Induction of CTLA-4 expression in CTCL cells. (A) Nonloaded DC-stimulated fresh CTCL cells and membrane CD3/cytoplasmic CTLA-4 expression was measured. Treg-cell induction was monitored by measuring the coexpression of (B) membrane CD3/cytoplasmic CTLA-4 and (C) membrane Vβ17 (antibody to the variable region of the beta chain of the TCR that is clonotypically expressed on the tumor cells)/cytoplasmic CTLA-4. (D) DCs pulsed with viable CTCL cells (incubated with CD4 antibody) provided a control for tumor-loaded DCs by measuring membrane CD3/cytoplasmic CTLA-4 and (E) membrane Vβ17/cytoplasmic CTLA-4. (F) Induced Treg cells coexpressed membrane CD25/cytoplasmic CTLA-4. Results are reported as percentage of positive cells and mean fluorescence intensity (MFI). Results are representative of 10 separate experiments performed on cells isolated from cultures of 8 different CTCL patients. (G) CTCL cells were induced to express cytoplasmic FoxP3, compared with isotype-matched control (1-color histogram), by overnight incubation with DCs loaded with apoptotic tumor cells in. (H) Isotype control (donkey antigoat PE) staining in CTCL cells is compared (i) with CTCL cocultivated overnight with DCs exposed to viable CTCL cells (ii) and CTCL cells isolated from 2 patients and cultured with DCs that had ingested apoptotic tumor cells (iii and iv). Histograms demonstrate side scatter gating on the lymphocyte population compared with FoxP3 PE staining.