Figure 6.
B-CLL cells treated with IL-21 plus CpG ODN can induce apoptosis of untreated, bystander B-CLL cells. Anti-granzyme B antibodies inhibit bystander B-CLL cell killing. (A) Purified B-CLL cells were split into 2 fractions. One fraction was stained with PKH-26, then incubated for 24 hours in IL-21 with or without CpG ODN. Unstained cells were maintained in culture without stimulus. The stained, treated cells were washed, added to the untreated cells, and cocultured for 2 days. Survival of the untreated cells (as indicated by lack of membrane dye) was analyzed by flow cytometry. Plotted are the B-CLL cell-survival rates for the untreated (ie, bystander) cells cultured at different ratios with treated cells. One representative experiment of 3 with similar results is shown. (B) Average B-CLL cell-survival rates for the untreated (ie, bystander) B-CLL cells cultured at a ratio of 1:2 with treated cells. Results are from 3 independent experiments. Error bars indicate SEM. (C) B-CLL cells from 3 patients were cultured for 4 days in the presence of IL-21 (100 ng/mL), CpG ODN (2.5 μg/mL), and anti-human granzyme B antibody ()ora control antibody (□). B-CLL cell survival was determined by FACS analysis using annexin V/PI staining and counterstaining with antibodies to CD19. Plotted are the mean B-CLL cell-survival rates in percent from 1 representative experiment of 3 with similar results. Error bars indicate SD.