RESUMEN
Acidic mammalian chitinase (CHIA) belongs to the 18-glycosidase family and is expressed in epithelial cells and certain immune cells (such as neutrophils and macrophages) in various organs. Under physiological conditions, as a hydrolase, CHIA can degrade chitin-containing pathogens, participate in Type 2 helper T (Th2)-mediated inflammation, and enhance innate and adaptive immunity to pathogen invasion. Under pathological conditions, such as rhinitis, ocular conjunctivitis, asthma, chronic atrophic gastritis, type 2 diabetes, and pulmonary interstitial fibrosis, CHIA expression is significantly changed. In addition, studies have shown that CHIA has an anti-apoptotic effect, promotes epithelial cell proliferation and maintains organ integrity, and these effects are not related to chitinase degradation. CHIA can also be used as a biomolecular marker in diseases such as chronic atrophic gastritis, dry eye, and acute kidney damage caused by sepsis. Analysis of the authoritative TCGA database shows that CHIA expression in gastric adenocarcinoma, liver cancer, renal clear cell carcinoma and other tumors is significantly downregulated compared with that in normal tissues, but the specific mechanism is unclear. This review is based on all surveys conducted to date and summarizes the expression patterns and functional diversity of CHIA in various organs. Understanding the physiological and pathophysiological relevance of CHIA in multiple organs opens new possibilities for disease treatment.
Asunto(s)
Encéfalo/enzimología , Quitinasas/metabolismo , Sistema Digestivo/enzimología , Ojo/enzimología , Riñón/enzimología , Sistema Respiratorio/enzimología , Animales , Encéfalo/fisiopatología , Diabetes Mellitus Tipo 2/enzimología , Diabetes Mellitus Tipo 2/fisiopatología , Sistema Digestivo/fisiopatología , Ojo/fisiopatología , Humanos , Riñón/fisiopatología , Sistema Respiratorio/fisiopatología , Transducción de SeñalRESUMEN
Pandemic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes coronavirus 19 disease (COVID-19) which presents a large spectrum of manifestations with fatal outcomes in vulnerable people over 70-years-old and with hypertension, diabetes, obesity, cardiovascular disease, COPD, and smoking status. Knowledge of the entry receptor is key to understand SARS-CoV-2 tropism, transmission and pathogenesis. Early evidence pointed to angiotensin-converting enzyme 2 (ACE2) as SARS-CoV-2 entry receptor. Here, we provide a critical summary of the current knowledge highlighting the limitations and remaining gaps that need to be addressed to fully characterize ACE2 function in SARS-CoV-2 infection and associated pathogenesis. We also discuss ACE2 expression and potential role in the context of comorbidities associated with poor COVID-19 outcomes. Finally, we discuss the potential co-receptors/attachment factors such as neuropilins, heparan sulfate and sialic acids and the putative alternative receptors, such as CD147 and GRP78.
Asunto(s)
Betacoronavirus/fisiología , Infecciones por Coronavirus/virología , Peptidil-Dipeptidasa A/fisiología , Neumonía Viral/virología , Acoplamiento Viral , Enzima Convertidora de Angiotensina 2 , Basigina/fisiología , COVID-19 , Comorbilidad , Infecciones por Coronavirus/epidemiología , Chaperón BiP del Retículo Endoplásmico , Regulación Enzimológica de la Expresión Génica , Heparitina Sulfato/fisiología , Humanos , Hipertensión/epidemiología , Hipertensión/fisiopatología , Neuropilina-1/fisiología , Oligopéptidos/fisiología , Especificidad de Órganos , Pandemias , Neumonía Viral/epidemiología , Unión Proteica , ARN Mensajero/biosíntesis , ARN Mensajero/genética , Receptores Virales , Sistema Renina-Angiotensina/fisiología , Sistema Respiratorio/enzimología , SARS-CoV-2 , Ácidos Siálicos/fisiología , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/fisiología , Internalización del VirusRESUMEN
The incidence, severity, and mortality of ongoing coronavirus infectious disease 19 (COVID-19) is greater in men compared with women, but the underlying factors contributing to this sex difference are still being explored. In the current study, using primary isolated human airway smooth muscle (ASM) cells from normal males versus females as a model, we explored the effect of estrogen versus testosterone in modulating the expression of angiotensin converting enzyme 2 (ACE2), a cell entry point for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Using confocal imaging, we found that ACE2 is expressed in human ASM. Furthermore, Western analysis of ASM cell lysates showed significantly lower ACE2 expression in females compared with males at baseline. In addition, ASM cells exposed to estrogen and testosterone for 24 h showed that testosterone significantly upregulates ACE2 expression in both males and females, whereas estrogen downregulates ACE2, albeit not significant compared with vehicle. These intrinsic and sex steroids induced differences may help explain sex differences in COVID-19.
Asunto(s)
Infecciones por Coronavirus/metabolismo , Peptidil-Dipeptidasa A/biosíntesis , Neumonía Viral/metabolismo , Sistema Respiratorio/metabolismo , Adulto , Anciano , Enzima Convertidora de Angiotensina 2 , COVID-19 , Células Cultivadas , Infecciones por Coronavirus/enzimología , Estrógenos/metabolismo , Estrógenos/farmacología , Femenino , Humanos , Masculino , Persona de Mediana Edad , Miocitos del Músculo Liso/citología , Miocitos del Músculo Liso/efectos de los fármacos , Miocitos del Músculo Liso/enzimología , Miocitos del Músculo Liso/metabolismo , Pandemias , Peptidil-Dipeptidasa A/metabolismo , Neumonía Viral/enzimología , Sistema Respiratorio/citología , Sistema Respiratorio/efectos de los fármacos , Sistema Respiratorio/enzimología , Factores Sexuales , Testosterona/metabolismo , Testosterona/farmacologíaRESUMEN
Tight barriers are crucial for animals. Insect respiratory cells establish barriers through their extracellular matrices. These chitinous-matrices must be soft and flexible to provide ventilation, but also tight enough to allow oxygen flow and protection against dehydration, infections, and environmental stresses. However, genes that control soft, flexible chitin-matrices are poorly known. We investigated the genes of the chitinolytic glycosylhydrolase-family 18 in the tracheal system of Drosophila melanogaster. Our findings show that five chitinases and three chitinase-like genes organize the tracheal chitin-cuticles. Most of the chitinases degrade chitin from airway lumina to enable oxygen delivery. They further improve chitin-cuticles to enhance tube stability and integrity against stresses. Unexpectedly, some chitinases also support chitin assembly to expand the tube lumen properly. Moreover, Chitinase2 plays a decisive role in the chitin-cuticle formation that establishes taenidial folds to support tube stability. Chitinase2 is apically enriched on the surface of tracheal cells, where it controls the chitin-matrix architecture independently of other known cuticular proteins or chitinases. We suppose that the principle mechanisms of chitin-cuticle assembly and degradation require a set of critical glycosylhydrolases for flexible and not-flexible cuticles. The same glycosylhydrolases support thick laminar cuticle formation and are evolutionarily conserved among arthropods.
Asunto(s)
Quitinasas/genética , Proteínas de Drosophila/genética , Genes de Insecto/genética , Hidrolasas/genética , Sistema Respiratorio/enzimología , Animales , Quitina/metabolismo , Quitinasas/fisiología , Proteínas de Drosophila/metabolismo , Proteínas de Drosophila/fisiología , Drosophila melanogaster/anatomía & histología , Drosophila melanogaster/enzimología , Drosophila melanogaster/genética , Genes de Insecto/fisiología , Hidrolasas/metabolismo , Hidrolasas/fisiología , Oxígeno/metabolismo , Sistema Respiratorio/anatomía & histología , Tráquea/anatomía & histología , Tráquea/enzimologíaRESUMEN
Human respiratory syncytial virus (RSV) is the leading viral cause of acute pediatric lower respiratory tract infections worldwide, with no available vaccine or effective antiviral drug. To gain insight into virus-host interactions, we performed a genome-wide siRNA screen. The expression of over 20,000 cellular genes was individually knocked down in human airway epithelial A549 cells, followed by infection with RSV expressing green fluorescent protein (GFP). Knockdown of expression of the cellular ATP1A1 protein, which is the major subunit of the Na+,K+-ATPase of the plasma membrane, had one of the strongest inhibitory effects on GFP expression and viral titer. Inhibition was not observed for vesicular stomatitis virus, indicating that it was RSV-specific rather than a general effect. ATP1A1 formed clusters in the plasma membrane very early following RSV infection, which was independent of replication but dependent on the attachment glycoprotein G. RSV also triggered activation of ATP1A1, resulting in signaling by c-Src-kinase activity that transactivated epidermal growth factor receptor (EGFR) by Tyr845 phosphorylation. ATP1A1 signaling and activation of both c-Src and EGFR were found to be required for efficient RSV uptake. Signaling events downstream of EGFR culminated in the formation of macropinosomes. There was extensive uptake of RSV virions into macropinosomes at the beginning of infection, suggesting that this is a major route of RSV uptake, with fusion presumably occurring in the macropinosomes rather than at the plasma membrane. Important findings were validated in primary human small airway epithelial cells (HSAEC). In A549 cells and HSAEC, RSV uptake could be inhibited by the cardiotonic steroid ouabain and the digitoxigenin derivative PST2238 (rostafuroxin) that bind specifically to the ATP1A1 extracellular domain and block RSV-triggered EGFR Tyr845 phosphorylation. In conclusion, we identified ATP1A1 as a host protein essential for macropinocytic entry of RSV into respiratory epithelial cells, and identified PST2238 as a potential anti-RSV drug.
Asunto(s)
Pinocitosis , Infecciones por Virus Sincitial Respiratorio/complicaciones , Virus Sincitial Respiratorio Humano/patogenicidad , Infecciones del Sistema Respiratorio/prevención & control , ATPasa Intercambiadora de Sodio-Potasio/antagonistas & inhibidores , Proteínas Virales/metabolismo , Internalización del Virus , Células A549 , Cardiotónicos/farmacología , Digitoxigenina/química , Digitoxigenina/farmacología , Células Epiteliales/efectos de los fármacos , Células Epiteliales/enzimología , Células Epiteliales/virología , Receptores ErbB/metabolismo , Ensayos Analíticos de Alto Rendimiento , Humanos , Ouabaína/farmacología , ARN Interferente Pequeño/genética , Infecciones por Virus Sincitial Respiratorio/virología , Sistema Respiratorio/efectos de los fármacos , Sistema Respiratorio/enzimología , Sistema Respiratorio/virología , Infecciones del Sistema Respiratorio/epidemiología , Infecciones del Sistema Respiratorio/virología , Transducción de Señal , ATPasa Intercambiadora de Sodio-Potasio/genética , Proteínas Virales/genéticaRESUMEN
This study investigated the co-localization of the Middle East respiratory syndrome coronavirus (MERS-CoV) and its receptor dipeptidyl peptidase-4 (DPP4) by immunohistochemistry (IHC) across respiratory and lymphoid organs of experimentally MERS-CoV infected pigs and llamas. Also, scanning electron microscopy was performed to assess the ciliary integrity of respiratory epithelial cells in both species. In pigs, on day 2 post-inoculation (p.i.), DPP4-MERS-CoV co-localization was detected in medial turbinate epithelium. On day 4 p.i., the virus/receptor co-localized in frontal and medial turbinate epithelial cells in pigs, and epithelial cells distributed unevenly through the whole nasal cavity and in the cervical lymph node in llamas. MERS-CoV viral nucleocapsid was mainly detected in upper respiratory tract sites on days 2 and 4 p.i. in pigs and day 4 p.i. in llamas. No MERS-CoV was detected on day 24 p.i. in any tissue by IHC. While pigs showed severe ciliary loss in the nasal mucosa both on days 2 and 4 p.i. and moderate loss in the trachea on days 4 and 24 p.i., ciliation of respiratory organs in llamas was not significantly affected. Obtained data confirm the role of DPP4 for MERS-CoV entry in respiratory epithelial cells of llamas. Notably, several nasal epithelial cells in pigs were found to express viral antigen but not DPP4, suggesting the possible existence of other molecule/s facilitating virus entry or down regulation of DPP4 upon infection.
Asunto(s)
Camélidos del Nuevo Mundo/virología , Infecciones por Coronavirus/veterinaria , Dipeptidil Peptidasa 4/metabolismo , Tejido Linfoide/enzimología , Coronavirus del Síndrome Respiratorio de Oriente Medio/patogenicidad , Sistema Respiratorio/enzimología , Enfermedades de los Porcinos/virología , Animales , Infecciones por Coronavirus/virología , Inmunohistoquímica/veterinaria , Microscopía Electrónica de Rastreo/veterinaria , ARN Viral/genética , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Receptores Virales/metabolismo , PorcinosRESUMEN
Rhinovirus (RV) infection is involved in acute exacerbations of asthma and chronic obstructive pulmonary disease (COPD). RV primarily infects upper and lower airway epithelium. Immunoproteasomes (IP) are proteolytic machineries with multiple functions including the regulation of MHC class I antigen processing during viral infection. However, the role of IP in RV infection has not been explored. We sought to investigate the expression and function of IP during airway RV infection. Primary human tracheobronchial epithelial (HTBE) cells were cultured at air-liquid interface (ALI) and treated with RV16, RV1B, or interferon (IFN)-λ in the absence or presence of an IP inhibitor (ONX-0914). IP gene (i.e. LMP2) deficient mouse tracheal epithelial cells (mTECs) were cultured for the mechanistic studies. LMP2-deficient mouse model was used to define the in vivo role of IP in RV infection. IP subunits LMP2 and LMP7, antiviral genes MX1 and OAS1 and viral load were measured. Both RV16 and RV1B significantly increased the expression of LMP2 and LMP7 mRNA and proteins, and IFN-λ mRNA in HTBE cells. ONX-0914 down-regulated MX1 and OAS1, and increased RV16 load in HTBE cells. LMP2-deficient mTECs showed a significant increase in RV1B load compared with the wild-type (WT) cells. LMP2-deficient (compared with WT) mice increased viral load and neutrophils in bronchoalveolar lavage (BAL) fluid after 24 h of RV1B infection. Mechanistically, IFN-λ induction by RV infection contributed to LMP2 and LMP7 up-regulation in HTBE cells. Our data suggest that IP are induced during airway RV infection, which in turn may serve as an antiviral and anti-inflammatory mechanism.
Asunto(s)
Células Epiteliales/inmunología , Infecciones por Picornaviridae/inmunología , Complejo de la Endopetidasa Proteasomal/inmunología , Rhinovirus/inmunología , Animales , Asma/enzimología , Asma/inmunología , Asma/virología , Línea Celular , Cisteína Endopeptidasas/genética , Cisteína Endopeptidasas/inmunología , Cisteína Endopeptidasas/metabolismo , Células Epiteliales/enzimología , Células Epiteliales/virología , Interacciones Huésped-Patógeno/efectos de los fármacos , Interacciones Huésped-Patógeno/inmunología , Humanos , Ratones Noqueados , Oligopéptidos/farmacología , Infecciones por Picornaviridae/enzimología , Infecciones por Picornaviridae/virología , Complejo de la Endopetidasa Proteasomal/metabolismo , Inhibidores de Proteasoma/farmacología , Sistema Respiratorio/enzimología , Sistema Respiratorio/inmunología , Sistema Respiratorio/virología , Rhinovirus/fisiologíaRESUMEN
RATIONALE: In lung cancer, upregulation of the PI3K (phosphoinositide 3-kinase) pathway is an early event that contributes to cell proliferation, survival, and tissue invasion. Upregulation of this pathway was recently described as associated with enrichment of the lower airways with bacteria identified as oral commensals. OBJECTIVES: We hypothesize that host-microbe interactions in the lower airways of subjects with lung cancer affect known cancer pathways. METHODS: Airway brushings were collected prospectively from subjects with lung nodules at time of diagnostic bronchoscopy, including 39 subjects with final lung cancer diagnoses and 36 subjects with noncancer diagnoses. In addition, samples from 10 healthy control subjects were included. 16S ribosomal RNA gene amplicon sequencing and paired transcriptome sequencing were performed on all airway samples. In addition, an in vitro model with airway epithelial cells exposed to bacteria/bacterial products was performed. MEASUREMENTS AND MAIN RESULTS: The composition of the lower airway transcriptome in the patients with cancer was significantly different from the control subjects, which included up-regulation of ERK (extracellular signal-regulated kinase) and PI3K signaling pathways. The lower airways of patients with lung cancer were enriched for oral taxa (Streptococcus and Veillonella), which was associated with up-regulation of the ERK and PI3K signaling pathways. In vitro exposure of airway epithelial cells to Veillonella, Prevotella, and Streptococcus led to upregulation of these same signaling pathways. CONCLUSIONS: The data presented here show that several transcriptomic signatures previously identified as relevant to lung cancer pathogenesis are associated with enrichment of the lower airway microbiota with oral commensals.
Asunto(s)
Neoplasias Pulmonares/enzimología , Microbiota/fisiología , Fosfatidilinositol 3-Quinasas/metabolismo , Sistema Respiratorio/enzimología , Regulación hacia Arriba/fisiología , Adulto , Anciano , Broncoscopía , Estudios Transversales , Femenino , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/microbiología , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Sistema Respiratorio/metabolismo , Sistema Respiratorio/microbiologíaRESUMEN
Two cAMP signaling compartments centered on adenylyl cyclase (AC) exist in human airway smooth muscle (HASM) cells, one containing ß2-adrenergic receptor AC6 and another containing E prostanoid receptor AC2. We hypothesized that different PDE isozymes selectively regulate cAMP signaling in each compartment. According to RNA-sequencing data, 18 of 24 PDE genes were expressed in primary HASM cells derived from age- and sex-matched donors with and without asthma. PDE8A was the third most abundant of the cAMP-degrading PDE genes, after PDE4A and PDE1A. Knockdown of PDE8A using shRNA evoked twofold greater cAMP responses to 1 µM forskolin in the presence of 3-isobutyl-1-methylxanthine. Overexpression of AC2 did not alter this response, but overexpression of AC6 increased cAMP responses an additional 80%. We examined cAMP dynamics in live HASM cells using a fluorescence sensor. PF-04957325, a PDE8-selective inhibitor, increased basal cAMP concentrations by itself, indicating a significant basal level of cAMP synthesis. In the presence of an AC inhibitor to reduce basal signaling, PF-04957325 accelerated cAMP production and increased the inhibition of cell proliferation induced by isoproterenol, but it had no effect on cAMP concentrations or cell proliferation regulated by prostaglandin E2. Lipid raft fractionation of HASM cells revealed PDE8A immunoreactivity in buoyant fractions containing caveolin-1 and AC5/6 immunoreactivity. Thus, PDE8 is expressed in lipid rafts of HASM cells, where it specifically regulates ß2-adrenergic receptor AC6 signaling without effects on signaling by the E prostanoid receptors 2/4-AC2 complex. In airway diseases such as asthma and chronic obstructive pulmonary disease, PDE8 may represent a novel therapeutic target to modulate HASM responsiveness and airway remodeling.
Asunto(s)
3',5'-AMP Cíclico Fosfodiesterasas/metabolismo , Adenilil Ciclasas/metabolismo , Asma/enzimología , AMP Cíclico/metabolismo , Músculo Liso/enzimología , Miocitos del Músculo Liso/enzimología , Receptores Adrenérgicos beta 2/metabolismo , Sistema Respiratorio/enzimología , 3',5'-AMP Cíclico Fosfodiesterasas/genética , Adenilil Ciclasas/genética , Remodelación de las Vías Aéreas (Respiratorias) , Asma/genética , Asma/patología , Asma/fisiopatología , Estudios de Casos y Controles , Proliferación Celular , Células Cultivadas , Humanos , Microdominios de Membrana/enzimología , Microdominios de Membrana/patología , Músculo Liso/patología , Músculo Liso/fisiopatología , Miocitos del Músculo Liso/patología , Receptores Adrenérgicos beta 2/genética , Sistema Respiratorio/patología , Sistema Respiratorio/fisiopatología , Sistemas de Mensajero Secundario , Factores de TiempoRESUMEN
The host defense system with lysozyme and secretory phospholipase A2 (sPLA2) was immunohistochemically investigated in rat respiratory tract under healthy conditions. In the nasal epithelium, a large number of non-ciliated and non-microvillous cells (NC) and a small number of goblet cells (GC) were immunopositive for lysozyme and sPLA2. A few acinar cells and almost all epithelial cells of intercalated ducts were immunopositive for both bactericidal substances in the nasal glands. In the laryngeal and tracheal epithelia, few NC and GC were immunopositive for both bactericidal substances. In the laryngeal and tracheal glands, a few acinar cells and most ductal epithelial cells were immunopositive for both bactericidal substances. In extra-pulmonary bronchus, small numbers of NC and GC were immunopositive for lysozyme and sPLA2, whereas few NC and no GC were immunopositive in the intra-pulmonary bronchus. No secretory source of either bactericidal substance was located in the bronchioles. In the alveolus, many glandular epithelial cells and alveolar macrophages were immunopositive for lysozyme but immunonegative for sPLA2. Moreover, lysozyme and sPLA2 were detected in the mucus layer and in the periciliary layer from the nose to the extra-pulmonary bronchus. These findings suggest that secretory sources of lysozyme and sPLA2 are distributed in almost all the respiratory tract. Their secretory products are probably transported to the pharynx and contribute to form the first line of defense against inhaled bacteria throughout the respiratory tract.
Asunto(s)
Muramidasa/metabolismo , Fosfolipasas A2 Secretoras/metabolismo , Sistema Respiratorio/citología , Sistema Respiratorio/enzimología , Animales , Inmunohistoquímica , Masculino , Ratas Wistar , Mucosa Respiratoria/citología , Mucosa Respiratoria/enzimología , Mucosa Respiratoria/metabolismoRESUMEN
To enter into airway epithelial cells, influenza, parainfluenza- and coronaviruses rely on host cell proteases for activation of the viral protein involved in membrane fusion. One protease, transmembrane protease serine 2 (TMPRSS2) was recently proven to be crucial for hemagglutinin cleavage of some human influenza viruses. Since the catalytic sites of the diverse serine proteases linked to influenza, parainfluenza- and coronavirus activation are structurally similar, active site inhibitors of these airway proteases could have broad therapeutic applicability against multiple respiratory viruses. Alternatively, superior selectivity could be achieved with allosteric inhibitors of TMPRSS2 or another critical protease. Though still in its infancy, airway protease inhibition represents an attractive host-cell targeting approach to combat respiratory viruses such as influenza.
Asunto(s)
Gripe Humana/tratamiento farmacológico , Péptido Hidrolasas/metabolismo , Inhibidores de Proteasas/uso terapéutico , Sistema Respiratorio/enzimología , Animales , Coronavirus/efectos de los fármacos , Infecciones por Coronavirus/tratamiento farmacológico , Interacciones Huésped-Patógeno , Humanos , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/fisiología , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/fisiología , Virus de la Influenza A/efectos de los fármacos , Ratones , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Sistema Respiratorio/efectos de los fármacos , Sistema Respiratorio/virología , Serina Endopeptidasas , Serina Proteasas/efectos de los fármacos , Inhibidores de Serina Proteinasa/uso terapéuticoRESUMEN
This study evaluated the effect of tin exposure on enzyme activity in the sea cucumber (Holothuria grisea Selenka, 1867). After exposure to 0 (control), 0.04, 0.08, or 0.12 mg L-1 tin, we tested the activities of total cholinesterase in longitudinal muscles, acid phosphatase in gonads and the respiratory tree, as well as alkaline phosphatase in the intestines during a 96-h bioassay. Regression analyses showed that all enzyme activities declined with increasing tin concentrations, except for acid phosphatase in the respiratory tree, which were similarly, inhibited at all tin concentrations. These results indicate that H. grisea is a potential bioindicator for seascape habitat monitoring programs, as its biochemical markers show sensitivity to trace elements that can indicate a rise in pollution levels.
Asunto(s)
Fosfatasa Ácida/antagonistas & inhibidores , Fosfatasa Alcalina/antagonistas & inhibidores , Inhibidores de la Colinesterasa/farmacología , Holothuria/enzimología , Estaño/farmacología , Animales , Relación Dosis-Respuesta a Droga , Gónadas/enzimología , Intestinos/enzimología , Músculos/enzimología , Sistema Respiratorio/enzimologíaRESUMEN
Chronic obstructive pulmonary disease (COPD) is characterized by persistent airflow limitation and lung inflammation resulting in a progressive decline in lung function whose principle cause is cigarette smoke. MAP3K19 is a novel kinase expressed predominantly by alveolar and interstitial macrophages and bronchial epithelial cells in the lung. We found that MAP3K19 mRNA was overexpressed in a limited sampling of lung tissue from COPD patients, and a closer examination found it to be overexpressed in bronchoalveolar macrophages from COPD patients, as well as the bronchial epithelium and inflammatory cells in the lamina propria. We further found MAP3K19 to be induced in various cell lines upon environmental stress, such as cigarette smoke, oxidative and osmotic stress. Exogenous expression of MAP3K19 in cells caused an upregulation of transcriptionally active NF-κB, and secretion of the chemokines CXCL-8, CCL-20 and CCL-7. Inhibition of MAP3K19 activity by siRNA or small molecular weight inhibitors caused a decrease in cigarette smoke-induced inflammation in various murine models, which included a decrease in pulmonary neutrophilia and KC levels. In a chronic cigarette smoke model, inhibition of MAP3K19 significantly attenuated emphysematous changes in airway parenchyma. Finally, in a viral exacerbation model, mice exposed to cigarette smoke and influenza A virus showed a decrease in pulmonary neutrophilia, pro-inflammatory cytokines and viral load upon inhibition of MAP3K19. Collectively, these results suggest that inhibition of MAP3K19 may represent a novel strategy to target COPD that promises to have a potential therapeutic benefit for patients.
Asunto(s)
Mediadores de Inflamación/metabolismo , Quinasas Quinasa Quinasa PAM/metabolismo , Neumonía/enzimología , Enfermedad Pulmonar Obstructiva Crónica/enzimología , Sistema Respiratorio/enzimología , Fumar/efectos adversos , Adulto , Anciano , Anciano de 80 o más Años , Animales , Western Blotting , Línea Celular Tumoral , Citocinas/metabolismo , Células Epiteliales/enzimología , Células Epiteliales/metabolismo , Femenino , Expresión Génica , Células HEK293 , Humanos , Quinasas Quinasa Quinasa PAM/genética , Macrófagos/enzimología , Masculino , Ratones , Ratones Endogámicos BALB C , Persona de Mediana Edad , FN-kappa B/metabolismo , Neumonía/etiología , Neumonía/genética , Enfermedad Pulmonar Obstructiva Crónica/genética , Enfermedad Pulmonar Obstructiva Crónica/metabolismo , Interferencia de ARN , Sistema Respiratorio/metabolismo , Sistema Respiratorio/patología , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Arachidonic acid (AA) is a polyunsaturated fatty acid that is present in the phospholipids of the cell membranes of the body and is abundant in the brain. Exogenously administered AA has been shown to affect brain metabolism and to exhibit cardiovascular and neuroendocrine actions. However, little is known regarding its respiratory actions and/or central mechanism of its respiratory effects. Therefore, the present study was designed to investigate the possible effects of centrally injected AA on respiratory system and the mediation of the central cyclooxygenase (COX) to thromboxane A2 (TXA2) signaling pathway on AA-induced respiratory effects in anaesthetized rats. Intracerebroventricular (i.c.v.) administration of AA induced dose- and time-dependent increase in tidal volume, respiratory rates and respiratory minute ventilation and also caused an increase in partial oxygen pressure (pO2) and decrease in partial carbon dioxide pressure (pCO2) in male anaesthetized Spraque Dawley rats. I.c.v. pretreatment with ibuprofen, a non-selective COX inhibitor, completely blocked the hyperventilation and blood gases changes induced by AA. In addition, central pretreatment with different doses of furegrelate, a TXA2 synthesis inhibitor, also partially prevented AA-evoked hyperventilation and blood gases effects. These data explicitly show that centrally administered AA induces hyperventilation with increasing pO2 and decreasing pCO2 levels which are mediated by the activation of central COX to TXA2 signaling pathway.
Asunto(s)
Ácido Araquidónico/farmacología , Prostaglandina-Endoperóxido Sintasas/metabolismo , Sistema Respiratorio/efectos de los fármacos , Sistema Respiratorio/enzimología , Tromboxano A2/metabolismo , Anestesia , Animales , Benzofuranos/farmacología , Dióxido de Carbono/sangre , Relación Dosis-Respuesta a Droga , Inhibidores Enzimáticos/farmacología , Ibuprofeno/farmacología , Inyecciones Intraventriculares , Masculino , Oxígeno/sangre , Ratas Sprague-Dawley , Respiración/efectos de los fármacos , Fármacos del Sistema Respiratorio/farmacología , Volumen de Ventilación Pulmonar/efectos de los fármacos , Volumen de Ventilación Pulmonar/fisiología , Factores de TiempoRESUMEN
Infections in cystic fibrosis (CF), often involving Pseudomonas aeruginosa, result from a dysregulated airway immunity where one hallmark is the accumulation of necrotic and apoptotic immune cells, in particular neutrophils. In addition, neutrophils actively release DNA, forming neutrophil extracellular traps (NETs) that contain antimicrobial proteins. Altogether, free DNA in complex with actin accumulates in the airway lumen, resulting in highly viscous sputum that provides an anionic matrix, binding cationic antimicrobial proteins. In this study, granulocyte chemotactic protein 2 (GCP-2)/CXCL6, a neutrophil-activating chemokine with bactericidal properties, was detected in the airway epithelium of CF patients and was also present in azurophilic and specific granules of neutrophils. Elastase of neutrophils, but not of P. aeruginosa, completely degraded CXCL6 (chemokine (C-X-C motif) ligand 6). In addition, CXCL6 colocalized with extracellular DNA in both CF sputa and in in vitro-formed NETs. In vitro, CXCL6 bound DNA with a KD of 2,500 nM. Interestingly, both the bactericidal and the receptor-activating properties of CXCL6 (against neutrophils) remained largely unaffected in the presence of DNA. However, the chemotactic properties of CXCL6 were reduced by the presence of DNA. Taken together, CXCL6 is expressed in CF, retaining its functional properties even after binding to the anionic scaffold that extracellular DNA provides in CF.
Asunto(s)
Fibrosis Quística/inmunología , ADN/inmunología , Trampas Extracelulares/inmunología , Elastasa de Leucocito/inmunología , Neutrófilos/inmunología , Sistema Respiratorio/inmunología , Proteínas Bacterianas/genética , Proteínas Bacterianas/inmunología , Proteínas Bacterianas/metabolismo , Estudios de Casos y Controles , Fibrosis Quística/enzimología , Fibrosis Quística/genética , Fibrosis Quística/patología , ADN/metabolismo , Trampas Extracelulares/química , Expresión Génica , Humanos , Elastasa de Leucocito/genética , Elastasa de Leucocito/metabolismo , Activación Neutrófila , Infiltración Neutrófila , Neutrófilos/enzimología , Neutrófilos/patología , Unión Proteica , Proteolisis , Pseudomonas aeruginosa/enzimología , Sistema Respiratorio/enzimología , Sistema Respiratorio/patología , Especificidad de la Especie , Esputo/química , Esputo/inmunologíaRESUMEN
Airway hyperresponsiveness (AHR) is a hallmark feature in asthma characterized by exaggerated airway contractile response to stimuli due to increased airway sensitivity and chronic airway remodeling. We have previously shown that allergen-induced AHR in mice is associated with aberrant DNA methylation in the lung genome, suggesting that AHR could be epigenetically regulated, and these changes might predispose the animals to asthma. Previous studies demonstrated that overexpression of phosphodiesterase 4D (PDE4D) is associated with increased AHR. However, epigenetic regulation of this gene in asthmatic airway smooth muscle cells (ASMCs) has not been examined. In this study, we aimed to examine the relationship between epigenetic regulation of PDE4D and ASMC phenotypes. We identified CpG site-specific hypomethylation at PDE4D promoter in human asthmatic ASMCs. We next used methylated oligonucleotides to introduce CpG site-specific methylation at PDE4D promoter and examined its effect on ASMCs. We showed that PDE4D methylation decreased cell proliferation and migration of asthmatic ASMCs. We further elucidated that methylated PDE4D decreased PDE4D expression in asthmatic ASMCs, increased cAMP level, and inhibited the aberrant increase in Ca(2+) level. Moreover, PDE4D methylation reduced the phosphorylation level of downstream effectors of Ca(2+) signaling, including myosin light chain kinase and p38. Taken together, our findings demonstrate that gene-specific epigenetic changes may predispose ASMCs to asthma through alterations in cell phenotypes. Modulation of ASMC phenotypes by methylated PDE4D oligonucleotides can reverse the aberrant ASMC functions to normal phenotypes. This has provided new insight to the development of novel therapeutic options for this debilitative disease.
Asunto(s)
Asma/enzimología , Asma/genética , Fosfodiesterasas de Nucleótidos Cíclicos Tipo 4/genética , Metilación de ADN , Epigénesis Genética , Miocitos del Músculo Liso/enzimología , Sistema Respiratorio/enzimología , Remodelación de las Vías Aéreas (Respiratorias) , Asma/patología , Calcio/metabolismo , Estudios de Casos y Controles , Movimiento Celular , Proliferación Celular , Células Cultivadas , Islas de CpG , AMP Cíclico/metabolismo , Estudios de Asociación Genética , Predisposición Genética a la Enfermedad , Humanos , Miocitos del Músculo Liso/patología , Quinasa de Cadena Ligera de Miosina/metabolismo , Fenotipo , Fosforilación , Regiones Promotoras Genéticas , Sistema Respiratorio/patología , Transducción de Señal , Transfección , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Dipeptidyl peptidase 4 (DPP4, CD26), a type II transmembrane ectopeptidase, is the receptor for the Middle Eastern respiratory syndrome coronavirus (MERS-CoV). MERS emerged in 2012 and has a high mortality associated with severe lung disease. A lack of autopsy studies from MERS fatalities has hindered understanding of MERS-CoV pathogenesis. We investigated the spatial and cellular localization of DPP4 to evaluate an association MERS clinical disease. DPP4 was rarely detected in the surface epithelium from nasal cavity to conducting airways with a slightly increased incidence in distal airways. DPP4 was also found in a subset of mononuclear leukocytes and in serous cells of submucosal glands. In the parenchyma, DPP4 was found principally in type I and II cells and alveolar macrophages and was also detected in vascular endothelium (eg, lymphatics) and pleural mesothelia. Patients with chronic lung disease, such as chronic obstructive pulmonary disease and cystic fibrosis, exhibited increased DPP4 immunostaining in alveolar epithelia (type I and II cells) and alveolar macrophages with similar trends in reactive mesothelia. This finding suggests that preexisting pulmonary disease could increase MERS-CoV receptor abundance and predispose individuals to MERS morbidity and mortality, which is consistent with current clinical observations. We speculate that the preferential spatial localization of DPP4 in alveolar regions may explain why MERS is characterized by lower respiratory tract disease.
Asunto(s)
Infecciones por Coronavirus , Dipeptidil Peptidasa 4/biosíntesis , Sistema Respiratorio/enzimología , Adolescente , Adulto , Anciano , Niño , Preescolar , Femenino , Humanos , Inmunohistoquímica , Lactante , Masculino , Persona de Mediana Edad , Adulto JovenRESUMEN
Interferon-stimulated genes (ISGs) act in concert to provide a tight barrier against viruses. Recent studies have shed light on the contribution of individual ISG effectors to the antiviral state, but most have examined those acting on early, intracellular stages of the viral life cycle. Here, we applied an image-based screen to identify ISGs inhibiting late stages of influenza A virus (IAV) infection. We unraveled a directly antiviral function for the gene SERPINE1, encoding plasminogen activator inhibitor 1 (PAI-1). By targeting extracellular airway proteases, PAI-1 inhibits IAV glycoprotein cleavage, thereby reducing infectivity of progeny viruses. This was biologically relevant for IAV restriction in vivo. Further, partial PAI-1 deficiency, attributable to a polymorphism in human SERPINE1, conferred increased susceptibility to IAV in vitro. Together, our findings reveal that manipulating the extracellular environment to inhibit the last step in a virus life cycle is an important mechanism of the antiviral response.
Asunto(s)
Virus de la Influenza A/fisiología , Inhibidor 1 de Activador Plasminogénico/metabolismo , Serpina E2/metabolismo , Animales , Línea Celular , Humanos , Inmunidad Innata , Ratones Endogámicos C57BL , Inhibidor 1 de Activador Plasminogénico/genética , Sistema Respiratorio/enzimología , Sistema Respiratorio/virología , Serina Proteasas/metabolismo , Serpina E2/genéticaRESUMEN
BACKGROUND: Balanced levels of proteases and anti-proteases are essential in host defense systems. In CF patients' lungs, elevated protease/anti-protease-ratios contribute to damage of airway tissue and premature death with the inherited disease. Little is known about upper airway protease equilibrium in CF. METHODS: Neutrophil elastase (NE), Secretory leukocyte protease inhibitor (SLPI), matrix metalloproteinase (MMP)9, tissue inhibitors of metalloproteinase (TIMP)1, cathepsin S (CTSS) and the corresponding cellular distribution were assessed in the nasal lavage (NL) and sputum of 40 CF patients. RESULTS: Concentrations of all proteases and anti-proteases were markedly higher in sputum than in NL (NE: 10-fold, SLPI: 5000-fold). Interestingly, the NE/SLPI ratio was 726-fold higher in NL compared to sputum, while the MMP9/TIMP1 ratio was 4.5-fold higher in sputum compared to NL. DISCUSSION: This first study to compare protease/anti-protease networks of CF upper and lower airways by NL and sputum reveals substantial differences between both compartments' immunological responses. This finding may have implications for sinonasal and pulmonary treatment, possibly leading to new therapeutic approaches.
Asunto(s)
Fibrosis Quística/metabolismo , Elastasa de Leucocito/metabolismo , Líquido del Lavado Nasal/química , Sistema Respiratorio/enzimología , Inhibidor Secretorio de Peptidasas Leucocitarias/metabolismo , Esputo/enzimología , Adolescente , Adulto , Anciano , Niño , Preescolar , Fibrosis Quística/patología , Ensayo de Inmunoadsorción Enzimática , Femenino , Humanos , Masculino , Persona de Mediana Edad , Líquido del Lavado Nasal/citología , Sistema Respiratorio/patología , Esputo/citología , Adulto JovenRESUMEN
MicroRNAs have emerged as regulators of smooth muscle cell phenotype with a role in smooth muscle-related disease. Studies have shown that miR-143 and miR-145 are the most highly expressed microRNAs in smooth muscle cells, controlling differentiation and function. The effect of miR-143/145 knockout has been established in the vasculature but not in smooth muscle from other organs. Using knockout mice we found that maximal contraction induced by either depolarization or phosphatase inhibition was reduced in vascular and airway smooth muscle but maintained in the urinary bladder. Furthermore, a reduction of media thickness and reduced expression of differentiation markers was seen in the aorta but not in the bladder. Supporting the view that phenotype switching depends on a tissue-specific target of miR-143/145, we found induction of angiotensin-converting enzyme in the aorta but not in the bladder where angiotensin-converting enzyme was expressed at a low level. Chronic treatment with angiotensin type-1 receptor antagonist restored contractility in miR-143/145-deficient aorta while leaving bladder contractility unaffected. This shows that tissue-specific targets are critical for the effects of miR-143/145 on smooth muscle differentiation and that angiotensin converting enzyme is one such target.