Expansion of BCR-ABL⁺senescent MgkL cells expressing TGF-β in colony formation assay. (A) The frequency of CD150⁺CD41⁺ MgkL cells of in vitro colony formation assay of BCR-ABL–transduced and nontransduced WT cells or DKO cells. The frequency was determined at the indicated time (n = 4 or 5). Senescence-associated (B) and SASP-related (C) gene expression. Either CD16/32⁺ myeloid-lineage or CD41⁺CD150⁺ MgkL cells were purified from in vitro colony formation assay of BCR-ABL–transduced and nontransduced WT cells or DKO cells on day 9. Total RNA was extracted from each cell population and subjected to qRT-PCR to determine the mRNA expression (n = 3 or 5). The expression level was calculated relative to the Gapdh gene and normalized to the averaged value of BCR-ABL⁻ myeloid-lineage cells. (D) Intracellular TGF-β1 expression. Cells were recovered from in vitro colony formation assay of BCR-ABL–transduced WT- (left) or DKO-derived KLS⁺ cells (right) on day 9 and subjected to intracellular TGF-β1 staining. Intracellular TGF-β1 expression was determined in each cell population using flow cytometry. MFIs were determined for each cell population (n = 4) and normalized to the averaged value of BCR-ABL⁻ total cells. Data are presented as the mean ± SD. Statistical significance was evaluated using the 1-way ANOVA with Tukey’s multiple-comparisons test. *P < .05; **P < .01.