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Sample records for mesophyll cells glutamine

  1. Amino acid transport across the tonoplast of vacuoles isolated from barley mesophyll protoplasts: Uptake of alanine, leucine, and glutamine

    Dietz, K.J.; Jaeger, R.; Kaiser, G.; Martinoia, E.

    1990-01-01

    Mesophyll protoplasts from leaves of well-fertilized barley (Hordeum vulgare L.) plants contained amino acids at concentrations as high as 120 millimoles per liter. With the exception of glutamic acid, which is predominantly localized in the cytoplasm, a major part of all other amino acids was contained inside the large central vacuole. Alanine, leucine, and glutamine are the dominant vacuolar amino acids in barley. Their transport into isolated vacuoles was studied using 14 C-labeled amino acids. Uptake was slow in the absence of ATP. A three- to sixfold stimulation of uptake was observed after addition of ATP or adenylyl imidodiphosphate an ATP analogue not being hydrolyzed by ATPases. Other nucleotides were ineffective in increasing the rate of uptake. ATP-Stimulated amino acid transport was not dependent on the transtonoplast pH or membrane potential. p-Chloromercuriphenylsulfonic acid and n-ethyl maleimide increased transport independently of ATP. Neutral amino acids such as valine or leucine effectively decreased the rate of alanine transport. Glutamine and glycine were less effective or not effective as competitive inhibitors of alanine transport. The results indicate the existence of a uniport translocator specific for neutral or basic amino acids that is under control of metabolic effectors

  2. Metabolomic Responses of Guard Cells and Mesophyll Cells to Bicarbonate

    Misra, Biswapriya B.; de Armas, Evaldo; Tong, Zhaohui; Chen, Sixue

    2015-01-01

    Anthropogenic CO2 presently at 400 ppm is expected to reach 550 ppm in 2050, an increment expected to affect plant growth and productivity. Paired stomatal guard cells (GCs) are the gate-way for water, CO2, and pathogen, while mesophyll cells (MCs) represent the bulk cell-type of green leaves mainly for photosynthesis. We used the two different cell types, i.e., GCs and MCs from canola (Brassica napus) to profile metabolomic changes upon increased CO2 through supplementation with bicarbonate (HCO3 -). Two metabolomics platforms enabled quantification of 268 metabolites in a time-course study to reveal short-term responses. The HCO3 - responsive metabolomes of the cell types differed in their responsiveness. The MCs demonstrated increased amino acids, phenylpropanoids, redox metabolites, auxins and cytokinins, all of which were decreased in GCs in response to HCO3 -. In addition, the GCs showed differential increases of primary C-metabolites, N-metabolites (e.g., purines and amino acids), and defense-responsive pathways (e.g., alkaloids, phenolics, and flavonoids) as compared to the MCs, indicating differential C/N homeostasis in the cell-types. The metabolomics results provide insights into plant responses and crop productivity under future climatic changes where elevated CO2 conditions are to take center-stage. PMID:26641455

  3. FUNCTION OF MALATDEHYDROGENASE COMPLEX OF MAIZE MESOPHYLL AND BUNDLE SHEATH CELLS UNDER SALT STRESS CONDITION

    Еprintsev А.Т.

    2006-12-01

    Full Text Available Salt-induced changes in malatdehydrogenase system activity make the essential contribution to cell adaptation to stress condition. The enzyme systems of C4-plants are most interesting due to their ability for adaptation to environment conditions. The role of separate components of malatdehydrogenase complex of mesophyll and bundle sheath cells of corn in formation of adaptive reaction in stressful conditions is investigated in presented work.The activation of all enzymes of malatdehydrogenase system and the subsequent decrease in their activity was observed in mesophyll durring the first stage of adaptation to salt influence. In bundle sheath cells such parameters are differed from control less essentially. Fast accumulation of piruvate in cells and malate in both investigated tissues was induced. The further salinity led to falling of concentration this intermediate. The concentration of piruvate was below control level, and it was raised by the end of an exposition.The results show that sodium chloride causes induction of Krebs-cycle in mesophyll and bundle sheath cells of corn and intensification of Hatch-Slack cycle. The described differences in function malatdehydrogenase systems of mesophyll and bundle sheath cells of leaves of corn under salinity mainly consist of the activity of enzymes of a studied complex in bundle sheath cells is subject to the minimal changes in comparison with mesophyll. Role of this enzymesystem in mechanisms of adaptive reaction of various tissues of corn to salt stress is discussed.

  4. Three-dimensional intracellular structure of a whole rice mesophyll cell observed with FIB-SEM.

    Oi, Takao; Enomoto, Sakiko; Nakao, Tomoyo; Arai, Shigeo; Yamane, Koji; Taniguchi, Mitsutaka

    2017-07-01

    Ultrathin sections of rice leaf blades observed two-dimensionally using a transmission electron microscope (TEM) show that the chlorenchyma is composed of lobed mesophyll cells, with intricate cell boundaries, and lined with chloroplasts. The lobed cell shape and chloroplast positioning are believed to enhance the area available for the gas exchange surface for photosynthesis in rice leaves. However, a cell image revealing the three-dimensional (3-D) ultrastructure of rice mesophyll cells has not been visualized. In this study, a whole rice mesophyll cell was observed using a focused ion beam scanning electron microscope (FIB-SEM), which provides many serial sections automatically, rapidly and correctly, thereby enabling 3-D cell structure reconstruction. Rice leaf blades were fixed chemically using the method for conventional TEM observation, embedded in resin and subsequently set in the FIB-SEM chamber. Specimen blocks were sectioned transversely using the FIB, and block-face images were captured using the SEM. The sectioning and imaging were repeated overnight for 200-500 slices (each 50 nm thick). The resultant large-volume image stacks ( x = 25 μm, y = 25 μm, z = 10-25 μm) contained one or two whole mesophyll cells. The 3-D models of whole mesophyll cells were reconstructed using image processing software. The reconstructed cell models were discoid shaped with several lobes around the cell periphery. The cell shape increased the surface area, and the ratio of surface area to volume was twice that of a cylinder having the same volume. The chloroplasts occupied half the cell volume and spread as sheets along the cell lobes, covering most of the inner cell surface, with adjacent chloroplasts in close contact with each other. Cellular and sub-cellular ultrastructures of a whole mesophyll cell in a rice leaf blade are demonstrated three-dimensionally using a FIB-SEM. The 3-D models and numerical information support the hypothesis that rice mesophyll

  5. Characterization of a light-controlled anion channel in the plasma membrane of mesophyll cells of pea

    Elzenga, J.T.M.; Volkenburgh Van, E

    In leaf mesophyll cells of pea (Pisum sativum) light induces a transient depolarization that is at least partly due to an increased plasma membrane conductance for anions. Several channel types were identified in the plasma membrane of protoplasts from mesophyll cells using the patch-clamp

  6. Factors affecting polyhydroxybutyrate accumulation in mesophyll cells of sugarcane and switchgrass

    2014-01-01

    Background Polyhydroxyalkanoates are linear biodegradable polyesters produced by bacteria as a carbon store and used to produce a range of bioplastics. Widespread polyhydroxyalkanoate production in C4 crops would decrease petroleum dependency by producing a renewable supply of biodegradable plastics along with residual biomass that could be converted into biofuels or energy. Increasing yields to commercial levels in biomass crops however remains a challenge. Previously, lower accumulation levels of the short side chain polyhydroxyalkanoate, polyhydroxybutyrate (PHB), were observed in the chloroplasts of mesophyll (M) cells compared to bundle sheath (BS) cells in transgenic maize (Zea mays), sugarcane (Saccharum sp.), and switchgrass (Panicum virgatum L.) leading to a significant decrease in the theoretical yield potential. Here we explore various factors which might affect polymer accumulation in mesophyll cells, including targeting of the PHB pathway enzymes to the mesophyll plastid and their access to substrate. Results The small subunit of Rubisco from pea effectively targeted the PHB biosynthesis enzymes to both M and BS chloroplasts of sugarcane and switchgrass. PHB enzyme activity was retained following targeting to M plastids and was equivalent to that found in the BS plastids. Leaf total fatty acid content was not affected by PHB production. However, when fatty acid synthesis was chemically inhibited, polymer accumulated in M cells. Conclusions In this study, we provide evidence that access to substrate and neither poor targeting nor insufficient activity of the PHB biosynthetic enzymes may be the limiting factor for polymer production in mesophyll chloroplasts of C4 plants. PMID:25209261

  7. Physiological implications of seasonal variation in membrane-associated calcium in red spruce mesophyll cells

    D.H. DeHayes; P.G. Schaberg; G.J. Hawley; C.H. Borer; J.R. Cumming; J.R. Strimbeck

    1997-01-01

    We examined the pattern of seasonal variation in total foliar calcium (Ca) pools and plasma membrane-associated Ca (mCa) in mesophyll cells of current-year and 1-year-old needles of red spruce (Picea rubens Sarg.) and the relationship between mCa and total foliar Ca on an individual plant and seasonal basis. Foliar samples were collected from...

  8. Effects of shading on the photosynthetic characteristics and mesophyll cell ultrastructure of summer maize.

    Ren, Baizhao; Cui, Haiyan; Camberato, James J; Dong, Shuting; Liu, Peng; Zhao, Bin; Zhang, Jiwang

    2016-08-01

    A field experiment was conducted to study the effects of shading on the photosynthetic characteristics and mesophyll cell ultrastructure of two summer maize hybrids Denghai605 (DH605) and Zhengdan958 (ZD958). The ambient sunlight treatment was used as control (CK) and shading treatments (40 % of ambient sunlight) were applied at different growth stages from silking (R1) to physiological maturity (R6) (S1), from the sixth leaf stage (V6) to R1 (S2), and from seeding to R6 (S3), respectively. The net photosynthetic rate (P n) was significantly decreased after shading. The greatest reduction of P n was found at S3 treatment, followed by S1 and S2 treatments. P n of S3 was decreased by 59 and 48 % for DH605, and 39 and 43 % for ZD958 at tasseling and milk-ripe stages, respectively, compared to that of CK. Additionally, leaf area index (LAI) and chlorophyll content decreased after shading. In terms of mesophyll cell ultrastructure, chloroplast configuration of mesophyll cells dispersed, and part of chloroplast swelled and became circular. Meanwhile, the major characteristics of chloroplasts showed poorly developed thylakoid structure at the early growth stage, blurry lamellar structure, loose grana, and a large gap between slices and warping granum. Then, plasmolysis occurred in mesophyll cells and the endomembrane system was destroyed, which resulted in the dissolution of cell membrane, karyotheca, mitochondria, and some membrane structures. The damaged mesophyll cell ultrastructure led to the decrease of photosynthetic capacity, and thus resulted in significant yield reduction by 45, 11, and 84 % in S1, S2, and S3 treatments, respectively, compared to that of CK.

  9. Distinct abscisic acid signaling pathways for modulation of guard cell versus mesophyll cell potassium channels revealed by expression studies in Xenopus laevis oocytes

    Sutton, F.; Paul, S. S.; Wang, X. Q.; Assmann, S. M.; Evans, M. L. (Principal Investigator)

    2000-01-01

    Regulation of guard cell ion transport by abscisic acid (ABA) and in particular ABA inhibition of a guard cell inward K(+) current (I(Kin)) is well documented. However, little is known concerning ABA effects on ion transport in other plant cell types. Here we applied patch clamp techniques to mesophyll cell protoplasts of fava bean (Vicia faba cv Long Pod) plants and demonstrated ABA inhibition of an outward K(+) current (I(Kout)). When mesophyll cell protoplast mRNA (mesophyll mRNA) was expressed in Xenopus laevis oocytes, I(Kout) was generated that displayed similar properties to I(Kout) observed from direct analysis of mesophyll cell protoplasts. I(Kout) expressed by mesophyll mRNA-injected oocytes was inhibited by ABA, indicating that the ABA signal transduction pathway observed in mesophyll cells was preserved in the frog oocytes. Co-injection of oocytes with guard cell protoplast mRNA and cRNA for KAT1, an inward K(+) channel expressed in guard cells, resulted in I(Kin) that was similarly inhibited by ABA. However, oocytes co-injected with mesophyll mRNA and KAT1 cRNA produced I(Kin) that was not inhibited by ABA. These results demonstrate that the mesophyll-encoded signaling mechanism could not substitute for the guard cell pathway. These findings indicate that mesophyll cells and guard cells use distinct and different receptor types and/or signal transduction pathways in ABA regulation of K(+) channels.

  10. Signal function of cytokinin 6-benzylaminopurine in the reaction of Triticum aestivum L. mesophyll cells to hyperthermia

    M. M. Musienko

    2014-12-01

    Full Text Available The signaling effect of 6-benzylaminopurine (BAP on leaf mesophyll cells of Triticum aestivum L. under hyperthermic conditions was studied­. It was found that BAP regulated photosynthetic pigment, hydrogen peroxide content and activity of antioxidant enzymes, namely superoxide dismutase, ascorbate peroxidase and catalase under high-temperature conditions. The additive effect of BAP and high temperature on the activation of cell antioxidant systems was demonstrated. BAP regulated reducing processes in mesophyll leaf cells under high-temperature conditions.

  11. Uptake of /sup 86/Rb/sup +/ into photoautotrophic mesophyll cells of Papaver somniferum

    Kaiser, W.M.; Jeschke, W.D.; Hartung, W.

    1982-06-01

    Uptake of /sup 86/Rb/sup +/, used as a tracer for potassium, into isolated photoautotrophic mesophyll cells of Papaver somniferum was weakly but consistently stimulated in the light. It showed mono-phasic saturation kinetics with a pH optimum of 7.0, a Vsub(max) of 6.7 ..mu..mol mg/sup -1/ Chl x h/sup -1/ and a Ksub(m) of 2.7 mmol l/sup -1/. Different anions as Cl/sup -/, NO/sub 3//sup -/ and PO/sub 4//sup 3 -/ had no effects on /sup 86/Rb/sup +/ uptake. Sodium ions influenced Rb/sup +/-uptake very weakly, indicating a high K/sup +/ -specificity of the mesophyll cell plasmalemma. Fusicoccin stimulated /sup 86/Rb/sup +/ -uptake strongly whereas abscisic acid inhibited uptake only following preincubation for two hours. Nitrite, CCCP and Dio-9 inhibited /sup 86/Rb/sup +/-uptake which gives evidence that this process is dependent on intact pH-gradients within the cells and on ATP-formation.

  12. Evidence for a specific glutamate/H+ cotransport in isolated mesophyll cells

    McCutcheon, S.L.; Bown, A.W.

    1987-01-01

    Mechanically isolated Asparagus sprengeri Regel mesophyll cells were suspended in 1 millimolar CaSO 4 . Immediate alkalinization of the medium occurred on the addition of 1 millimolar concentrations of L-glutamate (Glu) and its analog L-methionine-D,L-sulfoximine (L-MSO). D-Glu and the L isomers of the protein amino acids did not elicit alkalinization. L-Glu dependent alkalinization was transient and acidification resumed after approximately 30 to 45 minutes. At pH 6.0, 5 millimolar L-Glu stimulated initial rates of alkalinization that varied between 1.3 to 4.1 nmol H + /10 6 cells minute. L-Glu dependent alkalinization was saturable, increased with decreasing pH, was inhibited by carbonyl cyanide-p-trichloromethoxyphenyl hydrazone (CCCP), and was not stimulated by light. Uptake of L-[U- 14 C]glutamate increased as the pH decreased from 6.5 to 5.5, and was inhibited by L-MSO. L-Glu had no influence on K + efflux. Although evidence for multiple amino acid/proton cotransport systems has been found in other tissues, the present report indicates that a highly specific L-Glu/proton uptake process is present in Asparagus mesophyll cells

  13. Air pollution effects on the ultrastructure of Phlomis fruticosa mesophyll cells

    Psaras, G.K.; Christodoulakis, N.S.

    1987-04-01

    Plant physiologists and environmental scientists suggest that a basic effect of air pollution on plants leads towards the minimization of their productivity. On the other hand the action of individual pollutants on intact plants has been studied from biochemical as well as structural viewpoint. Thus the study of plant responses to SO/sub 2/ exposure revealed that this agent causes acute and chronic injury. Chronic injury results in chlorosis and subsequent necrosis due to destruction of chlorophylls and final chloroplast lysis. It has been documented that ultrastructural characteristics of leaves are affected prior to any visible injury. Electron microscope examination of SO/sub 2/ fumigated plant-attached leaves of Vicia faba revealed chloroplast thylakoids starting to swell whilst photosynthesis rate was drastically reduced. The first light microscope-detected effects of air pollution on the leaf structure of plants common in natural ecosystems of Athens metropolitan area, have been reported. A chlorosis phenomenon in Urginea maritima leaves as well as an indication of detrimental effects of Phlomis fruticosa mesophyll chloroplasts were documented. In this work further investigation has been undertaken in order to elucidate the precise effects of air pollution on the ultrastructure of the photosynthesizing mesophyll cells.

  14. Vacuolar Localization of Endoproteinases EP(1) and EP(2) in Barley Mesophyll Cells.

    Thayer, S S; Huffaker, R C

    1984-05-01

    The localization of two previously characterized endoproteinases (EP(1) and EP(2)) that comprise more than 95% of the protease activity in primary Hordeum vulgare L. var Numar leaves was determined. Intact vacuoles released from washed mesophyll protoplasts by gentle osmotic shock and increase in pH, were purified by flotation through a four-step Ficoll gradient. These vacuoles contained endoproteinases that rapidly degraded purified barley ribulose-1,5-bisphosphate carboxylase (RuBPCase) substrate. Breakdown products and extent of digestion of RuBPCase were determined using 12% polyacrylamide-sodium dodecyl sulfate gels. Coomassie brilliant blue- or silver-stained gels were scanned, and the peaks were integrated to provide quantitative information. The characteristics of the vacuolar endoproteinases (e.g. sensitivity to various inhibitors and activators, and the molecular weights of the breakdown products, i.e. peptide maps) closely resembled those of purified EP(1) and partially purified EP(2). It is therefore concluded that EP(1) and EP(2) are localized in the vacuoles of mesophyll cells.

  15. Macroautophagy and microautophagy in relation to vacuole formation in mesophyll cells of Dendrobium tepals.

    van Doorn, Wouter G; Kirasak, Kanjana; Ketsa, Saichol

    2015-04-01

    Prior to flower opening, mesophyll cells at the vascular bundles of Dendrobium tepals showed a large increase in vacuolar volume, partially at the expense of the cytoplasm. Electron micrographs indicated that this increase in vacuolar volume was mainly due to vacuole fusion. Macroautophagous structures typical of plant cells were observed. Only a small part of the decrease in cytoplasmic volume seemed due to macroautophagy. The vacuoles contained vesicles of various types, including multilamellar bodies. It was not clear if these vacuolar inclusions were due to macroautophagy or microautophagy. Only a single structure was observed of a protruding vacuole, indicating microautophagy. It is concluded that macroautophagy occurs in these cells but its role in vacuole formation seems small, while a possible role of microautophagy in vacuole formation might be hypothesized. Careful labeling of organelle membranes seems required to advance our insight in plant macro- and microautophagy and their roles in vacuole formation. Copyright © 2015 Elsevier GmbH. All rights reserved.

  16. Importance of glutamine metabolism in leukemia cells by energy production through TCA cycle and by redox homeostasis.

    Goto, Mineaki; Miwa, Hiroshi; Shikami, Masato; Tsunekawa-Imai, Norikazu; Suganuma, Kazuto; Mizuno, Shohei; Takahashi, Miyuki; Mizutani, Motonori; Hanamura, Ichiro; Nitta, Masakazu

    2014-07-01

    Some cancer cells depend on glutamine despite of pronounced glycolysis. We examined the glutamine metabolism in leukemia cells, and found that HL-60 cells most depended on glutamine in the 4 acute myelogenous leukemia (AML) cell lines examined: growth of HL-60 cells was most suppressed by glutamine deprivation and by inhibition of glutaminolysis, which was rescued by tricarboxylic acid (TCA) cycle intermediate, oxaloacetic acid. Glutamine is also involved in antioxidant defense function by increasing glutathione. Glutamine deprivation suppressed the glutathione content and elevated reactive oxygen species most evidently in HL-60 cells. Glutamine metabolism might be a therapeutic target in some leukemia.

  17. Pyruvate carboxylase is required for glutamine-independent growth of tumor cells

    Cheng, Tzuling; Sudderth, Jessica; Yang, Chendong; Mullen, Andrew R.; Jin, Eunsook S.; Matés, José M.; DeBerardinis, Ralph J.

    2011-01-01

    Tumor cells require a constant supply of macromolecular precursors, and interrupting this supply has been proposed as a therapeutic strategy in cancer. Precursors for lipids, nucleic acids, and proteins are generated in the tricarboxylic acid (TCA) cycle and removed from the mitochondria to participate in biosynthetic reactions. Refilling the pool of precursor molecules (anaplerosis) is therefore crucial to maintain cell growth. Many tumor cells use glutamine to feed anaplerosis. Here we studied how “glutamine-addicted” cells react to interruptions of glutamine metabolism. Silencing of glutaminase (GLS), which catalyzes the first step in glutamine-dependent anaplerosis, suppressed but did not eliminate the growth of glioblastoma cells in culture and in vivo. Profiling metabolic fluxes in GLS-suppressed cells revealed induction of a compensatory anaplerotic mechanism catalyzed by pyruvate carboxylase (PC), allowing the cells to use glucose-derived pyruvate rather than glutamine for anaplerosis. Although PC was dispensable when glutamine was available, forcing cells to adapt to low-glutamine conditions rendered them absolutely dependent on PC for growth. Furthermore, in other cell lines, measuring PC activity in nutrient-replete conditions predicted dependence on specific anaplerotic enzymes. Cells with high PC activity were resistant to GLS silencing and did not require glutamine for survival or growth, but displayed suppressed growth when PC was silenced. Thus, PC-mediated, glucose-dependent anaplerosis allows cells to achieve glutamine independence. Induction of PC during chronic suppression of glutamine metabolism may prove to be a mechanism of resistance to therapies targeting glutaminolysis. PMID:21555572

  18. Effects of water stress on photosynthetic electron transport, photophosphorylation, and metabolite levels of Xanthium strumarium mesophyll cells.

    Sharkey, T D; Badger, M R

    1982-12-01

    Several component processes of photosynthesis were measured in osmotically stressed mesophyll cells of Xanthium strumarium L. The ribulose-1,5-bisphosphate regeneration capacity was reduced by water stress. Photophoshorylation was sensitive to water stress but photosynthetic electron transport was unaffected by water potentials down to-40 bar (-4 MPa). The concentrations of several intermediates of the photosynthetic carbon-reduction cycle remained relatively constant and did not indicate that ATP supply was limiting photosynthesis in the water-stressed cells.

  19. Ultrastructural response of cabbage outer leaf mesophyll cells (Brassica oleracea L. to excess of nickel

    Jolanta Molas

    2014-01-01

    Full Text Available Changes in the structure and in the ultrastructure of cabbage outer leaf mesophyll cells [Brassica oleracea L.] cv. Sława from Enkhouizen were examined by means of light and electron microscopy. The examined plants were grown on the basic Murashige and Skoog medium with addition of excesive concentrations of nickel (added as NiSO4 x 7H2O,i.e. Ni 5, Ni 10 and Ni 20 mg/dm3. In Ni 5 mg samples mainly adaptation changes to the conditions of stress were observed. These changes were manifested by the increase of cytoplasm content and by cytoplasm vacuolization, by the increase of nucleus and nucleous volume, nucleolus vacuolization, the increase of plasmalemma invaginations and of the amount of rough ER, by the central arrangement of smooth ER and of the thylakoids of chloroplasts; it was also shown by the growth of the number of mitochondria and of peroxisomes in the cell. In Ni 10 mg samples, apart from adaptation changes, such as the increase of the nucleus volume, increase of plasmalemma invaginations, cytoplasm and nucleolus vacuolization, degeneration changes were also observed. They concerned mainly the nucleus (the increasing amount of condensed chromatin, ER (swelling and fragmentation of rER and sER, mitochondrium (swelling and reduction of cristae, Golgi apparatus (disintegration and decay and chloroplasts (changes of shape, swelling and reduction of thylakoids, disappearance of starch and presence of big plastoglobuli. In Ni 20 mg samples cell protoplasts were in different stages of degeneration and the cell organelles that were identifiable, were usually damaged.

  20. Inorganic carbon uptake during photosynthesis. II. Uptake by isolated Asparagus mesophyll cells during isotopic disequilibrium

    Espie, G.S.; Owttrim, G.W.; Colman, B.

    1986-01-01

    The species of inorganic carbon (CO 2 or HCO 3 - ) taken up as a source of substrate for photosynthetic fixation by isolated Asparagus sprengeri mesophyll cells is investigated. Discrimination between CO 2 or HCO 3 - transport, during steady state photosynthesis, is achieved by monitoring the changes (by 14 C fixation) which occur in the specific activity of the intracellular pool of inorganic carbon when the inorganic carbon present in the suspending medium is in a state of isotopic disequilibrium. Quantitative comparisons between theoretical (CO 2 or HCO 3 - transport) and experimental time-courses of 14 C incorporation, over the pH range of 5.2 to 7.5, indicate that the specific activity of extracellular CO 2 , rather than HCO 3 - , is the appropriate predictor of the intracellular specific activity. It is concluded, therefore, that CO 2 is the major source of exogenous inorganic carbon taken up by Asparagus cells. However, at high pH (8.5), a component of net DIC uptake may be attributable to HCO 3 - transport, as the incorporation of 14 C during isotopic disequilibrium exceeds the maximum possible incorporation predicted on the basis of CO 2 uptake alone. The contribution of HCO 3 - to net inorganic carbon uptake (pH 8.5) is variable, ranging from 5 to 16%, but is independent of the extracellular HCO 3 - concentration. The evidence for direct HCO 3 - transport is subject to alternative explanations and must, therefore, be regarded as equivocal. Nonlinear regression analysis of the rate of 14 C incorporation as a function of time indicates the presence of a small extracellular resistance to the diffusion of CO 2 , which is partially alleviated by a high extracellular concentration of HCO 3 -

  1. Stomatal responses to flooding of the intercellular air spaces suggest a vapor-phase signal between the mesophyll and the guard cells.

    Sibbernsen, Erik; Mott, Keith A

    2010-07-01

    Flooding the intercellular air spaces of leaves with water was shown to cause rapid closure of stomata in Tradescantia pallida, Lactuca serriola, Helianthus annuus, and Oenothera caespitosa. The response occurred when water was injected into the intercellular spaces, vacuum infiltrated into the intercellular spaces, or forced into the intercellular spaces by pressurizing the xylem. Injecting 50 mm KCl or silicone oil into the intercellular spaces also caused stomata to close, but the response was slower than with distilled water. Epidermis-mesophyll grafts for T. pallida were created by placing the epidermis of one leaf onto the exposed mesophyll of another leaf. Stomata in these grafts opened under light but closed rapidly when water was allowed to wick between epidermis and the mesophyll. When epidermis-mesophyll grafts were constructed with a thin hydrophobic filter between the mesophyll and epidermis stomata responded normally to light and CO(2). These data, when taken together, suggest that the effect of water on stomata is caused partly by dilution of K(+) in the guard cell and partly by the existence of a vapor-phase signal that originates in the mesophyll and causes stomata to open in the light.

  2. Stomatal Responses to Flooding of the Intercellular Air Spaces Suggest a Vapor-Phase Signal Between the Mesophyll and the Guard Cells1[OA

    Sibbernsen, Erik; Mott, Keith A.

    2010-01-01

    Flooding the intercellular air spaces of leaves with water was shown to cause rapid closure of stomata in Tradescantia pallida, Lactuca serriola, Helianthus annuus, and Oenothera caespitosa. The response occurred when water was injected into the intercellular spaces, vacuum infiltrated into the intercellular spaces, or forced into the intercellular spaces by pressurizing the xylem. Injecting 50 mm KCl or silicone oil into the intercellular spaces also caused stomata to close, but the response was slower than with distilled water. Epidermis-mesophyll grafts for T. pallida were created by placing the epidermis of one leaf onto the exposed mesophyll of another leaf. Stomata in these grafts opened under light but closed rapidly when water was allowed to wick between epidermis and the mesophyll. When epidermis-mesophyll grafts were constructed with a thin hydrophobic filter between the mesophyll and epidermis stomata responded normally to light and CO2. These data, when taken together, suggest that the effect of water on stomata is caused partly by dilution of K+ in the guard cell and partly by the existence of a vapor-phase signal that originates in the mesophyll and causes stomata to open in the light. PMID:20472750

  3. Consequences of C4 differentiation for chloroplast membrane proteomes in maize mesophyll and bundle sheath cells.

    Majeran, Wojciech; Zybailov, Boris; Ytterberg, A Jimmy; Dunsmore, Jason; Sun, Qi; van Wijk, Klaas J

    2008-09-01

    Chloroplasts of maize leaves differentiate into specific bundle sheath (BS) and mesophyll (M) types to accommodate C(4) photosynthesis. Chloroplasts contain thylakoid and envelope membranes that contain the photosynthetic machineries and transporters but also proteins involved in e.g. protein homeostasis. These chloroplast membranes must be specialized within each cell type to accommodate C(4) photosynthesis and regulate metabolic fluxes and activities. This quantitative study determined the differentiated state of BS and M chloroplast thylakoid and envelope membrane proteomes and their oligomeric states using innovative gel-based and mass spectrometry-based protein quantifications. This included native gels, iTRAQ, and label-free quantification using an LTQ-Orbitrap. Subunits of Photosystems I and II, the cytochrome b(6)f, and ATP synthase complexes showed average BS/M accumulation ratios of 1.6, 0.45, 1.0, and 1.33, respectively, whereas ratios for the light-harvesting complex I and II families were 1.72 and 0.68, respectively. A 1000-kDa BS-specific NAD(P)H dehydrogenase complex with associated proteins of unknown function containing more than 15 proteins was observed; we speculate that this novel complex possibly functions in inorganic carbon concentration when carboxylation rates by ribulose-bisphosphate carboxylase/oxygenase are lower than decarboxylation rates by malic enzyme. Differential accumulation of thylakoid proteases (Egy and DegP), state transition kinases (STN7,8), and Photosystem I and II assembly factors was observed, suggesting that cell-specific photosynthetic electron transport depends on post-translational regulatory mechanisms. BS/M ratios for inner envelope transporters phosphoenolpyruvate/P(i) translocator, Dit1, Dit2, and Mex1 were determined and reflect metabolic fluxes in carbon metabolism. A wide variety of hundreds of other proteins showed differential BS/M accumulation. Mass spectral information and functional annotations are

  4. Early local differentiation of the cell wall matrix defines the contact sites in lobed mesophyll cells of Zea mays.

    Giannoutsou, E; Sotiriou, P; Apostolakos, P; Galatis, B

    2013-10-01

    The morphogenesis of lobed mesophyll cells (MCs) is highly controlled and coupled with intercellular space formation. Cortical microtubule rings define the number and the position of MC isthmi. This work investigated early events of MC morphogenesis, especially the mechanism defining the position of contacts between MCs. The distributions of plasmodesmata, the hemicelluloses callose and (1 → 3,1 → 4)-β-d-glucans (MLGs) and the pectin epitopes recognized by the 2F4, JIM5, JIM7 and LM6 antibodies were studied in the cell walls of Zea mays MCs. Matrix cell wall polysaccharides were immunolocalized in hand-made sections and in sections of material embedded in LR White resin. Callose was also localized using aniline blue in hand-made sections. Plasmodesmata distribution was examined by transmission electron microscopy. Before reorganization of the dispersed cortical microtubules into microtubule rings, particular bands of the longitudinal MC walls, where the MC contacts will form, locally differentiate by selective (1) deposition of callose and the pectin epitopes recognized by the 2F4, LM6, JIM5 and JIM7 antibodies, (2) degradation of MLGs and (3) formation of secondary plasmodesmata clusterings. This cell wall matrix differentiation persists in cell contacts of mature MCs. Simultaneously, the wall bands between those of future cell contacts differentiate with (1) deposition of local cell wall thickenings including cellulose microfibrils, (2) preferential presence of MLGs, (3) absence of callose and (4) transient presence of the pectins identified by the JIM5 and JIM7 antibodies. The wall areas between cell contacts expand determinately to form the cell isthmi and the cell lobes. The morphogenesis of lobed MCs is characterized by the early patterned differentiation of two distinct cell wall subdomains, defining the sites of the future MC contacts and of the future MC isthmi respectively. This patterned cell wall differentiation precedes cortical microtubule

  5. Kinetics of Ca2+- and ATP-dependent, voltage-controlled anion conductance in the plasma membrane of mesophyll cells of Pisum sativum

    Elzenga, J.T.M.; van Volkenburgh, E.

    Whole-cell patch-clamp techniques were used to measure anion currents through the plasma membrane of protoplasts of mesophyll cells of expanding pea (Pisum sativum L.) leaves. Voltage-induced changes of the currents could be modelled with single exponential activation and deactivation kinetics. The

  6. A new mechanism for the regulation of stomatal aperture size in intact leaves: accumulation of mesophyll-derived sucrose in the guard-cell wall of Vicia faba

    Lu, P.; Outlaw, W.H. Jr.; Smith, B.G.; Freed, G.A.

    1997-01-01

    At various times after pulse-labeling broad bean (Vicia faba L.) leaflets with 14CO2, whole-leaf pieces and rinsed epidermal peels were harvested and subsequently processed for histochemical analysis. Cells dissected from whole leaf retained apoplastic contents, whereas those from rinsed peels contained only symplastic contents. Sucrose (Suc)-specific radioactivity peaked (111 GBq mol-1) in palisade cells at 20 min. In contrast, the 14C content and Suc-specific radioactivity were very low in guard cells for 20 min, implying little CO2 incorporation; both then peaked at 40 min. The guard-cell apoplast had a high maximum Suc-specific radioactivity (204 GBq mol-1) and a high Suc influx rate (0.05 pmol stoma-1 min-1). These and other comparisons implied the presence of (a) multiple Suc pools in mesophyll cells, (b) a localized mesophyll-apoplast region that exchanges with phloem and stomata, and (c) mesophyll-derived Suc in guard-cell walls sufficient to diminish stomatal opening by approximately 3 micrometers. Factors expected to enhance Suc accumulation in guard-cell walls are (a) high transpiration rate, which closes stomata, and (b) high apoplastic Suc concentration, which is elevated when mesophyll Suc efflux exceeds translocation. Therefore, multiple physiological factors are integrated in the attenuation of stomatal aperture size by this previously unrecognized mechanism

  7. Intracellular position of mitochondria and chloroplasts in bundle sheath and mesophyll cells of C3 grasses in relation to photorespiratory CO2 loss

    Yuto Hatakeyama

    2016-10-01

    Full Text Available In C3 plants, photosynthetic efficiency is reduced by photorespiration. A part of CO2 fixed during photosynthesis in chloroplasts is lost from mitochondria during photorespiration by decarboxylation of glycine by glycine decarboxylase (GDC. Thus, the intracellular position of mitochondria in photosynthetic cells is critical to the rate of photorespiratory CO2 loss. We investigated the intracellular position of mitochondria in parenchyma sheath (PS and mesophyll cells of 10 C3 grasses from 3 subfamilies (Ehrhartoideae, Panicoideae, and Pooideae by immunostaining for GDC and light and electron microscopic observation. Immunostaining suggested that many mitochondria were located in the inner half of PS cells and on the vacuole side of chloroplasts in mesophyll cells. Organelle quantification showed that 62–75% of PS mitochondria were located in the inner half of cells, and 62–78% of PS chloroplasts were in the outer half. In mesophyll cells, 61–92% of mitochondria were positioned on the vacuole side of chloroplasts and stromules. In PS cells, such location would reduce the loss of photorespiratory CO2 by lengthening the path of CO2 diffusion and allow more efficient fixation of CO2 from intercellular spaces. In mesophyll cells, it would facilitate scavenging by chloroplasts of photorespiratory CO2 released from mitochondria. Our data suggest that the PS cells of C3 grasses have already acquired an initial structure leading to proto-Kranz and further C3–C4 intermediate anatomy. We also found that in the Pooideae, organelle positioning in PS cells on the phloem side resembles that in mesophyll cells.

  8. Cyst(e)ine Is the Transport Metabolite of Assimilated Sulfur from Bundle-Sheath to Mesophyll Cells in Maize Leaves1

    Burgener, Marta; Suter, Marianne; Jones, Stephanie; Brunold, Christian

    1998-01-01

    The intercellular distribution of the enzymes and metabolites of assimilatory sulfate reduction and glutathione synthesis was analyzed in maize (Zea mays L. cv LG 9) leaves. Mesophyll cells and strands of bundle-sheath cells from second leaves of 11-d-old maize seedlings were obtained by two different mechanical-isolation methods. Cross-contamination of cell preparations was determined using ribulose bisphosphate carboxylase (EC 4.1.1.39) and nitrate reductase (EC 1.6.6.1) as marker enzymes for bundle-sheath and mesophyll cells, respectively. ATP sulfurylase (EC 2.7.7.4) and adenosine 5′-phosphosulfate sulfotransferase activities were detected almost exclusively in the bundle-sheath cells, whereas GSH synthetase (EC 6.3.2.3) and cyst(e)ine, γ-glutamylcysteine, and glutathione were located predominantly in the mesophyll cells. Feeding experiments using [35S]sulfate with intact leaves indicated that cyst(e)ine was the transport metabolite of reduced sulfur from bundle-sheath to mesophyll cells. This result was corroborated by tracer experiments, which showed that isolated bundle-sheath strands fed with [35S]sulfate secreted radioactive cyst(e)ine as the sole thiol into the resuspending medium. The results presented in this paper show that assimilatory sulfate reduction is restricted to the bundle-sheath cells, whereas the formation of glutathione takes place predominantly in the mesophyll cells, with cyst(e)ine functioning as a transport metabolite between the two cell types. PMID:9536048

  9. Glutamine deficiency induces DNA alkylation damage and sensitizes cancer cells to alkylating agents through inhibition of ALKBH enzymes.

    Tran, Thai Q; Ishak Gabra, Mari B; Lowman, Xazmin H; Yang, Ying; Reid, Michael A; Pan, Min; O'Connor, Timothy R; Kong, Mei

    2017-11-01

    Driven by oncogenic signaling, glutamine addiction exhibited by cancer cells often leads to severe glutamine depletion in solid tumors. Despite this nutritional environment that tumor cells often experience, the effect of glutamine deficiency on cellular responses to DNA damage and chemotherapeutic treatment remains unclear. Here, we show that glutamine deficiency, through the reduction of alpha-ketoglutarate, inhibits the AlkB homolog (ALKBH) enzymes activity and induces DNA alkylation damage. As a result, glutamine deprivation or glutaminase inhibitor treatment triggers DNA damage accumulation independent of cell death. In addition, low glutamine-induced DNA damage is abolished in ALKBH deficient cells. Importantly, we show that glutaminase inhibitors, 6-Diazo-5-oxo-L-norleucine (DON) or CB-839, hypersensitize cancer cells to alkylating agents both in vitro and in vivo. Together, the crosstalk between glutamine metabolism and the DNA repair pathway identified in this study highlights a potential role of metabolic stress in genomic instability and therapeutic response in cancer.

  10. Cell wall matrix polysaccharide distribution and cortical microtubule organization: two factors controlling mesophyll cell morphogenesis in land plants.

    Sotiriou, P; Giannoutsou, E; Panteris, E; Apostolakos, P; Galatis, B

    2016-03-01

    This work investigates the involvement of local differentiation of cell wall matrix polysaccharides and the role of microtubules in the morphogenesis of mesophyll cells (MCs) of three types (lobed, branched and palisade) in the dicotyledon Vigna sinensis and the fern Asplenium nidus. Homogalacturonan (HGA) epitopes recognized by the 2F4, JIM5 and JIM7 antibodies and callose were immunolocalized in hand-made leaf sections. Callose was also stained with aniline blue. We studied microtubule organization by tubulin immunofluorescence and transmission electron microscopy. In both plants, the matrix cell wall polysaccharide distribution underwent definite changes during MC differentiation. Callose constantly defined the sites of MC contacts. The 2F4 HGA epitope in V. sinensis first appeared in MC contacts but gradually moved towards the cell wall regions facing the intercellular spaces, while in A. nidus it was initially localized at the cell walls delimiting the intercellular spaces, but finally shifted to MC contacts. In V. sinensis, the JIM5 and JIM7 HGA epitopes initially marked the cell walls delimiting the intercellular spaces and gradually shifted in MC contacts, while in A. nidus they constantly enriched MC contacts. In all MC types examined, the cortical microtubules played a crucial role in their morphogenesis. In particular, in palisade MCs, cortical microtubule helices, by controlling cellulose microfibril orientation, forced these MCs to acquire a truncated cone-like shape. Unexpectedly in V. sinensis, the differentiation of colchicine-affected MCs deviated completely, since they developed a cell wall ingrowth labyrinth, becoming transfer-like cells. The results of this work and previous studies on Zea mays (Giannoutsou et al., Annals of Botany 2013; 112: : 1067-1081) revealed highly controlled local cell wall matrix differentiation in MCs of species belonging to different plant groups. This, in coordination with microtubule-dependent cellulose microfibril

  11. Determination of glutamine and glutamic acid in mammalian cell cultures using tetrathiafulvalene modified enzyme electrodes.

    Mulchandani, A; Bassi, A S

    1996-01-01

    Tetrathiafulvalene (TTF) mediated amperometric enzyme electrodes have been developed for the monitoring of L-glutamine and L-glutamic acid in growing mammalian cell cultures. The detection of glutamine was accomplished by a coupled enzyme system comprised of glutaminase plus glutamate oxidase, while the detection of glutamic acid was carried out by a single enzyme, glutamate oxidase. The appropriate enzyme(s) were immoblized on the Triton-X treated surface of tetrathiafulvalene modified carbon paste electrodes by adsorption, in conjunction with entrapment by an electrochemically deposited copolymer film of 1,3-phenylenediamine and resorcinol. Operating conditions for the glutamine enzyme electrode were optimized with respect to the amount of enzymes immoblized, pH, temperature and mobile phase flow rate for operation in a flow injection (FIA) system. When applied to glutamine and glutamic acid measurements in mammalian cell culture in FIA, the results obtained with enzyme electrodes were in excellent agreement with those determined by enzymatic analysis.

  12. Regional tumour glutamine supply affects chromatin and cell identity

    Højfeldt, Jonas W; Helin, Kristian

    2016-01-01

    Limited perfusion of solid tumours produces a nutrient-deprived tumour core microenvironment. Low glutamine levels in the tumour core are now shown to lead to reduced levels of α-ketoglutarate and decreased histone demethylase activity, thereby promoting a less differentiated and more therapy-res...

  13. Mitochondrial Sirt3 supports cell proliferation by regulating glutamine-dependent oxidation in renal cell carcinoma

    Choi, Jieun; Koh, Eunjin; Lee, Yu Shin; Lee, Hyun-Woo; Kang, Hyeok Gu; Yoon, Young Eun; Han, Woong Kyu; Choi, Kyung Hwa; Kim, Kyung-Sup

    2016-01-01

    Clear cell renal carcinoma (RCC), the most common malignancy arising in the adult kidney, exhibits increased aerobic glycolysis and low mitochondrial respiration due to von Hippel-Lindau gene defects and constitutive hypoxia-inducible factor-α expression. Sirt3 is a major mitochondrial deacetylase that mediates various types of energy metabolism. However, the role of Sirt3 as a tumor suppressor or oncogene in cancer depends on cell types. We show increased Sirt3 expression in the mitochondrial fraction of human RCC tissues. Sirt3 depletion by lentiviral short-hairpin RNA, as well as the stable expression of the inactive mutant of Sirt3, inhibited cell proliferation and tumor growth in xenograft nude mice, respectively. Furthermore, mitochondrial pyruvate, which was used for oxidation in RCC, might be derived from glutamine, but not from glucose and cytosolic pyruvate, due to depletion of mitochondrial pyruvate carrier and the relatively high expression of malic enzyme 2. Depletion of Sirt3 suppressed glutamate dehydrogenase activity, leading to impaired mitochondrial oxygen consumption. Our findings suggest that Sirt3 plays a tumor-progressive role in human RCC by regulating glutamine-derived mitochondrial respiration, particularly in cells where mitochondrial usage of cytosolic pyruvate is severely compromised. -- Highlights: •Sirt3 is required for the maintenance of RCC cell proliferation. •Mitochondrial usage of cytosolic pyruvate is severely compromised in RCC. •Sirt3 supports glutamine-dependent oxidation in RCC.

  14. Mitochondrial Sirt3 supports cell proliferation by regulating glutamine-dependent oxidation in renal cell carcinoma

    Choi, Jieun; Koh, Eunjin; Lee, Yu Shin; Lee, Hyun-Woo; Kang, Hyeok Gu [Department of Biochemistry and Molecular Biology, Brain Korea 21 PLUS Project for Medical Sciences, Institute of Genetic Science, Integrated Genomic Research Center for Metabolic Regulation, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Yoon, Young Eun; Han, Woong Kyu [Department of Urology, Urological Science Institute, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of); Choi, Kyung Hwa [Department of Urology, CHA Bundang Medical Center, CHA University, Seongnam 463-712 (Korea, Republic of); Kim, Kyung-Sup, E-mail: KYUNGSUP59@yuhs.ac [Department of Biochemistry and Molecular Biology, Brain Korea 21 PLUS Project for Medical Sciences, Institute of Genetic Science, Integrated Genomic Research Center for Metabolic Regulation, Yonsei University College of Medicine, Seoul 120-752 (Korea, Republic of)

    2016-06-03

    Clear cell renal carcinoma (RCC), the most common malignancy arising in the adult kidney, exhibits increased aerobic glycolysis and low mitochondrial respiration due to von Hippel-Lindau gene defects and constitutive hypoxia-inducible factor-α expression. Sirt3 is a major mitochondrial deacetylase that mediates various types of energy metabolism. However, the role of Sirt3 as a tumor suppressor or oncogene in cancer depends on cell types. We show increased Sirt3 expression in the mitochondrial fraction of human RCC tissues. Sirt3 depletion by lentiviral short-hairpin RNA, as well as the stable expression of the inactive mutant of Sirt3, inhibited cell proliferation and tumor growth in xenograft nude mice, respectively. Furthermore, mitochondrial pyruvate, which was used for oxidation in RCC, might be derived from glutamine, but not from glucose and cytosolic pyruvate, due to depletion of mitochondrial pyruvate carrier and the relatively high expression of malic enzyme 2. Depletion of Sirt3 suppressed glutamate dehydrogenase activity, leading to impaired mitochondrial oxygen consumption. Our findings suggest that Sirt3 plays a tumor-progressive role in human RCC by regulating glutamine-derived mitochondrial respiration, particularly in cells where mitochondrial usage of cytosolic pyruvate is severely compromised. -- Highlights: •Sirt3 is required for the maintenance of RCC cell proliferation. •Mitochondrial usage of cytosolic pyruvate is severely compromised in RCC. •Sirt3 supports glutamine-dependent oxidation in RCC.

  15. Early H2O2 Accumulation in Mesophyll Cells Leads to Induction of Glutathione during the Hyper-Sensitive Response in the Barley-Powdery Mildew Interaction1

    Vanacker, Helene; Carver, Tim L.W.; Foyer, Christine H.

    2000-01-01

    H2O2 production and changes in glutathione, catalase, and peroxidase were followed in whole-leaf extracts from the susceptible (AlgS [Algerian/4* (F14) Man.(S)]; ml-a1 allele) and resistant (AlgR [Algerian/4* (F14) Man.(R)]; Ml-a1 allele) barley (Hordeum vulgare) isolines between 12 and 24 h after inoculation with powdery mildew (Blumeria graminis [DC]. Speer [syn. Erysiphe graminis DC] f.sp hordei Marchal). Localized papilla responses and cell death hypersensitive responses were not observed within the same cell. In hypersensitive response sites, H2O2 accumulation first occurred in the mesophyll underlying the attacked epidermal cell. Subsequently, H2O2 disappeared from the mesophyll and accumulated around attacked epidermal cells. In AlgR, transient glutathione oxidation coincided with H2O2 accumulation in the mesophyll. Subsequently, total foliar glutathione and catalase activities transiently increased in AlgR. These changes, absent from AlgS, preceded inoculation-dependent increases in peroxidase activity that were observed in both AlgR and AlgS at 18 h. An early intercellular signal precedes H2O2, and this elicits anti-oxidant responses in leaves prior to events leading to death of attacked cells. PMID:10938348

  16. Nonnutritive effects of glutamine.

    Roth, Erich

    2008-10-01

    Glutamine is the most abundant free amino acid of the human body. Besides its role as a constituent of proteins and its importance in amino acid transamination, glutamine has regulatory capacity in immune and cell modulation. Glutamine deprivation reduces proliferation of lymphocytes, influences expression of surface activation markers on lymphocytes and monocytes, affects the production of cytokines, and stimulates apoptosis. Moreover, glutamine administration seems to have a positive effect on glucose metabolism in the state of insulin resistance. Glutamine influences a variety of different molecular pathways. Glutamine stimulates the formation of heat shock protein 70 in monocytes by enhancing the stability of mRNA, influences the redox potential of the cell by enhancing the formation of glutathione, induces cellular anabolic effects by increasing the cell volume, activates mitogen-activated protein kinases, and interacts with particular aminoacyl-transfer RNA synthetases in specific glutamine-sensing metabolism. Glutamine is applied under clinical conditions as an oral, parenteral, or enteral supplement either as the single amino acid or in the form of glutamine-containing dipeptides for preventing mucositis/stomatitis and for preventing glutamine-deficiency in critically ill patients. Because of the high turnover rate of glutamine, even high amounts of glutamine up to a daily administration of 30 g can be given without any important side effects.

  17. Effects of herbicides on /sup 14/CO/sub 2/ fixation in isolated mesophyll cells from Beta vulgaris (sugar beet) and Chenopodium album

    Baumann, G; Guenther, G [Paedagogische Hochschule Karl Liebknecht, Potsdam (German Democratic Republic)

    1979-01-01

    10/sup -4/ - 10/sup -6/ molar solutions of herbicides (atrazine, 2,4-D, desmetryne, diallate, diquat, feuron, lenacil, NaTa, paraquat, phenmedipham, prometryne, propham, pyrazone, and simazine) cause similar inhibitory effects on the photosynthetic /sup 14/CO/sub 2/ fixation in isolated mesophyll cells from Chenopodium album and Beta vulgaris. Correlatdion between inhibition and herbicide resistance of the whole plants could be realized for lenacil only.

  18. Identification of the TaBTF3 gene in wheat (Triticum aestivum L.) and the effect of its silencing on wheat chloroplast, mitochondria and mesophyll cell development.

    Ma, Hong-Zhen; Liu, Guo-Qin; Li, Cheng-Wei; Kang, Guo-Zhang; Guo, Tian-Cai

    2012-10-05

    The full-length cDNA (882bp) and DNA (1742bp) sequences encoding a basic transcription factor 3, designated as TaBTF3, were first isolated from common wheat (Triticum aestivum L.). Subcellular localization studies revealed that the TaBTF3 protein was mainly located in the cytoplasm and nucleus. In TaBTF3-silenced transgenic wheat seedlings obtained using the Virus-induced gene silencing (VIGS) method, the chlorophyll pigment content was markedly reduced. However, the malonaldehyde (MDA) and H(2)O(2) contents were enhanced, and the structure of the wheat mesophyll cell was seriously damaged. Furthermore, transcripts of the chloroplast- and mitochondrial-encoded genes were significantly reduced in TaBTF3-silenced transgenic wheat plants. These results suggest that the TaBTF3 gene might function in the development of the wheat chloroplast, mitochondria and mesophyll cell. This paper is the first report to describe the involvement of TaBTF3 in maintaining the normal plant mesophyll cell structure. Copyright © 2012 Elsevier Inc. All rights reserved.

  19. Intracellular compartimentation of abscisic acid (ABA) in guard cells and mesophyll cells under exposure to SO sub 2. Kompartimentierung von Abscisinsaeure (ABA) in Schliess- und Mesophyllzellen unter SO sub 2 -Belastung

    Baier, M.; Daeter, W.; Hartung, W. (Wuerzburg Univ. (Germany, F.R.). Lehrstuhl fuer Botanik 1)

    1989-07-01

    The effect of SO{sub 2} on the intracellular compartimentation of ABA in guard cells and mesophyll cells of Valerianella locusta was investigated, using the efflux compartmental analysis, as described by Behl and Hartung (1986). The cytoplasmic ABA content of the guard cells was reduced drastically by 6 {mu}molxm{sup -3} SO{sub 2} (20% of the controls). The vacuolar content was decreased less dramatically (70% of the controls). The ABA distribution of mesophyll cells remained uneffected by 6 {mu}molxm{sup -3} SO{sub 2}. The SO{sub 2} effects are explained by an acidification of the compartments. (orig.).

  20. Structural characterization of a mixed-linkage glucan deficient mutant reveals alteration in cellulose microfibril orientation in rice coleoptile mesophyll cell walls

    Andreia Michelle Smith-Moritz

    2015-08-01

    Full Text Available The CELLULOSE SYNTHASE-LIKE F6 (CslF6 gene was previously shown to mediate the biosynthesis of mixed-linkage glucan (MLG, a cell wall polysaccharide that is hypothesized to be a tightly associated with cellulose and also have a role in cell expansion in the primary cell wall of young seedlings in grass species. We have recently shown that loss-of-function cslf6 rice mutants do not accumulate MLG in most vegetative tissues. Despite the absence of a structurally important polymer, MLG, these mutants are unexpectedly viable and only show a moderate growth compromise compared to wild type. Therefore these mutants are ideal biological systems to test the current grass cell wall model. In order to gain a better understanding of the role of MLG in the primary wall, we performed in-depth compositional and structural analyses of the cell walls of three day-old rice seedlings using various biochemical and novel microspectroscopic approaches. We found that cellulose content as well as matrix polysaccharide composition was not significantly altered in the MLG deficient mutant. However, we observed a significant change in cellulose microfibril bundle organization in mesophyll cell walls of the cslf6 mutant. Using synchrotron source Fourier Transform Mid-Infrared Spectromicroscopy for high-resolution imaging, we determined that the bonds associated with cellulose and arabinoxylan, another major component of the primary cell was of grasses, were in a lower energy configuration compared to wild type, suggesting a slightly weaker primary wall in MLG deficient mesophyll cells. Taken together, these results suggest that MLG may influence cellulose deposition in mesophyll cell walls without significantly affecting anisotropic growth thus challenging MLG importance in cell wall expansion.

  1. PDHA1 gene knockout in prostate cancer cells results in metabolic reprogramming towards greater glutamine dependence

    Li, Yaqing; Li, Xiaoran; Li, Xiaoli; Zhong, Yali; Ji, Yasai; Yu, Dandan; Zhang, Mingzhi; Wen, Jian-Guo; Zhang, Hongquan; Goscinski, Mariusz Adam; Nesland, Jahn M.; Suo, Zhenhe

    2016-01-01

    Alternative pathways of metabolism endowed cancer cells with metabolic stress. Inhibiting the related compensatory pathways might achieve synergistic anticancer results. This study demonstrated that pyruvate dehydrogenase E1α gene knockout (PDHA1 KO) resulted in alterations in tumor cell metabolism by rendering the cells with increased expression of glutaminase1 (GLS1) and glutamate dehydrogenase1 (GLUD1), leading to an increase in glutamine-dependent cell survival. Deprivation of glutamine induced cell growth inhibition, increased reactive oxygen species and decreased ATP production. Pharmacological blockade of the glutaminolysis pathway resulted in massive tumor cells apoptosis and dysfunction of ROS scavenge in the LNCaP PDHA1 KO cells. Further examination of the key glutaminolysis enzymes in human prostate cancer samples also revealed that higher levels of GLS1 and GLUD1 expression were significantly associated with aggressive clinicopathological features and poor clinical outcome. These insights supply evidence that glutaminolysis plays a compensatory role for cell survival upon alternative energy metabolism and targeting the glutamine anaplerosis of energy metabolism via GLS1 and GLUD1 in cancer cells may offer a potential novel therapeutic strategy. PMID:27462778

  2. Glutamine-derived 2-hydroxyglutarate is associated with disease progression in plasma cell malignancies

    Gonsalves, Wilson I.; Hitosugi, Taro; Ghosh, Toshi; Jevremovic, Dragan; Petterson, Xuan-Mai; Wellik, Linda; Kumar, Shaji K.; Nair, K. Sreekumaran

    2018-01-01

    The production of the oncometabolite 2-hydroxyglutarate (2-HG) has been associated with c-MYC overexpression. c-MYC also regulates glutamine metabolism and drives progression of asymptomatic precursor plasma cell (PC) malignancies to symptomatic multiple myeloma (MM). However, the presence of 2-HG and its clinical significance in PC malignancies is unknown. By performing 13C stable isotope resolved metabolomics (SIRM) using U[13C6]Glucose and U[13C5]Glutamine in human myeloma cell lines (HMCLs), we show that 2-HG is produced in clonal PCs and is derived predominantly from glutamine anaplerosis into the TCA cycle. Furthermore, the 13C SIRM studies in HMCLs also demonstrate that glutamine is preferentially utilized by the TCA cycle compared with glucose. Finally, measuring the levels of 2-HG in the BM supernatant and peripheral blood plasma from patients with precursor PC malignancies such as smoldering MM (SMM) demonstrates that relatively elevated levels of 2-HG are associated with higher levels of c-MYC expression in the BM clonal PCs and with a subsequent shorter time to progression (TTP) to MM. Thus, measuring 2-HG levels in BM supernatant or peripheral blood plasma of SMM patients offers potential early identification of those patients at high risk of progression to MM, who could benefit from early therapeutic intervention. PMID:29321378

  3. Addiction to Coupling of the Warburg Effect with Glutamine Catabolism in Cancer Cells

    Bradley Smith

    2016-10-01

    Full Text Available Metabolic reprogramming is critical to oncogenesis, but the emergence and function of this profound reorganization remain poorly understood. Here we find that cooperating oncogenic mutations drive large-scale metabolic reprogramming, which is both intrinsic to cancer cells and obligatory for the transition to malignancy. This involves synergistic regulation of several genes encoding metabolic enzymes, including the lactate dehydrogenases LDHA and LDHB and mitochondrial glutamic pyruvate transaminase 2 (GPT2. Notably, GPT2 engages activated glycolysis to drive the utilization of glutamine as a carbon source for TCA cycle anaplerosis in colon cancer cells. Our data indicate that the Warburg effect supports oncogenesis via GPT2-mediated coupling of pyruvate production to glutamine catabolism. Although critical to the cancer phenotype, GPT2 activity is dispensable in cells that are not fully transformed, thus pinpointing a metabolic vulnerability specifically associated with cancer cell progression to malignancy.

  4. Glutamine deficiency induces DNA alkylation damage and sensitizes cancer cells to alkylating agents through inhibition of ALKBH enzymes.

    Thai Q Tran

    2017-11-01

    Full Text Available Driven by oncogenic signaling, glutamine addiction exhibited by cancer cells often leads to severe glutamine depletion in solid tumors. Despite this nutritional environment that tumor cells often experience, the effect of glutamine deficiency on cellular responses to DNA damage and chemotherapeutic treatment remains unclear. Here, we show that glutamine deficiency, through the reduction of alpha-ketoglutarate, inhibits the AlkB homolog (ALKBH enzymes activity and induces DNA alkylation damage. As a result, glutamine deprivation or glutaminase inhibitor treatment triggers DNA damage accumulation independent of cell death. In addition, low glutamine-induced DNA damage is abolished in ALKBH deficient cells. Importantly, we show that glutaminase inhibitors, 6-Diazo-5-oxo-L-norleucine (DON or CB-839, hypersensitize cancer cells to alkylating agents both in vitro and in vivo. Together, the crosstalk between glutamine metabolism and the DNA repair pathway identified in this study highlights a potential role of metabolic stress in genomic instability and therapeutic response in cancer.

  5. Effect of Hf on the fine structure of mesophyll cells from Glycine max, Merr

    Wei, L.; Miller, G.W.

    1972-04-01

    A series of ultrastructural changes were observed in soybean leaves fumigated with 40 to 50 ppb of hydrogen fluoride. In the cytoplasm the presence of small vacuoles was the first noticeable initial change. The fragmentation of the vacuolar membrane occurred either simultaneously or followed immediately. Lipid-droplet-like globules and numerous vesicles occurred subsequently in the cytoplasm and increased as the injury became more severe. There was a decrease in polysomes and a detachment of ribosome from the rough endoplasmic reticulum. Free ribosome concentration also decreased as the injury became severe. Mitochondrial modifications involving dilation of outer and cristae membranes followed by reduction of both cristae number and matrix electron density and the disappearance of mitochondrial granules were observed in the chlorotic leaves. Electron dense inclusions accumulated in some mitochondria as well. The first noticeable change observed in the chloroplast was the presence of clusters of phytoferritin granules within the stoma after only 2 days of fumigation. Alterations in nuclear structures were observed in later stages of injury. Numerous small electron dense particles were found on various types of membranes in cells of severely chlorotic leaves. They were distributed on outer mitochondrial membranes, endoplasmic reticula, dictyosomes, tonoplasts, plasmalemma, nuclear envelopes, and disintegrating organelles and vesicles, but were never observed on membranes of chloroplasts and microbodies. The presence of fluoride has attracted the attention of many workers primarily in certain industrial areas where the emitted atmospheric fluoride concentrates and is accumulated by plants initiating injury. 6 references.

  6. Increasing Leaf Vein Density via Mutagenesis in Rice Results in an Enhanced Rate of Photosynthesis, Smaller Cell Sizes and Can Reduce Interveinal Mesophyll Cell Number

    Aryo B. Feldman

    2017-11-01

    Full Text Available Improvements to leaf photosynthetic rates of crops can be achieved by targeted manipulation of individual component processes, such as the activity and properties of RuBisCO or photoprotection. This study shows that simple forward genetic screens of mutant populations can also be used to rapidly generate photosynthesis variants that are useful for breeding. Increasing leaf vein density (concentration of vascular tissue per unit leaf area has important implications for plant hydraulic properties and assimilate transport. It was an important step to improving photosynthetic rates in the evolution of both C3 and C4 species and is a foundation or prerequisite trait for C4 engineering in crops like rice (Oryza sativa. A previous high throughput screen identified five mutant rice lines (cv. IR64 with increased vein densities and associated narrower leaf widths (Feldman et al., 2014. Here, these high vein density rice variants were analyzed for properties related to photosynthesis. Two lines were identified as having significantly reduced mesophyll to bundle sheath cell number ratios. All five lines had 20% higher light saturated photosynthetic capacity per unit leaf area, higher maximum carboxylation rates, dark respiration rates and electron transport capacities. This was associated with no significant differences in leaf thickness, stomatal conductance or CO2 compensation point between mutants and the wild-type. The enhanced photosynthetic rate in these lines may be a result of increased RuBisCO and electron transport component amount and/or activity and/or enhanced transport of photoassimilates. We conclude that high vein density (associated with altered mesophyll cell length and number is a trait that may confer increased photosynthetic efficiency without increased transpiration.

  7. Improving the efficiency of CHO cell line generation using glutamine synthetase gene knockout cells.

    Fan, Lianchun; Kadura, Ibrahim; Krebs, Lara E; Hatfield, Christopher C; Shaw, Margaret M; Frye, Christopher C

    2012-04-01

    Although Chinese hamster ovary (CHO) cells, with their unique characteristics, have become a major workhorse for the manufacture of therapeutic recombinant proteins, one of the major challenges in CHO cell line generation (CLG) is how to efficiently identify those rare, high-producing clones among a large population of low- and non-productive clones. It is not unusual that several hundred individual clones need to be screened for the identification of a commercial clonal cell line with acceptable productivity and growth profile making the cell line appropriate for commercial application. This inefficiency makes the process of CLG both time consuming and laborious. Currently, there are two main CHO expression systems, dihydrofolate reductase (DHFR)-based methotrexate (MTX) selection and glutamine synthetase (GS)-based methionine sulfoximine (MSX) selection, that have been in wide industrial use. Since selection of recombinant cell lines in the GS-CHO system is based on the balance between the expression of the GS gene introduced by the expression plasmid and the addition of the GS inhibitor, L-MSX, the expression of GS from the endogenous GS gene in parental CHOK1SV cells will likely interfere with the selection process. To study endogenous GS expression's potential impact on selection efficiency, GS-knockout CHOK1SV cell lines were generated using the zinc finger nuclease (ZFN) technology designed to specifically target the endogenous CHO GS gene. The high efficiency (∼2%) of bi-allelic modification on the CHO GS gene supports the unique advantages of the ZFN technology, especially in CHO cells. GS enzyme function disruption was confirmed by the observation of glutamine-dependent growth of all GS-knockout cell lines. Full evaluation of the GS-knockout cell lines in a standard industrial cell culture process was performed. Bulk culture productivity improved two- to three-fold through the use of GS-knockout cells as parent cells. The selection stringency was

  8. Effects of potentially acidic air pollutants on the intracellular distribution and transport of plant growth regulators in mesophyll cells of leaves. Consequences on stress- and developmental physiology

    Kremer, H.; Pfanz, H.; Hartung, W.

    1987-07-11

    The influence of SO/sub 2/ on the intracellular distribution of abscisic acid (ABA) and indole-acetic acid (IAA) in mesophyll cells of Picea abies, Tsuga americana and Hordeum vulgare was investigated. The compartmentation of ABA and IAA depends on intracellular pH-gradients. The hydrophilic anions ABA and IAA are accumulated in the alkaline cell compartments cytosol and chloroplasts, which act as anion traps for weak acids. Uptake of sulfur dioxide into leaves leads to an acidification of alkaline cell compartments, thus decreasing intracellular pH-gradients. Consequently this results in an increased release of plant growth regulators from the cell interior into the apoplast. Therefore the target cells of plant hormones i.e. meristems and stomates are exposed to altered hormone concentrations. Obviously this influences the regulation of cellular metabolism plant development and growth.

  9. [Effects of light intensities after anthesis on the photosynthetic characteristics and chloroplast ultrastructure in mesophyll cell of summer maize (Zea mays L. )].

    Gao, Jia; Cui, Hai Yan; Shi, Jian Guo; Dong, Shu Ting; Liu, Peng; Zhao, Bin; Zhang, Ji Wang

    2018-03-01

    We examined the changes of photosynthetic characteristics and chloroplast ultrastructure in mesophyll cell of summer maize in response to different light intensities in the field, with the summer maize hybrid Denghai 605 as experimental material. Two treatments of both shading (S) and increasing light (L) from flowering to physiological maturity stage were designed, with the ambient sunlight treatment as control (CK). Under shading treatment, poorly developed thylakoid structure, blurry lamellar structure, loose granum, large gap between slices and warping granum were the major characteristics in chloroplast. Meanwhile, photosynthetic rate (P n ), transpiration rate, stomatal conductance, chlorophyll content, and actual photo-chemical efficiency (Φ PSII ) decreased, whereas the maximal photochemical efficiency and non-photochemical quenching increased, which resulted in decreases in grain yield under shading treatment. However, a better development was observed in chloroplasts for L treatment, with the number of grana and lamellae increased and lamellae arranged compactly. In addition, P n and Φ PSII increased under L treatment, which increased grain yield. The chloroplast arrangement dispersed in mesophyll cells and chloroplast ultrastructure was destroyed after shading, and then chlorophyll synthesis per unit leaf area and photosynthetic capacity decreased. In contrast, the number of grana and lamellae increased and lamellae arranged compactly after increasing light, which are beneficial for corn yield.

  10. Influence of pH on the /sup 14/C-labelling pattern after photosynthesis of suspended leaf slices and isolated mesophyll cells from chenopodium album in NaH/sup 14/CO/sub 3/

    Baumann, G; Guenther, G [Paedagogische Hochschule Karl Liebknecht, Potsdam (German Democratic Republic). Sektion Chemie/Biologie

    1983-01-01

    Photosynthetic fixation of /sup 14/C from solutions of NaH/sup 14/CO/sub 3/ (at constant concentrations of free CO/sub 2/) by suspended leaf slices or isolated mesophyll cells from Chenopodium album is increased with increasing pH. Above all, the incorporation of radioactivity into amino acids and malate is stimulated. A direct uptake of HCO/sub 3/ ions and its fixation by PEP carboxylase is suggested. Isolated mesophyll cells showed at pH 7.3 a higher rate of photosynthesis than at pH 5.0.

  11. Glutamate/glutamine metabolism coupling between astrocytes and glioma cells: neuroprotection and inhibition of glioma growth.

    Yao, Pei-Sen; Kang, De-Zhi; Lin, Ru-Ying; Ye, Bing; Wang, Wei; Ye, Zu-Cheng

    2014-07-18

    Glioma glutamate release has been shown to promote the growth of glioma cells and induce neuronal injuries from epilepsy to neuronal death. However, potential counteractions from normal astrocytes against glioma glutamate release have not been fully evaluated. In this study, we investigated the glutamate/glutamine cycling between glioma cells and astrocytes and their impact on neuronal function. Co-cultures of glioma cells with astrocytes (CGA) in direct contact were established under different mix ratio of astrocyte/glioma. Culture medium conditioned in these CGAs were sampled for HPLC measurement, for neuronal ratiometric calcium imaging, and for neuronal survival assay. We found: (1) High levels of glutaminase expression in glioma cells, but not in astrocytes, glutaminase enables glioma cells to release large amount of glutamate in the presence of glutamine. (2) Glutamate levels in CGAs were directly determined by the astrocyte/glioma ratios, indicating a balance between glioma glutamate release and astrocyte glutamate uptake. (3) Culture media from CGAs of higher glioma/astrocyte ratios induced stronger neuronal Ca(2+) response and more severe neuronal death. (4) Co-culturing with astrocytes significantly reduced the growth rate of glioma cells. These results indicate that normal astrocytes in the brain play pivotal roles in glioma growth inhibition and in reducing neuronal injuries from glioma glutamate release. However, as tumor growth, the protective role of astrocytes gradually succumb to glioma cells. Copyright © 2014 Elsevier Inc. All rights reserved.

  12. Cell Death-Autophagy Loop and Glutamate-Glutamine Cycle in Amyotrophic Lateral Sclerosis

    Shu Yuan

    2017-07-01

    Full Text Available Although we know that amyotrophic lateral sclerosis (ALS is correlated with the glutamate-mediated corticomotor neuronal hyperexcitability, detailed ALS pathology remains largely unexplained. While a number of drugs have been developed, no cure exists so far. Here, we propose a hypothesis of neuronal cell death—incomplete autophagy positive-feedback loop—and summarize the role of the neuron-astrocyte glutamate-glutamine cycle in ALS. The disruption of these two cycles might ideally retard ALS progression. Cerebrovascular injuries (such as multiple embolization sessions and strokes induce neuronal cell death and the subsequent autophagy. ALS impairs autophagosome-lysosome fusion and leads to magnified cell death. Trehalose rescues this impaired fusion step, significantly delaying the onset of the disease, although it does not affect the duration of the disease. Therefore, trehalose might be a prophylactic drug for ALS. Given that a major part of neuronal glutamate is converted from glutamine through neuronal glutaminase (GA, GA inhibitors may decrease the neuronal glutamate accumulation, and, therefore, might be therapeutic ALS drugs. Of these, Ebselen is the most promising one with strong antioxidant properties.

  13. Alanyl-glutamine dipeptide restores the cytoprotective stress proteome of mesothelial cells exposed to peritoneal dialysis fluids.

    Kratochwill, Klaus; Boehm, Michael; Herzog, Rebecca; Lichtenauer, Anton Michael; Salzer, Elisabeth; Lechner, Michael; Kuster, Lilian; Bergmeister, Konstantin; Rizzi, Andreas; Mayer, Bernd; Aufricht, Christoph

    2012-03-01

    Exposure of mesothelial cells to peritoneal dialysis fluids (PDF) results in cytoprotective cellular stress responses (CSR) that counteract PDF-induced damage. In this study, we tested the hypothesis that the CSR may be inadequate in relevant models of peritoneal dialysis (PD) due to insufficient levels of glutamine, resulting in increased vulnerability against PDF cytotoxicity. We particularly investigated the role of alanyl-glutamine (Ala-Gln) dipeptide on the cytoprotective PDF stress proteome. Adequacy of CSR was investigated in two human in vitro models (immortalized cell line MeT-5A and mesothelial cells derived from peritoneal effluent of uraemic patients) following exposure to heat-sterilized glucose-based PDF (PD4-Dianeal, Baxter) diluted with medium and, in a comparative proteomics approach, at different levels of glutamine ranging from depletion (0 mM) via physiological (0.7 mM) to pharmacological levels (8 mM administered as Ala-Gln). Despite severe cellular injury, expression of cytoprotective proteins was dampened upon PDF exposure at physiological glutamine levels, indicating an inadequate CSR. Depletion of glutamine aggravated cell injury and further reduced the CSR, whereas addition of Ala-Gln at pharmacological level restored an adequate CSR, decreasing cellular damage in both PDF exposure systems. Ala-Gln specifically stimulated chaperoning activity, and cytoprotective processes were markedly enhanced in the PDF stress proteome. Taken together, this study demonstrates an inadequate CSR of mesothelial cells following PDF exposure associated with low and physiological levels of glutamine, indicating a new and potentially relevant pathomechanism. Supplementation of PDF with pharmacological doses of Ala-Gln restored the cytoprotective stress proteome, resulting in improved resistance of mesothelial cells to exposure to PDF. Future work will study the clinical relevance of CSR-mediated cytoprotection.

  14. Glutamine alimentation in catabolic state.

    Boelens, P G; Nijveldt, R J; Houdijk, A P; Meijer, S; van Leeuwen, P A

    2001-09-01

    Glutamine should be reclassified as a conditionally essential amino acid in the catabolic state because the body's glutamine expenditures exceed synthesis and low glutamine levels in plasma are associated with poor clinical outcome. After severe stress, several amino acids are mobilized from muscle tissue to supply energy and substrate to the host. Glutamine is one of the most important amino acids that provide this function. Glutamine acts as the preferred respiratory fuel for lymphocytes, hepatocytes and intestinal mucosal cells and is metabolized in the gut to citrulline, ammonium and other amino acids. Low concentrations of glutamine in plasma reflect reduced stores in muscle and this reduced availability of glutamine in the catabolic state seems to correlate with increased morbidity and mortality. Adding glutamine to the nutrition of clinical patients, enterally or parenterally, may reduce morbidity. Several excellent clinical trials have been performed to prove efficacy and feasibility of the use of glutamine supplementation in parenteral and enteral nutrition. The increased intake of glutamine has resulted in lower septic morbidity in certain critically ill patient populations. This review will focus on the efficacy and the importance of glutamine supplementation in diverse catabolic states.

  15. Glutamine Synthetase: Localization Dictates Outcome

    Alessandra Castegna

    2018-02-01

    Full Text Available Glutamine synthetase (GS is the adenosine triphosphate (ATP-dependent enzyme that catalyses the synthesis of glutamine by condensing ammonium to glutamate. In the circulatory system, glutamine carries ammonia from muscle and brain to the kidney and liver. In brain reduction of GS activity has been suggested as a mechanism mediating neurotoxicity in neurodegenerative disorders. In cancer, the delicate balance between glutamine synthesis and catabolism is a critical event. In vitro evidence, confirmed in vivo in some cases, suggests that reduced GS activity in cancer cells associates with a more invasive and aggressive phenotype. However, GS is known to be highly expressed in cells of the tumor microenvironment, such as fibroblasts, adipocytes and immune cells, and their ability to synthesize glutamine is responsible for the acquisition of protumoral phenotypes. This has opened a new window into the complex scenario of the tumor microenvironment, in which the balance of glutamine consumption versus glutamine synthesis influences cellular function. Since GS expression responds to glutamine starvation, a lower glutamine synthesizing power due to the absence of GS in cancer cells might apply a metabolic pressure on stromal cells. This event might push stroma towards a GS-high/protumoral phenotype. When referred to stromal cells, GS expression might acquire a ‘bad’ significance to the point that GS inhibition might be considered a conceivable strategy against cancer metastasis.

  16. Aging Reduces an ERRalpha-Directed Mitochondrial Glutaminase Expression Suppressing Glutamine Anaplerosis and Osteogenic Differentiation of Mesenchymal Stem Cells.

    Huang, Tongling; Liu, Renzhong; Fu, Xuekun; Yao, Dongsheng; Yang, Meng; Liu, Qingli; Lu, William W; Wu, Chuanyue; Guan, Min

    2017-02-01

    Aging deteriorates osteogenic capacity of mesenchymal stem/stromal cells (MSCs), contributing to imbalanced bone remodeling and osteoporosis. Glutaminase (Gls) catabolizes glutamine into glutamate at the first step of mitochondrial glutamine (Gln)-dependent anaplerosis which is essential for MSCs upon osteogenic differentiation. Estrogen-related receptor α (ERRα) regulates genes required for mitochondrial function. Here, we found that ERRα and Gls are upregulated by osteogenic induction in human MSCs (hMSCs). In contrast, osteogenic differentiation capacity and glutamine consumption of MSCs, as well as ERRα, Gls and osteogenic marker genes are significantly reduced with age. We demonstrated that ERRα binds to response elements on Gls promoter and affects glutamine anaplerosis through transcriptional induction of Gls. Conversely, mTOR inhibitor rapamycin, ERRα inverse agonist compound 29 or Gls inhibitor BPTES leads to reduced Gln anaplerosis and deteriorated osteogenic differentiation of hMSCs. Importantly, overexpression of ERRα or Gls restored impairment by these inhibitors. Finally, we proved that compensated ERRα or Gls expression indeed potentiated Gln anaplerosis and osteogenic capability of elderly mice MSCs in vitro. Together, we establish that Gls is a novel ERRα target gene and ERRα/Gls signaling pathway plays an important role in osteogenic differentiation of MSCs, providing new sights into novel regenerative therapeutics development. Our findings suggest that restoring age-related mitochondrial Gln-dependent anaplerosis may be beneficial for degenerative bone disorders such as osteoporosis. Stem Cells 2017;35:411-424. © 2016 AlphaMed Press.

  17. L-Glutamine in vitro Modulates some Immunomodulatory Properties of Bone Marrow Mesenchymal Stem Cells.

    Dos Santos, Guilherme Galvão; Hastreiter, Araceli Aparecida; Sartori, Talita; Borelli, Primavera; Fock, Ricardo Ambrósio

    2017-08-01

    Glutamine (GLUT) is a nonessential amino acid that can become conditionally essential under stress conditions, being able to act in the modulation of the immune responses. Mesenchymal stem cells (MSCs) are known to their capability in the modulation of immune responses through cell-cell contact and by the secretion of soluble factors. Considering that GLUT is an immunonutrient and little is known about the influence of GLUT on the capability of MSCs to modulate immune cells, this work aims to investigate how variations in GLUT concentrations in vitro could affect some immunomodulatory properties of MSCs. In order to evaluate the effects of GLUT on MSCs immunomodulatory properties, cell proliferation rates, the expression of NFκB and STAT-3, and the production of IL-1β, IL-6, IL-10, TGF-β and TNF-α by MSCs were assessed. Based on our findings, GLUT at high doses (10 mM) augmented the proliferation of MSCs and modulated immune responses by decreasing levels of pro-inflammatory cytokines, such as IL-1β and IL-6, and by increasing levels of anti-inflammatory cytokines IL-10 and TGF-β. In addition, MSCs cultured in higher GLUT concentrations (10 mM) expressed lower levels of NF-κB and higher levels of STAT-3. Furthermore, conditioned media from MSCs cultured at higher GLUT concentrations (10 mM) reduced lymphocyte and macrophage proliferation, increased IL-10 production by both cells types, and decreased IFN-γ production by lymphocytes. Overall, this study showed that 10 mM of GLUT is able to modify immunomodulatory properties of MSCs.

  18. Incorporation of uridine-H3 into healthy and tobacco necrosis virus-infected mesophyll cells of Chenopodium amaranticolor

    Faccioli, G.; Rubies-Autonel, C.

    1975-01-01

    Tritiated uridine was selectively incorporated into the nucleus, nucleolus and cytoplasm of actinomycin D-treated Chenopodium amaranticolor cells locally infected with a strain of tobacco necrosis virus (TNV), 3 days after inoculation. Healthy cells did not show such an incorporation. Chloroplasts, in both types of cells, were free of label. Treatment with pancreatic ribonuclease removed the label completely in the majority of nuclei and nucleoli of infected cells. Since infectivity tests showed that AMD treatment increased virus multiplication by 10-12%, it is conceivable to think that the incorporation observed was due to virus synthesis. Preliminary infectivity experiments also showed that treatment of the cells with cycloheximide inhibited virus multiplication up to 80%, while chloramphenicol increased such multiplication. Our results lead to the conclusion that nucleus, nucleolus and cytoplasm but not chloroplasts are the sites involved in the synthesis of TNV. (orig.) [de

  19. Glutamine domain of the chimeric protein, CAD, that initiates pyrimidine biosynthesis in mammalian cells

    Kelly, R.E.; Kim, H.; Evans, D.R.

    1986-01-01

    Glutamine dependent carbamyl phosphate synthesis, the first step in mammalian de novo pyrimidine biosynthesis, is catalyzed by a 240 kDa chimeric protein, CAD, that also has the aspartate transcarbamylase and dihydroorotase activities. The complex was found to have a separate glutaminase activity of 0.04 μmol/min/mg, that increased five fold in the presence of bicarbonate and ATP. To determine whether the glutaminase activity, which provides ammonia for carbamyl phosphate synthesis, is associated with a separate structural domain (GLN), CAD was subjected to controlled proteolysis with elastase. The glutaminase, glutamine and ammonia dependent carbamyl phosphate synthetase activities, as well as the partial reactions; carbamyl phosphate dependent ATP synthesis and bicarbonate dependent ATPase, were correlated with the concentration of the various proteolytic fragments that accumulated in the digest. While the glutamine dependent carbamyl phosphate synthetase was rapidly inactivated, the glutaminase activity was found to be very resistant to proteolysis. The glutamine binding site of CAD was also specifically modified with 6-diazo-5-oxo-L-norleucine (DON). The modification was accompanied by a loss of both glutaminase and glutamine dependent carbamyl phosphate synthetase activities. Bicarbonate and ATP increased the rate of reaction of CAD with DON, while glutamine protected against inactivation. The stoichiometry of the reaction and the identity of the modified proteolytic fragments was determined using 14 C labelled DON

  20. The Glutamine-Glutamate/GABA Cycle

    Walls, Anne B; Waagepetersen, Helle S; Bak, Lasse Kristoffer

    2015-01-01

    information about glutamine transfer. The present review will give information about glutamine trafficking and the tools used to map it as exemplified by discussions of published work employing brain cell cultures as well as intact animals. It will be documented that considerably more glutamine is transferred...

  1. Plant, cell, and molecular mechanisms of abscisic-acid regulation of stomatal apertures. A new mechanism for the regulation of stomatal-aperture size in intact leaves: Accumulation of mesophyll-derived sucrose in the guard-cell wall of Vicia faba L.

    Lu, P.; Outlaw, W.H. Jr.; Smith, B.G.; Freed, G.A.

    1996-12-31

    At various times after pulse labeling Vicia faba L. leaflets with {sup 14}CO{sub 2}, whole-leaf pieces and rinsed epidermal peels were harvested and subsequently processed for histochemical analysis. Cells dissected from whole leaf retained apoplastic contents whereas those from rinsed peels contained only cytoplastic contents. Sucrose specific radioactivity peaked in palisade cells, 111 GBq{center_dot}mol{sup {minus}1}, at 20 min. In contrast, the {sup 14}C content and sucrose specific radioactivity were very low in guard cells for 20 min, implying little CO{sub 2} incorporation; both then peaked at 40 min. The guard-cell apoplast had a high maximum sucrose specific radioactivity and a high sucrose influx rate. These and other comparisons implied the presence of (a) multiple sucrose pools in mesophyll cells, (b) a localized mesophyll-apoplast region that exchanges with phloem and stomata, and (c) mesophyll-derived sucrose in guard-cell walls sufficient to diminish stomatal opening by {approximately} 4 {micro}m. Factors expected to enhance sucrose accumulation in guard-cell walls are (a) high transpiration rate, which closes stomata, and (b) high apoplastic sucrose concentration, which is elevated when mesophyll-sucrose efflux exceeds translocation. Therefore, multiple physiological factors are integrated in the attenuation of stomatal-aperture size by this previously unrecognized mechanism.

  2. Glutamine synthetase gene knockout-human embryonic kidney 293E cells for stable production of monoclonal antibodies.

    Yu, Da Young; Lee, Sang Yoon; Lee, Gyun Min

    2018-05-01

    Previously, it was inferred that a high glutamine synthetase (GS) activity in human embryonic kidney (HEK) 293E cells results in elevated resistance to methionine sulfoximine (MSX) and consequently hampers GS-mediated gene amplification and selection by MSX. To overcome this MSX resistance in HEK293E cells, a GS-knockout HEK293E cell line was generated using the CRISPR/Cas9 system to target the endogenous human GS gene. The GS-knockout in the HEK293E cell line (RK8) was confirmed by Western blot analysis of GS and by observation of glutamine-dependent growth. Unlike the wild type HEK293E cells, the RK8 cells were successfully used as host cells to generate a recombinant HEK293E cell line (rHEK293E) producing a monoclonal antibody (mAb). When the RK8 cells were transfected with the GS expression vector containing the mAb gene, rHEK293E cells producing the mAb could be selected in the absence as well as in the presence of MSX. The gene copies and mRNA expression levels of the mAb in rHEK293E cells were also quantified using qRT-PCR. Taken together, the GS-knockout HEK293E cell line can be used as host cells to generate stable rHEK293E cells producing a mAb through GS-mediated gene selection in the absence as well as in the presence of MSX. © 2018 Wiley Periodicals, Inc.

  3. Cancerous epithelial cell lines shed extracellular vesicles with a bimodal size distribution that is sensitive to glutamine inhibition

    Santana, Steven Michael; Kirby, Brian J; Antonyak, Marc A; Cerione, Richard A

    2014-01-01

    Extracellular shed vesicles (ESVs) facilitate a unique mode of cell–cell communication wherein vesicle uptake can induce a change in the recipient cell's state. Despite the intensity of ESV research, currently reported data represent the bulk characterization of concentrated vesicle samples with little attention paid to heterogeneity. ESV populations likely represent diversity in mechanisms of formation, cargo and size. To better understand ESV subpopulations and the signaling cascades implicated in their formation, we characterize ESV size distributions to identify subpopulations in normal and cancerous epithelial cells. We have discovered that cancer cells exhibit bimodal ESV distributions, one small-diameter and another large-diameter population, suggesting that two mechanisms may govern ESV formation, an exosome population and a cancer-specific microvesicle population. Altered glutamine metabolism in cancer is thought to fuel cancer growth but may also support metastatic niche formation through microvesicle production. We describe the role of a glutaminase inhibitor, compound 968, in ESV production. We have discovered that inhibiting glutamine metabolism significantly impairs large-diameter microvesicle production in cancer cells. (paper)

  4. Comprehensive characterization of glutamine synthetase-mediated selection for the establishment of recombinant CHO cells producing monoclonal antibodies

    Noh, Soo Min; Shin, Seunghyeon; Min Lee, Gyun

    2018-01-01

    To characterize a glutamine synthetase (GS)-based selection system, monoclonal antibody (mAb) producing recombinant CHO cell clones were generated by a single round of selection at various methionine sulfoximine (MSX) concentrations (0, 25, and 50 μM) using two different host cell lines (CHO-K1...... and GS-knockout CHO). Regardless of the host cell lines used, the clones selected at 50 μM MSX had the lowest average specific growth rate and the highest average specific production rates of toxic metabolic wastes, lactate and ammonia. Unlike CHO-K1, high producing clones could be generated...... in the absence of MSX using GS-knockout CHO with an improved selection stringency. Regardless of the host cell lines used, the clones selected at various MSX concentrations showed no significant difference in the GS, heavy chain, and light chain gene copies (P > 0.05). Furthermore, there was no correlation...

  5. Glutamine synthetase activity in solanaceous cell suspensions accumulating alkaloids or not. 13C NMR and enzymatic assay

    Mesnard, F.; Marty, D.; Monti, J.P.; Gillet-Manceau, F.; Fliniaux, M.A.

    1999-01-01

    The metabolism of labelled pyruvate followed by 13 C NMR and the measure of glutamine synthetase (GS) showed, according to previous results, a high activity of this enzyme in suspension cells of Nicotiana plumbaginifolia. This activity could derive glutamate from the alkaloid synthesizing pathways. However, a recent work showed that the rate of the GS gene transcription was inversely proportional to the Gln/Glu ratio. The measures of Gln and Glu concentrations in Nicotiana plumbaginifolia cells revealed that high GS activity correlates with the weak value of Gln/Glu ratio. Therefore, the hypothesis of GS dysfunction for the non-biosynthesis of alkaloids in N. plumbaginifolia suspension cells can be discarded. This conclusion is strengthened by the results obtained when using a GS inhibitor. (author)

  6. Oral feeding with L-Glutamine and Nucleotides: impact on some GALT (gut associated lymphoid tissue parameters and cell proliferation/death rates in weaning piglets

    V. Bontempo

    2011-03-01

    Full Text Available Dietary supplementation with glutamine and nucleotides may be useful during piglets weaning, when a rough passage from milk suckling to a solid feed may cause a strong reduction of the length of intestinal villi height and the depth of the crypts, and consequently of the intestinal digestive and absorptive capacities (Van Beers-Schreurs et al., 1998. Glutamine stimulates cell proliferation and activates protein kinases, suggesting that it could control the regularly alternating cellular apoptosis/proliferation sequence (Rhoads et al., 2000...

  7. Inhibition of nitrogen-fixing activity of the cyanobiont affects the localization of glutamine synthetase in hair cells of Azolla.

    Uheda, Eiji; Maejima, Kazuhiro

    2009-10-15

    In the Azolla-Anabaena association, the host plant Azolla efficiently incorporates and assimilates ammonium ions that are released from the nitrogen-fixing cyanobiont, probably via glutamine synthetase (GS; EC 6.3.1.2) in hair cells, which are specialized cells protruding into the leaf cavity. In order to clarify the regulatory mechanism underlying ammonium assimilation in the Azolla-Anabaena association, Azolla plants were grown under an argon environment (Ar), in which the nitrogen-fixing activity of the cyanobiont was inhibited specifically and completely. The localization of GS in hair cells was determined by immunoelectron microscopy and quantitative analysis of immunogold labeling. Azolla plants grew healthily under Ar when nitrogen sources, such as NO(3)(-) and NH(4)(+), were provided in the growth medium. Both the number of cyanobacterial cells per leaf and the heterocyst frequency of the plants under Ar were similar to those of plants in a nitrogen environment (N(2)). In hair cells of plants grown under Ar, regardless of the type of nitrogen source provided, only weak labeling of GS was observed in the cytoplasm and in chloroplasts. In contrast, in hair cells of plants grown under N(2), abundant labeling of GS was observed in both sites. These findings indicate that specific inhibition of the nitrogen-fixing activity of the cyanobiont affects the localization of GS isoenzymes. Ammonium fixed and released by the cyanobiont could stimulate GS synthesis in hair cells. Simultaneously, the abundant GS, probably GS1, in these cells, could assimilate ammonium rapidly.

  8. Carbon isotope ratios of epidermal and mesophyll tissues from leaves of C3 and CAM plants

    Nishida, K.; Roksandic, Z.; Osmond, B.

    1981-01-01

    The δ 13 C values for epidermal and mesophyll tissues of two C 3 plants, Commelina communis and Tulipa gesneriana, and a CAM plant, Kalanchoē daigremontiana, were measured. The values for the tissues of both C 3 plants were similar. In young leaves of Kalanchoē, the epidermis and the mesophyll showed S 13 C values which were nearly identical, and similar to those found in C 3 plants. However, markedly more negative values for epidermal compared to mesophyll tissue, were obtained in the mature Kalanchoē leaf. This is consistent with the facts that the epidermis in a CAM leaf is formed when leaves engage in C 3 photosynthesis and that subsequent dark CO 2 fixation in guard cells or mesophyll cells makes only a small contribution to total epidermal carbon

  9. Glutamine Supplementation Attenuates Expressions of Adhesion Molecules and Chemokine Receptors on T Cells in a Murine Model of Acute Colitis

    Yu-Chen Hou

    2014-01-01

    Full Text Available Background. Migration of T cells into the colon plays a major role in the pathogenesis in inflammatory bowel disease. This study investigated the effects of glutamine (Gln supplementation on chemokine receptors and adhesion molecules expressed by T cells in mice with dextran sulfate sodium- (DSS- induced colitis. Methods. C57BL/6 mice were fed either a standard diet or a Gln diet replacing 25% of the total nitrogen. After being fed the diets for 5 days, half of the mice from both groups were given 1.5% DSS in drinking water to induce colitis. Mice were killed after 5 days of DSS exposure. Results. DSS colitis resulted in higher expression levels of P-selectin glycoprotein ligand- (PSGL- 1, leukocyte function-associated antigen- (LFA- 1, and C-C chemokine receptor type 9 (CCR9 by T helper (Th and cytotoxic T (Tc cells, and mRNA levels of endothelial adhesion molecules in colons were upregulated. Gln supplementation decreased expressions of PSGL-1, LFA-1, and CCR9 by Th cells. Colonic gene expressions of endothelial adhesion molecules were also lower in Gln-colitis mice. Histological finding showed that colon infiltrating Th cells were less in the DSS group with Gln administration. Conclusions. Gln supplementation may ameliorate the inflammation of colitis possibly via suppression of T cell migration.

  10. Blocking anaplerotic entry of glutamine into the TCA cycle sensitizes K-Ras mutant cancer cells to cytotoxic drugs.

    Saqcena, M; Mukhopadhyay, S; Hosny, C; Alhamed, A; Chatterjee, A; Foster, D A

    2015-05-14

    Cancer cells undergo a metabolic transformation that allows for increased anabolic demands, wherein glycolytic and tricarboxylic acid (TCA) cycle intermediates are shunted away for the synthesis of biological molecules required for cell growth and division. One of the key shunts is the exit of citrate from the mitochondria and the TCA cycle for the generation of cytosolic acetyl-coenzyme A that can be used for fatty acid and cholesterol biosynthesis. With the loss of mitochondrial citrate, cancer cells rely on the 'conditionally essential' amino acid glutamine (Q) as an anaplerotic carbon source for TCA cycle intermediates. Although Q deprivation causes G1 cell cycle arrest in non-transformed cells, its impact on the cancer cell cycle is not well characterized. We report here a correlation between bypass of the Q-dependent G1 checkpoint and cancer cells harboring K-Ras mutations. Instead of arresting in G1 in response to Q-deprivation, K-Ras-driven cancer cells arrest in either S- or G2/M-phase. Inhibition of K-Ras effector pathways was able to revert cells to G1 arrest upon Q deprivation. Blocking anaplerotic utilization of Q mimicked Q deprivation--causing S- and G2/M-phase arrest in K-Ras mutant cancer cells. Significantly, Q deprivation or suppression of anaplerotic Q utilization created synthetic lethality to the cell cycle phase-specific cytotoxic drugs, capecitabine and paclitaxel. These data suggest that disabling of the G1 Q checkpoint could represent a novel vulnerability of cancer cells harboring K-Ras and possibly other mutations that disable the Q-dependent checkpoint.

  11. Comprehensive characterization of glutamine synthetase-mediated selection for the establishment of recombinant CHO cells producing monoclonal antibodies.

    Noh, Soo Min; Shin, Seunghyeon; Lee, Gyun Min

    2018-03-29

    To characterize a glutamine synthetase (GS)-based selection system, monoclonal antibody (mAb) producing recombinant CHO cell clones were generated by a single round of selection at various methionine sulfoximine (MSX) concentrations (0, 25, and 50 μM) using two different host cell lines (CHO-K1 and GS-knockout CHO). Regardless of the host cell lines used, the clones selected at 50 μM MSX had the lowest average specific growth rate and the highest average specific production rates of toxic metabolic wastes, lactate and ammonia. Unlike CHO-K1, high producing clones could be generated in the absence of MSX using GS-knockout CHO with an improved selection stringency. Regardless of the host cell lines used, the clones selected at various MSX concentrations showed no significant difference in the GS, heavy chain, and light chain gene copies (P > 0.05). Furthermore, there was no correlation between the specific mAb productivity and these three gene copies (R 2  ≤ 0.012). Taken together, GS-mediated gene amplification does not occur in a single round of selection at a MSX concentration up to 50 μM. The use of the GS-knockout CHO host cell line facilitates the rapid generation of high producing clones with reduced production of lactate and ammonia in the absence of MSX.

  12. Feedback inhibition of ammonium (methylammonium) ion transport in Escherichia coli by glutamine and glutamine analogs

    Jayakumar, A.; Hong, J.S.; Barnes, E.M. Jr.

    1987-01-01

    When cultured with glutamate or glutamine as the nitrogen source, Escherichia coli expresses a specific ammonium (methylammonium) transport system. Over 95% of the methylammonium transport activity in washed cells was blocked by incubation with 100 μM L-glutamine in the presence of chloramphenicol (100 μg/ml). The inhibition of transport by L-glutamine was noncompetitive with respect to the [ 14 C]methylammonium substrate. D-Glutamine had no significant effect. The glutamine analogs γ-L-glutamyl hydroxamate and γ-L-glutamyl hydrazide were also noncompetitive inhibitors of methylammonium transport, suggesting that glutamine metabolism is not required. The role of the intracellular glutamine pool in the regulation of ammonium transport was investigated by using mutants carrying defects in the operon of glnP, the gene for the glutamine transporter. The glnP mutants had normal rates of methylammonium transport but were refractory to glutamine inhibition. Glycylglycine, a noncompetitive inhibitor of methylammonium uptake in wild-type cells, was equipotent in blocking transport in glnP mutants. Although ammonium transport is also subject to repression by growth of E. coli in the presence of ammonia, this phenomenon is unrelated to glutamine inhibition

  13. Growth factors regulate glutamine synthetase activity in ...

    Khaled

    2012-07-10

    Jul 10, 2012 ... glutamate and ammonia, which in turn, cells are supplied with ammonia ... out to determine the maximum growth time at which cells will be .... Western blot technique for detection the glutamine synthetase enzyme. Lane 1;.

  14. De Novo Glutamine Synthesis

    He, Qiao; Shi, Xinchong; Zhang, Linqi; Yi, Chang; Zhang, Xuezhen

    2016-01-01

    Purpose: The aim of this study was to investigate the role of de novo glutamine (Gln) synthesis in the proliferation of C6 glioma cells and its detection with 13N-ammonia. Methods: Chronic Gln-deprived C6 glioma (0.06C6) cells were established. The proliferation rates of C6 and 0.06C6 cells were measured under the conditions of Gln deprivation along with or without the addition of ammonia or glutamine synthetase (GS) inhibitor. 13N-ammonia uptake was assessed in C6 cells by gamma counting and in rats with C6 and 0.06C6 xenografts by micro–positron emission tomography (PET) scanning. The expression of GS in C6 cells and xenografts was assessed by Western blotting and immunohistochemistry, respectively. Results: The Gln-deprived C6 cells showed decreased proliferation ability but had a significant increase in GS expression. Furthermore, we found that low concentration of ammonia was sufficient to maintain the proliferation of Gln-deprived C6 cells, and 13N-ammonia uptake in C6 cells showed Gln-dependent decrease, whereas inhibition of GS markedly reduced the proliferation of C6 cells as well as the uptake of 13N-ammoina. Additionally, microPET/computed tomography exhibited that subcutaneous 0.06C6 xenografts had higher 13N-ammonia uptake and GS expression in contrast to C6 xenografts. Conclusion: De novo Gln synthesis through ammonia–glutamate reaction plays an important role in the proliferation of C6 cells. 13N-ammonia can be a potential metabolic PET tracer for Gln-dependent tumors. PMID:27118759

  15. Glutamine reduces myocardial cell apoptosis in a rat model of sepsis by promoting expression of heat shock protein 90.

    Li, Wanxia; Tao, Shaoyu; Wu, Qinghua; Wu, Tao; Tao, Ran; Fan, Jun

    2017-12-01

    Myocardial cell injury and cardiac myocyte apoptosis are associated with sepsis. Glutamine (Gln) has been reported to repair myocardial cell injury. The aim of this study was to explore the role of Gln on cardiac myocytes in a cecal ligation and puncture (CLP) model of sepsis in Wistar rats. Following induction of sepsis in a CLP rat model, viral encoding heat shock protein 90 (Hsp90) gene and Hsp90dsDNA were designed to express and knockdown Hsp90, respectively. Rat cardiac tissues were examined histologically, and apoptosis was detected by terminal deoxynucleotidyl transferase dUTP nick end labeling staining. The expression of B-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein, Hsp90, p53 upregulated modulator of apoptosis, and p53 was measured by western blotting and real-time polymerase chain reaction. Caspase-3, caspase-8, and caspase-9 were detected by enzyme-linked immunosorbent assay. Rat cardiac myocyte damage induced by CLP was reduced by Gln treatment and Hsp90 overexpression, and these changes were reversed by Hsp90 knockdown. Bcl-2 expression, Bcl-2-associated X protein, p53, p53 upregulated modulator of apoptosis, caspase-8, caspase-9, and caspase-3 activities were significantly upregulated in the CLP model, which were reduced by Gln treatment and Hsp90 overexpression. Gln reduced apoptosis of cardiac myocytes in a rat model of sepsis, by promoting Hsp90 expression. Further studies are needed to determine the possible therapeutic action of Gln in sepsis in human tissue. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Glutamine supplementation suppresses herpes simplex virus reactivation.

    Wang, Kening; Hoshino, Yo; Dowdell, Kennichi; Bosch-Marce, Marta; Myers, Timothy G; Sarmiento, Mayra; Pesnicak, Lesley; Krause, Philip R; Cohen, Jeffrey I

    2017-06-30

    Chronic viral infections are difficult to treat, and new approaches are needed, particularly those aimed at reducing reactivation by enhancing immune responses. Herpes simplex virus (HSV) establishes latency and reactivates frequently, and breakthrough reactivation can occur despite suppressive antiviral therapy. Virus-specific T cells are important to control HSV, and proliferation of activated T cells requires increased metabolism of glutamine. Here, we found that supplementation with oral glutamine reduced virus reactivation in latently HSV-1-infected mice and HSV-2-infected guinea pigs. Transcriptome analysis of trigeminal ganglia from latently HSV-1-infected, glutamine-treated WT mice showed upregulation of several IFN-γ-inducible genes. In contrast to WT mice, supplemental glutamine was ineffective in reducing the rate of HSV-1 reactivation in latently HSV-1-infected IFN-γ-KO mice. Mice treated with glutamine also had higher numbers of HSV-specific IFN-γ-producing CD8 T cells in latently infected ganglia. Thus, glutamine may enhance the IFN-γ-associated immune response and reduce the rate of reactivation of latent virus infection.

  17. De Novo Glutamine Synthesis: Importance for the Proliferation of Glioma Cells and Potentials for Its Detection With 13N-Ammonia.

    He, Qiao; Shi, Xinchong; Zhang, Linqi; Yi, Chang; Zhang, Xuezhen; Zhang, Xiangsong

    2016-01-01

    The aim of this study was to investigate the role of de novo glutamine (Gln) synthesis in the proliferation of C6 glioma cells and its detection with (13)N-ammonia. Chronic Gln-deprived C6 glioma (0.06C6) cells were established. The proliferation rates of C6 and 0.06C6 cells were measured under the conditions of Gln deprivation along with or without the addition of ammonia or glutamine synthetase (GS) inhibitor. (13)N-ammonia uptake was assessed in C6 cells by gamma counting and in rats with C6 and 0.06C6 xenografts by micro-positron emission tomography (PET) scanning. The expression of GS in C6 cells and xenografts was assessed by Western blotting and immunohistochemistry, respectively. The Gln-deprived C6 cells showed decreased proliferation ability but had a significant increase in GS expression. Furthermore, we found that low concentration of ammonia was sufficient to maintain the proliferation of Gln-deprived C6 cells, and (13)N-ammonia uptake in C6 cells showed Gln-dependent decrease, whereas inhibition of GS markedly reduced the proliferation of C6 cells as well as the uptake of (13)N-ammoina. Additionally, microPET/computed tomography exhibited that subcutaneous 0.06C6 xenografts had higher (13)N-ammonia uptake and GS expression in contrast to C6 xenografts. De novo Gln synthesis through ammonia-glutamate reaction plays an important role in the proliferation of C6 cells. (13)N-ammonia can be a potential metabolic PET tracer for Gln-dependent tumors. © The Author(s) 2016.

  18. Glutamine deprivation induces interleukin-8 expression in ataxia telangiectasia fibroblasts.

    Kim, Min-Hyun; Kim, Aryung; Yu, Ji Hoon; Lim, Joo Weon; Kim, Hyeyoung

    2014-05-01

    To investigate whether glutamine deprivation induces expression of inflammatory cytokine interleukin-8 (IL-8) by determining NF-κB activity and levels of oxidative indices (ROS, reactive oxygen species; hydrogen peroxide; GSH, glutathione) in fibroblasts isolated from patients with ataxia telangiectasia (A-T). We used A-T fibroblasts stably transfected with empty vector (Mock) or with human full-length ataxia telangiectasia mutated (ATM) cDNA (YZ5) and mouse embryonic fibroblasts (MEFs) transiently transfected with ATM small interfering RNA (siRNA) or with non-specific control siRNA. The cells were cultured with or without glutamine or GSH. ROS levels were determined using a fluorescence reader and confocal microscopy. IL-8 or murine IL-8 homolog, keratinocyte chemoattractant (KC), and hydrogen peroxide levels in the medium were determined by enzyme-linked immunosorbent assay and colorimetric assay. GSH level was assessed by enzymatic assay, while IL-8 (KC) mRNA level was measured by reverse transcription-polymerase chain reaction (RT-PCR) and/or quantitative real-time PCR. NF-κB DNA-binding activity was determined by electrophoretic mobility shift assay. Catalase activity and ATM protein levels were determined by O2 generation and Western blotting. While glutamine deprivation induced IL-8 expression and increased NF-κB DNA-binding activity in Mock cells, both processes were decreased by treatment of cells with glutamine or GSH or both glutamine and GSH. Glutamine deprivation had no effect on IL-8 expression or NF-κB DNA-binding activity in YZ5 cells. Glutamine-deprived Mock cells had higher oxidative stress indices (increases in ROS and hydrogen peroxide, reduction in GSH) than glutamine-deprived YZ5 cells. In Mock cells, glutamine deprivation-induced oxidative stress indices were suppressed by treatment with glutamine or GSH or both glutamine and GSH. GSH levels and catalase activity were lower in Mock cells than YZ5 cells. MEFs transfected with ATM siRNA and

  19. Limitations to the development of recombinant human embryonic kidney 293E cells using glutamine synthetase-mediated gene amplification: Methionine sulfoximine resistance.

    Yu, Da Young; Noh, Soo Min; Lee, Gyun Min

    2016-08-10

    To investigate the feasibility of glutamine synthetase (GS)-mediated gene amplification in HEK293 cells for the high-level stable production of therapeutic proteins, HEK293E cells were transfected by the GS expression vector containing antibody genes and were selected at various methionine sulfoximine (MSX) concentrations in 96-well plates. For a comparison, CHOK1 cells were transfected by the same GS expression vector and selected at various MSX concentrations. Unlike CHOK1 cells, HEK293E cells producing high levels of antibodies were not selected at all. For HEK293E cells, the number of wells with the cell pool did not decrease with an increase in the concentration of MSX up to 500μM MSX. A q-RT-PCR analysis confirmed that the antibody genes in the HEK293E cells, unlike the CHOK1 cells, were not amplified after increasing the MSX concentration. It was found that the GS activity in HEK293E cells was much higher than that in CHOK1 cells (PMSX and therefore hampers GS-mediated gene amplification by MSX. Thus, in order to apply the GS-mediated gene amplification system to HEK293 cells, the endogenous GS expression level in HEK293 cells needs to be minimized by knock-out or down-regulation methods. Copyright © 2016 Elsevier B.V. All rights reserved.

  20. The influence of nitric oxide and mercury chloride on leaf mesophyll structure under natural drought conditions

    Mykola M. Musiyenko

    2012-03-01

    Full Text Available It is established that under natural drought conditions starch was accumulated in the central part of chloroplasts of mesophyll cells and chloroplasts were localized on the periphery of cells at plasmalemma. After treatment wheat plants by nitric oxide donor the decreasing of starch deposits number and close contacts between chloroplasts were indicated, elongated nucleus was localized in the centre of cells. After treatment wheat plant by mercury chloride chloroplasts in the cells lost their oval shape and contacts, increased eventually deposition of starch, indicating the acceleration of aging tissues. Thus, nitric oxide in drought conditions reduced the destructive effect of drought on mesophyll cells, and mercury chloride caused deformation of the membrane cell.

  1. Exogenous transglutaminase improves multiple-stress tolerance in Lactococcus lactis and other lactic acid bacteria with glutamine and lysine in the cell wall.

    Li, Yu; Kan, Zhipeng; You, Yuanli; Gao, Xueling; Wang, Zhigeng; Fu, Ruiyan

    2015-12-01

    To increase the resistance of ingested bacteria to multiple environmental stresses, the role of transglutaminase in Lactococcus lactis and possible mechanisms of action were explored. L. lactis grown with transglutaminase exhibited significantly higher resistance to bile salts, stimulated gastric juice, antibiotics, NaCl, and cold stress compared to the control (cultured without transglutaminase), with no negative influence on cell growth. Transmission electron microscopy revealed that the cell walls of L. lactis cultured with 9 U transglutaminase/ml were approx. 1.9-times thicker than the control. Further analysis demonstrated that the multi-resistant phenotype was strain-specific; that is, it occurred in bacteria with the presence of glutamine and lysine in the peptidoglycan. Supplementation of culture media with transglutaminase is an effective, simple, and inexpensive strategy to protect specific ingested bacteria against multiple environmental challenges.

  2. Endogenous glutamine decrease is associated with pancreatic cancer progression.

    Roux, Cecilia; Riganti, Chiara; Borgogno, Sammy Ferri; Curto, Roberta; Curcio, Claudia; Catanzaro, Valeria; Digilio, Giuseppe; Padovan, Sergio; Puccinelli, Maria Paola; Isabello, Monica; Aime, Silvio; Cappello, Paola; Novelli, Francesco

    2017-11-10

    Pancreatic ductal adenocarcinoma (PDAC) is becoming the second leading cause of cancer-related death in the Western world. The mortality is very high, which emphasizes the need to identify biomarkers for early detection. As glutamine metabolism alteration is a feature of PDAC, its in vivo evaluation may provide a useful tool for biomarker identification. Our aim was to identify a handy method to evaluate blood glutamine consumption in mouse models of PDAC. We quantified the in vitro glutamine uptake by Mass Spectrometry (MS) in tumor cell supernatants and showed that it was higher in PDAC compared to non-PDAC tumor and pancreatic control human cells. The increased glutamine uptake was paralleled by higher activity of most glutamine pathway-related enzymes supporting nucleotide and ATP production. Free glutamine blood levels were evaluated in orthotopic and spontaneous mouse models of PDAC and other pancreatic-related disorders by High-Performance Liquid Chromatography (HPLC) and/or MS. Notably we observed a reduction of blood glutamine as much as the tumor progressed from pancreatic intraepithelial lesions to invasive PDAC, but was not related to chronic pancreatitis-associated inflammation or diabetes. In parallel the increased levels of branched-chain amino acids (BCAA) were observed. By contrast blood glutamine levels were stable in non-tumor bearing mice. These findings demonstrated that glutamine uptake is measurable both in vitro and in vivo . The higher in vitro avidity of PDAC cells corresponded to a lower blood glutamine level as soon as the tumor mass grew. The reduction in circulating glutamine represents a novel tool exploitable to implement other diagnostic or prognostic PDAC biomarkers.

  3. Metabolic Alterations Caused by KRAS Mutations in Colorectal Cancer Contribute to Cell Adaptation to Glutamine Depletion by Upregulation of Asparagine Synthetase

    Kosuke Toda

    2016-11-01

    Full Text Available A number of clinical trials have shown that KRAS mutations of colorectal cancer (CRC can predict a lack of responses to anti-epidermal growth factor receptor–based therapy. Recently, there have been several studies to elucidate metabolism reprogramming in cancer. However, it remains to be investigated how mutated KRAS can coordinate the metabolic shift to sustain CRC tumor growth. In this study, we found that KRAS mutation in CRC caused alteration in amino acid metabolism. KRAS mutation causes a marked decrease in aspartate level and an increase in asparagine level in CRC. Using several human CRC cell lines and clinical specimens of primary CRC, we demonstrated that the expression of asparagine synthetase (ASNS, an enzyme that synthesizes asparagine from aspartate, was upregulated by mutated KRAS and that ASNS expression was induced by KRAS-activated signaling pathway, in particular PI3K-AKT-mTOR pathway. Importantly, we demonstrated that KRAS-mutant CRC cells could become adaptive to glutamine depletion through asparagine biosynthesis by ASNS and that asparagine addition could rescue the inhibited growth and viability of cells grown under the glutamine-free condition in vitro. Notably, a pronounced growth suppression of KRAS-mutant CRC was observed upon ASNS knockdown in vivo. Furthermore, combination of L-asparaginase plus rapamycin markedly suppressed the growth of KRAS-mutant CRC xenografts in vivo, whereas either L-asparaginase or rapamycin alone was not effective. These results indicate ASNS might be a novel therapeutic target against CRCs with mutated KRAS.

  4. Glutamine and glutamate as vital metabolites

    Newsholme P.

    2003-01-01

    Full Text Available Glucose is widely accepted as the primary nutrient for the maintenance and promotion of cell function. This metabolite leads to production of ATP, NADPH and precursors for the synthesis of macromolecules such as nucleic acids and phospholipids. We propose that, in addition to glucose, the 5-carbon amino acids glutamine and glutamate should be considered to be equally important for maintenance and promotion of cell function. The functions of glutamine/glutamate are many, i.e., they are substrates for protein synthesis, anabolic precursors for muscle growth, they regulate acid-base balance in the kidney, they are substrates for ureagenesis in the liver and for hepatic and renal gluconeogenesis, they act as an oxidative fuel for the intestine and cells of the immune system, provide inter-organ nitrogen transport, and act as precursors of neurotransmitter synthesis, of nucleotide and nucleic acid synthesis and of glutathione production. Many of these functions are interrelated with glucose metabolism. The specialized aspects of glutamine/glutamate metabolism of different glutamine-utilizing cells are discussed in the context of glucose requirements and cell function.

  5. Neuronal Cell Death Induced by Mechanical Percussion Trauma in Cultured Neurons is not Preceded by Alterations in Glucose, Lactate and Glutamine Metabolism.

    Jayakumar, A R; Bak, L K; Rama Rao, K V; Waagepetersen, H S; Schousboe, A; Norenberg, M D

    2016-02-01

    Traumatic brain injury (TBI) is a devastating neurological disorder that usually presents in acute and chronic forms. Brain edema and associated increased intracranial pressure in the early phase following TBI are major consequences of acute trauma. On the other hand, neuronal injury, leading to neurobehavioral and cognitive impairments, that usually develop months to years after single or repetitive episodes of head trauma, are major consequences of chronic TBI. The molecular mechanisms responsible for TBI-induced injury, however, are unclear. Recent studies have suggested that early mitochondrial dysfunction and subsequent energy failure play a role in the pathogenesis of TBI. We therefore examined whether oxidative metabolism of (13)C-labeled glucose, lactate or glutamine is altered early following in vitro mechanical percussion-induced trauma (5 atm) to neurons (4-24 h), and whether such events contribute to the development of neuronal injury. Cell viability was assayed using the release of the cytoplasmic enzyme lactate dehydrogenase (LDH), together with fluorescence-based cell staining (calcein and ethidium homodimer-1 for live and dead cells, respectively). Trauma had no effect on the LDH release in neurons from 1 to 18 h. However, a significant increase in LDH release was detected at 24 h after trauma. Similar findings were identified when traumatized neurons were stained with fluorescent markers. Additionally (13)C-labeling of glutamate showed a small, but statistically significant decrease at 14 h after trauma. However, trauma had no effect on the cycling ratio of the TCA cycle at any time-period examined. These findings indicate that trauma does not cause a disturbance in oxidative metabolism of any of the substrates used for neurons. Accordingly, such metabolic disturbance does not appear to contribute to the neuronal death in the early stages following trauma.

  6. Stomatal conductance, mesophyll conductance, and transpiration efficiency in relation to leaf anatomy in rice and wheat genotypes under drought.

    Ouyang, Wenjing; Struik, Paul C; Yin, Xinyou; Yang, Jianchang

    2017-11-02

    Increasing leaf transpiration efficiency (TE) may provide leads for growing rice like dryland cereals such as wheat (Triticum aestivum). To explore avenues for improving TE in rice, variations in stomatal conductance (gs) and mesophyll conductance (gm) and their anatomical determinants were evaluated in two cultivars from each of lowland, aerobic, and upland groups of Oryza sativa, one cultivar of O. glaberrima, and two cultivars of T. aestivum, under three water regimes. The TE of upland rice, O. glaberrima, and wheat was more responsive to the gm/gs ratio than that of lowland and aerobic rice. Overall, the explanatory power of the particular anatomical trait varied among species. Low stomatal density mostly explained the low gs in drought-tolerant rice, whereas rice genotypes with smaller stomata generally responded more strongly to drought. Compared with rice, wheat had a higher gm, which was associated with thicker mesophyll tissue, mesophyll and chloroplasts more exposed to intercellular spaces, and thinner cell walls. Upland rice, O. glaberrima, and wheat cultivars minimized the decrease in gm under drought by maintaining high ratios of chloroplasts to exposed mesophyll cell walls. Rice TE could be improved by increasing the gm/gs ratio via modifying anatomical traits. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  7. Glutamine synthetase activity in solanaceous cell suspensions accumulating alkaloids or not. {sup 13}C NMR and enzymatic assay; Activite de la glutamine synthetase dans des suspensions cellulaires de solanacees productrices ou non d'alcaloides. RMN du {sup 13}C et dosage enzymatique

    Mesnard, F.; Marty, D.; Monti, J.P. [Faculte de Pharmacie, 80 - Amiens (France). Laboratoire de Biophysique, Groupe de Recherche des Biomolecules: micro-environnement et Metabolisme; Gillet-Manceau, F.; Fliniaux, M.A. [Faculte de Pharmacie, 80 - Amiens (France). Laboratoire de Phytotechnologie

    1999-09-01

    The metabolism of labelled pyruvate followed by {sup 13}C NMR and the measure of glutamine synthetase (GS) showed, according to previous results, a high activity of this enzyme in suspension cells of Nicotiana plumbaginifolia. This activity could derive glutamate from the alkaloidsynthesizing pathways. However, a recent work showed that the rate of the GS gene transcription was inversely proportional to the Gln/Glu ratio. The measures of Gln and Glu concentrations in Nicotiana plumbaginifolia cells revealed that high GS activity correlates with the weak value of Gln/Glu ratio. Therefore, the hypothesis of GS dysfunction for the non-biosynthesis of alkaloids in N. plumbaginifolia suspension cells can be discarded. This conclusion is strengthened by the results obtained when using a GS inhibitor. (author)

  8. ATP-dependent and NAD-dependent modification of glutamine synthetase from Rhodospirillum rubrum in vitro

    Woehle, D.L.; Lueddecke, B.A.; Ludden, P.W.

    1990-01-01

    Glutamine synthetase from the photosynthetic bacterium Rhodospirillum rubrum is the target of both ATP- and NAD-dependent modification. Incubation of R. rubrum cell supernatant with [α- 32 P]NAD results in the labeling of glutamine synthetase and two other unidentified proteins. Dinitrogenase reductase ADP-ribosyltransferase does not appear to be responsible for the modification of glutamine synthetase or the unidentified proteins. The [α- 32 P]ATP- and [α- 32 P] NAD-dependent modifications of R. rubrum glutamine synthetase appear to be exclusive and the two forms of modified glutamine synthetase are separable on two-dimensional gels. Loss of enzymatic activity by glutamine synthetase did not correlate with [α- 32 P]NAD labeling. This is in contrast to inactivation by nonphysiological ADP-ribosylation of other glutamine synthetases by an NAD:arginine ADP-ribosyltransferase from turkey erythrocytes. A 32 P-labeled protein spot comigrates with the NAD-treated glutamine synthetase spot when glutamine synthetase purified from H 3 32 PO 4 -grown cells is analyzed on two-dimensional gels. The adenylylation site of R. rubrum glutamine synthetase has been determined to be Leu-(Asp)-Tyr-Leu-Pro-Pro-Glu-Glu-Leu-Met; the tyrosine residue is the site of modification

  9. Glutamine acts as a neuroprotectant against DNA damage, beta-amyloid and H2O2-induced stress.

    Jianmin Chen

    Full Text Available Glutamine is the most abundant free amino acid in the human blood stream and is 'conditionally essential' to cells. Its intracellular levels are regulated both by the uptake of extracellular glutamine via specific transport systems and by its intracellular synthesis by glutamine synthetase (GS. Adding to the regulatory complexity, when extracellular glutamine is reduced GS protein levels rise. Unfortunately, this excess GS can be maladaptive. GS overexpression is neurotoxic especially if the cells are in a low-glutamine medium. Similarly, in low glutamine, the levels of multiple stress response proteins are reduced rendering cells hypersensitive to H(2O(2, zinc salts and DNA damage. These altered responses may have particular relevance to neurodegenerative diseases of aging. GS activity and glutamine levels are lower in the Alzheimer's disease (AD brain, and a fraction of AD hippocampal neurons have dramatically increased GS levels compared with control subjects. We validated the importance of these observations by showing that raising glutamine levels in the medium protects cultured neuronal cells against the amyloid peptide, Aβ. Further, a 10-day course of dietary glutamine supplementation reduced inflammation-induced neuronal cell cycle activation, tau phosphorylation and ATM-activation in two different mouse models of familial AD while raising the levels of two synaptic proteins, VAMP2 and synaptophysin. Together, our observations suggest that healthy neuronal cells require both intracellular and extracellular glutamine, and that the neuroprotective effects of glutamine supplementation may prove beneficial in the treatment of AD.

  10. Effects of glutamine, taurine and their association on inflammatory pathway markers in macrophages.

    Sartori, Talita; Galvão Dos Santos, Guilherme; Nogueira-Pedro, Amanda; Makiyama, Edson; Rogero, Marcelo Macedo; Borelli, Primavera; Fock, Ricardo Ambrósio

    2018-06-01

    The immune system is essential for the control and elimination of infections, and macrophages are cells that act as important players in orchestrating the various parts of the inflammatory/immune response. Amino acids play important role in mediating functionality of the inflammatory response, especially mediating macrophages functions and cytokines production. We investigated the influence of glutamine, taurine and their association on the modulation of inflammatory pathway markers in macrophages. The RAW 264.7 macrophage cell line was cultivated in the presence of glutamine and taurine and proliferation rates, cell viability, cell cycle phases, IL-1α, IL-6, IL-10 and TNF-α as well as H 2 O 2 production and the expression of the transcription factor, NFκB, and its inhibitor, IκBα, were evaluated. Our results showed an increase in viable cells and increased proliferation rates of cells treated with glutamine concentrations over 2 mM, as well as cells treated with both glutamine and taurine. The cell cycle showed a higher percentage of cells in the phases S, G2 and M when they were treated with 2 or 10 mM glutamine, or with glutamine and taurine in cells stimulated with lipopolysaccharide. The pNFκB/NFκB showed reduced ratio expression when cells were treated with 10 mM of glutamine or with glutamine in association with taurine. These conditions also resulted in reduced TNF-α, IL-1α and H 2 O 2 production, and higher production of IL-10. These findings demonstrate that glutamine and taurine are able to modulate macrophages inflammatory pathways, and that taurine can potentiate the effects of glutamine, illustrating their immunomodulatory properties.

  11. Influence of oral glutamine supplementation on survival outcomes of patients treated with concurrent chemoradiotherapy for locally advanced non-small cell lung cancer

    Topkan, Erkan; Parlak, Cem; Topuk, Savas; Pehlivan, Berrin

    2012-01-01

    Glutamine (Gln) supplementation during concurrent chemoradiotherapy (C-CRT) effectively reduces the incidence and severity of acute radiation-induced esophagitis (RIE). However, there are concerns that Gln might stimulate tumor growth, and therefore negatively impact the outcomes of anticancer treatment. We retrospectively investigated the effect of co-administration of oral Gln during C-CRT on survival outcomes of patients with stage IIIB non-small cell lung carcinoma (NSCLC). We additionally evaluated role of oral Gln in preventing C-CRT-induced weight change, acute and late toxicities. The study included 104 patients: 56 (53.8%) received prophylactic powdered Gln (Gln+) orally at a dose of 10 g/8 h and 48 (46.2%) did not receive Gln (Gln-) and served as controls. The prescribed radiation dose to the planning target volume was 66 Gy in 2-Gy fractions. Primary endpoints of progression-free survival (PFS), local/regional progression-free survival (LRPFS), and overall survival (OS) were correlated with status of Gln supplementation. Oral Gln was well tolerated except for mild nausea/vomiting in 14 (25.0%) patients. There was no C-CRT-related acute or late grade 4–5 toxicity. Administration of Gln was associated with a decrease in the incidence of grade 3 acute radiation-induced esophagitis (RIE) (7.2% vs. 16.7% for Gln+ vs. Gln-; p=0.02) and late-RIE (0% vs. 6.3%; p=0.06), a reduced need for unplanned treatment breaks (7.1% vs. 20.8%; p=0.04), and reduced incidence of weight loss (44.6% vs. 72.9%; p=0.002). At a median follow-up of 24.2 months (range 9.2-34.4) the median OS, LRPFS, and PFS for Gln+ vs. Gln- cohorts were 21.4 vs. 20.4 (p=0.35), 14.2 vs.11.3 (p=0.16), and 10.2 vs. 9.0 months (p=0.11), respectively. In our study, supplementation with Gln during C-CRT had no detectable negative impact on tumor control and survival outcomes in patients with Stage IIIB NSCLC. Furthermore, Gln appeared to have a beneficial effect with respect to prevention of weight loss

  12. Reductive glutamine metabolism by IDH1 mediates lipogenesis under hypoxia

    Metallo, Christian M.; Gameiro, Paulo A.; Bell, Eric L.; Mattaini, Katherine R.; Yang, Juanjuan; Hiller, Karsten; Jewell, Christopher M.; Johnson, Zachary R.; Irvine, Darrell J.; Guarente, Leonard; Kelleher, Joanne K.; Vander Heiden, Matthew G.; Iliopoulos, Othon; Stephanopoulos, Gregory

    2013-01-01

    Acetyl coenzyme A (AcCoA) is the central biosynthetic precursor for fatty acid synthesis and protein acetylation. In the conventional view of mammalian cell metabolism, AcCoA is primarily generated from glucose-derived pyruvate through the citrate shuttle and adenosine triphosphate citrate lyase (ACL) in the cytosol1-3. However, proliferating cells that exhibit aerobic glycolysis and those exposed to hypoxia convert glucose to lactate at near stoichiometric levels, directing glucose carbon away from the tricarboxylic acid cycle (TCA) and fatty acid synthesis4. Although glutamine is consumed at levels exceeding that required for nitrogen biosynthesis5, the regulation and utilization of glutamine metabolism in hypoxic cells is not well understood. Here we show that human cells employ reductive metabolism of alpha-ketoglutarate (αKG) to synthesize AcCoA for lipid synthesis. This isocitrate dehydrogenase 1 (IDH1) dependent pathway is active in most cell lines under normal culture conditions, but cells grown under hypoxia rely almost exclusively on the reductive carboxylation of glutamine-derived αKG for de novo lipogenesis. Furthermore, renal cell lines deficient in the von Hippel-Lindau (VHL) tumor suppressor protein preferentially utilize reductive glutamine metabolism for lipid biosynthesis even at normal oxygen levels. These results identify a critical role for oxygen in regulating carbon utilization in order to produce AcCoA and support lipid synthesis in mammalian cells. PMID:22101433

  13. The effect of glutamine infusion on the inflammatory response and HSP70 during human experimental endotoxaemia

    Andreasen, Anne Sofie; Pedersen-Skovsgaard, Theis; Mortensen, Ole Hartvig

    2009-01-01

    was associated with alterations in white blood cell and differential counts, tumour necrosis factor-alpha, IL-6, temperature and heart rate, but glutamine affected neither the endotoxin-induced change in these variables nor the expression of HSP70 in BMNCs. CONCLUSIONS: Endotoxin reduced plasma glutamine...

  14. Disruption of stomatal lineage signaling or transcriptional regulators has differential effects on mesophyll development, but maintains coordination of gas exchange.

    Dow, Graham J; Berry, Joseph A; Bergmann, Dominique C

    2017-10-01

    Stomata are simultaneously tasked with permitting the uptake of carbon dioxide for photosynthesis while limiting water loss from the plant. This process is mainly regulated by guard cell control of the stomatal aperture, but recent advancements have highlighted the importance of several genes that control stomatal development. Using targeted genetic manipulations of the stomatal lineage and a combination of gas exchange and microscopy techniques, we show that changes in stomatal development of the epidermal layer lead to coupled changes in the underlying mesophyll tissues. This coordinated response tends to match leaf photosynthetic potential (V cmax ) with gas-exchange capacity (g smax ), and hence the uptake of carbon dioxide for water lost. We found that different genetic regulators systematically altered tissue coordination in separate ways: the transcription factor SPEECHLESS (SPCH) primarily affected leaf size and thickness, whereas peptides in the EPIDERMAL PATTERNING FACTOR (EPF) family altered cell density in the mesophyll. It was also determined that interlayer coordination required the cell-surface receptor TOO MANY MOUTHS (TMM). These results demonstrate that stomata-specific regulators can alter mesophyll properties, which provides insight into how molecular pathways can organize leaf tissues to coordinate gas exchange and suggests new strategies for improving plant water-use efficiency. © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  15. Infection of potato mesophyll protoplasts with five plant viruses.

    Barker, H; Harrison, B D

    1982-12-01

    Methods are described for preparing potato mesophyll protoplasts that are suitable for infection with inocula of virus nucleoprotein or RNA. The protoplasts could be infected with four sap-transmissible viruses (tobacco mosaic, tobacco rattle, tobacco ringspot and tomato black ring viruses) and with potato leafroll virus, which is not saptransmissible. No differences were observed in ability to infect protoplasts with potato leafroll virus strains differing either in virulence in intact plants or in aphid transmissibility.

  16. Effect of L-glutamine levels in piglets diets challenged with Escherichia coli lipopolysacharides

    Arturo Pardo L.

    2014-09-01

    Full Text Available Objective. To evaluate the effect of different levels of L-glutamine on weaned and immunologically challenged piglets with Escherichia coli lipopolysaccharides (LPS on performance parameters, serum cortisol and defense cells. Materials and methods. Four levels of L –glutamine were evaluated (0, 1.0, 1.5, 2.0% as well as the addition, or no addition, of LPS (0.3μg. 96 piglets were used (48 castrated males and 48 females of Agroceres x PenArlan lineage, with an initial age of 21 days and 6.06±0.852 kg live weight. An experimental design was used on randomized blocks in a factorial setting 4 x 2 (levels of L- glutamine with or without challenge. Results. Cubic effect was shown for daily weight gain of unchallenged animals, and was better with the addition of 0.41% L- glutamine. Feed conversion improved with increased levels of L -glutamine for challenged animals. In the evaluation of defense cells, there was interaction of leukocytes with the levels of L- glutamine and the immune challenge. Eosinophils and lymphocytes showed a quadratic effect for the levels of L –glutamine, with a maximum value of 1.30% and 0.5%, respectively. Conclusions. L -glutamine supplementation of up to 2% in the diet improves feed conversion and favors the immune serum of weaned piglets challenged with LPS of E. coli.

  17. Glutamine deprivation enhances antitumor activity of 3-bromopyruvate through the stabilization of monocarboxylate transporter-1.

    Cardaci, Simone; Rizza, Salvatore; Filomeni, Giuseppe; Bernardini, Roberta; Bertocchi, Fabio; Mattei, Maurizio; Paci, Maurizio; Rotilio, Giuseppe; Ciriolo, Maria Rosa

    2012-09-01

    Anticancer drug efficacy might be leveraged by strategies to target certain biochemical adaptations of tumors. Here we show how depriving cancer cells of glutamine can enhance the anticancer properties of 3-bromopyruvate, a halogenated analog of pyruvic acid. Glutamine deprival potentiated 3-bromopyruvate chemotherapy by increasing the stability of the monocarboxylate transporter-1, an effect that sensitized cells to metabolic oxidative stress and autophagic cell death. We further elucidated mechanisms through which resistance to chemopotentiation by glutamine deprival could be circumvented. Overall, our findings offer a preclinical proof-of-concept for how to employ 3-bromopyruvate or other monocarboxylic-based drugs to sensitize tumors to chemotherapy. ©2012 AACR.

  18. Benzo[a]pyrene affects Jurkat T cells in the activated state via the antioxidant response element dependent Nrf2 pathway leading to decreased IL-2 secretion and redirecting glutamine metabolism

    Murugaiyan, Jayaseelan; Rockstroh, Maxie; Wagner, Juliane [Department of Proteomics, Helmholtz-Centre for Environmental Research — UFZ, Permoserstr. 15, 04318 Leipzig (Germany); Baumann, Sven [Department of Metabolomics, Helmholtz-Centre for Environmental Research — UFZ, Permoserstr. 15, 04318 Leipzig (Germany); Schorsch, Katrin [Department of Proteomics, Helmholtz-Centre for Environmental Research — UFZ, Permoserstr. 15, 04318 Leipzig (Germany); Trump, Saskia; Lehmann, Irina [Department of Environmental Immunology, Helmholtz-Centre for Environmental Research — UFZ, Permoserstr. 15, 04318 Leipzig (Germany); Bergen, Martin von [Department of Proteomics, Helmholtz-Centre for Environmental Research — UFZ, Permoserstr. 15, 04318 Leipzig (Germany); Department of Environmental Immunology, Helmholtz-Centre for Environmental Research — UFZ, Permoserstr. 15, 04318 Leipzig (Germany); Department of Biotechnology, Chemistry and Environmental Engineering, Aalborg University, Aalborg (Denmark); Tomm, Janina M., E-mail: Janina.tomm@ufz.de [Department of Proteomics, Helmholtz-Centre for Environmental Research — UFZ, Permoserstr. 15, 04318 Leipzig (Germany)

    2013-06-15

    There is a clear evidence that environmental pollutants, such as benzo[a]pyrene (B[a]P), can have detrimental effects on the immune system, whereas the underlying mechanisms still remain elusive. Jurkat T cells share many properties with native T lymphocytes and therefore are an appropriate model to analyze the effects of environmental pollutants on T cells and their activation. Since environmental compounds frequently occur at low, not acute toxic concentrations, we analyzed the effects of two subtoxic concentrations, 50 nM and 5 μM, on non- and activated cells. B[a]P interferes directly with the stimulation process as proven by an altered IL-2 secretion. Furthermore, B[a]P exposure results in significant proteomic changes as shown by DIGE analysis. Pathway analysis revealed an involvement of the AhR independent Nrf2 pathway in the altered processes observed in unstimulated and stimulated cells. A participation of the Nrf2 pathway in the change of IL-2 secretion was confirmed by exposing cells to the Nrf2 activator tBHQ. tBHQ and 5 μM B[a]P caused similar alterations of IL-2 secretion and glutamine/glutamate metabolism. Moreover, the proteome changes in unstimulated cells point towards a modified regulation of the cytoskeleton and cellular stress response, which was proven by western blotting. Additionally, there is a strong evidence for alterations in metabolic pathways caused by B[a]P exposure in stimulated cells. Especially the glutamine/glutamate metabolism was indicated by proteome pathway analysis and validated by metabolite measurements. The detrimental effects were slightly enhanced in stimulated cells, suggesting that stimulated cells are more vulnerable to the environmental pollutant model compound B[a]P. - Highlights: • B[a]P affects the proteome of Jurkat T cells also at low concentrations. • Exposure to B[a]P (50 nM, 5 μM) did not change Jurkat T cell viability. • Both B[a]P concentrations altered the IL-2 secretion of stimulated cells.

  19. Endogenous glutamine production in critically ill patients: the effect of exogenous glutamine supplementation.

    Mori, Maiko; Rooyackers, Olav; Smedberg, Marie; Tjäder, Inga; Norberg, Ake; Wernerman, Jan

    2014-04-14

    Glutamine rate of appearance (Ra) may be used as an estimate of endogenous glutamine production. Recently a technique employing a bolus injection of isotopically labeled glutamine was introduced, with the potential to allow for multiple assessments of the glutamine Ra over time in critically ill patients, who may not be as metabolically stable as healthy individuals. Here the technique was used to evaluate the endogenous glutamine production in critically ill patients in the fed state with and without exogenous glutamine supplementation intravenously. Mechanically ventilated patients (n = 11) in the intensive care unit (ICU) were studied on two consecutive days during continuous parenteral feeding. To allow the patients to be used as their own controls, they were randomized for the reference measurement during basal feeding without supplementation, before or after the supplementation period. Glutamine Ra was determined by a bolus injection of 13C-glutamine followed by a period of frequent sampling to establish the decay-curve for the glutamine tracer. Exogenous glutamine supplementation was given by intravenous infusion of a glutamine containing dipeptide, L-alanyl-L-glutamine, 0.28 g/kg during 20 hours. A 14% increase of endogenous glutamine Ra was seen at the end of the intravenous supplementation period as compared to the basal measurements (P = 0.009). The bolus injection technique to measure glutamine Ra to estimate the endogenous production of glutamine in critically ill patients was demonstrated to be useful for repetitive measurements. The hypothesized attenuation of endogenous glutamine production during L-alanyl-L-glutamine infusion given as a part of full nutrition was not seen.

  20. The glutamate/GABA-glutamine cycle

    Bak, Lasse K; Schousboe, Arne; Waagepetersen, Helle S

    2006-01-01

    Neurons are metabolically handicapped in the sense that they are not able to perform de novo synthesis of neurotransmitter glutamate and gamma-aminobutyric acid (GABA) from glucose. A metabolite shuttle known as the glutamate/GABA-glutamine cycle describes the release of neurotransmitter glutamate...... or GABA from neurons and subsequent uptake into astrocytes. In return, astrocytes release glutamine to be taken up into neurons for use as neurotransmitter precursor. In this review, the basic properties of the glutamate/GABA-glutamine cycle will be discussed, including aspects of transport and metabolism...... of intercellular transfer of ammonia produced in neurons (when glutamine is deamidated to glutamate) and utilized in astrocytes (for amidation of glutamate) when the glutamate/GABA-glutamine cycle is operating. A main objective of this review is to endorse the view that the glutamate/GABA-glutamine cycle must...

  1. Glutamine Triggers and Potentiates Glucagon-Like Peptide-1 Secretion by Raising Cytosolic Ca2+ and cAMP

    Tolhurst, Gwen; Zheng, Yue; Parker, Helen E.; Habib, Abdella M.; Reimann, Frank; Gribble, Fiona M.

    2011-01-01

    L-glutamine stimulates glucagon-like peptide 1 (GLP-1) secretion in human subjects and cell lines. As recent advances have enabled the study of primary GLP-1–releasing L cells, this study aimed to characterize glutamine-sensing pathways in native murine L cells. L cells were identified using transgenic mice with cell-specific expression of fluorescent markers. Cells were studied in primary colonic cultures from adult mice, or purified by flow cytometry for expression analysis. Intracellular C...

  2. Glutamine granule-supplemented enteral nutrition maintains immunological function in severely burned patients.

    Peng, Xi; Yan, Hong; You, Zhongyi; Wang, Pei; Wang, Shiliang

    2006-08-01

    Glutamine is an important energy source for immune cells. It is a necessary nutrient for cell proliferation, and serves as specific fuel for lymphocytes, macrophages, and enterocytes when it is present in appropriate concentrations. The purpose of this clinical study was to observe the effects of enteral nutrition supplemented with glutamine granules on immunologic function in severely burned patients. Forty-eight severely burned patients (total burn surface area 30-75%, full thickness burn area 20-58%) who met the requirements of the protocol joined this double-blind randomized controlled clinical trail. Patients were randomly divided into two groups: burn control group (B group, 23 patients) and glutamine treated group (Gln group, 25 patients). There was isonitrogenous and isocaloric intake in both groups, Gln and B group patents were given glutamine granules or placebo (glycine) at 0.5 g/kgd for 14 days with oral feeding or tube feeding, respectively. The plasma level of glutamine and several indices of immunologic function including lymphocyte transformation ratio, neutrophil phagocytosis index (NPI), CD4/CD8 ratio, the content of immunoglobulin, complement C3, C4 and IL-2 levels were determined. Moreover, wound healing rate of burn area was observed and then hospital stay was recorded. The results showed significantly reduced plasma glutamine and damaged immunological function after severe burn Indices of cellular immunity function were remarkably decreased from normal controls. After taking glutamine granules for 14 days, plasma glutamine concentration was significantly higher in Gln group than that in B group (607.86+/-147.25 micromol/L versus 447.63+/-132.38 micromol/L, P0.05). In addition, wound healing was better and hospital stay days were reduced in Gln group (46.59+/-12.98 days versus 55.68+/-17.36 days, Pburn; supplemented glutamine granules with oral feeding or tube feeding abate the degree of immunosuppression, improve immunological function

  3. Interaction of E. coli DNA with tobacco mesophyll protoplasts

    Heyn, R.F.

    1975-01-01

    This chapter is part of a dissertation dealing with the interaction of DNA with protoplasts. Having established the length of time during which tobacco mesophyll protoplasts do not synthesize DNA following their isolation, it is important to know the extent of DNA uptake just before the onset of DNA synthesis (and possible integration) and to find optimal conditions for this uptake. Therefore, the association of E. coli DNA with tobacco protoplasts was studied. Care should be taken with the interpretation of ''uptake'' results: adsorption phenomena play a very important role and may do so at the plasmalemma of naked protoplasts. To solve the problems involved, the use of radiation-damaged DNA was attempted. With E. coli DNA possessing a large number of thymine containing pyrimidine dimers, the loss of dimers from DNA recovered from treated protoplasts was tested in order to obtain an indication of ''real'' uptake. The results are reported

  4. Estimation of mesophyll conductance to CO2 flux by three different methods

    Loreto, F.; Harley, P.C.; Di Marco, G.; Sharkey, T.D.

    1992-01-01

    The resistance to diffusion of CO2 from the intercellular airspaces within the leaf through the mesophyll to the sites of carboxylation during photosynthesis was measured using three different techniques, The three techniques include a method based on discrimination against the heavy stable isotope of carbon, 13C, and two modeling methods. The methods rely upon different assumptions, but the estimates of mesophyll conductance were similar with all three methods. The mesophyll conductance of leaves from a number of species was about 1.4 times the stomatal conductance for CO2 diffusion determined in unstressed plants at high light. The relatively low CO2 partial pressure inside chloroplasts of plants with a low mesophyll conductance did not lead to enhanced O2 sensitivity of photosynthesis because the low conductance caused a significant drop in the chloroplast CO2 Partial pressure upon switching to low O2. We found no correlation between mesophyll conductance and the ratio of internal leaf area to leaf surface area and only a weak correlation between mesophyll conductance and the proportion of leaf volume occupied by air. Mesophyll conductance was independent of CO2 and O2 partial pressure during the measurement, indicating that a true physical parameter, independent of biochemical effects, was being measured. No evidence for accumulating mechanisms was found. Some plants, notably Citrus aurantium and Simmondsia chinensis, had very low conductances that limit the rate of photosynthesis these plants can attain at atmospheric CO2 level

  5. The metabolism of L-arginine and its significance for the biosynthesis of endothelium-derived relaxing factor: L-glutamine inhibits the generation of L-arginine by cultured endothelial cells

    Sessa, W.C.; Hecker, M.; Mitchell, J.A.; Vane, J.R.

    1990-01-01

    The mechanism by which L-glutamine (L-Gln) inhibits the release of endothelium-derived factor from bovine aortic cultured endothelial cells was investigated. The intracellular concentration of L-arginine (L-Arg) in Arg-depleted endothelial cells was inversely related to the level of L-Gln. Removal of L-Gln from the culture medium (usually containing L-Gln at 2 mM) abolished the inhibitory effect of the culture medium on L-Arg generation. L-Gln (0.2 and 2 mM) but not D-Gln inhibited the generation of L-Arg by both Arg-depleted and nondepleted endothelial cells. L-Gln did not interfere with the uptake of L-Arg or the metabolism of L-Arg-L-Phe to L-Arg but inhibited the formation of L-Arg from L-citrulline (L-Cit), L-Cit-L-Phe, and N G -monomethyl-L-arginine. L-Gln also inhibited the conversion of L-[ 14 C]Cit to L-[ 14 C]Arg by Arg-depleted endothelial cells. However, L-Gln did not inhibit the conversion of L-argininosuccinic acid to L-Arg by endothelial cell homogenates. Thus, L-Gln interferes with the conversion of L-Cit to L-Arg probably by acting on argininosuccinate synthetase rather than argininosuccinate lyase. L-Gln also inhibited the generation of L-Arg by the monocyte-macrophage cell line J774 but had no effect on the conversion of L-Cit to L-Arg by these cells. As the release of endothelium-derived relaxing factor from cultured and non-cultured endothelial cells is limited by the availability of L-Arg, endogenous L-Gln may play a regulatory role in the biosynthesis of endothelium-derived relaxing factor

  6. Early Administration of Glutamine Protects Cardiomyocytes from Post-Cardiac Arrest Acidosis

    Yan-Ren Lin

    2016-01-01

    Full Text Available Postcardiac arrest acidosis can decrease survival. Effective medications without adverse side effects are still not well characterized. We aimed to analyze whether early administration of glutamine could improve survival and protect cardiomyocytes from postcardiac arrest acidosis using animal and cell models. Forty Wistar rats with postcardiac arrest acidosis (blood pH < 7.2 were included. They were divided into study (500 mg/kg L-alanyl-L-glutamine, n=20 and control (normal saline, n=20 groups. Each of the rats received resuscitation. The outcomes were compared between the two groups. In addition, cardiomyocytes derived from human induced pluripotent stem cells were exposed to HBSS with different pH levels (7.3 or 6.5 or to culture medium (control. Apoptosis-related markers and beating function were analyzed. We found that the duration of survival was significantly longer in the study group (p<0.05. In addition, in pH 6.5 or pH 7.3 HBSS buffer, the expression levels of cell stress (p53 and apoptosis (caspase-3, Bcl-xL markers were significantly lower in cardiomyocytes treated with 50 mM L-glutamine than those without L-glutamine (RT-PCR. L-glutamine also increased the beating function of cardiomyocytes, especially at the lower pH level (6.5. More importantly, glutamine decreased cardiomyocyte apoptosis and increased these cells’ beating function at a low pH level.

  7. Differential gene expression and transport functionality in the bundle sheath versus mesophyll - a potential role in leaf mineral homeostasis.

    Wigoda, Noa; Pasmanik-Chor, Metsada; Yang, Tianyuan; Yu, Ling; Moshelion, Menachem; Moran, Nava

    2017-06-01

    Under fluctuating ambient conditions, the ability of plants to maintain hydromineral homeostasis requires the tight control of long distance transport. This includes the control of radial transport within leaves, from veins to mesophyll. The bundle sheath is a structure that tightly wraps around leaf vasculature. It has been suggested to act as a selective barrier in the context of radial transport. This suggestion is based on recent physiological transport assays of bundle sheath cells (BSCs), as well as the anatomy of these cells.We hypothesized that the unique transport functionality of BSCs is apparent in their transcriptome. To test this, we compared the transcriptomes of individually hand-picked protoplasts of GFP-labeled BSCs and non-labeled mesophyll cells (MCs) from the leaves of Arabidopsis thaliana. Of the 90 genes differentially expressed between BSCs and MCs, 45% are membrane related and 20% transport related, a prominent example being the proton pump AHA2. Electrophysiological assays showed that the major AKT2-like membrane K+ conductances of BSCs and MCs had different voltage dependency ranges. Taken together, these differences may cause simultaneous but oppositely directed transmembrane K+ fluxes in BSCs and MCs, in otherwise similar conditions. © The Author 2017. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  8. Novel efficient methods for measuring mesophyll anatomical characteristics from fresh thick sections using stereology and confocal microscopy: application on acid rain-treated Norway spruce needles

    Albrechtová, Jana; Janáček, Jiří; Lhotáková, Zuzana; Radochová, Barbora; Kubínová, Lucie

    2007-01-01

    Roč. 58, č. 6 (2007), s. 1451-1461 ISSN 0022-0957 R&D Projects: GA AV ČR IAA5011810; GA AV ČR(CZ) IAA600110507; GA MŠk(CZ) LC06063 Institutional research plan: CEZ:AV0Z50110509; CEZ:AV0Z60050516 Keywords : mesophyll * stereology * confocal microscopy Subject RIV: EA - Cell Biology Impact factor: 3.917, year: 2007

  9. Cytosolic glutamine synthetase in barley

    Thomsen, Hanne Cecilie

    remobilisation from ageing plant parts. Thus, GS is highly involved in determining crop yield and NUE. The major objective of this PhD project was to investigate the NUE properties of transgenic barley designed to constitutively overexpress a GS1 isogene (HvGS1.1). These transgenic lines exhibited an increased...... for N demand. Of the GS isogenes, only the transcript levels of root HvGS1.1 increased when plants were transferred from high to low N. This change coincided with an increase in total GS activity. Pronounced diurnal variation was observed for root nitrate transporter genes and GS isogenes in both root...... fertilizer requirement. The enzyme glutamine synthetase (GS) has been a major topic in plant nitrogen research for decades due to its central role in plant N metabolism. The cytosolic version of this enzyme (GS1) plays an important role in relation to primary N assimilation as well as in relation to N...

  10. Plasma Glutamine Concentrations in Liver Failure.

    Gunnel Helling

    Full Text Available Higher than normal plasma glutamine concentration at admission to an intensive care unit is associated with an unfavorable outcome. Very high plasma glutamine levels are sometimes seen in both acute and chronic liver failure. We aimed to systematically explore the relation between different types of liver failure and plasma glutamine concentrations.Four different groups of patients were studies; chronic liver failure (n = 40, acute on chronic liver failure (n = 20, acute fulminant liver failure (n = 20, and post-hepatectomy liver failure (n = 20. Child-Pugh and Model for End-stage Liver Disease (MELD scores were assessed as indices of liver function. All groups except the chronic liver failure group were followed longitudinally during hospitalisation. Outcomes were recorded up to 48 months after study inclusion.All groups had individuals with very high plasma glutamine concentrations. In the total group of patients (n = 100, severity of liver failure correlated significantly with plasma glutamine concentration, but the correlation was not strong.Liver failure, regardless of severity and course of illness, may be associated with a high plasma glutamine concentration. Further studies are needed to understand whether high glutamine levels should be regarded as a biomarker or as a contributor to symptomatology in liver failure.

  11. Glutamine supplementation maintains intramuscular glutamine concentrations and normalizes lymphocyte function in infected early weaned pigs.

    Yoo, S S; Field, C J; McBurney, M I

    1997-11-01

    Numerous studies in humans and rats have shown that glutamine supplementation during stressful conditions has favorable outcomes. However, the requirements for glutamine during weaning are unknown. Thus, the effects of glutamine supplementation in healthy and infected weaned pigs were investigated. At 21 d of age, pigs were weaned to an elemental diet supplemented with glutamine (+Gln) or an isonitrogenous diet containing nonessential amino acids (-Gln). At 26 d of age, pigs were intraperitoneally injected with Escherichia coli (+Ecoli) or buffered saline (-Ecoli) and killed at 28 d of age. Infection decreased (P Ecoli+Gln pigs were greater (P Ecoli-Gln pigs and not different than those of noninfected pigs. Hence, glutamine supplementation maintained muscular glutamine concentrations and normalized lymphocyte function in infected pigs.

  12. Introduction to the Glutamate-Glutamine Cycle

    Sonnewald, Ursula; Schousboe, Arne

    2016-01-01

    . This is metabolically impossible unless it is assumed that at least two distinct pools of these amino acids exist. This combined with the finding that the enzyme synthesizing glutamine from glutamate was expressed in astrocytes but not in neurons formed the basis of the notion that a cycle must exist in which glutamate......The term 'glutamate-glutamine cycle' was coined several decades ago based on the observation that using certain (14)C-labeled precursors for studies of brain metabolism the specific radioactivity of glutamine generated from glutamate was higher than that of glutamate, its immediate precursor...... released from neurons is transported into astrocytes, converted to glutamine which is subsequently returned to neurons and converted to glutamate by an enzyme the activity of which is much higher in neurons than in astrocytes. Originally this cycle was supposed to function in a stoichiometric fashion...

  13. effects of enteral glutamine supplementation on reduction

    Objective: To determine the effect of enteral glutamine in reducing the incidence of ... in use. These modalities include among others; topical antibacterial agents, early excision of eschar, and ... in the burns unit and plastic surgery ward 4D of.

  14. Introduction to the Glutamate-Glutamine Cycle

    Sonnewald, Ursula; Schousboe, Arne

    2016-01-01

    released from neurons is transported into astrocytes, converted to glutamine which is subsequently returned to neurons and converted to glutamate by an enzyme the activity of which is much higher in neurons than in astrocytes. Originally this cycle was supposed to function in a stoichiometric fashion......The term 'glutamate-glutamine cycle' was coined several decades ago based on the observation that using certain (14)C-labeled precursors for studies of brain metabolism the specific radioactivity of glutamine generated from glutamate was higher than that of glutamate, its immediate precursor....... This is metabolically impossible unless it is assumed that at least two distinct pools of these amino acids exist. This combined with the finding that the enzyme synthesizing glutamine from glutamate was expressed in astrocytes but not in neurons formed the basis of the notion that a cycle must exist in which glutamate...

  15. The Glutamine Transporters and Their Role in the Glutamate/GABA-Glutamine Cycle

    Leke, Renata; Schousboe, Arne

    2016-01-01

    in this neural communication, i.e., the transporters responsible for glutamine efflux from astrocytes and influx into the neurons, such as the members of the SNAT, LAT, y(+)LAT, and ASC families of transporters. The SNAT family consists of the transporter isoforms SNAT3 and SNAT5 that are related to efflux from......Glutamine is a key amino acid in the CNS, playing an important role in the glutamate/GABA-glutamine cycle (GGC). In the GGC, glutamine is transferred from astrocytes to neurons, where it will replenish the inhibitory and excitatory neurotransmitter pools. Different transporters participate...... the astrocytic compartment, and SNAT1 and SNAT2 that are associated with glutamine uptake into the neuronal compartment. The isoforms SNAT7 and SNAT8 do not have their role completely understood, but they likely also participate in the GGC. The isoforms LAT2 and y(+)LAT2 facilitate the exchange of neutral amino...

  16. The intestinal T cell response to alpha-gliadin in adult celiac disease is focused on a single deamidated glutamine targeted by tissue transglutaminase

    Arentz-Hansen, H; Körner, R; Molberg, O

    2000-01-01

    The great majority of patients that are intolerant of wheat gluten protein due to celiac disease (CD) are human histocompatibility leukocyte antigen (HLA)-DQ2(+), and the remaining few normally express HLA-DQ8. These two class II molecules are chiefly responsible for the presentation of gluten...... for T cell recognition. Gluten-specific T cell lines from 16 different adult patients all responded to one or both of these deamidated peptides, indicating that these epitopes are highly relevant to disease pathology. Binding studies showed that the deamidated peptides displayed an increased affinity...

  17. The utilization of glutamine by the retina: an autoradiographic and metabolic study

    Voaden, M.J.; Lake, N.; Marshall, J.; Morjaria, B.

    1978-01-01

    The cells able to accumulate exogenously applied [ 3 H] glutamine in rat, cat, frog, pigeon and guinea pig retinas have been located by autoradiography, and the fate of the labelled glutamine, as regards its incorporation into aspartic, glutamic and γ-amino-butyric acids, followed for 60 min. The results support the notion of glutamine as a precursor of transmitter amino acids in some neurones. In particular, it would appear to be a source of a relatively stable pool of GABA which may be located, with species variation, in amacrine or ganglion cells. In the pigeon retina glutamate pool incorporates and retains a major percentage of the label, and perikarya in the middle of the inner nuclear layer of the tissue are predominantly labelled. (author)

  18. Utilization of glutamine by the retina: an autoradiographic and metabolic study

    Voaden, M J; Lake, N; Marshall, J; Morjaria, B [Institute of Ophthalmology, London (UK). Dept. of Visual Science

    1978-10-01

    The cells able to accumulate exogenously applied (/sup 3/H) glutamine in rat, cat, frog, pigeon and guinea pig retinas have been located by autoradiography, and the fate of the labelled glutamine, as regards its incorporation into aspartic, glutamic and ..gamma..-amino-butyric acids, followed for 60 min. The results support the notion of glutamine as a precursor of transmitter amino acids in some neurones. In particular, it would appear to be a source of a relatively stable pool of GABA which may be located, with species variation, in amacrine or ganglion cells. In the pigeon retina glutamate pool incorporates and retains a major percentage of the label, and perikarya in the middle of the inner nuclear layer of the tissue are predominantly labelled.

  19. Introduction to the Glutamate-Glutamine Cycle.

    Sonnewald, Ursula; Schousboe, Arne

    2016-01-01

    The term 'glutamate-glutamine cycle' was coined several decades ago based on the observation that using certain 14 C-labeled precursors for studies of brain metabolism the specific radioactivity of glutamine generated from glutamate was higher than that of glutamate, its immediate precursor. This is metabolically impossible unless it is assumed that at least two distinct pools of these amino acids exist. This combined with the finding that the enzyme synthesizing glutamine from glutamate was expressed in astrocytes but not in neurons formed the basis of the notion that a cycle must exist in which glutamate released from neurons is transported into astrocytes, converted to glutamine which is subsequently returned to neurons and converted to glutamate by an enzyme the activity of which is much higher in neurons than in astrocytes. Originally this cycle was supposed to function in a stoichiometric fashion but more recent research has seriously questioned this.This volume of Advances in Neurobiology is intended to provide a detailed discussion of recent developments in research aimed at delineating the functional roles of the cycle taking into account that in order for this system to work there must be a tight coupling between metabolism of glutamate in astrocytes, transfer of glutamine to neurons and de novo synthesis of glutamine in astrocytes. To understand this, knowledge about the activity and regulation of the enzymes and transporters involved in these processes is required and as can be seen from the table of contents these issues will be dealt with in detail in the individual chapters of the book.

  20. [Effect of glutamine on small intestinal repair in weanling rats after chronic diarrhea].

    Huang, Zu-xiong; Ye, Li-yan; Zheng, Zhi-yong; Chen, Xin-min; Ren, Rong-na; Tong, Guo-yuan

    2005-05-01

    To investigate the nutrient effect of glutamine on small intestinal repair in weanling rats after chronic diarrhea. Forty 21-day-old wistar rats were randomly divided into five groups (8 in each). Animal model of chronic diarrhea was induced by a lactose enriched diet in the weanling Wistar rat, normal control group was fed with a standard semipurified diet, and after 14 days the rats in both groups were killed to test the establishment of the model. After the establishment of the model, the other groups were fed with the standard semipurified diet to recover for 7 days, and were randomly divided into three groups: non-intervention group, glutamine (Gln)-intervention group and control group. Glutamine concentrations in blood was detected by high-performance liquid chromatography (HPLC). Morphological changes including villus height and villus surface area of the jejunum were measured under a light microscope and electron microscope, expression of proliferating cell nuclear antigen (PCNA) as an index of cell proliferation was observed using immunohistochemical staining and image analysis. The diarrhea rate in model group was 100 percent, average diarrhea index was 1.16 +/- 0.06, but both diarrhea rate and average diarrhea index in control group were 0 (P body weight, plasma Gln concentration, villus height, villus surface area and expression of PCNA in non-intervened group compared with the control group (P body weight, villus height and villus surface area in Gln-intervened group compared with control group (P 0.05). And compared with non-intervened group, except for body weight (P > 0.05), plasma glutamine, villus height, villus surface area and expression of PCNA were all significantly increased in Gln-intervened group. Chronic diarrhea can induce malnutrition and reduce the villus height, villus surface area, expression of PCNA and plasm glutamine concentration. Oral glutamine could improve the proliferation of crypt cell and promote repair of intestinal mucosa

  1. The Glutamine Transporters and Their Role in the Glutamate/GABA-Glutamine Cycle

    Leke, Renata; Schousboe, Arne

    2016-01-01

    in this neural communication, i.e., the transporters responsible for glutamine efflux from astrocytes and influx into the neurons, such as the members of the SNAT, LAT, y(+)LAT, and ASC families of transporters. The SNAT family consists of the transporter isoforms SNAT3 and SNAT5 that are related to efflux from...... and translational mechanisms, which are induced by several determinants such as amino acid deprivation, hormones, pH, and the activity of different signaling pathways. Dysfunctional glutamine transporter activity has been associated with the pathophysiological mechanisms of certain neurologic diseases......, such as Hepatic Encephalopathy and Manganism. However, there might also be other neuropathological conditions associated with an altered GGC, in which glutamine transporters are dysfunctional. Hence, it appears to be of critical importance that the physiological and pathological aspects of glutamine transporters...

  2. effects of standard and/or glutamine dipeptide and/or omega-3 fatty

    2013-02-01

    Feb 1, 2013 ... effects of these to neutrophil functions and IL-8 levels are compared. Methods: ... acid solution to S-O group and TPN with omega 3 fatty acids solution and glutamine to S-D-O group ..... exudation involving multiple cell types.

  3. Glutamine methylation in histone H2A is an RNA-polymerase-I-dedicated modification

    Tessarz, Peter; Santos-Rosa, Helena; Robson, Sam C

    2014-01-01

    as the methyltransferase in yeast and demonstrate that fibrillarin is the orthologue enzyme in human cells. Glutamine methylation of H2A is restricted to the nucleolus. Global analysis in yeast, using an H2AQ105me-specific antibody, shows that this modification is exclusively enriched over the 35S ribosomal DNA...

  4. Supplementation with a combination of beta-hydroxy-beta-methylbutyrate (HMB), arginine, and glutamine is safe and could improve hematological parameters.

    Rathmacher, J A; Nissen, S; Panton, L; Clark, R H; Eubanks May, P; Barber, A E; D'Olimpio, J; Abumrad, N N

    2004-01-01

    Combining the amino acids arginine and glutamine with the leucine metabolite beta-hydroxy-beta-methylbutyrate (HMB) has been shown to reverse lean tissue loss in cancer and acquired immunodeficiency syndrome (AIDS) patients. Although each of these nutrients has been shown to be safe, the safety of this mixture has not been reported. Three double-blind studies examined the safety of the combination of HMB, arginine and glutamine on blood chemistries, hematology, emotional profile, and adverse events. Study 1 was conducted in healthy adult males (n = 34), study 2 was in HIV patients with AIDS-associated weight loss (n = 43), and study 3 was in cancer patients with wasting (n = 32). Volunteers were assigned to either a placebo or a mixture of 3 g HMB, 14 g arginine, and 14 g glutamine per day. Across the 3 studies, HMB, arginine, and glutamine supplementation was not associated with any adverse indicators of health. The only significant changes noted were positive indicators of health status. HMB, arginine, and glutamine supplementation was associated with an improvement in emotional profile (p = .05), a decreased feeling of weakness (p = .03), and increased red blood cells, hemoglobin, hematocrit, lymphocytes, and eosinophils (p HMB, arginine, and glutamine supplementation, which was possibly caused by the additional nitrogen consumed or to the fact that ureagenesis is influenced by arginine and glutamine supplementation. These results show that HMB, arginine, and glutamine can be safely used to treat muscle wasting associated with AIDS and cancer.

  5. Deprive to kill: Glutamine closes the gate to anticancer monocarboxylic drugs

    Cardaci, Simone; Ciriolo, Maria Rosa

    2012-01-01

    Killing properties of antitumor drugs can be enhanced by strategies targeting biochemical adaptations of cancer cells. Recently, we reported that depriving cancer cells of glutamine is a feasible approach to enhance antitumor effects of the alkylating analog of pyruvic acid, 3-bromopyruvate, which rely on the induction of autophagic cell death by metabolic-oxidative stress. 3-bromopyruvate chemopotentiation is the result of its increased intracellular uptake mediated by the monocarboxylate tr...

  6. Ammonium absorption mechanism of rice seedling roots and 15N-labelling pattern of their glutamine-amide group, 2

    Arima, Yasuhiro; Kumazawa, Kikuo

    1975-01-01

    The processes of producing glutamine and asparagine at the initial stage of the absorption and assimilation of ammonia in rice seedling roots were examined in relation to glutamic acid, aspartic acid and ammonia by 15 N-labelling method. When ( 15 NH 4 ) 2 SO 4 was absorbed into the roots, 15 N concentration appeared very high in glutamine-amide radical and ammonia. It was also higher in amide radical than in amino radical in both glutamine and asparagine, while 15 N concentration in the amino radical of glutamine and asparagine were far lower than that of corresponding glutamine acid and aspartic acid. From these facts, glutamine-amide radical seems to be produced directly from the ammonia in culture media at the contact point of root cells and the culture media, while there is some possibility that asparagine-amide radical is formed from other amino compounds than ammonia. Also the amino radical of aspartic acid seems to be produced not only by the transamination from glutamic acid but also by the reductive amination of oxalautic acid by ammonium. (Kobatake, H.)

  7. GLUTAMINE AND HYPERAMMONEMIC CRISES IN PATIENTS WITH UREA CYCLE DISORDERS

    Lee, B.; Diaz, G.A.; Rhead, W.; Lichter-Konecki, U.; Feigenbaum, A.; Berry, S.A.; Le Mons, C.; Bartley, J.; Longo, N.; Nagamani, S.C.; Berquist, W.; Gallagher, R.C.; Harding, C.O.; McCandless, S.E.; Smith, W.; Schulze, A.; Marino, M.; Rowell, R.; Coakley, D.F.; Mokhtarani, M.; Scharschmidt, B.F.

    2016-01-01

    Blood ammonia and glutamine levels are used as biomarkers of control in patients with urea cycle disorders (UCDs). This study was undertaken to evaluate glutamine variability and utility as a predictor of hyperammonemic crises (HACs) in UCD patients. Methods The relationships between glutamine and ammonia levels and the incidence and timing of HACs were evaluated in over 100 adult and pediatric UCD patients who participated in clinical trials of glycerol phenylbutyrate. Results The median (range) intra-subject 24-hour coefficient of variation for glutamine was 15% (8–29%) as compared with 56% (28%–154%) for ammonia, and the correlation coefficient between glutamine and concurrent ammonia levels varied from 0.17 to 0.29. Patients with baseline (fasting) glutamine values >900 µmol/L had higher baseline ammonia levels (mean [SD]: 39.6 [26.2] µmol/L) than patients with baseline glutamine ≤900 µmol/L (26.6 [18.0] µmol/L). Glutamine values >900 µmol/L during the study were associated with an approximately 2-fold higher HAC risk (odds ratio [OR]=1.98; p=0.173). However, glutamine lost predictive significance (OR=1.47; p=0.439) when concomitant ammonia was taken into account, whereas the predictive value of baseline ammonia ≥ 1.0 upper limit of normal (ULN) was highly statistically significant (OR=4.96; p=0.013). There was no significant effect of glutamine >900 µmol/L on time to first HAC crisis (hazard ratio [HR]=1.14; p=0.813), but there was a significant effect of baseline ammonia ≥ 1.0 ULN (HR=4.62; p=0.0011). Conclusions The findings in this UCD population suggest that glutamine is a weaker predictor of HACs than ammonia and that the utility of the predictive value of glutamine will need to take into account concurrent ammonia levels. PMID:26586473

  8. Effects of L-glutamine supplementation on the myenteric neurons from the duodenum and cecum of diabetic rats

    Jacqueline Nelisis Zanoni

    2011-03-01

    Full Text Available CONTEXT: Peripheral neuropathy is one of the chronic complications of diabetes mellitus and is directly related to gastrointestinal consequences of the disease. Myenteric neurons are affected in some pathological conditions such as diabetic neuropathy. The imbalance between cellular antioxidants and free radicals, leading to an increase in oxidative stress, is considered one of the main factors responsible for neuronal damages in diabetes. Drugs that reduce the oxidative stress may play a significant role in the treatment of neurological complications of diabetes mellitus. OBJECTIVE: To evaluate the effect of L-glutamine supplementation on the myenteric neurons from the cecum and duodenum of Wistar rats with streptozotocin-induced diabetes mellitus. METHODS: The animals were divided in four groups (n = 5: non-treated normoglycemics, normoglycemics treated with L-glutamine, non-treated diabetics and diabetics treated with L-glutamine from the 4th day of diabetes induction on. The amino acid L-glutamine was added to their diet at 1%. Giemsa's technique was employed to stain the myenteric neurons. We determined the cell body area of 500 neurons in each group studied. The quantitative analysis was performed by sampling in an area of 16.6 mm² in the cecum and 3.6 mm² in the duodenum of each animal. RESULTS: After the supplementation with L-glutamine in the duodenum, we observed a preservation of neuronal density in groups normoglycemic and diabetic (P<0.05. We also observed a preservation of the cell bodies area in diabetic animals (group treated with L-glutamine (P<0.05. In the cecum, that preservation was not evident. CONCLUSION: Supplementation with L-glutamine (1% promoted a neuroprotective effect on the myenteric neurons from the duodenum of rats, both in terms of natural aging and of diabetes mellitus.

  9. Glutamine-Elicited Secretion of Glucagon-Like Peptide 1 Is Governed by an Activated Glutamate Dehydrogenase.

    Andersson, Lotta E; Shcherbina, Liliya; Al-Majdoub, Mahmoud; Vishnu, Neelanjan; Arroyo, Claudia Balderas; Aste Carrara, Jonathan; Wollheim, Claes B; Fex, Malin; Mulder, Hindrik; Wierup, Nils; Spégel, Peter

    2018-03-01

    Glucagon-like peptide 1 (GLP-1), secreted from intestinal L cells, glucose dependently stimulates insulin secretion from β-cells. This glucose dependence prevents hypoglycemia, rendering GLP-1 analogs a useful and safe treatment modality in type 2 diabetes. Although the amino acid glutamine is a potent elicitor of GLP-1 secretion, the responsible mechanism remains unclear. We investigated how GLP-1 secretion is metabolically coupled in L cells (GLUTag) and in vivo in mice using the insulin-secreting cell line INS-1 832/13 as reference. A membrane-permeable glutamate analog (dimethylglutamate [DMG]), acting downstream of electrogenic transporters, elicited similar alterations in metabolism as glutamine in both cell lines. Both DMG and glutamine alone elicited GLP-1 secretion in GLUTag cells and in vivo, whereas activation of glutamate dehydrogenase (GDH) was required to stimulate insulin secretion from INS-1 832/13 cells. Pharmacological inhibition in vivo of GDH blocked secretion of GLP-1 in response to DMG. In conclusion, our results suggest that nonelectrogenic nutrient uptake and metabolism play an important role in L cell stimulus-secretion coupling. Metabolism of glutamine and related analogs by GDH in the L cell may explain why GLP-1 secretion, but not that of insulin, is activated by these secretagogues in vivo. © 2017 by the American Diabetes Association.

  10. Protective effects of L-glutamine against toxicity of deltamethrin in the cerebral tissue

    Varol S

    2016-04-01

    Full Text Available Sefer Varol, Hasan Hüseyin Özdemir, Mehmet Uğur Çevik, Yaşar Altun, Ibrahim Ibiloğlu, Aysun Ekinci, Aslıhan Okan Ibiloğlu, Metin Balduz, Demet Arslan, Recep Tekin, Fesih Aktar, Mehmet Ufuk Aluçlu Department of Neurology, Faculty of Medicine, Dicle University, Diyarbakir, Turkey Background: Deltamethrin (DLM is a broad-spectrum synthetic dibromo-pyrethroid pesticide that is widely used for agricultural and veterinary purposes. However, human exposure to the pesticide leads to neurotoxicity. Glutamine is one of the principal, free intracellular amino acids and may also be an antioxidant. This study was undertaken in order to examine the neuroprotective and antioxidant potential of L-glutamine against DLM toxicity in female Wistar albino rats. Materials and methods: The rats were divided into the following groups (n=10: Group I: control (distilled water; 10 mL/kg, po one dose, Group II: L-glutamine (1.5 g/kg, po one dose, Group III: DLM (35 mg/kg, po one dose, and Group IV: DLM (35 mg/kg, po one dose and L-glutamine (1.5 g/kg, po one dose after 4 hours. Total oxidant status (TOS, total antioxidant status (TAS, tumor necrosis factor-α, interleukin (IL-1β, and IL-6 levels and apoptosis were evaluated in brain tissue. Results: DLM-treated animals had a significant increase in brain biochemical parameters, as well as TOS and TAS. Furthermore, the histopathological examination showed neuronal cell degeneration in the cerebral tissue. L-Glutamine treatment decreased the elevated brain levels of TOS and neuronal cell degeneration. There was no difference in tumor necrosis factor-α, IL-1β, and IL-6 levels between the groups. Conclusion: L-Glutamine may reduce the toxic effects of DLM in the cerebral tissue through antioxidant properties. Keywords: deltamethrin, L-glutamine, rat

  11. Glutamine Supplementation in Sick Children: Is It Beneficial?

    Elise Mok

    2011-01-01

    Full Text Available The purpose of this review is to provide a critical appraisal of the literature on Glutamine (Gln supplementation in various conditions or illnesses that affect children, from neonates to adolescents. First, a general overview of the proposed mechanisms for the beneficial effects of Gln is provided, and subsequently clinical studies are discussed. Despite safety, studies are conflicting, partly due to different effects of enteral and parenteral Gln supplementation. Further insufficient evidence is available on the benefits of Gln supplementation in pediatric patients. This includes premature infants, infants with gastrointestinal disease, children with Crohn's disease, short bowel syndrome, malnutrition/diarrhea, cancer, severe burns/trauma, Duchenne muscular dystrophy, sickle cell anemia, cystic fibrosis, and type 1 diabetes. Moreover, methodological issues have been noted in some studies. Further mechanistic data is needed along with large randomized controlled trials in select populations of sick children, who may eventually benefit from supplemental Gln.

  12. The cell wall stress response in Aspergillus niger involves increased expression of the glutamine: Fructose-6-phosphate amidotransferase-encoding gene (gfaA) and increased deposition of chitin in the cell wall

    Ram, A.F.J.; Arentshorst, M.; Damveld, R.A.; Kuyk, P.A. van; Klis, F.M.; Hondel, C.A.M.J.J. van den

    2004-01-01

    Perturbation of cell wall synthesis in Saccharomyces cerevisiae, either by mutations in cell wall synthesis-related genes or by adding compounds that interfere with normal cell wall assembly, triggers a compensatory response to ensure cell wall integrity. This response includes an increase in chitin

  13. Antagonizing Bcl-2 family members sensitizes neuroblastoma and Ewing's sarcoma to an inhibitor of glutamine metabolism.

    Rachelle R Olsen

    Full Text Available Neuroblastomas (NBL and Ewing's sarcomas (EWS together cause 18% of all pediatric cancer deaths. Though there is growing interest in targeting the dysregulated metabolism of cancer as a therapeutic strategy, this approach has not been fully examined in NBL and EWS. In this study, we first tested a panel of metabolic inhibitors and identified the glutamine antagonist 6-diazo-5-oxo-L-norleucine (DON as the most potent chemotherapeutic across all NBL and EWS cell lines tested. Myc, a master regulator of metabolism, is commonly overexpressed in both of these pediatric malignancies and recent studies have established that Myc causes cancer cells to become "addicted" to glutamine. We found DON strongly inhibited tumor growth of multiple tumor lines in mouse xenograft models. In vitro, inhibition of caspases partially reversed the effects of DON in high Myc expressing cell lines, but not in low Myc expressing lines. We further showed that induction of apoptosis by DON in Myc-overexpressing cancers is via the pro-apoptotic factor Bax. To relieve inhibition of Bax, we tested DON in combination with the Bcl-2 family antagonist navitoclax (ABT-263. In vitro, this combination caused an increase in DON activity across the entire panel of cell lines tested, with synergistic effects in two of the N-Myc amplified neuroblastoma cell lines. Our study supports targeting glutamine metabolism to treat Myc overexpressing cancers, such as NBL and EWS, particularly in combination with Bcl-2 family antagonists.

  14. Glutamine Synthetase Deficiency in Murine Astrocytes Results in Neonatal Death

    He, Youji; Hakvoort, Theodorus B. M.; Vermeulen, Jacqueline L. M.; Labruyère, Wilhelmina T.; de Waart, D. Rudi; van der Hel, W. Saskia; Ruijter, Jan M.; Uylings, Harry B. M.; Lamers, Wouter H.

    2010-01-01

    Glutamine synthetase (GS) is a key enzyme in the "glutamine-glutamate cycle" between astrocytes and neurons, but its function in vivo was thus far tested only pharmacologically. Crossing GS(fl/lacZ) or GS(fl/f)l mice with hGFAP-Cre mice resulted in prenatal excision of the GS(fl) allele in

  15. Large-scale protein-protein interaction analysis in Arabidopsis mesophyll protoplasts by split firefly luciferase complementation.

    Li, Jian-Feng; Bush, Jenifer; Xiong, Yan; Li, Lei; McCormack, Matthew

    2011-01-01

    Protein-protein interactions (PPIs) constitute the regulatory network that coordinates diverse cellular functions. There are growing needs in plant research for creating protein interaction maps behind complex cellular processes and at a systems biology level. However, only a few approaches have been successfully used for large-scale surveys of PPIs in plants, each having advantages and disadvantages. Here we present split firefly luciferase complementation (SFLC) as a highly sensitive and noninvasive technique for in planta PPI investigation. In this assay, the separate halves of a firefly luciferase can come into close proximity and transiently restore its catalytic activity only when their fusion partners, namely the two proteins of interest, interact with each other. This assay was conferred with quantitativeness and high throughput potential when the Arabidopsis mesophyll protoplast system and a microplate luminometer were employed for protein expression and luciferase measurement, respectively. Using the SFLC assay, we could monitor the dynamics of rapamycin-induced and ascomycin-disrupted interaction between Arabidopsis FRB and human FKBP proteins in a near real-time manner. As a proof of concept for large-scale PPI survey, we further applied the SFLC assay to testing 132 binary PPIs among 8 auxin response factors (ARFs) and 12 Aux/IAA proteins from Arabidopsis. Our results demonstrated that the SFLC assay is ideal for in vivo quantitative PPI analysis in plant cells and is particularly powerful for large-scale binary PPI screens.

  16. Large-scale protein-protein interaction analysis in Arabidopsis mesophyll protoplasts by split firefly luciferase complementation.

    Jian-Feng Li

    Full Text Available Protein-protein interactions (PPIs constitute the regulatory network that coordinates diverse cellular functions. There are growing needs in plant research for creating protein interaction maps behind complex cellular processes and at a systems biology level. However, only a few approaches have been successfully used for large-scale surveys of PPIs in plants, each having advantages and disadvantages. Here we present split firefly luciferase complementation (SFLC as a highly sensitive and noninvasive technique for in planta PPI investigation. In this assay, the separate halves of a firefly luciferase can come into close proximity and transiently restore its catalytic activity only when their fusion partners, namely the two proteins of interest, interact with each other. This assay was conferred with quantitativeness and high throughput potential when the Arabidopsis mesophyll protoplast system and a microplate luminometer were employed for protein expression and luciferase measurement, respectively. Using the SFLC assay, we could monitor the dynamics of rapamycin-induced and ascomycin-disrupted interaction between Arabidopsis FRB and human FKBP proteins in a near real-time manner. As a proof of concept for large-scale PPI survey, we further applied the SFLC assay to testing 132 binary PPIs among 8 auxin response factors (ARFs and 12 Aux/IAA proteins from Arabidopsis. Our results demonstrated that the SFLC assay is ideal for in vivo quantitative PPI analysis in plant cells and is particularly powerful for large-scale binary PPI screens.

  17. Structure of the Dispase Autolysis-inducing Protein from Streptomyces mobaraensis and Glutamine Cross-linking Sites for Transglutaminase.

    Fiebig, David; Schmelz, Stefan; Zindel, Stephan; Ehret, Vera; Beck, Jan; Ebenig, Aileen; Ehret, Marina; Fröls, Sabrina; Pfeifer, Felicitas; Kolmar, Harald; Fuchsbauer, Hans-Lothar; Scrima, Andrea

    2016-09-23

    Transglutaminase from Streptomyces mobaraensis (MTG) is an important enzyme for cross-linking and modifying proteins. An intrinsic substrate of MTG is the dispase autolysis-inducing protein (DAIP). The amino acid sequence of DAIP contains 5 potential glutamines and 10 lysines for MTG-mediated cross-linking. The aim of the study was to determine the structure and glutamine cross-linking sites of the first physiological MTG substrate. A production procedure was established in Escherichia coli BL21 (DE3) to obtain high yields of recombinant DAIP. DAIP variants were prepared by replacing four of five glutamines for asparagines in various combinations via site-directed mutagenesis. Incorporation of biotin cadaverine revealed a preference of MTG for the DAIP glutamines in the order of Gln-39 ≫ Gln-298 > Gln-345 ∼ Gln-65 ≫ Gln-144. In the structure of DAIP the preferred glutamines do cluster at the top of the seven-bladed β-propeller. This suggests a targeted cross-linking of DAIP by MTG that may occur after self-assembly in the bacterial cell wall. Based on our biochemical and structural data of the first physiological MTG substrate, we further provide novel insight into determinants of MTG-mediated modification, specificity, and efficiency. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Abscisic Acid Induces Rapid Reductions in Mesophyll Conductance to Carbon Dioxide.

    Giuseppe Sorrentino

    Full Text Available The rate of photosynthesis (A of plants exposed to water deficit is a function of stomatal (gs and mesophyll (gm conductance determining the availability of CO2 at the site of carboxylation within the chloroplast. Mesophyll conductance often represents the greatest impediment to photosynthetic uptake of CO2, and a crucial determinant of the photosynthetic effects of drought. Abscisic acid (ABA plays a fundamental role in signalling and co-ordination of plant responses to drought; however, the effect of ABA on gm is not well-defined. Rose, cherry, olive and poplar were exposed to exogenous ABA and their leaf gas exchange parameters recorded over a four hour period. Application with ABA induced reductions in values of A, gs and gm in all four species. Reduced gm occurred within one hour of ABA treatment in three of the four analysed species; indicating that the effect of ABA on gm occurs on a shorter timescale than previously considered. These declines in gm values associated with ABA were not the result of physical changes in leaf properties due to altered turgor affecting movement of CO2, or caused by a reduction in the sub-stomatal concentration of CO2 (Ci. Increased [ABA] likely induces biochemical changes in the properties of the interface between the sub-stomatal air-space and mesophyll layer through the actions of cooporins to regulate the transport of CO2. The results of this study provide further evidence that gm is highly responsive to fluctuations in the external environment, and stress signals such as ABA induce co-ordinated modifications of both gs and gm in the regulation of photosynthesis.

  19. Excess Diffuse Light Absorption in Upper Mesophyll Limits CO2 Drawdown and Depresses Photosynthesis.

    Earles, J Mason; Théroux-Rancourt, Guillaume; Gilbert, Matthew E; McElrone, Andrew J; Brodersen, Craig R

    2017-06-01

    In agricultural and natural systems, diffuse light can enhance plant primary productivity due to deeper penetration into and greater irradiance of the entire canopy. However, for individual sun-grown leaves from three species, photosynthesis is actually less efficient under diffuse compared with direct light. Despite its potential impact on canopy-level productivity, the mechanism for this leaf-level diffuse light photosynthetic depression effect is unknown. Here, we investigate if the spatial distribution of light absorption relative to electron transport capacity in sun- and shade-grown sunflower ( Helianthus annuus ) leaves underlies its previously observed diffuse light photosynthetic depression. Using a new one-dimensional porous medium finite element gas-exchange model parameterized with light absorption profiles, we found that weaker penetration of diffuse versus direct light into the mesophyll of sun-grown sunflower leaves led to a more heterogenous saturation of electron transport capacity and lowered its CO 2 concentration drawdown capacity in the intercellular airspace and chloroplast stroma. This decoupling of light availability from photosynthetic capacity under diffuse light is sufficient to generate an 11% decline in photosynthesis in sun-grown but not shade-grown leaves, primarily because thin shade-grown leaves similarly distribute diffuse and direct light throughout the mesophyll. Finally, we illustrate how diffuse light photosynthetic depression could overcome enhancement in canopies with low light extinction coefficients and/or leaf area, pointing toward a novel direction for future research. © 2017 American Society of Plant Biologists. All Rights Reserved.

  20. Patterns of indole alkaloids synthesis in response to heat shock, 5-azacytidine and Na-butyrate treatment of cultured catharanthus roseus mesophyll protoplasts

    Saleem, M.; Cutler, A.J.

    1986-01-01

    Alkaloids of C. roseus are in high demand for therapeutic and other reasons. Cultured Catharanthus cells can produce limited quantities of these alkaloids. The authors have found that cultured mesophyll protoplasts in the presence of 14 C-Tryptamine are capable of synthesizing alkaloids. The pattern of alkaloids synthesis changes when protoplasts are subjected to a heat shock at 37 0 C. The heat shocked protoplasts incorporated 33% more 14 C-Tryptamine and produced 3 new types of alkaloids. Treatment of protoplasts with 5-azacytidine, a DNA hypomethylating agent and Na-butyrate which induces hyperacetylation of histones produced qualitative and quantitative changes in the alkaloid pattern. Four new alkaloids following the above treatments were detected by TLC and HPLC of the extracts. It is suggested that the alkaloid pattern of the cultured protoplasts can be altered by treatment with compounds known as regulators of gene expression. Work is in progress to isolate and identify these new alkaloids

  1. Effects of honey, glutamine and their combination on canine small ...

    olayemitoyin

    and adaptation, evidenced by increased in residual bowel Villi height (27.71µm), ... advantage over glutamine or honey and may be a preferred treatment for short bowel syndrome patients. .... good general health based on complete clinical.

  2. Protoplast isolation, transient transformation of leaf mesophyll protoplasts and improved Agrobacterium-mediated leaf disc infiltration of Phaseolus vulgaris: tools for rapid gene expression analysis.

    Nanjareddy, Kalpana; Arthikala, Manoj-Kumar; Blanco, Lourdes; Arellano, Elizabeth S; Lara, Miguel

    2016-06-24

    Phaseolus vulgaris is one of the most extensively studied model legumes in the world. The P. vulgaris genome sequence is available; therefore, the need for an efficient and rapid transformation system is more imperative than ever. The functional characterization of P. vulgaris genes is impeded chiefly due to the non-amenable nature of Phaseolus sp. to stable genetic transformation. Transient transformation systems are convenient and versatile alternatives for rapid gene functional characterization studies. Hence, the present work focuses on standardizing methodologies for protoplast isolation from multiple tissues and transient transformation protocols for rapid gene expression analysis in the recalcitrant grain legume P. vulgaris. Herein, we provide methodologies for the high-throughput isolation of leaf mesophyll-, flower petal-, hypocotyl-, root- and nodule-derived protoplasts from P. vulgaris. The highly efficient polyethylene glycol-mannitol magnesium (PEG-MMG)-mediated transformation of leaf mesophyll protoplasts was optimized using a GUS reporter gene. We used the P. vulgaris SNF1-related protein kinase 1 (PvSnRK1) gene as proof of concept to demonstrate rapid gene functional analysis. An RT-qPCR analysis of protoplasts that had been transformed with PvSnRK1-RNAi and PvSnRK1-OE vectors showed the significant downregulation and ectopic constitutive expression (overexpression), respectively, of the PvSnRK1 transcript. We also demonstrated an improved transient transformation approach, sonication-assisted Agrobacterium-mediated transformation (SAAT), for the leaf disc infiltration of P. vulgaris. Interestingly, this method resulted in a 90 % transformation efficiency and transformed 60-85 % of the cells in a given area of the leaf surface. The constitutive expression of YFP further confirmed the amenability of the system to gene functional characterization studies. We present simple and efficient methodologies for protoplast isolation from multiple P

  3. Glutamine and glutamate: Nonessential or essential amino acids?

    Malcolm Watford

    2015-09-01

    Full Text Available Glutamine and glutamate are not considered essential amino acids but they play important roles in maintaining growth and health in both neonates and adults. Although glutamine and glutamate are highly abundant in most feedstuffs there is increasing evidence that they may be limiting during pregnancy, lactation and neonatal growth, particularly when relatively low protein diets are fed. Supplementation of diets with glutamine, glutamate or both at 0.5 to 1.0% to both suckling and recently weaned piglets improves intestinal and immune function and results in better growth. In addition such supplementation to the sow prevents some of the loss of lean body mass during lactation, and increases milk glutamine content. However, a number of important questions related to physiological condition, species under study and the form and amount of the supplements need to be addressed before the full benefits of glutamine and glutamate supplementation in domestic animal production can be realized. Keywords: Amino acid, Glutamate, Glutamine, Lactation, Pregnancy, Growth

  4. Effect of rapid weight loss and glutamine supplementation on immunosuppression of combat athletes: a double-blind, placebo-controlled study.

    Tritto, Aline C C; Amano, Mariane T; De Cillo, Maria E; Oliveira, Vinicius A; Mendes, Sandro H; Yoshioka, Caroline; Roschel, Hamilton; Camara, Niels Olsen S; Gualano, Bruno; Artioli, Guilherme G

    2018-02-01

    The role of plasma glutamine concentration and glutamine supplementation on immunosuppression was investigated in combat athletes. Twenty-three male athletes were randomly assigned to receive glutamine (21 g/day, n=12) or placebo (ovalbumin, n=11) for 10 days. Six athletes who did not lose weight served as controls. Athletes were assessed 21 days before (-21d), 1 day before (-1d) and 5 days after (+5d) a competition. Weight reduction was similar between glutamine (-8.2%± 4.1%) and placebo (-8.5%±2.4%) and negligible in control (-0.6%±1.4%). In both weight-loss groups, the majority of athletes reported symptoms of upper respiratory symptoms, as assessed by the Wisconsin upper respiratory symptom survey questionnaire. Only two athletes reported symptoms in the control group. Immune cell function remained unchanged throughout the study except for an increase in neutrophil phagocytic activity (placebo: -21d=5,251±2,986; -1d=17,428±22,374; +5d=21,125±21,934; glutamine: -21d=6,096±3,549; -1d=11,029±17,113; +5d=28,186±21,032 FI) and a minor change in monocyte phagocytic activity (placebo: -21d=4,421±3,634; -1d=3,329±6,283; +5d=3,243± 2,553; glutamine: -21d=4,051±3,186; -1d=3,106±2,625; +5d=4,981± 4,598) in both glutamine and placebo after weight loss. Plasma glutamine and cortisol remained unchanged across the study. creatine kinase levels were increased in placebo (-21d=125.2±54.1; -1d=187.2± 73.5; +5d=111.3±59.1 U/L) but not in glutamine (-21d=136.2±58.2; -1d= 168.8±65.0; +5d=129.7±64.0 U/L). Rapid weight loss increased the frequency and severity of infection symptoms, but this was neither associated with plasma glutamine depletion nor counteracted by glutamine supplementation.

  5. Glutamine methylation in histone H2A is an RNA-polymerase-I-dedicated modification

    Tessarz, Peter; Santos-Rosa, Helena; Robson, Sam C.; Sylvestersen, Kathrine B.; Nelson, Christopher J.; Nielsen, Michael L.; Kouzarides, Tony

    2014-01-01

    Nucleosomes are decorated with numerous post-translational modifications capable of influencing many DNA processes. Here we describe a new class of histone modification, methylation of glutamine, occurring on yeast histone H2A at position 105 (Q105) and human H2A at Q104. We identify Nop1 as the methyltransferase in yeast and demonstrate that fibrillarin is the orthologue enzyme in human cells. Glutamine methylation of H2A is restricted to the nucleolus. Global analysis in yeast, using an H2AQ105me-specific antibody, shows that this modification is exclusively enriched over the 35S ribosomal DNA transcriptional unit. We show that the Q105 residue is part of the binding site for the histone chaperone FACT (facilitator of chromatin transcription) complex. Methylation of Q105 or its substitution to alanine disrupts binding to FACT in vitro. A yeast strain mutated at Q105 shows reduced histone incorporation and increased transcription at the ribosomal DNA locus. These features are phenocopied by mutations in FACT complex components. Together these data identify glutamine methylation of H2A as the first histone epigenetic mark dedicated to a specific RNA polymerase and define its function as a regulator of FACT interaction with nucleosomes.

  6. Lymphocyte Glucose and Glutamine Metabolism as Targets of the Anti-Inflammatory and Immunomodulatory Effects of Exercise

    Frederick Wasinski

    2014-01-01

    Full Text Available Glucose and glutamine are important energetic and biosynthetic nutrients for T and B lymphocytes. These cells consume both nutrients at high rates in a function-dependent manner. In other words, the pathways that control lymphocyte function and survival directly control the glucose and glutamine metabolic pathways. Therefore, lymphocytes in different functional states reprogram their glucose and glutamine metabolism to balance their requirement for ATP and macromolecule production. The tight association between metabolism and function in these cells was suggested to introduce the possibility of several pathologies resulting from the inability of lymphocytes to meet their nutrient demands under a given condition. In fact, disruptions in lymphocyte metabolism and function have been observed in different inflammatory, metabolic, and autoimmune pathologies. Regular physical exercise and physical activity offer protection against several chronic pathologies, and this benefit has been associated with the anti-inflammatory and immunomodulatory effects of exercise/physical activity. Chronic exercise induces changes in lymphocyte functionality and substrate metabolism. In the present review, we discuss whether the beneficial effects of exercise on lymphocyte function in health and disease are associated with modulation of the glucose and glutamine metabolic pathways.

  7. New perspectives on glutamine synthetase in grasses.

    Swarbreck, Stéphanie M; Defoin-Platel, M; Hindle, M; Saqi, M; Habash, Dimah Z

    2011-02-01

    Members of the glutamine synthetase (GS) gene family have now been characterized in many crop species such as wheat, rice, and maize. Studies have shown that cytosolic GS isoforms are involved in nitrogen remobilization during leaf senescence and emphasized a role in seed production particularly in small grain crop species. Data from the sequencing of genomes for model crops and expressed sequence tag (EST) libraries from non-model species have strengthened the idea that the cytosolic GS genes are organized in three functionally and phylogenetically conserved subfamilies. Using a bioinformatic approach, the considerable publicly available information on high throughput gene expression was mined to search for genes having patterns of expression similar to GS. Interesting new hypotheses have emerged from searching for co-expressed genes across multiple unfiltered experimental data sets in rice. This approach should inform new experimental designs and studies to explore the regulation of the GS gene family further. It is expected that understanding the regulation of GS under varied climatic conditions will emerge as an important new area considering the results from recent studies that have shown nitrogen assimilation to be critical to plant acclimation to high CO(2) concentrations.

  8. Asymmetrical effects of mesophyll conductance on fundamental photosynthetic parameters and their relationships estimated from leaf gas exchange measurements

    Most previous analyses of leaf gas exchange measurements assumed an infinite value of mesophyll conductance (gm) and thus equaled CO2 partial pressures in the substomatal cavity and chloroplast. Yet an increasing number of studies have recognized that gm is finite and there is a drawdown of CO2 part...

  9. Stomatal conductance, mesophyll conductance, and trans piration efficiency in relation to leaf anatomy in rice and wheat genotypes under drought

    Ouyang, Wenjing; Struik, Paul C.; Yin, Xinyou; Yang, Jianchang

    2017-01-01

    Increasing leaf transpiration efficiency (TE) may provide leads for growing rice like dryland cereals such as wheat (Triticum aestivum). To explore avenues for improving TE in rice, variations in stomatal conductance (g s) and mesophyll conductance (g m) and their anatomical determinants were

  10. Mesophyll conductance to CO2 transport estimated by two independent methods: effect of variable CO2 concentration and abscisic acid

    Vrábl, D.; Vašková, M.; Hronková, Marie; Flexas, J.; Šantrůček, Jiří

    2009-01-01

    Roč. 60, č. 8 (2009), s. 2315-2323 ISSN 0022-0957 R&D Projects: GA AV ČR(CZ) IAA601410505 Institutional research plan: CEZ:AV0Z50510513 Keywords : Carbon dioxide * mesophyll conductance * Helianthus annuus Subject RIV: ED - Physiology Impact factor: 4.271, year: 2009

  11. Hypoxia-Like Signatures Induced by BCR-ABL Potentially Alter the Glutamine Uptake for Maintaining Oxidative Phosphorylation.

    Pallavi Sontakke

    Full Text Available The Warburg effect is probably the most prominent metabolic feature of cancer cells, although little is known about the underlying mechanisms and consequences. Here, we set out to study these features in detail in a number of leukemia backgrounds. The transcriptomes of human CB CD34+ cells transduced with various oncogenes, including BCR-ABL, MLL-AF9, FLT3-ITD, NUP98-HOXA9, STAT5A and KRASG12V were analyzed in detail. Our data indicate that in particular BCR-ABL, KRASG12V and STAT5 could impose hypoxic signaling under normoxic conditions. This coincided with an upregulation of glucose importers SLC2A1/3, hexokinases and HIF1 and 2. NMR-based metabolic profiling was performed in CB CD34+ cells transduced with BCR-ABL versus controls, both cultured under normoxia and hypoxia. Lactate and pyruvate levels were increased in BCR-ABL-expressing cells even under normoxia, coinciding with enhanced glutaminolysis which occurred in an HIF1/2-dependent manner. Expression of the glutamine importer SLC1A5 was increased in BCR-ABL+ cells, coinciding with an increased susceptibility to the glutaminase inhibitor BPTES. Oxygen consumption rates also decreased upon BPTES treatment, indicating a glutamine dependency for oxidative phosphorylation. The current study suggests that BCR-ABL-positive cancer cells make use of enhanced glutamine metabolism to maintain TCA cell cycle activity in glycolytic cells.

  12. Variability in mesophyll conductance between barley genotypes, and effects on transpiration efficiency and carbon isotope discrimination.

    Barbour, Margaret M; Warren, Charles R; Farquhar, Graham D; Forrester, Guy; Brown, Hamish

    2010-07-01

    Leaf internal, or mesophyll, conductance to CO(2) (g(m)) is a significant and variable limitation of photosynthesis that also affects leaf transpiration efficiency (TE). Genotypic variation in g(m) and the effect of g(m) on TE were assessed in six barley genotypes (four Hordeum vulgare and two H. bulbosum). Significant variation in g(m) was found between genotypes, and was correlated with photosynthetic rate. The genotype with the highest g(m) also had the highest TE and the lowest carbon isotope discrimination as recorded in leaf tissue (Delta(p)). These results suggest g(m) has unexplored potential to provide TE improvement within crop breeding programmes.

  13. Durum and bread wheat differ in their ability to retain potassium in leaf mesophyll: implications for salinity stress tolerance.

    Wu, Honghong; Shabala, Lana; Zhou, Meixue; Shabala, Sergey

    2014-10-01

    Understanding the intrinsic mechanisms involved in the differential salinity tolerance between bread wheat and durum wheat is essential for breeding salt-tolerant varieties to cope with the global salinity issue threatening future food supply. In the past, higher salinity tolerance in bread wheat compared with durum wheat has been attributed to its better ability to exclude Na(+) from uptake. Here we show that another mechanism, namely more superior K(+) retention ability in the leaf mesophyll, also contributes to this difference. A strong positive correlation (R(2) > 0.41, P varieties. However, while the above correlation was strong in bread wheat, it was statistically insignificant in durum wheat. Consistent with these findings, a significantly higher relative leaf K(+) content was found in bread wheat than in durum wheat. In contrast to root tissues, the role of voltage-gated K(+) channels in K(+) retention in the wheat mesophyll was relatively small, and non-selective cation channels played a major role in controlling intracellular K(+) homeostasis. Moreover, a significant negative correlation between NaCl-induced mesophyll H(+) flux and mesophyll K(+) retention was found, and interpreted as a compensatory mechanism employed by sensitive varieties to regain K(+) leaked into the apoplast. It is concluded that bread wheat and durum wheat show different strategies of coping with salinity, and that targeting mechanisms conferring K(+) retention in the leaf mesophyll may be a promising way to improve the overall salinity tolerance in these species. © The Author 2014. Published by Oxford University Press on behalf of Japanese Society of Plant Physiologists. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  14. Glutaminase enzyme biosensor for determination of glutamine in cerebrospinal fluid, human serum and l-glutamine capsule

    Bagriyanik, D.B.; Karakus, E

    2014-01-01

    Ammonium-selective glutamine biosensor was prepared by immobilizing glutaminase on poly(vinylchloride) (PVC) ammonium membrane electrode containing palmitic acid prepared by using nonactine. The response of glutamine biosensor was linear over the concentration range of 1.0x10-11.0x10-4M and slope was Nernstian. We determined optimum working conditions of the biosensor such as buffer concentration, buffer pH, lifetime, response time, linear working range and other response characteristics. The optimum buffer concentration and pH of proposed glutamine biosensor were determined as 20mM and pH 7.5, respectively. The interference effects of some ions and amino acids that may be present in body fluids were also investigated. The Km and Vmax values of glutaminase were determined. Additionally, glutamine assay in several biological samples such as healthy human serum, cerebrospinal fluid (CSF) and commercial glutamine capsule were also successfully carried out by using the standard addition method. The results were good agreement with previously reported values. (author)

  15. Insecticidal activity of glufosinate through glutamine depletion in a caterpillar.

    Kutlesa, N J; Caveney, S

    2001-01-01

    The herbicide glufosinate-ammonium (GLA) is a competitive inhibitor of glutamine synthetase (GS), an enzyme converting glutamate to glutamine in both plants and animals. Because GS is essential for ammonia detoxification in plants, GLA treatment disrupts photorespiration by causing a build-up of ammonia and a loss of glutamine in plant tissues. This study reports that GLA applied to leaf surfaces is also toxic to 5th-instar caterpillars of the skipper butterfly Calpodes ethlius (LD50 = 400 mg kg-1). After ingesting GLA, caterpillars stopped feeding and became dehydrated through a loss of rectal function. Caterpillars showed symptoms of neurotoxicity, such as proleg tremors, body convulsions and complete paralysis before death. Incubation of several tissues isolated from normal feeding-stage caterpillars with the GS substrates glutamate and ammonium showed that GLA inhibited GS activity in vitro. Within 24 h of ingesting GLA, caterpillars had a greatly reduced glutamine content and the ammonium ion levels had more than doubled. Injection of ammonium chloride into non-GLA-treated caterpillars had no deleterious effect, suggesting that glutamine depletion, and not a rise in body ammonium, was the primary cause of GLA toxicity following GS inhibition. This was supported by the observation that the onset of the symptoms of GLA poisoning could be postponed by giving GLA-fed caterpillars several subsequent daily injections of glutamine. The effective GLA dose fed to 5th-instar caterpillars in this study was comparable to the amount that might realistically by acquired from feeding on GLA-treated crops.

  16. N-acetylcysteine prevents HIV gp 120-related damage of human cultured astrocytes: correlation with glutamine synthase dysfunction

    Costa Nicola

    2007-12-01

    Full Text Available Abstract Background HIV envelope gp 120 glycoprotein is released during active HIV infection of brain macrophages thereby generating inflammation and oxidative stress which contribute to the development of the AIDS-Dementia Complex (ADC. Gp120 has also been found capable to generate excitotoxic effect on brain tissue via enhancement of glutamatergic neurotransmission, leading to neuronal and astroglial damage, though the mechanism is still to be better understood. Here we investigated on the effect of N-acetylcysteine (NAC, on gp120-induced damage in human cultured astroglial cells and the possible contribution of gp120-related reacting oxygen species (ROS in the imbalanced activity of glutamine synthase (GS, the enzyme that metabolizes glutamate into glutamine within astroglial cells playing a neuroprotective role in brain disorders. Results Incubation of Lipari human cultured astroglial cells with gp 120 (0.1–10 nM produced a significant reduction of astroglial cell viability and apoptosis as evaluated by TUNEL reaction and flow cytometric analysis (FACS. This effect was accompanied by lipid peroxidation as detected by means of malondialdehyde assay (MDA. In addition, gp 120 reduced both glutamine concentration in astroglial cell supernatants and GS expression as detected by immunocytochemistry and western blotting analysis. Pre-treatment of cells with NAC (0.5–5 mM, dose-dependently antagonised astroglial apoptotic cell death induced by gp 120, an effect accompanied by significant attenuation of MDA accumulation. Furthermore, both effects were closely associated with a significant recovery of glutamine levels in cell supernatants and by GS expression, thus suggesting that overproduction of free radicals might contribute in gp 120-related dysfunction of GS in astroglial cells. Conclusion In conclusion, the present experiments demonstrate that gp 120 is toxic to astroglial cells, an effect accompanied by lipid peroxidation and by altered

  17. Effects of hyperthyroidism and hypothyroidism on glutamine metabolism by skeletal muscle of the rat.

    Parry-Billings, M; Dimitriadis, G D; Leighton, B; Bond, J; Bevan, S J; Opara, E; Newsholme, E A

    1990-01-01

    1. The effects of hyperthyroidism and hypothyroidism on the concentrations of glutamine and other amino acids in the muscle and plasma and on the rates of glutamine and alanine release from incubated isolated stripped soleus muscle of the rat were investigated. 2. Hyperthyroidism decreased the concentration of glutamine in soleus muscle but was without effect on that in the gastrocnemius muscle or in the plasma. Hyperthyroidism also increased markedly the rate of release of glutamine from the...

  18. Molecular Mechanisms of Glutamine Synthetase Mutations that Lead to Clinically Relevant Pathologies.

    Benedikt Frieg

    2016-02-01

    Full Text Available Glutamine synthetase (GS catalyzes ATP-dependent ligation of ammonia and glutamate to glutamine. Two mutations of human GS (R324C and R341C were connected to congenital glutamine deficiency with severe brain malformations resulting in neonatal death. Another GS mutation (R324S was identified in a neurologically compromised patient. However, the molecular mechanisms underlying the impairment of GS activity by these mutations have remained elusive. Molecular dynamics simulations, free energy calculations, and rigidity analyses suggest that all three mutations influence the first step of GS catalytic cycle. The R324S and R324C mutations deteriorate GS catalytic activity due to loss of direct interactions with ATP. As to R324S, indirect, water-mediated interactions reduce this effect, which may explain the suggested higher GS residual activity. The R341C mutation weakens ATP binding by destabilizing the interacting residue R340 in the apo state of GS. Additionally, the mutation is predicted to result in a significant destabilization of helix H8, which should negatively affect glutamate binding. This prediction was tested in HEK293 cells overexpressing GS by dot-blot analysis: Structural stability of H8 was impaired through mutation of amino acids interacting with R341, as indicated by a loss of masking of an epitope in the glutamate binding pocket for a monoclonal anti-GS antibody by L-methionine-S-sulfoximine; in contrast, cells transfected with wild type GS showed the masking. Our analyses reveal complex molecular effects underlying impaired GS catalytic activity in three clinically relevant mutants. Our findings could stimulate the development of ATP binding-enhancing molecules by which the R324S mutant can be repaired extrinsically.

  19. Metabolic Cooperation of Glucose and Glutamine Is Essential for the Lytic Cycle of Obligate Intracellular Parasite Toxoplasma gondii*

    Nitzsche, Richard; Zagoriy, Vyacheslav; Lucius, Richard; Gupta, Nishith

    2015-01-01

    Toxoplasma gondii is a widespread protozoan parasite infecting nearly all warm-blooded organisms. Asexual reproduction of the parasite within its host cells is achieved by consecutive lytic cycles, which necessitates biogenesis of significant energy and biomass. Here we show that glucose and glutamine are the two major physiologically important nutrients used for the synthesis of macromolecules (ATP, nucleic acid, proteins, and lipids) in T. gondii, and either of them is sufficient to ensure ...

  20. Metabolic Cooperation of Glucose and Glutamine Is Essential for the Lytic Cycle of Obligate Intracellular Parasite Toxoplasma gondii.

    Nitzsche, Richard; Zagoriy, Vyacheslav; Lucius, Richard; Gupta, Nishith

    2016-01-01

    Toxoplasma gondii is a widespread protozoan parasite infecting nearly all warm-blooded organisms. Asexual reproduction of the parasite within its host cells is achieved by consecutive lytic cycles, which necessitates biogenesis of significant energy and biomass. Here we show that glucose and glutamine are the two major physiologically important nutrients used for the synthesis of macromolecules (ATP, nucleic acid, proteins, and lipids) in T. gondii, and either of them is sufficient to ensure the parasite survival. The parasite can counteract genetic ablation of its glucose transporter by increasing the flux of glutamine-derived carbon through the tricarboxylic acid cycle and by concurrently activating gluconeogenesis, which guarantee a continued biogenesis of ATP and biomass for host-cell invasion and parasite replication, respectively. In accord, a pharmacological inhibition of glutaminolysis or oxidative phosphorylation arrests the lytic cycle of the glycolysis-deficient mutant, which is primarily a consequence of impaired invasion due to depletion of ATP. Unexpectedly, however, intracellular parasites continue to proliferate, albeit slower, notwithstanding a simultaneous deprivation of glucose and glutamine. A growth defect in the glycolysis-impaired mutant is caused by a compromised synthesis of lipids, which cannot be counterbalanced by glutamine but can be restored by acetate. Consistently, supplementation of parasite cultures with exogenous acetate can amend the lytic cycle of the glucose transport mutant. Such plasticity in the parasite's carbon flux enables a growth-and-survival trade-off in assorted nutrient milieus, which may underlie the promiscuous survival of T. gondii tachyzoites in diverse host cells. Our results also indicate a convergence of parasite metabolism with cancer cells. © 2016 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. Metabolic Cooperation of Glucose and Glutamine Is Essential for the Lytic Cycle of Obligate Intracellular Parasite Toxoplasma gondii*

    Nitzsche, Richard; Zagoriy, Vyacheslav; Lucius, Richard; Gupta, Nishith

    2016-01-01

    Toxoplasma gondii is a widespread protozoan parasite infecting nearly all warm-blooded organisms. Asexual reproduction of the parasite within its host cells is achieved by consecutive lytic cycles, which necessitates biogenesis of significant energy and biomass. Here we show that glucose and glutamine are the two major physiologically important nutrients used for the synthesis of macromolecules (ATP, nucleic acid, proteins, and lipids) in T. gondii, and either of them is sufficient to ensure the parasite survival. The parasite can counteract genetic ablation of its glucose transporter by increasing the flux of glutamine-derived carbon through the tricarboxylic acid cycle and by concurrently activating gluconeogenesis, which guarantee a continued biogenesis of ATP and biomass for host-cell invasion and parasite replication, respectively. In accord, a pharmacological inhibition of glutaminolysis or oxidative phosphorylation arrests the lytic cycle of the glycolysis-deficient mutant, which is primarily a consequence of impaired invasion due to depletion of ATP. Unexpectedly, however, intracellular parasites continue to proliferate, albeit slower, notwithstanding a simultaneous deprivation of glucose and glutamine. A growth defect in the glycolysis-impaired mutant is caused by a compromised synthesis of lipids, which cannot be counterbalanced by glutamine but can be restored by acetate. Consistently, supplementation of parasite cultures with exogenous acetate can amend the lytic cycle of the glucose transport mutant. Such plasticity in the parasite's carbon flux enables a growth-and-survival trade-off in assorted nutrient milieus, which may underlie the promiscuous survival of T. gondii tachyzoites in diverse host cells. Our results also indicate a convergence of parasite metabolism with cancer cells. PMID:26518878

  2. Impact of dietary glutamine on amino acid digestibility values and ...

    Nebonid F. Namroud

    2017-05-22

    May 22, 2017 ... optimum growth during and after the critical period of birds' life. However ... For the first 18 days of incubation, the temperature was set at. 37.8 ºC ... Segments were cut longitudinally and flushed with cold phosphate-buffered saline (PBS). For ...... Muscle glutamine depletion in the intensive care unit. Int. J.

  3. Intravenous glutamine enhances COX-2 activity giving cardioprotection.

    McGuinness, Jonathan

    2009-03-01

    Preconditioning, a highly evolutionary conserved endogenous protective response, provides the most powerful form of anti-infarct protection known. We investigated whether acute intravenous glutamine, through an effect on cyclooxygenase (COX)-2 and heat shock protein (HSP) 72, might induce preconditioning.

  4. Synthesis of Biobased Succinonitrile from Glutamic Acid and Glutamine

    Lammens, T.M.; Nôtre, Le J.; Franssen, M.C.R.; Scott, E.L.; Sanders, J.P.M.

    2011-01-01

    Succinonitrile is the precursor of 1,4-diaminobutane, which is used for the industrial production of polyamides. This paper describes the synthesis of biobased succinonitrile from glutamic acid and glutamine, amino acids that are abundantly present in many plant proteins. Synthesis of the

  5. Antioxidant defence of L-glutamine on mitochondrial function in ...

    Myocardial infarction is a major public health concern and the leading cause of death all over the world. A better understanding of the processes involved in myocardial infarction has stimulated the search for biomolecules, which could limit the myocardial injury. We determined the protective activity of L-glutamine on ...

  6. Glutamine-enriched enteral diet increases renal arginine production

    Houdijk, A. P.; van Leeuwen, P. A.; Teerlink, T.; FLINKERBUSCH, E. L.; Boermeester, M. A.; Sauerwein, H. P.; Wesdorp, R. I.

    1994-01-01

    Arginine (Arg) is generated in the kidney by the conversion of circulating citrulline. The most important source for circulating citrulline is the metabolism of glutamine (Gln) by the gut. In this study, we investigated the influence of an enteral diet enriched with Gln on renal Arg synthesis in the

  7. Glutamine prevents oxidative stress in a model of portal hypertension.

    Zabot, Gilmara Pandolfo; Carvalhal, Gustavo Franco; Marroni, Norma Possa; Licks, Francielli; Hartmann, Renata Minuzzo; da Silva, Vinícius Duval; Fillmann, Henrique Sarubbi

    2017-07-07

    To evaluate the protective effects of glutamine in a model of portal hypertension (PH) induced by partial portal vein ligation (PPVL). Male Wistar rats were housed in a controlled environment and were allowed access to food and water ad libitum . Twenty-four male Wistar rats were divided into four experimental groups: (1) control group (SO) - rats underwent exploratory laparotomy; (2) control + glutamine group (SO + G) - rats were subjected to laparotomy and were treated intraperitoneally with glutamine; (3) portal hypertension group (PPVL) - rats were subjected to PPVL; and (4) PPVL + glutamine group (PPVL + G) - rats were treated intraperitoneally with glutamine for seven days. Local injuries were determined by evaluating intestinal segments for oxidative stress using lipid peroxidation and the activities of glutathione peroxidase (GPx), endothelial nitric oxide synthase (eNOS) and inducible nitric oxide synthase (iNOS) after PPVL. Lipid peroxidation of the membrane was increased in the animals subjected to PH ( P 0.05). The activity of the antioxidant enzyme GTx was decreased in the gut of animals subjected to PH compared with that in the control group of animals not subjected to PH ( P 0.05). At least 10 random, non-overlapping images of each histological slide with 200 × magnification (44 pixel = 1 μm) were captured. The sum means of all areas, of each group were calculated. The mean areas of eNOS staining for both of the control groups were similar. The PPVL group showed the largest area of staining for eNOS. The PPVL + G group had the second highest amount of staining, but the mean value was much lower than that of the PPVL group ( P < 0.01). For iNOS, the control (SO) and control + G (SO + G) groups showed similar areas of staining. The PPVL group contained the largest area of iNOS staining, followed by the PPVL + G group; however, this area was significantly smaller than that of the group that underwent PH without glutamine ( P < 0.01). Treatment with

  8. Nano-nutrition of chicken embryos. The effect of silver nanoparticles and glutamine on molecular responses, and the morphology of pectoral muscle

    Sawosz, Filip; Pineda, Lane Manalili; Hotowy, Anna Malgorzata

    2012-01-01

    Background: It has been demonstrated that concentrations of certain amino acids in the egg, in late-term embryos, are not sufficient to fully support embryonic development. One of the methods to assure an adequate nutrient content in the egg is in ovo administration of nutrients, which increases...... and vascular endothelial growth factor. We have therefore tested if silver nanoparticles can affect muscle development of chicken embryos and, furthermore, if they can be used in in ovo nutrition as carriers of nutrients e.g. glutamine into muscle cells. Methods: 160 broiler eggs were randomly divided...... into the control group (Control) without injection and injected groups with hydrocolloids of nanoparticles of silver (Nano-Ag), glutamine (Glu) and the complex of nanoparticles of silver and glutamine (Nano-Ag/Glu). The embryos were evaluated on day 20 of incubation. Samples of the breast muscles were collected...

  9. Prevention of gastrointestinal injury by using glutamine in cardiac surgery

    С. М. Ефремов

    2015-10-01

    Full Text Available A pilot double-blind, placebo-controlled, randomized study The aim of this study was to evaluate the efficiency of perioperative administration of glutamine to preserve intestinal integrity in patients undergoing cardiac surgery. 24 patients scheduled for elective coronary artery bypass surgery under cardiopulmonary bypass were included in this prospective, randomized, double-blind placebo controlled pilot study. 12 patients were randomized to receive glutamine (20% solution of N(2-L-alanyl-L-glutamine 0.4 g/kg a day, while the remaining 12 patients received an equivalent placebo dose (0.9% solution of NaCl. Infusion of glutamine/placebo was started after the induction of anesthesia and was continued for 24 hours. The primary end-point was dynamics of plasma concentration of a specific marker of intestinal damage, intestinal fatty acid binding protein (I-FABP. The secondary end-points were liver fatty acid binding protein (L-FABP, alpha glutathione s-transferase (aGST, heat shock protein 70 (HSP 70. There were no between-group differences of all the studied biochemical parameters at any stage of the study. Plasma I-FABP levels (median [25-75 percentile] were markedly elevated during CPB and remained the same postoperatively: 962 (577-2 067 and 883 (444-1 625 г/ml 5 min after un-clamping of aorta, 2203 (888-3 429 and 1 560 (506-2 657 г/ml 2 hours post-bypass, 897 (555-1 424 and 794 (505-951 г/ml 6 hours post-bypass in the GLN and control groups respectively. Perioperative administration of glutamine in dose of 0.4 g/kg a day does not appear to preserve intestinal integrity in low risk cardiac surgery patients.

  10. Intravenous glutamine does not modify leucocyte count but shortens duration of mucositis after bone marrow transplant

    María Belén Andrade Hernández

    2018-04-01

    Full Text Available Rationale: Intravenous administration of Glutamine dipeptides (Gln has been proposed as treatment of oral mucositis following a bone marrow transplant (BMT. Objective: To establish the effects of intravenous Gln supplementation upon the severity of oral mucositis after BMT. Study design: Retrospective, analytical. Study serie: Records from 25 patients (Males: 56.0%; BMT cause: Leukemia: 64.0% who developed oral mucositis (Grades III – IV: 48.0% after BMT (Autologous: 44.0% at the "Juan Tanca Marengo" Hospital (Guayaquil, Ecuador between 2009 – 2017. Glutamine source: Dipeptiven©®: 13 grams of Gln suspended in 100 milliliters of a 20% solution of the alanine-glutamine dipeptide (Fresenius-Kabi©®, Germany. Materials and Methods: Gln-treated patients received 3(4.0% of the treatment leg, 5 (20.0%; 6 (12.0%; 7 (48.0%; or 10 (16.0% doses of the dipeptide until resolution of the symptoms. Impact of Gln was estimated from changes observed in the severity and duration of mucositis, white blood cell counts, and body weight regarding 25 non-Gln treated patients (Males: 68.0%; Leukemias: 32.0%; Autologous graft: 68.0%; Grade III – IV mucositis: 48.0%. Results: Intravenously-administered Gln shortened duration of oral mucositis: Gln-Treated: 12.5 ± 5.1 days vs. Non-Gln Treated: 21.3 ± 17.8 days (p < 0.05. Also, intravenous Gln marginally ameliorated loss of body weight: Gln-Treated -4.5 ± 5.5% vs. Non-Gln Treated: -7.5 ± 5.7% (p = 0.07. Conclusions: Intravenous Gln administration shortens duration of oral mucositis following BMT. Gln effect might be translated to a lesser weight loss in patients with oral mucositis.

  11. A simple and effective method to encapsulate tobacco mesophyll protoplasts to maintain cell viability

    Rong Lei

    2015-01-01

    • It is very convenient to change or collect the solution without mechanically disturbing the protoplasts. This simple and effective silica sol–gel/alginate two-step immobilization of protoplasts in Transwell has great potential for applications in genetic transformation, metabolite production, and migration assays.

  12. A study on the isolation of protoplasts from mesophyll cells of Dendrobium Queen Pink

    Aqeel, R.; Zehra, M.; Kazmi, S. K.; Khan, S.

    2016-01-01

    Protoplasts were successfully isolated from one month old In vitro grown plantlets of Dendrobium cultivar Queen pink. The enzyme solution used was composed of 1 percent Cellulase Onozuka R-10, 0.5 percent Macerozyme R-10, 0.1 percent Pectinase, 0.3 M mannitol, 10 mM CaCl/sub 2/.2H/sub 2/O and 10 mM 2 (N-morpholino)-ethanesulfonic acid (MES) at pH 5.8. Protoplast highest yield with 15.7x104 protoplasts per 1.5 gm freshly chopped leaves were obtained when digested in enzyme solution for 4 hrs on a rotary shaker with an agitation speed of 45 rpm in dark conditions. Protoplasts were filtered with 45 micro m nylon sieve and washed with 0.3 M mannitol solution supplemented with 10 mM CaCl/sub 2/.2H/sub 2/O and 10 mM MES, and purified with 0.3 M sucrose solution gradient. Purification of protoplasts on a sucrose mannitol gradient yielded clean protoplasts that were free from debris. (author)

  13. Energy metabolism and glutamate-glutamine cycle in the brain: a stoichiometric modeling perspective.

    Massucci, Francesco A; DiNuzzo, Mauro; Giove, Federico; Maraviglia, Bruno; Castillo, Isaac Perez; Marinari, Enzo; De Martino, Andrea

    2013-10-10

    The energetics of cerebral activity critically relies on the functional and metabolic interactions between neurons and astrocytes. Important open questions include the relation between neuronal versus astrocytic energy demand, glucose uptake and intercellular lactate transfer, as well as their dependence on the level of activity. We have developed a large-scale, constraint-based network model of the metabolic partnership between astrocytes and glutamatergic neurons that allows for a quantitative appraisal of the extent to which stoichiometry alone drives the energetics of the system. We find that the velocity of the glutamate-glutamine cycle (Vcyc) explains part of the uncoupling between glucose and oxygen utilization at increasing Vcyc levels. Thus, we are able to characterize different activation states in terms of the tissue oxygen-glucose index (OGI). Calculations show that glucose is taken up and metabolized according to cellular energy requirements, and that partitioning of the sugar between different cell types is not significantly affected by Vcyc. Furthermore, both the direction and magnitude of the lactate shuttle between neurons and astrocytes turn out to depend on the relative cell glucose uptake while being roughly independent of Vcyc. These findings suggest that, in absence of ad hoc activity-related constraints on neuronal and astrocytic metabolism, the glutamate-glutamine cycle does not control the relative energy demand of neurons and astrocytes, and hence their glucose uptake and lactate exchange.

  14. Energy metabolism and glutamate-glutamine cycle in the brain: a stoichiometric modeling perspective

    2013-01-01

    Background The energetics of cerebral activity critically relies on the functional and metabolic interactions between neurons and astrocytes. Important open questions include the relation between neuronal versus astrocytic energy demand, glucose uptake and intercellular lactate transfer, as well as their dependence on the level of activity. Results We have developed a large-scale, constraint-based network model of the metabolic partnership between astrocytes and glutamatergic neurons that allows for a quantitative appraisal of the extent to which stoichiometry alone drives the energetics of the system. We find that the velocity of the glutamate-glutamine cycle (Vcyc) explains part of the uncoupling between glucose and oxygen utilization at increasing Vcyc levels. Thus, we are able to characterize different activation states in terms of the tissue oxygen-glucose index (OGI). Calculations show that glucose is taken up and metabolized according to cellular energy requirements, and that partitioning of the sugar between different cell types is not significantly affected by Vcyc. Furthermore, both the direction and magnitude of the lactate shuttle between neurons and astrocytes turn out to depend on the relative cell glucose uptake while being roughly independent of Vcyc. Conclusions These findings suggest that, in absence of ad hoc activity-related constraints on neuronal and astrocytic metabolism, the glutamate-glutamine cycle does not control the relative energy demand of neurons and astrocytes, and hence their glucose uptake and lactate exchange. PMID:24112710

  15. Effects of hyperthyroidism and hypothyroidism on glutamine metabolism by skeletal muscle of the rat.

    Parry-Billings, M; Dimitriadis, G D; Leighton, B; Bond, J; Bevan, S J; Opara, E; Newsholme, E A

    1990-01-01

    1. The effects of hyperthyroidism and hypothyroidism on the concentrations of glutamine and other amino acids in the muscle and plasma and on the rates of glutamine and alanine release from incubated isolated stripped soleus muscle of the rat were investigated. 2. Hyperthyroidism decreased the concentration of glutamine in soleus muscle but was without effect on that in the gastrocnemius muscle or in the plasma. Hyperthyroidism also increased markedly the rate of release of glutamine from the incubated soleus muscle. 3. Hypothyroidism decreased the concentrations of glutamine in the gastrocnemius muscle and plasma but was without effect on that in soleus muscle. Hypothyroidism also decreased markedly the rate of glutamine release from the incubated soleus muscle. 4. Thyroid status was found to have marked effects on the rate of glutamine release by skeletal muscle per se, and may be important in the control of this process in both physiological and pathological conditions. PMID:2268261

  16. The importance of cytosolic glutamine synthetase in nitrogen assimilation and recycling

    Bernard, S.M.; Habash, D.Z.

    2009-07-02

    Glutamine synthetase assimilates ammonium into amino acids, thus it is a key enzyme for nitrogen metabolism. The cytosolic isoenzymes of glutamine synthetase assimilate ammonium derived from primary nitrogen uptake and from various internal nitrogen recycling pathways. In this way, cytosolic glutamine synthetase is crucial for the remobilization of protein-derived nitrogen. Cytosolic glutamine synthetase is encoded by a small family of genes that are well conserved across plant species. Members of the cytosolic glutamine synthetase gene family are regulated in response to plant nitrogen status, as well as to environmental cues, such as nitrogen availability and biotic/abiotic stresses. The complex regulation of cytosolic glutamine synthetase at the transcriptional to post-translational levels is key to the establishment of a specific physiological role for each isoenzyme. The diverse physiological roles of cytosolic glutamine synthetase isoenzymes are important in relation to current agricultural and ecological issues.

  17. Regulation of the glutamine transporter SN1 by extracellular pH and intracellular sodium ions

    Broeer, A.; Broeer, S.; Setiawan, I.; Lang, F.

    2001-01-01

    Full text: SN1 has recently been identified as one of the major glutamine transporters in hepatocytes and brain astrocytes. It appears to be the molecular correlate of the system N amino acid transporter. Two different transport mechanisms have been proposed for this transporter. Either an electroneutral mechanism, in which glutamine uptake is coupled to an exchange of 1Na + and 1H + , or an electrogenic mechanism coupled to the exchange of 2Na + against 1H + . This study was performed to solve the discrepancies and to investigate the reversibility of the transporter. When expressed in Xenopus laevis oocytes glutamine uptake activity increased strongly with increasing pH. In agreement with the pH-dependence we found that uptake of glutamine was accompanied by an alkalization of the cytosol, indicating that SN1 mediates Glutamine/H + -Antiport. Uptake of glutamine into oocytes was Na + -dependent. Analysis of the Na + -dependence of glutamine transport and Flux studies using 22 Na + indicated that two or more sodium ions were cotransported together with glutamine. However, at the same time intracellular Na + was exchanged against extracellular Na + . Taken together with the results of the pH-dependence it is proposed that SN1 mediates a Na + /Na + -exchange and a Na + /H + -exchange, both being coupled to the transport of glutamine. In agreement with this mechanism we found that acidic pH caused a reversal of the transporter. To investigate the source of the glutamine-induced inward currents, we compared inward currents generated by the 1Na + /glutamine cotransporter ATA1 with those generated by SN1. Currents induced by glutamine uptake in SN1 expressing oocytes were only a fraction of the currents induced by glutamine in ATA1 expressing oocytes, indicating that they were not generated by a stoichiometric uptake of ions. It is concluded that SN1 is tightly regulated by pH and intracellular Na + -ions and is capable of mediating glutamine uptake and release

  18. Effect of the replacement of aspartic acid/glutamic acid residues with asparagine/glutamine residues in RNase He1 from Hericium erinaceus on inhibition of human leukemia cell line proliferation.

    Kobayashi, Hiroko; Motoyoshi, Naomi; Itagaki, Tadashi; Suzuki, Mamoru; Inokuchi, Norio

    2015-01-01

    RNase He1 from Hericium erinaceus, a member of the RNase T1 family, has high identity with RNase Po1 from Pleurotus ostreatus with complete conservation of the catalytic sequence. However, the optimal pH for RNase He1 activity is lower than that of RNase Po1, and the enzyme shows little inhibition of human tumor cell proliferation. Hence, to investigate the potential antitumor activity of recombinant RNase He1 and to possibly enhance its optimum pH, we generated RNase He1 mutants by replacing 12 Asn/Gln residues with Asp/Glu residues; the amino acid sequence of RNase Po1 was taken as reference. These mutants were then expressed in Escherichia coli. Using site-directed mutagenesis, we successfully modified the optimal pH for enzyme activity and generated a recombinant RNase He1 that inhibited the proliferation of cells in the human leukemia cell line. These properties are extremely important in the production of anticancer biologics that are based on RNase activity.

  19. Measurement of gross photosynthesis, respiration in the light, and mesophyll conductance using H218O labeling.

    Gauthier, Paul Pg; Battle, Mark O; Griffin, Kevin L; Bender, Michael L

    2018-03-27

    A fundamental challenge in plant physiology is independently determining the rates of gross O2 production by photosynthesis and O2 consumption by respiration, photorespiration, and other processes. Previous studies on isolated chloroplasts or leaves have separately constrained net and gross O2 production (NOP and GOP, respectively) by labeling ambient O2 with 18O while leaf water was unlabeled. Here, we describe a method to accurately measure GOP and NOP of whole detached leaves in a cuvette as a routine gas exchange measurement. The petiole is immersed in water enriched to a δ18O of ~9,000‰, and leaf water is labeled through the transpiration stream. Photosynthesis transfers 18O from H2O to O2. GOP is calculated from the increase in δ18O of O2 as air passes through the cuvette. NOP is determined from the increase in O2/N2. Both terms are measured by isotope ratio mass spectrometry. CO2 assimilation and other standard gas exchange parameters are also measured. Reproducible measurements are made on a single leaf for more than 15 hours. We used this method to measure the light response curve of NOP and GOP in Phaseolus vulgaris at 21% and 2% O2. We then used these data to examine the O2/CO2 ratio of net photosynthesis, the light response curve of mesophyll conductance, and the apparent inhibition of respiration in the light (Kok effect) at both oxygen levels. The results are discussed in the context of evaluating the technique as a tool to study and understand leaf physiological traits. {copyright, serif} 2018 American Society of Plant Biologists. All rights reserved.

  20. Disruption of the glutamate-glutamine cycle involving astrocytes in an animal model of depression for males and females

    Virginie Rappeneau

    2016-12-01

    Full Text Available Background: Women are twice as likely as men to develop major depression (MD. The brain mechanisms underlying this sex disparity are not clear. Disruption of the glutamate-glutamine cycle has been implicated in psychiatric disturbances. This study identifies sex-based impairments in the glutamate-glutamine cycle involving astrocytes using an animal model of depression. Methods: Male and female adult Long-Evans rats were exposed to chronic social defeat stress (CSDS for 21 days, using a modified resident-intruder paradigm. Territorial aggression was used for males and maternal aggression was used for females to induce depressive-like deficits for intruders. The depressive-like phenotype was assessed with intake for saccharin solution, weight gain, estrous cycle, and corticosterone (CORT. Behaviors displayed by the intruders during daily encounters with residents were characterized. Rats with daily handling were used as controls for each sex. Ten days after the last encounter, both the intruders and controls were subjected to a no-net-flux in vivo microdialysis to assess glutamate accumulation and extracellular glutamine in the nucleus accumbens (NAc. The contralateral hemispheres were used for determining changes in astrocytic markers, including glial fibrillary acidic protein (GFAP and glutamate transporter-1 (GLT-1. Results: Both male and female intruders reduced saccharin intake over the course of CSDS, compared to their pre-stress period and to their respective controls. Male intruders exhibited submissive/defensive behaviors to territorial aggression by receiving sideways threats and bites. These males showed reductions in striatal GLT-1 and spontaneous glutamine in the NAc, compared to controls. Female intruders exhibited isolated behaviors to maternal aggression, including immobility, rearing, and self-grooming. Their non-reproductive days were extended. Also, they showed reductions in prefrontal and accumbal GFAP+ cells and prefrontal GLT

  1. Disruption of the Glutamate–Glutamine Cycle Involving Astrocytes in an Animal Model of Depression for Males and Females

    Rappeneau, Virginie; Blaker, Amanda; Petro, Jeff R.; Yamamoto, Bryan K.; Shimamoto, Akiko

    2016-01-01

    Background: Women are twice as likely as men to develop major depression. The brain mechanisms underlying this sex disparity are not clear. Disruption of the glutamate–glutamine cycle has been implicated in psychiatric disturbances. This study identifies sex-based impairments in the glutamate–glutamine cycle involving astrocytes using an animal model of depression. Methods: Male and female adult Long-Evans rats were exposed to chronic social defeat stress (CSDS) for 21 days, using a modified resident-intruder paradigm. Territorial aggression was used for males and maternal aggression was used for females to induce depressive-like deficits for intruders. The depressive-like phenotype was assessed with intake for saccharin solution, weight gain, estrous cycle, and corticosterone (CORT). Behaviors displayed by the intruders during daily encounters with residents were characterized. Rats with daily handling were used as controls for each sex. Ten days after the last encounter, both the intruders and controls were subjected to a no-net-flux in vivo microdialysis to assess glutamate accumulation and extracellular glutamine in the nucleus accumbens (NAc). The contralateral hemispheres were used for determining changes in astrocytic markers, including glial fibrillary acidic protein (GFAP) and glutamate transporter-1 (GLT-1). Results: Both male and female intruders reduced saccharin intake over the course of CSDS, compared to their pre-stress period and to their respective controls. Male intruders exhibited submissive/defensive behaviors to territorial aggression by receiving sideways threats and bites. These males showed reductions in striatal GLT-1 and spontaneous glutamine in the NAc, compared to controls. Female intruders exhibited isolated behaviors to maternal aggression, including immobility, rearing, and selfgrooming. Their non-reproductive days were extended. Also, they showed reductions in prefrontal and accumbal GFAP+ cells and prefrontal GLT-1, compared to

  2. Addition of Alanyl-Glutamine to Dialysis Fluid Restores Peritoneal Cellular Stress Responses - A First-In-Man Trial.

    Klaus Kratochwill

    Full Text Available Peritonitis and ultrafiltration failure remain serious complications of chronic peritoneal dialysis (PD. Dysfunctional cellular stress responses aggravate peritoneal injury associated with PD fluid exposure, potentially due to peritoneal glutamine depletion. In this randomized cross-over phase I/II trial we investigated cytoprotective effects of alanyl-glutamine (AlaGln addition to glucose-based PDF.In a prospective randomized cross-over design, 20 stable PD outpatients underwent paired peritoneal equilibration tests 4 weeks apart, using conventional acidic, single chamber 3.86% glucose PD fluid, with and without 8 mM supplemental AlaGln. Heat-shock protein 72 expression was assessed in peritoneal effluent cells as surrogate parameter of cellular stress responses, complemented by metabolomics and functional immunocompetence assays.AlaGln restored peritoneal glutamine levels and increased the primary outcome heat-shock protein expression (effect 1.51-fold, CI 1.07-2.14; p = 0.022, without changes in peritoneal ultrafiltration, small solute transport, or biomarkers reflecting cell mass and inflammation. Further effects were glutamine-like metabolomic changes and increased ex-vivo LPS-stimulated cytokine release from healthy donor peripheral blood monocytes. In patients with a history of peritonitis (5 of 20, AlaGln supplementation decreased dialysate interleukin-8 levels. Supplemented PD fluid also attenuated inflammation and enhanced stimulated cytokine release in a mouse model of PD-associated peritonitis.We conclude that AlaGln-supplemented, glucose-based PD fluid can restore peritoneal cellular stress responses with attenuation of sterile inflammation, and may improve peritoneal host-defense in the setting of PD.

  3. Modulation of the nuclear factor-kappa B (NF-κB) signalling pathway by glutamine in peritoneal macrophages of a murine model of protein malnutrition.

    da Silva Lima, Fabiana; Rogero, Marcelo Macedo; Ramos, Mayara Caldas; Borelli, Primavera; Fock, Ricardo Ambrósio

    2013-06-01

    Protein malnutrition affects resistance to infection by impairing the inflammatory response, modifying the function of effector cells, such as macrophages. Recent studies have revealed that glutamine-a non-essential amino acid, which could become conditionally essential in some situations like trauma, infection, post-surgery and sepsis-is able to modulate the synthesis of cytokines. The aim of this study was to evaluate the effect of glutamine on the expression of proteins involved in the nuclear factor-kappa B (NF-κB) signalling pathway of peritoneal macrophages from malnourished mice. Two-month-old male Balb/c mice were submitted to protein-energy malnutrition (n = 10) with a low-protein diet containing 2 % protein, whereas control mice (n = 10) were fed a 12 % protein-containing diet. The haemogram and analysis of plasma glutamine and corticosterone were evaluated. Peritoneal macrophages were pre-treated in vitro with glutamine (0, 0.6, 2 and 10 mmol/L) for 24 h and then stimulated with 1.25 μg LPS for 30 min, and the synthesis of TNF-α and IL-1α and the expression of proteins related to the NF-κB pathway were evaluated. Malnourished animals had anaemia, leucopoenia, lower plasma glutamine and increased corticosterone levels. TNF-α production of macrophages stimulated with LPS was significantly lower in cells from malnourished animals when cultivated in supraphysiological (2 and 10 mmol/L) concentrations of glutamine. Further, glutamine has a dose-dependent effect on the activation of macrophages, in both groups, when stimulated with LPS, inducing a decrease in TNF-α and IL-1α production and negatively modulating the NF-κB signalling pathway. These data lead us to infer that the protein malnutrition state interferes with the activation of macrophages and that higher glutamine concentrations, in vitro, have the capacity to act negatively in the NF-κB signalling pathway.

  4. Time-resolved transcriptome analysis of Bacillus subtilis responding to valine, glutamate, and glutamine.

    Bang-Ce Ye

    Full Text Available Microorganisms can restructure their transcriptional output to adapt to environmental conditions by sensing endogenous metabolite pools. In this paper, an Agilent customized microarray representing 4,106 genes was used to study temporal transcript profiles of Bacillus subtilis in response to valine, glutamate and glutamine pulses over 24 h. A total of 673, 835, and 1135 amino-acid-regulated genes were identified having significantly changed expression at one or more time points in response to valine, glutamate, and glutamine, respectively, including genes involved in cell wall, cellular import, metabolism of amino-acids and nucleotides, transcriptional regulation, flagellar motility, chemotaxis, phage proteins, sporulation, and many genes of unknown function. Different amino acid treatments were compared in terms of both the global temporal profiles and the 5-minute quick regulations, and between-experiment differential genes were identified. The highlighted genes were analyzed based on diverse sources of gene functions using a variety of computational tools, including T-profiler analysis, and hierarchical clustering. The results revealed the common and distinct modes of action of these three amino acids, and should help to elucidate the specific signaling mechanism of each amino acid as an effector.

  5. Activation of the TOR Signalling Pathway by Glutamine Regulates Insect Fecundity.

    Zhai, Yifan; Sun, Zhongxiang; Zhang, Jianqing; Kang, Kui; Chen, Jie; Zhang, Wenqing

    2015-05-29

    The target of rapamycin (TOR) positively controls cell growth in response to nutrients such as amino acids. However, research on the specific nutrients sensed by TOR is limited. Glutamine (Gln), a particularly important amino acid involved in metabolism in organisms, is synthesised and catalysed exclusively by glutamine synthetase (GS), and our previous studies have shown that Gln may regulate fecundity in vivo levels of the brown planthopper (BPH) Nilaparvata lugens. Until now, it has remained unclear whether Gln activates or inhibits the TOR signalling pathway. Here, we performed the combined analyses of iTRAQ (isobaric tags for relative and absolute quantification) and DGE (tag-based digital gene expression) data in N. lugens at the protein and transcript levels after GS RNAi, and we found that 52 pathways overlap, including the TOR pathway. We further experimentally demonstrate that Gln activates the TOR pathway by promoting the serine/threonine protein kinase AKT and inhibiting the 5'AMP-activated protein kinase AMPK phosphorylation activity in the pest. Furthermore, TOR regulates the fecundity of N. lugens probably by mediating vitellogenin (Vg) expression. This work is the first report that Gln activates the TOR pathway in vivo.

  6. Glutamine synthetase gene evolution: A good molecular clock

    Pesole, G.; Lanvave, C.; Saccone, C.; Bozzetti, M.P.; Preparata, G.

    1991-01-01

    Glutamine synthetase gene evolution in various animals, plants, and bacteria was evaluated by a general stationary Markov model. The evolutionary process proved to be unexpectedly regular even for a time span as long as that between the divergence of prokaryotes from eukaryotes. This enabled us to draw phylogenetic trees for species whose phylogeny cannot be easily reconstructed from the fossil record. The calculation of the times of divergence of the various organelle-specific enzymes led us to hypothesize that the pea and bean chloroplast genes for these enzymes originated from the duplication of nuclear genes as a result of the different metabolic needs of the various species. The data indicate that the duplication of plastid glutamine synthetase genes occurred long after the endosymbiotic events that produced the organelles themselves

  7. Sensitive, specific radioisotope assay for L-glutamine-D-fructose-6- phosphat aminotransferase

    Callahan, M.; Tourian, A.; Hung, W.Y.

    1981-01-01

    A sensitive and specific radioassay for L-glutamine-D-fructose-6-phosphate aminotransferase (EC 5.3.1.19) activity is presented. Picomoles of product are measurable, and the assay can be applied to systems having limited quantities of available protein, particularly in extracts of either cell or organ cultures. The assay is at least 10,000 times more sensitive under K 1 concentrations of fructose 6-phosphate than the modified Elson-Morgan colorimetric assay and 20 times more sensitive under saturating conditions of fructose 6-phosphate. As little as 0.5 μg of cell-extract protein will yield measurable product. In contrast, 280 μg of crude-extract protein from colon is required with the modified Elson-Morgan colorimetric assay

  8. l-glutamine and l-alanine supplementation increase glutamine-glutathione axis and muscle HSP-27 in rats trained using a progressive high-intensity resistance exercise.

    Leite, Jaqueline Santos Moreira; Raizel, Raquel; Hypólito, Thaís Menezes; Rosa, Thiago Dos Santos; Cruzat, Vinicius Fernandes; Tirapegui, Julio

    2016-08-01

    In this study we investigated the chronic effects of oral l-glutamine and l-alanine supplementation, either in their free or dipeptide form, on glutamine-glutathione (GLN-GSH) axis and cytoprotection mediated by HSP-27 in rats submitted to resistance exercise (RE). Forty Wistar rats were distributed into 5 groups: sedentary; trained (CTRL); and trained supplemented with l-alanyl-l-glutamine, l-glutamine and l-alanine in their free form (GLN+ALA), or free l-alanine (ALA). All trained animals were submitted to a 6-week ladder-climbing protocol. Supplementations were offered in a 4% drinking water solution for 21 days prior to euthanasia. Plasma glutamine, creatine kinase (CK), myoglobin (MYO), and erythrocyte concentration of reduced GSH and glutathione disulfide (GSSG) were measured. In tibialis anterior skeletal muscle, GLN-GSH axis, thiobarbituric acid reactive substances (TBARS), and the expression of heat shock factor 1 (HSF-1), 27-kDa heat shock protein (HSP-27), and glutamine synthetase were determined. In CRTL animals, high-intensity RE reduced muscle glutamine levels and increased GSSG/GSH rate and TBARS, as well as augmented plasma CK and MYO levels. Conversely, l-glutamine-supplemented animals showed an increase in plasma and muscle levels of glutamine, with a reduction in GSSG/GSH rate, TBARS, and CK. Free l-alanine administration increased plasma glutamine concentration and lowered muscle TBARS. HSF-1 and HSP-27 were high in all supplemented groups when compared with CTRL (p alanine, in both a free or dipeptide form, improve the GLN-GSH axis and promote cytoprotective effects in rats submitted to high-intensity RE training.

  9. [Imbalance of system of glutamin - glutamic acid in the placenta and amniotic fluid at placental insufficiency].

    Pogorelova, T N; Gunko, V O; Linde, V A

    2014-01-01

    Metabolism of glutamine and glutamic acid has been investigated in the placenta and amniotic fluid under conditions of placental insufficiency. The development of placental insufficiency is characterized by the increased content of glutamic acid and a decrease of glutamine in both placenta and amniotic fluid. These changes changes were accompanied by changes in the activity of enzymes involved in the metabolism of these amino acids. There was a decrease in glutamate dehydrogenase activity and an increase in glutaminase activity with the simultaneous decrease of glutamine synthetase activity. The compensatory decrease in the activity of glutamine keto acid aminotransferase did not prevent a decrease in the glutamine level. The impairments in the system glutamic acid-glutamine were more pronounced during the development of premature labor.

  10. Mesophyll conductance in Zea mays responds transiently to CO2 availability: implications for transpiration efficiency in C4 crops.

    Kolbe, Allison R; Cousins, Asaph B

    2018-03-01

    Mesophyll conductance (g m ) describes the movement of CO 2 from the intercellular air spaces below the stomata to the site of initial carboxylation in the mesophyll. In contrast with C 3 -g m , little is currently known about the intraspecific variation in C 4 -g m or its responsiveness to environmental stimuli. To address these questions, g m was measured on five maize (Zea mays) lines in response to CO 2 , employing three different estimates of g m . Each of the methods indicated a significant response of g m to CO 2 . Estimates of g m were similar between methods at ambient and higher CO 2 , but diverged significantly at low partial pressures of CO 2 . These differences are probably driven by incomplete chemical and isotopic equilibrium between CO 2 and bicarbonate under these conditions. Carbonic anhydrase and phosphoenolpyruvate carboxylase in vitro activity varied significantly despite similar values of g m and leaf anatomical traits. These results provide strong support for a CO 2 response of g m in Z. mays, and indicate that g m in maize is probably driven by anatomical constraints rather than by biochemical limitations. The CO 2 response of g m indicates a potential role for facilitated diffusion in C 4 -g m . These results also suggest that water-use efficiency could be enhanced in C 4 species by targeting g m . © 2017 The Authors. New Phytologist © 2017 New Phytologist Trust.

  11. Photosynthetic response of an alpine plant, Rhododendron delavayi Franch, to water stress and recovery: the role of mesophyll conductance

    Yanfei eCai

    2015-12-01

    Full Text Available Rhododendron delavayi Franch is an evergreen shrub or small tree with large scarlet flowers that makes it highly attractive as an ornamental species. The species is native to southwest China and southeast Asia, especially the Himalayan region, showing good adaptability and tolerance to drought. To understand the water stress coping mechanisms of R. delavayi, we analysed the plant’s photosynthetic performance during water stress and recovery. In particular, we looked at the regulation of stomatal (gs and mesophyll conductance (gm, and maximum rate of carboxylation (Vcmax. After four days of water stress treatment, the net CO2 assimilation rate (AN declined slightly while gs and gm were not affected and stomatal limitation (SL was therefore negligible. At this stage mesophyll conductance limitation (MCL and biochemical limitation (BL constituted the main limitation factors. After eight days of water stress treatment, AN, gs and gm had decreased notably. At this stage SL increased markedly and MCL even more so, while BL remained relatively constant. After re-watering, the recovery of AN, gs and gm was rapid, although remaining below the levels of the control plants, while Vcmax fully regained control levels after three days of re-watering. MCL remained the main limitation factor irrespective of the degree of photosynthetic recovery. In conclusion, in our experiment MCL was the main photosynthetic limitation factor of R. delavayi under water stress and during the recovery phase, with the regulation of gm probably being the result of interactions between the environment and leaf anatomical features.

  12. Changes in photosynthesis, mesophyll conductance to CO2, and isoprenoid emissions in Populus nigra plants exposed to excess nickel

    Velikova, Violeta; Tsonev, Tsonko; Loreto, Francesco; Centritto, Mauro

    2011-01-01

    Poplar (Populus nigra) plants were grown hydroponically with 30 and 200 μM Ni (Ni 30 and Ni 200 ). Photosynthesis limitations and isoprenoid emissions were investigated in two leaf types (mature and developing). Ni stress significantly decreased photosynthesis, and this effect depended on the leaf Ni content, which was lower in mature than in developing leaves. The main limitations to photosynthesis were attributed to mesophyll conductance and metabolism impairment. In Ni-stressed developing leaves, isoprene emission was significantly stimulated. We attribute such stimulation to the lower chloroplastic [CO 2 ] than in control leaves. However chloroplastic [CO 2 ] did not control isoprene emission in mature leaves. Ni stress induced the emission of cis-β-ocimene in mature leaves, and of linalool in both leaf types. Induced biosynthesis and emission of isoprenoids reveal the onset of antioxidant processes that may also contribute to reduce Ni stress, especially in mature poplar leaves. - Graphical abstract: Visible damage caused by Ni treatment. 1 - Ni 0 (control plants); 2 - Ni 200 ; M = mature and D = developing Populus nigra leaves. Display Omitted Highlights: → We study the effect of Ni pollution on photosynthesis and isoprenoid emissions. → Ni stress significantly decreases photosynthesis. The main limitations are attributed to mesophyll conductance and metabolism impairment. → Constitutive isoprene emission was significantly stimulated in Ni-stressed leaves. Exposure to enhanced Ni concentration induces cis-beta-ocimene and linalool emissions. - The study reveals consequences of Ni stress on plant physiology, namely increasing diffusional limitation to photosynthesis and isoprenoid emissions.

  13. Comparative proteomics of chloroplasts envelopes from bundle sheath and mesophyll chloroplasts reveals novel membrane proteins with a possible role in C4-related metabolite fluxes and development.

    Kalpana eManandhar-Shrestha

    2013-03-01

    Full Text Available As the world population grows, our need for food increases drastically. Limited amounts of arable land lead to a competition between food and fuel crops, while changes in the global climate may impact future crop yields. Thus, a second green revolution will need a better understanding of the processes essential for plant growth and development. One approach toward the solution of this problem is to better understand regulatory and transport processes in C4 plants. C4 plants display an up to 10-fold higher apparent CO2 assimilation and higher yields while maintaining high water use efficiency. This requires differential regulation of mesophyll (M and bundle sheath (BS chloroplast development as well as higher metabolic fluxes of photosynthetic intermediates between cells and across chloroplast envelopes. While previous analyses of overall chloroplast membranes have yielded significant insight, our comparative proteomics approach using enriched BS and M chloroplast envelopes of Zea mays allowed us to identify 37 proteins of unknown function that have not been seen in these earlier studies. We identified 280 proteins, 84% of which are known/predicted to be present in chloroplasts (cp. 74% have a known or predicted membrane association. 21 membrane proteins were 2-15 times more abundant in BS cells, while 36 proteins were more abundant in M cp envelopes. These proteins could represent additional candidates of proteins essential for development or metabolite transport processes in C4 plants. RT-PCR confirmed differential expression of thirteen candidate genes. Cp association was confirmed using GFP labeling. Genes for a PIC-like protein and an ER-AP-like protein show an early transient increase in gene expression during the transition to light. In addition, PIC gene expression is increased in the immature part of the leaf and was lower in the fully developed parts of the leaf, suggesting a need for/incorporation of the protein during chloroplast

  14. Effects of alanyl-glutamine supplementation on the small intestinal mucosa barrier in weaned piglets

    Shen Xing

    2017-02-01

    Full Text Available Objective The study was to investigate the effects of alanyl-glutamine (Ala-Gln and glutamine (Gln supplementation on the intestinal mucosa barrier in piglets. Methods A total of 180 barrows with initial weight 10.01±0.03 kg were randomly allocated to three treatments, and each treatment consisted of three pens and twenty pigs per pen. The piglets of three groups were fed with control diet [0.62% alanine (Ala], Ala-Gln diet (0.5% Ala-Gln, Gln diet (0.34% Gln and 0.21% Ala, respectively. Results The results showed that in comparison with control diet, dietary Ala-Gln supplementation increased the height of villi in duodenum and jejunum (p<0.05, Gln supplementation increased the villi height of jejunum (p<0.05, Ala-Gln supplementation up-regulated the mRNA expressions of epidermal growth factor receptor and insulin-like growth factor 1 receptor in jejunal mucosa (p<0.05, raised the mRNA expressions of Claudin-1, Occludin, zonula occludens protein-1 (ZO-1 and the protein levels of Occludin, ZO-1 in jejunal mucosa (p<0.05, Ala-Gln supplementation enlarged the number of goblet cells in duodenal and ileal epithelium (p<0.05, Gln increased the number of goblet cells in duodenal epithelium (p<0.05 and Ala-Gln supplementation improved the concentrations of secretory immunoglobulin A and immunoglobulin G in the jejunal mucosa (p<0.05. Conclusion These results demonstrated that dietary Ala-Gln supplementation could maintain the integrity of small intestine and promote the functions of intestinal mucosa barriers in piglets.

  15. Physiological hypercortisolemia increases proteolysis, glutamine, and alanine production

    Darmaun, D.; Matthews, D.E.; Bier, D.M.

    1988-01-01

    Physiological elevations of plasma cortisol levels, as are encountered in stress and severe trauma, were produced in six normal subjects by infusing them with hydrocortisone for 64 h. Amino acid kinetics were measured in the postabsorptive state using three 4-h infusions of L-[1- 13 C]leucine, L-phenyl[ 2 H 5 ]phenylalanine, L-[2- 15 N]glutamine, and L-[1- 13 C]alanine tracers (1) before, (2) at 12 h, and (3) at 60 h of cortisol infusion. Before and throughout the study, the subjects ate a normal diet of adequate protein and energy intake. The cortisol infusion raised plasma cortisol levels significantly from 10 ± 1 to 32 ± 4 μg/dl, leucine flux from 83 ± 3 to 97 ± 3 μmol·kg -1 ·h -1 , and phenylalanine flux from 34 ± 1 to 39 ± 1 (SE) μmol·kg -1 ·h -1 after 12 h of cortisol infusion. These increases were maintained until the cortisol infusion was terminated. These nearly identical 15% increases in two different essential amino acid appearance rates are reflective of increased whole body protein breakdown. Glutamine flux rose by 12 h of cortisol infusion and remained elevated at the same level at 64 h. The increase in flux was primarily due to a 55% increase in glutamine de novo synthesis. Alanine flux increased with acute hypercortisolemia and increased further at 60 h of cortisol infusion, a result primarily of increased alanine de novo synthesis. Insulin, alanine, and lactate plasma levels responded similarly with significant rises between the acute and chronic periods of cortisol infusion. Thus hypercortisolemia increases both protein breakdown and the turnover of important nonessential amino acids for periods of up to 64 h

  16. Distinctive properties and expression profiles of glutamine synthetase from a plant symbiotic fungus.

    Montanini, Barbara; Betti, Marco; Márquez, Antonio J; Balestrini, Raffaella; Bonfante, Paola; Ottonello, Simone

    2003-01-01

    The nucleotide sequences reported in this paper have been submitted to the GenBank(R)/EBI Nucleotide Sequence Databases with accession numbers AF462037 (glutamine synthetase) and AF462032 (glutamate synthase). Nitrogen retrieval and assimilation by symbiotic ectomycorrhizal fungi is thought to play a central role in the mutualistic interaction between these organisms and their plant hosts. Here we report on the molecular characterization of the key N-assimilation enzyme glutamine synthetase from the mycorrhizal ascomycete Tuber borchii (TbGS). TbGS displayed a strong positive co-operativity ( n =1.7+/-0.29) and an unusually high S(0.5) value (54+/-16 mM; S(0.5) is the substrate concentration value at which v =(1/2) V (max)) for glutamate, and a correspondingly low sensitivity towards inhibition by the glutamate analogue herbicide phosphinothricin. The TbGS mRNA, which is encoded by a single-copy gene in the Tuber genome, was up-regulated in N-starved mycelia and returned to basal levels upon resupplementation of various forms of N, the most effective of which was nitrate. Both responses were accompanied by parallel variations of TbGS protein amount and glutamine synthetase activity, thus indicating that TbGS levels are primarily controlled at the pre-translational level. As revealed by a comparative analysis of the TbGS mRNA and of the mRNAs for the metabolically related enzymes glutamate dehydrogenase and glutamate synthase, TbGS is not only the sole messenger that positively responds to N starvation, but also the most abundant under N-limiting conditions. A similar, but even more discriminating expression pattern, with practically undetectable glutamate dehydrogenase mRNA levels, was observed in fruitbodies. The TbGS mRNA was also found to be expressed in symbiosis-engaged hyphae, with distinctively higher hybridization signals in hyphae that were penetrating among and within root cells. PMID:12683951

  17. Functional and Transcriptomic Characterization of Peritoneal Immune-Modulation by Addition of Alanyl-Glutamine to Dialysis Fluid.

    Herzog, Rebecca; Kuster, Lilian; Becker, Julia; Gluexam, Tobias; Pils, Dietmar; Spittler, Andreas; Bhasin, Manoj K; Alper, Seth L; Vychytil, Andreas; Aufricht, Christoph; Kratochwill, Klaus

    2017-07-24

    Peritonitis remains a major cause of morbidity and mortality during chronic peritoneal dialysis (PD). Glucose-based PD fluids reduce immunological defenses in the peritoneal cavity. Low concentrations of peritoneal extracellular glutamine during PD may contribute to this immune deficit. For these reasons we have developed a clinical assay to measure the function of the immune-competent cells in PD effluent from PD patients. We then applied this assay to test the impact on peritoneal immune-competence of PD fluid supplementation with alanyl-glutamine (AlaGln) in 6 patients in an open-label, randomized, crossover pilot trial (EudraCT 2012-004004-36), and related the functional results to transcriptome changes in PD effluent cells. Ex-vivo stimulation of PD effluent peritoneal cells increased release of interleukin (IL) 6 and tumor necrosis factor (TNF) α. Both IL-6 and TNF-α were lower at 1 h than at 4 h of the peritoneal equilibration test but the reductions in cytokine release were attenuated in AlaGln-supplemented samples. AlaGln-supplemented samples exhibited priming of IL-6-related pathways and downregulation of TNF-α upstream elements. Results from measurement of cytokine release and transcriptome analysis in this pilot clinical study support the conclusion that suppression of PD effluent cell immune function in human subjects by standard PD fluid is attenuated by AlaGln supplementation.

  18. Differential inhibition of adenylylated and deadenylylated forms of M. tuberculosis glutamine synthetase as a drug discovery platform

    Theron, Anjo

    2017-10-01

    Full Text Available mechanism, so the adenylylated form of bacterial glutamine synthetase is of particular interest. Previously published reports show that, when M. tuberculosis glutamine synthetase is expressed in Escherichia coli, the E. coli adenylyl transferase does...

  19. Plasma glutamine is a minor precursor for the synthesis of citrulline: A multispecies study

    Glutamine is considered the main precursor for citrulline synthesis in many species, including humans. The transfer of 15N from 2[15N]-glutamine to citrulline has been used as evidence for this precursor-product relationship. However, work in mice has shown that nitrogen and carbon tracers follow di...

  20. Proximal tubule-specific glutamine synthetase deletion alters basal and acidosis-stimulated ammonia metabolism

    Lee, Hyun-Wook; Osis, Gunars; Handlogten, Mary E.; Lamers, Wouter H.; Chaudhry, Farrukh A.; Verlander, Jill W.; Weiner, I. David

    2016-01-01

    Glutamine synthetase (GS) catalyzes the recycling of NH4 (+) with glutamate to form glutamine. GS is highly expressed in the renal proximal tubule (PT), suggesting ammonia recycling via GS could decrease net ammoniagenesis and thereby limit ammonia available for net acid excretion. The purpose of

  1. Quantitation and localisation of (in vitro) transglutaminase-catalysed glutamine hydroxylation using mass spectrometry

    Piersma, S.R.; Pijpekamp, A. van de; Wijngaards, G.; Gruppen, H.; Boumans, H.

    2002-01-01

    A mass spectrometric approach was chosen to quantify and localise in vitro enzymatically modified glutamine (Gln) residues in a glutamine-rich protein. This protein (named dB1), a cloned domain of the high molecular weight wheat glutenin subunit Dx5, was modified by microbial transglutaminase

  2. The role of glutamine transport in metabolism in the brain cortical tissue slice

    Hare, N.; Bubb, W.A.; Rae, C.; Broeer, S.

    2001-01-01

    The widely accepted 'glutamate/glutamine cycle' holds that glutamate released as a neurotransmitter in the brain is taken up by surrounding astrocytes, converted to neuro-inactive glutamine and transported back to neurons for reconversion to glutamate. Little, however, is known about the role of glutamine transport in this process. The situation is complicated by the fact that glutamine is transported by a variety of general amino-acid transporters of low specificity. The role of these transporters in flux of glutamine through the glutamate/glutamine cycle was investigated by 13 C NMR monitoring of the flux of C from [3- 13 C]L-lactate in guinea pig cortical tissue slices in the presence of competitive inhibitors of the A-type(α-(methylamino)isobutyrate; MeAIB) and N-type (histidine) transporters. The presence of each inhibitor (10 mM) produced no significant decrease in total metabolite pool size but resulted in a significant decrease in flux of [ 13 C] into the neurotransmitters glutamate and GABA and also into glutamine and alanine. The factional enrichment of glutamate and GABA was also significantly lower. By contrast there was no effect on the amount of [ 13 C] incorporated into aspartate isotopomers which may represent a predominantly astrocyte-labelled pool. These results are consistent with involvement of glutamine transporters in the recycling of synaptic glutamate by demonstrating partial blockage of incorporation of [ 13 C] label into neuronal metabolites

  3. Effects of glutamine on performance and intestinal mucosa morphometry of broiler chickens vaccinated against coccidiosis

    Brenda Carla Luquetti

    2016-08-01

    Full Text Available ABSTRACT This study aimed to assess the effects of glutamine as feed additive on performance and intestinal mucosa morphometry of broiler chickens vaccinated against coccidiosis. A total of 400 day-old male chicks were randomly assigned to four treatments (NVNG – no vaccination, no glutamine supplementation; NVG – no vaccination, glutamine supplementation (10 g kg−1; VNG – vaccination, no glutamine supplementation; VG – vaccination, glutamine supplementation replicated four times with 25 birds per replicate. A commercial sprayed-on vaccine against coccidiosis containing Eimeria acervulina, E. maxima, E. mivati, and E. tenella was administered at the hatchery. Broiler performance was evaluated from 1-28 days, and morphometric parameters were analyzed at 14, 21, and 28 days of age. Body weight gain and feed intake were negatively affected by vaccination, but not by glutamine. Vaccination increased crypt depth in the duodenum and jejunum at 21 and 28 days. In conclusion, this study showed that glutamine was not able to increase weight gain of broiler chickens, irrespective of whether the animals were vaccinated or not against coccidiosis. Glutamine supplementation was able to improve feed conversion in vaccinated birds suggesting trophic effect on intestinal epithelium improving.

  4. Irradiation temperature effect on glutamine (spectrophotometric readout) dosimeter

    Gupta, B.L.; Narayan, G.R.; Nilekani, S.R.; Bhat, R.M.

    2001-01-01

    For the dose estimation using glutamine dosimeter, 20 mg L-glutamine powder is dissolved in 10 ml of a solution which contains 2x10 -3 mol dm -3 ferrous ammonium sulphate and 10 -4 mol dm -3 xylenol orange in aerated aqueous 0.033 mol dm -3 sulphuric acid (FX). The plot of absorbance at 549 nm against dose is non-linear, however, the plot of 1/absorbance vs. 1/dose is linear. The slope of this linear plot changes above an absorbance of 0.3 corresponding to a dose of about 15 kGy. The response of the dosimeter is independent of irradiation temperature in the temperature range of 23-30 deg. C within the uncertainty of dose measurement. Below 23 deg. C, the absorbance decreases by 1.23% per deg. sign C decrease in temperature while between 30 and 40 deg. C, the absorbance increases by 0.75% per deg. C increase in temperature. Above 40 deg. C, the absorbance increases by 0.2% per deg. C only. The absorbance is corrected for the irradiation temperature and the dose is read from a calibration graph at 25 deg. C. Either a linear or polynomial relation can be used for the dose calculation

  5. Influence of glutamine on the effect of resistance exercise on cardiac ANP in rats

    Romeu Rodrigues de Souza

    2015-03-01

    Full Text Available Various nutritional supplements (herbs, vitamins, and micronutrients improve responses and adaptations to resistance exercise. ANP is a heart hormone that contributes to fluid, electrolyte and blood pressure homeostasis through its natriuretic and vasodilative actions. In the present study, the adaptation of ANP in response to resistance exercise was investigated in rats supplemented with glutamine for five weeks. The results showed that supplementation with glutamine did not influence the number of ANP granules per atrial cardiocyte in sedentary animals. In exercised-trained rats, the number and diameter of the granules was significantly higher in comparison with the control group and in exercised animals supplemented with glutamine there was significant increase in the number and diameter of ANP granules compared with controls. Altogether, these data indicated that in resistance exercise rats, glutamine significantly enhances cardiac ANP thus implicating the beneficial effects of glutamine supplementation to the ANP system.

  6. Synthesis, characterization, and biological evaluation of poly(L-γ-glutamyl-glutamine-paclitaxel nanoconjugate

    Sang Van

    2010-10-01

    Full Text Available Sang Van1, Sanjib K Das1, Xinghe Wang1, Zhongling Feng1, Yi Jin1, Zheng Hou1, Fu Chen1, Annie Pham1, Nan Jiang1, Stephen B Howell2, Lei Yu11Nitto Denko Technical Corporation, Oceanside, CA, USA; 2Moores Cancer Center, University of California, La Jolla, San Diego, CA, USAAbstract: The purpose of this study was to develop a novel, highly water-soluble poly(L-γ-glutamyl-glutamine-paclitaxel nanoconjugate (PGG-PTX that would improve the therapeutic index of paclitaxel (PTX. PGG-PTX is a modification of poly(L-glutamic acid-paclitaxel conjugate (PGA-PTX in which an additional glutamic acid has been added to each glutamic side chain in the polymer. PGG-PTX has higher water-solubility and faster dissolution than PGA-PTX. Unlike PGA-PTX, PGG-PTX self-assembles into nanoparticles, whose size remains in the range of 12–15 nm over the concentration range from 25 to 2,000 µg/mL in saline. Its critical micellar concentration in saline was found to be ~25 µg/mL. The potency of PGG-PTX when tested in vitro against the human lung cancer H460 cell line was comparable to other known polymer-PTX conjugates. However, PGG-PTX possesses lower toxicity compared with PGA-PTX in mice. The maximum tolerated dose of PGG-PTX was found to be 350 mg PTX/kg, which is 2.2-fold higher than the maximum tolerated dose of 160 mg PTX/kg reported for the PGA-PTX. This result indicates that PGG-PTX was substantially less toxic in vivo than PGA-PTX.Keywords: nanoconjugates, poly(L-glutamic acid, poly(L-γ-glutamyl-glutamine-paclitaxel, nanoparticles, anticancer

  7. Effect of intermittent glutamine supplementation on skeletal muscle is not long-lasting in very old rats.

    Meynial-Denis, D; Beaufrère, A-M; Mignon, M; Patureau Mirand, P

    2013-01-01

    Muscle is the major site for glutamine synthesis via glutamine synthetase (GS). This enzyme is increased 1.5-2 fold in 25-27-mo rats and may be a consequence of aging-induced stress. This stimulation is similar to the induction observed following a catabolic state such as glucocorticoid treatment (6 to 24 months). Although oral glutamine supply regulates the plasma glutamine level, nothing is known if this supplementation is interrupted before the experiment. Adult (8-mo) and very old (27-mo) female rats were exposed to intermittent glutamine supplementation for 50 % of their age lifetime. Treated rats received glutamine added to their drinking water and control rats water alone but the effect of glutamine supplementation was only studied 15 days after the last supplementation. Glutamine pretreatment discontinued 15 days before the experiment increased plasma glutamine to ~ 0.6 mM, a normal value in very old rats. However, it failed to decrease the up-regulated GS activity in skeletal muscle from very old rats. Our results suggest that long-term treatment with glutamine started before advanced age but discontinued 15 days before rat sacrifice is effective in increasing plasma glutamine to recover basal adult value and in maintaining plasma glutamine in very old rats, but has no long-lasting effect on the GS activity of skeletal muscle with advanced age.

  8. Prefrontal changes in the glutamate-glutamine cycle and neuronal/glial glutamate transporters in depression with and without suicide

    Zhao, J.; Verwer, R.W.H.; van Wamelen, D.J.; Qi, X.R.; Gao, S.F.; Lucassen, P.J.; Swaab, D.F.

    2016-01-01

    There are indications for changes in glutamate metabolism in relation to depression or suicide. The glutamate-glutamine cycle and neuronal/glial glutamate transporters mediate the uptake of the glutamate and glutamine. The expression of various components of the glutamate-glutamine cycle and the

  9. The amino acid transporters of the glutamate/GABA-glutamine cycle and their impact on insulin and glucagon secretion

    Monica eJenstad

    2013-12-01

    Full Text Available Intercellular communication is pivotal in optimising and synchronising cellular responses to keep internal homeostasis and to respond adequately to external stimuli. In the central nervous system (CNS, glutamatergic and GABAergic signals are postulated to be dependent on the glutamate/GABA-glutamine (GGG cycle for vesicular loading of neurotransmitters, for inactivating the signal and for the replenishment of the neurotransmitters. Islets of Langerhans release the hormones insulin and glucagon, but share similarities with CNS cells in for example transcriptional control of development and differentiation, and chromatin methylation. Interestingly, proteins involved in the CNS in secretion of the neurotransmitters and emitting their responses as well as the regulation of these processes, are also found in islet cells. Moreover, high levels of glutamate, GABA and glutamine and their respective vesicular and plasma membrane transporters have been shown in the islet cells and there is emerging support for these amino acids and their transporters playing important roles in the maturation and secretion of insulin and glucagon. In this review, we will discuss the feasibility of recent data in the field in relation to the biophysical properties of the transporters (Slc1, Slc17, Slc32 and Slc38 and physiology of hormone secretion in islets of Langerhans.

  10. Guard cell zeaxanthin tracks photosynthetically active radiation and stomatal apertures in Vicia faba leaves

    Srivastava, A.; Zeiger, E.

    1995-01-01

    Zeaxanthin, antheraxanthin and violaxanthin concentrations in guard cells from sonicated abaxial epidermal peels of Vicia faba were measured from dawn to dusk, and compared with concentrations in mesophyll tissue of the same leaves. Measured changes in guard cell zeaxanthin and violaxanthin concentrations indicate that guard cells operate the xanthophyll cycle throughout the day. Mesophyll tissue had no detectable zeaxanthin at dawn, whereas guard cells had 30–50 mmol mol −1 chlorophyll a+b. On a chlorophyll basis, maximal zeaxanthin levels were 3–4 fold higher in guard cells than in mesophyll cells. Zeaxanthin concentrations tracked levels of photosynthetically active radiation (PAR) in both mesophyll and guard cells. In the mesophyll, most of the zeaxanthin changes occurred in mid-morning and mid-afternoon. In guard cells, zeaxanthin concentrations changed nearly linearly with PAR in the early morning and late afternoon, and closely tracked PAR levels throughout the day. Guard cell zeaxanthin concentrations were also closely correlated with stomatal apertures. The close relationship between zeaxanthin concentrations and PAR levels in guard cells indicates that zeaxanthin is well suited to function as a molecular photosensor in stomatal movements. (author)

  11. Nano-Nutrition of Chicken Embryos. The Effect of in Ovo Administration of Diamond Nanoparticles and l-Glutamine on Molecular Responses in Chicken Embryo Pectoral Muscles

    Marta Grodzik

    2013-11-01

    Full Text Available It has been demonstrated that the content of certain amino acids in eggs is not sufficient to fully support embryonic development. One possibility to supply the embryo with extra nutrients and energy is in ovo administration of nutrients. Nanoparticles of diamond are highly biocompatible non-toxic carbonic structures, and we hypothesized that bio-complexes of diamond nanoparticles with l-glutamine may affect molecular responses in breast muscle. The objective of the investigation was to evaluate the effect of diamond nanoparticle (ND and l-glutamine (Gln on expression of growth and differentiation factors of chicken embryo pectoral muscles. ND, Gln, and Gln/ND solutions (50 mg/L were injected into fertilized broiler chicken eggs at the beginning of embryogenesis. Muscle tissue was dissected at day 20 of incubation and analysed for gene expression of FGF2, VEGF-A, and MyoD1. ND and especially Gln/ND up-regulated expression of genes related to muscle cell proliferation (FGF2 and differentiation (MyoD1. Furthermore, the ratio between FGF2 and MyoD1 was highest in the Gln/ND group. At the end of embryogenesis, Gln/ND enhanced both proliferation and differentiation of pectoral muscle cells and differentiation dominated over proliferation. These preliminary results suggest that the bio-complex of glutamine and diamond nanoparticles may accelerate growth and maturation of muscle cells.

  12. Reduced Hsp70 and Glutamine in Pediatric Severe Malaria Anemia

    Kempaiah, Prakasha; Dokladny, Karol; Karim, Zachary

    2016-01-01

    by decreased HSPA1A, a heat shock protein (Hsp) 70 coding gene. Hsp70 is a ubiquitous chaperone that regulates Nuclear Factor-kappa B (NF-κB) signaling and production of pro-inflammatory cytokines known to be important in malaria pathogenesis (e.g., IL-1β, IL-6 and TNF-α). Since the role of host Hsp70...... in malaria pathogenesis is unexplored, we investigated Hsp70 and molecular pathways in children with SMA. Validation experiments revealed that leukocytic HSP70 transcripts were reduced in SMA relative to non-severe malaria, and that intraleukocytic hemozoin (PfHz) was associated with lower HSP70. HSP70...... was correlated with reticulocyte production and Hb. Since glutamine (Gln) up-regulates Hsp70, modulates NF-κB activation, and attenuates over-expression of pro-inflammatory cytokines, circulating Gln was measured in children with malaria. Reduced Gln was associated with increased risk of developing SMA...

  13. Elevated glutamine/glutamate ratio in cerebrospinal fluid of first episode and drug naive schizophrenic patients

    Lindström Leif H

    2005-01-01

    Full Text Available Abstract Background Recent magnetic resonance spectroscopy (MRS studies report that glutamine is altered in the brains of schizophrenic patients. There were also conflicting findings on glutamate in cerebrospinal fluid (CSF of schizophrenic patients, and absent for glutamine. This study aims to clarify the question of glutamine and glutamate in CSF of first episode and drug naive schizophrenic patients. Method Levels of glutamine and glutamate in CSF of 25 first episode and drug-naive male schizophrenic patients and 17 age-matched male healthy controls were measured by a high performance liquid chromatography. Results The ratio (126.1 (median, 117.7 ± 27.4 (mean ± S.D. of glutamine to glutamate in the CSF of patients was significantly (z = -3.29, p = 0.001 higher than that (81.01 (median, 89.1 ± 22.5 (mean ± S.D. of normal controls although each level of glutamine and glutamate in patients was not different from that of normal controls. Conclusion Our data suggests that a disfunction in glutamate-glutamine cycle in the brain may play a role in the pathophysiology of schizophrenia.

  14. Lack of functional benefit with glutamine versus placebo in Duchenne muscular dystrophy: a randomized crossover trial.

    Elise Mok

    Full Text Available Oral glutamine decreases whole body protein breakdown in Duchenne muscular dystrophy (DMD. We evaluated the functional benefit of 4 months oral glutamine in DMD.30 ambulant DMD boys were included in this double-blind, randomized crossover trial with 2 intervention periods: glutamine (0.5 g/kg/d and placebo, 4 months each, separated by a 1-month wash-out, at 3 outpatient clinical investigation centers in France. Functional benefit was tested by comparing glutamine versus placebo on change in walking speed at 4 months. Secondary outcome measures were: 2-minute walk test, work, power, muscle mass (urinary creatinine, markers of myofibrillar protein breakdown (urinary 3-methyl-histidine/creatinine, serum creatine phospho-kinase, body composition (fat free mass, fat mass percentage, safety and oral nutrient intake. There was no improvement in the primary end point (walking speed or in secondary measures of muscle function (2-minute walk test, work, power in the glutamine group compared with placebo. However, subjects receiving glutamine or placebo showed no deterioration in functional measures over the course of the 9-month trial. No differences in muscle mass, markers of protein breakdown or serum creatine phosho-kinase were observed, except for a blunted increase in fat free mass in the glutamine group which led to a greater increase in fat mass percentage. Glutamine was safe and well-tolerated.This trial did not identify additional benefit of 4 months oral glutamine over placebo on muscle mass or function in ambulatory DMD boys. Although apparently safe, current data cannot support routine supplementation in this population as a whole, until further research proves otherwise.(ClinicalTrials.gov NCT00296621.

  15. Prolonged continuous intravenous infusion of the dipeptide L-alanine- L-glutamine significantly increases plasma glutamine and alanine without elevating brain glutamate in patients with severe traumatic brain injury.

    Nägeli, Mirjam; Fasshauer, Mario; Sommerfeld, Jutta; Fendel, Angela; Brandi, Giovanna; Stover, John F

    2014-07-02

    Low plasma glutamine levels are associated with worse clinical outcome. Intravenous glutamine infusion dose- dependently increases plasma glutamine levels, thereby correcting hypoglutaminemia. Glutamine may be transformed to glutamate which might limit its application at a higher dose in patients with severe traumatic brain injury (TBI). To date, the optimal glutamine dose required to normalize plasma glutamine levels without increasing plasma and cerebral glutamate has not yet been defined. Changes in plasma and cerebral glutamine, alanine, and glutamate as well as indirect signs of metabolic impairment reflected by increased intracranial pressure (ICP), lactate, lactate-to-pyruvate ratio, electroencephalogram (EEG) activity were determined before, during, and after continuous intravenous infusion of 0.75 g L-alanine-L-glutamine which was given either for 24 hours (group 1, n = 6) or 5 days (group 2, n = 6) in addition to regular enteral nutrition. Lab values including nitrogen balance, urea and ammonia were determined daily. Continuous L-alanine-L-glutamine infusion significantly increased plasma and cerebral glutamine as well as alanine levels, being mostly sustained during the 5 day infusion phase (plasma glutamine: from 295 ± 62 to 500 ± 145 μmol/ l; brain glutamine: from 183 ± 188 to 549 ± 120 μmol/ l; plasma alanine: from 327 ± 91 to 622 ± 182 μmol/ l; brain alanine: from 48 ± 55 to 89 ± 129 μmol/ l; p alanine-L-glutamine infusion (0.75 g/ kg/ d up to 5 days) increased plasma and brain glutamine and alanine levels. This was not associated with elevated glutamate or signs of potential glutamate-mediated cerebral injury. The increased nitrogen load should be considered in patients with renal and hepatic dysfunction. Clinicaltrials.gov NCT02130674. Registered 5 April 2014.

  16. Clinical and protein metabolic efficacy of glutamine granules-supplemented enteral nutrition in severely burned patients.

    Peng, Xi; Yan, Hong; You, Zhongyi; Wang, Pei; Wang, Shiliang

    2005-05-01

    As an abundant amino acid in the human body, glutamine has many important metabolic roles that may protect or promote tissue integrity and enhance the immune system. A relative deficiency of glutamine in such patients could compromise recovery and result in prolonged illness and an increase in late mortality. The purpose of this clinical study is to observe the effects of enteral supplement with glutamine granules on protein metabolism in severely burned patients. Forty-eight severe burn patients (total burn surface area 30-75%, full thickness burn area 20-58%) who met the requirements of the protocol joined this double-blind randomized controlled clinical trial. Patients were randomly divided into two groups: burn control group (B group, 23 patients) and glutamine treated group (Gln group, 25 patients). There was isonitrogenous and isocaloric intake in both groups, glutamine and B group patents were supplemented with glutamine granules or placebo (glycine) at 0.5 g/kg per day for 14 days with oral feeding or tube feeding, respectively. The level of plasma glutamine, plasma protein content, urine nitrogen and urine 3-methylhistidine (3-MTH) excretion were determined, wound healing rate of the burned area and hospital stay were recorded. The results showed that there were significant reductions in plasma glutamine level and abnormal protein metabolism. After supplement with glutamine granules for 14 days, the plasma glutamine concentration was significantly higher than that in B group (607.86+/-147.25 micromol/L versus 447.63+/-132.38 micromol/L, P0.05). On the other hand, the amount of urine nitrogen and 3-MTH excreted in Gln group were significantly lower than that in B group. In addition, wound healing was faster and hospital stay days were shorter in Gln group than B group (46.59+/-12.98 days versus 55.68+/-17.36 days, P<0.05). These indicated that supplement glutamine granules with oral feeding or tube feeding could abate the degree of glutamine depletion

  17. Glutamine's protection against cellular injury is dependent on heat shock factor-1.

    Morrison, Angela L; Dinges, Martin; Singleton, Kristen D; Odoms, Kelli; Wong, Hector R; Wischmeyer, Paul E

    2006-06-01

    Glutamine (GLN) has been shown to protect cells, tissues, and whole organisms from stress and injury. Enhanced expression of heat shock protein (HSP) has been hypothesized to be responsible for this protection. To date, there are no clear mechanistic data confirming this relationship. This study tested the hypothesis that GLN-mediated activation of the HSP pathway via heat shock factor-1 (HSF-1) is responsible for cellular protection. Wild-type HSF-1 (HSF-1(+/+)) and knockout (HSF-1(-/-)) mouse fibroblasts were used in all experiments. Cells were treated with GLN concentrations ranging from 0 to 16 mM and exposed to heat stress injury in a concurrent treatment model. Cell viability was assayed with phenazine methosulfate plus tetrazolium salt, HSP-70, HSP-25, and nuclear HSF-1 expression via Western blot analysis, and HSF-1/heat shock element (HSE) binding via EMSA. GLN significantly attenuated heat-stress induced cell death in HSF-1(+/+) cells in a dose-dependent manner; however, the survival benefit of GLN was lost in HSF-1(-/-) cells. GLN led to a dose-dependent increase in HSP-70 and HSP-25 expression after heat stress. No inducible HSP expression was observed in HSF-1(-/-) cells. GLN increased unphosphorylated HSF-1 in the nucleus before heat stress. This was accompanied by a GLN-mediated increase in HSF-1/HSE binding and nuclear content of phosphorylated HSF-1 after heat stress. This is the first demonstration that GLN-mediated cellular protection after heat-stress injury is related to HSF-1 expression and cellular capacity to activate an HSP response. Furthermore, the mechanism of GLN-mediated protection against injury appears to involve an increase in nuclear HSF-1 content before stress and increased HSF-1 promoter binding and phosphorylation.

  18. Gene expression, cellular localisation and function of glutamine synthetase isozymes in wheat (Triticum aestivum L.)

    Bernard, Stéphanie M.; Møller, Anders Laurell Blom; Dionisio, Giuseppe

    2008-01-01

    ). Phylogenetic analysis showed that the wheat GS sub-families together with the GS genes from other monocotyledonous species form four distinct clades. Immunolocalisation studies in leaves, stems and rachis in plants at flowering showed GS protein to be present in parenchyma, phloem companion and perifascicular......We present the first cloning and study of glutamine synthetase (GS) genes in wheat (Triticum aestivum L.). Based on sequence analysis, phylogenetic studies and mapping data, ten GS sequences were classified into four sub-families: GS2 (a, b and c), GS1 (a, b and c), GSr (1 and 2) and GSe (1 and 2...... sheath cells. In situ localisation confirmed that GS1 transcripts were present in the perifascicular sheath cells whilst those for GSr were confined to the vascular cells. Studies of the expression and protein profiles showed that all GS sub-families were differentially expressed in the leaves, peduncle...

  19. Ion exclusion chromatography for the purification of L-glutamine; Ion haijo chromatography ni yoru L-glutamine no seiseiho no kenkyu

    Ito, H.; Nishi, A.; Naruse, M. [Ajinomoto Co. Inc., Kawasaki (Japan)

    1998-09-05

    Ion exclusion chromatography for the purification of L-glutamine is studied. L-glutamine is usually produced by fermentation and used in pharmaceuticals. By using a model solution of L-glutamine and L-glutamic acid, the optimum cation exchange resin is examined. As a result of the experiments, it is found that a cation exchange resin which has smaller crosslinkage and smaller diameter is better. Ammonium sulfate, L-glutamic acid and pyrrolidonecarboxylic acid, which are usually contained in fermentation broth as impurities, are effectively separated by this method. Moreover, the experimental data of the chromatography is expressed fairly well by the differential equations which express the mass transfer in the fixed bed. 8 refs., 5 figs., 2 tabs.

  20. Lack of cardioprotection from metabolic support with glutamine or glutamate in a porcine coronary occlusion model

    Kristensen, Jens; Mæng, Michael; Mortensen, Ulrik

    2005-01-01

    vascular resistance, while glutamate preserved cardiac output during infusion. CONCLUSION: Substrate supplementation with the anaplerotic precursors glutamine and glutamate is ineffective as adjunctive therapy for severe myocardial ischemia. Beneficial effects documented in less complex experimental...

  1. Effects of Standard and/or Glutamine Dipeptide and/or Omega-3 ...

    Supplemented Parenteral Nutrition on Neutrophil Functions, Interleukin-8 Level and Length of ... Standard TPN and glutamine and lipid emulsion with omega 3 fatty acids were given to colorectal cancer patients and the effects of these to neutrophil ...

  2. Serum Glutamine Levels as a Potential Diagnostic Biomarker in Sepsis following Surgery for Peritonitis.

    Yang, Chun-Ju; Huang, Ting-Shuo; Lee, Tung-Liang; Yang, Kang-Chung; Yuan, Shin-Sheng; Lu, Ruey-Hwa; Hsieh, Chung-Ho; Shyu, Yu-Chiau

    2017-12-31

    Few diagnostic biomarkers for sepsis after emergency peritonitis surgery are available to clinicians, and, thus, it is important to develop new biomarkers for patients undergoing this procedure. We investigated whether serum glutamine and selenium levels could be diagnostic biomarkers of sepsis in individuals recovering from emergency peritonitis surgery. From February 2012 to March 2013, patients who had peritonitis diagnosed at the emergency department and underwent emergency surgery were screened for eligibility. Serum glutamine and selenium levels were obtained at pre-operative, post-operative and recovery time points. The average level of pre-operation serum glutamine was significantly different from that on the recovery day (0.317 ± 0.168 vs. 0.532 ± 0.155 mM, P peritonitis. We recommend including glutamine as a biomarker for sepsis severity assessment in addition to the commonly used clinical indicators.

  3. Influence of Drought on Mesophyll Resistance to CO2 Diffusion and its Impact on Water-Use Efficiency in Trees

    Guo, J.; Beverly, D.; Cook, C.; Ewers, B. E.; Williams, D. G.

    2015-12-01

    The resistance to CO2 diffusion inside leaves (mesophyll resistance; rm) during photosynthesis is often comparable in magnitude to stomatal diffusion resistance, and varies among species and across environmental conditions. Consequently, photosynthesis is strongly limited by rm at low internal CO2 partial pressures, such that its variation may determine patterns of leaf water-use efficiency (WUE). Reduction in stomatal conductance with drought typically increases WUE, but also decreases photosynthesis. In theory, the decrease in photosynthesis could be countered by reduction in rm while maintaining high WUE. It is still uncertain how drought-related changes in rm affect short- and long-term WUE strategies of different tree species. We conducted field observations of instantaneous WUE and 13C discrimination in two dominant conifer species (Pinus contorta and Picea engelmannii) in SE Wyoming over the seasonal dry-down period in the summer of 2015. rm was examined by on-line 13C discrimination using isotope laser spectroscopy. Controlled environment studies on three conifer species (P. contorta, P. engelmannii, and Abies lasiocarpa) and one angiosperm (Populus tremuloides) are in progress. We hypothesize that the plasticity of rm in response to drought accounts for significant adjustments in photosynthetic capacity and WUE. Needle leaf conifers are known to have relatively high rm, and we expect them to show greater improvements in photosynthesis and WUE when rm is decreased compared to angiosperm tree species.

  4. Asymmetrical effects of mesophyll conductance on fundamental photosynthetic parameters and their relationships estimated from leaf gas exchange measurements.

    Sun, Ying; Gu, Lianhong; Dickinson, Robert E; Pallardy, Stephen G; Baker, John; Cao, Yonghui; DaMatta, Fábio Murilo; Dong, Xuejun; Ellsworth, David; Van Goethem, Davina; Jensen, Anna M; Law, Beverly E; Loos, Rodolfo; Martins, Samuel C Vitor; Norby, Richard J; Warren, Jeffrey; Weston, David; Winter, Klaus

    2014-04-01

    Worldwide measurements of nearly 130 C3 species covering all major plant functional types are analysed in conjunction with model simulations to determine the effects of mesophyll conductance (g(m)) on photosynthetic parameters and their relationships estimated from A/Ci curves. We find that an assumption of infinite g(m) results in up to 75% underestimation for maximum carboxylation rate V(cmax), 60% for maximum electron transport rate J(max), and 40% for triose phosphate utilization rate T(u) . V(cmax) is most sensitive, J(max) is less sensitive, and T(u) has the least sensitivity to the variation of g(m). Because of this asymmetrical effect of g(m), the ratios of J(max) to V(cmax), T(u) to V(cmax) and T(u) to J(max) are all overestimated. An infinite g(m) assumption also limits the freedom of variation of estimated parameters and artificially constrains parameter relationships to stronger shapes. These findings suggest the importance of quantifying g(m) for understanding in situ photosynthetic machinery functioning. We show that a nonzero resistance to CO2 movement in chloroplasts has small effects on estimated parameters. A non-linear function with gm as input is developed to convert the parameters estimated under an assumption of infinite gm to proper values. This function will facilitate gm representation in global carbon cycle models. © 2013 John Wiley & Sons Ltd.

  5. Effects of various glutamine concentrations on gene expression and steviol glycosides accumulation in Stevia rebaudiana Bertoni.

    Esmaeili, Fatemeh; Ghaheri, Matin; Kahrizi, Danial; Mansouri, Mohsen; Safavi, Seyed Mehdi; Ghorbani, Tayebeh; Muhammadi, Sarre; Rahmanian, Elham; Vaziri, Siavash

    2018-02-10

    Stevia rebaudiana Bertoni is one of the most important biologically sourced and low-calorie sweeteners that contains a lots of Steviol glycosides. Tissue culture is the best method for propagation of stevia and micro nutrients can affect both morphological traits and steviol glycosides production. In the present study, we investigated the effect of different concentrations of glutamine (10, 20, 30 and 40 g/l) on expression of UGT74G1 and UGT76G1 genes and stevioside and rebaudioside A accumulation in the leaves of stevia under in vitro conditions. The highest level of expression for UGT74G1 (1.000 Total lab unit) was seen at plants grown in MS media without glutamine and the highest gene expression level for UGT76G1 (1.321 Total lab unit) was observed at plants grown in 2% glutamine. Based on HPLC results, the highest amount of stevioside (22.74) was accumulated in plants which were under 3% glutamine treatment and the lowest production level of stevioside (16.19) was resulted under MS (0 glutamine) medium. The highest rebaudioside A (12.19) accumulation was observed under 2% glutamine treatment and the lowest accumulation of rebaudioside A (8.41) was seen at plants grown in MS medium.

  6. Evidence for an operative glutamine translocator in chloroplasts from maritime pine (Pinus pinaster Ait.) cotyledons.

    Claros, M G; Aguilar, M L; Cánovas, F M

    2010-09-01

    In higher plants, ammonium is assimilated into amino acids through the glutamine synthetase (GS)/glutamate synthase (GOGAT) cycle. This metabolic cycle is distributed in different cellular compartments in conifer seedlings: glutamine synthesis occurs in the cytosol and glutamate synthesis within the chloroplast. A method for preparing intact chloroplasts of pine cotyledons is presented with the aim of identifying a glutamine-glutamate translocator. Glutamine-glutamate exchange has been studied using the double silicone layer system, suggesting the existence of a translocator that imports glutamine into the chloroplast and exports glutamate to the cytoplasm. The translocator identified is specific for glutamine and glutamate, and the kinetic constants for both substrates indicate that it is unsaturated at intracellular concentrations. Thus, the experimental evidence obtained supports the model of the GS/GOGAT cycle in developing pine seedlings that accounts for the stoichiometric balance of metabolites. As a result, the efficient assimilation of free ammonia produced by photorespiration, nitrate reduction, storage protein mobilisation, phenylpropanoid pathway or S-adenosylmethionine synthesis is guaranteed.

  7. Pilot study with a glutamine-supplemented enteral formula in critically ill infants

    Barbosa Eliana

    1999-01-01

    Full Text Available Seriously ill infants often display protein-calorie malnutrition due to the metabolic demands of sepsis and respiratory failure. Glutamine has been classified as a conditionally essential amino acid, with special usefulness in critical patients. Immunomodulation, gut protection, and prevention of protein depletion are mentioned among its positive effects in such circumstances. With the intent of evaluating the tolerance and clinical impact of a glutamine supplement in seriously ill infants, a prospective randomized study was done with nine patients. Anthropometric and biochemical determinations were made, and length of stay in the intensive care unit (ICU, in the hospital, and under artificial ventilation, and septic morbidity and mortality were tabulated. Infants in the treatment group (n=5 were enterally administered 0.3 g/kg of glutamine, whereas controls received 0.3 g/kg of casein during a standard period of five days. Septic complications occurred in 75% of the controls (3/4 versus 20% of the glutamine-treated group (1/5, p<=0.10, and two patients in the control group died of bacterial infections (50% vs. 0%, p<=0.10. Days in the ICU, in the hospital, and with ventilation numerically favored glutamine therapy, although without statistical significance. The supplements were usually well tolerated, and no patient required discontinuation of the program. The conclusion was that glutamine supplementation was safe and tended to be associated with less infectious morbidity and mortality in this high-risk population.

  8. Assay of glutamine phosphoribosylpyrophosphate amidotransferase using [1-14C]phosphoribosylpyrophosphate

    Ross, G.R.; Idriss, S.D.; Willis, R.C.; Seegmiller, J.E.

    1983-01-01

    Glutamine phosphoribosylpyrophosphate amidotransferase (EC 2.4.2.14) catalyzes the transfer of the amide group of glutamine to 5-phospho-α-D-ribose-1-pyrophosphate. It is the first enzyme committed to the synthesis of purines by the de novo pathway. Previous assays of enzyme activity have either measured the phosphoribosylpyrophosphate-dependent disappearance of radioactive glutamine or have linked this reaction to subsequent steps in the purine pathway. A new assay for activity of the enzyme by directly measuring the synthesis of the product of the reaction, 5-β-phosphoribosyl-1-amine, using [1- 14 C]phosphoribosylpyrophosphate as substrate is described. Substrate and product are separated by thin-layer chromatography and identified by autoradiography. Glutamine or ammonia may be used as substrates; the apparent K/sub m/ values of the human lymphoblast enzyme are 0.46 mM for glutamine and 0.71 mM for ammonia. GMP is a considerably more potent inhibitor of the human lymphoblast enzyme than is AMP; 6-diazo-5-oxo-L-norleucine inhibits only glutamine-dependent activity and has no effect on ammonia-dependent activity

  9. Effect of glutamine synthetase inhibition on brain and interorgan ammonia metabolism in bile duct ligated rats.

    Fries, Andreas W; Dadsetan, Sherry; Keiding, Susanne; Bak, Lasse K; Schousboe, Arne; Waagepetersen, Helle S; Simonsen, Mette; Ott, Peter; Vilstrup, Hendrik; Sørensen, Michael

    2014-03-01

    Ammonia has a key role in the development of hepatic encephalopathy (HE). In the brain, glutamine synthetase (GS) rapidly converts blood-borne ammonia into glutamine which in high concentrations may cause mitochondrial dysfunction and osmolytic brain edema. In astrocyte-neuron cocultures and brains of healthy rats, inhibition of GS by methionine sulfoximine (MSO) reduced glutamine synthesis and increased alanine synthesis. Here, we investigate effects of MSO on brain and interorgan ammonia metabolism in sham and bile duct ligated (BDL) rats. Concentrations of glutamine, glutamate, alanine, and aspartate and incorporation of (15)NH(4)(+) into these amino acids in brain, liver, muscle, kidney, and plasma were similar in sham and BDL rats treated with saline. Methionine sulfoximine reduced glutamine concentrations in liver, kidney, and plasma but not in brain and muscle; MSO reduced incorporation of (15)NH(4)(+) into glutamine in all tissues. It did not affect alanine concentrations in any of the tissues but plasma alanine concentration increased; incorporation of (15)NH(4)(+) into alanine was increased in brain in sham and BDL rats and in kidney in sham rats. It inhibited GS in all tissues examined but only in brain was an increased incorporation of (15)N-ammonia into alanine observed. Liver and kidney were important for metabolizing blood-borne ammonia.

  10. Fibronectin-integrin signaling is required for L-glutamine's protection against gut injury.

    Stefanie Niederlechner

    Full Text Available Extracellular matrix (ECM stabilization and fibronectin (FN-Integrin signaling can mediate cellular protection. L-glutamine (GLN is known to prevent apoptosis after injury. However, it is currently unknown if ECM stabilization and FN-Integrin osmosensing pathways are related to GLN's cell protective mechanism in the intestine.IEC-6 cells were treated with GLN with or without FN siRNA, integrin inhibitor GRGDSP, control peptide GRGESP or ERK1/2 inhibitors PD98059 and UO126 under basal and stressed conditions. Cell survival measured via MTS assay. Phosphorylated and/or total levels of cleaved caspase-3, cleaved PARP, Bax, Bcl-2, heat shock proteins (HSPs, ERK1/2 and transcription factor HSF-1 assessed via Western blotting. Cell size and F-actin morphology quantified by confocal fluorescence microscopy and intracellular GLN concentration by LC-MS/MS.GLN's prevention of FN degradation after hyperthermia attenuated apoptosis. Additionally, inhibition of FN-Integrin interaction by GRGDSP and ERK1/2 kinase inhibition by PD98059 inhibited GLN's protective effect. GRGDSP attenuated GLN-mediated increases in ERK1/2 phosphorylation and HSF-1 levels. PD98059 and GRGDSP also decreased HSP levels after GLN treatment. Finally, GRGDSP attenuated GLN-mediated increases in cell area size and disrupted F-actin assembly, but had no effect on intracellular GLN concentrations.Taken together, this data suggests that prevention of FN degradation and the FN-Integrin signaling play a key role in GLN-mediated cellular protection. GLN's signaling via the FN-Integrin pathway is associated with HSP induction via ERK1/2 and HSF-1 activation leading to reduced apoptosis after gut injury.

  11. Design of composite microparticle systems based on pectin and waste material of propolis for modified l-alanyl-l-glutamine release and with immunostimulant activity.

    Villa Nova, Mônica; Ratti, Bianca A; Herculano, Leandro S; Bittencourt, Paulo R S; Novello, Cláudio R; Bazotte, Roberto Barbosa; Lautenschlager, Sueli de Oliveira Silva; Bruschi, Marcos Luciano

    2017-12-12

    Catabolic conditions like acquired immunodeficiency syndrome, cancer, and burn can cause immunosuppression. Amino acids such as alanine and glutamine are essential for the activity of the immune system. Propolis is immunostimulant and the waste of propolis extraction has been reused with technological and therapeutic purposes. Therefore, this study describes the association of propolis byproduct extract (BPE) with pectin to prepare spray-dried microparticles containing the dipeptide l-alanyl-l-glutamine as stimulant systems of neutrophils. The use of a factorial design allowed selecting the best formulation, which was characterized by morphology, size, and entrapment efficiency analyses. In addition, the systems were characterized by thermal and X-ray diffraction analysis, Fourier-transform infrared spectroscopy, in vitro drug release, and in vitro cytotoxicity and stimulation test of neutrophils. Small well-structured microparticles with good entrapment efficiency values were achieved. Thermal stability of formulation was observed, and it was proved that pectin, BPE and l-alanyl-l-glutamine were dispersed throughout the matrix. The drug was released from the microparticles during 24 h governed by swelling and diffusion. The drug-loaded formulations showed a significant stimulating effect on neutrophils. These structures could increase the activity of immune cells, and other in vitro and in vivo studies should be performed in the future.

  12. Effects of creep feeding and supplemental glutamine or glutamine plus glutamate (Aminogut) on pre- and post-weaning growth performance and intestinal health of piglets

    2013-01-01

    Background Creep feeding is used to stimulate piglet post-weaning feed consumption. L-Glutamine (GLN) is an important source of fuel for intestinal epithelial cells. The objective of this study was to determine the impact of creep feeding and adding GLN or AminoGut (AG; containing glutamine + glutamate) to pre- and post-weaning diets on pig performance and intestinal health. Litters (N = 120) were allotted to four treatments during 14–21 d of lactation: 1) No creep feed (NC, n = 45); 2) creep fed control diet (CFCD, n = 45); 3) creep fed 1% GLN (CFGLN, n = 15); 4) creep fed .88% AG (CFAG, n = 15). After weaning, the NC and CFCD groups were sub-divided into three groups (n = 15 each), receiving either a control nursery diet (NC-CD, CFCD-CD) or a diet supplemented with either GLN (NC-GLN, CFCD-GLN) or with AG (NC-AG, CFCD-AG). Litters that were creep fed with diets containing GLN or AG also were supplemented with those amino acids in the nursery diets (CFGLN-GLN, CFAG-AG). Glutamine was added at 1% in all three post-weaning diet phases and AG was added at .88% in phase 1 and 2 and at .66% in phase 3. Results Feed conversion (feed/gain) showed means among treatment means close to significance (P = 0.056) and Tukey’s test for pairwise mean comparisons showed that Pigs in the CFGLN-GLN group had the best feed conversion (feed/gain) in the first three-week period post-weaning, exceeding (P = 0.044) controls (CFCD-CD) by 34%. The NC-AG group had (P = 0.02) the greatest feed intake in the last three week of the study, exceeding controls (CFCD-CD) by 12%. CFGLN-GLN, CFCD-GLN and sow reared (SR) pigs had the greatest (P = 0.049) villi height exceeding the CFCD-AG group by 18%, 20% and 19% respectively. The CFAG-AG group had the deepest (P = 0.001) crypts among all treatments. CFGLN-GLN, CFCD-GLN and SR groups had the greatest (P = 0.001) number of cells proliferating (PCNA) exceeding those in the NC-CD group by 43%, 54

  13. Effects of creep feeding and supplemental glutamine or glutamine plus glutamate (Aminogut) on pre- and post-weaning growth performance and intestinal health of piglets.

    Cabrera, Rafael A; Usry, James L; Arrellano, Consuelo; Nogueira, Eduardo T; Kutschenko, Marianne; Moeser, Adam J; Odle, Jack

    2013-08-03

    Creep feeding is used to stimulate piglet post-weaning feed consumption. L-Glutamine (GLN) is an important source of fuel for intestinal epithelial cells. The objective of this study was to determine the impact of creep feeding and adding GLN or AminoGut (AG; containing glutamine + glutamate) to pre- and post-weaning diets on pig performance and intestinal health. Litters (N = 120) were allotted to four treatments during 14-21 d of lactation: 1) No creep feed (NC, n = 45); 2) creep fed control diet (CFCD, n = 45); 3) creep fed 1% GLN (CFGLN, n = 15); 4) creep fed .88% AG (CFAG, n = 15). After weaning, the NC and CFCD groups were sub-divided into three groups (n = 15 each), receiving either a control nursery diet (NC-CD, CFCD-CD) or a diet supplemented with either GLN (NC-GLN, CFCD-GLN) or with AG (NC-AG, CFCD-AG). Litters that were creep fed with diets containing GLN or AG also were supplemented with those amino acids in the nursery diets (CFGLN-GLN, CFAG-AG). Glutamine was added at 1% in all three post-weaning diet phases and AG was added at .88% in phase 1 and 2 and at .66% in phase 3. Feed conversion (feed/gain) showed means among treatment means close to significance (P = 0.056) and Tukey's test for pairwise mean comparisons showed that Pigs in the CFGLN-GLN group had the best feed conversion (feed/gain) in the first three-week period post-weaning, exceeding (P = 0.044) controls (CFCD-CD) by 34%. The NC-AG group had (P = 0.02) the greatest feed intake in the last three week of the study, exceeding controls (CFCD-CD) by 12%. CFGLN-GLN, CFCD-GLN and sow reared (SR) pigs had the greatest (P = 0.049) villi height exceeding the CFCD-AG group by 18%, 20% and 19% respectively. The CFAG-AG group had the deepest (P = 0.001) crypts among all treatments. CFGLN-GLN, CFCD-GLN and SR groups had the greatest (P = 0.001) number of cells proliferating (PCNA) exceeding those in the NC-CD group by 43%, 54% and 63% respectively

  14. Controlling the prion propensity of glutamine/asparagine-rich proteins.

    Paul, Kacy R; Ross, Eric D

    2015-01-01

    The yeast Saccharomyces cerevisiae can harbor a number of distinct prions. Most of the yeast prion proteins contain a glutamine/asparagine (Q/N) rich region that drives prion formation. Prion-like domains, defined as regions with high compositional similarity to yeast prion domains, are common in eukaryotic proteomes, and mutations in various human proteins containing prion-like domains have been linked to degenerative diseases, including amyotrophic lateral sclerosis. Here, we discuss a recent study in which we utilized two strategies to generate prion activity in non-prion Q/N-rich domains. First, we made targeted mutations in four non-prion Q/N-rich domains, replacing predicted prion-inhibiting amino acids with prion-promoting amino acids. All four mutants formed foci when expressed in yeast, and two acquired bona fide prion activity. Prion activity could be generated with as few as two mutations, suggesting that many non-prion Q/N-rich proteins may be just a small number of mutations from acquiring aggregation or prion activity. Second, we created tandem repeats of short prion-prone segments, and observed length-dependent prion activity. These studies demonstrate the considerable progress that has been made in understanding the sequence basis for aggregation of prion and prion-like domains, and suggest possible mechanisms by which new prion domains could evolve.

  15. Neurotransmitter synthesis from CNS glutamine for central control of breathing

    Hoop, B.; Systrom, D.; Chiang, C.H.; Shih, V.E.; Kazemi, H.

    1986-01-01

    The maximum rate at which CNS glutamine (GLN) derived from glutamate (GLU) can be sequestered for synthesis of neurotransmitter GLU and/or γ-aminobutyric acid (GABA) has been determined in pentobarbital-anesthetized dogs. A total of 57 animals were studied under normal, hypoxic (Pa/sub O2/ greater than or equal to 20 mmHg), or hypercapnic (Pa/sub CO2/ less than or equal to 71 mm Hg) conditions. Thirteen of these were bilaterally vagotomized and carotid body denervated and studied only under normoxic or hypoxic conditions. In 5 animals cerebrospinal fluid GLN transfer rate constant k was measured using 13 N-ammonia tracer. Measured cerebral cortical (CC) and medullary (MED) GLN concentrations c are found to vary with GLU metabolic rate r according to c-C/sub m/r/(r+R), where r, the product of k and corresponding tissue GLU concentration, is assumed equal to the maximum GLN metabolic rate via pathways other than for neurotransmitter synthesis. The constants C/sub m/ and R are the predicted maximum GLN concentration and its maximum rate of sequestration for neurotransmitter synthesis, respectively. For both CNS tissue types in all animals, C/sub m/ = 20.9 +- 7.4 (SD) mmoles/kg wet wt(mM) and R = 6.2 +- 2.3 mM/min. These values are consistent with results obtained in anesthetized rats

  16. Glutamine Attenuates Acute Lung Injury Caused by Acid Aspiration

    Chih-Cheng Lai

    2014-08-01

    Full Text Available Inadequate ventilator settings may cause overwhelming inflammatory responses associated with ventilator-induced lung injury (VILI in patients with acute respiratory distress syndrome (ARDS. Here, we examined potential benefits of glutamine (GLN on a two-hit model for VILI after acid aspiration-induced lung injury in rats. Rats were intratracheally challenged with hydrochloric acid as a first hit to induce lung inflammation, then randomly received intravenous GLN or lactated Ringer’s solution (vehicle control thirty min before different ventilator strategies. Rats were then randomized to receive mechanical ventilation as a second hit with a high tidal volume (TV of 15 mL/kg and zero positive end-expiratory pressure (PEEP or a low TV of 6 mL/kg with PEEP of 5 cm H2O. We evaluated lung oxygenation, inflammation, mechanics, and histology. After ventilator use for 4 h, high TV resulted in greater lung injury physiologic and biologic indices. Compared with vehicle treated rats, GLN administration attenuated lung injury, with improved oxygenation and static compliance, and decreased respiratory elastance, lung edema, extended lung destruction (lung injury scores and lung histology, neutrophil recruitment in the lung, and cytokine production. Thus, GLN administration improved the physiologic and biologic profiles of this experimental model of VILI based on the two-hit theory.

  17. Conformation-specific spectroscopy of capped glutamine-containing peptides: role of a single glutamine residue on peptide backbone preferences.

    Walsh, Patrick S; Dean, Jacob C; McBurney, Carl; Kang, Hyuk; Gellman, Samuel H; Zwier, Timothy S

    2016-04-28

    The conformational preferences of a series of short, aromatic-capped, glutamine-containing peptides have been studied under jet-cooled conditions in the gas phase. This work seeks a bottom-up understanding of the role played by glutamine residues in directing peptide structures that lead to neurodegenerative diseases. Resonant ion-dip infrared (RIDIR) spectroscopy is used to record single-conformation infrared spectra in the NH stretch, amide I and amide II regions. Comparison of the experimental spectra with the predictions of calculations carried out at the DFT M05-2X/6-31+G(d) level of theory lead to firm assignments for the H-bonding architectures of a total of eight conformers of four molecules, including three in Z-Gln-OH, one in Z-Gln-NHMe, three in Ac-Gln-NHBn, and one in Ac-Ala-Gln-NHBn. The Gln side chain engages actively in forming H-bonds with nearest-neighbor amide groups, forming C8 H-bonds to the C-terminal side, C9 H-bonds to the N-terminal side, and an amide-stacked geometry, all with an extended (C5) peptide backbone about the Gln residue. The Gln side chain also stabilizes an inverse γ-turn in the peptide backbone by forming a pair of H-bonds that bridge the γ-turn and stabilize it. Finally, the entire conformer population of Ac-Ala-Gln-NHBn is funneled into a single structure that incorporates the peptide backbone in a type I β-turn, stabilized by the Gln side chain forming a C7 H-bond to the central amide group in the β-turn not otherwise involved in a hydrogen bond. This β-turn backbone structure is nearly identical to that observed in a series of X-(AQ)-Y β-turns in the protein data bank, demonstrating that the gas-phase structure is robust to perturbations imposed by the crystalline protein environment.

  18. Plasma glutamine levels before cardiac surgery are related to post-surgery infections; an observational study

    Hanneke Buter

    2016-11-01

    Full Text Available Abstract Background A low plasma glutamine level was found in 34% of patients after elective cardiothoracic surgery. This could be a result of the inflammation caused by surgical stress or the use of extracorporeal circulation (ECC. But it is also possible that plasma glutamine levels were already lowered before surgery and reflect an impaired metabolic state and a higher likelihood to develop complications. In the present study plasma glutamine levels were measured before and after cardiac surgery and we questioned whether there is a relation between plasma glutamine levels and duration of ECC and the occurrence of postoperative infections. Methods We performed a single-centre prospective, observational study in a closed-format, 20-bed, mixed ICU in a tertiary teaching hospital. We included consecutive patients after elective cardiac surgery with use of extracorporeal circulation. Blood samples were collected on the day prior to surgery and at admission on the ICU. The study was approved by the local Medical Ethics Committee (Regional Review Committee Patient-related Research, Medical Centre Leeuwarden, nWMO 115, April 28th 2015. Results Ninety patients were included. Pre-operative plasma glutamine level was 0.42 ± 0.10 mmol/l and post-operative 0.38 ± 0.09 mmol/l (p < 0.001. There was no relation between duration of extracorporeal circulation or aortic occlusion time and changes in plasma glutamine levels. A logistic regression analysis showed a significant correlation between the presence of a positive culture during the post-operative course and pre-operative plasma glutamine levels (p = 0.04. Conclusion Plasma glutamine levels are significantly lower just after cardiac surgery compared to pre-operative levels. We did not find a relation between the decrease in plasma glutamine levels and the duration of extracorporeal circulation or aortic clamp time. There was a correlation between pre-operative plasma glutamine levels

  19. The effect of free glutamine and peptide ingestion on the rate of muscle glycogen resynthesis in man

    Van Hall, Gerrit; Saris, W H; van de Schoor, P A

    2000-01-01

    hydrolysate (26% glutamine) and 3) a whey hydrolysate (6.6% glutamine). Plasma glutamine, decreased by approximately 20% during recovery with ingestion of the control drink, no changes with ingestion of the protein hydrolysates drinks, and a 2-fold increase with ingestion of the free glutamine drinks....... The rate of glycogen resynthesis was not significantly different in the four tests: 28 +/- 5, 26 +/- 6, 33 +/- 4, and 34 +/- 3 mmol glucosyl units x kg(-1) dry weight muscle x h(-1) for the control, glutamine, wheat- and whey hydrolysate ingestion, respectively. It is concluded that ingestion...... of a glutamine/carbohydrate mixture does not increase the rate of glycogen resynthesis in muscle. Glycogen resynthesis rates were higher, although not statistically significant, after ingestion of the drink containing the wheat (21 +/- 8%) and whey protein hydrolysate (20 +/- 6%) compared to ingestion...

  20. The effect of glutamine administration on urinary ammonium excretion in normal subjects and patients with renal disease.

    Welbourne, T; Weber, M; Bank, N

    1972-07-01

    The effect of acute changes in the delivery rate of glutamine to the kidney on urinary ammonium excretion was studied in man. Healthy subjects and patients with intrinsic renal disease were studied under three different acid-base conditions: unaltered acid-base balance; NH(4)Cl-induced acidosis; and NaHCO(3)-induced alkalosis. Anhydrous L-glutamine was administered orally in a single dose of 260 mmoles during each of these three acid-base states. We found that endogenous venous plasma glutamine concentration fell during acidosis and rose during alkalosis in both healthy subjects and patients with renal disease. In healthy subjects, orally administered glutamine raised plasma glutamine concentration markedly over a 2-3 hr period. This was accompanied by an increase in urinary ammonium excretion and a rise in urine pH under normal acid-base conditions and during metabolic acidosis. No increase in ammonium excretion occurred when glutamine was administered during metabolic alkalosis in spite of an equivalent rise in plasma glutamine concentration. In patients with renal disease, endogenous venous plasma glutamine concentration was lower than in healthy subjects, perhaps as a result of mild metabolic acidosis. Acute oral glutamine loading failed to increase urinary ammonium excretion significantly during either unaltered acid-base conditions or after NH(4)Cl-induced acidosis, even though plasma glutamine rose as high as in healthy subjects. We conclude from these observations that glutamine delivery to the kidney is a rate-limiting factor for ammonium excretion in healthy subjects, both before and after cellular enzyme adaptation induced by metabolic acidosis. In contrast, in patients with renal disease, glutamine delivery is not rate-limiting for ammonium excretion. Presumably other factors, such as surviving renal mass and the activity of intracellular enzymes necessary for ammonia synthesis limit ammonium excretion in these patients.

  1. Effects of L-glutamine supplementation on maternal and fetal hemodynamics in gestating ewes exposed to alcohol

    Sawant, Onkar B.; Ramadoss, Jayanth; Hankins, Gary D.; Wu, Guoyao; Washburn, Shannon E.

    2014-01-01

    Not much is known about effects of gestational alcohol exposure on maternal and fetal cardiovascular adaptations. This study determined whether maternal binge alcohol exposure and L-glutamine supplementation could affect maternal-fetal hemodynamics and fetal regional brain blood flow during the brain growth spurt period. Pregnant sheep were randomly assigned to one of four groups: saline control, alcohol (1.75–2.5 g/kg body weight), glutamine (100 mg/kg body weight) or alcohol + glutamine. A ...

  2. GLUTAMIN MEMPERCEPAT WAKTU PEMULIHAN JUMLAH SEL LIMFOSIT LIEN SETELAH AKTIVITAS FISIK MAKSIMAL PADA MENCIT (GLUTAMINE SHORTENS RECOVERY TIME OF LIEN LYMPHOCYTES AFTER EXCESSIVE PHYSICAL ACTIVITY IN MICE

    I Made Jawi

    2007-06-01

    Full Text Available The immunologic? system? of the body requires? suitable recovery time after? excessive physical? activity. The recovery? time of spleen lymphocytes after? excessive? physical activity? in one? investigation was? 3? days. The? aim? of this? research is to identify?the role of? glutamine? in shortening? the recovery time of? spleen? lymphocytes? after?excessive physical activity. The? study? was conducted? on? 70? adults? Balb/c mice which? were? divided? into? 7? groups, with? a randomized control? group post-test? only design. In this? study? an observation? was made on? spleen? lymphocytes? of control, after? excessive? physical activity(in the the? form of swimming? until? near? drowning with glutamine,? without glutamine ?and? after? the? recovery time of 1 and 2 days? of? each of? the 10 mice. Spleen? lymphocytes? were? counted in spleen preparation by mikroskop. The data obtained were tested? by? one way Anova. The findings? showed? that the number of spleen? lymphocytes significantly decrease after? excessive? physical activity?? in the?glutamine? and? non glutamine groups ( p < 0,05.The number of spleen? lymphocytes? was? not different as compered to control group or returned? to normal after recovery? time? of 1? day? in? the? glutamine group (p > 0,05 .?In the nonglutamine group the number of spleen lymphocytes was different from control group until 2 days (p<0,05 . From this finding it can be concluded that glutamine? shortens the recovery time of spleen lymphocytes? after? excessive? physical activity in mice.

  3. Glutamine reduces postprandial glycemia and augments the glucagon-like peptide-1 response in type 2 diabetes patients

    Samocha-Bonet, Dorit; Wong, Olivia; Synnott, Emma-Leigh

    2011-01-01

    Impaired glucagon-like peptide (GLP-1) secretion or response may contribute to ineffective insulin release in type 2 diabetes. The conditionally essential amino acid glutamine stimulates GLP-1 secretion in vitro and in vivo. In a randomized, crossover study, we evaluated the effect of oral...... glutamine, with or without sitagliptin (SIT), on postprandial glycemia and GLP-1 concentration in 15 type 2 diabetes patients (glycated hemoglobin 6.5 ± 0.6%). Participants ingested a low-fat meal (5% fat) after receiving either water (control), 30 g l-glutamine (Gln-30), 15 g L-glutamine (Gln-15), 100 mg...... concentration and limiting postprandial glycemia in type 2 diabetes....

  4. Differential inhibition of adenylylated and deadenylylated forms of M. tuberculosis glutamine synthetase as a drug discovery platform.

    A Theron

    Full Text Available Glutamine synthetase is a ubiquitous central enzyme in nitrogen metabolism that is controlled by up to four regulatory mechanisms, including adenylylation of some or all of the twelve subunits by adenylyl transferase. It is considered a potential therapeutic target for the treatment of tuberculosis, being essential for the growth of Mycobacterium tuberculosis, and is found extracellularly only in the pathogenic Mycobacterium strains. Human glutamine synthetase is not regulated by the adenylylation mechanism, so the adenylylated form of bacterial glutamine synthetase is of particular interest. Previously published reports show that, when M. tuberculosis glutamine synthetase is expressed in Escherichia coli, the E. coli adenylyl transferase does not optimally adenylylate the M. tuberculosis glutamine synthetase. Here, we demonstrate the production of soluble adenylylated M. tuberulosis glutamine synthetase in E. coli by the co-expression of M. tuberculosis glutamine synthetase and M. tuberculosis adenylyl transferase. The differential inhibition of adenylylated M. tuberulosis glutamine synthetase and deadenylylated M. tuberulosis glutamine synthetase by ATP based scaffold inhibitors are reported. Compounds selected on the basis of their enzyme inhibition were also shown to inhibit M. tuberculosis in the BACTEC 460TB™ assay as well as the intracellular inhibition of M. tuberculosis in a mouse bone-marrow derived macrophage assay.

  5. Differential inhibition of adenylylated and deadenylylated forms of M. tuberculosis glutamine synthetase as a drug discovery platform.

    Theron, A; Roth, R L; Hoppe, H; Parkinson, C; van der Westhuyzen, C W; Stoychev, S; Wiid, I; Pietersen, R D; Baker, B; Kenyon, C P

    2017-01-01

    Glutamine synthetase is a ubiquitous central enzyme in nitrogen metabolism that is controlled by up to four regulatory mechanisms, including adenylylation of some or all of the twelve subunits by adenylyl transferase. It is considered a potential therapeutic target for the treatment of tuberculosis, being essential for the growth of Mycobacterium tuberculosis, and is found extracellularly only in the pathogenic Mycobacterium strains. Human glutamine synthetase is not regulated by the adenylylation mechanism, so the adenylylated form of bacterial glutamine synthetase is of particular interest. Previously published reports show that, when M. tuberculosis glutamine synthetase is expressed in Escherichia coli, the E. coli adenylyl transferase does not optimally adenylylate the M. tuberculosis glutamine synthetase. Here, we demonstrate the production of soluble adenylylated M. tuberulosis glutamine synthetase in E. coli by the co-expression of M. tuberculosis glutamine synthetase and M. tuberculosis adenylyl transferase. The differential inhibition of adenylylated M. tuberulosis glutamine synthetase and deadenylylated M. tuberulosis glutamine synthetase by ATP based scaffold inhibitors are reported. Compounds selected on the basis of their enzyme inhibition were also shown to inhibit M. tuberculosis in the BACTEC 460TB™ assay as well as the intracellular inhibition of M. tuberculosis in a mouse bone-marrow derived macrophage assay.

  6. Greater efficiency of water use in poplar clones having a delayed response of mesophyll conductance to drought.

    Théroux Rancourt, Guillaume; Éthier, Gilbert; Pepin, Steeve

    2015-02-01

    Improvement of water use efficiency is a key objective to improve the sustainability of cultivated plants, especially fast growing species with high water consumption like poplar. It is well known that water use efficiency (WUE) varies considerably among poplar genotypes, and it was recently suggested that the use of the mesophyll-to-stomatal conductance ratio (gm/gs) would be an appropriate trait to improve WUE. The responses of 7-week-old cuttings of four hybrid poplar clones and one native Balsam poplar (Populus balsamifera L.) to a water stress-recovery cycle were examined to evaluate the relation between the gm/gs ratio and transpiration efficiency (TE), a leaf-level component of WUE. A contrasting gs response to water stress was observed among the five clones, from stomatal closure early on during soil drying up to limited closure in Balsam poplar. However in the hybrids, the decline in gm was consistently delayed by a few days compared with gs. Moreover, in the most water use-efficient hybrids, the recovery following rehydration occurred faster for gm than for gs. Thus, the delay in the response of gm to drought and its faster recovery upon rewatering increased the gm/gs of the hybrids and this ratio scaled positively with TE. Our results support the use of the gm/gs ratio to select genotypes with improved WUE, and the notion that breeding strategies focusing mainly on stomatal responses to soil drying should also look for a strong curvilinearity between net carbon assimilation rate and gs, the indication of a significant increase in gm/gs in the earlier stages of stomatal closure. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  7. Brain glutamine synthesis requires neuronal-born aspartate as amino donor for glial glutamate formation.

    Pardo, Beatriz; Rodrigues, Tiago B; Contreras, Laura; Garzón, Miguel; Llorente-Folch, Irene; Kobayashi, Keiko; Saheki, Takeyori; Cerdan, Sebastian; Satrústegui, Jorgina

    2011-01-01

    The glutamate-glutamine cycle faces a drain of glutamate by oxidation, which is balanced by the anaplerotic synthesis of glutamate and glutamine in astrocytes. De novo synthesis of glutamate by astrocytes requires an amino group whose origin is unknown. The deficiency in Aralar/AGC1, the main mitochondrial carrier for aspartate-glutamate expressed in brain, results in a drastic fall in brain glutamine production but a modest decrease in brain glutamate levels, which is not due to decreases in neuronal or synaptosomal glutamate content. In vivo (13)C nuclear magnetic resonance labeling with (13)C(2)acetate or (1-(13)C) glucose showed that the drop in brain glutamine is due to a failure in glial glutamate synthesis. Aralar deficiency induces a decrease in aspartate content, an increase in lactate production, and lactate-to-pyruvate ratio in cultured neurons but not in cultured astrocytes, indicating that Aralar is only functional in neurons. We find that aspartate, but not other amino acids, increases glutamate synthesis in both control and aralar-deficient astrocytes, mainly by serving as amino donor. These findings suggest the existence of a neuron-to-astrocyte aspartate transcellular pathway required for astrocyte glutamate synthesis and subsequent glutamine formation. This pathway may provide a mechanism to transfer neuronal-born redox equivalents to mitochondria in astrocytes.

  8. Glutamine Supplemented Parenteral Nutrition to Prevent Ventilator-Associated Pneumonia in the Intensive Care Unit

    Meltem Türkay Aydoğmuş

    2012-12-01

    Full Text Available Objective: Ventilator-associated pneumonia (VAP is a form of nosocomial pneumonia that increases patient morbidity and mortality, length of hospital stay, and healthcare costs. Glutamine preserves the intestinal mucosal structure, increases immune function, and reduces harmful changes in gut permeability in patients receiving total parenteral nutrition (TPN. We hypothesized that TPN supplemented by glutamine might prevent the development of VAP in patients on mechanical ventilator support in the intensive care unit (ICU. Material and Methods: With the approval of the ethics committee and informed consent from relatives, 60 patients who were followed in the ICU with mechanical ventilator support were included in our study. Patients were divided into three groups. The first group received enteral nutrition (n=20, and the second was prescribed TPN (n=20 while the third group was given glutamine-supplemented TPN (n=20. C-reactive protein (CRP, sedimentation rate, body temperature, development of purulent secretions, increase in the amount of secretions, changes in the characteristics of secretions and an increase in requirement of deep tracheal aspiration were monitored for seven days by daily examination and radiographs. Results: No statistically significant difference was found among groups in terms of development of VAP (p=0.622. Conclusion: Although VAP developed at a lower rate in the glutamine-supplemented TPN group, no statistically significant difference was found among any of the groups. Glutamine-supplemented TPN may have no superiority over unsupplemented enteral and TPN in preventing VAP.

  9. Glutamine supplemented parenteral nutrition to prevent ventilator-associated pneumonia in the intensive care unit.

    Aydoğmuş, Meltem Türkay; Tomak, Yakup; Tekin, Murat; Katı, Ismail; Hüseyinoğlu, Urfettin

    2012-12-01

    Ventilator-associated pneumonia (VAP) is a form of nosocomial pneumonia that increases patient morbidity and mortality, length of hospital stay, and healthcare costs. Glutamine preserves the intestinal mucosal structure, increases immune function, and reduces harmful changes in gut permeability in patients receiving total parenteral nutrition (TPN). We hypothesized that TPN supplemented by glutamine might prevent the development of VAP in patients on mechanical ventilator support in the intensive care unit (ICU). With the approval of the ethics committee and informed consent from relatives, 60 patients who were followed in the ICU with mechanical ventilator support were included in our study. Patients were divided into three groups. The first group received enteral nutrition (n=20), and the second was prescribed TPN (n=20) while the third group was given glutamine-supplemented TPN (n=20). C-reactive protein (CRP), sedimentation rate, body temperature, development of purulent secretions, increase in the amount of secretions, changes in the characteristics of secretions and an increase in requirement of deep tracheal aspiration were monitored for seven days by daily examination and radiographs. No statistically significant difference was found among groups in terms of development of VAP (p=0.622). Although VAP developed at a lower rate in the glutamine-supplemented TPN group, no statistically significant difference was found among any of the groups. Glutamine-supplemented TPN may have no superiority over unsupplemented enteral and TPN in preventing VAP.

  10. Response to Dietary Supplementation of Glutamine in Broiler Chickens Subjected to Transportation Stress

    Majid SHAKERI

    2016-07-01

    Full Text Available The main purpose of this study was to determine effects of glutamine supplementation on performance and blood parameters including Hsp70 and acute phase protein when chicken were subjected to transportation stress. A total of four hundred day-old-male cobb-500 chicks were obtained directly from a local hatchery. The chicks were allotted to two groups as: immediate placement (1 hour after hatching with access to feed and water and placement after 24h transportation without access to feed and water. The experiment consisted of a factorial arrangement of 2 different diets and 2 different time of placement. Chicks from each placement group were fed either basal diet or basal diet + 1% glutamine from 1 to 21 days of age. The results indicated that dietary glutamine improved the body weight gain and feed conversion ratio significantly when chicks were subjected to delayed or immediate placement. In conclusion, supplementing chicken with glutamine in diet can reduce negative effects of delayed access to feed and water during transportation. Moreover, APP concentration and HSP70 level were positively affected when chicks supplemented with glutamine in the diet.

  11. Carbon dioxide diffusion across stomata and mesophyll and photo-biochemical processes as affected by growth CO2 and phosphorus nutrition in cotton.

    Singh, Shardendu K; Badgujar, Girish; Reddy, Vangimalla R; Fleisher, David H; Bunce, James A

    2013-06-15

    Nutrients such as phosphorus may exert a major control over plant response to rising atmospheric carbon dioxide concentration (CO2), which is projected to double by the end of the 21st century. Elevated CO2 may overcome the diffusional limitations to photosynthesis posed by stomata and mesophyll and alter the photo-biochemical limitations resulting from phosphorus deficiency. To evaluate these ideas, cotton (Gossypium hirsutum) was grown in controlled environment growth chambers with three levels of phosphate (Pi) supply (0.2, 0.05 and 0.01mM) and two levels of CO2 concentration (ambient 400 and elevated 800μmolmol(-1)) under optimum temperature and irrigation. Phosphate deficiency drastically inhibited photosynthetic characteristics and decreased cotton growth for both CO2 treatments. Under Pi stress, an apparent limitation to the photosynthetic potential was evident by CO2 diffusion through stomata and mesophyll, impairment of photosystem functioning and inhibition of biochemical process including the carboxylation efficiency of ribulose-1,5-bisphosphate carboxylase/oxyganase and the rate of ribulose-1,5-bisphosphate regeneration. The diffusional limitation posed by mesophyll was up to 58% greater than the limitation due to stomatal conductance (gs) under Pi stress. As expected, elevated CO2 reduced these diffusional limitations to photosynthesis across Pi levels; however, it failed to reduce the photo-biochemical limitations to photosynthesis in phosphorus deficient plants. Acclimation/down regulation of photosynthetic capacity was evident under elevated CO2 across Pi treatments. Despite a decrease in phosphorus, nitrogen and chlorophyll concentrations in leaf tissue and reduced stomatal conductance at elevated CO2, the rate of photosynthesis per unit leaf area when measured at the growth CO2 concentration tended to be higher for all except the lowest Pi treatment. Nevertheless, plant biomass increased at elevated CO2 across Pi nutrition with taller plants

  12. Nitrogen Starvation and TorC1 Inhibition Differentially Affect Nuclear Localization of the Gln3 and Gat1 Transcription Factors Through the Rare Glutamine tRNACUG in Saccharomyces cerevisiae

    Tate, Jennifer J.; Rai, Rajendra; Cooper, Terrance G.

    2015-01-01

    A leucine, leucyl-tRNA synthetase–dependent pathway activates TorC1 kinase and its downstream stimulation of protein synthesis, a major nitrogen consumer. We previously demonstrated, however, that control of Gln3, a transcription activator of catabolic genes whose products generate the nitrogenous precursors for protein synthesis, is not subject to leucine-dependent TorC1 activation. This led us to conclude that excess nitrogen-dependent down-regulation of Gln3 occurs via a second mechanism that is independent of leucine-dependent TorC1 activation. A major site of Gln3 and Gat1 (another GATA-binding transcription activator) control occurs at their access to the nucleus. In excess nitrogen, Gln3 and Gat1 are sequestered in the cytoplasm in a Ure2-dependent manner. They become nuclear and activate transcription when nitrogen becomes limiting. Long-term nitrogen starvation and treatment of cells with the glutamine synthetase inhibitor methionine sulfoximine (Msx) also elicit nuclear Gln3 localization. The sensitivity of Gln3 localization to glutamine and inhibition of glutamine synthesis prompted us to investigate the effects of a glutamine tRNA mutation (sup70-65) on nitrogen-responsive control of Gln3 and Gat1. We found that nuclear Gln3 localization elicited by short- and long-term nitrogen starvation; growth in a poor, derepressive medium; Msx or rapamycin treatment; or ure2Δ mutation is abolished in a sup70-65 mutant. However, nuclear Gat1 localization, which also exhibits a glutamine tRNACUG requirement for its response to short-term nitrogen starvation or growth in proline medium or a ure2Δ mutation, does not require tRNACUG for its response to rapamycin. Also, in contrast with Gln3, Gat1 localization does not respond to long-term nitrogen starvation. These observations demonstrate the existence of a specific nitrogen-responsive component participating in the control of Gln3 and Gat1 localization and their downstream production of nitrogenous precursors

  13. 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings

    Thakkar, Katharine N; Rösler, Lara; Wijnen, Jannie P; Boer, Vincent O.; Klomp, Dennis W J; Cahn, Wiepke; Kahn, René S; Neggers, Sebastiaan F W

    BACKGROUND: The N-methyl-D-aspartate receptor hypofunction model of schizophrenia predicts dysfunction in both glutamatergic and gamma-aminobutyric acidergic (GABAergic) transmission. We addressed this hypothesis by measuring GABA, glutamate, glutamine, and the sum of glutamine plus glutamate

  14. The role of mesophyll conductance during water stress and recovery in tobacco (Nicotiana sylvestris): acclimation or limitation?

    Galle, Alexander; Florez-Sarasa, Igor; Tomas, Magdalena; Pou, Alicia; Medrano, Hipolito; Ribas-Carbo, Miquel; Flexas, Jaume

    2009-01-01

    While the responses of photosynthesis to water stress have been widely studied, acclimation to sustained water stress and recovery after re-watering is poorly understood. In particular, the factors limiting photosynthesis under these conditions, and their possible interactions with other environmental conditions, are unknown. To assess these issues, changes of photosynthetic CO(2) assimilation (A(N)) and its underlying limitations were followed during prolonged water stress and subsequent re-watering in tobacco (Nicotiana sylvestris) plants growing under three different climatic conditions: outdoors in summer, outdoors in spring, and indoors in a growth chamber. In particular, the regulation of stomatal conductance (g(s)), mesophyll conductance to CO(2) (g(m)), leaf photochemistry (chlorophyll fluorescence), and biochemistry (V(c,max)) were assessed. Leaf gas exchange and chlorophyll fluorescence data revealed that water stress induced a similar degree of stomatal closure and decreased A(N) under all three conditions, while V(c,max) was unaffected. However, the behaviour of g(m) differed depending on the climatic conditions. In outdoor plants, g(m) strongly declined with water stress, but it recovered rapidly (1-2 d) after re-watering in spring while it remained low many days after re-watering in summer. In indoor plants, g(m) initially declined with water stress, but then recovered to control values during the acclimation period. These differences were reflected in different velocities of recovery of A(N) after re-watering, being the slowest in outdoor summer plants and the fastest in indoor plants. It is suggested that these differences among the experiments are related to the prevailing climatic conditions, i.e. to the fact that stress factors other than water stress have been superimposed (e.g. excessive light and elevated temperature). In conclusion, besides g(s), g(m) contributes greatly to the limitation of photosynthesis during water stress and during

  15. Dose intercomparison for 400–500 keV electrons using FWT-60 film and glutamine (spectrophotometric readout) dosimeters

    Gupta, B. L.; Nilekani, S. R.; Gehringer, P.

    1986-01-01

    This paper describes the dose and the depth dose measurements with FWT-60 film and glutamine (Spectrophotometric readout) dosimeters for 400–500 keV electrons. The glutamine powder was spread uniformly in polyethylene bags and the powder thickness in each bag was 5 mg cm−2. Both techniques show...

  16. Cerebral glucose metabolism and the glutamine cycle as detected by in vivo and in vitro 13C NMR spectroscopy.

    García-Espinosa, María A; Rodrigues, Tiago B; Sierra, Alejandra; Benito, Marina; Fonseca, Carla; Gray, Heather L; Bartnik, Brenda L; García-Martín, María L; Ballesteros, Paloma; Cerdán, Sebastián

    2004-01-01

    We review briefly 13C NMR studies of cerebral glucose metabolism with an emphasis on the roles of glial energetics and the glutamine cycle. Mathematical modeling analysis of in vivo 13C turnover experiments from the C4 carbons of glutamate and glutamine are consistent with: (i) the glutamine cycle being the major cerebral metabolic route supporting glutamatergic neurotransmission, (ii) glial glutamine synthesis being stoichiometrically coupled to glycolytic ATP production, (iii) glutamine serving as the main precursor of neurotransmitter glutamate and (iv) glutamatergic neurotransmission being supported by lactate oxidation in the neurons in a process accounting for 60-80% of the energy derived from glucose catabolism. However, more recent experimental approaches using inhibitors of the glial tricarboxylic acid (TCA) cycle (trifluoroacetic acid, TFA) or of glutamine synthase (methionine sulfoximine, MSO) reveal that a considerable portion of the energy required to support glutamine synthesis is derived from the oxidative metabolism of glucose in the astroglia and that a significant amount of the neurotransmitter glutamate is produced from neuronal glucose or lactate rather than from glial glutamine. Moreover, a redox switch has been proposed that allows the neurons to use either glucose or lactate as substrates for oxidation, depending on the relative availability of these fuels under resting or activation conditions, respectively. Together, these results suggest that the coupling mechanisms between neuronal and glial metabolism are more complex than initially envisioned.

  17. Maternal L-glutamine supplementation prevents prenatal alcohol exposure-induced fetal growth restriction in an ovine model.

    Sawant, Onkar B; Wu, Guoyao; Washburn, Shannon E

    2015-06-01

    Prenatal alcohol exposure is known to cause fetal growth restriction and disturbances in amino acid bioavailability. Alterations in these parameters can persist into adulthood and low birth weight can lead to altered fetal programming. Glutamine has been associated with the synthesis of other amino acids, an increase in protein synthesis and it is used clinically as a nutrient supplement for low birth weight infants. The aim of this study was to explore the effect of repeated maternal alcohol exposure and L-glutamine supplementation on fetal growth and amino acid bioavailability during the third trimester-equivalent period in an ovine model. Pregnant sheep were randomly assigned to four groups, saline control, alcohol (1.75-2.5 g/kg), glutamine (100 mg/kg, three times daily) or alcohol + glutamine. In this study, a weekend binge drinking model was followed where treatment was done 3 days per week in succession from gestational day (GD) 109-132 (normal term ~147). Maternal alcohol exposure significantly reduced fetal body weight, height, length, thoracic girth and brain weight, and resulted in decreased amino acid bioavailability in fetal plasma and placental fluids. Maternal glutamine supplementation successfully mitigated alcohol-induced fetal growth restriction and improved the bioavailability of glutamine and glutamine-related amino acids such as glycine, arginine, and asparagine in the fetal compartment. All together, these findings show that L-glutamine supplementation enhances amino acid availability in the fetus and prevents alcohol-induced fetal growth restriction.

  18. Effect of carbohydrate supplementation on plasma glutamine during prolonged exercise and recovery

    Van Hall, Gerrit; Saris, W H; Wagenmakers, A J

    1998-01-01

    . Eight well-trained subjects cycled at an alternating workload of 50 and 80% Wmax until exhaustion (59 to 140 min). During the exercise bout the subjects received either water (control) or a carbohydrate (CHO) drink (83 g CHO x l(-1), 2 ml x kg(-1) per kg body weight every 15 min). Plasma glutamine......%) decreased below the pre-exercise level. The plasma alanine and the total amino acid concentration was still suppressed after 7 h of recovery. In conclusion, carbohydrate supplementation had neither an effect during exercise nor during recovery on the concentration of plasma glutamine or other amino acids...

  19. Effect of glutamine synthetase inhibition on brain and interorgan ammonia metabolism in bile duct ligated rats

    Fries, Andreas W; Dadsetan, Sherry; Keiding, Susanne

    2014-01-01

    , and aspartate and incorporation of (15)NH4(+) into these amino acids in brain, liver, muscle, kidney, and plasma were similar in sham and BDL rats treated with saline. Methionine sulfoximine reduced glutamine concentrations in liver, kidney, and plasma but not in brain and muscle; MSO reduced incorporation...... of (15)NH4(+) into glutamine in all tissues. It did not affect alanine concentrations in any of the tissues but plasma alanine concentration increased; incorporation of (15)NH4(+) into alanine was increased in brain in sham and BDL rats and in kidney in sham rats. It inhibited GS in all tissues examined...

  20. Structure-Function Relationship of Transporters in the Glutamate–Glutamine Cycle of the Central Nervous System

    Mariko Kato Hayashi

    2018-04-01

    Full Text Available Many kinds of transporters contribute to glutamatergic excitatory synaptic transmission. Glutamate is loaded into synaptic vesicles by vesicular glutamate transporters to be released from presynaptic terminals. After synaptic vesicle release, glutamate is taken up by neurons or astrocytes to terminate the signal and to prepare for the next signal. Glutamate transporters on the plasma membrane are responsible for transporting glutamate from extracellular fluid to cytoplasm. Glutamate taken up by astrocyte is converted to glutamine by glutamine synthetase and transported back to neurons through glutamine transporters on the plasma membranes of the astrocytes and then on neurons. Glutamine is converted back to glutamate by glutaminase in the neuronal cytoplasm and then loaded into synaptic vesicles again. Here, the structures of glutamate transporters and glutamine transporters, their conformational changes, and how they use electrochemical gradients of various ions for substrate transport are summarized. Pharmacological regulations of these transporters are also discussed.

  1. Arginyl-glutamine dipeptide or docosahexaenoic acid attenuates hyperoxia-induced small intestinal injury in neonatal mice.

    Li, Nan; Ma, Liya; Liu, Xueyan; Shaw, Lynn; Li Calzi, Sergio; Grant, Maria B; Neu, Josef

    2012-04-01

    Supplementation studies of glutamine, arginine, and docosahexaenoic acid (DHA) have established the safety of each of these nutrients in neonates; however, the potential for a more stable and soluble dipeptide, arginyl-glutamine (Arg-Gln) or DHA with anti-inflammatory properties, to exert benefits on hyperoxia-induced intestinal injury has not been investigated. Arg-Gln dipeptide has been shown to prevent retinal damage in a rodent model of oxygen-induced injury. The objective of the present study was to investigate whether Arg-Gln dipeptide or DHA could also attenuate markers of injury and inflammation to the small intestine in this same model. Seven-day-old mouse pups were placed with their dams in 75% oxygen for 5 days. After 5 days of hyperoxic exposure (P7-P12), pups were removed from hyperoxia and allowed to recover in atmospheric conditions for 5 days (P12-P17). Mouse pups received Arg-Gln (5g·kg·day) or DHA (5g·kg·day) or vehicle orally started on P12 through P17. Distal small intestine (DSI) histologic changes, myeloperoxidase (MPO), lactate dehydrogenase (LDH), inflammatory cytokines, and tissue apoptosis were evaluated. Hyperoxic mice showed a greater distortion of overall villus structure and with higher injury score (PDHA supplementation groups were more similar to the room air control group. Supplementation of Arg-Gln or DHA reduced hyperoxia-induced MPO activity (PDHA returned LDH activity to the levels of control. Hyperoxia induced apoptotic cell death in DSIs, and both Arg-Gln and DHA reversed this effect (PDHA may limit some inflammatory and apoptotic processes involved in hyperoxic-induced intestinal injury in neonatal mice.

  2. The glutamate-glutamine(GABA cycle: importance of late postnatal development and potential reciprocal interactions between biosynthesis and degradation

    Leif eHertz

    2013-05-01

    Full Text Available The gold standard for studies of glutamate-glutamine(GABA cycling and its connections to brain biosynthesis from glucose of glutamate and GABA and their subsequent metabolism are the elegant in vivo studies by 13C magnetic resonance spectroscopy (NMR, showing the large fluxes in the cycle. However, simpler experiments in intact brain tissue (e.g. immunohistochemistry, brain slices, cultured brain cells and mitochondria have also made important contributions to the understanding of details, mechanisms and functional consequences of glutamate/GABA biosynthesis and degradation. The purpose of this review is to attempt to integrate evidence from different sources regarding i the enzyme(s responsible for the initial conversion of -ketoglutarate to glutamate; ii the possibility that especially glutamate oxidation is essentially confined to astrocytes; and iii the ontogenetically very late onset and maturation of glutamine-glutamate(GABA cycle function. Pathway models based on the functional importance of aspartate for glutamate synthesis suggest the possibility of interacting pathways for biosynthesis and degradation of glutamate and GABA and the use of transamination as the default mechanism for initiation of glutamate oxidation. The late development and maturation are related to the late cortical gliogenesis and convert brain cortical function from being purely neuronal to becoming neuronal-astrocytic. This conversion is associated with huge increases in energy demand and production, and the character of potentially incurred gains of function are discussed. These may include alterations in learning mechanisms, in mice indicated by lack of pairing of odor learning with aversive stimuli in newborn animals but the development of such an association 10-12 days later. The possibility is suggested that analogous maturational changes may contribute to differences in the way learning is accomplished in the newborn human brain and during later development.

  3. Astrocytes and Glutamate Homoeostasis in Alzheimer's Disease: A Decrease in Glutamine Synthetase, But Not in Glutamate Transporter-1, in the Prefrontal Cortex

    Magdalena Kulijewicz-Nawrot

    2013-09-01

    Full Text Available Astrocytes control tissue equilibrium and hence define the homoeostasis and function of the CNS (central nervous system. Being principal homoeostatic cells, astroglia are fundamental for various forms of neuropathology, including AD (Alzheimer's disease. AD is a progressive neurodegenerative disorder characterized by the loss of cognitive functions due to specific lesions in mnesic-associated regions, including the mPFC (medial prefrontal cortex. Here, we analyzed the expression of GS (glutamine synthetase and GLT-1 (glutamate transporter-1 in astrocytes in the mPFC during the progression of AD in a triple-transgenic mouse model (3xTg-AD. GS is an astrocyte-specific enzyme, responsible for the intracellular conversion of glutamate into glutamine, whereas the removal of glutamate from the extracellular space is accomplished mainly by astroglia-specific GLT-1. We found a significant decrease in the numerical density (Nv, cells/mm3 of GS-positive astrocytes from early to middle ages (1–9 months; at the age of 1 month by 17%, 6 months by 27% and 9 months by 27% when compared with control animals in parallel with a reduced expression of GS (determined by Western blots, which started at the age of 6 months and was sustained up to 12 months of age. We did not, however, find any changes in the expression of GLT-1, which implies an intact glutamate uptake mechanism. Our results indicate that the decrease in GS expression may underlie a gradual decline in the vital astrocyte-dependent glutamate–glutamine conversion pathway, which in turn may compromise glutamate homoeostasis, leading towards failures in synaptic connectivity with deficient cognition and memory.

  4. Quantitative analysis of 15N labeled positional isomers of glutamine and citrulline via electrospray ionization tandem mass spectrometry of their dansyl derivatives

    The enteral metabolism of glutamine and citrulline are intertwined because, while glutamine is one of the main fuel sources for the enterocyte, citrulline is one of its products. It has been shown that the administration of 15N labeled glutamine results in the incorporation of the 15N label into cit...

  5. The effect of glutamine-enriched enteral nutrition on intestinal permeability in very-low-birth-weight infants : A randomized controlled trial

    van den Berg, Anemone; Fetter, Willem P. F.; Westerbeek, Elisabeth A. M.; van der Vegt, Ina M.; van der Molen, Hilda R. A.; van Elburg, Ruurd M.

    2006-01-01

    Background: Very-low-birth-weight (VLBW) infants are susceptible to glutamine depletion. Glutamine depletion has negative effects on intestinal integrity. The lower infection rate in VLBW infants receiving glutamine-enriched enteral nutrition may originate from improved intestinal integrity, as

  6. The effect of glutamine-enriched enteral nutrition on intestinal permeability in very-low-birth-weight infants: A randomized controlled trial

    van den Berg, Anemone; Fetter, Willem P. F.; Westerbeek, Elisabeth A. M.; van der Vegt, Ina M.; van der Molen, Hilda R. A.; van Elburg, Ruurd M.

    2006-01-01

    Background: Very-low-birth-weight (VLBW) infants are susceptible to glutamine depletion. Glutamine depletion has negative effects on intestinal integrity. The lower infection rate in VLBW infants receiving glutamine-enriched enteral nutrition may originate from improved intestinal integrity, as

  7. Using tunable diode laser spectroscopy to measure carbon isotope discrimination and mesophyll conductance to CO₂ diffusion dynamically at different CO₂ concentrations.

    Tazoe, Youshi; VON Caemmerer, Susanne; Estavillo, Gonzalo M; Evans, John R

    2011-04-01

    In C₃ leaves, the mesophyll conductance to CO₂ diffusion, g(m) , determines the drawdown in CO₂ concentration from intercellular airspace to the chloroplast stroma. Both g(m) and stomatal conductance limit photosynthetic rate and vary in response to the environment. We investigated the response of g(m) to changes in CO₂ in two Arabidopsis genotypes (including a mutant with open stomata, ost1), tobacco and wheat. We combined measurements of gas exchange with carbon isotope discrimination using tunable diode laser absorption spectroscopy with a CO₂ calibration system specially designed for a range of CO₂ and O₂ concentrations. CO₂ was initially increased from 200 to 1000 ppm and then decreased stepwise to 200 ppm and increased stepwise back to 1000 ppm, or the sequence was reversed. In 2% O₂ a step increase from 200 to 1000 ppm significantly decreased g(m) by 26-40% in all three species, whereas following a step decrease from 1000 to 200 ppm, the 26-38% increase in g(m) was not statistically significant. The response of g(m) to CO₂ was less in 21% O₂. Comparing wild type against the ost1 revealed that mesophyll and stomatal conductance varied independently in response to CO₂. We discuss the effects of isotope fractionation factors on estimating g(m) . © 2011 Blackwell Publishing Ltd.

  8. A novel glutamine-rich putative transcriptional adaptor protein (TIG-1), preferentially expressed in placental and bone-marrow tissues.

    Abraham, S; Solomon, W B

    2000-09-19

    We used a subtractive hybridization protocol to identify novel expressed sequence tags (ESTs) corresponding to mRNAs whose expression was induced upon exposure of the human leukemia cell line K562 to the phorbol ester 12-O-tetradecanolyphorbol-13-acetate (TPA). The complete open reading frame of one of the novel ESTs, named TIG-1, was obtained by screening K562 cell and placental cDNA libraries. The deduced open reading frame of the TIG-1 cDNA encodes for a glutamine repeat-rich protein with a predicted molecular weight of 63kDa. The predicted open reading frame also contains a consensus bipartite nuclear localization signal, though no specific DNA-binding domain is found. The corresponding TIG-1 mRNA is ubiquitously expressed. Placental tissue expresses the TIG-1 mRNA 200 times more than the lowest expressing tissues such as kidney and lung. There is also preferential TIG-1 mRNA expression in cells of bone-marrow lineage.In-vitro transcription/translation of the TIG-1 cDNA yielded a polypeptide with an apparent molecular weight of 97kDa. Using polyclonal antibodies obtained from a rabbit immunized with the carboxy-terminal portion of bacterially expressed TIG-1 protein, a polypeptide with molecular weight of 97kDa was identified by Western blot analyses of protein lysates obtained from K562 cells. Cotransfection assays of K562 cells, using a GAL4-TIG-1 fusion gene and GAL4 operator-CAT, indicate that the TIG-1 protein may have transcriptional regulatory activity when tethered to DNA. We hypothesize that this novel glutamine-rich protein participates in a protein complex that regulates gene transcription. It has been demonstrated by Naar et al. (Naar, A.M., Beaurang, P.A., Zhou, S., Abraham, S., Solomon, W.B., Tjian, R., 1999, Composite co-activator ARC mediates chromatin-directed transcriptional activation. Nature 398, 828-830) that the amino acid sequences of peptide fragments obtained from a polypeptide found in a complex of proteins that alters chromatin

  9. Reaction Mechanism of Mycobacterium Tuberculosis Glutamine Synthetase Using Quantum Mechanics/Molecular Mechanics Calculations.

    Moreira, Cátia; Ramos, Maria J; Fernandes, Pedro Alexandrino

    2016-06-27

    This paper is devoted to the understanding of the reaction mechanism of mycobacterium tuberculosis glutamine synthetase (mtGS) with atomic detail, using computational quantum mechanics/molecular mechanics (QM/MM) methods at the ONIOM M06-D3/6-311++G(2d,2p):ff99SB//B3LYP/6-31G(d):ff99SB level of theory. The complete reaction undergoes a three-step mechanism: the spontaneous transfer of phosphate from ATP to glutamate upon ammonium binding (ammonium quickly loses a proton to Asp54), the attack of ammonia on phosphorylated glutamate (yielding protonated glutamine), and the deprotonation of glutamine by the leaving phosphate. This exothermic reaction has an activation free energy of 21.5 kcal mol(-1) , which is consistent with that described for Escherichia coli glutamine synthetase (15-17 kcal mol(-1) ). The participating active site residues have been identified and their role and energy contributions clarified. This study provides an insightful atomic description of the biosynthetic reaction that takes place in this enzyme, opening doors for more accurate studies for developing new anti-tuberculosis therapies. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  10. Glutamine-enriched enteral diet increases splanchnic blood flow in the rat

    Houdijk, A. P.; van Leeuwen, P. A.; Boermeester, M. A.; van Lambalgen, T.; Teerlink, T.; FLINKERBUSCH, E. L.; Sauerwein, H. P.; Wesdorp, R. I.

    1994-01-01

    The hemodynamic consequences of glutamine (Gln)-enriched nutrition have not been investigated. This study investigates the effects of a Gln-enriched enteral diet on organ blood flows and systemic hemodynamics. Male Fischer 344 rats (n = 24) were randomized to a group that received a 12.5% (wt/wt)

  11. Combined Effects of Tauroursodeoxycholic Acid and Glutamine on Bacterial Translocation in Obstructive Jaundiced Rats

    Ahmet Rahmi Hatipoğlu

    2013-12-01

    Full Text Available Background: Bacterial Translocation is believed to be an important factor on mortality and morbidity in Obstructive Jaundiced. Aims: We investigated the probable or estimated positive effects of tauroursodeoxycholic acid, which has antibacterial and regulatory effects on intestinal flora, together with glutamine on BT in an experimental obstructive jaundiced rat model. Study Design: Animal experimentation. Methods: Forty adult, male, Sprague Dawley rats were used in this study. Animals were randomised and divided into five groups of eight each: sham (Sh; control (common bile duct ligation, CBDL; and supplementation groups administered tauroursodeoxycholic acid (CBDL+T, glutamine (CBDL+G, or tauroursodeoxycholic acid plus glutamine (CBDL+TG. Blood and liver, spleen, MLN, and ileal samples were taken via laparotomy under sterile conditions for investigation of bacterial translocation and intestinal mucosal integrity and hepatic function tests on the tenth postoperative day. Results: There were statistically significant differences in BT rates in all samples except the spleen of the CBDL+TG group compared with the CBDL group (p=0.041, p=0.026, and p=0.041, respectively. Conclusion: It is essential to protect hepatic functions besides maintaining intestinal mucosal integrity in the active struggle against BT occurring in obstructive jaundice. The positive effect on intestinal mucosal integrity can be increased if glutamine is used with tauroursodeoxycholic acid, which also has hepatoprotective and immunomodulatory features.

  12. Changes in Activities of Glutamine Synthetase during Grain Filling and Their Relation to Rice Quality

    Zheng-xun JIN

    2007-09-01

    Full Text Available Four japonica rice varieties differed in cooking and eating qualities were used in a pot experiment to study the relationship between the activities of glutamine synthetase during grain filling and rice quality. The activities of glutamine synthetase gradually increased and then declined as a single peak curve in the course of grain filling. The 15th day after heading was a turning point, before which the enzymatic activities in the inferior rice varieties with high protein content were higher than those in the superior rice varietie with low protein content, and after which it was converse. The activity of glutamine synthetase in grain was correlated with the taste meter value, peak viscosity and breakdown negatively at the early stage of grain filling whereas positively at the middle and late stages. Moreover, it was correlated with the protein content of rice grain and setback positively at the early stage and negatively at the middle and late stages. The correlation degree varied with the course of grain filling. From 15 days to 20 days after heading was a critical stage, in which the direction of correlation between the activity of glutamine synthetase and taste meter value and RVA properties of rice changed.

  13. Assessing the extent of bone degradation using glutamine deamidation in collagen.

    Wilson, Julie; van Doorn, Nienke L; Collins, Matthew J

    2012-11-06

    Collagen peptides are analyzed using a low-cost, high-throughput method for assessing deamidation using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS). For each chosen peptide, the theoretical distribution is calculated and the measured distribution for each sample compared with this to determine the extent of glutamine deamidation. The deamidation of glutamine (Q) to glutamic acid (E) results in a mass shift of +0.984 Da. Thus, from the resolution of our data, the second peak in the isotope distribution for a peptide containing one glutamine residue coincides with the first peak of the isotope distribution for the peptide in which the residue is deamidated. A genetic algorithm is used to determine the extent of deamidation that gives the best fit to the measured distribution. The method can be extended to peptides containing more than one glutamine residue. The extent of protein degradation assessed in this way could be used, for example, to assess the damage of collagen, and screen samples for radiocarbon dating and DNA analysis.

  14. L-Glutamine Metabolism Is Not A Major Source Of Increased Free ...

    10 weeks (2K-1C) and 4 weeks (1K-1C) respectively after renal artery clamping, clipped rats exhibited elevated blood pressures (P<0.001), which was sustained under anaesthesia. No significant difference in plasma glutamine levels were found in hypertensive rats compared to controls (11.3±1.3 mg/l in 2K-1C vs.

  15. Brain MRS glutamine as a biomarker to guide therapy of hyperammonemic coma.

    O'Donnell-Luria, Anne H; Lin, Alexander P; Merugumala, Sai K; Rohr, Frances; Waisbren, Susan E; Lynch, Rebecca; Tchekmedyian, Vatche; Goldberg, Aaron D; Bellinger, Andrew; McFaline-Figueroa, J Ricardo; Simon, Tracey; Gershanik, Esteban F; Levy, Bruce D; Cohen, David E; Samuels, Martin A; Berry, Gerard T; Frank, Natasha Y

    2017-05-01

    Acute idiopathic hyperammonemia in an adult patient is a life-threatening condition often resulting in a rapid progression to irreversible cerebral edema and death. While ammonia-scavenging therapies lower blood ammonia levels, in comparison, clearance of waste nitrogen from the brain may be delayed. Therefore, we used magnetic resonance spectroscopy (MRS) to monitor cerebral glutamine levels, the major reservoir of ammonia, in a gastric bypass patient with hyperammonemic coma undergoing therapy with N-carbamoyl glutamate and the ammonia-scavenging agents, sodium phenylacetate and sodium benzoate. Improvement in mental status mirrored brain glutamine levels, as coma persisted for 48h after plasma ammonia normalized. We hypothesize that the slower clearance for brain glutamine levels accounts for the delay in improvement following initiation of treatment in cases of chronic hyperammonemia. We propose MRS to monitor brain glutamine as a noninvasive approach to be utilized for diagnostic and therapeutic monitoring purposes in adult patients presenting with idiopathic hyperammonemia. Copyright © 2017 Elsevier Inc. All rights reserved.

  16. Expression of glutamine transporter isoforms in cerebral cortex of rats with chronic hepatic encephalopathy

    Leke, Renata; Escobar, Thayssa D.C.; Rama Rao, Kakulavarapu V.

    2015-01-01

    Hepatic encephalopathy (HE) is a neuropsychiatric disorder that occurs due to acute and chronic liver diseases, the hallmark of which is the increased levels of ammonia and subsequent alterations in glutamine synthesis, i.e. conditions associated with the pathophysiology of HE. Under physiological...

  17. Glutamine-enriched enteral nutrition in very low-birth-weight infants

    van den Berg, Anemone; van Zwol, Annelies; Moll, Henriëtte A.; Fetter, Willem P. F.; van Elburg, Ruurd M.

    2007-01-01

    Objective: To determine the effect of glutamine-enriched enteral nutrition in very low- birth- weight infants on the incidence of allergic and infectious diseases during the first year of life. Design: Follow- up study. Setting: Tertiary care hospital. Participants: All surviving infants who

  18. Deamidation of asparagine and glutamine residues in proteins and peptides: structural determinants and analytical methodology

    Bischoff, Rainer; Kolbe, H.V.

    1994-01-01

    Non-enzymatic deamidation of asparagine and glutamine residues in proteins and peptides are reviewed by first outlining the well-described reaction mechanism involving cyclic imide intermediates, followed by a discussion of structural features which influence the reaction rate. The second and major

  19. Deamidation Reactions of Asparagine- and Glutamine-Containing Dipeptides Investigated by Ion Spectroscopy

    Kempkes, L.J.M.; Martens, J.; Grzetic, J.; Berden, G.; Oomens, J.

    2016-01-01

    Deamidation is a major fragmentation channel upon activation by collision induced dissociation (CID) for protonated peptides containing glutamine (Gln) and asparagine (Asn) residues. Here, we investigate these NH3-loss reactions for four Asn- and Gln-containing protonated peptides in terms of the

  20. Noncoding RNA of Glutamine Synthetase I Modulates Antibiotic Production in Streptomyces coelicolor A3(2)

    D'Alia, Davide; Nieselt, Kay; Steigele, Stephan; Mueller, Jonas; Verburg, Ilse; Takano, Eriko; Alia, Davide D’; Müller, Jonas

    Overexpression of antisense chromosomal cis-encoded noncoding RNAss (ncRNAs) in glutamine synthetase I resulted in a decrease in growth, protein synthesis, and antibiotic production in Streptomyces coelicolor. In addition, we predicted 3,597 cis-encoded ncRNAs and validated 13 of them

  1. Evidence for Tautomerisation of Glutamine in BLUF Blue Light Receptors by Vibrational Spectroscopy and Computational Chemistry

    Domratcheva, Tatiana; Hartmann, Elisabeth; Schlichting, Ilme; Kottke, Tilman

    2016-01-01

    BLUF (blue light sensor using flavin) domains regulate the activity of various enzymatic effector domains in bacteria and euglenids. BLUF features a unique photoactivation through restructuring of the hydrogen-bonding network as opposed to a redox reaction or an isomerization of the chromophore. A conserved glutamine residue close to the flavin chromophore plays a central role in the light response, but the underlying modification is still unclear. We labelled this glutamine with 15N in two representative BLUF domains and performed time-resolved infrared double difference spectroscopy. The assignment of the signals was conducted by extensive quantum chemical calculations on large models with 187 atoms reproducing the UV-vis and infrared signatures of BLUF photoactivation. In the dark state, the comparatively low frequency of 1,667 cm−1 is assigned to the glutamine C=O accepting a hydrogen bond from tyrosine. In the light state, the signature of a tautomerised glutamine was extracted with the C=N stretch at ~1,691 cm−1 exhibiting the characteristic strong downshift by 15N labelling. Moreover, an indirect isotope effect on the flavin C4=O stretch was found. We conclude that photoactivation of the BLUF receptor does not only involve a rearrangement of hydrogen bonds but includes a change in covalent bonds of the protein. PMID:26947391

  2. Efeito da suplementação com L-alanil-L-glutamina sobre a resposta de hipersensibilidade do tipo tardio em ratos submetidos ao treinamento intenso Effect of L-glutamine and L-alanyl-L-glutamine supplementation on the response to delayed-type hypersensitivity test (DTH in rats submitted to intense training

    Marcelo Macedo Rogero

    2002-12-01

    Full Text Available O treinamento intenso e o exercício exaustivo podem ocasionar imunossupressão em atletas por meio da diminuição da concentração plasmática de glutamina. O presente estudo verificou inicialmente o efeito da suplementação com L-glutamina e L-alanilL-glutamina sobre a resposta ao teste de hipersensibilidade do tipo tardio (HTT em ratos submetidos ao treinamento intenso em natação durante seis semanas. Posteriormente, foi avaliado o efeito dessas intervenções nutricionais sobre a contagem total e porcentual de leucócitos e concentração sérica de anticorpos IgG anti-albumina de soro bovino, em animais submetidos ao teste de exaustão e recuperados durante o período de 3 horas. Não houve efeito do treinamento e da suplementação sobre a resposta ao teste de HTT. Animais suplementados apresentaram maior concentração de glutamina no plasma (PIntense training and exhaustive exercise may cause immunesupression in athletes by reducing plasma glutamine concentration. Initially, this study verified the effect of L-glutamine and L-alanyl-L-glutamine supplementation on the response to delayed-type hypersensitivity test (DTH in rats submitted to intense swimming training for six weeks. Later on, we assessed the effect of these nutritional interventions on total and differential white blood cell counts and on concentration of anti-bovine serum albumin IgG antibodies, in animals submitted to exhaustion test and a three-hour recovery period. There was no effect of training and supplementation on the response to DTH. Supplemented animals presented greatest plasma glutamine concentration (p<0.05, though this increase in glutaminemia did not interfere on the serum IgG antibody concentration. The recovery period after intense exercise resulted in decreased glutaminemia as compared with the values obtained immediately after exhaustion test (p<0.05. Increase in corticosterone levels induced by strenuous exercises led to leukocytosis, neutrophilia

  3. Alanyl-glutamine attenuates 5-fluorouracil-induced intestinal mucositis in apolipoprotein E-deficient mice

    C.V. Araújo

    2015-06-01

    Full Text Available Apolipoprotein E (APOE=gene, apoE=protein is a known factor regulating the inflammatory response that may have regenerative effects during tissue recovery from injury. We investigated whether apoE deficiency reduces the healing effect of alanyl-glutamine (Ala-Gln treatment, a recognized gut-trophic nutrient, during tissue recovery after 5-FU-induced intestinal mucositis. APOE-knockout (APOE-/- and wild-type (APOE+/+ C57BL6J male and female mice (N=86 were given either Ala-Gln (100 mM or phosphate buffered saline (PBS by gavage 3 days before and 5 days after a 5-fluorouracil (5-FU challenge (450 mg/kg, via intraperitoneal injection. Mouse body weight was monitored daily. The 5-FU cytotoxic effect was evaluated by leukometry. Intestinal villus height, villus/crypt ratio, and villin expression were monitored to assess recovery of the intestinal absorptive surface area. Crypt length, mitotic, apoptotic, and necrotic crypt indexes, and quantitative real-time PCR for insulin-like growth factor-1 (IGF-1 and B-cell lymphoma 2 (Bcl-2 intestinal mRNA transcripts were used to evaluate intestinal epithelial cell turnover. 5-FU challenge caused significant weight loss and leukopenia (P<0.001 in both mouse strains, which was not improved by Ala-Gln. Villus blunting, crypt hyperplasia, and reduced villus/crypt ratio (P<0.05 were found in all 5-FU-challenged mice but not in PBS controls. Ala-Gln improved villus/crypt ratio, crypt length and mitotic index in all challenged mice, compared with PBS controls. Ala-Gln improved villus height only in APOE-/- mice. Crypt cell apoptosis and necrotic scores were increased in all mice challenged by 5-FU, compared with untreated controls. Those scores were significantly lower in Ala-Gln-treated APOE+/+ mice than in controls. Bcl-2 and IGF-1 mRNA transcripts were reduced only in the APOE-/--challenged mice. Altogether our findings suggest APOE-independent Ala-Gln regenerative effects after 5-FU challenge.

  4. Alanyl-glutamine attenuates 5-fluorouracil-induced intestinal mucositis in apolipoprotein E-deficient mice

    Araújo, C.V.; Lazzarotto, C.R.; Aquino, C.C.; Figueiredo, I.L.; Costa, T.B.; Oliveira Alves, L.A. de; Ribeiro, R.A.; Bertolini, L.R.; Lima, A.A.M.; Brito, G.A.C.; Oriá, R.B.

    2015-01-01

    Apolipoprotein E (APOE=gene, apoE=protein) is a known factor regulating the inflammatory response that may have regenerative effects during tissue recovery from injury. We investigated whether apoE deficiency reduces the healing effect of alanyl-glutamine (Ala-Gln) treatment, a recognized gut-trophic nutrient, during tissue recovery after 5-FU-induced intestinal mucositis. APOE-knockout (APOE -/- ) and wild-type (APOE +/+ ) C57BL6J male and female mice (N=86) were given either Ala-Gln (100 mM) or phosphate buffered saline (PBS) by gavage 3 days before and 5 days after a 5-fluorouracil (5-FU) challenge (450 mg/kg, via intraperitoneal injection). Mouse body weight was monitored daily. The 5-FU cytotoxic effect was evaluated by leukometry. Intestinal villus height, villus/crypt ratio, and villin expression were monitored to assess recovery of the intestinal absorptive surface area. Crypt length, mitotic, apoptotic, and necrotic crypt indexes, and quantitative real-time PCR for insulin-like growth factor-1 (IGF-1) and B-cell lymphoma 2 (Bcl-2) intestinal mRNA transcripts were used to evaluate intestinal epithelial cell turnover. 5-FU challenge caused significant weight loss and leukopenia (P<0.001) in both mouse strains, which was not improved by Ala-Gln. Villus blunting, crypt hyperplasia, and reduced villus/crypt ratio (P<0.05) were found in all 5-FU-challenged mice but not in PBS controls. Ala-Gln improved villus/crypt ratio, crypt length and mitotic index in all challenged mice, compared with PBS controls. Ala-Gln improved villus height only in APOE -/- mice. Crypt cell apoptosis and necrotic scores were increased in all mice challenged by 5-FU, compared with untreated controls. Those scores were significantly lower in Ala-Gln-treated APOE +/+ mice than in controls. Bcl-2 and IGF-1 mRNA transcripts were reduced only in the APOE -/- -challenged mice. Altogether our findings suggest APOE-independent Ala-Gln regenerative effects after 5-FU challenge

  5. Alanyl-glutamine attenuates 5-fluorouracil-induced intestinal mucositis in apolipoprotein E-deficient mice

    Araújo, C.V. [Laboratório da Biologia da Cicatrização, Ontogenia e Nutrição de Tecidos, INCT - Instituto de Biomedicina do Semiárido Brasileiro, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil); Lazzarotto, C.R. [Laboratório de Biologia Molecular e do Desenvolvimento, Universidade de Fortaleza, Fortaleza, CE (Brazil); Aquino, C.C.; Figueiredo, I.L.; Costa, T.B.; Oliveira Alves, L.A. de [Laboratório da Biologia da Cicatrização, Ontogenia e Nutrição de Tecidos, INCT - Instituto de Biomedicina do Semiárido Brasileiro, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil); Ribeiro, R.A. [Laboratório da Inflamação e Câncer, INCT - Instituto de Biomedicina do Semiárido Brasileiro, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil); Bertolini, L.R. [Laboratório de Biologia Molecular e do Desenvolvimento, Universidade de Fortaleza, Fortaleza, CE (Brazil); Lima, A.A.M. [Laboratório de Doenças Infecciosas, INCT - Instituto de Biomedicina do Semiárido Brasileiro, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil); Brito, G.A.C. [Laboratório da Inflamação e Câncer, INCT - Instituto de Biomedicina do Semiárido Brasileiro, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil); Oriá, R.B. [Laboratório da Biologia da Cicatrização, Ontogenia e Nutrição de Tecidos, INCT - Instituto de Biomedicina do Semiárido Brasileiro, Faculdade de Medicina, Universidade Federal do Ceará, Fortaleza, CE (Brazil)

    2015-04-28

    Apolipoprotein E (APOE=gene, apoE=protein) is a known factor regulating the inflammatory response that may have regenerative effects during tissue recovery from injury. We investigated whether apoE deficiency reduces the healing effect of alanyl-glutamine (Ala-Gln) treatment, a recognized gut-trophic nutrient, during tissue recovery after 5-FU-induced intestinal mucositis. APOE-knockout (APOE{sup -/-}) and wild-type (APOE{sup +/+}) C57BL6J male and female mice (N=86) were given either Ala-Gln (100 mM) or phosphate buffered saline (PBS) by gavage 3 days before and 5 days after a 5-fluorouracil (5-FU) challenge (450 mg/kg, via intraperitoneal injection). Mouse body weight was monitored daily. The 5-FU cytotoxic effect was evaluated by leukometry. Intestinal villus height, villus/crypt ratio, and villin expression were monitored to assess recovery of the intestinal absorptive surface area. Crypt length, mitotic, apoptotic, and necrotic crypt indexes, and quantitative real-time PCR for insulin-like growth factor-1 (IGF-1) and B-cell lymphoma 2 (Bcl-2) intestinal mRNA transcripts were used to evaluate intestinal epithelial cell turnover. 5-FU challenge caused significant weight loss and leukopenia (P<0.001) in both mouse strains, which was not improved by Ala-Gln. Villus blunting, crypt hyperplasia, and reduced villus/crypt ratio (P<0.05) were found in all 5-FU-challenged mice but not in PBS controls. Ala-Gln improved villus/crypt ratio, crypt length and mitotic index in all challenged mice, compared with PBS controls. Ala-Gln improved villus height only in APOE{sup -/-} mice. Crypt cell apoptosis and necrotic scores were increased in all mice challenged by 5-FU, compared with untreated controls. Those scores were significantly lower in Ala-Gln-treated APOE{sup +/+} mice than in controls. Bcl-2 and IGF-1 mRNA transcripts were reduced only in the APOE{sup -/-}-challenged mice. Altogether our findings suggest APOE-independent Ala-Gln regenerative effects after 5-FU

  6. Arginine Deprivation Inhibits the Warburg Effect and Upregulates Glutamine Anaplerosis and Serine Biosynthesis in ASS1-Deficient Cancers

    Jeff Charles Kremer

    2017-01-01

    Full Text Available Targeting defects in metabolism is an underutilized strategy for the treatment of cancer. Arginine auxotrophy resulting from the silencing of argininosuccinate synthetase 1 (ASS1 is a common metabolic alteration reported in a broad range of aggressive cancers. To assess the metabolic effects that arise from acute and chronic arginine starvation in ASS1-deficient cell lines, we performed metabolite profiling. We found that pharmacologically induced arginine depletion causes increased serine biosynthesis, glutamine anaplerosis, oxidative phosphorylation, and decreased aerobic glycolysis, effectively inhibiting the Warburg effect. The reduction of glycolysis in cells otherwise dependent on aerobic glycolysis is correlated with reduced PKM2 expression and phosphorylation and upregulation of PHGDH. Concurrent arginine deprivation and glutaminase inhibition was found to be synthetic lethal across a spectrum of ASS1-deficient tumor cell lines and is sufficient to cause in vivo tumor regression in mice. These results identify two synthetic lethal therapeutic strategies exploiting metabolic vulnerabilities of ASS1-negative cancers.

  7. Impaired Hippocampal Glutamate and Glutamine Metabolism in the db/db Mouse Model of Type 2 Diabetes Mellitus

    Andersen, Jens Velde; Nissen, Jakob Dahl; Christensen, Sofie Kjellerup

    2017-01-01

    Type 2 diabetes mellitus (T2DM) is a risk factor for the development of Alzheimer's disease, and changes in brain energy metabolism have been suggested as a causative mechanism. The aim of this study was to investigate the cerebral metabolism of the important amino acids glutamate and glutamine...... significantly reduced 13C labeling in glutamate, glutamine, GABA, citrate, and aspartate from metabolism of [U-13C]glutamate. Additionally, reduced 13C labeling were observed in GABA, citrate, and aspartate from [U-13C]glutamine metabolism in hippocampal slices of db/db mice when compared to controls. None...

  8. Sleep Deprivation in Humans, Immunodepression and Glutamine Supplementation

    Castell, Linda M; Gough, Elizabeth; Cardenas, Rebecca; Miller, James C

    2005-01-01

    ... (I) Are the cytokines linked with eosinophils neutrophils and lymphocytes cell types which are known to be affected by sleep deprivation changed in terms of intracellular cytokine production? (2...

  9. Changes in photosynthesis, mesophyll conductance to CO{sub 2}, and isoprenoid emissions in Populus nigra plants exposed to excess nickel

    Velikova, Violeta, E-mail: violet@obzor.bio21.bas.bg [Bulgarian Academy of Sciences, Acad. M. Popov Institute of Plant Physiology, Acad. G. Bonchev, Bl. 21, 1113 Sofia (Bulgaria); Tsonev, Tsonko [Bulgarian Academy of Sciences, Acad. M. Popov Institute of Plant Physiology, Acad. G. Bonchev, Bl. 21, 1113 Sofia (Bulgaria); Loreto, Francesco [Consiglio Nazionale delle Ricerche, Istituto per la Protezione delle Piante, Via Madonna del Piano 10, 50019 Sesto Fiorentino, Firenze (Italy); Centritto, Mauro [Consiglio Nazionale delle Ricerche, Istituto di Biologia Agroambientale e Forestale, 00015 Monterotondo Scalo (RM) (Italy)

    2011-05-15

    Poplar (Populus nigra) plants were grown hydroponically with 30 and 200 {mu}M Ni (Ni{sub 30} and Ni{sub 200}). Photosynthesis limitations and isoprenoid emissions were investigated in two leaf types (mature and developing). Ni stress significantly decreased photosynthesis, and this effect depended on the leaf Ni content, which was lower in mature than in developing leaves. The main limitations to photosynthesis were attributed to mesophyll conductance and metabolism impairment. In Ni-stressed developing leaves, isoprene emission was significantly stimulated. We attribute such stimulation to the lower chloroplastic [CO{sub 2}] than in control leaves. However chloroplastic [CO{sub 2}] did not control isoprene emission in mature leaves. Ni stress induced the emission of cis-{beta}-ocimene in mature leaves, and of linalool in both leaf types. Induced biosynthesis and emission of isoprenoids reveal the onset of antioxidant processes that may also contribute to reduce Ni stress, especially in mature poplar leaves. - Graphical abstract: Visible damage caused by Ni treatment. 1 - Ni{sub 0} (control plants); 2 - Ni{sub 200}; M = mature and D = developing Populus nigra leaves. Display Omitted Highlights: > We study the effect of Ni pollution on photosynthesis and isoprenoid emissions. > Ni stress significantly decreases photosynthesis. The main limitations are attributed to mesophyll conductance and metabolism impairment. > Constitutive isoprene emission was significantly stimulated in Ni-stressed leaves. Exposure to enhanced Ni concentration induces cis-beta-ocimene and linalool emissions. - The study reveals consequences of Ni stress on plant physiology, namely increasing diffusional limitation to photosynthesis and isoprenoid emissions.

  10. Effects of short-term dietary restriction and glutamine supplementation in vitro on the modulation of inflammatory properties.

    C de Oliveira, Dalila; Santos, Ed Wilson; Nogueira-Pedro, Amanda; Xavier, José Guilherme; Borelli, Primavera; Fock, Ricardo Ambrósio

    2018-04-01

    Dietary restriction (DR) is a nutritional intervention that exerts profound effects on biochemical and immunologic parameters, modulating some inflammatory properties. Glutamine (GLN) is a conditionally essential amino acid that can modulate inflammatory properties. However, there is a lack of data evaluating the effects of DR and GLN supplementation, especially in relation to inflammatory cytokine production and the expression of transcription factors such as nuclear factor (NF)-κB. We subjected 3-mo-old male Balb/c mice to DR by reducing their food intake by 30%. DR animals lost weight and showed reduced levels of serum triacylglycerols, glucose, cholesterol, and calcium as well as a reduction in bone density. Additionally, blood, peritoneal, and spleen cellularity were reduced, lowering the number of peritoneal F4/80- and CD86-positive cells and the total number of splenic CD4- and CD8-positive cells. The production of interleukin (IL)-10 and the expression of NF-κB in splenic cells were not affected by DR or by GLN supplementation. However, peritoneal macrophages from DR animals showed reduced IL-12 and tumor necrosis factor-α production and increased IL-10 production with reduced phosphorylation of NF-κB expression. Additionally, GLN was able to modulate cytokine production by peritoneal cells from the control group, although no effects were observed in cells from the DR group. DR induces biochemical and immunologic changes, in particular by reducing IL-12 and tumor necrosis factor-α production by macrophages and clearly upregulating IL-10 production, whereas GLN supplementation did not modify these parameters in cells from DR animals. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Interaction of PHM, PHI and 24-glutamine PHI with human VIP receptors from colonic epithelium: comparison with rat intestinal receptors

    Laburthe, M.; Couvineau, A.; Rouyer-Fessard, C.; Moroder, L.

    1985-01-01

    PHM, the human counterpart of porcine Peptide Histidine Isoleucine amide (PHI), is shown to be a VIP agonist with low potency on human VIP receptors located in colonic epithelial cell membranes. Its potency is identical to that of PHI but by 3 orders of magnitude lower than that of VIP itself in inhibiting 125 I-VIP binding and in stimulating adenylate cyclase activity. This contrasts markedly with the behavior of PHI on rat VIP receptors located in intestinal epithelial cell membranes where PHI is a potent agonist with a potency that is 1/5 that of VIP. In another connection, the authors show that 24-glutamine PHI has the same affinity as 24-glutamic acid PHI (the natural peptide) for rat or human VIP receptors. These results indicate that while PHI may exert some physiological function through its interaction with VIP receptors in rodents, its human counterpart PHM is a very poor agonist of VIP in human. Furthermore, they show that the drastic change in position 24 of PHI (neutral versus acid residue) does not affect the activity of PHI, at least on VIP receptors. 21 references, 1 figure

  12. Effect of Nutrient Dilution and Glutamine Supplementation on Growth Performance, Small Intestine Morphology and Immune Response of Broilers

    majid gheshlagh olyayee

    2016-11-01

    Full Text Available Introduction Glutamine (Gln, a semi-essential or conditionally essential amino acid, is an abundant amino acid in plasma and skeletal muscle. It is the main energy substrate for cells that undergo intense replication, such as enterocytes, lymphocytes, macrophages, neutrophils and kidney cells and plays an important role in their function and homeostasis. Apart from providing nitrogen for protein synthesis, Gln is a precursor for nucleic acids, nucleotides, hexose amines, the nitric oxide precursor arginine (Arg, and the major antioxidant-glutathione. It plays a central role in nitrogen transport between tissues, specifically from muscle to gut, kidney, and liver. In addition to its role as a gluconeogenic substrate in the liver, kidney, and intestine, Gln is involved in the renal handling of ammonia, serving as a regulator of acid base homeostasis. So the aim of this study was to evaluate the effect of nutrient dilution and L- glutamine (Gln supplementation on growth performance, intestine morphology and immune response of broilers during starter (0 to 10 days, growth (11 to 24 days and finisher (25 to 42 days periods. Materials and methods A total of 320 one-day-old male Ross 308 broiler chicks were randomly assigned to eight treatments with 4 replicates and 10 chicks per each. In this study two levels of nutrient dilution (Ross 308 broiler nutrition recommendation and 5% diluted and 4 levels of Gln supplementation (0, 0.5, 1 and 1.5% were used in a completely randomized design as factorial arrangement 2×4. Growth performance was measured periodically. In order to investigate jejenual histomorphology such as villus height, depth of crypt, villus height to depth of crypt ratio, villus width, muscle layer thickness and epithelium thickness, on day 42 after 4 h fasting, one bird per each replicate was randomly selected, slaughtered and 1 cm of middle section of jejenum was cut. Cellular immune response was assessed in 40-d-old chick using the in

  13. Effects of adding different levels of Glutamine to modified Beltsville extender on the survival of frozen rooster semen.

    Khiabani, Aytak Bakhshayesh; Moghaddam, Gholamali; Kia, Hossein Daghigh

    2017-09-01

    The aim of the present study was to investigate the effects of l-glutamine on the quality of frozen-thawed rooster semen. Semen samples were collected from eight mature roosters (Ross 308). After initial semen assessments, samples of adequate quality were mixed together and diluted with modified Beltsville extender without l-glutamine (control) and supplemented with 2.5, 5, and 7.5mM l-glutamine. Semen straws were subjected to cryopreservation and evaluated twice at 15-day intervals. After thawing, sperm viability, total and progressive sperm motilities were measured by Eosin-Nigrosine and Computer-Aided Sperm Analysis (CASA), respectively. The results showed that sperm functions decreased on day 30 compared to day 15. The extender supplemented with 5mM glutamine improved (prooster sperm to be frozen for longer. Copyright © 2017 Elsevier B.V. All rights reserved.

  14. Exogenous glutamine increases lipid accumulation in developing seeds of castor bean (Ricinus communis L. cultured in vitro

    Zhang Yang

    2015-01-01

    Full Text Available This report describes biomass production and compositional changes of developing castor seeds in response to change in the nitrogen resource (glutamine of the medium. During the early developmental period (24-36 days after pollination, oil was found to initially accumulate in the developing seeds. Carbohydrates and oil were inversely related after glutamine provision (35 mM, in the culture medium. [U-14C] sucrose labeling was used to investigate the effect of metabolic fluxes among different storage materials. Addition of glutamine led to a 7% increase of labeling in lipids and an inverse decrease of labeling in carbohydrates. It was postulated that changes in the glutamine concentration in the medium are likely to influence the partitioning of resources between the various storage products, especially carbohydrates and oil. These observations will contribute to a better understanding of assimilate partitioning in developing castor seeds and the development of molecular strategies to improve castor bean seed quality and plant breeding studies.

  15. Immune and cell modulation by amino acids.

    Roth, Erich

    2007-10-01

    Sir David Cuthbertson was the first to define metabolic alterations in post-aggression syndrome (PAS). From basic measurements of nitrogen loss and total protein synthesis/degradation, the current research has moved to genomics, proteomics and metabolomics. In this respect, first evidence was accumulated for the influence of acute catabolism, immobilisation by bed rest and sarcopenia of old age on the muscle-cell genome and proteome. Moreover, in post-aggression syndrome specific amino acids such as glutamine, arginine, glycine, taurine, tryptophan and cysteine are used for cell and immune modulation. Our laboratory has focused on the regulative capacity of glutamine. Glutamine deficiency as found in post-aggression syndrome reduces lymphocyte proliferation, alters monocyte/macrophage activity, decreases the formation of heat-shock proteins, stimulates cell apoptosis, shifts the cellular redox potential by altering the glutathione synthesis and increases the activity of the AMPK system. Investigating the molecular effect of glutamine on Hsp 70 induction, we tested the glutamine dependence on the formation of transfer-RNA and of heat-shock factor 1 (HSF 1), and on transcription and translation of Hsp 70. We could demonstrate that glutamine stabilises the mRNA of Hsp 70 thereby prolonging its half-life. The lecture also discusses the principal molecular targets of administered arginine, glycine, cysteine, taurine and tryptophan.

  16. Initiation of Swarming Motility by Proteus mirabilis Occurs in Response to Specific Cues Present in Urine and Requires Excess l-Glutamine

    Armbruster, Chelsie E.; Hodges, Steven A.

    2013-01-01

    Proteus mirabilis, a leading cause of catheter-associated urinary tract infection (CaUTI), differentiates into swarm cells that migrate across catheter surfaces and medium solidified with 1.5% agar. While many genes and nutrient requirements involved in the swarming process have been identified, few studies have addressed the signals that promote initiation of swarming following initial contact with a surface. In this study, we show that P. mirabilis CaUTI isolates initiate swarming in response to specific nutrients and environmental cues. Thirty-three compounds, including amino acids, polyamines, fatty acids, and tricarboxylic acid (TCA) cycle intermediates, were tested for the ability to promote swarming when added to normally nonpermissive media. l-Arginine, l-glutamine, dl-histidine, malate, and dl-ornithine promoted swarming on several types of media without enhancing swimming motility or growth rate. Testing of isogenic mutants revealed that swarming in response to the cues required putrescine biosynthesis and pathways involved in amino acid metabolism. Furthermore, excess glutamine was found to be a strict requirement for swarming on normal swarm agar in addition to being a swarming cue under normally nonpermissive conditions. We thus conclude that initiation of swarming occurs in response to specific cues and that manipulating concentrations of key nutrient cues can signal whether or not a particular environment is permissive for swarming. PMID:23316040

  17. Glutamina: aspectos bioquímicos, metabólicos, moleculares e suplementação Glutamine: biochemical, metabolic, molecular aspects and supplementation

    Vinicius Fernandes Cruzat

    2009-10-01

    Full Text Available A glutamina é o aminoácido livre mais abundante no plasma e no tecido muscular. Nutricionalmente é classificada como um aminoácido não essencial, uma vez que pode ser sintetizada pelo organismo a partir de outros aminoácidos. A glutamina está envolvida em diferentes funções, tais como a proliferação e desenvolvimento de células, o balanço acidobásico, o transporte da amônia entre os tecidos, a doação de esqueletos de carbono para a gliconeogênese, a participação no sistema antioxidante e outras. Por meio de técnicas de biologia molecular, estudos demonstram que a glutamina pode também influenciar diversas vias de sinalização celular, em especial a expressão de proteínas de choque térmico (HSPs. As HSPs contribuem para a manutenção da homeostasia da célula na presença de agentes estressores, tais como as espécies reativas de oxigênio (ERO. Em situações de elevado catabolismo muscular, como após exercícios físicos intensos e prolongados, a concentração de glutamina pode tornar-se reduzida. A menor disponibilidade desse aminoácido pode diminuir a resistência da célula a lesões, levando a processos de apoptose celular. Por essas razões, a suplementação com L-glutamina, tanto na forma livre, quanto como dipeptídeo, tem sido investigada. Alguns aspectos bioquímicos, metabólicos e mecanismos moleculares da glutamina, bem como os efeitos de sua suplementação, são abordados no presente trabalho.Glutamine is the most frequent free amino acid in the serum and muscular tissue. Nutritionally, it is classified as a non-essential amino acid, once it can be synthesized by the body from other amino acids. Glutamine is involved in different functions, such as cell proliferation and development, basic acid balance, ammonia transportation between tissues, carbon skeleton donation to the gluconeogenesis, participation in the antioxidant system, among others. Molecular biology techniques show that it may also

  18. Knockout of GAD65 has major impact on synaptic GABA synthesized from astrocyte-derived glutamine

    Walls, Anne Byriel; Eyjolfsson, Elvar M.; Smeland, Olav B.

    2011-01-01

    γ-Aminobutyric acid (GABA) synthesis from glutamate is catalyzed by glutamate decarboxylase (GAD) of which two isoforms, GAD65 and GAD67, have been identified. The GAD65 has repeatedly been shown to be important during intensified synaptic activity. To specifically elucidate the significance of G...... glutamine both via direct synthesis and via a pathway involving mitochondrial metabolism. Furthermore, a severe neuronal hypometabolism, involving glycolysis and tricarboxylic acid (TCA) cycle activity, was observed in cerebral cortex of GAD65 knockout mice.......65 for maintenance of the highly compartmentalized intracellular and intercellular GABA homeostasis, GAD65 knockout and corresponding wild-type mice were injected with [1-(13)C]glucose and the astrocyte-specific substrate [1,2-(13)C]acetate. Synthesis of GABA from glutamine in the GABAergic synapses...

  19. The impact of use of Glutamine on patients with head and neck tumors in radiotherapy and chemotherapy treatment

    Boligon, Caroline Schardong; Huth, Adriane

    2011-01-01

    Introduction: patients with head and neck neoplasia usually show malnutrition or a nutritional risk, because of common symptoms like: dysphagia, odynophagia and xerostomia. Objective: this study aimed to verify the impact of using amino glutamine in patients with head and neck neoplasia and under radiotherapy and chemotherapy treatment concomitantly. Methods: the research was quantitative, cross-sectional, descriptive and exploratory. The data was collected from nutritional evaluation, and patients chart consultation. The patients were divided in a control group (without use of glutamine) and a test group (with use of glutamine). 16 patients, 13 of which were men and three were women, participated in the research. Results: The control group presented mucositis grades I to IV while patients who used the amino glutamine showed mucositis grades I to II only. It could be observed that the Nutritional Risk Index decreased, which represents higher nutritional risk in patients from the control group only. In patients who used glutamine, this decrease was not significant. Conclusion: these results suggest that the use of glutamine in patients with head and neck tumors and under antineoplastic therapy helps to maintain their nutritional stage and to prevent mucositis throughout their treatment, mainly grades III and IV, which prevents adequate and regular eating and nourishment. (author)

  20. Phage display selection of efficient glutamine-donor substrate peptides for transglutaminase 2.

    Keresztessy, Zsolt; Csosz, Eva; Hársfalvi, Jolán; Csomós, Krisztián; Gray, Joe; Lightowlers, Robert N; Lakey, Jeremy H; Balajthy, Zoltán; Fésüs, László

    2006-11-01

    Understanding substrate specificity and identification of natural targets of transglutaminase 2 (TG2), the ubiquitous multifunctional cross-linking enzyme, which forms isopeptide bonds between protein-linked glutamine and lysine residues, is crucial in the elucidation of its physiological role. As a novel means of specificity analysis, we adapted the phage display technique to select glutamine-donor substrates from a random heptapeptide library via binding to recombinant TG2 and elution with a synthetic amine-donor substrate. Twenty-six Gln-containing sequences from the second and third biopanning rounds were susceptible for TG2-mediated incorporation of 5-(biotinamido)penthylamine, and the peptides GQQQTPY, GLQQASV, and WQTPMNS were modified most efficiently. A consensus around glutamines was established as pQX(P,T,S)l, which is consistent with identified substrates listed in the TRANSDAB database. Database searches showed that several proteins contain peptides similar to the phage-selected sequences, and the N-terminal glutamine-rich domain of SWI1/SNF1-related chromatin remodeling proteins was chosen for detailed analysis. MALDI/TOF and tandem mass spectrometry-based studies of a representative part of the domain, SGYGQQGQTPYYNQQSPHPQQQQPPYS (SnQ1), revealed that Q(6), Q(8), and Q(22) are modified by TG2. Kinetic parameters of SnQ1 transamidation (K(M)(app) = 250 microM, k(cat) = 18.3 sec(-1), and k(cat)/K(M)(app) = 73,200) classify it as an efficient TG2 substrate. Circular dichroism spectra indicated that SnQ1 has a random coil conformation, supporting its accessibility in the full-length parental protein. Added together, here we report a novel use of the phage display technology with great potential in transglutaminase research.

  1. The Effectiveness of Combined Use of Antioxidant and Glutamine in Abdominal Sepsis

    V. V. Nazaretyan

    2017-01-01

    Full Text Available Aim of the study: the effectiveness of concomitant use of antioxidant therapy with antioxidant 2-ethyl-6- methyl-3-hydroxypyridine succinate (mexidol and intensive nutritional support with glutamine in patients with abdominal sepsis (AS. Materials and methods. 170 patients with abdominal sepsis (AS involved in the study were separated into two groups. Patients of group 1 (control group, n=70 received basic treatment. Patients from group 2 (n=100 were divided into 2 subgroups. Patients from the subgroup 21 (n=70, in additon to the basic treatment, received intravenously, by drop infusion, mexidol (2000 mg per day and dipeptiven (27.5 g per day, patients from subgroup 22 (n=30 additionally to that received per os glutamine. Survival analysis was carried out according to the Kaplan-Meier method with using of the Cox's F-test and Mantel-Cox test for testing of statistical hypotheses. Results. Treatment outcomes analysis showed that in the basic group 2, mortality was lower than in the control group 1. A statistically significant increase of cumulative part in the survivors was revealed using mexidol and glutamine. Conclusion. Concomitant intravenous administration of medications had positive effects on treatment outcomes. Following on from the analysis results, we may suggest that the pair mexidol + dipeptiven interrupts the cascade of development of abdominal sepsis and contributes to avoiding a critical condition during sepsis.

  2. Evolution of the Maillard Reaction in Glutamine or Arginine-Dextrinomaltose Model Systems

    Silvia Pastoriza

    2016-12-01

    Full Text Available Enteral formulas are foods designed for medical uses to feed patients who are unable to eat normally. They are prepared by mixing proteins, amino acids, carbohydrates and fats and submitted to sterilization. During thermal treatment, the Maillard reaction takes place through the reaction of animo acids with reducing sugars. Thus, although glutamine and arginine are usually added to improve the nutritional value of enteral formulas, their final concentration may vary. Thus, in the present paper the early, intermediate, and advanced states of the Maillard reaction were studied in model systems by measuring loss of free amino acids through the decrease of fluorescence intensity with o-phtaldialdehyde (OPA, 5-Hydroximethylfurfural (HMF, furfural, glucosylisomaltol, fluorescence, and absorbance at 420 nm. The systems were prepared by mixing glutamine or arginine with dextrinomaltose (similar ingredients to those used in special enteral formula, and heated at 100 °C, 120 °C and 140 °C for 0 to 30 min. The recorded changes in the concentration of furanic compounds was only useful for longer heating times of high temperatures, while absorbance and fluorescence measurements were useful in all the assayed conditions. In addition, easiness and sensitivity of absorbance and fluorescence make them useful techniques that could be implemented as indicators for monitoring the manufacture of special enteral formulas. Glucosylisomaltol is a useful indicator to monitor the manufacture of glutamine-enriched enteral formulas.

  3. Is long term creatine and glutamine supplementation effective in enhancing physical performance of military police officers?

    da Silveira, Celismar Lázaro; de Souza, Thiago Siqueira Paiva; Batista, Gilmário Ricarte; de Araújo, Adenilson Targino; da Silva, Júlio César Gomes; de Sousa, Maria do Socorro Cirilo; Marta, Carlos; Garrido, Nuno Domingo

    2014-09-29

    The objective of this study was to analyze the effect of supplementation with creatine and glutamine on physical fitness of military police officers. Therefore, an experimental double blind study was developed, with the final sample composed by 32 men randomly distributed into three groups: a group supplemented with creatine (n=10), glutamine (n=10) and a placebo group (n=12) and evaluated in three distinct moments, in an interval of three months (T1, T2 and T3). The physical training had a weekly frequency of 5 sessions × 90 min, including strength exercises, local muscular resistance, flexibility and both aerobic and anaerobic capacity. After analyzing the effect of time, group and interaction (group × time) for measures that indicated the physical capabilities of the subjects, a significant effect of time for the entire variable was identified (p0,05). In face of the results it was concluded that supplementation with creatine and glutamine showed no ergogenic effect on physical performance in military police officers.

  4. Glutamine prevents gastric oxidative stress in an animal model of portal hypertension gastropathy.

    Marques, Camila; Mauriz, José L; Simonetto, Douglas; Marroni, Claudio A; Tuñon, María J; González-Gallego, Javier; Marrón, Norma P

    2011-01-01

    Portal hypertension (PHI) is a clinical syndrome characterized by increases of the blood flow and/or of the vascular resistance in the portal system. A direct consequence of PHI can appearance different lesions on the gastric mucosa and submucosa, cumulatively termed portal hypertensive gastropathy (PHG). To investigate the effects of glutamine on oxidative stress in an experimental model of PHG induced by partial portal vein ligation (PPVL). Portal pressure, transaminase and alkaline phosphatase activity were quantified. Gastric tissue damage was assessed by histological analysis. Oxidative stress was measured by quantification of cytosolic concentration of thiobarbituric acid reactive substances (TBARS), hydroperoxide-initiated chemiluminescence (QL), and nitric oxide (NO) production. Moreover, activities of the antioxidant enzymes superoxide dismutase (SOD), glutathione peroxidase (GPx), and catalase (CAT) were analyzed. Transaminase and alkaline phosphatase activities were not significantly modified by PPVL, indicating absence of liver injury. Histological analysis of gastric sections showed a lost of normal architecture, with edema and vasodilatation. TBARS, QL, and NO production were significantly increased in PPVL animals. A reduction of SOD activity was found. Glutamine administration markedly alleviated histological abnormalities and oxidative stress, normalized SOD activity, and blocked NO overproduction. Our results confirm that the use of molecules with antioxidant capacity can provide protection of the gastric tissue in portal hypertension. Glutamine treatment can be useful to reduce the oxidative damage induced by PHI on gastric tissue.

  5. The standard enthalpies of formation of L-asparagine and L-{alpha}-glutamine

    Contineanu, Iulia, E-mail: icontineanu@yahoo.com [Institute of Physical Chemistry ' I.G. Murgulescu' , Spl. Independentei 202, Bucharest (Romania); Neacsu, Ana, E-mail: anna_matache@yahoo.com [Institute of Physical Chemistry ' I.G. Murgulescu' , Spl. Independentei 202, Bucharest (Romania); Perisanu, Stefan T., E-mail: stefan.perisanu@upb.ro [Laboratory of General Chemistry, Polytechnic University of Bucharest, str. Polizu nr. 1, Bucuresti (Romania)

    2010-01-10

    The energies of combustion of L-asparagine and L-{alpha}-glutamine were measured in a static bomb adiabatic calorimeter. Corrections were made for the heats due to the ignition of sample and for the nitric acid formation. The derived enthalpies of formation in solid state of asparagine monohydrate, nonhydrated asparagine and glutamine are respectively -1084.1 {+-} 3.0, -788.1 {+-} 4.7 and -834.3 {+-} 4.6 kJ mol{sup -1}. The data of enthalpy of formation are compared with the literature values and with estimated values by means of group additivity, using parameters recommended by Domalski and Hearing. The discrepancies between experimental and calculated values are explained by considering the number and strength of intermolecular hydrogen bonds. The dehydration of asparagine monohydrate and the possible melting of the three amino acids were investigated by means of DSC. Glutamine melts without considerable decomposition at about 182 {sup o}C, while asparagines decompose during the fusion process (208 {sup o}C).

  6. Effects of Glutamine and Alanine Supplementation on Central Fatigue Markers in Rats Submitted to Resistance Training

    Audrey Yule Coqueiro

    2018-01-01

    Full Text Available Recent evidence suggests that increased brain serotonin synthesis impairs performance in high-intensity intermittent exercise and specific amino acids may modulate this condition, delaying fatigue. This study investigated the effects of glutamine and alanine supplementation on central fatigue markers in rats submitted to resistance training (RT. Wistar rats were distributed in: sedentary (SED, trained (CON, trained and supplemented with alanine (ALA, glutamine and alanine in their free form (G + A, or as dipeptide (DIP. Trained groups underwent a ladder-climbing exercise for eight weeks, with progressive loads. In the last 21 days, supplementations were offered in water with a 4% concentration. Albeit without statistically significance difference, RT decreased liver glycogen, and enhanced the concentrations of plasma glucose, free fatty acids (FFA, hypothalamic serotonin, and ammonia in muscle and the liver. Amino acids affected fatigue parameters depending on the supplementation form. G + A prevented the muscle ammonia increase by RT, whereas ALA and DIP augmented ammonia and glycogen concentrations in muscle. DIP also increased liver ammonia. ALA and G + A reduced plasma FFA, whereas DIP increased this parameter, free tryptophan/total tryptophan ratio, hypothalamic serotonin, and the serotonin/dopamine ratio. The supplementations did not affect physical performance. In conclusion, glutamine and alanine may improve or impair central fatigue markers depending on their supplementation form.

  7. Is Long Term Creatine and Glutamine Supplementation Effective in Enhancing Physical Performance of Military Police Officers?

    Lázaro da Silveira Celismar

    2014-12-01

    Full Text Available The objective of this study was to analyze the effect of supplementation with creatine and glutamine on physical fitness of military police officers. Therefore, an experimental double blind study was developed, with the final sample composed by 32 men randomly distributed into three groups: a group supplemented with creatine (n=10, glutamine (n=10 and a placebo group (n=12 and evaluated in three distinct moments, in an interval of three months (T1, T2 and T3. The physical training had a weekly frequency of 5 sessions x 90 min, including strength exercises, local muscular resistance, flexibility and both aerobic and anaerobic capacity. After analyzing the effect of time, group and interaction (group x time for measures that indicated the physical capabilities of the subjects, a significant effect of time for the entire variable was identified (p0,05. In face of the results it was concluded that supplementation with creatine and glutamine showed no ergogenic effect on physical performance in military police officers.

  8. Evolution of the Maillard Reaction in Glutamine or Arginine-Dextrinomaltose Model Systems.

    Pastoriza, Silvia; Rufián-Henares, José Ángel; García-Villanova, Belén; Guerra-Hernández, Eduardo

    2016-12-07

    Enteral formulas are foods designed for medical uses to feed patients who are unable to eat normally. They are prepared by mixing proteins, amino acids, carbohydrates and fats and submitted to sterilization. During thermal treatment, the Maillard reaction takes place through the reaction of animo acids with reducing sugars. Thus, although glutamine and arginine are usually added to improve the nutritional value of enteral formulas, their final concentration may vary. Thus, in the present paper the early, intermediate, and advanced states of the Maillard reaction were studied in model systems by measuring loss of free amino acids through the decrease of fluorescence intensity with o -phtaldialdehyde (OPA), 5-Hydroximethylfurfural (HMF), furfural, glucosylisomaltol, fluorescence, and absorbance at 420 nm. The systems were prepared by mixing glutamine or arginine with dextrinomaltose (similar ingredients to those used in special enteral formula), and heated at 100 °C, 120 °C and 140 °C for 0 to 30 min. The recorded changes in the concentration of furanic compounds was only useful for longer heating times of high temperatures, while absorbance and fluorescence measurements were useful in all the assayed conditions. In addition, easiness and sensitivity of absorbance and fluorescence make them useful techniques that could be implemented as indicators for monitoring the manufacture of special enteral formulas. Glucosylisomaltol is a useful indicator to monitor the manufacture of glutamine-enriched enteral formulas.

  9. Mineral nitrogen sources differently affect root glutamine synthetase isoforms and amino acid balance among organs in maize.

    Prinsi, Bhakti; Espen, Luca

    2015-04-03

    Glutamine synthetase (GS) catalyzes the first step of nitrogen assimilation in plant cell. The main GS are classified as cytosolic GS1 and plastidial GS2, of which the functionality is variable according to the nitrogen sources, organs and developmental stages. In maize (Zea mays L.) one gene for GS2 and five genes for GS1 subunits are known, but their roles in root metabolism are not yet well defined. In this work, proteomic and biochemical approaches have been used to study root GS enzymes and nitrogen assimilation in maize plants re-supplied with nitrate, ammonium or both. The plant metabolic status highlighted the relevance of root system in maize nitrogen assimilation during both nitrate and ammonium nutrition. The analysis of root proteomes allowed a study to be made of the accumulation and phosphorylation of six GS proteins. Three forms of GS2 were identified, among which only the phosphorylated one showed an accumulation trend consistent with plastidial GS activity. Nitrogen availabilities enabled increments in root total GS synthetase activity, associated with different GS1 isoforms according to the nitrogen sources. Nitrate nutrition induced the specific accumulation of GS1-5 while ammonium led to up-accumulation of both GS1-1 and GS1-5, highlighting co-participation. Moreover, the changes in thermal sensitivity of root GS transferase activity suggested differential rearrangements of the native enzyme. The amino acid accumulation and composition in roots, xylem sap and leaves deeply changed in response to mineral sources. Glutamine showed the prevalent changes in all nitrogen nutritions. Besides, the ammonium nutrition was associated with an accumulation of asparagine and reducing sugars and a drop in glutamic acid level, significantly alleviated by the co-provision with nitrate. This work provides new information about the multifaceted regulation of the GS enzyme in maize roots, indicating the involvement of specific isoenzymes/isoforms, post

  10. Establishing in vitro Zinnia elegans cell suspension culture with high tracheary elements differentiation

    Twumasi, P.; Schel, J.H.N.; Ieperen, van W.; Woltering, E.J.; Emons, A.M.C.

    2009-01-01

    The Zinnia elegans mesophyll cell culture is a useful system for xylogenesis studies. The system is associated with highly synchronous tracheary element (TE) differentiation, making it more suitable for molecular studies requiring larger amounts of molecular isolates, such as mRNA and proteins and

  11. The effects of glutamine/asparagine content on aggregation and heterologous prion induction by yeast prion-like domains.

    Shattuck, Jenifer E; Waechter, Aubrey C; Ross, Eric D

    2017-07-04

    Prion-like domains are low complexity, intrinsically disordered domains that compositionally resemble yeast prion domains. Many prion-like domains are involved in the formation of either functional or pathogenic protein aggregates. These aggregates range from highly dynamic liquid droplets to highly ordered detergent-insoluble amyloid-like aggregates. To better understand the amino acid sequence features that promote conversion to stable, detergent-insoluble aggregates, we used the prediction algorithm PAPA to identify predicted aggregation-prone prion-like domains with a range of compositions. While almost all of the predicted aggregation-prone domains formed foci when expressed in cells, the ability to form the detergent-insoluble aggregates was highly correlated with glutamine/asparagine (Q/N) content, suggesting that high Q/N content may specifically promote conversion to the amyloid state in vivo. We then used this data set to examine cross-seeding between prion-like proteins. The prion protein Sup35 requires the presence of a second prion, [PIN + ], to efficiently form prions, but this requirement can be circumvented by the expression of various Q/N-rich protein fragments. Interestingly, almost all of the Q/N-rich domains that formed SDS-insoluble aggregates were able to promote prion formation by Sup35, highlighting the highly promiscuous nature of these interactions.

  12. [Cardioprotective effects of glutamine in patients with ischemic heart disease operated under conditions of extracorporeal blood circulation].

    Lomivorotov, V V; Efremov, S M; Shmyrev, V A; Ponomarev, D N; Sviatchenko, A V; Kniaz'kova, L G

    2012-01-01

    It was conducted a study of glutamine cardioptotective effects during perioperative use in patients with ischemic heart disease, operated under CB. Exclusion criteria were: left ventricular ejection fraction less than 50%, diabetes melitus, myocardial infarction less than 3 months ago, Patients of the study group (n=25) had glutamine (20% solution N(2)-L-alanine-L-glutamine ("Dipeptiven" Fresenius Kabi, Germany); 0.4 g/kg/day. Patients of control group (n=25) received placebo (0.9% NaCl solution). The main indicators were the dynamics of troponin I, as well as central hemodynamics parameters. On the 1-st day after operation the concentration of troponin I was significantly lower in the glutamine-group compared placebo-group (1.280 (0.840-2.230) 2.410 (1.060-6.600) ng/ml; p=0.035). 4 hours after CB in a glutamine-group also had significantly large indicators of cardiac index (2.58 (2.34-2.91) l/min/m2 vs 2.03 (1.76-2.32)) l/min/m2; p=0,002) and stroke index (32.8 (27.8-36.0.) ml/m2 vs 26.1 (22.6-31.8) ml/m2; p=0.023). Systemic vascular resistance index was significantly lower in glutamine-group (1942 (1828-2209) dyn x s/cm(-5)/m2 vs 2456 (2400-3265) dyn x s/cm(-5)/m2; p=0.001). Conclusion. Perioperative use of N(2)-L-alanine-L-glutamine during the first 24 hours ofperioperative period gives cardioprotective effect in patients with ischemic heart disease operated under CB.

  13. The structures of cytosolic and plastid-located glutamine synthetases from Medicago truncatula reveal a common and dynamic architecture

    Torreira, Eva; Seabra, Ana Rita; Marriott, Hazel; Zhou, Min; Llorca, Óscar; Robinson, Carol V.; Carvalho, Helena G.; Fernández-Tornero, Carlos; Pereira, Pedro José Barbosa

    2014-01-01

    The experimental models of dicotyledonous cytoplasmic and plastid-located glutamine synthetases unveil a conserved eukaryotic-type decameric architecture, with subtle structural differences in M. truncatula isoenzymes that account for their distinct herbicide resistance. The first step of nitrogen assimilation in higher plants, the energy-driven incorporation of ammonia into glutamate, is catalyzed by glutamine synthetase. This central process yields the readily metabolizable glutamine, which in turn is at the basis of all subsequent biosynthesis of nitrogenous compounds. The essential role performed by glutamine synthetase makes it a prime target for herbicidal compounds, but also a suitable intervention point for the improvement of crop yields. Although the majority of crop plants are dicotyledonous, little is known about the structural organization of glutamine synthetase in these organisms and about the functional differences between the different isoforms. Here, the structural characterization of two glutamine synthetase isoforms from the model legume Medicago truncatula is reported: the crystallographic structure of cytoplasmic GSII-1a and an electron cryomicroscopy reconstruction of plastid-located GSII-2a. Together, these structural models unveil a decameric organization of dicotyledonous glutamine synthetase, with two pentameric rings weakly connected by inter-ring loops. Moreover, rearrangement of these dynamic loops changes the relative orientation of the rings, suggesting a zipper-like mechanism for their assembly into a decameric enzyme. Finally, the atomic structure of M. truncatula GSII-1a provides important insights into the structural determinants of herbicide resistance in this family of enzymes, opening new avenues for the development of herbicide-resistant plants

  14. The structures of cytosolic and plastid-located glutamine synthetases from Medicago truncatula reveal a common and dynamic architecture

    Torreira, Eva [Centro de Investigaciones Biológicas – CSIC, Ramiro de Maeztu 9, 28040 Madrid (Spain); Seabra, Ana Rita [IBMC – Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto (Portugal); Marriott, Hazel; Zhou, Min [University of Oxford, South Parks Road, Oxford OX1 3QZ (United Kingdom); Llorca, Óscar [Centro de Investigaciones Biológicas – CSIC, Ramiro de Maeztu 9, 28040 Madrid (Spain); Robinson, Carol V. [University of Oxford, South Parks Road, Oxford OX1 3QZ (United Kingdom); Carvalho, Helena G. [IBMC – Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto (Portugal); Fernández-Tornero, Carlos, E-mail: cftornero@cib.csic.es [Centro de Investigaciones Biológicas – CSIC, Ramiro de Maeztu 9, 28040 Madrid (Spain); Pereira, Pedro José Barbosa, E-mail: cftornero@cib.csic.es [IBMC – Instituto de Biologia Molecular e Celular, Universidade do Porto, Rua do Campo Alegre 823, 4150-180 Porto (Portugal); Centro de Investigaciones Biológicas – CSIC, Ramiro de Maeztu 9, 28040 Madrid (Spain)

    2014-04-01

    The experimental models of dicotyledonous cytoplasmic and plastid-located glutamine synthetases unveil a conserved eukaryotic-type decameric architecture, with subtle structural differences in M. truncatula isoenzymes that account for their distinct herbicide resistance. The first step of nitrogen assimilation in higher plants, the energy-driven incorporation of ammonia into glutamate, is catalyzed by glutamine synthetase. This central process yields the readily metabolizable glutamine, which in turn is at the basis of all subsequent biosynthesis of nitrogenous compounds. The essential role performed by glutamine synthetase makes it a prime target for herbicidal compounds, but also a suitable intervention point for the improvement of crop yields. Although the majority of crop plants are dicotyledonous, little is known about the structural organization of glutamine synthetase in these organisms and about the functional differences between the different isoforms. Here, the structural characterization of two glutamine synthetase isoforms from the model legume Medicago truncatula is reported: the crystallographic structure of cytoplasmic GSII-1a and an electron cryomicroscopy reconstruction of plastid-located GSII-2a. Together, these structural models unveil a decameric organization of dicotyledonous glutamine synthetase, with two pentameric rings weakly connected by inter-ring loops. Moreover, rearrangement of these dynamic loops changes the relative orientation of the rings, suggesting a zipper-like mechanism for their assembly into a decameric enzyme. Finally, the atomic structure of M. truncatula GSII-1a provides important insights into the structural determinants of herbicide resistance in this family of enzymes, opening new avenues for the development of herbicide-resistant plants.

  15. ISOLATION OF MESOPHYLL PROTOPLASTS FROM MEDITERRANEAN WOODY PLANTS FOR THE STUDY OF DNA INTEGRITY UNDER ABIOTIC STRESS

    Elena Kuzminsky

    2016-08-01

    Full Text Available Abiotic stresses have considerable negative impact on Mediterranean plant ecosystems and better comprehension of the genetic control of response and adaptation of trees to global changes is urgently needed. The Single Cell Gel Electrophoresis assay could be considered a good estimator of DNA damage in an individual eukaryotic cell. This method has been mainly employed in animal tissues, because the plant cell wall represents an obstacle for the extraction of nuclei; moreover, in Mediterranean woody species, especially in the sclerophyll plants, this procedure can be quite difficult because of the presence of sclerenchyma and hardened cells. On the other hand, these plants represent an interesting material to be studied because of the ability of these plants to tolerate abiotic stress. For instance, holm oak (Quercus ilex L. has been selected as the model plant to identify critical levels of O3 for Southern European forests. Consequently, a quantitative method for the evaluation of cell injury of leaf tissues of this species is required. Optimal conditions for high-yield nuclei isolation were obtained by using protoplast technology and a detailed description of the method is provided and discussed. White poplar (Populus alba L. was used as an internal control for protoplast isolation. Such a method has not been previously reported in newly fully developed leaves of holm oak. This method combined with Single Cell Gel Electrophoresis assay represents a new tool for testing the DNA integrity of leaf tissues in higher plants under stress conditions.

  16. L-glutamine supplementation prevents the development of experimental diabetic cardiomyopathy in streptozotocin-nicotinamide induced diabetic rats.

    Sachin L Badole

    Full Text Available The objective of the present investigation was to evaluate the effect of L-glutamine on cardiac myopathy in streptozotocin-nicotinamide induced diabetic rats. Diabetes was induced in overnight fasted Sprague Dawely rats by using intraperitonial injection of streptozotocin (55 mg/kg. Nicotinamide (100 mg/kg, i.p. was administered 20 min before administration of streptozotocin. Experimental rats were divided into Group I: non-diabetic control (distilled water; 10 ml/kg, p.o., II: diabetic control (distilled water, 10 ml/kg, p.o., III: L-glutamine (500 mg/kg, p.o. and IV: L-glutamine (1000 mg/kg, p.o.. All groups were diabetic except group I. The plasma glucose level, body weight, electrocardiographic abnormalities, hemodynamic changes and left ventricular contractile function, biological markers of cardiotoxicity, antioxidant markers were determined after 4 months after STZ with nicotinamide injection. Histopathological changes of heart tissue were carried out by using H and E stain. L-glutamine treatment improved the electrocardiographic, hemodynamic changes; LV contractile function; biological markers; oxidative stress parameters and histological changes in STZ induced diabetic rats. Results from the present investigation demonstrated that L-glutamine has seemed a cardioprotective activity.

  17. Characterization of amino acid metabolism by cultured rat kidney cells: Study with 15N

    Nissim, I.; States, B.; Yudkoff, M.; Segal, S.

    1987-01-01

    The present study evaluates the metabolism of glutamine and glutamate by normal rat kidney (NRK) cells. The major aim was to evaluate the effect of acute acidosis on the metabolism of amino acid and ammonia formation by cultured NRK cells. Experiments at either pH 7.0 or 7.4 were conducted with phosphate-buffered saline supplemented with either 1 mM [5- 15 N]glutamine, [2- 15 N]glutamine, or [ 15 N]glutamate. Incubation with either glutamine or glutamate as a precursor showed that production of ammonia and glucose was increased significantly at pH 7.0 vs 7.4. In experiments with [5- 15 N]glutamine, the authors found that ∼57 and 43% of ammonia N was derived from 5-N of glutamine at pH 7.4 and 7.0, respectively. Three major metabolic pathways of [2- 15 N]glutamine or [ 15 N]glutamate disposal were identified: (1) transamination reactions involving the pH-independent formation of [ 15 N] aspartate and [ 15 N]alanine; (2) the synthesis of [6- 15 NH 2 ]adenine nucleotide, a process more active at pH 7.4 vs. 7.0; and (3) glutamine synthesis from [ 15 N]glutamate, especially at pH 7.4. The data indicate that NRK cells in culture consume glutamine and glutamate and generate ammonia and various amino acids, depending on the H + concentration in the media. The studies suggest that these cell lines may provide a useful model for studying various aspects of the effect of pH on rat renal ammoniagenesis

  18. Cell-specific vacuolar calcium storage mediated by CAX1 regulates apoplastic calcium concentration, gas exchange, and plant productivity in Arabidopsis.

    Conn, Simon J; Gilliham, Matthew; Athman, Asmini; Schreiber, Andreas W; Baumann, Ute; Moller, Isabel; Cheng, Ning-Hui; Stancombe, Matthew A; Hirschi, Kendal D; Webb, Alex A R; Burton, Rachel; Kaiser, Brent N; Tyerman, Stephen D; Leigh, Roger A

    2011-01-01

    The physiological role and mechanism of nutrient storage within vacuoles of specific cell types is poorly understood. Transcript profiles from Arabidopsis thaliana leaf cells differing in calcium concentration ([Ca], epidermis 60 mM) were compared using a microarray screen and single-cell quantitative PCR. Three tonoplast-localized Ca(2+) transporters, CAX1 (Ca(2+)/H(+)-antiporter), ACA4, and ACA11 (Ca(2+)-ATPases), were identified as preferentially expressed in Ca-rich mesophyll. Analysis of respective loss-of-function mutants demonstrated that only a mutant that lacked expression of both CAX1 and CAX3, a gene ectopically expressed in leaves upon knockout of CAX1, had reduced mesophyll [Ca]. Reduced capacity for mesophyll Ca accumulation resulted in reduced cell wall extensibility, stomatal aperture, transpiration, CO(2) assimilation, and leaf growth rate; increased transcript abundance of other Ca(2+) transporter genes; altered expression of cell wall-modifying proteins, including members of the pectinmethylesterase, expansin, cellulose synthase, and polygalacturonase families; and higher pectin concentrations and thicker cell walls. We demonstrate that these phenotypes result from altered apoplastic free [Ca(2+)], which is threefold greater in cax1/cax3 than in wild-type plants. We establish CAX1 as a key regulator of apoplastic [Ca(2+)] through compartmentation into mesophyll vacuoles, a mechanism essential for optimal plant function and productivity.

  19. Deletion of Type I glutamine synthetase deregulates nitrogen metabolism and increases ethanol production in Clostridium thermocellum

    Rydzak, Thomas [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Garcia, David [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Stevenson, David M. [Univ. of Wisconsin, Madison, WI (United States). Dept. of Bacteriology; Sladek, Margaret [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Klingeman, Dawn M. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Holwerda, Evert K. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division; Dartmouth College, Hanover, NH (United States). Thayer School of Engineering; Amador-Noguez, Daniel [Univ. of Wisconsin, Madison, WI (United States). Dept. of Bacteriology; Brown, Steven D. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center; Guss, Adam M. [Oak Ridge National Lab. (ORNL), Oak Ridge, TN (United States). Biosciences Division, BioEnergy Science Center

    2017-05-01

    Clostridium thermocellum rapidly deconstructs cellulose and ferments resulting hydrolysis products into ethanol and other products, and is thus a promising platform organism for the development of cellulosic biofuel production via consolidated bioprocessing. And while recent metabolic engineering strategies have targeted eliminating canonical fermentation products (acetate, lactate, formate, and H2), C. thermocellum also secretes amino acids, which has limited ethanol yields in engineered strains to approximately 70% of the theoretical maximum. To decrease amino acid secretion, we attempted to reduce ammonium assimilation by deleting the Type I glutamine synthetase (glnA) in C. thermocellum. Deletion of glnA reduced levels of secreted valine and total amino acids by 53% and 44% respectively, and increased ethanol yields by 53%. RNA-seq analysis revealed that genes encoding the RNF-complex were more highly expressed in ΔglnA and may have a role in improving NADH-availability for ethanol production. While a significant up-regulation of genes involved in nitrogen assimilation and urea uptake suggested that deletion of glnA induces a nitrogen starvation response, metabolomic analysis showed an increase in intracellular glutamine and α-ketoglutarate levels indicative of nitrogen-rich conditions. Here, we propose that deletion of glnA causes deregulation of nitrogen metabolism, leading to overexpression of nitrogen metabolism genes and, in turn, elevated glutamine/α-ketoglutarate levels. Here we demonstrate that perturbation of nitrogen assimilation is a promising strategy to redirect flux from the production of nitrogenous compounds toward biofuels in C. thermocellum.

  20. Combined enteral infusion of glutamine, carbohydrates, and antioxidants modulates gut protein metabolism in humans.

    Coëffier, Moïse; Claeyssens, Sophie; Lecleire, Stéphane; Leblond, Jonathan; Coquard, Aude; Bôle-Feysot, Christine; Lavoinne, Alain; Ducrotté, Philippe; Déchelotte, Pierre

    2008-11-01

    Available data suggest that nutrients can affect intestinal protein metabolism, which contributes to the regulation of gut barrier function. We aimed to assess whether an oral nutritional supplement (ONS) containing glutamine (as the dipeptide Ala-Gln), carbohydrates, and antioxidants would modulate duodenal protein metabolism in healthy humans. Thirty healthy control subjects were included and, over a period of 5 h, received by nasogastric tube either saline or ONS providing 11.7 kcal/kg as 0.877 g Ala-Gln/kg, 3.9 g carbohydrates/kg, and antioxidants (29.25 mg vitamin C/kg, 9.75 mg vitamin E/kg, 195 microg beta-carotene/kg, 5.85 mg Se/kg, and 390 microg Zn/kg) or glutamine (0.585 g/kg, 2.34 kcal/kg). Simultaneously, a continuous intravenous infusion of l-[1-(13)C]-leucine was done until endoscopy. Leucine enrichment was assessed by using gas chromatography-mass spectrometric analysis, and mucosal fractional synthesis rate was calculated by using intracellular amino acid enrichment as precursor. Mucosal proteolytic pathways were also evaluated. ONS infusion resulted in a doubling increase (P < 0.01) of duodenal fractional synthesis rate and a significant (P < 0.05) decrease in cathepsin D-mediated proteolysis compared with saline, whereas proteasome and Ca(2+)-dependent activities were unaffected. ONS infusion significantly (P < 0.01) decreased duodenal glutathione but not glutathione disulfide concentrations or the ratio of glutathione to glutathione disulfide. Insulinemia increased after ONS infusion, whereas plasma essential amino acids decreased. Infusion of glutamine alone did not reproduce ONS effects. ONS infusion improves duodenal protein balance in healthy humans. Further investigations are needed to study the origin of these effects and to evaluate ONS supply in stressed persons.

  1. Deletion of Type I glutamine synthetase deregulates nitrogen metabolism and increases ethanol production in Clostridium thermocellum.

    Rydzak, Thomas; Garcia, David; Stevenson, David M; Sladek, Margaret; Klingeman, Dawn M; Holwerda, Evert K; Amador-Noguez, Daniel; Brown, Steven D; Guss, Adam M

    2017-05-01

    Clostridium thermocellum rapidly deconstructs cellulose and ferments resulting hydrolysis products into ethanol and other products, and is thus a promising platform organism for the development of cellulosic biofuel production via consolidated bioprocessing. While recent metabolic engineering strategies have targeted eliminating canonical fermentation products (acetate, lactate, formate, and H 2 ), C. thermocellum also secretes amino acids, which has limited ethanol yields in engineered strains to approximately 70% of the theoretical maximum. To investigate approaches to decrease amino acid secretion, we attempted to reduce ammonium assimilation by deleting the Type I glutamine synthetase (glnA) in an essentially wild type strain of C. thermocellum. Deletion of glnA reduced levels of secreted valine and total amino acids by 53% and 44% respectively, and increased ethanol yields by 53%. RNA-seq analysis revealed that genes encoding the RNF-complex were more highly expressed in ΔglnA and may have a role in improving NADH-availability for ethanol production. While a significant up-regulation of genes involved in nitrogen assimilation and urea uptake suggested that deletion of glnA induces a nitrogen starvation response, metabolomic analysis showed an increase in intracellular glutamine levels indicative of nitrogen-rich conditions. We propose that deletion of glnA causes deregulation of nitrogen metabolism, leading to overexpression of nitrogen metabolism genes and, in turn, elevated glutamine levels. Here we demonstrate that perturbation of nitrogen assimilation is a promising strategy to redirect flux from the production of nitrogenous compounds toward biofuels in C. thermocellum. Copyright © 2017. Published by Elsevier Inc.

  2. Persistent negative temperature response of mesophyll conductance in red raspberry (Rubus idaeus L.) leaves under both high and low vapour pressure deficits: a role for abscisic acid?

    Qiu, Changpeng; Ethier, Gilbert; Pepin, Steeve; Dubé, Pascal; Desjardins, Yves; Gosselin, André

    2017-09-01

    The temperature dependence of mesophyll conductance (g m ) was measured in well-watered red raspberry (Rubus idaeus L.) plants acclimated to leaf-to-air vapour pressure deficit (VPDL) daytime differentials of contrasting amplitude, keeping a fixed diurnal leaf temperature (T leaf ) rise from 20 to 35 °C. Contrary to the great majority of g m temperature responses published to date, we found a pronounced reduction of g m with increasing T leaf irrespective of leaf chamber O 2 level and diurnal VPDL regime. Leaf hydraulic conductance was greatly enhanced during the warmer afternoon periods under both low (0.75 to 1.5 kPa) and high (0.75 to 3.5 kPa) diurnal VPDL regimes, unlike stomatal conductance (g s ), which decreased in the afternoon. Consequently, the leaf water status remained largely isohydric throughout the day, and therefore cannot be evoked to explain the diurnal decrease of g m . However, the concerted diurnal reductions of g m and g s were well correlated with increases in leaf abscisic acid (ABA) content, thus suggesting that ABA can induce a significant depression of g m under favourable leaf water status. Our results challenge the view that the temperature dependence of g m can be explained solely from dynamic leaf anatomical adjustments and/or from the known thermodynamic properties of aqueous solutions and lipid membranes.​. © 2017 John Wiley & Sons Ltd.

  3. Regulation of photosynthesis and stomatal and mesophyll conductance under water stress and recovery in olive trees: correlation with gene expression of carbonic anhydrase and aquaporins.

    Perez-Martin, Alfonso; Michelazzo, Chiara; Torres-Ruiz, Jose M; Flexas, Jaume; Fernández, José E; Sebastiani, Luca; Diaz-Espejo, Antonio

    2014-07-01

    The hypothesis that aquaporins and carbonic anhydrase (CA) are involved in the regulation of stomatal (g s) and mesophyll (g m) conductance to CO2 was tested in a short-term water-stress and recovery experiment in 5-year-old olive plants (Olea europaea) growing outdoors. The evolution of leaf gas exchange, chlorophyll fluorescence, and plant water status, and a quantitative analysis of photosynthesis limitations, were followed during water stress and recovery. These variables were correlated with gene expression of the aquaporins OePIP1.1 and OePIP2.1, and stromal CA. At mild stress and at the beginning of the recovery period, stomatal limitations prevailed, while the decline in g m accounted for up to 60% of photosynthesis limitations under severe water stress. However, g m was restored to control values shortly after rewatering, facilitating the recovery of the photosynthetic rate. CA was downregulated during water stress and upregulated after recovery. The use of structural equation modelling allowed us to conclude that both OePIP1.1 and OePIP2.1 expression could explain most of the variations observed for g s and g m. CA expression also had a small but significant effect on g m in olive under water-stress conditions. © The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.

  4. Mesophyll conductance decreases in the wild type but not in an ABA-deficient mutant (aba1) of Nicotiana plumbaginifolia under drought conditions.

    Mizokami, Yusuke; Noguchi, Ko; Kojima, Mikiko; Sakakibara, Hitoshi; Terashima, Ichiro

    2015-03-01

    Under drought conditions, leaf photosynthesis is limited by the supply of CO2 . Drought induces production of abscisic acid (ABA), and ABA decreases stomatal conductance (gs ). Previous papers reported that the drought stress also causes the decrease in mesophyll conductance (gm ). However, the relationships between ABA content and gm are unclear. We investigated the responses of gm to the leaf ABA content [(ABA)L ] using an ABA-deficient mutant, aba1, and the wild type (WT) of Nicotiana plumbaginifolia. We also measured leaf water potential (ΨL ) because leaf hydraulics may be related to gm . Under drought conditions, gm decreased with the increase in (ABA)L in WT, whereas both (ABA)L and gm were unchanged by the drought treatment in aba1. Exogenously applied ABA decreased gm in both WT and aba1 in a dose-dependent manner. ΨL in WT was decreased by the drought treatment to -0.7 MPa, whereas ΨL in aba1 was around -0.8 MPa even under the well-watered conditions and unchanged by the drought treatment. From these results, we conclude that the increase in (ABA)L is crucial for the decrease in gm under drought conditions. We discuss possible relationships between the decrease in gm and changes in the leaf hydraulics. © 2014 John Wiley & Sons Ltd.

  5. Antioxidant properties of glutamine and its role in VEGF-Akt pathways in portal hypertension gastropathy.

    Marques, Camila; Licks, Francielli; Zattoni, Ingrid; Borges, Beatriz; de Souza, Luiz Eduardo Rizzo; Marroni, Claudio Augusto; Marroni, Norma Possa

    2013-07-28

    To investigate the effects of glutamine on oxidative/nitrosative stress and the vascular endothelial growth factor (VEGF)-Akt-endothelial nitric oxide synthase (eNOS) signaling pathway in an experimental model of portal hypertension induced by partial portal vein ligation (PPVL). Portal hypertension was induced by PPVL. The PPVL model consists of a partial obstruction of the portal vein, performed using a 20 G blunt needle as a guide, which is gently removed after the procedure. PPVL model was performed for 14 d beginning treatment with glutamine on the seventh day. On the fifteenth day, the mesenteric vein pressure was checked and the stomach was removed to test immunoreactivity and oxidative stress markers. We evaluated the expression and the immunoreactivity of proteins involved in the VEGF-Akt-eNOS pathway by Western blotting and immunohistochemical analysis. Oxidative stress was measured by quantification of the cytosolic concentration of thiobarbituric acid reactive substances (TBARS) as well as the levels of total glutathione (GSH), superoxide dismutase (SOD) activity, nitric oxide (NO) production and nitrotyrosine immunoreactivity. All data are presented as the mean ± SE. The production of TBARS and NO was significantly increased in PPVL animals. A reduction of SOD activity was detected in PPVL + G group. In the immunohistochemical analyses of nitrotyrosine, Akt and eNOS, the PPVL group exhibited significant increases, whereas decreases were observed in the PPVL + G group, but no difference in VEGF was detected between these groups. Western blotting analysis detected increased expression of phosphatidylinositol-3-kinase (PI3K), P-Akt and eNOS in the PPVL group compared with the PPVL + G group, which was not observed for the expression of VEGF when comparing these groups. Glutamine administration markedly alleviated oxidative/nitrosative stress, normalized SOD activity, increased levels of total GSH and blocked NO overproduction as well as the formation of

  6. BLOOD AMMONIA AND GLUTAMINE AS PREDICTORS OF HYPERAMMONEMIC CRISES IN UREA CYCLE DISORDER PATIENTS

    Lee, Brendan; Diaz, George A.; Rhead, William; Lichter-Konecki, U.; Feigenbaum, Annette; Berry, Susan A.; Le Mons, C.; Bartley, James A; Longo, Nicola; Nagamani, Sandesh C.; Berquist, William; Gallagher, Renata; Bartholomew, Dennis; Harding, Cary O.; Korson, Mark S.; McCandless, Shawn E.; Smith, Wendy; Cederbaum, Stephen; Wong, Derek; Merritt, J. Lawrence; Schulze, A.; Vockley, Gerard.; Kronn, David; Zori, Roberto; Summar, Marshall; Milikien, D.A.; Marino, M.; Coakley, D.F.; Mokhtarani, M.; Scharschmidt, B.F.

    2014-01-01

    Purpose To examine predictors of ammonia exposure and hyperammonemic crises (HAC) in patients with urea cycle disorders (UCDs). Methods The relationships between fasting ammonia, daily ammonia exposure, and HACs were analyzed in >100 UCD patients. Results Fasting ammonia correlated strongly with daily ammonia exposure (r=0.764, p200% (purea nitrogen. Fasting glutamine correlated weakly with AUC0-24 and was not a significant predictor of HACs. Conclusions Fasting ammonia correlates strongly and positively with daily ammonia exposure and with the risk and rate of HACs, suggesting that UCD patients may benefit from tight ammonia control. PMID:25503497

  7. Glutamine and ornithine alpha-ketoglutarate supplementation on malate dehydrogenases expression in hepatectomized rats

    Guimarães Filho, Artur; Cunha, Rodrigo Maranguape Silva da; Vasconcelos, Paulo Roberto Leitão de; Guimarães, Sergio Botelho

    2014-01-01

    PURPOSE: To evaluate the relative gene expression (RGE) of cytosolic (MDH1) and mitochondrial (MDH2) malate dehydrogenases enzymes in partially hepatectomized rats after glutamine (GLN) or ornithine alpha-ketoglutarate (OKG) suplementation. METHODS: One-hundred and eight male Wistar rats were randomly distributed into six groups (n=18): CCaL, GLNL and OKGL and fed calcium caseinate (CCa), GLN and OKG, 0.5g/Kg by gavage, 30 minutes before laparotomy. CCaH, GLNH and OKGH groups were likewise fe...

  8. Randomised trial of glutamine and selenium supplemented parenteral nutrition for critically ill patients. Protocol Version 9, 19 February 2007 known as SIGNET (Scottish Intensive care Glutamine or seleNium Evaluative Trial

    Vale Luke D

    2007-09-01

    Full Text Available Abstract Background Mortality rates in the Intensive Care Unit and subsequent hospital mortality rates in the UK remain high. Infections in Intensive Care are associated with a 2–3 times increased risk of death. It is thought that under conditions of severe metabolic stress glutamine becomes "conditionally essential". Selenium is an essential trace element that has antioxidant and anti-inflammatory properties. Approximately 23% of patients in Intensive Care require parenteral nutrition and glutamine and selenium are either absent or present in low amounts. Both glutamine and selenium have the potential to influence the immune system through independent biochemical pathways. Systematic reviews suggest that supplementing parenteral nutrition in critical illness with glutamine or selenium may reduce infections and mortality. Pilot data has shown that more than 50% of participants developed infections, typically resistant organisms. We are powered to show definitively whether supplementation of PN with either glutamine or selenium is effective at reducing new infections in critically ill patients. Methods/design 2 × 2 factorial, pragmatic, multicentre, double-blind, randomised controlled trial. The trial has an enrolment target of 500 patients. Inclusion criteria include: expected to be in critical care for at least 48 hours, aged 16 years or over, patients who require parenteral nutrition and are expected to have at least half their daily nutritional requirements given by that route. Allocation is to one of four iso-caloric, iso-nitrogenous groups: glutamine, selenium, both glutamine & selenium or no additional glutamine or selenium. Trial supplementation is given for up to seven days on the Intensive Care Unit and subsequent wards if practicable. The primary outcomes are episodes of infection in the 14 days after starting trial nutrition and mortality. Secondary outcomes include antibiotic usage, length of hospital stay, quality of life and

  9. Randomised trial of glutamine and selenium supplemented parenteral nutrition for critically ill patients. Protocol Version 9, 19 February 2007 known as SIGNET (Scottish Intensive care Glutamine or seleNium Evaluative Trial).

    Andrews, Peter J D; Avenell, Alison; Noble, David W; Campbell, Marion K; Battison, Claire G; Croal, Bernard L; Simpson, William G; Norrie, John; Vale, Luke D; Cook, Jonathon; de Verteuil, Robyn; Milne, Anne C

    2007-09-20

    Mortality rates in the Intensive Care Unit and subsequent hospital mortality rates in the UK remain high. Infections in Intensive Care are associated with a 2-3 times increased risk of death. It is thought that under conditions of severe metabolic stress glutamine becomes "conditionally essential". Selenium is an essential trace element that has antioxidant and anti-inflammatory properties. Approximately 23% of patients in Intensive Care require parenteral nutrition and glutamine and selenium are either absent or present in low amounts. Both glutamine and selenium have the potential to influence the immune system through independent biochemical pathways. Systematic reviews suggest that supplementing parenteral nutrition in critical illness with glutamine or selenium may reduce infections and mortality. Pilot data has shown that more than 50% of participants developed infections, typically resistant organisms. We are powered to show definitively whether supplementation of PN with either glutamine or selenium is effective at reducing new infections in critically ill patients. 2 x 2 factorial, pragmatic, multicentre, double-blind, randomised controlled trial. The trial has an enrollment target of 500 patients. Inclusion criteria include: expected to be in critical care for at least 48 hours, aged 16 years or over, patients who require parenteral nutrition and are expected to have at least half their daily nutritional requirements given by that route. Allocation is to one of four iso-caloric, iso-nitrogenous groups: glutamine, selenium, both glutamine & selenium or no additional glutamine or selenium. Trial supplementation is given for up to seven days on the Intensive Care Unit and subsequent wards if practicable. The primary outcomes are episodes of infection in the 14 days after starting trial nutrition and mortality. Secondary outcomes include antibiotic usage, length of hospital stay, quality of life and cost-effectiveness. To date more than 285 patients have been

  10. Small glutamine-rich tetratricopeptide repeat-containing protein alpha is present in human ovaries but may not be differentially expressed in relation to polycystic ovary syndrome.

    Butler, Miriam S; Yang, Xing; Ricciardelli, Carmela; Liang, Xiaoyan; Norman, Robert J; Tilley, Wayne D; Hickey, Theresa E

    2013-06-01

    To evaluate the expression and function of small glutamine-rich tetratricopeptide repeat-containing protein alpha (SGTA), an androgen receptor (AR) molecular chaperone, in human ovarian tissues. Examine the effect of SGTA on AR subcellular localization in granulosa tumor cells (KGN) and SGTA expression in ovarian tissues. University-based research laboratory. Archived tissues from premenopausal women and granulosa cells from infertile women receiving assisted reproduction. None. AR subcellular localization and SGTA protein or mRNA levels. SGTA and AR proteins were expressed in the cytoplasm of KGN cells and exposure to androgen stimulated AR nuclear localization. SGTA protein knockdown increased AR nuclear localization at low (0-0.1 nmol/L) but not high (1-10 nmol/L) concentrations of androgen hormone. In ovarian tissues, SGTA was localized to the cytoplasm of granulosa cells at all stages of folliculogenesis and in thecal cells of antral follicles. SGTA protein levels were similar when comparing primordial and primary follicles within core biopsies (n = 40) from women with and without polycystic ovary syndrome (PCOS). Likewise, SGTA mRNA levels were not significantly different in granulosa cells from preovulatory follicles after hyperstimulation of women with and without PCOS. SGTA is present in human ovaries and has the potential to modulate AR signalling, but it may not be differentially expressed in PCOS. Copyright © 2013 American Society for Reproductive Medicine. All rights reserved.

  11. Effects of glutamine pretreatment on learning and memory in heat-exposed rats

    Shenghao Zhao; Lei Wang; Qin Wang; Siyi Wang; Chundi Deng; Xianfei Xie; Youe Yan; Hui Wang

    2008-01-01

    BACKGROUND: Glutamine (Gln) pretreatment can protect neural cells from injuries due to heat, ischemia, hypoxia, endotoxemia, and inflammatory factors.OBJECTIVE: To observe the effects of Gln pretreatment on learning and memory, survival time, and rectal temperature in heat-exposed rats.DESIGN, TIME AND SETTING: The present randomized grouping, neurobehavioral experiment was performed at the Laboratory of Department of Pharmacology, Basic School of Medicine, Wuhan University between March and September 2007.MATERIALS: Twenty-four healthy, Wistar rats were included in this study. SPX-160B biochemistry incubator (Shanghai Experimental Equipment Co., Ltd., China), probe electronic thermometer (11000 type, Maikepai Science and Technology Co., Ltd., China), Y-type maze box used in conjunction with MG-2 maze stimulator (Zhangjiagang Biomedical Instrument Factory, China), L-Gin (Batch No. 061218, 5 g/bottle, prepared into 10% aqueous solution, Amresco Company, USA) were used.METHODS: Twenty-four rats were randomly and evenly divided into 3 groups: heat-exposed, Gln low-lose, and Gln high-dose. Following learning and memory testing with the Y-maze, rats in the heat-exposed group were subjected to heat injury (40.5-41.5℃) in a biochemistry incubator. Rectal temperature was measured every 5 minutes. Thirty-five minutes after heat exposure, rats were removed and placed in the Y-type maze to test learning and memory again. Subsequently, the rats were returned to the same environment of thermal stimulation until they died. Rat survival time was recorded. Subsequent to learning and memory testing, rats in the Gln low-dose and high-dose groups received an i.p. injection of Gln (0.4 g/kg and 0.8 g/kg, respectively), and were exposed to heat injury. The remaining experimental procedures remained the same as for the heat-exposed group.MAIN OUTCOME MEASURES: Rat learning and memory, rectal temperature, and survival time in heat exposure environment.RESULTS: (1) In the Y

  12. Nutrition Modulation of Cardiotoxicity and Anticancer Efficacy Related to Doxorubicin Chemotherapy by Glutamine and ω-3 Polyunsaturated Fatty Acids

    Xue, Hongyu; Ren, Wenhua; Denkinger, Melanie; Schlotzer, Ewald; Wischmeyer, Paul E.

    2015-01-01

    Background Doxorubicin (DOX) has been one of the most effective antitumor agents against a broad spectrum of malignancies. However, DOX-induced cardiotoxicity forms the major cumulative dose-limiting factor. Glutamine and ω-3 polyunsaturated fatty acids (PUFAs) are putatively cardioprotective during various stresses and/or have potential chemosensitizing effects during cancer chemotherapy. Methods Antitumor activity and cardiotoxicity of DOX treatment were evaluated simultaneously in a MatBIII mammary adenocarcinoma tumor-bearing rat model treated with DOX (cumulative dose 12 mg/kg). Single or combined treatment of parenteral glutamine (0.35 g/kg) and ω-3 PUFAs (0.19 g/kg eicosapentaenoic acid and 0.18 g/kg docosahexaenoic acid) was administered every other day, starting 6 days before chemotherapy initiation until the end of study (day 50). Results Glutamine alone significantly prevented DOX-related deterioration of cardiac function, reduced serum cardiac troponin I levels, and diminished cardiac lipid peroxidation while not affecting tumor inhibition kinetics. Single ω-3 PUFA treatment significantly enhanced antitumor activity of DOX associated with intensified tumoral oxidative stress and enhanced tumoral DOX concentration while not potentiating cardiac dysfunction or increasing cardiac oxidative stress. Intriguingly, providing glutamine and ω-3 PUFAs together did not consistently confer a greater benefit; conversely, individual benefits on cardiotoxicity and chemosensitization were mostly attenuated or completely lost when combined. Conclusions Our data demonstrate an interesting differentiality or even dichotomy in the response of tumor and host to single parenteral glutamine and ω-3 PUFA treatments. The intriguing glutamine × ω-3 PUFA interaction observed draws into question the common assumption that there are additive benefits of combinations of nutrients that are beneficial on an individual basis. PMID:25888676

  13. Effects of L-glutamine supplementation on maternal and fetal hemodynamics in gestating ewes exposed to alcohol.

    Sawant, Onkar B; Ramadoss, Jayanth; Hankins, Gary D; Wu, Guoyao; Washburn, Shannon E

    2014-08-01

    Not much is known about effects of gestational alcohol exposure on maternal and fetal cardiovascular adaptations. This study determined whether maternal binge alcohol exposure and L-glutamine supplementation could affect maternal-fetal hemodynamics and fetal regional brain blood flow during the brain growth spurt period. Pregnant sheep were randomly assigned to one of four groups: saline control, alcohol (1.75-2.5 g/kg body weight), glutamine (100 mg/kg body weight) or alcohol + glutamine. A chronic weekend binge drinking paradigm between gestational days (GD) 99 and 115 was utilized. Fetuses were surgically instrumented on GD 117 ± 1 and studied on GD 120 ± 1. Binge alcohol exposure caused maternal acidemia, hypercapnea, and hypoxemia. Fetuses were acidemic and hypercapnic, but not hypoxemic. Alcohol exposure increased fetal mean arterial pressure, whereas fetal heart rate was unaltered. Alcohol exposure resulted in ~40 % reduction in maternal uterine artery blood flow. Labeled microsphere analyses showed that alcohol induced >2-fold increases in fetal whole brain blood flow. The elevation in fetal brain blood flow was region-specific, particularly affecting the developing cerebellum, brain stem, and olfactory bulb. Maternal L-glutamine supplementation attenuated alcohol-induced maternal hypercapnea, fetal acidemia and increases in fetal brain blood flow. L-Glutamine supplementation did not affect uterine blood flow. Collectively, alcohol exposure alters maternal and fetal acid-base balance, decreases uterine blood flow, and alters fetal regional brain blood flow. Importantly, L-glutamine supplementation mitigates alcohol-induced acid-base imbalances and alterations in fetal regional brain blood flow. Further studies are warranted to elucidate mechanisms responsible for alcohol-induced programming of maternal uterine artery and fetal circulation adaptations in pregnancy.

  14. Glutamine supplementation of parenteral nutrition does not improve intestinal permeability, nitrogen balance, or outcome in newborns and infants undergoing digestive-tract surgery: results from a double-blind, randomized, controlled trial

    E.W. Steyerberg (Ewout); F.W.J. Hazebroek (Frans); M. Mourik; G.J.J.M. Borsboom (Gerard); T. Rietveld (Trinet); J.G.M. Huijmans (Jan); D. Tibboel (Dick); M.J.I.J. Albers (Marcel)

    2005-01-01

    textabstractOBJECTIVE: To assess the effect of isocaloric isonitrogenous parenteral glutamine supplementation on intestinal permeability and nitrogen loss in newborns and infants after major digestive-tract surgery. SUMMARY BACKGROUND DATA: Glutamine supplementation in critically

  15. Suplementação enteral e parenteral com glutamina em neonatos pré-termo e com baixo peso ao nascer Enteral and parenteral supplementation with glutamine in preterm and low-birth-weight neonates

    Maria Carolina Borges

    2008-03-01

    Full Text Available A glutamina é o aminoácido livre mais abundante no sangue e no músculo esquelético, bem como é o principal substrato energético para células de elevado turnover, como enterócitos e leucócitos. Adicionalmente, a glutamina representa o principal aminoácido transferido para o feto pela placenta e, juntamente com o glutamato, constituem os aminoácidos mais abundantes no leite materno. Todavia, bebês nascidos prematuramente sofrem interrupção abrupta do fornecimento placentário de glutamina, o que acarreta em dependência exclusiva da síntese endógena ou do fornecimento exógeno deste aminoácido. Aliado a isso, neonatos pré-termo (PT e com baixo peso ao nascer (BPN, freqüentemente, recebem apenas nutrição parenteral total nas primeiras semanas de vida, a qual não contém glutamina. Cabe ainda destacar que esses bebês possuem pouca massa muscular e, portanto, seus estoques de glutamina são limitados. Uma vez que neonatos PT e com BPN estão sujeitos a intenso crescimento e a inúmeros estresses fisiológicos, é possível que a glutamina seja um nutriente condicionalmente essencial nessa fase da vida, fato que estimulou a realização de estudos com a finalidade de avaliar os possíveis benefícios clínicos da suplementação enteral e parenteral com glutamina em neonatos PT e com BPN.Glutamine is the most abundant amino acid found in the blood and skeletal muscle, and is the principal energetic substrate for cells with a high turnover, such as enterocytes and leucocytes. Furthermore, glutamine is the most important amino acid that is passed to the foetus via the placenta, and together with glutamate, is the most abundant amino acid in maternal milk. Preterm infants suffer an abrupt interruption in the supply of glutamine via the placenta, which leads to an exclusive dependence on the endogenous synthesis or the exogenous supply of this amino acid. Preterm neonates (PT as well as low-birth-weight neonates (LBW frequently

  16. Quantitative analysis of 15N-labeled positional isomers of glutamine and citrulline via electrospray ionization tandem mass spectrometry of their dansyl derivatives.

    Marini, Juan C

    2011-05-15

    The enteral metabolisms of glutamine and citrulline are intertwined because, while glutamine is one of the main fuel sources for the enterocyte, citrulline is one of its products. It has been shown that the administration of (15)N-labeled glutamine results in the incorporation of the (15)N label into citrulline, but it is not clear which of the three nitrogen groups of citrulline is actually labeled. To determine the (15)N-enrichment of the positional isomers of glutamine and citrulline, a rapid liquid chromatography/tandem mass spectrometry (LC/MS/MS) method was developed. The amino acids were analyzed as their dansyl derivatives. The product ion resulting from the loss of NH(3) from the omega carbon allows for the determination of the enrichment of the ureido (citrulline) or amido groups (glutamine). The protonated pyrrolidine (citrulline) or 5-oxopyrrolidine (glutamine) product ion contains the 2-N (amino group) and is used to determine its enrichment. The method described showed no ion suppression and a wide dynamic range ranging from 1.3 picomoles to 2 nanomoles for citrulline. Background samples and standards resulted in enrichments not different from those theoretically expected. The enrichment curves for the different glutamine and citrulline isotopomers were linear (R(2)  > 0.998) over the range of enrichments studied. The method developed provides an additional insight into the metabolism of glutamine and citrulline tracing the precursor-product relationship between these two amino acids. Copyright © 2011 John Wiley & Sons, Ltd.

  17. A randomised trial of enteral glutamine supplementation for very preterm children showed no beneficial or adverse long-term neurodevelopmental outcomes

    Twilhaar, E. Sabrina; de Kieviet, Jorrit F.; Oosterlaan, Jaap; van Elburg, Ruurd M.

    2017-01-01

    This study evaluated the long-term effects of enteral glutamine supplementation on neurodevelopmental outcomes of a Dutch cohort of very preterm children at 13 years of age. The cohort was enrolled in a randomised placebo-controlled trial between 2001-2003 in which infants received glutamine- or

  18. Some amino acids levels: glutamine,glutamate, and homocysteine, in plasma of children with chronic kidney disease.

    Fadel, Fatina I; Elshamaa, Manal F; Essam, Rascha G; Elghoroury, Eman A; El-Saeed, Gamila S M; El-Toukhy, Safinaz E; Ibrahim, Mona Hamed

    2014-03-01

    The high prevalence of protein-energy malnutrition is a critical issue for patients with chronic kidney disease (CKD). Serum albumin is the most commonly used nutritional marker. Another index is plasma amino acid (AA) profile. Of these, the plasma levels of glutamine, glutamate and homocysteine, correlate well with nutritional status. We measured some plasma AAs in children with different stages CKD to provide information in monitoring the therapeutic strategy, particularly in AA supplementary therapy or protein restriction. Three amino acids were evaluated along with albumin and high sensitivity C-reactive protein (hs-CRP) in 30 patients with advanced CKD stages 4 and 5. They were divided into two groups undergoing conservative treatment (CT) (n=15) or hemodialysis (HD) (n=15). An additional group of patients with nephrotic syndrome [CKD stage 2] was also studied to assess the alterations of plasma free amino acids with the early stage of CKD. Another 30 age- and sex-matched healthy children served as controls. A significant increase in plasma concentration of amino acid glutamine was observed in children with advanced CKD stages 4 and 5 when compared with controls (P=0.02).Plasma glutamine level was significantly higher in ESRD children on HD than in children with nephrotic syndrome (P=0.02). We did not find a significant difference between HD children and CT children as regard to glutamine level. Notable differences were in the plasma homocysteine level detected in the CKD groups patients, which was greater than that in controls (P=0.0001). Plasma homocysteine level was significantly higher in children on HD than in children with nephrotic syndrome (P=0.01). A significant differences was observed in hs-CRP levels between the CKD groups and the controls (P=0.04). Albumin levels were lower in CKD groups than in controls (p=0.01). Glutamine showed significant positive correlations with blood urea level (r=0.84, P=0.002) and blood ammonia level (r=0.72, P=0

  19. Glutamine and ornithine alpha-ketoglutarate supplementation on malate dehydrogenases expression in hepatectomized rats.

    Guimarães Filho, Artur; Cunha, Rodrigo Maranguape Silva da; Vasconcelos, Paulo Roberto Leitão de; Guimarães, Sergio Botelho

    2014-06-01

    To evaluate the relative gene expression (RGE) of cytosolic (MDH1) and mitochondrial (MDH2) malate dehydrogenases enzymes in partially hepatectomized rats after glutamine (GLN) or ornithine alpha-ketoglutarate (OKG) suplementation. One-hundred and eight male Wistar rats were randomly distributed into six groups (n=18): CCaL, GLNL and OKGL and fed calcium caseinate (CCa), GLN and OKG, 0.5 g/Kg by gavage, 30 minutes before laparotomy. CCaH, GLNH and OKGH groups were likewise fed 30 minutes before 70% partial hepatectomy. Blood and liver samples were collected three, seven and 14 days after laparotomy/hepatectomy for quantification of MDH1/MDH2 enzymes using the real-time polymerase chain reaction (PCR) methodology. Relative enzymes expression was calculated by the 2-(ΔΔC)T method using the threshold cycle (CT) value for normalization. MDH1/MDH2 RGE was not different in hepatectomized rats treated with OKG compared to rats treated with CCa. However, MDH1/MDH2 RGE was greater on days 3 (321:1/26.48:1) and 7 (2.12:1/2.48:1) while MDH2 RGE was greater on day 14 (7.79:1) in hepatectomized rats treated with GLN compared to control animals. Glutamine has beneficial effects in liver regeneration in rats by promoting an up-regulation of the MDH1 and MDH2 relative gene expression.

  20. Glutamine/glutamate (Glx) concentration in prefrontal cortex predicts reversal learning performance in the marmoset.

    Lacreuse, Agnès; Moore, Constance M; LaClair, Matthew; Payne, Laurellee; King, Jean A

    2018-07-02

    This study used Magnetic Resonance Spectroscopy (MRS) to identify potential neurometabolitic markers of cognitive performance in male (n = 7) and female (n = 8) middle-aged (∼5 years old) common marmosets (Callithrix jacchus). Anesthetized marmosets were scanned with a 4.7 T/40 cm horizontal magnet equipped with 450 mT/m magnetic field gradients and a 20 G/cm magnetic field gradient insert, within 3 months of completing the CANTAB serial Reversal Learning task. Neurometabolite concentrations of N-Acetyl Asparate, Myo-Inositol, Choline, Phosphocreatine + creatine, Glutamate and Glutamine were acquired from a 3 mm 3 voxel positioned in the Prefrontal Cortex (PFC). Males acquired the reversals (but not simple discriminations) faster than the females. Higher PFC Glx (glutamate + glutamine) concentration was associated with faster acquisition of the reversals. Interestingly, the correlation between cognitive performance and Glx was significant in males, but not in females. These results suggest that MRS is a useful tool to identify biochemical markers of cognitive performance in the healthy nonhuman primate brain and that biological sex modulates the relationship between neurochemical composition and cognition. Copyright © 2018 Elsevier B.V. All rights reserved.

  1. Proximal tubule-specific glutamine synthetase deletion alters basal and acidosis-stimulated ammonia metabolism

    Lee, Hyun-Wook; Osis, Gunars; Handlogten, Mary E.; Lamers, Wouter H.; Chaudhry, Farrukh A.; Verlander, Jill W.

    2016-01-01

    Glutamine synthetase (GS) catalyzes the recycling of NH4+ with glutamate to form glutamine. GS is highly expressed in the renal proximal tubule (PT), suggesting ammonia recycling via GS could decrease net ammoniagenesis and thereby limit ammonia available for net acid excretion. The purpose of the present study was to determine the role of PT GS in ammonia metabolism under basal conditions and during metabolic acidosis. We generated mice with PT-specific GS deletion (PT-GS-KO) using Cre-loxP techniques. Under basal conditions, PT-GS-KO increased urinary ammonia excretion significantly. Increased ammonia excretion occurred despite decreased expression of key proteins involved in renal ammonia generation. After the induction of metabolic acidosis, the ability to increase ammonia excretion was impaired significantly by PT-GS-KO. The blunted increase in ammonia excretion occurred despite greater expression of multiple components of ammonia generation, including SN1 (Slc38a3), phosphate-dependent glutaminase, phosphoenolpyruvate carboxykinase, and Na+-coupled electrogenic bicarbonate cotransporter. We conclude that 1) GS-mediated ammonia recycling in the PT contributes to both basal and acidosis-stimulated ammonia metabolism and 2) adaptive changes in other proteins involved in ammonia metabolism occur in response to PT-GS-KO and cause an underestimation of the role of PT GS expression. PMID:27009341

  2. Stereospecific assignment of the asparagine and glutamine sidechain amide protons in proteins from chemical shift analysis

    Harsch, Tobias; Schneider, Philipp; Kieninger, Bärbel; Donaubauer, Harald; Kalbitzer, Hans Robert, E-mail: hans-robert.kalbitzer@biologie.uni-regensburg.de [University of Regensburg, Institute of Biophysics and Physical Biochemistry and Centre of Magnetic Resonance in Chemistry and Biomedicine (Germany)

    2017-02-15

    Side chain amide protons of asparagine and glutamine residues in random-coil peptides are characterized by large chemical shift differences and can be stereospecifically assigned on the basis of their chemical shift values only. The bimodal chemical shift distributions stored in the biological magnetic resonance data bank (BMRB) do not allow such an assignment. However, an analysis of the BMRB shows, that a substantial part of all stored stereospecific assignments is not correct. We show here that in most cases stereospecific assignment can also be done for folded proteins using an unbiased artificial chemical shift data base (UACSB). For a separation of the chemical shifts of the two amide resonance lines with differences ≥0.40 ppm for asparagine and differences ≥0.42 ppm for glutamine, the downfield shifted resonance lines can be assigned to H{sup δ21} and H{sup ε21}, respectively, at a confidence level >95%. A classifier derived from UASCB can also be used to correct the BMRB data. The program tool AssignmentChecker implemented in AUREMOL calculates the Bayesian probability for a given stereospecific assignment and automatically corrects the assignments for a given list of chemical shifts.

  3. Glutamine synthetase in Medicago truncatula, unveiling new secrets of a very old enzyme

    Ana Rita Seabra

    2015-07-01

    Full Text Available Glutamine Synthetase (GS catalyses the first step at which nitrogen is brought into cellular metabolism and is also involved in the reassimilation of ammonium released by a number of metabolic pathways. Due to its unique position in plant nitrogen metabolism, GS plays essential roles in all aspects of plant development, from germination to senescence, and is a key component of nitrogen use efficiency (NUE and plant yield. Understanding the mechanisms regulating GS activity is therefore of utmost importance and a great effort has been dedicated to understand how GS is regulated in different plant species. The present review summarizes exciting recent developments concerning the structure and regulation of glutamine synthetase isoenzymes, using the model legume Medicago truncatula. These include the understanding of the structural determinants of both the cytosolic and plastid located isoenzymes, the existence of a seed-specific GS gene unique to M. truncatula and closely related species and the discovery that GS isoenzymes are regulated by nitric oxide at the post-translational level. The data is discussed and integrated with the potential roles of the distinct GS isoenzymes within the whole plant context.

  4. Perioperative Alanyl-Glutamine-Supplemented Parenteral Nutrition in Chronic Radiation Enteritis Patients With Surgical Intestinal Obstruction: A Prospective, Randomized, Controlled Study.

    Yao, Danhua; Zheng, Lei; Wang, Jian; Guo, Mingxiao; Yin, Jianyi; Li, Yousheng

    2016-04-01

    A prospective, randomized, controlled study was performed to evaluate the effects of perioperative alanyl-glutamine-supplemented parenteral nutrition (PN) support on the immunologic function, intestinal permeability, and nutrition status of surgical patients with chronic radiation enteritis (CRE)-induced intestinal obstruction. Patients who received 0.4 g/kg/d alanyl-glutamine and isonitrogenous PN were assigned to an alanyl-glutamine-supplemented PN (Gln-PN) group and a control group, respectively. Serum levels of alanine aminotransferase and glutamine, body fat mass (FM), immunologic function, and intestinal permeability were measured before and after surgery. Serum glutamine levels of the Gln-PN group significantly exceeded that of the control group (P nutrition state and intestinal motility of surgical patients with CRE-induced intestinal obstruction. © 2015 American Society for Parenteral and Enteral Nutrition.

  5. Mid-Season Leaf Glutamine Predicts End-Season Maize Grain Yield and Nitrogen Content in Response to Nitrogen Fertilization under Field Conditions

    Travis Goron

    2017-06-01

    Full Text Available After uptake in cereal crops, nitrogen (N is rapidly assimilated into glutamine (Gln and other amino acids for transport to sinks. Therefore Gln has potential as an improved indicator of soil N availability compared to plant N demand. Gln has primarily been assayed to understand basic plant physiology, rather than to measure plant/soil-N under field conditions. It was hypothesized that leaf Gln at early-to-mid season could report the N application rate and predict end-season grain yield in field-grown maize. A three-year maize field experiment was conducted with N application rates ranging from 30 to 218 kg ha−1. Relative leaf Gln was assayed from leaf disk tissue using a whole-cell biosensor for Gln (GlnLux at the V3-V14 growth stages. SPAD (Soil Plant Analysis Development and NDVI (Normalized Difference Vegetation Index measurements were also performed. When sampled at V6 or later, GlnLux glutamine output consistently correlated with the N application rate, end-season yield, and grain N content. Yield correlation outperformed GreenSeekerTM NDVI, and was equivalent to SPAD chlorophyll, indicating the potential for yield prediction. Additionally, depleting soil N via overplanting increased GlnLux resolution to the earlier V5 stage. The results of the study are discussed in the context of luxury N consumption, leaf N remobilization, senescence, and grain fill. The potential and challenges of leaf Gln and GlnLux for the study of crop N physiology, and future N management are also discussed.

  6. Competition among oxidizable substrates in brains of young and adult rats. Dissociated cells.

    Roeder, L M; Tildon, J T; Holman, D C

    1984-01-01

    The rates of conversion of D-(-)-3-hydroxy[3-14C]butyrate, [3-14C]acetoacetate, [6-14C]glucose and [U-14C]glutamine into 14CO2 were measured in the presence and absence of alternative oxidizable substrates in intact dissociated cells from the brains of young and adult rats. When unlabelled glutamine was added to [6-14C]glucose or unlabelled glucose was added to [U-14C]glutamine, the rate of 14CO2 production was decreased in both young and adult rats. The rate of oxidation of 3-hydroxy[3-14C]b...

  7. Longevity of guard cell chloroplasts in falling leaves: implication for stomatal function and cellular aging.

    Zeiger, E; Schwartz, A

    1982-11-12

    Guard cell chloroplasts in senescing leaves from 12 species of perennial trees and three species of annual plants survived considerably longer than their mesophyll counterparts. In Ginkgo biloba, stomata from yellow leaves opened during the day and closed at night; guard cell chloroplasts from these leaves showed fluorescence transients associated with electron transport and photophosphorylation. These findings indicate that guard cell chloroplasts are highly conserved throughout the life-span of the leaf and that leaves retain stomatal control during senescence.

  8. Dysregulation of the Glutamine Transporter Slc38a3 (SNAT3 and Ammoniagenic Enzymes in Obese, Glucose-Intolerant Mice

    Stephanie M. Busque

    2014-08-01

    Full Text Available Background/Aims: Uric acid nephrolithiasis is prevalent among patients with type 2 diabetes and metabolic syndrome; it is correlated with an acidic urine and lower urinary ammonium excretion and is likely associated with insulin resistance. Insulin stimulates ammoniagenesis in renal cell lines via increased phosphate-dependent glutaminase (PDG activity and glutamine metabolism. Ammonium excretion into the proximal tubule is mediated at least in part by the Na+/H+-exchanger NHE3 and in the collecting duct involving the Rhesus protein RhCG. Here we tested, whether obesity and insulin resistance in a diet-induced mouse model could contribute to deranged ammonium excretion. Methods: Obesity was induced by diet in mice and the impact on key molecules of proximal tubular ammoniagenesis and urinary acid excretion tested. Results: Diet-induced obesity was confirmed by pathological intraperitoneal glucose tolerance tests (IPGTT. Three groups of mice were compared: control mice; obese, glucose-intolerant with abnormal IPGTT (O-GI; or moderate weight with normal IPGTT (Non-Responders, NR. Basal urinary ammonium excretion did not differ among groups. However, acid loading increased urinary ammonium excretion in all groups, but to a lesser extent in the O-GI group. SNAT3 mRNA expression was enhanced in both obese groups. PDG expression was elevated only in acid-loaded O-GI mice, whereas PEPCK was enhanced in both O-GI and NR groups given NH4CI. NHE activity in the brush border membrane of the proximal tubule was strongly reduced in the O-GI group whereas RhCG expression was similar. Conclusion: In sum, obesity and glucose intolerance impairs renal ammonium excretion in response to NH4CI feeding most likely through reduced NHE activity. The stimulation of SNAT3 and ammoniagenic enzyme expression may be compensatory but futile.

  9. Possible role of glutamine synthetase in the NO signaling response in root nodules by contributing to the antioxidant defenses

    Liliana Santos Silva

    2013-09-01

    Full Text Available Nitric oxide (NO is emerging as an important regulatory player in the Rhizobium-legume symbiosis. The occurrence of NO during several steps of the symbiotic interaction suggests an important, but yet unknown, signaling role of this molecule for root nodule formation and functioning. The identification of the molecular targets of NO is key for the assembly of the signal transduction cascade that will ultimately help to unravel NO function. We have recently shown that the key nitrogen assimilatory enzyme Glutamine Synthetase (GS is a molecular target of NO in root nodules of Medicago truncatula, being post-translationally regulated by tyrosine nitration in relation to nitrogen fixation. In functional nodules of M. truncatula NO formation has been located in the bacteroid containing cells of the fixation zone, where the ammonium generated by bacterial nitrogenase is released to the plant cytosol and assimilated into the organic pools by plant GS. We propose that the NO-mediated GS post-translational inactivation is connected to nitrogenase inhibition induced by NO and is related to metabolite channeling to boost the nodule antioxidant defenses. Glutamate, a substrate for GS activity is also the precursor for the synthesis of glutathione (GSH, which is highly abundant in root nodules of several plant species and known to play a major role in the antioxidant defense participating in the ascorbate/GSH cycle. Existing evidence suggests that upon NO-mediated GS inhibition, glutamate could be channeled for the synthesis of GSH. According to this hypothesis, GS would be involved in the NO-signaling responses in root nodules and the NO-signaling events would meet the nodule metabolic pathways to provide an adaptive response to the inhibition of symbiotic nitrogen fixation by reactive nitrogen species (RNS.

  10. Selection of metastatic breast cancer cells based on adaptability of their metabolic state.

    Balraj Singh

    Full Text Available A small subpopulation of highly adaptable breast cancer cells within a vastly heterogeneous population drives cancer metastasis. Here we describe a function-based strategy for selecting rare cancer cells that are highly adaptable and drive malignancy. Although cancer cells are dependent on certain nutrients, e.g., glucose and glutamine, we hypothesized that the adaptable cancer cells that drive malignancy must possess an adaptable metabolic state and that such cells could be identified using a robust selection strategy. As expected, more than 99.99% of cells died upon glutamine withdrawal from the aggressive breast cancer cell line SUM149. The rare cells that survived and proliferated without glutamine were highly adaptable, as judged by additional robust adaptability assays involving prolonged cell culture without glucose or serum. We were successful in isolating rare metabolically plastic glutamine-independent (Gln-ind variants from several aggressive breast cancer cell lines that we tested. The Gln-ind cells overexpressed cyclooxygenase-2, an indicator of tumor aggressiveness, and they were able to adjust their glutaminase level to suit glutamine availability. The Gln-ind cells were anchorage-independent, resistant to chemotherapeutic drugs doxorubicin and paclitaxel, and resistant to a high concentration of a COX-2 inhibitor celecoxib. The number of cells being able to adapt to non-availability of glutamine increased upon prior selection of cells for resistance to chemotherapy drugs or resistance to celecoxib, further supporting a linkage between cellular adaptability and therapeutic resistance. Gln-ind cells showed indications of oxidative stress, and they produced cadherin11 and vimentin, indicators of mesenchymal phenotype. Gln-ind cells were more tumorigenic and more metastatic in nude mice than the parental cell line as judged by incidence and time of occurrence. As we decreased the number of cancer cells in xenografts, lung metastasis

  11. Selection of Metastatic Breast Cancer Cells Based on Adaptability of Their Metabolic State

    Singh, Balraj; Tai, Karen; Madan, Simran; Raythatha, Milan R.; Cady, Amanda M.; Braunlin, Megan; Irving, LaTashia R.; Bajaj, Ankur; Lucci, Anthony

    2012-01-01

    A small subpopulation of highly adaptable breast cancer cells within a vastly heterogeneous population drives cancer metastasis. Here we describe a function-based strategy for selecting rare cancer cells that are highly adaptable and drive malignancy. Although cancer cells are dependent on certain nutrients, e.g., glucose and glutamine, we hypothesized that the adaptable cancer cells that drive malignancy must possess an adaptable metabolic state and that such cells could be identified using a robust selection strategy. As expected, more than 99.99% of cells died upon glutamine withdrawal from the aggressive breast cancer cell line SUM149. The rare cells that survived and proliferated without glutamine were highly adaptable, as judged by additional robust adaptability assays involving prolonged cell culture without glucose or serum. We were successful in isolating rare metabolically plastic glutamine-independent (Gln-ind) variants from several aggressive breast cancer cell lines that we tested. The Gln-ind cells overexpressed cyclooxygenase-2, an indicator of tumor aggressiveness, and they were able to adjust their glutaminase level to suit glutamine availability. The Gln-ind cells were anchorage-independent, resistant to chemotherapeutic drugs doxorubicin and paclitaxel, and resistant to a high concentration of a COX-2 inhibitor celecoxib. The number of cells being able to adapt to non-availability of glutamine increased upon prior selection of cells for resistance to chemotherapy drugs or resistance to celecoxib, further supporting a linkage between cellular adaptability and therapeutic resistance. Gln-ind cells showed indications of oxidative stress, and they produced cadherin11 and vimentin, indicators of mesenchymal phenotype. Gln-ind cells were more tumorigenic and more metastatic in nude mice than the parental cell line as judged by incidence and time of occurrence. As we decreased the number of cancer cells in xenografts, lung metastasis and then primary

  12. Comparative conventional- and quantum dot-labelling strategies for LPS binding site detection in Arabidopsis thaliana mesophyll protoplasts

    Londiwe Siphephise Mgcina

    2015-05-01

    Full Text Available Lipopolysaccharide (LPS from Gram-negative bacteria is recognized as a microbe-associated molecular pattern (MAMP and not only induces an innate immune response in plants, but also stimulates the development of characteristic defense responses. However, identification and characterization of a cell surface LPS-receptor/binding site, as described in mammals, remains elusive in plants. As an amphiphilic, macromolecular lipoglycan, intact LPS potentially contains three MAMP-active regions, represented by the O-polysaccharide chain, the core and the lipid A. Binding site studies with intact labelled LPS were conducted in Arabidopsis thaliana protoplasts and quantified using flow cytometry fluorescence changes. Qdots, which allow non-covalent, hydrophobic labelling were used as a novel strategy in this study and compared to covalent, hydrophilic labelling with Alexa 488. Affinity for LPS-binding sites was clearly demonstrated by concentration-, temperature- and time-dependent increases in protoplast fluorescence following treatment with the labelled LPS. Moreover, this induced fluorescence increase was convincingly reduced following pre-treatment with excess unlabeled LPS, thereby indicating reversibility of LPS binding. Inhibition of the binding process is also reported using endo- and exocytosis inhibitors. Here, we present evidence for the anticipated presence of LPS-specific binding sites in Arabidopsis protoplasts, and furthermore propose Qdots as a more sensitive LPS-labelling strategy in comparison to the conventional Alexa 488 hydrazide label for binding studies.

  13. Oral supplementations with L-glutamine or L-alanyl-L-glutamine do not change metabolic alterations induced by long-term high-fat diet in the B6.129F2/J mouse model of insulin resistance.

    Bock, Patricia Martins; Krause, Mauricio; Schroeder, Helena Trevisan; Hahn, Gabriela Fernandes; Takahashi, Hilton Kenji; Schöler, Cinthia Maria; Nicoletti, Graziella; Neto, Luiz Domingos Zavarize; Rodrigues, Maria Inês Lavina; Bruxel, Maciel Alencar; Homem de Bittencourt, Paulo Ivo

    2016-01-01

    In this work, we aimed to investigate the effects of long-term supplementations with L-glutamine or L-alanyl-L-glutamine in the high-fat diet (HFD)-fed B6.129SF2/J mouse model over insulin sensitivity response and signaling, oxidative stress markers, metabolism and HSP70 expression. Mice were fed in a standard low-fat diet (STA) or a HFD for 20 weeks. In the 21th week, mice from the HFD group were allocated in five groups and supplemented for additional 8 weeks with different amino acids: HFD control group (HFD-Con), HFD + dipeptide L-alanyl-L-glutamine group (HFD-Dip), HFD + L-alanine group (HFD-Ala), HFD + L-glutamine group (HFD-Gln), or the HFD + L-alanine + L-glutamine (in their free forms) group (HFD-Ala + Gln). HFD induced higher body weight, fat pad, fasted glucose, and total cholesterol in comparison with STA group. Amino acid supplementations did not induce any modifications in these parameters. Although insulin tolerance tests indicated insulin resistance in all HFD groups, amino acid supplementations did not improve insulin sensitivity in the present model. There were also no significant differences in the immunocontents of insulin receptor, Akt, and Toll-like receptor-4. Notably, total 70 kDa heat shock protein (HSP72 + HSP73) contents in the liver was markedly increased in HFD-Con group as compared to STA group, which might suggest that insulin resistance is only in the beginning. Apparently, B6.129SF2/J mice are more resistant to the harmful effects of HFD through a mechanism that may include gut adaptation, reducing the absorption of nutrients, including amino acids, which may explain the lack of improvements in our intervention.

  14. Systematic replacement of lysine with glutamine and alanine in Escherichia coli malate synthase G: effect on crystallization

    Anstrom, David M.; Colip, Leslie; Moshofsky, Brian; Hatcher, Eric; Remington, S. James

    2005-01-01

    Alanine and glutamine mutations were made to the same 15 lysine positions on the surface of E. coli malate synthase G and the impact on crystallization observed. The results support lysine replacement for improvement of crystallization and provide insight into site selection and type of amino-acid replacement. Two proposals recommend substitution of surface lysine residues as a means to improve the quality of protein crystals. In proposal I, substitution of lysine by alanine has been suggested to improve crystallization by reducing the entropic cost of ordering flexible side chains at crystal contacts. In proposal II, substitution of lysine by residues more commonly found in crystal contacts, such as glutamine, has been proposed to improve crystallization. 15 lysine residues on the surface of Escherichia coli malate synthase G, distributed over a variety of secondary structures, were individually mutated to both alanine and glutamine. For 28 variants, detailed studies of the effect on enzymatic activity and crystallization were conducted. This has permitted direct comparison of the relative effects of the two types of mutations. While none of the variants produced crystals suitable for X-ray structural determination, small crystals were obtained in a wide variety of conditions, in support of the general approach. Glutamine substitutions were found to be more effective than alanine in producing crystals, in support of proposal II. Secondary structure at the site of mutation does not appear to play a major role in determining the rate of success

  15. Time dependent effects of haloperidol on glutamine and GABA homeostasis and astrocyte activity in the rat brain

    Konopaske, Glenn T.; Bolo, Nicolas R.; Basu, Alo C.; Renshaw, Perry F.; Coyle, Joseph T.

    2013-01-01

    Rationale Schizophrenia is a severe, persistent, and fairly common mental illness. Haloperidol is widely used and is effective against the symptoms of psychosis seen in schizophrenia. Chronic oral haloperidol administration decreased the number of astrocytes in the parietal cortex of macaque monkeys (Konopaske et al. Biol Psych, 2008). Since astrocytes play a key role in glutamate metabolism, chronic haloperidol administration was hypothesized to modulate astrocyte metabolic function and glutamate homeostasis. Objectives This study investigated the effects of chronic haloperidol administration on astrocyte metabolic activity and glutamate, glutamine, and GABA homeostasis. Methods We used ex vivo 13C magnetic resonance spectroscopy along with high performance liquid chromatography after [1-13C]glucose and [1,2-13C]acetate administration to analyze forebrain tissue from rats administered oral haloperidol for 1 or 6 months. Results Administration of haloperidol for 1 month produced no changes in 13C labeling of glutamate, glutamine, or GABA, or in their total levels. However, a 6 month haloperidol administration increased 13C labeling of glutamine by [1,2-13C]acetate. Moreover, total GABA levels were also increased. Haloperidol administration also increased the acetate/glucose utilization ratio for glutamine in the 6 month cohort. Conclusions Chronic haloperidol administration in rats appears to increase forebrain GABA production along with astrocyte metabolic activity. Studies exploring these processes in subjects with schizophrenia should take into account the potential confounding effects of antipsychotic medication treatment. PMID:23660600

  16. Effects of combined pulse electromagnetic field stimulation plus glutamine on the healing of colonic anastomosis in rats.

    Girgin, Sadullah; Gedik, Ercan; Ozturk, Hayrettin; Akpolat, Veysi; Akbulut, Veysi; Kale, Ebru; Buyukbayram, Huseyin; Celik, Salih

    2009-04-01

    An experimental study was designed to investigate the effect of combined pulse electromagnetic field (PEMF) stimulation plus glutamine administration on colonic anastomosis. Anastomosis of the left colon was performed in 28 rats, which were divided into four groups; Group 1: normal resection anastomosis plus oral 50 mg/kg/day glutamine; Group 2: normal resection anastomosis plus PEMF stimulation plus oral 50 mg/kg/day glutamine; Group 3: normal resection anastomosis plus PEMF stimulation; Group 4: normal resection anastomosis. On the seventh postoperative day, the animals were killed and the bursting pressure and tissue hydroxyproline concentration of the anastomosis were analyzed and compared. The mean anastomotic bursting pressure in Group 2 was significantly higher than in Groups 1 and 4. On the other hand, the mean anastomotic bursting pressure in Group 1 was significantly higher than in Group 4. The collagen deposition and the fibroblast infiltration were significantly increased on the seventh day in Group 3 compared the other groups. On the other hand, Groups 1 and 2 had higher scores for collagen deposition and fibroblast infiltration than Group 4. In conclusion, burst pressures, hydroxyproline, and histologic features (fibroblast infiltration and collagen deposition) were improved in the PEMF group, and both PEMF and glutamine-enriched nutrition provide a significant gain in the strength of colonic anastomoses in rats.

  17. Advances in animal cell recombinant protein production: GS-NS0 expression system.

    Barnes, L M; Bentley, C M; Dickson, A J

    2000-02-01

    The production of recombinant proteins using mammalian cell expression systems is of growing importance within biotechnology, largely due to the ability of specific mammalian cells to carry out post-translational modifications of the correct fidelity. The Glutamine Synthetase-NS0 system is now one such industrially important expression system.Glutamine synthetase catalyses the formation ofglutamine from glutamate and ammonia. NS0 cellscontain extremely low levels of endogenous glutaminesynthetase activity, therefore exogenous glutaminesynthetase can be used efficiently as a selectablemarker to identify successful transfectants in theabsence of glutamine in the media. In addition, theinclusion of methionine sulphoximine, an inhibitor ofglutamine synthetase activity, enables furtherselection of those clones producing relatively highlevels of transfected glutamine synthetase and henceany heterologous gene which is coupled to it. Theglutamine synthetase system technology has been usedfor research and development purposes during thisdecade and its importance is clearly demonstrated nowthat two therapeutic products produced using thissystem have reached the market place.

  18. Interactions of glutamine dipeptides with sodium dodecyl sulfate in aqueous solution measured by volume, conductivity, and fluorescence spectra

    Yan Zhenning, E-mail: yanzzn@zzu.edu.cn [Department of Chemistry, Zhengzhou University, Zhengzhou, Henan 450001 (China); Sun Ximeng; Li Weiwei; Li Yu [Department of Chemistry, Zhengzhou University, Zhengzhou, Henan 450001 (China); Wang Jianji [Department of Chemistry, Henan Normal University, Xinxiang, Henan 453007 (China)

    2011-10-15

    Highlights: > Ion-ion and ion-polar group interactions are dominant interactions. > The SDS addition and temperature increase cause a dehydration effect on dipeptides. > The addition of dipeptide in water decreases the c{sub cmc} of SDS. > Enthalpy-entropy compensation takes place during micellization. > Micelle aggregation number was decreased by addition of glutamine dipeptides. - Abstract: Densities, conductivities, and fluorescence spectra of {l_brace}sodium dodecyl sulfate (SDS) + glutamine dipeptide + water{r_brace} mixtures were measured as a function of temperature. The density data have been utilized to calculate apparent molar volumes, standard partial molar volumes (V{sub 2,{phi}}{sup o}), standard partial molar volumes of transfer from water to aqueous SDS solutions ({Delta}{sub t}V{sup o}), the hydration number, partial molar expansibility (E{sub {phi}}{sup o}), and Hepler's constant of glutamine dipeptides. The critical micellar concentration (c{sub cmc}) and the degree of counterion dissociation of SDS micelles obtained from electrical conductivity data have been estimated at various concentrations of glutamine dipeptide. Thermodynamic parameters of micellization of SDS in aqueous dipeptide solutions have been determined from c{sub cmc} values and an enthalpy-entropy compensation effect was observed for the ternary systems. The pyrene fluorescence spectra were used to study the change of micropolarity produced by the interaction of SDS with glutamine dipeptide, and the aggregation behavior of SDS. The results have been interpreted in terms of solute-solvent interactions and structural changes in the mixed solutions.

  19. Blood ammonia and glutamine as predictors of hyperammonemic crises in patients with urea cycle disorder.

    Lee, Brendan; Diaz, George A; Rhead, William; Lichter-Konecki, Uta; Feigenbaum, Annette; Berry, Susan A; Le Mons, Cindy; Bartley, James A; Longo, Nicola; Nagamani, Sandesh C; Berquist, William; Gallagher, Renata; Bartholomew, Dennis; Harding, Cary O; Korson, Mark S; McCandless, Shawn E; Smith, Wendy; Cederbaum, Stephen; Wong, Derek; Merritt, J Lawrence; Schulze, Andreas; Vockley, Jerry; Vockley, Gerard; Kronn, David; Zori, Roberto; Summar, Marshall; Milikien, Douglas A; Marino, Miguel; Coakley, Dion F; Mokhtarani, Masoud; Scharschmidt, Bruce F

    2015-07-01

    The aim of this study was to examine predictors of ammonia exposure and hyperammonemic crises in patients with urea cycle disorders. The relationships between fasting ammonia, daily ammonia exposure, and hyperammonemic crises were analyzed in >100 patients with urea cycle disorders. Fasting ammonia correlated strongly with daily ammonia exposure (r = 0.764; P 200% (P urea cycle disorder subtype, dietary protein intake, or blood urea nitrogen. Fasting glutamine correlated weakly with daily ammonia exposure assessed as 24-hour area under the curve and was not a significant predictor of hyperammonemic crisis. Fasting ammonia correlates strongly and positively with daily ammonia exposure and with the risk and rate of hyperammonemic crises, suggesting that patients with urea cycle disorder may benefit from tight ammonia control.

  20. Possible links between intestinal permeablity and food processing: a potential therapeutic niche for glutamine

    Jean Robert Rapin

    2010-01-01

    Full Text Available Increased intestinal permeability is a likely cause of various pathologies, such as allergies and metabolic or even cardiovascular disturbances. Intestinal permeability is found in many severe clinical situations and in common disorders such as irritable bowel syndrome. In these conditions, substances that are normally unable to cross the epithelial barrier gain access to the systemic circulation. To illustrate the potential harmfulness of leaky gut, we present an argument based on examples linked to protein or lipid glycation induced by modern food processing. Increased intestinal permeability should be largely improved by dietary addition of compounds, such as glutamine or curcumin, which both have the mechanistic potential to inhibit the inflammation and oxidative stress linked to tight junction opening. This brief review aims to increase physician awareness of this common, albeit largely unrecognized, pathology, which may be easily prevented or improved by means of simple nutritional changes.

  1. Inhibitory plant serpins with a sequence of three glutamine residues in the reactive center

    Hejgaard, Jørn

    2005-01-01

    Serpins appear to be ubiquitous in eukaryotes, except fungi, and are also present in some bacteria, archaea and viruses. Inhibitory serpins with a glutamine as the reactive-center P1 residue have been identified exclusively in a few plant species. Unique serpins with a reactive center sequence...... of three Gln residues at P3-P1 or P2-P1' were isolated from barley and wheat grain, respectively. Barley BSZ3 was an irreversible inhibitor of chymotrypsin, with a second-order association rate constant for complex formation k(a)' of the order of 10(4) M-1 s(-1) ; however, only a minor fraction...... of the serpin molecules reacted with chymotrypsin, with the majority insensitive to cleavage in the reactive center loop. Wheat WSZ3 was cleaved specifically at P8 Thr and was not an inhibitor of chymotrypsin. These reactive-center loops may have evolved conformations that are optimal as inhibitory baits...

  2. Possible Links between Intestinal Permeablity and Food Processing: A Potential Therapeutic Niche for Glutamine

    Rapin, Jean Robert; Wiernsperger, Nicolas

    2010-01-01

    Increased intestinal permeability is a likely cause of various pathologies, such as allergies and metabolic or even cardiovascular disturbances. Intestinal permeability is found in many severe clinical situations and in common disorders such as irritable bowel syndrome. In these conditions, substances that are normally unable to cross the epithelial barrier gain access to the systemic circulation. To illustrate the potential harmfulness of leaky gut, we present an argument based on examples linked to protein or lipid glycation induced by modern food processing. Increased intestinal permeability should be largely improved by dietary addition of compounds, such as glutamine or curcumin, which both have the mechanistic potential to inhibit the inflammation and oxidative stress linked to tight junction opening. This brief review aims to increase physician awareness of this common, albeit largely unrecognized, pathology, which may be easily prevented or improved by means of simple nutritional changes. PMID:20613941

  3. Inhibition of Glutamine Synthetase: A Potential Drug Target in Mycobacterium tuberculosis

    Sherry L. Mowbray

    2014-08-01

    Full Text Available Tuberculosis is an infectious disease caused by Mycobacterium tuberculosis. Globally, tuberculosis is second only to AIDS in mortality and the disease is responsible for over 1.3 million deaths each year. The impractically long treatment schedules (generally 6–9 months and unpleasant side effects of the current drugs often lead to poor patient compliance, which in turn has resulted in the emergence of multi-, extensively- and totally-drug resistant strains. The development of new classes of anti-tuberculosis drugs and new drug targets is of global importance, since attacking the bacterium using multiple strategies provides the best means to prevent resistance. This review presents an overview of the various strategies and compounds utilized to inhibit glutamine synthetase, a promising target for the development of drugs for TB therapy.

  4. Labeling of amino acid (L-Glutamine) with technetium-99m and biological evaluation

    Ahmed, S. S.; Abass, S. A. E.; Karim, L. M.; Risan, R. H.; Abdulkarim, H. M.; Zia, M. A.; Manjy, A.N.; Hamed, S. A.

    2012-12-01

    The study of labeling of some amino acid 9 9mT c has been carried before (1, 2). Some were found to be chemically valuable, while some others had no clinical value although, they provided US with knowledge on the mechanism of labeling and biodistribution. In this paper we experimented the possibility of labeling L-Glutamine with Tc 9 '9 m and studied its biological behavior the study revealed that the optimal pH for the formation of higher percentage of the complex is about (2.5-2.55). Biodistribution carried in balb/c mice showed that the complex accumulates in soft tissue of the abdomen. This fining may indicate good clinical value in diagnosis of soft tissue tumors of lesion. (Author)

  5. Reduced parietooccipital white matter glutamine measured by proton magnetic resonance spectroscopy in treated graves' disease patients

    Danielsen, Else Rubæk; Elberling, T.V.; Rasmussen, Åse Krogh

    2008-01-01

    CONTEXT: Graves' disease is an autoimmune disease of the thyroid gland. Patients often have affective and cognitive complaints, whether these disappear after treatment remains disputed. OBJECTIVE: Our objective was to evaluate cerebral biochemistry in acute and treated Graves' disease. DESIGN: We...... conducted a prospective study, investigating volunteers once and patients before and 1 yr after treatment. SETTING: The study was performed at a radiology department, a memory disorder clinic, and two endocrinology clinics. PATIENTS AND OTHER PARTICIPANTS: Of 53 consecutively referred, newly diagnosed...... and a battery of biochemical, affective, and cognitive tests were used. RESULTS: Previously reported findings of reduced choline and myo-inositol in acute Graves' disease were confirmed and reversibility was demonstrated. Parieto-occipital white matter glutamine was and remained significantly reduced (P

  6. Effects of Dietary Glutamine Supplementation on the Body Composition and Protein Status of Early-Weaned Mice Inoculated with Mycobacterium bovis Bacillus Calmette-Guerin

    Rogero, Marcelo Macedo; Borges, Maria Carolina; de Castro, Inar Alves; Pires, Ivanir S. O.; Borelli, Primavera; Tirapegui, Julio

    2011-01-01

    Glutamine, one of the most abundant amino acids found in maternal milk, favors protein anabolism. Early-weaned babies are deprived of this source of glutamine, in a period during which endogenous biosynthesis may be insufficient for tissue needs in states of metabolic stress, mainly during infections. The objective of this study was to verify the effects of dietary glutamine supplementation on the body composition and visceral protein status of early-weaned mice inoculated with Mycobacterium bovis Bacillus Calmette-Guérin (BCG). Mice were weaned early on their 14th day of life and seperated into two groups, one of which was fed a glutamine-free diet (n = 16) and the other a glutamine-supplemented diet (40 g/kg diet) (n = 16). At 21 days of age, some mice were intraperitoneally injected with BCG. Euthanasia was performed at the 28th day of age. BCG inoculation significantly reduced body weight (P < 0.001), lean mass (P = 0.002), water (P = 0.006), protein (P = 0.007) and lipid content (P = 0.001) in the carcass. Dietary glutamine supplementation resulted in a significant increase in serum IGF-1 (P = 0.019) and albumin (P = 0.025) concentration, muscle protein concentration (P = 0.035) and lipid content (P = 0.002) in the carcass. In conclusion, dietary glutamine supplementation had a positive influence on visceral protein status but did not affect body composition in early-weaned mice inoculated with BCG. PMID:22254124

  7. Effects of Dietary Glutamine Supplementation on the Body Composition and Protein Status of Early-Weaned Mice Inoculated with Mycobacterium bovis Bacillus Calmette-Guerin

    Ivanir S. O. Pires

    2011-08-01

    Full Text Available Glutamine, one of the most abundant amino acids found in maternal milk, favors protein anabolism. Early-weaned babies are deprived of this source of glutamine, in a period during which endogenous biosynthesis may be insufficient for tissue needs in states of metabolic stress, mainly during infections. The objective of this study was to verify the effects of dietary glutamine supplementation on the body composition and visceral protein status of early-weaned mice inoculated with Mycobacterium bovis Bacillus Calmette-Guérin (BCG. Mice were weaned early on their 14th day of life and seperated into two groups, one of which was fed a glutamine-free diet (n = 16 and the other a glutamine-supplemented diet (40 g/kg diet (n = 16. At 21 days of age, some mice were intraperitoneally injected with BCG. Euthanasia was performed at the 28th day of age. BCG inoculation significantly reduced body weight (P < 0.001, lean mass (P = 0.002, water (P = 0.006, protein (P = 0.007 and lipid content (P = 0.001 in the carcass. Dietary glutamine supplementation resulted in a significant increase in serum IGF-1 (P = 0.019 and albumin (P = 0.025 concentration, muscle protein concentration (P = 0.035 and lipid content (P = 0.002 in the carcass. In conclusion, dietary glutamine supplementation had a positive influence on visceral protein status but did not affect body composition in early-weaned mice inoculated with BCG.

  8. Vaccatides: Antifungal Glutamine-Rich Hevein-Like Peptides from Vaccaria hispanica

    Ka H. Wong

    2017-06-01

    Full Text Available Hevein and hevein-like peptides are disulfide-constrained chitin-binding cysteine-rich peptides. They are divided into three subfamilies, 6C-, 8C-, and 10C-hevein-like peptides, based on the number of cysteine residues. In addition, hevein-like peptides can exist in two forms, short and long. The long C-terminal form found in hevein and 10C-hevein-like peptides contain a C-terminal protein cargo. In contrast, the short form without a protein cargo is found in all three subfamilies. Here, we report the discovery and characterization of two novel glutamine-rich and protein cargo-free 8C-hevein-like peptides, vaccatides vH1 and vH2, from Vaccaria hispanica of the Caryophyllaceae family. Proteomic analyses showed that the vaccatides are 40–41 amino acids in length and contain a chitin-binding domain. NMR determination revealed that vaccatide vH2 displays a highly compact structure with a N-terminal cystine knot and an addition C-terminal disulfide bond. Stability studies showed that this compact structure renders vaccatide vH2 resistant to thermal, chemical and proteolytic degradation. The chitin-binding vH2 was shown to inhibit the mycelium growth of four phyto-pathogenic fungal strains with IC50 values in the micromolar range. Our findings show that vaccatides represent a new family of 8C-hevein-like peptides, which are protein cargo-free and glutamine-rich, characteristics that differentiate them from the prototypic hevein and the 10C-hevein-like peptides. In summary, this study enriches the existing library of hevein-like peptides and provides insight into their molecular diversity in sequence, structure and biosynthesis. Additionally, their highly disulfide-constrained structure could be used as a scaffold for developing metabolically and orally active peptidyl therapeutics.

  9. Glutamine-dependent carbamoyl-phosphate synthetase and other enzyme activities related to the pyrimidine pathway in spleen of Squalus acanthias (spiny dogfish).

    Anderson, P M

    1989-01-01

    The first two steps of urea synthesis in liver of marine elasmobranchs involve formation of glutamine from ammonia and of carbamoyl phosphate from glutamine, catalysed by glutamine synthetase and carbamoyl-phosphate synthetase, respectively [Anderson & Casey (1984) J. Biol. Chem. 259, 456-462]; both of these enzymes are localized exclusively in the mitochondrial matrix. The objective of this study was to establish the enzymology of carbamoyl phosphate formation and utilization for pyrimidine nucleotide biosynthesis in Squalus acanthias (spiny dogfish), a representative elasmobranch. Aspartate carbamoyltransferase could not be detected in liver of dogfish. Spleen extracts, however, had glutamine-dependent carbamoyl-phosphate synthetase, aspartate carbamoyltransferase, dihydro-orotase, and glutamine synthetase activities, all localized in the cytosol; dihydro-orotate dehydrogenase, orotate phosphoribosyltransferase, and orotidine-5'-decarboxylase activities were also present. Except for glutamine synthetase, the levels of all activities were very low. The carbamoyl-phosphate synthetase activity is inhibited by UTP and is activated by 5-phosphoribosyl 1-pyrophosphate. The first three enzyme activities of the pyrimidine pathway were eluted in distinctly different positions during gel filtration chromatography under a number of different conditions; although complete proteolysis of inter-domain regions of a multifunctional complex during extraction cannot be excluded, the evidence suggests that in dogfish, in contrast to mammalian species, these three enzymes of the pyrimidine pathway exist as individual polypeptide chains. These results: (1) establish that dogfish express two different glutamine-dependent carbamoyl-phosphate synthetase activities, (2) confirm the report [Smith, Ritter & Campbell (1987) J. Biol. Chem. 262, 198-202] that dogfish express two different glutamine synthetases, and (3) provide indirect evidence that glutamine may not be available in liver for

  10. Selective 15N labeling and direct observation by NMR of the active-site glutamine of Fe-containing superoxide dismutase

    Vance, Carrie K.; Kang, Young M.; Miller, Anne-Frances

    1997-01-01

    The glutamine in position 69 is one of only three conserved active-site amino acid differences between Fe- and Mn-containing superoxide dismutases (SODs). We have refined the conditions for extremely selective labeling of the side chains of glutamine with 15N, and thus obtained dramatically simplified spectra, despite the large size of Fe-SOD. The improved resolution afforded by such highly specific labeling permits the use of direct 15N detection to observe and assign Gln 69, even though its distance to the paramagnetic Fe2+ is only 5A. Selective glutamine side-chain labeling is inexpensive and has general utility for large (and paramagnet-containing) proteins

  11. Mesophyll cell-sucking herbivores (Cicadellidae: Typhlocybinae) on rainforest trees in New Guinea: local and regional diversity of a taxonomically unexplored guild

    Baje, L.; Stewart, A. J. A.; Novotný, Vojtěch

    2014-01-01

    Roč. 39, č. 3 (2014), s. 325-333 ISSN 0307-6946 R&D Projects: GA ČR GAP505/10/0673; GA ČR GA206/09/0115; GA MŠk(CZ) LH11008 Grant - others:National Science Foundation(US) DEB 0515678; European Social Fund(CZ) CZ.1.07/2.3.00/20.0064; UK Darwin Initiative(GB) 14/054 Institutional support: RVO:60077344 Keywords : Auchenorrhyncha * effective specialisation * food webs Subject RIV: EH - Ecology, Behaviour Impact factor: 1.699, year: 2014 http://onlinelibrary.wiley.com/doi/10.1111/een.12104/pdf

  12. Plastidic Phosphoglucose Isomerase Is an Important Determinant of Starch Accumulation in Mesophyll Cells, Growth, Photosynthetic Capacity, and Biosynthesis of Plastidic Cytokinins in Arabidopsis

    Bahaji, A.; Sanchez-Lopez, A.M.; De Diego, N.; Munoz, F.J.; Humplík, J.F.; Novák, Ondřej; Spíchal, L.; Doležal, K.; Pozueta-Romero, J.

    2015-01-01

    Roč. 10, č. 3 (2015) E-ISSN 1932-6203 R&D Projects: GA MŠk(CZ) LO1204 Institutional support: RVO:61389030 Keywords : ADP-GLUCOSE PYROPHOSPHORYLASE * PENTOSE-PHOSPHATE PATHWAY * POSTTRANSLATIONAL REDOX-MODIFICATION Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 3.057, year: 2015

  13. LncRNA TUG1 sponges miR-145 to promote cancer progression and regulate glutamine metabolism via Sirt3/GDH axis.

    Zeng, Bing; Ye, Huilin; Chen, Jianming; Cheng, Di; Cai, Canfeng; Chen, Guoxing; Chen, Xiang; Xin, Haiyang; Tang, Chaoming; Zeng, Jun

    2017-12-26

    Long noncoding RNAs (lncRNAs) are important regulators in cancer progression. Deregulation of the lncRNA taurine upregulated gene 1 (TUG1) predicts poor prognosis and is implicated in the development of several cancers. In this study, we investigated the role of TUG1 in the pathogenesis of intrahepatic cholangiocarcinoma (ICC). We found that TUG1 is upregulated in ICC samples, which correlates with poor prognosis and adverse clinical pathological characteristics. Knockdown of TUG1 inhibited the proliferation, motility, and invasiveness of cultured ICC cells, and decreased tumor burden in a xenograft mouse model. When we explored the mechanisms underlying these effects, we found that TUG1 acts as an endogenous competing RNA (ceRNA) that 'sponges' miR-145, thereby preventing the degradation of Sirt3 mRNA and increasing expression of Sirt3 and GDH proteins. Accordingly, glutamine consumption, α-KG production, and ATP levels were dramatically decreased by TUG1 knockdown in ICC cells, and this effect was reversed by miR-145 inhibition. These findings indicate that the TUG1/miR-145/Sirt3/GDH regulatory network may provide a novel therapeutic strategy for treatment of ICC.

  14. Oral glutamine increases circulating glucagon-like peptide 1, glucagon, and insulin concentrations in lean, obese, and type 2 diabetic subjects

    Greenfield, Jerry R; Farooqi, I Sadaf; Keogh, Julia M

    2008-01-01

    objective was to determine whether glutamine increases circulating GLP-1 and GIP concentrations in vivo and, if so, whether this is associated with an increase in plasma insulin. DESIGN: We recruited 8 healthy normal-weight volunteers (LEAN), 8 obese individuals with type 2 diabetes or impaired glucose...... plasma insulin concentrations. Glutamine stimulated glucagon secretion in all 3 study groups. CONCLUSION: Glutamine effectively increases circulating GLP-1, GIP, and insulin concentrations in vivo and may represent a novel therapeutic approach to stimulating insulin secretion in obesity and type 2......BACKGROUND: Incretin hormones, such as glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP), play an important role in meal-related insulin secretion. We previously demonstrated that glutamine is a potent stimulus of GLP-1 secretion in vitro. OBJECTIVE: Our...

  15. 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings

    Thakkar, Katharine N; Rösler, Lara; Wijnen, Jannie P

    2017-01-01

    BACKGROUND: The N-methyl-D-aspartate receptor hypofunction model of schizophrenia predicts dysfunction in both glutamatergic and gamma-aminobutyric acidergic (GABAergic) transmission. We addressed this hypothesis by measuring GABA, glutamate, glutamine, and the sum of glutamine plus glutamate...... concentrations in vivo in patients with schizophrenia using proton magnetic resonance spectroscopy at 7T, which allows separation of metabolites that would otherwise overlap at lower field strengths. In addition, we investigated whether altered levels of GABA, glutamate, glutamine, and the sum of glutamine plus...... glutamate reflect genetic vulnerability to schizophrenia by including healthy first-degree relatives. METHODS: Proton magnetic resonance spectroscopy at 7T was performed in 21 patients with chronic schizophrenia who were taking medication, 23 healthy first-degree relatives of patients with schizophrenia...

  16. Reduced brain glutamine in female varsity rugby athletes after concussion and in non-concussed athletes after a season of play.

    Schranz, Amy L; Manning, Kathryn Y; Dekaban, Gregory A; Fischer, Lisa; Jevremovic, Tatiana; Blackney, Kevin; Barreira, Christy; Doherty, Timothy J; Fraser, Douglas D; Brown, Arthur; Holmes, Jeff; Menon, Ravi S; Bartha, Robert

    2018-04-01

    The purpose of this study was to use non-invasive proton magnetic resonance spectroscopy (MRS) and diffusion tensor imaging (DTI) to monitor changes in prefrontal white matter metabolite levels and tissue microstructure in female rugby players with and without concussion (ages 18-23, n = 64). Evaluations including clinical tests and 3 T MRI were performed at the beginning of a season (in-season) and followed up at the end of the season (off-season). Concussed athletes were additionally evaluated 24-72 hr (n = 14), three months (n = 11), and six months (n = 8) post-concussion. Reduced glutamine at 24-72 hr and three months post-concussion, and reduced glutamine/creatine at three months post-concussion were observed. In non-concussed athletes (n = 46) both glutamine and glutamine/creatine were lower in the off-season compared to in-season. Within the MRS voxel, an increase in fractional anisotropy (FA) and decrease in radial diffusivity (RD) were also observed in the non-concussed athletes, and correlated with changes in glutamine and glutamine/creatine. Decreases in glutamine and glutamine/creatine suggest reduced oxidative metabolism. Changes in FA and RD may indicate neuroinflammation or re-myelination. The observed changes did not correlate with clinical test scores suggesting these imaging metrics may be more sensitive to brain injury and could aid in assessing recovery of brain injury from concussion. © 2017 Wiley Periodicals, Inc.

  17. Glutamine and alanine-induced differential expression of intracellular IL-6, IL-8, and TNF-α in LPS-stimulated monocytes in human whole-blood.

    Raspé, C; Czeslick, E; Weimann, A; Schinke, C; Leimert, A; Kellner, P; Simm, A; Bucher, M; Sablotzki, A

    2013-04-01

    To investigate the effects of the commonly-used immunomodulators l-glutamine, l-alanine, and the combination of both l-alanyl-l-glutamine (Dipeptamin(®)) on intracellular expression of IL-6, IL-8, and TNF-α during endotoxemia, lipopolysaccharide (LPS)-stimulated human monocytes in a whole blood system were investigated by flow cytometry. Whole blood of twenty-seven healthy volunteers was stimulated with LPS and incubated with three different amino acid solutions (1. l-glutamine, 2. l-alanine, 3. l-alanyl-l-glutamine, each concentration 2 mM, 5 mM, incubation time 3 h). CD14(+) monocytes were phenotyped in whole-blood and intracellular expression of cytokines was assessed by flow cytometry. Our investigations showed for the first time in whole blood probes, imitating best physiologically present cellular interactions, that l-glutamine caused a dose-independent inhibitory effect on IL-6 and TNF-α production in human monocytes stimulated with LPS. However, l-alanine had contrary effects on IL-6 expression, significantly upregulating expression of IL-6 in LPS-treated monocytes. The impact of l-alanine on the expression of TNF-α was comparable with glutamine. Neither amino acid was able to affect IL-8 production in LPS-stimulated monocytes. The combination of both did not influence significantly IL-6 and IL-8 expression in monocytes during endotoxemia, however strongly reduced TNF-α production. For the regulation of TNF-α, l-glutamine, l-alanine and the combination of both show a congruent and exponentiated downregulating effect during endotoxemia, for the modulation of IL-6, l-glutamine and l-alanine featured opposite regulation leading to a canceling impact of each other when recombining both amino acids. Copyright © 2013 Elsevier Ltd. All rights reserved.

  18. Effects of Glutamine on Gastric Emptying of Low- and High-Nutrient Drinks in Healthy Young Subjects—Impact on Glycaemia

    Yang T. Du

    2018-06-01

    Full Text Available Glutamine is a potent stimulus for the release of glucagon-like peptide-1, which increases postprandial insulin and slows gastric emptying (GE. We determined the effects of glutamine on GE of, and glycaemic responses to, low- and high-nutrient drinks in eight healthy males (mean age 21.6 ± 0.7 years and BMI 22.9 ± 0.7 kg/m2. Participants were studied on four occasions on which they consumed either a low-nutrient (beef soup; 18 kcal or high-nutrient (75 g dextrose; 255 kcal drink, each with or without 30 g of glutamine (120 kcal, in a randomised, crossover design. GE (2D ultrasound, blood glucose and plasma insulin concentrations were measured concurrently. Glutamine slowed GE (half emptying time (T50 of both low- (45 ± 3 min vs. 26 ± 2 min, p < 0.001, and high-nutrient, (100 ± 5 min vs. 77 ± 5 min, p = 0.03 drinks, however, there was no effect on GE of the high nutrient drinks when expressed as kcal/min (3.39 ± 0.21 kcal/min vs. 3.81 ± 0.20 kcal/min, p = 0.25. There was no change in blood glucose after the low-nutrient drinks with or without glutamine, despite a slight increase in plasma insulin with glutamine (p = 0.007. The rise in blood glucose following the high-nutrient drink (p = 0.0001 was attenuated during the first 60 min by glutamine (p = 0.007. We conclude that in healthy subjects, glutamine slows GE of both low- and high-nutrient drinks comparably and attenuates the rise in blood glucose after the high-nutrient glucose drink.

  19. Contribution of extracellular glutamine as an anaplerotic substrate to neuronal metabolism: a re-evaluation by multinuclear NMR spectroscopy in primary cultured neurons.

    Shokati, Touraj; Zwingmann, Claudia; Leibfritz, Dieter

    2005-10-01

    Multinuclear NMR spectroscopy is used to investigate the effect of glutamine on neuronal glucose metabolism. Primary neurons were incubated with [1-(13C)]glucose in the absence or presence of glutamine (2 mM) and/or NH4Cl (5 mM). After ammonia-treatment, the concentrations of high-energy phosphates decreased up to 84% of control, which was aggravated in glutamine-containing medium (up to 42% of control). These effects could not be attributed to changes in mitochondrial glucose oxidation. Withdrawal of glutamine decreased amino acid concentrations, e.g. of glutamate to 53%, but also considerably lessened the 13C enrichment in [4-(13C)]glutamate to 8.3% of control, and decreased the 13C-enrichment in acetyl-CoA entering the Krebs cycle (P neuronal glutamate stores, glutamate formation is mainly attributed to its de novo synthesis from glucose. Furthermore, mitochondrial glucose metabolism strongly depends on the supply of carbons from glutamine, indicating that exogenous glutamine is a well-suited substrate to replenish neuronal Krebs cycle intermediates.

  20. 15N-enrichments of ammonia and glutamine in blood after infusion of 15N-ammonia in chickens fed low or high protein diet

    Karasawa, Yutaka; Koh, Katsuki

    1985-01-01

    In this experment, the blood ammonia and glutamine amide came from infused ammonia were determined when N-15 labeled ammonium acetate was intraportally infused into the chickens fed 5 or 20 % protein diet. The data obtained indicated that the infused ammonia was taken into blood glutamine amide, and also accumulated in blood as it is, in both dietary groups. 10 to 12 months old White Leghorn male birds were used. The experimental diet was fed once a day for 5 days to the birds weighting about 1.2 kg by 35 g per kg body weight. The experimental diet was consumed within 40 min in all cases. Cardiac and portal catheterization were performed for blood collection and ammonia infusion, respectively. After finishing the infusion, blood samples were taken to analyze the ammonia and glutamine contents and their N-15 enrichment. Statistical difference was not observed in the appearance of N-15 in ammonia and glutamine amide between two dietary groups. The N-15 enrichment in blood ammonia and the amide of plasma glutamine, and the calculated exogenous nitrogen in the ammonia and glutamine amide tended to be more in the 5 % protein diet group than the other. (Kako, I.)

  1. Inhibition of glutamine synthesis induces glutamate dehydrogenase-dependent ammonia fixation into alanine in co-cultures of astrocytes and neurons.

    Dadsetan, Sherry; Bak, Lasse K; Sørensen, Michael; Keiding, Susanne; Vilstrup, Hendrik; Ott, Peter; Leke, Renata; Schousboe, Arne; Waagepetersen, Helle S

    2011-09-01

    It has been previously demonstrated that ammonia exposure of neurons and astrocytes in co-culture leads to net synthesis not only of glutamine but also of alanine. The latter process involves the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT). In the present study it was investigated if the glutamine synthetase (GS) inhibitor methionine sulfoximine (MSO) would enhance alanine synthesis by blocking the GS-dependent ammonia scavenging process. Hence, co-cultures of neurons and astrocytes were incubated for 2.5h with [U-(13)C]glucose to monitor de novo synthesis of alanine and glutamine in the absence and presence of 5.0 mM NH(4)Cl and 10 mM MSO. Ammonia exposure led to increased incorporation of label but not to a significant increase in the amount of these amino acids. However, in the presence of MSO, glutamine synthesis was blocked and synthesis of alanine increased leading to an elevated content intra- as well as extracellularly of this amino acid. Treatment with MSO led to a dramatic decrease in glutamine content and increased the intracellular contents of glutamate and aspartate. The large increase in alanine during exposure to MSO underlines the importance of the GDH and ALAT biosynthetic pathway for ammonia fixation, and it points to the use of a GS inhibitor to ameliorate the brain toxicity and edema induced by hyperammonemia, events likely related to glutamine synthesis. Copyright © 2011 Elsevier Ltd. All rights reserved.

  2. Interactions of MK-801 with glutamate-, glutamine- and methamphetamine-evoked release of [3H]dopamine from striatal slices

    Bowyer, J.F.; Scallet, A.C.; Holson, R.R.; Lipe, G.W.; Slikker, W. Jr.; Ali, S.F.

    1991-01-01

    The interactions of MK-801 [(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d] cyclohepten-5,10-imine], glutamate and glutamine with methamphetamine (METH)-evoked release of [ 3 H]dopamine were assessed in vitro to determine whether MK-801 inhibition of METH neurotoxicity might be mediated presynaptically, and to evaluate the effects of glutamatergic stimulation on METH-evoked dopamine release. MK-801 inhibition of glutamate- or METH-evoked dopamine release might reduce synaptic dopamine levels during METH exposure and decrease the formation of 6-hydroxydopamine or other related neurotoxins. Without Mg 2+ present, 40 microM and 1 mM glutamate evoked a N-methyl-D-aspartate receptor-mediated [ 3 H]dopamine and [ 3 H]metabolite (tritium) release of 3 to 6 and 12 to 16% of total tritium stores, respectively, from striatal slices. With 1.50 mM Mg 2+ present, 10 mM glutamate alone or in combination with the dopamine uptake blocker nomifensine released only 2.1 or 4.2%, respectively, of total tritium stores, and release was only partially dependent on N-methyl-D-aspartate-type glutamate receptors. With or without 1.50 mM Mg 2+ present, 0.5 or 5 microM METH evoked a substantial release of tritium (5-8 or 12-21% of total stores, respectively). METH-evoked dopamine release was not affected by 5 microM MK-801 but METH-evoked release was additive with glutamate-evoked release. Without Mg 2+ present, 1 mM glutamine increased glutamate release and induced the release of [ 3 H]dopamine and metabolites. Both 0.5 and 5 microM METH also increased tritium release with 1 mM glutamine present. When striatal slices were exposed to 5 microM METH this glutamine-evoked release of glutamate was increased more than 50%

  3. Supplementation of diets for weaned piglets withL-Valine and L-Glutamine+ L-Glutamic acid

    Naiara Diedrich Rodrigues

    2015-10-01

    Full Text Available The objective of this study was to evaluate the effects of supplementation of diets for weaned piglets with L-valine and L-glutamine + L-glutamic acid on performance, frequency of diarrhea, organ weight, digesta pH, intestinal morphology, and economic viability. Seventy-two piglets with a live weight of 7.53 ± 0.84 kg and 24 days of age were used. The animals were submitted to the following four treatments from 24 to 46 days of age: diet not supplemented with amino acids (control diet, CD; diet supplemented with glutamine + glutamic acid (GD; diet supplemented with glutamine + glutamic acid + valine (GVD, and diet supplemented with valine (VD. Two sequential phases (pre-initial I and pre-initial II with a duration of 12 and 11 days, respectively, were established. A completely randomized design, consisting of six repetitions and three pigs per experimental unit, was used. Nine days after weaning, at 32 days of age, a piglet per pen was slaughtered for the evaluation of organ weight, digesta pH and intestinal morphology. All animals received a single diet from days 47 to 65. No effects on performance were observed during the pre-initial phases I and II; however, when the whole study period was considered (24 to 65 days of age, piglets fed GVD consumed less feed and exhibited better feed conversion than animals of the VD group. With respect to morphometric parameters, GD provided a greater ileal crypt depth than CD and VD. There was an economic advantage of diets supplemented with L-valine and L-glutamine + L-glutamic acid, validating their use in diets for weaned piglets until 46 days of age.

  4. Crystal structures of mammalian glutamine synthetases illustrate substrate-induced conformational changes and provide opportunities for drug and herbicide design.

    Krajewski, Wojciech W; Collins, Ruairi; Holmberg-Schiavone, Lovisa; Jones, T Alwyn; Karlberg, Tobias; Mowbray, Sherry L

    2008-01-04

    Glutamine synthetase (GS) catalyzes the ligation of glutamate and ammonia to form glutamine, with concomitant hydrolysis of ATP. In mammals, the activity eliminates cytotoxic ammonia, at the same time converting neurotoxic glutamate to harmless glutamine; there are a number of links between changes in GS activity and neurodegenerative disorders, such as Alzheimer's disease. In plants, because of its importance in the assimilation and re-assimilation of ammonia, the enzyme is a target of some herbicides. GS is also a central component of bacterial nitrogen metabolism and a potential drug target. Previous studies had investigated the structures of bacterial and plant GSs. In the present publication, we report the first structures of mammalian GSs. The apo form of the canine enzyme was solved by molecular replacement and refined at a resolution of 3 A. Two structures of human glutamine synthetase represent complexes with: a) phosphate, ADP, and manganese, and b) a phosphorylated form of the inhibitor methionine sulfoximine, ADP and manganese; these structures were refined to resolutions of 2.05 A and 2.6 A, respectively. Loop movements near the active site generate more closed forms of the eukaryotic enzymes when substrates are bound; the largest changes are associated with the binding of the nucleotide. Comparisons with earlier structures provide a basis for the design of drugs that are specifically directed at either human or bacterial enzymes. The site of binding the amino acid substrate is highly conserved in bacterial and eukaryotic GSs, whereas the nucleotide binding site varies to a much larger degree. Thus, the latter site offers the best target for specific drug design. Differences between mammalian and plant enzymes are much more subtle, suggesting that herbicides targeting GS must be designed with caution.

  5. The Effect of Single Portion Glutamine Supplement Consumption on Injury Indices of Muscle After Eccentric Resistance Exercise

    Azadeh Najarzadeh; Hadi Atarod; Hasan Mozaffari-Khosravi; Ali Dehghani; Foad Asjodi

    2015-01-01

    Abstract Background: Delayed muscular soreness after resistance exercises or eccentric trainings is probably because of muscle damage and injury. Nutrition by playing a crucial role in both protein synthesize and catabolism can influence the extent of muscle injury. The objective of this study was to assess the effect of single portion of Glutamine supplement consumption on injury indices of muscle after a session eccentric resistance exercise. Materials and Methods: this study used a ...

  6. The Effect of Single Portion Glutamine Supplement Consumption on Injury Indices of Muscle After Eccentric Resistance Exercise

    Azadeh Najarzadeh

    2015-07-01

    Full Text Available Abstract Background: Delayed muscular soreness after resistance exercises or eccentric trainings is probably because of muscle damage and injury. Nutrition by playing a crucial role in both protein synthesize and catabolism can influence the extent of muscle injury. The objective of this study was to assess the effect of single portion of Glutamine supplement consumption on injury indices of muscle after a session eccentric resistance exercise. Materials and Methods: this study used a randomized, double blind design that consisted of 80 volvnteer non-athletic males (aged 22.2±2.2years, height 175±5 cm, weight 71/64±9 kg, body mass index 23/2±2/2 kg/m2, and body fat 17/5±2/4%. A total of 40 participants were divided randomly into 2 groups, supplement group (receiving 0/1 g/kg Body weight/ day Glutamine and placebo group (receiving 0/1 g/kg Body weight/ day Maltodextrin. Serum keratine kinase (CK was determined by photometric method, muscle pain and knee joint range of motion were measured using, respectively, a standard scale of PAS and goniometer before, 24 and 48 hours after a resistance test involving knee flexion. Results: Glutamine supplement consumption caused no significant differences in CK levels reduction in none of the measured times, but it reduced the muscle pain at the times of 24 and 48 hours in comparison with the placebo group. In addition, the knee joint range of motion was significantly improved at 24 hours after the test. Conclusion: It seems that this dose of Glutamine supplementation can reduce the apparent signs apart from muscle injury indices reduction.

  7. Enzymatic-fluorometric analyses for glutamine, glutamate and free amino groups in protein-free plasma and milk

    Larsen, Torben; Fernández, Carlos J.

    2017-01-01

    This Technical Research Communication describes new analytical methods for free, unbound glutamic acid and glutamine in protein-free blood plasma and milk and introduces the use of quantitation of free amino groups in the same matrices for descriptive and analytical purposes. The present enzymatic......-fluorometric methods are easily performed within one working day, allowing for ‘high throughput’ assays of animal trials. These assays could support and enable further studies in lactation physiology with the objective of improved metabolic health....

  8. Intravenous Glutamine Administration Modulates TNF-α/IL-10 Ratio and Attenuates NFkB Phosphorylation in a Protein Malnutrition Model.

    Santos, Andressa Cristina Antunes; Correia, Carolina Argondizo; de Oliveira, Dalila Cunha; Nogueira-Pedro, Amanda; Borelli, Primavera; Fock, Ricardo Ambrosio

    2016-12-01

    Protein malnutrition (PM) is a major public health problem in developing countries, affecting the inflammatory response and increasing susceptibility to opportunistic infections. For this reason, an adequate nutritional intervention can improve the quality of life of patients. Glutamine (GLN) is a nonessential amino acid, but can be considered "conditionally essential" for macrophage function in stress situations, in which it plays a role in the improvement of the inflammatory response. Concerning this issue, in the current study, it was of interest to evaluate some biological aspects of peritoneal cells from a protein malnutrition (PM) mouse model challenged with lipopolysaccharide (LPS) and treated intravenously with GLN. Two-month-old male Balb/c mice were subjected to a low-protein diet (2 % protein) and stimulated intravenously with LPS 1 h prior to the injection of 0.75 mg/kg GLN. Malnourished animals showed a reduced number of total peritoneal cells. Malnourished animals stimulated with LPS or LPS plus GLN did not show differences in peritoneal cell counts; however, the control group showed increased cellularity after LPS stimulus, which was reversed after GLN injection. Further, in the animals from both groups stimulated with LPS, GLN decreased the circulating levels of TNF-α and the levels of TNF-α produced by peritoneal cells; additionally, GLN decreased the IL-10 circulating levels in the malnourished animals stimulated with LPS. In addition, peritoneal cells of the control and malnourished groups stimulated with LPS showed a negative modulation of the NFkB signaling pathway after GLN injection. In conclusion, this study shows that GLN has the capacity to reduce TNF-α synthesis as well as to act as a negative regulator of NFkB phosphorylation, leading to a positive outcome in the control of TNF-α production.

  9. Proton extrusion is an essential signalling component in the HR of epidermal single cells in the barley-powdery mildew interaction

    Zhou, F.S.; Andersen, C.H.; Burhenne, K.

    2000-01-01

    We propose a model for activation of the epidermal cell hypersensitive response (HR) in the barley/powdery mildew interaction. The model suggests that the plasma membrane proton pump (H+-ATPase) of epidermal cells is activated following penetration by an avirulent powdery mildew fungus...... in the incompatible interaction; (4) race-specific proton extrusion is observed underneath epidermal tissue detached from leaves inoculated 15 h earlier; and (5) treatment of leaves with fusicoccin, an activator of the plasma membrane H+-ATPase, increases the number of HR-cells in the compatible interaction........ This will cause an acidification of the apoplast towards the mesophyll cells, thereby activating generation of H2O2 from the mesophyll, which subsequently triggers the epidermal cell to undergo HR. The model is supported by the following data: (1) the earliest HR-related H2O2 is found in the attachment zones...

  10. Evaluating Factor XIII Specificity for Glutamine-Containing Substrates Using a MALDI-TOF Mass Spectrometry Assay

    Doiphode, Prakash G.; Malovichko, Marina V.; Mouapi, Kelly Njine; Maurer, Muriel C.

    2014-01-01

    Activated Factor XIII (FXIIIa) catalyzes the formation of γ-glutamyl-ε-lysyl cross-links within the fibrin blood clot network. Although several cross-linking targets have been identified, the characteristic features that define FXIIIa substrate specificity are not well understood. To learn more about how FXIIIa selects its targets, a matrix-assisted laser desorption ionization – time of flight mass spectrometry (MALDI-TOF MS) based assay was developed that could directly follow the consumption of a glutamine-containing substrate and the formation of a cross-linked product with glycine ethylester. This FXIIIa kinetics assay is no longer reliant on a secondary coupled reaction, on substrate labeling, or on detecting the final deacylation portion of the transglutaminase reaction. With the MALDI-TOF MS assay, glutamine-containing peptides derived from α2-antiplasmin, S. Aureus fibronectin binding protein A, and thrombin activatable fibrinolysis inhibitor were examined directly. Results suggest that the FXIIIa active site surface responds to changes in substrate residues following the reactive glutamine. The P-1 substrate position is sensitive to charge character and the P-2 and P-3 to the broad FXIIIa substrate specificity pockets. The more distant P-8 to P-11 region serves as a secondary substrate anchoring point. New knowledge on FXIIIa specificity may be used to design better substrates or inhibitors of this transglutaminase. PMID:24751466

  11. Mitochondrial citrulline synthesis from ammonia and glutamine in the liver of ureogenic air-breathing catfish, Clarias batrachus (Linnaeus).

    Kharbuli, Zaiba Y; Biswas, Kuheli; Saha, Nirmalendu

    2007-12-01

    The possible synthesis of citrulline, a rate limiting step for urea synthesis via the ornithine-urea cycle (OUC) in teleosts was tested both in the presence of ammonia and glutamine as nitrogen-donating substrates by the isolated liver mitochondria of ureogenic air-breathing walking catfish, C. batrachus. Both ammonia and glutamine could be used as nitrogen-donating substrates for the synthesis of citrulline by the isolated liver mitochondria, since the rate of citrulline synthesis was almost equal in presence of both the substrates. The citrulline synthesis by the isolated liver mitochondria requires succinate at a concentration of 0.1 mM as an energy source, and also requires the involvement of intramitochondrial carbonic anhydrase activity for supplying HCO3 as another substrate for citrulline synthesis. The rate of citrulline synthesis was further stimulated significantly by the isolated liver mitochondria of the fish after pre-exposure to 25 mM NH4Cl for 7 days. Due to possessing this biochemical adaptational strategy leading to the amelioration of ammonia toxicity mainly by channeling ammonia directly and/or via the formation of glutamine to the OUC, this air-breathing catfish could succeed in surviving in high external ammonia, which it faces in its natural habitat in certain seasons of the year.

  12. Plant nutritional status modulates glutamine synthetase levels in ripe tomatoes (Solanum lycopersicum cv. Micro-Tom).

    Scarpeci, Telma E; Marro, Martin L; Bortolotti, Santiago; Boggio, Silvana B; Valle, Estela M

    2007-02-01

    Tomato (Solanum lycopersicum) fruit ripening implies that chloroplastic proteins are degraded and new proteins are synthesized. Supplementary nutrition is frequently required when tomato plants begin to fruit and continues until the end of the plant's life cycle. Ammonium assimilation is crucial in these fruit maturation and ripening processes. Glutamine synthetase (GS; EC 6.3.1.2), the main ammonium-fixing enzyme in plants, could not be detected in red fruits of several tomato varieties when growing under standard nutrition. In this paper, we analyze the influence of the nutritional status on the ammonium assimilation capacity of ripe tomato (cv. Micro-Tom) fruit. For this purpose, GS expression and protein profiles were followed in mature green and red fruits harvested from plants grown under standard or supplemented nutrition. Under standard nutrient regime (weekly supplied with 0.5 x Hoagland solution) GS activity was found in chloroplasts (GS2) of mature green fruits, but it was not detected either in the chromoplasts or in the cytosol of red fruits. When plants were shifted to a supplemented nutritional regime (daily supplied with 0.5 x Hoagland solution), GS was found in red fruits. Also, cytosolic transcripts (gs1) preferentially accumulated in red fruits under high nutrition. These results indicate that mature green Micro-Tom fruits assimilate ammonia through GS2 under standard nutrition, while ripe red fruits accumulate GS1 under high nutrition, probably in order to assimilate the extra N-compounds made available through supplemented nutrition.

  13. Glutamine synthetase activity and glutamate uptake in hippocampus and frontal cortex in portal hypertensive rats

    Acosta, Gabriela Beatriz; Fernández, María Alejandra; Roselló, Diego Martín; Tomaro, María Luján; Balestrasse, Karina; Lemberg, Abraham

    2009-01-01

    AIM: To study glutamine synthetase (GS) activity and glutamate uptake in the hippocampus and frontal cortex (FC) from rats with prehepatic portal vein hypertension. METHODS: Male Wistar rats were divided into sham-operated group and a portal hypertension (PH) group with a regulated stricture of the portal vein. Animals were sacrificed by decapitation 14 d after portal vein stricture. GS activity was determined in the hippocampus and FC. Specific uptake of radiolabeled L-glutamate was studied using synaptosome-enriched fractions that were freshly prepared from both brain areas. RESULTS: We observed that the activity of GS increased in the hippocampus of PH rats, as compared to control animals, and decreased in the FC. A significant decrease in glutamate uptake was found in both brain areas, and was more marked in the hippocampus. The decrease in glutamate uptake might have been caused by a deficient transport function, significantly and persistent increase in this excitatory neurotransmitter activity. CONCLUSION: The presence of moderate ammonia blood levels may add to the toxicity of excitotoxic glutamate in the brain, which causes alterations in brain function. Portal vein stricture that causes portal hypertension modifies the normal function in some brain regions. PMID:19533812

  14. Disrupted glutamate-glutamine cycle in acute encephalopathy with biphasic seizures and late reduced diffusion

    Takanashi, Jun-ichi; Terai, Masaru [Tokyo Women' s Medical University Yachiyo Medical Center, Department of Pediatrics, Yachiyo-shi (Japan); Mizuguchi, Masashi [The University of Tokyo, Department of Developmental Medical Sciences, Graduate School of Medicine, Tokyo (Japan); Barkovich, A.J. [University of California San Francisco, Department of Radiology and Biomedical Imaging, San Francisco, CA (United States)

    2015-11-15

    Acute encephalopathy with biphasic seizures and late reduced diffusion (AESD) is the most common subtype of infectious pediatric encephalopathy in Japan. It is sometimes difficult to make an early diagnosis of AESD; excitotoxicity is postulated to be the pathogenesis based on elevated glutamine (Gln) and glutamate (Glu) complex (Glx = Glu + Gln) observed on MR spectroscopy. It is uncertain whether Gln or Glu contributes to the elevated Glx, or whether MR spectroscopy is useful for an early diagnosis. Five Japanese patients with AESD (three boys and two girls, 1 year of age) were enrolled in this study. MR spectroscopy was acquired from the frontal white matter (repetition time (TR) of 5000 ms, echo time (TE) of 30 ms) with a 1.5- or 3.0-T scanner. MR spectroscopy was performed four times for two patients, three times for one patient, and two times for two patients. Quantification of Glu and Gln was performed using LCModel. Glu was elevated in three of four studies on days 1-4 and became normal or low afterward. Gln was normal in three studies on days 1-2, elevated in all seven studies on days 4-12, and became normal or low afterward. These findings suggest that MR spectroscopy may be useful for an early diagnosis. Acute Glu elevation changes to subacute Gln elevation, suggesting that a disrupted Glu-Gln cycle may play an important role. (orig.)

  15. Sleep spindles are related to schizotypal personality traits and thalamic glutamine/glutamate in healthy subjects.

    Lustenberger, Caroline; O'Gorman, Ruth L; Pugin, Fiona; Tüshaus, Laura; Wehrle, Flavia; Achermann, Peter; Huber, Reto

    2015-03-01

    Schizophrenia is a severe mental disorder affecting approximately 1% of the worldwide population. Yet, schizophrenia-like experiences (schizotypy) are very common in the healthy population, indicating a continuum between normal mental functioning and the psychosis found in schizophrenic patients. A continuum between schizotypy and schizophrenia would be supported if they share the same neurobiological origin. Two such neurobiological markers of schizophrenia are: (1) a reduction of sleep spindles (12-15 Hz oscillations during nonrapid eye movement sleep), likely reflecting deficits in thalamo-cortical circuits and (2) increased glutamine and glutamate (Glx) levels in the thalamus. Thus, this study aimed to investigate whether sleep spindles and Glx levels are related to schizotypal personality traits in healthy subjects. Twenty young male subjects underwent 2 all-night sleep electroencephalography recordings (128 electrodes). Sleep spindles were detected automatically. After those 2 nights, thalamic Glx levels were measured by magnetic resonance spectroscopy. Subjects completed a magical ideation scale to assess schizotypy. Sleep spindle density was negatively correlated with magical ideation (r = -.64, P .1). The common relationship of sleep spindle density with schizotypy and thalamic Glx levels indicates a neurobiological overlap between nonclinical schizotypy and schizophrenia. Thus, sleep spindle density and magical ideation may reflect the anatomy and efficiency of the thalamo-cortical system that shows pronounced impairment in patients with schizophrenia. © The Author 2014. Published by Oxford University Press on behalf of the Maryland Psychiatric Research Center. All rights reserved. For permissions, please email: journals.permissions@oup.com.

  16. Growth hormone increases and maturation decreases glutamine synthetase turnover rate in rat liver

    Lin, C.K.

    1985-01-01

    An investigation was made of the effect of hypophysectomy and growth hormone (GH) replacement regimen (1 mg/100 g twice daily for 30 days); and maturation (from 25 up to 90 days) on the liver and brain glutamine synthetase (GS) mass and turnover rates in rats. The first order decay rate of enzyme 14 C radioactivity was determined between 1 and 4 days to obtain the half-life (T/sub 1/2/) of GS. The hepatic GS mass was determined by immunoassay. GS turnover (GS/sub s/) was calculated from T/sub 1/2/ and the GS mass (i.e., K = 0.693/T/sub 1/2/; GS/sub s/ = K x GS mass). It was concluded that: (1) GS specific activity is not decreased by hypophysectomy or increased by GH. These results suggested that observed endocrine induced changes in GS are due to changes in GS mass. (2) The liver GS turnover rate is significantly reduced by hypophysectomy and increased by GH replacement. It was proposed that GH specifically enhances synthesis of GS in the liver. (3) Maturation (25, 40, 60, and 90 days) decreases GS turnover rate in both liver and brain of normal rats. This similar effect of maturation suggests that the observed age induced decline in GS turnover rate is not related to GH in all tissues

  17. Kynurenine aminotransferase III and glutamine transaminase L are identical enzymes that have cysteine S-conjugate β-lyase activity and can transaminate L-selenomethionine.

    Pinto, John T; Krasnikov, Boris F; Alcutt, Steven; Jones, Melanie E; Dorai, Thambi; Villar, Maria T; Artigues, Antonio; Li, Jianyong; Cooper, Arthur J L

    2014-11-07

    Three of the four kynurenine aminotransferases (KAT I, II, and IV) that synthesize kynurenic acid, a neuromodulator, are identical to glutamine transaminase K (GTK), α-aminoadipate aminotransferase, and mitochondrial aspartate aminotransferase, respectively. GTK/KAT I and aspartate aminotransferase/KAT IV possess cysteine S-conjugate β-lyase activity. The gene for the former enzyme, GTK/KAT I, is listed in mammalian genome data banks as CCBL1 (cysteine conjugate beta-lyase 1). Also listed, despite the fact that no β-lyase activity has been assigned to the encoded protein in the genome data bank, is a CCBL2 (synonym KAT III). We show that human KAT III/CCBL2 possesses cysteine S-conjugate β-lyase activity, as does mouse KAT II. Thus, depending on the nature of the substrate, all four KATs possess cysteine S-conjugate β-lyase activity. These present studies show that KAT III and glutamine transaminase L are identical enzymes. This report also shows that KAT I, II, and III differ in their ability to transaminate methyl-L-selenocysteine (MSC) and L-selenomethionine (SM) to β-methylselenopyruvate (MSP) and α-ketomethylselenobutyrate, respectively. Previous studies have identified these seleno-α-keto acids as potent histone deacetylase inhibitors. Methylselenol (CH3SeH), also purported to have chemopreventive properties, is the γ-elimination product of SM and the β-elimination product of MSC catalyzed by cystathionine γ-lyase (γ-cystathionase). KAT I, II, and III, in part, can catalyze β-elimination reactions with MSC generating CH3SeH. Thus, the anticancer efficacy of MSC and SM will depend, in part, on the endogenous expression of various KAT enzymes and cystathionine γ-lyase present in target tissue coupled with the ability of cells to synthesize in situ either CH3SeH and/or seleno-keto acid metabolites. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.

  18. Chloroplast behaviour and interactions with other organelles in Arabidopsis thaliana pavement cells.

    Barton, Kiah A; Wozny, Michael R; Mathur, Neeta; Jaipargas, Erica-Ashley; Mathur, Jaideep

    2018-01-29

    Chloroplasts are a characteristic feature of green plants. Mesophyll cells possess the majority of chloroplasts and it is widely believed that, with the exception of guard cells, the epidermal layer in most higher plants does not contain chloroplasts. However, recent observations on Arabidopsis thaliana have shown a population of chloroplasts in pavement cells that are smaller than mesophyll chloroplasts and have a high stroma to grana ratio. Here, using stable transgenic lines expressing fluorescent proteins targeted to the plastid stroma, plasma membrane, endoplasmic reticulum, tonoplast, nucleus, mitochondria, peroxisomes, F-actin and microtubules, we characterize the spatiotemporal relationships between the pavement cell chloroplasts (PCCs) and their subcellular environment. Observations on the PCCs suggest a source-sink relationship between the epidermal and the mesophyll layers, and experiments with the Arabidopsis mutants glabra2 ( gl2 ) and immutans ( im ), which show altered epidermal plastid development, underscored their developmental plasticity. Our findings lay down the foundation for further investigations aimed at understanding the precise role and contributions of PCCs in plant interactions with the environment. © 2018. Published by The Company of Biologists Ltd.

  19. The role of disease-linked residue glutamine-913 in support of the structure and function of the human electrogenic sodium/bicarbonate cotransporter NBCe1-A.

    Myers, Evan J; Marshall, Aniko; Parker, Mark D

    2018-02-15

    Mutations in the sodium bicarbonate cotransporter NBCe1 (SLC4A4) cause proximal renal tubular acidosis (pRTA). We recently described a novel pRTA mutation p.Gln913Arg (Q913R), inherited in compound heterozygous form with p.Arg510His (R510H). Q913R causes intracellular retention of NBCe1 and a 'gain of function' Cl - leak. To learn more about the importance of glutamine at position 913, we substituted a variety of alternative amino-acid residues (Cys, Glu, Lys, Leu, Ser) at position 913. Studying cRNA-injected Xenopus oocytes by voltage clamp, we find that most de novo mutants exhibit close-to-normal NBCe1 activity; only Q913K expresses a Cl - leak. Studying transiently-transfected, polarised kidney cells by fluorescence microscopy we find that most de novo mutants (except Q913E) are intracellularly retained. A 3D homology model predicts that Gln913 is located in the gating domain of NBCe1 and neighbours the 3D space occupied by another pRTA-associated residue (Arg881), highlighting an important and conformationally-sensitive region of NBCe1. We conclude that the intracellular retention of Q913R is caused by the loss of Gln at position 913, but that the manifestation of the Cl - leak is related to the introduction of Arg at position 913. Our findings will inform future studies to elucidate the nature and the consequences of the leak.

  20. Local differentiation of cell wall matrix polysaccharides in sinuous pavement cells: its possible involvement in the flexibility of cell shape.

    Sotiriou, P; Giannoutsou, E; Panteris, E; Galatis, B; Apostolakos, P

    2018-03-01

    The distribution of homogalacturonans (HGAs) displaying different degrees of esterification as well as of callose was examined in cell walls of mature pavement cells in two angiosperm and two fern species. We investigated whether local cell wall matrix differentiation may enable pavement cells to respond to mechanical tension forces by transiently altering their shape. HGA epitopes, identified with 2F4, JIM5 and JIM7 antibodies, and callose were immunolocalised in hand-made or semithin leaf sections. Callose was also stained with aniline blue. The structure of pavement cells was studied with light and transmission electron microscopy (TEM). In all species examined, pavement cells displayed wavy anticlinal cell walls, but the waviness pattern differed between angiosperms and ferns. The angiosperm pavement cells were tightly interconnected throughout their whole depth, while in ferns they were interconnected only close to the external periclinal cell wall and intercellular spaces were developed between them close to the mesophyll. Although the HGA epitopes examined were located along the whole cell wall surface, the 2F4- and JIM5- epitopes were especially localised at cell lobe tips. In fern pavement cells, the contact sites were impregnated with callose and JIM5-HGA epitopes. When tension forces were applied on leaf regions, the pavement cells elongated along the stretching axis, due to a decrease in waviness of anticlinal cell walls. After removal of tension forces, the original cell shape was resumed. The presented data support that HGA epitopes make the anticlinal pavement cell walls flexible, in order to reversibly alter their shape. Furthermore, callose seems to offer stability to cell contacts between pavement cells, as already suggested in photosynthetic mesophyll cells. © 2017 German Society for Plant Sciences and The Royal Botanical Society of the Netherlands.

  1. Phloem sap proteins from Cucurbita maxima and Ricinus communis have the capacity to traffic cell to cell through plasmodesmata

    Balachandran, Suchandra; Xiang, Yu; Schobert, Christian; Thompson, Gary A.; Lucas, William J.

    1997-01-01

    In angiosperms, the functional enucleate sieve tube system of the phloem appears to be maintained by the surrounding companion cells. In this study, we tested the hypothesis that polypeptides present within the phloem sap traffic cell to cell from the companion cells, where they are synthesized, into the sieve tube via plasmodesmata. Coinjection of fluorescently labeled dextrans along with size-fractionated Cucurbita maxima phloem proteins, ranging in size from 10 to 200 kDa, as well as injection of individual fluorescently labeled phloem proteins, provided unambiguous evidence that these proteins have the capacity to interact with mesophyll plasmodesmata in cucurbit cotyledons to induce an increase in size exclusion limit and traffic cell to cell. Plasmodesmal size exclusion limit increased to greater than 20 kDa, but less than 40 kDa, irrespective of the size of the injected protein, indicating that partial protein unfolding may be a requirement for transport. A threshold concentration in the 20–100 nM range was required for cell-to-cell transport indicating that phloem proteins have a high affinity for the mesophyll plasmodesmal binding site(s). Parallel experiments with glutaredoxin and cystatin, phloem sap proteins from Ricinus communis, established that these proteins can also traffic through cucurbit mesophyll plasmodesmata. These results are discussed in terms of the requirements for regulated protein trafficking between companion cells and the sieve tube system. As the threshold value for plasmodesmal transport of phloem sap proteins falls within the same range as many plant hormones, the possibility is discussed that some of these proteins may act as long-distance signaling molecules. PMID:9391168

  2. Phloem sap proteins from Cucurbita maxima and Ricinus communis have the capacity to traffic cell to cell through plasmodesmata.

    Balachandran, S; Xiang, Y; Schobert, C; Thompson, G A; Lucas, W J

    1997-12-09

    In angiosperms, the functional enucleate sieve tube system of the phloem appears to be maintained by the surrounding companion cells. In this study, we tested the hypothesis that polypeptides present within the phloem sap traffic cell to cell from the companion cells, where they are synthesized, into the sieve tube via plasmodesmata. Coinjection of fluorescently labeled dextrans along with size-fractionated Cucurbita maxima phloem proteins, ranging in size from 10 to 200 kDa, as well as injection of individual fluorescently labeled phloem proteins, provided unambiguous evidence that these proteins have the capacity to interact with mesophyll plasmodesmata in cucurbit cotyledons to induce an increase in size exclusion limit and traffic cell to cell. Plasmodesmal size exclusion limit increased to greater than 20 kDa, but less than 40 kDa, irrespective of the size of the injected protein, indicating that partial protein unfolding may be a requirement for transport. A threshold concentration in the 20-100 nM range was required for cell-to-cell transport indicating that phloem proteins have a high affinity for the mesophyll plasmodesmal binding site(s). Parallel experiments with glutaredoxin and cystatin, phloem sap proteins from Ricinus communis, established that these proteins can also traffic through cucurbit mesophyll plasmodesmata. These results are discussed in terms of the requirements for regulated protein trafficking between companion cells and the sieve tube system. As the threshold value for plasmodesmal transport of phloem sap proteins falls within the same range as many plant hormones, the possibility is discussed that some of these proteins may act as long-distance signaling molecules.

  3. DNA binding, anti-tumour activity and reactivity toward cell thiols of ...

    leukemia cell line L1210 than alkene 1. The incubation of ... intracellular target of anti-cancer drugs, so any interac- ... lymphocytic leukemia cells (L1210) and human ovarian cancer cell ..... was also supplemented with 2 mM glutamine. The.

  4. Thirteen-week oral toxicity study of L-glutamine in rats.

    Tsubuku, Shoji; Hatayama, Kazuhisa; Mawatari, Kazunori; Smriga, Miro; Kimura, Takeshi

    2004-01-01

    L-Glutamine (Gln) is a semiessential amino acid used in enteral feeding in critically ill patients, and is contained in numerous dietary supplements available to the general public. This study evaluated toxicological effects of Gln in male and female Sprague-Dawley rats. Gln produced by Ajinomoto Co. (Tokyo, Japan) was incorporated into a standard diet at doses equal to 1.25%, 2.5%, and 5.0% (w/w), respectively. A control group of rats received only a standard diet. All diets were administered ad libitum for 13 consecutive weeks. To examine recoverability of any potential effects, the administration period was followed by a 5-week recovery period, during which only the standard diet was provided to all animals. Throughout the administration and recovery periods, no deaths were observed, and no changes in diet consumption, ophthalmologic findings, gross pathology, and histopathology were detected. Several changes in urine parameters (total protein, urine pH, and a positive incidence (+/-) of ketone bodies) were observed in the 2.5% and 5.0% groups at the end of the administration period. Minor increases were found in hematology parameters for the 5.0% group (platelet count, gamma-globulin, lactate dehydrogenase [LDH]), but all changes were within physiological range. No effects of administration were observed in the 1.25% group. The no-observed-adverse-effect level (NOAEL) for Gln was estimated at 1.25% for both genders (males 0.83 +/- 0.01 g/kg/day; females, 0.96 +/- 0.06 g/kg/day).

  5. Glutamate-glutamine and GABA in brain of normal aged and patients with cognitive impairment.

    Huang, Dandan; Liu, Dan; Yin, Jianzhong; Qian, Tianyi; Shrestha, Susan; Ni, Hongyan

    2017-07-01

    To explore the changes of glutamate-glutamine (Glx) and gamma-aminobutyric acid (GABA) in the brain in normal old age and cognitive impairment using magnetic resonance spectroscopy (MRS). Seventeen normal young controls (NYC), 15 normal elderly controls (NEC), 21 patients with mild cognitive impairment (MCI) and 17 with Alzheimer disease (AD) patients were included in this study. Glx and GABA+ levels in the anterior cingulate cortex (ACC) and right hippocampus (rHP) were measured by using a MEGA-PRESS sequence. Glx/Cr and GABA+/Cr ratios were compared between NYC and NEC and between the three elderly groups using analysis of covariance (ANCOVA); the tissue fractions of voxels were used as covariates. The relationships between metabolite ratios and cognitive performance were analysed using Spearman correlation coefficients. For NEC and NYC groups, Glx/Cr and GABA+/Cr ratios were lower in NEC in ACC and rHP. For the three elderly groups, Glx/Cr ratio was lower in AD in ACC compared to NEC and MCI; Glx/Cr ratio was lower in AD in rHP compared to NEC. There was no significant decrease for GABA+/Cr ratio. Glx and GABA levels may decrease simultaneously in normal aged, and Glx level decreased predominantly in AD, and it is helpful in the early diagnosis of AD. • Glx and GABA levels may decrease simultaneously in normal aged. • Glx level may decrease predominantly in Alzheimer disease. • The balance in excitatory-inhibitory systems may be broken in AD. • Decreased Glx level may be helpful in early diagnosis of AD.

  6. Relationship among Glutamine, γ-Aminobutyric Acid, and Social Cognition in Autism Spectrum Disorders

    Sikoglu, Elif M.; Hodge, Steven M.; Edden, Richard A.E.; Foley, Ann; Kennedy, David N.; Moore, Constance M.; Frazier, Jean A.

    2015-01-01

    Abstract Objective: An imbalance of excitatory and inhibitory neurotransmission in autism spectrum disorder (ASD) has been proposed. We compared glutamate (Glu), glutamine (Gln), and γ-aminobutyric acid (GABA) levels in the anterior cingulate cortex (ACC) of 13 males with ASD and 14 typically developing (TD) males (ages 13–17), and correlated these levels with intelligence quotient (IQ) and measures of social cognition. Methods: Social cognition was evaluated by administration of the Social Responsiveness Scale (SRS) and the Reading the Mind in the Eyes Test (RMET). We acquired proton magnetic resonance spectroscopy (1H-MRS) data from the bilateral ACC using the single voxel point resolved spectroscopy sequence (PRESS) to quantify Glu and Gln, and Mescher–Garwood point-resolved spectroscopy sequence (MEGA-PRESS) to quantify GABA levels referenced to creatine (Cr). Results: There were higher Gln levels (p=0.04), and lower GABA/Cre levels (p=0.09) in the ASD group than in the TD group. There was no difference in Glu levels between groups. Gln was negatively correlated with RMET score (rho=−0.62, p=0.001) and IQ (rho=−0.56, p=0.003), and positively correlated with SRS scores (rho=0.53, p=0.007). GABA/Cre levels were positively correlated with RMET score (rho=0.34, p=0.09) and IQ (rho=0.36, p=0.07), and negatively correlated with SRS score (rho=−0.34, p=0.09). Conclusions: These data suggest an imbalance between glutamatergic neurotransmission and GABA-ergic neurotransmission in ASD. Higher Gln levels and lower GABA/Cre levels were associated with lower IQ and greater impairments in social cognition across groups. PMID:25919578

  7. Oral Supplementation with Beta-Hydroxy-Beta-Methylbutyrate, Arginine, and Glutamine Improves Lean Body Mass in Healthy Older Adults.

    Ellis, Amy C; Hunter, Gary R; Goss, Amy M; Gower, Barbara A

    2018-04-19

    Oral intake of beta-hydroxy-beta-methylbutyrate (HMB), arginine, and glutamine may ameliorate muscle loss by stimulating protein synthesis and decreasing protein degradation while simultaneously decreasing inflammation. Previous studies provide evidence for improvement in body composition with dietary supplementation of these ingredients among patients with muscle-wasting diseases. The objectives of this study were to examine the effects of this amino acid mixture on lean body mass, muscle volume, and physical function among healthy older adults. Thirty-one community-dwelling men and women, aged 65-89 years, were randomized to either two oral doses of the amino acid supplement (totaling 3 g HMB, 14 g arginine, 14 g glutamine) or placebo daily for six months. At baseline and month six, lean body mass was measured by air displacement plethysmography, dual-energy X-ray absorptiometry (DXA), and four-compartment model. Muscle volume of quadriceps was quantified by magnetic resonance imaging (MRI), and participants performed a battery of tests to assess physical function. As compared to the placebo group, the treatment group exhibited improvement in a timed stair climb (p =.016) as well as significant increases in lean body mass by all methods of assessment (p lean mass in the supplement group only (p =.035). However, no change was observed in MRI-derived quadriceps volume. Dietary supplementation with HMB, arginine, and glutamine improved total body lean mass among a small sample of healthy older adults. Further research is indicated to elucidate mechanisms of action and to determine whether supplementation may benefit frail elders. Registered under ClinicalTrials.gov identifier no. NCT01057082.

  8. Uptake and metabolism of L-[3H]glutamate and L-[3H]glutamine in adult rat cerebellar slices

    de Barry, J.; Vincendon, G.; Gombos, G.

    1983-01-01

    Using very low concentrations (1 mumol range) of L-2-3-[ 3 H]glutamate, ( 3 H-Glu) or L-2-3-[ 3 H]glutamine ( 3 H-Gln), the authors have previously shown by autoradiography that these amino acids were preferentially taken up in the molecular layer of the cerebellar cortex. Furthermore, the accumulation of 3 H-Glu was essentially glial in these conditions. Uptake and metabolism of either ( 3 H-Glu) or ( 3 H-Gln) were studied in adult rat cerebellar slices. Both amino acids were rapidly converted into other metabolic compounds: after seven minutes of incubation in the presence of exogenous 3 H-Glu, 70% of the tissue accumulated radioactivity was found to be in compounds other than glutamate. The main metabolites were Gln (42%), alpha-ketoglutarate (25%) and GABA (1,4%). In the presence of exogenous 3 H-Gln the rate of metabolism was slightly slower (50% after seven minutes of incubation) and the metabolites were also Glu (29%), alpha-ketoglutarate (15%) and GABA (5%). Using depolarizing conditions (56 mM KCl) with either exogenous 3 H-Glu or 3 H-Gln, the radioactivity was preferentially accumulated in glutamate compared to control. From these results we conclude: i) there are two cellular compartments for the neurotransmission-glutamate-glutamine cycle; one is glial, the other neuronal; ii) these two cellular compartments contain both Gln and Glu; iii) transmitter glutamate is always in equilibrium with the so-called ''metabolic'' pool of glutamate; iv) the regulation of the glutamate-glutamine cycle occurs at least at two different levels: the uptake of glutamate and the enzymatic activity of the neuronal glutaminase

  9. Separation and determination of acetyl-glutamine enantiomers by HPLC–MS and its application in pharmacokinetic study

    Xiaoxiao Zhang

    2017-10-01

    Full Text Available A high-performance liquid chromatography coupled with mass spectrometry (HPLC–MS method was established for the separation and determination of acetyl-glutamine enantiomers (acetyl-L-glutamine and acetyl-D-glutamine simultaneously. Baseline separation was achieved on Chiralpak AD-H column (250 mm × 4.6 mm, 5 µm. n-Hexane (containing 0.1% acetic acid and ethanol (75:25, v/v were used as mobile phase at a flow rate of 0.6 mL/min. The detection was operated in the negative ion mode with an ESI source. [M-H]− m/z 187.0540 for enantiomers and [M-H]− m/z 179.0240 for aspirin (IS were selected as detecting ions. The linear range of the calibration curve for each enantiomer was 0.05–40 µg/mL. The precision of this method at concentrations of 0.5–20 µg/mL was within 7.23%, and the accuracy was 99.81%–107.81%. The precision at LOQ (0.05 µg/mL was between 16.28% and 17.56%, which was poor than that at QC levels. The average extraction recovery was higher than 85% for both enantiomers at QC levels. The pharmacokinetics of enantiomers was found to be stereoselective. There was not chiral inversion in vivo or in vitro between enantiomers.

  10. Post-Transcriptional Regulation Prevents Accumulation of Glutathione Reductase Protein and Activity in the Bundle Sheath Cells of Maize1

    Pastori, Gabriela M.; Mullineaux, Philip M.; Foyer, Christine H.

    2000-01-01

    Glutathione reductase (GR; EC 1.6.4.2) activity was assayed in bundle sheath and mesophyll cells of maize (Zea mays L. var H99) from plants grown at 20°C, 18°C, and 15°C. The purity of each fraction was determined by measuring the associated activity of the compartment-specific marker enzymes, Rubisco and phosphoenolpyruvate carboxylase, respectively. GR activity and the abundance of GR protein and mRNA increased in plants grown at 15°C and 18°C compared with those grown at 20°C. In all cases GR activity was found only in mesophyll fractions of the leaves, with no GR activity being detectable in bundle sheath extracts. Immunogold labeling with GR-specific antibodies showed that the GR protein was exclusively localized in the mesophyll cells of leaves at all growth temperatures, whereas GR transcripts (as determined by in situ hybridization techniques) were observed in both cell types. These results indicate that post-transcriptional regulation prevents GR accumulation in the bundle sheath cells of maize leaves. The resulting limitation on the capacity for regeneration of reduced glutathione in this compartment may contribute to the extreme chilling sensitivity of maize leaves. PMID:10712529

  11. Simultaneous infusion of glutamine and branched-chain amino acids (BCAA) to septic rats does not have more favorable effect on protein synthesis in muscle, liver, and small intestine than separate infusions.

    Holecek, Milan; Muthny, Tomas; Kovarik, Miroslav; Sispera, Ludek

    2006-01-01

    Glutamine and branched-chain amino acids (BCAA; valine, leucine, and isoleucine) are used as nutrition supplements in the treatment of proteocatabolic illness. We hypothesized that simultaneous administration of BCAA and glutamine affects protein metabolism more significantly than separate administration. In the present study, we evaluated their effect on protein synthesis in skeletal muscle, liver, and jejunum of septic rats. Twenty-four hours after induction of sepsis by subcutaneous injection of turpentine, the rats were infused for 6 hours with 5 mL of 1.75% glutamine, 1.75% BCAA, 1.75% glutamine+BCAA, or saline solution. The control group consisted of intact rats infused with saline. Protein synthesis was measured at the end of infusion by a "flooding method" with [3,4,5-(3)H]phenylalanine. In turpentine-treated animals, we observed a decrease in glutamine concentration in blood plasma and skeletal muscle, a decrease in BCAA concentration in liver and jejunum, and a decrease in protein synthesis in all tissues. Glutamine or glutamine+BCAA infusion increased glutamine concentration in plasma and muscle and stimulated protein synthesis in the liver. The BCAA infusion enhanced concentrations of BCAA in plasma and tissues, but the effect of BCAA on protein synthesis was insignificant. Synergistic effect of simultaneous infusion of glutamine and BCAA on protein synthesis was not observed. We conclude that glutamine infusion to rats with septic injury may significantly improve impaired protein synthesis in the liver and that there is no synergistic effect of glutamine and BCAA infusion on protein synthesis in skeletal muscle, liver, and jejunum.

  12. Glycyl-glutamine in nucleus accumbens reduces ethanol intake in alcohol preferring (P) rats.

    Resch, Garth E; Shridharani, Shyam; Millington, William R; Garris, David R; Simpson, C Wayne

    2005-10-05

    Opioid peptides and glycyl-glutamine (Gly-Gln) have been implicated in the control of ethanol consumption. A recognized beta-endorphin cleavage product, Gly-Gln, inhibits voluntary alcohol consumption when microinjected into the nucleus accumbens (AcbSh) of P rats. To evaluate the site-specific efficacy of Gly-Gln on ethanol consumption following AcbSh application, ethanol preferring (P) rats were allowed to establish individual baseline ethanol/water consumption utilizing a voluntary self-administration paradigm. Subsequent to baseline ethanol consumption being established, bilateral guide cannulae were stereotaxically implanted +1 mm dorsal to the AcbSh for subsequent Gly-Gln (100 nmol/microl) or saline vehicle (1 microl) injections. Alcohol intake, body weight, and water intake were measured at 24 h post-injection intervals. Unilateral Gly-Gln injections reduced ethanol consumption 35.6% (P < 0.05) from pre-established baseline consumption (6.24 +/- 0.64 g/kg to 4.06 +/- 0.28 g/kg). Bilateral Gly-Gln injections further reduced consumption to 51.9% (6.4 +/- 1.0 g/kg to 3.08 +/- 0.65 g/kg at 24 h (P < 0.01) below established baseline values within 24 h without significant changes in body weight or water consumption. Also, the amino acid constituents of the dipeptide had no influence on ethanol consumption behavior; however, Gly-Gln efficacy was shown to be comparable to central beta-endorphin-(1-27) or intraperitoneal (i.p.) naltrexone-induced suppression of ethanol intake. These data indicate that the AcbSh exhibits a site-specific sensitivity to the suppressive actions of Gly-Gln or beta-endorphin-(1-27) injections that modulate voluntary ethanol consumption in P rats. These findings support the broader concept that select forebrain opioid-responsive neural sites may influence the development or expression of alcohol abuse syndromes in animal models or humans.

  13. L-alanyl-L-glutamine ingestion maintains performance during a competitive basketball game

    Hoffman Jay R

    2012-03-01

    Full Text Available Abstract Background The purpose of this study was to examine the efficacy of L-alanyl-L-glutamine (AG ingestion on basketball performance, including jump power, reaction time, shooting accuracy and fatigue. Methods Ten women (21.2 ± 1.6 years; height: 177.8 ± 8.7 cm; body mass: 73.5 ± 8.0 kg, all scholarship NCAA Division I basketball players, volunteered for this study. Subjects participated in four trials, each consisting of a 40-min basketball game with controlled time-outs for rehydration. During the first trial (DHY subjects were not allowed to rehydrate, and the total weight lost during the contest was used to determine fluid replenishment during the subsequent three trials. During one trial subjects consumed only water (W, while during the other two trials subjects consumed the AG supplement mixed in water using either a low dose (1 g per 500 ml (AG1 or high dose (2 g per 500 ml (AG2 concentration. All data assessed prior to and following each game were converted into a Δ score (Post results - Pre results. All performance data were then analyzed using a one-way repeated measures analysis of variance. Results During DHY subjects lost 1.72 ± 0.42 kg (2.3% of their body mass. No differences in fluid intake (1.55 ± 0.43 L were seen between rehydration trials. A 12.5% (p = 0.016 difference in basketball shooting performance was noted between DHY and AG1 and an 11.1% (p = 0.029 difference was seen between AG1 and W. Visual reaction time was significantly greater following AG1 (p = 0.014 compared to DHY. Differences (p = 0.045 in fatigue, as determined by player loads, were seen only between AG2 and DHY. No differences were seen in peak or mean vertical jump power during any trial. Conclusion Rehydration with AG appears to maintain basketball skill performance and visual reaction time to a greater extent than water only.

  14. L-alanyl-L-glutamine ingestion maintains performance during a competitive basketball game.

    Hoffman, Jay R; Williams, David R; Emerson, Nadia S; Hoffman, Mattan W; Wells, Adam J; McVeigh, Daniele M; McCormack, William P; Mangine, Gerald T; Gonzalez, Adam M; Fragala, Maren S

    2012-03-07

    The purpose of this study was to examine the efficacy of L-alanyl-L-glutamine (AG) ingestion on basketball performance, including jump power, reaction time, shooting accuracy and fatigue. Ten women (21.2 ± 1.6 years; height: 177.8 ± 8.7 cm; body mass: 73.5 ± 8.0 kg), all scholarship NCAA Division I basketball players, volunteered for this study. Subjects participated in four trials, each consisting of a 40-min basketball game with controlled time-outs for rehydration. During the first trial (DHY) subjects were not allowed to rehydrate, and the total weight lost during the contest was used to determine fluid replenishment during the subsequent three trials. During one trial subjects consumed only water (W), while during the other two trials subjects consumed the AG supplement mixed in water using either a low dose (1 g per 500 ml) (AG1) or high dose (2 g per 500 ml) (AG2) concentration. All data assessed prior to and following each game were converted into a Δ score (Post results - Pre results). All performance data were then analyzed using a one-way repeated measures analysis of variance. During DHY subjects lost 1.72 ± 0.42 kg (2.3%) of their body mass. No differences in fluid intake (1.55 ± 0.43 L) were seen between rehydration trials. A 12.5% (p = 0.016) difference in basketball shooting performance was noted between DHY and AG1 and an 11.1% (p = 0.029) difference was seen between AG1 and W. Visual reaction time was significantly greater following AG1 (p = 0.014) compared to DHY. Differences (p = 0.045) in fatigue, as determined by player loads, were seen only between AG2 and DHY. No differences were seen in peak or mean vertical jump power during any trial. Rehydration with AG appears to maintain basketball skill performance and visual reaction time to a greater extent than water only.

  15. A conserved glutamine plays a central role in LOV domain signal transmission and duration

    Nash, Abigail I.; Ko, Wen-Huang; Harper, Shannon M.; Gardner, Kevin H.

    2009-01-01

    Light is a key stimulus for plant biological functions, several of which are controlled by light-activated kinases known as phototropins, a group of kinases that contain two light-sensing domains (LOV, Light-Oxygen-Voltage domains) and a C-terminal serine/threonine kinase domain. The second sensory domain, LOV2, plays a key role in regulating kinase enzymatic activity via the photochemical formation of a covalent adduct between a LOV2 cysteine residue and an internally-bound flavin mononucleotide (FMN) chromophore. Subsequent conformational changes in LOV2 lead to the unfolding of a peripheral Jα helix, and ultimately, phototropin kinase activation. To date, the mechanism coupling bond formation and helix dissociation has remained unclear. Previous studies found that a conserved glutamine residue (Q513 in the Avena sativa phototropin 1 LOV2 (AsLOV2) domain) switches its hydrogen-bonding pattern with FMN upon light stimulation. Located in the immediate vicinity of the FMN binding site, this Gln residue is provided by the Iβ strand that interacts with the Jα helix, suggesting a route for signal propagation from the core of the LOV domain to its peripheral Jα helix. To test whether Q513 plays a key role in tuning the photochemical and transduction properties of AsLOV2, we designed two point mutations, Q513L and Q513N, and monitored the effects on the chromophore and protein using a combination of UV-visible absorbance and circular dichroism spectroscopy, limited proteolysis, and solution NMR. The results show that these mutations significantly dampen the changes between the dark and lit state AsLOV2 structures, leaving the protein in a pseudo-dark state (Q513L) or a pseudo-lit state (Q513N) conformation. Further, both mutations changed the photochemical properties of this receptor, particularly the lifetime of the photoexcited signaling states. Together, these data establish that this residue plays a central role in both spectral tuning and signal propagation from

  16. Risk-Conferring Glutamatergic Genes and Brain Glutamate Plus Glutamine in Schizophrenia

    Juan R. Bustillo

    2017-06-01

    Full Text Available BackgroundThe proton magnetic resonance spectroscopy (1H-MRS signals from glutamate (or the combined glutamate and glutamine signal—Glx have been found to be greater in various brain regions in people with schizophrenia. Recently, the Psychiatric Genetics Consortium reported that several common single-nucleotide polymorphisms (SNPs in glutamate-related genes confer increased risk of schizophrenia. Here, we examined the relationship between presence of these risk polymorphisms and brain Glx levels in schizophrenia.Methods1H-MRS imaging data from an axial, supraventricular tissue slab were acquired in 56 schizophrenia patients and 67 healthy subjects. Glx was measured in gray matter (GM and white matter (WM regions. The genetic data included six polymorphisms genotyped across an Illumina 5M SNP array. Only three of six glutamate as well as calcium-related SNPs were available for examination. These included three glutamate-related polymorphisms (rs10520163 in CLCN3, rs12704290 in GRM3, and rs12325245 in SLC38A7, and three calcium signaling polymorphisms (rs1339227 in RIMS1, rs7893279 in CACNB2, and rs2007044 in CACNA1C. Summary risk scores for the three glutamate and the three calcium polymorphisms were calculated.ResultsGlx levels in GM positively correlated with glutamate-related genetic risk score but only in younger (≤36 years schizophrenia patients (p = 0.01. Glx levels did not correlate with calcium risk scores. Glx was higher in the schizophrenia group compared to levels in controls in GM and WM regardless of age (p < 0.001.ConclusionElevations in brain Glx are in part, related to common allelic variants of glutamate-related genes known to increase the risk for schizophrenia. Since the glutamate risk scores did not differ between groups, some other genetic or environmental factors likely interact with the variability in glutamate-related risk SNPs to contribute to an increase in brain Glx early in the illness.

  17. Zinc, vitamin A, and glutamine supplementation in Brazilian shantytown children at risk for diarrhea results in sex-specific improvements in verbal learning

    Aldo A. M. Lima

    2013-01-01

    Full Text Available OBJECTIVE: To identify the impact of supplemental zinc, vitamin A, and glutamine, alone or in combination, on long-term cognitive outcomes among Brazilian shantytown children with low median height-for-age z-scores. METHODS: A randomized, double-blind, placebo-controlled trial was conducted in children aged three months to nine years old from the urban shanty compound community of Fortaleza, Brazil. Demographic and anthropometric information was assessed. The random treatment groups available for cognitive testing (total of 167 children were: (1 placebo, n = 25; (2 glutamine, n = 23; (3 zinc, n = 18; (4 vitamin A, n = 19; (5 glutamine+zinc, n = 20; (6 glutamine+vitamin A, n = 21; (7 zinc+vitamin A, n = 23; and (8 glutamine+zinc+vitamin A, n = 18. Neuropsychological tests were administered for the cognitive domains of non-verbal intelligence and abstraction, psychomotor speed, verbal memory and recall ability, and semantic and phonetic verbal fluency. Statistical analyses were performed using SPSS, version 16.0. ClinicalTrials.gov: NCT00133406. RESULTS: Girls receiving a combination of glutamine, zinc, and vitamin A had higher mean age-adjusted verbal learning scores than girls receiving only placebo (9.5 versus 6.4, p = 0.007 and girls receiving zinc+vitamin A (9.5 versus 6.5, p = 0.006. Similar group differences were not found between male study children. CONCLUSIONS: The findings suggest that combination therapy offers a sex-specific advantage on tests of verbal learning, similar to that seen among female patients following traumatic brain injury.

  18. Conformation-Specific IR and UV Spectroscopy of the Amino Acid Glutamine: Amide-Stacking and Hydrogen Bonding in AN Important Residue in Neurodegenerative Diseases

    Walsh, Patrick S.; Dean, Jacob C.; Zwier, Timothy S.

    2014-06-01

    Glutamine plays an important role in several neurodegenerative diseases including Huntington's disease (HD) and Alzheimer's disease (AD). An intriguing aspect of the structure of glutamine is its incorporation of an amide group in its side chain, thereby opening up the possibility of forming amide-amide H-bonds between the peptide backbone and side chain. In this study the conformational preferences of two capped gluatamines Z(carboxybenzyl)-Glutamine-X (X=OH, NHMe) are studied under jet-cooled conditions in the gas phase in order to unlock the intrinsic structural motifs that are favored by this flexible sidechain. Conformational assignments are made by comparing the hydride stretch ( 3100-3700 cm-1) and amide I and II ( 1400-1800 cm-1) resonant ion-dip infrared spectra with predictions from harmonic frequency calculations. Assigned structures will be compared to previously published results on both natural and unnatural residues. Particular emphasis will be placed on the comparison between glutamine and unconstrained γ-peptides due to the similar three-carbon spacing between backbone and side chain in glutamine to the backbone spacing in γ-peptides. The ability of the glutamine side-chain to form amide stacked conformations will be a main focus, along with the prevalence of extended backbone type structures. W. H. James, III, C W. Müller, E. G. Buchanan, M. G. D. Nix, L. Guo, L. Roskop, M. S. Gordon, L. V. Slipchenko, S. H. Gellman, and T. S. Zwier, J. Am. Chem. Soc., 2009, 131(40), 14243-14245.

  19. 7T Proton Magnetic Resonance Spectroscopy of Gamma-Aminobutyric Acid, Glutamate, and Glutamine Reveals Altered Concentrations in Patients With Schizophrenia and Healthy Siblings.

    Thakkar, Katharine N; Rösler, Lara; Wijnen, Jannie P; Boer, Vincent O; Klomp, Dennis W J; Cahn, Wiepke; Kahn, René S; Neggers, Sebastiaan F W

    2017-03-15

    The N-methyl-D-aspartate receptor hypofunction model of schizophrenia predicts dysfunction in both glutamatergic and gamma-aminobutyric acidergic (GABAergic) transmission. We addressed this hypothesis by measuring GABA, glutamate, glutamine, and the sum of glutamine plus glutamate concentrations in vivo in patients with schizophrenia using proton magnetic resonance spectroscopy at 7T, which allows separation of metabolites that would otherwise overlap at lower field strengths. In addition, we investigated whether altered levels of GABA, glutamate, glutamine, and the sum of glutamine plus glutamate reflect genetic vulnerability to schizophrenia by including healthy first-degree relatives. Proton magnetic resonance spectroscopy at 7T was performed in 21 patients with chronic schizophrenia who were taking medication, 23 healthy first-degree relatives of patients with schizophrenia, and 24 healthy nonrelatives. Glutamate, glutamine, and GABA were measured cortically and subcortically in bilateral basal ganglia and occipital cortex. Patients with schizophrenia had reduced cortical GABA compared with healthy relatives and the combined sample of healthy relatives and healthy nonrelatives, suggesting that altered GABAergic systems in schizophrenia are associated with either disease state or medication effects. Reduced cortical glutamine relative to healthy control subjects was observed in patients with schizophrenia and the combined sample of healthy relatives and patients with schizophrenia, suggesting that altered glutamatergic metabolite levels are associated with illness liability. No group differences were found in the basal ganglia. Taken together, these findings are consistent with alterations in GABAergic and glutamatergic systems in patients with schizophrenia and provide novel insights into these systems in healthy relatives. Copyright © 2016 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

  20. Inhibition of glutamine synthesis induces glutamate dehydrogenase-dependent ammonia fixation into alanine in co-cultures of astrocytes and neurons

    Dadsetan, Sherry; Bak, Lasse Kristoffer; Sørensen, Michael

    2011-01-01

    study it was investigated if the glutamine synthetase (GS) inhibitor methionine sulfoximine (MSO) would enhance alanine synthesis by blocking the GS-dependent ammonia scavenging process. Hence, co-cultures of neurons and astrocytes were incubated for 2.5h with [U-(13)C]glucose to monitor de novo......It has been previously demonstrated that ammonia exposure of neurons and astrocytes in co-culture leads to net synthesis not only of glutamine but also of alanine. The latter process involves the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT). In the present...

  1. Mixed ligand complexes of essential metal ions with L-glutamine and succinic acid in SLS-water mixtures

    Bindu Hima Gandham

    2012-01-01

    Full Text Available Speciation of mixed ligand complexes of Co(II, Ni(II and Cu(II with L-glutamine and succinic acid was studied in varying amounts (0.0-2.5% w/v of sodium lauryl sulphate in aqueous solutions maintaining an ionic strength of 0.16 mol L-1 (NaCl at 303.0 K. Titrations were carried out in the presence of different relative concentrations (M : L : X = 1 : 2 : 2, 1 : 4 : 2, 1 : 2 : 4 of metal (M to L-glutamine (L to succinic acid (X with sodium hydroxide. Stability constants of ternary complexes were refined with MINIQUAD75. The best-fit chemical models were selected based on statistical parameters and residual analysis. The species detected were ML2X, MLX, MLXH and MLXH2 for Co(II, Ni(II and Cu(II. Extra stability of ternary complexes compared to their binary complexes was believed to be due to electrostatic interactions of the side chains of ligands, charge neutralization, chelate effect, stacking interactions and hydrogen bonding. The species distribution with pH at different compositions of SLS and plausible equilibria for the formation of species were also presented.

  2. The role of adrenal hormones in the response of glutamine synthetase to fasting in adult and old rats.

    Mezzarobba, V; Torrent, A; Leydier, I; Alles, S; Brajon, B; Mignon, M; Attaix, D; Meynial-Denis, D

    2003-12-01

    During fasting, skeletal muscle exports increased amounts of glutamine (Gln) while increasing the production of this amino acid by glutamine synthetase (GS) in order to maintain the intramuscular Gln pool. Glucocorticoid hormones are believed to be the principal mediators of GS induction during stress conditions. The aim of this study was to evaluate (1) the effect of fasting on GS activity and expression in skeletal muscle during aging and consequently, (2) the role of glucocorticoids in fasting-induced GS activity. Male Wistar rats (6-, 22-month old) were fasted for 5 days and both the activity and expression of GS were measured in tibialis anterior muscle. To better demonstrate the role of glucocorticoids in the response of GS to fasting, we suppressed their action by RU38486 administration (a potent glucocorticoid antagonist) and their production by adrenalectomy in fed and fasted rats. An increase in fasting-induced GS activity was observed in skeletal muscles from both adult and aged rats. Adrenalectomy, but surprisingly not RU38486, suppressed the fasting-induced increase in GS activity and expression. The data clearly show that the GS responsiveness to fasting was not modified by aging in skeletal muscle.

  3. Perinatal supplementation of 4-phenylbutyrate and glutamine attenuates endoplasmic reticulum stress and improves colonic epithelial barrier function in rats born with intrauterine growth restriction.

    Désir-Vigné, Axel; Haure-Mirande, Vianney; de Coppet, Pierre; Darmaun, Dominique; Le Dréan, Gwenola; Segain, Jean-Pierre

    2018-05-01

    Intrauterine growth restriction (IUGR) can affect the structure and function of the intestinal barrier and increase digestive disease risk in adulthood. Using the rat model of maternal dietary protein restriction (8% vs. 20%), we found that the colon of IUGR offspring displayed decreased mRNA expression of epithelial barrier proteins MUC2 and occludin during development. This was associated with increased mRNA expression of endoplasmic reticulum (ER) stress marker XBP1s and increased colonic permeability measured in Ussing chambers. We hypothesized that ER stress contributes to colonic barrier alterations and that perinatal supplementation of dams with ER stress modulators, phenylbutyrate and glutamine (PG) could prevent these defects in IUGR offspring. We first demonstrated that ER stress induction by tunicamycin or thapsigargin increased the permeability of rat colonic tissues mounted in Ussing chamber and that PG treatment prevented this effect. Therefore, we supplemented the diet of control and IUGR dams with PG during gestation and lactation. Real-time polymerase chain reaction and histological analysis of colons from 120-day-old offspring revealed that perinatal PG treatment partially prevented the increased expression of ER stress markers but reversed the reduction of crypt depth and goblet cell number in IUGR rats. In dextran sodium sulfate-induced injury and recovery experiments, the colon of IUGR rats without perinatal PG treatment showed higher XBP1s mRNA levels and histological scores of inflammation than IUGR rats with perinatal PG treatment. In conclusion, these data suggest that perinatal supplementation with PG could alleviate ER stress and prevent epithelial barrier dysfunction in IUGR offspring. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Arginine, N-carbamylglutamate, and glutamine exert protective effects against oxidative stress in rat intestine

    Liang Xiao

    2016-09-01

    Full Text Available The objective of the current study is to evaluate the effects of dietary supplementation with arginine (ARG, N-carbamylglutamate (NCG, and glutamine (GLN on rat intestinal morphology and antioxidant status under oxidative stress. Rats were fed for 30 d with one of the following iso-nitrogenous diets: basal diet (BD, BD plus 1% ARG, BD plus 0.1% NCG, and BD plus 1% GLN. On day 28, half of the rats fed BD were intraperitoneally injected with 12 mg/kg body weight of diquat (DT; i.e., the DT group and the other half was intraperitoneally injected with sterile solution (i.e., the control group. The other diet groups were intraperitoneally injected with 12 mg/kg body weight of DT (i.e., DT + 1% GLN [DT + GLN], DT + 1% ARG [DT + ARG], and DT + 0.1% NCG [DT + NCG]. Rat jejunum samples obtained at 48 h after DT injection were analyzed. Results showed that DT significantly decreased catalase (CAT activity and glutathione (GSH content by 58.25% and 56.57%, respectively, and elevated malondialdehyde (MDA content and crypt depth (CD by 19.39% and 22.13%, respectively, in the jejunum (P < 0.05, relative to the control group. Compared with the DT group, the DT + GLN group exhibited significantly improved villus height (VH, villus width (VW, villus surface area (VSA, CD and total antioxidant capacity (T-AOC activity (P < 0.05; the DT + ARG group exhibited significantly increased the ratio of VH to CD (H:D and T-AOC activity (P < 0.05; the DT + GLN, DT + ARG and DT + NCG groups exhibited significantly enhanced CAT activity and GSH content as well as decreased MDA content (P < 0.05. Moreover, VH, VW, VSA, CD and GSH content in the DT + GLN group were higher whereas MDA content was lower compared with the corresponding values observed in both the DT + ARG and the DT + NCG groups (P < 0.05. The H:D ratio in the DT + ARG group significantly increased compared with that in the DT + NCG and DT + GLN groups (P < 0

  5. Nitrogen Control in Pseudomonas aeruginosa : A Role for Glutamine in the Regulation of the Synthesis of NADP-Dependent Glutamate Dehydrogenase, Urease and Histidase

    Janssen, Dick B.; Herst, Patricia M.; Joosten, Han M.L.J.; Drift, Chris van der

    1981-01-01

    In Pseudomonas aeruginosa the formation of urease, histidase and some other enzymes involved in nitrogen assimilation is repressed by ammonia in the growth medium. The key metabolite in this process appears to be glutamine or a product derived from it, since ammonia and glutamate did not repress

  6. Hyperhydricity Phenomena Problem in Embryonic Callus of Date Palm, Solving by Glutamine and NH4+: No3- Ratio in Basal Nutrient Medium

    El-Dawayaty, M.M.; Zayed, Z.E.; Abdel-Gelil, L.M.

    2012-01-01

    Hyperhydricity is a serious problem faced in vitro date palm propagation which directly effects on the commercial production. So we try to solve this problem by studying, the effect of glutamine as the organic source of nitrogen and NH 4 + :NO 3- ratio as the inorganic source of nitrogen to decrease hyperhydricity phenomena and to produce normal somatic embryos of date palm cv. Gondila.Vitrified embryonic callus were inoculated on MS basal nutrient medium modified with glutamine levels and NH 4 + : NO 3 - ratio. Five concentration ratios of NH 4 : NO 3 (10:15, 15:10, 0:20, 20:0, 0:0 ml/l) were used with 0.1 mg/l NAA for 8 weeks throughout 2 recultures. There were gradually increasing in the percentage of vitrified embryonic callus differentiation to normal somatic embryos by increasing glutamine levels from 0.0 to 400 mg/l. Glutamine at the lowest level (50 mg/l) increased significantly the number of vitrified somatic embryos. Ammonium as the sole source of N resulted in depression in somatic embryos differentiation and escalated the frequency of hyperhydricity whereas,if nitrate was used as the sole N source, somatic embryos good quality were produced and hyperhydricity was eliminated

  7. Molecular identification and characterisation of the glycine transporter (GLYT1) and the glutamine/glutamate transporter (ASCT2) in the rat lens

    Lim, Julie; Lorentzen, Karen Axelgaard; Kistler, Joerg

    2006-01-01

    Glutathione (GSH) is an essential antioxidant required for the maintenance of lens transparency. In the lens, GSH is maintained at unusually high concentrations as a result of direct GSH uptake and/or intracellular de novo synthesis from its precursor amino acids; cysteine, glycine and glutamine/...

  8. Effects of L-glutamine on rectal temperature and some markers of oxidative stress in Red Sokoto goats during the hot-dry season.

    Ocheja, Ohiemi Benjamin; Ayo, Joseph Olusegun; Aluwong, Tagang; Minka, Ndazo Salka

    2017-08-01

    The experiment investigated the ameliorative effects of L-glutamine administration on rectal temperature (RT), erythrocyte osmotic fragility (EOF), serum antioxidant enzyme activities and malondialdehyde (MDA) concentration in Red Sokoto goats during the hot-dry season. Twenty eight healthy Red Sokoto goats, comprising 14 experimental (administered 0.2 g/kg of L-glutamine dissolved in 10 mL of distilled water, once daily for 21 days) and 14 control (administered equivalent of distilled water) goats served as subjects. Rectal temperature (measured at 6:00, 13:00 and 18:00 h) and blood samples (taken at 8:00 h) were obtained from all subjects weekly, before, during and after L-glutamine administration. Data obtained were compared using one-way repeated-measures ANOVA, followed by Tukey's post-hoc test. The dry-bulb temperature, relative humidity and temperature-humidity index for the study period ranged between 24.0 and 37.5 °C, 26.0 and 84.0% and 73.0 and 86.3, respectively. L-glutamine administration decreased (P heat-stressed goats during the hot-dry season.

  9. A randomised trial of enteral glutamine supplementation for very preterm children showed no beneficial or adverse long-term neurodevelopmental outcomes

    Twilhaar, E.S.; de Kieviet, J.F.; Oosterlaan, J.; van Elburg, R.M.

    2017-01-01

    Aim This study evaluated the long-term effects of enteral glutamine supplementation on neurodevelopmental outcomes of a Dutch cohort of very preterm children at 13 years of age. Methods The cohort was enrolled in a randomised placebo-controlled trial between 2001 and 2003 in which infants received

  10. A randomised trial of enteral glutamine supplementation for very preterm children showed no beneficial or adverse long-term neurodevelopmental outcomes.

    Twilhaar, E Sabrina; de Kieviet, Jorrit F; Oosterlaan, Jaap; van Elburg, Ruurd M

    2018-04-01

    This study evaluated the long-term effects of enteral glutamine supplementation on neurodevelopmental outcomes of a Dutch cohort of very preterm children at 13 years of age. The cohort was enrolled in a randomised placebo-controlled trial between 2001 and 2003 in which infants received glutamine- or alanine-supplemented enteral nutrition during the first month of life. Participants were invited for follow-up at a mean age of 13.30 years. Motor, neurocognitive, academic and behavioural outcomes were assessed in 61 children. No differences were found between the groups regarding motor, intellectual, academic and behavioural functioning. Forward span visuospatial working memory performance was better in the controls (crude/adjusted model: d = 0.67/0.64, p = 0.02/0.02), but no difference was found for backward span. After the data were adjusted for confounders, the groups differed regarding parent-rated attention (crude/adjusted model: d = 0.47/0.73, p = 0.07/0.003), but both groups scored within the normal range. This was the first study on the long-term effects of enteral glutamine supplementation on the neurodevelopmental outcomes of very preterm children. Our study provided no evidence that enteral glutamine supplementation had any beneficial or adverse effects on the children's motor, neurocognitive, academic and behavioural outcomes at 13 years of age. ©2017 Foundation Acta Paediatrica. Published by John Wiley & Sons Ltd.

  11. Survey of Synergistic Effect of L-carnitine with Glutamine on Body Composition and Dietary Intake in Soccer Players: A Double-blind, Randomized Clinical Trial

    Mohammad Hozoori

    2016-10-01

    Full Text Available Background: The present study was conducted to investigate the possible effects of L-carnitine and glutamine and their synergistic effects on male soccer athletes. Methods: 28 male soccer players (21.1 ± 0.7 y were enrolled in a randomized pre and post intervention, double-blind design. Before the intervention, their performances were assessed by Bruce protocol, and their body composition was measured with the body composition analyzer. Then, athletes were randomly allocated into four groups: 2 g L-glutamine, 2 g L-carnitine, 2 g L-carnitine + 2 g L-glutamine and placebo. Supplements were prescribed for 21 days and after three weeks, athletes' performances and body composition were re-evaluated. Results: The results showed that body weight, body fat percentage, lean muscle mass, and dietary intake made no significant changes in different groups of athletes. In between groups comparison, results did not significantly change in any performance indices. However, in L-carnitine supplement group, the results of pre and post intervention showed that the running distance and maximal oxygen uptake (VO2max increased significantly while the subjective sense of fatigue decreased significantly. Conclusions: Based on our findings, a three-week prescription of separateor combined glutamine and L-carnitine, had no effects on body composition or dietary intake in soccer players. But, the athletes' energy intake was more than the one reported in other studies. Although further studies are required to assess these effects on athletic performance.

  12. Growth hormone enhances fat-free mass and glutamine availability in patients with short-bowel syndrome: an ancillary double-blind, randomized crossover study.

    Seguy, David; Darmaun, Dominique; Duhamel, Alain; Thuillier, François; Cynober, Luc; Cortot, Antoine; Gottrand, Frédéric; Messing, Bernard

    2014-09-01

    Benefits of recombinant human growth hormone (rhGH) alone or combined with glutamine in patients with intestinal failure because of short-bowel syndrome remain controversial. We explored effects of rhGH on whole-body protein metabolism in patients with short-bowel syndrome with intestinal failure (SBS-IF) to gain insight into its mechanism of action. Eight stable hyperphagic patients with severe SBS-IF received, in a double-blind, randomized crossover study, low-dose rhGH (0.05 mg · kg⁻¹ · d⁻¹) and a placebo for two 3-wk periods. Leucine and glutamine kinetics under fasting and fed conditions, fat-free mass (FFM), and serum insulin were determined on the final day of each treatment. rhGH increased FFM and nonoxidative leucine disposal (NOLD; an index of protein synthesis) (P de novo synthesis (P de novo synthesis and intestinal absorption increase glutamine availability over the physiologic range, suggesting that beneficial effects of rhGH in hyperphagic patients might be achieved without glutamine supplementation. © 2014 American Society for Nutrition.

  13. Examination of the efficacy of acute L-alanyl-L-glutamine ingestion during hydration stress in endurance exercise

    Yamamoto Linda M

    2010-02-01

    Full Text Available Abstract Background The effect of acute L-alanyl-L-glutamine (AG; Sustamine™ ingestion on performance changes and markers of fluid regulation, immune, inflammatory, oxidative stress, and recovery was examined in response to exhaustive endurance exercise, during and in the absence of dehydration. Methods Ten physically active males (20.8 ± 0.6 y; 176.8 ± 7.2 cm; 77.4 ± 10.5 kg; 12.3 ± 4.6% body fat volunteered to participate in this study. During the first visit (T1 subjects reported to the laboratory in a euhydrated state to provide a baseline (BL blood draw and perform a maximal exercise test. In the four subsequent randomly ordered trials, subjects dehydrated to -2.5% of their baseline body mass. For T2, subjects achieved their goal weight and were not rehydrated. During T3 - T5, subjects reached their goal weight and then rehydrated to 1.5% of their baseline body mass by drinking either water (T3 or two different doses (T4 and T5 of the AG supplement (0.05 g·kg-1 and 0.2 g·kg-1, respectively. Subjects then exercised at a workload that elicited 75% of their VO2 max on a cycle ergometer. During T2 - T5 blood draws occurred once goal body mass was achieved (DHY, immediately prior to the exercise stress (RHY, and immediately following the exercise protocol (IP. Resting 24 hour (24P blood samples were also obtained. Blood samples were analyzed for glutamine, potassium, sodium, aldosterone, arginine vasopressin (AVP, C-reactive protein (CRP, interleukin-6 (IL-6, malondialdehyde (MDA, testosterone, cortisol, ACTH, growth hormone and creatine kinase. Statistical evaluation of performance, hormonal and biochemical changes was accomplished using a repeated measures analysis of variance. Results Glutamine concentrations for T5 were significantly higher at RHY and IP than T2 - T4. When examining performance changes (difference between T2 - T5 and T1, significantly greater times to exhaustion occurred during T4 (130.2 ± 340.2 sec and T5 (157.4

  14. Ammonium assimilation in rice based on the occurrence of 15N and inhibition of glutamine synthetase activity

    Magalhaes, J. R.; Huber, D. M.; Lee, T. C.; Tsai, C. Y.

    1995-07-01

    Assimilation of ammonium (NH4) into free amino acids and total reduced nitrogen (N) was monitored in both roots and shoots of two-week old rice seedlings supplied with 5 mM 99% (15NH4)2SO4 in aerated hydroponic culture with or without a 2 h preincubation with 1 mM methionine sulfoximine (MSX) an inhibitor of glutamine synthetase (GS) activity. 15NH4 was not assimilated into amino acids when the GS/GOGAT (glutamate synthase) cycle was inhibited by MSX. Inhibition of glutamine synthetase (GS) activity in roots with MSX increased both the amount of NH4 and the abundance of 15N labeled NH4. In contrast, the amount of Gln and Glu, and their proportions as 15N, decreased in roots when GS activity was inhibited. This research confirms the importance of GS/GOGAT in NH4 assimilation in rice roots. 15N-labeled studies indicate that NH4 ions incorporated by roots of rice are transformed primarily into glutamine (Gin) and glutamic acid (Glu) before being converted to other amino acids through transamination. The formation of amino acids such as aspartic acid (Asp) and alanine (Ala) directly from free NH4 in roots also has been reported. Translocation of free NH4 to plant shoots, based on the concentration of free NH4 in xylem exudate, has been reported in tomato, although NH4 in shoots primarily originates from nitrate reduction in the shoot. Photorespiration also can contribute to the accumulation of NH4 in leaves. The GS/GOGAT cycle appears to be primarily responsible for the assimilation of exogenously supplied NH4 and NH4 derived from nitrate reduction in leaves, as well as NH4 derived from photorespiration. Genetic evidence cited to support this conclusion includes the lethal effect of photorespiratory conditions on plant mutants deficient in chloroplast-localized GS and GOGAT activities, and the rapid accumulation of free NH4 in GS-deficient mutants under photorespiratory conditions. The present study was initiated to quantify the in vivo amino acid synthesis in rice

  15. Ammonium assimilation in rice based on the occurrence of 15N and inhibition of glutamine synthetase activity

    Magalhaes, J.R.; Huber, D.M.; Lee, T.C.; Tsai, C.Y.

    1995-01-01

    Assimilation of ammonium (NH4) into free amino acids and total reduced nitrogen (N) was monitored in both roots and shoots of two-week old rice seedlings supplied with 5 mM 99% (15NH4)2SO4 in aerated hydroponic culture with or without a 2 h preincubation with 1 mM methionine sulfoximine (MSX) an inhibitor of glutamine synthetase (GS) activity. 15NH4 was not assimilated into amino acids when the GS/GOGAT (glutamate synthase) cycle was inhibited by MSX. Inhibition of glutamine synthetase (GS) activity in roots with MSX increased both the amount of NH4 and the abundance of 15N labeled NH4. In contrast, the amount of Gln and Glu, and their proportions as 15N, decreased in roots when GS activity was inhibited. This research confirms the importance of GS/GOGAT in NH4 assimilation in rice roots. 15N-labeled studies indicate that NH4 ions incorporated by roots of rice are transformed primarily into glutamine (Gin) and glutamic acid (Glu) before being converted to other amino acids through transamination. The formation of amino acids such as aspartic acid (Asp) and alanine (Ala) directly from free NH4 in roots also has been reported. Translocation of free NH4 to plant shoots, based on the concentration of free NH4 in xylem exudate, has been reported in tomato, although NH4 in shoots primarily originates from nitrate reduction in the shoot. Photorespiration also can contribute to the accumulation of NH4 in leaves. The GS/GOGAT cycle appears to be primarily responsible for the assimilation of exogenously supplied NH4 and NH4 derived from nitrate reduction in leaves, as well as NH4 derived from photorespiration. Genetic evidence cited to support this conclusion includes the lethal effect of photorespiratory conditions on plant mutants deficient in chloroplast-localized GS and GOGAT activities, and the rapid accumulation of free NH4 in GS-deficient mutants under photorespiratory conditions. The present study was initiated to quantify the in vivo amino acid synthesis in rice

  16. [2,4-(13)C]β-hydroxybutyrate metabolism in astrocytes and C6 glioblastoma cells.

    Eloqayli, Haytham; Melø, Torun M; Haukvik, Anne; Sonnewald, Ursula

    2011-08-01

    This study was undertaken to determine if the ketogenic diet could be useful for glioblastoma patients. The hypothesis tested was whether glioblastoma cells can metabolize ketone bodies. Cerebellar astrocytes and C6 glioblastoma cells were incubated in glutamine and serum free medium containing [2,4-(13)C]β-hydroxybutyrate (BHB) with and without glucose. Furthermore, C6 cells were incubated with [1-(13)C]glucose in the presence and absence of BHB. Cell extracts were analyzed by mass spectrometry and media by (1)H magnetic resonance spectroscopy and HPLC. Using [2,4-(13)C]BHB and [1-(13)C]glucose it could be shown that C6 cells, in analogy to astrocytes, had efficient mitochondrial activity, evidenced by (13)C labeling of glutamate, glutamine and aspartate. However, in the presence of glucose, astrocytes were able to produce and release glutamine, whereas this was not accomplished by the C6 cells, suggesting lack of anaplerosis in the latter. We hypothesize that glioblastoma cells kill neurons by not supplying the necessary glutamine, and by releasing glutamate.

  17. Comparison of the eight weeks of supplementation Creatine and Glutamine consumption along with resistance exercise on the level of ALP in female mice

    A eskandari

    2015-11-01

    Full Text Available Background and purpose: in recent years, in order to improve power, speed, the increase in the volume of the musculature, preventing sports injuries and maintain the muscle performance athletes use from different resistance exercises and food supplements. In this regard, present study has been conducted with the aim of comparison the influence of an 8 week period consumption of creatine (2 gr.kg-1.day-1 in 1st week and 0.48 gr.kg-1.day-1during 2nd to 8th weeks and glutamine (1 gr.kg-1.day-1 from first to eighth weeks along with resistance exercise on level of ALP of female mice. Materials and methods: This experimental study was done on 80 Small adult female mice of Surrey species (28 ± 5 gram. The animals were randomly divided into 8 groups of: resistance exercise, resistance exercise + creatine, resistance exercise + glutamine, resistance exercise + glutamine + creatine, creatine, glutamine, creatine + glutamine and control groups (N= 10. Resistance exercise (5 days a week was including: climbing (4 sets, 5 times repetition with two minutes rest between the sets from a ladder (with the height of one meter and including 26 steps and bearing 30 percent of the weight of the Mouse body (hanging from tail in the first week and the increasing it up to 200 percent of body weight till the last week of the experiment. During 48 hours after the last practice session of resistance exercise, the blood sample was taken and the the level of ALP has been measured. Findings:The results showed that the level of ALP enzyme in creatine + glutamine + resistance exercise groug had been increased in comparison with the control group (144.3 ± 15.86 in comparison with 234.7 ± 25.69 U.L-1 P < 0.05. Conclusion: The results of this research indicate Creatine and Glutamine supplementation consumption along with resistance exercise increases in the level of ALP enzyme in the liver of mice.

  18. A meta-analysis of trials using the intention to treat principle for glutamine supplementation in critically ill patients with burn.

    Lin, Jiun-Jie; Chung, Xiu-Juan; Yang, Chung-Yih; Lau, Hui-Ling

    2013-06-01

    During critical illness, the demand for glutamine may exceed that which can be mobilized from muscle stores. Infections increase mortality, morbidity, length-of-stay, antibiotic usage and the cost of care. This is a major health care issue. RCTs were identified from the electronic databases: the Cochrane Library, MEDLINE, PubMed web of knowledge and hand searching journals. The trials compared the supplementation with glutamine and non-supplementation in burn. Statistical analysis was performed using RevMan5.1 software, from Cochrane Collaboration. 216 papers showed a match, in the keyword search. Upon screening the title, reading the abstract and the entire article, only four RCTs, involving 155 patients, were included. For both the glutamine group and control group, total burn surface area (TBSA) (MD=2.02, 95% CI -2.17, 6.21, p=0.34) was similar. Glutamine supplementation was associated with a statistically significant decrease in the number of patients with gram-negative bacteremia (OR 0.27, 95% CI 0.08-0.92, p=0.04) and hospital mortality (OR=0.13, 95% CI 0.03, 0.51, p=0.004), however, no statistical difference was noted between groups, for the other results. Glutamine supplemented nutrition can be associated with a reduction in mortality in hospital, complications due to gram-negative bacteremia in burn patients. Further larger and better quality trials are required, in order to determine whether any differences are statistically and clinically important. Copyright © 2012 Elsevier Ltd and ISBI. All rights reserved.

  19. Conformational changes induced by Mg2+ on the multiple forms of glutamine synthetase from Bacillus brevis Bb G1

    Suja Abraham

    2013-01-01

    Full Text Available Conformational changes play an important role in the function of proteins. Glutamine synthetase, an important enzyme of nitrogen metabolism, was purified under sporulating (GSala and non-sporulating (GSpyr conditions and the effect of Mg2+ on these multiple forms was studied by fluorescence spectroscopy to detect possible conformational changes that occur in the presenceof Mg2+. The substantial changes in the fluorescence emission maximum, fluorescence intensity and lifetime that occur in the presence of different concentrations of Mg2+, indicated major changes in molecular conformations in both forms of this enzyme. The fluorescent changes produced by the effect of Mg2+ in GSala was much more prominent than in GSpyr. These observations strongly support the possibility that GSala and GSpyr undergoes a conformational change on binding with Mg2+.

  20. The transcriptional activator NtrC controls the expression and activity of glutamine synthetase in Herbaspirillum seropedicae.

    Persuhn, D C; Souza, E M; Steffens, M B; Pedrosa, F O; Yates, M G; Rigo, L U

    2000-11-15

    The role of the Ntr system in Herbaspirillum seropedicae was determined via ntrB and ntrC mutants. Three phenotypes were identified in these mutants: Nif(-), deficiency in growth using nitrate, and low glutamine synthetase (GS) activity. All phenotypes were restored by the plasmid pKRT1 containing the intact glnA, ntrB and ntrC genes of H. seropedicae. The promoter region of glnA was subcloned into a beta-galactosidase fusion vector and the results suggested that NtrC positively regulates the glnA promoter in response to low nitrogen. The H. seropedicae ntrC and ntrB mutant strains showed a deficiency of adenylylation/deadenylylation of GS, indicating that NtrC and NtrB are involved in both transcription and activity control of GS in this organism.

  1. Functional specialization of one copy of glutamine phosphoribosyl pyrophosphate amidotransferase in ureide production from symbiotically fixed nitrogen in Phaseolus vulgaris.

    Coleto, Inmaculada; Trenas, Almudena T; Erban, Alexander; Kopka, Joachim; Pineda, Manuel; Alamillo, Josefa M

    2016-08-01

    Purines are essential molecules formed in a highly regulated pathway in all organisms. In tropical legumes, the nitrogen fixed in the nodules is used to generate ureides through the oxidation of de novo synthesized purines. Glutamine phosphoribosyl pyrophosphate amidotransferase (PRAT) catalyses the first committed step of de novo purine synthesis. In Phaseolus vulgaris there are three genes coding for PRAT. The three full-length sequences, which are intron-less genes, were cloned, and their expression levels were determined under conditions that affect the synthesis of purines. One of the three genes, PvPRAT3, is highly expressed in nodules and protein amount and enzymatic activity in these tissues correlate with nitrogen fixation activity. Inhibition of PvPRAT3 gene expression by RNAi-silencing and subsequent metabolomic analysis of the transformed roots shows that PvPRAT3 is essential for the synthesis of ureides in P. vulgaris nodules. © 2016 John Wiley & Sons Ltd.

  2. The neuropharmacology of L-theanine(N-ethyl-L-glutamine): a possible neuroprotective and cognitive enhancing agent.

    Nathan, Pradeep J; Lu, Kristy; Gray, M; Oliver, C

    2006-01-01

    L-theanine (N-ethyl-L-glutamine) or theanine is a major amino acid uniquely found in green tea. L-theanine has been historically reported as a relaxing agent, prompting scientific research on its pharmacology. Animal neurochemistry studies suggest that L-theanine increases brain serotonin, dopamine, GABA levels and has micromolar affinities for AMPA, Kainate and NMDA receptors. In addition has been shown to exert neuroprotective effects in animal models possibly through its antagonistic effects on group 1 metabotrophic glutamate receptors. Behavioural studies in animals suggest improvement in learning and memory. Overall, L-theanine displays a neuropharmacology suggestive of a possible neuroprotective and cognitive enhancing agent and warrants further investigation in animals and humans.

  3. A diet containing whey protein, glutamine, and TGFbeta modulates gut protein metabolism during chemotherapy-induced mucositis in rats.

    Boukhettala, Nabile; Ibrahim, Ayman; Claeyssens, Sophie; Faure, Magali; Le Pessot, Florence; Vuichoud, Jacques; Lavoinne, Alain; Breuillé, Denis; Déchelotte, Pierre; Coëffier, Moïse

    2010-08-01

    Mucositis, a common side effect of chemotherapy, is characterized by compromised digestive function, barrier integrity and immune competence. Our aim was to evaluate the impact of a specifically designed diet Clinutren Protect (CP), which contains whey proteins, TGFbeta-rich casein, and free glutamine, on mucositis in rats. Mucositis was induced by three consecutive injections (day 0, day 1, day 2) of methotrexate (2.5 mg/kg). Rats had free access to CP or placebo diets from days -7 to 9. In the placebo diet, whey proteins and TGFbeta-rich casein were replaced by TGFbeta-free casein and glutamine by alanine. Intestinal parameters were assessed at day 3 and 9. Values, expressed as mean +/- SEM, were compared using two-way ANOVA. At day 3, villus height was markedly decreased in the placebo (296 +/- 11 microm) and CP groups (360 +/- 10 microm) compared with controls (464 +/- 27 microm), but more markedly in the placebo as compared to CP group. The intestinal damage score was also reduced in the CP compared with the placebo group. Glutathione content increased in the CP compared with the placebo group (2.2 +/- 0.2 vs. 1.7 +/- 0.2 micromol/g tissue). Gut protein metabolism was more affected in the placebo than in the CP group. The fractional synthesis rate was decreased in the placebo group (93.8 +/- 4.9%/day) compared with controls (121.5 +/- 12.1, P < 0.05), but not in the CP group (106.0 +/- 13.1). In addition, at day 9, rats exhibited improved body weight and food intake recovery in the CP compared to the placebo group. Clinutren Protect feeding reduces intestinal injury in the acute phase of methotrexate-induced mucositis in rats and improves recovery.

  4. Combined glutamate and glutamine levels in pain-processing brain regions are associated with individual pain sensitivity.

    Zunhammer, Matthias; Schweizer, Lauren M; Witte, Vanessa; Harris, Richard E; Bingel, Ulrike; Schmidt-Wilcke, Tobias

    2016-10-01

    The relationship between glutamate and γ-aminobutyric acid (GABA) levels in the living human brain and pain sensitivity is unknown. Combined glutamine/glutamate (Glx), as well as GABA levels can be measured in vivo with single-voxel proton magnetic resonance spectroscopy. In this cross-sectional study, we aimed at determining whether Glx and/or GABA levels in pain-related brain regions are associated with individual differences in pain sensitivity. Experimental heat, cold, and mechanical pain thresholds were obtained from 39 healthy, drug-free individuals (25 men) according to the quantitative sensory testing protocol and summarized into 1 composite measure of pain sensitivity. The Glx levels were measured using point-resolved spectroscopy at 3 T, within a network of pain-associated brain regions comprising the insula, the anterior cingulate cortex, the mid-cingulate cortex, the dorsolateral prefrontal cortex, and the thalamus. GABA levels were measured using GABA-edited spectroscopy (Mescher-Garwood point-resolved spectroscopy) within the insula, the anterior cingulate cortex, and the mid-cingulate cortex. Glx and/or GABA levels correlated positively across all brain regions. Gender, weekly alcohol consumption, and depressive symptoms were significantly associated with Glx and/or GABA levels. A linear regression analysis including all these factors indicated that Glx levels pooled across pain-related brain regions were positively associated with pain sensitivity, whereas no appreciable relationship with GABA was found. In sum, we show that the levels of the excitatory neurotransmitter glutamate and its precursor glutamine across pain-related brain regions are positively correlated with individual pain sensitivity. Future studies will have to determine whether our findings also apply to clinical populations.

  5. The association between glutamine repeats in the androgen receptor gene and personality traits in dromedary camel (Camelus dromedarius).

    Ramadan, Sherif; Nowier, Amira M; Hori, Yusuke; Inoue-Murayama, Miho

    2018-01-01

    Temperament traits such as fearfulness are important as they define an animal's responses to its environment and handling. The increasing automation of daily tasks and growing population limits contact between camels and humans. Such limitations contribute to fear of humans and changes in physical environment. Monoamine oxidase A (MAOA) and androgen receptor (AR) genes are important candidates associated with mammal personality. In our analysis, MAOA exon 15 showed no polymorphism but a novel polymorphism was seen in the camel AR exon 1; 16, 17, 18, and 19 glutamine repeats were detected. We genotyped 138 camels belonging to four Egyptian breeds: Maghrabi (n = 90), Sudani (n = 15), Somali (n = 23), and Baladi (n = 10) for AR. Out of the 90 genotyped Maghrabi camels, we evaluated responses of 33 and 32 mature females to a novel object and exposure to an unfamiliar person, respectively. AR gene showed a significant association based on the principal component (PC) score, which indicated the fear of human touch, and the PC score indicates fear during interaction with novel objects. Individuals carrying a shorter genotype in homozygote (S/S) were found to be more fearful. Furthermore, we found that Sudani and Somali breeds had a higher frequency of shorter genotype (S/S), which was associated with increased fearfulness. These findings reflect the behavioral tendency and consequently, affect the use of this breed. This is the first report showing the role of AR glutamine repeats influencing a behavioral trait in dromedary camels and leading to inter-breed differences. Fear-related traits reported here are important because camels cope with various types of stresses and fear, resulting from the demands of intensive production systems and racing events. However, further studies, employing functional genomics and linkage analysis are necessary for confirming the relationship between fearfulness and genetic variation.

  6. The association between glutamine repeats in the androgen receptor gene and personality traits in dromedary camel (Camelus dromedarius.

    Sherif Ramadan

    Full Text Available Temperament traits such as fearfulness are important as they define an animal's responses to its environment and handling. The increasing automation of daily tasks and growing population limits contact between camels and humans. Such limitations contribute to fear of humans and changes in physical environment. Monoamine oxidase A (MAOA and androgen receptor (AR genes are important candidates associated with mammal personality. In our analysis, MAOA exon 15 showed no polymorphism but a novel polymorphism was seen in the camel AR exon 1; 16, 17, 18, and 19 glutamine repeats were detected. We genotyped 138 camels belonging to four Egyptian breeds: Maghrabi (n = 90, Sudani (n = 15, Somali (n = 23, and Baladi (n = 10 for AR. Out of the 90 genotyped Maghrabi camels, we evaluated responses of 33 and 32 mature females to a novel object and exposure to an unfamiliar person, respectively. AR gene showed a significant association based on the principal component (PC score, which indicated the fear of human touch, and the PC score indicates fear during interaction with novel objects. Individuals carrying a shorter genotype in homozygote (S/S were found to be more fearful. Furthermore, we found that Sudani and Somali breeds had a higher frequency of shorter genotype (S/S, which was associated with increased fearfulness. These findings reflect the behavioral tendency and consequently, affect the use of this breed. This is the first report showing the role of AR glutamine repeats influencing a behavioral trait in dromedary camels and leading to inter-breed differences. Fear-related traits reported here are important because camels cope with various types of stresses and fear, resulting from the demands of intensive production systems and racing events. However, further studies, employing functional genomics and linkage analysis are necessary for confirming the relationship between fearfulness and genetic variation.

  7. Medicago truncatula contains a second gene encoding a plastid located glutamine synthetase exclusively expressed in developing seeds

    Seabra Ana R

    2010-08-01

    Full Text Available Abstract Background Nitrogen is a crucial nutrient that is both essential and rate limiting for plant growth and seed production. Glutamine synthetase (GS, occupies a central position in nitrogen assimilation and recycling, justifying the extensive number of studies that have been dedicated to this enzyme from several plant sources. All plants species studied to date have been reported as containing a single, nuclear gene encoding a plastid located GS isoenzyme per haploid genome. This study reports the existence of a second nuclear gene encoding a plastid located GS in Medicago truncatula. Results This study characterizes a new, second gene encoding a plastid located glutamine synthetase (GS2 in M. truncatula. The gene encodes a functional GS isoenzyme with unique kinetic properties, which is exclusively expressed in developing seeds. Based on molecular data and the assumption of a molecular clock, it is estimated that the gene arose from a duplication event that occurred about 10 My ago, after legume speciation and that duplicated sequences are also present in closely related species of the Vicioide subclade. Expression analysis by RT-PCR and western blot indicate that the gene is exclusively expressed in developing seeds and its expression is related to seed filling, suggesting a specific function of the enzyme associated to legume seed metabolism. Interestingly, the gene was found to be subjected to alternative splicing over the first intron, leading to the formation of two transcripts with similar open reading frames but varying 5' UTR lengths, due to retention of the first intron. To our knowledge, this is the first report of alternative splicing on a plant GS gene. Conclusions This study shows that Medicago truncatula contains an additional GS gene encoding a plastid located isoenzyme, which is functional and exclusively expressed during seed development. Legumes produce protein-rich seeds requiring high amounts of nitrogen, we postulate

  8. N-Linked Glycosylation is an Important Parameter for Optimal Selection of Cell Lines Producing Biopharmaceutical Human IgG

    van Berkel, Patrick H. C.; Gerritsen, Jolanda; Perdok, Gerrard; Valbjorn, Jesper; Vink, Tom; van de Winkel, Jan G. J.; Parren, Paul W. H. I.

    2009-01-01

    We studied the variations in N-linked glycosylation of human IgG molecules derived from 105 different stable cell lines each expressing one of the six different antibodies. Antibody expression was based on glutamine synthetase selection technology in suspension growing CHO-KISV cells. The glycans

  9. Effect of nerve growth factor on the synthesis of amino acids in PC12 cells

    Zielke, H.R.; Tildon, J.T.; Kauffman, F.C.; Baab, P.J.

    1989-01-01

    Radioactive short-chain fatty acids preferentially label glutamine relative to glutamate in brain due to compartmentation of glutamine and glutamate. To determine whether this phenomenon occurs in a single cell culture model, we examined the effect of fatty acid chain length on the synthesis as well as pool size of selected amino acids in rat pheochromocytoma PC12 cells, a cell culture model of the large glutamate compartment in neurons. Intracellular 14C-amino acids were quantitated by HPLC, and the incorporation of [U-14C]-glucose, [1-14C]-butyrate, [1-14C]-octanoate, and [1-14C]-palmitate into five amino acids was measured in native and NGF-treated PC12 cells. NGF pretreatment decreased the intracellular concentration of amino acids as did addition of fatty acids but these effects were not additive. Specific activities of amino acids in native cells labelled by 14C-octanoate were 1,300 DPM/nmol, 490 DPM/nmol, 200 DPM/nmol, and 110 DPM/nmol for glutamate, aspartate, glutamine, and serine, respectively. No radioactivity was detected in alanine. Similar specific activities were noted when 14C-butyrate was the precursor; however, there was at least 5-fold less if 14C-palmitate was the precursor. Pretreatment of cells with NGF decreased the specific activity of amino acids by 25-65%. Specific activities of amino acids synthesized from 14C-glucose decreased in the following order: glutamate, 1,640 DPM/nmol; aspartate, 1,210 DPM/nmol; alanine, 580 DPM/nmol; glutamine, 275 DPM/nmol; and serine, 80 DPM/nmol for native cells. NGF pretreatment decreased the specific activities of glutamate and glutamine, but not of the other 3 amino acids. The preferred precursor for glutamate synthesis in native PC12 cells was glucose followed by octanoate, butyrate and palmitate (16:6:3:1)

  10. Efeito da suplementação de glutamina sobre variáveis hematológicas e do estado nutricional de ratas desnutridas Glutamine supplementation effect in hematological parameters and nutritional status of rats submitted to malnutrition protocol

    Natália Cristina Lima Rodrigues

    2007-12-01

    diet. METHODS: Total of 20 Wistar females rats, 21 days of age, were submitted or not to the protein-calorie malnutrition induced by diarrhea, using an enriched lactose diet 60% for 15 days. After malnutrition period, eutrophic or malnourished animals remained for 30 days provided with a variety diet (standard and experimental. According to nutritional status and treatment, animals were randomized into four distinct groups: 1 GC: animals eutrophic + standard diet, 2 GD: malnourished animals + enriched lactose diet (60%, 3 GDGLN: malnourished animals + enriched glutamine diet (2%; and 4 GDP: malnourished animals + standard diet. After the 30 days of nutritional treatment, blood samples were obtained by cardiac puncture for biochemical (total protein, albumin and urea and hematological evaluations (red and white blood cells quantification. RESULTS: After 15 days of experimental study, weight gain in the respective groups GD (46.4 ± 2.60 g, GDGLN (39.2 ± 8.9 g and GDP (33.2 ± 11.5 g was lower once compared to the control (64 ± 4.24 g, P <0.05. Biochemical examinations showed a significant difference in the urea dosage in GD as compared to other groups (33.4 ± 4.77 mg / dL, P<0.05. The leukocytes counting of GD (3.68 ± 1.0 cel/mm3 x103 showed reduced comparing to the rest of the groups (P<0.05, while red blood cell counting had not presented significant difference. CONCLUSION: Malnutrition affects leukocytes cell number, although such deleterious counting blood effect can be reversed by simple re-administration of nutrition, regardless of glutamine supplementation. The intake of oral glutamine does not affect the weight gain, biochemical variables of nutritional status, or leukocytes number while malnourished due to enriched lactose diet.

  11. The Aspergillus niger MADS-box transcription factor RlmA is required for cell wall reinforcement in response to cell wall stress.

    Damveld, R.A.; Arentshorst, M.; Franken, A.; Vankuyk, P.A.; Klis, F.M.; van den Hondel, C.A.; Ram, A.F.

    2005-01-01

    In Aspergillus niger, the genes coding for glutamine:fructose-6-phosphate amidotransferase (gfaA) and ¿-1,3-glucan synthase (agsA) are induced in response to cell wall stress. In silico analysis of the promoter region of the two genes revealed the presence of putative DNA binding sites for

  12. Ataxin-1 with an expanded glutamine tract alters nuclear matrix-associated structures

    Skinner, P J; Koshy, B T; Cummings, C J

    1997-01-01

    a similar pattern of nuclear localization; with expanded ataxin-1 occurring in larger structures that are fewer in number than those of normal ataxin-1. Colocalization studies show that mutant ataxin-1 causes a specific redistribution of the nuclear matrix-associated domain containing promyelocytic...... the subcellular localization of wild-type human ataxin-1 (the protein encoded by the SCA1 gene) and mutant ataxin-1 in the Purkinje cells of transgenic mice. We found that ataxin-1 localizes to the nuclei of cerebellar Purkinje cells. Normal ataxin-1 localizes to several nuclear structures approximately 0.......5 microm across, whereas the expanded ataxin-1 localizes to a single approximately 2-microm structure, before the onset of ataxia. Mutant ataxin-1 localizes to a single nuclear structure in affected neurons of SCA1 patients. Similarly, COS-1 cells transfected with wild-type or mutant ataxin-1 show...

  13. Activity of the lactate-alanine shuttle is independent of glutamate-glutamine cycle activity in cerebellar neuronal-astrocytic cultures

    Bak, Lasse K; Sickmann, Helle M; Schousboe, Arne

    2004-01-01

    The glutamate-glutamine cycle describes the neuronal release of glutamate into the synaptic cleft, astrocytic uptake, and conversion into glutamine, followed by release for use as a neuronal glutamate precursor. This only explains the fate of the carbon atoms, however, and not that of the ammonia....... Recently, a role for alanine has been proposed in transfer of ammonia between glutamatergic neurons and astrocytes, denoted the lactate-alanine shuttle (Waagepetersen et al. [ 2000] J. Neurochem. 75:471-479). The role of alanine in this context has been studied further using cerebellar neuronal cultures...... and corresponding neuronal-astrocytic cocultures. A superfusion paradigm was used to induce repetitively vesicular glutamate release by N-methyl-D-aspartate (NMDA) in the neurons, allowing the relative activity dependency of the lactate-alanine shuttle to be assessed. [(15)N]Alanine (0.2 mM), [2-(15)N]/[5-(15)N...

  14. Specificity of exogenous acetate and glutamate as astrocyte substrates examined in acute brain slices from female mice using methionine sulfoximine (MSO) to inhibit glutamine synthesis

    Andersen, Jens Velde; McNair, Laura Frendrup; Schousboe, Arne

    2017-01-01

    Removal of endogenously released glutamate is mediated primarily by astrocytes and exogenous (13) C-labeled glutamate has been applied to study glutamate metabolism in astrocytes. Likewise, studies have clearly established the relevance of (13) C-labeled acetate as an astrocyte specific metabolic...... cortical slices from female NMRI mice were incubated in media containing [1,2-(13) C]acetate or [U-(13) C]glutamate, with or without methionine sulfoximine (MSO) to inhibit glutamine synthetase (GS). Tissue extracts were analyzed by gas chromatography-mass spectrometry. Blocking GS abolished the majority...... of glutamine (13) C-labeling from [1,2-(13) C]acetate as intended. However, (13) C-labeling of GABA was only 40-50% reduced by MSO, suggesting considerable neuronal uptake of acetate. Moreover, labeling of glutamate from [1,2-(13) C]acetate in the presence of MSO exceeded the level probable from exclusive...

  15. Software sensors as a tool for optimization of animal-cell cultures

    Dorresteijn, R.C.

    1997-01-01

    In this thesis software sensors are introduced that predict the biomass activity and the concentrations of glucose, glutamine, lactic acid, and ammonium on line, The software sensors for biomass activity, glucose and lactic acid can be applied for any type of animal cell that is grown in a

  16. Application of CRISPR/Cas9 Genome Editing to Improve Recombinant Protein Production in CHO Cells

    Grav, Lise Marie; Julie la Cour Karottki, Karen; Lee, Jae Seong

    2017-01-01

    and yields. In this chapter, we present our protocol on how to use the genome editing tool Clustered Regularly Interspaced Short Palindromic Repeat (CRISPR)/CRISPR-associated protein 9 (Cas9) to knockout engineering target genes in CHO cells. As an example, we refer to the glutamine synthetase (GS...

  17. Regulation of glutamine synthetase isoforms in two differentially drought-tolerant rice (Oryza sativa L.) cultivars under water deficit conditions.

    Singh, Kamal Krishna; Ghosh, Shilpi

    2013-02-01

    KEY MESSAGE : The regulation of GS isoforms by WD was organ specific. Two GS isoforms i.e. OsGS1;1 and OsGS2 were differentially regulated in IR-64 (drought-sensitive) and Khitish (drought-tolerant) cultivars of rice. Water deficit (WD) has adverse effect on rice (Oryza sativa L.) and acclimation requires essential reactions of primary metabolism to continue. Rice plants utilize ammonium as major nitrogen source, which is assimilated into glutamine by the reaction of Glutamine synthetase (GS, EC 6.3.1.2). Rice plants possess one gene (OsGS2) for chloroplastic GS2 and three genes (OsGS1;1, OsGS1;2 and OsGS1;3) for cytosolic GS1. Here, we report the effect of WD on regulation of GS isoforms in drought-sensitive (cv. IR-64) and drought-tolerant (cv. Khitish) rice cultivars. Under WD, total GS activity in root and leaf decreased significantly in IR-64 seedlings in comparison to Khitish seedlings. The reduced GS activity in IR-64 leaf was mainly due to decrease in GS2 activity, which correlated with decrease in corresponding transcript and polypeptide contents. GS1 transcript and polypeptide accumulated in leaf during WD, however, GS1 activity was maintained at a constant level. Total GS activity in stem of both the varieties was insensitive to WD. Among GS1 genes, OsGS1;1 expression was differently regulated by WD in the two rice varieties. Its transcript accumulated more abundantly in IR-64 leaf than in Khitish leaf. Following WD, OsGS1;1 mRNA level in stem and root tissues declined in IR-64 and enhanced in Khitish. A steady OsGS1;2 expression patterns were noted in leaf, stem and root of both the cultivars. Results suggest that OsGS2 and OsGS1;1 expression may contribute to drought tolerance of Khitish cultivar under WD conditions.

  18. Creatine, Glutamine plus Glutamate, and Macromolecules Are Decreased in the Central White Matter of Premature Neonates around Term.

    Meriam Koob

    Full Text Available Preterm birth represents a high risk of neurodevelopmental disabilities when associated with white-matter damage. Recent studies have reported cognitive deficits in children born preterm without brain injury on MRI at term-equivalent age. Understanding the microstructural and metabolic underpinnings of these deficits is essential for their early detection. Here, we used diffusion-weighted imaging and single-voxel 1H magnetic resonance spectroscopy (MRS to compare brain maturation at term-equivalent age in premature neonates with no evidence of white matter injury on conventional MRI except diffuse excessive high-signal intensity, and normal term neonates. Thirty-two infants, 16 term neonates (mean post-conceptional age at scan: 39.8±1 weeks and 16 premature neonates (mean gestational age at birth: 29.1±2 weeks, mean post-conceptional age at scan: 39.2±1 weeks were investigated. The MRI/MRS protocol performed at 1.5T involved diffusion-weighted MRI and localized 1H-MRS with the Point RESolved Spectroscopy (PRESS sequence. Preterm neonates showed significantly higher ADC values in the temporal white matter (P<0.05, the occipital white matter (P<0.005 and the thalamus (P<0.05. The proton spectrum of the centrum semiovale was characterized by significantly lower taurine/H2O and macromolecules/H2O ratios (P<0.05 at a TE of 30 ms, and reduced (creatine+phosphocreatine/H2O and (glutamine+glutamate/H2O ratios (P<0.05 at a TE of 135 ms in the preterm neonates than in full-term neonates. Our findings indicate that premature neonates with normal conventional MRI present a delay in brain maturation affecting the white matter and the thalamus. Their brain metabolic profile is characterized by lower levels of creatine, glutamine plus glutamate, and macromolecules in the centrum semiovale, a finding suggesting altered energy metabolism and protein synthesis.

  19. Molecular evolution of glutamine synthetase II: Phylogenetic evidence of a non-endosymbiotic gene transfer event early in plant evolution

    Tartar Aurélien

    2010-06-01

    Full Text Available Abstract Background Glutamine synthetase (GS is essential for ammonium assimilation and the biosynthesis of glutamine. The three GS gene families (GSI, GSII, and GSIII are represented in both prokaryotic and eukaryotic organisms. In this study, we examined the evolutionary relationship of GSII from eubacterial and eukaryotic lineages and present robust phylogenetic evidence that GSII was transferred from γ-Proteobacteria (Eubacteria to the Chloroplastida. Results GSII sequences were isolated from four species of green algae (Trebouxiophyceae, and additional green algal (Chlorophyceae and Prasinophytae and streptophyte (Charales, Desmidiales, Bryophyta, Marchantiophyta, Lycopodiophyta and Tracheophyta sequences were obtained from public databases. In Bayesian and maximum likelihood analyses, eubacterial (GSIIB and eukaryotic (GSIIE GSII sequences formed distinct clades. Both GSIIB and GSIIE were found in chlorophytes and early-diverging streptophytes. The GSIIB enzymes from these groups formed a well-supported sister clade with the γ-Proteobacteria, providing evidence that GSIIB in the Chloroplastida arose by horizontal gene transfer (HGT. Bayesian relaxed molecular clock analyses suggest that GSIIB and GSIIE coexisted for an extended period of time but it is unclear whether the proposed HGT happened prior to or after the divergence of the primary endosymbiotic lineages (the Archaeplastida. However, GSIIB genes have not been identified in glaucophytes or red algae, favoring the hypothesis that GSIIB was gained after the divergence of the primary endosymbiotic lineages. Duplicate copies of the GSIIB gene were present in Chlamydomonas reinhardtii, Volvox carteri f. nagariensis, and Physcomitrella patens. Both GSIIB proteins in C. reinhardtii and V. carteri f. nagariensis had N-terminal transit sequences, indicating they are targeted to the chloroplast or mitochondrion. In contrast, GSIIB proteins of P. patens lacked transit sequences, suggesting

  20. The impact of use of Glutamine on patients with head and neck tumors in radiotherapy and chemotherapy treatment; O Impacto do uso de Glutamina em pacientes com tumores de cabeca e pescoco em tratamento radioterapico e quimioterapico

    Boligon, Caroline Schardong, E-mail: caronut@bol.com.b [Hospital de Caridade de Ijui, RS (Brazil); Huth, Adriane, E-mail: adriane.huth@unijui.edu.b [UNIJUI, RS (Brazil). Dept. Ciencias da Saude

    2011-07-01

    Introduction: patients with head and neck neoplasia usually show malnutrition or a nutritional risk, because of common symptoms like: dysphagia, odynophagia and xerostomia. Objective: this study aimed to verify the impact of using amino glutamine in patients with head and neck neoplasia and under radiotherapy and chemotherapy treatment concomitantly. Methods: the research was quantitative, cross-sectional, descriptive and exploratory. The data was collected from nutritional evaluation, and patients chart consultation. The patients were divided in a control group (without use of glutamine) and a test group (with use of glutamine). 16 patients, 13 of which were men and three were women, participated in the research. Results: The control group presented mucositis grades I to IV while patients who used the amino glutamine showed mucositis grades I to II only. It could be observed that the Nutritional Risk Index decreased, which represents higher nutritional risk in patients from the control group only. In patients who used glutamine, this decrease was not significant. Conclusion: these results suggest that the use of glutamine in patients with head and neck tumors and under antineoplastic therapy helps to maintain their nutritional stage and to prevent mucositis throughout their treatment, mainly grades III and IV, which prevents adequate and regular eating and nourishment. (author)

  1. Effects of Supplementation with BCAA and L-glutamine on Blood Fatigue Factors and Cytokines in Juvenile Athletes Submitted to Maximal Intensity Rowing Performance.

    Koo, Ga Hee; Woo, Jinhee; Kang, Sungwhun; Shin, Ki Ok

    2014-08-01

    [Purpose] This study was conducted to understand the impacts of BCAA (branched-chain amino acid) and glutamine supplementation on the degree of blood fatigue factor stimulation and cytokines along with performance of exercise at the maximal intensity. [Subjects] Five male juvenile elite rowing athletes participated in this study as the subjects; they took 3 tests and received placebo supplementation (PS), BCAA supplementation (BS), and glutamine supplementation (GS). [Methods] The exercise applied in the tests was 2,000 m of rowing at the maximal intensity using an indoor rowing machine, and blood samples were collected 3 times, while resting, at the end of exercise, and after 30 min of recovery, to analyze the blood fatigue factors (lactate, phosphorous, ammonia, creatine kinase (CK)) and blood cytokines (IL (interleukin)-6, 8, 15). [Results] The results of the analysis showed that the levels of blood phosphorous in the BS and GS groups at the recovery stage were decreased significantly compared with at the end of exercise, and the level of CK appeared lower in the GS group alone at recovery stage than at the end of exercise. The level of blood IL-15 in the PS and BS groups appeared higher at the end of exercise compared with the resting stage. [Conclusion] It seemed that glutamine supplementation had a positive effect on the decrease in fatigue factor stimulation at the recovery stage after maximal intensity exercise compared with supplementation with the placebo or BCAA. Besides, pre-exercise glutamine supplementation seemed to help enhance immune function and the defensive inflammatory reaction.

  2. Age-dependent decrease in glutamine synthetase expression in the hippocampal astroglia of the triple transgenic Alzheimer's disease mouse model: Mechanism for deficient glutamatergic transmission?

    Olabarria, M.; Noristani, H. N.; Verkhratsky, Alexei; Rodríguez Arellano, Jose Julio

    2011-01-01

    Roč. 6, č. 1 (2011), s. 55-63 ISSN 1750-1326 R&D Projects: GA ČR GA309/09/1696; GA ČR(CZ) GAP304/11/0184; GA ČR GA305/08/1384; GA ČR GA309/08/1381 Institutional research plan: CEZ:AV0Z50390703 Keywords : astroglia * glutamine synthetase * Alzheimer ?'?s disease Subject RIV: FH - Neurology Impact factor: 4.278, year: 2011

  3. The effect of oral supplementation with a combination of beta-hydroxy-beta-methylbutyrate, arginine and glutamine on wound healing: a retrospective analysis of diabetic haemodialysis patients

    Sipahi, Savas; Gungor, Ozkan; Gunduz, Mehmet; Cilci, Mehmet; Demirci, Mustafa Cahit; Tamer, Ali

    2013-01-01

    Background Diabetes is an important reason for end-stage renal failure and diabetic foot wounds worsen the life qualities of these patients. Protein and amino acid support accelerates the wound healing. The purpose of this retrospective study is to examine the effect of beta-hydroxy-beta-methylbutyrate, arginine and glutamine (Abound?) supplementation on the wound healing. Methods A total of 11 diabetic dialysis patients were included in this retrospective study aiming to evaluate the effect ...

  4. Chronic probiotic supplementation with or without glutamine does not influence the eHsp72 response to a multi-day ultra-endurance exercise event.

    Marshall, Hannah; Chrismas, Bryna Catherine Rose; Suckling, Craig Anthony; Roberts, Justin D; Foster, Josh; Taylor, Lee

    2017-08-01

    Probiotic and glutamine supplementation increases tissue Hsp72, but their influence on extracellular Hsp72 (eHsp72) has not been investigated. The aim of this study was to investigate the effect of chronic probiotic supplementation, with or without glutamine, on eHsp72 concentration before and after an ultramarathon. Thirty-two participants were split into 3 independent groups, where they ingested probiotic capsules (PRO; n = 11), probiotic + glutamine powder (PGLn; n = 10), or no supplementation (CON; n = 11), over a 12-week period prior to commencement of the Marathon des Sables (MDS). eHsp72 concentration in the plasma was measured at baseline, 7 days pre-race, 6-8 h post-race, and 7 days post-race. The MDS increased eHsp72 concentrations by 124% (F [1,3] = 22.716, p 0.05). In conclusion, the MDS caused a substantial increase in eHsp72 concentration, indicating high levels of systemic stress. However, chronic PRO or PGLn supplementation did not affect eHsp72 compared with control pre- or post-MDS. Given the role of eHsp72 in immune activation, the commercially available supplements used in this study are unlikely to influence this cascade.

  5. Brain alanine formation as an ammonia-scavenging pathway during hyperammonemia: effects of glutamine synthetase inhibition in rats and astrocyte–neuron co-cultures

    Dadsetan, Sherry; Kukolj, Eva; Bak, Lasse K; Sørensen, Michael; Ott, Peter; Vilstrup, Hendrik; Schousboe, Arne; Keiding, Susanne; Waagepetersen, Helle S

    2013-01-01

    Hyperammonemia is a major etiological toxic factor in the development of hepatic encephalopathy. Brain ammonia detoxification occurs primarily in astrocytes by glutamine synthetase (GS), and it has been proposed that elevated glutamine levels during hyperammonemia lead to astrocyte swelling and cerebral edema. However, ammonia may also be detoxified by the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT) leading to trapping of ammonia in alanine, which in vivo likely leaves the brain. Our aim was to investigate whether the GS inhibitor methionine sulfoximine (MSO) enhances incorporation of 15NH4+ in alanine during acute hyperammonemia. We observed a fourfold increased amount of 15NH4 incorporation in brain alanine in rats treated with MSO. Furthermore, co-cultures of neurons and astrocytes exposed to 15NH4Cl in the absence or presence of MSO demonstrated a dose-dependent incorporation of 15NH4 into alanine together with increased 15N incorporation in glutamate. These findings provide evidence that ammonia is detoxified by the concerted action of GDH and ALAT both in vivo and in vitro, a mechanism that is accelerated in the presence of MSO thereby reducing the glutamine level in brain. Thus, GS could be a potential drug target in the treatment of hyperammonemia in patients with hepatic encephalopathy. PMID:23673435

  6. Brain alanine formation as an ammonia-scavenging pathway during hyperammonemia: effects of glutamine synthetase inhibition in rats and astrocyte-neuron co-cultures.

    Dadsetan, Sherry; Kukolj, Eva; Bak, Lasse K; Sørensen, Michael; Ott, Peter; Vilstrup, Hendrik; Schousboe, Arne; Keiding, Susanne; Waagepetersen, Helle S

    2013-08-01

    Hyperammonemia is a major etiological toxic factor in the development of hepatic encephalopathy. Brain ammonia detoxification occurs primarily in astrocytes by glutamine synthetase (GS), and it has been proposed that elevated glutamine levels during hyperammonemia lead to astrocyte swelling and cerebral edema. However, ammonia may also be detoxified by the concerted action of glutamate dehydrogenase (GDH) and alanine aminotransferase (ALAT) leading to trapping of ammonia in alanine, which in vivo likely leaves the brain. Our aim was to investigate whether the GS inhibitor methionine sulfoximine (MSO) enhances incorporation of (15)NH4(+) in alanine during acute hyperammonemia. We observed a fourfold increased amount of (15)NH4 incorporation in brain alanine in rats treated with MSO. Furthermore, co-cultures of neurons and astrocytes exposed to (15)NH4Cl in the absence or presence of MSO demonstrated a dose-dependent incorporation of (15)NH4 into alanine together with increased (15)N incorporation in glutamate. These findings provide evidence that ammonia is detoxified by the concerted action of GDH and ALAT both in vivo and in vitro, a mechanism that is accelerated in the presence of MSO thereby reducing the glutamine level in brain. Thus, GS could be a potential drug target in the treatment of hyperammonemia in patients with hepatic encephalopathy.

  7. An ultra­high field Magnetic Resonance Spectroscopy study of post exercise brain lactate, glutamate and glutamine change in the human brain.

    Andrea eDennis

    2015-12-01

    Full Text Available During strenuous exercise there is a progressive increase in lactate uptake and metabolism into the brain as workload and plasma lactate levels increase. Although it is now widely accepted that the brain can metabolise lactate, few studies have directly measured brain lactate following vigorous exercise. Here, we used ultra-high field Magnetic Resonance Spectroscopy of the brain to obtain static measures of brain lactate, as well as brain glutamate and glutamine after vigorous exercise. The aims of our experiment were to (a track the changes in brain lactate following recovery from exercise and, (b to simultaneously measure the signals from brain glutamate and glutamine. The results of our experiment showed that vigorous exercise resulted in a significant increase in brain lactate. Furthermore, both glutamate and glutamine were successfully resolved, and as expected, although contrary to some previous reports, we did not observe any significant change in either amino acid after exercise. We did however observe a negative correlation between glutamate and a measure of fitness. These results support the hypothesis that peripherally-derived lactate is taken up by the brain when available. Our data additionally highlight the potential of ultra-high field magnetic resonance spectroscopy as a non-invasive way of measuring multiple brain metabolite changes with exercise.

  8. Natural HLA-B*2705 Protein Ligands with Glutamine as Anchor Motif

    Infantes, Susana; Lorente, Elena; Barnea, Eilon; Beer, Ilan; Barriga, Alejandro; Lasala, Fátima; Jiménez, Mercedes; Admon, Arie; López, Daniel

    2013-01-01

    The presentation of short viral peptide antigens by human leukocyte antigen (HLA) class I molecules on cell surfaces is a key step in the activation of cytotoxic T lymphocytes, which mediate the killing of pathogen-infected cells or initiate autoimmune tissue damage. HLA-B27 is a well known class I molecule that is used to study both facets of the cellular immune response. Using mass spectrometry analysis of complex HLA-bound peptide pools isolated from large amounts of HLA-B*2705+ cells, we identified 200 naturally processed HLA-B*2705 ligands. Our analyses revealed that a change in the position (P) 2 anchor motif was detected in the 3% of HLA-B*2705 ligands identified. B*2705 class I molecules were able to bind these six GlnP2 peptides, which showed significant homology to pathogenic bacterial sequences, with a broad range of affinities. One of these ligands was able to bind with distinct conformations to HLA-B27 subtypes differentially associated with ankylosing spondylitis. These conformational differences could be sufficient to initiate autoimmune damage in patients with ankylosing spondylitis-associated subtypes. Therefore, these kinds of peptides (short, with GlnP2, and similar low affinity to all HLA-B27 subtypes tested but with unlike conformations in differentially ankylosing spondylitis-associated subtypes) must not be excluded from future researches involving potential arthritogenic peptides. PMID:23430249

  9. Glutamate/glutamine concentrations in the dorsal anterior cingulate vary with Post-Traumatic Stress Disorder symptoms.

    Harnett, Nathaniel G; Wood, Kimberly H; Ference, Edward W; Reid, Meredith A; Lahti, Adrienne C; Knight, Amy J; Knight, David C

    2017-08-01

    Trauma and stress-related disorders (e.g., Acute Stress Disorder; ASD and Post-Traumatic Stress Disorder; PTSD) that develop following a traumatic event are characterized by cognitive-affective dysfunction. The cognitive and affective functions disrupted by stress disorder are mediated, in part, by glutamatergic neural systems. However, it remains unclear whether neural glutamate concentrations, measured acutely following trauma, vary with ASD symptoms and/or future PTSD symptom expression. Therefore, the current study utilized proton magnetic resonance spectroscopy ( 1 H-MRS) to investigate glutamate/glutamine (Glx) concentrations within the dorsal anterior cingulate cortex (ACC) of recently (i.e., within one month) traumatized individuals and non-traumatized controls. Although Glx concentrations within dorsal ACC did not differ between recently traumatized and non-traumatized control groups, a positive linear relationship was observed between Glx concentrations and current stress disorder symptoms in traumatized individuals. Further, Glx concentrations showed a positive linear relationship with future stress disorder symptoms (i.e., assessed 3 months post-trauma). The present results suggest glutamate concentrations may play a role in both acute and future post-traumatic stress symptoms following a traumatic experience. The current results expand our understanding of the neurobiology of stress disorder and suggest glutamate within the dorsal ACC plays an important role in cognitive-affective dysfunction following a traumatic experience. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. Differences in the roles of a glutamine amidotransferase subunit of pyridoxal 5'-phosphate synthase between Bacillus circulans and Bacillus subtilis.

    Itagaki, Shiori; Haga, Minami; Oikawa, Yuji; Sakoda, Ayaka; Ohke, Yoshie; Sawada, Hiroshi; Eguchi, Tadashi; Tamegai, Hideyuki

    2013-01-01

    BtrC2 of the butirosin producer Bacillus circulans is a non-catalytic subunit of 2-deoxy-scyllo-inosose (DOI) synthase that is involved in butirosin biosynthesis, and also a homolog of glutamine amidotransferase subunit (PdxT) of pyridoxal 5'-phosphate (PLP) synthase of Bacillus subtilis. BtrC2 has been found to have functions in B. circulans both in primary and secondary metabolism. In this study, we investigated the properties of PdxT of B. subtilis in order to determine whether the property of enzyme stabilization is universal among PdxT homologs. Complementation with PdxT in the btrC2 disruptant of B. circulans restored the growth and short-term production of antibiotics, but long-term production of antibiotics cannot be restored. Additionally, PdxT did not bind physically with or stabilize BtrC. Our results indicate that the function of BtrC2 in secondary metabolism is specific properties, not universal among PdxT homologs.

  11. Glutamine and glutamic acid supplementation enhances performance of broiler chickens under the hot and humid tropical condition

    Joshua O. Olubodun

    2015-02-01

    Full Text Available Day-old (day 1 commercial broiler chickens were fed i basal diet (control, ii basal diet +0.5% AminoGut (AG, or iii basal diet +1% AG from 1 to 42 d of age under the hot and humid tropical environment. AminoGut is a commercial dietary supplement containing a mixture of L-glutamine (Gln and L-glutamic (Glu acid. Weight gain and feed conversion ratio during the starter (1 to 21 d and overall (1 to 42 d periods improved linearly and quadratically with AG supplementation when compared to control. Supplementing birds with AG significantly reduced overall mortality rate. At 21 and 42 d of age, intestinal (duodenum and ileum villi height and crypt depth showed both linear and quadratic positive responses to AG supplementation. Intestinal amylase activity increased linearly and quadratically on d 21, and linearly only on d 42. In conclusion, Gln and Glu supplementation was beneficial in improving the growth performance and survivability of broiler chickens under the hot and humid tropical environment.

  12. Structural asymmetry of phosphodiesterase-9, potential protonation of a glutamic acid, and role of the invariant glutamine.

    Jing Hou

    2011-03-01

    Full Text Available PDE9 inhibitors show potential for treatment of diseases such as diabetes. To help with discovery of PDE9 inhibitors, we performed mutagenesis, kinetic, crystallographic, and molecular dynamics analyses on the active site residues of Gln453 and its stabilizing partner Glu406. The crystal structures of the PDE9 Q453E mutant (PDE9Q453E in complex with inhibitors IBMX and (S-BAY73-6691 showed asymmetric binding of the inhibitors in two subunits of the PDE9Q453E dimer and also the significant positional change of the M-loop at the active site. The kinetic analysis of the Q453E and E406A mutants suggested that the invariant glutamine is critical for binding of substrates and inhibitors, but is unlikely to play a key role in the differentiation between substrates of cGMP and cAMP. The molecular dynamics simulations suggest that residue Glu406 may be protonated and may thus explain the hydrogen bond distance between two side chain oxygens of Glu453 and Glu406 in the crystal structure of the PDE9Q453E mutant. The information from these studies may be useful for design of PDE9 inhibitors.

  13. Origins of domestication and polyploidy in oca (Oxalis Tuberosa: Oxalidaceae). 2. Chloroplast-expressed glutamine synthetase data.

    Emshwiller, Eve; Doyle, Jeff J

    2002-07-01

    In continuing study of the origins of the octoploid tuber crop oca, Oxalis tuberosa Molina, we used phylogenetic analysis of DNA sequences of the chloroplast-active (nuclear encoded) isozyme of glutamine synthetase (ncpGS) from cultivated oca, its allies in the "Oxalis tuberosa alliance," and other Andean Oxalis. Multiple ncpGS sequences found within individuals of both the cultigen and a yet unnamed wild tuber-bearing taxon of Bolivia were separated by molecular cloning, but some cloned sequences appeared to be artifacts of polymerase chain reaction (PCR) recombination and/or Taq error. Nonetheless, three classes of nonrecombinant sequences each joined a different part of the O. tuberosa alliance clade on the ncpGS gene tree. Octoploid oca shares two sequence classes with the Bolivian tuber-bearing taxon (of unknown ploidy level). Fixed heterozygosity of these two sequence classes in all ocas sampled suggests that they represent homeologous loci and that oca is allopolyploid. A third sequence class, found in eight of nine oca plants sampled, might represent a third homeologous locus, suggesting that oca may be autoallopolyploid, and is shared with another wild tuber-bearing species, tetraploid O. picchensis of southern Peru. Thus, ncpGS data identify these two taxa as the best candidates as progenitors of cultivated oca.

  14. Metabolic flux profiling of MDCK cells during growth and canine adenovirus vector production.

    Carinhas, Nuno; Pais, Daniel A M; Koshkin, Alexey; Fernandes, Paulo; Coroadinha, Ana S; Carrondo, Manuel J T; Alves, Paula M; Teixeira, Ana P

    2016-03-23

    Canine adenovirus vector type 2 (CAV2) represents an alternative to human adenovirus vectors for certain gene therapy applications, particularly neurodegenerative diseases. However, more efficient production processes, assisted by a greater understanding of the effect of infection on producer cells, are required. Combining [1,2-(13)C]glucose and [U-(13)C]glutamine, we apply for the first time (13)C-Metabolic flux analysis ((13)C-MFA) to study E1-transformed Madin-Darby Canine Kidney (MDCK) cells metabolism during growth and CAV2 production. MDCK cells displayed a marked glycolytic and ammoniagenic metabolism, and (13)C data revealed a large fraction of glutamine-derived labelling in TCA cycle intermediates, emphasizing the role of glutamine anaplerosis. (13)C-MFA demonstrated the importance of pyruvate cycling in balancing glycolytic and TCA cycle activities, as well as occurrence of reductive alphaketoglutarate (AKG) carboxylation. By turn, CAV2 infection significantly upregulated fluxes through most central metabolism, including glycolysis, pentose-phosphate pathway, glutamine anaplerosis and, more prominently, reductive AKG carboxylation and cytosolic acetyl-coenzyme A formation, suggestive of increased lipogenesis. Based on these results, we suggest culture supplementation strategies to stimulate nucleic acid and lipid biosynthesis for improved canine adenoviral vector production.

  15. Dynamics of glutamine synthetase activity in the cycle of chlorella development

    Selivanova, K.M.; Evstigneeva, Z.G.; Kretovich, V.L.

    1980-01-01

    The zinc role in lysosomes and lysosomal membranes of an irradiated tumour tissue is elucidated in the experiments on rats. The animals with inoculated sarcoma have been exposed to local irradiation (the 1000 R dose per a tumour) on the 14th day of the tumour development. Quantitative zinc determination is carried out by the neutron-activation analysis method on thermal neutrons. It is established that zinc being in the lysosomal membrane structure of the tumour cells plays the definite biological role in the tumour lysosome functions

  16. Double-blinded, placebo-controlled trial on intravenous L-alanyl-L-glutamine in the incidence of oral mucositis following chemoradiotherapy in patients with head-and-neck cancer

    Cerchietti, Leandro C.A.; Navigante, Alfredo H.; Lutteral, Maribel A.; Castro, Monica A.; Kirchuk, Ricardo; Bonomi, Marcelo; Cabalar, Maria Esther; Roth, Berta; Negretti, Graciela; Sheinker, Beatriz; Uchima, Patricia

    2006-01-01

    Purpose: We performed this double-blinded, placebo-controlled study to determine the safety and efficacy of L-alanyl-L-glutamine in the prevention of mucositis in patients with head-and-neck cancer. Methods and Materials: Thirty-two patients with head-and-neck cancer were treated with chemoradiotherapy (CRT) (radiotherapy daily up to 70 Gy plus cisplatin/5-fluoruracil once a week) and were asked to participate. Twenty-nine patients received the CRT schedule and were double-blindly assigned to receive either intravenous L-alanyl-L-glutamine 0.4 g/kg weight/day or an equal volume of saline (placebo) during chemotherapy days. Results: Fourteen patients received L-alanyl-L-glutamine and 15 received placebo. Mucositis was assessed by the Objective Mucositis Score (OMS) and the World Health Organization (WHO) grading system. There was a significant difference in incidence of mucositis developed in patients receiving placebo compared with those who received L-alanyl-L-glutamine (p = 0.035). The number of patients with severe objective mucositis (OMS >1.49) was higher in the placebo group compared with the L-alanyl-L-glutamine group (67% vs. 14%, p 0.007). L-alanyl-L-glutamine patients experienced less pain (three highest Numeric Rating Scale scores of 1.3/10 vs. 6.3/10 respectively, p = 0.008) and need for feeding tubes (14% vs. 60% respectively, p = 0.020) compared with placebo patients. No adverse effects related to the drug or the infusions were noted in either group. Conclusion: For patients with head-and-neck cancer receiving CRT, intravenous L-alanyl-L-glutamine may be an effective preventive measure to decrease the severity of mucositis

  17. Characterization of energy and neurotransmitter metabolism in cortical glutamatergic neurons derived from human induced pluripotent stem cells

    Aldana, Blanca I; Zhang, Yu; Lihme, Maria Fog

    2017-01-01

    pathways in neurons derived from human induced pluripotent stem cells (hiPSC). With this aim, cultures of hiPSC-derived neurons were incubated with [U-(13)C]glucose, [U-(13)C]glutamate or [U-(13)C]glutamine. Isotopic labeling in metabolites was determined using gas chromatography coupled to mass...

  18. Light-stimulated cell expansion in bean (Phaseolus vulgaris L.) leaves. I. Growth can occur without photosynthesis

    Van Volkenburgh, E.; Cleland, R. E.

    1990-01-01

    Cell expansion in dicotyledonous leaves is strongly stimulated by bright white light (WL), at least in part as a result of light-induced acidification of the cell walls. It has been proposed that photosynthetic reactions are required for light-stimulated transport processes across plasma membranes of leaf cells, including proton excretion. The involvement of photosynthesis in growth and wall acidification of primary leaves of bean has been tested by inhibiting photosynthesis in two ways: by reducing chlorophyll content of intact plants with tentoxin (TX) and by treating leaf discs with 3-(3,4-dichlorophenyl)-1,1-dimethylurea (DCMU). Exposure to bright WL stimulated growth of intact leaves of TX-treated plants. Discs excised from green as well as from TX-or DCMU-treated leaves also responded by growing faster in WL, as long as exogenous sucrose was supplied to the photosynthetically inhibited tissues. The WL caused acidification of the epidermal surface of intact TX-leaves, but acidification of the incubation medium by mesophyll cells only occurred when photosynthesis was not inhibited. It is concluded that light-stimulated cell enlargement of bean leaves, and the necessary acidification of epidermal cell walls, are mediated by a pigment other than chlorophyll. Light-induced proton excretion by mesophyll cells, on the other hand, may require both a photosynthetic product (or exogenous sugars) and a non-photosynthetic light effect.

  19. Temporal fluxomics reveals oscillations in TCA cycle flux throughout the mammalian cell cycle.

    Ahn, Eunyong; Kumar, Praveen; Mukha, Dzmitry; Tzur, Amit; Shlomi, Tomer

    2017-11-06

    Cellular metabolic demands change throughout the cell cycle. Nevertheless, a characterization of how metabolic fluxes adapt to the changing de