FIGURE 3: Treatments using B7.1wa-DCs and multiligand-DCs result in modulation of immune response to BcAg in vivo.

BcAg-pulsed engineered DCs were injected i.v. once into 10-week-old pre-diabetic female NOD mice (2×10⁶ cells/mouse; single injection). A) Cohorts of mice (6 mice/group) were euthanized 15-days post-treatment, PnLN cells were labeled with CFSE, cultured in the presence of BcAg for 72h and examined for CFSE dilution by FACS. Fresh PnLN cells were also cultured in the presence of PMA/Ionomycin and Brefeldin A for 4h and subjected to intracellular staining and FACS to detect cytokine positive cell frequencies. In addition, fresh cells were examined for Foxp3+ cell frequencies by FACS. CD4+ T cells were gated for these analyses. B) Spleen cells from additional cohorts of mice (5 mice/group) were labeled using CFSE, stimulated using anti-CD3 antibody (2 μg/ml) for 72h, and CFSE dilution was examined by FACS. Supernatants from these cultures were tested for secreted cytokines by multiplex assay. P-value by Mann-Whitney test. All P-values are in comparison with control-DC group. This experiment was repeated once using an additional 4 mice/group and obtained comparable results.