BACKGROUND:Bronchiolitis is a common viral respiratory disease of infants, with severity ranging from mild symptoms, such as coryza and feeding difficulties, to fulminant respiratory failure. Endotracheal administration of exogenous surfactant has been shown in small studies to improve gas exchange in critically ill infants with bronchiolitis. We aimed to investigate the safety and efficacy of endotracheal poractant alfa for treating critical bronchiolitis compared with a sham procedure. METHODS:BESS was a multicentre, blinded, randomised, sham-controlled, parallel-group, phase 2, superiority trial with exploratory mechanism evaluation studies. The trial was done in 15 paediatric intensive care units in the devolved National Health Service (NHS) of England, Scotland, and Northern Ireland. Preterm and term-born infants younger than 26 weeks of gestationally corrected age admitted to hospitals with bronchiolitis requiring invasive mechanical ventilation (IMV) were randomly assigned (1:1) to receive up to three doses of endotracheal poractant alfa (Curosurf) or sham intervention, allocated through web-based randomisation. Randomisation was stratified by duration of IMV before randomisation (<24 h and ≥24 h) and by site. The infants and their families, clinical care staff, Liverpool Clinical Trial Centre staff, and members of the site research teams were masked to treatment allocations. Endotracheal poractant alfa was given initially at 200 mg/kg, followed by 100 mg/kg at 12 h intervals. The primary endpoint was the duration of IMV from randomisation to final successful extubation. All infants who were successfully extubated were included in the intention-to-treat analysis. Safety outcomes were analysed in infants who had received at least one trial intervention. This trial was registered prospectively with ISRCTN (ISRCTN11746266) and EudraCT (2018-001169-18), and is completed. FINDINGS:The trial was completed after six recruitment seasons. Between Dec 18, 2018, to March 31, 2024, 1009 infants were assessed for eligibility, 232 of whom were randomly assigned to receive either endotracheal poractant alfa (n=115) or a sham intervention (n=117). 130 (56%) of 232 infants were male and 102 (44%) were female. Three infants were withdrawn from the study. None were lost to follow-up. The median duration of IMV was 64·9 h (IQR 43·2-92·1) in the endotracheal poractant alfa group and 62·0 h (39·3-95·1) in the sham intervention group. The geometric mean ratio was 1·02 (95% CI 0·84-1·24; t-test p=0·86). No clinically significant safety issues were associated with endotracheal poractant alfa and there were no deaths. INTERPRETATION:Poractant alfa, administered endotracheally to infants with early critical bronchiolitis, although safe, did not reduce the duration of IMV compared with the sham intervention. Therefore, our findings suggest that it should not be used for this indication at this dose and administration method. FUNDING:UK National Institute for Health and Care Research, UK Research and Innovation Medical Research Council, Chief Scientist Office Scotland, Health and Social Care Research and Development Division Northern Ireland, and Chiesi Farmaceutici, Italy.

Rapid point of care assessment of pulmonary surfactant composition by measuring the lecithin/sphingomyelin (L/S) ratio could improve management of patients with neonatal respiratory distress syndrome (nRDS). Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) offers a practical route to making such measurements, but the influence of the sample solvent prior to drying on measurement repeatability is poorly understood. We compare films dried from dichloromethane (DCM) and water (AQ) solvents (DCM-dry route vs. AQ-dry route) by ATR-FTIR and show that spectra from the AQ-dry route increased the signal-to-noise ratio (SNR) of a representative (2920 cm-1) absorption peak for the mixture from 20.13 to 128.20 and for human endotracheal aspirate (ETA) from 6.33 to 8.13. A mixed nested analysis of variance (ANOVA) showed that drying route accounted for 89.52% of mixture peak height variance and reduced percent relative standard deviation (%RSD) from 23.5% to 16.2%, corroborated by multivariate analysis for ETA. We further demonstrate that partial least squares regression (PLSR) models trained on AQ-dry mixture spectra predicted L/S (R2 = 0.91; root mean square error (RMSE) = 0.31) with 95% prediction interval grey-zone interpretation around L/S = 2.2, complemented by a receiver operating characteristic area under the curve (ROC-AUC) of 0.978.

COV-SARS-2 targets alveolar type II cells. As these cells synthesise lung surfactant, we hypothesised that surfactant dysfunction may contribute to development of ARDS in COVID-19. Here we report turnover of surfactant delivered to patients ventilated for severe COVID-19 using a novel breath-synchronized nebulizer compared with a control group. Endogenous surfactant status, turnover and half-life of administered surfactant and tracheal aspirate (TA) phospholipid metabolism were analysed by lipidomic mass spectrometry. At enrolment shortly after intubation, TA analysis (n = 20) showed markedly reduced concentrations of surfactant phospholipids, consistent with surfactant depletion. In a dose-range study in 12 ventilated COVID-19 patients with ARDS, administered surfactant resulted mean 20-fold (range 6.4 and 60.3) excess over endogenous lipid, providing proof of concept for effective nebulization, with a very rapid turnover (median half-life 7.8, range 0.4 to 20.8 h). Neither the rate of endogenous TA phosphatidylcholine (PC) synthesis nor the composition of newly synthesised PC, determined by incorporation of methyl-D9-choline, were significantly altered by exogenous surfactant nebulization. An inverse correlation between the fractional synthesis of dipalmitoyl phosphatidylcholine and inflammatory status suggested that a significant portion of endogenous TA phospholipid was derived from non-surfactant sources. This analysis is the first direct demonstration of surfactant deficiency in COVID-19; while exogenous surfactant can correct this deficiency, its rapid turnover suggests that prolonged treatment with surfactant will be needed.

Supplemental oxygen is an essential therapy during critical illness. However, patients with severe hypoxemic respiratory failure and/or acute respiratory distress syndrome often require high oxygen concentrations, exposing lungs to alveolar hyperoxia despite systemic hypoxemia, with consequent pulmonary oxygen toxicity. Pulmonary oxygen toxicity causes disruption of surfactant, which is essential for maintenance of alveolar functional anatomy, as well as efficient and effective gas exchange and immune regulation. This surfactant dysregulation can increase alveolar surface tension, causing alveolar collapse with atelectasis, resulting in poor lung compliance and impaired gas exchange. Hyperoxia-induced lung injury mechanisms may interact with mechanisms of harm associated with infections and mechanical ventilation. The intricate relationship between these different, interrelated stressors and altered surfactant metabolism and function is not yet delineated, particularly in humans. This review examines the current understanding of hyperoxia-induced surfactant dysregulation. We discuss potential mechanisms, including biochemical/compositional and functional changes to lipids and proteins including surfactant proteins A and D, epithelial atrophy, impaired surfactant synthesis/metabolism, redox imbalances, phospholipase A2, and altered macrophage clearance. Key areas for future research are outlined, emphasizing the need for clinically relevant human models that discriminate between the effects of oxygen therapy dose and duration, as well as other iatrogenic effects and underlying disease processes. We propose a roadmap to progress current knowledge and outline opportunities for well-designed human studies, novel surfactant preparations resistant to functional inhibition and breakdown, and technological developments, with the potential for leveraging these to identify innovative biomarkers, individualized therapeutic targets, and novel therapies in the future.

Vibrational spectroscopy, in the form of Raman and mid-IR spectroscopies, is a powerful, label-free technique that can identify and quantify individual components of a complex chemical mixture which makes it ideal for analysis of biosamples. Vibrational spectroscopic measurements are quick, simple and easy to perform, enabling rapid screening, and can be employed for the diagnosis of multiple conditions at once and can be employed for the diagnosis of multiple conditions at once. We explore the use of vibrational spectroscopy for rapid point-of-care diagnosis by considering we study the diagnosis of neonatal respiratory distress syndrome of preterm babies. We are developing on-chip spectroscopy using low-cost and highly sensitive disposable chips. Together with machine learning, it will further simplify biomarker detection in critical care for rapid and early diagnosis of disease.

Access to distal airway samples to assess respiratory diseases is not straightforward and requires invasive procedures such as bronchoscopy and bronchoalveolar lavage. The particles in exhaled air (PExA) device provides a non-invasive means of assessing small airways; it captures distal airway particles (PEx) sized around 0.5–7 μm and contains particles of respiratory tract lining fluid (RTLF) that originate during airway closure and opening. The PExA device can count particles and measure particle mass according to their size. The PEx particles can be analysed for metabolites on various analytical platforms to quantitatively measure targeted and untargeted lung specific markers of inflammation. As such, the measurement of distal airway components may help to evaluate acute and chronic inflammatory conditions such as asthma, chronic obstructive pulmonary disease, acute respiratory distress syndrome, and more recently, acute viral infections such as COVID-19. PExA may provide an alternative to traditional methods of airway sampling, such as induced sputum, tracheal aspirate, or bronchoalveolar lavage. The measurement of specific biomarkers of airway inflammation obtained directly from the RTLF by PExA enables a more accurate and comprehensive understanding of pathophysiological changes at the molecular level in patients with acute and chronic lung diseases.

Lipids and their mediators have important regulatory functions in many cellular processes, including the innate antiviral response. The aim of this study was to compare the lipid membrane composition of in vitro differentiated primary bronchial epithelial cells (PBECs) with ex vivo bronchial brushings and to establish whether any changes in the lipid membrane composition affect antiviral defense of cells from donors without and with severe asthma. Using mass spectrometry, we showed that the lipid membrane of in vitro differentiated PBECs was deprived of polyunsaturated fatty acids (PUFAs) compared to ex vivo bronchial brushings. Supplementation of the culture medium with arachidonic acid (AA) increased the PUFA-content to more closely match the ex vivo membrane profile. Rhinovirus (RV16) infection of AA-supplemented cultures from healthy donors resulted in significantly reduced viral replication while release of inflammatory mediators and prostaglandin E2 (PGE2) was significantly increased. Indomethacin, an inhibitor of prostaglandin-endoperoxide synthases, suppressed RV16-induced PGE2 release and significantly reduced CXCL-8/IL-8 release from AA-supplemented cultures indicating a link between PGE2 and CXCL8/IL-8 release. In contrast, in AA-supplemented cultures from severe asthmatic donors, viral replication was enhanced whereas PTGS2 expression and PGE2 release were unchanged and CXCL8/IL-8 was significantly reduced in response to RV16 infection. While the PTGS2/COX-2 pathway is initially pro-inflammatory, its downstream products can promote symptom resolution. Thus, reduced PGE2 release during an RV-induced severe asthma exacerbation may lead to prolonged symptoms and slower recovery. Our data highlight the importance of reflecting the in vivo lipid profile in in vitro cell cultures for mechanistic studies.

Mammalian cell membranes composed of a mixture of glycerophospholipids, the relative composition of individual phospholipids and the dynamic flux vary between cells. In addition to their structural role, membrane phospholipids are involved in cellular signalling and immunomodulatory functions. In this study, we investigate the molecular membrane composition and dynamic flux of phosphatidylcholines in CD15+ leucocytes and CD3+ lymphocytes extracted from patients with acute respiratory distress syndrome (ARDS). We identified compositional variations between these cell types, where CD15+ cells had relatively higher quantities of alkyl-acyl PC species and CD3+ cells contained more arachidonoyl-PC species. There was a significant loss of arachidonoyl-PC in CD3+ cells in ARDS patients. Moreover, there were significant changes in PC composition and the methyl-D9 enrichment of individual molecular species in CD15+ cells from ARDS patients. This is the first study to perform an in vivo assessment of membrane composition and dynamic changes in immunological cells from ARDS patients.

Early diagnosis of neonatal respiratory distress syndrome (nRDS) is important in reducing the mortality of preterm babies. Knowledge of the ratio of two components of lung surfactant, dipalmitoylphosphatidylcholine (DPPC), and sphingomyelin (SM) can be used as biomarkers of lung maturity and inform treatment. Raman spectroscopy is a powerful tool to analyze vibrational spectra of organic molecules which requires only limited sample preparation steps and, unlike IR spectroscopy, is not masked by water absorption. In this paper, we explore the potential of using Raman spectroscopy as a tool to estimate the ratio of DPPC and SM from aqueous vesicles of binary mixture of DPPC and SM. We demonstrate that the ratio of DPPC and SM can be estimated by estimating the ratio of intensity of CO stretch of DPPC and CC stretch of SM as well as CO stretch of DPPC and amide I of SM. Further, we employ a partial least squares regression (PLSR) model to automate the estimation and demonstrate that PLSR method can predict the DPPC and SM ratio with an R-2 value of 0.968.

Acute lung Injury leads to alterations in surfactant lipid composition and metabolism. Although several mechanisms contribute to dysregulated surfactant metabolism, studies investigating in vivo surfactant metabolism are limited. The aim of this study is to characterise surfactant phospholipid composition and flux utilising a stable isotope labelling technique in mechanically ventilated paediatric patients. Paediatric patients (<16 years of age) received 3.6 mg/kg intravenous methyl-D9-choline chloride followed by the endotracheal instillation of 100 mg/kg of exogenous surfactant after 24 h. Bronchioalveolar fluid samples were taken at baseline and 12, 24, 36, 48, 72 and 96 h after methyl-D9-choline infusion. Nine participants (median age of 48 days) were recruited. The primary phosphatidylcholine (PC) composition consisted of PC16:0/16:0 or DPPC (32.0 ± 4.5%). Surfactant supplementation resulted in a 30% increase in DPPC. Methyl-D9 PC enrichment was detected after 12 h and differed significantly between patients, suggesting variability in surfactant synthesis/secretion by the CDP-choline pathway. Peak enrichment was achieved (0.94 ± 0.15% of total PC) at 24 h after methyl-D9-choline infusion. There was a trend towards reduced enrichment with the duration of mechanical ventilation prior to study recruitment; however, this was not statistically significant (p = 0.19). In this study, we demonstrated the fractional molecular composition and turnover of surfactant phospholipids, which was highly variable between patients.

Background: The anti-IgE monoclonal antibody omalizumab is widely used for severe asthma. This study aimed to identify biomarkers that predict clinical improvement during 1 year of omalizumab treatment. Methods: One-year open-label Study of Mechanisms of action of Omalizumab in Severe Asthma (SoMOSA) involving 216 patients with severe (Global Initiative for Asthma step 4/5) uncontrolled atopic asthma (at least two severe exacerbations in the previous year) taking high-dose inhaled corticosteroids and long-acting β-agonists with or without maintenance oral corticosteroids. It had two phases: 0-16 weeks, to assess early clinical improvement by Global Evaluation of Therapeutic Effectiveness (GETE); and 16-52 weeks, to assess late responses based on ⩾50% reduction in exacerbations or mOCS dose. All participants provided samples (exhaled breath, blood, sputum, urine) before and after 16 weeks of omalizumab treatment. Measurements and Main Results: A total of 191 patients completed phase 1; 63% had early improvement. Of 173 who completed phase 2, 69% had reduced exacerbations by ⩾50% and 57% (37 of 65) taking mOCSs had reduced their dose by ⩾50%. The primary outcomes 2,3-dinor-11-β-PGF2α, GETE score, and standard clinical biomarkers (blood and sputum eosinophils, exhaled nitric oxide, serum IgE) did not predict either clinical response. Five volatile organic compounds and five plasma lipid biomarkers strongly predicted the ⩾50% reduction in exacerbations (receiver operating characteristic areas under the curve of 0.780 and 0.922, respectively) and early responses (areas under the curve of 0.835 and 0.949, respectively). In an independent cohort, gas chromatography/mass spectrometry biomarkers differentiated between severe and mild asthma. Conclusions: This is the first discovery of omics biomarkers that predict improvement in asthma with biologic agent treatment. Prospective validation and development for clinical use is justified.

Neonatal respiratory distress syndrome (nRDS) is a challenging condition to diagnose which can lead to delays in receiving appropriate treatment. Mid infrared (IR) spectroscopy is capable of measuring the concentrations of two diagnostic nRDS biomarkers, lecithin (L) and sphingomyelin (S) with the potential for point of care (POC) diagnosis and monitoring. The effects of varying other lipid species present in lung surfactant on the mid IR spectra used to train machine learning models are explored. This study presents a lung lipid model of five lipids present in lung surfactant and varies each in a systematic approach to evaluate the ability of machine learning models to predict the lipid concentrations, the L/S ratio and to quantify the uncertainty in the predictions using the jackknife + -after-bootstrap and variant bootstrap methods. We establish the L/S ratio can be determined with an uncertainty of approximately ±0.3 mol/mol and we further identify the 5 most prominent wavenumbers associated with each machine learning model.

Background MecROX is a mechanistic sub-study of the UK-ROX trial which was designed to evaluate the clinical and cost-effectiveness of a conservative approach to oxygen therapy for invasively ventilated adults in intensive care. This is based on the scientific rationale that excess oxygen is harmful. Epithelial cell damage with alveolar surfactant deficiency is characteristic of hyperoxic acute lung injury. Additionally, hyperoxaemia (excess blood oxygen levels) may exacerbate whole-body oxidative stress leading to cell death, autophagy, mitochondrial dysfunction, bioenergetic failure and multi-organ failure resulting in poor clinical outcomes. However, there is a lack of in-vivo human models evaluating the mechanisms that underpin oxygen-induced organ damage in mechanically ventilated patients. Aim The aim of the MecROX mechanistic sub-study is to assess lung surfactant composition and global systemic redox status to provide a mechanistic and complementary scientific rationale to the UK-ROX trial findings. The objectives are to quantify in-vivo surfactant composition, synthesis, and metabolism with markers of oxidative stress and systemic redox disequilibrium (as evidenced by alterations in the ‘reactive species interactome’) to differentiate between groups of conservative and usual oxygen targets. Methods and design After randomisation into the UK-ROX trial, 100 adult participants (50 in the conservative and 50 in usual care group) will be recruited at two trial sites. Blood and endotracheal samples will be taken at 0, 48 and 72 hours following an infusion of 3 mg/kg methyl-D9-choline chloride. This is a non-radioactive, stable isotope of choline (vitamin), which has been extensively used to study surfactant phospholipid kinetics in humans. This study will mechanistically evaluate the in-vivo surfactant synthesis and breakdown (by hydrolysis and oxidation), oxidative stress and redox disequilibrium from sequential plasma and bronchial samples using an array of analytical platforms. We will compare conservative and usual oxygenation groups according to the amount of oxygen administered. Trial registration: ISRCTN ISRCTN61929838, 27/03/2023 https://doi.org/10.1186/ISRCTN61929838.

Maternal obesity is associated with increased risk of diabetes, cardiovascular disease and hypertension in adult offspring. Midlife hypercholesterolemia and hypertension are risk factors for Alzheimer's disease, suggesting that the ageing brain may be impacted by early life environment. We found that exposure to a high fat diet during gestation and lactation induced changes in multiple components of the neurovascular unit, including a downregulation in apolipoprotein E and fibronectin, an upregulation in markers of astrocytes and perivascular macrophages and altered blood vessel morphology in the brains of adult mice. Feeding of high fat diet after weaning increased lipid droplets in the brain and influenced the fatty acid composition of phosphatidylcholine and phosphatidylethanolamine species, but did not affect the neurovascular unit. Sustained high fat diet over the entire lifespan resulted in additional decreases in levels of pericytes and collagen IV, changes in phospholipid composition and impaired perivascular clearance of Beta-amyloid (A-Beta) from the brain. In humans, vascular A-Beta load was significantly increased in the brains of aged individuals with a history of hypercholesterolemia. These results support a critical role for early dietary influence on the brain vasculature across the lifespan, with consequences for the development of age-related cerebrovascular and neurodegenerative diseases.

Aqueous lipid solutions were analyzed by mid-IR and Raman spectroscopies to establish a diagnosis approach for neonatal respiratory distress syndrome. We show the ability to predict lipid ratios around the diagnostic cutoff of 2.2.

SARS-CoV-2 directly targets alveolar epithelial cells and can lead to surfactant deficiency. Early reports suggested surfactant replacement may be effective in improving outcomes. The aim of the study to assess the feasibility and efficacy of nebulized surfactant in mechanically ventilated COVID-19 patients. Patients were randomly assigned to receive open-labelled bovine nebulized surfactant or control (ratio 3-surfactant: 2-control). This was an exploratory dose–response study starting with 1080 mg of surfactant delivered at 3 time points (0, 8 and 24 h). After completion of 10 patients, the dose was reduced to 540 mg, and the frequency of nebulization was increased to 5/6 time points (0, 12, 24, 36, 48, and an optional 72 h) on the advice of the Trial Steering Committee. The co-primary outcomes were improvement in oxygenation (change in PaO2/FiO2 ratio) and ventilation index at 48 h. 20 patients were recruited (12 surfactant and 8 controls). Demographic and clinical characteristics were similar between groups at presentation. Nebulized surfactant administration was feasible. There was no significant improvement in oxygenation at 48 h overall. There were also no differences in secondary outcomes or adverse events. Nebulized surfactant administration is feasible in mechanically ventilated patients with COVID-19 but did not improve measures of oxygenation or ventilation.

Acute hypoxic respiratory failure (AHRF) is a prominent feature of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) critical illness. The severity of gas exchange impairment correlates with worse prognosis, and AHRF requiring mechanical ventilation is associated with substantial mortality. Persistent impaired gas exchange leading to hypoxemia often warrants the prolonged administration of a high fraction of inspired oxygen (FiO2). In SARS-CoV-2 AHRF, systemic vasculopathy with lung microthrombosis and microangiopathy further exacerbates poor gas exchange due to alveolar inflammation and oedema. Capillary congestion with microthrombosis is a common autopsy finding in the lungs of patients who die with coronavirus disease 2019 (COVID-19)-associated acute respiratory distress syndrome. The need for a high FiO2 to normalise arterial hypoxemia and tissue hypoxia can result in alveolar hyperoxia. This in turn can lead to local alveolar oxidative stress with associated inflammation, alveolar epithelial cell apoptosis, surfactant dysfunction, pulmonary vascular abnormalities, resorption atelectasis, and impairment of innate immunity predisposing to secondary bacterial infections. While oxygen is a life-saving treatment, alveolar hyperoxia may exacerbate pre-existing lung injury. In this review, we provide a summary of oxygen toxicity mechanisms, evaluating the consequences of alveolar hyperoxia in COVID-19 and propose established and potential exploratory treatment pathways to minimise alveolar hyperoxia.

Acute respiratory distress syndrome (ARDS) is a major cause of hypoxemic respiratory failure in adults, leading to the requirement for mechanical ventilation and poorer outcomes. Dysregulated surfactant metabolism and function are characteristic of ARDS. A combination of alveolar epithelial damage leading to altered surfactant synthesis, secretion, and breakdown with increased functional inhibition from overt alveolar inflammation contributes to the clinical features of poor alveolar compliance and alveolar collapse. Quantitative and qualitative alterations in the bronchoalveolar lavage and tracheal aspirate surfactant composition contribute to ARDS pathogenesis. Compared to neonatal respiratory distress syndrome (nRDS), replacement studies of exogenous surfactants in adult ARDS suggest no survival benefit. However, these studies are limited by disease heterogeneity, variations in surfactant preparations, doses, and delivery methods. More importantly, the lack of mechanistic understanding of the exact reasons for dysregulated surfactant remains a significant issue. Moreover, studies suggest an extremely short half-life of replaced surfactant, implying increased catabolism. Refining surfactant preparations and delivery methods with additional co-interventions to counteract surfactant inhibition and degradation has the potential to enhance the biophysical characteristics of surfactant in vivo.

Background Surfactant phospholipid (PL) composition plays an important role in lung diseases. We compared the PL composition of non-invasively collected exhaled breath particles (PEx) with bronchoalveolar lavage (BAL) and induced sputum (ISP) at baseline and following endotoxin (LPS) challenges. Methods PEx and BAL were collected from ten healthy nonsmoking participants before and after segmental LPS challenge. Four weeks later, PEx and ISP were sampled in the week before and after a whole lung LPS inhalation challenge. PL composition was analysed using mass spectrometry. Results The overall PL composition of BAL, ISP and PEx was similar, with PC(32:0) and PC(34:1) representing the largest fractions in all three sample types (baseline PC(32:0) geometric mean mol%: 52.1, 56.9, and 51.7, PC(34:1) mol%: 11.7, 11.9 and 11.4, respectively). Despite this similarity, PEx PL composition was more closely related to BAL than to ISP. For most lipids comparable inter-individual differences in BAL, ISP, and PEx were found. PL composition of PEx was repeatable. The most pronounced increase following segmental LPS challenge was detected for SM(d34:1) in BAL (0.24 to 0.52 mol%) and following inhalation LPS challenge in ISP (0.45 to 0.68 mol%). An increase of SM(d34:1) following segmental LPS challenge was also detectable in PEx (0.099 to 0.103 mol%). The inhalation challenge did not change PL composition of PEx. Conclusion Our data supports the peripheral origin of PEx. The lack of PL changes in PEx after inhalation challenge might to be due to the overall weaker response of inhaled LPS which primarily affects the larger airways. Compared with BAL, which always contains lining fluid from both peripheral lung and central airways, PEx analysis might add value as a selective and non-invasive method to investigate peripheral airway PL composition. Trial registration NCT03044327, first posted 07/02/2017.

Background: Asthma is a chronic respiratory disease with significant heterogeneity in its clinical presentation and pathobiology. There is need for improved understanding of respiratory lipid metabolism in asthma patients and its relation to observable clinical features.Objective: We performed a comprehensive, prospective, cross-sectional analysis of the lipid composition of induced sputum supernatant obtained from asthma patients with a range of disease severities, as well as from healthy controls.Methods: Induced sputum supernatant was collected from 211 adults with asthma and 41 healthy individuals enrolled onto the U-BIOPRED (Unbiased Biomarkers for the Prediction of Respiratory Disease Outcomes) study. Sputum lipidomes were characterized by semiquantitative shotgun mass spectrometry and clustered using topologic data analysis to identify lipid phenotypes.Results: Shotgun lipidomics of induced sputum supernatant revealed a spectrum of 9 molecular phenotypes, highlighting not just significant differences between the sputum lipidomes of asthma patients and healthy controls, but also within the asthma patient population. Matching clinical, pathobiologic, proteomic, and transcriptomic data helped inform the underlying disease processes. Sputum lipid phenotypes with higher levels of nonendogenous, cell-derived lipids were associated with significantly worse asthma severity, worse lung function, and elevated granulocyte counts.Conclusion: We propose a novel mechanism of increased lipid loading in the epithelial lining fluid of asthma patients resulting from the secretion of extracellular vesicles by granulocytic inflammatory cells, which could reduce the ability of pulmonary surfactant to lower surface tension in asthmatic small airways, as well as compromise its role as an immune regulator. (J Allergy Clin Immunol 2023;152:117-25.)