Dexamethasone is a widely used anti-inflammatory agent for a broad spectrum of diseases. The effectiveness of this agent is thought to be due to the capacity to modulate cytokine production in inflammatory cells. We examined the effects of dexamethasone on expressions of interferon gamma (IFN-gamma) and interleukin 4 (IL-4) by human peripheral blood mononuclear cells (PBMCs) in response to interleukin 12 (IL-12). Dexamethasone (10-5 M) inhibited IFN-gamma secretion, through direct suppression of IFN-gamma, IL-12 receptor (IL-12R) -beta1, and -beta2 expressions. Conversely dexamethasone increased IL-4 secretion as well as IL-4 expressions by PBMCs in response to IL-12. In addition, dexamethasone increased expression of suppressor of cytokine signalling (SOCS)-1, which inhibits JAK-STAT pathway of IL-12R signalling. The result of our study suggested that dexamethasone directly inhibited IFN-gamma expression, through suppression of IL-12 signalling and indirectly increases IL-4 expression, through suppression of IFN-gamma expression.
Dexamethasone* ; Humans ; Interferons ; Interleukin-12* ; Interleukin-4* ; Receptors, Interleukin-12

IL-4 and IL-13 are closely related cytokines that are produced by Th2 cells. However, IL-4 and IL-13 have different effects on the development of asthma phenotypes. Here, we evaluated downstream molecular mechanisms involved in the development of Th2 type asthma phenotypes. A murine model of Th2 asthma was used that involved intraperitoneal sensitization with an allergen (ovalbumin) plus alum and then challenge with ovalbumin alone. Asthma phenotypes, including airway-hyperresponsiveness (AHR), lung inflammation, and immunologic parameters were evaluated after allergen challenge in mice deficient in candidate genes. The present study showed that methacholine AHR and lung inflammation developed in allergen-challenged IL-4-deficient mice but not in allergen-challenged IL-13-deficient mice. In addition, the production of OVA-specific IgG2a and IFN-gamma-inducible protein (IP)-10 was also impaired in the absence of IL-13, but not of IL-4. Lung-targeted IFN-gamma over-expression in the airways enhanced methacholine AHR and non-eosinophilic inflammation; in addition, these asthma phenotypes were impaired in allergen-challenged IFN-gamma-deficient mice. Moreover, AHR, non-eosinophilic inflammation, and IFN-gamma expression were impaired in allergen-challenged IL-12Rbeta2- and STAT4-deficient mice; however, AHR and non-eosinophilic inflammation were not impaired in allergen-challenged IL-4Ralpha-deficient mice, and these phenomena were accompanied by the enhanced expression of IL-12 and IFN-gamma. The present data suggest that IL-13-mediated asthma phenotypes, such as AHR and non-eosinophilic inflammation, in the Th2 type asthma are dependent on the IL-12-STAT4-IFN-gamma axis, and that these asthma phenotypes are independent of IL-4Ralpha-mediated signaling.
Allergens/immunology ; Animals ; Asthma/complications/*immunology/pathology/physiopathology ; Bronchial Hyperreactivity/complications/immunology/pathology ; Disease Models, Animal ; Interferon-gamma/*immunology ; Interleukin-12/*immunology ; Interleukin-12 Receptor beta 2 Subunit/metabolism ; Interleukin-13/deficiency/*immunology ; Interleukin-4/deficiency ; Methacholine Chloride ; Mice ; Mice, Transgenic ; Models, Immunological ; Organ Specificity ; Pneumonia/complications/immunology/pathology ; Receptors, Cell Surface/metabolism ; STAT4 Transcription Factor/*metabolism ; Signal Transduction/*immunology ; Th2 Cells/*immunology

Genetic Predisposition to Disease ; Humans ; Interleukin-12 Subunit p40 ; physiology ; Receptors, Interferon ; physiology ; Receptors, Interleukin-12 ; physiology ; Toll-Like Receptors ; physiology ; Tuberculosis ; genetics ; immunology

BACKGROUND: The type 2 deviated immunological state is predominant in lepromatous leprosy. Erythema nodosum leprosum (ENL) is an immune-complex mediated reaction that typically occurs in lepromatous leprosy. To date, the serum levels of tumor necrosis factor (TNF)-alpha, interleukin (IL)-2 receptor, IL-10, IL-1beta, IL-1 receptor antagonist and monocyte chemoattractant protein-1 (MCP-1) were reported to be higher in lepromatous leprosy. TNF-alpha is also known to be higher in ENL, which is reduced after thalidomide treatment. However the serum type 2 chemokine levels in lepromatous leprosy patients have not been reported. METHODS: The serum levels of the type 2 chemokines such as thymus and activation-regulated chemokine (TARC), macrophage-derived chemokine (MDC) and eotaxin together with IL-12 and IL-10 in the sera from leprosy patients were detected using an enzyme-linked solvent assay (ELISA) method. RESULTS: The Serum TARC, MDC, eotaxin, IL-10 and IL-12 levels in lepromatous leprosy patients were not significantly different from the normal control levels. The serum levels were not significantly different between the paucibacillary group and multibacillary group. The serum TARC or MDC levels in the ENL patients were more reduced after a treatment containing thalidomide. CONCLUSION: The type 2 chemokines are not related to the severity of lepromatous leprosy. The larger reducing effect of the TARC or MDC levels in ENL patients by a treatment containing thalidomide suggests the potential role of these chemokines in the development of ENL and the therapeutic mechanism of thalidomide.
Chemokine CCL17 ; Chemokine CCL2 ; Chemokine CCL22 ; Chemokines* ; Erythema Nodosum ; Humans ; Interleukin-1 ; Interleukin-10 ; Interleukin-12 ; Interleukins ; Leprosy ; Leprosy, Lepromatous* ; Receptors, Interleukin-10 ; Thalidomide ; Tumor Necrosis Factor-alpha

BACKGROUND AND OBJECTIVES: The etiology and pathogenesis of nasal polyp are still ill-defined and have been debated for many years. Recently, the identification and localization of mRNA of cytokines, chemokines, and growth factors that may be involved in the formation of nasal polyp have been studied. But, transcription factors that control the expressions of cytokines have not been studied. The purpose of this study is to investigate IL-12 and IL-4 mRNA in the polyps of the patients with allergy-associated and nonallergy-associated chronic sinusitis and compared it with controls. IL-12 receptor and IRF-1, c-maf and GATA-3 which are transcription factors of IFN-gamma, IL-4 and IL-5, respectively were also studied. MATERIALS AND METHOD: Nasal polyp tissues were surgically obtained from two groups of patients with chronic sinusitis: those who had allergic rhinitis (n=) and those without allergy (n=3). The normal nasal mucosa from inferior turbinate were obtained from normal subjects. IL-12p35, IL-12p40, IL-12Rbeta2, IRF-1, IL-4, GATA-3 and c-maf mRNA expression were analysed by means of the reverse transcription and polymerase chain reaction and southern blot in three groups. RESULTS: The expression of IL-12p40, IL-12Rbeta2, IRF-1 mRNA were significantly higher in the nonallergic polyp group than in the control group (p<0.05). GATA-3 mRNA was significantly expressed in allergic polyp group than in the control group (p<0.05). CONCLUSION: These results suggest IL-12, IL-12Rbeta2 and IRF-1 may be involved in nonallergic polyp formation. GATA-3 may contribute to allergic polyp formation.
Blotting, Southern ; Chemokines ; Cytokines ; Gene Expression* ; Humans ; Hypersensitivity ; Intercellular Signaling Peptides and Proteins ; Interleukin-12 ; Interleukin-12 Subunit p35 ; Interleukin-12 Subunit p40 ; Interleukin-4 ; Interleukin-5 ; Nasal Mucosa ; Nasal Polyps* ; Polymerase Chain Reaction ; Polyps ; Receptors, Interleukin-12 ; Reverse Transcription ; Rhinitis ; RNA, Messenger ; Sinusitis ; Transcription Factors ; Turbinates

BACKGROUND: Mycobacterial antigens released as PIM, LM, LAM, lipoproteins and other cellular factors may contribute to macrophage and dendritic cell activation through pattern recognition receptors such as TLRs. In this study, we assessed cytokine production and ERK activation with stimulation of several major mycobacterial antigens. METHODS: Purified mycobacterial antigens (10, 22, 30, 38kappaDa) and recombinant antigens (6, 16, 19, 38kappaDa, Ag85A antigen) were studied. The production of cytokines (TNF-alpha, IL-12, IL-6) was measured by ELISA. The ERK activation was detected by western blotting. The expression of TLR2 or TLR4 was measured by flow cytometry. RESULTS: Among purified antigens only 30kappaDa antigen induced production of IL-6 or TNF-alpha in THP-1 macrophage cells. When THP-1 macrophage cells were treated with 30kappaDa antigen, phosphorylation of ERK was detected. ERK activation also occurred in TLR2 transfectant HEK293 cells with 30kappaDa antigen stimulation. CONCLUSION: 30kappaDa antigen is one of the major mycobacterial antigens inducing cytokine production and MAP kinases phosphorylation in macrophages.
Blotting, Western ; Cytokines ; Dendritic Cells ; Enzyme-Linked Immunosorbent Assay ; Flow Cytometry ; HEK293 Cells ; Interleukin-12 ; Interleukin-6* ; Lipoproteins ; Macrophages* ; Phosphorylation ; Phosphotransferases ; Receptors, Pattern Recognition ; Tumor Necrosis Factor-alpha*

BACKGROUND: Dendritic cell (DC)-based vaccines are currently being evaluated as a novel strategy for tumor vaccination and immunotherapy. However, inducing long-term regression in established tumor-implanted mice is difficult. Here, we show that deoxypohophyllotoxin (DPT) induces maturation and activation of bone marrow-derived DCs via Toll-like receptor (TLR) 4 activation of MAPK and NF-kappaB. METHODS: The phenotypic and functional maturation of DPT-treated DCs was assessed by flow cytometric analysis and cytokine production, respectively. DPT-treated DCs was also used for mixed leukocyte reaction to evaluate T cell-priming capacity and for tumor regression against melanoma. RESULTS: DPT promoted the activation of CD8+ T cells and the Th1 immune response by inducing IL-12 production in DCs. In a B16F10 melanoma-implanted mouse model, we demonstrated that DPT-treated DCs (DPT-DCs) enhance immune priming and regression of an established tumor in vivo. Furthermore, migration of DPT-DCs to the draining lymph nodes was induced via CCR7 upregulation. Mice that received DPT-DCs displayed enhanced antitumor therapeutic efficacy, which was associated with increased IFN-gamma production and induction of cytotoxic T lymphocyte activity. CONCLUSION: These findings strongly suggest that the adjuvant effect of DPT in DC vaccination is associated with the polarization of T effector cells toward a Th1 phenotype and provides a potential therapeutic antitumor immunity.
Animals ; Dendritic Cells ; Immunotherapy ; Interleukin-12 ; Lymph Nodes ; Lymphocyte Culture Test, Mixed ; Lymphocytes ; Mice ; Phenotype ; Podophyllotoxin ; T-Lymphocytes ; Toll-Like Receptors ; Up-Regulation ; Vaccination ; Vaccines

Since cancer has become the second most common cause of death, next to heart disease and approximately 20% of human population dies from cancer, it is much desired to develop therapeutic anti-tumor vaccine with safety and efficacy. Here we investigated the immunostimulatory effects of B16 freezing/thawing (F/T) anti-tumor vaccine (hereafter F/T vaccine), one of whole cell anti-tumor vaccines. To this end, we took advantage of the IL12 p40 reporter system which is designed for monitoring the induction of IL12 expression via the detection of co-expressed yellow fluorescent protein. First, we examined whether F/T vaccine can induce IL12 expression using bone marrow-derived dendritic cells (BMDCs) from IL12 p40 reporter mice. Second, we examined whether F/T vaccine can change the expression level of MHC molecules and co-stimulatory molecules during the activation of dendritic cells. Third, to dissect what component of F/T vaccines accounts for the immunostimulatory activities, we examined the effect of F/T vaccine on BMDC activation after treating it with DNase or proteinase. Lastly, we used MyD88 knockout mice to investigate whether F/T vaccine activates BMDCs in a TLRdependent manner. We found that treatment of BMDCs with F/T vaccine induced IL12 expression as well as the increase of MHC II expression and co-stimulatory molecules such as CD86. Interestingly, we also found that F/T vaccine increased CD1d expression on BMDCs, which may influence the activation of natural killer T cells known to be involved in anti-tumor immune responses. In addition, we found that treatment of F/T vaccine with proteinase but not DNase abolished its immunostimulatory effect, indicating that proteins in F/T vaccine mainly have its adjuvant activity. Furthermore, the activation of BMDCs with F/T vaccine was dependent on MyD88 adaptor molecule. Taken together, our findings in this study demonstrated that the F/T vaccine might be one of the valuable reagents to provide a new insight for underlying mechanism of whole-cell anti-tumor vaccines and an important clue for the development of better therapeutic anti-cancer vaccines.
Animals ; Cause of Death ; Dendritic Cells ; Deoxyribonucleases ; Heart Diseases ; Humans ; Indicators and Reagents ; Interleukin-12 ; Mice ; Mice, Knockout ; Natural Killer T-Cells ; Toll-Like Receptors ; Vaccines

Interleukin-12 (IL-12), consisting of p35 and p40, plays important roles in linking innate and adaptive immunity. While p35 is constitutively expressed, IL-12 p40 gene expression is induced upon activation by Toll-like receptor ligands. Recently, with gene targeting technology, the cytokine IL-12 p40 reporter mouse has been developed to express the p40 gene linked via a viral IRES element with yellow fluorescence protein (YFP) fluorescent reporter. We investigated whether this novel system would be useful to reveal IL-12 p40-producing immune cells. We first investigated whether macrophages and dendritic cells from these mice faithfully reported p40 induction. Next, we tested if microglial cells, macrophages in the brain, could induce IL-12 p40. Finally we tested whether B cells could produce IL-12 p40 because there were very few reports for IL-12 production by B cells. Our results confirmed that macrophages and dendritic cells are main producer of IL-12 p40. Then, we found that microglial cells could produce IL-12 p40 upon stimulation with various TLR ligands. Finally we found that a subset of B cells could produce IL-12 p40 in TLR9-dependent manner. Taken all together, our system will be a valuable tool to identify the type of immune cells that produce IL-12 p40.
Adaptive Immunity ; Animals ; B-Lymphocytes ; Brain ; Corynebacterium ; Dendritic Cells ; Fluorescence ; Gene Expression ; Gene Targeting ; Interleukin-12 ; Ligands ; Macrophages ; Mice ; Microglia ; Toll-Like Receptors

OBJECTIVES: To assess innate and humoral immune responses in middle ear effusion of obese pediatric patients with otitis media with effusion (OME). METHODS: We evaluated 219 children with OME, of whom 21 were obese and 198 were non-obese. We compared the expression in middle ear effusion of mRNAs encoding toll-like receptors (TLR) 2, 4, 5, and 9; nucleotide-binding oligomerization domains (NOD) 1 and 2; retinoic acid-inducible gene (RIG)-I; interleukins (IL)-6, -10, and -12; interferon (IFN)-gamma; and tumor necrosis factor (TNF)-alpha mRNAs. We also compared the expression of immunoglobulins IgG, IgA, and IgM and the bacterial detection rate in the two groups. RESULTS: TLR2-mediated expression of IL-6 mRNA, TLR4-mediated expression of IL-6 and IL-10 mRNA, TLR5-mediated expression of IL-6, IL-10, and TNF-alpha mRNA, TLR9-mediated expression of IL-6 mRNA, and NOD2-mediated expression of IL-6, IL-12, and TNF-alpha mRNA were significantly lower in obese than in non-obese children (P<0.05). However, concentrations of IgG, IgA, and IgM in middle ear effusion were lower in obese than in non-obese children, but none of these differences was significant (P>0.05). CONCLUSION: Mean body mass index was higher and pattern-recognition receptor-mediated cytokine mRNA expression was lower in obese than in non-obese children with OME.
Bacteria ; Body Mass Index ; Child* ; Humans ; Immunity, Humoral ; Immunoglobulin A ; Immunoglobulin G ; Immunoglobulin M ; Immunoglobulins ; Interferons ; Interleukin-10 ; Interleukin-12 ; Interleukin-6 ; Interleukins ; Obesity ; Otitis Media with Effusion* ; Otitis Media* ; Otitis* ; RNA, Messenger* ; Toll-Like Receptors ; Tumor Necrosis Factor-alpha