C/EBP inhibits the growth of Flt3-ITD–transformed cells. (A) Western blot analysis of transfected 32D-Flt3-ITD cells. The 32D-Flt3-ITD stable cells were transfected with an ER-inducible C/EBPδ construct or empty vector. Cells were puromycin selected. Lysates were probed with anti-C/EBPδ and antiactin antibody. (B) Growth curve of C/EBPδ-expressing 32D-Flt3-ITD cells. 32D-Flt3-ITD cells with C/EBPδ or control cells were seeded in suspension cultures at the density of 2 × 105/mL in the presence of β-estradiol or ethanol. Cells were counted every 24 hours by trypan blue exclusion method. The results are shown as the mean ± SD of 2 independent experiments. (C) Colony assays were performed with 32D-Flt3-ITD cells expressing the C/EBPδ ER construct. A total of 1000 cells were seeded per dish with either β-estradiol or ethanol. Colonies were analyzed on day 8. Each bar represents the mean ± SD of a representative triplicate experiment. (D) Wright-Giemsa–stained cytospin slides under light microscopy. C/EBPδ or the empty vector–transduced 32D-Flt3-ITD cells were incubated with β-estradiol for induction of C/EBPδ. Representative morphologic changes seen at day 7 of β-estradiol treatment are shown.
