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1.
PLoS Pathog ; 17(9): e1009878, 2021 09.
Artigo em Inglês | MEDLINE | ID: mdl-34473805

RESUMO

SARS-CoV-2 fine-tunes the interferon (IFN)-induced antiviral responses, which play a key role in preventing coronavirus disease 2019 (COVID-19) progression. Indeed, critically ill patients show an impaired type I IFN response accompanied by elevated inflammatory cytokine and chemokine levels, responsible for cell and tissue damage and associated multi-organ failure. Here, the early interaction between SARS-CoV-2 and immune cells was investigated by interrogating an in vitro human peripheral blood mononuclear cell (PBMC)-based experimental model. We found that, even in absence of a productive viral replication, the virus mediates a vigorous TLR7/8-dependent production of both type I and III IFNs and inflammatory cytokines and chemokines, known to contribute to the cytokine storm observed in COVID-19. Interestingly, we observed how virus-induced type I IFN secreted by PBMC enhances anti-viral response in infected lung epithelial cells, thus, inhibiting viral replication. This type I IFN was released by plasmacytoid dendritic cells (pDC) via an ACE-2-indipendent but Neuropilin-1-dependent mechanism. Viral sensing regulates pDC phenotype by inducing cell surface expression of PD-L1 marker, a feature of type I IFN producing cells. Coherently to what observed in vitro, asymptomatic SARS-CoV-2 infected subjects displayed a similar pDC phenotype associated to a very high serum type I IFN level and induction of anti-viral IFN-stimulated genes in PBMC. Conversely, hospitalized patients with severe COVID-19 display very low frequency of circulating pDC with an inflammatory phenotype and high levels of chemokines and pro-inflammatory cytokines in serum. This study further shed light on the early events resulting from the interaction between SARS-CoV-2 and immune cells occurring in vitro and confirmed ex vivo. These observations can improve our understanding on the contribution of pDC/type I IFN axis in the regulation of the anti-viral state in asymptomatic and severe COVID-19 patients.


Assuntos
COVID-19/imunologia , Células Dendríticas/classificação , Interferon Tipo I/metabolismo , SARS-CoV-2/imunologia , Adulto , Idoso de 80 Anos ou mais , Infecções Assintomáticas , Linhagem Celular Tumoral , Células Dendríticas/imunologia , Células Dendríticas/virologia , Células Epiteliais/citologia , Feminino , Hospitalização , Humanos , Interferon Tipo I/imunologia , Pulmão/citologia , Masculino , Pessoa de Meia-Idade , Neuropilina-1/metabolismo , Fenótipo , Índice de Gravidade de Doença , Receptor 7 Toll-Like/metabolismo
2.
Cancer Sci ; 112(11): 4490-4500, 2021 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-34537997

RESUMO

Various cancer vaccines have been developed to generate and amplify antigen-specific T cell responses against malignancy. Among them, in situ vaccination is one of the most practical types as it can trigger immune responses without previous antigen identification. Here we reported a novel in situ vaccine by intratumoral injection of imiquimod and OX40 agonist. In mice bearing hepatic carcinoma, both the injected tumor and the noninjected tumor in the distant lesion of the same mice were suppressed after vaccination. Further studies found that this in situ vaccine triggered systemic tumor-specific responses, with one-fold increase of effector memory T cells properties and stronger toxicity of lymphocytes in spleen. Besides, we found that imiquimod upregulated the expression of OX40 on CD4+ T cells and thus enhanced the effectiveness of OX40 agonist. Five immune-positive-related pathways were activated after vaccination. This in situ vaccine caused little harm to normal organs and provided long-term protection against the same syngeneic tumor rechallenge. Due to its effectiveness, feasibility and safety, this strategy could potentially be applied to various types of late-stage solid tumors and worthy of further clinical research.


Assuntos
Antineoplásicos/uso terapêutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Vacinas Anticâncer/uso terapêutico , Imiquimode/uso terapêutico , Neoplasias Hepáticas/tratamento farmacológico , Receptores OX40/agonistas , Animais , Antineoplásicos/administração & dosagem , Antineoplásicos/efeitos adversos , Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Linfócitos T CD4-Positivos/efeitos dos fármacos , Linfócitos T CD4-Positivos/metabolismo , Vacinas Anticâncer/administração & dosagem , Vacinas Anticâncer/efeitos adversos , Feminino , Imiquimode/administração & dosagem , Imiquimode/efeitos adversos , Memória Imunológica/efeitos dos fármacos , Imunoterapia , Injeções Intralesionais/métodos , Neoplasias Hepáticas/imunologia , Glicoproteínas de Membrana/metabolismo , Camundongos , Receptores OX40/metabolismo , Linfócitos T/efeitos dos fármacos , Receptor 7 Toll-Like/metabolismo , Microambiente Tumoral/efeitos dos fármacos , Microambiente Tumoral/imunologia , Vacinação/métodos
3.
Dermatology ; 237(6): 847-856, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34511591

RESUMO

BACKGROUND: The innate immune system is recognized as an essential aspect of COVID-19 pathogenesis. Toll-like receptors (TLRs) are important in inducing antiviral response, triggering downstream production of interferons (IFNs). Certain loss-of-function variants in TLR7 are associated with increased COVID-19 disease severity, and imiquimod (ImiQ) is known to have immunomodulating effects as an agonist of TLR7. Given that topical imiquimod (topImiQ) is indicated for various dermatologic conditions, it is necessary for dermatologists to understand the interplay between innate immunity mechanisms and the potential role of ImiQ in COVID-19, with a particular focus on TLR7. SUMMARY: Our objective was to survey recent peer-reviewed scientific literature in the PubMed database, examine relevant evidence, and elucidate the relationships between IFNs, TLR7, the innate immune system, and topImiQ in the context of COVID-19. Despite limited studies on this topic, current evidence supports the critical role of TLRs in mounting a strong immune response against COVID-19. Of particular interest to dermatologists, topImiQ can result in systemic upregulation of the immune system via activation of TLR7. Key Message: Given the role of TLR7 in the systemic activation of the immune system, ImiQ, as a ligand of the TLR7 receptor, may have potential therapeutic benefit as a topical immunomodulatory treatment for COVID-19.


Assuntos
COVID-19/prevenção & controle , Imiquimode/administração & dosagem , Imunidade Inata , Interferons/administração & dosagem , SARS-CoV-2 , Receptor 7 Toll-Like/metabolismo , Regulação para Cima/efeitos dos fármacos , Adjuvantes Imunológicos/farmacologia , Administração Tópica , Animais , Antivirais/administração & dosagem , COVID-19/epidemiologia , COVID-19/metabolismo , Humanos
4.
J Immunol ; 207(5): 1448-1455, 2021 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-34362834

RESUMO

Identification of the receptors involved in innate immune recognition of Staphylococcus aureus, a major cause of morbidity and mortality in humans, is essential to develop alternative strategies to treat infections caused by antibiotic-resistant strains. In the current study, we examine the role of endosomal TLRs, which sense the presence of prokaryotic-type nucleic acids, in anti-staphylococcal host defenses using infection models involving genetically defective mice. Single deficiencies in TLR7, 9, or 13 resulted in mild or no decrease in host defenses. However, the simultaneous absence of TLR7, 9, and 13 resulted in markedly increased susceptibility to cutaneous and systemic S. aureus infection concomitantly with decreased production of proinflammatory chemokines and cytokines, neutrophil recruitment to infection sites, and reduced production of reactive oxygen species. This phenotype was significantly more severe than that of mice lacking TLR2, which senses the presence of staphylococcal lipoproteins. Notably, the combined absence of TLR7, 9, and 13 resulted in complete abrogation of IL-12 p70 and IFN-ß responses to staphylococcal stimulation in macrophages. Taken together, our data highlight the presence of a highly integrated endosomal detection system, whereby TLR7, 9, and 13 cooperate in sensing the presence of staphylococcal nucleic acids. We demonstrate that the combined absence of these receptors cannot be compensated for by cell surface-associated TLRs, such as TLR2, or cytosolic receptors. These data may be useful to devise strategies aimed at stimulating innate immune receptors to treat S. aureus infections.


Assuntos
Endossomos/metabolismo , Glicoproteínas de Membrana/metabolismo , Neutrófilos/imunologia , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/fisiologia , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Receptores Toll-Like/metabolismo , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Feminino , Humanos , Imunidade Inata , Mediadores da Inflamação/metabolismo , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor 7 Toll-Like/genética , Receptor Toll-Like 9/genética , Receptores Toll-Like/genética
5.
Sci Rep ; 11(1): 16348, 2021 08 11.
Artigo em Inglês | MEDLINE | ID: mdl-34381063

RESUMO

Systemic lupus erythematosus (SLE) is a chronic autoimmune disease that affects multiple organs. Recent studies suggest relevance between cysteine protease cathepsin S (CTSS) expression and SLE. To investigate the mechanism of CTSS in SLE, CTSS-overexpressing transgenic (TG) mice were generated, and induced lupus-like symptoms. Eight months later, the TG mice spontaneously developed typical SLE symptoms regardless of the inducement. Furthermore, we observed increased toll-like receptor 7 (TLR7) expression with increased monocyte and neutrophil populations in the TG mice. In conclusion, overexpression of CTSS in mice influences TLR7 expression, autoantibodies and IFN-α, which leads to an autoimmune reaction and exacerbates lupus-like symptoms.


Assuntos
Catepsinas/metabolismo , Interferon-alfa/metabolismo , Glicoproteínas de Membrana/metabolismo , Receptor 7 Toll-Like/metabolismo , Regulação para Cima/fisiologia , Animais , Autoanticorpos , Feminino , Humanos , Lúpus Eritematoso Sistêmico/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos ICR , Camundongos Transgênicos , Monócitos/metabolismo , Neutrófilos/metabolismo
6.
Front Immunol ; 12: 686060, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34211474

RESUMO

Toll-like receptor (TLR) signaling is critical for defense against pathogenic infection, as well as for modulating tissue development. Activation of different TLRs triggers common inflammatory responses such as cytokine induction. Here, we reveal differential impacts of TLR3 and TLR7 signaling on transcriptomic profiles in bone marrow-derived macrophages (BMDMs). Apart from self-regulation, TLR3, but not TLR7, induced expression of other TLRs, suggesting that TLR3 activation globally enhances innate immunity. Moreover, we observed diverse influences of TLR3 and TLR7 signaling on genes involved in methylation, caspase and autophagy pathways. We compared endogenous TLR3 and TLR7 by using CRISPR/Cas9 technology to knock in a dual Myc-HA tag at the 3' ends of mouse Tlr3 and Tlr7. Using anti-HA antibodies to detect endogenous tagged TLR3 and TLR7, we found that both TLRs display differential tissue expression and posttranslational modifications. C-terminal tagging did not impair TLR3 activity. However, it disrupted the interaction between TLR7 and myeloid differentiation primary response 88 (MYD88), the Tir domain-containing adaptor of TLR7, which blocked its downstream signaling necessary to trigger cytokine and chemokine expression. Our study demonstrates different properties for TLR3 and TLR7, and also provides useful mouse models for further investigation of these two RNA-sensing TLRs.


Assuntos
Epitopos/metabolismo , Macrófagos/metabolismo , Glicoproteínas de Membrana/fisiologia , Neurônios/metabolismo , Receptor 3 Toll-Like/fisiologia , Receptor 7 Toll-Like/fisiologia , Animais , Quimiocinas/metabolismo , Citocinas/metabolismo , Epitopos/imunologia , Feminino , Perfilação da Expressão Gênica , Imunidade Inata , Macrófagos/imunologia , Masculino , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Fator 88 de Diferenciação Mieloide/metabolismo , Fator 88 de Diferenciação Mieloide/fisiologia , Transdução de Sinais , Receptor 3 Toll-Like/genética , Receptor 3 Toll-Like/metabolismo , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo
7.
Sci Rep ; 11(1): 15231, 2021 07 27.
Artigo em Inglês | MEDLINE | ID: mdl-34315922

RESUMO

Human Vγ9Vδ2 T cells recognize pyrophosphates produced by microbes and transformed cells and play a role in anti-infective immunity and tumor surveillance. Toll-like receptors (TLR) are pattern recognition receptors in innate immune cells which sense microbial structures including nucleic acids. Given that γδ T cells are in clinical development for application in cellular cancer immunotherapy and TLR ligands have potent adjuvant activity, we investigated the co-stimulatory role of selected TLR ligands in γδ T-cell activation. Here we have used recently described RNA ligands for TLR7 and TLR8 together with Vγ9Vδ2 T-cell specific pyrophosphate antigens to analyze the rapid cytokine induction in Vδ2 T cells as well as the accessory cell requirements. While TLR8- as well as TLR7/8-specific RNA did not induce IFN-γ in Vδ2 T cells on their own, they provided strong co-stimulation for Vδ2 T cells within peripheral blood mononuclear cells in the presence of additional T-cell receptor activation. In contrast, TLR7 ligands were ineffective. Purified γδ T cells did not directly respond to TLR8 co-stimulation but required the presence of monocytes. Further experiments revealed a critical role of IL-1ß and IL-18, and to a slightly lesser extent of IL-12p70, in the co-stimulation of Vδ2 T cells by TLR8 and TLR7/8 RNA ligands. Results of intracellular cytokine expression were validated by ELISA analysis of cytokines in cell culture supernatants. The cell context-dependent adjuvant activity of TLR8 and TLR7/8 RNA ligands described here might be important for the future optimization of γδ T-cell based cancer immunotherapy.


Assuntos
Citocinas/biossíntese , Linfócitos Intraepiteliais/metabolismo , Monócitos/metabolismo , RNA/metabolismo , Receptor 8 Toll-Like/metabolismo , Imunidade Adaptativa , Células Cultivadas , Citocinas/metabolismo , Humanos , Imunidade Inata , Linfócitos Intraepiteliais/imunologia , Ligantes , Receptor 7 Toll-Like/metabolismo
8.
Oncology ; 99(9): 589-600, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34139707

RESUMO

BACKGROUND: Colorectal cancer (CRC), the third most common cancer globally, caused 881,000 cancer deaths in 2018. Toll-like receptors (TLRs), the primary sensors of pathogen-associated molecular patterns and damage-associated molecular patterns, activate innate and adaptive immune systems and participate in the development of an inflammatory tumor microenvironment. We aimed to explore the prognostic value of TLR3, TLR5, TLR7, and TLR9 tissue expressions in CRC patients. METHODS: Using immunohistochemistry, we analyzed tissue microarray samples from 825 CRC patients who underwent surgery between 1982 and 2002 at the Department of Surgery, Helsinki University Hospital, Finland. After analyzing a pilot series of 205 tissue samples, we included only TLR5 and TLR7 in the remainder of the patient series. We evaluated the associations between TLR5 and TLR7 tissue expressions, clinicopathologic variables, and survival. Using the Kaplan-Meier method, we generated survival curves, determining significance using the log-rank test. Univariate and multivariate survival analyses relied on the Cox proportional hazards model. RESULTS: The 5-year disease-specific survival was 55.9% among TLR5-negative (95% confidence interval [CI] 50.6-61.2%) and 61.9% (95% CI 56.6-67.2%; p = 0.011, log-rank test) among TLR5-positive patients. In the Cox multivariate survival analysis adjusted for age, sex, stage, location, and grade, positive TLR5 immunoexpression (hazard ratio [HR] 0.74; 95% CI 0.59-0.92; p = 0.007) served as an independent positive prognostic factor. TLR7 immunoexpression exhibited no prognostic value in the survival analysis across the entire cohort (HR 0.97; 95% CI 0.78-1.20; p = 0.754) nor in subgroup analyses. CONCLUSIONS: We show for the first time that a high TLR5 tumor tissue expression associates with a better prognosis in CRC patients.


Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Colorretais/metabolismo , Receptor 5 Toll-Like/metabolismo , Idoso , Neoplasias Colorretais/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Prognóstico , Estudos Retrospectivos , Análise de Sobrevida , Receptor 7 Toll-Like/metabolismo
9.
Cells ; 10(5)2021 05 12.
Artigo em Inglês | MEDLINE | ID: mdl-34066164

RESUMO

The B cell scaffold protein with ankyrin repeats (BANK1) is expressed primarily in B cells and with multiple but discrete roles in B cell signaling, including B cell receptor signaling, CD40-related signaling, and Toll-like receptor (TLR) signaling. The gene for BANK1, located in chromosome 4, has been found to contain genetic variants that are associated with several autoimmune diseases and also other complex phenotypes, in particular, with systemic lupus erythematosus. Common genetic variants are associated with changes in BANK1 expression in B cells, while rare variants modify their capacity to bind efferent effectors during signaling. A BANK1-deficient model has shown the importance of BANK1 during TLR7 and TLR9 signaling and has confirmed its role in the disease. Still, much needs to be done to fully understand the function of BANK1, but the main conclusion is that it may be the link between different signaling functions within the B cells and they may act to synergize the various pathways within a cell. With this review, we hope to enhance the interest in this molecule.


Assuntos
Proteínas Adaptadoras de Transdução de Sinal/genética , Doenças Autoimunes/genética , Linfócitos B/metabolismo , Proteínas de Membrana/genética , Polimorfismo Genético , Proteínas Adaptadoras de Transdução de Sinal/fisiologia , Alelos , Animais , Doenças Autoimunes/metabolismo , Autoimunidade , Antígenos CD40/metabolismo , Células Dendríticas , Variação Genética , Humanos , Proteínas de Membrana/fisiologia , Camundongos , Fenótipo , Risco , Transdução de Sinais , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/metabolismo , Quinases da Família src/genética , Quinases da Família src/metabolismo
10.
J Clin Exp Hematop ; 61(2): 85-92, 2021 Jun 05.
Artigo em Inglês | MEDLINE | ID: mdl-33994431

RESUMO

Necrotizing lymphadenitis (NEL) is a self-limited systemic disease exhibiting characteristic clinical features. The pathogenesis of the disease remains unclear, but it may be associated with viral infection. In lymph nodes affected by this disease, innumerable plasmacytoid dendritic cells produce interferon-α when triggered by certain viral stimuli. IFN-α presents antigens causing the transformation of CD8+ cells into immunoblasts and apoptosis of CD4+ cells. From the perspective of innate immunity, UNC93B1, an endoplasmic reticulum (ER)-resident protein, associates more strongly with TLR9 than TLR7. Homeostasis is maintained under normal conditions. However, in NEL, TLR 7 was observed more than TLR 9, possibly because mutant type UNC93B1 associates more tightly with TLR7. The inhibitory effects against TLR7 by TLR9 were reported to disappear. It is likely that more TLR7 than TLR9 is transported from the ER to endolysosomes. In conclusion, overexpression of TLR7, an innate immune sensor of microbial single-stranded RNA, is inferred. Consequently, NEL may be induced.


Assuntos
Células Dendríticas/patologia , Linfadenite/patologia , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Adolescente , Adulto , Criança , Pré-Escolar , Células Dendríticas/metabolismo , Feminino , Humanos , Linfadenite/metabolismo , Masculino , Pessoa de Meia-Idade , Transporte Proteico , Receptor 7 Toll-Like/análise , Receptor Toll-Like 9/análise , Adulto Jovem
11.
J Med Chem ; 64(11): 7507-7532, 2021 06 10.
Artigo em Inglês | MEDLINE | ID: mdl-34048243

RESUMO

Activation of the toll-like receptors 7 and 8 has emerged as a promising strategy for cancer immunotherapy. Herein, we report the design and synthesis of a series of pyrido[3,2-d]pyrimidine-based toll-like receptor 7/8 dual agonists that exhibited potent and near-equivalent agonistic activities toward TLR7 and TLR8. In vitro, compounds 24e and 25a significantly induced the secretion of IFN-α, IFN-γ, TNF-α, IL-1ß, IL-12p40, and IP-10 in human peripheral blood mononuclear cell assays. In vivo, compounds 24e, 24m, and 25a significantly suppressed tumor growth in CT26 tumor-bearing mice by remodeling the tumor microenvironment. Additionally, compounds 24e, 24m, and 25a markedly improved the antitumor activity of PD-1/PD-L1 blockade. In particular, compound 24e combined with the anti-PD-L1 antibody led to complete tumor regression. These results demonstrated that TLR7/8 agonists (24e, 24m, and 25a) held great potential as single agents or in combination with PD-1/PD-L1 blockade for cancer immunotherapy.


Assuntos
Antineoplásicos/química , Desenho de Fármacos , Piridinas/química , Pirimidinas/química , Receptor 7 Toll-Like/agonistas , Receptor 8 Toll-Like/agonistas , Animais , Anticorpos Monoclonais/uso terapêutico , Antineoplásicos/metabolismo , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Sítios de Ligação , Humanos , Imunoterapia , Interleucina-1beta/metabolismo , Leucócitos Mononucleares/citologia , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Simulação de Acoplamento Molecular , Neoplasias/patologia , Neoplasias/terapia , Piridinas/metabolismo , Piridinas/farmacologia , Piridinas/uso terapêutico , Pirimidinas/metabolismo , Pirimidinas/farmacologia , Pirimidinas/uso terapêutico , Relação Estrutura-Atividade , Receptor 7 Toll-Like/metabolismo , Receptor 8 Toll-Like/metabolismo , Microambiente Tumoral , Fator de Necrose Tumoral alfa/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
12.
Front Immunol ; 12: 680068, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34025679

RESUMO

Toll-like receptors (TLRs) play critical roles in regulating the abnormal activation of the immune cells resulting in the pathogenesis of inflammation and autoimmune diseases. Pyruvate kinase M2 (PKM2), which governs the last step of glycolysis, is involved in multiple cellular processes and pathological conditions. However, little is known about the involvement of PKM2 in regulating TLR-mediated inflammation and autoimmunity. Herein, we investigated the role of PKM2 in the activation of the TLR pathways and the pathogenesis of inflammation and autoimmune diseases. The activation of TLR4, TLR7 and TLR9 pathways was found to induce the up-regulation of PKM2 expression in macrophages, dendritic cells (DCs) and B cells. The over-expression of PKM2 promotes the activation of TLR4, TLR7 and TLR9 pathways while interference with the PKM2 expression or the addition of the PKM2 inhibitor (PKM-IN) markedly inhibited the activation of TLR4, TLR7 and TLR9 pathways. Mechanistically, PKM2 augmented the activation of TLR4, TLR7 and TLR9 pathways by promoting the activation of the proline-rich tyrosine kinase 2 (Pyk2). Intriguingly, the PKM2 inhibitor PKM2-IN significantly protected the mice from the endotoxic shock mediated by the TLR4-agonist LPS. Additionally, it alleviated the progression in the TLR7-agonist imiquimod-mediated lupus mice and spontaneous lupus MRL/lpr mice. Moreover, PKM2 expression was highly elevated in the monocytes, DCs and B cells from systemic lupus erythematous (SLE) patients compared with those from the healthy donors. Besides, the PKM2 expression level was positively correlated with the degree of activation of these immune cells. In summary, PKM2 contributed to TLR-mediated inflammation and autoimmunity and can be a valuable target to control inflammation and autoimmunity.


Assuntos
Autoimunidade , Proteínas de Transporte/metabolismo , Quinase 2 de Adesão Focal/metabolismo , Inflamação/etiologia , Inflamação/metabolismo , Proteínas de Membrana/metabolismo , Hormônios Tireóideos/metabolismo , Receptores Toll-Like/metabolismo , Animais , Doenças Autoimunes/etiologia , Doenças Autoimunes/metabolismo , Linfócitos B/imunologia , Linfócitos B/metabolismo , Biomarcadores , Proteínas de Transporte/antagonistas & inibidores , Sobrevivência Celular , Modelos Animais de Doenças , Suscetibilidade a Doenças , Feminino , Inflamação/diagnóstico , Lúpus Eritematoso Sistêmico/imunologia , Lúpus Eritematoso Sistêmico/metabolismo , Lúpus Eritematoso Sistêmico/patologia , Ativação Linfocitária/imunologia , Macrófagos/imunologia , Macrófagos/metabolismo , Proteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos MRL lpr , Modelos Biológicos , Inibidores de Proteínas Quinases/farmacologia , Transdução de Sinais/efeitos dos fármacos , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo
13.
J Innate Immun ; 13(6): 345-358, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34058746

RESUMO

Regulation of proinflammatory cytokine expression is critical in the face of single-stranded RNA (ssRNA) virus infections. Many viruses, including coronavirus and influenza virus, wreak havoc on the control of cytokine expression, leading to the formation of detrimental cytokine storms. Understanding the regulation and interplay between inflammatory cytokines is critical to the identification of targets involved in controlling the induction of cytokine expression. In this study, we focused on how the antiviral cytokine interleukin-27 (IL-27) regulates signal transduction downstream of Toll-like receptor 7 (TLR7) and TLR8 ligation, which recognize endosomal single-stranded RNA. Given that IL-27 alters bacterial-sensing TLR expression on myeloid cells and can inhibit replication of single-stranded RNA viruses, we investigated whether IL-27 affects expression and function of TLR7 and TLR8. Analysis of IL-27-treated THP-1 monocytic cells and THP-1-derived macrophages revealed changes in mRNA and protein expression of TLR7 and TLR8. Although treatment with IL-27 enhanced TLR7 expression, only TLR8-mediated cytokine secretion was amplified. Furthermore, we demonstrated that imiquimod, a TLR7 agonist, inhibited cytokine and chemokine production induced by a TLR8 agonist, TL8-506. Delineating the immunomodulatory role of IL-27 on TLR7 and TLR8 responses provides insight into how myeloid cell TLR-mediated responses are regulated during virus infection.


Assuntos
Interleucina-27/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Receptor 7 Toll-Like/imunologia , Receptor 8 Toll-Like/imunologia , Citocinas/imunologia , Humanos , Imunomodulação , Inflamação , RNA Mensageiro/metabolismo , Transdução de Sinais , Células THP-1 , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo , Receptor 8 Toll-Like/genética , Receptor 8 Toll-Like/metabolismo
14.
Front Immunol ; 12: 635018, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33936047

RESUMO

Objective: Bacterial and viral infectious triggers are linked to spondyloarthritis (SpA) including psoriatic arthritis (PsA) development, likely via dendritic cell activation. We investigated spinal entheseal plasmacytoid dendritic cells (pDCs) toll-like receptor (TLR)-7 and 9 activation and therapeutic modulation, including JAK inhibition. We also investigated if COVID-19 infection, a potent TLR-7 stimulator triggered PsA flares. Methods: Normal entheseal pDCs were characterized and stimulated with imiquimod and CpG oligodeoxynucleotides (ODN) to evaluate TNF and IFNα production. NanoString gene expression assay of total pDCs RNA was performed pre- and post- ODN stimulation. Pharmacological inhibition of induced IFNα protein was performed with Tofacitinib and PDE4 inhibition. The impact of SARS-CoV2 viral infection on PsA flares was evaluated. Results: CD45+HLA-DR+CD123+CD303+CD11c- entheseal pDCs were more numerous than blood pDCs (1.9 ± 0.8% vs 0.2 ± 0.07% of CD45+ cells, p=0.008) and showed inducible IFNα and TNF protein following ODN/imiquimod stimulation and were the sole entheseal IFNα producers. NanoString data identified 11 significantly upregulated differentially expressed genes (DEGs) including TNF in stimulated pDCs. Canonical pathway analysis revealed activation of dendritic cell maturation, NF-κB signaling, toll-like receptor signaling and JAK/STAT signaling pathways following ODN stimulation. Both tofacitinib and PDE4i strongly attenuated ODN induced IFNα. DAPSA scores elevations occurred in 18 PsA cases with SARS-CoV2 infection (9.7 ± 4 pre-infection and 35.3 ± 7.5 during infection). Conclusion: Entheseal pDCs link microbes to TNF/IFNα production. SARS-CoV-2 infection is associated with PsA Flares and JAK inhibition suppressed activated entheseal plasmacytoid dendritic Type-1 interferon responses as pointers towards a novel mechanism of PsA and SpA-related arthropathy.


Assuntos
Artrite Psoriásica/complicações , COVID-19/complicações , Células Dendríticas/metabolismo , Interferon-alfa/metabolismo , Janus Quinases/antagonistas & inibidores , Adjuvantes Imunológicos/farmacologia , Adulto , Idoso , COVID-19/genética , COVID-19/metabolismo , Biologia Computacional , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Células Dendríticas/efeitos dos fármacos , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/genética , Humanos , Imiquimode/farmacologia , Janus Quinases/metabolismo , Masculino , Pessoa de Meia-Idade , NF-kappa B/metabolismo , Oligonucleotídeos/farmacologia , Inibidores da Fosfodiesterase 4/farmacologia , Piperidinas/farmacologia , Inibidores de Proteínas Quinases/farmacologia , Pirimidinas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Transcriptoma , Fator de Necrose Tumoral alfa/metabolismo
15.
Front Immunol ; 12: 636427, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33897689

RESUMO

Macrophage activation and osteoclastogenesis are hallmarks of inflammatory osteolysis and may be targeted by the local application of liquid platelet-rich fibrin (PRF). Liquid PRF is produced by a hard spin of blood in the absence of clot activators and anticoagulants, thereby generating an upper platelet-poor plasma (PPP) layer, a cell-rich buffy coat layer (BC; termed concentrated-PRF or C-PRF), and the remaining red clot (RC) layer. Heating PPP has been shown to generate an albumin gel (Alb-gel) that when mixed back with C-PRF generates Alb-PRF having extended working properties when implanted in vivo. Evidence has demonstrated that traditional solid PRF holds a potent anti-inflammatory capacity and reduces osteoclastogenesis. Whether liquid PRF is capable of also suppressing an inflammatory response and the formation of osteoclasts remains open. In the present study, RAW 264.7 and primary macrophages were exposed to lipopolysaccharides (LPS), lactoferrin, and agonists of Toll-like receptors (TLR3 and TLR7) in the presence or absence of lysates prepared by freeze-thawing of liquid PPP, BC, Alb-gel, and RC. For osteoclastogenesis, primary macrophages were exposed to receptor activator of nuclear factor kappa B ligand (RANKL), macrophage colony-stimulating factor (M-CSF), and human transforming growth factor-ß1 (TGF-ß1) in the presence or absence of PPP, BC, Alb-gel, RC lysates and hemoglobin. We show here that it is mainly the lysates prepared from PPP and BC that consistently reduced the agonist-induced expression of interleukin 6 (IL6) and cyclooxygenase-2 (COX2) in macrophages, as determined by RT-PCR and immunoassay. With respect to osteoclastogenesis, lysates from PPP and BC but also from RC, similar to hemoglobin, reduced the expression of osteoclast marker genes tartrate-resistant acid phosphatase (TRAP) and cathepsin K, as well as TRAP histochemical staining. These findings suggest that liquid PRF holds a potent in vitro heat-sensitive anti-inflammatory activity in macrophages that goes along with an inhibition of osteoclastogenesis.


Assuntos
Inflamação/prevenção & controle , Ativação de Macrófagos , Macrófagos/metabolismo , Osteoclastos/metabolismo , Osteogênese , Plasma Rico em Plaquetas/metabolismo , Animais , Imiquimode/farmacologia , Inflamação/sangue , Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Lactoferrina/farmacologia , Lipopolissacarídeos/farmacologia , Ativação de Macrófagos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Glicoproteínas de Membrana/agonistas , Glicoproteínas de Membrana/metabolismo , Camundongos , Osteoclastos/efeitos dos fármacos , Osteoclastos/imunologia , Osteogênese/efeitos dos fármacos , Fenótipo , Poli I-C/farmacologia , Células RAW 264.7 , Receptor 3 Toll-Like/agonistas , Receptor 3 Toll-Like/metabolismo , Receptor 7 Toll-Like/agonistas , Receptor 7 Toll-Like/metabolismo
16.
Mol Immunol ; 134: 183-191, 2021 06.
Artigo em Inglês | MEDLINE | ID: mdl-33812250

RESUMO

Interferon regulatory factor 7 (IRF7) is a crucial regulator of type I interferons (IFNs) against pathogen infections and plays a significant role in the endosomal Toll-like receptor signaling (namely, TLR7 and TLR9) in plasmacytoid dendritic cells (pDCs). In this study, we identify MEKK3, one of the MAP3K kinase, as a potent stimulator of IRF7 upon cellular activation of the TLR7/9 signaling pathways to induce various type I IFNs. The knockdown of MEKK3 in vivo substantially impairs type I IFN induction and increases susceptibility to HSV-1 infection in mice. Overexpression of MEKK3 significantly activates IRF7 to trigger strong induction of type I IFNs, while cells deficient in MEKK3 expression show abrogated innate immune responses to TLR7/TLR9 ligands stimulation. We confirmed that the IFNs' induction is due to a MEKK3 and IRF7 interaction; it leads to the phosphorylation of IRF7 at multiple sites. Moreover, endogenous MEKK3 can bind and phosphorylate IRF7 after TLR9 activation by its specific ligand CpG DNA. It is the first time to report the role of MEKK3 on type I IFN, which indicates crosstalk between MAP3K activation and type I IFNs' induction in the endosomal Toll-like receptor pathways.


Assuntos
Fator Regulador 7 de Interferon/metabolismo , Interferon Tipo I/biossíntese , MAP Quinase Quinase Quinase 3/metabolismo , Receptor 7 Toll-Like/metabolismo , Receptor Toll-Like 9/metabolismo , Animais , Linhagem Celular , Feminino , Humanos , Fator Regulador 7 de Interferon/imunologia , Interferon Tipo I/imunologia , MAP Quinase Quinase Quinase 3/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Transdução de Sinais/imunologia , Receptor 7 Toll-Like/imunologia , Receptor Toll-Like 9/imunologia
17.
J Ethnopharmacol ; 275: 114063, 2021 Jul 15.
Artigo em Inglês | MEDLINE | ID: mdl-33813013

RESUMO

ETHNOPHARMACOLOGICAL RELEVANCE: Fufang-Yinhua-Jiedu Granules (FFYH) optimized from a Yin-Qiao-San, as traditional Chinese medicine (TCM), was used to treat influenza and upper respiratory tract infection and was recommended for the prevention and treatment of SARS in 2003 and current COVID-19 in Anhui Province in 2020. AIM OF STUDY: In the clinical studies, FFYH was very effective for the treatment of influenza, but the mechanism of action against influenza A virus remains unclear. In the present study, we investigated the antiviral effect of FFYH against influenza A virus in vitro and vivo. Moreover, the potential mechanism of FFYH against influenza A virus in vivo was investigated for the first time. MATERIALS AND METHODS: CPE inhibition assay and HA assay were used to evaluate the in vitro antiviral effects of FFYH against influenza A virus H1N1, H3N2, H5N1, H7N9 and H9N2. Mice were used to evaluate the antiviral effect of FFYH in vivo with ribavirin and lianhuaqingwen as positive controls. RT-PCR was used to quantify the mRNA transcription of TNF-α, IL-6, IFN-γ, IP10, and IL-1ß mRNA. ELISA was used to examine the expression of inflammatory factors such as TNF-α, IL-6, IFN-γ, IP10, and IL-1ß in sera. The blood parameters were analyzed with auto hematology analyzer. Moreover, the potential mechanism of FFYH against influenza A virus in vivo was also investigated. RESULTS: FFYH showed a broad-spectrum of antiviral activity against H1N1, H3N2, H5N1, H7N9, and H9N2 influenza A viruses. Furthermore, FFYH dose-dependently increased the survival rate, significantly prolonged the median survival time of mice, and markedly reduced lung injury caused by influenza A virus. Also, FFYH significantly improve the sick signs, food taken, weight loss, blood parameters, lung index, and lung pathological changes. Moreover, FFYH could markedly inhibit the inflammatory cytokine expression of TNF-α, IL-6, IFN-γ, IP10, IL-10, and IL-1ß mRNA or protein via inhibition of the TLR7/MyD88/NF-κB signaling pathway in vivo. CONCLUSION: FFYH not only showed a broad-spectrum of anti-influenza virus activity in vitro, but also exhibited a significant protective effect against lethal influenza virus infection in vivo. Furthermore, our results indicated that the in vivo antiviral effect of FFYH against influenza virus may be attributed to suppressing the expression of inflammatory cytokines via regulating the TLR7/MyD88/NF-κB signaling pathway. These findings provide evidence for the clinical treatment of influenza A virus infection with FFYH.


Assuntos
Anti-Inflamatórios/farmacologia , Antivirais/farmacologia , Medicamentos de Ervas Chinesas/farmacologia , Vírus da Influenza A/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Glicoproteínas de Membrana/metabolismo , Fator 88 de Diferenciação Mieloide/metabolismo , Infecções por Orthomyxoviridae/tratamento farmacológico , Receptor 7 Toll-Like/metabolismo , Células A549 , Animais , Citocinas/genética , Citocinas/metabolismo , Modelos Animais de Doenças , Cães , Interações Hospedeiro-Patógeno , Humanos , Mediadores da Inflamação/metabolismo , Vírus da Influenza A/patogenicidade , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/virologia , Células Madin Darby de Rim Canino , Camundongos Endogâmicos ICR , NF-kappa B/metabolismo , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologia , Transdução de Sinais , Replicação Viral/efeitos dos fármacos
18.
Int J Mol Sci ; 22(5)2021 Mar 04.
Artigo em Inglês | MEDLINE | ID: mdl-33806288

RESUMO

Although the cause of progressive neurodegeneration is often unclear, neuronal death can occur through several mechanisms. In conditions such as Alzheimer's or alcohol use disorder (AUD), Toll-like receptor (TLR) induction is observed with neurodegeneration. However, links between TLR activation and neurodegeneration are lacking. We report a role of apoptotic neuronal death in AUD through TLR7-mediated induction of death receptor signaling. In postmortem human cortex, a two-fold increase in apoptotic terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining in neurons was found in AUD versus controls. This occurred with the increased expression of TLR7 and tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) death receptors. Binge ethanol treatment in C57BL/6 mice increased TLR7 and induced neuronal apoptosis in cortical regions that was blocked by TLR7 antagonism. Mechanistic studies in primary organotypic brain slice culture (OBSC) found that the inhibition of TLR7 and its endogenous ligand let-7b blocked ethanol-induced neuronal cell death. Both IMQ and ethanol induced the expression of TRAIL and its death receptor. In addition, TRAIL-neutralizing monoclonal antibodies blocked both imiquimod (IMQ) and ethanol induced neuronal death. These findings implicate TRAIL as a mediator of neuronal apoptosis downstream of TLR7 activation. TLR7 and neuronal apoptosis are implicated in other neurodegenerative diseases, including Alzheimer's disease. Therefore, TRAIL may represent a therapeutic target to slow neurodegeneration in multiple diseases.


Assuntos
Alcoolismo/metabolismo , Alcoolismo/patologia , Degeneração Neural/metabolismo , Degeneração Neural/patologia , Ligante Indutor de Apoptose Relacionado a TNF/metabolismo , Adulto , Animais , Apoptose , Consumo Excessivo de Bebidas Alcoólicas/genética , Consumo Excessivo de Bebidas Alcoólicas/metabolismo , Consumo Excessivo de Bebidas Alcoólicas/patologia , Encéfalo/metabolismo , Encéfalo/patologia , Estudos de Casos e Controles , Caspase 3/metabolismo , Modelos Animais de Doenças , Humanos , Inflamação/metabolismo , Inflamação/patologia , Masculino , Glicoproteínas de Membrana/antagonistas & inibidores , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/genética , MicroRNAs/metabolismo , Pessoa de Meia-Idade , Modelos Neurológicos , Neurônios/metabolismo , Neurônios/patologia , Córtex Pré-Frontal/metabolismo , Córtex Pré-Frontal/patologia , Transdução de Sinais , Técnicas de Cultura de Tecidos , Receptor 7 Toll-Like/antagonistas & inibidores , Receptor 7 Toll-Like/metabolismo , Adulto Jovem
19.
Front Immunol ; 12: 637659, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33767707

RESUMO

Objectives: To identify the importance of the Toll-like receptor (TLR) pathway using B cell high-throughput sequencing and to explore the participation of the TLR7 signaling pathway in primary Sjogren's syndrome (pSS)-associated thrombocytopenia in patient and mouse models. Methods: High-throughput gene sequencing and bioinformatic analyses were performed for 9 patients: 3 patients with pSS and normal platelet counts, 3 patients with pSS-associated thrombocytopenia, and 3 healthy controls. Twenty-four patients with pSS were recruited for validation. Twenty-four non-obese diabetic (NOD) mice were divided into the TLR7 pathway inhibition (CA-4948), activation (Resiquimod), and control groups. Serum, peripheral blood, bone marrow, and submandibular glands were collected for thrombocytopenia and TLR7 pathway analysis. Results: Seven hub genes enriched in the TLR pathway were identified. Compared to that in control patients, the expression of interleukin (IL)-8 and TLR7 pathway molecules in B-cells was higher in patients with pSS-associated thrombocytopenia. Platelet counts exhibited a negative correlation with serum IL-1ß and IL-8 levels. In NOD mice, CA-4948/Resiquimod treatment induced the downregulation/upregulation of the TLR7 pathway, leading to consistent elevation/reduction of platelet counts. Megakaryocyte counts in the bone marrow showed an increasing trend in the Resiquimod group, with more naked nuclei. The levels of IL-1ß and IL-8 in the serum and submandibular gland tissue increased in the Resiquimod group compared with that in CA-4948 and control groups. Conclusion: pSS-associated thrombocytopenia may be a subset of the systemic inflammatory state as the TLR7 signaling pathway was upregulated in B cells of patients with pSS-associated thrombocytopenia, and activation of the TLR7 pathway led to a thrombocytopenia phenotype in NOD mice.


Assuntos
Linfócitos B/imunologia , Interleucina-1beta/sangue , Interleucina-8/sangue , Síndrome de Sjogren/patologia , Trombocitopenia/imunologia , Receptor 7 Toll-Like/metabolismo , Animais , Plaquetas/citologia , Feminino , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imidazóis/farmacologia , Masculino , Camundongos , Camundongos Endogâmicos NOD , Pessoa de Meia-Idade , Neutrófilos/imunologia , Contagem de Plaquetas , Transdução de Sinais/fisiologia , Síndrome de Sjogren/imunologia , Trombocitopenia/patologia , Receptor 7 Toll-Like/antagonistas & inibidores
20.
Commun Biol ; 4(1): 276, 2021 03 03.
Artigo em Inglês | MEDLINE | ID: mdl-33658617

RESUMO

In this work, we are reporting that "Shock and Kill", a therapeutic approach designed to eliminate latent HIV from cell reservoirs, is extrapolatable to cancer therapy. This is based on the observation that malignant cells express a spectrum of human endogenous retroviral elements (HERVs) which can be transcriptionally boosted by HDAC inhibitors. The endoretroviral gene HERV-V2 codes for an envelope protein, which resembles syncytins. It is significantly overexpressed upon exposure to HDAC inhibitors and can be effectively targeted by simultaneous application of TLR7/8 agonists, triggering intrinsic apoptosis. We demonstrated that this synergistic cytotoxic effect was accompanied by the functional disruption of the TLR7/8-NFκB, Akt/PKB, and Ras-MEK-ERK signalling pathways. CRISPR/Cas9 ablation of TLR7 and HERV-V1/V2 curtailed apoptosis significantly, proving the pivotal role of these elements in driving cell death. The effectiveness of this new approach was confirmed in ovarian tumour xenograft studies, revealing a promising avenue for future cancer therapies.


Assuntos
Adjuvantes Imunológicos/farmacologia , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Retrovirus Endógenos/efeitos dos fármacos , Inibidores de Histona Desacetilases/farmacologia , Neoplasias Ovarianas/tratamento farmacológico , Receptor 7 Toll-Like/agonistas , Ativação Viral/efeitos dos fármacos , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Depsipeptídeos/farmacologia , Retrovirus Endógenos/genética , Retrovirus Endógenos/metabolismo , Feminino , Humanos , Imiquimode/farmacologia , Imunidade Inata/efeitos dos fármacos , Camundongos Nus , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/virologia , Pteridinas/farmacologia , Transdução de Sinais , Receptor 7 Toll-Like/genética , Receptor 7 Toll-Like/metabolismo , Células Tumorais Cultivadas , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/metabolismo , Vorinostat/farmacologia , Ensaios Antitumorais Modelo de Xenoenxerto
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