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Superoxide, peroxynitrite and oxidative/nitrative stress in inflammation

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A considerable body of evidence suggests that formation of potent reactive oxygen species and resulting oxidative/nitrative stress play a major role in acute and chronic inflammation and pain. Much of the knowledge in this field has been gathered by the use of pharmacological and genetic approaches. In this mini review, we will evaluate recent advances made towards understanding the roles of reactive oxygen species in inflammation, focusing in particular on superoxide and peroxynitrite. Given the limited space to cover this broad topic, here we will refer the reader to comprehensive review articles whenever possible.
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Proteins: Structure and Function 965
Superoxide, peroxynitrite and oxidative/nitrative
stress in inflammation
D. Salvemini*
1
, T.M. Doyle* and S. Cuzzocrea†‡
*Department of Internal Medicine, Division of Pulmonary, Critical Care and Sleep Medicine, Saint Louis University School of Medicine, 3635 Vista Avenue, St.
Louis, MO 63110-0250, U.S.A., Department of Clinical and Experimental Medicine and Pharmacology, University of Messina, Messina, Italy, and IRCCS
(Istituti di Ricovero e Cura a Carattere Scientifico) Centro Neurolesi ‘Bonino-Pulejo’, Messina, Italy
Abstract
A considerable body of evidence suggests that formation of potent reactive oxygen species and resulting
oxidative/nitrative stress play a major role in acute and chronic inflammation and pain. Much of the
knowledge in this field has been gathered by the u se of pharmacological and genetic approaches. In this
mini review, we will evaluate recent advances made towards understanding the roles of reactive oxygen
species in inflammation, focusing in particular on superoxide and peroxynitrite. Given the limited space to
cover this broad topic, here we will refer the reader to comprehensive review articles whenever possible.
Superoxide
Superoxide is formed from various sources, including normal
cellular respiration, activated polymorphonuclear leucocytes,
endothelial cells and mitochondrial electron flux [1,2]. It
is now well appreciated that, under physiological circum-
stances, the biological reactivity of superoxide is tightly
controlled by SOD (superoxide dismutase) enzymes.
These include the manganese-complexed enzyme in mito-
chondria (MnSOD) and the copper/zinc-complexed enzyme
(CuZnSOD) present in the cytosol and extracellular surfaces
[3]. During acute and chronic inflammations, superoxide
is produced at rates that overwhelm the capacity of the
endogenous SOD enzyme defence system to remove it,
resulting in superoxide-mediated injury [4,5]. Important pro-
inflammatory roles for superoxide include: (i) endothelial
cell damage and increased microvascular permeability [6,7];
(ii) up-regulation of adhesion molecules such as ICAM-1
(intercellular adhesion molecule 1) and P-selectin (through
mechanisms not yet defined) that r ecruit neutrophils to
sites of inflammation [8,9]; (iii) autocatalytic destruction
of neurotransmitters and hormones such as noradrenaline
(norepinephrine) and adrenaline (epinephrine) respectively
[10]; (iv) lipid peroxidation and oxidation; (v) DNA
damage [11]; and (vi) activation of PARP [poly(ADP-ribose)
polymerase] [12]. PARP activation in particular is important
in inflammation and is a target for therapeutic inter-
vention; inhibitors of PARP activation such as nicotinamide
and 3-aminobenzamide attenuate both acute and chronic
inflammatory processes [12]. Superoxide also activates redox-
sensitive transcription factors including NF-κ B (nuclear
factor κ B) and AP-1 (activator protein 1) that in turn
regulate genes encoding various pro-inflammatory and pro-
Key words: inflammation, pain, peroxynitrite, reactive oxygen species, superoxide, tyrosine
nitration.
Abbreviations used: ALS, amyotrophic lateral sclerosis; PARP, poly(ADP-ribose) polymerase;
SOD, superoxide dismutase.
1
To whom correspondence should be addressed (email salvemd@slu.edu).
nociceptive cytokines [9,13–17]. In addition, superoxide
interacts with and destroys the biological activity of nitric
oxide [18] (Figure 1). Importantly, the rate of interaction
between superoxide and nitric oxide is faster than the rate
of removal of superoxide by SOD [19]. As a consequence,
important anti-inflammatory and tissue-protective properties
of nitric oxide may be attenuated, notably maintenance of
blood vessel tone, inhibition of platelet adhesion/aggregation
and cytoprotection in numerous organs (including heart,
intestine and kidney) [20]. Nitric oxide is known to mediate
several of its beneficial effects through the activation of cyclo-
oxygenase and subsequent release of beneficial and anti-
inflammatory prostaglandins [21,22]. By interacting with
superoxide, nitric oxide loses its ability to activate this
enzyme.
Taken together, it is therefore not surprising that removal
of superoxide by exogenous administration of SOD has been
shown to be beneficial in a broad range of inflammatory
diseases, both preclinically and clinically [5,23–26]. In
addition, the native SOD enzyme attenuates intestinal injury
induced by alcohol [27], Helicobacter pylori [28] and non-
steroidal anti-inflammatory drugs including indomethacin,
diclofenac and flurbiprofen [29]. When tested in humans
in various clinical trials, bovine CuZnSOD (Orgotein
R
)
showed promising anti-inflammatory effects under acute and
chronic conditions including rheumatoid arthritis, osteo-
arthritis as well as the deleterious side effects associated with
chemotherapy and radiation therapy [24]. Thus the use of
the native enzyme in clinical trials supports the concept that
removal of superoxide is beneficial. However, there were
drawbacks associated with its use, primarily the non-
human origin of the bovine enzyme. This inevitably led to
immunological reactions causing its removal from the market,
except in Spain where it is still clinically used to prevent
radiation-induced side effects [24].
Based on the concept that removal of superoxide modulates
the course of inflammation, several low-molecular-mass
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966 Biochemical Society Transactions (2006) Volume 34, part 5
Figure 1 Interaction between superoxide and nitric oxide
In health, the levels of superoxide are well controlled by endogenous SOD, thus not interfering with the beneficial anti-
inflammatory and cytoprotective effects of nitric oxide (NO) released from the constitutive nitric oxide enzyme (cNOS). In
disease states, MnSOD is nitrated and inactivated, favouring superoxide (O
2
) to increase. Superoxide interacts with NO,
destroying its beneficial effects. Over time, the levels of superoxide and NO [generated by induction of the inducible form
of NO synthase (iNOS)] increase greatly, tilting the balance O
2
/NO towards formation of the potent pro-inflammatory and
pro-apoptotic mediator peroxynitrite (ONOO
).
mimetics of the SOD enzymes have been developed t hat
could overcome some of the limitations associated with
bovine CuZnSOD for potential use as pharmaceuticals. These
include manganese-based metalloporphyrins such as
MnTBAP [Mn(III)tetrakis (4-benzoic acid) porphyrin], the
Mn(III) salen complexes such as EUK-8 [manganese N,N
-
bis(salicyldene)ethylenediamine chloride] and EUK-134
[manganese 3-methoxy-N ,N
-bis(salicyldene)ethylenedi-
amine chloride], the nitroxides such as TEMPOL (4-
hydroxy-2,2,6,6-tetramethylpiperidine-N -oxyl) and the
Mn(II)(penta-azamacrocyclic ligand)-based complexes
SC-55858, M40403 and M40401 [24,30–33] (Figure 2). Of
these, M40403 advanced to Phase II clinical studies in the
U.S.A. [34]. Numerous preclinical studies have shown
these low-molecular-mass antioxidants to be potently anti-
inflammatory in animal models of tissue injury and acute/
chronic inflammation [5,24,35]. In addition, recent results
have demonstrated that removal of superoxide and super-
oxide-derived reactive species is an attractive strategy to
inhibit peripheral and central sensitization associated with
several pain states. Thus these agents as well as other
antioxidants have been reported to be effective against acute
inflammatory pain [36,37], glutamate-induced hyperalgesia
[38], neuropathic pain [39–44], trigeminal pain, fibromyalgia
and temporomandibular joint dysfunction [45,46], chronic
pancreatitis [47], post-irradiation of breast cancer fibrosis
[48] and morphine-induced hyperalgesia and associated
anti-nociceptive tolerance [49,50].
Peroxynitrite and oxidative/nitrative
stress
Besides destroying the biological activity of nitric oxide (a key
anti-inflammatory and cytoprotective agent), a fundamental
consequence of the interaction between superoxide and
nitric oxide is in situ formation of peroxynitrite [51], a
potent cytotoxic and pro-inflammatory molecule [12,52–55]
(Figure 3). Thus removal of peroxynitrite by agents such as
FeTMPS [5,10,15,20-tetrakis(2,4,6-trimethyl-3,5-disulfonat-
ophenyl)-porphyrinato iron (III)] [53,55,56], FP-15 [57–59]
or WW85 [60] results in cytoprotective and anti-inflam-
matory effects. Although the actions of peroxynitrite in
inflammation are numerous, a mechanism that is receiving
greatest attention is its ability to nitrate protein tyrosine
groups that modify the functional activity of key proteins
[61] (Figure 3).
At least two well-described pathways can lead to such
protein nitration, one involving peroxynitrite [51] and the
other utilizing H
2
O
2
and myeloperoxidase [62]. Involve-
ment of these pathways in a particular pathophysiological
setting can be dissected pharmacologically with agents that
remove superoxide or nitric oxide, thereby preventing
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Proteins: Structure and Function 967
Figure 2 Some examples of SOD mimetics
Several classes of synthetic SOD mimetics have been developed and shown to be effective in various animal models of
acute/chronic inflammation and pain
Figure 3 Peroxynitrite-mediated tyrosine nitration plays a key role in inflammation and pain
Nitration can be focused on specific tyrosine residues on proteins and potentially results in modification, loss ()orgain()
of function.
the formation of peroxynitrite and peroxynitrite-mediated
protein nitration [36,38,63,64]. Interestingly, these agents do
not inhibit nitration driven by reactions other than peroxy-
nitrite and thus are useful pharmacological tools to ex-
plore the pathophysiological consequences of peroxynitrite-
driven nitration. Biologic nitration of tyrosine to form
3-nitrotyrosine is associated with numerous diseases in-
cluding transplant rejection, lung infection, central nervous
system and ocular inflammations, septic shock, cancer and
neurological disorders [ALS (amyotrophic lateral sclerosis),
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2006 Biochemical Society
968 Biochemical Society Transactions (2006) Volume 34, part 5
stroke, Parkinson’s disease] [19]. Recently, our groups linked
nitration and inactivation of MnSOD to the development
of peripheral and central pain sensitization [36,38]. Proteins
typically are composed of 4% tyrosine residues although
chemical nitration of isolated proteins modifies only a subset
of these. The basis for t his selectivity is not fully understood
[61,65]. Although there is no clearly defined mechanism of
removal of this modification, there is evidence that such a
signal could be turned off by either protein degradation
or denitration. With respect to the latter, a specific activity
termed denitrase, that removes the nitro group without
degrading the protein, has been shown to occur in tissues
[66]. Identification of the molecular basis for such an activity
awaits further study.
The advent of proteomics and the development of
numerous immunological and analytical methodologies have
revealed that tyrosine nitration is limited to specific proteins
[61]. Nitration can be focused on specific tyrosine residues
on proteins and potentially result in modification, loss or
gain of function [67–69]. A key example of lost enzyme
activity due to nitration in vivo is mitochondrial MnSOD,
the enzyme that normally keeps concentrations of superoxide
under tight control [3]. This protein is nitrated by per-
oxynitrite on Tyr-34 by an Mn-catalysed process that leads
to enzyme inactivation [70]. The importance of MnSOD
nitration/inactivation is highlighted by an overwhelming
body of evidence linking nitration of this enzyme to diseases
driven by overt production of peroxynitrite. These include
ischaemia and reperfusion injury, organ transplantation,
shock and inflammation, neurodegeneration in Alzheimer’s
disease, ALS or AIDS dementia complex [71–73] and pain
[36,38].
Nitration and inactivation of mitochondrial MnSOD
favour the accumulation of peroxynitrite [65], which then
nitrates and alters additional proteins and receptors perpetu-
ating and extending the initial damage. For example, nitration
of the mitochondrial proteins aconitase, cytochrome c,
voltage-dependent anion channel, ATPase and succinyl-CoA
oxoacid-CoA transferase [65,74] disrupts mitochondrial
metabolism. Such a mitochondrial injury in turn triggers
apoptotic signalling of cell death. Importantly, the enhanced
peroxidative activity of nitrated cytochrome c [65,67] may
further contribute to oxidative damage. Mitochondrial
dysfunction is thought to represent an important component
of various cardiovascular [75] and neurological disorders
[76–78] as well as neuropathic pain states [79].
Other important consequences of protein tyrosine nitra-
tion are being evaluated in various settings. For example,
glutamate transporters and glutamine synthase play a central
role in regulating the homoeostasis of extracellular glutamate
and its metabolic fate. These proteins are also nitrated by
peroxynitrite [80–84]. Nitration of glutamate transporters
by peroxynitrite inhibits their ability to transport glutamate
from the synaptic cleft to the neurons where it is then
metabolized to non-toxic glutamine by glutamine synthase
[85]. Nitration of the active site tyrosine residue (Tyr-160)
on glutamine synthase by peroxynitrite destroys its en-
zymatic activity [82–84]. As a consequence, excitotoxic
and neurotoxic glutamate accumulates in the synaptic cleft
where it is not metabolized, and within neurons, thereby
leading to neurotoxicity. In addition, peroxynitrite nitrates
tyrosine residues present on the NMDA receptor subunits,
an event leading to constant potentiation of synaptic currents,
calcium influx and ultimately neuronal excitotoxicity [86–88].
Therefore optimal glutamatergic transmission can be com-
promised by nitration, potentially contributing to the severity
of stroke, spinal cord injury, neuropathic pain and opiate-
induced hyperalgesia and tolerance all of which are known
to be impacted by the excessive presence of glutamate
[89–97]
Functional alteration of proteins through nitration can
also have a significant impact in cardiovascular diseases. In
particular we now know that the endothelial enzyme prosta-
cyclin synthase is nitrated by peroxynitrite at haem-
adjacent Tyr-430 in a process catalysed by the active site
haem-thiolate involving transient ferryl species [98,99].
Importantly, this enzyme co-localizes with the endothelial
form of nitric oxide synthase in the caveolae, which may
accentuate it as a sensitive and critical target of peroxy-
nitrite. Recent experiments indicate that inflammation of
arterial walls during atherosclerosis results in rapid prosta-
cyclin synthase nitration by a peroxynitrite-dependent
mechanism [99]. Prostacyclin, the product of this enzyme, is
a powerful anti-thrombotic and vasodilatory mediator that
is known to also limit vascular remodelling and cholesterol
uptake [100,101]. Thus nitration of prostacyclin synthase
by peroxynitrite reduces local prostacyclin while leaving
concentrations of thromboxane A
2
, a potent pro-thrombotic
and vasoconstricting mediator also known to promote
vascular remodelling [100,101], relatively unaffected. Thus
peroxynitrite nitration of prostacyclin synthase removes a
protective constraint on thrombogenesis, hypertension and
atherogenesis, by favouring accumulation of thromboxane
A
2
[102]. The risk of thrombotic events in cardiovascular
disorders could therefore be increased as a consequence of
peroxynitrite-mediated protein nitration.
Clearly, examples of this nature will grow as we begin to
unravel, understand and appreciate the impact that protein
nitration has in pathophysiological conditions.
Conclusions and prospectus
Superoxide and peroxynitrite play fundamental roles in
inflammation that makes them critical for the development of
novel anti-inflammatory agents. Protein tyrosine nitration is
gaining considerable interest in biomedical research, because
it can alter protein function, is associated with acute and
chronic disease states and can be a predictor of disease risk. A
comprehensive and focused approach, utilizing proteomics,
biochemical, pharmacological and genetic strategies to
understand the functional relevance of protein tyrosine
nitration, will identify novel targets that are pivotal to the
management of human diseases.
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2006 Biochemical Society
Proteins: Structure and Function 969
We thank Dr A.J. Lechner (Department of Pharmacological and
Physiological Sciences, Saint Louis University School of Medicine)
for his invaluable input.
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2006 Biochemical Society
... As well, NO mediates a wide range of both antitumor and antimicrobial activities [16][17][18][19]. Nevertheless, the exorbitant production of NO and/or several of its reaction products (e.g., peroxynitrite ( − OONO)) has been implicated in a vast range of pathological circumstances, such as chronic inflammation and infection conditions [20][21][22], septic shock syndrome [23][24], diabetes [25][26][27], Parkinson's [28][29] and Alzheimer's [30][31][32] diseases, and cancer [18,21,[33][34]. In light of these adverse effects, physiological NO concentrations must be tightly regulated, especially with respect to that of dioxygen (O 2 ) in order to minimize the generation of some highly reactive nitrogen oxide species (RNOS) such as peroxynitrite; peroxynitrite is known to produce via diffusion limited reactivity of superoxide anions and NO. ...
... These final electronic absorption features are in line with those of the previously known six-coordinate heme ferric nitrite complexes with an axially coordinated NO ligand [129][130], indicating the formation of the six-coordinate [(NO)(F 20 TPP)Fe III (NO 2 )] in this case (Scheme 3). This six-coordinate complex is only stable at cryogenic temperatures under an NO atmosphere [118,131], and warming up to room temperature and argon sparging results in the five-coordinate heme ferric These NO reactivities were also analyzed by 2 H NMR spectroscopy using the F 20 and NO at − 80 • C displayed a diamagnetic peak at δ pyrrole = 8.8 ppm (Fig. 3), and was EPR-silent (Fig. S5), lending credence to its assignment as the aforementioned six-coordinate [(NO)(F 20 TPP)Fe III (NO 2 )] complex; i.e., unpaired spins on the heme Fe III center and NO are antiferromagnetically coupled to give an overall diamagnetic spin system. Accordingly, following argon purging at RT, the conversion of [(NO) (F 20 TPP)Fe III (NO 2 )] to [(F 20 TPP)Fe III (NO 2 )] was evidenced by the paramagnetic shifting of the 2 H NMR signal to δ pyrrole = 80.7 ppm (Fig. 3), which is accompanied the formation of a new intense high-spin Fe III EPR feature at g = 5.8 (Figs. ...
Article
Dioxygen activating heme enzymes have long predicted to be powerhouses for nitrogen oxide interconversion, especially for nitric oxide (NO) oxidation which has far-reaching biological and/or environmental impacts. Lending credence, reactivity of NO with high-valent heme‑oxygen intermediates of globin proteins has recently been implicated in the regulation of a variety of pivotal physiological events such as modulating catalytic activities of various heme enzymes, enhancing antioxidant activity to inhibit oxidative damage, controlling inflammatory and infectious properties within the local heme environments, and NO scavenging. To reveal insights into such crucial biological processes, we have investigated low temperature NO reactivities of two classes of synthetic high-valent heme intermediates, Compound-II and Compound-I. In that, Compound-II rapidly reacts with NO yielding the six-coordinate (NO bound) heme ferric nitrite complex, which upon warming to room temperature converts into the five-coordinate heme ferric nitrite species. These ferric nitrite complexes mediate efficient substrate oxidation reactions liberating NO; i.e., shuttling NO2⁻ back to NO. In contrast, Compound-I and NO proceed through an oxygen-atom transfer process generating the strong nitrating agent NO2, along with the corresponding ferric nitrosyl species that converts to the naked heme ferric parent complex upon warmup. All reaction components have been fully characterized by UV–vis, ²H NMR and EPR spectroscopic methods, mass spectrometry, elemental analyses, and semi-quantitative determination of NO2⁻ anions. The clean, efficient, potentially catalytic NOx interconversions driven by high-valent heme species presented herein illustrate the strong prospects of a heme enzyme/O2/NOx dependent unexplored territory that is central to human physiology, pathology, and therapeutics.
... Moreover, hypomagnesemia causes a systemic stress response through neuroendocrinological pathways (change or activation of acetylcholine, catecholamines, release of substance P (SP), and activation of renin-angiotensin system RAAS) [170,176]. The inflammation affects pro-atherogenic changes in the metabolism of lipoproteins (increase lipoproteins oxidation), endothelial dysfunction (increased production of peroxynitrite, which damages cellular biomolecules and structures) [183,184], and influences host metabolism, e.g., lipid peroxidation [170,185]. The second mechanism is associated with increased reactive oxygen species, promoting membrane oxidation and the nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) production. ...
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Magnesium (Mg) is an essential nutrient for maintaining vital physiological functions. It is involved in many fundamental processes, and Mg deficiency is often correlated with negative health outcomes. On the one hand, most western civilizations consume less than the recommended daily allowance of Mg. On the other hand, a growing body of evidence has indicated that chronic hypomagnesemia may be implicated in the pathogenesis of various metabolic disorders such as overweight and obesity, insulin resistance (IR) and type 2 diabetes mellitus (T2DM), hypertension (HTN), changes in lipid metabolism, and low-grade inflammation. High Mg intake with diet and/or supplementation seems to prevent chronic metabolic complications. The protective action of Mg may include limiting the adipose tissue accumulation, improving glucose and insulin metabolism, enhancing endothelium-dependent vasodilation, normalizing lipid profile, and attenuating inflammatory processes. Thus, it currently seems that Mg plays an important role in developing metabolic disorders associated with obesity, although more randomized controlled trials (RCTs) evaluating Mg supplementation strategies are needed. This work represents a review and synthesis of recent data on the role of Mg in the pathogenesis of metabolic disorders.
... On the contrary, CP induces endothelial dysfunction and increases the inducible NO synthase (iNOS) level [61]. erefore, excessive NO reacts with superoxide and produces peroxynitrite which is a potent vasoconstrictor and easily causes oxidative damage to cellular structures [62,63]. In this study, the serum levels of nitrite in female rats treated with CP alone were more than male rats, probably due to increasing iNOS level and the presence of estrogen. ...
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Background: Cisplatin (CP) is widely used to treat various kinds of malignancies, but to avoid its side effects of nephrotoxicity and hypomagnesemia, magnesium supplementation is a subject of debate. The current study was designed to determine the protective role of intravenous magnesium sulfate (MgSO4) against intravenous administration of CP in male and female rats. Method: In this case-control experimental study, 80 Wistar male and female rats in 12 groups of experiments were subjected to receive intravenous administration of CP accompanied with intravenous infusion of different doses (1, 3, and 10 mg/ml solution) of MgSO4 and were compared with the control groups. Results: CP administration increased blood urea nitrogen (BUN), creatinine (Cr), kidney tissue damage score (KTDS), and kidney weight (KW), and they were attenuated by the mid-dose of MgSO4 supplementation in female rats. However, in male rats, the increase of Cr, BUN, KTDS, and KW induced by CP was ameliorated by low, mid-, and high doses of MgSO4 supplements. The levels of these markers were significantly different between male and female rats in the mid-dose of MgSO4-treated group (BUN: P=0.002, Cr: P=0.005, KTDS: P=0.002, and KW: P=0.031). CP reduced clearance of Cr (ClCr) in both male and female rats significantly compared to the control group of saline alone (P male = 0.002 and P female = 0.001), and the mid- and high doses of MgSO4 supplements improved ClCr in female rats. There were also sex differences in ClCr in mid- (P=0.05) and high (P=0.032) doses of MgSO4-treated groups. CP accompanied with the mid-dose of MgSO4 supplement reduced the KTDS (P male = 0.04 and P female = 0.004) and KW (P male = 0.002 and P female = 0.042) in both male and female rats significantly when compared with the CP-alone-treated group, while there were also significant differences between the sexes (KTDS: P=0.002 and KW: P=0.031). CP accompanied with three different doses of MgSO4 supplements did not improve the serum levels of lactate dehydrogenase, urine level of sodium, malondialdehyde, urine flow, and nitrite statistically when compared with the CP-alone-treated group. Conclusion: The renal protective effect of MgSO4 could be dose and gender related.
... For example, targeted studies of isolated mouse alveolar macrophages have shown that 4-aPDD activates TRPV4 to promote Ca 2+ influx and subsequent release of NO and superoxide (11). The combination of NO and superoxide can produce peroxynitrite, a strong oxidant involved in pathogen defense and inflammation (113,114). Untargeted global profiling of TRPV4-induced macrophage phenotypes could help to address important questions of how and why TRPV4 can have both pro-and anti-inflammatory responses, and further understand the underlying mechanisms involved. ...
Article
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Transient receptor potential vanilloid 4 (TRPV4) is a non-selective mechanosensitive ion channel expressed by various macrophage populations. Recent reports have characterized the role of TRPV4 in shaping the activity and phenotype of macrophages to influence the innate immune response to pathogen exposure and inflammation. TRPV4 has been studied extensively in the context of inflammation and inflammatory pain. Although TRPV4 activity has been generally described as pro-inflammatory, emerging evidence suggests a more complex role where this channel may also contribute to anti-inflammatory activities. However, detailed understanding of how TRPV4 may influence the initiation, maintenance, and resolution of inflammatory disease remains limited. This review highlights recent insights into the cellular processes through which TRPV4 contributes to pathological conditions and immune processes, with a focus on macrophage biology. The potential use of high-throughput and omics methods as an unbiased approach for studying the functional outcomes of TRPV4 activation is also discussed.
... It has cytotoxic activity against invading pathogens, and by oxidation and nitration reactions, cytotoxicity also affects mitochondrial function and induces cell death [34]. Peroxynitrite interacts via indirect, radical-mediated, and direct oxidative reaction mechanisms with macromolecules and this results in biological responses, including modulation of cell signaling pathways, acute and chronic inflammation and pain, increased synthesis of pro-inflammatory cytokines via NF-kB, and necrosis or apoptosis in cells [32,35,36]. The biological targets of peroxynitrite include tryptophan methionine and cysteine residues, centers of transition metal centers [37], erythrocytes, and oxidizes oxyhemoglobin [38], myoglobin [39], and cytochrome c [40]. ...
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Free radical contained one or more unpaired electrons in its valence shell, thus making it unstable, short-lived and highly reactive specie. Excessive generation of these free radicals ultimately leads to oxidative stress causing oxidation and damage to significant macromolecules in the living system and essentially disrupting signal transduction pathways and antioxidants equilibrium. At lower concentrations, ROS serves as “second messengers” influencing many physiological processes in the cell. However, at higher concentrations beyond cell capacity causes oxidative stress, which contributes to much human pathology such as diabetes, cancer, Parkinson’s disease, cardiovascular diseases, cataract, asthma, hypertension, atherosclerosis, arthritis and Alzheimer’s disease. Signaling pathways such as NF-κB, MAPKs, PI3K/Akt/ mTOR and Keap1-Nrf2-ARE modulates the detrimental effects of oxidative stress by increasing the expression of cellular antioxidant defenses, phase II detoxification enzymes and decreased production of ROS. Free radicals such as H2O2 are indeed needed for the advancement of cell cycle as these molecules influences DNA, proteins and enzymes in the cell cycle pathway. In the course of cell cycle progression, the cellular redox environment becomes more oxidized moving from G1 phase, becomes higher in G2/M and moderate in S phase. Signals in the form of an increase in cellular pro-oxidant levels are required and these signals are often terminated by a rise in the amount of antioxidants and MnSOD with a decrease in the level of cyclin D1 proteins. Therefore, understanding the mechanism of cell cycle redox regulation will help in therapy of many diseases.
... Oxidative stress appears to induce ME/CFS symptoms, associated with reduced blood flow and neuroinflammation. An infection (the most frequently reported trigger for ME/CFS onset) has been linked to peroxynitrite production, a proinflammatory oxygen/nitrogen species (57,58), triggering neuroinflammation. This can lead to the production of isoprostanes and cause vasoconstriction when the level of antioxidants is insufficient (59). ...
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Background: Myalgic Encephalomyelitis/Chronic Fatigue Syndrome (ME/CFS) is a multisystem medical condition with heterogeneous symptom expression. Currently, there is no effective cure or treatment for the standard care of patients. A variety of ME/CFS symptoms can be linked to the vital life functions of the brainstem, the lower extension of the brain best known as the hub relaying information back and forth between the cerebral cortex and various parts of the body. Objective/Methods: Over the past decade, Magnetic Resonance Imaging (MRI) studies have emerged to understand ME/CFS with interesting findings, but there has lacked a synthesized evaluation of what has been found thus far regarding the involvement of the brainstem. We conducted this study to review and evaluate the recent MRI findings via a literature search of the MEDLINE database, from which 11 studies met the eligibility criteria. Findings: Data showed that MRI studies frequently reported structural changes in the white and gray matter. Abnormalities of the functional connectivity within the brainstem and with other brain regions have also been found. The studies have suggested possible mechanisms including astrocyte dysfunction, cerebral perfusion impairment, impaired nerve conduction, and neuroinflammation involving the brainstem, which may at least partially explain a substantial portion of the ME/CFS symptoms and their heterogeneous presentations in individual patients. Conclusions: This review draws research attention to the role of the brainstem in ME/CFS, helping enlighten future work to uncover the pathologies and mechanisms of this complex medical condition, for improved management and patient care.
... These sites are not perfect even under normal conditions and can leak electrons out of the transport chain [90,91] (Figure 2). The leaked electrons can then partially reduce oxygen to form superoxide anion, which is the precursor of all other reactive oxygen species including H 2 O 2 , hydroxyl radical, and peroxynitrite [92,93] (Figure 3). Additionally, dihydrolipoamide dehydrogenase involved enzyme complexes such as pyruvate dehydrogenase, α-ketoglutarate dehydrogenase, and branched chain amino acid dehydrogenase can also produce superoxide anion in a variety of experimental and pathological conditions [94][95][96][97]. ...
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Cadmium is a nonessential metal that has heavily polluted the environment due to human activities. It can be absorbed into the human body via the gastrointestinal tract, respiratory tract, and the skin, and can cause chronic damage to the kidneys. The main site where cadmium accumulates and causes damage within the nephrons is the proximal tubule. This accumulation can induce dysfunction of the mitochondrial electron transport chain, leading to electron leakage and production of reactive oxygen species (ROS). Cadmium may also impair the function of NADPH oxidase, resulting in another source of ROS. These ROS together can cause oxidative damage to DNA, proteins, and lipids, triggering epithelial cell death and a decline in kidney function. In this article, we also reviewed evidence that the antioxidant power of plant extracts, herbal medicines, and pharmacological agents could ameliorate cadmium-induced kidney injury. Finally, a model of cadmium-induced kidney injury, centering on the notion that oxidative damage is a unifying mechanism of cadmium renal toxicity, is also presented. Given that cadmium exposure is inevitable, further studies using animal models are warranted for a detailed understanding of the mechanism underlying cadmium induced ROS production, and for the identification of more therapeutic targets.
... However, excessive ROS cause an imbalance in the prooxidant-antioxidant homeostasis and thus change signal transduction pathways. Similarly, residual RNS activates the expulsion of nitrogen intermediates by inducing nitrative stress (Salvemini et al. 2006). Being unstable biomolecules, ROS and RNS are not suitable for direct measurement. ...
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One of the major impacts of climate change has been the marked rise in global temperature. Recently, we demonstrated that high temperatures (1-week exposure) disrupt prooxidant-antioxidant homeostasis and promote cellular apoptosis in the American oyster. In this study, we evaluated the effects of seasonal sea surface temperature (SST) on tissue morphology, extrapallial fluid (EPF) conditions, heat shock protein-70 (HSP70), dinitrophenyl protein (DNP, an indicator of reactive oxygen species, ROS), 3-nitrotyrosine protein (NTP, an indicator of RNS), catalase (CAT), superoxide dismutase (SOD) protein expressions, and cellular apoptosis in gills and digestive glands of oysters collected on the southern Texas coast during the winter (15 °C), spring (24 °C), summer (30 °C), and fall (27 °C). Histological observations of both tissues showed a notable increase in mucus production and an enlargement of the digestive gland lumen with seasonal temperature rise, whereas biochemical analyses exhibited a significant decrease in EPF pH and protein concentration. Immunohistochemical analyses showed higher expression of HSP70 along with the expression of DNP and NTP in oyster tissues during summer. Intriguingly, CAT and SOD protein expressions exhibited significant upregulation with rising seasonal temperatures (15 to 27 °C), which decreased significantly in summer (30 °C), leaving oysters vulnerable to oxidative and nitrative damage. qRT-PCR analysis revealed a significant increase in HSP70 mRNA levels in oyster tissues during the warmer seasons. In situ TUNNEL assay showed a significant increase in apoptotic cells in seasons with high temperature. These results suggest that elevated SST induces oxidative/nitrative stress through the overproduction of ROS/RNS and disrupts the antioxidant system which promotes cellular apoptosis in oysters.
Article
Peroxynitrite (ONOO-), a highly reactive oxygen species (ROS), is implicated with many physiological and pathological processes including cancer, neurodegenerative diseases and inflammation. In this regard, developing effective tools for highly selective tracking of ONOO- is urgently needed. Herein, we constructed a concise and specific fluorescent probe NA-ONOO for sensing ONOO- by conjugating an ONOO--specific recognition group ((4-methoxyphenylthio)carbonyl, a thiocarbonate derivative) with a naphthalene fluorophore. The probe, NA-ONOO, was in a dark state because the high electrophilicity of (4-methoxyphenylthio)carbonyl disturbs the intramolecular charge transfer (ICT) in the fluorophore. Upon treatment with ONOO-, the fluorescent emission was sharply boosted (quantum yield Φ: 3% to 56.6%) owing to an ONOO- triggered release of (4-methoxyphenylthio)carbonyl from NA-ONOO. Optical analyses showed that NA-ONOO presented high selectivity and sensitivity toward ONOO-. With good cell permeability and biocompatibility, the NA-ONOO probe was successfully applied to imaging and tracing exogenous and endogenous ONOO- in living cells and zebrafish. The probe NA-ONOO presents a new recognition group and a promising method for further investigating ONOO- in living systems.
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With the continuous growth of the human population and new challenges in the quality of life, it is more important than ever to diagnose diseases and pathologies with high accuracy, sensitivity and in different scenarios from medical implants to the operation room. Although conventional methods of diagnosis revolutionized healthcare, alternative analytical methods are making their way out of academic labs into clinics. In this regard, surface-enhanced Raman spectroscopy (SERS) developed immensely with its capability to achieve single-molecule sensitivity and high-specificity in the last two decades, and now it is well on its way to join the arsenal of physicians. This review discusses how SERS is becoming an essential tool for the clinical investigation of pathologies including inflammation, infections, necrosis/apoptosis, hypoxia, and cancer. We critically discuss the strategies reported so far in nanoparticle assembly, functionalization, non-metallic substrates, colloidal solutions and how these techniques improve SERS characteristics during pathology diagnoses like sensitivity, selectivity, and detection limit. Moreover, it is crucial to introduce SERS’ recent developments and future perspectives as a biomedical analytical method. We finally discuss the challenges that remain as bottlenecks for a routine SERS implementation in the medical room from in vitro to in vivo applications. The review showcases the adaptability and versatility of SERS to resolve pathological processes by covering various experimental and analytical methods and the specific spectral features and analysis results achieved by these methods.
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An enzyme which catalyzes the dismutation of superoxide radicals (O2·⁻ + O2·⁻ + 2H⁺ → O2 + H2O2) has been purified by a simple procedure from bovine erythrocytes. This enzyme, called superoxide dismutase, contains 2 eq of copper per mole of enzyme. The copper may be reversibly removed, and it is required for activity. Superoxide dismutase has been shown to be identical with the previously described copper-containing erythrocuprein (human) and hemocuprein (bovine). Stable solutions of the superoxide radical were generated by the electrolytic reduction of O2 in an aprotic solvent, dimethylformamide. Slow infusion of such solutions into buffered aqueous media permitted the demonstration that O2·⁻ can reduce ferricytochrome c and tetranitromethane, and that superoxide dismutase, by competing for the superoxide radicals, can markedly inhibit these reactions. Superoxide dismutase was used to show that the oxidation of epinephrine to adrenochrome by milk xanthine oxidase is mediated by the superoxide radical. An assay of several tissues indicates that superoxide dismutase is widely distributed within mammalian organisms.
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The formation of the powerful oxidant peroxynitrite (PN) from the reaction of superoxide anion with nitric oxide has been shown to be a kinetically favored reaction contributing to cellular injury and death at sites of tissue inflammation. The PN molecule is highly reactive causing lipid peroxidation as well as nitration of both free and protein-bound tyrosine. We present evidence for the pharmacological manipulation of PN with decomposition catalysts capable of converting it to nitrate. In target cells challenged with exogenously added synthetic PN, a series of metalloporphyrin catalysts (5,10,15,20-tetrakis(2,4,6-trimethyl-3,3-disulfonatophenyl)porphyrinato iron (III) (FeTMPS); 5,10,15,20-tetrakis(4-sulfonatophenyl)porphyrinato iron (III) (FeTPPS); 5,10,15,20-tetrakis(N-methyl-4′-pyridyl)porphyrinato iron (III) (FeTMPyP)) provided protection against PN-mediated injury with EC50 values for each compound 30–50-fold below the final concentration of PN added. Cytoprotection was correlated with a reduction in the level of measurable nitrotyrosine. In addition, we found our catalysts to be cytoprotective against endogenously generated PN in endotoxin-stimulated RAW 264.7 cells as well as in dissociated cultures of hippocampal neurons and glia that had been exposed to cytokines. Our studies thus provide compelling evidence for the involvement of peroxynitrite in cytokine-mediated cellular injury and suggest the therapeutic potential of PN decomposition catalysts in reducing cellular damage at sites of inflammation.
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1. Previous results suggest that glutamine synthesis in brain could be modulated by nitrix oxide. The aim of this work was to assess this possibility. 2. As glutamine synthetase in brain is located mainly in astrocytes, we used primary cultures of astrocytes to assess the effects of increasing or decreasing nitrix oxide levels on glutamine synthesis in intact astrocytes. 3. Nitric oxide levels were decreased by adding nitroarginine, an inhibitor of nitric oxide synthase. To increase nitric oxide we used S-nitroso-N-acetylpenicillamine, a nitric oxide generating agent. 4. It is shown that S-nitroso-N-acetylpenicillamine decreases glutamine synthesis in intact astrocytes by ≈40–50%. Nitroarginine increases glutamine synthesis slightly in intact astrocytes. 5. These results indicate that brain glutamine synthesis may be modulated in vivo by nitric oxide.
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A major feature of septic shock is the development of a vascular crisis characterized by nonresponsiveness to sympathetic vasoconstrictor agents and the subsequent irreversible fall in blood pressure. In addition, sepsis, like other inflammatory conditions, results in a large increase in the production of free radicals, including superoxide anions (O2⨪) within the body. Here we show that O2⨪ reacts with catecholamines deactivating them in vitro. Moreover, this deactivation would appear to account for the hyporeactivity to exogenous catecholamines observed in sepsis, because administration of a superoxide dismutase (SOD) mimetic to a rat model of septic shock to remove excess O2⨪ restored the vasopressor responses to norepinephrine. This treatment with the SOD mimetic also reversed the hypotension in these animals; suggesting that deactivation of endogenous norepinephrine by O2⨪ contributes significantly to this aspect of the vascular crisis. Indeed, the plasma concentrations of both norepinephrine and epinephrine in septic rats treated with the SOD mimetic were significantly higher than in untreated rats. Interestingly, the plasma concentrations for norepinephrine and epinephrine were inversely related to the plasma concentrations of adrenochromes, the product of the autoxidation of catecholamines initiated by O2⨪. We propose, therefore, that the use of a SOD mimetic represents a new paradigm for the treatment of septic shock. By removing O2⨪, exogenous and endogenous catecholamines are protected from autoxidation. As a result, both hyporeactivity and hypotension are reversed, generation of potentially toxic adrenochromes is reduced, and survival rate is improved.
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Poly(ADP-ribose) polymerase-1 (PARP-1) is a member of the PARP enzyme family consisting of PARP-1 and several recently identified novel poly( ADP-ribosylating) enzymes. PARP-1 is an abundant nuclear protein functioning as a DNA nick-sensor enzyme. Upon binding to DNA breaks, activated PARP cleaves NAD(+) into nicotinamide and ADP-ribose and polymerizes the latter onto nuclear acceptor proteins including histones, transcription factors, and PARP itself. Poly(ADP-ribosylation) contributes to DNA repair and to the maintenance of genomic stability. On the other hand, oxidative stress-induced overactivation of PARP consumes NAD(+) and consequently ATP, culminating in cell dysfunction or necrosis. This cellular suicide mechanism has been implicated in the pathomechanism of stroke, myocardial ischemia, diabetes, diabetes-associated cardiovascular dysfunction, shock, traumatic central nervous system injury, arthritis, colitis, allergic encephalomyelitis, and various other forms of inflammation. PARP has also been shown to associate with and regulate the function of several transcription factors. Of special interest is the enhancement by PARP of nuclear factor kappaB-mediated transcription, which plays a central role in the expression of inflammatory cytokines, chemokines, adhesion molecules, and inflammatory mediators. Herein we review the double-edged sword roles of PARP in DNA damage signaling and cell death and summarize the underlying mechanisms of the anti-inflammatory effects of PARP inhibitors. Moreover, we discuss the potential use of PARP inhibitors as anticancer agents, radiosensitizers, and antiviral agents.
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Manganese superoxide dismutase (MnSOD) is essential for life as dramatically illustrated by the neonatal lethality of mice that are deficient in MnSOD. In addition, mice expressing only 50% of the normal compliment of MnSOD demonstrate increased susceptibility to oxidative stress and severe mitochondrial dysfunction resulting from elevation of reactive oxygen species. Thus, it is important to know the status of both MnSOD protein levels and activity in order to assess its role as an important regulator of cell biology. Numerous studies have shown that MnSOD can be induced to protect against pro-oxidant insults resulting from cytokine treatment, ultraviolet light, irradiation, certain tumors, amyotrophic lateral sclerosis, and ischemia/reperfusion. In addition, overexpression of MnSOD has been shown to protect against pro-apoptotic stimuli as well as ischemic damage. Conversely, several studies have reported declines in MnSOD activity during diseases including cancer, aging, progeria, asthma, and transplant rejection. The precise biochemical/molecular mechanisms involved with this loss in activity are not well understood. Certainly, MnSOD gene expression or other defects could play a role in such inactivation. However, based on recent findings regarding the susceptibility of MnSOD to oxidative inactivation, it is equally likely that post-translational modification of MnSOD may account for the loss of activity. Our laboratory has recently demonstrated that MnSOD is tyrosine nitrated and inactivated during human kidney allograft rejection and human pancreatic ductal adenocarcinoma. We have determined that peroxynitrite (ONOO-) is the only known biological oxidant competent to inactivate enzymatic activity, to nitrate critical tyrosine residues, and to induce dityrosine formation in MnSOD. Tyrosine nitration and inactivation of MnSOD would lead to increased levels of superoxide and concomitant increases in ONOO- within the mitochondria which, could lead to tyrosine nitration/oxidation of key mitochondrial proteins and ultimately mitochondrial dysfunction and cell death. This article assesses the important role of MnSOD activity in various pathological states in light of this potentially lethal positive feedback cycle involving oxidative inactivation.
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The relative contributions of superoxide anion (O2−) and peroxynitrite (PN) were evaluated in the pathogenesis of intestinal microvascular damage caused by the intravenous injection of E. coli lipopolysaccharide (LPS) in rats. The superoxide dismutase mimetic (SODm) SC-55858 and the active peroxynitrite decomposition catalysts 5,10,15,20-tetrakis(2,4,6-trimethyl-3,5-disulphonatophenyl)-porphyrinato iron (III) and 5,10,15,20-tetrakis(N-methyl-4′-pyridyl)-porphyrinato iron (III) (FeTMPS, FeTMPyP respectively) were used to assess the roles of O2− and PN respectively. The intravenous injection of LPS elicited an inflammatory response that was characterized by a time-dependent infiltration of neutrophils, lipid peroxidation, microvascular leakage (indicative of microvascular damage), and epithelial cell injury in both the duodenum and jejunum. Administration of the SODm SC-55858, FeTMPS or FeTMPyP at 3 h post LPS reduced the subsequent increase in microvascular leakage, lipid peroxidation and epithelial cell injury. Inactive peroxynitrite decomposition catalysts exhibited no protective effects. Only, SC-55858 inhibited neutrophil infiltration. Our results suggest that O2− and peroxynitrite play a significant role in the pathogenesis of duodenal and intestinal injury during endotoxaemia and that their removal by SODm and peroxynitrite decomposition catalysts offers a novel approach to the treatment of septic shock or clinical conditions of gastrointestinal inflammation. Furthermore, the remarkable protection of the intestinal epithelium by these agents suggests their use during chemo- and radiation therapy, cancer treatments characterized by gastrointestinal damage. Potential mechanisms through which these radicals evoke damage are discussed. British Journal of Pharmacology (1999) 127, 685–692; doi:10.1038/sj.bjp.0702604
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