SRA/CD204 suppresses immunostimulatory activity of TLR4-engaged DCs in vitro. (A) Increased proliferation of OT-I cells primed by LPS-activated SRA^−/− DCs. BM-DCs derived from WT or SRA^−/− mice were pulsed with OVA protein (10 μg/mL) for 3 hours and subsequently activated with or without LPS for 2 hours. Cells were cocultured with OT-I cells for 72 hours. OT-I cell proliferation was measured by ³H-thymidine incorporation assays (*P < .005, LPS-treated SRA^−/− DC vs LPS-treated WT DC). (B) OVA-pulsed, LPS-stimulated BM-DCs were cocultured with 5 μM CFSE-labeled OT-I cells. OT-I cell proliferation was assessed on day 3 by FACS based on the dilution of CFSE intensity. Representative histograms from 2 independent experiments are shown (P < .001, SRA^−/− vs WT). (C) Increased IL-2 production by SRA^−/− DC-stimulated OT-I cells. After incubation, the levels of IL-2 in the supernatant were determined by ELISA. (D) DCs were pulsed with OVA_257-264 (1 μg/mL) and activated with LPS. After coculturing with OT-I cells, IL-2 levels were determined using an ELISA. Data shown as mean plus or minus SD (*P < .005, SRA^−/− vs WT). (E) Enhanced immunostimulatory capability of DCs from immunized SRA/CD204 KO mice. Splenic DCs were isolated by magnetic selection from immunized WT or SRA^−/− mice, and cocultured with OT-I cells for 3 days. ³H-TdR incorporation assays were used to determine OT-I cell proliferation was assayed (*P < .01, SRA^−/− vs WT). (F) Increased Pmel cell proliferation by LPS-stimulated SRA^−/− DCs. DCs were pulsed with gp100 protein antigens (20 μg/mL), and treated with or without LPS. DCs were then cocultured with gp100-specific CD8⁺ T cells purified from Pmel transgenic mice, followed by ³H-TdR incorporation assays (*P < .005, LPS-treated SRA^−/− DC vs LPS-treated WT DC). Representative results from 2 independent experiments are shown.