Clonogenicity, gene expression and phenotype during neutrophil versus erythroid differentiation of cytokine-stimulated CD34+ human marrow cells in vitro.
With the objective to correlate clonogenicity, gene expression and phenotype during differentiation, human bone marrow CD34(+) cells were cultured in vitro to stimulate erythroid or neutrophil development, and sorted into five subpopulations according to their surface expression of CD15/CD33 and blood group antigen A/CD117 respectively. Sorted cells were cultured in methylcellulose and analysed by real-time reverse transcription polymerase chain reaction for expression of neutrophil and erythroid marker genes. Surface expression of CD15 coincided with restriction to neutrophil/monocyte differentiation and A antigen with restriction to erythroid differentiation. GATA-2 mRNA was down-regulated during both neutrophil and erythroid maturation, whereas GATA-1, SCL, ABO, erythropoietin receptor, Kell, glycophorin A, beta-globin and alpha-haemoglobin stabilizing protein were up-regulated during erythroid differentiation and silenced during neutrophil differentiation. CCAAT/enhancer-binding protein (C/EBP)-alpha, PU.1, granulocyte colony-stimulating factor receptor, PR3, C/EBP-epsilon and lactoferrin were sequentially expressed during neutrophil differentiation but rapidly down-regulated during the early erythroid stages. Nuclear factor erythroid-derived 2 (NF-E2) and glycophorin C were expressed both during neutrophil and erythroid differentiation. Our data support the notion of early expression of several lineage-associated genes prior to actual lineage commitment, defined by surface expression of CD15 and A antigen as markers for definitive neutrophil/monocyte and erythroid differentiation respectively. Previous findings, primarily from cell lines and mouse models, have been extended to adult human haematopoiesis.
Edvardsson
Louise
L
Department of Haematology, Biomedical Centre, Lund University, Lund, Sweden.
Dykes
Josefina
J
Olsson
Martin L
ML
Olofsson
Tor
T
D000328:Adult; D018952:Antigens, CD34 / Q000378:metabolism*; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D016207:Cytokines / Q000494:pharmacology*; D004912:Erythrocytes / Q000166:cytology*; D015672:Erythroid Precursor Cells / Q000166:cytology; D005434:Flow Cytometry; D015870:Gene Expression / Q000502:physiology*; D005838:Genotype; D006801:Humans; D066298:In Vitro Techniques; D009504:Neutrophils / Q000166:cytology*; D020133:Reverse Transcriptase Polymerase Chain Reaction
D018952:Antigens, CD34; D016207:Cytokines