Severe respiratory syncytial virus (RSV) infection is a major cause of morbidity and mortality in infants <2 years-old. Here we describe that high-fiber diet protects mice from RSV infection. This effect was dependent on intestinal microbiota and production of acetate. Oral administration of acetate mediated interferon-β (IFN-β) response by increasing expression of interferon-stimulated genes in the lung. These effects were associated with reduction of viral load and pulmonary inflammation in RSV-infected mice. Type 1 IFN signaling via the IFN-1 receptor (IFNAR) was essential for acetate antiviral activity in pulmonary epithelial cell lines and for the acetate protective effect in RSV-infected mice. Activation of Gpr43 in pulmonary epithelial cells reduced virus-induced cytotoxicity and promoted antiviral effects through IFN-β response. The effect of acetate on RSV infection was abolished in Gpr43^−/− mice. Our findings reveal antiviral effects of acetate involving IFN-β in lung epithelial cells and engagement of GPR43 and IFNAR.

GPIs isolated from Toxoplasma gondii, as well as a chemically synthesized GPI lacking the lipid moiety, activated a reporter gene in Chinese hamster ovary cells expressing TLR4, while the core glycan and lipid moieties cleaved from the GPIs activated both TLR4- and TLR2-expressing cells. MyD88, but not TLR2, TLR4, or CD14, is absolutely needed to trigger TNF-α production by macrophages exposed to T. gondii GPIs. Importantly, TNF-α response to GPIs was completely abrogated in macrophages from TLR2/4-double-deficient mice. MyD88^−/− mice were more susceptible to death than wild-type (WT), TLR2^−/−, TLR4^−/−, TLR2/4^−/−, and CD14^−/− mice infected with the ME-49 strain of T. gondii. The cyst number was higher in the brain of TLR2/4^−/−, but not TLR2^−/−, TLR4^−/−, and CD14^−/−, mice, as compared with WT mice. Upon infection with the ME-49 strain of T. gondii, we observed no decrease of IL-12 and IFN-γ production in TLR2-, TLR4-, or CD14-deficient mice. Indeed, splenocytes from T. gondii-infected TLR2^−/− and TLR2/4^−/− mice produced more IFN-γ than cells from WT mice in response to in vitro stimulation with parasite extracts enriched in GPI-linked surface proteins. Together, our results suggest that both TLR2 and TLR4 receptors may participate in the host defense against T. gondii infection through their activation by the GPIs and could work together with other MyD88-dependent receptors, like other TLRs or even IL-18R or IL-1R, to obtain an effective host response against T. gondii infection.

Recognition of viruses by pattern recognition receptors (PRRs) causes interferon-β (IFN-β) induction, a key event in the anti-viral innate immune response, and also a target of viral immune evasion. Here the vaccinia virus (VACV) protein C6 is identified as an inhibitor of PRR-induced IFN-β expression by a functional screen of select VACV open reading frames expressed individually in mammalian cells. C6 is a member of a family of Bcl-2-like poxvirus proteins, many of which have been shown to inhibit innate immune signalling pathways. PRRs activate both NF-κB and IFN regulatory factors (IRFs) to activate the IFN-β promoter induction. Data presented here show that C6 inhibits IRF3 activation and translocation into the nucleus, but does not inhibit NF-κB activation. C6 inhibits IRF3 and IRF7 activation downstream of the kinases TANK binding kinase 1 (TBK1) and IκB kinase-ε (IKKε), which phosphorylate and activate these IRFs. However, C6 does not inhibit TBK1- and IKKε-independent IRF7 activation or the induction of promoters by constitutively active forms of IRF3 or IRF7, indicating that C6 acts at the level of the TBK1/IKKε complex. Consistent with this notion, C6 immunoprecipitated with the TBK1 complex scaffold proteins TANK, SINTBAD and NAP1. C6 is expressed early during infection and is present in both nucleus and cytoplasm. Mutant viruses in which the C6L gene is deleted, or mutated so that the C6 protein is not expressed, replicated normally in cell culture but were attenuated in two in vivo models of infection compared to wild type and revertant controls. Thus C6 contributes to VACV virulence and might do so via the inhibition of PRR-induced activation of IRF3 and IRF7. A key event in the innate immune response to virus infection is the detection of pathogen-associated molecular patterns (PAMPs) such as viral DNA and RNA by cellular pattern recognition receptors (PRRs). This leads to expression of interferon-β (IFN-β) by an infected cell. Many viruses have evolved mechanisms to evade the induction of IFN-β. Here a screen of poorly characterized vaccinia virus (VACV) proteins identified protein C6 as an inhibitor of IFN-β induction by PRRs. Data presented show that C6 prevents the activation of the transcription factors IRF3 and IRF7 by the kinases TBK1 and IKKε, which are key components at the point of convergence of several PRR signalling pathways. C6 interacts with the scaffold proteins NAP1, TANK and SINTBAD, which are components of the protein complexes containing TBK1 and IKKε, and this interaction might modulate the activity of these kinases. C6 is expressed early during infection and contributes to virulence because viruses that do not express C6 are attenuated in two in vivo models compared to wild type and revertant control viruses.

Tissue-resident macrophages are the most abundant immune cell population in healthy adipose tissue. Adipose tissue macrophages (ATMs) change during metabolic stress and are thought to contribute to metabolic syndrome. Here, we studied ATM subpopulations in steady state and in response to nutritional and infectious challenges. We found that tissue-resident macrophages from healthy epididymal white adipose tissue (eWAT) tightly associate with blood vessels, displaying very high endocytic capacity. We refer to these cells as vasculature-associated ATMs (VAMs). Chronic high-fat diet (HFD) results in the accumulation of a monocyte-derived CD11c⁺CD64⁺ double-positive (DP) macrophage eWAT population with a predominant anti-inflammatory/detoxifying gene profile, but reduced endocytic function. In contrast, fasting rapidly and reversibly leads to VAM depletion, while acute inflammatory stress induced by pathogens transiently depletes VAMs and simultaneously boosts DP macrophage accumulation. Our results indicate that ATM populations dynamically adapt to metabolic stress and inflammation, suggesting an important role for these cells in maintaining tissue homeostasis.

Drug‐induced liver injury (DILI) is an important cause of acute liver failure, with limited therapeutic options. During DILI, oncotic necrosis with concomitant release and recognition of intracellular content amplifies liver inflammation and injury. Among these molecules, self‐DNA has been widely shown to trigger inflammatory and autoimmune diseases; however, whether DNA released from damaged hepatocytes accumulates into necrotic liver and the impact of its recognition by the immune system remains elusive. Here we show that treatment with two different hepatotoxic compounds (acetaminophen and thioacetamide) caused DNA release into the hepatocyte cytoplasm, which occurred in parallel with cell deathin vitro. Administration of these compoundsin vivocaused massive DNA deposition within liver necrotic areas, together with an intravascular DNA coating. Using confocal intravital microscopy, we revealed that liver injury due to acetaminophen overdose led to a directional migration of neutrophils to DNA‐rich areas, where they exhibit an active patrolling behavior. DNA removal by intravenous DNASE1 injection or ablation of Toll‐like receptor 9 (TLR9)‐mediated sensing significantly reduced systemic inflammation, liver neutrophil recruitment, and hepatotoxicity. Analysis of liver leukocytes by flow cytometry revealed that emigrated neutrophils up‐regulated TLR9 expression during acetaminophen‐mediated necrosis, and these cells sensed and reacted to extracellular DNA by activating the TLR9/NF‐κB pathway. Likewise, adoptive transfer of wild‐type neutrophils to TLR9^−/−mice reversed the hepatoprotective phenotype otherwise observed in TLR9 absence.Conclusion: Hepatic DNA accumulation is a novel feature of DILI pathogenesis. Blockage of DNA recognition by the innate immune system may constitute a promising therapeutic venue. (Hepatology2015;61:348–360)

Interferon-induced proteins with tetratricopeptide repeats (IFITs) are highly expressed during the cell-intrinsic immune response to viral infection. IFIT1 inhibits translation by binding directly to the 5′ end of foreign RNAs, particularly those with non-self cap structures, precluding the recruitment of the cap-binding eukaryotic translation initiation factor 4F and ribosome recruitment. The presence of IFIT1 imposes a requirement on viruses that replicate in the cytoplasm to maintain mechanisms to avoid its restrictive effects. Interaction of different IFIT family members is well described, but little is known of the molecular basis of IFIT association or its impact on function. Here, we reconstituted different complexes of IFIT1, IFIT2 and IFIT3 in vitro, which enabled us to reveal critical aspects of IFIT complex assembly. IFIT1 and IFIT3 interact via a YxxxL motif present in the C-terminus of each protein. IFIT2 and IFIT3 homodimers dissociate to form a more stable heterodimer that also associates with IFIT1. We show for the first time that IFIT3 stabilizes IFIT1 protein expression, promotes IFIT1 binding to a cap0 Zika virus reporter mRNA and enhances IFIT1 translation inhibition. This work reveals molecular aspects of IFIT interaction and provides an important missing link between IFIT assembly and function.