Science.gov

Sample records for high-isoelectric-point superoxide dismutase1cw

  1. Downregulation of high-isoelectric-point extracellular superoxide dismutase mediates alterations in the metabolism of reactive oxygen species and developmental disturbances in hybrid aspen.

    PubMed

    Srivastava, Vaibhav; Schinkel, Helga; Witzell, Johanna; Hertzberg, Magnus; Torp, Mikaela; Srivastava, Manoj Kumar; Karpinska, Barbara; Melzer, Michael; Wingsle, Gunnar

    2007-01-01

    Transgenic hybrid aspen (Populus tremula L. x P. tremuloides Michx.) plants expressing a high-isoelectric-point superoxide dismutase (hipI-SOD) gene in antisense orientation were generated to investigate its function. Immunolocalization studies showed the enzyme to be localized extracellularly, in the secondary cell wall of xylem vessels and phloem fibers. The antisense lines of hipI-SOD exhibited a distinct phenotype; growth rate was reduced, stems were thinner and leaves smaller than in wild-type (WT) plants. The abundance of hipI-SOD was reduced in the bark and xylem of plants from these antisense lines. The vascular tissue of transgenic lines became lignified earlier than in WT plants and also showed an increased accumulation of reactive oxygen species (ROS). Xylem fibers and vessels were shorter and thinner in the transgenic lines than in WT plants. The total phenolic content was enhanced in the antisense lines. Furthermore, microarray analysis indicated that several enzymes involved in cell signaling, lignin biosynthesis and stress responses were upregulated in apical vascular tissues of transgenic plants. The upregulation of selected genes involved in lignin biosynthesis was also verified by real-time PCR. The results suggest that, in the transgenic plants, a premature transition into maturation occurs and the process is discussed in terms of the effects of increased accumulation of ROS due to reduced expression of hipI-SOD during development and differentiation.

  2. Alternative Splicing Studies of the Reactive Oxygen Species Gene Network in Populus Reveal Two Isoforms of High-Isoelectric-Point Superoxide Dismutase1[C][W

    PubMed Central

    Srivastava, Vaibhav; Srivastava, Manoj Kumar; Chibani, Kamel; Nilsson, Robert; Rouhier, Nicolas; Melzer, Michael; Wingsle, Gunnar

    2009-01-01

    Recent evidence has shown that alternative splicing (AS) is widely involved in the regulation of gene expression, substantially extending the diversity of numerous proteins. In this study, a subset of expressed sequence tags representing members of the reactive oxygen species gene network was selected from the PopulusDB database to investigate AS mechanisms in Populus. Examples of all known types of AS were detected, but intron retention was the most common. Interestingly, the closest Arabidopsis (Arabidopsis thaliana) homologs of half of the AS genes identified in Populus are not reportedly alternatively spliced. Two genes encoding the protein of most interest in our study (high-isoelectric-point superoxide dismutase [hipI-SOD]) have been found in black cottonwood (Populus trichocarpa), designated PthipI-SODC1 and PthipI-SODC2. Analysis of the expressed sequence tag libraries has indicated the presence of two transcripts of PthipI-SODC1 (hipI-SODC1b and hipI-SODC1s). Alignment of these sequences with the PthipI-SODC1 gene showed that hipI-SODC1b was 69 bp longer than hipI-SODC1s due to an AS event involving the use of an alternative donor splice site in the sixth intron. Transcript analysis showed that the splice variant hipI-SODC1b was differentially expressed, being clearly expressed in cambial and xylem, but not phloem, regions. In addition, immunolocalization and mass spectrometric data confirmed the presence of hipI-SOD proteins in vascular tissue. The functionalities of the spliced gene products were assessed by expressing recombinant hipI-SOD proteins and in vitro SOD activity assays. PMID:19176719

  3. pH-Responsive poly(itaconic acid-co-N-vinylpyrrolidone) hydrogels with reduced ionic strength loading solutions offer improved oral delivery potential for high isoelectric point-exhibiting therapeutic proteins

    PubMed Central

    Koetting, Michael C.; Peppas, Nicholas A.

    2014-01-01

    pH-Responsive hydrogels comprised of itaconic acid copolymerized with N-vinylpyrrolidone (P(IA-co-NVP)) were synthesized and tested as carriers for the oral delivery of high isoelectric point (pI) exhibiting therapeutic proteins. Swelling studies show that P(IA-co-NVP) hydrogels exhibit significantly greater and faster pH-responsive swelling than previously studied methacrylic acid-based hydrogels, achieving up to 68% greater equilibrium swelling and 10.4 times greater swelling in time-limited experiments. Using salmon calcitonin as a model high pI protein therapeutic, we show that P(IA-co-NVP) hydrogels exhibit significantly greater delivery potential than methacrylic acid-based hydrogels. Additionally, we show that utilizing a lower ionic strength solution during drug loading significantly improves drug delivery potential for high pI therapeutics. By using a 1.5 mM PBS buffer rather than the standard 150 mM PBS buffer during loading, up to 83 times as much calcitonin can be delivered in neutral conditions, with up to a 9.6 fold improvement in percent release. Using P(IA-co-NVP) hydrogel microparticles and a low ionic strength loading solution, up to 48 μg calcitonin/mg hydrogel can be delivered in small intestinal conditions. Based on expected absorption in the small intestine, this is sufficient delivery potential for achieving therapeutic dosage via a single, regularly-sized pill taken daily. PMID:24853463

  4. Student award for outstanding research winner in the Ph.D. category for the 2017 society for biomaterials annual meeting and exposition, april 5-8, 2017, Minneapolis, Minnesota: Characterization of protein interactions with molecularly imprinted hydrogels that possess engineered affinity for high isoelectric point biomarkers.

    PubMed

    Clegg, John R; Zhong, Justin X; Irani, Afshan S; Gu, Joann; Spencer, David S; Peppas, Nicholas A

    2017-06-01

    Molecularly imprinted polymers (MIPs) with selective affinity for protein biomarkers could find extensive utility as environmentally robust, cost-efficient biomaterials for diagnostic and therapeutic applications. In order to develop recognitive, synthetic biomaterials for prohibitively expensive protein biomarkers, we have developed a molecular imprinting technique that utilizes structurally similar, analogue proteins. Hydrogel microparticles synthesized by molecular imprinting with trypsin, lysozyme, and cytochrome c possessed an increased affinity for alternate high isoelectric point biomarkers both in isolation and plasma-mimicking adsorption conditions. Imprinted and non-imprinted P(MAA-co-AAm-co-DEAEMA) microgels containing PMAO-PEGMA functionalized polycaprolactone nanoparticles were net-anionic, polydisperse, and irregularly shaped. MIPs and control non-imprinted polymers (NIPs) exhibited regions of Freundlich and BET isotherm adsorption behavior in a range of non-competitive protein solutions, where MIPs exhibited enhanced adsorption capacity in the Freundlich isotherm regions. In a competitive condition, imprinting with analogue templates (trypsin, lysozyme) increased the adsorption capacity of microgels for cytochrome c by 162% and 219%, respectively, as compared to a 122% increase provided by traditional bulk imprinting with cytochrome c. Our results suggest that molecular imprinting with analogue protein templates is a viable synthetic strategy for enhancing hydrogel-biomarker affinity and promoting specific protein adsorption behavior in biological fluids. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1565-1574, 2017. © 2017 Wiley Periodicals, Inc.

  5. Air Revitalization Using Superoxides

    NASA Technical Reports Server (NTRS)

    Wydeven, Theodore; Wood, Peter C.; Spitze, L. A.

    1988-01-01

    Pellets made from powder mixtures of potassium superoxide, KO2, and calcium superoxide, Ca(O2)2, proven markedly superior to pellets of pure KO2 for adding O2 to and removing CO2 from atmospheric-pressure flow of humidified CO2 in He. Superoxides used extensively to supply O2 and scrub CO2 in variety of ambient-pressure life-support applications, including portable self-contained breathing apparatuses, spacecraft, and undersea submersible craft.

  6. Superoxide flashes in single mitochondria.

    PubMed

    Wang, Wang; Fang, Huaqiang; Groom, Linda; Cheng, Aiwu; Zhang, Wanrui; Liu, Jie; Wang, Xianhua; Li, Kaitao; Han, Peidong; Zheng, Ming; Yin, Jinhu; Wang, Weidong; Mattson, Mark P; Kao, Joseph P Y; Lakatta, Edward G; Sheu, Shey-Shing; Ouyang, Kunfu; Chen, Ju; Dirksen, Robert T; Cheng, Heping

    2008-07-25

    In quiescent cells, mitochondria are the primary source of reactive oxygen species (ROS), which are generated by leakiness of the electron transport chain (ETC). High levels of ROS can trigger cell death, whereas lower levels drive diverse and important cellular functions. We show here by employing a newly developed mitochondrial matrix-targeted superoxide indicator, that individual mitochondria undergo spontaneous bursts of superoxide generation, termed "superoxide flashes." Superoxide flashes occur randomly in space and time, exhibit all-or-none properties, and provide a vital source of superoxide production across many different cell types. Individual flashes are triggered by transient openings of the mitochondrial permeability transition pore stimulating superoxide production by the ETC. Furthermore, we observe a flurry of superoxide flash activity during reoxygenation of cardiomyocytes after hypoxia, which is inhibited by the cardioprotective compound adenosine. We propose that superoxide flashes could serve as a valuable biomarker for a wide variety of oxidative stress-related diseases.

  7. Synthesis of calcium superoxide

    NASA Technical Reports Server (NTRS)

    Rewick, R. T.; Blucher, W. G.; Estacio, P. L.

    1972-01-01

    Efforts to prepare Ca(O2) sub 2 from reactions of calcium compounds with 100% O3 and with O(D-1) atoms generated by photolysis of O3 at 2537 A are described. Samples of Ca(OH) sub 2, CaO, CaO2, Ca metal, and mixtures containing suspected impurities to promote reaction have been treated with excess O3 under static and flow conditions in the presence and absence of UV irradiation. Studies with KO2 suggest that the superoxide anion is stable to radiation at 2537 A but reacts with oxygen atoms generated by the photolysis of O3 to form KO3. Calcium superoxide is expected to behave in an analogous.

  8. Superoxide Dismutase Assays

    DTIC Science & Technology

    1987-06-16

    rapidly catalyzed the dismutation of Superoxide free radical anions in the reaction : enzyme 0 * + 0’~ + 2H > O...its rate of dismutation both enzymically and spontaneously. By allowing the generator reaction to proceed for a specific time period before the addi...concentration is adjusted to give a reaction rate so that the change at 550 rm is 0.02 to 0.04 OD/min. Stock of Xanthine Oxidase BC 1.1.3.22 is at

  9. OnPLS integration of transcriptomic, proteomic and metabolomic data shows multi-level oxidative stress responses in the cambium of transgenic hipI- superoxide dismutase Populus plants

    PubMed Central

    2013-01-01

    Background Reactive oxygen species (ROS) are involved in the regulation of diverse physiological processes in plants, including various biotic and abiotic stress responses. Thus, oxidative stress tolerance mechanisms in plants are complex, and diverse responses at multiple levels need to be characterized in order to understand them. Here we present system responses to oxidative stress in Populus by integrating data from analyses of the cambial region of wild-type controls and plants expressing high-isoelectric-point superoxide dismutase (hipI-SOD) transcripts in antisense orientation showing a higher production of superoxide. The cambium, a thin cell layer, generates cells that differentiate to form either phloem or xylem and is hypothesized to be a major reason for phenotypic perturbations in the transgenic plants. Data from multiple platforms including transcriptomics (microarray analysis), proteomics (UPLC/QTOF-MS), and metabolomics (GC-TOF/MS, UPLC/MS, and UHPLC-LTQ/MS) were integrated using the most recent development of orthogonal projections to latent structures called OnPLS. OnPLS is a symmetrical multi-block method that does not depend on the order of analysis when more than two blocks are analysed. Significantly affected genes, proteins and metabolites were then visualized in painted pathway diagrams. Results The main categories that appear to be significantly influenced in the transgenic plants were pathways related to redox regulation, carbon metabolism and protein degradation, e.g. the glycolysis and pentose phosphate pathways (PPP). The results provide system-level information on ROS metabolism and responses to oxidative stress, and indicate that some initial responses to oxidative stress may share common pathways. Conclusion The proposed data evaluation strategy shows an efficient way of compiling complex, multi-platform datasets to obtain significant biological information. PMID:24341908

  10. OnPLS integration of transcriptomic, proteomic and metabolomic data shows multi-level oxidative stress responses in the cambium of transgenic hipI- superoxide dismutase Populus plants.

    PubMed

    Srivastava, Vaibhav; Obudulu, Ogonna; Bygdell, Joakim; Löfstedt, Tommy; Rydén, Patrik; Nilsson, Robert; Ahnlund, Maria; Johansson, Annika; Jonsson, Pär; Freyhult, Eva; Qvarnström, Johanna; Karlsson, Jan; Melzer, Michael; Moritz, Thomas; Trygg, Johan; Hvidsten, Torgeir R; Wingsle, Gunnar

    2013-12-17

    Reactive oxygen species (ROS) are involved in the regulation of diverse physiological processes in plants, including various biotic and abiotic stress responses. Thus, oxidative stress tolerance mechanisms in plants are complex, and diverse responses at multiple levels need to be characterized in order to understand them. Here we present system responses to oxidative stress in Populus by integrating data from analyses of the cambial region of wild-type controls and plants expressing high-isoelectric-point superoxide dismutase (hipI-SOD) transcripts in antisense orientation showing a higher production of superoxide. The cambium, a thin cell layer, generates cells that differentiate to form either phloem or xylem and is hypothesized to be a major reason for phenotypic perturbations in the transgenic plants. Data from multiple platforms including transcriptomics (microarray analysis), proteomics (UPLC/QTOF-MS), and metabolomics (GC-TOF/MS, UPLC/MS, and UHPLC-LTQ/MS) were integrated using the most recent development of orthogonal projections to latent structures called OnPLS. OnPLS is a symmetrical multi-block method that does not depend on the order of analysis when more than two blocks are analysed. Significantly affected genes, proteins and metabolites were then visualized in painted pathway diagrams. The main categories that appear to be significantly influenced in the transgenic plants were pathways related to redox regulation, carbon metabolism and protein degradation, e.g. the glycolysis and pentose phosphate pathways (PPP). The results provide system-level information on ROS metabolism and responses to oxidative stress, and indicate that some initial responses to oxidative stress may share common pathways. The proposed data evaluation strategy shows an efficient way of compiling complex, multi-platform datasets to obtain significant biological information.

  11. Models of Superoxide Dismutases

    SciTech Connect

    Cabelli, Diane E.; Riley, Dennis; Rodriguez, Jorge A.; Valentine, Joan Selverstone; Zhu, Haining

    1998-05-20

    In this review we have focused much of our discussion on the mechanistic details of how the native enzymes function and how mechanistic developments/insights with synthetic small molecule complexes possessing SOD activity have influenced our understanding of the electron transfer processes involved with the natural enzymes. A few overriding themes have emerged. Clearly, the SOD enzymes operate at near diffusion controlled rates and to achieve such catalytic turnover activity, several important physical principles must be operative. Such fast electron transfer processes requires a role for protons; i.e., proton-coupled electron transfer (''H-atom transfer'') solves the dilemma of charge separation developing in the transition state for the electron transfer step. Additionally, outer-sphere electron transfer is likely a most important pathway for manganese and iron dismutases. This situation arises because the ligand exchange rates on these two ions in water never exceed {approx}10{sup +7} s{sup -1}; consequently, 10{sup +9} catalytic rates require more subtle mechanistic insights. In contrast, copper complexes can achieve diffusion controlled (>10{sup +9}) exchange rates in water; thus inner-sphere electron transfer processes are more likely to be operative in the Cu/Zn enzymes. Recent studies have continued to expand our understanding of the mechanism of action of this most important class of redox active enzymes, the superoxide dismutases, which have been critical in the successful adaptation of life on this planet to an oxygen-based metabolism. The design of SOD mimic drugs, synthetic models compounds that incorporate this superoxide dismutase catalytic activity and are capable of functioning in vivo, offers clear potential benefits in the control of diseases, ranging from the control of neurodegenerative conditions, such as Parkinson's or Alzheimer's disease, to cancer.

  12. Ionol (BHT) produces superoxide anion.

    PubMed

    Smirnova, E G; Lyubimov, Yu I; Malinina, T G; Lyubimova, E Yu; Alexandrushkina, N I; Vanyushin, B F; Kolesova, G M; Yaguzhinsky, L S

    2002-11-01

    In aqueous medium etiolated wheat seedlings release superoxide anion (O2*-). Interaction of a synthetic antioxidant, butylated hydroxytoluene (BHT, ionol), with oxygen in the aqueous medium is accompanied by O2*- formation. This suggests that under certain conditions BHT behaves as a prooxidant. A natural antioxidant, superoxide dismutase (SOD), and also a wound healing preparation, emulsified denatured placenta (EDP), do not exhibit the prooxidant properties. In contrast to BHT, they reduce O2*- production by the etiolated wheat seedling system.

  13. Superoxide and Peroxynitrite in Atherosclerosis

    NASA Astrophysics Data System (ADS)

    White, C. Roger; Brock, Tommy A.; Chang, Ling-Yi; Crapo, James; Briscoe, Page; Ku, David; Bradley, William A.; Gianturco, Sandra H.; Gore, Jeri; Freeman, Bruce A.; Tarpey, Margaret M.

    1994-02-01

    The role of reactive oxygen species in the vascular pathology associated with atherosclerosis was examined by testing the hypothesis that impaired vascular reactivity results from the reaction of nitric oxide (^.NO) with superoxide (O^-_2), yielding the oxidant peroxynitrite (ONOO^-). Contractility studies were performed on femoral arteries from rabbits fed a cholesterol-supplemented diet. Cholesterol feeding shifted the EC50 for acetylcholine (ACh)-induced relaxation and impaired the maximal response to ACh. We used pH-sensitive liposomes to deliver CuZn superoxide dismutase (SOD; superoxide:superoxide oxidoreductase, EC 1.15.1.1) to critical sites of ^.NO reaction with O^-_2. Intravenously injected liposomes (3000 units of SOD per ml) augmented ACh-induced relaxation in the cholesterol-fed group to a greater extent than in controls. Quantitative immunocytochemistry demonstrated enhanced distribution of SOD in both endothelial and vascular smooth muscle cells as well as in the extracellular matrix. SOD activity in vessel homogenates of liposome-treated rabbits was also increased. Incubation of β very low density lipoprotein with ONOO^- resulted in the rapid formation of conjugated dienes and thiobarbituric acid-reactive substances. Our results suggest that the reaction of O^-_2 with ^.NO is involved in the development of atherosclerotic disease by yielding a potent mediator of lipoprotein oxidation, as well as by limiting ^.NO stimulation of vascular smooth muscle guanylate cyclase activity.

  14. Economical synthesis of potassium superoxide

    NASA Technical Reports Server (NTRS)

    Bell, A. T.; Sadhukhan, P.

    1979-01-01

    High-frequency discharge in oxygen can be used to prepare superoxides of alkali and alkaline-earth metals. Since no direct-current discharge at the electrodes is present, no sputtering can contaminate the product, hence a high conversion efficiency.

  15. Superoxide dismutase: an evolutionary puzzle

    SciTech Connect

    Lee, Y.M.; Friedman, D.J.; Ayala, F.J.

    1985-02-01

    The authors have obtained the complete amino acid sequence of copper/zinc-containing superoxide dismutase (SOD, superoxide:superoxide oxidoreductase, EC 1.15.1.1) from Drosophila melanogaster. The sequence of this enzyme is also known for man, horse, cow, and the yeast Saccharomyces cerevisiae. The rate of evolution of this enzyme is far from constant. The number of amino acid substitutions per 100 residues per 100 million years is 30.9 when the three mammals are compared to each other, 10.6 when Drosophila is compared to the three mammals, and 5.8 when the yeast is compared to the four animals. The first value represents one of the fastest evolutionary rates for any protein, the second is similar to the globin rate, and the third is similar to some cytochromes and other slowly evolving proteins. Hence, SOD is not acceptable evolutionary clock. Another peculiarity of this enzyme is that a two-amino-acid deletion must have occurred independently in the lineages going to the cow and to Drosophila. The authors conclude that using the primary structure of a single gene or protein to time evolutionary events or to reconstruct phylogenetic relationships is potentially fraught with error.

  16. Superoxide dismutases in chronic gastritis.

    PubMed

    Švagelj, Dražen; Terzić, Velimir; Dovhanj, Jasna; Švagelj, Marija; Cvrković, Mirta; Švagelj, Ivan

    2016-04-01

    Human gastric diseases have shown significant changes in the activity and expression of superoxide dismutase (SOD) isoforms. The aim of this study was to detect Mn-SOD activity and expression in the tissue of gastric mucosa, primarily in chronic gastritis (immunohistochemical Helicobacter pylori-negative gastritis, without other pathohistological changes) and to evaluate their possible connection with pathohistological diagnosis. We examined 51 consecutive outpatients undergoing endoscopy for upper gastrointestinal symptoms. Patients were classified based on their histopathological examinations and divided into three groups: 51 patients (archive samples between 2004-2009) with chronic immunohistochemical Helicobacter pylori-negative gastritis (mononuclear cells infiltration were graded as absent, moderate, severe) divided into three groups. Severity of gastritis was graded according to the updated Sydney system. Gastric tissue samples were used to determine the expression of Mn-SOD with anti-Mn-SOD Ab immunohistochemically. The Mn-SOD expression was more frequently present in specimens with severe and moderate inflammation of gastric mucosa than in those with normal mucosa. In patients with normal histological finding, positive immunoreactivity of Mn-SOD was not found. Our results determine the changes in Mn-SOD expression occurring in the normal gastric mucosa that had undergone changes in the intensity of chronic inflammatory infiltrates in the lamina propria.

  17. DPI induces mitochondrial superoxide-mediated apoptosis.

    PubMed

    Li, Nianyu; Ragheb, Kathy; Lawler, Gretchen; Sturgis, Jennie; Rajwa, Bartek; Melendez, J Andres; Robinson, J Paul

    2003-02-15

    The iodonium compounds diphenyleneiodonium (DPI) and diphenyliodonium (IDP) are well-known phagocyte NAD(P)H oxidase inhibitors. However, it has been shown that at high concentrations they can inhibit the mitochondrial respiratory chain as well. Since inhibition of the mitochondrial respiratory chain has been shown to induce superoxide production and apoptosis, we investigated the effect of iodonium compounds on mitochondria-derived superoxide and apoptosis. Mitochondrial superoxide production was measured on both cultured cells and isolated rat-heart submitochondrial particles. Mitochondria function was examined by monitoring mitochondrial membrane potential. Apoptotic pathways were studied by measuring cytochrome c release and caspase 3 activation. Apoptosis was characterized by detecting DNA fragmentation on agarose gel and measuring propidium iodide- (PI-) stained subdiploid cells using flow cytometry. Our results showed that DPI could induce mitochondrial superoxide production. The same concentration of DPI induced apoptosis by decreasing mitochondrial membrane potential and releasing cytochrome c. Addition of antioxidants or overexpression of MnSOD significantly reduced DPI-induced mitochondrial damage, cytochrome c release, caspase activation, and apoptosis. These observations suggest that DPI can induce apoptosis via induction of mitochondrial superoxide. DPI-induced mitochondrial superoxide production may prove to be a useful model to study the signaling pathways of mitochondrial superoxide.

  18. Mast cell release of superoxide

    SciTech Connect

    Dileepan, K.N.; Simpson, K.M.; Stechschulte, D.J.

    1987-05-01

    The ability of rat serosal mast cells (MC) to release superoxide (O/sub 2//sup -/) upon activation by immunologic and nonimmunologic stimuli was investigated. Purified MC (90-95%) were either sensitized with monoclonal IgE reactive against dinitrophenyl bovine serum albumin (DNP-BSA) and challenged with DNP-BSA, or naive MC were treated with compound 48/80 or ionophore A23187. O/sub 2//sup -/ release was measured by O/sub 2//sup -/ dismutase (SOD)-sensitive reduction of cytochrome C and MC activation was assessed by the release of histamine or (/sup 14/C)5-hydroxytryptamine (5HT). The results reveal that activation of MC by 48/80 or immunologic challenge does not release O/sub 2//sup -/, although these stimuli induce substantial release of histamine and 5HT (40-70%). In contrast, A23187 released O/sub 2//sup -/ (3-6 nMols/10/sup 6/ MC) and histamine (40-80%). In mixed cell preparations containing MC and macrophages (M0), activation of MC with 48/80 resulted in inhibition of M0 O/sub 2//sup -/ release. The MC-mediated inhibition of O/sub 2//sup -/ production was not due to histamine or 5HT, but was due to MC-granule SOD. MC contain abundant quantities of SOD and, therefore, release O/sub 2//sup -/ only when its production exceeds the intracellular SOD threshold following activation with selective stimuli. In addition, the apparent differences in the mode and site of action of various stimuli on MC may contribute to the discriminative release of O/sub 2//sup -/.

  19. Metal Uptake by Manganese Superoxide Dismutase

    PubMed Central

    Whittaker, James W.

    2009-01-01

    Manganese superoxide dismutase is an important antioxidant defense metalloenzyme that protects cells from damage by the toxic oxygen metabolite, superoxide free radical, formed as an unavoidable by-product of aerobic metabolism. Many years of research have gone into understanding how the metal cofactor interacts with small molecules in its catalytic role. In contrast, very little is presently known about how the protein acquires its metal cofactor, an important step in the maturation of the protein and one that is absolutely required for its biological function. Recent work is beginning to provide insight into the mechanisms of metal delivery to manganese superoxide dismutase in vivo and in vitro. PMID:19699328

  20. Oxygen plasmas used to synthesize superoxides

    NASA Technical Reports Server (NTRS)

    Hollahan, J. R.; Wydeven, T.

    1972-01-01

    Production of alkali metal superoxides by interaction of molecular oxygen with alkali metals or their salts is discussed. Diagram of reactor to show components and operating principles is provided. Analysis of chemical reactions involved is developed.

  1. Biological Superoxide In Manganese Oxide Formation

    NASA Astrophysics Data System (ADS)

    Hansel, C.; Learman, D.; Zeiner, C.; Santelli, C. M.

    2011-12-01

    Manganese (Mn) oxides are among the strongest sorbents and oxidants within the environment, controlling the fate and transport of numerous elements and the degradation of recalcitrant carbon. Both bacteria and fungi mediate the oxidation of Mn(II) to Mn(III/IV) oxides but the genetic and biochemical mechanisms responsible remain poorly understood. Furthermore, the physiological basis for microbial Mn(II) oxidation remains an enigma. We have recently reported that a common marine bacterium (Roseobacter sp. AzwK-3b) oxidizes Mn(II) via reaction with extracellular superoxide (O2-) produced during exponential growth. Here we expand this superoxide-mediated Mn(II) oxidation pathway to fungi, introducing a surprising homology between prokaryotic and eukaryotic metal redox processes. For instance, Stibella aciculosa, a common soil Ascomycete filamentous fungus, precipitates Mn oxides at the base of asexual reproductive structures (synnemata) used to support conidia (Figure 1). This distribution is a consequence of localized production of superoxide (and it's dismutation product hydrogen peroxide, H2O2), leading to abiotic oxidation of Mn(II) by superoxide. Disruption of NADPH oxidase activity using the oxidoreductase inhibitor DPI leads to diminished cell differentiation and subsequent Mn(II) oxidation inhibition. Addition of Cu(II) (an effective superoxide scavenger) leads to a concentration dependent decrease in Mn oxide formation. We predict that due to the widespread production of extracellular superoxide within the fungal and likely bacterial kingdoms, biological superoxide may be an important contributor to the cycling of Mn, as well as other metals (e.g., Hg, Fe). Current and future explorations of the genes and proteins involved in superoxide production and Mn(II) oxidation will ideally lend insight into the physiological and biochemical basis for these processes.

  2. Periplasmic Superoxide Dismutase in Meningococcal Pathogenicity

    PubMed Central

    Wilks, Kathryn E.; Dunn, Kate L. R.; Farrant, Jayne L.; Reddin, Karen M.; Gorringe, Andrew R.; Langford, Paul R.; Kroll, J. Simon

    1998-01-01

    Meningococcal sodC encodes periplasmic copper- and zinc-cofactored superoxide dismutase (Cu,Zn SOD) which catalyzes the conversion of the superoxide radical anion to hydrogen peroxide, preventing a sequence of reactions leading to production of toxic hydroxyl free radicals. From its periplasmic location, Cu,Zn SOD was inferred to acquire its substrate from outside the bacterial cell and was speculated to play a role in preserving meningococci from the action of microbicidal oxygen free radicals produced in the context of host defense. A sodC mutant was constructed by allelic exchange and was used to investigate the role of Cu,Zn SOD in pathogenicity. Wild-type and mutant meningococci grew at comparable rates and survived equally long in aerobic liquid culture. The mutant showed no increased sensitivity to paraquat, which generates superoxide within the cytosol, but was approximately 1,000-fold more sensitive to the toxicity of superoxide generated in solution by the xanthine/xanthine oxidase system. These data support a role for meningococcal Cu,Zn SOD in protection against exogenous superoxide. In experiments to translate this into a role in pathogenicity, wild-type and mutant organisms were used in an intraperitoneal mouse infection model. The sodC mutant was significantly less virulent. We conclude that periplasmic Cu,Zn SOD contributes to the virulence of Neisseria meningitidis, most likely by reducing the effectiveness of toxic oxygen host defenses. PMID:9423860

  3. 2-methoxyestradiol does not inhibit superoxide dismutase.

    PubMed

    Kachadourian, R; Liochev, S I; Cabelli, D E; Patel, M N; Fridovich, I; Day, B J

    2001-08-15

    It has been reported in the literature that the endogenous estrogen metabolite 2-methoxyestradiol (2-ME) inhibits both manganese and copper,zinc superoxide dismutases (Mn and Cu,Zn SODs) and that this mechanism is responsible for 2-ME's ability to kill cancer cells. In fact, as demonstrated using several SOD assays including pulse radiolysis, 2-ME does not inhibit SOD but rather interferes with the SOD assay originally used. Nevertheless, as confirmed by aconitase inactivation measurements and lactate dehydrogenase release in human leukemia HL-60 cells, 2-ME does increase superoxide production in these cells and is more toxic than its non-O-methylated precursor 2-hydroxyestradiol. Other mechanisms previously suggested in the literature may explain 2-ME's ability to increase intracellular superoxide levels in tumor cells. Copyright 2001 Academic Press.

  4. Process for the preparation of calcium superoxide

    NASA Technical Reports Server (NTRS)

    Ballou, E. V.; Wood, P. C.; Wydeven, T. J.; Spitze, L. A. (Inventor)

    1978-01-01

    Calcium superoxide is prepared in high yields by spreading a quantity of calcium peroxide diperoxyhydrate on the surface of a container, positioning said container in a vacuum chamber on a support structure through which a coolant fluid can be circulated, partially evacuating said vacuum chamber, allowing the temperature of the diperoxyhydrate to reach the range of about 0 to about 40 C; maintaining the temperature selected for a period of time sufficient to complete the disproproriation of the diperoxyhydrate to calcium superoxide, calcium hydroxide, oxygen, and water; constantly and systematically removing the water as it is formed by sweeping the reacting material with a current of dry inert gas and/or by condensation of said water on a cold surface; backfilling the chamber with a dry inert gas; and finally, recovering the calcium superoxide produced.

  5. Induction of Superoxide Dismutase by Molecular Oxygen

    PubMed Central

    Gregory, Eugene M.; Fridovich, Irwin

    1973-01-01

    Oxygen induces superoxide dismutase in Streptococcus faecalis and in Escherichia coli B. S. faecalis grown under 20 atm of O2 had 16 times more of this enzyme than did anaerobically grown cells. In the case of E. coli, changing the conditions of growth from anaerobic to 5 atm of O2 caused a 25-fold increase in the level of superoxide dismutase. Induction of this enzyme was a response to O2 rather than to pressure, since 20 atm of N2 was without effect. Induction of superoxide dismutase was a rapid process, and half of the maximal level was reached within 90 min after N2-grown cells of S. faecalis were exposed to 20 atm of O2 at 37 C. S. faecalis did not contain perceptible levels of catalase under any of the growth conditions investigated by Stanier, Doudoroff, and Adelberg (23), and the concentration of catalase in E. coli was not affected by the presence of O2 during growth. S. faecalis, which had been grown under 100% O2 and which therefore contained an elevated level of superoxide dismutase, was more resistant of 46 atm of O2 than were cells which had been grown under N2. E. coli grown under N2 contained as much superoxide dismutase as did S. faecalis grown under 1 atm of O2. The E. coli which had been grown under N2 was as resistant to the deleterious effects of 50 atm of O2 as was S. faecalis which had been grown under 1 atm of O2. These results are consistent with the proposal that the peroxide radical is an important agent of the toxicity of oxygen and that superoxide dismutase may be a component of the systems which have been evolved to deal with this potential toxicity. Images PMID:4196244

  6. In vitro inhibition of superoxide anion production and superoxide dismutase activity by zinc in human spermatozoa.

    PubMed

    Gavella, M; Lipovac, V; Vucić, M; Sverko, V

    1999-08-01

    The in vitro effect of zinc on superoxide anion (O2-) generation and on SOD-like activity in spermatozoa of infertile men was investigated. The formation of superoxide anion was stimulated by NADPH and the level of superoxide anion was measured by the reduction of ferricytochrome c. Both Percoll-isolated (n = 14) and washed spermatozoa (n = 14) exposed to 1 mmol/L zinc (60 min, 37 degrees C), released less (p < 0.002 and p < 0.04, respectively) superoxide anions than did zinc-untreated spermatozoa. These results implicate a possible role for zinc as a scavenger of excessive superoxide anions produced by defective spermatozoa in semen after ejaculation. Additionally, zinc was found to dose-dependently inhibit superoxide dismutase (SOD)-like activity of spermatozoa in vitro. The inhibition of SOD-like activity by an equal concentration of zinc (1 mmol/L) was less pronounced in oligospermic (p < 0.002; n = 16) and asthenozoospermic (p < 0.0005; n = 20) than in normozoospermic samples (p < 0.0001; n = 20). This differential ability of zinc to inhibit SOD-like activity may be relevant to the physiological function of spermatozoa in fertilization. The evidence that zinc may elicit an inhibition of both superoxide anion production and SOD-like activity in human spermatozoa, indicate the existence of novel, zinc-related mechanism(s) involved in the oxidative events occurring after ejaculation, with a possible modulatory effect on germ cell function.

  7. Diagnosis of superoxide anion radical induced in liquids by atmospheric-pressure plasma using superoxide dismutase

    NASA Astrophysics Data System (ADS)

    Tani, Atsushi; Fukui, Satoshi; Ikawa, Satoshi; Kitano, Katsuhisa

    2015-01-01

    To confirm the formation of the superoxide anion radical (O2-•) in liquids by atmospheric-pressure plasma, we investigated plasma-induced radical species in water using the electron spin resonance (ESR) spin-trapping technique combined with two proteins: superoxide dismutase (SOD), which has enzymatic activity to quench the superoxide anion radical, and bovine serum albumin (BSA), which does not have this enzymatic activity. Different setups of contact and non-contact atmospheric-pressure helium plasma were tested with an additional supply of oxygen gas. For each setup of plasma, no superoxide anion adduct ESR signal was observed in the aqueous solution with SOD, whereas the ESR signal appears in the samples with BSA and without any additive proteins. This means that a superoxide anion radical in the solution is sufficiently quenched by SOD before the formation of the spin adduct. The superoxide anion radical is actually induced in an aqueous solution by atmospheric-pressure plasma when ambient gases contain oxygen.

  8. Superoxide Dismutases and Reactive Oxygen Species

    SciTech Connect

    Cabelli, D.E.

    2011-01-01

    The 'free radical theory' of aging was introduced over a half-century ago. In this theory, much of the deleterious effects of aging were attributed to the cumulative buildup of damage from reactive oxygen species. When discussing reactive oxygen species (ROS) in aerobic systems, both superoxide radicals (O{sub 2}{sup -}) and superoxide dismutases (SODs) are considered to play prominent roles. O{sub 2}{sup -} is formed by attachment of the electron to oxygen (O{sub 2}) that is present in tens to hundreds of micromolar concentration in vivo. SODs are enzymes that serve to eliminate O{sub 2}{sup -} by rapidly converting it to O{sub 2} and hydrogen peroxide (H{sub 2}O{sub 2}). Both the radical and the enzyme will be discussed with the focus on the systems that are present in humans.

  9. Role of extracellular superoxide dismutase in hypertension.

    PubMed

    Gongora, Maria Carolina; Qin, Zhenyu; Laude, Karine; Kim, Ha Won; McCann, Louise; Folz, J Rodney; Dikalov, Sergey; Fukai, Tohru; Harrison, David G

    2006-09-01

    We previously found that angiotensin II-induced hypertension increases vascular extracellular superoxide dismutase (ecSOD), and proposed that this is a compensatory mechanism that blunts the hypertensive response and preserves endothelium-dependent vasodilatation. To test this hypothesis, we studied ecSOD-deficient mice. ecSOD(-/-) and C57Blk/6 mice had similar blood pressure at baseline; however, the hypertension caused by angiotensin II was greater in ecSOD(-/-) compared with wild-type mice (168 versus 147 mm Hg, respectively; P<0.01). In keeping with this, angiotensin II increased superoxide and reduced endothelium-dependent vasodilatation in small mesenteric arterioles to a greater extent in ecSOD(-/-) than in wild-type mice. In contrast to these findings in resistance vessels, angiotensin II paradoxically improved endothelium-dependent vasodilatation, reduced intracellular and extracellular superoxide, and increased NO production in aortas of ecSOD(-/-) mice. Whereas aortic expression of endothelial NO synthase, Cu/ZnSOD, and MnSOD were not altered in ecSOD(-/-) mice, the activity of Cu/ZnSOD was increased by 80% after angiotensin II infusion. This was associated with a concomitant increase in expression of the copper chaperone for Cu/ZnSOD in the aorta but not in the mesenteric arteries. Moreover, the angiotensin II-induced increase in aortic reduced nicotinamide-adenine dinucleotide phosphate oxidase activity was diminished in ecSOD(-/-) mice as compared with controls. Thus, during angiotensin II infusion, ecSOD reduces hypertension, minimizes vascular superoxide production, and preserves endothelial function in resistance arterioles. We also identified novel compensatory mechanisms involving upregulation of copper chaperone for Cu/ZnSOD, increased Cu/ZnSOD activity, and decreased reduced nicotinamide-adenine dinucleotide phosphate oxidase activity in larger vessels. These compensatory mechanisms preserve large vessel function when ecSOD is absent in

  10. Extracellular superoxide dismutase of boar seminal plasma.

    PubMed

    Kowalowka, M; Wysocki, P; Fraser, L; Strzezek, J

    2008-08-01

    Superoxide dismutase (SOD) is an enzymatic component of the antioxidant defense system that protects spermatozoa by catalysing the dismutation of superoxide anions to hydrogen peroxide and oxygen. Age and season effects on SOD activity in the seminal plasma were measured in boars at the onset of 8 months through a 35-month period. It was found that age-related changes in SOD activity in the seminal plasma were markedly higher in boars less than 2 years of age. However, it appeared that SOD activity was established at the early sexual maturity age (8-12 months). There were variations in SOD activity throughout the season, being significantly higher in spring and autumn than in summer. A secretory extracellular form of SOD (EC-SOD) was purified to homogeneity (350-fold) from boar seminal plasma, using a three-step purification protocol (affinity chromatography followed by ion exchange and ceramic hydroxyapatite chromatography). The molecular properties and specificity of SOD (molecular mass, isoelectric point, optimum pH, thermostability and susceptibility to inhibitors) confirmed that the purified enzyme is an extracellular form of Cu/Zn-superoxide dismutase occurring in boar seminal plasma. The results of this study indicate that EC-SOD is an important antioxidant enzyme of boar seminal plasma, which plays an important physiological role in counteracting oxidative stress in spermatozoa.

  11. Superoxide decay kinetics in the southern ocean.

    PubMed

    Heller, Maija I; Croot, Peter L

    2010-01-01

    Measurements of superoxide (O(2)(-)) reaction kinetics were made during a transect with the research icebreaker Polarstern (ANT24-3) in the Antarctic through the Drake Passage in austral autumn 2008. Our sampling strategy was designed to investigate the sinks of superoxide in Polar waters; principally through reactions with dissolved organic matter (DOM) or metals (copper and iron). We modified an existing chemiluminescence flow injection system using methyl Cypridina luciferin analog (MCLA) for the detection of O(2)(-) and added O(2)(-) using KO(2) as the source. Our results indicate that O(2)(-) in ambient seawater had a half-life ranging from 9.3 to 194 s. DTPA additions to seawater, to remove the effects of reactions with metals, revealed O(2)(-) decay rates consistent with a second order reaction, indicating that the dismutation reaction dominated and that reactions with DOM were not significant. Titrations of seawater by the addition of nanomolar amounts of iron or copper revealed the importance of organic chelation of Fe and/or Cu in controlling the reactivity with O(2)(-). Throughout the water column reactions with Cu appeared to be the major sink for superoxide in the Southern Ocean. This new strategy suggests an alternative approach for speciation measurements of Fe and Cu in seawater.

  12. The structural biochemistry of the superoxide dismutases

    PubMed Central

    Perry, J.J.P.; Shin, D.S.; Getzoff, E.D.; Tainer, J.A.

    2011-01-01

    The discovery of superoxide dismutases (SODs), which convert superoxide radicals to molecular oxygen and hydrogen peroxide, has been termed the most important discovery of modern biology never to win a Nobel Prize. Here, we review the reasons this discovery has been underappreciated, as well as discuss the robust results supporting its premier biological importance and utility for current research. We highlight our understanding of SOD function gained through structural biology analyses, which reveal important hydrogen-bonding schemes and metal-binding motifs. These structural features create remarkable enzymes that promote catalysis at faster than diffusion-limited rates by using electrostatic guidance. These architectures additionally alter the redox potential of the active site metal center to a range suitable for the superoxide disproportionation reaction and protect against inhibition of catalysis by molecules such as phosphate. SOD structures may also control their enzymatic activity through product inhibition; manipulation of these product inhibition levels has the potential to generate therapeutic forms of SOD. Markedly, structural destabilization of the SOD architecture can lead to disease, as mutations in Cu,ZnSOD may result in familial amyotrophic lateral sclerosis, a relatively common, rapidly progressing and fatal neurodegenerative disorder. We describe our current understanding of how these Cu,ZnSOD mutations may lead to aggregation/fibril formation, as a detailed understanding of these mechanisms provides new avenues for the development of therapeutics against this so far untreatable neurodegenerative pathology. PMID:19914407

  13. A modular trigger for the development of selective superoxide probes.

    PubMed

    Yu, Zuo Hang; Chung, Clive Yik-Sham; Tang, Fung Kit; Brewer, Thomas F; Au-Yeung, Ho Yu

    2017-09-05

    We report here a new bioinspired copper-based strategy of superoxide sensing and the development of sensitive (>90-fold fluorescence turn-on) and selective superoxide probes for imaging variations in the endogenous superoxide level in various live mammalian cells (HEK293T, HeLa and A431).

  14. Scavenging of superoxide anions by lecithinized superoxide dismutase in HL-60 cells.

    PubMed

    Ishihara, Tsutomu; Shibui, Misaki; Hoshi, Takaya; Mizushima, Tohru

    2016-01-01

    Superoxide dismutase covalently bound to four lecithin molecules (PC-SOD) has been found to have beneficial therapeutic effects in animal models of various diseases. However, the mechanism underlying these improved therapeutic effects has not yet been elucidated. It has previously been shown that PC-SOD localizes on the plasma membrane and in the lysosomes of cells. In this study, we evaluated the superoxide anion-scavenging activity of PC-SOD in HL-60 human promyelocytic leukemia cells. Compared to SOD, PC-SOD had only 17% scavenging activity in cell-free systems. Nevertheless, by analyzing enzyme activities in cell suspensions containing PC-SOD or SOD, PC-SOD and SOD showed almost equal activity for scavenging extracellular superoxide anions produced by HL-60 cells. Furthermore, the activity for scavenging extracellular superoxide anions increased with increased amount of PC-SOD on the plasma membrane. Moreover, PC-SOD exhibited no obvious inhibitory effect on the scavenging of intracellular superoxide anions. These results suggested that the association of PC-SOD with the plasma membrane plays a key role in its beneficial therapeutic effects. Thus, this finding may provide a rationale for selecting target diseases for PC-SOD treatment.

  15. Superoxide Dismutase in the Symbiont Anabaena azollae Strasb. 1

    PubMed Central

    Canini, A.; Galiazzo, F.; Rotilio, G.; Caiola, M. Grilli

    1991-01-01

    Superoxide dismutase was investigated in the symbiont Anabaena azollae Strasb. living in Azolla filiculoides Lam. In vegetative cells, three isoenzymatic forms of superoxide dismutase, containing manganese, iron, and the hybrid iron-manganese, respectively, were present. Hybrid superoxide dismutase, detected for the first time in cyanobacteria, was 7% of the total superoxide dismutase present in vegetative cells. All three superoxide dismutase forms increased in the Anabaena vegetative cells obtained from irradiated plants grown in winter. In heterocysts, only an iron superoxide dismutase was present, which amounted to 25% of total vegetative cell superoxide dismutase activity. Hybrid superoxide dismutase appeared in heterocysts after irradiation. In vegetative cells of Anabaena from plants grown in summer, the basal level of total superoxide dismutase increased by 60% as compared with winter, and was unaffected by irradiation. The levels of superoxide dismutase in heterocysts from control and exposed plants grown in summer were comparable to those observed in heterocysts obtained from the plants grown during winter. No direct correlation was found between nitrogenase activity and superoxide dismutase in heterocysts. The presence of cyanophycin granules, either within the heterocyst pore channel or close to the transversal septum of vegetative cells, suggested a mechanism to stop communications between vegetative cells and heterocysts. ImagesFigure 1Figure 3Figure 4Figure 5 PMID:16668392

  16. Differential effects of superoxide dismutase and superoxide dismutase/catalase mimetics on human breast cancer cells.

    PubMed

    Shah, Manisha H; Liu, Guei-Sheung; Thompson, Erik W; Dusting, Gregory J; Peshavariya, Hitesh M

    2015-04-01

    Reactive oxygen species (ROS) such as superoxide and hydrogen peroxide (H2O2) have been implicated in development and progression of breast cancer. In the present study, we have evaluated the effects of the superoxide dismutase (SOD) mimetic MnTmPyP and the SOD/catalase mimetic EUK 134 on superoxide and H2O2 formation as well as proliferation, adhesion, and migration of MCF-7 and MDA-MB-231 cells. Superoxide and H2O2 production was examined using dihydroethidium and Amplex red assays, respectively. Cell viability and adhesion were measured using a tetrazolium-based MTT assay. Cell proliferation was determined using trypan blue assay. Cell cycle progression was analyzed using flow cytometry. Clonal expansion of a single cell was performed using a colony formation assay. Cell migration was measured using transwell migration assay. Dual luciferase assay was used to determine NF-κB reporter activity. EUK 134 effectively reduced both superoxide and H2O2, whereas MnTmPyP removed superoxide but enhanced H2O2 formation. EUK 134 effectively attenuated viability, proliferation, clonal expansion, adhesion, and migration of MCF-7 and MDA-MB-231 cells. In contrast, MnTmPyP only reduced clonal expansion of MCF-7 and MDA-MB-231 cells but had no effect on adhesion and cell cycle progression. Tumor necrosis factor-alpha-induced NF-κB activity was reduced by EUK 134, whereas MnTmPyP enhanced this activity. These data indicate that the SOD mimetic MnTmPyP and the SOD/catalase mimetic EUK 134 exert differential effects on breast cancer cell growth. Inhibition of H2O2 signaling using EUK 134-like compound might be a promising approach to breast cancer therapy.

  17. Superoxide Radical Lifetime on the Martian Surface

    NASA Astrophysics Data System (ADS)

    Zent, A. P.; Ichimura, A.; Quinn, R. C.

    2005-08-01

    We have examined the formation and stability of the superoxide radical O2-, which has been hypothesized as a potential Mars oxidant. Rutile (TiO2) was heated to ˜ 400 degrees C under vacuum. The samples were tipped off in ampules under 8-9 torr O2, photolyzed with a Hg lamp for 30 minutes; EPR spectra were immediately obtained at 77K. The signature of O2- was clearly observed in the rutile. The sealed ampules were stored at room temperature for up to 2 weeks without any decrease in the number of spins. The same process, applied to rutile that was not baked out yielded superoxide signals that could not be detected once the photolyzing flux was cut off. To examine the effects of partial dehydration, we carried out the same series of experiments on rutile that was baked out at 200 degrees C. This material showed decay of superoxide spins to zero in less than 10 minutes. This qualitative pattern is also observed in experiments on anatase (Attwood, et al., , 2003). We hypothesize that O2- can be stabilized against reaction with H2O and OH by crystalline surface defects. On hydrated surfaces, O2- must compete for stabilizing sites, and the population is quickly extinguished; in dehydrated samples, it can migrate to stabilizing defects. Once sorbed, the O2- radical is stable in the presence of H2O. OMEGA Mars Express data (Poullet et al, 2005) suggest one to several percent adsorbed H2O across the Martian surface, which will significantly decrease O2- lifetime. One possibility for subsurface stabilization of O2- can be postulated based on EPR spectra of anatase, exposed to H2O2 in our lab in 1996, and which in 2005 shows the signature of O2-. Evidently, H2O2 can convert to stable O2- on some surfaces. This hypothesis might allow subsurface diffusion of H2O2, followed by conversion to O2-.

  18. Subsarcolemmal and interfibrillar mitochondria display distinct superoxide production profiles.

    PubMed

    Crochemore, C; Mekki, M; Corbière, C; Karoui, A; Noël, R; Vendeville, C; Vaugeois, J-M; Monteil, C

    2015-03-01

    Cardiac subsarcolemmal mitochondria (SSM) and interfibrillar mitochondria (IFM) subpopulations display distinct biochemical, morphological, and functional characteristics. Moreover, they appear to be differently influenced during cardiac pathologies or toxic injuries. Although mitochondrial reactive oxygen species seem to play a critical role in cardiac function and diseases, limited information exists about the superoxide production characteristics of these mitochondrial subpopulations. In this work, using direct measurement of superoxide by electron paramagnetic resonance, we showed that differences in superoxide production profiles were present between cardiac IFM and SSM, in terms of intensity and major sites of superoxide generation. In SSM incubated with glutamate plus malate as substrates, the total observed superoxide levels were significantly higher than those observed with IFM, with an important contribution of the NADH-oxidizing site of complex I (site If) and the quinol-oxidizing site of complex III (site IIIQ0). In both IFM and SSM, succinate leads to similar rates of total superoxide levels with a substantial role for contribution of reverse electron transfer. Finally, using two spin probes with different membrane permeabilities, our data on complex III showed direct intra- and extra-mitochondrial superoxide release whereas complex I- and II-dependent superoxide were exclusively released inside the mitochondria, confirming previous studies. Feasibility of this approach to measure intra- and extra-mitochondrial superoxide levels and to characterize distinct superoxide production profiles of cardiac IFM and SSM has been demonstrated.

  19. The peroxidase activity of mitochondrial superoxide dismutase.

    PubMed

    Ansenberger-Fricano, Kristine; Ganini, Douglas; Mao, Mao; Chatterjee, Saurabh; Dallas, Shannon; Mason, Ronald P; Stadler, Krisztian; Santos, Janine H; Bonini, Marcelo G

    2013-01-01

    Manganese superoxide dismutase (MnSOD) is an integral mitochondrial protein known as a first-line antioxidant defense against superoxide radical anions produced as by-products of the electron transport chain. Recent studies have shaped the idea that by regulating the mitochondrial redox status and H(2)O(2) outflow, MnSOD acts as a fundamental regulator of cellular proliferation, metabolism, and apoptosis, thereby assuming roles that extend far beyond its proposed antioxidant functions. Accordingly, allelic variations of MnSOD that have been shown to augment levels of MnSOD in mitochondria result in a 10-fold increase in prostate cancer risk. In addition, epidemiologic studies indicate that reduced glutathione peroxidase activity along with increases in H(2)O(2) further increase cancer risk in the face of MnSOD overexpression. These facts led us to hypothesize that, like its Cu,ZnSOD counterpart, MnSOD may work as a peroxidase, utilizing H(2)O(2) to promote mitochondrial damage, a known cancer risk factor. Here we report that MnSOD indeed possesses peroxidase activity that manifests in mitochondria when the enzyme is overexpressed. Copyright © 2012 Elsevier Inc. All rights reserved.

  20. Superoxide reduction by a superoxide reductase lacking the highly conserved lysine residue

    SciTech Connect

    Teixeira, Miguel; Cabelli, Diane; Pinto, Ana F.; Romao, Celia V.; Pinto, Liliana C.; Huber, Harald; Saraiva, Ligia M.; Todorovic, Smilja

    2014-12-05

    Superoxide reductases (SORs) are the most recently identified superoxide detoxification systems, being found in microorganisms from the three domains of life. These enzymes are characterized by a catalytic mononuclear iron site, with one cysteine and four histidine ligands of the ferrous active form. A lysine residue in the –EKHVP– motif, located close to the active site, has been considered to be essential for the enzyme function, by contributing to the positive surface patch that attracts the superoxide anion and by controlling the chemistry of the catalytic mechanism through a hydrogen bond network. However, we show here that this residue is substituted by non-equivalent amino acids in several putative SORs from Archaea and unicellular Eukarya. In this work, we focus on mechanistic and spectroscopic studies of one of these less common enzymes, the SOR from the hyperthermophilic crenarchaeon Ignicoccus hospitalis. We employ pulse radiolysis fast kinetics and spectroscopic approaches to study the wild-type enzyme (₋E₂₃T₂₄HVP₋), and two mutants, T24K and E23A, the later mimicking enzymes lacking both the lysine and glutamate (a ferric ion ligand) of the motif. The efficiency of the wild type protein and mutants in reducing superoxide is comparable to other SORs, revealing the robustness of these enzymes to single mutations.

  1. Superoxide reduction by a superoxide reductase lacking the highly conserved lysine residue

    DOE PAGES

    Teixeira, Miguel; Cabelli, Diane; Pinto, Ana F.; ...

    2014-12-05

    Superoxide reductases (SORs) are the most recently identified superoxide detoxification systems, being found in microorganisms from the three domains of life. These enzymes are characterized by a catalytic mononuclear iron site, with one cysteine and four histidine ligands of the ferrous active form. A lysine residue in the –EKHVP– motif, located close to the active site, has been considered to be essential for the enzyme function, by contributing to the positive surface patch that attracts the superoxide anion and by controlling the chemistry of the catalytic mechanism through a hydrogen bond network. However, we show here that this residue ismore » substituted by non-equivalent amino acids in several putative SORs from Archaea and unicellular Eukarya. In this work, we focus on mechanistic and spectroscopic studies of one of these less common enzymes, the SOR from the hyperthermophilic crenarchaeon Ignicoccus hospitalis. We employ pulse radiolysis fast kinetics and spectroscopic approaches to study the wild-type enzyme (₋E₂₃T₂₄HVP₋), and two mutants, T24K and E23A, the later mimicking enzymes lacking both the lysine and glutamate (a ferric ion ligand) of the motif. The efficiency of the wild type protein and mutants in reducing superoxide is comparable to other SORs, revealing the robustness of these enzymes to single mutations.« less

  2. Superoxide reduction by a superoxide reductase lacking the highly conserved lysine residue.

    PubMed

    Pinto, Ana F; Romão, Célia V; Pinto, Liliana C; Huber, Harald; Saraiva, Lígia M; Todorovic, Smilja; Cabelli, Diane; Teixeira, Miguel

    2015-01-01

    Superoxide reductases (SORs) are the most recently identified superoxide detoxification systems, being found in microorganisms from the three domains of life. These enzymes are characterized by a catalytic mononuclear iron site, with one cysteine and four histidine ligands of the ferrous active form. A lysine residue in the -EKHVP- motif, located close to the active site, has been considered to be essential for the enzyme function, by contributing to the positive surface patch that attracts the superoxide anion and by controlling the chemistry of the catalytic mechanism through a hydrogen bond network. However, we show here that this residue is substituted by non-equivalent amino acids in several putative SORs from Archaea and unicellular Eukarya. In this work, we focus on mechanistic and spectroscopic studies of one of these less common enzymes, the SOR from the hyperthermophilic Crenarchaeon Ignicoccus hospitalis. We employ pulse radiolysis fast kinetics and spectroscopic approaches to study the wild-type enzyme (-E23T24HVP-), and two mutants, T24K and E23A, the later mimicking enzymes lacking both the lysine and glutamate (a ferric ion ligand) of the motif. The efficiency of the wild-type protein and mutants in reducing superoxide is comparable to other SORs, revealing the robustness of these enzymes to single mutations.

  3. Superoxide production and decay in the subtropical North Pacific

    NASA Astrophysics Data System (ADS)

    Roe, K.; Voelker, B. M.; Hansel, C. M.

    2012-12-01

    Reactive oxygen species (ROS), which include superoxide and hydrogen peroxide, can be generated through photochemical reactions or biological activity in seawater. The generation of ROS, especially superoxide, by photochemical or biological processes can influence trace metal speciation and cycling in the ocean since superoxide can react quickly with metals (Cu and Fe) and is capable of both oxidation and reduction of trace metals. In this study superoxide was detected and measured in the oligotrophic waters at station ALOHA by a MCLA chemiluminescence flow injection method. The superoxide concentrations ranged between 0.037-0.099 nM, had observed decay rates of 0.004-0.014 s-1, and production rates of 0.88-4.81 nM hr-1 during a 16 day period during July 2012. The influence of biological activity vs photochemical production on superoxide concentration, decay and production rates are discussed.

  4. Chemical reactivity and biological effects of superoxide radicals

    NASA Astrophysics Data System (ADS)

    Chuaqui, C. A.; Petkau, A.

    The chemical reactivity of the superoxide radical is described in terms of its dismutation to oxygen, its basicity and ability to act as a nucleophile in aprotic media, and in its capacity to mediate one electron transfer reactions and undergo conversion to other species of active oxygen. The biological role of the superoxide radical is discussed in terms of its diffusion and reactivity within membranes, its potential to inactivate enzymes and elicit DNA damage in human granulocytes activated by phorbol esters. Mechanisms for reduction of oxygen to the superoxide radical by riboflavin, nitroaromatic compounds, nitrofurazone and oxazolinone are described. Finally, inactivation of bone marrow progenitor cells by superoxide radicals generated photochemically is discussed to emphasize the sensitizing effect of the medium, the sequential toxicity of superoxide and hydrogen peroxide, and the protective effects of superoxide dismutase and catalase while acting in their respective time frames.

  5. A mitochondrial superoxide theory for oxidative stress diseases and aging.

    PubMed

    Indo, Hiroko P; Yen, Hsiu-Chuan; Nakanishi, Ikuo; Matsumoto, Ken-Ichiro; Tamura, Masato; Nagano, Yumiko; Matsui, Hirofumi; Gusev, Oleg; Cornette, Richard; Okuda, Takashi; Minamiyama, Yukiko; Ichikawa, Hiroshi; Suenaga, Shigeaki; Oki, Misato; Sato, Tsuyoshi; Ozawa, Toshihiko; Clair, Daret K St; Majima, Hideyuki J

    2015-01-01

    Fridovich identified CuZnSOD in 1969 and manganese superoxide dismutase (MnSOD) in 1973, and proposed "the Superoxide Theory," which postulates that superoxide (O2 (•-)) is the origin of most reactive oxygen species (ROS) and that it undergoes a chain reaction in a cell, playing a central role in the ROS producing system. Increased oxidative stress on an organism causes damage to cells, the smallest constituent unit of an organism, which can lead to the onset of a variety of chronic diseases, such as Alzheimer's, Parkinson's, amyotrophic lateral sclerosis and other neurological diseases caused by abnormalities in biological defenses or increased intracellular reactive oxygen levels. Oxidative stress also plays a role in aging. Antioxidant systems, including non-enzyme low-molecular-weight antioxidants (such as, vitamins A, C and E, polyphenols, glutathione, and coenzyme Q10) and antioxidant enzymes, fight against oxidants in cells. Superoxide is considered to be a major factor in oxidant toxicity, and mitochondrial MnSOD enzymes constitute an essential defense against superoxide. Mitochondria are the major source of superoxide. The reaction of superoxide generated from mitochondria with nitric oxide is faster than SOD catalyzed reaction, and produces peroxynitrite. Thus, based on research conducted after Fridovich's seminal studies, we now propose a modified superoxide theory; i.e., superoxide is the origin of reactive oxygen and nitrogen species (RONS) and, as such, causes various redox related diseases and aging.

  6. Superoxide-dependent oxidation of melatonin by myeloperoxidase.

    PubMed

    Ximenes, Valdecir F; Silva, Sueli de O; Rodrigues, Maria R; Catalani, Luiz H; Maghzal, Ghassan J; Kettle, Anthony J; Campa, Ana

    2005-11-18

    Myeloperoxidase uses hydrogen peroxide to oxidize numerous substrates to hypohalous acids or reactive free radicals. Here we show that neutrophils oxidize melatonin to N(1)-acetyl-N(2)-formyl-5-methoxykynuramine (AFMK) in a reaction that is catalyzed by myeloperoxidase. Production of AFMK was highly dependent on superoxide but not hydrogen peroxide. It did not require hypochlorous acid, singlet oxygen, or hydroxyl radical. Purified myeloperoxidase and a superoxide-generating system oxidized melatonin to AFMK and a dimer. The dimer would result from coupling of melatonin radicals. Oxidation of melatonin was partially inhibited by catalase or superoxide dismutase. Formation of AFMK was almost completely eliminated by superoxide dismutase but weakly inhibited by catalase. In contrast, production of melatonin dimer was enhanced by superoxide dismutase and blocked by catalase. We propose that myeloperoxidase uses superoxide to oxidize melatonin by two distinct pathways. One pathway involves the classical peroxidation mechanism in which hydrogen peroxide is used to oxidize melatonin to radicals. Superoxide adds to these radicals to form an unstable peroxide that decays to AFMK. In the other pathway, myeloperoxidase uses superoxide to insert dioxygen into melatonin to form AFMK. This novel activity expands the types of oxidative reactions myeloperoxidase can catalyze. It should be relevant to the way neutrophils use superoxide to kill bacteria and how they metabolize xenobiotics.

  7. Superoxide Free Radicals Are Produced in Glyoxysomes 1

    PubMed Central

    Sandalio, Luisa M.; Fernández, Victor M.; Rupérez, Francisco L.; Del Río, Luis A.

    1988-01-01

    The production of superoxide free radicals in pellet and supernatant fractions of glyoxysomes, specialized plant peroxisomes from watermelon (Citrullus vulgaris Schrad.) cotyledons, was investigated. Upon inhibition of the endogenous superoxide dismutase, xanthine, and hypoxanthine induced in glyoxysomal supernatants the generation of O2− radicals and this was inhibited by allopurinol. In glyoxysomal pellets, NADH stimulated the generation of superoxide radicals. Superoxide production by purines was due to xanthine oxidase, which was found predominantly in the matrix of glyoxysomes. The generation of O2− radicals in glyoxysomes by endogenous metabolites suggests new active oxygen-related roles for glyoxysomes, and for peroxisomes in general, in cellular metabolism. PMID:16666081

  8. Induction of superoxide dismutases in Photobacterium leiognathi.

    PubMed

    Kobayashi, H; Tonokawa, H; Fukasawa, S; Yamakura, F

    1991-01-01

    We investigated the induction of Cu,Zn-SOD (bacteriocuprein) and Fe-SOD in Photobacterium leiognathi DK-A1 which was isolated from the light organ of the squid, Droteuthis kensaki. The induction of superoxide dismutases depended on the addition of paraquat to the medium. Induction of SOD by paraquat was attributed mostly to the bacteriocuprein by measuring of the activities of both SODs by using densitometry of isoelectrofocusing gel. When paraquat was added to the culture at various times in the early log phase of growth, the most efficient induction of the SODs, which was measured at the time of harvesting the cells (17 hours after inoculation), was observed when paraquat was added at 60 min after the inoculation. Catalase was not significantly induced by the addition of paraquat or increasing of oxygen concentration. We developed an assay of SOD by modification of a cytochrome c-xanthine oxidase method using a computer equipped absorption spectrophotometer.

  9. Regulation of pancreatic cancer growth by superoxide.

    PubMed

    Du, Juan; Nelson, Elke S; Simons, Andrean L; Olney, Kristen E; Moser, Justin C; Schrock, Hannah E; Wagner, Brett A; Buettner, Garry R; Smith, Brian J; Teoh, Melissa L T; Tsao, Ming-Sound; Cullen, Joseph J

    2013-07-01

    K-ras mutations have been identified in up to 95% of pancreatic cancers, implying their critical role in the molecular pathogenesis. Expression of K-ras oncogene in an immortalized human pancreatic ductal epithelial cell line, originally derived from normal pancreas (H6c7), induced the formation of carcinoma in mice. We hypothesized that K-ras oncogene correlates with increased non-mitochondrial-generated superoxide (O 2.-), which could be involved in regulating cell growth contributing to tumor progression. In the H6c7 cell line and its derivatives, H6c7er-Kras+ (H6c7 cells expressing K-ras oncogene), and H6c7eR-KrasT (tumorigenic H6c7 cells expressing K-ras oncogene), there was an increase in hydroethidine fluorescence in cell lines that express K-ras. Western blots and activity assays for the antioxidant enzymes that detoxify O 2.- were similar in these cell lines suggesting that the increase in hydroethidine fluorescence was not due to decreased antioxidant capacity. To determine a possible non-mitochondrial source of the increased levels of O 2.-, Western analysis demonstrated the absence of NADPH oxidase-2 (NOX2) in H6c7 cells but present in the H6c7 cell lines expressing K-ras and other pancreatic cancer cell lines. Inhibition of NOX2 decreased hydroethidine fluorescence and clonogenic survival. Furthermore, in the cell lines with the K-ras oncogene, overexpression of superoxide dismutases that detoxify non-mitochondrial sources of O 2.-, and treatment with the small molecule O 2.- scavenger Tempol, also decreased hydroethidine fluorescence, inhibited clonogenic survival and inhibited growth of tumor xenografts. Thus, O 2.- produced by NOX2 in pancreatic cancer cells with K-ras, may regulate pancreatic cancer cell growth. Copyright © 2012 Wiley Periodicals, Inc.

  10. Revisiting the reactions of superoxide with glutathione and other thiols.

    PubMed

    Winterbourn, Christine C

    2016-04-01

    The reaction between GSH and superoxide has long been of interest in the free radical biology. Early studies were confusing, as some reports suggested that the reaction could be a major pathway for superoxide removal whereas others questioned whether it happened at all. Further research by several investigators, including Helmut Sies, was required to clarify this complex reaction. We now know that superoxide does react with GSH, but the reaction is relatively slow and occurs mostly by a chain reaction that consumes oxygen and regenerates superoxide. Most of the GSH is converted to GSSG, with a small amount of sulfonic acid. As shown by Sies and colleagues, singlet oxygen is a by-product. Although removal of superoxide by GSH may be a minor pathway, GSH and superoxide have a strong physiological connection. GSH is an efficient free radical scavenger, and when it does so, thiyl radicals are generated. These further react to generate superoxide. Therefore, radical scavenging by GSH and other thiols is a source of superoxide and hydrogen peroxide, and to be an antioxidant pathway, there must be efficient removal of these species.

  11. Superoxide anion production by human neutrophils activated by Trichomonas vaginalis.

    PubMed

    Song, Hyun-Ouk; Ryu, Jae-Sook

    2013-08-01

    Neutrophils are the predominant inflammatory cells found in vaginal discharges of patients infected with Trichomonas vaginalis. In this study, we examined superoxide anion (O2 (.-)) production by neutrophils activated by T. vaginalis. Human neutrophils produced superoxide anions when stimulated with either a lysate of T. vaginalis, its membrane component (MC), or excretory-secretory product (ESP). To assess the role of trichomonad protease in production of superoxide anions by neutrophils, T. vaginalis lysate, ESP, and MC were each pretreated with a protease inhibitor cocktail before incubation with neutrophils. Superoxide anion production was significantly decreased by this treatment. Trichomonad growth was inhibited by preincubation with supernatants of neutrophils incubated for 3 hr with T. vaginalis lysate. Furthermore, myeloperoxidase (MPO) production by neutrophils was stimulated by live trichomonads. These results indicate that the production of superoxide anions and MPO by neutrophils stimulated with T. vaginalis may be a part of defense mechanisms of neutrophils in trichomoniasis.

  12. Superoxide dismutase activity in thermally stressed Staphylococcus aureus.

    PubMed Central

    Bucker, E R; Martin, S E

    1981-01-01

    The effects of heat and NaCl on the activity of superoxide dismutase from Staphylococcus aureus were examined. A linear decrease in superoxide dismutase activity occurred when S. aureus MF-31 cells were thermally stressed for 90 min at 52% C in 100 mM potassium phosphate buffer (pH 7.2). After 20 min of heating, only 5% of the superoxide dismutase activity was lost. Heating for 60, 90 and 120 min resulted in decreases of approximately 10, 22, and 68%, respectively. The rates of thermal inactivation of superoxide dismutase from S. aureus strains 196E and 210 were similar and slightly greater than those of strains MF-31, S-6, and 181. The addition of NaCl before or after heating resulted in increased losses of superoxide dismutase activity. PMID:7235693

  13. Manganese Superoxide Dismutase in Cancer Prevention

    PubMed Central

    Robbins, Delira

    2014-01-01

    Abstract Significance: Cancer is the second leading cause of death in the United States. Considering the quality of life and treatment cost, the best way to fight against cancer is to prevent or suppress cancer development. Cancer is preventable as indicated by human papilloma virus (HPV) vaccination and tamoxifen/raloxifen treatment in breast cancer prevention. The activities of superoxide dismutases (SODs) are often lowered during early cancer development, making it a rational candidate for cancer prevention. Recent Advances: SOD liposome and mimetics have been shown to be effective in cancer prevention animal models. They've also passed safety tests during early phase clinical trials. Dietary supplement-based SOD cancer prevention provides another opportunity for antioxidant-based cancer prevention. New mechanistic studies have revealed that SOD inhibits not only oncogenic activity, but also subsequent metabolic shifts during early tumorigenesis. Critical Issues: Lack of sufficient animal model studies targeting specific cancers; and lack of clinical trials and support from pharmaceutical industries also hamper efforts in further advancing SOD-based cancer prevention. Future Directions: To educate and obtain support from our society that cancer is preventable. To combine SOD-based therapeutics with other cancer preventive agents to obtain synergistic effects. To formulate a dietary supplementation-based antioxidant approach for cancer prevention. Lastly, targeting specific populations who are prone to carcinogens, which can trigger oxidative stress as the mechanism of carcinogenesis. Antioxid. Redox Signal. 20, 1628–1645. PMID:23706068

  14. Superoxide Dismutase as an Anaerobic Polypeptide 1

    PubMed Central

    Monk, Lorna S.; Fagerstedt, Kurt V.; Crawford, Robert M. M.

    1987-01-01

    The perennating organ, the rhizome, was chosen for examination of response to anoxia in the species Iris pseudacorus L., Iris germanica L. var Quechei, and Glyceria maxima (Hartm.) Holmberg. These monocots are known to differ in their tolerance of anoxia. Intact rhizomes were subjected to periods of prolonged anoxia of up to 28 days and superoxide dismutase (SOD) activity was determined in a 48 hour postanoxic recovery phase. Tests were performed to ensure the accuracy of the measured enzyme activities. In the most anoxia tolerant species, I. pseudacorus, SOD activity rose continuously during the period of imposed anoxia, and levels were maintained in the postanoxic recovery phases: 28 days brought about a 13-fold increase to 1576 U SOD per milligram protein. Small increases were found in the less anoxia tolerant I. germanica during anoxic/postanoxic phases, while a drop in activity was recorded in the least anoxia tolerant G. maxima. However, initial levels in G. maxima were more than twice as high as in the other two species. Experiments applying cycloheximide to anoxic rhizome slices of I. pseudacorus inhibited the increase in SOD activity. This indicates that SOD is, paradoxically, induced under anoxia and we suggest that in this species SOD is one of the enzymes identified as anaerobic polypeptides. The significance of the induction of an `oxygen-protecting' enzyme during complete oxygen deprivation is discussed with regard to a possible critical role during recovery from anoxic stress. PMID:16665795

  15. Manganese Superoxide Dismutase: Guardian of the Powerhouse

    PubMed Central

    Holley, Aaron K.; Bakthavatchalu, Vasudevan; Velez-Roman, Joyce M.; St. Clair, Daret K.

    2011-01-01

    The mitochondrion is vital for many metabolic pathways in the cell, contributing all or important constituent enzymes for diverse functions such as β-oxidation of fatty acids, the urea cycle, the citric acid cycle, and ATP synthesis. The mitochondrion is also a major site of reactive oxygen species (ROS) production in the cell. Aberrant production of mitochondrial ROS can have dramatic effects on cellular function, in part, due to oxidative modification of key metabolic proteins localized in the mitochondrion. The cell is equipped with myriad antioxidant enzyme systems to combat deleterious ROS production in mitochondria, with the mitochondrial antioxidant enzyme manganese superoxide dismutase (MnSOD) acting as the chief ROS scavenging enzyme in the cell. Factors that affect the expression and/or the activity of MnSOD, resulting in diminished antioxidant capacity of the cell, can have extraordinary consequences on the overall health of the cell by altering mitochondrial metabolic function, leading to the development and progression of numerous diseases. A better understanding of the mechanisms by which MnSOD protects cells from the harmful effects of overproduction of ROS, in particular, the effects of ROS on mitochondrial metabolic enzymes, may contribute to the development of novel treatments for various diseases in which ROS are an important component. PMID:22072939

  16. Manganese superoxide dismutase in cancer prevention.

    PubMed

    Robbins, Delira; Zhao, Yunfeng

    2014-04-01

    Cancer is the second leading cause of death in the United States. Considering the quality of life and treatment cost, the best way to fight against cancer is to prevent or suppress cancer development. Cancer is preventable as indicated by human papilloma virus (HPV) vaccination and tamoxifen/raloxifen treatment in breast cancer prevention. The activities of superoxide dismutases (SODs) are often lowered during early cancer development, making it a rational candidate for cancer prevention. SOD liposome and mimetics have been shown to be effective in cancer prevention animal models. They've also passed safety tests during early phase clinical trials. Dietary supplement-based SOD cancer prevention provides another opportunity for antioxidant-based cancer prevention. New mechanistic studies have revealed that SOD inhibits not only oncogenic activity, but also subsequent metabolic shifts during early tumorigenesis. Lack of sufficient animal model studies targeting specific cancers; and lack of clinical trials and support from pharmaceutical industries also hamper efforts in further advancing SOD-based cancer prevention. To educate and obtain support from our society that cancer is preventable. To combine SOD-based therapeutics with other cancer preventive agents to obtain synergistic effects. To formulate a dietary supplementation-based antioxidant approach for cancer prevention. Lastly, targeting specific populations who are prone to carcinogens, which can trigger oxidative stress as the mechanism of carcinogenesis.

  17. Peroxynitrite-mediated tyrosine nitration catalyzed by superoxide dismutase.

    PubMed

    Ischiropoulos, H; Zhu, L; Chen, J; Tsai, M; Martin, J C; Smith, C D; Beckman, J S

    1992-11-01

    Peroxynitrite (ONOO-), the reaction product of superoxide (O2-) and nitric oxide (NO), may be a major cytotoxic agent produced during inflammation, sepsis, and ischemia/reperfusion. Bovine Cu,Zn superoxide dismutase reacted with peroxynitrite to form a stable yellow protein-bound adduct identified as nitrotyrosine. The uv-visible spectrum of the peroxynitrite-modified superoxide dismutase was highly pH dependent, exhibiting a peak at 438 nm at alkaline pH that shifts to 356 nm at acidic pH. An equivalent uv-visible spectrum was obtained by Cu,Zn superoxide dismutase treated with tetranitromethane. The Raman spectrum of authentic nitrotyrosine was contained in the spectrum of peroxynitrite-modified Cu,Zn superoxide dismutase. The reaction was specific for peroxynitrite because no significant amounts of nitrotyrosine were formed with nitric oxide (NO), nitrogen dioxide (NO2), nitrite (NO2-), or nitrate (NO3-). Removal of the copper from the Cu,Zn superoxide dismutase prevented formation of nitrotyrosine by peroxynitrite. The mechanism appears to involve peroxynitrite initially reacting with the active site copper to form an intermediate with the reactivity of nitronium ion (NO2+), which then nitrates tyrosine on a second molecule of superoxide dismutase. In the absence of exogenous phenolics, the rate of nitration of tyrosine followed second-order kinetics with respect to Cu,Zn superoxide dismutase concentration, proceeding at a rate of 1.0 +/- 0.1 M-1.s-1. Peroxynitrite-mediated nitration of tyrosine was also observed with the Mn and Fe superoxide dismutases as well as other copper-containing proteins.

  18. Magnetoreception through Cryptochrome May Involve Superoxide

    PubMed Central

    Solov'yov, Ilia A.; Schulten, Klaus

    2009-01-01

    Abstract In the last decades, it has been demonstrated that many animal species orient in the Earth magnetic field. One of the best-studied examples is the use of the geomagnetic field by migratory birds for orientation and navigation. However, the biophysical mechanism underlying animal magnetoreception is still not understood. One theory for magnetoreception in birds invokes the so-called radical-pair model. This mechanism involves a pair of reactive radicals, whose chemical fate can be influenced by the orientation with respect to the magnetic field of the Earth through Zeeman and hyperfine interactions. The fact that the geomagnetic field is weak, i.e., ∼0.5 G, puts a severe constraint on the radical pair that can establish the magnetic compass sense. For a noticeable change of the reaction yield in a redirected geomagnetic field, the hyperfine interaction has to be as weak as the Earth field Zeeman interaction, i.e., unusually weak for an organic compound. Such weak hyperfine interaction can be achieved if one of the radicals is completely devoid of this interaction as realized in a radical pair containing an oxygen molecule as one of the radicals. Accordingly, we investigate here a possible radical pair-based reaction in the photoreceptor cryptochrome that reduces the protein's flavin group from its signaling state FADH• to the inactive state FADH– (which reacts to the likewise inactive FAD) by means of the superoxide radical, O2•–. We argue that the spin dynamics in the suggested reaction can act as a geomagnetic compass and that the very low physiological concentration (nM-μM) of otherwise toxic O2•– is sufficient, even favorable, for the biological function. PMID:19527640

  19. Manganese superoxide dismutase: beyond life and death

    PubMed Central

    Holley, Aaron K.; Dhar, Sanjit Kumar; Xu, Yong

    2010-01-01

    Manganese superoxide dismutase (MnSOD) is a nuclear-encoded antioxidant enzyme that localizes to the mitochondria. Expression of MnSOD is essential for the survival of aerobic life. Transgenic mice expressing a luciferase reporter gene under the control of the human MnSOD promoter demonstrate that the level of MnSOD is reduced prior to the formation of cancer. Overexpression of MnSOD in transgenic mice reduces the incidences and multiplicity of papillomas in a DMBA/TPA skin carcinogenesis model. However, MnSOD deficiency does not lead to enhanced tumorigenicity of skin tissue similarly treated because MnSOD can modulate both the p53-mediated apoptosis and AP-1-mediated cell proliferation pathways. Apoptosis is associated with an increase in mitochondrial levels of p53 suggesting a link between MnSOD deficiency and mitochondrial-mediated apoptosis. Activation of p53 is preventable by application of a SOD mimetic (MnTE-2-PyP5+). Thus, p53 translocation to mitochondria and subsequent inactivation of MnSOD explain the observed mitochondrial dysfunction that leads to transcription-dependent mechanisms of p53-induced apoptosis. Administration of MnTE-2-PyP5+ following apoptosis but prior to proliferation leads to suppression of protein carbonyls and reduces the activity of AP-1 and the level of the proliferating cellular nuclear antigen, without reducing the activity of p53 or DNA fragmentation following TPA treatment. Remarkably, the incidence and multiplicity of skin tumors are drastically reduced in mice that receive MnTE-2-PyP5+ prior to cell proliferation. The results demonstrate the role of MnSOD beyond its essential role for survival and suggest a novel strategy for an antioxidant approach to cancer intervention. PMID:20454814

  20. Mitochondrial Flashes: Dump Superoxide and Dance with Protons Now.

    PubMed

    Demaurex, Nicolas; Schwarzländer, Markus

    2016-09-20

    Transient changes in the physiology of individual mitochondria have recently drawn much interest. The use of a circular permuted yellow fluorescent protein (cpYFP) to monitor mitochondrial flashes and their interpretation as superoxide bursts has added confusion, however. Reviewing mitochondrial flashes in this Forum, Wang et al. again deem cpYFP to be a specific and reversible superoxide indicator, dismissing evidence that purified cpYFP is insensitive to superoxide. This interpretation lacks reproducible evidence and conflicts with the parsimony principle. We offer a constructive, transparent pathway to reach definitive clarification of contradictory reports. Antioxid. Redox Signal. 25, 550-551.

  1. Superoxide radical generation and Mn- and Cu-Zn superoxide dismutases activities in human leukemic cells.

    PubMed

    Kato, Masahiko; Minakami, Hisanori; Kuroiwa, Minoru; Kobayashi, Yasuyuki; Oshima, Shigeru; Kozawa, Kunihisa; Morikawa, Akihiro; Kimura, Hirokazu

    2003-03-01

    Mn- and Cu-Zn superoxide dismutase (SOD) activities and generation of superoxide radicals (O(2) (-)) were assessed in leukemic cells from 10 patients with acute myeloid or monocytic leukemia (AML) and 10 patients with acute lymphoblastic leukemia (ALL), using a sensitive, specific chemiluminescence method. Leukemic cells were classified according to the French-American-British classification. M4 AML cells from two patients produced some O(2) (-) upon stimulation with opsonized zymosan (OZ), phorbol myristate acetate (PMA) or N-formyl-methionyl-leucyl-phenylalanine (FMLP), but less than normal granulocytes or monocytes. M5b AML cells from one patient produced as much O(2) (-) in response to these stimulants as normal monocytes. No O(2) (-) generation was induced in other types of leukemic cells. Total SOD activity in AML cells was significantly greater in normal granulocytes, but was only half of the activity in ALL cells. Mn-SOD in AML cells was very low or undetectable. These results suggest that except in M5b cells, decreased O(2) (-) production may contribute to susceptibility to infections in AML patients. Decreased Mn-SOD activity in AML cells may predispose them to oxidative stress. Copyright 2002 John Wiley & Sons, Ltd.

  2. Phenotypes of mice lacking extracellular superoxide dismutase and copper- and zinc-containing superoxide dismutase.

    PubMed

    Sentman, Marie-Louise; Granström, Micael; Jakobson, Håkan; Reaume, Andrew; Basu, Samar; Marklund, Stefan L

    2006-03-17

    Mice lacking the secreted extracellular superoxide dismutase (EC-SOD) or the cytosolic copper- and zinc-containing SOD (CuZn-SOD) show relatively mild phenotypes. To explore the possibility that the isoenzymes have partly overlapping functions, single and double knockout mice were examined. The absence of EC-SOD was found to be without effect on the lifespan of mice, and the reduced lifespan of CuZn-SOD knockouts was not further shortened by EC-SOD deficiency. The urinary excretion of isoprostanes was increased in CuZn-SOD knockout mice, and plasma thiobarbituric acid-reactive substances levels were elevated in EC-SOD knockout mice. These oxidant stress markers showed potentiated increases in the absence of both isoenzymes. Other alterations were mainly found in CuZn-SOD knockout mice, such as halved glutathione peroxidase activity in the tissues examined and increased glutathione and iron in the liver. There were no changes in tissue content of the alternative superoxide scavenger ascorbate, but there was a 25% reduction in ascorbate in blood plasma in mice lacking CuZn-SOD. No increase was found in the urinary excretion of the terminal metabolites of NO, nitrite, and nitrate in any of the genotypes. In conclusion, apart from the increases in the global urinary and plasma oxidant stress markers, our phenotype studies revealed no other evidence that the copper- and zinc-containing SOD isoenzymes have overlapping roles.

  3. Negative air ions as a source of superoxide

    NASA Astrophysics Data System (ADS)

    Goldstein, Naum I.; Goldstein, Roman N.; Merzlyak, Mark N.

    1992-06-01

    The physico-chemical characteristics and possible formation mechanisms of negative air ions are considered. It was found that the products of oxygen and nitrogen negative ionization reduce ferricytochrome c and nitroblue tetrazolium, and that these reactions were inhibited by superoxide dismutase. The interaction of negatively ionized oxygen with water led to hydrogen peroxide accumulation, which was inhibited by tetranitromethane or catalase. Nitrogen ionization under these conditions caused the formation of the hydrated electron e{aq/—} and the superoxide anion O{2/—}. The data obtained indicate that the biological activity of negative air ions may be dependent on superoxide. The generation of reactive oxygen ions in the gas phase and also at a gas/water interface is described. A scheme for superoxide production under oxygen and nitrogen ionization is proposed.

  4. Superoxide-dependent cerebrovascular effects of homocysteine.

    PubMed

    Zhang, F; Slungaard, A; Vercellotti, G M; Iadecola, C

    1998-06-01

    Recent evidence indicates that elevated plasma levels of homocysteine are a risk factor for ischemic cerebrovascular diseases. However, little is known about cerebrovascular effects of homocysteine. Homocysteine could impair cerebrovascular function by metal-catalyzed production of activated oxygen species. We studied whether homocysteine, in the presence of Cu2+, alters reactivity of cerebral circulation and, if so, whether this effect depends on O-2 generation. In halothane-anesthetized rats the parietal cortex was exposed and superfused with Ringer solution. Cerebrocortical blood flow (CBF) was monitored by a laser-Doppler probe. With Ringer solution superfusion, CBF increased with hypercapnia (+134 +/- 7%; PCO2 = 50-60 mmHg) and topical application of 10 microM ACh (+35 +/- 3%), the NO donor S-nitroso-N-acetylpenicillamine (SNAP, 500 microM; +66 +/- 6%), or 1 mM papaverine (+100 +/- 6%; n = 5). Superfusion with 40 microM Cu2+ alone did not perturb resting CBF or responses to hypercapnia, ACh, SNAP, or papaverine (P > 0.05, n = 5). However, superfusion of homocysteine-Cu2+ reduced resting CBF (-28 +/- 4%) and attenuated (P < 0.05) responses to hypercapnia (-31 +/- 9%), ACh (-73 +/- 6%), or SNAP (-48 +/- 4%), but not papaverine. The effect was observed only at 1 mM homocysteine. Cerebrovascular effects of homocysteine-Cu2+ were prevented by coadministration of superoxide dismutase (SOD; 1,000 U/ml; n = 5). SOD alone did not affect resting CBF or CBF reactivity (n = 5). The observation that homocysteine-Cu2+ attenuates the response to hypercapnia, ACh, and SNAP, but not the NO-independent vasodilator papaverine, suggests that homocysteine-Cu2+ selectively impairs NO-related cerebrovascular responses. The fact that SOD prevents such impairment indicates that the effect of homocysteine is O-2 dependent. The data support the conclusion that O-2, generated by the reaction of homocysteine with Cu2+, inhibits NO-related cerebrovascular responses by scavenging NO

  5. Suppressors of superoxide production from mitochondrial complex III

    PubMed Central

    Orr, Adam L.; Vargas, Leonardo; Turk, Carolina N.; Baaten, Janine E.; Matzen, Jason T.; Dardov, Victoria J.; Attle, Stephen J.; Li, Jing; Quackenbush, Douglas C.; Goncalves, Renata L. S.; Perevoshchikova, Irina V.; Petrassi, H. Michael; Meeusen, Shelly L.; Ainscow, Edward K.; Brand, Martin D.

    2015-01-01

    Mitochondrial electron transport drives ATP synthesis but also generates reactive oxygen species (ROS), which are both cellular signals and damaging oxidants. Superoxide production by respiratory complex III is implicated in diverse signaling events and pathologies but its role remains controversial. Using high-throughput screening we identified compounds that selectively eliminate superoxide production by complex III without altering oxidative phosphorylation; they modulate retrograde signaling including cellular responses to hypoxic and oxidative stress. PMID:26368590

  6. The crystal structure of superoxide dismutase from Plasmodium falciparum

    PubMed Central

    Boucher, Ian W; Brzozowski, Andrzej M; Brannigan, James A; Schnick, Claudia; Smith, Derek J; Kyes, Sue A; Wilkinson, Anthony J

    2006-01-01

    Background Superoxide dismutases (SODs) are important enzymes in defence against oxidative stress. In Plasmodium falciparum, they may be expected to have special significance since part of the parasite life cycle is spent in red blood cells where the formation of reactive oxygen species is likely to be promoted by the products of haemoglobin breakdown. Thus, inhibitors of P. falciparum SODs have potential as anti-malarial compounds. As a step towards their development we have determined the crystal structure of the parasite's cytosolic iron superoxide dismutase. Results The cytosolic iron superoxide dismutase from P. falciparum (PfFeSOD) has been overexpressed in E. coli in a catalytically active form. Its crystal structure has been solved by molecular replacement and refined against data extending to 2.5 Å resolution. The structure reveals a two-domain organisation and an iron centre in which the metal is coordinated by three histidines, an aspartate and a solvent molecule. Consistent with ultracentrifugation analysis the enzyme is a dimer in which a hydrogen bonding lattice links the two active centres. Conclusion The tertiary structure of PfFeSOD is very similar to those of a number of other iron-and manganese-dependent superoxide dismutases, moreover the active site residues are conserved suggesting a common mechanism of action. Comparison of the dimer interfaces of PfFeSOD with the human manganese-dependent superoxide dismutase reveals a number of differences, which may underpin the design of parasite-selective superoxide dismutase inhibitors. PMID:17020617

  7. Superoxide-mediated dissolution of amorphous ferric oxyhydroxide in seawater.

    PubMed

    Fujii, Manabu; Rose, Andrew L; Waite, T David; Omura, Tatsuo

    2006-02-01

    We have investigated the kinetics of superoxide-mediated dissolution of amorphous ferric oxyhydroxide (AFO) in seawater by spectrophotometrically examining the rate of formation of a ferrous-ferrozine complex (Fe(II)(FZ)3) with particular attention given to the effect of aging and iron concentration on the rate of superoxide-mediated dissolution of AFO. The production rates of Fe(II)(FZ)3 decreased with aging of AFO for iron concentrations from 50 to 500 nM, indicating that changes to the chemical and physical properties of AFO affected the reactivity of inorganic ferric iron species with superoxide. A kinetic model developed by assuming that Fe(II)' formation is preceded by the thermal dissolution of AFO provided a good description of the Fe(II)(FZ)3 production rates over time. First-order rate constants for Fe(II)' formation were found to depend on the total iron concentration, suggesting that superoxide-mediated Fe(II) reduction is affected not only by the rate of thermal dissolution of AFO but also by the rate of AFO precipitation. The reported high rates of superoxide production by both photochemical and biotic pathways in aquatic systems coupled with the ability of superoxide to dissolve freshly formed ferric oxides suggest that this process may have a significant impact on the biogeochemical cycling of iron, especially if organisms have an affinity for ferrous iron.

  8. Clastogenic Factors as Potential Biomarkers of Increased Superoxide Production

    PubMed Central

    Emerit, Ingrid

    2007-01-01

    The formation of clastogenic factors (CF) and their damaging effects are mediated by superoxide, since superoxide dismutase is regularly protective. CF are produced via superoxide and stimulate the production of superoxide by monocytes and neutrophils. This results in a selfsustaining and longlasting process of clastogenesis, which may exceed the DNA repair system and ultimately lead to cancer (Emerit, 1994). An increased cancer risk is indeed observed in conditions accompanied by CF formation. These include irradiated persons, patients with chronic inflammatory diseases, HIV-infected persons and the chromosomal breakage syndromes ataxia telangiectasia, Bloom’s syndrome and Fanconi’s anemia. Biochemical analysis has identified lipid peroxidation products, arachidonic acid metabolites, nucleotides of inosine and cytokines, in particular tumor necrosis factor alpha, as the clastogenic and also superoxide stimulating components of CF. Due to their chromosome damaging effects, these oxidants can be detected with classical cytogenetic techniques. Their synergistic action renders the CF-test particularly sensitive for the detection of a pro-oxidant state. Correlations were observed between CF and other biomarkers of oxidative stress such as decreases in total plasma thiols or increases in TBARS or chemiluminescence. Correlations between CF and disease activity, between CF and radiation exposure, suggest the study of CF for monitoring these conditions. CF may also be useful as biochemical markers and intermediate endpoints for the evaluation of promising antioxidant drugs. CF formation represents a link between chronic inflammation and carcinogenesis. Prophylactic use of superoxide scavengers as anticarcinogens is therefore suggested. PMID:19662223

  9. Efficiency of superoxide anions in the inactivation of selected dehydrogenases

    NASA Astrophysics Data System (ADS)

    Rodacka, Aleksandra; Serafin, Eligiusz; Puchala, Mieczyslaw

    2010-09-01

    The most ubiquitous of the primary reactive oxygen species, formed in all aerobes, is the superoxide free radical. It is believed that the superoxide anion radical shows low reactivity and in oxidative stress it is regarded mainly as an initiator of more reactive species such as rad OH and ONOO -. In this paper, the effectiveness of inactivation of selected enzymes by radiation-generated superoxide radicals in comparison with the effectiveness of the other products of water radiolysis is examined. We investigate three enzymes: glyceraldehyde-3-phosphate dehydrogenase (GAPDH), alcohol dehydrogenase (ADH) and lactate dehydrogenase (LDH). We show that the direct contribution of the superoxide anion radical to GAPDH and ADH inactivation is significant. The effectiveness of the superoxide anion in the inactivation of GAPDH and ADG was only 2.4 and 2.8 times smaller, respectively, in comparison with hydroxyl radical. LDH was practically not inactivated by the superoxide anion. Despite the fact that the studied dehydrogenases belong to the same class of enzymes (oxidoreductases), all have a similar molecular weight and are tetramers, their susceptibility to free-radical damage varies. The differences in the radiosensitivity of the enzymes are not determined by the basic structural parameters analyzed. A significant role in inactivation susceptibility is played by the type of amino acid residues and their localization within enzyme molecules.

  10. Production of superoxide and activity of superoxide dismutase in rabbit epididymal spermatozoa.

    PubMed

    Holland, M K; Alvarez, J G; Storey, B T

    1982-12-01

    Mature rabbit spermatozoa from the cauda epididymidis suspended in potassium Tris phosphate buffer at 24 degrees C produced O2.-, as measured by reduction of acetylated ferricytochrome c, with an intrinsic rate of 0.20 nmol/min per 10(8) cells. This rate increased to 1.80 nmol/min per 10(8) cells in the presence of 10 mM cyanide. These spermatozoa contain 2.8 units per 10(8) cells of superoxide dismutase activity, 95% of which is sensitive, and 5% of which is insensitive, to cyanide inhibition. These activities correspond to the cytosolic Cu-Zn form and the mitochondrial Mn form of the dismutase, respectively. Only the cyanide-sensitive form is released from the sperm on hypo-osmotic treatment or sonication. Hypo-osmotically treated rabbit epididymal spermatozoa produced O2.- with an intrinsic rate of 0.24 nmol/min per 10(8) cells, which increased to 0.58 nmol/min per 10(8) cells in the presence of 10 mM cyanide. Both intact and hypo-osmotically treated cells react with O2.- in a second order reaction as inferred from the hyperbolic dependence on cell concentration of O2.- production rate in both the absence and presence of cyanide. The second order rate constant for this reaction with intact cells, kS, was calculated to be 22.9 X 10(-8) (cells/ml)-1 min-1 in its absence. For hypo-osmotically treated cells, the values of kS were 10.8 X 10(-8) (cells/ml)-1 min-1 and 8.2 X 10(-8) (cells/ml) -1 min-1, respectively. Since hypo-osmotically treated cells have lost much of their plasma membrane, the lower value of kS for the treated cells implies that this membrane is one site of reaction of O2.- with the cells. The increase in kS in the presence of cyanide, which inhibits superoxide dismutase and so increases O2.- production, suggests that the cells become more reactive with O2.- as its production rate increase, as would be expected for the occurrence of radical chain oxidation. This in turn suggests that superoxide dismutase plays a major role in protecting rabbit sperm

  11. A flow-system comparison of the reactivities of calcium superoxide and potassium superoxide with carbon dioxide and water vapor

    NASA Technical Reports Server (NTRS)

    Wood, P. C.; Ballou, E. V.; Spitze, L. A.; Wydeven, T.

    1982-01-01

    A single pass flow system was used to test the reactivity of calcium superoxide with respiratory gases and the performance was compared to that of potassium superoxide. The KO2 system is used by coal miners as a self-contained unit in rescue operations. Particular attention was given to the reactivity with carbon dioxide and water vapor at different temperatures and partial pressures of oxygen, carbon dioxide, and water vapor. The calcium superoxide beds were found to absorb CO2 and H2O vapor, releasing O2. The KO2 bed, however, released O2 at twice the rate of CO2 absorption at 37 C. It is concluded that the calcium superoxide material is not a suitable replacement for the KO2 bed, although Ca(O2)2 may be added to the KO2 bed to enhance the CO2 absorption.

  12. Skeletal muscle contractions induce acute changes in cytosolic superoxide, but slower responses in mitochondrial superoxide and cellular hydrogen peroxide.

    PubMed

    Pearson, Timothy; Kabayo, Tabitha; Ng, Rainer; Chamberlain, Jeffrey; McArdle, Anne; Jackson, Malcolm J

    2014-01-01

    Skeletal muscle generation of reactive oxygen species (ROS) is increased following contractile activity and these species interact with multiple signaling pathways to mediate adaptations to contractions. The sources and time course of the increase in ROS during contractions remain undefined. Confocal microscopy with specific fluorescent probes was used to compare the activities of superoxide in mitochondria and cytosol and the hydrogen peroxide content of the cytosol in isolated single mature skeletal muscle (flexor digitorum brevis) fibers prior to, during, and after electrically stimulated contractions. Superoxide in mitochondria and cytoplasm were assessed using MitoSox red and dihydroethidium (DHE) respectively. The product of superoxide with DHE, 2-hydroxyethidium (2-HE) was acutely increased in the fiber cytosol by contractions, whereas hydroxy-MitoSox showed a slow cumulative increase. Inhibition of nitric oxide synthases increased the contraction-induced formation of hydroxy-MitoSox only with no effect on 2-HE formation. These data indicate that the acute increases in cytosolic superoxide induced by contractions are not derived from mitochondria. Data also indicate that, in muscle mitochondria, nitric oxide (NO) reduces the availability of superoxide, but no effect of NO on cytosolic superoxide availability was detected. To determine the relationship of changes in superoxide to hydrogen peroxide, an alternative specific approach was used where fibers were transduced using an adeno-associated viral vector to express the hydrogen peroxide probe, HyPer within the cytoplasmic compartment. HyPer fluorescence was significantly increased in fibers following contractions, but surprisingly followed a relatively slow time course that did not appear directly related to cytosolic superoxide. These data demonstrate for the first time temporal and site specific differences in specific ROS that occur in skeletal muscle fibers during and after contractile activity.

  13. Sevoflurane exposure generates superoxide but leads to decreased superoxide during ischemia and reperfusion in isolated hearts.

    PubMed

    Kevin, Leo G; Novalija, Enis; Riess, Matthias L; Camara, Amadou K S; Rhodes, Samhita S; Stowe, David F

    2003-04-01

    Reactive oxygen species (ROS) are largely responsible for cardiac injury consequent to ischemia and reperfusion, but, paradoxically, there is evidence suggesting that anesthetics induce preconditioning (APC) by generating ROS. We hypothesized that sevoflurane generates the ROS superoxide (O(2)(.-)), that APC attenuates O(2)(.-) formation during ischemia, and that this attenuation is reversed by bracketing APC with the O(2)(.-) scavenger manganese (III) tetrakis (4-benzoic acid) porphyrin chloride (MnTBAP) or the putative mitochondrial adenosine triphosphate-sensitive potassium (mK(ATP)) channel blocker 5-hydroxydecanoate (5-HD). O(2)(.-) was measured continuously in guinea pig hearts by using dihydroethidium. Sevoflurane was administered alone (APC), with MnTBAP, or with 5-HD before 30 min of ischemia and 120 min of reperfusion. Control hearts underwent no pretreatment. Sevoflurane directly increased O(2)(.-); this was blocked by MnTBAP but not by 5-HD. O(2)(.-) increased during ischemia and during reperfusion. These increases in O(2)(.-) were attenuated in the APC group, but this was prevented by MnTBAP or 5-HD. We conclude that sevoflurane directly induces O(2)(.-) formation but that O(2)(.-) formation is decreased during subsequent ischemia and reperfusion. The former effect appears independent of mK(ATP) channels, but not the latter. Our study indicates that APC is initiated by ROS that in turn cause mK(ATP) channel opening. Although there appears to be a paradoxical role for ROS in triggering and mediating APC, a possible mechanism is offered. Reactive oxygen species (ROS) are implicated in triggering anesthetic preconditioning (APC). The ROS superoxide (O(2)(.-)) was measured continuously in guinea pig isolated hearts. Sevoflurane directly increased O(2)(.-) but led to attenuated O(2)(.-) formation during ischemia. This demonstrates triggering of APC by ROS and clarifies the mechanism of cardioprotection during ischemia.

  14. Superoxide modulates myogenic contractions of mouse afferent arterioles.

    PubMed

    Lai, En Yin; Wellstein, Anton; Welch, William J; Wilcox, Christopher S

    2011-10-01

    Reactive oxygen species enhance or impair autoregulation. Because superoxide is a vasoconstrictor, we tested the hypothesis that stretch generates superoxide that mediates myogenic responses. Increasing perfusion pressure of mouse isolated perfused renal afferent arterioles from 40 to 80 mm Hg reduced their diameter by 13.3±1.8% (P<0.001) and increased reactive oxygen species (ethidium: dihydroethidium fluorescence) by 9.8±2.3% (P<0.05). Stretch-induced fluorescence was reduced significantly (P<0.05) by incubation with Tempol (3.7±0.8%), pegylated superoxide dismutase (3.2±1.0%), or apocynin (3.5±0.9%) but not by pegylated catalase, L-nitroarginine methylester, or Ca(2+)-free medium, relating it to Ca(2+)-independent vascular superoxide. Compared with vehicle, basal tone and myogenic contractions were reduced significantly (P<0.05) by pegylated superoxide dismutase (5.4±0.8), Tempol (4.1±1.0%), apocynin (1.0±1.3%), and diphenyleneiodinium (3.9±0.9%) but not by pegylated catalase (10.1±1.6%). L-Nitroarginine methylester enhanced basal tone, but neither it (15.8±3.3%) nor endothelial NO synthase knockout (10.2±1.8%) significantly changed myogenic contractions. Tempol had no further effect after superoxide dismutase but remained effective after catalase. H(2)O(2) >50 μmol/L caused contractions but at 25 μmol/L inhibited myogenic responses (7.4±0.8%; P<0.01). In conclusion, increasing the pressure within afferent arterioles led to Ca(2+)-independent increased vascular superoxide production from nicotinamide adenine dinucleotide phosphate oxidase, which enhanced myogenic contractions largely independent of NO, whereas H(2)O(2) impaired pressure-induced contractions but was not implicated in the normal myogenic response.

  15. Superoxide Dismutase and Oxygen Toxicity in a Eukaryote

    PubMed Central

    Gregory, Eugene M.; Goscin, Stephen A.; Fridovich, Irwin

    1974-01-01

    Saccharomyces cerevisiae var. ellipsoideus contained 6.5 times more superoxide dismutase and 2.3 times more catalase when grown under 100% O2 than when grown anaerobically. Growth under oxygen caused equal increases in both the cyanide-sensitive and the cyanide-insensitive superoxide dismutases of this organism. Experience with other eukaryotes has shown that cyanide sensitivity is a property of the cupro-zinc superoxide dismutase of the cytosol, whereas cyanide insensitivity is a property of the corresponding mangani-enzyme found in mitochondria. Cu2+, which has been shown to increase the radioresistance of yeast, also caused an increase of both of the superoxide dismutases of S. cerevisiae. Yeast which had been grown under 1 atm of O2 were more resistant toward the lethal effects of 20 atm of O2 than were yeast which had been grown in the absence of O2. Escherichia coli K-12 his− responded to growth under 1 atm of O2 by increasing its content of catalase and of peroxidase, but not of superoxide dismutase. This contrasts with E. coli B, which was previously shown to respond to O2 by a striking increase in superoxide dismutase. E. coli K-12 his− did not gain resistance toward 20 atm of O2 because of having been grown under 1 atm of O2. Once again, this contrasts with the behavior of E. coli B. These data indicate that, in both prokaryotes and in eukaryotes, superoxide dismutase is an important component of the defenses against oxygen toxicity. PMID:4590469

  16. Photodynamic production of superoxide in vitro by altertoxins in the presence of reducing agents.

    PubMed

    Hartman, P E; Suzuki, C K; Stack, M E

    1989-01-01

    Superoxide production by the three 4,9-dihydroxyperylene-3,10-quinone fungal toxins, altertoxins I, II, and III, was stimulated on illumination with broad-spectrum light. As determined previously for cercosporin, superoxide production by illuminated altertoxins was increased by the addition of the reducing substances ergothioneine or urate; ascorbate also effectively increased superoxide production. Illuminated urate alone engendered some superoxide production.

  17. Cytokines directly induce degranulation and superoxide production from human eosinophils.

    PubMed

    Horie, S; Gleich, G J; Kita, H

    1996-08-01

    Cytokines are implicated in allergic diseases and can modulate effector functions of eosinophils stimulated by another agonist. However, little is known about the capacity of cytokines to directly trigger eosinophil degranulation. We attempted to determine whether cytokines can directly induce degranulation and superoxide production from eosinophils. Eosinophils from normal donors were incubated with various cytokines in albumin-coated tissue culture plates for 4 hours. To quantitate degranulation, the amounts of eosinophil-derived neurotoxin in supernatants were measured by radioimmunoassay. In addition, superoxide production was measured by superoxide dismutase-inhibitable reduction of cytochrome c. IL-5, IL-3, granulocyte-macrophage colony-stimulating factor, tumor necrosis factor- alpha, and RANTES all induced eosinophil degranulation. Granulocyte-macrophage colony-stimulating factor was the most potent and induced eosinophil-derived neurotoxin release comparable to that induced by secretory IgA beads, one of the most potent secretagogues for eosinophils. In addition, IL-5 and tumor necrosis factor- alpha were synergistic in their induction of eosinophil degranulation. In contrast, IL-1, IL-8, interferon- gamma, and macrophage inflammatory protein-1 alpha did not induce degranulation. Finally, IL-5, IL-3, granulocyte-macrophage colony-stimulating factor, and tumor necrosis factor- alpha, but not RANTES, also induced superoxide production from eosinophils. Certain cytokines directly induce eosinophil degranulation and superoxide production in vitro. Therefore these cytokines may be important in the release of toxic granule proteins from eosinophils in allergic diseases.

  18. Formation of manganese oxides by bacterially generated superoxide

    NASA Astrophysics Data System (ADS)

    Learman, D. R.; Voelker, B. M.; Vazquez-Rodriguez, A. I.; Hansel, C. M.

    2011-02-01

    Manganese oxide minerals are among the strongest sorbents and oxidants in the environment. The formation of these minerals controls the fate of contaminants, the degradation of recalcitrant carbon, the cycling of nutrients and the activity of anaerobic-based metabolisms. Oxidation of soluble manganese(II) ions to manganese(III/IV) oxides has been primarily attributed to direct enzymatic oxidation by microorganisms. However, the physiological reason for this process remains unknown. Here we assess the ability of a common species of marine bacteria-Roseobacter sp. AzwK-3b-to oxidize manganese(II) in the presence of chemical and biological inhibitors. We show that Roseobacter AzwK-3b oxidizes manganese(II) by producing the strong and versatile redox reactant superoxide. The oxidation of manganese(II), and concomitant production of manganese oxides, was inhibited in both the light and dark in the presence of enzymes and metals that scavenge superoxide. Oxidation was also inhibited by various proteases, enzymes that break down bacterial proteins, confirming that the superoxide was bacterially generated. We conclude that bacteria can oxidize manganese(II) indirectly, through the enzymatic generation of extracellular superoxide radicals. We suggest that dark bacterial production of superoxide may be a driving force in metal cycling and mineralization in the environment.

  19. The superoxide reductase from the early diverging eukaryote Giardia intestinalis.

    PubMed

    Testa, Fabrizio; Mastronicola, Daniela; Cabelli, Diane E; Bordi, Eugenio; Pucillo, Leopoldo P; Sarti, Paolo; Saraiva, Lígia M; Giuffrè, Alessandro; Teixeira, Miguel

    2011-10-15

    Unlike superoxide dismutases (SODs), superoxide reductases (SORs) eliminate superoxide anion (O(2)(•-)) not through its dismutation, but via reduction to hydrogen peroxide (H(2)O(2)) in the presence of an electron donor. The microaerobic protist Giardia intestinalis, responsible for a common intestinal disease in humans, though lacking SOD and other canonical reactive oxygen species-detoxifying systems, is among the very few eukaryotes encoding a SOR yet identified. In this study, the recombinant SOR from Giardia (SOR(Gi)) was purified and characterized by pulse radiolysis and stopped-flow spectrophotometry. The protein, isolated in the reduced state, after oxidation by superoxide or hexachloroiridate(IV), yields a resting species (T(final)) with Fe(3+) ligated to glutamate or hydroxide depending on pH (apparent pK(a)=8.7). Although showing negligible SOD activity, reduced SOR(Gi) reacts with O(2)(•-) with a pH-independent second-order rate constant k(1)=1.0×10(9) M(-1) s(-1) and yields the ferric-(hydro)peroxo intermediate T(1); this in turn rapidly decays to the T(final) state with pH-dependent rates, without populating other detectable intermediates. Immunoblotting assays show that SOR(Gi) is expressed in the disease-causing trophozoite of Giardia. We propose that the superoxide-scavenging activity of SOR in Giardia may promote the survival of this air-sensitive parasite in the fairly aerobic proximal human small intestine during infection.

  20. Characterization of superoxide production by isolated pea thylakoids

    SciTech Connect

    Grace, S.; Osmond, B. )

    1991-05-01

    During photosynthesis chloroplasts univalently reduce molecular oxygen to superoxide through autoxidations in the electron transport chain. Cytochrome c reduction was used to assay superoxide production in illuminate pea thylakoids under a variety of conditions. Superoxide dismutase was found to inhibit the reaction by 80%, indicating that cytochrome c reduction is primarily mediated by superoxide. This was further supported by the observation that the highest rates of cytochrome c reduction occurred in the presence of methyl viologen, an autoxidizable redox carrier that accepts electrons from photosystem I. The reaction was fully suppressed by DCMU, demonstrating a requirement for electron transport. In the presence of the plastoquinone antagonist DBMIB the rate of cytochrome c reduction increased substantially. This indicates that under conditions where electron transport to photosystem I is blocked, autoxidation reactions can occur on the reducing side of photosystem II to maintain Q{sub A} in the oxidized state. Superoxide production at sites other than the reducing side of photosystem I may thus represent an important pathway for dissipating excess excitation energy.

  1. Superoxide production in aprotic interior of chloroplast thylakoids.

    PubMed

    Takahashi, M; Asada, K

    1988-12-01

    The site of superoxide production in spinach thylakoids was found to be the aprotic interior of the thylakoid membranes near the P700 chlorophyll a protein at the reaction center of photosystem I complexes. This conclusion was drawn from the following findings. (i) Cytochrome c reduction by illuminated thylakoids, which was confirmed to be superoxide dependent by the failure of this reaction to occur in anaerobiosis, was completely inhibited by a dibutyl catechol, but partially inhibited by a hydrophilic disulfonated derivative. (ii) P700 chlorophyll a proteins were preferentially iodinated by lactoperoxidase by the use of hydrogen peroxide that was derived from the disproportionation of superoxides in illuminated thylakoids. (iii) Hydrogen peroxide production and oxygen uptake were induced by ammonium chloride, a proton conductor that can permeate through thylakoid membranes, but whole superoxide in the bulk solution was oxidized back to molecular oxygen by cytochrome c. The effective concentration of ammonium chloride decreased to one-sixtieth of the original, when an ammonium ion ionophore, nonactin, was added. Thus, the weak acid allowed superoxide to yield hydrogen peroxide disproportionately in the thylakoid membrane interior.

  2. Evidence for increased mitochondrial superoxide production in Down syndrome.

    PubMed

    Capone, George; Kim, Peter; Jovanovich, Sole; Payne, Lynda; Freund, Lisa; Welch, Kimberly; Miller, Elizabeth; Trush, Michael

    2002-05-03

    Respiring mitochondria represent the major source of superoxide production in most cells, and superoxide anions function as direct precursors of hydrogen peroxide formation within mitochondria. We use a lucigenen-derived chemiluminescence (LDCL) assay to test the hypothesis that intramitochondrial superoxide production is altered in young children with DS. We also measured the levels of two serum markers of lipid peroxidation, lipid peroxides (LOOH), and malondialdehyde as thiobarbituric acid reactive substances (TBARS), to determine if superoxide levels correlate with in vivo measures of lipid peroxidation. A three-group, cross-sectional design was utilized which allowed us to compare young children with DS to children with cognitive impairment (CI) of unknown etiology, and typically developing (Nl) children. Data was analyzed using Pearson's zero-order correlations and multivariate analysis of variance (MANOVA) with Bonferroni correction for multiple comparisons. DS subjects had significantly elevated LDCL signal compared to Nl subjects (p = .03), but did not differ significantly from CI subjects. This study provides new evidence regarding an important source of reactive oxygen species in trisomy 21. The role of the mitochondria in superoxide anion production and the mechanisms underlying its generation in DS deserves further study.

  3. The sites and topology of mitochondrial superoxide production

    PubMed Central

    Brand, Martin D.

    2010-01-01

    Mitochondrial superoxide production is an important source of reactive oxygen species in cells, and may cause or contribute to ageing and the diseases of ageing. Seven major sites of superoxide production in mammalian mitochondria are known and widely accepted. In descending order of maximum capacity they are the ubiquinone binding sites in complex I (site IQ) and complex III (site IIIQo), glycerol 3-phosphate dehydrogenase, the flavin in complex I (site IF), the electron transferring flavoprotein:Q oxidoreductase (ETFQOR) of fatty acid beta oxidation, and pyruvate and 2-oxoglutarate dehydrogenases. None of these sites is fully characterized and for some we only have sketchy information. The topology of the sites is important because it determines whether or not a site will produce superoxide in the mitochondrial matrix and be able to damage mitochondrial DNA. All sites produce superoxide in the matrix; site IIIQo and glycerol 3-phosphate dehydrogenase also produce superoxide to the intermembrane space. The relative contribution of each site to mitochondrial reactive oxygen species generation in the absence of electron transport inhibitors is unknown in isolated mitochondria, in cells or in vivo, and may vary considerably with species, tissue, substrate, energy demand and oxygen tension. PMID:20064600

  4. Superoxide dismutase and O2 lethality in Bacteroides fragilis.

    PubMed Central

    Privalle, C T; Gregory, E M

    1979-01-01

    Exposure of midlog Bacteroides fragils (VPI 2393) to 2% O2-98% N2 caused a three- to fivefold increase in superoxide dismutase specific activity within the cells. The increase in specific activity was completed within 90 min after exposure to oxygen and was dependent upon protein synthesis. Cells containing the higher superoxide dismutase level were more resistant to the effects of 5 atm of oxygen tension than were cells containing the lower level of superoxide dismutase but were equally resistant to 5 atm of nitrogen tension. Similar results were observed upon comparing viability experiments with B. fragilis and B. vulgatus. Superoxide dismutase activity in sonic extracts of B. fragilis was rapidly inactivated by exposure to 5 mM H2O2 and was inhibited by 1 mM NaN3 but not 5 mM NaCN. The inhibition pattern is identical to the pattern demonstrated for the purified iron-containing enzyme from Escherichia coli B and suggests that the superoxide dismutase in B. fragilis is an iron enzyme. PMID:438129

  5. Superoxide generation from human polymorphonuclear leukocytes by liposome-encapsulated hemoglobin.

    PubMed

    Abe, H; Ikebuchi, K; Niwa, K; Inanami, O; Kuwabara, M; Fujihara, M; Hirayama, J; Ikeda, H

    2001-07-01

    We investigated the interactions between liposome-encapsulated hemoglobin (Neo Red Cells: NRC) and human polymorphonuclear leukocytes as assessed by superoxide generation. NRC triggered superoxide generation from neutrophils in a dose-dependent manner. Empty liposomes also induced superoxide production of neutrophils. Superoxide generation of neutrophils induced by phorbol myristate acetate (PMA) was delayed but intensified both by NRC and empty liposomes. The intensity of superoxide generation induced by NRC was smaller than that by the empty liposomes. As NRC contained superoxide dismutase (SOD) that was copurified with hemoglobin from red blood cells and its activity remained, SOD contained in NRC may partially eliminate superoxide.

  6. Induction of Cu,Zn-superoxide dismutase after cortical contusion injury during hypothermia.

    PubMed

    Fukuhara, T; Nishio, S; Ono, Y; Kawauchi, M; Asari, S; Ohmoto, T

    1994-09-19

    To determine the effect of hypothermia on superoxide injury after cerebral contusion, the induction of Cu,Zn-superoxide dismutase was examined 6 h after contusion in rats using Northern blotting. Cu,Zn-superoxide dismutase gene expression increased at the periphery of the contusion, which may indicate the severity of the superoxide stimulus. This increase was preserved after contusion under hypothermia, which may show that superoxide injury is still severe although brain edema is decreased.

  7. Bleaching of the red anthocyanin induced by superoxide radical.

    PubMed

    Yamasaki, H; Uefuji, H; Sakihama, Y

    1996-08-01

    Red anthocyanin prepared from petals of Hibiscus rosa-sinensis L. was photobleached in the EDTA-riboflavin system. The rate of bleaching monitored at 565 nm depended on the light intensity and EDTA concentrations. Anaerobic conditions and/or addition of superoxide dismutase prevented the bleaching of anthocyanin, whereas mannitol and catalase did not. A similar bleaching was observed under dark conditions in the xanthine-xanthine oxidase system. The results indicate that anthocyanin is bleached by the nonenzymatic reaction with the superoxide radical and suggest that the pigment can function as an antioxidant. The antioxidative efficiency of cyanidin to superoxide was 10-fold higher than that of cyanidin-3-sophoroside as a Hibiscus anthocyanin.

  8. Regulation of superoxide dismutase synthesis in Candida albicans.

    PubMed

    Gunasekaran, U; Yang, R; Gunasekaran, M

    1998-01-01

    The synthesis of superoxide dismutase [SOD: EC 1.15.1.1] in response to various cultural conditions was examined in Candida albicans, an opportunistic yeast which causes candidiasis in immunosuppressed patients. SOD plays an important role in protecting cells from teh oxidative damage of superoxide radicals. Maximum SOD activity was found after 72 hrs of yeast growth. The optimum pH and temperature for the SOD activity were 7 and 40 degrees C, respectively. The major SOD activity was found in the cytosol fraction and the level of extracellular SOD was very low. The enzyme was stimulated to varying degrees by cholic acid, procaine and tocopherol. On the basis of inhibitor studies and other enzyme properties, the isolated enzyme from C. albicans is identified as a copper and zinc superoxide dismutase.

  9. The preparation of calcium superoxide from calcium peroxide diperoxyhydrate

    NASA Technical Reports Server (NTRS)

    Ballou, E. V.; Wood, P. C.; Spitze, L. A.; Wydeven, T.

    1977-01-01

    There is interest in solid materials containing a high percentage of stored oxygen for use in emergency breathing apparatus for miners and as auxiliary oxygen sources for astronauts. In theory, the amount of available oxygen in calcium superoxide, Ca(O2)2 is higher than in potassium superoxide, KO2, and its availability during use should be unhindered by the formation of a low melting and hydrous coating. The decomposition of solid calcium peroxide diperoxyhydrate, CaO2.2H2O2 has been studied, using an apparatus which allows good control of the critical reaction parameters. Samples have been prepared showing apparent superoxide contents in excess of those previously reported and higher than the theoretical 58.4% expected from a disproportionation reaction.

  10. Bactericidal Activity of Superoxide Anion and of Hydrogen Peroxide: Investigations Employing Dialuric Acid, a Superoxide-Generating Drug

    PubMed Central

    DeChatelet, Lawrence R.; Shirley, Pamela S.; Goodson, Phillip R.; McCall, Charles E.

    1975-01-01

    The addition of dialuric acid (a superoxide-generating drug) to a suspension of resting human neutrophils resulted in a stimulation of cellular hexose monophosphate shunt activity. Measurement of oxygen consumption demonstrated a rapid rate of oxygen uptake by the drug alone in aqueous solution. The subsequent addition of catalase (but not of superoxide dismutase) resulted in a substantial release of oxygen, indicating that H2O2 was accumulating in the media. The generation of O2− by the drug was verified by measuring the rate of reduction of cytochrome c by the drug in the presence and absence of authentic superoxide dismutase. The growth of Escherichia coli B and Staphylococcus aureus was inhibited in vitro by addition of the drug to a suitable culture media; the degree of inhibition was dose dependent. This inhibition of bacterial growth could be completely reversed by the addition of as little as 5 μg of purified catalase, but was not affected by concentrations of superoxide dismutase that were 2,000-fold higher. These results suggest that the dialuric acid-induced stimulation of hexose monophosphate shunt activity in neutrophils is due to accumulated H2O2. Further, the contribution of superoxide to the bactericidal activity towards E. coli and S. aureus is probably minimal compared with that of H2O2. PMID:170855

  11. Constraints on superoxide mediated formation of manganese oxides

    PubMed Central

    Learman, Deric R.; Voelker, Bettina M.; Madden, Andrew S.; Hansel, Colleen M.

    2013-01-01

    Manganese (Mn) oxides are among the most reactive sorbents and oxidants within the environment, where they play a central role in the cycling of nutrients, metals, and carbon. Recent discoveries have identified superoxide (O2−) both of biogenic and abiogenic origin as an effective oxidant of Mn(II) leading to the formation of Mn oxides. Here we examined the conditions under which abiotically produced superoxide led to oxidative precipitation of Mn and the solid-phases produced. Oxidized Mn, as both aqueous Mn(III) and Mn(III/IV) oxides, was only observed in the presence of active catalase, indicating that hydrogen peroxide (H2O2), a product of the reaction of O2− with Mn(II), inhibits the oxidation process presumably through the reduction of Mn(III). Citrate and pyrophosphate increased the yield of oxidized Mn but decreased the amount of Mn oxide produced via formation of Mn(III)-ligand complexes. While complexing ligands played a role in stabilizing Mn(III), they did not eliminate the inhibition of net Mn(III) formation by H2O2. The Mn oxides precipitated were highly disordered colloidal hexagonal birnessite, similar to those produced by biotically generated superoxide. Yet, in contrast to the large particulate Mn oxides formed by biogenic superoxide, abiotic Mn oxides did not ripen to larger, more crystalline phases. This suggests that the deposition of crystalline Mn oxides within the environment requires a biological, or at least organic, influence. This work provides the first direct evidence that, under conditions relevant to natural waters, oxidation of Mn(II) by superoxide can occur and lead to formation of Mn oxides. For organisms that oxidize Mn(II) by producing superoxide, these findings may also point to other microbially mediated processes, in particular enzymatic hydrogen peroxide degradation and/or production of organic ligand metabolites, that allow for Mn oxide formation. PMID:24027565

  12. Ferric human neuroglobin scavenges superoxide to form oxy adduct.

    PubMed

    Yamashita, Taku; Hafsi, Leila; Masuda, Eri; Tsujino, Hirofumi; Uno, Tadayuki

    2014-01-01

    Neuroglobin (Ngb) is the third member of the vertebrate globin family, and the structure was solved as a typical globin fold with a b-type heme. Although it has been proposed that Ngb could be involved in neuroprotection against oxidative stress, the protective mechanism has not been fully identified yet. In order to clarify functions under hypoxic condition, in this study, we focused on the scavenger activity of human Ngb (hNgb) against superoxide. The activity of hNgb for superoxide was evaluated to be 7.4 µM for IC50, the half maximal inhibitory concentration. The result indicates that hNgb can be an anti-oxidant, and the value was almost the same as that of ascorbic acid. In addition, we characterized oxidation states of a heme iron in superoxide-treated hNgb with spectroscopic measurements. Superoxide-treated hNgb in the ferric form was readily converted to the oxygenated ferrous form, and the result suggested that ferric hNgb could scavenge superoxide by change of an oxidation state in a heme iron. Moreover, mutational experiments were performed, and the each variant mutated at 46 and 55 positions suggested a disulfide bond between Cys46 and Cys55 could be essential to be sensors for oxidative stress with the direct binding of superoxide. As a consequence, we concluded that redox changes of the heme iron and the disulfide bond could regulate neuroprotective functions of hNgb, and it suggests that hNgb can afford protection against hypoxic and ischemic stress in the brain.

  13. Ethanol stimulates superoxide production and inhibits phorbol ester induced superoxide production in alveolar macrophages

    SciTech Connect

    Dorio, R.J.; Hoek, J.B.; Forman, H.J.; Rubin, E.

    1986-05-01

    Ethanol stimulates superoxide (O/sub 2//sup -/) production in rat alveolar macrophages. Increasing the ethanol concentration from 75 to 500 mM produces a linear dose response curve, generating between 10 and 30 pmol O/sub 2//sup -//min/10/sup 6/ cells. Thus, ethanol is a weak agonist of O/sub 2//sup -/ in these cells. Pretreatment with ethanol in the same concentration range results in a dose and time dependent inhibition of O/sub 2//sup -/ production by phorbol-12-myristate-13-acetate (PMA). 100 mM ethanol inhibits PMA (100 ng/ml)-induced O/sub 2//sup -/ production by 60% after 5 minutes and by 80% after 30 minutes of preincubation. At lower concentrations (10-25 mM), however, ethanol causes a synergistic stimulation of PMA-induced O/sub 2//sup -/ production. Preincubation for 15 minutes with 10 mM ethanol results in a 20% increase in PMA-induced O/sub 2//sup -/ production. Synergism between PMA and ethanol is seen at ethanol concentrations which do not result in O/sub 2//sup -/ production by ethanol alone. This synergism is abolished by a 15 minute preincubation of the cells in EGTA. Thus, ethanol acts as a weak agonist for O/sub 2//sup -/ production and interacts significantly with PMA-induced stimulation of O/sub 2//sup -/ production.

  14. Reactions between microhydrated superoxide anions and formic acid.

    PubMed

    Ryding, Mauritz Johan; Fernández, Israel; Uggerud, Einar

    2017-08-30

    Reactions between water clusters containing the superoxide anion, O2˙(-)(H2O)n (n = 0-4), and formic acid, HCO2H, were studied experimentally in vacuo and modelled using quantum chemical methods. Encounters between microhydrated superoxide and formic acid were found to result in a number of different reactions, including (a) proton transfer, (b) ligand exchange, (c) H2-elimination (affording microhydrated CO4˙(-)), and (d) dihydrogen transfer (affording H2O2 and microhydrated CO2˙(-)). The effect of reactant-ion hydration on reaction rates was investigated and the involved reaction mechanisms were elucidated.

  15. A novel murrel Channa striatus mitochondrial manganese superoxide dismutase: gene silencing, SOD activity, superoxide anion production and expression.

    PubMed

    Arockiaraj, Jesu; Palanisamy, Rajesh; Bhatt, Prasanth; Kumaresan, Venkatesh; Gnanam, Annie J; Pasupuleti, Mukesh; Kasi, Marimuthu

    2014-12-01

    We have reported the molecular characterization including gene silencing, superoxide activity, superoxide anion production, gene expression and molecular characterization of a mitochondrial manganese superoxide dismutase (mMnSOD) from striped murrel Channa striatus (named as CsmMnSOD). The CsmMnSOD polypeptide contains 225 amino acids with a molecular weight of 25 kDa and a theoretical isoelectric point of 8.3. In the N-terminal region, CsmMnSOD carries a mitochondrial targeting sequence and a superoxide dismutases (SOD) Fe domain (28-109), and in C-terminal region, it carries another SOD Fe domain (114-220). The CsmMnSOD protein sequence shared significant similarity with its homolog of MnSOD from rock bream Oplegnathus fasciatus (96%). The phylogenetic analysis showed that the CsmMnSOD fell in the clade of fish mMnSOD group. The monomeric structure of CsmMnSOD possesses 9 α-helices (52.4%), 3 β-sheets (8.8%) and 38.8% random coils. The highest gene expression was noticed in liver, and its expression was inducted with fungal (Aphanomyces invadans) and bacterial (Aeromonas hydrophila) infections. The gene silencing results show that the fish that received dsRNA exhibited significant (P < 0.05) changes in expression when compared to their non-injected and fish physiological saline-injected controls. The SOD activity shows that the activity increases with the spread of infection and decreases once the molecule controls the pathogen. The capacity of superoxide anion production was determined by calculating the granular blood cell count during infection in murrel. It shows that the infection influenced the superoxide radical production which plays a major role in killing the pathogens. Overall, this study indicated the defense potentiality of CsmMnSOD; however, further research is necessary to explore its capability at protein level.

  16. Studies with primaquine in vitro: superoxide radical formation and oxidation of haemoglobin.

    PubMed Central

    Summerfield, M; Tudhope, G R

    1978-01-01

    1. The production of superoxide radicals from primaquine diphosphate in aqueous solution has been demonstrated, using as indicator the reduction of cytochrome C with inhibition of the reaction by superoxide dismutase. 2. Primaquine-mediated oxidation of haemoglobin to methaemoglobin was reduced by the addition of catalase and increased by superoxide dismutase. Mannitol, a hydroxyl radical scavenger, abolished the increase in methaemoglobin observed in the presence of superoxide dismutase. EDTA reduced the oxidation of haemoglobin with and without superoxide dismutase. 3. Although the oxidation of haemoglobin in the presence of primaquine includes the effects of hydrogen peroxide, superoxide and hydroxyl radicals and metal ions, the results indicate that hydrogen peroxide, rather than the superoxide radical, is the main oxidizing species. The increase in haemoglobin oxidation occurring with superoxide dismutase may result from the augmented rate of hydrogen peroxide formation from superoxide radicals. PMID:212091

  17. Coelenterazine analogs as chemiluminescent probe for superoxide anion.

    PubMed

    Teranishi, K; Shimomura, O

    1997-06-15

    Eleven new coelenterazine analogs containing the 3,7-dihydroimidazo[1,2-alpha]pyrazin-3-one structure were synthesized. The superoxide-triggered chemiluminescence of these compounds was investigated using the hypoxanthine-xanthine oxidase system in comparison with four known compounds. The results showed that an alkyl substitution at the position 5 of the imidazopyrazinone ring causes a drastic decrease in the superoxide-dependent chemiluminescence intensity, whereas a dimethylene bridge added between the position 5 and the phenyl group bound to the position 6 dramatically increases the luminescence intensity, indicating the potential usefulness of this type of compound as a probe for superoxide anion. The luminescence intensity of the bridged analog was 33 times greater than that of MCLA [2-methyl-6-(4-methoxyphenyl)-3, 7-dihydroimidazo[1,2-alpha]pyrazin-3-one], the most sensitive superoxide probe of Cypridina luciferin-type. Two of the analogs synthesized, each with a covalently bound cyclodextrin, had a good solubility in water, an advantage in actual use. Moreover, one of them having a beta-cyclodextrin group showed a unique property; its luminescence was little affected by various substances in the environment.

  18. Role of nitric oxide and superoxide in Giardia lamblia killing.

    PubMed

    Fernandes, P D; Assreuy, J

    1997-01-01

    Giardia lamblia trophozoites were incubated for 2 h with activated murine macrophages, nitric oxide (NO) donors or a superoxide anion generator (20 mU/ml xanthine oxidase plus 1 mM xanthine). Activated macrophages were cytotoxic to Giardia trophozoites (approximately 60% dead trophozoites). The effect was inhibited (> 90%) by an NO synthase inhibitor (200 microM) and unaffected by superoxide dismutase (SOD, 300 U/ml). Giardia trophozoites were killed by the NO donors, S-nitroso-acetyl-penicillamine (SNAP) and sodium nitroprusside (SNP) in a dose-dependent manner (LD50 300 and 50 microM, respectively). A dual NO-superoxide anion donor, 3-morpholino-sydnonimine hydrochloride (SIN-1), did not have a killing effect in concentrations up to 1 mM. However, when SOD (300 U/ml) was added simultaneously with SIN-1 to Giardia, a significant trophozoite-killing effect was observed (approximately 35% dead trophozoites at 1 mM). The mixtures of SNAP or SNP with superoxide anion, which yields peroxynitrite, abolished the trophozoite killing induced by NO donors. Authentic peroxynitrite only killed trophozoites at very high concentrations (3 mM). These results indicate that NO accounts for Giardia trophozoites killing and this effect is not mediated by peroxynitrite.

  19. Cu/Zn superoxide dismutases in developing cotton fibers

    USDA-ARS?s Scientific Manuscript database

    Hydrogen peroxide (H2O2) and other reactive oxygen species (ROS) are important signaling molecules in diverse physiological processes. Previously, we discovered superoxide dismutase (SOD) activity in extracellular protein preparations from fiber-bearing cotton (Gossypium hirsutum L.) seeds. We sho...

  20. Therapeutic effect of lecithinized superoxide dismutase on pulmonary emphysema.

    PubMed

    Tanaka, Ken-Ichiro; Tanaka, Yuta; Miyazaki, Yuri; Namba, Takushi; Sato, Keizo; Aoshiba, Kazutetsu; Azuma, Arata; Mizushima, Tohru

    2011-09-01

    No medication exists that clearly improves the mortality of chronic obstructive pulmonary disease (COPD). Oxidative molecules, in particular superoxide anions, play important roles in the COPD-associated abnormal inflammatory response and pulmonary emphysema, which arises because of an imbalance in proteases and antiproteases and increased apoptosis. Superoxide dismutase (SOD) catalyzes the dismutation of superoxide anions. Lecithinized human Cu/Zn- SOD (PC-SOD) has overcome a number of the clinical limitations of SOD, including low tissue affinity and low stability in plasma. In this study, we examine the effect of PC-SOD on elastase-induced pulmonary emphysema, an animal model of COPD. The severity of the pulmonary inflammatory response and emphysema in mice was assessed by various criteria, such as the number of leukocytes in the bronchoalveolar lavage fluid and the enlargement of airspace. Not only intravenous administration but also inhalation of PC-SOD suppressed elastase-induced pulmonary inflammation, emphysema, and dysfunction. Inhalation of PC-SOD suppressed the elastase-induced increase in the pulmonary level of superoxide anions and apoptosis. Inhalation of PC-SOD also suppressed elastase-induced activation of proteases and decreased in the level of antiproteases and expression of proinflammatory cytokines and chemokines. We also found that inhalation of PC-SOD suppressed cigarette smoke-induced pulmonary inflammation. The results suggest that PC-SOD protects against pulmonary emphysema by decreasing the pulmonary level of superoxide anions, resulting in the inhibition of inflammation and apoptosis and amelioration of the protease/antiprotease imbalance. We propose that inhalation of PC-SOD would be therapeutically beneficial for COPD.

  1. Superoxide dismutase-inhibitible reduction of cytochrome c by the alloxan radical. Implications for alloxan cytotoxicity.

    PubMed Central

    Winterbourn, C C

    1982-01-01

    Cytochrome c was reduced when superoxide was generated from xanthine oxidase in the presence of alloxan, and by the reaction of alloxan and with reduced glutathione. In each case, most of the reduction was inhibited by superoxide dismutase, but considerably more enzyme was required than with superoxide alone. This indicates that the superoxide dismutase-inhibitible cytochrome c reduction was mainly due to a direct reaction with the alloxan radical, and implies that other reactions that are inhibited by superoxide dismutase could be due to either alloxan radicals or superoxide. PMID:6299273

  2. Extracellular superoxide anion production contributes to the virulence of Xanthomonas oryzae pv. oryzae.

    PubMed

    Li, Xin; Pang, Xinyue; Zhi, Dejuan; Wang, Jinsheng; Li, Minquan; Li, Hongyu

    2009-02-01

    Endogenous superoxide anion production was determined by electron spin resonance in wild-type strains and avrXa7 mutants of Xanthomonas oryzae pv. oryzae. The localization of superoxide anion was carried out in the intra- and extra-cellular fractions. Results showed the presence of superoxide anion in multi-locations of X. oryzae pv. oryzae cells. The extracellular fraction was the major location of superoxide anion production. Furthermore, a positive relationship was shown between the levels of endogenous superoxide anion and the virulence of strains. These indubitable results suggested that the superoxide anion contributes to the virulence of X. oryzae pv. oryzae.

  3. Novel mechanisms for superoxide-scavenging activity of human manganese superoxide dismutase determined by the K68 key acetylation site.

    PubMed

    Lu, Jiaqi; Cheng, Kuoyuan; Zhang, Bo; Xu, Huan; Cao, Yuanzhao; Guo, Fei; Feng, Xudong; Xia, Qing

    2015-08-01

    Superoxide is the primary reactive oxygen species generated in the mitochondria. Manganese superoxide dismutase (SOD2) is the major enzymatic superoxide scavenger present in the mitochondrial matrix and one of the most crucial reactive oxygen species-scavenging enzymes in the cell. SOD2 is activated by sirtuin 3 (SIRT3) through NAD(+)-dependent deacetylation. However, the exact acetylation sites of SOD2 are ambiguous and the mechanisms underlying the deacetylation-mediated SOD2 activation largely remain unknown. We are the first to characterize SOD2 mutants of the acetylation sites by investigating the relative enzymatic activity, structures, and electrostatic potential of SOD2 in this study. These SOD2 mutations affected the superoxide-scavenging activity in vitro and in HEK293T cells. The lysine 68 (K68) site is the most important acetylation site contributing to SOD2 activation and plays a role in cell survival after paraquat treatment. The molecular basis underlying the regulation of SOD2 activity by K68 was investigated in detail. Molecular dynamics simulations revealed that K68 mutations induced a conformational shift of residues located in the active center of SOD2 and altered the charge distribution on the SOD2 surface. Thus, the entry of the superoxide anion into the coordinated core of SOD2 was inhibited. Our results provide a novel mechanistic insight, whereby SOD2 acetylation affects the structure and charge distribution of SOD2, its tetramerization, and p53-SOD2 interactions of SOD2 in the mitochondria, which may play a role in nuclear-mitochondrial communication during aging.

  4. Real time monitoring of superoxide dynamics in vivo through fluorescent proteins using a sensitive fiber probe

    NASA Astrophysics Data System (ADS)

    Chang, Yu-Chung; Ken, Chuian-Fu; Hsu, Che-Wei; Liu, Ya-Ging

    2014-03-01

    Superoxide anion is the primary oxygen free radical generated in mitochondria that causes intracellular oxidative stress. The lack of a method to directly monitor superoxide concentration in vivo in real time has severely hindered our understanding on its pathophysiology. We made transgenic zebrafish to specifically express fluorescent proteins, which are recently developed as reversible superoxide-specific indicators, in the liver. A fiber-optic fluorescent probe was used to noninvasively monitor superoxide generation in the liver in real time. The fish were placed in microfluidic channels for manipulation and reagents administration. Several superoxide-inducing and scavenging reagents were administrated onto the fish to investigate their effects on superoxide anion balancing. The biochemical dynamics of superoxide due to the application reagents were revealed in the transient behaviors of fluorescence time courses. With the ability to monitor superoxide dynamics in vivo in real time, this method can be used as an in vivo pharmaceutical screening platform.

  5. Cupric yersiniabactin is a virulence-associated superoxide dismutase mimic.

    PubMed

    Chaturvedi, Kaveri S; Hung, Chia S; Giblin, Daryl E; Urushidani, Saki; Austin, Anthony M; Dinauer, Mary C; Henderson, Jeffrey P

    2014-02-21

    Many Gram-negative bacteria interact with extracellular metal ions by expressing one or more siderophore types. Among these, the virulence-associated siderophore yersiniabactin (Ybt) is an avid copper chelator, forming stable cupric (Cu(II)-Ybt) complexes that are detectable in infected patients. Here we show that Ybt-expressing E. coli are protected from intracellular killing within copper-replete phagocytic cells. This survival advantage is highly dependent upon the phagocyte respiratory burst, during which superoxide is generated by the NADPH oxidase complex. Chemical fractionation links this phenotype to a previously unappreciated superoxide dismutase (SOD)-like activity of Cu(II)-Ybt. Unlike previously described synthetic copper-salicylate (Cu(II)-SA) SOD mimics, the salicylate-based natural product Cu(II)-Ybt retains catalytic activity at physiologically plausible protein concentrations. These results reveal a new virulence-associated adaptation based upon spontaneous assembly of a non-protein catalyst.

  6. Extracellular Superoxide Dismutase: Growth Promoter or Tumor Suppressor?

    PubMed Central

    Laukkanen, Mikko O.

    2016-01-01

    Extracellular superoxide dismutase (SOD3) gene transfer to tissue damage results in increased healing, increased cell proliferation, decreased apoptosis, and decreased inflammatory cell infiltration. At molecular level, in vivo SOD3 overexpression reduces superoxide anion (O2−) concentration and increases mitogen kinase activation suggesting that SOD3 could have life-supporting characteristics. The hypothesis is further strengthened by the observations showing significantly increased mortality in conditional knockout mice. However, in cancer SOD3 has been shown to either increase or decrease cell proliferation and survival depending on the model system used, indicating that SOD3-derived growth mechanisms are not completely understood. In this paper, the author reviews the main discoveries in SOD3-dependent growth regulation and signal transduction. PMID:27293512

  7. Cupric Yersiniabactin Is a Virulence-Associated Superoxide Dismutase Mimic

    PubMed Central

    2013-01-01

    Many Gram-negative bacteria interact with extracellular metal ions by expressing one or more siderophore types. Among these, the virulence-associated siderophore yersiniabactin (Ybt) is an avid copper chelator, forming stable cupric (Cu(II)-Ybt) complexes that are detectable in infected patients. Here we show that Ybt-expressing E. coli are protected from intracellular killing within copper-replete phagocytic cells. This survival advantage is highly dependent upon the phagocyte respiratory burst, during which superoxide is generated by the NADPH oxidase complex. Chemical fractionation links this phenotype to a previously unappreciated superoxide dismutase (SOD)-like activity of Cu(II)-Ybt. Unlike previously described synthetic copper-salicylate (Cu(II)-SA) SOD mimics, the salicylate-based natural product Cu(II)-Ybt retains catalytic activity at physiologically plausible protein concentrations. These results reveal a new virulence-associated adaptation based upon spontaneous assembly of a non-protein catalyst. PMID:24283977

  8. Preliminary crystallographic analysis of the Megavirus superoxide dismutase

    PubMed Central

    Lartigue, Audrey; Philippe, Nadège; Jeudy, Sandra; Abergel, Chantal

    2012-01-01

    Megavirus chilensis, a close relative of the Mimivirus giant virus, is able to replicate in Acanthamoeba castellanii. The first step of viral infection involves the internalization of the virions in host vacuoles. It has been experimentally demonstrated that Mimivirus particles contain many proteins capable of resisting oxidative stress, as encountered in the phagocytic process. These proteins are conserved in Megavirus, which has an additional gene (Mg277) encoding a putative superoxide dismutase. The Mg277 ORF product was overexpressed in Escherichia coli, purified and crystallized. A SAD data set was collected to 2.24 Å resolution at the selenium peak wavelength on the BM30 beamline at the ESRF from a single crystal of selenomethionine-substituted recombinant superoxide dismutase protein. PMID:23192047

  9. [Generation of superoxides during the interaction of melanins with oxygen].

    PubMed

    Lapina, V A; Dontsov, A E; Ostrovskiĭ, M A

    1984-10-01

    The rate of nitroblue tetrazolium (NBT) reduction by dihydroxyphenylalanine-melanin, pheomelanin and retinal pigment epithelium melanosomes under aerobic conditions (pH 7.4) is low both in the dark and upon illumination, but increases drastically in the presence of cetyltrimethylammonium bromide (CTAB). Under these conditions, the light insignificantly stimulates NBT reduction (1.3-fold). The reaction is effectively inhibited by superoxide dismutase. This suggests that superoxide anions (O2-. are formed as intermediate reaction products in the course of NBT reduction by melanins. At alkaline values of pH (greater than or equal to 9.0), the O2-.-dependent reduction of NBT can also take place in the absence of CTAB. In contrast with oxidation of photoreduced riboflavin, the melanin oxidation by O2 cannot induce lipid peroxidation. It is concluded that O2-. generation via melanin oxidation of melanosomes occurs only under non-physiological conditions and can hardly take place in vivo.

  10. Superoxide dismutase protects Escherichia coli against killing by human serum.

    PubMed

    McManus, D C; Josephy, P D

    1995-02-20

    To assess the role of superoxide dismutase in protecting Escherichia coli from killing by human serum and neutrophils, we constructed isogenic, smooth-lipopolysaccharide K-12 strains, either sod wild-type, delta sodA, or delta sodA delta sodB. The delta sodA delta sodB strain was killed by serum much more readily than either the wild-type or delta sodA strain. After allowing for this serum sensitivity difference, the delta sodA delta sodB strain also showed increased susceptibility to phagocytic killing by human neutrophils. These results indicate that superoxide dismutase protects E. coli from killing by serum (complement system) and by human neutrophils, possibly by a role in maintaining bacterial membrane structure.

  11. The Superoxide Reductase from the Early Diverging Eukaryote Giardia Intestinalis

    SciTech Connect

    Cabelli, D.E.; Testa, F.; Mastronicola, D.; Bordi, E.; Pucillo, L.P.; Sarti, P.; Saraiva, L.M.; Giuffre, A.; Teixeira, M.

    2011-10-15

    Unlike superoxide dismutases (SODs), superoxidereductases (SORs) eliminate superoxide anion (O{sub 2}{sup {sm_bullet}-}) not through its dismutation, but via reduction to hydrogen peroxide (H{sub 2}O{sub 2}) in the presence of an electron donor. The microaerobic protist Giardia intestinalis, responsible for a common intestinal disease in humans, though lacking SOD and other canonical reactive oxygen species-detoxifying systems, is among the very few eukaryotes encoding a SOR yet identified. In this study, the recombinant SOR from Giardia (SOR{sub Gi}) was purified and characterized by pulse radiolysis and stopped-flow spectrophotometry. The protein, isolated in the reduced state, after oxidation by superoxide or hexachloroiridate(IV), yields a resting species (T{sub final}) with Fe{sup 3+} ligated to glutamate or hydroxide depending on pH (apparent pK{sub a} = 8.7). Although showing negligible SOD activity, reduced SOR{sub Gi} reacts with O{sub 2}{sup {sm_bullet}-} with a pH-independent second-order rate constant k{sub 1} = 1.0 x 10{sup 9} M{sup -1} s{sup -1} and yields the ferric-(hydro)peroxo intermediate T{sub 1}; this in turn rapidly decays to the T{sub final} state with pH-dependent rates, without populating other detectable intermediates. Immunoblotting assays show that SOR{sub Gi} is expressed in the disease-causing trophozoite of Giardia. We propose that the superoxide-scavenging activity of SOR in Giardia may promote the survival of this air-sensitive parasite in the fairly aerobic proximal human small intestine during infection.

  12. Calcineurin transgenic mice have mitochondrial dysfunction and elevated superoxide production.

    PubMed

    Sayen, M R; Gustafsson, Asa B; Sussman, Mark A; Molkentin, Jeffery D; Gottlieb, Roberta A

    2003-02-01

    Introduction of the constitutively active calcineurin gene into neonatal rat cardiomyocytes by adenovirus resulted in decreased mitochondrial membrane potential (P < 0.05). Infection of H9c2 cells with calcineurin adenovirus resulted in increased superoxide production (P < 0.001). Transgenic mice with cardiac-specific expression of a constitutively active calcineurin cDNA (CalTG mice) exhibit a two- to threefold increase in heart size that progresses to heart failure. We prepared mitochondria enriched for the subsarcolemmal population from the hearts of CalTG mice and transgene negative littermates (control). Intact, well-coupled mitochondria prepared from one to two mouse hearts at a time yielded sufficient material for functional studies. Mitochondrial oxygen consumption was measured with a Clark-type oxygen electrode with substrates for mitochondrial complex II (succinate) and complex IV [tetramethylpentadecane (TMPD)/ascorbate]. CalTG mice exhibited a maximal rate of electron transfer in heart mitochondria that was reduced by approximately 50% (P < 0.002) without a loss of respiratory control. Mitochondrial respiration was unaffected in tropomodulin-overexpressing transgenic mice, another model of cardiomyopathy. Western blotting for mitochondrial electron transfer subunits from mitochondria of CalTG mice revealed a 20-30% reduction in subunit 3 of complex I (ND3) and subunits I and IV of cytochrome oxidase (CO-I, CO-IV) when normalized to total mitochondrial protein or to the adenine nucleotide transporter (ANT) and compared with littermate controls (P < 0.002). Impaired mitochondrial electron transport was associated with high levels of superoxide production in the CalTG mice. Taken together, these data indicate that calcineurin signaling affects mitochondrial energetics and superoxide production. The excessive production of superoxide may contribute to the development of cardiac failure.

  13. Copper complexes of bioactive ligands with superoxide dismutase activity.

    PubMed

    Khalid, Huma; Hanif, Muhammad; Hashmi, Muhammad Ali; Mahmood, Tariq; Ayub, Khurshid; Monim-Ul-Mehboob, Muhammad

    2013-11-01

    Free radicals or reactive oxygen species (ROS) are highly toxic and their damaging effects result in a variety of detrimental health issues such as neurodegenerative, cardiovascular and age-related diseases. Human body has evolved an effective defense system including superoxide dismutase (SOD) and catalase against the toxicity of these free radicals. SOD is a metalloenzyme and it acts as an excellent antioxidant to protect the body from superoxide radicals that are generated in the biological system. However, the clinical use of SOD is limited due to its short in vivo life span, and its large size that hampered its penetration across the cell membranes. Pharmaceuticals that provide ROS scavenging systems are the most effective when the production of ROS exceeds the scavenging capacity of endogenous SOD as a result of aging or pathological processes. Inspired by the Nature, scientists have designed metal-based mimics of the superoxide dismutase. This review focuses on different copper complexes that are developed from bioactive ligands and mimic the protecting action of the SOD.

  14. Superoxide generation and cytotactic response of irradiated neutrophils

    SciTech Connect

    Eastlund, D.T.; Charbonneau, T.T.

    1988-07-01

    Irradiation of blood components has been used to prevent transfusion-related graft-versus-host disease (GVHD) in immunocompromised patients. This study was designed to determine the effect of irradiation on neutrophil aggregation, chemotaxis, and superoxide generation. Purified neutrophils were irradiated with a Cesium source at four doses ranging from 0 to 17,500 rads. Formyl-methionyl-leucyl-phenylalanine (FMLP) and zymosan-treated serum (ZTS) cytotaxin-induced chemotaxis and migration were determined in the agarose assay. Neutrophil aggregation to FMLP was determined by aggregometry. Superoxide generation and random migration were not affected by irradiation at doses up to 17,500 rads. When compared to nonirradiated controls, the chemotactic response to ZTS remained normal, with an insignificant decline from 174 +/- 31.0 to 150 +/- 42.3 (mean +/- SD) units. The chemotactic response to FMLP declined insignificantly, from 228 +/- 31.3 at 0 rad to 207 +/- 26.4 at 17,500 rads. The aggregation response to FMLP remained within the normal range but declined from 0.78 +/- 0.11 to 0.61 +/- 0.18. At the radiation doses currently used to reduce the risk of transfusion-related GVHD, neutrophil superoxide generation and chemotactic response remain essentially normal.

  15. A new method to prevent degradation of lithium-oxygen batteries: reduction of superoxide by viologen.

    PubMed

    Yang, L; Frith, J T; Garcia-Araez, N; Owen, J R

    2015-01-31

    Lithium-oxygen battery development is hampered by degradation reactions initiated by superoxide, which is formed in the pathway of oxygen reduction to peroxide. This work demonstrates that the superoxide lifetime is drastically decreased upon addition of ethyl viologen, which catalyses the reduction of superoxide to peroxide.

  16. Reactions of superoxide dismutases with HS(-)/H2S and superoxide radical anion: An in vitro EPR study.

    PubMed

    Bolić, Bojana; Mijušković, Ana; Popović-Bijelić, Ana; Nikolić-Kokić, Aleksandra; Spasić, Snežana; Blagojević, Duško; Spasić, Mihajlo B; Spasojević, Ivan

    2015-12-01

    Interactions of hydrogen sulfide (HS(-)/H2S), a reducing signaling species, with superoxide dimutases (SOD) are poorly understood. We applied low-T EPR spectroscopy to examine the effects of HS(-)/H2S and superoxide radical anion O2.- on metallocenters of FeSOD, MnSOD, and CuZnSOD. HS(-)/H2S did not affect FeSOD, whereas active centers of MnSOD and CuZnSOD were open to this agent. Cu(2+) was reduced to Cu(1+), while manganese appears to be released from MnSOD active center. Untreated and O2.- treated FeSOD and MnSOD predominantly show 5 d-electron systems, i.e. Fe(3+) and Mn(2+). Our study provides new details on the mechanisms of (patho)physiological effects of HS(-)/H2S.

  17. A mitochondrial superoxide signal triggers increased longevity in Caenorhabditis elegans.

    PubMed

    Yang, Wen; Hekimi, Siegfried

    2010-12-07

    The nuo-6 and isp-1 genes of C. elegans encode, respectively, subunits of complex I and III of the mitochondrial respiratory chain. Partial loss-of-function mutations in these genes decrease electron transport and greatly increase the longevity of C. elegans by a mechanism that is distinct from that induced by reducing their level of expression by RNAi. Electron transport is a major source of the superoxide anion (O(⋅) (-)), which in turn generates several types of toxic reactive oxygen species (ROS), and aging is accompanied by increased oxidative stress, which is an imbalance between the generation and detoxification of ROS. These observations have suggested that the longevity of such mitochondrial mutants might result from a reduction in ROS generation, which would be consistent with the mitochondrial oxidative stress theory of aging. It is difficult to measure ROS directly in living animals, and this has held back progress in determining their function in aging. Here we have adapted a technique of flow cytometry to directly measure ROS levels in isolated mitochondria to show that the generation of superoxide is elevated in the nuo-6 and isp-1 mitochondrial mutants, although overall ROS levels are not, and oxidative stress is low. Furthermore, we show that this elevation is necessary and sufficient to increase longevity, as it is abolished by the antioxidants NAC and vitamin C, and phenocopied by mild treatment with the prooxidant paraquat. Furthermore, the absence of effect of NAC and the additivity of the effect of paraquat on a variety of long- and short-lived mutants suggest that the pathway triggered by mitochondrial superoxide is distinct from previously studied mechanisms, including insulin signaling, dietary restriction, ubiquinone deficiency, the hypoxic response, and hormesis. These findings are not consistent with the mitochondrial oxidative stress theory of aging. Instead they show that increased superoxide generation acts as a signal in young

  18. Detoxification of superoxide without production of H2O2: antioxidant activity of superoxide reductase complexed with ferrocyanide.

    PubMed

    Molina-Heredia, Fernando P; Houée-Levin, Chantal; Berthomieu, Catherine; Touati, Danièle; Tremey, Emilie; Favaudon, Vincent; Adam, Virgile; Nivière, Vincent

    2006-10-03

    The superoxide radical O(2)(-.) is a toxic by-product of oxygen metabolism. Two O(2)(-.) detoxifying enzymes have been described so far, superoxide dismutase and superoxide reductase (SOR), both forming H2O2 as a reaction product. Recently, the SOR active site, a ferrous iron in a [Fe(2+) (N-His)(4) (S-Cys)] pentacoordination, was shown to have the ability to form a complex with the organometallic compound ferrocyanide. Here, we have investigated in detail the reactivity of the SOR-ferrocyanide complex with O(2)(-.) by pulse and gamma-ray radiolysis, infrared, and UV-visible spectroscopies. The complex reacts very efficiently with O(2)(-.). However, the presence of the ferrocyanide adduct markedly modifies the reaction mechanism of SOR, with the formation of transient intermediates different from those observed for SOR alone. A one-electron redox chemistry appears to be carried out by the ferrocyanide moiety of the complex, whereas the SOR iron site remains in the reduced state. Surprisingly, the toxic H2O2 species is no longer the reaction product. Accordingly, in vivo experiments showed that formation of the SOR-ferrocyanide complex increased the antioxidant capabilities of SOR expressed in an Escherichia coli sodA sodB recA mutant strain. Altogether, these data describe an unprecedented O(2)(-.) detoxification activity, catalyzed by the SOR-ferrocyanide complex, which does not conduct to the production of the toxic H2O2 species.

  19. Superoxide Generation and Its Involvement in the Growth of Mycobacterium smegmatis

    PubMed Central

    Yeware, Amar M.; Shurpali, Ketaki D.; Athalye, Meghana C.; Sarkar, Dhiman

    2017-01-01

    Superoxide generation is inevitable in aerobic organisms, most of which have developed mechanisms to detoxify superoxides. However, its significance has not been clearly understood in mycobacteria. This study demonstrates that NADH oxidase is the major source of superoxide in Mycobacterium smegmatis and elucidates the involvement of superoxide in M. smegmatis growth. The maximum inhibition of superoxide generation was observed in the presence of diphenyleneiodonium chloride (DPI), an NADH oxidase inhibitor, compared to other standard inhibitors. After incubation for 24 h, the number of colony forming units (CFUs) was reduced by 6.8 log10 compared to the untreated culture. The inhibitory effect of DPI on M. smegmatis was reversed when the same culture was supplemented with menadione and pyrogallol, which are superoxide generators. Thus, this study reports the source of superoxide generation and its involvement in the growth of M. smegmatis. PMID:28194149

  20. Cu,Zn Superoxide Dismutase is a Peroxisomal Enzyme in Human Fibroblast and Hepatoma Cells

    NASA Astrophysics Data System (ADS)

    Keller, Gilbert-Andre; Warner, Thomas G.; Steimer, Kathelyn S.; Hallewell, Robert A.

    1991-08-01

    The intracellular localization of Cu,Zn superoxide dismutase (superoxide:superoxide oxidoreductase, EC 1.15.1.1) has been examined by immunofluorescence using four monoclonal anti-Cu,Zn superoxide dismutase antibodies raised against a recombinant human Cu,Zn superoxide dismutase derivative produced and purified from Escherichia coli. Colocalization with catalase, a peroxisomal matrix enzyme, was used to demonstrate the peroxisomal localization of Cu,Zn superoxide dismutase in human fibroblasts and hepatoma cells. In the fibroblasts of Zellweger syndrome patients, the enzyme is not transported to the peroxisomal ghosts but, like catalase, remains in the cytoplasm. In addition, immunocryoelectron microscopy of yeast cells expressing human Cu,Zn superoxide dismutase showed that the enzyme is translocated to the peroxisomes.

  1. Healing of colonic ischemic anastomoses in the rat: role of superoxide radicals.

    PubMed

    Garcia, J G; Criado, F J; Persona, M A; Alonso, A G

    1998-07-01

    The aim of this study was to evaluate the role of superoxide radicals in the healing of ischemic colonic anastomoses in the rat. Adult male Wistar rats were used in a factorial design with two factors (normal or ischemic colonic anastomoses) each having two levels (treatment with saline or allopurinol). Colonic anastomoses were performed either in normal or previously devascularized colons (ischemic anastomoses) at identical locations, using the same technique. On the fourth postoperative day, animals were killed, and specimens were taken for determinations. Ischemic anastomoses displayed significant increases in superoxide radical (assayed as superoxide anion), superoxide dismutase, and glutathione peroxidase concentrations. Bursting strength and hydroxyproline levels were also significantly lower in these anastomoses. Allopurinol administration elicited a significant decrease in superoxide anions and raised both bursting strength and hydroxyproline levels only in ischemic anastomoses. Superoxide radicals are involved in the delay in healing of ischemic anastomoses. Allopurinol lowers superoxide anion production and has beneficial effects on the cicatrization of ischemic anastomoses.

  2. Mechanical overloading causes mitochondrial superoxide and SOD2 imbalance in chondrocytes resulting in cartilage degeneration.

    PubMed

    Koike, Masato; Nojiri, Hidetoshi; Ozawa, Yusuke; Watanabe, Kenji; Muramatsu, Yuta; Kaneko, Haruka; Morikawa, Daichi; Kobayashi, Keiji; Saita, Yoshitomo; Sasho, Takahisa; Shirasawa, Takuji; Yokote, Koutaro; Kaneko, Kazuo; Shimizu, Takahiko

    2015-06-25

    Mechanical stress and aging are major risk factors of cartilage degeneration. Human studies have previously reported that oxidative damage increased, while SOD2 protein was reciprocally downregulated in osteoarthritic degenerated cartilage. However, it remains unclear whether mitochondrial superoxide imbalance in chondrocytes causes cartilage degeneration. We herein demonstrate that mechanical loading promoted mitochondrial superoxide generation and selective Sod2 downregulation in chondrocytes in vivo and that mitochondrial superoxide inducer also downregulated Sod2 expression in chondrocytes in vitro. A genetically manipulated model revealed that Sod2 deficiency in chondrocytes also resulted in mitochondrial superoxide overproduction and dysfunction, thus leading to cartilage degeneration. Intra-articular injection of a permeable antioxidant effectively suppressed the mechanical loading-induced mitochondrial superoxide generation and cartilage degeneration in mice. Our findings demonstrate that mitochondrial superoxide plays a pivotal role in the development and progression of osteoarthritis, and the mitochondrial superoxide balance may therefore be a promising target for the treatment of cartilage degeneration.

  3. A lithium–oxygen battery based on lithium superoxide

    DOE PAGES

    Lu, Jun; Jung Lee, Yun; Luo, Xiangyi; ...

    2016-01-11

    Batteries based on sodium superoxide and on potassium superoxide have recently been reported(1-3). But, there have been no reports of a battery based on lithium superoxide (LiO2), despite much research(4-8) into the lithium-oxygen (Li-O2) battery because of its potential high energy density. Several studies(9-16) of Li-O2 batteries have found evidence of LiO2 being formed as one component of the discharge product along with lithium peroxide (Li2O2). In addition, theoretical calculations have indicated that some forms of LiO2 may have a long lifetime(17). Our studies also suggest that it might be possible to form LiO2 alone for use in a battery.more » However, solid LiO2 has been difficult to synthesize in pure form(18) because it is thermodynamically unstable with respect to disproportionation, giving Li2O2 (refs 19, 20). We show that crystalline LiO2 can be stabilized in a Li-O2 battery by using a suitable graphene-based cathode. Various characterization techniques reveal no evidence for the presence of Li2O2. A novel templating growth mechanism involving the use of iridium nanoparticles on the cathode surface may be responsible for the growth of crystalline LiO2. Our results demonstrate that the LiO2 formed in the Li-O2 battery is stable enough for the battery to be repeatedly charged and discharged with a very low charge potential (about 3.2 volts). We also anticipate that this discovery will lead to methods of synthesizing and stabilizing LiO2, which could open the way to high-energy-density batteries based on LiO2 as well as to other possible uses of this compound, such as oxygen storage.« less

  4. Balance between Endogenous Superoxide Stress and Antioxidant Defenses

    PubMed Central

    Gort, Amy Strohmeier; Imlay, James A.

    1998-01-01

    Cells devoid of cytosolic superoxide dismutase (SOD) suffer enzyme inactivation, growth deficiencies, and DNA damage. It has been proposed that the scant superoxide (O2−) generated by aerobic metabolism harms even cells that contain abundant SOD. However, this idea has been difficult to test. To determine the amount of O2− that is needed to cause these defects, we modulated the O2− concentration inside Escherichia coli by controlling the expression of SOD. An increase in O2− of more than twofold above wild-type levels substantially diminished the activity of labile dehydratases, an increase in O2− of any more than fourfold measurably impaired growth, and a fivefold increase in O2− sensitized cells to DNA damage. These results indicate that E. coli constitutively synthesizes just enough SOD to defend biomolecules against endogenous O2− so that modest increases in O2− concentration diminish cell fitness. This conclusion is in excellent agreement with quantitative predictions based upon previously determined rates of intracellular O2− production, O2− dismutation, dehydratase inactivation, and enzyme repair. The vulnerability of bacteria to increased intracellular O2− explains the widespread use of superoxide-producing drugs as bactericidal weapons in nature. E. coli responds to such drugs by inducing the SoxRS regulon, which positively regulates synthesis of SOD and other defensive proteins. However, even toxic amounts of endogenous O2− did not activate SoxR, and SoxR activation by paraquat was not at all inhibited by excess SOD. Therefore, in responding to redox-cycling drugs, SoxR senses some signal other than O2−. PMID:9515906

  5. A superoxide dismutase of metacestodes of Taenia taeniaeformis.

    PubMed

    Leid, R W; Suquet, C M

    1986-03-01

    Superoxide dismutase was purified from Taenia taeniaeformis metacestodes by sequential ion exchange chromatography on quaternary-amino-ethyl-cellulose, gel filtration chromatography on ACA 44 and ion exchange chromatography on DEAE-cellulose, followed by chromatofocusing on polybuffer exchanger 94. This isolation procedure resulted in the detection of a single protein-staining band on alkaline gels, coincident with enzyme activity. We have, however, detected what appear to be two peaks of enzyme activity within this single protein-staining band. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis using gradient slab gels and analysis under reducing conditions, resulted in the detection of only one protein at an apparent Mr of 16,600, while analysis under non-reducing conditions, gave a single protein of an apparent Mr of 64,000. The isoelectric point of the purified protein is 6.6. Boiling for 3 min completely destroyed the enzyme, whereas incubation for 2 h at 37 degrees C resulted in the loss of 56% of the enzymic activity. Incubation with 10 mM KCN resulted in 83% inhibition of the enzyme. We have detected up to 168 U ml-1 of enzyme activity in the cyst fluid surrounding the parasite in situ. This is the first instance in which any parasite superoxide dismutase has been purified to apparent homogeneity. Parasite-mediated enzymic destruction of superoxide anion can not only protect against oxygen toxicity as a result of normal parasite respiratory processes but also may serve as yet another mechanism used by tissue-dwelling parasites to evade host immunologic attack.

  6. Purification and characterization of superoxide dismutase from chicken liver.

    PubMed

    Oztürk-Urek, R; Tarhan, L

    2001-02-01

    Superoxide dismutase (SOD; EC 1.15.1.1) is an enzyme that protects against oxidative stress from superoxide radicals in living cells. This enzyme has been isolated, purified and partially characterized from chicken liver. The following steps were carried out in order to purify chicken liver SOD. Initially, the liver was homogenized and hemoglobin was removed. Subsequently protein precipitation was effected with (NH(4))(2)SO(4), methanol, (NH(4))(2)SO(4)-methanol and polyethylene glycol methods. The product from polyethylene glycol-3350 precipitation was found to have the highest SOD activity. Polyethylene glycol was removed by chromatography using a PD-10 column. After passing through an ultrafilter, the superoxide dismutase was fractionated by DEAE-ion chromatography and then Sephadex G-75 gel filtration chromatography. During this purification procedure, a specific activity of 4818.2 IU/mg was reached, corresponding to 285.8-fold purification. The purified enzyme, which was characterized as cyanide-sensitive SOD, contained two subunits having Cu and Zn elements with a molecular weight of 16000+/-500 for each. The optimum pH of purified CuZnSOD was determined to be 8.9. The enzyme was found to have good pH stability in the pH range 6.0-7.5 at 25 degrees C over a 2-h incubation period and displayed good thermal stability up to 45 degrees C at pH 7.4 over a 1-h incubation period. The SOD enzyme was not inhibited by DTT and beta-mercaptoethanol, but inhibited by CN(-) and H(2)O(2). In the presence of 2 mM iodoacetamide, the enzyme showed an approximately 40% activity loss. Finally, the inhibitory effect of ionic strength on SOD was also investigated.

  7. Autoxidation of ubiquinol-6 is independent of superoxide dismutase.

    PubMed

    Schultz, J R; Ellerby, L M; Gralla, E B; Valentine, J S; Clarke, C F

    1996-05-28

    Ubiquinone (Q) is an essential, lipid soluble, redox component of the mitochondrial respiratory chain. Much evidence suggests that ubiquinol (QH2) functions as an effective antioxidant in a number of membrane and biological systems by preventing peroxidative damage to lipids. It has been proposed that superoxide dismutase (SOD) may protect QH2 form autoxidation by acting either directly as a superoxide-semiquinone oxidoreductase or indirectly by scavenging superoxide. In this study, such an interaction between QH2 and SOD was tested by monitoring the fluorescence of cis-parinaric acid (cPN) incorporated phosphatidylcholine (PC) liposomes. Q6H2 was found to prevent both fluorescence decay and generation of lipid peroxides (LOOH) when peroxidation was initiated by the lipid-soluble azo initiator DAMP, dimethyl 2,2'-azobis (2-methylpropionate), while Q6 or SOD alone had no inhibitory effect. Addition of either SOD or catalase to Q6H2-containing liposomes had little effect on the rate of peroxidation even when incubated in 100% O2. Hence, the autoxidation of QH2 is a competing reaction that reduces the effectiveness of QH2 as an antioxidant and was not slowed by either SOD or catalase. The in vivo interaction of SOD and QH2 was also tested by employing yeast mutant strains harboring deletions in either CuZnSOD and/or MnSOD. The sod mutant yeast strains contained the same percent Q6H2 per cell as wild-type cells. These results indicate that the autoxidation of QH2 is independent of SOD.

  8. A lithium-oxygen battery based on lithium superoxide.

    PubMed

    Lu, Jun; Lee, Yun Jung; Luo, Xiangyi; Lau, Kah Chun; Asadi, Mohammad; Wang, Hsien-Hau; Brombosz, Scott; Wen, Jianguo; Zhai, Dengyun; Chen, Zonghai; Miller, Dean J; Jeong, Yo Sub; Park, Jin-Bum; Fang, Zhigang Zak; Kumar, Bijandra; Salehi-Khojin, Amin; Sun, Yang-Kook; Curtiss, Larry A; Amine, Khalil

    2016-01-21

    Batteries based on sodium superoxide and on potassium superoxide have recently been reported. However, there have been no reports of a battery based on lithium superoxide (LiO2), despite much research into the lithium-oxygen (Li-O2) battery because of its potential high energy density. Several studies of Li-O2 batteries have found evidence of LiO2 being formed as one component of the discharge product along with lithium peroxide (Li2O2). In addition, theoretical calculations have indicated that some forms of LiO2 may have a long lifetime. These studies also suggest that it might be possible to form LiO2 alone for use in a battery. However, solid LiO2 has been difficult to synthesize in pure form because it is thermodynamically unstable with respect to disproportionation, giving Li2O2 (refs 19, 20). Here we show that crystalline LiO2 can be stabilized in a Li-O2 battery by using a suitable graphene-based cathode. Various characterization techniques reveal no evidence for the presence of Li2O2. A novel templating growth mechanism involving the use of iridium nanoparticles on the cathode surface may be responsible for the growth of crystalline LiO2. Our results demonstrate that the LiO2 formed in the Li-O2 battery is stable enough for the battery to be repeatedly charged and discharged with a very low charge potential (about 3.2 volts). We anticipate that this discovery will lead to methods of synthesizing and stabilizing LiO2, which could open the way to high-energy-density batteries based on LiO2 as well as to other possible uses of this compound, such as oxygen storage.

  9. Evaluation of five imidazopyrazinone-type chemiluminescent superoxide probes and their application to the measurement of superoxide anion generated by Listeria monocytogenes.

    PubMed

    Shimomura, O; Wu, C; Murai, A; Nakamura, H

    1998-05-01

    Superoxide-triggered chemiluminescence of five new imidazopyrazinone derivatives was investigated using the hypoxanthine-xanthine oxidase system as the source of superoxide anion. The results showed that they are highly sensitive and have favorable properties in measuring superoxide anion. With those new probes, the generation of superoxide anion from the bacteria Listeria monocytogenes was examined. The results confirmed the previous report that L. monocytogenes is an unusual organism that extracellularly and continuously generates a high level of superoxide anion in the presence of acetaldehyde. The data indicated that two of the probes, 3,7-dihydro-2-methyl-6-phenylethynylimidazo[1,2-a]pyrazin-3- one (4) and its methoxy derivative (5), are highly sensitive and useful in the measurements of superoxide anion and are clearly superior to 3,7-dihydro-2-methyl-6-(4-methoxyphenyl)imidazo[1,2-a]pyrazin-3-on e (MCLA), which-has been generally considered the most sensitive superoxide probe in the past. When tested at a probe concentration of 3.3 microM, the luminescence response and the signal-background ratio of compound 4 were 1.5 and 2.5 times those of MCLA, respectively, and the signal-background ratio of compound 5 was almost 15 times that of MCLA, though the luminescence response of this compound was slightly lower than that of MCLA. The low probe concentration used enhances the usefulness of probes in the measurements of superoxide in functioning biological systems.

  10. Theoretical determination of the alkali-metal superoxide bond energies

    NASA Technical Reports Server (NTRS)

    Partridge, Harry; Bauschlicher, Charles W., Jr.; Sodupe, Mariona; Langhoff, Stephen R.

    1992-01-01

    The bond dissociation energies for the alkali-metal superoxides have been computed using extensive Gaussian basis sets and treating electron correlation at the modified coupled-pair functional level. Our computed D0 values are 61.4, 37.2, 40.6, and 38.4 kcal/mol for LiO2, NaO2, KO2, and RbO2, respectively. These values, which are expected to be lower bounds and accurate to 2 kcal/mol, agree well with some of the older flame data, but rule out several recent experimental measurements.

  11. Production of superoxide dismutases from Proteus mirabilis and Proteus vulgaris.

    PubMed

    Dayton, T M; Diefenbach, K A; Fuller, M L; Valtos, J; Niederhoffer, E C

    1996-04-01

    Proteus mirabilis and Proteus vulgaris expressed a combination of superoxide dismutase (Sod) activities, which was assigned to FeSod1, FeSod2 and MnSod for P. mirabilis, and FeSod, MnSod and CuZnSod for P. vulgaris. Production of the Sod proteins was dependent on the availability of iron, whether cells were grown under anaerobiosis or aerobiosis and growth phase. Nalidixic acid and chloramphenicol inhibited cell growth and the iron- and dioxygen-dependent production of Sod. These results support the involvement of metal ions and redox status in the production of Proteus Sods.

  12. Zinc hydroxide stimulates superoxide production by rat alveolar macrophages.

    PubMed

    Ogino, K; Izumi, Y; Ishiyama, H; Murata, T; Kobayashi, H; Houbara, T

    1992-06-30

    The effect of zinc hydroxide on superoxide (O2-) production by rat alveolar macrophages was determined by chemiluminescence and by cytochrome c reduction. Zinc ions had no effect on the chemiluminescence of unstimulated alveolar macrophages. By contrast, zinc hydroxide (ZnOH2), a neutralized form of zinc ions, increased the chemiluminescence level and O2- release. Increased O2- release was inhibited by pertussis toxin, isoquinoline sulfonamide and pretreatment with EGTA. These findings indicate that zinc hydroxide formation from zinc compounds can stimulate the O2- production by alveolar macrophages by receptor-mediated and Ca(2+)-dependent process.

  13. Water induced dismutation of superoxide anion generates singlet molecular oxygen.

    PubMed

    Corey, E J; Mehrotra, M M; Khan, A U

    1987-06-15

    Direct spectroscopic measurement of 1268 nm singlet oxygen emission from KO2 suspensions at room temperature in three non-protonic solvents--CCl4, Cl2FCCClF2, and C6F14 by the action of water is reported. The results clearly show that the singlet oxygen generation is due to a water induced reaction, and suggest that one role of the enzyme superoxide dismutase may be the protection of biological structures, for example, lipid membranes, from degradation by singlet oxygen.

  14. Theoretical determination of the alkali-metal superoxide bond energies

    NASA Technical Reports Server (NTRS)

    Partridge, Harry; Bauschlicher, Charles W., Jr.; Sodupe, Mariona; Langhoff, Stephen R.

    1992-01-01

    The bond dissociation energies for the alkali-metal superoxides have been computed using extensive Gaussian basis sets and treating electron correlation at the modified coupled-pair functional level. Our computed D0 values are 61.4, 37.2, 40.6, and 38.4 kcal/mol for LiO2, NaO2, KO2, and RbO2, respectively. These values, which are expected to be lower bounds and accurate to 2 kcal/mol, agree well with some of the older flame data, but rule out several recent experimental measurements.

  15. Methylglyoxal as a scavenger for superoxide anion-radical.

    PubMed

    Shumaev, K B; Lankin, V Z; Konovalova, G G; Grechnikova, M A; Tikhaze, A K

    2016-07-01

    Methylglyoxal at a concentration of 5 mM caused a significant inhibition of superoxide anion radical (O2 (·-)) comparable to the effect of Tirone. In the process of O2 (·-) generation in the system of egg phosphatidylcholine liposome peroxidation induced by the azo-initiator AIBN, a marked inhibition of chemiluminescence in the presence of 100 mM methylglyoxal was found. At the same time, methylglyoxal did not inhibit free radical peroxidation of low-density lipoprotein particles, which indicates the absence of interaction with methylglyoxal alkoxyl and peroxyl polyenoic lipid radicals. These findings deepen information about the role of methylglyoxal in the regulation of free radical processes.

  16. Recycling ferrous sulfate via super-oxidant synthesis

    NASA Astrophysics Data System (ADS)

    Kanari, N.; Evrard, O.; Neveux, N.; Ninane, L.

    2001-11-01

    Hydrated ferrous sulfate, a by-product of the titanium-dioxide and steel-surface-treatment industries, is usually disposed of as waste at a significant extra cost for these industries. Due to tight environmental regulations in the European countries, waste disposal of ferrous sulfate will not be an acceptable solution in the near future. Consequently, the waste will have to be treated. Recently, ferrous sulfate was successfully used to synthesize a novel superoxidant material (potassium ferrate) containing iron in hexavalent state (FeVI). With ferrates synthesis, innovative applications are possible in different industrial sectors, such as treatment of water and wastewater and effluent decontamination.

  17. Detoxification of superoxide without production of H2O2: Antioxidant activity of superoxide reductase complexed with ferrocyanide

    PubMed Central

    Molina-Heredia, Fernando P.; Houée-Levin, Chantal; Berthomieu, Catherine; Touati, Danièle; Tremey, Emilie; Favaudon, Vincent; Adam, Virgile; Nivière, Vincent

    2006-01-01

    The superoxide radical O2·̅ is a toxic by-product of oxygen metabolism. Two O2·̅ detoxifying enzymes have been described so far, superoxide dismutase and superoxide reductase (SOR), both forming H2O2 as a reaction product. Recently, the SOR active site, a ferrous iron in a [Fe2+ (N-His)4 (S-Cys)] pentacoordination, was shown to have the ability to form a complex with the organometallic compound ferrocyanide. Here, we have investigated in detail the reactivity of the SOR–ferrocyanide complex with O2·̅ by pulse and γ-ray radiolysis, infrared, and UV-visible spectroscopies. The complex reacts very efficiently with O2·̅. However, the presence of the ferrocyanide adduct markedly modifies the reaction mechanism of SOR, with the formation of transient intermediates different from those observed for SOR alone. A one-electron redox chemistry appears to be carried out by the ferrocyanide moiety of the complex, whereas the SOR iron site remains in the reduced state. Surprisingly, the toxic H2O2 species is no longer the reaction product. Accordingly, in vivoexperiments showed that formation of the SOR–ferrocyanide complex increased the antioxidant capabilities of SOR expressed in an Escherichia coli sodA sodB recA mutant strain. Altogether, these data describe an unprecedented O2·̅ detoxification activity, catalyzed by the SOR–ferrocyanide complex, which does not conduct to the production of the toxic H2O2 species. PMID:17001016

  18. Superoxide anion-induced pain and inflammation depends on TNFα/TNFR1 signaling in mice.

    PubMed

    Yamacita-Borin, Fabiane Y; Zarpelon, Ana C; Pinho-Ribeiro, Felipe A; Fattori, Victor; Alves-Filho, Jose C; Cunha, Fernando Q; Cunha, Thiago M; Casagrande, Rubia; Verri, Waldiceu A

    2015-09-25

    Inhibition of tumor necrosis factor-alpha (TNFα) and superoxide anion production reduces inflammation and pain. The present study investigated whether superoxide anion-induced pain depends on TNFα signaling and the role of superoxide anion in TNFα-induced hyperalgesia to clarify the interrelation between these two mediators in the context of pain. Intraplantar injection of a superoxide anion donor (potassium superoxide) induced mechanical hyperalgesia (0.5-5h after injection), neutrophil recruitment (myeloperoxidase activity), and overt pain-like behaviors (paw flinching, paw licking, and abdominal writhings) in wild-type mice. Tumor necrosis factor receptor 1 deficiency (TNFR1-/-) and treatment of wild-type mice with etanercept (a soluble TNFR2 receptor that inhibits TNFα actions) inhibited superoxide anion-induced pain-like behaviors. TNFR1(-/-) mice were also protected from superoxide anion donor-induced oxidative stress, suggesting the role of this pathway in the maintenance of oxidative stress. Finally, we demonstrated that Apocynin (an NADPH oxidase inhibitor) or Tempol (a superoxide dismutase mimetic) treatment inhibited TNFα-induced paw mechanical hyperalgesia and neutrophil recruitment (myeloperoxidase activity). These results demonstrate that TNFα/TNFR1 signaling is important in superoxide anion-triggered pain and that TNFα/TNFR1 signaling amplifies the oxidative stress triggered by superoxide anion, which contributes to sustaining pain and inflammation.

  19. Superoxide dismutase SOD-1 modulates C. elegans pathogen avoidance behavior

    PubMed Central

    Horspool, Alexander M.; Chang, Howard C.

    2017-01-01

    The C. elegans nervous system mediates protective physiological and behavioral responses amid infection. However, it remains largely unknown how the nervous system responds to reactive oxygen species (ROS) activated by pathogenic microbes during infection. Here, we show superoxide dismutase-1 (SOD-1), an enzyme that converts superoxide into less toxic hydrogen peroxide and oxygen, functions in the gustatory neuron ASER to mediate C. elegans pathogen avoidance response. When C. elegans first encounters pathogenic bacteria P. aeruginosa, SOD-1 is induced in the ASER neuron. After prolonged P. aeruginosa exposure, ASER-specific SOD-1 expression is diminished. In turn, C. elegans starts to vacate the pathogenic bacteria lawn. Genetic knockdown experiments reveal that pathogen-induced ROS activate sod-1 dependent behavioral response non cell-autonomously. We postulate that the delayed aversive response to detrimental microbes may provide survival benefits by allowing C. elegans to temporarily utilize food that is tainted with pathogens as an additional energy source. Our data offer a mechanistic insight into how the nervous system mediates food-seeking behavior amid oxidative stress and suggest that the internal state of redox homeostasis could underlie the behavioral response to harmful microbial species. PMID:28322326

  20. Superoxide Dismutase Mimics: Chemistry, Pharmacology, and Therapeutic Potential

    PubMed Central

    Rebouças, Júlio S.; Spasojević, Ivan

    2010-01-01

    Abstract Oxidative stress has become widely viewed as an underlying condition in a number of diseases, such as ischemia–reperfusion disorders, central nervous system disorders, cardiovascular conditions, cancer, and diabetes. Thus, natural and synthetic antioxidants have been actively sought. Superoxide dismutase is a first line of defense against oxidative stress under physiological and pathological conditions. Therefore, the development of therapeutics aimed at mimicking superoxide dismutase was a natural maneuver. Metalloporphyrins, as well as Mn cyclic polyamines, Mn salen derivatives and nitroxides were all originally developed as SOD mimics. The same thermodynamic and electrostatic properties that make them potent SOD mimics may allow them to reduce other reactive species such as peroxynitrite, peroxynitrite-derived CO3·−, peroxyl radical, and less efficiently H2O2. By doing so SOD mimics can decrease both primary and secondary oxidative events, the latter arising from the inhibition of cellular transcriptional activity. To better judge the therapeutic potential and the advantage of one over the other type of compound, comparative studies of different classes of drugs in the same cellular and/or animal models are needed. We here provide a comprehensive overview of the chemical properties and some in vivo effects observed with various classes of compounds with a special emphasis on porphyrin-based compounds. Antioxid. Redox Signal. 13, 877–918. PMID:20095865

  1. Superoxide overproduction and kidney fibrosis: a new animal model

    PubMed Central

    Guimarães-Souza, Nadia Karina; Yamaleyeva, Liliya Marsovna; Lu, Baisong; Ramos, Ana Claudia Mallet de Souza; Bishop, Colin Edward; Andersson, Karl Erik

    2015-01-01

    Objective To establish whether the mutation in the Immp2L gene induces renal fibrosis and whether aging exacerbates renal morphology in mice. Methods Female mutant mice with mutation in the inner mitochondrial membrane peptidase 2-like protein at 3 and 18 months of age were used. Renal fibrosis was analyzed using classic fibrosis score, Masson’s trichrome staining, and analysis of profibrotic markers using real time polymerase chain reaction (superoxide dismutase 1, metalloproteinase-9, erythropoietin, transforming growth factor beta), and immunostaining (fibroblasts and Type IV collagen). Oxidative stress markers were determined by immunohistochemistry. The number of renal apoptotic cells was determined. Renal function was estimated by serum creatinine. Results Young mutant mice had significantly more glomerulosclerosis than age-matched mice (p=0.034). Mutant mice had more tubular casts (p=0.025), collagen deposition (p=0.019), and collagen type IV expression (p<0.001). Superoxide dismutase 1 expression was significantly higher in young mutants (p=0.038). Old mutants exhibited significantly higher expression of the fibroblast marker and macrophage marker (p=0.007 and p=0.012, respectively). The real time polymerase chain reaction of metalloproteinase-9 and erythropoietin were enhanced 2.5- and 6-fold, respectively, in old mutants. Serum creatinine was significantly higher in old mutants (p<0.001). Conclusion This mutation altered renal architecture by increasing the deposition of extracellular matrix, oxidative stress, and inflammation, suggesting a protective role of Immp2L against renal fibrosis. PMID:25993073

  2. Superoxide overproduction and kidney fibrosis: a new animal model.

    PubMed

    Guimarães-Souza, Nadia Karina; Yamaleyeva, Liliya Marsovna; Lu, Baisong; Ramos, Ana Claudia Mallet de Souza; Bishop, Colin Edward; Andersson, Karl Erik

    2015-01-01

    To establish whether the mutation in the Immp2L gene induces renal fibrosis and whether aging exacerbates renal morphology in mice. Female mutant mice with mutation in the inner mitochondrial membrane peptidase 2-like protein at 3 and 18 months of age were used. Renal fibrosis was analyzed using classic fibrosis score, Masson's trichrome staining, and analysis of profibrotic markers using real time polymerase chain reaction (superoxide dismutase 1, metalloproteinase-9, erythropoietin, transforming growth factor beta), and immunostaining (fibroblasts and Type IV collagen). Oxidative stress markers were determined by immunohistochemistry. The number of renal apoptotic cells was determined. Renal function was estimated by serum creatinine. Young mutant mice had significantly more glomerulosclerosis than age-matched mice (p=0.034). Mutant mice had more tubular casts (p=0.025), collagen deposition (p=0.019), and collagen type IV expression (p<0.001). Superoxide dismutase 1 expression was significantly higher in young mutants (p=0.038). Old mutants exhibited significantly higher expression of the fibroblast marker and macrophage marker (p=0.007 and p=0.012, respectively). The real time polymerase chain reaction of metalloproteinase-9 and erythropoietin were enhanced 2.5- and 6-fold, respectively, in old mutants. Serum creatinine was significantly higher in old mutants (p<0.001). This mutation altered renal architecture by increasing the deposition of extracellular matrix, oxidative stress, and inflammation, suggesting a protective role of Immp2L against renal fibrosis.

  3. A superoxide sensor based on a multilayer cytochrome c electrode.

    PubMed

    Beissenhirtz, Moritz K; Scheller, Frieder W; Lisdat, Fred

    2004-08-15

    A novel multilayer cytochrome c electrode for the quantification of superoxide radical concentrations is introduced. The electrode consists of alternating layers of cytochrome c and poly(aniline(sulfonic acid)) on a gold wire electrode. The formation of multilayer structures was proven by SPR experiments. Assemblies with 2-15 protein layers showed electrochemical communication with the gold electrode. For every additional layer, a substantial increase in electrochemically active cytochrome c (cyt. c) was found. For electrodes of more than 10 layers, the increase was more than 1 order of magnitude as compared to monolayer electrode systems. Thermodynamic and kinetic parameters of the electrodes were characterized. The mechanism of electron transfer within the multilayer assembly was studied, with results suggesting a protein-protein electron-transfer model. Electrodes of 2-15 layers were applied to the in vitro quantification of enzymatically generated superoxide, showing superior sensitivity as compared to a monolayer-based sensor. An electrode with 6 cyt. c/PASA layers showed the highest sensitivity of the systems studied, giving an increase in sensitivity of half an order of magnitude versus the that of the monolayer electrode. The stability of the system was optimized using thermal treatment, resulting in no loss in sensor signal or protein loading after 10 successive measurements or 2 days of storage.

  4. A lithium-oxygen battery based on lithium superoxide.

    SciTech Connect

    Lu, Jun; Lee, Yun Jung; Luo, Xiangyi; Lau, Kah Chun; Wen, Jianguo; Wang, Hsien-Hau; Zhai, Dengyun; Miller, Dean; Jeong, Yo-Sub; Park, Jin-Bum; Curtiss, Larry A.; Amine, Khalil

    2016-01-11

    Although the superoxide of lithium (LiO2) is believed to be a key intermediate in Li-O2 batteries leading to the formation of lithium peroxide, LiO2 has never been observed in its pure state. In this work, we provide evidence that use of a cathode based on a reduced graphene oxide with Ir nanoparticles in a Li-O2 battery results in a LiO2 discharge product formed by single electron transfer without further electron transfer or disproportionation to form Li2O2. High energy X-ray diffraction (HE-XRD) patterns indicates the presence of crystalline LiO2 with no evidence of Li2O2 or Li2O. The HEXRD studies as a function of time also show that LiO2 can be stable in its crystalline form after one week of aging in the presence of electrolyte. The results provide evidence that LiO2 is stable enough that it can be repeatedly charged and discharged with a very low charge potential (~3.2 V) and may open the avenue for a lithium superoxide-based battery.

  5. Superoxide dismutase from the extremely halophilic archaebacterium Halobacterium cutirubrum.

    PubMed Central

    May, B P; Dennis, P P

    1987-01-01

    Halobacterium cutirubrum, a member of the archaebacteria, contains one superoxide dismutase (EC 1.15.1.1). This enzyme functions in the high-ionic-strength intracellular environment and protects the organism against the toxic effects of the superoxide anion. The enzyme has been purified to about 90% homogeneity by a four-step procedure which never removes it from conditions of high ionic strength. The subunits of the purified enzyme have a molecular weight of 25,000 and are possibly in tetrameric association. The enzyme shows anomalously high resistance to azide inhibition and sensitivity to inactivation by hydrogen peroxide. Metal analysis indicates 0.2 atom of Mn, less than 0.03 atom of Cu, and less than 0.001 atom of Fe per subunit. The low content of Mn may explain the low specific activity found for this enzyme compared with that of eubacterial enzymes. Optimum activity occurs in 2 M KCl; KCl gives about twice as much activity as NaCl over the range of 2 to 4 M. The enzyme appears to be related to those isolated from other archaebacteria but also exhibits several novel features. Images PMID:3104309

  6. The Cu, Zn Superoxide Dismutase: Not Only a Dismutase Enzyme

    PubMed Central

    Mondola, Paolo; Damiano, Simona; Sasso, Anna; Santillo, Mariarosaria

    2016-01-01

    The Cu,Zn superoxide dismutase (SOD1) is an ubiquitary cytosolic dimeric carbohydrate free molecule, belonging to a family of isoenzymes involved in the scavenger of superoxide anions. This effect certainly represents the main and well known function ascribed to this enzyme. Here we highlight new aspects of SOD1 physiology that point out some inedited effects of this enzyme in addition to the canonic role of oxygen radical enzymatic dismutation. In the last two decades our research group produced many data obtained in in vitro studies performed in many cellular lines, mainly neuroblastoma SK-N-BE cells, indicating that this enzyme is secreted either constitutively or after depolarization induced by high extracellular K+ concentration. In addition, we gave many experimental evidences showing that SOD1 is able to stimulate, through muscarinic M1 receptor, pathways involving ERK1/2, and AKT activation. These effects are accompanied with an intracellular calcium increase. In the last part of this review we describe researches that link deficient extracellular secretion of mutant SOD1G93A to its intracellular accumulation and toxicity in NSC-34 cells. Alternatively, SOD1G93A toxicity has been attributed to a decrease of Km for H2O2 with consequent OH radical formation. Interestingly, this last inedited effect of SOD1G93A could represent a gain of function that could be involved in the pathogenesis of familial Amyotrophic Lateral Sclerosis (fALS). PMID:27965593

  7. Superoxide dismutase activity in normal and preeclamptic placentas.

    PubMed

    Wiktor, H; Kankofer, M

    1998-12-01

    A deficiency in superoxide dismutase (SOD) activity in preeclamptic placentas can lead to an excess of superoxide radicals and may be responsible for the development and the severity of preeclampsia (PE). Our studies were undertaken in order to determine placental SOD activity and to investigate their association with the development and the severity of PE. The activity of SOD was determined using a spectrophotometric method in 22 placentas from normal term pregnancies (group K), 24 placentas from pregnancies complicated by severe PE (group PE), and 21 placentas from pregnancies complicated by severe PE and intrauterine growth retardation (IUGR) (group PEI). Mean activity of SOD (MSOD) in 45 preeclamptic placentas 3.89 +/- 1.32 (M +/- SD) was significantly lower (P = 0.008) as compared to MSOD in the group K (6.75 +/- 1.96). MSOD in the PEI group (3.5 +/- 1.29) was significantly lower (P = 0.03) as compared to MSOD in the group K. MSOD in the PE group (4.23 +/- 1.25) was lower than MSOD in the group K, but this difference was not statistically significant (P = 0.11). MSOD in the group PEI was lower as compared to MSOD in the PE group, however this difference was not statistically significant (P = 0.23). The studies revealed decreased SOD activity in preeclamptic placentas in comparison to normal placentas.

  8. Determination of superoxide dismutase mimetic activity in common culinary herbs.

    PubMed

    Chohan, Magali; Naughton, Declan P; Opara, Elizabeth I

    2014-01-01

    Under conditions of oxidative stress, the removal of superoxide, a free radical associated with chronic inflammation, is catalysed by superoxide dismutase (SOD). Thus in addition to acting as an antioxidant, SOD may also be utilized as an anti-inflammatory agent. Some plant derived foods have been shown to have SOD mimetic (SODm) activity however it is not known if this activity is possessed by culinary herbs which have previously been shown to possess both antioxidant and anti-inflammatory properties. The aim of the study was to ascertain if the culinary herbs rosemary, sage and thyme possess SODm activity, and to investigate the influence of cooking and digestion on this activity. Transition metal ion content was also determined to establish if it could likely contribute to any SODm activity detected. All extracts of uncooked (U), cooked (C) and cooked and digested (C&D) herbs were shown to possess SODm activity, which was significantly correlated with previously determined antioxidant and anti-inflammatory activities of these herbs. SODm activity was significantly increased following (C) and (C&D) for rosemary and sage only. The impact of (C) and (C&D) on the SODm for thyme may have been influenced by its transition metal ion content. SODm activity may contribute to the herbs' antioxidant and anti-inflammatory activities however the source and significance of this activity need to be established.

  9. Molecular characterization of two superoxide dismutases from Hydra vulgaris

    PubMed Central

    Dash, Bhagirathi; Metz, Richard; Huebner, Henry J.; Porter, Weston; Phillips, Timothy D.

    2007-01-01

    Apparent full-length cDNA sequences coding for manganese superoxide dismutase (HvMnSOD) and extracellular superoxide dismutase (HvEC-SOD) were isolated from Hydra vulgaris in order to understand their expression and 3D structures; and explore their possibility of being used as for biomarkers for environmental stress and toxicity. The deduced HvMnSOD protein consists of 219 amino acids of which first 21 amino acids constitute a presumed mitochondria-targeting signal peptide whereas HvEC-SOD protein consists of 189 amino acids of which first 19 amino acids constitute a presumed signal peptide. Molecular model generated for HvMnSOD displayed the N-terminal long alpha antiparallel hairpin and the C-terminal mixed alpha/beta fold characteristic of MnSODs and that for HvEC-SOD displayed the characteristic CuZnSOD beta-barrel fold. Hydrae subjected to thermal, starvation, metal and oxidative stress responded by regulating MnSOD and EC-SOD mRNA transcription. These results indicated that these genes are involved in the cellular stress response and (anti)oxidative processes triggered by stressor and contaminant exposure. Hence the expression of these SODs in hydra may have potential as molecular biomarkers for assessing stress, toxicity and pro-oxidant quality of chemicals and aquatic environmental quality. PMID:17150313

  10. The Cu, Zn Superoxide Dismutase: Not Only a Dismutase Enzyme.

    PubMed

    Mondola, Paolo; Damiano, Simona; Sasso, Anna; Santillo, Mariarosaria

    2016-01-01

    The Cu,Zn superoxide dismutase (SOD1) is an ubiquitary cytosolic dimeric carbohydrate free molecule, belonging to a family of isoenzymes involved in the scavenger of superoxide anions. This effect certainly represents the main and well known function ascribed to this enzyme. Here we highlight new aspects of SOD1 physiology that point out some inedited effects of this enzyme in addition to the canonic role of oxygen radical enzymatic dismutation. In the last two decades our research group produced many data obtained in in vitro studies performed in many cellular lines, mainly neuroblastoma SK-N-BE cells, indicating that this enzyme is secreted either constitutively or after depolarization induced by high extracellular K(+) concentration. In addition, we gave many experimental evidences showing that SOD1 is able to stimulate, through muscarinic M1 receptor, pathways involving ERK1/2, and AKT activation. These effects are accompanied with an intracellular calcium increase. In the last part of this review we describe researches that link deficient extracellular secretion of mutant SOD1(G93A) to its intracellular accumulation and toxicity in NSC-34 cells. Alternatively, SOD1(G93A) toxicity has been attributed to a decrease of Km for H2O2 with consequent OH radical formation. Interestingly, this last inedited effect of SOD1(G93A) could represent a gain of function that could be involved in the pathogenesis of familial Amyotrophic Lateral Sclerosis (fALS).

  11. Metabolic alterations in yeast lacking copper-zinc superoxide dismutase.

    PubMed

    Sehati, Sadaf; Clement, Matthew H S; Martins, Jake; Xu, Lei; Longo, Valter D; Valentine, Joan S; Gralla, Edith B

    2011-06-01

    Yeast lacking copper-zinc superoxide dismutase (sod1∆) have a number of oxygen-dependent defects, including auxotrophies for lysine and methionine and sensitivity to oxygen. Here we report additional defects in metabolic regulation. Under standard growth conditions with glucose as the carbon source, yeast undergo glucose repression in which mitochondrial respiration is deemphasized, energy is mainly derived from glycolysis, and ethanol is produced. When glucose is depleted, the diauxic shift is activated, in which mitochondrial respiration is reemphasized and stress resistance increases. We find that both of these programs are adversely affected by the lack of Sod1p. Key events in the diauxic shift do not occur and sod1∆ cells do not utilize ethanol and stop growing. The ability to shift to growth on ethanol is gradually lost as time in culture increases. In early stages of culture, sod1∆ cells consume more oxygen and have more mitochondrial mass than wild-type cells, indicating that glucose repression is not fully activated. These changes are at least partially dependent on the activity of the Hap2,3,4,5 complex, as indicated by CYC1-lacZ reporter assays. These changes may indicate a role for superoxide in metabolic signaling and regulation and/or a role for glucose derepression in defense against oxidative stress. Copyright © 2011 Elsevier Inc. All rights reserved.

  12. Conformationally gated metal uptake by apo-manganese superoxide dismutase

    PubMed Central

    Whittaker, Mei M.; Whittaker, James W.

    2009-01-01

    Metal uptake by apo-manganese superoxide dismutase in vitro is a complex process exhibiting multiphase “gated” reaction kinetics and a striking sigmoidal temperature profile that has led to a model of conformationally gated metal binding, requiring conversion between “closed” and “open” forms. The present work systematically explores the structural determinants of metal binding in both WT apoprotein and mutational variants as a test of mechanistic models. The pH dependence of metallation under physiological conditions (37°C) shows it is linked to ionization of a single proton with a pKa of 7.7. Size exclusion chromatography demonstrates that the apoprotein is dimeric even when it is fully converted to the open form. The role of molecular motions in metal binding has been probed by using disulfide engineering to introduce covalent constraints into the protein. While restricting motion at domain interfaces has no effect, constraining the subunit interface significantly perturbs metal uptake, but does not prevent the process. Mutagenesis of residues in the active site environment results in a dramatic shift in the transition temperature by as much as 20°C or loss of pH-sensitivity. Based on these results, a mechanism for metal uptake by manganese superoxide dismutase is proposed involving reorientation of active site residues to form a metal entry channel. PMID:18841998

  13. Biologically active metal-independent superoxide dismutase mimics

    SciTech Connect

    Mitchell, J.B.; Samuni, A.; Krishna, M.C.; DeGraff, W.G.; Ahn, M.S.; Samuni, U.; Russo, A. )

    1990-03-20

    Superoxide dismutase (SOD) is an enzyme that detoxifies superoxide (O2.-), a potentially toxic oxygen-derived species. Attempts to increase intracellular concentrations of SOD by direct application are complicated because SOD, being a relatively large molecule, does not readily cross cell membranes. We have identified a set of stable nitroxides that possess SOD-like activity, have the advantage of being low molecular weight, membrane permeable, and metal independent, and at pH 7.0 have reaction rate constants with O2.- ranging from 1.1 x 10(3) to 1.3 x 10(6) M-1 s-1. These SOD mimics protect mammalian cells from damage induced by hypoxanthine/xanthine oxidase and H{sub 2}O{sub 2}, although they exhibit no catalase-like activity. In addition, the nitroxide SOD mimics rapidly oxidize DNA-FeII and thus may interrupt the Fenton reaction and prevent formation of deleterious OH radicals and/or higher oxidation states of metal ions. Whether by SOD-like activity and/or interception of an electron from redox-active metal ions they protect cells from oxidative stress and may have use in basic and applied biological studies.

  14. Mechanism of Action of Sulforaphane as a Superoxide Radical Anion and Hydrogen Peroxide Scavenger by Double Hydrogen Transfer: A Model for Iron Superoxide Dismutase.

    PubMed

    Prasad, Ajit Kumar; Mishra, P C

    2015-06-25

    The mechanism of action of sulforaphane as a scavenger of superoxide radical anion (O2(•-)) and hydrogen peroxide (H2O2) was investigated using density functional theory (DFT) in both gas phase and aqueous media. Iron superoxide dismutase (Fe-SOD) involved in scavenging superoxide radical anion from biological media was modeled by a complex consisting of the ferric ion (Fe(3+)) attached to three histidine rings. Reactions related to scavenging of superoxide radical anion by sulforaphane were studied using DFT in the presence and absence of Fe-SOD represented by this model in both gas phase and aqueous media. The scavenging action of sulforaphane toward both superoxide radical anion and hydrogen peroxide was found to involve the unusual mechanism of double hydrogen transfer. It was found that sulforaphane alone, without Fe-SOD, cannot scavenge superoxide radical anion in gas phase or aqueous media efficiently as the corresponding reaction barriers are very high. However, in the presence of Fe-SOD represented by the above-mentioned model, the scavenging reactions become barrierless, and so sulforaphane scavenges superoxide radical anion by converting it to hydrogen peroxide efficiently. Further, sulforaphane was found to scavenge hydrogen peroxide also very efficiently by converting it into water. Thus, the mechanism of action of sulforaphane as an excellent antioxidant has been unravelled.

  15. The Influence of Extracellular Superoxide on Iron Redox Chemistry and Bioavailability to Aquatic Microorganisms

    PubMed Central

    Rose, Andrew L.

    2012-01-01

    Superoxide, the one-electron reduced form of dioxygen, is produced in the extracellular milieu of aquatic microbes through a range of abiotic chemical processes and also by microbes themselves. Due to its ability to promote both oxidative and reductive reactions, superoxide may have a profound impact on the redox state of iron, potentially influencing iron solubility, complex speciation, and bioavailability. The interplay between iron, superoxide, and oxygen may also produce a cascade of other highly reactive transients in oxygenated natural waters. For microbes, the overall effect of reactions between superoxide and iron may be deleterious or beneficial, depending on the organism and its chemical environment. Here I critically discuss recent advances in understanding: (i) sources of extracellular superoxide in natural waters, with a particular emphasis on microbial generation; (ii) the chemistry of reactions between superoxide and iron; and (iii) the influence of these processes on iron bioavailability and microbial iron nutrition. PMID:22514548

  16. Some dinophycean red tide plankton species generate a superoxide scavenging substance.

    PubMed

    Sato, Emiko; Niwano, Yoshimi; Matsuyama, Yukihiko; Kim, Daekyung; Nakashima, Takuji; Oda, Tatsuya; Kohno, Masahiro

    2007-03-01

    Recent studies indicate that some raphidophycean red tide flagellates produce substances able to scavenge superoxide, whereas there have been no reports on superoxide scavenger production by dinophycean red tide flagellates. In this study, we examined the superoxide-scavenging activity of aqueous extracts from dinophycean red tide flagellates, Gymnodinium spp., Scrippsiella trochoidea, and Karenia sp., by a luminol analog L-012-dependent chemiluminescence (CL) method and an electron spin resonance (ESR)-spin trapping method, and compared the activity to that of raphidophycean red tide flagellates, Chattonella spp., Heterosigma akashiwo, and Fibrocapsa japonica. In the experiment applying the L-012-dependent CL method, only the aqueous extracts from raphidophycean red tide flagellates showed superoxide-scavenging activity. On the other hand, applying the ESR-spin trapping method, we found that the aqueous extracts from dinophycean red tide flagellates also showed superoxide-scavenging activity. This is the first report on the production of a superoxide-scavenger by dinophycean red tide flagellates.

  17. Superoxide Mediates the Toxicity of Paraquat for Chinese Hamster Ovary Cells

    NASA Astrophysics Data System (ADS)

    Bagley, Ann C.; Krall, Judith; Lynch, Robert E.

    1986-05-01

    The roles of superoxide and H2O2 in the cytotoxicity of paraquat were assessed in Chinese hamster ovary cells. Neither catalase nor superoxide dismutase inhibited the loss of ability to form colonies when added to the medium. When introduced into the cells, superoxide dismutase but not catalase inhibited the toxicity of paraquat. That superoxide dismutase acted by its known catalytic action is shown by the loss of inhibition when the enzyme was inactivated by H2O2 before being introduced into the cells. The lack of inhibition by catalase, by dimethyl sulfoxide, and by desferoxamine suggests that the toxicity is not mediated by a reaction between H2O2 and superoxide to engender the hydroxyl radical. Exposure of Chinese hamster ovary cells to paraquat may be a suitable means to determine the effects of superoxide anion in cultured cells and the ways in which cells can resist this toxic action.

  18. Superoxide dismutase abolishes the platelet-derived growth factor-induced release of prostaglandin E2 by blocking induction of nitric oxide synthase: role of superoxide.

    PubMed

    Kelner, M J; Uglik, S F

    1995-09-10

    The ability of platelet-derived growth factor (PDGF) to induce prostaglandin E2 (PGE2) release in fibroblasts is abolished when copper-zinc superoxide dismutase activity is increased by transfection of an expression vector. The effect is specific to copper-zinc superoxide dismutase as glutathione peroxidase-overexpressing NIH3T3 cells, again produced by transfection of an expression vector, retain the ability to release PGE2 in response to growth factor stimulation. The defect in PDGF-induced PGE2 release occurs prior to action of prostaglandin H synthase/cyclooxygenase as release of arachadonic acid (in response to PDGF) does not occur in the superoxide dismutase-overexpressing clones. The defect in PDGF-induced release of PGE2 in superoxide dismutase-overexpressing clones differs from the defect found in pEJ-ras-transformed clones. The parent cells, the glutathione peroxidase-expressing cells, and the superoxide dismutase-overexpressing cells all release PGE2 in response to exogenous nitric oxide, whereas the pEJ-ras-transformed cells do not. The glutathione peroxidase-expressing cells also retained the ability to release nitrite in response to PDGF, whereas the superoxide dismutase-expressing clones do not. PDGF stimulates nitric oxide synthase activity in NIH3T3 cells, but not in the superoxide dismutase-expressing clones. These results indicate that superoxide dismutase overexpression blocks the PDGF-induced release of PGE2 by blocking induction of nitric oxide synthase. This indicates that the increase of nitric oxide synthase induced by PDGF is mediated in part by production of superoxide. These findings link cellular oxygen radical homeostasis to three different classes of messenger molecules (growth factors, nitric oxide, and prostaglandins).

  19. A lithium–oxygen battery based on lithium superoxide

    SciTech Connect

    Lu, Jun; Jung Lee, Yun; Luo, Xiangyi; Chun Lau, Kah; Asadi, Mohammad; Wang, Hsien-Hau; Brombosz, Scott; Wen, Jianguo; Zhai, Dengyun; Chen, Zonghai; Miller, Dean J.; Sub Jeong, Yo; Park, Jin-Bum; Zak Fang, Zhigang; Kumar, Bijandra; Salehi-Khojin, Amin; Sun, Yang-Kook; Curtiss, Larry A.; Amine, Khalil

    2016-01-11

    Batteries based on sodium superoxide and on potassium superoxide have recently been reported(1-3). But, there have been no reports of a battery based on lithium superoxide (LiO2), despite much research(4-8) into the lithium-oxygen (Li-O2) battery because of its potential high energy density. Several studies(9-16) of Li-O2 batteries have found evidence of LiO2 being formed as one component of the discharge product along with lithium peroxide (Li2O2). In addition, theoretical calculations have indicated that some forms of LiO2 may have a long lifetime(17). Our studies also suggest that it might be possible to form LiO2 alone for use in a battery. However, solid LiO2 has been difficult to synthesize in pure form(18) because it is thermodynamically unstable with respect to disproportionation, giving Li2O2 (refs 19, 20). We show that crystalline LiO2 can be stabilized in a Li-O2 battery by using a suitable graphene-based cathode. Various characterization techniques reveal no evidence for the presence of Li2O2. A novel templating growth mechanism involving the use of iridium nanoparticles on the cathode surface may be responsible for the growth of crystalline LiO2. Our results demonstrate that the LiO2 formed in the Li-O2 battery is stable enough for the battery to be repeatedly charged and discharged with a very low charge potential (about 3.2 volts). We also anticipate that this discovery will lead to methods of synthesizing and stabilizing LiO2, which could open the way to high-energy-density batteries based on LiO2 as well as to other possible uses of this compound, such as oxygen storage.

  20. The role of superoxide anion radical in the reduction of ferritin iron by xanthine oxidase.

    PubMed Central

    Williams, D M; Lee, G R; Cartwright, G E

    1974-01-01

    Superoxide dismutase exerted a pronounced inhibitory effect upon xanthine oxidase-mediated reduction of iron in ferritin, ferric chloride, or ferric ADP. Maximal inhibition was observed when the superoxide dismutase concentration was only about 1% of that found in normal porcine liver. These observations indicate that superoxide anion radical is an intermediate in the reduction of iron by xanthine oxidase in vitro but not in vivo. PMID:11344583

  1. Cadmium activates both diphenyleneiodonium- and rotenone-sensitive superoxide production in barley root tips.

    PubMed

    Tamás, Ladislav; Mistrík, Igor; Zelinová, Veronika

    2016-12-01

    Mild Cd stress-activated diphenyleneiodonium-sensitive superoxide production is utilized in root morphogenic responses, while severe Cd stress-induced robust rotenone-sensitive superoxide generation may lead to cell and root death. In barley, even a few minute exposure of roots to Cd concentration higher than 10 µM evoked a strong superoxide generation in the root transition zone. This superoxide generation was strongly inhibited by the inhibition of mitochondrial electron flow into complex III in the presence of the mitochondrial complex I inhibitor rotenone. Similarly, the superoxide generation induced by antimycin A, an inhibitor of mitochondrial complex III, was considerably reduced by rotenone, suggesting the involvement of complex III also in the severe Cd stress-induced superoxide generation. This severe Cd stress-induced superoxide generation was followed by an extensive cell death in this part of the root tip, which similar to the superoxide generation, was eliminated by rotenone co-treatment. In turn, mild Cd stress-induced diphenyleneiodonium (DPI)-sensitive superoxide generation was observed only in the post-stressed roots, suggesting that it is not directly associated with Cd toxicity. Diphenyleneiodonium, an inhibitor of NADPH oxidase, markedly inhibited the mild Cd stress-induced radial expansion of root apex, indicating that enhanced DPI-sensitive superoxide production is required for rapid isotropic cell growth. Severe Cd stress, probably through the inhibition of complex III, caused a rapid and robust superoxide generation leading to cell and/or root death. By contrast, mild Cd stress did not evoke oxidative stress, and the enhanced DPI-sensitive superoxide generation is utilized in adaptive morphogenic responses.

  2. Superoxide disproportionation driven by zinc complexes with various steric and electrostatic properties.

    PubMed

    Wada, Akira; Jitsukawa, Koichiro; Masuda, Hideki

    2013-11-18

    Attractive models: Synthetic Zn(II) complexes were investigated as models of copper-zinc superoxide dismutase. Superoxide underwent a unique disproportionation reaction in the electrostatic sphere of the complexes (see picture; bpy=2,2'-bipyridyl). The effectiveness of the Zn(II) complexes in inducing the disproportionation of superoxide depended on both the Lewis acidity and the coordination geometry of the Zn center.

  3. Superoxide Induces Neutrophil Extracellular Trap Formation in a TLR-4 and NOX-Dependent Mechanism

    PubMed Central

    Al-Khafaji, Ahmed B; Tohme, Samer; Yazdani, Hamza Obaid; Miller, David; Huang, Hai; Tsung, Allan

    2016-01-01

    Neutrophils constitute the early innate immune response to perceived infectious and sterile threats. Neutrophil extracellular traps (NETs) are a novel mechanism to counter pathogenic invasion and sequelae of ischemia, including cell death and oxidative stress. Superoxide is a radical intermediate of oxygen metabolism produced by parenchymal and nonparenchymal hepatic cells, and is a hallmark of oxidative stress after liver ischemia-reperfusion (I/R). While extracellular superoxide recruits neutrophils to the liver and initiates sterile inflammatory injury, it is unknown whether superoxide induces the formation of NETs. We hypothesize that superoxide induces NET formation through a signaling cascade involving Toll-like receptor 4 (TLR-4) and neutrophil NADPH oxidase (NOX). We treated neutrophils with extracellular superoxide and observed NET DNA release, histone H3 citrullination and increased levels of MPO-DNA complexes occurring in a TLR-4–dependent manner. Inhibition of superoxide generation by allopurinol and inhibition of NOX by diphenyleneiodonium prevented NET formation. When mice were subjected to warm liver I/R, we found significant NET formation associated with liver necrosis and increased serum ALT in TLR-4 WT but not TLR-4 KO mice. To reduce circulating superoxide, we pretreated mice undergoing I/R with allopurinol and N-acetylcysteine, which resulted in decreased NETs and ameliorated liver injury. Our study demonstrates a requirement for TLR-4 and NOX in superoxide-induced NETs, and suggests involvement of superoxide-induced NETs in pathophysiologic settings. PMID:27453505

  4. Singlet oxygen production in the reaction of superoxide with organic peroxides.

    PubMed

    MacManus-Spencer, Laura A; Edhlund, Betsy L; McNeill, Kristopher

    2006-01-20

    [reaction: see text] A selective chemiluminescent probe for singlet oxygen has been employed to detect and quantify singlet oxygen in the reactions of superoxide with organic peroxides. The production of singlet oxygen has been quantified in the reaction of superoxide with benzoyl peroxide (BP). No singlet oxygen was detected in the reactions of superoxide with cumyl peroxide, tert-butyl peroxide, or tert-butyl hydroperoxide. On the basis of these results and on the temperature dependence of the reaction, we proposed a mechanism for singlet oxygen formation in the reaction of superoxide with BP.

  5. A simplified hydroethidine method for fast and accurate detection of superoxide production in isolated mitochondria.

    PubMed

    Back, Patricia; Matthijssens, Filip; Vanfleteren, Jacques R; Braeckman, Bart P

    2012-04-01

    Because superoxide is involved in various physiological processes, many efforts have been made to improve its accurate quantification. We optimized and validated a superoxide-specific and -sensitive detection method. The protocol is based on fluorescence detection of the superoxide-specific hydroethidine (HE) oxidation product, 2-hydroxyethidium. We established a method for the quantification of superoxide production in isolated mitochondria without the need for acetone extraction and purification chromatography as described in previous studies. Copyright © 2012 Elsevier Inc. All rights reserved.

  6. Production of superoxide ions by leukocytes of the American alligator (Alligator mississippiensis).

    PubMed

    Merchant, Mark; Williams, Stetson; Hardy, Ross

    2009-01-01

    This study was conducted to characterize the production of superoxide ions by leukocytes in whole blood of the American alligator (Alligator mississippiensis). We used WST-1, a tetrazolium salt which can be reduced to a water-soluble formazan compound with high molar absorptivity at 438 nm, to probe the production of superoxide by alligator leukocytes. Incubation of alligator whole blood with WST-1 resulted in a time- and concentration-dependent increase in absorbance of the plasma at 438 nm. The reduction of WST-1 was inhibited in a concentration-dependent manner by superoxide dismutase, an enzyme that catalyzes the reduction of superoxide to peroxide, confirming that the reduction of WST-1 was due to the presence of superoxide. Treatment of whole blood with nitrotetrazolium blue (NBT) resulted in the staining of heterophils and monocytes, enforcing the idea that that the production of superoxide is due to the presence of leukocytes, and not other blood cell components. It is interesting to note that the production of superoxide by the alligator leukocytes required no external stimulation while human leukocytes must be stimulated with an immunological challenge before producing superoxide. This is the first report of the production of superoxide as an innate immune mechanism in crocodilians.

  7. Permeability transition pore-mediated mitochondrial superoxide flashes regulate cortical neural progenitor differentiation.

    PubMed

    Hou, Yan; Mattson, Mark P; Cheng, Aiwu

    2013-01-01

    In the process of neurogenesis, neural progenitor cells (NPCs) cease dividing and differentiate into postmitotic neurons that grow dendrites and an axon, become excitable, and establish synapses with other neurons. Mitochondrial biogenesis and aerobic metabolism provide energy substrates required to support the differentiation, growth and synaptic activity of neurons. Mitochondria may also serve signaling functions and, in this regard, it was recently reported that mitochondria can generate rapid bursts of superoxide (superoxide flashes), the frequency of which changes in response to environmental conditions and signals including oxygen levels and Ca(2+) fluxes. Here we show that the frequency of mitochondrial superoxide flashes increases as embryonic cerebral cortical neurons differentiate from NPCs, and provide evidence that the superoxide flashes serve a signaling function that is critical for the differentiation process. The superoxide flashes are mediated by mitochondrial permeability transition pore (mPTP) opening, and pharmacological inhibition of the mPTP suppresses neuronal differentiation. Moreover, superoxide flashes and neuronal differentiation are inhibited by scavenging of mitochondrial superoxide. Conversely, manipulations that increase superoxide flash frequency accelerate neuronal differentiation. Our findings reveal a regulatory role for mitochondrial superoxide flashes, mediated by mPTP opening, in neuronal differentiation.

  8. Selective superoxide generation within mitochondria by the targeted redox cycler MitoParaquat.

    PubMed

    Robb, Ellen L; Gawel, Justyna M; Aksentijević, Dunja; Cochemé, Helena M; Stewart, Tessa S; Shchepinova, Maria M; Qiang, He; Prime, Tracy A; Bright, Thomas P; James, Andrew M; Shattock, Michael J; Senn, Hans M; Hartley, Richard C; Murphy, Michael P

    2015-12-01

    Superoxide is the proximal reactive oxygen species (ROS) produced by the mitochondrial respiratory chain and plays a major role in pathological oxidative stress and redox signaling. While there are tools to detect or decrease mitochondrial superoxide, none can rapidly and specifically increase superoxide production within the mitochondrial matrix. This lack impedes progress, making it challenging to assess accurately the roles of mitochondrial superoxide in cells and in vivo. To address this unmet need, we synthesized and characterized a mitochondria-targeted redox cycler, MitoParaquat (MitoPQ) that comprises a triphenylphosphonium lipophilic cation conjugated to the redox cycler paraquat. MitoPQ accumulates selectively in the mitochondrial matrix driven by the membrane potential. Within the matrix, MitoPQ produces superoxide by redox cycling at the flavin site of complex I, selectively increasing superoxide production within mitochondria. MitoPQ increased mitochondrial superoxide in isolated mitochondria and cells in culture ~a thousand-fold more effectively than untargeted paraquat. MitoPQ was also more toxic than paraquat in the isolated perfused heart and in Drosophila in vivo. MitoPQ enables the selective generation of superoxide within mitochondria and is a useful tool to investigate the many roles of mitochondrial superoxide in pathology and redox signaling in cells and in vivo.

  9. Superoxide dismutase activity in radioresistant tissues of irradiated rabbits.

    PubMed

    Stoklasová, A; Kovárová, H; Ledvina, M

    1992-01-01

    The activities of Cu, Zn-containing superoxide dismutase were studied in radioresistant tissues (liver, brain, erythrocytes) of whole-body irradiated rabbits with 6.0 Gy and 24.0 Gy with local shielding. No significant changes were observed after irradiation with 6.0 Gy. Both the changes in Cu, Zn-SOD activity and the protein concentrations were more pronounced after exposure to 24.0 Gy with local shielding of the head and abdominal region. The dose on the shielded regions was about 6.0 Gy. Local shielding of rabbits irradiated with a lethal dose 24.0 Gy influenced positively the survival of animals. However, the decrease in SOD activity on 60th day after irradiation seems to be unfavourable for further survival of rabbits, if we accept that SOD content in tissue is maintained at a rather constant level.

  10. Reduced superoxide dismutase activity in xeroderma pigmentosum fibroblasts

    SciTech Connect

    Nishigori, C.; Miyachi, Y.; Imamura, S.; Takebe, H. )

    1989-10-01

    This study was performed in order to assess the possible protective effect of superoxide dismutase (SOD) on ultraviolet (UV) damage in xeroderma pigmentosum (XP) fibroblasts. SOD activity in fibroblasts originating from seven xeroderma pigmentosum (XP) patients was significantly lower than that in normal cells (p less than 0.005). Average SOD activity in XP cells belonging to complementation group A was 3.68 +/- 0.54 (n = 7) and that in normal human cells was 5.79 +/- 1.59 (n = 6). Addition of SOD before and during UV irradiation (UVB and UVC) to the cells caused no change in the amount of unscheduled DNA synthesis and UV survival. A possible involvement of reduced SOD in XP and a possible protective effect by SOD on UV damage is discussed.

  11. Preclinical trials of human erythrocyte superoxide dismutase injection.

    PubMed

    Zhou, C; Fang, Y; Jiang, D; Yang, S; Lu, X; Sui, J; Li, P; Ren, J

    2000-07-01

    To assess the quality of human erythrocyte superoxide dismutase (SOD) injection reaching the official standard for its clinical uses. Human erythrocyte SOD injection prepared by McCord-Fridovich's method without column chromatography but with some modifications was used in preclinical trials, to observe the general pharmacology and pharmacodynamics of the product. The quality of human erythrocyte SOD injection conformed to the official standard of a biological product, which was found to be non-toxic and did not have any effects on the central and autonomic nervous systems as well as cardiovascular and respiratory systems. The efficacy of anti-inflammation and promotion of immuno-regulation especially on carrageenan and adjuvant-induced polyarthritis were shown in animals. Human erythrocyte SOD injection is appropriate for prophylactic and therapeutic uses in clinical trials.

  12. Superoxide dismutase amplifies organismal sensitivity to ionizing radiation

    SciTech Connect

    Scott, M.D.; Meshnick, S.R.; Eaton, J.W.

    1989-02-15

    Although increased superoxide dismutase (SOD) activity is often associated with enhanced resistance of cells and organisms to oxidant challenges, few direct tests of the antioxidant importance of this enzyme have been carried out. To assess the importance of SOD in defending against gamma-radiation, we employed Escherichia coli with deficient, normal, and super-normal enzyme activities. Surprisingly, the radiation sensitivity of E. coli actually increases as bacterial SOD activity increases. Elevated intracellular SOD activity sensitizes E. coli to radiation-induced mortality, whereas SOD-deficient bacteria show normal or decreased radiosensitivity. Toxic effects of activated oxygen species are involved in this phenomenon; bacterial SOD activity has no effect on radiation sensitivity under anaerobic conditions or on the lethality of other, non-oxygen-dependent, toxins such as ultraviolet radiation.

  13. Stability of ALS-related Superoxide Dismutase Protein variants

    NASA Astrophysics Data System (ADS)

    Lusebrink, Daniel; Plotkin, Steven

    Superoxide dismutase (SOD1) is a metal binding, homodimeric protein, whose misfolding is implicated in the neurodegenerative disease amyotrophic lateral sclerosis (ALS). Monomerization is believed to be a key step in the propagation of the disease. The dimer stability is often difficult to measure experimentally however, because it is entangled with protein unfolding and metal loss. We thus computationally investigate the dimer stability of mutants of SOD1 known to be associated with ALS. We report on systematic trends in dimer stability, as well as intriguing allosteric communication between mutations and the dimer interface. We study the dimer stabilities in molecular dynamics simulations and obtain the binding free energies of the dimers from pulling essays. Mutations are applied in silicoand we compare the differences of binding free energies compared to the wild type.

  14. Metalloantibiotic Mn(II)-bacitracin complex mimicking manganese superoxide dismutase

    SciTech Connect

    Piacham, Theeraphon; Isarankura-Na-Ayudhya, Chartchalerm; Nantasenamat, Chanin; Yainoy, Sakda; Ye Lei; Buelow, Leif; Prachayasittikul, Virapong . E-mail: mtvpr@mucc.mahidol.ac.th

    2006-03-24

    Superoxide dismutase (SOD) activities of various metallobacitracin complexes were evaluated using the riboflavin-methionine-nitro blue tetrazolium assay. The radical scavenging activity of various metallobacitracin complexes was shown to be higher than those of the negative controls, e.g., free transition metal ions and metal-free bacitracin. The SOD activity of the complex was found to be in the order of Mn(II) > Cu(II) > Co(II) > Ni(II). Furthermore, the effect of bacitracin and their complexation to metals on various microorganisms was assessed by antibiotic susceptibility testing. Moreover, molecular modeling and quantum chemical calculation of the metallobacitracin complex was performed to evaluate the correlation of electrostatic charge of transition metal ions on the SOD activity.

  15. Basal superoxide as a sex-specific immune constraint

    PubMed Central

    Tobler, Michael; Healey, Mo; Wilson, Mark; Olsson, Mats

    2011-01-01

    There is increasing evidence that reactive oxygen species (ROS), a group of unstable and highly reactive chemical molecules, play a key role in regulating and maintaining life-history trade-offs. Upregulation of ROS in association with immune activation is costly because it may result in an imbalance between pro- and antioxidants and, hence, oxidative damage. Previous research aimed at quantifying this cost has mostly focused on changes in the pro-/antioxidant balance subsequent to an immune response. Here, we test the hypothesis that systemic ROS may constrain immune activation. We show that systemic, pre-challenge superoxide (SO) levels are negatively related to the strength of the subsequent immune response towards the mitogen phytohaemagglutinin in male, but not female painted dragon lizards (Ctenophorus pictus). We therefore suggest that systemic SO constrains immune activation in painted dragon males. We speculate that this may be due to sex-specific selection pressures on immune investment. PMID:21632618

  16. Strain variation in bacteriocuprein superoxide dismutase from symbiotic Photobacterium leiognathi.

    PubMed

    Dunlap, P V; Steinman, H M

    1986-02-01

    Photobacterium leiognathi ATCC 25521 (the type strain and light-organ symbiont of ponyfish) is one of the few bacteria that produces a copper-zinc superoxide dismutase, termed bacteriocuprein. We enzymologically and immunologically characterized the bacteriocuprein superoxide dismutases in sonicates from the type strain and nine additional strains of P. leiognathi, each isolated from the light organ of a separate ponyfish specimen, representing seven ponyfish species. The results indicate considerable strain variation. (i) The level of bacteriocuprein enzymatic activity varied greatly among strains from different species of ponyfish. In four of the nine strains, activity was low or undetectable, while in five strains it was comparable to that in the type strain. (ii) The bacteriocuprein in one strain had a specific activity much lower than that of the type strain, and in another strain, no bacteriocuprein activity and no cross-reactive polypeptide were detectable. (iii) A new electrophoretic variant, which migrated slower than that of strains from fish captured in Thailand and Japan, was identified in strains from fish captured in the Philippine Islands. (iv) Enzymological and immunological differences were observed in bacteriocupreins of strains from male and female specimens of the same ponyfish species, for the two species in which specimens of both sexes were examined. These observations raise the possibility that specific variations in the bacteriocupreins of P. leiognathi might be characteristic of the species, geographical source, or sex of the ponyfish host. Thus, the data indicate that the possibility of strain variation should be considered when other species are screened for bacteriocupreins.

  17. Superoxide and the production of oxidative DNA damage.

    PubMed Central

    Keyer, K; Gort, A S; Imlay, J A

    1995-01-01

    The conventional model of oxidative DNA damage posits a role for superoxide (O2-) as a reductant for iron, which subsequently generates a hydroxyl radical by transferring the electron to H2O2. The hydroxyl radical then attacks DNA. Indeed, mutants of Escherichia coli that lack superoxide dismutase (SOD) were 10-fold more vulnerable to DNA oxidation by H2O2 than were wild-type cells. Even the pace of DNA damage by endogenous oxidants was great enough that the SOD mutants could not tolerate air if enzymes that repair oxidative DNA lesions were inactive. However, DNA oxidation proceeds in SOD-proficient cells without the involvement of O2-, as evidenced by the failure of SOD overproduction or anaerobiosis to suppress damage by H2O2. Furthermore, the mechanism by which excess O2- causes damage was called into question when the hypersensitivity of SOD mutants to DNA damage persisted for at least 20 min after O2- had been dispelled through the imposition of anaerobiosis. That behavior contradicted the standard model, which requires that O2- be present to rereduce cellular iron during the period of exposure to H2O2. Evidently, DNA oxidation is driven by a reductant other than O2-, which leaves the mechanism of damage promotion by O2- unsettled. One possibility is that, through its well-established ability to leach iron from iron-sulfur clusters, O2- increases the amount of free iron that is available to catalyze hydroxyl radical production. Experiments with iron transport mutants confirmed that increases in free-iron concentration have the effect of accelerating DNA oxidation. Thus, O2- may be genotoxic only in doses that exceed those found in SOD-proficient cells, and in those limited circumstances it may promote DNA damage by increasing the amount of DNA-bound iron. PMID:7592468

  18. Pharmacological induction of vascular extracellular superoxide dismutase expression in vivo.

    PubMed

    Oppermann, Marc; Balz, Vera; Adams, Volker; Dao, Vu Thao-Vi; Bas, Murat; Suvorava, Tatsiana; Kojda, Georg

    2009-07-01

    Pentaerythritol tetranitrate (PETN) treatment reduces progression of atherosclerosis and endothelial dysfunction and decreases oxidation of low-density lipoprotein (LDL) in rabbits. These effects are associated with decreased vascular superoxide production, but the underlying molecular mechanisms remain unknown. Previous studies demonstrated that endogenous nitric oxide could regulate the expression of extracellular superoxide dismutase (ecSOD) in conductance vessels in vivo. We investigated the effect of PETN and overexpression of endothelial nitric oxide synthase (eNOS(++)) on the expression and activity of ecSOD. C57BL/6 mice were randomized to receive placebo or increasing doses of PETN for 4 weeks and eNOS(++) mice with a several fold higher endothelial-specific eNOS expression were generated. The expression of ecSOD was determined in the lung and aortic tissue by real-time PCR and Western blot. The ecSOD activity was measured using inhibition of cytochrome C reduction. There was no effect of PETN treatment or eNOS overexpression on ecSOD mRNA in the lung tissue, whereas ecSOD protein expression increased from 2.5-fold to 3.6-fold (P < 0.05) by 6 mg PETN/kg body weight (BW)/day and 60 mg PETN/kg BW/day, respectively. A similar increase was found in aortic homogenates. eNOS(++) lung cytosols showed an increase of ecSOD protein level of 142 +/- 10.5% as compared with transgene-negative littermates (P < 0.05), which was abolished by N(omega)-nitro-L-arginine treatment. In each animal group, the increase of ecSOD expression was paralleled by an increase of ecSOD activity. Increased expression and activity of microvascular ecSOD are likely induced by increased bioavailability of vascular nitric oxide. Up-regulation of vascular ecSOD may contribute to the reported antioxidative and anti-atherosclerotic effects of PETN.

  19. Effects of oxidative stress on expression of extracellular superoxide dismutase, CuZn-superoxide dismutase and Mn-superoxide dismutase in human dermal fibroblasts.

    PubMed

    Strålin, P; Marklund, S L

    1994-03-01

    To determine the effect of oxidative stress on expression of extracellular superoxide dismutase (EC-SOD), CuZn-SOD and Mn-SOD, two fibroblast lines were exposed for periods of up to 4 days to a wide concentration range of oxidizing agents: xanthine oxidase plus hypoxanthine, paraquat, pyrogallol, alpha-naphthoflavone, hydroquinone, catechol, Fe2+ ions, Cu2+ ions, buthionine sulphoximine, diethylmaleate, t-butyl hydroperoxide, cumene hydroperoxide, selenite, citiolone and high oxygen partial pressure. The cell lines were cultured both under serum starvation and at a serum concentration that permitted growth. Under no condition was there any evidence of EC-SOD induction. Instead, the agents uniformly, dose-dependently and continuously reduced EC-SOD expression. We interpret the effect to be due to toxicity. Enhancement of the protection against oxidative stress by addition of CuZn-SOD, catalase and low concentrations of selenite did not influence the expression of any of the SOD isoenzymes. Removal of EC-SOD from cell surfaces by heparin also did not influence SOD expression. Mn-SOD was moderately induced by high doses of the first 11 oxidants. Apart from reduction at high toxic doses, there were no significant effects on the CuZn-SOD activity by any of the treatments. Thus EC-SOD, previously shown to be profoundly influenced by inflammatory cytokines, was not induced by its substrate or other oxidants. In a similar fashion, Mn-SOD, previously shown to be greatly induced and depressed by cytokines, was only moderately influenced by oxidants. We suggest that the regulation of these SOD isoenzymes in mammalian tissues primarily occurs in a manner co-ordinated by cytokines, rather than as a response of individual cells to oxidants.

  20. Detection of Non-Photochemical Superoxide in Coastal and Open Ocean Seawater: Particulate Versus Dissolved Sources

    NASA Astrophysics Data System (ADS)

    Roe, K. L.; Rand, T.; Hansel, C. M.; Voelker, B. M.

    2016-02-01

    Superoxide radical (O2-) could have a significant effect on marine metal redox chemistry, but little data exists on its marine concentrations. In this study, we measured superoxide steady-state concentrations in both filtered and unfiltered samples collected near the California coast and at Station ALOHA. Particle-generated superoxide, defined as the difference between unfiltered and filtered concentrations, ranged from undetectable to 0.019 nM at Station ALOHA and from undetectable to 0.052 nM in samples from the southern California Current. We also show that a transient superoxide signal is generated during filtering, an artifact that may have affected previously reported concentrations of particle-generated superoxide in the ocean. High concentrations of superoxide (range) were measured in filtered samples from ALOHA station and the California Current, raising concerns about possible sources of background signals. Further study of background signals revealed that some superoxide production occurs even in artificial seawater and very aged filtered seawater samples, and that a small additional background signal is generated as the sample travels from the container to the flow cell where it is mixed with reagent for CL analysis. However, filtered seawater samples collected from the Scripps Pier had significantly higher superoxide production rates than those measured in artificial seawater, and production rates in unfiltered samples were no higher than those in filtered samples. Therefore, production by dissolved sources was the dominant non-photochemical source of superoxide in these samples. Production rates decreased in the presence of DTPA, suggesting involvement of metal ions in superoxide production. Laboratory experiments with natural organic matter (NOM) indicate that superoxide formation occurs during oxidation of reduced moieties of NOM by oxygen.

  1. Membrane Potential Greatly Enhances Superoxide Generation by the Cytochrome bc1 Complex Reconstituted into Phospholipid Vesicles*

    PubMed Central

    Rottenberg, Hagai; Covian, Raul; Trumpower, Bernard L.

    2009-01-01

    The mitochondrial cytochrome bc1 complex (ubiquinol/cytochrome c oxidoreductase) is generally thought to generate superoxide anion that participates in cell signaling and contributes to cellular damage in aging and degenerative disease. However, the isolated, detergent-solubilized bc1 complex does not generate measurable amounts of superoxide except when inhibited by antimycin. In addition, indirect measurements of superoxide production by cells and isolated mitochondria have not clearly resolved the contribution of the bc1 complex to the generation of superoxide by mitochondria in vivo, nor did they establish the effect, if any, of membrane potential on superoxide formation by this enzyme complex. In this study we show that the yeast cytochrome bc1 complex does generate significant amounts of superoxide when reconstituted into phospholipid vesicles. The rate of superoxide generation by the reconstituted bc1 complex increased exponentially with increased magnitude of the membrane potential, a finding that is compatible with the suggestion that membrane potential inhibits electron transfer from the cytochrome bL to bH hemes, thereby promoting the formation of a ubisemiquinone radical that interacts with oxygen to generate superoxide. When the membrane potential was further increased, by the addition of nigericin or by the imposition of a diffusion potential, the rate of generation of superoxide was further accelerated and approached the rate obtained with antimycin. These findings suggest that the bc1 complex may contribute significantly to superoxide generation by mitochondria in vivo, and that the rate of superoxide generation can be controlled by modulation of the mitochondrial membrane potential. PMID:19478336

  2. Membrane potential greatly enhances superoxide generation by the cytochrome bc1 complex reconstituted into phospholipid vesicles.

    PubMed

    Rottenberg, Hagai; Covian, Raul; Trumpower, Bernard L

    2009-07-17

    The mitochondrial cytochrome bc(1) complex (ubiquinol/cytochrome c oxidoreductase) is generally thought to generate superoxide anion that participates in cell signaling and contributes to cellular damage in aging and degenerative disease. However, the isolated, detergent-solubilized bc(1) complex does not generate measurable amounts of superoxide except when inhibited by antimycin. In addition, indirect measurements of superoxide production by cells and isolated mitochondria have not clearly resolved the contribution of the bc(1) complex to the generation of superoxide by mitochondria in vivo, nor did they establish the effect, if any, of membrane potential on superoxide formation by this enzyme complex. In this study we show that the yeast cytochrome bc(1) complex does generate significant amounts of superoxide when reconstituted into phospholipid vesicles. The rate of superoxide generation by the reconstituted bc(1) complex increased exponentially with increased magnitude of the membrane potential, a finding that is compatible with the suggestion that membrane potential inhibits electron transfer from the cytochrome b(L) to b(H) hemes, thereby promoting the formation of a ubisemiquinone radical that interacts with oxygen to generate superoxide. When the membrane potential was further increased, by the addition of nigericin or by the imposition of a diffusion potential, the rate of generation of superoxide was further accelerated and approached the rate obtained with antimycin. These findings suggest that the bc(1) complex may contribute significantly to superoxide generation by mitochondria in vivo, and that the rate of superoxide generation can be controlled by modulation of the mitochondrial membrane potential.

  3. Molecular characterization of a manganese superoxide dismutase and copper/zinc superoxide dismutase from the mussel Mytilus galloprovincialis.

    PubMed

    Wang, Qing; Yuan, Zeyi; Wu, Huifeng; Liu, Feng; Zhao, Jianmin

    2013-05-01

    The full-length cDNA sequences coding respectively for a manganese superoxide dismutase (Mg-MnSOD) and copper/zinc superoxide dismutase (Mg-CuZnSOD) were cloned from Mytilus galloprovincialis. Mg-MnSOD and Mg-CuZnSOD cDNAs encoded a polypeptide of 228 and 211 amino acids, respectively. Sequence analysis indicated Mg-MnSOD was a mitochondrial MnSOD and Mg-CuZnSOD was an intracellular CuZnSOD. Multiple alignment analysis showed that both Mg-MnSOD and Mg-CuZnSOD sequences had the common features conserved in MnSODs and CuZnSODs, respectively. Phylogenetic analysis revealed that Mg-MnSOD clustered together with MnSODs from other mollusks, whereas Mg-CuZnSOD clustered with other mollusk intracellular CuZnSODs with a wider phylogenetic distance. By quantitative real-time RT-PCR (qPCR) analysis, both Mg-MnSOD and Mg-CuZnSOD transcripts were detected in all tissues examined with the highest expression level in hepatopancreas. Following bacterial challenge, the expression level of Mg-MnSOD and Mg-CuZnSOD increased first and subsequently decreased to the original level in hemocytes. In hepatopancreas, Mg-CuZnSOD mRNA was up-regulated significantly at 72 h and 96 h post challenge, while the level of Mg-MnSOD transcript had no significant change. Therefore, Mg-MnSOD and Mg-CuZnSOD expressions were inducible and they were probably involved in the immune response against bacterial challenge. These results suggest that these SODs may play important roles in the immune defense system of M. galloprovincialis and perhaps contribute to the protective effects against oxidative stress in this mussel.

  4. Pluronic-Modified Superoxide Dismutase 1 (SOD1) Attenuates Angiotensin II-Induced Increase in Intracellular Superoxide in Neurons

    PubMed Central

    Yi, Xiang; Zimmerman, Matthew C.; Yang, Ruifang; Tong, Jing; Vinogradov, Serguei; Kabanov, Alexander V.

    2010-01-01

    Overexpressing superoxide dismutase 1 (SOD1; also called Cu/ZnSOD), an intracellular superoxide (O2•−) scavenging enzyme, in central neurons inhibits angiotensin II (AngII) intra-neuronal signaling and normalizes cardiovascular dysfunction in diseases associated with enhanced AngII signaling in the brain including hypertension and heart failure. However, the blood-brain barrier (BBB) and neuronal cell membranes impose tremendous impediment for the delivery of SOD1 to central neurons, which hinders the potential therapeutic impact of SOD1 treatment on these diseases. To address this, we developed conjugates of SOD1 with poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) block copolymer (Pluronic) (SOD1-P85 and SOD1-L81), which retained significant SOD1 enzymatic activity. The modified SOD1 effectively scavenged xanthine oxidase/hypoxanthine-derived O2•−, as determined in HPLC and the measurement of 2-hydroxyethidium. Using catecholaminergic (CATH.a) neurons, we observed an increase in neuronal uptake of SOD1-Pluronic after 1, 6, or 24 hrs, compared to neurons treated with pure SOD1 or PEG-SOD1. Importantly, without inducing neuronal toxicity, SOD1-Pluronic conjugates significantly inhibited AngII-induced increases in intra-neuronal O2•−-levels. These data indicate that SOD1-Pluronic conjugates penetrate neuronal cell membranes, which results in elevated intracellular levels of functional SOD1. Pluronic conjugation may be a new delivery system for SOD1 into central neurons and therapeutically beneficial for AngII-related cardiovascular diseases. PMID:20493251

  5. Immobilization of superoxide dismutase on Pt-Pd/MWCNTs hybrid modified electrode surface for superoxide anion detection.

    PubMed

    Zhu, Xiang; Niu, Xiangheng; Zhao, Hongli; Tang, Jie; Lan, Minbo

    2015-05-15

    Monitoring of reactive oxygen species like superoxide anion (O2(∙-)) turns to be of increasing significance considering their potential damages to organism. In the present work, we fabricated a novel O2(∙-) electrochemical sensor through immobilizing superoxide dismutase (SOD) onto a Pt-Pd/MWCNTs hybrid modified electrode surface. The Pt-Pd/MWCNTs hybrid was synthesized via a facile one-step alcohol-reduction process, and well characterized by transmission electron microscopy, X-ray photoelectron spectroscopy and X-ray diffraction. The immobilization of SOD was accomplished using a simple drop-casting method, and the performance of the assembled enzyme-based sensor for O2(∙-) detection was systematically investigated by several electrochemcial techniques. Thanks to the specific biocatalysis of SOD towards O2(∙-) and the Pt-Pd/MWCNTs - promoted fast electron transfer at the fabricated interface, the developed biosensor exhibits a fast, selective and linear amperometric response upon O2(∙-) in the concentration scope of 40-1550 μM (R(2)=0.9941), with a sensitivity of 0.601 mA cm(-2) mM(-1) and a detection limit of 0.71 μM (S/N=3). In addition, the favorable biocompatibility of this electrode interface endows the prepared biosensor with excellent long-term stability (a sensitivity loss of only 3% over a period of 30 days). It is promising that the proposed sensor will be utilized as an effective tool to quantitatively monitor the dynamic changes of O2(∙-) in biological systems.

  6. Effect of superoxide and superoxide-generating systems on the prooxidant effect of iron in oil emulsion and raw turkey homogenates.

    PubMed

    Ahn, D U; Kim, S M

    1998-09-01

    Mechanisms of superoxide.O2--generating systems on the pro-oxidant effect of iron from various sources were studied. Reaction mixtures were prepared with distilled water, oil emulsion, or meat homogenates. Free ionic iron (ferrous and ferric), ferritin and hemoglobin (Hb) were used as iron sources, and KO2 and xanthine oxidase (XOD) systems were used to produce .O2-. Thiobarbituric acid reactive substances (TBARS) values and iron contents of the reaction mixtures were determined. Ferric iron and ferritin, in the presence or absence of superoxide-generating systems, had no catalytic effect on the oxidation of oil emulsion but became pro-oxidants when reducing agent (ascorbate) was present. Ferrous iron and Hb had strong catalytic effects on the oxidation of oil emulsion as shown by TBARS values. Superoxide and H2O2, generated from superoxide-generating systems, oxidized ferrous iron and ascorbate, and lowered the pro-oxidant effect of ferrous iron in oil emulsion. Addition of ferric or ferrous iron increased but Hb did not have any effect on the TBARS values of raw meat homogenates. The reaction mechanisms of superoxide and the superoxide-generating systems on the prooxidant effect of various iron sources indicated that .O2- was a strong oxidizer rather than a reducing agent, and the antioxidant effect of XOD system in oil was caused by the oxidation of ferrous iron to the ferric form by .O2- and/or H2O2.

  7. On the selectivity of superoxide dismutase mimetics and its importance in pharmacological studies

    PubMed Central

    Muscoli, Carolina; Cuzzocrea, Salvatore; Riley, Dennis P; Zweier, Jay L; Thiemermann, Christoph; Wang, Zhi-Qiang; Salvemini, Daniela

    2003-01-01

    The list of pathophysiological conditions associated with the overproduction of superoxide expands every day. Much of the knowledge compiled on the role of this radical in disease has been gathered using the native superoxide dismutase enzyme and, more recently, by the use of superoxide dismutase knockout models or transgenic models that overexpress the various isoforms of the enzyme. Although the native enzyme has shown promising anti-inflammatory properties in both preclinical and clinical studies, there were drawbacks and issues associated with its use as a therapeutic agent and pharmacological tool. Based on the concept that removal of superoxide modulates the course of inflammation, synthetic, low-molecular-weight mimetics of the superoxide dismutase enzymes that could overcome some of the limitations associated with the use of the native enzyme have been designed. In this review, we will discuss the advances made using various superoxide dismutase mimetics that led to the proposal that superoxide (and/or the product of its interaction with nitric oxide, peroxynitrite) is an important mediator of inflammation, and to the conclusion that superoxide dismutase mimetics can be utilized as therapeutic agents in diseases of various etiologies. The importance of the selectivity of such compounds in pharmacological studies will be discussed. PMID:14522841

  8. Do Superoxide Dismutase (SOD) and Catalase (CAT) protect Cells from DNA Damage Induced by Active Arsenicals?

    EPA Science Inventory

    Superoxide dismutase (SOD) catalyzes the conversion of superoxide to hydrogen peroxide, which can be converted to water and oxygen through the action of catalase. Heterozygous mice of strain B6: 129S7-SodltmlLeb/J were obtained from Jackson Laboratories and bred to produce offspr...

  9. Superoxide Dismutase Protects Cells from DNA Damage Induced by Trivalent Methylated Arsenicals

    EPA Science Inventory

    Superoxide dismutase (SOD) catalyzes the conversion of superoxide to hydrogen peroxide. Heterozygous mice of strain B6; 129S7-Sod1(tm1Leb)/J were obtained from Jackson Laboratories and bred to produce offspring that were heterozygous (+/Sod1(tm1Leb)), homozygous wild-type (+/+), ...

  10. Do Superoxide Dismutase (SOD) and Catalase (CAT) protect Cells from DNA Damage Induced by Active Arsenicals?

    EPA Science Inventory

    Superoxide dismutase (SOD) catalyzes the conversion of superoxide to hydrogen peroxide, which can be converted to water and oxygen through the action of catalase. Heterozygous mice of strain B6: 129S7-SodltmlLeb/J were obtained from Jackson Laboratories and bred to produce offspr...

  11. The French Paradox: Determining the Superoxide-Scavenging Capacity of Red Wine and Other Beverages

    ERIC Educational Resources Information Center

    Logan, Barry A.; Hammond, Matthew P.; Stormo, Benjamin M.

    2008-01-01

    Plant-derived phenolic compounds such as those found in red wine, tea, and certain fruit juices may protect against cardiovascular disease by detoxifying (scavenging) superoxide and other unstable reactive oxygen species. We present a laboratory exercise that can be used to assess the superoxide-scavenging capacity of beverages. Among the…

  12. Isolation and characterization of Cu,Zn superoxide dismutase of the shark Prionace glauca.

    PubMed

    Galtieri, A; Natoli, G; Lania, A; Calabrese, L

    1986-01-01

    A Cu,Zn superoxide dismutase was purified for the first time from an elasmobranch species (Prionace glauca) and showed the following differences with respect to other animal superoxide dismutases. The enzyme displays a low isoelectric point. The enzyme activity is unusually independent of ionic strength. The isolated enzyme has 30% of its copper in the reduced state.

  13. A biologically effective fullerene (C60) derivative with superoxide dismutase mimetic properties.

    PubMed

    Ali, Sameh S; Hardt, Joshua I; Quick, Kevin L; Kim-Han, Jeong Sook; Erlanger, Bernard F; Huang, Ting-Ting; Epstein, Charles J; Dugan, Laura L

    2004-10-15

    Superoxide, a potentially toxic by-product of cellular metabolism, may contribute to tissue injury in many types of human disease. Here we show that a tris-malonic acid derivative of the fullerene C60 molecule (C3) is capable of removing the biologically important superoxide radical with a rate constant (k(C3)) of 2 x 10(6) mol(-1) s(-1), approximately 100-fold slower than the superoxide dismutases (SOD), a family of enzymes responsible for endogenous dismutation of superoxide. This rate constant is within the range of values reported for several manganese-containing SOD mimetic compounds. The reaction between C3 and superoxide was not via stoichiometric "scavenging," as expected, but through catalytic dismutation of superoxide, indicated by lack of structural modifications to C3, regeneration of oxygen, production of hydrogen peroxide, and absence of EPR-active (paramagnetic) products, all consistent with a catalytic mechanism. A model is proposed in which electron-deficient regions on the C60 sphere work in concert with malonyl groups attached to C3 to electrostatically guide and stabilize superoxide, promoting dismutation. We also found that C3 treatment of Sod2(-/-) mice, which lack expression of mitochondrial manganese superoxide dismutase (MnSOD), increased their life span by 300%. These data, coupled with evidence that C3 localizes to mitochondria, suggest that C3 functionally replaces MnSOD, acting as a biologically effective SOD mimetic.

  14. The French Paradox: Determining the Superoxide-Scavenging Capacity of Red Wine and Other Beverages

    ERIC Educational Resources Information Center

    Logan, Barry A.; Hammond, Matthew P.; Stormo, Benjamin M.

    2008-01-01

    Plant-derived phenolic compounds such as those found in red wine, tea, and certain fruit juices may protect against cardiovascular disease by detoxifying (scavenging) superoxide and other unstable reactive oxygen species. We present a laboratory exercise that can be used to assess the superoxide-scavenging capacity of beverages. Among the…

  15. Superoxide Dismutase Protects Cells from DNA Damage Induced by Trivalent Methylated Arsenicals

    EPA Science Inventory

    Superoxide dismutase (SOD) catalyzes the conversion of superoxide to hydrogen peroxide. Heterozygous mice of strain B6; 129S7-Sod1(tm1Leb)/J were obtained from Jackson Laboratories and bred to produce offspring that were heterozygous (+/Sod1(tm1Leb)), homozygous wild-type (+/+), ...

  16. Volume reduction of nonaqueous media contaminated with a highly halogenated model compound using superoxide.

    PubMed

    Furman, Olha S; Teel, Amy L; Watts, Richard J

    2010-02-10

    Highly halogenated organic compounds, which include polychlorinated biphenyls (PCBs) and polychlorinated dibenzo-p-dioxins (PCDDs) formed during the synthesis of pentachlorophenol and chlorophenoxy herbicides, are often found as contaminants in less toxic nonaqueous media, such as waste oil, oily sludges, or biosolids. Superoxide is highly reactive with halogenated compounds when both are dissolved in nonaqueous media; however, superoxide is most economically generated in water, where it is unreactive with most organic compounds. Superoxide reactivity was investigated in organic solvent-water systems as a basis for treating halogenated contaminants in less toxic nonaqueous media. Such a process could potentially render a contaminated oil or sludge nonhazardous, providing a mechanism for waste volume reduction. Increasing amounts of water added to acetone and dimethyl sulfoxide systems decreased the activity of superoxide in the solvent, but enough activity remained for effective treatment. Superoxide was then generated in the aqueous phase of two-phase water-organic solvent systems, and significant superoxide activity was achieved in the organic media with the addition of phase transfer catalysts (PTCs) to transfer superoxide into the nonaqueous phase. The results of this research demonstrate that superoxide, which can be generated in water electrochemically or through the catalytic decomposition of peroxygens, has the potential to be transferred to oils, sludges, and other less toxic nonaqueous media to destroy highly refractory contaminants such as PCBs, PCDDs, and other halogenated contaminants.

  17. ROLE OF COPPER,ZINC-SUPEROXIDE DISMUTASE IN CATALYZING NITROTYROSINE FORMATION IN MURINE LIVER

    USDA-ARS?s Scientific Manuscript database

    The solely known function of Cu,Zn-superoxide dismutase (SOD1) is to catalyze the dismutation of superoxide anion into hydrogen peroxide. Our objective was to determine if SOD1 catalyzed murine liver protein nitration induced by acetaminophen (APAP) and lipopolysaccharide (LPS). Liver and plasma ...

  18. Pharmacological induction of vascular extracellular superoxide dismutase expression in vivo

    PubMed Central

    Oppermann, Marc; Balz, Vera; Adams, Volker; Thao-Vi Dao, Vu; Bas, Murat; Suvorava, Tatsiana; Kojda, Georg

    2009-01-01

    Pentaerythritol tetranitrate (PETN) treatment reduces progression of atherosclerosis and endothelial dysfunction and decreases oxidation of low-density lipoprotein (LDL) in rabbits. These effects are associated with decreased vascular superoxide production, but the underlying molecular mechanisms remain unknown. Previous studies demonstrated that endogenous nitric oxide could regulate the expression of extracellular superoxide dismutase (ecSOD) in conductance vessels in vivo. We investigated the effect of PETN and overexpression of endothelial nitric oxide synthase (eNOS++) on the expression and activity of ecSOD. C57BL/6 mice were randomized to receive placebo or increasing doses of PETN for 4 weeks and eNOS++ mice with a several fold higher endothelial-specific eNOS expression were generated. The expression of ecSOD was determined in the lung and aortic tissue by real-time PCR and Western blot. The ecSOD activity was measured using inhibition of cytochrome C reduction. There was no effect of PETN treatment or eNOS overexpression on ecSOD mRNA in the lung tissue, whereas ecSOD protein expression increased from 2.5-fold to 3.6-fold (P < 0.05) by 6 mg PETN/kg body weight (BW)/day and 60 mg PETN/kg BW/day, respectively. A similar increase was found in aortic homogenates. eNOS++ lung cytosols showed an increase of ecSOD protein level of 142 ± 10.5% as compared with transgene-negative littermates (P < 0.05), which was abolished by Nω-nitro-L-arginine treatment. In each animal group, the increase of ecSOD expression was paralleled by an increase of ecSOD activity. Increased expression and activity of microvascular ecSOD are likely induced by increased bioavailability of vascular nitric oxide. Up-regulation of vascular ecSOD may contribute to the reported antioxidative and anti-atherosclerotic effects of PETN. PMID:19320775

  19. Sensitive spectrophotometric method for the determination of superoxide dismutase activity in tissue extracts

    SciTech Connect

    Paoletti, F.; Aldinucci, D.; Mocali, A.; Caparrini, A.

    1986-05-01

    Superoxide dismutase (EC 1.15.1.1) has been assayed by a spectrophotometric method based on the inhibition of a superoxide-driven NADH oxidation. The assay consists of a purely chemical reaction sequence which involves EDTA. Mn(II), mercaptoethanol, and molecular oxygen, requiring neither auxiliary enzymes nor sophisticated equipment. The method is very flexible and rapid and is applicable with high sensitivity to the determination of both pure and crude superoxide dismutase preparations. The decrease of the rate of NADH oxidation is a function of enzyme concentration, and saturation levels are attainable. Fifty percent inhibition, corresponding to one unit of the enzyme, is produced by approximately 15 ng of pure superoxide dismutase. Experiments on rat liver cytosol have shown the specificity of the method for superoxide dismutase. Moreover, common cellular components do not interfere with the measurement, except for hemoglobin when present at relatively high concentrations. The assay is performed at physiological pH and is unaffected by catalase.

  20. Light-induced production of singlet oxygen and superoxide by the fungal toxin, cercosporin.

    PubMed

    Daub, M E; Hangarter, R P

    1983-11-01

    Cercosporin, a toxin produced by members of the fungal genus Cercospora, is a photosensitizing compound which rapidly kills plant cells in the light. We have found that cercosporin, when activated by light in the presence of oxygen, is able to generate both singlet oxygen and superoxide ions. Cercosporin, when illuminated in the presence of O(2), reacted with cholesterol to form the 5alpha-hydroperoxide of cholesterol which is only produced by reaction with singlet oxygen. Cercosporin, in the presence of light, O(2), and a reducing substrate, was also able to reduce p-nitro blue tetrazolium chloride, a compound readily reduced by superoxide. Superoxide dismutase, a scavenger of superoxide, inhibited this reaction. Production of both singlet oxygen and superoxide by cercosporin must be considered when studying the possible mechanisms of resistance to cercosporin.

  1. Inhibitory effects of cardols and related compounds on superoxide anion generation by xanthine oxidase.

    PubMed

    Masuoka, Noriyoshi; Nihei, Ken-ichi; Maeta, Ayami; Yamagiwa, Yoshiro; Kubo, Isao

    2015-01-01

    5-Pentadecatrienylresorcinol, isolated from cashew nuts and commonly known as cardol (C₁₅:₃), prevented the generation of superoxide radicals catalysed by xanthine oxidase without the inhibition of uric acid formation. The inhibition kinetics did not follow the Michelis-Menten equation, but instead followed the Hill equation. Cardol (C₁₀:₀) also inhibited superoxide anion generation, but resorcinol and cardol (C₅:₀) did not inhibit superoxide anion generation. The related compounds 3,5-dihydroxyphenyl alkanoates and alkyl 2,4-dihydroxybenzoates, had more than a C9 chain, cooperatively inhibited but alkyl 3,5-dihydroxybenzoates, regardless of their alkyl chain length, did not inhibit the superoxide anion generation. These results suggested that specific inhibitors for superoxide anion generation catalysed by xanthine oxidase consisted of an electron-rich resorcinol group and an alkyl chain having longer than C9 chain.

  2. Relationship of superoxide production to cytoplasmic free calcium in human monocytes.

    PubMed Central

    Scully, S P; Segel, G B; Lichtman, M A

    1986-01-01

    Calcium has been proposed as an intracellular second messenger for activation of secretion, phagocytosis, and the oxidative burst of neutrophils. We have examined the role of calcium in human monocyte activation. Concanavalin A (Con A)-stimulated monocytes displayed an increment in cytoplasmic ionized calcium at 31 +/- 6 s and the onset of superoxide production at 61 +/- 9 s. The increase in cytoplasmic calcium invariably preceded the onset of superoxide production. If the external calcium concentration was reduced to less than 28 nM by the addition of 10 mM EGTA, superoxide production was not diminished at 5 min; however, superoxide production decreased thereafter. The Con A-evoked increment in cytoplasmic ionized calcium was blunted upon the addition of EGTA and decreased further with time. Both the production of superoxide and the Con A-evoked increment in cytoplasmic ionized calcium displayed a 50% inhibition after 15 min of calcium depletion and were completely inhibited after 60 min. Total cell calcium fell from 0.7 to 0.5 fmol/cell, and the basal level of ionized calcium fell from 83 to 30 nM after 60 min. Histidine, a strong chelator of divalent cations other than calcium and magnesium, had no effect on monocyte superoxide production or on ionized calcium concentrations, indicating that EGTA inhibition was due to cell calcium depletion. In calcium-depleted cells, Con A did not evoke superoxide production until calcium was restored to the incubation medium. The restoration of calcium to Con A-treated, calcium-depleted monocytes permitted a rapid rise in the cytoplasmic ionized calcium, and the production of superoxide within 9 s. These data suggest that an increase in ionized cytoplasmic calcium is necessary for the activation of monocyte superoxide production by Con A. The rise in ionized calcium in response to Con A results, in part, from an internal redistribution of calcium, which is sufficient to permit superoxide generation. PMID:3007579

  3. Increased production of superoxide anion contributes to dysfunction of the arteriovenous fistula

    PubMed Central

    Tsapenko, Mykola V.; d'Uscio, Livius V.; Grande, Joseph P.; Croatt, Anthony J.; Hernandez, Melissa C.; Ackerman, Allan W.; Katusic, Zvonimir S.

    2012-01-01

    Vascular access dysfunction causes morbidity in hemodialysis patients. This study examined the generation and pathobiological significance of superoxide anion in a rat femoral arteriovenous fistula (AVF). One week after AVF creation, there was increased production of superoxide anion accompanied by decreased total superoxide dismutase (SOD) and Cu/Zn SOD activities and induction of the redox-sensitive gene heme oxygenase-1. Immunohistochemical studies of nitrotyrosine formation demonstrated that peroxynitrite, a product of superoxide anion and nitric oxide, was present in increased amounts in endothelial and smooth muscle cells in the AVF. Because uncoupled NOS isoforms generate superoxide anion, and NOS coupling requires tetrahydrobiopterin (BH4) as a cofactor, we assessed NOS uncoupling by determining the ratio of BH4 to dihydrobiopterin (BH2); the BH4-to-BH2 ratio was markedly attenuated in the AVF. Because Src is a vasculopathic signaling species upstream and downstream of superoxide anion, such expression was evaluated; expression of Src and phosphorylated Src was both markedly increased in the AVF. Expression of NADPH oxidase (NOX) 1, NOX2, NOX4, cyclooxygenase (COX) 1, COX2, p47phox, and p67phox was all unchanged, as assessed by Western analyses, thereby suggesting that these proteins may not be involved in increased production of superoxide anion. Finally, administration of tempol, a superoxide anion scavenger, decreased neointima formation in the juxta-anastomotic venous segment and improved AVF blood flow. We conclude that the AVF exhibits increased superoxide anion generation that may reflect the combined effects of decreased scavenging by SOD and increased generation by uncoupled NOS, and that enhanced superoxide anion production promotes juxta-anastomotic stenosis and impairs AVF function. PMID:22993073

  4. Bosentan, a mixed endothelin receptor antagonist, inhibits superoxide anion-induced pain and inflammation in mice.

    PubMed

    Serafim, Karla G G; Navarro, Suelen A; Zarpelon, Ana C; Pinho-Ribeiro, Felipe A; Fattori, Victor; Cunha, Thiago M; Alves-Filho, Jose C; Cunha, Fernando Q; Casagrande, Rubia; Verri, Waldiceu A

    2015-11-01

    Bosentan is a mixed endothelin receptor antagonist widely used to treat patients with pulmonary arterial hypertension, and the emerging literature suggests bosentan as a potent anti-inflammatory drug. Superoxide anion is produced in large amounts during inflammation, stimulates cytokine production, and thus contributes to inflammation and pain. However, it remains to be determined whether endothelin contributes to the inflammatory response triggered by the superoxide anion. The present study investigated the effects of bosentan in a mouse model of inflammation and pain induced by potassium superoxide, a superoxide anion donor. Male Swiss mice were treated with bosentan (10-100 mg/kg) by oral gavage, 1 h before potassium superoxide injection, and the inflammatory response was evaluated locally and at spinal cord (L4-L6) levels. Bosentan (100 mg/kg) inhibited superoxide anion-induced mechanical and thermal hyperalgesia, overt pain-like behavior (abdominal writhings, paw flinching, and licking), paw edema, myeloperoxidase activity (neutrophil marker) in the paw skin, and leukocyte recruitment in the peritoneal cavity. Bosentan also inhibited superoxide anion-induced interleukin-1 beta (IL-1β) and tumor necrosis factor alpha (TNF-α) production, while it enhanced IL-10 production in the paw skin and spinal cord. Bosentan inhibited the reduction of antioxidant capacity (reduced glutathione, ferric reducing antioxidant power, and ABTS radical scavenging ability) induced by the superoxide anion. Finally, we demonstrated that intraplantar injection of potassium superoxide induces the mRNA expression of prepro-endothelin-1 in the paw skin and spinal cord. In conclusion, our results demonstrated that superoxide anion-induced inflammation, pain, cytokine production, and oxidative stress depend on endothelin; therefore, these responses are amenable to bosentan treatment.

  5. The superoxide anion donor, potassium superoxide, induces pain and inflammation in mice through production of reactive oxygen species and cyclooxygenase-2.

    PubMed

    Maioli, N A; Zarpelon, A C; Mizokami, S S; Calixto-Campos, C; Guazelli, C F S; Hohmann, M S N; Pinho-Ribeiro, F A; Carvalho, T T; Manchope, M F; Ferraz, C R; Casagrande, R; Verri, W A

    2015-04-01

    It is currently accepted that superoxide anion (O2•-) is an important mediator in pain and inflammation. The role of superoxide anion in pain and inflammation has been mainly determined indirectly by modulating its production and inactivation. Direct evidence using potassium superoxide (KO2), a superoxide anion donor, demonstrated that it induced thermal hyperalgesia, as assessed by the Hargreaves method. However, it remains to be determined whether KO2 is capable of inducing other inflammatory and nociceptive responses attributed to superoxide anion. Therefore, in the present study, we investigated the nociceptive and inflammatory effects of KO2. The KO2-induced inflammatory responses evaluated in mice were: mechanical hyperalgesia (electronic version of von Frey filaments), thermal hyperalgesia (hot plate), edema (caliper rule), myeloperoxidase activity (colorimetric assay), overt pain-like behaviors (flinches, time spent licking and writhing score), leukocyte recruitment, oxidative stress, and cyclooxygenase-2 mRNA expression (quantitative PCR). Administration of KO2 induced mechanical hyperalgesia, thermal hyperalgesia, paw edema, leukocyte recruitment, the writhing response, paw flinching, and paw licking in a dose-dependent manner. KO2 also induced time-dependent cyclooxygenase-2 mRNA expression in the paw skin. The nociceptive, inflammatory, and oxidative stress components of KO2-induced responses were responsive to morphine (analgesic opioid), quercetin (antioxidant flavonoid), and/or celecoxib (anti-inflammatory cyclooxygenase-2 inhibitor) treatment. In conclusion, the well-established superoxide anion donor KO2 is a valuable tool for studying the mechanisms and pharmacological susceptibilities of superoxide anion-triggered nociceptive and inflammatory responses ranging from mechanical and thermal hyperalgesia to overt pain-like behaviors, edema, and leukocyte recruitment.

  6. Role of superoxide in the germination of Bacillus anthracis endospores.

    PubMed

    Baillie, Les; Hibbs, Stephen; Tsai, Pei; Cao, Guan-Liang; Rosen, Gerald M

    2005-04-01

    The spore forming Gram-positive bacterium Bacillus anthracis, the causative agent of anthrax, has achieved notoriety due to its use as a bioterror agent. In the environment, B. anthracis exists as a dormant endospore. Germination of endospores during their internalization within the myeloid phagocyte, and the ability of those endospores to survive exposure to antibacterial killing mechanisms such as superoxide (O(2)*-, is a key initial event in the infective process. We report herein that endospores exposed to fluxes of O(2)*- typically found in stimulated phagocytes had no effect on viability. Further endospores of the Sterne strain of B. anthracis were found to scavenge O(2)*-, which may enhance the ability of the bacterium to survive within the hostile environment of the phagolysosome. Most intriguing was the observation that endospore germination was stimulated by a flux of O(2)*- as low as 1 microM/min. Data presented herein suggest that B. anthracis may co-opt O(2)*- which is produced by stimulated myeloid phagocytes and is an essential element of host immunity, as a necessary step in productive infection of the host.

  7. Amelioration of hepatic reperfusion injury by superoxide dismutase and catalase

    SciTech Connect

    Clemens, M.G.; Burke, F.; Chaudry, I.H.

    1986-03-05

    Oxygen-derived free radicals have been implicated in reperfusion injury in various tissues. The present study determined if enzymatic scavenging of free radicals could improve recovery of hepatic function following ischemia. Livers from fasted rats were perfused with Krebs-HCO/sub 3/ buffer with substrates for gluconeogenesis for 30 min (control) followed by 60 min warm ischemia and 90 min reperfusion. At the beginning and end of ischemia the liver was flushed with buffered Ringer's with superoxide dismutase + catalase (150,000 U/L each)(SOD) or without additions (Untreated). Bile flow and glucose release were monitored during control and reperfusion periods and tissue sampled at the end of the experiment to determine tissue water and electrolytes. Bile flow and gluconeogenesis were markedly depressed after ischemia in both groups. At the end of 90 min reperfusion bile flow in Untreated and SOD were 23 +/- 6 and 46 +/- 8 ..mu..l/15 min (20% and 41% of control respectively, p < .01). Gluconeogenesis recovered to 83 +/- 4% of control in Untreated vs 103 +/- 6% with SOD (p < .05). Tissue water and electrolytes were not different. These results suggest that generation of oxygen-derived free radicals contributes to functional deficits in the liver following ischemia and that these defects can be attenuated by enzymatic scavenging.

  8. NADPH Oxidase-Dependent Superoxide Production in Plant Reproductive Tissues.

    PubMed

    Jiménez-Quesada, María J; Traverso, José Á; Alché, Juan de Dios

    2016-01-01

    In the life cycle of a flowering plant, the male gametophyte (pollen grain) produced in the anther reaches the stigmatic surface and initiates the pollen-pistil interaction, an important step in plant reproduction, which ultimately leads to the delivery of two sperm cells to the female gametophyte (embryo sac) inside the ovule. The pollen tube undergoes a strictly apical expansion characterized by a high growth rate, whose targeting should be tightly regulated. A continuous exchange of signals therefore takes place between the haploid pollen and diploid tissue of the pistil until fertilization. In compatible interactions, theses processes result in double fertilization to form a zygote (2n) and the triploid endosperm. Among the large number of signaling mechanisms involved, the redox network appears to be particularly important. Respiratory burst oxidase homologs (Rbohs) are superoxide-producing enzymes involved in a broad range of processes in plant physiology. In this study, we review the latest findings on understanding Rboh activity in sexual plant reproduction, with a particular focus on the male gametophyte from the anther development stages to the crowning point of fertilization. Rboh isoforms have been identified in both the male and female gametophyte and have proven to be tightly regulated. Their role at crucial points such as proper growth of pollen tube, self-incompatibility response and eventual fertilization is discussed.

  9. Copper, ceruloplasmin, superoxide dismutase and iron parameters in Parkinson's disease.

    PubMed

    Tórsdóttir, G; Kristinsson, J; Sveinbjörnsdóttir, S; Snaedal, J; Jóhannesson, T

    1999-11-01

    In a previous study we found copper dyshomeostasis in patients with Alzheimer's disease. In this study, levels of copper in plasma, of ceruloplasmin in serum and ceruloplasmin oxidative activity as well as superoxide dismutase (SOD) activity in erythrocytes were determined in 40 patients with Parkinson's disease and their healthy age- and gender-matched controls. Copper concentrations did not differ significantly in the two groups, whereas both ceruloplasmin concentrations and ceruloplasmin oxidative activity were significantly lower in the patients, also relative to ceruloplasmin mass. SOD activity was not significantly different in the two groups but decreased significantly with the duration of disease. The same was found for ceruloplasmin oxidative activity. Ceruloplasmin oxidative activity and SOD activity did not decrease with age. Levels of serum iron, serum ferritin and total iron binding capacity were determined in about 30 of the patients and an equal number of controls and were not found to differ. Transferrin levels were significantly lower in the patients than in their controls but, conversely, the transferrin saturation was significantly higher in the patients. The results indicate that patients with Alzheimer's disease and Parkinson's disease have defective ceruloplasmin and SOD activities in common and that these defects are not necessarily associated with major disturbances in iron homeostasis.

  10. Superoxide scavenging activity of pirfenidone-iron complex

    SciTech Connect

    Mitani, Yoshihiro; Sato, Keizo Muramoto, Yosuke; Karakawa, Tomohiro; Kitamado, Masataka; Iwanaga, Tatsuya; Nabeshima, Tetsuji; Maruyama, Kumiko; Nakagawa, Kazuko; Ishida, Kazuhiko; Sasamoto, Kazumi

    2008-07-18

    Pirfenidone (PFD) is focused on a new anti-fibrotic drug, which can minimize lung fibrosis etc. We evaluated the superoxide (O{sub 2}{sup {center_dot}}{sup -}) scavenging activities of PFD and the PFD-iron complex by electron spin resonance (ESR) spectroscopy, luminol-dependent chemiluminescence assay, and cytochrome c reduction assay. Firstly, we confirmed that the PFD-iron complex was formed by mixing iron chloride with threefold molar PFD, and the complex was stable in distillated water and ethanol. Secondary, the PFD-iron complex reduced the amount of O{sub 2}{sup {center_dot}}{sup -} produced by xanthine oxidase/hypoxanthine without inhibiting the enzyme activity. Thirdly, it also reduced the amount of O{sub 2}{sup {center_dot}}{sup -} released from phorbor ester-stimulated human neutrophils. PFD alone showed few such effects. These results suggest the possibility that the O{sub 2}{sup {center_dot}}{sup -} scavenging effect of the PFD-iron complex contributes to the anti-fibrotic action of PFD used for treating idiopathic pulmonary fibrosis.

  11. Status of Superoxide Dismutase in Transfusion Dependent Thalassaemia

    PubMed Central

    Rujito, Lantip; Mulatsih, Sri; Sofro, Abdul Salam M.

    2015-01-01

    Background: Thalassemia is a collection of genetic impairments in beta and alpha genes causing various states of anemia. Severe types of the disease need lifelong transfusions, leading to oxidant-antioxidant disturbance due to massive iron deposits. Aims: The aim of this study was to assess the antioxidant enzyme Superoxide Dismutase (SOD) and ferritin levels of thalassemia major patients in a peripheral health facility. Materials and Methods: Two hundred and nine probands were recruited and performed laboratory experiments for SOD and Ferritin levels. Chelation administration and clinical score were taken from interviewing the family and from medical report data. Results: The study showed that SOD intensity was lower (162.41 u/ml) compared to the normal cutoff point (P = 0.001), while the mean of Ferritin levels was ten times over the normal value (4226,67 ng/dl). Observations also reported that chelation medicine was not administrated properly. Conclusions: The data indicates that thalassemic patients have oxidant-antioxidant uproar due to oxidative stress. Monitored chelating administration, selective antioxidant, and a well-balanced diet may prevent oxidative injury. PMID:26110130

  12. Superoxide Dismutase 1 Nanozyme for Treatment of Eye Inflammation

    PubMed Central

    Kost, Olga A.; Beznos, Olga V.; Davydova, Nina G.; Manickam, Devika S.; Nikolskaya, Irina I.; Guller, Anna E.; Binevski, Petr V.; Chesnokova, Natalia B.; Shekhter, Anatoly B.; Klyachko, Natalia L.; Kabanov, Alexander V.

    2016-01-01

    Use of antioxidants to mitigate oxidative stress during ocular inflammatory diseases has shown therapeutic potential. This work examines a nanoscale therapeutic modality for the eye on the base of antioxidant enzyme, superoxide dismutase 1 (SOD1), termed “nanozyme.” The nanozyme is produced by electrostatic coupling of the SOD1 with a cationic block copolymer, poly(L-lysine)-poly(ethyleneglycol), followed by covalent cross-linking of the complexes with 3,3′-dithiobis(sulfosuccinimidylpropionate) sodium salt. The ability of SOD1 nanozyme as well as the native SOD1 to reduce inflammatory processes in the eye was examined in vivo in rabbits with immunogenic uveitis. Results suggested that topical instillations of both enzyme forms demonstrated anti-inflammatory activity; however, the nanozyme was much more effective compared to the free enzyme in decreasing uveitis manifestations. In particular, we noted statistically significant differences in such inflammatory signs in the eye as the intensities of corneal and iris edema, hyperemia of conjunctiva, lens opacity, fibrin clots, and the protein content in aqueous humor. Clinical findings were confirmed by histological data. Thus, SOD1-containing nanozyme is potentially useful therapeutic agent for the treatment of ocular inflammatory disorders. PMID:26697135

  13. Superoxide Production by Digitonin-Stimulated Guinea Pig Granulocytes

    PubMed Central

    Cohen, Harvey J.; Chovaniec, Margaret E.

    1978-01-01

    N-ethylmaleimide, divalent cations, ethylene glycol bis (β aminoethyl ether) N,N,N′,N′,-tetraacetate, 2-deoxyglucose, cyanide, and dinitrophenol were examined for their effect on the ability of guinea pig granulocytes to generate superoxide (O2−) when stimulated by digitonin. N-ethylmaleimide (1 mM) inhibits only when added before complete activation of the O2− generating system, and at lower concentrations (0.05-0.2 mM) slows the activation process. Ca++ is required for maximum O2− generation, and Mg++ decreases the amount of Ca++ required. Ethylene glycol bis (β aminoethyl ether) N,N,N′,N′,-tetraacetate (10 mM) inhibits only if added before complete activation. Incubation of cells in 2-DOG causes a time- and concentration-dependent inhibition of O2− generation. It also increases the time required for activation of this system. Cyanide and dinitrophenol increase the rate of O2− production. However, when these compounds are added to cells whose O2− production is partially inhibited by incubation in 2-deoxyglucose, complete inhibition results. If cyanide or dinitrophenol is added after activation of 2-deoxyglucose-treated cells, no further inhibition occurs. On the basis of the above results, we conclude that the activation of the O2− generating system is N-ethylmaleimide sensitive, Ca++ dependent, and energy requiring, but that the activity of the enzyme system in the cell is not. PMID:207722

  14. Superoxide dismutase activity of Cu-bound prion protein

    NASA Astrophysics Data System (ADS)

    Hodak, Miroslav; Lu, Wenchang; Bernholc, Jerry

    2009-03-01

    Misfolding of the prion protein, PrP, has been linked to a group of neurodegenerative diseases, including the mad cow disease in cattle and the Creutzfeldt-Jakob disease in humans. The normal function of PrP is still unknown, but it was found that the PrP can efficiently bind Cu(II) ions. Early experiments suggested that Cu-PrP complex possesses significant superoxide dismutase (SOD) activity, but later experiments failed to confirm it and at present this issue remains unresolved. Using a recently developed hybrid DFT/DFT method, which combines Kohn-Sham DFT for the solute and its first solvation shells with orbital-free DFT for the remainder of the solvent, we have investigated SOD activity of PrP. The PrP is capable of incorporating Cu(II) ions in several binding modes and our calculations find that each mode has a different SOD activity. The highest activity found is comparable to those of well-known SOD proteins, suggesting that the conflicting experimental results may be due to different bindings of Cu(II) in those experiments.

  15. Superoxide dismutase 3 attenuates experimental Th2-driven allergic conjunctivitis.

    PubMed

    Lee, Hyun Jung; Kim, Bo-Mi; Shin, Soojung; Kim, Tae-Yoon; Chung, So-Hyang

    2017-03-01

    Allergic conjunctivitis is an inflammatory eye disease mediated by Th2 type immune response. The role of extracellular superoxide dismutase 3 (SOD3) in immune response and allergic conjunctival inflammation was examined in a murine model for experimental allergic conjunctivitis (EAC). Allergic conjunctivitis was induced in mice by allergen challenge with ovalbumin in alum via the conjunctival sac. SOD3 was topically applied and allergy indicators were compared. Clinical signs associated with conjunctivitis, such as OVA-specific IgE production, IgG1/G2a ratio and eosinophil infiltration, were drastically reduced in mice treated with SOD3. They also had less dendritic cells and CD4(+) T cells in conjunctiva than controls. Attenuated allergic inflammation was accredited to reduced Th2 type cytokine responses and increased Treg cytokine in draining lymph node. The characteristics of EAC were attributed to the absence of SOD3. Our findings suggest that SOD3 might be considered as a potential target for Th2-driven allergic conjunctival inflammation. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Biodegradable polycaprolactone (PCL) nanosphere encapsulating superoxide dismutase and catalase enzymes.

    PubMed

    Singh, Sushant; Singh, Abhay Narayan; Verma, Anil; Dubey, Vikash Kumar

    2013-12-01

    Biodegradable polycaprolactone (PCL) nanosphere encapsulating superoxide dismutase (SOD) and catalase (CAT) were successfully synthesized using double emulsion (w/o/w) solvent evaporation technique. Characterization of the nanosphere using dynamic light scattering, field emission scanning electron microscope, and Fourier transform infrared spectroscopy revealed a spherical-shaped nanosphere in a size range of 812 ± 64 nm with moderate protein encapsulation efficiency of 55.42 ± 3.7 % and high in vitro protein release. Human skin HaCat cells were used for analyzing antioxidative properties of SOD- and CAT-encapsulated PCL nanospheres. Oxidative stress condition in HaCat cells was optimized with exposure to hydrogen peroxide (H2O2; 1 mM) as external stress factor and verified through reactive oxygen species (ROS) analysis using H2DCFDA dye. PCL nanosphere encapsulating SOD and CAT together indicated better antioxidative defense against H2O2-induced oxidative stress in human skin HaCat cells in comparison to PCL encapsulating either SOD or CAT alone as well as against direct supplement of SOD and CAT protein solution. Increase in HaCat cells SOD and CAT activities after treatment hints toward uptake of PCL nanosphere into the human skin HaCat cells. The result signifies the role of PCL-encapsulating SOD and CAT nanosphere in alleviating oxidative stress.

  17. Reversible activation of the neutrophil superoxide generating system by hexachlorocyclohexane: correlation with effects on a subcellular superoxide-generating fraction.

    PubMed

    English, D; Schell, M; Siakotos, A; Gabig, T G

    1986-07-01

    gamma-Hexachlorocyclohexane was found to exert profound effects on the phosphatidylinositol cycle, cytosolic calcium level, and the respiratory burst of human neutrophils. Exposure of neutrophils prelabelled with 32P to 4 X 10(-4) M gamma-hexachlorocyclohexane almost tripled radioactivity in phosphatidic acid and correspondingly decreased radioactivity in phosphatidylinositol 4,5 bisphosphate. Under similar conditions, gamma-hexachlorocyclohexane evoked the generation of superoxide at a rate of over 11 nmol/min/10(6) cells and more than doubled cytosolic-free calcium concentration as monitored by Quin-2 fluorescence. Because intermediates of the phosphatidylinositol cycle, via increases in available calcium levels or activated protein kinase C, are considered potential second messengers for activation of the NADPH-dependent O-2-generating system, we compared neutrophil responses to gamma-hexachlorocyclohexane with responses to phorbol myristate acetate, an activator of protein kinase C with well known effects on neutrophils. Like phorbol myristate acetate, gamma-hexachlorocyclohexane induced neutrophil degranulation but was not an effective chemotactic stimulus. The ability of gamma-hexachlorocyclohexane to induce a pattern of oxidative activation in neutrophil cytoplasts similar to that in intact cells indicated that concurrent degranulation was not required for sustained O-2 generation in response to this agent. When neutrophils or neutrophil cytoplasts exposed to gamma-hexachlorocyclohexane were centrifuged and resuspended in stimulus-free medium, O-2 generation ceased entirely but could be reinitiated by addition of the same stimulus. This finding was in contrast to the continued O-2 production by phorbol myristate acetate-stimulated neutrophils similarly washed and resuspended in stimulus-free medium. Unlike subcellular fractions of phorbol myristate acetate-stimulated neutrophils, corresponding fractions prepared from gamma

  18. Erythrocyte copper chaperone for superoxide dismutase and superoxide dismutase as biomarkers for hepatic copper concentrations in Labrador retrievers.

    PubMed

    Dirksen, K; Roelen, Y S; van Wolferen, M E; Kruitwagen, H S; Penning, L C; Burgener, I A; Spee, B; Fieten, H

    2016-12-01

    Hereditary hepatic copper accumulation in Labrador retrievers leads to hepatitis with fibrosis and eventually cirrhosis. The development of a non-invasive blood-based biomarker for copper status in dogs could be helpful in identifying dogs at risk and to monitor copper concentrations during treatment. In this study, two cellular copper metabolism proteins, Cu/Zn superoxide dismutase (SOD1) and its chaperone (copper chaperone for SOD1, CCS) were measured in erythrocytes and tested for association with hepatic copper concentrations in 15 Labrador retrievers with normal or increased hepatic copper concentrations. Antibodies against CCS and SOD1 were applicable for use in canine specimens. This was demonstrated by the loss of immune-reactive bands for CCS and SOD1 in siRNA treated canine bile duct epithelial cells. Erythrocyte CCS and CCS/SOD1 ratios were decreased 2.37 (P <0.001) and 3.29 (P <0.001) fold in the high copper group compared to the normal copper group. Erythrocyte CCS and CCS/SOD1 ratio are potential new biomarkers for hepatic copper concentrations in Labrador retrievers and could facilitate early diagnosis and treatment monitoring for copper-associated hepatitis in dogs. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Chaperonin 20 might be an iron chaperone for superoxide dismutase in activating iron superoxide dismutase (FeSOD)

    PubMed Central

    Kuo, Wen-Yu; Huang, Chien-Hsun; Jinn, Tsung-Luo

    2013-01-01

    Activation of Cu/Zn superoxide dismutases (CuZnSODs) is aided by Cu incorporation and disulfide isomerization by Cu chaperone of SOD (CCS). As well, an Fe-S cluster scaffold protein, ISU, might alter the incorporation of Fe or Mn into yeast MnSOD (ySOD2), thus leading to active or inactive ySOD2. However, metallochaperones involved in the activation of FeSODs are unknown. Recently, we found that a chloroplastic chaperonin cofactor, CPN20, could mediate FeSOD activity. To investigate whether Fe incorporation in FeSOD is affected by CPN20, we used inductively coupled plasma mass spectrometry to analyze the ability of CPN20 to bind Fe. CPN20 could bind Fe, and the Fe binding to FeSOD was increased with CPN20 incubation. Thus, CPN20 might be an Fe chaperone for FeSOD activation, a role independent of its well-known co-chaperonin activity. PMID:23299425

  20. Biologically Relevant Mechanism For Catalytic Removal of Superoxide by Simple Manganese Compounds

    SciTech Connect

    Barnese K.; Cabelli D.; Gralla, E.B.; Valentine, J.S.

    2012-05-01

    Nonenzymatic manganese was first shown to provide protection against superoxide toxicity in vivo in 1981, but the chemical mechanism responsible for this protection subsequently became controversial due to conflicting reports concerning the ability of Mn to catalyze superoxide disproportionation in vitro. In a recent communication, we reported that low concentrations of a simple Mn phosphate salt under physiologically relevant conditions will indeed catalyze superoxide disproportionation in vitro. We report now that two of the four Mn complexes that are expected to be most abundant in vivo, Mn phosphate and Mn carbonate, can catalyze superoxide disproportionation at physiologically relevant concentrations and pH, whereas Mn pyrophosphate and citrate complexes cannot. Additionally, the chemical mechanisms of these reactions have been studied in detail, and the rates of reactions of the catalytic removal of superoxide by Mn phosphate and carbonate have been modeled. Physiologically relevant concentrations of these compounds were found to be sufficient to mimic an effective concentration of enzymatic superoxide dismutase found in vivo. This mechanism provides a likely explanation as to how Mn combats superoxide stress in cellular systems.

  1. Manganese Superoxide Dismutase Protects against 6-Hydroxydopamine Injury in Mouse Brains*

    PubMed Central

    Callio, Jason; Oury, Tim D.; Chu, Charleen T.

    2007-01-01

    Dopaminergic neurons of the substantia nigra are susceptible to toxin-based insults. Intrastriatal injection of 6-hydroxydopamine results in selective toxicity to these neurons. A mechanistic role for reactive oxygen species is supported by observations that antioxidants confer protection from 6-hydroxydopamine. Although cell culture studies have suggested extracellular or nonmitochondrial mechanisms in 6-hydroxydopamine toxicity, the compartmentalization of oxidative injury mechanisms is incompletely defined in vivo. Transgenic mice overexpressing mitochondrial manganese superoxide dismutase or extracellular superoxide dismutase received unilateral intrastriatal injections of 6-hydroxydopamine. Mice that overexpress manganese superoxide dismutase showed significantly smaller striatal lesions than littermate controls. There were no differences in nonspecific striatal injury associated with contralateral vehicle injection. Manganese superoxide dismutase overexpression also protected against loss of neuronal cell bodies in the substantia nigra. In contrast, mice overexpressing extracellular superoxide dismutase showed no protection from 6-hydroxydopamine toxicity in either brain region. Protection of the nigrostriatal system by overexpression of manganese super-oxide dismutase supports a role for mitochondrially derived superoxide in 6-hydroxydopamine toxicity. Mitochondrial oxidative stress appears to be a common mechanism among diverse models of Parkinson disease, whether involving toxins, mutated genes, or cybrid cells containing patient mitochondria. Antioxidant therapies that target this subcellular compartment may prove promising. PMID:15755737

  2. Effects of asbestos and silica on superoxide anion production in the guinea pig alveolar macrophage

    SciTech Connect

    Roney, P.L.

    1988-01-01

    This study examined the effect of asbestos and silica on the activation pathway of the guinea pig alveolar macrophage. Activation of macrophages by physiological agents results in stimulation of phospholipase C causing phosphatidyl inositol turnover and intracellular calcium mobilization. Phosphatidyl inositol turnover produces diacylglycerol which activates protein kinase C causing superoxide anion production. Chrysotile stimulated alveolar macrophages to produce superoxide anion. This stimulation proceeded via phospholipase C, since chrysotile stimulated phosphatidyl inositol turnover and intracellular calcium mobilization. The possible involvement of a coupling protein was evaluated by pretreating cells with pertussis toxin. Potential binding sites for chrysotile stimulation were examined using a series of nine lectins. Chrysotile-stimulated superoxide anion production was blocked by pretreatment with lectins which bound to mannose, fucose, or N-acetylgalactosamine. In addition, incubation with the N-acetylglucosamine, but not by lectins which bound to mannose, fucose, or N-acetylgalactosamine. In addition, incubation with the N-acetylglucosamine polymer, chitin, inhibited chrysotile-stimulated superoxide anion production, suggesting that chrysotile stimulated superoxide anion production by binding to N-acetylglucosamine residues. On the other hand, silica did not stimulate superoxide anion production. The effect of silica on agonist stimulation of this pathway was examined using two stimulants of superoxide anion production, N-formyl-nle-leu-phe (FNLP, which stimulates through phospholipase C) and phorbol-12,13-dibutyrate (which directly activates protein kinase C).

  3. AMPK dysregulation promotes diabetes-related reduction of superoxide and mitochondrial function.

    PubMed

    Dugan, Laura L; You, Young-Hyun; Ali, Sameh S; Diamond-Stanic, Maggie; Miyamoto, Satoshi; DeCleves, Anne-Emilie; Andreyev, Aleksander; Quach, Tammy; Ly, San; Shekhtman, Grigory; Nguyen, William; Chepetan, Andre; Le, Thuy P; Wang, Lin; Xu, Ming; Paik, Kacie P; Fogo, Agnes; Viollet, Benoit; Murphy, Anne; Brosius, Frank; Naviaux, Robert K; Sharma, Kumar

    2013-11-01

    Diabetic microvascular complications have been considered to be mediated by a glucose-driven increase in mitochondrial superoxide anion production. Here, we report that superoxide production was reduced in the kidneys of a steptozotocin-induced mouse model of type 1 diabetes, as assessed by in vivo real-time transcutaneous fluorescence, confocal microscopy, and electron paramagnetic resonance analysis. Reduction of mitochondrial biogenesis and phosphorylation of pyruvate dehydrogenase (PDH) were observed in kidneys from diabetic mice. These observations were consistent with an overall reduction of mitochondrial glucose oxidation. Activity of AMPK, the major energy-sensing enzyme, was reduced in kidneys from both diabetic mice and humans. Mitochondrial biogenesis, PDH activity, and mitochondrial complex activity were rescued by treatment with the AMPK activator 5-aminoimidazole-4-carboxamide-1-β-D-ribofuranoside (AICAR). AICAR treatment induced superoxide production and was linked with glomerular matrix and albuminuria reduction in the diabetic kidney. Furthermore, diabetic heterozygous superoxide dismutase 2 (Sod2(+/-)) mice had no evidence of increased renal disease, and Ampka2(-/-) mice had increased albuminuria that was not reduced with AICAR treatment. Reduction of mitochondrial superoxide production with rotenone was sufficient to reduce AMPK phosphorylation in mouse kidneys. Taken together, these results demonstrate that diabetic kidneys have reduced superoxide and mitochondrial biogenesis and activation of AMPK enhances superoxide production and mitochondrial function while reducing disease activity.

  4. Mn(II) oxidation by an ascomycete fungus is linked to superoxide production during asexual reproduction.

    PubMed

    Hansel, Colleen M; Zeiner, Carolyn A; Santelli, Cara M; Webb, Samuel M

    2012-07-31

    Manganese (Mn) oxides are among the most reactive minerals within the environment, where they control the bioavailability of carbon, nutrients, and numerous metals. Although the ability of microorganisms to oxidize Mn(II) to Mn(III/IV) oxides is scattered throughout the bacterial and fungal domains of life, the mechanism and physiological basis for Mn(II) oxidation remains an enigma. Here, we use a combination of compound-specific chemical assays, microspectroscopy, and electron microscopy to show that a common Ascomycete filamentous fungus, Stilbella aciculosa, oxidizes Mn(II) to Mn oxides by producing extracellular superoxide during cell differentiation. The reactive Mn oxide phase birnessite and the reactive oxygen species superoxide and hydrogen peroxide are colocalized at the base of asexual reproductive structures. Mn oxide formation is not observed in the presence of superoxide scavengers (e.g., Cu) and inhibitors of NADPH oxidases (e.g., diphenylene iodonium chloride), enzymes responsible for superoxide production and cell differentiation in fungi. Considering the recent identification of Mn(II) oxidation by NADH oxidase-based superoxide production by a common marine bacterium (Roseobacter sp.), these results introduce a surprising homology between some prokaryotic and eukaryotic organisms in the mechanisms responsible for Mn(II) oxidation, where oxidation appears to be a side reaction of extracellular superoxide production. Given the versatility of superoxide as a redox reactant and the widespread ability of fungi to produce superoxide, this microbial extracellular superoxide production may play a central role in the cycling and bioavailability of metals (e.g., Hg, Fe, Mn) and carbon in natural systems.

  5. Mn(II) oxidation by an ascomycete fungus is linked to superoxide production during asexual reproduction

    SciTech Connect

    Hansel, C. M.; Zeiner, C. A.; Santelli, C. M.; Webb, S. M.

    2012-07-16

    Manganese (Mn) oxides are among the most reactive minerals within the environment, where they control the bioavailability of carbon, nutrients, and numerous metals. Although the ability of microorganisms to oxidize Mn(II) to Mn(III/IV) oxides is scattered throughout the bacterial and fungal domains of life, the mechanism and physiological basis for Mn(II) oxidation remains an enigma. Here, we use a combination of compound-specific chemical assays, microspectroscopy, and electron microscopy to show that a common Ascomycete filamentous fungus, Stilbella aciculosa, oxidizes Mn(II) to Mn oxides by producing extracellular superoxide during cell differentiation. The reactive Mn oxide phase birnessite and the reactive oxygen species superoxide and hydrogen peroxide are colocalized at the base of asexual reproductive structures. Mn oxide formation is not observed in the presence of superoxide scavengers (e.g., Cu) and inhibitors of NADPH oxidases (e.g., diphenylene iodonium chloride), enzymes responsible for superoxide production and cell differentiation in fungi. Considering the recent identification of Mn(II) oxidation by NADH oxidase-based superoxide production by a common marine bacterium (Roseobacter sp.), these results introduce a surprising homology between some prokaryotic and eukaryotic organisms in the mechanisms responsible for Mn(II) oxidation, where oxidation appears to be a side reaction of extracellular superoxide production. Finally, given the versatility of superoxide as a redox reactant and the widespread ability of fungi to produce superoxide, this microbial extracellular superoxide production may play a central role in the cycling and bioavailability of metals (e.g., Hg, Fe, Mn) and carbon in natural systems.

  6. Ceruloplasmin copper induces oxidant damage by a redox process utilizing cell-derived superoxide as reductant

    NASA Technical Reports Server (NTRS)

    Mukhopadhyay, C. K.; Fox, P. L.

    1998-01-01

    Oxidative damage by transition metals bound to proteins may be an important pathogenic mechanism. Ceruloplasmin (Cp) is a Cu-containing plasma protein thought to be involved in oxidative modification of lipoproteins. We have previously shown that Cp increased cell-mediated low-density lipoprotein (LDL) oxidation by a process requiring cell-derived superoxide, but the underlying chemical mechanism(s) is (are) unknown. We now show that superoxide reduction of Cp Cu is a critical reaction in cellular LDL oxidation. By bathocuproine disulfonate (BCS) binding and by superoxide utilization, we showed that exogenous superoxide reduces a single Cp Cu atom, the same Cu required for LDL oxidation. The Cu atom remained bound to Cp during the redox cycle. Three avenues of evidence showed that vascular cells reduce Cp Cu by a superoxide-dependent process. The 2-fold higher rate of Cp Cu reduction by smooth muscle cells (SMC) compared to endothelial cells (EC) was consistent with their relative rates of superoxide release. Furthermore, Cp Cu reduction by cells was blocked by Cu,Zn superoxide dismutase (SOD1). Finally, the level of superoxide produced by EC and SMC was sufficient to cause the amount of Cu reduction observed. An important role of Cp Cu reduction in LDL oxidation was suggested by results showing that SOD1 inhibited Cp Cu reduction and LDL oxidation by SMC with equal potency, while tumor necrosis factor-alpha stimulated both processes. In summary, these results show that superoxide is a critical cellular reductant of divalent transition metals involved in oxidation, and that protein-bound Cu is a substrate for this reaction. The role of these mechanisms in oxidative processes in vivo has yet to be defined.

  7. Ceruloplasmin copper induces oxidant damage by a redox process utilizing cell-derived superoxide as reductant

    NASA Technical Reports Server (NTRS)

    Mukhopadhyay, C. K.; Fox, P. L.

    1998-01-01

    Oxidative damage by transition metals bound to proteins may be an important pathogenic mechanism. Ceruloplasmin (Cp) is a Cu-containing plasma protein thought to be involved in oxidative modification of lipoproteins. We have previously shown that Cp increased cell-mediated low-density lipoprotein (LDL) oxidation by a process requiring cell-derived superoxide, but the underlying chemical mechanism(s) is (are) unknown. We now show that superoxide reduction of Cp Cu is a critical reaction in cellular LDL oxidation. By bathocuproine disulfonate (BCS) binding and by superoxide utilization, we showed that exogenous superoxide reduces a single Cp Cu atom, the same Cu required for LDL oxidation. The Cu atom remained bound to Cp during the redox cycle. Three avenues of evidence showed that vascular cells reduce Cp Cu by a superoxide-dependent process. The 2-fold higher rate of Cp Cu reduction by smooth muscle cells (SMC) compared to endothelial cells (EC) was consistent with their relative rates of superoxide release. Furthermore, Cp Cu reduction by cells was blocked by Cu,Zn superoxide dismutase (SOD1). Finally, the level of superoxide produced by EC and SMC was sufficient to cause the amount of Cu reduction observed. An important role of Cp Cu reduction in LDL oxidation was suggested by results showing that SOD1 inhibited Cp Cu reduction and LDL oxidation by SMC with equal potency, while tumor necrosis factor-alpha stimulated both processes. In summary, these results show that superoxide is a critical cellular reductant of divalent transition metals involved in oxidation, and that protein-bound Cu is a substrate for this reaction. The role of these mechanisms in oxidative processes in vivo has yet to be defined.

  8. How the location of superoxide generation influences the β-cell response to nitric oxide.

    PubMed

    Broniowska, Katarzyna A; Oleson, Bryndon J; McGraw, Jennifer; Naatz, Aaron; Mathews, Clayton E; Corbett, John A

    2015-03-20

    Cytokines impair the function and decrease the viability of insulin-producing β-cells by a pathway that requires the expression of inducible nitric oxide synthase (iNOS) and generation of high levels of nitric oxide. In addition to nitric oxide, excessive formation of reactive oxygen species, such as superoxide and hydrogen peroxide, has been shown to cause β-cell damage. Although the reaction of nitric oxide with superoxide results in the formation of peroxynitrite, we have shown that β-cells do not have the capacity to produce this powerful oxidant in response to cytokines. When β-cells are forced to generate peroxynitrite using nitric oxide donors and superoxide-generating redox cycling agents, superoxide scavenges nitric oxide and prevents the inhibitory and destructive actions of nitric oxide on mitochondrial oxidative metabolism and β-cell viability. In this study, we show that the β-cell response to nitric oxide is regulated by the location of superoxide generation. Nitric oxide freely diffuses through cell membranes, and it reacts with superoxide produced within cells and in the extracellular space, generating peroxynitrite. However, only when it is produced within cells does superoxide attenuate nitric oxide-induced mitochondrial dysfunction, gene expression, and toxicity. These findings suggest that the location of radical generation and the site of radical reactions are key determinants in the functional response of β-cells to reactive oxygen species and reactive nitrogen species. Although nitric oxide is freely diffusible, its biological function can be controlled by the local generation of superoxide, such that when this reaction occurs within β-cells, superoxide protects β-cells by scavenging nitric oxide.

  9. Antioxidant Capacity and Superoxide Dismutase Activity in Adrenoleukodystrophy.

    PubMed

    Turk, Bela R; Theisen, Benjamin E; Nemeth, Christina L; Marx, Joel S; Shi, Xiaohai; Rosen, Melissa; Jones, Richard O; Moser, Ann B; Watkins, Paul A; Raymond, Gerald V; Tiffany, Carol; Fatemi, Ali

    2017-05-01

    X-linked adrenoleukodystrophy (ALD) may switch phenotype to the fatal cerebral form (ie, cerebral ALD [cALD]), the cause of which is unknown. Determining differences in antioxidant capacity and superoxide dismutase (SOD) levels between phenotypes may allow for the generation of a clinical biomarker for predicting the onset of cALD, as well as initiating a more timely lifesaving therapy. To identify variations in the levels of antioxidant capacity and SOD activity between ALD phenotypes in patients with cALD or adrenomyeloneuropathy (AMN), heterozygote female carriers, and healthy controls and, in addition, correlate antioxidant levels with clinical outcome scores to determine a possible predictive value. Samples of monocytes and blood plasma were prospectively collected from healthy controls, heterozygote female carriers, and patients with AMN or cALD. We are counting each patient as 1 sample in our study. Because adrenoleukodystrophy is an X-linked disease, the affected group populations of cALD and AMN are all male. The heterozygote carriers are all female. The samples were assayed for total antioxidant capacity and SOD activity. The data were collected in an academic hospital setting. Eligibility criteria included patients who received a diagnosis of ALD and heterozygote female carriers, both of which groups were compared with age-matched controls. The prospective samples (n = 30) were collected between January 2015 to January 2016, and existing samples were collected from tissue storage banks at the Kennedy Krieger Institute (n = 30). The analyses were performed during the first 3 months of 2016. Commercially available total antioxidant capacity and SOD assays were performed on samples of monocytes and blood plasma and correlated with magnetic resonance imaging severity score. A reduction in antioxidant capacity was shown between the healthy controls (0.225 mmol trolox equivalent) and heterozygote carriers (0.181 mmol trolox equivalent), and significant

  10. A Manganese Superoxide Dismutase (SOD2)-Mediated Adaptive Response

    PubMed Central

    Grdina, David J.; Murley, Jeffrey S.; Miller, Richard C.; Mauceri, Helena J.; Sutton, Harold G.; Thirman, Michael J.; Li, Jian Jian; Woloschak, Gayle E.; Weichselbaum, Ralph R.

    2013-01-01

    Very low doses of ionizing radiation, 5 to 100 mGy, can induce adaptive responses characterized by elevation in cell survival and reduction in micronuclei formation. Utilizing these end points, RKO human colon carcinoma and transformed mouse embryo fibroblasts (MEF), wild-type or knockout cells missing TNF receptors 1 and 2 (TNFR1−R2−), and C57BL/6 and TNFR1−R2− knockout mice, we demonstrate that intact TNF signaling is required for induction of elevated manganese superoxide dismutase (SOD2) activity (P < 0.001) and the subsequent expression of these SOD2-mediated adaptive responses when cells are challenged at a later time with 2 Gy. In contrast, amifostine’s free thiol form WR1065 can directly activate NF-κB giving rise to elevated SOD2 activity 24 h later and induce an adaptive response in both MEF wild-type and TNF signaling defective TNFR1−R2− cells. Transfection of cells with SOD2 siRNA completely abolishes both the elevation in SOD2 activity and expression of the adaptive responses. These results were confirmed in vivo using a micronucleus assay in splenocytes derived from C57BL/6 and TNFR1−R2− knockout mice that were exposed to 100 mGy or 400 mg/kg amifostine 24 h prior to exposure to a 2 Gy whole-body dose. A dose of 100 mGy also conferred enhanced protection to C57BL/6 mice exposed 24 h later to 100 mg/kg of N-Ethyl-N-nitrosourea (ENU). While very low radiation doses require an intact TNF signaling process to induce a SOD2-mediated adaptive response, amifostine can induce a similar adaptive response in both TNF receptor competent and knockout cells, respectively. PMID:23237540

  11. Mineral status and superoxide dismutase enzyme activity in Alzheimer's disease.

    PubMed

    Rodrigues, Gilmara Péres; Cozzolino, Silvia Maria Franciscato; Marreiro, Dilina do Nascimento; Caldas, Daniele Rodrigues Carvalho; da Silva, Kelcylene Gomes; de Sousa Almondes, Kaluce Gonçalves; Neto, José Machado Moita; Pimentel, José Alexandre Coelho; de Carvalho, Cecília Maria Resende Gonçalves; Nogueira, Nadir do Nascimento

    2017-12-01

    The study evaluated the dietary intake of zinc and copper, as measured by plasma and erythrocyte concentrations, the Cu/Zn ratio and measure the erythrocyte superoxide dismutase enzyme (eSOD) activity and the relationship between these markers and the degree of dementia in elderly individuals with and without Alzheimer's Disease (AD). A total of 93 elderly people aged 60-94 years were divided into two groups: with AD (n=44) and without AD (n=49). The NINCDS-ADRDA criteria were used for diagnosing AD, and dementia staging was determined using the Clinical Dementia Rating (CDR) scale. The dietary intake of Zn and Cu was obtained from a standard 3-day food record. Plasma and erythrocyte concentrations of the minerals were determined by flame atomic absorption spectrophotometry and by measuring eSOD activity in an automatic biochemical analyzer. The results showed dietary intake of Zn and Cu above the reference values with no differences observed between the two groups (p>0.05). Plasma and erythrocyte normocupremia as well as alteration in the Zn pool, with its reduced plasma concentrations and high in the erythrocytes, were observed in both groups (p>0.05). The plasma Cu/Zn ratio were not significantly different in patients with and without AD (p>0.05). The eSOD activity was high in both patient groups (p>0.05). However, among elderly patients with AD there was a positive correlation between this marker and dementia severity. According to our study results, we conclude that plasma and erythrocyte concentrations of Cu and Zn, as well as Cu/Zn ratio among elderly individuals is not related to Alzheimer's Disease. However, antioxidant activity of eSOD is associated with dementia severity. Copyright © 2017 Elsevier GmbH. All rights reserved.

  12. Superoxide dismutase and oxygen toxicity defenses in the genus Neisseria.

    PubMed Central

    Archibald, F S; Duong, M N

    1986-01-01

    Among aerotolerant cells, Neisseria gonorrhoeae is very unusual because despite its obligately aerobic lifestyle and frequent isolation from purulent exudates containing polymorphonuclear leukocytes vigorously evolving O2- and H2O2, it contains no superoxide dismutase (SOD). Strains (14) of N. gonorrhoeae were compared with each other and with strains of Neisseria meningitidis, Neisseria mucosa, and Neisseria subflava under identical growth conditions for their contents of the oxy-protective enzymes catalase, peroxidase, and SOD, as well as respiratory chain proteins and activity. The absence of SOD from N. gonorrhoeae strains was demonstrated under a variety of oxygen-stress conditions. The neisserial species showed very different SOD, catalase, and peroxidase profiles. These profiles correlated well with the tolerance of the species to various intra- and extracellular oxygen insults. The high tolerance of N. gonorrhoeae for extracellular O2- and H2O2 appeared to be due to very high constitutive levels of peroxidase and catalase activity combined with a cell envelope impervious to O2-. Nevertheless, N. gonorrhoeae 19424 was much more sensitive to an intracellular flux of O2- than were the other (SOD-containing) neisserial species. The responses of N. gonorrhoeae and N. meningitidis respiratory and oxy-protective enzymes to growth under high and low oxygen tensions were followed, and a novel response, the apparent repression of the respiratory chain intermediates, respiration, and SOD, peroxidase, and catalase activity, was observed. The gonococcal catalase was partially purified and characterized. The results suggest that the very active terminal oxidase, low pO2 natural habitat, O2-stable catalase, and possibly the high glutathione content of the organism explain its aerobic survival in the absence of SOD. PMID:3943903

  13. Characterization of the Single Superoxide Dismutase of Staphylococcus xylosus

    PubMed Central

    Barrière, Charlotte; Brückner, Reinhold; Talon, Régine

    2001-01-01

    Staphylococcus xylosus is a facultative anaerobic bacterium used as a starter culture for fermented meat products. In an attempt to analyze the antioxidant capacities of this organism, the superoxide dismutase (SOD) was characterized. S. xylosus contains a single cytoplasmic SOD, which was not inhibited by H2O2. The SOD activity in crude extracts was completely lost upon metal depletion, but it could be recovered by manganese and very weakly by iron. It is therefore suggested that the S. xylosus SOD is a manganese-preferring enzyme. The corresponding gene, sod, was isolated from a genomic library of S. xylosus DNA and complemented the growth defect of an Escherichia coli SOD-deficient mutant. As deduced from the nucleotide sequence, sod encodes a protein of 199 amino acids with a molecular mass of 22.5 kDa. Two transcriptional start sites 25 and 120 bp upstream of the sod start codon were identified. A terminator-like structure downstream of the gene suggested a monocistronic sod mRNA. Regulation of sod expression was studied using fusions of the sod promoters to a genomic promoterless β-galactosidase gene. The sod expression was not affected by manganese and increased slightly with paraquat. It was induced during stationary phase in a complex medium but not in a chemically defined medium. To investigate the physiological role of SOD, a mutant devoid of SOD activity was constructed. Growth experiments showed that sod is not essential for aerobic growth in complex medium. However, in chemically defined medium without leucine, isoleucine, and valine, the sod mutant hardly grew, in contrast to the wild-type strain. In addition, the mutant was sensitive to hyperbaric oxygen and to paraquat. Therefore, sod plays an important role in the protection of S. xylosus from oxidative stress. PMID:11526011

  14. [The superoxide theory of pathogenesis and therapy of immune disorders].

    PubMed

    Lebedev, V V

    2004-01-01

    On the basis of the understanding that there are common development mechanisms for the inflammatory and immune reactions it was established that the activity of the oxidant-antioxidant system (OAS) correlates not only with a severity of the inflammatory reaction but also with a degree of immune disorders. Such disorders were studied in patients with endogenous uveitis and with cancer of the esophagus or uterine cervix, i.e. those nosological forms, which are normally accompanied by OAS decompensation, which comprised a lower activity of primary antioxidants (superoxides of dismutase, catalase, lactoferrin, ceruloplasmin etc.) in patients with pronounced immune disorders. Moreover, a lower content of secondary antioxidants, like vitamin A, ascorbic acid and tocopherol, was registered in the blood of patients with immune disorders. The suppression of the antioxidant system was concomitant with an essentially increased level of lipid peroxidation in all patients. Besides, it was noted that there were intensifying signs of immune disorders primarily observed during irradiation chemotherapy. In this context, a clear-cut correlation was established, in monitoring the body immune status, between degrees of free-radical formation and lipid peroxidation, on the one hand, and an activity of detoxication-system antioxidants, on the other hand,. The OAS correction by direct or indirect-action antioxidants normally improves the clinical course of immune impairments. The indirect-action antioxidants, e.g. synthetic regulatory peptide "Imunofan", induce the increasing activity of primary endogenous antioxidants. An activation of the detoxication antioxidant system, brings about, in such cases, a lower content of inflammation mediators, a recovery of cell-immunity indices and lower parameters of body auto-sensitization. Finally, the antioxidant system in patients with chronic inflammatory or oncological disorders, when recovered, ensures the correction of cell immunity and cuts

  15. The Interaction of Mitochondrial Iron with Manganese Superoxide Dismutase*

    PubMed Central

    Naranuntarat, Amornrat; Jensen, Laran T.; Pazicni, Samuel; Penner-Hahn, James E.; Culotta, Valeria C.

    2009-01-01

    Superoxide dismutase 2 (SOD2) is one of the rare mitochondrial enzymes evolved to use manganese as a cofactor over the more abundant element iron. Although mitochondrial iron does not normally bind SOD2, iron will misincorporate into Saccharomyces cerevisiae Sod2p when cells are starved for manganese or when mitochondrial iron homeostasis is disrupted by mutations in yeast grx5, ssq1, and mtm1. We report here that such changes in mitochondrial manganese and iron similarly affect cofactor selection in a heterologously expressed Escherichia coli Mn-SOD, but not a highly homologous Fe-SOD. By x-ray absorption near edge structure and extended x-ray absorption fine structure analyses of isolated mitochondria, we find that misincorporation of iron into yeast Sod2p does not correlate with significant changes in the average oxidation state or coordination chemistry of bulk mitochondrial iron. Instead, small changes in mitochondrial iron are likely to promote iron-SOD2 interactions. Iron binds Sod2p in yeast mutants blocking late stages of iron-sulfur cluster biogenesis (grx5, ssq1, and atm1), but not in mutants defective in the upstream Isu proteins that serve as scaffolds for iron-sulfur biosynthesis. In fact, we observed a requirement for the Isu proteins in iron inactivation of yeast Sod2p. Sod2p activity was restored in mtm1 and grx5 mutants by depleting cells of Isu proteins or using a dominant negative Isu1p predicted to stabilize iron binding to Isu1p. In all cases where disruptions in iron homeostasis inactivated Sod2p, we observed an increase in mitochondrial Isu proteins. These studies indicate that the Isu proteins and the iron-sulfur pathway can donate iron to Sod2p. PMID:19561359

  16. Intermolecular transmission of superoxide dismutase 1 misfolding in living cells

    PubMed Central

    Grad, Leslie I.; Guest, Will C.; Yanai, Anat; Pokrishevsky, Edward; O'Neill, Megan A.; Gibbs, Ebrima; Semenchenko, Valentyna; Yousefi, Masoud; Wishart, David S.; Plotkin, Steven S.; Cashman, Neil R.

    2011-01-01

    Human wild-type superoxide dismutase-1 (wtSOD1) is known to coaggregate with mutant SOD1 in familial amyotrophic lateral sclerosis (FALS), in double transgenic models of FALS, and in cell culture systems, but the structural determinants of this process are unclear. Here we molecularly dissect the effects of intracellular and cell-free obligately misfolded SOD1 mutant proteins on natively structured wild-type SOD1. Expression of the enzymatically inactive, natural familial ALS SOD1 mutations G127X and G85R in human mesenchymal and neural cell lines induces misfolding of wild-type natively structured SOD1, as indicated by: acquisition of immunoreactivity with SOD1 misfolding-specific monoclonal antibodies; markedly enhanced protease sensitivity suggestive of structural loosening; and nonnative disulfide-linked oligomer and multimer formation. Expression of G127X and G85R in mouse cell lines did not induce misfolding of murine wtSOD1, and a species restriction element for human wtSOD1 conversion was mapped to a region of sequence divergence in loop II and β-strand 3 of the SOD1 β-barrel (residues 24–36), then further refined surprisingly to a single tryptophan residue at codon 32 (W32) in human SOD1. Time course experiments enabled by W32 restriction revealed that G127X and misfolded wtSOD1 can induce misfolding of cell-endogenous wtSOD1. Finally, aggregated recombinant G127X is capable of inducing misfolding and protease sensitivity of recombinant human wtSOD1 in a cell-free system containing reducing and chelating agents; cell-free wtSOD1 conversion was also restricted by W32. These observations demonstrate that misfolded SOD1 can induce misfolding of natively structured wtSOD1 in a physiological intracellular milieu, consistent with a direct protein–protein interaction. PMID:21930926

  17. EPR Detection of Cellular and Mitochondrial Superoxide Using Cyclic Hydroxylamines

    PubMed Central

    Dikalov, Sergey I.; Kirilyuk, Igor A.; Voinov, Maxim; Grigor’ev, Igor A.

    2014-01-01

    Superoxide (O2•) has been implicated in the pathogenesis of many human diseases but detection of the O2• radicals in biological systems is limited due to inefficiency of O2• spin trapping and lack of site-specific information. In this work we studied production of extracellular, intracellular and mitochondrial O2• in neutrophils, cultured endothelial cells and isolated mitochondria using new set of cationic, anionic and neutral hydroxylamine spin probes with various lipophilicity and cell permeability. Cyclic hydroxylamines rapidly react with O2• producing stable nitroxides and allowed site-specific O2• detection in intracellular, extracellular and mitochondrial compartments. Negatively charged 1-hydroxy-4-phosphono-oxy-2,2,6,6-tetramethylpiperidine (PP-H) and positively charged 1-hydroxy-2,2,6,6-tetramethylpiperidin-4-yl-trimethylammonium (CAT1-H) detected only extramitochondrial O2•. Inhibition of EPR signal by SOD2 overexpression showed that mitochondria targeted mitoTEMPO-H detected intramitochondrial O2• both in isolated mitochondria and intact cells. Both 1-hydroxy-3-carboxy-2,2,5,5-tetramethylpyrrolidine (CP-H) and 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine (CM-H) detected increase in cytoplasm O2• stimulated by PMA but only CM-H and mitoTEMPO-H showed increase in rotenone-induced mitochondrial O2•. These data show that new set of hydroxylamine spin probes provide unique information about site-specific production of O2• radical in extracellular or intracellular compartments, cytoplasm or mitochondria. PMID:21128732

  18. Portacaval shunting causes differential mitochondrial superoxide production in brain regions.

    PubMed

    Kosenko, Elena A; Tikhonova, Lyudmila A; Alilova, Gubidat A; Montoliu, Carmina; Barreto, George E; Aliev, Gjumrakch; Kaminsky, Yury G

    2017-09-27

    The portacaval shunting (PCS) prevents portal hypertension and recurrent bleeding of esophageal varices. On the other hand, it can induce chronic hyperammonemia and is considered to be the best model of mild hepatic encephalopathy (HE). Pathogenic mechanisms of HE and dysfunction of the brain in hyperammonemia are not fully elucidated, but it was originally suggested that the pathogenetic defect causes destruction of antioxidant defense which leads to an increase in the production of reactive oxygen species (ROS) and the occurrence of oxidative stress. In order to gain insight into the pathogenic mechanisms of HE in the brain tissue, we investigated the effects of PCS in rats on free radicals production and activity levels of antioxidant and prooxidant enzymes in mitochondria isolated from different brain areas. We found that O2(·-) production, activities of Mn-superoxide dismutase (Mn-SOD), glutathione peroxidase (GPx), glutathione reductase (GR), glutathione transferase (GT), nitric oxide synthase (NOS), and levels of carbonylated proteins differed between the four brain regions both in the amount and response to PCS. In PCS rats, Mn-SOD activity in the cerebellum was significantly decreased, and remained unchanged in the neocortex, hippocampus and striatum compared with that in sham-operated animals. Among the four brain regions in control rats, the levels of the carbonyl groups in mitochondrial proteins were maximal in the cerebellum. 4 weeks after PCS, the content of carbonylated proteins were higher only in mitochondria of this brain region. Under control conditions, O2(·-) production by submitochondrial particles in the cerebellum was significantly higher than in other brain regions, but was significantly increased in each brain region from PCS animals. Indeed, the production of O2(·-) by submitochondrial particles correlated with mitochondrial ammonia levels in the four brain regions of control and PCS-animals. These findings are the first to suggest

  19. EPR detection of cellular and mitochondrial superoxide using cyclic hydroxylamines.

    PubMed

    Dikalov, Sergey I; Kirilyuk, Igor A; Voinov, Maxim; Grigor'ev, Igor A

    2011-04-01

    Superoxide (O₂ⁱ⁻) has been implicated in the pathogenesis of many human diseases, but detection of the O(2)(•-) radicals in biological systems is limited due to inefficiency of O₂ⁱ⁻ spin trapping and lack of site-specific information. This work studied production of extracellular, intracellular and mitochondrial O₂ⁱ⁻ in neutrophils, cultured endothelial cells and isolated mitochondria using a new set of cationic, anionic and neutral hydroxylamine spin probes with various lipophilicity and cell permeability. Cyclic hydroxylamines rapidly react with O₂ⁱ⁻, producing stable nitroxides and allowing site-specific cO₂ⁱ⁻ detection in intracellular, extracellular and mitochondrial compartments. Negatively charged 1-hydroxy-4-phosphono-oxy-2,2,6,6-tetramethylpiperidine (PP-H) and positively charged 1-hydroxy-2,2,6,6-tetramethylpiperidin-4-yl-trimethylammonium (CAT1-H) detected only extramitochondrial O₂ⁱ⁻. Inhibition of EPR signal by SOD2 over-expression showed that mitochondria targeted mitoTEMPO-H detected intramitochondrial O₂ⁱ⁻ both in isolated mitochondria and intact cells. Both 1-hydroxy-3-carboxy-2,2,5,5-tetramethylpyrrolidine (CP-H) and 1-hydroxy-3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine (CM-H) detected an increase in cytoplasm O₂ⁱ⁻ stimulated by PMA, but only CM-H and mitoTEMPO-H showed an increase in rotenone-induced mitochondrial O₂ⁱ⁻. These data show that a new set of hydroxylamine spin probes provide unique information about site-specific production of the O₂ⁱ⁻ radical in extracellular or intracellular compartments, cytoplasm or mitochondria.

  20. Naloxone inhibits superoxide but not enzyme release by human neutrophils

    SciTech Connect

    Simpkins, C.; Alailima, S.; Tate, E.

    1986-03-01

    The release of toxic oxygen metabolites and enzymes by phagocytic cells is thought to play a role in the multisystemic tissue injury of sepsis. Naloxone protects septic animals. We have found that at concentrations administered to animals (10/sup -7/ to 10/sup -4/M), naloxone inhibited (p < .001) the release of superoxide (O/sub 2//sup -/) by human neutrophils (HN), stimulated with N-formyl methionyl leucyl phenylalanine (FMLP). Naloxone had no effect on cell viability. Maximum inhibition was 65% of the total O/sub 2//sup -/ released (13.1 nMoles/8 min/320,000 cells). FMLP-stimulated release of beta-glucoronidase or lysozyme was not altered by naloxone. Naloxone had no effect on the binding of /sup 3/H FMLP to HN. Using /sup 3/H naloxone and various concentrations of unlabeled naloxone higher affinity (K/sub D/ = 12nM) and lower affinity (K/sub D/ = 4.7 x 10/sup -5/) binding sites were detected. The K/sub D/ of the low affinity site corresponded to the ED/sub 50/ for naloxone inhibition of O/sub 2//sup -/ (1 x 10/sup -5/M). Binding to this low affinity site was decreased by (+) naloxone, beta-endorphin and N acetyl beta-endorphin, but not by leu-enkephalin, thyrotropin releasing factor, prostaglandin D/sub 2/ or E/sub 2/. Conclusions: (1) naloxone inhibits FMLP-stimulated O/sub 2/ but not enzyme release, (2) this inhibition is not due to alteration of FMLP receptor binding, (3) naloxone may act via a low affinity binding site which is ligand specific, and (4) a higher affinity receptor is present on HN.

  1. Superoxide dismutase 2 as a marker to differentiate tuberculous pleural effusions from malignant pleural effusions.

    PubMed

    Wang, Maoshui; Zhang, Zhiqiang; Wang, Xinfeng

    2014-01-01

    Our previous study demonstrated that superoxide dismutase levels were higher in tuberculous pleural effusions than in malignant pleural effusions, but that this difference could not be used to discriminate between the two. The objective of the present study was to investigate the levels of superoxide dismutase 2 in pleural effusions and to evaluate the diagnostic significance of pleural effusion superoxide dismutase 2. Superoxide dismutase 2 concentrations were determined in pleural effusions from 54 patients with tuberculous pleural effusion and 33 with malignant pleural effusion using an enzyme-linked immunosorbent assay (ELISA) kit. Pleural effusion interferon gamma and tumor necrosis factor alpha levels were also analyzed by ELISA. The Mann-Whitney U test was used to evaluate the significance of differences. Associations between superoxide dismutase 2 concentrations and sex, age and smoking habits were assessed using Spearman's or Pearson's correlation coefficient analysis. Receiver operator characteristic analysis was performed to evaluate the value of superoxide dismutase 2 levels in the discrimination of tuberculous pleural effusion from malignant pleural effusion. Superoxide dismutase 2 levels were significantly higher in patients with tuberculous pleural effusion compared with those with malignant pleural effusion (p<0.05). When superoxide dismutase 2 was used to differentiate between tuberculous pleural effusions and malignant pleural effusions, the area under the receiver operator characteristic curve was 0.909 (95% confidence interval, 0.827-0.960; p<0.01). With a cut-off value of 54.2 ng/mL, the sensitivity, specificity, positive likelihood ratio and negative likelihood ratio were 75.8% (95%CI: 57.7-88.9%), 98.1% (95%CI: 90.1-99.7%), 40.91 and 0.25, respectively. Furthermore, significant correlations between pleural effusion superoxide dismutase 2 and interferon gamma (r=0.579, p<0.01) and between pleural effusion superoxide dismutase 2 and tumor

  2. Production of superoxide in chloroplast thylakoid membranes ESR study with cyclic hydroxylamines of different lipophilicity.

    PubMed

    Kozuleva, Marina; Klenina, Irina; Proskuryakov, Ivan; Kirilyuk, Igor; Ivanov, Boris

    2011-04-06

    Accumulation of nitroxide radicals, DCP· or TMT·, under illumination of a thylakoid suspension containing either hydrophilic, DCP-H, or lipophilic, TMT-H, cyclic hydroxylamines that have high rate constants of the reaction with superoxide radicals, was measured using ESR. A slower accumulation of TMT· in contrast with DCP· accumulation was explained by re-reduction of TMT· by the carriers of the photosynthetic electron transport chain within the membrane. Superoxide dismutase suppressed TMT· accumulation to a lesser extent than DCP· accumulation. The data are interpreted as evidencing the production of intramembrane superoxide in thylakoids. Copyright © 2011 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  3. Endomorphins 1 and 2 modulate chemotaxis, phagocytosis and superoxide anion production by microglia.

    PubMed

    Azuma, Y; Ohura, K; Wang, P L; Shinohara, M

    2001-09-03

    We evaluate the role of endomorphins 1 and 2 on microglial functions. Endomorphins 1 and 2 blocked phagocytosis of Escherichia coli. In addition, both markedly inhibited chemotaxis toward zymosan-activated serum. In contrast, when microglia was preincubated with these endomorphins, followed by incubation with LPS before stimulation with phorbol 12-myristate 13-acetate (PMA) at 200 nM, they potentiated superoxide anion production. Furthermore, when microglia was preincubated with these endomorphins together with PMA at 20 nM, followed by stimulation with PMA at 200 nM, superoxide anion production was potentiated. These results suggest that endomorphins 1 and 2 modulate phagocytosis, chemotaxis and superoxide anion production by microglia.

  4. Complex I generated, mitochondrial matrix-directed superoxide is released from the mitochondria through voltage dependent anion channels.

    PubMed

    Lustgarten, Michael S; Bhattacharya, Arunabh; Muller, Florian L; Jang, Youngmok C; Shimizu, Takahiko; Shirasawa, Takuji; Richardson, Arlan; Van Remmen, Holly

    2012-06-08

    Mitochondrial complex I has previously been shown to release superoxide exclusively towards the mitochondrial matrix, whereas complex III releases superoxide to both the matrix and the cytosol. Superoxide produced at complex III has been shown to exit the mitochondria through voltage dependent anion channels (VDAC). To test whether complex I-derived, mitochondrial matrix-directed superoxide can be released to the cytosol, we measured superoxide generation in mitochondria isolated from wild type and from mice genetically altered to be deficient in MnSOD activity (TnIFastCreSod2(fl/fl)). Under experimental conditions that produce superoxide primarily by complex I (glutamate/malate plus rotenone, GM+R), MnSOD-deficient mitochondria release ∼4-fold more superoxide than mitochondria isolated from wild type mice. Exogenous CuZnSOD completely abolished the EPR-derived GM+R signal in mitochondria isolated from both genotypes, evidence that confirms mitochondrial superoxide release. Addition of the VDAC inhibitor DIDS significantly reduced mitochondrial superoxide release (∼75%) in mitochondria from either genotype respiring on GM+R. Conversely, inhibition of potential inner membrane sites of superoxide exit, including the matrix face of the mitochondrial permeability transition pore and the inner membrane anion channel did not reduce mitochondrial superoxide release in the presence of GM+R in mitochondria isolated from either genotype. These data support the concept that complex I-derived mitochondrial superoxide release does indeed occur and that the majority of this release occurs through VDACs.

  5. Radiation resistance and the CuZn superoxide dismutase, Mn superoxide dismutase, catalase, and glutathione peroxidase activities of seven human cell lines.

    PubMed

    Marklund, S L; Westman, N G; Roos, G; Carlsson, J

    1984-10-01

    CuZn superoxide dismutase, Mn superoxide dismutase, catalase, and glutathione peroxidase form the primary enzymic defense against toxic oxygen reduction metabolites in cells. To test the importance of these protective enzymes in the cellular radiation response, the enzymic activities of seven different human cell lines were determined in parallel with their clonogenic survival characteristics. A positive correlation between the content of glutathione peroxidase in cell lines and their extrapolation numbers (n) and quasithreshold doses (Dq) was detected. Between the cellular contents of the other enzymes and D0, n, and Dq no positive correlations could be established. An interesting finding was a very high Mn superoxide dismutase content in a malignant mesothelioma cell line P7, which had an extremely high D0, 5.0 Gy.

  6. Mitochondrial superoxide radicals mediate programmed cell death in Trypanosoma cruzi: cytoprotective action of mitochondrial iron superoxide dismutase overexpression

    PubMed Central

    Piacenza, Lucía; Irigoín, Florencia; Alvarez, María Noel; Peluffo, Gonzalo; Taylor, Martin C.; Kelly, John M.; Wilkinson, Shane R.; Radi, Rafael

    2006-01-01

    Trypanosoma cruzi undergo PCD (programmed cell death) under appropriate stimuli, the mechanisms of which remain to be established. In the present study, we show that stimulation of PCD in T. cruzi epimastigotes by FHS (fresh human serum) results in rapid (<1 h) externalization of phosphatidylserine and depletion of the low molecular mass thiols dihydrotrypanothione and glutathione. Concomitantly, enhanced generation of oxidants was established by EPR and immuno-spin trapping of radicals using DMPO (5,5-dimethylpyrroline-N-oxide) and augmentation of the glucose flux through the pentose phosphate pathway. In the early period (<20 min), changes in mitochondrial membrane potential and inhibition of respiration, probably due to the impairment of ADP/ATP exchange with the cytosol, were observed, conditions that favour the generation of O2•−. Accelerated rates of mitochondrial O2•− production were detected by the inactivation of the redox-sensitive mitochondrial aconitase and by oxidation of a mitochondrial-targeted probe (MitoSOX). Importantly, parasites overexpressing mitochondrial FeSOD (iron superoxide dismutase) were more resistant to the PCD stimulus, unambiguously indicating the participation of mitochondrial O2•− in the signalling process. In summary, FHS-induced PCD in T. cruzi involves mitochondrial dysfunction that causes enhanced O2•− formation, which leads to cellular oxidative stress conditions that trigger the initiation of PCD cascades; moreover, overexpression of mitochondrial FeSOD, which is also observed during metacyclogenesis, resulted in cytoprotective effects. PMID:17168856

  7. Cryo-Trapping the Distorted Octahedral Reaction Intermediate of Manganese Superoxide Dismutase

    NASA Technical Reports Server (NTRS)

    Borgstahl, Gloria; Snell, Edward H.; Rose, M. Franklin (Technical Monitor)

    2000-01-01

    Superoxide dismutase protects organisms from potentially damaging oxygen radicals by catalyzing the disproportion of superoxide to oxygen and hydrogen peroxide. We report the use of cryogenic temperatures to kinetically trap the 6th ligand bound to the active site of manganese superoxide dismutase. Using cryocrystallography and synchrotron radiation, we describe at 1.55A resolution the six-coordinate, distorted octahedral geometry assumed by the active site during catalysis and compare it to the room temperature, five-coordinate trigonal-bipyramidal active site. Gateway residues Tyr34, His30 and a tightly bound water molecule are implicated in closing off the active site and blocking the escape route of superoxide during dismutation.

  8. Structures of two superoxide dismutases from Bacillus anthracis reveal a novel active centre

    SciTech Connect

    Boucher, Ian W.; Kalliomaa, Anne K.; Levdikov, Vladimir M.; Blagova, Elena V.; Fogg, Mark J.; Brannigan, James A. Wilson, Keith S.; Wilkinson, Anthony J.

    2005-07-01

    The crystal structures of two manganese superoxide dismutases from B. anthracis were solved by X-ray crystallography using molecular replacement. The BA4499 and BA5696 genes of Bacillus anthracis encode proteins homologous to manganese superoxide dismutase, suggesting that this organism has an expanded repertoire of antioxidant proteins. Differences in metal specificity and quaternary structure between the dismutases of prokaryotes and higher eukaryotes may be exploited in the development of therapeutic antibacterial compounds. Here, the crystal structure of two Mn superoxide dismutases from B. anthracis solved to high resolution are reported. Comparison of their structures reveals that a highly conserved residue near the active centre is substituted in one of the proteins and that this is a characteristic feature of superoxide dismutases from the B. cereus/B. anthracis/B. thuringiensis group of organisms.

  9. Cryo-Trapping the Distorted Octahedral Reaction Intermediate of Manganese Superoxide Dismutase

    NASA Technical Reports Server (NTRS)

    Borgstahl, Gloria; Snell, Edward H.

    2000-01-01

    Superoxide dismutase protects organisms from potentially damaging oxygen radicals by catalyzing the disproportion of superoxide to oxygen and hydrogen peroxide. We report the use of cryogenic temperatures to kinetically trap the 6th ligand bound to the active site of manganese superoxide dismutase. Using cryocrystallography and synchrotron radiation, we describe at 1.55A resolution the six-coordinate, distorted octahedral geometry assumed by the active site during catalysis and compare it to the room temperature, five-coordinate trigonal-bipyramidal active site. Gateway residues Tyr34, His30 and a tightly bound water molecule are implicated in closing off the active site and blocking the escape route of superoxide during dismutation.

  10. Hydrogen peroxide efflux from muscle mitochondria underestimates matrix superoxide production: a correction using glutathione depletion

    PubMed Central

    TREBERG, Jason R.; QUINLAN, Casey L.; BRAND, Martin D.

    2010-01-01

    Summary The production of H2O2 by isolated mitochondria is frequently used as a measure of mitochondrial superoxide formation. Matrix superoxide dismutase quantitatively converts matrix superoxide to H2O2. However, matrix enzymes such as the glutathione peroxidases can consume H2O2 and compete with efflux of H2O2, causing an underestimate of superoxide production. To assess this underestimate we depleted matrix glutathione in rat skeletal muscle mitochondria by more than 90% by pretreatment with 1-chloro-2,4-dintrobenzene (CDNB). The pretreatment protocol strongly diminished the mitochondrial capacity to consume exogenous H2O2, consistent with decreased peroxidase capacity, but avoided direct stimulation of superoxide production from complex I. It elevated the observed rates of H2O2 formation from matrix-directed superoxide up to two-fold from several sites of production, defined by substrates and electron transport inhibitors, over a wide range of control rates, from 0.2 to 2.5 nmol H2O2 • min−1 • mg protein−1. Similar results were obtained when glutathione was depleted using monochlorobimane or when soluble matrix peroxidase activity was removed by preparation of submitochondrial particles. The data indicate that the increased H2O2 efflux observed with CDNB pretreatment was a result of glutathione depletion and compromised peroxidase activity. A hyperbolic correction curve was constructed, making H2O2 efflux a more quantitative measure of matrix superoxide production. For rat muscle mitochondria, the correction equation was: [CDNB pretreated rate = control rate + (1.43*(control rate))/(0.55+control rate)]. These results have significant ramifications for the rates and topology of superoxide production by isolated mitochondria. PMID:20491900

  11. Decreased Superoxide Production in Macrophages of Long-lived p66Shc Knock-out Mice*

    PubMed Central

    Tomilov, Alexey A.; Bicocca, Vincent; Schoenfeld, Robert A.; Giorgio, Marco; Migliaccio, Enrica; Ramsey, Jon J.; Hagopian, Kevork; Pelicci, Pier Giuseppe; Cortopassi, Gino A.

    2010-01-01

    A decrease in reactive oxygen species (ROS) production has been associated with extended life span in animal models of longevity. Mice deficient in the p66Shc gene are long-lived, and their cells are both resistant to oxidative stress and produce less ROS. Our microarray analysis of p66Shc(−/−) mouse tissues showed alterations in transcripts involved in heme and superoxide production and insulin signaling. Thus, we carried out analysis of ROS production by NADPH oxidase (PHOX) in macrophages of control and p66Shc knock-out mice. p66Shc(−/−) mice had a 40% reduction in PHOX-dependent superoxide production. To confirm whether the defect in superoxide production was a direct consequence of p66Shc deficiency, p66Shc was knocked down with siRNA in the macrophage cell line RAW264, and a 30% defect in superoxide generation was observed. The pathway of PHOX-dependent superoxide generation was investigated. PHOX protein levels were not decreased in mutant macrophages; however, the rate and extent of phosphorylation of p47phox was decreased in mutants, as was membrane translocation of the complex. Consistently, phosphorylation of protein kinase Cδ, Akt, and ERK (the kinases responsible for phosphorylation of p47phox) was decreased. Thus, p66Shc deficiency causes a defect in activation of the PHOX complex that results in decreased superoxide production. p66Shc-deficient mice have recently been observed to be resistant to atherosclerosis and to oxidant injury in kidney and brain. Because phagocyte-derived superoxide is often a component of oxidant injury and inflammation, we suggest that the decreased superoxide production by PHOX in p66Shc-deficient mice could contribute significantly to their relative protection from oxidant injury and consequent longevity. PMID:19892704

  12. Purification of human copper, zinc superoxide dismutase by copper chelate affinity chromatography

    SciTech Connect

    Weslake, R.J.; Chesney, S.L.; Petkau, A.; Friesen, A.D.

    1986-05-15

    Copper, zinc superoxide dismutase was isolated from human red blood cell hemolysate by DEAE-Sepharose and copper chelate affinity chromatography. Enzyme preparations had specific activities ranging from 3400 to 3800 U/mg and recoveries were approximately 60% of the enzyme activity in the lysate. Copper chelate affinity chromatography resulted in a purification factor of about 60-fold. The homogeneity of the superoxide dismutase preparation was analyzed by sodium dodecyl sulfate-gel electrophoresis, analytical gel filtration chromatography, and isoelectric focusing.

  13. Luminol-amplified chemiluminescence detects mainly superoxide anion produced by human neutrophils

    PubMed Central

    Bedouhène, Samia; Moulti-Mati, Farida; Hurtado-Nedelec, Margarita; Dang, Pham My-Chan; El-Benna, Jamel

    2017-01-01

    Reactive oxygen species (ROS) are produced by numerous biological systems and by several phagocytes such as neutrophils and macrophages. ROS include mostly superoxide anion, hydrogen peroxide, singlet oxygen and hydroxyl radical, which are involved in a variety of biological processes such as immunity, inflammation, apoptosis and cell signaling. Thus, there is a need for a sensitive and reliable method to measure ROS. The luminol-amplified chemiluminescence technique is widely used to measure ROS production by neutrophils; however, it is unclear which ROS species are detected by this technique. In this study, we show that Xanthine/Xanthine oxidase (XXO), a known superoxide-producing system, stimulated a luminol-amplified chemiluminescence in the absence of horseradish peroxidase (HRPO), while the presence of HRPO enhanced the response. Both reactions were inhibited by superoxide dismutase (SOD), but not by catalase, confirming that superoxide anion, and not hydrogen peroxide, is the species oxidizing luminol to produce chemiluminescence. Glucose/Glucose oxidase (GGO), a known hydrogen peroxide-producing system, did not induce luminol-amplified chemiluminescence in the absence of HRPO; however, addition of HRPO resulted in a chemiluminescence response, which was inhibited by catalase, but not by SOD. Myeloperoxidase (MPO), isolated from human neutrophils, was also able to enhance the superoxide- and hydrogen peroxide-dependent luminol-amplified chemiluminescence. The production of ROS by stimulated human neutrophils was detected by luminol-amplified chemiluminescence, which was only partially inhibited by SOD and catalase. Interestingly, adding HRPO to stimulated neutrophils increased the luminol-amplified chemiluminescence, which was strongly inhibited by SOD, but not by catalase. These results show that (a) luminol-amplified chemiluminescence is able to detect superoxide anion in the absence of peroxidases, but not hydrogen peroxide; (b) in the presence of

  14. Angiotensin-(1-7) blocks the angiotensin II-stimulated superoxide production.

    PubMed

    Polizio, Ariel Héctor; Gironacci, Mariela Mercedes; Tomaro, Maria Luján; Peña, Clara

    2007-07-01

    Angiotensin (Ang)-(1-7), a bioactive compound of the renin-angiotensin system, exerts effects leading to blood pressure reduction which counterbalance Ang II pressor actions. The present study was conducted to examine Ang-(1-7) and Ang II effects on superoxide anion production in rat aorta using the lucigenin chemiluminescence method. Ang II dose-dependently increased superoxide anion formation when compared to control levels; a maximal increase (2.5-fold) was observed with 1 x 10(-10)M peptide concentration. The Ang II-stimulated superoxide formation was blocked by 1 x 10(-10)M losartan, the specific AT(1) receptor antagonist, but not by 1 x 10(-10)M PD 123319, the AT(2) receptor antagonist, suggesting that the increased superoxide levels caused by Ang II are mediated through AT(1) receptors activation. The Ang II-stimulated superoxide production was not modified by 2 x 10(-8)M allopurinol or 1 x 10(-7)M indomethacin, but was completely abolished by NAD(P)H oxidase inhibitors: 1 x 10(-8)M diphenylene iodonium, or 2 x 10(-8)M apocynin, demonstrating that NAD(P)H oxidase participates in such response. In contrast to Ang II, Ang-(1-7) concentrations ranging 1 x 10(-12) to 1 x 10(-6)M did not modify superoxide anion levels, but prevented the Ang II-enhanced superoxide production. In conclusion, we demonstrated that Ang-(1-7) blocks the pro-oxidant effects of Ang II, thus reducing the superoxide anion production and delaying the hypertension development.

  15. Ras modulation of superoxide activates ERK-dependent fibronectin expression in diabetes-induced renal injuries.

    PubMed

    Lin, C-L; Wang, F-S; Kuo, Y-R; Huang, Y-T; Huang, H-C; Sun, Y-C; Kuo, Y-H

    2006-05-01

    Although previous studies have demonstrated that diabetic nephropathy is attributable to early extracellular matrix accumulation in glomerular mesangial cells, the molecular mechanism by which high glucose induces matrix protein deposition remains not fully elucidated. Rat mesangial cells pretreated with or without inhibitors were cultured in high-glucose or advanced glycation end product (AGE) conditions. Streptozotocin-induced diabetic rats were given superoxide dismutase (SOD)-conjugated propylene glycol to scavenge superoxide. Transforming growth factor (TGF)-beta1, fibronectin expression, Ras, ERK, p38, and c-Jun activation of glomerular mesangial cells or urinary albumin secretion were assessed. Superoxide, not nitric oxide or hydrogen peroxide, mediated high glucose- and AGE-induced TGF-beta1 and fibronectin expression. Pretreatment with diphenyliodonium, not allopurinol or rotenone, reduced high-glucose and AGE augmentation of superoxide synthesis and fibronection expression. High glucose and AGEs rapidly enhanced Ras activation and progressively increased cytosolic ERK and nuclear c-Jun activation. Inhibiting Ras by manumycin A reduced the stimulatory effects of high glucose and AGEs on superoxide and fibronectin expression. SOD or PD98059 pretreatment reduced high-glucose and AGE promotion of ERK and c-Jun activation. Exogenous SOD treatment in diabetic rats significantly attenuated diabetes induction of superoxide, urinary albumin excretion, 8-hydroxy-2'-deoxyguanosine, TGF-beta1, and fibronectin immunoreactivities in renal glomerular mesangial cells. Ras induction of superoxide activated ERK-dependent fibrosis-stimulatory factor and extracellular matrix gene transcription of mesangial cells. Reduction of oxidative stress by scavenging superoxide may provide an alternative strategy for controlling diabetes-induced early renal injury.

  16. Photosensitization with alpha-terthienyl: the formation of superoxide ion in aqueous media.

    PubMed

    Kagan, J; Bazin, M; Santus, R

    1989-04-01

    It is well known that alpha-terthienyl generates singlet oxygen in organic solvents with high quantum yields. In an aqueous medium, the production of superoxide radical-anion is readily detected by comparing the reduction of ferricytochrome c or nitro blue tetrazolium in the presence and in the absence of superoxide dismutase. Electron transfer reactions from the electronically excited sensitizer are also detected in an argon atmosphere.

  17. Superoxide enhances Ca2+ entry through L-type channels in the renal afferent arteriole.

    PubMed

    Vogel, Paul A; Yang, Xi; Moss, Nicholas G; Arendshorst, William J

    2015-08-01

    Reactive oxygen species regulate cardiovascular and renal function in health and disease. Superoxide participates in acute calcium signaling in afferent arterioles and renal vasoconstriction produced by angiotensin II, endothelin, thromboxane, and pressure-induced myogenic tone. Known mechanisms by which superoxide acts include quenching of nitric oxide and increased ADP ribosyl cyclase/ryanodine-mediated calcium mobilization. The effect(s) of superoxide on other calcium signaling pathways in the renal microcirculation is poorly understood. The present experiments examined the acute effect of superoxide generated by paraquat on calcium entry pathways in isolated rat afferent arterioles. The peak increase in cytosolic calcium concentration caused by KCl (40 mmol/L) was 99±14 nmol/L. The response to this membrane depolarization was mediated exclusively by L-type channels because it was abolished by nifedipine but was unaffected by the T-type channel blocker mibefradil. Paraquat increased superoxide production (dihydroethidium fluorescence), tripled the peak response to KCl to 314±68 nmol/L (P<0.001) and doubled the plateau response. These effects were abolished by tempol and nitroblue tetrazolium, but not by catalase, confirming actions of superoxide and not of hydrogen peroxide. Unaffected by paraquat and superoxide was calcium entry through store-operated calcium channels activated by thapsigargin-induced calcium depletion of sarcoplasmic reticular stores. Also unresponsive to paraquat was ryanodine receptor-mediated calcium-induced calcium release from the sarcoplasmic reticulum. Our results provide new evidence that superoxide enhances calcium entry through L-type channels activated by membrane depolarization in rat cortical afferent arterioles, without affecting calcium entry through store-operated entry or ryanodine receptor-mediated calcium mobilization.

  18. Cu(II)-disulfide complexes display simultaneous superoxide dismutase- and catalase-like activities.

    PubMed

    Aliaga, Margarita E; Andrade-Acuña, Daniela; López-Alarcón, Camilo; Sandoval-Acuña, Cristián; Speisky, Hernán

    2013-12-01

    Superoxide is a potentially toxic by-product of cellular metabolism. We have addressed here the in vitro ability of complexes formed between copper(II) ions and various biologically-occurring disulfides (RSSR: oxidized glutathione, cystine, homocystine and α-lipoic acid) to react with superoxide. The studied complexes were found to react with superoxide (generated by a xanthine/xanthine oxidase system) at rate constants (kCu(II)-RSSR) close to 10(6)M(-1)s(-1), which are three orders of magnitude lower than that reported for superoxide dismutase (SOD) but comparable to that of several other copper-containing complexes reported as SOD mimetics. The interaction between the tested Cu(II)-RSSR and superoxide, led to the generation and recovery of concentrations of hydrogen peroxide and oxygen that were, respectively, below and above those theoretically-expected from a sole SOD mimetic action. Interestingly, oxygen was generated when the Cu(II)-RSSR complexes were directly incubated with hydrogen peroxide. Taken together, these results reveal that the Cu(II)-RSSR complexes not only have the capacity to dismutate superoxide but also to simultaneously act like catalase mimetic molecules. When added to superoxide-overproducing mitochondria (condition attained by its exposure to diclofenac), three of the tested complexes were able (2-4μM), not only to totally restore, but also to lower below the basal level the mitochondrial production of superoxide. The present study is first in reporting on the potential of Cu(II)-disulfide complexes to act as SOD and catalase like molecules, suggesting a potential for these types of molecules to act as such under physiological and/or oxidative-stress conditions.

  19. Chemiluminescence Imaging of Superoxide Anion Detects Beta-Cell Function and Mass

    PubMed Central

    Bronsart, Laura L.; Stokes, Christian; Contag, Christopher H.

    2016-01-01

    Superoxide anion is produced during normal cellular respiration and plays key roles in cellular physiology with its dysregulation being associated with a variety of diseases. Superoxide anion is a short-lived molecule and, therefore, its homeostatic regulation and role in biology and disease requires dynamic quantification with fine temporal resolution. Here we validated coelenterazine as a reporter of intracellular superoxide anion concentration and used it as a dynamic measure both in vitro and in vivo. Chemiluminescence was dependent upon superoxide anion levels, including those produced during cellular respiration, and concentrations varied both kinetically and temporally in response to physiologically relevant fluctuations in glucose levels. In vivo imaging with coelenterazine revealed that beta cells of the pancreas have increased levels of superoxide anion, which acted as a measure of beta-cell function and mass and could predict the susceptibility of mice to diabetes mellitus. Glucose response and regulation are key elements of cellular physiology and organismal biology, and superoxide anion appears to play a fundamental and dynamic role in both of these processes. PMID:26752052

  20. A novel superoxide dismutase from Cicer arietinum L. seedlings: isolation, purification and characterization.

    PubMed

    Singh, Sushant; Singh, Abhay Narayan; Verma, Anil; Dubey, Vikash Kumar

    2013-07-01

    Superoxide dismutase is an important enzyme with various therapeutic applications. Search of a new source of superoxide dismutase with novel properties has significant importance. The current work reports purification of a novel superoxide dismutase enzyme with unique characteristics. A copper zinc superoxide dismutase (Cu-Zn SOD) was purified and characterized from Cicer arietinum L. seedlings germinated under aluminium (Al+3) stress. The specific activity of purified protein was 158 units/mg with 28 fold purification. The superoxide dismutase is a homodimeric protein with approx subunit molecular weight of 33.27 kDa. The enzyme is identified as Cu-Zn category of superoxide dismutase, reflected by H2O2 induced inhibition of in-gel activity and presence of quantifiable copper and zinc ions. The optimum pH range for purified Cu-Zn SOD activity was observed within 6.5-8.5 (highest at pH 8.0) and the pH stability was in the range of 6.0-8.5. The enzyme was more stable at low temperature (below 30°C) and the Km of purified Cu-Zn SOD for riboflavin as substrate was 10.16 ± 2.5 µM. The N-terminal amino acid sequence showed homology at conserved residues with other plant Cu-Zn SODs.

  1. Neonatal developmental pattern of superoxide dismutase and aniline hydroxylase in rat lung

    SciTech Connect

    Kakkar, P.; Jaffery, F.N.; Viswanathan, P.N.

    1986-10-01

    The developmental biology of superoxide dismutase and aniline hydroxylase was followed in rat lungs from prenatal stage to 3 months old. Total superoxide dismutase activity as determined by spectrophotometry as well as electrophoresis was high in the prenatal rat lung, decreased in the first 24 hr postpartum, increased within 7 days, and then decreased gradually to adult levels. On polyacrylamide gel electrophoresis only two isozymic forms of superoxide dismutase were located as achromatic zones in the fetal lung. In the adult rat lung, there were three molecular forms of superoxide dismutase, two in the postmitochondrial supernatant and one in the mitochondrial fraction. Unlike superoxide dismutase, aniline hydroxylase was detectable only after 5 days of age and the activity exhibited a gradual increase afterward up to 1 month of age. The developmental pattern of superoxide dismutase and aniline hydroxylase activities in lung may be significant in understanding the mechanism of body defenses and their regulatory modulations in response to toxic air pollutants and environmental stress.

  2. Osmotic stress stimulates generation of superoxide anion by spermatozoa in horses.

    PubMed

    Burnaugh, L; Ball, B A; Sabeur, K; Thomas, A D; Meyers, S A

    2010-02-01

    The objective of this study was to examine the interplay between osmotic and oxidative stress as well as to determine mechanisms by which osmotic stress increases superoxide generation in spermatozoa of horses. Superoxide production, as measured by dihydroethidium (DHE), increased when spermatozoa of horses were incubated under either hyperosmotic or hyposmotic conditions. This increase in superoxide production was inhibited by the MAP kinase p38 inhibitor, SB203580, and by the superoxide scavenger, tiron. Incubation of spermatozoa under hyperosmotic conditions increased overall protein tyrosine phosphorylation as measured by western blotting techniques; however, a similar increase was not detected when spermatozoa were incubated under hyposmotic conditions. The general protein kinase C (PKC) and protein tyrosine kinase (PTK) inhibitor staurosporine inhibited (P<0.05) tyrosine phosphorylation in samples from cells under hyperosmotic conditions. In addition, the NADPH oxidase inhibitor diphenyleneiodonium (DPI) also inhibited (P<0.05) protein tyrosine phosphorylation in cells under hyperosmotic conditions. In summary, these data indicate that incubation of equine spermatozoa under both hyposmotic and hyperosmotic conditions can increase superoxide anion generation. Under hyperosmotic conditions, this increased generation of superoxide anion was accompanied by increased protein tyrosine phosphorylation.

  3. Apparent hydroxyl radical production by peroxynitrite: implications for endothelial injury from nitric oxide and superoxide.

    PubMed Central

    Beckman, J S; Beckman, T W; Chen, J; Marshall, P A; Freeman, B A

    1990-01-01

    Superoxide dismutase reduces injury in many disease processes, implicating superoxide anion radical (O2-.) as a toxic species in vivo. A critical target of superoxide may be nitric oxide (NO.) produced by endothelium, macrophages, neutrophils, and brain synaptosomes. Superoxide and NO. are known to rapidly react to form the stable peroxynitrite anion (ONOO-). We have shown that peroxynitrite has a pKa of 7.49 +/- 0.06 at 37 degrees C and rapidly decomposes once protonated with a half-life of 1.9 sec at pH 7.4. Peroxynitrite decomposition generates a strong oxidant with reactivity similar to hydroxyl radical, as assessed by the oxidation of deoxyribose or dimethyl sulfoxide. Product yields indicative of hydroxyl radical were 5.1 +/- 0.1% and 24.3 +/- 1.0%, respectively, of added peroxynitrite. Product formation was not affected by the metal chelator diethyltriaminepentaacetic acid, suggesting that iron was not required to catalyze oxidation. In contrast, desferrioxamine was a potent, competitive inhibitor of peroxynitrite-initiated oxidation because of a direct reaction between desferrioxamine and peroxynitrite rather than by iron chelation. We propose that superoxide dismutase may protect vascular tissue stimulated to produce superoxide and NO. under pathological conditions by preventing the formation of peroxynitrite. PMID:2154753

  4. Chemiluminescence Imaging of Superoxide Anion Detects Beta-Cell Function and Mass.

    PubMed

    Bronsart, Laura L; Stokes, Christian; Contag, Christopher H

    2016-01-01

    Superoxide anion is produced during normal cellular respiration and plays key roles in cellular physiology with its dysregulation being associated with a variety of diseases. Superoxide anion is a short-lived molecule and, therefore, its homeostatic regulation and role in biology and disease requires dynamic quantification with fine temporal resolution. Here we validated coelenterazine as a reporter of intracellular superoxide anion concentration and used it as a dynamic measure both in vitro and in vivo. Chemiluminescence was dependent upon superoxide anion levels, including those produced during cellular respiration, and concentrations varied both kinetically and temporally in response to physiologically relevant fluctuations in glucose levels. In vivo imaging with coelenterazine revealed that beta cells of the pancreas have increased levels of superoxide anion, which acted as a measure of beta-cell function and mass and could predict the susceptibility of mice to diabetes mellitus. Glucose response and regulation are key elements of cellular physiology and organismal biology, and superoxide anion appears to play a fundamental and dynamic role in both of these processes.

  5. Macroporous mesh of nanoporous gold in electrochemical monitoring of superoxide release from skeletal muscle cells.

    PubMed

    Banan Sadeghian, Ramin; Han, Jiuhui; Ostrovidov, Serge; Salehi, Sahar; Bahraminejad, Behzad; Ahadian, Samad; Chen, Mingwei; Khademhosseini, Ali

    2017-02-15

    Real-time monitoring of metabolically relevant biochemicals released in minuscule amounts is of utmost diagnostic importance. Superoxide anion as a primary member of reactive oxygen species, has physiological and pathological effects that depend on its concentration and release rate. Here we present fabrication and successfully testing of a highly sensitive electrochemical biosensor featuring a three-dimensional macroporous mesh of nanoporous gold tailored to measure the dynamics of extracellular superoxide concentration. Wide and accessible surface of the mesh combined with high porosity of the thin nanoporous gold coating enables capturing the analyte in pico- to nano-molar ranges. The mesh is functionalized with cytochrome-c (cyt-c) and incorporated as a working electrode to measure the release rate of drug-induced superoxides from C2C12 cells through a porous membrane. The device displays a considerably improved superoxide sensitivity of 7.29nAnM(-)(1)cm(-)(2) and a low level of detection of 70pM. Such sensitivity is orders of magnitude higher than any similar enzyme-based electrochemical superoxide sensor and is attributed to the facile diffusion of the analyte through the well-spread nanofeatured gold skin. Superoxide generation rates captured from monolayer myoblast cultures containing about 4×10(4) cells, varied from 1.0 to 9.0nMmin(-)(1) in a quasi-linear fashion as a function of drug concentration. This work provides a platform for the development of highly sensitive molecular electrochemical biosensors.

  6. The primary structure of Cu-Zn superoxide dismutase from Photobacterium leiognathi: evidence for a separate evolution of Cu-Zn superoxide dismutase in bacteria.

    PubMed

    Steffens, G J; Bannister, J V; Bannister, W H; Flohé, L; Günzler, W A; Kim, S M; Otting, F

    1983-06-01

    The complete amino-acid sequence of the copper-zinc superoxide dismutase of the Photobacterium leiognathi was determined. The fragmentation strategy employed included cyanogen bromide cleavage at its methionine residues and the only tryptophan residue. The S-carboxymethylated chain was further cleaved by means of trypsin, in order to obtain overlapping fragments. For sequence determination automated solid or liquid-phase techniques of Edman degradation were used. C-Terminal amino acids of the entire chain were determined after treatment with carboxypeptidase A. Comparison of the primary structure of this bacterial Cu-Zn superoxide dismutase with the established amino-acid sequences of the other eukaryotic Cu-Zn superoxide dismutases revealed clear homologies. Correspondingly, the Cu-Zn-binding amino-acid residues of the active centre were localized: His45, His47, His70, His79, His125 and Asp91. The two cysteine residues in position 52 and 147 were homologous to the cysteine residues, modelling the essential intrachain disulfide bridge of the corresponding bovine enzyme. As only 25-30% of aligned sequence positions were found to be identical, the enzyme of P. leiognathi shows only a remote phylogenetic relationship towards eukaryotic Cu-Zn superoxide dismutases. When compared to the established phylogenetic tree of the cytochrome c family, this indicates a separate evolution of Cu-Zn superoxide dismutase in Photobacterium. Therefore, a natural gene transfer from the eukaryotic host (ponyfish) to the prokaryotic photobacterium, which Martin and Fridovich postulated 1981 (J. Biol. Chem. 256, 6080-6089) on the basis of amino-acid compositions, can be excluded.

  7. NFAT is required for spontaneous pulmonary hypertension in superoxide dismutase 1 knockout mice

    PubMed Central

    Ramiro-Diaz, Juan Manuel; Nitta, Carlos H.; Maston, Levi D.; Codianni, Simon; Giermakowska, Wieslawa; Resta, Thomas C.

    2013-01-01

    Elevated reactive oxygen species are implicated in pulmonary hypertension (PH). Superoxide dismutase (SOD) limits superoxide bioavailability, and decreased SOD activity is associated with PH. A decrease in SOD activity is expected to increase superoxide and reduce hydrogen peroxide levels. Such an imbalance of superoxide/hydrogen peroxide has been implicated as a mediator of nuclear factor of activated T cells (NFAT) activation in epidermal cells. We have shown that NFATc3 is required for chronic hypoxia-induced PH. However, it is unknown whether NFATc3 is activated in the pulmonary circulation in a mouse model of decreased SOD1 activity and whether this leads to PH. Therefore, we hypothesized that an elevated pulmonary arterial superoxide/hydrogen peroxide ratio activates NFATc3, leading to PH. We found that SOD1 knockout (KO) mice have elevated pulmonary arterial wall superoxide and decreased hydrogen peroxide levels compared with wild-type (WT) littermates. Right ventricular systolic pressure (RVSP) was elevated in SOD1 KO and was associated with pulmonary arterial remodeling. Vasoreactivity to endothelin-1 was also greater in SOD1 KO vs. WT mice. NFAT activity and NFATc3 nuclear localization were increased in pulmonary arteries from SOD1 KO vs. WT mice. Administration of A-285222 (selective NFAT inhibitor) decreased RVSP, arterial wall thickness, vasoreactivity, and NFAT activity in SOD1 KO mice to WT levels. The SOD mimetic, tempol, also reduced NFAT activity, NFATc3 nuclear localization, and RVSP to WT levels. These findings suggest that an elevated superoxide/hydrogen peroxide ratio activates NFAT in pulmonary arteries, which induces vascular remodeling and increases vascular reactivity leading to PH. PMID:23475768

  8. HIV-1 Nef regulates the release of superoxide anions from human macrophages

    PubMed Central

    2005-01-01

    The NADPH oxidase enzymatic complex participates in the oxidative burst by producing ROS (reactive oxygen species). Altered levels of ROS production may have pathogenetic implications due to the loss of some innate immune functions such as oxidative burst and phagocytosis. Considering that HIV-1 Nef protein plays a primary role in AIDS pathogenesis, by affecting the immune system, we sought to dissect possible effects of Nef on the release of superoxide anions. We show here that the inducible expression of Nef in human phagocytic cells modulates the superoxide release in a biphasic manner. In particular, an early Nef-induced increase of the superoxide release was followed by a dramatic decrease starting from 10 h after the Nef induction. This was observed whatever the presence of cell activators such as GM-CSF (granulocyte/macrophage colony-stimulating factor) or fMLP (N-formyl-L-methionyl-L-leucyl-L-phenylalanine). Whereas the early increase in superoxide release is probably the result of the already described Nef-dependent activation of PAK-2 (p21-activated kinase 2)–Rac2, we were interested in investigating the mechanisms underlying the late inhibition of superoxide release observed originally. In this regard, we individuated at least three independent requirements for the Nef-induced blockade of superoxide release: (i) the active protein synthesis; (ii) both the membrane localization and the interaction with endocytotic machinery of Nef; and (iii) the release of soluble factor(s). Moreover, we observed that IL-10 (interleukin-10) inhibits superoxide release, whereas its depletion restored NADPH oxidase activity. We propose that the cell membrane-to-lysosome Nef transit leads to the synthesis and release of soluble factor(s) and, among them, IL-10 might significantly contribute to the inhibition of NAPDH oxidase activity. PMID:15847608

  9. Lung Extracellular Superoxide Dismutase Overexpression Lessens Bleomycin-Induced Pulmonary Hypertension and Vascular Remodeling

    PubMed Central

    Van Rheen, Zachary; Fattman, Cheryl; Domarski, Shannon; Majka, Susan; Klemm, Dwight; Stenmark, Kurt R.; Nozik-Grayck, Eva

    2011-01-01

    Interstitial lung disease is a devastating disease in humans that can be further complicated by the development of secondary pulmonary hypertension. Accumulating evidence indicates that the oxidant superoxide can contribute to the pathogenesis of both interstitial lung disease and pulmonary hypertension. We used a model of pulmonary hypertension secondary to bleomycin-induced pulmonary fibrosis to test the hypothesis that an imbalance in extracellular superoxide and its antioxidant defense, extracellular superoxide dismutase, will promote pulmonary vascular remodeling and pulmonary hypertension. We exposed transgenic mice overexpressing lung extracellular superoxide dismutase and wild-type littermates to a single dose of intratracheal bleomycin, and evaluated the mice weekly for up to 35 days. We assessed pulmonary vascular remodeling and the expression of several genes critical to lung fibrosis, as well as pulmonary hypertension and mortality. The overexpression of extracellular superoxide dismutase protected against late remodeling within the medial, adventitial, and intimal layers of the vessel wall after the administration of bleomycin, and attenuated pulmonary hypertension at the same late time point. The overexpression of extracellular superoxide dismutase also blocked the early up-regulation of two key genes in the lung known to be critical in pulmonary fibrosis and vascular remodeling, the transcription factor early growth response–1 and transforming growth factor–β. The overexpression of extracellular superoxide dismutase attenuated late pulmonary hypertension and significantly improved survival after exposure to bleomycin. These data indicate an important role for an extracellular oxidant/antioxidant imbalance in the pathogenesis of pulmonary vascular remodeling associated with secondary pulmonary hypertension attributable to bleomycin-induced lung fibrosis. PMID:20539010

  10. Magnetic Resonance Imaging of Mitochondrial Dysfunction and Metabolic Activity, Accompanied by Overproduction of Superoxide.

    PubMed

    Bakalova, Rumiana; Georgieva, Ekaterina; Ivanova, Donika; Zhelev, Zhivko; Aoki, Ichio; Saga, Tsuneo

    2015-12-16

    This study shows that a mitochondria-penetrating nitroxide probe (mito-TEMPO) allows detection of superoxide and visualization of mitochondrial dysfunction in living cells due to the effect of T1 shortening in MRI. Mitochondrial dysfunction was induced by treatment of cells with rotenone and 2-methoxyestradiol (2-ME/Rot). The MRI measurements were performed on 7T MRI. The 2-ME/Rot-treated cells were characterized by overproduction of superoxide, which was confirmed by a conventional dihydroethidium test. In the presence of mito-TEMPO, the intensity of MRI signal in 2-ME/Rot-treated cells was ∼30-40% higher, in comparison with that in untreated cells or culture media. In model (cell-free) systems, we observed that superoxide, but not hydrogen peroxide, increased the intensity of T1-weighted MRI signal of mito-TEMPO. Moreover, the superoxide restores the T1-weighted MRI contrast of mito-TEMPOH, a noncontrast (diamagnetic) analogue of mito-TEMPO. This was also confirmed by using EPR spectroscopy. The results demonstrate that superoxide radical is involved in the enhancement of T1-weighted MRI contrast in living cells, in the absence and presence of mito-TEMPO. This report gives a direction for discovering new opportunities for functional MRI, for detection of metabolic activity, accompanied by overproduction of superoxide, as well as by disturbance of the balance between superoxide and hydrogen peroxide, a very important approach to clarify the fine molecular mechanisms in the regulation of many pathologies. The visualization of mitochondrial activity in real-time can be crucial to clarify the molecular mechanism of the functional MRI in its commonly accepted definition, as a method for detection of neurovascular coupling.

  11. Species-specific control of external superoxide levels by the coral holobiont during a natural bleaching event

    NASA Astrophysics Data System (ADS)

    Diaz, Julia M.; Hansel, Colleen M.; Apprill, Amy; Brighi, Caterina; Zhang, Tong; Weber, Laura; McNally, Sean; Xun, Liping

    2016-12-01

    The reactive oxygen species superoxide (O2.-) is both beneficial and detrimental to life. Within corals, superoxide may contribute to pathogen resistance but also bleaching, the loss of essential algal symbionts. Yet, the role of superoxide in coral health and physiology is not completely understood owing to a lack of direct in situ observations. By conducting field measurements of superoxide produced by corals during a bleaching event, we show substantial species-specific variation in external superoxide levels, which reflect the balance of production and degradation processes. Extracellular superoxide concentrations are independent of light, algal symbiont abundance and bleaching status, but depend on coral species and bacterial community composition. Furthermore, coral-derived superoxide concentrations ranged from levels below bulk seawater up to ~120 nM, some of the highest superoxide concentrations observed in marine systems. Overall, these results unveil the ability of corals and/or their microbiomes to regulate superoxide in their immediate surroundings, which suggests species-specific roles of superoxide in coral health and physiology.

  12. Species-specific control of external superoxide levels by the coral holobiont during a natural bleaching event

    PubMed Central

    Diaz, Julia M.; Hansel, Colleen M.; Apprill, Amy; Brighi, Caterina; Zhang, Tong; Weber, Laura; McNally, Sean; Xun, Liping

    2016-01-01

    The reactive oxygen species superoxide (O2·−) is both beneficial and detrimental to life. Within corals, superoxide may contribute to pathogen resistance but also bleaching, the loss of essential algal symbionts. Yet, the role of superoxide in coral health and physiology is not completely understood owing to a lack of direct in situ observations. By conducting field measurements of superoxide produced by corals during a bleaching event, we show substantial species-specific variation in external superoxide levels, which reflect the balance of production and degradation processes. Extracellular superoxide concentrations are independent of light, algal symbiont abundance and bleaching status, but depend on coral species and bacterial community composition. Furthermore, coral-derived superoxide concentrations ranged from levels below bulk seawater up to ∼120 nM, some of the highest superoxide concentrations observed in marine systems. Overall, these results unveil the ability of corals and/or their microbiomes to regulate superoxide in their immediate surroundings, which suggests species-specific roles of superoxide in coral health and physiology. PMID:27924868

  13. Endothelin-1 critically influences cardiac function via superoxide-MMP9 cascade

    PubMed Central

    Hathaway, Catherine K.; Grant, Ruriko; Hagaman, John R.; Hiller, Sylvia; Li, Feng; Xu, Longquan; Chang, Albert S.; Madden, Victoria J.; Bagnell, C. Robert; Rojas, Mauricio; Kim, Hyung-Suk; Wu, Bingruo; Zhou, Bin; Smithies, Oliver; Kakoki, Masao

    2015-01-01

    We have generated low-expressing and high-expressing endothelin-1 genes (L and H) and have bred mice with four levels of expression: L/L, ∼20%; L/+, ∼65%; +/+ (wild type), 100%; and H/+, ∼350%. The hypomorphic L allele can be spatiotemporally switched to the hypermorphic H allele by Cre-loxP recombination. Young adult L/L and L/+ mice have dilated cardiomyopathy, hypertension, and increased plasma volumes, together with increased ventricular superoxide levels, increased matrix metalloproteinase 9 (Mmp9) expression, and reduced ventricular stiffness. H/+ mice have decreased plasma volumes and significantly heavy stiff hearts. Global or cardiomyocyte-specific switching expression from L to H normalized the abnormalities already present in young adult L/L mice. An epithelial sodium channel antagonist normalized plasma volume and blood pressure, but only partially corrected the cardiomyopathy. A superoxide dismutase mimetic made superoxide levels subnormal, reduced Mmp9 overexpression, and substantially improved cardiac function. Genetic absence of Mmp9 also improved cardiac function, but increased superoxide remained. We conclude that endothelin-1 is critical for maintaining normal contractile function, for controlling superoxide and Mmp9 levels, and for ensuring that the myocardium has sufficient collagen to prevent overstretching. Even a modest (∼35%) decrease in endothelin-1 gene (Edn1) expression is sufficient to cause cardiac dysfunction. PMID:25848038

  14. Yeast and Mammalian Metallothioneins Functionally Substitute for Yeast Copper-Zinc Superoxide Dismutase

    NASA Astrophysics Data System (ADS)

    Tamai, Katherine T.; Gralla, Edith B.; Ellerby, Lisa M.; Valentine, Joan S.; Thiele, Dennis J.

    1993-09-01

    Copper-zinc superoxide dismutase catalyzes the disproportionation of superoxide anion to hydrogen peroxide and dioxygen and is thought to play an important role in protecting cells from oxygen toxicity. Saccharomyces cerevisiae strains lacking copper-zinc superoxide dismutase, which is encoded by the SOD1 gene, are sensitive to oxidative stress and exhibit a variety of growth defects including hypersensitivity to dioxygen and to superoxide-generating drugs such as paraquat. We have found that in addition to these known phenotypes, SOD1-deletion strains fail to grow on agar containing the respiratory carbon source lactate. We demonstrate here that expression of the yeast or monkey metallothionein proteins in the presence of copper suppresses the lactate growth defect and some other phenotypes associated with SOD1-deletion strains, indicating that copper metallothioneins substitute for copper-zinc superoxide dismutase in vivo to protect cells from oxygen toxicity. Consistent with these results, we show that yeast metallothionein mRNA levels are dramatically elevated under conditions of oxidative stress. Furthermore, in vitro assays demonstrate that yeast metallothionein, purified or from whole-cell extracts, exhibits copper-dependent antioxidant activity. Taken together, these data suggest that both yeast and mammalian metallothioneins may play a direct role in the cellular defense against oxidative stress by functioning as antioxidants.

  15. Measurement of NAD(P)H oxidase-derived superoxide with the luminol analogue L-012.

    PubMed

    Daiber, Andreas; August, Michael; Baldus, Stephan; Wendt, Maria; Oelze, Matthias; Sydow, Karsten; Kleschyov, Andrei L; Munzel, Thomas

    2004-01-01

    In the present study we sought to determine the ability of the chemiluminescence dye 8-amino-5-chloro-7-phenylpyridol[3,4-d]pyridazine-1,4-(2H,3H)dione sodium salt (L-012) to detect superoxide in different biological systems. In human whole blood or isolated leukocytes, the sensitivity of the luminol analogue L-012 to detect superoxide was higher as compared with luminol, lucigenin, coelenterazine, and the fluorescence dye dihydroethidine. In isolated leukocytes as well as aortic rings from control (New Zealand White) and hyperlipidemic (Watanabe heritable hyperlipidemic) rabbits, L-012-enhanced chemiluminescence was successful in detecting differences in superoxide formation under basal conditions and on stimulation with the direct activator of protein kinase C, phorbol 12,13-dibutyrate (PDBu). The effects of PDBu were abrogated by gliotoxin and inhibitors of protein kinase C such as chelerythrine, identifying NAD(P)H oxidase as the significant superoxide source. Experiments using electron paramagnetic resonance and the spin trap 5-diethoxyphosphoryl-5-methyl-1-pyrroline-N-oxide revealed that in contrast to lucigenin, L-012 is not subject to redox cycling. These findings indicate that L-012-enhanced chemiluminescence represents a sensitive and reliable probe to detect superoxide in whole blood, inflammatory cells, and vascular tissue.

  16. Nitric oxide synthesis inhibition induces leukocyte adhesion via superoxide and mast cells.

    PubMed

    Kubes, P; Kanwar, S; Niu, X F; Gaboury, J P

    1993-10-01

    Recent work has demonstrated that inhibition of nitric oxide production with various nitric oxide synthesis inhibitors (L-NAME, L-NMMA) initiate leukocyte adhesion to postcapillary venules. The objective of this study was to elucidate the mechanism (or mechanisms) that promote the L-NAME-induced leukocyte response. Intravital microscopy was used to examine 25-40 microns venules in the rat mesentery. Nitric oxide synthesis was inhibited with L-NAME and leukocyte adhesion was observed over the first 60 min. The fourfold increase in leukocyte adhesion was independent of alterations in venular red blood cell velocity. The adhesion was superoxide-mediated inasmuch as superoxide dismutase (SOD) abolished the rise in leukocyte adhesion associated with nitric oxide synthesis inhibition. Ketotifen, a mast cell stabilizer, also abolished the rise in leukocyte adhesion induced by L-NAME. Histology revealed that mast cell degranulation occurred only in animals treated with L-NAME but not in animals pretreated with SOD or ketotifen. This observation suggests that mast cells become activated in the absence of nitric oxide production and superoxide contributes to the mast cell activation. The L-NAME-induced leukocyte adhesion could be reproduced by infusing hypoxanthine/xanthine oxidase (a superoxide generating system) or compound 48/80 (an activator of mast cells) and both responses were attenuated by ketotifen. These data suggest that inhibition of nitric oxide synthesis results in a superoxide and mast cell-dependent leukocyte adhesion.

  17. [Involvement of carbonate/bicarbonate ions in the superoxide-generating reaction of adrenaline autoxidation].

    PubMed

    Sirota, T V

    2015-01-01

    An important role of carbonate/bicarbonate ions has been recognized in the superoxide generating reaction of adrenaline autooxidation in an alkaline buffer (a model of quinoid adrenaline oxidation in the body). It is suggested that these ions are directly involved not only in formation of superoxide anion radical (О(2)(-)) but also other radicals derived from the carbonate/bicarbonate buffer. Using various buffers it was shown that the rate of accumulation of adrenochrome, the end product of adrenaline oxidation, and the rate of О(2)(-)· formation depend on concentration of carbonate/bicarbonate ions in the buffer and that these ions significantly accelerate adrenaline autooxidation thus demonstrating prooxidant properties. The detectable amount of diformazan, the product of nitro blue tetrazolium (NBT) reduction, was significantly higher than the amount of adrenochrome formed; taking into consideration the literature data on О(2)(-)· detection by NBT it is suggested that adrenaline autooxidation is accompanied by one-electron reduction not only of oxygen dissolved in the buffer and responsible for superoxide formation but possible carbon dioxide also dissolved in the buffer as well as carbonate/bicarbonate buffer components leading to formation of corresponding radicals. The plots of the dependence of the inhibition of adrenochrome and diformazan formation on the superoxide dismutase concentration have shown that not only superoxide radicals are formed during adrenaline autooxidation. Since carbonate/bicarbonate ions are known to be universally present in the living nature, their involvement in free radical processes proceeding in the organism is discussed.

  18. Detection of drug-induced, superoxide-mediated cell damage and its prevention by antioxidants.

    PubMed

    Horáková, K; Sovcíková, A; Seemannová, Z; Syrová, D; Busányová, K; Drobná, Z; Ferencík, M

    2001-03-15

    The mode of the cytotoxic activity of three benzo(c)fluorene derivatives was characterized. The observed morphological changes of lysosomes or variations of mitochondrial activity are assumed to be the consequence of cell protection against oxidative damage and/or the part of the damage process. To establish the relationship between the quantity of superoxide (O2*-) generated and the degree of damage resulting from O2*-, a simple system based on measurement of 3-(4-iodophenyl)-2-(4-nitrophenyl)-5-phenyltetrazolium chloride (INT) reductase activity in the presence of superoxide dismutase (SOD) was used. The functionality of the chosen battery of in vitro tests was proved using several known superoxide inducers: cyclosporin A (CsA) and benzo(a)pyrene (BP), as well as noninducers: citrinin (CT) and cycloheximide (CH). From the results followed that the cell growth tests are much better indices of toxicity than the other tests. The model system for the evaluation of the protective capacity of antioxidants against superoxide-induced cytotoxicity included simultaneous exposure of HeLa cells to cytotoxic drugs and to quercetin (Qe), an antioxidant of plant origin. The complete abolishment of the inhibition of cell proliferation and clonogenic survival was concluded to be due to the protective effect of the antioxidant. These observations correlated with the decrease of superoxide content as estimated by the INT-reductase assay in the presence of SOD using the same model system, as well as with the increase of intracellular SOD content and its activity.

  19. Nitration and Inactivation of Manganese Superoxide Dismutase in Chronic Rejection of Human Renal Allografts

    NASA Astrophysics Data System (ADS)

    MacMillan-Crow, L. A.; Crow, John P.; Kerby, Jeffrey D.; Beckman, Joseph S.; Thompson, John A.

    1996-10-01

    Inflammatory processes in chronic rejection remain a serious clinical problem in organ transplantation. Activated cellular infiltrate produces high levels of both superoxide and nitric oxide. These reactive oxygen species interact to form peroxynitrite, a potent oxidant that can modify proteins to form 3-nitrotyrosine. We identified enhanced immunostaining for nitrotyrosine localized to tubular epithelium of chronically rejected human renal allografts. Western blot analysis of rejected tissue demonstrated that tyrosine nitration was restricted to a few specific polypeptides. Immunoprecipitation and amino acid sequencing techniques identified manganese superoxide dismutase, the major antioxidant enzyme in mitochondria, as one of the targets of tyrosine nitration. Total manganese superoxide dismutase protein was increased in rejected kidney, particularly in the tubular epithelium; however, enzymatic activity was significantly decreased. Exposure of recombinant human manganese superoxide dismutase to peroxynitrite resulted in a dose-dependent (IC50 = 10 μ M) decrease in enzymatic activity and concomitant increase in tyrosine nitration. Collectively, these observations suggest a role for peroxynitrite during development and progression of chronic rejection in human renal allografts. In addition, inactivation of manganese superoxide dismutase by peroxynitrite may represent a general mechanism that progressively increases the production of peroxynitrite, leading to irreversible oxidative injury to mitochondria.

  20. Ab initio molecular dynamics of the reaction of quercetin with superoxide radical

    NASA Astrophysics Data System (ADS)

    Lespade, Laure

    2016-08-01

    Superoxide plays an important role in biology but in unregulated concentrations it is implicated in a lot of diseases such as cancer or atherosclerosis. Antioxidants like flavonoids are abundant in plant and are good scavengers of superoxide radical. The modeling of superoxide scavenging by flavonoids from the diet still remains a challenge. In this study, ab initio molecular dynamics of the reaction of the flavonoid quercetin toward superoxide radical has been carried out using Car-Parrinello density functional theory. The study has proven different reactant solvation by modifying the number of water molecules surrounding superoxide. The reaction consists in the gift of a hydrogen atom of one of the hydroxyl groups of quercetin to the radical. When it occurs, it is relatively fast, lower than 100 fs. Calculations show that it depends largely on the environment of the hydroxyl group giving its hydrogen atom, the geometry of the first water layer and the presence of a certain number of water molecules in the second layer, indicating a great influence of the solvent on the reactivity.

  1. Superoxide scavenging by thiol/copper complex of captopril--an EPR spectroscopy study.

    PubMed

    Reguli, J; Misík, V

    1995-02-01

    Scavenging of superoxide radical by angiotensin converting enzyme (ACE) inhibitor captopril (CAP), a thiol compound, was studied by several investigators and the results were contradictory; while some reported a high superoxide scavenging activity of CAP others found that CAP removed superoxide inefficiently. In this work we show that in the presence of copper ions the apparent rate of superoxide removal by CAP (molar ratio CAP:CuSO4 4:1) was two orders of magnitude higher (approximately 1.5 x 10(5) M-1s-1 at pH 7.4) than the literature value for superoxide scavenging by CAP alone (< 10(3) M-1s-1 at pH 7.4). We presume that in the presence of copper ions a CAP/copper complex with a SOD-mimicking activity is being formed. Similar results were also obtained with another thiol glutathione (GSH). The possible role of the CAP/copper complexes in the anti-inflammatory effect of CAP is discussed.

  2. Catalytic superoxide scavenging by metal complexes of the calcium chelator EGTA and contrast agent EHPG.

    PubMed

    Fisher, Anna E O; Hague, Theresa A; Clarke, Charlotte L; Naughton, Declan P

    2004-10-08

    Metal ion chelators widely used in experimental protocols and clinical diagnosis are generally assumed to be inert. We previously reported that the ubiquitous chelator EDTA has high levels of superoxide suppressing activity. Here, we report that the common chelators calcium chelator EGTA and contrast agent EHPG have significant activities in suppressing superoxide levels depending on the nature of metal ion chelated. The most active species is Mn(II)-EGTA which exhibited an IC50 value of 0.19 microM for superoxide destruction. In addition, IC50 values for Mn(II)-EHPG and 2Cu(II)-EGTA were 0.69 and 0.60 microM, respectively. In conclusion, Mn(II) and Cu(II) complexes of the common chelators EGTA and EHPG exhibit considerable superoxide scavenging activities. Caution should be employed in their use in biological systems where superoxide has a key role and they may be useful for the development of catalytic anti-oxidants. Copyright 2004 Elsevier Inc.

  3. A Printed Superoxide Dismutase Coated Electrode for the Study of Macrophage Oxidative Burst

    PubMed Central

    Hiatt, Leslie A.; McKenzie, Jennifer R.; Deravi, Leila F.; Harry, Reese S.; Wright, David W.; Cliffel, David E.

    2012-01-01

    The miniaturization of electrochemical sensors allows for the minimally invasive and cost effective examination of cellular responses at a high efficacy rate. In this work, an ink-jet printed superoxide dismutase electrode was designed, characterized, and utilized as a novel microfluidic device to examine the metabolic response of a 2D layer of macrophage cells. Since superoxide production is one of the first indicators of oxidative burst, macrophage cells were exposed within the microfluidic device to phorbol myristate acetate (PMA), a known promoter of oxidative burst, and the production of superoxide was measured. A 46 ± 19% increase in current was measured over a 30 min time period demonstrating successful detection of sustained macrophage oxidative burst, which corresponds to an increase in the superoxide production rate by 9 ± 3 attomoles/cell/sec. Linear sweep voltammetry was utilized to show the selectivity of this sensor for superoxide over hydrogen peroxide. This novel controllable microfluidic system can be used to study the impact of multiple effectors from a large number of bacteria or other invaders along a 2D layer of macrophages, providing an in vitro platform for improved electrochemical studies of metabolic responses. PMID:22257735

  4. Superoxide dismutase and catalase conjugated to polyethylene glycol increases endothelial enzyme activity and oxidant resistance

    SciTech Connect

    Beckman, J.S.; Minor, R.L. Jr.; White, C.W.; Repine, J.E.; Rosen, G.M.; Freeman, B.A.

    1988-05-15

    Covalent conjugation of superoxide dismutase and catalase with polyethylene glycol (PEG) increases the circulatory half-lives of these enzymes from <10 min to 40 h, reduced immunogenicity, and decreases sensitivity to proteolysis. Because PEG has surface active properties and can induce cell fusion, the authors hypothesized that PEG conjugation could enhance cell binding and association of normally membrane-impermeable enzymes. Incubation of cultured porcine aortic endothelial cells with /sup 125/I-PEG-catalase or /sup 125/I-PEG-superoxide dismutase produced a linear, concentration-dependent increase in cellular enzyme activity and radioactivity. Fluorescently labeled PEG-superoxide dismutase incubated with endothelial cells showed a vesicular localization. Mechanical injury to cell monolayers, which is known to stimulate endocytosis, further increased the uptake of fluorescent PEG-superoxide dismutase. Addition of PEG and PEG-conjugated enzymes perturbed the spin-label binding environment, indicative of producing an increase in plasma membrane fluidity. Thus, PEG conjugation to superoxide dismutase and catalase enhances cell association of these enzymes in a manner which increases cellular enzyme activities and provides prolonged protection from partially reduced oxygen species.

  5. Discriminative protection against hydroxyl and superoxide anion radicals by quercetin in human leucocytes in vitro.

    PubMed

    Wilms, Lonneke C; Kleinjans, Jos C S; Moonen, Edwin J C; Briedé, Jacob J

    2008-03-01

    Antioxidants play a vital role in the cellular protection against oxidative damage. Quercetin is a well-investigated antioxidant and known to be able to protect against cellular oxidative DNA damage. In this study, we tried to relate the protection by quercetin pre-treatment against oxidative DNA damage in human leucocytes in vitro to the interaction of quercetin in solution with hydroxyl and superoxide anion radicals as measured by electron spin resonance (ESR) spectrometry, using DMPO as a spin trap. Further, scavenging capacity of quercetin-treated leucocytes in vitro was evaluated by ESR spectrometry. Quercetin appears capable of protecting human leucocytes against oxidative DNA damage caused by hydrogen peroxide in a dose-dependent manner. The protection of leucocytes against superoxides is ambiguous. Incubation concentrations of quercetin (1, 10, and 50 microM) reduced levels of superoxide-induced oxidative DNA damage, while at 100 microM the amount of damage was increased. These results are supported by ESR-findings on quercetin in solution, also showing a prooxidant effect at 100 microM. ESR spectroscopy showed rate constant values for the reaction kinetics of quercetin in lowering iron-dependent hydroxyl radical formation and NADH-dependent superoxide anion formation of respectively 3.2 x 10(12)M(-1)s(-1) and 1.1 x 10(4)M(-1)s(-1). This shows that quercetin is a more potent inhibitor of hydroxyl radical formation than a scavenger of superoxide anions.

  6. Extensive screening for edible herbal extracts with potent scavenging activity against superoxide anions.

    PubMed

    Saito, Keita; Kohno, Masahiro; Yoshizaki, Fumihiko; Niwano, Yoshimi

    2008-06-01

    To search for edible herbal extracts with potent antioxidant activity, we conducted a large scale screening based on the superoxide scavenging activity. That is, scavenging activity against superoxide anions were extensively screened from ethanol extracts of approximately 1,000 kinds of herbs by applying an electron spin resonance (ESR)-spin trapping method. Among them we chose four edible herbal extracts with prominently potent ability to reduce the signal intensity of 5,5-dimethyl-1-pyrroline-N-oxide (DMPO)-OOH, a spin adduct formed by DMPO and superoxide anion. They are the extracts from Punica granatum (Peel), Syzygium aromaticum (Bud), Mangifera indica (Kernel), and Phyllanthus emblica (Fruit), and are allowed to be used as foodstuffs according to the Japanese legal regulation. The ESR-spin trapping method coupled with steady state kinetic analysis showed that all of the four extracts directly scavenge superoxide anions, and that the superoxide scavenging potential of any of the extracts was comparable to that of L-ascorbic acid. Furthermore, polyphenol determination indicates that the activity is at least in part attributable to polyphenols. These results with such large scale screening might give useful information when choosing a potent antioxidant as a foodstuff.

  7. Bz-423 superoxide signals B cell apoptosis via Mcl-1, Bak, and Bax.

    PubMed

    Blatt, Neal B; Boitano, Anthony E; Lyssiotis, Costas A; Opipari, Anthony W; Glick, Gary D

    2009-10-15

    Bz-423 is a pro-apoptotic 1,4-benzodiazepine with therapeutic properties in murine models of lupus demonstrating selectivity for autoreactive lymphocytes. Bz-423 modulates the F(1)F(0)-ATPase, inducing the formation of superoxide within the mitochondrial respiratory chain, which then functions as a second messenger initiating apoptosis. In order to understand some of the features that contribute to the increased sensitivity of lymphocytes, we report the signaling pathway engaged by Bz-423 in a Burkitt lymphoma cell line (Ramos). Following the generation of superoxide, Bz-423-induced apoptosis requires the activation of Bax and Bak to induce mitochondrial outer membrane permeabilization and cytochrome c release. Knockdown of the BH3-only proteins Bad, Bim, Bik, and Puma inhibits Bz-423 apoptosis, suggesting that these proteins serve as upstream sensors of the oxidant stress induced by Bz-423. Treatment with Bz-423 results in superoxide-dependent Mcl-1 degradation, implicating this protein as the link between Bz-423-induced superoxide and Bax and Bak activation. In contrast to fibroblasts, B cell death induced by Bz-423 is independent of c-Jun N-terminal kinase. These results demonstrate that superoxide generated from the mitochondrial respiratory chain as a consequence of a respiratory transition can signal a specific apoptotic response that differs across cell types.

  8. Metabolic stability of superoxide adducts derived from newly developed cyclic nitrone spin traps.

    PubMed

    Bézière, Nicolas; Hardy, Micael; Poulhès, Florent; Karoui, Hakim; Tordo, Paul; Ouari, Olivier; Frapart, Yves-Michel; Rockenbauer, Antal; Boucher, Jean-Luc; Mansuy, Daniel; Peyrot, Fabienne

    2014-02-01

    Reactive oxygen species are by-products of aerobic metabolism involved in the onset and evolution of various pathological conditions. Among them, the superoxide radical is of special interest as the origin of several damaging species such as H2O2, hydroxyl radical, or peroxynitrite (ONOO(-)). Spin trapping coupled with ESR is a method of choice to characterize these species in chemical and biological systems and the metabolic stability of the spin adducts derived from reaction of superoxide and hydroxyl radicals with nitrones is the main limit to the in vivo application of the method. Recently, new cyclic nitrones bearing a triphenylphosphonium or permethylated β-cyclodextrin moiety have been synthesized and their spin adducts demonstrated increased stability in buffer. In this article, we studied the stability of the superoxide adducts of four new cyclic nitrones in the presence of liver subcellular fractions and biologically relevant reductants using an original setup combining a stopped-flow device and an ESR spectrometer. The kinetics of disappearance of the spin adducts were analyzed using an appropriate simulation program. Our results highlight the interest of the new spin trapping agents CD-DEPMPO and CD-DIPPMPO for specific detection of superoxide with high stability of the superoxide adducts in the presence of liver microsomes.

  9. Endothelin-1 critically influences cardiac function via superoxide-MMP9 cascade.

    PubMed

    Hathaway, Catherine K; Grant, Ruriko; Hagaman, John R; Hiller, Sylvia; Li, Feng; Xu, Longquan; Chang, Albert S; Madden, Victoria J; Bagnell, C Robert; Rojas, Mauricio; Kim, Hyung-Suk; Wu, Bingruo; Zhou, Bin; Smithies, Oliver; Kakoki, Masao

    2015-04-21

    We have generated low-expressing and high-expressing endothelin-1 genes (L and H) and have bred mice with four levels of expression: L/L, ∼20%; L/+, ∼65%; +/+ (wild type), 100%; and H/+, ∼350%. The hypomorphic L allele can be spatiotemporally switched to the hypermorphic H allele by Cre-loxP recombination. Young adult L/L and L/+ mice have dilated cardiomyopathy, hypertension, and increased plasma volumes, together with increased ventricular superoxide levels, increased matrix metalloproteinase 9 (Mmp9) expression, and reduced ventricular stiffness. H/+ mice have decreased plasma volumes and significantly heavy stiff hearts. Global or cardiomyocyte-specific switching expression from L to H normalized the abnormalities already present in young adult L/L mice. An epithelial sodium channel antagonist normalized plasma volume and blood pressure, but only partially corrected the cardiomyopathy. A superoxide dismutase mimetic made superoxide levels subnormal, reduced Mmp9 overexpression, and substantially improved cardiac function. Genetic absence of Mmp9 also improved cardiac function, but increased superoxide remained. We conclude that endothelin-1 is critical for maintaining normal contractile function, for controlling superoxide and Mmp9 levels, and for ensuring that the myocardium has sufficient collagen to prevent overstretching. Even a modest (∼35%) decrease in endothelin-1 gene (Edn1) expression is sufficient to cause cardiac dysfunction.

  10. Catalytic spectrofluorimetric determination of superoxide anion radical and superoxide dismutase activity using N, N-dimethylaniline as the substrate for horseradish peroxidase (HRP)

    NASA Astrophysics Data System (ADS)

    Tang, Bo; Wang, Yan; Chen, Zhen-zhen

    2002-10-01

    The coupled reaction of N, N-dimethylaniline (DMA) with 4-aminoantipyrine (4-AAP) using superoxide anion radical (O 2-) as oxidizing agent under the catalysis of horseradish peroxidase (HRP) was studied. Based on the reaction, O 2- produced by irradiating Vitamin B 2, (V B2) was spectrophotometricly determined at 554 nm. The linear range of this method was 1.8×10 -6-1.2×10 -4 mol l -1 with a detection limit of 5.3×10 -7 mol l -1. The effect of interferences on the determination of O 2- was investigated. The proposed method was successfully applied to the determination of superoxide dismutase (SOD) activity in human blood and mouse blood.

  11. Multiconfigurational and DFT analyses of the electromeric formulation and UV-vis absorption spectra of the superoxide adduct of ferrous superoxide reductase.

    PubMed

    Attia, Amr A A; Cioloboc, Daniela; Lupan, Alexandru; Silaghi-Dumitrescu, Radu

    2016-12-01

    The putative initial adduct of ferrous superoxide reductase (SOR) with superoxide has been alternatively formulated as ferric-peroxo or ferrous-superoxo. The ~600-nm UV-vis absorption band proposed to be assigned to this adduct (either as sole intermediate in the SOR catalytic cycle, or as one of the two intermediates) has recently been interpreted as due to a ligand-to-metal charge transfer, involving thiolate and superoxide in a ferrous complex, contrary to an alternative assignment as a predominantly cysteine thiolate-to-ferric charge transfer in a ferric-peroxo electromer. In an attempt to clarify the electromeric formulation of this adduct, we report a computational study using a multiconfigurational complete active space self-consistent field (MC-CASSCF) wave function approach as well as modelling the UV-vis absorption spectra with time-dependent density functional theory (TD-DFT). The MC-CASSCF calculations disclose a weak interaction between iron and the dioxygenic ligand and a dominant configuration with an essentially ferrous-superoxo character. The computed UV-vis absorption spectra reveal a marked dependence on the choice of density functional - both in terms of location of bands and in terms of orbital contributors. For the main band in the visible region, besides the recently reported thiolate-to-superoxide charge transfer, a more salient, and less functional-dependent, feature is a thiolate-to-ferric iron charge transfer, consistent with a ferric-peroxo electromer. By contrast, the computed UV-vis spectra of a ferric-hydroperoxo SOR model match distinctly better (and with no qualitative dependence on the DFT methodology) the 600-nm band as due to a mainly thiolate-to-ferric character - supporting the assignment of the SOR "600-nm intermediate" as a S=5/2 ferric-hydroperoxo species.

  12. Oxidation of Hantzsch 1,4-dihydropyridines of pharmacological significance by electrogenerated superoxide.

    PubMed

    Ortiz, M E; Núñez-Vergara, L J; Camargo, C; Squella, J A

    2004-03-01

    To study the reaction of a series of Hantzsch dihydropyridines with pharmacological significance such as, nifedipine, nitrendipine, nisoldipine, nimodipine, isradipine and felodipine, with electrogenerated superoxide in order to identify products and postulate a mechanism. The final pyridine derivatives were separated and identified by gas chromatography/mass spectrometry (GC-MS). The intermediates, anion dihydropyridine and the HO2*/HO2- species, were observed from voltammetric studies and controlled potential electrolysis was used to electrogenerate O2*-. The current work reveals that electrogenerated superoxide can quantitatively oxidize Hantzsch dihydropyridines to produce the corresponding aromatized pyridine derivatives. Our results indicate that the aromatization of Hantzsch dihydropyridines by superoxide is initiated by proton transfer from the N1-position on the 1,4-dihydropyridine ring to give the corresponding anion dihydropyridine, which readily undergoes further homogeneous oxidations to provide the final aromatized products. The oxidation of the anionic species of the dihydropyridine is more easily oxidized than the parent compound.

  13. Superoxide metabolism is correlated to the post-anoxic injury of soybean (Glycine max) roots

    SciTech Connect

    Bolles, C.S.; Van Toai, T.T. )

    1990-05-01

    Post-anoxic injury of root tips of soybean seedlings is more severe following a very short (1 hour) period of anoxia than a longer (3-5 hour) period. Anaerobic incubation of root tips in the presence of 100 mM ascorbate, an antioxidant and free-radical-scavenging compound, alleviates the detrimental post-anoxia effects of a very short anoxic treatment. Extracts of root tips which have been treated anoxically for 1 hour have an elevated capacity to produce superoxide anions when subsequently exposed to air, than extracts from seedlings treated anoxically for longer time. Changes in superoxide dismutase (SOD) enzyme activity and SOD-specific RNA sequences will be presented. The results support that post-anoxic injury occurs in soybean roots and that SOD plays a role in the detoxification of superoxide anions.

  14. Dehydroepiandrosterone inhibits the spontaneous release of superoxide radical by alveolar macrophages in vitro in asbestosis

    SciTech Connect

    Rom, W.N.; Harkin, T. )

    1991-08-01

    Asbestosis is characterized by an alveolar macrophage alveolitis with injury and fibrosis of the lower respiratory tract. Alveolar macrophages recovered by bronchoalveolar lavage spontaneously release exaggerated amounts of oxidants including superoxide anion and hydrogen peroxide that may mediate alveolar epithelial cell injury. Dehydroepiandrosterone (DHEA) is a normally occurring adrenal androgen that inhibits glucose-6-phosphate dehydrogenase, the initial enzyme in the pentose phosphate shunt necessary for NADPH generation and superoxide anion formation. In this regard, the authors hypothesized that DHEA may reduce asbestos-induced oxidant release. DHEA added in vitro to alveolar macrophages lavaged from 11 nonsmoking asbestos workers significantly reduced superoxide anion release. DHEA is an antioxidant and potential anticarcinogenic agent that may have a therapeutic role in reducing the increased oxidant burden in asbestos-induced alveolitis of the lower respiratory tract.

  15. Endogenous antioxidant defense induction by melon superoxide dismutase reduces cardiac hypertrophy in spontaneously hypertensive rats.

    PubMed

    Carillon, Julie; Rugale, Caroline; Rouanet, Jean-Max; Cristol, Jean-Paul; Lacan, Dominique; Jover, Bernard

    2014-08-01

    We assessed the influence of SODB, a melon superoxide dismutase (SOD), on left ventricular (LV) hypertrophy in SHR. SODB (4 or 40U SOD) was given orally for 4 or 28 days to SHR. For each treatment period, LV weight index (LVWI) and cardiomyocytes size were measured. SOD, glutathione peroxidase (GPx) and catalase expressions, and LV production and presence of superoxide anion were determined. Pro-inflammatory markers were also measured. SODB reduced LVWI and cardiomyocytes size after 4 or 28 days. Cardiac SOD and GPx increased by 30-40% with SODB. The presence but not production of superoxide anion was significantly reduced by SODB. No effect of SODB was detected on inflammatory status in any group. The beneficial effect of SODB on cardiac hypertrophy seems to be related to the stimulation of endogenous antioxidant defense, suggesting that SODB may be of interest as a dietary supplementation during conventional antihypertensive therapy.

  16. Superoxide dismutase 1 acts as a nuclear transcription factor to regulate oxidative stress resistance

    PubMed Central

    Tsang, Chi Kwan; Liu, Yuan; Thomas, Janice; Zhang, Yanjie; Zheng, X. F. Steven

    2015-01-01

    Summary Superoxide dismutase 1 (Sod1) has been known for nearly half a century for catalysis of superoxide to hydrogen peroxide. Here we report a new Sod1 function in oxidative signaling: in response to elevated endogenous and exogenous reactive oxygen species (ROS), Sod1 rapidly relocates into the nucleus, which is important for maintaining genomic stability. Interestingly, H2O2 is sufficient to promote Sod1 nuclear localization, indicating that it is responding to general ROS rather than Sod1 substrate superoxide. ROS signaling is mediated by Mec1/ATM and its effector Dun1/Cds1 kinase, through Dun1 interaction with Sod1 and regulation of Sod1 by phosphorylation at S60, 99. In the nucleus, Sod1 binds to the promoters and regulates the expression of oxidative resistance and repair genes. Altogether, our study unravels an unorthodox function of Sod1 as a transcription factor and elucidates the regulatory mechanism for its localization. PMID:24647101

  17. Copper, Zinc Superoxide Dismutase is Primarily a Cytosolic Protein in Human Cells

    NASA Astrophysics Data System (ADS)

    Crapo, James D.; Oury, Tim; Rabouille, Catherine; Slot, Jan W.; Chang, Ling-Yi

    1992-11-01

    The intracellular localization of human copper, zinc superoxide dismutase (Cu,Zn-SOD; superoxide:superoxide oxidoreductase, EC 1.15.1.1) was evaluated by using EM immunocytochemistry and both isolated human cell lines and human tissues. Eight monoclonal antibodies raised against either native or recombinant human Cu,Zn-SOD and two polyclonal antibodies raised against either native or recombinant human Cu,Zn-SOD were used. Fixation with 2% paraformaldehyde/0.2% glutaraldehyde was found necessary to preserve normal distribution of the protein. Monoclonal antibodies were less effective than polyclonal antibodies in recognizing the antigen after adequate fixation of tissue. Cu,Zn-SOD was found widely distributed in the cell cytosol and in the cell nucleus, consistent with it being a soluble cytosolic protein. Mitochondria and secretory compartments did not label for this protein. In human cells, peroxisomes showed a labeling density slightly less than that of cytoplasm.

  18. [Production of superoxide radicals with pulse radiolysis of water with high linear energy transfer].

    PubMed

    Baldacchino, G; Trupin-Wasselin, V; Bouffard, S; Balanzat, E; Gardès-Albert, M; Abedinzadeh, Z; Jore, D; Deycard, S; Hickel, B

    2001-02-01

    The radiolysis of water with heavy ions of high linear energy transfer (LET) (-dE/dx) is characterized, in deaerated medium, by the production of superoxide anions, the radiolytic yields of which increase with the LET. Radiobiological interest in such radical species comes from the oxidative stress which may be generated by their dismutation in O2 and H2O2 in anoxic medium (radiotherapy with heavy ions). A brief review of the measurements of superoxide free radicals in aqueous solution by indirect or direct methods is presented. Moreover, some experimental results obtained by pulse radiolysis with Ar18+ ions (TEL = 290 keV x microm(-1)), are described. The interpretation of the kinetics takes into account the superoxide absorbance and that of hydrogen peroxide, which is present at the millisecond time scale.

  19. Effect of pH on the conversion of superoxide to hydroxyl free radicals

    SciTech Connect

    Baker, M.S.; Gebicki, J.M.

    1984-10-01

    The conversion of superoxide (O-.2) to the hydroxyl (HO.) free radical by superoxide-driven Fenton reactions was measured by the formation of hydroxylated derivatives from benzoate. Among a range of catalysts required for the conversion, the Fe3+EDTA complex was the most effective. The effect of superoxide dismutase and catalase indicated that O-.2 and H2O2 were essential reactants, while the formation of authentic HO. was confirmed by the inhibiting capacities of formate, t-butanol, and mannitol. The conversion of O-.2 to HO. was tested over a broad pH range, and was found to be highest at pH 4.8 whether Fe3+EDTA or free Fe3+ were used as the catalysts. When Fe3+EDTA was used at the optimum pH, every HO. produced required 3.7 O-.2 radicals, close to the theoretical limit of one HO. from every three O-.2 radicals generated.

  20. Characterization of the Bacillus stearothermophilus manganese superoxide dismutase gene and its ability to complement copper/zinc superoxide dismutase deficiency in Saccharomyces cerevisiae.

    PubMed Central

    Bowler, C; Van Kaer, L; Van Camp, W; Van Montagu, M; Inzé, D; Dhaese, P

    1990-01-01

    Recombinant clones containing the manganese superoxide dismutase (MnSOD) gene of Bacillus stearothermophilus were isolated with an oligonucleotide probe designed to match a part of the previously determined amino acid sequence. Complementation analyses, performed by introducing each plasmid into a superoxide dismutase-deficient mutant of Escherichia coli, allowed us to define the region of DNA which encodes the MnSOD structural gene and to identify a promoter region immediately upstream from the gene. These data were subsequently confirmed by DNA sequencing. Since MnSOD is normally restricted to the mitochondria in eucaryotes, we were interested (i) in determining whether B. stearothermophilus MnSOD could function in eucaryotic cytosol and (ii) in determining whether MnSOD could replace the structurally unrelated copper/zinc superoxide dismutase (Cu/ZnSOD) which is normally found there. To test this, the sequence encoding bacterial MnSOD was cloned into a yeast expression vector and subsequently introduced into a Cu/ZnSOD-deficient mutant of the yeast Saccharomyces cerevisiae. Functional expression of the protein was demonstrated, and complementation tests revealed that the protein was able to provide tolerance at wild-type levels to conditions which are normally restrictive for this mutant. Thus, in spite of the evolutionary unrelatedness of these two enzymes, Cu/ZnSOD can be functionally replaced by MnSOD in yeast cytosol. Images FIG. 2 FIG. 4 FIG. 5 PMID:2407726

  1. Manganese superoxide dismutase, but not CuZn superoxide dismutase, is highly expressed in the granulomas of pulmonary sarcoidosis and extrinsic allergic alveolitis.

    PubMed

    Lakari, E; Pääkkö, P; Kinnula, V L

    1998-08-01

    The role of antioxidant defense mechanisms in the pathogenesis of granulomatous human lung diseases remains open to investigation. In this study we investigated the immunoreactivity of two important superoxide radical scavenging intracellular antioxidant enzymes, manganese superoxide dismutase (MnSOD) and copperzinc superoxide dismutase (CuZnSOD), in pulmonary sarcoidosis and extrinsic allergic alveolitis. In histologically normal lung MnSOD was variable but mostly positive in the cells of bronchial epithelium, alveolar epithelium especially in type II pneumocytes, and alveolar macrophages. Copperzinc SOD showed positive immunoreactivity most markedly in the bronchial epithelium. The biopsies of 22 patients with pulmonary sarcoidosis and 10 with extrinsic allergic alveolitis indicated that MnSOD was highly stained in the granulomas of both diseases, with 60 to 100% of the granulomas showing intensive immunoreactivity. Western blots conducted on the cell samples of bronchoalveolar lavage (BAL) fluid revealed significantly higher amounts of MnSOD in sarcoidosis and extrinsic allergic alveolitis than in the controls. Immunohistochemistry on the cells obtained from BAL fluid showed positive immunoreactivity of MnSOD in the macrophages but not in the lymphocytes. In contrast, copperzinc SOD was not induced in either of these diseases. We conclude that MnSOD is highly expressed in the granulomas of pulmonary sarcoidosis and extrinsic allergic alveolitis, and variable but mostly positive in alveolar macrophages, possibly owing to cytokine mediated induction during the granuloma formation.

  2. Oxidative stress induced by loss of Cu,Zn-superoxide dismutase (SOD1) or superoxide-generating herbicides causes axonal degeneration in mouse DRG cultures

    PubMed Central

    Fischer, Lindsey R.

    2014-01-01

    Axonal degeneration is a common pathologic feature in peripheral neuropathy, neurodegenerative disease, and normal aging. Oxidative stress may be an important mechanism of axonal degeneration, but is underrepresented among current experimental models. To test the effects of loss of the antioxidant enzyme Cu,Zn-superoxide dismutase (SOD1) on axon survival, we cultured dorsal root ganglion (DRG) neurons from SOD1 knockout mice. Beginning as early as 48–72 h, we observed striking degeneration of Sod1−/− axons that was prevented by introduction of human SOD1 and was attenuated by antioxidant treatment. To test susceptibility to increased superoxide production, we exposed wild-type DRGs to the redox-cycling herbicides paraquat and diquat (DQ). Dose-dependent axon degeneration was observed, and toxicity of DQ was exacerbated by SOD1 deficiency. MTT staining suggested that DRG axons are more susceptible to injury than their parent cell bodies in both paradigms. Taken together, these data demonstrate susceptibility of DRG axons to oxidative stress-mediated injury due to loss of SOD1 or excess superoxide production. These in vitro models provide a novel means of investigating oxidative stress-mediated injury to axons, to improve our understanding of axonal redox control and dysfunction in peripheral neuropathy. PMID:20039174

  3. Mitochondrial generation of superoxide and hydrogen peroxide as the source of mitochondrial redox signaling.

    PubMed

    Brand, Martin D

    2016-11-01

    This review examines the generation of reactive oxygen species by mammalian mitochondria, and the status of different sites of production in redox signaling and pathology. Eleven distinct mitochondrial sites associated with substrate oxidation and oxidative phosphorylation leak electrons to oxygen to produce superoxide or hydrogen peroxide: oxoacid dehydrogenase complexes that feed electrons to NAD(+); respiratory complexes I and III, and dehydrogenases, including complex II, that use ubiquinone as acceptor. The topologies, capacities, and substrate dependences of each site have recently clarified. Complex III and mitochondrial glycerol 3-phosphate dehydrogenase generate superoxide to the external side of the mitochondrial inner membrane as well as the matrix, the other sites generate superoxide and/or hydrogen peroxide exclusively in the matrix. These different site-specific topologies are important for redox signaling. The net rate of superoxide or hydrogen peroxide generation depends on the substrates present and the antioxidant systems active in the matrix and cytosol. The rate at each site can now be measured in complex substrate mixtures. In skeletal muscle mitochondria in media mimicking muscle cytosol at rest, four sites dominate, two in complex I and one each in complexes II and III. Specific suppressors of two sites have been identified, the outer ubiquinone-binding site in complex III (site IIIQo) and the site in complex I active during reverse electron transport (site IQ). These suppressors prevent superoxide/hydrogen peroxide production from a specific site without affecting oxidative phosphorylation, making them excellent tools to investigate the status of the sites in redox signaling, and to suppress the sites to prevent pathologies. They allow the cellular roles of mitochondrial superoxide/hydrogen peroxide production to be investigated without catastrophic confounding bioenergetic effects. They show that sites IIIQo and IQ are active in cells and

  4. Hyperglycemic switch from mitochondrial nitric oxide to superoxide production in endothelial cells.

    PubMed

    Brodsky, Sergey V; Gao, Shujuan; Li, Hong; Goligorsky, Michael S

    2002-11-01

    The accumulated ultrastructural and biochemical evidence is highly suggestive of the existence of mitochondrial nitric oxide (NO) synthase (mtNOS), where local production of NO regulates the electron transport along the respiratory chain. Here, the functional competence of mtNOS in situ in a living cell was examined using an intravital fluorescent NO indicator, 4,5-diaminofluorescein, employing a new procedure for loading it into the mitochondria to demonstrate local NO generation in undisrupted endothelial cells and in isolated mitochondria as well as in human embryonic kidney cells stably expressing endothelial NOS. With the use of this approach, we showed that endothelial cells incubated in the presence of high concentration of D-glucose (but not L-glucose) are characterized by the reduced NO synthetic function of mitochondria despite the unaltered abundance of the enzyme. In parallel, mitochondrial generation of superoxide was augmented in endothelial cells incubated in the presence of a high concentration of D-glucose. Both the NO generation and superoxide production in hyperglycemic environment could be restored to control levels by treating cells with a cell-permeable superoxide dismutase mimetic. In addition, enhanced mitochondrial superoxide production could be suppressed with an inhibitor of NOS in stimulated endothelial cells. In conclusion, the data 1) provide direct evidence of mitochondrial NO production in endothelial cells, 2) demonstrate its suppression and enhanced superoxide generation in hyperglycemic environment, and 3) provide evidence that "uncoupled" mtNOS represents an important source of superoxide anions in endothelial cells incubated in high glucose-containing medium.

  5. Production of superoxide/H2O2 by dihydroorotate dehydrogenase in rat skeletal muscle mitochondria.

    PubMed

    Hey-Mogensen, Martin; Goncalves, Renata L S; Orr, Adam L; Brand, Martin D

    2014-07-01

    Dehydrogenases that use ubiquinone as an electron acceptor, including complex I of the respiratory chain, complex II, and glycerol-3-phosphate dehydrogenase, are known to be direct generators of superoxide and/or H2O2. Dihydroorotate dehydrogenase oxidizes dihydroorotate to orotate and reduces ubiquinone to ubiquinol during pyrimidine metabolism, but it is unclear whether it produces superoxide and/or H2O2 directly or does so only indirectly from other sites in the electron transport chain. Using mitochondria isolated from rat skeletal muscle we establish that dihydroorotate oxidation leads to superoxide/H2O2 production at a fairly high rate of about 300pmol H2O2·min(-1)·mg protein(-1) when oxidation of ubiquinol is prevented and complex II is uninhibited. This H2O2 production is abolished by brequinar or leflunomide, known inhibitors of dihydroorotate dehydrogenase. Eighty percent of this rate is indirect, originating from site IIF of complex II, because it can be prevented by malonate or atpenin A5, inhibitors of complex II. In the presence of inhibitors of all known sites of superoxide/H2O2 production (rotenone to inhibit sites in complex I (site IQ and, indirectly, site IF), myxothiazol to inhibit site IIIQo in complex III, and malonate plus atpenin A5 to inhibit site IIF in complex II), dihydroorotate dehydrogenase generates superoxide/H2O2, at a small but significant rate (23pmol H2O2·min(-1)·mg protein(-1)), from the ubiquinone-binding site. We conclude that dihydroorotate dehydrogenase can generate superoxide and/or H2O2 directly at low rates and is also capable of indirect production at higher rates from other sites through its ability to reduce the ubiquinone pool.

  6. Production of superoxide/hydrogen peroxide by the mitochondrial 2-oxoadipate dehydrogenase complex.

    PubMed

    Goncalves, Renata L S; Bunik, Victoria I; Brand, Martin D

    2016-02-01

    In humans, mutations in dehydrogenase E1 and transketolase domain containing 1 (DHTKD1) are associated with neurological abnormalities and accumulation of 2-oxoadipate, 2-aminoadipate, and reactive oxygen species. The protein encoded by DHTKD1 has sequence and structural similarities to 2-oxoglutarate dehydrogenase, and the 2-oxoglutarate dehydrogenase complex can produce superoxide/H2O2 at high rates. The DHTKD1 enzyme is hypothesized to catalyze the oxidative decarboxylation of 2-oxoadipate, a shared intermediate of the degradative pathways for tryptophan, lysine and hydroxylysine. Here, we show that rat skeletal muscle mitochondria can produce superoxide/H2O2 at high rates when given 2-oxoadipate. We identify the putative mitochondrial 2-oxoadipate dehydrogenase complex as one of the sources and characterize the conditions that favor its superoxide/H2O2 production. Rates increased at higher NAD(P)H/NAD(P)(+) ratios and were higher at each NAD(P)H/NAD(P)(+) ratio when 2-oxoadipate was present, showing that superoxide/H2O2 was produced during the forward reaction from 2-oxoadipate, but not in the reverse reaction from NADH in the absence of 2-oxoadipate. The maximum capacity of the 2-oxoadipate dehydrogenase complex for production of superoxide/H2O2 is comparable to that of site IF of complex I, and seven, four and almost two-fold lower than the capacities of the 2-oxoglutarate, pyruvate and branched-chain 2-oxoacid dehydrogenase complexes, respectively. Regulation by ADP and ATP of H2O2 production driven by 2-oxoadipate was very different from that driven by 2-oxoglutarate, suggesting that site AF of the 2-oxoadipate dehydrogenase complex is a new source of superoxide/H2O2 associated with the NADH isopotential pool in mitochondria.

  7. Elevated peripheral blood mononuclear cell-derived superoxide production in healthy young black men.

    PubMed

    Deo, Shekhar H; Holwerda, Seth W; Keller, David M; Fadel, Paul J

    2015-03-01

    Several studies have demonstrated that blacks exhibit elevations in systemic oxidative stress. However, the source(s) and mechanism(s) contributing to the elevation in oxidative stress remain unclear. Given that peripheral blood mononuclear cells (PBMCs) can be a major source of NADPH oxidase-derived superoxide production, we tested the hypothesis that young black men demonstrate greater superoxide production and NADPH oxidase expression in PBMCs compared with whites. PBMCs were freshly isolated from whole blood in young normotensive black (n = 18) and white (n = 16) men. Intracellular superoxide production in PBMCs was measured using dihydroethidium fluorescence, protein expression of NADPH oxidase subunits, gp91(phox) (membranous) and p47(phox) (cytosolic) in PBMCs were assessed using Western blot analysis, and plasma protein carbonyls were measured as a marker of systemic oxidative stress. Black men showed elevated intracellular superoxide production (4.3 ± 0.5 vs. 2.0 ± 0.6 relative fluorescence units; black men vs. white men, P < 0.05), increased protein expression for gp91(phox) and p47(phox) (e.g., p47(phox): 1.1 ± 0.2, black men vs. 0.4 ± 0.1, white men, P < 0.05) in PBMCs and higher circulating protein carbonyl levels (22 ± 4 vs. 14 ± 2 nmol/ml; black men vs. white men, P < 0.05). Interestingly, a positive family history of hypertension in black men did not further enhance PBMC-derived intracellular superoxide production or NADPH oxidase subunit protein expression. These findings indicate that black men exhibit greater resting PBMC-derived superoxide production and an upregulation of the NADPH oxidase pathway with a possible contribution to increases in systemic oxidative stress.

  8. Effect of superoxide derived from lucifer yellow CH on voltage-gated currents of mouse taste bud cells.

    PubMed

    Takeuchi, Keita; Yoshii, Kiyonori

    2008-06-01

    Lucifer yellow CH (LY), a fluorescent membrane-impermeable cell marker dye, has been routinely loaded into cells through recording electrodes to visualize these cells after electrophysiological investigation, without considering its pharmacological effect. Recently, we showed that the exposure of cells loaded with LY to light for microscopy produced unidentified radical species that retarded the inactivation of voltage-gated Na+ currents irreversibly (Higure Y et al. 2003). Here, we show that superoxide dismutase, an enzyme that decomposes superoxide, reverses the retardation effect, which assures that superoxide is the unidentified radical species. The estimated mean lifetime of superoxide in recording electrodes (in the absence of cytoplasm) is approximately 6 min, and hence, the Na+ currents are retarded even in the dark, when LY is exposed to light before being loaded into the cell. Superoxide has no effect on voltage-gated Cl- currents. These results show that superoxide action on ion channels is rather selective. The breakdown of superoxide inside cells and the effect of endogenous superoxide on the superoxide-susceptible channels are discussed.

  9. The SoxRS response of Escherichia coli is directly activated by redox-cycling drugs rather than by superoxide

    PubMed Central

    Gu, Mianzhi; Imlay, James A.

    2011-01-01

    When Escherichia coli is exposed to redox-cycling drugs, its SoxR transcription factor is activated by oxidation of its [2Fe–2S] cluster. In aerobic cells these drugs generate superoxide, and because superoxide dismutase (SOD) is a member of the SoxRS regulon, superoxide was initially thought to be the activator of SoxR. Its many-gene regulon was therefore believed to comprise a defense against superoxide stress. However, we found that abundant superoxide did not effectively activate SoxR in an SOD− mutant, that overproduced SOD could not suppress activation by redox-cycling drugs, and that redox-cycling drugs were able to activate SoxR in anaerobic cells as long as alternative respiratory acceptors were provided. Thus superoxide is not the signal that SoxR senses. Indeed, redox-cycling drugs directly oxidized the cluster of purified SoxR in vitro, while superoxide did not. Redox-cycling drugs are excreted by both bacteria and plants. Their toxicity does not require superoxide, as they poisoned E. coli under anaerobic conditions, in part by oxidizing dehydratase iron-sulfur clusters. Under these conditions SoxRS induction was protective. Thus it is physiologically appropriate that the SoxR protein directly senses redox-cycling drugs rather than superoxide. PMID:21226770

  10. Distribution of Iron-Containing Superoxide Dismutase in Vascular Plants 12

    PubMed Central

    Bridges, Susan M.; Salin, Marvin L.

    1981-01-01

    Superoxide dismutases (EC 1.15.1.1) in vascular plants representing different evolutionary levels were characterized using polyacrylamide gel electrophoresis. The three forms of the enzyme were distinguished from each other based on the following criteria: a) the Cu-Zn enzyme is sensitive to cyanide wherease the Fe and Mn enzymes are not; and b) the Cu-Zn and Fe enzymes are inhibited by H2O2 whereas the Mn enzyme is H2O2-resistant. Of the 43 plant families investigated, the Fe-containing superoxide dismutase was found in three families: Gingkoaceae, Nymphaceae, and Cruciferae. PMID:16661901

  11. Superoxide dismutase transgenes in sugarbeets confer resistance to oxidative agents and the fungus C. beticola.

    PubMed

    Tertivanidis, Konstantinos; Goudoula, Catherine; Vasilikiotis, Christos; Hassiotou, Efthymia; Perl-Treves, Rafael; Tsaftaris, Athanasios

    2004-06-01

    Sugarbeets carrying superoxide dismutase transgenes were developed in order to investigate the possibility of enhancing their resistance to oxidative stress. Binary T-DNA vectors carrying the chloroplastic and cytosolic superoxide dismutase genes from tomato, were used for Agrobacterium-mediated transformation of sugarbeet petioles. The transgenic plants were subjected to treatments known to cause oxidative stress, such as the herbicide methyl viologen and a natural photosensitizer toxin produced by the fungus Cercospora beticola, namely cercosporin. The transgenic plants exhibited increased tolerance to methyl viologen, to pure cercosporin, as well as to leaf infection with the fungus C. beticola.

  12. The Single Superoxide Dismutase of Rhodobacter capsulatus Is a Cambialistic, Manganese-Containing Enzyme

    PubMed Central

    Tabares, Leandro C.; Bittel, Cristian; Carrillo, Néstor; Bortolotti, Ana; Cortez, Néstor

    2003-01-01

    The phototrophic bacterium Rhodobacter capsulatus contains a single, oxygen-responsive superoxide dismutase (SODRc) homologous to iron-containing superoxide dismutase enzymes. Recombinant SODRc, however, displayed higher activity after refolding with Mn2+, especially when the pH of the assay mixture was raised. SODRc isolated from Rhodobacter cells also preferentially contains manganese, but metal discrimination depends on the culture conditions, with iron fractions increasing from 7% in aerobic cultures up to 40% in photosynthetic cultures. Therefore, SODRc behaves as a Mn-containing dismutase with cambialistic properties. PMID:12730184

  13. Effect of superoxide anion scavenger on rat hearts with chronic intermittent hypoxia.

    PubMed

    Pai, Peiying; Lai, Ching Jung; Lin, Ching-Yuang; Liou, Yi-Fan; Huang, Chih-Yang; Lee, Shin-Da

    2016-04-15

    Only very limited information regarding the protective effects of the superoxide anion scavenger on chronic intermittent hypoxia-induced cardiac apoptosis is available. The purpose of this study is to evaluate the effects of the superoxide anion scavenger on cardiac apoptotic and prosurvival pathways in rats with sleep apnea. Forty-two Sprague-Dawley rats were divided into three groups, rats with normoxic exposure (Control, 21% O2, 1 mo), rats with chronic intermittent hypoxia exposure (Hypoxia, 3-7% O2vs. 21% O2per 40 s cycle, 8 h per day, 1 mo), and rats with pretreatment of the superoxide anion scavenger and chronic intermittent hypoxia exposure (Hypoxia-O2 (-)-Scavenger, MnTMPyP pentachloride, 1 mg/kg ip per day; 3-7% O2vs. 21% O2per 40 s cycle, 8 h per day, 1 mo) at 5-6 mo of age. After 1 mo, the protein levels and apoptotic cells of excised hearts from three groups were measured by Western blotting and terminal deoxynucleotide transferase-mediated dUTP nick end labeling (TUNEL) assay. The superoxide anion scavenger decreased hypoxia-induced myocardial architecture abnormalities, left ventricular hypertrophy, and TUNEL-positive apoptosis. The superoxide anion scavenger decreased hypoxia-induced Fas ligand, Fas death receptors, Fas-associated death domain (FADD), activated caspase-8, and activated caspase-3 (Fas-dependent apoptotic pathway) as well as Bad, activated caspase-9 and activated caspase-3 (mitochondria-dependent apoptotic pathway), endonuclease G (EndoG), apoptosis-inducing factor (AIF), and TUNEL-positive apoptosis. The superoxide anion scavenger increased IGF-1, IGF-1R, p-PI3k, p-Akt, p-Bad, Bcl-2, and Bcl-xL (survival pathway). Our findings imply that the superoxide anion scavenger might prevent cardiac Fas-mediated and mitochondrial-mediated apoptosis and enhance the IGF-1-related survival pathway in chronic intermittent hypoxia. The superoxide anion scavenger may prevent chronic sleep apnea-enhanced cardiac apoptotic pathways and enhances

  14. Evidence that the reactivity of the martian soil is due to superoxide ions

    NASA Technical Reports Server (NTRS)

    Yen, A. S.; Kim, S. S.; Hecht, M. H.; Frant, M. S.; Murray, B.

    2000-01-01

    The Viking Landers were unable to detect evidence of life on Mars but, instead, found a chemically reactive soil capable of decomposing organic molecules. This reactivity was attributed to the presence of one or more as-yet-unidentified inorganic superoxides or peroxides in the martian soil. Using electron paramagnetic resonance spectroscopy, we show that superoxide radical ions (O2-) form directly on Mars-analog mineral surfaces exposed to ultraviolet radiation under a simulated martian atmosphere. These oxygen radicals can explain the reactive nature of the soil and the apparent absence of organic material at the martian surface.

  15. Evidence that the reactivity of the martian soil is due to superoxide ions.

    PubMed

    Yen, A S; Kim, S S; Hecht, M H; Frant, M S; Murray, B

    2000-09-15

    The Viking Landers were unable to detect evidence of life on Mars but, instead, found a chemically reactive soil capable of decomposing organic molecules. This reactivity was attributed to the presence of one or more as-yet-unidentified inorganic superoxides or peroxides in the martian soil. Using electron paramagnetic resonance spectroscopy, we show that superoxide radical ions (O2-) form directly on Mars-analog mineral surfaces exposed to ultraviolet radiation under a simulated martian atmosphere. These oxygen radicals can explain the reactive nature of the soil and the apparent absence of organic material at the martian surface.

  16. Construction of a Fusion Enzyme Exhibiting Superoxide Dismutase and Peroxidase Activity.

    PubMed

    Sharapov, M G; Novoselov, V I; Ravin, V K

    2016-04-01

    A chimeric gene construct encoding human peroxiredoxin 6 and Mn-superoxide dismutase from Escherichia coli was developed. Conditions for expression of the fusion protein in E. coli cell were optimized. Fusing of the enzymes into a single polypeptide chain with peroxiredoxin 6 at the N-terminus (PSH) did not affect their activities. On the contrary, the chimeric protein with reverse order of enzymes (SPH) was not obtained in a water-soluble active form. The active chimeric protein (PSH) exhibiting both peroxidase and superoxide dismutase activities was prepared and its physicochemical properties were characterized.

  17. Extraction of erythrocyte enzymes for the preparation of polyhemoglobin-catalase-superoxide dismutase.

    PubMed

    Gu, Jingsong; Chang, Thomas Ming Swi

    2009-01-01

    In sustained severe ischemia, reperfusion with oxygen carriers may result in ischemia-reperfusion injuries because of the release of damaging oxygen radicals. A nanobiotechnology-based polyhemogloin-calatase-superoxide dismutase can prevent this because the oxygen carrier, polyhemoglobin, is linked to antioxidant enzymes, catalase and superoxide dismutase. However, these antioxidant enzymes come from nonhuman sources and recombinant human enzymes are expensive. This paper describes our study on extracting these enzymes from red blood cells and analyzing the amount of enzymes needed for adequate protection from ischemia-reperfusion.

  18. Serine 1179 Phosphorylation of Endothelial Nitric Oxide Synthase Increases Superoxide Generation and Alters Cofactor Regulation.

    PubMed

    Peng, Hu; Zhuang, Yugang; Harbeck, Mark C; He, Donghong; Xie, Lishi; Chen, Weiguo

    2015-01-01

    Endothelial nitric oxide synthase (eNOS) is responsible for maintaining systemic blood pressure, vascular remodeling and angiogenesis. In addition to producing NO, eNOS can also generate superoxide (O2-.) in the absence of the cofactor tetrahydrobiopterin (BH4). Previous studies have shown that bovine eNOS serine 1179 (Serine 1177/human) phosphorylation critically modulates NO synthesis. However, the effect of serine 1179 phosphorylation on eNOS superoxide generation is unknown. Here, we used the phosphomimetic form of eNOS (S1179D) to determine the effect of S1179 phosphorylation on superoxide generating activity, and its sensitivity to regulation by BH4, Ca2+, and calmodulin (CAM). S1179D eNOS exhibited significantly increased superoxide generating activity and NADPH consumption compared to wild-type eNOS (WT eNOS). The superoxide generating activities of S1179D eNOS and WT eNOS did not differ significantly in their sensitivity to regulation by either Ca2+ or CaM. The sensitivity of the superoxide generating activity of S1179D eNOS to inhibition by BH4 was significantly reduced compared to WT eNOS. In eNOS-overexpressing 293 cells, BH4 depletion with 10mM DAHP for 48 hours followed by 50ng/ml VEGF for 30 min to phosphorylate eNOS S1179 increased ROS accumulation compared to DAHP-only treated cells. Meanwhile, MTT assay indicated that overexpression of eNOS in HEK293 cells decreased cellular viability compared to control cells at BH4 depletion condition (P<0.01). VEGF-mediated Serine 1179 phosphorylation further decreased the cellular viability in eNOS-overexpressing 293 cells (P<0.01). Our data demonstrate that eNOS serine 1179 phosphorylation, in addition to enhancing NO production, also profoundly affects superoxide generation: S1179 phosphorylation increases superoxide production while decreasing sensitivity to the inhibitory effect of BH4 on this activity.

  19. Evidence that the reactivity of the martian soil is due to superoxide ions

    NASA Technical Reports Server (NTRS)

    Yen, A. S.; Kim, S. S.; Hecht, M. H.; Frant, M. S.; Murray, B.

    2000-01-01

    The Viking Landers were unable to detect evidence of life on Mars but, instead, found a chemically reactive soil capable of decomposing organic molecules. This reactivity was attributed to the presence of one or more as-yet-unidentified inorganic superoxides or peroxides in the martian soil. Using electron paramagnetic resonance spectroscopy, we show that superoxide radical ions (O2-) form directly on Mars-analog mineral surfaces exposed to ultraviolet radiation under a simulated martian atmosphere. These oxygen radicals can explain the reactive nature of the soil and the apparent absence of organic material at the martian surface.

  20. Parasitization by Scleroderma guani influences expression of superoxide dismutase genes in Tenebrio molitor

    USDA-ARS?s Scientific Manuscript database

    Superoxide dismutase (SOD) is an antioxidant enzyme involved in detoxifying reactive oxygen species. In this study, we identified genes encoding the extracellular and intracellular copper-zinc SODs (ecCuZnSOD and icCuZnSOD) and a manganese SOD (MnSOD) in the yellow mealworm beetle, Tenebrio molitor....

  1. Exogenous superoxide dismutase may lose its antidotal ability on rice leaves

    USDA-ARS?s Scientific Manuscript database

    Leaf diffusates of the resistant rice cultivars suppressed spore germination of blast fungus (Magnaporthe grisea). Bovine Cu-Zn superoxide dismutase (SOD) added to the diffusate abolished its toxicity. However, the enzyme added to the inoculum did not affect the toxicity of the diffusate. Even the s...

  2. Periplasmic superoxide dismutase SodCI of Salmonella binds peptidoglycan to remain tethered within the periplasm.

    PubMed

    Tidhar, Avital; Rushing, Marcus D; Kim, Byoungkwan; Slauch, James M

    2015-09-01

    Salmonellae survive and propagate in macrophages to cause serious systemic disease. Periplasmic superoxide dismutase plays a critical role in this survival by combating phagocytic superoxide. Salmonella Typhimurium strain 14028 produces two periplasmic superoxide dismutases: SodCI and SodCII. Although both proteins are produced during infection, only SodCI is functional in the macrophage phagosome. We have previously shown that SodCI, relative to SodCII, is both protease resistant and tethered within the periplasm and that either of these properties is sufficient to allow a SodC to protect against phagocytic superoxide. Tethering is defined as remaining cell-associated after osmotic shock or treatment with cationic antimicrobial peptides. Here we show that SodCI non-covalently binds peptidoglycan. SodCI binds to Salmonella and Bacillus peptidoglycan, but not peptidoglycan from Staphylococcus. Moreover, binding can be inhibited by a diaminopimelic acid containing tripeptide, but not a lysine containing tripeptide, showing that the protein recognizes the peptide portion of the peptidoglycan. Replacing nine amino acids in SodCII with the corresponding residues from SodCI confers tethering, partially delineating an apparently novel peptidoglycan binding domain. These changes in sequence increase the affinity of SodCII for peptidoglycan fragments to match that of SodCI and allow the now tethered SodCII to function during infection.

  3. Superoxide production is inversely related to complex I activity in inherited complex I deficiency.

    PubMed

    Verkaart, Sjoerd; Koopman, Werner J H; van Emst-de Vries, Sjenet E; Nijtmans, Leo G J; van den Heuvel, Lambertus W P J; Smeitink, Jan A M; Willems, Peter H G M

    2007-03-01

    Deficiency of NADH:ubiquinone oxidoreductase or complex I (CI) is the most common cause of disorders of the oxidative phosphorylation system in humans. Using life cell imaging and blue-native electrophoresis we quantitatively compared superoxide production and CI amount and activity in cultured skin fibroblasts of 7 healthy control subjects and 21 children with inherited isolated CI deficiency. Thirteen children had a disease causing mutation in one of the nuclear-encoded CI subunits, whereas in the remainder the genetic cause of the disease is not yet established. Superoxide production was significantly increased in all but two of the patient cell lines. An inverse relationship with the amount and residual activity of CI was observed. In agreement with this finding, rotenone, a potent inhibitor of CI activity, dose-dependently increased superoxide production in healthy control cells. Also in this case an inverse relationship with the residual activity of CI was observed. In sharp contrast, however, rotenone did not decrease the amount of CI. The data presented show that superoxide production is increased in inherited CI deficiency and that this increase is primarily a consequence of the reduction in cellular CI activity and not of a further leakage of electrons from mutationally malformed complexes.

  4. Differential response of maize catalases and superoxide dismutases to the photoactivated fungal toxin cercosporin.

    PubMed

    Williamson, J D; Scandalios, J G

    1992-05-01

    Many fungi of the genus Cercospora produce a light-induced, photoactivated polyketide toxin called cercosporin. In the presence of light an excited form (triplet state) of the toxin molecule is produced which, depending on the reducing potential of the environment, reacts with molecular oxygen to produce singlet oxygen and/or superoxide radicals. In this paper a system is presented for analysis of antioxidant defense gene response using purified cercosporin under conditions demonstrated to favor superoxide formation. Under the assay conditions employed, changes in total catalase activity, as well as individual isozyme protein levels generally mirrored the changes observed in corresponding steady-state RNA levels in response to applied cercosporin. In contrast, while transcript accumulation for most maize superoxide dismutases increased dramatically, both total superoxide dismutase activity and individual isozyme protein levels remained constant in all toxin treatments. In one case, the analyses indicated that there are two distinct transcripts that hybridize with a gene-specific probe for Sod3. These two transcripts responded differentially to applied toxin (levels of the larger transcript increased while the smaller decreased), whereas corresponding steady-state levels for the SOD-3 isozyme proteins remained constant. This suggests that protein turnover might play a role in the response of these SODs to activated oxygen species.

  5. Role of superoxide in the photochemical reduction of iron in seawater

    NASA Astrophysics Data System (ADS)

    Rose, Andrew L.; Waite, T. David

    2006-08-01

    We have conducted a series of laboratory studies to investigate the generation of ferrous iron and reactive oxygen species when solutions of seawater containing natural organic matter (NOM) and ferric iron are exposed to simulated sunlight. Total ferrous iron and hydrogen peroxide were measured at nanomolar concentrations with high temporal resolution using chemiluminescence-based methods. In all cases, ferrous iron concentrations rapidly peaked at several nanomoles per litre after a few minutes, and then declined over time, while hydrogen peroxide concentrations increased in a non-linear manner. Although concentrations of both species depended on the concentration of NOM, hydrogen peroxide concentrations were only minimally affected by the presence of iron. Increasing the NOM concentration while the total iron concentration was maintained constant led to an increase in the maximum ferrous iron concentration, suggesting that superoxide-mediated reduction of iron may be a major pathway for ferrous iron formation. This was supported by measurements of superoxide production from irradiation of NOM in the absence of iron and kinetic calculations, as well as an experiment in which superoxide dismutase was added. Further analysis of the data suggested that dissolved oxygen and photo-produced hydrogen peroxide were the primary oxidants of the Fe(II) formed. Thus we propose that superoxide and ferrous iron may be intricately coupled in the system, and that their generation is determined by the supply of NOM available to harvest light and donate electrons.

  6. Copper complexes of 1,10-phenanthroline and related compounds as superoxide dismutase mimetics.

    PubMed

    Bijloo, G J; van der Goot, H; Bast, A; Timmerman, H

    1990-11-01

    In a preliminary study we tested CuSO4.5H2O, (Cu(II]2[3,5-diisopropylsalicylate]4.2H2O and a number of copper complexes of substituted 1,10-phenanthrolines for superoxide anion dismutase activity. It appeared that this activity depends on the ligands involved and might be governed by the redox potential of the Cu(I) complex/Cu(II) complex couple. The strong superoxide anion dismutase activity of Cu(II)[DMP]2 complex can be expected considering its high redox potential. Rather surprisingly is the superoxide anion dismutase activity of the Cu(I)[DMP]2 complex since it involves oxidation to Cu(II)[DMP]2 complex. From regression analysis it was established that steric and field effects of the substituents of the investigated phenanthrolines play an important role in SOD activity and therefore it is concluded that complex formation is important for the superoxide dismutase-like activity.

  7. NADH induces the generation of superoxide radicals in leaf peroxisomes. [Pisum sativum L

    SciTech Connect

    del Rio, L.A.; Sandalio, L.M.; Palma, J.M. ); Fernandez, V.M.; Ruperez, F.L. )

    1989-03-01

    In peroxisomes isolated from pea leaves (Pisum sativum L.) the production of superoxide free radicals (O{sub 2}{sup {minus}}) by xanthine and NADH was investigated. In peroxisomal membranes, 100 micromolar NADH induced the production of O{sub 2}{sup {minus}} radicals. In the soluble fractions of peroxisomes, no generation of O{sub 2}{sup {minus}} radicals was observed by incubation with either NADH or xanthine, although xanthine oxidase was found located predominantly in the matrix of peroxisomes. The failure of xanthine to induce superoxide generation was probably due to the inability to fully suppress the endogenous Mn-superoxide dismutase activity by inhibitors which were inactive against xanthine oxidase. The generation of superoxide radicals in leaf peroxisomes together with the recently described production of these oxygen radicals in glyoxysomes suggests that O{sub 2}{sup {minus}} generation could be a common metabolic property of peroxisomes and further supports the existence of active oxygen-related roles for peroxisomes in cellular metabolism.

  8. Superoxide dismutase expression and oxidative damage in a case of myopathy in brown pelicans (Pelecanus occidentalis).

    PubMed

    Giri, Dipak K; Miller, Debra L; Thompson, Larry J; Mailler, Lesley; Styer, Eloise; Baldwin, Charles

    2007-05-01

    Four brown pelicans (Pelecanus occidentalis) housed at a rehabilitation facility were found dead after a 3-day history of muscle weakness and after being fed for about 2 weeks from a recent shipment of fish. The birds had pale streaking of the skeletal and heart muscles. Microscopically, the skeletal muscle, and to a lesser extent the cardiac muscle, had severe myocyte degeneration and necrosis characterized by microvacuolation with loss of cross-striations, condensation of cytoplasm, fragmentation, mineralization, and inflammatory cell infiltrates consisting of multinucleated cells, macrophages, and few heterophils. The findings were consistent with myopathy, and a nutritional myopathy caused by eating rancid fish was suspected. Immunohistochemical staining revealed abundant immunoreactive copper zinc superoxide dismutase and manganese superoxide dismutase either as diffuse homogeneous precipitates or granular aggregates in the cytoplasm of affected cells. Immunoreactivity was directly related to degree of cellular damage as estimated by light microscopic examination. We suggest that the lack of protection, despite upregulation of superoxide dismutase, is most likely attributable to supersaturation of oxidants beyond the capacity of superoxide dismutases to scavenge.

  9. [Plasma and red blood cell superoxide dismutase activity in patients with different stages of essential hypertension].

    PubMed

    Simić, Dragan; Perunicić, Jovan; Lasica, Ratko; Ivanović, Branislava; Matić, Danica; Kalimanovska-Ostrić, Dimitra; Vranić, Ivana; Medenica, Miroslav; Mimić-Oka, Jasmina; Simić, Tatjana

    2005-01-01

    It has been suggested that superoxide dismutase (SOD) plays an important role in endothelial dysfunction in essential hypertension (EH), by competing with nitric oxide for superoxide, thus influencing nitric oxide bioavailability. To answer the question of whether endothelial dysfunction is consequence of altered SOD expression we determined SOD activity in patients with different stages of EH. In this study 45 EH patients and 25 normotensive subjects were included. EH patients were divided into the three groups according to the guidelines of European Society of Hypertension. SOD activity was determined spectrophotometrically in RBC and plasma of EH patients and controls. The results obtained have shown that all groups of EH patients exhibit lower SOD activity than control normotensive subjects. Significant correlation between SOD activity and both diastolic (p<0.05, r=-0.394) and systolic blood pressure (p<0.05, r=-0.356) was found. Lowering of SOD activity in patients with different stages of EH leads to inefficient detoxification of superoxide in EH. An excess of superoxide of both cellular and extracellular origin takes part in enhanced degradation of nitric oxide and altered vasodilation, and consequent endothelial dysfunction.

  10. Novel reversible and selective nerve agent simulant detection in conjunction with superoxide "turn-on" probing.

    PubMed

    Jang, Yoon Jeong; Murale, Dhiraj P; Churchill, David G

    2014-04-07

    Herein, we present fluorescein as a reversible fluorescent sensor for nerve agent simulants diethylchlorophosphate (DCP), diethyl methylphosphonate (DEMP), and diethyl cyanophosphonate (DECP). The superoxide allows for an "off-on" mechanism to regenerate fluorescein. The order of decrease in fluorescence intensity for nerve agent simulants is DCP > DEMP ≫ DECP.

  11. Titanium Dioxide Nanoparticles Increase Superoxide Anion Production by Acting on NADPH Oxidase.

    PubMed

    Masoud, Rawand; Bizouarn, Tania; Trepout, Sylvain; Wien, Frank; Baciou, Laura; Marco, Sergio; Houée Levin, Chantal

    2015-01-01

    Titanium dioxide (TiO2) anatase nanoparticles (NPs) are metal oxide NPs commercialized for several uses of everyday life. However their toxicity has been poorly investigated. Cellular internalization of NPs has been shown to activate macrophages and neutrophils that contribute to superoxide anion production by the NADPH oxidase complex. Transmission electron micrososcopy images showed that the membrane fractions were close to the NPs while fluorescence indicated an interaction between NPs and cytosolic proteins. Using a cell-free system, we have investigated the influence of TiO2 NPs on the behavior of the NADPH oxidase. In the absence of the classical activator molecules of the enzyme (arachidonic acid) but in the presence of TiO2 NPs, no production of superoxide ions could be detected indicating that TiO2 NPs were unable to activate by themselves the complex. However once the NADPH oxidase was activated (i.e., by arachidonic acid), the rate of superoxide anion production went up to 140% of its value without NPs, this effect being dependent on their concentration. In the presence of TiO2 nanoparticles, the NADPH oxidase produces more superoxide ions, hence induces higher oxidative stress. This hyper-activation and the subsequent increase in ROS production by TiO2 NPs could participate to the oxidative stress development.

  12. Copper-Zinc Superoxide Dismutase: A Unique Biological "Ligand" for Bioinorganic Studies.

    ERIC Educational Resources Information Center

    Valentine, Joan Selverstone; de Freitas, Duarte Mota

    1985-01-01

    Discusses superoxide dismutase (SOD) research and the properties of copper, zinc (Cu, Zn)-SOD. Emphasizes the controversy concerning the role of Cu,Zn-SOD and other SOD enzymes as protective agents in reactions involving dioxygen metabolism, and the properties of Cu, Zn-SOD that make it an interesting biological ligand for physical studies of…

  13. Hydrogen peroxide induce modifications of human extracellular superoxide dismutase that results in enzyme inhibition.

    PubMed

    Gottfredsen, Randi H; Larsen, Ulrike G; Enghild, Jan J; Petersen, Steen V

    2013-01-01

    Superoxide dismutase (EC-SOD) controls the level of superoxide in the extracellular space by catalyzing the dismutation of superoxide into hydrogen peroxide and molecular oxygen. In addition, the enzyme reacts with hydrogen peroxide in a peroxidase reaction which is known to disrupt enzymatic activity. Here, we show that the peroxidase reaction supports a site-specific bond cleavage. Analyses by peptide mapping and mass spectrometry shows that oxidation of Pro112 supports the cleavage of the Pro112-His113 peptide bond. Substitution of Ala for Pro112 did not inhibit fragmentation, indicating that the oxidative fragmentation at this position is dictated by spatial organization and not by side-chain specificity. The major part of EC-SOD inhibited by the peroxidase reaction was not fragmented but found to encompass oxidations of histidine residues involved in the coordination of copper (His98 and His163). These oxidations are likely to support the dissociation of copper from the active site and thus loss of enzymatic activity. Homologous modifications have also been described for the intracellular isozyme, Cu/Zn-SOD, reflecting the almost identical structures of the active site within these enzymes. We speculate that the inactivation of EC-SOD by peroxidase activity plays a role in regulating SOD activity in vivo, as even low levels of superoxide will allow for the peroxidase reaction to occur.

  14. Contribution of human manganese superoxide dismutase tyrosine 34 to structure and catalysis.

    PubMed

    Perry, J Jefferson P; Hearn, Amy S; Cabelli, Diane E; Nick, Harry S; Tainer, John A; Silverman, David N

    2009-04-21

    Superoxide dismutase (SOD) enzymes are critical in controlling levels of reactive oxygen species (ROS) that are linked to aging, cancer, and neurodegenerative disease. Superoxide (O(2)(*-)) produced during respiration is removed by the product of the SOD2 gene, the homotetrameric manganese superoxide dismutase (MnSOD). Here, we examine the structural and catalytic roles of the highly conserved active-site residue Tyr34, based upon structure-function studies of MnSOD enzymes with mutations at this site. Substitution of Tyr34 with five different amino acids retained the active-site protein structure and assembly but caused a substantial decrease in the catalytic rate constant for the reduction of superoxide. The rate constant for formation of the product inhibition complex also decreases but to a much lesser extent, resulting in a net increase in the level of product inhibited form of the mutant enzymes. Comparisons of crystal structures and catalytic rates also suggest that one mutation, Y34V, interrupts the hydrogen-bonded network, which is associated with a rapid dissociation of the product-inhibited complex. Notably, with three of the Tyr34 mutants, we also observe an intermediate in catalysis, which has not been reported previously. Thus, these mutants establish a means of trapping a catalytic intermediate that promises to help elucidate the mechanism of catalysis.

  15. Effect of stilbene derivative on superoxide generation and enzyme release from human neutrophils in vitro

    PubMed Central

    Pečivová, Jana; Harmatha, Juraj; Sviteková, Klára; Nosáľ, Radomír

    2012-01-01

    Neutrophils represent the body′s primary line of defense against invading pathogens. They most rapidly reach the site of injury or infection, liberate antimicrobial proteins, proteases and produce reactive oxygen species. Prolonged or excessive liberation of these very effective and toxic substances could intensify the inflammatory process and enhance tissue damage in many diseases, such as allergies, infections and rheumatoid arthritis. Pterostilbene belongs to stilbenoids, structural analogues of resveratrol, which act as natural protective agents in defending the plant against viral and microbial attack. It possesses anticancerous, antidiabetic and anti-inflammatory properties. The study provides new information on the effect of pterostilbene [0.01–100 µmol/l] on superoxide generation in and myeloperoxidase (MPO) release from azurophil granules of isolated human neutrophils. PMA [1µmol/l], which activates NADPH-oxidase via protein kinase C, was used for stimulation of neutrophils Unstimulated cells showed neither superoxide generation nor myelopereoxidase release after preincubation with the drug studied. Pterostilbene dose dependently decreased superoxide generation in and MPO release from stimulated human neutrophils, however a significant decrease was recorded only in the concentration 100 µmol/l. The effect of pterostilbene was more pronounced on superoxide generation in comparison to MPO release. Our results suggest that the effect of pterostilbene may prove beneficial in controlling inflammation. PMID:23118590

  16. Sympathetic denervation caused by long-term noradrenaline infusions; prevention by desipramine and superoxide dismutase.

    PubMed

    Albino Teixeira, A; Azevedo, I; Branco, D; Rodrigues-Pereira, E; Osswald, W

    1989-05-01

    1. The effects of continuous intravenous infusion of noradrenaline (0.01 and 0.1 microgram kg-1 h-1) were studied in both the infused lateral saphenous vein and the contralateral saphenous vein of normal dogs. Noradrenaline, saline, noradrenaline + desipramine or noradrenaline + superoxide dismutase were infused using Alzet osmotic minipumps. 2. After a 5 day infusion period, the noradrenaline content in plasma and in both saphenous veins was determined, and the venous tissues submitted to light microscope morphometry and ultrastructural study and used for the determination of their O-methylation capacity (with [3H]-isoprenaline as a substrate). 3. Noradrenaline caused dose-dependent damage to the sympathetic nerve endings of the lateral saphenous veins. Concomitant changes in extraneuronal structure and function were observed (hypertrophy of smooth muscle cells, nuclear dysmorphy, thickening of the vessel wall, impairment in O-methylation capacity). 4. Desipramine and superoxide dismutase prevented or reduced the effects of noradrenaline on both the morphological and the biochemical parameters; the protection afforded by superoxide dismutase was more marked than that by desipramine. 5. It is concluded that moderately high doses of noradrenaline exert a 6-hydroxydopamine-like effect and that this chemical sympathectomy is partially or totally prevented by desipramine or superoxide dismutase. The data suggest that a substance derived from noradrenaline, in the formation of which free oxygen radicals are involved and which is subject to neuronal uptake, is the chemical entity responsible for the neurotoxic effect observed.

  17. Nutritional status of zinc and activity superoxide dismutase in chronic renal patients undergoing hemodialysis.

    PubMed

    Noleto Magalhães, R C; Guedes Borges de Araujo, C; Batista de Sousa Lima, V; Machado Moita Neto, J; do Nascimento Nogueira, N; do Nascimento Marreiro, D

    2011-01-01

    Chronic kidney disease promotes changes in the zinc nutritional status and in the antioxidant defense system. This study assessed the relationship between the parameters of the zinc nutritional status and the activity of superoxide dismutase in patients with chronic renal failure who are receiving hemodialysis. 134 individuals, aged between 18 and 85 years, were divided into two groups: case group (hemodialyzed patients, n = 63) and control group (n = 71). Zinc concentrations in plasma and erythrocytes were determined using the flame atomic absorption spectrophotometry technique. The activity of superoxide dismutase enzyme was determined according to Ransod kit. The mean values of plasma zinc were 62.02 ± 13.59 μg/dL and 65.58 ± 8.88 μg/dL, and for erythrocytary zinc the values were 54.52 ± 22.82 μgZn/gHb and 48.01 ± 15.08 μgZn/gHb for the chronic renal patients and the control group, respectively. The activity of superoxide dismutase was significantly lower in patients when compared with the control group (p < 0.05). The activity of superoxide dismutase in patients with chronic renal failure undergoing hemodialysis, which is influenced by zinc concentracions, was significantly lower. There was an inadequate response of this enzyme to oxidative stress in patients undergoing hemodialysis.

  18. SUPEROXIDE-DEPENDENT IRON UPTAKE: A NEW ROLE FOR ANION EXCHANGE PROTEIN 2

    EPA Science Inventory

    Lung cells import iron across the plasma membrane as ferrous (Fe2+) ion by incompletely understood mechanisms. We tested the hypothesis that human bronchial epithelial (HBE) cells import non-transferrin-bound iron (NTBI) using superoxide-dependent ferri-reductase activity involvi...

  19. Expression of Pyrococcus furiosus superoxide reductase in Arabidopsis enhances heat tolerance.

    PubMed

    Im, Yang Ju; Ji, Mikyoung; Lee, Alice; Killens, Rushyannah; Grunden, Amy M; Boss, Wendy F

    2009-10-01

    Plants produce reactive oxygen species (ROS) in response to environmental stresses sending signaling cues, which, if uncontrolled, result in cell death. Like other aerobic organisms, plants have ROS-scavenging enzymes, such as superoxide dismutase (SOD), which removes superoxide anion radical (O(2)(-)) and prevents the production and buildup of toxic free radicals. However, increasing the expression of cytosolic SODs is complex, and increasing their production in vivo has proven to be challenging. To avoid problems with endogenous regulation of gene expression, we expressed a gene from the archaeal hyperthermophile Pyrococcus furiosus that reduces O(2)(-). P. furiosus uses superoxide reductase (SOR) rather than SOD to remove superoxide. SOR is a thermostable enzyme that reduces O(2)(-) in a one-electron reduction without producing oxygen. We show that P. furiosus SOR can be produced as a functional enzyme in planta and that plants producing SOR have enhanced tolerance to heat, light, and chemically induced ROS. Stress tolerance in the SOR-producing plants correlates positively with a delayed increase in ROS-sensitive transcripts and a decrease in ascorbate peroxidase activity. The SOR plants provide a good model system to study the impact of cytosolic ROS on downstream signaling in plant growth and development. Furthermore, this work demonstrates that this synthetic approach for reducing cytosolic ROS holds promise as a means for improving stress tolerance in crop plants.

  20. Inhibition of the Ca sup 2+ -ATPase of vascular smooth muscle sarcoplasmic reticulum by superoxide radicals

    SciTech Connect

    Suzuki, Yuichiro; Ford, G.D. )

    1991-03-15

    The effect of oxygen free radicals generated by hypoxanthine plus xanthine oxidase on the Ca{sup 2+}-ATPase of sarcoplasmic reticulum from bovine aortic smooth muscle were studied. Exogenous hypoxanthine plus xanthine oxidase produced an hypoxanthine concentration dependent inhibition of the Ca{sup 2+}-ATPase. The inhibition could be completely blocked by superoxide dismutase but not by either mannitol or deferoxamine. Direct addition of reagent hydrogen peroxide in the {mu}M range did not cause significant inhibition. These results suggest that superoxide is the primary damaging species. Additionally, 1.16 {plus minus} 0.17 mU/g wet wt of xanthine oxidase activity were detected in the post-nuclear supernatant of bovine aortic smooth muscle, suggesting the existence of a possible intracellular source of superoxide. This value was calculated to be approximately 5 mU/ml by using a usual value of vascular smooth muscle cellular volume. Thus the level of endogenous xanthine oxidase resident in vascular smooth muscle is comparable with the level of exogenous xanthine oxidase used in the present study. These findings suggest a potential role of xanthine oxidase-generated superoxide in free radical injury to vascular smooth muscle.

  1. Copper-Zinc Superoxide Dismutase: A Unique Biological "Ligand" for Bioinorganic Studies.

    ERIC Educational Resources Information Center

    Valentine, Joan Selverstone; de Freitas, Duarte Mota

    1985-01-01

    Discusses superoxide dismutase (SOD) research and the properties of copper, zinc (Cu, Zn)-SOD. Emphasizes the controversy concerning the role of Cu,Zn-SOD and other SOD enzymes as protective agents in reactions involving dioxygen metabolism, and the properties of Cu, Zn-SOD that make it an interesting biological ligand for physical studies of…

  2. NADH Induces the Generation of Superoxide Radicals in Leaf Peroxisomes 1

    PubMed Central

    del Río, Luis A.; Fernández, Víctor M.; Rupérez, Francisco L.; Sandalio, Luisa M.; Palma, José M.

    1989-01-01

    In peroxisomes isolated from pea leaves (Pisum sativum L.) the production of superoxide free radicals (O2−) by xanthine and NADH was investigated. In peroxisomal membranes, 100 micromolar NADH induced the production of O2− radicals. In the soluble fractions of peroxisomes, no generation of O2− radicals was observed by incubation with either NADH or xanthine, although xanthine oxidase was found located predominantly in the matrix of peroxisomes. The failure of xanthine to induce superoxide generation was probably due to the inability to fully suppress the endogenous Mn-superoxide dismutase activity by inhibitors which were inactive against xanthine oxidase. The generation of superoxide radicals in leaf peroxisomes together with the recently described production of these oxygen radicals in glyoxysomes (LM Sandalio, VM Fernández, FL Rupérez, LA del Río [1988] Plant Physiol 87: 1-4) suggests that O2− generation could be a common metabolic property of peroxisomes and further supports the existence of active oxygen-related rôles for peroxisomes in cellular metabolism. PMID:16666612

  3. SUPEROXIDE-DEPENDENT IRON UPTAKE: A NEW ROLE FOR ANION EXCHANGE PROTEIN 2

    EPA Science Inventory

    Lung cells import iron across the plasma membrane as ferrous (Fe2+) ion by incompletely understood mechanisms. We tested the hypothesis that human bronchial epithelial (HBE) cells import non-transferrin-bound iron (NTBI) using superoxide-dependent ferri-reductase activity involvi...

  4. Bz-423 superoxide signals apoptosis via selective activation of JNK, Bak, and Bax.

    PubMed

    Blatt, Neal B; Boitano, Anthony E; Lyssiotis, Costas A; Opipari, Anthony W; Glick, Gary D

    2008-11-01

    Bz-423 is a proapoptotic 1,4-benzodiazepine with potent therapeutic properties in murine models of lupus and psoriasis. Bz-423 modulates the F(1)F(0)-ATPase, inducing the formation of superoxide within the mitochondrial respiratory chain, which then functions as a second messenger initiating apoptosis. Herein, we report the signaling pathway activated by Bz-423 in mouse embryonic fibroblasts containing knockouts of key apoptotic proteins. Bz-423-induced superoxide activates cytosolic ASK1 and its release from thioredoxin. A mitogen-activated protein kinase cascade follows, leading to the specific phosphorylation of JNK. JNK signals activation of Bax and Bak which then induces mitochondrial outer membrane permeabilization to cause the release of cytochrome c and a commitment to apoptosis. The response of these cells to Bz-423 is critically dependent on both superoxide and JNK activation as antioxidants and the JNK inhibitor SP600125 prevents Bax translocation, cytochrome c release, and cell death. These results demonstrate that superoxide generated from the mitochondrial respiratory chain as a consequence of a respiratory transition can signal a sequential and specific apoptotic response. Collectively, these data suggest that the selectivity of Bz-423 observed in vivo results from cell-type specific differences in redox balance and signaling by ASK1 and Bcl-2 proteins.

  5. Mitochondrial superoxide in osteocytes perturbs canalicular networks in the setting of age-related osteoporosis

    PubMed Central

    Kobayashi, Keiji; Nojiri, Hidetoshi; Saita, Yoshitomo; Morikawa, Daichi; Ozawa, Yusuke; Watanabe, Kenji; Koike, Masato; Asou, Yoshinori; Shirasawa, Takuji; Yokote, Koutaro; Kaneko, Kazuo; Shimizu, Takahiko

    2015-01-01

    Osteocytes are major bone cells that play a crucial role in maintaining the quality of and healing damage to bone tissue. The number of living osteocytes and canalicular networks declines in an age-dependent manner. However, the pathological effects of mitochondrial redox imbalances on osteocytes and bone metabolism have not been fully elucidated. We generated mice lacking mitochondrial superoxide dismutase 2 (Sod2) in osteocytes. Like an aged bone, Sod2 depletion in the osteocytes positively enhanced the production of cellular superoxide in vivo. A bone morphological analysis demonstrated that the Sod2-deficient femurs showed remarkable bone loss in an age-dependent manner. Interestingly, Sod2 loss induced markedly disorganized osteocytic canalicular networks and decreased the number of live osteocytes. Furthermore, Sod2 deficiency significantly suppressed bone formation and increased bone resorption concomitant with the upregulation of sclerostin and receptor activator of NF-κB ligand (RANKL). In vitro experiments also revealed that treatment with paraquat, a superoxide inducer in mitochondria, promoted the RANKL expression via, in part, ERK phosphorylation. These findings demonstrate that the mitochondrial superoxide induced in osteocytes by Sod2 ablation causes age-related bone loss due to the impairment of canalicular networks and bone metabolism via the deregulation of the sclerostin and RANKL expression. PMID:25779629

  6. Molecular Cloning and Expression of Sequence Variants of Manganese Superoxide Dismutase Genes from Wheat

    USDA-ARS?s Scientific Manuscript database

    Reactive oxygen species (ROS) are very harmful to living organisms due to the potential oxidation of membrane lipids, DNA, proteins, and carbohydrates. Transformed E.coli strain QC 871, superoxide dismutase (SOD) double-mutant, with three sequence variant MnSOD1, MnSOD2, and MnSOD3 manganese supero...

  7. Molecular cloning and biochemical characterization of iron superoxide dismutase from the rodent malaria parasite Plasmodium vinckei.

    PubMed

    Prakash, Kirtika; Goyal, Manish; Soni, Awakash; Siddiqui, Arif Jamal; Bhardwaj, Jyoti; Puri, Sunil K

    2014-12-01

    Plasmodium parasite utilizes superoxide dismutase family proteins to limit the toxicity of reactive oxygen species, such as produced through hemoglobin degradation. These proteins play an important role in parasite survival during intra-erythrocytic phase. We have identified, and biochemically characterized a putative iron dependent superoxide dismutase from rodent malaria parasite Plasmodium vinckei (PvSOD1). The recombinant PvSOD1 protein was purified to homogeneity through a combination of affinity and gel filtration chromatography. Crosslinking, Native-PAGE and FPLC gel filtration analyses documented that PvSOD1 exists as a dimer in solution, a common feature shared by other Fe-SODs. PvSOD1 is cytosolic in localization and its expression is comparatively higher during trophozoite as compared to that of ring and schizont stages. Enzymatic activity of recombinant PvSOD1 was validated using conventional zymogram analyses and xanthine-xanthine oxidase system. Under optimal conditions, PvSOD1 was highly active and catalyzed the dismutation of superoxide radicals. Furthermore, PvSOD1 showed activity over a broad range of pH and temperature. Inhibition studies suggested that PvSOD1 was inactivated by hydrogen peroxide, and peroxynitrite, but not by cyanide and azide. Since, PvSOD1 plays a central role in oxidative defense mechanism, therefore, characterization of PvSOD1 will be exploited in the screening of new superoxide dismutase inhibitors for their antimalarial activity.

  8. Curcumin Rescues Diabetic Renal Fibrosis by Targeting Superoxide-Mediated Wnt Signaling Pathways.

    PubMed

    Ho, Cheng; Hsu, Yung-Chien; Lei, Chen-Chou; Mau, Shu-Ching; Shih, Ya-Hsueh; Lin, Chun-Liang

    2016-03-01

    The purposes of this study were to investigate whether curcumin can weaken diabetic nephropathy by modulating both oxidative stress and renal injury from Wnt signaling mediation. Wnt5a/β-catenin depression and induction of superoxide synthesis are associated with high glucose (HG) induced transforming growth factor (TGF)-β1 and fibronectin expression in mesangial cells. Curcumin resumes HG depression of Wnt/β-catenin signaling and alleviates HG induction of superoxide, TGF-β1 and fibronectin expression in renal mesangial cell. Exogenous curcumin alleviated urinary total proteinuria and serum superoxide level in diabetic rats. Based on laser-captured microdissection for quantitative real-time polymerase chain reaction, it was found that diabetes significantly increased TGF-β1 and fibronectin expression in line with depressed Wnt5a expression. Curcumin treatment reduced the TGF-β1 and fibronectin activation and the inhibiting effect of diabetes on Wnt5a/β-catenin expression in renal glomeruli. Immunohistochemistry showed that curcumin treatment significantly reduced 8-hydroxy-2'-deoxyguanosine, TGF-β1 and fibronectin, and was in line with the restoration of the suppressed Wnt5a expression immunoreactivities in glomeruli of diabetic rats. Curcumin alleviated extracellular matrix accumulation in diabetic nephropathy by not only preventing the diabetes-mediated superoxide synthesis but also resuming downregulation of Wnt/β-catenin signaling. These findings suggest that regulation of Wnt activity by curcumin is a feasible alternative strategy to rescue diabetic renal injury.

  9. Induction of peroxidases and superoxide dismutases in transformed embryogenic calli of alfalfa (Medicago sativa L.)

    USDA-ARS?s Scientific Manuscript database

    Activities of peroxidase (POD) and superoxide dismutase (SOD) enzymes were analyzed in non-regenerative transformed embryogenic lines of alfalfa (Medicago sativa L.) carrying wound-inducible oryzacystatin I (OC-I), wound-inducible oryzacystatin I antisense (OC-Ias) or hygromycin phosphotransferase (...

  10. Superoxide dismutase activity in mesocarp tissue from divergent Cucumis melo L. genotypes

    USDA-ARS?s Scientific Manuscript database

    Muskmelon (Cucumis melo L.) fruit matrix is unique among plant foods in being able to provide a protective medium in which the antioxidant activity of the enzyme superoxide dismutase (SOD) is preserved during the digestive process, and therefore, being able to elicit in vivo pharmacological effects ...

  11. The preparation of calcium superoxide for air breathing and scrubbing applications

    NASA Technical Reports Server (NTRS)

    Ballou, E. V.; Wood, P. C.; Spitze, L. A.; Wydeven, T.

    1976-01-01

    There is interest in the preparation of high-purity calcium superoxide as an oxygen source for breathing apparatus because both the available oxygen and the capacity for carbon dioxide removal, per unit weight of superoxide, are higher than that of a number of other chemical oxygen sources. A review of earlier findings shows that the general method used by Vol'nov and coworkers for the decomposition of calcium peroxide diperoxyhydrate can yield preparations containing more than 58.4% calcium superoxide maximum predicted for an equimolar disproportionation reaction. The decomposition of solid calcium peroxide diperoxyhydrate is studied using an apparatus that allows good control of the critical reaction parameters. The removal of water from decomposing calcium peroxide diperoxyhydrate, before the same water has an opportunity to back react with the calcium superoxide formed in the reaction, constitutes the rationale of the experiments. Even with allowance for the anomalies observed in the analytical results, the yields appear to be in the 65+ percent range, and optimization of the experimental variables is still being pursued.

  12. Glucose-6-phosphate dehydrogenase-derived NADPH fuels superoxide production in the failing heart

    USDA-ARS?s Scientific Manuscript database

    In the failing heart, NADPH oxidase and uncoupled NO synthase utilize cytosolic NADPH to form superoxide. NADPH is supplied principally by the pentose phosphate pathway, whose rate-limiting enzyme is glucose 6-phosphate dehydrogenase (G6PD). Therefore, we hypothesized that cardiac G6PD activation dr...

  13. Mitochondrial superoxide in osteocytes perturbs canalicular networks in the setting of age-related osteoporosis.

    PubMed

    Kobayashi, Keiji; Nojiri, Hidetoshi; Saita, Yoshitomo; Morikawa, Daichi; Ozawa, Yusuke; Watanabe, Kenji; Koike, Masato; Asou, Yoshinori; Shirasawa, Takuji; Yokote, Koutaro; Kaneko, Kazuo; Shimizu, Takahiko

    2015-03-16

    Osteocytes are major bone cells that play a crucial role in maintaining the quality of and healing damage to bone tissue. The number of living osteocytes and canalicular networks declines in an age-dependent manner. However, the pathological effects of mitochondrial redox imbalances on osteocytes and bone metabolism have not been fully elucidated. We generated mice lacking mitochondrial superoxide dismutase 2 (Sod2) in osteocytes. Like an aged bone, Sod2 depletion in the osteocytes positively enhanced the production of cellular superoxide in vivo. A bone morphological analysis demonstrated that the Sod2-deficient femurs showed remarkable bone loss in an age-dependent manner. Interestingly, Sod2 loss induced markedly disorganized osteocytic canalicular networks and decreased the number of live osteocytes. Furthermore, Sod2 deficiency significantly suppressed bone formation and increased bone resorption concomitant with the upregulation of sclerostin and receptor activator of NF-κB ligand (RANKL). In vitro experiments also revealed that treatment with paraquat, a superoxide inducer in mitochondria, promoted the RANKL expression via, in part, ERK phosphorylation. These findings demonstrate that the mitochondrial superoxide induced in osteocytes by Sod2 ablation causes age-related bone loss due to the impairment of canalicular networks and bone metabolism via the deregulation of the sclerostin and RANKL expression.

  14. URBAN PARTICLE-INDUCED PULMONARY ARTERY CONSTRUCTION IS MEDIATED BY SUPEROXIDE PRODUCTION

    EPA Science Inventory

    URBAN PARTICLE-INDUCED PULMONARY ARTERY CONSTRICTION IS MEDIATED BY SUPEROXIDE PRODUCTION.Jacqueline D. Carter, Zhuowei Li, Lisa A. Dailey, Yuh-Chin T. Huang. CEMALB, University of North Carolina, and ORD, US EPA, Chapel Hill, North Carolina.

    Exposure to particulate matter...

  15. Titanium Dioxide Nanoparticles Increase Superoxide Anion Production by Acting on NADPH Oxidase

    PubMed Central

    Trepout, Sylvain; Wien, Frank; Marco, Sergio

    2015-01-01

    Titanium dioxide (TiO2) anatase nanoparticles (NPs) are metal oxide NPs commercialized for several uses of everyday life. However their toxicity has been poorly investigated. Cellular internalization of NPs has been shown to activate macrophages and neutrophils that contribute to superoxide anion production by the NADPH oxidase complex. Transmission electron micrososcopy images showed that the membrane fractions were close to the NPs while fluorescence indicated an interaction between NPs and cytosolic proteins. Using a cell-free system, we have investigated the influence of TiO2 NPs on the behavior of the NADPH oxidase. In the absence of the classical activator molecules of the enzyme (arachidonic acid) but in the presence of TiO2 NPs, no production of superoxide ions could be detected indicating that TiO2 NPs were unable to activate by themselves the complex. However once the NADPH oxidase was activated (i.e., by arachidonic acid), the rate of superoxide anion production went up to 140% of its value without NPs, this effect being dependent on their concentration. In the presence of TiO2 nanoparticles, the NADPH oxidase produces more superoxide ions, hence induces higher oxidative stress. This hyper-activation and the subsequent increase in ROS production by TiO2 NPs could participate to the oxidative stress development. PMID:26714308

  16. Superoxide anion generation and oxidative stress in methylmercury-induced endothelial toxicity in vitro.

    PubMed

    Ghizoni, Heloisa; de Souza, Viviane; Straliotto, Marcos Raniel; de Bem, Andreza Fabro; Farina, Marcelo; Hort, Mariana Appel

    2017-02-01

    Emerging evidence has pointed to mercury exposure as a risk factor for hypertension, atherosclerosis, myocardial infarction and coronary heart disease. However, the underlying mechanisms are not well understood. This study investigated potential toxic effects of low concentrations of methylmercury (MeHg) in cultured bovine aortic endothelial cells (BAECs) and the possible involvement of reactive species, particularly superoxide anion, in mediating such toxicity. MeHg treatment increased the oxidation of 2',7'-dichlorodihydrofluorescein diacetate (a general probe for reactive species) and dihydroethidium, a specific probe for superoxide anion. MeHg-induced 2',7'-dichlorodihydrofluorescein diacetate and dihydroethidium oxidations were significantly decreased by apocynin, an inhibitor of the enzyme NADPH oxidase, which represents a main source of superoxide anion in endothelial cells. MeHg treatment significantly disrupted mitochondrial membrane potential and this event was also reversed by apocynin. MeHg treatment also decreased glutathione levels and this event preceded glutathione peroxidase inhibition, which was observed only at 24h after treatment. These results indicate that MeHg induces oxidative stress in cultured BAECs and that this event is related to the production of superoxide anion. Moreover, the observed protective effects of apocynin in BAECs suggest the potential involvement of NADPH-oxidase in MeHg-induced endothelial dysfunction, which represents a pivotal event in most cardiovascular diseases.

  17. Involvement of Extracellular Cu/Zn Superoxide Dismutase in Cotton Fiber Primary and Secondary Cell Wall Biosynthesis

    USDA-ARS?s Scientific Manuscript database

    Extracellular Cu/Zn superoxide dismutases (CSDs) that catalyze the conversion of superoxide to hydrogen peroxide have been suggested to be involved in lignification of secondary walls in spinach, pine and aspen. In cotton fibers, hydrogen peroxide was proposed to be involved in the induction of seco...

  18. NADPH oxidase (NOX) 1 mediates cigarette smoke-induced superoxide generation in rat vascular smooth muscle cells.

    PubMed

    Chang, Kyung-Hwa; Park, Jung-Min; Lee, Chang Hoon; Kim, Bumseok; Choi, Kyung-Chul; Choi, Seong-Jin; Lee, Kyuhong; Lee, Moo-Yeol

    2017-02-01

    Smoking is a well-established risk factor for cardiovascular diseases. Oxidative stress is one of the common etiological factors, and NADPH oxidase (NOX) has been suggested as a potential mediator of oxidative stress. In this study, cigarette smoke (CS)-induced superoxide production was characterized in vascular smooth muscle cells (VSMC). CS was prepared in forms of cigarette smoke extract (CSE) and total particulate matter (TPM). Several molecular probes for reactive oxygen species were trialed, and dihydroethidium (DHE) and WST-1 were chosen for superoxide detection considering the autofluorescence, light absorbance, and peroxidase inhibitory activity of CS. Both CSE and TPM generated superoxide in a VSMC culture system by stimulating cells to produce superoxide and by directly producing superoxide in the aqueous solution. NOX, specifically NOX1 was found to be an important cellular source of superoxide through experiments with the NOX inhibitors diphenyleneiodonium (DPI) and VAS2870 as well as isoform-specific NOX knockdown. NOX inhibitors and the superoxide dismutase mimetic TEMPOL reduced the cytotoxicity of CSE, thus suggesting the contribution of NOX1-derived superoxide to cytotoxicity. Since NOX1 is known to mediate diverse pathological processes in the vascular system, NOX1 may be a critical effector of cardiovascular toxicity caused by smoking.

  19. Superoxide microsensor integrated into a Sensing Cell Culture Flask microsystem using direct oxidation for cell culture application.

    PubMed

    Flamm, H; Kieninger, J; Weltin, A; Urban, G A

    2015-03-15

    A new electrochemical sensor system for reliable and continuous detection of superoxide radical release from cell culture was developed utilizing direct oxidation of superoxide on polymer covered gold microelectrodes. Direct superoxide oxidation was demonstrated to provide robust measurement principle for sensitive and selective reactive oxygen species (ROS) quantification without the need for biocomponent supported conversion. Sensor performance was investigated by using artificial enzymatic superoxide production revealing a sensitivity of 2235AM(-1)m(-2). An electrode protection layer with molecular weight cut-off property from adsorbed linear branched polyethylenimine was successfully introduced for long term and selectivity improvement. Thin-film based sensor chip fabrication with implemented three-electrode setup and full integration into the technological platform Sensing Cell Culture Flask was described. Cell culturing directly on-chip and free radical release by phorbol-12-myristate-13-acetate (PMA) stimulation was demonstrated using T-47D human breast cancer carcinoma cell model. Transient extracellular superoxide production upon stimulation was successfully observed from amperometric monitoring. Signal inhibition from scavenging of extracellular superoxide by specific superoxide dismutase (SOD) showed the applicability for selective in vitro ROS determination. The results confirm the possibility of direct superoxide oxidation, with exclusion of the main interfering substances uric acid and hydrogen peroxide. This offers new insights into the development of reliable and robust ROS sensors.

  20. Acceleration of diabetic renal injury in the superoxide dismutase knockout mouse: effects of tempol.

    PubMed

    DeRubertis, Frederick R; Craven, Patricia A; Melhem, Mona F

    2007-09-01

    Indices of renal injury and oxidative stress were examined in mice with deficiency of cytosolic Cu(2+)/Zn(2+) superoxide dismutase (SOD1-/-, KO) and their wild-type (WT) littermates with streptozotocin-induced diabetes. After 5 weeks of diabetes, KO diabetic (D) but not WT-D mice developed marked albuminuria, increases in glomerular content of transforming growth factor beta, collagen alpha1(IV), and nitrotyrosine, and higher glomerular superoxide compared with corresponding values in nondiabetics. After 5 months of diabetes, increases in these parameters, mesangial matrix expansion, renal cortical malondialdehyde content, and severity of tubulointerstitial injury were all significantly greater, whereas cortical glutathione was lower, in KO-D than in WT-D. In contrast to WT-D, after 4 weeks of diabetes, KO-D mice did not develop the increase in inulin clearance (C(In)) characteristic of early diabetes. The nitric oxide synthase inhibitor N(omega)-nitro-l-arginine methylester suppressed C(In) in WT-D, but had no effect on C(In) in KO-D. Treatment of KO-D with the SOD mimetic tempol for 4 weeks suppressed albuminuria, increases in glomerular transforming growth factor beta, collagen alpha1(IV), nitrotyrosine, and glomerular superoxide, and concurrently increased C(In). The latter action of tempol in KO-D was blocked by the N(omega)-nitro-l-arginine methylester. The findings provide support for a role for superoxide and its metabolism by SOD1 in the pathogenesis of renal injury in diabetes in vivo, and implicate increased interaction of superoxide with nitric oxide as a pathogenetic factor.

  1. Superoxide dismutase reduces the impairment of endothelium-dependent relaxation in the spontaneously hypertensive rat aorta.

    PubMed

    Sekiguchi, Fumiko; Yanamoto, Aiko; Sunano, Satoru

    2004-04-01

    The involvement of the superoxide anion in endothelium-dependent relaxation (EDR) was examined in noradrenaline-contracted aortic smooth muscle preparations isolated from normotensive Wistar Kyoto rats (WKY) and stroke-prone spontaneously hypertensive rats (SHRSP). Acetylcholine (ACh, 10(-9)-10(-5) M) induced EDR in both WKY and SHRSP preparations in a concentration-dependent manner, but with a significantly smaller amplitude in those from SHRSP than in those from WKY. The ACh-induced EDR was inhibited by N(omega)-nitro-L-arginine (L-NOARG), in a concentration-dependent manner, both in WKY and SHRSP. The EDR produced in WKY in the presence of 3 x 10(-6) M L-NOARG was similar in magnitude to that produced in SHRSP in the absence of L-NOARG. Superoxide dismutase (SOD, 300 units/ml) increased the amplitude of EDR in SHRSP but not in WKY, with no alteration of the threshold or of the maximal amplitude. The maximal amplitude of EDR produced in SHRSP in the presence of SOD was still smaller than that in WKY. In WKY, a possible involvement of superoxide in the EDR was examined in aortae whose EDR was partially inhibited by treatment with a subthreshold concentration (3 x 10 (-6) M) of L-NOARG. In the L-NOARG-conditioned aorta, the reduced EDR was partially but significantly recovered by SOD. These results suggest that the impaired EDR in aortae of SHRSP may be causally related to a higher production of superoxide. The L-NOARG-induced inhibition of EDR in WKY may be produced, in part, by the reduction of effective NO due to its destruction by superoxide.

  2. Superoxide flashes: illuminating new insights into cardiac ischemia/reperfusion injury.

    PubMed

    Sheu, Shey-Shing; Wang, Wang; Cheng, Heping; Dirksen, Robert T

    2008-11-01

    Although the mitochondrial permeability transition pore (mPTP) was first discovered almost 30 years ago [1], it did not attract significant research attention until the 1990's when several studies implicated mPTP in apoptosis [2]. Today, the dogma suggests that opening of mPTP is detrimental to the cell and mPTP activation is widely thought to contribute to disease in cancer, neurodegenerative diseases, stroke, muscular dystrophy, and cardiac reperfusion injury [3]. Multiple factors including Ca(2+), OH(-), P(i), cyclophilin D, reactive oxygen and nitrogen species (ROS and RNS) trigger mPTP opening [4]. However, whether mPTP activation feeds back to alter mitochondrial ROS generation remains unclear. We recently demonstrated that under normal conditions, individual mitochondria undergo spontaneous transient bursts of quantal superoxide generation, termed "superoxide flashes" [5]. Superoxide flashes are observed in all cell types investigated to date and are triggered by a surprising functional coupling between mPTP activation and electron transport chain (ETC) dependent superoxide production. Additionally, reoxgenation following anoxia leads to uncontrolled superoxide flash genesis in cardiomyocytes. This positive feedback mechanism for mPTP/ETC-dependent ROS generation may drive localized redox signaling in individual mitochondria under physiological conditions, and when left unchecked, contribute to global cellular oxidative stress under pathological conditions in cardiac disease. The mPTP activity-dependent cell life and death determination imposes new challenges and opportunities in the pursuit of therapeutic agents for treating diseases in which oxidative stress has been implicated such as cardiac ischemia-reperfusion injury.

  3. Diosgenin inhibits superoxide generation in FMLP-activated mouse neutrophils via multiple pathways.

    PubMed

    Lin, Y; Jia, R; Liu, Y; Gao, Y; Zeng, X; Kou, J; Yu, B

    2014-12-01

    Diosgenin possesses anti-inflammatory and anticancer properties. Activated neutrophils produce high concentrations of the superoxide anion which is involved in the pathophysiology of inflammation-related diseases and cancer. In the present study, the inhibitory effect and possible mechanisms of diosgenin on superoxide generation were investigated in mouse bone marrow neutrophils. Diosgenin potently and concentration-dependently inhibited the extracellular and intracellular superoxide anion generation in Formyl-Met-Leu-Phe (FMLP)- activated neutrophils, with IC50 values of 0.50 ± 0.08 μM and 0.66 ± 0.13 μM, respectively. Such inhibition was not mediated by scavenging the superoxide anion or by a cytotoxic effect. Diosgenin inhibited the phosphorylation of p47phox and membrane translocation of p47phox and p67phox, and thus blocking the assembly of nicotinamide adenine dinucleotide phosphate oxidase. Moreover, cellular cyclic adenosine monophosphate (cAMP) levels and protein kinase A (PKA) expression were also effectively increased by diosgenin. It attenuated FMLP-induced increase of phosphorylation of cytosolic phospholipase A (cPLA2), p21-activated kinase (PAK), Akt, p38 mitogen-activated protein kinase (p38MAPK), extracellular signal-regulated kinase (ERK1/2), and c-Jun N-terminal kinase (JNK). Our data indicate that diosgenin exhibits inhibitory effects on superoxide anion production through the blockade of cAMP, PKA, cPLA2, PAK, Akt and MAPKs signaling pathways. The results may explain the clinical implications of diosgenin in the treatment of inflammation-related disorders.

  4. Sources of superoxide/H2O2 during mitochondrial proline oxidation.

    PubMed

    Goncalves, Renata L S; Rothschild, Daniel E; Quinlan, Casey L; Scott, Gary K; Benz, Christopher C; Brand, Martin D

    2014-01-01

    p53 Inducible gene 6 (PIG6) encodes mitochondrial proline dehydrogenase (PRODH) and is up-regulated several fold upon p53 activation. Proline dehydrogenase is proposed to generate radicals that contribute to cancer cell apoptosis. However, there are at least 10 mitochondrial sites that can produce superoxide and/or H2O2, and it is unclear whether proline dehydrogenase generates these species directly, or instead drives production by other sites. Amongst six cancer cell lines, ZR75-30 human breast cancer cells had the highest basal proline dehydrogenase levels, and mitochondria isolated from ZR75-30 cells consumed oxygen and produced H2O2 with proline as sole substrate. Insects use proline oxidation to fuel flight, and mitochondria isolated from Drosophila melanogaster were even more active with proline as sole substrate than ZR75-30 mitochondria. Using mitochondria from these two models we identified the sites involved in formation of superoxide/H2O2 during proline oxidation. In mitochondria from Drosophila the main sites were respiratory complexes I and II. In mitochondria from ZR75-30 breast cancer cells the main sites were complex I and the oxoglutarate dehydrogenase complex. Even with combinations of substrates and respiratory chain inhibitors designed to minimize the contributions of other sites and maximize any superoxide/H2O2 production from proline dehydrogenase itself, there was no significant direct contribution of proline dehydrogenase to the observed H2O2 production. Thus proline oxidation by proline dehydrogenase drives superoxide/H2O2 production, but it does so mainly or exclusively by providing anaplerotic carbon for other mitochondrial dehydrogenases and not by producing superoxide/H2O2 directly.

  5. Detection of superoxide production in stimulated and unstimulated living cells using new cyclic nitrone spin traps.

    PubMed

    Abbas, Kahina; Hardy, Micael; Poulhès, Florent; Karoui, Hakim; Tordo, Paul; Ouari, Olivier; Peyrot, Fabienne

    2014-06-01

    Reactive oxygen species (ROS), including superoxide anion and hydrogen peroxide (H2O2), have a diverse array of physiological and pathological effects within living cells depending on the extent, timing, and location of their production. For measuring ROS production in cells, the ESR spin trapping technique using cyclic nitrones distinguishes itself from other methods by its specificity for superoxide and hydroxyl radical. However, several drawbacks, such as the low spin trapping rate and the spontaneous and cell-enhanced decomposition of the spin adducts to ESR-silent products, limit the application of this method to biological systems. Recently, new cyclic nitrones bearing a triphenylphosphonium (Mito-DIPPMPO) or a permethylated β-cyclodextrin moiety (CD-DIPPMPO) have been synthesized and their spin adducts demonstrated increased stability in buffer. In this study, a comparison of the spin trapping efficiency of these new compounds with commonly used cyclic nitrone spin traps, i.e., 5,5-dimethyl-1-pyrroline N-oxide (DMPO), and analogs BMPO, DEPMPO, and DIPPMPO, was performed on RAW 264.7 macrophages stimulated with phorbol 12-myristate 13-acetate. Our results show that Mito-DIPPMPO and CD-DIPPMPO enable a higher detection of superoxide adduct, with a low (if any) amount of hydroxyl adduct. CD-DIPPMPO, especially, appears to be a superior spin trap for extracellular superoxide detection in living macrophages, allowing measurement of superoxide production in unstimulated cells for the first time. The main rationale put forward for this extreme sensitivity is that the extracellular localization of the spin trap prevents the reduction of the spin adducts by ascorbic acid and glutathione within cells.

  6. Unraveling the role of animal heme peroxidases in superoxide mediated Mn oxide formation

    NASA Astrophysics Data System (ADS)

    Learman, D. R.; Hansel, C. M.

    2013-12-01

    Manganese(III,IV) oxides are important in the environment as they can impact the fate of a broad range of nutrients (e.g. carbon and phosphate) and contaminates (e.g. lead and chromium). Bacteria play a valuable role in the production of Mn oxides, yet the mechanisms and physiological reasons remain unclear. Roseobacter sp. AzwK-3b, an organism within the abundant and ubiquitous Roseobacter clade, has recently been shown to oxidize Mn(II) via a novel pathway that involves enzymatic extracellular superoxide production. However, in reactions with only Mn(II) and abiotically generated superoxide, we find superoxide alone is not enough to produce Mn(III,IV) oxides. Scavenging of the byproduct hydrogen peroxide (via the addition of catalase) is required to generate Mn oxides via abiotic reaction of Mn(II) with superoxide. Thus, R. AzwK-3b must produce superoxide and also scavenge hydrogen peroxide to form Mn oxides. Further, in-gel Mn(II) oxidation assay revealed a protein band that could generate Mn oxides in the presence of soluble Mn(II). This Mn(II)-oxidizing protein band was excised from the gel and the peptides identified via mass spectrometry. An animal heme peroxidase (AHP) was the predominant protein found in this band. This protein is homologous to the AHPs previously implicated as a Mn(II)-oxidizing enzyme within the Alphaproteobacteria, Erythrobacter SD-21 and Aurantimonas manganoxydans strain SI85-9A1. Currently, protein expression of the AHPs in R. AzwK-3b is being examined to determine if expression is correlated with Mn(II) concentration or oxidative stress. Our data suggests that AHPs do not directly oxidize Mn(II) but rather plays a role in scavenging hydrogen peroxide and/or producing an organic Mn(III) ligand that complexes Mn(III) and likely aids in Mn oxide precipitation.

  7. Detection of superoxide radicals in tomato plants exposed to salinity, drought, cold and heavy metal stress using CMC-G-SOD biosensor.

    PubMed

    Kocabay, Ozge; Emregul, Emel; Aydın, Semra Soydan; Aras, Sumer

    2013-10-01

    A novel highly sensitive electrochemical carboxymethylcellulose-gelatin-superoxide dismutase biosensor was used for the determination of superoxide radicals enhancement in tomato plants exposed to salinity, drought, cold and heavy metal stress. The variations in superoxide radicals depending on abiotic stress was determined using biosensor. The superoxide radical production with regard to control rapidly was increased in tomato plants exposed to salinity, drought, cold and heavy metal stress. The superoxide radical enhancement in tomato plants exposed to salinity, drought, cold and heavy metal stress was successfully determined using carboxymethylcellulose-gelatin-superoxide dismutase biosensor.

  8. Over-expressed copper/zinc superoxide dismutase localizes to mitochondria in neurons inhibiting the angiotensin II-mediated increase in mitochondrial superoxide.

    PubMed

    Li, Shumin; Case, Adam J; Yang, Rui-Fang; Schultz, Harold D; Zimmerman, Matthew C

    2013-01-01

    Angiotensin II (AngII) is the main effector peptide of the renin-angiotensin system (RAS), and contributes to the pathogenesis of cardiovascular disease by exerting its effects on an array of different cell types, including central neurons. AngII intra-neuronal signaling is mediated, at least in part, by reactive oxygen species, particularly superoxide (O2 (•-)). Recently, it has been discovered that mitochondria are a major subcellular source of AngII-induced O2 (•-). We have previously reported that over-expression of manganese superoxide dismutase (MnSOD), a mitochondrial matrix-localized O2 (•-) scavenging enzyme, inhibits AngII intra-neuronal signaling. Interestingly, over-expression of copper/zinc superoxide dismutase (CuZnSOD), which is believed to be primarily localized to the cytoplasm, similarly inhibits AngII intra-neuronal signaling and provides protection against AngII-mediated neurogenic hypertension. Herein, we tested the hypothesis that CuZnSOD over-expression in central neurons localizes to mitochondria and inhibits AngII intra-neuronal signaling by scavenging mitochondrial O2 (•-). Using a neuronal cell culture model (CATH.a neurons), we demonstrate that both endogenous and adenovirus-mediated over-expressed CuZnSOD (AdCuZnSOD) are present in mitochondria. Furthermore, we show that over-expression of CuZnSOD attenuates the AngII-mediated increase in mitochondrial O2 (•-) levels and the AngII-induced inhibition of neuronal potassium current. Taken together, these data clearly show that over-expressed CuZnSOD in neurons localizes in mitochondria, scavenges AngII-induced mitochondrial O2 (•-), and inhibits AngII intra-neuronal signaling.

  9. Kinetic investigation of catalytic disproportionation of superoxide ions in the non-aqueous electrolyte used in Li–air batteries

    DOE PAGES

    Wang, Qiang; Zheng, Dong; McKinnon, Meaghan E.; ...

    2014-10-28

    Superoxide reacts with carbonate solvents in Li–air batteries. Tris(pentafluorophenyl)borane is found to catalyze a more rapid superoxide (O2-) disproportionation reaction than the reaction between superoxide and propylene carbonate (PC). With this catalysis, the negative impact of the reaction between the electrolyte and O2-produced by the O2 reduction can be minimized. A simple kinetic study using ESR spectroscopy was reported to determine reaction orders and rate constants for the reaction between PC and superoxide, and the disproportionation of superoxide catalyzed by Tris(pentafluorophenyl)borane and Li ions. As a result, the reactions are found to be first order and the rate constants aremore » 0.033 s-1 M-1, 0.020 s-1 M-1and 0.67 s-1M-1 for reactions with PC, Li ion and Tris(pentafluorophenyl)borane, respectively.« less

  10. Positive control of a global antioxidant defense regulon activated by superoxide-generating agents in Escherichia coli.

    PubMed Central

    Greenberg, J T; Monach, P; Chou, J H; Josephy, P D; Demple, B

    1990-01-01

    Escherichia coli responds to superoxide-generating agents by inducing approximately 40 proteins. We have identified a genetic locus, soxR (superoxide response), that positively regulates 9 of these proteins during superoxide stress. Induction under soxR control is at the transcriptional level, as shown with lac fusions to five paraquat-inducible promoters. Members of the soxR regulon include at least three proteins with demonstrable antioxidant roles: Mn-containing superoxide dismutase (which destroys superoxide radicals), endonuclease IV (which repairs radical-induced damages in DNA), and glucose-6-phosphate dehydrogenase (which produces NADPH). Induction of the soxR regulon also leads to diminished levels of the major outer membrane protein OmpF and alteration of the small-subunit ribosomal protein S6. These latter changes confer resistance to a variety of antibiotics. The soxR regulon may thus operate as an inducible defense against xenobiotics in general. Images PMID:1696718

  11. Dynamics of Superoxide Production and Decay in Natural Trichodesmium Colonies from the Sargasso Sea: Implications for Cell Signaling

    NASA Astrophysics Data System (ADS)

    Hansel, C. M.; Buchwald, C.; Diaz, J. M.; Dyhrman, S.; Van Mooy, B. A. S.

    2014-12-01

    Reactive oxygen species (ROS) are key players in the biogeochemistry of the ocean, where they serve a critical role in the cycling of carbon and metals. Research in the past decade has introduced phytoplankton and, most recently, heterotrophic bacteria as significant sources of ROS, including superoxide, within both photic and aphotic regions of the ocean. ROS are both beneficial and detrimental to life. For instance, superoxide is a vital inter- and intra-cellular signaling molecule, yet at high concentrations it induces lipid peroxidation and initiates programmed cell death (PCD). In fact, superoxide has been implicated in PCD in the nitrogen-fixing diazotroph Trichodesmium, presumably leading to the demise of blooms within oligotrophic marine systems. Here, we explore the rates of superoxide production and decay by natural Trichodesmium populations obtained from various surface waters in the Sargasso Sea. We investigate also the role of light and colony density and morphology (puff v. raft) on superoxide fluxes. We find that Trichodesmium colonies produce extracellular superoxide at extremely high rates in the dark that are on par with those of the toxic raphidophyte Chattonella. The rates of superoxide production, however, rapidly decline with increasing cell density pointing to a role for superoxide in cell signaling in these organisms. We also find extremely rapid extracellular superoxide degradation by Trichodesmium. Together, this likely reflects a need for these organisms to maintain ROS at levels that will support signaling but below the threshold level that triggers PCD or oxidative damage. We also show differences in the effect of light on superoxide fluxes as a function of Trichodesmium colony morphology, suggesting differences in either colony physiology or associated bacterial symbionts. These findings point to complex physiological, ecological, and physical influences on ROS dynamics in phytoplankton that require further exploration.

  12. Superoxide radicals increase transforming growth factor-{beta}1 and collagen release from human lung fibroblasts via cellular influx through chloride channels

    SciTech Connect

    Qi Shufan Hartog, Gertjan J.M. den; Bast, Aalt

    2009-05-15

    Reactive oxygen species (ROS) have been implicated in the pathogenesis of fibrosis. However, it remains unclear which ROS is the major cause. We hypothesize that superoxide elicits specific toxicity to human lung fibroblasts and plays an important role in the development of pulmonary fibrosis. In this study, superoxide generated from xanthine and xanthine oxidase activated lung fibroblasts by increasing the release of TGF-{beta}1 and collagen. This was associated with increased levels of intracellular superoxide. SOD and tempol, by scavenging respectively extracellular and intracellular superoxide, prevented the activation of fibroblasts induced by exposure to exogenous superoxide, whereas catalase did not. Moreover, hydrogen peroxide did not activate fibroblasts. Apparently, superoxide rather than hydrogen peroxide is involved in the regulation of TGF-{beta}1 and collagen release in lung fibroblasts. The chloride channel blocker, DIDS, inhibited the increase of intracellular superoxide levels induced by exogenous superoxide and consequently prevented the activation of fibroblasts. This suggests that the cellular influx of superoxide through chloride channels is essential for superoxide-induced activation of fibroblasts. ERK1/2 and p38 MAPKs are involved in the intracellular pathway leading to superoxide-induced fibroblasts activation. Superoxide possesses until now undiscovered specific pro-fibrotic properties in human lung fibroblasts. This takes place via the cellular influx of superoxide through chloride channels rather than via the formation of hydrogen peroxide.

  13. Superoxide release by confluent endothelial cells, an electron spin resonance (ESR) study

    NASA Astrophysics Data System (ADS)

    Barbacanne, M.-A.; Margeat, E.; Arnal, J.-F.; Nepveu, F.; Souchard, J.-P.

    1999-01-01

    In the present study we used ESR to detect the release of oxygen radicals by endothelial cells stimulated with calcium ionophore A23187. Dimethyl-1-pyrroline-N-oxide (DMPO) was used as a spin trap. Although the observed adduct (DMPO-OH) suggested the presence of the hydroxyl radical, the use of superoxide dismutase and catalase revealed that superoxide anion was released in the medium. Superoxide production was more efficient when the cells were post-confluent for a few days. The release of superoxide was 3-fold greater in growth arrested cells (D6-D9) than in proliferating cells (D0). Although two inhibitors of the mitochondrial respiratory chain carbanyl cyanide m-chlorophenylhydrazone (CCCP), antimycine decreased the ESR signal by 35%, the use of superoxide dismutase (SOD) and tumor necrosis factor (TNF) suggested that the release of O2- occurred in the cell membrane. The physiological significance of this extracellular superoxide release by post-confluent cells deserves further study. Ce travail présente une étude par RSE de la libération des radicaux oxygénés par les cellules endothéliales bovines (BAEC) sous l'effet de l'ionophore calcique A23187. Le diméthyl-1-pyrroline-N-oxyde (DMPO) est utilisé comme piégeur de spin. Bien que l'adduit formé (DMPO-OH) semble traduire la présence du radical hydroxyle .OH, l'utilisation de superoxyde dismutase et de catalase a révélé que les cellules endothéliales libéraient l'anion superoxyde. La production du radical superoxyde est plus abondante lorsque les cellules sont à confluence depuis plusieurs jours. Lorsque les cellules sont entre J6 et J9, la production de superoxyde est trois fois supérieure à celle observée lorsque les cellules sont en prolifération (J0). Bien que deux inhibiteurs de la chaîne mitochondriale 1-carbonyldinitrile-m-chlorophenylhydrazone (CCCP), antimycineinhibent de 35 % le signal RPE, l'utilisation de superoxyde dismutase (SOD) et du tumor necrosis factor (TNF) sugg

  14. Ethylene Improves Root System Development under Cadmium Stress by Modulating Superoxide Anion Concentration in Arabidopsis thaliana

    PubMed Central

    Abozeid, Ann; Ying, Zuojia; Lin, Yingchao; Liu, Jia; Zhang, Zhonghua; Tang, Zhonghua

    2017-01-01

    This work aims at identifying the effects of ethylene on the response of Arabidopsis thaliana root system to cadmium chloride (CdCl2) stress. Two ethylene-insensitive mutants, ein2-5 and ein3-1eil1-1, were subjected to (25, 50, 75, and 100 μM) CdCl2 concentrations, from which 75 μM concentration decreased root growth by 40% compared with wild type Col-0 as a control. Ethylene biosynthesis increased in response to CdCl2 treatment. The length of primary root and root tip in ein2-5 and ein3-1eil1-1 decreased compared with wild type after CdCl2 treatment, suggesting that ethylene play a role in root system response to Cd stress. The superoxide concentration in roots of ein2-5 and ein3-1eil1-1 was greater than in wild type seedlings under Cd stress. Application of exogenous 1-aminocyclopropane-1-carboxylic acid (ACC) (a precursor of ethylene biosynthesis) in different concentrations (0.01, 0.05 and 0.5 μM) decreased superoxide accumulation in Col-0 root tips and increased the activities of superoxide dismutase (SOD) isoenzymes under Cd stress. This result was reversed with 5 μM of aminoisobutyric acid AIB (an inhibitor of ethylene biosynthesis). Moreover, it was accompanied by increase in lateral roots number and root hairs length, indicating the essential role of ethylene in modulating root system development by controlling superoxide accumulation through SOD isoenzymes activities. The suppressed Cd-induced superoxide accumulation in wild type plants decreased the occurrence of cells death while programmed cell death (PCD) was initiated in the root tip zone, altering root morphogenesis (decreased primary root length, more lateral roots and root hairs) to minimize the damage caused by Cd stress, whereas this response was absent in the ein2-5 and ein3-1eil1-1 seedlings. Hence, ethylene has a role in modulating root morphogenesis during CdCl2 stress in A. thaliana by increasing the activity of SOD isoenzymes to control superoxide accumulation. PMID:28286514

  15. Extracellular Production and Degradation of Superoxide in the Coral Stylophora pistillata and Cultured Symbiodinium

    PubMed Central

    Saragosti, Eldad; Tchernov, Dan; Katsir, Adi; Shaked, Yeala

    2010-01-01

    Background Reactive oxygen species (ROS) are thought to play a major role in cell death pathways and bleaching in scleractinian corals. Direct measurements of ROS in corals are conspicuously in short supply, partly due to inherent problems with ROS quantification in cellular systems. Methodology/Principal Findings In this study we characterized the dynamics of the reactive oxygen species superoxide anion radical (O2−) in the external milieu of the coral Stylophora pistillata. Using a sensitive, rapid and selective chemiluminesence-based technique, we measured extracellular superoxide production and detoxification activity of symbiont (non-bleached) and aposymbiont (bleached) corals, and of cultured Symbiodinium (from clades A and C). Bleached and non-bleached Stylophora fragments were found to produce superoxide at comparable rates of 10−11–10−9 mol O2− mg protein−1 min−1 in the dark. In the light, a two-fold enhancement in O2− production rates was observed in non-bleached corals, but not in bleached corals. Cultured Symbiodinium produced superoxide in the dark at a rate of . Light was found to markedly enhance O2− production. The NADPH Oxidase inhibitor Diphenyleneiodonium chloride (DPI) strongly inhibited O2− production by corals (and more moderately by algae), possibly suggesting an involvement of NADPH Oxidase in the process. An extracellular O2− detoxifying activity was found for bleached and non-bleached Stylophora but not for Symbiodinium. The O2− detoxifying activity was partially characterized and found to resemble that of the enzyme superoxide dismutase (SOD). Conclusions/Significance The findings of substantial extracellular O2− production as well as extracellular O2− detoxifying activity may shed light on the chemical interactions between the symbiont and its host and between the coral and its environment. Superoxide production by Symbiodinium possibly implies that algal bearing corals are more susceptible to an internal

  16. A novel amperometric biosensor for superoxide anion based on superoxide dismutase immobilized on gold nanoparticle-chitosan-ionic liquid biocomposite film.

    PubMed

    Wang, Lu; Wen, Wei; Xiong, Huayu; Zhang, Xiuhua; Gu, Haoshuang; Wang, Shengfu

    2013-01-03

    A novel superoxide anion (O(2)(-)) biosensor is proposed based on the immobilization of copper-zinc superoxide dismutase (SOD) in a gold nanoparticle-chitosan-ionic liquid (GNPs-CS-IL) biocomposite film. The SOD-based biosensor was constructed by one-step ultrasonic electrodeposition of GNP-CS-IL composite onto glassy carbon electrode (GCE), followed by immobilization of SOD on the modified electrode. Surface morphologies of a set of representative films were characterized by scanning electron microscopy. The electrochemical performance of the biosensor was evaluated by cyclic voltammetry and chronoamperometry. A pair of quasi-reversible redox peaks of SOD with a formal potential of 0.257V was observed at SOD/GNPs-CS-IL/GCE in phosphate buffer solution (PBS, 0.1M, pH 7.0). The effects of varying test conditions on the electrochemical behavior of the biosensor were investigated. Furthermore, several electrochemical parameters were calculated in detail. Based on the biomolecule recognition of the specific reactivity of SOD toward O(2)(-), the developed biosensor exhibited a fast amperometric response (<5s), wide linear range (5.6-2.7×10(3)nM), low detection limit (1.7nM), and excellent selectivity for the real-time measurement of O(2)(-). The proposed method is promising for estimating quantitatively the dynamic changes of O(2)(-) in biological systems. Copyright © 2012 Elsevier B.V. All rights reserved.

  17. Alteration of cellular phenotype and responses to oxidative stress by manganese superoxide dismutase and a superoxide dismutase mimic in RWPE-2 human prostate adenocarcinoma cells.

    PubMed

    Zhong, Weixiong; Yan, Tao; Webber, Mukta M; Oberley, Terry D

    2004-06-01

    To study biologic effects of increased manganese superoxide dismutase (MnSOD) on cell behavior, we overexpressed MnSOD in a human prostate cancer cell line RWPE-2 by cDNA transfection. Stable transfectants of MnSOD showed a two- to threefold increase in MnSOD protein and enzymatic activity and a decrease in growth rate with prolonged cell population doubling times. Western blot analysis showed a 1.5- to twofold increase in the cyclin-dependent kinase inhibitor p21(Waf1) in MnSOD transfectants. Overexpression of MnSOD resulted in a seven- to eightfold increase in reduced glutathione (GSH), 18- to 26-fold increase in oxidized glutathione (GSSG), and a two- to threefold decrease in the ratio of GSH to GSSG. MnSOD-overexpressing cells showed an increase in sensitivity to the cytotoxicity of buthionine sulfoximine, a glutathione-depleting agent, and vitamin C, but a decrease in sensitivity to sodium selenite. Treatment with a superoxide dismutase (SOD) mimic MnTMPyP resulted in similar effects of MnSOD overexpression on cell responses to vitamin C and selenium. These data demonstrate that overexpression of MnSOD or treatment with SOD mimics can result in antioxidant or prooxidant effects in cells, depending on the presence of other antioxidants and prooxidants. MnSOD also has redox regulatory effects on cell growth and gene expression. These findings suggest that MnSOD and SOD mimics have the potential for cancer prevention or treatment.

  18. Quenching of superoxide ions by curcumin. A mechanistic study in acetonitrile.

    PubMed

    Toniolo, Rosanna; Di Narda, Francesca; Susmel, Sabina; Martelli, Mario; Martelli, Laura; Bontempelli, Gino

    2002-03-01

    The quenching of superoxide ions, O2.-, by curcumin has been studied by electrogenerating this anion radical from oxygen dissolved in acetonitrile solvent (that is, at best, a mimic of the lipofilic layer of biological membranes), containing known amounts of curcumin. Voltammetric tests, combined with coulometric and spectrophotometric measurements, pointed out that each mol of curcumin is able to react with six mols of such anion radical, through a process initiated by an acid-base step, which provides the perhydroxyl radical, HO2.; that disproportionates rapidly to the anionic form of hydrogen peroxide, HO2-, and oxygen, which is thus partially regenerated. At the same time, curcumin is converted to the corresponding three-charged anion. The strict resemblance existing between the mechanism of the rapid superoxide radical decay caused by curcumin and that involved in the presence of the superoxodismutase enzyme (SOD) is also underlined.

  19. N-Glycosylation is essential for the secretion of extracellular superoxide dismutase.

    PubMed

    Ota, Fumi; Kizuka, Yasuhiko; Kitazume, Shinobu; Adachi, Tetsuo; Taniguchi, Naoyuki

    2016-10-01

    Extracellular superoxide dismutase (EC-SOD or SOD3) protects against various oxidative stress-related diseases by scavenging reactive superoxides in the extracellular space. It is the only SOD isozyme that is secreted and glycosylated (at asparagine 89). However, the physiological roles of its glycosylation are poorly understood. In this study, we found that the glycosylation site on EC-SOD is well conserved and that a glycosylation-deficient EC-SOD mutant retains its enzymatic activity, but is not secreted. This impairment in secretion may, in part, be due to the ability of the mutants to form unusual higher order oligomers. Our findings reveal that the glycan modification is a key regulator of EC-SOD secretion and contributes to the understanding of the roles of glycans in EC-SOD-related diseases.

  20. Biphasic Superoxide Generation in Potato Tubers. A Self-Amplifying Response to Stress1

    PubMed Central

    Johnson, S.M.; Doherty, S.J.; Croy, R.R.D.

    2003-01-01

    Potato (Solanum tuberosum) cultivars differ quantitatively in their responses to mechanical stress including the ability to synthesize melanin pigments in tuber tissues. Investigations into the cellular events induced by mechanical stress on tuber tissues have shown that an early cellular response is a significant and rapid synthesis of superoxide radicals. This burst of radical production distinctively displays a reproducible biphasic pattern over time with peaks of generation at 2 and 5 h. A concomitant consequence of the generation of these free radicals is elevated levels of oxidatively modified tuber proteins. Both radical generation and protein modification vary between cultivars but both are directly proportional to the amount of melanin pigments produced. Cell-free extracts of mechanically stressed tissues, pectic fragments, and scission products generated from cell walls are able to induce superoxide generation in non-stressed tissues, indicating the participation of a biologically active factor that induces a further a phase of radical synthesis. PMID:12644693

  1. Alterations in superoxide dismutase and catalase in Fusarium oxysporum during starvation-induced differentiation.

    PubMed

    Kono, Y; Yamamoto, H; Takeuchi, M; Komada, H

    1995-07-20

    Vegetative hyphae of Fusarium oxysporum differentiate into chlamydospore by triggering with carbon-starvation. The current changes in the cellular detoxifying defenses against superoxide and hydrogen peroxide: superoxide dismutase (SOD) and catalase, were examined. Although there was a little change in catalase, a dramatic change in SOD was observed during the differentiation. In vegetative hyphae of F. oxysporum f. sp. raphani, three isozymes of SOD, all of which were not inhibited by hydrogen peroxide and cyanide, were present whereas in chlamydospore an isoenzyme, which was inhibited by hydrogen peroxide but not by cyanide, was present. Thus, as differentiation proceeded, Mn-type SOD disappeared and an Fe-type SOD appeared. The results suggest that the Fe-type SOD is specifically expressed during chlamydospore formation and that active intermediates of oxygen and/or its scavenging enzymes participate in the differentiation of Fusarium oxysporum.

  2. Superoxide generation from the reduction of oxygen at the carbon-oil-water triple phase boundary.

    PubMed

    Nissim, Rita; Compton, Richard G

    2013-07-28

    The reduction of oxygen is studied in aqueous solutions of pH 6.22-8.01, at a carbon paste electrode fabricated from dioctyl phthalate (oil) and graphite. Two two-electron voltammetric waves are usually seen on carbon electrodes, associated with the formation of hydrogen peroxide and water, respectively. However, an additional signal is seen on the carbon paste electrode, which can attributed to the initial formation of the superoxide radical anion, O2˙(-). Data is presented to show that the predominant source of oxygen for this reaction is that dissolved in the carbon paste material, rather than in the aqueous solution, and that the superoxide is likely formed at the graphite-oil-water triple phase boundary. Kinetic and thermodynamic parameters for the O2/O2˙(-) redox couple are reported.

  3. Correlation of Electronic and Geometric Structure in Mononuclear Copper(II) Superoxide Complexes

    PubMed Central

    Ginsbach, Jake W.; Peterson, Ryan L.; Cowley, Ryan E.; Karlin, Kenneth D.; Solomon, Edward I.

    2013-01-01

    The geometry of mononuclear copper(II) superoxide complexes has been shown to determine their ground state where side-on bonding leads to a singlet ground state and end-on complexes have triplet ground states. In apparent contrast to this trend, the recently synthesized (HIPT3tren)CuII–O2•− (1) was proposed to have an end-on geometry and a singlet ground state. However, re-examination of 1 with resonance Raman (rR), magnetic circular dichroism (MCD), and 2H NMR spectroscopy indicates that 1 is in fact an end-on superoxide species with a triplet ground state that results from the single CuII–O2•− bonding interaction being weaker than the spin pairing energy. PMID:24164429

  4. Preliminary neutron diffraction analysis of challenging human manganese superoxide dismutase crystals.

    PubMed

    Azadmanesh, Jahaun; Trickel, Scott R; Weiss, Kevin L; Coates, Leighton; Borgstahl, Gloria E O

    2017-04-01

    Superoxide dismutases (SODs) are enzymes that protect against oxidative stress by dismutation of superoxide into oxygen and hydrogen peroxide through cyclic reduction and oxidation of the active-site metal. The complete enzymatic mechanisms of SODs are unknown since data on the positions of hydrogen are limited. Here, methods are presented for large crystal growth and neutron data collection of human manganese SOD (MnSOD) using perdeuteration and the MaNDi beamline at Oak Ridge National Laboratory. The crystal from which the human MnSOD data set was obtained is the crystal with the largest unit-cell edge (240 Å) from which data have been collected via neutron diffraction to sufficient resolution (2.30 Å) where hydrogen positions can be observed.

  5. The effects of superoxide dismutase knockout on the oxidative stress parameters and survival of mouse erythrocytes.

    PubMed

    Grzelak, Agnieszka; Kruszewski, Marcin; Macierzyńska, Ewa; Piotrowski, Łukasz; Pułaski, Łukasz; Rychlik, Błazej; Bartosz, Grzegorz

    2009-01-01

    The erythrocytes of 12-month old Sod1 (-/-) mice showed an increased level of reactive oxygen species (ROS), as estimated by the degree of dihydroethidine and dihydrorhodamine oxidation, and the increased level of Heinz bodies. No indices of severe oxidative stress were found in the red blood cells and blood plasma of Sod1 (-/-) mice as judged from the lack of significant changes in the levels of erythrocyte and plasma glutathione, plasma protein thiol and carbonyl groups and thiobarbituric-acid reactive substances in the blood plasma. However, a decreased erythrocyte lifespan, increased reticulocyte count and splenomegaly were noted, indicating the importance of superoxide dismutase for maintaining erythrocyte viability. The levels of erythrocyte ROS and Heinz bodies and the reticulocyte count were indistinguishable in Sod1 (+/+) and Sod1 (+/-) mice, suggesting that a superoxide dismutase activity decrease to half of its normal value may be sufficient to secure the protective effects of the enzyme.

  6. Observation of Superoxide Production During Catalysis of Bacillus subtilis Oxalate Decarboxylase at pH4

    PubMed Central

    Twahir, Umar T.; Stedwell, Corey N.; Lee, Cory T.; Richards, Nigel G. J.; Polfer, Nicolas C.; Angerhofer, Alexander

    2015-01-01

    This contribution describes the trapping of the hydroperoxyl radical at a pH of 4 during turnover of wild-type oxalate decarboxylase and its T165V mutant using the spin trap BMPO. Radicals were detected and identified by a combination of EPR and mass spectrometry. Superoxide, or its conjugate acid, the hydroperoxyl radical, is expected as an intermediate in the decarboxylation and oxidation reactions of the oxalate monoanion both of which are promoted by oxalate decarboxylase. Another intermediate, the carbon dioxide radical anion was also observed. The quantitative yields of superoxide trapping is similar in the wild type and the mutant while it is significantly different for the trapping of the carbon dioxide radical anion. This suggests that the two radicals are released from different sites of the protein. PMID:25526893

  7. Free-radical chain oxidation of 2-nitropropane initiated and propagated by superoxide.

    PubMed Central

    Kuo, C F; Fridovich, I

    1986-01-01

    The superoxide radical O2.-, whether produced by the xanthine/xanthine oxidase reaction or infused as KO2, solubilized by a crown ether in dry dimethyl sulphoxide, initiated a free-radical chain oxidation of anionic 2-nitropropane. Superoxide dismutase, but not catalase, inhibited oxidation of the nitroalkane. Xanthine oxidase suffered a syncatalytic inactivation, during the co-oxidation of 2-nitropropane, which was reversed by dialysis. Cyanide exacerbated this syncatalytic inactivation and rendered it irreversible. The frequently observed oxidations of nitroalkanes by flavoenzymes now need to be re-examined to clarify the extent to which O2.--initiated free-radical chain oxidation contributed to the overall nitroalkane oxidation. PMID:3026320

  8. Intracellular localization of the superoxide dismutases of Escherichia coli: a reevaluation.

    PubMed Central

    Britton, L; Fridovich, I

    1977-01-01

    All of the superoxide dismutase isozymes of Escherichia coli have been shown to occur in the cell matrix, and none have been found in the periplasm. This was the case with both E. coli B and E. coli K-12, whether grown on a low phosphate medium or on a Trypticase soy-yeast extract medium. Alkaline phosphatase was used as a marker of the periplasm; adenosine deaminase and glucose 6-phosphate dehydrogenase were used as matrix markers, and consistent results were obtained by osmotic shock, spheroplast formation, and use of a diazonium salt that penetrates the periplasm but cannot cross the plasma membrane. A previous report that the iron-containing superoxide dismutase of E. coli is a periplasmic enzyme is now seen to have been in error. PMID:330499

  9. Iron superoxide dismutases in eukaryotic pathogens: new insights from Apicomplexa and Trypanosoma structures.

    PubMed

    Phan, Isabelle Q H; Davies, Douglas R; Moretti, Nilmar Silvio; Shanmugam, Dhanasekaran; Cestari, Igor; Anupama, Atashi; Fairman, James W; Edwards, Thomas E; Stuart, Kenneth; Schenkman, Sergio; Myler, Peter J

    2015-05-01

    Prior studies have highlighted the potential of superoxide dismutases as drug targets in eukaryotic pathogens. This report presents the structures of three iron-dependent superoxide dismutases (FeSODs) from Trypanosoma cruzi, Leishmania major and Babesia bovis. Comparison with existing structures from Plasmodium and other trypanosome isoforms shows a very conserved overall fold with subtle differences. In particular, structural data suggest that B. bovis FeSOD may display similar resistance to peroxynitrite-mediated inactivation via an intramolecular electron-transfer pathway as previously described in T. cruzi FeSOD isoform B, thus providing valuable information for structure-based drug design. Furthermore, lysine-acetylation results in T. cruzi indicate that acetylation occurs at a position close to that responsible for the regulation of acetylation-mediated activity in the human enzyme.

  10. Preliminary neutron diffraction analysis of challenging human manganese superoxide dismutase crystals

    PubMed Central

    Azadmanesh, Jahaun; Trickel, Scott R.; Borgstahl, Gloria E. O.

    2017-01-01

    Superoxide dismutases (SODs) are enzymes that protect against oxidative stress by dismutation of superoxide into oxygen and hydrogen peroxide through cyclic reduction and oxidation of the active-site metal. The complete enzymatic mechanisms of SODs are unknown since data on the positions of hydrogen are limited. Here, methods are presented for large crystal growth and neutron data collection of human manganese SOD (MnSOD) using perdeuteration and the MaNDi beamline at Oak Ridge National Laboratory. The crystal from which the human MnSOD data set was obtained is the crystal with the largest unit-cell edge (240 Å) from which data have been collected via neutron diffraction to sufficient resolution (2.30 Å) where hydrogen positions can be observed. PMID:28368283

  11. The preparation of calcium superoxide in a flowing gas stream and fluidized bed

    NASA Technical Reports Server (NTRS)

    Wood, P. C.; Ballou, E. V.; Spitze, L. A.; Wydeven, T.

    1980-01-01

    Superoxides can be used as sources of chemically stored oxygen in emergency breathing apparatus. The work reported here describes the use of a low-pressure nitrogen gas sweep through the reactant bed, for temperature control and water vapor removal. For a given set of gas temperature, bed thickness, and reaction time values, the highest purity calcium superoxide, Ca(O2)2, was obtained at the highest space velocity of the nitrogen gas sweep. The purity of the product was further increased by flow conditions that resulted in the fluidization of the reactant bed. However, scale-up of the low-pressure fluidized bed process was limited to the formation of agglomerates of reactant particles, which hindered thermal control by the flowing gas stream. A radiofrequency flow discharge inside the reaction chamber prevented agglomeration, presumably by dissipation of the static charges on the fluidized particles.

  12. p38-MAPK- and Caspase-3-Mediated Superoxide-Induced Apoptosis of Rat Hepatic Stellate Cells: Reversal by Retinoic Acid

    PubMed Central

    Jameel, Noor Mohamed; Thirunavukkarasu, Chinnasamy; Wu, Tong; Watkins, Simon C.; Friedman, Scott L.; Gandhi, Chandrashekhar R.

    2010-01-01

    Reactive oxygen species (ROS) activate retinoid-containing quiescent hepatic stellate cells (qHSCs) to retinoid-deficient fibrogenic myofibroblast-like cells (aHSCs). However, ROS also cause apoptosis of aHSCs, and apoptotic aHSCs are observed in inflammatory fibrotic liver. Here, we investigated mechanisms of the effects of oxidative stress on the survival of qHSCs and aHSCs. HSCs from normal rat liver were used after overnight culture (qHSCs), or in 3–5 passages (aHSCs). For in vivo induction of oxidative stress, tert-butylhydroperoxide was injected into control and CCl4-induced cirrhotic rats. Spontaneous caspase-3 activation and apoptosis, observed in cultured qHSCs, decreased with time and were unaffected by superoxide. In contrast, superoxide caused caspase-3 and p38-MAPK activation, reduction in Bcl-xL expression, and apoptosis in aHSCs. Inhibition of caspase-3 and p38-MAPK did not affect the viability of qHSCs in the absence or presence of superoxide, but inhibited superoxide-induced death of aHSCs. Glutathione (GSH) level and activities of superoxide dismutase (SOD), catalase and glutathione peroxidase (GPx) were lower in aHSCs than qHSCs. Superoxide increased GSH content, and activities of SOD, catalase and GPx in qHSCs but not in aHSCs. Incubation of 13-cis-retinoic acid (RA)-treated aHSCs with superoxide increased their GSH content significantly, and prevented superoxide-induced p38-MAPK and caspase-3 activation while dramatically reducing the extent of apoptosis. Finally, oxidative stress induced in vivo caused apoptosis of aHSCs in cirrhotic but not of qHSCs in control rats. These results suggest that the absence of retinoids render aHSCs susceptible to superoxide-induced apoptosis via caspase-3 and p38-MAPK activation. PMID:18792915

  13. Indirect detection of superoxide in RAW 264.7 macrophage cells using microchip electrophoresis coupled to laser-induced fluorescence.

    PubMed

    de Campos, Richard P S; Siegel, Joseph M; Fresta, Claudia G; Caruso, Giuseppe; da Silva, José A F; Lunte, Susan M

    2015-09-01

    Superoxide, a naturally produced reactive oxygen species (ROS) in the human body, is involved in many pathological and physiological signaling processes. However, if superoxide formation is left unregulated, overproduction can lead to oxidative damage to important biomolecules, such as DNA, lipids, and proteins. Superoxide can also lead to the formation of peroxynitrite, an extremely hazardous substance, through its reaction with endogenously produced nitric oxide. Despite its importance, quantitative information regarding superoxide production is difficult to obtain due to its high reactivity and low concentrations in vivo. MitoHE, a fluorescent probe that specifically reacts with superoxide, was used in conjunction with microchip electrophoresis (ME) and laser-induced fluorescence (LIF) detection to investigate changes in superoxide production by RAW 264.7 macrophage cells following stimulation with phorbol 12-myristate 13-acetate (PMA). Stimulation was performed in the presence and absence of the superoxide dismutase (SOD) inhibitors, diethyldithiocarbamate (DDC) and 2-metoxyestradiol (2-ME). The addition of these inhibitors resulted in an increase in the amount of superoxide specific product (2-OH-MitoE(+)) from 0.08 ± 0.01 fmol (0.17 ± 0.03 mM) in native cells to 1.26 ± 0.06 fmol (2.5 ± 0.1 mM) after PMA treatment. This corresponds to an approximately 15-fold increase in intracellular concentration per cell. Furthermore, the addition of 3-morpholino-sydnonimine (SIN-1) to the cells during incubation resulted in the production of 0.061 ± 0.006 fmol (0.12 ± 0.01 mM) of 2-OH-MitoE(+) per cell on average. These results demonstrate that indirect superoxide detection coupled with the use of SOD inhibitors and a separation method is a viable method to discriminate the 2-OH-MitoE(+) signal from possible interferences.

  14. [Role of superoxide anion radicals in the bacterial corrosion of metals].

    PubMed

    Belov, D V; Kalinina, A A; Sokolova, T N; Smirnov, V F; Chelnokova, M V; Kartashov, V R

    2012-01-01

    It was found that seven strains of bacteria can cause corrosion damage to aluminum, its alloys, and zinc. With respect to the studied metals, the most active bacteria were Proteus vulgaris 1212 and Pseudomonas aeruginosa 969. Superoxide anion radicals were demonstrated to play a role in the initiation of corrosive damage to aluminum and zinc, while bacterial exometabolites participate in the later stages of this process.

  15. Mitochondrial superoxide flashes: metabolic biomarkers of skeletal muscle activity and disease

    PubMed Central

    Wei, Lan; Salahura, Gheorghe; Boncompagni, Simona; Kasischke, Karl A.; Protasi, Feliciano; Sheu, Shey-Shing; Dirksen, Robert T.

    2011-01-01

    Mitochondrial superoxide flashes (mSOFs) are stochastic events of quantal mitochondrial superoxide generation. Here, we used flexor digitorum brevis muscle fibers from transgenic mice with muscle-specific expression of a novel mitochondrial-targeted superoxide biosensor (mt-cpYFP) to characterize mSOF activity in skeletal muscle at rest, following intense activity, and under pathological conditions. Results demonstrate that mSOF activity in muscle depended on electron transport chain and adenine nucleotide translocase functionality, but it was independent of cyclophilin-D-mediated mitochondrial permeability transition pore activity. The diverse spatial dimensions of individual mSOF events were found to reflect a complex underlying morphology of the mitochondrial network, as examined by electron microscopy. Muscle activity regulated mSOF activity in a biphasic manner. Specifically, mSOF frequency was significantly increased following brief tetanic stimulation (18.1±1.6 to 22.3±2.0 flashes/1000 μm2·100 s before and after 5 tetani) and markedly decreased (to 7.7±1.6 flashes/1000 μm2·100 s) following prolonged tetanic stimulation (40 tetani). A significant temperature-dependent increase in mSOF frequency (11.9±0.8 and 19.8±2.6 flashes/1000 μm2·100 s at 23°C and 37°C) was observed in fibers from RYR1Y522S/WT mice, a mouse model of malignant hyperthermia and heat-induced hypermetabolism. Together, these results demonstrate that mSOF activity is a highly sensitive biomarker of mitochondrial respiration and the cellular metabolic state of muscle during physiological activity and pathological oxidative stress.—Wei, L., Salahura, G., Boncompagni, S., Kasischke, K. A., Protasi, F., Sheu, S.-S., Dirksen, R. T. Mitochondrial superoxide flashes: metabolic biomarkers of skeletal muscle activity and disease. PMID:21646399

  16. Nox4-generated superoxide drives angiotensin II-induced neural stem cell proliferation.

    PubMed

    Topchiy, Elena; Panzhinskiy, Evgeniy; Griffin, W Sue T; Barger, Steven W; Das, Mita; Zawada, W Michael

    2013-01-01

    Reactive oxygen species (ROS) have been reported to affect neural stem cell self-renewal and therefore may be important for normal development and may influence neurodegenerative processes when ROS activity is elevated. To determine if increasing production of superoxide, via activation of NADPH oxidase (Nox), increases neural stem cell proliferation, 100 nM angiotensin II (Ang II) - a strong stimulator of Nox - was applied to cultures of a murine neural stem cell line, C17.2. Twelve hours following a single treatment with Ang II, there was a doubling of the number of neural stem cells. This increase in neural stem cell numbers was preceded by a gradual elevation of superoxide levels (detected by dihydroethidium fluorescence) from the steady state at 0, 5, and 30 min and gradually increasing from 1 h to the maximum at 12 h, and returning to baseline at 24 h. Ang II-dependent proliferation was blocked by the antioxidant N-acetyl-L-cysteine. Confocal microscopy revealed the presence of two sources of intracellular ROS in C17.2 cells: (i) mitochondrial and (ii) extramitochondrial; the latter indicative of the involvement of one or more specific isoforms of Nox. Of the Nox family, mRNA expression for one member, Nox4, is abundant in neural stem cell cultures, and Ang II treatment resulted in elevation of the relative levels of Nox4 protein. SiRNA targeting of Nox4 mRNA reduced both the constitutive and Ang II-induced Nox4 protein levels and attenuated Ang II-driven increases in superoxide levels and stem cell proliferation. Our findings are consistent with our hypothesis that Ang II-induced proliferation of neural stem cells occurs via Nox4-generated superoxide, suggesting that an Ang II/Nox4 axis is an important regulator of neural stem cell self-renewal and as such may fine-tune normal, stress- or disease-modifying neurogenesis.

  17. Nox4-generated superoxide drives angiotensin II-induced neural stem cell proliferation

    PubMed Central

    Topchiy, Elena; Panzhinskiy, Evgeniy; Griffin, W. Sue T.; Barger, Steven W.; Das, Mita; Zawada, W. Michael

    2013-01-01

    Reactive oxygen species (ROS) have been reported to affect neural stem cell self-renewal and therefore may be important for normal development and may influence neurodegenerative processes when ROS activity is elevated. To determine if increasing production of superoxide, via activation of NADPH oxidase (Nox), increases neural stem cell proliferation, 100nM angiotensin II (Ang II) – a strong stimulator of Nox – was applied to cultures of a murine neural stem cell line C17.2. Twelve hours following a single treatment with Ang II there was a doubling of the number of neural stem cells. This increase in neural stem cell numbers was preceded by a gradual elevation of superoxide levels (detected by dihydroethidium, DHE, fluorescence) from the steady state at 0, 5, and 30 minutes and gradually increasing from one hour to the maximum at 12 h, and returning to baseline at 24 h. Ang II-dependent proliferation was blocked by the antioxidant N-acetyl-L-cysteine (NAC). Confocal microscopy revealed the presence of two sources of intracellular ROS in C17.2 cells: i) mitochondrial and ii) extramitochondrial; the latter indicative of involvement of one or more specific isoforms of Nox. Of the Nox family, mRNA expression for one member, Nox4, is abundant in neural stem cell cultures, and Ang II treatment resulted in elevation of the relative levels of Nox4 protein. SiRNA targeting of Nox4 mRNA reduced both the constitutive and Ang II-induced Nox4 protein levels and attenuated Ang II-driven increases in superoxide levels and stem cell proliferation. Our findings are consistent with our hypothesis that Ang II-induced proliferation of neural stem cells occurs via Nox4-generated superoxide, suggesting that an Ang II/Nox4 axis is an important regulator of neural stem cell self-renewal and as such may fine-tune normal or stress- or disease-modifying neurogenesis. PMID:23751520

  18. Superoxide production by a manganese-oxidizing bacterium facilitates iodide oxidation.

    PubMed

    Li, Hsiu-Ping; Daniel, Benjamin; Creeley, Danielle; Grandbois, Russell; Zhang, Saijin; Xu, Chen; Ho, Yi-Fang; Schwehr, Kathy A; Kaplan, Daniel I; Santschi, Peter H; Hansel, Colleen M; Yeager, Chris M

    2014-05-01

    The release of radioactive iodine (i.e., iodine-129 and iodine-131) from nuclear reprocessing facilities is a potential threat to human health. The fate and transport of iodine are determined primarily by its redox status, but processes that affect iodine oxidation states in the environment are poorly characterized. Given the difficulty in removing electrons from iodide (I(-)), naturally occurring iodide oxidation processes require strong oxidants, such as Mn oxides or microbial enzymes. In this study, we examine iodide oxidation by a marine bacterium, Roseobacter sp. AzwK-3b, which promotes Mn(II) oxidation by catalyzing the production of extracellular superoxide (O2(-)). In the absence of Mn(2+), Roseobacter sp. AzwK-3b cultures oxidized ∼90% of the provided iodide (10 μM) within 6 days, whereas in the presence of Mn(II), iodide oxidation occurred only after Mn(IV) formation ceased. Iodide oxidation was not observed during incubations in spent medium or with whole cells under anaerobic conditions or following heat treatment (boiling). Furthermore, iodide oxidation was significantly inhibited in the presence of superoxide dismutase and diphenylene iodonium (a general inhibitor of NADH oxidoreductases). In contrast, the addition of exogenous NADH enhanced iodide oxidation. Taken together, the results indicate that iodide oxidation was mediated primarily by extracellular superoxide generated by Roseobacter sp. AzwK-3b and not by the Mn oxides formed by this organism. Considering that extracellular superoxide formation is a widespread phenomenon among marine and terrestrial bacteria, this could represent an important pathway for iodide oxidation in some environments.

  19. Superoxide Production by a Manganese-Oxidizing Bacterium Facilitates Iodide Oxidation

    PubMed Central

    Li, Hsiu-Ping; Daniel, Benjamin; Creeley, Danielle; Grandbois, Russell; Zhang, Saijin; Xu, Chen; Ho, Yi-Fang; Schwehr, Kathy A.; Kaplan, Daniel I.; Santschi, Peter H.; Hansel, Colleen M.

    2014-01-01

    The release of radioactive iodine (i.e., iodine-129 and iodine-131) from nuclear reprocessing facilities is a potential threat to human health. The fate and transport of iodine are determined primarily by its redox status, but processes that affect iodine oxidation states in the environment are poorly characterized. Given the difficulty in removing electrons from iodide (I−), naturally occurring iodide oxidation processes require strong oxidants, such as Mn oxides or microbial enzymes. In this study, we examine iodide oxidation by a marine bacterium, Roseobacter sp. AzwK-3b, which promotes Mn(II) oxidation by catalyzing the production of extracellular superoxide (O2−). In the absence of Mn2+, Roseobacter sp. AzwK-3b cultures oxidized ∼90% of the provided iodide (10 μM) within 6 days, whereas in the presence of Mn(II), iodide oxidation occurred only after Mn(IV) formation ceased. Iodide oxidation was not observed during incubations in spent medium or with whole cells under anaerobic conditions or following heat treatment (boiling). Furthermore, iodide oxidation was significantly inhibited in the presence of superoxide dismutase and diphenylene iodonium (a general inhibitor of NADH oxidoreductases). In contrast, the addition of exogenous NADH enhanced iodide oxidation. Taken together, the results indicate that iodide oxidation was mediated primarily by extracellular superoxide generated by Roseobacter sp. AzwK-3b and not by the Mn oxides formed by this organism. Considering that extracellular superoxide formation is a widespread phenomenon among marine and terrestrial bacteria, this could represent an important pathway for iodide oxidation in some environments. PMID:24561582

  20. Deletion of Aldose Reductase from Mice Inhibits Diabetes-Induced Retinal Capillary Degeneration and Superoxide Generation

    PubMed Central

    Tang, Jie; Du, Yunpeng; Petrash, J. Mark; Sheibani, Nader; Kern, Timothy S.

    2013-01-01

    Purpose Pharmacologic inhibition of aldose reductase (AR) previously has been studied with respect to diabetic retinopathy with mixed results. Since drugs can have off-target effects, we studied the effects of AR deletion on the development and molecular abnormalities that contribute to diabetic retinopathy. Since recent data suggests an important role for leukocytes in the development of the retinopathy, we determined also if AR in leukocytes contributes to leukocyte-mediated death of retinal endothelial cells in diabetes. Methods Wild-type (WT; C57BL/6J) and AR deficient (AR−/−) mice were made diabetic with streptozotocin. Mice were sacrificed at 2 and 10 months of diabetes to evaluate retinal vascular histopathology, to quantify retinal superoxide production and biochemical and physiological abnormalities in the retina, and to assess the number of retinal endothelial cells killed by blood leukocytes in a co-culture system. Results Diabetes in WT mice developed the expected degeneration of retinal capillaries, and increased generation of superoxide by the retina. Leukocytes from diabetic WT mice also killed more retinal endothelial cells than did leukocytes from nondiabetic animals (p<0.0001). Deletion of AR largely (P<0.05) inhibited the diabetes-induced degeneration of retinal capillaries, as well as the increase in superoxide production by retina. AR-deficiency significantly inhibited the diabetes-induced increase in expression of inducible nitric oxide synthase (iNOS) in retina, but had no significant effect on expression of intercellular adhesion molecule-1 (ICAM-1), phosphorylated p38 MAPK, or killing of retinal endothelial cells by leukocytes. Conclusions AR contributes to the degeneration of retinal capillaries in diabetic mice. Deletion of the enzyme inhibits the diabetes-induced increase in expression of iNOS and of superoxide production, but does not correct a variety of other pro-inflammatory abnormalities associated with the development of

  1. Thermodynamic and kinetic considerations for the reaction of semiquinone radicals to form superoxide and hydrogen peroxide.

    PubMed

    Song, Yang; Buettner, Garry R

    2010-09-15

    The quinone/semiquinone/hydroquinone triad (Q/SQ(*-)/H(2)Q) represents a class of compounds that has great importance in a wide range of biological processes. The half-cell reduction potentials of these redox couples in aqueous solutions at neutral pH, E degrees ', provide a window to understanding the thermodynamic and kinetic characteristics of this triad and their associated chemistry and biochemistry in vivo. Substituents on the quinone ring can significantly influence the electron density "on the ring" and thus modify E degrees' dramatically. E degrees' of the quinone governs the reaction of semiquinone with dioxygen to form superoxide. At near-neutral pH the pK(a)'s of the hydroquinone are outstanding indicators of the electron density in the aromatic ring of the members of these triads (electrophilicity) and thus are excellent tools to predict half-cell reduction potentials for both the one-electron and two-electron couples, which in turn allow estimates of rate constants for the reactions of these triads. For example, the higher the pK(a)'s of H(2)Q, the lower the reduction potentials and the higher the rate constants for the reaction of SQ(*-) with dioxygen to form superoxide. However, hydroquinone autoxidation is controlled by the concentration of di-ionized hydroquinone; thus, the lower the pK(a)'s the less stable H(2)Q to autoxidation. Catalysts, e.g., metals and quinone, can accelerate oxidation processes; by removing superoxide and increasing the rate of formation of quinone, superoxide dismutase can accelerate oxidation of hydroquinones and thereby increase the flux of hydrogen peroxide. The principal reactions of quinones are with nucleophiles via Michael addition, for example, with thiols and amines. The rate constants for these addition reactions are also related to E degrees'. Thus, pK(a)'s of a hydroquinone and E degrees ' are central to the chemistry of these triads. Copyright 2010 Elsevier Inc. All rights reserved.

  2. Bioactivity screening of Pinus brutia bark extracts: superoxide dismutase-like and nitric oxide scavenging effects.

    PubMed

    Cretu, Elena; Miron, S D; Miron, Anca

    2013-01-01

    In the Eastern Mediterranean region the wood of Pinus brutia Ten. is used as building material, the bark being the main waste of the wood production processes. A study was designed to explore possible medicinal applications for the bark waste. This paper reports on the superoxide dismutase-like and NO scavenging effects of bark extracts. Bark extracts (a raw hydromethanolic extract and its fractions) were initially screened regarding their ability to scavenge DPPH radical. The superoxide dismutase-like and NO scavenging effects were further evaluated. Catechin and quercetin were the positive controls in all antioxidant assays. According to the EC50 values, all extracts (11.8 +/- 0.1 - 21.10 +/- 0.05 microg/ml) efficiently scavenged DPPH radical in comparison to quercetin (3.13 +/- 0.05 microg/ml) and catechin (6.36 +/- 0.05 microg/ml). The raw extract, diethyl ether and ethyl acetate fractions (449.46 +/- 1.75, 115.43 +/- 0.25 and 278.3 +/- 2.3 microg/ml, respectively) exhibited higher superoxide dismutase-like effects in comparison to catechin (> 645.1 microg/ml). In NO scavenging assay, the raw extract, ethyl acetate and aqueous fractions (160.63 +/- 0.85, 162.96 +/- 0.52 and 160.23 +/- 2.35 microg/ml, respectively) showed a scavenging ability similar to that of quercetin (156.76 +/- 5.05 microg/ml) and higher than the one developed by catechin (242.66 +/- 7.65 microg/ml). As superoxide anion and NO are important mediators in inflammation, our results support a possible use of Pinus brutia bark waste to develop nutraceuticals with efficiency in disorders involving oxidative and inflammatory stress.

  3. Behavioral and Neurotransmitter Abnormalities in Mice Deficient for Parkin, DJ-1 and Superoxide Dismutase

    PubMed Central

    Hennis, Meghan R.; Seamans, Katherine W.; Marvin, Marian A.; Casey, Bradford H.; Goldberg, Matthew S.

    2013-01-01

    Parkinson’s disease (PD) is a progressive neurodegenerative disease characterized by loss of neurons in the substantia nigra that project to the striatum and release dopamine. The cause of PD remains uncertain, however, evidence implicates mitochondrial dysfunction and oxidative stress. Although most cases of PD are sporadic, 5-10% of cases are caused by inherited mutations. Loss-of-function mutations in Parkin and DJ-1 were the first to be linked to recessively inherited Parkinsonism. Surprisingly, mice bearing similar loss-of-function mutations in Parkin and DJ-1 do not show age-dependent loss of nigral dopaminergic neurons or depletion of dopamine in the striatum. Although the normal cellular functions of Parkin and DJ-1 are not fully understood, we hypothesized that loss-of-function mutations in Parkin and DJ-1 render cells more sensitive to mitochondrial dysfunction and oxidative stress. To test this hypothesis, we crossed mice deficient for Parkin and DJ-1 with mice deficient for the mitochondrial antioxidant protein Mn-superoxide dismutase (SOD2) or the cytosolic antioxidant protein Cu-Zn-superoxide dismutase (SOD1). Aged Parkin-/-DJ-1-/- and Mn-superoxide dismutase triple deficient mice have enhanced performance on the rotorod behavior test. Cu/Zn-superoxide dismutase triple deficient mice have elevated levels of dopamine in the striatum in the absence of nigral cell loss. Our studies demonstrate that on a Parkin/DJ-1 null background, mice that are also deficient for major antioxidant proteins do not have progressive loss of dopaminergic neurons but have behavioral and striatal dopamine abnormalities. PMID:24386432

  4. New water-soluble Mn-porphyrin with catalytic activity for superoxide dismutation and peroxynitrite decomposition.

    PubMed

    Asayama, Shoichiro; Nakajima, Takumi; Kawakami, Hiroyoshi

    2011-07-01

    We have synthesized a new water-soluble cationic Mn-porphyrin with catalytic activity for both superoxide dismutation and peroxynitrite decomposition. The resulting Mn-porphyrin also showed higher stability for reactive oxygen species such as hydrogen peroxide and lower cytotoxicity, when compared with a control normal Mn-porphyrin. Furthermore, the new porphyrin recovered the viability of lipopolysaccharide-stimulated macrophage RAW 264.7 cells but the control Mn-porphyrin did not.

  5. Absence of superoxide dismutase activity causes nuclear DNA fragmentation during the aging process

    SciTech Connect

    Muid, Khandaker Ashfaqul; Karakaya, Hüseyin Çaglar; Koc, Ahmet

    2014-02-07

    Highlights: • Aging process increases ROS accumulation. • Aging process increases DNA damage levels. • Absence of SOD activity does not cause DNA damage in young cells. • Absence of SOD activity accelerate aging and increase oxidative DNA damages during the aging process. - Abstract: Superoxide dismutases (SOD) serve as an important antioxidant defense mechanism in aerobic organisms, and deletion of these genes shortens the replicative life span in the budding yeast Saccharomyces cerevisiae. Even though involvement of superoxide dismutase enzymes in ROS scavenging and the aging process has been studied extensively in different organisms, analyses of DNA damages has not been performed for replicatively old superoxide dismutase deficient cells. In this study, we investigated the roles of SOD1, SOD2 and CCS1 genes in preserving genomic integrity in replicatively old yeast cells using the single cell comet assay. We observed that extend of DNA damage was not significantly different among the young cells of wild type, sod1Δ and sod2Δ strains. However, ccs1Δ mutants showed a 60% higher amount of DNA damage in the young stage compared to that of the wild type cells. The aging process increased the DNA damage rates 3-fold in the wild type and more than 5-fold in sod1Δ, sod2Δ, and ccs1Δ mutant cells. Furthermore, ROS levels of these strains showed a similar pattern to their DNA damage contents. Thus, our results confirm that cells accumulate DNA damages during the aging process and reveal that superoxide dismutase enzymes play a substantial role in preserving the genomic integrity in this process.

  6. Increased superoxide levels in ganglia and sympathoexcitation are involved in sarafotoxin 6c-induced hypertension.

    PubMed

    Li, Melissa; Dai, Xiaoling; Watts, Stephanie; Kreulen, David; Fink, Gregory

    2008-11-01

    Endothelin (ET) type B receptors (ET(B)R) are expressed in multiple tissues and perform different functions depending on their location. ET(B)R mediate endothelium-dependent vasodilation, clearance of circulating ET, and diuretic effects; all of these should produce a fall in arterial blood pressure. However, we recently showed that chronic activation of ET(B)R in rats with the selective agonist sarafotoxin 6c (S6c) causes sustained hypertension. We have proposed that one mechanism of this effect is constriction of capacitance vessels. The current study was performed to determine whether S6c hypertension is caused by increased generation of reactive oxygen species (ROS) and/or activation of the sympathetic nervous system. The model used was continuous 5-day infusion of S6c into male Sprague-Dawley rats. No changes in superoxide anion levels in arteries and veins were found in hypertensive S6c-treated rats. However, superoxide levels were increased in sympathetic ganglia from S6c-treated rats. In addition, superoxide levels in ganglia increased progressively the longer the animals received S6c. Treatment with the antioxidant tempol impaired S6c-induced hypertension and decreased superoxide levels in ganglia. Acute ganglion blockade lowered blood pressure more in S6c-treated rats than in vehicle-treated rats. Although plasma norepinephrine levels were not increased in S6c hypertension, surgical ablation of the celiac ganglion plexus, which provides most of the sympathetic innervation to the splanchnic organs, significantly attenuated hypertension development. The results suggest that S6c-induced hypertension is partially mediated by sympathoexcitation to the splanchnic organs driven by increased oxidative stress in prevertebral sympathetic ganglia.

  7. Superoxide and its metabolism during germination and axis growth of Vigna radiata (L.) Wilczek seeds

    PubMed Central

    Singh, Khangembam Lenin; Chaudhuri, Abira; Kar, Rup Kumar

    2014-01-01

    Involvement of reactive oxygen species in regulation of plant growth and development is recently being demonstrated with various results depending on the experimental system and plant species. Role of superoxide and its metabolism in germination and axis growth was investigated in case of Vigna radiata seeds, a non-endospermous leguminous species having epigeal germination, by studying the effect of different reactive oxygen species (ROS) inhibitors, distribution of O2•ˉ and H2O2 and ROS enzyme profile in axes. Germination percentage and axis growth were determined under treatment with ROS inhibitors and scavengers. Localization of O2•ˉ and H2O2 was done using nitroblue tetrazolium (NBT) and 3,3′,5,5′-tetramethyl benzidine dihydrochloride hydrate (TMB), respectively. Apoplastic level of O2•ˉ was monitored by spectrophotometric analysis of bathing medium of axes. Profiles of NADPH oxidase and superoxide dismutase (SOD) were studied by in-gel assay. Germination was retarded by treatments affecting ROS level except H2O2 scavengers, while axis growth was retarded by all. Superoxide synthesis inhibitor and scavenger prevented H2O2 accumulation in axes in later phase as revealed from TMB staining. Activity of Cu/Zn SOD1 was initially high and declined thereafter. Superoxide being produced in apoplast possibly by NADPH oxidase activity is further metabolized to •OH via H2O2. Germination process depends possibly on •OH production in the axes. Post-germinative axis growth requires O2•ˉ while the differentiating zone of axis (radicle) requires H2O2 for cell wall stiffening. PMID:25763616

  8. Superoxide production during reduction of molecular oxygen by assimilatory nitrate reductase.

    PubMed

    Barber, M J; Kay, C J

    1996-02-15

    Assimilatory NADH:nitrate reductase catalyzes the transfer of reducing equivalents from NADH to molecular oxygen. Initial rate studies performed under conditions of optimal pH (8.0) and constant ionic strength (mu = 0.2) revealed that the maximal rate of activity with molecular oxygen was 0.5% (0.44 mumol NADH consumed/min/nmol heme) with a Km for O2 of 586 microM. NADH:molecular oxygen reductase activity exhibited a pH optimum of 9.2, was inhibited by cyanide, and was unaffected by changes in ionic strength or the presence of phosphate ions. Spectroscopic studies indicated NADH:molecular oxygen reductase activity resulted in the production of the superoxide radical, detected as the formation of adrenochrome from epinephrine and by the formation of adrenochrome from epinephrine and by the reduction of nitroblue tetrazolium, both of which could be inhibited by the addition of superoxide dismutase and were unaffected by the addition of catalase. Direct observation of superoxide production using spin-trapping in combination with EPR spectroscopy resulted in the detection of the spin adduct 5.5-dimethyl-5-hydroxy-1-pyrrolidinyloxy (DMPO-OH). The formation of this spin adduct was abolished either in the absence of nitrate reductase, NADH, or DMPO or the the addition of superoxide dismutase or nitrate and was greatly reduced by the presence of cyanide. Inclusion of catalase or ethanol had no effect on the formation of the spin adduct. These results indicate that nitrate reductase can utilize molecular oxygen as an electron acceptor and that the product, O2.(-), is primarily generated via the Mopterin center.

  9. Neelaredoxin, an iron-binding protein from the syphilis spirochete, Treponema pallidum, is a superoxide reductase.

    PubMed

    Jovanović, T; Ascenso, C; Hazlett, K R; Sikkink, R; Krebs, C; Litwiller, R; Benson, L M; Moura, I; Moura, J J; Radolf, J D; Huynh, B H; Naylor, S; Rusnak, F

    2000-09-15

    Treponema pallidum, the causative agent of venereal syphilis, is a microaerophilic obligate pathogen of humans. As it disseminates hematogenously and invades a wide range of tissues, T. pallidum presumably must tolerate substantial oxidative stress. Analysis of the T. pallidum genome indicates that the syphilis spirochete lacks most of the iron-binding proteins present in many other bacterial pathogens, including the oxidative defense enzymes superoxide dismutase, catalase, and peroxidase, but does possess an orthologue (TP0823) for neelaredoxin, an enzyme of hyperthermophilic and sulfate-reducing anaerobes shown to possess superoxide reductase activity. To analyze the potential role of neelaredoxin in treponemal oxidative defense, we examined the biochemical, spectroscopic, and antioxidant properties of recombinant T. pallidum neelaredoxin. Neelaredoxin was shown to be expressed in T. pallidum by reverse transcriptase-polymerase chain reaction and Western blot analysis. Recombinant neelaredoxin is a 26-kDa alpha(2) homodimer containing, on average, 0.7 iron atoms/subunit. Mössbauer and EPR analysis of the purified protein indicates that the iron atom exists as a mononuclear center in a mixture of high spin ferrous and ferric oxidation states. The fully oxidized form, obtained by the addition of K(3)(Fe(CN)(6)), exhibits an optical spectrum with absorbances at 280, 320, and 656 nm; the last feature is responsible for the protein's blue color, which disappears upon ascorbate reduction. The fully oxidized protein has a A(280)/A(656) ratio of 10.3. Enzymatic studies revealed that T. pallidum neelaredoxin is able to catalyze a redox equilibrium between superoxide and hydrogen peroxide, a result consistent with it being a superoxide reductase. This finding, the first description of a T. pallidum iron-binding protein, indicates that the syphilis spirochete copes with oxidative stress via a primitive mechanism, which, thus far, has not been described in pathogenic

  10. Antibacterial Efficacy of Super-Oxidized Water on Enterococcus faecalis Biofilms in Root Canal

    PubMed Central

    Zan, Recai; Alacam, Tayfun; Hubbezoglu, Ihsan; Tunc, Tutku; Sumer, Zeynep; Alici, Oguzhan

    2016-01-01

    Background The success of endodontic treatment depends on a few crucial factors. One of these factors is the complete chemomechanic preparation of root canal against various bacteria. In particular, the effect of resistant bacteria may cause intense pain with flare-up and formation of periapical lesions. Therefore, the strong effect of irrigants plays an important role in terms of the complete elimination of these bacteria to achieve long-term successful treatment. Objectives The aim of this study was to investigate the antibacterial effects of super-oxidized water (SPO) in root canals infected with Enterococcus faecalis biofilms. Methods One hundred twenty single-root, premolar teeth were selected. Initially, the teeth were prepared and then disinfected. E. faecalis were inoculated and kept at 37°C for 24 hours in the root canals. The re-inoculation procedure was repeated on the first, fourth, seventh, and tenth days. The infected root canals were divided into one negative (saline) and one positive (sodium hypochlorite) control group and four experimental groups (super-oxidized water: 1, 2, 3, or 5 minutes) (n = 20). Paper points were placed in the root canals to control and evaluate the biofilm formation. Biofilms were counted on blood agar plates, and data was evaluated and statistically analyzed using one-way ANOVA and Tukey’s test. Results Although sodium hypochlorite (NaOCl) showed no statistically significant difference when compared with three and five minutes of SPO irrigation (P > 0.05), NaOCl showed statistically significant differences among all other groups (P < 0.05). Conclusions Super-oxidized water indicated a remarkable and similar bactericidal effect to that of traditional NaOCl against E. faecalis biofilms. In terms of successful endodontic treatment approaches, super-oxidized water may be used as an effective irrigation solution in clinics. PMID:27800142

  11. Methamphetamine toxicity is attenuated in mice that overexpress human manganese superoxide dismutase.

    PubMed

    Maragos, W F; Jakel, R; Chesnut, D; Pocernich, C B; Butterfield, D A; St Clair, D; Cass, W A

    2000-09-29

    We have investigated methamphetamine (MA) toxicity in transgenic mice that overexpress the human form of mitochondrial manganese superoxide dismutase (MnSOD). Our results reveal a significant reduction in the long-term depletion of striatal dopamine and protein oxidation following repeated administration of MA in transgenic vs. non-transgenic littermates. These findings support the notion that ROS contribute to MA-induced brain damage and suggest that mitochondria may play an important role in this form of neurodegeneration.

  12. Higher macrophage superoxide anion production in coronary artery disease (CAD) patients with Type D personality.

    PubMed

    Zuccarella-Hackl, Claudia; von Känel, Roland; Thomas, Livia; Kuebler, Peggy; Schmid, Jean-Paul; Mattle, Heinrich P; Mono, Marie-Louise; Rieben, Robert; Wiest, Roland; Wirtz, Petra H

    2016-06-01

    Type D personality (Type D) is an independent psychosocial risk factor for poor cardiac prognosis and increased mortality in patients with cardiovascular disease (CVD), but the involved mechanisms are poorly understood. Macrophages play a pivotal role in atherosclerosis, the process underlying coronary artery disease (CAD). We investigated macrophage superoxide anion production in production in CAD patients with and without Type D. We studied 20 male CAD patients with Type D (M:66.7±9.9years) and 20 age-matched male CAD patients without Type D (M:67.7±8.5years). Type D was measured using the DS14 questionnaire with the two subscales 'negative affectivity' and 'social inhibition'. We assessed macrophage superoxide anion production using the WST-1 assay. All analyses were controlled for potential confounders. CAD patients with Type D showed higher superoxide anion production compared to CAD patients without Type D (F(1,38)=15.57, p<0.001). Complementary analyses using the Type D subscales 'negative affectivity' and 'social inhibition', and their interaction as continuous measures, showed that both Type D subscales (negative affectivity: (ß=0.48, p=0.002, R(2)=0.227); social inhibition: (ß=0.46, p=0.003, R(2)=0.208)) and their interaction (ß=0.36, p=0.022, R(2)=0.130) were associated with higher WST-1 reduction scores. Results remained significant when controlling for classical CVD risk factors (i.e. body mass index, mean arterial blood pressure), atherosclerosis severity (i.e. intima media thickness, presence of carotid plaques), and psychological factors (depressive symptom severity, chronic stress). Our results indicate higher macrophage superoxide anion production in CAD patients with Type D compared to those without Type D. This may suggest a mechanism contributing to increased morbidity and mortality in CAD patients with Type D. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Superoxide and respiratory coupling in mitochondria of insulin-deficient diabetic rats.

    PubMed

    Herlein, Judith A; Fink, Brian D; O'Malley, Yunxia; Sivitz, William I

    2009-01-01

    Mitochondrial reactive oxygen species have been implicated in both diabetic complications and the progression of the underlying diabetic state. However, it is not clear whether mitochondria of diabetic origin are intrinsically altered to generate excess reactive oxygen species independent of the surrounding diabetic milieu. Mitochondria were isolated from gastrocnemius, heart, and liver of 2-wk and 2-month streptozotocin diabetic rats and controls. We rigidly quantified mitochondrial superoxide, respiration and ATP production, respiratory coupling, the expression of several proteins with antioxidant properties, and the redox state of glutathione. Both fluorescent assessment and electron paramagnetic spectroscopy revealed that superoxide production was unchanged or reduced in the 2-month diabetic mitochondria compared with controls. Kinetic analysis of the proton leak showed that diabetic heart and muscle mitochondria were actually more coupled compared with control despite an approximate 2- to 4-fold increase in uncoupling protein-3 content. Adenine nucleotide translocator type 1 expression was reduced by approximately 50% in diabetic muscle mitochondria. Catalase was significantly up-regulated in muscle and heart tissue and in heart mitochondria, whereas glutathione peroxidase expression was increased in liver mitochondria of diabetic rats. We conclude that gastrocnemius, heart, and liver mitochondria of streptozotocin diabetic rats are not irrevocably altered toward excess superoxide production either by complex I or complex III. Moreover, gastrocnemius and heart mitochondria demonstrate increased, not decreased, respiratory coupling. Mitochondria of insulin-deficient diabetic rats do show signs of adaptation to antecedent oxidative stress manifested as tissue-specific enzyme and uncoupling protein expression but remain remarkably robust with respect to superoxide production.

  14. Equine digital veins are more sensitive to superoxide anions than digital arteries.

    PubMed

    Lapo, Rock Allister; Gogny, Marc; Chatagnon, Gérard; Lalanne, Valérie; Harfoush, Khaled; Assane, Moussa; Desfontis, Jean-Claude; Mallem, Mohamed Yassine

    2014-10-05

    This work was designed to investigate (i) the effect of superoxide dismutase (SOD) inhibition on endothelial function and (ii) the free radical-induced endothelial dysfunction in equine digital veins (EDVs) and equine digital arteries (EDAs) isolated from healthy horses. EDV and EDA rings were suspended in a 5 ml organ bath containing Krebs solution. After a 60 min equilibration period, EDV and EDA rings were contracted with phenylephrine. Then, cumulative concentration-response curves (CCRCs) to acetylcholine were performed. In both EDVs and EDAs, acetylcholine (1 nM to 10 µM) produced concentration-dependent relaxation. We investigated the influence of SOD inhibition by diethyldithiocarbamate (DETC; 100 µM), a CuZnSOD inhibitor, on EDAs and EDVs relaxant responses to acetylcholine. Acetylcholine -mediated relaxation was impaired by DETC only in EDVs. SOD activity assayed by a xanthine-xanthine oxidase method was higher in EDAs compared with EDVs (P<0.05). CCRCs to acetylcholine established in the presence of pyrogallol (30 µM) or homocysteine (20 µM), two superoxide anions generating systems showed that in both EDVs and EDAs, the acetylcholine-mediated relaxation was significantly impaired by pyrogallol and homocysteine. This impairment was more pronounced in EDVs than in EDAs. Moreover, the pyrogallol-induced impairment of acetylcholine-mediated relaxation was potentiated by DETC to a greater extent in EDVs. We concluded that due to the lower activity of SOD, EDVs are more sensitive to superoxide anions than EDAs. So, any alteration of superoxide anions metabolism is likely to have a more important impact on venous rather than arterial relaxation.

  15. Superoxide and its metabolism during germination and axis growth of Vigna radiata (L.) Wilczek seeds.

    PubMed

    Singh, Khangembam Lenin; Chaudhuri, Abira; Kar, Rup Kumar

    2014-01-01

    Involvement of reactive oxygen species in regulation of plant growth and development is recently being demonstrated with various results depending on the experimental system and plant species. Role of superoxide and its metabolism in germination and axis growth was investigated in case of Vigna radiata seeds, a non-endospermous leguminous species having epigeal germination, by studying the effect of different reactive oxygen species (ROS) inhibitors, distribution of O2(•)- and H2O2 and ROS enzyme profile in axes. Germination percentage and axis growth were determined under treatment with ROS inhibitors and scavengers. Localization of O2(•)- and H2O2 was done using nitroblue tetrazolium (NBT) and 3,3',5,5'-tetramethyl benzidine dihydrochloride hydrate (TMB), respectively. Apoplastic level of O2(•)- was monitored by spectrophotometric analysis of bathing medium of axes. Profiles of NADPH oxidase and superoxide dismutase (SOD) were studied by in-gel assay. Germination was retarded by treatments affecting ROS level except H2O2 scavengers, while axis growth was retarded by all. Superoxide synthesis inhibitor and scavenger prevented H2O2 accumulation in axes in later phase as revealed from TMB staining. Activity of Cu/Zn SOD1 was initially high and declined thereafter. Superoxide being produced in apoplast possibly by NADPH oxidase activity is further metabolized to (•)OH via H2O2. Germination process depends possibly on (•)OH production in the axes. Post-germinative axis growth requires O2(•)- while the differentiating zone of axis (radicle) requires H2O2 for cell wall stiffening.

  16. Oxygen activation at the plasma membrane: relation between superoxide and hydroxyl radical production by isolated membranes.

    PubMed

    Heyno, Eiri; Mary, Véronique; Schopfer, Peter; Krieger-Liszkay, Anja

    2011-07-01

    Production of reactive oxygen species (hydroxyl radicals, superoxide radicals and hydrogen peroxide) was studied using EPR spin-trapping techniques and specific dyes in isolated plasma membranes from the growing and the non-growing zones of hypocotyls and roots of etiolated soybean seedlings as well as coleoptiles and roots of etiolated maize seedlings. NAD(P)H mediated the production of superoxide in all plasma membrane samples. Hydroxyl radicals were only produced by the membranes of the hypocotyl growing zone when a Fenton catalyst (FeEDTA) was present. By contrast, in membranes from other parts of the seedlings a low rate of spontaneous hydroxyl radical formation was observed due to the presence of small amounts of tightly bound peroxidase. It is concluded that apoplastic hydroxyl radical generation depends fully, or for the most part, on peroxidase localized in the cell wall. In soybean plasma membranes from the growing zone of the hypocotyl pharmacological tests showed that the superoxide production could potentially be attributed to the action of at least two enzymes, an NADPH oxidase and, in the presence of menadione, a quinone reductase.

  17. Superoxide radical formation by pure complex I (NADH:ubiquinone oxidoreductase) from Yarrowia lipolytica.

    PubMed

    Galkin, Alexander; Brandt, Ulrich

    2005-08-26

    Generation of reactive oxygen species (ROS) is increasingly recognized as an important cellular process involved in numerous physiological and pathophysiological processes. Complex I (NADH:ubiquinone oxidoreductase) is considered as one of the major sources of ROS within mitochondria. Yet, the exact site and mechanism of superoxide production by this large membrane-bound multiprotein complex has remained controversial. Here we show that isolated complex I from Yarrowia lipolytica forms superoxide at a rate of 0.15% of the rate measured for catalytic turnover. Superoxide production is not inhibited by ubiquinone analogous inhibitors. Because mutant complex I lacking a detectable iron-sulfur cluster N2 exhibited the same rate of ROS production, this terminal redox center could be excluded as a source of electrons. From the effect of different ubiquinone derivatives and pH on this side reaction of complex I we concluded that oxygen accepts electrons from FMNH2 or FMN semiquinone either directly or via more hydrophilic ubiquinone derivatives.

  18. A Zostera marina manganese superoxide dismutase gene involved in the responses to temperature stress.

    PubMed

    Liu, Jiao; Tang, Xuexi; Wang, You; Zang, Yu; Zhou, Bin

    2016-01-10

    Superoxide dismutase (SOD) is an essential enzyme playing a pivotal role in the protection mechanism against oxidative stress by reducing superoxide radicals. In the present study, the full-length cDNA sequence of manganese superoxide dismutase was identified from Zostera marina (ZmMnSOD) via raid amplification of cDNA ends (RACE) technique and expressed sequence tags (ESTs) analysis. The open reading frame (ORF) encoded a polypeptide of 254 amino acid residues, which shared 69%-77% similarity with previous identified SODs. Analysis of the deduced amino acid revealed conserved features, including functional domains, signature motifs and metal binding sites. Phylogenetic analysis revealed that ZmMnSOD was closer to the SODs from angiosperm than those from other organisms. The mRNA expression level of ZmMnSOD at different temperatures was investigated using real-time PCR and it was significantly up-regulated from 5°C to 15°C, and then dramatically down-regulated. The recombinant ZmMnSOD protein was purified and exhibited Mn(2+) ions dependency specific enzymatic activity and strong antioxidant activity over a wide temperature range. All these results indicate that ZmMnSOD is an authentic member of the plant SOD family and may play important roles in minimizing the effect of oxidative damage in Z. marina against temperature stress and affect the adaptability of Z. marina to global warming. Copyright © 2015 Elsevier B.V. All rights reserved.

  19. Rutin inhibits proliferation, attenuates superoxide production and decreases adhesion and migration of human cancerous cells.

    PubMed

    Ben Sghaier, Mohamed; Pagano, Alessandra; Mousslim, Mohamed; Ammari, Youssef; Kovacic, Hervé; Luis, José

    2016-12-01

    Lung and colorectal cancer are the principal causes of death in the world. Rutin, an active flavonoid compound, is known for possessing a wide range of biological activities. In this study, we examined the effect of rutin on the viability, superoxide anion production, adhesion and migration of human lung (A549) and colon (HT29 and Caco-2) cancer cell lines. In order to control the harmlessness of the tested concentrations of rutin, the viability of cancer cell lines was assessed using a 3-(4,5-dimethylthiazol- 2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) assay. ROS generation was measured by lucigenin chemiluminescence detecting superoxide ions. To investigate the effect of rutin on the behavior of human lung and colon cancer cell lines, we performed adhesion assays, using various purified extracellular matrix (ECM) proteins. Finally, in vitro cell migration assays were explored using modified Boyden chambers. The viability of cancerous cells was inhibited by rutin. It also significantly attenuated the superoxide production in HT29 cells. In addition, rutin affected adhesion and migration of A549 and HT29 cell. These findings indicate that rutin, a natural molecule, might have potential as anticancer agent against lung and colorectal carcinogenesis.

  20. Superoxide-hydrogen peroxide imbalance interferes with colorectal cancer cells viability, proliferation and oxaliplatin response.

    PubMed

    Azzolin, Verônica Farina; Cadoná, Francine Carla; Machado, Alencar Kolinski; Berto, Maiquidieli Dal; Barbisan, Fernanda; Dornelles, Eduardo Bortoluzzi; Glanzner, Werner Giehl; Gonçalves, Paulo Bayard; Bica, Claudia Giugliano; da Cruz, Ivana Beatrice Mânica

    2016-04-01

    The role of superoxide dismutase manganese dependent enzyme (SOD2) in colorectal cancer is presently insufficiently understood. Some studies suggest that high SOD2 levels found in cancer tissues are associated with cancer progression. However, thus far, the role of colorectal cancer superoxide-hydrogen peroxide imbalance has not yet been studied. Thus, in order to address this gap in extant literature, we performed an in vitro analysis using HT-29 colorectal cell line exposed to paraquat, which generates high superoxide levels, and porphyrin, a SOD2 mimic molecule. The effect of these drugs on colorectal cancer cell response to oxaliplatin was evaluated. At 0.1 μM concentration, both drugs exhibited cytotoxic and antiproliferative effect on colorectal cancer cells. However, this effect was more pronounced in cells exposed to paraquat. Paraquat also augmented the oxaliplatin cytotoxic and antiproliferative effects by increasing the number of apoptosis events, thus causing the cell cycle arrest in the S and M/G2 phases. The treatments were also able to differentially modulate genes related to apoptosis, cell proliferation and antioxidant enzyme system. However, the effects were highly variable and the results obtained were inconclusive. Nonetheless, our findings support the hypothesis that imbalance caused by increased hydrogen peroxide levels could be beneficial to cancer cell biology. Therefore, the use of therapeutic strategies to decrease hydrogen peroxide levels mainly during oxaliplatin chemotherapy could be clinically important to the outcomes of colorectal cancer treatment.

  1. Irradiation Enhances Hippocampus-Dependent Cognition in Mice Deficient in Extracellular Superoxide Dismutase

    PubMed Central

    Raber, Jacob; Villasana, Laura; Rosenberg, Jenna; Zou, Yani; Huang, Ting Ting; Fike, John R.

    2009-01-01

    The effects of ionizing irradiation on the brain are associated with oxidative stress. While oxidative stress following irradiation is generally viewed as detrimental for hippocampal function, it might have beneficial effects as part of an adaptive or preconditioning response to a subsequent challenge. Here we show that in contrast to what is seen in wild-type mice, irradiation enhances hippocampus-dependent cognitive measures in mice lacking extracellular superoxide dismutase. These outcomes were associated with genotype-dependent effects on measures of oxidative stress. When cortices and hippocampi were analyzed for nitrotyrosine formation as an index of oxidative stress, the levels were chronically elevated in mice lacking extracellular superoxide dismutase. However, irradiation caused a greater increase in nitrotyrosine levels in wild-type mice than mice lacking extracellular superoxide dismutase. These paradoxical genotype-dependent effects of irradiation on measures of oxidative stress and cognitive function underscore potential beneficial effects associated with chronic oxidative stress if it exists prior to a secondary insult such as irradiation. PMID:20020436

  2. The induction of manganese superoxide dismutase in response to stress in Nicotiana plumbaginifolia.

    PubMed Central

    Bowler, C; Alliotte, T; De Loose, M; Van Montagu, M; Inzé, D

    1989-01-01

    Superoxide dismutases (SODs) are metalloproteins that catalyse the dismutation of superoxide radicals to oxygen and hydrogen peroxide. The enzyme has been found in all aerobic organisms examined, where it plays a major role in the defence against toxic reduced oxygen species which are generated in many biological oxidations. Here we report the complete primary structure of a plant manganese superoxide dismutase (MnSOD), deduced from a cDNA clone of Nicotiana plumbaginifolia. The plant protein is highly homologous to MnSODs from other organisms and also contains an N-terminal leader sequence resembling a transit peptide for mitochondrial targeting. The location of the mature protein within the mitochondria has been demonstrated by subcellular fractionation experiments. We have analysed the expression profile of this MnSOD and found that it is dramatically induced during stress conditions, most notably in tissue culture as a result of sugar metabolism and also as part of the pathogenesis response of the plant, being induced by ethylene, salicylic acid, and Pseudomonas syringae infection. This induction is always accompanied by an increase in cytochrome oxidase activity, which suggests a specific protective role for MnSOD during conditions of increased mitochondrial respiration. Images PMID:2540959

  3. Desacetyluvaricin induces S phase arrest in SW480 colorectal cancer cells through superoxide overproduction.

    PubMed

    Xue, Jun-Yi; Zhou, Guang-Xiong; Chen, Tianfeng; Gao, Si; Choi, Mei-Yuk; Wong, Yum-Shing

    2014-03-01

    Annonaceous acetogenins (ACGs) are a group of fatty acid-derivatives with potent anticancer effects. In the present study, we found desacetyluvaricin (Dau) exhibited notable in vitro antiproliferative effect on SW480 human colorectal carcinoma cells with IC50 value of 14 nM. The studies on the underlying mechanisms revealed that Dau inhibited the cancer cell growth through induction of S phase cell cycle arrest from 11.3% (control) to 33.2% (160 nM Dau), which was evidenced by the decreased protein expression of cyclin A Overproduction of superoxide, intracellular DNA damage, and inhibition of MEK/ERK signaling pathway, were also found involved in cells exposed to Dau. Moreover, pre-treatment of the cells with ascorbic acid significantly prevented the Dau-induced overproduction of superoxide, DNA damage and cell cycle arrest. Taken together, our results suggest that Dau induces S phase arrest in cancer cells by firstly superoxide overproduction and subsequently the involvement of various signaling pathways.

  4. Extracellular superoxide dismutase is necessary to maintain renal blood flow during sepsis development.

    PubMed

    Constantino, Larissa; Galant, Letícia Selinger; Vuolo, Francieli; Guarido, Karla Lorena; Kist, Luiza Wilges; de Oliveira, Giovanna Medeiros Tavares; Pasquali, Matheus Augusto de Bittencourt; de Souza, Cláudio Teodoro; da Silva-Santos, José Eduardo; Bogo, Maurício Reis; Moreira, José Cláudio Fonseca; Ritter, Cristiane; Dal-Pizzol, Felipe

    2017-12-01

    Extracellular superoxide dismutase (ECSOD) protects nitric oxide (NO) bioavailability by decreasing superoxide levels and preventing peroxynitrite generation, which is important in maintaining renal blood flow and in preventing acute kidney injury. However, the profile of ECSOD expression after sepsis is not fully understood. Therefore, we intended to evaluate the content and gene expression of superoxide dismutase (SOD) isoforms in the renal artery and their relation to renal blood flow. Sepsis was induced in Wistar rats by caecal ligation and perforation. Several times after sepsis induction, renal blood flow (12, 24 and 48 h); the renal arterial content of SOD isoforms, nitrotyrosine, endothelial and inducible nitric oxide synthase (e-NOS and i-NOS), and phosphorylated vasodilator-stimulated phosphoprotein (pVASP); and SOD activity (3, 6 and 12 h) were measured. The influence of a SOD inhibitor was also evaluated. An increase in ECSOD content was associated with decreased 3-nitrotyrosine levels. These events were associated with an increase in pVASP content and maintenance of renal blood flow. Moreover, previous treatment with a SOD inhibitor increased nitrotyrosine content and reduced renal blood flow. ECSOD appears to have a major role in decreasing peroxynitrite formation in the renal artery during the early stages of sepsis development, and its application can be important in renal blood flow control and maintenance during septic insult.

  5. Concerted double proton-transfer electron-transfer between catechol and superoxide radical anion.

    PubMed

    Quintero-Saumeth, Jorge; Rincón, David A; Doerr, Markus; Daza, Martha C

    2017-09-20

    We have carried out a computational study on the reactivity of catechol (1,2-dihydroxybenzene) towards superoxide radical anion (O2˙(-)) in water, N,N-dimethylformamide (DMF), pentyl ethanoate (PEA) and vacuum using density functional theory and the coupled cluster method. Five reaction mechanisms were studied: (i) sequential proton transfer followed by hydrogen atom transfer (PT-HT), (ii) sequential hydrogen atom transfer followed by proton transfer (HT-PT), (iii) single electron transfer (SET), (iv) radical adduct formation (RAF) and (v) concerted double proton-transfer electron-transfer (denoted as global reaction, GR). Our results show that catechol and superoxide do not react via a sequential reaction mechanism (initial PT, initial HAT or SET). Instead, the reaction proceeds via a concerted double proton-transfer electron-transfer mechanism yielding hydrogen peroxide and catechol radical anion. The protons are transferred asynchronously between the σ orbitals of the catechol oxygen atoms to superoxide, while the electron is transferred between oxygen π orbitals in the same direction. The calculated rate constants in aqueous media agree with the experimental values reported in the literature. This suggests that the mechanism proposed in this work is adequate to describe this reaction. In addition, our results show that the reaction exhibits a large tunneling effect.

  6. Transient hyperoxic reoxygenation reduces cytochrome C oxidase activity by increasing superoxide dismutase and nitric oxide.

    PubMed

    Arab, Amina; Wang, Jin; Bausch, Kathrin; von Schmädel, Katharina; Bode, Christoph; Hehrlein, Christoph

    2010-04-09

    Oxygen therapies have been shown to be cytoprotective in a dose-dependent fashion. Previously, we have characterized the protective effects of moderate hyperoxia on cell viability of ischemic human cardiomyocytes and their mitochondrial membrane potential by transient addition of oxygenated perfluorocarbons to the cell medium. Now, we report that the activity and expression of cytochrome c oxidase (COX) after prolonged ischemia depend on the amount of oxygen delivered during reoxygenation. Transient hyperoxia during reoxygenation results in a decrease of COX activity by 62 +/- 15% and COX expression by 67 +/- 5%, when hyperoxic tensions of approximately = 300 mm Hg are reached in the cell medium. This decrease in COX expression is prevented by the inhibition of inducible nitric-oxide synthase (iNOS). Immunoblot analysis of ischemic human cardiomyocytes revealed that hyperoxic reoxygenation causes a 2-fold increase of iNOS, leading to a rise in nitric oxide production by 140 +/- 45%. Hyperoxic reoxygenation is further responsible for a 2-fold activation of hydrogen peroxide production and an increase in cytosolic superoxide dismutase expression by 35 +/- 10%. NADPH availability has no effect on the hyperoxia-induced decrease of superoxide. Overall, these results indicate that transient hyperoxic reoxygenation in optimal concentrations increases the level of nitric oxide by activation of iNOS and superoxide dismutase, thereby inducing respiration arrest in mitochondria of ischemic cardiomyocytes.

  7. Assessment of antioxidants status and superoxide dismutase activity in HIV-infected children.

    PubMed

    Pugliese, Camila; Patin, Rose Vega; Palchetti, Cecilia Zanin; Claudio, Cristiane Chiantelli; Gouvêa, Aída de Fátima Thomé Barbosa; Succi, Regina Célia de Menezes; Amancio, Olga Maria Silverio; Cozzolino, Silvia Maria Franciscato; Oliveira, Fernanda Luisa Ceragioli

    2014-01-01

    This study aims to assess the nutritional status of selenium, copper and zinc; and also the erythrocyte superoxide dismutase activity of HIV-infected children compared to a control group. A cross-sectional study was carried out with prepubertal HIV-infected children (n=51) and their healthy siblings (n=32). All biochemical measurements including plasma selenium, serum copper levels, serum and erythrocyte zinc levels and erythrocyte superoxide dismutase activity were evaluated according to dietary, clinical and biochemical parameters. Compared to the control group, the HIV-infected children had lower z-score values for height-for-age (p=0.0006), higher prevalence of stunting (11.8%) (p=0.047), lower selenium levels (p=0.0006) and higher copper levels (p=0.019). No difference was found concerning superoxide dismutase activity (p>0.05). The HIV-infected group presented a higher proportion (45.1%) of children with zinc intakes below the estimated average requirement (p=0.014); however, no association with zinc biochemical parameters was found. HIV-infected children have an inadequate selenium and copper nutritional status, which could influence the progression to AIDS. An adequate micronutrient status could improve the clinical conditions in these patients and minimize free radical production and cellular oxidative stress. Copyright © 2014. Published by Elsevier Editora Ltda.

  8. Superoxide Dismutase, Catalase, and α-Tocopherol Content of Stored Potato Tubers 1

    PubMed Central

    Spychalla, James P.; Desborough, Sharon L.

    1990-01-01

    Activated oxygen or oxygen free radical mediated damage to plants has been established or implicated in many plant stress situations. The extent of activated oxygen damage to potato (Solanum tuberosum L.) tubers during low temperature storage and long-term storage is not known. Quantitation of oxygen free radical mediated damage in plant tissues is difficult. However, it is comparatively easy to quantitate endogenous antioxidants, which detoxify potentially damaging forms of activated oxygen. Three tuber antioxidants, superoxide dismutase, catalase, and α-tocopherol were assayed from four potato cultivars stored at 3°C and 9°C for 40 weeks. Tubers stored at 3°C demonstrated increased superoxide dismutase activities (up to 72%) compared to tubers stored at 9°C. Time dependent increases in the levels of superoxide dismutase, catalase, and α-tocopherol occurred during the course of the 40 week storage. The possible relationship between these increases in antioxidants and the rate of activated oxygen production in the tubers is discussed. PMID:16667819

  9. Superoxide Dismutase, Catalase, and alpha-Tocopherol Content of Stored Potato Tubers.

    PubMed

    Spychalla, J P; Desborough, S L

    1990-11-01

    Activated oxygen or oxygen free radical mediated damage to plants has been established or implicated in many plant stress situations. The extent of activated oxygen damage to potato (Solanum tuberosum L.) tubers during low temperature storage and long-term storage is not known. Quantitation of oxygen free radical mediated damage in plant tissues is difficult. However, it is comparatively easy to quantitate endogenous antioxidants, which detoxify potentially damaging forms of activated oxygen. Three tuber antioxidants, superoxide dismutase, catalase, and alpha-tocopherol were assayed from four potato cultivars stored at 3 degrees C and 9 degrees C for 40 weeks. Tubers stored at 3 degrees C demonstrated increased superoxide dismutase activities (up to 72%) compared to tubers stored at 9 degrees C. Time dependent increases in the levels of superoxide dismutase, catalase, and alpha-tocopherol occurred during the course of the 40 week storage. The possible relationship between these increases in antioxidants and the rate of activated oxygen production in the tubers is discussed.

  10. Superoxide dismutase, catalase, and. alpha. -tocopherol content of stored potato tubers. [Solanum tuberosum L

    SciTech Connect

    Spychalla, J.P.; Desborough, S.L. )

    1990-11-01

    Activated oxygen or oxygen free radical mediated damage to plants has been established or implicated in many plant stress situations. The extent of activated oxygen damage to potato (Solanum tuberosum L.) tubers during low temperature storage and long-term storage is not known. Quantitation of oxygen free radical mediated damage in plant tissues is difficult. However, it is comparatively easy to quantitate endogenous antioxidants, which detoxify potentially damaging forms of activated oxygen. Three tuber antioxidants, superoxide dismutase, catalase, and {alpha}-tocopherol were assayed from four potato cultivars stored at 3{degree}C and 9{degree}C for 40 weeks. Tubers stored at 3{degree}C demonstrated increased superoxide dismutase activities (up to 72%) compared to tubers stored at 9{degree}C. Time dependent increases in the levels of superoxide dismutase, catalase, and {alpha}-tocopherol occurred during the course of the 40 week storage. The possible relationship between these increases in antioxidants and the rate of activated oxygen production in the tubers is discussed.

  11. Mitochondrial complex I deactivation is related to superoxide production in acute hypoxia.

    PubMed

    Hernansanz-Agustín, Pablo; Ramos, Elena; Navarro, Elisa; Parada, Esther; Sánchez-López, Nuria; Peláez-Aguado, Laura; Cabrera-García, J Daniel; Tello, Daniel; Buendia, Izaskun; Marina, Anabel; Egea, Javier; López, Manuela G; Bogdanova, Anna; Martínez-Ruiz, Antonio

    2017-08-01

    Mitochondria use oxygen as the final acceptor of the respiratory chain, but its incomplete reduction can also produce reactive oxygen species (ROS), especially superoxide. Acute hypoxia produces a superoxide burst in different cell types, but the triggering mechanism is still unknown. Herein, we show that complex I is involved in this superoxide burst under acute hypoxia in endothelial cells. We have also studied the possible mechanisms by which complex I could be involved in this burst, discarding reverse electron transport in complex I and the implication of PTEN-induced putative kinase 1 (PINK1). We show that complex I transition from the active to 'deactive' form is enhanced by acute hypoxia in endothelial cells and brain tissue, and we suggest that it can trigger ROS production through its Na(+)/H(+) antiporter activity. These results highlight the role of complex I as a key actor in redox signalling in acute hypoxia. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  12. Structure of glycosylated Cu/Zn-superoxide dismutase from Kluyveromyces yeast NBIMCC 1984

    NASA Astrophysics Data System (ADS)

    Dolashka-Angelova, Pavlina; Moshtanska, Vesela; Kujumdzieva, Anna; Atanasov, Boris; Petrova, Vencislava; Voelter, Wolfgang; Beeumen, Jozef Van

    2010-09-01

    The primary structure of Cu/Zn-superoxide dismutase from Kluyveromyces marxianus NBIMCC 1984 was elucidated by N-terminal sequence analysis of the intact protein and by determination of the amino acid sequences of tryptic peptides by MALDI-TOF-TOF tandem mass spectrometry. The molecular mass of one subunit of the homodimer SOD, containing 152 amino acid residues, was calculated to be 15858.3 Da while a value of 17096.63 Da was obtained by MALDI-TOF MS. This difference is explained by the presence of N-glycosylation of one linkage site, -Asn-Ile/Leu-Thr-, and a glycan chain with the structure Hex 5 GlcNAc 2. Glycosylation of K.marxianus superoxide dismutase is a post-translational modification. Recent developments in mass spectrometry have enabled detailed structural analyses of covalent modifications of proteins. Therefore, in this paper, we introduce a covalent modification of Cu/Zn-SOD from K. marxianus NBIMCC 1984, by analysis of the enzymatic liberated N-glycan from the enzyme using MALDI-TOF and tandem mass spectrometry on a Q-Trap mass spectrometer. This is the first report of the structure of the oligosaccharide of a naturally-glycosylated superoxide dismutase, determined by mass spectrometry.

  13. Feijoa sellowiana Berg fruit juice: anti-inflammatory effect and activity on superoxide anion generation.

    PubMed

    Monforte, Maria T; Fimiani, Vincenzo; Lanuzza, Francesco; Naccari, Clara; Restuccia, Salvatore; Galati, Enza M

    2014-04-01

    Feijoa sellowiana Berg var. coolidge fruit juice was studied in vivo for the anti-inflammatory activity by carrageenin-induced paw edema test and in vitro for the effects on superoxide anion release from neutrophils in human whole blood. The fruit juice was analyzed by the high-performance liquid chromatography method, and quercetin, ellagic acid, catechin, rutin, eriodictyol, gallic acid, pyrocatechol, syringic acid, and eriocitrin were identified. The results showed a significant anti-inflammatory activity of F. sellowiana fruit juice, sustained also by an effective antioxidant activity observed in preliminary studies on 1,1-diphenyl-2-picrylhydrazyl (DPPH) test. In particular, the anti-inflammatory activity edema inhibition is significant since the first hour (44.11%) and persists until the fifth hour (44.12%) of the treatment. The effect on superoxide anion release was studied in human whole blood, in the presence of activators affecting neutrophils by different mechanisms. The juice showed an inhibiting response on neutrophils basal activity in all experimental conditions. In stimulated neutrophils, the higher inhibition of superoxide anion generation was observed at concentration of 10(-4) and 10(-2) mg/mL in whole blood stimulate with phorbol-myristate-13-acetate (PMA; 20% and 40%) and with N-formyl-methionyl-leucyl-phenylalanine (FMLP; 15% and 48%). The significant reduction of edema and the inhibition of O2(-) production, occurring mainly through interaction with protein-kinase C pathway, confirm the anti-inflammatory effect of F. sellowiana fruit juice.

  14. Kinetics of superoxide scavenging by dismutase enzymes and manganese mimics determined by electron spin resonance.

    PubMed Central

    Gray, B; Carmichael, A J

    1992-01-01

    This study presents an e.s.r. assay for superoxide dismutase (SOD). Enzymic reactions were studied in which Cu,Zn-SOD, Mn-SOD and Fe-SOD each competed with the spin trap 5,5-dimethyl-1-pyrroline 1-oxide (DMPO) for superoxide anion (O2-) at pH 7.8 O2- from dissolved KO2 (potassium superoxide) in dimethyl sulphoxide was added directly to the enzyme solutions containing DMPO. The results show that, in this competition reaction system, the kinetics of the reactions between the enzymes and O2- follow a function y = f[( SOD]0.5). The rate constant, kSOD = 6.4 x 10(9) M-1. S-1, determined for Cu,Zn-SOD is approximately an order of magnitude larger than those for Mn-SOD and Fe-SOD. A comparative study of reported SOD mimics, including Mn2+, MnO2-desferrioxamine mesylate (Desferal) and MnO2-Desferal-ascorbate, was done. The results show that solutions of these complexes are approximately three orders of magnitude less active than Cu,Zn-SOD and approximately two orders of magnitude less active than Mn-SOD or Fe-SOD. The results also suggest that the reactivity toward O2- in solutions of these complexes originates from the Mn2+ present and not from the MnO2-Desferal complexes. PMID:1311175

  15. Cloning, Purification, and Characterization of Recombinant Human Extracellular Superoxide Dismutase in SF9 Insect Cells.

    PubMed

    Shrestha, Pravesh; Yun, Ji-Hye; Kim, Woo Taek; Kim, Tae-Yoon; Lee, Weontae

    2016-03-01

    A balance between production and degradation of reactive oxygen species (ROS) is critical for maintaining cellular homeostasis. Increased levels of ROS during oxidative stress are associated with disease conditions. Antioxidant enzymes, such as extracellular superoxide dismutase (EC-SOD), in the extracellular matrix (ECM) neutralize the toxicity of superoxide. Recent studies have emphasized the importance of EC-SOD in protecting the brain, lungs, and other tissues from oxidative stress. Therefore, EC-SOD would be an excellent therapeutic drug for treatment of diseases caused by oxidative stress. We cloned both the full length (residues 1-240) and truncated (residues 19-240) forms of human EC-SOD (hEC-SOD) into the donor plasmid pFastBacHTb. After transposition, the bacmid was transfected into the Sf9-baculovirus expression system and the expressed hEC-SOD purified using FLAG-tag. Western blot analysis revealed that hEC-SOD is present both as a monomer (33 kDa) and a dimer (66 kDa), as detected by the FLAG antibody. A water-soluble tetrazolium (WST-1) assay showed that both full length and truncated hEC-SOD proteins were enzymatically active. We showed that a potent superoxide dismutase inhibitor, diethyldithiocarbamate (DDC), inhibits hEC-SOD activity.

  16. Structure and gene expression of the E. coli Mn-superoxide dismutase gene.

    PubMed Central

    Takeda, Y; Avila, H

    1986-01-01

    Superoxide dismutase is an enzyme which converts superoxide O2- to hydrogen peroxide. Using a single synthetic oligonucleotide 33mer, we screened the E. coli DNA library and isolated a clone containing the E. coli manganese-superoxide dismutase gene. We determined the DNA sequence. The analysis of the DNA sequence and in vivo as well as in vitro transcription has shown the following. The DNA sequence suggests two possible promoters. However, only one of them seems active during normal aerobic growth. Purified RNA polymerase initiates in vitro transcription from the same promoter. It is not clear whether the second promoter is functional. It is possible that this promoter could be activated under different growth conditions. There is an inverted repeat sequence which could form a stem-loop structure downstream of the translation stop codon TAA of the Mn-SOD gene. The results of the analysis of in vivo and in vitro RNA have shown that this is the transcription termination signal. Thus, the Mn-SOD gene constitutes a single gene operon. There is an almost perfect 19 base palindrome at the -35 region. The position and the size of the palindrome suggest that this could be a regulatory site. Images PMID:3520487

  17. Synergistic triggering of superoxide flashes by mitochondrial Ca2+ uniport and basal reactive oxygen species elevation.

    PubMed

    Hou, Tingting; Zhang, Xing; Xu, Jiejia; Jian, Chongshu; Huang, Zhanglong; Ye, Tao; Hu, Keping; Zheng, Ming; Gao, Feng; Wang, Xianhua; Cheng, Heping

    2013-02-15

    Mitochondrial superoxide flashes reflect a quantal, bursting mode of reactive oxygen species (ROS) production that arises from stochastic, transient opening of the mitochondrial permeability transition pore (mPTP) in many types of cells and in living animals. However, the regulatory mechanisms and the exact nature of the flash-coupled mPTP remain poorly understood. Here we demonstrate a profound synergistic effect between mitochondrial Ca(2+) uniport and elevated basal ROS production in triggering superoxide flashes in intact cells. Hyperosmotic stress potently augmented the flash activity while simultaneously elevating mitochondrial Ca(2+) and ROS. Blocking mitochondrial Ca(2+) transport by knockdown of MICU1 or MCU, newly identified components of the mitochondrial Ca(2+) uniporter, or scavenging mitochondrial basal ROS markedly diminished the flash response. More importantly, whereas elevating Ca(2+) or ROS production alone was inefficacious in triggering the flashes, concurrent physiological Ca(2+) and ROS elevation served as the most powerful flash activator, increasing the flash incidence by an order of magnitude. Functionally, superoxide flashes in response to hyperosmotic stress participated in the activation of JNK and p38. Thus, physiological levels of mitochondrial Ca(2+) and ROS synergistically regulate stochastic mPTP opening and quantal ROS production in intact cells, marking the flash as a coincidence detector of mitochondrial Ca(2+) and ROS signals.

  18. Dietary copper supplements modulate aortic superoxide dismutase, nitric oxide and atherosclerosis

    PubMed Central

    Lamb, David J; Tickner, Michelle L; Hourani, Susanna M O; Ferns, Gordon A A

    2005-01-01

    The objective was to test the hypothesis that dietary copper inhibits atherosclerosis by inducing superoxide dismutase (SOD) and potentiating nitric oxide (NO). New Zealand White rabbits were fed either a cholesterol diet (n = 8) or a cholesterol diet containing 0.02% copper acetate (n = 8) for 13 weeks. We found that the intimal area was significantly smaller in the animals supplemented with copper (P < 0.005), although integrated plasma cholesterol levels were not significantly different. This was associated with a significant increase in aortic copper content (P < 0.05), SOD activity (P < 0.05) and Cu/Zn SOD mRNA (P < 0.05) and a significant decrease in nitrotyrosine content (P < 0.05). Furthermore, there was a positive correlation between aortic copper content and SOD activity (P < 0.005, R2 = 0.83) and a negative correlation between aortic superoxide dimutase activity and nitrotyrosine content (P < 0.005, R2 = 0.93). In organ bath experiments, the relaxation of precontracted carotid artery rings to calcium ionophore was greater in animals supplemented with copper. No difference in response to sodium nitroprusside was observed. These data suggest that in the cholesterol-fed rabbit, copper supplements inhibit the progression of atherosclerosis by increasing SOD expression, thereby reducing the interaction of NO with superoxide, and hence potentiating NO-mediated pathways that may protect against atherosclerosis. PMID:16045547

  19. Neurodegeneration, myocardial injury, and perinatal death in mitochondrial superoxide dismutase-deficient mice.

    PubMed Central

    Lebovitz, R M; Zhang, H; Vogel, H; Cartwright, J; Dionne, L; Lu, N; Huang, S; Matzuk, M M

    1996-01-01

    Manganese superoxide dismutase (SOD2) converts superoxide to oxygen plus hydrogen peroxide and serves as the primary defense against mitochondrial superoxide. Impaired SOD2 activity in humans has been associated with several chronic diseases, including ovarian cancer and type I diabetes, and SOD2 overexpression appears to suppress malignancy in cultured cells. We have produced a line of SOD2 knockout mice (SOD2m1BCM/SOD2m1BCM) that survive up to 3 weeks of age and exhibit several novel pathologic phenotypes including severe anemia, degeneration of neurons in the basal ganglia and brainstem, and progressive motor disturbances characterized by weakness, rapid fatigue, and circling behavior. In addition, SOD2m1BCM/SOD2m1BCM mice older than 7 days exhibit extensive mitochondrial injury within degenerating neurons and cardiac myocytes. Approximately 10% of SOD2m1BCM/SOD2m1BCM mice exhibit markedly enlarged and dilated hearts. These observations indicate that SOD2 deficiency causes increased susceptibility to oxidative mitochondrial injury in central nervous system neurons, cardiac myocytes, and other metabolically active tissues after postnatal exposure to ambient oxygen concentrations. Our SOD2-deficient mice differ from a recently described model in which homozygotes die within the first 5 days of life with severe cardiomyopathy and do not exhibit motor disturbances, central nervous system injury, or ultrastructural evidence of mitochondrial injury. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 PMID:8790408

  20. Stimulation of superoxide production increases fungicidal action of miconazole against Candida albicans biofilms

    PubMed Central

    De Cremer, Kaat; De Brucker, Katrijn; Staes, Ines; Peeters, Annelies; Van den Driessche, Freija; Coenye, Tom; Cammue, Bruno P. A.; Thevissen, Karin

    2016-01-01

    We performed a whole-transcriptome analysis of miconazole-treated Candida albicans biofilms, using RNA-sequencing. Our aim was to identify molecular pathways employed by biofilm cells of this pathogen to resist action of the commonly used antifungal miconazole. As expected, genes involved in sterol biosynthesis and genes encoding drug efflux pumps were highly induced in biofilm cells upon miconazole treatment. Other processes were affected as well, including the electron transport chain (ETC), of which eight components were transcriptionally downregulated. Within a diverse set of 17 inhibitors/inducers of the transcriptionally affected pathways, the ETC inhibitors acted most synergistically with miconazole against C. albicans biofilm cells. Synergy was not observed for planktonically growing C. albicans cultures or when biofilms were treated in oxygen-deprived conditions, pointing to a biofilm-specific oxygen-dependent tolerance mechanism. In line, a correlation between miconazole’s fungicidal action against C. albicans biofilm cells and the levels of superoxide radicals was observed, and confirmed both genetically and pharmacologically using a triple superoxide dismutase mutant and a superoxide dismutase inhibitor N-N′-diethyldithiocarbamate, respectively. Consequently, ETC inhibitors that result in mitochondrial dysfunction and affect production of reactive oxygen species can increase miconazole’s fungicidal activity against C. albicans biofilm cells. PMID:27272719

  1. The Basic Understanding of Lithium Superoxide in Li-O2 Battery

    NASA Astrophysics Data System (ADS)

    Lau, Kah Chun; Zhai, Dengyun; Wang, Hsien-Hau; Luo, Xiangyi; Wen, Jianguo; Miller, Dean; Redfern, Paul; Lu, Jun; Curtiss, Larry; Amine, Khalil

    The electrochemical and chemical processes that involved in Li-O2 battery are complex, and depend heavily on electrode materials, electrolytes, interfaces, and cell operating conditions. In non-aqueous Li-O2 battery, the main discharge products are commonly known to be lithium peroxide (Li2O2) , and possibly some other parasitic components (i.e. Li2CO3, LiOH, Li2O). However, the superoxide intermediates and lithium superoxide (O2-, LiO2) which are commonly known to be metastable can also be found as reported. Relative to these compounds (i.e. Li2CO3, Li2O,LiOH,Li2O2) in discharge products, little is known about LiO2. To have a basic understanding of lithium superoxide, both theoretical studies and experimental characterizations are important. In this presentation, the recent developments, studies and findings of this exotic species will be discussed. This work was primarily supported by the U.S. Department of Energy under Contract DE-AC02-06CH11357 from the Vehicle Technologies Office, Department of Energy, Office of Energy Efficiency and Renewable Energy.

  2. Novel synthesis approach for stable sodium superoxide (NaO2) nanoparticles for LPG sensing application

    NASA Astrophysics Data System (ADS)

    Nemade, Kailash; Waghuley, Sandeep

    2017-05-01

    The synthesis of stable superoxide is still great challenge for the researchers working in the field of materials science. Through this letter, we report the novel and simple synthesis approach for the preparation of stable sodium superoxide (NaO2) nanoparticles. NaO2 nanoparticles were prepared by a spray pyrolysis technique, under oxygen rich environment for gas sensing application. The texture characterizations show that as-obtained NaO2 nanoparticles have high structural purity. Most importantly, NaO2 nanoparticles exhibits higher sensing response, shorter response time and recovery time, low operating temperature and good stability during sensing of liquefied petroleum gas (LPG). The main accomplishment of present work is that as-fabricated sensor has low operating temperature (423 K), which is below auto-ignition temperature of LPG. The gas sensing mechanism of NaO2 nanoparticles was discussed without the conventional oxygen bridging mechanism. Through this short communication, LPG sensing application of stable sodium superoxide nanoparticle is explored.

  3. Topical Application of TAT-Superoxide Dismutase in Acupoints LI 20 on Allergic Rhinitis

    PubMed Central

    Zheng, Mei-Feng; Zhou, Jian-Wu; Ke, Li-Jing

    2016-01-01

    Reactive oxygen species are products of cellular metabolism and assigned important roles in biomedical science as deleterious factors in pathologies. In fact, some studies have shown that the therapeutic benefits of taking antioxidants were limited and the potential for therapeutic intervention remains unclear. New evidences showed that ROS have some ability of intercellular transportation. For treating allergic rhinitis, as a novel intracellular superoxide quencher, TAT-SOD applied to acupoints LI 20 instead of directly to nasal cavity can be used to test that. TTA group apply TAT-SOD cream prepared by adding purified TAT-SOD to the vehicle cream to acupoints LI 20, while placebo group used the vehicle cream instead. TTN group applied the same TAT-SOD cream directly to nasal cavity three times daily. Symptom scores were recorded at baseline and days 8 and 15. For the overall efficacy rate, TTA group was 81.0%, while placebo group was 5.9% and TTN was 0%. Malondialdehyde levels decreased observably in TTA group, and superoxide dismutase, catalase, and glutathione peroxidase levels remained basically unaffected. Enzymatic scavenging of the intracellular superoxide at acupoints LI 20 proved to be effective in treating allergic rhinitis, while no improvement was observed with the placebo group and TTN group. PMID:28119757

  4. Induction and inactivation of catalase and superoxide dismutase of Escherichia coli by ozone

    SciTech Connect

    Whiteside, C.; Hassan, H.M.

    1987-09-01

    Oxyradicals have been implicated in ozone (O/sub 3/) toxicity and in other oxidant stress. In this study, we investigated the effects of O/sub 3/ on the biosynthesis of the antioxidant enzymes catalase and superoxide dismutase in Escherichia coli to determine their role in the defense against ozone toxicity. Inhibition of growth and loss of viability were observed in cultures exposed to ozone. Results also showed an increase in the activities of catalase and superoxide dismutase in cultures exposed to ozone, which was shown to be due to true induction rather than activation of preexisting apoproteins. Cessation of O/sub 3/ exposure resulted in 30 min of continual high rate of catalase biosynthesis followed by a gradual decrease in the level of the enzyme approaching that of control cultures. This decrease was attributed to a concomitant cessation of de novo enzyme synthesis and dilution of preexisting enzyme by cellular growth. Ozonation of cell-free extracts showed that superoxide dismutase and catalase are subject to oxidative inactivation by ozone. In vivo induction of these enzymes may represent an adaptive response evolved to protect cells against ozone toxicity.

  5. Quantitative determination of superoxide in plant leaves using a modified NBT staining method.

    PubMed

    Bournonville, Carlos F Grellet; Díaz-Ricci, Juan Carlos

    2011-01-01

    In plants, the ROS (reactive oxygen species) level is tightly regulated because their accumulation produces irreversible damage leading to cell death. However, ROS accumulation plays a key role in plant signaling under biotic or abiotic stress. Although various methods were reported to evaluate ROS accumulation, they are restricted to model plants or provide only qualitative information. Develop a simple method to quantify superoxide radicals produced in plant tissues, based on the selective extraction of the formazan produced after nitroblue tetrazolium (NBT) reduction in histochemical staining. Plant leaves were stained with a standard NBT method and the formazan precipitated in tissues was selectively extracted using chloroform. The organic phase was dried and formazan residue dissolved in dimethylsulfoxide-potassium hydroxide and quantified by spectrophotometry. The method was tested in strawberry plant leaves under different stressing conditions. Formazan extracted from leaves subjected to stress conditions showed similar absorption spectra to those obtained from standard solutions using pure formazan. Calibration curves showed a linear relationship between absorbance and formazan amounts, within the range 0.5-8 µg. Outcomes suggested that formazan was retained in the solid residue of leaf tissues. This protocol allowed us to quantify superoxide radicals produced under different stress conditions. Chloroform allowed a selective formazan extraction and removal of potential endogenous, exogenous or procedural artefacts that may interfere with the quantitative determination. This protocol can be used to quantify the superoxide produced in plant tissues using any traditional qualitative NBT histochemical staining method. Copyright © 2011 John Wiley & Sons, Ltd.

  6. Nitric Oxide and Superoxide Anion Balance in Rats Exposed to Chronic and Long Term Intermittent Hypoxia

    PubMed Central

    Siques, Patricia; López de Pablo, Ángel Luis; Brito, Julio; Arribas, Silvia M.; Naveas, Nelson; González, M. Carmen; León-Velarde, Fabiola; López, M. Rosario

    2014-01-01

    Work at high altitude in shifts exposes humans to a new form of chronic intermittent hypoxia, with still unknown health consequences. We have established a rat model resembling this situation, which develops a milder form of right ventricular hypertrophy and pulmonary artery remodelling compared to continuous chronic exposure. We aimed to compare the alterations in pulmonary artery nitric oxide (NO) availability induced by these forms of hypoxia and the mechanisms implicated. Rats were exposed for 46 days to normoxia or hypobaric hypoxia, either continuous (CH) or intermittent (2 day shifts, CIH2x2), and assessed: NO and superoxide anion availability (fluorescent indicators and confocal microscopy); expression of phosphorylated endothelial NO synthase (eNOS), NADPH-oxidase (p22phox), and 3-nitrotyrosine (western blotting); and NADPH-oxidase location (immunohistochemistry). Compared to normoxia, (1) NO availability was reduced and superoxide anion was increased in both hypoxic groups, with a larger effect in CH, (2) eNOS expression was only reduced in CH, (3) NADPH-oxidase was similarly increased in both hypoxic groups, and (4) 3-nitrotyrosine was increased to a larger extent in CH. In conclusion, intermittent hypoxia reduces NO availability through superoxide anion destruction, without reducing its synthesis, while continuous hypoxia affects both, producing larger nitrosative damage which could be related to the more severe cardiovascular alterations. PMID:24719876

  7. Nanostructured cobalt phosphates as excellent biomimetic enzymes to sensitively detect superoxide anions released from living cells.

    PubMed

    Wang, Min-Qiang; Ye, Cui; Bao, Shu-Juan; Xu, Mao-Wen; Zhang, Yan; Wang, Ling; Ma, Xiao-Qing; Guo, Jun; Li, Chang-Ming

    2017-01-15

    Monitoring superoxide anion radicals in living cells has been attracting much academic and industrial interest due to the dual roles of the radicals. Herein, we synthesized a novel nanostructured cobalt phosphate nanorods (Co3(PO4)2 NRs) with tunable pore structure using a simple and effective micro-emulsion method and explored their potential utilization in the electrochemical sensing of superoxide anions. As an analytical and sensing platform, the nanoscale biomimetic enzymes Co3(PO4)2 NRs exhibited excellent selectivity and sensitivity towards superoxide anion (O2(•-)) with a low detection limit (2.25nM), wide linear range (5.76-5396nM), and long-term stability. Further, the nanoscale biomimetic enzyme could be efficiently applied in situ to electrochemically detect O2(•-) released from human malignant melanoma cells and normal keratinocyte, showing excellent real time quantitative detection capability. This material open up exciting opportunities for implementing biomimetic enzymes in nanoscale transition metal phosphates and designing enzyme-free biosensors with much higher sensitivity and durability in health and disease analysis than those of natural one.

  8. Purification and characterization of iron-cofactored superoxide dismutase from Enteromorpha linza

    NASA Astrophysics Data System (ADS)

    Lü, Mingsheng; Cai, Ruanhong; Wang, Shujun; Liu, Zhaopu; Jiao, Yuliang; Fang, Yaowei; Zhang, Xiaoxin

    2013-11-01

    A superoxide dismutase was purified from Enteromorpha linza using a simple and safe procedure, which comprised phosphate buffer extraction, ammonium sulphate precipitation, ion exchange chromatography on Q-sepharose column, and gel filtration chromatography on Superdex 200 10/300GL. The E. linza superoxide dismutase ( ElSOD) was purified 103.6-fold, and a yield of 19.1% and a specific activity of 1 750 U/mg protein were obtained. The SDS-PAGE exhibited ElSOD a single band near 23 kDa and the gel filtration study showed ElSOD's molecular weight is near 46 kDa in nondenatured condition, indicating it's a homodimeric protein. El SOD is an iron-cofactored superoxide dismutase (Fe-SOD) because it was inhibited by hydrogen peroxide, insensitive to potassium cyanide. The optimal temperature for its maximal enzyme activity was 35°C, and it still had 29.8% relative activity at 0°C, then ElSOD can be classified as a cold-adapted enzyme. ElSOD was stable when temperature was below 40°C or the pH was within the range of 5-10. The first 11 N-terminal amino acids of ElSOD were ALELKAPPYEL, comparison of its N-terminal sequence with other Fe-SOD N-terminal sequences at the same position suggests it is possibly a chloroplastic Fe-SOD.

  9. Chlorovirus PBCV-1 Encodes an Active Copper-Zinc Superoxide Dismutase

    PubMed Central

    Kang, Ming; Duncan, Garry A.; Kuszynski, Charles; Oyler, George; Zheng, Jiayin; Becker, Donald F.

    2014-01-01

    ABSTRACT Superoxide dismutases (SODs) are metalloproteins that protect organisms from toxic reactive oxygen species by catalyzing the conversion of superoxide anion to hydrogen peroxide and molecular oxygen. Chlorovirus PBCV-1 encodes a 187-amino-acid protein that resembles a Cu-Zn SOD with all of the conserved amino acid residues for binding copper and zinc (named cvSOD). cvSOD has an internal Met that results in a 165-amino-acid protein (named tcvSOD). Both cvSOD and tcvSOD recombinant proteins inhibited nitroblue tetrazolium reduction of superoxide anion generated in a xanthine-xanthine oxidase system in solution. tcvSOD was chosen for further characterization because it was easier to produce. Recombinant tcvSOD also inhibited a riboflavin photochemical reduction system in a polyacrylamide gel assay, which was blocked by the Cu-Zn SOD inhibitor cyanide but not by azide, which inhibits Fe and Mn SODs. A kcat/Km value for cvSOD was determined by stop-flow spectrophotometry as 1.28 × 108 M−1 s−1, suggesting that cvSOD-catalyzed O2− dismutation was not a diffusion controlled encounter. The cvsod gene was expressed as a late gene, and cvSOD activity was detected in purified virions. Superoxide accumulated rapidly during virus infection, and circumstantial evidence indicates that cvSOD aids its decomposition to benefit virus replication. Cu-Zn SOD homologs have been described to occur in 3 other families of large DNA viruses, poxviruses, baculoviruses, and mimiviruses, which group as a clade. Interestingly, cvSOD does not group in the same clade as the other virus SODs but instead groups in an expanded clade that includes Cu-Zn SODs from many cellular organisms. IMPORTANCE Virus infection often leads to an increase in toxic reactive oxygen species in the host, which can be detrimental to virus replication. Viruses have developed various ways to overcome this barrier. As reported in this article, the chloroviruses often encode and package a functional Cu

  10. The design of synthetic superoxide dismutase mimetics: seven-coordinate water soluble manganese(ii) and iron(ii) complexes and their superoxide dismutase-like activity studies.

    PubMed

    Singh, Ovender; Tyagi, Nidhi; Olmstead, Marilyn M; Ghosh, Kaushik

    2017-10-06

    Bio-inspired manganese [Mn(N5Py)(H2O)(CH3OH)](ClO4)2 (1) and iron [Fe(N5Py)(H2O)(ClO4)]ClO4 (2) complexes derived from a pentadentate ligand (N5Py = 2,6-bis((E)-1-phenyl-2-(pyridin-2-ylmethylene)hydrazinyl)pyridine) framework containing a N5 binding motif were synthesized and characterized using different spectroscopic methods. The molecular structures of complexes 1 and 2 were determined by X-ray crystallography. These complexes were found to be stable under physiological conditions and exhibited an excellent superoxide dismutase (SOD) activity. The SOD activity was determined by a xanthine-xanthine oxidase-nitro blue tetrazolium assay and the IC50 values were determined to be 1.53 and 2.09 μM, respectively.

  11. Compartmentalized oxidative stress in dopaminergic cell death induced by pesticides and complex I inhibitors: Distinct roles of superoxide anion and superoxide dismutases

    PubMed Central

    Rodriguez-Rocha, Humberto; Garcia-Garcia, Aracely; Pickett, Chillian; Sumin, Li; Jones, Jocelyn; Chen, Han; Webb, Brian; Choi, Jae; Zhou, You; Zimmerman, Matthew C.; Franco, Rodrigo

    2013-01-01

    The loss of dopaminergic neurons induced by the parkinsonian toxins paraquat, rotenone and 1-methyl-4-phenylpyridinium (MPP+) is associated with oxidative stress. However, controversial reports exist regarding the source/compartmentalization of reactive oxygen species (ROS) generation and its exact role in cell death. We aimed to determine in detail the role of superoxide anion (O2•−), oxidative stress and their subcellular compartmentalization in dopaminergic cell death induced by parkinsonian toxins. Oxidative stress and ROS formation was determined in the cytosol, intermembrane (IMS) and mitochondrial matrix compartments, using dihydroethidine derivatives, the redox sensor roGFP, as well as electron paramagnetic resonance spectroscopy. Paraquat induced an increase in ROS and oxidative stress in both the cytosol and mitochondrial matrix prior to cell death. MPP+ and rotenone primarily induced an increase in ROS and oxidative stress in the mitochondrial matrix. No oxidative stress was detected at the level of the IMS. In contrast to previous studies, overexpression of manganese superoxide dismutase (MnSOD) or copper/zinc SOD (CuZnSOD) had no effect on ROS steady state levels, lipid peroxidation, loss of mitochondrial membrane potential (ΔΨm) and dopaminergic cell death induced by MPP+ or rotenone. In contrast, paraquat-induced oxidative stress and cell death were selectively reduced by MnSOD overexpression, but not by CuZnSOD or manganese-porphyrins. However, MnSOD also failed to prevent ΔΨm loss. Finally, paraquat, but not MPP+ or rotenone, induced the transcriptional activation the redox-sensitive antioxidant response elements (ARE) and nuclear factor kappa-B (NF-κB). These results demonstrate a selective role of mitochondrial O2•− in dopaminergic cell death induced by paraquat, and show that toxicity induced by the complex I inhibitors rotenone and MPP+ does not depend directly on mitochondrial O2•− formation. PMID:23602909

  12. The involvement of superoxide and iron ions in the NADPH-dependent lipid peroxidation in human placental mitochondria.

    PubMed

    Klimek, J

    1988-01-19

    Incubation of human term placental mitochondria with Fe2+ and a NADPH-generating system initiated high levels of lipid peroxidation, as measured by the production of malondialdehyde. Malondialdehyde formation was accompanied by a corresponding decrease of the unsaturated fatty acid content. This NADPH-dependent lipid peroxidation was strongly inhibited by superoxide dismutase and singlet oxygen scavengers, markedly stimulated by paraquat, but was not affected by hydroxyl radical scavengers. Catalase enhanced the production of malondialdehyde by placental mitochondria. The effects of catalase and hydroxyl radical scavengers suggest that the initiation of NADPH-dependent lipid peroxidation is not dependent upon the hydroxyl radical produced via an iron-catalyzed Fenton reaction. These studies provide evidence that hydrogen peroxide strongly inhibits NADPH-dependent mitochondrial lipid peroxidation. The inhibitory effect of superoxide dismutase and stimulatory effect of paraquat, which was abolished by the addition of superoxide dismutase, suggests that superoxide may promote NADPH-dependent lipid peroxidation in human placental mitochondria.

  13. Activation of Mitochondrial Uncoupling Protein 4 and ATP-Sensitive Potassium Channel Cumulatively Decreases Superoxide Production in Insect Mitochondria.

    PubMed

    Slocińska, Malgorzata; Rosinski, Grzegorz; Jarmuszkiewicz, Wieslawa

    2016-01-01

    It has been evidenced that mitochondrial uncoupling protein 4 (UCP4) and ATP-regulated potassium channel (mKATP channel) of insect Gromphadorhina coqereliana mitochondria decrease superoxide anion production. We elucidated whether the two energy-dissipating systems work together on a modulation of superoxide level in cockroach mitochondria. Our data show that the simultaneous activation of UCP4 by palmitic acid and mKATP channel by pinacidil revealed a cumulative effect on weakening mitochondrial superoxide formation. The inhibition of UCP4 by GTP (and/or ATP) and mKATP channel by ATP elevated superoxide production. These results suggest a functional cooperation of both energy-dissipating systems in protection against oxidative stress in insects.

  14. [Superoxide dismutase and catalase activities in carotenoid-synthesizing fungi Blakeslea trispora and Neurospora crassa under the oxidative stress].

    PubMed

    Gessler, N N; Sokolov, A V; Bykhovskiĭ, V Ia; Belozerskaia, T A

    2002-01-01

    The addition of menadione into the medium during cultivation of Neurospora crassa in the dark activated its constitutive superoxide dismutase. Exposure to light not only activated superoxide dismutase and catalase, but also increased the content of neurosporaxanthin. Superoxide dismutase activity in the mixed (+/-) mycelium of Blakeslea trispora synthesizing beta-carotene in the dark was much lower than that in Neurospora crassa. The superoxide dismutase activity further decreased in oxidative stress. The catalase activity decreased with an increase in the content of beta-carotene. Our results indicate that neurosporaxanthin possesses photoprotective properties in Neurospora crassa. In Blakeslea trispora (+/-) fungi, this compound acts as a major antioxidant during inactivation of enzymes that detoxify reactive oxygen species.

  15. A two-photon fluorescent probe for exogenous and endogenous superoxide anion imaging in vitro and in vivo.

    PubMed

    Li, Run-Qing; Mao, Zhi-Qiang; Rong, Lei; Wu, Nian; Lei, Qi; Zhu, Jing-Yi; Zhuang, Lin; Zhang, Xian-Zheng; Liu, Zhi-Hong

    2017-01-15

    Herein, we report a novel quinoline derivative-based two-photon fluorescent probe 6-(dimethylamino)quinoline-2-benzothiazoline (HQ), which is capable of tracking superoxide anion in organisms with specific "turn-on" fluorescence response based on extension of π-conjugations and moderate ICT process. The probe exhibited favorable photophysical properties, a broad linear range and high photostability. It can specifically detect superoxide anion with a significant fluorescence enhancement and great linearity from 0 to 500μM in PBS buffer. Furthermore, HQ shows low cytotoxicity and excellent photostability toward living cells and organisms, which was able to monitor endogenous superoxide anion fluxes in living cells and in vivo. For the first time, endogenous superoxide anion in lung inflammation was visualized successfully by using HQ through two-photon microscopy, and the probe HQ shows great potential for fast in-situ detecting of inflammatory response in live organisms.

  16. Overproduction of Petunia Chloroplastic Copper/Zinc Superoxide Dismutase Does Not Confer Ozone Tolerance in Transgenic Tobacco 1

    PubMed Central

    Pitcher, Lynne H.; Brennan, Eileen; Hurley, Arthur; Dunsmuir, Pamela; Tepperman, James M.; Zilinskas, Barbara A.

    1991-01-01

    Transgenic tobacco (Nicotiana tabacum cultivar W38) plants that overproduce petunia chloroplastic Cu/Zn superoxide dismutase were exposed to ozone dosages that injure control tobacco plants. Based on foliar injury ratings, there was no consistent protection provided to the transgenic plants. These data indicate that an increase in the chloroplastic Cu/Zn superoxide dismutase alone is not sufficient to reduce ozone toxicity. ImagesFigure 1 PMID:16668407

  17. Hydroethidine- and Mito-SOX-derived red fluorescence is not a reliable indicator of intracellular superoxide formation: Another inconvenient truth

    PubMed Central

    Zielonka, Jacek; Kalyanaraman, B.

    2010-01-01

    Hydroethidine (or dihydroethidium) (HE) is the most popular fluorogenic probe used for detecting intracellular superoxide radical anion. The reaction between superoxide and HE generates a highly specific red fluorescent product, 2-hydroxyethidium (2-OH-E+). In biological systems, another red fluorescent product, ethidium (E+), is also formed, usually at a much higher concentration than 2-OH-E+. In this article, we have reviewed the methods to selectively detect the superoxide-specific product (2-OH-E+) and the factors affecting its levels in cellular and biological systems. The most important conclusion of the present review is that it is nearly impossible to assess the intracellular levels of the superoxide specific product, 2-OH-E+, using the confocal microscopy or other fluorescence-based microscopic assays and that it is essential to measure by HPLC the intracellular HE and other oxidation products of HE, in addition to 2-OH-E+, in order to fully understand the origin of red fluorescence. The chemical reactivity of mitochondria-targeted hydroethidine (Mito-HE, MitoSOX Red ®) with superoxide is similar to the reactivity of HE with superoxide and therefore, all of the limitations attributed to the HE assay are applicable to Mito-HE (or Mito-SOX) as well. PMID:20116425

  18. Production of superoxide radical in reductive metabolism of a synthetic food-coloring agent, indigocarmine, and related compounds.

    PubMed

    Kohno, Yoichi; Kitamura, Shigeyuki; Yamada, Tsuyoshi; Sugihara, Kazumi; Ohta, Shigeru

    2005-06-24

    Indigocarmine, which is widely used as a synthetic colouring agent for foods and cosmetics in many countries, was reduced to its leuco form and decolorized by rat liver microsomes with NADPH under anaerobic conditions. The reductase activity was enhanced in liver microsomes of phenobarbital-treated rats, and inhibited by diphenyliodonium chloride, a NADPH-cytochrome P450 reductase (P450 reductase) inhibitor, but was not inhibited by SKF 525-A or carbon monoxide. Indigocarmine reductase activity was exhibited by purified rat P450 reductase. In contrast, when indigocarmine was incubated with rat liver microsomes and NADPH under aerobic conditions, superoxide radical was produced and its production was inhibited by superoxide dismutase and diphenyliodonium chloride. When indigocarmine was incubated with purified rat P450 reductase in the presence of NADPH, superoxide radical production was enhanced 17.7-fold (similar to the enhancement of indigocarmine-reducing ability) as compared with that of rat liver microsomes. A decrease of one molecule of NADPH was accompanied with formation of about two molecules of superoxide radical. P450 reductase exhibited little reductase activity towards indigo and tetrabromoindigo, which also afforded little superoxide radical under aerobic conditions. These results indicate that indigocarmine is reduced by P450 reductase to its leuco form, and superoxide radical is produced by autoxidation of the leuco form, through a mechanism known as futile redox cycling.

  19. Roles of superoxide and myeloperoxidase in ascorbate oxidation in stimulated neutrophils and H2O2-treated HL60 cells.

    PubMed

    Parker, Amber; Cuddihy, Sarah L; Son, Tae G; Vissers, Margreet C M; Winterbourn, Christine C

    2011-10-01

    Ascorbate is present at high concentrations in neutrophils and becomes oxidized when the cells are stimulated. We have investigated the mechanism of oxidation by studying cultured HL60 cells and isolated neutrophils. Addition of H(2)O(2) to ascorbate-loaded HL60 cells resulted in substantial oxidation of intracellular ascorbate. Oxidation was myeloperoxidase-dependent, but not attributable to hypochlorous acid, and can be explained by myeloperoxidase (MPO) exhibiting direct ascorbate peroxidase activity. When neutrophils were stimulated with phorbol myristate acetate, about 40% of their intracellular ascorbate was oxidized over 20 min. Ascorbate loss required NADPH oxidase activity but in contrast to the HL60 cells did not involve myeloperoxidase. It did not occur when exogenous H(2)O(2) was added, was not inhibited by myeloperoxidase inhibitors, and was the same for normal and myeloperoxidase-deficient cells. Neutrophil ascorbate loss was enhanced when endogenous superoxide dismutase was inhibited by cyanide or diethyldithiocarbamate and appears to be due to oxidation by superoxide. We propose that in HL60 cells, MPO-dependent ascorbate oxidation occurs because cellular ascorbate can access newly synthesized MPO before it becomes packaged in granules: a mechanism not possible in neutrophils. In neutrophils, we estimate that ascorbate is capable of competing with superoxide dismutase for a small fraction of the superoxide they generate and propose that the superoxide responsible is likely to come from previously identified sites of intracellular NADPH oxidase activity. We speculate that ascorbate might protect the neutrophil against intracellular effects of superoxide generated at these sites.

  20. Rapid reaction of superoxide with insulin-tyrosyl radicals to generate a hydroperoxide with subsequent glutathione addition.

    PubMed

    Das, Andrew B; Nauser, Thomas; Koppenol, Willem H; Kettle, Anthony J; Winterbourn, Christine C; Nagy, Péter

    2014-05-01

    Tyrosine (Tyr) residues are major sites of radical generation during protein oxidation. We used insulin as a model to study the kinetics, mechanisms, and products of the reactions of radiation-induced or enzyme-generated protein-tyrosyl radicals with superoxide to demonstrate the feasibility of these reactions under oxidative stress conditions. We found that insulin-tyrosyl radicals combined to form dimers, mostly via the tyrosine at position 14 on the α chain (Tyr14). However, in the presence of superoxide, dimerization was largely outcompeted by the reaction of superoxide with insulin-tyrosyl radicals. Using pulse radiolysis, we measured a second-order rate constant for the latter reaction of (6±1) × 10(8) M(-1) s(-1) at pH 7.3, representing the first measured rate constant for a protein-tyrosyl radical with superoxide. Mass-spectrometry-based product analyses revealed the addition of superoxide to the insulin-Tyr14 radical to form the hydroperoxide. Glutathione efficiently reduced the hydroperoxide to the corresponding monoxide and also subsequently underwent Michael addition to the monoxide to give a diglutathionylated protein adduct. Although much slower, conjugation of the backbone amide group can form a bicyclic Tyr-monoxide derivative, allowing the addition of only one glutathione molecule. These findings suggest that Tyr-hydroperoxides should readily form on proteins under oxidative stress conditions where protein radicals and superoxide are both generated and that these should form addition products with thiol compounds such as glutathione.

  1. High-Content Imaging Assays for Identifying Compounds that Generate Superoxide and Impair Mitochondrial Membrane Potential in Adherent Eukaryotic Cells.

    PubMed

    Billis, Puja; Will, Yvonne; Nadanaciva, Sashi

    2014-02-19

    Reactive oxygen species (ROS) are constantly produced in cells as a result of aerobic metabolism. When there is an excessive production of ROS and the cell's antioxidant defenses are overwhelmed, oxidative stress occurs. The superoxide anion is a type of ROS that is produced primarily in mitochondria but is also generated in other regions of the cell including peroxisomes, endoplasmic reticulum, plasma membrane, and cytosol. Here, a high-content imaging assay using the dye dihydroethidium is described for identifying compounds that generate superoxide in eukaryotic cells. A high-content imaging assay using the fluorescent dye tetramethylrhodamine methyl ester is also described to identify compounds that impair mitochondrial membrane potential in eukaryotic cells. The purpose of performing both assays is to identify compounds that (1) generate superoxide at lower concentrations than they impair mitochondrial membrane potential, (2) impair mitochondrial membrane potential at lower concentrations than they generate superoxide, (3) generate superoxide and impair mitochondrial function at similar concentrations, and (4) do not generate superoxide or impair mitochondrial membrane potential during the duration of the assays.

  2. The Phylogeny and Active Site Design of Eukaryotic Copper-only Superoxide Dismutases

    SciTech Connect

    Peterson, Ryan L.; Galaleldeen, Ahmad; Villarreal, Johanna; Taylor, Alexander B.; Cabelli, Diane E.; Hart, P. John; Culotta, Valeria C.

    2016-08-17

    In eukaryotes the bimetallic Cu/Zn superoxide dismutase (SOD) enzymes play important roles in the biology of reactive oxygen species by disproportionating superoxide anion. We reported that the fungal pathogen Candida albicans expresses a novel copper-only SOD, known as SOD5, that lacks the zinc cofactor and electrostatic loop (ESL) domain of Cu/Zn-SODs for substrate guidance. In spite of these abnormalities, C. albicans SOD5 can disproportionate superoxide at rates limited only by diffusion. Here we demonstrate that this curious copper-only SOD occurs throughout the fungal kingdom as well as in phylogenetically distant oomycetes or “pseudofungi” species. It is the only form of extracellular SOD in fungi and oomycetes, in stark contrast to the extracellular Cu/Zn-SODs of plants and animals. Through structural biology and biochemical approaches we demonstrate that these copper-only SODs have evolved with a specialized active site consisting of two highly conserved residues equivalent to SOD5 Glu-110 and Asp-113. The equivalent positions are zinc binding ligands in Cu/Zn-SODs and have evolved in copper-only SODs to control catalysis and copper binding in lieu of zinc and the ESL. Similar to the zinc ion in Cu/Zn-SODs, SOD5 Glu-110 helps orient a key copper-coordinating histidine and extends the pH range of enzyme catalysis. Furthermore, SOD5 Asp-113 connects to the active site in a manner similar to that of the ESL in Cu/Zn-SODs and assists in copper cofactor binding. Copper-only SODs are virulence factors for certain fungal pathogens; thus this unique active site may be a target for future anti-fungal strategies.

  3. Predicted effects of nitric oxide and superoxide on the vasoactivity of the afferent arteriole.

    PubMed

    Layton, Anita T; Edwards, Aurélie

    2015-10-15

    We expanded a published mathematical model of an afferent arteriole smooth muscle cell in rat kidney (Edwards A, Layton, AT. Am J Physiol Renal Physiol 306: F34-F48, 2014) to understand how nitric oxide (NO) and superoxide (O(2)(-)) modulate the arteriolar diameter and its myogenic response. The present model includes the kinetics of NO and O(2)(-) formation, diffusion, and reaction. Also included are the effects of NO and its second messenger cGMP on cellular Ca²⁺ uptake and efflux, Ca²⁺-activated K⁺ currents, and myosin light chain phosphatase activity. The model considers as well pressure-induced increases in O(2)(-) production, O(2)(-)-mediated regulation of L-type Ca²⁺ channel conductance, and increased O(2)(-) production in spontaneous hypertensive rats (SHR). Our results indicate that elevated O(2)(-) production in SHR is sufficient to account for observed differences between normotensive and hypertensive rats in the response of the afferent arteriole to NO synthase inhibition, Tempol, and angiotensin II at baseline perfusion pressures. In vitro, whether the myogenic response is stronger in SHR remains uncertain. Our model predicts that if mechanosensitive cation channels are not modulated by O(2)(-), then fractional changes in diameter induced by pressure elevations should be smaller in SHR than in normotensive rats. Our results also suggest that most NO diffuses out of the smooth muscle cell without being consumed, whereas most O(2)(-) is scavenged, by NO and superoxide dismutase. Moreover, the predicted effects of superoxide on arteriolar constriction are not predominantly due to its scavenging of NO.

  4. The Phylogeny and Active Site Design of Eukaryotic Copper-only Superoxide Dismutases.

    PubMed

    Peterson, Ryan L; Galaleldeen, Ahmad; Villarreal, Johanna; Taylor, Alexander B; Cabelli, Diane E; Hart, P John; Culotta, Valeria C

    2016-09-30

    In eukaryotes the bimetallic Cu/Zn superoxide dismutase (SOD) enzymes play important roles in the biology of reactive oxygen species by disproportionating superoxide anion. Recently, we reported that the fungal pathogen Candida albicans expresses a novel copper-only SOD, known as SOD5, that lacks the zinc cofactor and electrostatic loop (ESL) domain of Cu/Zn-SODs for substrate guidance. Despite these abnormalities, C. albicans SOD5 can disproportionate superoxide at rates limited only by diffusion. Here we demonstrate that this curious copper-only SOD occurs throughout the fungal kingdom as well as in phylogenetically distant oomycetes or "pseudofungi" species. It is the only form of extracellular SOD in fungi and oomycetes, in stark contrast to the extracellular Cu/Zn-SODs of plants and animals. Through structural biology and biochemical approaches we demonstrate that these copper-only SODs have evolved with a specialized active site consisting of two highly conserved residues equivalent to SOD5 Glu-110 and Asp-113. The equivalent positions are zinc binding ligands in Cu/Zn-SODs and have evolved in copper-only SODs to control catalysis and copper binding in lieu of zinc and the ESL. Similar to the zinc ion in Cu/Zn-SODs, SOD5 Glu-110 helps orient a key copper-coordinating histidine and extends the pH range of enzyme catalysis. SOD5 Asp-113 connects to the active site in a manner similar to that of the ESL in Cu/Zn-SODs and assists in copper cofactor binding. Copper-only SODs are virulence factors for certain fungal pathogens; thus this unique active site may be a target for future anti-fungal strategies. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  5. Changes in superoxide dismutase mRNA expression by streptozotocin-induced diabetes.

    PubMed Central

    Kamata, K.; Kobayashi, T.

    1996-01-01

    1. Experiments were designed to investigate the involvement of superoxide anions in the attenuated endothelium-dependent relaxation of the rat aorta from streptozotocin (STZ)-induced diabetic rats. 2. The endothelium-dependent relaxation responses to acetylcholine (ACh, 10(-7) M) in helical strips of the aorta precontracted with noradrenaline (NA, 5 x 10(-3) approximately 3 x 10(-7) M) were significantly decreased in STZ-induced diabetic rats. The recovery phase of the relaxation after single administration of ACh in the STZ-induced diabetic rats was more rapid than those in control vessels. 3. Preincubation of aortic strips with superoxide dismutase (SOD, 60 u ml-1) normalized the recovery phase of the relaxation of diabetic aorta after single administration of ACh, whereas catalase (150 u ml-1) or indomethacin (10(-5) M) had no effects on the relaxation. 4. SOD (180 u ml-1) caused relaxation in NA precontracted aortic strips and the degree of the SOD-induced relaxation was significantly greater in diabetic aorta as compared with age-matched control vessels. 5. When the changes in mRNA expressions of Mn-SOD or Cu-Zn-SOD were observed, Mn-SOD mRNA expression was markedly decreased, and Cu-Zn-SOD was slightly decreased in diabetic aorta. 6. These results suggest that the rapid destruction of NO by superoxide anions may occur in the STZ-induced diabetic rats, and this may be due to a decrease in mRNA expression of Mn-SOD or Cu-Zn-SOD. Images Figure 4 PMID:8894182

  6. Cu,Zn superoxide dismutase from Photobacterium leiognathi is an hyperefficient enzyme.

    PubMed

    Stroppolo, M E; Sette, M; O'Neill, P; Polizio, F; Cambria, M T; Desideri, A

    1998-09-01

    The catalytic rate constant of recombinant Photobacterium leiognathi Cu,Zn superoxide dismutase has been determined as a function of pH by pulse radiolysis. At pH 7 and low ionic strength (I = 0.02 M) the catalytic rate constant is 8.5 x 10(9) M-1 s-1, more than two times the value found for all the native eukaryotic Cu,Zn superoxide dismutases investigated to date. Similarly, Brownian dynamics simulations indicate an enzyme-substrate association rate more than two times higher than that found for bovine Cu,Zn superoxide dismutase. Titration of the paramagnetic contribution to the water proton relaxation rate of the P. leiognathi with increasing concentration of halide ions with different radii indicates that the proteic channel delimiting the active site is wider than 4.4 A. This is at variance with that found on the eukariotic enzymes, and provides a rationale for the high catalytic rate of the bacterial enzyme. Evidence for solvent exposure of the active site different from that observed in the eukaryotic enzyme is suggested from the pH dependence of the water proton relaxation rate and of the EPR spectrum line shape, which indicate the occurrence of a prototropic equilibrium at pH 9.1 and 9.0, respectively. The pH dependence of the P. leiognathi catalytic rate has a trend different from that observed in the bovine enzyme, indicating that groups differently exposed to the solvent are involved in the modulation of the enzyme-substrate encounter.

  7. Mitochondrial superoxide flashes: metabolic biomarkers of skeletal muscle activity and disease.

    PubMed

    Wei, Lan; Salahura, Gheorghe; Boncompagni, Simona; Kasischke, Karl A; Protasi, Feliciano; Sheu, Shey-Shing; Dirksen, Robert T

    2011-09-01

    Mitochondrial superoxide flashes (mSOFs) are stochastic events of quantal mitochondrial superoxide generation. Here, we used flexor digitorum brevis muscle fibers from transgenic mice with muscle-specific expression of a novel mitochondrial-targeted superoxide biosensor (mt-cpYFP) to characterize mSOF activity in skeletal muscle at rest, following intense activity, and under pathological conditions. Results demonstrate that mSOF activity in muscle depended on electron transport chain and adenine nucleotide translocase functionality, but it was independent of cyclophilin-D-mediated mitochondrial permeability transition pore activity. The diverse spatial dimensions of individual mSOF events were found to reflect a complex underlying morphology of the mitochondrial network, as examined by electron microscopy. Muscle activity regulated mSOF activity in a biphasic manner. Specifically, mSOF frequency was significantly increased following brief tetanic stimulation (18.1 ± 1.6 to 22.3 ± 2.0 flashes/1000 μm²·100 s before and after 5 tetani) and markedly decreased (to 7.7 ± 1.6 flashes/1000 μm²·100 s) following prolonged tetanic stimulation (40 tetani). A significant temperature-dependent increase in mSOF frequency (11.9 ± 0.8 and 19.8 ± 2.6 flashes/1000 μm²·100 s at 23°C and 37°C) was observed in fibers from RYR1(Y522S/WT) mice, a mouse model of malignant hyperthermia and heat-induced hypermetabolism. Together, these results demonstrate that mSOF activity is a highly sensitive biomarker of mitochondrial respiration and the cellular metabolic state of muscle during physiological activity and pathological oxidative stress

  8. Macrophage Stimulating Protein (MSP) evokes superoxide anion production by human macrophages of different origin

    PubMed Central

    Brunelleschi, Sandra; Penengo, Lorenza; Lavagno, Luisa; Santoro, Claudio; Colangelo, Donato; Viano, Ilario; Gaudino, Giovanni

    2001-01-01

    Macrophage Stimulating Protein (MSP), a serum factor related to Hepatocyte Growth Factor, was originally discovered to stimulate chemotaxis of murine resident peritoneal macrophages. MSP is the ligand for Ron, a member of the Met subfamily of tyrosine kinase receptors. The effects of MSP on human macrophages and the role played in human pathophysiology have long been elusive.We show here that human recombinant MSP (hrMSP) evokes a dose-dependent superoxide anion production in human alveolar and peritoneal macrophages as well as in monocyte-derived macrophages, but not in circulating human monocytes. Consistently, the mature Ron protein is expressed by the MSP responsive cells but not by the unresponsive monocytes. The respiratory burst evoked by hrMSP is quantitatively higher than the one induced by N-formylmethionyl-leucyl-phenylalanine and similar to phorbol myristate acetate-evoked one.To investigate the mechanisms involved in NADPH oxidase activation, leading to superoxide anion production, different signal transduction inhibitors were used. By using the non selective tyrosine kinase inhibitor genistein, the selective c-Src inhibitor PP1, the tyrosine phosphatase inhibitor sodium orthovanadate, the phosphatidylinositol 3-kinase inhibitor wortmannin, the p38 inhibitor SB203580, the MEK inhibitor PD098059, we demonstrate that hrMSP-evoked superoxide production is mediated by tyrosine kinase activity, requires the activation of Src but not of PI 3-kinase. We also show that MAP kinase and p38 signalling pathways are involved.These results clearly indicate that hrMSP induces the respiratory burst in human macrophages but not in monocytes, suggesting for the MSP/Ron complex a role of activator as well as of possible marker for human mature macrophages. PMID:11704649

  9. Ergothioneine products derived by superoxide oxidation in endothelial cells exposed to high-glucose.

    PubMed

    Servillo, Luigi; D'Onofrio, Nunzia; Casale, Rosario; Cautela, Domenico; Giovane, Alfonso; Castaldo, Domenico; Balestrieri, Maria Luisa

    2017-03-12

    Ergothioneine (Egt), 2-mercapto-L-histidine betaine (ESH), is a dietary component acting as antioxidant and cytoprotectant. In vitro studies demonstrated that Egt, a powerful scavenger of hydroxyl radicals, superoxide anion, hypochlorous acid and peroxynitrite, protects vascular function against oxidative damages, thus preventing endothelial dysfunction. In order to delve the peculiar oxidative behavior of Egt, firstly identified in cell free-systems, experiments were designed to identify the Egt oxidation products when endothelial cells (EC) benefit of its protection against high-glucose (hGluc). HPLC-ESI-MS/MS analyses revealed a decrease in the intracellular GSH levels and an increase in the ophthalmic acid (OPH) levels during hGluc treatment. Interestingly, in the presence of Egt, the decrease of the GSH levels was lower than in cells treated with hGluc alone, and this effect was paralleled by lower OPH levels. Egt was also effective in reducing the cytotoxicity of H2O2 and paraquat (PQT), an inducer of superoxide anion production, showing a similar time-dependent pattern of GSH and OPH levels, although with peaks occurring at different times. Importantly, Egt oxidation generated not only hercynine (EH) but also the sulfonic acid derivative (ESO3H) whose amounts were dependent on the oxidative stress employed. Furthermore, cell-free experiments confirmed the formation of both EH and ESO3H when Egt was reacted with superoxide anion. In summary, these data, by identifying the EH and ESO3H formation in EC exposed to hGluc, highlight the cellular antioxidant properties of Egt, whose peculiar redox behavior makes it an attractive candidate for the prevention of oxidative stress-associated endothelial dysfunction during hyperglycemia.

  10. The effect of superoxide dismutase mimetic and catalase on the quality of postthawed goat semen.

    PubMed

    Shafiei, Mojtaba; Forouzanfar, Mohsen; Hosseini, Sayyed Morteza; Esfahani, Mohammad Hossein Nasr

    2015-05-01

    Manganese(III) meso-tetrakis(N-ethylpyridinium-2-yl)porphyrin chloride (MnTE) is a cell-permeable superoxide dismutase mimetic agent which can convert superoxide to hydrogen peroxide (H2O2). Supplementation of MnTE to a commercial semen extender can protect sperm from superoxide but not H2O2. Therefore, we proposed that addition of catalase (0.0, 200, or 400 IU/mL) in combination with MnTE (0.1 μM) may further improve the cryopreservation efficiency of goat semen in commercially optimized freezing media such as Andromed. Therefore, ejaculates were obtained from three adult bucks twice a week during the breeding season and diluted with Andromed supplemented with or without MnTE and catalase and were frozen in liquid nitrogen. Sperm parameters and reactive oxygen species contents were evaluated 2 hours after dilution (before freezing) and after freezing/thawing. The results revealed that all the treatments significantly (P ≤ 0.05) improved sperm motility, viability, and membrane integrity after freezing and reduced reactive oxygen species content compared with the control group, but maximum improvement was obtained in MnTE + 400 IU/mL catalase. In addition, supplementation with these antioxidants significantly (P ≤ 0.05) increases the cleavage rate after IVF. In conclusion, the results of present study suggest that addition of antioxidant MnTE or catalase to commercial optimized media, such as Andromed, improves total motility, membrane integrity, and viability of goat semen samples after thawing. But the degree of improvement for these parameters significantly (P ≤ 0.05) higher when MnTE and catalase were simultaneously added to the cryopreservation media.

  11. Superoxide (Electro)Chemistry on Well-Defined Surfaces in Organic Environments

    DOE PAGES

    Genorio, Bostjan; Staszak-Jirkovský, Jakub; Assary, Rajeev S.; ...

    2016-02-09

    Efficient chemical transformations in energy conversion and storage systems depend on understanding superoxide anion (O2–) electrochemistry at atomic and molecular levels. Here, in this work, a combination of experimental and theoretical techniques are used for rationalizing, and ultimately understanding, the complexity of superoxide anion (electro)chemistry in organic environments. By exploring the O2 + e– ↔ O2– reaction on well-characterized metal single crystals (Au, Pt, Ir), Pt single crystal modified with a single layer of graphene (Graphene@Pt(111)), and glassy carbon (GC) in 1,2 dimethoxyethane (DME) electrolytes, we demonstrate that (i) the reaction is an outer-sphere process; (ii) the reaction product O2–more » can “attack” any part of the DME molecule, i.e., the C–O bond via nucleophilic reaction and the C–H bond via radical hydrogen abstraction; (iii) the adsorption of carbon-based decomposition products and the extent of formation of a “solid electrolyte interface” (“SEI”) increases in the same order as the reactivity of the substrate, i.e., Pt(hkl)/Ir(hkl) » Au(hkl)/GC > Gaphene@Pt(111); and (iv) the formation of the “SEI” layer leads to irreversible superoxide electrochemistry on Pt(hkl) and Ir(hkl) surfaces. In conclusion, we believe this fundamental insight provides a pathway for the rational design of stable organic solvents that are urgently needed for the development of a new generation of reliable and affordable battery systems.« less

  12. The Phylogeny and Active Site Design of Eukaryotic Copper-only Superoxide Dismutases

    SciTech Connect

    Peterson, Ryan L.; Galaleldeen, Ahmad; Villarreal, Johanna; Taylor, Alexander B.; Cabelli, Diane E.; Hart, P. John; Culotta, Valeria C.

    2016-08-17

    In eukaryotes the bimetallic Cu/Zn superoxide dismutase (SOD) enzymes play important roles in the biology of reactive oxygen species by disproportionating superoxide anion. We reported that the fungal pathogen Candida albicans expresses a novel copper-only SOD, known as SOD5, that lacks the zinc cofactor and electrostatic loop (ESL) domain of Cu/Zn-SODs for substrate guidance. In spite of these abnormalities, C. albicans SOD5 can disproportionate superoxide at rates limited only by diffusion. Here we demonstrate that this curious copper-only SOD occurs throughout the fungal kingdom as well as in phylogenetically distant oomycetes or “pseudofungi” species. It is the only form of extracellular SOD in fungi and oomycetes, in stark contrast to the extracellular Cu/Zn-SODs of plants and animals. Through structural biology and biochemical approaches we demonstrate that these copper-only SODs have evolved with a specialized active site consisting of two highly conserved residues equivalent to SOD5 Glu-110 and Asp-113. The equivalent positions are zinc binding ligands in Cu/Zn-SODs and have evolved in copper-only SODs to control catalysis and copper binding in lieu of zinc and the ESL. Similar to the zinc ion in Cu/Zn-SODs, SOD5 Glu-110 helps orient a key copper-coordinating histidine and extends the pH range of enzyme catalysis. Furthermore, SOD5 Asp-113 connects to the active site in a manner similar to that of the ESL in Cu/Zn-SODs and assists in copper cofactor binding. Copper-only SODs are virulence factors for certain fungal pathogens; thus this unique active site may be a target for future anti-fungal strategies.

  13. The Phylogeny and Active Site Design of Eukaryotic Copper-only Superoxide Dismutases

    DOE PAGES

    Peterson, Ryan L.; Galaleldeen, Ahmad; Villarreal, Johanna; ...

    2016-08-17

    In eukaryotes the bimetallic Cu/Zn superoxide dismutase (SOD) enzymes play important roles in the biology of reactive oxygen species by disproportionating superoxide anion. We reported that the fungal pathogen Candida albicans expresses a novel copper-only SOD, known as SOD5, that lacks the zinc cofactor and electrostatic loop (ESL) domain of Cu/Zn-SODs for substrate guidance. In spite of these abnormalities, C. albicans SOD5 can disproportionate superoxide at rates limited only by diffusion. Here we demonstrate that this curious copper-only SOD occurs throughout the fungal kingdom as well as in phylogenetically distant oomycetes or “pseudofungi” species. It is the only form ofmore » extracellular SOD in fungi and oomycetes, in stark contrast to the extracellular Cu/Zn-SODs of plants and animals. Through structural biology and biochemical approaches we demonstrate that these copper-only SODs have evolved with a specialized active site consisting of two highly conserved residues equivalent to SOD5 Glu-110 and Asp-113. The equivalent positions are zinc binding ligands in Cu/Zn-SODs and have evolved in copper-only SODs to control catalysis and copper binding in lieu of zinc and the ESL. Similar to the zinc ion in Cu/Zn-SODs, SOD5 Glu-110 helps orient a key copper-coordinating histidine and extends the pH range of enzyme catalysis. Furthermore, SOD5 Asp-113 connects to the active site in a manner similar to that of the ESL in Cu/Zn-SODs and assists in copper cofactor binding. Copper-only SODs are virulence factors for certain fungal pathogens; thus this unique active site may be a target for future anti-fungal strategies.« less

  14. Expression of bovine superoxide dismutase in Drosophila melanogaster augments resistance of oxidative stress.

    PubMed Central

    Reveillaud, I; Niedzwiecki, A; Bensch, K G; Fleming, J E

    1991-01-01

    Superoxide dismutases (SOD) play a major role in the intracellular defense against oxygen radical damage to aerobic cells. In eucaryotes, the cytoplasmic form of the enzyme is a 32-kDa dimer containing two copper and two zinc atoms (CuZn SOD) that catalyzes the dismutation of the superoxide anion (O2-) to H2O2 and O2. Superoxide-mediated damage has been implicated in a number of biological processes, including aging and cancer; however, it is not certain whether endogenously elevated levels of SOD will reduce the pathological events resulting from such damage. To understand the in vivo relationship between an efficient dismutation of O2- and oxidative injury to biological structures, we generated transgenic strains of Drosophila melanogaster overproducing CuZn SOD. This was achieved by microinjecting Drosophila embryos with P-elements containing bovine CuZn SOD cDNA under the control of the Drosophila actin 5c gene promoter. Adult flies of the resulting transformed lines which expressed both mammalian and Drosophila CuZn SOD were then used as a novel model for evaluating the role of oxygen radicals in aging. Our data show that expression of enzymatically active bovine SOD in Drosophila flies confers resistance to paraquat, an O2(-)-generating compound. This is consistent with data on adult mortality, because there was a slight but significant increase in the mean lifespan of several of the transgenic lines. The highest level of expression of the active enzyme in adults was 1.60 times the normal value. Higher levels may have led to the formation of toxic levels of H2O2 during development, since flies that died during the process of eclosion showed an unusual accumulation of lipofuscin (age pigment) in some of their cells. In conclusion, our data show that free-radical detoxification has a minor by positive effect on mean longevity for several strains. Images PMID:1899285

  15. Superoxide (Electro)Chemistry on Well-Defined Surfaces in Organic Environments

    SciTech Connect

    Genorio, Bostjan; Staszak-Jirkovský, Jakub; Assary, Rajeev S.; Connell, Justin G.; Strmcnik, Dusan; Diesendruck, Charles E.; Lopes, Pietro P.; Stamenkovic, Vojislav R.; Moore, Jeffrey S.; Curtiss, Larry A.; Markovic, Nenad M.

    2016-02-09

    Efficient chemical transformations in energy conversion and storage systems depend on understanding superoxide anion (O2) electrochemistry at atomic and molecular levels. Here, in this work, a combination of experimental and theoretical techniques are used for rationalizing, and ultimately understanding, the complexity of superoxide anion (electro)chemistry in organic environments. By exploring the O2 + e ↔ O2 reaction on well-characterized metal single crystals (Au, Pt, Ir), Pt single crystal modified with a single layer of graphene (Graphene@Pt(111)), and glassy carbon (GC) in 1,2 dimethoxyethane (DME) electrolytes, we demonstrate that (i) the reaction is an outer-sph