Objective:To investigate the effects of β-glucan on the activation and functions of mouse bone marrow-derived dendritic cells (BMDCs) induced under different conditions.Methods:Mouse bone marrow from the tibia and fibula was in vitro induced into FL-DCs and GM-DCs by FMS-like tyrosine kinase 3 ligand (Flt-3L) and granulocyte-macrophage colony-stimulating factor (GM-CSF) in combination with IL-4 into, respectively. These cells were stimulated with whole glucan particles (WGP) and flow cytometry was performed to detect the expression of surface molecules such as CD40, CD80, MHCⅠ, MHCⅡ, CD8α, and CD11b on them. ELISA was used to detect the levels of IL-12p40, TNF-α, IL-6, and IL-10 in cell culture supernatants. The expression of each cytokine and chemokine receptor at the mRNA level was detected by real-time fluorescence quantitative PCR. CD8 + and CD4 + T cells in OT-Ⅰ and OT-Ⅱ mice were sorted out by magnetic bead sorting kit and co-cultured with FL-DCs and GM-DCs in the presence of ovalbumin (OVA). And then, the proliferation and differentiation of T cells were detected by flow cytometry. Results:WGP significantly promoted the expression of CD40, CD80, MHCⅠ, MHCⅡ and CD8α on FL-DCs and GM-DCs ( P<0.05). The secretion of IL-12p40 and TNF-α by FL-DCs and GM-DCs was significantly enhanced after WGP treatment ( P<0.05), while there are significant differences in the secretion of IL-6 and IL-10 between FL-DCs and GM-DCs. Real-time fluorescence quantitative PCR showed that WGP promoted the expression of chemokine receptors CXCR5 and CCR7 by GM-DCs ( P<0.001 and P<0.01), and the expression of CCR7 by FL-DCs ( P<0.0001). WGP promoted CD4 + T cells to secrete more IFN-γ and IL-17α when they were co-cultured with FL-DCs or GM-DCs ( P<0.05). Conclusions:WGP induces the maturation of BMDCs and improves the ability of BMDCs to activate effector T cells. Besides, WGP prompts the differentiation of BMDCs that are induced under different conditions to different Th cells.

Objective:To investigate the effect of tripartite motif-containing 23 (Trim23) on the differentiation and maturation of dendritic cells and the possible mechanism.Methods:Mouse bone marrow-derived dendritic cells (BMDCs) were prepared from bone marrow cells of C57BL/6 mice with the presence of Flt3L. Real-time quantitative PCR and Western blot were used to detect the expression of Trim23 in BMDCs after LPS stimulation. An overexpression vector for full-length Trim23 (Trim23 OE) was constructed and transfected into BMDCs, and the pcDNA3.1 empty vector was used as control. Flow cytometry was used to detect the expression of CD80, CD86, CD40 and MHCⅡ on the surface of vector-transfected BMDCs after LPS stimulation and ELISA was used to detect the secretion of IL-12p40, TNF-α, IL-6 and IL-10 by these cells. CD8 + and CD4 + T cells were isolated from spleen and lymph nodes of OT-Ⅰ and OT-Ⅱ mice by magnetic beads and co-cultured with LPS-treated BMDCs in the presence of ovalbumin (OVA). Flow cytometry was used to detect the proliferation and differentiation of CD8 + and CD4 + T cells. Western blot was performed to analyze the phosphorylation of p38, ERK1/2 and AKT in BMDCs. Two overexpression vectors for Trim23 mutants lacking RING or ARF domain (Trim23 ΔRING and Trim23 ΔARF) were constructed and transfected into BMDCs. Then flow cytometry and ELISA were used to detect the expression of surface molecules and cytokines. Results:The expression of Trim23 in BMDCs was significantly down-regulated after LPS stimulation. The expression of MHCⅡ, CD86 and CD80 and the secretion of TNF-α and IL-6 decreased significantly in BMDCs overexpressing Trim23. Furthermore, overexpression of Trim23 inhibited the ability of BMDCs to induce the proliferation and differentiation of CD4 + T cells and the proliferation of CD8 + T cells. Western blot showed that the phosphorylation of p38 and ERK1/2 decreased significantly in Trim23-overexpressing BMDCs. Compared with wildtype Trim23, overexpression of Trim23 ΔRING had no significant influence on the expression of surface molecules (MHCⅡ and CD86) and the secretion of cytokines (TNF-α and IL-6) in BMDCs stimulated by LPS. Conclusions:Trim23 overexpression inhibited the maturation and immune activation of BMDCs via MAPK signal pathway and its RING domain. This study provided reference for targeting Trim23 to improve the immune response of dendritic cell-based tumor vaccines.