BACKGROUNDChronic obstructive pulmonary disease (COPD) is characterized by progressive loss of lung function and local and systemic inflammation, in which CD8+ T-cells are believed to play a key role. Activated CD8+ T-cells differentiate into distinct subpopulations, including interferon-γ (IFN-γ)-producing Tc1 and interleukin (IL)-17-producing Tc17 cells. Recent evidence indicates that Tc17 cells exhibit considerable plasticity and may convert into IL-17/IFN-γ-double producing (Tc17/IFN-γ) cells when driven by inflammatory conditions. The aim of this study was to investigate the Tc17/IFN-γ subpopulation in peripheral blood of patients with COPD and to evaluate their potential roles in this disease.

METHODSPeripheral blood samples were collected from 15 never-smokers, 23 smokers with normal lung function, and 25 patients with COPD (Global Initiative for Chronic Obstructive Lung Disease 2-4). Proportions of the IL-17/IFN-γ-double expressing subpopulation were assessed using flow cytometry. Plasma concentrations of cytokines favoring Tc17/IFN-γ differentiation were measured by enzyme-linked immunosorbent assay.

RESULTSPatients with COPD had higher proportions of Tc17 cells and Tc17/IFN-γ cells in the peripheral blood than smokers and never-smokers. The plasticity of Tc17 cells was higher than that of Th17 cells. The percentages of Tc17 cells and Tc17/IFN-γ cells showed negative correlations with forced expiratory volume in 1 s % predicted value (r = -0.418, P = 0.03; r = -0.596, P = 0.002, respectively). The plasma concentrations of IL-6, transforming growth factor-β1, and IL-12 were significantly higher in patients with COPD compared with smokers and never-smokers.

CONCLUSIONSPeripheral Tc17 cells are increased and more likely to convert to Tc17/IFN-γ cells in COPD, suggesting that Tc17 cell plasticity may be involved in persistent inflammation of the disease.

Aged ; Female ; Forced Expiratory Volume ; Humans ; Interferon-gamma ; biosynthesis ; Interleukin-12 ; blood ; Interleukin-6 ; blood ; Lung ; physiopathology ; Male ; Middle Aged ; Pulmonary Disease, Chronic Obstructive ; immunology ; physiopathology ; Th17 Cells ; immunology

Dendritic cells (DCs) comprise two functionally distinct subsets: plasmacytoid DCs (pDCs) and myeloid DCs (mDCs). pDCs are specialized in rapid and massive secretion of type I interferon (IFN-I) in response to nucleic acids through Toll like receptor (TLR)-7 or TLR-9. In this report, we characterized a CD56(+) DC population that express typical pDC markers including CD123 and BDCA2 but produce much less IFN-I comparing with pDCs. In addition, CD56(+) DCs cluster together with mDCs but not pDCs by genome-wide transcriptional profiling. Accordingly, CD56(+) DCs functionally resemble mDCs by producing IL-12 upon TLR4 stimulation and priming naïve T cells without prior activation. These data suggest that the CD56(+) DCs represent a novel mDC subset mixed with some pDC features. A CD4(+)CD56(+) hematological malignancy was classified as blastic plasmacytoid dendritic cell neoplasm (BPDCN) due to its expression of characteristic molecules of pDCs. However, we demonstrated that BPDCN is closer to CD56(+) DCs than pDCs by global gene-expression profiling. Thus, we propose that the CD4(+)CD56(+) neoplasm may be a tumor counterpart of CD56(+) mDCs but not pDCs.
Biomarkers ; metabolism ; CD56 Antigen ; genetics ; immunology ; Cell Lineage ; genetics ; immunology ; Dendritic Cells ; immunology ; metabolism ; pathology ; Gene Expression ; Hematologic Neoplasms ; genetics ; immunology ; pathology ; Humans ; Immunophenotyping ; Interferon Type I ; biosynthesis ; metabolism ; Interleukin-12 ; biosynthesis ; metabolism ; Interleukin-3 Receptor alpha Subunit ; genetics ; immunology ; Lectins, C-Type ; genetics ; immunology ; Membrane Glycoproteins ; genetics ; immunology ; Myeloid Cells ; immunology ; metabolism ; pathology ; Receptors, Immunologic ; genetics ; immunology ; Terminology as Topic ; Toll-Like Receptor 4 ; genetics ; immunology ; Toll-Like Receptor 7 ; genetics ; immunology ; Toll-Like Receptor 9 ; genetics ; immunology

Legionella bacterium, an intracellular pathogen of mononuclear phagocytes, causes acute fatal pneumonia, especially in patients with impaired cellular immune responses. Until recently, however, the toll-like receptor (TLR) engagement of bacterial proteins derived from Legionella is uncertain. We previously showed that a 19-kDa highly conserved peptidoglycan-associated lipoprotein (PAL) of Legionella pneumophila induced the PAL-specific B cell and T cell responses in mice. In this study, we observed that the rPAL antigen of L. pneumophila, as an effector molecule, activated murine macrophages via TLR2 and produced proinflammatory cytokines such as IL-6 and TNF-alpha. In both BALB/c and TLR4-deficient C3H/HeJ mice, pretreatment of macrophages with anti-TLR2 mAb showed severely impaired cytokine production in response to the rPAL. In addition, in vitro the rPAL treatment increased the cell surface expression of CD40, CD80, CD86 and MHC I/II molecules. We further showed that the synthetic CpG-oligodeoxynucleotides (CpG ODN) coadministered with the rPAL enhanced IL-12 and IL-6 production and expression of CD40, CD80 and MHC II compared to the rPAL treatment alone. In conclusions, these results indicate that Legionella PAL might activate macrophages via a TLR2-dependent mechanism which thus induce cytokine production and expression of costimulatory and MHC molecules.
Animals ; Antigens, CD/immunology/metabolism ; Bacterial Outer Membrane Proteins/*pharmacology ; Cells, Cultured ; Female ; Histocompatibility Antigens Class II/immunology/metabolism ; Host-Pathogen Interactions ; Interleukin-12/biosynthesis ; Interleukin-6/biosynthesis ; Legionella pneumophila/*immunology/metabolism ; Legionnaires' Disease/immunology/metabolism ; Lipoproteins/*pharmacology ; Macrophage Activation/drug effects/immunology ; Macrophages, Peritoneal/drug effects/immunology/*metabolism ; Mice ; Mice, Inbred BALB C ; Mice, Inbred C3H ; Mice, Inbred C57BL ; Toll-Like Receptor 2/*metabolism ; Tumor Necrosis Factor-alpha/biosynthesis

Bacille Calmette-Guerin (BCG) induces potent Th1 responses with the help of interleukin (IL)-10 and IL-12 released from dendritic cells (DCs), and suppresses Th2- associated allergic reactions. However, there are still some controversies on therapeutic effects of BCG in asthmatics. This study investigated whether BCG administration to DCs suppresses IL-5 production from T cells in atopic asthmatics. DCs derived from peripheral blood of subjects were cultured with or without BCG and Dermatophagoides farinae extract. Some DCs were co-cultured with T cells in the presence of BCG or the above culture supernatants. In the atopic asthmatics, BCG significantly increased IL-10 and IL-12 production from DCs. In the presence of D. farinae extract, BCG further increased IL-10 production. BCG-induced IL-10 production was significantly higher in the atopics (n=14) than in the non-atopics (n=9). Both BCG and the BCG-treated DCs culture supernatant significantly increased IFN-gamma production from T cells. Both BCG and the supernatant from DCs+BCG+D. farinae co-cultures significantly decreased IL-5 production (all p<0.05), but the supernatant from DCs+BCG co-cultures did not. In conclusion, administration of BCG together with D. farinae extract effectively decreased IL-5 production from T cells, probably through the action of IL-10 and IL-12 released from DCs in D. farinaesensitive asthmatics.
Adult ; Asthma/*immunology ; BCG Vaccine/*immunology ; Dendritic Cells/*immunology ; Female ; Humans ; Interferon-gamma/biosynthesis ; Interleukin-10/biosynthesis ; Interleukin-12/biosynthesis ; Interleukin-5/*biosynthesis ; Lymphocyte Activation ; Male ; T-Lymphocytes/*immunology

BACKGROUNDSurvivin is a rather specific gene in tumor tissue. We transfected dendritic cells (DCs) with recombinant adenovirus (Ad) containing survivin gene and granulocyte-macrophage colony-stimulating factor (GM-CSF) gene and tested the inducing effect of the transfected DCs on cytotoxic T lymphocytes (CTL) to kill leukemic cells.

METHODSAfter derived from the peripheral, DCs was assayed by mixed leukocyte reaction (MLR) tests. Lactate dehydrogenase (LDH) release test was used to evaluate cytotoxicity of CTL.

RESULTSExpression of survivin in transfected DCs was confirmed by Western blotting analysis. GM-CSF expression was confirmed by enzyme-linked immunosorbent assay (ELISA). In MLR assay, DCs coinfected with Ad-survivin and Ad-GM-CSF induced higher allogeneic lymphocyte reaction than control DCs at ratios of 1:5, 1:10, 1:50 and 1:100. DCs coinfected with Ad-survivin and Ad-GM-CSF had much higher activity of CTL to HL-60 cells than DCs infected with Ad-survivin only, Ad-GM-CSF only, or control DCs. Levels of interleukin-12 (IL-12) and interferon gamma (IFN-gamma) in lymphocyte supernatants containing DCs coinfected with Ad-survivin and Ad-GM-CSF were significantly higher than those in the control group.

CONCLUSIONDCs coinfected with Ad-survivin and Ad-GM-CSF induce much higher anti-leukemic response in vitro than those infected with either factor. Therefore, adenovirus vectors containing survivin and GM-CSF genes may be promising vaccine candidates for leukemia therapy.

Adenoviridae ; genetics ; Cytotoxicity, Immunologic ; Dendritic Cells ; physiology ; ultrastructure ; Genetic Therapy ; Granulocyte-Macrophage Colony-Stimulating Factor ; genetics ; HL-60 Cells ; Humans ; Inhibitor of Apoptosis Proteins ; Interferon-gamma ; biosynthesis ; Interleukin-12 ; biosynthesis ; Leukemia ; therapy ; Lymphocyte Activation ; Microtubule-Associated Proteins ; genetics ; Neoplasm Proteins ; genetics ; T-Lymphocytes, Cytotoxic ; immunology ; Transfection

OBJECTIVETo observe the changes in the activity of dendritic cells (DCs) after carcino-embryonic antigen (CEA) gene transfection mediated by recombinant adeno-associated virus type2 (rAAV) and tumor cell lysate.

METHODSImmature DCs isolated from peripheral blood monocytes of HLA-A11-positive healthy volunteers were infected with the rAAV carrying CEA gene or loaded with tumor cell lysate. The surface markers of the DCs such as CD40, CD 1alpha, and CD86 were analyzed by flow cytometry. Interleukin-12 (IL-12) in the supernatants of DCs and interferon-gamma (IFN-gamma) released by the cytotoxic T lymphocytes (CTLs) were determined by ELISA detection kit. The specific killing activity of CTL against LoVo cells was assessed by MTT assay.

RESULTSThe DCs following antigen loading with the two methods both highly expressed CD40, CD86 and IL-12, and induced specific CTL that specifically recognized and killed LoVo cells, but the killing effect resulting from rAAV infection of the DCs was much better than that induced by tumor cell lysate loading.

CONCLUSIONBoth methods of antigen loading can induce mature DCs from peripheral blood monocyte cells, but rAAV infection of the DCs can be more effective than tumor cells lysate loading. DCs infected with rAAV may have the potential to serve as an adjuvant immunotherapy for patients with colorectal carcinoma.

B7-2 Antigen ; metabolism ; CD40 Antigens ; metabolism ; Cancer Vaccines ; biosynthesis ; immunology ; Carcinoembryonic Antigen ; genetics ; Cell Line, Tumor ; Colorectal Neoplasms ; therapy ; Dendritic Cells ; immunology ; metabolism ; Dependovirus ; genetics ; Genetic Vectors ; Humans ; Interleukin-12 ; metabolism ; Transfection

OBJECTIVETo investigate the characteristics of circulating type II pre-dendritic cells (pDC2) and evaluate its role in patients with chronic hepatitis B virus infection.

METHODSThe quantitative alterations of pDC2 in 27 chronic HBV-infected patients as treated group and 15 healthy individuals as a control group were analyzed by using flow cytometry based on the comparison of CD4+/CD8+ ratios of T lymphocyte subsets between the two groups. The IFN-alpha-producing ability of pDC2 after incubation was determined by ELISA.

RESULTSThe percentage of pDC2 (0.096 +/- 0.086) from the peripheral blood in chronic HBV-infected patients were significantly lower than that (0.304 +/- 0.093) from the normal controls (P less than 0.001) while the CD4+/CD8+ ratios were higher than those in normal controls (P less than 0.01). The values of IFN-alpha-producing function and IL-12 of circulating pDC2 in chronic HBV-infected patients group were significantly lower than those in healthy subjects (P < 0.001). The percentage of pDC2 and CD4+/CD8+ ratios were higher in the patients positive for HBV DNA in sera than those in patients negative for HBV DNA in sera (P < 0.01).

CONCLUSIONThe decreased number of circulating pDC2 and IFN-alpha-producing function from peripheral blood in patients with chronic hepatitis B virus infection may result in the decline of host immune response, which may partially contribute to the disease progress of HBV infection and existence of viral genomic DNA in patient's sera.

Adolescent ; Adult ; CD4-CD8 Ratio ; Cell Count ; DNA, Viral ; blood ; genetics ; Dendritic Cells ; cytology ; immunology ; metabolism ; Enzyme-Linked Immunosorbent Assay ; Female ; Hepatitis B virus ; genetics ; growth & development ; Hepatitis B, Chronic ; blood ; immunology ; virology ; Humans ; Interleukin-12 ; biosynthesis ; Male ; Middle Aged ; T-Lymphocyte Subsets ; cytology ; immunology ; Young Adult

For treating Leishmania major infection in BALB/c mice, we used thalidomide in conjunction with glucantime. Groups of mice were challenged with 5 x 10(3) metacyclic promastigotes of L. major subcutaneously. A week after the challenge, drug treatment was started and continued for 12 days. Thalidomide was orally administrated 30 mg/kg/day and glucantime was administrated intraperitoneally (200 mg/kg/day). It was shown that the combined therapy is more effective than single therapies with each one of the drugs since the foot pad swelling in the group of mice received thalidomide and glucantime was significantly decreased (0.9 +/- 0.2 mm) compared to mice treated with either glucantime, thalidomide, or carrier alone (1.2 +/- 0.25, 1.4 +/- 0.3, and 1.7 +/- 0.27 mm, respectively). Cytokine study showed that the effect of thalidomide was not dependent on IL-12; however, it up-regulated IFN-gamma and down-regulated IL-10 production. Conclusively, thalidomide seems promising as a conjunctive therapy with antimony in murine model of visceral leishmaniasis.
Time Factors ; Thalidomide/pharmacology/*therapeutic use ; Organometallic Compounds/pharmacology/*therapeutic use ; Mice, Inbred BALB C ; Mice ; Meglumine/pharmacology/*therapeutic use ; Leishmaniasis, Visceral/*drug therapy/immunology ; Leishmania major/*drug effects ; Interleukin-12/analysis/biosynthesis ; Interleukin-10/analysis/biosynthesis ; Interferon Type II/analysis/biosynthesis/drug effects ; Immunosuppressive Agents/pharmacology/*therapeutic use ; Female ; Drug Therapy, Combination ; Disease Progression ; Disease Models, Animal ; Cells, Cultured ; Antiprotozoal Agents/pharmacology/*therapeutic use ; Animals

OBJECTIVETo study the effect of bifidobacterium genomic DNA on umbilical cord blood mononuclear cell (CBMC), and investigate the immunoregulation of bifidobacterium DNA and explore possible mechanisms by which bifidobacterium acts against allergic reaction.

METHODSBifidobacterium genomic DNA (bDNA) and human DNA (hDNA) were extracted with phenol/chloroform/isoamyl alcohol and stored at -20 degrees C for later use. Parts of bDNA were completely digested with DNaseI (d-bDNA) at 37 degrees C. CBMCs were isolated with Ficoll from umbilical cord blood and incubated at 37 degrees C in a 5% CO2 humidified incubator. These cells were divided into four groups, control group: without any stimulant; bDNA group: stimulated with 25 microg/ml bDNA; d-bDNA group: stimulated with 25 microg/ml d-bDNA; hDNA group: stimulated with 25 microg/ml hDNA. The cells were stimulated with different stimulants in vitro, at the end of incubation culture supernatant and cells were collected. IL-12 and IL-10 levels in the culture supernatant were measured by enzyme linked immuno sorbent assay (ELISA); cells secreting IL-4 and IFN-gamma were counted by enzyme linked immunospot (ELISPOT) assay; and total RNA was isolated from the cells to assay T-bet and GATA3 mRNA expression levels by reverse transcription polymerase chain reaction (RT-PCR).

RESULTSSix hours after stimulation there was no significant difference in IL-12 level in supernatant among the four groups; 12 hours after stimulation, IL-12 level in supernatant of bDNA treated group was significantly higher than that of each of the other groups, so were the results obtained at 24 hours and 48 hours after stimulation (P < 0.05). No significant difference could be detected in IL-12 level in supernatant among the other 3 groups. On the other hand, 6 hours after stimulation there was no significant difference in IL-10 level in supernatant among the four groups. But 12 and 24 hours after stimulation IL-10 level in supernatant of bDNA treated group was lower than that of each of the other groups, but the difference was not statistically significant. The count of IFN-gamma secreting cells of bDNA treated group was higher than that of the other groups, while IL-4 secteting cells of bDNA treated group were lower than that of the other groups. After bDNA stimulation, nuclear factor T-box expressed in T cells (T-bet) mRNA expression was conspicuously enhanced as compared to the other three groups (P < 0.05). GATA3 mRNA transcription in CBMC had no significant change after bDNA stimulation.

CONCLUSIONbDNA could promote secretion of Th1 type cytokine IL-12, while Th2 type cytokine IL-10 level of cell supernatant was decreased. bDNA could stimulate secretion of IFN-gamma by CBMC and inhibit secretion of IL-4. T-bet mRNA expression was highly enhanced after bDNA stimulation. bDNA could upregulate Th1 type response, which may be one of important mechanisms by which bifidobacterium inhibit allergic response.

Bifidobacterium ; cytology ; genetics ; Cell Culture Techniques ; DNA, Bacterial ; biosynthesis ; metabolism ; pharmacology ; Enzyme-Linked Immunosorbent Assay ; Fetal Blood ; cytology ; immunology ; GATA3 Transcription Factor ; genetics ; Humans ; Infant, Newborn ; Interferon-gamma ; immunology ; secretion ; Interleukin-10 ; immunology ; secretion ; Interleukin-12 ; immunology ; secretion ; Interleukin-4 ; immunology ; secretion ; Leukocytes, Mononuclear ; immunology ; secretion ; RNA, Messenger ; isolation & purification ; Reverse Transcriptase Polymerase Chain Reaction ; T-Box Domain Proteins ; genetics ; Th1 Cells ; drug effects ; immunology ; secretion

OBJECTIVETo study the changes of IL-12 produced by dendritic cells in peripheral blood in children with Henoch-Schonlein purpura (HSP), and to explore its influence on TH1/TH2 balance in order to elucidate its significance in the pathogenesis of HSP.

METHODSThe levels of interferon-gamma (IFN-gamma), interleukin-4 (IL-4) and interleukin-12 (IL-12) in plasma were determined by ELISA in 60 HSP children (HSP group) and 21 healthy children (Control group). Peripheral blood mononuclear cells (PBMC) of 22 HSP patients and 21 healthy children were cultured in vitro and then were transformed into dendritic cells. The levels of IL-12 in the supernatant were detected by ELISA and the positive expression rate of CD1a(+) was detected by indirect immunofluorescence procedure.

RESULTS1) The levels of IFN-gamma and the ratio of IFN-gamma/IL-4 in plasma of the HSP group were lower than those of the Control group (IFN-gamma 30.59 +/- 11.27 pg/mL vs 43.38 +/- 19.19 pg/mL; IFN-gamma/IL-4 ratio 0.70 +/- 0.28 vs 1.33 +/- 0.57) (P < 0.01). The levels of IL-12 in the HSP group were also lower than those of the Control group (153.95 +/- 91.88 pg/mL vs 323.06 +/- 162.34 pg/mL; P < 0.01). In contrast, the levels of IL-4 were higher than those of the Control group (45.08 +/- 9.19 pg/mL vs 32.95 +/- 7.10 pg/mL; P < 0.01). The plasma levels of IL-12 positively correlated with the IFN-gamma levels (r=0.52, P < 0.01) and the ratio of IFN-gamma/IL-4 (r=0.43, P < 0.01) in the HSP group. 2) The IL-12 levels in the supernatant of the HSP group were lower than those of the Control group (357.06 +/- 153.56 pg/mL vs 489.80 +/- 213.45 pg/mL; P < 0.05), and had a positive correlation with the plasma IL-12 levels (r=0.74, P < 0.01). 3) The positive expression rate of CD1a(+) of the HSP group was lower than that of the Control group [(27.42 +/- 10.75)% vs (35.68 +/- 12.18)%; P < 0.05], and positively correlated with the IL-12 levels in the supernatants (r=0.57, P < 0.01) and in plasma (r=0.68, P < 0.01).

CONCLUSIONSThere was an imbalance of TH1/TH2 in HSP children. The decrease of TH1 function had a positive correlation with the low levels of IL-12 in plasma, while the latter correlated closely with decreased number and / or function of dendritic cells, suggesting that the decreased number and / or function of dendritic cells in peripheral blood resulted in the imbalance of TH1/TH2 indirectly.

Adolescent ; Antigens, CD1 ; analysis ; Cells, Cultured ; Child ; Child, Preschool ; Dendritic Cells ; immunology ; Female ; Humans ; Interferon-gamma ; blood ; Interleukin-12 ; biosynthesis ; blood ; Interleukin-4 ; blood ; Male ; Purpura, Schoenlein-Henoch ; immunology ; Th1 Cells ; immunology ; Th2 Cells ; immunology