Cytokines ; physiology ; Humans ; Interferon-gamma ; physiology ; Interleukin-1 ; physiology ; Interleukin-10 ; physiology ; Interleukin-12 ; physiology ; Interleukin-6 ; physiology ; Interleukin-8 ; physiology ; Rotavirus Infections ; immunology ; Tumor Necrosis Factor-alpha ; physiology

OBJECTIVEPrevious studies have shown that bacillus calmette-guerin (BCG) can deviate TH2 response toward TH1 response, resulting in a suppressive effect on the development of asthma/atopy. This study examined the effect of BCG treatment on regulatory T cells in asthmatic mice to investigate the possible mechanism.

METHODSKunming mice were sensitized and challenged with ovalbumin (OVA) to establish asthmatic models. Asthmatic mice were injected intradermally with BCG five days before and after sensitization. After 24 hrs of last challenge, bronchoaveolar lavage fluid (BALF) and peripheral blood were collected . The total cells and eosinophils were counted in the BALF. The percentage of CD4(+) CD25(+) in peripheral blood was detected with flow cytometry. Single spleen cell suspension was prepared and cultured in 1640 medium for 48 hrs and then the cytokine IL-10 level in the supernatant was determined using ELISA. The mice which were challenged with normal saline were used as the Normal control group.

RESULTSThe number of total cells and eosinophils in BALF in asthmatic mice [(27.27 +/- 5.36) x 10(7)/L and (6.59 +/- 1.32) x 10(7)/L respectively] were more than in the Normal control group [(1.52 +/- 0.36) x 10(7)/L and zero respectively] (P < 0.01). The number of total cells and eosinophils in BALF in asthmatic mice were reduced after BCG treatment [(13.71 +/- 3.17) x 10(7)/L and (1.43 +/- 0.37) x 10(7)/L respectively] (P < 0.01). The percentage of CD4(+) CD25(+) in peripheral blood of asthmatic mice [(11.59 +/- 1.33)%] was noticeably lower than that of the Control group [(13.66 +/- 1.68)%] (P < 0.01), but increased significantly in asthmatic mice after BCG treatment [(14.40 +/- 2.70)%] (P < 0.05). The IL-10 level in spleen cell supernatant in the BCG-treated group (7.79 +/- 1.34 pg/mL) also increased compared with that in the untreated asthmatic mice (5.54 +/- 0.66 pg/mL) (P < 0.01).

CONCLUSIONSBCG can markedly inhibit the airway inflammation in asthmatic mice possibly by promoting the production of regulatory T cells.

Animals ; Asthma ; immunology ; therapy ; BCG Vaccine ; therapeutic use ; Bronchoalveolar Lavage Fluid ; cytology ; Interleukin-10 ; analysis ; physiology ; Male ; Mice ; T-Lymphocytes, Regulatory ; immunology ; Toll-Like Receptor 2 ; physiology

Cell Survival ; drug effects ; Cells, Cultured ; Dendritic Cells ; drug effects ; immunology ; physiology ; Humans ; Immunophenotyping ; Interleukin-10 ; pharmacology ; Interleukin-12 ; genetics ; secretion ; Interleukin-12 Subunit p35 ; Interleukin-12 Subunit p40 ; Protein Subunits ; genetics ; secretion ; RNA, Messenger ; analysis

BACKGROUNDExperimental autoimmune myocarditis (EAM) in rats is a T-cell-mediated disorder. The initiation and maintenance of autoimmune responses in EAM depend on the maturation state of dendritic cells. IL-10 is a pleiotrophic immunomodulatory cytokine that functions at different levels of the immune response, so it has emerged as a promising therapeutic factor for the treatment of autoimmune/inflammatory diseases. This study was designed to test the hypothesis that IL-10 gene modified bone marrow-derived immature dendritic cells (iDCs) ameliorate EAM and to explore the underlying mechanisms.

METHODSEAM was induced using the methods of cardiac myosin immunization on day 0 and day 7. Immature and mature bone marrow-derived dendritic cells (BMDCs) were generated without or with the stimulation by lipopolysaccharide (LPS) and the phenotype was analyzed by flow cytometry. Some of the iDCs were transfected by pcDNA3-IL-10 plasmid. 2 x 10(6)/per rat mature DC (mDC), immature DC (iDC), pcDNA3 transfected iDC, pcDNA3-IL-10 transfected iDC or phosphate buffered saline (PBS) were injected intravenously for treatment 5 days after the first immunization. On day 21, HE staining was performed to detect the myocardial inflammation and T lymphocyte proliferation assay was used to determine the effects of IL-10 gene transfected iDC on autoreactive T cell proliferation. Expression of IkappaB, the inhibitor of NF-kappaB pathway, was determined by Western blot.

RESULTSBMDCs generated in a medium supplemented with granulocyte-macrophage-colony-stimulating factor (GM-CSF) were relatively immature, as determined by flow cytometry. However, stimulation with LPS induced these cells to become mature (m) DCs with higher levels of surface major histocompatibility complex (MHC)-II and costimulatory molecules. Intravenous administration of iDCs, especially pcDNA3-IL-10 transfected iDC, ameliorated the histopathological severity of the myosin induced-EAM, and the effect was lost after the DCs underwent maturation induced by in vitro exposure to LPS. IL-10 gene modified iDC inhibited the antigen specific T cell responses towards cardiac myosin. IkappaB protein was up-regulated significantly in the IL-10 gene modified iDC group.

CONCLUSIONSIL-10 gene modified iDC induced antigen-specific tolerance in EAM. The underlying mechanisms may be related to costimulatory molecules down-regulation and NF-kappaB pathway inhibition.

Animals ; Autoimmune Diseases ; immunology ; Dendritic Cells ; physiology ; Immune Tolerance ; Interleukin-10 ; genetics ; Lymphocyte Activation ; Male ; Myocarditis ; immunology ; Myosins ; immunology ; NF-kappa B ; physiology ; Rats ; Rats, Inbred Lew ; Signal Transduction ; Transfection

OBJECTIVETo investigate the effect and its mechanism of reducing graft-versus-host disease (GVHD) by in vitro blockade of CD(40)-CD(40)L pathway in vitro, the donor T lymphocytes cultured in vitro with anti-CD(40)L mAb were transfused in bone marrow transplantation (BMT) GVHD mouse model.

METHODSC57BL/6(H-2b) spleen T cells were isolated as responder cells, and BALB/c(H-2d) spleen cells as stimulator cells. They were cocultured with or without Anti-CD(40)L mAb as anti-CD(40)L mAb group and control group, respectively. At day 5, the mixed lymphocyte response (MLR)-culture cells mixed with bone marrow cells and transfused respectively into the TBI conditioned recipient mice. The mice were divided into two groups: group A, bone marrow cells (2 x 10(6)) and spleen T lymphocytes (2 x 10(6)) from MLR control group; group B, bone marrow cells (2 x 10(6)) and spleen T lymphocytes (2 x 10(6)) from MLR anti-CD(40)L mAb group. The GVHD incidence and hematopoietic reconstitution were observed. Peripheral blood sera and spleen cells of the recipients mice were harvested at scheduled time points for the measurement of cytokines and T cell immunophenotyping with flow cytometry.

RESULTSThe incidence of GVHD in group A was 100% (10/10), and in group B was 20% (2/10). The percentage of H-2D(b) positive cells in group B (n = 8) was (93.54 +/- 2.32)% at day 40 after transplantation. The levels of cytokines in serum from group B were significantly lower than those from group A (P < 0.05). The expressions of CD(4)(+), CD(8)(+), CD(4)(+)CD(25)(+), CD(8)(+)CD(25)(+), CD(4)(+)CD(69)(+), CD(8)(+)CD(69)(+) and CD(4)(+)CD(40)L(+) were lower in group B than in group A (P < 0.05). The expressions of CD(8)(+)CD(40)L(+) and CD(4)(+)CD(45)RA(+) were similar in the two groups (P > 0.05).

CONCLUSIONBlockade of CD(40)-CD(40)L interaction in vitro could induce immune tolerance in vivo, reduce aGVHD in aGVHD mice model and form chimerism, which was mediated by inhibiting the Th1 and Th2 cytokines production, inducing tolerance of CD(4)(+) and CD(8)(+) cells to alloantigens. The obstruction of T cells activation after tolerance happened mainly at the early and mature phase of T cells activation. These provided the experimental basis for the use of anti-CD(40)L mAb in the clinical transplantation to prevent aGVHD.

Animals ; Antibodies, Monoclonal ; therapeutic use ; Bone Marrow Transplantation ; adverse effects ; CD40 Antigens ; physiology ; CD40 Ligand ; immunology ; physiology ; Graft vs Host Disease ; prevention & control ; Interleukin-10 ; blood ; Interleukin-4 ; blood ; Male ; Mice ; Mice, Inbred C57BL

BACKGROUNDBAFF, the B cell activation factor, is a member of the tumor necrosis factor (TNF) ligand family that binds to BCMA, TACI, and BAFF-R. Previous studies have shown that members of the TNF family are detected in human placental trophoblast cells, but the expression patterns of BAFF involved in human decidua and the differential expression of BAFF between normal pregnancy and miscarriage are still incompletely documented or unknown. This study was designed to investigate the expression of BAFF and BAFF-R in the trophoblast and decidua of normal early pregnant women and recurrent spontaneous abortion (RSA) patients.

METHODSForty-five patients with RSA and 45 normal pregnant women were included in this study. By reverse transcriptase-polymerase chain reaction (RT-PCR), Western blotting and immunohistochemical experiments, we explored the expression of BAFF and BAFF-R in the maternal-fetal interface of normal early pregnant women and RSA patients.

RESULTSAnalysis by RT-PCR and Western blotting revealed that BAFF was detected in both trophoblast and decidua of all the samples, and the expression level was higher in the tissues of normal early pregnant women (P<0.05) than that of recurrent spontaneous abortion patients under the same gestational weeks. Messages for BAFF-R were absent. Immunohistochemical experiments showed that expression of BAFF was cell-specific which was localized to villous cytotrophoblast and syncytiotrophoblast cells in trophoblast and to stromal cells in decidua. Whereas BAFF was prominent on the trophoblast and decidua of normal early pregnant women, it was decreased in the tissues of RSA patients.

CONCLUSIONSBAFF might steer maternal leukocytes away from a harmful immune response and toward a favorable one and play a potentially vital role for successful pregnancy.

Abortion, Habitual ; metabolism ; B-Cell Activating Factor ; analysis ; genetics ; physiology ; Decidua ; chemistry ; metabolism ; Female ; Humans ; Immunohistochemistry ; Interleukin-10 ; genetics ; Pregnancy ; RNA, Messenger ; analysis ; Th1 Cells ; immunology ; Th2 Cells ; immunology ; Trophoblasts ; chemistry ; metabolism

BACKGROUNDMast cells are implicated in the development of irritable bowel syndrome (IBS), which is associated with the activation of the "neural-immune" system. The aim of this study was to investigate the role of mast cells in the remodeling of cholinergic and peptidergic neurotransmitters induced by acute cold restriction stress (ACRS) post infection (PI) using mast cell deficient rats (Ws/Ws) and their wild-type controls (+/+).

METHODSTransient intestinal infection was initiated by giving 1500 Trichinella spiralis (T.S.) larvae by gavage. ACRS was induced for 2 hours at day 100 PI. Samples of terminal ilea were prepared for H&E staining, mast cell counting and activation and assessment of IL-1beta and IL-10.

RESULTSWhen infected, both strains of rats experienced an acute infectious stage followed by a recovery. Histological scores were significantly higher in infected rats compared with those of the non-infected controls at day 10 PI (10 day-PI vs. control: +/+: 2.75+/-0.17 vs. 0.42+/-0.09; Ws/Ws: 2.67+/-0.67 vs. 0.50+/-0.34; P<0.01). In +/+ rats, post-infection ACRS induced the formation of low-grade inflammation, represented by the imbalance of IL-1beta and IL-10 (IL-1beta: PI+ACRS vs. control: (1812.24+/-561.61) vs. (1275.97+/-410.21) pg/g, P<0.05; IL-10: PI+ACRS vs. control: (251.9+/-39.8) vs. (255.3+/-24.7) pg/g, P>0.05), accompanied by hyperplasia and activation of mast cells (PI+ACRS vs. control: 58.8+/-19.2 vs. 28.0+/-7.6; P<0.01). The balance between acetylcholine (ACh) and substance P (SP) was also disturbed (ACh: PI+ACRS vs. control: (743.94+/-238.72) vs. (1065.68+/-256.46) pg/g, P<0.05; SP: PI+ACRS vs. control: (892.60+/-231.12) vs. (696.61+/-148.61) pg/g, P<0.05). Nevertheless, similar changes of IL-1beta/IL-10 and ACh/SP were not detected in Ws/Ws rats.

CONCLUSIONThe imbalance of ACh/SP, together with the activation of mucosal immunity induced by post-infection ACRS were lacking in mast cell deficient rats, which supports the premise that mast cells play an important role in cholinergic and peptidergic remodeling in the ileum of rats.

Acetylcholine ; metabolism ; Animals ; Enzyme-Linked Immunosorbent Assay ; Ileum ; metabolism ; Interleukin-10 ; metabolism ; Interleukin-1beta ; metabolism ; Intestines ; immunology ; metabolism ; parasitology ; Male ; Mast Cells ; cytology ; metabolism ; physiology ; ultrastructure ; Microscopy, Electron, Transmission ; Neurotransmitter Agents ; metabolism ; Radioimmunoassay ; Rats ; Substance P ; metabolism ; Trichinella spiralis ; physiology ; Trichinellosis ; immunology

PURPOSE: The function of regulatory B lymphocytes is known to be abnormal in inflammatory diseases. However, a recent study indicates that IL-10+ B cells seem to be expanded in rheumatoid arthritis (RA). Therefore, the state of IL-10+ B cells in the peripheral blood from RA patients and healthy controls were investigated. MATERIALS AND METHODS: CD19+ cells in peripheral blood mononuclear cells were purified from blood samples of RA patients and age and gender-matched healthy controls, and stimulated with CD40 ligand and CpG for 48 hours. Then, intracellular IL-10 in CD19+ cells was analyzed using flow cytometry. RESULTS: There was no significant difference in the proportion of IL-10+ B cells between 10 RA patients and 10 healthy controls (RA, 0.300+/-0.07 vs. healthy control 0.459+/-0.07, p=0.114). The proportion of induced IL-10+ B cells to total B cells in RA patients was significantly higher than those in controls (RA, 4.44+/-3.44% vs. healthy control 2.44+/-1.64%, p=0.033). However, the proportion of IL-10+ B cells to total B cells correlated negatively with disease activity in RA patients (r=-0.398, p=0.040). Erythrocyte sedimentation rate or C-reactive protein or medication was not associated with the proportion of IL-10+ B cells. CONCLUSION: The proportion of induced IL-10+ B cell increased in RA patients compared to healthy control, however, negatively correlated with disease activity in RA.
Adult ; Aged ; Antigens, CD19/metabolism ; Arthritis, Rheumatoid/blood/*immunology/pathology ; B-Lymphocytes, Regulatory/metabolism/*physiology ; Biological Markers/blood ; Female ; Humans ; Interleukin-10/metabolism ; Male ; Middle Aged ; Severity of Illness Index

Genetic Predisposition to Disease ; genetics ; Genotype ; HLA Antigens ; genetics ; Hepatitis B virus ; genetics ; physiology ; Hepatitis B, Chronic ; genetics ; immunology ; virology ; Humans ; Interleukin-10 ; genetics ; Mutation ; Polymorphism, Single Nucleotide ; Promoter Regions, Genetic ; genetics ; Tumor Necrosis Factor-alpha ; genetics

A degree of brain inflammation is required for repair of damaged tissue, but excessive inflammation causes neuronal cell death. Here, we observe that IL-10 is expressed in LPS-injected rat cerebral cortex, contributing to neuronal survival. Cells immunopositive for IL-10 were detected as early as 8 h post-injection and persisted for up to 3 d, in parallel with the expression of IL-1beta, TNF-alpha, and iNOS. Double immunofluorescence staining showed that IL-10 expression was localized mainly in activated microglia. Next, we examined the neuroprotective effects of IL-10 using IL-10 neutralizing antibody (IL-10NA). Blockade of IL-10 action caused a significant loss of neurons both 3 d and 7 d after LPS injection. Further, the induction of mRNA species encoding IL-1beta, TNF-alpha, and iNOS, reactive oxygen species (ROS) production, and NADPH oxidase activation, increased after co-injection of LPS and IL-10NA, compared to the levels seen after injection of LPS alone. Taken together, these results clearly suggest that LPS-induced endogenous expression of IL-10 in microglia contributes to neuronal survival by inhibiting brain inflammation.
Animals ; Cerebral Cortex/drug effects/*pathology ; Fluorescent Antibody Technique ; Interleukin-10/immunology/*physiology ; Lipopolysaccharides/*pharmacology ; Microglia/cytology/*metabolism ; Nerve Degeneration/pathology/*prevention & control ; Neurons/cytology/drug effects/*metabolism ; Nitric Oxide Synthase/genetics/metabolism ; Rats ; Rats, Sprague-Dawley ; Reactive Oxygen Species/metabolism