pDCs are susceptible to the killing effect of bortezomib. (A) PBMCs (2 × 10⁶ cells/2 mL) were cultured without or with the indicated concentrations of bortezomib for 6 hours and washed to remove bortezomib. The cell concentration was then adjusted to 1 × 10⁶/mL and further cultured for 18 hours without bortezomib (Wash [+]). Alternatively, PBMCs (2 × 10⁶ cells/2 mL) were cultured without or with bortezomib for 24 hours (Wash [−]). After staining the PBMCs with mAbs to identify each cell population and with propidium iodide to exclude dead cells, viable cell numbers were counted using Flow-Count Fluorospheres by flow cytometry. Cell numbers in the presence of bortezomib relative to those in the absence of bortezomib were expressed as a percentage. Mo indicates monocytes. (B) Purified pDCs (4 × 10⁴ cells/200 μL) were cultured without or with the indicated concentrations of bortezomib for 3 hours, and stimulated with 0.5μM ODN2216 in the presence of bortezomib for 24 hours. The cells were stained with annexin V and propidium iodide and analyzed by flow cytometry. The numbers indicate the percentages of annexin V and propidium iodide double-negative viable cells. *P < .05; **P < .01; ***P < .001. The data are shown as means ± SE of 3 independent experiments. P values refer to the comparison between the data obtained without bortezomib and those obtained with each concentration of bortezomib.