, venules, and parenchymal cells. In vivo measurements of perivascular NO concentration ([NO]) were made with NO-sensitive microelectrodes during normoxic and reduced oxygen availability. During normoxia, intestinal first-order arteriolar [NO] was 397 ± 26 nM ( n = 5), paired venular [NO] was 298 ± 34 nM ( n = 5), and parenchymal cell [NO] was 138 ± 36 nM ( n = 3). During reduced oxygen availability, arteriolar and venular [NO] significantly increased to 695 ± 79 nM ( n = 5) and 534 ± 66 nM ( n = 5), respectively, whereas parenchymal [NO] remained unchanged at 144 ± 34 nM ( n = 4). During reduced oxygenation, arteriolar and venular diameters increased by 15 ± 3% and 14 ± 5%, respectively: NG-nitro-l-arginine methyl ester strongly suppressed the dilation to lower periarteriolar Po2. Micropipette injection of a CO2 embolus into arterioles significantly attenuated arteriolar dilation and suppressed NO release in response to reduced oxygen availability. These results indicated that in rat intestine, reduced oxygen availability increased both arteriolar and venular NO and that the main site of NO release under these conditions was from endothelial cells.

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