Continuous positive airway pressure therapy suppresses inflammatory cytokines and improves glucocorticoid responsiveness in patients with obstructive sleep apnea and asthma: A case-control study.

CONTEXT: Asthma and obstructive sleep apnea (OSA) are prevalent respiratory disorders that frequently coexist. Continuous positive airway pressure (CPAP) therapy is the standard treatment for OSA. However, its effects on systemic inflammation and glucocorticoid responsiveness in OSA patients with asthma are largely unknown. AIMS: To examine the potential role of CPAP therapy in reducing systemic inflammation and improving glucocorticoid responsiveness in asthmatic patients with OSA. SETTINGS AND DESIGN: A case-control study was conducted at the respiratory and sleep clinics involving patients with OSA and patients with asthma and OSA. METHODS: The levels of inflammatory asthma biomarkers (interleukin [IL]-4, IL-17A, IL-8, IL-2, and interferon-γ [IFN-γ]), and glucocorticoid receptors (GR)-α and GR-ß, were determined to compare systemic inflammation and glucocorticoid responsiveness between pre- and post-1-month CPAP treatment in both groups. STATISTICAL ANALYSIS: The Wilcoxon signed-rank test was used to compare inflammatory biomarkers before and after CPAP therapy. P < 0.05 considered statistically significant. The analysis was performed using SPSS. RESULTS: Recruited patients (n = 47), 51% (n = 24) had OSA and 49% (n = 23), had OSA with asthma. Interestingly, the blood levels of IL-17 and IL-8 were significantly decreased post-CPAP therapy in OSA patients, whereas IL-4, IL-17, and IFN-γ were significantly reduced post-CPAP treatment in OSA patients with asthma. Remarkably, CPAP therapy improved glucocorticoid responsiveness in asthmatic patients with OSA, but not in the OSA group and an increase in the GR-α/GR-ß ratio was noted post-CPAP therapy. CONCLUSIONS: Continuous positive airway pressure therapy improved responsiveness to glucocorticoid treatment and demonstrated a suppressive effect on proinflammatory cytokines in asthmatics with OSA.

Enhanced mitophagy in bronchial fibroblasts from severe asthmatic patients.

BACKGROUND: Sub-epithelial fibrosis is a characteristic feature of airway remodeling in asthma which correlates with disease severity. Current asthma medications are ineffective in treating fibrosis. In this study, we aimed to investigate the mitochondrial phenotype in fibroblasts isolated from airway biopsies of non-asthmatic and severe asthmatic subjects by examining mitophagy as a mechanism contributing to fibroblast persistence and thereby, fibrosis in severe asthma. METHODS: Bioinformatics analysis of publicly available transcriptomic data was performed to identify the top enriched pathways in asthmatic fibroblasts. Endogenous expression of mitophagy markers in severe asthmatic and non-asthmatic fibroblasts was determined using qRT-PCR, western blot and immunofluorescence. Mitophagy flux was examined by using lysosomal protease inhibitors, E64d and pepstatin A. Mitochondrial membrane potential and metabolic activity were also evaluated using JC-1 assay and MTT assay, respectively. RESULTS: Bioinformatics analysis revealed the enrichment of Pink/Parkin-mediated mitophagy in asthmatic fibroblasts compared to healthy controls. In severe asthmatic fibroblasts, the differential expression of mitophagy genes, PINK1 and PRKN, was accompanied by the accumulation of PINK1, Parkin and other mitophagy proteins at baseline. The further accumulation of endogenous LC3BII, p62 and PINK1 in the presence of E64d and pepstatin A in severe asthmatic fibroblasts reinforced their enhanced mitophagy flux. Significantly reduced mitochondrial membrane potential and metabolic activity were also demonstrated at baseline confirming the impairment in mitochondrial function in severe asthmatic fibroblasts. Interestingly, these fibroblasts displayed neither an apoptotic nor senescent phenotype but a pro-fibrotic phenotype with an adaptive survival mechanism triggered by increased AMPKα phosphorylation and mitochondrial biogenesis. CONCLUSIONS: Our results demonstrated a role for mitophagy in the pathogenesis of severe asthma where the enhanced turnover of damaged mitochondria may contribute to fibrosis in severe asthma by promoting the persistence and pro-fibrotic phenotype of fibroblasts.

Neutrophils from severe asthmatic patients induce epithelial to mesenchymal transition in healthy bronchial epithelial cells.

BACKGROUND: Asthma is a heterogenous disease characterized by chronic inflammation and airway remodeling. An increase in the severity of airway remodeling is associated with a more severe form of asthma. There is increasing interest in the epithelial to mesenchymal transition process and mechanisms involved in the differentiation and repair of the airway epithelium, especially as they apply to severe asthma. Growing evidence suggests that Epithelial-Mesenchymal transition (EMT) could contribute to airway remodeling and fibrosis in asthma. Severe asthmatic patients with remodeled airways have a neutrophil driven inflammation. Neutrophils are an important source of TGF-ß1, which plays a role in recruitment and activation of inflammatory cells, extracellular matrix (ECM) production and fibrosis development, and is a potent inducer of EMT. OBJECTIVE: As there is little data examining the contribution of neutrophils and/or their mediators to the induction of EMT in airway epithelial cells, the objective of this study was to better understand the potential role of neutrophils in severe asthma in regards to EMT. METHODS: We used an in vitro system to investigate the neutrophil-epithelial cell interaction. We obtained peripheral blood neutrophils from severe asthmatic patients and control subjects and examined for their ability to induce EMT in primary airway epithelial cells. RESULTS: Our data indicate that neutrophils from severe asthmatic patients induce changes in morphology and EMT marker expression in bronchial epithelial cells consistent with the EMT process when co-cultured. TGF-ß1 levels in the culture medium of severe asthmatic patients were increased compared to that from co-cultures of non-asthmatic neutrophils and epithelial cells. CONCLUSIONS AND CLINICAL RELEVANCE: As an inducer of EMT and an important source of TGF-ß1, neutrophils may play a significant role in the development of airway remodeling and fibrosis in severe asthmatic airways.

Steroid-insensitive ERK1/2 activity drives CXCL8 synthesis and neutrophilia by airway smooth muscle.

Severe or refractory asthma affects 5 to 15% of all patients with asthma, but is responsible for more than half of the health burden associated with the disease. Severe asthma is characterized by a dramatic increase in smooth muscle and airway inflammation. Although glucocorticoids are the mainstay of treatment in asthma, they are unable to fully control the disease in individuals with severe asthma. We found that airway smooth muscle cells (ASMCs) from individuals with severe asthma showed elevated activities of the ERK1/ERK2 and p38 MAPK pathways despite treatment with oral and inhaled glucocorticoids, which increased the expression of DUSP1, a phosphatase shown to limit p38 MAPK activity. In ex vivo ASMCs, TNF-α but not IL-17A induced expression of the neutrophil chemoattractant CXCL8. Moreover, TNF-α led to up-regulation of the ERK1/ERK2 and p38 MAPKs pathways, with only the latter being sensitive to pretreatment with the glucocorticoid dexamethasone. In contrast to epithelial and endothelial cells, TNF-α-stimulated CXCL8 synthesis was dependent on ERK1/ERK2 but not on p38 MAPK. Moreover, suppressing ERK1/ERK2 activation prevented neutrophil recruitment by ASMCs, whereas suppressing p38 MAPK activity had no impact. Taken together, these results highlight the ERK1/ERK2 MAPK cascade as a novel and attractive target in severe asthma because the activation of this pathway is insensitive to the action of glucocorticoids and is involved in neutrophil recruitment, contributing the to inflammation seen in the disease.

Increased expression of ADAM33 and ADAM8 with disease progression in asthma.

BACKGROUND: ADAM33, a disintegrin and metalloproteinase 33 gene, has been identified as a risk factor for asthma and bronchial hyperresponsiveness and has been postulated as a gene for airway remodeling. ADAM8 is strongly induced by allergens and T(H)2 cytokines in the lung in experimental asthma. OBJECTIVES: To assess the importance of these genes in asthma pathogenesis and to investigate whether expression relates to disease severity or deterioration in lung function, we measured the mRNA and protein expression of both genes in bronchial biopsies of subjects with asthma and control subjects. METHODS: RNA was extracted from frozen endobronchial biopsies of mild, moderate, and severe adults with asthma and controls. Subjects with moderate and severe asthma were taking corticosteroids. The mRNA transcript of both genes was measured by real time RT-PCR using specific primers. Protein expression was examined by immunohistochemistry on paraffin sections. RESULTS: ADAM33 mRNA expression was significantly higher in both moderate and severe asthma compared with mild asthma (P < .05) and controls. Immunostaining for ADAM33 was increased in the epithelium, submucosal cells, and smooth muscle in severe asthma compared with mild disease and controls. ADAM8 mRNA expression was significantly increased in all asthma groups compared with controls. Increased inflammatory cells stained positive for ADAM8 in both moderate (P < .05) and severe asthma (P < .005) compared with mild disease. CONCLUSIONS: These results demonstrate increased expression of both ADAM genes as asthma severity increases. CLINICAL IMPLICATIONS: These genes may contribute to the remodeling process that occurs with asthma progression and may have implications for future treatment in severe disease.