Hemoglobin switching in unicellular erythroid culture of sibling erythroid burst-forming units: kit ligand induces a dose-dependent fetal hemoglobin reactivation potentiated by sodium butyrate.
Mechanisms underlying fetal hemoglobin (HbF) reactivation in adult life have not been elucidated; particularly, the role of growth factors (GFs) is controversial. Interestingly, histone deacetylase (HD) inhibitors (sodium butyrate, NaB, trichostatin A, TSA) reactivate HbF. We developed a novel model system to investigate HbF reactivation: (1) single hematopoietic progenitor cells (HPCs) were seeded in serum-free unilineage erythroid culture; (2) the 4 daughter cells (erythroid burst-forming units, [BFU-Es]), endowed with equivalent proliferation/differentiation and HbF synthesis potential, were seeded in 4 unicellular erythroid cultures differentially treated with graded dosages of GFs and/or HD inhibitors; and (3) HbF levels were evaluated in terminal erythroblasts by assay of F cells and gamma-globin content (control levels, 2.4% and 1.8%, respectively, were close to physiologic values). HbF was moderately enhanced by interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor treatment (up to 5%-8% gamma-globin content), while sharply reactivated in a dose-dependent fashion by c-kit ligand (KL) and NaB (20%-23%). The stimulatory effects of KL on HbF production and erythroid cell proliferation were strictly correlated. A striking increase of HbF was induced by combined addition of KL and NaB or TSA (40%-43%). This positive interaction is seemingly mediated via different mechanisms: NaB and TSA may modify the chromatin structure of the beta-globin gene cluster; KL may activate the gamma-globin promoter via up-modulation of tal-1 and possibly FLKF transcription factors. These studies indicate that KL plays a key role in HbF reactivation in adult life. Furthermore, combined KL and NaB administration may be considered for sickle cell anemia and beta-thalassemia therapy.
Gabbianelli
M
M
Department of Hematology and Oncology, Istituto Superiore di Sanità, Rome, Italy.
Testa
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Massa
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Pelosi
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Sposi
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Riccioni
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Luchetti
L
L
Peschle
C
C
D000328:Adult; D002087:Butyrates / Q000494:pharmacology*; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D004912:Erythrocytes / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D004921:Erythropoietin / Q000494:pharmacology; D005319:Fetal Hemoglobin / Q000096:biosynthesis*; D005812:Genes, Switch; D005914:Globins / Q000096:biosynthesis; D016178:Granulocyte-Macrophage Colony-Stimulating Factor / Q000494:pharmacology; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects / Q000502:physiology*; D006441:Hemoglobin A / Q000096:biosynthesis*; D006801:Humans; D007377:Interleukin-3 / Q000494:pharmacology; D008297:Male; D011994:Recombinant Proteins / Q000494:pharmacology; D019089:Stem Cell Factor / Q000494:pharmacology*
D002087:Butyrates; D007377:Interleukin-3; D011994:Recombinant Proteins; D019089:Stem Cell Factor; D004921:Erythropoietin; D016178:Granulocyte-Macrophage Colony-Stimulating Factor; D005914:Globins; D006441:Hemoglobin A; D005319:Fetal Hemoglobin