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Immunology
A Simple Multistep Protocol for Differentiating Human Induced Pluripotent Stem Cells into Functional Macrophages.
May 15, 2018   Methods In Molecular Biology (Clifton, N.J.)
Mukherjee C, Hale C, Mukhopadhyay S
A Simple Multistep Protocol for Differentiating Human Induced Pluripotent Stem Cells into Functional Macrophages.
May 15, 2018
Methods In Molecular Biology (Clifton, N.J.)
Macrophages differentiated from human induced pluripotent stem cells (hiPSCs) provide an alternative new tool overcoming some of the limitations of existing models for human macrophages, such as human macrophage-like cell lines and primary monocyte-derived macrophages. A combination of different cytokines and growth factors can differentiate hiPSCs toward myeloid lineage. Here we describe a simple multistep protocol for differentiating hiPSCs into functional macrophages. This includes derivation of three germ-line containing embryoid bodies (EBs) from iPSCs, generation of myeloid precursors from EBs, and finally maturation of myeloid precursors into functional macrophages. Technical procedure and specific culture conditions associated with each of these steps are discussed in detail.
TRIM29 Negatively Regulates the Type I IFN Production in Response to RNA Virus.
Jun 19, 2018   Journal Of Immunology (Baltimore, Md. : 1950)
Xing J, Zhang A, Minze LJ, Li XC, Zhang Z
TRIM29 Negatively Regulates the Type I IFN Production in Response to RNA Virus.
Jun 19, 2018
Journal Of Immunology (Baltimore, Md. : 1950)
The innate immunity is critically important in protection against virus infections, and in the case of RNA viral infections, the signaling mechanisms that initiate robust protective innate immunity without triggering autoimmune inflammation remain incompletely defined. In this study, we found the E3 ligase TRIM29 was specifically expressed in poly I:C-stimulated human myeloid dendritic cells. The induced TRIM29 played a negative role in type I IFN production in response to poly I:C or dsRNA virus reovirus infection. Importantly, the challenge of wild-type mice with reovirus led to lethal infection. In contrast, deletion of TRIM29 protected the mice from this developing lethality. Additionally, TRIM29-/- mice have lower titers of reovirus in the heart, intestine, spleen, liver, and brain because of elevated production of type I IFN. Mechanistically, TRIM29 was shown to interact with MAVS and subsequently induce its K11-linked ubiquitination and degradation. Taken together, TRIM29 regulates negatively the host innate immune response to RNA virus, which could be employed by RNA viruses for viral pathogenesis.
Recent advances in the discovery and development of TLR ligands as novel therapeutics for chronic HBV and HIV infections.
Jun 14, 2018   Expert Opinion On Drug Discovery
Du K, Liu J, Broering R, Zhang X, Yang D, Dittmer U, Lu M
Recent advances in the discovery and development of TLR ligands as novel therapeutics for chronic HBV and HIV infections.
Jun 14, 2018
Expert Opinion On Drug Discovery
INTRODUCTION: Toll-like receptor (TLR) ligands remain as promising antiviral drug candidates for the treatment of chronic viral infections. Basic research on the mechanisms of antiviral activity of TLR ligands in preclinical animal models and clinical testing of drug candidates have been carried out in recent years. Areas covered: This review provides an overview of the preclinical and clinical testing of TLR ligands in two major viral infections: hepatitis B virus (HBV) and human immunodeficiency virus (HIV). Recent results have further demonstrated the potent antiviral activity of various TLR ligands . A TLR7 agonist is in clinical trials for the treatment of chronic HBV infection while a HBV vaccine using a TLR9 ligand as an adjuvant has proven to be superior to conventional HBV vaccines and has been approved for clinical use. Generally, TLR activation may achieve viral control mainly by promoting adaptive immunity to viral proteins. Expert opinion: Recent research in this field indicates that TLR ligands could be developed as clinically effective drugs if the obstacles concerning toxicity and application routes are overcome. TLR-mediated promotion of adaptive immunity is a major issue for future studies and will determine the future development of TLR ligands as drugs for immunomodulation.
Evolution, expression, and characterisation of liver-expressed antimicrobial peptide genes in ancient chondrostean sturgeons.
Jun 12, 2018   Fish & Shellfish Immunology
Zhang S, Xu Q, Du H, Qi Z, Li Y, Huang J, Di J, Wei Q
Evolution, expression, and characterisation of liver-expressed antimicrobial peptide genes in ancient chondrostean sturgeons.
Jun 12, 2018
Fish & Shellfish Immunology
Liver-expressed antimicrobial peptide 2 (leap-2) is an evolutionarily ancient molecule that acts as the key component in vertebrate innate immunity against invading pathogens. Leap-2 has been identified and characterised in several teleosts, but not yet in chondrosteans. Herein, the complete coding sequences of leap-2b and leap-2c were identified from expressed sequence tags (ESTs) isolated from Dabry's sturgeon (Acipenser dabryanus) and Chinese sturgeon (A. sinensis), designated as adleap-2b, adleap-2c, asleap-2b, and asleap-2c, respectively. Adleap-2b and adleap-2c sequences share 98% and 100% sequence identity with asleap-2b, and asleap-2c, respectively. Sequence alignment revealed that all four genes contain four cysteine residues, conserved in all fish leap-2 homologs, that form two disulfide bonds. Comparative analysis of the exon-intron structure revealed a three exon/two intron structure for that leap-2 genes in animals, but intron 1 is much longer in sturgeons than in other species. The adleap-2c gene was expressed mainly in the liver of Dabry's sturgeon, and transcription of adleap-2c was significantly up-regulated (p 
Microevolutionary selection dynamics acting on immune genes of the green veined white butterfly, Pieris napi.
May 17, 2018   Molecular Ecology
Keehnen NLP, Hill J, Nylin S, Wheat CW
Microevolutionary selection dynamics acting on immune genes of the green veined white butterfly, Pieris napi.
May 17, 2018
Molecular Ecology
Insects rely on their innate immune system to successfully mediate complex interactions with their microbiota, as well as the microbes present in the environment. Previous work has shown that components of the canonical immune gene repertoire evolve rapidly and have evolutionary characteristics originating from interactions with fast-evolving microorganisms. Although these interactions are likely to vary among populations, there is a poor understanding of the microevolutionary dynamics of immune genes, especially in non-Dipteran insects. Here we use the full set of canonical insect immune genes to investigate microevolutionary dynamics acting on these genes between and among populations by comparing three allopatric populations of the Green Veined White butterfly, Pieris napi (Linné; Lepidoptera, Pieridae). Immune genes showed increased genetic diversity compared to genes from the rest of the genome and various functional categories exhibited different types of signatures of selection, at different evolutionary scales, presenting a complex pattern of selection dynamics. Signatures of balancing selection were identified in 10 genes, and 17 genes appear to be under positive selection. Genes involved with the cellular arm of the immune response as well as the Toll pathway appear to be enriched among our outlier loci, regardless of functional category. This suggests that the targets of selection might focus upon an entire pathway, more than on functional subsets across pathways. Our microevolutionary results are similar to previously observed macroevolutionary patterns from diverse taxa, suggesting that either the immune system is robust to dramatic differences in life history and microbial communities, or that diverse microbes exert similar selection pressures. This article is protected by copyright. All rights reserved.
Structure and Function of the Transmembrane Domain of NsaS, an Antibiotic Sensing Histidine Kinase in Staphylococcus aureus.
Jun 20, 2018   Journal Of The American Chemical Society
Bhate MP, Lemmin T, Kuenze G, Mensa B, Ganguly S, Peters JM, Schmidt N, Pelton JG, Gross CA, Meiler J, DeGrado WF
Structure and Function of the Transmembrane Domain of NsaS, an Antibiotic Sensing Histidine Kinase in Staphylococcus aureus.
Jun 20, 2018
Journal Of The American Chemical Society
NsaS is one of four intramembrane histidine kinases (HKs) in Staphylococcus aureus that mediate the pathogen's response to membrane active antimicrobials and human innate immunity. We describe the first integrative structural study of NsaS using a combination of solution state NMR spectroscopy, chemical-cross-linking, molecular modeling and dynamics. Three key structural features emerge: First, NsaS has a short N-terminal amphiphilic helix that anchors its transmembrane (TM) bundle into the inner leaflet of the membrane such that it might sense neighboring proteins or membrane deformations. Second, the transmembrane domain of NsaS is a 4-helix bundle with significant dynamics and structural deformations at the membrane interface. Third, the intracellular linker connecting the TM domain to the cytoplasmic catalytic domains of NsaS is a marginally stable helical dimer, with one state likely to be a coiled-coil. Data from chemical shifts, heteronuclear NOE, H/D exchange measurements and molecular modeling suggest that this linker might adopt different conformations during antibiotic induced signaling.
A Pelota-like gene regulates root development and defence responses in rice.
May 17, 2018   Annals Of Botany
Ding W, Wu J, Ye J, Zheng W, Wang S, Zhu X, Zhou J, Pan Z, Zhang B, Zhu S
A Pelota-like gene regulates root development and defence responses in rice.
May 17, 2018
Annals Of Botany
Background and Aims: Pelota (Pelo) are evolutionarily conserved genes reported to be involved in ribosome rescue, cell cycle control and meiotic cell division. However, there is little known about their function in plants. The aim of this study was to elucidate the function of an ethylmethane sulphonate (EMS)-derived mutation of a Pelo-like gene in rice (named Ospelo). Methods: A dysfunctional mutant was used to characterize the function of OsPelo. Analyses of its expression and sub-cellular localization were performed. The whole-genome transcriptomic change in leaves of Ospelo was also investigated by RNA sequencing. Key Results: The Ospelo mutant showed defects in root system development and spotted leaves at early seedling stages. Map-based cloning revealed that the mutation occurred in the putative Pelo gene. OsPelo was found to be expressed in various tissues throughout the plant, and the protein was located in mitochondria. Defence responses were induced in the Ospelo mutant, as shown by enhanced resistance to the bacterial pathogen Xanthomonas oryzae pv. oryzae, coupled with upregulation of three pathogenesis-related marker genes. In addition, whole-genome transcriptome analysis showed that OsPelo was significantly associated with a number of biological processes, including translation, metabolism and biotic stress response. Detailed analysis showed that activation of a number of innate immunity-related genes might be responsible for the enhanced disease resistance in the Ospelo mutant. Conclusions: These results demonstrate that OsPelo positively regulates root development while its loss of function enhances pathogen resistance by pre-activation of defence responses in rice.
AID/APOBEC-like cytidine deaminases are ancient innate immune mediators in invertebrates.
May 22, 2018   Nature Communications
Liu MC, Liao WY, Buckley KM, Yang SY, Rast JP, Fugmann SD
AID/APOBEC-like cytidine deaminases are ancient innate immune mediators in invertebrates.
May 22, 2018
Nature Communications
In the course of both innate and adaptive immunity, cytidine deaminases within the activation induced cytidine deaminase (AID)/apolipoprotein B editing complex (APOBEC) family modulate immune responses by mutating specific nucleic acid sequences of hosts and pathogens. The evolutionary emergence of these mediators, however, seems to coincide precisely with the emergence of adaptive immunity in vertebrates. Here, we show a family of genes in species within two divergent invertebrate phyla-the echinoderm Strongylocentrotus purpuratus and the brachiopod Lingula anatina-that encode proteins with similarities in amino acid sequence and enzymatic activities to the vertebrate AID/APOBECs. The expression of these invertebrate factors is enriched in tissues undergoing constant, direct interactions with microbes and can be induced upon pathogen challenge. Our findings suggest that AID/APOBEC proteins, and their function in immunity, emerged far earlier than previously thought. Thus, cytidine deamination is probably an ancient innate immune mechanism that predates the protostome/deuterostome divergence.
TLR-adjuvanted nanoparticle vaccines differentially influence the quality and longevity of responses to malaria antigen Pfs25.
May 17, 2018   JCI Insight
Thompson EA, Ols S, Miura K, Rausch K, Narum DL,   . . . . . .   , Long CA, Duffy PE, Johnston L, O'Neil CP, Loré K
TLR-adjuvanted nanoparticle vaccines differentially influence the quality and longevity of responses to malaria antigen Pfs25.
May 17, 2018
JCI Insight
Transmission-blocking vaccines (TBVs) are considered an integral element of malaria eradication efforts. Despite promising evaluations of Plasmodium falciparum Pfs25-based TBVs in mice, clinical trials have failed to induce robust and long-lived Ab titers, in part due to the poorly immunogenic nature of Pfs25. Using nonhuman primates, we demonstrate that multiple aspects of Pfs25 immunity were enhanced by antigen encapsulation in poly(lactic-co-glycolic acid)-based [(PLGA)-based] synthetic vaccine particles (SVP[Pfs25]) and potent TLR-based adjuvants. SVP[Pfs25] increased Ab titers, Pfs25-specific plasmablasts, circulating memory B cells, and plasma cells in the bone marrow when benchmarked against the clinically tested multimeric form Pfs25-EPA given with GLA-LSQ. SVP[Pfs25] also induced the first reported Pfs25-specific circulating Th1 and Tfh cells to our knowledge. Multivariate correlative analysis indicated several mechanisms for the improved Ab responses. While Pfs25-specific B cells were responsible for increasing Ab titers, T cell responses stimulated increased Ab avidity. The innate immune activation differentially stimulated by the adjuvants revealed a strong correlation between type I IFN polarization, induced by R848 and CpG, and increased Ab half-life and longevity. Collectively, the data identify ways to improve vaccine-induced immunity to poorly immunogenic proteins, both by the choice of antigen and adjuvant formulation, and highlight underlying immunological mechanisms.
Exosome-delivered microRNAs promote IFN-α secretion by human plasmacytoid DCs via TLR7.
May 17, 2018   JCI Insight
Salvi V, Gianello V, Busatto S, Bergese P, Andreoli L, D'Oro U, Zingoni A, Tincani A, Sozzani S, Bosisio D
Exosome-delivered microRNAs promote IFN-α secretion by human plasmacytoid DCs via TLR7.
May 17, 2018
JCI Insight
The excessive production of type I IFNs is a hallmark and a main pathogenic mechanism of many autoimmune diseases, including systemic lupus erythematosus (SLE). In these pathologies, the sustained secretion of type I IFNs is dependent on the improper activation of plasmacytoid DCs (pDCs) by self-nucleic acids. However, the nature and origin of pDC-activating self-nucleic acids is still incompletely characterized. Here, we report that exosomes isolated from the plasma of SLE patients can activate the secretion of IFN-α by human blood pDCs in vitro. This activation requires endosomal acidification and is recapitulated by microRNAs isolated from exosomes, suggesting that exosome-delivered microRNAs act as self-ligands of innate single-stranded endosomal RNA sensors. By using synthetic microRNAs, we identified an IFN induction motif that is responsible for the TLR7-dependent activation, maturation, and survival of human pDCs. These findings identify exosome-delivered microRNAs as potentially novel TLR7 endogenous ligands able to induce pDC activation in SLE patients. Therefore, microRNAs may represent novel pathogenic mediators in the onset of autoimmune reactions and potential therapeutic targets in the treatment of type I IFN-mediated diseases.
The PERK Arm of the Unfolded Protein Response Negatively Regulates Transmissible Gastroenteritis Virus Replication by Suppressing Protein Translation and Promoting Type I IFN Production.
May 17, 2018   Journal Of Virology
Xue M, Fu F, Ma Y, Zhang X, Li L, Feng L, Liu P
The PERK Arm of the Unfolded Protein Response Negatively Regulates Transmissible Gastroenteritis Virus Replication by Suppressing Protein Translation and Promoting Type I IFN Production.
May 17, 2018
Journal Of Virology
Coronavirus replication is closely associated with the endoplasmic reticulum (ER), the primary cellular organelle for protein synthesis, folding, and modification. ER stress is a common consequence in coronavirus-infected cells. However, how the virus-induced ER stress influences coronavirus replication and pathogenesis remains controversial. Here, we demonstrated that infection with the alphacoronavirus transmissible gastroenteritis virus (TGEV) induced ER stress and triggered the unfolded protein response (UPR) in vitro and in vivo, and ER stress negatively regulated TGEV replication in vitro Although TGEV infection activated all three UPR pathways (ATF6, IRE1, and PERK), the virus-triggered UPR suppressed TGEV replication in both ST and IPEC-J2 cells primarily through activation of the PERK-eIF2α axis, as shown by functional studies with overexpression, siRNA, or specific chemical inhibitors. Moreover, we demonstrated that PERK-eIF2α axis-mediated inhibition of TGEV replication is through phosphorylated eIF2α-induced overall attenuation of protein translation. In addition to direct inhibition of viral production, the PERK-eIF2α pathway activated NF-κB and then facilitated type I IFN production, resulting in TGEV suppression. Taken together, our results suggest that the TGEV-triggered PERK-eIF2α pathway negatively regulates TGEV replication and represents a vital aspect of host innate responses to invading pathogens.IMPORTANCE The induction of ER stress is a common outcome in cells infected with coronaviruses. The UPR initiated by ER stress is actively involved in viral replication and modulates the host innate responses to the invading viruses, but these underlying mechanisms remain incompletely understood. We show here that infection with the alphacoronavirus TGEV elicited ER stress in vitro and in vivo, and the UPR PERK-eIF2α branch was predominantly responsible for the suppression of TGEV replication by ER stress. Furthermore, the PERK-eIF2α axis inhibited TGEV replication through direct inhibition of viral proteins due to global translation inhibition and type I IFN induction. These findings highlight a critical role of the UPR PERK-eIF2α pathway in modulating the host innate immunity and coronavirus replication.
Lysophospholipids induce innate immune transdifferentiation of endothelial cells, resulting in prolonged endothelial activation.
May 17, 2018   The Journal Of Biological Chemistry
Li X, Wang L, Fang P, Sun Y, Jiang X, Wang H, Yang XF
Lysophospholipids induce innate immune transdifferentiation of endothelial cells, resulting in prolonged endothelial activation.
May 17, 2018
The Journal Of Biological Chemistry
Innate immune cells express danger-associated molecular pattern (DAMP) receptors, T-cell co-stimulation/co-inhibition receptors, and the major histocompatibility complex II (MHC-II). We have recently proposed that endothelial cells can serve as innate immune cells, but the molecular mechanisms involved still await discovery. Here, we investigated whether human aortic endothelial cells (HAECs) could be transdifferentiated into innate immune cells by exposing them to hyperlipidemia-upregulated DAMP molecules, i.e. lysophospholipids. Performing RNA-Seq analysis of lysophospholipid-treated HAECs, we found that lysophosphatidylcholine (LPC) and lysophosphatidylinositol (LPI) regulate largely distinct gene programs, as revealed by principal component analysis. Metabolically, LPC up-regulated genes that are involved in cholesterol biosynthesis, presumably through sterol regulatory element-binding protein 2 (SREBP2). By contrast, LPI up-regulated gene transcripts critical for the metabolism of glucose, lipids, and amino acids. Of note, we found that LPC and LPI both induce adhesion molecules, cytokines, and chemokines in HAECs, which are all classic markers of endothelial cell activation. Moreover, LPC and LPI shared the ability to transdifferentiate HAECs into innate immune cells, including induction of potent DAMP receptors, such as CD36 molecule, T-cell co-stimulation/co-inhibition receptors, and MHC-II proteins. The induction of these innate-immunity signatures by lysophospholipids correlated with their ability to induce up-regulation of cytosolic calcium and mitochondrial reactive oxygen species. In conclusion, lysophospholipids such as LPC and LPI induce innate immune cell transdifferentiation in HAECs. The concept of prolonged endothelial activation, discovered here, is relevant for designing new strategies for managing cardiovascular diseases.
Tobacco-induced suppression of the vascular response to dental plaque.
May 16, 2018   Molecular Oral Microbiology
Buduneli N, Scott DA
Tobacco-induced suppression of the vascular response to dental plaque.
May 16, 2018
Molecular Oral Microbiology
Cigarette smoking presents oral health professionals with a clinical and research conundrum: reduced periodontal vascular responsiveness to the oral biofilm accompanied by increased susceptibility to destructive periodontal diseases. This presents a significant problem, hampering diagnosis and complicating treatment planning. The aim of this review is to summarize contemporary hypotheses that help explain mechanistically the phenomenon of a suppressed bleeding response to dysbiotic plaque in the periodontia of smokers. The influence of smoke exposure on angiogenesis, innate cell function, the production of inflammatory mediators including cytokines and proteases, tobacco-bacterial interactions and potential genetic predisposition, are discussed. This article is protected by copyright. All rights reserved.
Immortalization of Murine Bone Marrow-Derived Macrophages.
May 15, 2018   Methods In Molecular Biology (Clifton, N.J.)
De Nardo D, Kalvakolanu DV, Latz E
Immortalization of Murine Bone Marrow-Derived Macrophages.
May 15, 2018
Methods In Molecular Biology (Clifton, N.J.)
Macrophages are specialized phagocytes that display a variety of important functions for the host immune system. They are particularly important for the recognition of exogenous and endogenous danger signals, forming the defensive front line as part of innate immune response. As such, murine macrophages are commonly used for in vitro cell-based assays examining the mechanisms of innate immune activation, which can require the ongoing breeding and housing of a large number of genetically modified mouse strains. Here, we describe a robust protocol for the generation of immortalized bone marrow-derived macrophages (iBMDMs) from primary murine bone marrow cells. We further provide general protocols for harvesting, freezing, and thawing of bone marrow cells, maintaining iBMDMs in culture and generation of monoclonal iBMDM populations by single-cell cloning.
Integrative omics to detect bacteremia in patients with febrile neutropenia.
May 27, 2018   PloS One
Kelly RS, Lasky-Su J, Yeung SJ, Stone RM, Caterino JM, Hagan SC, Lyman GH, Baden LR, Glotzbecker BE, Coyne CJ, Baugh CW, Pallin DJ
Integrative omics to detect bacteremia in patients with febrile neutropenia.
May 27, 2018
PloS One
BACKGROUND: Cancer chemotherapy-associated febrile neutropenia (FN) is a common condition that is deadly when bacteremia is present. Detection of bacteremia depends on culture, which takes days, and no accurate predictive tools applicable to the initial evaluation are available. We utilized metabolomics and transcriptomics to develop multivariable predictors of bacteremia among FN patients. METHODS: We classified emergency department patients with FN and no apparent infection at presentation as bacteremic (cases) or not (controls), according to blood culture results. We assessed relative metabolite abundance in plasma, and relative expression of 2,560 immunology and cancer-related genes in whole blood. We used logistic regression to identify multivariable predictors of bacteremia, and report test characteristics of the derived predictors. RESULTS: For metabolomics, 14 bacteremic cases and 25 non-bacteremic controls were available for analysis; for transcriptomics we had 7 and 22 respectively. A 5-predictor metabolomic model had an area under the receiver operating characteristic curve of 0.991 (95%CI: 0.972,1.000), 100% sensitivity, and 96% specificity for identifying bacteremia. Pregnenolone steroids were more abundant in cases and carnitine metabolites were more abundant in controls. A 3-predictor gene expression model had corresponding results of 0.961 (95%CI: 0.896,1.000), 100%, and 86%. Genes involved in innate immunity were differentially expressed. CONCLUSIONS: Classifiers derived from metabolomic and gene expression data hold promise as objective and accurate predictors of bacteremia among FN patients without apparent infection at presentation, and can provide insights into the underlying biology. Our findings should be considered illustrative, but may lay the groundwork for future biomarker development.
Antifungal Innate Immunity: A Perspective from the Last 10 Years.
May 16, 2018   Journal Of Innate Immunity
Salazar F, Brown GD
Antifungal Innate Immunity: A Perspective from the Last 10 Years.
May 16, 2018
Journal Of Innate Immunity
Fungal pathogens can rarely cause diseases in immunocompetent individuals. However, commensal and normally nonpathogenic environmental fungi can cause life-threatening infections in immunocompromised individuals. Over the last few decades, there has been a huge increase in the incidence of invasive opportunistic fungal infections along with a worrying increase in antifungal drug resistance. As a consequence, research focused on understanding the molecular and cellular basis of antifungal immunity has expanded tremendously in the last few years. This review will provide an overview of the most exciting recent advances in innate antifungal immunity, discoveries that are helping to pave the way for the development of new strategies that are desperately needed to combat these devastating diseases.
Dietary Fiber Confers Protection against Flu by Shaping Ly6c- Patrolling Monocyte Hematopoiesis and CD8+ T Cell Metabolism.
May 16, 2018   Immunity
Trompette A, Gollwitzer ES, Pattaroni C, Lopez-Mejia IC, Riva E, Pernot J, Ubags N, Fajas L, Nicod LP, Marsland BJ
Dietary Fiber Confers Protection against Flu by Shaping Ly6c- Patrolling Monocyte Hematopoiesis and CD8+ T Cell Metabolism.
May 16, 2018
Immunity
Dietary fiber protects against chronic inflammatory diseases by dampening immune responses through short-chain fatty acids (SCFAs). Here we examined the effect of dietary fiber in viral infection, where the anti-inflammatory properties of SCFAs in principle could prevent protective immunity. Instead, we found that fermentable dietary fiber increased survival of influenza-infected mice through two complementary mechanisms. High-fiber diet (HFD)-fed mice exhibited altered bone marrow hematopoiesis, characterized by enhanced generation of Ly6c- patrolling monocytes, which led to increased numbers of alternatively activated macrophages with a limited capacity to produce the chemokine CXCL1 in the airways. Blunted CXCL1 production reduced neutrophil recruitment to the airways, thus limiting tissue immunopathology during infection. In parallel, diet-derived SCFAs boosted CD8+ T cell effector function by enhancing cellular metabolism. Hence, dietary fermentable fiber and SCFAs set an immune equilibrium, balancing innate and adaptive immunity so as to promote the resolution of influenza infection while preventing immune-associated pathology.
Intestinal Epithelial Wnt Signaling Mediates Acetylcholine-Triggered Host Defense against Infection.
May 23, 2018   Immunity
Labed SA, Wani KA, Jagadeesan S, Hakkim A, Najibi M, Irazoqui JE
Intestinal Epithelial Wnt Signaling Mediates Acetylcholine-Triggered Host Defense against Infection.
May 23, 2018
Immunity
Regulated antimicrobial peptide expression in the intestinal epithelium is key to defense against infection and to microbiota homeostasis. Understanding the mechanisms that regulate such expression is necessary for understanding immune homeostasis and inflammatory disease and for developing safe and effective therapies. We used Caenorhabditis elegans in a preclinical approach to discover mechanisms of antimicrobial gene expression control in the intestinal epithelium. We found an unexpected role for the cholinergic nervous system. Infection-induced acetylcholine release from neurons stimulated muscarinic signaling in the epithelium, driving downstream induction of Wnt expression in the same tissue. Wnt induction activated the epithelial canonical Wnt pathway, resulting in the expression of C-type lectin and lysozyme genes that enhanced host defense. Furthermore, the muscarinic and Wnt pathways are linked by conserved transcription factors. These results reveal a tight connection between the nervous system and the intestinal epithelium, with important implications for host defense, immune homeostasis, and cancer.
The Chaperone UNC93B1 Regulates Toll-like Receptor Stability Independently of Endosomal TLR Transport.
May 16, 2018   Immunity
Pelka K, Bertheloot D, Reimer E, Phulphagar K, Schmidt SV,   . . . . . .   , Haas A, Brinkmann MM, Marshak-Rothstein A, Meissner F, Latz E
The Chaperone UNC93B1 Regulates Toll-like Receptor Stability Independently of Endosomal TLR Transport.
May 16, 2018
Immunity
Unc-93 homolog B1 (UNC93B1) is a key regulator of nucleic acid (NA)-sensing Toll-like receptors (TLRs). Loss of NA-sensing TLR responses in UNC93B1-deficient patients facilitates Herpes simplex virus type 1 (HSV-1) encephalitis. UNC93B1 is thought to guide NA-sensing TLRs from the endoplasmic reticulum (ER) to their respective endosomal signaling compartments and to guide the flagellin receptor TLR5 to the cell surface, raising the question of how UNC93B1 mediates differential TLR trafficking. Here, we report that UNC93B1 regulates a step upstream of the differential TLR trafficking process. We discovered that UNC93B1 deficiency resulted in near-complete loss of TLR3 and TLR7 proteins in primary splenic mouse dendritic cells and macrophages, showing that UNC93B1 is critical for maintaining TLR expression. Notably, expression of an ER-retained UNC93B1 version was sufficient to stabilize TLRs and largely restore endosomal TLR trafficking and activity. These data are critical for an understanding of how UNC93B1 can regulate the function of a broad subset of TLRs.
Tissue Damage Signaling Is a Prerequisite for Protective Neutrophil Recruitment to Microbial Infection in Zebrafish.
May 16, 2018   Immunity
Huang C, Niethammer P
Tissue Damage Signaling Is a Prerequisite for Protective Neutrophil Recruitment to Microbial Infection in Zebrafish.
May 16, 2018
Immunity
Tissue damage and infection are deemed likewise triggers of innate immune responses. But whereas neutrophil responses to microbes are generally protective, neutrophil recruitment into damaged tissues without infection is deleterious. Why neutrophils respond to tissue damage and not just to microbes is unknown. Is it a flaw of the innate immune system that persists because evolution did not select against it, or does it provide a selective advantage? Here we dissect the contribution of tissue damage signaling to antimicrobial immune responses in a live vertebrate. By intravital imaging of zebrafish larvae, a powerful model for innate immunity, we show that prevention of tissue damage signaling upon microbial ear infection abrogates leukocyte chemotaxis and reduces animal survival, at least in part, through suppression of cytosolic phospholipase A2 (cPla2), which integrates tissue damage- and microbe-derived cues. Thus, microbial cues are insufficient, and damage signaling is essential for antimicrobial neutrophil responses in zebrafish.
Functional Alleles of Chicken BG Genes, Members of the Butyrophilin Gene Family, in Peripheral T Cells.
May 19, 2018   Frontiers In Immunology
Chen L, Fakiola M, Staines K, Butter C, Kaufman J
Functional Alleles of Chicken BG Genes, Members of the Butyrophilin Gene Family, in Peripheral T Cells.
May 19, 2018
Frontiers In Immunology
γδ T cells recognize a wide variety of ligands in mammals, among them members of the butyrophilin (BTN) family. Nothing is known about γδ T cell ligands in chickens, despite there being many such cells in blood and lymphoid tissues, as well as in mucosal surfaces. The major histocompatibility complex (MHC) of chickens was discovered because of polymorphic BG genes, part of the BTN family. All but two BG genes are located in the BG region, oriented head-to-tail so that unequal crossing-over has led to copy number variation (CNV) as well as hybrid (chimeric) genes, making it difficult to identify true alleles. One approach is to examine BG genes expressed in particular cell types, which likely have the same functions in different BG haplotypes and thus can be considered "functional alleles." We cloned nearly full-length BG transcripts from peripheral T cells of four haplotypes (B2, B15, B19, and B21), and compared them to the BG genes of the B12 haplotype that previously were studied in detail. A dominant BG gene was found in each haplotype, but with significant levels of subdominant transcripts in three haplotypes (B2, B15, and B19). For three haplotypes (B15, B19, and B21), most sequences are closely-related to BG8, BG9, and BG12 from the B12 haplotype. We found that variation in the extracellular immunoglobulin-variable-like (Ig-V) domain is mostly localized to the membrane distal loops but without evidence for selection. However, variation in the cytoplasmic tail composed of many amino acid heptad repeats does appear to be selected (although not obviously localized), consistent with an intriguing clustering of charged and polar residues in an apparent α-helical coiled-coil. By contrast, the dominantly-expressed BG gene in the B2 haplotype is identical to BG13 from the B12 haplotype, and most of the subdominant sequences are from the BG5-BG7-BG11 clade. Moreover, alternative splicing leading to intron read-through results in dramatically truncated cytoplasmic tails, particularly for the dominantly-expressed BG gene of the B2 haplotype. The approach of examining "functional alleles" has yielded interesting data for closely-related genes, but also thrown up unexpected findings for at least one haplotype.
RECON-Dependent Inflammation in Hepatocytes Enhances Listeria monocytogenes Cell-to-Cell Spread.
May 24, 2018   MBio
McFarland AP, Burke TP, Carletti AA, Glover RC, Tabakh H, Welch MD, Woodward JJ
RECON-Dependent Inflammation in Hepatocytes Enhances Listeria monocytogenes Cell-to-Cell Spread.
May 24, 2018
MBio
The oxidoreductase RECON is a high-affinity cytosolic sensor of bacterium-derived cyclic dinucleotides (CDNs). CDN binding inhibits RECON's enzymatic activity and subsequently promotes inflammation. In this study, we sought to characterize the effects of RECON on the infection cycle of the intracellular bacterium Listeria monocytogenes, which secretes cyclic di-AMP (c-di-AMP) into the cytosol of infected host cells. Here, we report that during infection of RECON-deficient hepatocytes, which exhibit hyperinflammatory responses, L. monocytogenes exhibits significantly enhanced cell-to-cell spread. Enhanced bacterial spread could not be attributed to alterations in PrfA or ActA, two virulence factors critical for intracellular motility and intercellular spread. Detailed microscopic analyses revealed that in the absence of RECON, L. monocytogenes actin tail lengths were significantly longer and there was a larger number of faster-moving bacteria. Complementation experiments demonstrated that the effects of RECON on L. monocytogenes spread and actin tail lengths were linked to its enzymatic activity. RECON enzyme activity suppresses NF-κB activation and is inhibited by c-di-AMP. Consistent with these previous findings, we found that augmented NF-κB activation in the absence of RECON caused enhanced L. monocytogenes cell-to-cell spread and that L. monocytogenes spread correlated with c-di-AMP secretion. Finally, we discovered that, remarkably, increased NF-κB-dependent inducible nitric oxide synthase expression and nitric oxide production were responsible for promoting L. monocytogenes cell-to-cell spread. The work presented here supports a model whereby L. monocytogenes secretion of c-di-AMP inhibits RECON's enzymatic activity, drives augmented NF-κB activation and nitric oxide production, and ultimately enhances intercellular spread.IMPORTANCE To date, bacterial CDNs in eukaryotes are solely appreciated for their capacity to activate cytosolic sensing pathways in innate immunity. However, it remains unclear whether pathogens that actively secrete CDNs benefit from this process. Here, we provide evidence that secretion of CDNs leads to enhancement of L. monocytogenes cell-to-cell spread. This is a heretofore-unknown role of these molecules and suggests L. monocytogenes may benefit from their secretion in certain contexts. Molecular characterization revealed that, surprisingly, nitric oxide was responsible for the enhanced spread. Pathogens act to prevent nitric oxide production or, like L. monocytogenes, they have evolved to resist its direct antimicrobial effects. This study provides evidence that intracellular bacterial pathogens not only tolerate nitric oxide, which is inevitably encountered during infection, but can also capitalize on the changes this pleiotropic molecule enacts on the host cell.
Influence of lipopolysaccharide on proinflammatory gene expression in human corneal, conjunctival and meibomian gland epithelial cells.
Jun 08, 2018   The Ocular Surface
Chen D, Sahin A, Kam WR, Liu Y, Darabad RR, Sullivan DA
Influence of lipopolysaccharide on proinflammatory gene expression in human corneal, conjunctival and meibomian gland epithelial cells.
Jun 08, 2018
The Ocular Surface
PURPOSE: Lipopolysaccharide (LPS), a bacterial endotoxin, is known to stimulate leuokotriene B4 (LTB4) secretion by human corneal (HCECs), conjunctival (HConjECs) and meibomian gland (HMGECs) epithelial cells. We hypothesize that this LTB4 effect represents an overall induction of proinflammatory gene expression in these cells. Our objective was to test this hypothesis. METHODS: Immortalized HCECs, HConjECs and HMGECs were cultured in the presence or absence of LPS (15 μg/ml) and ligand binding protein (LBP; 150 ng/ml). Cells were then processed for RNA isolation and the analysis of gene expression by using Illumina BeadChips, background subtraction, cubic spline normalization and GeneSifter software. RESULTS: Our findings show that LPS induces a striking increase in proinflammatory gene expression in HCECs and HConjECs. These cellular reactions are associated with a significant up-regulation of genes associated with inflammatory and immune responses (e.g. IL-1β, IL-8, and tumor necrosis factor), including those related to chemokine and Toll-like receptor signaling pathways, cytokine-cytokine receptor interactions, and chemotaxis. In contrast, with the exception of Toll-like signaling and associated innate immunity pathways, almost no proinflammatory ontologies were upregulated by LPS in HMGECs. CONCLUSIONS: Our results support our hypothesis that LPS stimulates proinflammatory gene expression in HCECs and HConjECs. However, our findings also show that LPS does not elicit such proinflammatory responses in HMGECs.
Polysaccharide PRM3 from Rhynchosia minima root enhances immune function through TLR4-NF-κB pathway.
Jun 08, 2018   Biochimica Et Biophysica Acta
Jia X, Liang Y, Zhang C, Wang K, Tu Y, Chen M, Li P, Wan JB, He C
Polysaccharide PRM3 from Rhynchosia minima root enhances immune function through TLR4-NF-κB pathway.
Jun 08, 2018
Biochimica Et Biophysica Acta
BACKGROUND: Polysaccharides, one of the active ingredients in herbal medicine, are proved to enhance innate immunity against infections. The aim of this study is to explore the immunoregulatory ability of polysaccharides from Rhynchosia minima root in vitro and in vivo. METHODS: Polysaccharide fractions of R. minima root were obtained by chromatographic column. The content of NO was measured by spectrophotometry. The levels of cytokines (tumor necrosis factor-α, TNF-α; interleukin-6, IL-6; and monocyte chemoattractant protein-1, MCP-1) were determined by enzyme-linked immuno-sorbent assay (ELISA) kits. The translocation of p65 into the nucleus was imaged by confocal microscopy. The mRNA expression of TNF-α, IL-6, and MCP-1 was determined by quantitative real-time PCR. T-lymphocyte subgroups of spleen from immunosuppressive mouse were evaluated by flow cytometry. RESULTS: PRM3 remarkably enhanced the phagocytic ability of macrophages and promoted the release of NO and the secretion of cytokines (TNF-α, IL-6, and MCP-1) from macrophages. Simultaneously, PRM3 potently activated NF-κB signaling pathway via Toll-like receptor 4 (TLR4). In addition, PRM3 obviously increased the levels of serum cytokines, markedly up-regulated the percentages of CD3+ and CD4+ T lymphocytes and the CD4+/CD8+ ratio of splenocytes, and effectively attenuated cyclophosphamide induced immunosuppression in mice. CONCLUSIONS: PRM3 profoundly enhanced the immune function in vitro and in vivo through TLR4-NF-κB pathway and is a promising candidate of immunopotentiator which could be applied in functional foods or drugs. GENERAL SIGNIFICANCE: This study reported a polysaccharide PRM3 from R. minima root exhibited potent immunoenhancing activity and significantly alleviated cyclophosphamide-induced immunosuppression through TLR4-NF-κB pathway.
Critical Role of Alternative M2 Skewing in miR-155 Deletion-Mediated Protection of Colitis.
May 20, 2018   Frontiers In Immunology
Li J, Zhang J, Guo H, Yang S, Fan W,   . . . . . .   , Yan P, Lin H, Luo X, Li H, Wu Y
Critical Role of Alternative M2 Skewing in miR-155 Deletion-Mediated Protection of Colitis.
May 20, 2018
Frontiers In Immunology
Inflammatory bowel disease (IBD) is associated with dysregulation of both innate and adaptive immune response in the intestine. MicroRNA (miR)-155 is frequently expressed and functions in many immune cell types. Besides its function in adaptive immunity, miR-155 is a key regulator of the innate immune response in macrophages, dendritic cells, and even in epithelia cells. Although the roles of miR-155 within T and B lymphocytes in colitis have been reported, its function in innate immune cells has not been thoroughly examined. In this study, the dextran sulfate sodium (DSS)-induced colitis model was established in wild-type (WT) and miR-155-/- mice. Our results showed that miR-155 deficiency in macrophages recapitulated the alleviated colitis feature of miR-155-/- mice and appeared to skew toward the alterative M2 phenotype. Notably, the predominance of M2 in colon can result in dampened intestinal immune cell proliferation and inhibit CD4 T cell polarization toward Th1 and Th17. Moreover, C/EBPβ and SOCS1 were demonstrated as two key functional targets in this process. We also provided evidence for use of miR-155 inhibitor to treat colitis. Collectively, the findings highlight the central role of alternative M2 skewing for miR-155 function in colitis and reveal that macrophages might be a main target for therapeutics.

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