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Tonal nitric oxide and health - A free radical and a scavenger of free radicals

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Abstract

Basal/tonal nitric oxide (NO) production helps maintain particular microenvironments, i.e., vascular. Besides NO's function in controlling the activation state of various tissues such as immune cells, its presence appears to modulate other free radical levels, i.e., H2O2, in these same tissues and indeed these processes may be one and the same. Thus, by being a free radical, along with the ability to scavenge other free radicals, NO is placed in a pivotal regulatory position. We surmise that in the absence of adequate NO release other free radicals may go 'unchecked' and, therefore, initiate tissue damage. Furthermore, under these circumstances, proinflammatory events will occur due to heightened cell sensitivity and a diminished control of NF-kappaB. In an excess situation, and one without an appropriate circumstance, i.e., microbial action, NO may become the harmful agent. Hence, balancing basal NO production in body compartments may represent a fundamental process in maintaining general, long-term health.
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Tonal nitric oxide and health – a free radical
and a scavenger of free radicals
Danielle Benz, Patrick Cadet, Kirk Mantione, Wei Zhu, George B. Stefano
Neuroscience Research Institute, State University of New York at Old Westbury, Old Westbury, New York, USA
Source of support: This work was in part supported by NIDA 09010 and the Cell Dynamics, Corp.
Summary:
Basal/tonal nitric oxide (NO) production helps maintain particular microenvironments, i.e.,
vascular. Besides NO’s function in controlling the activation state of various tissues such as
immune cells, its presence appears to modulate other free radical levels, i.e., H
2
O
2
, in these
same tissues and indeed these processes may be one and the same. Thus, by being a free radi-
cal, along with the ability to scavenge other free radicals, NO is placed in a pivotal regulatory
position. We surmise that in the absence of adequate NO release other free radicals may go
‘unchecked’ and, therefore, initiate tissue damage. Furthermore, under these circumstances,
proinflammatory events will occur due to heightened cell sensitivity and a diminished control
of NF-κB. In an excess situation, and one without an appropriate circumstance, i.e., microbial
action, NO may become the harmful agent. Hence, balancing basal NO production in body
compartments may represent a fundamental process in maintaining general, long-term
health.
key words:
nitric oxide • free radical • NF-kB • basal nitric oxide • tonal nitric oxide • antioxidant •
immunocytes • vascular endothelial cells
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Review Article
Signature: Med Sci Monit, 2002; 8(1): RA1
PMID: XXXXXXXX
RA1-2
Received: 2001.12.15
Accepted: 2001.12.21
Published: XXXX
Author’s address:
George B. Stefano, Neuroscience Research Institute, State University of New York at Old Westbury, Old Westbury,
New York 11568, USA, e-mail: gstefano@optonline.net
Authors’ Contribution:
A
Study Design
B
Data Collection
C
Statistical Analysis
D
Data Interpretation
E
Manuscript Preparation
F
Literature Search
G
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Med Sci Monit, 2002; 8(1): CR1
Benz D et al – Tonal nitric oxide and health: a free radical and a scavenger…
NITRIC OXIDE
Nitric oxide (NO) signaling occurs in diverse systems
including the immune, cardiovascular and nervous
[1–6]. It also occurs in evolutionarily diverse organisms
[7,8]. NO is produced from L-arginine by the enzyme
NO synthase (NOS) [4,9], which occurs in three forms:
endothelial (e), neuronal (n) and inducible (i) NOS.
NO derived from e- or -n (constitutively [c] expressed
forms) cNOS may occur in two functional forms: the
first is always present at low ‘tonal’ or ‘basal’ levels
which can be increased slightly for a short time in
response to various biological signals [5] such as acetyl-
choline (ACH). This brief enhanced release of cNOS
derived NO can have profound physiological actions
that are evident long after NO levels have returned to
the basal level of production [10]. In this regard,
immune and vascular endothelial cells can be down reg-
ulated by NO see [7]. We have hypothesized that certain
classes of cells are always activated and thus can respond
to immediate microenvironmental changes [7]. We fur-
ther speculate that basal or tonal NO levels may provide
a major pathway to dampen these cells sensitivity to
microenvironmental ‘noise’ that would otherwise non-
specifically and inappropriately lead to increased activa-
tion [7]. In this regard, NO may modulate the threshold
required for activation of these cells [7] and the magni-
tude of the subsequent response [11]. Diminished NO
levels would then represent a disinhibition process that
results in an overcoming of the inhibitory influence by
changing the level of NO production and the corre-
sponding levels of excitatory signals required for cellu-
lar activation (see [7]).
NO AND NF-KB
As an example of tonal NO’s significance, the transcrip-
tion factor NF-kB plays a pivotal role in regulation of
gene expression induced by inflammatory mediators
such as cytokines and adhesion molecules [12]. NO has
been associated with NF-kB inactivation [13-17]. Our
previous reports document the effect of NO on NF-kB
[7,18–20]. NF-kB mediated transcriptional activation of
many proinflammatory signal molecules/genes is inhibit-
ed by NO in a variety of cells including monocytes
[7,18–23]. Thus, NO, aside from its cGMP influences,
profoundly impacts DNA events leading to proinflam-
matory events.
NO AS AN ANTIOXIDANT FREE RADICAL
The intensity and diversity of current research regard-
ing NO demonstrate the complexity of the interactions
of this simple molecule. NO, a free radical, has actually
been shown to be a beneficial antioxidant against reac-
tive oxygen species (ROS), such as H
2
O
2
and O
2
–
,
[24,25]. When L-arginine, a precursor of NO, was
administered to rats in which experimental allergic
encephalitis (EAE) was induced, increased levels of NO
were shown to be correlated to a decrease in superoxide
and hydrogen peroxide levels, demonstrating a role as a
protective molecule [26]. There is also evidence of a role
in the production of ROS. Varying rates of endogenous
NO production resulted in a reciprocal correlation of
released H
2
O
2
in rat liver mitochondria. This seems to
be due to a regulation of O
2
consumption at the level of
the cytochrome oxidase [27]. It has been found that the
antioxidant properties of NO can be greatly increased
by the activation of specific pathways leading to
increased endogenous antioxidant production or down
regulated pro-inflammatory responses [28].
Much research has also been done to examine NO’s role
in decreasing lipid peroxidation [24,29–31], but this
process appears to be determined by the relative con-
centration of NO as compared to the reactive oxidant
species. When it is in excess of the ROS, then lipid oxi-
dation is decreased; but when NO levels drop below
that of the ROS, lipid oxidation reactions propagate
[32]. Nitric oxide has also been demonstrated to protect
cells from tert-butyl hydroperoxide (tBOOH), a com-
pound of lipid peroxidation. The generation of
tBOOH-derived free radicals and tBOOH-induced
cytotoxicity were both attenuated by endogenously pro-
duced or exogenously added NO [33-35]. In human
erythroleukemia cells, t-BuOOH-induced oxoferryl and
t-BuO alkoxyl radicals were chemically reduced by NO
[34].
From the earlier discussion it becomes clear that the
basal level of NO, derived from cNOS, may serve as the
key modulator regulating a complex cascading process
associated with maintaining cell health [7,36]. It
becomes important to determine how a particular
microenvironment may alter basal NO levels because, in
turn, we learn how NO functions, varied by circum-
stance. NO has the potential to interact with oxygen,
metals and other free radicals [37]. NO can also form
peroxynitrite (ONOO
–
) and dinitrogen trioxide (N
2
O
3
),
following an interaction with the superoxide radical (O
2
–
)
and oxygen, respectively [38]. In this regard, NO’s
direct effect is felt when its level is low and of short
duration, such as that occurring under physiological
conditions, including the right PH [38]. For example,
NO interaction with the heme proteins represents the
activation of soluble guanylyl cyclase (sGC) and/or
cyclooxygenase (COX) [39-41]. This last interaction is
important in the regulation of a proinflammatory
process [41]. At low concentrations (e.g., when it is scav-
enged), NO modulates the redox form of COX, con-
verting the ferrous iron to the ferric active form, acting
also as a scavenger of superoxide [38]. In addition, NO
has the ability to inhibit lipoxygenase, as noted earlier
[42]. It can reversibly inhibit the heme moiety of
cytochrome P-450, preventing the binding of oxygen to
the catalytic sites [43,44].
Interestingly, at low NO levels, H
2
O
2
can be consumed
to yield HNO
2
[38,45], suggesting that H
2
O
2
might
serve to control NO levels [38]. Indeed, the activation of
monocytes, with interferon ς for 24hr, results in the
appearance of activated ameboid monocytes as opposed
to inactive cells despite the production of high levels of
NO. Cell activation is abrogated in the presence of cata-
lase or superoxide dismutase, suggesting that H
2
O
2
RA1-4
Med Sci Monit, 2002; 8(1): CR1Review Article
inhibition of NO suppression represents an important
regulator of cellular activation [46]. Thus, in the
absence of H
2
O
2
, NO activity may be unregulated
whereas in the absence of NO, H
2
O
2
may generate tis-
sue damage and disruption in energy metabolism as evi-
dent in Alzheimer’s Disease [7,36]. In any case, the
basal/tonal level of NO may represent a specific signal to
maintaining ‘cell’ health.
Mitochondria represent a NO target due to the fact that
NO is an inhibitor of cytochrome oxidase of the electron
transport process [47-52], which suggests a NO role in
modulating oxygen utilization [47]. The inhibition of
cNOS-derived NO increases oxygen consumption in
many animal species [53-57]. This last fact is critical to
our NO hypothesis concerning alterations in basal NO
levels since its regulatory process may be stopped (see
earlier discussion). Furthermore, a NOS isoform,
mtNOS, is present in mitochondria [48,58], suggesting
an important modulatory function as well.
Heme proteins (e.g, hemoglobin, cytochromes, etc.)
reacting with H
2
O
2
results in ferryl cation (FE
4+
=O), a
toxic substance [59]. However, once in contact with NO,
this compound is reduced (FE
3+
+ NO
2
–
) [38], demon-
strating again a NO antioxidant action. NO also has the
potential to diminish the formation of OH
•
, again,
demonstrating an antioxidant action [60]. This scaveng-
ing property gives NO a major intracellular and extra-
cellular action against oxidative stress [38,61–67]. Here,
we note that in the absence of NO, these reactive chemi-
cal species may cause tissue damage associated with a
pathological progression.
In summary, it would appear that basal/tonal NO pro-
duction helps maintain particular microenvironments,
i.e., vascular see [7,68,69]. Besides NO’s function in con-
trolling the activation state of various tissues, such as
immune cells, its presence may also control free radical
levels in these same tissues and indeed these processes
may be one. Hence, by being a free radical, along with
the ability to scavenge other free radicals, NO is placed
in a pivotal regulatory position. We surmise that in the
absence of adequate NO release, other free radicals may
go ‘unchecked’ and, therefore, initiate tissue damage
see [68,69]. Furthermore, under these circumstances,
proinflammatory events will occur due to heightened
cell sensitivity and a diminished control of NF-kB. In an
excess situation and one without an appropriate circum-
stance, i.e., antimicrobial action, NO, by itself, may
become the harmful agent (see above). Thus, balancing
and maintaining basal NO production, exerting a tonal
effect, in microenvironments may represent a funda-
mental process in maintaining general health over the
long term.
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  • Jul 2021
The World Health Organization declared the SARS-CoV-2 outbreak a Public Health Emergency of International Concern at the end of January 2020 and a pandemic two months later. The virus primarily spreads between humans via respiratory droplets, and is the causative agent of Coronavirus Disease 2019 (COVID-19), which can vary in severity, from asymptomatic or mild disease (the vast majority of the cases) to respiratory failure, multi-organ failure, and death. Recently, several vaccines were approved for emergency use against SARS-CoV-2. However, their worldwide availability is acutely limited, and therefore, SARS-CoV-2 is still expected to cause significant morbidity and mortality in the upcoming year. Hence, additional countermeasures are needed, particularly pharmaceutical drugs that are widely accessible, safe, scalable, and affordable. In this comprehensive review, we target the prophylactic arena, focusing on small-molecule candidates. In order to consolidate a potential list of such medications, which were categorized as either antivirals, repurposed drugs, or miscellaneous, a thorough screening for relevant clinical trials was conducted. A brief molecular and/or clinical background is provided for each potential drug, rationalizing its prophylactic use as an antiviral or inflammatory modulator. Drug safety profiles are discussed, and current medical indications and research status regarding their relevance to COVID-19 are shortly reviewed. In the near future, a significant body of information regarding the effectiveness of drugs being clinically studied for COVID-19 is expected to accumulate, in addition to information regarding the efficacy of prophylactic treatments.
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... Several recent studies have revealed that anticancer activities of flavonoids may be mediated through pro-oxidant action [61,62] Cancer cells exhibit higher and more persistent oxidative stress and are more susceptible to being killed by drugs that boost increased ROS levels, such as some flavonoids [63][64][65][66] Whether a flavonoid acts as anti-or pro-oxidant depends on its dose, cell type and also culture conditions [65,67,68] Nitric oxide (NO) is an integral part of ROS produced by nitric oxide synthases [69][70][71]. Excessive ROS may cause irreversible damage to DNA, leading to cell death [72,73] ROS stimulates autophagy by regulation of ATG4 and stress signaling pathways [74]. Exogenous NO induces autophagy [75,76]. ...
Açai (Euterpe oleracea Mart.) Seed Extract Induces ROS Production and Cell Death in MCF-7 Breast Cancer Cell Line
Article
Full-text available
Abstract: Euterpe oleracea Mart. (açai) is a native palm from the Amazon region. There are various chemical constituents of açai with bioactive properties. This study aimed to evaluate the chemical composition and cytotoxic effects of açai seed extract on breast cancer cell line (MCF-7). Global Natural Products Social Molecular Networking (GNPS) was applied to identify chemical compounds present in açai seed extract. LC-MS/MS and molecular networking were employed to detect the phenolic compounds of açai. The antioxidant activity of açai seed extract was measured by DPPH assay. MCF-7 breast cancer cell line viability was evaluated by MTT assay. Cell death was evaluated by flow cytometry and time-lapse microscopy. Autophagy was evaluated by orange acridin immunofluorescence assay. Reactive oxygen species (ROS) production was evaluated by DAF assay. From the molecular networking, fifteen compounds were identified, mainly phenolic compounds. The açai seed extract showed cytotoxic effects against MCF-7, induced morphologic changes in the cell line by autophagy and increased the ROS production pathway. The present study suggests that açai seed extract has a high cytotoxic capacity and may induce autophagy by increasing ROS production in breast cancer. Apart from its antioxidant activity, flavonoids with high radical scavenging activity present in açai also generated NO (nitric oxide), contributing to its cytotoxic effect and autophagy induction.
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... Uncapacitated spermatozoa produce low levels of NO, whereas under capacitation conditions, a time-dependent increase in NO synthesis has been observed (Belen Herrero et al., 2000), so the effect of arginine as an antioxidant may delay the aging process. Benz et al. (2002) reported that since nitric oxide being a free radical along with the ability to scavenge other free radicals, it is placed in a Pivotal regulatory position. They speculated that in the absence of adequate nitric oxide release, other free radicals may initiate cells damage. ...
Effect of adding arginine in different concentrations on some physical properties of poor motile bull sperms during different months
Article
Full-text available
  • Dec 2012
In order to investigate the effect of adding arginine on poor motile bull sperms, this study was conducted in Artificial Insemination Center, Iraq, from December, 2008 to July, 2009. 114 ejaculates with poor motile sperms estimated (30 to 55%) were collected by artificial vagina (AV) from 10 Holstein bulls and extended with Tris-yolk-fructose extender supplemented with different concentrations of arginine (0, 0.002, 0.003, 0.004, 0.005 and 0.006 M/ml) for the determination of spermatozoa motility, dead and abnormalities percentages, and acrosoma abnormalities percentage. Results showed that adding different concentrations of arginine to poor motile bull sperms extended with Tris-yolk-fructose extender gradually increased the motility of spermatozoa but the best concentrations was 0.005 and 0.006 M/ml with significant (P < 0.05) differences when compared with the control during all month of this study, and months December and January gave better significant (P < 0.05) quality of semen compared with April, May, June and July, and last two months (June and July) gave lower significant (P < 0.05) different motility and high significant (P < 0.05) different percentage of dead, abnormality and abnormal acrosoma when compared with other months.
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... By acting as an electron donor or electron acceptor, this molecule can react with proteins, modifying their enzymatic and transcriptional activities [22,23]. Under oxidative stress conditions, NO can function as an antioxidant against reactive oxygen species (ROS), such as hydrogen peroxide (H 2 O 2 ) and the superoxide anion (O 2 − ), creating reactive nitrogen species (RNS); these molecules are responsible for DNA nitrosation, nitration and deamination, which lead to DNA instability [24,25]. ...
Lopinavir-NO, a nitric oxide-releasing HIV protease inhibitor, suppresses the growth of melanoma cells in vitro and in vivo
Article
Full-text available
We generated a nitric oxide (NO)-releasing derivative of the anti-HIV protease inhibitor lopinavir by linking the NO moiety to the parental drug. We investigated the effects of lopinavir and its derivative lopinavir-NO on melanoma cell lines in vitro and in vivo. Lopinavir-NO exhibited a twofold stronger anticancer action than lopinavir in vitro. These results were successfully translated into syngeneic models of melanoma in vivo, where a significant reduction in tumour volume was observed only in animals treated with lopinavir-NO. Both lopinavir and lopinavir-NO inhibited cell proliferation and induced the trans-differentiation of melanoma cells to Schwann-like cells. In melanoma cancer cell lines, both lopinavir and lopinavir-NO induced morphological changes, minor apoptosis and reactive oxygen species (ROS) production. However, caspase activation and autophagy were detected only in B16 cells, indicating a cell line-specific treatment response. Lopinavir-NO released NO intracellularly, and NO neutralization restored cell viability. Treatment with lopinavir-NO induced only a transient activation of Akt and inhibition of P70S6 kinase. The results of this study identify lopinavir-NO as a promising candidate for further clinical trials in melanoma and possibly other solid tumours.
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... This specific pattern is also known today as "conserved transcriptional response to adversity" (CTRA) and is considered a molecular signature of chronic stress [26], which is ultimately a part of the biological stress response (SR) already described. While the acute inflammatory response is a time-limited adaptation of the body to increased activity of the immune system to fight injury or infection, the chronic inflammatory response is a maladaptive condition [15,27,28]. This is associated with an increased risk of certain cancers and neurodegenerative and cardiovascular diseases, as well as asthma, arthritis, and mental disorders [29,30]. ...
Chromosomal Processes in Mind-Body Medicine: Chronic Stress, Cell Aging, and Telomere Length
Article
Full-text available
Stress affects cellular aging and inflammatory and chromosomal processes, including telomere length, thereby potentially compromising health and facilitating disease onset and progression. Stress-related diseases and strategies to manage stress usually require integrative or behavioral therapeutic approaches that also operate on cellular levels. Mind-body medicine (MBM) uses the interaction between the mind, body, behavior, and the environment to correct physical and psychological malfunctions, thus ameliorating disease states and improving health. The relaxation response (RR) is a physiological opponent of stress and the stress response (SR) (i.e., fight-or-flight response), also invoking molecular anti-stress processes. Techniques that elicit the RR are at the core of practically all MBM interventions. We surmise that these techniques can also affect chromosomal and telomere processes, molecular aging, and the modulation of inflammatory states on cellular levels.
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... This specific pattern is also known today as "conserved transcriptional response to adversity" (CTRA) and is considered a molecular signature of chronic stress [26], which is ultimately a part of the biological stress response (SR) already described. While the acute inflammatory response is a time-limited adaptation of the body to increased activity of the immune system to fight injury or infection, the chronic inflammatory response is a maladaptive condition [15,27,28]. This is associated with an increased risk of certain cancers and neurodegenerative and cardiovascular diseases, as well as asthma, arthritis, and mental disorders [29,30]. ...
Chromosomal Processes in Mind-Body Medicine: Chronic Stress, Cell Aging, and Telomere Length
Article
Full-text available
  • Sep 2018
Stress affects cellular aging and inflammatory and chromosomal processes, including telomere length, thereby potentially compromising health and facilitating disease onset and progression. Stress-related diseases and strategies to manage stress usually require integrative or behavioral therapeutic approaches that also operate on cellular levels. Mind-body medicine (MBM) uses the interaction between the mind, body, behavior, and the environment to correct physical and psychological malfunctions, thus ameliorating disease states and improving health. The relaxation response (RR) is a physiological opponent of stress and the stress response (SR) (i.e., fight-or-flight response), also invoking molecular anti-stress processes. Techniques that elicit the RR are at the core of practically all MBM interventions. We surmise that these techniques can also affect chromosomal and telomere processes, molecular aging, and the modulation of inflammatory states on cellular levels.
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... It has been demonstrated that some gene polymorphisms, such as the polymorphism in the promoter region of endothelial nitric oxide synthase, may increase the risk of TAO development (5). Because nitric oxide (NO) is responsible for vascular tone and inhibition Rep of platelet activation and aggregation, as well as inhibition of vascular oxidative stress (6,7), it has been suggested that NO may play a role in the pathophysiology of TAO (5). Due to the critical role of NO in the body, it is little wonder that NO has a backup system: the so-called heme oxygenase (HMOX) system (8). ...
The Status of Nitric Oxide and its Backup, Heme Oxygenase 1, in Thromboangiitis Obliterans
Article
Full-text available
  • Apr 2018
Background: Until recently, a gene polymorphism in the promoter region of endothelial nitric oxide synthase has been suggested as a risk factor for thromboangiitis obliterans (TAO) development. The aim of this study was to compare the metabolites of nitric oxide (NO) and its backup, heme-oxygenase-1 (HMOX1), between TAO patients and those of a smoking control group matched by race, age, sex, and smoking habits. Methods: Twenty-four male Caucasian TAO patients and 20 male Caucasian controls enrolled in the study. Their smoking habits were matched based on the serum cotinine levels of 17 of the TAO patients and the 20 controls. A colorimetric kit was used to measure NO, and an enzyme-linked immunosorbent assay kit was used to measure cotinine and HMOX1 levels. Results: The mean serum level of NO metabolites in the TAO group was significantly less than in the controls (p = 0.03) and also significantly less in the patients with below-knee amputations than in non-amputees (p= 0.018). Also, HMOX1 was significantly greater in the TAO patients than in the controls (p= 0.01). No significant correlation was found between NO and HMOX1 (p = 0.054). Conclusion: Nitric oxide may play a pivotal role in TAO development and its outcome. However, the intact HMOX1 pathway may demonstrate the unique role of NO, which cannot be compensated for by HMOX1 and whose absence may make patients susceptible to developing TAO. In addition, another pathway besides NO, with influence on vascular tone and hemostasis, might be involved in TAO development, such as the autonomic nervous system. Further studies are suggested regarding these issues.
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... Another important characteristic of the cell is its morphology which may change due to reasons such as nuclear, chromatin, and cytoskeleton damage. Overproduction of NO is considered to result in free radicals (15,30,31) and it has been shown to induce apoptosis at concentrations higher than physiologic levels (32,33). Thus, in this study, we have investigated cell morphology under the influence of SNP. ...
Sodium Nitroprusside Changed The Metabolism of Mesenchymal Stem Cells to An Anaerobic State while Viability and Proliferation Remained Intact
Article
Full-text available
  • Mar 2017
Objective We used sodium nitroprusside (SNP), a nitric oxide (NO) releasing molecule, to understand its effect on viability and proliferation of rat bone marrow mesenchymal stem cells (BM-MSCs). Materials and Methods This experimental study evaluated the viability and morphology of MSCs in the presence of SNP (100 to 2000 ?M) at 1, 5, and 15 hours. We chose the 100, 1000, and 2000 ?M concentrations of SNP for one hour exposure for further analyses. Cell proliferation was investigated by the colony forming assay and population doubling number (PDN). Na?, K?, and Ca?? levels as well as activities of lactate dehydrogenase (LDH), alkaline phosphatase (ALP), aspartate transaminase (AST), and alanine transaminase (ALT) were measured. Results The viability of MSCs dose-dependently reduced from 750 ?M at one hour and 250 ?M at 5 and 15 hours. The 100 ?M caused no change in viability, however we observed a reduction in the cytoplasmic area at 5 and 15 hours. This change was not observed at one hour. The one hour treatment with 100 ?M of SNP reduced the mean colony numbers but not the diameter when the cells were incubated for 7 and 14 days. In addition, one hour treatment with 100 ?M of SNP significantly reduced ALT, AST, and ALP activities whereas the activity of LDH increased when incubated for 24 hours. The same treatment caused an increase in Ca?? and reduction in Na? content. The 1000 and 2000 ?M concentrations reduced all the factors except Ca?? and LDH which increased. Conclusion The high dose of SNP, even for a short time, was toxic. The low dose was safe with respect to viability and proliferation, especially over a short time. However elevated LDH activity might increase anaerobic metabolism.
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... Nitric oxide (NO) is considered a biologic mediator; plays a vital role in different biologic processes; and is an essential component in the fields of physiology, biochemistry, immunology, and neuroscience [2,3]. Nitric oxide and polyamines are very important for implantation and development of embryos and extra-embryonic membranes; however, mechanisms regulating their biosynthesis in the uterus, embryos, and extra-embryonic membranes are mostly unknown [4]. ...
The nitric oxide serum level and combined utero placental thickness in buffalo (Bubalus bubalis) affected by pregnancy pathology
Article
  • Aug 2016
The present study aimed mainly to study the level of nitric oxide (NO) and combined utero placental thickness (CTUP) in buffaloes affected by pregnancy pathology. Females (n = 104) were classified into three main groups: non-pregnant (n = 10), healthy pregnant (n = 54), and pathologically pregnant (n = 40). The healthy pregnant animals were sub-grouped according to the stage of pregnancy to early stage (n = 15), mid-stage (n = 15), late stage (n = 19), and full term (n = 5). The animals in which pregnancy was associated with placental pathology were subgrouped to uterine torsion (n = 27), hydro-allantois (n = 5), and abortion (n = 8). Blood samples were collected from all animals to estimate the NO level using Griess reaction test. CTUP was measured at the most caudal part of the pregnant horn using a rectal ultrasound probe. Placental tissue samples were collected from all pregnant animals just after delivery or abortion for histopathologic section. The results revealed that the level of NO was higher (P ≤ 0.05) in pregnant than in non-pregnant animals (6.89±0.18 µM). Additionally, NO level was decreased significantly at full term (18.26±0.27 µM) when compared to the other stages of pregnancy. Moreover, there was a decrease (P ≤ 0.05) in the level of NO in cases of uterine torsion (22.22±0.46 µM) and hydro-allantois (24.20±0.07 µM) in comparison with the normal pregnant buffalo. The CTUP was significantly increased as pregnancy progressed. The CTUP in cases of uterine torsion (18.2±3.3 mm) was higher (P < 0.05) than in healthy pregnant buffalo at the same stage of pregnancy. Additionally, CTUP significantly differed between different types of uterine torsion. There was no significant difference between CTUP in hydro-allantois and abortion cases besides healthy pregnancy. Histopathologic examination revealed a great proliferation and hyperplasia of cytotrophoblastic cells only in uterine torsion and hydro-allantois cases. In conclusion, NO level and CTUP can be used as indicators for pregnancy pathology that is usually associated with presence of cytotrophoblastic cells and placental insufficiency.
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Nitric oxide-induced S-nitrosylation of glyceraldehyde-3-phosphate dehydrogenase inhibits enzymatic activity and increases endogenous ADP-ribosylation
Article
Full-text available
  • Jan 1993
Using conditions that produced chronic inflammation in rat liver, we were able to find a correlation between induction of nitric oxide production and inhibition of glyceraldehyde-3-phosphate dehydrogenase (GAPDH; EC 1.2.1.12). This enzyme is a tetramer composed of identical M(r) 37,000 subunits. The tetramer contains 16 thiol groups, four of which are essential for enzymatic activity. Our information indicates that four thiol groups are S-nitrosylated by exposure to authentic nitric oxide (NO) gas. Furthermore, NO decreased GAPDH activity while increasing its auto-ADP-ribosylation. Reduced nicotinamide adenine dinucleotide and dithiothreitol are required for the S-nitrosylation of GAPDH caused by the NO-generating compound sodium nitroprusside. Our results suggests that a new and important action of nitric oxide on cells is the S-nitrosylation and inactivation of GAPDH. S-Nitrosylation of GAPDH may be a key covalent modification of multiple regulatory consequences in chronic liver inflammation.
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INDUCTION AND STABILIZATION OF I-KAPPA-B-ALPHA BY NITRIC-OXIDE MEDIATES INHIBITION OF NF-KAPPA-B
Article
  • Jun 1995
To determine the mechanism(s) by which the endogenous mediator nitric oxide (NO) inhibits the activation of transcription factor NF-kappa B, we stimulated human vascular endothelial cells with tumor necrosis factor-alpha in the presence of two NO donors, sodium nitroprusside and S-nitrosoglutathione. Electrophoretic mobility shift assays demonstrated that both NO donors inhibited NF-kappa B activation by tumor necrosis factor-alpha. This effect was not mediated by guanylyl cyclase activation since the cGMP analogue 8-bromo-cGMP had no similar effect. Inhibition of endogenous constitutive NO production by L-N-monomethylarginine, however, activated NF-kappa B, suggesting tonic inhibition of NF-kappa B under basal conditions. NO had little or no effects on other nuclear binding proteins such as AP-1 and GATA. Immunoprecipitation studies showed that NO stabilized the NF-kappa B inhibitor, I kappa B alpha, by preventing its degradation from NF-kappa B. NO also increased the mRNA expression of I kappa B alpha, but not NF-kappa B subunits, p65 or p50, and transfection experiments with a chloramphenicol acetyltransferase reporter gene linked to the I kappa B alpha promoter suggested transcriptional induction of I kappa B alpha by NO. me propose that the induction and stabilization of I kappa B alpha by NO are important mechanisms by which NO inhibits NF-kappa B and attenuate atherogenesis.
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Nitric oxide and mitochondrial respiration
Article
  • Aug 1997
We and others have found that physiological/pathological levels of nitric oxide (NO) reversibly inhibit mitochondrial respiration, by binding to cytochrome oxidase in competition with oxygen. We measured O2 and NO concentrations simultaneously using specific electrodes, and showed that nanomolar NO reversibly inhibits oxygen consumption of isolated cytochrome oxidase, mitochondria, synaptosomes, and cells. Oxygen consumption of nerve terminals isolated from brain (synaptosomes) was half inhibited by 270nM NO when the oxygen concentration is 150μM, but half inhibited by 60nM NO when the oxygen concentration was 30μM (a roughly physiological level of oxygen). Cultured astrocytes, activated to express the inducible form of NO synthase produced up to 1μM which strongly and reversibly inhibited cellular respiration via the inhibition of cytochrome oxidase. Mitochondria caused rapid NO breakdown, which may be due to reaction with cytochrome oxidase and/or mitochondrial superoxide. Long-term exposure to NO can cause irreversible damage to mitochondria and cells, which may be mediated by reactive oxygen species or peroxynitrite. We have found that NO reversibly inhibits catalase, with a Ki of 0.3μM NO, and that superoxide dismutase catalyses the reaction of NO and H2O2 to produce peroxynitrite. These reactions might contribute to the cytotoxicity of NO.
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Morphine Inhibits NF-κB Nuclear Binding in Human Neutrophils and Monocytes by a Nitric Oxide–dependent Mechanism
Article
The transcription factor NF-kappaB plays a pivotal role in gene expression of inflammatory mediators such as cytokines or adhesion molecules. NF-kappaB-mediated transcriptional activation of these genes is inhibited by nitric oxide (NO) in a variety of cells, including monocytes. Morphine mediates NO release in a naloxone antagonizable manner in monocytes and neutrophils. The influence of morphine on NF-kappaB activation was investigated in a whole-blood flow cytometric assay. A specific antibody against the p65 subunit of NF-kappaB was used and detected by fluoresceine-isothiocyanate-labeled anti-immunoglobulin G. Nuclei were stained with propidium iodide. Leukocyte subpopulations were evaluated by gating on neutrophils and monocytes. The median fluorescence channel was determined. Different morphine concentrations (50 nm, 50 microm, 1 mm) and incubation intervals (10-150 min) were used. Morphine inhibits lipopolysaccharide-induced NF-kappaB nuclear binding in human blood neutrophils and monocytes in a time-, concentration-, and naloxone-sensitive-dependent manner. Similar effects were achieved with the NO donor S-nitroso-N-acetyl-pencillamine and the antioxidant N-acetyl-cysteine. The NO synthase inhibitors Nomega-nitro-l-arginine-methyl-esther and Nomega-nitro-l-arginine completely abolished the morphine-induced attenuation of NF-kappaB nuclear binding, demonstrating that the inhibitory action is mediated by NO release. Morphine causes immunosuppression, at least in part, via the NO-stimulated depression of NF-kappaB nuclear binding.
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Free Radicals and Antioxidants in the Year 2000: A Historical Look to the Future
Article
In the late 1950's free radicals and antioxidants were almost unheard of in the clinical and biological sciences but chemists had known about them for years in the context of radiation, polymer and combustion technology. Daniel Gilbert, Rebeca Gerschman and their colleagues related the toxic effects of elevated oxygen levels on aerobes to those of ionizing radiation, and proposed that oxygen toxicity is due to free radical formation, in a pioneering paper in 1956. Biochemistry owes much of its early expansion to the development and application of chromatographic and electrophoretic techniques, especially as applied to the study of proteins. Thus, superoxide dismutase (SOD) enzymes (MnSOD, CuZnSOD, FeSOD) were quickly identified. By the 1980's Molecular Biology had evolved from within biochemistry and microbiology to become a dominant new discipline, with DNA sequencing, recombinant DNA technology, cloning, and the development of PCR representing milestones in its advance. As a biological tool to explore reaction mechanisms, SOD was a unique and valuable asset. Its ability to inhibit radical reactions leading to oxidative damage in vitro often turned out to be due to its ability to prevent reduction of iron ions by superoxide. Nitric oxide (NO·) provided the next clue as to how SOD might be playing a critical biological role. Although NO· is sluggish in its reactions with most biomolecules it is astoundingly reactive with free radicals, including superoxide. Overall, this high reactivity of NO· with radicals may be beneficial in vivo, e.g. by scavenging peroxyl radicals and inhibiting lipid peroxidation. If reactive oxygen species are intimately involved with the redox regulation of cell functions, as seems likely from current evidence, it may be easier to understand why attempts to change antioxidant balance in aging experiments have failed. The cell will adapt to maintain its redox balance. Indeed, transgenic animals over-expressing antioxidants show some abnormalities of function. There must therefore be a highly complex interrelationship between dietary, constitutive, and inducible antioxidants within the body, under genetic control. The challenge for the new century is to be able to understand these relationships, and how to manipulate them to our advantage to prevent and treat disease.
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Oxygen-Dependent Regulation of Mitochondrial Energy Metabolism by Nitric Oxide
Article
  • Nov 1995
To elucidate the role of nitric oxide (NO) in the regulation of mitochondrial function, the effect of NO on energy transfer reactions was examined under different oxygen tensions. Mitochondrial respiration was remarkably inhibited by NO resulting in the inhibition of ATP synthesis in a concentration-dependent manner. In the presence of succinate, respiration, respiratory control by ADP, and ATP synthesis recovered completely at certain times after adding NO. The inhibitory action of NO continued significantly longer under physiologically low oxygen concentrations (such as the cytosolic level) than at high concentrations. In the presence of various substrates, such as pyruvate malate, succinate, and ascorbate tetramethyl paraphenylenediamine, NO also inhibited the uncoupled respiration in an oxygen- and concentration-dependent manner. The inhibition of respiration by NO was stoichiometrically suppressed by oxyhemoglobin. When added to a mitochondrial suspension, NO rapidly disappeared from the medium particularly at high oxygen tension. However, the rate of NO disappearance was significantly lower under low oxygen tension. Thus, under cytosolic oxygen concentration, NO might play an important role in the regulation of mitochondrial energy metabolism.
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Basal nitric oxide limits immune, nervous and cardiovascular excitation: Human endothelia express a mu opiate receptor
Article
Nitric oxide (NO) is a major signaling molecule in the immune, cardiovascular and nervous systems. The synthesizing enzyme, nitric oxide synthase (NOS) occurs in three forms: endothelial (e), neuronal (n) and inducible (i) NOS. The first two are constitutively expressed. We surmise that in many tissues there is a basal level of NO and that the actions of several signaling molecules initiate increases in cNOS-derived NO to enhance momentary basal levels that exerts inhibitory cellular actions, via cellular conformational changes. It is our contention that much of the literature concerning the actions of NO really deal with i-NOS-derived NO. We make the case that cNOS is responsible for a basal or ‘tonal’ level of NO; that this NO keeps particular types of cells in a state of inhibition and that activation of these cells occurs through disinhibition. Furthermore, naturally occurring signaling molecules such as morphine, anandamide, interleukin-10 and 17-β-estradiol appear to exert, in part, their beneficial physiological actions, i.e., immune and endothelial down regulation by the stimulation of cNOS. In regard to opiates, we demonstrate the presence of a human endothelial mu opiate receptor by RT-PCR and sequence determination, further substantiating the role of opiates in vascular coupling to NO release. Taken together, cNOS derived NO enhances basal NO actions, i.e., cellular activation state, and these actions are further enhanced by iNOS derived NO.
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Mechanisms of the pro- and anti-oxidant action of nitric oxide in atherosclerosis
Article
  • Sep 2000
The association of nitric oxide (NO) with cardiovascular disease has long been recognized and the extensive research on this topic has revealed both pro- and anti-atherosclerotic effects. While these contradictory findings were initially perplexing recent studies offer molecular mechanisms for the integration of these data in the context of our current understanding of the biochemistry of NO. The essential findings are that the biochemical properties of NO allow its exploitation as both a cell signaling molecule, through its interaction with redox centers in heme proteins, and an extremely rapid reaction with other biologically relevant free radicals. The direct reaction of NO with free radicals can have either pro- or antioxidant effects. In the cell, antioxidant properties of NO can be greatly amplified by the activation of signal transduction pathways that lead to the increased synthesis of endogenous antioxidants or down regulate responses to pro-inflammatory stimuli. These findings will be discussed in the context of atherosclerosis.
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Nitric oxide as antioxidant
Article
  • Sep 1991
Benzoate monohydroxy compounds, and in particular salicylate, were produced during interaction of ferrous complexes with hydrogen peroxide (Fenton reaction) in a N2 environment. These reactions were inhibited when Fe complexes were flushed, prior to the addition in the model system, by nitric oxide. Methionine oxidation to ethylene by Fenton reagents was also inhibited by nitric oxide. Myoglobin in several forms such as metmyoglobin, oxymyoglobin, and nitric oxide-myoglobin were interacted with an equimolar concentration of hydrogen peroxide. Spectra changes in the visible region and the changes in membrane (microsomes) lipid peroxidation by the accumulation of thiobarbituric acid-reactive substances (TBA-RS) were determined. The results showed that metmyoglobin and oxymyoglobin were activated by H2O2 to ferryl myoglobin, which initiates membrane lipid peroxidation; but not nitric oxide-myoglobin, which, during interaction with H2O2, did not form ferryl but metmyoglobin which only poorly affected lipid peroxidation. It is assumed that nitric oxide, liganded to ferrous complexes, acts to prevent the prooxidative reaction of these complexes with H2O2.
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