Effect of Active Oxygen Radicals on Protein and Carbohydrate Moieties of Recombinant Human Erythropoietin

Uchida, Eiji; Morimoto, Kazushige; Kawasaki, Nana; Y, Izaki; Abdu, Aliyu; Hayakawa, Tetsuo

doi:10.3109/10715769709065769

Cited by 58 publications

(39 citation statements)

References 32 publications

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“…This result is in line with the lower heme levels in b-thalassemic late erythropoiesis than in normal late erythropoiesis. In addition we treated the control cells with 16 μM H2O2, as previously reported by Uchida et al, 24 and found a marked reduction in ALAS-2 activity and in heme content compared to the activity of normal controls at day 14 of culture (data not shown).…”

Section: D-aminolevulinate Synthase-2 Activity Is Reduced In B-thalassupporting

confidence: 54%

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Oxidative stress modulates heme synthesis and induces peroxiredoxin-2 as a novel cytoprotective response in -thalassemic erythropoiesis

Franceschi¹,

Bertoldi²,

Falco³

et al. 2011

Haematologica

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The online version of this article has a Supplementary Appendix. Backgroundb-thalassemic syndromes are inherited red cell disorders characterized by severe ineffective erythropoiesis and increased levels of reactive oxygen species whose contribution to b-thalassemic anemia is only partially understood. Design and MethodsWe studied erythroid precursors from normal and b-thalassemic peripheral CD34+ cells in twophase liquid culture by proteomic, reverse transcriptase polymerase chain reaction and immunoblot analyses. We measured intracellular reactive oxygen species, heme levels and the activity of d-aminolevulinate-synthase-2. We exposed normal cells and K562 cells with silenced peroxiredoxin-2 to H2O2 and generated a recombinant peroxiredoxin-2 for kinetic measurements in the presence of H2O2 or hemin. ResultsIn b-thalassemia the increased production of reactive oxygen species was associated with down-regulation of heme oxygenase-1 and biliverdin reductase and up-regulation of peroxiredoxin-2. In agreement with these observations in b-thalassemic cells we found decreased heme levels related to significantly reduced activity of the first enzyme of the heme pathway, d-aminolevulinate synthase-2 without differences in its expression. We demonstrated that the activity of recombinant d-aminolevulinate synthase-2 is inhibited by both reactive oxygen species and hemin as a protective mechanism in b-thalassemic cells. We then addressed the question of the protective role of peroxiredoxin-2 in erythropoiesis by exposing normal cells to oxidative stress and silencing peroxiredoxin-2 in human erythroleukemia K562 cells. We found that peroxiredoxin-2 expression is up-regulated in response to oxidative stress and required for K562 cells to survive oxidative stress. We then showed that peroxiredoxin-2 binds heme in erythroid precursors with high affinity, suggesting a possible multifunctional cytoprotective role of peroxiredoxin-2 in b-thalassemia. ConclusionsIn b-thalassemic erythroid cells the reduction of d-aminolevulinate synthase-2 activity and the increased expression of peroxiredoxin-2 might represent two novel stress-response protective systems.Key words: b-thalassemia, heme biosynthesis, oxidative stress, ROS, peroxiredoxin-2.Citation: De Franceschi L, Bertoldi M, De Falco L, Santos Franco S, Ronzoni L, Turrini F, Colancecco A, Camaschella C, Cappellini MD, and Iolascon A. Oxidative stress modulates heme synthesis and induces peroxiredoxin-2 as a novel cytoprotective response in b-thalassemic erythropoiesis. Haematologica 2011;96(11):1595-1604. doi:10.3324/haematol.2011 This is an open-access paper.Oxidative stress modulates heme synthesis and induces peroxiredoxin-2 as a novel cytoprotective response in b-thalassemic erythropoiesis

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Section: D-aminolevulinate Synthase-2 Activity Is Reduced In B-thalassupporting

confidence: 54%

“…24 Details on cytoflourimetric analysis are reporetd in the Online Supplementary Design and Methods. Whenever indicated, PRDX2 was immunoprecipitated from cell lysates (11¥10 6 cells).…”

Section: Peroxiredoxin-2: Cytofluorimetric and Immunoprecipitation Assaymentioning

confidence: 99%

Oxidative stress modulates heme synthesis and induces peroxiredoxin-2 as a novel cytoprotective response in -thalassemic erythropoiesis

Franceschi¹,

Bertoldi²,

Falco³

et al. 2011

Haematologica

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“…The glycosylated chains confer some of the biological activity to the EPO molecule, which is stabilized by the carbohydrate chains. The oligosaccharides in EPO may protect the protein structure from oxygen radical destruction (Uchida et al, 1997). In addition, the N-and O-linked chains may be necessary for the cellular secretion of EPO (Krantz, 1991).…”

Section: Discussionmentioning

confidence: 99%

Hematopoietic Factor Erythropoietin Fosters Neuroprotection through Novel Signal Transduction Cascades

Chong¹,

Kang²,

Maiese

2002

J Cereb Blood Flow Metab

187

143

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Summary:In addition to promoting the survival, proliferation, and differentiation of immature erythroid cells, erythropoietin and the erythropoietin receptor have recently been shown to modulate cellular signal transduction pathways that extend beyond the erythropoietic function of erythropoietin. In particular, erythropoietin has been linked to the prevention of programmed cell death in neuronal systems. Although this work is intriguing, the underlying molecular mechanisms that serve to mediate neuroprotection by erythropoietin are not well understood. Further analysis illustrates that erythropoietin modulates two distinct components of programmed cell death that involve the degradation of DNA and the externalization of cellular membrane phosphatidylserine residues. Initiation of the cascades that modulate protection by erythropoietin and its receptor may begin with the activation of the Janus tyrosine kinase 2 protein. Subsequent downstream mechanisms appear to lead to the activation of multiple signal transduction pathways that include transcription factor STAT5 (signal transducers and activators of transcription), Bcl-2, protein kinase B, cysteine proteases, mitogen-activated protein kinases, proteintyrosine phosphatases, and nuclear factor-B. New knowledge of the cellular pathways regulated by erythropoietin in neuronal environments will potentially solidify the development and initiation of therapeutic strategies against nervous system disorders.

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“…The glycosylated chains are important for the biological activity of EPO and can protect EPO from oxygen radical degradation. The carbohydrate chains stabilize EPO (Toyoda, et al, 2000) and the oligosaccharides in EPO may protect the protein from oxygen radical activity (Uchida, et al, 1997). The N-glycosylated chains are believed to contribute to the thermal stability of EPO (Tsuda, et al, 1988).…”

Section: Epo Expression Structure and Receptor Role In Cells And Timentioning

confidence: 99%

Erythropoietin and Oxidative Stress

Maiese

Chong

Hou

et al. 2008

CNR

108

113

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Unmitigated oxidative stress can lead to diminished cellular longevity, accelerated aging, and accumulated toxic effects for an organism. Current investigations further suggest the significant disadvantages that can occur with cellular oxidative stress that can lead to clinical disability in a number of disorders, such as myocardial infarction, dementia, stroke, and diabetes. New therapeutic strategies are therefore sought that can be directed toward ameliorating the toxic effects of oxidative stress. Here we discuss the exciting potential of the growth factor and cytokine erythropoietin for the treatment of diseases such as cardiac ischemia, vascular injury, neurodegeneration, and diabetes through the modulation of cellular oxidative stress. Erythropoietin controls a variety of signal transduction pathways during oxidative stress that can involve Janus-tyrosine kinase 2, protein kinase B, signal transducer and activator of transcription pathways, Wnt proteins, mammalian forkhead transcription factors, caspases, and nuclear factor κB. Yet, the biological effects of erythropoietin may not always be beneficial and may be poor tolerated in a number of clinical scenarios, necessitating further basic and clinical investigations that emphasize the elucidation of the signal transduction pathways controlled by erythropoietin to direct both successful and safe clinical care. KeywordsAlzheimer's disease; Akt; angiogenesis; apoptosis; cancer; cardiac; caspases; diabetes; endothelial; erythropoietin; forkhead; FoxO; GSK-3β; inflammation; mitochondria; NF-κB; renal; STATs; Wnt OXIDATIVE STRESSInitial work in pathways that can lead to oxidative stress by early investigators observed that increased metabolic rates could be detrimental to animals in an elevated oxygen environment. More current studies point to the potential aging mechanisms and accumulated toxic effects for an organism that are tied to oxidative stress (Maiese, et al., 2008a NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript in organisms, or at least the rate of oxygen consumption in organisms, has intrigued several investigators. Pearl proposed that increased exposure to oxygen through an increased metabolic rate could lead to a shortened life span (Pearl, 1928). Subsequent work by multiple investigators has furthered this hypothesis by demonstrating that increased metabolic rates could be detrimental to animals in an elevated oxygen environment . When one moves to more current work, oxygen free radicals and mitochondrial DNA mutations have become associated with oxidative stress injury, aging mechanisms, and accumulated toxicity for an organism (Yui and Matsuura, 2006).Oxygen free radicals can be generated in elevated quantities during the reduction of oxygen and subsequently lead to cell injury and apoptosis. Oxidative stress occurs as a result of the development of reactive oxygen species that consist of oxygen free radicals and other chemical entities. These agents can involve superoxide free radicals, hydrogen peroxide, sin...

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Effect of Active Oxygen Radicals on Protein and Carbohydrate Moieties of Recombinant Human Erythropoietin

Cited by 58 publications

References 32 publications

Oxidative stress modulates heme synthesis and induces peroxiredoxin-2 as a novel cytoprotective response in -thalassemic erythropoiesis

Oxidative stress modulates heme synthesis and induces peroxiredoxin-2 as a novel cytoprotective response in -thalassemic erythropoiesis

Hematopoietic Factor Erythropoietin Fosters Neuroprotection through Novel Signal Transduction Cascades

Erythropoietin and Oxidative Stress

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