The subunits of IL-12, originating from two distinct cells, can functionally synergize to protect against pathogen dissemination in vivo.

Cytokines are typically single gene products, except for the heterodimeric interleukin (IL)-12 family. The two subunits (IL-12p40 and IL-12p35) of the prototype IL-12 are known to be simultaneously co-expressed in activated myeloid cells, which secrete the fully active heterodimer to promote interferon (IFN)γ production in innate and adaptive cells. We find that chimeric mice containing mixtures of cells that can only express either IL-12p40 or IL-12p35, but not both together, generate functional IL-12. This alternate two-cell pathway requires IL-12p40 from hematopoietic cells to extracellularly associate with IL-12p35 from radiation-resistant cells. The two-cell mechanism is sufficient to propel local T cell differentiation in sites distal to the initial infection and helps control systemic dissemination of a pathogen, although not parasite burden, at the site of infection. Broadly, this suggests that early secretion of IL-12p40 monomers by sentinel cells at the infection site may help prepare distal host tissues for potential pathogen arrival.

Safety and Immunogenicity of ChAd63/MVA Pfs25-IMX313 in a Phase I First-in-Human Trial.

Background: Transmission blocking vaccines targeting the sexual-stages of the malaria parasite could play a major role to achieve elimination and eradication of malaria. The Plasmodium falciparum Pfs25 protein (Pfs25) is the most clinically advanced candidate sexual-stage antigen. IMX313, a complement inhibitor C4b-binding protein that forms heptamers with the antigen fused to it, improve antibody responses. This is the first time that viral vectors have been used to induce antibodies in humans against an antigen that is expressed only in the mosquito vector. Methods: Clinical trial looking at safety and immunogenicity of two recombinant viral vectored vaccines encoding Pfs25-IMX313 in healthy malaria-naive adults. Replication-deficient chimpanzee adenovirus serotype 63 (ChAd63) and the attenuated orthopoxvirus modified vaccinia virus Ankara (MVA), encoding Pfs25-IMX313, were delivered by the intramuscular route in a heterologous prime-boost regimen using an 8-week interval. Safety data and samples for immunogenicity assays were taken at various time-points. Results: The reactogenicity of the vaccines was similar to that seen in previous trials using the same viral vectors encoding other antigens. The vaccines were immunogenic and induced both antibody and T cell responses against Pfs25, but significant transmission reducing activity (TRA) was not observed in most volunteers by standard membrane feeding assay. Conclusion: Both vaccines were well tolerated and demonstrated a favorable safety profile in malaria-naive adults. However, the transmission reducing activity of the antibodies generated were weak, suggesting the need for an alternative vaccine formulation. Trial Registration: Clinicaltrials.gov NCT02532049.

Design of a Novel Multi-Epitope Vaccine Against Echinococcus granulosus in Immunoinformatics.

All the time, echinococcosis is a global zoonotic disease which seriously endangers public health all over the world. In order to speed up the development process of anti-Echinococcus granulosus vaccine, at the same time, it can also save economic cost. In this study, immunoinformatics tools and molecular docking methods were used to predict and screen the antigen epitopes of Echinococcus granulosus, to design a multi-epitope vaccine containing B- and T-cell epitopes. The multi-epitope vaccine could activate B lymphocytes to produce specific antibodies theoretically, which could protect the human body against Echinococcus granulosus infection. It also could activate T lymphocytes and clear the infected parasites in the body. In this study, four CD8+ T-cell epitopes, three CD4+ T-cell epitopes and four B-cell epitopes of Protein EgTeg were identified by immunoinformatics methods. Meanwhile, three CD8+ T-cell epitopes, two CD4+ T-cell epitopes and four B-cell epitopes of Protein EgFABP1 were identified. We constructed the multi-epitope vaccine using linker proteins. The study based on the traditional methods of antigen epitope prediction, further optimized the prediction results combined with molecular docking technology and improved the precision and accuracy of the results. Finally, in vivo and in vitro experiments had verified that the vaccine designed in this study had good antigenicity and immunogenicity.

Dendritic cell engineered cTXN as new vaccine prospect against L. donovani.

Dendritic cells (DCs), as antigen-presenting cells, can reportedly be infected withLeishmaniaparasites and hence provide a better option to trigger T-cell primary immune responses and immunological memory. We consistently primed DCs during culture with purified recombinant cytosolic tryparedoxin (rcTXN) and then evaluated the vaccine prospect of presentation of rcTXN against VL in BALB/c mice. We reported earlier the immunogenic properties of cTXN antigen derived fromL. donovani when anti-cTXN antibody was detected in the sera of kala-azar patients. It was observed that cTXN antigen, when used as an immunogen with murine DCs acting as a vehicle, was able to induce complete protection against VL in an infected group of immunized mice. This vaccination triggered splenic macrophages to produce more IL-12 and GM-CSF, and restricted IL-10 release to a minimum in an immunized group of infected animals. Concomitant changes in T-cell responses against cTXN antigen were also noticed, which increased the release of protective cytokine-like IFN-γ under the influence of NF-κß in the indicated vaccinated group of animals. All cTXN-DCs-vaccinated BALB/c mice survived during the experimental period of 120 days. The results obtained in our study suggest that DCs primed with cTXN can be used as a vaccine prospect for the control of visceral leishmaniasis.

Designing peptide-based vaccine candidates for Plasmodium falciparum erythrocyte binding antigen 175.

Plasmodium falciparum leads to a virulent form of malaria. Progress has been achieved in understanding the mechanisms involved in the malarial infection, still there is no effective vaccine to prevent severe infection. An effective vaccine against malaria should be one which can induce immune responses against multiple epitopes in the context of predominantly occurring HLA alleles. In this study, an integrated approach was employed to identify promiscuous peptides of a well-defined sequence of erythrocyte binding antigen-175 and promiscuous peptides for HLA alleles were designed using bioinformatics tools. A peptide with 15 amino acids (ILAIAIYESRILKRK) was selected based on its high binding affinity score and synthesized. This promiscuous peptide was used as stimulating antigen in lymphoproliferative responses to evaluate the cellular immune response. It was observed this peptide evokes lymphoproliferative and cytokine responses in individuals naturally exposed to the malaria parasite. The intensity of PBMCs proliferation was observed to be higher in sera obtained from P. falciparum exposed as compared to unexposed healthy individuals, suggesting earlier recognition of peptide of this region by T cells. Furthermore, the binding mode of HLA-peptide complex and their interaction may lead to a rational and selective peptide-based vaccine candidate design approach which can be used as a malaria prophylaxis.

Innate immune responses to malaria-infected erythrocytes in pregnant women: Effects of gravidity, malaria infection, and geographic location.

BACKGROUND: Malaria in pregnancy causes maternal, fetal and neonatal morbidity and mortality, and maternal innate immune responses are implicated in pathogenesis of these complications. The effects of malaria exposure and obstetric and demographic factors on the early maternal immune response are poorly understood. METHODS: Peripheral blood mononuclear cell responses to Plasmodium falciparum-infected erythrocytes and phytohemagglutinin were compared between pregnant women from Papua New Guinea (malaria-exposed) with and without current malaria infection and from Australia (unexposed). Elicited levels of inflammatory cytokines at 48 h and 24 h (interferon γ, IFN-γ only) and the cellular sources of IFN-γ were analysed. RESULTS: Among Papua New Guinean women, microscopic malaria at enrolment did not alter peripheral blood mononuclear cell responses. Compared to samples from Australia, cells from Papua New Guinean women secreted more inflammatory cytokines tumor necrosis factor-α, interleukin 1ß, interleukin 6 and IFN-γ; p<0.001 for all assays, and more natural killer cells produced IFN-γ in response to infected erythrocytes and phytohemagglutinin. In both populations, cytokine responses were not affected by gravidity, except that in the Papua New Guinean cohort multigravid women had higher IFN-γ secretion at 24 h (p = 0.029) and an increased proportion of IFN-γ+ Vδ2 γδ T cells (p = 0.003). Cytokine levels elicited by a pregnancy malaria-specific CSA binding parasite line, CS2, were broadly similar to those elicited by CD36-binding line P6A1. CONCLUSIONS: Geographic location and, to some extent, gravidity influence maternal innate immunity to malaria.

In the presence of Trypanosoma cruzi antigens, activated peripheral T lymphocytes retained in the liver induce a proinflammatory phenotypic and functional shift in intrahepatic T lymphocyte.

In secondary lymphoid organs, pathogen-derived and endogenous danger molecules are recognized by pattern recognition receptors, leading to adaptive proinflammatory immune responses. This conceptual rule does not apply directly to the liver, as hepatic immune cells tolerate gut-derived bacterial molecules from the flora. Therefore, the recognition of danger and proinflammatory stimuli differs between the periphery and the liver. However, the tolerant nature of the liver must be overcome in the case of infections or cancer, for example. The central paradigm is the basis for danger recognition and the balance between inflammation and tolerance in the liver. Here, we observed functional integration, with activated peripheral T lymphocytes playing a role in the induction of a proinflammatory environment in the liver in the presence of Trypanosoma cruzi antigens. When only parasite extract was orally administered, it led to the up-regulation of hepatic tolerance markers, but oral treatment plus adoptively transferred activated splenic T lymphocytes led to a proinflammatory response. Moreover, treated/recipient mice showed increased levels of TNF, IFN-γ, IL-6, and CCL2 in the liver and increased numbers of effector and/or effector memory T lymphocytes and F4/80+ cells. There was a reduction in FoxP3+ Treg cells, NKT cells, and γδ T lymphocytes with increased liver damage in the presence of activated peripheral T cells. Our results show that the induction of a proinflammatory liver response against T. cruzi danger molecules is at least partially dependent on cooperation with activated peripheral T cells.

Muscular Sarcocystosis in Sheep Associated With Lymphoplasmacytic Myositis and Expression of Major Histocompatibility Complex Class I and II.

Sarcocystosis is a protozoal disease affecting a wide range of animals. The aims of this study were to characterize the following in sheep: (1) the muscle pathology in Sarcocystis infection, (2) the inflammatory infiltrate and its relationship to severity of infection, and (3) immune markers expressed by parasitized muscle fibers and parasitic cysts. Skeletal muscle samples from 78 sheep slaughtered in southern Italy were snap frozen and analyzed by histopathology, immunohistochemistry, and immunofluorescence. Polymerase chain reaction (PCR) and sequencing were used for Sarcocystis species identification. All 40 muscle samples tested were PCR-positive for Sarcocystis tenella. Histologically, cysts were identified in 76/78 cases (97%), associated with an endomysial infiltrate of lymphocytes and plasma cells. The T cells were predominantly CD8+, with fewer CD4+ or CD79α+ cells. Eosinophils were absent. Notably, sarcolemmal immunopositivity for major histocompatibility complex (MHC) I and II was found in 76/78 cases (97%) and 75/78 cases (96%), respectively, both in samples with and in those without evident inflammatory infiltrate. The number of cysts was positively correlated with inflammation. In addition, MHC I was detected in 55/78 cyst walls (72%), and occasionally co-localized with the membrane-associated protein dystrophin. The findings suggest that muscle fibers respond to the presence of cysts by expression of MHC I and II. The possible role of MHC I and II in the inflammatory response and on the cyst wall is also discussed.

Trypanosoma brucei gambiense excreted/secreted factors impair lipopolysaccharide-induced maturation and activation of human monocyte-derived dendritic cells.

Trypanosoma brucei gambiense, an extracellular eukaryotic flagellate parasite, is the main etiological agent of human African trypanosomiasis (HAT) or sleeping sickness. Dendritic cells (DCs) play a pivotal role at the interface between innate and adaptive immune response and are implicated during HAT. In this study, we investigated the effects of T gambiense and its excreted/secreted factors (ESF) on the phenotype of human monocyte-derived DCs (Mo-DCs). Mo-DCs were cultured with trypanosomes, lipopolysaccharide (LPS), ESF derived from T gambiense bloodstream strain Biyamina (MHOM/SD/82), or both ESF and LPS. Importantly, ESF reduced the expression of the maturation markers HLA-DR and CD83, as well as the secretion of IL-12, TNF-alpha and IL-10, in LPS-stimulated Mo-DCs. During mixed-leucocyte reactions, LPS- plus ESF-exposed DCs induced a non-significant decrease in the IFN-gamma/IL-10 ratio of CD4 + T-cell cytokines. Based on the results presented here, we raise the hypothesis that T gambiense has developed an immune escape strategy through the secretion of paracrine mediators in order to limit maturation and activation of human DCs. The identification of the factor(s) in the T gambiense ESF and of the DCs signalling pathway(s) involved may be important in the development of new therapeutic targets.

Histamine releasing factor and elongation factor 1 alpha secreted via malaria parasites extracellular vesicles promote immune evasion by inhibiting specific T cell responses.

Protozoan pathogens secrete nanosized particles called extracellular vesicles (EVs) to facilitate their survival and chronic infection. Here, we show the inhibition by Plasmodium berghei NK65 blood stage-derived EVs of the proliferative response of CD4+ T cells in response to antigen presentation. Importantly, these results were confirmed in vivo by the capacity of EVs to diminish the ovalbumin-specific delayed type hypersensitivity response. We identified two proteins associated with EVs, the histamine releasing factor (HRF) and the elongation factor 1α (EF-1α) that were found to have immunosuppressive activities. Interestingly, in contrast to WT parasites, EVs from genetically HRF- and EF-1α-deficient parasites failed to inhibit T cell responses in vitro and in vivo. At the level of T cells, we demonstrated that EVs from WT parasites dephosphorylate key molecules (PLCγ1, Akt, and ERK) of the T cell receptor signalling cascade. Remarkably, immunisation with EF-1α alone or in combination with HRF conferred a long-lasting antiparasite protection and immune memory. In conclusion, we identified a new mechanism by which P. berghei-derived EVs exert their immunosuppressive functions by altering T cell responses. The identification of two highly conserved immune suppressive factors offers new conceptual strategies to overcome EV-mediated immune suppression in malaria-infected individuals.

Preventive CTLA-4-Ig Treatment Reduces Hepatic Egg Load and Hepatic Fibrosis in Schistosoma mansoni-Infected Mice.

BACKGROUND: Hepatic fibrosis and granuloma formation as a consequence of tissue entrapped eggs produced by female schistosomes characterize the pathology of Schistosoma mansoni infection. We have previously shown that single-sex infection with female schistosomes mitigates hepatic fibrosis after secondary infection. This was associated with an increased expression of cytotoxic T-lymphocyte-associated protein-4 (CTLA-4), known as a negative regulator of T cell activation. Based on these findings, we hypothesized that administration of agonistic CTLA-4-Ig (Belatacept) is capable to prevent and/or treat hepatic fibrosis during schistosomiasis. METHODS: Mice were infected with 50 S. mansoni cercariae and CTLA-4-Ig, or appropriated control-Ig was administered for 4 weeks. Preventive treatment started 4 weeks after infection, before onset of egg production, and therapeutic treatment started 8 weeks after infection when hepatic fibrosis was already established. RESULTS: When given early after infection, livers of CTLA-4-Ig-treated mice showed significantly reduced collagen deposition and decreased expression of profibrotic genes in comparison to controls. In addition, administration of CTLA-4-Ig suppressed the inflammatory T cell response in infected mice. If therapy was started at a later time point when fibrogenesis was initiated, CTLA-4-Ig had no impact on hepatic fibrosis. CONCLUSION: We could demonstrate that an early preventive administration of CTLA-4-Ig suppresses effector T cell function and therefore ameliorates liver fibrosis. CTLA-4-Ig administration after onset of egg production fails to treat hepatic fibrosis.

The ectodomains of the lymphocyte scavenger receptors CD5 and CD6 interact with tegumental antigens from Echinococcus granulosus sensu lato and protect mice against secondary cystic echinococcosis.

BACKGROUND: Scavenger Receptors (SRs) from the host's innate immune system are known to bind multiple ligands to promote the removal of non-self or altered-self targets. CD5 and CD6 are two highly homologous class I SRs mainly expressed on all T cells and the B1a cell subset, and involved in the fine tuning of activation and differentiation signals delivered by the antigen-specific receptors (TCR and BCR, respectively), to which they physically associate. Additionally, CD5 and CD6 have been shown to interact with and sense the presence of conserved pathogen-associated structures from bacteria, fungi and/or viruses. METHODOLOGY/PRINCIPAL FINDINGS: We report herein the interaction of CD5 and CD6 lymphocyte surface receptors with Echinococcus granulosus sensu lato (s.l.). Binding studies show that both soluble and membrane-bound forms of CD5 and CD6 bind to intact viable protoscoleces from E. granulosus s.l. through recognition of metaperiodate-resistant tegumental components. Proteomic analyses allowed identification of thioredoxin peroxidase for CD5, and peptidyl-prolyl cis-trans isomerase (cyclophilin) and endophilin B1 (antigen P-29) for CD6, as their potential interactors. Further in vitro assays demonstrate that membrane-bound or soluble CD5 and CD6 forms differentially modulate the pro- and anti-inflammatory cytokine release induced following peritoneal cells exposure to E. granulosus s.l. tegumental components. Importantly, prophylactic infusion of soluble CD5 or CD6 significantly ameliorated the infection outcome in the mouse model of secondary cystic echinococcosis. CONCLUSIONS/SIGNIFICANCE: Taken together, the results expand the pathogen binding properties of CD5 and CD6 and provide novel evidence for their therapeutic potential in human cystic echinococcosis.

Impact of Trypanosoma cruzi infection on nitric oxide synthase and arginase expression and activity in young and elderly mice.

Elderly organisms are more susceptible to infectious diseases. However, the impact of aging on antiparasitic mechanisms, especially the nitric oxide pathway, is poorly understood. Using an integrated in vivo and in vitro model, we compared the severity of Trypanosoma cruzi infection in young and elderly (8 or 72 weeks old) mice. Forty C57BL/6 mice were randomized into four groups: Y-inf, young infected; Yn-inf, young uninfected; A-inf, aged infected; An-inf, aged uninfected. Parasitemia was measured daily, and animals were euthanized after 15 days of infection. Trypanosoma cruzi-induced inflammatory processes were analyzed in blood and heart samples, as well as in bone marrow-derived macrophages (BMDMs) co-cultured with splenocytes isolated from young or elderly mice. Our results indicated upregulated IgG2b and IL-17 production in elderly animals, which was not sufficient to reduce parasitemia, parasitic load and myocarditis to levels observed in young animals. The higher susceptibility of elderly mice to T. cruzi infection was accompanied by reduced cardiac inducible nitric oxide synthase (iNOS) gene expression, nitric oxide (NO) and IFN-γ levels, as well as an antagonistic upregulation of arginase-1 expression and arginase activity. The same responses were observed when BMDMs co-cultured with splenocytes from elderly mice were stimulated with T. cruzi antigens. Our findings indicate that elderly mice were more susceptible to T. cruzi infection, which was potentially related to an attenuated response to antigenic stimulation, inhibition of iNOS gene expression and NO production, and antagonistic upregulation of arginase gene expression and activity, which created favorable conditions for heart parasitism and myocarditis development.

Anti-Tumor Necrosis Factor α Therapeutics Differentially Affect Leishmania Infection of Human Macrophages.

Tumor necrosis factor α (TNFα) drives the pathophysiology of human autoimmune diseases and consequently, neutralizing antibodies (Abs) or Ab-derived molecules directed against TNFα are essential therapeutics. As treatment with several TNFα blockers has been reported to entail a higher risk of infectious diseases such as leishmaniasis, we established an in vitro model based on Leishmania-infected human macrophages, co-cultured with autologous T-cells, for the analysis and comparison of anti-TNFα therapeutics. We demonstrate that neutralization of soluble TNFα (sTNFα) by the anti-TNFα Abs Humira®, Remicade®, and its biosimilar Remsima® negatively affects infection as treatment with these agents significantly reduces Leishmania-induced T-cell proliferation and increases the number of infected macrophages. By contrast, we show that blockade of sTNFα by Cimzia® does not affect T-cell proliferation and infection rates. Moreover, compared to Remicade®, treatment with Cimzia® does not impair the expression of cytolytic effector proteins in proliferating T-cells. Our data demonstrate that Cimzia® supports parasite control through its conjugated polyethylene glycol (PEG) moiety as PEGylation of Remicade® improves the clearance of intracellular Leishmania. This effect can be linked to complement activation, with levels of complement component C5a being increased upon treatment with Cimzia® or a PEGylated form of Remicade®. Taken together, we provide an in vitro model of human leishmaniasis that allows direct comparison of different anti-TNFα agents. Our results enhance the understanding of the efficacy and adverse effects of TNFα blockers and they contribute to evaluate anti-TNFα therapy for patients living in countries with a high prevalence of leishmaniasis.

Prolactin as immune cell regulator in Toxocara canis somatic larvae chronic infection.

Toxocariasis is a zoonotic disease produced by ingestion of larval Toxocara spp. eggs. Prolactin (PRL) has been considered to have an important role in Toxocara canis infection. Recent evidence has found that PRL directly can increase parasite growth and differentiation of T. canis The present study, evaluated the effect of high PRL levels on the immune system's response and parasites clearance in chronic infection. Our results showed that hyperprolactinemia did not affect the number of larvae recovered from several tissues in rats. Parasite-specific antibody production, showed no difference between the groups. Lung tissue presented eosinophilic granulomas typical of a chronic infection in all the experimental groups. Flow cytometry analysis was made in order to determine changes in the percentage of innate and adaptive immune cell subpopulations in the spleen, peripheric (PLN) and mesenteric (MLN) lymphatic nodes. The results showed a differential effect of PRL and infection on different immune compartments in the percent of total T cells, T helper cells, T cytotoxic cells, B cells, NK cells, and Tγδ cells. To our knowledge, for the first time it is demonstrated that PRL can have an immunomodulatory role during T. canis chronic infection in the murine host.

Extent and Dynamics of Polymorphism in the Malaria Vaccine Candidate Plasmodium falciparum Reticulocyte-Binding Protein Homologue-5 in Kalifabougou, Mali.

Reticulocyte-binding homologues (RH) are a ligand family that mediates merozoite invasion of erythrocytes in Plasmodium falciparum. Among the five members of this family identified so far, only P. falciparum reticulocyte-binding homologue-5 (PfRH5) has been found to be essential for parasite survival across strains that differ in virulence and route of host-cell invasion. Based on its essential role in invasion and early evidence of sequence conservation, PfRH5 has been prioritized for development as a vaccine candidate. However, little is known about the extent of genetic variability of RH5 in the field and the potential impact of such diversity on clinical outcomes or on vaccine evasion. Samples collected during a prospective cohort study of malaria incidence conducted in Kalifabougou, in southwestern Mali, were used to estimate genetic diversity, measure haplotype prevalence, and assess the within-host dynamics of PfRH5 variants over time and in relation to clinical malaria. A total of 10 nonsynonymous polymorphic sites were identified in the Pfrh5 gene, resulting in 13 haplotypes encoding unique protein variants. Four of these variants have not been previously observed. Plasmodium falciparum reticulocyte-binding homologue-5 had low amino acid haplotype (h = 0.58) and nucleotide (π = 0.00061) diversity. By contrast to other leading blood-stage malaria vaccine candidate antigens, amino acid differences were not associated with changes in the risk of febrile malaria in consecutive infections. Conserved B- and T-cell epitopes were identified. These results support the prioritization of PfRH5 for possible inclusion in a broadly cross-protective vaccine.

Histopathological and immunohistochemical characterisation of hepatic granulomas in Leishmania donovani-infected BALB/c mice: a time-course study.

BACKGROUND: Visceral leishmaniasis (VL) is a neglected tropical disease (NTD), caused by the intracellular protozoan parasites Leishmania donovani and Leishmania infantum. Symptomatic VL is considered fatal when left untreated. At present, there is no effective vaccine licensed for human use and available chemotherapies have limitations. Understanding the local immune mechanisms required for the control of infection is a key factor for developing effective vaccines and therapeutics. METHODS: We have investigated the development of the typical granulomatous lesions in the liver in experimental VL over time, together with the local immune responses. BALB/c mice were infected intravenously with a dose of 2 × 107 L. donovani amastigotes (MHOM/ET/67/HU3) and sacrificed at 15, 35 and 63 days post-infection (dpi). Histopathology and immunohistochemical techniques were used for the detection of Leishmania antigen, selected cell types including B and T lymphocytes, macrophages and neutrophils (CD45R-B220+, CD3+, F4/80+ and Ly-6G+) and iNOS. RESULTS: Granulomatous lesions were identified as early as 15 dpi in the livers of all infected animals. Three categories were used to classify liver granulomas (immature, mature and clear). Clear granulomas were exclusively detected from 35 dpi onwards. Kupffer cells (F4/80+) were predominant in immature granulomas, regardless of the dpi. Nonetheless, the highest expression was found 63 dpi. Positive staining for iNOS was mainly observed in the cytoplasm of fused Kupffer cells and the highest expression observed at 35 dpi. T cells (CD3+) and B cells (CD45R-B220+) were predominant in more advanced granuloma stages, probably related to the establishment of acquired immunity. Neutrophils (Ly-6G+) were predominantly observed in mature granulomas with the highest expression at 15 dpi. Neutrophils were lower in numbers compared to other cell types, particularly at later time points. CONCLUSIONS: Our results reflect the role of macrophages during the early stage of infection and the establishment of a lymphocytic response to control the infection in more advanced stages.

Acquired Protective Immunity in Atlantic Salmon Salmo salar against the Myxozoan Kudoa thyrsites Involves Induction of MHIIß+ CD83+ Antigen-Presenting Cells.

The histozoic myxozoan parasite Kudoa thyrsites causes postmortem myoliquefaction and is responsible for economic losses to salmon aquaculture in the Pacific Northwest. Despite its importance, little is known about the host-parasite relationship, including the host response to infection. The present work sought to characterize the immune response in Atlantic salmon during infection, recovery, and reexposure to K. thyrsites After exposure to infective seawater, infected and uninfected smolts were sampled three times over 4,275 degree-days. Histological analysis revealed infection severity decreased over time in exposed fish, while in controls there was no evidence of infection. Following a secondary exposure of all fish, severity of infection in the controls was similar to that measured in exposed fish at the first sampling time but was significantly reduced in reexposed fish, suggesting the acquisition of protective immunity. Using immunohistochemistry, we detected a population of MHIIß+ cells in infected muscle that followed a pattern of abundance concordant with parasite prevalence. Infiltration of these cells into infected myocytes preceded destruction of the plasmodium and dissemination of myxospores. Dual labeling indicated a majority of these cells were CD83+/MHIIß+ Using reverse transcription-quantitative PCR, we detected significant induction of cellular effectors, including macrophage/dendritic cells (mhii/cd83/mcsf), B cells (igm/igt), and cytotoxic T cells (cd8/nkl), in the musculature of infected fish. These data support a role for cellular effectors such as antigen-presenting cells (monocyte/macrophage and dendritic cells) along with B and T cells in the acquired protective immune response of Atlantic salmon against K. thyrsites.

Gamma Interferon Mediates Experimental Cerebral Malaria by Signaling within Both the Hematopoietic and Nonhematopoietic Compartments.

Experimental cerebral malaria (ECM) is a gamma interferon (IFN-γ)-dependent syndrome. However, whether IFN-γ promotes ECM through direct and synergistic targeting of multiple cell populations or by acting primarily on a specific responsive cell type is currently unknown. Here, using a panel of cell- and compartment-specific IFN-γ receptor 2 (IFN-γR2)-deficient mice, we show that IFN-γ causes ECM by signaling within both the hematopoietic and nonhematopoietic compartments. Mechanistically, hematopoietic and nonhematopoietic compartment-specific IFN-γR signaling exerts additive effects in orchestrating intracerebral inflammation, leading to the development of ECM. Surprisingly, mice with specific deletion of IFN-γR2 expression on myeloid cells, T cells, or neurons were completely susceptible to terminal ECM. Utilizing a reductionist in vitro system, we show that synergistic IFN-γ and tumor necrosis factor (TNF) stimulation promotes strong activation of brain blood vessel endothelial cells. Combined, our data show that within the hematopoietic compartment, IFN-γ causes ECM by acting redundantly or by targeting non-T cell or non-myeloid cell populations. Within the nonhematopoietic compartment, brain endothelial cells, but not neurons, may be the major target of IFN-γ leading to ECM development. Collectively, our data provide information on how IFN-γ mediates the development of cerebral pathology during malaria infection.

Implications of NOVA1 suppression within the microenvironment of gastric cancer: association with immune cell dysregulation.

BACKGROUND: The neuronal splicing factor neuro-oncological ventral antigen 1 (NOVA1) is enriched in normal fibroblasts. Stromal spindle cells such as fibroblasts are major components of tissue inflammation and tertiary lymphoid structures within the microenvironment that contribute to the survival and growth of cancer cells. In the present study, we investigated changes of NOVA1 expression in tertiary lymphoid structures in early and advanced gastric cancer microenvironments in terms of tumor progression and immune regulation. METHODS: Using immunohistochemistry, we analyzed NOVA1 expression in tumor cells, T cells, and stromal spindle cells as well as infiltrating densities of CD3+ T cells, forkhead box P3 positive (FOXP3+) regulatory T cells, CD68+ macrophages, CD163+ M2 macrophages, and myeloperoxidase-positive neutrophils in 396 surgically resected gastric cancer tissues. RESULTS: Suppressed NOVA1 expression in tumor cells, T cells, and stromal spindle cells was closely related to decreased infiltration of FOXP3+ regulatory T cells, increased infiltration of CD68+ macrophages and CD163+ M2 macrophages, more advanced tumor stage, and inferior overall survival rate. In addition, low infiltration of CD3+ T cells and FOXP3+ regulatory T cells and high infiltration of CD68+ macrophages were associated with inferior overall survival. Specifically, weak NOVA1 expression in tumor cells was independently related to more advanced tumor stage and inferior overall survival. CONCLUSIONS: NOVA1 suppression was frequently noted in the gastric cancer microenvironment, and attenuated NOVA1 expression in tumor cells was associated with tumor progression and poor prognosis. This finding seems to be related to immune dysfunction through changes in the immune cell composition of T cells and macrophages.