The inflammasome recognizes cytosolic microbial and host DNA and triggers an innate immune response.

The innate immune system recognizes nucleic acids during infection and tissue damage. Whereas viral RNA is detected by endosomal toll-like receptors (TLR3, TLR7, TLR8) and cytoplasmic RIG-I and MDA5, endosomal TLR9 and cytoplasmic DAI bind DNA, resulting in the activation of nuclear factor-kappaB and interferon regulatory factor transcription factors. However, viruses also trigger pro-inflammatory responses, which remain poorly defined. Here we show that internalized adenoviral DNA induces maturation of pro-interleukin-1beta in macrophages, which is dependent on NALP3 and ASC, components of the innate cytosolic molecular complex termed the inflammasome. Correspondingly, NALP3- and ASC-deficient mice display reduced innate inflammatory responses to adenovirus particles. Inflammasome activation also occurs as a result of transfected cytosolic bacterial, viral and mammalian (host) DNA, but in this case sensing is dependent on ASC but not NALP3. The DNA-sensing pro-inflammatory pathway functions independently of TLRs and interferon regulatory factors. Thus, in addition to viral and bacterial components or danger signals in general, inflammasomes sense potentially dangerous cytoplasmic DNA, strengthening their central role in innate immunity.

Stem cell-related "self-renewal" signature and high epidermal growth factor receptor expression associated with resistance to concomitant chemoradiotherapy in glioblastoma.

PURPOSE: Glioblastomas are notorious for resistance to therapy, which has been attributed to DNA-repair proficiency, a multitude of deregulated molecular pathways, and, more recently, to the particular biologic behavior of tumor stem-like cells. Here, we aimed to identify molecular profiles specific for treatment resistance to the current standard of care of concomitant chemoradiotherapy with the alkylating agent temozolomide. PATIENTS AND METHODS: Gene expression profiles of 80 glioblastomas were interrogated for associations with resistance to therapy. Patients were treated within clinical trials testing the addition of concomitant and adjuvant temozolomide to radiotherapy. RESULTS: An expression signature dominated by HOX genes, which comprises Prominin-1 (CD133), emerged as a predictor for poor survival in patients treated with concomitant chemoradiotherapy (n = 42; hazard ratio = 2.69; 95% CI, 1.38 to 5.26; P = .004). This association could be validated in an independent data set. Provocatively, the HOX cluster was reminiscent of a "self-renewal" signature (P = .008; Gene Set Enrichment Analysis) recently characterized in a mouse leukemia model. The HOX signature and EGFR expression were independent prognostic factors in multivariate analysis, adjusted for the O-6-methylguanine-DNA methyltransferase (MGMT) methylation status, a known predictive factor for benefit from temozolomide, and age. Better outcome was associated with gene clusters characterizing features of tumor-host interaction including tumor vascularization and cell adhesion, and innate immune response. CONCLUSION: This study provides first clinical evidence for the implication of a "glioma stem cell" or "self-renewal" phenotype in treatment resistance of glioblastoma. Biologic mechanisms identified here to be relevant for resistance will guide future targeted therapies and respective marker development for individualized treatment and patient selection.

Tuning the immune response of dendritic cells to surface-assembled poly(I:C) on microspheres through synergistic interactions between phagocytic and TLR3 signaling.

The artificial dsRNA polyriboinosinic acid-polyribocytidylic acid, poly(I:C), is a potent adjuvant candidate for vaccination, as it strongly drives cell-mediated immunity. However, because of its effects on non-immune bystander cells, poly(I:C) administration may bear danger for the development of autoimmune diseases. Thus poly(I:C) should be applied in the lowest dose possible. We investigated microspheres carrying surface-assembled poly(I:C) as a two-in-one adjuvant formulation to stimulate maturation of monocyte-derived dendritic cells (MoDCs). Negatively charged polystyrene microspheres were equipped with a poly(ethylene glycol) corona through electrostatically driven surface assembly of a library of polycationic poly(l-lysine)-graft-poly(ethylene glycol) copolymers, PLL-g-PEG. Stable surface assembly of poly(I:C) was achieved by incubation of polymer-coated microspheres in an aqueous poly(I:C) solution. Surface-assembled poly(I:C) exhibited a strongly enhanced efficacy to stimulate maturation of MoDCs by up to two orders of magnitude, as compared to free poly(I:C). Multiple phagocytosis events were the key factor to enhance the efficacy. The cytokine secretion pattern of MoDCs after exposure to surface-assembled poly(I:C) differed from that of free poly(I:C), while their ability to stimulate T cell proliferation was similar. Overall, phagocytic signaling plays an important role in defining the resulting immune response to such two-in-one adjuvant formulations.

Mutations in penicillin-binding protein (PBP) genes and in non-PBP genes during selection of penicillin-resistant Streptococcus gordonii.

Penicillin resistance in Streptococcus spp. involves multiple mutations in both penicillin-binding proteins (PBPs) and non-PBP genes. Here, we studied the development of penicillin resistance in the oral commensal Streptococcus gordonii. Cyclic exposure of bacteria to twofold-increasing penicillin concentrations selected for a progressive 250- to 500-fold MIC increase (from 0.008 to between 2 and 4 microg/ml). The major MIC increase (> or = 35-fold) was related to non-PBP mutations, whereas PBP mutations accounted only for a 4- to 8-fold additional increase. PBP mutations occurred in class B PBPs 2X and 2B, which carry a transpeptidase domain, but not in class A PBP 1A, 1B, or 2A, which carry an additional transglycosylase domain. Therefore, we tested whether inactivation of class A PBPs affected resistance development in spite of the absence of mutations. Deletion of PBP 1A or 2A profoundly slowed down resistance development but only moderately affected resistance in already highly resistant mutants (MIC = 2 to 4 microg/ml). Thus, class A PBPs might facilitate early development of resistance by stabilizing penicillin-altered peptidoglycan via transglycosylation, whereas they might be less indispensable in highly resistant mutants which have reestablished a penicillin-insensitive cell wall-building machinery. The contribution of PBP and non-PBP mutations alone could be individualized in DNA transformation. Both PBP and non-PBP mutations conferred some level of intrinsic resistance, but combining the mutations synergized them to ensure high-level resistance (> or = 2 microg/ml). The results underline the complexity of penicillin resistance development and suggest that inhibition of transglycosylase might be an as yet underestimated way to interfere with early resistance development.

Viral superantigen-induced negative selection of TCR transgenic CD4+ CD8+ thymocytes depends on activation, but not proliferation.

T-cell negative selection, a process by which intrathymic immunological tolerance is induced, involves the apoptosis-mediated clonal deletion of potentially autoreactive T cells. Although different experimental approaches suggest that this process is triggered as the result of activation-mediated cell death, the signal transduction pathways underlying this process is not fully understood. In the present report we have used an in vitro system to analyze the cell activation and proliferation requirements for the deletion of viral superantigen (SAg)-reactive Vbeta8.1 T-cell receptor (TCR) transgenic (TG) thymocytes. Our results indicate that in vitro negative selection of viral SAg-reactive CD4+ CD8+ thymocytes is dependent on thymocyte activation but does not require the proliferation of the negatively signaled thymocytes.

MyD88 and TLR9 dependent immune responses mediate resistance to Leishmania guyanensis infections, irrespective of Leishmania RNA virus burden.

Infections with Leishmania parasites of the Leishmania Viannia subgenus give rise to both localized cutaneous (CL), and metastatic leishmaniasis. Metastasizing disease forms including disseminated (DCL) and mutocutaneous (MCL) leishmaniasis result from parasitic dissemination and lesion formation at sites distal to infection and have increased inflammatory responses. The presence of Leishmania RNA virus (LRV) in L. guyanensis parasites contributes to the exacerbation of disease and impacts inflammatory responses via activation of TLR3 by the viral dsRNA. In this study we investigated other innate immune response adaptor protein modulators and demonstrated that both MyD88 and TLR9 played a crucial role in the development of Th1-dependent healing responses against L. guyanensis parasites regardless of their LRV status. The absence of MyD88- or TLR9-dependent signaling pathways resulted in increased Th2 associated cytokines (IL-4 and IL-13), which was correlated with low transcript levels of IL-12p40. The reliance of IL-12 was further confirmed in IL12AB-/- mice, which were completely susceptible to infection. Protection to L. guyanensis infection driven by MyD88- and TLR9-dependent immune responses arises independently to those induced due to high LRV burden within the parasites.

The number of Toll-like receptor 9-agonist motifs in the adenovirus genome correlates with induction of dendritic cell maturation by adenovirus immune complexes.

Adenovirus serotype 5 (Ad5) vectors and specific neutralizing antibodies (NAbs) generate immune complexes (ICs) which are potent inducers of dendritic cell (DC) maturation. Here we show that ICs generated with rare Ad vector serotypes, such as Ad26 and Ad35, which are lead candidates in HIV vaccine development, are poor inducers of DC maturation and that their potency in inducing DC maturation strongly correlated with the number of Toll-like receptor 9 (TLR9)-agonist motifs present in the Ad vector's genome. In addition, we showed that antihexon but not antifiber antibodies are responsible for the induction of Ad IC-mediated DC maturation.

Conditional manipulation of sex ratios by ant workers: A test of kin selection theory

Variable queen mating frequencies provide a unique opportunity to study the resolution of worker-queen conflict over sex ratio in social Hymenoptera, because the conflict is maximal in colonies headed by a singly mated queen and is weak or nonexistent in colonies headed by a multiply mated queen. In the wood ant Formica exsecta, workers in colonies with a singly mated queen, but not those in colonies with a multiply mated queen, altered the sex ratio of queen-laid eggs by eliminating males to preferentially raise queens. By this conditional response to queen mating frequency, workers enhance their inclusive fitness.

Community-wide plasmid gene mobilization and selection.

Plasmids have long been recognized as an important driver of DNA exchange and genetic innovation in prokaryotes. The success of plasmids has been attributed to their independent replication from the host's chromosome and their frequent self-transfer. It is thought that plasmids accumulate, rearrange and distribute nonessential genes, which may provide an advantage for host proliferation under selective conditions. In order to test this hypothesis independently of biases from culture selection, we study the plasmid metagenome from microbial communities in two activated sludge systems, one of which receives mostly household and the other chemical industry wastewater. We find that plasmids from activated sludge microbial communities carry among the largest proportion of unknown gene pools so far detected in metagenomic DNA, confirming their presumed role of DNA innovators. At a system level both plasmid metagenomes were dominated by functions associated with replication and transposition, and contained a wide variety of antibiotic and heavy metal resistances. Plasmid families were very different in the two metagenomes and grouped in deep-branching new families compared with known plasmid replicons. A number of abundant plasmid replicons could be completely assembled directly from the metagenome, providing insight in plasmid composition without culturing bias. Functionally, the two metagenomes strongly differed in several ways, including a greater abundance of genes for carbohydrate metabolism in the industrial and of general defense factors in the household activated sludge plasmid metagenome. This suggests that plasmids not only contribute to the adaptation of single individual prokaryotic species, but of the prokaryotic community as a whole under local selective conditions.

Cost-effectiveness analysis of immune-modulating nutritional support for gastrointestinal cancer patients.

BACKGROUND & AIM: Immune-modulating nutritional formula containing arginine, omega-3 fatty acids and nucleotides has been demonstrated to decrease complications and length of stay in surgical patients. This study aims at assessing the impact of immune-modulating formula on hospital costs in gastrointestinal cancer surgical patients in Switzerland. METHOD: Based on a previously published meta-analysis, the relative risks of overall and infectious complications with immune-modulating versus standard nutrition formula were computed. Swiss hospital costs of patients undergoing gastrointestinal cancer surgery were retrieved. A method was developed to compute the patients' severity level, not taking into account the complications from the surgery. Incremental costs of complications were computed for both treatment groups, and sensitivity analyses were carried out. RESULTS: Relative risk of complications with pre-, peri- and post-operative use of immune-modulating formula was 0.69 (95%CI 0.58-0.83), 0.62 (95%CI 0.53-0.73) and 0.73 (95%CI 0.35-0.96) respectively. The estimated average contribution of complications to the cost of stay was CHF 14,949 (euro10,901) per patient (95%CI 10,712-19,186), independently of case's severity. Based on this cost, immune-modulating nutritional support decreased costs of hospital stay by CHF 1638 to CHF 2488 per patient (euro1195-euro1814). Net hospital savings were present for baseline complications rates as low as 5%. CONCLUSION: Immune-modulating nutritional solution is a cost-saving intervention in gastrointestinal cancer patients. The additional cost of immune-modulating formula are more than offset by savings associated with decreased treatment of complications.

Immune response to MMTV infection.

Mouse mammary tumor virus (MMTV) has developed a strategy of exploitation of the immune response. It infects dendritic cells and B cells and requires this infection to establish an efficient chronic infection. This allows transmission of infection to the mammary gland, production in milk and infection of the next generation via lactation. The elaborate strategy developed by MMTV utilizes several key elements of the normal immune response. Starting with the infection and activation of dendritic cells and B cells leading to the expression of a viral superantigen followed by professional superantigen-mediated priming of naive polyclonal T cells by dendritic cells and induction of superantigen-mediated T cell B cell collaboration results in long-lasting germinal center formation and production of long-lived B cells that can later carry the virus to the mammary gland epithelium. Later in life it can induce transformation of mammary gland epithelium by integrating close to proto-oncogenes leading to their overexpression. Genes encoding proteins of the Wnt-pathway are preferential targets. This review will put these effects in the context of a normal immune response and summarize important facts on MMTV biology.

Microbial influences on epithelial integrity and immune function as a basis for inflammatory diseases.

Certain autoimmune diseases as well as asthma have increased in recent decades, particularly in developed countries. The hygiene hypothesis has been the prevailing model to account for this increase; however, epidemiology studies also support the contribution of diet and obesity to inflammatory diseases. Diet affects the composition of the gut microbiota, and recent studies have identified various molecules and mechanisms that connect diet, the gut microbiota, and immune responses. Herein, we discuss the effects of microbial metabolites, such as short chain fatty acids, on epithelial integrity as well as immune cell function. We propose that dysbiosis contributes to compromised epithelial integrity and disrupted immune tolerance. In addition, dietary molecules affect the function of immune cells directly, particularly through lipid G-protein coupled receptors such as GPR43.

Antibody-mediated and cellular immune responses induced in naive volunteers by vaccination with long synthetic peptides derived from the Plasmodium vivax circumsporozoite protein.

Plasmodium vivax circumsporozoite (CS) protein is a leading malaria vaccine candidate. We describe the characterization of specific immune responses induced in 21 malaria-naive volunteers vaccinated with long synthetic peptides derived from the CS protein formulated in Montanide ISA 720. Both antibody- and cell-mediated immune responses were analyzed. Antibodies were predominantly of IgG1 and IgG3 isotypes, recognized parasite proteins on the immunofluorescent antibody test, and partially blocked sporozoite invasion of hepatoma cell lines in vitro. Peripheral blood mononuclear cells from most volunteers (94%) showed IFN-γ production in vitro upon stimulation with both long signal peptide and short peptides containing CD8+ T-cell epitopes. The relatively limited sample size did not allow conclusions about HLA associations with the immune responses observed. In summary, the inherent safety and tolerability together with strong antibody responses, invasion blocking activity, and the IFN-γ production induced by these vaccine candidates warrants further testing in a phase II clinical trial.