RESUMEN
Silver nanoparticles (AgNPs) are a potential antiviral agent due to their ability to disrupt the viral particle or alter the virus metabolism inside the host cell. In vitro, AgNPs exhibit antiviral activity against the most common human respiratory viruses. However, their capacity to modulate immune responses during respiratory viral infections has yet to be explored. This study demonstrates that administering AgNPs directly into the lungs prior to infection can reduce viral loads and therefore virus-induced cytokines in mice infected with influenza virus or murine pneumonia virus. The prophylactic effect was diminished in mice with depleted lymphoid cells. We showed that AgNPs-treatment resulted in the recruitment and activation of lymphocytes in the lungs, particularly natural killer (NK) cells. Mechanistically, AgNPs enhanced the ability of alveolar macrophages to promote both NK cell migration and IFN-γ production. By contrast, following infection, in mice treated with AgNPs, NK cells exhibited decreased activation, indicating that these nanoparticles can regulate the potentially deleterious activation of these cells. Overall, the data suggest that AgNPs may possess prophylactic antiviral properties by recruiting and controlling the activation of lymphoid cells through interaction with alveolar macrophages.
Asunto(s)
Células Asesinas Naturales , Pulmón , Nanopartículas del Metal , Infecciones por Orthomyxoviridae , Plata , Animales , Plata/química , Plata/farmacología , Nanopartículas del Metal/química , Pulmón/virología , Pulmón/patología , Pulmón/efectos de los fármacos , Infecciones por Orthomyxoviridae/prevención & control , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Infecciones por Orthomyxoviridae/virología , Ratones , Células Asesinas Naturales/efectos de los fármacos , Macrófagos Alveolares/efectos de los fármacos , Macrófagos Alveolares/metabolismo , Macrófagos Alveolares/virología , Ratones Endogámicos C57BL , Linfocitos/efectos de los fármacos , Linfocitos/metabolismo , Antivirales/farmacología , Antivirales/uso terapéutico , Femenino , Activación de Linfocitos/efectos de los fármacosRESUMEN
Introduction: Although many studies have underscored the importance of T cells, phenotypically and functionally, fewer have studied the functions of myeloid cells in COVID disease. In particular, the potential role of myeloid cells such as monocytes and low-density neutrophils (LDNs) in innate responses and particular in the defense against secondary bacterial infections has been much less documented. Methods: Here, we compared, in a longitudinal study, healthy subjects, idiopathic fibrosis patients, COVID patients who were either hospitalized/moderate (M-) or admitted to ICU (COV-ICU) and patients in ICU hospitalized for other reasons (non-COV-ICU). Results: We show that COVID patients have an increased proportion of low-density neutrophils (LDNs), which produce high levels of proteases (particularly, NE, MMP-8 and MMP-9) (unlike non-COV-ICU patients), which are partly responsible for causing type II alveolar cell damage in co-culture experiments. In addition, we showed that M- and ICU-COVID monocytes had reduced responsiveness towards further live Pseudomonas aeruginosa (PAO1 strain) infection, an important pathogen colonizing COVID patients in ICU, as assessed by an impaired secretion of myeloid cytokines (IL-1, TNF, IL-8, ). By contrast, lymphoid cytokines (in particular type 2/type 3) levels remained high, both basally and post PAO1 infection, as reflected by the unimpaired capacity of T cells to proliferate, when stimulated with anti-CD3/CD28 beads. Discussion: Overall, our results demonstrate that COVID circulatory T cells have a biased type 2/3 phenotype, unconducive to proper anti-viral responses and that myeloid cells have a dual deleterious phenotype, through their LDN-mediated damaging effect on alveolar cells and their impaired responsiveness (monocyte-mediated) towards bacterial pathogens such as P. aeruginosa.
Asunto(s)
COVID-19 , Monocitos , Neutrófilos , Infecciones por Pseudomonas , Pseudomonas aeruginosa , SARS-CoV-2 , Humanos , COVID-19/inmunología , Pseudomonas aeruginosa/inmunología , Monocitos/inmunología , Masculino , Femenino , Persona de Mediana Edad , SARS-CoV-2/inmunología , SARS-CoV-2/fisiología , Infecciones por Pseudomonas/inmunología , Neutrófilos/inmunología , Anciano , Citocinas/metabolismo , Citocinas/inmunología , Adulto , Estudios Longitudinales , Leucocitos Mononucleares/inmunología , Pulmón/inmunología , Pulmón/patología , Pulmón/microbiologíaRESUMEN
Epidemiological studies showed a positive association between exposure to PM2.5 and the severity of influenza virus infection. However, the mechanisms by which PM2.5 can disrupt antiviral defence are still unclear. From this perspective, the objective of this study was to evaluate the effects of PM2.5 on antiviral signalling in the respiratory epithelium using the bronchial Calu-3 cell line grown at the air-liquid interface. Pre-exposure to PM2.5 before infection with the influenza virus was investigated, as well as a co-exposure. Although a physical interaction between the virus and the particles seems possible, no effect of PM2.5 on viral replication was observed during co-exposure, although a downregulation of IFN-ß release was associated to PM2.5 exposure. However, pre-exposure slightly increased the viral nucleoprotein production and the pro-inflammatory response. Conversely, the level of the myxovirus resistance protein A (MxA), an interferon-stimulated gene (ISG) induced by IFN-ß, was reduced. Therefore, these results suggest that pre-exposure to PM2.5 could alter the antiviral response of bronchial epithelial cells, increasing their susceptibility to viral infection.
Asunto(s)
Gripe Humana , Orthomyxoviridae , Virosis , Humanos , Interferones , Gripe Humana/genética , Gripe Humana/metabolismo , Mucosa Respiratoria , Antivirales , Epitelio/metabolismo , Material Particulado/toxicidadRESUMEN
One billion people worldwide get flu every year, including patients with non-small cell lung cancer (NSCLC). However, the impact of acute influenza A virus (IAV) infection on the composition of the tumor microenvironment (TME) and the clinical outcome of patients with NSCLC is largely unknown. We set out to understand how IAV load impacts cancer growth and modifies cellular and molecular players in the TME. Herein, we report that IAV can infect both tumor and immune cells, resulting in a long-term protumoral effect in tumor-bearing mice. Mechanistically, IAV impaired tumor-specific T-cell responses, led to the exhaustion of memory CD8+ T cells and induced PD-L1 expression on tumor cells. IAV infection modulated the transcriptomic profile of the TME, fine-tuning it toward immunosuppression, carcinogenesis, and lipid and drug metabolism. Consistent with these data, the transcriptional module induced by IAV infection in tumor cells in tumor-bearing mice was also found in human patients with lung adenocarcinoma and correlated with poor overall survival. In conclusion, we found that IAV infection worsened lung tumor progression by reprogramming the TME toward a more aggressive state.
Asunto(s)
Carcinoma de Pulmón de Células no Pequeñas , Virus de la Influenza A , Gripe Humana , Neoplasias Pulmonares , Infecciones por Orthomyxoviridae , Humanos , Animales , Ratones , Neoplasias Pulmonares/patología , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/patología , Microambiente Tumoral , Linfocitos T CD8-positivos , Pulmón , Infecciones por Orthomyxoviridae/patologíaRESUMEN
Eosinophils have been previously shown to be able to regulate early humoral responses during systemic vaccination. Here we investigated the role of eosinophils during pulmonary vaccination, comparing vaccine-induced responses in eosinophil-deficient (ΔdblGATA) and wild-type mice using a Th2 adjuvant. We observed that eosinophils were needed to induce a complete vaccine response, thereby eliciting specific antibody-secreting plasma cells in the regional lymph nodes and antibody secretion in the BAL at the early stage of the immune response. Reintroduction of eosinophils in the lungs of ΔdblGATA mice during the priming stage enhanced both specific IgM and IgG plasma cells but not specific IgA plasma cells. Upon vaccination, eosinophils migrated to the lungs and secreted cytokines involved in B-cell activation, which might promote antibody production. Importantly, however, the absence of eosinophils did not impair late immune responses in a prime/boost protocol because, in that setup, we uncovered a compensating mechanism involving a Th17 pathway. In conclusion, our data demonstrate for the first time a new role for eosinophils during lung mucosal vaccination, whereby they accelerate early immune responses (IgM and IgG) while regulating IgA production at the late stages.
Asunto(s)
Formación de Anticuerpos , Eosinófilos , Ratones , Animales , Eosinófilos/metabolismo , Pulmón/patología , Vacunación , Inmunoglobulina G , Inmunoglobulina M , Inmunoglobulina A/metabolismo , Ratones Endogámicos BALB C , Inmunidad MucosaRESUMEN
Pseudomonas aeruginosa (P.a) is a pathogen causing significant morbidity and mortality, particularly in hospital patients undergoing ventilation and in individuals with cystic fibrosis. Although we and others have investigated mechanisms used by P.a to subvert innate immunity, relatively less is known about the potential strategies used by this bacterium to fight the adaptive immune system and, in particular, T cells. Here, using RAG KO (devoid of 'classical' αß and γδ TCR T lymphocytes) and double RAG γC KO mice (devoid of T, NK and ILC cells), we demonstrate that the lymphocytic compartment is important to combat P.a (PAO1 strain). Indeed, we show that PAO1 load was increased in double RAG γC KO mice. In addition, we show that PAO1 down-regulates IL-23 and IL-22 protein accumulation in the lungs of infected mice while up-regulating their RNA production, thereby pointing towards a specific post-transcriptional regulatory mechanism not affecting other inflammatory mediators. Finally, we demonstrate that an adenovirus-mediated over-expression of IL-1, IL-23 and IL-7 induced lung neutrophil and lymphocytic influx and rescued mice against P.a-induced lethality in all WT, RAG γC KO and RAG γC KO RAG-deficient mice, suggesting that this regimen might be of value in 'locally immunosuppressed' individuals such as cystic fibrosis patients.
Asunto(s)
Fibrosis Quística , Infecciones por Pseudomonas , Animales , Fibrosis Quística/genética , Fibrosis Quística/metabolismo , Interleucina-23/metabolismo , Interleucinas , Pulmón/metabolismo , Ratones , Ratones Noqueados , Infecciones por Pseudomonas/metabolismo , Pseudomonas aeruginosa , Interleucina-22RESUMEN
Pseudomonas aeruginosa (P.a) infections are a major public health issue in ventilator-associated pneumoniae, cystic fibrosis, and chronic obstructive pulmonary disease exacerbations. P.a is multidrug resistant, and there is an urgent need to develop new therapeutic approaches. Here, we evaluated the effect of direct pulmonary transplantation of gene-modified (elafin and interleukin [IL]-6) syngeneic macrophages in a mouse model of acute P.a infection. Wild-type (WT) or Elafin-transgenic (eTg) alveolar macrophages (AMs) or bone marrow-derived macrophages (BMDMs) were recovered from bronchoalveolar lavage or generated from WT or eTg mouse bone marrow. Cells were modified with adenovirus IL-6 (Ad-IL-6), characterized in vitro, and transferred by oropharyngeal instillation in the lungs of naive mice. The protective effect was assessed during P.a acute infection (survival studies, mechanistic studies of the inflammatory response). We show that a single bolus of genetically modified syngeneic AMs or BMDMs provided protection in our P.a-induced model. Mechanistically, Elafin-modified AMs had an IL-6-IL-10-IL-4R-IL-22-antimicrobial molecular signature that, in synergy with IL-6, enhanced epithelial cell proliferation and tissue repair in the alveolar unit. We believe that this innovative cell therapy strategy could be of value in acute bacterial infections in the lung.
Asunto(s)
Infecciones por Pseudomonas , Animales , Elafina , Inmunoterapia , Interleucina-6/genética , Pulmón/microbiología , Macrófagos , Macrófagos Alveolares , Ratones , Ratones Endogámicos C57BL , Infecciones por Pseudomonas/genética , Infecciones por Pseudomonas/microbiología , Infecciones por Pseudomonas/terapia , Pseudomonas aeruginosa/genéticaRESUMEN
Influenza virus infection leads to severe and complicated disease, particularly in patients with lung cancer. It alters the tumor microenvironment (TME), which may potentiate lung cancer progression and disrupt responses to antitumoral treatments. Consequently, influenza vaccination and antiviral treatments should be recommended to all patients with lung cancer.
Asunto(s)
Antivirales/uso terapéutico , Virus de la Influenza A/inmunología , Vacunas contra la Influenza/administración & dosificación , Gripe Humana/terapia , Neoplasias Pulmonares/mortalidad , Progresión de la Enfermedad , Humanos , Gripe Humana/complicaciones , Gripe Humana/inmunología , Gripe Humana/virología , Pulmón/inmunología , Pulmón/patología , Pulmón/virología , Neoplasias Pulmonares/complicaciones , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/patología , Oncología Médica/normas , Guías de Práctica Clínica como Asunto , Microambiente Tumoral/inmunología , Vacunación/normasRESUMEN
Individuals with impaired immune responses, such as ventilated and cystic fibrosis patients are often infected with Pseudomonas aeruginosa (P.a) bacteria, and a co-infection with the Influenza virus (IAV) is often present. It has been known for many years that infection with IAV predisposes the host to secondary bacterial infections (such as Streptococcus pneumoniae or Staphylococcus aureus), and there is an abundance of mechanistic studies, including those studying the role of desensitization of TLR signaling, type I IFN- mediated impairment of neutrophil chemokines and antimicrobial production, attenuation of IL1ß production etc., showing this. However, little is known about the mechanistic events underlying the potential deleterious synergy between Influenza and P.a co-infections. We demonstrate here in vitro in epithelial cells and in vivo in three independent models (two involving mice given IAV +/- P.a, and one involving mice given IAV +/- IL-1ß) that IAV promotes secondary P.a-mediated lung disease or augmented IL-1ß-mediated inflammation. We show that IAV-P.a-mediated deleterious responses includes increased matrix metalloprotease (MMP) activity, and MMP-9 in particular, and that the use of the MMP inhibitor improves lung resilience. Furthermore, we show that IAV post-transcriptionally inhibits the antimicrobial/anti-protease molecule elafin/trappin-2, which we have shown previously to be anti-inflammatory and to protect the host against maladaptive neutrophilic inflammation in P.a infections. Our work highlights the capacity of IAV to promote further P.a-mediated lung damage, not necessarily through its interference with host resistance to the bacterium, but by down-regulating tissue resilience to lung inflammation instead. Our study therefore suggests that restoring tissue resilience in clinical settings where IAV/P.a co-exists could prove a fruitful strategy.
Asunto(s)
Coinfección/inmunología , Elafina/metabolismo , Virus de la Influenza A/inmunología , Metaloproteinasa 9 de la Matriz/metabolismo , Pseudomonas aeruginosa/inmunología , Animales , Línea Celular , Coinfección/inducido químicamente , Coinfección/metabolismo , Fibrosis Quística/inmunología , Citocinas/metabolismo , Susceptibilidad a Enfermedades/metabolismo , Células Epiteliales/metabolismo , Humanos , Inflamación/inducido químicamente , Inflamación/inmunología , Pulmón/inmunología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Modelos Animales , Neumonía/metabolismo , Infecciones Estafilocócicas/inmunologíaRESUMEN
Most of current influenza virus vaccines fail to develop a strong immunity at lung mucosae (site of viral entry) due to sub-optimal vaccination protocols (e.g. inactivated virus administered by parenteral injections). Mucosal immunity could be improved by using locally-delivered vaccines containing appropriate adjuvants. Here we show, in a mouse model, that inclusion of silver nanoparticles (AgNPs) in virus-inactivated flu vaccine resulted in reduction of viral loads and prevention of excessive lung inflammation following influenza infection. Concomitantly, AgNPs enhanced specific IgA secreting plasma cells and antibodies titers, a hallmark of successful mucosal immunity. Moreover, vaccination in the presence of AgNPs but not with gold nanoparticles, protected mice from lethal flu. Compared with other commercial adjuvants (squalene/oil-based emulsion) or silver salts, AgNPs stimulated stronger antigen specific IgA production with lower toxicity by promoting bronchus-associated lymphoid tissue (BALT) neogenesis, and acted as a bona fide mucosal adjuvant.
Asunto(s)
Adyuvantes Inmunológicos/farmacología , Inmunidad Mucosa , Inmunoglobulina A/metabolismo , Vacunas contra la Influenza/inmunología , Gripe Humana/inmunología , Tejido Linfoide/inmunología , Nanopartículas del Metal/química , Plata/química , Animales , Bronquios/inmunología , Perros , Centro Germinal/efectos de los fármacos , Centro Germinal/metabolismo , Humanos , Inmunidad Mucosa/efectos de los fármacos , Inflamación/patología , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/inmunología , Tejido Linfoide/efectos de los fármacos , Células de Riñón Canino Madin Darby , Nanopartículas del Metal/ultraestructura , Ratones Endogámicos C57BL , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/prevención & control , Infecciones por Orthomyxoviridae/virología , VacunaciónRESUMEN
Silver nanoparticles (AgNPs) are microbicidal agents which could be potentially used as an alternative to antivirals to treat human infectious diseases, especially influenza virus infections where antivirals have generally proven unsuccessful. However, concerns about the use of AgNPs on humans arise from their potential toxicity, although mechanisms are not well-understood. We show here, in the context of an influenza virus infection of lung epithelial cells, that AgNPs down-regulated influenza induced CCL-5 and -IFN-ß release (two cytokines important in antiviral immunity) through RIG-I inhibition, while enhancing IL-8 production, a cytokine important for mobilizing host antibacterial responses. AgNPs activity was independent of coating and was not observed with gold nanoparticles. Down-stream analysis indicated that AgNPs disorganized the mitochondrial network and prevented the antiviral IRF-7 transcription factor influx into the nucleus. Importantly, we showed that the modulation of RIG-I-IRF-7 pathway was concomitant with inhibition of either classical or alternative autophagy (ATG-5- and Rab-9 dependent, respectively), depending on the epithelial cell type used. Altogether, this demonstration of a AgNPs-mediated functional dichotomy (down-regulation of IFN-dependent antiviral responses and up-regulation of IL-8-dependent antibacterial responses) may have practical implications for their use in the clinic.
Asunto(s)
Antivirales/farmacología , Células Epiteliales/efectos de los fármacos , Pulmón/efectos de los fármacos , Nanopartículas del Metal/química , Mitocondrias/efectos de los fármacos , Orthomyxoviridae/efectos de los fármacos , Plata/farmacología , Tretinoina/farmacología , Animales , Antivirales/química , Autofagia/efectos de los fármacos , Línea Celular Tumoral , Perros , Células Epiteliales/metabolismo , Células Epiteliales/virología , Humanos , Pulmón/metabolismo , Pulmón/virología , Células de Riñón Canino Madin Darby/efectos de los fármacos , Células de Riñón Canino Madin Darby/virología , Pruebas de Sensibilidad Microbiana , Mitocondrias/metabolismo , Plata/química , Tretinoina/químicaRESUMEN
BACKGROUND: Pseudomonas aeruginosa lung infections are a huge problem in ventilator-associated pneumonia, cystic fibrosis (CF) and in chronic obstructive pulmonary disease (COPD) exacerbations. This bacterium secretes virulence factors that may subvert host innate immunity. OBJECTIVE: We evaluated the effect of P. aeruginosa elastase LasB, an important virulence factor secreted by the type II secretion system, on ion transport, innate immune responses and epithelial repair, both in vitro and in vivo. METHODS: Wild-type (WT) or cystic fibrosis transmembrane conductance regulator (CFTR)-mutated epithelial cells (cell lines and primary cells from patients) were treated with WT or ΔLasB pseudomonas aeruginosa O1 (PAO1) secretomes. The effect of LasB and PAO1 infection was also assessed in vivo in murine models. RESULTS: We showed that LasB was the most abundant protein in WT PAO1 secretomes and that it decreased epithelial CFTR expression and activity. In airway epithelial cell lines and primary bronchial epithelial cells, LasB degraded the immune mediators interleukin (IL)-6 and trappin-2, an important epithelial-derived antimicrobial molecule. We further showed that an IL-6/STAT3 signalling pathway was downregulated by LasB, resulting in inhibition of epithelial cell repair. In mice, intranasally instillated LasB induced significant weight loss, inflammation, injury and death. By contrast, we showed that overexpression of IL-6 and trappin-2 protected mice against WT-PAO1-induced death, by upregulating IL-17/IL-22 antimicrobial and repair pathways. CONCLUSIONS: Our data demonstrate that PAO1 LasB is a major P. aeruginosa secreted factor that modulates ion transport, immune response and tissue repair. Targeting this virulence factor or upregulating protective factors such as IL-6 or antimicrobial molecules such as trappin-2 could be beneficial in P. aeruginosa-infected individuals.
Asunto(s)
Proteínas Bacterianas , Regulador de Conductancia de Transmembrana de Fibrosis Quística/fisiología , Fibrosis Quística/inmunología , Células Epiteliales/fisiología , Inmunidad Innata/fisiología , Interleucina-6/fisiología , Metaloendopeptidasas , Animales , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones TransgénicosRESUMEN
According to the WHO, and despite reduction in mortality rates, there were an estimated 438 000 malaria deaths in 2015. Therefore new antimalarials capable of limiting organ damage are still required. We show that systemic and lung adenovirus (Ad)-mediated over-expression of trappin-2 (T-2) an antibacterial molecule with anti-inflammatory activity, increased mice survival following infection with the cerebral malaria-inducing Plasmodium berghei ANKA (PbANKA) strain. Systemically, T-2 reduced PbANKA sequestration in spleen, lung, liver and brain, associated with a decrease in pro-inflammatory cytokines (eg TNF-α in spleen and lung) and an increase in IL-10 production in the lung. Similarly, local lung instillation of Ad-T-2 resulted in a reduced organ parasite sequestration and a shift towards an anti-inflammatory/repair response, potentially implicating monocytes in the protective phenotype. Relatedly, we demonstrated in vitro that human monocytes incubated with Plasmodium falciparum-infected red blood cells (Pf-iRBCs) and IgGs from hyper-immune African human sera produced T-2 and that the latter colocalized with merozoites and inhibited Pf multiplication. This array of data argues for the first time for the potential therapeutic usefulness of this host defense peptide in human malaria patients, with the aim to limit acute lung injury and respiratory distress syndrom often observed during malaria episodes.
Asunto(s)
Antiinfecciosos/uso terapéutico , Antiparasitarios/uso terapéutico , Elafina/uso terapéutico , Malaria Cerebral/tratamiento farmacológico , Malaria Cerebral/parasitología , Plasmodium berghei/efectos de los fármacos , Administración Intranasal , Animales , Antiinfecciosos/farmacología , Antiparasitarios/farmacología , Citocinas/genética , Citocinas/metabolismo , Modelos Animales de Enfermedad , Elafina/farmacología , Eritrocitos/parasitología , Femenino , Humanos , Malaria Cerebral/sangre , Merozoítos/metabolismo , Ratones Endogámicos C57BL , Monocitos/metabolismo , Parasitemia/tratamiento farmacológico , Parasitemia/parasitología , Parasitemia/patología , Plasmodium falciparum/crecimiento & desarrollo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factor de Transcripción STAT3/metabolismoRESUMEN
BACKGROUND: The lung epithelium constitutes the first barrier against invading pathogens and also a major surface potentially exposed to nanoparticles. In order to ensure and preserve lung epithelial barrier function, the alveolar compartment possesses local defence mechanisms that are able to control bacterial infection. For instance, alveolar macrophages are professional phagocytic cells that engulf bacteria and environmental contaminants (including nanoparticles) and secrete pro-inflammatory cytokines to effectively eliminate the invading bacteria/contaminants. The consequences of nanoparticle exposure in the context of lung infection have not been studied in detail. Previous reports have shown that sequential lung exposure to nanoparticles and bacteria may impair bacterial clearance resulting in increased lung bacterial loads, associated with a reduction in the phagocytic capacity of alveolar macrophages. RESULTS: Here we have studied the consequences of SiO2 nanoparticle exposure on Pseudomonas aeruginosa clearance, Pseudomonas aeruginosa-induced inflammation and lung injury in a mouse model of acute pneumonia. We observed that pre-exposure to SiO2 nanoparticles increased mice susceptibility to lethal pneumonia but did not modify lung clearance of a bioluminescent Pseudomonas aeruginosa strain. Furthermore, internalisation of SiO2 nanoparticles by primary alveolar macrophages did not reduce the capacity of the cells to clear Pseudomonas aeruginosa. In our murine model, SiO2 nanoparticle pre-exposure preferentially enhanced Pseudomonas aeruginosa-induced lung permeability (the latter assessed by the measurement of alveolar albumin and IgM concentrations) rather than contributing to Pseudomonas aeruginosa-induced lung inflammation (as measured by leukocyte recruitment and cytokine concentration in the alveolar compartment). CONCLUSIONS: We show that pre-exposure to SiO2 nanoparticles increases mice susceptibility to lethal pneumonia but independently of macrophage phagocytic function. The deleterious effects of SiO2 nanoparticle exposure during Pseudomonas aeruginosa-induced pneumonia are related to alterations of the alveolar-capillary barrier rather than to modulation of the inflammatory responses.
Asunto(s)
Permeabilidad Capilar/efectos de los fármacos , Nanopartículas/toxicidad , Neumonía Bacteriana/inducido químicamente , Infecciones por Pseudomonas/inducido químicamente , Pseudomonas aeruginosa/patogenicidad , Alveolos Pulmonares/efectos de los fármacos , Óxidos de Selenio/toxicidad , Animales , Líquido del Lavado Bronquioalveolar/química , Líquido del Lavado Bronquioalveolar/citología , Líquido del Lavado Bronquioalveolar/microbiología , Citocinas/análisis , Inmunoglobulina M/análisis , Exposición por Inhalación , Macrófagos Alveolares/efectos de los fármacos , Macrófagos Alveolares/inmunología , Masculino , Ratones Endogámicos C57BL , Nanopartículas/química , Tamaño de la Partícula , Fagocitosis/efectos de los fármacos , Neumonía Bacteriana/inmunología , Neumonía Bacteriana/microbiología , Infecciones por Pseudomonas/inmunología , Infecciones por Pseudomonas/microbiología , Alveolos Pulmonares/irrigación sanguínea , Óxidos de Selenio/química , Propiedades de Superficie , Análisis de SupervivenciaRESUMEN
Because of an increasing exposure to environmental and occupational nanoparticles (NPs), the potential risk of these materials for human health should be better assessed. Since one of the main routes of entry of NPs is via the lungs, it is of paramount importance to further characterize their impact on the respiratory system. Here, we have studied the uptake of fluorescently labeled SiO2 NPs (50 and 100 nm) by epithelial cells (NCI-H292) and alveolar macrophages (MHS) in the presence or absence of pulmonary surfactant. The quantification of NP uptake was performed by measuring cell-associated fluorescence using flow cytometry and spectrometric techniques in order to identify the most suitable methodology. Internalization was shown to be time and dose dependent, and differences in terms of uptake were noted between epithelial cells and macrophages. In the light of our observations, we conclude that flow cytometry is a more reliable technique for the study of NP internalization, and importantly, that the hydrophobic fraction of lung surfactant is critical for downregulating NP uptake in both cell types.
Asunto(s)
Productos Biológicos/farmacología , Monitoreo del Ambiente/métodos , Células Epiteliales/metabolismo , Macrófagos Alveolares/metabolismo , Nanopartículas/administración & dosificación , Tamaño de la Partícula , Fosfolípidos/farmacología , Surfactantes Pulmonares/farmacología , Animales , Línea Celular , Citometría de Flujo/métodos , Humanos , Pulmón/metabolismo , Ratones , Microscopía Confocal , Dióxido de Silicio/metabolismo , Espectrofotometría/métodosRESUMEN
Pseudomonas aeruginosa is an opportunistic pathogen involved in nosocomial infections. Flagellin is a P. aeruginosa virulence factor involved in host response to this pathogen. We examined the role of flagellin in P. aeruginosa-induced mucus secretion. Using a mouse model of pulmonary infection we showed that PAK, a wild type strain of P. aeruginosa, induced airway mucus secretion and mucin muc5ac expression at higher levels than its flagellin-deficient mutant (ΔFliC). PAK induced expression of MUC5AC and MUC2 in both human airway epithelial NCI-H292 cell line and in primary epithelial cells. In contrast, ΔFliC infection had lower to no effect on MUC5AC and MUC2 expressions. A purified P. aeruginosa flagellin induced MUC5AC expression in parallel to IL-8 secretion in NCI-H292 cells. Accordingly, ΔFliC mutant stimulated IL-8 secretion at significantly lower levels compared to PAK. Incubation of NCI-H292 cells with exogenous IL-8 induced MUC5AC expression and pre-incubation of these cells with an anti-IL-8 antibody abrogated flagellin-mediated MUC5AC expression. Silencing of TLR5 and Naip, siRNA inhibited both flagellin-induced MUC5AC expression and IL-8 secretion. Finally, inhibition of ERK abolished the expression of both PAK- and flagellin-induced MUC5AC. We conclude that: (i) flagellin is crucial in P. aeruginosa-induced mucus hyper-secretion through TLR5 and Naip pathways; (ii) this process is mediated by ERK and amplified by IL-8. Our findings help understand the mechanisms involved in mucus secretion during pulmonary infectious disease induced by P. aeruginosa, such as in cystic fibrosis.
Asunto(s)
Flagelina/metabolismo , Moco/metabolismo , Infecciones por Pseudomonas/metabolismo , Pseudomonas aeruginosa/metabolismo , Mucosa Respiratoria/metabolismo , Transducción de Señal , Animales , Línea Celular , Femenino , Flagelina/inmunología , Regulación de la Expresión Génica/inmunología , Humanos , Interleucina-8/biosíntesis , Interleucina-8/inmunología , Ratones , Mucina 5AC/biosíntesis , Mucina 5AC/inmunología , Mucina 2/biosíntesis , Mucina 2/inmunología , Moco/inmunología , Proteína Inhibidora de la Apoptosis Neuronal/inmunología , Proteína Inhibidora de la Apoptosis Neuronal/metabolismo , Infecciones por Pseudomonas/inmunología , Pseudomonas aeruginosa/inmunología , Pseudomonas aeruginosa/patogenicidad , Mucosa Respiratoria/inmunología , Mucosa Respiratoria/microbiología , Receptor Toll-Like 5/inmunología , Receptor Toll-Like 5/metabolismoRESUMEN
Trappin-2/Elafin is a potent serine protease inhibitor which prevents excessive damage under inflammatory status. This "alarm-antiprotease" is locally expressed by epithelial cells and immune cells such as macrophages and γδ T cells. It has also been proven to modulate a wide range of parameters that are critical for the inflammation process like modulating the NFκB pathway, cytokine secretion and cell recruitment. In addition, Trappin-2/Elafin was shown to possess anti-microbial properties against different classes of pathogens including viruses, fungi and bacteria. Studies also linked Trappin-2/Elafin to either susceptibility or protection against inflammatory disease and infections, even though the mechanisms remains poorly understood. This review will discuss some of the pleiotropic effects displayed by Trappin-2/Elafin, and the properties that could be used to prevent infection or to protect against inflammation.
Asunto(s)
Elafina/inmunología , Inmunidad/inmunología , Inflamación/inmunología , Infecciones Bacterianas/inmunología , Elafina/metabolismo , Células Epiteliales/inmunología , Células Epiteliales/metabolismo , Humanos , Inflamación/metabolismo , Mediadores de Inflamación/inmunología , Mediadores de Inflamación/metabolismo , Modelos Inmunológicos , Micosis/inmunología , Linfocitos T/inmunología , Linfocitos T/metabolismo , Virosis/inmunologíaRESUMEN
Mucins, the main glycoproteins present within mucus, modulate the rheologic properties of airways and participate in lung defense. They are thought to be able to trap and eliminate microorganisms from the lung. Among the mucins secreted in the lung, MUC5AC is the most prominent factor secreted by surface epithelial cells. Although much is known about the signaling pathways involved in the regulation of MUC5AC by host factors such as cytokines or proteases, less is known about the pathways triggered by microorganisms and, specifically, by influenza A virus (IAV). We therefore set up experiments to dissect the molecular mechanisms responsible for the potential modulation of MUC5AC by IAV. Using epithelial cells, C57/Bl6 mice, and IAV strains, we measured MUC5AC expression at the RNA and protein levels, specificity protein 1 (Sp1) activation, and protease activity. Intermediate molecular partners were confirmed using pharmacological inhibitors, blocking antibodies, and small interfering (si)RNAs. We showed in vitro and in vivo that IAV up-regulates epithelial cell-derived MUC5AC and Muc5ac expression in mice, both at transcriptional (through the induction of Sp1) and translational levels. In addition, we determined that this induction was dependent on a protease-epithelial growth factor receptor-extracellular regulated kinase-Sp1 signaling cascade, involving in particular the human airway trypsin. Our data point to MUC5AC as a potential modulatory mechanism by which the lung epithelia respond to IAV infection, and we dissect, for the first time to the best of our knowledge, the molecular partners involved. Future experiments using MUC5AC-targeted strategies should help further unravel the pathophysiological consequences of IAV-induced MUC5AC expression for lung homeostasis.
Asunto(s)
Receptores ErbB/metabolismo , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Virus de la Influenza A/metabolismo , Pulmón/metabolismo , Mucina 5AC/biosíntesis , Péptido Hidrolasas/genética , Factor de Transcripción Sp1/metabolismo , Proteínas ADAM/genética , Proteínas ADAM/metabolismo , Proteína ADAM17 , Animales , Línea Celular Tumoral , Células Epiteliales/metabolismo , Células Epiteliales/virología , Receptores ErbB/genética , Quinasas MAP Reguladas por Señal Extracelular/genética , Humanos , Gripe Humana/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Mucina 5AC/genética , Mucina 5AC/metabolismo , Péptido Hidrolasas/metabolismo , Transducción de Señal , Factor de Transcripción Sp1/genética , Tripsina/genética , Tripsina/metabolismo , Regulación hacia Arriba , Replicación Viral/genéticaRESUMEN
Eicosanoids are metabolites of arachidonic acid produced by cyclooxygenases (COXs) or lipoxygenases (LOXs). They mediate inflammation and mucus secretion in chronic pulmonary inflammatory diseases. The gel-forming mucin MUC5AC is over-expressed in the airways of patients with these diseases. MUC5AC expression is mediated by an extracellular signal-regulated kinase (ERK)/Sp1 dependent mechanism. Our aim was to study the role of eicosanoids and their signalling pathways in MUC5AC expression. Inhibitors of 12-LOX, but not those of COX, 5-LOX or 15-LOX, reduce MUC5AC expression induced by phorbol myristate acetate (PMA) in the bronchial epithelial cell line NCI-H292. These inhibitors also abrogate the production of whole mucus by cell monolayers. Two forms of 12-LOX (R and S) exist in mammals. Using siRNAs we show that 12R-LOX but not 12S-LOX is involved in MUC5AC expression induced by PMA, lipopolysaccharide or transforming growth factor-α. 12R-LOX also participates in MUC2 and MUC5B expression, although to a lesser extent than for MUC5AC. Contrarily, 12R-LOX silencing does not modify interleukin-8 production. 12-LOX inhibitors reduce ERK activation and Sp1 translocation induced by PMA. Moreover, the 12R-LOX product 12(R)-hydroxyeicosatetraenoic acid, induces MUC5AC expression, ERK activation and Sp1 translocation. 12R-LOX is involved in MUC5AC expression. This occurs via ERK- and Sp1-signalling pathways.
Asunto(s)
Araquidonato 12-Lipooxigenasa/metabolismo , Mucina 5AC/biosíntesis , Mucosa Respiratoria/metabolismo , Ácido 12-Hidroxi-5,8,10,14-Eicosatetraenoico/metabolismo , Araquidonato 12-Lipooxigenasa/genética , Carcinógenos/farmacología , Línea Celular , Inhibidores de la Ciclooxigenasa/farmacología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/metabolismo , Silenciador del Gen , Humanos , Interleucina-8/biosíntesis , Lipopolisacáridos/farmacología , Inhibidores de la Lipooxigenasa/farmacología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Mucina 2/biosíntesis , Mucina 5B/biosíntesis , Moco/metabolismo , Transporte de Proteínas , Mucosa Respiratoria/efectos de los fármacos , Factor de Transcripción Sp1/metabolismo , Acetato de Tetradecanoilforbol/farmacología , Factor de Crecimiento Transformador alfa/farmacologíaRESUMEN
Surfactant protein A (SP-A), a member of the collectin family originally described as a major component of lung surfactant, plays an important role in the modulation of lung host defense. A new interest in SP-A is provided by the link between fetal lung development and the timing of labor in the mouse. In the present review, we discuss some of the known features of SP-A such as biological functions, signaling pathways involved and the recent developments showing that SP-A bind and serve as a signal in the female genital tract. Therefore, such reports support a new paradigm involving SP-A as a multifunctional protein in the parturition process.