Figure 6.

dNP2-ctCTLA-4 inhibits Th2 cell differentiation in vitro. (a, b) CD4⁺CD62L^hiCD44⁻ naive CD4 T cells were isolated from 8-week-old BALB/c mice using magnetic activated cell sorting (MACS). The cells were incubated for 6 days in a 96-well plate with Th2 cell-skewing medium that contained coated anti-CD3/CD28 antibodies (0.1 μg ml⁻¹), anti-IFN-γ neutralizing antibody (5 μg ml⁻¹), IL-2 (50 U ml⁻¹) and IL-4 (30 ng ml⁻¹). Simultaneously, 1 μM of dNP2-EGFP, dNP2-ctCTLA-4 or PBS was added to the cell culture at day 0. On day 6, the cells were analyzed by flow cytometry after intracellular staining with anti-mouse IL-4-PE and anti-mouse IFN-γ-FITC FACS antibodies. (a) Dot plots are representative of three independent experiments. (b) The percentage of CD4⁺IL-4⁺ differentiated Th2 cells was analyzed. Results are presented as the mean±s.d. (n=3). Non-activated (NA): negative control group, which was not stimulated with anti-TcR antibodies nor cytokines. *P<0.05, **P<0.01.