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. 2014 Mar 28;192(9):4233–4241. doi: 10.4049/jimmunol.1302850

FIGURE 2.

FIGURE 2.

TLR2 agonist–conjugated peptides could be cross-presented by BMDCs via a TAP-independent pathway. BMDCs cultured from C57BL/6 WT or TAPKO mice were incubated with 1 μM Pam2IDG for 3 h, and free peptides were then washed away. (A) Seven days after C57BL/6 mice were immunized with BMDCs via i.v. injection, the cytotoxic ability of Ag-specific T cells was assessed via an in vivo killing assay. The specific lysis percentages of the target cells were determined by flow cytometry. The data represent the average of two independent experiments. (B) TC-1 tumor-bearing mice (six per group) were immunized with 2 × 105 peptide-pulsed BMDCs via i.v. injection. Average tumor sizes are shown (cm3). The data are expressed as the mean ± SD. WT BMDCs were pretreated with 25 μM chloroquine (CQ) or 5 μM cathepsin S (CS) inhibitor for 30 min and then pulsed with 1 μM Pam2IDG (C) or RAH (D) for 2.5 h at 37°C. After free peptides were washed away, 2 × 104 BMDCs were cocultured for 48 h with 2 × 105 CD8+ T cells that were purified from the splenocytes of RAH/IFA-immunized mice. IFN-γ–secreting cells were then detected by an ELISPOT assay. WT (E) or TAPKO (F) BMDCs were pretreated as in (C) but pulsed with 1 μM Pam2EQL for another 2.5 h at 37°C. After free peptides were washed away, 2 × 104 BMDCs were cocultured for 48 h with 2 × 105 CD8+ T cells that were purified from OT-I splenocytes. Cell proliferation was determined using a [3H]thymidine incorporation assay. The data shown are representative of two independent experiments. **p < 0.01.