IL-21 enhances dendritic cell ability to induce interferon-gamma production by natural killer T cells

Interleukin (IL)-21 shows pleiotropic effects on the proliferation, differentiation, and effector functions of leukocytes. However, the influence of IL-21 on dendritic cell (DC) activation of natural killer T (NKT) cells has not yet been elucidated. In the present study, we examined the effect of IL-21 on murine myeloid DC ability to induce NKT cell production of interferon-γ (IFN-γ) and IL-4. Pretreatment of DCs with IL-21 and α-galactosylceramide (α-GalCer), an NKT cell-specific ligand, resulted in the enhanced ability of the DCs to induce NKT cell production of IFN-γ but not IL-4 in vitro compared to DCs pretreated with α-GalCer alone. A similar effect of IL-21 was observed when DCs pretreated with IL-21 and α-GalCer in vitro were transferred into naïve mice. Direct administration of IL-21 to the mice also enhanced IFN-γ production after injection of α-GalCer. Thus, IL-21 can modify DC ability to selectively enhance NKT cell production of IFN-γ upon stimulation with α-GalCer.     

Growth ofChlamydia pneumoniaein cultured human peripheral blood mononuclear cells and induction of a cytokine response

In vitroinfection of freshly isolated human peripheral blood mononuclear cells (HPBMC) withChlamydia pneumoniaewas found to induce a production of tumour necrosis factorα(TNF-α), interleukin-1 β (IL-1β), interleukin-6 (IL-6) and interferonα(IFN-α). The secretion was dependent on the amount of infecting chlamydiae and most of it occurred during the first 12 to 24 h. Lipopolysaccharide (LPS) ofSalmonella minnesotaRechemotype, used as a positive control for HPBMC activation, induced a release of TNF-α, IL-1β and IL-6, but not of IFN-α, similar to the effect ofC. pneumoniae. Viable chlamydiae could not be recovered from HPBMCs infected immediately after their isolation, whereas HPBMCs which were culturedin vitrofor 3 to 9 days before infection were able to maintain the growth ofC. pneumoniae. Growth inside HPBMCs as well as induction of cytokine response may have a role in the pathogenesis ofC. pneumoniaeinfection.     

Induction of TNF-α mRNA in murine macrophages by virulent and avirulent strains ofSalmonella choleraesuisserovar Typhimurium and serovar Choleraesuis

TNF-αmRNA induction in murine macrophages by virulent and avirulentSalmonellastrains was measuredin vitroby RT-PCR method. Virulence plasmid-cured strains ofS. choleraesuisserovar Typhimurium and serovar Choleraesuis, andrpoS-defective mutant ofS. choleraesuisserovar Typhimurium induced significantly higher level of TNF-αmRNA than their parent (virulent) strains in macrophages of C3H/HeN mice. When macrophages of LPS-low responder (C3H/HeJ) mice were used, the difference of the induction level was not observed, indicating that LPS was involved in the enhanced level of TNF-αmRNA induction by avirulentSalmonellastrains. LPSs from virulent and avirulent strains were analysed, but no difference was found for cytokine-inducing activity, and chemical properties. Those results suggested that avirulentSalmonellastrains were damaged more easily, and released more LPS in macrophages to enhance TNF-αinduction.     

IMMUNOSTIMULATING EFFECTS OF ACIDIC POLYSACCHARIDES EXTRACT OF PANAX GINSENG ON MACROPHAGE FUNCTION

ABSTRACT

The root of Panax ginseng C. A. Meyer is one of the most popular natural tonics in oriental countries. In this study, we have isolated polysaccharide fraction of Panax ginseng (ginsan) and examined its effect on the function of murine peritoneal macrophages. When macrophages were treated with ginsan, cytotoxic activity against B16 melanoma cells was significantly induced. In addition, the levels of cytokines, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), IL-6 and lnterferon-γ (IFN-γ) were increased and the production of reactive oxygen/nitrogen components such as nitric oxide (NO) and hydrogen peroxide (H₂O₂) was enhanced. Moreover, phagocytic activity was induced in ginsan-treated macrophages compared to the control. The expression of CD14 and l-A^b on murine peritoneal macrophages was increased by the treatment with ginsan, while the expression of CD11b was decreased. Taken together, these results suggest that ginsan has an immunopotentiating effects on macrophages and these abilities could be used clinically for the treatment of diseases such as cancer.     

Production of soluble ICAM-1 from human endothelial cells induced by IL-1β and TNF-α

The present study was designed to establish the effects of cytokines on soluble ICAM-1 (sICAM-1) production by human endothelial cells (EC) and ICAM-1 expression on these cells and the effects of purified sICAM-1 on lympho-cyte-EC adhesion. Expression of ICAM-1 and production of sICAM-1 were measured by a specific ELISA method. ICAM-1 expression was enhanced by IL-1β, TNF-α, and most effectively by IFN-γ. IL-4, IL-6, M-CSF, or GM-CSF showed no effects on ICAM-1 expression. IL-4 (100 units/ml) or IL-6(100 units/ml) abolished the enhancing effect of IL-1β, while TNF-α (1, 10, 100 units/ml) synergized with IL-1β to promote ICAM-1 expression in EC. In contrast with the transient increase of cell-associated ICAM-1 expression after activiation by IL-1β, which peaked 40 h poststimulation and declined thereafter, sICAM-1 continued to accumulate in culture supernatants even after 48 h poststimulation in IL-1β-stimulated EC. IL-1β treatment resulted in an increase in adhesion. sICAM-1, purified from cell-free supernatants obtained after a 48-h culture of EC in IL-1β by affinity chromatography using monoclonal ICAM-1 antibody coupled to Sepharose beads, significantly inhibited lymphocyte EC adhesion. Preincubation of lymphocytes with conditioned medium of EC cultured with 100 units/ml IL-1β for 48 h, which contained a considerable amount of sICAM-1, resulted in a significant inhibition of lymphocyte adhesion to IL-1β-stimulated EC. These results suggest that there is a cumulative increase in sICAM-1 concentration in the vicinity of cytokine-stimulated EC and that this sICAM-1 modulates ICAM-1-mediated cell to cell interaction.     

Discriminators of mouse bladder response to intravesical Bacillus Calmette-Guerin (BCG)

Background

Intravesical Bacillus Calmette-Guerin (BCG) is an effective treatment for bladder superficial carcinoma and it is being tested in interstitial cystitis patients, but its precise mechanism of action remains poorly understood. It is not clear whether BCG induces the release of a unique set of cytokines apart from its pro-inflammatory effects. Therefore, we quantified bladder inflammatory responses and alterations in urinary cytokine protein induced by intravesical BCG and compared the results to non-specific pro-inflammatory stimuli (LPS and TNF-α). We went further to determine whether BCG treatment alters cytokine gene expression in the urinary bladder.

Methods

C57BL/6 female mice received four weekly instillations of BCG, LPS, or TNF-α. Morphometric analyses were conducted in bladders isolated from all groups and urine was collected for multiplex analysis of 18 cytokines. In addition, chromatin immune precipitation combined with real-time polymerase chain reaction assay (CHIP/Q-PCR) was used to test whether intravesical BCG would alter bladder cytokine gene expression.

Results

Acute BCG instillation induced edema which was progressively replaced by an inflammatory infiltrate, composed primarily of neutrophils, in response to weekly administrations. Our morphological analysis suggests that these polymorphonuclear neutrophils are of prime importance for the bladder responses to BCG. Overall, the inflammation induced by BCG was higher than LPS or TNF-α treatment but the major difference observed was the unique granuloma formation in response to BCG. Among the cytokines measured, this study highlighted the importance of IL-1β, IL-2, IL-3, IL-4, IL-6, IL-10, IL-17, GM-CSF, KC, and Rantes as discriminators between generalized inflammation and BCG-specific inflammatory responses. CHIP/Q-PCR indicates that acute BCG instillation induced an up-regulation of IL-17A, IL-17B, and IL-17RA, whereas chronic BCG induced IL-17B, IL-17RA, and IL-17RB.

Conclusion

To the best of our knowledge, the present work is the first to report that BCG induces an increase in the IL-17 family genes. In addition, BCG induces a unique type of persisting bladder inflammation different from TNF-α, LPS, and, most likely, other classical pro-inflammatorystimuli.     

Pathophysiological role of nitric oxide in rat experimental colitis

Overproduction of nitric oxide (NO) by inducible nitric oxide synthase (iNOS) may contribute to the pathophysiology of ulcerative colitis. A 2,4,6-trinitrobenzenesulfonic acid sodium salt (TNBS) colitis model was established to examine the effect of selective iNOS inhibition, by S-(2-aminoethyl) isothiouronium bromide (ITU), on colonic mucosal cell damage and inflammation. Rats, killed 7 days after TNBS, had increased colonic mucosal levels of iNOS and interleukin-8 (IL-8), in addition to severe colonic inflammation which was characterized by significantly increased colon weight, damage score and colonic myeloperoxidase activity (MPO) (a marker of neutrophil influx). TNBS-treated rats had markedly decreased body weight and thymus weight. Administration of colitic rats with ITU significantly inhibited iNOS activity/expression and tended to reduce mucosal levels of IL-8, but no effect on MPO activity was observed. Following ITU therapy, colitic rats had reduced colonic damage and losses in body weight and thymus weight were reversed. Improvement of TNBS colitis by ITU suggested that excess NO, produced by iNOS, may have contributed to the initiation/ amplification of colonic disease, by mechanisms including enhancement of IL-8 releaseNO-mediated enhancement of pro-inflammatory cytokine release was further investigated in vitro. Lipopolysaccharide (LPS) and interferon-γ (IFN-γ) stimulated release of nitrite, lactate dehydrogenase (LDH), TNFα, IL-1β and IL-8 from rat peritoneal macrophages, all of which were significantly reduced by ITU. This suggests that NO-mediated cell damage enhances pro-inflammatory mediator release from macrophages. In addition, enhancement of IL-8 and TNFα release was also partially NO-dependent in activated peritoneal neutrophils. Therefore, the amelioration of TNBS colitis by ITU could include inhibition of NO-mediated pro-inflammatory cytokine release.     

Balance of proinflammatory and antiinflammatory cytokines in mice immunized with Escherichia coli and correlation with mortality after lethal challenge

The balance of proinflammatory and antiinflammatory cytokines, their correlation with endotoxin levels and mortality rate after lethal challenge of Escherichia coli was investigated in mice immunized weekly for 8 weeks with formalin-killed E. coli either untreated or treated with 0.5×minimal inhibitory concentration of aztreonam. Control mice treated in parallel with saline, died within 24 h after challenge with 100× lethal dose (LD₅₀) of viable E. coli O6:K–. Mice immunized with antibiotic-treated bacteria showed a significantly higher survival than mice immunized with untreated E. coli. Cytokines were not detected in the sera of control mice during the entire period of immunization. At 90 min after immunization, mice immunized with antibiotic-treated E. coli showed tumor necrosis factor-α (TNF-α) levels significantly lower and interleukin (IL)-6 levels significantly higher (P<0.05) than mice immunized with untreated E. coli, while comparable levels of interferon-γ (IFN-γ) were measured in both groups. TNF-α and IL-10 levels measured 90 min after lethal challenge correlated with the mortality rate observed in each group (r=0.96 for TNF-α and 0.94 for IL-10). IL-6 levels correlated with survival (r = 0.95), while IFN-γ serum levels did not differ in the two immunized groups, but were significantly higher than those measured in the control mice. IL-4 was detected only after challenge of mice immunized with antibiotic-treated bacteria. Comparable levels of circulating endotoxin were measured after lethal challenge in both control and immunized mice. These data showed that in the presence of a protective immune response the survival of immunized mice was correlated with an early alteration of cytokine expression pattern including enhanced secretion of IL-4, IL-6 and IFN-γ, and reduced secretion of TNF-α and IL-10. Received: 5 January 1998     

泡球蚴原头蚴抗原和卡介苗对Th1/Th2转录因子和细胞因子的影响

目的探讨泡球蚴原头蚴抗原和卡介苗免疫小鼠对泡球蚴攻击感染的调节机制。方法用实时定量聚合酶链反应检测鼠脾组织中GATA-3及T-bet的mRNA表达水平;酶联免疫吸附法检测鼠血清中白介素4和γ干扰素的含量。结果卡介苗免疫攻击组与PBS对照组的转录因子(T-bet mRNA)和其标志性细胞因子(INF-γ)的表达量,差异有统计学意义(P<0.05)。结论实验证明卡介苗(BCG)有上调Th1型免疫反应的作用,用BCG可以干预或治疗由泡球蚴抗原诱导的晚期泡球蚴(AE)动物的免疫抑制状态。     

An essential role for endogenous interferon-gamma in the generation of protective T cells against Mycobacterium bovis BCG in mice.

Protective CD4+ T cells against Mycobacterium bovis bacillus Calmette-Guérin (BCG), which are characterized by the ability to produce interferon-gamma (IFN-gamma), could be induced by immunization of mice with viable BCG but not with killed BCG. A high level of IFN-gamma mRNA was observed when normal spleen cells were stimulated with viable but not killed BCG. By comparing mice immunized with either viable or killed M. bovis BCG, it was found that a high level of IFN-gamma mRNA was expressed only after immunization with viable BCG. This finding prompted us to investigate the effect of neutralizing the IFN-gamma on the final generation of protective T cells against M. bovis BCG. When endogenous IFN-gamma was neutralized by administration of anti-IFN-gamma monoclonal antibody in mice immunized with viable BCG, the generation of protective T cells was significantly impaired, as revealed by the adoptive transfer of spleen T cells. The generation of BCG-specific, IFN-gamma-producing T cells was also abolished. These results clearly demonstrate that endogenous IFN-gamma actually plays a critical role in the generation of protective T cells against M. bovis BCG in vivo. Moreover, this study suggests that the lack of IFN-gamma-inducing ability is responsible for the inability of killed M. bovis BCG to induce protective T cells in mice.     

IL-15 complexes induce NK- and T-cell responses independent of type I IFN signaling during rhinovirus infection

Rhinoviruses are among the most common viruses to infect man, causing a range of serious respiratory diseases including exacerbations of asthma and COPD. Type I IFN and IL-15 are thought to be required for antiviral immunity; however, their function during rhinovirus infection in vivo is undefined. In RV-infected human volunteers, IL-15 protein expression in fluid from the nasal mucosa and in bronchial biopsies was increased. In mice, RV induced type I IFN-dependent expressions of IL-15 and IL-15Rα, which in turn were required for NK- and CD8⁺ T-cell responses. Treatment with IL-15–IL-15Rα complexes (IL-15c) boosted RV-induced expression of IL-15, IL-15Rα, IFN-γ, CXCL9, and CXCL10 followed by recruitment of activated, IFN-γ-expressing NK, CD8⁺, and CD4⁺ T cells. Treating infected IFNAR1^−/− mice with IL-15c similarly increased IL-15, IL-15Rα, IFN-γ, and CXCL9 (but not CXCL10) expression also followed by NK-, CD8⁺-, and CD4⁺-T-cell recruitment and activation. We have demonstrated that type I IFN-induced IFN-γ and cellular immunity to RV was mediated by IL-15 and IL-15Rα. Importantly, we also show that IL-15 could be induced via a type I IFN-independent mechanism by IL-15 complex treatment, which in turn was sufficient to drive IFN-γ expression and lymphocyte responses.     

Listeria monocytogenes (delta-actA mutant) infection in tumor necrosis factor receptor p55-deficient neonatal mice

Using TNF receptor 1 knock out (TNFR1KO) mice, we investigated the role played by TNFR1 in immune regulation during neonatal listeriosis. Induction of protective immune response in wild type pups resulted in the prompt control of infection with an attenuated ?actA mutant Listeria monocytogenes, accompanied by enhanced hepatic expression of mRNA for IFN-γ, TNF-α, and IL-10. Conversely, the lack of TNFR1 signalling in TNFR1KO neonatal mice resulted in substantial changes in the profile of inflammatory mediators and ultimately fatal outcome of the infected pups. Despite remarkable increase in indoleamine 2, 3-dioxygenase (IDO) and inducible nitric oxide synthase (iNOS) mRNA detected in the liver of TNFR1KO mice, bacterial proliferation was unrestrained. Increased mRNA expression of IDO, iNOS, TNF-α, IFN-γ, MCP-1, and MIP-1α was found in the spleens of infected KO mice, and in the brains mRNA encoding iNOS, IDO, IFN-γ, IL-12p40, IL-10, and RANTES was also upregulated. Large necrotic lesions consisting of granulocytes and macrophages were scattered throughout the liver of these mice. TNFR1KO neonates were unable to clear neutrophils and switch from the innate immune response to a specific reaction mediated by T cells. These results prove that TNF-α signalling is crucial and irreplaceable in antilisterial protection during the neonatal period.     

Recombinant BCG-Rv1767 amount determines, in vivo, antigen-specific T cells location,frequency, and protective outcome

One possibility to improve the efficacy of BCG vaccine against TB is to create a recombinant BCG (r-BCG), increasing the expression of mycobacterial antigens, to ameliorate the response to BCG. Here we describe a new r-BCG expressing the gene Rv1767, induced by Mycobacterium tuberculosis during its survival in human macrophages. The r-BCG elicited a specific T cells response in Balb/c mice higher than wt BCG. The r-BCG amount used to immunise mice determined diverse Th1/Th2 equilibriums, which was not the same in spleen and Lymph Nodes. Differences in cytokines production were found for IL-10, IL-4, TNF-α, and Arginase-1, which, in some conditions, resulted higher in r-BCG as compared to wt BCG-immunised mice.     

Distinct expression of interleukin 17, tumor necrosis factor α, transforming growth factor β, and forkhead box P3 in acute rejection after kidney transplantation

The kidney transplant is the main therapeutic alternative for end-stage kidney disease, and rejection is a major complication. The expression of proinflammatory cytokines is related to graft loss, whereas anti-inflammatory cytokines are associated with graft protection. The objective of this study is to evaluate the “in situ” expression of cytokines T helper 1 (tumor necrosis factor α [TNF-α]), T helper 17 (interleukin 17 [IL-17]), and regulatory T cell (transforming growth factor β [TGF-β]) and the expression of forkhead box P3 (FoxP3) in allograft kidney. We evaluated in situ expression of cytokines in allograft kidney under rejection process by indirect immunohistochemistry. Eighteen renal graft biopsies were from patients with episodes of rejection. The in situ expression of IL-17, TNF-α, and TGF-β was significantly higher in patients with acute rejection when compared with the control group. In contrast, analysis of FoxP3 expression showed few positive cells in patients with acute rejection compared with the control group. The results suggest that the expression of proinflammatory cytokines (IL-17 and TNF-α) contributes to the mechanisms of kidney transplant rejection. The increase in TGF-β expression might be an attempt to establish a process of immunoregulation or even to induce higher production of IL-17. The last hypothesis is supported by the observation of a reduced expression of FoxP3 and elevated levels of IL-17.