Regulation Of Innate Immune Responses Research Articles

SIRT6 Acts as a Negative Regulator in Dengue Virus-Induced Inflammatory Response by Targeting the DNA Binding Domain of NF-κB p65.

Dengue virus (DENV) is a mosquito-borne single-stranded RNA virus causing human disease with variable severity. The production of massive inflammatory cytokines in dengue patients has been associated with dengue disease severity. However, the regulation of these inflammatory responses remains unclear. In this study, we report that SIRT6 is a negative regulator of innate immune responses during DENV infection. Silencing of Sirt6 enhances DENV-induced proinflammatory cytokine and chemokine production. Overexpression of SIRT6 inhibits RIG-I-like receptor (RLR) and Toll-like receptor 3 (TLR3) mediated NF-κB activation. The sirtuin core domain of SIRT6 is required for the inhibition of NF-κB p65 function. SIRT6 interacts with the DNA binding domain of p65 and competes with p65 to occupy the Il6 promoter during DENV infection. Collectively, our study demonstrates that SIRT6 negatively regulates DENV-induced inflammatory response via RLR and TLR3 signaling pathways.

Modulating ADME Properties by Fluorination: MK2 Inhibitors with Improved Oral Exposure.

MAP-activated protein kinase 2 (MK2) plays an important role in the regulation of innate immune response as well as in cell survival upon DNA damage. Despite its potential for the treatment of inflammation and cancer, to date no MK2 low molecular weight inhibitors have reached the clinic, mainly due to inadequate absorption, distribution, metabolism, and excretion (ADME) properties. We describe here an approach based on specifically placed fluorine within a recently described pyrrole-based MK2 inhibitor scaffold for manipulation of its physicochemical and ADME properties. While preserving target potency, the novel fluoro-derivatives showed greatly improved permeability as well as enhanced solubility and reduced in vivo clearance leading to significantly increased oral exposure.

TRIM29 negatively controls antiviral immune response through targeting STING for degradation

Innate immune system is armed by several lines of pattern recognition receptors to sense various viral infection and to initiate antiviral immune response. This process is under a tight control and the negative feedback induced by infection and/or inflammation is critical to maintain immune homoeostasis and to prevent autoimmune disorders, however, the molecular mechanism is not fully understood. Here we report TRIM29, a ubiquitin E3 ligase, functions as an inducible negative regulator of innate immune response triggered by DNA virus and cytosolic DNA. DNA virus and cytosolic DNA stimulation induce TRIM29 expression robustly in macrophages and dendritic cells, although the basal level of TRIM29 is undetectable in those cells. TRIM29 deficiency elevates IFN-I and proinflammatory cytokine production upon viral DNA and cytosolic dsDNA stimulation. Consistently, in vivo experiments show that TRIM29-deficient mice are more resistant to HSV-1 infection than WT controls, indicated by better survival rate and reduced viral load in organs. Mechanism studies suggest that STING–TBK1–IRF3 signaling pathway in TRIM29 KO cells is significantly enhanced and the degradation of STING is impaired. Furthermore, we identify that TRIM29 targets STING for K48 ubiquitination and degradation. This study reveals TRIM29 as a crucial negative regulator in immune response to DNA virus and cytosolic DNA, preventing potential damage caused by overcommitted immune responses.

DeSUMOylation of MKK7 kinase by the SUMO2/3 protease SENP3 potentiates lipopolysaccharide-induced inflammatory signaling in macrophages

Protein SUMOylation has been reported to play a role in innate immune response, but the enzymes, substrates, and consequences of the specific inflammatory signaling events are largely unknown. Reactive oxygen species (ROS) are abundantly produced during macrophage activation and required for Toll-like receptor 4 (TLR4)–mediated inflammatory signaling. Previously, we demonstrated that SENP3 is a redox-sensitive SUMO2/3 protease. To explore any links between reversible SUMOylation and ROS-related inflammatory signaling in macrophage activation, we generated mice with Senp3 conditional knock-out in myeloid cells. In bacterial lipopolysaccharide (LPS)-induced in vitro and in vivo inflammation models, we found that SENP3 deficiency markedly compromises the activation of TLR4 inflammatory signaling and the production of proinflammatory cytokines in macrophages exposed to LPS. Moreover, Senp3 conditional knock-out mice were significantly less susceptible to septic shock. Of note, SENP3 deficiency was associated with impairment in JNK phosphorylation. We found that MKK7, which selectively phosphorylates JNK, is a SENP3 substrate and that SENP3-mediated deSUMOylation of MKK7 may favor its binding to JNK. Importantly, ROS-dependent SENP3 accumulation and MKK7 deSUMOylation rapidly occurred after LPS stimulation. In conclusion, our findings indicate that SENP3 potentiates LPS-induced TLR4 signaling via deSUMOylation of MKK7 leading to enhancement in JNK phosphorylation and the downstream events. Therefore this work provides novel mechanistic insights into redox regulation of innate immune responses.

The Emerging Role of Pattern Recognition Receptors in the Pathogenesis of Malaria.

Despite a global effort to develop an effective vaccine, malaria is still a significant health problem. Much of the pathology of malaria is immune mediated. This suggests that host immune responses have to be finely regulated. The innate immune system initiates and sets the threshold of the acquired immune response and determines the outcome of the disease. Yet, our knowledge of the regulation of innate immune responses during malaria is limited. Theoretically, inadequate activation of the innate immune system could result in unrestrained parasite growth. Conversely, hyperactivation of the innate immune system, is likely to cause excessive production of proinflammatory cytokines and severe pathology. Toll-like receptors (TLRs) have emerged as essential receptors which detect signature molecules and shape the complex host response during malaria infection. This review will highlight the mechanisms by which Plasmodium components are recognized by innate immune receptors with particular emphasis on TLRs. A thorough understanding of the complex roles of TLRs in malaria may allow the delineation of pathological versus protective host responses and enhance the efficacy of anti-malarial treatments and vaccines.

Elevated sodium chloride drives type I interferon signaling in macrophages and increases antiviral resistance

Type I IFN production and signaling in macrophages play critical roles in innate immune responses. High salt (i.e. high concentrations of NaCl) has been proposed to be an important environmental factor that influences immune responses in multiple ways. However, it remains unknown whether high salt regulates type I IFN production and signaling in macrophages. Here, we demonstrated that high salt promoted IFNβ production and its signaling in both human and mouse macrophages, and consequentially primed macrophages for strengthened immune sensing and signaling when challenged with viruses or viral nucleic acid analogues. Using both pharmacological inhibitors and RNA interference we showed that these effects of high salt on IFNβ signaling were mediated by the p38 MAPK/ATF2/AP1 signaling pathway. Consistently, high salt increased resistance to vesicle stomatitis virus (VSV) infection in vitro. In vivo data indicated that a high-salt diet protected mice from lethal VSV infection. Taken together, these results identify high salt as a crucial regulator of type I IFN production and signaling, shedding important new light on the regulation of innate immune responses.

Enhanced survival of BCG-stimulated dendritic cells: involvement of anti-apoptotic proteins and NF-κB.

ABSTRACTBCG (Bacillus Calmette-Guérin) is the only available vaccine against TB and is also used for the treatment of superficial bladder cancer. BCG-mediated protection against TB and bladder cancer has been shown to rely on its ability to induce superior CD4+ and CD8+ T cell responses. As the magnitude of T cell responses is defined by dendritic cell (DC) lifespan, we examined the effect of BCG on DC survival and its underlying mechanisms. It was observed that BCG stimulation enhanced DC survival and prolonged DC lifespan in a dose-dependent manner. Live BCG led to a higher DC survival compared with heat-killed BCG. FITC-Annexin V staining showed that BCG promoted DC survival by inhibiting apoptosis. Consistently, higher expressions of anti-apoptotic proteins Bcl-2 and Bcl-xL were observed in BCG-stimulated DCs. Pharmacological inhibition of Bcl-2 and Bcl-xL drastically reduced the DC survival efficacy of BCG. Comparable survival of BCG-stimulated wild-type and MyD88−/− DCs suggested that MyD88 signaling is dispensable for BCG-induced DC survival. NF-κB is one of the key regulators of innate immune responses. We observed that pharmacological inhibition of NF-κB abrogated BCG-mediated increase in DC survival and expression of anti-apoptotic proteins. These findings provide a novel insight into the effect of BCG on DC physiology.

5,5-Dimethyl-1-pyrroline N -oxide Modulates Transcriptome and Interactome Towards Dampening Innate Immune Response in RAW 264.7 cells

The nitrone spin trap 5,5-dimethyl-1-pyrroline N-oxide (DMPO) was originally synthesized for the study of free radicals. However we found that DMPO can also prevent inflammatory activation in RAW264.7 cells primed with lipopolysaccharide (LPS). Additionally, DMPO prevents lung damage, adipose tissue inflammation, systemic inflammation and insulin resistance in a mouse model of acute distress respiratory syndrome induced by intratracheal instillation with LPS. These effects of DMPO are more likely caused by changes in gene expression in innate immune cells. Thus herein we tested whether DMPO by itself can change the transcriptome and interactome in RAW 264.7 cells, as a model of macrophage. To accomplish our goal we incubated RAW 264.7 cells with (DMPO) or without (control) 50 mM DMPO for 6h. The transcriptome was analyzed using micro-array (Illumina) and corroborated by Nanostring (nCounter) technology. Bioinformatics analysis showed 79 differentially expressed genes (DEGs) in DMPO vs control comparison (ONE-way ANOVA; FDR = 0.05). DAVID databases for identifying enriched Gene Onthology terms and Ingenuity Pathway Analysis (IPA) for functional analysis showed that DMPO DEGs were consistent with a negative regulation of innate immune response. Functional analysis indicated that IRF7 and TLRs were related (predicted inhibitions) to the observed transcriptomic effects of DMPO. Functional data analyses are consistent with DMPO dampening LPS-induced inflammatory activation of RAW 264.7 cells by down regulating several genes related to the innate immune response. These effects were also supported by Nanostring data showing that DMPO downregulates the expression of inflammation markers (CCL2, IFNβ, COX-2 and iNOS). Remarkably, our data suggest that DMPO by itself shifts the transcriptomic profile of RAW264.7 cells towards a negative modulation of innate immune response.

The Relationship between Mitochondria and NLRP3 Inflammasome

Mitochondria participate in various intracellular metabolic pathways such as generating intracellular ATP, synthesizing several essential molecules, regulating calcium homeostasis, and producing the cell’s reactive oxygen species (ROS). Emerging studies have demonstrated newly discovered roles of mitochondria, which participate in the regulation of innate immune responses by modulating NLRP3 inflammasomes. Here, we review the recently proposed pathways to be involved in mitochondria-mediated regulation of inflammasome activation and inflammation: 1) mitochondrial ROS, 2) calcium mobilization, 3) nicotinamide adenine dinucleotide (NAD+) reduction, 4) cardiolipin, 5) mitofusin, 6) mitochondrial DNA, 7) mitochondrial antiviral signaling protein. Furthermore, we highlight the significance of mitophagy as a negative regulator of mitochondrial damage and NLRP3 inflammasome activation, as potentially helpful therapeutic approaches which could potentially address uncontrolled inflammation.

MKP-1 negative regulates Staphylococcus aureus induced inflammatory responses in Raw264.7 cells: roles of PKA-MKP-1 pathway and enhanced by rolipram

MAP phosphatases (MKP)-1 acts as an important regulator of innate immune response through a mechanism of control and attention both MAPK and NF-κB molecules during bacterial infection. However, the regulatory role of MKP-1 in the interplay between MAPK and NFκB pathway molecules is still not fully understood. In present study, we showed a direct interactions of p38, ERK or IκBα with MKP-1, and demonstrated that MKP-1 was a pivotal feedback control for both MAP kinases and NF-κB pathway in response to S. aureus. In addition, we found that rolipram had anti-inflammatory activity and repressed IκBα activation induced by S. aureus via PKA-MKP-1 pathway. Our report also demonstrated that PKA-cα can directly bind to IκBα upon S. aureus stimulation, which influenced the downstream signaling of PKA pathway, including altered the expression of MKP-1. These results presented a novel mechanism of PKA and IκB pathway, which may be targeted for treating S. aureus infection.

Arginine methylation regulates c-Myc–dependent transcription by altering promoter recruitment of the acetyltransferase p300

Protein arginine methyltransferase 1 (PRMT1) is an essential enzyme controlling about 85% of the total cellular arginine methylation in proteins. We have shown previously that PRMT1 is an important regulator of innate immune responses and that it is required for M2 macrophage differentiation. c-Myc is a transcription factor that is critical in regulating cell proliferation and also regulates the M2 transcriptional program in macrophages. Here, we sought to determine whether c-Myc in myeloid cells is regulated by PRMT1-dependent arginine methylation. We found that PRMT1 activity was necessary for c-Myc binding to the acetyltransferase p300. PRMT1 inhibition decreased p300 recruitment to c-Myc target promoters and increased histone deacetylase 1 (HDAC1) recruitment, thereby decreasing transcription at these sites. Moreover, PRMT1 inhibition blocked c-Myc-mediated induction of several of its target genes, including peroxisome proliferator-activated receptor γ (PPARG) and mannose receptor C-type 1 (MRC1), suggesting that PRMT1 is necessary for c-Myc function in M2 macrophage differentiation. Of note, in primary human blood monocytes, p300-c-Myc binding was strongly correlated with PRMT1 expression, and in liver sections, PRMT1, c-Myc, and M2 macrophage levels were strongly correlated with each other. Both PRMT1 levels and M2 macrophage numbers were significantly lower in livers from individuals with a history of spontaneous bacterial peritonitis, known to have defective cellular immunity. In conclusion, our findings demonstrate that PRMT1 is an important regulator of c-Myc function in myeloid cells. PRMT1 loss in individuals with cirrhosis may contribute to their immune defects.

Imbalance between innate antiviral and pro-inflammatory immune responses may contribute to different outcomes involving low- and highly pathogenic avian influenza H5N3 infections in chickens.

In order to gain further insight into the early virus-host interactions associated with highly pathogenic avian influenza virus infections in chickens, genome-wide expression profiling of chicken lung and brain was carried out at 24 and 72 h post-inoculation (h p.i.). For this purpose two recombinant H5N3 viruses were utilized, each possessing a polybasic HA0 cleavage site but differing in pathogenicity. The original rH5N3 P0 virus, which has a low-pathogenic phenotype, was passaged six times through chickens to give rise to the derivative rH5N3 P6 virus, which is highly pathogenic (Diederich S, Berhane Y, Embury-Hyatt C, Hisanaga T, Handel K et al.J Virol 2015;89:10724-10734). The gene-expression profiles in lung were similar for both viruses, although they varied in magnitude. While both viruses produced systemic infections, differences in clinical disease progression and viral tissue loads, particularly in brain, where loads of rH5N3 P6 were three orders of magnitude higher than rH5N3 P0 at 72 .p.i., were observed. Although genes associated with gene ontology (GO) categories INFα and INFβ biosynthesis, regulation of innate immune response, response to exogenous dsRNA, defence response to virus, positive regulation of NF-κB import into the nucleus and positive regulation of immune response were up-regulated in rH5N3 P0 and rH5N3 P6 brains, fold changes were higher for rH5N3 P6. The additional up-regulation of genes associated with cytokine production, inflammasome and leukocyte activation, and cell-cell adhesion detected in rH5N3 P6 versus rH5N3 P0 brains, suggested that the balance between antiviral and pro-inflammatory innate immune responses leading to acute CNS inflammation might explain the observed differences in pathogenicity.

Identification of a TLR2-regulated gene signature associated with tumor cell growth in gastric cancer

Toll-like receptors (TLRs) are key regulators of innate immune responses, and their dysregulation is observed in numerous inflammation-associated malignancies, including gastric cancer (GC). However, the identity of specific TLRs and their molecular targets which promote the pathogenesis of human GC is ill-defined. Here, we sought to determine the clinical utility of TLR2 in human GC. TLR2 mRNA and protein expression levels were elevated in >50% of GC patient tumors across multiple ethnicities. TLR2 was also widely expressed among human GC cell lines, and DNA microarray-based expression profiling demonstrated that the TLR2-induced growth responsiveness of human GC cells corresponded with the up-regulation of six anti-apoptotic (BCL2A1, BCL2, BIRC3, CFLAR, IER3, TNFAIP3) and down-regulation of two tumor suppressor (PDCD4, TP53INP1) genes. The TLR2-mediated regulation of these anti-apoptotic and tumor suppressor genes was also supported by their increased and reduced expression, respectively, in two independent genetic GC mouse models (gp130F/F and Gan) characterized by high tumor TLR2 expression. Notably, enrichment of this TLR2-regulated gene signature also positively correlated with augmented TLR2 expression in human GC tumors, and served as an indicator of poor patient survival. Furthermore, treatment of gp130F/F and cell line-derived xenograft (MKN1) GC mouse models with a humanized anti-TLR2 antibody suppressed gastric tumor growth, which was coincident with alterations to the TLR2-driven gene signature. Collectively, our study demonstrates that in the majority of GC patients, elevated TLR2 expression is associated with a growth-potentiating gene signature which predicts poor patient outcomes, thus supporting TLR2 as a promising therapeutic target in GC.

Loss of CCR5 signaling results in early NK cell and neutrophil recruitment into the liver and increased hepatic inflammation in a mouse model of acute hepatitis B

Abstract Human genetic studies demonstrate that a non-functional CCR5 receptor sequence variant is associated with enhanced control of acute hepatitis B virus (HBV) infection. To better understand the role of CCR5 in immune responses to HBV, we employed a mouse model of acute HBV in which Ccr5 +/+(WT) or Ccr5 −/−(KO) C57BL/6 mice were intravenously infected with adenovirus containing an overlapping HBV1.3 construct (AdHBV) or vector control (AdShuttle). Mice (N=4–6/group) were euthanized at days 2, 3, 7, 10, and 14 post injection. Increased plasma ALT levels were observed in both Ccr5 WT and KO AdHBV mice at day 7, but levels were significantly higher in the Ccr5 KO compared to WT mice by day 14 (554 vs 274 U/L, p < 0.05). Plasma ALT levels were not significantly different between Ccr5 WT and KO AdShuttle mice. Analysis of intrahepatic leukocytes by flow cytometry revealed significantly increased intrahepatic CD11b+ NK cells (26.3 vs 6.9, % of live leukocytes, p< 0.01) and neutrophils (7.3 vs 1.3, % of live leukocytes, p< 0.05) at day 3 in Ccr5 KO compared to WT AdHBV mice. By day 7, these differences in cell populations were no longer present. At Day 14, significantly higher levels of intrahepatic Ly6c+ macrophages were detected in Ccr5 KO compared to WT AdHBV mice (17.0 vs 6.4, % of live leukocytes, p< 0.01). These differences in intrahepatic leukocytes were not observed in AdShuttle mice. Loss of Ccr5 function did not significantly alter intrahepatic CD4+ and CD8+ populations at the time points investigated. Taken together, these findings suggest that loss of Ccr5 signaling increases intrahepatic innate cells and inflammation during acute HBV infection, providing evidence for a role of CCR5 in regulation of innate immune responses to acute HBV infection.

Innate immune response in acute lung injury following hemorrhagic shock

Abstract Trauma is the major cause of death for Americans under the age of 46 years. Hemorrhagic shock accounts for up to 40% of trauma-related deaths. Hemorrhagic shock evokes an acute, non-specific, systemic inflammatory response syndrome (SIRS) resulting in the damage to multiple organs. Acute lung injury (ALI) is one of the most serious complications in traumatic patients, however, the immunological mechanisms in ALI are still not well understood. Recently studies suggest that mitochondria play an important role in physical injury, leading to the onset of the SIRS. In order to address the mechanistic basis of ALI following hemorrhagic shock, we subjected rats to severe hemorrhage and shock (hemorrhagic injury - HI) or sham procedure and lung tissue was tested. We demonstrate that NLRX1, a mitochondria-targeted protein, that negatively regulates innate immunity and cell death responses, was markedly decreased in HI lung. The decrease of NLRX1 was concomitant with the activation of NF/κB and TBK1 signaling pathways. Lung histochemistry was used to establish pathologic effect on the lung. Significant increases in pulmonary expression of inflammatory signaling molecules IL-6, IL-1β, IL-10, TNF-α and MIP-1α further confirmed HI induced pulmonary inflammation and injury. Our results suggest NLRX1 participate in the pathogenesis of HI related pulmonary diseases and support its important role in the regulation of innate immune response following hemorrhagic shock.

Innate immune signaling in Drosophila is regulated by transforming growth factor β (TGFβ)-activated kinase (Tak1)-triggered ubiquitin editing

Coordinated regulation of innate immune responses is necessary in all metazoans. In Drosophila the Imd pathway detects Gram-negative bacterial infections through recognition of diaminopimelic acid (DAP)-type peptidoglycan and activation of the NF-κB precursor Relish, which drives robust antimicrobial peptide gene expression. Imd is a receptor-proximal adaptor protein homologous to mammalian RIP1 that is regulated by proteolytic cleavage and Lys-63-polyubiquitination. However, the precise events and molecular mechanisms that control the post-translational modification of Imd remain unclear. Here, we demonstrate that Imd is rapidly Lys-63-polyubiquitinated at lysine residues 137 and 153 by the sequential action of two E2 enzymes, Ubc5 and Ubc13-Uev1a, in conjunction with the E3 ligase Diap2. Lys-63-ubiquitination activates the TGFβ-activated kinase (Tak1), which feeds back to phosphorylate Imd, triggering the removal of Lys-63 chains and the addition of Lys-48 polyubiquitin. This ubiquitin-editing process results in the proteasomal degradation of Imd, which we propose functions to restore homeostasis to the Drosophila immune response.

Modulation of experimental arthritis by vagal sensory and central brain stimulation

Articular inflammation is a major clinical burden in multiple inflammatory diseases, especially in rheumatoid arthritis. Biological anti-rheumatic drug therapies are expensive and increase the risk of systemic immunosuppression, infections, and malignancies. Here, we report that vagus nerve stimulation controls arthritic joint inflammation by inducing local regulation of innate immune response. Most of the previous studies of neuromodulation focused on vagal regulation of inflammation via the efferent peripheral pathway toward the viscera. Here, we report that vagal stimulation modulates arthritic joint inflammation through a novel “afferent” pathway mediated by the locus coeruleus (LC) of the central nervous system. Afferent vagal stimulation activates two sympatho-excitatory brain areas: the paraventricular hypothalamic nucleus (PVN) and the LC. The integrity of the LC, but not that of the PVN, is critical for vagal control of arthritic joint inflammation. Afferent vagal stimulation suppresses articular inflammation in the ipsilateral, but not in the contralateral knee to the hemispheric LC lesion. Central stimulation is followed by subsequent activation of joint sympathetic nerve terminals inducing articular norepinephrine release. Selective adrenergic beta-blockers prevent the effects of articular norepinephrine and thereby abrogate vagal control of arthritic joint inflammation. These results reveals a novel neuro-immune brain map with afferent vagal signals controlling side-specific articular inflammation through specific inflammatory-processing brain centers and joint sympathetic innervations.

Blockade of Notch signaling promotes acetaminophen-induced liver injury.

Liver injury after experimental acetaminophen treatment is mediated both by direct hepatocyte injury through a P450-generated toxic metabolite and indirectly by activated liver Kupffer cells and neutrophils. This study was designed to investigate the role of Notch signaling in the regulation of innate immune responses in acetaminophen (APAP)-induced liver injury. Using a mouse model of APAP-induced liver injury, wild-type (WT) and toll-like receptor 4 knockout (TLR4 KO) mice were injected intraperitoneally with APAP or PBS. Some animals were injected with γ-secretase inhibitor DAPT or DMSO vehicle. For the in vitro study, bone marrow-derived macrophages (BMMs) were transfected with Notch1 siRNA, TLR4 siRNA, and non-specific (NS) siRNA and stimulated with LPS. Indeed, paracetamol/acetaminophen-induced liver damage was worse after Notch blockade with DAPT in wild-type mice, which was accompanied by significantly increased ALT levels, diminished hairy and enhancer of split-1 (Hes1), and phosphorylated Stat3 and Akt but enhanced high mobility group box 1 (HMGB1), TLR4, NF-κB, and NLRP3 activation after APAP challenge. Mice receiving DAPT increased macrophage and neutrophil accumulation and hepatocellular apoptosis. However, TLR4 KO mice that received DAPT reduced APAP-induced liver damage and NF-κB, NLRP3, and cleaved caspase-1 activation. BMMs transfected with Notch1 siRNA reduced Hes1 and phosphorylated Stat3 and Akt but augmented HMGB1, TLR4, NF-κB, and NLRP3. Furthermore, TLR4 siRNA knockdown resulted in decreased NF-κB and NLRP3 and cleaved caspase-1 and IL-1β levels following LPS stimulation. These results demonstrate that Notch signaling regulates innate NLRP3 inflammasome activation through regulation of HMGB1/TLR4/NF-κB activation in APAP-induced liver injury. Our novel findings underscore the critical role of the Notch1-Hes1 signaling cascade in the regulation of innate immunity in APAP-triggered liver inflammation. This might imply a novel therapeutic potential for the drug-induced damage-associated lethal hepatitis.

TLR10 suppresses the activation and differentiation of monocytes with effects on DC-mediated adaptive immune responses.

TLRs are important pattern-recognition receptors involved in the activation of innate immune responses against foreign pathogens. TLR10 is the only TLR family member without a known ligand, signaling pathway, or clear cellular function. Previous work has shown that TLR10 suppresses proinflammatory cytokine production in response to TLR agonists in a mixed human mononuclear cell population. We report that TLR10 is preferentially expressed on monocytes and suppresses proinflammatory cytokine production resulting from either TLR or CD40 stimulation. TLR10 engagement affects both the MAPK and Akt signaling pathways, leading to changes in the transcriptome of isolated human monocytes. Differentiation of monocytes into dendritic cells in the presence of an αTLR10 mAb reduced the expression of maturation markers and the induction of proinflammatory cytokines, again in response to either TLR or CD40 stimulation. Finally, in coculture experiments, TLR10 differentiated dendritic cells exhibited a decreased capacity to activate T cells as measured by IL-2 and IFN-γ production. These data demonstrate that TLR10 is a novel regulator of innate immune responses and of the differentiation of primary human monocytes into effective dendritic cells.

A NLRC3-like gene from blunt snout bream (Megalobrama amblycephala): Molecular characterization, expression and association with resistance to Aeromonas hydrophila infection

NLRC (the nucleotide-oligomerization domain (NOD)-like receptor subfamily C) consists of teleost-specific NLRs (NOD-like receptors) and plays pivotal roles in microbial recognition and regulation of innate immune response. In this study, we cloned and characterized a NLRC3-like gene (MamNLRC3-like) from blunt snout bream (Megalobrama amblycephala) by using the quantitative real-time PCR method, and analyzed the correlation between its polymorphisms and resistance to Aeromonas hydrophila infection. The full length cDNA of MamNLRC3-like was 2863 bp, with a 5′-UTR of 169 bp, ORF of 2301 bp and 3′-UTR of 393 bp, encoding 766 amino acid residues. MamNLRC3-like consisted of a conserved NACHT domain, a fish-specific NACHT associated domain (FISNA) and a PYRIN effective domain. During early developmental stages, MamNLRC3-like was most highly expressed at 39.4 hpf (hours post fertilization, hatching stage) and constitutively detected in various healthy tissues. The expression of MamNLRC3-like was strongly induced by A. hydrophila infection and peaked in the head kidney, liver, intestine, and trunk kidney at 12 h post infection. Six SNPs (single nucleotide polymorphisms) from MamNLRC3-like, with 2 in introns and 4 in exons, were identified by direct sequencing, in which 6515C/T was a novel non-synonymous mutation. HRM (high resolution melting) genotyping of the 6 loci showed that locus 6515C/T SNP was associated with the resistance/susceptibility of blunt snout bream to A. hydrophila infection (P < 0.01). The CC genotype fish were more resistant than CT carriers (P < 0.01). This study suggests that the MamNLRC3-like gene might play an important role in the innate immunity of blunt snout bream and could be used as a candidate marker for genetic selection of A. hydrophila-resistant blunt snout bream.