Publisher Correction: Tumor-necrosis factor impairs CD4+ T cell-mediated immunological control in chronic viral infection.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Tumor-necrosis factor impairs CD4(+) T cell-mediated immunological control in chronic viral infection.

Persistent viral infections are characterized by the simultaneous presence of chronic inflammation and T cell dysfunction. In prototypic models of chronicity--infection with human immunodeficiency virus (HIV) or lymphocytic choriomeningitis virus (LCMV)--we used transcriptome-based modeling to reveal that CD4(+) T cells were co-exposed not only to multiple inhibitory signals but also to tumor-necrosis factor (TNF). Blockade of TNF during chronic infection with LCMV abrogated the inhibitory gene-expression signature in CD4(+) T cells, including reduced expression of the inhibitory receptor PD-1, and reconstituted virus-specific immunity, which led to control of infection. Preventing signaling via the TNF receptor selectively in T cells sufficed to induce these effects. Targeted immunological interventions to disrupt the TNF-mediated link between chronic inflammation and T cell dysfunction might therefore lead to therapies to overcome persistent viral infection.

On-surface synthesis of graphene nanoribbons with zigzag edge topology.

Graphene-based nanostructures exhibit electronic properties that are not present in extended graphene. For example, quantum confinement in carbon nanotubes and armchair graphene nanoribbons leads to the opening of substantial electronic bandgaps that are directly linked to their structural boundary conditions. Nanostructures with zigzag edges are expected to host spin-polarized electronic edge states and can thus serve as key elements for graphene-based spintronics. The edge states of zigzag graphene nanoribbons (ZGNRs) are predicted to couple ferromagnetically along the edge and antiferromagnetically between the edges, but direct observation of spin-polarized edge states for zigzag edge topologies--including ZGNRs--has not yet been achieved owing to the limited precision of current top-down approaches. Here we describe the bottom-up synthesis of ZGNRs through surface-assisted polymerization and cyclodehydrogenation of specifically designed precursor monomers to yield atomically precise zigzag edges. Using scanning tunnelling spectroscopy we show the existence of edge-localized states with large energy splittings. We expect that the availability of ZGNRs will enable the characterization of their predicted spin-related properties, such as spin confinement and filtering, and will ultimately add the spin degree of freedom to graphene-based circuitry.

NK cell-intrinsic FcεRIγ limits CD8+ T-cell expansion and thereby turns an acute into a chronic viral infection.

During viral infection, tight regulation of CD8+ T-cell functions determines the outcome of the disease. Recently, others and we determined that the natural killer (NK) cells kill hyperproliferative CD8+ T cells in the context of viral infection, but molecules that are involved in shaping the regulatory capability of NK cells remain virtually unknown. Here we used mice lacking the Fc-receptor common gamma chain (FcRγ, FcεRIγ, Fcer1g-/- mice) to determine the role of Fc-receptor and NK-receptor signaling in the process of CD8+ T-cell regulation. We found that the lack of FcRγ on NK cells limits their ability to restrain virus-specific CD8+ T cells and that the lack of FcRγ in Fcer1g-/- mice leads to enhanced CD8+ T-cell responses and rapid control of the chronic docile strain of the lymphocytic choriomeningitis virus (LCMV). Mechanistically, FcRγ stabilized the expression of NKp46 but not that of other killer cell-activating receptors on NK cells. Although FcRγ did not influence the development or activation of NK cell during LCMV infection, it specifically limited their ability to modulate CD8+ T-cell functions. In conclusion, we determined that FcRγ plays an important role in regulating CD8+ T-cell functions during chronic LCMV infection.

Fragile X mental retardation protein protects against tumour necrosis factor-mediated cell death and liver injury.

OBJECTIVE: The Fragile X mental retardation (FMR) syndrome is a frequently inherited intellectual disability caused by decreased or absent expression of the FMR protein (FMRP). Lack of FMRP is associated with neuronal degradation and cognitive dysfunction but its role outside the central nervous system is insufficiently studied. Here, we identify a role of FMRP in liver disease. DESIGN: Mice lacking Fmr1 gene expression were used to study the role of FMRP during tumour necrosis factor (TNF)-induced liver damage in disease model systems. Liver damage and mechanistic studies were performed using real-time PCR, Western Blot, staining of tissue sections and clinical chemistry. RESULTS: Fmr1null mice exhibited increased liver damage during virus-mediated hepatitis following infection with the lymphocytic choriomeningitis virus. Exposure to TNF resulted in severe liver damage due to increased hepatocyte cell death. Consistently, we found increased caspase-8 and caspase-3 activation following TNF stimulation. Furthermore, we demonstrate FMRP to be critically important for regulating key molecules in TNF receptor 1 (TNFR1)-dependent apoptosis and necroptosis including CYLD, c-FLIPS and JNK, which contribute to prolonged RIPK1 expression. Accordingly, the RIPK1 inhibitor Necrostatin-1s could reduce liver cell death and alleviate liver damage in Fmr1null mice following TNF exposure. Consistently, FMRP-deficient mice developed increased pathology during acute cholestasis following bile duct ligation, which coincided with increased hepatic expression of RIPK1, RIPK3 and phosphorylation of MLKL. CONCLUSIONS: We show that FMRP plays a central role in the inhibition of TNF-mediated cell death during infection and liver disease.

A dual role for hepatocyte-intrinsic canonical NF-κB signaling in virus control.

BACKGROUND & AIMS: Hepatic innate immune control of viral infections has largely been attributed to Kupffer cells, the liver-resident macrophages. However, hepatocytes, the parenchymal cells of the liver, also possess potent immunological functions in addition to their known metabolic functions. Owing to their abundance in the liver and known immunological functions, we aimed to investigate the direct antiviral mechanisms employed by hepatocytes. METHODS: Using lymphocytic choriomeningitis virus (LCMV) as a model of liver infection, we first assessed the role of myeloid cells by depletion prior to infection. We investigated the role of hepatocyte-intrinsic innate immune signaling by infecting mice lacking canonical NF-κB signaling (IkkßΔHep) specifically in hepatocytes. In addition, mice lacking hepatocyte-specific interferon-α/ß signaling-(IfnarΔHep), or interferon-α/ß signaling in myeloid cells-(IfnarΔMyel) were infected. RESULTS: Here, we demonstrate that LCMV activates NF-κB signaling in hepatocytes. LCMV-triggered NF-κB activation in hepatocytes did not depend on Kupffer cells or TNFR1 signaling but rather on Toll-like receptor signaling. LCMV-infected IkkßΔHep livers displayed strongly elevated viral titers due to LCMV accumulation within hepatocytes, reduced interferon-stimulated gene (ISG) expression, delayed intrahepatic immune cell influx and delayed intrahepatic LCMV-specific CD8+ T cell responses. Notably, viral clearance and ISG expression were also reduced in LCMV-infected primary hepatocytes lacking IKKß, demonstrating a hepatocyte-intrinsic effect. Similar to livers of IkkßΔHep mice, enhanced hepatocytic LCMV accumulation was observed in livers of IfnarΔHep mice, whereas IfnarΔMyel mice were able to control LCMV infection. Hepatocytic NF-κB signaling was also required for efficient ISG induction in HDV-infected dHepaRG cells and interferon-α/ß-mediated inhibition of HBV replication in vitro. CONCLUSIONS: Together, these data show that hepatocyte-intrinsic NF-κB is a vital amplifier of interferon-α/ß signaling, which is pivotal for strong early ISG responses, immune cell infiltration and hepatic viral clearance. LAY SUMMARY: Innate immune cells have been ascribed a primary role in controlling viral clearance upon hepatic infections. We identified a novel dual role for NF-κB signaling in infected hepatocytes which was crucial for maximizing interferon responses and initiating adaptive immunity, thereby efficiently controlling hepatic virus replication.

Tumor Necrosis Factor-Mediated Survival of CD169+ Cells Promotes Immune Activation during Vesicular Stomatitis Virus Infection.

Innate immune activation is essential to mount an effective antiviral response and to prime adaptive immunity. Although a crucial role of CD169+ cells during vesicular stomatitis virus (VSV) infections is increasingly recognized, factors regulating CD169+ cells during viral infections remain unclear. Here, we show that tumor necrosis factor is produced by CD11b+ Ly6C+ Ly6G+ cells following infection with VSV. The absence of TNF or TNF receptor 1 (TNFR1) resulted in reduced numbers of CD169+ cells and in reduced type I interferon (IFN-I) production during VSV infection, with a severe disease outcome. Specifically, TNF triggered RelA translocation into the nuclei of CD169+ cells; this translocation was inhibited when the paracaspase MALT-1 was absent. Consequently, MALT1 deficiency resulted in reduced VSV replication, defective innate immune activation, and development of severe disease. These findings indicate that TNF mediates the maintenance of CD169+ cells and innate and adaptive immune activation during VSV infection.IMPORTANCE Over the last decade, strategically placed CD169+ metallophilic macrophages in the marginal zone of the murine spleen and lymph nodes (LN) have been shown to play a very important role in host defense against viral pathogens. CD169+ macrophages have been shown to activate innate and adaptive immunity via "enforced virus replication," a controlled amplification of virus particles. However, the factors regulating the CD169+ macrophages remain to be studied. In this paper, we show that after vesicular stomatitis virus infection, phagocytes produce tumor necrosis factor (TNF), which signals via TNFR1, and promote enforced virus replication in CD169+ macrophages. Consequently, lack of TNF or TNFR1 resulted in defective immune activation and VSV clearance.

B Cell-Mediated Maintenance of Cluster of Differentiation 169-Positive Cells Is Critical for Liver Regeneration.

The liver has an extraordinary capacity to regenerate through activation of key molecular pathways. However, central regulators controlling liver regeneration remain insufficiently studied. Here, we show that B cell-deficient animals failed to induce sufficient liver regeneration after partial hepatectomy (PHx). Consistently, adoptive transfer of B cells could rescue defective liver regeneration. B cell-mediated lymphotoxin beta production promoted recovery from PHx. Absence of B cells coincided with loss of splenic cluster of differentiation 169-positive (CD169+ ) macrophages. Moreover, depletion of CD169+ cells resulted in defective liver regeneration and decreased survival, which was associated with reduced hepatocyte proliferation. Mechanistically, CD169+ cells contributed to liver regeneration by inducing hepatic interleukin-6 (IL-6) production and signal transducer and activator of transcription 3 activation. Accordingly, treatment of CD169+ cell-depleted animals with IL-6/IL-6 receptor rescued liver regeneration and severe pathology following PHx. Conclusion: We identified CD169+ cells to be a central trigger for liver regeneration, by inducing key signaling pathways important for liver regeneration.

Imperatorin ameliorates lipopolysaccharide induced memory deficit by mitigating proinflammatory cytokines, oxidative stress and modulating brain-derived neurotropic factor.

BACKGROUND: Lipopolysaccharide (LPS), an endotoxin from the outer membrane of Gram negative bacteria has been reported to cause neuroinflammation and learning and memory deficits. There are reports describing the beneficial effects of Imperatorin (IMP), a naturally occurring furanocoumarin in central nervous system (CNS) disorders such as anxiety and epilepsy. OBJECTIVE: In the current study, we investigated whether IMP protects against LPS mediated memory deficits and neuroinflammation. METHODS: Mice pretreated with IMP (5, 10 mg/kg po) were administered LPS (250 µg/kg ip) for 7 days. Memory was evaluated in the Morris water maze (MWM) and Y maze. The mice were euthanised and different biochemical assessments were carried out to measure oxidative stress and acetyl choline esterase (AChE). Further, evaluation of proinflammatory cytokines such as tumor necrosis factor (TNF-α) and interleukin-6 (IL-6) levels and brain derived neurotrophic factor (BDNF) in hippocampus and cortex of brain were performed. RESULTS: LPS administration caused poor memory retention in both, MWM and Y maze, and caused distinct oxidative stress since decrease in superoxide dismutase (SOD), reduced glutathione (GSH) levels and increased lipid peroxidation were observed. Also, a significant rise was observed in the levels of AChE. Moreover, a rise in TNF-α and IL-6 levels and depleted levels of BDNF were noted. IMP pretreatment reversed LPS induced behavioral and memory disturbances and significantly decreased the oxidative stress and AChE levels. It also reduced TNF-α and IL-6 levels and caused a significant upregulation of BDNF levels. CONCLUSION: Present study highlights the potential neuroprotective role of IMP against LPS mediated cognitive impairment and neuroinflammation.

T-cell STAT3 is required for the maintenance of humoral immunity to LCMV.

STAT3 is a critical transcription factor activated downstream of cytokine signaling and is integral for the function of multiple immune cell types. Human mutations in STAT3 cause primary immunodeficiency resulting in impaired control of a variety of infections, including reactivation of latent viruses. In this study, we investigate how T-cell functions of STAT3 contribute to responses to viral infection by inducing chronic lymphocytic choriomeningitis virus (LCMV) infection in mice lacking STAT3 specifically in T cells. Although mice with conditional disruption of STAT3 in T cells were able to mount early responses to viral infection similar to control animals, including expansion of effector T cells, we found generation of T-follicular helper (Tfh) cells to be impaired. As a result, STAT3 T cell deficient mice produced attenuated germinal center reactions, and did not accumulate bone marrow virus specific IgG-secreting cells, resulting in failure to maintain levels of virus-specific IgG or mount neutralizing responses to LCMV in the serum. These effects were associated with reduced control of viral replication and prolonged infection. Our results demonstrate the importance of STAT3 in T cells for the generation of functional long-term humoral immunity to viral infections.

Storage of Erythrocytes Induces Suicidal Erythrocyte Death.

BACKGROUND/AIMS: Similar to apoptosis of nucleated cells, red blood cells (RBC) can undergo suicidal cell death - called eryptosis. It is characterized by cell shrinkage and phosphatidylserine translocation. Eryptosis is triggered by an increase of intracellular calcium concentration due to activation of nonselective cation channels. The cation channels and consequently eryptosis are inhibited by erythropoietin. Eryptotic RBC are engulfed by macrophages and thus rapidly cleared from circulating blood. In this study, we explored whether storage of RBC influences the rate of eryptosis. METHODS: Flow cytometry was employed to quantify phosphatidylserine exposing erythrocytes from annexin V binding and cytosolic Ca2+ activity from Fluo-3 fluorescence. Clearance of stored murine RBC was tested by injection of carboxyfluorescein succinimidyl ester (CFSE)-labelled erythrocytes. RESULTS: Storage for 42 days significantly increased the percentage of phosphatidylserine exposing and haemolytic erythrocytes, an effect blunted by removal of extracellular calcium. Phosphatidylserine exposure could be inhibited by addition of erythropoietin. Upon transfusion, the clearance of murine CFSE-labelled RBC from circulating blood was significantly higher following storage for 10 days when compared to 2 days of storage. CONCLUSION: Storage of RBC triggers eryptosis by Ca2+ and erythropoietin sensitive mechanisms.

Deficiency of the B cell-activating factor receptor results in limited CD169+ macrophage function during viral infection.

UNLABELLED: The B cell-activating factor (BAFF) is critical for B cell development and humoral immunity in mice and humans. While the role of BAFF in B cells has been widely described, its role in innate immunity remains unknown. Using BAFF receptor (BAFFR)-deficient mice, we characterized BAFFR-related innate and adaptive immune functions following infection with vesicular stomatitis virus (VSV) and lymphocytic choriomeningitis virus (LCMV). We identified a critical role for BAFFR signaling in the generation and maintenance of the CD169(+) macrophage compartment. Consequently, Baffr(-) (/) (-) mice exhibited limited induction of innate type I interferon production after viral infection. Lack of BAFFR signaling reduced virus amplification and presentation following viral infection, resulting in highly reduced antiviral adaptive immune responses. As a consequence, BAFFR-deficient mice showed exacerbated and fatal disease after viral infection. Mechanistically, transient lack of B cells in Baffr(-) (/) (-) animals resulted in limited lymphotoxin expression, which is critical for maintenance of CD169(+) cells. In conclusion, BAFFR signaling affects both innate and adaptive immune activation during viral infections. IMPORTANCE: Viruses cause acute and chronic infections in humans resulting in millions of deaths every year. Innate immunity is critical for the outcome of a viral infection. Innate type I interferon production can limit viral replication, while adaptive immune priming by innate immune cells induces pathogen-specific immunity with long-term protection. Here, we show that BAFFR deficiency not only perturbed B cells, but also resulted in limited CD169(+) macrophages. These macrophages are critical in amplifying viral particles to trigger type I interferon production and initiate adaptive immune priming. Consequently, BAFFR deficiency resulted in reduced enforced viral replication, limited type I interferon production, and reduced adaptive immunity compared to BAFFR-competent controls. As a result, BAFFR-deficient mice were predisposed to fatal viral infections. Thus, BAFFR expression is critical for innate immune activation and antiviral immunity.

Water Formation for the Metalation of Porphyrin Molecules on Oxidized Cu(111).

Herein the formation of water molecules in the intermediate step of the redox reaction of porphyrins self-metalation on O/Cu(111) is demonstrated. Photoemission measurements show that the temperature on which porphyrins pick-up a substrate metal atom on O/Cu(111) is reduced by about 185±15 K with respect to the pure Cu(111). DFT calculations clearly indicate that the formation of a water molecule is less expensive than the formation of H2 on the O/Cu(111) substrate and, in some cases, it can be also exothermic.

Electrochemical low valent cobalt-catalyzed addition of aryl and vinyl chlorides to α-ketoamides via C-Cl bond activation.

The development of an electrochemical cobalt catalyzed C-Cl bond activation at room temperature for the nucleophilic addition of aryl and vinyl chlorides to α-ketoamides is described. The overall method operates through an electrochemically induced low valent cobalt catalyst that oxidatively adds to aryl or vinyl chlorides affording medicinally important 3-hydroxy oxindole and 3-hydroxypyrrolidinone scaffolds. The development of an enantioselective version using a chiral pyrox ligand is also demonstrated.

Serologic evidence of avian influenza H9N2 and paramyxovirus type 1 infection in emus (Dromaius novaehollandiae) in India.

An avian influenza (AI) surveillance was undertaken in Maharashtra state, India during the period 2010-2011. There are no reports of AI surveillance in emus from India. A total of 202 blood samples and 467 tracheal and cloacal swabs were collected from eight emu farms. A hemagglutination inhibition (HI) assay was performed for detection of antibodies against AI H5N1, H7N1, H9N2, and avian paramyxovirus type 1 (APMV-1) viruses. A microneutralization (MN) assay was performed to confirm the presence of neutralizing antibodies against AI H9N2 and to compare with HI assays. A total of 28.2% and 28.7% of samples were positive for antibodies against AI H9N2 by HI and MN assays, respectively, using > or = 1:40 as a cut-off titer; 15.3% samples were positive for APMV-1 by HI assay using a > or = 1:10 cut-off titer. Seropositivity of AI H9N2 was nil in the grower (<1 yr) age group and highest (78%) in the breeder (2-3 yr) age group, whereas seropositivity against APMV-1 was observed in all age groups. Performance of both HI and MN assays was similar, suggesting the utility of using the MN assay along with HI assay for surveillance studies. This is the first report of the seroprevalence of AI H9N2 and APMV-1 in emus in India.

Synthesis and antibacterial activity of a new series of 3-[3-(substituted phenyl)-1-phenyl-1H-pyrazol-5-yl]-2H-chromen-2-one derivatives.

A series of 3-[3-(substituted phenyl)-1-phenyl-1H-pyrazol-5-yl]-2H-chromen-2-one (4a-k) were synthesized by reaction of 3-[2,3-dibromo-3-(substituted phenyl)propanoyl]-2H-chromen-2-one (3 a-k) with phenyl hydrazine in presence of triethylamine in absolute ethanol, characterized by spectral data and screened for their in vitro antibacterial activity against gram-positive and gram-negative bacteria. Among the series, compounds 4d, 4h and 4i displayed an encouraging antibacterial activity profile as compared to reference standard drug ciprofloxacin against tested bacterial strains.

Paired Electrolysis for Decarboxylative Cyanation: 4-CN-Pyridine, a Versatile Nitrile Source.

A decarboxylative cyanation of amino acids under paired electrochemical reaction conditions has been developed. 4-CN-pyridine was found to be a new and effective cyanation reagent under catalyst-free conditions. Mechanistic studies support a nucleophilic reaction pathway, and the cyanation protocol can be applied to diverse substrates including N,N-dialkyl aniline and indole derivatives.

Progress on Thin Film Freezing Technology for Dry Powder Inhalation Formulations.

The surface drying process is an important technology in the pharmaceutical, biomedical, and food industries. The final stage of formulation development (i.e., the drying process) faces several challenges, and overall mastering depends on the end step. The advent of new emerging technologies paved the way for commercialization. Thin film freezing (TFF) is a new emerging freeze-drying technique available for various treatment modalities in drug delivery. TFF has now been used for the commercialization of pharmaceuticals, food, and biopharmaceutical products. The present review highlights the fundamentals of TFF along with modulated techniques used for drying pharmaceuticals and biopharmaceuticals. Furthermore, we have covered various therapeutic applications of TFF technology in the development of nanoformulations, dry powder for inhalations and vaccines. TFF holds promise in delivering therapeutics for lung diseases such as fungal infection, bacterial infection, lung dysfunction, and pneumonia.

BAFF Attenuates Immunosuppressive Monocytes in the Melanoma Tumor Microenvironment.

Emerging evidence indicates B-cell activating factor (BAFF, Tnfsf13b) to be an important cytokine for antitumor immunity. In this study, we generated a BAFF-overexpressing B16.F10 melanoma cell model and found that BAFF-expressing tumors grow more slowly in vivo than control tumors. The tumor microenvironment (TME) of BAFF-overexpressing tumors had decreased myeloid infiltrates with lower PD-L1 expression. Monocyte depletion and anti-PD-L1 antibody treatment confirmed the functional importance of monocytes for the phenotype of BAFF-mediated tumor growth delay. RNA sequencing analysis confirmed that monocytes isolated from BAFF-overexpressing tumors were characterized by a less exhaustive phenotype and were enriched for in genes involved in activating adaptive immune responses and NF-κB signaling. Evaluation of patients with late-stage metastatic melanoma treated with inhibitors of the PD-1/PD-L1 axis demonstrated a stratification of patients with high and low BAFF plasma levels. Patients with high BAFF levels experienced lower responses to anti-PD-1 immunotherapies. In summary, these results show that BAFF, through its effect on tumor-infiltrating monocytes, not only impacts primary tumor growth but can serve as a biomarker to predict response to anti-PD-1 immunotherapy in advanced disease. SIGNIFICANCE: The BAFF cytokine regulates monocytes in the melanoma microenvironment to suppress tumor growth, highlighting the importance of BAFF in antitumor immunity.