SummaryBackground Autophagy and neutrophil extracellular DNA traps (NETs) are implicated in asthma; however, their roles in asthma pathogenesis have not been elucidated. Objectives We compared autophagy and NET production levels from peripheral blood neutrophils (PBNs) of patients with severe asthma (SA) and non-severe asthma (NSA). Additionally, we investigated the inflammatory effects of NETs on human airway epithelial cells (AECs) and peripheral blood eosinophils (PBEs). Methods Peripheral blood neutrophils from patients with SA (n = 30) and NSA (n = 38) were treated with interleukin (IL)-8 (100 ng/mL). Autophagy (light chain 3-II expression) and NET production levels were evaluated by Western blot, immunofluorescence microscopy, and PicoGreen assay. The effects of NETs on AECs were assessed by investigating cell death, cell detachment, expression of occludin and claudin-1, and IL-8 production; the effects of NETs on PBEs were examined by investigating the activation and release of eosinophil cationic protein (ECP) and eosinophil-derived neurotoxin (EDN). Results Untreated and IL-8-treated PBNs from the SA group produced higher autophagy and NET levels compared with those from the NSA group (P < 0.01). IL-8 increased autophagy and NET levels in PBNs from the SA group, but not from the NSA group. NET levels were correlated with autophagy levels in PBNs (P < 0.001). IL-8-induced NET production levels negatively were correlated with FEV1/FVC (r = À0.700, P = 0.016). NETs induced cell death, detachment, degradation of occludin and claudin-1, and IL-8 production from AECs. Higher levels of NET-induced ECP and EDN were released from PBEs in SA compared with NSA groups. Conclusions and Clinical Relevance Neutrophil autophagy and NETs could enhance asthma severity by damaging airway epithelium and triggering inflammatory responses of AECs and PBEs. Modulating neutrophil autophagy and NET production may be a new target therapy for SA.

Autophagy could play a role in the pathogenesis of severe asthma. Autophagy modulation may be a novel therapeutic target for conventional therapy-resistant severe asthma.

SummaryBackground To date, there has been no reliable in vitro test to diagnose aspirin-exacerbated respiratory disease (AERD). Objective To investigate potential diagnostic biomarkers for AERD using metabolomic analysis. Methods An untargeted profile of serum from asthmatics in the first cohort (group 1) comprising 45 AERD, 44 patients with aspirin-tolerant asthma (ATA), and 28 normal controls was developed using the ultra-high-performance liquid chromatography (UHPLC)/QToF MS system. Metabolites that discriminate AERD from ATA were quantified in both serum and urine, which were collected before (baseline) and after the lysine-aspirin bronchoprovocation test (Lys-ASA BPT). The serum metabolites were validated in the second cohort (group 2) comprising 50 patients with AERD and 50 patients with ATA. Results A clear discrimination of metabolomes was found between patients with AERD and ATA. In group 1, serum levels of LTE 4 and LTE 4 /PGF 2 a ratio before and after the Lys-ASA BPT were significantly higher in patients with AERD than in patients with ATA (P < 0.05 for each), and urine baseline levels of these two metabolites were significantly higher in patients with AERD. Significant differences of serum metabolite levels between patients with AERD and ATA were replicated in group 2 (P < 0.05 for each). Moreover, serum baseline levels of LTE 4 and LTE 4 /PGF 2 a ratio discriminated AERD from ATA with 70.5%/71.6% sensitivity and 41.5%/62.8% specificity, respectively (AUC = 0.649 and 0.732, respectively P < 0.001 for each). Urine baseline LTE 4 levels were significantly correlated with the fall in FEV 1 % after the Lys-ASA BPT in patients with AERD (P = 0.008, r = 0.463). Conclusions and Clinical Relevance Serum metabolite level of LTE 4 and LTE 4 /PGF 2 a ratio was identified as potential in vitro diagnostic biomarkers for AERD using the UHPLC/QToF MS system, which were closely associated with major pathogenetic mechanisms underlying AERD.

Background The long‐term goals of asthma treatment are to achieve well control of symptoms and to minimize the future risk of asthma exacerbation. Identifying biomarkers for uncontrolled asthma is important for improving the asthma outcome. This study aimed to investigate the association of the levels of eosinophil‐derived neurotoxin (EDN) with asthma control status in specific asthma phenotype, aspirin‐exacerbated respiratory disease (AERD), and aspirin‐tolerant asthma (ATA). Methods A total of 136 adult asthmatics, including 47 asthmatics with AERD and 89 asthmatics with ATA, were enrolled. Plasma, sputum, and urine were collected at enrollment and the levels of EDN were measured by the K‐EDN ELISA kit. Urinary leukotriene E4 (LTE 4 ) level was measured using liquid chromatography–mass spectrometry (LC‐MS)/MS methods. Asthma control status was evaluated according to the GINA guideline, asthma control test and asthma control questionnaire scores. Results In the total study subjects, sputum levels of EDN as well as of urine and plasma EDN showed significantly higher levels in patients with uncontrolled asthma than in those with well‐controlled or partly‐controlled asthma (ANOVA, p < 0.001); in patients with AERD, the sputum EDN levels showed significant correlations with ACT, ACQ, and AQLQ scores ( p = 0.010, r = −0.536, p = 0.001, r = 0.665, and p < 0.001, r = −0.691, respectively), while no differences were noted in patients with ATA. Sputum EDN level was the only significant factor for ACT, ACQ, and AQLQ scores in patients with AERD ( p = 0.001, p < 0.001, and p < 0.001, respectively) in the multivariate analysis adjusting for age, sex, peripheral eosinophil count, and urine LTE 4 . The ROC curve analysis demonstrated that sputum EDN can predict uncontrolled asthma with 80% sensitivity and 88.2% specificity for ACT ≤ 19 (area under the ROC curve [AUC] = 0.824, p = 0.019); 71.4% sensitivity and 86.7% specificity for ACQ ≥ 1.5 (AUC = 0.752, p = 0.049) only in AERD patients. Conclusion The level of sputum EDN may be a potential biomarker for identifying the asthma control status in patients with AERD.