Stromal cells decrease Bcl-2/Bcl-x_Ldependence of multiple myeloma. (A) Myeloma patient bone marrow aspirates were divided into either a buffy coat fraction containing myeloma and stromal cells or CD138⁺ purified myeloma cells and then treated with increasing concentrations of ABT-737 for 24 hours. Apoptosis of myeloma cells was measured by flow cytometry following staining for annexin V-FITC to calculate the IC₅₀ under the 2 conditions. Plasma cells were identified by staining with CD38 and CD45 (CD38⁺, CD45⁻). The IC₅₀ of myeloma in the presence of stromal cells is plotted along the y-axis, and the IC₅₀ of CD138⁺ purified cells is plotted on the x-axis. Each dot represents a single patient sample. Dashed lines represent the cutoff for ABT-737 sensitivity at 0.5 μM. (B) Hs-5 stromal cells and conditioned media induce resistance to ABT-737 but not bortezomib or arsenic trioxide in myeloma cell lines. MM.1s, 8226, KMS18, OPM2, and U266 were treated with the indicated drugs and concentrations for 24 hours in the presence or absence of Hs-5 cells or conditioned media (50%) before staining with CD38 to identify plasma cells and annexin V to measure apoptosis. (C) KMS21BM and OCI-My5 were treated with ABT-199 at the indicated concentrations for 24 hours in the presence or absence of Hs-5 cells or conditioned media (50%) before staining with CD38 to identify plasma cells and annexin V to measure apoptosis. Data are presented as the mean ± standard error (SE) of 3 independent experiments (*P < .05; **P < .01; ***P < .001).