DC-SIGN (CD209) gene promoter polymorphisms in a Brazilian population and their association with human T-cell lymphotropic virus type 1 infection

This study evaluated four polymorphisms located in the DC-SIGN (CD209) gene promoter region (positions -336, -332 -201 and -139) in DNA samples from four Brazilian ethnic groups (Caucasians, Afro-Brazilian, Asians and Amerindians) to establish the population distribution of these single-nucleotide polymorphisms (SNPs) and correlated DC-SIGN polymorphisms and infection in samples from human T-cell lymphotropic virus type 1 (HTLV-1)-infected individuals. To identify CD209 SNPs, 452 bp of the CD209 promoter region were sequenced and the genotype and allelic frequencies were evaluated. This is the first study to show genetic polymorphism in the CD209 gene in distinct Brazilian ethnic groups with the distribution of allelic and genotypic frequency. The results showed that -336A and -139A SNPs were quite common in Asians and that the -201T allele was not observed in Caucasians, Asians or Amerindians. No significant differences were observed between individuals with HTLV-1 disease and asymptomatic patients. However, the -336A variant was more frequent in HTLV-1 -infected patients [HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), 80%; healthy asymptomatic HTLV-1 carriers, 90 %] than in the control group (70 %) [P=0.0197, odds ratio (OR)=2.511, 95 % confidence interval (CI)=1.218-5.179). In addition, the -139A allele was found to be associated with protection against HTLV-1 infection (P=0.0037, OR=0.3758, 95% CI=0.1954-0.7229) when the HTLV-1 -infected patients as a whole were compared with the healthy-control group. These observations suggest that the -139A allele may be associated with HTLV-1 infection, although no significant association was observed among asymptomatic and HAM/TSP patients. In conclusion, the variation observed in SNPs -336 and -139 indicates that this lectin may be of crucial importance in the susceptibility/transmission of HTLV-1 infections.

HMGB1-derived peptide acts as adjuvant inducing immune responses to peptide and protein antigen

There is a need for new adjuvants that will induce immune responses to subunit vaccines. We show that a short peptide, named Hp91, whose sequence corresponds to an area within the endogenous molecule high mobility group box (HMGB1) protein 1 potentiates cellular immune responses to peptide antigen and cellular and humoral immune responses to protein antigen in vivo. Hp91 promoted the in vivo production of the immunomodulatory cytokines, IFN-gamma, TNF-alpha, IL-6, and IL-12 (p70), as well as antigen-specific activation of CD8+ T cells. These results demonstrate the ability of a short immunostimulatory peptide to serve as an adjuvant for subunit vaccines. (C) 2010 Elsevier Ltd. All rights reserved.

Prostaglandin mediates IL-23/IL-17-induced neutrophil migration in inflammation by inhibiting IL-12 and IFN gamma production

IL-23/IL-17-induced neutrophil recruitment plays a pivotal role in rheumatoid arthritis (RA). However, the mechanism of the neutrophil recruitment is obscure. Here we report that prostaglandin enhances the IL-23/IL-17-induced neutrophil migration in a murine model of RA by inhibiting IL-12 and IFN gamma production. Methylated BSA (mBSA) and IL-23-induced neutrophil migration was inhibited by anti-IL-23 and anti-IL-17 antibodies, COX inhibitors, IL-12, or IFN gamma but was enhanced by prostaglandin E(2) (PGE(2)). IL-23-induced IL-17 production was increased by PGE(2) and suppressed by COX-inhibition or IL-12. Furthermore, COX inhibition failed to reduce IL-23-induced neutrophil migration in IL-12- or IFN gamma-deficient mice. IL-17-induced neutrophil migration was not affected by COX inhibitors, IL-12, or IFN gamma but was inhibited by MK886 (a leukotriene synthesis inhibitor), anti-TNF alpha, anti-CXCL1, and anti-CXCL5 antibodies and by repertaxin (a CXCR1/2 antagonist). These treatments all inhibited mBSA- or IL-23-induced neutrophil migration. IL-17 induced neutrophil chemotaxis through a CXC chemokines-dependent pathway. Our results suggest that prostaglandin plays an important role in IL-23-induced neutrophil migration in arthritis by enhancing IL-17 synthesis and by inhibiting IL-12 and IFN gamma production. We thus provide a mechanism for the pathogenic role of the IL-23/IL-17 axis in RA and also suggest an additional mechanism of action for nonsteroidal anti-inflammatory drugs.

IL-4 regulates susceptibility to intestinal inflammation in murine food allergy

Cardoso CR, Provinciatto PR, Godoi DF, Ferreira BR, Teixeira G, Rossi MA, Cunha FQ, Silva JS. IL-4 regulates susceptibility to intestinal inflammation in murine food allergy. Am J Physiol Gastrointest Liver Physiol 296: G593-G600, 2009. First published January 8, 2009; doi:10.1152/ajpgi.90431.2008.-Allergies involve a state of immediate hypersensitivity to antigens, including food proteins. The mechanism underlying the initiation and development of allergic responses involves IL-4 that directly induces the differentiation of committed effector Th2 lymphocytes. Although it is clear that Th2 responses play a pivotal role in the development of allergic responses, it remains unclear which mechanisms are involved in the development of the intestinal damages observed in food allergy. Accordingly, this work aimed to study the role of Th2/IL-4-dependent responses in the development of food allergy and intestinal pathology. C57BL/6 wild-type (WT) and IL-4(-/-) mice were sensitized with peanut proteins, challenged with peanut seeds, and followed for the development of food allergy and intestinal inflammation. Results demonstrated that exposure to peanut seeds led to weight loss in WT but not in IL-4(-/-) mice that preserved gut integrity with no signs of mucosal inflammation. These animals presented increased levels of IgG2a in sera, suggesting a role for allergic antibodies in the pathogenesis of WT animals. Most importantly, results also showed that lack of IL-4 modulated gut mucosal response in food allergy through diminished expression of TNF-alpha mRNA, increased Th1 IFN-gamma, IL-12p40, regulatory cytokines, and Foxp3, demonstrating their relevance in the control of allergic inflammatory processes, especially in the intestine. Finally, this study highlighted some of the complex mechanisms involved in the pathogenesis of allergic responses to food antigens in the gut, thereby providing valuable tools for directing novel therapeutic or preventive strategies to the control of allergic enteropathy.

Phlebotomine salivas inhibit immune inflammation-induced neutrophil migration via an autocrine DC-derived PGE(2)/IL-10 sequential pathway

In the present study, we investigated whether saliva from Phlebotomus papatasi and Phlebotomus duboscqi inhibited antigen-induced neutrophil migration and the mechanisms involved in these effects. The pretreatment of immunized mice with salivary gland extracts (SGE) of both phlebotomines inhibited OVA challenge-induced neutrophil migration and release of the neutrophil chemotactic mediators, MIP-1 alpha, TNF-alpha, and leukotriene B-4 (LTB4). Furthermore, SGE treatment enhanced the production of anti-inflammatory mediators, IL-10 and PGE(2). SGE treatments failed to inhibit neutrophil migration and MIP-1 alpha and LTB4 production in IL-10(-/-) mice, also failing in mice treated with nonselective (indomethacin) or selective (rofecoxibe) cyclooxygenase (COX) inhibitors. COX inhibition resulted in diminished SGE-induced IL-10 production, and PGE(2) release triggered by SGE remained increased in IL-10(-/-) mice, suggesting that prostanoids are acting through an IL-10-dependent mechanism. SGE treatments in vivo reduced the OVA-induced lymphoproliferation of spleen-derived cells. Further, the in vitro incubation of bone marrow-derived dendritic cells (DC) with SGE inhibited the proliferation of CD4(+) T cells from OVA-immunized mice, which was reversed by indomethacin and anti-IL-10 antibody treatments. Supporting these results, SGE induced the production of PGE(2) and IL-10 by DC, which were blocked by COX inhibition. These effects were associated with the reduction of DC-membrane expression of MHC-II and CD86 by SGE treatment. Altogether, the results showed that Phlebotomine saliva inhibits immune inflammation-induced neutrophil migration by an autocrine DC sequential production of PGE(2)/IL-10, suggesting that the saliva constituents might be promising therapeutic molecules to target immune inflammatory diseases.

NO EVIDENCE OF PATHOLOGICAL AUTOIMMUNITY FOLLOWING MYCOBACTERIUM LEPRAE HEAT-SHOCK PROTEIN 65-DNA VACCINATION IN MICE

Heat-shock proteins (HSPs) are currently one of the most promising targets for the development of immunotherapy against tumours and autoimmune disorders. This protein family has the capacity to activate or modulate the function of different immune system cells. They induce the activation of monocytes, macrophages and dendritic cells, and contribute to cross-priming, an important mechanism of presentation of exogenous antigen in the context of MHC class I molecules, These various immunological properties of HSP have encouraged their use in several clinical trials. Nevertheless, an important issue regarding these proteins is whether the high homology among HSPs across different species may trigger the breakdown of immune tolerance and induce autoimmune diseases. We have developed a DNA vaccine codifying the Mycobacterium leprae Hsp65 (DNAhsp65), which showed to be highly immunogenic and protective against experimental tuberculosis. Here, we address the question of whether DNAhsp65 immunization could induce pathological autoimmunity in mice. Our results show that DNAhsp65 vaccination induced antibodies that can recognize the human Hsp60 but did not induce harmful effects in 16 different organs analysed by histopathology up to 210 days after vaccination. We also showed that anti-DNA antibodies were not elicited after DNA vaccination. The results are important for the development of both HSP and DNA-based immunomodulatory agents.

Cytokine Signatures of Innate and Adaptive Immunity in 17DD Yellow Fever Vaccinated Children and Its Association With the Level of Neutralizing Antibody

Background. The live attenuated yellow fever (YF) vaccines have been available for decades and are considered highly effective and one of the safest vaccines worldwide. Methods. The impact of YF-17DD-antigens recall on cytokine profiles of YF-17DD-vaccinated children were characterized using short-term cultures of whole blood samples and single-cell flow cytometry. This study enrolled seroconverters and nonseroconverters after primovaccination (PV-PRNT(+) and PV-PRNT(-)), seroconverters after revaccination (RV-PRNT(+)), and unvaccinated volunteers (UV-PRNT(-)). Results. The analysis demonstrated in the PV-PRNT(+) group a balanced involvement of pro-inflammatory/regulatory adaptive immunity with a prominent participation of innate immunity pro-inflammatory events (IL-12(+) and TNF-alpha(+) NEU and MON). Using the PV-PRNT(+) cytokine signature as a reference profile, PV-PRNT(+) presented a striking lack of innate immunity proinflammatory response along with an increased adaptive regulatory profile (IL-4(+) CD4(+) T cells and IL-10(+) and IL-5(+) CD8(+) T cells). Conversely, the RV-PRNT(+) shifted the overall cytokine signatures toward an innate immunity pro-inflammatory profile and restored the adaptive regulatory response. Conclusions. The data demonstrated that the overall cytokine signature was associated with the levels of PRNT antibodies with a balanced innate/adaptive immunity with proinflammatory/regulatory profile as the hallmark of PV-PRNT(MEDIUM+), whereas a polarized regulatory response was observed in PV-PRNT(-) and a prominent proinflammatory signature was the characteristic of PV-PRNT(HIGH+).

Recognition by toll-like receptor 2 induces antigen-presenting cell activation and Th1 programming during infection by Neospora caninum

Neospora caninum is an apicomplexan parasite responsible for major economic losses due to abortions in cattle. Toll-like receptors (TLRs) sense specific microbial products and direct downstream signaling pathways in immune cells, linking innate, and adaptive immunity. Here, we analyze the role of TLR2 on innate and adaptive immune responses during N. caninum infection. Inflammatory peritoneal macrophages and bone marrow-derived dendritic cells exposed to N. caninum-soluble antigens presented an upregulated expression of TLR2. Increased receptor expression was correlated to TLR2/MyD88-dependent antigen-presenting cell maturation and pro-inflammatory cytokine production after stimulation by antigens. Impaired innate responses observed after infection of mice genetically deficient for TLR2((-/-)) was followed by downregulation of adaptive T helper 1 (Th1) immunity, represented by diminished parasite-specific CD4(+) and CD8(+) T-cell proliferation, IFN-gamma:interleukin (IL)-10 ratio, and IgG subclass synthesis. In parallel, TLR2(-/-) mice presented higher parasite burden than wild-type (WT) mice at acute and chronic stages of infection. These results show that initial recognition of N. caninum by TLR2 participates in the generation of effector immune responses against N. caninum and imply that the receptor may be a target for future prophylactic strategies against neosporosis. Immunology and Cell Biology (2010) 88, 825-833; doi:10.1038/icb.2010.52; published online 20 April 2010

Cytokines in periodontal disease: where to from here?

Numerous studies have attempted to elucidate the cytokine networks involved in chronic periodontitis, often with conflicting results. A variety of techniques were used to study cells in situ, cells extracted from gingival tissues, peripheral blood mononuclear cells, purified cell populations, and T cell lines and clones. Bacterial components, including sonicates, killed cells, outer membrane components, and purified antigens, have all been used to stimulate cells in vitro, making comparisons of cytokine profiles difficult. As it is likely that different cells are present at different disease stages, the inability to determine disease activity clinically is a major limitation of all these studies. In the context of tissue destruction, cytokines such as IL-1, IL-6 and IL-18 are likely to be important, as are their regulating cytokines IL-10 and IL-11. In terms of the nature of the inflammatory infiltrate, two apparently conflicting hypotheses have emerged: one based on direct observations of human lesions, the other based on animal experimentation and the inability to demonstrate IL-4 mRNA in gingival extracts. In the first of these, Th1 responses are responsible for the stable lesion, while in the second Th2 responses are considered protective. Using Porphyromonas gingivalis specific T cell lines we have shown a tendency for IFN-gamma production rather than LL-I or IL-10 when antigen is presented with peripheral blood mononuclear cells which may contain dendritic cells. It is likely that the nature of the antigen-presenting cell is fundamental in determining the nature of the cytokine profile, which may in turn open up possibilities for new therapeutic modalities.

Rat forming incisor requires a rigorous ECM remodeling modulated by MMP/RECK balance

Reversion-inducing-cysteine-rich protein with Kazal motifs (RECK) is a single membrane-anchored MMP-regulator and regulates matrix metalloproteinases (MMP) 2, 9 and 14. In turn, MMPs are endopeptidases that play a pivotal role in remodeling ECM. In this work, we decided to evaluate expression pattern of RECK in growing rat incisor during, specifically focusing out amelogenesis process. Based on different kinds of ameloblasts, our results showed that RECK expression was conducted by secretory and post-secretory ameloblasts. At the secretory phase, RECK was localized in the infra-nuclear region of the ameloblast, outer epithelium, near blood vessels, and in the stellate reticulum. From the transition to the maturation phases, RECK was strongly expressed by non-epithelial immuno-competent cells (macrophages and/or dendritic-like cells) in the papillary layer. From the transition to the maturation stage, RECK expression was increased. RECK mRNA was amplified by RT-PCR from whole enamel organ. Here, we verified the presence of RECK mRNA during all stages of amelogenesis. These events were governed by ameloblasts and by non-epithelial cells residents in the enamel organ. Concluding, we found differential expression of MMPs-2, -9 and RECK in the different phases of amelogenesis, suggesting that the tissue remodeling is rigorously controlled during dental mineralization.

Genomic organization of the CC chemokine MIP-3 alpha/CCL20/LARC/EXODUS/SCYA20, showing gene structure, splice variants, and chromosome localization

We describe the genomic organization of a recently identified CC chemokine, MIP3 alpha /CCL20 (HGMW-approved symbol SCYA20). The MIP-3 alpha /CCL20 gene was cloned and sequenced, revealing a four exon, three intron structure, and was localized by FISK analysis to 2q35-q36. Two distinct cDNAs were identified, encoding two forms of MIP-3 alpha /CCL20, Ala MLP-3 alpha /CCL20 and Ser MIP-3 alpha /CCL20, that differ by one amino acid at the predicted signal peptide cleavage site. Examination of the sequence around the boundary of intron 1 and exon 2 showed that use of alternative splice acceptor sites could give rise to Ata MIP-3 alpha /CCL20 or Ser MIP-3 alpha /CCL20. Both forms of MIP-3cr/CCL20 were chemically synthesized and tested for biological activity. Both flu antigen plus IL-a-activated CD4(+) and CD8(+) T lymphoblasts and cord blood-derived dendritic cells responded to Ser and Ala MIP-3 alpha /CCL20. T lymphocytes exposed only to IL-2 responded inconsistently, while no response was detected in naive T lymphocytes, monocytes, or neutrophils. The biological activity of Ser MIP-3 alpha /CCL20 and Ala MIP-3 alpha /CCL20 and the tissue-specific preference of different splice acceptor sites are not yet known. (C) 2001 Academic Press.

Association of clinical, radiological and synovial immunopathological responses to anti-rheumatic treatment in rheumatoid arthritis

Objectives. To compare immunohistochemical scoring with clinical scoring and radiology for the assessment of rheumatoid arthritis (RA) disease activity, synovial tissue (ST) biopsied arthroscopically was assessed from 18 patients before and after commencement of disease-modifying anti-rheumatic drug (DMARD) therapy. Methods. Lymphocytes, macrophages, differentiated dendritic cells (DC), vascularity, tumour necrosis factor (TNF)alpha and interleukin-1 beta levels were scored. Clinical status was scored using the American College of Rheumatology (ACR) core set and serial radiographs were scored using the Larsen and Sharp methods. Histopathological evidence of activity included infiltration by lymphocytes, DC, macrophages. tissue vascularity, and expression of lining and sublining TNF alpha. These indices co-varied across the set of ST biopsies and were combined as a synovial activity score for each biopsy. Results. The change in synovial activity with treatment correlated with the ACR clinical response and with decreased radiological progression by the Larsen score, The ACR response to DMARD therapy. the change in synovial activity score and the slowing of radiological progression were each greatest in patients with high initial synovial vascularity. Conclusions. The data demonstrate an association between clinical, radiological and synovial immunopathological responses to anti-rheumatic treatment in RA. High ST vascularity may predict favourable clinical and radiological responses to treatment.

Peripheral T-Cell tolerance defined through transgenic mouse studies

Selection in the thymus restricted by MHC and self-peptide shapes the diverse reactivities of the T-cell population which subsequently seeds into the peripheral tissues, in anticipation of the universe of pathogen antigens to which the organism may be exposed. A necessary corollary is the potential for T-cell self-reactivity (autoimmunity) in the periphery. Transgenic mouse models in which transgene expression in the thymus is prevented or excluded, have been particularly useful for determining the immunological outcome when T-cells encounter transgene-encoded 'self' antigen in peripheral tissues. Data suggest that non-mutually exclusive mechanisms of T-cells 'ignoring' self-antigen, T-cell deletion, T-cell anergy and T-cell immunoregulation have evolved to prevent self-reactivity while maintaining T-cell diversity. The peripheral T-cell repertoire, far from being static following maturation through the thymus, is in a dynamic stated determined by these peripheral selective and immunoregulatory influences. This article reviews the evidence with particular reference to CD8+ive T-cells.

Nonspecific down-regulation of CD8+ T-Cell responses in mice expressing human Papillomavirus Type 16 E7 oncoprotein from the keratin-14 promoter

The E7 oncoprotein of human papillomavirus 16 (HPV16) transforms basal and suprabasal cervical epithelial cells and is a tumor-specific antigen in cervical carcinoma, to which immunotherapeutic strategies aimed at cytotoxic T-lymphocyte (CTL) induction are currently directed. By quantifying major histocompatibility complex class I tetramer-binding T cells and CTL in mice expressing an HPV16 E7 transgene from the keratin-l l (K14) promoter in basal and suprabasal keratinocytes and in thymic cortical epithelium, we show that antigen responsiveness of both E7- and non-E7-specific CD8(+) cells is down-regulation compared to non-E7 transgenic control mice. We show that the effect is specific for E7, and not another transgene, expressed from the K14 promoter, Down-regulation did not involve deletion of CD8(+) T cells of high affinity or high avidity, and T-cell receptor (TCR) VP-chain usage and TCR receptor density were similar in antigen-responsive cells from E7 transgenic and non-E7 transgenic mice. These data indicate that E7 expressed chronically from the K14 promoter nonspecifically down-regulates CD8+ T-cell responses. The in vitro data correlated with the failure of immunized E7 transgenic mice to control the growth of an E7-expressing tumor challenge, We have previously shown that E7-directed CTL down-regulation correlates with E7 expression in peripheral but not thymic epithelium (T, Dean et al., J, Virol. 73:6166-6170, 1999), The findings have implications for the immunological consequences of E7-expressing tumor development and E7-directed immunization strategies. Generically, the findings illustrate a T-cell immunomodulatory function for a virally encoded human oncoprotein.

Characterization of a human synovial cell antigen: VCAM-1 and inflammatory arthritis

The contribution of synovial cells to the pathogenesis of rheumatoid arthritis (RA) is only partly understood. Monoclonal antibody (mAb) 1D5 is one of very few mAb ever raised against RA synovial cells in order to study the biology of these cells. Studies on the expression pattern and structural features of the 1D5 Ag suggest that 1D5 recognizes human vascular cell adhesion molecule-1 (VCAM-1), which is an intercellular adhesion molecule. Vascular cell adhesion molecule-1 may be involved in a number of crucial intercellular interactions in RA.