Reactive Oxygen Species And Reactive Nitrogen Species Research Articles

Imaging of innate immunity activation in vivo with a redox-tuned PET reporter.

High-redox-potential reactive oxygen species and reactive nitrogen species (ROS/RNS), generated by NADPH oxidase-2 (NOX2), myeloperoxidase (MPO) and related enzymes, are key effector molecules of innate immunity. High-redox-potential radicals are difficult to distinguish by imaging from less potent ROS/RNS functioning as background biological signaling molecules. Here we present 4-[18F]fluoro-1-naphthol ([18F]4FN), a redox-tuned radiopharmaceutical that selectively binds proteins and cells when oxidized by products of human MPO plus H2O2, but not H2O2 alone, and can be detected using positron emission tomography (PET). Activating HL-60 neutrophil-like human cells with phorbol ester (PMA) caused [18F]4FN retention five-fold over unstimulated cells. An MPO-specific inhibitor (4-ABAH) blocked cellular retention by more than 95%. [18F]4FN PET/CT imaging discriminated inflammatory foci in vivo in three murine models of activated innate immunity: endotoxin-induced toxic shock, PMA-induced contact dermatitis and lipopolysaccharide-induced ankle arthritis. 4-ABAH and Cybb-/- (Nox2-/-) gene deletion strongly abrogated [18F]4FN retention in vivo. Thus, [18F]4FN shows promise as a robust reporter of innate immunity activation by PET/CT.

Study of antioxidant activities, acceptability, and shelf life prediction of Ciplukan (Physalis angulata L.) juice drinks

Antioxidants are free radical scavenger compounds that can reduce or prevent the adverse effects of reactive oxygen species and reactive nitrogen species (ROS/RNS). This study aims to study the antioxidant activity, acceptability, and shelf life of fruit extract drinks and ciplukan buds (Physalis angulata L.). The experimental design used factorial completely randomized design (FCRD), with two factors, namely the ratio of juice extracts and ciplukan buds (a) and stabilizer concentrations (b), three replications. The organoleptic analyzed by hedonic test, antioxidant activity by DPPH method, and total phenolic contents, total flavonoid contents and alkaloid using each of the Folin-Ciocalteau, aluminum chloride and gravimetric methods. The estimated shelf life is calculated using the Arrhenius method at 28°C for 28 days. The results showed that the ratio of fruit extracts and buds (a) affected the preference for taste, aroma, and color attributes. The interaction between juice extract ratio with a bud (a) and stabilizer concentration, (b) influences antioxidant activity, color attributes, and viscosity. The best ciplukan drink samples based on organoleptic calculations are juice with fruit extracts with buds 15:1 and stabilizer concentration of 0.1% (a3b1) and fruit extracts with buds 20:1 and stabilizer concentration of 0.1% (a4b1). The highest total phenolic and flavonoid contents obtained sample code a3b1, while the highest alkaloid content from code sample a4b1. Based on the estimated shelf life of total microbes, ciplukan bud extract has a shelf life of 9 months at 28°C.

Visualizing Peroxynitrite in Microvessels of the Brain with Stroke Using an Engineered Highly Specific Fluorescent Probe

Stroke is one of the leading causes of death and disability in the world, which is associated with malfunction of reactive oxygen species and reactive nitrogen species (ROS/RNS) in cerebral microvessels. In vivo monitoring these species, such as ONOO-, with high selectivity in stroke process is of great significance for early diagnoses and therapies of the disease. Herein, by engineering an indoline-2,3-dione moiety as the recognizing domain, we proposed a novel fluorescence probe Rd-PN2 with highly specific response toward ONOO-, even in the coexistence of other ROS/RNS with high concentration. Rd-PN2 showed high sensitivity and reaction speed in response to ONOO- and exhibited satisfying performances in tracking the endogenously generated ONOO- in living cells and zebrafish. Accordingly, Rd-PN2 can furnish real-time and in vivo visualizing of ONOO- in cerebral microvessels of mice with ischemic and hemorrhagic strokes under two-photon microscopy. This work presented a precisely modulated fluorescence probe for real-time visualizing of ONOO- production in cerebral micovessels, which will also help to acquire more accurate information in the studies of ONOO- functions in the future.

Effect of Nitrosative Stress on the S-Nitroso-Proteome of Paracoccidioides brasiliensis.

The fungi Paracoccidioides brasiliensis and Paracoccidioides lutzii are the causative agents of paracoccidioidomycosis (PCM), a systemic mycosis endemic to Latin America. This fungus is considered a facultative intracellular pathogen that is able to survive and replicate inside macrophages. The survival of the fungus during infection depends on its adaptability to various conditions, such as nitrosative/oxidative stress produced by the host immune cells, particularly alveolar macrophages. Currently, there is little knowledge about the Paracoccidioides spp. signaling pathways involved in the fungus evasion mechanism of the host defense response. However, it is known that some of these pathways are triggered by reactive oxygen species and reactive nitrogen species (ROS/RNS) produced by host cells. Considering that the effects of NO (nitric oxide) on pathogens are concentration dependent, such effects could alter the redox state of cysteine residues by influencing (activating or inhibiting) a variety of protein functions, notably S-nitrosylation, a highly important NO-dependent posttranslational modification that regulates cellular functions and signaling pathways. It has been demonstrated by our group that P. brasiliensis yeast cells proliferate when exposed to low NO concentrations. Thus, this work investigated the modulation profile of S-nitrosylated proteins of P. brasiliensis, as well as identifying S-nitrosylation sites after treatment with RNS. Through mass spectrometry analysis (LC-MS/MS) and label-free quantification, it was possible to identify 474 proteins in the S-nitrosylated proteome study. With this approach, we observed that proteins treated with NO at low concentrations presented a proliferative response pattern, with several proteins involved in cellular cycle regulation and growth being activated. These proteins appear to play important roles in fungal virulence. On the other hand, fungus stimulated by high NO concentrations exhibited a survival response pattern. Among these S-nitrosylated proteins we identified several potential molecular targets for fungal disease therapy, including cell wall integrity (CWI) pathway, amino acid and folic acid metabolisms. In addition, we detected that the transnitrosylation/denitrosylation redox signaling are preserved in this fungus. Finally, this work may help to uncover the beneficial and antifungal properties of NO in the P. brasiliensis and point to useful targets for the development of antifungal drugs.

Effects of Mercury, Lead, Arsenic and Zinc to Human Renal Oxidative Stress and Functions: A Review

Background: Heavy metals such as mercury (Hg), lead (Pb) and arsenic (As) are elements that promote the generation of reactive oxygen species (ROS) and reactive nitrogen species (RNS) involved in the etiology of oxidative stress. They cause oxidative damage to membrane lipids, proteins and DNA, thereby activating pathways of apoptosis and tissue degeneration. Some chemical species derived from these metals include methyl mercury (CH3Hg+), tetraethyl lead [(CH3CH2)4Pb], arsenate (AsO43-) and arsenite (AsO2-), all of which have the capacity to induce oxidative stress and renal damage. Purpose: To comprehensively review the effects of Hg, Pb, As and Zinc (Zn) on renal oxidative status. Methods: Literature survey was done using key words such as heavy metals, oxidative stress and kidney damage using search engines for free scientific publications such as PubMed database, FreeFullPDF.com and Google Scheler. Results: It was revealed that Hg, Pb and As contribute significantly to oxidative stress by stimulating the generation of free radicals, oxidation of biomolecules, deregulation of pro-oxidant proteins, and activation of pro-inflammatory molecules, which ultimately lead to renal damage. There is a strong association between exposure to these heavy metals and chronic renal damage, since their bioaccumulation deregulates glomerular filtration and tubular secretion due to excessive production of ROS and activation of apoptotic pathways. However, studies have shown that Zn possesses renoprotective and antioxidant effects, and its deficiency leads to oxidative stress. Conclusion: The results of this survey suggest that deficiencies of Hg, Pb, As and Zn produce different degrees of oxidative damage which negatively impact on renal health.

Production characteristics of reactive oxygen/nitrogen species in water using atmospheric pressure discharge plasmas

A pulsed discharge, a DC corona discharge, and a plasma jet are separately generated above a water surface, and reactive oxygen species and reactive nitrogen species (ROS/RNS) in the water are investigated. ROS/RNS in water after the sparging of the off-gas of a packed-bed dielectric barrier discharge (PB-DBD) are also investigated. H2O2, NO2−, and NO3− are detected after plasma exposure and only NO3− after off-gas sparging. Short-lifetime species in plasma are found to play an important role in H2O2 and NO2− production and long-lifetime species in NO3− production. NOx may inhibit H2O2 production through OH consumption to produce HNO2 and HNO3. O3 does not contribute to ROS/RNS production. The pulsed plasma exposure is found to be effective for the production of H2O2 and NO2−, and the off-gas sparging of the PB-DBD for the production of NO3−.

L-Glutathione enhances antioxidant capacity of hyaluronic acid and modulates expression of pro-inflammatory cytokines in human fibroblast-like synoviocytes.

Intra-articular injection of hyaluronic acid (HA) has been widely accepted for the treatment of osteoarthritis (OA) in early stage. l-Glutathione (GSH), an antioxidant, has an anti-inflammatory effect on protecting cells from reactive oxygen species and reactive nitrogen species (ROS/RNS). In this study, the therapeutic effects of HA (0.1%) supplemented with GSH (0, 5, 10, and 20% in weight ratios to HA) on human fibroblast-like synoviocytes (FLSs) were evaluated. The results showed that cell morphology and glycosaminoglycan production of FLSs were not changed under treatments. However, the addition of HA + 20% GSH significantly decreased cell survival (p < 0.001) relative to other groups. Relative to un-stimulated FLSs, interleukin-1 beta (IL-1β) stimulation significantly decreased the total antioxidant capacity (p < 0.001) of cells. The antioxidant capacity was restored and the intracellular ROS/RNS was decreased in HA or HA + GSH-treated FLSs. Real-time PCR analysis revealed the mRNA levels of IL-1β, tumor necrosis factor-alpha, and matrix metalloproteinase-3 were down-regulated significantly (all p < 0.05) when FLSs cultured in HA or HA + GSH. IL-6 mRNA expressions were down-regulated significantly in HA and HA + 5% GSH groups (both p < 0.05) but up-regulated when HA supplemented with 10% and 20% GSH (both p < 0.01). In addition, the protein levels of IL-1β were further decreased with significant differences (both p < 0.05) in the HA + 10% GSH and HA + 20% GSH groups when compared to FLSs cultured in normal medium. In conclusion, HA supplemented with GSH improves antioxidant capacity and modulates pro-inflammatory cytokines expressions in FLSs. GSH has the potential to augment the effect of viscosupplementation using HA on OA patients. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2071-2079, 2016.

Cytoprotective Role of Nrf2 in Electrical Pulse Stimulated C2C12 Myotube.

Regular physical exercise is central to a healthy lifestyle. However, exercise-related muscle contraction can induce reactive oxygen species and reactive nitrogen species (ROS/RNS) production in skeletal muscle. The nuclear factor-E2-related factor-2 (Nrf2) transcription factor is a cellular sensor for oxidative stress. Regulation of nuclear Nrf2 signaling regulates antioxidant responses and protects organ structure and function. However, the role of Nrf2 in exercise- or contraction-induced ROS/RNS production in skeletal muscle is not clear. In this study, using differentiated C2C12 cells and electrical pulse stimulation (EPS) of muscle contraction, we explored whether Nrf2 plays a role in the skeletal muscle response to muscle contraction-induced ROS/RNS. We found that EPS (40 V, 1 Hz, 2 ms) stimulated ROS/RNS accumulation and Nrf2 activation. We also showed that expression of NQO1, HO-1 and GCLM increased after EPS-induced muscle contraction and was remarkably suppressed in cells with Nrf2 knockdown. We also found that the antioxidant N-acetylcysteine (NAC) significantly attenuated Nrf2 activation after EPS, whereas the nitric oxide synthetase inhibitor Nω-nitro-L-arginine methyl ester (L-NAME) did not. Furthermore, Nrf2 knockdown after EPS markedly decreased ROS/RNS redox potential and cell viability and increased expression of the apoptosis marker Annexin V in C2C12 myotubes. These results indicate that Nrf2 activation and expression of Nrf2 regulated-genes protected muscle against the increased ROS caused by EPS-induced muscle contraction. Thus, our findings suggest that Nrf2 may be a key factor for preservation of muscle function during muscle contraction.

Redox Control of Platelet Functions in Physiology and Pathophysiology

An imbalance between the production and the detoxification of reactive oxygen species and reactive nitrogen species (ROS/RNS) can be implicated in many pathological processes. Platelets are best known as primary mediators of hemostasis and can be either targets of ROS/RNS or generate radicals during cell activation. These conditions can dramatically affect platelet physiology, leading even, as an ultimate event, to the cell number modification. In this case, pathological conditions such as thrombocytosis (promoted by increased cell number) or thrombocytopenia and myelodysplasia (promoted by cell decrease mediated by accelerated apoptosis) can occur. Usually, in peripheral blood, ROS/RNS production is balanced by the rate of oxidant elimination. Under this condition, platelets are in a nonadherent "resting" state. During endothelial dysfunction or under pathological conditions, ROS/RNS production increases and the platelets respond with specific biochemical and morphologic changes. Mitochondria are at the center of these processes, being able to both generate ROS/RNS, that drive redox-sensitive events, and respond to ROS/RNS-mediated changes of the cellular redox state. Irregular function of platelets and enhanced interaction with leukocytes and endothelial cells can contribute to pathogenesis of atherosclerotic and thrombotic events. The relationship between oxidative stress, platelet death, and the activation-dependent pathways that drive platelet pro-coagulant activity is unclear and deserves to be explored. Expanding knowledge about how platelets can mediate hemostasis and modulate inflammation may lead to novel and effective therapeutic strategies for the long and growing list of pathological conditions that involve both thrombosis and inflammation.

Exploiting Oxidative Microenvironments in the Body as Triggers for Drug Delivery Systems

Reactive oxygen species and reactive nitrogen species (ROS/RNS) play an important role in cell signaling pathways. However, the increased production of these species may disrupt cellular homeostasis, giving rise to pathological conditions. Biomaterials that are responsive to ROS/RNS can be strategically used to specifically release therapeutics and diagnostic agents to regions undergoing oxidative stress. Many nanocarriers intended to exploit redox micro-environments as triggers for drug release, summarized and compared in this review, have recently been developed. We describe these carriers' chemical structures, strategies for payload protection and oxidation-selective release, and ROS/RNS sensitivity as tested in initial studies. ROS/RNS are unstable, so reliable measures of their concentrations in various conditions are scarce. Combined with the dearth of materials shown to respond to physiologically relevant levels of ROS/RNS, evaluations of their true sensitivity are difficult. Oxidation-responsive nanocarriers developed thus far show tremendous potential for applicability in vivo; however, the sensitivity of these chemistries needs to be fine tuned to enable responses to physiological levels of ROS and RNS.

Antioxidant properties of Neu2000 on mitochondrial free radicals and oxidative damage

Neu2000 [2-hydroxy-5-(2,3,5,6-tetrafluoro-4 trifluoromethylbenzylamino) benzoic acid] is a dual-acting neuroprotective agent that functions both as a noncompetitive N-methyl-d-aspartate (NMDA) receptor antagonist and a free radical scavenger. In the present study, we investigated the scavenging activity of Neu2000 on various classes of reactive oxygen species and reactive nitrogen species (ROS/RNS) as well as its efficacy for reducing free radicals and oxidative stress/damage induced in spinal cord mitochondrial preparations. Neu2000 exerted scavenging activity against superoxide, nitric oxide, and hydroxyl radicals, and efficiently scavenged peroxynitrite. In the mitochondrial studies, Neu2000 markedly inhibited ROS/RNS and hydrogen peroxide levels following antimycin treatment. In addition, Neu2000 effectively scavenged hydroxyl radicals generated by iron(III)-ascorbate, reduced protein carbonyl formation mediated by hydroxyl radicals and peroxynitrite, and prevented glutathione oxidation caused by tert-butyl hydroperoxide in isolated mitochondria. Interestingly, incubation of isolated mitochondria with Neu2000 followed by centrifugation and removal of the supernatant also resulted in a concentration-dependent decrease in lipid peroxidation. This observation suggests that Neu2000 enters mitochondria to target free radicals or indirectly affects mitochondrial function in a manner that promotes antioxidant activity. The results of the present study demonstrate that Neu2000 possesses potent in vitro antioxidant activity due, most likely, to its active phenoxy group.