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Sample records for accumulation myeloperoxidase activity

  1. Bioluminescence imaging of myeloperoxidase activity in vivo

    PubMed Central

    Gross, Shimon; Gammon, Seth T; Moss, Britney L; Rauch, Daniel; Harding, John; Heinecke, Jay W; Ratner, Lee; Piwnica-Worms, David

    2010-01-01

    The myeloperoxidase (MPO) system of activated phagocytes is central to normal host defense mechanisms, and dysregulated MPO contributes to the pathogenesis of inflammatory disease states ranging from atherosclerosis to cancer. Here we show that upon systemic administration, the small molecule luminol enables noninvasive bioluminescence imaging (BLI) of MPO activity in vivo. Luminol-BLI allowed quantitative longitudinal monitoring of MPO activity in animal models of acute dermatitis, mixed allergic contact hypersensitivity, focal arthritis and spontaneous large granular lymphocytic tumors. Bioluminescence colocalized with histological sites of inflammation and was totally abolished in gene-deleted Mpo−/− mice, despite massive tissue infiltration of neutrophils and activated eosinophils, indicating that eosinophil peroxidase did not contribute to luminol-BLI in vivo. Thus, luminol-BLI provides a noninvasive, specific and highly sensitive optical readout of phagocyte-mediated MPO activity in vivo and may enable new diagnostic applications in a wide range of acute and chronic inflammatory conditions. PMID:19305414

  2. Bioluminescence imaging of myeloperoxidase activity in vivo.

    PubMed

    Gross, Shimon; Gammon, Seth T; Moss, Britney L; Rauch, Daniel; Harding, John; Heinecke, Jay W; Ratner, Lee; Piwnica-Worms, David

    2009-04-01

    The myeloperoxidase (MPO) system of activated phagocytes is central to normal host defense mechanisms, and dysregulated MPO contributes to the pathogenesis of inflammatory disease states ranging from atherosclerosis to cancer. Here we show that upon systemic administration, the small molecule luminol enables noninvasive bioluminescence imaging (BLI) of MPO activity in vivo. Luminol-BLI allowed quantitative longitudinal monitoring of MPO activity in animal models of acute dermatitis, mixed allergic contact hypersensitivity, focal arthritis and spontaneous large granular lymphocytic tumors. Bioluminescence colocalized with histological sites of inflammation and was totally abolished in gene-deleted Mpo(-/-) mice, despite massive tissue infiltration of neutrophils and activated eosinophils, indicating that eosinophil peroxidase did not contribute to luminol-BLI in vivo. Thus, luminol-BLI provides a noninvasive, specific and highly sensitive optical readout of phagocyte-mediated MPO activity in vivo and may enable new diagnostic applications in a wide range of acute and chronic inflammatory conditions.

  3. Cyanate-mediated inhibition of neutrophil myeloperoxidase activity.

    PubMed Central

    Qian, M; Eaton, J W; Wolff, S P

    1997-01-01

    Cyanate (CNO-) forms spontaneously in solutions containing urea, and is present in urine and the body fluids of uraemic patients. We have explored the possibility that CNO- might be one of the unknown substances responsible for the reported impairment, by urine and uraemic plasma, of neutrophil oxidative metabolism (especially as measured by luminol-enhanced chemiluminescence). Luminol-enhanced chemiluminescence generated by human neutrophils derives predominantly from the activity of myeloperoxidase (MPO) which produces hypochlorous acid from H2O2 and Cl-. We hypothesized that CNO- (which resembles the 'pseudohalide' thiocyanate, an alternative substrate for MPO) might somehow interfere with the activity of MPO. In support of this, we find: (i) CNO- inhibits both peroxidative and halogenating activities of MPO and also inhibits the enzyme within intact human neutrophils; (ii) the inhibition is H2O2-dependent, irreversible, accompanied by covalent addition of [14C]CNO- (or a carbon-containing fragment thereof) to the enzyme; (iii) CNO- also inhibits Cl-/H2O2/MPO-mediated bacterial killing. Impairment of this arm of neutrophil bactericidal activity by CNO- formed from urea may be one factor in the risk of urinary-tract infection associated with urinary stasis and perhaps in the generalized increase in susceptibility to infection in uraemic patients. PMID:9337863

  4. Myeloperoxidase reduces the opsonizing activity of immunoglobulin G and complement component C3b.

    PubMed

    Coble, B I; Dahlgren, C; Hed, J; Stendahl, O

    1984-12-20

    The effect of myeloperoxidase, hydrogen peroxide (H2O2) and a halide (Cl) on the opsonizing molecules in immunoglobulin G (IgG) and complement factor C3b was assayed. At concentrations of the enzyme (1 microgram/ml) that can be found in the extracellular fluid during inflammation, the myeloperoxidase-H2O2-Cl system inhibited the opsonizing effect of IgG and C3b measured as phagocytic uptake and superoxide generation. The effect was related to the enzymatic peroxidative activity of the protein. The presence of albumin (10 mg/ml) reduced the effect of myeloperoxidase with 10-20%. Taurine, which in the presence of myeloperoxidase-H2O2-Cl forms hydrophilic chloramines, and D-penicillamine, which scavenges HOCl, neutralize the inhibitory effect of myeloperoxidase. This suggests that either hypochlorous acid or lipophilic chloramines may exert its effect by oxidizing free sulphydryl groups exposed on the opsonizing ligands. Since the myeloperoxidase-H2O2-halide system also affects chemotactic factors, leukotrienes, proteinases and membrane receptors, the system may in several ways affect the development of the inflammatory response.

  5. Inhibition of Myeloperoxidase Activity in Cystic Fibrosis Sputum by Peptide Inhibitor of Complement C1 (PIC1)

    PubMed Central

    Hair, Pamela S.; Sass, Laura A.; Krishna, Neel K.

    2017-01-01

    Myeloperoxidase is the major peroxidase enzyme in neutrophil granules and implicated in contributing to inflammatory lung damage in cystic fibrosis. Free myeloperoxidase is present in cystic fibrosis lung fluid and generates hypochlorous acid. Here we report a new inhibitor of myeloperoxidase activity, Peptide Inhibitor of Complement C1 (PIC1). Using TMB as the oxidizing substrate, PIC1 inhibited myeloperoxidase activity in cystic fibrosis sputum soluble fractions by an average of a 3.4-fold decrease (P = 0.02). PIC1 also dose-dependently inhibited myeloperoxidase activity in a neutrophil lysate or purified myeloperoxidase by up to 28-fold (P < 0.001). PIC1 inhibited myeloperoxidase activity similarly, on a molar basis, as the specific myeloperoxidase inhibitor 4-Aminobenzoic acid hydrazide (ABAH) for various oxidizing substrates. PIC1 was able to protect the heme ring of myeloperoxidase from destruction by NaOCl, assayed by spectral analysis. PIC1 incubated with oxidized TMB reversed the oxidation state of TMB, as measured by absorbance at 450 nm, with a 20-fold reduction in oxidized TMB (P = 0.02). This result was consistent with an antioxidant mechanism for PIC1. In summary, PIC1 inhibits the peroxidase activity of myeloperoxidase in CF sputum likely via an antioxidant mechanism. PMID:28135312

  6. Myeloperoxidase activity and the oxidized proteins in blood neutrophils of patients with pneumonia.

    PubMed

    Muravlyova, Larissa; Molotov-Luchanskiy, Vilen; Bakirova, Ryszhan; Klyuyev, Dmitriy; Demidchik, Ludmila; Kolesnikova, Yevgeniya

    2014-10-01

    The main purpose of our investigation was to study myeloperoxidase activity and concentration of oxidized proteins in blood neutrophils of patients with ambulant pneumonia and secondary pneumonia which has arisen on a background of chronic obstructive pulmonary disease (COPD). Patients were divided into 2 groups. 17 patients with ambulant pneumonia moderate severity and respiratory insufficiency of grade 2 were included in the 1-st group. 20 COPD patients with secondary pneumonia moderate severity and with respiratory insufficiency of grade 2 were included in the 2-nd group. The control group consisted of 15 healthy subjects. The reactive protein carbonyl derivates, advanced oxidation protein products (AOPP) and myeloperoxidase activity were detected in neutrophils. In neutrophils of 1-st group patients the augmentation of reactive protein carbonyl derivates was observed in comparison with healthy ones. In neutrophils of 2-nd group patients the slight decrease of reactive protein carbonyl derivates was observed in comparison with healthy ones (by 17%). In neutrophils of 2-nd group patients the significant increasing AOPP in comparison with healthy ones (p <0.01) and 1 group patients (p <0.05) was fixed. Myeloperoxidase activity was higher in neutrophils of 1-th group patients in comparison with healthy ones. In neutrophils of 2-nd group patients myeloperoxidase activity was higher in comparison with the same of 1 group patients (by 67%, p <0.05). Our results showed the different direction of oxidized proteins formation neutrophils of patients with primary and secondary pneumonia. Besides that the varied degree of myeloperoxidase activity was fixed. Our results require more detailed understanding because they can reflect peculiar mechanisms of pneumonia development and determine the characteristics of their progression.

  7. Novel bis-arylalkylamines as myeloperoxidase inhibitors: Design, synthesis, and structure-activity relationship study.

    PubMed

    Aldib, Iyas; Gelbcke, Michel; Soubhye, Jalal; Prévost, Martine; Furtmüller, Paul G; Obinger, Christian; Elfving, Betina; Alard, Ibaa Chikh; Roos, Goedele; Delporte, Cédric; Berger, Gilles; Dufour, Damien; Zouaoui Boudjeltia, Karim; Nève, Jean; Dufrasne, Francois; Van Antwerpen, Pierre

    2016-11-10

    Human myeloperoxidase (MPO) plays an important role in innate immunity but also aggravates tissue damage by oxidation of biomolecules at sites of inflammation. As a result from a recent high-throughput virtual screening approach for MPO inhibitors, bis-2,2'-[(dihydro-1,3(2H,4H) pyrimidinediyl)bis(methylene)]phenol was detected as a promising lead compound for inhibition of the MPO-typical two-electron oxidation of chloride to hypochlorous acid (IC50 = 0.5 μM). In the present pharmacomodulation study, 37 derivatives of this lead compound were designed and synthesized driven by comprehensive docking studies and the impact on the chlorination activity of MPO. We describe the structural requirements for optimum (i) binding to the heme periphery and (ii) inhibition capacity. Finally, the best three inhibitors (bis-arylalkylamine derivatives) were probed for interaction with the MPO redox intermediates Compound I and Compound II. Determined apparent bimolecular rate constants together with determination of reduction potential and nucleophilicity of the selected compounds allowed us to propose a mechanism of inhibition. The best inhibitor was found to promote the accumulation of inactive form of MPO-Compound II and has IC50 = 54 nM, demonstrating the successful approach of the drug design. Due to the similarity of ligand interactions between MPO and serotonine transporter, the selectivity of this inhibitor was also tested on the serotonin transporter providing a selectivity index of 14 (KiSERT/IC50MPO).

  8. Modulatory activities of Agelanthus dodoneifolius (Loranthaceae) extracts on stimulated equine neutrophils and myeloperoxidase activity.

    PubMed

    Boly, Raïnatou; Dessy, Stéphanie; Kohnen, Stephan; Kini, Félix; Lompo, Marius; Mouithys-Mickalad, Ange; Guissou, Innocent Pierre; Dubois, Jacques; Deby-Dupont, Ginette; Serteyn, Didier; Franck, Thierry

    2011-08-01

    Agelanthus dodoneifolius DC Danser (Loranthaceae) is used for the treatment of various diseases including asthma. The aqueous and hydroalcoholic extracts have been reported to have anti-inflammatory, spasmolytic and bronchorelaxant activities. The present study investigates the effects of the aqueous decoction and the diethyl ether, ethyl acetate and butanolic fractions of Agelanthus dodoneifolius DC Danser (Loranthaceae) on reactive oxygen species (ROS) production and myeloperoxidase (MPO) release by phorbol 12-myristate 13-acetate (PMA)-stimulated equine neutrophils and on purified equine MPO activity. ROS production and MPO release by the PMA-stimulated neutrophils were measured by the lucigenin-enhanced chemiluminescence and ELISA assays, respectively. Specific immunological extraction followed by enzymatic detection (SIEFED) was used to specifically measure the equine MPO activity. Identification and quantification of the individual and total phenolic and flavonoid compounds were performed using UPLC-MS/MS equipment and colorimetric methods involving Folin-Ciocalteu and AlCl₃, respectively. All the tested extracts displayed dose-dependent inhibitory effects on the oxidant activities of neutrophils; a stronger effect was observed with the organic fractions than the aqueous decoction. These findings could be correlated with a high content of phenolic and flavonoid compounds. The results confirm the previously shown anti-inflammatory effect of Agelanthus dodoneifolius and its potential use for the treatment of neutrophil-dependent inflammatory diseases.

  9. Characterization and Antioxidant Properties of Six Algerian Propolis Extracts: Ethyl Acetate Extracts Inhibit Myeloperoxidase Activity

    PubMed Central

    Boufadi, Yasmina Mokhtaria; Soubhye, Jalal; Riazi, Ali; Rousseau, Alexandre; Vanhaeverbeek, Michel; Nève, Jean; Boudjeltia, Karim Zouaoui; Van Antwerpen, Pierre

    2014-01-01

    Because propolis contains many types of antioxidant compounds such as polyphenols and flavonoids, it can be useful in preventing oxidative damages. Ethyl acetate extracts of propolis from several Algerian regions show high activity by scavenging free radicals, preventing lipid peroxidation and inhibiting myeloperoxidase (MPO). By fractioning and assaying ethyl acetate extracts, it was observed that both polyphenols and flavonoids contribute to these activities. A correlation was observed between the polyphenol content and the MPO inhibition. However, it seems that kaempferol, a flavonoid, contributes mainly to the MPO inhibition. This molecule is in a high amount in the ethyl acetate extract and demonstrates the best efficiency towards the enzyme with an inhibiting concentration at 50% of 4 ± 2 μM. PMID:24514562

  10. Nanocrystal quantum dot-conjugated anti-myeloperoxidase antibody as the detector of activated neutrophils.

    PubMed

    Hoshino, Akiyoshi; Nagao, Tomokazu; Nakasuga, Akira; Ishida-Okawara, Akiko; Suzuki, Kazuo; Yasuhara, Masato; Yamamoto, Kenji

    2007-12-01

    Fluorescent nanocrystal quantum dots (QDs) have been applied to a wide range of biological studies by taking advantage of their fluorescence properties. Here we show that QDs conjugated with antibody against neutrophil peroxidase, myeloperoxidase (MPO). We designed a novel method to conjugate QDs to antibody without losing any antibody function including their antigen recognizing and Fc-receptor binding activities. When we applied anti-MPO antibody (Ab) with conventional organic probes in the case of immunostaining of living cells, the antibodies lost their fluorescence because of MPO enzymic activity to produce reactive oxygen species. Our QD-conjugated anti-MPO (alpha-MPO-QDs) can detect MPO on the surface of activated neutrophils. In addition, anti-MPO-QDs did not react to the inactivated neutrophils. In conclusion, we demonstrated that antibody visualized the expression of MPO on the neutrophil surface after stimulation with proinflammatory cytokines. Taken together, these techniques have the possibility that QDs can reveal the activation of neutrophils by immunostaining and flow cytometric analysis as a powerful tool for diagnosis of the neutrophil activation in vitro.

  11. A rapid bioluminescence assay for measuring myeloperoxidase activity in human plasma

    PubMed Central

    Goiffon, Reece J.; Martinez, Sara C.; Piwnica-Worms, David

    2015-01-01

    Myeloperoxidase (MPO) is a circulating cardiovascular disease (CVD) biomarker used to estimate clinical risk and patient prognosis. Current enzyme-linked immunosorbent assays (ELISA) for MPO concentration are costly and time-intensive. Here we report a novel bioluminescence assay, designated MPO activity on a polymer surface (MAPS), for measuring MPO activity in human plasma samples using the bioluminescent substrate L-012. The method delivers a result in under an hour and is resistant to confounding effects from endogenous MPO inhibitors. In a pilot clinical study, we compared MAPS and two clinical ELISAs using 72 plasma samples from cardiac catheterization patients. Results from parallel MAPS and ELISAs were concordant within 2±11 μg l−1 MPO with similar uncertainty and reproducibility. Results between parallel MAPS and ELISA were in better agreement than those between independent ELISAs. MAPS may provide an inexpensive and rapid assay for determining MPO activity in plasma samples from patients with CVD or potentially other immune and inflammatory disorders. PMID:25666092

  12. Alkalinity of Neutrophil Phagocytic Vacuoles Is Modulated by HVCN1 and Has Consequences for Myeloperoxidase Activity

    PubMed Central

    Levine, Adam P.; Duchen, Michael R.; de Villiers, Simon; Rich, Peter R.; Segal, Anthony W.

    2015-01-01

    The NADPH oxidase of neutrophils, essential for innate immunity, passes electrons across the phagocytic membrane to form superoxide in the phagocytic vacuole. Activity of the oxidase requires that charge movements across the vacuolar membrane are balanced. Using the pH indicator SNARF, we measured changes in pH in the phagocytic vacuole and cytosol of neutrophils. In human cells, the vacuolar pH rose to ~9, and the cytosol acidified slightly. By contrast, in Hvcn1 knock out mouse neutrophils, the vacuolar pH rose above 11, vacuoles swelled, and the cytosol acidified excessively, demonstrating that ordinarily this channel plays an important role in charge compensation. Proton extrusion was not diminished in Hvcn1-/- mouse neutrophils arguing against its role in maintaining pH homeostasis across the plasma membrane. Conditions in the vacuole are optimal for bacterial killing by the neutral proteases, cathepsin G and elastase, and not by myeloperoxidase, activity of which was unphysiologically low at alkaline pH. PMID:25885273

  13. The importance of myeloperoxidase enzyme activity in the pathogenesis of Crimean-Congo haemorrhagic fever.

    PubMed

    Guven, F M K; Aydin, H; Yildiz, G; Engin, A; Celik, V K; Bakir, D; Deveci, K

    2013-03-01

    Crimean-Congo haemorrhagic fever (CCHF) is a disease with a severe course including acute viral haemorrhagic fever, ecchymosis, thrombocytopenia, hepatic function disorder and high mortality. Myeloperoxidase (MPO) is an enzyme located in neutrophil granulocytes and plays an important role in the destruction of phagocytosed micro-organisms. The aim of this study was to analyse MPO enzyme activity in CCHF cases compared with a control group. A total of 47 randomly selected CCHF patients admitted to the Department of Infectious Diseases of Cumhuriyet University Hospital in Sivas, Turkey, were studied, and as a control group, 41 age- and sex-matched individuals without any systemic disease were included in this study. MPO enzyme activity was measured in plasma and leukocytes for both groups by the ELISA method. MPO plasma and MPO leukocyte values were calculated as 57.62 ± 8.85 and 44.84 ± 9.71 in CCHF patients, and 0.79 ± 0.29 and 0.49 ± 0.11 in the controls, respectively. MPO enzyme activity was statistically significantly higher in patients with CCHF when compared to the control group. In conclusion, MPO enzyme activity is directly related to the activation of phagocytic leukocytes, and increases in both the plasma and leukocytes in CCHF patients. The increase of the MPO enzyme activity in leukocytes due to viral load leads to the destruction of the leukocyte. It is thought that MPO enzyme activity in plasma was higher in CCHF patients due to the destruction of leukocytes. MPO enzyme activity may be important in terms of the prognosis in patients with CCHF; however, more extensive studies are required on this subject.

  14. Increased Myeloperoxidase Activity and Protein Nitration Are Indicators of Inflammation in Patients with Chagas' Disease▿

    PubMed Central

    Dhiman, Monisha; Estrada-Franco, Jose Guillermo; Pando, Jasmine M.; Ramirez-Aguilar, Francisco J.; Spratt, Heidi; Vazquez-Corzo, Sara; Perez-Molina, Gladys; Gallegos-Sandoval, Rosa; Moreno, Roberto; Garg, Nisha Jain

    2009-01-01

    In this study, we investigated whether inflammatory responses contribute to oxidative/nitrosative stress in patients with Chagas' disease. We used three tests (enzyme-linked immunosorbent assay, immuno-flow cytometry, and STAT-PAK immunochromatography) to screen human serum samples (n = 1,481) originating from Chiapas, Mexico, for Trypanosoma cruzi-specific antibodies. We identified 121 subjects who were seropositive for T. cruzi-specific antibodies, a finding indicative of an 8.5% seroprevalence in the rural population from Chiapas. Seropositive and seronegative subjects were examined for plasma levels of biomarkers of inflammation, i.e., myeloperoxidase (MPO), inducible nitric oxide synthase (iNOS), and xanthine oxidase (XOD), as well as for oxidative (advanced oxidation protein products [AOPPs]) and nitrosative (3-nitrotyrosine [3NT]) biomarkers. The seropositive subjects exhibited a significant increase in MPO activity and protein level, the indicator of neutrophil activation. Subsequently, a corresponding increase in AOPP contents, formed by MPO-dependent hypochlorous acid and chloramine formation, was noted in seropositive subjects. The plasma level of 3NT was significantly increased in seropositive subjects, yet we observed no change in XOD activity (O2− source) and nitrate/nitrite contents (denotes iNOS activation and NO production), which implied that direct peroxynitrite formation does not contribute to increased nitrosative damage in chagasic subjects. Instead, a positive correlation between increased MPO activity and protein 3NT formation was observed, which suggested to us that MPO-dependent formation of nitrylchloride that occurs in the presence of physiological NO and O2− concentrations contributes to protein nitration. Overall, our data demonstrate that T. cruzi-induced neutrophil activation is pathological and contributes to MPO-mediated collateral protein oxidative and nitrosative damage in human patients with Chagas' disease. Therapies

  15. Diffused Intra-Oocyte Hydrogen Peroxide Activates Myeloperoxidase and Deteriorates Oocyte Quality.

    PubMed

    Khan, Sana N; Shaeib, Faten; Najafi, Tohid; Kavdia, Mahendra; Gonik, Bernard; Saed, Ghassan M; Goud, Pravin T; Abu-Soud, Husam M

    2015-01-01

    Hydrogen peroxide (H2O2) is a relatively long-lived signaling molecule that plays an essential role in oocyte maturation, implantation, as well as early embryonic development. Exposure to relatively high levels of H2O2 functions efficiently to accelerate oocyte aging and deteriorate oocyte quality. However, little precise information exists regarding intra-oocyte H2O2 concentrations, and its diffusion to the oocyte milieu. In this work, we utilized an L-shaped amperometric integrated H2O2-selective probe to directly and quantitatively measure the real-time intra-oocyte H2O2 concentration. This investigation provides an exact measurement of H2O2 in situ by reducing the possible loss of H2O2 caused by diffusion or reactivity with other biological systems. This experiment suggests that the intra-oocyte H2O2 levels of oocytes obtained from young animals are reasonably high and remained constant during the procedure measurements. However, the intra-oocyte H2O2 concentration dropped significantly (40-50% reduction) in response to catalase pre-incubation, suggesting that the measurements are truly H2O2 based. To further confirm the extracellular diffusion of H2O2, oocytes were incubated with myeloperoxidase (MPO), and the diffused H2O2 triggered MPO chlorinating activity. Our results show that the generated hypochlorous acid (HOCl) facilitated the deterioration in oocyte quality, a process that could be prevented by pre-incubating the oocytes with melatonin, which was experimentally proven to be oxidized utilizing HPLC methods. This study is the first to demonstrate direct quantitative measurement of intracellular H2O2, and its extracellular diffusion and activation of MPO as well as its impact on oocyte quality. These results may help in designing more accurate treatment plans in assisted reproduction under inflammatory conditions.

  16. The Impact of Myeloperoxidase and Activated Macrophages on Metaphase II Mouse Oocyte Quality

    PubMed Central

    Shaeib, Faten; Khan, Sana N.; Thakur, Mili; Kohan-Ghadr, Hamid-Reza; Drewlo, Sascha; Saed, Ghassan M.; Pennathur, Subramaniam; Abu-Soud, Husam M.

    2016-01-01

    Myeloperoxidase (MPO), an abundant heme-containing enzyme present in neutrophils, monocytes, and macrophages, is produced in high levels during inflammation, and associated with poor reproductive outcomes. MPO is known to generate hypochlorous acid (HOCl), a damaging reactive oxygen species (ROS) utilizing hydrogen peroxide (H2O2) and chloride (Cl-). Here we investigate the effect of activated immune cells and MPO on oocyte quality. Mouse metaphase II oocytes were divided into the following groups: 1) Incubation with a catalytic amount of MPO (40 nM) for different incubation periods in the presence of 100 mM Cl- with and without H2O2 and with and without melatonin (100 μM), at 37°C (n = 648/648 total number of oocytes in each group for oocytes with and without cumulus cells); 2) Co-cultured with activated mouse peritoneal macrophage and neutrophils cells (1.0 x 106 cells/ml) in the absence and presence of melatonin (200 μM), an MPO inhibitor/ROS scavenger, for different incubation periods in HTF media, at 37°C (n = 200/200); 3) Untreated oocytes incubated for 4 hrs as controls (n = 73/64). Oocytes were then fixed, stained and scored based on the microtubule morphology and chromosomal alignment. All treatments were found to negatively affect oocyte quality in a time dependent fashion as compared to controls. In all cases the presence of cumulus cells offered no protection; however significant protection was offered by melatonin. Similar results were obtained with oocytes treated with neutrophils. This work provides a direct link between MPO and decreased oocyte quality. Therefore, strategies to decrease MPO mediated inflammation may influence reproductive outcomes. PMID:26982351

  17. Diffused Intra-Oocyte Hydrogen Peroxide Activates Myeloperoxidase and Deteriorates Oocyte Quality

    PubMed Central

    Khan, Sana N.; Shaeib, Faten; Najafi, Tohid; Kavdia, Mahendra; Gonik, Bernard; Saed, Ghassan M.; Goud, Pravin T.; Abu-Soud, Husam M.

    2015-01-01

    Hydrogen peroxide (H2O2) is a relatively long-lived signaling molecule that plays an essential role in oocyte maturation, implantation, as well as early embryonic development. Exposure to relatively high levels of H2O2 functions efficiently to accelerate oocyte aging and deteriorate oocyte quality. However, little precise information exists regarding intra-oocyte H2O2 concentrations, and its diffusion to the oocyte milieu. In this work, we utilized an L-shaped amperometric integrated H2O2-selective probe to directly and quantitatively measure the real-time intra-oocyte H2O2 concentration. This investigation provides an exact measurement of H2O2 in situ by reducing the possible loss of H2O2 caused by diffusion or reactivity with other biological systems. This experiment suggests that the intra-oocyte H2O2 levels of oocytes obtained from young animals are reasonably high and remained constant during the procedure measurements. However, the intra-oocyte H2O2 concentration dropped significantly (40-50% reduction) in response to catalase pre-incubation, suggesting that the measurements are truly H2O2 based. To further confirm the extracellular diffusion of H2O2, oocytes were incubated with myeloperoxidase (MPO), and the diffused H2O2 triggered MPO chlorinating activity. Our results show that the generated hypochlorous acid (HOCl) facilitated the deterioration in oocyte quality, a process that could be prevented by pre-incubating the oocytes with melatonin, which was experimentally proven to be oxidized utilizing HPLC methods. This study is the first to demonstrate direct quantitative measurement of intracellular H2O2, and its extracellular diffusion and activation of MPO as well as its impact on oocyte quality. These results may help in designing more accurate treatment plans in assisted reproduction under inflammatory conditions. PMID:26197395

  18. A sensitive and selective assay for chloramine production by myeloperoxidase.

    PubMed

    Dypbukt, Jeannette M; Bishop, Cynthia; Brooks, Wendy M; Thong, Bob; Eriksson, Håkan; Kettle, Anthony J

    2005-12-01

    We describe a new assay for the chlorination activity of myeloperoxidase and detection of chloramines. Chloramines were detected by using iodide to catalyze the oxidation of either 3,3',5,5'-tetramethylbenzidine (TMB) or dihydrorhodamine to form strongly absorbing or fluorescent products, respectively. With TMB as little as 1 muM taurine chloramine could be detected. The sensitivity of the dihydrorhodamine assay was about 10-fold greater. The chlorination activity of myeloperoxidase was measured by trapping hypochlorous acid with taurine and subsequently using iodide to promote the oxidation reactions of the accumulated taurine chloramine. A similar approach was used to detect hypochlorous acid production by stimulated human neutrophils. Iodide-dependent catalysis distinguished N-chloramines from N-bromamines. This allows for discrimination between heme peroxidases that generate either hypochlorous acid or hypobromous acid. The assay has distinct advantages over existing assays for myeloperoxidase with regard to sensitivity, specificity, and its ease and versatility of use.

  19. δ-Aminolevulinate Dehydratase Activity is Stimulated in a MPTP Mouse Model of Parkinson's Disease: Correlation with Myeloperoxidase Activity.

    PubMed

    Sampaio, Tuane Bazanella; Marcondes Sari, Marcel Henrique; Pesarico, Ana Paula; Nogueira, Cristina Wayne

    2016-09-21

    Myeloperoxidase (MPO) is an inducible heme peroxidase responsive to some stress situations. It is already known that its activity is stimulated in neurodegenerative disorders and in the animal model of parkinson's disease (PD) induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). By contrast, the role of δ-aminolevulinate dehydratase (δ-ALA-D), an essential enzyme for heme synthesis, has not been investigated in the MPTP model. The aim of this study was to investigate the involvement of striatal δ-ALA-D activity in an acute model of PD, induced by MPTP, in C57Bl/6 mice and its correlation with MPO activity. Animals received four MPTP injections (20 mg/kg, i.p.) or saline (vehicle) to induce a PD model. 7 days after MPTP administration, the motor function was evaluated through rotarod and challenging beam tests in mice. Afterward, mice were killed, and the striata were removed for biochemical analyses. MPTP-treated mice showed impairment in motor skills, such as balance and motor coordination. Furthermore, there was a reduction of tyrosine hydroxylase levels in these animals, which characterizes the dopaminergic lesion. Striatal δ-ALA-D activity was stimulated by MPTP, as well as the MPO activity, and a significant positive correlation between δ-ALA-D and MPO activities was also demonstrated. These data suggest that δ-ALA-D activity could be stimulated due to the requirement of heme groups by peroxidases. Therefore, this study demonstrated for the first time the involvement of striatal δ-ALA-D activity in the MPTP model and its correlation with the MPO activity.

  20. A novel fluorescent probe for the detection of myeloperoxidase activity in atherosclerosis-associated macrophages

    PubMed Central

    Shepherd, Joanna; Hilderbrand, Scott A.; Waterman, Peter; Heinecke, Jay W.; Weissleder, Ralph; Libby, Peter

    2009-01-01

    SUMMARY The myeloperoxidase (MPO)-derived oxidant hypochlorous acid (HOCl/OCl−) is implicated in the pathogenesis of atherosclerosis and other inflammatory states. We have synthesized a novel imaging probe, sulfonaphthoaminophenyl fluorescein (SNAPF), that selectively reacts to HOCl. SNAPF detects HOCl produced by stimulated MPO-expressing cells cultured from human whole blood, as well as HOCl from bone marrow (BM)-derived macrophages isolated from transgenic mice that express human MPO. Two lines of evidence indicate that SNAPF permits the in vivo imaging of HOCl production. First, we used this approach to demonstrate HOCl production by neutrophils in experimental murine peritonitis. Second, we detected HOCl production by MPO expressing cells in human atherosclerotic arteries. Thus, fluorescence reflectance imaging by SNAPF may provide a valuable non-invasive molecular imaging tool for implicating HOCl and MPO in the damage of inflamed tissues. PMID:18022561

  1. Perpetuation of inflammation associated with experimental arthritis: the role of macrophage activation by neutrophilic myeloperoxidase.

    PubMed Central

    Gelderman, M P; Stuart, R; Vigerust, D; Fuhrmann, S; Lefkowitz, D L; Allen, R C; Lefkowitz, S S; Graham, S

    1998-01-01

    Rheumatoid arthritis (RA) is characterized by an abnormal cellular and cytokine infiltration of inflamed joints. This study addresses a previously unrecognized interaction between neutrophilic-myeloperoxidase (MPO) and macrophages (Mphi) which could explain the perpetuation of inflammation associated with RA. A monoarticular arthritis was induced in female Lewis rats by injection of streptococcal cell wall extracts (PG-APS). After swelling and erythema subsided, joints were re-injected with one of the following: porcine MPO or partially inactivated MPO (iMPO). Injection with either MPO or iMPO induced a 'flare' of experimental RA. Blocking the Mphi-mannose receptor by mannans, ablated exacerbation of disease. These results indicate that MPO or iMPO can play a pivotal role in the perpetuation but not initiation of this RA model. PMID:9927230

  2. Polyphenol Content and Modulatory Activities of Some Tropical Dietary Plant Extracts on the Oxidant Activities of Neutrophils and Myeloperoxidase

    PubMed Central

    Tsumbu, Cesar N.; Deby-Dupont, Ginette; Tits, Monique; Angenot, Luc; Frederich, Michel; Kohnen, Stephane; Mouithys-Mickalad, Ange; Serteyn, Didier; Franck, Thierry

    2012-01-01

    Young leaves of Manihot esculenta Crantz (Euphorbiaceae), Abelmoschus esculentus (Malvaceae), Hibiscus acetosella (Malvaceae) and Pteridium aquilinum (Dennstaedtiaceae) are currently consumed as green vegetables by peoples in sub-Saharan Africa, Latin America, Asia and their migrants living in Western Europe. Sub-Saharan peoples use Manihot, Abelmoschus and Hibiscus also in the folk medicine to alleviate fever and pain, in the treatment of conjunctivitis, rheumatism, hemorrhoid, abscesses, ... The present study investigates the effects of aqueous extracts of those plants on the production of reactive oxygen species (ROS) and the release of myeloperoxidase (MPO) by equine neutrophils activated with phorbol 12-myristate 13-acetate (PMA). The ROS production was measured by lucigenin-enhanced chemiluminescence (CL), and the release of total MPO by an ELISA method. The study also investigates the effect of the extracts on the activity of MPO by studying its nitration activity on tyrosine and by using a new technique called SIEFED (Specific Immunological Extraction Followed by Enzymatic Detection) that allows studying the direct interaction of compounds with the enzyme. In all experiments, the aqueous extracts of the plants developed concentration-dependent inhibitory effects. A moderate heat treatment did not significantly modify the inhibitory capacity of the extracts in comparison to not heated ones. Total polyphenol and flavonoid contents were determined with an HPLC-UV/DAD analysis and a spectroscopic method using Folin-Ciocalteu reagent. Some polyphenols with well-known antioxidant activities (caffeic acid, chlorogenic acid, hyperoside, rosmarinic acid and rutin) were found in the extracts and may partly explain the inhibitory activities observed. The role of those dietary and medicinal plants in the treatment of ROS-dependent inflammatory diseases could have new considerations for health. PMID:22312276

  3. Myeloperoxidase and cardiovascular disease.

    PubMed

    Nicholls, Stephen J; Hazen, Stanley L

    2005-06-01

    Myeloperoxidase (MPO) is a leukocyte-derived enzyme that catalyzes the formation of a number of reactive oxidant species. In addition to being an integral component of the innate immune response, evidence has emerged that MPO-derived oxidants contribute to tissue damage during inflammation. MPO-catalyzed reactions have been attributed to potentially proatherogenic biological activities throughout the evolution of cardiovascular disease, including during initiation, propagation, and acute complication phases of the atherosclerotic process. As a result, MPO and its downstream inflammatory pathways represent attractive targets for both prognostication and therapeutic intervention in the prophylaxis of atherosclerotic cardiovascular disease.

  4. Myeloperoxidase targets oxidative host attacks to Salmonella and prevents collateral tissue damage.

    PubMed

    Schürmann, Nura; Forrer, Pascal; Casse, Olivier; Li, Jiagui; Felmy, Boas; Burgener, Anne-Valérie; Ehrenfeuchter, Nikolaus; Hardt, Wolf-Dietrich; Recher, Mike; Hess, Christoph; Tschan-Plessl, Astrid; Khanna, Nina; Bumann, Dirk

    2017-01-23

    Host control of infections crucially depends on the capability to kill pathogens with reactive oxygen species (ROS). However, these toxic molecules can also readily damage host components and cause severe immunopathology. Here, we show that neutrophils use their most abundant granule protein, myeloperoxidase, to target ROS specifically to pathogens while minimizing collateral tissue damage. A computational model predicted that myeloperoxidase efficiently scavenges diffusible H2O2 at the surface of phagosomal Salmonella and converts it into highly reactive HOCl (bleach), which rapidly damages biomolecules within a radius of less than 0.1 μm. Myeloperoxidase-deficient neutrophils were predicted to accumulate large quantities of H2O2 that still effectively kill Salmonella, but most H2O2 would leak from the phagosome. Salmonella stimulation of neutrophils from normal and myeloperoxidase-deficient human donors experimentally confirmed an inverse relationship between myeloperoxidase activity and extracellular H2O2 release. Myeloperoxidase-deficient mice infected with Salmonella had elevated hydrogen peroxide tissue levels and exacerbated oxidative damage of host lipids and DNA, despite almost normal Salmonella control. These data show that myeloperoxidase has a major function in mitigating collateral tissue damage during antimicrobial oxidative bursts, by converting diffusible long-lived H2O2 into highly reactive, microbicidal and locally confined HOCl at pathogen surfaces.

  5. Circulating DNA and myeloperoxidase indicate disease activity in patients with thrombotic microangiopathies.

    PubMed

    Fuchs, Tobias A; Kremer Hovinga, Johanna A; Schatzberg, Daphne; Wagner, Denisa D; Lämmle, Bernhard

    2012-08-09

    Thrombotic microangiopathies (TMAs) are a group of life-threatening disorders characterized by thrombocytopenia, fragmentation of erythrocytes, and ischemic organ damage. Genetic disorders, autoimmune disease, and cancer are risk factors for TMAs, but an additional, unknown trigger is needed to bring about acute disease. Recent studies suggest that DNA and histones are released during inflammation or infection and stimulate coagulation, thrombosis, thrombocytopenia, and organ damage in mice. We show that extracellular DNA and histones as well as markers of neutrophils are present in acute TMAs. Analysis of plasma from TMA patients of different clinical categories revealed elevated levels of DNA-histone complexes and myeloperoxidase (MPO) from neutrophil granules as well as S100A8/A9, a heterocomplex abundant in neutrophil cytosol. During therapy of thrombotic thrombocytopenic purpura, a subtype of TMAs often associated with severe ADAMTS13 (a disintegrin and metalloproteinase with thrombospondin type 1 motifs, member 13) deficiency, plasma DNA and MPO were inversely correlated with platelet counts, and their levels indicated amelioration or exacerbation of the disease. ADAMTS13 deficiency together with increased levels of plasma DNA and MPO were characteristic for acute thrombotic thrombocytopenic purpura. A minor infection often precedes acute TMA and extracellular DNA and histones released during the inflammatory response could provide the second hit, which precipitates acute TMA in patients with pre-existing risk factors.

  6. Myeloperoxidase Enhances Etoposide and Mitoxantrone-Mediated DNA Damage: A Target for Myeloprotection in Cancer Chemotherapy

    PubMed Central

    Atwal, Mandeep; Lishman, Emma L.; Austin, Caroline A.

    2017-01-01

    Myeloperoxidase is expressed exclusively in granulocytes and immature myeloid cells and transforms the topoisomerase II (TOP2) poisons etoposide and mitoxantrone to chemical forms that have altered DNA damaging properties. TOP2 poisons are valuable and widely used anticancer drugs, but they are associated with the occurrence of secondary acute myeloid leukemias. These factors have led to the hypothesis that myeloperoxidase inhibition could protect hematopoietic cells from TOP2 poison-mediated genotoxic damage and, therefore, reduce the rate of therapy-related leukemia. We show here that myeloperoxidase activity leads to elevated accumulation of etoposide- and mitoxantrone-induced TOP2A and TOP2B-DNA covalent complexes in cells, which are converted to DNA double-strand breaks. For both drugs, the effect of myeloperoxidase activity was greater for TOP2B than for TOP2A. This is a significant finding because TOP2B has been linked to genetic damage associated with leukemic transformation, including etoposide-induced chromosomal breaks at the MLL and RUNX1 loci. Glutathione depletion, mimicking in vivo conditions experienced during chemotherapy treatment, elicited further MPO-dependent increase in TOP2A and especially TOP2B-DNA complexes and DNA double-strand break formation. Together these results support targeting myeloperoxidase activity to reduce genetic damage leading to therapy-related leukemia, a possibility that is enhanced by the recent development of novel specific myeloperoxidase inhibitors for use in inflammatory diseases involving neutrophil infiltration. PMID:27974636

  7. Evaluation of the serum catalase and myeloperoxidase activities in chronic arsenic-exposed individuals and concomitant cytogenetic damage

    SciTech Connect

    Banerjee, Mayukh; Banerjee, Nilanjana; Ghosh, Pritha; Das, Jayanta K.; Basu, Santanu; Sarkar, Ajoy K.; States, J. Christopher; Giri, Ashok K.

    2010-11-15

    Chronic arsenic exposure through contaminated drinking water is a major environmental health issue. Chronic arsenic exposure is known to exert its toxic effects by a variety of mechanisms, of which generation of reactive oxygen species (ROS) is one of the most important. A high level of ROS, in turn, leads to DNA damage that might ultimately culminate in cancer. In order to keep the level of ROS in balance, an array of enzymes is present, of which catalase (CAT) and myeloperoxidase (MPO) are important members. Hence, in this study, we determined the activities of these two enzymes in the sera and chromosomal aberrations (CA) in peripheral blood lymphocytes in individuals exposed and unexposed to arsenic in drinking water. Arsenic in drinking water and in urine was used as a measure of exposure. Our results show that individuals chronically exposed to arsenic have significantly higher CAT and MPO activities and higher incidence of CA. We found moderate positive correlations between CAT and MPO activities, induction of CA and arsenic in urine and water. These results indicate that chronic arsenic exposure causes higher CAT and MPO activities in serum that correlates with induction of genetic damage. We conclude that the serum levels of these enzymes might be used as biomarkers of early arsenic exposure induced disease much before the classical dermatological symptoms of arsenicosis begin to appear.

  8. Antioxidant activity of minimally processed red chicory (Cichorium intybus L.) evaluated in xanthine oxidase-, myeloperoxidase-, and diaphorase-catalyzed reactions.

    PubMed

    Lavelli, Vera

    2008-08-27

    Minimally processed red chicory products (Cichorium intybus L. var. silvestre) were studied for their polyphenol content and antioxidant activity evaluated by using the synthetic 2,2-diphenyl-1-(2,4,6-trinitrophenyl)hydrazyl radical and three model reactions catalyzed by relevant enzymatic sources of reactive oxygen species, namely, xanthine oxidase, myeloperoxidase, and diaphorase. Products were analyzed at the time of production and after storage at 4 degrees C within either a gas permeable film or a gas barrier film. The antioxidant activity and contents of hydroxycinnamic acids and flavonoids decreased by less than 20% during storage of the minimally processed red chicory products. Total phenolics were significantly correlated with the antioxidant activity evaluated with both the synthetic radical and the enzyme-catalyzed reactions. On a molar basis, red chicory phenolics were as efficient as the reference compound Trolox in scavenging the synthetic radical. However, red chicory phenolics had a much higher inhibitory activity than Trolox in the model enzymatic systems.

  9. Collagen catabolism through Coll2-1 and Coll2-1NO2 and myeloperoxidase activity in marathon runners.

    PubMed

    Henrotin, Yves; Labasse, Alain; Franck, Thierry; Bosseloir, Alain; Bury, Thierry; Deberg, Michelle

    2013-12-01

    To determine the influence of marathon on the serum levels of two markers of cartilage degradation, Coll2-1 and its nitrated form, Coll2-1NO2, and of a marker of neutrophils activation, the myeloperoxidase (MPO). Coll2-1, Coll2-1NO2, total and active MPO were measured in 98 marathon runners without joint pain and with an average age of 47 years. Sera were taken at rest right before the departure and within 30 min after the marathon. The subjects were submitted to a questionnaire concerning their physical activity and their life style. The levels of Coll2-1, Coll2-1NO2 and active MPO were not affected by age, body mass index, sex or performance. The levels of total MPO were higher in female than in male (p < 0.05), but were not affected by the other parameters. After the marathon, Coll2-1 and Coll2-1NO2 concentrations were slightly but systematically decreased. The total and active MPO concentrations were increased by 2 to 3-fold in comparison to the pre-marathon values (p < 0.001 for total and active MPO). The active MPO/total MPO ratio was significantly enhanced after the marathon (p < 0.001). The variation of total MPO during the marathon was negatively correlated with the training time per week (r = -0.34; p = 0.009). The serum levels of Coll2-1 and Coll2-1NO2 were slightly decreased by marathon, indicating that intensive running could reduce cartilage catabolism. Furthermore, Coll2-1NO2 was not correlated with the total and active MPO indicating that Coll2-1 nitration did not result of a systemic oxidative phenomenon but reflects local changes.

  10. Impact of myeloperoxidase-LDL interactions on enzyme activity and subsequent posttranslational oxidative modifications of apoB-100

    PubMed Central

    Delporte, Cédric; Boudjeltia, Karim Zouaoui; Noyon, Caroline; Furtmüller, Paul G.; Nuyens, Vincent; Slomianny, Marie-Christine; Madhoun, Philippe; Desmet, Jean-Marc; Raynal, Pierre; Dufour, Damien; Koyani, Chintan N.; Reyé, Florence; Rousseau, Alexandre; Vanhaeverbeek, Michel; Ducobu, Jean; Michalski, Jean-Claude; Nève, Jean; Vanhamme, Luc; Obinger, Christian; Malle, Ernst; Van Antwerpen, Pierre

    2014-01-01

    Oxidation of LDL by the myeloperoxidase (MPO)-H2O2-chloride system is a key event in the development of atherosclerosis. The present study aimed at investigating the interaction of MPO with native and modified LDL and at revealing posttranslational modifications on apoB-100 (the unique apolipoprotein of LDL) in vitro and in vivo. Using amperometry, we demonstrate that MPO activity increases up to 90% when it is adsorbed at the surface of LDL. This phenomenon is apparently reflected by local structural changes in MPO observed by circular dichroism. Using MS, we further analyzed in vitro modifications of apoB-100 by hypochlorous acid (HOCl) generated by the MPO-H2O2-chloride system or added as a reagent. A total of 97 peptides containing modified residues could be identified. Furthermore, differences were observed between LDL oxidized by reagent HOCl or HOCl generated by the MPO-H2O2-chloride system. Finally, LDL was isolated from patients with high cardiovascular risk to confirm that our in vitro findings are also relevant in vivo. We show that several HOCl-mediated modifications of apoB-100 identified in vitro were also present on LDL isolated from patients who have increased levels of plasma MPO and MPO-modified LDL. In conclusion, these data emphasize the specificity of MPO to oxidize LDL. PMID:24534704

  11. Sunscreen protection against ultraviolet-induced oxidative stress: evaluation of reduced glutathione levels, metalloproteinase secretion, and myeloperoxidase activity.

    PubMed

    Vilela, F M P; Fonseca, Y M; Vicentini, F T M C; Fonseca, M J V

    2013-11-01

    Several studies have demonstrated the skin protection by sunscreens considering the aspects skin penetration, photostability, and protection against erythema and sunburn. However, little is known about the effect of topically applied sunscreen formulations on the antioxidant defense, metalloproteinases, and inflammatory processes of skin in response to UVR exposure. Therefore, this study aimed to investigate the use of a cream gel formulation containing the UV filters benzophenone-3, octyl methoxycinnamate, and octyl salicylate to prevent skin damage from a single dose of UVR (2.87 J/cm2). This protective effect was evaluated in vivo by measuring the following biochemical parameters: reduced glutathione levels, secretion of matrix metalloproteinases, and myeloperoxidase activity. The results showed that the sunscreen formulation, despite having sun protection factor (SPF) 15, was not completely effective to protect the skin against GSH depletion, MMP-9 secretion and the inflammatory process induced by UVR. These results demonstrate the importance of analyzing UV-altered biochemical parameters of skin in order to propose new sunscreen formulations that can completely protect skin against UVR-induced damage.

  12. Mechanistic characterization of a 2-thioxanthine myeloperoxidase inhibitor and selectivity assessment utilizing click chemistry--activity-based protein profiling.

    PubMed

    Ward, Jessica; Spath, Samantha N; Pabst, Brandon; Carpino, Philip A; Ruggeri, Roger B; Xing, Gang; Speers, Anna E; Cravatt, Benjamin F; Ahn, Kay

    2013-12-23

    Myeloperoxidase (MPO) is a heme peroxidase that catalyzes the production of hypochlorous acid. Despite a high level of interest in MPO as a therapeutic target, there have been limited reports about MPO inhibitors that are suitable for evaluating MPO in pharmacological studies. 2-Thioxanthine, 3-(2-ethoxypropyl)-2-thioxo-2,3-dihydro-1H-purin-6(9H)-one (A), has recently been reported to inhibit MPO by covalently modifying the heme prosthetic group. Here we report a detailed mechanistic characterization demonstrating that A possesses all the distinguishing features of a mechanism-based inactivator. A is a time-dependent MPO inhibitor and displays saturable inactivation kinetics consistent with a two-step mechanism of inactivation and a potency (k(inact)/K(I) ratio) of 8450 ± 780 M⁻¹ s⁻¹. MPO inactivation by A is dependent on MPO catalysis and is protected by substrate. A reduces MPO compound I to compound II with a second-order rate constant of (0.801 ± 0.056) × 10⁶ M⁻¹ s⁻¹, and its irreversible inactivation of MPO occurs prior to release of the activated inhibitory species. Despite its relatively high selectivity against a broad panel of more than 100 individual targets, including enzymes, receptors, transporters, and ion channels, we demonstrate that A labels multiple other protein targets in the presence of MPO. By synthesizing an alkyne analogue of A and utilizing click chemistry-activity-based protein profiling, we present that the MPO-activated inhibitory species can diffuse away to covalently modify other proteins, as reflected by the relatively high partition ratio of A, which we determined to be 15.6. This study highlights critical methods that can guide the discovery and development of next-generation MPO inhibitors.

  13. Myeloperoxidase: Bridging the gap in neurodegeneration.

    PubMed

    Ray, R S; Katyal, Anju

    2016-09-01

    Neurodegenerative conditions present a group of complex disease pathologies mostly due to unknown aetiology resulting in neuronal death and permanent neurological disability. Any undesirable stress to the brain, disrupts homeostatic balance, through a remarkable convergence of pathophysiological changes and immune dysregulation. The crosstalk between inflammatory and oxidative mechanisms results in the release of neurotoxic mediators apparently spearheaded by myeloperoxidase derived from activated microglia, astrocytes, neurons as well as peripheral inflammatory cells. These isolated entities combinedly have the potential to flare up and contribute significantly to neuropathology and disease progression. Recent, clinicopathological evidence support the association of myeloperoxidase and its cytotoxic product, hypochlorous acid in a plethora of neurodegenerative diseases including Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, Multiple sclerosis, Stroke, Epilepsy etc. But the biochemical and mechanistic insights into myeloperoxidase mediated neuroinflammation and neuronal death is still an uncharted territory. The current review outlines the emerging recognition of myeloperoxidase in neurodegeneration, which may offer novel therapeutic and diagnostic targets for neurodegenerative disorders.

  14. Measurement of equine myeloperoxidase (MPO) activity in synovial fluid by a modified MPO assay and evaluation of joint diseases - an initial case study.

    PubMed

    Fietz, S; Bondzio, A; Moschos, A; Hertsch, B; Einspanier, R

    2008-06-01

    The aim of this study was to develop a specific myeloperoxidase (MPO) activity assay in the synovial fluid of horses and investigate whether MPO activity is increased in different forms of joint diseases. Synovial fluid samples were taken from affected joints from horses with osteoarthritis, chronic non-septic arthritis and septic arthritis, and from healthy control horses. MPO activity was measured using a specific modified o-dianisidine-assay containing 4-aminobenzoic acid hydrazide as a potent and specific inhibitor of the MPO. This assay is characterized by high reproducibility. The results reveal only a slight elevation of MPO activity in the synovial fluid of horses with osteoarthritis and chronic non-septic arthritis. However, in the cases of septic arthritis a significant increase in MPO activity was found when compared to the controls. In conclusion the first field study suggests that synovial fluid MPO may be used as a marker for septic arthritis in horses.

  15. Inhibition by polyphenolic phytochemicals and sulfurous compounds of the formation of 8-chloroguanosine mediated by hypochlorous acid, human myeloperoxidase, and activated human neutrophils.

    PubMed

    Nakano, Toshiki; Masuda, Mitsuharu; Suzuki, Toshinori; Ohshima, Hiroshi

    2012-01-01

    Hypochlorous acid (HOCl) produced by myeloperoxidase (MPO) of activated neutrophils can react with nucleic acid bases to form chlorinated nucleosides such as 8-chloroguanosine (Cl-Guo). Chlorination is enhanced by nicotine. We investigated the effects of various natural antioxidants including polyphenolic phytochemicals on the formation of Cl-Guo by HOCl in the presence and the absence of nicotine. Polyphenols, including catechins, curcumin, resveratrol, silibinin, and sulfurous compound α-lipoic acid, were found to inhibit both HOCl- and human MPO-induced Cl-Guo formation dose-dependently. Among the test compounds, (-)-epigallocatechin gallate (EGCG) showed the strongest inhibitory effect. Cl-Guo formation, mediated by activated human neutrophils in the presence of nicotine, was inhibited by EGCG, silibinin, and α-lipoic acid. These results suggest that polyphenols and sulfurous compounds have the potential to inhibit the induction of nucleobase damage mediated by chlorination, with possible application to reducing DNA damage associated with inflammation and cigarette-smoke inhalation.

  16. Myeloperoxidase-Related Chlorination Activity Is Positively Associated with Circulating Ceruloplasmin in Chronic Heart Failure Patients: Relationship with Neurohormonal, Inflammatory, and Nutritional Parameters

    PubMed Central

    Cabassi, Aderville; Binno, Simone Maurizio; Tedeschi, Stefano; Graiani, Gallia; Galizia, Cinzia; Bianconcini, Michele; Coghi, Pietro; Fellini, Federica; Ruffini, Livia; Govoni, Paolo; Piepoli, Massimo; Perlini, Stefano; Regolisti, Giuseppe; Fiaccadori, Enrico

    2015-01-01

    Rationale. Heart failure (HF) is accompanied by the development of an imbalance between oxygen- and nitric oxide-derived free radical production leading to protein nitration. Both chlorinating and peroxidase cycle of Myeloperoxidase (MPO) contribute to oxidative and nitrosative stress and are involved in tyrosine nitration of protein. Ceruloplasmin (Cp) has antioxidant function through its ferroxidase I (FeOxI) activity and has recently been proposed as a physiological defense mechanism against MPO inappropriate actions. Objective. We investigated the relationship between plasma MPO-related chlorinating activity, Cp and FeOxI, and nitrosative stress, inflammatory, neurohormonal, and nutritional biomarkers in HF patients. Methods and Results. In chronic HF patients (n = 81, 76 ± 9 years, NYHA Class II (26); Class III (29); Class IV (26)) and age-matched controls (n = 17, 75 ± 11 years, CTR), plasma MPO chlorinating activity, Cp, FeOxI, nitrated protein, free Malondialdehyde, BNP, norepinephrine, hsCRP, albumin, and prealbumin were measured. Plasma MPO chlorinating activity, Cp, BNP, norepinephrine, and hsCRP were increased in HF versus CTR. FeOxI, albumin, and prealbumin were decreased in HF. MPO-related chlorinating activity was positively related to Cp (r = 0.363, P < 0.001), nitrated protein, hsCRP, and BNP and inversely to albumin. Conclusions. Plasma MPO chlorinated activity is increased in elderly chronic HF patients and positively associated with Cp, inflammatory, neurohormonal, and nitrosative parameters suggesting a role in HF progression. PMID:26539521

  17. 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.

  18. Redundant contribution of myeloperoxidase-dependent systems to neutrophil-mediated killing of Escherichia coli.

    PubMed Central

    Rosen, H; Michel, B R

    1997-01-01

    Neutrophil microbicidal activity is a consequence of overlapping antimicrobial systems that vary in prominence according to the conditions of the neutrophil-microbe interaction, the nature of the microbe, and its metabolic state. In this study, normal, myeloperoxidase-deficient, and respiratory burst-deficient (chronic granulomatous disease [CGD]) neutrophils killed Escherichia coli with equivalent, high efficiencies. Killing by CGD and myeloperoxidase-deficient neutrophils was not augmented by supplements, such as exogenous H2O2 and myeloperoxidase, directed at ameliorating their metabolic defects, suggesting that nonoxidative microbicidal systems were sufficient for a full microbicidal effect. Neutrophils with an intact myeloperoxidase antimicrobial system (normal or appropriately supplemented deficient cells) were capable of rapidly suppressing E. coli DNA synthesis, while unsupplemented CGD or myeloperoxidase-deficient cells were far less effective, indicating that the myeloperoxidase system was active in normal neutrophils. The degree of DNA synthesis inhibition by myeloperoxidase-sufficient neutrophils could account, in a cell-free system, for most of the observed microbicidal activity. While the myeloperoxidase system was active and probably bactericidal, it was not rate limiting for microbicidal activity and appears to have been redundant with other microbicidal systems in the cell. Rapid and extensive inhibition of bacterial DNA synthesis appears to be an indicator of myeloperoxidase activity in neutrophils. PMID:9317024

  19. Myeloperoxidase, a catalyst for lipoprotein oxidation, is expressed in human atherosclerotic lesions.

    PubMed Central

    Daugherty, A; Dunn, J L; Rateri, D L; Heinecke, J W

    1994-01-01

    Oxidatively modified lipoproteins have been implicated in atherogenesis, but the mechanisms that promote oxidation in vivo have not been identified. Myeloperoxidase, a heme protein secreted by activated macrophages, generates reactive intermediates that oxidize lipoproteins in vitro. To explore the potential role of myeloperoxidase in the development of atherosclerosis, we determined whether the enzyme was present in surgically excised human vascular tissue. In detergent extracts of atherosclerotic arteries subjected to Western blotting, a rabbit polyclonal antibody monospecific for myeloperoxidase detected a 56-kD protein, the predicted molecular mass of the heavy subunit. Both the immunoreactive protein and authentic myeloperoxidase bound to a lectin-affinity column; after elution with methyl mannoside their apparent molecular masses were indistinguishable by nondenaturing size-exclusion chromatography. Peroxidase activity in detergent extracts of atherosclerotic lesions likewise bound to a lectin column and eluted with methyl mannoside. Moreover, eluted peroxidase generated the cytotoxic oxidant hypochlorous acid (HOCl), indicating that enzymatically active myeloperoxidase was present in lesions. Patterns of immunostaining of arterial tissue with antihuman myeloperoxidase antibodies were similar to those produced by an antimacrophage antibody, and were especially prominent in the shoulder region of transitional lesions. Intense foci of myeloperoxidase immunostaining also appeared adjacent to cholesterol clefts in lipid-rich regions of advanced atherosclerotic lesions. These findings identify myeloperoxidase as a component of human vascular lesions. Because this heme protein can generate reactive species that damage lipids and proteins, myeloperoxidase may contribute to atherogenesis by catalyzing oxidative reactions in the vascular wall. Images PMID:8040285

  20. A Phytochemical-rich Multivitamin-multimineral Supplement Is Bioavailable and Reduces Serum Oxidized Low-density Lipoprotein, Myeloperoxidase, and Plasminogen Activator Inhibitor-1 in a Four-week Pilot trial of Healthy Individuals

    PubMed Central

    Desai, Anuradha; Lamb, Joseph J.; Chang, Jyh-Lurn; Darland, Gary; Konda, Veera R.

    2014-01-01

    Background: A multivitamin-multimineral supplement combined with a diverse blend of bioactive phytochemicals may provide additional antioxidant capacity and anti-inflammatory property for overall health. This convenient feature may be useful for individuals who want to increase their intake of phytochemicals. Methods: We conducted a pilot study in 15 healthy individuals (8 women and 7 men, mean age 41.7±14.9 years, mean body mass index 28.0±5.6) to investigate the effects of this novel formulation on biomarkers associated with oxidative stress and inflammation. After a 2-week diet that limited intake of fruits and vegetables to 2 servings/day, participants continued with the same restricted diet but began consuming 2 tablets of the study product for the subsequent 4 weeks. Fasting blood samples collected at Week 2 and Week 6 were analyzed and compared using paired t-tests for levels of carotenoids, folate, vitamin B12, homocysteine, oxidized low-density lipoprotein cholesterol (oxLDL), high-sensitivity C-reactive protein (hs-CRP), F2-isoprostane, plasminogen activator inhibitor-1 (PAI-1), and myeloperoxidase. Noninvasive peripheral arterial tonometry (EndoPAT) was also measured. Results: After 4 weeks of supplementation, plasma levels of carotenoids, folate, and vitamin B12, but not homocysteine, were significantly increased (P<.05). Serum levels of oxLDL, PAI-1 and myeloperoxidase were significantly reduced (P<.05), but F2-isoprostane, hs-CRP, and EndoPAT measures were unchanged compared with baseline. The study product was well tolerated. Conclusions: This nutritional supplement is bioavailable as indicated by the significant increase in plasma carotenoids, vitamin B12, and folate levels and may provide health benefits by significantly reducing serum levels of oxLDL, myeloperoxidase, and PAI-1 in healthy individuals. PMID:24808980

  1. Neutrophil myeloperoxidase destruction by ultraviolet irradiation

    SciTech Connect

    Hanker, J.; Giammara, B.; Strauss, G.

    1988-01-01

    The peroxidase activity of enriched leukocyte preparations on coverslips was determined cytochemically with a newly developed method. The techniques utilizes diaminobenzidine medium and cupric nitrate intensification and is suitable for analysis with light microscopy, SEM, and TEM. Blood specimens from control individuals were studied with and without in vitro UV irradiation and compared with those from psoriasis patients exposed therapeutically to various types of UV in phototherapy. All UV irradiated samples showed diminished neutrophil myeloperoxidase (MP) activity although that of the principal eosinophil peroxidase was unaffected. The SEMs supported the contention that decreased neutrophil MP activity might be related to UV induced degranulation. It is believed to be possible, eventually, to equate the observed MP degranulation effect after UV irradiation with diminished ability to fight bacterial infections.

  2. Neutrophil-derived microparticles induce myeloperoxidase-mediated damage of vascular endothelial cells

    PubMed Central

    2014-01-01

    Background Upon activation neutrophil releases microparticles - small plasma membrane vesicles that contain cell surface proteins and cytoplasmic matter, with biological activities. In this study we investigated the potential role of myeloperoxidase in the endothelial cell injury caused by neutrophil-derived microparticles. Results Microparticles were produced by activating human neutrophils with a calcium ionophore and characterized by flow cytometry and transmission and scanning electron microscopy. Myeloperoxidase activity was measured by luminol-dependent chemiluminescence. Neutrophil microparticles-induced injuries and morphological alterations in human umbilical vein endothelial cells (HUVECs) were evaluated by microscopy and flow cytometry. Neutrophil microparticles were characterized as structures bounded by lipid bilayers and were less than 1 μm in diameter. The microparticles also expressed CD66b, CD62L and myeloperoxidase, which are all commonly expressed on the surface of neutrophils, as well as exposition of phosphatidylserine. The activity of the myeloperoxidase present on the microparticles was confirmed by hypochlorous acid detection. This compound is only catalyzed by myeloperoxidase in the presence of hydrogen peroxide and chloride ion. The addition of sodium azide or taurine inhibited and reduced enzymatic activity, respectively. Exposure of HUVEC to neutrophil microparticles induced a loss of cell membrane integrity and morphological changes. The addition of sodium azide or myeloperoxidase-specific inhibitor-I consistently reduced the injury to the endothelial cells. Taurine addition reduced HUVEC morphological changes. Conclusions We have demonstrated the presence of active myeloperoxidase in neutrophil microparticles and that the microparticle-associated myeloperoxidase cause injury to endothelial cells. Hence, the microparticle-associated myeloperoxidase-hydrogen peroxide-chloride system may contribute to widespread endothelial cell damage

  3. Deconstruction of Activity-Dependent Covalent Modification of Heme in Human Neutrophil Myeloperoxidase by Multistage Mass Spectrometry (MS[superscript 4])

    SciTech Connect

    Geoghegan, Kieran F.; Varghese, Alison H.; Feng, Xidong; Bessire, Andrew J.; Conboy, James J.; Ruggeri, Roger B.; Ahn, Kay; Spath, Samantha N.; Filippov, Sergey V.; Conrad, Steven J.; Carpino, Philip A.; Guimarães, Cristiano R.W.; Vajdos, Felix F.

    2013-03-07

    Myeloperoxidase (MPO) is known to be inactivated and covalently modified by treatment with hydrogen peroxide and agents similar to 3-(2-ethoxypropyl)-2-thioxo-2,3-dihydro-1H-purin-6(9H)-one (1), a 254.08 Da derivative of 2-thioxanthine. Peptide mapping by liquid chromatography and mass spectrometry detected modification by 1 in a labile peptide-heme-peptide fragment of the enzyme, accompanied by a mass increase of 252.08 Da. The loss of two hydrogen atoms was consistent with mechanism-based oxidative coupling. Multistage mass spectrometry (MS{sup 4}) of the modified fragment in an ion trap/Orbitrap spectrometer demonstrated that 1 was coupled directly to heme. Use of a 10 amu window delivered the full isotopic envelope of each precursor ion to collision-induced dissociation, preserving definitive isotopic profiles for iron-containing fragments through successive steps of multistage mass spectrometry. Iron isotope signatures and accurate mass measurements supported the structural assignments. Crystallographic analysis confirmed linkage between the methyl substituent of the heme pyrrole D ring and the sulfur atom of 1. The final orientation of 1 perpendicular to the plane of the heme ring suggested a mechanism consisting of two consecutive one-electron oxidations of 1 by MPO. Multistage mass spectrometry using stage-specific collision energies permits stepwise deconstruction of modifications of heme enzymes containing covalent links between the heme group and the polypeptide chain.

  4. Accumulator

    NASA Technical Reports Server (NTRS)

    Fenwick, J. R.; Karigan, G. H. (Inventor)

    1977-01-01

    An accumulator particularly adapted for use in controlling the pressure of a stream of fluid in its liquid phase utilizing the fluid in its gaseous phase was designed. The accumulator is characterized by a shell defining a pressure chamber having an entry throat for a liquid and adapted to be connected in contiguous relation with a selected conduit having a stream of fluid flowing through the conduit in its liquid phase. A pressure and volume stabilization tube, including an array of pressure relief perforations is projected into the chamber with the perforations disposed adjacent to the entry throat for accommodating a discharge of the fluid in either gaseous or liquid phases, while a gas inlet and liquid to gas conversion system is provided, the chamber is connected with a source of the fluid for continuously pressuring the chamber for controlling the pressure of the stream of liquid.

  5. Myeloperoxidase in chronic kidney disease.

    PubMed

    Madhusudhana Rao, A; Anand, Usha; Anand, C V

    2011-01-01

    Numerous lines of evidence implicate a role of myeloperoxidase (MPO) in the pathogenesis of cardiovascular disease (CVD). It is a well accepted fact that patients with chronic kidney disease (CKD) are at an increased risk for CVD. MPO is a pro-oxidant enzyme which could be involved in the increased susceptibility of these patients to CVD. Hence, the levels of plasma MPO was determined in healthy controls as well as in patients with CKD [stratified with the level of their kidney failure as CKD stages II-V (end stage renal disease)]. Plasma MPO was assayed by a spectrophotometric method. Serum urea and creatinine were estimated on a clinical chemistry analyzer using standard laboratory procedures. The mean plasma MPO levels were significantly lower with advancing stages of renal failure (P < 0.001). There was a positive correlation between MPO and GFR (r = +0.89, P < 0.001) and a negative correlation with urea (r = -0.85, P < 0.001) and creatinine (r = -0.82, P < 0.001). While an inverse association was observed between plasma MPO and urea in CKD patients, such an association was not observed in control subjects (P = 0.43). In conclusion, the decline in plasma MPO levels may be due to the inhibitory effect of uraemic toxins on the enzyme.

  6. Myeloperoxidase: a target for new drug development?

    PubMed

    Malle, E; Furtmüller, P G; Sattler, W; Obinger, C

    2007-11-01

    Myeloperoxidase (MPO), a member of the haem peroxidase-cyclooxygenase superfamily, is abundantly expressed in neutrophils and to a lesser extent in monocytes and certain type of macrophages. MPO participates in innate immune defence mechanism through formation of microbicidal reactive oxidants and diffusible radical species. A unique activity of MPO is its ability to use chloride as a cosubstrate with hydrogen peroxide to generate chlorinating oxidants such as hypochlorous acid, a potent antimicrobial agent. However, evidence has emerged that MPO-derived oxidants contribute to tissue damage and the initiation and propagation of acute and chronic vascular inflammatory disease. The fact that circulating levels of MPO have been shown to predict risks for major adverse cardiac events and that levels of MPO-derived chlorinated compounds are specific biomarkers for disease progression, has attracted considerable interest in the development of therapeutically useful MPO inhibitors. Today, detailed information on the structure of ferric MPO and its complexes with low- and high-spin ligands is available. This, together with a thorough understanding of reaction mechanisms including redox properties of intermediates, enables a rationale attempt in developing specific MPO inhibitors that still maintain MPO activity during host defence and bacterial killing but interfere with pathophysiologically persistent activation of MPO. The various approaches to inhibit enzyme activity of MPO and to ameliorate adverse effects of MPO-derived oxidants will be discussed. Emphasis will be put on mechanism-based inhibitors and high-throughput screening of compounds as well as the discussion of physiologically useful HOCl scavengers.

  7. Myeloperoxidase binds to and kills Entamoeba histolytica trophozoites.

    PubMed

    Pacheco-Yépez, J; Rivera-Aguilar, V; Barbosa-Cabrera, E; Rojas Hernández, S; Jarillo-Luna, R A; Campos-Rodríguez, R

    2011-05-01

    During amebic invasion, neutrophils are a key component in either protecting against invading trophozoites or contributing to tissue damage. Upon degranulating or being lysed, neutrophils release toxic substances that can kill amebas as well as damage host tissue. In a previous study we identified a protein from nonspecifically stimulated peritoneal exudates of hamster that has peroxidase and marked amebicidal activity. In the current study we analyzed the in vitro amebicidal effect of purified hamster myeloperoxidase (MPO). The results demonstrate that MPO must bind directly to the surface of Entamoeba histolytica trophozoites in order to carry out amebicidal activity by using the H(2) O(2) produced by the amebas themselves. Myeloperoxidase-incubated amebas showed important morphological and ultrastructural alterations that increased with incubation time. Changes included an increase of vacuoles in the cytoplasm, a decrease of glycogen, alterations of nuclear morphology and disturbances in the plasma membrane culminating in complete ameba destruction.

  8. Myeloperoxidase deficiency attenuates nitrogen mustard-induced skin injuries

    PubMed Central

    Jain, Anil K; Tewari-Singh, Neera; Inturi, Swetha; Orlicky, David J.; White, Carl W.; Agarwal, Rajesh

    2014-01-01

    The pathologic mechanisms of skin injuries, following the acute inflammatory response induced by vesicating agents sulfur mustard (SM) and nitrogen mustard (NM) exposure, are poorly understood. Neutrophils which accumulate at the site of injury, abundantly express myeloperoxidase (MPO), a heme protein that is implicated in oxidant-related antimicrobial and cytotoxic responses. Our previous studies have shown that exposure to SM analog 2-chloroethyl ethyl sulfide (CEES) or NM results in an inflammatory response including increased neutrophilic infiltration and MPO activity. To further define the role of neutrophil-derived MPO in NM-induced skin injury, here we used a genetic approach and examined the effect of NM exposure (12 h and 24 h) on previously established injury endpoints in C57BL/6J wild type (WT) and B6.129X1-MPOtm1Lus/J mice (MPO KO), homozygous null for MPO gene. NM exposure caused a significant increase in skin bi-fold thickness, epidermal thickness, microvesication, DNA damage and apoptosis in WT mice compared to MPO KO mice. MPO KO mice showed relatively insignificant effect. Similarly, NM-induced increases in the expression of inflammatory and proteolytic mediators, including COX-2, iNOS and MMP-9 in WT mice, while having a significantly lower effect in MPO KO mice. Collectively, these results show that MPO, which generates microbicidal oxidants, plays an important role in NM-induced skin injuries. This suggests the development of mechanism-based treatments against NM- and SM-induced skin injuries that inhibit MPO activity and attenuate MPO-derived oxidants. PMID:24631667

  9. Myeloperoxidase activity and its corresponding mRNA expression as well as gene polymorphism in the population living in the coal-burning endemic fluorosis area in Guizhou of China.

    PubMed

    Zhang, Ting; Shan, Ke-Ren; Tu, Xi; He, Yan; Pei, Jin-Jing; Guan, Zhi-Zhong

    2013-06-01

    The myeloperoxidase (MPO) activity and its corresponding mRNA expression as well as gene polymorphism were investigated in the population who live in the endemic fluorosis area. In the study, 150 people were selected from the coal-burning endemic fluorosis area and 150 normal persons from the non-fluorosis area in Guizhou province of China. The blood samples were collected from these people. The activity of MPO in the plasma was determined by spectrophotometer; the expression of MPO mRNA was measured by employing real-time polymerase chain reaction; DNAs were extracted from the leucocytes in blood and five SNP genotypes of MPO promoter gene detected by a multiplex genotyping method, adapter-ligation-mediated allele-specific amplification. The results showed that the MPO activity and its corresponding mRNA in blood were significantly increased in the population living in the area of fluorosis. The different genotype frequencies of MPO, including -1228G/A, -585T/C, -463G/A, and -163C/T, and the three haplotypes with higher frequencies, including -163C-463G-585T-1228G-1276T, -163C-463G-585T-1228G-1276C, and -163C-463G-585T-1228A-1276T, were significantly associated with fluorosis. The results indicated that the elevated activity of MPO induced by endemic fluorosis may be connected in mechanism to the stimulated expression of MPO mRNA and the changed gene polymorphism.

  10. Mechanism of reaction of myeloperoxidase with nitrite.

    PubMed

    Burner, U; Furtmuller, P G; Kettle, A J; Koppenol, W H; Obinger, C

    2000-07-07

    Myeloperoxidase (MPO) is a major neutrophil protein and may be involved in the nitration of tyrosine residues observed in a wide range of inflammatory diseases that involve neutrophils and macrophage activation. In order to clarify if nitrite could be a physiological substrate of myeloperoxidase, we investigated the reactions of the ferric enzyme and its redox intermediates, compound I and compound II, with nitrite under pre-steady state conditions by using sequential mixing stopped-flow analysis in the pH range 4-8. At 15 degrees C the rate of formation of the low spin MPO-nitrite complex is (2.5 +/- 0.2) x 10(4) m(-1) s(-1) at pH 7 and (2.2 +/- 0.7) x 10(6) m(-1) s(-1) at pH 5. The dissociation constant of nitrite bound to the native enzyme is 2.3 +/- 0.1 mm at pH 7 and 31.3 +/- 0.5 micrometer at pH 5. Nitrite is oxidized by two one-electron steps in the MPO peroxidase cycle. The second-order rate constant of reduction of compound I to compound II at 15 degrees C is (2.0 +/- 0.2) x 10(6) m(-1) s(-1) at pH 7 and (1.1 +/- 0.2) x 10(7) m(-1) s(-1) at pH 5. The rate constant of reduction of compound II to the ferric native enzyme at 15 degrees C is (5.5 +/- 0.1) x 10(2) m(-1) s(-1) at pH 7 and (8.9 +/- 1.6) x 10(4) m(-1) s(-1) at pH 5. pH dependence studies suggest that both complex formation between the ferric enzyme and nitrite and nitrite oxidation by compounds I and II are controlled by a residue with a pK(a) of (4.3 +/- 0.3). Protonation of this group (which is most likely the distal histidine) is necessary for optimum nitrite binding and oxidation.

  11. Rational drug design applied to myeloperoxidase inhibition.

    PubMed

    Van Antwerpen, P; Zouaoui Boudjeltia, K

    2015-06-01

    Rational drug design is a general approach using protein-structure technique in which the discovery of a ligand can be driven either by chance, screening, or rational theory. Myeloperoxidase (MPO) was rapidly identified as a therapeutical target because of its involvement in chronic inflammatory syndromes. In this context, the research of MPO inhibitors was intensified and development of new chemical entities was rationally driven by the research of ligands that enter into the MPO catalytic pocket. Actually, as soon as crystallography data of MPO have become available and its structure was virtually designed, the rational drug design has been applied to this peroxidase. Pharmaceutical industries and academic laboratories apply rational drug design on MPO by either optimizing known inhibitors or searching new molecules by high-throughput virtual screening. By these ways, they were able to find efficient MPO inhibitors and understand their interactions with the enzyme. During this quest of MPO inhibition, it appears that Glu268 is a crucial residue in order to optimize ligand-target interaction. This amino acid should be carefully considered by medicinal chemist when they design inhibitors interfering with MPO activity.

  12. Most proteinase3- and myeloperoxidase-antineutrophil cytoplasmic antibodies enzyme-linked immunosorbent assays perform less well in treated small-vessel vasculitis than in active disease.

    PubMed

    Savige, Judy; Trevisin, Michelle; Hayman, Matthew; Pollock, Wendy

    2009-06-01

    Antineutrophil cytoplasmic antibodies (ANCA) levels have been thought to follow disease activity, with levels being high at presentation, declining with treatment and increasing just before relapse. However, we have shown that ANCA often persist for many years in patients with clinically inactive Wegener's granulomatosis. ANCA assays are less sensitive for treated disease than for active disease, and the levels in treated patients produce different results in different assay systems. ANCA often persist for years without relapse, and the risk of relapse probably depend on levels that are critical for any individual patient. The capture enzyme-linked immunosorbent assays may be more sensitive in detecting early relapse. Relapse is more common when ANCA levels are high but, although elevated, ANCA levels are lower in relapse than at presentation. Standardized ANCA levels for the definitions of remission and relapse may not be possible, and the optimal ANCA testing protocol for treated disease remains unclear.

  13. Thioxo-dihydroquinazolin-one Compounds as Novel Inhibitors of Myeloperoxidase

    PubMed Central

    2015-01-01

    Myeloperoxidase (MPO) is a key antimicrobial enzyme, playing a normal role in host defense, but also contributing to inflammatory conditions including neuroinflammatory diseases such as Parkinson’s and Alzheimer’s. We synthesized and characterized more than 50 quinazolin-4(1H)-one derivatives and showed that this class of compounds inhibits MPO with IC50 values as low as 100 nM. Representative compounds showed partially reversible inhibition that was competitive with respect to Amplex Red substrate and did not result in the accumulation of MPO Compound II. Members of this group show promise for therapeutic development for the treatment of diseases in which inflammation plays a pathogenic role. PMID:26487910

  14. Blood pressure reduction following accumulated physical activity in prehypertensive

    PubMed Central

    Saxena, Yogesh; Gupta, Rani; Moinuddin, Arsalan; Narwal, Ravinder

    2016-01-01

    Context: Accumulated moderate physical activity (PA) for 30 min in a day is the only recommended treatment of prehypertension. Objective: We investigated autonomic modulation as a possible mechanism for the decrease in blood pressure (BP) during the rest periods in each 10 min session of PA. Design, Setting, and Participants: We conducted a single-blind randomized multi-arm control trial on 40 prehypertensive (pre-HT) young male adults. Methods: Participants were randomly divided by using random number table into four groups. Control (no intervention); Group 1 (walking at 50% of predicted VO2 peak); Group 2 (walking at 60% of predicted VO2 peak); Group 3 (walking at 70% of predicted VO2 peak). BP, heart rate variability (HRV), and heart rate recovery 1 min (HRR 1 min) were measured at baseline and during the rest period after each session of 10 min over 30 min of accumulated physical activity (PAcumm). Results: Significant diastolic BP (DBP) reduction (P < 0.001) was observed during the rest period after each session of PAcumm in intervention groups. An average reduction in DBP was more in pre-HT undertaking PAcumm at 70% of predicted VO2 Peak. Decrease in the mean value of low-frequency (LF) and LF/high-frequency ratio was observed following PAcumm in all intervention groups irrespective of the intensity of PA. No significant association of reduction of BP with HRV and HRR 1 s was observed. Conclusion: Reduction in BP was observed during the rest period after each 10 min session of PAcumm irrespective of the intensity of PA. Autonomic modulation does not seem to be the possible mechanism for the reduction in BP during the sessions. PMID:27843840

  15. Fecal Myeloperoxidase as a Biomarker for Inflammatory Bowel Disease

    PubMed Central

    Hansberry, David R; Shah, Kush; Agarwal, Nitin

    2017-01-01

    Inflammatory bowel disease (IBD) is a chronic condition involving the inflammation of the colon and small intestine. IBD affects as many as 1.4 million people in the U.S. alone and costs the health care industry over $1.7 billion annually. Managing IBD normally requires invasive and often discomforting diagnostic tests. In an effort to alleviate the painful and costly nature of traditional diagnosis, there has been increasing research initiative focused on noninvasive biomarkers. PubMed, provided by the United States National Library of Medicine (NLM) at the National Institutes of Health, was utilized with the following search terms: 1) myeloperoxidase (MPO) 2), inflammatory bowel disease (IBD), and 3) neutrophils. The following terms were used interchangeably with search terms 1-3: 4) costs, 5) biomarkers, 6) review, and 7) etiology. In the context of IBD, myeloperoxidase (MPO), a lysosomal protein found in neutrophils, may serve as a viable biomarker for assessing disease status. Several studies demonstrated increased levels of neutrophils in patients with active IBD. Furthermore, studies have found significantly higher levels of MPO in patients with active IBD compared to patients without IBD as well as patients with inactive IBD. MPO is also expressed in higher concentrations in patients with more severe forms of IBD. When measuring treatment efficacy, MPO levels are indicative of the quality of response. MPO may serve as an important diagnostic and prognostic tool in assessing IBD status.  PMID:28286723

  16. Leukocyte myeloperoxidase deficiency and disseminated candidiasis: the role of myeloperoxidase in resistance to Candida infection

    PubMed Central

    Lehrer, Robert I.; Cline, Martin J.

    1969-01-01

    The neutrophils and monocytes of a patient with disseminated candidiasis were found to lack detectable levels of the lysosomal enzyme myeloperoxidase (MPO), although they had normal levels of other granule-associated enzymes. Leukocytes from one of the patient's sisters also lacked detectable MPO; leukocytes from his four sons contained approximately one-third of mean normal peroxidase levels. Neither the patient nor his affected relatives had experienced frequent or unusual bacterial infections. The phagocytic activity of the patient's MPO-deficient neutrophils was intact, and the cells displayed normal morphologic and metabolic responses to phagocytosis. In contrast to normal leukocytes which killed 30.5±7.3% of ingested Candida albicans in 1 hr, however, the patient's neutrophils killed virtually none. His leukocytes also failed to kill the strain of C. albicans recovered from his lesions, as well as other Candida species. These MPO-deficient neutrophils killed Serratia marcescens and Staphylococens aureus 502A at an abnormally slow rate, requiring 3-4 hr to achieve the bactericidal effect attained by normal leukocytes after 45 min. No other abnormalities in his cellular or humoral immune responses were demonstrated. These findings suggest that hereditary MPO deficiency is transmitted as an autosomal recessive characteristic, that the homozygous state conveys enhanced susceptibility to disseminated candidiasis, and that MPO is necessary for candidacidal activity in human neutrophils. Although lending support to the suggested bactericidal role of MPO in leukocytes, the data indicate that alternative bactericidal mechanisms, effective in the absence of MPO, are functionally dominant in the human neutrophil. Images PMID:5796360

  17. Myeloperoxidase-produced Genomic DNA-centered Radicals and Protection by Resveratrol

    EPA Science Inventory

    Myeloperoxidase (MPO) released by activated neutrophils, production of hypochlorous acid (HOCI) and oxidation of the genomic DNA in epithelial cells is thought to initiate and promote carcinogenesis. In this study we applied the 5,5-dimethyl-l-pyrroline N-oxide (DMPO)-based i;nmu...

  18. Preparation of Monoclonal Antibodies and a Simple Myeloperoxidase-Immunosorbent Assay for Detecting Human Myeloperoxidase.

    PubMed

    Bian, Zhi-Ping; Li, Xiong-Zhi; Wu, Heng-Fang; Xu, Jin-Dan; Gu, Chun-Rong; Chen, Xiang-Jian; Yang, Di

    2016-04-01

    Myeloperoxidase (MPO), a leukocyte hemoprotein released from neutrophils, is thought to be a potential participant in plaque formation and plaque rupture. Therefore, MPO is regarded as an early marker predicting the risk for atherosclerosis, especially for coronary artery disease and acute coronary syndrome. We generated hybridoma clones 1E3 and 3E8 secreting monoclonal antibodies (mAbs) specific to human MPO. BALB/c mice were immunized with MPO protein purified from human neutrophils. Splenocytes from these mice were fused with the mouse myeloma cell line SP2/0. Based on isotyping of the mAbs, both clones 1E3 and 3E8 were referred to the IgG1 subclass. The specificities of 1E3 and 3E8 were assessed by enzyme-linked immunosorbent assay (ELISA), and only 3E8 was confirmed by western blot. We developed a simple MPO-immunosorbent assay (MPO-ISA) on microplate based on both the immune activity and peroxidase activity of MPO. The mAb secreted by clone 3E8 was chosen as coating antibody to capture the plasma MPO without interfering with the peroxidase activity of MPO. Then, tetramethylbenzidine substrate was added to the microwell directly, catalyzed by captured MPO, and a colored product was formed. The simple MPO-ISA test has a sensitivity of 3.68 ng/mL. The linear concentration of MPO-ISA for commercial MPO standard ranged to 250 ng/mL. The average recovery rate is 101.02%. The imprecision within-day was <10% at three different MPO levels. The imprecision between-day was <10% at low and middle MPO levels and varied to 14.61% at the high MPO level. We found that the established MPO-ISA can detect the plasma MPO from human and cavy, but not from mouse and rat. Compared with the commercial human MPO ELISA assay, the MPO-ISA can be used to detect the natural human MPO protein, but not recombinant MPO polypeptides. The generated mAbs and MPO-ISA test may be useful tools to assess risk for inflammation and cardiac events.

  19. Anti-myeloperoxidase antibodies attenuate the monocyte response to LPS and shape macrophage development.

    PubMed

    Popat, Reena J; Hakki, Seran; Thakker, Alpesh; Coughlan, Alice M; Watson, Julie; Little, Mark A; Spickett, Corinne M; Lavender, Paul; Afzali, Behdad; Kemper, Claudia; Robson, Michael G

    2017-01-26

    Anti-neutrophil cytoplasmic antibody (ANCA) vasculitis is characterized by the presence of autoantibodies to myeloperoxidase and proteinase-3, which bind monocytes in addition to neutrophils. While a pathological effect on neutrophils is acknowledged, the impact of ANCA on monocyte function is less well understood. Using IgG from patients we investigated the effect of these autoantibodies on monocytes and found that anti-myeloperoxidase antibodies (MPO-ANCA) reduced both IL-10 and IL-6 secretion in response to LPS. This reduction in IL-10 and IL-6 depended on Fc receptors and enzymatic myeloperoxidase and was accompanied by a significant reduction in TLR-driven signaling pathways. Aligning with changes in TLR signals, oxidized phospholipids, which function as TLR4 antagonists, were increased in monocytes in the presence of MPO-ANCA. We further observed that MPO-ANCA increased monocyte survival and differentiation to macrophages by stimulating CSF-1 production. However, this was independent of myeloperoxidase enzymatic activity and TLR signaling. Macrophages differentiated in the presence of MPO-ANCA secreted more TGF-β and further promoted the development of IL-10- and TGF-β-secreting CD4(+) T cells. Thus, MPO-ANCA may promote inflammation by reducing the secretion of antiinflammatory IL-10 from monocytes, and MPO-ANCA can alter the development of macrophages and T cells to potentially promote fibrosis.

  20. Anti-myeloperoxidase antibodies attenuate the monocyte response to LPS and shape macrophage development

    PubMed Central

    Popat, Reena J.; Hakki, Seran; Coughlan, Alice M.; Watson, Julie; Little, Mark A.; Spickett, Corinne M.; Lavender, Paul; Afzali, Behdad; Kemper, Claudia; Robson, Michael G.

    2017-01-01

    Anti-neutrophil cytoplasmic antibody (ANCA) vasculitis is characterized by the presence of autoantibodies to myeloperoxidase and proteinase-3, which bind monocytes in addition to neutrophils. While a pathological effect on neutrophils is acknowledged, the impact of ANCA on monocyte function is less well understood. Using IgG from patients we investigated the effect of these autoantibodies on monocytes and found that anti-myeloperoxidase antibodies (MPO-ANCA) reduced both IL-10 and IL-6 secretion in response to LPS. This reduction in IL-10 and IL-6 depended on Fc receptors and enzymatic myeloperoxidase and was accompanied by a significant reduction in TLR-driven signaling pathways. Aligning with changes in TLR signals, oxidized phospholipids, which function as TLR4 antagonists, were increased in monocytes in the presence of MPO-ANCA. We further observed that MPO-ANCA increased monocyte survival and differentiation to macrophages by stimulating CSF-1 production. However, this was independent of myeloperoxidase enzymatic activity and TLR signaling. Macrophages differentiated in the presence of MPO-ANCA secreted more TGF-β and further promoted the development of IL-10– and TGF-β–secreting CD4+ T cells. Thus, MPO-ANCA may promote inflammation by reducing the secretion of antiinflammatory IL-10 from monocytes, and MPO-ANCA can alter the development of macrophages and T cells to potentially promote fibrosis. PMID:28138552

  1. From Dynamic Combinatorial Chemistry to in Vivo Evaluation of Reversible and Irreversible Myeloperoxidase Inhibitors.

    PubMed

    Soubhye, Jalal; Gelbcke, Michel; Van Antwerpen, Pierre; Dufrasne, François; Boufadi, Mokhtaria Yasmina; Nève, Jean; Furtmüller, Paul G; Obinger, Christian; Zouaoui Boudjeltia, Karim; Meyer, Franck

    2017-02-09

    The implementation of dynamic combinatorial libraries allowed the determination of highly active reversible and irreversible inhibitors of myeloperoxidase (MPO) at the nanomolar level. Docking experiments highlighted the interaction between the most active ligands and MPO, and further kinetic studies defined the mode of inhibition of these compounds. Finally, in vivo evaluation showed that one dose of irreversible inhibitors is able to suppress the activity of MPO after inducing inflammation.

  2. EFFECT OF VAPOR-PHASE BIOREACTOR OPERATION ON BIOMASS ACCUMULATION, DISTRIBUTION, AND ACTIVITY. (R826168)

    EPA Science Inventory

    Excess biomass accumulation and activity loss in vapor-phase bioreactors (VPBs) can lead to unreliable long-term operation. In this study, temporal and spatial variations in biomass accumulation, distribution and activity in VPBs treating toluene-contaminated air were monitored o...

  3. 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.

  4. Categorisation through evidence accumulation in an active vision system

    NASA Astrophysics Data System (ADS)

    Mirolli, Marco; Ferrauto, Tomassino; Nolfi, Stefano

    2010-12-01

    In this paper, we present an artificial vision system that is trained with a genetic algorithm for categorising five different kinds of images (letters) of different sizes. The system, which has a limited field of view, can move its eye so as to explore the images visually. The analysis of the system at the end of the training process indicates that correct categorisation is achieved by (1) exploiting sensory-motor coordination so as to experience stimuli that facilitate discrimination, and (2) integrating perceptual and/or motor information over time through a process of accumulation of partially conflicting evidence. We discuss our results with respect to the possible different strategies for categorisation and to the possible roles that action can play in perception.

  5. Myeloperoxidase-antineutrophil Cytoplasmic Antibodies with Cytoplasmic Fluorescence Pattern.

    PubMed

    Chhabra, Seema; Minz, Ranjana Walker; Goyal, Lekha; Sharma, Nidhi

    2010-01-01

    We report here two rare cases of myeloperoxidase-antineutrophil cytoplasmic antibody (MPO-ANCA)-positive Wegener's granulomatosis (limited variant) which deceptively produced a cytoplasmic (C-ANCA) pattern on indirect immunofluorescence.

  6. Ascorbate in aqueous humor protects against myeloperoxidase-induced oxidation.

    PubMed Central

    Rosenbaum, J. T.; Howes, E. L.; English, D.

    1985-01-01

    Chemotactic factors can cause polymorphonuclear leukocytes to release the contents of azurophilic granules, including the enzymes beta-glucuronidase and myeloperoxidase. In the presence of aqueous humor from the anterior chamber of the rabbit eye, the supernatant from stimulated leukocytes contains beta-glucuronidase, but myeloperoxidase is not detectable. Studies with aqueous humor and partially purified human myeloperoxidase suggest that this phenomenon is not due to a failure of enzyme release. The factor responsible for the inability to detect MPO in the assay system is heat-labile, dialyzable, and reversed by ascorbate oxidase. Comparable assay inhibition is produced by ascorbic acid at a concentration present in either human or rabbit aqueous humor. The ability of aqueous humor to protect against myeloperoxidase-induced oxidation may contribute to several diverse phenomena, including the susceptibility of the eye to Candida infection and a prolonged half-life for several inflammatory mediators in the anterior chamber. PMID:2992283

  7. Myeloperoxidase, paraoxonase-1, and HDL form a functional ternary complex.

    PubMed

    Huang, Ying; Wu, Zhiping; Riwanto, Meliana; Gao, Shengqiang; Levison, Bruce S; Gu, Xiaodong; Fu, Xiaoming; Wagner, Matthew A; Besler, Christian; Gerstenecker, Gary; Zhang, Renliang; Li, Xin-Min; DiDonato, Anthony J; Gogonea, Valentin; Tang, W H Wilson; Smith, Jonathan D; Plow, Edward F; Fox, Paul L; Shih, Diana M; Lusis, Aldons J; Fisher, Edward A; DiDonato, Joseph A; Landmesser, Ulf; Hazen, Stanley L

    2013-09-01

    Myeloperoxidase (MPO) and paraoxonase 1 (PON1) are high-density lipoprotein-associated (HDL-associated) proteins mechanistically linked to inflammation, oxidant stress, and atherosclerosis. MPO is a source of ROS during inflammation and can oxidize apolipoprotein A1 (APOA1) of HDL, impairing its atheroprotective functions. In contrast, PON1 fosters systemic antioxidant effects and promotes some of the atheroprotective properties attributed to HDL. Here, we demonstrate that MPO, PON1, and HDL bind to one another, forming a ternary complex, wherein PON1 partially inhibits MPO activity, while MPO inactivates PON1. MPO oxidizes PON1 on tyrosine 71 (Tyr71), a modified residue found in human atheroma that is critical for HDL binding and PON1 function. Acute inflammation model studies with transgenic and knockout mice for either PON1 or MPO confirmed that MPO and PON1 reciprocally modulate each other's function in vivo. Further structure and function studies identified critical contact sites between APOA1 within HDL, PON1, and MPO, and proteomics studies of HDL recovered from acute coronary syndrome (ACS) subjects revealed enhanced chlorotyrosine content, site-specific PON1 methionine oxidation, and reduced PON1 activity. HDL thus serves as a scaffold upon which MPO and PON1 interact during inflammation, whereupon PON1 binding partially inhibits MPO activity, and MPO promotes site-specific oxidative modification and impairment of PON1 and APOA1 function.

  8. Myeloperoxidase, paraoxonase-1, and HDL form a functional ternary complex

    PubMed Central

    Huang, Ying; Wu, Zhiping; Riwanto, Meliana; Gao, Shengqiang; Levison, Bruce S.; Gu, Xiaodong; Fu, Xiaoming; Wagner, Matthew A.; Besler, Christian; Gerstenecker, Gary; Zhang, Renliang; Li, Xin-Min; DiDonato, Anthony J.; Gogonea, Valentin; Tang, W.H. Wilson; Smith, Jonathan D.; Plow, Edward F.; Fox, Paul L.; Shih, Diana M.; Lusis, Aldons J.; Fisher, Edward A.; DiDonato, Joseph A.; Landmesser, Ulf; Hazen, Stanley L.

    2013-01-01

    Myeloperoxidase (MPO) and paraoxonase 1 (PON1) are high-density lipoprotein–associated (HDL-associated) proteins mechanistically linked to inflammation, oxidant stress, and atherosclerosis. MPO is a source of ROS during inflammation and can oxidize apolipoprotein A1 (APOA1) of HDL, impairing its atheroprotective functions. In contrast, PON1 fosters systemic antioxidant effects and promotes some of the atheroprotective properties attributed to HDL. Here, we demonstrate that MPO, PON1, and HDL bind to one another, forming a ternary complex, wherein PON1 partially inhibits MPO activity, while MPO inactivates PON1. MPO oxidizes PON1 on tyrosine 71 (Tyr71), a modified residue found in human atheroma that is critical for HDL binding and PON1 function. Acute inflammation model studies with transgenic and knockout mice for either PON1 or MPO confirmed that MPO and PON1 reciprocally modulate each other’s function in vivo. Further structure and function studies identified critical contact sites between APOA1 within HDL, PON1, and MPO, and proteomics studies of HDL recovered from acute coronary syndrome (ACS) subjects revealed enhanced chlorotyrosine content, site-specific PON1 methionine oxidation, and reduced PON1 activity. HDL thus serves as a scaffold upon which MPO and PON1 interact during inflammation, whereupon PON1 binding partially inhibits MPO activity, and MPO promotes site-specific oxidative modification and impairment of PON1 and APOA1 function. PMID:23908111

  9. Myeloperoxidase-catalyzed oxidation of chloroacetonitrile to cyanide.

    PubMed

    Abdel-Naim, Ashraf B; Mohamadin, Ahmed M

    2004-02-02

    Chloroacetonitrile (CAN) is a disinfection by-product of chlorination of drinking water. Epidemiological studies indicate that it might present a potential hazard to human health. The present work provides an evidence for CAN activation to cyanide (CN-) by myeloperoxidase (MPO)/hydrogen peroxide (H2O2)/chloride (Cl-) system in vitro. Optimum conditions for the oxidation of CAN to CN- were characterized with respect to pH, temperature and time of incubation as well as CAN, MPO, H2O2 and KCl concentrations in incubation mixtures. The kinetic parameters governing the reaction; maximum velocity (Vmax) and Michaelis-Menten constant (Km) were assessed. Oxidation of CAN to CN- by NaOCl alone was shown. Addition of the MPO inhibitors; sodium azide (NaN3), 4-amino benzoic acid hydrazine (ABAH) or indomethacin to the reaction mixtures resulted in a significant decrease in the rate of CAN oxidation. Inclusion of the antioxidant enzyme catalase (CAT) in the incubation mixtures resulted in a significant decrease in the rate of CAN oxidation and CN- formation. Addition of the sulfhydryl compounds; glutathione (GSH), N-acetyl-L-cysteine (NAC), L-cysteine or D-penicillamine significantly enhanced the rate of CN- release. In conclusion, MPO/H2O2/Cl- system has the ability of oxidizing CAN to CN-. The present results represent a novel pathway for CAN activation and might be important in explaining CAN-induced toxicity.

  10. Thiocyanate and chloride as competing substrates for myeloperoxidase.

    PubMed Central

    van Dalen, C J; Whitehouse, M W; Winterbourn, C C; Kettle, A J

    1997-01-01

    The neutrophil enzyme myeloperoxidase uses H2O2 to oxidize chloride, bromide, iodide and thiocyanate to their respective hypohalous acids. Chloride is considered to be the physiological substrate. However, a detailed kinetic study of its substrate preference has not been undertaken. Our aim was to establish whether myeloperoxidase oxidizes thiocyanate in the presence of chloride at physiological concentrations of these substrates. We determined this by measuring the rate of H2O2 loss in reactions catalysed by the enzyme at various concentrations of each substrate. The relative specificity constants for chloride, bromide and thiocyanate were 1:60:730 respectively, indicating that thiocyanate is by far the most favoured substrate for myeloperoxidase. In the presence of 100 mM chloride, myeloperoxidase catalysed the production of hypothiocyanite at concentrations of thiocyanate as low as 25 microM. With 100 microM thiocyanate, about 50% of the H2O2 present was converted into hypothiocyanite, and the rate of hypohalous acid production equalled the sum of the individual rates obtained when each of these anions was present alone. The rate of H2O2 loss catalysed by myeloperoxidase in the presence of 100 mM chloride doubled when 100 microM thiocyanate was added, and was maximal with 1mM thiocyanate. This indicates that at plasma concentrations of thiocyanate and chloride, myeloperoxidase is far from saturated. We conclude that thiocyanate is a major physiological substrate of myeloperoxidase, regardless of where the enzyme acts. As a consequence, more consideration should be given to the oxidation products of thiocyanate and to the role they play in host defence and inflammation. PMID:9359420

  11. Variation in sorbitol accumulation and polyol-pathway activity in cultured human proximal tubule cells.

    PubMed

    Flath, M C; Bylander, J E; Sens, D A

    1992-09-01

    The polyol pathway is present in tissues of several organs where its activation may participate in the development of diabetic complications. We measured the accumulation of polyol-pathway intermediates in HPT cells isolated from 21 different human kidneys from nondiabetic individuals. When exposed to 27.5 mM glucose in the growth media, cells isolated from approximately 75% of individuals (accumulators) accumulated sorbitol within 1-4 days, whereas 25% (nonaccumulators) accumulated only negligible amounts, even when the period of exposure was extended to 2 wk. Surprisingly, measurement of the activities of the polyol-pathway enzymes showed no difference in the levels of either AR or SDH between accumulators and nonaccumulators, even when the conversion of galactose to galactitol was used to measure AR activity in intact cells independently of SDH. Measurement of sorbitol in the growth media indicated that nonaccumulators were not releasing sorbitol into the growth media. Fructose levels in the conditioned growth media were 4 times higher in the sorbitol-accumulating cells. Together, these results indicate that the tendency of cells from an individual to accumulate significant amounts of sorbitol may reflect the cells' ability to metabolize sorbitol in steps subsequent to the polyol pathway.

  12. Payload drug vs. nanocarrier biodegradation by myeloperoxidase- and peroxynitrite-mediated oxidations: pharmacokinetic implications

    NASA Astrophysics Data System (ADS)

    Seo, Wanji; Kapralov, Alexandr A.; Shurin, Galina V.; Shurin, Michael R.; Kagan, Valerian E.; Star, Alexander

    2015-05-01

    With the advancement of nanocarriers for drug delivery into biomedical practice, assessments of drug susceptibility to oxidative degradation by enzymatic mechanisms of inflammatory cells become important. Here, we investigate oxidative degradation of a carbon nanotube-based drug carrier loaded with Doxorubicin. We employed myeloperoxidase-catalysed and peroxynitrite-mediated oxidative conditions to mimic the respiratory burst of neutrophils and macrophages, respectively. In addition, we revealed that the cytostatic and cytotoxic effects of free Doxorubicin, but not nanotube-carried drug, on melanoma and lung carcinoma cell lines were abolished in the presence of tumor-activated myeloid regulatory cells that create unique myeloperoxidase- and peroxynitrite-induced oxidative conditions. Both ex vivo and in vitro studies demonstrate that the nanocarrier protects the drug against oxidative biodegradation.With the advancement of nanocarriers for drug delivery into biomedical practice, assessments of drug susceptibility to oxidative degradation by enzymatic mechanisms of inflammatory cells become important. Here, we investigate oxidative degradation of a carbon nanotube-based drug carrier loaded with Doxorubicin. We employed myeloperoxidase-catalysed and peroxynitrite-mediated oxidative conditions to mimic the respiratory burst of neutrophils and macrophages, respectively. In addition, we revealed that the cytostatic and cytotoxic effects of free Doxorubicin, but not nanotube-carried drug, on melanoma and lung carcinoma cell lines were abolished in the presence of tumor-activated myeloid regulatory cells that create unique myeloperoxidase- and peroxynitrite-induced oxidative conditions. Both ex vivo and in vitro studies demonstrate that the nanocarrier protects the drug against oxidative biodegradation. Electronic supplementary information (ESI) available: Experimental details and data from characterization of materials synthesis and degradation studies. See DOI: 10

  13. Gold enrichment in active geothermal systems by accumulating colloidal suspensions

    NASA Astrophysics Data System (ADS)

    Hannington, Mark; Harðardóttir, Vigdis; Garbe-Schönberg, Dieter; Brown, Kevin L.

    2016-04-01

    The origins of high-grade hydrothermal ore deposits are debated, but active geothermal systems provide important clues to their formation. The highest concentrations of gold are found in geothermal systems with direct links to island arc magmatism. Yet, similar concentrations have also been found in the absence of any input from arc magmas, for example, in the Reykjanes geothermal field, Iceland. Here we analyse brine samples taken from deep wells at Reykjanes and find that gold concentrations in the reservoir zone have increased over the past seven years from an average of 3 ppb to 14 ppb. The metal concentrations greatly exceed the maximum solubility of gold in the reservoir under saturated conditions and are now nearly two orders of magnitude higher than in mid-ocean ridge black smoker fluids--the direct analogues of Reykjanes deep liquids. We suggest that ongoing extraction of brine, the resulting pressure drop, and increased boiling have caused gold to drop out of solution and become trapped in the reservoir as a colloidal suspension. This process may explain how the stock of metal in the reservoirs of fossil geothermal systems could have increased over time and thus become available for the formation of gold-rich ore deposits.

  14. Sequence analysis, characterization and tissue distribution of channel catfish (Ictalurus punctatus Rafinesque, 1818) myeloperoxidase cDNA

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Myeloperoxidase (EC 1.11.1.7), a heme-containing lysosomal glycoprotein, is found predominantly in azurophilic granules of neutrophils. This enzyme upon activation catalyzes hydrogen peroxide in the presence of various halide ions to form hypohalous acids. Subsequently, these reagents are able to ...

  15. Reaction of myeloperoxidase compound I with chloride, bromide, iodide, and thiocyanate.

    PubMed

    Furtmüller, P G; Burner, U; Obinger, C

    1998-12-22

    Myeloperoxidase plays a fundamental role in oxidant production by neutrophils. The enzyme uses hydrogen peroxide to oxidize chloride (Cl-), bromide (Br-), iodide (I-), and the pseudohalide thiocyanate (SCN-) to their respective hypohalous acids. This study for the first time presents transient kinetic measurements of the oxidation of these halides and thiocyanate by the myeloperoxidase intermediate compound I, using the sequential mixing stopped-flow technique. At pH 7 and 15 degrees C, the two-electron reduction of compound I to the native enzyme by Cl- has a second-order rate constant of (2.5 +/- 0.3) x 10(4) M(-1) s(-1), whereas reduction of compound I by SCN- has a second-order rate constant of (9.6 +/- 0.5) x 10(6) M(-1) s(-1). Iodide [(7.2 +/- 0.7) x 10(6) M(-1) s(-1)] is shown to be a better electron donor for compound I than Br- [(1.1 +/- 0.1) x 10(6) M(-1) s(-1)]. The pH dependence studies suggest that compound I reduction by (pseudo-)halides is controlled by a residue with a pKa of about 4.6. The protonation of this group is necessary for optimum (pseudo-)halide anion oxidation. These transient kinetic results are underlined by steady-state spectral and kinetic investigations. SCN- is shown to be most effective in shifting the system myeloperoxidase/hydrogen peroxide from the peroxidatic cycle to the halogenation cycle, whereas iodide is shown to be more effective than bromide which in turn is much more effective than chloride. Decreasing pH increases the rate of this transition. Our results show that thiocyanate is an important substrate of myeloperoxidase in most environments and that hypothiocyanate is likely to contribute to leukocyte antimicrobial activity.

  16. The molecular mechanism for interaction of ceruloplasmin and myeloperoxidase

    NASA Astrophysics Data System (ADS)

    Bakhautdin, Bakytzhan; Bakhautdin, Esen Göksöy

    2016-04-01

    Ceruloplasmin (Cp) is a copper-containing ferroxidase with potent antioxidant activity. Cp is expressed by hepatocytes and activated macrophages and has been known as physiologic inhibitor of myeloperoxidase (MPO). Enzymatic activity of MPO produces anti-microbial agents and strong prooxidants such as hypochlorous acid and has a potential to damage host tissue at the sites of inflammation and infection. Thus Cp-MPO interaction and inhibition of MPO has previously been suggested as an important control mechanism of excessive MPO activity. Our aim in this study was to identify minimal Cp domain or peptide that interacts with MPO. We first confirmed Cp-MPO interaction by ELISA and surface plasmon resonance (SPR). SPR analysis of the interaction yielded 30 nM affinity between Cp and MPO. We then designed and synthesized 87 overlapping peptides spanning the entire amino acid sequence of Cp. Each of the peptides was tested whether it binds to MPO by direct binding ELISA. Two of the 87 peptides, P18 and P76 strongly interacted with MPO. Amino acid sequence analysis of identified peptides revealed high sequence and structural homology between them. Further structural analysis of Cp's crystal structure by PyMOL software unfolded that both peptides represent surface-exposed sites of Cp and face nearly the same direction. To confirm our finding we raised anti-P18 antisera in rabbit and demonstrated that this antisera disrupts Cp-MPO binding and rescues MPO activity. Collectively, our results confirm Cp-MPO interaction and identify two nearly identical sites on Cp that specifically bind MPO. We propose that inhibition of MPO by Cp requires two nearly identical sites on Cp to bind homodimeric MPO simultaneously and at an angle of at least 120 degrees, which, in turn, exerts tension on MPO and results in conformational change.

  17. Evaluation of Antiradical and Anti-Inflammatory Activities of Ethyl Acetate and Butanolic Subfractions of Agelanthus dodoneifolius (DC.) Polhill & Wiens (Loranthaceae) Using Equine Myeloperoxidase and Both PMA-Activated Neutrophils and HL-60 Cells

    PubMed Central

    Boly, Rainatou; Franck, Thierry; Kohnen, Stephan; Lompo, Marius; Guissou, Innocent Pierre; Dubois, Jacques; Serteyn, Didier; Mouithys-Mickalad, Ange

    2015-01-01

    The ethyl acetate and n-butanolic subfractions of Agelanthus dodoneifolius were investigated for their antioxidant and antimyeloperoxidase (MPO) activities. The reactive oxygen species (ROS) generation was assessed by lucigenin-enhanced chemiluminescence (CL) and dichlorofluorescein- (DCF-) induced fluorescence techniques from phorbol myristate acetate- (PMA-) stimulated equine neutrophils and human myeloid cell line HL-60, respectively. In parallel, the effects of the tested subfractions were evaluated on the total MPO release by stimulated neutrophils and on the specific MPO activity by means of immunological assays. The results showed the potent activity of the butanolic subfraction, at least in respect of the chemiluminescence test (IC50 = 0.3 ± 0.1 µg/mL) and the ELISA and SIEFED assays (IC50 = 2.8 ± 1.2 µg/mL and 1.3 ± 1.0 µg/mL), respectively. However, the ethyl acetate subfraction was found to be the most potent in the DCF assay as at the highest concentration, DCF fluorescence intensity decreases of about 50%. Moreover, we demonstrated that the ethyl acetate subfraction was rich in catechin (16.51%) while it was not easy to identify the main compounds in the butanolic subfraction using the UPLC-MS/MS technique. Nevertheless, taken together, our results provide evidence that Agelanthus dodoneifolius subfractions may represent potential sources of natural antioxidants and of antimyeloperoxidase compounds. PMID:25821497

  18. Melatonin Prevents Myeloperoxidase Heme Destruction and the Generation of Free Iron Mediated by Self-Generated Hypochlorous Acid

    PubMed Central

    Shaeib, Faten; Khan, Sana N.; Ali, Iyad; Najafi, Tohid; Maitra, Dhiman; Abdulhamid, Ibrahim; Saed, Ghassan M.; Pennathur, Subramaniam; Abu-Soud, Husam M.

    2015-01-01

    Myeloperoxidase (MPO) generated hypochlorous acid (HOCl) formed during catalysis is able to destroy the MPO heme moiety through a feedback mechanism, resulting in the accumulation of free iron. Here we show that the presence of melatonin (MLT) can prevent HOCl-mediated MPO heme destruction using a combination of UV-visible photometry, hydrogen peroxide (H2O2)-specific electrode, and ferrozine assay techniques. High performance liquid chromatography (HPLC) analysis showed that MPO heme protection was at the expense of MLT oxidation. The full protection of the MPO heme requires the presence of a 1:2 MLT to H2O2 ratio. Melatonin prevents HOCl–mediated MPO heme destruction through multiple pathways. These include competition with chloride, the natural co-substrate; switching the MPO activity from a two electron oxidation to a one electron pathway causing the buildup of the inactive Compound II, and its subsequent decay to MPO-Fe(III) instead of generating HOCl; binding to MPO above the heme iron, thereby preventing the access of H2O2 to the catalytic site of the enzyme; and direct scavenging of HOCl. Collectively, in addition to acting as an antioxidant and MPO inhibitor, MLT can exert its protective effect by preventing the release of free iron mediated by self-generated HOCl. Our work may establish a direct mechanistic link by which MLT exerts its antioxidant protective effect in chronic inflammatory diseases with MPO elevation. PMID:25835505

  19. Hippocampal activation is associated with longitudinal amyloid accumulation and cognitive decline.

    PubMed

    Leal, Stephanie L; Landau, Susan M; Bell, Rachel K; Jagust, William J

    2017-02-08

    The amyloid hypothesis suggests that beta-amyloid (Aβ) deposition leads to alterations in neural function and ultimately to cognitive decline in Alzheimer's disease. However, factors that underlie Aβ deposition are incompletely understood. One proposed model suggests that synaptic activity leads to increased Aβ deposition. More specifically, hyperactivity in the hippocampus may be detrimental and could be one factor that drives Aβ deposition. To test this model, we examined the relationship between hippocampal activity during a memory task using fMRI and subsequent longitudinal change in Aβ using PIB-PET imaging in cognitively normal older adults. We found that greater hippocampal activation at baseline was associated with increased Aβ accumulation. Furthermore, increasing Aβ accumulation mediated the influence of hippocampal activation on declining memory performance, demonstrating a crucial role of Aβ in linking hippocampal activation and memory. These findings support a model linking increased hippocampal activation to subsequent Aβ deposition and cognitive decline.

  20. Inhibition of Myeloperoxidase- and Neutrophil-Mediated Hypochlorous Acid Formation in Vitro and Endothelial Cell Injury by (-)-Epigallocatechin Gallate.

    PubMed

    Tian, Rong; Ding, Yun; Peng, Yi-Yuan; Lu, Naihao

    2017-04-10

    Myeloperoxidase (MPO) plays important roles in various diseases through its unique chlorinating activity to catalyze excess hypochlorous acid (HOCl) formation. Epidemiological studies indicate an inverse correlation between plant polyphenol consumption and the incidence of cardiovascular diseases. Here we showed that (-)-epigallocatechin gallate (EGCG), the main flavonoid present in green tea, dose-dependently inhibited MPO-mediated HOCl formation in vitro (chlorinating activities of MPO: 50.2 ± 5.7% for 20 μM EGCG versus 100 ± 5.6% for control, P < 0.01). UV-vis spectral and docking studies indicated that EGCG bound to the active site (heme) of MPO and resulted in the accumulation of compound II, which was unable to produce HOCl. This flavonoid also effectively inhibited HOCl generation in activated neutrophils (HOCl formation: 65.0 ± 5.6% for 20 μM EGCG versus 100 ± 6.2% for control, P < 0.01) without influencing MPO and Nox2 release and superoxide formation, suggesting that EGCG specifically inhibited MPO but not NADPH oxidase activity in activated neutrophils. Moreover, EGCG inhibited MPO (or neutrophil)-mediated HOCl formation in human umbilical vein endothelial cells (HUVEC) culture and accordingly protected HUVEC from MPO (or neutrophil)-induced injury (P < 0.05, all cases), although it did not induce cytotoxicity to HUVEC (P > 0.05, all cases). Our results indicate that dietary EGCG is an effective and specific inhibitor of MPO activity and may participate in the regulation of immune responses at inflammatory sites.

  1. The effect of nitrite on aerobic phosphate uptake and denitrifying activity of phosphate-accumulating organisms.

    PubMed

    Yoshida, Y; Takahashi, K; Saito, T; Tanaka, K

    2006-01-01

    An anaerobic/aerobic/anoxic/aerobic sequencing batch reactor (SBR) was operated with municipal wastewater to investigate the effect of nitrite on biological phosphorus removal (BPR). When nitrite accumulated, aerobic phosphate uptake activity significantly decreased and, in case of hard exposure to nitrite, BPR severely deteriorated. The interesting observation was that the relative anoxic activity of phosphate accumulating organisms (PAOs) increased after nitrite exposure. Moreover batch tests of aerobic phosphate uptake in the presence/absence of nitrite indicated that PAOs with the higher relative anoxic activity are less sensitive to nitrite exposure. From these results, we concluded that BPR is sensitive to nitrite exposure, but BPR containing PAOs with the higher relative anoxic activity is possibly more stable against nitrite than BPR containing PAOs with the lower relative anoxic activity.

  2. Relationships Between Photosynthetic Activity and Silica Accumulation with Ages of Leaf in Sasa veitchii (Poaceae, Bambusoideae)

    PubMed Central

    Motomura, Hiroyuki; Hikosaka, Kouki; Suzuki, Mitsuo

    2008-01-01

    Background and Aims Bamboos have long-lived, evergreen leaves that continue to accumulate silica throughout their life. Silica accumulation has been suggested to suppress their photosynthetic activity. However, nitrogen content per unit leaf area (Narea), an important determinant of maximum photosynthetic capacity per unit leaf area (Pmax), decreases as leaves age and senescence. In many species, Pmax decreases in parallel with the leaf nitrogen content. It is hypothesized that if silica accumulation affects photosynthesis, then Pmax would decrease faster than Narea, leading to a decrease in photosynthetic rate per unit leaf nitrogen (photosynthetic nitrogen use efficiency, PNUE) with increasing silica content in leaves. Methods The hypothesis was tested in leaves of Sasa veitchii, which have a life span of 2 years and accumulate silica up to 41 % of dry mass. Seasonal changes in Pmax, stomatal conductance, Narea and silica content were measured for leaves of different ages. Key Results Although Pmax and PNUE were negatively related with silica content across leaves of different ages, the relationship between PNUE and silica differed depending on leaf age. In second-year leaves, PNUE was almost constant although there was a large increase in silica content, suggesting that leaf nitrogen was a primary factor determining the variation in Pmax and that silica accumulation did not affect photosynthesis. PNUE was strongly and negatively correlated with silica content in third-year leaves, suggesting that silica accumulation affected photosynthesis of older leaves. Conclusions Silica accumulation in long-lived leaves of bamboo did not affect photosynthesis when the silica concentration of a leaf was less than 25 % of dry mass. Silica may be actively transported to epidermal cells rather than chlorenchyma cells, avoiding inhibition of CO2 diffusion from the intercellular space to chloroplasts. However, in older leaves with a larger silica content, silica was also

  3. ANTIMICROBIAL ACTIVITY OF COPPER AND ZINC ACCUMULATED BY EASTERN OYSTER AMEBOCYTES

    EPA Science Inventory

    Fisher, William S. Submitted. Antimicrobial Activity of Copper and Zinc Accumulated by Eastern Oyster Amebocytes. J. Shellfish Res. 54 p. (ERL,GB 1196).

    The distribution of eastern oysters Crassostrea virginica near terrestrial watersheds has led to a general impression t...

  4. Impact on growth and aflatoxin B1 accumulation by Kluyveromyces isolates at different water activity conditions.

    PubMed

    Penna, Mariángeles La; Etcheverry, Miriam

    2006-11-01

    This study showed the impact on germination, mycelial growth and aflatoxin B(1) accumulation when interacting Aspergillus aflatoxigenic strains with Kluyveromyces isolates and the effect of water activity on this relationship. Isolates Y(14) and Y(16) reduced the percentage of germination of all Aspergillus strains and decrease germ tube elongation rate at majority of water activity assayed. Similarly they produced an increase of germination lag phase and lag phase of growth beside decreased growth rate of all Aspergillus strains. At water activities 0.994, 0.982, 0.955 and 0.937, no aflatoxins were produced in paired cultures with isolates Y(25,) Y(22), Y(16), and Y(14), and Kluyveromyces isolates Y(14) and Y(16) impact both growth and aflatoxin accumulation at wide range of water activity.

  5. SMA Selectively Codes the Active Accumulation of Temporal, Not Spatial, Magnitude.

    PubMed

    Coull, Jennifer T; Charras, Pom; Donadieu, Maxime; Droit-Volet, Sylvie; Vidal, Franck

    2015-11-01

    Estimating duration depends on the sequential integration (accumulation) of temporal information in working memory. Using fMRI, we directly compared the accumulation of information in temporal versus spatial domains. Participants estimated either the duration or distance of the dynamic trajectory of a moving dot or, in a control condition, a static line stimulus. Comparing the duration versus distance of static lines activated an extensive cortico-striatal network. By contrast, comparing the duration versus distance of dynamic trajectories, both of which required sequential integration of information, activated SMA alone. Indeed, activity in SMA, as well as right inferior occipital cortex, increased parametrically as a function of stimulus duration and also correlated with individual differences in the propensity to overestimate stimulus duration. By contrast, activity in primary visual cortex increased parametrically as a function of stimulus distance. Crucially, a direct comparison of the parametric responses to duration versus distance revealed that activity in SMA increased incrementally as a function of stimulus duration but not as a function of stimulus distance. Collectively, our results indicate that SMA responds to the active accumulation of information selectively in the temporal domain.

  6. Luminol-dependent photoemission from single neutrophil stimulated by phorbol ester and calcium ionophore--role of degranulation and myeloperoxidase

    SciTech Connect

    Suematsu, M.; Oshio, C.; Miura, S.; Suzuki, M.; Houzawa, S.; Tsuchiya, M.

    1988-08-30

    Luminol-dependent photonic burst from phorbol ester-treated single neutrophil was visually investigated by using an ultrasensitive photonic image intensifier microscope. Neutrophils stimulated by phorbol myristate acetate (0.1 microgram/ml) alone produced a negligible level of photonic activities in the presence of luminol (10 micrograms/ml). The additional application of 0.1 microM Ca2+ ionophore A23187 induced explosive changes of photonic burst corresponding to the distribution of neutrophils, and these photonic activities were gradually spread to extracellular space. Sodium azide, which prevents myeloperoxidase activity, inhibited Ca2+ ionophore-induced photonic burst from phorbol ester-treated neutrophil. These findings suggest a prerequisite role of degranulation and myeloperoxidase release in luminol-dependent photoemission from stimulated neutrophils.

  7. Inhibition of the alpha-ketoglutarate dehydrogenase complex by the myeloperoxidase products, hypochlorous acid and mono-N-chloramine.

    PubMed

    Jeitner, Thomas M; Xu, Hui; Gibson, Gary E

    2005-01-01

    Abstract alpha-Ketoglutarate dehydrogenase (KGDHC) complex activity is diminished in a number of neurodegenerative disorders and its diminution in Alzheimer Disease (AD) is thought to contribute to the major loss of cerebral energy metabolism that accompanies this disease. The loss of KGDHC activity appears to be predominantly due to post-translation modifications. Thiamine deficiency also results in decreased KGDHC activity and a selective neuronal loss. Recently, myeloperoxidase has been identified in the activated microglia of brains from AD patients and thiamine-deficient animals. Myeloperoxidase produces a powerful oxidant, hypochlorous acid that reacts with amines to form chloramines. The aim of this study was to investigate the ability of hypochlorous acid and chloramines to inhibit the activity of KGDHC activity as a first step towards investigating the role of myeloperoxidase in AD. Hypochlorous acid and mono-N-chloramine both inhibited purified and cellular KGDHC and the order of inhibition of the purified complex was hypochlorous acid (1x) > mono-N-chloramine (approximately 50x) > hydrogen peroxide (approximately 1,500). The inhibition of cellular KGDHC occurred with no significant loss of cellular viability at all exposure times that were examined. Thus, hypochlorous acid and chloramines have the potential to inactivate a major target in neurodegeneration.

  8. A Clickable Analogue of Ketamine Retains NMDA Receptor Activity, Psychoactivity, and Accumulates in Neurons

    PubMed Central

    Emnett, Christine; Li, Hairong; Jiang, Xiaoping; Benz, Ann; Boggiano, Joseph; Conyers, Sara; Wozniak, David F.; Zorumski, Charles F.; Reichert, David E.; Mennerick, Steven

    2016-01-01

    Ketamine is a psychotomimetic and antidepressant drug. Although antagonism of cell-surface NMDA receptors (NMDARs) may trigger ketamine’s psychoactive effects, ketamine or its major metabolite norketamine could act intracellularly to produce some behavioral effects. To explore the viability of this latter hypothesis, we examined intracellular accumulation of novel visualizable analogues of ketamine/norketamine. We introduced an alkyne “click” handle into norketamine (alkyne-norketamine, A-NK) at the key nitrogen atom. Ketamine, norketamine, and A-NK, but not A-NK-amide, showed acute and persisting psychoactive effects in mice. This psychoactivity profile paralleled activity of the compounds as NMDAR channel blockers; A-NK-amide was inactive at NMDARs, and norketamine and A-NK were active but ~4-fold less potent than ketamine. We incubated rat hippocampal cells with 10 μM A-NK or A-NK-amide then performed Cu2+ catalyzed cycloaddition of azide-Alexa Fluor 488, which covalently attaches the fluorophore to the alkyne moiety in the compounds. Fluorescent imaging revealed intracellular localization of A-NK but weak A-NK-amide labeling. Accumulation was not dependent on membrane potential, NMDAR expression, or NMDAR activity. Overall, the approach revealed a correlation among NMDAR activity, intracellular accumulation/retention, and behavioral effects. Thus, we advance first generation chemical biology tools to aid in the identification of ketamine targets. PMID:27982047

  9. Accumulated Bending Energy Elicits Neutral Sphingomyelinase Activity in Human Red Blood Cells

    PubMed Central

    López, David J.; Egido-Gabas, Meritxell; López-Montero, Iván; Busto, Jon V.; Casas, Josefina; Garnier, Marie; Monroy, Francisco; Larijani, Banafshé; Goñi, Félix M.; Alonso, Alicia

    2012-01-01

    We propose that accumulated membrane bending energy elicits a neutral sphingomyelinase (SMase) activity in human erythrocytes. Membrane bending was achieved by osmotic or chemical processes, and SMase activity was assessed by quantitative thin-layer chromatography, high-performance liquid chromatography, and electrospray ionization-mass spectrometry. The activity induced by hypotonic stress in erythrocyte membranes had the pH dependence, ion dependence, and inhibitor sensitivity of mammalian neutral SMases. The activity caused a decrease in SM contents, with a minimum at 6 min after onset of the hypotonic conditions, and then the SM contents were recovered. We also elicited SMase activity by adding lysophosphatidylcholine externally or by generating it with phospholipase A2. The same effect was observed upon addition of chlorpromazine or sodium deoxycholate at concentrations below the critical micellar concentration, and even under hypertonic conditions. A unifying factor of the various agents that elicit this SMase activity is the accumulated membrane bending energy. Both hypo-and hypertonic conditions impose an increased curvature, whereas the addition of surfactants or phospholipase A2 activation increases the outer monolayer area, thus leading to an increased bending energy. The fact that this latent SMase activity is tightly coupled to the membrane bending properties suggests that it may be related to the general phenomenon of stress-induced ceramide synthesis and apoptosis. PMID:22824271

  10. Accumulated bending energy elicits neutral sphingomyelinase activity in human red blood cells.

    PubMed

    López, David J; Egido-Gabas, Meritxell; López-Montero, Iván; Busto, Jon V; Casas, Josefina; Garnier, Marie; Monroy, Francisco; Larijani, Banafshé; Goñi, Félix M; Alonso, Alicia

    2012-05-02

    We propose that accumulated membrane bending energy elicits a neutral sphingomyelinase (SMase) activity in human erythrocytes. Membrane bending was achieved by osmotic or chemical processes, and SMase activity was assessed by quantitative thin-layer chromatography, high-performance liquid chromatography, and electrospray ionization-mass spectrometry. The activity induced by hypotonic stress in erythrocyte membranes had the pH dependence, ion dependence, and inhibitor sensitivity of mammalian neutral SMases. The activity caused a decrease in SM contents, with a minimum at 6 min after onset of the hypotonic conditions, and then the SM contents were recovered. We also elicited SMase activity by adding lysophosphatidylcholine externally or by generating it with phospholipase A(2). The same effect was observed upon addition of chlorpromazine or sodium deoxycholate at concentrations below the critical micellar concentration, and even under hypertonic conditions. A unifying factor of the various agents that elicit this SMase activity is the accumulated membrane bending energy. Both hypo-and hypertonic conditions impose an increased curvature, whereas the addition of surfactants or phospholipase A(2) activation increases the outer monolayer area, thus leading to an increased bending energy. The fact that this latent SMase activity is tightly coupled to the membrane bending properties suggests that it may be related to the general phenomenon of stress-induced ceramide synthesis and apoptosis.

  11. Myeloperoxidase-dependent lipid peroxidation promotes the oxidative modification of cytosolic proteins in phagocytic neutrophils.

    PubMed

    Wilkie-Grantham, Rachel P; Magon, Nicholas J; Harwood, D Tim; Kettle, Anthony J; Vissers, Margreet C; Winterbourn, Christine C; Hampton, Mark B

    2015-04-10

    Phagocytic neutrophils generate reactive oxygen species to kill microbes. Oxidant generation occurs within an intracellular phagosome, but diffusible species can react with the neutrophil and surrounding tissue. To investigate the extent of oxidative modification, we assessed the carbonylation of cytosolic proteins in phagocytic neutrophils. A 4-fold increase in protein carbonylation was measured within 15 min of initiating phagocytosis. Carbonylation was dependent on NADPH oxidase and myeloperoxidase activity and was inhibited by butylated hydroxytoluene and Trolox, indicating a role for myeloperoxidase-dependent lipid peroxidation. Proteomic analysis of target proteins revealed significant carbonylation of the S100A9 subunit of calprotectin, a truncated form of Hsp70, actin, and hemoglobin from contaminating erythrocytes. The addition of the reactive aldehyde 4-hydroxynonenal (HNE) caused carbonylation, and HNE-glutathione adducts were detected in the cytosol of phagocytic neutrophils. The post-translational modification of neutrophil proteins will influence the functioning and fate of these immune cells in the period following phagocytic activation, and provides a marker of neutrophil activation during infection and inflammation.

  12. Myeloperoxidase-dependent Lipid Peroxidation Promotes the Oxidative Modification of Cytosolic Proteins in Phagocytic Neutrophils*

    PubMed Central

    Wilkie-Grantham, Rachel P.; Magon, Nicholas J.; Harwood, D. Tim; Kettle, Anthony J.; Vissers, Margreet C.; Winterbourn, Christine C.; Hampton, Mark B.

    2015-01-01

    Phagocytic neutrophils generate reactive oxygen species to kill microbes. Oxidant generation occurs within an intracellular phagosome, but diffusible species can react with the neutrophil and surrounding tissue. To investigate the extent of oxidative modification, we assessed the carbonylation of cytosolic proteins in phagocytic neutrophils. A 4-fold increase in protein carbonylation was measured within 15 min of initiating phagocytosis. Carbonylation was dependent on NADPH oxidase and myeloperoxidase activity and was inhibited by butylated hydroxytoluene and Trolox, indicating a role for myeloperoxidase-dependent lipid peroxidation. Proteomic analysis of target proteins revealed significant carbonylation of the S100A9 subunit of calprotectin, a truncated form of Hsp70, actin, and hemoglobin from contaminating erythrocytes. The addition of the reactive aldehyde 4-hydroxynonenal (HNE) caused carbonylation, and HNE-glutathione adducts were detected in the cytosol of phagocytic neutrophils. The post-translational modification of neutrophil proteins will influence the functioning and fate of these immune cells in the period following phagocytic activation, and provides a marker of neutrophil activation during infection and inflammation. PMID:25697357

  13. Looking for phosphate-accumulating bacteria in activated sludge processes: a multidisciplinary approach.

    PubMed

    Tarayre, Cédric; Charlier, Raphaëlle; Delepierre, Anissa; Brognaux, Alison; Bauwens, Julien; Francis, Frédéric; Dermience, Michaël; Lognay, Georges; Taminiau, Bernard; Daube, Georges; Compère, Philippe; Meers, Erik; Michels, Evi; Delvigne, Frank

    2017-01-29

    Over the past decades, an increasing need in renewable resources has progressively appeared. This trend concerns not only fossil fuels but also mineral resources. Wastewater and sewage sludge contain significant concentrations in phosphate and can be considered as a fertilizer source of the utmost importance. In wastewater treatment plants, the biological uptake of phosphate is performed by a specific microbiota: the phosphate-accumulating organisms. These microorganisms are recovered in sewage sludge. Here, we aimed to investigate the occurrence of phosphate accumulators in four wastewater treatment plants. A 16S metagenetic analysis identified the main bacterial phyla extracted from the aerobic treatment: α-Proteobacteria, β-Proteobacteria, and Sphingobacteria. An enrichment stage was performed to stimulate the specific growth of phosphate-accumulating bacteria in an acetate medium. An analysis of metabolic activities of sulfur and phosphorus highlighted strong modifications related to phosphorus and much less distinguishable effects with sulfur. A solid acetate medium containing 5-Br-4-Cl-3-indolyl phosphate was used to select potential phosphate-accumulating bacteria from the enriched consortia. The positive strains have been found to belong in the genera Acinetobacter, Corynebacterium, and Pseudomonas. Finally, electron microscopy was applied to the strains and allowed to confirm the presence of polyphosphate granules. Some of these bacteria contained granules the size of which exceeded 100 nm.

  14. Cellular Accumulation, Localization, and Activity of a Synthetic Cyclopeptamine in Fungi

    PubMed Central

    Capobianco, John O.; Zakula, Dorothy; Frost, David J.; Goldman, Robert C.; Li, Leping; Klein, Larry L.; Lartey, Paul A.

    1998-01-01

    A novel synthetic cyclopeptamine, A172013, rapidly accumulated by passive diffusion into Candida albicans CCH442. Drug influx could not be totally facilitated by the membrane-bound target, β-(1,3)-glucan synthase, since accumulation was unsaturable at drug concentrations up to 10 μg/ml (about 1.6 × 10−7 molecules/cell), or 25× MIC. About 55 and 23% of the cell-incorporated drug was associated with the cell wall and protoplasts, respectively. Isolated microsomes contained 95% of the protoplast-associated drug, which was fully active against glucan synthesis in vitro. Drug (0.1 μg/ml) accumulation was rapid and complete after 5 min in several fungi tested, including a lipopeptide/cyclopeptamine-resistant strain of C. albicans (LP3-1). The compound penetrated to comparable levels in both yeast and hyphal forms of C. albicans, and accumulation in Aspergillus niger was 20% that in C. albicans. These data indicated that drug-cell interactions were driven by the amphiphilic nature of the compound and that the cell wall served as a major drug reservoir. PMID:9527791

  15. Visual cortical activity reflects faster accumulation of information from cortically blind fields

    PubMed Central

    Martin, Tim; Das, Anasuya; Huxlin, Krystel R.

    2012-01-01

    Brain responses (from functional magnetic resonance imaging) and components of information processing were investigated in nine cortically blind observers performing a global direction discrimination task. Three of these subjects had responses in perilesional cortex in response to blind field stimulation, whereas the others did not. We used the EZ-diffusion model of decision making to understand how cortically blind subjects make a perceptual decision on stimuli presented within their blind field. We found that these subjects had slower accumulation of information in their blind fields as compared with their good fields and to intact controls. Within cortically blind subjects, activity in perilesional tissue, V3A and hMT+ was associated with a faster accumulation of information for deciding direction of motion of stimuli presented in the blind field. This result suggests that the rate of information accumulation is a critical factor in the degree of impairment in cortical blindness and varies greatly among affected individuals. Retraining paradigms that seek to restore visual functions might benefit from focusing on increasing the rate of information accumulation. PMID:23169923

  16. Hippocampal activation is associated with longitudinal amyloid accumulation and cognitive decline

    PubMed Central

    Leal, Stephanie L; Landau, Susan M; Bell, Rachel K; Jagust, William J

    2017-01-01

    The amyloid hypothesis suggests that beta-amyloid (Aβ) deposition leads to alterations in neural function and ultimately to cognitive decline in Alzheimer’s disease. However, factors that underlie Aβ deposition are incompletely understood. One proposed model suggests that synaptic activity leads to increased Aβ deposition. More specifically, hyperactivity in the hippocampus may be detrimental and could be one factor that drives Aβ deposition. To test this model, we examined the relationship between hippocampal activity during a memory task using fMRI and subsequent longitudinal change in Aβ using PIB-PET imaging in cognitively normal older adults. We found that greater hippocampal activation at baseline was associated with increased Aβ accumulation. Furthermore, increasing Aβ accumulation mediated the influence of hippocampal activation on declining memory performance, demonstrating a crucial role of Aβ in linking hippocampal activation and memory. These findings support a model linking increased hippocampal activation to subsequent Aβ deposition and cognitive decline. DOI: http://dx.doi.org/10.7554/eLife.22978.001 PMID:28177283

  17. Distinctive role of activated tumor-associated macrophages in photosensitizer accumulation

    NASA Astrophysics Data System (ADS)

    Korbelik, Mladen; Krosl, Gorazd

    1995-05-01

    Cells dissociated from tumors (carcinomas and sarcomas) growing subcutaneously in mice that have been administered Photofrin or other photosensitizers were analyzed by flow cytometry. Monoclonal antibodies were used for identification of major cellular populations contained in these tumors. The results demonstrate that a subpopulation of tumor-associated macrophages (TAMs) is unique among tumor cell populations in that it excels in the accumulation of very high levels of photosensitizers. These macrophages showed an increased expression of interleukin 2 receptor, which is indicative of their activated state. since macrophages were reported to concentrate in the periphery of human neoplasms, it is suggested that activates TAMs are the determinants of tumor-localized photosensitizer fluorescence.

  18. Effect of acetic acid on lipid accumulation by glucose-fed activated sludge cultures

    SciTech Connect

    Mondala, Andro; Hernandez, Rafael; French, Todd; McFarland, Linda; Sparks, Darrell; Holmes, William; Haque, Monica

    2012-01-01

    The effect of acetic acid, a lignocellulose hydrolysis by-product, on lipid accumulation by activated sludge cultures grown on glucose was investigated. This was done to assess the possible application of lignocellulose as low-cost and renewable fermentation substrates for biofuel feedstock production. Results: Biomass yield was reduced by around 54% at a 2 g L -1 acetic acid dosage but was increased by around 18% at 10 g L -1 acetic acid dosage relative to the control run. The final gravimetric lipid contents at 2 and 10 g L -1 acetic acid levels were 12.5 + 0.7% and 8.8 + 3.2% w/w, respectively, which were lower than the control (17.8 + 2.8% w/w). However, biodiesel yields from activated sludge grown with acetic acid (5.6 + 0.6% w/w for 2 g L -1 acetic acid and 4.2 + 3.0% w/w for 10 g L -1 acetic acid) were higher than in raw activated sludge (1-2% w/w). The fatty acid profiles of the accumulated lipids were similar with conventional plant oil biodiesel feedstocks. Conclusions: Acetic acid enhanced biomass production by activated sludge at high levels but reduced lipid production. Further studies are needed to enhance acetic acid utilization by activated sludge microorganisms for lipid biosynthesis.

  19. Plasma membrane overgrowth causes fibrotic collagen accumulation and immune activation in Drosophila adipocytes

    PubMed Central

    Zang, Yiran; Wan, Ming; Liu, Min; Ke, Hongmei; Ma, Shuangchun; Liu, Lu-Ping; Ni, Jian-Quan; Carlos Pastor-Pareja, José

    2015-01-01

    Many chronic diseases are associated with fibrotic deposition of Collagen and other matrix proteins. Little is known about the factors that determine preferential onset of fibrosis in particular tissues. Here we show that plasma membrane (PM) overgrowth causes pericellular Collagen accumulation in Drosophila adipocytes. We found that loss of Dynamin and other endocytic components causes pericellular trapping of outgoing Collagen IV due to dramatic cortex expansion when endocytic removal of PM is prevented. Deposits also form in the absence of negative Toll immune regulator Cactus, excess PM being caused in this case by increased secretion. Finally, we show that trimeric Collagen accumulation, downstream of Toll or endocytic defects, activates a tissue damage response. Our work indicates that traffic imbalances and PM topology may contribute to fibrosis. It also places fibrotic deposits both downstream and upstream of immune signaling, consistent with the chronic character of fibrotic diseases. DOI: http://dx.doi.org/10.7554/eLife.07187.001 PMID:26090908

  20. Regulation of Activation-associated MicroRNA Accumulation Rates during Monocyte-to-macrophage Differentiation*

    PubMed Central

    Eigsti, Renee L.; Sudan, Bayan; Wilson, Mary E.; Graff, Joel W.

    2014-01-01

    Circulating monocytes recruited to tissues can differentiate into macrophages and adopt unique gene expression programs in response to environmental cues. We recently described the regulated expression of several microRNAs (miRNAs) in polarized human monocyte-derived macrophages (MDMs). Basal expression of these activation-associated miRNAs was low in monocytes relative to MDMs. As development occurs in the context of specific cellular environments, we hypothesized that the rate of miRNA accumulation would be modified in the presence of microbial or cellular products during monocyte-to-macrophage differentiation. Indeed, LPS treatment augmented the accumulation of miR-146a and miR-155, whereas IL-4 treatment augmented the accumulation of miR-193b and miR-222 during development. In contrast, some stimuli repressed accumulation of specific miRNAs including interferons (IFNs) (miR-27a, miR-125a-5p, and miR-222), IL-4 (miR-125a-5p), and LPS (miR-27a). RT-PCR-based expression profiling of monocytes differentiated with distinct methods showed that activation-associated miRNAs and markers of macrophage polarization were substantially altered in MDMs differentiated in the presence of non-monocytic peripheral blood mononuclear cells due in part to NF-κB and STAT1 pathway activation. Expression of several of these miRNAs was regulated at a preprocessing step because the expression of the primary miRNAs, but not Dicer, correlated with mature miRNA expression. We conclude that a set of miRNAs is regulated during MDM differentiation, and the rate is uniquely modified for each miRNA by environmental factors. The low basal expression of activation-associated miRNAs in monocytes and their dynamic rates of accumulation during MDM differentiation permit monocytes to tailor miRNA profiles in peripheral tissues during differentiation to macrophages. PMID:25148686

  1. Proton accumulation and ATPase activity in Golgi apparatus-enriched vesicles from rat liver

    SciTech Connect

    Yeh, H.I.; van Rossum, G.D. )

    1991-03-01

    We have studied the mechanism by which liver Golgi apparatus maintains the acidity of its contents, using a subcellular fraction from rat liver highly enriched in Golgi marker enzymes. Proton accumulation (measured by quenching of acridine-orange fluorescence) and anion-dependent ATPase were characterized and compared. Maximal ATPase and proton accumulation required ATP; GTP and other nucleotides gave 10% to 30% of maximal activity. Among anions, Cl- and Br- approximately doubled the activities; others were much less effective. Half-maximal increase of ATPase and H+ uptake required 55 mmol/L and 27 mmol/L Cl-, respectively. In predominantly chloride media, SCN- and NO3- markedly inhibited H+ uptake. Nitrate competitively inhibited both the chloride-dependent ATPase (apparent Ki 6 mmol/L) and proton uptake (apparent Ki 2 mmol/L). Nitrate and SCN- also inhibited uptake of 36Cl. Replacing K+ with Na+ had no effect on the initial rate of proton uptake but somewhat reduced the steady state attained. Replacement of K+ with NH4+ and choline reduced proton uptake without affecting ATPase. The ATPase and H+ uptake were supported equally well by Mg2+ or Mn2+. The ATPase was competitively inhibited by 4-acetamido-4'-isothiocyano-stilbene-2,2'-disulfonic acid (apparent Ki 39 mumol/L). Other agents inhibiting both H+ uptake and ATPase were N-ethylmaleimide, N,N'-dicyclohexylcarbodiimide, chlorpromazine, diethylstilbestrol, Zn2+, Co2+ and Cu2+. In the Cl- medium, accumulated protons were released by ionophores at the relative rates, monensin = nigericin greater than valinomycin greater than carbonyl cyanide mchlorophenylhydrazone; the last of these also reduced ATPase activity. In the absence of Cl-, monensin and valinomycin both stimulated the ATPase. These results show a close association between ATPase activity and acidification of liver Golgi vesicles.

  2. Contribution of myeloperoxidase and inducible nitric oxide synthase to pathogenesis of psoriasis

    PubMed Central

    Dilek, Aziz Ramazan; Taşkın, Yakup; Erkinüresin, Taşkın; Yalçın, Ömer; Saral, Yunus

    2016-01-01

    Introduction Histological changes of psoriasis include invasion of neutrophils into the epidermis and formation of Munro abscesses in the epidermis. Neutrophils are the predominant white blood cells in circulation when stimulated; they discharge the abundant myeloperoxidase (MPO) enzyme that uses hydrogen peroxide to oxidize chloride for killing ingested bacteria. Aim To investigate the contribution of neutrophils to the pathogenesis of psoriasis at the blood and tissue levels through inducible nitric oxide synthase (iNOS) and MPO. Material and methods A total of 50 adult patients with a chronic plaque form of psoriasis and 25 healthy controls were enrolled to this study. Serum MPO and iNOS levels were measured using ELISA method. Two biopsy specimens were taken in each patient from the center of the lesion and uninvolved skin. Immunohistochemistry was performed for MPO and iNOS on both normal and psoriasis vulgaris biopsies. Results While a significant difference between serum myeloperoxidase levels were detected, a similar statistical difference between participants in the serum iNOS levels was not found. In immunohistochemistry, intensely stained leukocytes with MPO and intensely staining with iNOS in psoriatic skin was observed. Conclusions Neutrophils in psoriasis lesions are actively producing MPO and this indirectly triggers the synthesis of iNOS. Targeting of MPO or synthesis of MPO in the lesion area may contribute to development of a new treatment option. PMID:28035220

  3. UV-B-Induced PR-1 Accumulation Is Mediated by Active Oxygen Species.

    PubMed

    Green, R.; Fluhr, R.

    1995-02-01

    Depletion of the stratospheric ozone layer may result in an increase in the levels of potentially harmful UV-B radiation reaching the surface of the earth. We have found that UV-B is a potent inducer of the plant pathogenesis-related protein PR-1 in tobacco leaves. UV-B fluences required for PR-1 accumulation are similar to those of other UV-B-induced responses. The UV-B-induced PR-1 accumulation was confined precisely to the irradiated area of the leaf but displayed no leaf tissue specificity. A study of some of the possible components of the signal transduction pathway between UV-B and PR-1 induction showed that photosynthetic processes are not essential, and photoreversible DNA damage is not involved. Antioxidants and cycloheximide were able to block the induction of PR-1 by UV-B, and treatment of leaves with a generator of reactive oxygen resulted in the accumulation of PR-1 protein. These results demonstrate an absolute requirement for active oxygen species and protein synthesis in this UV-B signal transduction pathway. In contrast, we also show that other elicitors, notably salicylic acid, are able to elicit PR-1 via nonreactive oxygen species-requiring pathways.

  4. Potential role of plasma myeloperoxidase level in predicting long-term outcome of acute myocardial infarction.

    PubMed

    Kaya, Mehmet Gungor; Yalcin, Ridvan; Okyay, Kaan; Poyraz, Fatih; Bayraktar, Nilufer; Pasaoglu, Hatice; Boyaci, Bulent; Cengel, Atiye

    2012-01-01

    We investigated the prognostic importance of plasma myeloperoxidase levels in patients with ST-elevation myocardial infarction (STEMI) at long-term follow-up, and we analyzed the correlations between plasma myeloperoxidase levels and other biochemical values. We evaluated 73 consecutive patients (56 men; mean age, 56 ± 11 yr) diagnosed with acute STEMI and 46 age- and sex-matched healthy control participants. Patients were divided into 2 groups according to the median myeloperoxidase level (Group 1: plasma myeloperoxidase ≤ 68 ng/mL; and Group 2: plasma myeloperoxidase > 68 ng/mL). Patients were monitored for the occurrence of major adverse cardiovascular events (MACE), which were defined as cardiac death; reinfarction; new hospital admission for angina; heart failure; and revascularization procedures. The mean follow-up period was 25 ± 16 months. Plasma myeloperoxidase levels were higher in STEMI patients than in control participants (82 ± 34 vs 20 ± 12 ng/mL; P = 0.001). Composite MACE occurred in 12 patients with high myeloperoxidase levels (33%) and in 4 patients with low myeloperoxidase levels (11%) (P = 0.02). The incidences of nonfatal recurrent myocardial infarction and verified cardiac death were higher in the high-myeloperoxidase group. In multivariate analysis, high plasma myeloperoxidase levels were independent predictors of MACE (odds ratio = 3.843; <95% confidence interval, 1.625-6.563; P = 0.003). High plasma myeloperoxidase levels identify patients with a worse prognosis after acute STEMI at 2-year follow-up. Evaluation of plasma myeloperoxidase levels might be useful in determining patients at high risk of death and MACE who can benefit from further aggressive treatment and closer follow-up.

  5. Photosynthesis Activates Plasma Membrane H+-ATPase via Sugar Accumulation1[OPEN

    PubMed Central

    Okumura, Masaki; Inoue, Shin-ichiro; Kuwata, Keiko

    2016-01-01

    Plant plasma membrane H+-ATPase acts as a primary transporter via proton pumping and regulates diverse physiological responses by controlling secondary solute transport, pH homeostasis, and membrane potential. Phosphorylation of the penultimate threonine and the subsequent binding of 14-3-3 proteins in the carboxyl terminus of the enzyme are required for H+-ATPase activation. We showed previously that photosynthesis induces phosphorylation of the penultimate threonine in the nonvascular bryophyte Marchantia polymorpha. However, (1) whether this response is conserved in vascular plants and (2) the process by which photosynthesis regulates H+-ATPase phosphorylation at the plasma membrane remain unresolved issues. Here, we report that photosynthesis induced the phosphorylation and activation of H+-ATPase in Arabidopsis (Arabidopsis thaliana) leaves via sugar accumulation. Light reversibly phosphorylated leaf H+-ATPase, and this process was inhibited by pharmacological and genetic suppression of photosynthesis. Immunohistochemical and biochemical analyses indicated that light-induced phosphorylation of H+-ATPase occurred autonomously in mesophyll cells. We also show that the phosphorylation status of H+-ATPase and photosynthetic sugar accumulation in leaves were positively correlated and that sugar treatment promoted phosphorylation. Furthermore, light-induced phosphorylation of H+-ATPase was strongly suppressed in a double mutant defective in ADP-glucose pyrophosphorylase and triose phosphate/phosphate translocator (adg1-1 tpt-2); these mutations strongly inhibited endogenous sugar accumulation. Overall, we show that photosynthesis activated H+-ATPase via sugar production in the mesophyll cells of vascular plants. Our work provides new insight into signaling from chloroplasts to the plasma membrane ion transport mechanism. PMID:27016447

  6. PiZ Mouse Liver Accumulates Polyubiquitin Conjugates That Associate with Catalytically Active 26S Proteasomes

    PubMed Central

    Haddock, Christopher J.; Blomenkamp, Keith; Gautam, Madhav; James, Jared; Mielcarska, Joanna; Gogol, Edward; Teckman, Jeffrey; Skowyra, Dorota

    2014-01-01

    Accumulation of aggregation-prone human alpha 1 antitrypsin mutant Z (AT-Z) protein in PiZ mouse liver stimulates features of liver injury typical of human alpha 1 antitrypsin type ZZ deficiency, an autosomal recessive genetic disorder. Ubiquitin-mediated proteolysis by the 26S proteasome counteracts AT-Z accumulation and plays other roles that, when inhibited, could exacerbate the injury. However, it is unknown how the conditions of AT-Z mediated liver injury affect the 26S proteasome. To address this question, we developed a rapid extraction strategy that preserves polyubiquitin conjugates in the presence of catalytically active 26S proteasomes and allows their separation from deposits of insoluble AT-Z. Compared to WT, PiZ extracts had about 4-fold more polyubiquitin conjugates with no apparent change in the levels of the 26S and 20S proteasomes, and unassembled subunits. The polyubiquitin conjugates had similar affinities to ubiquitin-binding domain of Psmd4 and co-purified with similar amounts of catalytically active 26S complexes. These data show that polyubiquitin conjugates were accumulating despite normal recruitment to catalytically active 26S proteasomes that were available in excess, and suggest that a defect at the 26S proteasome other than compromised binding to polyubiquitin chain or peptidase activity played a role in the accumulation. In support of this idea, PiZ extracts were characterized by high molecular weight, reduction-sensitive forms of selected subunits, including ATPase subunits that unfold substrates and regulate access to proteolytic core. Older WT mice acquired similar alterations, implying that they result from common aspects of oxidative stress. The changes were most pronounced on unassembled subunits, but some subunits were altered even in the 26S proteasomes co-purified with polyubiquitin conjugates. Thus, AT-Z protein aggregates indirectly impair degradation of polyubiquitinated proteins at the level of the 26S proteasome

  7. Theaflavins attenuate hepatic lipid accumulation through activating AMPK in human HepG2 cells.

    PubMed

    Lin, Chih-Li; Huang, Hsiu-Chen; Lin, Jen-Kun

    2007-11-01

    Black tea is one of the world's most popular beverages, and its health-promoting effects have been intensively investigated. The antiobesity and hypolipidemic effects of black tea have attracted increasing interest, but the mechanisms underlying these phenomena remain unclear. In the present study, the black tea major component theaflavins were assessed for their hepatic lipid-lowering potential when administered in fatty acid overload conditions both in cell culture and in an animal experimental model. We found that theaflavins significantly reduced lipid accumulation, suppressed fatty acid synthesis, and stimulated fatty acid oxidation. Furthermore, theaflavins also inhibited acetyl-coenzyme A carboxylase activities by stimulating AMP-activated protein kinase (AMPK) through the LKB1 and reactive oxygen species pathways. These observations support the idea that AMPK is a critical component of decreased hepatic lipid accumulation by theaflavin treatments. Our results show that theaflavins are bioavailable both in vitro and in vivo and may be active in the prevention of fatty liver and obesity.

  8. Endogenous nitric oxide accumulation is involved in the antifungal activity of Shikonin against Candida albicans

    PubMed Central

    Liao, Zebin; Yan, Yu; Dong, Huaihuai; Zhu, Zhenyu; Jiang, Yuanying; Cao, Yingying

    2016-01-01

    The aim of the present study was to investigate the role of nitric oxide (NO) in the antifungal activity of Shikonin (SK) against Candida albicans (C. albicans) and to clarify the underlying mechanism. The results showed that the NO donors S-nitrosoglutathione (GSNO) and L-arginine could enhance the antifungal activity of SK, whereas the NO production inhibitor Nω-nitro-L-arginine methyl ester (L-NAME) attenuated antifungal action. Using the fluorescent dye 3-amino,4-aminomethyl-2′, 7-difluorescein, diacetate (DAF-FM DA), we found that the accumulation of NO in C. albicans was increased markedly by SK in a time- and dose-dependent manner. In addition, the results of real-time reverse transcription-PCR (RT-PCR) demonstrated that the transcription level of YHB1 in C. albicans was greatly increased upon incubation of SK. Consistently, the YHB1-null mutant (yhb1Δ/Δ) exhibited a higher susceptibility to SK than wild-type cells. In addition, although the transcription level of CTA4 in C. albicans was not significantly changed when exposed to SK, the CTA4-null mutant (cta4Δ/Δ) was more susceptible to SK. Collectively, SK is the agent found to execute its antifungal activity directly via endogenous NO accumulation, and NO-mediated damage is related to the suppression of YHB1 and the function of CTA4. PMID:27530748

  9. Oxidation of guaiacol by myeloperoxidase: a two-electron-oxidized guaiacol transient species as a mediator of NADPH oxidation.

    PubMed Central

    Capeillère-Blandin, C

    1998-01-01

    The present study was first aimed at a complete steady-state kinetic analysis of the reaction between guaiacol (2-methoxyphenol) and the myeloperoxidase (MPO)/H2O2 system, including a description of the isolation and purification of MPO from human polymorphonuclear neutrophil cells. Secondly, the overall reaction of the oxidation of NADPH, mediated by the reactive intermediates formed from the oxidation of guaiacol in the MPO/H2O2 system, was analysed kinetically. The presence of guaiacol stimulates the oxidation of NADPH by the MPO/H2O2 system in a concentration-dependent manner. Concomitantly, the accumulation of biphenoquinone (BQ), the final steady-state product of guaiacol oxidation, is lowered, and even inhibited completely, at high concentrations of NADPH. Under these conditions, the stoichiometry of NADPH:H2O2 is 1, and the oxidation rate of NADPH approximates to that of the rate of guaiacol oxidation by MPO. The effects of the presence of superoxide dismutase, catalase and of anaerobic conditions on the overall oxidation of NADPH have also been examined, and the data indicated that superoxide formation did not occur. The final product of NADPH oxidation was shown to be enzymically active NADP+, while guaiacol was generated continuously from the reaction between NADPH and oxidized guaiacol product. In contrast, similar experiments performed on the indirect, tyrosine-mediated oxidation of NADPH by MPO showed that a propagation of the free radical chain was occurring, with generation of both O2(-.) and H2O2. BQ, in itself, was able to spontaneously oxidize NADPH, but neither the rate nor the stoichiometry of the reaction could account for the NADPH-oxidation process involved in the steady-state peroxidation cycle. These results provide evidence that the oxidation of NADPH does not involve a free nucleotide radical intermediate, but that this is probably due to a direct electron-transfer reaction between NADPH and a two-electron-oxidized guaiacol intermediate

  10. Rapid adaptation of activated sludge bacteria into a glycogen accumulating biofilm enabling anaerobic BOD uptake.

    PubMed

    Hossain, Md Iqbal; Paparini, Andrea; Cord-Ruwisch, Ralf

    2017-03-01

    Glycogen accumulating organisms (GAO) are known to allow anaerobic uptake of biological oxygen demand (BOD) in activated sludge wastewater treatment systems. In this study, we report a rapid transition of suspended activated sludge biomass to a GAO dominated biofilm by selective enrichment using sequences of anaerobic loading followed by aerobic exposure of the biofilm to air. The study showed that within eight weeks, a fully operational, GAO dominated biofilm had developed, enabling complete anaerobic BOD uptake at a rate of 256mg/L/h. The oxygen uptake by the biofilm directly from the atmosphere had been calculated to provide significant energy savings. This study suggests that wastewater treatment plant operators can convert activated sludge systems readily into a "passive aeration" biofilm that avoids costly oxygen transfer to bulk wastewater solution. The described energy efficient BOD removal system provides an opportunity to be coupled with novel nitrogen removal processes such as anammox.

  11. Intracellular Accumulation of Glycine in Polyphosphate-Accumulating Organisms in Activated Sludge, a Novel Storage Mechanism under Dynamic Anaerobic-Aerobic Conditions

    PubMed Central

    Nguyen, Hien Thi Thu; Kristiansen, Rikke; Vestergaard, Mette; Wimmer, Reinhard

    2015-01-01

    Dynamic anaerobic-aerobic feast-famine conditions are applied to wastewater treatment plants to select polyphosphate-accumulating organisms to carry out enhanced biological phosphorus removal. Acetate is a well-known substrate to stimulate this process, and here we show that different amino acids also are suitable substrates, with glycine as the most promising. 13C-labeled glycine and nuclear magnetic resonance (NMR) were applied to investigate uptake and potential storage products when activated sludge was fed with glycine under anaerobic conditions. Glycine was consumed by the biomass, and the majority was stored intracellularly as free glycine and fermentation products. Subsequently, in the aerobic phase without addition of external substrate, the stored glycine was consumed. The uptake of glycine and oxidation of intracellular metabolites took place along with a release and uptake of orthophosphate, respectively. Fluorescence in situ hybridization combined with microautoradiography using 3H-labeled glycine revealed uncultured actinobacterial Tetrasphaera as a dominant glycine consumer. Experiments with Tetrasphaera elongata as representative of uncultured Tetrasphaera showed that under anaerobic conditions it was able to take up labeled glycine and accumulate this and other labeled metabolites to an intracellular concentration of approximately 4 mM. All components were consumed under subsequent aerobic conditions. Intracellular accumulation of amino acids seems to be a novel storage strategy for polyphosphate-accumulating bacteria under dynamic anaerobic-aerobic feast-famine conditions. PMID:25956769

  12. Triggered activity in atrial fibres of canine coronary sinus: role of extracellular potassium accumulation and depletion.

    PubMed Central

    Henning, B; Kline, R P; Siegal, M S; Wit, A L

    1987-01-01

    1. Bursts of triggered activity can be induced in atrial fibres of the canine coronary sinus exposed to catecholamines. During a triggered burst there is an initial acceleration of rate accompanied by depolarization of the maximum diastolic potential (m.d.p.) followed by slowing of the rate and termination accompanied by hyperpolarization. 2. We have used extracellular K+-sensitive micro-electrodes (potassium ISE) to monitor extracellular K+ concentration ([K+]o) during and following triggered activity, while simultaneously measuring membrane potential with conventional intracellular micro-electrodes. 3. We found that the initial increase in rate during triggered activity is accompanied by increased [K+]o and depolarization. Later rate slowing and m.d.p. hyperpolarization is accompanied by decline of extracellular K+ accumulation. Following termination of triggered activity, extracellular K+ depletion occurred. 4. The decline of [K+]o and slowing of rate are known responses to enhanced Na+-K+ pump activation, as is the post-triggering depletion of extracellular K+. 5. Strophanthidin, which blocks the Na+-K+ pump, also blocks the [K+]o decline, the slowing of rate seen towards the end of the triggered episode, and the post-triggering depletion of extracellular K+. 6. Separate experiments studying the effects of elevated bath K+ and depolarizing current on triggering rate and delayed after-depolarization amplitude support our hypothesis that the rate profile of the triggered episode is to a large extent controlled by variations in m.d.p. subsequent to extracellular K+ accumulation and Na+-K+ pump activation. PMID:3656124

  13. Structure-activity relationships of oligo-beta-glucoside elicitors of phytoalexin accumulation in soybean.

    PubMed Central

    Cheong, J J; Birberg, W; Fügedi, P; Pilotti, A; Garegg, P J; Hong, N; Ogawa, T; Hahn, M G

    1991-01-01

    The abilities of a family of chemically synthesized oligo-beta-glucosides, ranging in size from hexamer to decamer, to induce phytoalexin accumulation in soybean cotyledons were investigated to determine which structural elements of the oligoglucosides are important for their biological activity. The results of the biological assays established that the following structural motif is necessary for the oligo-beta-glucosides to have high elicitor activity: [formula; see text] The branched trisaccharide at the nonreducing end of the oligoglucosides was found to be essential for maximum elicitor activity. Substitution of either the nonreducing terminal backbone glucosyl residue or the side-chain glucosyl residue closest to the nonreducing end with glucosaminyl or N-acetylglucosaminyl residues reduced the elicitor activity of the oligoglucosides between 10-fold and 10,000-fold. Elicitor activity was also reduced 1000-fold if the two side-chain glucosyl residues were attached to adjacent backbone glucosyl residues rather than to glucosyl residues separated by an unbranched residue. In contrast, modifications of the reducing terminal glucosyl residue of an elicitor-active hepta-beta-glucoside by conjugation with tyramine and subsequent iodination had no significant effect on the elicitor activity of the hepta-beta-glucoside. These results demonstrate that oligo-beta-glucosides must have a specific structure to trigger the signal transduction pathway, which ultimately leads to the de novo synthesis of phytoalexins in soybean. PMID:1840904

  14. Rapid Changes on Sediment Accumulation Rates within Submarine Canyons Caused By Bottom Trawling Activities

    NASA Astrophysics Data System (ADS)

    Puig, P.; Masque, P.; Martin, J.; Paradis, S.; Juan, X.; Toro, M.; Palanques, A.

    2014-12-01

    The physical disturbance of the marine sedimentary environments by commercial bottom trawling is a matter of concern. The direct physical effects of this fishing technique include scraping and ploughing of the seabed and increases of the near-bottom water turbidity by sediment resuspension. However, the quantification of the sediment that has been resuspended by this anthropogenic activity over years and has been ultimately exported across the margin remains largely unaddressed. The analysis of sediment accumulation rates from sediment cores collected along the axes of several submarine canyons in the Catalan margin (northwestern Mediterranean) has allowed to estimate the contribution of this anthropogenic activity to the present-day sediment dynamics. 210Pb chronologies, occasionally supported by 137Cs dating, indicate a rapid increase of sediment accumulation rates since the 1970s, in coincidence with a strong impulse in the industrialization of the trawling fleets of this region. Such increase has been associated to the enhanced delivery of sediment resuspended by trawlers from the shelves and upper slope regions towards the canyon's interior, and to the rapid technical development at that time, in terms of engine power and gear size. This change has been observed in La Fonera (or Palamós) Canyon at depths greater than 1700 m, while in other canyons it is restricted to shallower regions (~1000 m in depth) closer to fishing grounds. Two sampling sites from La Fonera and Foix submarine canyons that exhibited high sediment accumulation rates (0.6-0.7 cm/y) were reoccupied several years after the first chronological analyses. These two new cores reveal a second and more rapid increase of sediment accumulation rates in both canyons occurring circa 2002 and accounting for about 2 cm/y. This second change at the beginning of the XXI century has been attributed to a preferential displacement of the trawling fleet towards slope fishing grounds surrounding submarine

  15. Epigenetic Salt Accumulation and Water Movement in the Active Layer of Central Yakutia in Eastern Siberia

    NASA Astrophysics Data System (ADS)

    Lopez Caceres, M.; Brouchkov, A.; Nakayama, H.; Takakai, F.; Fedorov, A.; Fukuda, M.

    2005-12-01

    Observations of soil moisture and salt content were conducted from May to August at Neleger station in Eastern Siberia. Seasonal changes of salt and soil moisture distribution in the active layer of larch forest (undisturbed) and a thermokarst depression known as alas (disturbed) were studied. Electric conductivity (ECe) of the intact forest revealed higher concentrations that increased with depth from the soil surface into the active layer and the underlying permafrost, 1 mS cm-1 at 1.1m to 2.6 mS cm-1 at 160 cm depth in the permafrost. However, maximum value of 5.4 mS cm-1 at 0.6 m depth was found in the dry area of alas. The concentration of ions, especially Na+, Mg2+, Ca2+, SO42-as well as HCO3- in the upper layers of this long-term disturbed site indicates the upward movement of ions together with water. Higher concentration of solutes was found in profiles with deeper seasonal thawing. The accumulation of salts in alas occurs from spring through the growing season. The low concentration of salt in the surface soil layers appears to be linked to leaching of salts by rainfall. There are substantial differences between water content and electric conductivity of soil in forest and alas. Modern salinization of the active layer in alas is epigenetic, and it happens in summer as a result of spring water collection and high summer evaporation; the gradual salt accumulation in alas in comparison to forest is controlled by annual balance of water and salts in the active layer. Present climatic trends point to continuous permafrost degradation in eastern Siberia increasing the risk of surface salinization which has already contributed to change the landscape by hindering the growth of forests.

  16. Epigenetic salt accumulation and water movement in the active layer of central Yakutia in eastern Siberia

    NASA Astrophysics Data System (ADS)

    Lopez, C. M. Larry; Brouchkov, A.; Nakayama, H.; Takakai, F.; Fedorov, A. N.; Fukuda, M.

    2007-01-01

    Observations of soil moisture and salt content were conducted from May to August at Neleger station in eastern Siberia. Seasonal changes of salt and soil moisture distribution in the active layer of larch forest (undisturbed) and a thermokarst depression known as an alas (disturbed) were studied. Electric conductivity ECe of the intact forest revealed higher concentrations that increased with depth from the soil surface into the active layer and the underlying permafrost: 1 mS cm-1 at 1.1 m, to 2.6 mS cm-1 at 160 cm depth in the permafrost. However, a maximum value of 5.4 mS cm-1 at 0.6 m depth was found in the dry area of the alas. The concentration of ions, especially Na+, Mg2+, Ca2+, SO42- and HCO3- in the upper layers of this long-term disturbed site, indicates the upward movement of ions together with water. A higher concentration of solutes was found in profiles with deeper seasonal thawing. The accumulation of salts in the alas occurs from spring through into the growing season. The low concentration of salt in the surface soil layers appears to be linked to leaching of salts by rainfall. There are substantial differences between water content and electric conductivity of soil in the forest and alas. Modern salinization of the active layer in the alas is epigenetic, and it happens in summer as a result of spring water collection and high summer evaporation; the gradual salt accumulation in the alas in comparison with the forest is controlled by the annual balance of water and salts in the active layer. Present climatic trends point to continuous permafrost degradation in eastern Siberia increasing the risk of surface salinization, which has already contributed to changing the landscape by hindering the growth of forest. Copyright

  17. Potential Role of Plasma Myeloperoxidase Level in Predicting Long-Term Outcome of Acute Myocardial Infarction

    PubMed Central

    Kaya, Mehmet Gungor; Yalcin, Ridvan; Okyay, Kaan; Poyraz, Fatih; Bayraktar, Nilufer; Pasaoglu, Hatice; Boyaci, Bulent; Cengel, Atiye

    2012-01-01

    We investigated the prognostic importance of plasma myeloperoxidase levels in patients with ST-elevation myocardial infarction (STEMI) at long-term follow-up, and we analyzed the correlations between plasma myeloperoxidase levels and other biochemical values. We evaluated 73 consecutive patients (56 men; mean age, 56 ±11 yr) diagnosed with acute STEMI and 46 age- and sex-matched healthy control participants. Patients were divided into 2 groups according to the median myeloperoxidase level (Group 1: plasma myeloperoxidase ≤68 ng/mL; and Group 2: plasma myeloperoxidase >68 ng/mL). Patients were monitored for the occurrence of major adverse cardiovascular events (MACE), which were defined as cardiac death; reinfarction; new hospital admission for angina; heart failure; and revascularization procedures. The mean follow-up period was 25 ± 16 months. Plasma myeloperoxidase levels were higher in STEMI patients than in control participants (82 ± 34 vs 20 ±12 ng/mL; P=0.001). Composite MACE occurred in 12 patients with high myeloperoxidase levels (33%) and in 4 patients with low myeloperoxidase levels (11%) (P=0.02). The incidences of nonfatal recurrent myocardial infarction and verified cardiac death were higher in the high-mye-loperoxidase group. In multivariate analysis, high plasma myeloperoxidase levels were independent predictors of MACE (odds ratio = 3.843; <95% confidence interval, 1.625–6.563; P=0.003). High plasma myeloperoxidase levels identify patients with a worse prognosis after acute STEMI at 2-year follow-up. Evaluation of plasma myeloperoxidase levels might be useful in determining patients at high risk of death and MACE who can benefit from further aggressive treatment and closer follow-up. PMID:22949765

  18. Along-strike Variations in Active Strain Accumulation in the Northwest Himalaya

    NASA Astrophysics Data System (ADS)

    Morell, K. D.; Ahmad, T.; Sandiford, M.; Codilean, A. T.; Fulop, R. H.

    2015-12-01

    The spatial distribution of channel steepness, erosion rate, and physiographic data highlight pronounced along-strike changes in active strain accumulation in the northwest Himalaya. In particular, the data suggest that the mid-crustal ramp of the Main Himalayan Thrust could merge along-strike with an active portion of the Main Boundary Thrust near longitude ~77º E. This along-strike change in active fault geometry also coincides with the lateral termination of both lesser and greater Himalayan sequences, a significant reduction in total shortening within the wedge, and pronounced variations in regional seismicity. Recent activity along extensional structures in the high Himalaya of this same region appears to have led to significant reorganization, modification and capture of the Sutlej River basin, one of the largest Himalayan river systems. Given the recent 2015 Gorkha earthquake along a comparable section ~500-km along strike, these new constraints on active fault architecture could have regional implications for how strain is partitioned along seismogenic faults in the northwest Himalaya.

  19. Irrigation with industrial wastewater activates antioxidant system and osmoprotectant accumulation in lettuce, turnip and tomato plants.

    PubMed

    Hashem, H A; Hassanein, R A; El-Deep, M H; Shouman, A I

    2013-09-01

    We focused on the impact of industrial wastes on the water quality of the El-Amia drain in Egypt and the effect of irrigation with industrial wastewater on the growth, cell membranes, photosynthetic pigment content, the antioxidant system and selected osmoprotectants (proline, total amino nitrogen and soluble sugars) in three crop plants: turnip, tomato and lettuce. Furthermore, the present work focused on the analysis of the heavy metal content and its accumulation in the studied plants. For this purpose, water samples were collected 1, 10 and 19 km from the beginning of the drain and used for irrigation, with fresh water as a control. We found that industrial wastewater contained significant amounts of heavy metals (Cd, Ni and Co) warranted a pollution problem as their amounts exceed the maximum recommended concentrations according to FAO guidelines for trace metals in irrigation water. The three crop plants accumulate significant amounts of heavy metals in their shoots and roots and showed a significant decrease in leaf area, fresh weight and dry weight of shoots and roots, accompanied by a marked reduction in photosynthetic pigment content and damage to cell membranes, as indicated by increased electrolyte leakage and a lower membrane stability index. Significant increases in the activities of antioxidant enzymes and in the glutathione, proline, soluble sugar and total amino nitrogen content in response to irrigation with wastewater may be defense mechanisms induced in response to heavy metal stress.

  20. Inhibition of myeloperoxidase: evaluation of 2H-indazoles and 1H-indazolones.

    PubMed

    Roth, Aaron; Ott, Sean; Farber, Kelli M; Palazzo, Teresa A; Conrad, Wayne E; Haddadin, Makhluf J; Tantillo, Dean J; Cross, Carroll E; Eiserich, Jason P; Kurth, Mark J

    2014-11-15

    Myeloperoxidase (MPO) produces hypohalous acids as a key component of the innate immune response; however, release of these acids extracellularly results in inflammatory cell and tissue damage. The two-step, one-pot Davis-Beirut reaction was used to synthesize a library of 2H-indazoles and 1H-indazolones as putative inhibitors of MPO. A structure-activity relationship study was undertaken wherein compounds were evaluated utilizing taurine-chloramine and MPO-mediated H2O2 consumption assays. Docking studies as well as toxicophore and Lipinski analyses were performed. Fourteen compounds were found to be potent inhibitors with IC50 values <1μM, suggesting these compounds could be considered as potential modulators of pro-oxidative tissue injury pertubated by the inflammatory MPO/H2O2/HOCl/HOBr system.

  1. Comparison of flash and accumulation mode in range-gated active imaging

    NASA Astrophysics Data System (ADS)

    Christnacher, Frank; Laurenzis, Martin; Schertzer, Stéphane

    2013-10-01

    Range-gated active imaging has significantly been improved in the recent past. Due to the availability of high power laser diodes around 800-860 nm, it is now possible to find off-the-shelf systems working with very sensitive light intensifier and laser diodes. On the other hand, eye-safe systems working around 1.5 μm suffer from a lack of intensified sensor in the SWIR band. The only existing intensified sensors require the use of high power pulsed laser sources for the illumination. Consequently, the type of source (diode or solid-state laser) gives fundamental differences between the two types of system. The first technique which uses laser diodes, μchip or fiber lasers, is called "accumulation" imaging. These sources are characterized by a low-pulse power and high repetition rate, mostly around a few tens of kHz. Here, each image is the result of the accumulation of hundred of pulses during the frame time. The second technique which uses a solid-state laser illumination is called "flash" imaging. Here, each image is the result of a unique high power illumination of the scene at low repetition rate, mostly around the video rate. In this paper, we investigate the theoretical and practical differences between these two imaging modes and its influence on image quality, on sensitivity to day light or stray light, on fog penetration capacity, on its sensitivity to turbulences and on laser safety (NOHD). For comparative experimental purposes, we've built a range-gated active imaging system which allows the investigation of both methods. We've carried out precise comparative studies between the two acquisition methods.

  2. Accumulated source imaging of brain activity with both low and high-frequency neuromagnetic signals

    PubMed Central

    Xiang, Jing; Luo, Qian; Kotecha, Rupesh; Korman, Abraham; Zhang, Fawen; Luo, Huan; Fujiwara, Hisako; Hemasilpin, Nat; Rose, Douglas F.

    2014-01-01

    Recent studies have revealed the importance of high-frequency brain signals (>70 Hz). One challenge of high-frequency signal analysis is that the size of time-frequency representation of high-frequency brain signals could be larger than 1 terabytes (TB), which is beyond the upper limits of a typical computer workstation's memory (<196 GB). The aim of the present study is to develop a new method to provide greater sensitivity in detecting high-frequency magnetoencephalography (MEG) signals in a single automated and versatile interface, rather than the more traditional, time-intensive visual inspection methods, which may take up to several days. To address the aim, we developed a new method, accumulated source imaging, defined as the volumetric summation of source activity over a period of time. This method analyzes signals in both low- (1~70 Hz) and high-frequency (70~200 Hz) ranges at source levels. To extract meaningful information from MEG signals at sensor space, the signals were decomposed to channel-cross-channel matrix (CxC) representing the spatiotemporal patterns of every possible sensor-pair. A new algorithm was developed and tested by calculating the optimal CxC and source location-orientation weights for volumetric source imaging, thereby minimizing multi-source interference and reducing computational cost. The new method was implemented in C/C++ and tested with MEG data recorded from clinical epilepsy patients. The results of experimental data demonstrated that accumulated source imaging could effectively summarize and visualize MEG recordings within 12.7 h by using approximately 10 GB of computer memory. In contrast to the conventional method of visually identifying multi-frequency epileptic activities that traditionally took 2–3 days and used 1–2 TB storage, the new approach can quantify epileptic abnormalities in both low- and high-frequency ranges at source levels, using much less time and computer memory. PMID:24904402

  3. Activity blockade and GABAA receptor blockade produce synaptic scaling through chloride accumulation in embryonic spinal motoneurons and interneurons.

    PubMed

    Lindsly, Casie; Gonzalez-Islas, Carlos; Wenner, Peter

    2014-01-01

    Synaptic scaling represents a process whereby the distribution of a cell's synaptic strengths are altered by a multiplicative scaling factor. Scaling is thought to be a compensatory response that homeostatically controls spiking activity levels in the cell or network. Previously, we observed GABAergic synaptic scaling in embryonic spinal motoneurons following in vivo blockade of either spiking activity or GABAA receptors (GABAARs). We had determined that activity blockade triggered upward GABAergic scaling through chloride accumulation, thus increasing the driving force for these currents. To determine whether chloride accumulation also underlies GABAergic scaling following GABAAR blockade we have developed a new technique. We expressed a genetically encoded chloride-indicator, Clomeleon, in the embryonic chick spinal cord, which provides a non-invasive fast measure of intracellular chloride. Using this technique we now show that chloride accumulation underlies GABAergic scaling following blockade of either spiking activity or the GABAAR. The finding that GABAAR blockade and activity blockade trigger scaling via a common mechanism supports our hypothesis that activity blockade reduces GABAAR activation, which triggers synaptic scaling. In addition, Clomeleon imaging demonstrated the time course and widespread nature of GABAergic scaling through chloride accumulation, as it was also observed in spinal interneurons. This suggests that homeostatic scaling via chloride accumulation is a common feature in many neuronal classes within the embryonic spinal cord and opens the possibility that this process may occur throughout the nervous system at early stages of development.

  4. Role of Myeloperoxidase in Patients with Chronic Kidney Disease

    PubMed Central

    Kisic, Bojana; Miric, Dijana; Dragojevic, Ilija; Rasic, Julijana; Popovic, Ljiljana

    2016-01-01

    Chronic kidney disease (CKD) is a worldwide public health problem. Patients with CKD have a number of disorders in the organism, and the presence of oxidative stress and systemic inflammation in these patients is the subject of numerous studies. Chronic inflammation joined with oxidative stress contributes to the development of numerous complications: accelerated atherosclerosis process and cardiovascular disease, emergence of Type 2 diabetes mellitus, development of malnutrition, anaemia, hyperparathyroidism, and so forth, affecting the prognosis and quality of life of patients with CKD. In this review we presented the potential role of the myeloperoxidase enzyme in the production of reactive/chlorinating intermediates and their role in oxidative damage to biomolecules in the body of patients with chronic kidney disease and end-stage renal disease. In addition, we discussed the role of modified lipoprotein particles under the influence of prooxidant MPO intermediates in the development of endothelial changes and cardiovascular complications in renal failure. PMID:27127544

  5. Utilizing G2/M retention effect to enhance tumor accumulation of active targeting nanoparticles

    PubMed Central

    Hu, Guanlian; Cun, Xingli; Ruan, Shaobo; Shi, Kairong; Wang, Yang; Kuang, Qifang; Hu, Chuan; Xiao, Wei; He, Qin; Gao, Huile

    2016-01-01

    In recent years, active targeting strategies by ligand modification have emerged to enhance tumor accumulation of NP, but their clinical application was strictly restricted due to the complex preparation procedures, poor stability and serious toxicity. An effective and clinical translational strategy is required to satisfy the current problems. Interestingly, the internalization of NP is intimately related with cell cycle and the expression of receptors is not only related with cancer types but also cell cycle progression. So the cellular uptake of ligand modified NP may be related with cell cycle. However, few investigations were reported about the relationship between cell cycle and the internalization of ligand modified NP. Herein, cellular uptake of folic acid (FA) modified NP after utilizing chemotherapeutic to retain the tumor cells in G2/M phase was studied and a novel strategy was designed to enhance the active targeting effect. In our study, docetaxel (DTX) notably synchronized cells in G2/M phase and pretreatment with DTX highly improved in vitro and in vivo tumor cell targeting effect of FA decorated NP (FANP). Since FA was a most common used tumor active targeting ligand, we believe that this strategy possesses broader prospects in clinical application for its simplicity and effectiveness. PMID:27273770

  6. Excess beta-catenin promotes accumulation of transcriptionally active p53.

    PubMed Central

    Damalas, A; Ben-Ze'ev, A; Simcha, I; Shtutman, M; Leal, J F; Zhurinsky, J; Geiger, B; Oren, M

    1999-01-01

    beta-catenin is a multifunctional protein, acting both as a structural component of the cell adhesion machinery and as a transducer of extracellular signals. Deregulated beta-catenin protein expression, due to mutations in the beta-catenin gene itself or in its upstream regulator, the adenomatous polyposis coli (APC) gene, is prevalent in colorectal cancer and in several other tumor types, and attests to the potential oncogenic activity of this protein. Increased expression of beta-catenin is an early event in colorectal carcinogenesis, and is usually followed by a later mutational inactivation of the p53 tumor suppressor. To examine whether these two key steps in carcinogenesis are interrelated, we studied the effect of excess beta-catenin on p53. We report here that overexpression of beta-catenin results in accumulation of p53, apparently through interference with its proteolytic degradation. This effect involves both Mdm2-dependent and -independent p53 degradation pathways, and is accompanied by augmented transcriptional activity of p53 in the affected cells. Increased p53 activity may provide a safeguard against oncogenic deregulation of beta-catenin, and thus impose a pressure for mutational inactivation of p53 during the later stages of tumor progression. PMID:10357817

  7. Effects of copper sulfate-oxidized or myeloperoxidase-modified LDL on lipid loading and programmed cell death in macrophages under hypoxia

    PubMed Central

    Vlaminck, Benoit; Calay, Damien; Genin, Marie; Sauvage, Aude; Ninane, Noelle; Zouaoui Boudjeltia, Karim; Raes, Martine; Michiels, Carine

    2014-01-01

    Atheromatous plaques contain heavily lipid-loaded macrophages that die, hence generating the necrotic core of these plaques. Since plaque instability and rupture is often correlated with a large necrotic core, it is important to understand the mechanisms underlying foam cell death. Furthermore, macrophages within the plaque are associated with hypoxic areas but little is known about the effect of low oxygen partial pressure on macrophage death. The aim of this work was to unravel macrophage death mechanisms induced by oxidized low-density lipoproteins (LDL) both under normoxia and hypoxia. Differentiated macrophages were incubated in the presence of native, copper sulfate-oxidized, or myeloperoxidase-modified LDL. The unfolded protein response, apoptosis, and autophagy were then investigated. The unfolded protein response and autophagy were triggered by myeloperoxidase-modified LDL and, to a larger extent, by copper sulfate-oxidized LDL. Electron microscopy observations showed that oxidized LDL induced excessive autophagy and apoptosis under normoxia, which were less marked under hypoxia. Myeloperoxidase-modified LDL were more toxic and induced a higher level of apoptosis. Hypoxia markedly decreased apoptosis and cell death, as marked by caspase activation. In conclusion, the cell death pathways induced by copper sulfate-oxidized and myeloperoxidase-modified LDL are different and are differentially modulated by hypoxia. PMID:27774474

  8. Fractional activation of accumulation-mode particles in warm continental stratiform clouds

    SciTech Connect

    Gillani, N.V.; Daum, P.H.; Schwartz, S.E.; Leaitch, W.R.; Strapp, J.W.; Isaac, G.A.

    1991-07-01

    The degree of activation of accumulation-mode particles (AMP) in clouds has been studied using continuous (1 second average) aircraft measurements of the number concentrations of cloud droplets (N{sub cd}, 2 to 35 {mu}m diameter) and of unactivated AMP (N{sub amp}, 0.17 to 2.07 {mu}m diameter) in cloud interstitial air. The magnitude and spatial variation of the activated fraction (F) of all measured particles (defined as F {triple_bond} N{sub cd}/N{sub tot}, where N{sub tot} = N{sub cd} + N{sub amp}) are investigated, based on measurements made during ten aircraft flights in non-precipitating warm continental stratiform clouds near Syracuse NY in the fall of 1984. Based on instantaneous observations throughout the clouds, the spatial distribution of F was found to be quite nonuniform. In general, F was low in cloud edges and where total particle loading was high and/or cloud convective activity was low. In the interior of clouds, the value of F exceeded 0.9 for 36% of the data, but was below 0.6 for 28%. Factors influencing F the most were the total particle loading (N{sub tot}) and the thermal stability of the cloud layer. The dependence of F on N{sub tot} in cloud interior was characterized by two distinct regimes. For N{sub tot} < 600 cm{sup {minus}3}, F was generally close to unity and relatively insensitive to N{sub tot}. For N{sub tot} > 800 cm{sup {minus}3}, F tended to decrease with increasing N{sub tot}. This decrease was greatest in a stable stratus deck embedded in a warm moist airmass. The results suggest that, in warm continental stratiform clouds, the process of particle activation becomes nonlinear and self-limiting at high particle loading. The degree of this nonlinearity depends on cloud convective activity (thermal instability).

  9. Fractional activation of accumulation-mode particles in warm continental stratiform clouds

    SciTech Connect

    Gillani, N.V. Associates, Inc., St. Louis, MO ); Daum, P.H.; Schwartz, S.E. ); Leaitch, W.R.; Strapp, J.W.; Isaac, G.A. . Cloud Physics Research Div.)

    1991-07-01

    The degree of activation of accumulation-mode particles (AMP) in clouds has been studied using continuous (1 second average) aircraft measurements of the number concentrations of cloud droplets (N[sub cd], 2 to 35 [mu]m diameter) and of unactivated AMP (N[sub amp], 0.17 to 2.07 [mu]m diameter) in cloud interstitial air. The magnitude and spatial variation of the activated fraction (F) of all measured particles (defined as F [triple bond] N[sub cd]/N[sub tot], where N[sub tot] = N[sub cd] + N[sub amp]) are investigated, based on measurements made during ten aircraft flights in non-precipitating warm continental stratiform clouds near Syracuse NY in the fall of 1984. Based on instantaneous observations throughout the clouds, the spatial distribution of F was found to be quite nonuniform. In general, F was low in cloud edges and where total particle loading was high and/or cloud convective activity was low. In the interior of clouds, the value of F exceeded 0.9 for 36% of the data, but was below 0.6 for 28%. Factors influencing F the most were the total particle loading (N[sub tot]) and the thermal stability of the cloud layer. The dependence of F on N[sub tot] in cloud interior was characterized by two distinct regimes. For N[sub tot] < 600 cm[sup [minus]3], F was generally close to unity and relatively insensitive to N[sub tot]. For N[sub tot] > 800 cm[sup [minus]3], F tended to decrease with increasing N[sub tot]. This decrease was greatest in a stable stratus deck embedded in a warm moist airmass. The results suggest that, in warm continental stratiform clouds, the process of particle activation becomes nonlinear and self-limiting at high particle loading. The degree of this nonlinearity depends on cloud convective activity (thermal instability).

  10. Mercury modulates selenium activity via altering its accumulation and speciation in garlic (Allium sativum).

    PubMed

    Zhao, Jiating; Hu, Yi; Gao, Yuxi; Li, Yufeng; Li, Bai; Dong, Yuanxing; Chai, Zhifang

    2013-06-01

    Combined pollution of selenium (Se) and mercury (Hg) has been known in Wanshan district (Guizhou Province, China). A better understanding of how Se and Hg interact in plants and the phytotoxicity thereof will provide clues about how to avoid or mitigate adverse effects of Se/Hg on local agriculture. In this study, the biological activity of Se has been investigated in garlic with or without Hg exposure. Se alone can promote garlic growth at low levels (<0.1 mg L(-1)), whereas it inhibits garlic growth at high levels (>1 mg L(-1)). The promotive effect of Se in garlic can be enhanced by low Hg exposure (<0.1 mg L(-1)). When both Se and Hg are at high levels, there is a general antagonistic effect between these two elements in terms of phytotoxicity. Inductively coupled plasma mass spectrometry (ICP-MS) data suggest that Se is mainly concentrated in garlic roots, compared to the leaves and the bulbs. Se uptake by garlic in low Se medium (<0.1 mg L(-1)) can be significantly enhanced as Hg exposure levels increase (P < 0.05), while it can be inhibited by Hg when Se exposure levels exceed 1 mg L(-1). The synchrotron radiation X-ray fluorescence (SRXRF) mapping further shows that Se is mainly concentrated in the stele of the roots, bulbs and the veins of the leaves, and Se accumulation in garlic can be reduced by Hg. The X-ray absorption near edge structure (XANES) study indicates that Se is mainly formed in C-Se-C form in garlic. Hg can decrease the content of inorganic Se mainly in SeO3(2-) form in garlic while increasing the content of organic Se mainly in C-Se-C form (MeSeCys and its derivatives). Hg-mediated changes in Se species along with reduced Se accumulation in garlic may account for the protective effect of Hg against Se phytotoxicity.

  11. Two chitinase-like proteins abundantly accumulated in latex of mulberry show insecticidal activity

    PubMed Central

    2010-01-01

    Background Plant latex is the cytoplasm of highly specialized cells known as laticifers, and is thought to have a critical role in defense against herbivorous insects. Proteins abundantly accumulated in latex might therefore be involved in the defense system. Results We purified latex abundant protein a and b (LA-a and LA-b) from mulberry (Morus sp.) and analyzed their properties. LA-a and LA-b have molecular masses of approximately 50 and 46 kDa, respectively, and are abundant in the soluble fraction of latex. Western blotting analysis suggested that they share sequence similarity with each other. The sequences of LA-a and LA-b, as determined by Edman degradation, showed chitin-binding domains of plant chitinases at the N termini. These proteins showed small but significant chitinase and chitosanase activities. Lectin RCA120 indicated that, unlike common plant chitinases, LA-a and LA-b are glycosylated. LA-a and LA-b showed insecticidal activities when fed to larvae of the model insect Drosophila melanogaster. Conclusions Our results suggest that the two LA proteins have a crucial role in defense against herbivorous insects, possibly by hydrolyzing their chitin. PMID:20109180

  12. Virucidal efficacy of treatment with photodynamically activated curcumin on murine norovirus bio-accumulated in oysters.

    PubMed

    Wu, Juan; Hou, Wei; Cao, Binbin; Zuo, Tao; Xue, Changhu; Leung, Albert Wingnang; Xu, Chuanshan; Tang, Qing-Juan

    2015-09-01

    Norovirus (NoV) is one of the most important seafood- and water-borne viruses, and is a major cause of acute gastroenteritis outbreaks. In the present study we investigated the effect of curcumin as a sensitizer to photodynamic treatment both in buffer and in oysters against murine norovirus 1 (MNV-1), a surrogate of NoV. MNV-1 cultured in buffer and MNV-1 bio-accumulated in oysters were irradiated with a novel LED light source with a wavelength of 470nm and an energy of 3.6J/cm(2). Inactivation of MNV-1 was investigated by plaque assays. After virus was extracted from the gut of oysters treated over a range of curcumin concentrations, the ultrastructural morphology of the virus was observed using electron microscopy, and the integrity of viral nucleic acids and stability of viral capsid proteins were also determined. Results showed that the infectivity of MNV-1 was significantly inhibited by 1-3logPFU/ml, with significant damage to viral nucleic acids in a curcumin dose-dependent manner after photodynamic activation. Virus morphology was altered after the photodynamic treatment with curcumin, presumably due to the change of the viral capsid protein structures. The data suggest that treatment of oysters with photodynamic activation of curcumin is a potentially efficacious and cost-effective method to inactivate food-borne NoV. Further studies are necessary to evaluate the toxicology of this approach in detail and perform sensory evaluation of the treated product.

  13. Calibration of denitrifying activity of polyphosphate accumulating organisms in an extended ASM2d model.

    PubMed

    García-Usach, F; Ribes, J; Ferrer, J; Seco, A

    2010-10-01

    This paper presents the results of an experimental study for the modelling and calibration of denitrifying activity of polyphosphate accumulating organisms (PAOs) in full-scale WWTPs that incorporate simultaneous nitrogen and phosphorus removal. The convenience of using different yields under aerobic and anoxic conditions for modelling biological phosphorus removal processes with the ASM2d has been demonstrated. Thus, parameter η(PAO) in the model is given a physical meaning and represents the fraction of PAOs that are able to follow the DPAO metabolism. Using stoichiometric relationships, which are based on assumed biochemical pathways, the anoxic yields considered in the extended ASM2d can be obtained as a function of their respective aerobic yields. Thus, this modification does not mean an extra calibration effort to obtain the new parameters. In this work, an off-line calibration methodology has been applied to validate the model, where general relationships among stoichiometric parameters are proposed to avoid increasing the number of parameters to calibrate. The results have been validated through a UCT scheme pilot plant that is fed with municipal wastewater. The good concordance obtained between experimental and simulated values validates the use of anoxic yields as well as the calibration methodology. Deterministic modelling approaches, together with off-line calibration methodologies, are proposed to assist in decision-making about further process optimization in biological phosphate removal, since parameter values obtained by off-line calibration give valuable information about the activated sludge process such as the amount of DPAOs in the system.

  14. Essential role of CD11a in CD8+ T-cell accumulation and activation in adipose tissue

    Technology Transfer Automated Retrieval System (TEKTRAN)

    T-cells, particularly CD8+ T-cells, are major participants in obesity-linked adipose tissue inflammation. We examined the mechanisms of CD8+ T-cell accumulation and activation in adipose tissue and the role of CD11a, a beta2 integrin. CD8+ T-cells in adipose tissue of obese mice showed activated phe...

  15. Induction of Mast Cell Accumulation by Tryptase via a Protease Activated Receptor-2 and ICAM-1 Dependent Mechanism

    PubMed Central

    Liu, Xin; Wang, Junling; Zhang, Huiyun; Zhan, Mengmeng; Chen, Hanqiu; Fang, Zeman; Xu, Chiyan; Chen, Huifang; He, Shaoheng

    2016-01-01

    Mast cells are primary effector cells of allergy, and recruitment of mast cells in involved tissue is one of the key events in allergic inflammation. Tryptase is the most abundant secretory product of mast cells, but little is known of its influence on mast cell accumulation. Using mouse peritoneal model, cell migration assay, and flow cytometry analysis, we investigated role of tryptase in recruiting mast cells. The results showed that tryptase induced up to 6.7-fold increase in mast cell numbers in mouse peritoneum following injection. Inhibitors of tryptase, an antagonist of PAR-2 FSLLRY-NH2, and pretreatment of mice with anti-ICAM-1, anti-CD11a, and anti-CD18 antibodies dramatically diminished tryptase induced mast cell accumulation. On the other hand, PAR-2 agonist peptides SLIGRL-NH2 and tc-LIGRLO-NH2 provoked mast cell accumulation following injection. These implicate that tryptase induced mast cell accumulation is dependent on its enzymatic activity, activation of PAR-2, and interaction between ICAM-1 and LFA-1. Moreover, induction of trans-endothelium migration of mast cells in vitro indicates that tryptase acts as a chemoattractant. In conclusion, provocation of mast cell accumulation by mast cell tryptase suggests a novel self-amplification mechanism of mast cell accumulation. Mast cell stabilizers as well as PAR-2 antagonist agents may be useful for treatment of allergic reactions. PMID:27378825

  16. Effect of Inoculation with Glomus versiforme on Cadmium Accumulation, Antioxidant Activities and Phytochelatins of Solanum photeinocarpum

    PubMed Central

    Tan, Shi-Yun; Jiang, Qiu-Yun; Zhuo, Feng; Liu, Hui; Wang, Yu-Tao; Li, Shao-Shan; Ye, Zhi-Hong; Jing, Yuan-Xiao

    2015-01-01

    The plant growth, phosphate acquisition, Cd translocation, phytochelatins (PCs) production and antioxidant parameters [superoxide dismutase (SOD), catalase (CAT), guaiacol peroxidase (POD), ascorbate peroxidase (APX), glutathione reductase (GR), glutathione (GSH), ascorbate (ASA) and malonaldehyde (MDA)] were investigated in Cd-hyperaccumulator Solanum photeinocarpum inoculated with Glomus versiforme BGC GD01C (Gv) in Cd-added soils (0, 5, 10, 20, 40 mg Cd kg-1 soil). Mycorrhizal colonization rates were generally high (from 77% to 94%), and hardly affected by Cd. Gv colonization significantly enhanced P acquisition, growth and total Cd uptakes in both shoots and roots of S. photeinocarpum at all Cd levels. Meanwhile, Gv symbiosis significantly increased Cd concentration in the roots, and decreased Cd concentration in the shoots at all Cd levels, which indicates that Gv could promote phytostabilization by enhancing Cd accumulation in the roots to inhibit its translocation to shoots and the “dilution effects” linked to an increase in plant dry matter yield and a reduced Cd partitioning to shoots. Moreover, the improvement of CAT, POD and APX activities in the leaves of mycorrhizal plants infers that Gv symbiosis helped S. photeinocarpum to relieve oxidative damage to biomolecules in Cd-contaminated soil. The evident decline of MDA content in the leaves of mycorrhizal plants indicates that Gv symbiosis evidently improved antioxidant activities, and the enhancement of PCs production in the leaves of mycorrhizal plants suggests that Gv-inoculated plant may be more efficient to relieve Cd phytotoxicity. Therefore, the possible mechanisms of Cd phytotoxicity alleviation by Gv can be concluded as the decline of Cd concentration in the shoots and the improvement of P acquisition, PCs production and activities of CAT, POD, APX in mycorrhizal plants. PMID:26176959

  17. Accumulate repeat accumulate codes

    NASA Technical Reports Server (NTRS)

    Abbasfar, Aliazam; Divsalar, Dariush; Yao, Kung

    2004-01-01

    In this paper we propose an innovative channel coding scheme called 'Accumulate Repeat Accumulate codes' (ARA). This class of codes can be viewed as serial turbo-like codes, or as a subclass of Low Density Parity Check (LDPC) codes, thus belief propagation can be used for iterative decoding of ARA codes on a graph. The structure of encoder for this class can be viewed as precoded Repeat Accumulate (RA) code or as precoded Irregular Repeat Accumulate (IRA) code, where simply an accumulator is chosen as a precoder. Thus ARA codes have simple, and very fast encoder structure when they representing LDPC codes. Based on density evolution for LDPC codes through some examples for ARA codes, we show that for maximum variable node degree 5 a minimum bit SNR as low as 0.08 dB from channel capacity for rate 1/2 can be achieved as the block size goes to infinity. Thus based on fixed low maximum variable node degree, its threshold outperforms not only the RA and IRA codes but also the best known LDPC codes with the dame maximum node degree. Furthermore by puncturing the accumulators any desired high rate codes close to code rate 1 can be obtained with thresholds that stay close to the channel capacity thresholds uniformly. Iterative decoding simulation results are provided. The ARA codes also have projected graph or protograph representation that allows for high speed decoder implementation.

  18. Accumulation of fossil fuels and metallic minerals in active and ancient rift lakes

    USGS Publications Warehouse

    Robbins, E.I.

    1983-01-01

    A study of active and ancient rift systems around the world suggests that accumulations of fossil fuels and metallic minerals are related to the interactions of processes that form rift valleys with those that take place in and around rift lakes. The deposition of the precursors of petroleum, gas, oil shale, coal, phosphate, barite, Cu-Pb-Zn sulfides, and uranium begins with erosion of uplifted areas, and the consequent input of abundant nutrients and solute loads into swamps and tectonic lakes. Hot springs and volcanism add other nutrients and solutes. The resulting high biological productivity creates oxidized/reduced interfaces, and anoxic and H2S-rich bottom waters which preserves metal-bearing organic tissues and horizons. In the depositional phases, the fine-grained lake deposits are in contact with coarse-grained beach, delta, river, talus, and alluvial fan deposits. Earthquake-induced turbidites also are common coarse-grained deposits of rift lakes. Postdepositional processes in rifts include high heat flow and a resulting concentration of the organic and metallic components that were dispersed throughout the lakebeds. Postdepositional faulting brings organic- and metal-rich sourcebeds in contact with coarse-grained host and reservoir rocks. A suite of potentially economic deposits is therefore a characteristic of rift valleys. ?? 1983.

  19. Ganglioside accumulation in activated glia in the developing brain: comparison between WT and GalNAcT KO mice.

    PubMed

    Saito, Mariko; Wu, Gusheng; Hui, Maria; Masiello, Kurt; Dobrenis, Kostantin; Ledeen, Robert W; Saito, Mitsuo

    2015-08-01

    Our previous studies have shown accumulation of GM2 ganglioside during ethanol-induced neurodegeneration in the developing brain, and GM2 elevation has also been reported in other brain injuries and neurodegenerative diseases. Using GM2/GD2 synthase KO mice lacking GM2/GD2 and downstream gangliosides, the current study explored the significance of GM2 elevation in WT mice. Immunohistochemical studies indicated that ethanol-induced acute neurodegeneration in postnatal day 7 (P7) WT mice was associated with GM2 accumulation in the late endosomes/lysosomes of both phagocytic microglia and increased glial fibrillary acidic protein (GFAP)-positive astrocytes. However, in KO mice, although ethanol induced robust neurodegeneration and accumulation of GD3 and GM3 in the late endosomes/lysosomes of phagocytic microglia, it did not increase the number of GFAP-positive astrocytes, and the accumulation of GD3/GM3 in astrocytes was minimal. Not only ethanol, but also DMSO, induced GM2 elevation in activated microglia and astrocytes along with neurodegeneration in P7 WT mice, while lipopolysaccharide, which did not induce significant neurodegeneration, caused GM2 accumulation mainly in lysosomes of activated astrocytes. Thus, GM2 elevation is associated with activation of microglia and astrocytes in the injured developing brain, and GM2, GD2, or other downstream gangliosides may regulate astroglial responses in ethanol-induced neurodegeneration.

  20. Ganglioside accumulation in activated glia in the developing brain: comparison between WT and GalNAcT KO mice

    PubMed Central

    Saito, Mariko; Wu, Gusheng; Hui, Maria; Masiello, Kurt; Dobrenis, Kostantin; Ledeen, Robert W.; Saito, Mitsuo

    2015-01-01

    Our previous studies have shown accumulation of GM2 ganglioside during ethanol-induced neurodegeneration in the developing brain, and GM2 elevation has also been reported in other brain injuries and neurodegenerative diseases. Using GM2/GD2 synthase KO mice lacking GM2/GD2 and downstream gangliosides, the current study explored the significance of GM2 elevation in WT mice. Immunohistochemical studies indicated that ethanol-induced acute neurodegeneration in postnatal day 7 (P7) WT mice was associated with GM2 accumulation in the late endosomes/lysosomes of both phagocytic microglia and increased glial fibrillary acidic protein (GFAP)-positive astrocytes. However, in KO mice, although ethanol induced robust neurodegeneration and accumulation of GD3 and GM3 in the late endosomes/lysosomes of phagocytic microglia, it did not increase the number of GFAP-positive astrocytes, and the accumulation of GD3/GM3 in astrocytes was minimal. Not only ethanol, but also DMSO, induced GM2 elevation in activated microglia and astrocytes along with neurodegeneration in P7 WT mice, while lipopolysaccharide, which did not induce significant neurodegeneration, caused GM2 accumulation mainly in lysosomes of activated astrocytes. Thus, GM2 elevation is associated with activation of microglia and astrocytes in the injured developing brain, and GM2, GD2, or other downstream gangliosides may regulate astroglial responses in ethanol-induced neurodegeneration. PMID:26063460

  1. Trichothecenes induce accumulation of glucosylceramide in neural cells by interfering with lactosylceramide synthase activity

    SciTech Connect

    Kralj, Ana; Gurgui, Mihaela; Koenig, Gabriele M.; Echten-Deckert, Gerhild van

    2007-11-15

    Trichothecenes are sesquiterpenoid metabolites produced by several fungal strains that impair human and animal health. Since sphingolipids were connected with fungal toxicity the aim of the present study was to test the influence of fungal metabolites on sphingolipid metabolism in neural cells. The crude extract of fungal strain Spicellum roseum induced accumulation of glucosylceramide (GlcCer), and simultaneous reduction of the formation of lactosylceramide (LacCer) and complex gangliosides in primary cultured neurons. Following a bioassay-guided fractionation of the respective fungal extract we could demonstrate that the two isolated trichothecene derivatives, 8-deoxy-trichothecin (8-dT) and trichodermol (Td-ol) were responsible for this effect. Thus, incubation of primary cultured neurons as well as of neuroblastoma B104 cells for 24 h with 30 {mu}M of either of the two fungal metabolites resulted in uncoupling of sphingolipid biosynthesis at the level of LacCer. For the observed reduction of LacCer synthase activity by about 90% cell integrity was crucial in both cell types. In neuroblastoma cells the amount of LacCer synthase mRNA was reduced in the presence of trichothecenes, whereas in primary cultured neurons this was not the case, suggesting a post-transcriptional mechanism of action in the latter cell type. The data also show that the compounds did not interfere with the translocation of GlcCer in neuroblastoma cells. Collectively, our results demonstrate that trichodermol and 8-deoxy-trichothecin inhibit LacCer synthase activity in a cell-type-specific manner.

  2. Arbuscular mycorrhizae improve low temperature tolerance in cucumber via alterations in H2O2 accumulation and ATPase activity.

    PubMed

    Liu, Airong; Chen, Shuangchen; Chang, Rui; Liu, Dilin; Chen, Haoran; Ahammed, Golam Jalal; Lin, Xiaomin; He, Chaoxing

    2014-11-01

    The combined effects of arbuscular mycorrhizal fungi (AMF) and low temperature (LT) on cucumber plants were investigated with respect to biomass production, H2O2 accumulation, NADPH oxidase, ATPase activity and related gene expression. Mycorrhizal colonization ratio was gradually increased after AMF-inoculation. However, LT significantly decreased mycorrhizal colonization ability and mycorrhizal dependency. Regardless of temperature, the total fresh and dry mass, and root activity of AMF-inoculated plants were significantly higher than that of the non-AMF control. The H2O2 accumulation in AMF-inoculated roots was decreased by 42.44% compared with the control under LT. H2O2 predominantly accumulated on the cell walls of apoplast but was hardly detectable in the cytosol or organelles of roots. Again, NADPH oxidase activity involved in H2O2 production was significantly reduced by AMF inoculation under LT. AMF-inoculation remarkably increased the activities of P-type H(+)-ATPase, P-Ca(2+)-ATPase, V-type H(+)-ATPase, total ATPase activity, ATP concentration and plasma membrane protein content in the roots under LT. Additionally, ATP concentration and expression of plasma membrane ATPase genes were increased by AMF-inoculation. These results indicate that NADPH oxidase and ATPase might play an important role in AMF-mediated tolerance to chilling stress, thereby maintaining a lower H2O2 accumulation in the roots of cucumber.

  3. Radionuclide activities, geochemistry, and accumulation rates of sediments in the Gulf of Thailand

    NASA Astrophysics Data System (ADS)

    Srisuksawad, Kanitha; Porntepkasemsan, Boonsom; Nouchpramool, Sunun; Yamkate, Pathom; Carpenter, Roy; Peterson, Michael L.; Hamilton, Terry

    1997-07-01

    Downcore concentration profiles of 210Pb, U, and Th isotopes, Al, Fe, Ti, Mn and Sc were measured in sediment box cores collected at 22 stations (16-70 m water depth) covering most of the Thai zone of the Gulf of Thailand. Distributions of excess 210Pb and the detrital elements were used to study spatial variations in sedimentary processes, mineralogy, and geochemistry between different regions of the gulf. Steady-state depositional concentrations and fluxes of excess 210Pb are 3-10 times lower in Gulf of Thailand sediments than in sediments from mid-latitudes in the northern hemisphere, reflecting lower 210Pb inputs from atmospheric fallout at 6-13°N latitude and from lower production of 210Pb from 226Ra in the shallower waters of the Gulf. U and Th concentrations are approximately 2-3 times higher than those in shelf sediments from mid-latitudes of North America, consistent with a higher proportion of granitic source rocks in the Thai environment. Downcore variations in 228Th/ 232Th activity ratios and in U activities reveal that exchange of interstitial and overlying waters and their dissolved chemicals occurs down to 20 cm in 8 of 10 cores. This benthic exchange may be important in budgets of fluxes of other soluble chemicals in this shallow shelf sea. A net flux of U isotopes from overlying water into Gulf of Thailand sediments occurs, in contrast to their release from sediments of the tropical Amazon shelf. Detectable levels of 137Cs were found only in sediments near the mouth of the largest river, the Chao Phraya. The detrital elements 232Th, 230Th, Al, Ti, and Sc all show relatively uniform downcore concentration profiles. This supports a key assumption in calculations of sediment accumulation rates from downcore profiles of 210Pb activity, that steady-state depositional conditions exist and that basic sediment mineralogy and grain size does not change. 210Pb model derived mass accumulation rates vary between 270 and 490 mg/cm 2 per year in the upper Gulf

  4. EVALUATION OF THIOL-BASED ANTIOXIDANT THERAPEUTICS IN CYSTIC FIBROSIS SPUTUM: FOCUS ON MYELOPEROXIDASE

    PubMed Central

    Vasu, Vihas T.; De Cruz, Sharon J.; Houghton, Jessica S.; Hayakawa, Keri A.; Morrissey, Brian M.; Cross, Carroll E.; Eiserich, Jason P.

    2010-01-01

    Neutrophil-dependent reactions catalyzed by myeloperoxidase (MPO) are thought to play important roles in the pulmonary pathobiology of cystic fibrosis (CF). Aerosolized thiol antioxidants such as glutathione (GSH) and N-acetylcysteine (NAC) are currently being utilized as therapeutics to modify CF respiratory tract oxidative processes. We hypothesized that MPO in CF airway lining fluids may be a target of such therapeutics. MPO activity in sputum from 21 adult CF patients was found to be inversely associated with lung function (FEV1). In contrast, systemic inflammation (assessed by plasma C-reactive protein) was not correlated with lung function. Ex vivo studies revealed that GSH and NAC effectively scavenged N-chloramines in sputum, and inhibited sputum MPO activity with potency exquisitely dependent upon MPO activity levels. Detailed kinetic analyses revealed that NAC and GSH inhibit MPO by distinct mechanisms. Activation of the key proinflammatory transcription factor NF-κB in cultured HBE1 cells was inhibited by GSH. Our findings reveal that MPO activity and its reactive products represent useful predictors of the doses of inhaled thiol antioxidants required to ameliorate airway oxidative stress and inflammation in CF patients, and provides mechanistic insight into the antioxidative/anti-inflammatory mechanisms of action of GSH and NAC when administered into the CF lung. PMID:20954832

  5. Ammonia activates pacC and patulin accumulation in an acidic environment during apple colonization by Penicillium expansum.

    PubMed

    Barad, Shiri; Espeso, Eduardo A; Sherman, Amir; Prusky, Dov

    2016-06-01

    Penicillium expansum, the causal agent of blue mould rot, causes severe post-harvest fruit maceration simultaneously with the secretion of d-gluconic acid (GLA) and the mycotoxin patulin in colonized tissue. The factor(s) inducing patulin biosynthesis during colonization of the host acidic environment is unclear. During the colonization of apple fruit in vivo and growth in culture, P. expansum secretes pH-modulating GLA and ammonia. Although patulin and its possible opportunistic precursor GLA accumulate together during fungal development, ammonia is detected on the colonized tissue's leading edge and after extended culture, close to patulin accumulation. Here, we demonstrate ammonia-induced transcript activation of the global pH modulator PacC and patulin accumulation in the presence of GLA by: (i) direct exogenous treatment of P. expansum growing on solid medium; (ii) direct exogenous treatment on colonized apple tissue; (iii) growth under self-ammonia production conditions with limited carbon; and (iv) analysis of the transcriptional response to ammonia of the patulin biosynthesis cluster. Ammonia induced patulin accumulation concurrently with the transcript activation of pacC and patulin biosynthesis cluster genes, indicating the regulatory effect of ammonia on pacC transcript expression under acidic conditions. Electrophoretic mobility shift assays using P. expansum PacC and antibodies to the different cleaved proteins showed that PacC is not protected against proteolytic signalling at pH 4.5 relative to pH 7.0, but NH4 addition did not further enhance its proteolytic cleavage. Ammonia enhanced the activation of palF transcript in the Pal pathway under acidic conditions. Ammonia accumulation in the host environment by the pathogen under acidic pH may be a regulatory cue for pacC activation, towards the accumulation of secondary metabolites, such as patulin.

  6. Myeloperoxidase-Oxidized LDLs Enhance an Anti-Inflammatory M2 and Antioxidant Phenotype in Murine Macrophages

    PubMed Central

    Sauvage, Aude; Van Steenbrugge, Martine

    2016-01-01

    Macrophages and oxidized LDLs play a key role in atherogenesis but their heterogeneity has been neglected up to now. Macrophages are prone to polarization and subsets of polarized macrophages have been described in atheromas. LDLs can be oxidized not only chemically by copper (Ox-LDLs) but also enzymatically by myeloperoxidase (MpOx-LDLs) resulting in oxidized LDLs poor in lipid peroxides. The effects of physiologically relevant myeloperoxidase-oxidized LDLs on macrophage polarization or on polarized macrophages remain largely unknown. In this study, the effects of LDLs on macrophage polarization were investigated by monitoring the expression of M1 and M2 genes following stimulation with native LDLs, Ox-LDLs, or MpOx-LDLs in RAW 264.7 cells. Except for MRC1, which is induced only by Ox-LDLs, MpOx-LDLs induced an overexpression of most of the selected marker genes at the mRNA level. MpOx-LDLs also modulate marker gene expression in polarized macrophages favoring notably anti-inflammatory Arg1 expression in M2 cells and also in the other phenotypes. Noteworthy, MpOx-LDLs were the most efficient to accumulate lipids intracellularly in (un)polarized macrophages whatever the phenotype. These data were largely confirmed in murine bone marrow-derived macrophages. Our data suggest that MpOx-LDLs were the most efficient to accumulate within cells and to enhance an anti-inflammatory and antioxidant phenotype in M2 cells and also in the other macrophage phenotypes. PMID:27656049

  7. The reactivity of myeloperoxidase compound I formed with hypochlorous acid.

    PubMed

    Furtmüller, P G; Burner, U; Jantschko, W; Regelsberger, G; Obinger, C

    2000-01-01

    The reaction of human myeloperoxidase (MPO) with hypochlorous acid (HOCl) was investigated by conventional stopped-flow spectroscopy at pH 5, 7, and 9. In the reaction of MPO with HOCl, compound I is formed. Its formation is strongly dependent on pH. HOCl (rather than OCl-) reacts with the unprotonated enzyme in its ferric state. Apparent second-order rate constants were determined to be 8.1 x 10(7) M(-1)s(-1) (pH 5), 2.0 x 10(8) M(-1)s(-1) (pH 7) and 2.0 x 10(6) M(-1)s(-1) (pH 9) at 15 degrees C. Furthermore, the kinetics and spectra of the reactions of halides and thiocyanate and of physiologically relevant one-electron donors (ascorbate, nitrite, tyrosine and hydrogen peroxide) with this compound I were investigated using the sequential-mixing technique. The results show conclusively that the redox intermediates formed upon addition of either hydrogen peroxide or hypochlorous acid to native MPO exhibit the same spectral features and reactivities and thus are identical. In stopped-flow investigations, the MPO/HOCl system has some advantage since: (i) in contrast to H2O2, HOCl cannot function as a one-electron donor of compound I; and (ii) MPO can easily be prevented from cycling by addition of methionine as HOCl scavenger. As a consequence, the observed absorbance changes are bigger and errors in data analysis are smaller.

  8. Myeloperoxidase: expressing inflammation and oxidative stress in cardiovascular disease.

    PubMed

    Anatoliotakis, Nikolaos; Deftereos, Spyridon; Bouras, Georgios; Giannopoulos, Georgios; Tsounis, Dimitrios; Angelidis, Christos; Kaoukis, Andreas; Stefanadis, Christodoulos

    2013-01-01

    Myeloperoxidase (MPO), a heme protein released by leukocytes, is one of the most widely studied during the last decades molecule that plays a crucial role in inflammation and oxidative stress in the cellular level. It has become increasingly recognized that MPO performs a very important role as part of the innate immune system through the formation of microbicidal reactive oxidants, whilst it affects the arterial endothelium with a number of mechanisms that include modification of net cellular cholesterol flux and impairment of Nitric Oxide (NO)-induced vascular relaxation. In that way, MPO is implicated into both the formation and propagation of atheromatosis and there is substantial evidence that it also promotes ischemia through destabilization of the vulnerable plaque. Numerous studies have added information on the notion that MPO and its oxidant products are part of the inflammatory cascade initiated by endothelial injury and they are significantly overproduced at the site of arterial inflammation. Subsequent studies achieved quantification of this observation showing significant elevations of the systemic levels of MPO in a wide spectrum of cardiovascular disease scenarios with acute coronary syndromes and heart failure being the most studied. This review highlights key-aspects of MPO's pathophysiological properties and summarizes the role of MPO as a diagnostic and prognostic tool for a number of cardiovascular pathologies.

  9. Myeloperoxidase-positive acute megakaryoblastic leukemia in a dog.

    PubMed

    Ferreira, Helena M T; Smith, Sionagh H; Schwartz, Anita M; Milne, Elspeth M

    2011-12-01

    A 16-month-old female spayed Labrador Retriever was referred to the University of Edinburgh for exercise intolerance, inappetence, and severe anemia. A CBC showed severe nonregenerative anemia and moderate numbers of atypical cells with morphologic features most consistent with megakaryoblastic origin. Similar cells were identified in a bone marrow aspirate and accounted for 23% of all nucleated cells. Atypical promegakaryocytes and megakaryocytes were also noted. Myelodysplastic syndrome affecting the megakaryocytic lineage was suspected. Cytologic examination of a fine-needle aspirate of the spleen revealed rare megakaryoblasts similar to those in blood and bone marrow. At necropsy, the bone marrow consisted of atypical megakaryoblasts and megakaryocytes that were also infiltrating spleen, liver, lymph nodes, renal perihilar tissue, and visceral adipose tissue, consistent with acute megakaryoblastic leukemia. Immunohistochemical analysis of splenic sections confirmed megakaryoblastic origin (immunoreactive for CD61 and von Willebrand factor). Some leukemic cells were also immunoreactive for myeloperoxidase (MPO). This aberrant immunophenotype suggested both megakaryocytic and granulocytic/monocytic differentiation of the leukemic cells. To our knowledge, this is the first report of MPO-positive acute megakaryoblastic leukemia in a dog.

  10. Proteinase 3-ANCA Vasculitis versus Myeloperoxidase-ANCA Vasculitis

    PubMed Central

    Hilhorst, Marc; van Paassen, Pieter

    2015-01-01

    In patients with GN or vasculitis, ANCAs are directed against proteinase 3 (PR3) or myeloperoxidase (MPO). The differences between PR3-ANCA-associated vasculitis (AAV) and MPO-AAV described in the past have been supplemented during the last decade. In this review, we discuss the differences between these two small-vessel vasculitides, focusing especially on possible etiologic and pathophysiologic differences. PR3-AAV is more common in northern parts of the world, whereas MPO-AAV is more common in southern regions of Europe, Asia, and the Pacific, with the exception of New Zealand and Australia. A genetic contribution has been extensively studied, and there is a high prevalence of the HLA-DPB1*04:01 allele in patients with PR3-AAV as opposed to patients with MPO-AAV and/or healthy controls. Histologically, MPO-AAV and PR3-AAV are similar but show qualitative differences when analyzed carefully. Clinically, both serotypes are difficult to distinguish, but quantitative differences are present. More organs are affected in PR3-AAV, whereas renal limited vasculitis occurs more often in patients with MPO-AAV. For future clinical trials, we advocate classifying patients by ANCA serotype as opposed to the traditional disease type classification. PMID:25956510

  11. Ethylene formation by polymorphonuclear leukocytes. Role of myeloperoxidase

    PubMed Central

    1978-01-01

    Ethylene formation from the thioethers, beta-methylthiopropionaldehyde (methional) and 2-keto-4-thiomethylbutyric acid by phagocytosing polymorphonuclear leukocytes (PMNs) was found to be largely dependent on myeloperoxidase (MPO). Conversion was less than 10% of normal when MPO-deficient PMNs were employed; formation by normal PMNs was inhibited by the peroxidase inhibitors, azide, and cyanide, and a model system consisting of MPO, H2O2, chloride (or bromide) and EDTA was found which shared many of the properties of the predominant PMN system. MPO-independent mechanisms of ethylene formation were also identified. Ethylene formation from methional by phagocytosing eosinophils and by H2O2 in the presence or absence of catalase was stimulated by azide. The presence of MPO-independent, azide-stimulable systems in the PMN preparations was suggested by the azide stimulation of ethylene formation from methional when MPO-deficient leukocytes were employed. Ethylene formation by dye-sensitized photooxidation was also demonstrated and evidence obtained for the involvement of singlet oxygen (1O2). These findings are discussed in relation to the participation of H2O2, hydroxyl radicals, the superoxide anion and 1O2 in the formation of ethylene by PMNs and by the MPO model system. PMID:212502

  12. Active hydrocarbon biosynthesis and accumulation in a green alga, Botryococcus braunii (race A).

    PubMed

    Hirose, Mana; Mukaida, Fukiko; Okada, Sigeru; Noguchi, Tetsuko

    2013-08-01

    Among oleaginous microalgae, the colonial green alga Botryococcus braunii accumulates especially large quantities of hydrocarbons. This accumulation may be achieved more by storage of lipids in the extracellular space rather than in the cytoplasm, as is the case for all other examined oleaginous microalgae. The stage of hydrocarbon synthesis during the cell cycle was determined by autoradiography. The cell cycle of B. braunii race A was synchronized by aminouracil treatment, and cells were taken at various stages in the cell cycle and cultured in a medium containing [(14)C]acetate. Incorporation of (14)C into hydrocarbons was detected. The highest labeling occurred just after septum formation, when it was about 2.6 times the rate during interphase. Fluorescent and electron microscopy revealed that new lipid accumulation on the cell surface occurred during at least two different growth stages and sites of cells. Lipid bodies in the cytoplasm were not prominent in interphase cells. These lipid bodies then increased in number, size, and inclusions, reaching maximum values just before the first lipid accumulation on the cell surface at the cell apex. Most of them disappeared from the cytoplasm concomitant with the second new accumulation at the basolateral region, where extracellular lipids continuously accumulated. The rough endoplasmic reticulum near the plasma membrane is prominent in B. braunii, and the endoplasmic reticulum was often in contact with both a chloroplast and lipid bodies in cells with increasing numbers of lipid bodies. We discuss the transport pathway of precursors of extracellular hydrocarbons in race A.

  13. Results of Topical Myeloperoxidase/Glucose Oxidase/Glucose in the Walker-Mason Burn Model

    DTIC Science & Technology

    2005-12-01

    The system consists of an enzyme, myeloperoxidase (MEPO) and the requisite substrates which have been optimized for the production of bactericidal ...consists of an enzyme, myeloperoxidase (MPO) and the requisite substrates which have been optimized for the production of bactericidal reactive oxygen...engineered to luminesce. The bacterial infection is monitored by non -invasive bioluminescent imaging as a means to assess the animals’ infection

  14. Neutrophil- and myeloperoxidase-mediated metabolism of reduced nimesulide: evidence for bioactivation.

    PubMed

    Yang, Min; Chordia, Mahendra D; Li, Fengping; Huang, Tao; Linden, Joel; Macdonald, Timothy L

    2010-11-15

    Nimesulide, a widely used nonsteroidal anti-inflammatory drug (NSAID), has been associated with rare idiosyncratic hepatotoxicity. The chemical mechanisms underlying the liver injury remain unknown. We have undertaken the detailed study of the metabolic pathways of nimesulide in an effort to identify potential reactive metabolites. A previous report from this laboratory has demonstrated that one of the known nimesulide metabolites, termed reduced nimesulide (M1), is further bioactivated by human liver microsomes (HLMs) to form a reactive diiminoquinone species M2. The formation of M2 was confirmed indirectly by trapping with N-acetylcysteine (NAC). The aim of this study was to explore the fate of M1 in an inflammatory environment created by the recruitment of leukocytes. Leukocytes upon activation produce hydrogen peroxide (H(2)O(2)) and other myeloperoxidase (MPO) products, such as hypochlorous acid (HOCl), that are capable of metabolite oxidation. We demonstrate here that the reduced nimesulide, M1, undergoes a facile oxidation with activated neutrophils or with MPO in the presence of H(2)O(2) or HOCl to produce a variety of reactive as well as stable metabolites. One major metabolite, M3, was also produced by HLM as determined by trapping with NAC. Other metabolites, for example, M6, M8, and M9, were unique to the myeloperoxidase, because of their mode of formation from activation of the amino group of reduced nimesulide. The structures of some of these reactive metabolites were proposed on the basis of liquid chromatography-tandem mass spectrometry analyses and established by their comparison with synthetic standards. Metabolite M6 is interesting because it provides clear evidence of amine activation and indicates the potential of the reactive intermediate of M1 to conjugate with protein nucleophiles. In summary, our results demonstrate that a known nimesulide metabolite could be bioactivated by MPO through a pathway distinct from HLM-mediated pathways and that

  15. The Role of Neutrophil Myeloperoxidase in Models of Lung Tumor Development

    PubMed Central

    Rymaszewski, Amy L.; Tate, Everett; Yimbesalu, Joannes P.; Gelman, Andrew E.; Jarzembowski, Jason A.; Zhang, Hao; Pritchard, Kirkwood A.; Vikis, Haris G.

    2014-01-01

    Chronic inflammation plays a key tumor-promoting role in lung cancer. Our previous studies in mice demonstrated that neutrophils are critical mediators of tumor promotion in methylcholanthrene (MCA)-initiated, butylated hydroxytoluene (BHT)-promoted lung carcinogenesis. In the present study we investigated the role of neutrophil myeloperoxidase (MPO) activity in this inflammation promoted model. Increased levels of MPO protein and activity were present in the lungs of mice administered BHT. Treatment of mice with N-acetyl lysyltyrosylcysteine amide (KYC), a novel tripeptide inhibitor of MPO, during the inflammatory stage reduced tumor burden. In a separate tumor model, KYC treatment of a Lewis Lung Carcinoma (LLC) tumor graft in mice had no effect on tumor growth, however, mice genetically deficient in MPO had significantly reduced LLC tumor growth. Our observations suggest that MPO catalytic activity is critical during the early stages of tumor development. However, during the later stages of tumor progression, MPO expression independent of catalytic activity appears to be required. Our studies advocate for the use of MPO inhibitors in a lung cancer prevention setting. PMID:24821130

  16. Apoptosis induced by capsaicin in prostate PC-3 cells involves ceramide accumulation, neutral sphingomyelinase, and JNK activation.

    PubMed

    Sánchez, Ana Maria; Malagarie-Cazenave, Sophie; Olea, Nuria; Vara, Diana; Chiloeches, Antonio; Díaz-Laviada, Inés

    2007-11-01

    Numerous studies have recently focused on the anticarcinogenic, antimutagenic, or chemopreventive activities of the main pungent component of red pepper, capsaicin (N-vanillyl-8-methyl-1-nonenamide). We have previously shown that, in the androgen-independent prostate cancer PC-3 cells, capsaicin inhibits cell growth and induces apoptosis through reactive oxygen species (ROS) generation [Apoptosis 11 (2006) 89-99]. In the present study, we investigated the signaling pathways involved in the antiproliferative effect of capsaicin. Here, we report that capsaicin apoptotic effect was mediated by ceramide generation which occurred by sphingomyelin hydrolysis. Using siRNA, we demonstrated that N-SMase expression is required for the effect of capsaicin on prostate cell viability. We then investigated the role of MAP kinase cascades, extracellular signal-regulated protein kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 MAPK, in the antiproliferative effect of capsaicin, and we confirmed that capsaicin could activate ERK and JNK but not p38 MAPK. Pharmacological inhibition of JNK kinase, as well as inhibition of ROS by the reducing agent N-acetylcysteine, prevented ceramide accumulation and capsaicin-induced cell death. However, inhibition of ceramide accumulation by the SMase inhibitor D609 did not modify JNK activation. These data reveal JNK as an upstream regulator of ceramide production. Capsaicin-promoted activation of ERK was prevented with all the inhibitors tested. We conclude that capsaicin induces apoptosis in PC-3 cells via ROS generation, JNK activation, ceramide accumulation, and second, ERK activation.

  17. Cholesteryl Ester Accumulation Induced by PTEN Loss and PI3K/AKT Activation Underlies Human Prostate Cancer Aggressiveness

    PubMed Central

    Yue, Shuhua; Li, Junjie; Lee, Seung-Young; Lee, Hyeon Jeong; Shao, Tian; Song, Bing; Cheng, Liang; Masterson, Timothy A.; Liu, Xiaoqi; Ratliff, Timothy L.; Cheng, Ji-Xin

    2014-01-01

    Summary Altered lipid metabolism is increasingly recognized as a signature of cancer cells. Enabled by label-free Raman spectromicroscopy, we performed quantitative analysis of lipogenesis at single cell level in human patient cancerous tissues. Our imaging data revealed an unexpected, aberrant accumulation of esterified cholesterol in lipid droplets of high-grade prostate cancer and metastases. Biochemical study showed that such cholesteryl ester accumulation was a consequence of loss of tumor suppressor PTEN and subsequent activation of PI3K/AKT pathway in prostate cancer cells. Furthermore, we found that such accumulation arose from significantly enhanced uptake of exogenous lipoproteins and required cholesterol esterification. Depletion of cholesteryl ester storage significantly reduced cancer proliferation, impaired cancer invasion capability, and suppressed tumor growth in mouse xenograft models with negligible toxicity. These findings open opportunities for diagnosing and treating prostate cancer by targeting the altered cholesterol metabolism. PMID:24606897

  18. Infection-induced type I interferons activate CD11b on B-1 cells for subsequent lymph node accumulation

    PubMed Central

    Waffarn, Elizabeth E.; Hastey, Christine J.; Dixit, Neha; Choi, Youn Soo; Cherry, Simon; Kalinke, Ulrich; Simon, Scott I.; Baumgarth, Nicole

    2016-01-01

    Innate-like B-1a lymphocytes rapidly redistribute to regional mediastinal lymph nodes (MedLN) during influenza infection to generate protective IgM. Here we demonstrate that influenza infection-induced type I interferons directly stimulate body cavity B-1 cells and are a necessary signal required for B-1 cell accumulation in MedLN. Vascular mimetic flow chamber studies show that type I interferons increase ligand-mediated B-1 cell adhesion under shear stress by inducing high-affinity conformation shifts of surface-expressed integrins. In vivo trafficking experiments identify CD11b as the non-redundant, interferon-activated integrin required for B-1 cell accumulation in MedLN. Thus CD11b on B-1 cells senses infection-induced innate signals and facilitates their rapid sequester into secondary lymphoid tissues, thereby regulating the accumulation of polyreactive IgM producers at sites of infection. PMID:26612263

  19. Plasma myeloperoxidase is inversely associated with endothelium-dependent vasodilation in elderly subjects with abnormal glucose metabolism.

    PubMed

    van der Zwan, Leonard P; Teerlink, Tom; Dekker, Jacqueline M; Henry, Ronald M A; Stehouwer, Coen D A; Jakobs, Cornelis; Heine, Robert J; Scheffer, Peter G

    2010-12-01

    Myeloperoxidase (MPO), a biomarker related to inflammation, oxidative stress, and nitric oxide scavenging, has been shown to impair endothelium-dependent vasodilation. Because elevated hydrogen peroxide concentrations in diabetic vessels may enhance MPO activity, we hypothesized that a stronger association of MPO with flow-mediated dilation (FMD) may be found in subjects with abnormal glucose metabolism. Myeloperoxidase concentrations were measured in EDTA plasma samples from participants of a population-based cohort study, including 230 subjects with normal glucose metabolism and 386 with abnormal glucose metabolism. Vascular function was expressed as FMD and nitroglycerin-mediated dilation of the brachial artery. In subjects with abnormal glucose metabolism, MPO was negatively associated with FMD (-20.9 [95% confidence interval {CI}, -41.7 to -0.2] -μm change in FMD per SD increment of MPO). This association remained significant after adjustment for nitroglycerin-mediated dilation (-31.1 [95% CI, -50.0 to -12.3]) and was not attenuated after further adjustment for established risk factors. In subjects with normal glucose metabolism, MPO was not significantly associated with FMD (2.0 [95% CI, -16.0 to 20.0]). In conclusion, in subjects with abnormal glucose metabolism, plasma levels of MPO are inversely associated with endothelium-dependent vasodilation, possibly reflecting enhancement of MPO activity by vascular oxidative stress.

  20. Potent inhibition of macrophage migration inhibitory factor (MIF) by myeloperoxidase-dependent oxidation of epicatechins.

    PubMed

    Dickerhof, Nina; Magon, Nicholas J; Tyndall, Joel D A; Kettle, Anthony J; Hampton, Mark B

    2014-09-01

    MIF (macrophage migration inhibitory factor) plays a central role in the promotion and maintenance of the inflammatory response. It is implicated in a number of inflammatory diseases including sepsis, arthritis and colitis, and in diseases with an inflammatory component, such as atherosclerosis, diabetes and cancer. MIF has an unusual N-terminal proline with catalytic activity, and targeting of this residue by small-molecule inhibitors has been shown to interfere with the biological activity of MIF. The objective of the present study was to determine if MIF was susceptible to modification by epicatechins, a group of dietary flavonoids with known anti-inflammatory properties. Epicatechins are substrates for peroxidases including neutrophil-derived MPO (myeloperoxidase). In the present study we show that oxidation of the catechol moiety of epicatechins to an ο-quinone by MPO generates potent MIF inhibitors. Near complete inhibition of MIF by the MPO/H2O2/epicatechin system was achieved at equimolar concentrations of epicatechin and MIF, even in the presence of other MPO substrates. We have characterized the modification introduced by oxidized (-)-epicatechin on MIF by LC-MS (liquid chromatography MS) and found it to occur at the N-terminal proline. We propose that MIF inhibition by oxidized epicatechins contributes to the anti-inflammatory activity of these compounds.

  1. Myeloperoxidase propagates damage and is a potential therapeutic target for subacute stroke

    PubMed Central

    Forghani, Reza; Kim, Hyeon Ju; Wojtkiewicz, Gregory R; Bure, Lionel; Wu, Yue; Hayase, Makoto; Wei, Ying; Zheng, Yi; Moskowitz, Michael A; Chen, John W

    2015-01-01

    Few effective treatment options exist for stroke beyond the hyperacute period. Radical generation and myeloperoxidase (MPO) have been implicated in stroke. We investigated whether pharmacologic reduction or gene deletion of this highly oxidative enzyme reduces infarct propagation and improves outcome in the transient middle cerebral artery occlusion mouse model (MCAO). Mice were treated with 4-aminobenzoic acid hydrazide (ABAH), a specific irreversible MPO inhibitor. Three treatment regimens were used: (1) daily throughout the 21-day observational period, (2) during the acute stage (first 24 hours), or (3) during the subacute stage (daily starting on day 2). We found elevated MPO activity in ipsilateral brain 3 to 21 days after ischemia. 4-Aminobenzoic acid hydrazide reduced enzyme activity by 30% to 40% and final lesion volume by 60% (P<0.01). The MPO-knockout (KO) mice subjected to MCAO also showed a similar reduction in the final lesion volume (P<0.01). The ABAH treatment or MPO-KO mice also improved neurobehavioral outcome (P<0.001) and survival (P=0.01), but ABAH had no additional beneficial effects in MPO-KO mice, confirming specificity of ABAH. Interestingly, inhibiting MPO activity during the subacute stage recapitulated most of the therapeutic benefit of continuous MPO inhibition, suggesting that MPO-targeted therapies could be useful when given after 24 hours of stroke onset. PMID:25515211

  2. Myeloperoxidase propagates damage and is a potential therapeutic target for subacute stroke.

    PubMed

    Forghani, Reza; Kim, Hyeon Ju; Wojtkiewicz, Gregory R; Bure, Lionel; Wu, Yue; Hayase, Makoto; Wei, Ying; Zheng, Yi; Moskowitz, Michael A; Chen, John W

    2015-03-01

    Few effective treatment options exist for stroke beyond the hyperacute period. Radical generation and myeloperoxidase (MPO) have been implicated in stroke. We investigated whether pharmacologic reduction or gene deletion of this highly oxidative enzyme reduces infarct propagation and improves outcome in the transient middle cerebral artery occlusion mouse model (MCAO). Mice were treated with 4-aminobenzoic acid hydrazide (ABAH), a specific irreversible MPO inhibitor. Three treatment regimens were used: (1) daily throughout the 21-day observational period, (2) during the acute stage (first 24 hours), or (3) during the subacute stage (daily starting on day 2). We found elevated MPO activity in ipsilateral brain 3 to 21 days after ischemia. 4-Aminobenzoic acid hydrazide reduced enzyme activity by 30% to 40% and final lesion volume by 60% (P<0.01). The MPO-knockout (KO) mice subjected to MCAO also showed a similar reduction in the final lesion volume (P<0.01). The ABAH treatment or MPO-KO mice also improved neurobehavioral outcome (P<0.001) and survival (P=0.01), but ABAH had no additional beneficial effects in MPO-KO mice, confirming specificity of ABAH. Interestingly, inhibiting MPO activity during the subacute stage recapitulated most of the therapeutic benefit of continuous MPO inhibition, suggesting that MPO-targeted therapies could be useful when given after 24 hours of stroke onset.

  3. Bioactivation of myelotoxic xenobiotics by human neutrophil myeloperoxidase

    SciTech Connect

    Roy, R.R.

    1989-01-01

    Many environmental pollutants and drugs are toxic to the bone marrow. Some of these xenobiotics may initiate toxicity after undergoing bioactivation to free radicals and/or other reactive electrophiles. Peroxidases are a group of enzymes that catalyze the one-electron oxidative bioactivation of a variety of xenobiotics in vitro. Myeloperoxidase (MPO) is a peroxidative enzyme found in very high concentration in the neutrophils of human bone marrow. In this study, human MPO was evaluated to determine its ability to catalyze the in vitro bioactivation of known bone marrow toxicants that contain the aromatic hydroxyl (Ar-OH), aromatic amine (Ar-N-R{sub 2}), or heterocyclic tertiary amine ({double bond}N-R) moieties. The formation of free radical metabolites during the MPO-catalyzed bioactivation of hydroquinone and catechol (benzene metabolites), mitoxantrone and ametantrone (antitumor drugs), and chlorpromazine and promazine (antipsychotic drugs) was demonstrated by EPR spectroscopy. The reactivity of the products formed during the MPO catalyzed bioactivation of ({sup 14}C)hydroquinone and ({sup 14}C)catechol was shown by their covalent binding to protein and DNA in vitro. The covalently binding metabolite in each case is postulated to be the quinone form of the xenobiotic. In addition, both GSH and NADH were oxidized by the reactive intermediate(s) formed during the MPO-catalyzed bioactivation of many of the bone marrow toxicants tested. It was also shown that p,p-biphenol stimulated the MPO catalyzed bioactivation of both hydroquinone and catechol, while p-cresol stimulated the MPO-catalyzed bioactivation of catechol.

  4. Dietary fructose feeding increases adipose methylglyoxal accumulation in rats in association with low expression and activity of glyoxalase-2.

    PubMed

    Masterjohn, Christopher; Park, Youngki; Lee, Jiyoung; Noh, Sang K; Koo, Sung I; Bruno, Richard S

    2013-08-21

    Methylglyoxal is a precursor to advanced glycation endproducts that may contribute to diabetes and its cardiovascular-related complications. Methylglyoxal is successively catabolized to D-lactate by glyoxalase-1 and glyoxalase-2. The objective of this study was to determine whether dietary fructose and green tea extract (GTE) differentially regulate methylglyoxal accumulation in liver and adipose, mediated by tissue-specific differences in the glyoxalase system. We fed six week old male Sprague-Dawley rats a low-fructose diet (10% w/w) or a high-fructose diet (60% w/w) containing no GTE or GTE at 0.5% or 1.0% for nine weeks. Fructose-fed rats had higher (P < 0.05) adipose methylglyoxal, but GTE had no effect. Plasma and hepatic methylglyoxal were unaffected by fructose and GTE. Fructose and GTE also had no effect on the expression or activity of glyoxalase-1 and glyoxalase-2 at liver or adipose. Regardless of diet, adipose glyoxalase-2 activity was 10.8-times lower (P < 0.05) than adipose glyoxalase-1 activity and 5.9-times lower than liver glyoxalase-2 activity. Adipose glyoxalase-2 activity was also inversely related to adipose methylglyoxal (r = -0.61; P < 0.05). These findings suggest that fructose-mediated adipose methylglyoxal accumulation is independent of GTE supplementation and that its preferential accumulation in adipose compared to liver is due to low constitutive expression of glyoxalase-2.

  5. Prelamin A Accumulation Attenuates Rac1 Activity and Increases the Intrinsic Migrational Persistence of Aged Vascular Smooth Muscle Cells

    PubMed Central

    Porter, Lauren J.; Holt, Mark R.; Soong, Daniel; Shanahan, Catherine M.; Warren, Derek T.

    2016-01-01

    Vascular smooth muscle cell (VSMC) motility is essential during both physiological and pathological vessel remodeling. Although ageing has emerged as a major risk factor in the development of cardiovascular disease, our understanding of the impact of ageing on VSMC motility remains limited. Prelamin A accumulation is known to drive VSMC ageing and we show that presenescent VSMCs, that have accumulated prelamin A, display increased focal adhesion dynamics, augmented migrational velocity/persistence and attenuated Rac1 activity. Importantly, prelamin A accumulation in proliferative VSMCs, induced by depletion of the prelamin A processing enzyme FACE1, recapitulated the focal adhesion, migrational persistence and Rac1 phenotypes observed in presenescent VSMCs. Moreover, lamin A/C-depleted VSMCs also display reduced Rac1 activity, suggesting that prelamin A influences Rac1 activity by interfering with lamin A/C function at the nuclear envelope. Taken together, these data demonstrate that lamin A/C maintains Rac1 activity in VSMCs and prelamin A disrupts lamin A/C function to reduce Rac1 activity and induce migrational persistence during VSMC ageing. PMID:27854297

  6. Neuronal chloride accumulation and excitatory GABA underlie aggravation of neonatal epileptiform activities by phenobarbital.

    PubMed

    Nardou, Romain; Yamamoto, Sumii; Chazal, Geneviève; Bhar, Asma; Ferrand, Nadine; Dulac, Olivier; Ben-Ari, Yehezkel; Khalilov, Ilgam

    2011-04-01

    Phenobarbital produces its anti-epileptic actions by increasing the inhibitory drive of γ-aminobutyric acid. However, following recurrent seizures, γ-aminobutyric acid excites neurons because of a persistent increase of chloride raising the important issue of whether phenobarbital could aggravate persistent seizures. Here we compared the actions of phenobarbital on initial and established ictal-like events in an in vitro model of mirror focus. Using the in vitro three-compartment chamber preparation with the two hippocampi and their commissural fibres placed in three different chambers, kainate was applied to one hippocampus and phenobarbital contralaterally, either after one ictal-like event or after many recurrent ictal-like events that produce an epileptogenic mirror focus. Field, perforated patch and single-channel recordings were used to determine the effects of γ-aminobutyric acid and their modulation by phenobarbital, and alterations of the chloride cotransporters were investigated using sodium-potassium-chloride cotransporter 1 and potassium chloride cotransporter 2 antagonists, potassium chloride cotransporter 2 immunocytochemistry and sodium-potassium-chloride cotransporter 1 knockouts. Phenobarbital reduced initial ictal-like events and prevented the formation of a mirror focus when applied from the start. In contrast, phenobarbital aggravated epileptiform activities when applied after many ictal-like events by enhancing the excitatory actions of γ-aminobutyric acid due to increased chloride. The accumulation of chloride and the excitatory actions of γ-aminobutyric acid in mirror foci neurons are mediated by the sodium-potassium-chloride cotransporter 1 chloride importer and by downregulation and internalization of the chloride-exporter potassium-chloride cotransporter 2. Finally, concomitant applications of the sodium-potassium-chloride cotransporter 1 antagonist bumetanide and phenobarbital decreased excitatory actions of γ-aminobutyric acid and

  7. Activity-dependent synaptic GRIP1 accumulation drives synaptic scaling up in response to action potential blockade.

    PubMed

    Gainey, Melanie A; Tatavarty, Vedakumar; Nahmani, Marc; Lin, Heather; Turrigiano, Gina G

    2015-07-07

    Synaptic scaling is a form of homeostatic plasticity that stabilizes neuronal firing in response to changes in synapse number and strength. Scaling up in response to action-potential blockade is accomplished through increased synaptic accumulation of GluA2-containing AMPA receptors (AMPAR), but the receptor trafficking steps that drive this process remain largely obscure. Here, we show that the AMPAR-binding protein glutamate receptor-interacting protein-1 (GRIP1) is essential for regulated synaptic AMPAR accumulation during scaling up. Synaptic abundance of GRIP1 was enhanced by activity deprivation, directly increasing synaptic GRIP1 abundance through overexpression increased the amplitude of AMPA miniature excitatory postsynaptic currents (mEPSCs), and shRNA-mediated GRIP1 knockdown prevented scaling up of AMPA mEPSCs. Furthermore, knockdown and replace experiments targeting either GRIP1 or GluA2 revealed that scaling up requires the interaction between GRIP1 and GluA2. Finally, GRIP1 synaptic accumulation during scaling up did not require GluA2 binding. Taken together, our data support a model in which activity-dependent trafficking of GRIP1 to synaptic sites drives the forward trafficking and enhanced synaptic accumulation of GluA2-containing AMPAR during synaptic scaling up.

  8. Expression of cadR Enhances its Specific Activity for Cd Detoxification and Accumulation in Arabidopsis.

    PubMed

    Li, Jingrui; Wei, Xuezhi; Yu, Pengli; Deng, Xin; Xu, Wenxiu; Ma, Mi; Zhang, Haiyan

    2016-08-01

    Cadmium (Cd) is a transition metal that is highly toxic in biological systems. Anthropogenic emissions of Cd have increased biogeochemical cycling and the amount of Cd in the biosphere. Here we studied the utility of a bacterial Cd-binding protein, CadR, for the remediation of Cd contamination. CadR was successfully targeted to chloroplasts using a constitutive Cauliflower mosaic virus (CaMV) 35S promoter or a shoot-specific Chl a/b-binding protein 2 gene (CAB2) promoter and an RbcS (small subunit of the Rubisco complex) transit peptide. Under short-term (2 d) exposure to Cd, the cadR transgenic plants showed up to a 2.9-fold Cd accumulation in roots compared with untransformed plants. Under medium term (7 d) exposure to Cd, the concentrations of Cd in leaves began to increase but there were no differences between the wild type and the cadR transgenic plants. Under long-term (16 d) exposure to Cd, the cadR transgenic plants accumulated greater amounts of Cd in leaves than the untransformed plants. Total Cd accumulation (µg per plant) in shoots and roots of the plants expressing cadR were significantly higher (up to 3.5-fold in shoots and 5.2-fold in roots) than those of the untransformed plants. We also found that targeting CadR to chloroplasts facilitated chloroplastic metal homeostasis and Chl b accumulation. Our results demonstrate that manipulating chelating capacity in chloroplasts or in the cytoplasm may be effective in modifying both the accumulation of and resistance to Cd.

  9. Myeloperoxidase scavenges peroxynitrite: A novel anti-inflammatory action of the heme enzyme.

    PubMed

    Koyani, Chintan N; Flemmig, Joerg; Malle, Ernst; Arnhold, Juergen

    2015-04-01

    Peroxynitrite, a potent pro-inflammatory and cytotoxic species, interacts with a variety of heme containing proteins. We addressed the question whether (i) the interaction of myeloperoxidase (MPO, an enzyme generating hypochlorous acid from hydrogen peroxide and chloride ions) with peroxynitrite affects the clearance of peroxynitrite, and (ii) if peroxynitrite could modulate the chlorinating activity of MPO. Our results show that this interaction promotes the decomposition of the highly reactive pro-inflammatory oxidant, whereby MPO Compound II (but not Compound I) is formed. The efficiency of MPO to remove peroxynitrite was enhanced by L-tyrosine, nitrite and (-)-epicatechin, substances known to reduce Compound II with high reaction rate. Next, peroxynitrite (added as reagent) diminished the chlorinating activity of MPO in the presence of hydrogen peroxide. Alternatively, SIN-1, a peroxynitrite donor, reduced hypochlorous acid formation by MPO, as measured by aminophenyl fluorescein oxidation (time kinetics) and taurine chloramine formation (end point measurement). At inflammatory loci, scavenging of peroxynitrite by MPO may overcome the uncontrolled peroxynitrite decomposition and formation of reactive species, which lead to cell/tissue damage.

  10. The roles of myeloperoxidase in coronary artery disease and its potential implication in plaque rupture.

    PubMed

    Teng, Nathaniel; Maghzal, Ghassan J; Talib, Jihan; Rashid, Imran; Lau, Antony K; Stocker, Roland

    2017-03-01

    Atherosclerosis is the main pathophysiological process underlying coronary artery disease (CAD). Acute complications of atherosclerosis, such as myocardial infarction, are caused by the rupture of vulnerable atherosclerotic plaques, which are characterized by thin, highly inflamed, and collagen-poor fibrous caps. Several lines of evidence mechanistically link the heme peroxidase myeloperoxidase (MPO), inflammation as well as acute and chronic manifestations of atherosclerosis. MPO and MPO-derived oxidants have been shown to contribute to the formation of foam cells, endothelial dysfunction and apoptosis, the activation of latent matrix metalloproteinases, and the expression of tissue factor that can promote the development of vulnerable plaque. As such, detection, quantification and imaging of MPO mass and activity have become useful in cardiac risk stratification, both for disease assessment and in the identification of patients at risk of plaque rupture. This review summarizes the current knowledge about the role of MPO in CAD with a focus on its possible roles in plaque rupture and recent advances to quantify and image MPO in plasma and atherosclerotic plaques.

  11. Myeloperoxidase scavenges peroxynitrite: A novel anti-inflammatory action of the heme enzyme

    PubMed Central

    Koyani, Chintan N.; Flemmig, Joerg; Malle, Ernst; Arnhold, Juergen

    2015-01-01

    Peroxynitrite, a potent pro-inflammatory and cytotoxic species, interacts with a variety of heme containing proteins. We addressed the question whether (i) the interaction of myeloperoxidase (MPO, an enzyme generating hypochlorous acid from hydrogen peroxide and chloride ions) with peroxynitrite affects the clearance of peroxynitrite, and (ii) if peroxynitrite could modulate the chlorinating activity of MPO. Our results show that this interaction promotes the decomposition of the highly reactive pro-inflammatory oxidant, whereby MPO Compound II (but not Compound I) is formed. The efficiency of MPO to remove peroxynitrite was enhanced by l-tyrosine, nitrite and (−)-epicatechin, substances known to reduce Compound II with high reaction rate. Next, peroxynitrite (added as reagent) diminished the chlorinating activity of MPO in the presence of hydrogen peroxide. Alternatively, SIN-1, a peroxynitrite donor, reduced hypochlorous acid formation by MPO, as measured by aminophenyl fluorescein oxidation (time kinetics) and taurine chloramine formation (end point measurement). At inflammatory loci, scavenging of peroxynitrite by MPO may overcome the uncontrolled peroxynitrite decomposition and formation of reactive species, which lead to cell/tissue damage. PMID:25731855

  12. The sulfonium ion linkage in myeloperoxidase. Direct spectroscopic detection by isotopic labeling and effect of mutation.

    PubMed

    Kooter, I M; Moguilevsky, N; Bollen, A; van der Veen, L A; Otto, C; Dekker, H L; Wever, R

    1999-09-17

    The heme group of myeloperoxidase is covalently linked via two ester bonds to the protein and a unique sulfonium ion linkage involving Met(243). Mutation of Met(243) into Thr, Gln, and Val, which are the corresponding residues of eosinophil peroxidase, lactoperoxidase, and thyroid peroxidase, respectively, and into Cys was performed. The Soret band in the optical absorbance spectrum in the oxidized mutants is now found at approximately 411 nm. Both the pyridine hemochrome spectra and resonance Raman spectra of the mutants are affected by the mutation. In the Met(243) mutants the affinity for chloride has decreased 100-fold. All mutants have lost their chlorination activity, except for the M243T mutant, which still has 15% activity left. By Fourier transform infared difference spectroscopy it was possible to specifically detect the (13)CD(3)-labeled methionyl sulfonium ion linkage. We conclude that the sulfonium ion linkage serves two roles. First, it serves as an electron-withdrawing substituent via its positive charge, and, second, together with its neighboring residue Glu(242), it appears to be responsible for the lower symmetry of the heme group and distortion from the planar conformation normally seen in heme-containing proteins.

  13. Niemann-Pick Type C2 Protein Mediates Hepatic Stellate Cells Activation by Regulating Free Cholesterol Accumulation.

    PubMed

    Twu, Yuh-Ching; Lee, Tzong-Shyuan; Lin, Yun-Lian; Hsu, Shih-Ming; Wang, Yuan-Hsi; Liao, Chia-Yu; Wang, Chung-Kwe; Liang, Yu-Chih; Liao, Yi-Jen

    2016-07-13

    In chronic liver diseases, regardless of their etiology, the development of fibrosis is the first step toward the progression to cirrhosis, portal hypertension, and hepatocellular carcinoma. Hepatic stellate cells (HSCs) are the main profibrogenic cells that promote the pathogenesis of liver fibrosis, and so it is important to identify the molecules that regulate HSCs activation and liver fibrosis. Niemann-Pick type C2 (NPC2) protein plays an important role in the regulation of intracellular cholesterol homeostasis by directly binding with free cholesterol. However, the roles of NPC2 in HSCs activation and liver fibrosis have not been explored in detail. Since a high-cholesterol diet exacerbates liver fibrosis progression in both rodents and humans, we propose that the expression of NPC2 affects free cholesterol metabolism and regulates HSCs activation. In this study, we found that NPC2 is decreased in both thioacetamide- and carbon tetrachloride-induced liver fibrosis tissues. In addition, NPC2 is expressed in quiescent HSCs, but its activation status is down-regulated. Knockdown of NPC2 in HSC-T6 cells resulted in marked increases in transforming growth factor-β1 (TGF-β1)-induced collagen type 1 α1 (Col1a1), α-smooth muscle actin (α-SMA) expression, and Smad2 phosphorylation. In contrast, NPC2 overexpression decreased TGF-β1-induced HSCs activation. We further demonstrated that NPC2 deficiency significantly increased the accumulation of free cholesterol in HSCs, increasing Col1a1 and α-SMA expression and activating Smad2, and leading to sensitization of HSCs to TGF-β1 activation. In contrast, overexpression of NPC2 decreased U18666A-induced free cholesterol accumulation and inhibited the subsequent HSCs activation. In conclusion, our study has demonstrated that NPC2 plays an important role in HSCs activation by regulating the accumulation of free cholesterol. NPC2 overexpression may thus represent a new treatment strategy for liver fibrosis.

  14. Niemann-Pick Type C2 Protein Mediates Hepatic Stellate Cells Activation by Regulating Free Cholesterol Accumulation

    PubMed Central

    Twu, Yuh-Ching; Lee, Tzong-Shyuan; Lin, Yun-Lian; Hsu, Shih-Ming; Wang, Yuan-Hsi; Liao, Chia-Yu; Wang, Chung-Kwe; Liang, Yu-Chih; Liao, Yi-Jen

    2016-01-01

    In chronic liver diseases, regardless of their etiology, the development of fibrosis is the first step toward the progression to cirrhosis, portal hypertension, and hepatocellular carcinoma. Hepatic stellate cells (HSCs) are the main profibrogenic cells that promote the pathogenesis of liver fibrosis, and so it is important to identify the molecules that regulate HSCs activation and liver fibrosis. Niemann-Pick type C2 (NPC2) protein plays an important role in the regulation of intracellular cholesterol homeostasis by directly binding with free cholesterol. However, the roles of NPC2 in HSCs activation and liver fibrosis have not been explored in detail. Since a high-cholesterol diet exacerbates liver fibrosis progression in both rodents and humans, we propose that the expression of NPC2 affects free cholesterol metabolism and regulates HSCs activation. In this study, we found that NPC2 is decreased in both thioacetamide- and carbon tetrachloride-induced liver fibrosis tissues. In addition, NPC2 is expressed in quiescent HSCs, but its activation status is down-regulated. Knockdown of NPC2 in HSC-T6 cells resulted in marked increases in transforming growth factor-β1 (TGF-β1)-induced collagen type 1 α1 (Col1a1), α-smooth muscle actin (α-SMA) expression, and Smad2 phosphorylation. In contrast, NPC2 overexpression decreased TGF-β1-induced HSCs activation. We further demonstrated that NPC2 deficiency significantly increased the accumulation of free cholesterol in HSCs, increasing Col1a1 and α-SMA expression and activating Smad2, and leading to sensitization of HSCs to TGF-β1 activation. In contrast, overexpression of NPC2 decreased U18666A-induced free cholesterol accumulation and inhibited the subsequent HSCs activation. In conclusion, our study has demonstrated that NPC2 plays an important role in HSCs activation by regulating the accumulation of free cholesterol. NPC2 overexpression may thus represent a new treatment strategy for liver fibrosis. PMID:27420058

  15. The ARIC carotid MRI study of blood cellular markers: an inverse association of monocyte myeloperoxidase content with peripheral arterial disease.

    PubMed

    Matijevic, Nena; Wu, Kenneth K; Nidkarni, Nivedita; Heiss, Gerardo; Folsom, Aaron R

    2011-04-01

    We evaluated the association of blood monocyte and platelet activation markers with the risk of peripheral artery disease (PAD) in a multicenter study of atherosclerosis among African American and Caucasian patients. Flow cytometric analysis of blood cells was performed in 1791 participants (209 cases with PAD and 1582 noncases) from the cross-sectional Atherosclerosis Risk in Communities Carotid Magnetic Resonance Imaging ([MRI] ARIC Carotid MRI) Study to assess platelet glycoproteins IIb and IIIa, P-selectin, CD40 ligand, platelet-leukocyte aggregates, monocyte lipopolysaccharide receptor, toll-like receptors (TLRs) 2 and 4, P-selectin glycoprotein ligand 1, cyclooxygenase 2, and myeloperoxidase. Multivariate regression analyses evaluated the association of cellular markers with the risk of PAD. After adjusting for age, race, and gender, platelet CD40L, and monocyte myeloperoxidase (mMPO) levels were significantly lower (P < .001), and monocyte TLR-4 levels were higher (P = .03) in patients with PAD. With additional adjustments for conventional risk factors, mMPO remained inversely and independently associated with the risk of PAD (odds ratio [OR]: 0.35, P = .01).

  16. The ARIC Carotid MRI Study of Blood Cellular Markers: An Inverse Association of Monocyte Myeloperoxidase Content With Peripheral Arterial Disease

    PubMed Central

    Matijevic (Aleksic), Nena; Wu, Kenneth K.; Nidkarni, Nivedita; Heiss, Gerardo; Folsom, Aaron R.

    2015-01-01

    We evaluated the association of blood monocyte and platelet activation markers with the risk of peripheral artery disease (PAD) in a multicenter study of atherosclerosis among African American and Caucasian patients. Flow cytometric analysis of blood cells was performed in 1791 participants (209 cases with PAD and 1582 noncases) from the cross-sectional Atherosclerosis Risk in Communities Carotid Magnetic Resonance Imaging ([MRI] ARIC Carotid MRI) Study to assess platelet glycoproteins IIb and IIIa, P-selectin, CD40 ligand, platelet–leukocyte aggregates, monocyte lipopolysaccharide receptor, toll-like receptors (TLRs) 2 and 4, P-selectin glycoprotein ligand 1, cyclooxygenase 2, and myeloperoxidase. Multivariate regression analyses evaluated the association of cellular markers with the risk of PAD. After adjusting for age, race, and gender, platelet CD40L, and monocyte myeloperoxidase (mMPO) levels were significantly lower (P < .001), and monocyte TLR-4 levels were higher (P = .03) in patients with PAD. With additional adjustments for conventional risk factors, mMPO remained inversely and independently associated with the risk of PAD (odds ratio [OR]: 0.35, P = .01). PMID:21406422

  17. Autoantibodies to myeloperoxidase aggravate mild anti-glomerular-basement-membrane-mediated glomerular injury in the rat.

    PubMed Central

    Heeringa, P.; Brouwer, E.; Klok, P. A.; Huitema, M. G.; van den Born, J.; Weening, J. J.; Kallenberg, C. G.

    1996-01-01

    Autoantibodies to myeloperoxidase (MPO) are present in sera from patients with various forms of vasculitis-associated glomerulonephritis. Evidence for a pathogenic role of anti-MPO antibodies has been provided mainly by in vitro studies. We studied the pathogenic role of autoantibodies to MPO in a rat model of mild immune-mediated glomerular injury. Brown Norway rats were immunized with human MPO in complete Freund's adjuvant or with complete Freund's adjuvant alone. At 2 weeks after immunization, rats had developed antibodies to human and rat MPO as detected by indirect immunofluorescence, enzyme-linked immunosorbent assay, and immunoprecipitation. At this time point, rats were intravenously injected with a subnephritogenic dose of 150 micrograms of rabbit anti-rat GBM. Rats were sacrificed at 4 hours, 24 hours, 4 days, and 10 days after antibody administration. Control immunized rats developed mild glomerulonephritis characterized by slight proteinuria at day 10 (14.8 +/- 8.1 mg/24 hours) and moderate intraglomerular accumulation of ED1+ macrophages. Crescent formation, tuft necrosis, and tubular atrophy were not observed in those rats. In contrast, rats immunized with MPO developed severe glomerulonephritis characterized by the early occurrence of severe hematuria, marked proteinuria at day 10 (76.2 +/- 18.2 mg/24 hours), and massive glomerular deposition of fibrin. Complement and rat IgG were present in insudative lesions, but no linear pattern along the glomerular capillary wall was observed. By light microscopy, severe glomerular lesions were found at day 10 consisting of crescent formation and fibrinoid necrosis of capillary loops. In the interstitium, tubular necrosis and atrophy and marked interstitial mononuclear infiltration were found in conclusion, autoantibodies to MPO severely aggravate subclinical anti-GBM disease demonstrating their in vivo pathogenic potential. Images Figure 1 Figure 3 Figure 6 Figure 7 Figure 8 Figure 10 PMID:8909258

  18. Accumulation fatty acids of in Chlorella vulgaris under heterotrophic conditions in relation to activity of acetyl-CoA carboxylase, temperature, and co-immobilization with Azospirillum brasilense

    NASA Astrophysics Data System (ADS)

    Leyva, Luis A.; Bashan, Yoav; Mendoza, Alberto; de-Bashan, Luz E.

    2014-10-01

    The relation between fatty acid accumulation, activity of acetyl-CoA carboxylase (ACC), and consequently lipid accumulation was studied in the microalgae Chlorella vulgaris co-immobilized with the plant growth-promoting bacterium Azospirillum brasilense under dark heterotrophic conditions with Na acetate as a carbon source. In C. vulgaris immobilized alone, cultivation experiments for 6 days showed that ACC activity is directly related to fatty acid accumulation, especially in the last 3 days. In co-immobilization experiments, A. brasilense exerted a significant positive effect over ACC activity, increased the quantity in all nine main fatty acids, increased total lipid accumulation in C. vulgaris, and mitigated negative effects of nonoptimal temperature for growth. No correlation between ACC activity and lipid accumulation in the cells was established for three different temperatures. This study demonstrated that the interaction between A. brasilense and C. vulgaris has a significant effect on fatty acid and lipid accumulation in the microalgae.

  19. Complement (C5)-derived chemotactic activity accounts for accumulation of polymorphonuclear leukocytes in cerebrospinal fluid of rabbits with pneumococcal meningitis.

    PubMed Central

    Ernst, J D; Hartiala, K T; Goldstein, I M; Sande, M A

    1984-01-01

    Experiments were performed to identify the chemoattractant for polymorphonuclear leukocytes that appears in the cerebrospinal fluid of rabbits with experimental pneumococcal meningitis. Meningitis was induced in anesthetized New Zealand white rabbits by injecting 10(4) cells of stationary-phase Streptococcus pneumoniae type III intracisternally. Before bacteria were injected, cerebrospinal fluid contained neither polymorphonuclear leukocytes nor chemotactic activity. Significant chemotactic activity for rabbit polymorphonuclear leukocytes was detected 12 h after inoculation with bacteria and was maximal after 18 to 20 h. Chemotactic activity appeared in cerebrospinal fluid while concentrations of pneumococci and total protein were increasing but before there was any accumulation of polymorphonuclear leukocytes. The chemotactic activity in cerebrospinal fluid was heat stable (56 degrees C for 30 min), eluted from Sephadex G-75 with a profile identical to that of the chemotactic activity in zymosan-activated rabbit serum, and was inhibited by treatment with antibodies to native human C5. In addition, preincubation of polymorphonuclear leukocytes with partially purified rabbit C5a selectively inhibited their subsequent chemotactic responses to cerebrospinal fluid. These data indicate that complement (C5)-derived chemotactic activity appears in cerebrospinal fluid during the course of experimental pneumococcal meningitis in rabbits and suggest that this activity accounts for the accumulation of polymorphonuclear leukocytes observed in this infection. PMID:6480117

  20. Novel phosphatidylethanolamine derivatives accumulate in circulation in hyperlipidemic ApoE−/− mice and activate platelets via TLR2

    PubMed Central

    Biswas, Sudipta; Xin, Liang; Panigrahi, Soumya; Zimman, Alejandro; Wang, Hua; Yakubenko, Valentin P.; Byzova, Tatiana V.; Salomon, Robert G.

    2016-01-01

    A prothrombotic state and increased platelet reactivity are common in dyslipidemia and oxidative stress. Lipid peroxidation, a major consequence of oxidative stress, generates highly reactive products, including hydroxy-ω-oxoalkenoic acids that modify autologous proteins generating biologically active derivatives. Phosphatidylethanolamine, the second most abundant eukaryotic phospholipid, can also be modified by hydroxy-ω-oxoalkenoic acids. However, the conditions leading to accumulation of such derivatives in circulation and their biological activities remain poorly understood. We now show that carboxyalkylpyrrole-phosphatidylethanolamine derivatives (CAP-PEs) are present in the plasma of hyperlipidemic ApoE−/− mice. CAP-PEs directly bind to TLR2 and induces platelet integrin αIIbβ3 activation and P-selectin expression in a Toll-like receptor 2 (TLR2)-dependent manner. Platelet activation by CAP-PEs includes assembly of TLR2/TLR1 receptor complex, induction of downstream signaling via MyD88/TIRAP, phosphorylation of IRAK4, and subsequent activation of tumor necrosis factor receptor–associated factor 6. This in turn activates the Src family kinases, spleen tyrosine kinase and PLCγ2, and platelet integrins. Murine intravital thrombosis studies demonstrated that CAP-PEs accelerate thrombosis in TLR2-dependent manner and that TLR2 contributes to accelerate thrombosis in mice in the settings of hyperlipidemia. Our study identified the novel end-products of lipid peroxidation, accumulating in circulation in hyperlipidemia and inducing platelet activation by promoting cross-talk between innate immunity and integrin activation signaling pathways. PMID:27015965

  1. Four new triterpenes and triterpene glycosides from the leaves of Ilex latifolia and their inhibitory activity on triglyceride accumulation.

    PubMed

    Wang, Cun-Qin; Li, Man-Mei; Zhang, Wei; Wang, Lei; Fan, Chun-Lin; Feng, Rui-Bing; Zhang, Xiao-Qi; Ye, Wen-Cai

    2015-10-01

    Two new triterpenes (1 and 2) and two new triterpene glycosides (3 and 4), along with six known triterpenes (5-10) were isolated from the leaves of Ilex latifolia. The structures of new compounds were elucidated on the basis of NMR, HR-MS, and X-ray diffraction analysis. Compounds 4 and 5 showed potent inhibitory activity on oleic acid/palmitic acid induced triglyceride accumulation on HepG2 cells.

  2. Transient and steady-state kinetics of the oxidation of substituted benzoic acid hydrazides by myeloperoxidase.

    PubMed

    Burner, U; Obinger, C; Paumann, M; Furtmüller, P G; Kettle, A J

    1999-04-02

    Myeloperoxidase is the most abundant protein in neutrophils and catalyzes the production of hypochlorous acid. This potent oxidant plays a central role in microbial killing and inflammatory tissue damage. 4-Aminobenzoic acid hydrazide (ABAH) is a mechanism-based inhibitor of myeloperoxidase that is oxidized to radical intermediates that cause enzyme inactivation. We have investigated the mechanism by which benzoic acid hydrazides (BAH) are oxidized by myeloperoxidase, and we have determined the features that enable them to inactivate the enzyme. BAHs readily reduced compound I of myeloperoxidase. The rate constants for these reactions ranged from 1 to 3 x 10(6) M-1 s-1 (15 degrees C, pH 7.0) and were relatively insensitive to the substituents on the aromatic ring. Rate constants for reduction of compound II varied between 6.5 x 10(5) M-1 s-1 for ABAH and 1.3 x 10(3) M-1 s-1 for 4-nitrobenzoic acid hydrazide (15 degrees C, pH 7.0). Reduction of both compound I and compound II by BAHs adhered to the Hammett rule, and there were significant correlations with Brown-Okamoto substituent constants. This indicates that the rates of these reactions were simply determined by the ease of oxidation of the substrates and that the incipient free radical carried a positive charge. ABAH was oxidized by myeloperoxidase without added hydrogen peroxide because it underwent auto-oxidation. Although BAHs generally reacted rapidly with compound II, they should be poor peroxidase substrates because the free radicals formed during peroxidation converted myeloperoxidase to compound III. We found that the reduction of ferric myeloperoxidase by BAH radicals was strongly influenced by Hansch's hydrophobicity constants. BAHs containing more hydrophilic substituents were more effective at converting the enzyme to compound III. This implies that BAH radicals must hydrogen bond to residues in the distal heme pocket before they can reduce the ferric enzyme. Inactivation of myeloperoxidase by BAHs

  3. CYLD, a deubiquitinase specific for lysine63-linked polyubiquitins, accumulates at the postsynaptic density in an activity-dependent manner

    SciTech Connect

    Dosemeci, Ayse; Thein, Soe; Yang, Yijung; Reese, Thomas S.; Tao-Cheng, Jung-Hwa

    2013-01-04

    Highlights: Black-Right-Pointing-Pointer CYLD is a deubiquitinase specific for lysine63-linked polyubiquitins. Black-Right-Pointing-Pointer Presence of CYLD in PSDs is established by biochemistry and immunoEM. Black-Right-Pointing-Pointer CYLD accumulates on PSDs upon depolarization of neurons. Black-Right-Pointing-Pointer Accumulation of CYLD at PSDs may regulate trafficking/degradation of synaptic proteins. -- Abstract: Polyubiquitin chains on proteins flag them for distinct fates depending on the type of polyubiquitin linkage. While lysine48-linked polyubiquitination directs proteins to proteasomal degradation, lysine63-linked polyubiquitination promotes different protein trafficking and is involved in autophagy. Here we show that postsynaptic density (PSD) fractions from adult rat brain contain deubiquitinase activity that targets both lysine48 and lysine63-linked polyubiquitins. Comparison of PSD fractions with parent subcellular fractions by Western immunoblotting reveals that CYLD, a deubiquitinase specific for lysine63-linked polyubiquitins, is highly enriched in the PSD fraction. Electron microscopic examination of hippocampal neurons in culture under basal conditions shows immunogold label for CYLD at the PSD complex in approximately one in four synapses. Following depolarization by exposure to high K+, the proportion of CYLD-labeled PSDs as well as the labeling intensity of CYLD at the PSD increased by more than eighty percent, indicating that neuronal activity promotes accumulation of CYLD at the PSD. An increase in postsynaptic CYLD following activity would promote removal of lysine63-polyubiquitins from PSD proteins and thus could regulate their trafficking and prevent their autophagic degradation.

  4. Galactinol synthase enzyme activity influences raffinose family oligosaccharides (RFO) accumulation in developing chickpea (Cicer arietinum L.) seeds.

    PubMed

    Gangola, Manu P; Jaiswal, Sarita; Kannan, Udhaya; Gaur, Pooran M; Båga, Monica; Chibbar, Ravindra N

    2016-05-01

    To understand raffinose family oligosaccharides (RFO) metabolism in chickpea (Cicer arietinum L.) seeds, RFO accumulation and corresponding biosynthetic enzymes activities were determined during seed development of chickpea genotypes with contrasting RFO concentrations. RFO concentration in mature seeds was found as a facilitator rather than a regulating step of seed germination. In mature seeds, raffinose concentrations ranged from 0.38 to 0.68 and 0.75 to 0.99 g/100 g, whereas stachyose concentrations varied from 0.79 to 1.26 and 1.70 to 1.87 g/100 g indicating significant differences between low and high RFO genotypes, respectively. Chickpea genotypes with high RFO concentration accumulated higher concentrations of myo-inositol and sucrose during early seed developmental stages suggesting that initial substrate concentrations may influence RFO concentration in mature seeds. High RFO genotypes showed about two to three-fold higher activity for all RFO biosynthetic enzymes compared to those with low RFO concentrations. RFO biosynthetic enzymes activities correspond with accumulation of individual RFO during seed development.

  5. Neutrophil myeloperoxidase and its substrates: formation of specific markers and reactive compounds during inflammation

    PubMed Central

    Kato, Yoji

    2016-01-01

    Myeloperoxidase is an inflammatory enzyme that generates reactive hypochlorous acid in the presence of hydrogen peroxide and chloride ion. However, this enzyme also uses bromide ion or thiocyanate as a substrate to form hypobromous or hypothiocyanous acid, respectively. These species play important roles in host defense against the invasion of microorganisms. In contrast, these enzyme products modify biomolecules in hosts during excess inflammation, indicating that the action of myeloperoxidase is both beneficial and harmful. Myeloperoxidase uses other endogenous compounds, such as serotonin, urate, and l-tyrosine, as substrates. This broad-range specificity may have some biological implications. Target molecules of this enzyme and its products vary, including low-molecular weight thiols, proteins, nucleic acids, and lipids. The modified products represent biomarkers of myeloperoxidase action. Moderate inhibition of this enzyme might be critical for the prevention/modulation of excess, uncontrolled inflammatory events. Some phytochemicals inhibit myeloperoxidase, which might explain the reductive effect caused by the intake of vegetables and fruits on cardiovascular diseases. PMID:27013775

  6. Biological reactivity of hypochlorous acid: implications for microbicidal mechanisms of leukocyte myeloperoxidase.

    PubMed Central

    Albrich, J M; McCarthy, C A; Hurst, J K

    1981-01-01

    Oxidative degradation of biological substrates by hypochlorous acid has been examined under reaction conditions similar to those found in active phagosomes. Iron sulfur proteins are bleached extremely rapidly, followed in decreasing order by beta-carotene, nucleotides, porphyrins, and heme proteins. Enzymes containing essential cysteine molecules are inactivated with an effectiveness that roughly parallels the nucleophilic reactivities of their sulfhydryl groups. Other compounds, including glucosamines, quinones, riboflavin, and, except for N-chlorination, phospholipids, are unreactive. Rapid irreversible oxidation of cytochromes, adenine nucleotides, and carotene pigments occurs when bacterial cells are exposed to exogenous hypochlorous acid; with Escherichia coli, titrimetric oxidation of cytochrome was found to coincide with loss of aerobic respiration. The occurrence of these cellular reactions implicates hypochlorous acid as a primary microbicide in myeloperoxidase-containing leukocytes; the reactivity patterns observed are consistent with the view that bactericidal action results primarily from loss of energy-linked respiration due to destruction of cellular electron transport chains and the adenine nucleotide pool. PMID:6264434

  7. Present-day Block Motions and Strain Accumulation on Active Faults in the Caribbean

    NASA Astrophysics Data System (ADS)

    Symithe, S. J.; Calais, E.; Freed, A. M.

    2014-12-01

    The quasi-frontal subduction of the north and south American plates under the Lesser Antilles and the left and right lateral strike-slip along the northern and southern margins of the Caribbean plate offer the opportunity to study the transition from subduction to strike-slip between major plates. In addition, the segmentation and degree of interplate coupling at the Lesser Antilles subduction is key to our understanding of the earthquake potential of a subduction whose length is similar to the rupture area of the Mw9.0, 2011, Tohoku earthquake in Japan. We used the block modeling approach described in Meade and Loveless (2009) to test the optimal block geometry for the northern, eastern and southern boundaries of the Caribbean plate. We solved for angular velocities for each block/plate and strain accumulation rates for all major faults in the region. Then we calculated the variations in interplate coupling along the subduction plate boundaries using the accumulated strain rates. We tested 11 different block geometries; they are all based on geological evidences unless they are suggested by discrepancies within the GPS and seismological data or by previously published results. We confirm the existence of the micro Gonave plate. The boundary between the Micro-Gonave plate and the Hispaniola crustal block is better suited along the Haitian-Thrust-Belt instead of the Neiba-Matheux fault. The interseismic GPS velocities do not show evidence for a distinct North Lesser Antilles block. We found a totally uncoupled section of the subduction starting from the Puerto-Rico trench to the end of the Lesser Antilles section. All the relative motion of the Caribbean block is lost aseismically along the boundary of that portion of the subduction. While we found strong coupling along the northern Hispaniola section, most of the deformation on this region is being accumulated along intrablock faults with very low strain (~2mm/yr) along the intraplate subduction interface. We also

  8. Reduction of lipid accumulation in white adipose tissues by Cassia tora (Leguminosae) seed extract is associated with AMPK activation.

    PubMed

    Tzeng, Thing-Fong; Lu, Hung-Jen; Liou, Shorong-Shii; Chang, Chia Ju; Liu, I-Min

    2013-01-15

    Natural herbal medications may be one answer to the worldwide epidemic of obesity. This study examines the effects of Cassia seed ethanol extract (CSEE) upon lipid accumulation in white adipose tissue (WAT). CSEE exhibited a significant concentration-dependent decrease in the intracellular accumulation of trigycerides in 3T3-L1 adipocytes. After being fed a high-fat diet (HFD) for 2 weeks, rats were fed CSEE (100, 200 or 300 mg/kg) once daily for 8 weeks. CSEE caused dose-related reductions in body weight gain (as well as plasma lipid levels and epididymal WAT sizes in HFD-fed rats). CSEE enhanced the phosphorylation of AMP-activated protein kinase (AMPK) and its primary downstream targeting enzyme, acetyl-CoA carboxylase, up-regulated gene expression of carnitine palmitoyl transferase 1, and down-regulated sterol regulatory element-binding protein 1 and fatty acid synthase protein levels in epididymal WAT of HFD-fed rats. CSEE could attenuate lipid accumulation in WAT via AMPK signaling pathway activation.

  9. Effects of waste activated sludge and surfactant addition on primary sludge hydrolysis and short-chain fatty acids accumulation.

    PubMed

    Ji, Zhouying; Chen, Guanlan; Chen, Yinguang

    2010-05-01

    This paper focused on the effects of waste activated sludge (WAS) and surfactant sodium dodecylbenzene sulfonate (SDBS) addition on primary sludge (PS) hydrolysis and short-chain fatty acids (SCFA) accumulation in fermentation. The results showed that sludge hydrolysis, SCFA accumulation, NH(4)(+)-N and PO(4)(3-)-P release, and volatile suspended solids (VSS) reduction were increased by WAS addition to PS, which were further increased by the addition of SDBS to the mixture of PS and WAS. Acetic, propionic and valeric acids were the top three SCFA in all experiments. Also, the fermentation liquids of PS, PS+WAS, and PS+WAS+SDBS were added, respectively, to municipal wastewater to examine their effects on biological municipal wastewater treatment, and the enhancement of both wastewater nitrogen and phosphorus removals was observed compared with no fermentation liquid addition.

  10. Inhibition of Morganella morganii Histidine Decarboxylase Activity and Histamine Accumulation in Mackerel Muscle Derived from Filipendula ulumaria Extracts.

    PubMed

    Nitta, Yoko; Yasukata, Fumiko; Kitamoto, Noritoshi; Ito, Mikiko; Sakaue, Motoyoshi; Kikuzaki, Hiroe; Ueno, Hiroshi

    2016-03-01

    Filipendula ulmaria, also known as meadowsweet, is an herb; its extract was examined for the prevention of histamine production, primarily that caused by contaminated fish. The efficacy of meadowsweet was assessed using two parameters: inhibition of Morganella morganii histidine decarboxylase (HDC) and inhibition of histamine accumulation in mackerel. Ellagitannins from F. ulmaria (rugosin D, rugosin A methyl ester, tellimagrandin II, and rugosin A) were previously shown to be potent inhibitors of human HDC; and in the present work, these compounds inhibited M. morganii HDC, with half maximal inhibitory concentration values of 1.5, 4.4, 6.1, and 6.8 μM, respectively. Application of the extracts (at 2 wt%) to mackerel meat yielded significantly decreased histamine accumulation compared with treatment with phosphate-buffered saline as a control. Hence, F. ulmaria exhibits inhibitory activity against bacterial HDC and might be effective for preventing food poisoning caused by histamine.

  11. A Fuzzy Logic Prompting Mechanism Based on Pattern Recognition and Accumulated Activity Effective Index Using a Smartphone Embedded Sensor.

    PubMed

    Liu, Chung-Tse; Chan, Chia-Tai

    2016-08-19

    Sufficient physical activity can reduce many adverse conditions and contribute to a healthy life. Nevertheless, inactivity is prevalent on an international scale. Improving physical activity is an essential concern for public health. Reminders that help people change their health behaviors are widely applied in health care services. However, timed-based reminders deliver periodic prompts suffer from flexibility and dependency issues which may decrease prompt effectiveness. We propose a fuzzy logic prompting mechanism, Accumulated Activity Effective Index Reminder (AAEIReminder), based on pattern recognition and activity effective analysis to manage physical activity. AAEIReminder recognizes activity levels using a smartphone-embedded sensor for pattern recognition and analyzing the amount of physical activity in activity effective analysis. AAEIReminder can infer activity situations such as the amount of physical activity and days spent exercising through fuzzy logic, and decides whether a prompt should be delivered to a user. This prompting system was implemented in smartphones and was used in a short-term real-world trial by seventeenth participants for validation. The results demonstrated that the AAEIReminder is feasible. The fuzzy logic prompting mechanism can deliver prompts automatically based on pattern recognition and activity effective analysis. AAEIReminder provides flexibility which may increase the prompts' efficiency.

  12. Reduced dermis thickness and AGE accumulation in diabetic abdominal skin.

    PubMed

    Niu, Yiwen; Cao, Xiaozan; Song, Fei; Xie, Ting; Ji, Xiaoyun; Miao, Mingyuan; Dong, Jiaoyun; Tian, Ming; Lin, Yuan; Lu, Shuliang

    2012-09-01

    Dermatological problems in diabetes might play an important role in the spontaneous ulcers and impaired wound healing that are seen in diabetic patients. Investigation of the cause of diabetic skin disorders is critical for identifying effective treatment. The abdominal full-thickness skin tissues of 33 patients (14 nondiabetic and 19 diabetic) were analyzed. The cell viability and malondialdehyde (MDA) production of fibroblasts were measured after advanced glycosylation end product (AGE)-bovine serum albumin (BSA) exposure. Cutaneous histological observation showed reduced thickness of the diabetic abdominal dermis with morphological characteristics of obscured multilayer epithelium and shortened, thinned, and disorganized collagen fibrils with focal chronic inflammatory cell infiltration when compared with controls of the same age. Accumulation of AGEs in diabetic skin was prominent. Less hydroxyproline, higher myeloperoxidase activity, and increased MDA content were detected in diabetic skin. In vitro, the time- and dose-dependent inhibitory effects of AGE-BSA on fibroblast viability as well as the fact that AGE-BSA could promote MDA production of fibroblasts were shown. It is shown that the accumulation of AGEs in diabetic skin tissue induces an oxidative damage of fibroblasts and acts as an important contributor to the thinner diabetic abdominal dermis. The authors believe that diabetic cutaneous properties at baseline may increase the susceptibility to injury, and diabetic wounds possess atypical origin in the repair process.

  13. Accumulate-Repeat-Accumulate-Accumulate-Codes

    NASA Technical Reports Server (NTRS)

    Divsalar, Dariush; Dolinar, Sam; Thorpe, Jeremy

    2004-01-01

    Inspired by recently proposed Accumulate-Repeat-Accumulate (ARA) codes [15], in this paper we propose a channel coding scheme called Accumulate-Repeat-Accumulate-Accumulate (ARAA) codes. These codes can be seen as serial turbo-like codes or as a subclass of Low Density Parity Check (LDPC) codes, and they have a projected graph or protograph representation; this allows for a high-speed iterative decoder implementation using belief propagation. An ARAA code can be viewed as a precoded Repeat-and-Accumulate (RA) code with puncturing in concatenation with another accumulator, where simply an accumulator is chosen as the precoder; thus ARAA codes have a very fast encoder structure. Using density evolution on their associated protographs, we find examples of rate-lJ2 ARAA codes with maximum variable node degree 4 for which a minimum bit-SNR as low as 0.21 dB from the channel capacity limit can be achieved as the block size goes to infinity. Such a low threshold cannot be achieved by RA or Irregular RA (IRA) or unstructured irregular LDPC codes with the same constraint on the maximum variable node degree. Furthermore by puncturing the accumulators we can construct families of higher rate ARAA codes with thresholds that stay close to their respective channel capacity thresholds uniformly. Iterative decoding simulation results show comparable performance with the best-known LDPC codes but with very low error floor even at moderate block sizes.

  14. Down-regulation of catalase activity allows transient accumulation of a hydrogen peroxide signal in Chlamydomonas reinhardtii.

    PubMed

    Michelet, Laure; Roach, Thomas; Fischer, Beat B; Bedhomme, Mariette; Lemaire, Stéphane D; Krieger-Liszkay, Anja

    2013-06-01

    In photosynthetic organisms, excess light is a stress that induces production of reactive oxygen species inside the chloroplasts. As a response, the capacity of antioxidative defence mechanisms increases. However, when cells of Chlamydomonas reinhardtii were shifted from dark to high light, a reversible partial inactivation of catalase activity was observed, which correlated with a transient increase in the level of H2 O2 in the 10 μm range. This concentration range seems to be necessary to activate H2 O2 -dependent signalling pathways stimulating the expression of H2 O2 responsive genes, such as the heat shock protein HSP22C. Catalase knock-down mutants had lost the transient accumulation of H2 O2 , suggesting that a decrease in catalase activity was the key element for establishing a transient H2 O2 burst. Catalase was inactivated by a one-electron event consistent with the reduction of a single cysteine. We propose that under high light intensity, the redox state of the photosynthetic electron transport chain is sensed and transmitted to the cytosol to regulate the catalase activity. This allows a transient accumulation of H2 O2 , inducing a signalling event that is transmitted to the nucleus to modulate the expression of chloroplast-directed protection enzymes.

  15. Tooth development in Ambystoma mexicanum: phosphatase activities, calcium accumulation and cell proliferation in the tooth-forming tissues.

    PubMed

    Wistuba, Joachim; Ehmcke, Jens; Clemen, Günter

    2003-06-01

    Prerequisites of tooth formation, cell proliferation in the tooth-forming tissues, calcium accumulation and the enzymatic activities of alkaline (ALP) and acid phosphatases (ACP) were investigated by immunohistochemical and histochemical methods in various developmental stages of the Mexican Axolotl, Ambystoma mexicanum. During the growth of replacement teeth, the tooth-forming tissues continually recruit cells from the surrounding regions. The basal layer of the oral epithelium, the dental lamina and sometimes even the outer enamel epithelium provide cells for the differentiated inner enamel epithelium, in which the active ameloblasts are localized. The differentiating odontoblasts are derived from proliferating cells situated basally to the replacement teeth in the mesenchymal tissue. When differentiation has started and the cells have become functional, proliferative activity can no longer be observed. Calcium is accumulated close to the site of mineralization in the inner enamel epithelium and in the odontoblasts as it is in mammals, elasmobranchii and teleostei. The activities of ACP and ALP related to the mineralization of the replacement teeth are separated spatially and not sequentially as they are in mammals. However, the results indicate a similar function of these enzymatic components in relation to tooth formation and maturation of mineral deposition. Most of the substantial processes related to tooth formation reported from other vertebrates occur in a manner similar to that in Ambystoma mexicanum, but there also seem to be basic mechanisms present that are realised in a unique way in this urodele.

  16. Spt5 accumulation at variable genes distinguishes somatic hypermutation in germinal center B cells from ex vivo-activated cells.

    PubMed

    Maul, Robert W; Cao, Zheng; Venkataraman, Lakshmi; Giorgetti, Carol A; Press, Joan L; Denizot, Yves; Du, Hansen; Sen, Ranjan; Gearhart, Patricia J

    2014-10-20

    Variable (V) genes of immunoglobulins undergo somatic hypermutation by activation-induced deaminase (AID) to generate amino acid substitutions that encode antibodies with increased affinity for antigen. Hypermutation is restricted to germinal center B cells and cannot be recapitulated in ex vivo-activated splenic cells, even though the latter express high levels of AID. This suggests that there is a specific feature of antigen activation in germinal centers that recruits AID to V genes which is absent in mitogen-activated cultured cells. Using two Igh knock-in mouse models, we found that RNA polymerase II accumulates in V regions in B cells after both types of stimulation for an extended distance of 1.2 kb from the TATA box. The paused polymerases generate abundant single-strand DNA targets for AID. However, there is a distinct accumulation of the initiating form of polymerase, along with the transcription cofactor Spt5 and AID, in the V region from germinal center cells, which is totally absent in cultured cells. These data support a model where mutations are prevalent in germinal center cells, but not in ex vivo cells, because the initiating form of polymerase is retained, which affects Spt5 and AID recruitment.

  17. Age-related decrease in cold-activated brown adipose tissue and accumulation of body fat in healthy humans.

    PubMed

    Yoneshiro, Takeshi; Aita, Sayuri; Matsushita, Mami; Okamatsu-Ogura, Yuko; Kameya, Toshimitsu; Kawai, Yuko; Miyagawa, Masao; Tsujisaki, Masayuki; Saito, Masayuki

    2011-09-01

    Brown adipose tissue (BAT) can be identified by (18)F-fluorodeoxyglucose (FDG)-positron emission tomography (PET) combined with X-ray computed tomography (CT) in adult humans. The objective of this study was to clarify the relationship between BAT and adiposity in healthy adult humans, particularly to test the idea that decreased BAT activity may be associated with body fat accumulation with age. One hundred and sixty-two healthy volunteers aged 20-73 years (103 males and 59 females) underwent FDG-PET/CT after 2-h cold exposure at 19 °C with light clothing. Cold-activated BAT was detected in 41% of the subjects (BAT-positive). Compared with the BAT-negative group, the BAT-positive group was younger (P < 0.01) and showed a lower BMI (P < 0.01), body fat content (P < 0.01), and abdominal fat (P < 0.01). The incidence of cold-activated BAT decreased with age (P < 0.01), being more than 50% in the twenties, but less than 10% in the fifties and sixties. The adiposity-related parameters showed some sex differences, but increased with age in the BAT-negative group (P < 0.01), while they remained unchanged from the twenties to forties in the BAT-positive group, in both sexes. These results suggest that decreased BAT activity may be associated with accumulation of body fat with age.

  18. EFFECT OF ACTIVE ACCUMULATION OF CALCIUM AND PHOSPHATE IONS ON THE STRUCTURE OF RAT LIVER MITOCHONDRIA.

    PubMed

    GREENAWALT, J W; ROSSI, C S; LEHNINGER, A L

    1964-10-01

    Rat liver mitochondria allowed to accumulate maximal amounts of Ca(++) and HPO(4) (=) ions from the suspending medium in vitro during respiration have a considerably higher specific gravity than normal mitochondria and may be easily separated from the latter by isopycnic centrifugation in density gradients of sucrose or cesium chloride. When the mitochondria are allowed to accumulate less than maximal amounts of Ca(++) and HPO(4) (=) from the medium, they have intermediate specific gravities which are roughly proportional to their content of calcium phosphate. Maximally "loaded" mitochondria are relatively homogeneous with respect to specific gravity. Correlated biochemical and electron microscopic studies show that Ca(++)-loaded mitochondria contain numerous dense granules, of which some 85 per cent are over 500 A in diameter. These granules are electron-opaque not only following fixation and staining with heavy metal reagents, but also following fixation with formaldehyde, demonstrating that the characteristic granules in Ca(++)-loaded mitochondria have intrinsic electron-opacity. The dense granules are almost always located within the inner compartment of the mitochondria and not in the space between the inner and outer membranes. They are frequently located at or near the cristae and they often show electron-transparent "cores." Such granules appear to be made up of clusters of smaller dense particles, but preliminary x-ray diffraction analysis and electron diffraction studies have revealed no evidence of crystallinity in the deposits. The electron-opaque granules decrease in number when the Ca(++)-loaded mitochondria are incubated with 2,4-dinitrophenol; simultaneously there is discharge of Ca(++) and phosphate from the mitochondria into the medium.

  19. Stress-induced changes in accumulation of sorbitol and in activities of concomitant enzymes in digestive gland of freshwater snail.

    PubMed

    Tsvetkov, I L; Konichev, A S

    2009-11-01

    Sorbitol content was determined in the digestive gland of freshwater snail (Viviparus viviparus L.) in different seasons and in a short-term experiment on the water temperature decrease and on intoxication with cadmium chloride. In the model experiments, changes in activities of enzymes involved in sorbitol metabolism (acid phosphatases, sorbitol dehydrogenase, and aldose reductase) were also studied. Sorbitol was accumulated by the snail in response to the temperature decrease (as a cryoprotectant) and under conditions of acute intoxication (as a probable metabolic regulator or a nonspecific protective factor). However, the mechanisms of this accumulation are different: on cold adaptation sorbitol is produced as a result of reduction of glucose under the influence of aldose reductase, and on intoxication sorbitol is mainly produced from fructose under the influence of sorbitol dehydrogenase. Pathways of the sorbitol accumulation and its re-involvement into metabolism are not always the same, and this might be a mechanism for regulation of carbohydrate metabolism (at the initial stage of glycolysis) on adaptation to unfavorable factors of the environment.

  20. [Pigment accumulation and functional activity of chloroplasts in common Pisum sativum L. mutants with low chlorophyll level (chlorotica)].

    PubMed

    Ladygin, V G

    2003-01-01

    Pea mutants chlorotica 2004 and 2014 with a low content of chlorophyll were studied. The mutant 2004 has light green leaves and stem, and the mutant 2014 has yellow green leaves and stem. They accumulate approximately 80 and 50% chlorophylls of the parent form of pea Torsdag cv. The content of carotene in carotenoids of the mutant 2004 was much lower, and the accumulation of lutein and violaxanthine was increased. The accumulation of all carotenoids in the mutant 2014 decreased almost proportionally to a decrease in the chlorophyll content. The rate of CO2 evolution in mutant chlorotica 2004 and 2014 was established to be lower. The quantum efficiency of photosynthesis in the mutants was 29-30% lower as compared to the control, and in hybrid plants it was 1.5-2-fold higher. It is assumed that the increase in the activity of the night-time respiration in gas exchange of chlorotica mutants and the drop of photosynthesis lead to a decrease in biomass increment. The results obtained allow us to conclude that the mutation of chlorotica 2004 and 2014 affects the genes controlling the formation and functioning of different components of the photosynthetic apparatus.

  1. Effect of TCDD on ACARAT activity and vitamin A accumulation in the kidney of male Sprague-Dawley rats

    SciTech Connect

    Jurek, M.A.; Powers, R.H.; Gilbert, L.C.; Aust, S.D.

    1987-05-01

    Previous studies have shown that rats treated with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) exhibit symptoms of vitamin A deficiency, including hypophagia, failure of normal growth, loss of hepatic vitamin A and accumulation of vitamin A in the kidney. They observed that male Sprague-Dawley rats treated with a single dose of TCDD gained less weight than control rats over a 12 day period. Treated rats showed a progressive loss of hepatic retinyl esters to levels 55% that of control rats. Treated rats accumulated renal vitamin A, with retinyl palmitate levels reaching 5.2x that of control animals, while retinol levels were elevated to 1.5x that of control rats. The ratio of retinyl palmitate to retinol was significantly greater in treated rats than in the control rats. Acyl CoA:Retinol Acyl Transferase (ACARART) activity was 2x greater in kidneys from treated rats, and positively correlated with retinyl palmitate concentrations. They suggest that accumulation of retinyl esters in the kidney occurs as a result of retinol esterification, in response to the TCDD-induced vitamin A deficiency.

  2. Citrus flavonoid, naringenin, increases locomotor activity and reduces diacylglycerol accumulation in skeletal muscle of obese ovariectomized mice

    PubMed Central

    Ke, Jia-Yu; Cole, Rachel M; Hamad, Essam M; Hsiao, Yung-Hsuan; Cotten, Bradley M; Powell, Kimerly A; Belury, Martha A

    2016-01-01

    Scope Estrogen deficiency has been associated with central obesity, muscle loss, and metabolic syndrome in postmenopausal women. This study assessed naringenin accumulation in tissues and investigated the hypothesis that naringenin reverses diet-induced metabolic disturbances in obese ovariectomized mice. Methods and results In study 1, we measured naringenin concentrations in plasma, liver, perigonadal and subcutaneous adipose tissues, and muscle of ovariectomized C57BL/6J female mice after 11 weeks of naringenin supplementation. Naringenin accumulated 5–12 times more in mice fed a 3% naringenin diet than in mice fed a 1% naringenin diet. In study 2, ovariectomized mice were fed a high-fat diet (60 kcal% fat) for 11 weeks and half of the mice were then supplemented with 3% naringenin for another 11 weeks. Dietary naringenin suppressed weight gain, lowered hyperglycemia, and decreased intra-abdominal adiposity evaluated by magnetic resonance imaging. Naringenin-fed mice exhibited elevated locomotor activity monitored by infrared beam breaks, maintained muscle mass, and reduced muscle diacylglycerol content. Real-time PCR analysis in muscle revealed decreased mRNA level for genes involved in de novo lipogenesis, lipolysis, and triglyceride synthesis/storage. Conclusions Long-term 3% naringenin supplementation resulted in significant naringenin accumulation in plasma and tissues, associated with attenuated metabolic dysregulation and muscle loss in obese ovariectomized mice. PMID:26573879

  3. Accumulate-Repeat-Accumulate-Accumulate Codes

    NASA Technical Reports Server (NTRS)

    Divsalar, Dariush; Dolinar, Samuel; Thorpe, Jeremy

    2007-01-01

    Accumulate-repeat-accumulate-accumulate (ARAA) codes have been proposed, inspired by the recently proposed accumulate-repeat-accumulate (ARA) codes. These are error-correcting codes suitable for use in a variety of wireless data-communication systems that include noisy channels. ARAA codes can be regarded as serial turbolike codes or as a subclass of low-density parity-check (LDPC) codes, and, like ARA codes they have projected graph or protograph representations; these characteristics make it possible to design high-speed iterative decoders that utilize belief-propagation algorithms. The objective in proposing ARAA codes as a subclass of ARA codes was to enhance the error-floor performance of ARA codes while maintaining simple encoding structures and low maximum variable node degree.

  4. ELLI-1, a novel germline protein, modulates RNAi activity and P-granule accumulation in Caenorhabditis elegans

    PubMed Central

    Andralojc, Karolina M.; Kelly, Ashley L.; Tanner, Paige C.

    2017-01-01

    Germ cells contain non-membrane bound cytoplasmic organelles that help maintain germline integrity. In C. elegans they are called P granules; without them, the germline undergoes partial masculinization and aberrant differentiation. One key P-granule component is the Argonaute CSR-1, a small-RNA binding protein that antagonizes accumulation of sperm-specific transcripts in developing oocytes and fine-tunes expression of proteins critical to early embryogenesis. Loss of CSR-1 complex components results in a very specific, enlarged P-granule phenotype. In a forward screen to identify mutants with abnormal P granules, ten alleles were recovered with a csr-1 P-granule phenotype, eight of which contain mutations in known components of the CSR-1 complex (csr-1, ego-1, ekl-1, and drh-3). The remaining two alleles are in a novel gene now called elli-1 (enlarged germline granules). ELLI-1 is first expressed in primordial germ cells during mid-embryogenesis, and continues to be expressed in the adult germline. While ELLI-1 forms cytoplasmic aggregates, they occasionally dock, but do not co-localize with P granules. Instead, the majority of ELLI-1 aggregates accumulate in the shared germline cytoplasm. In elli-1 mutants, several genes that promote RNAi and P-granule accumulation are upregulated, and embryonic lethality, sterility, and RNAi resistance in a hypomorphic drh-3 allele is enhanced, suggesting that ELLI-1 functions with CSR-1 to modulate RNAi activity, P-granule accumulation, and post-transcriptional expression in the germline. PMID:28182654

  5. Casitas B-cell lymphoma (Cbl) proteins protect mammary epithelial cells from proteotoxicity of active c-Src accumulation

    PubMed Central

    Mukhopadhyay, Chandrani; Triplett, Aleata; Bargar, Tom; Heckman, Carol; Wagner, Kay-Uwe

    2016-01-01

    Casitas B-cell lymphoma (Cbl) family ubiquitin ligases negatively regulate tyrosine kinase-dependent signal transduction by promoting degradation of active kinases. We and others previously reported that loss of Cbl functions caused hyperproliferation in lymphoid and hematopoietic systems. Unexpectedly, Cbl deletion in Cbl-b–null, Cbl-c–null primary mouse mammary epithelial cells (MECs) (Cbl triple-deficiency) induced rapid cell death despite enhanced MAP kinase and AKT activation. Acute Cbl triple-deficiency elicited distinct transcriptional and biochemical responses with partial overlap with previously described cellular reactions to unfolded proteins and oxidative stress. Although the levels of reactive oxygen species were comparable, detergent-insoluble protein aggregates containing phosphorylated c-Src accumulated in Cbl triple-deficient MECs. Treatment with a broad-spectrum kinase inhibitor dasatinib blocked protein aggregate accumulation and restored in vitro organoid formation. This effect is most likely mediated through c-Src because Cbl triple-deficient MECs were able to form organoids upon shRNA-mediated c-Src knockdown. Taking these data together, the present study demonstrates that Cbl family proteins are required to protect MECs from proteotoxic stress-induced cell death by promoting turnover of active c-Src. PMID:27930322

  6. Plasma Hypoxanthine-Guanine Phosphoribosyl Transferase Activity in Bottlenose Dolphins Contributes to Avoiding Accumulation of Non-recyclable Purines

    PubMed Central

    López-Cruz, Roberto I.; Crocker, Daniel E.; Gaxiola-Robles, Ramón; Bernal, Jaime A.; Real-Valle, Roberto A.; Lugo-Lugo, Orlando; Zenteno-Savín, Tania

    2016-01-01

    Marine mammals are exposed to ischemia/reperfusion and hypoxia/reoxygenation during diving. During oxygen deprivation, adenosine triphosphate (ATP) breakdown implies purine metabolite accumulation, which in humans is associated with pathological conditions. Purine recycling in seals increases in response to prolonged fasting and ischemia. Concentrations of metabolites and activities of key enzymes in purine metabolism were examined in plasma and red blood cells from bottlenose dolphins (Tursiops truncatus) and humans. Hypoxanthine and inosine monophosphate concentrations were higher in plasma from dolphins than humans. Plasma hypoxanthine-guanine phosphoribosyl transferase (HGPRT) activity in dolphins suggests an elevated purine recycling rate, and a mechanism for avoiding accumulation of non-recyclable purines (xanthine and uric acid). Red blood cell concentrations of hypoxanthine, adenosine diphosphate, ATP and guanosine triphosphate were lower in dolphins than in humans; adenosine monophosphate and nicotinamide adenine dinucleotide concentrations were higher in dolphins. HGPRT activity in red blood cells was higher in humans than in dolphins. The lower concentrations of purine catabolism and recycling by-products in plasma from dolphins could be beneficial in providing substrates for recovery of ATP depleted during diving or vigorous swimming. These results suggest that purine salvage in dolphins could be a mechanism for delivering nucleotide precursors to tissues with high ATP and guanosine triphosphate requirements. PMID:27375492

  7. Spontaneous frameshift mutations in Saccharomyces cerevisiae: accumulation during DNA replication and removal by proofreading and mismatch repair activities.

    PubMed Central

    Greene, C N; Jinks-Robertson, S

    2001-01-01

    The accumulation of frameshift mutations during DNA synthesis is determined by the rate at which frameshift intermediates are generated during DNA polymerization and the efficiency with which frameshift intermediates are removed by DNA polymerase-associated exonucleolytic proofreading activity and/or the postreplicative mismatch repair machinery. To examine the relative contributions of these factors to replication fidelity in Saccharomyces cerevisiae, we determined the reversion rates and spectra of the lys2 Delta Bgl +1 frameshift allele. Wild-type and homozygous mutant diploid strains with all possible combinations of defects in the exonuclease activities of DNA polymerases delta and epsilon (conferred by the pol3-01 and pol2-4 alleles, respectively) and in mismatch repair (deletion of MSH2) were analyzed. Although there was no direct correlation between homopolymer run length and frameshift accumulation in the wild-type strain, such a correlation was evident in the triple mutant strain lacking all repair capacity. Furthermore, examination of strains defective in one or two repair activities revealed distinct biases in the removal of the corresponding frameshift intermediates by exonucleolytic proofreading and/or mismatch repair. Finally, these analyses suggest that the mismatch repair machinery may be important for generating some classes of frameshift mutations in yeast. PMID:11560887

  8. Traffic and industrial activities around Riyadh cause the accumulation of heavy metals in legumes: A case study.

    PubMed

    Alyemeni, Mohammed Nasser; Almohisen, Ibrahem A A

    2014-04-01

    The objective of this study was to analyse the effect of the continuously increasing anthropogenic activities around Riyadh, Saudi Arabia on the accumulation of heavy metals in leguminous crops. This study determined whether four legume crops, Pisum sativum L., Vicia faba L., Glycine max and Vigna sinensis, could accumulate the heavy metals Cu, Mn, Pb and Zn in their leaves, pods and grains during the summer when grown under conditions with ambient air pollution from heavy traffic and industrial activities in Riyadh, Saudi Arabia. The effect of the air pollution was examined by quantifying the protein and trace element Cu, Mn, Pb and Zn concentrations in the leaves, pods and grains of the four plant species. Analysis of the results indicated that air pollution significantly increased the heavy metal concentrations in the leaves, pods and grains. Toxic concentrations of the heavy metals were found in the plants grown at L3, L4 and L5. In conclusion, the air pollution increases as the traffic, industrial activities and population density increase.

  9. Activation of the unfolded protein response pathway causes ceramide accumulation in yeast and INS-1E insulinoma cells.

    PubMed

    Epstein, Sharon; Kirkpatrick, Clare L; Castillon, Guillaume A; Muñiz, Manuel; Riezman, Isabelle; David, Fabrice P A; Wollheim, Claes B; Riezman, Howard

    2012-03-01

    Sphingolipids are not only important components of membranes but also have functions in protein trafficking and intracellular signaling. The LCB1 gene encodes a subunit of the serine palmitoyltransferase, which is responsible for the first step of sphingolipid synthesis. Here, we show that activation of the unfolded protein response (UPR) can restore normal ceramide levels and viability in yeast cells with a conditional defect in LCB1. Dependence on UPR was demonstrated by showing the HAC1-dependence of the suppression. A similar induction of ceramides by UPR seems to take place in mammalian cells. In rat pancreatic INS-1E cells, UPR activation induces the transcription of the CerS6 gene, which encodes a ceramide synthase. This correlates with the specific accumulation of ceramide with a C16 fatty acyl chain upon UPR activation. Therefore, our study reveals a novel connection between UPR induction and ceramide synthesis that seems to be conserved between yeast and mammalian cells.

  10. An extraovarian protein accumulated in mosquito oocytes is a carboxypeptidase activated in embryos

    SciTech Connect

    Wenlong Cho; Deitsch, K.W.; Raikhel, A.S. )

    1991-12-01

    The authors report a phenomenon previously unknown for oviparous animals; in Aedes aegypti mosquitoes a serine carboxypeptidase is synthesized extraovarially and then internalized by oocytes. The cDNA encoding mosquito vitellogenic carboxypeptidase (VCP) was cloned and sequenced. The VCP cDNA hybridizes to a 1.5-kilobase mRNA present only in the fat body of vitellogenic females. The deduced amino acid sequence of VCP shares significant homology with members of the serine carboxypeptidase family. Binding assays using a serine protease inhibitor, ({sup 3}H)diisopropyl fluorophosphate, showed that VCP is activated in eggs at the onset of embryonic development. Activation of VCP is associated with the reduction in its size from 53 kDa (inactive proenzyme) to 48 kDa (active enzyme). The active, 48-kDa, form of VCP is maximally present at the middle of embryonic development and disappears by the end.

  11. [Combined effects of copper and simulated acid rain on copper accumulation, growth, and antioxidant enzyme activities of Rumex acetosa].

    PubMed

    He, Shan-Ying; Gao, Yong-Jie; Shentu, Jia-Li; Chen, Kun-Bai

    2011-02-01

    A pot experiment was conducted to study the combined effects of Cu (0-1500 mg x kg(-1)) and simulated acid rain (pH 2.5-5.6) on the copper accumulation, growth, and antioxidant enzyme activities of Rumex acetosa. With the increasing concentration of soil Cu, the Cu accumulation in R. acetosa increased, being higher in root than in stem and leaf. The exposure to low pH acid rain promoted the Cu uptake by R. acetosa. With the increase of soil Cu concentration and/or of acid rain acidity, the biomass of R. acetosa decreased, leaf and root MDA contents increased and had good correlation with soil Cu concentration, and the SOD and POD activities in leaf and root displayed a decreasing trend after an initial increase. This study showed that R. acetosa had a strong adaptive ability to Cu and acid rain stress, exhibiting a high application potential in the remediation of Cu-contaminated soil in acid rain areas.

  12. Free cholesterol accumulation in macrophage membranes activates Toll-like receptors and p38 mitogen-activated protein kinase and induces cathepsin K.

    PubMed

    Sun, Yu; Ishibashi, Minako; Seimon, Tracie; Lee, Mingsum; Sharma, Sudarshana M; Fitzgerald, Katherine A; Samokhin, Andriy O; Wang, Yibin; Sayers, Scott; Aikawa, Masanori; Jerome, W Gray; Ostrowski, Michael C; Bromme, Dieter; Libby, Peter; Tabas, Ira A; Welch, Carrie L; Tall, Alan R

    2009-02-27

    The molecular events linking lipid accumulation in atherosclerotic plaques to complications such as aneurysm formation and plaque disruption are poorly understood. BALB/c-Apoe(-/-) mice bearing a null mutation in the Npc1 gene display prominent medial erosion and atherothrombosis, whereas their macrophages accumulate free cholesterol in late endosomes and show increased cathepsin K (Ctsk) expression. We now show increased cathepsin K immunostaining and increased cysteinyl proteinase activity using near infrared fluorescence imaging over proximal aortas of Apoe(-/-), Npc1(-/-) mice. In mechanistic studies, cholesterol loading of macrophage plasma membranes (cyclodextrin-cholesterol) or endosomal system (AcLDL+U18666A or Npc1 null mutation) activated Toll-like receptor (TLR) signaling, leading to sustained phosphorylation of p38 mitogen-activated protein kinase and induction of p38 targets, including Ctsk, S100a8, Mmp8, and Mmp14. Studies in macrophages from knockout mice showed major roles for TLR4, following plasma membrane cholesterol loading, and for TLR3, after late endosomal loading. TLR signaling via p38 led to phosphorylation and activation of the transcription factor Microphthalmia transcription factor, acting at E-box elements in the Ctsk promoter. These studies suggest that free cholesterol enrichment of either plasma or endosomal membranes in macrophages leads to activation of signaling via various TLRs, prolonged p38 mitogen-activated protein kinase activation, and induction of Mmps, Ctsk, and S100a8, potentially contributing to plaque complications.

  13. The effect of dissolved oxygen on PHB accumulation in activated sludge cultures.

    PubMed

    Third, Katie A; Newland, Mark; Cord-Ruwisch, Ralf

    2003-04-20

    Nitrogen removal from wastewater is often limited by the availability of reducing power to perform denitrification, especially when treating wastewaters with a low carbon:nitrogen ratio. In the increasingly popular sequencing batch reactor (SBR), bacteria have the opportunity to preserve reducing power from incoming chemical oxygen demand (COD) as poly-beta-hydroxybutyrate (PHB). The current study uses laboratory experiments and mathematical modeling in an attempt to generate a better understanding of the effect of oxygen on microbial conversion of COD into PHB. Results from a laboratory SBR with acetate as the organic carbon source showed that the aerobic acetate uptake process was oxygen-dependent, producing higher uptake rates at higher dissolved oxygen (DO) supply rates. However, at the lower DO supply rates (k(L)a 6 to 16 h(-1), 0 mg L(-1) DO), a higher proportion of the substrate was preserved as PHB than at higher DO supply rates (k(L)a 30, 51 h(-1), DO >0.9 mg L(-1)). Up to 77% of the reducing equivalents available from acetate were converted to PHB under oxygen limitation (Y(PHB/Ac) 0.68 Cmol/Cmol), as opposed to only 54% under oxygen-excess conditions (Y(PHB/Ac) 0.48 Cmol/Cmol), where a higher fraction of acetate was used for biomass growth. It was calculated that, by oxygen management during the feast phase, the amount of PHB preserved (1.4 Cmmol L(-1) PHB) accounted for an additional denitrification potential of up to 18 mg L(-1) nitrate-nitrogen. The trends of the effect of oxygen (and hence ATP availability) on PHB accumulation could be reproduced by the simulation model, which was based on biochemical stoichiometry and maximum rates obtained from experiments. Simulated data showed that, at low DO concentrations, the limited availability of adenosine triphosphate (ATP) prevented significant biomass growth and most ATP was used for acetate transport into the cell. In contrast, high DO supply rates provided surplus ATP and hence higher growth rates

  14. Ethylene-Induced Vinblastine Accumulation Is Related to Activated Expression of Downstream TIA Pathway Genes in Catharanthus roseus

    PubMed Central

    Wang, Xi; Pan, Ya-Jie; Chang, Bo-Wen; Hu, Yan-Bo; Guo, Xiao-Rui; Tang, Zhong-Hua

    2016-01-01

    We selected different concentrations of ethephon, to stress C. roseus. We used qRT-PCR and HPLC followed by PCA to obtain comprehensive profiling of the vinblastine biosynthesis in response to ethephon. Based on our findings, the results showed that the high concentration of ethephon had a positive effect at both transcriptional and metabolite level. Meanwhile, there was a remarkable decrease of hydrogen peroxide content and a promoted peroxidase activity in leaves. The loading plot combination with correlation analysis suggested that CrPrx1 could be regarded as a positive regulator and interacts with ethylene response factor (ERF) to play a key role in vinblastine content and peroxidase (POD) activity. This study provides the foundation for a better understanding of the regulation and accumulation of vinblastine in response to ethephon. PMID:27314017

  15. Ethylene-Induced Vinblastine Accumulation Is Related to Activated Expression of Downstream TIA Pathway Genes in Catharanthus roseus.

    PubMed

    Wang, Xi; Pan, Ya-Jie; Chang, Bo-Wen; Hu, Yan-Bo; Guo, Xiao-Rui; Tang, Zhong-Hua

    2016-01-01

    We selected different concentrations of ethephon, to stress C. roseus. We used qRT-PCR and HPLC followed by PCA to obtain comprehensive profiling of the vinblastine biosynthesis in response to ethephon. Based on our findings, the results showed that the high concentration of ethephon had a positive effect at both transcriptional and metabolite level. Meanwhile, there was a remarkable decrease of hydrogen peroxide content and a promoted peroxidase activity in leaves. The loading plot combination with correlation analysis suggested that CrPrx1 could be regarded as a positive regulator and interacts with ethylene response factor (ERF) to play a key role in vinblastine content and peroxidase (POD) activity. This study provides the foundation for a better understanding of the regulation and accumulation of vinblastine in response to ethephon.

  16. HCV core protein induces hepatic lipid accumulation by activating SREBP1 and PPAR{gamma}

    SciTech Connect

    Kim, Kook Hwan; Hong, Sung Pyo; Kim, KyeongJin; Park, Min Jung; Kim, Kwang Jin; Cheong, JaeHun . E-mail: molecule85@pusan.ac.kr

    2007-04-20

    Hepatic steatosis is a common feature in patients with chronic hepatitis C virus (HCV) infection. HCV core protein plays an important role in the development of hepatic steatosis in HCV infection. Because SREBP1 (sterol regulatory element binding protein 1) and PPAR{gamma} (peroxisome proliferators-activated receptor {gamma}) are involved in the regulation of lipid metabolism of hepatocyte, we sought to determine whether HCV core protein may impair the expression and activity of SREBP1 and PPAR{gamma}. In this study, it was demonstrated that HCV core protein increases the gene expression of SREBP1 not only in Chang liver, Huh7, and HepG2 cells transiently transfected with HCV core protein expression plasmid, but also in Chang liver-core stable cells. Furthermore, HCV core protein enhanced the transcriptional activity of SREBP1. In addition, HCV core protein elevated PPAR{gamma} transcriptional activity. However, HCV core protein had no effect on PPAR{gamma} gene expression. Finally, we showed that HCV core protein stimulates the genes expression of lipogenic enzyme and fatty acid uptake associated protein. Therefore, our finding provides a new insight into the mechanism of hepatic steatosis by HCV infection.

  17. Effects of Age and Experience on Physical Activity Accumulation during Kin-Ball

    ERIC Educational Resources Information Center

    Hastie, Peter A.; Langevin, Francois; Wadsworth, Danielle

    2011-01-01

    With a specific agenda of creating a fun activity that emphasized teamwork, cooperation, and sportsmanship, Mario Demers, a Canadian physical education professor, created Kin-Ball in the mid 1980s. The game involves three teams of four players each in which a large ball (4 feet diameter and 2.2 pounds weight (1.22 m and 1 kg, respectively) is sent…

  18. Kinetic Modeling of the Arabidopsis Cryptochrome Photocycle: FADHo Accumulation Correlates with Biological Activity

    PubMed Central

    Procopio, Maria; Link, Justin; Engle, Dorothy; Witczak, Jacques; Ritz, Thorsten; Ahmad, Margaret

    2016-01-01

    Cryptochromes are flavoprotein photoreceptors with multiple signaling roles during plant de-etiolation and development. Arabidopsis cryptochromes (cry1 and cry2) absorb light through an oxidized flavin (FADox) cofactor which undergoes reduction to both FADH° and FADH− redox states. Since the FADH° redox state has been linked to biological activity, it is important to estimate its concentration formed upon illumination in vivo. Here we model the photocycle of isolated cry1 and cry2 proteins with a three-state kinetic model. Our model fits the experimental data for flavin photoconversion in vitro for both cry1 and cry2, providing calculated quantum yields which are significantly lower in cry1 than for cry2. The model was applied to the cryptochrome photocycle in vivo using biological activity in plants as a readout for FADH° concentration. The fit to the in vivo data provided quantum yields for cry1 and cry2 flavin reduction similar to those obtained in vitro, with decreased cry1 quantum yield as compared to cry2. These results validate our assumption that FADH° concentration correlates with biological activity. This is the first reported attempt at kinetic modeling of the cryptochrome photocycle in relation to macroscopic signaling events in vivo, and thereby provides a theoretical framework to the components of the photocycle that are necessary for cryptochrome response to environmental signals. PMID:27446119

  19. [Changed accumulation of active ingredient in different localities and growth period of Hemsleya zhejiangensis (Cucurbitaceae)].

    PubMed

    Yang, Wang-Wei; Lei, Zu-Pei; Wang, Wei-Min; Liang, Wei-qing; Zhou, Wei-Qing; Jin, Xiao-Feng

    2014-08-01

    In this paper, the content of moisture, ethanol-soluble extractives, total saponins and polysaccharide of different tuber samples of Hemsleya zhejiangensis, from different localities, years and seasons, were detected based upon Chinese Pharmacopoeia 2010 version. The samples of roots, stems and leaves in summer were detected as well. The results are mainly as follows. (1)With tuber quality increasing, the content of total saponins increased and then decreased. The individual quality of tubers getting 594.06 g, the content of total saponins reached the peak. (2) The content of active ingredients in different localities was significantly different, and the population of Wuyanling had the maximum content of total saponins and polysaccharide. (3) The content of active ingredients revealed stability between the years 2012 and 2013, but the content of polysaccharide was significantly different. The content in 2012 was higher than that of 2013. (4) The content of active ingredients reached the peak in autumn, which was the best harvest season. (5) Among different component content detection of nutritional organs, tubers had the maximum content of ethanol-soluble extractives, total saponins and polysaccharide. Leaves also contained higher content of ethanol-soluble extractives and total saponins than roots and stems. All of these provide theoretical basis for plant, harvest and production of H. zhejiangensis, which is an endemic, rare, and endangered medicinal plants.

  20. Geomorphic signatures of glacial activity in the Alba Patera volcanic province: Implications for recent frost accumulation on Mars

    NASA Astrophysics Data System (ADS)

    Sinha, Rishitosh K.; Murty, Sripada V. S.

    2013-08-01

    landforms lying within impact craters on Mars have led to the identification of two mechanisms for their formation: (1) intermittent deposition of atmospherically emplaced snow/ice during past spin-axis/orbital conditions and (2) flow of debris-covered ice-rich deposits. The maximum presence of the young ice/snow-rich features (thermal contraction crack polygons, gullies, arcuate ridges, and lobate debris tongues) was observed on the pole-facing slope, indicating that this slope was the preferred site for ice/snow accumulation (during the last 10 Ma). In this study, we investigated 30 craters lying in the Alba Patera volcanic province in the latitudinal bands between 45°N and 32.4°N. Morphological comparison of the younger ice/snow-rich features in these craters led us to conclude that glacial/periglacial features in Alba Patera are mainly present within pole-facing slopes of craters lying within 45°N-39°N. The craters lying within 40.2°N-40°N did not show any glacial/periglacial features. We suggest that the formation of these young ice/snow-rich features follows the same orientation trends as those of other older (>10 Ma) glacial features (debris-covered ice/snow-rich large deposits at the base of the crater wall) in the region. The present work has revealed that the onset of physical processes that result in the formation of glacial/periglacial landforms is also dependent on the changes in elevation ranges of the investigated craters in Alba Patera. Our results confirm past inferences for accumulation of ice/snow on Mars and suggest that the period of ice/snow accumulation activity in Alba Patera occurred throughout the Amazonian and lasted until the recent past, i.e., 2.1-0.4 Ma.

  1. Linkage of protein kinase C-beta activation and intracellular interleukin-2 accumulation in human naive CD4 T cells.

    PubMed Central

    Hassan, J; Rainsford, E; Reen, D J

    1997-01-01

    A critical role for protein kinase C (PKC) in signal transduction events has been well established. Moreover, studies of regulation in PKC levels suggest participation in mediating long-term cellular functions. Protein kinase C-beta (PKC-beta) has been reported to be involved in interleukin-2 (IL-2) synthesis in T lymphocytes. In this study, the role of PKC-beta in intracellular accumulation of IL-2 was investigated using specific inhibitors. Preincubation with two different PKC inhibitors, one specific for classical isotypes (alpha and beta I) Go6976, and one which inhibits both classical and non-classical isotypes, GF109203X, caused a complete block in cytoplasmic IL-2 accumulation when naive CD4 T cells were stimulated in the presence of CD2+CD28+phorbol myristate acetate (PMA). In contrast, preincubation with up to 1000 ng/ml of cyclosporin A (CsA) resulted in a reduction in the intracellular IL-2 detected, as observed by a decrease in the proportion of positive cells as well as a fall in the mean fluorescence intensity (MFI). CsA did not influence PKC-beta translocation. Flow cytometric assessments of PKC-beta and its isoforms beta I and beta II correlated with Western blotting analysis and these results were further supported by the use of PKC-beta-positive (HUT 78) and -negative (BW5147) T-cell lines. Using the specific inhibitors, Go6976 and GF109203X, the findings in this study suggest that activation and translocation of PKC-beta is critical for accumulation of intracellular IL-2. The influence of CsA in reducing but not blocking IL-2 synthesis is discussed. PMA-induced down-regulation of the CD4 antigen was observed in the presence of Go6976 and but not GF109203X, suggesting regulation by non-classical PKC isoforms. Images Figure 4 PMID:9497487

  2. Effect of self-alkalization on nitrite accumulation in a high-rate denitrification system: Performance, microflora and enzymatic activities.

    PubMed

    Li, Wei; Shan, Xiao-Yu; Wang, Zhi-Yao; Lin, Xiao-Yu; Li, Chen-Xu; Cai, Chao-Yang; Abbas, Ghulam; Zhang, Meng; Shen, Li-Dong; Hu, Zhi-Qiang; Zhao, He-Ping; Zheng, Ping

    2016-01-01

    The self-alkalization of denitrifying automatic circulation (DAC) reactor resulted in a large increase of pH up to 9.20 and caused a tremendous accumulation of nitrite up to 451.1 ± 49.0 mgN L(-1) at nitrate loading rate (NLR) from 35 kgN m(-3) d(-1) to 55 kgN m(-3) d(-1). The nitrite accumulation was greatly relieved even at the same NLR once the pH was maintained at 7.6 ± 0.2 in the system. Enzymatic assays indicated that the long-term bacterial exposure to high pH significantly inhibited the activity of copper type nitrite reductase (NirK) rather than the cytochrome cd1 type nitrite reductase (NirS). The terminal restriction fragment length polymorphism (T-RFLP) analysis revealed that the dominant denitrifying bacteria shifted from the NirS-containing Thauear sp. 27 to the NirK-containing Hyphomicrobium nitrativorans strain NL23 during the self-alkalization. The significant nitrite accumulation in the high-rate denitrification system could be therefore, due to the inhibition of Cu-containing NirK by high pH from the self-alkalization. The results suggest that the NirK-containing H. nitrativorans strain NL23 could be an ideal functional bacterium for the conversion of nitrate to nitrite, i.e. denitritation, which could be combined with anaerobic ammonium oxidation (Anammox) to develop a new process for nitrogen removal from wastewater.

  3. Accumulation of lipochitin oligosaccharides and NodD-activating compounds in an efficient plant--Rhizobium nodulation assay.

    PubMed

    Tak, Teun; van Spronsen, Paulina C; Kijne, Jan W; van Brussel, Anton A N; Boot, Kees J M

    2004-07-01

    During legume plant--Rhizobium spp. interactions, leading to the formation of nitrogen-fixing root nodules, the two major determinants of host plant-specificity are plant-produced nod gene inducers (NodD protein activating compounds) and bacterial lipochitin oligosaccharides (LCOs or Nod factors). In a time course, we describe the accumulation of LCOs in an efficient nodulation assay with Vicia sativa subsp. nigra and Rhizobium leguminosarum, in connection with the presence of NodD-activating compounds in the exudate of V. sativa roots. Relatively small amounts of both LCOs and NodD-activating compounds were found to be required for initiation of nodulation during the first days after inoculation. A strong increase in the amount of NodRlv-V[18:4,Ac] LCOs preceded root infection and nodule primordium formation. In contrast to the situation with non-nodulating rhizobia and nonmitogenic LCOs, the amount of NodD-activating compounds in the culture medium remained small after addition of nodulating rhizobia or mitogenic LCOs. Furthermore, addition of nodulating rhizobia or mitogenic LCOs resulted in nearly complete inhibition of root hair formation and elongation, whereas nonmitogenic LCOs stimulated root hair growth. Retention of NodD-activating compounds in the root may inhibit root hair growth.

  4. meso-Dihydroguaiaretic acid inhibits hepatic lipid accumulation by activating AMP-activated protein kinase in human HepG2 cells.

    PubMed

    Lee, Myoung-Su; Kim, Kyung Jin; Kim, Daeyoung; Lee, Kyung-Eun; Hwang, Jae-Kwan

    2011-01-01

    Hepatic lipid accumulation is a major risk factor for dyslipidemia, nonalcoholic fatty liver disease, and insulin resistance. The present study was conducted to evaluate hypolipidemic effects of meso-dihydroguaiaretic acid (MDA), anti-oxidative and anti-inflammatory compound isolated from the Myristica fragrans HOUTT., by oil red O staining, reverse transcription-polymerase chain reaction (RT-PCR), and Western blot. MDA significantly inhibited insulin-induced hepatic lipid accumulation in a dose-dependent manner. The lipid-lowering effect of MDA was accompanied by increased expression of proteins involved in fatty acid oxidation and decreased expression of lipid synthetic proteins. In addition, MDA activated AMP-activated protein kinase (AMPK) as determined by phosphorylation of acetyl-CoA carboxylase (ACC), a downstream target of AMPK. The effects of MDA on lipogenic protein expression were suppressed by pretreatment with compound C, an AMPK inhibitor. Taken together, these findings show that MDA inhibits insulin-induced lipid accumulation in human HepG2 cells by suppressing expression of lipogenic proteins through AMPK signaling, suggesting a potent lipid-lowering agent.

  5. Double-Edge Sword of Sustained ROCK Activation in Prion Diseases through Neuritogenesis Defects and Prion Accumulation.

    PubMed

    Alleaume-Butaux, Aurélie; Nicot, Simon; Pietri, Mathéa; Baudry, Anne; Dakowski, Caroline; Tixador, Philippe; Ardila-Osorio, Hector; Haeberlé, Anne-Marie; Bailly, Yannick; Peyrin, Jean-Michel; Launay, Jean-Marie; Kellermann, Odile; Schneider, Benoit

    2015-08-01

    In prion diseases, synapse dysfunction, axon retraction and loss of neuronal polarity precede neuronal death. The mechanisms driving such polarization defects, however, remain unclear. Here, we examined the contribution of RhoA-associated coiled-coil containing kinases (ROCK), key players in neuritogenesis, to prion diseases. We found that overactivation of ROCK signaling occurred in neuronal stem cells infected by pathogenic prions (PrPSc) and impaired the sprouting of neurites. In reconstructed networks of mature neurons, PrPSc-induced ROCK overactivation provoked synapse disconnection and dendrite/axon degeneration. This overactivation of ROCK also disturbed overall neurotransmitter-associated functions. Importantly, we demonstrated that beyond its impact on neuronal polarity ROCK overactivity favored the production of PrPSc through a ROCK-dependent control of 3-phosphoinositide-dependent kinase 1 (PDK1) activity. In non-infectious conditions, ROCK and PDK1 associated within a complex and ROCK phosphorylated PDK1, conferring basal activity to PDK1. In prion-infected neurons, exacerbated ROCK activity increased the pool of PDK1 molecules physically interacting with and phosphorylated by ROCK. ROCK-induced PDK1 overstimulation then canceled the neuroprotective α-cleavage of normal cellular prion protein PrPC by TACE α-secretase, which physiologically precludes PrPSc production. In prion-infected cells, inhibition of ROCK rescued neurite sprouting, preserved neuronal architecture, restored neuronal functions and reduced the amount of PrPSc. In mice challenged with prions, inhibition of ROCK also lowered brain PrPSc accumulation, reduced motor impairment and extended survival. We conclude that ROCK overactivation exerts a double detrimental effect in prion diseases by altering neuronal polarity and triggering PrPSc accumulation. Eventually ROCK emerges as therapeutic target to combat prion diseases.

  6. Double-Edge Sword of Sustained ROCK Activation in Prion Diseases through Neuritogenesis Defects and Prion Accumulation

    PubMed Central

    Alleaume-Butaux, Aurélie; Nicot, Simon; Pietri, Mathéa; Baudry, Anne; Dakowski, Caroline; Tixador, Philippe; Ardila-Osorio, Hector; Haeberlé, Anne-Marie; Bailly, Yannick; Peyrin, Jean-Michel; Launay, Jean-Marie; Kellermann, Odile; Schneider, Benoit

    2015-01-01

    In prion diseases, synapse dysfunction, axon retraction and loss of neuronal polarity precede neuronal death. The mechanisms driving such polarization defects, however, remain unclear. Here, we examined the contribution of RhoA-associated coiled-coil containing kinases (ROCK), key players in neuritogenesis, to prion diseases. We found that overactivation of ROCK signaling occurred in neuronal stem cells infected by pathogenic prions (PrPSc) and impaired the sprouting of neurites. In reconstructed networks of mature neurons, PrPSc-induced ROCK overactivation provoked synapse disconnection and dendrite/axon degeneration. This overactivation of ROCK also disturbed overall neurotransmitter-associated functions. Importantly, we demonstrated that beyond its impact on neuronal polarity ROCK overactivity favored the production of PrPSc through a ROCK-dependent control of 3-phosphoinositide-dependent kinase 1 (PDK1) activity. In non-infectious conditions, ROCK and PDK1 associated within a complex and ROCK phosphorylated PDK1, conferring basal activity to PDK1. In prion-infected neurons, exacerbated ROCK activity increased the pool of PDK1 molecules physically interacting with and phosphorylated by ROCK. ROCK-induced PDK1 overstimulation then canceled the neuroprotective α-cleavage of normal cellular prion protein PrPC by TACE α-secretase, which physiologically precludes PrPSc production. In prion-infected cells, inhibition of ROCK rescued neurite sprouting, preserved neuronal architecture, restored neuronal functions and reduced the amount of PrPSc. In mice challenged with prions, inhibition of ROCK also lowered brain PrPSc accumulation, reduced motor impairment and extended survival. We conclude that ROCK overactivation exerts a double detrimental effect in prion diseases by altering neuronal polarity and triggering PrPSc accumulation. Eventually ROCK emerges as therapeutic target to combat prion diseases. PMID:26241960

  7. Palmitic Acid Reduces Circulating Bone Formation Markers in Obese Animals and Impairs Osteoblast Activity via C16-Ceramide Accumulation.

    PubMed

    Alsahli, Ahmad; Kiefhaber, Kathryn; Gold, Tziporah; Muluke, Munira; Jiang, Hongfeng; Cremers, Serge; Schulze-Späte, Ulrike

    2016-05-01

    Obesity and impaired lipid metabolism increase circulating and local fatty acid (FA) levels. Our previous studies showed that a high high-saturated -fat diet induced greater bone loss in mice than a high high-unsaturated-fat diet due to increased osteoclast numbers and activity. The impact of elevated FA levels on osteoblasts is not yet clear. We induced obesity in 4 week old male mice using a palmitic acid (PA)- or oleic acid (OA)-enriched high fat high-fat diet (HFD) (20 % of calories from FA), and compared them to mice on a normal (R) caloric diet (10 % of calories from FA). We collected serum to determine FA and bone metabolism marker levels. Primary osteoblasts were isolated; cultured in PA, OA, or control (C) medium; and assessed for mineralization activity, gene expression, and ceramide levels. Obese animals in the PA and OA groups had significantly lower serum levels of bone formation markers P1NP and OC compared to normal weight animals (*p < 0.001), with the lowest marker levels in animals on an PA-enriched HFD (*p < 0.001). Accordingly, elevated levels of PA significantly reduced osteoblast mineralization activity in vitro (*p < 0.05). Elevated PA intake significantly increased C16 ceramide accumulation. This accumulation was preventable through inhibition of SPT2 (serine palmitoyl transferase 2) using myriocin. Elevated levels of PA reduce osteoblast function in vitro and bone formation markers in vivo. Our findings suggest that saturated PA can compromise bone health by affecting osteoblasts, and identify a potential mechanism through which obesity promotes bone loss.

  8. Smoke exposure causes endoplasmic reticulum stress and lipid accumulation in retinal pigment epithelium through oxidative stress and complement activation.

    PubMed

    Kunchithapautham, Kannan; Atkinson, Carl; Rohrer, Bärbel

    2014-05-23

    Age-related macular degeneration (AMD) is a complex disease caused by genetic and environmental factors, including genetic variants in complement components and smoking. Smoke exposure leads to oxidative stress, complement activation, endoplasmic reticulum (ER) stress, and lipid dysregulation, which have all been proposed to be associated with AMD pathogenesis. Here we examine the effects of smoke exposure on the retinal pigment epithelium (RPE). Mice were exposed to cigarette smoke or filtered air for 6 months. RPE cells grown as stable monolayers were exposed to 5% cigarette smoke extract (CSE). Effects of smoke were determined by biochemical, molecular, and histological measures. Effects of the alternative pathway (AP) of complement and complement C3a anaphylatoxin receptor signaling were analyzed using knock-out mice or specific inhibitors. ER stress markers were elevated after smoke exposure in RPE of intact mice, which was eliminated in AP-deficient mice. To examine this relationship further, RPE monolayers were exposed to CSE. Short term smoke exposure resulted in production and release of complement C3, the generation of C3a, oxidative stress, complement activation on the cell membrane, and ER stress. Long term exposure to CSE resulted in lipid accumulation, and secretion. All measures were reversed by blocking C3a complement receptor (C3aR), alternative complement pathway signaling, and antioxidant therapy. Taken together, our results provide clear evidence that smoke exposure results in oxidative stress and complement activation via the AP, resulting in ER stress-mediated lipid accumulation, and further suggesting that oxidative stress and complement act synergistically in the pathogenesis of AMD.

  9. Ultraviolet Radiation-Elicited Enhancement of Isoflavonoid Accumulation, Biosynthetic Gene Expression, and Antioxidant Activity in Astragalus membranaceus Hairy Root Cultures.

    PubMed

    Jiao, Jiao; Gai, Qing-Yan; Wang, Wei; Luo, Meng; Gu, Cheng-Bo; Fu, Yu-Jie; Ma, Wei

    2015-09-23

    In this work, Astragalus membranaceus hairy root cultures (AMHRCs) were exposed to ultraviolet radiation (UV-A, UV-B, and UV-C) for promoting isoflavonoid accumulation. The optimum enhancement for isoflavonoid production was achieved in 34-day-old AMHRCs elicited by 86.4 kJ/m(2) of UV-B. The resulting isoflavonoid yield was 533.54 ± 13.61 μg/g dry weight (DW), which was 2.29-fold higher relative to control (232.93 ± 3.08 μg/g DW). UV-B up-regulated the transcriptional expressions of all investigated genes involved in isoflavonoid biosynthetic pathway. PAL and C4H were found to be two potential key genes that controlled isoflavonoid biosynthesis. Moreover, a significant increase was noted in antioxidant activity of extracts from UV-B-elicited AMHRCs (IC50 values = 0.85 and 1.08 mg/mL) in comparison with control (1.38 and 1.71 mg/mL). Overall, this study offered a feasible elicitation strategy to enhance isoflavonoid accumulation in AMHRCs and also provided a basis for metabolic engineering of isoflavonoid biosynthesis in the future.

  10. Acidobacteria dominate the active bacterial communities of Arctic tundra with widely divergent winter-time snow accumulation and soil temperatures.

    PubMed

    Männistö, Minna K; Kurhela, Emilia; Tiirola, Marja; Häggblom, Max M

    2013-04-01

    The timing and extent of snow cover is a major controller of soil temperature and hence winter-time microbial activity and plant diversity in Arctic tundra ecosystems. To understand how snow dynamics shape the bacterial communities, we analyzed the bacterial community composition of windswept and snow-accumulating shrub-dominated tundra heaths of northern Finland using DNA- and RNA-based 16S rRNA gene community fingerprinting (terminal restriction fragment polymorphism) and clone library analysis. Members of the Acidobacteria and Proteobacteria dominated the bacterial communities of both windswept and snow-accumulating habitats with the most abundant phylotypes corresponding to subdivision (SD) 1 and 2 Acidobacteria in both the DNA- and RNA-derived community profiles. However, different phylotypes within Acidobacteria were found to dominate at different sampling dates and in the DNA- vs. RNA-based community profiles. The results suggest that different species within SD1 and SD2 Acidobacteria respond to environmental conditions differently and highlight the wide functional diversity of these organisms even within the SD level. The acidic tundra soils dominated by ericoid shrubs appear to select for diverse stress-tolerant Acidobacteria that are able to compete in the nutrient poor, phenolic-rich soils. Overall, these communities seem stable and relatively insensitive to the predicted changes in the winter-time snow cover.

  11. Effects of sludge retention time, carbon and initial biomass concentrations on selection process: From activated sludge to polyhydroxyalkanoate accumulating cultures.

    PubMed

    Chen, Zhiqiang; Huang, Long; Wen, Qinxue; Zhang, Huichao; Guo, Zirui

    2017-02-01

    Four sequence batch reactors (SBRs) fed by fermented sugar cane wastewater were continuously operated under the aerobic dynamic feeding (ADF) mode with different configurations of sludge retention time (SRT), carbon and initial biomass concentrations to enrich polyhydroxyalkanoate (PHA) accumulating mixed microbial cultures (MMCs) from municipal activated sludge. The stability of SBRs was investigated besides the enrichment performance. The microbial community structures of the enriched MMCs were analyzed using terminal restriction fragment length polymorphism (T-RFLP). The optimum operating conditions for the enrichment process were: SRT of 5days, carbon concentration of 2.52g COD/L and initial biomass concentration of 3.65g/L. The best enrichment performance in terms of both operating stability and PHA storage ability of enriched cultures (with the maximum PHA content and PHA storage yield (YPHA/S) of 61.26% and 0.68mg COD/mg COD, respectively) was achieved under this condition. Effects of the SRT, carbon concentration and initial biomass concentration on the PHA accumulating MMCs selection process were discussed respectively. A new model including the segmentation of the enrichment process and the effects of SRT on each phase was proposed.

  12. Effect of magnetic field on the accumulation of polyhydroxyalkanoates (PHAs) by microorganism in activated sludge.

    PubMed

    Zhu, Sai-Chang; Xu, Zhen-Lan; Meng, Hui-Juan; Zhou, Jun; Chen, Hong

    2012-08-01

    The effect of static magnetic field on polyhydroxyalkanoates (PHAs) syntheses by activated sludge under aerobic dynamic feeding (ADF) was evaluated in sequence batch reactors (SBR), with magnetic field intensities of 42, 21, 11 and 7 millitesla (mT) exposure in the feast, feast-famine and famine periods, respectively, and one control group without magnetic field exposure. Under each level of magnetic field intensity, the effect of magnetic field exposed in the famine period to PHAs syntheses was most significant in comparison with that in the feast or feast-famine period. Maximal hydroxybutyrate (HB) and (HV) yield occurred at 21 and 11 mT, respectively, and the minimal yield occurred at 42 mT during exposure in the famine period. The maximum biodegradable rate constant of PHA was noted at 11 mT during exposure in the famine period.

  13. Growth rate, protein accumulation, and catabolic enzyme activity of skeletal muscles of galliform birds.

    PubMed

    Shea, Russell E; Olson, John M; Ricklefs, Robert E

    2007-01-01

    We measured the mass and several potential indices of functional capacity of the leg and pectoral muscles through 21 d of age in chicks of three species of galliform birds and the domesticated turkey. The study was conducted to test the hypothesis that the growth rate of a tissue is inversely related to its capacity for mature function across species. We measured the proportion of protein and the activities of the catabolic enzymes citrate synthase (CS), pyruvate kinase (PK), and beta -hydroxy-acyl-CoA-dehydrogenase (HOAD) and estimated exponential growth rate (EGR) from growth increments. EGR was negatively related to proportion of protein, PK, and HOAD and positively related to CS activity. In a multiple regression, EGR was uniquely related only to proportion of protein; it was higher in pectoral muscles and increased in this order: wild turkey

  14. Effect of soluble complement receptor-1 on neutrophil accumulation after traumatic brain injury in rats.

    PubMed

    Kaczorowski, S L; Schiding, J K; Toth, C A; Kochanek, P M

    1995-09-01

    As part of the acute inflammatory response, neutrophils accumulate in the central nervous system after injury. Recently, a soluble human recombinant complement receptor (sCR1; BRL 55730; T Cell Sciences, Inc., Cambridge, MA, U.S.A.) has been developed that inhibits the activation of both the classical and the alternative pathways of complement. sCR1 attenuates the effects of the acute inflammatory response in several models of injury outside the central nervous system. The role of complement in traumatic brain injury, however, remains undefined. We hypothesized that treatment with sCR1 would attenuate neutrophil accumulation in the brain after cerebral trauma. Using a randomized, blinded protocol, 18 anesthetized Sprague-Dawley rats were pre-treated with sCR1 or saline (control) at both 2 h and 2 min before trauma (weight drop) to the exposed right parietal cortex. A third dose of sCR1 (or saline) was given 6 h after trauma. Coronal brain sections centered on the site of trauma were obtained at 24 h after trauma and analyzed for myeloperoxidase (MPO) activity as a marker of neutrophil accumulation. Complete blood counts with differential were obtained before treatment with sCR1 and at 24 h after trauma. At 24 h after trauma, brain MPO activity was reduced by 41% in sCR1-treated rats compared with control rats [0.1599 +/- 0.102 versus 0.2712 +/- 0.178 U/g (mean +/- SD); p = 0.02]. The neutrophil count in peripheral blood increased approximately twofold in both groups. Neutrophil accumulation occurring in the brain after trauma is inhibited by sCR1 treatment. This suggests that complement activation is involved in the local inflammatory response to traumatic brain injury and plays an important role in neutrophil accumulation in the injured brain.

  15. HIV-2 genomic RNA accumulates in stress granules in the absence of active translation.

    PubMed

    Soto-Rifo, Ricardo; Valiente-Echeverria, Fernando; Rubilar, Paulina S; Garcia-de-Gracia, Francisco; Ricci, Emiliano P; Limousin, Taran; Décimo, Didier; Mouland, Andrew J; Ohlmann, Théophile

    2014-11-10

    During the post-transcriptional events of the HIV-2 replication cycle, the full-length unspliced genomic RNA (gRNA) is first used as an mRNA to synthesize Gag and Gag-Pol proteins and then packaged into progeny virions. However, the mechanisms responsible for the coordinate usage of the gRNA during these two mutually exclusive events are poorly understood. Here, we present evidence showing that HIV-2 expression induces stress granule assembly in cultured cells. This contrasts with HIV-1, which interferes with stress granules assembly even upon induced cellular stress. Moreover, we observed that the RNA-binding protein and stress granules assembly factor TIAR associates with the gRNA to form a TIAR-HIV-2 ribonucleoprotein (TH2RNP) complex localizing diffuse in the cytoplasm or aggregated in stress granules. Although the assembly of TH2RNP in stress granules did not require the binding of the Gag protein to the gRNA, we observed that increased levels of Gag promoted both translational arrest and stress granule assembly. Moreover, HIV-2 Gag also localizes to stress granules in the absence of a 'packageable' gRNA. Our results indicate that the HIV-2 gRNA is compartmentalized in stress granules in the absence of active translation prior to being selected for packaging by the Gag polyprotein.

  16. Honokiol activates the LKB1–AMPK signaling pathway and attenuates the lipid accumulation in hepatocytes

    SciTech Connect

    Seo, Min Suk; Kim, Jung Hwan; Kim, Hye Jung; Chang, Ki Churl; Park, Sang Won

    2015-04-15

    Honokiol is a bioactive neolignan compound isolated from the species of Magnolia. This study was designed to elucidate the cellular mechanism by which honokiol alleviates the development of non-alcoholic steatosis. HepG2 cells were treated with honokiol for 1 h, and then exposed to 1 mM free fatty acid (FFA) for 24 h to simulate non-alcoholic steatosis in vitro. C57BL/6 mice were fed with a high-fat diet for 28 days, and honokiol (10 mg/kg/day) was daily treated. Honokiol concentration-dependently attenuated intracellular fat overloading and triglyceride (TG) accumulation in FFA-exposed HepG2 cells. These effects were blocked by pretreatment with an AMP-activated protein kinase (AMPK) inhibitor. Honokiol significantly inhibited sterol regulatory element-binding protein-1c (SREBP-1c) maturation and the induction of lipogenic proteins, stearoyl-CoA desaturase-1 (SCD-1) and fatty acid synthase (FAS) in FFA-exposed HepG2 cells, but these effects were blocked by pretreatment of an AMPK inhibitor. Honokiol induced AMPK phosphorylation and subsequent acetyl-CoA carboxylase (ACC) phosphorylation, which were inhibited by genetic deletion of liver kinase B1 (LKB1). Honokiol stimulated LKB1 phosphorylation, and genetic deletion of LKB1 blocked the effect of honokiol on SREBP-1c maturation and the induction of SCD-1 and FAS proteins in FFA-exposed HepG2 cells. Honokiol attenuated the increases in hepatic TG and lipogenic protein levels and fat accumulation in the mice fed with high-fat diet, while significantly induced LKB1 and AMPK phosphorylation. Taken together, our findings suggest that honokiol has an anti-lipogenic effect in hepatocytes, and this effect may be mediated by the LKB1–AMPK signaling pathway, which induces ACC phosphorylation and inhibits SREBP-1c maturation in hepatocytes. - Highlights: • Honokiol attenuates lipid accumulation induced by free fatty acid in hepatocyte. • Honokiol inhibits the increase in lipogenic enzyme levels induced by free fatty

  17. {delta}-ALAD activity variations in red blood cells in response to lead accumulation in rock doves (Columba livia)

    SciTech Connect

    Gonzalez, M.; Tejedor, M.C.

    1992-10-01

    The enzyme {delta}-aminolevulinic acid dehydratase ({delta}-ALAD, E.C. 4.2.1.24), catalyses the second step of the haeme biosynthetic pathway and is required to maintain the haemoglobin and cytochrome content in red cells. {delta}-ALAD is not only found in bone marrow cells, the major site of haeme synthesis, but also in circulating erythrocytes and other tissues. An inverse correlation was found between {delta}-ALAD activity in red blood cells and lead concentration in the blood. The degree of {delta}-ALAD inhibition in erythrocytes has been widely accepted as a standard bioassay to detect acute and chronic lead exposure in humans and in avians. The value of this parameter as an indicator for environmental lead has been often reported in doves and Scanlon. In lead-treated rats, an increase in {delta}-ALAD activity in bone marrow cells and in blood samples was shown by radioimmunoassay at 5 and 9 days after the treatment. Similarly, the amount of {delta}-ALAD seems to be more sensitive to lead in avian species than in mammals, the usefulness of blood {delta}-ALAD activity as an index of lead exposure has already been questioned by Hutton in the pigeon and by Jaffe et al. in humans. The present investigation studied the toxic effects of lead on rock dove red blood cell {delta}-ALAD activity in two situations: in doves treated with lead acetate in the laboratory and in doves exposed to the environment of Alcala de Henares. The final lead blood concentrations were lower in the environmental than in the laboratory doves. {delta}-ALAD activity in bone marrow cells and the relationships between lead accumulation and enzyme activity in red cells, are examined. 20 refs., 5 figs., 1 tab.

  18. A Caleosin-Like Protein with Peroxygenase Activity Mediates Aspergillus flavus Development, Aflatoxin Accumulation, and Seed Infection

    PubMed Central

    Almousally, Ibrahem; Shaban, Mouhnad; Blee, Elizabeth

    2015-01-01

    Caleosins are a small family of calcium-binding proteins endowed with peroxygenase activity in plants. Caleosin-like genes are present in fungi; however, their functions have not been reported yet. In this work, we identify a plant caleosin-like protein in Aspergillus flavus that is highly expressed during the early stages of spore germination. A recombinant purified 32-kDa caleosin-like protein supported peroxygenase activities, including co-oxidation reactions and reduction of polyunsaturated fatty acid hydroperoxides. Deletion of the caleosin gene prevented fungal development. Alternatively, silencing of the gene led to the increased accumulation of endogenous polyunsaturated fatty acid hydroperoxides and antioxidant activities but to a reduction of fungal growth and conidium formation. Two key genes of the aflatoxin biosynthesis pathway, aflR and aflD, were downregulated in the strains in which A. flavus PXG (AfPXG) was silenced, leading to reduced aflatoxin B1 production in vitro. Application of caleosin/peroxygenase-derived oxylipins restored the wild-type phenotype in the strains in which AfPXG was silenced. PXG-deficient A. flavus strains were severely compromised in their capacity to infect maize seeds and to produce aflatoxin. Our results uncover a new branch of the fungal oxylipin pathway and may lead to the development of novel targets for controlling fungal disease. PMID:26116672

  19. Lipoxygenase activity and proline accumulation in leaves and roots of olive trees in response to drought stress.

    PubMed

    Sofo, Adriano; Dichio, Bartolomeo; Xiloyannis, Cristos; Masia, Andrea

    2004-05-01

    The olive tree (Olea europaea L.) is commonly grown in the Mediterranean basin and is able to resist severe and prolonged drought. Levels of proline (PRO) and malondialdehyde (MDA), and the lipoxygenase (LOX) activity were determined in 2-year-old olive plants (cv. 'Coratina') grown in environmental conditions characterized by high temperatures and high photosynthetic photon flux density levels and gradually subjected to a controlled water deficit for 20 days. Before and during the experimental period, leaf and root samples were collected and analysed for PRO and MDA. The levels of PRO increased in parallel with the severity of drought stress in both leaves and roots. Significant increases of LOX activity and MDA content were also observed during the progressive increment of drought stress in both leaf and root tissues. Measurements of transpiration and photosynthetic rate, stomatal conductance and substomatal CO(2) concentration were carried out during the experiment. The accumulation of PRO indicates a possible role of PRO in drought tolerance. The increases of MDA content and LOX activity show that the water deficit is associated with lipid peroxidation mechanisms.

  20. Comparison of antimycobacterial activity of grepafloxacin against Mycobacterium avium with that of levofloxacin: accumulation of grepafloxacin in human macrophages.

    PubMed

    Hirota, M; Totsu, T; Adachi, F; Kamikawa, K; Watanabe, J; Kanegasaki, S; Nakata, K

    2001-03-01

    The bactericidal activity of two new quinolones, grepafloxacin and levofloxacin, against five strains of Mycobacterium avium was investigated in vitro. The minimum inhibitory concentrations (MICs) of these two quinolones, determined by the broth microdilution method, were comparable for all strains tested. In contrast, grepafloxacin suppressed the intracellular growth of all the strains in monocyte-derived macrophages more strongly than levofloxacin, when the cells infected with these strains were incubated for 7 days in the presence of various concentrations of the two new quinolones. To find the reason for the strengthened intracellular killing activity of grepafloxacin, we determined the ratio of the concentration of the new quinolones in the cells to that in the medium (C/M concentration ratio). The C/M concentration ratio of grepafloxacin was increased to 34.7 by 7 days, whereas that of levofloxacin at 7 days was only 12.3. These data suggested that a higher level of intraphagocytic accumulation of grepafloxacin endows it with greater mycobactericidal activity.

  1. Mutagenesis of conserved active site residues of dihydrolipoamide succinyltransferase enhances the accumulation of α-ketoglutarate in Yarrowia lipolytica.

    PubMed

    Guo, Hongwei; Madzak, Catherine; Du, Guocheng; Zhou, Jingwen

    2016-01-01

    α-Ketoglutarate (α-KG) is an important intermediate in the tricarboxylic acid cycle and has broad applications. The mitochondrial ketoglutarate dehydrogenase (KGDH) complex catalyzes the oxidation of α-KG to succinyl-CoA. Disruption of KGDH, which may enhance the accumulation of α-KG theoretically, was found to be lethal to obligate aerobic cells. In this study, individual overexpression of dihydrolipoamide succinyltransferase (DLST), which serves as the inner core of KGDH, decreased overall activity of the enzyme complex. Furthermore, two conserved active site residues of DLST, His419, and Asp423 were identified. In order to determine whether these residues are engaged in enzyme reaction or not, these two conserved residues were individually mutated. Analysis of the kinetic parameters of the enzyme variants provided evidence that the catalytic reaction of DLST depended on residues His419 and Asp423. Overexpression of mutated DLST not only impaired balanced assembly of KGDH, but also disrupted the catalytic integrity of the enzyme complex. Replacement of the Asp423 residue by glutamate increased extracellular α-KG by 40 % to 50 g L(-1) in mutant strain. These observations uncovered catalytic roles of two conserved active site residues of DLST and provided clues for effective metabolic strategies for rational carbon flux control for the enhanced production of α-KG and related bioproducts.

  2. Transgenic tobacco plants expressing siRNA targeted against the Mungbean yellow mosaic virus transcriptional activator protein gene efficiently block the viral DNA accumulation.

    PubMed

    Shanmugapriya, Gnanasekaran; Das, Sudhanshu Sekhar; Veluthambi, Karuppannan

    2015-06-01

    Mungbean yellow mosaic virus (MYMV) is a bipartite begomovirus that infects many pulse crops such as blackgram, mungbean, mothbean, Frenchbean, and soybean. We tested the efficacy of the transgenically expressed intron-spliced hairpin RNA gene of the transcriptional activator protein (hpTrAP) in reducing MYMV DNA accumulation. Tobacco plants transformed with the MYMV hpTrAP gene accumulated 21-22 nt siRNA. Leaf discs of the transgenic plants, agroinoculated with the partial dimers of MYMV, displayed pronounced reduction in MYMV DNA accumulation. Thus, silencing of the TrAP gene, a suppressor of gene silencing, emerged as an effective strategy to control MYMV.

  3. Structural characterization of the isoenzymatic forms of human myeloperoxidase: evaluation of the iron-containing prosthetic group.

    PubMed

    Wright, J; Bastian, N; Davis, T A; Zuo, C; Yoshimoto, S; Orme-Johnson, W H; Tauber, A I

    1990-01-01

    Myeloperoxidase (MPO) from human neutrophils has been purified and found to exist in three isoenzymatic forms, resolved by ion exchange chromatography. In addition to differences in subunit size and cellular compartmentalization of the isoenzymes, differences have been reported in their activity and susceptibility to inhibition. The structural basis of these isoenzymes is unclear; we attempted to further define their functional characteristics and structural identity. First, we measured respective enzymatic activity using a panel of substrates; MPO I was found to have lower activity with some substrates (pyrogallol, guaiacol, potassium iodide [KI]), but similar activity to the other isoenzymes with 4-aminoantipyrine. These studies confirm that MPO I is enzymatically distinct from MPO II and MPO III. Next, we examined the structural basis of these differences by evaluating the iron-containing prosthetic group in each form using electron paramagnetic resonance (EPR) and determination of the pyridine hemochrome. No significant difference between the isoenzymes was noted in these parameters, suggesting that the prosthetic group is the same in each protein. The cause for any difference in enzymatic activity must lie then in variations extrinsic to the heme, and based on previous studies of the gene and protein analysis, the posttranslational modification of MPO must account for these isoenzymatic species.

  4. Oxidation of hydroquinone, 2,3-dimethylhydroquinone and 2,3,5-trimethylhydroquinone by human myeloperoxidase.

    PubMed

    Burner, U; Krapfenbauer, G; Furtmüller, P G; Regelsberger, G; Obinger, C

    2000-01-01

    Myeloperoxidase is very susceptible to reducing radicals because the reduction potential of the ferric/ferrous redox couple is much higher compared with other peroxidases. Semiquinone radicals are known to reduce heme proteins. Therefore, the kinetics and spectra of the reactions of p-hydroquinone, 2,3-dimethylhydroquinone and 2,3,5-trimethylhydroquinone with compounds I and II were investigated using both sequential-mixing stopped-flow techniques and conventional spectrophotometric measurements. At pH 7 and 15 degrees C the rate constants for compound I reacting with p-hydroquinone, 2,3-dimethylhydroquinone and 2,3,5-trimethylhydroquinone were determined to be 5.6+/-0.4 x 10(7) M(-1)s(-1), 1.3+/-0.1 x 10(6) M(-1)s(-1) and 3.1+/-0.3 x 10(6) M(-1)s(-1), respectively. The corresponding reaction rates for compound II reduction were calculated to be 4.5+/-0.3 x 10(6) M(-1)s(-1), 1.9+/-0.1 x 10(5) M(-1)s(-1) and 4.5+/-0.2 x 10(4) M(-1)s(-1), respectively. Semiquinone radicals, produced by compounds I and II in the classical peroxidation cycle, promote compound III (oxymyeloperoxidase) formation. We could monitor formation of ferrous myeloperoxidase as well as its direct transition to compound II by addition of molecular oxygen. Formation of ferrous myeloperoxidase is shown to depend strongly on the reduction potential of the corresponding redox couple benzoquinone/semiquinone. With 2,3-dimethylhydroquinone and 2,3,5-trimethylhydroquinone as substrate, myeloperoxidase is extremely quickly trapped as compound III. These MPO-typical features could have potential in designing specific drugs which inhibit the production of hypochlorous acid and consequently attenuate inflammatory tissue damage.

  5. Cytoplasmic-anti-neutrophil cytoplasmic antibodies targeting myeloperoxidase in Wegener's granulomatosis: a rare phenomenon.

    PubMed

    Venkatesh, Bhavana M; Joshi, Sangeeta; Adhikary, Ranjeeta

    2014-01-01

    Wegener's granulomatosis (WG) patients can rarely have antineutrophil cytoplasmic antibodies (ANCAs) directed against myeloperoxidase (MPO), producing a cytoplasmic pattern on indirect immunofluorescence (IIF). This has important implications in the diagnosis and pathophysiology of the disease. We present to you a report of three cases of WG, demonstrating a cytoplasmic-ANCA pattern on indirect IIF, but directed against MPO. It is necessary to diagnose a patient taking into account both the autoimmune test results and the clinical features.

  6. Selective CB2 receptor activation ameliorates EAE by reducing Th17 differentiation and immune cell accumulation in the CNS

    PubMed Central

    Kong, Weimin; Li, Hongbo; Tuma, Ronald F.; Ganea, Doina

    2013-01-01

    CB2, the cannabinoid receptor expressed primarily on hematopoietic cells and activated microglia, mediates the immunoregulatory functions of cannabinoids. The involvement of CB2 in EAE has been demonstrated by using both endogenous and exogenous ligands. We showed previously that CB2 selective agonists inhibit leukocyte rolling and adhesion to CNS microvasculature and ameliorate clinical symptom in both chronic and remitting-relapsing EAE models. Here we showed that Gp1a, a highly selective CB2 agonist, with a four log higher affinity for CB2 than CB1, reduced clinical scores and facilitated recovery in EAE in conjunction with long term reduction in demyelination and axonal loss. We also established that Gp1a affected EAE through at least two different mechanisms, i.e. an early effect on Th1/Th17 differentiation in peripheral immune organs, and a later effect on the accumulation of pathogenic immune cells in the CNS, associated with reductions in the expression of CNS and T cell chemokine receptors, chemokines and adhesion molecules. This is the first report on the in vivo CB2-mediated Gp1a inhibition of Th17/Th1 differentiation. We also confirmed the Gp1a-induced inhibition of Th17/Th1 differentiation in vitro, both in non-polarizing and polarizing conditions. The CB2-induced inhibition of Th17 differentiation is highly relevant in view of recent studies emphasizing the importance of pathogenic self-reactive Th17 cells in EAE/MS. In addition, the combined effect on Th17 differentiation and immune cell accumulation into the CNS, emphasize the relevance of CB2 selective ligands as potential therapeutic agents in neuroinflammation. PMID:24342422

  7. High concentrations of myeloperoxidase in the equine uterus as an indicator of endometritis.

    PubMed

    Parrilla-Hernandez, Sonia; Ponthier, Jérôme; Franck, Thierry Y; Serteyn, Didier D; Deleuze, Stéfan C

    2014-04-15

    Intraluminal fluid and excessive abnormal hyperedema are regularly used for the diagnosis of endometritis in the mare, which is routinely confirmed by the presence of neutrophils on endometrial smears. Studies show a relation between neutrophils and myeloperoxidase (MPO), an enzyme contained in and released by neutrophils during degranulation or after cell lysis. This enzyme has been found in many fluids and tissues, and associated with different inflammatory pathologies in the horse. The aims of this study were to assess the presence and concentration of MPO in the equine uterus, and to investigate its relation with neutrophils, and other clinical signs of endometritis. Mares (n = 51) were evaluated for the presence of intraluminal fluid and excessive endometrial edema before breeding, and a small volume lavage and cytology samples were obtained. From 69 cycles, supernatant of the uterine flushes was analyzed with a specific equine MPO ELISA assay to measure MPO concentration. Cytology samples were used for the diagnosis of endometritis. Myeloperoxidase was present in the uterus of all estrus mares in highly variable concentrations. Myeloperoxidase concentrations were significantly (P < 0.05) higher in samples with positive cytologies and in the presence of intraluminal fluid. Occasionally, some samples with negative cytologies showed high MPO concentration, but the opposite was never observed. Cycles presenting hyperedema weren't associated with high concentration of MPO, intraluminal fluid, or positive cytology, making it a poor diagnostic tool of endometritis.

  8. Activity in inferior parietal and medial prefrontal cortex signals the accumulation of evidence in a probability learning task.

    PubMed

    d'Acremont, Mathieu; Fornari, Eleonora; Bossaerts, Peter

    2013-01-01

    In an uncertain environment, probabilities are key to predicting future events and making adaptive choices. However, little is known about how humans learn such probabilities and where and how they are encoded in the brain, especially when they concern more than two outcomes. During functional magnetic resonance imaging (fMRI), young adults learned the probabilities of uncertain stimuli through repetitive sampling. Stimuli represented payoffs and participants had to predict their occurrence to maximize their earnings. Choices indicated loss and risk aversion but unbiased estimation of probabilities. BOLD response in medial prefrontal cortex and angular gyri increased linearly with the probability of the currently observed stimulus, untainted by its value. Connectivity analyses during rest and task revealed that these regions belonged to the default mode network. The activation of past outcomes in memory is evoked as a possible mechanism to explain the engagement of the default mode network in probability learning. A BOLD response relating to value was detected only at decision time, mainly in striatum. It is concluded that activity in inferior parietal and medial prefrontal cortex reflects the amount of evidence accumulated in favor of competing and uncertain outcomes.

  9. Arabidopsis V-ATPase activity at the tonoplast is required for efficient nutrient storage but not for sodium accumulation

    PubMed Central

    Krebs, Melanie; Beyhl, Diana; Görlich, Esther; Al-Rasheid, Khaled A. S.; Marten, Irene; Stierhof, York-Dieter; Hedrich, Rainer; Schumacher, Karin

    2010-01-01

    The productivity of higher plants as a major source of food and energy is linked to their ability to buffer changes in the concentrations of essential and toxic ions. Transport across the tonoplast is energized by two proton pumps, the vacuolar H+-ATPase (V-ATPase) and the vacuolar H+-pyrophosphatase (V-PPase); however, their functional relation and relative contributions to ion storage and detoxification are unclear. We have identified an Arabidopsis mutant in which energization of vacuolar transport solely relies on the activity of the V-PPase. The vha-a2 vha-a3 double mutant, which lacks the two tonoplast-localized isoforms of the membrane-integral V-ATPase subunit VHA-a, is viable but shows day-length-dependent growth retardation. Nitrate content is reduced whereas nitrate assimilation is increased in the vha-a2 vha-a3 mutant, indicating that vacuolar nitrate storage represents a major growth-limiting factor. Zinc is an essential micronutrient that is toxic at excess concentrations and is detoxified via a vacuolar Zn2+/H+-antiport system. Accordingly, the double mutant shows reduced zinc tolerance. In the same way the vacuolar Na+/H+-antiport system is assumed to be an important component of the system that removes sodium from the cytosol. Unexpectedly, salt tolerance and accumulation are not affected in the vha-a2 vha-a3 double mutant. In contrast, reduction of V-ATPase activity in the trans-Golgi network/early endosome (TGN/EE) leads to increased salt sensitivity. Taken together, our results show that during gametophyte and embryo development V-PPase activity at the tonoplast is sufficient whereas tonoplast V-ATPase activity is limiting for nutrient storage but not for sodium tolerance during vegetative and reproductive growth. PMID:20133698

  10. Myeloperoxidase: a front-line defender against phagocytosed microorganisms

    PubMed Central

    Klebanoff, Seymour J.; Kettle, Anthony J.; Rosen, Henry; Winterbourn, Christine C.; Nauseef, William M.

    2013-01-01

    Successful immune defense requires integration of multiple effector systems to match the diverse virulence properties that members of the microbial world might express as they initiate and promote infection. Human neutrophils—the first cellular responders to invading microbes—exert most of their antimicrobial activity in phagosomes, specialized membrane-bound intracellular compartments formed by ingestion of microorganisms. The toxins generated de novo by the phagocyte NADPH oxidase and delivered by fusion of neutrophil granules with nascent phagosomes create conditions that kill and degrade ingested microbes. Antimicrobial activity reflects multiple and complex synergies among the phagosomal contents, and optimal action relies on oxidants generated in the presence of MPO. The absence of life-threatening infectious complications in individuals with MPO deficiency is frequently offered as evidence that the MPO oxidant system is ancillary rather than essential for neutrophil-mediated antimicrobial activity. However, that argument fails to consider observations from humans and KO mice that demonstrate that microbial killing by MPO-deficient cells is less efficient than that of normal neutrophils. We present evidence in support of MPO as a major arm of oxidative killing by neutrophils and propose that the essential contribution of MPO to normal innate host defense is manifest only when exposure to pathogens overwhelms the capacity of other host defense mechanisms. PMID:23066164

  11. Insulin Resistance in PCOS Patients Enhances Oxidative Stress and Leukocyte Adhesion: Role of Myeloperoxidase

    PubMed Central

    Victor, Victor M.; Rovira-Llopis, Susana; Bañuls, Celia; Diaz-Morales, Noelia; Martinez de Marañon, Arantxa; Rios-Navarro, Cesar; Alvarez, Angeles; Gomez, Marcelino; Rocha, Milagros; Hernández-Mijares, Antonio

    2016-01-01

    Cardiovascular diseases and oxidative stress are related to polycystic ovary syndrome (PCOS) and insulin resistance (IR). We have evaluated the relationship between myeloperoxidase (MPO) and leukocyte activation in PCOS patients according to homeostatic model assessment of IR (HOMA-IR), and have explored a possible correlation between these factors and endocrine and inflammatory parameters. This was a prospective controlled study conducted in an academic medical center. The study population consisted of 101 PCOS subjects and 105 control subjects. We divided PCOS subjects into PCOS non-IR (HOMA-IR<2.5) and PCOS IR (HOMA-IR>2.5). Metabolic and anthropometric parameters, total and mitochondrial reactive oxygen species (ROS) production, MPO levels, interactions between human umbilical vein endothelial cells and leukocytes, adhesion molecules (E-selectin, ICAM-1 and VCAM-1) and proinflammatory cytokines (IL-6 and TNF-α) were evaluated. Oxidative stress was observed in PCOS patients, in whom there was an increase in total and mitochondrial ROS production and MPO levels. Enhanced rolling flux and adhesion, and a decrease in polymorphonuclear cell rolling velocity were also detected in PCOS subjects. Increases in IL-6 and TNF-α and adhesion molecules (E-selectin, ICAM-1 and VCAM-1) were also observed, particularly in the PCOS IR group, providing evidence that inflammation and oxidative stress are related in PCOS patients. HOMA-IR was positively correlated with hsCRP (p<0.001, r = 0.304), ROS production (p<0.01, r = 0.593), leukocyte rolling flux (p<0.05, r = 0.446), E-selectin (p<0.01, r = 0.436) and IL-6 (p<0.001, r = 0.443). The results show an increase in the rate of ROS and MPO levels in PCOS patients in general, and particularly in those with IR. Inflammation in PCOS induces leukocyte-endothelium interactions and a simultaneous increase in IL-6, TNF-α, E-selectin, ICAM-1 and VCAM-1. These conditions are aggravated by the presence of IR. PMID:27007571

  12. Insulin Resistance in PCOS Patients Enhances Oxidative Stress and Leukocyte Adhesion: Role of Myeloperoxidase.

    PubMed

    Victor, Victor M; Rovira-Llopis, Susana; Bañuls, Celia; Diaz-Morales, Noelia; Martinez de Marañon, Arantxa; Rios-Navarro, Cesar; Alvarez, Angeles; Gomez, Marcelino; Rocha, Milagros; Hernández-Mijares, Antonio

    2016-01-01

    Cardiovascular diseases and oxidative stress are related to polycystic ovary syndrome (PCOS) and insulin resistance (IR). We have evaluated the relationship between myeloperoxidase (MPO) and leukocyte activation in PCOS patients according to homeostatic model assessment of IR (HOMA-IR), and have explored a possible correlation between these factors and endocrine and inflammatory parameters. This was a prospective controlled study conducted in an academic medical center. The study population consisted of 101 PCOS subjects and 105 control subjects. We divided PCOS subjects into PCOS non-IR (HOMA-IR<2.5) and PCOS IR (HOMA-IR>2.5). Metabolic and anthropometric parameters, total and mitochondrial reactive oxygen species (ROS) production, MPO levels, interactions between human umbilical vein endothelial cells and leukocytes, adhesion molecules (E-selectin, ICAM-1 and VCAM-1) and proinflammatory cytokines (IL-6 and TNF-α) were evaluated. Oxidative stress was observed in PCOS patients, in whom there was an increase in total and mitochondrial ROS production and MPO levels. Enhanced rolling flux and adhesion, and a decrease in polymorphonuclear cell rolling velocity were also detected in PCOS subjects. Increases in IL-6 and TNF-α and adhesion molecules (E-selectin, ICAM-1 and VCAM-1) were also observed, particularly in the PCOS IR group, providing evidence that inflammation and oxidative stress are related in PCOS patients. HOMA-IR was positively correlated with hsCRP (p<0.001, r = 0.304), ROS production (p<0.01, r = 0.593), leukocyte rolling flux (p<0.05, r = 0.446), E-selectin (p<0.01, r = 0.436) and IL-6 (p<0.001, r = 0.443). The results show an increase in the rate of ROS and MPO levels in PCOS patients in general, and particularly in those with IR. Inflammation in PCOS induces leukocyte-endothelium interactions and a simultaneous increase in IL-6, TNF-α, E-selectin, ICAM-1 and VCAM-1. These conditions are aggravated by the presence of IR.

  13. Myeloperoxidase-mediated protein lysine oxidation generates 2-aminoadipic acid and lysine nitrile in vivo.

    PubMed

    Lin, Hongqiao; Levison, Bruce S; Buffa, Jennifer A; Huang, Ying; Fu, Xiaoming; Wang, Zeneng; Gogonea, Valentin; DiDonato, Joseph A; Hazen, Stanley L

    2017-03-01

    Recent studies reveal 2-aminoadipic acid (2-AAA) is both elevated in subjects at risk for diabetes and mechanistically linked to glucose homeostasis. Prior studies also suggest enrichment of protein-bound 2-AAA as an oxidative post-translational modification of lysyl residues in tissues associated with degenerative diseases of aging. While in vitro studies suggest redox active transition metals or myeloperoxidase (MPO) generated hypochlorous acid (HOCl) may produce protein-bound 2-AAA, the mechanism(s) responsible for generation of 2-AAA during inflammatory diseases are unknown. In initial studies we observed that traditional acid- or base-catalyzed protein hydrolysis methods previously employed to measure tissue 2-AAA can artificially generate protein-bound 2-AAA from an alternative potential lysine oxidative product, lysine nitrile (LysCN). Using a validated protease-based digestion method coupled with stable isotope dilution LC/MS/MS, we now report protein bound 2-AAA and LysCN are both formed by hypochlorous acid (HOCl) and the MPO/H2O2/Cl(-) system of leukocytes. At low molar ratio of oxidant to target protein N(ε)-lysine moiety, 2-AAA is formed via an initial N(ε)-monochloramine intermediate, which ultimately produces the more stable 2-AAA end-product via sequential generation of transient imine and semialdehyde intermediates. At higher oxidant to target protein N(ε)-lysine amine ratios, protein-bound LysCN is formed via initial generation of a lysine N(ε)-dichloramine intermediate. In studies employing MPO knockout mice and an acute inflammation model, we show that both free and protein-bound 2-AAA, and in lower yield, protein-bound LysCN, are formed by MPO in vivo during inflammation. Finally, both 2-AAA and to lesser extent LysCN are shown to be enriched in human aortic atherosclerotic plaque, a tissue known to harbor multiple MPO-catalyzed protein oxidation products. Collectively, these results show that MPO-mediated oxidation of protein lysyl

  14. Energy release, beam attenuation radiation damage, gas production and accumulation of long-lived activity in Pb, Pb-Bi and Hg targets

    SciTech Connect

    Shubin, Yu.N.

    1996-06-01

    The calculation and analysis of the nuclei concentrations and long-lived residual radioactivity accumulated in Pb, Pb-Bi and Hg targets irradiated by 800 MeV, 30 mA proton beam have been performed. The dominating components to the total radioactivity of radionuclides resulting from fission and spallation reactions and radiative capture by both target nuclei and accumulated radioactive nuclei for various irradiation and cooling times were analyzed. The estimations of spectral component contributions of neutron and proton fluxes to the accumulated activity were carried out. The contributions of fission products to the targets activity and partial activities of main long-lived fission products to the targets activity and partial activities of main long-lived fission products were evaluated. The accumulation of Po isotopes due to reactions induced by secondary alpha-particles were found to be important for the Pb target as compared with two-step radiative capture. The production of Tritium in the targets and its contribution to the total targets activity was considered in detail. It is found that total activities of both targets are close to one another.

  15. Microbial activity promoted with organic carbon accumulation in macroaggregates of paddy soils under long-term rice cultivation

    NASA Astrophysics Data System (ADS)

    Liu, Yalong; Wang, Ping; Ding, Yuanjun; Lu, Haifei; Li, Lianqing; Cheng, Kun; Zheng, Jufeng; Filley, Timothy; Zhang, Xuhui; Zheng, Jinwei; Pan, Genxing

    2016-12-01

    While soil organic carbon (SOC) accumulation and stabilization has been increasingly the focus of ecosystem properties, how it could be linked to soil biological activity enhancement has been poorly assessed. In this study, topsoil samples were collected from a series of rice soils shifted from salt marshes for 0, 50, 100, 300 and 700 years from a coastal area of eastern China. Soil aggregates were fractioned into different sizes of coarse sand (200-2000 µm), fine sand (20-200 µm), silt (2-20 µm) and clay (< 2 µm), using separation with a low-energy dispersion protocol. Soil properties were determined to investigate niche specialization of different soil particle fractions in response to long-term rice cultivation, including recalcitrant and labile organic carbon, microbial diversity of bacterial, archaeal and fungal communities, soil respiration and enzyme activity. The results showed that the mass proportion both of coarse-sand (2000-200 µm) and clay (< 2 µm) fractions increased with prolonged rice cultivation, but the aggregate size fractions were dominated by fine-sand (200-20 µm) and silt (20-2 µm) fractions across the chronosequence. SOC was highly enriched in coarse-sand fractions (40-60 g kg-1) and moderately in clay fractions (20-25 g kg-1), but was depleted in silt fractions (˜ 10 g kg-1). The recalcitrant carbon pool was higher (33-40 % of SOC) in both coarse-sand and clay fractions than in fine-sand and silt fractions (20-29 % of SOC). However, the ratio of labile organic carbon (LOC) to SOC showed a weakly decreasing trend with decreasing size of aggregate fractions. Total soil DNA (deoxyribonucleic acid) content in the size fractions followed a similar trend to that of SOC. Despite the largely similar diversity between the fractions, 16S ribosomal gene abundance of bacteria and of archaeal were concentrated in both coarse-sand and clay fractions. Being the highest generally in coarse-sand fractions, 18S rRNA gene abundance of fungi decreased

  16. Activation of Na+/K+-ATPase attenuates high glucose-induced H9c2 cell apoptosis via suppressing ROS accumulation and MAPKs activities by DRm217.

    PubMed

    Yan, Xiaofei; Xun, Meng; Li, Jing; Wu, Litao; Dou, Xiaojuan; Zheng, Jin

    2016-10-01

    Hyperglycemia is one of the major factors responsible for the myocardial apoptosis and dysfunction in diabetes. Many studies have proved that there is a close relationship between decreased Na(+)/K(+)-ATPase activity and diabetic cardiomyopathy. However, the effect of directly activated Na(+)/K(+)-ATPase on high glucose-induced myocardial injury is still unknown. Here we found that DRm217, a Na(+)/K(+)-ATPase's DR-region specific monoclonal antibody and direct activator, could prevent high glucose-induced H9c2 cell injury, reactive oxygen species (ROS) release, and mitochondrial dysfunction. High glucose-treatment decreased Na(+)/K(+)-ATPase activity and increased intracellular Ca(2+) level, whereas DRm217 increased Na(+)/K(+)-ATPase activity and alleviated Ca(2+) overload. Inhibition of Ca(2+) overload or closing sodium calcium exchanger (NCX channel) could reverse high glucose-induced ROS increasing and cell injury. In addition, DRm217 could significantly attenuate high glucose-induced p38, JNK and ERK1/2 phosphorylation, which were involved in high glucose-induced cell injury and ROS accumulation. Our findings suggest that DRm217 may protect against the deleterious effects of high glucose in the heart. Prevention of high glucose-induced myocardial cell injury by specific Na(+)/K(+)-ATPase activator may be an attractive therapeutic option.

  17. Cadmium accumulation retard activity of functional components of photo assimilation and growth of rice cultivars amended with vermicompost.

    PubMed

    Sebastian, Abin; Prasad, M N V

    2013-01-01

    Cadmium (Cd) uptake mediated alterations in functional components of photo assimilation during conversion of cow dung and poultry cast to vermicompost were studied in two Indian rice cultivars; MO 16 and MTU 7029. It was found that higher amount of Cd accumulate in plants grown in soil amended with vermicompost which in turn damaged functional components in photo assimilation. Enhancement of root growth was recognized as reason for Cd accumulation. Metabolic alterations noticed among plants were not taken place during application of raw materials used for vermicomposting such as cow dung and poultry cast amendment. Rice varieties accumulated Cd differentially where MTU 7029 accumulated more Cd compare to MO 16. It was also noticed that existence of negative correlation between zinc status of the plant and Cd accumulation.

  18. Macrophage migration inhibitory factor (MIF) is rendered enzymatically inactive by myeloperoxidase-derived oxidants but retains its immunomodulatory function.

    PubMed

    Dickerhof, Nina; Schindler, Lisa; Bernhagen, Jürgen; Kettle, Anthony J; Hampton, Mark B

    2015-12-01

    Macrophage migration inhibitory factor (MIF) is an important player in the regulation of the inflammatory response. Elevated plasma MIF is found in sepsis, arthritis, cystic fibrosis and atherosclerosis. Immunomodulatory activities of MIF include the ability to promote survival and recruitment of inflammatory cells and to amplify pro-inflammatory cytokine production. MIF has an unusual nucleophilic N-terminal proline with catalytic tautomerase activity. It remains unclear whether tautomerase activity is required for MIF function, but small molecules that inhibit tautomerase activity also inhibit the pro-inflammatory activities of MIF. A prominent feature of the acute inflammatory response is neutrophil activation and production of reactive oxygen species, including myeloperoxidase (MPO)-derived hypochlorous acid and hypothiocyanous acid. We hypothesized that MPO-derived oxidants would oxidize the N-terminal proline of MIF and alter its biological activity. MIF was exposed to hypochlorous acid and hypothiocyanous acid and the oxidative modifications on MIF were examined by LC-MS/MS. Imine formation and carbamylation was observed on the N-terminal proline in response to MPO-dependent generation of hypochlorous and hypothiocyanous acid, respectively. These modifications led to a complete loss of tautomerase activity. However, modified MIF still increased CXCL-8/IL-8 production by peripheral blood mononuclear cells (PBMCs) and blocked neutrophil apoptosis, indicating that tautomerase activity is not essential for these biological functions. Pre-treatment of MIF with hypochlorous acid protected the protein from covalent modification by the MIF inhibitor 4-iodo-6-phenylpyrimidine (4-IPP). Therefore, oxidant generation at inflammatory sites may protect MIF from inactivation by more disruptive electrophiles, including drugs designed to target the tautomerase activity of MIF.

  19. MKT-077, a novel rhodacyanine dye in clinical trials, exhibits anticarcinoma activity in preclinical studies based on selective mitochondrial accumulation.

    PubMed

    Koya, K; Li, Y; Wang, H; Ukai, T; Tatsuta, N; Kawakami, M; Shishido; Chen, L B

    1996-02-01

    MKT-077 (formerly known as FJ-776) is a newly synthesized, highly water-soluble ( > 200 mg/ml) rhodacyanine dye that exhibits significant antitumor activity in a variety of model systems. In culture, MKT-077 inhibits the growth of five human cancer cell lines (colon carcinoma CX-1, breast carcinoma MCF-7, pancreatic carcinoma (CRL 1420, bladder transitional cell carcinoma EJ, and melanoma LOX) but not monkey kidney CV-1, an indicator cell line for normal epithelial cells. In nude mice, MKT-077 inhibits the growth of s.c. implanted human renal carcinoma A498 and human prostate carcinoma DU145 and prolongs the survival of mice bearing i.p. implanted human melanoma LOX (tumor:control = 344%). Subcellular localization indicates that MKT-077 is taken up and retained by mitochondria, and flow cytometric analysis suggests that CX-1 cells take up MKT-077 to a much greater extent than CV-1 cells. Quantitation of MKT-077 uptake by ethanol extraction shows that CX-1 cells accumulate 65-fold more MKT-077 than do CV-1 cells. MKT-077 is the first delocalized lipophilic cation with a favorable pharmacological and toxicological profile in preclinical studies. MKT-077 is now being investigated in Phase I clinical trials.

  20. Age‐related remodeling of small arteries is accompanied by increased sphingomyelinase activity and accumulation of long‐chain ceramides

    PubMed Central

    Ohanian, Jacqueline; Liao, Aiyin; Forman, Simon P.; Ohanian, Vasken

    2014-01-01

    Abstract The structure and function of large arteries alters with age leading to increased risk of cardiovascular disease. Age‐related large artery remodeling and arteriosclerosis is associated with increased collagen deposition, inflammation, and endothelial dysfunction. Bioactive sphingolipids are known to regulate these processes, and are also involved in aging and cellular senescence. However, less is known about age‐associated alterations in small artery morphology and function or whether changes in arterial sphingolipids occur in aging. We show that mesenteric small arteries from old sheep have increased lumen diameter and media thickness without a change in media to lumen ratio, indicative of outward hypertrophic remodeling. This remodeling occurred without overt changes in blood pressure or pulse pressure indicating it was a consequence of aging per se. There was no age‐associated change in mechanical properties of the arteries despite an increase in total collagen content and deposition of collagen in a thickened intima layer in arteries from old animals. Analysis of the sphingolipid profile showed an increase in long‐chain ceramide (C14–C20), but no change in the levels of sphingosine or sphingosine‐1‐phosphate in arteries from old compared to young animals. This was accompanied by a parallel increase in acid and neutral sphingomyelinase activity in old arteries compared to young. This study demonstrates remodeling of small arteries during aging that is accompanied by accumulation of long‐chain ceramides. This suggests that sphingolipids may be important mediators of vascular aging. PMID:24872355

  1. Effect of a microbiota activator on accumulated ammonium and microbial community structure in a pilot-scale membrane bioreactor.

    PubMed

    Sato, Yuya; Hori, Tomoyuki; Navarro, Ronald R; Ronald, Navarro R; Habe, Hiroshi; Ogata, Atsushi

    2015-01-01

    Microbiota activators (MAs) have been used to improve the reactor performances of biological wastewater treatment processes. In this study, to remove ammonium (NH4(+)) accumulated during the pre-operation of a pilot-scale membrane bioreactor (MBR) under high-organic-loading conditions, an MA was added to the MBR system and the resulting changes in reactor performances and microbial communities were monitored for 12 days. The NH4(+) concentrations in the sludge and effluent decreased (from 427 to 246 mg/L in the sludge (days 1-9)), and mixed liquor suspended solid increased (from 6,793 to 11,283 mg/L (days 1-12)) after the addition of MA. High-throughput Illumina sequencing of 16S rRNA genes revealed that the microbial community structure changed along with the NH4(+) removal resulting from the MA addition. In particular, the relative abundance of an Acidovorax-related operational taxonomic unit (OTU) increased significantly, accounting for approximately 50% of the total microbial population at day 11. In contrast, the ammonia-oxidizing bacteria and archaea showed low abundances (<0.05%), and no anaerobic ammonia oxidizers were detected. These results suggested that the Acidovorax-related OTU was mainly involved in the NH4(+) removal in the MBR, probably due to its ammonia-assimilating metabolism.

  2. An inhibitor of thrombin activated fibrinolysis inhibitor (TAFI) can reduce extracellular matrix accumulation in an in vitro model of glucose induced ECM expansion.

    PubMed

    Atkinson, J M; Pullen, N; Johnson, T S

    2013-06-24

    Chronic kidney disease (CKD) is characterised by the pathological accumulation of extracellular matrix (ECM) proteins leading to progressive kidney scarring via glomerular and tubular basement membrane expansion. Increased ECM synthesis and deposition, coupled with reduced ECM breakdown contribute to the elevated ECM level in CKD. Previous pre-clinical studies have demonstrated that increased plasmin activity has a beneficial effect in the protein overload model of CKD. As plasmin activation is downregulated by the action of the thrombin activated fibrinolytic inhibitor (TAFI), we tested the hypothesis that inhibition of TAFI might increase plasmin activity and reduce ECM accumulation in an in vitro model of glucose induced ECM expansion. Treatment of NRK52E tubular epithelial cells with increasing concentrations of glucose resulted in a 40% increase in TAFI activity, a 38% reduction in plasmin activity and a subsequent increase in ECM accumulation. In this model system, application of the previously reported TAFI inhibitor UK-396082 [(2S)-5-amino-2-[(1-n-propyl-1H-imidazol-4-yl)methyl]pentanoic acid] caused a reduction in TAFI activity, increased plasmin activity and induced a parallel decrease in ECM levels. In contrast, RNAi knockdown of plasmin resulted in an increase in ECM levels. The data presented here indicate that high glucose induces TAFI activity, inhibiting plasmin activation which results in elevated ECM levels in tubular epithelial cells. The results support the hypothesis that UK-396082 is able to reduce TAFI activity, normalising plasmin activity and preventing excess ECM accumulation suggesting that TAFI inhibition may have potential as an anti-scarring strategy in CKD.

  3. Early-successional ectomycorrhizal fungi effectively support extracellular enzyme activities and seedling nitrogen accumulation in mature forests.

    PubMed

    Nicholson, Bailey A; Jones, Melanie D

    2017-04-01

    After stand-replacing disturbance, regenerating conifer seedlings become colonized by different ectomycorrhizal fungi (EMF) than the locally adapted EMF communities present on seedlings in mature forests. We studied whether EMF species that colonized subalpine fir (Abies lasiocarpa) seedlings in clearcuts differed from those that colonized seedlings in adjacent mature forests with respect to mycorrhizoplane extracellular enzyme activities (EEAs) and N status of the seedlings. We tested two alternate hypotheses: (1) that EEAs would differ between the two EMF communities, with higher activities associated with forest-origin communities, and (2) that acclimation to soil environment was considerable enough that EEAs would be determined primarily by the soil type in which the ectomycorrhizas were growing. Naturally colonized fir seedlings were reciprocally transplanted between clearcuts and forests, carrying different EMF communities with them. EEAs were influenced more by destination environment than by EMF community. EEAs were as high in early-successional as in late-successional communities in both destination environments. Buds of clearcut-origin seedlings had the same or higher N contents as forest seedlings after a growing season in either environment. These results indicate that (i) symbiotic EMF and/or their associated microbial communities demonstrate substantial ability to acclimate to new field environments; (ii) the ability to produce organic matter-degrading enzymes is not a trait that necessarily distinguishes early- and late-successional EMF communities in symbiosis; (iii) early-successional EMF are as capable of supporting seedling N accumulation in forest soils as late-successional EMF; and (iv) disturbed ecosystems where early-successional EMF are present should have high resilience for organic matter degradation.

  4. Certain forms of matrix metalloproteinase-9 accumulate in the extracellular space after microdialysis probe implantation and middle cerebral artery occlusion/reperfusion.

    PubMed

    Planas, Anna M; Justicia, Carles; Solé, Sònia; Friguls, Bibiana; Cervera, Alvaro; Adell, Albert; Chamorro, Angel

    2002-08-01

    Matrix metalloproteinases (MMPs) are activated in focal cerebral ischemia. The activation of MMP-9 is involved in blood-brain barrier breakdown and tissue remodeling. The MMPs are released to the extracellular space, but the form and fate of secreted enzymes in brain are unknown. Using microdialysis in vivo, the authors studied whether ischemia-induced MMP-9 in brain tissue was related to free MMP-9 in the extracellular fluid. A microdialysis probe was placed into the right striatum and microdialysis was initiated 24 hours later in controls (n = 7). One hour prior to microdialysis, a group of rats (n = 7) was subjected to 1-hour occlusion of the right middle cerebral artery, followed by reperfusion. Dialysates were collected at discrete time points up to 24 hours, and subjected to zymography and Western blot analysis. The MMP-9 was released after ischemia and accumulated in the extracellular space at 24 hours (P < 0.05). Free MMP-9 forms include mainly the 95-kd proform, and, to a lesser extent, dimers and cleaved active forms (70 kd), but not the 88-kd form found in tissue. Probe implantation and microdialysis increased free MMP-9 in the dialysate. This increase was concomitant with neutrophil infiltration after the mechanical lesion, as myeloperoxidase was found by means of Western blot analysis in the brain hemisphere subjected to microdialysis (P < 0.005), and immunohistochemistry revealed the presence of myeloperoxidase stain surrounding the site of probe implantation. The results suggest that certain forms of MMP-9 are released and accumulate in the extracellular space after brain injury, and that vascular alterations and neutrophil recruitment elicit MMP-9 activation in the brain after focal ischemia and trauma.

  5. Comparison of Patterns of Accumulation of Ribulose Bisphosphate Carboxylase Antigen and Catalytic Activity and Measurement of Antigen Half-Life during the Cell Cycle of Chlorella sorokiniana1

    PubMed Central

    Toman, P. David; Schmidt, Robert R.

    1985-01-01

    By use of specific immunochemical procedures, ribulose-1,5-bisphosphate carboxylase (RuBPCase), antigen and catalytic activity were shown to have coincident step-patterns of accumulation during the cell cycle of Chlorella sorokiniana. Pulse-chase studies, employing radioactive sulfate, were performed during the period of rapid accumulation of enzyme activity and during the period of constant enzyme activity in the cell cycle. No degradation of RuBPCase antigen could be detected during either of these cell cycle periods. Thus, the step-pattern of accumulation of RuBPCase activity resulted from periodic synthesis of an enzyme that was stable under steady-state cell cycle conditions. Although inhibition of protein synthesis by cycloheximide, at different times in the cell cycle in the light, resulted in rapid decay of RuBPCase activity, this loss in activity occurred without detectable loss in enzyme antigen. When synchronous cells were placed into the dark, to slow the rate of protein synthesis in the absence of cycloheximide, the levels of enzyme antigen and activity decreased by 30 and 50%, respectively, during the 10-hour dark period. Thus, in C. sorokiniana changes in RuBPCase activity do not necessarily reflect parallel changes in enzyme antigen, particularly when cell growth is perturbed by changes from steady-state cultural conditions. PMID:16664496

  6. Oxidative inactivation of alpha 1-proteinase inhibitor by alveolar macrophages from healthy smokers requires the presence of myeloperoxidase.

    PubMed

    Wallaert, B; Gressier, B; Aerts, C; Mizon, C; Voisin, C; Mizon, J

    1991-11-01

    The aim of this work was to study the ability of human alveolar macrophages (AM) of 10 healthy smokers to inactivate alpha 1-proteinase inhibitor (alpha 1PI). Purified alpha 1PI was incubated for 45 min, with human alveolar macrophages before and after stimulation by phorbol myristate acetate (PMA) or opsonized zymosan. As a positive control, the same experiments were performed in parallel with blood human neutrophils (PMN). Results are expressed as percentage of inactivation of alpha 1PI as evaluated from its inhibitory activity against porcine pancreatic elastase. A strong correlation (r = 0.99) was shown when inhibitory activity of alpha 1PI was evaluated against porcine pancreatic elastase or human neutrophil elastase. Unstimulated AM (1.57 +/- 0.9%) as well as stimulated AM (PMA: 1 +/- 0.4%; zymosan: 3 +/- 0.6%) were unable to inactivate alpha 1PI. Gel electrophoresis of alpha 1PI demonstrated that AM before or after stimulation induced a slight proteolysis of alpha 1PI, whereas both cleaved and complexed alpha 1PI were found when alpha 1PI was incubated with activated PMN. Both unstimulated (22 +/- 2.6%) and activated PMN (PMA: 91.7 +/- 4.7%; zymosan: 90 +/- 5.5%) were responsible for a significant inactivation of alpha 1PI. Catalase, in contrast to superoxide dismutase, was responsible for a near complete protection of alpha 1PI inactivation by PMN. To better determine the role of PMN secretory products, especially myeloperoxidase (MPO), we also investigated the effect of zymosan-activated PMN supernatants or of purified MPO on the alpha 1PI-AM reaction. MPO assay in PMN supernatants demonstrated that activated neutrophils released significant amounts of MPO (16.8 +/- 4.1 U/ml), whereas MPO was undetectable in activated AM supernatants.(ABSTRACT TRUNCATED AT 250 WORDS)

  7. Citrate Accumulation-Related Gene Expression and/or Enzyme Activity Analysis Combined With Metabolomics Provide a Novel Insight for an Orange Mutant

    PubMed Central

    Guo, Ling-Xia; Shi, Cai-Yun; Liu, Xiao; Ning, Dong-Yuan; Jing, Long-Fei; Yang, Huan; Liu, Yong-Zhong

    2016-01-01

    ‘Hong Anliu’ (HAL, Citrus sinensis cv. Hong Anliu) is a bud mutant of ‘Anliu’ (AL), characterized by a comprehensive metabolite alteration, such as lower accumulation of citrate, high accumulation of lycopene and soluble sugars in fruit juice sacs. Due to carboxylic acid metabolism connects other metabolite biosynthesis and/or catabolism networks, we therefore focused analyzing citrate accumulation-related gene expression profiles and/or enzyme activities, along with metabolic fingerprinting between ‘HAL’ and ‘AL’. Compared with ‘AL’, the transcript levels of citrate biosynthesis- and utilization-related genes and/or the activities of their respective enzymes such as citrate synthase, cytosol aconitase and ATP-citrate lyase were significantly higher in ‘HAL’. Nevertheless, the mitochondrial aconitase activity, the gene transcript levels of proton pumps, including vacuolar H+-ATPase, vacuolar H+-PPase, and the juice sac-predominant p-type proton pump gene (CsPH8) were significantly lower in ‘HAL’. These results implied that ‘HAL’ has higher abilities for citrate biosynthesis and utilization, but lower ability for the citrate uptake into vacuole compared with ‘AL’. Combined with the metabolites-analyzing results, a model was then established and suggested that the reduction in proton pump activity is the key factor for the low citrate accumulation and the comprehensive metabolite alterations as well in ‘HAL’. PMID:27385485

  8. Accumulate Repeat Accumulate Coded Modulation

    NASA Technical Reports Server (NTRS)

    Abbasfar, Aliazam; Divsalar, Dariush; Yao, Kung

    2004-01-01

    In this paper we propose an innovative coded modulation scheme called 'Accumulate Repeat Accumulate Coded Modulation' (ARA coded modulation). This class of codes can be viewed as serial turbo-like codes, or as a subclass of Low Density Parity Check (LDPC) codes that are combined with high level modulation. Thus at the decoder belief propagation can be used for iterative decoding of ARA coded modulation on a graph, provided a demapper transforms the received in-phase and quadrature samples to reliability of the bits.

  9. Endocrine activity of persistent organic pollutants accumulated in human silicone implants--Dosing in vitro assays by partitioning from silicone.

    PubMed

    Gilbert, Dorothea; Mayer, Philipp; Pedersen, Mikael; Vinggaard, Anne Marie

    2015-11-01

    Persistent organic pollutants (POPs) accumulated in human tissues may pose a risk for human health by interfering with the endocrine system. This study establishes a new link between actual human internal POP levels and the endocrine active dose in vitro, applying partitioning-controlled dosing from silicone to the H295R steroidogenesis assay: (1) Measured concentrations of POPs in silicone breast implants were taken from a recent study and silicone disks were loaded according to these measurements. (2) Silicone disks were transferred into H295R cell culture plates in order to control exposure of the adrenal cells by equilibrium partitioning. (3) Hormone production of the adrenal cells was measured as toxicity endpoint. 4-Nonylphenol was used for method development, and the new dosing method was compared to conventional solvent-dosing. The two dosing modes yielded similar dose-dependent hormonal responses of H295R cells. However, with the partitioning-controlled freely dissolved concentrations (Cfree) as dose metrics, dose-response curves were left-shifted by two orders of magnitude relative to spiked concentrations. Partitioning-controlled dosing of POPs resulted in up to 2-fold increases in progestagen and corticosteroid levels at Cfree of individual POPs in or below the femtomolar range. Silicone acted not only as source of the POPs but also as a sorption sink for lipophilic hormones, stimulating the cellular hormone production. Methodologically, the study showed that silicone can be used as reference partitioning phase to transfer in vivo exposure in humans (silicone implants) to in vitro assays (partition-controlled dosing). The main finding was that POPs at the levels at which they are found in humans can interfere with steroidogenesis in a human adrenocortical cell line.

  10. Methyl jasmonate induces lauric acid omega-hydroxylase activity and accumulation of CYP94A1 transcripts but does not affect epoxide hydrolase activities in vicia sativa seedlings

    PubMed

    Pinot; Benveniste; Sala n JP; Durst

    1998-12-01

    Treatment of etiolated Vicia sativa seedlings by the plant hormone methyl jasmonate (MetJA) led to an increase of cytochrome P450 content. Seedlings that were treated for 48 h in a 1 mM solution of MetJA stimulated omega-hydroxylation of 12:0 (lauric acid) 14-fold compared with the control (153 versus 11 pmol min-1 mg-1 protein, respectively). Induction was dose dependent. The increase of activity (2.7-fold) was already detectable after 3 h of treatment. Activity increased as a function of time and reached a steady level after 24 h. Northern-blot analysis revealed that the transcripts coding for CYP94A1, a fatty acid omega-hydroxylase, had already accumulated after 1 h of exposure to MetJA and was maximal between 3 and 6 h. Under the same conditions, a study of the enzymatic hydrolysis of 9,10-epoxystearic acid showed that both microsomal and soluble epoxide hydrolase activities were not affected by MetJA treatment.

  11. Degradation of endothelial cell matrix heparan sulfate proteoglycan by elastase and the myeloperoxidase-H2O2-chloride system.

    PubMed Central

    Klebanoff, S. J.; Kinsella, M. G.; Wight, T. N.

    1993-01-01

    The degradation of the heparan sulfate proteoglycans of subendothelial matrix by neutrophil elastase and the myeloperoxidase-H2O2-chloride system added separately, sequentially, or together at pH 4.5 to 7.5 was determined by the release of lower molecular weight 35S-labeled material. Elastase alone and the myeloperoxidase system alone caused degradation, and when 4-hour exposure to elastase was followed by 15 minutes of exposure to the myeloperoxidase system, the effect was greater than additive. A greater than additive effect was not observed when elastase followed the myeloperoxidase system or the two were added together. Chloride (or sulfate) alone increased the release of 35S-labeled material from elastase-treated matrix, although the effect of 0.1 M chloride was not as great as that observed when an equivalent concentration of chloride was combined with myeloperoxidase and H2O2. The release of these systems at sites of adherence of neutrophils to glomerular basement membrane may contribute to neutrophil-associated proteinuria. Images Figure 1 Figure 2 Figure 5 Figure 7 PMID:8395774

  12. Myeloperoxidase deficiency enhances zymosan phagocytosis associated with up-regulation of surface expression of CD11b in mouse neutrophils.

    PubMed

    Fujimoto, Kenta; Motowaki, Takehiro; Tamura, Naoya; Aratani, Yasuaki

    2016-12-01

    Myeloperoxidase (MPO), a major component of neutrophils, catalyzes the production of hypochlorous acid (HOCl) from hydrogen peroxide and chloride anion. Phagocytosis is a critical event induced by neutrophils for host defense and inflammation. Interestingly, we found that MPO-deficient (MPO(-/-)) neutrophils engulfed larger amounts of zymosan than wild-type neutrophils. Blocking of the CD11b subunit of complement receptor 3 (CR3) as well as inhibition of focal adhesion kinase (FAK) and extracellular signal-regulated kinase (ERK) dramatically reduced zymosan phagocytosis. In contrast, blocking of dectin-1, toll-like receptor 2 (TLR2), or spleen tyrosine kinase (Syk) had no significant effects on phagocytosis. Western blotting analysis showed that inhibition of FAK decreased the phosphorylation of ERK1/2, indicating that ERK1/2 is a downstream regulator of FAK in neutrophils. Importantly, we found that cell surface expression of CD11b and phosphorylation of ERK1/2 was significantly higher in zymosan-stimulated MPO(-/-) neutrophils than in zymosan-stimulated wild-type neutrophils. Pretreatment with the MPO inhibitor 4-aminobenzoic acid hydrazide dramatically enhanced both zymosan phagocytosis and the surface expression of CD11b in wild-type neutrophils, but not in MPO(-/-) neutrophils. Collectively, these results strongly suggest that up-regulation of the CD11b/FAK/ERK signaling pathway due to absence of MPO enhances the zymosan phagocytic activity of mouse neutrophils.

  13. Relationships between Photosynthetically Active Radiation, Nocturnal Acid Accumulation, and CO2 Uptake for a Crassulacean Acid Metabolism Plant, Opuntia ficus-indica1

    PubMed Central

    Nobel, Park S.; Hartsock, Terry L.

    1983-01-01

    The influences of photosynthetically active radiation (PAR) and water status on nocturnal Crassulacean acid metabolism (CAM) were quantitatively examined for a widely cultivated cactus, Opuntia ficus-indica (L.) Miller. When the total daily PAR was maintained at 10 moles photons per square meter per day but the instantaneous PAR level varied, the rate of nocturnal H+ accumulation (tissue acidification) became 90% saturated near 700 micromoles per square meter per second, a PAR level typical for similar light saturation of C3 photosynthesis. The total nocturnal H+ accumulation and CO2 uptake reached 90% of maximum for a total daily PAR of about 22 moles per square meter per day. Light compensation occurred near 0 moles per square meter per day for nocturnal H+ accumulation and 4 moles per square meter per day for CO2 uptake. Above a total daily PAR of 36 moles per square meter per day or for an instantaneous PAR of 1150 micromoles per square meter per second for more than 6 hours, the nocturnal H+ accumulation actually decreased. This inhibition, which occurred at PAR levels just above those occurring in the field, was accompanied by a substantial decrease in chlorophyll content over a 1-week period. A minimum ratio of H+ accumulated to CO2 taken up of 2.5 averaged over the night occurred for a total daily PAR of 31 moles per square meter per day under wet conditions. About 2 to 6 hours into the night under such conditions, a minimum H+-to-CO2 ratio of 2.0 was observed. Under progressively drier conditions, both nocturnal H+ accumulation and CO2 uptake decreased, but the H+-to-CO2 ratio increased. A ratio of two H+ per CO2 is consistent with the H+ production accompanying the conversion of starch to malic acid, and it apparently occurs for O. ficus-indica when CAM CO2 uptake is strongly favored over respiratory activity. PMID:16662802

  14. Role of Myeloperoxidase Oxidants in the Modulation of Cellular Lysosomal Enzyme Function: A Contributing Factor to Macrophage Dysfunction in Atherosclerosis?

    PubMed Central

    Ismael, Fahd O.; Barrett, Tessa J.; Sheipouri, Diba; Brown, Bronwyn E.; Davies, Michael J.; Hawkins, Clare L.

    2016-01-01

    Low-density lipoprotein (LDL) is the major source of lipid within atherosclerotic lesions. Myeloperoxidase (MPO) is present in lesions and forms the reactive oxidants hypochlorous acid (HOCl) and hypothiocyanous acid (HOSCN). These oxidants modify LDL and have been strongly linked with the development of atherosclerosis. In this study, we examined the effect of HOCl, HOSCN and LDL pre-treated with these oxidants on the function of lysosomal enzymes responsible for protein catabolism and lipid hydrolysis in murine macrophage-like J774A.1 cells. In each case, the cells were exposed to HOCl or HOSCN or LDL pre-treated with these oxidants. Lysosomal cathepsin (B, L and D) and acid lipase activities were quantified, with cathepsin and LAMP-1 protein levels determined by Western blotting. Exposure of J774A.1 cells to HOCl or HOSCN resulted in a significant decrease in the activity of the Cys-dependent cathepsins B and L, but not the Asp-dependent cathepsin D. Cathepsins B and L were also inhibited in macrophages exposed to HOSCN-modified, and to a lesser extent, HOCl-modified LDL. No change was seen in cathepsin D activity or the expression of the cathepsin proteins or lysosomal marker protein LAMP-1. The activity of lysosomal acid lipase was also decreased on treatment of macrophages with each modified LDL. Taken together, these results suggest that HOCl, HOSCN and LDL modified by these oxidants could contribute to lysosomal dysfunction and thus perturb the cellular processing of LDL, which could be important during the development of atherosclerosis. PMID:27997605

  15. Decreased cytochrome-c oxidase activity and lack of age-related accumulation of mitochondrial DNA deletions in the brains of schizophrenics

    SciTech Connect

    Cavelier, L.; Jazin, E.E.; Eriksson, I.

    1995-09-01

    Defects in mitochondrial energy production have been implicated in several neurodegenerative disorders, such as Parkinson disease and amyotrophic lateral sclerosis. To study the contribution of mitochondrial defects to Alzheimer disease and schizophrenia, cytochrome-c oxidase (COX) activity and levels of the mtDNA{sup 4977} deletion in postmortem brain tissue specimens of patients were compared with those of asymptomatic age-matched controls. No difference in COX activity was observed between Alzheimer patients and controls in any of five brain regions investigated. In contrast, schizophrenic patients had a 63% reduction of the COX activity in the nucleus caudatus (P<0.0001) and a 43% reduction in the cortex gyrus frontalis (P<0.05) as compared to controls. The average levels of the mtDNA{sup 4977} deletion did not differ significantly between Alzheimer patients and controls, and the deletion followed similar modes of accumulation with age in the two groups. In contrast, no age-related accumulation of mtDNA deletions was found in schizophrenic patients. The reduction in COX activity in schizophrenic patients did not correlate with changes in the total amount of mtDNA or levels of the mtDNA{sup 4977} deletion. The lack of age-related accumulation of the mtDNA{sup 4977} deletion and reduction in COX activity suggest that a mitochondrial dysfunction may be involved in the pathogenesis of schizophrenia. 41 refs., 3 figs., 1 tab.

  16. Distributions and activities of ammonia oxidizing bacteria and polyphosphate accumulating organisms in a pumped-flow biofilm reactor.

    PubMed

    Wu, Guangxue; Nielsen, Michael; Sorensen, Ketil; Zhan, Xinmin; Rodgers, Michael

    2009-10-01

    The spatial distributions and activities of ammonia oxidizing bacteria (AOB) and polyphosphate accumulating organisms (PAOs) were investigated for a novel laboratory-scale sequencing batch pumped-flow biofilm reactor (PFBR) system that was operated for carbon, nitrogen and phosphorus removal. The PFBR comprised of two 16.5l tanks (Reactors 1 and 2), each with a biofilm module of 2m(2) surface area. To facilitate the growth of AOB and PAOs in the reactor biofilms, the influent wastewater was held in Reactor 1 under stagnant un-aerated conditions for 6 h after feeding, and was then pumped over and back between Reactors 1 and 2 for 12 h, creating aerobic conditions in the two reactors during this period; as a consequence, the biofilm in Reactor 2 was in an aerobic environment for almost all the 18.2 h operating cycle. A combination of micro-sensor measurements, molecular techniques, batch experiments and reactor studies were carried out to analyse the performance of the PFBR system. After 100 days operation at a filtered chemical oxygen demand (COD(f)) loading rate of 3.46 g/m(2) per day, the removal efficiencies were 95% COD(f), 87% TN(f) and 74% TP(f). While the PFBR microbial community structure and function were found to be highly diversified with substantial AOB and PAO populations, about 70% of the phosphorus release potential and almost 100% of the nitrification potential were located in Reactors 1 and 2, respectively. Co-enrichment of AOB and PAOs was realized in the Reactor 2 biofilm, where molecular analyses revealed unexpected microbial distributions at micro-scale, with population peaks of AOB in a 100-250 microm deep sub-surface zone and of PAOs in the 0-150 microm surface zone. The micro-distribution of AOB coincided with the position of the nitrification peak identified during micro-sensor analyses. The study demonstrates that enrichment of PAOs can be realized in a constant or near constant aerobic biofilm environment. Furthermore, the findings suggest

  17. Transcriptional activation of a MYB gene controls the tissue-specific anthocyanin accumulation in a purple cauliflower mutant

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Flavonoids such as anthocyanins possess significant health benefits to humans and play important physiological roles in plants. An interesting Purple gene mutation in cauliflower confers an abnormal pattern of anthocyanin accumulation, giving intense purple color in very young leaves, curds, and see...

  18. T-cell accumulation and regulated on activation, normal T cell expressed and secreted upregulation in adipose tissue in obesity

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Obesity is associated with chronic inflammation, which includes increased macrophage accumulation in adipose tissue (AT) and upregulation of chemokines and cytokines. T cells also play important roles in chronic inflammatory diseases such as atherosclerosis but have not been well studied in obesity....

  19. Excess copper induces accumulation of hydrogen peroxide and increases lipid peroxidation and total activity of copper-zinc superoxide dismutase in roots of Elsholtzia haichowensis.

    PubMed

    Zhang, Hongxiao; Xia, Yan; Wang, Guiping; Shen, Zhenguo

    2008-01-01

    The effects of excess copper (Cu) on the accumulation of hydrogen peroxide (H2O2) and antioxidant enzyme activities in roots of the Cu accumulator Elsholtzia haichowensis Sun were investigated. Copper at 100 and 300 microM significantly increased the concentrations of malondialdehyde and H2O2, and the activities of catalase (E.C. 1.11.1.6), ascorbate peroxidase (E.C. 1.11.1.11), guaiacol peroxidase (GPOD, E.C. 1.11.1.7) and superoxide dismutase (SOD, E.C. 1.15.1.1). Isoenzyme pattern and inhibitor studies showed that, among SOD isoforms, only copper-zinc superoxide dismutase (CuZn-SOD) increased. Excess Cu greatly increased the accumulation of superoxide anion (O2 (.-)) and H2O2 in E. haichowensis roots. This study also provides the first cytochemical evidence of an accumulation of H2O2 in the root cell walls as a consequence of Cu treatments. Experiments with diphenyleneiodonium as an inhibitor of NADPH oxidase, 1,2-dihydroxybenzene-3,5-disulphonic acid as an O2 (.-) scavenger, and N-N-diethyldithiocarbamate as an inhibitor of SOD showed that the source of H2O2 in the cell walls could partially be NADPH oxidase. The enzyme can use cytosolic NADPH to produce O2 (.-), which rapidly dismutates to H2O2 by SOD. Apoplastic GPOD and CuZn-SOD activities were induced in roots of E. haichowensis with 100 microM Cu suggesting that these two antioxidant enzymes may be responsible for H2O2 accumulation in the root apoplast.

  20. Defects in leukocyte-mediated initiation of lipid peroxidation in plasma as studied in myeloperoxidase-deficient subjects: systematic identification of multiple endogenous diffusible substrates for myeloperoxidase in plasma.

    PubMed

    Zhang, Renliang; Shen, Zhongzhou; Nauseef, William M; Hazen, Stanley L

    2002-03-01

    More than a decade ago it was demonstrated that neutrophil activation in plasma results in the time-dependent formation of lipid hydroperoxides through an unknown, ascorbate-sensitive pathway. It is now shown that the mechanism involves myeloperoxidase (MPO)-dependent use of multiple low-molecular-weight substrates in plasma, generating diffusible oxidant species. Addition of activated human neutrophils (from healthy subjects) to plasma (50%, vol/vol) resulted in the peroxidation of endogenous plasma lipids by catalase-, heme poison-, and ascorbate-sensitive pathways, as assessed by high-performance liquid chromatography (HPLC) with on-line electrospray ionization tandem mass spectrometric analysis of free and lipid-bound 9-HETE and 9-HODE. In marked contrast, neutrophils isolated from multiple subjects with MPO deficiency failed to initiate peroxidation of plasma lipids, but they did so after supplementation with isolated human MPO. MPO-dependent use of a low-molecular-weight substrate(s) in plasma for initiating lipid peroxidation was illustrated by demonstrating that the filtrate of plasma (10-kd MWt cutoff) could supply components required for low-density lipoprotein lipid peroxidation in the presence of MPO and H(2)O(2). Subsequent HPLC fractionation of plasma filtrate (10-kd MWt cutoff) by sequential column chromatography identified nitrite, tyrosine, and thiocyanate as major endogenous substrates and 17 beta-estradiol as a novel minor endogenous substrate in plasma for MPO in promoting peroxidation of plasma lipids. These results strongly suggest that the MPO-H(2)O(2) system of human leukocytes serves as a physiological mechanism for initiating lipid peroxidation in vivo.

  1. Increased Variation in Adh Enzyme Activity in Drosophila Mutation-Accumulation Experiment Is Not Due to Transposable Elements at the Adh Structural Gene

    PubMed Central

    Aquadro, C. F.; Tachida, H.; Langley, C. H.; Harada, K.; Mukai, T.

    1990-01-01

    We present here a molecular analysis of the region surrounding the structural gene encoding alcohol dehydrogenase (Adh) in 47 lines of Drosophila melanogaster that have each accumulated mutations for 300 generations. While these lines show a significant increase in variation of alcohol dehydrogenase enzyme activity compared to control lines, we found no restriction map variation in a 13-kb region including the complete Adh structural gene and roughly 5 kb of both 5' and 3' sequences. Thus, the rapid accumulation of ADH activity variation after 28,200 allele generations does not appear to have been due to the mobilization of transposable elements into or out of the Adh structural gene region. PMID:1963870

  2. The activity of HYDROPEROXIDE LYASE 1 regulates accumulation of galactolipids containing 12-oxo-phytodienoic acid in Arabidopsis

    PubMed Central

    Nilsson, Anders K.; Fahlberg, Per; Johansson, Oskar N.; Hamberg, Mats; Andersson, Mats X.; Ellerström, Mats

    2016-01-01

    Arabidopsis produces galactolipids containing esters of 12-oxo-phytodienoic acid (OPDA) and dinor-12-oxo-phytodienoic acid (dnOPDA). These lipids are referred to as arabidopsides and accumulate in response to abiotic and biotic stress. We explored the natural genetic variation found in 14 different Arabidopsis accessions to identify genes involved in the formation of arabidopsides. The accession C24 was identified as a poor accumulator of arabidopsides whereas the commonly used accession Col-0 was found to accumulate comparably large amounts of arabidopsides in response to tissue damage. A quantitative trait loci analysis of an F2 population created from a cross between C24 and Col-0 located a region on chromosome four strongly linked to the capacity to form arabidopsides. Expression analysis of HYDROPEROXIDE LYASE 1 (HPL1) showed large differences in transcript abundance between accessions. Transformation of Col-0 plants with the C24 HPL1 allele under transcriptional regulation of the 35S promoter revealed a strong negative correlation between HPL1 expression and arabidopside accumulation after tissue damage, thereby strengthening the view that HPL1 competes with ALLENE OXIDE SYNTHASE (AOS) for lipid-bound hydroperoxide fatty acids. We further show that the last step in the synthesis of galactolipid-bound OPDA and dnOPDA from unstable allene oxides is exclusively enzyme-catalyzed and not the result of spontaneous cyclization. Thus, the results presented here together with previous studies suggest that all steps in arabidopside biosynthesis are enzyme-dependent and apparently all reactions can take place with substrates being esterified to galactolipids. PMID:27422994

  3. Glucose inhibits root meristem growth via ABA INSENSITIVE 5, which represses PIN1 accumulation and auxin activity in Arabidopsis.

    PubMed

    Yuan, Ting-Ting; Xu, Heng-Hao; Zhang, Kun-Xiao; Guo, Ting-Ting; Lu, Ying-Tang

    2014-06-01

    Glucose functions as a hormone-like signalling molecule that modulates plant growth and development in Arabidopsis thaliana. However, the role of glucose in root elongation remains elusive. Our study demonstrates that high concentrations of glucose reduce the size of the root meristem zone by repressing PIN1 accumulation and thereby reducing auxin levels. In addition, we verified the involvement of ABA INSENSITIVE 5 (ABI5) in this process by showing that abi5-1 is less sensitive to glucose than the wild type, whereas glucose induces ABI5 expression and the inducible overexpression of ABI5 reduces the size of the root meristem zone. Furthermore, the inducible overexpression of ABI5 in PIN1::PIN1-GFP plants reduces the level of PIN1-GFP, but glucose reduces the level of PIN1-GFP to a lesser extent in abi5-1 PIN1::PIN1-GFP plants than in the PIN1::PIN1-GFP control, suggesting that ABI5 is involved in glucose-regulated PIN1 accumulation. Taken together, our data suggest that ABI5 functions in the glucose-mediated inhibition of the root meristem zone by repressing PIN1 accumulation, thus leading to reduced auxin levels in roots.

  4. Monoacylglycerol O-acyltransferase 1 is regulated by peroxisome proliferator-activated receptor γ in human hepatocytes and increases lipid accumulation

    SciTech Connect

    Yu, Jung Hwan; Lee, Yoo Jeong; Kim, Hyo Jung; Choi, Hyeonjin; Choi, Yoonjeong; Seok, Jo Woon; Kim, Jae-woo

    2015-05-08

    Monoacylglycerol O-acyltransferase (MGAT) is an enzyme that is involved in triglyceride synthesis by catalyzing the formation of diacylglycerol from monoacylglycerol and fatty acyl CoAs. Recently, we reported that MGAT1 has a critical role in hepatic TG accumulation and that its suppression ameliorates hepatic steatosis in a mouse model. However, the function of MGAT enzymes in hepatic lipid accumulation has not been investigated in humans. Unlike in rodents, MGAT3 as well as MGAT1 and MGAT2 are present in humans. In this study, we evaluated the differences between MGAT subtypes and their association with peroxisome proliferator-activated receptor γ (PPARγ), a regulator of mouse MGAT1 expression. In human primary hepatocytes, basal expression of MGAT1 was lower than that of MGAT2 or MGAT3, but was strongly induced by PPARγ overexpression. A luciferase assay as well as an electromobility shift assay revealed that human MGAT1 promoter activity is driven by PPARγ by direct binding to at least two regions of the promoter in 293T and HepG2 cells. Moreover, siRNA-mediated suppression of MGAT1 expression significantly attenuated lipid accumulation by PPARγ overexpression in HepG2 cells, as evidenced by oil-red-O staining. These results suggest that human MGAT1 has an important role in fatty liver formation as a target gene of PPARγ, and blocking MGAT1 activity could be an efficient therapeutic way to reduce nonalcoholic fatty liver diseases in humans. - Highlights: • PPARγ promotes MGAT1 expression in human primary hepatocytes. • PPARγ directly regulates MGAT1 promoter activity. • Human MGAT1 promoter has at least two PPARγ-binding elements. • Inhibition of MGAT1 expression attenuates hepatic lipid accumulation in humans.

  5. Tlr4-mutant mice are resistant to acute alcohol-induced sterol-regulatory element binding protein activation and hepatic lipid accumulation.

    PubMed

    Zhang, Zhi-Hui; Liu, Xiao-Qian; Zhang, Cheng; He, Wei; Wang, Hua; Chen, Yuan-Hua; Liu, Xiao-Jing; Chen, Xi; Xu, De-Xiang

    2016-09-15

    Previous studies demonstrated that acute alcohol intoxication caused hepatic lipid accumulation. The present study showed that acute alcohol intoxication caused hepatic lipid accumulation in Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic sterol-regulatory element binding protein (SREBP)-1, a transcription factor regulating fatty acid and triglyceride (TG) synthesis, was activated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic Fas, Acc, Scd-1 and Dgat-2, the key genes for fatty acid and TG synthesis, were up-regulated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Additional experiment showed that hepatic MyD88 was elevated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic NF-κB was activated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Moreover, hepatic GSH content was reduced and hepatic MDA level was elevated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic CYP2E1 was elevated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic p67phox and gp91phox, two NADPH oxidase subunits, were up-regulated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Alpha-phenyl-N-t-butylnitrone (PBN), a free radical spin-trapping agent, protected against alcohol-induced hepatic SREBP-1 activation and hepatic lipid accumulation. In conclusion, Tlr4-mutant mice are resistant to acute alcohol-induced hepatic SREBP-1 activation and hepatic lipid accumulation.

  6. Tlr4-mutant mice are resistant to acute alcohol-induced sterol-regulatory element binding protein activation and hepatic lipid accumulation

    PubMed Central

    Zhang, Zhi-Hui; Liu, Xiao-Qian; Zhang, Cheng; He, Wei; Wang, Hua; Chen, Yuan-Hua; Liu, Xiao-Jing; Chen, Xi; Xu, De-Xiang

    2016-01-01

    Previous studies demonstrated that acute alcohol intoxication caused hepatic lipid accumulation. The present study showed that acute alcohol intoxication caused hepatic lipid accumulation in Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic sterol-regulatory element binding protein (SREBP)-1, a transcription factor regulating fatty acid and triglyceride (TG) synthesis, was activated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic Fas, Acc, Scd-1 and Dgat-2, the key genes for fatty acid and TG synthesis, were up-regulated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Additional experiment showed that hepatic MyD88 was elevated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic NF-κB was activated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Moreover, hepatic GSH content was reduced and hepatic MDA level was elevated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic CYP2E1 was elevated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Hepatic p67phox and gp91phox, two NADPH oxidase subunits, were up-regulated in alcohol-treated Tlr4-wild-type mice but not in Tlr4-mutant mice. Alpha-phenyl-N-t-butylnitrone (PBN), a free radical spin-trapping agent, protected against alcohol-induced hepatic SREBP-1 activation and hepatic lipid accumulation. In conclusion, Tlr4-mutant mice are resistant to acute alcohol-induced hepatic SREBP-1 activation and hepatic lipid accumulation. PMID:27627966

  7. Tyrosine decarboxylase activity of enterococci grown in media with different nutritional potential: tyramine and 2-phenylethylamine accumulation and tyrDC gene expression.

    PubMed

    Bargossi, Eleonora; Tabanelli, Giulia; Montanari, Chiara; Lanciotti, Rosalba; Gatto, Veronica; Gardini, Fausto; Torriani, Sandra

    2015-01-01

    The ability to accumulate tyramine and 2-phenylethylamine by two strains of Enterococcus faecalis and two strains Enterococcus faecium was evaluated in two cultural media added or not with tyrosine. All the enterococcal strains possessed a tyrosine decarboxylase (tyrDC) which determined tyramine accumulation in all the conditions tested, independently on the addition of high concentration of free tyrosine. Enterococci differed in rate and level of biogenic amines accumulation. E. faecalis EF37 and E. faecium FC12 produced tyramine in high amount since the exponential growth phase, while 2-phenylethylamine was accumulated when tyrosine was depleted. E. faecium FC12 and E. faecalis ATCC 29212 showed a slower tyraminogenic activity which took place mainly in the stationary phase up to 72 h of incubation. Moreover, E. faecalis ATCC 29212 produced 2-phenylethylamine only in the media without tyrosine added. In BHI added or not with tyrosine the tyrDC gene expression level differed considerably depending on the strains and the growth phase. In particular, the tyrDC gene expression was high during the exponential phase in rich medium for all the strains and subsequently decreased except for E. faecium FC12. Even if tyrDC presence is common among enterococci, this study underlines the extremely variable decarboxylating potential of strains belonging to the same species, suggesting strain-dependent implications in food safety.

  8. Reduction of exportin 6 activity leads to actin accumulation via failure of RanGTP restoration and NTF2 sequestration in the nuclei of senescent cells.

    PubMed

    Park, Su Hyun; Park, Tae Jun; Lim, In Kyoung

    2011-04-15

    We have previously reported that G-actin accumulation in nuclei is a universal phenomenon of cellular senescence. By employing primary culture of human diploid fibroblast (HDF) and stress-induced premature senescence (SIPS), we explored whether the failure of actin export to cytoplasm is responsible for actin accumulation in nuclei of senescent cells. Expression of exportin 6 (Exp6) and small G-protein, Ran, was significantly reduced in the replicative senescence, but not yet in SIPS, whereas nuclear import of actin by cofilin was already increased in SIPS. After treatment of young HDF cells with H(2)O(2), rapid reduction of nuclear RanGTP was observed along with cytoplasmic increase of RanGDP. Furthermore, significantly reduced interaction of Exp6 with RanGTP was found by GST-Exp6 pull-down analysis. Failure of RanGTP restoration was accompanied with inhibition of ATP synthesis and NTF2 sequestration in the nuclei along with accordant change of senescence morphology. Indeed, knockdown of Exp6 expression significantly increased actin molecule in the nuclei of young HDF cells. Therefore, actin accumulation in nuclei of senescent cells is most likely due to the failure of RanGTP restoration with ATP deficiency and NTF2 accumulation in nuclei, which result in the decrease of actin export via Exp6 inactivation, in addition to actin import by cofilin activation.

  9. Tyrosine decarboxylase activity of enterococci grown in media with different nutritional potential: tyramine and 2-phenylethylamine accumulation and tyrDC gene expression

    PubMed Central

    Bargossi, Eleonora; Tabanelli, Giulia; Montanari, Chiara; Lanciotti, Rosalba; Gatto, Veronica; Gardini, Fausto; Torriani, Sandra

    2015-01-01

    The ability to accumulate tyramine and 2-phenylethylamine by two strains of Enterococcus faecalis and two strains Enterococcus faecium was evaluated in two cultural media added or not with tyrosine. All the enterococcal strains possessed a tyrosine decarboxylase (tyrDC) which determined tyramine accumulation in all the conditions tested, independently on the addition of high concentration of free tyrosine. Enterococci differed in rate and level of biogenic amines accumulation. E. faecalis EF37 and E. faecium FC12 produced tyramine in high amount since the exponential growth phase, while 2-phenylethylamine was accumulated when tyrosine was depleted. E. faecium FC12 and E. faecalis ATCC 29212 showed a slower tyraminogenic activity which took place mainly in the stationary phase up to 72 h of incubation. Moreover, E. faecalis ATCC 29212 produced 2-phenylethylamine only in the media without tyrosine added. In BHI added or not with tyrosine the tyrDC gene expression level differed considerably depending on the strains and the growth phase. In particular, the tyrDC gene expression was high during the exponential phase in rich medium for all the strains and subsequently decreased except for E. faecium FC12. Even if tyrDC presence is common among enterococci, this study underlines the extremely variable decarboxylating potential of strains belonging to the same species, suggesting strain-dependent implications in food safety. PMID:25914676

  10. Reduction of exportin 6 activity leads to actin accumulation via failure of RanGTP restoration and NTF2 sequestration in the nuclei of senescent cells

    SciTech Connect

    Park, Su Hyun; Park, Tae Jun; Lim, In Kyoung

    2011-04-15

    We have previously reported that G-actin accumulation in nuclei is a universal phenomenon of cellular senescence. By employing primary culture of human diploid fibroblast (HDF) and stress-induced premature senescence (SIPS), we explored whether the failure of actin export to cytoplasm is responsible for actin accumulation in nuclei of senescent cells. Expression of exportin 6 (Exp6) and small G-protein, Ran, was significantly reduced in the replicative senescence, but not yet in SIPS, whereas nuclear import of actin by cofilin was already increased in SIPS. After treatment of young HDF cells with H{sub 2}O{sub 2}, rapid reduction of nuclear RanGTP was observed along with cytoplasmic increase of RanGDP. Furthermore, significantly reduced interaction of Exp6 with RanGTP was found by GST-Exp6 pull-down analysis. Failure of RanGTP restoration was accompanied with inhibition of ATP synthesis and NTF2 sequestration in the nuclei along with accordant change of senescence morphology. Indeed, knockdown of Exp6 expression significantly increased actin molecule in the nuclei of young HDF cells. Therefore, actin accumulation in nuclei of senescent cells is most likely due to the failure of RanGTP restoration with ATP deficiency and NTF2 accumulation in nuclei, which result in the decrease of actin export via Exp6 inactivation, in addition to actin import by cofilin activation.

  11. Myeloperoxidase acts as a source of free iron during steady-state catalysis by a feedback inhibitory pathway

    PubMed Central

    Maitra, Dhiman; Shaeib, Faten; Abdulhamid, Ibrahim; Abdulridha, Rasha M.; Saed, Ghassan M.; Diamond, Michael P.; Pennathur, Subramaniam; Abu-Soud, Husam M.

    2013-01-01

    Myeloperoxidase (MPO) is a heme-containing enzyme that generates hypochlorous acid (HOCl) from chloride (Cl−) and hydrogen peroxide (H2O2). It is implicated in the pathology of several chronic inflammatory conditions such as cardiovascular and pulmonary diseases and cancer. Recently we have shown that HOCl can destroy the heme prosthetic group of hemoproteins. Here, we investigated whether the HOCl formed during steady-state catalysis is able to destroy the MPO heme moiety and thereby function as a major source of free iron. UV–visible spectra and H2O2-specific electrode measurements recorded during steady-state HOCl synthesis by MPO showed that the degree of MPO heme destruction increased after multiple additions of H2O2 (10 μM), precluding the enzyme from functioning at maximum activity (80–90% inhibition). MPO heme destruction occurred only in the presence of Cl−. Stopped-flow measurements revealed that the HOCl-mediated MPO heme destruction was complex and occurred through transient ferric species whose formation and decay kinetics indicated it participates in heme destruction along with subsequent free iron release. MPO heme depletion was confirmed by the buildup of free iron utilizing the ferrozine assay. Hypochlorous acid, once generated, first equilibrates in the solution as a whole before binding to the heme iron and initiating heme destruction. Eliminating HOCl from the MPO milieu by scavenging HOCl, destabilizing the MPO–Compound I–Cl complex that could be formed during catalysis, and/or inhibiting MPO catalytic activity partially or completely protects MPO from HOCl insults. Collectively, this study elucidates the bidirectional relationship between MPO and HOCl, which highlights the potential role of MPO as a source of free iron. PMID:23624305

  12. Maintenance of luminal pH and protease activity in lysosomes/late endosomes by vacuolar ATPase in chlorpromazine-treated RAW264 cells accumulating phospholipids.

    PubMed

    Hamaguchi, Ryohei; Haginaka, Jun; Tanimoto, Toshiko; Kuroda, Yukihiro

    2014-02-01

    Cationic amphiphilic drugs (CADs) inhibit phospholipases competitively/uncompetitively. It has also been reported that CADs spontaneously accumulate in acidic organelles and increase their luminal pH, which may lead to deactivation of phospholipid-metabolising enzymes, causing cellular phospholipid accumulation. Recently, however, contradictory results have also been reported in that the luminal pH is not increased by CAD treatment. In this study, we examined whether the lysosomal/late endosomal acidic pH was maintained by vacuolar ATPase (v-ATPase) after treatment with chlorpromazine (CPZ) as a model CAD. The activity of lysosomal protease after CPZ treatment was also measured. Oregon Green-dextran-tetramethylrhodamine conjugate was employed to determine the luminal pH of the lysosomes/late endosomes in RAW264 cells. The luminal pH remained acidic after treatment with CPZ for 23 h, and the lysosomal protease activity was not decreased by 5-min CPZ treatment. Co-treatment with CPZ and bafilomycin A1 (v-ATPase inhibitor) raised the luminal pH. These results suggest that the lysosomal/late endosomal pH is not affected by a 23-h CPZ treatment. In addition, lysosomal enzymes presumably maintain their activity when CPZ accumulates. Our results imply that the pH homeostasis in lysosomes/late endosomes is strictly maintained even after a longer treatment with CADs.

  13. Development of a novel transgenic rice with hypocholesterolemic activity via high-level accumulation of the α' subunit of soybean β-conglycinin.

    PubMed

    Cabanos, Cerrone; Kato, Naoki; Amari, Yoshiki; Fujiwara, Keigo; Ohno, Tomoki; Shimizu, Kousuke; Goto, Tsuyoshi; Shimada, Masaya; Kuroda, Masaharu; Masuda, Taro; Takaiwa, Fumio; Utsumi, Shigeru; Nagaoka, Satoshi; Maruyama, Nobuyuki

    2014-08-01

    Soybean 7S globulin, known as β-conglycinin, has been shown to regulate human plasma cholesterol and triglyceride levels. Furthermore, the α' subunit of β-conglycinin has specifically been shown to possess low-density lipoprotein (LDL)-cholesterol-lowering activity. Therefore, accumulation of the α' subunit of β-conglycinin in rice seeds could lead to the production of new functional rice that could promote human health. Herein, we used the low-glutelin rice mutant 'Koshihikari' (var. a123) and suppressed its glutelins and prolamins, the major seed storage proteins of rice, by RNA interference. The accumulation levels of the α' subunit in the lines with suppressed glutelin and prolamin levels were >20 mg in 1 g of rice seeds, which is considerably higher than those in previous studies. Oral administration of the transgenic rice containing the α' subunit exhibited a hypocholesterolemic activity in rats; the serum total cholesterol and LDL cholesterol levels were significantly reduced when compared to those of the control rice (var. a123). The cholesterol-lowering action by transgenic rice accumulating the α' subunit induces a significant increase in fecal bile acid excretion and a tendency to increase in fecal cholesterol excretion. This is the first report that transgenic rice exhibits a hypocholesterolemic activity in rats in vivo by using the β-conglycinin α' subunit.

  14. CD28/B7 Deficiency Attenuates Systolic Overload-Induced Congestive Heart Failure, Myocardial and Pulmonary Inflammation, and Activated T Cell Accumulation in the Heart and Lungs.

    PubMed

    Wang, Huan; Kwak, Dongmin; Fassett, John; Hou, Lei; Xu, Xin; Burbach, Brandon J; Thenappan, Thenappan; Xu, Yawei; Ge, Jun-Bo; Shimizu, Yoji; Bache, Robert J; Chen, Yingjie

    2016-09-01

    The inflammatory response regulates congestive heart failure (CHF) development. T cell activation plays an important role in tissue inflammation. We postulate that CD28 or B7 deficiency inhibits T cell activation and attenuates CHF development by reducing systemic, cardiac, and pulmonary inflammation. We demonstrated that chronic pressure overload-induced end-stage CHF in mice is characterized by profound accumulation of activated effector T cells (CD3(+)CD44(high) cells) in the lungs and a mild but significant increase of these cells in the heart. In knockout mice lacking either CD28 or B7, there was a dramatic reduction in the accumulation of activated effector T cells in both hearts and lungs of mice under control conditions and after transverse aortic constriction. CD28 or B7 knockout significantly attenuated transverse aortic constriction-induced CHF development, as indicated by less increase of heart and lung weight and less reduction of left ventricle contractility. CD28 or B7 knockout also significantly reduced transverse aortic constriction-induced CD45(+) leukocyte, T cell, and macrophage infiltration in hearts and lungs, lowered proinflammatory cytokine expression (such as tumor necrosis factor-α and interleukin-1β) in lungs. Furthermore, CD28/B7 blockade by CTLA4-Ig treatment (250 μg/mouse every 3 days) attenuated transverse aortic constriction-induced T cell activation, left ventricle hypertrophy, and left ventricle dysfunction. Our data indicate that CD28/B7 deficiency inhibits activated effector T cell accumulation, reduces myocardial and pulmonary inflammation, and attenuates the development of CHF. Our findings suggest that strategies targeting T cell activation may be useful in treating CHF.

  15. Development of a Certified Reference Material for myeloperoxidase-anti-neutrophil cytoplasmic autoantibodies (MPO-ANCA).

    PubMed

    Monogioudi, Evanthia; Hutu, Dana Petronela; Martos, Gustavo; Sheldon, Joanna; Schimmel, Heinz; Meroni, Pier Luigi; Zegers, Ingrid

    2016-06-01

    A serum Certified Reference Material (CRM) for supporting reliable autoimmune diagnostics was recently released by the Institute for Reference Materials and Measurements (IRMM) of the Joint Research Centre of the European Commission. It was produced in collaboration with a Working Group on the Harmonisation of Autoimmune Tests of the International Federation of Clinical Chemistry and Laboratory Medicine (IFCC WG-HAT). This material is aimed at facilitating the standardisation of measurements of anti-myeloperoxidase immunoglobulin G antibodies. The CRM could be used as a common calibrant by clinicians and manufacturers thereby significantly improving the comparability of results from commercial immunoassays used for IgG anti-MPO measurements. This paper provides information on the new CRM and its intended use.

  16. [Use of the granulocytic myeloperoxidase release reaction to diagnose food additive allergies].

    PubMed

    Titova, N D

    2011-03-01

    Adverse reactions to food additives are difficult to diagnose due to the diversity of mechanisms involved in their realization and to the absence of reasonably reliable methods for their determination. Eighty-three patients with allergic diseases were examined using the granulocytic myeloperoxidase release reaction (MRR) to diagnose intolerance reactions to food additives (E102, E122, E124, E132, E110, E2111). MRR revealed leukocyte hypersensitivity to tartrazine in 10.8%, sunset yellow in 4.8%, ponceau in 13.2%, indigo carmine in 8.4%, carmoisine and benzoate in 9.6%. The findings were correlated with history data and the levels of IgE antibodies to these dyes. The practical use of the proposed MRR method makes it possible to enhance the accuracy of diagnosis of allergy to food additives.

  17. Carbonic anhydrase activity and photosynthetic rate in the tree species Paulownia tomentosa Steud. Effect of dimethylsulfoxide treatment and zinc accumulation in leaves.

    PubMed

    Lazova, Galia N; Naidenova, Tsveta; Velinova, Katya

    2004-03-01

    The enzyme carbonic anhydrase (CA) (EC 4.2.1.1) catalyzes the reversible conversion of CO2 to HCO3- and has been shown to be involved in photosynthesis. The enzyme has been shown in animals, plants, eubacteria and viruses, but similar reports on the evidence for CA activity in tree plants does not be appear to be available. In the preliminary analyses of the work, the CA activity in leaf extracts from the tree species Paulownia tomentosa Steud. (introduced in Bulgaria) is described. A connection between CA activity and the rate of photosynthetic CO2 fixation is shown. In the second portion of the work, the effect of 10(-4) mol/L and 10(-2) mol/L dimethylsulfoxide (DMSO) on the zinc accumulation in leaves is demonstrated. It is suggested that CA activity is an indicator of the level of physiologically active zinc in leaves of P. tomentosa Steud. A connection between the process of zinc accumulation in leaves and the activity of the enzymes CA and glycolate oxidase (GO) (EC 1.1.3.1) is established.

  18. Myeloperoxidase amplified high glucose-induced endothelial dysfunction in vasculature: Role of NADPH oxidase and hypochlorous acid.

    PubMed

    Tian, Rong; Ding, Yun; Peng, Yi-Yuan; Lu, Naihao

    2017-03-11

    Nicotinamide adenine dinucleotide phosphate (NADPH) oxidase-derived reactive oxygen species (ROS) such as superoxide and hydrogen peroxide (H2O2), have emerged as important molecules in the pathogenesis of diabetic endothelial dysfunction. Additionally, neutrophils-derived myeloperoxidase (MPO) and MPO-catalyzed hypochlorous acid (HOCl) play important roles in the vascular injury. However, it is unknown whether MPO can use vascular-derived ROS to induce diabetic endothelial dysfunction. In the present study, we demonstrated that NADPH oxidase was the main source of ROS formation in high glucose-cultured human umbilical vein endothelial cells (HUVECs), and played a critical role in high glucose-induced endothelial dysfunction such as cell apoptosis, loss of cell viability and reduction of nitric oxide (NO). However, the addition of MPO could amplify the high glucose-induced endothelial dysfunction which was inhibited by the presence of apocynin (NADPH oxidase inhibitor), catalase (H2O2 scavenger), or methionine (HOCl scavenger), demonstrating the contribution of NADPH oxidase-H2O2-MPO-HOCl pathway in the MPO/high glucose-induced vascular injury. In high glucose-incubated rat aortas, MPO also exacerbated the NADPH oxidase-induced impairment of endothelium-dependent relaxation. Consistent with these in vitro data, in diabetic rat aortas, both MPO expresion and NADPH oxidase activity were increased while the endothelial function was simultaneously impaired. The results suggested that vascular-bound MPO could amplify high glucose-induced vascular injury in diabetes. MPO-NADPH oxidase-HOCl may represent an important pathogenic pathway in diabetic vascular diseases.

  19. The vinyl-sulfonium bond in human myeloperoxidase: Impact on compound I formation and reduction by halides and thiocyanate

    SciTech Connect

    Zederbauer, Martina; Furtmueller, Paul Georg; Ganster, Bernadette; Moguilevsky, Nicole; Obinger, Christian . E-mail: christian.obinger@boku.ac.at

    2007-05-04

    In human myeloperoxidase (MPO) the heme is covalently attached to the protein via two ester linkages and a unique sulfonium ion linkage between the sulfur atom of Met243 and the {beta}-carbon of the vinyl ring on pyrrole ring A. Here, we have investigated the variant Met243Val produced in Chinese hamster ovary cells in order to elucidate the role of the electron withdrawing sulfonium bond in compound I formation and reduction. Disruption of this MPO-typical bond causes a blue-shifted UV-vis spectrum and an increase in the heme flexibility. This had no impact on compound I formation mediated by hydrogen peroxide (2.2 x 10{sup 7} M{sup -1} s{sup -1} at pH 7.0 and 25 {sup o}C). Compared with wild-type recombinant MPO the cyanide association rate with ferric Met243Val was significantly enhanced as were also the calculated apparent bimolecular compound I reduction rates by iodide (>10{sup 8} M{sup -1} s{sup -1}) and thiocyanate (>10{sup 8} M{sup -1} s{sup -1}). By contrast, the overall chlorination and bromination activities were decreased by 98.1% and 87.4%, respectively, compared with the wild-type protein. Compound I reduction by chloride was slower than compound I decay to a compound II-like species (0.4 s{sup -1}), whereas compound I reduction by bromide was about 10-times slower (1.3 x 10{sup 4} M{sup -1} s{sup -1}) than the wild-type rate. These findings are discussed with respect to the known crystal structure of MPO and its bromide complex as well as the known redox chemistry of its intermediates and substrates.

  20. Defects in the Expression of Chloroplast Proteins Leads to H2O2 Accumulation and Activation of Cyclic Electron Flow around Photosystem I.

    PubMed

    Strand, Deserah D; Livingston, Aaron K; Satoh-Cruz, Mio; Koepke, Tyson; Enlow, Heather M; Fisher, Nicholas; Froehlich, John E; Cruz, Jeffrey A; Minhas, Deepika; Hixson, Kim K; Kohzuma, Kaori; Lipton, Mary; Dhingra, Amit; Kramer, David M

    2016-01-01

    We describe a new member of the class of mutants in Arabidopsis exhibiting high rates of cyclic electron flow around photosystem I (CEF), a light-driven process that produces ATP but not NADPH. High cyclic electron flow 2 (hcef2) shows strongly increased CEF activity through the NADPH dehydrogenase complex (NDH), accompanied by increases in thylakoid proton motive force (pmf), activation of the photoprotective qE response, and the accumulation of H2O2. Surprisingly, hcef2 was mapped to a non-sense mutation in the TADA1 (tRNA adenosine deaminase arginine) locus, coding for a plastid targeted tRNA editing enzyme required for efficient codon recognition. Comparison of protein content from representative thylakoid complexes, the cytochrome bf complex, and the ATP synthase, suggests that inefficient translation of hcef2 leads to compromised complex assembly or stability leading to alterations in stoichiometries of major thylakoid complexes as well as their constituent subunits. Altered subunit stoichiometries for photosystem I, ratios and properties of cytochrome bf hemes, and the decay kinetics of the flash-induced thylakoid electric field suggest that these defect lead to accumulation of H2O2 in hcef2, which we have previously shown leads to activation of NDH-related CEF. We observed similar increases in CEF, as well as increases in H2O2 accumulation, in other translation defective mutants. This suggests that loss of coordination in plastid protein levels lead to imbalances in photosynthetic energy balance that leads to an increase in CEF. These results taken together with a large body of previous observations, support a general model in which processes that lead to imbalances in chloroplast energetics result in the production of H2O2, which in turn activates CEF. This activation could be from either H2O2 acting as a redox signal, or by a secondary effect from H2O2 inducing a deficit in ATP.

  1. Defects in the Expression of Chloroplast Proteins Leads to H2O2 Accumulation and Activation of Cyclic Electron Flow around Photosystem I

    PubMed Central

    Strand, Deserah D.; Livingston, Aaron K.; Satoh-Cruz, Mio; Koepke, Tyson; Enlow, Heather M.; Fisher, Nicholas; Froehlich, John E.; Cruz, Jeffrey A.; Minhas, Deepika; Hixson, Kim K.; Kohzuma, Kaori; Lipton, Mary; Dhingra, Amit; Kramer, David M.

    2017-01-01

    We describe a new member of the class of mutants in Arabidopsis exhibiting high rates of cyclic electron flow around photosystem I (CEF), a light-driven process that produces ATP but not NADPH. High cyclic electron flow 2 (hcef2) shows strongly increased CEF activity through the NADPH dehydrogenase complex (NDH), accompanied by increases in thylakoid proton motive force (pmf), activation of the photoprotective qE response, and the accumulation of H2O2. Surprisingly, hcef2 was mapped to a non-sense mutation in the TADA1 (tRNA adenosine deaminase arginine) locus, coding for a plastid targeted tRNA editing enzyme required for efficient codon recognition. Comparison of protein content from representative thylakoid complexes, the cytochrome bf complex, and the ATP synthase, suggests that inefficient translation of hcef2 leads to compromised complex assembly or stability leading to alterations in stoichiometries of major thylakoid complexes as well as their constituent subunits. Altered subunit stoichiometries for photosystem I, ratios and properties of cytochrome bf hemes, and the decay kinetics of the flash-induced thylakoid electric field suggest that these defect lead to accumulation of H2O2 in hcef2, which we have previously shown leads to activation of NDH-related CEF. We observed similar increases in CEF, as well as increases in H2O2 accumulation, in other translation defective mutants. This suggests that loss of coordination in plastid protein levels lead to imbalances in photosynthetic energy balance that leads to an increase in CEF. These results taken together with a large body of previous observations, support a general model in which processes that lead to imbalances in chloroplast energetics result in the production of H2O2, which in turn activates CEF. This activation could be from either H2O2 acting as a redox signal, or by a secondary effect from H2O2 inducing a deficit in ATP. PMID:28133462

  2. Increasing sediment accumulation rates in La Fonera (Palamós) submarine canyon axis and their relationship with bottom trawling activities

    NASA Astrophysics Data System (ADS)

    Puig, P.; Martín, J.; Masqué, P.; Palanques, A.

    2015-10-01

    Previous studies conducted in La Fonera (Palamós) submarine canyon (NW Mediterranean) found that trawling activities along the canyon flanks cause resuspension and transport of sediments toward the canyon axis. 210Pb chronology supported by 137Cs dating applied to a sediment core collected at 1750 m in 2002 suggested a doubling of the sediment accumulation rate since the 1970s, coincident with the rapid industrialization of the local trawling fleet. The same canyon area has been revisited a decade later, and new data are consistent with a sedimentary regime shift during the 1970s and also suggest that the accumulation rate during the last decade could be greater than expected, approaching ~2.4 cm yr-1 (compared to ~0.25 cm yr-1 pre-1970s). These results support the hypothesis that commercial bottom trawling can substantially affect sediment dynamics and budgets on continental margins, eventually initiating the formation of anthropogenic depocenters in submarine canyon environments.

  3. Cellular accumulation and pharmacodynamic evaluation of the intracellular activity of CEM-101, a novel fluoroketolide, against Staphylococcus aureus, Listeria monocytogenes, and Legionella pneumophila in human THP-1 macrophages.

    PubMed

    Lemaire, Sandrine; Van Bambeke, Françoise; Tulkens, Paul M

    2009-09-01

    CEM-101 is a novel fluoroketolide with lower MICs than those of telithromycin and macrolides. Our aim was to assess the cellular accumulation and intracellular activity of CEM-101 using models developed for analyzing the pharmacokinetics and pharmacological properties of antibiotics against phagocytized bacteria. We used THP-1 macrophages and Staphylococcus aureus (ATCC 25923 [methicillin (meticillin) sensitive]), Listeria monocytogenes (strain EGD), and Legionella pneumophila (ATCC 33153). CEM-101 reached cellular-to-extracellular-concentration ratios of about 350 within 24 h (versus approximately 20, 30, and 160 for telithromycin, clarithromycin, and azithromycin, respectively). This intracellular accumulation was suppressed by incubation at a pH of < or = 6 and by monensin (proton ionophore) and was unaffected by verapamil (P-glycoprotein inhibitor; twofold accumulation increase for azithromycin) or gemfibrozil. While keeping with the general properties of the macrolide antibiotics in terms of maximal efficacy (Emax; approximately 1-log10-CFU decrease compared to the postphagocytosis inoculum after a 24-h incubation), CEM-101 showed significantly greater potency against phagocytized S. aureus than telithromycin, clarithromycin, and azithromycin (for which the 50% effective concentration [EC50] and static concentrations were about 3-, 6-, and 15-fold lower, respectively). CEM-101 was also about 50-fold and 100-fold more potent than azithromycin against phagocytized L. monocytogenes and L. pneumophila, respectively. These differences in EC50s and static concentrations between drugs were minimized when data were expressed as multiples of the MIC, demonstrating the critical role of intrinsic drug activity (MIC) in eliciting the antibacterial intracellular effects, whereas accumulation per se was unimportant. CEM-101 should show enhanced in vivo potency if used at doses similar to those of the comparators tested here.

  4. Wnt/β-Catenin Signaling Activation beyond Robust Nuclear β-Catenin Accumulation in Nondysplastic Barrett’s Esophagus: Regulation via Dickkopf-112

    PubMed Central

    Lyros, Orestis; Rafiee, Parvaneh; Nie, Linghui; Medda, Rituparna; Jovanovic, Nebojsa; Otterson, Mary F.; Behmaram, Behnaz; Gockel, Ιnes; Mackinnon, Alexander; Shaker, Reza

    2015-01-01

    INTRODUCTION: Wnt/β-catenin signaling activation has been reported only during the late steps of Barrett’s esophagus (BE) neoplastic progression, but not in BE metaplasia, based on the absence of nuclear β-catenin. However, β-catenin transcriptional activity has been recorded in absence of robust nuclear accumulation. Thus, we aimed to investigate the Wnt/β-catenin signaling in nondysplastic BE. METHODS: Esophageal tissues from healthy and BE patients without dysplasia were analyzed for Wnt target gene expression by quantitative reverse transcription polymerase chain reaction (qRT-PCR) and immunohistochemistry. Esophageal squamous (EPC1-& EPC2-hTERT), BE metaplastic (CP-A), and adenocarcinoma (OE33) cell lines were characterized for Wnt activation by qRT-PCR, Western blot, and luciferase assay. Wnt activity regulation was examined by using recombinant Wnt3a and Dickkopf-1 (Dkk1) as well as Dkk1 short interfering RNA. RESULTS: Wnt target genes (AXIN2, c-MYC, Cyclin D1, Dkk1) and Wnt3a were significantly upregulated in nondysplastic BE compared with squamous mucosa. Elevated levels of dephosphorylated β-catenin were detected in nondysplastic BE. Nuclear active β-catenin and TOPflash activity were increased in CP-A and OE33 cells compared with squamous cells. Wnt3a-mediated β-catenin signaling activation was abolished by Dkk1 in CP-A cells. TOPFlash activity was elevated following Dkk1 silencing in CP-A but not in OE33 cells. Dysplastic and esophageal adenocarcinoma tissues demonstrated further Dkk1 and AXIN2 overexpression. CONCLUSIONS: Despite the absence of robust nuclear accumulation, β-catenin is transcriptionally active in nondysplastic BE. Dkk1 overexpression regulates β-catenin signaling in BE metaplastic but not in adenocarcinoma cells, suggesting that early perturbation of Dkk1-mediated signaling suppression may contribute to BE malignant transformation. PMID:26297437

  5. Dietary freshwater clam (Corbicula fluminea) extract suppresses accumulation of hepatic lipids and increases in serum cholesterol and aminotransferase activities induced by dietary chloretone in rats.

    PubMed

    Chijimatsu, Takeshi; Umeki, Miki; Kobayashi, Satoru; Kataoka, Yutaro; Yamada, Koji; Oda, Hiroaki; Mochizuki, Satoshi

    2015-01-01

    We investigated the ameliorative effect of freshwater clam extract (FCE) on fatty liver, hypercholesterolemia, and liver injury in rats exposed to chloretone. Furthermore, we examined the effects of major FCE components (fat and protein fractions) to determine the active components in FCE. Chloretone increased serum aminotransferase activities and led to hepatic lipid accumulation. Serum aminotransferase activities and hepatic lipid content were lower in rats fed total FCE or fat/protein fractions of FCE. Expression of fatty acid synthase and fatty acid desaturase genes was upregulated by chloretone. Total FCE and fat/protein fractions of FCE suppressed the increase in gene expression involved in fatty acid synthesis. Serum cholesterol levels increased twofold upon chloretone exposure. Total FCE or fat/protein fractions of FCE showed hypocholesterolemic effects in rats with hypercholesterolemia induced by chloretone. These suggest that FCE contains at least two active components against fatty liver, hypercholesterolemia, and liver injury in rats exposed to chloretone.

  6. Ubiquitin C-terminal hydrolase-l1 activity induces polyubiquitin accumulation in podocytes and increases proteinuria in rat membranous nephropathy.

    PubMed

    Meyer-Schwesinger, Catherine; Meyer, Tobias N; Sievert, Henning; Hoxha, Elion; Sachs, Marlies; Klupp, Eva-Maria; Münster, Silvia; Balabanov, Stefan; Carrier, Lucie; Helmchen, Udo; Thaiss, Friedrich; Stahl, Rolf A K

    2011-05-01

    Ubiquitin C-terminal hydrolase L1 (UCH-L1), a key protease of the ubiquitin-proteasome system (UPS), is associated with neurodegenerative diseases and cancer. Recently, de novo expression of UCH-L1 was described in podocytes in patients with membranous nephropathy (MN), in which UCH-L1 expression correlated with increased ubiquitin content. The objective of the present study was to investigate the role of UCH-L1 in ubiquitin homeostasis and proteasomal degradation in a rat model of MN. After disease induction, UCH-L1 expression increased in podocytes and coincided with decreased glomerular monoubiquitin content. After an initial increase in proteasomal activity, the UPS was impaired. In addition to an increase of ubiquitin in podocytes, aggregates were observed 1 year after disease induction, as in MN in human beings. Inhibition of UCH-L1 hydrolase function in MN reduced UPS impairment and ameliorated proteinuria. In contrast, inhibition of proteasomal activity enhanced UPS impairment, resulting in increased proteinuria. Stable UCH-L1 overexpression in cultured podocytes resulted in accumulation of monoubiquitin and polyubiquitin proteins. In contrast, stable knock-down of UCH-L1 reduced monoubiquitin and polyubiquitin proteins and significantly increased proteasomal activity, indicating that the observed effects in rat MN also occurred in cultured podocytes. These data demonstrate that UCH-L1 activity results in polyubiquitin accumulation, proteasome inhibition, and disease aggravation in experimental models of MN.

  7. A Chimeric Cfh Transgene Leads to Increased Retinal Oxidative Stress, Inflammation, and Accumulation of Activated Subretinal Microglia in Mice

    PubMed Central

    Aredo, Bogale; Li, Tao; Chen, Xiao; Zhang, Kaiyan; Wang, Cynthia Xin-Zhao; Gou, Darlene; Zhao, Biren; He, Yuguang; Ufret-Vincenty, Rafael L.

    2015-01-01

    Purpose. Variants of complement factor H (Cfh) affecting short consensus repeats (SCRs) 6 to 8 increase the risk of age-related macular degeneration. Our aim was to explore the effect of expressing a Cfh variant on the in vivo susceptibility of the retina and RPE to oxidative stress and inflammation, using chimeric Cfh transgenic mice (chCfhTg). Methods. The chCfhTg and age-matched C57BL/6J (B6) mice were subjected to oxidative stress by either normal aging, or by exposure to a combination of oral hydroquinone (0.8% HQ) and increased light. Eyes were collected for immunohistochemistry of RPE–choroid flat mounts and of retinal sections, ELISA, electron microscopy, and RPE/microglia gene expression analysis. Results. Aging mice to 2 years led to an increased accumulation of basal laminar deposits, subretinal microglia/macrophages (MG/MΦ) staining for CD16 and for malondialdehyde (MDA), and MDA-modified proteins in the retina in chCfhTg compared to B6 mice. The chCfhTg mice maintained on HQ diet and increased light showed greater deposition of basal laminar deposits, more accumulation of fundus spots suggestive of MG/MΦ, and increased deposition of C3d in the sub-RPE space, compared to controls. In addition, chCfhTg mice demonstrated upregulation of NLRP3, IP-10, CD68, and TREM-2 in the RNA isolates from RPE/MG/MΦ. Conclusions. Expression of a Cfh transgene introducing a variant in SCRs 6 to 8 was sufficient to lead to increased retinal/RPE susceptibility to oxidative stress, a proinflammatory MG/MΦ phenotype, and a proinflammatory RPE/MG/MΦ gene expression profile in a transgenic mouse model. Our data suggest that altered interactions of Cfh with MDA-modified proteins may be relevant in explaining the effects of the Cfh variant. PMID:26030099

  8. Intensity of macrolide anti-inflammatory activity in J774A.1 cells positively correlates with cellular accumulation and phospholipidosis.

    PubMed

    Munić, Vesna; Banjanac, Mihailo; Koštrun, Sanja; Nujić, Krunoslav; Bosnar, Martina; Marjanović, Nikola; Ralić, Jovica; Matijašić, Mario; Hlevnjak, Mario; Eraković Haber, Vesna

    2011-09-01

    Some macrolide antibiotics were reported to inhibit interleukin-6 (IL6) and prostaglandin-E2 (PGE(2)) production by bacterial lipopolysaccharide (LPS) stimulated J774A.1 cells. Macrolides are also known to accumulate in cells and some were proven inducers of phospholipidosis. In the present study, with a set of 18 mainly 14- and 15-membered macrolides, we have investigated whether these macrolide induced phenomena in J774A.1 cells are connected. In LPS-stimulated J774A.1 cells, the extent of inhibition of proinflammatory markers (IL6 and PGE(2)) by macrolides significantly correlated with their extent of accumulation in cells, as well as with the induction of phospholipidosis, and cytotoxic effects in prolonged culture (with correlation coefficients (R) ranging from 0.78 to 0.93). The effects observed were related to macrolide binding to phospholipids (CHI IAM), number of positively charged centres, and were inversely proportional to the number of hydrogen bond donors. Similar interdependence of effects was obtained with chloroquine and amiodarone, whereas for dexamethasone and indomethacin these effects were not linked. The observed macrolide induced phenomena in J774A.1 cells were reversible and elimination of the macrolides from the culture media prevented phospholipidosis and the development of cytotoxicity in long-term cultures. Based on comparison with known clinical data, we conclude that LPS-stimulated J774A.1 cells in presented experimental setup are not a representative cellular model for the evaluation of macrolide anti-inflammatory potential in clinical trials. Nevertheless, our study shows that, at least in in vitro models, binding to biological membranes may be the crucial factor of macrolide mechanism of action.

  9. Diversity of Arbuscular Mycorrhizal Fungi Associated with a Sb Accumulator Plant, Ramie (Boehmeria nivea), in an Active Sb Mining.

    PubMed

    Wei, Yuan; Chen, ZhiPeng; Wu, FengChang; Li, JiNing; ShangGuan, YuXian; Li, FaSheng; Zeng, Qing Ru; Hou, Hong

    2015-08-01

    Arbuscular mycorrhizal fungi (AMF) have great potential for assisting heavy metal hyperaccumulators in the remediation of contaminated soils. However, little information is available about the symbiosis of AMF associated with an antimony (Sb) accumulator plant under natural conditions. Therefore, the objective of this study was to investigate the colonization and molecular diversity of AMF associated with the Sb accumulator ramie (Boehmeria nivea) growing in Sb-contaminated soils. Four Sb mine spoils and one adjacent reference area were selected from Xikuangshan in southern China. PCR-DGGE was used to analyze the AMF community composition in ramie roots. Morphological identification was also used to analyze the species in the rhizosphere soil of ramie. Results obtained showed that mycorrhizal symbiosis was established successfully even in the most heavily polluted sites. From the unpolluted site Ref to the highest polluted site T4, the spore numbers and AMF diversity increased at first and then decreased. Colonization increased consistently with the increasing Sb concentrations in the soil. A total of 14 species were identified by morphological analysis. From the total number of species, 4 (29%) belonged to Glomus, 2 (14%) belonged to Acaulospora, 2 (14%) belonged to Funneliformis, 1 (7%) belonged to Claroideoglomus, 1 (7%) belonged to Gigaspora, 1 (7%) belonged to Paraglomus, 1 (7%) belonging to Rhizophagus, 1 (7%) belonging to Sclervocystis, and 1 (7%) belonged to Scutellospora. Some AMF sequences were present even in the most polluted site. Morphological identification and phylogenetic analysis both revealed that most species were affiliated withGlomus, suggesting that Glomus was the dominant genus in this AMF community. This study demonstrated that ramie associated with AMF may have great potential for remediation of Sb-contaminated soils.

  10. Myeloperoxidase oxidized LDL interferes with endothelial cell motility through miR-22 and heme oxygenase 1 induction: possible involvement in reendothelialization of vascular injuries.

    PubMed

    Daher, Jalil; Martin, Maud; Rousseau, Alexandre; Nuyens, Vincent; Fayyad-Kazan, Hussein; Van Antwerpen, Pierre; Courbebaisse, Guy; Martiat, Philippe; Badran, Bassam; Dequiedt, Frank; Zouaoui Boudjeltia, Karim; Vanhamme, Luc

    2014-01-01

    Cardiovascular disease linked to atherosclerosis is the leading cause of death worldwide. Atherosclerosis is mainly linked to dysfunction in vascular endothelial cells and subendothelial accumulation of oxidized forms of LDL. In the present study, we investigated the role of myeloperoxidase oxidized LDL (Mox-LDL) in endothelial cell dysfunction. We studied the effect of proinflammatory Mox-LDL treatment on endothelial cell motility, a parameter essential for normal vascular processes such as angiogenesis and blood vessel repair. This is particularly important in the context of an atheroma plaque, where vascular wall integrity is affected and interference with its repair could contribute to progression of the disease. We investigated in vitro the effect of Mox-LDL on endothelial cells angiogenic properties and we also studied the signalling pathways that could be affected by analysing Mox-LDL effect on the expression of angiogenesis-related genes. We report that Mox-LDL inhibits endothelial cell motility and tubulogenesis through an increase in miR-22 and heme oxygenase 1 expression. Our in vitro data indicate that Mox-LDL interferes with parameters associated with angiogenesis. They suggest that high LDL levels in patients would impair their endothelial cell capacity to cope with a damaged endothelium contributing negatively to the progression of the atheroma plaque.

  11. Size effect of elemental selenium nanoparticles (Nano-Se) at supranutritional levels on selenium accumulation and glutathione S-transferase activity.

    PubMed

    Peng, Dungeng; Zhang, Jinsong; Liu, Qingliang; Taylor, Ethan Will

    2007-10-01

    It has been shown that 36 nm Nano-Se has lower toxicity than selenite or selenomethionine, but these forms of selenium (Se) all possess similar ability to increase selenoenzyme levels. The size of nanoparticles plays an important role in their biological activity: as expected, 5-200 nm Nano-Se can directly scavenge free radicals in vitro in a size-dependent fashion. However, in Se-deficient cells and Se-deficient mice, the size effect of Nano-Se on increasing selenoenzymes and liver Se disappears unexpectedly. We hypothesize that under conditions of Se deficiency, the avidity of Se uptake mechanisms may be increased to maintain the biosynthesis of selenoenzymes, which are fundamental for redox homeostasis. This increased avidity may override the potential advantage of small size Nano-Se seen under Se-replete conditions, thereby eliminating the size effect. Once selenoenzymes have been saturated, Se uptake mechanisms may downregulate; accordingly, the size effect of Nano-Se can then reappear. To test this hypothesis, Se-deficient mice were administered either 36 or 90 nm Nano-Se at supranutritional doses, in both a short-term model and a single-dose model. Under these conditions, Nano-Se showed a size effect on Se accumulation and glutathione S-transferase (GST) activity. A size effect of Nano-Se was found in 15 out of 18 total comparisons between sizes at the same dose and time in the two models. Furthermore, the magnitude of the size effect was more prominent on Se accumulation than on GST activity. GST is strictly regulated by transcriptional and translational mechanisms, so its increase in activity normally does not exceed 3-fold. In contrast, the homeostasis of Se accumulation is not as tightly controlled. In the present experiments, GST activity had reached or was approaching saturation, but liver Se was far below saturation. Therefore, our results strongly suggest that the saturation profile of the tested biomarker has an impact on the size effect of Nano

  12. The response of Hordeum spontaneum desert ecotype to drought and excessive light intensity is characterized by induction of O2 dependent photochemical activity and anthocyanin accumulation.

    PubMed

    Eppel, Amir; Keren, Nir; Salomon, Eitan; Volis, Sergei; Rachmilevitch, Shimon

    2013-03-01

    The goal of the current research was to study the role of anthocyanin accumulation, O(2)-related photochemical processes and non-photochemical quenching (NPQ) in the response of desert and Mediterranean plants to drought and excessive light. Plants of Hordeum spontaneum were collected from Mediterranean and desert environments and were subjected to terminal drought for 25 days and then measured for PSII yield at 2 and 21% O(2), NPQ, net carbon assimilation, stomatal conductance, leaf relative water content (LRWC), anthocyanin concentration and leaf absorbance. Under terminal drought, LRWC, carbon assimilation and stomatal conductance decreased similarly and significantly in both the Mediterranean and the desert ecotypes. Anthocyanin accumulated more in the desert ecotype than in the Mediterranean ecotype. NPQ increased more in the Mediterranean ecotype as compared with the desert ecotype. PSII yield decreased significantly in the Mediterranean ecotype under drought and was much lower than in the desert ecotype under drought. The relatively high PSII yield under drought in the desert ecotype was O(2) dependent. The response of the H. spontaneum ecotype from a desert environment to drought stress was characterized by anthocyanin accumulation and induction of O(2) dependent photochemical activity, while the response of the Mediterranean ecotype was based on a higher induction of NPQ.

  13. Artesunate has its enhancement on antibacterial activity of β-lactams via increasing the antibiotic accumulation within methicillin-resistant Staphylococcus aureus (MRSA).

    PubMed

    Jiang, Weiwei; Li, Bin; Zheng, Xinchuan; Liu, Xin; Pan, Xichun; Qing, Rongxin; Cen, Yanyan; Zheng, Jiang; Zhou, Hong

    2013-06-01

    Methicillin-resistant Staphylococcus aureus (MRSA) has now emerged as a predominant and serious pathogen because of its resistance to a large group of antibiotics, leading to high morbidity and mortality. Therefore, to develop new agents against resistance is urgently required. Previously, artesunate (AS) was found to enhance the antibacterial effect of β-lactams against MRSA. In this study, AS was first found to increase the accumulation of antibiotics (daunorubicin and oxacillin) within MRSA by laser confocal microscopy and liquid chromatography-tandem MS method, suggesting the increased antibiotics accumulation might be related to the enhancement of AS on antibiotics. Furthermore, AS was found not to destroy the cell structure of MRSA by transmission electron microscope. AS was found to inhibit gene expressions of important efflux pumps such as NorA, NorB and NorC, but not MepA, SepA and MdeA. In conclusion, our results showed that AS was capable of enhancing the antibacterial activity of β-lactams via increasing antibiotic accumulations within MRSA through inhibiting gene expressions of efflux pumps such as NorA, NorB and NorC, but did not destroy the cell structure of MRSA. AS could be further investigated as a candidate drug for treatment of MRSA infection.

  14. Functional Analysis of Arabidopsis Mutants Points to Novel Roles for Glutathione in Coupling H2O2 to Activation of Salicylic Acid Accumulation and Signaling

    PubMed Central

    Han, Yi; Chaouch, Sejir; Mhamdi, Amna; Queval, Guillaume; Zechmann, Bernd

    2013-01-01

    Abstract Aims: Through its interaction with H2O2, glutathione is a candidate for transmission of signals in plant responses to pathogens, but identification of signaling roles is complicated by its antioxidant function. Using a genetic approach based on a conditional catalase-deficient Arabidopsis mutant, cat2, this study aimed at establishing whether GSH plays an important functional role in the transmission of signals downstream of H2O2. Results: Introducing the cad2 or allelic mutations in the glutathione synthesis pathway into cat2 blocked H2O2-triggered GSH oxidation and accumulation. While no effects on NADP(H) or ascorbate were observed, and H2O2-induced decreases in growth were maintained, blocking GSH modulation antagonized salicylic acid (SA) accumulation and SA-dependent responses. Other novel double and triple mutants were produced and compared with cat2 cad2 at the levels of phenotype, expression of marker genes, nontargeted metabolite profiling, accumulation of SA, and bacterial resistance. Most of the effects of the cad2 mutation on H2O2-triggered responses were distinct from those produced by mutations for GLUTATHIONE REDUCTASE1 (GR1) or NONEXPRESSOR OF PATHOGENESIS-RELATED GENES 1 (NPR1), and were linked to compromised induction of ISOCHORISMATE SYNTHASE1 (ICS1) and ICS1-dependent SA accumulation. Innovation: A novel genetic approach was used in which GSH content or antioxidative capacity was independently modified in an H2O2 signaling background. Analysis of new double and triple mutants allowed us to infer previously undescribed regulatory roles for GSH. Conclusion: In parallel to its antioxidant role, GSH acts independently of NPR1 to allow increased intracellular H2O2 to activate SA signaling, a key defense response in plants. Antioxid. Redox Signal. 18, 2106–2121. PMID:23148658

  15. Inhibition of the host proteasome facilitates papaya ringspot virus accumulation and proteosomal catalytic activity is modulated by viral factor HcPro.

    PubMed

    Sahana, Nandita; Kaur, Harpreet; Basavaraj; Tena, Fatima; Jain, Rakesh Kumar; Palukaitis, Peter; Canto, Tomas; Praveen, Shelly

    2012-01-01

    The ubiquitin/26S proteasome system plays an essential role not only in maintaining protein turnover, but also in regulating many other plant responses, including plant-pathogen interactions. Previous studies highlighted different roles of the 20S proteasome in plant defense during virus infection, either indirectly through viral suppressor-mediated degradation of Argonaute proteins, affecting the RNA interference pathway, or directly through modulation of the proteolytic and RNase activity of the 20S proteasome, a component of the 20S proteasome, by viral proteins, affecting the levels of viral proteins and RNAs. Here we show that MG132, a cell permeable proteasomal inhibitor, caused an increase in papaya ringspot virus (PRSV) accumulation in its natural host papaya (Carica papaya). We also show that the PRSV HcPro interacts with the papaya homologue of the Arabidopsis PAA (α1 subunit of the 20S proteasome), but not with the papaya homologue of Arabidopsis PAE (α5 subunit of the 20S proteasome), associated with the RNase activity, although the two 20S proteasome subunits interacted with each other. Mutated forms of PRSV HcPro showed that the conserved KITC54 motif in the N-terminal domain of HcPro was necessary for its binding to PAA. Co-agroinfiltration assays demonstrated that HcPro expression mimicked the action of MG132, and facilitated the accumulation of bothtotal ubiquitinated proteins and viral/non-viral exogenous RNA in Nicotiana benthamiana leaves. These effects were not observed by using an HcPro mutant (KITS54), which impaired the HcPro - PAA interaction. Thus, the PRSV HcPro interacts with a proteasomal subunit, inhibiting the action of the 20S proteasome, suggesting that HcPro might be crucial for modulating its catalytic activities in support of virus accumulation.

  16. Subcellular cadmium distribution and antioxidant enzymatic activities in the leaves of two castor (Ricinus communis L.) cultivars exhibit differences in Cd accumulation.

    PubMed

    Zhang, Hanzhi; Guo, Qingjun; Yang, Junxing; Shen, Jianxiu; Chen, Tongbin; Zhu, Guangxu; Chen, Hui; Shao, Chunyan

    2015-10-01

    The aims of this study were: (1) the study of cadmium (Cd) accumulation and toxicity in different castor cultivars (Ricinus communis L.); (2) to investigate changes in antioxidant enzymatic activities and the subcellular distribution of Cd in young and old leaves from two different castor cultivars, after exposure to two different Cd concentrations, and explore the underlying mechanism of Cd detoxification focusing on antioxidant enzymes and subcellular compartmentalization. The Cd concentration, toxicity, and subcellular distribution, as well as superoxide dismutase (SOD), catalase (CAT), and peroxidase (POD) activities were measured in Zibo-3 and Zibo-9 cultivars after exposure to two different concentrations of Cd (2mg/L and 5mg/L) for 10 days. This research revealed Cd accumulation characteristics in castor are root>stem>young leaf>old leaf. Castor tolerance was Cd dose exposure and the cultivars themselves dependent. Investigation of subcellular Cd partitioning showed that Cd accumulated mainly in the heat stable protein (HSP) and cellular debris fractions, followed by the Cd rich granule (MRG), heat denatured protein (HDP), and organelle fractions. With increasing Cd concentration in nutrient solution, the decreased detoxified fractions (BDM) and the increased Cd-sensitive fractions (MSF) in young leaves may indicate the increased Cd toxicity in castor cultivars. The BDM-Cd fractions or MSF-Cd in old leaves may be linked with Cd tolerance of different cultivars of castor. The antioxidant enzymes that govern Cd detoxification were not found to be active in leaves. Taken together, these results indicate Cd tolerance and toxicity in castor can be explained by subcellular partitioning.

  17. Activation of the complement system and accumulation of hemoglobin-haptoglobin complexes in plasma during an adverse reaction to penicillin treatment.

    PubMed

    Brandslund, I; Svehag, S E; Teisner, B; Hyltoft Petersen, P

    1983-01-01

    A patient treated with penicillin intravenously developed a serum sickness-like reaction. Classical pathway complement (C) activation was indicated by quantitation of the split products C3c and C3d as well as demonstration of C4 conversion. Circulating immune complexes could, however, not be detected by the solid-phase Clq and PEG-precipitation methods. A concomitant accumulation of circulating hemoglobin-haptoglobin complexes and a marked fall in serum-fibronectin concentrations suggested saturation of the reticuloendothelial system. The plasma became a deep-red color, and a diffuse intravascular coagulation followed. The patient recovered completely upon discontinuation of penicillin administration.

  18. Accumulation of Epstein-Barr virus (EBV) BMRF1 protein EA-D during latent EBV activation of Burkitt's lymphoma cell line Raji.

    PubMed

    Ohashi, Makoto; Horie, Kazutaka; Hoshikawa, Yoshiko; Nagata, Keiko; Osaki, Mistuhiko; Ito, Hisao; Sairenji, Takeshi

    2007-02-01

    As a new model to elucidate molecular mechanisms in Epstein-Barr virus (EBV) activation, we tested the tetracycline-inducible (Tet-On)/BZLF1-oriP plasmid system in Raji cells. Cells transfected with this Tet-On plasmid did not activate EBV by doxycycline and surprisingly EBV latency was disrupted with large amounts of BMRF1 protein (EA-D) being accumulated in the cells. Brilliant EA-D fluorescence was markedly condensed in small sized cells, intra-cellular vesicles, and extra-cellular particles. Scanning electron microscopy demonstrated the extra-cellular particles to be covered with a membrane. EA-D molecules of 58, 50, 48, and 44kDa were expressed in the cells. The high (58 and 50kDa) and low (48 and 44kDa) EA-D molecules appeared in the early and late stages, respectively. Low EA-D molecules were detected mostly in EA-D positive cells separated into the heaviest density layer of a discontinuous Percoll gradient. Such molecules could be created from high EA-D molecules by protein phosphatase treatment. The EA-D molecules that appeared similar were detected in EBV-activated P3HR-1 and Akata cells. Several hypotheses concerning the accumulation of EA-D molecules of various polymorphic forms and their phosphorylation/dephosphorylation in this model system are presented, with possible biological and clinical relevance.

  19. HLA-G promotes myeloid-derived suppressor cell accumulation and suppressive activity during human pregnancy through engagement of the receptor ILT4.

    PubMed

    Köstlin, Natascha; Ostermeir, Anna-Lena; Spring, Bärbel; Schwarz, Julian; Marmé, Alexander; Walter, Christina B; Poets, Christian F; Gille, Christian

    2017-02-01

    Establishing and maintaining maternal-fetal tolerance is essential for a successful pregnancy; failure of immunological adaptation to pregnancy leads to severe complications such as abortion or preterm delivery. Myeloid-derived suppressor cells (MDSCs) are innate immune cells that suppress T-cell responses, expand during pregnancy and thus may play a role in tolerance induction. Human leucocyte antigen G (HLA-G) is a major histocompatibility complex (MHC) I molecule with immune-modulatory properties, which is expressed during pregnancy. Here, we investigated the impact of HLA-G on MDSCs accumulation and activation in pregnant women. We demonstrate that granulocytic MDSCs (GR-MDSCs) express receptors for HLA-G, namely immunoglobulin-like transcript (ILT) 2 and 4, and that ILT4-expression by GR-MDSCs is regulated during pregnancy. Stimulation with soluble HLA-G (sHLA-G) increased suppressive activity of GR-MDSCs, induced MDSCs from peripheral blood mononuclear cells (PBMCs) and led to phosphorylation of the signal transducer and activator of transcription 3 (STAT3) and induction of indoleamine-2,3-dioxygenase (IDO) in myeloid cells. Effects of sHLA-G on MDSC accumulation were mediated through ILT4. These results suggest an interaction between MDSCs and HLA-G in humans as a potential mechanism for maintaining maternal-fetal tolerance. Modulating MDSC function during pregnancy via HLA-G might provide new opportunities for a therapeutic manipulation of immunological pregnancy complications.

  20. Effect of water withdrawal on formation of free radical, proline accumulation and activities of antioxidant enzymes in ZAT12-transformed transgenic tomato plants.

    PubMed

    Chandra Rai, Avinash; Singh, Major; Shah, Kavita

    2012-12-01

    Water stress often leads to the accumulation of reactive oxygen species (ROS) and their excessive production alters the activities of enzymes involved in their removal. ZAT12 is a member of stress-responsive C(2)H(2) type Zinc Finger Protein (ZFP) reported to control the expression of several stress-activated genes in plants through ROS signaling. The ZAT12-transformed tomato lines (cv. H-86 variety Kashi Vishesh) when subjected to water withdrawal for 7, 14 and 21 days revealed significant and consistent changes in activities of enzymes SOD, CAT, APX, GR and POD paralleled with an increased proline levels. Unlike that in wild-type tomato, the leaf superoxide anion and hydrogen peroxide concentrations in the transformed tomato plants did not alter much, suggesting a well regulated formation of free radicals suppressing oxidative stress in the latter. Results suggest BcZAT12-transformed tomato lines ZT1, ZT2 and ZT6 to be better adapted to drought stress tolerance by accumulation of osmolyte proline and increased antioxidant response triggered by the ZAT12 gene. Therefore, the ZAT12-transformed tomato cv. H-86 lines will prove useful for higher yield of tomato crop in regions affected with severe drought stress.

  1. Active accumulation of very diluted biomolecules by nano-dispensing for easy detection below the femtomolar range

    NASA Astrophysics Data System (ADS)

    Grilli, S.; Miccio, L.; Gennari, O.; Coppola, S.; Vespini, V.; Battista, L.; Orlando, P.; Ferraro, P.

    2014-11-01

    Highly sensitive detection of biomolecules is of paramount interest in many fields including biomedicine, safety and eco-pollution. Conventional analyses use well-established techniques with detection limits ~1 pM. Here we propose a pyro-concentrator able to accumulate biomolecules directly onto a conventional binding surface. The operation principle is relatively simple but very effective. Tiny droplets are drawn pyro-electro-dynamically and released onto a specific site, thus increasing the sensitivity. The reliability of the technique is demonstrated in case of labelled oligonucleotides diluted serially. The results show the possibility to detect very diluted oligonucleotides, down to a few hundreds of attomoles. Excellent results are shown also in case of a sample of clinical interest, the gliadin, where a 60-fold improved detection limit is reached, compared with standard ELISA. This method could open the way to a mass-based technology for sensing molecules at very low concentrations, in environmental as well as in diagnostics applications.

  2. Chlorination of N-acetyltyrosine with HOCl, chloramines, and myeloperoxidase-hydrogen peroxide-chloride system.

    PubMed

    Drabik, G; Naskalski, J W

    2001-01-01

    N-acetyl-L-tyrosine (N-acTyr), with the alpha amine residue blocked by acetylation, can mimic the reactivity of exposed tyrosyl residues incorporated into polypeptides. In this study chlorination of N-acTyr residue at positions 3 and 5 in reactions with NaOCl, chloramines and the myeloperoxidase (MPO)-H2O2-Cl- chlorinating system were invesigated. The reaction of N-acTyr with HOCl/OCl- depends on the reactant concentration ratio employed. At the OCl-/N-acTyr (molar) ratio 1:4 and pH 5.0 the chlorination reaction yield is about 96% and 3-chlorotyrosine is the predominant reaction product. At the OCl-/N-acTyr molar ratio 1:1.1 both 3-chlorotyrosine and 3,5-dichlorotyrosine are formed. The yield of tyrosine chlorination depends also on pH, amounting to 100% at pH 5.5, 91% at pH 4.5 and 66% at pH 3.0. Replacing HOCl/OCl- by leucine/chloramine or alanine/chloramine in the reaction system, at pH 4.5 and 7.4, produces trace amount of 3-chlorotyrosine with the reaction yield of about 2% only. Employing the MPO-H2O2-Cl- chlorinating system at pH 5.4, production of a small amount of N-acTyr 3-chloroderivative was observed, but the reaction yield was low due to the rapid inactivation of MPO in the reaction system. The study results indicate that direct chlorination of tyrosyl residues which are not incorporated into the polypeptide structure occurs with excess HOCl/OCl- in acidic media. Due to the inability of the myeloperoxidase-H2O2-Cl- system to produce high enough HOCl concentrations, the MPO-mediated tyrosyl residue chlorination is not effective. Semistable amino-acid chloramines also appeared not effective as chlorine donors in direct tyrosyl chlorination.

  3. Functional characterization of NAC55 transcription factor from oilseed rape (Brassica napus L.) as a novel transcriptional activator modulating reactive oxygen species accumulation and cell death.

    PubMed

    Niu, Fangfang; Wang, Chen; Yan, Jingli; Guo, Xiaohua; Wu, Feifei; Yang, Bo; Deyholos, Michael K; Jiang, Yuan-Qing

    2016-09-01

    NAC transcription factors (TFs) are plant-specific and play important roles in development, responses to biotic and abiotic cues and hormone signaling. So far, only a few NAC genes have been reported to regulate cell death. In this study, we identified and characterized a NAC55 gene isolated from oilseed rape (Brassica napus L.). BnaNAC55 responds to multiple stresses, including cold, heat, abscisic acid (ABA), jasmonic acid (JA) and a necrotrophic fungal pathogen Sclerotinia sclerotiorum. BnaNAC55 has transactivation activity and is located in the nucleus. BnaNAC55 is able to form homodimers in planta. Unlike ANAC055, full-length BnaNAC55, but not either the N-terminal NAC domain or C-terminal regulatory domain, induces ROS accumulation and hypersensitive response (HR)-like cell death when expressed both in oilseed rape protoplasts and Nicotiana benthamiana. Furthermore, BnaNAC55 expression causes obvious nuclear DNA fragmentation. Moreover, quantitative reverse transcription PCR (qRT-PCR) analysis identified that the expression levels of multiple genes regulating ROS production and scavenging, defense response as well as senescence are significantly induced. Using a dual luciferase reporter assay, we further confirm that BnaNAC55 could activate the expression of a few ROS and defense-related gene expression. Taken together, our work has identified a novel NAC TF from oilseed rape that modulates ROS accumulation and cell death.

  4. Rifampicin-Induced Hepatic Lipid Accumulation: Association with Up-Regulation of Peroxisome Proliferator-Activated Receptor γ in Mouse Liver

    PubMed Central

    Zhang, Da-Gang; Li, Lu; Chen, Xi; Xu, De-Xiang

    2016-01-01

    Previous study found that rifampicin caused intrahepatic cholestasis. This study investigated the effects of rifampicin on hepatic lipid metabolism. Mice were orally administered with rifampicin (200 mg/kg) daily for different periods. Results showed that serum TG level was progressively reduced after a short elevation. By contrast, hepatic TG content was markedly increased in rifampicin-treated mice. An obvious hepatic lipid accumulation, as determined by Oil Red O staining, was observed in mice treated with rifampicin for more than one week. Moreover, mRNA levels of Fas, Acc and Scd-1, several key genes for fatty acid synthesis, were elevated in rifampicin-treated mice. In addition, the class B scavenger receptor CD36 was progressively up-regulated by rifampicin. Interestingly, hepatic SREBP-1c and LXR-α, two important transcription factors that regulate genes for hepatic fatty acid synthesis, were not activated by rifampicin. Instead, hepatic PXR was rapidly activated in rifampicin-treated mice. Hepatic PPARγ, a downstream target of PXR, was transcriptionally up-regulated. Taken together, the increased hepatic lipid synthesis and uptake of fatty acids from circulation into liver jointly contribute to rifampicin-induced hepatic lipid accumulation. The increased uptake of fatty acids from circulation into liver might be partially attributed to rifampicin-induced up-regulation of PPARγ and its target genes. PMID:27806127

  5. Data on amyloid precursor protein accumulation, spontaneous physical activity, and motor learning after traumatic brain injury in the triple-transgenic mouse model of Alzheimer׳s disease.

    PubMed

    Kishimoto, Yasushi; Shishido, Hajime; Sawanishi, Mayumi; Toyota, Yasunori; Ueno, Masaki; Kubota, Takashi; Kirino, Yutaka; Tamiya, Takashi; Kawai, Nobuyuki

    2016-12-01

    This data article contains supporting information regarding the research article entitled "Traumatic brain injury accelerates amyloid-β deposition and impairs spatial learning in the triple-transgenic mouse model of Alzheimer׳s disease" (H. Shishido, Y. Kishimoto, N. Kawai, Y. Toyota, M. Ueno, T. Kubota, Y. Kirino, T. Tamiya, 2016) [1]. Triple-transgenic (3×Tg)-Alzheimer׳s disease (AD) model mice exhibited significantly poorer spatial learning than sham-treated 3×Tg-AD mice 28 days after traumatic brain injury (TBI). Correspondingly, amyloid-β (Aβ) deposition within the hippocampus was significantly greater in 3×Tg-AD mice 28 days after TBI. However, data regarding the short-term and long-term influences of TBI on amyloid precursor protein (APP) accumulation in AD model mice remain limited. Furthermore, there is little data showing whether physical activity and motor learning are affected by TBI in AD model mice. Here, we provide immunocytochemistry data confirming that TBI induces significant increases in APP accumulation in 3×Tg-AD mice at both 7 days and 28 days after TBI. Furthermore, 3×Tg-AD model mice exhibit a reduced ability to acquire conditioned responses (CRs) during delay eyeblink conditioning compared to sham-treated 3×Tg-AD model mice 28 days after TBI. However, physical activity and motor performance are not significantly changed in TBI-treated 3×Tg-AD model mice.

  6. Intraneuronal Aβ accumulation induces hippocampal neuron hyperexcitability through A-type K(+) current inhibition mediated by activation of caspases and GSK-3.

    PubMed

    Scala, Federico; Fusco, Salvatore; Ripoli, Cristian; Piacentini, Roberto; Li Puma, Domenica Donatella; Spinelli, Matteo; Laezza, Fernanda; Grassi, Claudio; D'Ascenzo, Marcello

    2015-02-01

    Amyloid β-protein (Aβ) pathologies have been linked to dysfunction of excitability in neurons of the hippocampal circuit, but the molecular mechanisms underlying this process are still poorly understood. Here, we applied whole-cell patch-clamp electrophysiology to primary hippocampal neurons and show that intracellular Aβ42 delivery leads to increased spike discharge and action potential broadening through downregulation of A-type K(+) currents. Pharmacologic studies showed that caspases and glycogen synthase kinase 3 (GSK-3) activation are required for these Aβ42-induced effects. Extracellular perfusion and subsequent internalization of Aβ42 increase spike discharge and promote GSK-3-dependent phosphorylation of the Kv4.2 α-subunit, a molecular determinant of A-type K(+) currents, at Ser-616. In acute hippocampal slices derived from an adult triple-transgenic Alzheimer's mouse model, characterized by endogenous intracellular accumulation of Aβ42, CA1 pyramidal neurons exhibit hyperexcitability accompanied by increased phosphorylation of Kv4.2 at Ser-616. Collectively, these data suggest that intraneuronal Aβ42 accumulation leads to an intracellular cascade culminating into caspases activation and GSK-3-dependent phosphorylation of Kv4.2 channels. These findings provide new insights into the toxic mechanisms triggered by intracellular Aβ42 and offer potentially new therapeutic targets for Alzheimer's disease treatment.

  7. Intraneuronal Aβ accumulation induces hippocampal neuron hyperexcitability through A-type K+ current inhibition mediated by activation of caspases and GSK-3

    PubMed Central

    Scala, Federico; Fusco, Salvatore; Ripoli, Cristian; Piacentini, Roberto; Li Puma, Domenica Donatella; Spinelli, Matteo; Laezza, Fernanda; Grassi, Claudio; D’Ascenzo, Marcello

    2016-01-01

    Amyloid β-protein (Aβ) pathologies have been linked to dysfunction of excitability in neurons of the hippocampal circuit, but the molecular mechanisms underlying this process are still poorly understood. Here, we applied whole-cell patch-clamp electrophysiology to primary hippocampal neurons and show that intracellular Aβ42 delivery leads to increased spike discharge and action potential broadening through downregulation of A-type K+ currents. Pharmacologic studies showed that caspases and glycogen synthase kinase 3 (GSK-3) activation are required for these Aβ42-induced effects. Extracellular perfusion and subsequent internalization of Aβ42 increase spike discharge and promote GSK-3-dependent phosphorylation of the Kv4.2 α-subunit, a molecular determinant of A-type K+ currents, at Ser-616. In acute hippocampal slices derived from an adult triple-transgenic Alzheimer’s mouse model, characterized by endogenous intracellular accumulation of Aβ42, CA1 pyramidal neurons exhibit hyperexcitability accompanied by increased phosphorylation of Kv4.2 at Ser-616. Collectively, these data suggest that intraneuronal Aβ42 accumulation leads to an intracellular cascade culminating into caspases activation and GSK-3-dependent phosphorylation of Kv4.2 channels. These findings provide new insights into the toxic mechanisms triggered by intracellular Aβ42 and offer potentially new therapeutic targets for Alzheimer’s disease treatment. PMID:25541422

  8. Influence of arbuscular mycorrhizal fungi and copper on growth, accumulation of osmolyte, mineral nutrition and antioxidant enzyme activity of pepper (Capsicum annuum L.).

    PubMed

    Abdel Latef, Arafat Abdel Hamed

    2011-08-01

    The effect of arbuscular mycorrhizal (AM) fungi inoculation on pepper (Capsicum annuum L. cv. Zhongjiao 105) plant growth and on some physiological parameters in response to increasing soil Cu concentrations was studied. Treatments consisted of inoculation or not with Glomus mosseae and the addition of Cu to soil at the concentrations of 0 (control), 2 (low), 4 (medium), and 8 (high) mM CuSO(4). AM fungal inoculation decreased Cu concentrations in plant organs and promoted biomass yields as well as the contents of chlorophyll, soluble sugar, total protein, and the concentrations of P, K, Ca, and Mg. Plants grown in high Cu concentration exhibited a Cu-induced proline accumulation and also an increase in total free amino acid contents; however, both were lower in mycorrhizal pepper. Cu-induced oxidative stress by increasing lipid peroxidation rates and the activity of superoxide dismutase, catalase, ascorbate peroxidase and glutathione reductase, and AM symbiosis enhanced these antioxidant enzyme activities and decreased oxidative damage to lipids. In conclusion G. mosseae was able to maintain an efficient symbiosis with pepper plants in contaminated Cu soils, improving plant growth under these conditions, which is likely to be due to reduced Cu accumulation in plant tissues, reduced oxidative stress and damage to lipids, or enhanced antioxidant capacity.

  9. Accumulation and activation of epidermal γδ T cells in a mouse model of chronic dermatitis is not required for the inflammatory phenotype.

    PubMed

    Sulcova, Jitka; Maddaluno, Luigi; Meyer, Michael; Werner, Sabine

    2015-09-01

    Chronic skin inflammation resulting from a defective epidermal barrier is a hallmark of atopic dermatitis (AD). We previously demonstrated that mice lacking FGF receptors 1 and 2 in keratinocytes (K5-R1/R2 mice) develop an AD-like chronic dermatitis as a result of an impaired epidermal barrier. Here, we show that γδ T cells, which rapidly respond to various insults, accumulate in the epidermis of K5-R1/R2 mice before the development of histological abnormalities. Their number and activation further increase as the phenotype progresses, most likely as a consequence of increased expression of Il-2 and Il-7 and the stress-induced proteins Rae-1, H60c, Mult1, PlexinB2, and Skint1. To determine the role of γδ T cells in the skin phenotype, we generated quadruple mutant K5-R1/-R2 mice lacking γδ T cells. Surprisingly, loss of γδ T cells did not or only marginally affect keratinocyte proliferation, epidermal thickness, epidermal barrier function, and accumulation and activation of different immune cells in the skin of K5-R1/R2 mice, possibly due to partial compensation by αβ T cells. These results demonstrate that γδ T cells do not contribute to the development or maintenance of chronic inflammation in response to a defect in the epidermal barrier.

  10. The dipeptide H-Trp-Glu-OH (WE) shows agonistic activity to peroxisome proliferator-activated protein-α and reduces hepatic lipid accumulation in lipid-loaded H4IIE cells.

    PubMed

    Jia, Yaoyao; Kim, Jong-Ho; Nam, Bora; Kim, Jiyoung; Lee, Ji Hae; Hwang, Kwang-Yeon; Lee, Sung-Joon

    2014-07-01

    Dipeptides digested from dietary proteins can be directly absorbed by the intestine and delivered to the circulatory system. However, the dipeptides' metabolic roles and biological activities are largely unknown. Lipid-loaded HII4E cells stimulated with H-Trp-Glu-OH (WE) exhibited reduced lipid accumulation, of which the effect was abolished by peroxisome proliferator-activated receptor (PPAR) α gene knock down. A luciferase assay showed that the WE dipeptide induced PPARα transactivation in a dose-dependent manner. Surface plasmon resonance and time-resolved fluorescence resonance energy transfer analyses demonstrated that WE interacts directly with the PPARα ligand binding domain (KD, 120 μM; EC50, 83 μM). Cells stimulated with WE induced PPARα and its responsive genes and increased cellular fatty acid uptake. In conclusion, WE reduces hepatic lipid accumulation in lipid-loaded hepatocytes via the activation of PPARα by a direct interaction.

  11. Changes in the viscosity of hyaluronic acid after exposure to a myeloperoxidase-derived oxidant

    SciTech Connect

    Baker, M.S.; Green, S.P.; Lowther, D.A.

    1989-04-01

    Both purified hyaluronic acid (HA) and bovine synovial fluid react with OCI-, the major oxidant produced by the myeloperoxidase (MPO)/H/sub 2/O/sub 2//CI- system, resulting in a decrease in their specific viscosity. This reaction is inhibited in the presence of excess methionine. H/sub 2/O/sub 2/ alone decreases the viscosity of HA, presumably by the Fenton reaction, in the absence (but not in the presence) of the iron chelator, diethyltriaminepentacetic acid (DETAPAC). In the presence of DETAPAC, incubation of HA with the complete MPO/H/sub 2/O/sub 2//CI- system lowered the viscosity of HA. Analysis of 3H-HA exposed to OCI- by gel filtration chromatography indicated that cleavage of HA occurred only at higher OCI- concentrations. We suggest that the reduction in viscosity of HA by the MPO/H/sub 2/O/sub 2//CI- system may be due to a combination of oxidative cleavage and changes in the conformation of the molecule. We speculate that the changes in the molecular size of rheumatoid synovial fluid HA may be due to the action of the neutrophil MPO/H/sub 2/O/sub 2//CI- system.

  12. Two-electron reduction and one-electron oxidation of organic hydroperoxides by human myeloperoxidase.

    PubMed

    Furtmüller, P G; Burner, U; Jantschko, W; Regelsberger, G; Obinger, C

    2000-11-03

    The reaction of native myeloperoxidase (MPO) and its redox intermediate compound I with hydrogen peroxide, ethyl hydroperoxide, peroxyacetic acid, t-butyl hydroperoxide, 3-chloroperoxybenzoic acid and cumene hydroperoxide was studied by multi-mixing stopped-flow techniques. Hydroperoxides are decomposed by MPO by two mechanisms. Firstly, the hydroperoxide undergoes a two-electron reduction to its corresponding alcohol and heme iron is oxidized to compound I. At pH 7 and 15 degrees C, the rate constant of the reaction between 3-chloroperoxybenzoic acid and ferric MPO was similar to that with hydrogen peroxide (1.8x10(7) M(-1) s(-1) and 1.4x10(7) M(-1) s(-1), respectively). With the exception of t-butyl hydroperoxide, the rates of compound I formation varied between 5.2x10(5) M(-1) s(-1) and 2.7x10(6) M(-1) s(-1). Secondly, compound I can abstract hydrogen from these peroxides, producing peroxyl radicals and compound II. Compound I reduction is shown to be more than two orders of magnitude slower than compound I formation. Again, with 3-chloroperoxybenzoic acid this reaction is most effective (6. 6x10(4) M(-1) s(-1) at pH 7 and 15 degrees C). Both reactions are controlled by the same ionizable group (average pK(a) of about 4.0) which has to be in its conjugated base form for reaction.

  13. Resonance Raman microspectroscopy of myeloperoxidase and cytochrome b558 in human neutrophilic granulocytes.

    PubMed Central

    Sijtsema, N M; Otto, C; Segers-Nolten, G M; Verhoeven, A J; Greve, J

    1998-01-01

    With (resonance) Raman microscospectroscopy, it is possible to investigate the chemical constitution of a very small volume (0.5 fl) in a living cell. We have measured resonance Raman spectra in the cytoplasm of living normal, myeloperoxidase (MPO)-deficient, and cytochrome b558-deficient neutrophils and in isolated specific and azurophilic granule fractions, using an excitation wavelength of 413.1 nm. Similar experiments were performed after reduction of the redox centers by the addition of sodium dithionite. The specific and azurophilic granules in both redox states appeared to have clearly distinguishable Raman spectra when exciting at a wavelength of 413.1 nm. The azurophilic granules and the cytochrome b558-deficient neutrophils showed Raman spectra similar to that of the isolated MPO. The spectra of the specific granules and the MPO-deficient neutrophils corresponded very well to published cytochrome b558 spectra. The resonance Raman spectrum of the cytoplasmic region of normal neutrophilic granulocytes could be fitted with a combination of the spectra of the specific and azurophilic granules, which shows that the Raman signal of neutrophilic granulocytes mainly originates from MPO and cytochrome b558, at an excitation wavelength of 413.1 nm. PMID:9635778

  14. Myeloperoxidase-antineutrophil cytoplasmic antibody-associated crescentic glomerulonephritis in autosomal dominant polycystic kidney disease

    PubMed Central

    2013-01-01

    Background Autosomal dominant polycystic kidney disease (ADPKD) is an inherited disorder that is characterized by the development of cysts in the kidneys and other organs. Urinary protein excretion is usually less than 1 g/day, and ADPKD is rarely associated with nephrotic syndrome or rapidly progressive glomerulonephritis (RPGN). To date, myeloperoxidase (MPO)-antineutrophil cytoplasmic antibody (ANCA)-associated crescentic glomerulonephritis (CrGN) has not been reported in a patient with ADPKD. Case presentations We report two cases of MPO-ANCA positive ADPKD. A 60-year-old Japanese woman (case 1) and a 54-year-old Japanese woman (case 2) presented with RPGN featuring severe proteinuria and microscopic hematuria. In both patients percutaneous needle biopsy of the kidney revealed MPO-ANCA-associated CrGN with a paucity of glomerular immunoglobulin staining. Each patient received intravenous methylprednisolone for 3 days, followed by oral prednisolone. Case 1 showed gradual improvement and has not progressed to end-stage renal disease (ESRD), but case 2 developed ESRD requiring hemodialysis within one month despite treatment. Conclusion These are the first two reported cases of MPO-ANCA-associated CrGN in patients with ADPKD. Our experience suggests that serial measurement of the ANCA titer and renal biopsy should be considered for accurate diagnosis and appropriate treatment of ADPKD patients who present with proteinuria, hematuria, and rapid decline of renal function. PMID:23617397

  15. Evaluation of serum myeloperoxidase concentration in dogs with heart failure due to chronic mitral valvular insufficiency.

    PubMed

    Park, Jong-In; Suh, Sang-Il; Hyun, Changbaig

    2017-01-01

    Myeloperoxidase (MPO) is a leukocyte-derived enzyme involved in the process of heart failure and is found to have good diagnostic and prognostic values in humans with chronic heart failure. This study evaluated the relationship between serum MPO levels and the severity of heart failure (HF) due to chronic mitral valvular insufficiency (CMVI) in dogs. Eighty-two client-owned dogs consisting of 69 dogs with different stages of HF due to CMVI and 13 age-matched healthy dogs were enrolled in this study. Serum MPO concentrations in the healthy and CMVI groups were determined by enzyme-linked immunosorbent assay (ELISA) using a canine-specific monoclonal anti-MPO antibody. Serum MPO concentrations were 273.3 ± 179.6 ng/L in the controls, 140.8 ± 114.1 ng/L in the International Small Animal Cardiac Health Council (ISACHC) I group, 109.0 ± 85.2 ng/L in the ISACHC II group, and 106.0 ± 42.3 ng/L in the ISACHC III group. Close negative correlation to serum MPO concentration was found in the severity of heart failure (ISACHC stage). Although this study found a modest relationship between serum MPO levels and the severity of HF due to CMVI in dogs, it also suggested that serum MPO levels decreased as the severity of HF increased.

  16. Immunohistochemical Expression of Myeloperoxidase in Placental Samples of Systematic Lupus Erythematosus Pregnancies

    PubMed Central

    Heidari, Zahra; Mahmoudzadeh Sagheb, Hamidreza; Sheibak, Nadia

    2016-01-01

    Objective: Reports of increased level of Myeloperoxidase (MPO) in plasma and placental extracts of Systematic Lupus Erythematosus (SLE) has been proposed that MPO may have an important role in this pregnancy complication. In present study immunohistochemical MPO expression was investigated in placental samples of SLE women compared with normal controls. Materials and methods: Ten patients with SLE were recruited as case group. Control group was selected from mothers with normal uncomplicated pregnancies. A monoclonal antibody specific for MPO was used for immunohistochemical staining and then the staining was quantified and differences between groups were compared using Mann-Whitney U test. Results: There were significant differences in the expression levels of MPO in the syncytiotrophoblast cells and the extravillous trophoblast cells between the control and SLE groups (p < 0.05). There were no significant differences in the expression level of MPO in the vascular endothelium and the relative number of the MPO-positive leukocytes in placental tissue between SLE and control groups (p > 0.05). Conclusion: The present study showed that MPO expression is increased in syncytiotrophoblast cells and the extravillous trophoblast cells of SLE placentas compared to healthy subjects. It seems that these changes are able to impress structure and function of placenta and survival of the fetus. PMID:27648095

  17. Quick staining technique for myeloperoxidase using potassium iodide and oxidized pyronine B.

    PubMed

    Chen, Wan-Xin; Zhu, Hong-Lin; Xue, Mei; Zhou, Hao; Zhao, Fei; Yan, Ni; Chen, Yan

    2014-01-01

    Myeloperoxidase (MPO) staining has been important for the cytomorphological diagnosis and classification of leukemia. A novel staining method for MPO and its clinical application are presented in the report. Pyronine B (PyB), serving as a chromogenic reagent, was pre-oxidized to obtain stable oxidized Pyronine B solution. The MPO working solution for oxidized pyronine B method consisted of phosphate buffer solution, potassium iodide (KI) solution, and oxidized Pyronine B solution. The positive products of the oxidized Pyronine B method of MPO staining were vibrant red particles located in cytoplasm and the nucleus was stained bluish green. Bone marrow smears from 229 patients with acute leukemia or with grossly normal bone marrow were stained by both oxidized Pyronine B method and the conventional Washburn benzidine staining and a comparison revealed no significant difference in the positive detection rate between the two techniques. The new method eliminates the influence of the varying amount of H₂O₂ on MPO staining. With this method, the reagents were more stable and the staining procedure was simple and time-saving. This MPO staining technique is a better alternative than the conventional benzidine-based methods.

  18. Pediatric case of crescentic post-streptococcal glomerulonephritis with myeloperoxidase anti-neutrophil cytoplasmic antibody.

    PubMed

    Kanai, Hiroaki; Sawanobori, Emi; Koizumi, Keiichi; Ohashi, Ryuji; Higashida, Kosuke

    2015-04-01

    Post-streptococcal glomerulonephritis (PSGN) generally has a good renal prognosis, and immunosuppressive therapies are not needed. However, a few patients present with severe acute kidney injury and extensive crescent formations. The etiology of such patients is not well known, and involvement of anti-neutrophil cytoplasmic antibodies is rarely reported. A 9-year-old girl with rapidly progressive nephritic syndrome was diagnosed with PSGN. A biopsy showed diffuse crescentic glomerulonephritis with immunoglobulin G and C3 deposits; moreover, humps were observed on electron microscopy. After she was administered methylprednisolone pulse therapy and intravenous cyclophosphamide, followed by prednisolone and azathioprine therapy, her urinary abnormalities improved and renal function normalized. However, the myeloperoxidase anti-neutrophil cytoplasmic antibody (MPO-ANCA) titers gradually increased. We speculated that PSGN may be augmented by increased MPO-ANCA levels. Therefore, the patient is currently being treated with losartan, enalapril, azathioprine, and prednisolone. Although the MPO-ANCA titer remains high, urinary findings show mild proteinuria and her renal function has been norma for 18 months since onset. A progressive clinical course and severe histological findings may indicate the involvement of ANCA in deterioration of condition in patients with PSGN. Furthermore, in such cases immunosuppressive therapies should be considered even in pediatric PSGN.

  19. Activation of PKR Causes Amyloid ß-Peptide Accumulation via De-Repression of BACE1 Expression

    PubMed Central

    ILL-Raga, Gerard; Palomer, Ernest; Wozniak, Matthew A.; Ramos-Fernández, Eva; Bosch-Morató, Mònica; Tajes, Marta; Guix, Francesc X.; Galán, José J.; Clarimón, Jordi; Antúnez, Carmen; Real, Luis M.; Boada, Mercé; Itzhaki, Ruth F.; Fandos, César; Muñoz, Francisco J.

    2011-01-01

    BACE1 is a key enzyme involved in the production of amyloid ß-peptide (Aß) in Alzheimer's disease (AD) brains. Normally, its expression is constitutively inhibited due to the presence of the 5′untranslated region (5′UTR) in the BACE1 promoter. BACE1 expression is activated by phosphorylation of the eukaryotic initiation factor (eIF)2-alpha, which reverses the inhibitory effect exerted by BACE1 5′UTR. There are four kinases associated with different types of stress that could phosphorylate eIF2-alpha. Here we focus on the double-stranded (ds) RNA-activated protein kinase (PKR). PKR is activated during viral infection, including that of herpes simplex virus type 1 (HSV1), a virus suggested to be implicated in the development of AD, acting when present in brains of carriers of the type 4 allele of the apolipoprotein E gene. HSV1 is a dsDNA virus but it has genes on both strands of the genome, and from these genes complementary RNA molecules are transcribed. These could activate BACE1 expression by the PKR pathway. Here we demonstrate in HSV1-infected neuroblastoma cells, and in peripheral nervous tissue from HSV1-infected mice, that HSV1 activates PKR. Cloning BACE1 5′UTR upstream of a luciferase (luc) gene confirmed its inhibitory effect, which can be prevented by salubrinal, an inhibitor of the eIF2-alpha phosphatase PP1c. Treatment with the dsRNA analog poly (I∶C) mimicked the stimulatory effect exerted by salubrinal over BACE1 translation in the 5′UTR-luc construct and increased Aß production in HEK-APPsw cells. Summarizing, our data suggest that PKR activated in brain by HSV1 could play an important role in the development of AD. PMID:21738672

  20. Accumulation of recalcitrant xylan in mushroom-compost is due to a lack of xylan substituent removing enzyme activities of Agaricus bisporus.

    PubMed

    Jurak, Edita; Patyshakuliyeva, Aleksandrina; Kapsokalyvas, Dimitris; Xing, Lia; van Zandvoort, Marc A M J; de Vries, Ronald P; Gruppen, Harry; Kabel, Mirjam A

    2015-11-05

    The ability of Agaricus bisporus to degrade xylan in wheat straw based compost during mushroom formation is unclear. In this paper, xylan was extracted from the compost with water, 1M and 4M alkali. Over the phases analyzed, the remaining xylan was increasingly substituted with (4-O-methyl-)glucuronic acid and arabinosyl residues, both one and two arabinosyl residues per xylosyl residue remained. In the 1M and 4M KOH soluble solids of spent compost, 33 and 49 out of 100 xylosyl residues, respectively, were substituted. The accumulation of glucuronic acid substituents matched with the analysis that the two A. bisporus genes encoding for α-glucuronidase activity (both GH115) were not expressed in the A. bisporus mycelium in the compost during fruiting. Also, in a maximum likelihood tree it was shown that it is not likely that A. bisporus possesses genes encoding for the activity to remove arabinose from xylosyl residues having two arabinosyl residues.

  1. Rare Earth Ion Mediated Fluorescence Accumulation on a Single Microbead: An Ultrasensitive Strategy for the Detection of Protein Kinase Activity at the Single-Cell Level.

    PubMed

    Zhang, Xiaobo; Liu, Chenghui; Wang, Honghong; Wang, Hui; Li, Zhengping

    2015-12-07

    A single microbead-based fluorescence imaging (SBFI) strategy that enables detection of protein kinase activity from single cell lysates is reported. We systematically investigated the ability of various rare earth (RE) ions, immobilized on the microbead, for specific capturing of kinase-induced phosphopeptides, and Dy(3+) was found to be the most prominent one. Through the efficient concentration of kinase-induced fluorescent phosphopeptides on a Dy(3+) -functionalized single microbead, kinase activity can be detected and quantified by reading the fluorescence on the microbead with a confocal fluorescence microscope. Owing to the extremely specific recognition of Dy(3+) towards phosphopeptides and the highly-concentrated fluorescence accumulation on only one microbead, ultrahigh sensitivity has been achieved for the SBFI strategy which allows direct kinase analysis at the single-cell level.

  2. VRK3-mediated nuclear localization of HSP70 prevents glutamate excitotoxicity-induced apoptosis and Aβ accumulation via enhancement of ERK phosphatase VHR activity

    PubMed Central

    Song, Haengjin; Kim, Wanil; Kim, Sung-Hoon; Kim, Kyong-Tai

    2016-01-01

    Most of neurodegenerative disorders are associated with protein aggregation. Glutamate-induced excitotoxicity and persistent extracellular signal-regulated kinase (ERK) activation are also implicated in neurodegenerative diseases. Here, we found that vaccinia-related kinase 3 (VRK3) facilitates nuclear localization of glutamate-induced heat shock protein 70 (HSP70). Nuclear HSP70 leads to enhancement of vaccinia H1-related phosphatase (VHR) activity via protein-protein interaction rather than its molecular chaperone activity, thereby suppressing excessive ERK activation. Moreover, glutamate-induced ERK activation stimulates the expression of HSP70 and VRK3 at the transcriptional level. Downregulation of either VRK3 or HSP70 rendered cells vulnerable to glutamate-induced apoptosis. Overexpression of HSP70 fused to a nuclear localization signal attenuated apoptosis more than HSP70 alone. The importance of nuclear localization of HSP70 in the negative regulation of glutamate-induced ERK activation was further confirmed in VRK3-deficient neurons. Importantly, we showed a positive correlation between levels of VRK3 and HSP70 in the progression of Alzheimer’s and Parkinson’s diseases in humans, and neurons with HSP70 nuclear localization exhibited less Aβ accumulation in brains from patients with Alzheimer’s disease. Therefore, HSP70 and VRK3 could potentially serve as diagnostic and therapeutic targets in neurodegenerative diseases. PMID:27941812

  3. Lysophosphatidic acid mediates the rapid activation of platelets and endothelial cells by mildly oxidized low density lipoprotein and accumulates in human atherosclerotic lesions.

    PubMed

    Siess, W; Zangl, K J; Essler, M; Bauer, M; Brandl, R; Corrinth, C; Bittman, R; Tigyi, G; Aepfelbacher, M

    1999-06-08

    Oxidized low density lipoprotein (LDL) is a key factor in the pathogenesis of atherosclerosis and its thrombotic complications, such as stroke and myocardial infarction. It activates endothelial cells and platelets through mechanisms that are largely unknown. Here, we show that lysophosphatidic acid (LPA) was formed during mild oxidation of LDL and was the active compound in mildly oxidized LDL and minimally modified LDL, initiating platelet activation and stimulating endothelial cell stress-fiber and gap formation. Antagonists of the LPA receptor prevented platelet and endothelial cell activation by mildly oxidized LDL. We also found that LPA accumulated in and was the primary platelet-activating lipid of atherosclerotic plaques. Notably, the amount of LPA within the human carotid atherosclerotic lesion was highest in the lipid-rich core, the region most thrombogenic and most prone to rupture. Given the potent biological activity of LPA on platelets and on cells of the vessel wall, our study identifies LPA as an atherothrombogenic molecule and suggests a possible strategy to prevent and treat atherosclerosis and cardiocerebrovascular diseases.

  4. Effect of lipopolysaccharide on protein accumulation by murine peritoneal macrophages: the correlation to activation for macrophage tumoricidal function

    SciTech Connect

    Tannenbaum, C.S.

    1987-01-01

    The protein synthetic patterns of tumoricidal murine peritoneal macrophage populations have been compared to those of non-tumoricidal populations utilizing two dimensional polyacrylamide gel electrophoresis (2D PAGE) of (/sup 35/S)-methionine-labeled proteins. While the protein synthetic patterns exhibited by resident, inflammatory and activated macrophages had numerous common features which distinguished them from the other normal non-macrophage cell types examined, unique proteins also distinguished each macrophage population from the others. Peritoneal macrophages elicited by treatment with heat killed Propionibacterium acnes, the live, attenuated Mycobacterium bovis strain BCG, Listeria monocytogenes and the protozoan flagellate Trypanosoma rhodesiense, all exhibited tumoricidal activity in 16h or 72h functional assays, and shared a common protein synthetic profile which differentiated them from the synthetic patterns characteristic of the non-tumoricidal resident and inflammatory macrophages.

  5. The involvement of polyphenols and peroxidase activities in heavy-metal accumulation by epidermal glands of the waterlily (Nymphaeaceae).

    PubMed

    Lavid, N; Schwartz, A; Yarden, O; Tel-Or, E

    2001-02-01

    Co-localization of polyphenols and peroxidase activity was demonstrated in epidermal glands of the waterlily (Nymphaea) by histochemistry. Total phenols, tannins and peroxidase activity were determined quantitatively in plant extracts. Polyphenols were partially identified and were found to consist mainly of hydrolyzable tannins, gallic and tannic acid derivatives. Nymphaea polyphenols were shown to chelate Cr, Hg, and Pb in vitro, and Cd-binding by polymerized polyphenols was demonstrated in leaves exposed to Cd in vivo. Both polyphenols and peroxidases were found at very high constitutive levels, which were not induced or altered by external conditions, such as light and heavy-metal stress. It is suggested that the polymerization of polyphenols by peroxidases, enhanced after heavy-metal uptake and detoxification, is responsible for the binding of heavy metals in Nymphaea epidermal glands.

  6. Active moss biomonitoring applied to an industrial site in Romania: relative accumulation of 36 elements in moss-bags.

    PubMed

    Culicov, O A; Mocanu, R; Frontasyeva, M V; Yurukova, L; Steinnes, E

    2005-09-01

    Active moss biomonitoring using the species Sphagnum girgensohnii was tested at a strongly polluted site in Romania (Baia Mare) according to a novel sampling design. Nine moss transplants from each of the two background areas (Dubna, Russia and Vitosha Mountain, Bulgaria) were deployed in parallel on balconies about 24 m above street level for 4 months. The samples were analyzed for 36 elements using instrumental neutron activation analysis (INAA). Based on the results obtained the sampling variability is discussed in relation to the analytical variability, and the relative uptake of the different elements is assessed. The moss-bags using Sphagnum girgensohnii demonstrate a high or a very high relative uptake for a majority of the 36 investigated elements, but the values depend on the initial element concentration in the moss. Moss leaves analyzed separately showed somewhat higher levels than stems for many elements. Practical considerations however still speak in favor of using the whole moss for transplants.

  7. Partial deficit of pantothenate kinase 2 catalytic activity in a case of tremor-predominant neurodegeneration with brain iron accumulation.

    PubMed

    Liang, Tsao-Wei; Truax, Adam C; Trojanowski, John Q; Lee, Virginia M-Y; Stern, Matthew B; Kotzbauer, Paul T

    2006-05-01

    We describe an atypical case of pantothenate kinase-associated neurodegeneration (PKAN) in which slowly progressive arm tremor was the predominant symptom beginning at the age of 25, with late-onset dystonia and dysarthria developing at the age of 50. Compound heterozygous mutations resulting in missense amino acid substitutions G521R and I529V were identified in the pantothenate kinase (PANK2) gene. We demonstrate that while the G521R mutation results in an unstable and inactive protein, the previously unreported I529V substitution has no apparent effect on the stability or catalytic activity of PanK2. The phenotype that results from this combination of mutations suggests that atypical presentations of PKAN may arise from partial deficits in PanK2 catalytic activity.

  8. Enhancing cell nucleus accumulation and DNA cleavage activity of anti-cancer drug via graphene quantum dots.

    PubMed

    Wang, Chong; Wu, Congyu; Zhou, Xuejiao; Han, Ting; Xin, Xiaozhen; Wu, Jiaying; Zhang, Jingyan; Guo, Shouwu

    2013-10-04

    Graphene quantum dots (GQDs) maintain the intrinsic layered structural motif of graphene but with smaller lateral size and abundant periphery carboxylic groups, and are more compatible with biological system, thus are promising nanomaterials for therapeutic applications. Here we show that GQDs have a superb ability in drug delivery and anti-cancer activity boost without any pre-modification due to their unique structural properties. They could efficiently deliver doxorubicin (DOX) to the nucleus through DOX/GQD conjugates, because the conjugates assume different cellular and nuclear internalization pathways comparing to free DOX. Also, the conjugates could enhance DNA cleavage activity of DOX markedly. This enhancement combining with efficient nuclear delivery improved cytotoxicity of DOX dramatically. Furthermore, the DOX/GQD conjugates could also increase the nuclear uptake and cytotoxicity of DOX to drug-resistant cancer cells indicating that the conjugates may be capable to increase chemotherapy efficacy of anti-cancer drugs that are suboptimal due to the drug resistance.

  9. Sodium chloride enhances cadmium tolerance through reducing cadmium accumulation and increasing anti-oxidative enzyme activity in tobacco.

    PubMed

    Zhang, Bing-Lin; Shang, Sheng-Hua; Zhang, Hai-Tao; Jabeen, Zahra; Zhang, Guo-Ping

    2013-06-01

    The effect of sodium chloride (NaCl) on cadmium (Cd) uptake, translocation, and oxidative stress was investigated using 2 tobacco cultivars differing in Cd tolerance. The growth inhibition of the tobacco plants exposed to Cd toxicity was in part alleviated by moderate addition of NaCl in the culture solution. Cadmium concentration of shoots and roots in the 2 cultivars increased with increasing Cd levels in the solution and decreased with the addition of NaCl. The addition of NaCl could alleviate the oxidative stress caused by Cd toxicity, as reflected by reduced production of malondialdehyde and recovered or enhanced activities of antioxidative enzymes catalase and glutathione peroxidase. The results also showed that the enhancement of antioxidative enzyme activity by NaCl for the tobacco plants exposed to Cd stress is related to induced Ca signaling.

  10. Activation of mTOR signaling leads to orthopedic surgery-induced cognitive decline in mice through β-amyloid accumulation and tau phosphorylation.

    PubMed

    Shen, Wenzhen; Lu, Keliang; Wang, Jiawan; Wu, Anshi; Yue, Yun

    2016-10-01

    Postoperative cognitive dysfunction (POCD) is a serious complication following surgery, however, the mechanism of POCD remains to be elucidated. Previous evidence has revealed that POCD may be associated with the pathogenesis of neurodegenerative processes. The mammalian target of rapamycin (mTOR) signaling pathway has been reported to be crucial in the pathophysiology of neurodegenerative diseases. However, the implications of mTOR in POCD remains to be fully elucidated. In the present study, western blotting and enzyme‑linked immunosorbent assay were used to determine the expression of mTOR and any associated downstream targets; contextual fear conditioning was used to estimate the learning and memory ability of mice. Using an animal model of orthopedic surgery, it was found that surgical injury impaired hippocampal‑dependent memory and enhanced the levels of phosphorylated mTOR at Serine‑2448, phosphorylated 70‑kDa ribosomal protein S6 kinase (p70S6K) at Threonine‑389 with accumulation of β‑amyloid (Aβ) and hyperphosphorylated tau at Serine-396, compared with the control group. Pretreatment with rapamycin, an mTOR inhibitor, restored the abnormal mTOR/p70S6K signaling induced by surgery, attenuated the accumulation of Aβ and reduced the phosphorylation of tau protein. Rapamycin also reversed the surgery‑induced cognitive dysfunction. The results of the present study suggested that the surgical stimulus activated mTOR/p70S6K signaling excessively, and that the inhibition of mTOR signaling with rapamycin may prevent postoperative cognitive deficits, partly through attenuating the accumulation of Aβ and hyperphosphorylation of tau protein.

  11. Impact of high dietary zinc on zinc accumulation, enzyme activity and proteomic profiles in the pancreas of piglets.

    PubMed

    Pieper, R; Martin, L; Schunter, N; Villodre Tudela, C; Weise, C; Klopfleisch, R; Zentek, J; Einspanier, R; Bondzio, A

    2015-04-01

    The exocrine pancreas plays an important role in zinc homeostasis. Feeding very high (2000-3000mgzinc/kg diet) levels of zinc oxide to piglets for short periods is a common practice in the swine industry to improve performance and prevent diseases. The impact on pancreatic function and possible side effects during long-term feeding of high dietary zinc levels are still poorly understood. A total of 54 weaned piglets were either fed with low (57mg/kg, LZn), normal (164mg/kg, NZn) or high (2425mg/kg, HZn) zinc concentration in the diets. After 4 weeks of feeding, ten piglets per treatment were euthanized and pancreas samples were taken. Tissue zinc concentration and metallothionein abundance was greater with HZn compared with NZn and LZn (P<0.05). Similarly, activity of α-amylase, lipase, trypsin and chymotrypsin was higher with HZn as compared with NZn and LZn diets (P<0.05), whereas elastase activity was unchanged. Total trolox equivalent antioxidative capacity of pancreas tissue was higher with HZn diets compared with the other treatments (P<0.05). Pancreatic protein profiles of NZn and HZn fed piglets were obtained by 2D-DIGE technique and revealed 15 differentially expressed proteins out of 2100 detected spots (P<0.05). The differentially expressed proteins aldose reductase, eukaryotic elongation factor II and peroxiredoxin III were confirmed by immunoblotting. Identified proteins include zinc finger-containing transcription factors and proteins mainly associated with oxidative stress response and signal transduction in HZn compared with NZn pigs. Histologic examination however showed no morphologic changes. The results suggest that long-term supply of very high dietary zinc increases zinc and metallothionein concentration, and digestive enzyme activity, but also triggers oxidative stress reactions in the pancreas of young pigs. The data provide new insights into pancreatic function under outbalanced zinc homeostasis.

  12. Hydrogen peroxide-induced antioxidant activities and cardiotonic glycoside accumulation in callus cultures of endemic Digitalis species.

    PubMed

    Cingoz, Gunce Sahin; Verma, Sandeep Kumar; Gurel, Ekrem

    2014-09-01

    The effect of hydrogen peroxide (H2O2) on callus cultures of four Digitalis species (Digitalis lamarckii, Digitalis trojana, Digitalis davisiana and Digitalis cariensis) increased catalase (CAT), superoxide dismutase (SOD), total phenolic, proline activity and cardiotonic glycoside production. Callus derived from hypocotyl explants was cultured on Murashige and Skoog medium supplemented with 0.25 mg L(-1) indole-3-acetic acid (IAA) and 0.5 mg L(-1) thidiazuron (TDZ). After a month of culture, callus was transferred to MS medium containing 10 mM H2O2 and then incubated for 6 h. The amount of five cardenolides (Lanatoside C, Digitoxin, Digoxigenin, Gitoxigenin and Digoxin) as well as CAT, SOD, total phenolic, proline activity from Digitalis species were compared. No digoxin was detected in all treatments and control groups. The total cardenolides estimated were in the order of D. lamarckii (586.65  μg g(-1) dw), D. davisiana (506.79 μg g(-1) dw), D. cariensis (376.60 μg g(-1) dw) and D. trojana (282.39 μg g(-1) dw). It was clear that H2O2 pre-treatment resulted in an increase in enzymatic and nonenzymatic antioxidants. However, a significant negative relationship between cardenolides production and overall activities of CAT, SOD, total phenolic and proline was evident. The described protocol here will be useful for the development of new strategies for a large-scale production of cardenolides.

  13. Anthocyanins inhibit high-glucose-induced cholesterol accumulation and inflammation by activating LXRα pathway in HK-2 cells.

    PubMed

    Du, Chunyang; Shi, Yonghong; Ren, Yunzhuo; Wu, Haijiang; Yao, Fang; Wei, Jinying; Wu, Ming; Hou, Yanjuan; Duan, Huijun

    2015-01-01

    The dysregulation of cholesterol metabolism and inflammation plays a significant role in the progression of diabetic nephropathy (DN). Anthocyanins are polyphenols widely distributed in food and exert various biological effects including antioxidative, anti-inflammatory, and antihyperlipidemic effects. However, it remains unclear whether anthocyanins are associated with DN, and the mechanisms involved in the reciprocal regulation of inflammation and cholesterol efflux are yet to be elucidated. In this study, we evaluated the regulation of cholesterol metabolism and the anti-inflammatory effects exerted by anthocyanins (cyanidin-3-O-β-glucoside chloride [C3G] or cyanidin chloride [Cy]) and investigated the underlying molecular mechanism of action using high-glucose (HG)-stimulated HK-2 cells. We found that anthocyanins enhanced cholesterol efflux and ABCA1 expression markedly in HK-2 cells. In addition, they increased peroxisome proliferator-activated receptor alpha (PPARα) and liver X receptor alpha (LXRα) expression and decreased the HG-induced expression of the proinflammatory cytokines intercellular adhesion molecule-1 (ICAM1), monocyte chemoattractant protein-1 (MCP1), and transforming growth factor-β1 (TGFβ1), as well as NFκB activation. Incubation with the PPARα-specific inhibitor GW6471 and LXRα shRNA attenuated the anthocyanin-mediated promotion of ABCA1 expression and cholesterol efflux, suggesting that anthocyanins activated PPARα-LXRα-ABCA1-dependent cholesterol efflux in HK-2 cells. Moreover, the knockout of LXRα abrogated the anti-inflammatory effect of anthocyanins, whereas the PPARα antagonist GW6471 does not have this effect. Further investigations revealed that LXRα might interfere with anthocyanin-induced decreased ICAM1, MCP1, and TGFβ1 expression by reducing the nuclear translocation of NFκB. Collectively, these findings suggest that blocking cholesterol deposition and inhibiting the LXRα pathway-induced inflammatory response

  14. Constant-load exercise decreases the serum concentration of myeloperoxidase in healthy smokers and smokers with COPD.

    PubMed

    Holz, Olaf; Roepcke, Stefan; Watz, Henrik; Tegtbur, Uwe; Lahu, Gezim; Hohlfeld, Jens M

    2015-01-01

    There is an ongoing demand for easily accessible biomarkers related to pathophysiological processes in chronic obstructive pulmonary disease (COPD). Short-term intense exercise is known to increase the peripheral blood levels of cytokines. Therefore, we tested the potential and the repeatability of an exercise challenge to amplify seven serum biomarkers (interleukin 6 [IL6], C-reactive protein [CRP], myeloperoxidase [MPO], leukotriene B4, soluble intercellular adhesion molecule 1, soluble vascular cell adhesion molecule 1, and von Willebrand factor [VWF]) in smokers with and without COPD. Twenty-three smokers with moderate COPD (GOLD 2) and 23 sex- and age-matched healthy smokers underwent up to 30-minute submaximal, constant-load exercise (75% of maximum work load) on two occasions separated by 4 weeks (second challenge n=19/20). Serum samples were obtained before, 5 minutes after the start, at the end of exercise (maximum 30 minutes or until exhaustion), and after additional 20 minutes of rest. The median (interquartile range) exercise time until exhaustion in the two challenges was 10.0 (4.0) minutes and 10.0 (8.0) minutes in smokers with COPD and 22.0 (16.0) minutes and 26.5 (14.5) minutes in healthy smokers. The exercise challenge significantly increased the serum concentrations of IL6 and VWF, but decreased the concentrations of MPO. Healthy smokers showed a significantly greater increase (at the end of exercise compared to before exercise) in IL6 (P=0.01) and a larger decline (P=0.03) in MPO. The overall profile of the serum markers during the exercise challenge was shown to be repeatable in the second challenge. In summary, intense load exercise is capable of changing the concentration of inflammatory and endothelial function markers. Especially, the decline in the level of MPO, a marker closely related to cardiovascular risk, appears to be of clinical interest, as the exercise-induced decline might be related to the beneficial effects of physical activity

  15. Both microtubule-stabilizing and microtubule-destabilizing drugs inhibit hypoxia-inducible factor-1alpha accumulation and activity by disrupting microtubule function.

    PubMed

    Escuin, Daniel; Kline, Erik R; Giannakakou, Paraskevi

    2005-10-01

    We have recently identified a mechanistic link between disruption of the microtubule cytoskeleton and inhibition of tumor angiogenesis via the hypoxia-inducible factor-1 (HIF-1) pathway. Based on this model, we hypothesized that other microtubule-targeting drugs may have a similar effect on HIF-1alpha. To test that hypothesis, we studied the effects of different clinically relevant microtubule-disrupting agents, including taxotere, epothilone B, discodermolide, vincristine, 2-methoxyestradiol, and colchicine. In all cases, HIF-1alpha protein, but not mRNA, was down-regulated in a drug dose-dependent manner. In addition, HIF-1alpha transcriptional activity was also inhibited by all drugs tested. To further examine whether these effects were dependent on microtubule network disruption, we tested the ability of epothilone B to inhibit HIF-1alpha protein in the human ovarian cancer cell line 1A9 and its beta-tubulin mutant epothilone-resistant subclone 1A9/A8. Our data showed that epothilone B treatment down-regulated HIF-1alpha protein in the parental 1A9 cells but had no effect in the resistant 1A9/A8 cells. These observations were confirmed by confocal microscopy, which showed impaired nuclear accumulation of HIF-1alpha in parental 1A9 cells at epothilone B concentrations that induced extensive microtubule stabilization. In contrast, epothilone B treatment had no effect on either microtubules or HIF-1alpha nuclear accumulation in the resistant 1A9/A8 cells. Furthermore, epothilone B inhibited HIF-1 transcriptional activity in 1A9 cells, as evidenced by a hypoxia response element-luciferase reporter assay, but had no effect on HIF-1 activity in the resistant 1A9/A8 cells. These data directly link beta-tubulin drug binding with HIF-1alpha protein inhibition. Our results further provide a strong rationale for testing taxanes and epothilones in clinical trials targeting HIF-1 in cancer patients.

  16. Accumulation of mutant alpha1-antitrypsin Z in the endoplasmic reticulum activates caspases-4 and -12, NFkappaB, and BAP31 but not the unfolded protein response.

    PubMed

    Hidvegi, Tunda; Schmidt, Bela Z; Hale, Pamela; Perlmutter, David H

    2005-11-25

    In alpha(1)-antitrypsin (alpha1AT) deficiency, a polymerogenic mutant form of the secretory glycoprotein alpha1AT, alpha1ATZ, is retained in the endoplasmic reticulum (ER) of liver cells. It is not yet known how this results in liver injury in a subgroup of deficient individuals and how the remainder of deficient individuals escapes liver disease. One possible explanation is that the "susceptible" subgroup is unable to mount the appropriate protective cellular responses. Here we examined the effect of mutant alpha1ATZ on several potential protective signaling pathways by using cell lines with inducible expression of mutant alpha1AT as well as liver from transgenic mice with liver-specific inducible expression of mutant alpha1AT. The results show that ER retention of polymerogenic mutant alpha1ATZ does not result in an unfolded protein response (UPR). The UPR can be induced in the presence of alpha1ATZ by tunicamycin excluding the possibility that the pathway has been disabled. In striking contrast, ER retention of nonpolymerogenic alpha1AT mutants does induce the UPR. These results indicate that the machinery responsible for activation of the UPR can distinguish the physical characteristics of proteins that accumulate in the ER in such a way that it can respond to misfolded but not relatively ordered polymeric structures. Accumulation of mutant alpha1ATZ does activate specific signaling pathways, including caspase-12 in mouse, caspase-4 in human, NFkappaB, and BAP31, a profile that was distinct from that activated by nonpolymerogenic alpha1AT mutants.

  17. Effect of Ethanol Accumulation on Porcine Interferon-α Production by Pichia pastoris and Activities of Key Enzymes in Carbon Metabolism.

    PubMed

    Ding, Jian; Gao, Minjie; Hou, Guoli; Liang, Kexue

    2015-08-01

    In production of porcine interferon α (pIFN-α) by Pichia pastoris, improper glycerol feeding strategy leads to ethanol accumulation in the last stage of growth phase. In the present study, taking two runs with low ethanol accumulation under 2 g/L as control group, effects of long-term (>4 h) and instantaneous high ethanol concentration (>10 g/L) on pIFN-α production, and activities of key enzymes in carbon metabolism were discussed. As a result, compared with control group, pIFN-α expression level was decreased about 4~12 folds under long-term high ethanol concentration, from the level above 3 g/L to the level under 1 g/L; pIFN-α expression level was decreased about 8 folds under instantaneous high ethanol concentration, reaching to the low level of 0.42 g/L. The low production of pIFN-α was caused by the severe inhibitory effect of ethanol on these enzymes.

  18. Hydrolysis and volatile fatty acids accumulation of waste activated sludge enhanced by the combined use of nitrite and alkaline pH.

    PubMed

    Huang, Cheng; Liu, Congcong; Sun, Xiuyun; Sun, Yinglu; Li, Rui; Li, Jiansheng; Shen, Jinyou; Han, Weiqing; Liu, Xiaodong; Wang, Lianjun

    2015-12-01

    Volatile fatty acids (VFAs) production from anaerobic digestion of waste activated sludge (WAS) is often limited by the slow hydrolysis and/or poor substrate availability. Increased attention has been given to enhance the hydrolysis and acidification of WAS recently. This study presented an efficient and green strategy based on the combined use of nitrite pretreatment and alkaline pH to stimulate hydrolysis and VFA accumulation from WAS. Results showed that both proteins and polysaccharides increased in the presence of nitrite, indicating the enhancement of sludge solubilization and hydrolysis processes. Mechanism investigations showed that nitrite pretreatment could disintegrate the sludge particle and disperse extracellular polymeric substances (EPS). Then, anaerobic digestion tests demonstrated VFA production increased with nitrite treatment. The maximal VFA accumulation was achieved with 0.1 g N/L nitrite dosage and pH 10.0 at a sludge retention time (SRT) of 7 days, which was much higher VFA production in comparison with the blank, sole nitrite pretreatment, or sole pH 10. The potential analysis suggested that the combined nitrite pretreatment and alkaline pH is capable of enhancing WAS digestion with a great benefit for biological nutrient removal (BNR).

  19. Old Mice Accumulate Activated Effector CD4 T Cells Refractory to Regulatory T Cell-Induced Immunosuppression

    PubMed Central

    Harpaz, Idan; Bhattacharya, Udayan; Elyahu, Yehezqel; Strominger, Itai; Monsonego, Alon

    2017-01-01

    Chronic low-grade inflammation and reduced lymphocyte potency are implicated in the pathogenesis of major illnesses associated with aging. Whether this immune phenotype results from a loss of cell-mediated regulation or intrinsic dysregulated function of effector T cells (Teffs) requires further research. Here, we report that, as compared with young C57BL6 mice, old mice show an increased frequency of CD4+CD62L− Teffs with a dysregulated activated phenotype and markedly increased effector functions. Analysis of the frequency and suppressive function of CD4+FoxP3+ regulatory T cells (Tregs) indicates an increase in the frequency of FoxP3+ T cells with aging which, however, occurs within the CD4+CD25− T cells. Furthermore, whereas Tregs from young and old mice similarly suppress Teffs from young mice, both have a compromised suppressive capacity of Teffs from old mice, a phenomenon which is partially recovered in the presence of IL-2-producing CD4+CD62L+ non-Teffs. Finally, we observed that Teff subsets from old mice are enriched with IL-17A-producing T cells and exhibit intrinsically dysregulated expression of genes encoding cell-surface molecules and transcription factors, which play a key role in T-cell activation and regulation. We, thus, demonstrate an age-related impairment in the regulation of effector CD4 T cells, which may underlie the higher risk for destructive inflammation associated with aging. PMID:28382033

  20. Old Mice Accumulate Activated Effector CD4 T Cells Refractory to Regulatory T Cell-Induced Immunosuppression.

    PubMed

    Harpaz, Idan; Bhattacharya, Udayan; Elyahu, Yehezqel; Strominger, Itai; Monsonego, Alon

    2017-01-01

    Chronic low-grade inflammation and reduced lymphocyte potency are implicated in the pathogenesis of major illnesses associated with aging. Whether this immune phenotype results from a loss of cell-mediated regulation or intrinsic dysregulated function of effector T cells (Teffs) requires further research. Here, we report that, as compared with young C57BL6 mice, old mice show an increased frequency of CD4+CD62L- Teffs with a dysregulated activated phenotype and markedly increased effector functions. Analysis of the frequency and suppressive function of CD4+FoxP3+ regulatory T cells (Tregs) indicates an increase in the frequency of FoxP3+ T cells with aging which, however, occurs within the CD4+CD25- T cells. Furthermore, whereas Tregs from young and old mice similarly suppress Teffs from young mice, both have a compromised suppressive capacity of Teffs from old mice, a phenomenon which is partially recovered in the presence of IL-2-producing CD4+CD62L+ non-Teffs. Finally, we observed that Teff subsets from old mice are enriched with IL-17A-producing T cells and exhibit intrinsically dysregulated expression of genes encoding cell-surface molecules and transcription factors, which play a key role in T-cell activation and regulation. We, thus, demonstrate an age-related impairment in the regulation of effector CD4 T cells, which may underlie the higher risk for destructive inflammation associated with aging.

  1. Interaction of Yna1 and Yna2 Is Required for Nuclear Accumulation and Transcriptional Activation of the Nitrate Assimilation Pathway in the Yeast Hansenula polymorpha.

    PubMed

    Silvestrini, Lucia; Rossi, Beatrice; Gallmetzer, Andreas; Mathieu, Martine; Scazzocchio, Claudio; Berardi, Enrico; Strauss, Joseph

    2015-01-01

    A few yeasts, including Hansenula polymorpha are able to assimilate nitrate and use it as nitrogen source. The genes necessary for nitrate assimilation are organised in this organism as a cluster comprising those encoding nitrate reductase (YNR1), nitrite reductase (YNI1), a high affinity transporter (YNT1), as well as the two pathway specific Zn(II)2Cys2 transcriptional activators (YNA1, YNA2). Yna1p and Yna2p mediate induction of the system and here we show that their functions are interdependent. Yna1p activates YNA2 as well as its own (YNA1) transcription thus forming a nitrate-dependent autoactivation loop. Using a split-YFP approach we demonstrate here that Yna1p and Yna2p form a heterodimer independently of the inducer and despite both Yna1p and Yna2p can occupy the target promoter as mono- or homodimer individually, these proteins are transcriptionally incompetent. Subsequently, the transcription factors target genes containing a conserved DNA motif (termed nitrate-UAS) determined in this work by in vitro and in vivo protein-DNA interaction studies. These events lead to a rearrangement of the chromatin landscape on the target promoters and are associated with the onset of transcription of these target genes. In contrast to other fungi and plants, in which nuclear accumulation of the pathway-specific transcription factors only occur in the presence of nitrate, Yna1p and Yna2p are constitutively nuclear in H. polymorpha. Yna2p is needed for this nuclear accumulation and Yna1p is incapable of strictly positioning in the nucleus without Yna2p. In vivo DNA footprinting and ChIP analyses revealed that the permanently nuclear Yna1p/Yna2p heterodimer only binds to the nitrate-UAS when the inducer is present. The nitrate-dependent up-regulation of one partner protein in the heterodimeric complex is functionally similar to the nitrate-dependent activation of nuclear accumulation in other systems.

  2. Interaction of Yna1 and Yna2 Is Required for Nuclear Accumulation and Transcriptional Activation of the Nitrate Assimilation Pathway in the Yeast Hansenula polymorpha

    PubMed Central

    Silvestrini, Lucia; Rossi, Beatrice; Gallmetzer, Andreas; Mathieu, Martine; Scazzocchio, Claudio; Berardi, Enrico; Strauss, Joseph

    2015-01-01

    A few yeasts, including Hansenula polymorpha are able to assimilate nitrate and use it as nitrogen source. The genes necessary for nitrate assimilation are organised in this organism as a cluster comprising those encoding nitrate reductase (YNR1), nitrite reductase (YNI1), a high affinity transporter (YNT1), as well as the two pathway specific Zn(II)2Cys2 transcriptional activators (YNA1, YNA2). Yna1p and Yna2p mediate induction of the system and here we show that their functions are interdependent. Yna1p activates YNA2 as well as its own (YNA1) transcription thus forming a nitrate-dependent autoactivation loop. Using a split-YFP approach we demonstrate here that Yna1p and Yna2p form a heterodimer independently of the inducer and despite both Yna1p and Yna2p can occupy the target promoter as mono- or homodimer individually, these proteins are transcriptionally incompetent. Subsequently, the transcription factors target genes containing a conserved DNA motif (termed nitrate-UAS) determined in this work by in vitro and in vivo protein-DNA interaction studies. These events lead to a rearrangement of the chromatin landscape on the target promoters and are associated with the onset of transcription of these target genes. In contrast to other fungi and plants, in which nuclear accumulation of the pathway-specific transcription factors only occur in the presence of nitrate, Yna1p and Yna2p are constitutively nuclear in H. polymorpha. Yna2p is needed for this nuclear accumulation and Yna1p is incapable of strictly positioning in the nucleus without Yna2p. In vivo DNA footprinting and ChIP analyses revealed that the permanently nuclear Yna1p/Yna2p heterodimer only binds to the nitrate-UAS when the inducer is present. The nitrate-dependent up-regulation of one partner protein in the heterodimeric complex is functionally similar to the nitrate-dependent activation of nuclear accumulation in other systems. PMID:26335797

  3. The Use of Delta Neutrophil Index and Myeloperoxidase Index for Predicting Acute Complicated Appendicitis in Children

    PubMed Central

    Kim, Oh Hyun; Cha, Yong Sung; Hwang, Sung Oh; Jang, Ji Young; Choi, Eun Hee; Kim, Hyung Il; Cha, KyoungChul; Kim, Hyun; Lee, Kang Hyun

    2016-01-01

    Background In children with acute appendicitis, 30% to 75% present with a complication, such as perforation, and the early diagnosis of complications is known to improve outcomes. Serum delta neutrophil index (DNI) and myeloperoxidase index (MPXI) are new inflammatory markers, and thus, in the present study, the authors evaluated the predictive values of these two markers for the presence of a complication in children with acute appendicitis. Methods This retrospective observational study was conducted on 105 consecutive children (<12 years old) with acute appendicitis treated over a 31-month period. DNI, MPXI, C-reactive protein (CRP), and white blood cells (WBCs) were measured in an emergency department and investigated with respect to their abilities to predict the presence of acute complicated appendicitis. Results Twenty-nine of the 105 patients (median age, 9 years) were allocated to the complicated group (27.6%) and 76 to the non-complicated group (72.4%). Median serum DNI and CRP were significantly higher in the complicated group [0% vs. 2.2%, p<0.001 and 0.65 mg/dL vs. 8.0 mg/dL, p<0.001], but median MPXI was not (p = 0.316). Area under curve (AUC) for the ability of serum DNI and CRP to predict the presence of acute complicated appendicitis were 0.738 and 0.840, respectively. Multiple logistic regression analyses showed initial CRP [odds ratio 1.301, 95% confidence interval (1.092–1.549), p = 0.003] significantly predicted the presence of a complication. The optimal cutoff for serum CRP was 4.0 mg/dL (sensitivity 69%, specificity 83%, AUC 0.840). Conclusions Although serum DNI values were significantly higher in children with acute complicated appendicitis, no evidence was obtained to support the notion that serum DNI or serum MPXI aid the differentiation of acute complicated and non-complicated appendicitis in the ED setting. PMID:26859663

  4. PRISMA-combined Myeloperoxidase -463G/A gene polymorphism and coronary artery disease

    PubMed Central

    Li, Yan-Yan; Wang, Hui; Qian, Jin; Kim, Hyun Jun; Wu, Jing-jing; Wang, Lian-sheng; Zhou, Chuan-wei; Yang, Zhi-Jian; Lu, Xin-Zheng

    2017-01-01

    Abstract Background: Myeloperoxidase (MPO) -463G/A gene polymorphism may be associated with an increased risk of developing coronary artery disease (CAD). Studies on the subject, however, do not provide a clear consensus. This meta-analysis was performed to explore the relationship between MPO gene -463G/A polymorphism and CAD risk. Methods: This meta-analysis combines data from 4744 subjects from 9 independent studies. By using fixed or random effect models, the pooled odds ratios (ORs) and their corresponding 95% confidence intervals (CIs) were assessed. Results: Our analysis found a significant association between MPO gene -463G/A polymorphism and CAD in the whole population under all genetic models: allelic (OR: 0.68, 95% CI: 0.54–0.85, P = 0.0009), recessive (OR: 0.41, 95% CI: 0.22–0.76, P = 0.005), dominant (OR: 0.682, 95% CI: 0.534–0.871, P = 0.002), homozygous (OR: 0.36, 95% CI: 0.16–0.79, P = 0.01), heterozygous genetic model (OR: 0.832, 95% CI: 0.733–0.945, P = 0.004), and additive (OR: 0.64, 95% CI: 0.46–0.90, P = 0.01), especially in the Chinese subgroup (P < 0.05). On the contrary, we found no such relationship in the non-Chinese subgroup (P > 0.05). Conclusion: The MPO gene -463G/A polymorphism is associated with CAD risk, especially within the Chinese population. The A allele of MPO gene -463G/A polymorphism might protect the people from suffering the CAD risk. PMID:28328864

  5. A spliceosome intermediate with loosely associated tri-snRNP accumulates in the absence of Prp28 ATPase activity

    PubMed Central

    Boesler, Carsten; Rigo, Norbert; Anokhina, Maria M.; Tauchert, Marcel J.; Agafonov, Dmitry E.; Kastner, Berthold; Urlaub, Henning; Ficner, Ralf; Will, Cindy L.; Lührmann, Reinhard

    2016-01-01

    The precise role of the spliceosomal DEAD-box protein Prp28 in higher eukaryotes remains unclear. We show that stable tri-snRNP association during pre-catalytic spliceosomal B complex formation is blocked by a dominant-negative hPrp28 mutant lacking ATPase activity. Complexes formed in the presence of ATPase-deficient hPrp28 represent a novel assembly intermediate, the pre-B complex, that contains U1, U2 and loosely associated tri-snRNP and is stalled before disruption of the U1/5′ss base pairing interaction, consistent with a role for hPrp28 in the latter. Pre-B and B complexes differ structurally, indicating that stable tri-snRNP integration is accompanied by substantial rearrangements in the spliceosome. Disruption of the U1/5′ss interaction alone is not sufficient to bypass the block by ATPase-deficient hPrp28, suggesting hPrp28 has an additional function at this stage of splicing. Our data provide new insights into the function of Prp28 in higher eukaryotes, and the requirements for stable tri-snRNP binding during B complex formation. PMID:27377154

  6. Hydrolase and fructosyltransferase activities implicated in the accumulation of different chain size fructans in three Asteraceae species.

    PubMed

    Itaya, Nair M; Asega, Amanda F; Carvalho, Maria Angela M; Figueiredo-Ribeiro, Rita de Cássia L

    2007-09-01

    Fructans are widely distributed in Asteraceae from floras with seasonal growth and are thought to be involved in drought and freezing tolerance, in addition to storage function. Reserve organs of Vernonia herbacea and Viguiera discolor, from the cerrado, and of the perennial herb Smallanthus sonchifolius, endemic to Andean region, store over 80% inulin, with different DP (35, 150, and 15, respectively). The fructan pattern in Asteraceae species could be explained by characteristics of their respective 1-FFTs. Hydrolases and fructosyltransferases from S. sonchifolius, V. herbacea and V. discolor were analyzed in plants at the same environmental conditions. The higher 1-FEH activities found in the species with lower DP, S. sonchifolius and V. herbacea reinforce the hypothesis of the involvement of 1-FEH in fructan profile and suggest that the high DP fructan of V. discolor is a consequence of the low affinity of its 1-FEH to the native long chain inulin. Long term incubation with sucrose suggested that the affinity of 1-FFT of V. discolor for 1-kestose is low when compared to that of V. herbacea. Indeed 1-FFT from V. discolor was shown to be an hDP 1-FFT, preferring longer inulins as acceptors. Conversely, 1-FFT from V. herbacea seems to have a higher affinity for short fructo-oligosaccharides, including 1-kestose, as acceptor substrates. Differences in fructan enzymes of the three Asteraceae provide new information towards the understanding of fructan metabolism and control of carbon flow between low and high DP fructans.

  7. Soil organic carbon (SOC) accumulation in rice paddies under long-term agro-ecosystem experiments in southern China - VI. Changes in microbial community structure and respiratory activity

    NASA Astrophysics Data System (ADS)

    Liu, D.; Liu, X.; Liu, Y.; Li, L.; Pan, G.; Crowley, D.; Tippkötter, R.

    2011-02-01

    Biological stabilization within accumulated soil organic carbon (SOC) has not been well understood, while its role in physical and chemical protection as well as of chemical recalcitrance had been addressed in Chinese rice paddies. In this study, topsoil samples were collected and respiratory activity measured in situ following rice harvest under different fertilization treatments of three long-term experimental sites across southern China in 2009. The SOC contents, microbial biomass carbon (SMBC) and nitrogen (SMBN) were analysed using chemical digestion and microbial community structure assessment via clony dilute plate counting methods. While SOC contents were consistently higher under compound chemical fertilization (Comp-Fert) or combined organic and inorganic fertilization (Comb-Fert) compared to N fertilization only (N-Fert), there was significantly higher fungal-bacterial ratio under Comb-Fert than under N-Fert and Comp-Fert. When subtracting the background effect under no fertilization treatment (Non-Fert), the increase both in SMBC and SMBN under fertilization treatment was found very significantly correlated to the increase in SOC over controls across the sites. Also, the ratio of culturable fungal to bacterial population numbers (F/B ratio) was well correlated with soil organic carbon contents in all samples across the sites studied. SOC accumulation favoured a build-up the microbial community with increasing fungal dominance in the rice paddies under fertilization treatments. While soil respiration rates were high under Comb-Fert as a result of enhanced microbial community build-up, the specific soil respiratory activity based on microbial biomass carbon was found in a significantly negatively correlation with the SOC contents for overall samples. Thus, a fungal-dominated microbial community seemed to slow SOC turnover, thereby favouring SOC accumulation under Comp-Fert or under Comb-Fert in the rice paddies. Therefore, the biological stabilization

  8. Cyclooxygenase-2 induction and prostaglandin E2 accumulation in squamous cell carcinoma as a consequence of epidermal growth factor receptor activation by imatinib mesylate.

    PubMed

    Johnson, Faye M; Yang, Peiying; Newman, Robert A; Donato, Nicholas J

    2004-12-01

    Imatinib mesylate is a novel anti-tumor agent useful in the clinical management of chronic myelogenous leukemia and gastrointestinal stromal tumors with minimal toxicity relative to other forms of cancer therapy. Its clinical activity and minimal toxicity are related to specific inhibition of cellular targets including BCR-ABL, platelet-derived growth factor receptor and c-kit kinases, resulting in the collapse of downstream signaling cascades important for transformation. In some patients, unexpected toxicities arise that are not associated with inhibition of any known cellular imatinib target. In this report, we investigated the effects of imatinib on squamous carcinoma cell signaling. Imatinib induced expression of COX-2 in a dose-dependent manner with concomitant accumulation of prostaglandin E2. COX-2 induction by imatinib was initiated through epidermal growth factor (EGF) receptor kinase activation and downstream signaling through mitogenic-activated protein kinase. COX-2 induction by imatinib was blocked by MEK1 or EGF receptor inhibition. Imatinib did not activate stressor cytokine-signaling pathways (p38 kinase, nuclear factor-kB nuclear translocation) or affect COX-1 expression. Imatinib failed to activate EGF receptor signals in other tumor types, suggesting that COX-2 induction in imatinib-treated cells is mediated through release of autocrine factors expressed or activated in squamous tumors. COX-2 induction by imatinib in squamous tumors derived from the head and neck region is unique with respect to other target-specific agents and may represent one of the unintended toxic effects of imatinib described in some patients.

  9. [The accumulation of the technogenic radionuclides by mushrooms in the area affected by the activity of the Krasnoyarsk mining-and-chemical industrial complex].

    PubMed

    Bolsunovskiĭ, A Ia; Dement'ev, D V; Bondareva, L G

    2006-01-01

    The radionuclide content was measured in mushrooms collected in different sites situated in the zone of the radiation influence on the Mining-and-Chemical Combine at Zheleznogorsk and on the control site, near Krasnoyarsk, in 2002-2004. The analytical investigations of fruiting bodies of 12 mushroom species have revealed three gamma-emitting radionuclides: 7Be, 40K (natural) and 137Cs (artificial). It was found that only three species contain 7Be; activity concentration of 40K is not species- and site-dependent, averaging 1600 Bq/kg. All collected samples contain 137Cs, and its accumulation by mushrooms is species-specific. Suillus concentrates more 137Cs activity than other species and can be used as a bioindicator of soil contamination with radiocesium. The average activity concentration of 137Cs in Suillus granulatus collected in the sites subjected only to aerosol discharges of the MCC is more than twice higher than 137Cs content of the mushrooms collected in the control site--"Krasnoyarsk". The maximum activity concentration of 137Cs in Suillus samples collected in the sites that receive 137Cs with the flood water is an order of magnitude higher, amounting to 8624 Bq/kg. The analysis of the radionuclide distribution in a mushroom shows that 40K activity concentration is the same in caps and stems of Suillus, and 137Cs concentration in Suillus caps is 1.7-2.3 times higher than in stems. Binding of radionuclides by mushroom biomass was determined by chemical fractionation; it was found that the highest activities of 137Cs and 40K are in the exchange-adsorption fraction (56 to 71% of the total content of a radionuclide) and in the organic fraction (23 to 37%). Calculations were made for determination of the coefficients of 40K and 137Cs transfer from the soil to the fruiting body of Suillus.

  10. Defense activation triggers differential expression of phospholipase-C (PLC) genes and elevated temperature induces phosphatidic acid (PA) accumulation in tomato

    PubMed Central

    Abd-El-Haliem, Ahmed; Meijer, Harold J.G.; Tameling, Wladimir I.L.; Vossen, Jack H.; Joosten, Matthieu H.A.J.

    2012-01-01

    Recently, we provided the first genetic evidence for the requirement of tomato PLC4 and PLC6 genes in defense activation and disease resistance. The encoded enzymes were catalytically active as they were able to degrade phosphatidylinositol (PI), thereby producing diacylglycerol (DG). Here we report differential PLC gene expression following the initiation of defense signaling by the interaction between Cladosporium fulvum resistance (R) protein Cf-4 and its matching effector Avr4 in tomato hybrid seedlings that express both Cf-4 and Avr4. Furthermore, we observed that PLC3 and PLC6 gene expression is upregulated by elevated temperature in the control seedlings. This upregulation coincides with an increase in the levels of phosphatidic acid (PA) and a decrease in the levels of PI and phosphatidylinositol phosphate (PIP). The decrease in PI and PIP levels matches with the activation of PLC. In addition, the levels of the structural phospholipids phosphatidylcholine (PC), phosphatidylethanolamine (PE) and phosphatidylglycerol (PG) declined transiently during recovery after the exposure to elevated temperature., Further studies will be required to explain the mechanism causing the sustained accumulation of PA during recovery, combined with a reduction in the levels of structural phospholipids. PMID:22899083

  11. Pancreatic adenocarcinoma up-regulated factor (PAUF) enhances the accumulation and functional activity of myeloid-derived suppressor cells (MDSCs) in pancreatic cancer.

    PubMed

    Song, Jinhoi; Lee, Jaemin; Kim, Jinsil; Jo, Seongyea; Kim, Yeon Jeong; Baek, Ji Eun; Kwon, Eun-Soo; Lee, Kwang-Pyo; Yang, Siyoung; Kwon, Ki-Sun; Kim, Dong-Uk; Kang, Tae Heung; Park, Yun-Yong; Chang, Suhwan; Cho, Hee Jun; Kim, Song Cheol; Koh, Sang Seok; Kim, Seokho

    2016-08-09

    Pancreatic cancer is characterized by an immunosuppressive tumor microenvironment (TME) with a profound immune infiltrate populated by a significant number of myeloid-derived suppressor cells (MDSCs). MDSCs have been increasingly recognized for their role in immune evasion and cancer progression as well as their potential as a target for immunotherapy. However, not much is known about the mechanisms regulating their behavior and function in the pancreatic TME. Here we report that pancreatic adenocarcinoma up-regulated factor (PAUF), a soluble protein involved in pancreatic tumorigenesis and metastasis, plays a role as an enhancer of tumor-infiltrating MDSC and its functional activity. We show that PAUF enhanced the accumulation of MDSCs in the spleen and tumor tissues of PAUF-overexpressing tumor cell-injected mice. In addition, PAUF was found to enhance the immunosuppressive function of MDSCs via the TLR4-mediated signaling pathway, which was demonstrated by PAUF-induced increased levels of arginase, nitric oxide (NO), and reactive oxygen species (ROS). The role of PAUF in modulating the functional properties of MDSCs was further demonstrated by the use of a PAUF-neutralizing antibody that caused a decreased number of tumor-infiltrating MDSCs and reduced MDSC immunosuppressive activity. The observations made in mice were confirmed in human pancreatic cancer patient-derived MDSCs, supporting the clinical relevance of our findings. Collectively, we conclude that the PAUF is a powerful and multifunctional promoter of tumor growth through increase and functional activation of MDSCs, suggesting therapeutic potential for targeting PAUF in pancreatic cancers.

  12. Photosynthetic responses, carbohydrate composition and invertase activity in fructan accumulating bryophytes (Porella platyphylla and Sphagnum flexuosum) under different environmental conditions (carbohydrate treatments, dark starvation, low temperature, desiccation).

    PubMed

    Marschall, Marianna

    2010-01-01

    Photosynthetic responses, the non-structural carbohydrate pool and its alterations, and the acid invertase activity under different environmental conditions in the fructan synthesising Porella platyphylla and Sphagnum flexuosum are discussed. Sucrose and fructan are the major soluble carbohydrates in both species. TLC showed that fructans form a homologous series of increasing DP in a similar manner to fructans in Angiosperms and belong to the inulin type. Exogenous sugars (glucose, fructose, sucrose) applied in light and dark resulted in down-regulation of photosynthetic activity, but a long period (1 week) of dark starvation did not cause a significant decrease in the photosynthetic capacity. Light and exogenous sugars increased soluble carbohydrate content due to fructan-accumulation. Dark starvation, desiccation and low temperature did not influence significantly the amount of the total soluble carbohydrates, indicating the existence of a well-buffered carbohydrate pool, although changes in the ratio of fructans of different molecular weight can be detected. Alterations in the activity of acid invertase correlated well with the changes of the amount of the main soluble carbohydrates, showing the role of the enzyme in general carbohydrate and fructan metabolism.

  13. A novel partial agonist of peroxisome proliferator-activated receptor-gamma (PPARgamma) recruits PPARgamma-coactivator-1alpha, prevents triglyceride accumulation, and potentiates insulin signaling in vitro.

    PubMed

    Burgermeister, Elke; Schnoebelen, Astride; Flament, Angele; Benz, Jörg; Stihle, Martine; Gsell, Bernard; Rufer, Arne; Ruf, Armin; Kuhn, Bernd; Märki, Hans Peter; Mizrahi, Jacques; Sebokova, Elena; Niesor, Eric; Meyer, Markus

    2006-04-01

    Partial agonists of peroxisome proliferator-activated receptor-gamma (PPARgamma), also termed selective PPARgamma modulators, are expected to uncouple insulin sensitization from triglyceride (TG) storage in patients with type 2 diabetes mellitus. These agents shall thus avoid adverse effects, such as body weight gain, exerted by full agonists such as thiazolidinediones. In this context, we describe the identification and characterization of the isoquinoline derivative PA-082, a prototype of a novel class of non-thiazolidinedione partial PPARgamma ligands. In a cocrystal with PPARgamma it was bound within the ligand-binding pocket without direct contact to helix 12. The compound displayed partial agonism in biochemical and cell-based transactivation assays and caused preferential recruitment of PPARgamma-coactivator-1alpha (PGC1alpha) to the receptor, a feature shared with other selective PPARgamma modulators. It antagonized rosiglitazone-driven transactivation and TG accumulation during de novo adipogenic differentiation of murine C3H10T1/2 mesenchymal stem cells. The latter effect was mimicked by overexpression of wild-type PGC1alpha but not its LXXLL-deficient mutant. Despite failing to promote TG loading, PA-082 induced mRNAs of genes encoding components of insulin signaling and adipogenic differentiation pathways. It potentiated glucose uptake and inhibited the negative cross-talk of TNFalpha on protein kinase B (AKT) phosphorylation in mature adipocytes and HepG2 human hepatoma cells. PGC1alpha is a key regulator of energy expenditure and down-regulated in diabetics. We thus propose that selective recruitment of PGC1alpha to favorable PPARgamma-target genes provides a possible molecular mechanism whereby partial PPARgamma agonists dissociate TG accumulation from insulin signaling.

  14. T47D Cells Expressing Myeloperoxidase Are Able to Process, Traffic and Store the Mature Protein in Lysosomes: Studies in T47D Cells Reveal a Role for Cys319 in MPO Biosynthesis that Precedes Its Known Role in Inter-Molecular Disulfide Bond Formation

    PubMed Central

    Laura, Richard P.; Dong, David; Reynolds, Wanda F.; Maki, Richard A.

    2016-01-01

    Among the human heme-peroxidase family, myeloperoxidase (MPO) has a unique disulfide-linked oligomeric structure resulting from multi-step processing of the pro-protein monomer (proMPO) after it exits the endoplasmic reticulum (ER). Related family members undergo some, but not all, of the processing steps involved with formation of mature MPO. Lactoperoxidase has its pro-domain proteolytically removed and is a monomer in its mature form. Eosinophil peroxidase undergoes proteolytic removal of its pro-domain followed by proteolytic separation into heavy and light chains and is a heterodimer. However, only MPO undergoes both these proteolytic modifications and then is further oligomerized into a heterotetramer by a single inter-molecular disulfide bond. The details of how and where the post-ER processing steps of MPO occur are incompletely understood. We report here that T47D breast cancer cells stably transfected with an MPO expression plasmid are able to efficiently replicate all of the processing steps that lead to formation of the mature MPO heterotetramer. MPO also traffics to the lysosome granules of T47D cells where it accumulates, allowing in-depth immunofluorescent microscopy studies of MPO trafficking and storage for the first time. Using this novel cell model we show that formation of MPO’s single inter-molecular disulfide bond can occur normally in the absence of the proteolytic events that lead to separation of the MPO heavy and light chains. We further demonstrate that Cys319, which forms MPO’s unique inter-molecular disulfide bond, is important for events that precede this step. Mutation of this residue alters the glycosylation and catalytic activity of MPO and blocks its entry into the endocytic pathway where proteolytic processing and disulfide bonding occur. Finally, using the endocytic trafficking of lysosomal hydrolases as a guide, we investigate the role of candidate receptors in the endocytic trafficking of MPO. PMID:26890638

  15. Kinetics of oxidation of aliphatic and aromatic thiols by myeloperoxidase compounds I and II.

    PubMed

    Burner, U; Jantschko, W; Obinger, C

    1999-01-29

    Myeloperoxidase (MPO) is the most abundant protein in neutrophils and plays a central role in microbial killing and inflammatory tissue damage. Because most of the non-steroidal anti-inflammatory drugs and other drugs contain a thiol group, it is necessary to understand how these substrates are oxidized by MPO. We have performed transient kinetic measurements to study the oxidation of 14 aliphatic and aromatic mono- and dithiols by the MPO intermediates, Compound I (k3) and Compound II (k4), using sequential mixing stopped-flow techniques. The one-electron reduction of Compound I by aromatic thiols (e.g. methimidazole, 2-mercaptopurine and 6-mercaptopurine) varied by less than a factor of seven (between 1.39 +/- 0.12 x 10(5) M(-1) s(-1) and 9.16 +/- 1.63 x 10(5) M(-1) s(-1)), whereas reduction by aliphatic thiols was demonstrated to depend on their overall net charge and hydrophobic character and not on the percentage of thiol deprotonation or redox potential. Cysteamine, cysteine methyl ester, cysteine ethyl ester and alpha-lipoic acid showed k3 values comparable to aromatic thiols, whereas a free carboxy group (e.g. cysteine, N-acetylcysteine, glutathione) diminished k3 dramatically. The one-electron reduction of Compound II was far more constrained by the nature of the substrate. Reduction by methimidazole, 2-mercaptopurine and 6-mercaptopurine showed second-order rate constants (k4) of 1.33 +/- 0.08 x 10(5) M(-1) s(-1), 5.25 +/- 0.07 x 10(5) M(-1) s(-1) and 3.03 +/- 0.07 x 10(3) M(-1) s(-1). Even at high concentrations cysteine, penicillamine and glutathione could not reduce Compound II, whereas cysteamine (4.27 +/- 0.05 x 10(3) M(-1) s(-1)), cysteine methyl ester (8.14 +/- 0.08 x 10(3) M(-1) s(-1)), cysteine ethyl ester (3.76 +/- 0.17 x 10(3) M(-1) s(-1)) and alpha-lipoic acid (4.78 +/- 0.07 x 10(4) M(-1) s(-1)) were demonstrated to reduce Compound II and thus could be expected to be oxidized by MPO without co-substrates.

  16. Altered neuronal mitochondrial coenzyme A synthesis in neurodegeneration with brain iron accumulation caused by abnormal processing, stability, and catalytic activity of mutant pantothenate kinase 2.

    PubMed

    Kotzbauer, Paul T; Truax, Adam C; Trojanowski, John Q; Lee, Virginia M-Y

    2005-01-19

    Mutations in the pantothenate kinase 2 (PANK2) gene have been identified in patients with neurodegeneration with brain iron accumulation (NBIA; formerly Hallervorden-Spatz disease). However, the mechanisms by which these mutations cause neurodegeneration are unclear, especially given the existence of multiple pantothenate kinase genes in humans and multiple PanK2 transcripts with potentially different subcellular localizations. We demonstrate that PanK2 protein is localized to mitochondria of neurons in human brain, distinguishing it from other pantothenate kinases that do not possess mitochondrial-targeting sequences. PanK2 protein translated from the most 5' start site is sequentially cleaved at two sites by the mitochondrial processing peptidase, generating a long-lived 48 kDa mature protein identical to that found in human brain extracts. The mature protein catalyzes the initial step in coenzyme A (CoA) synthesis but displays feedback inhibition in response to species of acyl CoA rather than CoA itself. Some, but not all disease-associated point mutations result in significantly reduced catalytic activity. The most common mutation, G521R, results in marked instability of the intermediate PanK2 isoform and reduced production of the mature isoform. These results suggest that NBIA is caused by altered neuronal mitochondrial lipid metabolism caused by mutations disrupting PanK2 protein levels and catalytic activity.

  17. Effects of Lantana camara leaf extract on the activity of superoxide dismutase and accumulation of H2O2 in water hyacinth leaf.

    PubMed

    Zheng, Hui-Qiong; Wei, Ning; Wang, Liu-Fa; He, Ping

    2006-04-01

    Water hyacinth (Eichhornia crassipes) is one of the most productive plants, but is also a troublesome weed in the world. In order to protect the public water system from chemical herbicides pollution, biological method has been suggested to control the growth and the reproduction of this weed. Lantana (Lantana camara L.) is an important weed of the family Verbenaceae and its leaf extract is highly toxic to water hyacinth. The results of this study showed that the extract of lantana leaves suppressed the emergence of leaf buds of water hyacinth plant, and caused the decay of its leaves by foliar spraying. In addition, the increase of SOD activity in water hyacinth leaves was in accordance with the accumulation of H(2)O(2) and the increase in degree of membrane peroxidation, while the activity of catalase, which might remove the excessive H(2)O(2) in water hyacinth leaves, was inhibited by treatment with lantana extract. At tissue level, high H(2)O(2) histochemical labeling was detected in guard cells after treatment with lantana extract. This overproduction of H(2)O(2) could kill the leaf cells and cause leaf necrosis in the treated plant. Therefore, the high toxicity of lantana leaf extract to water hyacinth might be due to oxidative stress.

  18. Autoantibody production in lpr/lpr gld/gld mice reflects accumulation of CD4+ effector cells that are resistant to regulatory T cell activity.

    PubMed

    Hondowicz, Brian D; Fields, Michele L; Nish, Simone A; Larkin, Joseph; Caton, Andrew J; Erikson, Jan

    2008-09-01

    In Fas/FasL-deficient mice anti-chromatin Ab production is T cell dependent and is not apparent until after 10 weeks of age. Early control of anti-chromatin antibodies may be due to the counterbalancing influence of Treg cells. Here we show that Treg cells block lpr/lpr gld/gld Th cells from providing help to anti-chromatin B cells in an in vivo transfer system. Interestingly, the percentage and absolute numbers of Foxp3+ Treg cells is elevated in BALB/c-lpr/lpr gld/gld mice and increases with age compared to BALB/c mice. The majority of Foxp3 expression is found in the B220- CD4+ T cell population, and Foxp3-expressing cells are localized in the splenic PALS (periarteriolar lymphocyte sheath). Strikingly, although the lack of functional Fas/FasL does not affect the ability of Treg cells to block Th cell proliferation, Treg cells can block the IFN-gamma differentiation of Th cells from BALB/c or young BALB-lpr/lpr gld/gld mice but not of pre-existing Th1 cells from older BALB/c-lpr/lpr gld/gld mice. Thus, we suggest autoantibody production is not caused by the lack of Treg cells but by a defect in activation-induced cell death that leads to the accumulation of T effector cells that are resistant to regulatory T cell activity.

  19. Bidens pilosa and its active compound inhibit adipogenesis and lipid accumulation via down-modulation of the C/EBP and PPARγ pathways

    PubMed Central

    Liang, Yu-Chuan; Yang, Meng-Ting; Lin, Chuan-Ju; Chang, Cicero Lee-Tian; Yang, Wen-Chin

    2016-01-01

    Obesity and its complications are a major global health problem. In this study, we investigated the anti-obesity effect and mechanism of an edible plant, Bidens pilosa, and its active constituent. We first assessed the long-term effect of B. pilosa on body composition, body weight, blood parameters in ICR mice. We observed that it significantly decreased fat content and increased protein content in ICR mice. Next, we verified the anti-obesity effect of B. pilosa in ob/ob mice. It effectively and dose-dependently reduced fat content, adipocyte size and/or body weight in mice. Moreover, mechanistic studies showed that B. pilosa inhibited the expression of peroxisome proliferator activated receptor γ (PPARγ), CCAAT/enhancer binding proteins (C/EBPs) and Egr2 in adipose tissue. Finally, we examined the effect of 2-β-D-glucopyranosyloxy-1-hydroxytrideca-5,7,9,11-tetrayne (GHT) on adipogenesis in adipocytes. We found that B. pilosa significantly decreased the adipogenesis and lipid accumulation. This decrease was associated with the down-regulation of expression of Egr2, C/EBPs, PPARγ, adipocyte Protein 2 (aP2) and adiponectin. In summary, this work demonstrated that B. pilosa and GHT suppressed adipogenesis and lipid content in adipocytes and/or animals via the down-regulation of the Egr2, C/EBPs and PPARγ pathways, suggesting a novel application of B. pilosa and GHT against obesity. PMID:27063434

  20. Expression of a constitutively active nitrate reductase variant in tobacco reduces tobacco-specific nitrosamine accumulation in cured leaves and cigarette smoke.

    PubMed

    Lu, Jianli; Zhang, Leichen; Lewis, Ramsey S; Bovet, Lucien; Goepfert, Simon; Jack, Anne M; Crutchfield, James D; Ji, Huihua; Dewey, Ralph E

    2016-07-01

    Burley tobaccos (Nicotiana tabacum) display a nitrogen-use-deficiency phenotype that is associated with the accumulation of high levels of nitrate within the leaf, a trait correlated with production of a class of compounds referred to as tobacco-specific nitrosamines (TSNAs). Two TSNA species, 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK) and N-nitrosonornicotine (NNN), have been shown to be strong carcinogens in numerous animal studies. We investigated the potential of molecular genetic strategies to lower nitrate levels in burley tobaccos by overexpressing genes encoding key enzymes of the nitrogen-assimilation pathway. Of the various constructs tested, only the expression of a constitutively active nitrate reductase (NR) dramatically decreased free nitrate levels in the leaves. Field-grown tobacco plants expressing this NR variant exhibited greatly reduced levels of TSNAs in both cured leaves and mainstream smoke of cigarettes made from these materials. Decreasing leaf nitrate levels via expression of a constitutively active NR enzyme represents an exceptionally promising means for reducing the production of NNN and NNK, two of the most well-documented animal carcinogens found in tobacco products.

  1. Perennial peanut (Arachis glabrata Benth.) leaves contain hydroxycinnamoyl-CoA:tartaric acid hydroxycinnamoyl transferase activity and accumulate hydroxycinnamoyl-tartaric acid esters.

    PubMed

    Sullivan, Michael L

    2014-05-01

    Many plants accumulate hydroxycinnamoyl esters to protect against abiotic and biotic stresses. Caffeoyl esters in particular can be substrates for endogenous polyphenol oxidases (PPOs). Recently, we showed that perennial peanut (Arachis glabrata Benth.) leaves contain PPO and identified one PPO substrate, caftaric acid (trans-caffeoyl-tartaric acid). Additional compounds were believed to be cis- and trans-p-coumaroyl tartaric acid and cis- and trans-feruloyl-tartaric acid, but lack of standards prevented definitive identifications. Here we characterize enzymatic activities in peanut leaves to understand how caftaric acid and related hydroxycinnamoyl esters are made in this species. We show that peanut leaves contain a hydroxycinnamoyl-CoA:tartaric acid hydroxycinnamoyl transferase (HTT) activity capable of transferring p-coumaroyl, caffeoyl, and feruloyl moieties from CoA to tartaric acid (specific activities of 11 ± 2.8, 8 ± 1.8, 4 ± 0.8 pkat mg(-1) crude protein, respectively). The HTT activity was used to make cis- and trans-p-coumaroyl- and -feruloyl-tartaric acid in vitro. These products allowed definitive identification of the corresponding cis- and trans-hydroxycinnamoyl esters extracted from leaves. We tentatively identified sinapoyl-tartaric acid as another major phenolic compound in peanut leaves that likely participates in secondary reactions with PPO-generated quinones. These results suggest hydroxycinnamoyl-tartaric acid esters are made by an acyltransferase, possibly a BAHD family member, in perennial peanut. Identification of a gene encoding HTT and further characterization of the enzyme will aid in identifying determinants of donor and acceptor substrate specificity for this important class of biosynthetic enzymes. An HTT gene could also provide a means by genetic engineering for producing caffeoyl- and other hydroxycinnamoyl-tartaric acid esters in forage crops that lack them.

  2. Clozapine-induced agranulocytosis and hereditary polymorphisms of clozapine metabolizing enzymes: no association with myeloperoxidase and cytochrome P4502D6.

    PubMed

    Dettling, M; Sachse, C; Müller-Oerlinghausen, B; Roots, I; Brockmöller, J; Rolfs, A; Cascorbi, I

    2000-11-01

    The pathomechanisms of most drug-induced agranulocytoses are unclear; however, there are some studies pointing to genetic determinants. Some drug-induced agranulocytoses such as clozapine-induced agranulocytosis (CA) may be regarded as an idiosyncratic drug reaction because of its preclinical and clinical characteristics. To study some aspects of the genetic background of CA further, polymorphisms of specific metabolizing enzyme systems of clozapine were examined. Thirty-one schizophrenic patients with CA and 77 schizophrenic comparison subjects without this adverse effect underwent genotyping of a recently discovered G(-463)A polymorphism of myeloperoxidase (MPO) gene and cytochrome P4502D6. Neither the MPO mutation nor specific genotypes of cytochrome P4502D6 were associated with CA. Both were equally distributed among CA patients and controls. Thus, our data suggest lack of evidence of an association of CA and genetically variable activity of these specific drug metabolizing enzymes; however, this may be due to statistical reasons only. Thus, further studies with greater CA samples are necessary to draw final conclusions about these genetically based hypotheses.

  3. Pancreatic adenocarcinoma up-regulated factor (PAUF) enhances the accumulation and functional activity of myeloid-derived suppressor cells (MDSCs) in pancreatic cancer

    PubMed Central

    Jo, Seongyea; Kim, Yeon Jeong; Baek, Ji Eun; Kwon, Eun-Soo; Lee, Kwang-Pyo; Yang, Siyoung; Kwon, Ki-Sun; Kim, Dong-Uk; Kang, Tae Heung; Park, Yun-Yong; Chang, Suhwan; Cho, Hee Jun; Kim, Song Cheol; Koh, Sang Seok; Kim, Seokho

    2016-01-01

    Pancreatic cancer is characterized by an immunosuppressive tumor microenvironment (TME) with a profound immune infiltrate populated by a significant number of myeloid-derived suppressor cells (MDSCs). MDSCs have been increasingly recognized for their role in immune evasion and cancer progression as well as their potential as a target for immunotherapy. However, not much is known about the mechanisms regulating their behavior and function in the pancreatic TME. Here we report that pancreatic adenocarcinoma up-regulated factor (PAUF), a soluble protein involved in pancreatic tumorigenesis and metastasis, plays a role as an enhancer of tumor-infiltrating MDSC and its functional activity. We show that PAUF enhanced the accumulation of MDSCs in the spleen and tumor tissues of PAUF-overexpressing tumor cell-injected mice. In addition, PAUF was found to enhance the immunosuppressive function of MDSCs via the TLR4-mediated signaling pathway, which was demonstrated by PAUF-induced increased levels of arginase, nitric oxide (NO), and reactive oxygen species (ROS). The role of PAUF in modulating the functional properties of MDSCs was further demonstrated by the use of a PAUF-neutralizing antibody that caused a decreased number of tumor-infiltrating MDSCs and reduced MDSC immunosuppressive activity. The observations made in mice were confirmed in human pancreatic cancer patient-derived MDSCs, supporting the clinical relevance of our findings. Collectively, we conclude that the PAUF is a powerful and multifunctional promoter of tumor growth through increase and functional activation of MDSCs, suggesting therapeutic potential for targeting PAUF in pancreatic cancers. PMID:27322081

  4. Effects of Decreased Vitamin D and Accumulated Uremic Toxin on Human CYP3A4 Activity in Patients with End-Stage Renal Disease

    PubMed Central

    Tsujimoto, Masayuki; Nagano, Yui; Hosoda, Satomi; Shiraishi, Asuka; Miyoshi, Ayaka; Hiraoka, Shima; Furukubo, Taku; Izumi, Satoshi; Yamakawa, Tomoyuki; Minegaki, Tetsuya; Nishiguchi, Kohshi

    2013-01-01

    In patients with end-stage renal disease, not only renal clearance but also hepatic clearance is known to be impaired. For instance, the concentration of erythromycin, a substrate of cytochrome P450 3A4 (CYP3A4), has been reported to be elevated in patients with end-stage renal disease. The purpose of this study is to elucidate the reason for the decrease in hepatic clearance in patients with end-stage renal disease. Deproteinized pooled sera were used to assess the effects of low-molecular-weight uremic toxins on CYP3A4 activity in human liver microsomes and human LS180 cells. Four uremic toxins (3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid, hippuric acid, indole-3-acetic acid, and 3-indoxyl sulfate) present at high concentrations in uremic serum were also studied. Simultaneous treatment of uremic serum (less than 10%) or uremic toxins did not affect testosterone 6β-hydroxylation in human liver microsomes. On the other hand, pretreatment of each serum activates CYP3A4 in LS180 cells, and the increased CYP3A4 activity in uremic serum-treated cells was smaller than normal serum-treated cells. In addition, CYP3A4 and CYP24A1 mRNA levels also increased in LS180 cells exposed to normal serum, and this effect was reduced in uremic serum-treated cells and in cells exposed to uremic serum added to normal serum. Furthermore, addition of 1,25-dihydroxyvitamin D to uremic serum partially restored the serum effect on CYP3A4 expression. The present study suggests that the decrease of 1,25-dihydroxyvitamin D and the accumulation of uremic toxins contributed to the decreased hepatic clearance of CYP3A4 substrates in patients with end-stage renal disease. PMID:23965431

  5. The fruit of Acanthopanax senticosus (Rupr. et Maxim.) Harms improves insulin resistance and hepatic lipid accumulation by modulation of liver adenosine monophosphate-activated protein kinase activity and lipogenic gene expression in high-fat diet-fed obese mice.

    PubMed

    Saito, Tetsuo; Nishida, Miyako; Saito, Masafumi; Tanabe, Akari; Eitsuka, Takahiro; Yuan, Shi-Hua; Ikekawa, Nobuo; Nishida, Hiroshi

    2016-10-01

    Obesity-associated insulin resistance is a major risk factor for most metabolic diseases, including dyslipidemia and type 2 diabetes. Acanthopanax senticosus (Rupr. et Maxim.) Harms (Goka) root has been used in traditional Chinese medicine for treatment of diabetes and other conditions; however, little is known about the effects of Goka fruit (GF). Goka fruit is rich in anthocyanin, which has beneficial effects on obesity and insulin resistance via activation of adenosine monophosphate-activated protein kinase (AMPK). We hypothesized that GF can improve obesity-associated insulin resistance. The aim of the present study was to investigate whether GF improves insulin resistance in high-fat diet (HFD)-induced obese mice. High-fat diet mice treated with GF (500 and 1000 mg/kg) for 12 weeks showed an improved glucose tolerance and insulin sensitivity, as well as reduced plasma insulin and liver lipid accumulation. Moreover, GF administration to HFD mice resulted in down-regulation of fatty acid synthase expression and up-regulation of cholesterol 7-alpha-hydroxylase expression in the liver. Notably, AMPK phosphorylation in the liver increased after GF administration. In summary, GF supplementation improved obesity-associated insulin resistance and hepatic lipid accumulation through modulation of AMPK activity and lipid metabolism-associated gene expression.

  6. Heme-linked ionizations of myeloperoxidase detected by Raman difference spectroscopy. A comparison with plant and yeast peroxidases.

    PubMed Central

    Stump, R F; Deanin, G G; Oliver, J M; Shelnutt, J A

    1987-01-01

    The pH-dependence of the oxidation state marker line v4 of human leucocyte myeloperoxidase is determined in the absence of chloride using Raman difference spectroscopy (RDS). A transition in the frequency of v4 with pK of 4.2 +/- 0.3 is found. The pK compares favorably with that previously determined by spectrophotometric titration and kinetic studies. The shift in v4 across the transition is -1.3 cm-1. The shift in v4 and other Raman marker lines indicates enhanced pi charge in the chlorin ring below the transition. The low frequencies of the oxidation state marker lines indicate that a structural change occurs near the chromophore, which results in the formation of a more pi-charge donating protein environment for the chlorin ring at low pH. The Raman results are discussed in terms of a proposed catalytic control mechanism based on charge stabilization of the energy of ring charge-depleted ferryl intermediates of the reaction with peroxide. The myeloperoxidase findings are compared with similar RDS results for ferrous horseradish peroxidase and ferric cytochrome c peroxidase. PMID:3034344

  7. Proteomic profile of aminoglutethimide-induced apoptosis in HL-60 cells: Role of myeloperoxidase and arylamine free radicals.

    PubMed

    Khan, Saifur R; Baghdasarian, Argishti; Nagar, Prarthna H; Fahlman, Richard; Jurasz, Paul; Michail, Karim; Aljuhani, Naif; Siraki, Arno G

    2015-09-05

    In this study, the cellular effects resulting from the metabolism of aminoglutethimide by myeloperoxidase were investigated. Human promyelocytic leukemia (HL-60) cells were treated with aminoglutethimide (AG), an arylamine drug that has a risk of adverse drug reactions, including drug-induced agranulocytosis. HL-60 cells contain abundant amounts of myeloperoxidase (MPO), a hemoprotein, which catalyzes one-electron oxidation of arylamines using H2O2 as a cofactor. Previous studies have shown that arylamine metabolism by MPO results in protein radical formation. The purpose of this study was to determine if pathways associated with a toxic response could be determined from conditions that produced protein radicals. Conditions for AG-induced protein radical formation (with minimal cytotoxicity) were optimized, and these conditions were used to carry out proteomic studies. We identified 43 proteins that were changed significantly upon AG treatment among which 18 were up-regulated and 25 were down-regulated. The quantitative proteomic data showed that AG peroxidative metabolism led to the down-regulation of critical anti-apoptotic proteins responsible for inhibiting the release of pro-apoptotic factors from the mitochondria as well as cytoskeletal proteins such as nuclear lamina. This overall pro-apoptotic response was confirmed with flow cytometry which demonstrated apoptosis to be the main mode of cell death, and this was attenuated by MPO inhibition. This response correlated with the intensity of AG-induced protein radical formation in HL-60 cells, which may play a role in cell death signaling mechanisms.

  8. Involvement of myeloperoxidase and NADPH oxidase in the covalent binding of amodiaquine and clozapine to neutrophils: implications for drug-induced agranulocytosis.

    PubMed

    Lobach, Alexandra R; Uetrecht, Jack

    2014-04-21

    Amodiaquine (AQ) and clozapine (CLZ) are associated with a relatively high incidence of idiosyncratic agranulocytosis, a reaction that is suspected to involve covalent binding of reactive metabolites to neutrophils. Previous studies have shown that both AQ and CLZ are oxidized to reactive intermediates in vitro by activated neutrophils or by the combination of hydrogen peroxide and myeloperoxidase (MPO). Neutrophil activation leads to an oxidative burst with activation of NADPH oxidase and the production of hydrogen peroxide. However, the importance of this pathway in covalent binding in vivo has not been examined. In this study, we found that the binding of both AQ and CLZ to neutrophils from MPO knockout mice ex vivo decreased approximately 2-fold compared to neutrophils from wild-type mice, whereas binding to activated neutrophils from gp91 knockout (NADPH oxidase null) mice decreased 6-7-fold. When the AQ studies were performed in vivo, again the binding was decreased in MPO knockout mice to about 50% of the binding in wild-type mice; however, covalent binding was significant in the absence of MPO. Surprisingly, there was no significant decrease in covalent binding of AQ to neutrophils in vivo in gp91 knockout mice. In addition, there was extensive binding of AQ to many types of bone marrow cells and to peripheral lymphocytes. These results indicate that MPO is not the only neutrophil enzyme involved in the oxidation of AQ and that NADPH oxidase is not the major source of peroxide. There was also no decrease in AQ binding to neutrophils in COX-1 or COX-2 knockout mice. We were not able to readily reproduce the AQ in vivo studies with CLZ because of its acute toxicity in mice. These are the first studies to examine the enzymes involved in the bioactivation of AQ by neutrophils in vivo.

  9. Flavonol glycosides with lipid accumulation inhibitory activity and simultaneous quantitative analysis of 15 polyphenols and caffeine in the flower buds of Camellia sinensis from different regions by LCMS.

    PubMed

    Morikawa, Toshio; Ninomiya, Kiyofumi; Miyake, Sohachiro; Miki, Yoshinobu; Okamoto, Masaki; Yoshikawa, Masayuki; Muraoka, Osamu

    2013-09-01

    A simultaneous quantitative analytical method for 15 major polyphenols, e.g. five catechins (1-5) and 10 flavonols (6-15), as functional constituents in the extracts of "tea flowers", the flower buds of Camellia sinensis (Theaceae), has been developed. The content of caffeine (16), which showed similar chromatographic behaviour under the analytical conditions, was also determined. To approve the validity of the newly developed protocol, thirteen extracts of the plant's flower buds collected from different regions, i.e. China, Taiwan, Japan and India, were evaluated. The results indicated that the assay was reproducible and precise, and could be readily underutilised for the quality evaluation of tea flowers on the basis of polyphenols' contents. It was noteworthy that the contents of two major constituents, kaempferol 3-O-β-D-glucopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→6)-β-D-glucopyranoside (10) and kaempferol 3-O-β-D-glucopyranosyl-(1→3)-α-L-rhamnopyranosyl-(1→6)-β-D-galactopyranoside (11), varied by region where the flower buds were produced. A new flavonol glycoside, chakaflavonoside B (17), which was isolated in the course of this analytical study, was found to show oleic acid-albumin-induced lipid accumulation inhibitory activity.

  10. Effect of solids retention time and temperature on waste activated sludge hydrolysis and short-chain fatty acids accumulation under alkaline conditions in continuous-flow reactors.

    PubMed

    Feng, Leiyu; Wang, Hua; Chen, Yinguang; Wang, Qin

    2009-01-01

    The effects of solids retention time (SRT) and temperature on waste activated sludge (WAS) hydrolysis and short-chain fatty acids (SCFAs) accumulation were investigated in a series of continuous-flow reactors at pH 10. The experimental results showed that the increase of either SRT or temperature benefited the hydrolysis of WAS and the production of SCFAs. The changes in SRT gave also impact on the percentage of acetic and propionic acids in the fermentative SCFAs, but little influence on that of the slightly long-chain SCFAs, such as n-butyric, iso-butyric, n-valeric and iso-valeric acids. Compared with the control (pH unadjusted) experiment, at SRT of 12d and temperature of 20 degrees C the concentration of SCFAs produced at pH 10 increased from 261.2 to 933.5mg COD/L, and the propionic acid percentage improved from 11.7 to 16.0%. It can be concluded from this investigation that the efficient continuous production of SCFAs at pH 10 is feasible.

  11. SirT1 knockdown potentiates radiation-induced bystander effect through promoting c-Myc activity and thus facilitating ROS accumulation.

    PubMed

    Xie, Yuexia; Tu, Wenzhi; Zhang, Jianghong; He, Mingyuan; Ye, Shuang; Dong, Chen; Shao, Chunlin

    2015-02-01

    Radiation-induced bystander effect (RIBE) has important implications for secondary cancer risk assessment during cancer radiotherapy, but the bystander signaling processes, especially under hypoxic condition, are still largely unclear. The present study found that micronuclei (MN) formation could be induced in the non-irradiated HL-7702 hepatocyte cells after being treated with the conditioned medium from irradiated hepatoma HepG2 and SK-Hep-1 cells under either normoxia or hypoxia. This bystander response was dramatically diminished or enhanced when the SirT1 gene of irradiated hepatoma cells was overexpressed or knocked down, respectively, especially under hypoxia. Meanwhile, SirT1 knockdown promoted transcriptional activity for c-Myc and facilitated ROS accumulation. But both of the increased bystander responses and ROS generation due to SirT1-knockdown were almost completely suppressed by c-Myc interference. Moreover, ROS scavenger effectively abolished the RIBE triggered by irradiated hepatoma cells even with SirT1 depletion. These findings provide new insights that SirT1 has a profound role in regulating RIBE where a c-Myc-dependent release of ROS may be involved.

  12. Activation of GPR55 Receptors Exacerbates oxLDL-Induced Lipid Accumulation and Inflammatory Responses, while Reducing Cholesterol Efflux from Human Macrophages

    PubMed Central

    Lanuti, Mirko; Talamonti, Emanuela; Maccarrone, Mauro; Chiurchiù, Valerio

    2015-01-01

    The G protein-coupled receptor GPR55 has been proposed as a new cannabinoid receptor associated with bone remodelling, nervous system excitability, vascular homeostasis as well as in several pathophysiological conditions including obesity and cancer. However, its physiological role and underlying mechanism remain unclear. In the present work, we demonstrate for the first time its presence in human macrophages and its increased expression in ox-LDL-induced foam cells. In addition, pharmacological activation of GPR55 by its selective agonist O-1602 increased CD36- and SRB-I-mediated lipid accumulation and blocked cholesterol efflux by downregulating ATP-binding cassette (ABC) transporters ABCA1 and ABCG1, as well as enhanced cytokine- and pro-metalloprotease-9 (pro-MMP-9)-induced proinflammatory responses in foam cells. Treatment with cannabidiol, a selective antagonist of GPR55, counteracted these pro-atherogenic and proinflammatory O-1602-mediated effects. Our data suggest that GPR55 could play deleterious role in ox-LDL-induced foam cells and could be a novel pharmacological target to manage atherosclerosis and other related cardiovascular diseases. PMID:25970609

  13. Intracellular chloride activities in canine tracheal epithelium. Direct evidence for sodium-coupled intracellular chloride accumulation in a chloride-secreting epithelium.

    PubMed Central

    Welsh, M J

    1983-01-01

    Canine tracheal epithelium secretes Cl via an electrogenic transport process that appears to apply to a wide variety of secretory epithelia. To examine the mechanisms involved, intracellular chloride activity, acCl, was measured with Cl-selective intracellular microelectrodes. The results indicate that when the rate of secretion was minimal acCl was 37 mM; with stimulation of secretion the intracellular voltage depolarized, but acCl was not significantly altered, at 39 mM. These findings indicate that: (a) Cl is accumulated across the basolateral membrane under nonsecreting and secreting conditions at an activity 3.8 and 2.4 times, respectively, that predicted for an equilibrium distribution; (b) Cl exit across the apical membrane may be passive with an electrochemical driving force of 22 mV; and (c) stimulation of secretion enhanced the rate of Cl entry across the basolateral membrane, since Cl transport increased without a change in acCl. In the absence of Na in the extracellular fluid, acCl approached the value expected for an equilibrium distribution. This finding suggests that "uphill" entry of Cl into the cell against its electrochemical gradient is dependent upon, and energized by, the entry of Na down its gradient. Submucosal bumetanide, a loop diuretic, also decreased the rate of Cl secretion and decreased acCl, indicating an inhibition of Cl entry. These findings indicate that Cl entry into the cell is directed against its electrochemical gradient and is mediated by a Na-coupled, bumetanide-inhibitable, transport process at the basolateral membrane and that Cl may exit passively down a favorable electrochemical gradient across the apical membrane. PMID:6853719

  14. Changes in growth, carbon and nitrogen enzyme activity and mRNA accumulation in the halophilic microalga Dunaliella viridis in response to NaCl stress

    NASA Astrophysics Data System (ADS)

    Wang, Dongmei; Wang, Weiwei; Xu, Nianjun; Sun, Xue

    2016-12-01

    Many species of microalga Dunaliella exhibit a remarkable tolerance to salinity and are therefore ideal for probing the effects of salinity. In this work, we assessed the effects of NaCl stress on the growth, activity and mRNA level of carbon and nitrogen metabolism enzymes of D. viridis. The alga could grow over a salinity range of 0.44 mol L-1 to 3.00 mol L-1 NaCl, but the most rapid growth was observed at 1.00 mol L-1 NaCl, followed by 2.00 mol L-1 NaCl. Paralleling these growth patterns, the highest initial and total Rubisco activities were detected in the presence of 1.00 mol L-1 NaCl, decreasing to 37.33% and 26.39% of those values, respectively, in the presence of 3.00 mol L-1 NaCl, respectively. However, the highest extracellular carbonic anhydrase (CA) activity was measured in the presence of 2.00 mol L-1 NaCl, followed by 1.00 mol L-1 NaCl. Different from the two carbon enzymes, nitrate reductase (NR) activity showed a slight change under different NaCl concentrations. At the transcriptional level, the mRNAs of Rubisco large subunit ( rbcL), and small subunit ( rbcS), attained their highest abundances in the presence of 1.00 and 2.00 mol L-1 NaCl, respectively. The CA mRNA accumulation was induced from 0.44 mol L-1 to 3.00 mol L-1 NaCl, but the NR mRNA showed the decreasing tendency with the increasing salinity. In conclusion, the growth and carbon fixation enzyme of Rubisco displayed similar tendency in response to NaCl stress, CA was proved be salt-inducible within a certain salinity range and NR showed the least effect by NaCl in D. viridis.

  15. Psoriasis and cardiovascular risk factors: increased serum myeloperoxidase and corresponding immunocellular overexpression by Cd11b+ CD68+ macrophages in skin lesions

    PubMed Central

    Cao, Lauren Y; Soler, David C; Debanne, Sarah M; Grozdev, Ivan; Rodriguez, Myriam E; Feig, Rivka L; Carman, Teresa L; Gilkeson, Robert C; Orringer, Carl E; Kern, Elizabeth F; McCormick, Thomas S; Cooper, Kevin D; Korman, Neil J

    2014-01-01

    Background: Recent studies report independent associations between psoriasis, cardiovascular (CV) events and risk factors. Blood Myeloperoxidase (MPO) from activated myeloid cells is associated with CV risk mainly through lipid oxidation, induction of endothelial dysfunction and release of IL-12 from macrophages. Objectives: To elucidate associations between psoriasis and conventional CV risk factors. Methods: We performed a cross-sectional study of 100 psoriasis patients and 53 controls, group matched on age, gender and body mass index, to assess levels of MPO in serum, as well as immunohistochemical staining from psoriasis skin lesions, psoriasis uninvolved skin, and normal skin. Results: Although the groups did not differ on waist circumference, glucose, cholesterol, triglycerides, creatinine or personal history of CV events, psoriasis patients had significantly higher waist-to-hip ratios, blood pressures, proportion of current smokers, and lower high density lipoprotein level than controls. Serum MPO level was elevated 2.5 fold (P<0.001) in psoriasis patients, even after adjusting for the CV risk factors on which the groups differed. MPO did correlate with coronary artery calcification, carotid plaque, carotid intima media thickness and flow mediated dilation, but did not correlate with psoriasis severity. However, MPO was highly expressed in lesional psoriatic skin and colocalized predominantly with CD45+ CD11b+ leukocytes. CD11b+ cell density correlated with circulation MPO levels. Conclusion: Lesional skin CD11b+ leukocytes activated to generate MPO may contribute to serum levels of MPO. Lesional CD11b+ cell activity may be an alternative measure of disease burden to PASI that underlies the MPO biomarker for systemic inflammation related to Cardiovascular Disease. PMID:24349618

  16. Organic UV filters inhibit multixenobiotic resistance (MXR) activity in Tetrahymena thermophila: investigations by the Rhodamine 123 accumulation assay and molecular docking.

    PubMed

    Gao, Li; Yuan, Tao; Cheng, Peng; Zhou, Chuanqi; Ao, Junjie; Wang, Wenhua; Zhang, Haimou

    2016-09-01

    Multixenobiotic resistance (MXR) transporters, which belong to ATP-binding cassette (ABC) family proteins, are present in living organisms as a first line of defense system against xenobiotics and environmental contaminants. The effects of six organic UV filters (4-methyl -benzylidene camphor, 4-MBC; benzophenone-3, BP-3; butyl methoxydibenzoyl-methane, BM-DBM; ethylhexyl methoxy cinnamate, EHMC; octocrylene, OC and homosalate, HMS) on multixenobiotic resistance (MXR) in Tetrahymena thermophila were investigated in this study. It was found that 4-MBC, BP-3 and BM-DBM could significantly inhibit activity of the MXR system, causing concentration dependent accumulation of rhodamine 123; while EHMC, OC and HMS had weak MXR inhibition. The IC50 (50 % inhibition concentration) values of 4-MBC, BP-3 and BM-DBM were 23.54, 40.59 and 26.37 μM, respectively, with inhibitory potentials of 23.1, 13.4 and 20.6 % relative to verapamil (VER, a model inhibitor of P-glycoprotein). Our results firstly provide the evidence for UV filters inhibition effect on MXR in aquatic organisms. In addition, it was revealed by molecular docking analysis that the selected six UV filters can occupy the same binding site on T. thermophila P-gp as VER does; and form H-bonds with residues Ser 328 and/or Asn 281. This study raises the awareness of aquatic ecological risk from the organic UV filters exposure, as they would be involved in potentiating toxic effects by chemosensitizing.

  17. Real-time monitoring of superoxide accumulation and antioxidant activity in a brain slice model using an electrochemical cytochrome c biosensor

    PubMed Central

    Ganesana, Mallikarjunarao; Erlichman, Joseph S.; Andreescu, Silvana

    2012-01-01

    The overproduction of reactive oxygen species and resulting damage are central to the pathology of many diseases. The study of the temporal and spatial accumulation of reactive oxygen species has been limited due to the lack of specific probes and techniques capable of continuous measurement. We demonstrate the use of a miniaturized electrochemical cytochrome C (Cyt C) biosensor for real-time measurements and quantitative assessment of superoxide production and inactivation by natural and engineered antioxidants in acutely prepared brain slices from mice. During control conditions, superoxide radicals produced from the hippocampal region of the brain in 400 μm thick sections were well within the range of detection of the electrode. Exposure of the slices to ischemic conditions increased the superoxide production two fold and measurements from the slices were stable over a 3–4 hour period. The stilbene derivative and anion channel inhibitor, 4,4′-diisothiocyano-2,2′-disulfonic stilbene (DIDS), markedly reduced the extracellular superoxide signal under control conditions suggesting that a transmembrane flux of superoxide into the extracellular space may occur as part of normal redox signaling. The specificity of the electrode for superoxide released by cells in the hippocampus was verified by the exogenous addition of superoxide dismutase (SOD) which decreased the superoxide signal in a dose-dependent manner. Similar results were seen with the addition of the SOD-mimetic, cerium oxide nanoparticles (nanoceria) where the superoxide anion radical scavenging activity of nanoceria with an average diameter of 15 nm was equivalent to 527 U of SOD for each 1 μg/ml of nanoceria added. This study demonstrates the potential of electrochemical biosensors for studying real-time dynamics of reactive oxygen species in a biological model and the utility of these measurements in defining the relative contribution of superoxide to oxidative injury. PMID:23085519

  18. Downregulation of a putative plastid PDC E1α subunit impairs photosynthetic activity and triacylglycerol accumulation in nitrogen-starved photoautotrophic Chlamydomonas reinhardtii

    PubMed Central

    Shtaida, Nastassia; Khozin-Goldberg, Inna; Solovchenko, Alexei; Chekanov, Konstantin; Didi-Cohen, Shoshana; Leu, Stefan; Cohen, Zvi; Boussiba, Sammy

    2014-01-01

    The chloroplast pyruvate dehydrogenase complex (cpPDC) catalyses the oxidative decarboxylation of pyruvate forming acetyl-CoA, an immediate primer for the initial reactions of de novo fatty acid (FA) synthesis. Little is known about the source of acetyl-CoA in the chloroplasts of photosynthetic microalgae, which are capable of producing high amounts of the storage lipid triacylglycerol (TAG) under conditions of nutrient stresses. We generated Chlamydomonas reinhardtii CC-1618 mutants with decreased expression of the PDC2_E1α gene, encoding the putative chloroplast pyruvate dehydrogenase subunit E1α, using artificial microRNA. A comparative study on the effects of PDC2_E1α silencing on FAs and TAG production in C. reinhardtii, grown photoautotrophically and mixotrophically, with and without a nitrogen source in the nutrient medium, was carried out. Reduced expression of PDC2 _E1α led to a severely hampered photoautotrophic growth phenotype with drastic impairment in TAG accumulation under nitrogen deprivation. In the presence of acetate, downregulation of PDC2_E1α exerted little to no effect on TAG production and photosynthetic activity. In contrast, under photoautotrophic conditions, especially in the absence of a nitrogen source, a dramatic decline in photosynthetic oxygen evolution and photosystem II quantum yield against a background of the apparent over-reduction of the photosynthetic electron chain was recorded. Our results suggest an essential role of cpPDC in the supply of carbon precursors for de novo FA synthesis in microalgae under conditions of photoautotrophy. A shortage of this supply is detrimental to the nitrogen-starvation-induced synthesis of storage TAG, an important carbon and energy sink in stressed Chlamydomonas cells, thereby impairing the acclimation ability of the microalga. PMID:25210079

  19. Diversity, abundance, and activity of archaeal populations in oil-contaminated groundwater accumulated at the bottom of an underground crude oil storage cavity.

    PubMed

    Watanabe, Kazuya; Kodama, Yumiko; Hamamura, Natsuko; Kaku, Nobuo

    2002-08-01

    Fluorescence in situ hybridization has shown that cells labeled with an Archaea-specific probe (ARCH915) accounted for approximately 10% of the total cell count in oil-contaminated groundwater accumulated at the bottom of an underground crude oil storage cavity. Although chemical analyses have revealed vigorous consumption of nitrate in cavity groundwater, the present study found that the methane production rate was higher than the nitrate consumption rate. To characterize the likely archaeal populations responsible for methane production in this system, fragments of 16S ribosomal DNA (rDNA) were amplified by PCR using eight different combinations of universal and Archaea-specific primers. Sequence analysis of 324 clones produced 23 different archaeal sequence types, all of which were affiliated with the kingdom EURYARCHAEOTA: Among them, five sequence types (KuA1, KuA6, KuA12, KuA16, and KuA22) were obtained in abundance. KuA1 and KuA6 were closely related to the known methanogens Methanosaeta concilii (99% identical) and Methanomethylovorans hollandica (98%), respectively. Although no closely related organism was found for KuA12, it could be affiliated with the family METHANOMICROBIACEAE: KuA16 and KuA22 showed substantial homology only to some environmental clones. Both of these branched deeply in the Euryarchaeota, and may represent novel orders. Quantitative competitive PCR showed that KuA12 was the most abundant, accounting for approximately 50% of the total archaeal rDNA copies detected. KuA1 and KuA16 also constituted significant proportions of the total archaeal rDNA copies (7 and 17%, respectively). These results suggest that limited species of novel archaea were enriched in the oil storage cavity. An estimate of specific methane production rates suggests that they were active methanogens.

  20. Glutamine accumulation and up-regulation of glutamine synthetase activity in the swamp eel, Monopterus albus (Zuiew), exposed to brackish water.

    PubMed

    Tok, Chia Y; Chew, Shit F; Peh, Wendy Y X; Loong, Ai M; Wong, Wai P; Ip, Yuen K

    2009-05-01

    The swamp eel, Monopterus albus, is an air-breathing teleost which typically lives in freshwater but can also be found in estuaries, where it has to deal with ambient salinity fluctuations. Unlike other teleosts, its gills are highly degenerate. Hence, it may have uncommon osmoregulatory adaptations, but no information is available on its osmoregulatory capacity and mechanisms at present. In this study M. albus was exposed to a 5 day progressive increase in salinity from freshwater (1 per thousand) to brackish water (25 per thousand) and subsequently kept in 25 per thousand water for a total of 4 days. The results indicate that M. albus switched from hyperosmotic hyperionic regulation in freshwater to a combination of osmoconforming and hypoosmotic hypoionic regulation in 25 per thousand water. Exposure to 25 per thousand water resulted in relatively large increases in plasma osmolality, [Na(+)] and [Cl(-)]. Consequently, fish exposed to 25 per thousand water had to undergo cell volume regulation through accumulation of organic osmolytes and inorganic ions. Increases in tissue free amino acid content were apparently the result of increased protein degradation, decreased amino acid catabolism, and increased synthesis of certain non-essential amino acids. Here we report for the first time that glutamine is the major organic osmolyte in M. albus. Glutamine content increased to a phenomenal level of > 12 micromol g(-1) and > 30 micromol g(-1) in the muscle and liver, respectively, of fish exposed to 25 per thousand water. There were significant increases in glutamine synthetase (GS) activity in muscle and liver of these fish. In addition, exposure to 25 per thousand water for 4 days led to significant increases in GS protein abundance in both muscle and liver, indicating that increases in the expression of GS mRNA could have occurred.

  1. Lipoprotein-associated phospholipase A2, myeloperoxidase and vascular endothelial growth factor - predictors of high vascular risk in respiratory bacterial infections.

    PubMed

    Seri, A; Marta, D S; Madalan, A; Popescu, M; Tiglea, A I; Moldoveanu, E

    2016-01-01

    Objective. Respiratory bacterial infections are associated with important coagulation disturbances that amplify the pulmonary lesions and determine a more severe course of the disease. The aim of our study was to investigate the correlation between the evolution of the general clinical parameters and the occurrence of thrombotic events on one side, and plasma levels of selected proteins involved in inflammation and coagulation on the other side, with the intent to establish and to validate a laboratory test panel for the assessment of the vascular risk in patients with bacterial respiratory infections. Methods. The study included 111 patients (divided into two groups, 61 without thrombosis and 50 with thrombosis) with bacterial respiratory infections and 30 healthy controls, age and gender-matched. The baseline evaluation of the patients included clinical, biological, and respiratory examination. LpPLA2 and MPO activities were measured by the spectrophotometric method. VEGF was quantified with an ELISA kit. Results. The collected data showed a correlation between the occurrence of superimposed thrombosis in respiratory infection patients, and the intensity of the inflammatory process, reflected by the increased MPO activity, and the dynamics of LpPLA2 and VEGF. Conclusion. Bacterial respiratory infections associate thrombotic vascular events of various degrees of severity, which correlate with the intensity of the inflammatory process, and the severity of endothelium dysfunction at the level of microcirculation. Starting from the recorded data, and based on the established severity scales in use, it is possible to compute a vascular risk score that takes into consideration the values of the three biomarkers under investigation. Abbreviations: COPD = chronic obstructive pulmonary disease,hsCRP = high sensitivity C reactive protein,EC = endothelial cells, ICAM-1 = intercellular adhesion molecule1, LpPLA2 = lipoprotein-associated phospholipase A2, MPO

  2. Lipoprotein-associated phospholipase A2, myeloperoxidase and vascular endothelial growth factor - predictors of high vascular risk in respiratory bacterial infections

    PubMed Central

    Seri, A; Marta, DS; Madalan, A; Popescu, M; Tiglea, AI; Moldoveanu, E

    2016-01-01

    Objective. Respiratory bacterial infections are associated with important coagulation disturbances that amplify the pulmonary lesions and determine a more severe course of the disease. The aim of our study was to investigate the correlation between the evolution of the general clinical parameters and the occurrence of thrombotic events on one side, and plasma levels of selected proteins involved in inflammation and coagulation on the other side, with the intent to establish and to validate a laboratory test panel for the assessment of the vascular risk in patients with bacterial respiratory infections. Methods. The study included 111 patients (divided into two groups, 61 without thrombosis and 50 with thrombosis) with bacterial respiratory infections and 30 healthy controls, age and gender-matched. The baseline evaluation of the patients included clinical, biological, and respiratory examination. LpPLA2 and MPO activities were measured by the spectrophotometric method. VEGF was quantified with an ELISA kit. Results. The collected data showed a correlation between the occurrence of superimposed thrombosis in respiratory infection patients, and the intensity of the inflammatory process, reflected by the increased MPO activity, and the dynamics of LpPLA2 and VEGF. Conclusion. Bacterial respiratory infections associate thrombotic vascular events of various degrees of severity, which correlate with the intensity of the inflammatory process, and the severity of endothelium dysfunction at the level of microcirculation. Starting from the recorded data, and based on the established severity scales in use, it is possible to compute a vascular risk score that takes into consideration the values of the three biomarkers under investigation. Abbreviations: COPD = chronic obstructive pulmonary disease,hsCRP = high sensitivity C reactive protein,EC = endothelial cells, ICAM-1 = intercellular adhesion molecule1, LpPLA2 = lipoprotein-associated phospholipase A2, MPO

  3. Metabolism of isoniazid by neutrophil myeloperoxidase leads to isoniazid-NAD(+) adduct formation: A comparison of the reactivity of isoniazid with its known human metabolites.

    PubMed

    Khan, Saifur R; Morgan, Andrew G M; Michail, Karim; Srivastava, Nutan; Whittal, Randy M; Aljuhani, Naif; Siraki, Arno G

    2016-04-15

    The formation of isonicotinyl-nicotinamide adenine dinucleotide (INH-NAD(+)) via the mycobacterial catalase-peroxidase enzyme, KatG, has been described as the major component of the mode of action of isoniazid (INH). However, there are numerous human peroxidases that may catalyze this reaction. The role of neutrophil myeloperoxidase (MPO) in INH-NAD(+) adduct formation has never been explored; this is important, as neutrophils are recruited at the site of tuberculosis infection (granuloma) through infected macrophages' cell death signals. In our studies, we showed that neutrophil MPO is capable of INH metabolism using electron paramagnetic resonance (EPR) spin-trapping and UV-Vis spectroscopy. MPO or activated human neutrophils (by phorbol myristate acetate) catalyzed the oxidation of INH and formed several free radical intermediates; the inclusion of superoxide dismutase revealed a carbon-centered radical which is considered to be the reactive metabolite that binds with NAD(+). Other human metabolites, including N-acetyl-INH, N-acetylhydrazine, and hydrazine did not show formation of carbon-centered radicals, and either produced no detectable free radicals, N-centered free radicals, or superoxide, respectively. A comparison of these free radical products indicated that only the carbon-centered radical from INH is reducing in nature, based on UV-Vis measurement of nitroblue tetrazolium reduction. Furthermore, only INH oxidation by MPO led to a new product (λmax=326nm) in the presence of NAD(+). This adduct was confirmed to be isonicotinyl-NAD(+) using LC-MS analysis where the intact adduct was detected (m/z=769). The findings of this study suggest that neutrophil MPO may also play a role in INH pharmacological activity.

  4. Impaired Clearance and Enhanced Pulmonary Inflammatory/Fibrotic Response to Carbon Nanotubes in Myeloperoxidase-Deficient Mice

    PubMed Central

    Shvedova, Anna A.; Kapralov, Alexandr A.; Feng, Wei Hong; Kisin, Elena R.; Murray, Ashley R.; Mercer, Robert R.; St. Croix, Claudette M.; Lang, Megan A.; Watkins, Simon C.; Konduru, Nagarjun V.; Allen, Brett L.; Conroy, Jennifer; Kotchey, Gregg P.; Mohamed, Bashir M.; Meade, Aidan D.; Volkov, Yuri; Star, Alexander; Fadeel, Bengt; Kagan, Valerian E.

    2012-01-01

    Advancement of biomedical applications of carbonaceous nanomaterials is hampered by their biopersistence and pro-inflammatory action in vivo. Here, we used myeloperoxidase knockout B6.129X1-MPO (MPO k/o) mice and showed that oxidation and clearance of single walled carbon nanotubes (SWCNT) from the lungs of these animals after pharyngeal aspiration was markedly less effective whereas the inflammatory response was more robust than in wild-type C57Bl/6 mice. Our results provide direct evidence for the participation of MPO – one of the key-orchestrators of inflammatory response – in the in vivo pulmonary oxidative biodegradation of SWCNT and suggest new ways to control the biopersistence of nanomaterials through genetic or pharmacological manipulations. PMID:22479306

  5. Hypoxia-inducible factor 1 mediates hypoxia-induced cardiomyocyte lipid accumulation by reducing the DNA binding activity of peroxisome proliferator-activated receptor {alpha}/retinoid X receptor

    SciTech Connect

    Belanger, Adam J.; Luo Zhengyu; Vincent, Karen A.; Akita, Geoffrey Y.; Cheng, Seng H.; Gregory, Richard J.; Jiang Canwen

    2007-12-21

    In response to cellular hypoxia, cardiomyocytes adapt to consume less oxygen by shifting ATP production from mitochondrial fatty acid {beta}-oxidation to glycolysis. The transcriptional activation of glucose transporters and glycolytic enzymes by hypoxia is mediated by hypoxia-inducible factor 1 (HIF-1). In this study, we examined whether HIF-1 was involved in the suppression of mitochondrial fatty acid {beta}-oxidation in hypoxic cardiomyocytes. We showed that either hypoxia or adenovirus-mediated expression of a constitutively stable hybrid form (HIF-1{alpha}/VP16) suppressed mitochondrial fatty acid metabolism, as indicated by an accumulation of intracellular neutral lipid. Both treatments also reduced the mRNA levels of muscle carnitine palmitoyltransferase I which catalyzes the rate-limiting step in the mitochondrial import of fatty acids for {beta}-oxidation. Furthermore, adenovirus-mediated expression of HIF-1{alpha}/VP16 in cardiomyocytes under normoxic conditions also mimicked the reduction in the DNA binding activity of peroxisome proliferator-activated receptor {alpha} (PPAR{alpha})/retinoid X receptor (RXR), in the presence or absence of a PPAR{alpha} ligand. These results suggest that HIF-1 may be involved in hypoxia-induced suppression of fatty acid metabolism in cardiomyocytes by reducing the DNA binding activity of PPAR{alpha}/RXR.

  6. Reduced miR-200b and miR-200c expression contributes to abnormal hepatic lipid accumulation by stimulating JUN expression and activating the transcription of srebp1

    PubMed Central

    Sun, Libo; Lu, Yonggang; Dou, Lin; Huang, Xiuqing; Sun, Mingxiao; Pang, Cheng; Qu, Jing; Liu, Guanghui; Li, Jian

    2016-01-01

    Previous studies indicated that miR-200s participated in IL-6-induced hepatic insulin resistance. However, the role of miR-200s in hepatic lipid accumulation has not been elucidated. Here we found that miR-200b and miR-200c were reduced in the steatotic livers of mice fed a high-fat diet (HFD) and patients with nonalcoholic fatty liver disease. This down-regulation was accompanied by an increase in the expression of lipogenic proteins such as sterol regulatory element-binding protein 1 (SREBP1) and fatty acid synthase (FAS). The suppression of miR-200b and miR-200c in Hep1-6 and NCTC1469 hepatocytes enhanced intracellular triglyceride levels, which were associated with increased SREBP-1 and FAS protein levels. In contrast, the over-expression of miR-200b and miR-200c suppressed lipid accumulation and reduced the expression of SREBP1 and FAS in Hep1-6 and NCTC1469 cells transfected with miR-200b or miR-200c mimics. Importantly, the up-regulation of miR-200b and miR-200c could reverse oleic acid/palmitic acid-induced lipid accumulation in hepatocytes. A luciferase reporter assay identified that miR-200b and miR-200c could directly bind the 3′UTR of jun. JUN activated the transcription of srebp1 to increase lipid accumulation. The data also demonstrated that increased miR-200b and miR-200c expression might be associated with sitagliptin-reduced hepatic lipid accumulation in mice fed a HFD. These findings suggest, for the first time, that reduced miR-200b and miR-200c expression contributes to abnormal hepatic lipid accumulation by stimulating JUN expression and activating the transcription of srebp1. PMID:27166182

  7. Solids Accumulation Scouting Studies

    SciTech Connect

    Duignan, M. R.; Steeper, T. J.; Steimke, J. L.

    2012-09-26

    The objective of Solids Accumulation activities was to perform scaled testing to understand the behavior of remaining solids in a Double Shell Tank (DST), specifically AW-105, at Hanford during multiple fill, mix, and transfer operations. It is important to know if fissionable materials can concentrate when waste is transferred from staging tanks prior to feeding waste treatment plants. Specifically, there is a concern that large, dense particles containing plutonium could accumulate in poorly mixed regions of a blend tank heel for tanks that employ mixing jet pumps. At the request of the DOE Hanford Tank Operations Contractor, Washington River Protection Solutions, the Engineering Development Laboratory of the Savannah River National Laboratory performed a scouting study in a 1/22-scale model of a waste staging tank to investigate this concern and to develop measurement techniques that could be applied in a more extensive study at a larger scale. Simulated waste tank solids: Gibbsite, Zirconia, Sand, and Stainless Steel, with stainless steel particles representing the heavier particles, e.g., plutonium, and supernatant were charged to the test tank and rotating liquid jets were used to mix most of the solids while the simulant was pumped out. Subsequently, the volume and shape of the mounds of residual solids and the spatial concentration profiles for the surrogate for heavier particles were measured. Several techniques were developed and equipment designed to accomplish the measurements needed and they included: 1. Magnetic particle separator to remove simulant stainless steel solids. A device was designed and built to capture these solids, which represent the heavier solids during a waste transfer from a staging tank. 2. Photographic equipment to determine the volume of the solids mounds. The mounds were photographed as they were exposed at different tank waste levels to develop a composite of topographical areas. 3. Laser rangefinders to determine the volume of

  8. Oxidative stress caused by ozone exposure induces β-amyloid 1-42 overproduction and mitochondrial accumulation by activating the amyloidogenic pathway.

    PubMed

    Hernández-Zimbrón, L F; Rivas-Arancibia, S

    2015-09-24

    Oxidative stress is a major risk factor for Alzheimer's disease (AD) that has been suggested to be the trigger of AD pathology. However, whether oxidative damage precedes and contributes directly to the intracellular accumulation of beta amyloid 1-42 (βA42) peptide remains a matter of debate. Chronic exposure to low doses of ozone similar to the levels during a day of high pollution in México City causes a state of oxidative stress that elicits progressive neurodegeneration in the hippocampi of rats. Several reports have demonstrated that the mitochondria are among the first organelles to be affected by oxidative stress and βA42 toxicity and act as sites of the accumulation of βA42, which affects energy metabolism. However, the mechanisms related to the neurodegeneration process and organelle damage that occur in conditions of chronic exposure to low doses of ozone have not been demonstrated. To analyze the effect of chronic ozone chronic exposure on changes in the production and accumulation of the βA42 and βA40 peptides in the mitochondria of hippocampal neurons of rats exposed to ozone, we examined the mitochondrial expression levels of Presenilins 1 and 2 and ADAM10 to detect changes related to the oxidative stress caused by low doses of ozone (0.25ppm). The results revealed significant accumulations of βA42 peptide in the mitochondrial fractions on days 60 and 90 of ozone exposure along with reductions in beta amyloid 1-40 accumulation, significant overexpressions of Pres2 and significant reductions in ADAM10 expression. Beta amyloid immunodetection revealed that there were some intracellular deposits of βA42 and that βA42 and the mitochondrial markers OPA1 and COX1 colocalized. These results indicate that the time of exposure to ozone and the accumulation of βA42 in the mitochondria of the hippocampal cells of rats were correlated. Our results suggest that the accumulation of the βA42 peptide may promote mitochondrial dysfunction due to its

  9. Reactivity of selenium-containing compounds with myeloperoxidase-derived chlorinating oxidants: Second-order rate constants and implications for biological damage.

    PubMed

    Carroll, Luke; Pattison, David I; Fu, Shanlin; Schiesser, Carl H; Davies, Michael J; Hawkins, Clare L

    2015-07-01

    Hypochlorous acid (HOCl) and N-chloramines are produced by myeloperoxidase (MPO) as part of the immune response to destroy invading pathogens. However, MPO also plays a detrimental role in inflammatory pathologies, including atherosclerosis, as inappropriate production of oxidants, including HOCl and N-chloramines, causes damage to host tissue. Low molecular mass thiol compounds, including glutathione (GSH) and methionine (Met), have demonstrated efficacy in scavenging MPO-derived oxidants, which prevents oxidative damage in vitro and ex vivo. Selenium species typically have greater reactivity toward oxidants compared to the analogous sulfur compounds, and are known to be efficient scavengers of HOCl and other hypohalous acids produced by MPO. In this study, we examined the efficacy of a number of sulfur and selenium compounds to scavenge a range of biologically relevant N-chloramines and oxidants produced by both isolated MPO and activated neutrophils and characterized the resulting selenium-derived oxidation products in each case. A dose-dependent decrease in the concentration of each N-chloramine was observed on addition of the sulfur compounds (cysteine, methionine) and selenium compounds (selenomethionine, methylselenocysteine, 1,4-anhydro-4-seleno-L-talitol, 1,5-anhydro-5-selenogulitol) studied. In general, selenomethionine was the most reactive with N-chloramines (k2 0.8-3.4×10(3)M(-1) s(-1)) with 1,5-anhydro-5-selenogulitol and 1,4-anhydro-4-seleno-L-talitol (k2 1.1-6.8×10(2)M(-1) s(-1)) showing lower reactivity. This resulted in the formation of the respective selenoxides as the primary oxidation products. The selenium compounds demonstrated greater ability to remove protein N-chloramines compared to the analogous sulfur compounds. These reactions may have implications for preventing cellular damage in vivo, particularly under chronic inflammatory conditions.

  10. Sucrose phosphate synthase activity and the co-ordination of carbon partitioning during sucrose and amino acid accumulation in desiccation-tolerant leaf material of the C4 resurrection plant Sporobolus stapfianus during dehydration.

    PubMed

    Whittaker, Anne; Martinelli, Tommaso; Farrant, Jill M; Bochicchio, Adriana; Vazzana, Concetta

    2007-01-01

    Both sucrose and amino acids accumulate in desiccation-tolerant leaf material of the C(4) resurrection plant, Sporobolus stapfianus Gandoger (Poaceae). The present investigation was aimed at examining sucrose phosphate synthase (SPS) activity and various metabolic checkpoints involved in the co-ordination of carbon partitioning between these competing pathways during dehydration. In the initial phase of dehydration, photosynthesis and starch content declined to immeasurable levels, whilst significant increases in hexose sugars, sucrose, and amino acids were associated with concomitant significant increases in SPS and pyruvate kinase (PK) activities, and maximal activity levels of phosphoenolpyruvate carboxylase (PEPCase), NADP-dependent isocitrate dehydrogenase (NADP-ICDH), and NADH-dependent glutamate synthase (NADH-GOGAT). The next phase of dehydration was characterized by changes in metabolism coinciding with net hexose sugar phosphorylation. This phase was characterized by a further significant increase in sucrose accumulation, with increased rates of net sucrose accumulation and maximum rates of SPS activity measured under both saturating and limiting (inhibitory) conditions. SPS protein was also increased. The stronger competitive edge of SPS for carbon entering glycolysis during hexose phosphorylation was also demonstrated by the further decrease in respiration and the simultaneous, significant decline in both PEPCase and PK activities. A decreased anabolic demand for 2-oxoglutarate (2OG), which remained constant, was shown by the co-ordinated decrease in GOGAT. It is proposed that the further increase in amino acids in this phase of dehydration may be in part attributable to the breakdown of insoluble proteins.

  11. Temporal resolution of misfolded prion protein transport, accumulation, glial activation, and neuronal death in the retinas of mice inoculated with scrapie

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Currently, there is a lack of pathologic landmarks to describe the progression of prion disease in vivo. The goal of this work was to determine the temporal relationship between the transport of misfolded prion protein from the brain to the retina, the accumulation of PrPSc in the retina, the respon...

  12. Perennial peanut (Arachis glabrata Benth.) leaves contain hydroxycinnamoyl-CoA:tartaric acid hydroxycinnamoyl transferase activity and accumulate hydroxycinnamoyl-tartaric acid esters

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Many plants accumulate hydroxycinnamoyl esters to protect against abiotic and biotic stresses. Caffeoyl esters, in particular, can be substrates for endogenous polyphenol oxidases (PPOs). Recently, we showed that perennial peanut (Arachis glabrata Benth.) leaves contain PPO and identified one PPO su...

  13. Quantification of fecal neutrophils by MPO determination (myeloperoxidase) in patients with invasive diarrhea. Cuantificación de neutrofilos fecales mediante la determinación de MPO (Mieloperoxidasa) en pacientes con diarrea invasiva.

    PubMed

    Bustos, D; Greco, G; Yapur, V; Facente, A; Di Carlo, M; Bustos, F; Dapia, L; Ascione, A; Negri, G

    2000-01-01

    Myeloperoxidase (MPO), a specific polymorphonuclear leukocyte enzyme, has been used previously to quantify the number of neutrophils in tissue. MPO activity was found to be linearly related to the number of neutrophil cells. In an attempt to use this method in leukocytes measuring in stool, fecal MPO was solubilized with hexadecyltrimethylammonium bromide and the MPO activity was measured by a dianisidine-H2O2 assay. Stools from 10 normal subjects and 39 patients with diarrhea produced by enteropathogenic bacteria were examined for leukocytes by MPO activity as well as microscopically using methylene blue stain, MPO activity was positive in 36 patients (92%) and leukocytes were present by microscopic observation in 30 (77%). Fecal leukocytes were not found in healthy controls and the MPO activity was undectable. Stool MPO activity had a range of from 1.6 to 2,830.0 x 10(3) UMPO per gram of feces (median 460.0). The number of neutrophils obtained through MPO activity had a range of 6.0 to 13,216.0/ mm3 (median 1,261.0). Fecal MPO activity is a simple biochemical assay for the detection and quantification of fecal leukocytes.

  14. Response times from ensembles of accumulators

    PubMed Central

    Zandbelt, Bram; Purcell, Braden A.; Palmeri, Thomas J.; Logan, Gordon D.; Schall, Jeffrey D.

    2014-01-01

    Decision-making is explained by psychologists through stochastic accumulator models and by neurophysiologists through the activity of neurons believed to instantiate these models. We investigated an overlooked scaling problem: How does a response time (RT) that can be explained by a single model accumulator arise from numerous, redundant accumulator neurons, each of which individually appears to explain the variability of RT? We explored this scaling problem by developing a unique ensemble model of RT, called e pluribus unum, which embodies the well-known dictum “out of many, one.” We used the e pluribus unum model to analyze the RTs produced by ensembles of redundant, idiosyncratic stochastic accumulators under various termination mechanisms and accumulation rate correlations in computer simulations of ensembles of varying size. We found that predicted RT distributions are largely invariant to ensemble size if the accumulators share at least modestly correlated accumulation rates and RT is not governed by the most extreme accumulators. Under these regimes the termination times of individual accumulators was predictive of ensemble RT. We also found that the threshold measured on individual accumulators, corresponding to the firing rate of neurons measured at RT, can be invariant with RT but is equivalent to the specified model threshold only when the rate correlation is very high. PMID:24550315

  15. The Unusual Acid-Accumulating Behavior during Ripening of Cherimoya (Annona cherimola Mill.) is Linked to Changes in Transcription and Enzyme Activity Related to Citric and Malic Acid Metabolism.

    PubMed

    González-Agüero, Mauricio; Tejerina Pardo, Luis; Zamudio, María Sofía; Contreras, Carolina; Undurraga, Pedro; Defilippi, Bruno G

    2016-04-25

    Cherimoya (Annona cherimola Mill.) is a subtropical fruit characterized by a significant increase in organic acid levels during ripening, making it an interesting model for studying the relationship between acidity and fruit flavor. In this work, we focused on understanding the balance between the concentration of organic acids and the gene expression and activity of enzymes involved in the synthesis and degradation of these metabolites during the development and ripening of cherimoya cv. "Concha Lisa". Our results showed an early accumulation of citric acid and other changes associated with the accumulation of transcripts encoding citrate catabolism enzymes. During ripening, a 2-fold increase in malic acid and a 6-fold increase in citric acid were detected. By comparing the contents of these compounds with gene expression and enzymatic activity levels, we determined that cytoplasmic NAD-dependent malate dehydrogenase (cyNAD-MDH) and mitochondrial citrate synthase (mCS) play important regulatory roles in the malic and citric acid biosynthetic pathways.

  16. Lithium stimulates glutamate "release" and inositol 1,4,5-trisphosphate accumulation via activation of the N-methyl-D-aspartate receptor in monkey and mouse cerebral cortex slices.

    PubMed Central

    Dixon, J F; Los, G V; Hokin, L E

    1994-01-01

    Beginning at therapeutic concentrations (1-1.5 mM), the anti-manic-depressive drug lithium stimulated the release of glutamate, a major excitatory neurotransmitter in the brain, in monkey cerebral cortex slices in a time- and concentration-dependent manner, and this was associated with increased inositol 1,4,5-trisphosphate [Ins(1,4,5)P3] accumulation. (+/-)-3-(2-Carboxypiperazin-4-yl)propyl-1-phosphoric acid (CPP), dizocilpine (MK-801), ketamine, and Mg(2+)-antagonists to the N-methyl-D-aspartate (NMDA) receptor/channel complex selectively inhibited lithium-stimulated Ins(1,4,5)P3 accumulation. Antagonists to cholinergic-muscarinic, alpha 1-adrenergic, 5-hydroxytryptamine2 (serotoninergic), and H1 histaminergic receptors had no effect. Antagonists to non-NMDA glutamate receptors had no effect on lithium-stimulated Ins(1,4,5)P3 accumulation. Possible reasons for this are discussed. Similar results were obtained in mouse cerebral cortex slices. Carbetapentane, which inhibits glutamate release, inhibited lithium-induced Ins(1,4,5)P3 accumulation in this model. It is concluded that the primary effect of lithium in the cerebral cortex slice model is stimulation of glutamate release, which, presumably via activation of the NMDA receptor, leads to Ca2+ entry. Ins(1,4,5)P3 accumulation increases due to the presumed increased influx of intracellular Ca2+, which activates phospholipase C. These effects may have relevance to the therapeutic action of lithium in the treatment of manic depression as well as its toxic effects, especially at lithium blood levels above 1.5 mM. Images PMID:8078888

  17. Tor-Sch9 deficiency activates catabolism of the ketone body-like acetic acid to promote trehalose accumulation and longevity.

    PubMed

    Hu, Jia; Wei, Min; Mirzaei, Hamed; Madia, Federica; Mirisola, Mario; Amparo, Camille; Chagoury, Shawna; Kennedy, Brian; Longo, Valter D

    2014-06-01

    In mammals, extended periods of fasting leads to the accumulation of blood ketone bodies including acetoacetate. Here we show that similar to the conversion of leucine to acetoacetate in fasting mammals, starvation conditions induced ketone body-like acetic acid generation from leucine in S. cerevisiae. Whereas wild-type and ras2Δ cells accumulated acetic acid, long-lived tor1Δ and sch9Δ mutants rapidly depleted it through a mitochondrial acetate CoA transferase-dependent mechanism, which was essential for lifespan extension. The sch9Δ-dependent utilization of acetic acid also required coenzyme Q biosynthetic genes and promoted the accumulation of intracellular trehalose. These results indicate that Tor-Sch9 deficiency extends longevity by switching cells to an alternative metabolic mode, in which acetic acid can be utilized for the storage of stress resistance carbon sources. These effects are reminiscent of those described for ketone bodies in fasting mammals and raise the possibility that the lifespan extension caused by Tor-S6K inhibition may also involve analogous metabolic changes in higher eukaryotes.

  18. Molecular chaperone Hsp90 stabilizes Pih1/Nop17 to maintain R2TP complex activity that regulates snoRNA accumulation

    PubMed Central

    Zhao, Rongmin; Kakihara, Yoshito; Gribun, Anna; Huen, Jennifer; Yang, Guocheng; Khanna, May; Costanzo, Michael; Brost, Renée L.; Boone, Charles; Hughes, Timothy R.; Yip, Christopher M.; Houry, Walid A.

    2008-01-01

    Hsp90 is a highly conserved molecular chaperone that is involved in modulating a multitude of cellular processes. In this study, we identify a function for the chaperone in RNA processing and maintenance. This functionality of Hsp90 involves two recently identified interactors of the chaperone: Tah1 and Pih1/Nop17. Tah1 is a small protein containing tetratricopeptide repeats, whereas Pih1 is found to be an unstable protein. Tah1 and Pih1 bind to the essential helicases Rvb1 and Rvb2 to form the R2TP complex, which we demonstrate is required for the correct accumulation of box C/D small nucleolar ribonucleoproteins. Together with the Tah1 cofactor, Hsp90 functions to stabilize Pih1. As a consequence, the chaperone is shown to affect box C/D accumulation and maintenance, especially under stress conditions. Hsp90 and R2TP proteins are also involved in the proper accumulation of box H/ACA small nucleolar RNAs. PMID:18268103

  19. EPA, an omega-3 fatty acid, induces apoptosis in human pancreatic cancer cells: role of ROS accumulation, caspase-8 activation, and autophagy induction.

    PubMed

    Fukui, Masayuki; Kang, Ki Sung; Okada, Kazushi; Zhu, Bao Ting

    2013-01-01

    In a recent study, we showed that eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), two common omega-3 fatty acids, can cause ROS accumulation and subsequently induce caspase-8-dependent apoptosis in human breast cancer cells (Kang et al. [2010], PLoS ONE 5: e10296). In this study, we showed that the pancreas has a unique ability to accumulate EPA at a level markedly higher than several other tissues analyzed. Based on this finding, we sought to further investigate the anticancer actions of EPA and its analog DHA in human pancreatic cancer cells using both in vitro and in vivo models. EPA and DHA were found to induce ROS accumulation and caspase-8-dependent cell death in human pancreatic cancer cells (MIA-PaCa-2 and Capan-2) in vitro. Feeding animals with a diet supplemented with 5% fish oil, which contains high levels of EPA and DHA, also strongly suppresses the growth of MIA-PaCa-2 human pancreatic cancer xenografts in athymic nude mice, by inducing oxidative stress and cell death. In addition, we showed that EPA can concomitantly induce autophagy in these cancer cells, and the induction of autophagy diminishes its ability to induce apoptotic cell death. It is therefore suggested that combination of EPA with an autophagy inhibitor may be a useful strategy in increasing the therapeutic effectiveness in pancreatic cancer.

  20. Effect of organic complexation on copper accumulation and toxicity to the estuarine red macroalga Ceramium tenuicorne: a test of the free ion activity model.

    PubMed

    Ytreberg, Erik; Karlsson, Jenny; Hoppe, Sabina; Eklund, Britta; Ndungu, Kuria

    2011-04-01

    Current water quality criteria (WQC) regulations on copper toxicity to biota are still based on total dissolved (<0.4 μm membrane filter) copper concentrations with a hardness modification for freshwaters. There are however ongoing efforts to incorporate metal speciation in WQC and toxicity regulations (such as the biotic ligand model-BLM) for copper and other metals. Here, we show that copper accumulation and growth inhibition of the Baltic macroalga Ceramium tenuicorne exposed to copper in artificial seawater at typical coastal and estuarine DOC concentrations (similar to 2-4 mg/L-C as fulvic acid) are better correlated to weakly complexed and total dissolved copper concentrations rather than the free copper concentration [Cu2+]. Our results using a combination of competitive ligand exchange-adsorptive cathodic stripping voltammetry (CLE-ACSV) measurements and model calculations (using visual MINTEQ incorporating the Stockholm Humic Model) show that copper accumulation in C. tenuicorne only correlates linearly well to [Cu2+] at relatively high [Cu2+] and in the absence of fulvic acid. Thus the FIAM fails to describe copper accumulation in C. tenuicorne at copper and DOC concentrations typical of most marine waters. These results seem to indicate that at ambient total dissolved copper concentration in coastal and estuarine waters, C. tenuicorne might be able to access a sizable fraction of organically complexed copper when free copper concentration to the cell membrane is diffusion limited.

  1. Impact of land-use and long-term (>150 years) charcoal accumulation on microbial activity, biomass and community structure in temperate soils (Belgium).

    NASA Astrophysics Data System (ADS)

    Hardy, Brieuc; Cornelis, Jean-Thomas; Dufey, Joseph E.

    2015-04-01

    In the last decade, biochar has been increasingly investigated as a soil amendment for long-term soil carbon sequestration while improving soil fertility. On the short term, biochar application to soil generally increases soil respiration as well as microbial biomass and activity and affects significantly the microbial community structure. However, such effects are relatively short-term and tend to vanish over time. In our study, we investigated the long-term impact of charcoal accumulation and land-use on soil biota in temperate haplic Luvisols developed in the loess belt of Wallonia (Belgium). Charcoal-enriched soils were collected in the topsoil of pre-industrial (>150 years old) charcoal kilns in forest (4 sites) and cropland (5 sites). The topsoil of the adjacent charcoal-unaffected soils was sampled in a comparable way. Soils were characterized (pH, total, organic and inorganic C, total N, exchangeable Ca, Mg, K, Na, cation exchange capacity and available P) and natural soil organic matter (SOM) and black carbon (BC) contents were determined by differential scanning calorimetry. After rewetting at pF 2.5, soils were incubated during 140 days at 20 °C. At 70 days of incubation, 10 g of each soil were freeze dried in order to measure total microbial biomass and community structure by PLFA analysis. The PLFA dataset was analyzed by principal component analysis (PCA) while soil parameters were used as supplementary variables. For both agricultural and forest soils, the respiration rate is highly related to the total microbial biomass (R²=0.90). Both soil respiration and microbial biomass greatly depend on the SOM content, which indicates that the BC pool is relatively inert microbiologically. Land-use explains most of the variance in the PLFA dataset, largely governing the first principal component of the ACP. In forest soils, we observe a larger proportion of gram + bacteria, actinomycetes and an increased bacteria:fungi ratio compared to cropland, where gram

  2. p38 Mitogen-activated protein kinase and c-Jun NH2-terminal protein kinase regulate the accumulation of a tight junction protein, ZO-1, in cell-cell contacts in HaCaT cells.

    PubMed

    Minakami, Masahiko; Kitagawa, Norio; Iida, Hiroshi; Anan, Hisashi; Inai, Tetsuichiro

    2015-02-01

    To investigate the involvement of stress-activated protein kinases, JNK and p38 MAPK, in the assembly of tight junctions in keratinocytes, we treated HaCaT cells with various combinations of SP600125 (an inhibitor of JNK), SB202190 (an inhibitor of p38 MAPK) and anisomycin (an activator of both JNK and p38 MAPK) and examined the localization of ZO-1, an undercoat constitutive protein of the tight junction. Short-term (8h) incubation with SP600125, SB202190 or anisomycin induced the accumulation of ZO-1 in the cell-cell contacts, with reduced ZO-1 staining in the cytoplasm, while only long-term (24h) incubation with SP600125 induced the accumulation of ZO-1. SP600125, SB202190 or SP600125 plus SB202190 treatment induced thin linear staining for ZO-1 in the cell-cell contacts. Anisomycin treatment induced thick and irregular linear staining for ZO-1, while anisomycin plus SP600125 treatment induced zipper-like staining for ZO-1. Anisomycin plus SB202190 treatment or anisomycin plus both SP600125 and SB202190 treatment for 8h failed to lead to the accumulation of ZO-1 in cell-cell contacts, but induced thin linear staining with several gaps 16 h after removal of these agents. These results suggest that the localization of ZO-1 in cell-cell contacts is differently regulated by activation and inhibition of JNK and/or p38 MAPK depending on the incubation period.

  3. Influence of chloride on modification of unsaturated phosphatidylcholines by the myeloperoxidase/hydrogen peroxide/bromide system.

    PubMed

    Panasenko, Oleg M; Vakhrusheva, Tatyana; Tretyakov, Vadim; Spalteholz, Holger; Arnhold, Juergen

    2007-01-01

    The leukocyte enzyme myeloperoxidase (MPO) is capable of catalyzing the oxidation of chloride and bromide ions, at physiological concentrations of these substrates, by hydrogen peroxide, generating hypochlorous acid (HOCl) and hypobromous acid (HOBr), respectively. Our previous results showed that the hypohalous acids formed react with double bonds in phosphatidylcholines (PCs) to produce chloro- and bromohydrins. Lysophosphatidylcholine (lyso-PC) is additionally formed in PCs with two or more double bonds. This study was conducted to determine the effect physiological chloride concentration (140 mM) has on the formation of bromohydrins and lyso-PC from unsaturated PC upon treatment with the myeloperoxidase/hydrogen peroxide/bromide (MPO/H2O2/Br-) system using physiological bromide concentrations (20-100 microM). The composition of reaction products was analyzed by matrix-assisted laser desorption and ionization time-of-flight mass spectrometry (MALDI-TOF MS). With monounsaturated PC, we demonstrated that the rate and extent of mono-bromohydrin formation were higher in the samples with 140 mM chloride compared to those with no added chloride. Moreover, mono-bromohydrin came to be the major product and no mono-chlorohydrin was observed already at 60 microM bromide. We attributed these effects to the involvement of HOBr arising from the reaction of MPO-derived HOCl with bromide rather than to the exchange of bromide with chlorine atoms of chlorohydrins or direct formation of HOBr by MPO. The presence of chloride shifted the pH optimum for mono-bromohydrin formation (pH 5.0) toward neutral values, and a significant yield of mono-bromohydrin was detected at physiological pH values (7.0-7.4). For polyunsaturated PC, chloride enhanced also lyso-PC production, the effect being pronounced at bromide concentrations below 40 microM. The results indicate that at physiological levels of chloride and bromide, chloride promotes MPO-mediated formation of bromohydrins and lyso

  4. Modulation of the Cellular Accumulation and Intracellular Activity of Daptomycin towards Phagocytized Staphylococcus aureus by the P-Glycoprotein (MDR1) Efflux Transporter in Human THP-1 Macrophages and Madin-Darby Canine Kidney Cells▿

    PubMed Central

    Lemaire, Sandrine; Van Bambeke, Françoise; Mingeot-Leclercq, Marie-Paule; Tulkens, Paul M.

    2007-01-01

    P-glycoprotein (P-gp; MDR1), a major efflux transporter, recognizes various antibiotics and is present in macrophages. We have examined its effect on the modulation of the intracellular accumulation and activity of daptomycin towards phagocytized Staphylococcus aureus (ATCC 25923) in human THP-1 macrophages, in comparison with MDCK epithelial cells (wild type and MDCK-MDR1 overexpressing P-gp; the bulk of the protein was immunodetected at the surface of all three cell types). Daptomycin displayed concentration-dependent intracellular activity (Hill equation pattern) in THP-1 and MDCK cells with (i) 50% effective drug extracellular concentration (EC50; relative potency) and static concentrations at 9 to 10 times the MIC and (ii) maximal efficacy (Emax; CFU decrease at infinite extracellular drug concentration) at 1.6 to 2 log compared to that of the postphagocytosis inoculum. Verapamil (100 μM) and elacridar (GF 120918; 0.5 μM), two known inhibitors of P-gp, decreased daptomycin EC50 (about threefold) in THP-1 and MDCK cells without affecting Emax. Daptomycin EC50 was about three- to fourfold higher and accumulation in MDCK-MDR1 commensurately lower than in wild-type cells. In THP-1 macrophages, (i) verapamil and ATP depletion increased, and ouabain (an inducer of mdr1 [the gene encoding P-gp] expression) decreased the accumulation of daptomycin in parallel with that of DiOC2 (a known substrate of P-gp); (ii) silencing mdr1 with duplex human mdr1 siRNAs reduced the cell content in immunoreactive P-gp to 15 to 30% of controls and caused an eight- to 13-fold increase in daptomycin accumulation. We conclude that daptomycin is subject to efflux from THP-1 macrophages and MDCK cells by P-gp, which reduces its intracellular activity against phagocytized S. aureus. PMID:17548493

  5. (Accumulation of methyl-deficient rat liver messenger ribonucleic acid on ethionine administration). Progress report. [Methyltransferase activity in Ehrlich ascites tumor cells and effects of phorbol ester on methyltransferase activity

    SciTech Connect

    Borek, E.

    1980-01-01

    Enzyme fractions were isolated from Ehrlich ascites cells which introduced methyl groups into methyl deficient rat liver mRNA and unmethylated vaccinia mRNA. The methyl groups were incorporated at the 5' end into cap 1 structures by the viral enzyme, whereas both cap 0 and cap 1 structures were formed by the Ehrlich ascites cell enzymes. Preliminary results indicate the presence of adenine N/sup 6/-methyltransferase activity in Ehrlich ascites cells. These results indicate that mRNA deficient in 5'-cap methylation and in internal methylation of adenine accumulated in rats on exposure to ethionine. The methyl-deficient mRNA isolated from the liver of ethionine-fed rats differed in its translational properties from mRNA isolated from control animals. Preliminary experiments indicate that single topical application of 17n moles of TPA to mouse skin altered tRNA methyltransferases. The extent of methylation was increased over 2-fold in mouse skin treated with TPA for 48 hours. These changes have been observed as early as 12 hours following TPA treatment. In contrast, the application of initiating dose of DMBA had no effect on these enzymes. It should be emphasized that the changes in tRNA methyltransferases produced by TPA are not merely an increase of the concentration of the enzyme, rather that they represent alterations of specificity of a battery of enzymes. In turn the change in enzyme specificity can produce alterations in the structure of tRNA. (ERB)

  6. Absence of myeloperoxidase and CD8 positive cells in colorectal cancer infiltrates identifies patients with severe prognosis

    PubMed Central

    Däster, Silvio; Eppenberger-Castori, Serenella; Hirt, Christian; Soysal, Savas D; Delko, Tarik; Nebiker, Christian A; Weixler, Benjamin; Amicarella, Francesca; Iezzi, Giandomenica; Governa, Valeria; Padovan, Elisabetta; Mele, Valentina; Sconocchia, Giuseppe; Heberer, Michael; Terracciano, Luigi; Kettelhack, Christoph; Oertli, Daniel; Spagnoli, Giulio C; von Holzen, Urs; Tornillo, Luigi; Droeser, Raoul A

    2015-01-01

    Colorectal cancer (CRC) infiltration by cells expressing myeloperoxidase (MPO) or CD8 positive T lymphocytes has been shown to be independently associated with favorable prognosis. We explored the relationship occurring between CD8+ and MPO+ cell CRC infiltration, its impact on clinical-pathological features and its prognostic significance in a tissue microarray (TMA) including 1,162 CRC. We observed that CRC showing high MPO+ cell infiltration are characterized by a prognosis as favorable as that of cancers with high CD8+ T cell infiltration. However, MPO+ and CD8+ CRC infiltrating cells did not synergize in determining a more favorable outcome, as compared with cancers showing MPOhigh/CD8low or MPOlow/CD8high infiltrates. Most importantly, we identified a subgroup of CRC with MPOlow/CD8low tumor infiltration characterized by a particularly severe prognosis. Intriguingly, although MPO+ and CD8+ cells did not co-localize in CRC infiltrates, an increased expression of TIA-1 and granzyme-B was detectable in T cells infiltrating CRC with high MPO+ cell density. PMID:26587320

  7. First steps in the standardization of immunoglobulin IgG myeloperoxidase-anti-neutrophil cytoplasmic antibody measurements.

    PubMed

    Hutu, D P; Tuddenham, E; Monogioudi, E; Meroni, P; Schimmel, H; Sheldon, J; Zegers, I

    2016-02-01

    The standardization of immunoassays for immunoglobulin (Ig)G myeloperoxidase-anti-neutrophil cytoplasmic antibodies (MPO-ANCA) could contribute to a more accurate diagnosis and follow-up of small vessels-associated vasculitis, a systemic autoimmune disorder that leads to necrosis of blood vessel walls. Despite significant efforts by different groups, the level of comparability of results from commercially available immunoassays used for IgG MPO-ANCA detection is still poor. Therefore, the potential for improvement using reference materials was assessed. The evaluation of a set of 30 patient samples with 11 assays showed that differences between assays result in different interpretations for individual patients. Only 10 of 30 patient samples had the same clinical interpretation among 11 assays applying the cut-off values provided by each respective manufacturer. The correlation between results from 13 different assays was assessed in a pairwise manner. The correlation between results from patient samples was systematically very good for combinations of seven of those assays. The correlation of results ranged from reasonable to good for combinations with four other assays, therefore it should be possible to improve the comparability of results using a commutable reference material for calibration. Feasibility studies were conducted in order to find a reference material format most suitable for a calibrator. Two sets of candidate reference materials were produced from different raw materials, and assessed according to their suitability. A final format was selected, and a candidate reference material was produced.

  8. Bacterial accumulation in viscosity gradients

    NASA Astrophysics Data System (ADS)

    Waisbord, Nicolas; Guasto, Jeffrey

    2016-11-01

    Cell motility is greatly modified by fluid rheology. In particular, the physical environments in which cells function, are often characterized by gradients of viscous biopolymers, such as mucus and extracellular matrix, which impact processes ranging from reproduction to digestion to biofilm formation. To understand how spatial heterogeneity of fluid rheology affects the motility and transport of swimming cells, we use hydrogel microfluidic devices to generate viscosity gradients in a simple, polymeric, Newtonian fluid. Using video microscopy, we characterize the random walk motility patterns of model bacteria (Bacillus subtilis), showing that both wild-type ('run-and-tumble') cells and smooth-swimming mutants accumulate in the viscous region of the fluid. Through statistical analysis of individual cell trajectories and body kinematics in both homogeneous and heterogeneous viscous environments, we discriminate passive, physical effects from active sensing processes to explain the observed cell accumulation at the ensemble level.

  9. Association among active seafloor deformation, mound formation, and gas hydrate growth and accumulation within the seafloor of the Santa Monica Basin, offshore California

    USGS Publications Warehouse

    Paull, C.K.; Normark, W.R.; Ussler, W.; Caress, D.W.; Keaten, R.

    2008-01-01

    Seafloor blister-like mounds, methane migration and gas hydrate formation were investigated through detailed seafloor surveys in Santa Monica Basin, offshore of Los Angeles, California. Two distinct deep-water (??? 800??m water depth) topographic mounds were surveyed using an autonomous underwater vehicle (carrying a multibeam sonar and a chirp sub-bottom profiler) and one of these was explored with the remotely operated vehicle Tiburon. The mounds are > 10??m high and > 100??m wide dome-shaped bathymetric features. These mounds protrude from crests of broad anticlines (~ 20??m high and 1 to 3??km long) formed within latest Quaternary-aged seafloor sediment associated with compression between lateral offsets in regional faults. No allochthonous sediments were observed on the mounds, except slumped material off the steep slopes of the mounds. Continuous streams of methane gas bubbles emanate from the crest of the northeastern mound, and extensive methane-derived authigenic carbonate pavements and chemosynthetic communities mantle the mound surface. The large local vertical displacements needed to produce these mounds suggests a corresponding net mass accumulation has occurred within the immediate subsurface. Formation and accumulation of pure gas hydrate lenses in the subsurface is proposed as a mechanism to blister the seafloor and form these mounds. ?? 2008 Elsevier B.V. All rights reserved.

  10. Chinese herbal medicine Yi-Gan-San decreases the lipid accumulation in mouse 3T3-L1 adipocytes by modulating the activities of transcription factors SREBP-1c and FoxO1.

    PubMed

    Izumi, Masayuki; Seki, Takashi; Iwasaki, Koh; Sakamoto, Kazuichi

    2009-09-01

    Abnormal lipid metabolism in adipose tissue is closely related to the occurrence and progression of a wide variety of metabolic syndromes. We have analyzed the pharmacological effects of Chinese herbal medicines on cell differentiation and lipid metabolism in adipocytes. Yi-Gan-San (YGS) is a Chinese herbal medicine that is effective in treating the behavioral and psychological symptoms of dementia; however, its physiological mechanism remains unclear. We analyzed the effects of YGS on lipid accumulation in mouse 3T3-L1 adipocytes. Adipocyte differentiation was induced in mouse 3T3-L1 preadipocytes by treatment with the mixture of dexamethasone, 3-iso-butyl-1-methylxanthine, and insulin, and cells were cultured for 8 days with Chinese herbal medicines, including YGS. YGS effectively reduced the lipid accumulation in the differentiated 3T3-L1 cells in a dose-dependent manner, but had no effect on cell viability. YGS also reduced the activity of glycerol-3-phosphate dehydrogenase, an enzyme involved in lipid synthesis. In contrast, YGS gave no noticeable effect on glucose uptake and fatty acid uptake in the differentiated 3T3-L1 cells. Moreover, we established the stably transfected 3T3-L1 cell lines, each of which expresses the luciferase reporter gene under the control of sterol regulatory element-binding protein-1c (SREBP-1c) or FoxO1. SREBP-1c is a transcription factor involved in fatty acid synthesis, and FoxO1 is a forkhead-type transcription factor involved in adipocyte differentiation. Using these cell lines, we showed that YGS reduced the transcriptional activity of SREBP-1c, whereas YGS increased the activity of FoxO1. Thus, YGS may suppress lipid synthesis and fat accumulation in adipocytes through modulating the activities of SREBP-1c and FoxO1.

  11. Chimpanzee accumulative stone throwing.

    PubMed

    Kühl, Hjalmar S; Kalan, Ammie K; Arandjelovic, Mimi; Aubert, Floris; D'Auvergne, Lucy; Goedmakers, Annemarie; Jones, Sorrel; Kehoe, Laura; Regnaut, Sebastien; Tickle, Alexander; Ton, Els; van Schijndel, Joost; Abwe, Ekwoge E; Angedakin, Samuel; Agbor, Anthony; Ayimisin, Emmanuel Ayuk; Bailey, Emma; Bessone, Mattia; Bonnet, Matthieu; Brazolla, Gregory; Buh, Valentine Ebua; Chancellor, Rebecca; Cipoletta, Chloe; Cohen, Heather; Corogenes, Katherine; Coupland, Charlotte; Curran, Bryan; Deschner, Tobias; Dierks, Karsten; Dieguez, Paula; Dilambaka, Emmanuel; Diotoh, Orume; Dowd, Dervla; Dunn, Andrew; Eshuis, Henk; Fernandez, Rumen; Ginath, Yisa; Hart, John; Hedwig, Daniela; Ter Heegde, Martijn; Hicks, Thurston Cleveland; Imong, Inaoyom; Jeffery, Kathryn J; Junker, Jessica; Kadam, Parag; Kambi, Mohamed; Kienast, Ivonne; Kujirakwinja, Deo; Langergraber, Kevin; Lapeyre, Vincent; Lapuente, Juan; Lee, Kevin; Leinert, Vera; Meier, Amelia; Maretti, Giovanna; Marrocoli, Sergio; Mbi, Tanyi Julius; Mihindou, Vianet; Moebius, Yasmin; Morgan, David; Morgan, Bethan; Mulindahabi, Felix; Murai, Mizuki; Niyigabae, Protais; Normand, Emma; Ntare, Nicolas; Ormsby, Lucy Jayne; Piel, Alex; Pruetz, Jill; Rundus, Aaron; Sanz, Crickette; Sommer, Volker; Stewart, Fiona; Tagg, Nikki; Vanleeuwe, Hilde; Vergnes, Virginie; Willie, Jacob; Wittig, Roman M; Zuberbuehler, Klaus; Boesch, Christophe

    2016-02-29

    The study of the archaeological remains of fossil hominins must rely on reconstructions to elucidate the behaviour that may have resulted in particular stone tools and their accumulation. Comparatively, stone tool use among living primates has illuminated behaviours that are also amenable to archaeological examination, permitting direct observations of the behaviour leading to artefacts and their assemblages to be incorporated. Here, we describe newly discovered stone tool-use behaviour and stone accumulation sites in wild chimpanzees reminiscent of human cairns. In addition to data from 17 mid- to long-term chimpanzee research sites, we sampled a further 34 Pan troglodytes communities. We found four populations in West Africa where chimpanzees habitually bang and throw rocks against trees, or toss them into tree cavities, resulting in conspicuous stone accumulations at these sites. This represents the first record of repeated observations of individual chimpanzees exhibiting stone tool use for a purpose other than extractive foraging at what appear to be targeted trees. The ritualized behavioural display and collection of artefacts at particular locations observed in chimpanzee accumulative stone throwing may have implications for the inferences that can be drawn from archaeological stone assemblages and the origins of ritual sites.

  12. Accumulation of the planets

    NASA Technical Reports Server (NTRS)

    Wetherill, G. W.

    1987-01-01

    In modeling the accumulation of planetesimals into planets, it is appropriate to distinguish between two stages: an early stage, during which approximately 10 km diameter planetesimals accumulate locally to form bodies approximate 10 to the 25th g in mass; and a later stage in which the approximately 10 to the 25th g planetesimals accumulate into the final planets. In the terrestrial planet region, an initial planetesimal swarm corresponding to the critical mass of dust layer gravitational instabilities is considered. In order to better understand the accumulation history of Mercury-sized bodies, 19 Monte-Carlo simulations of terrestrial planet growth were calculated. A Monte Carlo technique was used to investigate the orbital evolution of asteroidal collision debris produced interior to 2.6 AU. It was found that there are two regions primarily responsible for production of Earth-crossing meteoritic material and Apollo objects. The same techniques were extended to include the origin of Earth-approaching asteroidal bodies. It is found that these same two resonant mechanisms predict a steady-state number of Apollo-Amor about 1/2 that estimated based on astronomical observations.

  13. Chimpanzee accumulative stone throwing

    PubMed Central

    Kühl, Hjalmar S.; Kalan, Ammie K.; Arandjelovic, Mimi; Aubert, Floris; D’Auvergne, Lucy; Goedmakers, Annemarie; Jones, Sorrel; Kehoe, Laura; Regnaut, Sebastien; Tickle, Alexander; Ton, Els; van Schijndel, Joost; Abwe, Ekwoge E.; Angedakin, Samuel; Agbor, Anthony; Ayimisin, Emmanuel Ayuk; Bailey, Emma; Bessone, Mattia; Bonnet, Matthieu; Brazolla, Gregory; Buh, Valentine Ebua; Chancellor, Rebecca; Cipoletta, Chloe; Cohen, Heather; Corogenes, Katherine; Coupland, Charlotte; Curran, Bryan; Deschner, Tobias; Dierks, Karsten; Dieguez, Paula; Dilambaka, Emmanuel; Diotoh, Orume; Dowd, Dervla; Dunn, Andrew; Eshuis, Henk; Fernandez, Rumen; Ginath, Yisa; Hart, John; Hedwig, Daniela; Ter Heegde, Martijn; Hicks, Thurston Cleveland; Imong, Inaoyom; Jeffery, Kathryn J.; Junker, Jessica; Kadam, Parag; Kambi, Mohamed; Kienast, Ivonne; Kujirakwinja, Deo; Langergraber, Kevin; Lapeyre, Vincent; Lapuente, Juan; Lee, Kevin; Leinert, Vera; Meier, Amelia; Maretti, Giovanna; Marrocoli, Sergio; Mbi, Tanyi Julius; Mihindou, Vianet; Moebius, Yasmin; Morgan, David; Morgan, Bethan; Mulindahabi, Felix; Murai, Mizuki; Niyigabae, Protais; Normand, Emma; Ntare, Nicolas; Ormsby, Lucy Jayne; Piel, Alex; Pruetz, Jill; Rundus, Aaron; Sanz, Crickette; Sommer, Volker; Stewart, Fiona; Tagg, Nikki; Vanleeuwe, Hilde; Vergnes, Virginie; Willie, Jacob; Wittig, Roman M.; Zuberbuehler, Klaus; Boesch, Christophe

    2016-01-01

    The study of the archaeological remains of fossil hominins must rely on reconstructions to elucidate the behaviour that may have resulted in particular stone tools and their accumulation. Comparatively, stone tool use among living primates has illuminated behaviours that are also amenable to archaeological examination, permitting direct observations of the behaviour leading to artefacts and their assemblages to be incorporated. Here, we describe newly discovered stone tool-use behaviour and stone accumulation sites in wild chimpanzees reminiscent of human cairns. In addition to data from 17 mid- to long-term chimpanzee research sites, we sampled a further 34 Pan troglodytes communities. We found four populations in West Africa where chimpanzees habitually bang and throw rocks against trees, or toss them into tree cavities, resulting in conspicuous stone accumulations at these sites. This represents the first record of repeated observations of individual chimpanzees exhibiting stone tool use for a purpose other than extractive foraging at what appear to be targeted trees. The ritualized behavioural display and collection of artefacts at particular locations observed in chimpanzee accumulative stone throwing may have implications for the inferences that can be drawn from archaeological stone assemblages and the origins of ritual sites. PMID:26923684

  14. Accumulation of CD62P during storage of apheresis platelet concentrates and the role of CD62P in transfusion-related acute lung injury.

    PubMed

    Tong, Shan; Wang, Haibao; Zhang, Ting; Chen, Linfeng; Liu, Bowei

    2015-11-01

    Transfusion-related acute lung injury (TRALI) is the leading cause of transfusion-associated morbidity and mortality. Activated platelets have important roles in TRALI and CD62P was identified to be an important indicator of platelet activation. However, the precise roles of CD62P in TRALI have remained elusive. The present study assessed CD62P accumulation during storage of apheresis platelet concentrates (A‑Plts) and established a mouse model of TRALI to further investigate the roles of CD62P in TRALI. The results showed that the CD62P concentration in A‑Plts was increased with the storage time. Mice were treated with monoclonal major histocompatibility complex (MHC)‑1 antibody to induce TRALI. The murine model of TRALI was successfully established as evidenced by pulmonary oedema, accompanied by decreased clearance of bronchoalveolar lavage fluid (BALF), increased pulmonary and systemic inflammation, elevated lung myeloperoxidase (MPO) activity as well as increased pulmonary and systemic coagulation in the TRALI group compared with those in the control group. To further determine the role of CD62P in TRALI, mice were treated with anti‑CD62P antibody to knockdown CD62P in vivo. It was found that pulmonary oedema, BALF clearance, pulmonary and systemic inflammation, MPO activity as well as pulmonary and systemic coagulation were decreased in the TRALI + anti‑CD62P antibody group compared with those in the TRALI + isotype antibody group. The present study supported the notion that CD62P is involved in mediating TRALI and may provide an important molecular basis for enhancing the clinical safety and effectiveness of platelet transfusion.

  15. Haptoglobin and myeloperoxidase (- G463A) gene polymorphisms in Brazilian sickle cell pat