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MicroRNAs (miRNAs) have been shown to play an important role in hematopoiesis. To elucidate the role of miRNAs in the early steps of hematopoiesis, we directly compared donor‐matched CD133 +  cells with the more differentiated CD34 + CD133 −  and CD34 − CD133 −  cells from bone marrow on the miRNA and mRNA level. Using quantitative whole genome miRNA microarray and sequencing‐based profiling, we found that between 109 (CD133 + ) and 216 (CD34 − CD133 − ) miRNAs were expressed. Quantification revealed that the 25 highest expressed miRNAs accounted for 73% of the total miRNA pool. miR‐142‐3p was the highest expressed miRNA with up to 2,000 copies per cell in CD34 + CD133 −  cells. Eighteen miRNAs were significantly differentially expressed between CD133 +  and CD34 + CD133 −  cells. We analyzed their biological role by examining the coexpression of miRNAs and its bioinformatically predicted mRNA targets and luciferase‐based reporter assays. We provide the first evidence for a direct regulation of CD133 by miR‐142‐3p as well as tropomyosin 1 and frizzled homolog 5 by miR‐29a. Overexpression of miRNAs in CD133 +  cells demonstrated that miR‐142‐3p has a negative influence on the overall colony‐forming ability. In conclusion, the miRNAs expressed differentially between the CD133 +  and CD34 + CD133 −  cells are involved in inhibition of differentiation, prevention of apoptosis, and cytoskeletal remodeling. These results are highly relevant for stem cell‐based therapies with CD133 +  cells and delineate for the first time how the stem cell character of CD133 +  cells is defined by the expression of specific miRNAs. S TEM  C ELLS  2011;29:847–857

stem cells

Combined Characterization of microRNA and mRNA Profiles Delineates Early Differentiation Pathways of CD133 +  and CD34 +  Hematopoietic Stem and Progenitor Cells