Background Neural respiratory drive (NRD) measurement, reflecting the balance between respiratory muscle load and capacity, is quantified using surface parasternal electromyography (EMGpara). EMGpara tracks recovery from severe exacerbations of chronic obstructive pulmonary disease (COPD). Among stable COPD participants, we hypothesised the existence of a relationship between NRD, breathlessness and airway obstruction. Study aims: (1) assessing the feasibility of measuring EMGpara in COPD participants with forced expiratory volume in 1 s (FEV1) ≥50% predicted in primary care; (2) investigating relationships between NRD measures, self-reported breathlessness, airflow obstruction severity and health-related quality of life (HRQoL).Methods Participants with stable mild/moderate COPD, using inhaled corticosteroid (ICS) therapy, were recruited from 20 general practices. Participants were randomly allocated to continue using ICS (maintenance group) or to withdraw ICS (withdrawal group) over 6 weeks. EMGpara, spirometry, self-reported breathlessness (modified Borg dyspnoea scale), COPD Assessment Test and Chronic Respiratory Disease Questionnaire Self-Administered Standardised were measured at baseline, 3- and 6-month follow-up. Bland-Altman plots examined agreement between serial measurements.Results Forty COPD participants were recruited: age 70±9.2 years; body mass index 26±5.3 kg/m2; FEV1 1.74±0.54 L; and FEV1% pred 69.6±14.0%. High-quality EMGpara data were obtained from 35 participants at baseline and 31 participants on three occasions. High intra-rater and inter-rater agreement for EMGpara (intraclass correlation coefficient >0.9) and moderate correlation between EMGpara and FEV1% predicted (r=−0.42; p=0.01) were recorded. No correlation was observed between resting EMGpara and breathlessness or HRQoL measures across the three time points.Conclusions EMGpara measurement is feasible in primary care. In this group of COPD patients, lung function was stable across the three time points and EMGpara was associated with the degree of airflow obstruction. In the resting stable state in mild/moderate disease, there was no association between EMGpara and participant-reported outcomes. Further work should investigate the utility of EMGpara in mild/moderate COPD participants during acute exacerbation and recovery.

BACKGROUND:Eyelid dermatitis (ED) is an interdisciplinary medical challenge affecting thousands of patients worldwide. ED management can be difficult in view of the numerous differential diagnoses and limited treatment options. We review the diagnostic work-up for ED patients, with a special focus on the latest innovative solutions. OBSERVATIONS:The diagnostic work-up of ED should include medical history, exposure analysis (direct contact, transfer by hands, airborne exposure, rarely ingestion) ocular complaints, clinical severity scores for eyelid manifestations, and consider general and specialized scoring systems. Patch testing and Schirmer test modifications can be used to delineate the underlying aetiology and to narrow the ED differential diagnosis. Metal release assays (nickel spot test, cobalt spot test) as well as gold jewelry avoidance can inform on clinically relevant metal allergy in selected cases. Repeated open application tests with cosmetic products can be used on the retroauricular skin. Transepidermal water loss (TEWL) measurements should be adapted for the eyelids. Further research on eyelid microbiome and transcriptomic biomarkers in the tear fluid and/or eyelid keratinocytes is required. Atopy patch testing with house dust mites (HDMs) can be helpful in selected cases but needs further standardization. Machine learning algorithms may aid image analysis for automated patch test readings and may leverage transcriptomic data for diagnostic classifications, particularly in ambiguous cases, and treatment monitoring in ED. CONCLUSIONS AND RELEVANCE:ED diagnosis can be challenging and may require the collaboration of ophthalmologists, dermatologists, allergists, and rheumatologists. Diagnostic innovations exist but their value in the diagnostic work-up is currently unclear.

Idiopathic pulmonary fibrosis (IPF) is a disease with a poor prognosis and no curative therapies. Fibroblast activation by transforming growth factor β1 (TGFβ1) and disrupted metabolic pathways, including the arginine–polyamine pathway, play crucial roles in IPF development. Polyamines are agonists of the calcium/cation-sensing receptor (CaSR), activation of which is detrimental for asthma and pulmonary hypertension, but its role in IPF is unknown. To address this question, we evaluated polyamine abundance using metabolomic analysis of IPF patient saliva. Furthermore, we examined CaSR functional expression in human lung fibroblasts (HLFs), assessed the anti-fibrotic effects of a CaSR antagonist, NPS2143, in TGFβ1-activated normal and IPF HLFs by RNA sequencing and immunofluorescence imaging, respectively; and NPS2143 effects on polyamine synthesis in HLFs by immunoassays. Our results demonstrate that polyamine metabolites are increased in IPF patient saliva. Polyamines activate fibroblast CaSR in vitro, elevating intracellular calcium concentration. CaSR inhibition reduced TGFβ1-induced polyamine and pro-fibrotic factor expression in normal and IPF HLFs. TGFβ1 directly stimulated polyamine release by HLFs, an effect that was blocked by NPS2143. This suggests that TGFβ1 promotes CaSR activation through increased polyamine expression, driving a pro-fibrotic response. By halting some polyamine-induced pro-fibrotic changes, CaSR antagonists exhibit disease-modifying potential in IPF onset and development.

Respiratory syncytial virus (RSV) infection has been associated with disruption of the airway epithelial barrier, potentially increasing the risk of asthma development. However, whether and how RSV and RSV-induced IL-33 contribute to this process are still unclear. In vivo, 7-day-old C57BL/6 mice were infected perinasally with RSV, then viral replication, lung inflammation and barrier integrity were evaluated at various time points postinfection. In vitro, human epithelial cells were infected with RSV in the presence or absence of IL-33, and the expression and localization of apical junction complex proteins (AJC) were assessed by western blot analysis and immunofluorescence staining. The involvement of components of the IL-33/ST2/MyD88 axis was further verified through blockade of endogenous IL-33 signaling and pharmacological inhibition of MyD88. Exposure to RSV infection resulted in impairment of the airway epithelial barrier, as indicated by reduced expression of tight junction proteins (ZO-1, Occludin) and adherents junction protein (E-cadherin) in the lung tissues. These effects on epithelial barrier disruption were significantly attenuated in St2-/- mice compared with wild-type controls. In vitro, the RSV-induced epithelial barrier disruption was exacerbated by topical application of exogenous IL-33, partially through activation of MyD88-mediated NF-κB signaling. Notably, knockdown of St2 by siRNA transfection or pharmacological inhibition of MyD88 partially restored the expression of E-cadherin, ZO-1 and Occludin in RSV-infected epithelial cells. RSV infection triggers robust IL-33 release from airway epithelial cells, leading to disrupted expression of AJC protein via activation of the MyD88-dependent NF-κB signaling pathway. These findings highlight the IL-33/ST2/MyD88 axis as a critical mediator of epithelial barrier dysfunction, which may represent a potential target for therapeutic intervention in RSV-mediated lung diseases.

IgE antibodies directed against cancer antigens have demonstrated potent anti-tumour effects in pre-clinical studies. MOv18 IgE, the first-in-class IgE recognising the cancer antigen folate receptor alpha (FRα), showed preliminary signs of efficacy in a Phase I trial. Treatment was well tolerated, with the most common adverse event being transient urticarial skin reactions. We investigated immunological and allergic response parameters associated with urticarial skin reactions in MOv18 IgE-treated patients. Expression of target antigen, FRα, and MOv18 IgE reactivity with FRα or any component in human skin was studied by immunohistochemistry, immunofluorescence and immuno-mass spectrometry. We conducted transcriptomic analyses in paired lesional and non-lesional skin biopsies from a patient who developed an urticarial skin reaction. Systemic immunological markers including cytokines, β-tryptase and basophil activation states were interrogated throughout the trial and contemporaneously with the skin reaction. Of the 24 IgE-treated patients, 62.5% developed transient urticarial skin reactions, with onset during the first infusion, diminishing with consecutive infusions and no β-tryptase elevation nor clinical features indicating allergic aetiology. No FRα expression or MOv18 IgE binding to human skin was identified. Lesional skin biopsies from a patient given the highest antibody dose revealed scattered eosinophils, neutrophils and mast cell degranulation, but no increased immune cell infiltration. Transcriptomic analysis indicated pro-inflammatory, but not allergic, pathway activation. No systemic allergic or hypersensitivity mediators or basophil activation were detected. Urticarial skin reactions following MOv18 IgE treatment were unlikely to result from allergic mechanisms or skin antigen recognition. The clinical presentation is consistent with infusion-related reactions commonly observed with monoclonal antibody treatments. EudraCT number: 2014-000070-19; ClinicalTrials.gov identifier: NCT02546921, registered 11/Sept/2015.

The data that support the findings of this study are available from the corresponding author upon reasonable request. Figures S1–S2. Table S1. Video S1. Video S2. Data S1. Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.

BACKGROUND:We have previously shown that asthma-like airways inflammation may be induced by topical exposure to respiratory tract pathogens such as S. pneumoniae (SP) in concert with epithelial alarmins such as IL-33. Details of the pathogenesis of this murine surrogate remain however unexplored. METHODS:Airways inflammation was induced by repeated, intranasal exposure of Il-4-/-, Rag1-/- and Rag2-/-Il2rg-/- mice (in which B lymphocyte IgE switching, adaptive and innate immunity are respectively ablated) as well as wild type mice to inactivated SP, IL-33 or both. Airways pathological changes were analysed, and the subsets and functions of locally accumulated ILC2s investigated by single cell RNA sequencing and flow cytometry. RESULTS:In the presence of IL-33, repeated exposure of the airways to inactivated SP caused marked eosinophil- and neutrophil-rich inflammation and local accumulation of ILC2s, which was retained in the Il-4-/- and Rag1-/- deficient mice but abolished in the Rag2-/-Il2rg-/- mice, an effect partly reversed by adoptive transfer of ILC2s. Single cell sequencing analysis of ILC2s recruited following SP and IL-33 exposure revealed a Klrg1+Ly6a+subset, expressing particularly elevated quantities of the pro-inflammatory cytokine IL-6, type 2 cytokines (IL-5 and IL-13) and MHC class II molecules, promoting type 2 inflammation as well as involved in neutrophil-mediated inflammatory responses. CONCLUSION:Local accumulation of KLRG1+Ly6a+ ILC2s in the lung tissue is a critical aspect of the pathogenesis of airways eosinophilic and neutrophil-rich inflammation induced by repeated exposure to SP in the presence of the epithelial alarmin IL-33.

Objective: In a previous study we have shown that, in the presence of interleukin (IL)-33, repeated, per-nasal challenge of murine airways with Streptococcus pneumoniae (S. pneumoniae) organisms induces human asthma-like airways inflammation. It is not clear, however, whether this effect is unique or manifest in response to other common respiratory pathogens.Methods: To explore this, airways of BALB/c mice were repeatedly challenged per-nasally with formaldehyde-inactivated bacterial bodies in the presence or absence of murine recombinant IL-33. Serum concentrations of S.pneumoniae, Moraxella catarrhalis (M.catarrhalis) and Haemophilus influenzae (H.influenzae) lysates-specific IgE were measured in patients with asthma and control subjects.Results: We showed that in the presence of IL-33, repeated, per-nasal airways exposure to the bodies of these bacteria induced airways hyperresponsiveness (AHR) in the experimental mice. This was accompanied by cellular infiltration into bronchoalveolar lavage fluid (BALF), eosinophilic infiltration and mucous hypertrophy of the lung tissue, with elevated local expression of some type 2 cytokines and elevated, specific IgG and IgE in the serum. The precise characteristics of the inflammation evoked by exposure to each bacterial species were distinguishable.Conclusions: These results suggest that in the certain circumstances, inhaled or commensal bacterial body antigens of both Gram-positive (S. pneumoniae) and Gram-negative (M. catarrhalis and H. influenzae) respiratory tract bacteria may initiate type 2 inflammation typical of asthma in the airways. In addition, we demonstrated that human asthmatic patients manifest elevated serum concentrations of M.catarrhalis- and H.influenzae-specific IgE.

Chronic obstructive pulmonary disease (COPD) is a highly prevalent chronic respiratory disease characterised by irreversible airways obstruction associated with chronic airways inflammation and remodelling, while the pathogenesis and the mechanistic differences between patients remain to be fully elucidated. We previously reported that alarmin cytokine IL-33 may contribute to the production of autoantibodies against respiratory epithelial cells. Here we expand the hypothesis that pulmonary autoimmune responses induced by airway microbiota also contribute to the progression of COPD. We focused on Edwardsiella tarda which we detected uniquely in the induced sputum of patients with acute exacerbations of COPD. Pernasal challenge of the airways of WT mice with supernatants of cultured E. tarda induced marked, elevated expression of IL-33 in the lung tissues. Immunisation of animals with supernatants of cultured E. tarda resulted in significantly elevated airways inflammation, the formation of tertiary lymphatic structures and significantly elevated proportions of T follicular helper T cells in the lung tissue and mediastinal lymph nodes. Interestingly, such challenge also induced production of IgG autoantibodies directed against lung tissue lysate, alveolar epithelial cell proteins and elastin fragment, while putrescine, one of metabolites generated by the bacterium, might play an important role in the autoantibody production. Furthermore, all of these effects were partly but significantly abrogated in mice with deletion of the IL-33 receptor ST2. Collectively, these data support the hypothesis that COPD is progressed at least partly by airways microbiota such as E. tarda initiating autoimmune attack of the airways epithelium mediated at least partly through the IL-33-ST2 axis.

<p>Description of additional experiments and procedures carried out in this study</p>

Gating strategy used in the characterization of tumor-infiltrating macrophages by flow cytometry

All antibodies approved for cancer therapy are monoclonal IgGs but the biology of IgE, supported by comparative preclinical data, offers the potential for enhanced effector cell potency. Here we report a Phase I dose escalation trial (NCT02546921) with the primary objective of exploring the safety and tolerability of MOv18 IgE, a chimeric first-in-class IgE antibody, in patients with tumours expressing the relevant antigen, folate receptor-alpha. The trial incorporated skin prick and basophil activation tests (BAT) to select patients at lowest risk of allergic toxicity. Secondary objectives were exploration of anti-tumour activity, recommended Phase II dose, and pharmacokinetics. Dose escalation ranged from 70 μg–12 mg. The most common toxicity of MOv18 IgE is transient urticaria. A single patient experienced anaphylaxis, likely explained by detection of circulating basophils at baseline that could be activated by MOv18 IgE. The BAT assay was used to avoid enrolling further patients with reactive basophils. The safety profile is tolerable and maximum tolerated dose has not been reached, with evidence of anti-tumour activity observed in a patient with ovarian cancer. These results demonstrate the potential of IgE therapy for cancer.

Demarcation of tumor islets and immune cell infiltrates in rat lung sections

Evaluation of FRalpha expression by tumor cell lines

<p>Table of Patient Characteristics</p>

Legends for Supplementary Figures S1-S4

Background: The emerging role of sphingosine-1-phosphate (S1P) in regulating smooth muscle functions has led to the exploration of the possibility that this sphingolipid could represent a potential therapeutic target in asthma and other lung diseases. Several studies in animal surrogates have suggested a role for S1P-mediated signaling in the regulation of airway smooth muscle (ASM) contraction, airway hyperresponsiveness, and airway remodeling, but evidence from human studies is lacking.Objective: We sought to compare the responsiveness of the airways to S1P in healthy and asthmatic individuals in vivo, in isolated human airways ex vivo, and in murine airways dissected from healthy and house dust mite (HDM)-sensitized animals. Methods: Airway responsiveness was measured by spirometry during inhalation challenges and by wire myography in airways isolated from human and mouse lungs. Thymidine incorporation and calcium mobilization assays were used to study human ASM cell responses.Results: S1P did not induce contraction of airways isolated from healthy and HDM-exposed mice, nor in human airways. Similarly, there was no airway constriction observed in healthy and asthmatic subjects in response to increasing concentrations of inhaled S1P. However, a 30-minute exposure to S1P induced a significant concentration-dependent enhancement of airway reactivity to methacholine and to histamine in murine and human airways, respectively. HDM-sensitized mice demonstrated a significant increase in methacholine responsiveness, which was not further enhanced by S1P treatment. S1P also concentration-dependently enhanced proliferation of human ASM cells, an effect mediated through S1P receptor type 2, as shown by selective antagonism and S1P receptor type 2 small-interfering RNA knockdown.Conclusions: Our data suggest that S1P released locally into the airways may be involved in the regulation of ASM hyperresponsiveness and hyperplasia, defining a novel target for future therapies. (J Allergy Clin Immunol 2023;152:1131-40.)

<p>Table of Healthy Volunteer Characteristics</p>

INTRODUCTION:Many asthmatic patients are exposed to cigarette smoke actively or passively, which contributes to asthma exacerbation and poor control. This study is to explore the effects of cigarette smoke on pathological changes in murine surrogate of asthma.METHODS:C57BL/6 mice were sensitised and challenged with ovalbumin (OVA) to establish a surrogate of asthma and then administered with cigarette smoke extract (CSE). Airways hyperresponsiveness (AHR) was measured using the Flexivent system. Histological staining (haematoxylin-eosin [HE], periodic acid Schiff [PAS], Congo red and Masson's trichrome) was employed to measure pathological changes in sections of lung tissue of experimental mice. Enzyme-linked immunosorbent assay (ELISA) was used to measure the concentrations of total and OVA-specific IgE, cytokines and chemokines (eotaxin-1, IL-13, IL-1β, TNF-α, IL-17A, IL-33) in the lung tissue homogenates. Immunoreactivity for vWF and α-SMA in lung tissue sections was detected by immunohistochemistry.RESULTS:Exposure of the animals to CSE significantly reduced OVA-induced AHR, the number of eosinophils in bronchoalveolar lavage fluid (BALF) and eosinophils infiltrating into the lung tissue, as well as concentrations of some cytokines in lung homogenate. In contrast, it significantly enhanced the number of macrophages and M2 in BALF, as well as collagen deposition, smooth muscle thickness and alveolar destruction in lung tissue.CONCLUSION:CSE inhibits OVA-induced AHR, changes inflammation 'phenotypes', while accelerates some aspects of airways remodelling, which might contribute to worse symptoms and be refractory to anti-inflammation therapies for asthmatics.

Set up of the tumor-bearing surrogate rat model