Protection of CLL B cells from fludarabine and MPS-induced apoptosis by imm-IgM. (A) Percentage of viable CLL cells determined by annexin V/PI staining after 48 hours in culture medium, in the presence of fludarabine, fludarabine and imm-IgM, or fludarabine and sol-IgM. Cells were preincubated for 3 hours with imm-IgM or sol-IgM prior to the addition of 20 μM fludarabine. Values represent mean of 7 U-CLL (left panel) and 7 M-CLL (right panel) cases. Error bars indicate SEM. F(ab′)2 fragments were used as sol-IgM in these experiments. (B) Cells were preincubated for 3 hours with sol-IgM-F(ab′)2 or imm-IgM prior to the addition of fludarabine (Flu, 20 μM) or MPS (20 μM). After 40 hours, cells were collected and analyzed for PARP cleavage and expression of Mcl-1 and Bcl-2 by immunoblotting. Membranes were stripped and reprobed with anti-β-actin to ensure equal protein loading. Of 8 experiments (5 M-CLL and 3 U-CLL cases), 2 representatives are presented.
