[BAD] EK9 - Abstracts



Articles

PhD Student


Credits: None available.

Itaconate regulates responses to inhaled aeroallergen

Gesa J. Albers1, Patricia P. Ogger1, Simone A. Walker1, John M. Halket2, Robert Gray2, Clare M. Lloyd1, Adam Byrne1
1National Heart and Lung Institute, Imperial College London, UK
2Mass Spectrometry Facility, King’s College London, UK

Background: Asthma is a chronic disease characterised by airway remodelling and hyperresponsiveness, mucus production and inflammation. Airway macrophages (AMs) are key sentinels of lung homeostasis and form the first line of defence against inhaled allergen. Increasing evidence suggests that changes in AM phenotype are underpinned by alterations in AM metabolism. The TCA cycle-derived metabolite itaconic acid (IA), synthesised by the enzyme aconitate decarboxylase (ACOD)-1, is a key regulator of macrophage function. Recently, we showed in human and murine models that IA limits pulmonary fibrosis, yet its role in regulating pulmonary responses to inhaled allergen is unknown. Here, we aimed to study the role of IA in murine models of allergic airway disease (AAD).
Methods: To characterise the kinetics of the Acod1/IA pathway in response to allergen, we measured lung Acod1 expression and BAL-IA levels after house dust mite (HDM) challenge. Next, we assessed Acod1 expression in murine AMs exposed to HDM ex vivo. Finally, to determine the role of IA in allergic airway responses, we treated WT or Acod1-/- mice with inhaled HDM and assessed disease pathology and inflammation.
Results: Continuous HDM exposure in mice resulted in augmented levels of BAL-IA and increased expression of lung Acod1. Ex vivo culture of AMs with HDM revealed enhanced Acod1 expression and a shift towards a more glycolytic phenotype. Finally, exposure of Acod1-/- mice to allergen led to an increased neutrophil-to-eosinophil ratio, compared to more eosinophilic WT controls, which was rescued by inhaled IA.
Conclusion: Our data indicate that the Acod1/IA pathway is highly induced during AAD and regulates the balance between neutrophil and eosinophil recruitment in response to inhaled allergen.

Author(s):

Assistant Professor


Credits: None available.

Modeling airway dysfunction in asthma using synthetic mucus biomaterials

As asthma worsens, occlusion of airways with mucus significantly contributes to airflow obstruction and reduced lung function. Recent evidence from clinical studies has shown mucus obtained from adults and children with asthma possesses altered mucin composition. However, how these changes alter the functional properties of the mucus gel is not yet fully understood. To study this, we have engineered a synthetic mucus biomaterial to closely mimic the properties of native mucus in health and disease. We demonstrate this model possesses comparable biophysical and transport properties to native mucus ex vivo collected from human subjects and in vitro isolated from human airway epithelial (HAE) tissue cultures. We found by systematically varying mucin composition that mucus gel viscoelasticity is enhanced when predominantly composed of mucin 5AC (MUC5AC), as is observed in asthma. As a result, asthma-like synthetic mucus gels are more slowly transported on the surface of HAE tissue cultures and at a similar rate to native mucus produced by HAE cultures stimulated with the type 2 cytokine IL-13, known to contribute to airway inflammation and MUC5AC hypersecretion in asthma. We also discovered the barrier function of asthma-like synthetic mucus towards influenza A virus was impaired as evidenced by the increased frequency of infection in MUC5AC-rich hydrogel coated HAE cultures. Together, this work establishes a biomaterial-based approach to understand airway dysfunction in asthma and related muco-obstructive lung diseases.

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):

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):

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):

Assistant Professor


Credits: None available.

Asthma and comorbidities, a harmful association, especially during COVID-19
Leila Laouar1,2,*, Sarah Boukellal2, Ali Adib Yaici2, Med Taib Makhloufi1,2, and Samya Taright1,2
1University Benyoucef Benkhedda, School of Medicine, Algiers, Algeria.
2Department of Pulmonary Diseases, Center Hospitalo-Universitaire Mustapha Pasha, Algiers, Algeria
* Correspondence should be addressed to: laouar_leila@yahoo.fr
 
Asthma management cannot depend solely on medication; instead, it requires the control of environmental exposures as well as the comorbidities that have a poor impact on the overall prognosis. Among these comorbidities, cardiovascular manifestations and metabolic disorders such as diabetes and obesity, are reminiscent of the risk factors underlying COVID-19 severity. Management of such comorbidities is critical in asthmatic patients during a pandemic, and particularly during the confinement period in which sedentary lifestyle, depression and eating disorders significantly intensify, contributing to weight gain and induction of cardiovascular diseases in asthmatic patients. The goal from this study is to determine the frequency of comorbidities and their impact on asthma control. To this aim, we have conducted a study using a cohort of 51 asthmatic patients monitored in the Tuberculosis and Chronic Respiratory Diseases Control Unit at Mustapha Pasha Hospital in Algiers, Algeria. In agreement with data studies published in the literature, we observed that women are at significantly increased risk of developing asthma with a male to female ratio of 0.31 and a median patient age of 58 years. Our data revealed a poor control of asthma in 35% of our cohort patients compared to 25-65% of cases reported in the literature, but a higher frequency of comorbidities in 88.2% of our patients compared to 15-86% of cases reported in the literature. Specifically, we report the following medical conditions in our cohort patients: 30% obesity (among obese patients, 47% exhibited poor asthma control); 60% allergic rhinitis (among patients with rhinitis, 25% exhibited poor control); 27% gastroesophageal reflux disease or GERD (among patients with GERD, 35% exhibited poor control); 30% hypertension (among hypertensive patients, 38% exhibited poor control); and 12% type 2 diabetes (among diabetic patients, 54% exhibited poor control). It should be noted that asthma control is largely conditioned by the proper use of the inhalation device, which remains a problem for the majority of asthmatic patients. However, this is not the case for our cohort patients, since most of them (> 75%) were properly trained for the use of such a medical device. Finally, in agreement with data studies published in the literature, we observed that smoking -whether passive or active- caused further exacerbations of asthma with lesser disease control. Although asthma is not among the top conditions associated with COVID-19 deaths (unlike cardio-metabolic disorders which are more frequently reported in severe COVID-19 cases); we cannot rule out the possibility that infection by COVID-19 could be responsible for an exacerbation of asthma, in particular in the presence of comorbidities which are reminiscent of the risk factors of severity and mortality by COVID-19. In this regard, recent published data on Algerian patients hospitalized for severe forms of COVID-19 reported a frequency of 8% of asthmatics among COVID-19 patients. Notably, these COVID-19 asthmatics suffered from a severe form of asthma associated with cardiovascular comorbidities. Therefore, achieving an optimal level of asthma control should be among the primary goals to attenuate COVID-19 severity. However, such a control is arduous due to the many associated factors involved in a cause-effect relationship with asthma including allergic rhinitis, GERD and obesity, in addition to the most common comorbidities such as diabetes. With all data considered, it remains unsolved whether the reported severity of COVID-19 in asthmatics is due to asthma manifestation itself or is the result of its underlying comorbidities. Future studies in this direction will elucidate the bias of causal links of asthma with certain comorbidities in COVID-19 patients.

Author(s):

Assistant Professor


Credits: None available.

Obesity dysregulates immunometabolic status in pediatric asthma and impacts vaccine responses
Sarah E. Henrickson1,2, Peyton Conrey2, Sasikanth Manne1,3, , Samir Sayed2, Kaitlin C. O’Boyle3 Bertram Bengsch1,†  , Ting Qian4, Ramin S. Herati1,5††­­†, Laura A. Vella1,6, Allison R. Greenplate1,3, Sam J.  McCright1,7, Cécile Alanio1,3, 12, Frank Mentch11, Kenneth E. Schmader8, Christopher F. Pastore9, Li-Yin Hun9, Scott E. Hensley1,10, De’Broski Herbert9, Aaron J. Masino4, Jorge Henao-Mejia1,7, Hakon Hakonarson11, Joshua D. Rabinowitz12, Susan E. Coffin6 and E. John Wherry1,3,12 
1Institute for Immunology, University of Pennsylvania, Philadelphia, PA.
2Division of Allergy-Immunology, Department of Pediatrics, Children’s Hospital of Philadelphia, Philadelphia, PA.
3Department of Systems Pharmacology and Translational Therapeutics, University of Pennsylvania, Philadelphia, PA.
4Department of Biomedical and Health Informatics, The Children’s Hospital of Philadelphia, Philadelphia, PA.
5Department of Medicine, University of Pennsylvania Perelman School of Medicine 
6Division of Infectious Disease, Department of Pediatrics, The Children’s Hospital of Philadelphia, Philadelphia, PA. 
7Department of Pathology, The Children’s Hospital of Philadelphia, Philadelphia, PA.
8Division of Geriatrics, Department of Medicine, Duke University Medical Center and Geriatric Research, Education, and Clinical Center, Durham VA Medical Center, Durham, NC.
9School of Veterinary Medicine, Department of Pathobiology, University of Pennsylvania, Philadelphia. PA
10Department of Microbiology, University of Pennsylvania, Philadelphia, PA.
11Center for Applied Genomics, Department of Pediatrics, The Children’s Hospital of Philadelphia, Philadelphia, PA.
12Parker Institute for Cancer Immunotherapy at University of Pennsylvania
13Department of Chemistry, Princeton University, Princeton, NJ.
†1Department of Medicine II, Gastroenterology, Hepatology, Endocrinology, and Infectious Diseases, University Medical Center Freiburg, Faculty of Medicine, Freiburg, Germany, and Signalling Research Centres BIOSS and CIBSS, University of Freiburg, Freiburg, Germany
††Department of Medicine, New York University, Grossman School of Medicine, New York City, NY.
 
Asthma and obesity are two of the most common chronic childhood diseases worldwide, with dramatically increasing prevalence over the last few decades. These diseases impact morbidity and mortality and strain health care systems financially. Asthma risk increases as body mass index (BMI) increases, suggesting a pathophysiological link. Both asthma and obesity are independently linked to altered immune status, however, it remains unclear how these diseases converge to affect pediatric immune function. To address this question, we investigated the immunometabolic profile in obese asthmatic (OA), non-obese asthmatic (A), obese non-asthmatic (O), and healthy control (HC) children using mass cytometry, serum metabolomics, cytokine analysis and clinical history. This multi-modal approach revealed two major forms of immune dysfunction in pediatric allergic OA: altered baseline T cell activation state (exhaustion-like) and increased type 2 immunity. OA had increased Th2 differentiation and decreased Th17 differentiation and these changes were associated with altered blood metabolites, including increased glutamate and decreased acetate. A mouse model of OA confirmed increased exhausted-like CD8 T cells compared to A and HC mice. Finally, immunometabolic dysregulation and altered T cell activation status in O and OA patients was linked to prolonged retention of humoral vaccine responses. These insights into the mechanistic links between metabolic alterations and immune dysfunction in OA may improve understanding of the severe asthma exacerbations secondary to viral upper respiratory tract infections seen in OA and provide opportunities for novel therapeutic approaches.

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