To characterize the transcriptional program that governs myeloid differentiation, we performed microarray analyses of cells derived from a G-CSF-dependent in vitro myeloid differentiation system. Using a high speed cell sorter, we purified CD34+/Lin− hematopoietic progenitors from wild type C57Bl/6 murine bone marrow cells cultured for 3 days in stem cell factor (SCF), thrombopoietin (TPO), interleukin-3 (IL3), and FLT3 ligand. These enriched progenitors were then treated for 7 days with G-CSF and SCF, which induces a coordinate wave of myeloid maturation. Linearly amplified cRNAs were prepared from cells harvested on each day of differentiation (d0-d7), labeled, and hybridized with Affymetrix Mouse 430v2.0 arrays. Comparing the percentage changes within two biological replicate sets by statistical analysis, we found that 51.5% of expressed probesets (15,693) were differentially regulated between day 0 and day 7. Among them, 10,627 probesets (34.9%) displayed expression levels higher than 500 (the mean expression level of all probesets was normalized to a value of 1500 before analysis) on at least one day. To avoid the high variability associated with probesets displaying low hybridization signals, we focused on these 10,627 probesets. The data showed the expected expression patterns for several genes that are known to be regulated during myeloid differentiation, including C/EBPa, Pu.1, c-Myb, CDP, c-Jun, M-CSF, GATA-1, AML1, CBFb, NE, MPO, MMP9, CD11b, lysozyme, and several others. In addition, the expression profiles were very consistent with the array data obtained using highly enriched human bone marrow-derived cells at early, mid, and late stages of myeloid development (

Theilgaard-Monch et al., Blood 105: 1785, 2005
). However, our data also includes expression patterns from earlier stages of myeloid differentiation, which may be important to identify genes that are specifically expressed in committed myeloid progenitors. We organized the expressed genes into several categories, including transcription factors, cell cycle regulators, apoptosis genes, receptor and ligand in signal transduction pathways, etc. Some genes relevant for the pathogenesis of acute myelogenous leukemia (AML) also showed dynamic changes during normal myeloid differentiation, including (but not limited to) AML1, Flt3, HoxA9, Npm1, Ncoa2, Crebbp, Etv6, Meis1, Bcl2, Mcl-1, P53, Junb, Myc, and others. This dataset will be invaluable for evaluating the molecular events that underlie normal myeloid differentiation, and for defining the normal expression patterns of genes that are disregulated during the development of myeloid malignancies.

Topics:
bcl-2 protein, cancer, cell cycle regulator, colony-stimulating factors, crossbreeding, flt3 ligand, gata1 protein, human, gelatinase b, gene expression profiling, granulocyte colony-stimulating factor

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2005, The American Society of Hematology
2005
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