Fig. 6.

Kinetic analysis of induction of tolerogenic dendritic cells (DCs). Non-obese diabetic (NOD) mice (4–6 weeks old) were treated with a variable number of injections (one to five) of α-galactosylceramide C26:0 (α-GalCer), C20:2 or vehicle [4 µg/dose spaced 2 days apart, intraperitoneally (i.p.)]. After 2 days of rest, spleen and pancreatic lymph node (PLN) suspensions were pooled and CD11c⁺ DCs from each group were sorted. Concurrently, the NOD-relevant V7 peptide (NRP-V7) peptide emulsified in incomplete Freund's adjuvant (IFA) was administered (100 µg/dose, i.p.) to a separate group of NOD mice to prime their CD8⁺ T cells. Fluorescence activated cell sorted (FACS) NRP-V7 peptide primed-CD8⁺ T cells were cultured with CD11c⁺ DCs in the presence of 100 µg/ml of NRP-V7 for 72 h. T cell proliferation and enzyme-linked immunosorbent assay (ELISA) assays of cytokine [interleukin (IL)-2, interferon (IFN)-γ, IL-4] concentrations in culture supernatants were then performed. Data are representative of two independent experiments yielding similar results and are expressed as mean ± standard deviation (s.d.). *Significant (P < 0·05) reduction compared to vehicle group.