[BAD] EK9 - Abstracts



Articles

PhD Student


Credits: None available.

Chronic unpredictable stress exacerbates allergic airway inflammation in mice
G. Dragunas*1,2,3, M.A. de Oliveira1, W. T. de Lima1, R. Gosens2,3, C.D. Munhoz1
 
1- Department of Pharmacology, Institute of Biomedical Sciences, University of São Paulo
2- Department of Molecular Pharmacology, University of Groningen
3 - Groningen Research Institute for Asthma and COPD (GRIAC), University Medical Center Groningen (UMCG), University of Groningen
 
Introduction
It is accepted that psychological stress can lead to asthma exacerbations (1). Amid COVID-19 pandemic, exposure to chronic and, hence, deleterious forms of psychological stress has become usual. Stress can be classically defined as a real or potential threat to one’s homeostasis, generating physiological responses, such as HPA and SNS axis activation (2). Studies in the literature are widely in agreement that stress induces neuroplastic changes in psychiatric disorders and some suggest that this might also happen in conditions as asthma (3,4). However, the knowledge concerning how stress increases asthma severity and if neuronal mechanisms play a role are scarce.
 
Methods
We applied a 12 days chronic unpredictable stress (CUS) paradigm in OVA sensitized mice followed by two daily OVA challenges to induce allergic airway inflammation. 24h after the last challenge, mice had lung functional parameters analyzed, were euthanized, bronchoalveolar lavage collected and the lungs and dorsal root ganglia (DRG) harvested. The tissues were submitted to histological and molecular assays.
 
Results
Exposure to 12 day-CUS increased cellular content recovered in BAL. This was paired to increased p65 NF-kB phosphorylation, TRPV1 and P2X3 receptors expression in DRG, but not in the lungs. Exposure to CUS before acute challenge to two OVA aerosol challenges significantly increased recovered cells in BAL. Opposite outcomes were observed after a single acute restraint stress (RS), as reduced cellularity in BAL and diminished airway resistance to methacholine. OVA+CUS group displayed increased NF-kB signaling and VCAM expression in the lungs.
 
Conclusions
Exposure to chronic stress can lead to allergic airway inflammation exacerbation in mice, whereas previous acute stress led to inflammation mitigation. Future experiments will determine differential cytokine and neurotrophic factor expression in the lungs and changes in innervation in the airways in response to chronic stress.
 
References
1.          Chen E, Miller GE. Stress and inflammation in exacerbations of asthma. Brain Behav Immun. 2007;21(8):993–9.
2.          de Kloet ER, Joëls M, Holsboer F. Stress and the brain: from adaptation to disease. Nat Rev Neurosci. 2005;6(6):463–75.
3.          Dragunas G, Woest ME, Nijboer S, Bos ST, van Asselt J, de Groot AP, et al. Cholinergic neuroplasticity in asthma driven by TrkB signaling. FASEB J. 2020;34(6):7703–17.
4.          Undem BJ, Taylor-Clark T. Mechanisms underlying the neuronal-based symptoms of allergy. J Allergy Clin Immunol. 2014;133(6):1521–34.
 

Author(s):

PhD student


Credits: None available.

Epidermal growth factor receptor in airway remodeling during allergic airway disease – divergent roles during early life and adulthood?

H. Stölting, S. A. Walker, M. C. Zarcone, F. Puttur, S. Saglani, C. M. Lloyd
National Heart and Lung Institute, Imperial College London - London (United Kingdom)

Airway remodelling is a key pathological feature of paediatric and adult asthma, but the underlying mechanisms remain poorly understood. However, their elucidation is crucial, since lung function deficits established in children with asthma persist into adulthood. Epidermal growth factor receptor (EGFR) was shown to be overexpressed in paediatric and adult asthmatics. In addition, several in vivo studies using rodent models of allergic airway disease (AAD) have described a role for EGFR signalling in driving impaired lung function and airway remodelling in adult animals. Here, we aimed to study the role of EGFR in early life AAD. 
Bronchial epithelial cells from non-asthmatic children cultured at air-liquid interface were shown to exhibit high mRNA levels for EGFR and its ligands. Exposure of these cells to the allergen house dust mite induced EGFR activation dose-dependently, as measured by Y1068 phosphorylation. qPCR analysis of flow-sorted murine lung cell populations during postnatal development similarly showed high EGFR expression in murine lung epithelial cells from neonatal and adult mice, and lung epithelial EGFR expression was confirmed by flow cytometry. Finally, a pharmacological inhibitor was used to block EGFR signalling in a neonatal model of AAD. Preliminary findings indicate that EGFR inhibition in neonatal mice resulted in worsened lung function, as measured by a 2-fold increase in airway resistance (AUC), without affecting overall inflammation, a finding we did not observe in a corresponding adult AAD model.
These results indicate that EGFR is present in lungs at all stages of life and that, in contrast to its widely described pathogenic contribution to airway remodelling of adult animals, signalling through EGFR may play a protective role during early life AAD. 

Author(s):

Senior Research Scientist, Professor


Credits: None available.

Cannabis compounds have both anti-inflammatory and pro-inflammatory activities in lung epithelial and macrophages while substantially increasing phagocytosis in vitro

Seegehalli M Anil1+, Nurit Shalev1+, Ajjampura C Vinayaka1+, Stalin Nadarajan1+, Dvory Namdar1, Eduard Belausov1, Irit Shoval2, Karthik Ananth Mani3, Guy Mechrez3, Hinanit Koltai1*
1 Institute of Plant Science, Agriculture Research Organization, Volcani Center, Rishon LeZion 7528809, Israel
2 The Mina & Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 5290002, Israel.
3 Institute for Postharvest and Food Science, Agriculture Research Organization, Volcani Center, Rishon LeZion 7528809, Israel
 
+ These authors contributed equally to this work
Cannabis sativa is used worldwide for medical purposes and is known to have anti-inflammatory activity, yet the potential for use of C. sativa compounds against Coronavirus disease 2019 (COVID-19)-like inflammation is unexplored. The purpose of this study was to examine the anti-inflammatory activity of cannabis on markers of immune responses associated with COVID-19 inflammation. An extract fraction from high cannabidiol (CBD) cannabis strain (FCBD) substantially reduced dose dependently interleukin 6 (IL-6) and interleukin-8 (IL-8) levels in an alveolar epithelial (A549) cell line. FCBD contained CBD, cannabigerol (CBG) and tetrahydrocannabivarin (THCV), and multiple terpenes. Treatments with FCBD and phytocannabinoid standards that compose FCBD (FCBD:std) reduced in a dose dependent way IL-6, IL-8, C-C Motif Chemokine Ligands (CCLs) 2 and 7 in the A549 cell line. It also reduced expression of angiotensin I converting enzyme 2 (ACE2), a receptor for SARS-CoV-2. Treatment with FCBD induced macrophage (differentiated KG1 cell line) polarization and phagocytosis in vitro, and increased expression of scavenger receptor CD36 and that of type II receptor for the Fc region of IgG (FcγRII). FCBD treatment also substantially increased IL-6 and IL-8 expression in macrophages. FCBD:std, while maintaining the anti-inflammatory activity in alveolar epithelial cells, led to reduced pro-inflammatory IL secretion in macrophages in comparison to FCBD and reduced level of phagocytosis. The phytocannabinoid mixture may show superior activity for reduction of lung inflammation over that of the cannabis fraction. Yet, as for now, users and healthcare personnel should avoid the use of cannabis for COVID-19 prevention or treatment.

Author(s):

Pediatric Pulmonary Fellow


Credits: None available.

Multicohort Analysis of Bronchial Epithelial Cell Gene Expression Classifies Asthma from Healthy

Authors: Ian Lee1,2,3, Ananthkrishnan Ganesan1,2, Purvesh Khatri1,2,*

1Institute for Immunity, Transplantation and Infection, School of Medicine, Stanford University, CA 94305
2 Center for Biomedical Informatics Research, Department of Medicine, School of Medicine, Stanford University, CA 94305
3 Stanford Pediatric Pulmonary Medicine, Stanford University, CA 94303

Asthma is a heterogeneous disease with variable clinical manifestations including wheezing, shortness of breath, cough, and airflow limitation varying over time. Previous transcriptome studies of airway epithelial cells in asthma compared to controls have identified hundreds of differentially expressed genes and characterized “T-helper cell 2 (Th2)-high” and “Th2-low” endotypes of asthma inflammation.

We hypothesized that integrating gene expression profiles of bronchial epithelial cells from patients with asthma across multiple studies would identify a robust gene signature that represents real-world biological and clinical heterogeneity in patients with asthma. We identified six data sets containing 486 whole transcriptome profiles of bronchial epithelial cells (BECs) from healthy controls (HCs) and patients with asthma of varying severity from at least four clinical centers in two countries. We arbitrarily chose four data sets comprised of 223 samples (HC=97, mild/moderate asthma=82, severe asthma=44) as the discovery data sets, and the remaining two consisting of 263 samples (HC=47, mild/moderate asthma=122, severe asthma=97) as the validation data sets. We calculated a Hedges’ g effect size (ES) for each gene. We applied leave-one-study-out analysis to avoid influence of a single data set, and used random effects inverse variance-based meta-analysis to integrate ES for each gene across the discovery data sets into a summary ES.

Using stringent selection criteria (FDR < 1%, absolute effect size > 0.6), we found 10 genes significantly differentially expressed between patients with asthma and healthy controls, including 5 over-expressed (POSTN, SERPINB2, TPRXL, CLCA1, CEACAM5) and 5 under-expressed (ACKR3, SCGB3A1, GMNN, CYP2A13, CNTD1) genes without between dataset heterogeneity.

We defined the asthma score of a sample as the difference between the geometric mean of over-expressed genes and that of the under-expressed genes. This simple classifier distinguished patients with asthma from healthy controls with an average area under the receiver operating characteristic (AUROC) curve of 0.87 (range: 0.79-0.91). Next, we validated this signature in the two independent data sets, where the asthma score distinguished patients with asthma from healthy controls with AUROC of 0.79 and 0.81. Across all discovery and validation cohorts, the asthma score increased with severity of asthma and was significantly positively correlated with it (Jonckheere-Terpstra trend test p-value < 0.05). We also observed a bimodal distribution of our asthma score in some data sets, which suggests the existence of endotypes of asthma that are not well classified by this gene signature and should be investigated further.

Our analysis identified a parsimonious gene set that distinguishes patients with asthma from a heterogeneous group of controls, including allergic rhinitis and former smokers, with high accuracy, and is positively correlated with severity of asthma. This could suggest a shared pathway despite the heterogeneity observed in patients with asthma. 

Author(s):

Research Associate Professor


Credits: None available.

Asthma-associated variants induce IL33 differential expression through a novel regulatory region

Ivy Aneas1, Donna C. Decker2, Débora R. Sobreira1, Noboru J. Sakabe1, Kelly M. Blaine2, Kevin M. Magnaye1, Selene M. Clay1, Carole Ober1, Anne I. Sperling2,3, Marcelo A. Nobrega1.
1- Department of Human Genetics, University of Chicago, Chicago, IL; 2- Department of Medicine, Section of Pulmonary and Critical Care Medicine, University of Chicago, Chicago, IL, 3- Committee on Immunology, University of Chicago, Chicago IL
Genome-wide association studies (GWAS) have implicated the IL33 locus in asthma, but the underlying mechanisms remain unclear. Here, we identify a 5 kb region within the GWAS-defined segment that acts as a strong regulatory element in vivo and in vitro. Chromatin conformation capture showed that this 5 kb region loops to the IL33 promoter, potentially regulating its expression. We show that genotype at the asthma-associated SNP rs1888909, located within the 5 kb region, is associated with IL33 gene expression in human airway epithelial cells and IL-33 protein expression in human plasma, potentially through differential binding of OCT-1 (POU2F1) to the asthma-risk allele. Our data demonstrate that asthma-associated variants at the IL33 locus mediate allele-specific regulatory activity and IL33 expression, providing a novel mechanism through which a regulatory SNP contributes to genetic risk of asthma.

Author(s):

Research associate


Credits: None available.

Evidence for an alternative IL13/IL13R mediated allergen response in lung in the absence of ILC2 cells and IL5

Jennifer Fraszczak*, Thannina Hamadou and Tarik Möröy*#&

* Institut de recherches cliniques de Montréal, québec, Canada
# Département de microbiologie, infectiologie et immunologie, Faculty of Medicine, Université de Montréal, Montreal, Canada
& Division of Experimental Medicine, McGill University, Montreal, Canada

Asthma is an inflammatory disease of the bronchial airways and is mainly caused by environmental factors such as air pollution and allergens. The development of resistance to existing therapies and increasing air pollution and exposure to allergens have made asthma a major health problem. The molecular mechanisms underlying an asthmatic reaction in the lung caused by airborne allergens are not well understood, but recent experimental evidence exists indicating that this reaction is mainly driven by type 2 innate lymphoid cells (ILC2) and CD4+ type 2 T helper cells (Th2). These cells support allergic reactions by producing cytokines such as IL4 or IL5 leading to eosinophilia. It has been shown that GFI1, a zinc finger transcription factor involved in hematopoiesis and inflammation, regulates many cells involved in the Th2 type immune response including ILC2 or Th2 T cells. Although Gfi1 deficient mice lack functional ILC2 and Th2 T cells and fail to produce IL5, they still show a robust response to allergens with features such as increased expression of mucus-associated genes and lung fibrosis. In particular, GFI1 KO mice still have an accumulation of  CD11c+SiglecFhigh eosinophils to the lungs in response to allergen. In addition, Gfi1 KO mice have a higher expression of Il13 mRNA in the lungs after allergen stimulation and Gfi1 null CD11c+SiglecFhigh eosinophils express Il13ra1. These data suggests that GFI1 KO mice can adapt their response to the allergen by using a different mechanism than WT mice. These findings may have implications for the currently used allergy therapies using anti-IL5 antibodies and may explain the occurrence of resistance towards this treatment.    

Author(s):

Postdoc


Credits: None available.

Pre-existing asthma does not enhance cytokine response and disease severity of COVID-19

Background: Both COVID-19 and asthma involve pulmonary inflammation, damage and dysfunction, and can result in respiratory failure and death.(1) A significant portion of COVID-19 sufferers have asthma comorbidity,(2) however, the impact of COVID-19 infections on patients with asthma is still unclear. In this study, we investigated the roles of pre-existing asthma as a comorbidity in disease severities of COVID-19 by measuring immune response in circulating cytokines.

Methods: Plasma samples and clinical information were collected from patients (total 80) with mild (25), severe (36) or critical (19) cases of COVID-19 at the John Radcliffe Hospital, Oxford, UK. The concentrations of 51 proteins in the plasma samples were measured with Luminex Kits (Bio-techne) using a Bio-Rad Bio-Plex® 200 Systems.

Results: A total of 16 pre-existing asthma patients were found (3 in mild, 10 in severe, and 3 in critical COVID-19).
In the comparison of circulating cytokines enrichment, no significant difference between COVID-19 patients with and without asthma was found in different disease severities and age groups. Indeed, a trend of slightly lower levels of CXCL10, CCL2, and IL-8 were observed in patients with asthma.
In comparing clinical traits within the same COVID-19 severity group, also no differences were observed between patients with or without asthma in terms of oxygen level, CRP, neutrophil counts, and length of hospital stay. Although the age distribution showed a slight increase with severity of COVID-19 (37.33±17.04 in mild, 55.60±16.12 in severe and 70.00±11.36 in critical) in the patients with asthma but not in the patients without asthma (69.91±20.15 in mild, 66.69±15.51 in severe and 55.38±12.47 in critical), the prevalence of asthma in COVID-19 severity groups (12.0% in mild, 27.8% in severe and 15.8% in critical) did not exhibit a clear correlation between asthma and the severities of COVID-19. Furthermore, the mortality ratio in the COVID-19 patients with asthma (12.5%) was not higher than that in patients without asthma (17.2%).

Conclusions: Based on this cohort study, pre-existing asthma was not associated with an enhanced cytokine storm after COVID-19 infection, and did not have strong effects on COVID-19 progression.

References
1. Menter T, et al. Postmortem examination of COVID-19 patients reveals diffuse alveolar damage with severe capillary congestion and variegated findings in lungs and other organs suggesting vascular dysfunction. Histopathology. 2020 May 4.
2. Docherty AB, et al. Features of 20133 UK patients in hospital with covid-19 using the ISARIC WHO Clinical Characterisation Protocol: prospective observational cohort study. BMJ. 2020;369:m1985.

Author(s):

Chief, Division of Allergy and Clinical Immunology


Credits: None available.

Leukotriene E4 Rapidly Amplifies Airway Type 2 Inflammation Through IL-25 and Endogenous Cysteinyl Leukotrienes

Tao Liu 1,2, Lora G. Bankova1,2, Nora A. Barrett1,2, Chunli Feng1, Junrui Lin1, Juying Lai1, and Joshua A. Boyce 1,2,3
1Division of Allergy and Clinical Immunology, Brigham and Women’s Hospital
Departments of 2Medicine and 3Pediatrics, Harvard Medical School
Boston, MA 02115
Abstract
Leukotriene E4 (LTE4), the stable metabolite of cysteinyl leukotrienes (cysLTs), accumulates in asthma, aspirin exacerbated respiratory disease (AERD) and chronic rhinosinusitis. Although it is the weakest direct bronchoconstrictor of the three cysLTs, LTE4 is disproportionately potent in the airways of subjects with asthma and AERD, and elicits both bronchial eosinophilia and mast cell (MC) activation when administered to asthmatic subjects by inhalation. We previously reported that LTE4 elicited expansion of IL-25-expressing tracheal brush cells (BrCs) and mild airway eosinophilia in naïve mice by a pathway requiring the type 3 cysteinyl leukotriene receptor (CysLT3R). We now report that LTE4 rapidly amplifies features of type 2 immunopathology by a pathway requiring all three cysLT receptors, and sequential actions of IL-25, IL-33, and endogenously generated LTC4.  A single inhaled dose of LTE4 administered to AERD-like Ptges-/- mice elicited rapid increases in lung ILC2s, PAS+ goblet cells, and DLCK1+ brush cells in the trachea.  LTE4 caused the rapid recruitment of platelet-adherent eosinophils to the lung, as well as sharp increases in airway resistance, release of mast cell activation products, and lung levels of IL-33. Deletion of CysLT3R, antibody neutralization of IL-25 and pharmacologic blockades of either CysLT1R or CysLT2R attenuated all features. Thus, LTE4 drives a potent innate type 2 response initiated by the bush cell products LTC4 and IL-25,  amplified by downstream ILC2 activation, and culminates in platelet-adherent eosinophil recruitment and platelet-dependent IL-33-driven mast cell activation. These findings suggest several avenues for therapeutic development, especially in AERD where the levels of LTE4 are especially high. 

Author(s):

Assistant Professor of Medicine, Medical Director of Clinical Asthma Research


Credits: None available.

Glucagon-like Peptide-1 Receptor Agonists decrease type-2 biomarker in asthma

Dinah Foer1,2, Patrick Beeler3, Jing Cui1,2, Joshua A. Boyce1,2, Elizabeth Karlson1,2, David W. Bates1,2, Katherine Cahill4
1Brigham and Women's Hospital, 2Harvard Medical School, Boston, MA, 3University Hospital of Zurich, Zurich, Switzerland, 4Vanderbilt University Medical Center, Nashville, TN.

Glucagon-like peptide-1 receptor agonists (GLP-1RA) are approved for the treatment of type II diabetes mellitus (DMII) and obesity. In murine models, GLP-1RA inhibit allergen- and viral-induced airway inflammation including airway interleukin (IL)-33 release, IL-13 and mucus production and hyperresponsiveness. Observational patient data supports an association between GLP-1RA use and decreased asthma exacerbations in patients with asthma and DMII. We hypothesized that GLP-1RA would decrease biomarkers pertinent to airway inflammation pathways in patients with asthma. To test this hypothesis, we obtained serum samples from adults with asthma and comorbid DMII in the Partners HealthCare Biobank treated with (N=43) or without a GLP-1RA (N=119) at the time of sample collection. Demographics, body mass index, asthma severity, glucose control, and comorbidities, confirmed by electronic health record chart review were extracted and a propensity score calculated based on GLP-1RA use included as a covariate in a logistic regression model. Serum periostin, total IgE, IL-6, IL-8, IL-33 and sST2 levels were measured. Periostin (Padi=.0006) was significantly decreased in GLP-1RA users than non-GLP-1RA users which included use of basal insulin, SGLT-2 inhibitors or sulfonylureas. There were no significant differences in total IgE (Padj =.12), IL-6 (Padj =.62), IL-8 (Padj =.41), sCD163 (Padj=.53), IL-33 (Padj =.91), and sST2 (Padj =.90). Periostin results were robust across asthma severity and gender subgroup analyses. Serum periostin, a known systemic biomarker of Type (T)2 cytokines IL-4 and IL-13 in airway inflammation, is significantly and selectively decreased in adults with asthma and comorbid DMII treated with GLP-1RAs. Our results support a role for GLP-1R signaling in airway inflammation and point to periostin as a possible biomarker for therapeutic use of GLP-1RAs in asthma.

Funding: NIH AI118804, Brigham and Women’s Hospital Department of Medicine Innovation Evergreen Award, Brigham Research Institute Pilot Award

Author(s):

Pediatrician


Credits: None available.

Attitudes, practices and knowledge of Macedonian primary health care physicians and pediatricians regarding Buteyko breathing technique as complementary method of asthma treatment

Authors:
1. Adrijana Ugrinoska Pandeva - Department for chronic respiratory diseases in children above 3 years, Institute for respiratory diseases in children Kozle, Skopje, Republic of North Macedonia
2. Aneta Lazarova - Primary, preventive and dental care centre " Gazi Baba, Public Health institute - Health Center Skopje, Republic of North Macedonia

Background: Asthma is a chronic inflammatory disease characterized by an obstruction of airflow which results in difficulty in breathing. Asthma affects 5-10% of the population. In adition to pharmacologic management many patients use complementary therapies. The Buteyko Breathing Technique (BBT) is distinctive breathing therapy that uses control breath and breath-holding exercises.

Aim: of this study is to evaluate the knowledge, attitudes and practices of primary health care physicians and pediatricians regarding Buteyko breathing technique as complementary method of asthma treatment.

Methods : The survey was conducted in September - October 2020 with the distribution of an anonymous survey questionnaire electronically in the form of Google document to 74 primary care physicians and pediatricians who work in various inpatient and outpatient units.  

Results : 42 respondents answered the questionnaire. The largest percentage of respondents are aged 36-45 y with 10-25 years of work experience. 78,6% work with patients with chronic respiratory diseases. 38,1% of patients use alternative medicine for asthma management, mostly homeopathy, yoga and phytothetapy.  Approximately half of physicians recommend a breathing technique most often yoga breathing. 21,4% of doctors know about Buteyko breathing technique and 7,1% practice that technique. Education for BBT was welcomed by most physicians ( 95,2%).

Conclusions : Macedonian primary health care physicians and pediatrician are familiar with some alternative therapies and breathing technique for relieving asthma symptoms, but only a few now about Buteyko breathing method.  There are educational needs for  Buteyko breathing technique in clinical practice.

Author(s):