Dataset Information

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

ABSTRACT:

Background

β-thalassemic syndromes are inherited red cell disorders characterized by severe ineffective erythropoiesis and increased levels of reactive oxygen species whose contribution to β-thalassemic anemia is only partially understood.

Design and methods

We studied erythroid precursors from normal and β-thalassemic peripheral CD34(+) cells in two-phase liquid culture by proteomic, reverse transcriptase polymerase chain reaction and immunoblot analyses. We measured intracellular reactive oxygen species, heme levels and the activity of δ-aminolevulinate-synthase-2. We exposed normal cells and K562 cells with silenced peroxiredoxin-2 to H(2)O(2) and generated a recombinant peroxiredoxin-2 for kinetic measurements in the presence of H(2)O(2) or hemin.

Results

In β-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 β-thalassemic cells we found decreased heme levels related to significantly reduced activity of the first enzyme of the heme pathway, δ-aminolevulinate synthase-2 without differences in its expression. We demonstrated that the activity of recombinant δ-aminolevulinate synthase-2 is inhibited by both reactive oxygen species and hemin as a protective mechanism in β-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 β-thalassemia.

Conclusions

In β-thalassemic erythroid cells the reduction of δ-aminolevulinate synthase-2 activity and the increased expression of peroxiredoxin-2 might represent two novel stress-response protective systems.

SUBMITTER: De Franceschi L

PROVIDER: S-EPMC3208676 | biostudies-literature | 2011 Nov

REPOSITORIES: biostudies-literature

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Publications

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

De Franceschi Lucia L Bertoldi Mariarita M De Falco Luigia L Santos Franco Sara S Ronzoni Luisa L Turrini Franco F Colancecco Alessandra A Camaschella Clara C Cappellini Maria Domenica MD Iolascon Achille A

Haematologica 20110712 11

<h4>Background</h4>β-thalassemic syndromes are inherited red cell disorders characterized by severe ineffective erythropoiesis and increased levels of reactive oxygen species whose contribution to β-thalassemic anemia is only partially understood.<h4>Design and methods</h4>We studied erythroid precursors from normal and β-thalassemic peripheral CD34(+) cells in two-phase liquid culture by proteomic, reverse transcriptase polymerase chain reaction and immunoblot analyses. We measured intracellula ...[more]

PMID: 21750082

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Project description:BackgroundIneffective erythropoiesis (IE) is the primary cause of anemia and associated pathologies in β-thalassemia. The characterization of IE is imbalance of erythroid proliferation and differentiation, resulting in increased erythroblast proliferation that fails to differentiate and gives rise to enucleate RBCs. MicroRNAs (miRs) are known to play important roles in hematopoiesis. miR-155 is a multifunctional molecule involved in both normal and pathological hematopoiesis, and its upregulation is observed in patients with β-thalassemia/HbE. However, the expression and function of miR-155, especially in β-thalassemia, have not yet been explored.MethodsTo study miR-155 expression in thalassemia, erythroblast subpopulations, CD45-CD71+Ter-119+ and CD45-CD71-Ter-119+ were collected from β IVSII-654 thalassemic bone marrow. Additionally, a two-phase culture of mouse bone marrow erythroid progenitor cells was performed. Expression of miR-155 and predicted mRNA target genes, c-myc, bach-1 and pu-1, were determined by quantitative reverse transcription (qRT)-polymerase chain reaction (PCR) and normalized to small nucleolar RNA (snoRNA) 202 and glyceraldehyde-3-phosphate dehydrogenase (GAPDH), respectively. To investigate the effect of miR-155 expression, erythroblasts were transfected with miR-inhibitor and -mimic in order to elevate and eliminate miR-155 expression, respectively. Erythroid cell differentiation was evaluated by Wright-Giemsa staining and flow cytometry.ResultsmiR-155 was upregulated, both in vivo and in vitro, during erythropoiesis in β-thalassemic mice. Our study revealed that gain- and loss of function of miR-155 were involved in erythroid proliferation and differentiation, and augmented proliferation and differentiation of thalassemic mouse erythroblasts may be associated with miR-155 upregulation. miR-155 upregulation in β-thalassemic mice significantly increased the percentage of basophilic and polychromatic erythroblasts. Conversely, a significant decrease in percentage of basophilic and polychromatic erythroblasts was observed in β-thalassemic mice transfected with anti-miR-155 inhibitor. We also examined the mRNA targets (c-myc, bach-1 and pu-1) of miR-155, which indicated that c-myc is a valid target gene of miR-155 that regulates erythroid differentiation.ConclusionmiR-155 regulates IE in β-thalassemia via c-myc expression controlling erythroblast proliferation and differentiation.

Dataset Information

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

Background

Design and methods

Results

Conclusions

Publications

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

Similar Datasets

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