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1.
Differential antibacterial control by neutrophil subsets.
Leliefeld, PHC, Pillay, J, Vrisekoop, N, Heeres, M, Tak, T, Kox, M, Rooijakkers, SHM, Kuijpers, TW, Pickkers, P, Leenen, LPH, et al
Blood advances. 2018;(11):1344-1355
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Abstract
Neutrophils comprise a heterogeneous population of cells essential for bacterial eradication, and defects in neutrophil function are associated with increased susceptibility to infection. In this study, neutrophils from healthy controls were shown to prevent bacterial proliferation for at least 48 hours when cocultured with methicillin-resistant Staphylococcus aureus (MRSA) in tissue-like scaffolds by establishing a bacteriostatic environment inside their phagolysosome. This intracellular bacterial containment is independent of reactive oxygen species because neutrophils that lack a functional nicotinamide adenine dinucleotide phosphate-oxidase complex displayed no defect in intracellular bacterial containment, whereas killing of the pathogen was impaired. During acute inflammation, a subset of CD16bright/CD62Ldim hypersegmented neutrophils displayed normal phagocytosis associated with a remarkably poor capacity to contain bacteria intracellularly. Conversely, CD16dim-banded neutrophils were the only neutrophil subset that adequately contained MRSA. These findings demonstrate a clear neutrophil heterogeneity in their antimicrobial capacity and the appearance of neutrophil subsets with a clear differentiation in functionality during acute inflammation. Furthermore, this study provides an evolutionary basis for the rapid release of banded neutrophils into the circulation during acute inflammation.
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2.
SK3 channel and mitochondrial ROS mediate NADPH oxidase-independent NETosis induced by calcium influx.
Douda, DN, Khan, MA, Grasemann, H, Palaniyar, N
Proceedings of the National Academy of Sciences of the United States of America. 2015;(9):2817-22
Abstract
Neutrophils cast neutrophil extracellular traps (NETs) to defend the host against invading pathogens. Although effective against microbial pathogens, a growing body of literature now suggests that NETs have negative impacts on many inflammatory and autoimmune diseases. Identifying mechanisms that regulate the process termed "NETosis" is important for treating these diseases. Although two major types of NETosis have been described to date, mechanisms regulating these forms of cell death are not clearly established. NADPH oxidase 2 (NOX2) generates large amounts of reactive oxygen species (ROS), which is essential for NOX-dependent NETosis. However, major regulators of NOX-independent NETosis are largely unknown. Here we show that calcium activated NOX-independent NETosis is fast and mediated by a calcium-activated small conductance potassium (SK) channel member SK3 and mitochondrial ROS. Although mitochondrial ROS is needed for NOX-independent NETosis, it is not important for NOX-dependent NETosis. We further demonstrate that the activation of the calcium-activated potassium channel is sufficient to induce NOX-independent NETosis. Unlike NOX-dependent NETosis, NOX-independent NETosis is accompanied by a substantially lower level of activation of ERK and moderate level of activation of Akt, whereas the activation of p38 is similar in both pathways. ERK activation is essential for the NOX-dependent pathway, whereas its activation is not essential for the NOX-independent pathway. Despite the differential activation, both NOX-dependent and -independent NETosis require Akt activity. Collectively, this study highlights key differences in these two major NETosis pathways and provides an insight into previously unknown mechanisms for NOX-independent NETosis.
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β-Carotene, α-tocopherol and ascorbic acid: differential profile of antioxidant, inflammatory status and regulation of gene expression in human mononuclear cells of diabetic donors.
de Oliveira, BF, Costa, DC, Nogueira-Machado, JA, Chaves, MM
Diabetes/metabolism research and reviews. 2013;(8):636-45
Abstract
BACKGROUND Diabetic patients are exposed to increased oxidative stress due to several mechanisms, mainly hyperglycaemia. Pathological processes, such as those in type 1 diabetes, include diminished activity of the antioxidant defense system(s) or excessive oxidative generation resulting in an oxidative/antioxidant imbalance and development of oxidative stress. METHODS The purpose of this study was to evaluate the production of reactive oxygen species (ROS) (chemiluminescence) and reduction capacity (MTT dye reduction), the expression of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase subunits, superoxide dismutase and catalase using quantitative reverse-transcriptase polymerase chain reaction, and the levels of cytokines [interleukin (IL)-6, tumour necrosis factor-α, IL-8, IL-10 and IL-4] by sandwich enzyme-linked immunosorbent assay in mononuclear cells from non-diabetic and diabetic donors treated with a vitamin complex (ascorbic acid, β-carotene and α-tocopherol) in two different concentrations ([A] = ascorbic acid = 0.08 µM, α-tocopherol = 0.04 µM, β-carotene = 0.0008 µM and [20A] = ascorbic acid = 1.6 µM, α-tocopherol = 0.82 µM, β-carotene = 0.016 µM). RESULTS Concentration [A] was antioxidant reducing ROS production, expression of NADPH oxidase subunits and pro-inflammatory cytokines while raising the expression of antioxidant enzymes and reducing pro-inflammatory cytokines in both groups. Concentration [20A] was pro-oxidant by raising ROS production, NADPH oxidase subunits and pro-inflammatory cytokines and reducing antioxidant enzymes and anti-inflammatory cytokines in the non-diabetic group but antioxidant in cells of type 1 diabetic patients by raising antioxidant enzymes and anti-inflammatory cytokines and reducing pro-inflammatory cytokines. CONCLUSION The vitamin complex has a dual effect, pro-oxidant and antioxidant, being also dose dependent with different profiles of cells of non-diabetic and type 1 diabetic patients.
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4.
Apocynin reduces reactive oxygen species concentrations in exhaled breath condensate in asthmatics.
Stefanska, J, Sarniak, A, Wlodarczyk, A, Sokolowska, M, Pniewska, E, Doniec, Z, Nowak, D, Pawliczak, R
Experimental lung research. 2012;(2):90-9
Abstract
Asthma is an inflammatory airway disease, and oxidative stress was proven to be involved in its pathogenesis. Apocynin effectively inhibits the main source of reactive oxygen species (ROS)-nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-by blocking its activation. The aim of this study was to investigate the effect of inhaled apocynin on ROS and RNS (reactive nitrogen species) concentration in 14 nonsmoking mild asthmatics. Effects of nebulized apocynin (0.5 mg/mL) were assessed in exhaled breath condensate (EBC) after 30, 60, and 120 minutes, and safety parameters have been analyzed. Apocynin significantly decreased H2O2 concentration in EBC in comparison with placebo after 60 and 120 minutes. Moreover, apocynin significantly reduced NO(-2) concentration 30 and 60 minutes after nebulization and caused a significant decrease of NO(-3) concentration in EBC 60 and 120 minutes after administration, comparing with placebo. No adverse events have been observed throughout the study. This research confirmed anti-inflammatory properties of nebulized apocynin, which might be an effective and safe drug in bronchial asthma.
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5.
Liver X receptor agonist alleviated high glucose-induced endothelial progenitor cell dysfunction via inhibition of reactive oxygen species and activation of AMP-activated protein kinase.
Li, X, Song, Y, Han, Y, Wang, D, Zhu, Y
Microcirculation (New York, N.Y. : 1994). 2012;(6):547-53
Abstract
OBJECTIVE Liver X receptors (LXRs) are key regulators of cholesterol homeostasis. Synthetic LXR agonists are anti-atherogenic and anti-inflammatory. However, the effect of LXR agonists on endothelial progenitor cell (EPC) function is largely unknown. Here, we explored the effect of the LXR agonist TO901317 (TO) on EPC biology and the underlying mechanisms. METHODS Endothelial progenitor cells were cultured in mannitol or 30 mm glucose (high glucose) for 24 hours. For TO treatments, cells were pretreated with TO (10 μm) for 12 hours, then mannitol or high glucose was added for an additional 24 hours. EPCs function, reactive oxygen species (ROS) release, and phosphorylation of adenosine monophosphate-activated protein kinase (AMPK) were analyzed. RESULTS TO could restore the high glucose-impaired adhesion and migration capacity of EPCs. High glucose impaired EPC-mediated angiogenesis, and TO reversed the impairment. TO also alleviated ROS release induced by high glucose. Western blot analysis revealed that high glucose downregulated the phosphorylation of AMPK and endothelial nitric oxide synthase, which could be reversed with TO treatment. Furthermore, inhibiting AMPK activation by compound C could abolish the protective effects of TO on EPCs. CONCLUSIONS TO had a protective effect on EPCs under high glucose by inhibiting ROS release and activating AMPK.
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Assessment of a standardized ROS production profile in humans by electron paramagnetic resonance.
Mrakic-Sposta, S, Gussoni, M, Montorsi, M, Porcelli, S, Vezzoli, A
Oxidative medicine and cellular longevity. 2012;:973927
Abstract
Despite the growing interest in the role of reactive oxygen species (ROS) in health and disease, reliable quantitative noninvasive methods for the assessment of oxidative stress in humans are still lacking. EPR technique, coupled to a specific spin probe (CMH: 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine) is here presented as the method of choice to gain a direct measurement of ROS in biological fluids and tissues. The study aimed at demonstrating that, differently from currently available "a posteriori" assays of ROS-induced damage by means of biomolecules (e.g., proteins and lipids) spin-trapping EPR provides direct evidence of the "instantaneous" presence of radical species in the sample and, as signal areas are proportional to the number of excited electron spins, lead to absolute concentration levels. Using a recently developed bench top continuous wave system (e-scan EPR scanner, Bruker) dealing with very low ROS concentration levels in small (50 μL) samples, we successfully monitored rapid ROS production changes in peripheral blood of athletes after controlled exercise and sedentary subjects after antioxidant supplementation. The correlation between EPR results and data obtained by various enzymatic assays (e.g., protein carbonyls and thiobarbituric acid reactive substances) was determined too. Synthetically, our method allows reliable, quick, noninvasive quantitative determination of ROS in human peripheral blood.
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7.
Diaphragmatic breathing reduces postprandial oxidative stress.
Martarelli, D, Cocchioni, M, Scuri, S, Pompei, P
Journal of alternative and complementary medicine (New York, N.Y.). 2011;(7):623-8
Abstract
OBJECTIVES A number of studies suggest that postprandial hyperglycemia produces oxidative stress, leading to complications associated with diabetes. However, hyperglycemia-induced oxidative stress may affect groups of people other than diabetics, such as smokers and athletes with specific diet plans. Based on previous reports that seated breathing meditation reduces hyperglycemia, the present study was designed to determine the effects of diaphragmatic breathing on postprandial plasma glycemia, insulin, oxidative stress, and antioxidant levels in athletes with normal glucose metabolism. DESIGN Data collected before and after consumption of a 900-calorie breakfast composed of 80% carbohydrates, 10% proteins, and 10% lipids were analyzed. Ten (10) minutes after the meal, 8 subjects spent 40 minutes performing diaphragmatic breathing in a quiet place. The other 8 subjects, representing the control group, spent the same time sitting in an equivalent quiet place reading a magazine. SUBJECTS Data from 16 amateur male cyclists age 30.12±4.9 years (±SD) were analyzed. Their mean height and weight were 177.81±5.3 cm and 71.40±5.2 kg, respectively. All subjects underwent a physical examination and were determined to be in good health. OUTCOME MEASURES Blood samples were collected immediately before the meal as well as 1 hour and 2 hours after the meal, and plasma levels of glucose, insulin, reactive oxygen metabolites, and biologic antioxidant potential were determined. Heart rate was also recorded. RESULTS Results show that in normal subjects, acute hyperglycemia induces free-radical production while reducing the antioxidant levels (p<0.05). Diaphragmatic breathing reduces heart rates (p<0.01), increases insulin (p<0.05), reduces glycemia (p<0.01), and reduces free-radical production as indicated by the higher antioxidants levels (p<0.05). CONCLUSIONS Diaphragmatic breathing, likely through the activation of the parasympathetic nervous system, increases insulin, reduces glycemia, and reduces reactive oxygen species production.
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Non-transferrin bound iron, cytokine activation and intracellular reactive oxygen species generation in hemodialysis patients receiving intravenous iron dextran or iron sucrose.
Pai, AB, Conner, T, McQuade, CR, Olp, J, Hicks, P
Biometals : an international journal on the role of metal ions in biology, biochemistry, and medicine. 2011;(4):603-13
Abstract
Intravenous (IV) iron supplementation is widely used to support erythropoeisis in hemodialysis patients. IV iron products are associated with oxidative stress that has been measured principally by circulating biomarkers such as products of lipid peroxidation. The pro-oxidant effects of IV iron are presumed to be due at least in part, by free or non-transferrin bound iron (NTBI). However, the effects of IV iron on intracellular redox status and downstream effectors is not known. This prospective, crossover study compared cytokine activation, reactive oxygen species generation and oxidative stress after single IV doses of iron sucrose and iron dextran. This was a prospective, open-label, crossover study. Ten patients with end-stage renal disease (ESRD) on hemodialysis and four age and sex-matched healthy were assigned to receive 100 mg of each IV iron product over 5 min in random sequence with a 2 week washout between products. Subjects were fasted and fed a low iron diet in the General Clinical Research Center at the University of New Mexico. Serum and plasma samples for IL-1, IL-6, TNF-α and IL-10 and NTBI were obtained at baseline, 60 and 240 min after iron infusion. Peripheral blood mononuclear cells (PBMC) were isolated at the same time points and stained with fluorescent probes to identify intracellular reactive oxygen species and mitochondrial membrane potential (Δψm) by flow cytometry. Lipid peroxidation was assessed by plasma F(2) isoprostane concentration. Mean ± SEM maximum serum NTBI values were significantly higher among patients receiving IS compared to ID (2.59 ± 0.31 and 1.0 ± 0.36 µM, respectively, P = 0.005 IS vs. ID) Mean ± SEM NTBI area under the serum concentration-time curve (AUC) was 3-fold higher after IS versus ID (202 ± 53 vs. 74 ± 23 µM*min/l, P = 0.04) in ESRD patients, indicating increased exposure to NTBI. IV iron administration was associated with increased pro-inflammatory cytokines. Serum IL-6 concentrations increased most profoundly, with a 2.6 and 2.1 fold increase from baseline in ESRD patients given IS and ID, respectively (P < 0.05 compared to baseline). In healthy controls, serum IL-6 was undetectable at baseline and after IV iron administration. Most ESRD patients had increased intracellular ROS generation, however, there was no difference between ID and IS. Only one healthy control had increased ROS generation post IV iron. All healthy controls experienced a loss of Δψm (100% with IS and 50% with ID). ESRD patients also had loss of Δψm with a nadir at 240 min. IS administration was associated with higher maximum serum NTBI concentrations compared to ID, however, the both compounds produced similar ROS generation and cytokine activation that was more pronounced among ESRD patients. The effect of IV iron-induced ROS production on pivotal signaling pathways needs to be explored.
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9.
Antioxidant activity of the new black vinegar "IZUMI".
Nagashima, M, Saito, K
The journal of nutrition, health & aging. 2010;(10):845-9
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Abstract
OBJECTIVES "IZUMI" is a new kind of vinegar resulting from an improvement in the manufacturing method of Kurosu, a traditional vinegar product made from unpolished rice. We evaluated the antioxidant activity of this new Kurosu by means of measuring the level of diacron-reactive oxygen metabolites (d-ROM), the biological antioxidant potential (BAP), as well as RBC deformability using the microchannel array flow method. PARTICIPANTS Ten healthy, untrained female volunteers participated in this study. MEASUREMENTS All subjects drank 50 ml of "IZUMI" on a daily basis, and blood samples were collected pre-"IZUMI" (I), after one month "IZUMI" consumption (II), and after two months "IZUMI" consumption (III). The subjects continuously wore a lifecorder during a 7-day period and the nutritional intake was measured before the initial blood sample collection. RESULTS There were no significant changes in weight, BMI, fat mass, or fat-free mass. There were no significant differences in daily energy consumption, physical activity and nutritional intake. Peripheral blood variables did not change significantly. The drinking of " IZUMI " increased serum BAP level gradually, and after 30 days it was significantly higher as compared to the pre-drinking level. The serum level of d-ROM and blood filtration time (BFT) decreased by drinking "IZUMI"; with d-ROM significantly lower than the pre-drinking level after 30 days and BFT significantly decreased after 60 days (all P < 0.05). CONCLUSIONS These results suggest that "IZUMI", a Kurosu containing a higher level of amino acids, increases antioxidant activity and reduces oxidative stress and blood filtration time in female subjects.
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Effects of the 'live high-train low' method on prooxidant/antioxidant balance on elite athletes.
Pialoux, V, Mounier, R, Rock, E, Mazur, A, Schmitt, L, Richalet, JP, Robach, P, Brugniaux, J, Coudert, J, Fellmann, N
European journal of clinical nutrition. 2009;(6):756-62
Abstract
BACKGROUND/OBJECTIVES We previously demonstrated that acute exposure to hypoxia (3 h at 3000 m) increased oxidative stress markers. Thus, by using the 'living high-training low' (LHTL) method, we further hypothesized that intermittent hypoxia associated with endurance training alters the prooxidant/antioxidant balance. SUBJECTS/METHODS Twelve elite athletes from the Athletic French Federation were subjected to 18-day endurance training. They were divided into two groups: one group (control group) trained at 1200 m and lived in hypoxia (2500-3000 m simulated altitude) and the second group trained and lived at 1200 m. The subjects performed an acute hypoxic test (10 min at 4800 m) before and immediately after the training. Plasma levels of advanced oxidation protein products (AOPP), malondialdehydes (MDA), ferric-reducing antioxidant power (FRAP), lipid-soluble antioxidants normalized for triacylglycerols, and cholesterol and retinol were measured before and after the 4800 m tests. RESULTS After the training, MDA and AOPP concentrations were decreased in response to the 4800 m test only for the control group. Eighteen days of LHTL induced a significant decrease of all antioxidant markers (FRAP, P=0.01; alpha-tocopherol, P=0.04; beta-carotene, P=0.01 and lycopene, P=0.02) for the runners. This imbalance between antioxidant and prooxidant might result from insufficient intakes in vitamins A and E. CONCLUSIONS The LHTL model characterized by the association of aerobic exercises and intermittent resting hypoxia exposures decreased the antioxidant status whereas the normoxic endurance training induced preconditioning mechanisms in response to the 4800 m test.