Ralf Dringen

Publications of Ralf Dringen

• Upregulation of Metallothioneins After Exposure of Cultured Primary Astrocytes to Silver Nanoparticles.

  Authors: Eva M Luther, Maike M Schmidt, Joerg Diendorf, Matthias Epple, Ralf Dringen

  Neurochemical research. 04/2012;

  To test for the prolonged consequences of a short transient exposure of astrocytes to silver nanoparticles (AgNP), cultured primary astrocytes were incubated for 4 h in the presence of AgNP and theTo test for the prolonged consequences of a short transient exposure of astrocytes to silver nanoparticles (AgNP), cultured primary astrocytes were incubated for 4 h in the presence of AgNP and the cell viability as well as various metabolic parameters were investigated during a subsequent incubation in AgNP-free medium. Acute exposure of astrocytes to AgNP led to a concentration-dependent increase in the specific cellular silver content to up to 46 nmol/mg protein, but did not compromise cell viability. During a subsequent incubation of the cells in AgNP-free medium, the cellular silver content of AgNP-treated astrocytes remained almost constant for up to 7 days. The cellular presence of AgNP did neither induce any delayed cell toxicity nor were alterations in cellular glucose consumption, lactate production or in the cellular ratio of glutathione to glutathione disulfide observed. However, Western blot analysis and immunocytochemical staining revealed that AgNP-treated astrocytes strongly upregulated the expression of metallothioneins. These results demonstrate that a prolonged presence of accumulated AgNP does not compromise the viability and the basal metabolism of cultured astrocytes and suggest that the upregulation of metallothioneins may help to prevent silver-mediated toxicity that could be induced by AgNP-derived silver ions.

• Formate generated by cellular oxidation of formaldehyde accelerates the glycolytic flux in cultured astrocytes.

  Authors: Ketki Tulpule, Ralf Dringen

  Glia. 04/2012; 60(4):582-93.

  Formaldehyde is a neurotoxic compound that can be endogenously generated in the brain. Because astrocytes play a key role in metabolism and detoxification processes in brain, we have investigated theFormaldehyde is a neurotoxic compound that can be endogenously generated in the brain. Because astrocytes play a key role in metabolism and detoxification processes in brain, we have investigated the capacity of these cells to metabolize formaldehyde using primary astrocyte-rich cultures as a model system. Application of formaldehyde to these cultures resulted in the appearance of formate in cells and in a time-, concentration- and temperature-dependent disappearance of formaldehyde from the medium that was accompanied by a matching extracellular accumulation of formate. This formaldehyde-oxidizing capacity of astrocyte cultures is likely to be catalyzed by alcohol dehydrogenase 3 and aldehyde dehydrogenase 2, because the cells of the cultures contain the mRNAs of these formaldehyde-oxidizing enzymes. In addition, exposure to formaldehyde increased both glucose consumption and lactate production by the cells. Both the strong increase in the cellular formate content and the increase in glycolytic flux were only observed after application of formaldehyde to the cells, but not after treatment with exogenous methanol or formate. The accelerated lactate production was not additive to that obtained for azide, a known inhibitor of complex IV of the respiratory chain, and persisted after removal of formaldehyde after a formaldehyde exposure for 1.5 h. These data demonstrate that cultured astrocytes efficiently oxidize formaldehyde to formate, which subsequently enhances glycolytic flux, most likely by inhibition of mitochondrial respiration. © 2012 Wiley Periodicals, Inc.

• Copper export from cultured astrocytes.

  Authors: Ivo F Scheiber, Maike M Schmidt, Ralf Dringen

  Neurochemistry international. 02/2012; 60(3):292-300.

  Copper is an essential trace metal that is required as a catalytic co-factor or a structural component of several important enzymes. However, since excess of copper can also harm cells due to itsCopper is an essential trace metal that is required as a catalytic co-factor or a structural component of several important enzymes. However, since excess of copper can also harm cells due to its potential to catalyse the generation of toxic reactive oxygen species, transport of copper and the cellular copper content are tightly regulated. Astrocytes are known to efficiently take up copper ions, but it was not known whether these cells are also able to export copper. Treatment of astrocyte-rich primary cultures for 24h with copper chloride caused a concentration-dependent increase in the specific cellular copper content. During further 24h incubation in the absence of copper chloride, the copper-loaded astrocytes remained viable and released up to 45% of the accumulated copper. The rate of copper export was proportional to the amount of cellular copper, was almost completely prevented by lowering the incubation temperature to 4°C and was partly prevented by the endocytosis inhibitor amiloride. Copper export is most likely mediated by the copper ATPase ATP7A, since this transporter is expressed in astrocyte cultures and its cellular location is strongly affected by the absence or the presence of extracellular copper. The potential of cultured astrocytes to export copper suggests that astrocytes provide neighbouring cells in brain with this essential trace element.

• The antiretroviral protease inhibitors indinavir and nelfinavir stimulate Mrp1-mediated GSH export from cultured brain astrocytes.

  Authors: Maria Brandmann, Ketki Tulpule, Maike M Schmidt, Ralf Dringen

  Journal of neurochemistry. 01/2012; 120(1):78-92.

  Combinations of antiretroviral drugs are successfully used for the treatment of acquired immune deficiency syndrome and reduce the incidence of severe human immunodeficiency virus (HIV)-associatedCombinations of antiretroviral drugs are successfully used for the treatment of acquired immune deficiency syndrome and reduce the incidence of severe human immunodeficiency virus (HIV)-associated dementia. To test whether such drugs affect the GSH metabolism of brain cells, we have exposed astrocyte-rich primary cultures to various antiretroviral compounds. Treatment of the cultures with the protease inhibitors indinavir or nelfinavir in low micromolar concentrations resulted in a time- and concentration-dependent depletion of cellular GSH from viable cells which was accompanied by a matching increase in the extracellular GSH content. In contrast, the reverse transcriptase inhibitors zidovudine, lamivudine, efavirenz or nevirapine did not alter cellular or extracellular GSH levels. Removal of indinavir from the medium by washing the cells terminated the stimulated GSH export immediately, while the nelfinavir-induced accelerated GSH export was maintained even after removal of nelfinavir. The stimulation of the GSH export from viable astrocytes by indinavir or nelfinavir was completely prevented by the application of MK571, an inhibitor of the multidrug resistance protein 1. These data demonstrate that indinavir and nelfinavir stimulate multidrug resistance protein 1-mediated GSH export from viable astrocytes and suggest that treatment of patients with such inhibitors may affect the GSH homeostasis in brain.

• Magnetic field-induced acceleration of the accumulation of magnetic iron oxide nanoparticles by cultured brain astrocytes.

  Authors: Marie-Christin Lamkowsky, Mark Geppert, Maike M Schmidt, Ralf Dringen

  Journal of biomedical materials research. Part A. 11/2011;

  Magnetic iron oxide nanoparticles (Fe-NPs) are considered for various biomedical and neurobiological applications that involve the presence of external magnetic fields. However, little is known onMagnetic iron oxide nanoparticles (Fe-NPs) are considered for various biomedical and neurobiological applications that involve the presence of external magnetic fields. However, little is known on the effects of a magnetic field on the uptake of such particles by brain cells. Cultured brain astrocytes accumulated dimercaptosuccinate-coated Fe-NP in a time-, temperature-, and concentration-dependent manner. This accumulation was strongly enhanced by the presence of the magnetic field generated by a permanent neodymium iron boron magnet that had been positioned below the cells. The magnetic field-induced acceleration of the accumulation of Fe-NP increased almost proportional to the strength of the magnetic field applied, increasing the cellular-specific iron content from an initial 10 nmol/mg protein within 4 h of incubation at 37°C to up to 12,000 nmol/mg protein. However, presence of a magnetic field also increased the amounts of iron that attached to the cells during incubation with Fe-NP at 4°C. These results suggest that the presence of an external magnetic field promotes in cultured astrocytes both the binding of Fe-NP to the cell membrane and the internalization of Fe-NP. © 2011 Wiley Periodicals, Inc. J Biomed Mater Res Part A:, 2011.

• Accumulation of silver nanoparticles by cultured primary brain astrocytes.

  Authors: Eva M Luther, Yvonne Koehler, Joerg Diendorf, Matthias Epple, Ralf Dringen

  Nanotechnology. 09/2011; 22(37):375101.

  Silver nanoparticles (AgNP) are components of various food industry products and are frequently used for medical equipment and materials. Although such particles enter the vertebrate brain, little isSilver nanoparticles (AgNP) are components of various food industry products and are frequently used for medical equipment and materials. Although such particles enter the vertebrate brain, little is known on their biocompatibility for brain cells. To study the consequences of an AgNP exposure of brain cells we have treated astrocyte-rich primary cultures with polyvinylpyrrolidone (PVP)-coated AgNP. The incubation of cultured astrocytes with micromolar concentrations of AgNP for up to 24 h resulted in a time- and concentration-dependent accumulation of silver, but did not compromise the cell viability nor lower the cellular glutathione content. In contrast, the incubation of astrocytes for 4 h with identical amounts of silver as AgNO(3) already severely compromised the cell viability and completely deprived the cells of glutathione. The accumulation of AgNP by astrocytes was proportional to the concentration of AgNP applied and significantly lowered by about 30% in the presence of the endocytosis inhibitors chloroquine or amiloride. Incubation at 4 °C reduced the accumulation of AgNP by 80% compared to the values obtained for cells that had been exposed to AgNP at 37 °C. These data demonstrate that viable cultured brain astrocytes efficiently accumulate PVP-coated AgNP in a temperature-dependent process that most likely involves endocytotic pathways.

• Treatment with iron oxide nanoparticles induces ferritin synthesis but not oxidative stress in oligodendroglial cells.

  Authors: Michaela C Hohnholt, Mark Geppert, Ralf Dringen

  Acta biomaterialia. 07/2011; 7(11):3946-54.

  Magnetic iron oxide nanoparticles (IONPs) have been used for a variety of neurobiological applications, although little is yet known as to the fate of such particles in brain cells. To address theseMagnetic iron oxide nanoparticles (IONPs) have been used for a variety of neurobiological applications, although little is yet known as to the fate of such particles in brain cells. To address these questions, we have exposed oligodendroglial OLN-93 cells to dimercaptosuccinate-coated IONPs. Treatment of the cells strongly increased the specific cellular iron content proportional to the IONP concentrations applied (0-1000 μM total iron as IONPs) up to 300-fold, but did not cause any acute cytotoxicity or induce oxidative stress. To investigate the potential of OLN-93 cells to liberate iron from the accumulated IONPs, we have studied the upregulation of the iron storage protein ferritin and the cell proliferation as cellular processes that depend on the availability of low-molecular-weight iron. The presence of IONPs caused a concentration-dependent increase in the amount of cellular ferritin and partially bypassed the inhibition of cell proliferation by the iron chelator deferoxamine. These data demonstrate that viable OLN-93 cells efficiently take up IONPs and suggest that these cells are able to use iron liberated from accumulated IONPs for their metabolism.

• Copper-treatment increases the cellular GSH content and accelerates GSH export from cultured rat astrocytes.

  Authors: Ivo F Scheiber, Ralf Dringen

  Neuroscience letters. 07/2011; 498(1):42-6.

  To test whether copper exposure affects astroglial glutathione (GSH) metabolism, we have exposed astrocyte-rich primary cultures with copper chloride in concentrations of up to 30 μM and investigatedTo test whether copper exposure affects astroglial glutathione (GSH) metabolism, we have exposed astrocyte-rich primary cultures with copper chloride in concentrations of up to 30 μM and investigated cellular and extracellular GSH contents. Cultured astrocytes accumulated copper in a concentration-dependent manner thereby increasing the specific cellular copper content within 24h up to sevenfold. The increase in the cellular copper content was accompanied by a proportional increase in the specific cellular GSH content that reached up to 165% of the values of cells that had been incubated without copper, while the low cellular content of GSH disulfide (GSSG) remained unaltered in copper-treated cells. Also the rate of GSH export was significantly increased after copper exposure reaching up to 177% of control values. The export of GSH from control and copper-treated astrocytes was lowered by more than 70%, if cells were incubated in presence of the multidrug-resistance protein (Mrp) 1 inhibitor MK571 or at a low incubation temperature of 4°C. These data demonstrate that copper accumulation stimulates GSH synthesis and accelerates Mrp1-mediated GSH export from cultured astrocytes. These processes are likely to contribute to the resistance of astrocytes against copper toxicity and could improve the supply of GSH precursors from astrocytes to neurons.

• Adsorption and reduction of glutathione disulfide on α-Al2O3 nanoparticles: experiments and modeling.

  Authors: Ralf Dringen, Yvonne Koehler, Ludmilla Derr, Giulia Tomba, Maike M Schmidt, Laura Treccani, Lucio Colombi Ciacchi, Kurosch Rezwan

  Langmuir : the ACS journal of surfaces and colloids. 06/2011; 27(15):9449-57.

  Glutathione disulfide (GSSG; γ-GluCysGly disulfide) was used as a physiologically relevant model molecule to investigate the fundamental adsorption mechanisms of polypeptides onto α-aluminaGlutathione disulfide (GSSG; γ-GluCysGly disulfide) was used as a physiologically relevant model molecule to investigate the fundamental adsorption mechanisms of polypeptides onto α-alumina nanoparticles. Its adsorption/desorption behavior was studied by enzymatic quantification of the bound GSSG combined with zeta potential measurements of the particles. The adsorption of GSSG to alumina nanoparticles was rapid, was prevented by alkaline pH, was reversed by increasing ionic strength, and followed a nearly ideal Langmuir isotherm with a standard Gibbs adsorption energy of -34.7 kJ/mol. Molecular dynamics simulations suggest that only one of the two glutathionyl moieties contained in GSSG binds stably to the nanoparticle surface. This was confirmed experimentally by the release of GSH from the bound GSSG upon reducing its disulfide bond with dithiothreitol. Our data indicate that electrostatic interactions via the carboxylate groups of one of the two glutathionyl moieties of GSSG are predominantly responsible for the binding of GSSG to the alumina surface. The results and conclusions presented here can provide a base for further experimental and modeling studies on the interactions of biomolecules with ceramic materials.

• The metabolism and toxicity of hemin in astrocytes.

  Authors: Theresa N Dang, Glenda M Bishop, Ralf Dringen, Stephen R Robinson

  Glia. 06/2011; 59(10):1540-50.

  Hemin is cytotoxic, and contributes to the brain damage that accompanies hemorrhagic stroke. In order to better understand the basis of hemin toxicity in astrocytes, the present study quantifiedHemin is cytotoxic, and contributes to the brain damage that accompanies hemorrhagic stroke. In order to better understand the basis of hemin toxicity in astrocytes, the present study quantified hemin metabolism and compared it to the pattern of cell death. Heme oxygenase-1 (HO-1) expression was first evident after 2 h incubation with hemin, with maximal expression being observed by 24 h. Despite the induction of HO-1, it was found that the proportion of hemin metabolized by astrocytes remained fairly constant throughout the 24 h period, with 70-80% of intracellular hemin remaining intact. A period of cell loss began after 2 h exposure to hemin, which gradually increased in severity to reach a maximum by 24 h. This cell loss could not be attenuated by the iron chelator, 1,10-phenanthroline, or by several antioxidant compounds (Trolox, N-acetyl-L-cysteine and N-tert-butyl-α-phenylnitrone), indicating that the mechanism of hemin toxicity does not involve iron. While these results make it unlikely that hemin toxicity is due to interactions with endogenous H(2)O(2), hemin toxicity was increased in the presence of supraphysiological levels of H(2)O(2) and this increase was ameliorated by PHEN, indicating that the iron released from hemin can be toxic under some pathological conditions. However, when H(2)O(2) is present at physiological levels, the toxicity of hemin appears to be caused by other mechanisms that may involve bilirubin and carbon monoxide in this model system.

• Effects of chlorinated acetates on the glutathione metabolism and on glycolysis of cultured astrocytes.

  Authors: Maike M Schmidt, Astrid Rohwedder, Ralf Dringen

  Neurotoxicity research. 05/2011; 19(4):628-37.

  The chlorinated acetates monochloroacetate (MCA), dichloroacetate (DCA), and trichloroacetate (TCA) are generated in water disinfection processes and are formed during metabolic detoxification ofThe chlorinated acetates monochloroacetate (MCA), dichloroacetate (DCA), and trichloroacetate (TCA) are generated in water disinfection processes and are formed during metabolic detoxification of industrial solvents such as trichloroethylene. In order to test for consequences of an exposure of brain cells to the different chlorinated acetates, glutathione levels and lactate production of primary astrocyte cultures were investigated as indicators for the potential of chlorinated acetates to disturb cellular detoxification processes and glucose metabolism, respectively. Application of MCA to cultured astrocytes caused a time and concentration dependent deprivation of cellular glutathione, inactivation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH) activity, and loss in cell viability with halfmaximal effects observed for MCA concentrations between 0.3 and 3 mM. In contrast, the presence of acetate, DCA, or TCA in a concentration of 10 mM did not compromise cell viability nor affect cellular glutathione content or GAPDH activity. However, the presence of DCA and TCA significantly lowered the rate of cellular lactate production in viable astrocytes. These data demonstrate that the extent of chlorination strongly determines the potential of chlorinated acetates to disturb glutathione and/or glucose metabolism of astrocytes.

• Accumulation of non-transferrin-bound iron by neurons, astrocytes, and microglia.

  Authors: Glenda M Bishop, Theresa N Dang, Ralf Dringen, Stephen R Robinson

  Neurotoxicity research. 04/2011; 19(3):443-51.

  Neurodegenerative conditions such as Alzheimer's disease, Parkinson's disease, and hemorrhagic stroke are associated with increased levels of non-transferrin-bound iron (NTBI) in the brain, which canNeurodegenerative conditions such as Alzheimer's disease, Parkinson's disease, and hemorrhagic stroke are associated with increased levels of non-transferrin-bound iron (NTBI) in the brain, which can promote Fenton chemistry. While all types of brain cells can take up NTBI, their efficiency of accumulation and capacity to withstand iron-mediated toxicity has not been directly compared. The present study assessed NTBI accumulation in cultures enriched in neurons, astrocytes, or microglia after exposure to ferric ammonium citrate (FAC). Microglia were found to be the most efficient in accumulating iron, followed by astrocytes, and then neurons. Exposure to 100 μM FAC for 24 h increased the specific iron content of cultured neurons, astrocytes, and microglial cells by 30-, 80-, and 100-fold, respectively. All cell types accumulated iron against the concentration gradient, resulting in intracellular iron concentrations that were several orders of magnitude higher than the extracellular iron concentrations. Accumulation of these large amounts of iron did not affect the viability of the cell cultures, indicating a high resistance to iron-mediated toxicity. These findings show that neurons, astrocytes and microglia cultured from neonatal mice all have the capacity to accumulate and safely store large quantities of iron, but that glial cells do this more efficiently than neurons. It is concluded that neurodegenerative conditions involving iron-mediated toxicity may be due to a failure of iron transport or storage mechanisms, rather than to the presence of high levels of NTBI.

• Uptake, metabolism and toxicity of hemin in cultured neurons.

  Authors: Theresa N Dang, Stephen R Robinson, Ralf Dringen, Glenda M Bishop

  Neurochemistry international. 03/2011;

  Following hemorrhagic stroke, red blood cells lyse and release neurotoxic hemin into the interstitial space. The present study investigates whether neurons can accumulate and metabolize hemin. WeFollowing hemorrhagic stroke, red blood cells lyse and release neurotoxic hemin into the interstitial space. The present study investigates whether neurons can accumulate and metabolize hemin. We demonstrate that cultured neurons express the heme carrier protein 1 (HCP1), and that this transporter appears to contribute to the time- and concentration-dependent accumulation of hemin by neurons. Although exposure of neurons to hemin stimulates the synthesis of the iron storage protein ferritin, approximately 80% of the hemin accumulated by neurons remains intact. Within 24h of incubation, substantial neurotoxicity was observed that was not attenuated by the cell permeable, selective ferrous iron chelator, 1,10-phenanthroline. These results demonstrate that while neurons efficiently accumulate hemin they slowly degrade it, and they support the conclusion that intact hemin is more neurotoxic than the iron released from the breakdown of hemin. Further investigations are required to determine the basis of this neurotoxicity.

• Copper accelerates glycolytic flux in cultured astrocytes.

  Authors: Ivo F Scheiber, Ralf Dringen

  Neurochemical research. 02/2011; 36(5):894-903.

  Astrocyte-rich primary cultures were used to investigate the consequences of a copper exposure on the glucose metabolism of astrocytes. After application of CuCl(2) (30 μM) the specific cellularAstrocyte-rich primary cultures were used to investigate the consequences of a copper exposure on the glucose metabolism of astrocytes. After application of CuCl(2) (30 μM) the specific cellular copper content increased from initial 1.5 ± 0.2 nmol/mg to a steady state level of 7.9 ± 0.9 nmol/mg within about 12 h. The copper accumulation was accompanied by a significant increase in the extracellular lactate concentration. The stimulating effect of copper on the lactate production remained after removal of extracellular copper. Copper treatment accelerated the rates of both glucose consumption and lactate production by about 60%. The copper induced acceleration of glycolytic flux was prevented by inhibition of protein synthesis, and additive to the stimulation of glycolysis observed for inhibitors of respiration or prolyl hydroxylases. A copper induced stimulation of glycolytic flux in astrocytes could have severe consequences for the glucose metabolism of the brain in conditions of copper overload.

• Formaldehyde stimulates Mrp1-mediated glutathione deprivation of cultured astrocytes.

  Authors: Ketki Tulpule, Ralf Dringen

  Journal of neurochemistry. 02/2011; 116(4):626-35.

  Formaldehyde (Fal) is an environmental neurotoxin that is also endogenously produced in brain. Since the tripeptide glutathione (GSH) plays an important role in detoxification processes in brainFormaldehyde (Fal) is an environmental neurotoxin that is also endogenously produced in brain. Since the tripeptide glutathione (GSH) plays an important role in detoxification processes in brain cells, we have investigated the consequences of a Fal exposure on the GSH metabolism of brain cells, using astrocyte-rich primary cultures as model system. Treatment of these cultures with Fal resulted in a rapid time- and concentration-dependent depletion of cellular GSH and a matching increase in the extracellular GSH content. Exposure of astrocytes to 1mm Fal for 3h did not compromise cell viability but almost completely deprived the cells of GSH. Half-maximal deprivation of cellular GSH was observed after application of 0.3mm Fal. This effect was rather specific for Fal, since methanol, formate or acetaldehyde did not affect cellular GSH levels. The Fal-stimulated GSH loss from viable astrocytes was completely prevented by semicarbazide-mediated chemical removal of Fal or by the application of MK571, an inhibitor of the multidrug resistance protein 1. These data demonstrate that Fal deprives astrocytes of cellular GSH by a multidrug resistance protein 1-mediated process.

• Uptake of dimercaptosuccinate-coated magnetic iron oxide nanoparticles by cultured brain astrocytes.

  Authors: Mark Geppert, Michaela C Hohnholt, Karsten Thiel, Sylvia Nürnberger, Ingo Grunwald, Kurosch Rezwan, Ralf Dringen

  Nanotechnology. 02/2011; 22(14):145101.

  Magnetic iron oxide nanoparticles (Fe-NP) are currently considered for various diagnostic and therapeutic applications in the brain. However, little is known on the accumulation and biocompatibilityMagnetic iron oxide nanoparticles (Fe-NP) are currently considered for various diagnostic and therapeutic applications in the brain. However, little is known on the accumulation and biocompatibility of such particles in brain cells. We have synthesized and characterized dimercaptosuccinic acid (DMSA) coated Fe-NP and have investigated their uptake by cultured brain astrocytes. DMSA-coated Fe-NP that were dispersed in physiological medium had an average hydrodynamic diameter of about 60 nm. Incubation of cultured astrocytes with these Fe-NP caused a time- and concentration-dependent accumulation of cellular iron, but did not lead within 6 h to any cell toxicity. After 4 h of incubation with 100-4000 µM iron supplied as Fe-NP, the cellular iron content reached levels between 200 and 2000 nmol mg⁻¹ protein. The cellular iron content after exposure of astrocytes to Fe-NP at 4 °C was drastically lowered compared to cells that had been incubated at 37 °C. Electron microscopy revealed the presence of Fe-NP-containing vesicles in cells that were incubated with Fe-NP at 37 °C, but not in cells exposed to the nanoparticles at 4 °C. These data demonstrate that cultured astrocytes efficiently take up DMSA-coated Fe-NP in a process that appears to be saturable and strongly depends on the incubation temperature.

• Glutathione synthesis and metabolism

  Authors: Maike M. Schmidt, Ralf Dringen

  01/2011; Springer Science.

• Exploring uncoupling proteins and antioxidant mechanisms under acute cold exposure in brains of fish.

  Authors: Yung-Che Tseng, Ruo-Dong Chen, Magnus Lucassen, Maike M Schmidt, Ralf Dringen, Doris Abele, Pung-Pung Hwang

  PloS one. 01/2011; 6(3):e18180.

  Exposure to fluctuating temperatures accelerates the mitochondrial respiration and increases the formation of mitochondrial reactive oxygen species (ROS) in ectothermic vertebrates including fish. ToExposure to fluctuating temperatures accelerates the mitochondrial respiration and increases the formation of mitochondrial reactive oxygen species (ROS) in ectothermic vertebrates including fish. To date, little is known on potential oxidative damage and on protective antioxidative defense mechanisms in the brain of fish under cold shock. In this study, the concentration of cellular protein carbonyls in brain was significantly increased by 38% within 1 h after cold exposure (from 28 °C to 18 °C) of zebrafish (Danio rerio). In addition, the specific activity of superoxide dismutase (SOD) and the mRNA level of catalase (CAT) were increased after cold exposure by about 60% (6 h) and by 60%-90% (1 and 24 h), respectively, while the specific glutathione content as well as the ratio of glutathione disulfide to glutathione remained constant and at a very low level. In addition, cold exposure increased the protein level of hypoxia-inducible factor (HIF) by about 50% and the mRNA level of the glucose transporter zglut3 in brain by 50%-100%. To test for an involvement of uncoupling proteins (UCPs) in the cold adaptation of zebrafish, five UCP members were annotated and identified (zucp1-5). With the exception of zucp1, the mRNA levels of the other four zucps were significantly increased after cold exposure. In addition, the mRNA levels of four of the fish homologs (zppar) of the peroxisome proliferator-activated receptor (PPAR) were increased after cold exposure. These data suggest that PPARs and UCPs are involved in the alterations observed in zebrafish brain after exposure to 18°C. The observed stimulation of the PPAR-UCP axis may help to prevent oxidative damage and to maintain metabolic balance and cellular homeostasis in the brains of ectothermic zebrafish upon cold exposure.

• Metabolic and physiological responses in tissues of the long-lived bivalve Arctica islandica to oxygen deficiency.

  Authors: Julia Strahl, Ralf Dringen, Maike M Schmidt, Silvia Hardenberg, Doris Abele

  Comparative biochemistry and physiology. Part A, Molecular & integrative physiology. 12/2010; 158(4):513-9.

  In Arctica islandica, a long lifespan is associated with low metabolic activity, and with a pronounced tolerance to low environmental oxygen. In order to study metabolic and physiological responsesIn Arctica islandica, a long lifespan is associated with low metabolic activity, and with a pronounced tolerance to low environmental oxygen. In order to study metabolic and physiological responses to low oxygen conditions vs. no oxygen in mantle, gill, adductor muscle and hemocytes of the ocean quahog, specimens from the German Bight were maintained for 3.5 days under normoxia (21 kPa=controls), hypoxia (2 kPa) or anoxia (0 kPa). Tissue levels of anaerobic metabolites octopine, lactate and succinate as well as specific activities of octopine dehydrogenase (ODH) and lactate dehydrogenase (LDH) were unaffected by hypoxic incubation, suggesting that the metabolism of A. islandica remains fully aerobic down to environmental oxygen levels of 2 kPa. PO(2)-dependent respiration rates of isolated gills indicated the onset of metabolic rate depression (MRD) below 5 kPa in A. islandica, while anaerobiosis was switched on in bivalve tissues only at anoxia. Tissue-specific levels of glutathione (GSH), a scavenger of reactive oxygen species (ROS), indicate no anticipatory antioxidant response takes place under experimental hypoxia and anoxia exposure. Highest specific ODH activity and a mean ODH/LDH ratio of 95 in the adductor muscle contrasted with maximal specific LDH activity and a mean ODH/LDH ratio of 0.3 in hemocytes. These differences in anaerobic enzyme activity patterns indicate that LDH and ODH play specific roles in different tissues of A. islandica which are likely to economize metabolism during anoxia and reoxygenation.

• 2-deoxyribose deprives cultured astrocytes of their glutathione.

  Authors: Maike M Schmidt, Helena Greb, Hendrik Koliwer-Brandl, Soerge Kelm, Ralf Dringen

  Neurochemical research. 11/2010; 35(11):1848-56.

  High concentrations of 2-deoxy-D-ribose (2dRib) have been reported to cause oxidative stress and to disturb the glutathione (GSH) metabolism of various cell types. Exposure of astrocyte-rich primaryHigh concentrations of 2-deoxy-D-ribose (2dRib) have been reported to cause oxidative stress and to disturb the glutathione (GSH) metabolism of various cell types. Exposure of astrocyte-rich primary cultures to millimolar concentrations of 2dRib or its stereoisomer 2-deoxy-L-ribose, but not the incubation with ribose, 2-deoxyglucose, glucose, fructose or saccharose, lowered the cellular GSH content in a time and concentration dependent manner. After exposure for 4 h to 30 mM 2dRib the cells contained 2dRib in a concentration of about 24 mM. Under these conditions 2dRib did not compromise cell viability and the ability of the cells to synthesise GSH, nor were the cellular ratio of glutathione disulfide (GSSG) to GSH and the extracellular concentrations of GSH or GSSG increased. These data demonstrate that 2dRib deprives viable cultured astrocytes of GSH and suggest that a cellular reaction of GSH with 2dRib or its metabolites is involved in the deprivation of astrocytic GSH.