Dose-response curve of CTLA-4 induction in CTCL cells. (A) Increasing numbers of DCs (103, 104, and 105 DCs/well) loaded with a constant number of apoptotic CTCL cells (▪) (2 × 105 cells/well) were used to stimulate a constant number of cocultured freshly purified CTCL cells (4 × 105 cells/well), isolated from the same original cultures. As controls, increasing numbers of nonloaded DCs (▴) or DCs pulsed with viable CTCL cells (treated with CD4 antibody, ♦) were added to the same number of freshly purified CTCL cells and were cocultured overnight. The percentage of cytoplasmic CTLA-4+ tumor cells was measured by flow cytometry. (B) CTCL cells and DCs were purified from cultures, and a constant number of DCs (1 × 106/well) were loaded with increasing numbers of apoptotic CTCL cells (6 × 104-6 × 106 CD3-treated apoptotic CTCL cells/well) and were used to stimulate 1.4 × 106 freshly purified responding CTCL cells. Results are presented as the percentage of cytoplasmic CTLA-4+ CTCL cells.
