scholarly journals Role of compartmentalized redox-active iron in hydrogen peroxide-induced DNA damage and apoptosis

2005 ◽  
Vol 387 (3) ◽  
pp. 703-710 ◽  
Author(s):  
Margarita TENOPOULOU ◽  
Paschalis-Thomas DOULIAS ◽  
Alexandra BARBOUTI ◽  
Ulf BRUNK ◽  
Dimitrios GALARIS
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Dna Damage ◽  
Nuclear Dna ◽  
Fluid Phase ◽  
Jurkat Cells ◽  
Acetoxymethyl Ester ◽  
Lysosomal Membranes ◽  
Active Iron ◽  
Redox Active

Jurkat cells in culture were exposed to oxidative stress in the form of continuously generated hydrogen peroxide, obtained by the addition of glucose oxidase to the medium. This treatment induced a rapid, dose-dependent increase in the ICIP (intracellular calcein-chelatable iron pool). Early destabilization of lysosomal membranes and subsequent nuclear DNA strand breaks were also observed, as evaluated by the Acridine Orange relocation test and the comet assay respectively. Somewhat later, these effects were followed by a lowered mitochondrial membrane potential, with release of cytochrome c and apoptosis-inducing factor. These events were all prevented if cells were pretreated with the potent iron chelator DFO (desferrioxamine) for a period of time (2–3 h) long enough to allow the drug to reach the lysosomal compartment following fluid-phase endocytosis. The hydrophilic calcein, a cleavage product of calcein acetoxymethyl ester following the action of cytosolic esterases, obviously does not penetrate intact lysosomal membranes, thus explaining why ICIP increased dramatically following lysosomal rupture. The rapid decrease in ICIP after addition of DFO to the medium suggests draining of cytosolic iron to the medium, rather than penetration of DFO through the plasma membrane. Most importantly, these observations directly connect oxidative stress and resultant DNA damage with lysosomal rupture and the release of redox-active iron into the cytosol and, apparently, the nucleus.

Lysosomal Redox-Active Iron Is Important for Oxidative Stress-Induced DNA Damage

2004 ◽  
Vol 1019 (1) ◽  
pp. 285-288 ◽  
Author(s):  
TINO KURZ ◽  
ALAN LEAKE ◽  
THOMAS ZGLINICKI ◽  
ULF T. BRUNK
Keyword(s):  
Oxidative Stress ◽  
Dna Damage ◽  
Active Iron ◽  
Redox Active

scholarly journals Relocalized redox-active lysosomal iron is an important mediator of oxidative-stress-induced DNA damage

2004 ◽  
Vol 378 (3) ◽  
pp. 1039-1045 ◽  
Author(s):  
Tino KURZ ◽  
Alan LEAKE ◽  
Thomas von ZGLINICKI ◽  
Ulf T. BRUNK
Keyword(s):  
Dna Damage ◽  
Free Radical ◽  
Oxidative Damage ◽  
Nuclear Dna ◽  
Membrane Integrity ◽  
Lysosomal Hydrolases ◽  
Free Radical Biol ◽  
Active Iron ◽  
Vacuolar Compartment ◽  
Redox Active

Oxidative damage to nuclear DNA is known to involve site-specific Fenton-type chemistry catalysed by redox-active iron or copper in the immediate vicinity of DNA. However, the presence of transition metals in the nucleus has not been shown convincingly. Recently, it was proposed that a major part of the cellular pool of loose iron is confined within the acidic vacuolar compartment [Yu, Persson, Eaton and Brunk (2003) Free Radical Biol. Med. 34, 1243–1252; Persson, Yu, Tirosh, Eaton and Brunk (2003) Free Radical Biol. Med. 34, 1295–1305]. Consequently, rupture of secondary lysosomes, as well as subsequent relocation of labile iron to the nucleus, could be an important intermediary step in the generation of oxidative damage to DNA. To test this concept we employed the potent iron chelator DFO (desferrioxamine) conjugated with starch to form an HMM-DFO (high-molecular-mass DFO complex). The HMM-DFO complex will enter cells only via fluid-phase endocytosis and remain within the acidic vacuolar compartment, thereby chelating redox-active iron exclusively inside the endosomal/lysosomal compartment. Both free DFO and HMM-DFO equally protected lysosomal-membrane integrity against H2O2-induced oxidative disruption. More importantly, both forms of DFO prevented H2O2-induced strand breaks in nuclear DNA, including telomeres. To exclude the possibility that lysosomal hydrolases, rather than iron, caused the observed DNA damage, limited lysosomal rupture was induced using the lysosomotropic detergent O-methyl-serine dodecylamine hydrochloride; subsequently, hardly any DNA damage was found. These observations suggest that rapid oxidative damage to cellular DNA is minimal in the absence of redox-active iron and that oxidant-mediated DNA damage, observed in normal cells, is mainly derived from intralysosomal iron translocated to the nucleus after lysosomal rupture.

scholarly journals Metallothionein protects against oxidative stress-induced lysosomal destabilization

2006 ◽  
Vol 394 (1) ◽  
pp. 275-283 ◽  
Author(s):  
Sarah K. Baird ◽  
Tino Kurz ◽  
Ulf T. Brunk
Keyword(s):  
Oxidative Stress ◽  
Fluid Phase ◽  
Iron Phosphate ◽  
Iron Chelator ◽  
Cellular Resistance ◽  
Lysosomal Compartment ◽  
Active Iron ◽  
Redox Active ◽  
Fluid Phase Endocytosis ◽  
X Irradiation

The introduction of apo-ferritin or the iron chelator DFO (desferrioxamine) conjugated to starch into the lysosomal compartment protects cells against oxidative stress, lysosomal rupture and ensuing apoptosis/necrosis by binding intralysosomal redox-active iron, thus preventing Fenton-type reactions and ensuing peroxidation of lysosomal membranes. Because up-regulation of MTs (metallothioneins) also generates enhanced cellular resistance to oxidative stress, including X-irradiation, and MTs were found to be capable of iron binding in an acidic and reducing lysosomal-like environment, we propose that these proteins might similarly stabilize lysosomes following autophagocytotic delivery to the lysosomal compartment. Here, we report that Zn-mediated MT up-regulation, assayed by Western blotting and immunocytochemistry, results in lysosomal stabilization and decreased apoptosis following oxidative stress, similar to the protection afforded by fluid-phase endocytosis of apo-ferritin or DFO. In contrast, the endocytotic uptake of an iron phosphate complex destabilized lysosomes against oxidative stress, but this was suppressed in cells with up-regulated MT. It is suggested that the resistance against oxidative stress, known to occur in MT-rich cells, may be a consequence of autophagic turnover of MT, resulting in reduced iron-catalysed intralysosomal peroxidative reactions.

Antioxidative Effect of Rhus javanica Linne Extract Against Hydrogen Peroxide or Menadione Induced Oxidative Stress and DNA Damage in HepG2Cells

2004 ◽  
Vol 9 (2) ◽  
pp. 150-155 ◽  
Author(s):  
Chi-Sung Chun ◽  
Ji-Hyun Kim ◽  
Hyun-Ae Lim ◽  
Ho-Yong Sohn ◽  
Kun-Ho Son ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Dna Damage ◽  
Antioxidative Effect

scholarly journals Escherichia coli MutM Suppresses Illegitimate Recombination Induced by Oxidative Stress

Genetics ◽  
1999 ◽  
Vol 151 (2) ◽  
pp. 439-446 ◽  
Author(s):  
Masaaki Onda ◽  
Katsuhiro Hanada ◽  
Hirokazu Kawachi ◽  
Hideo Ikeda
Keyword(s):  
Oxidative Stress ◽  
Escherichia Coli ◽  
Hydrogen Peroxide ◽  
Dna Damage ◽  
Double Strand Breaks ◽  
Illegitimate Recombination ◽  
Recombination Analysis ◽  
Wild Type ◽  
Strand Breaks ◽  
In The Wild

Abstract DNA damage by oxidative stress is one of the causes of mutagenesis. However, whether or not DNA damage induces illegitimate recombination has not been determined. To study the effect of oxidative stress on illegitimate recombination, we examined the frequency of λbio transducing phage in the presence of hydrogen peroxide and found that this reagent enhances illegitimate recombination. To clarify the types of illegitimate recombination, we examined the effect of mutations in mutM and related genes on the process. The frequency of λbio transducing phage was 5- to 12-fold higher in the mutM mutant than in the wild type, while the frequency in the mutY and mutT mutants was comparable to that of the wild type. Because 7,8-dihydro-8-oxoguanine (8-oxoG) and formamido pyrimidine (Fapy) lesions can be removed from DNA by MutM protein, these lesions are thought to induce illegitimate recombination. Analysis of recombination junctions showed that the recombination at Hotspot I accounts for 22 or 4% of total λbio transducing phages in the wild type or in the mutM mutant, respectively. The preferential increase of recombination at nonhotspot sites with hydrogen peroxide in the mutM mutant was discussed on the basis of a new model, in which 8-oxoG and/or Fapy residues may introduce double-strand breaks into DNA.

In vitro study on the effects of iron sucrose, ferric gluconate and iron dextran on redox-active iron and oxidative stress

2011 ◽  
Vol 60 (07) ◽  
pp. 459-465
Author(s):  
Brigitte Sturm ◽  
Hannes Steinkellner ◽  
Nina Ternes ◽  
Hans Goldenberg ◽  
Barbara Scheiber-Mojdehkar
Keyword(s):  
Oxidative Stress ◽  
In Vitro Study ◽  
Iron Sucrose ◽  
Vitro Study ◽  
Iron Dextran ◽  
Active Iron ◽  
Redox Active ◽  
And Oxidative Stress

Oxidative Stress and Vaso-Occlusion in Sickle Cell Disease: Role of Activated Leukocytes and Redox Active Iron.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3165-3165 ◽  
Author(s):  
John D. Belcher ◽  
Hemachandra Mahaseth ◽  
Thomas E. Welch ◽  
Felix Boakye-Agyeman ◽  
Robert P. Hebbel ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Sickle Cell Disease ◽  
Blood Cell ◽  
Sickle Cell ◽  
Ambient Air ◽  
Acute Chest Syndrome ◽  
Cell Disease ◽  
Active Iron ◽  
Redox Active ◽  
Hypoxia Reoxygenation

Abstract Sickle cell disease (SCD) is a disease of oxidative stress. We and others have demonstrated increased oxidative stress, inflammation, endothelial cell activation and white blood cell counts in human patients and transgenic murine models of SCD. Leukocytosis in SCD is associated with increases in the incidence of pain crisis, acute chest syndrome, stroke and mortality. We hypothesize that reactive oxygen species (ROS) derived from leukocytes and excess redox active iron promote vascular inflammation and vaso-occlusion. Leukocytes were activated in S+S-Antilles sickle mice compared to normal C57BL/6 control mice as measured by the percentage of leukocytes expressing CD11b on their surface in ambient air (25.4% vs. 19.3%, p<0.05) and after exposure of mice to hypoxia-reoxygenation (31.7% vs. 23.0%, p<0.05). In addition, resting leukocytes from S+S-Antilles mice produce 1.8-fold more H2O2 than normal mice (p<0.05) as measured by Amplex Red (10-acetyl-3,7-dihydroxyphenoxazine) fluorescence. These leukocyte oxidants are especially toxic in the presence of excess redox active iron. Histopathology of the lungs and livers of 10 week old S+S-Antilles and BERK sickle mice showed red blood cell (RBC) congestion compared to normal. In addition, the sickle livers had multiple areas of infarction and inflammatory leukocyte infiltration. The heme contents of S+S-Antilles sickle lungs and livers were increased by 37- and 4.9-fold, respectively, compared to normals (p<0.05 for both organs). Furthermore, there was significantly more chelatable iron that is potentially redox active as measured by Ferene-S in sickle lungs (21.0-fold, p<0.05) and livers (2.4-fold, p<0.05) compared to normals. Thus, these data demonstrate there is an explosive pro-oxidative environment in sickle mice. These excess oxidants lead to NF-kB activation, VCAM-1 and ICAM-1 expression, and increased oxidative injury, as seen histopathologically by nitro-tyrosine and dihydroethidium staining in organs. Hypoxia-reoxygenation, which induces RBC sickling and enhances ROS production in sickle mice, causes an increase in leukocyte rolling (4.4-fold, S+S-Antilles vs. normal, p<0.05) and adhesion (6.5-fold, p<0.05). Hypoxia-reoxygenation induces transient vaso-occlusion in 12% and 24% of the subcutaneous venules of S+S-Antilles and BERK mice respectively. No vessels become static in normal mice (p<0.05 sickle vs. normal). Hypoxia-reoxygenation-induced vaso-occlusion can be inhibited by antibodies to P-selectin, VCAM-1 or ICAM-1. Furthermore, scavenging ROS with the SOD and catalase mimetic, polynitroxyl albumin or the iron chelator Trimidox, inhibited hypoxia-reoxygenation-induced vaso-occlusion (p<0.05). We conclude that oxidative stress derived from activated leukocytes and excess redox active iron plays a critical role in promoting vaso-occlusion and organ injury in SCD. We speculate that iron chelators, leukocyte adhesion molecule blockade and anti-oxidants will modulate vaso-occlusion in patients with SCD.

scholarly journals Vitamin E Attenuates Oxidative Stress Induced by Intravenous Iron in Patients on Hemodialysis

2000 ◽  
Vol 11 (3) ◽  
pp. 539-549
Author(s):  
JOHANNES M. ROOB ◽  
GHOLAMALI KHOSCHSORUR ◽  
ANDREAS TIRAN ◽  
JÖRG H. HORINA ◽  
HERWIG HOLZER ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Lipid Peroxidation ◽  
Vitamin E ◽  
Oral Dose ◽  
Single Oral Dose ◽  
Intravenous Iron ◽  
Hemodialysis Session ◽  
Active Iron ◽  
Redox Active ◽  
Vitamin E Supplementation

Abstract. Intravenous iron application to anemic patients on hemodialysis leads to an “oversaturation” of transferrin. As a result, non-transferrin-bound, redox-active iron might induce lipid peroxidation. To test the hypothesis that vitamin E attenuates lipid peroxidation in patients receiving 100 mg of iron(II) hydroxide sucrose complex intravenously during a hemodialysis session, 22 patients were investigated in a randomized cross-over design, either with or without a single oral dose of 1200 IU of all-rac-α-tocopheryl acetate taken 6 h before the hemodialysis session. Blood was drawn before and 30, 60, 90, 135, and 180 min after the start of the iron infusion, and areas under the curve (AUC0-180 min) of ratios of plasma malondialdehyde (MDA) to cholesterol and plasma total peroxides to cholesterol (two markers of lipid peroxidation) were determined as the outcome variables. At baseline of the session without vitamin E supplementation, plasma α-tocopherol concentrations (27.6 ± 1.8 μmol/L) and ratios of α-tocopherol to cholesterol (5.88 ± 1.09 mmol/mol) were normal, plasma MDA concentrations were above normal (1.20 ± 0.28 μmol/L), and bleomycin-detectable iron (BDI), indicating the presence of redox-active iron, was not detectable. Upon iron infusion, BDI and MDA concentrations increased significantly (P < 0.001). BDI concentrations explained the increase over baseline in MDA concentrations (MDA = 1.29 + 0.075 × BDI). Vitamin E supplementation, leading to a 68% increase in plasma α-tocopherol concentrations, significantly reduced the AUC0-180 min of MDA to cholesterol (P = 0.004) and peroxides to cholesterol (P = 0.002). These data demonstrate that a single oral dose of vitamin E attenuates lipid peroxidation in patients on hemodialysis receiving intravenous iron. Given that intravenous iron is applied repeatedly to patients on hemodialysis, this therapeutic approach may protect against oxidative stress-related degenerative disease in the long term.

Abstract 265: Assessment Of Oxidative DNA Double Strand Break And Repair In Cardiomyocytes

2013 ◽  
Vol 113 (suppl_1) ◽  
Author(s):  
Bo Ye ◽  
Ning Hou ◽  
Lu Xiao ◽  
Haodong Xu ◽  
Faqian Li
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
Dna Damage ◽  
Dna Repair ◽  
Stress Responses ◽  
Strand Break ◽  
Neonatal Rat ◽  
Rat Cardiomyocytes ◽  
Repair Protein ◽  
Dna Repair Protein

Backgrounds: DNA damage occurs in cardiomyocytes during normal cellular metabolism and is significantly increased under cardiac stresses. How cardiomyocytes repair their DNA damage, especially DNA double strand breaks (DSBs), remains undetermined. We assessed DSBs caused by oxidative stress. More importantly, we investigated the spatiotemporal dynamics of DNA repair protein assembly/disassembly in DNA damage sites. Methods: Cultured neonatal rat cardiomyocytes were treated with different doses of hydrogen peroxide (H2O2) for 30 minutes to assess DNA damage response (DDR). To investigate the dynamics of DDR, cells were treated with 200 uM H2O2 and followed up to 72 hours. DSBs were evaluated by counting DNA damage foci after staining with antibody against histone H2AX phosphorylation at serine 139 (g-H2AX). The dynamics and posttranslational modification of DNA repair proteins were determined by Western blotting, immunolabeling, and confocal microscopy. Result: g-H2AX was proportionally increased to H2O2 dosage. Discrete nuclear g-H2AX foci were seen 30 minutes after hydrogen peroxide treatment with 50 uM, but became pannuclear when H2O2 was above 400 uM. At 200 uM of hydrogen peroxide, g-H2AX started to increase at 15 minutes and reached to highest levels at 60 minutes with up to 70 nuclear foci, started to decline at 2 hours, and returned to basal levels at 24 hours. DDR transducer kinase, ataxia telangiectasia mutated (ATM) was activated at 5 minutes with increased phosphorylation at serine 1981 (pATM) which started to decrease at 24 hours, but remained elevated up to 48 hours. Another DDR transducer kinase, ATM and Rad3-related (ATR) showed a biphasic activation at 30 minutes and 8 hours. ATM and ATR colocalized with g-H2AX. DNA damage mediator proteins such as MRN complex and p53BP1 were also recruited to sites of DNA damage at g-H2AX foci. Conclusions: DSBs and their repair have emerged as a new frontier of stress responses. Newly developed methods for studying g-H2AX and DNA repair protein dynamics can be explored to investigate DDR to oxidative stress in cardiomyocytes.

scholarly journals Constituents of French Marigold (Tagetes patulaL.) Flowers Protect Jurkat T-Cells against Oxidative Stress

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
Irakli Chkhikvishvili ◽  
Tamar Sanikidze ◽  
Nunu Gogia ◽  
Maia Enukidze ◽  
Marine Machavariani ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Hydrogen Peroxide ◽  
T Cells ◽  
Radical Scavenging ◽  
Jurkat Cells ◽  
Cellular Mechanisms ◽  
Jurkat T Cells ◽  
Radical Scavenging Capacity ◽  
Scavenging Capacity ◽  
Anti Inflammatory

The flowers of French marigold (Tagetes patulaL.) are widely used in folk medicine, in particular for treating inflammation-related disorders. However, cellular mechanisms of this activity demand further investigation. In the present work, we studied the potential ofT. patulacompounds to alleviate the oxidative stress in hydrogen peroxide-challenged human lymphoblastoid Jurkat T-cells. Crude extracts of marigold flowers and purified fractions containing flavonoids patuletin, quercetagetin, and quercetin and their derivatives, as well as the carotenoid lutein, were brought in contact with Jurkat cells challenged with 25 or 50 μM H2O2. Hydrogen peroxide caused oxidative stress in the cells, manifested as generation of superoxide and peroxyl radicals, reduced viability, arrested cell cycle, and enhanced apoptosis. The stress was alleviated by marigold ingredients that demonstrated high radical-scavenging capacity and enhanced the activity of antioxidant enzymes involved in neutralization of reactive oxygen species. Flavonoid fraction rich in quercetin and quercetagetin showed the highest cytoprotective activity, while patuletin in high dose exerted a cytotoxic effect associated with its anticancer potential.T. patulacompounds enhanced the production of anti-inflammatory and antioxidant interleukin-10 (IL-10) in Jurkat cells. Both direct radical-scavenging capacity and stimulation of protective cellular mechanisms can underlay the anti-inflammatory properties of marigold flowers.

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