IFN-α induces differentiation and exhaustion of CML stem cells via induction of C/EBPβ in vitro. (A) Experimental protocol of serial replating assay. Lin⁻ c-kit⁺ Sca-1⁺ GFP⁺ cells were purified from WT or C/EBPβ KO BM cells transduced with MIG-BCR-ABL, and then subjected to a colony-forming assay in the presence or absence of 100 U/mL IFN-α. (B) Numbers of colonies at the indicated rounds of plating. First-round colonies were counted on day 7; second- and third-round colonies were counted on days 10 and 14, respectively. Colony numbers are normalized against the input (10 000 cells per dish). Data are means ± SD of triplicates. Representative data from 2 independent experiments are shown. (C) May-Giemsa staining of cells harvested from second-round colonies derived from WT and C/EBPβ KO cells transduced with BCR-ABL (left and right panels, respectively) (scale bars, 20 μm). (D) Representative flow cytometric data of cells recovered from the second-round colonies. CD11b expression levels are shown as histograms with red (control) or blue (+IFN-α) lines.