RESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly become a global pandemic. In addition to the acute pulmonary symptoms of coronavirus disease (COVID-19) (the disease associated with SARS-CoV-2 infection), pulmonary and distal coagulopathies have caused morbidity and mortality in many patients. Currently, the molecular pathogenesis underlying COVID-19-associated coagulopathies are unknown. Identifying the molecular basis of how SARS-CoV-2 drives coagulation is essential to mitigating short- and long-term thrombotic risks of sick and recovered patients with COVID-19. We aimed to perform coagulation-focused transcriptome analysis of in vitro infected primary respiratory epithelial cells, patient-derived bronchial alveolar lavage cells, and circulating immune cells during SARS-CoV-2 infection. Our objective was to identify transcription-mediated signaling networks driving coagulopathies associated with COVID-19. We analyzed recently published experimentally and clinically derived bulk or single-cell RNA sequencing datasets of SARS-CoV-2 infection to identify changes in transcriptional regulation of blood coagulation. We also confirmed that the transcriptional expression of a key coagulation regulator was recapitulated at the protein level. We specifically focused our analysis on lung tissue-expressed genes regulating the extrinsic coagulation cascade and the plasminogen activation system. Analyzing transcriptomic data of in vitro infected normal human bronchial epithelial cells and patient-derived bronchial alveolar lavage samples revealed that SARS-CoV-2 infection induces the extrinsic blood coagulation cascade and suppresses the plasminogen activation system. We also performed in vitro SARS-CoV-2 infection experiments on primary human lung epithelial cells to confirm that transcriptional upregulation of tissue factor, the extrinsic coagulation cascade master regulator, manifested at the protein level. Furthermore, infection of normal human bronchial epithelial cells with influenza A virus did not drive key regulators of blood coagulation in a similar manner as SARS-CoV-2. In addition, peripheral blood mononuclear cells did not differentially express genes regulating the extrinsic coagulation cascade or plasminogen activation system during SARS-CoV-2 infection, suggesting that they are not directly inducing coagulopathy through these pathways. The hyperactivation of the extrinsic blood coagulation cascade and the suppression of the plasminogen activation system in SARS-CoV-2-infected epithelial cells may drive diverse coagulopathies in the lung and distal organ systems. Understanding how hosts drive such transcriptional changes with SARS-CoV-2 infection may enable the design of host-directed therapeutic strategies to treat COVID-19 and other coronaviruses inducing hypercoagulation.
Asunto(s)
Células Epiteliales Alveolares/metabolismo , Trastornos de la Coagulación Sanguínea/metabolismo , COVID-19/metabolismo , Regulación de la Expresión Génica , SARS-CoV-2/metabolismo , Transducción de Señal , Transcripción Genética , Células Epiteliales Alveolares/patología , Células Epiteliales Alveolares/virología , Trastornos de la Coagulación Sanguínea/etiología , Trastornos de la Coagulación Sanguínea/patología , COVID-19/complicaciones , COVID-19/patología , Línea Celular , Femenino , Humanos , Virus de la Influenza A/metabolismo , Gripe Humana/complicaciones , Gripe Humana/metabolismo , Gripe Humana/patología , MasculinoRESUMEN
Electronic nicotine delivery system (ENDS) use is outpacing our understanding of its potential harmful effects. Homeostasis of the lung is maintained through proper balance of cell death, efferocytic clearance, and phagocytosis of pathogens. To investigate whether ENDS use has the potential to alter this balance, we developed physiologically relevant ENDS exposure paradigms for lung epithelial cells and primary macrophages. In our studies, cells were exposed directly to aerosol made from carefully controlled components with and without nicotine. We found that ENDS aerosol exposure led to apoptosis, secondary necrosis, and necrosis in lung epithelial cell models. In contrast, macrophages died mostly by apoptosis and inflammatory caspase-mediated cell death when exposed to ENDS aerosol. The clearance of dead cells and pathogens by efferocytosis and phagocytosis, respectively, is an important process in maintaining a healthy lung. To investigate the impact of ENDS aerosol on macrophage function independent of general toxicity, we used an exposure time that did not induce cell death in primary macrophages. Exposure to ENDS aerosol containing nicotine inhibited nearly all phagocytic and greatly reduced the efferocytic abilities of primary macrophages. When challenged with a bacterial pathogen, there was decreased bacterial clearance. The presence of nicotine in the ENDS aerosol increased its toxicity and functional impact; however, nicotine exposure alone did not have any deleterious effects. These data demonstrate that ENDS aerosol exposure could lead to increased epithelial cell and macrophage death in the lung and impair important macrophage functions that are essential for maintenance of lung function.
Asunto(s)
Muerte Celular/efectos de los fármacos , Sistemas Electrónicos de Liberación de Nicotina , Células Epiteliales/efectos de los fármacos , Macrófagos/efectos de los fármacos , Aerosoles/farmacología , Apoptosis/efectos de los fármacos , Línea Celular , Humanos , Pulmón/efectos de los fármacos , Nicotina/farmacología , Fagocitosis/efectos de los fármacosRESUMEN
The innate immune system is responsible for many important functions in the body including responding to infection, clearing cancerous cells, healing wounds, and removing foreign substances. Although many of these functions happen simultaneously in life, most laboratory studies of the innate immune response focus on one activity. How the innate immune system responds to concurrent insults in different parts of the body is not well understood. This study explores the impact of a lung infection on the cutaneous wound healing process. We used two complimentary models of injury: the excisional tail wound and subcutaneous implantation of polyvinyl alcohol (PVA) sponges. These models allow for assessment of the rate of closure and measurement of cellular and cytokine responses during acute wound healing, respectively. When mice with these healing wounds were infected with influenza A virus (IAV) in the lung there was a delay in wound healing. The viral lung infection suppressed the innate immune response in a healing wound, including cellular infiltrate, chemokines, growth factors, and cytokines. However, there was not a global immune suppression as there was an increase in inflammation systemically in mice with both infection and healing wounds compared to mice with only wounds or IAV infection. In addition, the lung immune response was largely unaffected indicating that responding to a lung infection is prioritized over a healing wound. This study introduces the concept of immune triage, in that when faced with multiple insults the immune system prioritizes responses. This paradigm likely applies to many situations that involve the innate immune system, and understanding how the innate immune system handles multiple insults is essential to understanding how it can efficiently clear pathogens while responding to other inflammatory events.
Asunto(s)
Tolerancia Inmunológica , Pulmón/virología , Infecciones por Orthomyxoviridae/inmunología , Infecciones del Sistema Respiratorio/inmunología , Piel/inmunología , Piel/lesiones , Animales , Inmunidad Innata/fisiología , Virus de la Influenza A/inmunología , Virus de la Influenza A/patogenicidad , Masculino , Ratones , Ratones Endogámicos C57BL , Infecciones por Orthomyxoviridae/virología , Infecciones del Sistema Respiratorio/patología , Infecciones del Sistema Respiratorio/virología , Piel/virología , Cicatrización de Heridas/fisiologíaRESUMEN
Tumour metastasis is the primary cause of mortality in cancer patients and remains the key challenge for cancer therapy. New therapeutic approaches to block inhibitory pathways of the immune system have renewed hopes for the utility of such therapies. Here we show that genetic deletion of the E3 ubiquitin ligase Cbl-b (casitas B-lineage lymphoma-b) or targeted inactivation of its E3 ligase activity licenses natural killer (NK) cells to spontaneously reject metastatic tumours. The TAM tyrosine kinase receptors Tyro3, Axl and Mer (also known as Mertk) were identified as ubiquitylation substrates for Cbl-b. Treatment of wild-type NK cells with a newly developed small molecule TAM kinase inhibitor conferred therapeutic potential, efficiently enhancing anti-metastatic NK cell activity in vivo. Oral or intraperitoneal administration using this TAM inhibitor markedly reduced murine mammary cancer and melanoma metastases dependent on NK cells. We further report that the anticoagulant warfarin exerts anti-metastatic activity in mice via Cbl-b/TAM receptors in NK cells, providing a molecular explanation for a 50-year-old puzzle in cancer biology. This novel TAM/Cbl-b inhibitory pathway shows that it might be possible to develop a 'pill' that awakens the innate immune system to kill cancer metastases.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/metabolismo , Células Asesinas Naturales/inmunología , Neoplasias Mamarias Experimentales/patología , Melanoma Experimental/patología , Metástasis de la Neoplasia/inmunología , Proteínas Proto-Oncogénicas c-cbl/metabolismo , Proteínas Tirosina Quinasas Receptoras/metabolismo , Ubiquitina-Proteína Ligasas/metabolismo , Proteínas Adaptadoras Transductoras de Señales/deficiencia , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Anticoagulantes/farmacología , Anticoagulantes/uso terapéutico , Femenino , Células Asesinas Naturales/efectos de los fármacos , Células Asesinas Naturales/metabolismo , Masculino , Neoplasias Mamarias Experimentales/tratamiento farmacológico , Neoplasias Mamarias Experimentales/genética , Neoplasias Mamarias Experimentales/inmunología , Melanoma Experimental/tratamiento farmacológico , Melanoma Experimental/genética , Melanoma Experimental/inmunología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Metástasis de la Neoplasia/tratamiento farmacológico , Metástasis de la Neoplasia/prevención & control , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas c-cbl/deficiencia , Proteínas Proto-Oncogénicas c-cbl/genética , Proteínas Tirosina Quinasas Receptoras/antagonistas & inhibidores , Ubiquitina-Proteína Ligasas/deficiencia , Ubiquitina-Proteína Ligasas/genética , Ubiquitinación , Warfarina/farmacología , Warfarina/uso terapéutico , Tirosina Quinasa c-Mer , Tirosina Quinasa del Receptor AxlRESUMEN
[This corrects the article DOI: 10.1371/journal.ppat.1006032.].
RESUMEN
Pathogen clearance and host resilience/tolerance to infection are both important factors in surviving an infection. Cells of the myeloid lineage play important roles in both of these processes. Neutrophils, monocytes, macrophages, and dendritic cells all have important roles in initiation of the immune response and clearance of bacterial pathogens. If these cells are not properly regulated they can result in excessive inflammation and immunopathology leading to decreased host resilience. Programmed cell death (PCD) is one possible mechanism that myeloid cells may use to prevent excessive inflammation. Myeloid cell subsets play roles in tissue repair, immune response resolution, and maintenance of homeostasis, so excessive PCD may also influence host resilience in this way. In addition, myeloid cell death is one mechanism used to control pathogen replication and dissemination. Many of these functions for PCD have been well defined in vitro, but the role in vivo is less well understood. We created a mouse that constitutively expresses the pro-survival B-cell lymphoma (bcl)-2 protein in myeloid cells (CD68(bcl2tg), thus decreasing PCD specifically in myeloid cells. Using this mouse model we explored the impact that decreased cell death of these cells has on infection with two different bacterial pathogens, Legionella pneumophila and Streptococcus pyogenes. Both of these pathogens target multiple cell death pathways in myeloid cells, and the expression of bcl2 resulted in decreased PCD after infection. We examined both pathogen clearance and host resilience and found that myeloid cell death was crucial for host resilience. Surprisingly, the decreased myeloid PCD had minimal impact on pathogen clearance. These data indicate that the most important role of PCD during infection with these bacteria is to minimize inflammation and increase host resilience, not to aid in the clearance or prevent the spread of the pathogen.
Asunto(s)
Apoptosis/inmunología , Enfermedad de los Legionarios/inmunología , Células Mieloides/inmunología , Infecciones Estreptocócicas/inmunología , Animales , Citometría de Flujo , Inmunidad Innata , Legionella pneumophila/inmunología , Ratones , Ratones Transgénicos , Streptococcus pyogenes/inmunologíaRESUMEN
Neuroinflammation can lead to chronic maladaptive pain affecting millions of people worldwide. Neurotransmitters, cytokines, and ion channels are implicated in neuro-immune cell signaling but their roles in specific behavioral responses are not fully elucidated. Voltage-gated CaV2.2 channel activity in skin controls rapid and transient heat hypersensitivity induced by intradermal capsaicin via IL-1α cytokine signaling. CaV2.2 channels are not, however, involved in mechanical hypersensitivity that developed in the same animal model. Here, we show that CaV2.2 channels are also critical for heat hypersensitivity induced by the intradermal (id) Complete Freund's Adjuvant (CFA) model of chronic neuroinflammation that involves ongoing cytokine signaling for days. Ongoing CFA-induced cytokine signaling cascades in skin lead to pronounced edema, and hypersensitivity to sensory stimuli. Peripheral CaV2.2 channel activity in skin is required for the full development and week-long time course of heat hypersensitivity induced by id CFA. CaV2.2 channels, by contrast, are not involved in paw edema and mechanical hypersensitivity. CFA induced increases in cytokines in hind paws including IL-6 which was dependent on CaV2.2 channel activity. Using IL-6 specific neutralizing antibodies, we show that IL-6 contributes to heat hypersensitivity and, neutralizing both IL-1α and IL-6 was even more effective at reducing the magnitude and duration of CFA-induced heat hypersensitivity. Our findings demonstrate a functional link between CaV2.2 channel activity and the release of IL-6 in skin and show that CaV2.2 channels have a privileged role in the induction and maintenance of heat hypersensitivity during chronic forms of neuroinflammation in skin.
RESUMEN
Neurons have the unique capacity to adapt output in response to changes in their environment. Within seconds, sensory nerve endings can become hypersensitive to stimuli in response to potentially damaging events. The underlying behavioral response is well studied, but several of the key signaling molecules that mediate sensory hypersensitivity remain unknown. We previously discovered that peripheral voltage-gated CaV2.2 channels in nerve endings in skin are essential for the rapid, transient increase in sensitivity to heat, but not to mechanical stimuli, that accompanies intradermal capsaicin. Here we report that the cytokine interleukin-1α (IL-1α), an alarmin, is necessary and sufficient to trigger rapid heat and mechanical hypersensitivity in skin. Of 20 cytokines screened, only IL-1α was consistently detected in hind paw interstitial fluid in response to intradermal capsaicin and, similar to behavioral sensitivity to heat, IL-1α levels were also dependent on peripheral CaV2.2 channel activity. Neutralizing IL-1α in skin significantly reduced capsaicin-induced changes in hind paw sensitivity to radiant heat and mechanical stimulation. Intradermal IL-1α enhances behavioral responses to stimuli and, in culture, IL-1α enhances the responsiveness of Trpv1-expressing sensory neurons. Together, our data suggest that IL-1α is the key cytokine that underlies rapid and reversible neuroinflammatory responses in skin.
RESUMEN
Neurons have the unique capacity to adapt output in response to changes in their environment. Within seconds, sensory nerve endings can become hypersensitive to stimuli in response to potentially damaging events. The underlying behavioral response is well studied, but several of the key signaling molecules that mediate sensory hypersensitivity remain unknown. We previously discovered that peripheral voltage-gated CaV2.2 channels in nerve endings in skin are essential for the rapid, transient increase in sensitivity to heat, but not to mechanical stimuli, that accompanies intradermal capsaicin. Here we report that the cytokine interleukin-1α (IL-1α), an alarmin, is necessary and sufficient to trigger rapid heat and mechanical hypersensitivity in skin. Of 20 cytokines screened, only IL-1α was consistently detected in hind paw interstitial fluid in response to intradermal capsaicin and, similar to behavioral sensitivity to heat, IL-1α levels were also dependent on peripheral CaV2.2 channel activity. Neutralizing IL-1α in skin significantly reduced capsaicin-induced changes in hind paw sensitivity to radiant heat and mechanical stimulation. Intradermal IL-1α enhances behavioral responses to stimuli and, in culture, IL-1α enhances the responsiveness of Trpv1-expressing sensory neurons. Together, our data suggest that IL-1α is the key cytokine that underlies rapid and reversible neuroinflammatory responses in skin.
Asunto(s)
Calor , Interleucina-1alfa , Animales , Ratones , Capsaicina/farmacología , Interleucina-1alfa/metabolismo , Células Receptoras Sensoriales , Piel , Canales de Calcio Tipo N/metabolismoRESUMEN
The lung is a complex and unique organ system whose biology is strongly influenced by environmental exposure, oxygen abundance, connection to extrapulmonary systems via a dense capillary network, and an array of immune cells that reside in the tissue at steady state. The lung also harbors a low biomass community of commensal microorganisms that are dynamic during both health and disease with the capacity to modulate regulatory immune responses during diseases such as cancer. Lung cancer is the third most common cancer worldwide with the highest mortality rate amongst cancers due to the difficulty of an early diagnosis. This review discusses the current body of work addressing the interactions between the lung microbiota and the immune system, and how these two components of the pulmonary system are linked to lung cancer development and outcomes. Bringing in lessons from broader studies examining the effects of the gut microbiota on cancer outcomes, we highlight many challenges and gaps in this nascent field.
Asunto(s)
Microbioma Gastrointestinal , Neoplasias Pulmonares , Microbiota , Humanos , Pulmón , Mucosa IntestinalRESUMEN
Mast et al. analyzed transcriptome data derived from RNA-sequencing (RNA-seq) of COVID-19 patient bronchoalveolar lavage fluid (BALF) samples, as compared to BALF RNA-seq samples from a study investigating microbiome and inflammatory interactions in obese and asthmatic adults (Mast et al., 2021). Based on their analysis of these data, Mast et al. concluded that mRNA expression of key regulators of the extrinsic coagulation cascade and fibrinolysis were significantly reduced in COVID-19 patients. Notably, they reported that the expression of the extrinsic coagulation cascade master regulator Tissue Factor (F3) remained unchanged, while there was an 8-fold upregulation of its cognate inhibitor Tissue Factor Pathway Inhibitor (TFPI). From this they conclude that "pulmonary fibrin deposition does not stem from enhanced local [tissue factor] production and that counterintuitively, COVID-19 may dampen [tissue factor]-dependent mechanisms in the lungs". They also reported decreased Activated Protein C (aPC) mediated anticoagulant activity and major increases in fibrinogen expression and other key regulators of clot formation. Many of these results are contradictory to findings in most of the field, particularly the findings regarding extrinsic coagulation cascade mediated coagulopathies. Here, we present a complete re-analysis of the data sets analyzed by Mast et al. This re-analysis demonstrates that the two data sets utilized were not comparable between one another, and that the COVID-19 sample set was not suitable for the transcriptomic analysis Mast et al. performed. We also identified other significant flaws in the design of their retrospective analysis, such as poor-quality control and filtering standards. Given the issues with the datasets and analysis, their conclusions are not supported.
Asunto(s)
COVID-19 , SARS-CoV-2 , Anticoagulantes , Humanos , Pulmón , Estudios Retrospectivos , TranscriptomaRESUMEN
Dendritic cells (DCs) are specialized phagocytes that internalize exogenous antigens and microbes at peripheral sites, and then migrate to lymphatic organs to display foreign peptides to naïve T cells. There are several examples where DCs have been shown to be more efficient at restricting the intracellular replication of pathogens compared to macrophages, a property that could prevent DCs from enhancing pathogen dissemination. To understand DC responses to pathogens, we investigated the mechanisms by which mouse DCs are able to restrict replication of the intracellular pathogen Legionella pneumophila. We show that both DCs and macrophages have the ability to interfere with L. pneumophila replication through a cell death pathway mediated by caspase-1 and Naip5. L. pneumophila that avoided Naip5-dependent responses, however, showed robust replication in macrophages but remained unable to replicate in DCs. Apoptotic cell death mediated by caspase-3 was found to occur much earlier in DCs following infection by L. pneumophila compared to macrophages infected similarly. Eliminating the pro-apoptotic proteins Bax and Bak or overproducing the anti-apoptotic protein Bcl-2 were both found to restore L. pneumophila replication in DCs. Thus, DCs have a microbial response pathway that rapidly activates apoptosis to limit pathogen replication.
Asunto(s)
Apoptosis , Células Dendríticas/microbiología , Células Dendríticas/fisiología , Legionella pneumophila/crecimiento & desarrollo , Enfermedad de los Legionarios/inmunología , Animales , Caspasa 1/metabolismo , Caspasa 3/metabolismo , Fragmentación del ADN , Células Dendríticas/inmunología , Enfermedad de los Legionarios/microbiología , Macrófagos/inmunología , Macrófagos/microbiología , Ratones , Ratones Endogámicos C57BL , Microscopía Fluorescente , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Transducción de Señal , Proteína Destructora del Antagonista Homólogo bcl-2/metabolismo , Proteína X Asociada a bcl-2/metabolismoRESUMEN
Few studies have demonstrated that innate lymphocytes play a major role in preventing spontaneous tumor formation. We evaluated the development of spontaneous tumors in mice lacking beta-2 microglobulin (beta2m; and thus MHC class I, CD1d, and CD16) and/or perforin, since these tumor cells would be expected to activate innate effector cells. Approximately half the cohort of perforin gene-targeted mice succumbed to spontaneous disseminated B cell lymphomas and in mice that also lacked beta2m, the lymphomas developed earlier (by more than 100 d) and with greater incidence (84%). B cell lymphomas from perforin/beta2m gene-targeted mice effectively primed cell-mediated cytotoxicity and perforin, but not IFN-gamma, IL-12, or IL-18, was absolutely essential for tumor rejection. Activated NK1.1+ and gammadeltaTCR+ T cells were abundant at the tumor site, and transplanted tumors were strongly rejected by either, or both, of these cell types. Blockade of a number of different known costimulatory pathways failed to prevent tumor rejection. These results reflect a critical role for NK cells and gammadeltaTCR+ T cells in innate immune surveillance of B cell lymphomas, mediated by as yet undetermined pathway(s) of tumor recognition.
Asunto(s)
Citotoxicidad Inmunológica/inmunología , Inmunidad Innata/inmunología , Células Asesinas Naturales/inmunología , Linfoma de Células B/inmunología , Linfocitos T/inmunología , Animales , Citometría de Flujo , Rechazo de Injerto/inmunología , Linfoma de Células B/genética , Glicoproteínas de Membrana/deficiencia , Glicoproteínas de Membrana/genética , Ratones , Ratones Endogámicos C57BL , Trasplante de Neoplasias , Neoplasias Experimentales/genética , Neoplasias Experimentales/inmunología , Perforina , Peritoneo/inmunología , Proteínas Citotóxicas Formadoras de Poros , Microglobulina beta-2/deficienciaRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has rapidly become a global pandemic. In addition to the acute pulmonary symptoms of COVID-19 (the disease associated with SARS-CoV-2 infection), pulmonary and distal coagulopathies have caused morbidity and mortality in many patients. Currently, the molecular pathogenesis underlying COVID-19 associated coagulopathies are unknown. While there are many theories for the cause of this pathology, including hyper inflammation and excess tissue damage, the cellular and molecular underpinnings are not yet clear. By analyzing transcriptomic data sets from experimental and clinical research teams, we determined that changes in the gene expression of genes important in the extrinsic coagulation cascade in the lung epithelium may be important triggers for COVID-19 coagulopathy. This regulation of the extrinsic blood coagulation cascade is not seen with influenza A virus (IAV)-infected NHBEs suggesting that the lung epithelial derived coagulopathies are specific to SARS-Cov-2 infection. This study is the first to identify potential lung epithelial cell derived factors contributing to COVID-19 associated coagulopathy.
RESUMEN
In the context of pulmonary infection, both hosts and pathogens have evolved a multitude of mechanisms to regulate the process of host cell death. The host aims to rapidly induce an inflammatory response at the site of infection, promote pathogen clearance, quickly resolve inflammation, and return to tissue homeostasis. The appropriate modulation of cell death in respiratory epithelial cells and pulmonary immune cells is central in the execution of all these processes. Cell death can be either inflammatory or anti-inflammatory depending on regulated cell death (RCD) modality triggered and the infection context. In addition, diverse bacterial pathogens have evolved many means to manipulate host cell death to increase bacterial survival and spread. The multitude of ways that hosts and bacteria engage in a molecular tug of war to modulate cell death dynamics during infection emphasizes its relevance in host responses and pathogen virulence at the host pathogen interface. This narrative review outlines several current lines of research characterizing bacterial pathogen manipulation of host cell death pathways in the lung. We postulate that understanding these interactions and the dynamics of intracellular and extracellular bacteria RCD manipulation, may lead to novel therapeutic approaches for the treatment of intractable respiratory infections.
Asunto(s)
Comunicación Celular/inmunología , Muerte Celular/inmunología , Bacterias Gramnegativas/patogenicidad , Bacterias Grampositivas/patogenicidad , Interacciones Huésped-Patógeno/inmunología , Neumonía Bacteriana/inmunología , Animales , Humanos , Inmunidad Innata , Macrófagos/inmunología , Neutrófilos/inmunología , Neumonía Bacteriana/microbiología , Mucosa Respiratoria/inmunología , VirulenciaRESUMEN
Wound healing is a complex process that requires the orderly progression of inflammation, granulation tissue formation, fibrosis, and resolution. Murine models provide valuable mechanistic insight into these processes; however, no single model fully addresses all aspects of the wound healing response. Instead, it is ideal to use multiple models to address the different aspects of wound healing. Here, two different methods that address diverse aspects of the wound healing response are described. In the first model, polyvinyl alcohol sponges are subcutaneously implanted along the mouse dorsum. Following sponge retrieval, cells can be isolated by mechanical disruption, and fluids can be extracted by centrifugation, thus allowing for a detailed characterization of cellular and cytokine responses in the acute wound environment. A limitation of this model is the inability to assess the rate of wound closure. For this, a tail skin excision model is utilized. In this model, a 10 mm x 3 mm rectangular piece of tail skin is excised along the dorsal surface, near the base of the tail. This model can be easily photographed for planimetric analysis to determine healing rates and can be excised for histological analysis. Both described methods can be utilized in genetically altered mouse strains, or in conjunction with models of comorbid conditions, such as diabetes, aging, or secondary infection, in order to elucidate wound healing mechanisms.
Asunto(s)
Vendajes , Alcohol Polivinílico/farmacología , Prótesis e Implantes , Piel/patología , Tejido Subcutáneo/patología , Cola (estructura animal)/patología , Cicatrización de Heridas , Enfermedad Aguda , Animales , Separación Celular , Modelos Animales de Enfermedad , Inflamación/patología , Masculino , Ratones Endogámicos C57BL , Tejido Subcutáneo/efectos de los fármacos , Cicatrización de Heridas/efectos de los fármacosRESUMEN
Neutrophil recruitment into the lung airspaces plays an important role in the containment and clearance of bacteria. Hemorrhagic shock, a complication of traumatic injury, induces immune dysfunction that compromises host defense and frequently leads to secondary infection. The objective of the current study was to determine whether prior hemorrhage impacts neutrophil recruitment in response to secondary Pseudomonas aeruginosa. Experiments were performed using a mouse model (C57BL/6) of respiratory infection by P. aeruginosa (strain PA103, 3â×â10 colony-forming units [CFUs]) that is delivered by intratracheal inhalation 24âh after hypovolemic hemorrhagic shock (fixed mean arterial blood pressure at 35âmmHg for 90âmin, Ringer's lactate infused as fluid resuscitation). By postmortem flow cytometry analyses of bronchoalveolar lavage fluid, we observe that prior hemorrhage attenuates the entry of neutrophils into the lung airspaces in response to P. aeruginosa. The reduction in neutrophil recruitment occurs in an amplified inflammatory environment, with elevated lung tissue levels of interleukin 6 and C-X-C motif ligand 1 in mice receiving hemorrhage prior to infection. As compared to either insult alone, outcome to sequential hemorrhage and respiratory infection includes enhanced mortality. The effect of prior hemorrhage on clearance of P. aeruginosa, as determined by quantifying bacterial CFUs in lung tissue, was not statistically significant at 24âh postinfection, but our data suggest that further inquiry may be needed to fully understand the potential impact of hemorrhagic shock on this process. These results suggest that changes in neutrophil recruitment may contribute to the immune dysfunction following hemorrhagic shock that renders the host susceptible to severe respiratory infection.
Asunto(s)
Hemorragia , Neutrófilos , Infecciones por Pseudomonas , Pseudomonas aeruginosa/inmunología , Infecciones del Sistema Respiratorio , Animales , Quimiocina CXCL1/inmunología , Hemorragia/complicaciones , Hemorragia/inmunología , Hemorragia/patología , Interleucina-6/inmunología , Pulmón/inmunología , Pulmón/microbiología , Pulmón/patología , Ratones , Neutrófilos/inmunología , Neutrófilos/patología , Infecciones por Pseudomonas/etiología , Infecciones por Pseudomonas/inmunología , Infecciones por Pseudomonas/patología , Infecciones del Sistema Respiratorio/etiología , Infecciones del Sistema Respiratorio/inmunología , Infecciones del Sistema Respiratorio/patologíaRESUMEN
Much research on infectious diseases focuses on clearing the pathogen through the use of antimicrobial drugs, the immune response, or a combination of both. Rapid clearance of pathogens allows for a quick return to a healthy state and increased survival. Pathogen-targeted approaches to combating infection have inherent limitations, including their pathogen-specific nature, the potential for antimicrobial resistance, and poor vaccine efficacy, among others. Another way to survive an infection is to tolerate the alterations to homeostasis that occur during a disease state through a process called host tolerance or resilience, which is independent from pathogen burden. Alterations in homeostasis during infection are numerous and include tissue damage, increased inflammation, metabolic changes, temperature changes, and changes in respiration. Given its importance and sensitivity, the lung is a good system for understanding host tolerance to infectious disease. Pneumonia is the leading cause of death for children under five worldwide. One reason for this is because when the pulmonary system is altered dramatically it greatly impacts the overall health and survival of a patient. Targeting host pathways involved in maintenance of pulmonary host tolerance during infection could provide an alternative therapeutic avenue that may be broadly applicable across a variety of pathologies. In this review, we will summarize recent findings on tolerance to host lung infection. We will focus on the involvement of innate immune responses in tolerance and how an initial viral lung infection may alter tolerance mechanisms in leukocytic, epithelial, and endothelial compartments to a subsequent bacterial infection. By understanding tolerance mechanisms in the lung we can better address treatment options for deadly pulmonary infections.
RESUMEN
Pneumonia is a world health problem and a leading cause of death, particularly affecting children and the elderly (1, 2). Bacterial pneumonia following infection with influenza A virus (IAV) is associated with increased morbidity and mortality but the mechanisms behind this phenomenon are not yet well-defined (3). Host resistance and tolerance are two processes essential for host survival during infection. Resistance is the host's ability to clear a pathogen while tolerance is the host's ability to overcome the impact of the pathogen as well as the host response to infection (4-8). Some studies have shown that IAV infection suppresses the immune response, leading to overwhelming bacterial loads (9-13). Other studies have shown that some IAV/bacterial coinfections cause alterations in tolerance mechanisms such as tissue resilience (14-16). In a recent analysis of nasopharyngeal swabs from patients hospitalized during the 2013-2014 influenza season, we have found that a significant proportion of IAV-infected patients were also colonized with Klebsiella oxytoca, a gram-negative bacteria known to be an opportunistic pathogen in a variety of diseases (17). Mice that were infected with K. oxytoca following IAV infection demonstrated decreased survival and significant weight loss when compared to mice infected with either single pathogen. Using this model, we found that IAV/K. oxytoca coinfection of the lung is characterized by an exaggerated inflammatory immune response. We observed early inflammatory cytokine and chemokine production, which in turn resulted in massive infiltration of neutrophils and inflammatory monocytes. Despite this swift response, the pulmonary pathogen burden in coinfected mice was similar to singly-infected animals, albeit with a slight delay in bacterial clearance. In addition, during coinfection we observed a shift in pulmonary macrophages toward an inflammatory and away from a tissue reparative phenotype. Interestingly, there was only a small increase in tissue damage in coinfected lungs as compared to either single infection. Our results indicate that during pulmonary coinfection a combination of seemingly modest defects in both host resistance and tolerance may act synergistically to cause worsened outcomes for the host. Given the prevalence of K. oxytoca detected in human IAV patients, these dysfunctional tolerance and resistance mechanisms may play an important role in the response of patients to IAV.
Asunto(s)
Coinfección , Interacciones Huésped-Patógeno , Virus de la Influenza A , Gripe Humana/microbiología , Infecciones por Klebsiella/microbiología , Klebsiella oxytoca , Animales , Modelos Animales de Enfermedad , Resistencia a la Enfermedad/inmunología , Interacciones Huésped-Patógeno/inmunología , Humanos , Tolerancia Inmunológica , Inmunidad Innata , Gripe Humana/inmunología , Infecciones por Klebsiella/inmunología , Leucocitos/inmunología , Leucocitos/metabolismo , Ratones , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/virología , Neumonía Bacteriana/inmunología , Neumonía Bacteriana/microbiología , Neumonía Viral/inmunología , Neumonía Viral/virologíaRESUMEN
Recently, two coronaviruses, severe acute respiratory syndrome coronavirus and Middle East respiratory syndrome coronavirus, have emerged to cause unusually severe respiratory disease in humans. Currently, there is a lack of effective antiviral treatment options or vaccine available. Given the severity of these outbreaks, and the possibility of additional zoonotic coronaviruses emerging in the near future, the exploration of different treatment strategies is necessary. Disease resilience is the ability of a given host to tolerate an infection, and to return to a state of health. This review focuses on exploring various host resilience mechanisms that could be exploited for treatment of severe acute respiratory syndrome coronavirus, Middle East respiratory syndrome coronavirus and other respiratory viruses that cause acute lung injury and acute respiratory distress syndrome.