PIM2 expression is regulated not only by the intrinsic axis, but also by extrinsic stimulations in MM cells. (A) MM.1S cells were treated with MS-275 at the indicated concentration in the presence of 10 ng/mL of IL-6 or patient-derived BMSC-CM (20% of total cell culture media) for 48 hours. The whole cell lysates extracted were subjected to immunoblotting with the indicated antibodies. (B-C) MM.1S cells were transduced with shLuc, shIRF4 (#1) (B), or shHDAC1 (#1) (C). Transduced cells were stimulated or cocultured with IL-6 (10 ng/mL) or BMSC-CM (20% of total cell culture media) for 48 hours, and whole cell lysates were then extracted. Lysates were subjected to immunoblotting using the indicated antibodies. β-Actin served as the loading control. Relative expression levels of each target, which are normalized to its loading control, are shown below for each immunoblotting image. (D) MM.1S cells were treated with MS-275 at the indicated concentration in the presence of IL-6 (10 ng/mL) or BMSC-CM (20% of total cell culture media) for 48 hours. Cell viability was assessed by the CCK-8 assay. Error bars show the SD of triplicates. ∗∗P < .01, ∗∗∗P < .001 significantly different from each condition of the MS-275 treatment (0, 0.5, 1, and 2 μM) in the absence of IL-6 or BMSC-CM; the Tukey-Kramer multiple comparison test.