RESUMO
Although animal models have been evaluated for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, none have fully recapitulated the lung disease phenotypes seen in humans who have been hospitalized. Here, we evaluate transgenic mice expressing the human angiotensin I-converting enzyme 2 (ACE2) receptor driven by the cytokeratin-18 (K18) gene promoter (K18-hACE2) as a model of SARS-CoV-2 infection. Intranasal inoculation of SARS-CoV-2 in K18-hACE2 mice results in high levels of viral infection in lungs, with spread to other organs. A decline in pulmonary function occurs 4 days after peak viral titer and correlates with infiltration of monocytes, neutrophils and activated T cells. SARS-CoV-2-infected lung tissues show a massively upregulated innate immune response with signatures of nuclear factor-κB-dependent, type I and II interferon signaling, and leukocyte activation pathways. Thus, the K18-hACE2 model of SARS-CoV-2 infection shares many features of severe COVID-19 infection and can be used to define the basis of lung disease and test immune and antiviral-based countermeasures.
Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/patologia , Imunidade Inata/imunologia , Peptidil Dipeptidase A/genética , Pneumonia Viral/patologia , Pneumonia/patologia , Enzima de Conversão de Angiotensina 2 , Animais , COVID-19 , Chlorocebus aethiops , Infecções por Coronavirus/imunologia , Modelos Animais de Doenças , Feminino , Humanos , Interferon Tipo I/imunologia , Interferon gama/imunologia , Queratina-18/genética , Leucócitos/imunologia , Ativação Linfocitária/imunologia , Masculino , Camundongos , Camundongos Transgênicos , Monócitos/imunologia , NF-kappa B/imunologia , Infiltração de Neutrófilos/imunologia , Neutrófilos/imunologia , Pandemias , Pneumonia/genética , Pneumonia/virologia , Pneumonia Viral/imunologia , Regiões Promotoras Genéticas/genética , SARS-CoV-2 , Linfócitos T/imunologia , Células Vero , Replicação Viral/imunologiaRESUMO
COVID-19, which is caused by infection with SARS-CoV-2, is characterized by lung pathology and extrapulmonary complications1,2. Type I interferons (IFNs) have an essential role in the pathogenesis of COVID-19 (refs 3-5). Although rapid induction of type I IFNs limits virus propagation, a sustained increase in the levels of type I IFNs in the late phase of the infection is associated with aberrant inflammation and poor clinical outcome5-17. Here we show that the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway, which controls immunity to cytosolic DNA, is a critical driver of aberrant type I IFN responses in COVID-19 (ref. 18). Profiling COVID-19 skin manifestations, we uncover a STING-dependent type I IFN signature that is primarily mediated by macrophages adjacent to areas of endothelial cell damage. Moreover, cGAS-STING activity was detected in lung samples from patients with COVID-19 with prominent tissue destruction, and was associated with type I IFN responses. A lung-on-chip model revealed that, in addition to macrophages, infection with SARS-CoV-2 activates cGAS-STING signalling in endothelial cells through mitochondrial DNA release, which leads to cell death and type I IFN production. In mice, pharmacological inhibition of STING reduces severe lung inflammation induced by SARS-CoV-2 and improves disease outcome. Collectively, our study establishes a mechanistic basis of pathological type I IFN responses in COVID-19 and reveals a principle for the development of host-directed therapeutics.
Assuntos
COVID-19/imunologia , COVID-19/patologia , Interferon Tipo I/imunologia , Proteínas de Membrana/metabolismo , Nucleotidiltransferases/metabolismo , SARS-CoV-2/imunologia , Animais , COVID-19/metabolismo , COVID-19/virologia , Células Cultivadas , DNA Mitocondrial/metabolismo , Modelos Animais de Doenças , Progressão da Doença , Células Endoteliais/patologia , Feminino , Regulação da Expressão Gênica/imunologia , Humanos , Imunidade Inata , Pulmão/imunologia , Pulmão/metabolismo , Pulmão/patologia , Pulmão/virologia , Macrófagos/imunologia , Proteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos C57BL , Pneumonia/imunologia , Pneumonia/metabolismo , Pneumonia/patologia , Pneumonia/virologia , SARS-CoV-2/patogenicidade , Transdução de Sinais , Pele/imunologia , Pele/metabolismo , Pele/patologiaRESUMO
Severe COVID-19 is characterized by persistent lung inflammation, inflammatory cytokine production, viral RNA and a sustained interferon (IFN) response, all of which are recapitulated and required for pathology in the SARS-CoV-2-infected MISTRG6-hACE2 humanized mouse model of COVID-19, which has a human immune system1-20. Blocking either viral replication with remdesivir21-23 or the downstream IFN-stimulated cascade with anti-IFNAR2 antibodies in vivo in the chronic stages of disease attenuates the overactive immune inflammatory response, especially inflammatory macrophages. Here we show that SARS-CoV-2 infection and replication in lung-resident human macrophages is a critical driver of disease. In response to infection mediated by CD16 and ACE2 receptors, human macrophages activate inflammasomes, release interleukin 1 (IL-1) and IL-18, and undergo pyroptosis, thereby contributing to the hyperinflammatory state of the lungs. Inflammasome activation and the accompanying inflammatory response are necessary for lung inflammation, as inhibition of the NLRP3 inflammasome pathway reverses chronic lung pathology. Notably, this blockade of inflammasome activation leads to the release of infectious virus by the infected macrophages. Thus, inflammasomes oppose host infection by SARS-CoV-2 through the production of inflammatory cytokines and suicide by pyroptosis to prevent a productive viral cycle.
Assuntos
COVID-19 , Inflamassomos , Macrófagos , SARS-CoV-2 , Enzima de Conversão de Angiotensina 2 , Animais , COVID-19/patologia , COVID-19/fisiopatologia , COVID-19/virologia , Humanos , Inflamassomos/metabolismo , Interleucina-1 , Interleucina-18 , Pulmão/patologia , Pulmão/virologia , Macrófagos/metabolismo , Macrófagos/patologia , Macrófagos/virologia , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Pneumonia/metabolismo , Pneumonia/virologia , Piroptose , Receptores de IgG , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidadeRESUMO
Immune responses are tightly regulated to ensure efficient pathogen clearance while avoiding tissue damage. Here we report that Setdb2 was the only protein lysine methyltransferase induced during infection with influenza virus. Setdb2 expression depended on signaling via type I interferons, and Setdb2 repressed expression of the gene encoding the neutrophil attractant CXCL1 and other genes that are targets of the transcription factor NF-κB. This coincided with occupancy by Setdb2 at the Cxcl1 promoter, which in the absence of Setdb2 displayed diminished trimethylation of histone H3 Lys9 (H3K9me3). Mice with a hypomorphic gene-trap construct of Setdb2 exhibited increased infiltration of neutrophils during sterile lung inflammation and were less sensitive to bacterial superinfection after infection with influenza virus. This suggested that a Setdb2-mediated regulatory crosstalk between the type I interferons and NF-κB pathways represents an important mechanism for virus-induced susceptibility to bacterial superinfection.
Assuntos
Histona-Lisina N-Metiltransferase/imunologia , NF-kappa B/imunologia , Infecções por Orthomyxoviridae/imunologia , Orthomyxoviridae/imunologia , Pneumonia/imunologia , Superinfecção/imunologia , Animais , Quimiocina CXCL1/imunologia , Suscetibilidade a Doenças , Feminino , Interferon Tipo I/imunologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Análise de Sequência com Séries de Oligonucleotídeos , Infecções por Orthomyxoviridae/enzimologia , Infecções por Orthomyxoviridae/virologia , Pneumonia/enzimologia , Pneumonia/virologia , RNA/química , RNA/genética , Reação em Cadeia da Polimerase em Tempo Real , Organismos Livres de Patógenos Específicos , Superinfecção/enzimologia , Superinfecção/microbiologiaRESUMO
The virus severe acute respiratory syndrome coronavirus 2, SARS-CoV-2, is the causative agent of the current COVID-19 pandemic. It possesses a large 30 kilobase (kb) genome that encodes structural, non-structural, and accessory proteins. Although not necessary to cause disease, these accessory proteins are known to influence viral replication and pathogenesis. Through the synthesis of novel infectious clones of SARS-CoV-2 that lack one or more of the accessory proteins of the virus, we have found that one of these accessory proteins, ORF8, is critical for the modulation of the host inflammatory response. Mice infected with a SARS-CoV-2 virus lacking ORF8 exhibit increased weight loss and exacerbated macrophage infiltration into the lungs. Additionally, infection of mice with recombinant SARS-CoV-2 viruses encoding ORF8 mutations found in variants of concern reveal that naturally occurring mutations in this protein influence disease severity. Our studies with a virus lacking this ORF8 protein and viruses possessing naturally occurring point mutations in this protein demonstrate that this protein impacts pathogenesis.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , SARS-CoV-2/genética , COVID-19/virologia , COVID-19/imunologia , COVID-19/patologia , COVID-19/genética , Camundongos , Humanos , Progressão da Doença , Proteínas Virais/genética , Proteínas Virais/metabolismo , Pulmão/virologia , Pulmão/patologia , Replicação Viral , Pneumonia/virologia , Pneumonia/patologia , Chlorocebus aethiops , Mutação , Células Vero , FemininoRESUMO
COVID-19 has affected more than half a billion people worldwide, with more than 6.3 million deaths, but the pathophysiological mechanisms involved in lethal cases and the host determinants that determine the different clinical outcomes are still unclear. In this study, we assessed lung autopsies of 47 COVID-19 patients and examined the inflammatory profiles, viral loads, and inflammasome activation. Additionally, we correlated these factors with the patient's clinical and histopathological conditions. Robust inflammasome activation was detected in the lungs of lethal cases of SARS-CoV-2. Experiments conducted on transgenic mice expressing hACE2 and infected with SARS-CoV-2 showed that Nlrp3-/- mice were protected from disease development and lethality compared to Nlrp3+/+ littermate mice, supporting the involvement of this inflammasome in disease exacerbation. An analysis of gene expression allowed for the classification of COVID-19 patients into two different clusters. Cluster 1 died with higher viral loads and exhibited a reduced inflammatory profile than Cluster 2. Illness time, mechanical ventilation time, pulmonary fibrosis, respiratory functions, histopathological status, thrombosis, viral loads, and inflammasome activation significantly differed between the two clusters. Our data demonstrated two distinct profiles in lethal cases of COVID-19, thus indicating that the balance of viral replication and inflammasome-mediated pulmonary inflammation led to different clinical outcomes. We provide important information to understand clinical variations in severe COVID-19, a process that is critical for decisions between immune-mediated or antiviral-mediated therapies for the treatment of critical cases of COVID-19.
Assuntos
COVID-19 , Pulmão , SARS-CoV-2 , Carga Viral , Replicação Viral , COVID-19/virologia , COVID-19/mortalidade , COVID-19/imunologia , COVID-19/patologia , Animais , Humanos , Camundongos , Feminino , Masculino , Pulmão/virologia , Pulmão/patologia , Pulmão/imunologia , Pessoa de Meia-Idade , Inflamassomos/imunologia , Inflamassomos/metabolismo , Idoso , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Camundongos Transgênicos , Pneumonia/virologia , Pneumonia/mortalidade , Pneumonia/imunologia , Pneumonia/patologia , Enzima de Conversão de Angiotensina 2/metabolismo , Enzima de Conversão de Angiotensina 2/genética , Camundongos Knockout , AdultoRESUMO
Acinetobacter baumannii, an opportunistic pathogen, poses a significant threat in intensive care units, leading to severe nosocomial infections. The rise of multi-drug-resistant strains, particularly carbapenem-resistant A. baumannii, has created formidable challenges for effective treatment. Given the prolonged development cycle and high costs associated with antibiotics, phages have garnered clinical attention as an alternative for combating infections caused by drug-resistant bacteria. However, the utilization of phage therapy encounters notable challenges, including the narrow host spectrum, where each phage targets a limited subset of bacteria, increasing the risk of phage resistance development. Additionally, uncertainties in immune system dynamics during treatment hinder tailoring symptomatic interventions based on patient-specific states. In this study, we isolated two A. baumannii phages from wastewater and conducted a comprehensive assessment of their potential applications. This evaluation included sequencing analysis, genome classification, pH and temperature stability assessments, and in vitro bacterial inhibition assays. Further investigations involved analyzing histological and cytokine alterations in rats undergoing phage cocktail treatment for pneumonia. The therapeutic efficacy of the phages was validated, and transcriptomic studies of rat lung tissue during phage treatment revealed crucial changes in the immune system. The findings from our study underscore the potential of phages for future development as a treatment strategy and offer compelling evidence regarding immune system dynamics throughout the treatment process.IMPORTANCEDue to the growing problem of multi-drug-resistant bacteria, the use of phages is being considered as an alternative to antibiotics, and the genetic safety and application stability of phages determine the potential of phage application. The absence of drug resistance genes and virulence genes in the phage genome can ensure the safety of phage application, and the fact that phage can remain active in a wide range of temperatures and pH is also necessary for application. In addition, the effect evaluation of preclinical studies is especially important for clinical application. By simulating the immune response situation during the treatment process through mammalian models, the changes in animal immunity can be observed, and the effect of phage therapy can be further evaluated. Our study provides compelling evidence that phages hold promise for further development as therapeutic agents for Acinetobacter baumannii infections.
Assuntos
Infecções por Acinetobacter , Acinetobacter baumannii , Bacteriófagos , Carbapenêmicos , Modelos Animais de Doenças , Terapia por Fagos , Acinetobacter baumannii/virologia , Acinetobacter baumannii/efeitos dos fármacos , Animais , Infecções por Acinetobacter/terapia , Infecções por Acinetobacter/microbiologia , Ratos , Terapia por Fagos/métodos , Carbapenêmicos/farmacologia , Bacteriófagos/fisiologia , Bacteriófagos/genética , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Farmacorresistência Bacteriana Múltipla , Masculino , Genoma Viral , Águas Residuárias , Pneumonia/terapia , Pneumonia/microbiologia , Pneumonia/virologiaRESUMO
Children are susceptible to influenza infections and can experience severe disease presentation due to a lack of or limited pre-existing immunity. Despite the disproportionate impact influenza has on this population, there is a lack of focus on pediatric influenza research, particularly when it comes to identifying the pathogenesis of long-term outcomes that persist beyond the point of viral clearance. In this study, juvenile outbred male and female mice were infected with influenza and analyzed following viral clearance to determine how sex impacts the persistent inflammatory responses to influenza. It was found that females maintained a broader cytokine response in the lung following clearance of influenza, with innate, type I and type II cytokine signatures in almost all mice. Males, on the other hand, had higher levels of IL-6 and other macrophage-related cytokines, but no evidence of a type I or type II response. The immune landscape was similar in the lungs between males and females postinfection, but males had a higher regulatory T cell to TH1 ratio compared with female mice. Cytokine production positively correlated with the frequency of TH1 cells and exudate macrophages, as well as the number of cells in the bronchoalveolar lavage fluid. Furthermore, female lungs were enriched for metabolites involved in the glycolytic pathway, suggesting glycolysis is higher in female lungs compared with males after viral clearance. These data suggest juvenile female mice have persistent and excessive lung inflammation beyond the point of viral clearance, whereas juvenile males had a more immunosuppressive phenotype.NEW & NOTEWORTHY This study identifies sex-based differences in persistent lung inflammation following influenza infection in an outbred, juvenile animal model of pediatric infection. These findings indicate the importance of considering sex and age as variable in infectious disease research.
Assuntos
Citocinas , Infecções por Orthomyxoviridae , Pneumonia , Caracteres Sexuais , Animais , Feminino , Masculino , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/virologia , Infecções por Orthomyxoviridae/patologia , Infecções por Orthomyxoviridae/metabolismo , Camundongos , Citocinas/metabolismo , Pneumonia/virologia , Pneumonia/patologia , Pneumonia/imunologia , Pneumonia/metabolismo , Pulmão/virologia , Pulmão/patologia , Pulmão/imunologia , Pulmão/metabolismo , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/patologia , Macrófagos/virologia , Fatores SexuaisRESUMO
RATIONALE: Respiratory virus-induced inflammation is the leading cause of asthma exacerbation, frequently accompanied by induction of interferon-stimulated genes (ISGs). How asthma-susceptibility genes modulate cellular response upon viral infection by fine-tuning ISG induction and subsequent airway inflammation in genetically susceptible asthma patients remains largely unknown. OBJECTIVES: To decipher the functions of gasdermin B (encoded by GSDMB) in respiratory virus-induced lung inflammation. METHODS: In two independent cohorts, we analysed expression correlation between GSDMB and ISG s. In human bronchial epithelial cell line or primary bronchial epithelial cells, we generated GSDMB-overexpressing and GSDMB-deficient cells. A series of quantitative PCR, ELISA and co-immunoprecipitation assays were performed to determine the function and mechanism of GSDMB for ISG induction. We also generated a novel transgenic mouse line with inducible expression of human unique GSDMB gene in airway epithelial cells and infected the mice with respiratory syncytial virus to determine the role of GSDMB in respiratory syncytial virus-induced lung inflammation in vivo. RESULTS: GSDMB is one of the most significant asthma-susceptibility genes at 17q21 and acts as a novel RNA sensor, promoting mitochondrial antiviral-signalling protein (MAVS)-TANK binding kinase 1 (TBK1) signalling and subsequent inflammation. In airway epithelium, GSDMB is induced by respiratory viral infections. Expression of GSDMB and ISGs significantly correlated in respiratory epithelium from two independent asthma cohorts. Notably, inducible expression of human GSDMB in mouse airway epithelium led to enhanced ISGs induction and increased airway inflammation with mucus hypersecretion upon respiratory syncytial virus infection. CONCLUSIONS: GSDMB promotes ISGs expression and airway inflammation upon respiratory virus infection, thereby conferring asthma risk in risk allele carriers.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal , Asma , Gasderminas , Proteínas Serina-Treonina Quinases , Transdução de Sinais , Animais , Humanos , Asma/metabolismo , Asma/genética , Camundongos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Serina-Treonina Quinases/genética , Camundongos Transgênicos , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Predisposição Genética para Doença , Infecções por Vírus Respiratório Sincicial/metabolismo , Infecções por Vírus Respiratório Sincicial/genética , Células Epiteliais/metabolismo , Linhagem Celular , Brônquios/metabolismo , Brônquios/patologia , Pneumonia/metabolismo , Pneumonia/genética , Pneumonia/virologia , Feminino , Pulmão/metabolismo , Pulmão/patologiaRESUMO
BACKGROUND: Pneumonia and lung cancer are both major respiratory diseases, and observational studies have explored the association between their susceptibility. However, due to the presence of potential confounders and reverse causality, the comprehensive causal relationships between pneumonia and lung cancer require further exploration. METHODS: Genome-wide association study (GWAS) summary-level data were obtained from the hitherto latest FinnGen database, COVID-19 Host Genetics Initiative resource, and International Lung Cancer Consortium. We implemented a bidirectional Mendelian randomization (MR) framework to evaluate the causal relationships between several specific types of pneumonia and lung cancer. The causal estimates were mainly calculated by inverse-variance weighted (IVW) approach. Additionally, sensitivity analyses were also conducted to validate the robustness of the causalty. RESULTS: In the MR analyses, overall pneumonia demonstrated a suggestive but modest association with overall lung cancer risk (Odds ratio [OR]: 1.21, 95% confidence interval [CI]: 1.01 - 1.44, P = 0.037). The correlations between specific pneumonia types and overall lung cancer were not as significant, including bacterial pneumonia (OR: 1.07, 95% CI: 0.91 - 1.26, P = 0.386), viral pneumonia (OR: 1.00, 95% CI: 0.95 - 1.06, P = 0.891), asthma-related pneumonia (OR: 1.18, 95% CI: 0.92 - 1.52, P = 0.181), and COVID-19 (OR: 1.01, 95% CI: 0.78 - 1.30, P = 0.952). Reversely, with lung cancer as the exposure, we observed that overall lung cancer had statistically crucial associations with bacterial pneumonia (OR: 1.08, 95% CI: 1.03 - 1.13, P = 0.001) and viral pneumonia (OR: 1.09, 95% CI: 1.01 - 1.19, P = 0.037). Sensitivity analysis also confirmed the robustness of these findings. CONCLUSION: This study has presented a systematic investigation into the causal relationships between pneumonia and lung cancer subtypes. Further prospective study is warranted to verify these findings.
Assuntos
COVID-19 , Estudo de Associação Genômica Ampla , Neoplasias Pulmonares , Análise da Randomização Mendeliana , Pneumonia , Humanos , Neoplasias Pulmonares/genética , Pneumonia/genética , Pneumonia/epidemiologia , Pneumonia/virologia , COVID-19/genética , COVID-19/complicações , COVID-19/virologia , COVID-19/epidemiologia , SARS-CoV-2/genética , Predisposição Genética para Doença , Polimorfismo de Nucleotídeo Único , Causalidade , Razão de Chances , Fatores de RiscoRESUMO
Acinetobacter baumannii, which is resistant to multiple drugs, is an opportunistic pathogen responsible for severe nosocomial infections. With no antibiotics available, phages have obtained clinical attention. However, since immunocompromised patients are often susceptible to infection, the appropriate timing of administration is particularly important. During this research, we obtained a lytic phage vB_AbaM_P1 that specifically targets A. baumannii. We then assessed its potential as a prophylactic treatment for lung infections caused by clinical strains. The virus experiences a period of inactivity lasting 30 min and produces approximately 788 particles during an outbreak. Transmission electron microscopy shows that vB_AbaM_P1 was similar to the Saclayvirus. Based on the analysis of high-throughput sequencing and bioinformatics, vB_AbaM_P1 consists of 107537 bases with a G + C content of 37.68%. It contains a total of 177 open reading frames and 14 tRNAs. No antibiotic genes were detected. In vivo experiments, using a cyclophosphamide-induced neutrophil deficiency model, tested the protective effect of phage on neutrophil-deficient rats by prophylactic application of phage. The use of phages resulted in a decrease in rat mortality caused by A. baumannii and a reduction in the bacterial burden in the lungs. Histologic examination of lung tissue revealed a decrease in the presence of immune cells. The presence of phage vB_AbaM_P1 had a notable impact on preventing A. baumannii infection, as evidenced by the decrease in oxidative stress in lung tissue and cytokine levels in serum. Our research offers more robust evidence for the early utilization of bacteriophages to mitigate A. baumannii infection. KEY POINTS: â¢A novel Saclayvirus phage infecting A. baumannii was isolated from sewage. â¢The whole genome was determined, analyzed, and compared to other phages. â¢Assaying the effect of phage in preventing infection in neutrophil-deficient models.
Assuntos
Infecções por Acinetobacter , Acinetobacter baumannii , Bacteriófagos , Genoma Viral , Acinetobacter baumannii/virologia , Acinetobacter baumannii/genética , Animais , Infecções por Acinetobacter/prevenção & controle , Infecções por Acinetobacter/microbiologia , Bacteriófagos/genética , Bacteriófagos/isolamento & purificação , Bacteriófagos/fisiologia , Ratos , Terapia por Fagos/métodos , Composição de Bases , Modelos Animais de Doenças , Genômica , Sequenciamento de Nucleotídeos em Larga Escala , Pulmão/virologia , Pulmão/microbiologia , Pneumonia/prevenção & controle , Pneumonia/microbiologia , Pneumonia/virologia , MasculinoRESUMO
Chemokine production by epithelial cells is important for neutrophil recruitment during viral infection, the appropriate regulation of which is critical for restraining inflammation and attenuating subsequent tissue damage. Epithelial cell expression of long noncoding RNAs (lncRNAs), RNA-binding proteins, and their functional interactions during viral infection and inflammation remain to be fully understood. Here, we identified an inducible lncRNA in the Cxcl2 gene locus, lnc-Cxcl2, which could selectively inhibit Cxcl2 expression in mouse lung epithelial cells but not in macrophages. lnc-Cxcl2-deficient mice exhibited increased Cxcl2 expression, enhanced neutrophils recruitment, and more severe inflammation in the lung after influenza virus infection. Mechanistically, nucleus-localized lnc-Cxcl2 bound to Cxcl2 promoter, recruited a ribonucleoprotein La, which inhibited the chromatin accessibility of chemokine promoters, and consequently inhibited Cxcl2 transcription in cis However, unlike mouse lnc-Cxcl2, human lnc-CXCL2-4-1 inhibited multiple immune cytokine expressions including chemokines in human lung epithelial cells. Together, our results demonstrate a self-protecting mechanism within epithelial cells to restrain chemokine and neutrophil-mediated inflammation, providing clues for better understanding chemokine regulation and epithelial cell function in lung viral infection.
Assuntos
Quimiocina CXCL2/genética , Infiltração de Neutrófilos/imunologia , Neutrófilos/imunologia , RNA Longo não Codificante/genética , Células A549 , Animais , Linhagem Celular Tumoral , Quimiocina CXCL2/metabolismo , Cromatina/metabolismo , Células Epiteliais/metabolismo , Células HEK293 , Ribonucleoproteínas Nucleares Heterogêneas Grupo L/genética , Humanos , Inflamação/prevenção & controle , Mediadores da Inflamação , Vírus da Influenza A/imunologia , Pulmão/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infecções por Orthomyxoviridae/imunologia , Infecções por Orthomyxoviridae/patologia , Pneumonia/imunologia , Pneumonia/patologia , Pneumonia/virologia , Regiões Promotoras Genéticas/genética , Células RAW 264.7 , Estomatite Vesicular/imunologia , Estomatite Vesicular/patologia , Vírus da Estomatite Vesicular Indiana/imunologiaRESUMO
OBJECTIVE: To identify the prevalence of respiratory syncytial virus (RSV) in a cohort of children under 5 years of age with World Health Organization (WHO)-defined pneumonia and the factors associated with developing severe RSV-associated community-acquired pneumonia (CAP) in primary care in a single centre in Northern Malawi. METHODS: The BIOmarkers TO diagnose PnEumonia (BIOTOPE) study was a prospective cohort study conducted from March to June 2016 that took place in a primary care centre in Northern Malawi. Data from this study was used to identify the characteristics of children under 5 years of age who presented with RSV and WHO-defined CAP. Means, standard deviations, medians and ranges were calculated for continuous variables. A univariate logistic regression was performed to examine the potential predictor variables. RESULTS: Four hundred and ninety-four infants presented with CAP and were eligible for inclusion in the study; RSV infection was detected in 205 (41.6%) of the infants. Eight factors were associated with increased risk for RSV CAP in the univariate model: age, born at term, presenting for care in June, crowded living environment, not being exclusively breastfed, not having received zinc or vitamin A supplementation in the last six months. Infants with RSV were more likely to have an oxygen saturation ≤92% compared to infants with other causes of pneumonia and more likely to have severe pneumonia as defined by the WHO. CONCLUSION: This study supports that RSV-associated CAP is linked to modifiable and non-modifiable risk factors; further research is indicated to determine which interventions would be most impactful. Developing and implementing an infant or maternal vaccine could be a cost-effective way to prevent RSV-associated CAP and mortality in developing nations. More research is needed to understand seasonal patterns of CAP and research over extended periods can offer valuable insights on host, environmental and pathogen-specific factors that contribute to RSV-associated CAP.
Assuntos
Infecções Comunitárias Adquiridas , Atenção Primária à Saúde , Infecções por Vírus Respiratório Sincicial , Humanos , Infecções por Vírus Respiratório Sincicial/epidemiologia , Infecções por Vírus Respiratório Sincicial/diagnóstico , Malaui/epidemiologia , Masculino , Feminino , Lactente , Estudos Prospectivos , Infecções Comunitárias Adquiridas/epidemiologia , Infecções Comunitárias Adquiridas/virologia , Pré-Escolar , Prevalência , Fatores de Risco , Vírus Sincicial Respiratório Humano/isolamento & purificação , Recém-Nascido , Pneumonia/epidemiologia , Pneumonia/virologia , Pneumonia/diagnóstico , Pneumonia Viral/epidemiologia , Pneumonia Viral/diagnósticoRESUMO
Toll-like receptor 3 (TLR3) and TLR7 genes are involved in the host immune response against viral infections including SARS-COV-2. This study aimed to investigate the association between the TLR3(rs3775290) and TLR7(rs179008) polymorphisms with the prognosis and susceptibility to COVID-19 pneumonia accompanying SARS-COV-2 infection. This case-control study included 236 individuals: 136 COVID-19 pneumonia patients and 100 age and sex-matched controls. Two polymorphisms (TLR3 rs3775290 and TLR7 rs179008) were genotyped by allelic discrimination through TaqMan real-time PCR. This study also investigated predictors of mortality in COVID-19 pneumonia through logistic regression. The mutant 'T/T' genotypes and the 'T' alleles of TLR3(rs3775290) and TLR7(rs179008) polymorphisms were significantly associated with increased risk of COVID-19 pneumonia. This study did not report association between the mutant 'T/T' genotypes of TLR3(rs3775290) and TLR7(rs179008) and the disease outcome. In multivariate analysis, the independent predictors of mortality in COVID-19 pneumonia were male sex, SPO2 ≤ 82%, INR > 1, LDH ≥ 1000 U/l, and lymphocyte count<900/mm3 (P < 0.05).
Assuntos
COVID-19/genética , Predisposição Genética para Doença/genética , Pneumonia/genética , Polimorfismo de Nucleotídeo Único , Receptor 3 Toll-Like/genética , Receptor 7 Toll-Like/genética , Idoso , Alelos , COVID-19/diagnóstico , COVID-19/virologia , Estudos de Casos e Controles , Feminino , Frequência do Gene , Genótipo , Humanos , Masculino , Pessoa de Meia-Idade , Pneumonia/diagnóstico , Pneumonia/virologia , Prognóstico , Curva ROC , Fatores de Risco , SARS-CoV-2/fisiologiaRESUMO
In December 2019, a new type of virus, coronavirus disease 2019 broke out globally and caused great harm. The virus mutates rapidly, and more research reports are urgently needed to increase our understanding of the disease. We found the reversed halo sign (RHS) occurred in the imaging manifestations of severe acute respiratory syndrome coronavirus 2 delta variant of concern pneumonia. In the absence of pathology, the mechanism is unknown. Therefore, we reported two cases of RHS and tried to speculate the pathological mechanism through multiple computed tomography follow-up comparisons to judge the prognosis of the disease.
Assuntos
COVID-19/diagnóstico por imagem , SARS-CoV-2/patogenicidade , COVID-19/patologia , COVID-19/virologia , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Pneumonia/diagnóstico por imagem , Pneumonia/patologia , Pneumonia/virologia , Prognóstico , SARS-CoV-2/isolamento & purificação , Tomografia Computadorizada por Raios XRESUMO
BACKGROUND: Coronavirus-induced disease 19 (COVID-19) infects more than three hundred and sixty million patients worldwide, and people with severe symptoms frequently die of acute respiratory distress syndrome (ARDS). Recent studies indicated that excessive neutrophil extracellular traps (NETs) contributed to immunothrombosis, thereby leading to extensive intravascular coagulopathy and multiple organ dysfunction. Thus, understanding the mechanism of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-induced NET formation would be helpful to reduce thrombosis and prevent ARDS in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. METHODS: We incubated SARS-CoV-2 with neutrophils in the presence or absence of platelets to observe NET formation. We further isolated extracellular vesicles from COVID-19 patients' sera (COVID-19-EVs) to examine their ability to induce NET formation. RESULTS: We demonstrated that antagonistic mAbs against anti-CLEC5A mAb and anti-TLR2 mAb can inhibit COVID-19-EVs-induced NET formation, and generated clec5a-/-/tlr2-/- mice to confirm the critical roles of CLEC5A and TLR2 in SARS-CoV-2-induced lung inflammation in vivo. We found that virus-free extracellular COVID-19 EVs induced robust NET formation via Syk-coupled C-type lectin member 5A (CLEC5A) and TLR2. Blockade of CLEC5A inhibited COVID-19 EVs-induced NETosis, and simultaneous blockade of CLEC5A and TLR2 further suppressed SARS-CoV-2-induced NETosis in vitro. Moreover, thromboinflammation was attenuated dramatically in clec5a-/-/tlr2-/- mice. CONCLUSIONS: This study demonstrates that SARS-CoV-2-activated platelets produce EVs to enhance thromboinflammation via CLEC5A and TLR2, and highlight the importance of CLEC5A and TLR2 as therapeutic targets to reduce the risk of ARDS in COVID-19 patients.
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COVID-19 , Lectinas Tipo C , Neutrófilos , Pneumonia , Síndrome do Desconforto Respiratório , SARS-CoV-2 , Trombose , Animais , Plaquetas/imunologia , Plaquetas/patologia , Plaquetas/virologia , COVID-19/sangue , COVID-19/imunologia , Humanos , Lectinas Tipo C/imunologia , Camundongos , Neutrófilos/imunologia , Neutrófilos/patologia , Neutrófilos/virologia , Pneumonia/imunologia , Pneumonia/patologia , Pneumonia/virologia , Receptores de Superfície Celular , Síndrome do Desconforto Respiratório/imunologia , Síndrome do Desconforto Respiratório/virologia , SARS-CoV-2/imunologia , Trombose/sangue , Trombose/imunologia , Trombose/virologia , Receptor 2 Toll-Like/imunologiaRESUMO
Background: This document provides evidence-based clinical practice guidelines on the diagnostic utility of nucleic acid-based testing of respiratory samples for viral pathogens other than influenza in adults with suspected community-acquired pneumonia (CAP).Methods: A multidisciplinary panel developed a Population-Intervention-Comparison-Outcome question, conducted a pragmatic systematic review, and applied Grading of Recommendations, Assessment, Development, and Evaluation methodology for clinical recommendations.Results: The panel evaluated the literature to develop recommendations regarding whether routine diagnostics should include nucleic acid-based testing of respiratory samples for viral pathogens other than influenza in suspected CAP. The evidence addressing this topic was generally adjudicated to be of very low quality because of risk of bias and imprecision. Furthermore, there was little direct evidence supporting a role for routine nucleic acid-based testing of respiratory samples in improving critical outcomes such as overall survival or antibiotic use patterns. However, on the basis of direct and indirect evidence, recommendations were made for both outpatient and hospitalized patients with suspected CAP. Testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection was not addressed in the literature at the time of the evidence review.Conclusions: The panel formulated and provided their rationale for recommendations on nucleic acid-based diagnostics for viral pathogens other than influenza for patients with suspected CAP.
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Infecções Comunitárias Adquiridas/virologia , DNA Viral/análise , Pneumonia/virologia , Sociedades Médicas , Vírus/genética , Infecções Comunitárias Adquiridas/diagnóstico , Humanos , Pneumonia/diagnósticoRESUMO
Emerging and re-emerging human coronaviruses (hCoVs) cause severe respiratory illness in humans, but the basis for lethal pneumonia in these diseases is not well understood. Alveolar macrophages (AMs) are key orchestrators of host antiviral defense and tissue tolerance during a variety of respiratory infections, and AM dysfunction is associated with severe COVID-19. In this study, using a mouse model of Middle East respiratory syndrome coronavirus (MERS-CoV) infection, we examined the role of AMs in MERS pathogenesis. Our results show that depletion of AMs using clodronate (CL) liposomes significantly increased morbidity and mortality in human dipeptidyl peptidase 4 knock-in (hDPP4-KI) mice. Detailed examination of control and AM-depleted lungs at different days postinfection revealed increased neutrophil activity but a significantly reduced MERS-CoV-specific CD4 T-cell response in AM-deficient lungs during later stages of infection. Furthermore, enhanced MERS severity in AM-depleted mice correlated with lung inflammation and lesions. Collectively, these data demonstrate that AMs are critical for the development of an optimal virus-specific T-cell response and controlling excessive inflammation during MERS-CoV infection.
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Infecções por Coronavirus , Macrófagos Alveolares , Coronavírus da Síndrome Respiratória do Oriente Médio , Pneumonia , Animais , Ácido Clodrônico , Infecções por Coronavirus/imunologia , Macrófagos Alveolares/imunologia , Camundongos , Camundongos Transgênicos , Pneumonia/imunologia , Pneumonia/virologiaRESUMO
Neutrophils are vital for antimicrobial defense; however, their role during viral infection is less clear. Furthermore, the molecular regulation of neutrophil fate and function at the viral infected sites is largely elusive. Here we report that BCL6 deficiency in myeloid cells exhibited drastically enhanced host resistance to severe influenza A virus (IAV) infection. In contrast to the notion that BCL6 functions to suppress innate inflammation, we find that myeloid BCL6 deficiency diminished lung inflammation without affecting viral loads. Using a series of Cre-transgenic, reporter, and knockout mouse lines, we demonstrate that BCL6 deficiency in neutrophils, but not in monocytes or lung macrophages, attenuated host inflammation and morbidity following IAV infection. Mechanistically, BCL6 bound to the neutrophil gene loci involved in cellular apoptosis in cells specifically at the site of infection. As such, BCL6 disruption resulted in increased expression of apoptotic genes in neutrophils in the respiratory tract, but not in the circulation or bone marrow. Consequently, BCL6 deficiency promoted tissue neutrophil apoptosis. Partial neutrophil depletion led to diminished pulmonary inflammation and decreased host morbidity. Our results reveal a previously unappreciated role of BCL6 in modulating neutrophil apoptosis at the site of infection for the regulation of host disease development following viral infection. Furthermore, our studies indicate that tissue-specific regulation of neutrophil survival modulates host inflammation and tissue immunopathology during acute respiratory viral infection.
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Vírus da Influenza A/patogenicidade , Neutrófilos/metabolismo , Infecções por Orthomyxoviridae/metabolismo , Pneumonia/metabolismo , Proteínas Proto-Oncogênicas c-bcl-6/metabolismo , Animais , Apoptose/fisiologia , Interações Hospedeiro-Patógeno/fisiologia , Pulmão/metabolismo , Pulmão/virologia , Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/virologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Infiltração de Neutrófilos/fisiologia , Neutrófilos/virologia , Pneumonia/virologia , Infecções Respiratórias/metabolismo , Infecções Respiratórias/virologiaRESUMO
Premature infants, especially those with bronchopulmonary dysplasia (BPD), develop recurrent severe respiratory viral illnesses. We have shown that hyperoxic exposure of immature mice, a model of BPD, increases lung IL-12-producing Clec9a+ CD103+ dendritic cells (DCs), pro-inflammatory responses, and airway hyperreactivity following rhinovirus (RV) infection. However, the requirement for CD103+ DCs and Clec9a, a DAMP receptor that binds necrotic cell cytoskeletal filamentous actin (F-actin), for RV-induced inflammatory responses has not been demonstrated. To test this, 2-day-old C57BL/6J, CD103+ DC-deficient Batf3-/- or Clec9agfp-/- mice were exposed to normoxia or hyperoxia for 14 days. Also, selected mice were treated with neutralizing antibody against CD103. Immediately after hyperoxia, the mice were inoculated with RV intranasally. We found that compared with wild-type mice, hyperoxia-exposed Batf3-/- mice showed reduced levels of IL-12p40, IFN-γ, and TNF-α, fewer IFN-γ-producing CD4+ T cells, and decreased airway responsiveness following RV infection. Similar effects were observed in anti-CD103-treated and Clec9agfp-/- mice. Furthermore, hyperoxia increased airway dead cell number and extracellular F-actin levels. Finally, studies in preterm infants with respiratory distress syndrome showed that tracheal aspirate CLEC9A expression positively correlated with IL12B expression, consistent with the notion that CLEC9A+ cells are responsible for IL-12 production in humans as well as mice. We conclude that CD103+ DCs and Clec9a are required for hyperoxia-induced pro-inflammatory responses to RV infection. In premature infants, Clec9a-mediated activation of CD103+ DCs may promote pro-inflammatory responses to viral infection, thereby driving respiratory morbidity.