Your browser doesn't support javascript.
Show: 20 | 50 | 100
Results 1 - 9 de 9
Filter
1.
Sci Rep ; 11(1): 24432, 2021 12 24.
Article in English | MEDLINE | ID: covidwho-1585772

ABSTRACT

Despite the initial success of some drugs and vaccines targeting COVID-19, understanding the mechanism underlying SARS-CoV-2 disease pathogenesis remains crucial for the development of further approaches to treatment. Some patients with severe Covid-19 experience a cytokine storm and display evidence of inflammasome activation leading to increased levels of IL-1ß and IL-18; however, other reports have suggested reduced inflammatory responses to Sars-Cov-2. In this study we have examined the effects of the Sars-Cov-2 envelope (E) protein, a virulence factor in coronaviruses, on inflammasome activation and pulmonary inflammation. In cultured macrophages the E protein suppressed inflammasome priming and NLRP3 inflammasome activation. Similarly, in mice transfected with E protein and treated with poly(I:C) to simulate the effects of viral RNA, the E protein, in an NLRP3-dependent fashion, reduced expression of pro-IL-1ß, levels of IL-1ß and IL-18 in broncho-alveolar lavage fluid, and macrophage infiltration in the lung. To simulate the effects of more advanced infection, macrophages were treated with both LPS and poly(I:C). In this setting the E protein increased NLRP3 inflammasome activation in both murine and human macrophages. Thus, the Sars-Cov-2 E protein may initially suppress the host NLRP3 inflammasome response to viral RNA while potentially increasing NLRP3 inflammasome responses in the later stages of infection. Targeting the Sars-Cov-2 E protein especially in the early stages of infection may represent a novel approach to Covid-19 therapy.


Subject(s)
Coronavirus Envelope Proteins/metabolism , Inflammasomes/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , SARS-CoV-2/metabolism , Animals , Bronchoalveolar Lavage Fluid/chemistry , COVID-19/pathology , COVID-19/virology , Coronavirus Envelope Proteins/genetics , Down-Regulation/drug effects , Endoplasmic Reticulum Stress , Humans , Inflammasomes/drug effects , Interleukin-1beta/genetics , Interleukin-1beta/metabolism , Janus Kinases/genetics , Janus Kinases/metabolism , Lipopolysaccharides/pharmacology , Macrophages/cytology , Macrophages/drug effects , Macrophages/metabolism , Mice , Mice, Inbred C57BL , Mice, Knockout , NLR Family, Pyrin Domain-Containing 3 Protein/deficiency , NLR Family, Pyrin Domain-Containing 3 Protein/genetics , Poly I-C/pharmacology , RNA, Viral/metabolism , SARS-CoV-2/drug effects , SARS-CoV-2/isolation & purification
2.
Chem Commun (Camb) ; 57(93): 12476-12479, 2021 Nov 23.
Article in English | MEDLINE | ID: covidwho-1500757

ABSTRACT

We identified small-molecule enhancers of cellular stress granules by observing molecular crowding of proteins and RNAs in a time-dependent manner. Hit molecules sensitized the IRF3-mediated antiviral mechanism in the presence of poly(I:C) and inhibited the replication of SARS-CoV-2 by inducing stress granule formation. Thus, modulating multimolecular crowding can be a promising strategy against SARS-CoV-2.


Subject(s)
Antiviral Agents/pharmacology , Benzopyrans/pharmacology , Cytoplasmic Granules/drug effects , Pyrazoles/pharmacology , SARS-CoV-2/drug effects , Virus Replication/drug effects , Animals , Antiviral Agents/chemistry , Benzopyrans/chemistry , Cell Line, Tumor , Chlorocebus aethiops , Cytoplasmic Granules/metabolism , Dose-Response Relationship, Drug , Drug Combinations , Humans , Interferon Regulatory Factor-3/metabolism , Lopinavir/pharmacology , Microbial Sensitivity Tests , Molecular Structure , Poly I-C/pharmacology , Pyrazoles/chemistry , Structure-Activity Relationship , Vero Cells
3.
J Immunol ; 207(9): 2310-2324, 2021 11 01.
Article in English | MEDLINE | ID: covidwho-1497461

ABSTRACT

IFN-γ, a proinflammatory cytokine produced primarily by T cells and NK cells, activates macrophages and engages mechanisms to control pathogens. Although there is evidence of IFN-γ production by murine macrophages, IFN-γ production by normal human macrophages and their subsets remains unknown. Herein, we show that human M1 macrophages generated by IFN-γ and IL-12- and IL-18-stimulated monocyte-derived macrophages (M0) produce significant levels of IFN-γ. Further stimulation of IL-12/IL-18-primed macrophages or M1 macrophages with agonists for TLR-2, TLR-3, or TLR-4 significantly enhanced IFN-γ production in contrast to the similarly stimulated M0, M2a, M2b, and M2c macrophages. Similarly, M1 macrophages generated from COVID-19-infected patients' macrophages produced IFN-γ that was enhanced following LPS stimulation. The inhibition of M1 differentiation by Jak inhibitors reversed LPS-induced IFN-γ production, suggesting that differentiation with IFN-γ plays a key role in IFN-γ induction. We subsequently investigated the signaling pathway(s) responsible for TLR-4-induced IFN-γ production in M1 macrophages. Our results show that TLR-4-induced IFN-γ production is regulated by the ribosomal protein S6 kinase (p70S6K) through the activation of PI3K, the mammalian target of rapamycin complex 1/2 (mTORC1/2), and the JNK MAPK pathways. These results suggest that M1-derived IFN-γ may play a key role in inflammation that may be augmented following bacterial/viral infections. Moreover, blocking the mTORC1/2, PI3K, and JNK MAPKs in macrophages may be of potential translational significance in preventing macrophage-mediated inflammatory diseases.


Subject(s)
Interferon-gamma/biosynthesis , Macrophages/drug effects , Poly I-C/pharmacology , COVID-19/immunology , Humans , JNK Mitogen-Activated Protein Kinases/antagonists & inhibitors , JNK Mitogen-Activated Protein Kinases/immunology , Lipopolysaccharides/antagonists & inhibitors , Lipopolysaccharides/pharmacology , MAP Kinase Kinase Kinases/antagonists & inhibitors , MAP Kinase Kinase Kinases/immunology , Macrophages/immunology , Phosphatidylinositol 3-Kinases/immunology , Ribosomal Protein S6 Kinases, 70-kDa/antagonists & inhibitors , Ribosomal Protein S6 Kinases, 70-kDa/immunology , TOR Serine-Threonine Kinases/antagonists & inhibitors , TOR Serine-Threonine Kinases/immunology , Toll-Like Receptor 4/agonists
4.
J Immunol ; 207(9): 2310-2324, 2021 11 01.
Article in English | MEDLINE | ID: covidwho-1436143

ABSTRACT

IFN-γ, a proinflammatory cytokine produced primarily by T cells and NK cells, activates macrophages and engages mechanisms to control pathogens. Although there is evidence of IFN-γ production by murine macrophages, IFN-γ production by normal human macrophages and their subsets remains unknown. Herein, we show that human M1 macrophages generated by IFN-γ and IL-12- and IL-18-stimulated monocyte-derived macrophages (M0) produce significant levels of IFN-γ. Further stimulation of IL-12/IL-18-primed macrophages or M1 macrophages with agonists for TLR-2, TLR-3, or TLR-4 significantly enhanced IFN-γ production in contrast to the similarly stimulated M0, M2a, M2b, and M2c macrophages. Similarly, M1 macrophages generated from COVID-19-infected patients' macrophages produced IFN-γ that was enhanced following LPS stimulation. The inhibition of M1 differentiation by Jak inhibitors reversed LPS-induced IFN-γ production, suggesting that differentiation with IFN-γ plays a key role in IFN-γ induction. We subsequently investigated the signaling pathway(s) responsible for TLR-4-induced IFN-γ production in M1 macrophages. Our results show that TLR-4-induced IFN-γ production is regulated by the ribosomal protein S6 kinase (p70S6K) through the activation of PI3K, the mammalian target of rapamycin complex 1/2 (mTORC1/2), and the JNK MAPK pathways. These results suggest that M1-derived IFN-γ may play a key role in inflammation that may be augmented following bacterial/viral infections. Moreover, blocking the mTORC1/2, PI3K, and JNK MAPKs in macrophages may be of potential translational significance in preventing macrophage-mediated inflammatory diseases.


Subject(s)
Interferon-gamma/biosynthesis , Macrophages/drug effects , Poly I-C/pharmacology , COVID-19/immunology , Humans , JNK Mitogen-Activated Protein Kinases/antagonists & inhibitors , JNK Mitogen-Activated Protein Kinases/immunology , Lipopolysaccharides/antagonists & inhibitors , Lipopolysaccharides/pharmacology , MAP Kinase Kinase Kinases/antagonists & inhibitors , MAP Kinase Kinase Kinases/immunology , Macrophages/immunology , Phosphatidylinositol 3-Kinases/immunology , Ribosomal Protein S6 Kinases, 70-kDa/antagonists & inhibitors , Ribosomal Protein S6 Kinases, 70-kDa/immunology , TOR Serine-Threonine Kinases/antagonists & inhibitors , TOR Serine-Threonine Kinases/immunology , Toll-Like Receptor 4/agonists
5.
J Neuroinflammation ; 18(1): 193, 2021 Sep 06.
Article in English | MEDLINE | ID: covidwho-1430430

ABSTRACT

BACKGROUND: The neuroimmune system is required for normal neural processes, including modulation of cognition, emotion, and adaptive behaviors. Aberrant neuroimmune activation is associated with dysregulation of memory and emotion, though the precise mechanisms at play are complex and highly context dependent. Sex differences in neuroimmune activation and function further complicate our understanding of its roles in cognitive and affective regulation. METHODS: Here, we characterized the physiological sickness and inflammatory response of the hippocampus following intracerebroventricular (ICV) administration of a synthetic viral mimic, polyinosinic:polycytidylic acid (poly I:C), in both male and female C57Bl/6N mice. RESULTS: We observed that poly I:C induced weight loss, fever, and elevations of cytokine and chemokines in the hippocampus of both sexes. Specifically, we found transient increases in gene expression and protein levels of IL-1α, IL-1ß, IL-4, IL-6, TNFα, CCL2, and CXCL10, where males showed a greater magnitude of response compared with females. Only males showed increased IFNα and IFNγ in response to poly I:C, whereas both males and females exhibited elevations of IFNß, demonstrating a specific sex difference in the anti-viral response in the hippocampus. CONCLUSION: Our data suggest that type I interferons are one potential node mediating sex-specific cytokine responses and neuroimmune effects on cognition. Together, these findings highlight the importance of using both males and females and analyzing a broad set of inflammatory markers in order to identify the precise, sex-specific roles for neuroimmune dysregulation in neurological diseases and disorders.


Subject(s)
Poly I-C , Sex Characteristics , Animals , Chemokines/metabolism , Cytokines/metabolism , Female , Hippocampus/metabolism , Male , Mice , Poly I-C/pharmacology
6.
Biochem Biophys Res Commun ; 565: 64-71, 2021 08 06.
Article in English | MEDLINE | ID: covidwho-1251023

ABSTRACT

Neutrophil extracellular traps (NETs) are extracellular webs of DNA, histones and granular contents that are released by neutrophils to control infections. However, NETs that is not properly regulated can propagate inflammation and thrombosis. It was recognized that viruses can induce NETs. As a synthetic analog of viral double-stranded (ds) RNA, polyinosinic-polycytidylic acid [poly(I:C)] is known to induce inflammation and thrombosis. However, whether and how poly(I:C) modulates NETs remains unclear. Here, we have demonstrated that poly(I:C) induced extracellular DNA traps in human neutrophils in a dose-dependent manner. Further, poly(I:C) or dsRNA virus elevated the levels of myeloperoxidase-DNA complexes and citrullinated histone H3, which are specific markers of NETs, in both neutrophil supernatants and mouse plasma. Interestingly, a potent peptidylarginine deiminase 4 (PAD4) inhibitor, BB-CL-Amidine (BB-CLA) or PAD4 knockdown effectively prevented poly(I:C)-induced NETs formation and release. In addition, BB-CLA abrogated poly(I:C)-triggered neutrophil activation and infiltration, and vascular permeability in lungs. BB-CLA also attenuated poly(I:C)-induced thrombocytopenia in circulation, fibrin deposition and thrombus formation in tissues. Taken together, these results suggest that viral mimetic poly(I:C) may induce NETs-dependent inflammation and thrombosis through PAD4, and that inhibiting PAD4 may become a good strategy to protect against viral infection-caused inflammation/thrombosis-related pathological conditions of diseases.


Subject(s)
Extracellular Traps/drug effects , Inflammation/metabolism , Neutrophils/drug effects , Poly I-C/pharmacology , Protein-Arginine Deiminase Type 4/metabolism , Thrombosis/metabolism , Amidines/pharmacology , Animals , Cells, Cultured , Chlorocebus aethiops , Humans , Inflammation/pathology , Male , Mice , Mice, Inbred C57BL , Neutrophil Activation/drug effects , Neutrophils/metabolism , Protein-Arginine Deiminase Type 4/antagonists & inhibitors , Thrombosis/pathology
8.
Kidney Blood Press Res ; 46(1): 74-83, 2021.
Article in English | MEDLINE | ID: covidwho-978847

ABSTRACT

INTRODUCTION: Various viruses including a novel coronavirus (SARS-CoV-2) can infect the kidney. When viruses invade the glomeruli from the bloodstream, glomerular endothelial cells (GECs) initiate the innate immune reactions. We investigated the expression of interferon (IFN)-induced protein with tetratricopeptide repeats (IFIT) 1/2/3, antiviral molecules, in human GECs treated with a toll-like receptor (TLR) 3 agonist. Role of IFIT1/2/3 in the expression of C-X-C motif chemokine ligand 10 (CXCL10) was also examined. METHODS: Human GECs were cultured and stimulated with polyinosinic-polycytidylic acid (poly IC), a synthetic TLR3 agonist. Real-time qPCR, Western blotting, and ELISA were used to examine the expression of IFIT1/2/3, IFN-ß, and CXCL10. RNA interference against IFN-ß or IFIT1/2/3 was also performed. RESULTS: Expression of IFIT1/2/3 and CXCL10 was induced by poly IC in GECs. The inductions were inhibited by RNA interfering of IFN-ß. Knockdown of IFIT1/2/3 decreased the CXCL10 expression. Knockdown of IFIT3 decreased the expression of IFIT1 and IFIT2 proteins. CONCLUSION: IFIT1/2/3 and CXCL10 were induced by poly IC via IFN-ß in GECs. IFIT1/2/3 may increase the expression of CXCL10 which induces lymphocyte chemotaxis and may inhibit the replication of infected viruses. These molecules may play a role in GEC innate immune reactions in response to viruses.


Subject(s)
Adaptor Proteins, Signal Transducing/biosynthesis , Apoptosis Regulatory Proteins/biosynthesis , Chemokine CXCL10/biosynthesis , Intracellular Signaling Peptides and Proteins/biosynthesis , Kidney Glomerulus/metabolism , RNA-Binding Proteins/biosynthesis , Toll-Like Receptor 3/agonists , Adaptor Proteins, Signal Transducing/genetics , Apoptosis Regulatory Proteins/genetics , Cells, Cultured , Chemokine CXCL10/genetics , Dose-Response Relationship, Drug , Endothelial Cells/drug effects , Endothelial Cells/metabolism , Gene Expression , Humans , Intracellular Signaling Peptides and Proteins/genetics , Kidney Glomerulus/cytology , Kidney Glomerulus/drug effects , Poly I-C/pharmacology , RNA-Binding Proteins/genetics , Toll-Like Receptor 3/metabolism
9.
Vet Microbiol ; 252: 108918, 2021 Jan.
Article in English | MEDLINE | ID: covidwho-909094

ABSTRACT

Porcine haemagglutinating encephalomyelitis virus (PHEV) is a member of coronavirus that causes acute infectious disease and high mortality in piglets. The transcription factor IRF3 is a central regulator of type I interferon (IFN) innate immune signalling. Here, we report that PHEV infection of RAW264.7 cells results in strong suppression of IFN-ß production in the early stage. A comparative analysis of the upstream effector of IFN-ß transcription demonstrated that deactivation of IRF3, but not p65 or ATF-2 proteins, is uniquely attributed to failure of early IFN-ß induction. Moreover, the RIG-I/MDA5/MAVS/TBK1-dependent protective response that regulates the IRF3 pathway is not disrupted by PHEV and works well underlying the deactivated IRF3-mediated IFN-ß inhibition. After challenge with poly(I:C), a synthetic analogue of dsRNA used to stimulate IFN-ß secretion in the TLR-controlled pathway, we show that PHEV and poly(I:C) regulate IFN-ß-induction via two different pathways. Collectively, our findings reveal that deactivation of IRF3 is a specific mechanism that contributes to termination of type I IFN signalling during early infection with PHEV independent of the conserved RIG-I/MAVS/MDA5/TBK1-mediated innate immune response.


Subject(s)
Betacoronavirus 1/immunology , Coronavirus Infections/veterinary , Interferon Regulatory Factor-3/genetics , Interferon-beta/immunology , Animals , Betacoronavirus 1/genetics , Coronavirus Infections/immunology , Coronavirus Infections/virology , Immunity, Innate , Interferon Regulatory Factor-3/immunology , Mice , Poly I-C/pharmacology , RAW 264.7 Cells , Signal Transduction/immunology
SELECTION OF CITATIONS
SEARCH DETAIL