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Effects of acute iron overload on Nrf2-related glutathione metabolism in rat brain

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Abstract

Iron (Fe) overload triggers free radical production and lipid peroxidation processes that may lead to cell death (ferroptosis). The hypothesis of this work was that acute Fe-dextran treatment triggers Nrf2-mediated antioxidant regulation in rat brain involving glutathione (GSH) metabolism. Over the initial 8 h after Fe-dextran administration (single dose of 500 mg Fe-dextran/kg), total Fe, malondialdehyde (MDA) content, glutathione peroxidase (GPx), GPx-Se dependent (GPx-Se) and glutathione S-transferases (GST) activities were increased in rat whole brain. The content of GSH and the activity of glutathione reductase (GR) showed decreases (p < 0.05) after 6 and 8 h post injection in cortex. A significant increase in nuclear Nrf2 protein levels over control values was achieved after 6 h of Fe-dextran administration, while no significant differences were observed in the cytosolic fraction. Nuclear Nrf2/cytosolic Nrf2 ratios showed enhancement (p < 0.05) after 6 h of Fe overload, suggesting a greater translocation of the factor to the nucleus. No significant differences were observed in the expression of Keap1 in nuclear or cytosolic extracts. It is concluded that acute Fe overload induces oxidative stress in rat brain with the concomitant lipid peroxidation increase and GSH depletion, leading to the elevation of Nrf2-controlled GPx, GPx-Se and GST protein expression as a protective adaptive response. Further studies are required to fully comprehend the complex network of interrelated processes keeping the balance of GSH functions as chelator, antioxidant and redox buffer in the understanding of the ferroptotic and hormetic mechanisms triggered by Fe overload in brain.

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References

  • Baitharu I, Jain V, Deep SN, Shroff S, Sahu JK, Naik PK, Ilavazhaganet G (2014) Withanolide A prevents neurodegeneration by modulating hippocampal glutathione biosynthesis during hypoxia. PLoS ONE 9:e105311

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Brumby PE, Massey V (1967) Determination of nonheme iron, total iron and cooper. Methods Enzymol 10:463–474

    Article  CAS  Google Scholar 

  • Candan N, Tuzmen N (2008) Very rapid quantification of malondialdehyde (MDA) in rat brain exposed to lead, aluminium and phenolic antioxidants by high-performance liquid chromatography-fluorescence detection. Neurotoxicology 29:708–713

    Article  CAS  PubMed  Google Scholar 

  • Carlberg I, Mannervik B (1985) Glutathione reductase. Methods Enzymol 113:484–490

    Article  CAS  PubMed  Google Scholar 

  • Chen X, Comish PB, Tang D, Kang R (2021) Characteristics and biomarkers of ferroptosis. Front Cell Dev Biol 9:637162. https://doi.org/10.3389/fcell.2021.637162

    Article  PubMed  PubMed Central  Google Scholar 

  • Cooper AJL, Pulsinelli WA, Duffy TE (1980) Glutathione and ascorbate during ischemia and postischemic reperfusion in rat brain. J Neurochem 35:1242–1245

    Article  CAS  PubMed  Google Scholar 

  • Dejanović B, Stevanovic I, Ninkovic M, Stojanovic I, Lavrnja I, Radicevic T, Pavlovic M (2016) Agmatine protection against chlorpromazine-induced forebrain cortex injury in rats. J Vet Sci 17:53–61

    Article  PubMed  PubMed Central  Google Scholar 

  • Dixon SJ, Lemberg KM, Lamprecht MR, Skouta R, Zaitsev EM, Gleason CE, Patel DN, Bauer AJ, Cantley AM, Yang WS, Morrison B 3rd, Stockwell BR (2012) Ferroptosis: an iron-dependent form of nonapoptotic cell death. Cell 149:1060–1072

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Fernández V, Vargas R, Castillo V, Cádiz N, Bastías D, Román S, Tapia G, Videla LA (2013) Reestablishment of ischemia-reperfusion liver injury by N-acetylcysteine administration prior to a preconditioning iron protocol. ScientificWorldJournal. https://doi.org/10.1155/2013/607285

    Article  PubMed  PubMed Central  Google Scholar 

  • Flohé L, Günzler WA (1984) Assays of glutathione peroxidase. Methods Enzymol 105:114–120

    Article  PubMed  Google Scholar 

  • Friedmann Angeli JP, Krysko DV, Conrad M (2019) Ferroptosis at the crossroads of cancer-acquired drug resistance and immune evasión. Nat Rev Cancer 19:405–414

    Article  CAS  PubMed  Google Scholar 

  • Galaris D, Barbouti A, Pantopoulos K (2019) Iron homeostasis and oxidative stress: An intimate relationship. Biochim Biophys Acta Mol Cell Res 1866:118535

    Article  CAS  PubMed  Google Scholar 

  • Gloire G, Legrand-Poels S, Piette J (2006) NF-κB activation by reactive oxygen secies: fifteen years later. Biochem Pharmacol 72:1493–1505

    Article  CAS  PubMed  Google Scholar 

  • Habig WH, Pabst MJ, Jakoby WB (1974) Glutathione S-transferases. The first enzymatic step in mercapturic acid formation. J Biol Chem 249:7130–7139

    Article  CAS  PubMed  Google Scholar 

  • Jozefczak M, Keunen E, Schat H, Bliek M, Hernandez LE, Carleer R, Remans T, Bohler S, Vangronsveld J, Cuypers A (2014) Differential response of Arabidopsis leaves and roots to cadmium: glutathione-related chelating capacity vs antioxidant capacity. Plant Physiol Biochem 83:1–9

    Article  CAS  PubMed  Google Scholar 

  • Kang YP, Mockabee-Macias A, Jiang C, Falzone A, Prieto-Farigua N, Stone E, Harris IS, DeNicola GM (2021) Non-canonical glutamate-cysteine ligase activity protects against ferroptosis. Cell Metab 33:174–189

    Article  CAS  PubMed  Google Scholar 

  • Kerins MJ, Ooi A (2018) The roles of NRF2 in modulating cellular iron homeostasis. Antioxid Redox Signal 29:1756–1773

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Kuang F, Liu J, Tang D, Kang R (2020) Oxidative damage and antioxidant defense in ferroptosis. Front Cell Dev Biol 8:586578

    Article  PubMed  PubMed Central  Google Scholar 

  • Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685

    Article  CAS  PubMed  Google Scholar 

  • Laurie SH, Tancock NP, McGrath SP, Sanders J (1991) Influence of complexation on the uptake by plants of iron, manganese, copper and zinc: I. Effect of EDTA in a multimetal and computer simulation study. Exp Bot 42:509–513

    Article  CAS  Google Scholar 

  • Lee OK, Jain AK, Papusha V, Jaiswal AK (2007) An autoregulatory loop between stress sensors INrf2 and Nrf2 controls their cellular abundance. J Biol Chem 282:36412–36420

    Article  CAS  PubMed  Google Scholar 

  • Lowry OH, Rosebrough NJ, Farr AL, Randall RJ (1951) Protein measurement with the Folin phenol reagent. J Biol Chem 193:265–275

    Article  CAS  PubMed  Google Scholar 

  • Maiorino M, Conrad M, Ursini F (2018) GPx4, lipid peroxidation, and cell death: discoveries, rediscoveries, and open issues. Antioxid Redox Signal 29:61–74

    Article  CAS  PubMed  Google Scholar 

  • Masaldan S, Bush AI, Devos D, Rolland AS, Moreau C (2019) Striking while the iron is hot: iron metabolism and Ferroptosis in neurodegeneration. Free Radic Biol Med 133:221–233

    Article  CAS  PubMed  Google Scholar 

  • Meister A, Anderson ME (1983) Glutathione. Annu Rev Biochem 52:711–760

    Article  CAS  PubMed  Google Scholar 

  • Moniruzzaman M, Ghosal I, Das D, Chakrabortys B (2018) Melatonin ameliorates H2O2-induced oxidative stress through modulation of Erk/Akt/NFκB pathway. Biol Res 51:17

    Article  PubMed  PubMed Central  CAS  Google Scholar 

  • Moon MS, McDevitt EI, Zhu J, Stanley B, Krzeminski J, Amin S, Aliaga C, Miller TG, Isom HC (2012) Elevated hepatic iron activates NF-E2-related factor 2-regulated pathway in a dietary iron overload mouse model. Toxicol Sci 129:74–85

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Niture SK, Kaspar JW, Shen J, Jaiswal AK (2010) Nrf2 signaling and cell survival. Toxicol Appl Pharmacol 244:37–42

    Article  CAS  PubMed  Google Scholar 

  • Piloni NE, Fernandez V, Videla LA, Puntarulo S (2013) Acute iron overload and oxidative stress in brain. Toxicology 314:174–182

    Article  CAS  PubMed  Google Scholar 

  • Piloni NE, Perazzo JC, Fernandez V, Videla LA, Puntarulo S (2016) Sub-chronic iron overload triggers oxidative stress development in rat brain: implications for cell protection. Biometals 29:119–130

    Article  CAS  PubMed  Google Scholar 

  • Piloni NE, Reiteri M, Hernando MP, Cervino CO, Puntarulo S (2017) Differential effect of acute iron overload on oxidative status and antioxidant content in areas of rat brain. Toxicol Pathol 45:1067–1076

    Article  CAS  PubMed  Google Scholar 

  • Piloni NE, Caro AA, Puntarulo S (2018) Iron overload prevents oxidative damage to rat brain after chlorpromazine administration. Biometals 31:561–570

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Piloni NE, Puntarulo S (2020) Effects of Fe chelators on Fe oxidative cellular metabolism. An update on the role in human health. SL Nutr Metabol 3:123

    Google Scholar 

  • Rae CD, Williams SR (2017) Glutathione in the human brain: review of its roles and measurement by magnetic resonance spectroscopy. Anal Biochem 15:127–143

    Article  CAS  Google Scholar 

  • Rehncrona S, Folbergrova J, Smith DS, Siesjo BK (1980) Influence of complete and pronounced incomplete cerebral ischemia and subsequent recirculation on cortical concentrations of oxidized and reduced glutathione in the rat. J Neurochem 34:477–486

    Article  CAS  PubMed  Google Scholar 

  • Riegman M, Sagie L, Galed C, Levin T, Steinberg N, Dixon SJ, Wiesner U, Bradbury MS, Niethammer P, Zaritsky A, Overholtzer M (2020) Ferroptosis occurs through an osmotic mechanism and propagates independently of cell rupture. Nat Cell Biol 22:1041–1048

    Article  CAS  Google Scholar 

  • Rodriguez-Ariza A, Toribio F, López-Barea J (1994) Rapid determination of glutathione status in fish liver using high-performance liquid chromatography and electrochemical detection. J Chromatog B 656:311–318

    Article  CAS  Google Scholar 

  • Sasazuki T, Okazaki T, Tada K, Sakon-Komazawa S, Katano M, Tanaka M, Yagita H, Okumura K, Tominaga N, Hayashizaki Y, Okazaki Y, Nakano H (2004) Genome wide analysis of TNF-inducible genes reveals that antioxidant enzymes are induced by TNF and responsible for elimination of ROS. Mol Immunol 41:547–551

    Article  CAS  PubMed  Google Scholar 

  • Schmidlin CJ, Dodson MB, Madhavan L, Zhang DD (2019) Redox regulation by NRF2 in aging and disease. Free Radic Biol Med 134:702–707

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Shah R, Shchepinov MS, Pratt DA (2018) Resolving the role of lipoxygenases in the initiation and execution of ferroptosis. ACS Cent Sci 4:387–396

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Stockwell BR, Friedmann-Angeli JP, Bayir H, Bush AI, Conrad M, Dixon SJ, Fulda S, Gascon S, Hatzios SK, Kagan VE, Noel K, Jiang X, Linkermann A, Murphy ME, Overholtzer M, Oyagi A, Pagnussat GC, Park J, Ran Q, Rosenfeld CS, Salnikow K, Tang D, Torti FM, Torti SV, Toyokuni S, Woerpel KA, Zhang DD (2017) Ferroptosis: a regulated cell death nexus linking metabolism, redox biology, and disease. Cell 171:273–285

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Smeyne M, Smeyne RJ (2013) Glutathione metabolism and Parkinson’s disease. Free Radic Biol Med 62:13–25

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Su LJ, Zhang JH, Gomez H, Murugan R, Hong X, Xu D, Jiang F, Peng ZY (2019) Reactive oxygen species-induced lipid peroxidation in apoptosis, autophagy, and ferroptosis. Oxid Med Cell Longev 2019:5080843

    Article  PubMed  PubMed Central  Google Scholar 

  • Sun Z, Wu T, Lau A, Birch CM, Zhang DD (2011) KPNA6 (importin α7)-mediated nuclear import of Keap1 represses the Nrf2-dependent antioxidant response. Mol Cell Biol 31:1800–1811

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Sun X, Ou Z, Chen R, Niu X, Chen D, Kang R, Tang D (2016) Activation of the p62-Keap1-NRF2 pathway protects against ferroptosis in hepatocellular carcinoma cells. Hepatology 63:173–184

    Article  CAS  PubMed  Google Scholar 

  • Tabatabaie T, Floyd RA (1994) Susceptibility of glutathione peroxidase and glutathione reductase to oxidative damage and the protective effect of spin trapping agents. Arch Biochem Bioph 314:112–119

    Article  CAS  Google Scholar 

  • Tebay LE, Robertsona H, Durantb ST, Vitalec SR, Penning TM, Dinkova-Kostovaa AT, Hayes JD (2015) Mechanisms of activation of the transcription factor Nrf2 by redox stressors, nutrient cues, and energy status and the pathways through which it attenuates degenerative disease. Free Radic Biol Med 88:108–146

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Towbin H, Staehelin T, Gordon J (1979) Electrophoretic transfer of proteins from polyacrylamide gels to nitrocellulose sheets: procedure and some applications. Proc Nat Acad Sci USA 76:4350–4354

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Vanden Berghe T, Linkermann A, Jouan-Lanhouet S, Walczak H, Vandenabeele P (2014) Regulated necrosis: the expanding network of non-apoptotic cell death pathways. Nat Rev Mol Cell Biol 15:135–147

    Article  CAS  PubMed  Google Scholar 

  • Wang D, Wang C, Wu H, Li Z, Ye Q (2016) Glutathione production by recombinant Escherichia coli expressing bifunctional glutathione synthetase. J Ind Microbiol Biotechnol 43:45–53

    Article  CAS  PubMed  Google Scholar 

  • Xie Y, Hou W, Song X, Yu Y, Huang J, Sun X, Kang R, Tang D (2016) Ferroptosis: process and function. Cell Death Differ 23:369–379

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  • Yang WS, Stockwell BR (2016) Ferroptosis: death by lipid peroxidation. Trends Cell Biol 26:165–176

    Article  CAS  PubMed  Google Scholar 

  • Yu H, Guo P, Xie X, Wang Y, Chen G (2017) Ferroptosis, a new form of cell death, and its relationships with tumourous diseases. J Cell Mol Med 21:648–657

    Article  CAS  PubMed  Google Scholar 

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Acknowledgement

This study was supported by Grants from the University of Buenos Aires (UBACyT 20020130100383BA) and CONICET (PIP 11220170100539CO) to S.P, and FONDECYT Chile (Grant 1150104) to LAV. S.P. is career investigator from CONICET.

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  1. Fisicoquímica-IBIMOL, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires, Junín 956, CAAD1113, Buenos Aires, Argentina

    Natacha E. Piloni & Susana Puntarulo

  2. Instituto de Bioquímica y Medicina Molecular (IBIMOL), CONICET-Universidad de Buenos Aires, Buenos Aires, Argentina

    Natacha E. Piloni & Susana Puntarulo

  3. Molecular and Clinical Pharmacology Program, Institute of Biomedical Sciences, Faculty of Medicine, University of Chile, Santiago, Chile

    Romina Vargas, Virginia Fernández & Luis A. Videla

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  1. Natacha E. Piloni
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Correspondence to Susana Puntarulo.

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Piloni, N.E., Vargas, R., Fernández, V. et al. Effects of acute iron overload on Nrf2-related glutathione metabolism in rat brain. Biometals 34, 1017–1027 (2021). https://doi.org/10.1007/s10534-021-00324-x

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