As shown in Fig. 4, TREM-2-deficient DCs had more I-AbhighCD86high mature cells than WT DCs after CpG DNA and Zymosan stimulation. Importantly, the maturation level of TREM-2-deficient DCs was very similar to that of DAP12-deficient DCs, suggesting that TREM-2 signaling is mediated by DAP12 in BMDCs. We also compared TREM-2-deficient DCs to those deficient in both DAP12 and FcRγ. Similar to what we found for cytokine production, TREM-2-deficient DCs showed less CpG DNA- and Zymosan-induced maturation than DAP12/FcRγ-deficient DCs. Interestingly,

whereas WT, DAP12-deficient and TREM-2-deficient DCs had a similar amount of maturation in the absence of stimulus, DCs lacking both DAP12 and FcRγ consistently had less learn more basal maturation even though they had the highest amount of stimulus-induced click here maturation (Fig. 4B). In conclusion, these results show that TREM-2/DAP12 signaling negatively regulates DC TLR responses. It has been reported that Siglec-H is involved in the negative regulation of type I IFN responses through DAP12 signaling in plasmacytoid DCs (pDCs) 20, 21.

Though TREM-2 is not expressed in pDCs (Ito and Hamerman, unpublished data), we hypothesized that TREM-2 may inhibit type I IFN production in conventional DCs, such as BMDCs. We assessed IFN-α4 and IFN-β expression by qRT-PCR in WT and TREM-2-deficient BMDCs after CpG DNA stimulation. Expression of mRNAs encoding both type I IFNs analyzed were higher in TREM-2-deficient BMDCs compared with WT BMDCs at 2 and

6 h after stimulation (Fig. 5A and B). As expected, TREM-2-deficient BMDCs also expressed more mRNA encoding IL-12 p40 (il12b) at 2 and 6 h after CpG DNA treatment than WT BMDCs (Fig. 5C). Intriguingly, IRF7 expression was not changed between WT and TREM-2-deficient BMDCs (Fig. 5D). IRF7 is induced by type I IFN stimulation and plays a major role in the positive feedback regulation of type I IFN expression 22, 23. We also measured IFN-β secretion after 16 h of CpG DNA stimulation by ELISA. TREM-2-deficient BMDCs secreted significantly more IFN-β protein than WT BMDCs after CpG DNA stimulation (Fig. 5E). These results suggest that increased type I IFN response in TREM-2-deficient Aldol condensation DCs was due to lack of TREM-2/DAP12 signaling at the primary TLR response phase. In conclusion, these results demonstrate that TREM-2 negatively regulates DC production of type I IFN in addition to IL-12 p70 and TNF in response to TLR ligation. Because TREM-2-deficient BMDCs matured more efficiently than WT BMDCs, we investigated whether the antigen-presenting activity of TREM-2-deficient DCs was higher than that of WT DCs. We co-cultured OVA peptide-pulsed BMDCs in the presence of high (100 nM) and low (25 nM) doses of CpG DNA with CFSE-labeled OT-II TCR transgenic CD4+ T cells for 72 h and detected CFSE dilution of CD4+ T cells by flow cytometry (Fig. 6A).