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Frontiers in Immunology 2018Extracellular vesicles (EV) can modulate the responses of cells to toll-like receptor (TLR) ligation; conversely, TLR ligands such as double-stranded RNA (dsRNA) can...
Extracellular vesicles (EV) can modulate the responses of cells to toll-like receptor (TLR) ligation; conversely, TLR ligands such as double-stranded RNA (dsRNA) can enhance the release of EV and influence of the composition and functions of EV cargos. Inflamed synovial joints in rheumatoid arthritis (RA) are rich in EV and extracellular RNA; besides, RNA released from necrotic synovial fluid cells can activate the TLR3 signaling in synovial fibroblasts (SFs) from patients with RA. Since EV occur prominently in synovial joints in RA and may contribute to the pathogenesis, we questioned whether EV can interact with dsRNA, a TLR3 ligand, and modify its actions in arthritis. We have used as model the effects on RA SFs, of EV released from monocyte U937 cells and peripheral blood mononuclear cells upon stimulation with Poly(I:C), a synthetic analog of dsRNA. We show that EV released from unstimulated cells and Poly(I:C)-stimulated U937 cells [Poly(I:C) EV] differ in size but bind similar amounts of Annexin V and express comparable levels of MAC-1, the receptor for dsRNA, on the vesicular membranes. Specifically, Poly(I:C) EV contain or associate with Poly(I:C) and at least partially protect Poly(I:C) from RNAse III degradation. Poly(I:C) EV shuttle Poly(I:C) to SFs and reproduce the proinflammatory and antiviral gene responses of SFs to direct stimulation with Poly(I:C). Poly(I:C) EV, however, halt the death receptor-induced apoptosis in SFs, thereby inverting the proapoptotic nature of Poly(I:C). These prosurvival effects sharply contrast with the high toxicity of cationic liposome-delivered Poly(I:C) and may reflect the route of Poly(I:C) delivery EV or the fine-tuning of Poly(I:C) actions by molecular cargo in EV. The demonstration that EV may safeguard extracellular dsRNA and allow dsRNA to exert antiapoptotic effects on SFs highlights the potential of EV to amplify the pathogenicity of dsRNA in arthritis beyond inflammation (by concurrently enhancing the expansion of the invasive synovial stroma).
Topics: Annexin A5; Arthritis, Rheumatoid; Cell Line, Tumor; Extracellular Vesicles; Fibroblasts; Humans; Macrophage-1 Antigen; Poly I-C; RNA, Double-Stranded; Ribonuclease III; Signal Transduction; Synovial Membrane; Toll-Like Receptor 3; U937 Cells
PubMed: 29434584
DOI: 10.3389/fimmu.2018.00028 -
Frontiers in Immunology 2022Retinoic acid inducible gene-I (RIG-I)-like receptors (RLRs) are viral RNA sensors that regulate host interferon (IFN)-mediated antiviral signaling. LGP2 (laboratory...
Retinoic acid inducible gene-I (RIG-I)-like receptors (RLRs) are viral RNA sensors that regulate host interferon (IFN)-mediated antiviral signaling. LGP2 (laboratory genetics and physiology 2) lacks the N-terminal caspase activation and recruitment domains (CARDs) responsible for signaling transduction in the other two RLR proteins, RIG-I and melanoma differentiation associated gene-5 (MDA5). How LGP2 regulates IFN signaling is controversial, and inconsistent results have often been obtained in overexpression assays when performed in fish cells and mammalian cells. Here we report that the differential sensitivity of fish cells and mammalian cells to poly(I:C) transfection conceals the function conservation of zebrafish and human LGP2. In fish cells, overexpression of zebrafish or human LGP2 initially activates IFN signaling in a dose-dependent manner, followed by inhibition at a critical threshold of LGP2 expression. A similar trend exists for LGP2-dependent IFN induction in response to stimulation by low and high concentrations of poly(I:C). In contrast, overexpression of zebrafish or human LGP2 alone in mammalian cells does not activate IFN signaling, but co-stimulation with very low or very high concentrations of poly(I:C) shows LGP2-dependent enhancement or inhibition of IFN signaling, respectively. Titration assays show that LGP2 promotes MDA5 signaling in mammalian cells mainly under low concentration of poly(I:C) and inhibits RIG-I/MDA5 signaling mainly under high concentration of poly(I:C). Our results suggest that fish and human LGP2s switch regulatory roles from a positive one to a negative one in increasing concentrations of poly(I:C)-triggered IFN response.
Topics: Animals; Antiviral Agents; Humans; Interferon-Induced Helicase, IFIH1; Interferons; Mammals; Poly I-C; RNA Helicases; Zebrafish
PubMed: 36059486
DOI: 10.3389/fimmu.2022.985792 -
Brain, Behavior, and Immunity Nov 2019The viral mimetic polyinosinic:polycytidylic acid (poly(I:C)) is increasingly used to induce maternal immune activation (mIA) to model neurodevelopmental disorders...
The viral mimetic polyinosinic:polycytidylic acid (poly(I:C)) is increasingly used to induce maternal immune activation (mIA) to model neurodevelopmental disorders (NDDs). Robust and reproducible phenotypes across studies are essential for the generation of models that will enhance our understanding of NDDs and enable the development of improved therapeutic strategies. However, differences in mIA-induced phenotypes using poly(I:C) have been widely observed, and this has prompted the reporting of useful and much needed methodological guidelines. Here, we perform a detailed investigation of molecular weight and endotoxin variations in poly(I:C) procured from two of the most commonly used suppliers, Sigma and InvivoGen. We demonstrate that endotoxin contamination and molecular weight differences in poly(I:C) composition lead to considerable variability in maternal IL-6 response in rats treated on gestational day (GD)15 and impact on fetal outcomes. Specifically, both endotoxin contamination and molecular weight predicted reductions in litter size on GD21. Further, molecular weight predicted a reduction in placental weight at GD21. While fetal body weight at GD21 was not affected by poly(I:C) treatment, male fetal brain weight was significantly reduced by poly(I:C), dependent on supplier. Our data are in agreement with recent reports of the importance of poly(I:C) molecular weight, and extend this work to demonstrate a key role of endotoxin on relevant phenotypic outcomes. We recommend that the source and batch numbers of poly(I:C) used should always be stated and that molecular weight variability and endotoxin contamination should be minimised for more robust mIA modelling.
Topics: Animals; Behavior, Animal; Cytokines; Endotoxins; Female; Fetus; Infectious Disease Transmission, Vertical; Litter Size; Male; Maternal Exposure; Neurodevelopmental Disorders; Poly I-C; Pregnancy; Prenatal Exposure Delayed Effects; Rats; Rats, Wistar; Reproducibility of Results
PubMed: 31415868
DOI: 10.1016/j.bbi.2019.08.006 -
Frontiers in Immunology 2023Both bacterial and viral diseases are a major threat to farmed fish. As the antiviral immune mechanisms in lumpfish ( L.) are poorly understood, lumpfish leukocytes were...
BACKGROUND
Both bacterial and viral diseases are a major threat to farmed fish. As the antiviral immune mechanisms in lumpfish ( L.) are poorly understood, lumpfish leukocytes were stimulated with poly(I:C), a synthetic analog of double stranded RNA, which mimic viral infections, and RNA sequencing was performed.
METHODS
To address this gap, we stimulated lumpfish leukocytes with poly(I:C) for 6 and 24 hours and did RNA sequencing with three parallels per timepoint. Genome guided mapping was performed to define differentially expressed genes (DEGs).
RESULTS
Immune genes were identified, and transcriptome-wide analyses of early immune responses showed that 376 and 2372 transcripts were significantly differentially expressed 6 and 24 hours post exposure (hpe) to poly(I:C), respectively. The most enriched GO terms when time had been accounted for, were immune system processes (GO:0002376) and immune response (GO:0006955). Analysis of DEGs showed that among the most highly upregulated genes were TLRs and genes belonging to the RIG-I signaling pathway, including LGP2, STING and MX, as well as IRF3 and IL12A. RIG-I was not identified, but analyses showed that genes encoding proteins involved in pathogen recognition, cell signaling, and cytokines of the TLR and RIG-I signaling pathway are mostly conserved in lumpfish when compared to mammals and other teleost species.
CONCLUSIONS
Our analyses unravel the innate immune pathways playing a major role in antiviral defense in lumpfish. The information gathered can be used in comparative studies and lay the groundwork for future functional analyses of immune and pathogenicity mechanisms. Such knowledge is also necessary for the development of immunoprophylactic measures for lumpfish, which is extensively cultivated for use as cleaner fish in the aquaculture for removal of sea lice from Atlantic salmon ( L.).
Topics: Animals; Transcriptome; Poly I-C; Perciformes; Antiviral Agents; Immunity; Mammals
PubMed: 37388730
DOI: 10.3389/fimmu.2023.1198211 -
Zoological Science Apr 2022Obscure puffer, , is an important aquaculture species in China, but the disease problem of this species seriously affects its production and causes huge economic losses....
Obscure puffer, , is an important aquaculture species in China, but the disease problem of this species seriously affects its production and causes huge economic losses. In order to reveal the molecular mechanism of disease resistance, polyinosinic-polycytidylic acid [poly(I:C)] was used to stimulate obscure puffer. At 0, 12, and 48 h (named To0, To12, and To48) after poly(I:C) challenge, the kidneys from obscure puffer were collected for transcriptome sequencing. A total of 54,816 transcripts was generated. Pairwise comparison of the sequencing libraries of tissue samples at these three time points revealed that the number of differentially expressed genes (DEGs) at To12 vs To0, To48 vs To0, and To48 vs To12 were 2039, 776, and 2579, respectively. Gene Ontology (GO) function classification analysis revealed that some DEGs were annotated to GO items for membrane, biological process, molecular function, and metabolic process. Kyoto Encyclopedia of Genes and Genomes pathway (KEGG) analysis of DEGs demonstrated that they mainly presented in immune-related pathways, such as Toll-like receptor signaling pathway, Retinoic acid-inducible gene-I-like receptor signaling pathway and NOD-like receptor signaling pathway. Then, eight genes were randomly selected from immune-related genes for real-time quantitative reverse transcription PCR verification (RT-qPCR), and 22 key immune DEGs were used to construct network functions. This study has obtained a large number of information resources about the transcriptome of obscure puffer, which can provide references for further research on the anti-virus response of obscure puffer.
Topics: Animals; Gene Expression Profiling; Kidney; Poly I-C; Takifugu; Transcriptome
PubMed: 35380191
DOI: 10.2108/zs210070 -
Contact Dermatitis Oct 2023Poly(I:C) is recognised by endosomal Toll-like receptor 3 (TLR3) and activates cytotoxic CD8(+) lymphocytes and natural killer (NK) cells. It has been shown that the...
INTRODUCTION
Poly(I:C) is recognised by endosomal Toll-like receptor 3 (TLR3) and activates cytotoxic CD8(+) lymphocytes and natural killer (NK) cells. It has been shown that the viral TLR3 agonist induces robust and long-lasting T-cell-mediated responses. In addition, TLR3 modulates the contact hypersensitivity reaction.
OBJECTIVE
This study aimed to determine whether poly(I:C) injection can induce NK-mediated hapten reactivity in mice.
METHODS
Mice were treated with poly(I:C), and their response to dinitrofluorobenzene hapten was measured by assessing ear swelling and serum interferon gamma (IFN-γ) production. Adoptive cell transfer and cell sorting were used to investigate the mechanism of the reaction, and the phenotype of poly(I:C)-activated liver NK cells was determined by flow cytometry analysis.
RESULTS
The results showed that poly(I:C) administration increased ear swelling, serum IFN-γ levels and the response to hapten in both immunocompetent and T- and B-cell-deficient mice. Only liver poly(I:C)-activated DX5(+) NK cells were able to transfer reactivity to hapten into a naive recipient. Induction of liver NK cells after poly(I:C) administration was TLR3/TRIF- and IFN-γ-dependent, interleukin 12-independent, and not modulated by MyD88.
CONCLUSION
This study provides new insights into how poly(I:C) stimulates NK-mediated reactivity to hapten and suggests that liver NK cells may modulate the immune response to non-pathogenic factors during viral infection.
Topics: Mice; Animals; Toll-Like Receptor 3; Ligands; Dermatitis, Allergic Contact; Killer Cells, Natural; Poly I-C; Interferon-gamma; Mice, Inbred C57BL
PubMed: 37463838
DOI: 10.1111/cod.14380 -
Current Topics in Behavioral... 2023The epidemiological literature reporting increased risk for neurodevelopmental and psychiatric disorders after prenatal exposure to maternal immune activation (MIA) is... (Review)
Review
The epidemiological literature reporting increased risk for neurodevelopmental and psychiatric disorders after prenatal exposure to maternal immune activation (MIA) is still evolving, and so are the attempts to model this association in animals. Epidemiological studies of MIA offer the advantage of directly evaluating human populations but are often limited in their ability to uncover pathogenic mechanisms. Animal models, on the other hand, are limited in their generalizability to psychiatric disorders but have made significant strides toward discovering causal relationships and biological pathways between MIA and neurobiological phenotypes. Like in any other model system, both planned and unplanned sources of variability exist in animal models of MIA. Therefore, the design, implementation, and interpretation of MIA models warrant a careful consideration of these sources, so that appropriate strategies can be developed to handle them satisfactorily. While every research group may have its own strategy to this aim, it is essential to report the methodological details of the chosen MIA model in order to enhance the transparency and comparability of models across research laboratories. Even though it poses a challenge for attempts to compare experimental findings across laboratories, variability does not undermine the utility of MIA models for translational research. In fact, variability and heterogenous outcomes in MIA models offer unique opportunities for new discoveries and developments in this field, including the identification of disease pathways and molecular mechanisms determining susceptibility and resilience to MIA. This review summarizes the most important sources of variability in animal models of MIA and discusses how model variability can be used to investigate neurobiological and immunological factors causing phenotypic heterogeneity in offspring exposed to MIA.
Topics: Humans; Pregnancy; Female; Animals; Behavior, Animal; Poly I-C; Disease Models, Animal; Prenatal Exposure Delayed Effects; Mental Disorders
PubMed: 36306055
DOI: 10.1007/7854_2022_398 -
Journal of Visualized Experiments : JoVE Aug 2022Maternal immune activation (MIA) during pregnancy is consistently linked to increased risk of neurodevelopmental and neuropsychiatric disorders in offspring. Animal...
Maternal immune activation (MIA) during pregnancy is consistently linked to increased risk of neurodevelopmental and neuropsychiatric disorders in offspring. Animal models of MIA are used to test causality, investigate mechanisms, and develop diagnostics and treatments for these disorders. Despite their widespread use, many MIA models suffer from a lack of reproducibility and almost all ignore two important aspects of this risk factor: (i) many offspring are resilient to MIA, and (ii) susceptible offspring can exhibit distinct combinations of phenotypes. To increase reproducibility and model both susceptibility and resilience to MIA, the baseline immunoreactivity (BIR) of female mice before pregnancy is used to predict which pregnancies will result in either resilient offspring or offspring with defined behavioral and molecular abnormalities after exposure to MIA. Here, a detailed method of inducing MIA via intraperitoneal (i.p.) injection of the double stranded RNA (dsRNA) viral mimic poly(I:C) at 12.5 days of gestation is provided. This method induces an acute inflammatory response in the dam, which results in perturbations in brain development in mice that map onto similarly impacted domains in human psychiatric and neurodevelopmental disorders (NDDs).
Topics: Animals; Behavior, Animal; Disease Models, Animal; Female; Humans; Mice; Poly I-C; Pregnancy; Prenatal Exposure Delayed Effects; Reproducibility of Results
PubMed: 36063000
DOI: 10.3791/64095 -
Pharmacology, Biochemistry, and Behavior Aug 2023Maternal polyinosinic-polycytidylic acid (Poly I:C) exposure leads to an increase in various proinflammatory cytokines and causes schizophrenia-like symptoms in...
RATIONALE
Maternal polyinosinic-polycytidylic acid (Poly I:C) exposure leads to an increase in various proinflammatory cytokines and causes schizophrenia-like symptoms in offspring. In recent years, group I metabotropic glutamate receptors (mGluRs) have emerged as a potential target in the pathophysiology of schizophrenia.
OBJECTIVES
The aim of our study was to investigate the behavioral and molecular changes by using the mGlu1 receptor positive allosteric modulator (PAM) agent RO 67-7476, and the negative allosteric modulator (NAM) agent JNJ 16259685 and the mGlu5 receptor PAM agent VU-29, and NAM agent fenobam in the Poly I:C-induced schizophrenia model in rats.
METHODS
Female Wistar albino rats were treated with Poly I:C on day 14 of gestation after mating. On the postnatal day (PND) 34-35, 56-57 and 83-84, behavioral tests were performed in the male offspring. On the PND84, brain tissue was collected and the level of proinflammatory cytokines was determined by ELISA method.
RESULTS
Poly I:C caused impairments in all behavioral tests and increased the levels of proinflammatory cytokines. While PAM agents caused significant improvements in prepulse inhibition (PPI), novel object recognition (NOR), spontaneous alternation and reference memory tests, they brought the levels of proinflammatory cytokines closer to the control group. NAM agents were ineffective on behavioral tests. It was observed that PAM agents significantly improved Poly I:C-induced disruption in behavioral and molecular analyses.
CONCLUSIONS
These results suggest that PAM agents, particularly the mGlu5 receptor VU-29, are also promising and could be a potential target in schizophrenia.
Topics: Rats; Animals; Male; Female; Rats, Wistar; Schizophrenia; Poly I-C; Brain; Prepulse Inhibition; Allosteric Regulation
PubMed: 37390974
DOI: 10.1016/j.pbb.2023.173593 -
Ecotoxicology and Environmental Safety Dec 2022Pathogenic microorganisms that are ubiquitous in the environment threaten human health and food safety. Polyinosinic:polycytidylic acid (Poly (I:C)) is a macromolecule...
Pathogenic microorganisms that are ubiquitous in the environment threaten human health and food safety. Polyinosinic:polycytidylic acid (Poly (I:C)) is a macromolecule with a double-stranded RNA structure, which is often used to simulate viruses. Our previous study found that Poly (I:C) maternal stimulation could affect the reproduction of laying hens and their offspring, but the underlying mechanism needed to be explored. In the present study, splenic transcriptomes were sequenced and analyzed from two groups (Poly (I:C) treatment as the challenged group and saline treatment as the control) and in three generations (maternal stimulated F0 hens, unchallenged F1 and F2 generations). The results showed that Poly (I:C) maternal stimulation affected gene expression patterns in laying hens and their offspring. A total of 27 differentially expressed genes (DEGs) with the same regulating trend were discovered in the F0 and F1 generations, indicating an influence of the intergenerational transmission effect. Functional enrichment analysis of Gene Ontology (GO) showed that lymphocyte differentiation, positive regulation of leukocyte differentiation, positive regulation of MAPK cascade, and T cell differentiation were the common biological processes between F0 and F1 generations, revealing Poly (I:C) could affect the immunity of the treated F0 hens and the unchallenged subsequent generations. Further study showed that pathways associated with growth, development, biosynthesis, and metabolism of F2 chicks were also affected by Poly (I:C) maternal stimulation. Correlation analysis between DEGs and reproductive traits revealed that PHLDA2 (pleckstrin homology-like domain family A member 2) and PODN (podocan) with inheritable effect were highly correlated with egg-laying rate and egg weight in F1 hens, suggesting their potential long-term role in regulating reproductive traits. ARHGAP40, FGB, HRH4, PHLDA2, PODN, NTSR1, and NMU were supposed to play important roles in regulating chickens' immunity and reproductive traits. This study reveals the far-reaching effect on transcriptome induced by Poly (I:C), reflecting the influence of the mother's living environment on the offspring. It is an important reference for future research into the multi-generational transmission of maternal stimulation and harmful environmental factors.
Topics: Humans; Animals; Female; Chickens; Poly I-C; Reproduction; Oviposition; Transcriptome
PubMed: 36288637
DOI: 10.1016/j.ecoenv.2022.114216