IL-23 and IL-12 are structurally similar and critical for the generation of efficient cellular immune responses. Toxoplasma gondii induces a strong cell-mediated immune response. However, little is known about IL-23 secretion profiles in T. gondii-infected immune cells in connection with IL-12. We compared the patterns of IL-23 and IL-12 production by THP-1 human monocytic cells in response to stimulation with live or heat-killed T. gondii tachyzoites, or with equivalent quantities of either T. gondii excretory/secretory proteins (ESP) or soluble tachyzoite antigen (STAg). IL-23 and IL-12 were significantly increased from 6 hr after stimulation with T. gondii antigens, and their secretions were increased with parasite dose-dependent manner. IL-23 concentrations were significantly higher than those of IL-12 at the same multiplicity of infection. IL-23 secretion induced by live parasites was significantly higher than that by heat-killed parasites, ESP, or STAg, whereas IL-12 secretion by live parasite was similar to those of ESP or STAg. However, the lowest levels of both cytokines were at stimulation with heat-killed parasites. These data indicate that IL-23 secretion patterns by stimulation with various kinds of T. gondii antigens at THP-1 monocytic cells are similar to those of IL-12, even though the levels of IL-23 induction were significantly higher than those of IL-12. The detailed kinetics induced by each T. gondii antigen were different from each other.
Antigens, Protozoan/*immunology ; Cell Line ; Humans ; Interleukin-12/*secretion ; Interleukin-23/*secretion ; Monocytes/*immunology/*parasitology ; Time Factors ; Toxoplasma/*immunology

OBJECTIVETo examine whether the enhanced expression of CD40L cDNA on murine ovarian cancer (OVHM) cells could induce the secretion of Th1 cytokines from dendritic cells (DC).

METHODSOVHM cells were transfected with the full-length mouse CD40L cDNA by lipofectamine 2000 and then G418 resistant cells as positive cells were selected. They were examined for their expression of CD40L with flow cytometry. Bone marrow cells were firstly depleted of erythrocytes, macrophages, T and B cells with PE-conjugated magnetic beads, and then cultured in 10% FCS RPMI 1640 medium supplemented with recombinant mouse GM-CSF and IL-4 for 10 days. PKH67-labeled tumor cells were cultured with DC, and then the stained cells were analyzed for the expression of MHC-I, MHC-II, CD80, CD86, CCR7 in DC with flow cytometry. The expression of p40, p19, p35, p28, EBI3 subunits, IL-18, IFN-gamma, Mig gene in cocultured DC-tumor cells were detected by RT-PCR.

RESULTSThe CD40L cDNA was successfully transfected into OVHM cells. Bone marrow-derived DCs, when cultured with CD40L/OVHM, formed clusters with the tumors and showed an upregulated expression of MHC- I, MHC-II, CD80, CD86, CCR7. Expression of the IL-12, IL-23, IL-27, IL-18, interferon-gamma (IFN-gamma) and Mig (monokine induced by IFN-gamma) genes was induced in the DCs that were cultured with CD40L/OVHM but not with OVHM cells.

CONCLUSIONThese data directly showed that the expression of CD40L on ovarian cancer cells facilitates the interaction between DCs and tumors, enhances the maturation of DCs, induces secretion of Th1 cytokines, especially for IL-12, IL-23 and IL-27, which maybe one of the possible antitumor mechanism for CD40L-transfected ovarian cancer cell line.

Animals ; CD40 Ligand ; genetics ; metabolism ; Cell Line, Tumor ; Cells, Cultured ; Coculture Techniques ; Cytokines ; secretion ; DNA, Complementary ; genetics ; Dendritic Cells ; cytology ; metabolism ; Female ; Interleukin-12 ; secretion ; Interleukin-23 ; secretion ; Interleukins ; secretion ; Mice ; Ovarian Neoplasms ; metabolism ; pathology ; Th1 Cells ; secretion ; Transfection

Anti-cancer drug bleomycin (BLM) can cause acute lung injury (ALI) which often results in pulmonary fibrosis due to a failure of resolving acute inflammatory response. The aim of this study is to investigate whether toll-like receptor (TLR) 2 mediates BLM-induced ALI, inflammation and fibrosis. BLM-induced dendritic cells (DCs) maturation was analyzed by flow cytometry and cytokine secretion was detected by the ELISA method. The expression and activity of p38 and ERK MAPK were determined with Western blotting. The roles of TLR2 in ALI, inflammation and fibrosis were investigated in C57BL/6 mice administered intratracheally with BLM. The results demonstrated that BLM-administered mice had higher expression of TLR2 (P<0.001) and its signaling molecules. Blocking TLR2 significantly inhibited the maturation of DCs and reversed BLM-stimulated secretion of cytokines in DCs, such as IL-6 (P<0.001), IL-17 (P<0.05) and IL-23 (P<0.05). TLR2 inhibition attenuated BLM-induced increase of inflammatory cells in bronchoalveolar lavage fluid (BALF), and reversed the immunosuppressive microenvironment by enhancing TH1 response (P<0.05) and inhibiting TH2 (P<0.001), Treg (P<0.01) and TH17 (P<0.01) responses. Importantly, blocking TLR2 in vivo significantly protected BLM-administered mice from pulmonary injury, inflammation and fibrosis and subsequently increased BLM-induced animal survival (from 50% to 92%). Therefore, TLR2 is a novel potential target for ALI and pulmonary fibrosis.
Acute Lung Injury ; chemically induced ; metabolism ; pathology ; Animals ; Bleomycin ; toxicity ; Bronchoalveolar Lavage Fluid ; Cells, Cultured ; Cytokines ; secretion ; Dendritic Cells ; cytology ; metabolism ; Inflammation ; chemically induced ; metabolism ; pathology ; Interleukin-17 ; secretion ; Interleukin-23 ; secretion ; Interleukin-6 ; secretion ; Lung ; metabolism ; MAP Kinase Signaling System ; Male ; Mice ; Mice, Inbred C57BL ; Pulmonary Fibrosis ; chemically induced ; metabolism ; pathology ; T-Lymphocytes, Regulatory ; drug effects ; Th1 Cells ; drug effects ; Th2 Cells ; drug effects ; Toll-Like Receptor 2 ; metabolism ; physiology