Cell-dependent Responses Research Articles

Protection against Streptococcus suis Serotype 2 Infection Using a Capsular Polysaccharide Glycoconjugate Vaccine.

Streptococcus suis serotype 2 is an encapsulated bacterium and one of the most important bacterial pathogens in the porcine industry. Despite decades of research for an efficient vaccine, none is currently available. Based on the success achieved with other encapsulated pathogens, a glycoconjugate vaccine strategy was selected to elicit opsonizing anti-capsular polysaccharide (anti-CPS) IgG antibodies. In this work, glycoconjugate prototypes were prepared by coupling S. suis type 2 CPS to tetanus toxoid, and the immunological features of the postconjugation preparations were evaluated in vivo In mice, experiments evaluating three different adjuvants showed that CpG oligodeoxyribonucleotide (ODN) induces very low levels of anti-CPS IgM antibodies, while the emulsifying adjuvants Stimune and TiterMax Gold both induced high levels of IgGs and IgM. Dose-response trials comparing free CPS with the conjugate vaccine showed that free CPS is nonimmunogenic independently of the dose used, while 25 μg of the conjugate preparation was optimal in inducing high levels of anti-CPS IgGs postboost. With an opsonophagocytosis assay using murine whole blood, sera from immunized mice showed functional activity. Finally, the conjugate vaccine showed immunogenicity and induced protection in a swine challenge model. When conjugated and administered with emulsifying adjuvants, S. suis type 2 CPS is able to induce potent IgM and isotype-switched IgGs in mice and pigs, yielding functional activity in vitro and protection against a lethal challenge in vivo, all features of a T cell-dependent response. This study represents a proof of concept for the potential of glycoconjugate vaccines in veterinary medicine applications against invasive bacterial infections.

The transcriptional coactivator Bob1 promotes the development of follicular T helper cells via Bcl6.

Follicular T helper (Tfh) cells are key regulators of the germinal center reaction and long‐term humoral immunity. Tfh cell differentiation requires the sustained expression of the transcriptional repressor Bcl6; however, its regulation in CD4+ T cells is incompletely understood. Here, we report that the transcriptional coactivator Bob1, encoded by the Pou2af1 gene, promotes Bcl6 expression and Tfh cell development. We found that Bob1 together with the octamer transcription factors Oct1/Oct2 can directly bind to and transactivate the Bcl6 and Btla promoters. Mixed bone marrow chimeras revealed that Bob1 is required for the expression of normal levels of Bcl6 and BTLA, thereby controlling the pool size and composition of the Tfh compartment in a T cell‐intrinsic manner. Our data indicate that T cell‐expressed Bob1 is directly involved in Tfh cell differentiation and required for mounting normal T cell‐dependent B‐cell responses.

Distinct courses of infection with Leishmania (L.) amazonensis are observed in BALB/c, BALB/c nude and C57BL/6 mice.

Leishmania (L.) amazonensis [L. (L.) amazonensis] is widely distributed in Brazil and its symptomatic infections usually lead to few localized lesions and sometimes to diffuse cutaneous form, with nodules throughout the body, anergy to parasite antigens and poor therapeutic response. The variability of these manifestations draws attention to the need for studies on the pathophysiology of infection by this species. In this study, we analysed the course and immunological aspects of L. (L.) amazonensis infection in BALB/c and C57BL/6 strains, both susceptible, but displaying different clinical courses, and athymic BALB/c nude, to illustrate the role of T cell dependent responses. We analysed footpad thickness and parasite burden by in vivo imaging. Furthermore, we evaluated the cellular profile and cytokine production in lymph nodes and the inflammatory infiltrates of lesions. Nude mice showed delayed lesion development and less inflammatory cells in lesions, but higher parasite burden than BALB/c and C57BL/6. BALB/c and C57BL/6 mice had similar parasite burdens, lesion sizes and infiltrates until 6 weeks after infection, and after that C57BL/6 mice controlled the infection. Small differences in parasite numbers were observed in C57BL/6 macrophages in vitro, indicating that in vivo milieu accounts for most differences in infection. We believe our results shed light on the role of host immune system in the course of L. (L.) amazonensis infection by comparing three mouse strains that differ in parasitaemia and inflammatory cells.

Survival analysis and microarray profiling identify Cd40 as a candidate for the Salmonella susceptibility locus, Ity5.

The outcome of infection with Salmonella Typhimurium in mouse models of human typhoid fever is dependent upon a coordinated complex immune response. A panel of recombinant congenic strains (RCS) derived from reciprocal backcross of A/J and C57BL/6J mice was screened for their susceptibility to Salmonella infection and two susceptibility loci, Ity4 (Immunity to Typhimurium locus 4) and Ity5, were identified. We validated Ity5 in a genetic environment free of the impact of Ity4 using a cross between A/J and 129S6. Using a time-series analysis of genome-wide transcription during infection, comparing A/J with AcB60 mice having a C57BL/6J-derived Ity5 interval, we have identified the differential expression of the positional candidate gene Cd40, Cd40-associated signaling pathways, and the differential expression of numerous genes expressed in neutrophils. CD40 is known to coordinate T cell-dependent B-cell responses and myeloid cell activation. In fact, CD40 signaling is altered in A/J mice as seen by impaired IgM upregulation during infection, decreased Ig class switching, neutropenia, reduced granulocyte recruitment in response to infection and inflammation, and decreased ERK1/2 activity. These results suggest that altered CD40 signaling and granulocyte recruitment in response to infection are responsible for the Ity5-associated Salmonella susceptibility of A/J mice.

Generation of a soluble recombinant trimeric form of bovine CD40L and its potential use as a vaccine adjuvant in cows

Vaccination is the most cost-effective way to control infectious diseases in cattle. However, many infectious diseases leading to severe economical losses worldwide still remain for which a really effective and safe vaccine is not available. These diseases are most often due to intracellular pathogens such as bacteria or viruses, which are, by their localization, protected from antibiotics and/or CD4+ T cell-dependent humoral responses. We therefore postulated that strategies leading to induction of not only CD4+ T cell responses but also CD8+ cytotoxic T lymphocyte (CTL) responses against infected cells should be privileged in the development of new vaccines against problematic intracellular pathogens in bovines. CD40 signaling in antigen-presenting cells may lead to the induction of robust CD4-independent CTL responses and several studies, especially in mice, have used CD40 stimulation to promote CD8+ T cell-mediated immunity. For example, we have recently shown that immunization of mice with heat-killed Staphylococcus aureus (HKSA) and agonistic anti-CD40 monoclonal antibodies elicits strong CTL responses capable of protecting mice from subsequent staphylococcal mastitis. Unfortunately, there is at present no tool available to efficiently stimulate CD40 in cattle. In this study, we therefore first produced a soluble recombinant trimeric form of the natural bovine CD40 ligand (sboCD40LT). We then observed that sboCD40LT was able to potently stimulate bovine cells in vitro. Finally, we provide evidence that immunization of cows with sboCD40LT combined with HKSA was able to significantly increase the number of both HKSA-specific CD4+ and CD8+ T cells in the draining lymph nodes. In conclusion, we suggest that this new molecular tool could help in the development of vaccine strategies against bovine diseases caused by intracellular pathogens.

CD21 -/low B cells: A Snapshot of a Unique B Cell Subset in Health and Disease.

B cells represent one of the cellular components of the immune system that protects the individual from invading pathogens. In response to the invader, these cells differentiate into plasma cells and produce large amounts of antibodies that bind to and eliminate the pathogen. A hallmark of autoimmune diseases is the production of autoantibodies i.e. antibodies that recognize self. Those that are considered pathogenic can damage tissues and organs, either by direct binding or when deposited as immune complexes. For decades, B cells have been considered to play a major role in autoimmune diseases by antibody production. However, as pathogenic autoantibodies appear to derive mainly from T cell dependent responses, T cells have been the focus for many years. The successful treatment of patients with autoimmune diseases with either B cell depletion therapy (rituximab) or inhibition of B cell survival (belimumab), suggested that not only the autoantibodies but also other B cell features are important. This has caused a surge of interest in B cells and their biology resulting in the identification of various subsets e.g. regulatory B cells, several memory B cell subsets etc. Also, in other conditions such as chronic viral infections and primary immunodeficiency, several B cell subsets with unique characteristics have been identified. In this review, we will discuss one of these subsets, a subset that is expanded in conditions characterized by chronic immune stimulation. This B cell subset lacks, or expresses low, surface levels of the complement receptor 2 (CD21) and has therefore been termed CD21(-/low) B cells.

Abstract A103: Intracellular immunotherapy of disseminated pancreatic cancer activates potent IFNγ and CD8+ T cell dependent anti-tumor responses

Abstract Innovative therapies with the ability to activate effective cellular-based immune mechanisms in conjunction with current treatment strategies have potential to combat highly aggressive solid and metastatic tumors such as pancreatic adenocarcinoma, which remains one of the most deadly cancers with an overall 5-year survival rate of 6% following diagnosis. In this study, we explored the immunotherapeutic mechanisms triggered by an avirulent and nonreplicating obligate intracellular parasite Toxoplasma gondii (cps strain), which preferentially invades myeloid cell types, in the disseminated pancreatic tumor environment. We hypothesized the strong induction of Th1 responses in the tumor environment triggered by cps may stimulate significant mechanisms that could halt or eliminate established pancreatic cancer. Pan02 cells were injected intraperitoneally into syngeneic C57BL/6 mice. Seven days later, these mice were treated with 2, 3, or 5 doses of cps or PBS, as a treatment control. Pan02 tumor-bearing mice treated with cps had a significant survival advantage over untreated mice. Additionally, treatment with heat-killed cps, which is unable to actively invade, led to loss of treatment efficacy. Following a single cps treatment CD11b+CD11c+ dendritic cell populations increased and CD11b+F4/80+CD11c- macrophage populations decreased in the peritoneum. Cps-invaded macrophages and dendritic cells in the tumor environment upregulated surface expression of CD80 and CD86, and there was an increase in local production of IL-12 up to 7 days post-treatment. Pan02 tumor-bearing mice unable to produce IL-12 lost protection mediated by cps treatment. Seven days post-cps treatment, both CD4+ and CD8+ T cells were significantly higher than within the tumor microenvironment of untreated mice. Local T cells had lower expression of CD62L and increased expression of CD44 and CD69 indicating activation of recruited T cells. Six and seven days post-cps treatment, local IFNγ production in Pan02 tumor-bearing mice increased and the numbers of IFNγ+ CD4+ and IFNγ+ CD8+ T cells were higher. Similar to IL-12, loss of IFNγ within the tumor microenvironment led to a loss of treatment efficacy. A lack of CD8+ T cells prior to and during therapy resulted in a loss of treatment efficacy, whereas CD4+ T cells and NK cells were not required for treatment efficacy. CD8+ T cells isolated from the spleens of treated animals 10 days after cps treatment robustly produced IFNγ ex-vivo when stimulated with Pan02 cells. This response appears to be tumor specific as stimulation with irrelevant tumor antigen did not induce an IFNγ response from the same CD8+ T cells. Cps-mediated activation of tumor-associated myeloid cells leads to increases in local IFNγ+ CD8+ T cells, which drive specific anti-tumor responses. Cps induction of host Th1 responses elicited in the tumor microenvironment demonstrates the capability of cps to be broadly applied as an immunotherapeutic against pancreatic tumors. Citation Format: Kiah L. Sanders, Barbara A. Fox, David J. Bzik. Intracellular immunotherapy of disseminated pancreatic cancer activates potent IFNγ and CD8+ T cell dependent anti-tumor responses. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Innovations in Research and Treatment; May 18-21, 2014; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2015;75(13 Suppl):Abstract nr A103.

Microbes taming mast cells: Implications for allergic inflammation and beyond

There is increasing awareness of a relationship between our microbiota and the pathogenesis of allergy and other inflammatory diseases. In investigating the mechanisms underlying microbiota modulation of allergy the focus has been on the induction phase; alterations in the phenotype and function of antigen presenting cells, induction of regulatory T cells and shifts in Th1/Th2 balance. However there is evidence that microbes can influence the effector phase of disease, specifically that certain potentially beneficial bacteria can attenuate mast cell activation and degranulation. Furthermore, it appears that different non-pathogenic bacteria can utilize distinct mechanisms to stabilize mast cells, acting locally though direct interaction with the mast cell at mucosal sites or attenuating systemic mast cell dependent responses, likely through indirect signaling mechanisms. The position of mast cells on the frontline of defense against pathogens also suggests they may play an important role in fostering the host–microbiota relationship. Mast cells are also conduits of neuro-immuo-endocrine communication, suggesting the ability of microbes to modulate cell responses may have implications for host physiology beyond immunology. Further investigation of mast cell regulation by non-pathogenic or symbiotic bacteria will likely lead to a greater understanding of host microbiota interaction and the role of the microbiome in health and disease.

Alarmin' Immunologists: IL-33 as a Putative Target for Modulating T Cell-Dependent Responses.

IL-33 is a known member of the IL-1 cytokine superfamily classically named “atypical” due to its diverse functions. The receptor for this cytokine is the ST2 chain (or IL-1RL1), part of the IL-1R family, and the accessory chain IL-1R. ST2 can be found as both soluble and membrane-bound forms, property that explains, at least in part, its wide range of functions. IL-33 has increasingly gained our attention as a potential target to modulate immune responses. At the beginning, it was known as one of the participants during the development of allergic states and other Th2-mediated responses and it is now accepted that IL-33 contributes to Th1-driven pathologies as demonstrated in animal models of experimental autoimmune encephalomyelitis (EAE), collagen-induced arthritis, and trinitrobenzene sulfonic acid-induced experimental colitis, among others. Interestingly, current data are placing IL-33 as a novel regulator of immune tolerance by affecting regulatory T cells (Tregs); although the mechanism is not fully understood, it seems that dendritic cells and myeloid suppressor-derived cells may be cooperating in the generation and/or establishment of IL-33-mediated tolerance. Here, we review the most updated literature on IL-33, its role on T cell biology, and its impact in immune tolerance.

Pim kinases significantly impact humoral immune responses (IRM12P.648)

Abstract The Provirus Integration site for Moloney murine leukemia virus (Pim) family of oncogenic serine/threonine kinases regulates a variety of metabolic functions in hematopoietic cells and lymphocytes. We examined the importance of Pim kinases in B cell activation following antigen stimulation. Humoral immune responses were significantly impaired in Pim 1 and 2 deficient mice (Pim 1-/-2-/-) T cell dependent responses were compromised while T cell independent type 1 and 2 antigen responses were virtually absent. Humoral deficiencies were owing to reduced production of antibody secreting cells due to the diminished induction of BLIMP-1 expression. The central role of Pim in these responses is consistent with our observation that Pims are induced downstream of multiple B cell activation receptors including CD40, BCR, and TLR4. We suggest that Pim 1 and 2 dependent signaling pathways are involved in B cell activation and their loss requires increased signaling through other pathways to reach an activation threshold. This hypothesis is further supported by the initial higher affinity of BCRs on Pim deficient B cells activated in a TD response and their inability to proliferate or undergo isotype switching in culture at the same rate as WT B cells when activator concentrations are decreased. Taken together, these data demonstrate a critical role for Pim kinases in affinity maturation, isotype switching, proliferation and the production of antibody secreting cells by B lymphocytes. .

A monomeric fab fragment capable of inducing CD3 conformational change increases T cell receptor reactivity to poorly immunogenic antigens (TUM2P.1020)

Abstract TCR interactions with agonist pMHCs induce a conformational change in the tails of the associated CD3 complex (CD3Δc) that contributes to efficient TCR/CD3 triggering and productive T cell responses. In contrast, TCR engagement by antagonistic pMHCs does not induce either CD3Δc or full TCR/CD3 signaling in mature T cells. Here, we have used a monovalent Fab fragment (Mono-Fab) specific for the murine CD3 complex to provide induction of CD3Δc to mature T cells engaging pMHC ligands. This Mono-Fab does not interfere with TCR capacity to bind pMHC and fails to elicit signaling in the absence of TCR/MHC interactions. Using OT-I Tg T cells and a collection of altered peptide ligands derived of the agonist peptide OVA, we show that addition of CD3Δc to OT-I T cells using Mono-Fab enables T cell responses to antagonist ligands that fail to induce CD3Δc on their own. Moreover, using a mouse model of metastatic melanoma, we show that Mono-Fab promotes in vivo T cell dependent anti-melanoma responses that are antigen-specific. This Mono-Fab can also be combined with established therapies targeting signals 2 and 3 involved in T cell activation for a synergistic effect. Our data suggests that in-trans CD3Δc provision decreases the activation threshold of T cells. We propose that targeting the CD3 complex for the induction of CD3Δc could be exploited to achieve efficient TCR/CD3 triggering by poorly immunogenic antigens in the context of vaccination against cancer and infectious diseases.

Aryl hydrocarbon receptor modulates the development and activity of innate-like B cells (HEM1P.226)

Abstract The Aryl hydrocarbon receptor (AhR) is a ligand-activated transcription factor that recognizes polyaromatic hydrocarbons. While the contributions of AhR ligands to the differentiation and functions of Th17/Treg cells are relatively well understood, little is known about AhR in B cells. Microarray and quantitative PCR analysis of B cell subsets revealed that B1 and marginal zone (MZ) B cells expressed higher levels of AhR compared to follicular (FO) B cells. We found ~ two-fold more B1 B cells, but fewer MZ B cells in AhR-/- mice compared to AhR+/+ mice. Following immunization with Streptococcus pneumonia, AhR-/- mice had significantly higher titers of anti-phosphorylcholine IgM antibodies. In contrast, after immunization with a model T cell-dependent antigen, Trinitrophenyl Keyhole Limpet Hemocyanin (TNP-KLH), AhR-/- mice had anti-TNP titers that were comparable to those in AhR+/+ mice. Further, we found significantly fewer B cells in C57BL/6 mice raised on AhR ligand free diet, and an increase in titers of IgM and IgG3 in mice following a switch to an AhR ligand free diet when compared to littermates raised and maintained on conventional diet. Taken together, these findings suggests that AhR plays a role in the development of B1 and MZ B cells and regulates T cell-independent but not T cell-dependent humoral response. Further studies using mice with a B cell specific deletion of AhR are underway to elucidate the mechanism by which AhR ligands influence humoral immunity.

Border patrol gone awry: lung NKT cell activation by F. tularensis exacerbates tularemia-like disease (MUC1P.908)

Abstract The respiratory mucosa is a major site for pathogen invasion and, hence, a site requiring constant immune surveillance. The semi-invariant natural killer T (NKT) cells are enriched within the lung vasculature. Despite optimal positioning for border patrol, the role of NKT cells in respiratory infectious diseases remains poorly understood. Hence, we assessed their function in a murine model of pulmonary tularemia—because tularemia is a sepsis-like proinflammatory disease and NKT cells are known to control the cellular and humoral responses underlying sepsis. Here we show for the first time that respiratory infection with Francisella tularensis live vaccine strain results in rapid recruitment of NKT cells to the lung interstitium. Interstitial NKT cell activation resulted in protective interferon-γ production, and affected both local and systemic proinflammatory responses. Hence, NKT cell-deficient mice showed reduced inflammatory cytokine and chemokine response yet survived the infection whilst their wild type counterparts did not. Strikingly, NKT cell-deficient mice had increased lymphocytic infiltration in the lungs that organized into structures resembling induced bronchus-associated lymphoid tissue (iBALT) at the peak of infection. Thus, NKT cell activation by F tularensis infection hampers iBALT formation, which in conjunction with NKT cell-dependent proinflammatory response causes severe pulmonary tularemia-like disease in mice.

An inhibitory receptor leukocyte mono-immunoglobulin-like receptor 3/CD300f deficiency aggravates DSS-induced colitis (MUC8P.737)

Abstract Leukocyte mono-immunoglobulin-like receptor 3 (LMIR3) (also called CD300f) is an inhibitory receptor. LMIR3 is mainly expressed in myeloid cells, including mast cells. The binding of LMIR3 to its ligand ceramide inhibits IgE- and mast cell-dependent allergic responses. It was recently reported that ATP-induced activation of mast cells via P2X7 purinoceptor contributes to the initiation and progression of intestinal inflammation. Here we show that LMIR3 is a negative regulator of dextran sodium sulfate (DSS)-induced colitis, a murine model of ulcerative colitis (UC). Notably, Lmir3-/- mice were highly susceptible to DSS-induced colitis. We found a prominent increase in inflammatory cells, including degranulated mast cells, in the colonic lamina propria of DSS-treated Lmir3-/- mice. Mast cell-deficient KitWsh/Wsh mice engrafted with LMIR3 deficient bone-marrow-derived mast cells (BMMCs) exhibited increased susceptibility to DSS-induced colitis as compared with those with wild-type (WT) BMMCs. Importantly, ceramide-LMIR3 binding suppressed activation of ATP-stimulated BMMCs. Consistently, treatment with ceramide liposomes improved DSS-induced colitis in WT mice, but not in Lmir3-/- mice. Taken together, these results indicated that LMIR3 negatively regulated DSS-induced colitis by inhibiting ATP-mediated mast cell activation through ceramide-LMIR3 binding. Accordingly, LMIR3-targeted ceramide liposomes would provide novel therapeutic strategies for UC.

FCRL5 regulates innate-like B cell natural antibody production and T cell-independent responses (IRM10P.607)

Abstract At homeostasis and during primary responses to pathogens, humoral host defense is chiefly coordinated by body cavity-derived B-1 and splenic marginal zone (MZ) B cells. These early responders have been termed “innate-like” due to their broadly-reactive natural antibody repertoires and rapid responses to T cell-independent (TI) antigens. However, what regulates these functions in these specialized cells remains unclear. Fc receptor-like (FCRL) molecules have homology to the classical FCRs for IgG and IgE, exhibit complex tyrosine-based regulation, and are preferentially expressed by B cells. Notably, these receptor-genes have been increasingly associated with immune disorders and human (h) FCRL3 has emerged as a risk factor in autoimmunity. To better understand hFCRL3’s role in B cells, we investigated its closest relative, mouse FCRL5 (mFCRL5), which shares homologous extracellular domains, binary tyrosine-based signaling, and innate-like B cell expression. Here we characterize the first in vivo model of FCRL function by examining Fcrl5 deficient mice. Our findings demonstrate that at homeostasis Fcrl5 deficient mice have normal total B cell numbers and subset frequencies, but enhanced natural antibody production. By contrast, Fcrl5 deficiency was found to impair TI humoral responses, but T cell-dependent responses remained intact. These data indicate a critical role for mFCRL5 in regulating innate-like B cell function at the interface of host defense and tolerance.

Role of CD154 in cancer pathogenesis and immunotherapy

Many factors and molecules have been investigated as potential players in the pathogenesis or immunosurveillance of cancer. Among these, CD154 has been recognized as a co-stimulatory molecule with high potential for treating cancer, in addition to its contribution in the development of the disease. CD154 was initially described for its pivotal role in T cell-dependent humoral responses via an interaction with its classical receptor, CD40. Subsequent studies showed that CD154 is also implicated in cell-mediated immunity and inflammation via an interaction with CD40 alone or in combination with newly identified receptors, members of the integrin family, leading to the development of chronic inflammatory and autoimmune diseases. In the current article, we present an overview of the role of CD154 as a potential etiological factor in tumors inducing proliferation of malignant cells, their rescue from apoptosis and their invasiveness. In addition, this review describes the immuno-regulatory functions of CD154 against cancer reflected by its stimulation of antigen-presenting cells and the subsequent activation of effector cells, its enhancement of malignant cells' immunogenicity, its modulation of immune settings around tumors, and its initiation of proliferation inhibiting effects in malignant cells. In vitro as well as in vivo studies are outlined and a particular attention is given to clinical studies and progress reached at this point. Findings reviewed herein will improve our knowledge of the role of the CD154 system in cancers from causative to immunotherapeutic functions, paving the way for the identification of new targets for prevention and/or treatment of malignant disorders.

Regulation of Leishmania (L.) amazonensis protein expression by host T cell dependent responses: differential expression of oligopeptidase B, tryparedoxin peroxidase and HSP70 isoforms in amastigotes isolated from BALB/c and BALB/c nude mice.

Leishmaniasis is an important disease that affects 12 million people in 88 countries, with 2 million new cases every year. Leishmania amazonensis is an important agent in Brazil, leading to clinical forms varying from localized (LCL) to diffuse cutaneous leishmaniasis (DCL). One interesting issue rarely analyzed is how host immune response affects Leishmania phenotype and virulence. Aiming to study the effect of host immune system on Leishmania proteins we compared proteomes of amastigotes isolated from BALB/c and BALB/c nude mice. The athymic nude mice may resemble patients with diffuse cutaneous leishmaniasis, considered T-cell hyposensitive or anergic to Leishmania´s antigens. This work is the first to compare modifications in amastigotes’ proteomes driven by host immune response. Among the 44 differentially expressed spots, there were proteins related to oxidative/nitrosative stress and proteases. Some correspond to known Leishmania virulence factors such as OPB and tryparedoxin peroxidase. Specific isoforms of these two proteins were increased in parasites from nude mice, suggesting that T cells probably restrain their posttranslational modifications in BALB/c mice. On the other hand, an isoform of HSP70 was increased in amastigotes from BALB/c mice. We believe our study may allow identification of potential virulence factors and ways of regulating their expression.

Ceramide-CD300f binding suppresses experimental colitis by inhibiting ATP-mediated mast cell activation

ObjectiveExtracellular ATP mediates mast cell-dependent intestinal inflammation via P2X7 purinoceptors. We have previously shown that CD300f (also called the leucocyte mono-immunoglobulin-like receptor 3 (LMIR3)) suppresses immunoglobulin E-dependent and mast cell-dependent...

Eosinophils are Required for Immune Responses Induced by Oral Immunization

Eosinophils are multifunctional leukocytes that reside in several tissues, most abundantly in the small intestinal lamina propria under the steady state. To date, the phenotypic and functional characteristics of small intestinal eosinophils have remained poorly understood. In this study, we found that proliferation of ovalbumin (OVA)-specific CD4 + T cells isolated from the mesenteric lymph nodes of eosinophil-deficient ΔdblGATA mice were decreased relative to wild-type mice after oral immunization with OVA and cholera toxin (CT), the typical mucosal adjuvant that induces CD4 + T cell-dependent responses. ΔdblGATA mice showed reduced mucosal secretion of OVA-specific IgA and IgG1 while maintaining a systemic level of anti-OVA IgG1 upon oral immunization with OVA and CT. These findings suggest that eosinophils might have a role in the modulation of T cell-mediated immune responses including mucosal antibody responses in the gastrointestinal tract following oral immunization.

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An inhibitory receptor LMIR3/CD300f is mainly expressed in myeloid cells, including mast cells and neutrophils. We have recently demonstrated that ceramide-LMIR3 binding inhibits IgE- and mast cell-dependent allergic responses. Sepsis remains a major clinical problem. Negative regulation of innate immunity is associated with sepsis progression. Here we identify the critical role of ceramide-LMIR3 binding in suppressing innate host responses. LMIR3−/− mice were protected against lethality after cecal ligation and puncture (CLP), a murine model of septic peritonitis. In the peritoneal cavity of CLP-operated LMIR3−/− mice, mast cells and recruited neutrophils released high levels of neutrophil chemoattractants, leading to enhanced recruitment of neutrophils that efficiently eliminated Escherichia coli. Ceramide-LMIR3 interaction suppressed such release from Escherichia coli-stimulated mast cells and neutrophils. Importantly, treatment with ceramide antibody or LMIR3-Fc, which disrupted the ceramide-LMIR3 interaction, prevented CLP-induced sepsis by profoundly stimulating neutrophil recruitment. Thus, LMIR3 is an attractive target for the treatment of sepsis.