title: Reintroduction of C/EBPalpha in leukemic CD34+ stem/progenitor cells impairs self-renewal and partially restores myelopoiesis.
abstract: The CCAAT/enhancer binding protein (C/EBP) alpha transcription factor is indispensable for myeloid differentiation. In various myeloid leukemias, C/EBPalpha is mutated or functionally impaired due to decreased C/EBPalpha expression or phosphorylation. In order to investigate the functional consequences of decreased C/EBPalpha function in AML, we reintroduced C/EBPalpha in primary CD34(+) sorted acute myeloid leukemia (AML) cells using a lentiviral approach. Self-renewal and differentiation of primary AML stem cells were studied on long-term MS5 cocultures. Activation of C/EBPalpha immediately led to a growth arrest in all AML cultures (N = 7), resulting in severely reduced expansion compared with control cultures. This growth arrest corresponded with enhanced myeloid differentiation as assessed by fluorescence-activated cell sorter (FACS) analysis for CD14, CD15, and CD11b. Myeloid differentiation was further confirmed by the up-regulation of neutrophil elastase and granulocyte colony-stimulating factor (G-CSF) receptor in C/EBPalpha transduced cells. C/EBPalpha-expressing AML CD34(+) cells failed to generate second and third leukemic cobblestone areas (L-CAs) in serial replating experiments, while control cultures could be sequentially passaged for more than 4 times, indicating that reintroduction of C/EBPalpha impaired the self-renewal capacity of the leukemic CD34(+) compartment. Together, our data indicate that low C/EBPalpha levels are necessary to maintain self-renewal and the immature character of AML stem cells.
authors.affiliation: Division of Hematology, Department of Medicine, University Medical Center Groningen, Groningen, The Netherlands.
authors.forename: Hein
authors.initials: H
authors.lastname: Schepers
authors.forename: Albertus T J
authors.initials: AT
authors.lastname: Wierenga
authors.forename: Djoke
authors.initials: D
authors.lastname: van Gosliga
authors.forename: Bart J L
authors.initials: BJ
authors.lastname: Eggen
authors.forename: Edo
authors.initials: E
authors.lastname: Vellenga
authors.forename: Jan Jacob
authors.initials: JJ
authors.lastname: Schuringa
mesh_terms: D018952:Antigens, CD34 / Q000378:metabolism*; D001752:Blast Crisis; D001854:Bone Marrow Cells / Q000473:pathology; D022763:CCAAT-Enhancer-Binding Protein-alpha / Q000235:genetics / Q000494:pharmacology*; D002453:Cell Cycle / Q000235:genetics; D002454:Cell Differentiation; D049109:Cell Proliferation; D002478:Cells, Cultured; D006801:Humans; D016086:Lentivirus / Q000235:genetics; D015470:Leukemia, Myeloid, Acute / Q000378:metabolism / Q000473:pathology; D038042:Myelopoiesis; D010766:Phosphorylation; D013234:Stem Cells / Q000166:cytology*; D017154:Stromal Cells / Q000473:pathology; D014161:Transduction, Genetic
chemical_list: D018952:Antigens, CD34; D022763:CCAAT-Enhancer-Binding Protein-alpha