Stroma-derived IL-6 reduces Bcl-2/Bcl-x_Ldependence. (A-C) MM.1s was treated with the indicated concentrations of ABT-737 for 24 hours, and myeloma cell death determined by staining with annexin V-FITC and CD38. Data are presented as the mean ± SE of 3 independent experiments (*P < .05; **P < .01; ***P < .001; ****P < .0001). (A) Cells were incubated in the presence or absence of Hs-5 cells, Hs-5 conditioned media (CM, 50%), conditioned media from the stromal cells of a myeloma patient (BMSC CM, 50%), or IL-6 (10 ng/mL). (B) Cells were incubated in the presence or absence of Hs-5 conditioned media (0.5%) and the indicated concentrations of neutralizing IL-6 antibody. (C) Cells were incubated in the presence or absence of Hs-5 cells or conditioned media (0.5% or 50%) and the neutralizing anti-IL-6 antibody and/or anti-IL-6 receptor blocking antibody. (D) MM.1s was incubated with BMSCs from a normal donor (NBMSC) in the presence or absence of a neutralizing anti-IL-6 antibody (30 μg/mL) and treated with the indicated concentrations of ABT-737 for 24 hours. Myeloma cell death was determined by staining with annexin V-FITC and CD38. (E) CD138⁺ plasma cells (IPC) purified from a myeloma patient bone marrow aspirate were treated with ABT-737 and IL-6 (10 ng/mL). The total pool of mononuclear cells isolated by Ficoll separation (BCPC) were also treated with ABT-737 and a neutralizing anti-IL-6 antibody (30 μg/mL). Cell viability was determined by flow cytometry after staining with annexin V-FITC. Plasma cells were identified by staining with CD38 and CD45 (CD38⁺, CD45⁻).