RESUMO
BACKGROUND: Although mucus plugging is a well-reported feature of asthma, whether asthma and type 2 inflammation affect mucociliary clearance (MCC) is unknown. RESEARCH QUESTION: Does type 2 inflammation influence mucus clearance rates in patients with mild asthma who are not receiving corticosteroids? STUDY DESIGN AND METHODS: The clearance rates of inhaled radiolabeled particles were compared between patients with mild asthma with low (n = 17) and high (n = 18) levels of T2 inflammation. Fraction exhaled nitric oxide (Feno) was used to prospectively segregate subjects into T2 Lo (Feno < 25 ppb) and T2 Hi (Feno > 35 ppb) cohorts. Bronchial brush samples were collected with fiber-optic bronchoscopy, and quantitative polymerase chain reaction was performed to measure expression of genes associated with T2 asthma. MCC rate comparisons were also made with a historical group of healthy control subjects (HCs, n = 12). RESULTS: The T2 Lo cohort demonstrated increased MCC when compared with both T2 Hi and historic HCs. MCC within the T2 Hi group varied significantly, with some subjects having low or zero clearance. MCC decreased with increasing expression of several markers of T2 airway inflammation (CCL26, NOS2, and POSTN) and with Feno. MUC5AC and FOXJ1 expression was similar between the T2Lo and T2Hi cohorts. INTERPRETATION: Increasing T2 inflammation was associated with decreasing MCC. High rates of MCC in T2 Lo subjects may indicate a compensatory mechanism present in mild disease but lost with high levels of inflammation. Future studies are required to better understand mechanisms and whether impairments in MCC in more severe asthma drive worse clinical outcomes.
Assuntos
Asma , Quimiocina CCL26/antagonistas & inibidores , Inflamação/imunologia , Depuração Mucociliar/imunologia , Óxido Nítrico Sintase Tipo II/análise , Absorção pelo Trato Respiratório/imunologia , Adulto , Asma/diagnóstico , Asma/imunologia , Asma/fisiopatologia , Testes de Provocação Brônquica/métodos , Broncoscopia/métodos , Moléculas de Adesão Celular , Correlação de Dados , Estudos Transversais , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Muco/metabolismo , Compostos Radiofarmacêuticos/farmacologia , Testes de Função Respiratória/métodos , Índice de Gravidade de DoençaRESUMO
Allergic airway disease is known to cause significant morbidity due to impaired mucociliary clearance, however the mechanism that leads to the mucus dysfunction is not entirely understood. Interleukin 13 (IL-13), a key mediator of Type 2 (T2) inflammation, profoundly alters the ion transport properties of airway epithelium. However, these electrophysiological changes cannot explain the thick, tenacious airway mucus that characterizes the clinical phenotype. Here we report that IL-13 dramatically increases the airway surface liquid (ASL) viscosity in cultured primary human bronchial epithelial cells and thereby inhibits mucus clearance. These detrimental rheological changes require ATP12A, a non-gastric H+/K+-ATPase that secretes protons into the ASL. ATP12A knockdown or inhibition prevented the IL-13 dependent increase in ASL viscosity but did not alter the ASL pH. We propose that ATP12A promotes airway mucus dysfunction in individuals with T2 inflammatory airway diseases and that ATP12A may be a novel therapeutic target to improve mucus clearance.
Assuntos
ATPase Trocadora de Hidrogênio-Potássio/metabolismo , Inflamação/patologia , Depuração Mucociliar/fisiologia , Muco/química , Mucosa Respiratória/patologia , Doenças Respiratórias/patologia , Células Cultivadas , Humanos , Inflamação/metabolismo , Interleucina-13/metabolismo , Mecânica Respiratória , Mucosa Respiratória/metabolismo , Doenças Respiratórias/metabolismoRESUMO
Epithelial Na(+) channel (ENaC) function is regulated by the intracellular Na(+) concentration ([Na(+)]i) through a process known as Na(+) feedback inhibition. Although this process is known to decrease the expression of proteolytically processed active channels on the cell surface, it is unknown how [Na(+)]i alters ENaC cleavage. We show here that [Na(+)]i regulates the posttranslational processing of ENaC subunits during channel biogenesis. At times when [Na(+)]i is low, ENaC subunits develop mature N-glycans and are processed by proteases. Conversely, glycan maturation and sensitivity to proteolysis are reduced when [Na(+)]i is relatively high. Surface channels with immature N-glycans were not processed by endogenous channel activating proteases, nor were they sensitive to cleavage by exogenous trypsin. Biotin chase experiments revealed that the immature surface channels were not converted into mature cleaved channels following a reduction in [Na(+)]i. The hypothesis that [Na(+)]i regulates ENaC maturation within the biosynthetic pathways is further supported by the finding that Brefeldin A prevented the accumulation of processed surface channels following a reduction in [Na(+)]i. Therefore, increased [Na(+)]i interferes with ENaC N-glycan maturation and prevents the channel from entering a state that allows proteolytic processing.
Assuntos
Canais Epiteliais de Sódio/metabolismo , Espaço Intracelular/metabolismo , Sódio/metabolismo , Canais Epiteliais de Sódio/biossíntese , Humanos , Peptídeo Hidrolases/metabolismo , Polissacarídeos/metabolismo , Processamento de Proteína Pós-TraducionalRESUMO
Pseudomonas aeruginosa is an opportunistic pathogen that significantly contributes to the mortality of patients with cystic fibrosis. Chronic infection by Pseudomonas induces sustained immune and inflammatory responses and damage to the airway. The ability of Pseudomonas to resist host defenses is aided, in part, by secreted proteases, which act as virulence factors in multiple modes of infection. Recent studies suggest that misregulation of protease activity in the cystic fibrosis lung may alter fluid secretion and pathogen clearance by proteolytic activation of the epithelial sodium channel (ENaC). To evaluate the possibility that proteolytic activation of ENaC may contribute to the virulence of Pseudomonas, primary human bronchial epithelial cells were exposed to P. aeruginosa and ENaC function was assessed by short circuit current measurements. Apical treatment with a strain known to express high levels of alkaline protease (AP) resulted in an increase in basal ENaC current and a loss of trypsin-inducible ENaC current, consistent with sustained activation of ENaC. To further characterize this AP-induced ENaC activation, AP was purified, and its folding, activity, and ability to activate ENaC were assessed. AP folding was efficient under pH and calcium conditions thought to exist in the airway surface liquid of normal and cystic fibrosis (CF) lungs. Short circuit measurements of ENaC in polarized monolayers indicated that AP activated ENaC in immortalized cell lines as well as post-transplant, primary human bronchial epithelial cells from both CF and non-CF patients. This activation was mapped to the γ-subunit of ENaC. Based on these data, patho-mechanisms associated with AP in the CF lung are proposed wherein secretion of AP leads to decreased airway surface liquid volume and a corresponding decrease in mucocilliary clearance of pulmonary pathogens.
Assuntos
Proteínas de Bactérias/metabolismo , Brônquios/metabolismo , Endopeptidases/metabolismo , Células Epiteliais/metabolismo , Canais Epiteliais de Sódio/metabolismo , Infecções por Pseudomonas/metabolismo , Pseudomonas aeruginosa/enzimologia , Animais , Brônquios/microbiologia , Brônquios/patologia , Linhagem Celular , Polaridade Celular , Fibrose Cística/metabolismo , Fibrose Cística/microbiologia , Fibrose Cística/patologia , Células Epiteliais/microbiologia , Células Epiteliais/patologia , Humanos , Camundongos , Infecções por Pseudomonas/microbiologia , Infecções por Pseudomonas/patologia , Pseudomonas aeruginosa/patogenicidadeRESUMO
Effective clearance of inhaled pathogens is the primary innate defense mechanism in the lung, and requires the maintenance of a proper airway surface liquid (ASL) volume to facilitate ciliary beat and optimize mucociliary clearance. Na(+) absorption via the epithelial sodium channel (ENaC) is tightly regulated and, together with chloride movement, provides the optimal osmotic gradients to absorb excessive fluid in the airway lumen while preventing excessive ASL dehydration, which would compromise mucus clearance from the lung. To absorb excessive fluid from the luminal surface, a local mechanism of ENaC activation allows for an increase in Na(+) absorption at times when the ASL volume is expanded. To help define these regulatory mechanisms, we examined the effects of ASL volume expansion on ENaC activity in primary human bronchial epithelial (HBE) cell cultures. We found that ENaC activity increases dramatically after rapid dilution of endogenous ASL. Approximately 35% of the increase in Na(+) absorption was attributable to activation of ENaC by proteases. The remainder of the increase in Na(+) current was prevented when membrane trafficking was disrupted with brefeldin A, nocodazole, or myosin light chain kinase inhibitors, demonstrating that trafficking is involved with ENaC regulation in the airway. These findings demonstrate that Na(+) absorption in the airway is acutely modulated by the coordinated trafficking of channels to the luminal surface and by the proteolytic activation of ENaC in response to ASL volume expansion.
Assuntos
Brônquios/metabolismo , Células Epiteliais/enzimologia , Canais Epiteliais de Sódio/metabolismo , Epitélio/enzimologia , Peptídeo Hidrolases/metabolismo , Absorção/efeitos dos fármacos , Aprotinina/farmacologia , Brônquios/patologia , Membrana Celular/metabolismo , Polaridade Celular/efeitos dos fármacos , Células Cultivadas , Fibrose Cística/metabolismo , Fibrose Cística/patologia , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Epitélio/efeitos dos fármacos , Epitélio/patologia , Meia-Vida , Humanos , Concentração Osmolar , Inibidores de Proteases/farmacologia , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Transporte Proteico/efeitos dos fármacos , Sódio/metabolismoRESUMO
O objetivo deste trabalho, foi verificar o impacto dos programas de assistência e das condições sócio-economicas e ambientais, sobre o crescimento e desenvolvimento dos pré-escolares.Realizou-se exame antropométrico, para verificar a prevalência de desnutrição aguda, obesidade e déficit altura/idade, de crianças na idade pré-escolar, assistidas e não assistidas por Instituiçöes de Atendimento ao Pré-escolar, residentes em áreas carentes e não carentes...