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Blood Mar 2024Protease activated receptors (PARs) are cleaved by coagulation proteases and thereby connect hemostasis with innate immune responses. Signaling of the tissue factor (TF)...
Protease activated receptors (PARs) are cleaved by coagulation proteases and thereby connect hemostasis with innate immune responses. Signaling of the tissue factor (TF) complex with factor VIIa (FVIIa) via PAR2 stimulates extracellular signal-regulated kinase (ERK) activation and cancer cell migration, but functions of cell autonomous TF-FVIIa signaling in immune cells are unknown. Here, we show that myeloid cell expression of FVII but not of FX is crucial for inflammatory cell recruitment to the alveolar space after challenge with the double-stranded viral RNA mimic polyinosinic:polycytidylic acid [Poly(I:C)]. In line with these data, genetically modified mice completely resistant to PAR2 cleavage but not FXa-resistant PAR2-mutant mice are protected from lung inflammation. Poly(I:C)-stimulated migration of monocytes/macrophages is dependent on ERK activation and mitochondrial antiviral signaling (MAVS) but independent of toll-like receptor 3 (TLR3). Monocyte/macrophage-synthesized FVIIa cleaving PAR2 is required for integrin αMβ2-dependent migration on fibrinogen but not for integrin β1-dependent migration on fibronectin. To further dissect the downstream signaling pathway, we generated PAR2S365/T368A-mutant mice deficient in β-arrestin recruitment and ERK scaffolding. This mutation reduces cytosolic, but not nuclear ERK phosphorylation by Poly(I:C) stimulation, and prevents macrophage migration on fibrinogen but not fibronectin after stimulation with Poly(I:C) or CpG-B, a single-stranded DNA TLR9 agonist. In addition, PAR2S365/T368A-mutant mice display markedly reduced immune cell recruitment to the alveolar space after Poly(I:C) challenge. These results identify TF-FVIIa-PAR2-β-arrestin-biased signaling as a driver for lung infiltration in response to viral nucleic acids and suggest potential therapeutic interventions specifically targeting TF-VIIa signaling in thrombo-inflammation.
Topics: Animals; Mice; Factor VIIa; Monocytes; Thromboplastin; Receptor, PAR-2; Signal Transduction; Extracellular Signal-Regulated MAP Kinases; Fibrinogen; beta-Arrestins
PubMed: 38096370
DOI: 10.1182/blood.2023021149 -
Biology Methods & Protocols 2023Mucosal vaccine for sublingual route was prepared with recombinant SARS-CoV-2 spike protein receptor binding domain (RBD) antigen and poly(I:C) adjuvant components. The...
Mucosal vaccine for sublingual route was prepared with recombinant SARS-CoV-2 spike protein receptor binding domain (RBD) antigen and poly(I:C) adjuvant components. The efficacy of this sublingual vaccine was examined using Cynomolgus macaques. Nine of the macaque monkeys were divided into three groups of three animals: control [just 400 µg poly(I:C) per head], low dose [30 µg RBD and 400 µg poly(I:C) per head], and high dose [150 µg RBD and 400 µg poly(I:C) per head], respectively. N-acetylcysteine (NAC), a mild reducing agent losing mucin barrier, was used to enhance vaccine delivery to mucosal immune cells. RBD-specific IgA antibody secreted in pituita was detected in two of three monkeys of the high dose group and one of three animals of the low dose group. RBD-specific IgG and/or IgA antibodies in plasma were also detected in these monkeys. These indicated that the sublingual vaccine stimulated mucosal immune response to produce antigen-specific secretory IgA antibodies in pituita and/or saliva. This sublingual vaccine also affected systemic immune response to produce IgG (IgA) in plasma. Little RBD-specific IgE was detected in plasma, suggesting no allergic antigenicity of this sublingual vaccine. Thus, SARS-CoV-2 sublingual vaccine consisting of poly(I:C) adjuvant showed reasonable efficacy in a non-human primate model.
PubMed: 37711440
DOI: 10.1093/biomethods/bpad017 -
Cellular and Molecular Life Sciences :... Jul 2023DExD/H-box helicase (DDX) 5 belongs to the DExD/H-box helicase family. DDX family members play differential roles in the regulation of innate antiviral immune response....
DExD/H-box helicase (DDX) 5 belongs to the DExD/H-box helicase family. DDX family members play differential roles in the regulation of innate antiviral immune response. However, whether DDX5 is involved in antiviral immunity remains unclear. In this study, we found that DDX5 serves as a negative regulator of type I interferon (IFN) response. Overexpression of DDX5 inhibited IFN production induced by Spring viremia of carp virus (SVCV) and poly(I:C) and enhanced virus replication by targeting key elements of the RLR signaling pathway (MAVS, MITA, TBK1, IRF3 and IRF7). Mechanistically, DDX5 directly interacted with TBK1 to promote its autophagy-mediated degradation. Moreover, DDX5 was shown to block the interaction between TRAF3 and TBK1, hence preventing nuclear translocation of IRF3. Together, these data shed light on the roles of DDX5 in regulating IFN response.
Topics: Protein Serine-Threonine Kinases; TNF Receptor-Associated Factor 3; Phosphorylation; Dichlorodiphenyl Dichloroethylene; Immunity, Innate; Interferon Type I; Interferon Regulatory Factor-3; Antiviral Agents
PubMed: 37462751
DOI: 10.1007/s00018-023-04860-2 -
Advanced Science (Weinheim,... Nov 2023The efficient activation of professional antigen-presenting cells-such as dendritic cells (DC)-in tumors and lymph nodes is critical for the design of next-generation...
The efficient activation of professional antigen-presenting cells-such as dendritic cells (DC)-in tumors and lymph nodes is critical for the design of next-generation cancer vaccines and may be able to provide anti-tumor effects by itself through immune stimulation. The challenge is to stimulate these cells without causing excessive toxicity. It is hypothesized that a multi-pronged combinatorial approach to DC stimulation would allow dose reductions of innate immune receptor-stimulating TLR3 agonists while enhancing drug efficacy. Here, a hybrid lipid nanoparticle (LNP) platform is developed and tested for double-stranded RNA (polyinosinic:polycytidylic acid for TLR3 agonism) and immune modulator (L-CANDI) delivery. This study shows that the ≈120 nm hybrid nanoparticles-in-nanoparticles effectively eradicate tumors by themselves and generate long-lasting, durable anti-tumor immunity in mouse models.
Topics: Animals; Mice; Toll-Like Receptor 3; Poly I-C; Cancer Vaccines; Neoplasms; Dendritic Cells
PubMed: 37814359
DOI: 10.1002/advs.202303576 -
Frontiers in Immunology 2023Viral pneumonia is a global health burden with a high mortality rate, especially in the elderly and in patients with underlying diseases. Recent studies have found that...
Viral pneumonia is a global health burden with a high mortality rate, especially in the elderly and in patients with underlying diseases. Recent studies have found that myeloid-derived suppressor cells (MDSCs) are abundant in these patient groups; however, their roles in the progression of viral pneumonia remain unclear. In this study, we observed a substantial increase in MDSCs in a mouse model of renal ischemia/reperfusion (I/R) injury and in older mice. When intranasal polyinosinic-polycytidylic acid (poly(I:C)) administration was used to mimic viral pneumonia, mice with renal I/R injury exhibited more severe lung inflammation than sham mice challenged with poly(I:C). In addition, MDSC depletion attenuated lung inflammation in mice with I/R injury. Similar results were obtained in older mice compared with those in young mice. Furthermore, adoptive transfer of in -differentiated MDSCs exacerbated poly(I:C)-induced lung inflammation. Taken together, these experimental results suggest that the increased proportion of MDSCs in mice with renal I/R injury and in older mice exacerbates poly(I:C)-induced lung inflammation. These findings have important implications for the treatment and prevention of severe lung inflammation caused by viral pneumonia.
Topics: Humans; Mice; Animals; Aged; Myeloid-Derived Suppressor Cells; Poly I-C; Kidney; Disease Models, Animal; Pneumonia, Viral
PubMed: 37818369
DOI: 10.3389/fimmu.2023.1243851 -
Drug Delivery and Translational Research Mar 2024Melanoma is the main cause of death among skin cancers and its incidence worldwide has been experiencing an appalling increase. However, traditional treatments lack...
Melanoma is the main cause of death among skin cancers and its incidence worldwide has been experiencing an appalling increase. However, traditional treatments lack effectiveness in advanced or metastatic patients. Immunotherapy, meanwhile, has been shown to be an effective treatment option, but the rate of cancers responding remains far from ideal. Here we have developed a personalized neoantigen peptide-based cancer vaccine by encapsulating patient derived melanoma neoantigens in polyethylenimine (PEI)-functionalised poly(lactic-co-glycolic acid) (PLGA) nanoparticles (NPs) and coating them with polyinosinic:polycytidylic acid (poly(I:C)). We found that PLGA NPs can be effectively modified to be coated with the immunoadjuvant poly(I:C), as well as to encapsulate neoantigens. In addition, we found that both dendritic cells (DCs) and lymphocytes were effectively stimulated. Moreover, the developed NP was found to have a better immune activation profile than NP without poly(I:C) or without antigen. Our results demonstrate that the developed vaccine has a high capacity to activate the immune system, efficiently maturing DCs to present the antigen of choice and promoting the activity of lymphocytes to exert their cytotoxic function. Therefore, the immune response generated is optimal and specific for the elimination of melanoma tumour cells.
PubMed: 38427275
DOI: 10.1007/s13346-024-01557-2 -
Heliyon Dec 2023Viruses have become a major threat to human health. Interferon-β (IFN-β) has a key role in the antivirus process, as it can increase the expression of...
Viruses have become a major threat to human health. Interferon-β (IFN-β) has a key role in the antivirus process, as it can increase the expression of antivirus-associated genes. Itaconate and its derivatives can regulate the immune response, secretion of inflammatory factors, and pyroptosis of macrophages. The effect of itaconate on IFN-β secretion of double-stranded RNA-induced macrophages are not well known. A derivative of itaconate, 4-octoyl itaconate (4-OI), was used to treat mouse bone marrow-derived macrophages (BMDM) induced with 100 μg/mL poly(I:C). The IFN-β concentration was detected through ELISA, and IFN-β mRNA expression was detected through quantitative PCR. High-throughput transcriptome sequencing was used to analyze changes in the BMDM transcriptome after 4-OI treatment. The Nrf2 expression was knocked down with siRNA.4-OI inhibited poly(I:C)-induced IFN-β secretion and mRNA expression in BMDM. Results of transcriptome sequencing revealed that 4-OI downregulated 1047 genes and upregulated 822 genes. GO and KEGG enrichment of differently expressed genes revealed that many downregulated genes were related to the anti-virus process, whereas many upregulated genes were related to metabolism. The Nrf2 inhibitor ML385 and Nrf2 siRNA could partially reverse the inhibitory effect of 4-OI. In conclusion, 4-octyl itaconate could inhibit the poly(I:C)-induced interferon-β secretion in BMDM partially by regulating Nrf2.
PubMed: 38076131
DOI: 10.1016/j.heliyon.2023.e23001 -
Fish & Shellfish Immunology Jan 2024Poly(I:C) is known as an agonist of the TLR3 receptor which could prime inflammation and elicit the host immune response, which is widely applied as adjuvant or...
Poly(I:C) is known as an agonist of the TLR3 receptor which could prime inflammation and elicit the host immune response, which is widely applied as adjuvant or antivirus treatment. However, the negative effects of poly(I:C) on regulating immune response to protect the host from inflammatory diseases remain largely unknown. Here, we establish an in vivo model to pre-treat zebrafish larvae with poly(I:C) at 2 dpf, then challenge them with LPS at 6 dpf, and find that poly(I:C) training could significantly alleviate the LPS challenge-induced septic shock and inflammatory phenotypes. Moreover, the poly(I:C)-trained larvae exhibit decreased number of macrophages, but not neutrophils, after secondary LPS challenge. Furthermore, training the larvae with poly(I:C) could elevate the transcripts of mTOR signaling and heighten the H3K4me3-mediated epigenetic modifications. And interestingly, we find that inhibiting the H3K4me3 modification, rather than mTOR signaling, could recover the number of macrophages in poly(I:C)-trained larvae, which is consistent with the observations of inflammatory phenotypes. Taken together, these results suggest that poly(I:C) training could induce epigenetic rewiring to mediate the anti-inflammatory response against secondary LPS challenge-induced septic shock through decreasing macrophages' number in vivo, which might expand our understanding of poly(I:C) in regulating fish immune response.
Topics: Animals; Lipopolysaccharides; Zebrafish; Larva; Shock, Septic; Inflammation; Anti-Inflammatory Agents; TOR Serine-Threonine Kinases
PubMed: 38092095
DOI: 10.1016/j.fsi.2023.109285 -
Pancreatology : Official Journal of the... Nov 2023IgG4-related autoimmune pancreatitis (AIP) is considered to be a T cell-mediated autoimmune disease. However, CD8 T cells have only received brief mention, and have yet...
BACKGROUND
IgG4-related autoimmune pancreatitis (AIP) is considered to be a T cell-mediated autoimmune disease. However, CD8 T cells have only received brief mention, and have yet to be completely studied. The study aimed to investigate the expression of signaling lymphocytic activation molecule family 7 (SLAMF7) on CD8 T cells and the features of SLAMF7CD8 T cells in MRL/Mp mice with AIP.
METHODS
A murine model of AIP was established by intraperitoneal injection with polyinosinic:polycytidylic acid (poly I:C) for 8 weeks. Dexamethasone treatment was daily administrated for the last 2 weeks during a 6-week course of poly I:C. SLAMF7 expression on CD8 T cells in the spleen and pancreas was detected by flow cytometry. Granzyme B (GZMB) and cytokines including IFN-γ, TNF-α, and IL-2, were monitored in an in vitro T cell activation assay. Dexamethasone suppression assays were performed to downregulate SLAMF7 expression on T cells upon T cell receptor stimulation.
RESULTS
AIP in MRL/Mp mice was induced by repeated intraperitoneal administration of poly I:C and CD8 T cells were increased in the inflamed pancreas. SLAMF7CD8 T cells were elevated in the spleen and pancreas of AIP mice. SLAMF7CD8 T subsets produced more GZMB, IFN-γ, TNF-α and IL-2 than SLAMF7CD8 T subsets. Dexamethasone treatment ameliorated pancreatic inflammatory and fibrosis of AIP. Dexamethasone could downregulate SLAMF7CD8 T cells and reduce GZMB, IFN-γ and TNF-α levels both in vitro and in vivo.
CONCLUSIONS
Increased SLAMF7CD8 T cells exhibit enhanced cytotoxicity and cytokines secretion capacity, which may be involved in the pathogenesis of AIP.
Topics: Mice; Animals; Autoimmune Pancreatitis; CD8-Positive T-Lymphocytes; Interleukin-2; Tumor Necrosis Factor-alpha; Autoimmune Diseases; Poly I-C; Dexamethasone; Signaling Lymphocytic Activation Molecule Family
PubMed: 37661465
DOI: 10.1016/j.pan.2023.08.005 -
ACS Pharmacology & Translational Science May 2024Polyriboinosinic acid-polyribocytidylic acid (Poly I:C) serves as a synthetic mimic of viral double-stranded dsRNA, capable of inducing apoptosis in numerous cancer...
Polyriboinosinic acid-polyribocytidylic acid (Poly I:C) serves as a synthetic mimic of viral double-stranded dsRNA, capable of inducing apoptosis in numerous cancer cells. Despite its potential, therapeutic benefits, the application of Poly I:C has been hindered by concerns regarding toxicity, stability, enzymatic degradation, and undue immune stimulation, leading to autoimmune disorders. To address these challenges, encapsulation of antitumor drugs within delivery systems such as cationic liposomes is often employed to enhance their efficacy while minimizing dosages. In this study, we investigated the potential of cationic liposomes to deliver Poly I:C into the Head and Neck 12 (HN12) cell line to induce apoptosis in the carcinoma cells and tumor model. Cationic liposomes made by the hydrodynamic focusing method surpass traditional methods by offering a continuous flow-based approach for encapsulating genes, which is ideal for efficient tumor delivery. DOTAP liposomes efficiently bind Poly I:C, confirmed by transmission electron microscopy images displaying their spherical morphology. Liposomes are easily endocytosed in HN12 cells, suggesting their potential for therapeutic gene and drug delivery in head and neck squamous carcinoma cells. Activation of apoptotic pathways involving MDA5, RIG-I, and TLR3 is evidenced by upregulated caspase-3, caspase-8, and IRF3 genes upon endocytosis of Poly(I:C)-encapsulated liposomes. Therapeutic evaluations revealed significant inhibition of tumor growth with Poly I:C liposomes, indicating the possibility of MDA5, RIG-I, and TLR3-induced apoptosis pathways via Poly I:C liposomes in HN12 xenografts in J:NU mouse models. Comparative histological analysis underscores enhanced cell death with Poly I:C liposomes, warranting further investigation into the precise mechanisms of apoptosis and inflammatory cytokine response in murine models for future research.
PubMed: 38751634
DOI: 10.1021/acsptsci.4c00121