ORCID Profile
0000-0003-2429-4165
Current Organisation
McMaster University
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Publisher: European Respiratory Society (ERS)
Date: 10-2020
DOI: 10.1183/23120541.00128-2020
Abstract: Cannabis smoking is the dominant route of delivery, with the airway epithelium functioning as the site of first contact. The endocannabinoid system is responsible for mediating the physiological effects of inhaled phytocannabinoids. The expression of the endocannabinoid system in the airway epithelium and contribution to normal physiological responses remains to be defined. To begin to address this knowledge gap, a curated dataset of 1090 unique human bronchial brushing gene expression profiles was created. The dataset included 616 healthy subjects, 136 subjects with asthma, and 338 subjects with COPD. A 32-gene endocannabinoid signature was analysed across all s les with sex and disease-specific analyses performed. Immunohistochemistry and immunoblots were performed to probe in situ and in vitro protein expression. CB 1 , CB 2 , and TRPV1 protein signal is detectable in human airway epithelial cells in situ and in vitro , justifying examining the downstream endocannabinoid pathway. Sex status was associated with differential expression of 7 of 32 genes. In contrast, disease status was associated with differential expression of 21 of 32 genes in people with asthma and 26 of 32 genes in people with COPD. We confirm at the protein level that TRPV1, the most differentially expressed candidate in our analyses, was upregulated in airway epithelial cells from people with asthma relative to healthy subjects. Our data demonstrate that the endocannabinoid system is expressed in human airway epithelial cells with expression impacted by disease status and minimally by sex. The data suggest that cannabis consumers may have differential physiological responses in the respiratory mucosa.
Publisher: European Respiratory Society (ERS)
Date: 10-12-2021
DOI: 10.1183/23120541.00705-2020
Abstract: Accessible in vitro models recapitulating the human airway that are amenable to study whole cannabis smoke exposure are needed for immunological and toxicological studies that inform public health policy and recreational cannabis use. In the present study, we developed and validated a novel three-dimensional (3D)-printed in vitro exposure system (IVES) that can be directly applied to study the effect of cannabis smoke exposure on primary human bronchial epithelial cells. Using commercially available design software and a 3D printer, we designed a four-chamber Transwell insert holder for exposures to whole smoke. COMSOL Multiphysics software was used to model gas distribution, concentration gradients, velocity profile and shear stress within IVES. Following simulations, primary human bronchial epithelial cells cultured at the air–liquid interface on Transwell inserts were exposed to whole cannabis smoke using a modified version of the Foltin puff procedure. Following 24 h, outcome measurements included cell morphology, epithelial barrier function, lactate dehydrogenase (LDH) levels, cytokine expression and gene expression. Whole smoke delivered through IVES possesses velocity profiles consistent with uniform gas distribution across the four chambers and complete mixing. Airflow velocity ranged between 1.0 and 1.5 µm·s −1 and generated low shear stresses ( Pa). Human airway epithelial cells exposed to cannabis smoke using IVES showed changes in cell morphology and disruption of barrier function without significant cytotoxicity. Cannabis smoke elevated interleukin-1 family cytokines and elevated CYP1A1 and CYP1B1 expression relative to control, validating IVES smoke exposure impacts in human airway epithelial cells at a molecular level. The growing legalisation of cannabis on a global scale must be paired with research related to potential health impacts of lung exposures. IVES represents an accessible, open-source, exposure system that can be used to model varying types of cannabis smoke exposures with human airway epithelial cells grown under air–liquid interface culture conditions.
Publisher: BMJ
Date: 16-11-2015
DOI: 10.1136/THORAXJNL-2015-207399
Abstract: Traffic-related air pollution has been shown to augment allergy and airway disease. However, the enhancement of allergenic effects by diesel exhaust in particular is unproven in vivo in the human lung, and underlying details of this apparent synergy are poorly understood. To test the hypothesis that a 2 h inhalation of diesel exhaust augments lower airway inflammation and immune cell activation following segmental allergen challenge in atopic subjects. 18 blinded atopic volunteers were exposed to filtered air or 300 µg PM 2.5 /m 3 of diesel exhaust in random fashion. 1 h post-exposure, diluent-controlled segmental allergen challenge was performed 2 days later, s les from the challenged segments were obtained by bronchoscopic lavage. S les were analysed for markers and modifiers of allergic inflammation (eosinophils, Th2 cytokines) and adaptive immune cell activation. Mixed effects models with ordinal contrasts compared effects of single and combined exposures on these end points. Diesel exhaust augmented the allergen-induced increase in airway eosinophils, interleukin 5 (IL-5) and eosinophil cationic protein (ECP) and the GSTT1 null genotype was significantly associated with the augmented IL-5 response. Diesel exhaust alone also augmented markers of non-allergic inflammation and monocyte chemotactic protein (MCP)-1 and suppressed activity of macrophages and myeloid dendritic cells. Inhalation of diesel exhaust at environmentally relevant concentrations augments allergen-induced allergic inflammation in the lower airways of atopic in iduals and the GSTT1 genotype enhances this response. Allergic in iduals are a susceptible population to the deleterious airway effects of diesel exhaust. NCT01792232.
Publisher: American Thoracic Society
Date: 08-2017
Publisher: American Thoracic Society
Date: 09-2005
Publisher: Public Library of Science (PLoS)
Date: 09-2017
Publisher: American Thoracic Society
Date: 09-2013
Publisher: Wiley
Date: 2009
DOI: 10.1111/J.1365-2222.2008.03109.X
Abstract: Asthma is a disease characterized by variable and reversible airway obstruction and is associated with airway inflammation, airway remodelling (including goblet cell hyperplasia, increased collagen deposition and increased smooth muscle mass) and increased airway responsiveness. It is believed that airway inflammation plays a critical role in the development of airway remodelling, with IL-13 and TGF-beta1 pathways being strongly associated with the disease progression. Mouse models of asthma are capable of recapitulating some components of asthma and have been used to look at both IL-13 and TGF-beta1 pathways, which use STAT6 and SMAD2 signalling molecules, respectively. Using brief and chronic models of allergen exposure, we utilized BALB/c and C57Bl/6 to explore the hypothesis that observed differences in responses to allergen between these mouse strains will involve fundamental differences in IL-13 and TGF-beta1 responses. The following outcome measurements were performed: airway physiology, bronchoalveolar lavage cell counts/cytokine analysis, histology, immunoblots and gene expression assays. We demonstrate in BALB/c mice an IL-13-dependent phosphorylation of STAT6, nuclear localized in inflammatory cells, which is associated with indices of airway remodelling and development of airway dysfunction. In BALB/c mice, phosphorylation of SMAD2 is delayed relative to STAT6 activation and also involves an IL-13-dependent mechanism. In contrast, despite an allergen-induced increase in IL-4, IL-13 and eosinophils, C57Bl/6 demonstrates a reduced and distinct pattern of phosphorylated STAT6, no SMAD2 phosphorylation changes and fail to develop indices of remodelling or changes in airway function. The activation of signalling pathways and nuclear translocation of signalling molecules downstream of IL-13 and TGF-beta1 further support the central role of these molecules in the pathology and dysfunction in animal models of asthma. Activation of signalling pathways downstream from IL-13 and TGF-beta1 may be more relevant in disease progression than elevations in airway inflammation alone.
Publisher: Springer Science and Business Media LLC
Date: 17-01-2019
DOI: 10.1038/S41598-018-36248-9
Abstract: ABC transporters are conserved in prokaryotes and eukaryotes, with humans expressing 48 transporters ided into 7 classes (ABCA, ABCB, ABCC, ABCD, ABDE, ABCF, and ABCG). Throughout the human body, ABC transporters regulate cAMP levels, chloride secretion, lipid transport, and anti-oxidant responses. We used a bioinformatic approach complemented with in vitro experimental methods for validation of the 48 known human ABC transporters in airway epithelial cells using bronchial epithelial cell gene expression datasets available in NCBI GEO from well-characterized patient populations of healthy subjects and in iduals that smoke cigarettes, or have been diagnosed with COPD or asthma, with validation performed in Calu-3 airway epithelial cells. Gene expression data demonstrate that ABC transporters are variably expressed in epithelial cells from different airway generations, regulated by cigarette smoke exposure ( ABCA13 , ABCB6 , ABCC1 , and ABCC3 ), and differentially expressed in in iduals with COPD and asthma ( ABCA13 , ABCC1 , ABCC2 , ABCC9) . An in vitro cell culture model of cigarette smoke exposure was able to recapitulate select observed in situ changes. Our work highlights select ABC transporter candidates of interest and a relevant in vitro model that will enable a deeper understanding of the contribution of ABC transporters in the respiratory mucosa in lung health and disease.
Publisher: AIP Publishing
Date: 28-04-2023
DOI: 10.1063/5.0134177
Abstract: Idiopathic pulmonary fibrosis (IPF) is a severe form of pulmonary fibrosis. IPF is a fatal disease with no cure and is challenging to diagnose. Unfortunately, due to the elusive etiology of IPF and a late diagnosis, there are no cures for IPF. Two FDA-approved drugs for IPF, nintedanib and pirfenidone, slow the progression of the disease, yet fail to cure or reverse it. Furthermore, most animal models have been unable to completely recapitulate the physiology of human IPF, resulting in the failure of many drug candidates in preclinical studies. In the last few decades, the development of new IPF drugs focused on changes at the cellular level, as it was believed that the cells were the main players in IPF development and progression. However, recent studies have shed light on the critical role of the extracellular matrix (ECM) in IPF development, where the ECM communicates with cells and initiates a positive feedback loop to promote fibrotic processes. Stemming from this shift in the understanding of fibrosis, there is a need to develop in vitro model systems that mimic the human lung microenvironment to better understand how biochemical and biomechanical cues drive fibrotic processes in IPF. However, current in vitro cell culture platforms, which may include substrates with different stiffness or natural hydrogels, have shortcomings in recapitulating the complexity of fibrosis. This review aims to draw a roadmap for developing advanced in vitro pulmonary fibrosis models, which can be leveraged to understand better different mechanisms involved in IPF and develop drug candidates with improved efficacy. We begin with a brief overview defining pulmonary fibrosis and highlight the importance of ECM components in the disease progression. We focus on fibroblasts and myofibroblasts in the context of ECM biology and fibrotic processes, as most conventional advanced in vitro models of pulmonary fibrosis use these cell types. We transition to discussing the parameters of the 3D microenvironment that are relevant in pulmonary fibrosis progression. Finally, the review ends by summarizing the state of the art in the field and future directions.
Publisher: Frontiers Media SA
Date: 24-11-2021
DOI: 10.3389/FBIOE.2021.773511
Abstract: Human lungs are organs with an intricate hierarchical structure and complex composition lungs also present heterogeneous mechanical properties that impose dynamic stress on different tissue components during the process of breathing. These physiological characteristics combined create a system that is challenging to model in vitro . Many efforts have been dedicated to develop reliable models that afford a better understanding of the structure of the lung and to study cell dynamics, disease evolution, and drug pharmacodynamics and pharmacokinetics in the lung. This review presents methodologies used to develop lung tissue models, highlighting their advantages and current limitations, focusing on 3D bioprinting as a promising set of technologies that can address current challenges. 3D bioprinting can be used to create 3D structures that are key to bridging the gap between current cell culture methods and living tissues. Thus, 3D bioprinting can produce lung tissue biomimetics that can be used to develop in vitro models and could eventually produce functional tissue for transplantation. Yet, printing functional synthetic tissues that recreate lung structure and function is still beyond the current capabilities of 3D bioprinting technology. Here, the current state of 3D bioprinting is described with a focus on key strategies that can be used to exploit the potential that this technology has to offer. Despite today’s limitations, results show that 3D bioprinting has unexplored potential that may be accessible by optimizing bioink composition and looking at the printing process through a holistic and creative lens.
Publisher: Wiley
Date: 04-04-2023
DOI: 10.1111/IMCB.12637
Abstract: Idiopathic pulmonary fibrosis (IPF) is a chronic, progressive, fibrotic interstitial lung disease of unknown etiology. The accumulation of macrophages is associated with disease pathogenesis. The unfolded protein response (UPR) has been linked to macrophage activation in pulmonary fibrosis. To date, the impact of activating transcription factor 6 alpha (ATF6α), one of the UPR mediators, on the composition and function of pulmonary macrophage subpopulations during lung injury and fibrogenesis is not fully understood. We began by examining the expression of Atf6α in IPF patients’ lung single‐cell RNA sequencing dataset, archived surgical lung specimens, and CD14 + circulating monocytes. To assess the impact of ATF6α on pulmonary macrophage composition and pro‐fibrotic function during tissue remodeling, we conducted an in vivo myeloid‐specific deletion of Atf6α . Flow cytometric assessments of pulmonary macrophages were carried out in C57BL/6 and myeloid specific ATF6α‐deficient mice in the context of bleomycin‐induced lung injury. Our results demonstrated that Atf6α mRNA was expressed in pro‐fibrotic macrophages found in the lung of a patient with IPF and in CD14 + circulating monocytes obtained from blood of a patient with IPF. After bleomycin administration, the myeloid‐specific deletion of Atf6α altered the pulmonary macrophage composition, expanding CD11b + subpopulations with dual polarized CD38 + CD206 + expressing macrophages. Compositional changes were associated with an aggravation of fibrogenesis including increased myofibroblast and collagen deposition. A further mechanistic ex vivo investigation revealed that ATF6α was required for CHOP induction and the death of bone marrow‐derived macrophages. Overall, our findings suggest a detrimental role for the ATF6α‐deficient CD11b + macrophages which had altered function during lung injury and fibrosis.
Publisher: Springer Science and Business Media LLC
Date: 24-06-2007
DOI: 10.1038/NM1604
Abstract: Gamma-aminobutyric acid (GABA) is an important neurotransmitter that, through the subtype A GABA receptor (GABAAR), induces inhibition in the adult brain. Here we show that an excitatory, rather than inhibitory, GABAergic system exists in airway epithelial cells. Both GABAARs and the GABA synthetic enzyme glutamic acid decarboxylase (GAD) are expressed in pulmonary epithelial cells. Activation of GABAARs depolarized these cells. The expression of GAD in the cytosol and GABAARs in the apical membranes of airway epithelial cells increased markedly when mice were sensitized and then challenged with ovalbumin, an approach for inducing allergic asthmatic reactions. Similarly, GAD and GABAARs in airway epithelial cells of humans with asthma increased after allergen inhalation challenge. Intranasal application of selective GABAAR inhibitors suppressed the hyperplasia of goblet cells and the overproduction of mucus induced by ovalbumin or interleukin-13 in mice. These findings show that a previously unknown epithelial GABAergic system has an essential role in asthma.
Publisher: American Thoracic Society
Date: 04-2004
Publisher: American Thoracic Society
Date: 04-2011
Publisher: American Thoracic Society
Date: 10-2013
Publisher: Springer Science and Business Media LLC
Date: 12-2017
Publisher: Springer Science and Business Media LLC
Date: 19-10-2021
DOI: 10.1186/S12931-021-01862-1
Abstract: Over 300 million people in the world live with asthma, resulting in 500,000 annual global deaths with future increases expected. It is estimated that around 50–80% of asthma exacerbations are due to viral infections. Currently, a combination of long-acting beta agonists (LABA) for bronchodilation and glucocorticoids (GCS) to control lung inflammation represent the dominant strategy for the management of asthma, however, it is still sub-optimal in 35–50% of moderate-severe asthmatics resulting in persistent lung inflammation, impairment of lung function, and risk of mortality. Mechanistically, LABA/GCS combination therapy results in synergistic efficacy mediated by intracellular cyclic adenosine monophosphate (cAMP). Increasing intracellular cAMP during LABA/GCS combination therapy via inhibiting phosphodiesterase 4 (PDE4) and/or blocking the export of cAMP by ATP Binding Cassette Transporter C4 (ABCC4), will potentiate anti-inflammatory responses of mainstay LABA/GCS therapy. Expression and localization experiments were performed using in situ hybridization and immunohistochemistry in human lung tissue from healthy subjects, while confirmatory transcript and protein expression analyses were performed in primary human airway epithelial cells and cell lines. Intervention experiments were performed on the human airway epithelial cell line, HBEC-6KT, by pre-treatment with combinations of LABA/GCS with PDE4 and/or ABCC4 inhibitors followed by Poly I:C or imiquimod challenge as a model for viral stimuli. Cytokine readouts for IL-6, IL-8, CXCL10/IP-10, and CCL5/RANTES were quantified by ELISA. Using archived human lung and human airway epithelial cells, ABCC4 gene and protein expression were confirmed in vitro and in situ. LABA/GCS attenuation of Poly I:C or imiquimod-induced IL-6 and IL-8 were potentiated with ABCC4 and PDE4 inhibition, which was greater when ABCC4 and PDE4 inhibition was combined. Modulation of cAMP levels had no impact on LABA/GCS modulation of Poly I:C-induced CXCL10/IP-10 or CCL5/RANTES. Modulation of intracellular cAMP levels by PDE4 or ABCC4 inhibition potentiates LABA/GCS efficacy in human airway epithelial cells challenged with viral stimuli. The data suggest further exploration of the value of adding cAMP modulators to mainstay LABA/GCS therapy in asthma for potentiated anti-inflammatory efficacy.
Publisher: Springer Science and Business Media LLC
Date: 14-04-2017
DOI: 10.1038/S41525-017-0015-6
Abstract: Pulmonary disease is the major cause of morbidity and mortality in patients with cystic fibrosis, a disease caused by mutations in the Cystic Fibrosis Transmembrane conductance Regulator (CFTR) gene. Heterogeneity in CFTR genotype–phenotype relationships in affected in iduals plus the escalation of drug discovery targeting specific mutations highlights the need to develop robust in vitro platforms with which to stratify therapeutic options using relevant tissue. Toward this goal, we adapted a fluorescence plate reader assay of apical CFTR -mediated chloride conductance to enable profiling of a panel of modulators on primary nasal epithelial cultures derived from patients bearing different CFTR mutations. This platform faithfully recapitulated patient-specific responses previously observed in the “gold-standard” but relatively low-throughput Ussing chamber. Moreover, using this approach, we identified a novel strategy with which to augment the response to an approved drug in specific patients. In proof of concept studies, we also validated the use of this platform in measuring drug responses in lung cultures differentiated from cystic fibrosis iPS cells. Taken together, we show that this medium throughput assay of CFTR activity has the potential to stratify cystic fibrosis patient-specific responses to approved drugs and investigational compounds in vitro in primary and iPS cell-derived airway cultures.
Publisher: Wiley
Date: 10-2019
DOI: 10.14814/PHY2.14249
Publisher: European Respiratory Society (ERS)
Date: 03-2008
DOI: 10.1183/09031936.00079307
Abstract: Asthma is characterised by an excessive airway narrowing in response to a variety of stimuli, called airway hyperresponsiveness (AHR). Previous comparisons between mouse strains have shown that increased velocity of airway narrowing correlates with baseline airway responsiveness. These data prompted the investigation into models of induced AHR to see whether airway narrowing dynamics correlated with in vivo responsiveness. In an attempt to reproduce some of the features of asthma, BALB/c mice were sensitised and subjected to either brief or chronic periods of allergen exposure. Brief exposure involved two challenges with intranasal chicken egg ovalbumin (OVA(in)). Chronic exposure involved six 2-day periods of OVA(in) challenges, each separated by 12 days. Control mice received intranasal saline challenges. Outcomes included videomicrometry of lung slices (magnitude and velocity of airway narrowing), in vivo respiratory physiology measurements and histological staining with morphometric analysis. Neither brief nor chronic allergen exposure resulted in greater airway narrowing and increased velocity compared with saline controls. Structural changes in the airway, such as goblet cell hyperplasia, subepithelial fibrosis and increased contractile tissue, were detected in mice chronically challenged with allergen. In conclusion, increased responsiveness to methacholine following allergen challenge may not be due to an intrinsic change to the smooth muscle per se, but rather to other changes in the lung, which ultimately manifest as an increase in respiratory resistance.
Publisher: American Thoracic Society
Date: 09-2011
Publisher: Cold Spring Harbor Laboratory
Date: 07-06-2021
DOI: 10.1101/2021.06.07.447351
Abstract: The angiotensin-converting enzyme 2 (ACE2) protein has been highly studied as a key catalytic regulator of the renin-angiotensin system (RAS), involved in fluid homeostasis and blood pressure modulation. In addition to its important physiological role as a broadly-expressed membrane-bound protein, ACE2 serves as a cell-surface receptor for some viruses - most notably, coronaviruses such as SARS-CoV and SARS-CoV-2. Differing levels of ACE2 expression may impact viral susceptibility and subsequent changes to expression may be a pathogenic mechanism of disease risk and manifestation. Therefore, an improved understanding of how ACE2 expression is regulated at the genomic and transcriptional level may help us understand not only how the effects of pre-existing conditions (e.g., chronic obstructive pulmonary disease) may manifest with increased COVID-19 incidence, but also the mechanisms that regulate ACE2 levels following viral infection. Here, we initially perform bioinformatic analyses of several datasets to generate hypotheses about ACE2 gene-regulatory mechanisms in the context of immune signaling and chronic oxidative stress. We then identify putative non-coding regulatory elements within ACE2 intronic regions as potential determinants of ACE2 expression activity. We perform functional validation of our computational predictions in vitro via targeted CRISPR-Cas9 deletions of the identified ACE2 cis -regulatory elements in the context of both immunological stimulation and oxidative stress conditions. We demonstrate that intronic ACE2 regulatory elements are responsive to both immune signaling and oxidative-stress pathways, and this contributes to our understanding of how expression of this gene may be modulated at both baseline and during immune challenge. Our work supports the further pursuit of these putative mechanisms in our understanding, prevention, and treatment of infection and disease caused by ACE2-utilizing viruses such as SARS-CoV, SARS-CoV-2, and future emerging SARS-related viruses. The recent emergence of the virus SARS-CoV-2 which has caused the COVID-19 pandemic has prompted scientists to intensively study how the virus enters human host cells. This work has revealed a key protein, ACE2, that acts as a receptor permitting the virus to infect cells. Much research has focused on how the virus physically interacts with ACE2, yet little is known on how ACE2 is turned on or off in human cells at the level of the DNA molecule. Understanding this level of regulation may offer additional ways to prevent or lower viral entry into human hosts. Here, we have examined the control of the ACE2 gene, the DNA sequence that instructs ACE2 protein receptor formation, and we have done so in the context of immune stimulation. We have indeed identified a number of DNA on/off switches for ACE2 that appear responsive to immuno-logical and oxidative stress. These switches may fine-tune how ACE2 is turned on or off before, during, and/or after infection by SARS-CoV-2 or other related coronaviruses. Our studies help pave the way for additional functional studies on these switches, and their potential therapeutic targeting in the future.
Publisher: Elsevier BV
Date: 12-2023
Publisher: Elsevier BV
Date: 10-2011
DOI: 10.1016/J.PUPT.2011.03.008
Abstract: Multiphoton microscopy has become a powerful imaging method for minimally invasive evaluation of extracellular matrix (ECM) and cellular structures deep within tissues in their native environments. This technology, which uses ultra-short femto-second laser pulses as the excitation source, is efficient in multiphoton excitation fluorescence (MPEF) of endogenously fluorescent macromolecular systems and induction of highly specific second harmonic generation (SHG) signals from non-centrosymmetric macromolecules such as fibrillar collagens. Both these signals can be captured simultaneously to provide spatially resolved 3D structural organization of ECM as well as cellular morphologies in lung or airway tissue with spectral specificity and sensitivity. These imaging modalities are minimally invasive since structures deep within tissues can be visualized without the need for tissue fixation and/or sectioning. Much of the traditional histological and chemical procedures associated with conventional microscopy methods, which may alter native structure of lung tissue s les, can be circumvented to generate more accurate 3D morphological and fine structural information. In addition to outlining basic principles associated with MPEF and SHG microscopy methods, this review reports potential uses of these high resolution imaging modalities in lung structural imaging. We place special emphasis on imaging 3D structural features of airways, visualizing and quantifying ECM remodeling associated with mouse asthma model as well as the potential uses for multiphoton microscopy in in vitro airway applications.
Publisher: Elsevier BV
Date: 11-2015
DOI: 10.1016/J.BIOCEL.2015.08.014
Abstract: The epithelium of asthmatics is characterized by reduced expression of E-cadherin and increased expression of the basal cell markers ck-5 and p63 that is indicative of a relatively undifferentiated repairing epithelium. This phenotype correlates with increased proliferation, compromised wound healing and an enhanced capacity to undergo epithelial-mesenchymal transition (EMT). The transcription factor β-catenin plays a vital role in epithelial cell differentiation and regeneration, depending on the co-factor recruited. Transcriptional programs driven by the β-catenin/CBP axis are critical for maintaining an undifferentiated and proliferative state, whereas the β-catenin 300 axis is associated with cell differentiation. We hypothesized that disrupting the β-catenin/CBP signaling axis would promote epithelial differentiation and inhibit EMT. We treated monolayer cultures of human airway epithelial cells with TGFβ1 in the presence or absence of the selective small molecule ICG-001 to inhibit β-catenin/CBP signaling. We used western blots to assess expression of an EMT signature, CBP, p300, β-catenin, fibronectin and ITGβ1 and scratch wound assays to assess epithelial cell migration. Snai-1 and -2 expressions were determined using q-PCR. Exposure to TGFβ1 induced EMT, characterized by reduced E-cadherin expression with increased expression of α-smooth muscle actin and EDA-fibronectin. Either co-treatment or therapeutic administration of ICG-001 completely inhibited TGFβ1-induced EMT. ICG-001 also reduced the expression of ck-5 and -19 independent of TGFβ1. Exposure to ICG-001 significantly inhibited epithelial cell proliferation and migration, coincident with a down regulation of ITGβ1 and fibronectin expression. These data support our hypothesis that modulating the β-catenin/CBP signaling axis plays a key role in epithelial plasticity and function.
Publisher: European Respiratory Society (ERS)
Date: 30-06-2015
Publisher: MDPI AG
Date: 07-03-2021
DOI: 10.3390/BIOMEDICINES9030267
Abstract: Despite the intricate involvement of the endocannabinoid system in various physiological processes, it remains one of the most under-studied biological systems of the human body. The scope of endocannabinoid signalling is widespread, ranging from modulation of immune responses in innate and adaptive immunity to gestational processes in female physiology. Cannabinoid receptors are ubiquitously distributed in reproductive tissues and are thought to play a role in regulating the immune–reproductive interactions required for successful pregnancy, specifically among uterine natural killer cells and placental extravillous trophoblasts. The use of cannabis during pregnancy, however, can perturb endocannabinoid homeostasis through effects mediated by its major constituents, Δ-9-tetrahydrocannabinol and cannabidiol. Decidualization of the endometrium, invasion, and angiogenesis may be impaired as a consequence, leading to clinical complications such as miscarriage and preecl sia. In this review, the crosstalk between endocannabinoid signalling in uterine natural killer cells and placental extravillous trophoblasts will be examined in healthy and complicated pregnancies. This lays a foundation for discussing the potential of targeting the endocannabinoid system for therapeutic benefit, particularly with regard to the emerging field of synthetic cannabinoids.
Publisher: Elsevier BV
Date: 05-2021
Publisher: Elsevier BV
Date: 06-2016
DOI: 10.1016/J.MOLIMM.2017.01.014
Abstract: Innate immune responses act as first line defences upon exposure to potentially noxious stimuli. The innate immune system has evolved numerous intracellular and extracellular receptors that undertake surveillance for potentially damaging particulates. Inflammasomes are intracellular innate immune multiprotein complexes that form and are activated following interaction with these stimuli. Inflammasome activation leads to the cleavage of pro-IL-1β and release of the pro-inflammatory cytokine, IL-1β, which initiates acute phase pro-inflammatory responses, and other responses are also involved (IL-18, pyroptosis). However, excessive activation of inflammasomes can result in chronic inflammation, which has been implicated in a range of chronic inflammatory diseases. The airways are constantly exposed to a wide variety of stimuli. Inflammasome activation and downstream responses clears these stimuli. However, excessive activation may drive the pathogenesis of chronic respiratory diseases such as severe asthma and chronic obstructive pulmonary disease. Thus, there is currently intense interest in the role of inflammasomes in chronic inflammatory lung diseases and in their potential for therapeutic targeting. Here we review the known associations between inflammasome-mediated responses and the development and exacerbation of chronic lung diseases.
Publisher: Wiley
Date: 15-05-2014
DOI: 10.1111/RESP.12316
Abstract: Airway epithelial cells represent the first line of defence against inhaled insults, including air pollution. Air pollution can activate innate immune signalling in airway epithelial cells leading to the production of soluble mediators that can influence downstream inflammatory cells. Our objective was to develop and validate a model of dendritic cell exposure to airway epithelial cell-conditioned media. After establishing the model, we explored how soluble mediators released from airway epithelial cells in response to air pollution influenced the phenotype of dendritic cells. Human airway epithelial cells were cultured under control and urban particulate matter (PM10) exposure conditions with or without pharmacological inhibitors of the uric acid pathway. Culture supernatants were collected for conditioned media experiments with peripheral blood mononuclear cell-derived dendritic cells analysed by flow cytometry. Monocytes derived from peripheral blood mononuclear cells cultured in interleukin-4 and granulocyte macrophage colony stimulating factor differentiated into immature dendritic cells that phenotypically differentiated into mature dendritic cells in response to conditioned media from phorbol myristate acetate-activated THP-1 monocytes. Exposure of immature dendritic cells to conditioned media from airway epithelial cells exposed to PM10 resulted in dendritic cell maturation that was independent of uric acid. We present a conditioned media model useful for interrogating the contribution of soluble mediators produced by airway epithelial cells to dendritic cell phenotype and function. Furthermore, we demonstrate that PM10 exposure induces airway epithelial cell production of soluble mediators that induce maturation of dendritic cells independent of uric acid.
Publisher: MDPI AG
Date: 26-01-2021
DOI: 10.3390/MI12020132
Abstract: Polydimethylsiloxane (PDMS) is a silicone-based synthetic material used in various biomedical applications due to its properties, including transparency, flexibility, permeability to gases, and ease of use. Though PDMS facilitates and assists the fabrication of complicated geometries at micro- and nano-scales, it does not optimally interact with cells for adherence and proliferation. Various strategies have been proposed to render PDMS to enhance cell attachment. The majority of these surface modification techniques have been offered for a static cell culture system. However, dynamic cell culture systems such as organ-on-a-chip devices are demanding platforms that recapitulate a living tissue microenvironment’s complexity. In organ-on-a-chip platforms, PDMS surfaces are usually coated by extracellular matrix (ECM) proteins, which occur as a result of a physical and weak bonding between PDMS and ECM proteins, and this binding can be degraded when it is exposed to shear stresses. This work reports static and dynamic coating methods to covalently bind collagen within a PDMS-based microfluidic device using polydopamine (PDA). These coating methods were evaluated using water contact angle measurement and atomic force microscopy (AFM) to optimize coating conditions. The biocompatibility of collagen-coated PDMS devices was assessed by culturing primary human bronchial epithelial cells (HBECs) in microfluidic devices. It was shown that both PDA coating methods could be used to bind collagen, thereby improving cell adhesion (approximately three times higher) without showing any discernible difference in cell attachment between these two methods. These results suggested that such a surface modification can help coat extracellular matrix protein onto PDMS-based microfluidic devices.
Publisher: Springer Science and Business Media LLC
Date: 12-2015
Publisher: Springer Science and Business Media LLC
Date: 13-10-2016
DOI: 10.1038/SREP35338
Abstract: There is an unmet need for effective new and innovative treatments for asthma. It is becoming increasingly evident that bacterial stimulation can have beneficial effects at attenuating allergic airway disease through immune modulation. Our aim was to test the ability of a novel inactivated microbe-derived therapeutic based on Klebsiella (KB) in a model of allergic airway disease in mice. BALB/c mice were exposed intranasally to house dust mite (HDM) for two weeks. Mice were treated prophylactically via subcutaneous route with either KB or placebo for one week prior to HDM exposure and throughout the two week exposure period. 24 hours after the last exposure, lungs were analysed for inflammatory cell infiltrate, gene expression, cytokine levels, goblet cell metaplasia, and serum was analysed for allergen-specific serum IgE levels. HDM exposed mice developed goblet cell hyperplasia, elevated allergen-specific serum IgE, airway eosinophilia, and a concomitant increase in T H 2 cytokines including IL-4, IL-13 and IL-5. Treatment with KB attenuated HDM-mediated airway eosinophilia, total bronchoalveolar lavage (BAL) cell numbers, BAL T H 2 cytokine production, and goblet cell metaplasia. Our prophylactic intervention study illustrates the potential of subcutaneous treatment with bacterial derived biologics as a promising approach for allergic airway disease treatment.
Publisher: Wiley
Date: 06-08-2018
Publisher: Informa UK Limited
Date: 23-02-2016
DOI: 10.3109/08958378.2016.1145770
Abstract: Epidemiological studies and animal research have suggested that air pollution may negatively impact the central nervous system (CNS). Controlled human exposure studies of the effect of air pollution on the brain have potential to enhance our understanding of this relationship and to inform potential biological mechanisms. Biomarkers of systemic and CNS inflammation may address whether air pollution exposure induces inflammation, with potential for CNS negative effects. Twenty-seven healthy adults were exposed to two conditions: filtered air (FA) and diesel exhaust (DE) (300 μg PM2.5/m(3)) for 120 min, in a double-blinded crossover study with exposures separated by four weeks. Prior to and at 0, 3, and 24 h following each exposure, serum and plasma were collected and analyzed for inflammatory cytokines interleukin 6 (IL-6) and tumour necrosis factor alpha (TNF-α), the astrocytic protein S100b, the neuronal cytoplasmic enzyme neuron-specific enolase (NSE), and serum brain-derived neurotrophic factor (BDNF). We hypothesized that IL-6, TNF-α, S100b and NSE would increase, and BDNF would decrease, following DE exposure. At no time-point following exposure to DE was a significant increase in concentration from baseline seen for IL-6, TNF-α, S100b, or NSE relative to FA exposure. Similarly, no significant decrease in BDNF concentration from baseline was seen following DE exposure, relative to FA. Furthermore, the repeated measures ANOVA considered for all time-points and biomarkers revealed no significant time-exposure interaction. These results suggest that short-term exposure to DE amongst healthy adults does not acutely affect the systemic or CNS biomarkers that we measured.
Publisher: Wiley
Date: 19-04-2010
DOI: 10.1111/J.1365-2222.2010.03458.X
Abstract: Background Asthma is a disease characterized by airway inflammation, remodelling and dysfunction. Airway inflammation contributes to remodelling, a term that is used to describe structural changes including goblet cell metaplasia (GCM), matrix deposition, and smooth muscle hyperplasia/hypertrophy. GCM has been implicated in asthma mortality by contributing to mucus plugs and leading to asphyxiation. In animal models, this process is highly dependent on IL-13. Recently, we have described an IL-13-dependent up-regulation of a GABAergic signalling system in airway epithelium that contributes to GCM. The mechanism by which IL-13 up-regulates GABA signalling in airway epithelium is unknown. Objectives To test the hypothesis that IL-4Ralpha signalling is required for allergen induced up-regulation of GABAergic signalling and GCM. Methods BALB/c mice were exposed to an acute house dust mite (HDM) protocol and received vehicle, anti-IL-4Ralpha-monoclonal antibody, or control antibody. Outcomes included airway responses to inhaled methacholine (MCh), histology for eosinophilia and GCM, phosphorylated STAT6 levels using immunohistochemistry and immunoblot, and glutamic acid decarboxylase (GAD) 65/67 and GABA(A)beta(2/3) receptor subunit expression using confocal microscopy. Results Acute HDM exposure resulted in increased airway responses to MCh, lung eosinophilia, STAT6 phosphorylation, elevations in GAD65/67 and GABA(A)beta(2/3) receptor expression, and GCM that were inhibited with anti-IL-4Ralpha-monoclonal treatment. Control antibody had no effect. Conclusion The IL-4Ralpha is required for allergen-induced up-regulation of a GABAergic system in airway epithelium implicated in GCM following acute HDM exposure.
Publisher: Wiley
Date: 12-10-2017
DOI: 10.1111/RESP.12908
Abstract: COPD is a major cause of global mortality and morbidity but current treatments are poorly effective. This is because the underlying mechanisms that drive the development and progression of COPD are incompletely understood. Animal models of disease provide a valuable, ethically and economically viable experimental platform to examine these mechanisms and identify biomarkers that may be therapeutic targets that would facilitate the development of improved standard of care. Here, we review the different established animal models of COPD and the various aspects of disease pathophysiology that have been successfully recapitulated in these models including chronic lung inflammation, airway remodelling, emphysema and impaired lung function. Furthermore, some of the mechanistic features, and thus biomarkers and therapeutic targets of COPD identified in animal models are outlined. Some of the existing therapies that suppress some disease symptoms that were identified in animal models and are progressing towards therapeutic development have been outlined. Further studies of representative animal models of human COPD have the strong potential to identify new and effective therapeutic approaches for COPD.
Publisher: Elsevier BV
Date: 07-2016
DOI: 10.1016/J.FREERADBIOMED.2016.04.202
Abstract: Isolated exposure to diesel exhaust (DE) or allergen can cause decrements in lung function that are impacted by the presence of genetic variants in the glutathione-S-transferase (GST) family but the effect of GST interactions with DE-allergen co-exposure on lung function is unknown. We aimed to assess the impact of DE and allergen co-exposure on lung function and the influence of GSTM1 or GSTT1 variation We used a blinded crossover study design with 17 atopic subjects exposed to filtered air (FA the control for DE) or DE for 2h. One hour following each exposure to DE or FA, bronchoscopy was performed to deliver a diluent-controlled segmental allergen challenge (SAC). Methacholine challenge and forced expiratory volume in 1s (FEV1) was performed pre-exposure (baseline airway responsiveness) and 24h post-exposure (effect of co-exposure). Additionally, FEV1 was performed hourly after DE/FA exposure and protein carbonyl content was measured in plasma as an oxidative stress marker. Changes in FEV1 from baseline were dependent on time following allergen exposure. DE, as opposed to FA, led to a significant change in FEV1 at 2h post-allergen exposure in GSTT1 variants only (24.5±19.6% reduction in GSTT1 null in iduals vs. 9.2±7.3% reduction in GSTT1 present in iduals). Moreover, plasma protein carbonyl level 4h after co-exposure was higher in the in iduals who have the GSTT1 null genotype. This suggests a gene-environment interaction that endangers susceptible populations co-exposed to DE and allergen.
Publisher: European Respiratory Society (ERS)
Date: 16-07-2020
DOI: 10.1183/13993003.01123-2020
Abstract: In December 2019, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) emerged, causing the coronavirus disease 2019 (COVID-19) pandemic. SARS-CoV, the agent responsible for the 2003 SARS outbreak, utilises angiotensin-converting enzyme 2 (ACE2) and transmembrane serine protease 2 (TMPRSS2) host molecules for viral entry. ACE2 and TMPRSS2 have recently been implicated in SARS-CoV-2 viral infection. Additional host molecules including ADAM17, cathepsin L, CD147 and GRP78 may also function as receptors for SARS-CoV-2. To determine the expression and in situ localisation of candidate SARS-CoV-2 receptors in the respiratory mucosa, we analysed gene expression datasets from airway epithelial cells of 515 healthy subjects, gene promoter activity analysis using the FANTOM5 dataset containing 120 distinct s le types, single cell RNA sequencing (scRNAseq) of 10 healthy subjects, proteomic datasets, immunoblots on multiple airway epithelial cell types, and immunohistochemistry on 98 human lung s les. We demonstrate absent to low ACE2 promoter activity in a variety of lung epithelial cell s les and low ACE2 gene expression in both microarray and scRNAseq datasets of epithelial cell populations. Consistent with gene expression, rare ACE2 protein expression was observed in the airway epithelium and alveoli of human lung, confirmed with proteomics. We present confirmatory evidence for the presence of TMPRSS2, CD147 and GRP78 protein in vitro in airway epithelial cells and confirm broad in situ protein expression of CD147 and GRP78 in the respiratory mucosa. Collectively, our data suggest the presence of a mechanism dynamically regulating ACE2 expression in human lung, perhaps in periods of SARS-CoV-2 infection, and also suggest that alternative receptors for SARS-CoV-2 exist to facilitate initial host cell infection.
Publisher: Springer Science and Business Media LLC
Date: 06-02-2017
DOI: 10.1038/SREP42214
Abstract: S ling various compartments within the lower airways to examine human bronchial epithelial cells (HBEC) is essential for understanding numerous lung diseases. Conventional methods to identify HBEC in bronchoalveolar lavage (BAL) and wash (BW) have throughput limitations in terms of efficiency and ensuring adequate cell numbers for quantification. Flow cytometry can provide high-throughput quantification of cell number and function in BAL and BW s les, while requiring low cell numbers. To date, a flow cytometric method to identify HBEC recovered from lower human airway s les is unavailable. In this study we present a flow cytometric method identifying HBEC as CD45 negative, EpCAM an-cytokeratin (pan-CK) double-positive population after excluding debris, doublets and dead cells from the analysis. For validation, the HBEC panel was applied to primary HBEC resulting in 98.6% of live cells. In healthy volunteers, HBEC recovered from BAL (2.3% of live cells), BW (32.5%) and bronchial brushing s les (88.9%) correlated significantly (p = 0.0001) with the manual microscopy counts with an overall Pearson correlation of 0.96 across the three s le types. We therefore have developed, validated, and applied a flow cytometric method that will be useful to interrogate the role of the respiratory epithelium in multiple lung diseases.
Publisher: Wiley
Date: 14-09-2023
DOI: 10.1002/CJCE.24581
Abstract: The incidence of respiratory diseases such as chronic obstructive pulmonary disease and pulmonary cancer is growing significantly around the world, making pulmonary disease one of the leading causes of mortality. However, the development of effective therapeutics for pulmonary diseases has been hindered by the lack of human‐mimetic physiological models that reliably emulate patient responses. Recent advances in technology and cell culture have led to the development of organoids and organ‐on‐a‐chip models that allow us to recapitulate the structure, cellular organization, and organ‐level responses of the target tissue in vitro. Here, we review the advances and milestones of lung organoid and lung‐on‐a‐chip models in the past decade and highlight their applications in mimicking pulmonary system development, physiology, disease, and regeneration. In addition, we discuss the ongoing challenges and the future prospects of integrating lung organoids and lung‐on‐a‐chip models to overcome current limitations and to enhance their physiological relevance. These human‐centric models are likely to provide important insights into pulmonary physiology and pathophysiology for drug discovery that complement and potentially replace traditional animal models.
Publisher: Elsevier BV
Date: 03-2018
Publisher: American Thoracic Society
Date: 06-2019
Publisher: Elsevier BV
Date: 10-2021
Publisher: Elsevier BV
Date: 10-2009
DOI: 10.1016/J.PUPT.2009.04.001
Abstract: Asthma is a complex disease that involves chronic inflammation and subsequent decline in airway function. The widespread use of animal models has greatly contributed to our understanding of the cellular and molecular pathways underlying human allergic asthma. Animal models of allergic asthma include smaller animal models which offer 'ease of use' and availability of reagents, and larger animal models that may be used to address aspects of allergic airways disease not possible in humans or smaller animal models. This review examines the application and suitability of various animal models for studying mechanisms of airway inflammation and tissue remodelling in allergic asthma, with a specific focus on airway smooth muscle.
Publisher: Springer Science and Business Media LLC
Date: 13-01-2021
DOI: 10.1038/S41598-020-79555-W
Abstract: Cystic fibrosis (CF) is a genetic disease characterized by CF transmembrane regulator (CFTR) dysfunction. With over 2000 CFTR variants identified, in addition to known patient to patient variability, there is a need for personalized treatment. The discovery of CFTR modulators has shown efficacy in certain CF populations, however there are still CF populations without valid therapeutic options. With evidence suggesting that single drug therapeutics are insufficient for optimal management of CF disease, there has been an increased pursuit of combinatorial therapies. Our aim was to test cyclic AMP (cAMP) modulation, through ATP Binding Cassette Transporter C4 (ABCC4) and phosphodiesterase-4 (PDE-4) inhibition, as a potential add-on therapeutic to a clinically approved CFTR modulator, VX-770, as a method for increasing CFTR activity. Human airway epithelial cells (Calu-3) were used to test the efficacy of cAMP modulation by ABCC4 and PDE-4 inhibition through a series of concentration–response studies. Our results showed that cAMP modulation, in combination with VX-770, led to an increase in CFTR activity via an increase in sensitivity when compared to treatment of VX-770 alone. Our study suggests that cAMP modulation has potential to be pursued as an add-on therapy for the optimal management of CF disease.
Publisher: Wiley
Date: 28-06-2017
DOI: 10.1111/IMR.12543
Abstract: Severe, steroid-resistant asthma is clinically and economically important since affected in iduals do not respond to mainstay corticosteroid treatments for asthma. Patients with this disease experience more frequent exacerbations of asthma, are more likely to be hospitalized, and have a poorer quality of life. Effective therapies are urgently required, however, their development has been h ered by a lack of understanding of the pathological processes that underpin disease. A major obstacle to understanding the processes that drive severe, steroid-resistant asthma is that the several endotypes of the disease have been described that are characterized by different inflammatory and immunological phenotypes. This heterogeneity makes pinpointing processes that drive disease difficult in humans. Clinical studies strongly associate specific respiratory infections with severe, steroid-resistant asthma. In this review, we discuss key findings from our studies where we describe the development of representative experimental models to improve our understanding of the links between infection and severe, steroid-resistant forms of this disease. We also discuss their use in elucidating the mechanisms, and their potential for developing effective therapeutic strategies, for severe, steroid-resistant asthma. Finally, we highlight how the immune mechanisms and therapeutic targets we have identified may be applicable to obesity-or pollution-associated asthma.
Publisher: Microbiology Society
Date: 12-2020
DOI: 10.1099/JGV.0.001491
Abstract: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) recently emerged to cause widespread infections in humans. SARS-CoV-2 infections have been reported in the Kingdom of Saudi Arabia, where Middle East respiratory syndrome coronavirus (MERS-CoV) causes seasonal outbreaks with a case fatality rate of ~37 %. Here we show that there exists a theoretical possibility of future recombination events between SARS-CoV-2 and MERS-CoV RNA. Through computational analyses, we have identified homologous genomic regions within the ORF1ab and S genes that could facilitate recombination, and have analysed co-expression patterns of the cellular receptors for SARS-CoV-2 and MERS-CoV, ACE2 and DPP4, respectively, to identify human anatomical sites that could facilitate co-infection. Furthermore, we have investigated the likely susceptibility of various animal species to MERS-CoV and SARS-CoV-2 infection by comparing known virus spike protein–receptor interacting residues. In conclusion, we suggest that a recombination between SARS-CoV-2 and MERS-CoV RNA is possible and urge public health laboratories in high-risk areas to develop diagnostic capability for the detection of recombined coronaviruses in patient s les.
Publisher: European Respiratory Society (ERS)
Date: 20-10-2022
Publisher: American Thoracic Society
Date: 07-2017
Publisher: American Thoracic Society
Date: 2022
Publisher: Elsevier BV
Date: 10-2009
DOI: 10.1016/J.PUPT.2008.12.004
Abstract: Clinical asthma is characterized by reversible airway obstruction which is commonly due to an exaggerated airway narrowing referred to as airway hyperresponsiveness (AHR). Although debate exists on the complex etiology of AHR, it is clear that airway smooth muscle (ASM) mediated airway narrowing is a major contributor to airway dysfunction. More importantly, it is now appreciated that smooth muscle is far from being a simple cell with only contractile ability properties. Rather, it is more versatile with the capacity to exhibit numerous cellular functions as it adapts to the microenvironment to which it is exposed. The emerging ability of in idual smooth muscle cells to undergo changes in their phenotype (phenotype plasticity) and function (functional plasticity) in response to physiological and pathological cues is an important and active area of research. This article provides a brief review of the current knowledge and emerging concepts in the field of ASM phenotype and function both under healthy and asthmatic conditions.
Publisher: Elsevier BV
Date: 12-2012
DOI: 10.1016/J.COI.2012.08.012
Abstract: The innate immunity function of the human airway epithelium is responsible for orchestrating defence against inhaled viruses, bacteria, fungi, allergens, pollution, and other environmental insults. Epithelial cells present a mechanically tight, pseudostratified, multi-cell barrier that secretes mucus, surfactants, and anti-microbial peptides to manage minor insults. Secondary to the mechanical impedances, cell surface and cytoplasmic pattern recognition receptors await detection of more aggressive insults. The differentiation state of the airway epithelium contributes to innate immunity by compartmentalizing receptors and mediator production. Activation of innate immune receptors triggers production of interferons, cytokines, and chemokines, which influence adaptive immune responses. Mounting evidence suggests that these responses are aberrant in asthma and may contribute to disease progression and exacerbations. In this review, we discuss the recent evidence supporting these statements, focusing primarily on data generated from using human s les.
Publisher: Springer Science and Business Media LLC
Date: 15-05-2017
Publisher: Elsevier BV
Date: 05-2016
DOI: 10.1038/MI.2015.104
Abstract: Exposure to particulate matter (PM), a major component of air pollution, contributes to increased morbidity and mortality worldwide. PM induces innate immune responses and contributes to allergic sensitization, although the mechanisms governing this process remain unclear. Lung mucosal uric acid has also been linked to allergic sensitization. The links among PM exposure, uric acid, and allergic sensitization remain unexplored. We therefore investigated the mechanisms behind PM-induced allergic sensitization in the context of lung mucosal uric acid. PM10 and house dust mite exposure selectively induced lung mucosal uric acid production and secretion in vivo, which did not occur with other challenges (lipopolysaccharide, virus, bacteria, or inflammatory/fibrotic stimuli). PM10-induced uric acid mediates allergic sensitization and augments antigen-specific T-cell proliferation, which is inhibited by uricase. We then demonstrate that human airway epithelial cells secrete uric acid basally and after stimulation through a previously unidentified mucosal secretion system. Our work discovers a previously unknown mechanism of air pollution-induced, uric acid-mediated, allergic sensitization that may be important in the pathogenesis of asthma.
Publisher: Frontiers Media SA
Date: 18-10-2022
DOI: 10.3389/FBIOE.2022.959335
Abstract: Integration of mechanical cues in conventional 2D or 3D cell culture platforms is an important consideration for in vivo and ex vivo models of lung health and disease. Available commercial and published custom-made devices are frequently limited in breadth of applications, scalability, and customization. Herein we present a technical report on an open-source, cell and tissue (CaT) stretcher, with modularity for different in vitro and ex vivo systems, that includes the following features: 1) Programmability for modeling different breathing patterns, 2) scalability to support low to high-throughput experimentation, and 3) modularity for submerged cell culture, organ-on-chips, hydrogels, and live tissues. The strategy for connecting the experimental cell or tissue s les to the stretching device were designed to ensure that traditional biomedical outcome measurements including, but not limited to microscopy, soluble mediator measurement, and gene and protein expression remained possible. Lastly, to increase the uptake of the device within the community, the system was built with economically feasible and available components. To accommodate erse in vitro and ex vivo model systems we developed a variety of chips made of compliant polydimethylsiloxane (PDMS) and optimized coating strategies to increase cell adherence and viability during stretch. The CaT stretcher was validated for studying mechanotransduction pathways in lung cells and tissues, with an increase in alpha smooth muscle actin protein following stretch for 24 h observed in independent submerged monolayer, 3D hydrogel, and live lung tissue experiments. We anticipate that the open-source CaT stretcher design will increase accessibility to studies of the dynamic lung microenvironment through direct implementation by other research groups or custom iterations on our designs.
Publisher: Cold Spring Harbor Laboratory
Date: 19-08-2023
DOI: 10.1101/2023.08.17.553712
Abstract: Polydimethylsiloxane (PDMS) is widely used for microfluidics fabrication in many disciplines due to its ease of use in soft lithography and its ability to bond liquid-tight seals. A variety of PDMS-PDMS bonding methods exist, but each may have limitations for some applications. For ex le, chemical bonding via oxygen plasma treatment is reliable but requires expensive equipment and specialized training. Here we present a rapid, low-cost, and accessible method for irreversible PDMS bonding in which flame treatment activates PDMS and Nitto 5302A adhesive surfaces. Using this technique, PDMS microchannels can be fabricated with a bonding integrity of up to 325 kPa burst pressure. This technique is suitable for fabricating organ-on-chip devices for cell line and primary cell culture, supporting cell viability and establishment of key cellular features in apical and basal compartments, while maintaining bonding integrity.
Publisher: Elsevier BV
Date: 08-2022
Publisher: The Company of Biologists
Date: 30-04-2009
DOI: 10.1242/DMM.001859
Abstract: Experimental mouse models of asthma have broadened our understanding of the mechanisms behind allergen-induced asthma. Typically, mouse models of allergic asthma explore responses to a single allergen however, patients with asthma are frequently exposed to, and tend to be allergic to, more than one allergen. The aim of the current study was to develop a new and more relevant mouse model of asthma by measuring the functional, inflammatory and structural consequences of chronic exposure to a combination of two different allergens, ovalbumin (OVA) and house dust mite (HDM), in comparison with either allergen alone. BALB/c mice were sensitized and exposed to OVA, HDM or the combination of HDM and OVA for a period of 10 weeks. Following allergen exposure, airway responsiveness was measured using the flexiVent small animal ventilator, and mice were assessed for indices of airway inflammation and remodeling at both 24 hours and 4 weeks after the final allergen exposure. Mice exposed to the HDM-OVA combination exhibited increased numbers of inflammatory cells in the bronchoalveolar lavage (BAL) when compared with mice exposed to a single allergen. Mice exposed to HDM-OVA also exhibited an elevated level of lung tissue mast cells compared with mice exposed to a single allergen. Following the resolution of inflammatory events, mice exposed to the allergen combination displayed an elevation in the maximal degree of total respiratory resistance (Max RRS) compared with mice exposed to a single allergen. Furthermore, trends for increases in indices of airway remodeling were observed in mice exposed to the allergen combination compared with a single allergen. Although concurrent exposure to HDM and OVA resulted in increased aspects of airway hyperresponsiveness, airway inflammation and airway remodeling when compared with exposure to each allergen alone, concurrent exposure did not result in a substantially more robust mouse model of allergic asthma than exposure to either allergen alone.
Publisher: Frontiers Media SA
Date: 16-09-2020
Publisher: Elsevier BV
Date: 06-2019
DOI: 10.1016/J.JACI.2019.04.012
Abstract: Every day, we breathe in more than 10,000 L of air that contains a variety of air pollutants that can pose negative consequences to lung health. The respiratory mucosa formed by the airway epithelium is the first point of contact for air pollution in the lung, functioning as a mechanical and immunologic barrier. Under normal circumstances, airway epithelial cells connected by tight junctions secrete mucus, airway surface lining fluid, host defense peptides, and antioxidants and express innate immune pattern recognition receptors to respond to inhaled foreign substances and pathogens. Under conditions of air pollution exposure, the defenses of the airway epithelium are compromised by reductions in barrier function, impaired host defense to pathogens, and exaggerated inflammatory responses. Central to the mechanical and immunologic changes induced by air pollution are activation of redox-sensitive pathways and a role for antioxidants in normalizing these negative effects. Genetic variants in genes important in epithelial cell function and phenotype contribute to a ersity of responses to air pollution in the population at the in idual and group levels and suggest a need for personalized approaches to attenuate the respiratory mucosal immune responses to air pollution.
Publisher: Elsevier BV
Date: 07-2022
Publisher: European Respiratory Society (ERS)
Date: 17-06-2021
DOI: 10.1183/23120541.00107-2021
Abstract: Cannabis is widely used for both recreational and medicinal purposes. Inhalation of combusted cannabis smoke is the most common mode of drug consumption, exposing the lungs to the pharmacologically active ingredients, including tetrahydrocannabinol (THC) and cannabidiol (CBD). While the relationship between cannabis smoke exposure and compromised respiratory health has yet to be sufficiently defined, previous investigations suggest that cannabis smoke may dysregulate pulmonary immunity. Presently, there exist few preclinical animal models that have been extensively validated for contemporary cannabis smoke exposure. To address this need, we developed a mouse model with readouts of total particulate matter, serum cannabinoid and carboxyhaemoglobin levels, lung cellular responses, and immune-mediator production. Using a commercially available smoke exposure system and a cannabis source material of documented THC/CBD composition, we exposed mice to a mean± sd total particulate matter of 698.89±66.09 µg·L −1 and demonstrate increases in serum cannabinoids and carboxyhaemoglobin. We demonstrate that cannabis smoke modulates immune cell populations and mediators in both male and female BALB/c mice. This modulation is highlighted by increases in airway and lung tissue macrophage populations, including tissue-resident alveolar macrophages, monocyte-derived alveolar macrophages, and interstitial macrophage subpopulations. No changes in airway or lung tissue infiltration of neutrophils were observed. Immune-mediator analysis indicated significant upregulation of macrophage-derived chemokine, thymus and activation-regulated chemokine, and vascular endothelial growth factor within the lung tissue of cannabis smoke-exposed mice. This accessible and reproducible smoke-exposure model provides a foundation to explore the impact of chronic cannabis exposures and/or co-exposures with pathogens of clinical relevance, such as influenza.
Publisher: Centers for Disease Control and Prevention (CDC)
Date: 09-2020
Publisher: Cold Spring Harbor Laboratory
Date: 09-01-2019
DOI: 10.1101/516294
Abstract: Global recreational cannabis use is a potentially important public health issue that would benefit from experimental evidence to inform policy, regulations, and in idual user practices. Comparative analyses between cannabis and tobacco smoke, the latter long reported to have negative impacts on respiratory health, may help provide context and provide clinically relevant evidence. To address this unmet need we performed a comparative study between cannabis and tobacco smoke exposure in the Calu-3 human airway epithelial cells using concentration-response and pharmacological intervention study designs with outcome measurements of cell viability, epithelial cell barrier function, cytokine profile, and transcriptomics. Our results demonstrate that cannabis smoke exposure reduces epithelial cell barrier function without impacting cell viability, accompanied by a cytokine profile associated with inflammation (elevated IL-6 and IL-8), barrier repair (elevated TGF-α and PDGF-AA) and suppressed antiviral immunity (decreased IP-10 and RANTES). Transcriptomic analyses revealed a cannabis smoke induced signature associated with suppressed antiviral genes and induction of oncogenic and oxidative stress pathways. Similar trends were observed for tobacco smoke exposure. A formoterol/budesonide intervention was unable to prevent cannabis smoke-induced reductions in antiviral pathways or normalize induction of oncogenic and oxidative stress responses. Our results show striking similarities between cannabis and tobacco smoke exposure on impairing barrier function, suppressing antiviral pathways, potentiating of pro-inflammatory mediators, and inducing oncogenic and oxidative stress gene expression signatures. Furthermore, we demonstrate that an intervention with formoterol and budesonide is unable to completely normalized cannabisinduced responses. Collectively our data suggest that cannabis smoke exposure is not innocuous and may possess many of the deleterious properties of tobacco smoke, warranting additional studies to support public policy, government regulations, and in idual user practices.
Publisher: Springer Science and Business Media LLC
Date: 21-09-2006
Publisher: Elsevier BV
Date: 09-2015
DOI: 10.1016/J.JACI.2015.01.019
Abstract: Inpp5d (Src homology 2 domain-containing inositol-5-phosphatase [Ship1])-deficient mice experience spontaneous airway inflammation and have enhanced sensitivity to allergen-induced airway inflammation. We hypothesized that lineage-specific deletion of Ship1 expression in cells known to be crucial for adaptive TH2 responses would uncover distinct roles that could either positively or negatively regulate susceptibility to allergic airway inflammation (AAI). Ship1 expression was deleted in B cells, T cells, or dendritic cells (DCs), and the resulting Ship1(ΔB cell), Ship1(ΔT cell), Ship1(ΔDC), or Ship1(F/F) (wild-type) control mice were evaluated in a model of house dust mite (HDM)-induced AAI. Unlike germline panhematopoietic Ship1 deletion, deletion of Ship1 selectively in either the B-cell, T-cell, or DC lineages did not result in spontaneous airway inflammation. Strikingly, although loss of Ship1 in the B-cell lineage did not affect HDM-induced AAI, loss of Ship1 in either of the T-cell or DC lineages protected mice from AAI by skewing the typical TH2 immune response toward a TH1 response. Although panhematopoietic deletion of Ship1 leads to spontaneous lung inflammation, selective deletion of Ship1 in T cells or DCs impairs the formation of an adaptive TH2 response and protects animals from HDM-induced AAI.
Publisher: American Thoracic Society
Date: 2015
Publisher: Royal Society of Chemistry (RSC)
Date: 2022
DOI: 10.1039/D1LC01141C
Abstract: We developed a subtractive manufacturing with swelling induced stochastic folding of sacrificial materials for fabricating complex perfusable tissues in high-throughput multi-well plate formats.
Publisher: Elsevier BV
Date: 06-2021
Publisher: American Chemical Society (ACS)
Date: 26-10-2021
Publisher: Wiley
Date: 08-12-2021
Abstract: In vitro wound healing assays are widely used to investigate cell migration during various physiologic and pathologic processes. However, traditional scratch‐based assays produce cell‐free areas that are not reproducible, whereas the alternate insert‐based exclusion method is expensive and time‐consuming. Here, a rapid, label‐free, insert‐free magnetic exclusion technique, where magnetic fields are used to create cell‐free areas is described. Suspensions of diamagnetic cells in a paramagnetic culture medium are seeded into microwells placed on an array of the coaxially arranged ring and cylinder magnets. In the presence of an inhomogeneous field, the magnetic susceptibility difference drives the cells toward an annular region of the lowest field strength. Annular cell aggregates of reproducible dimensions are produced on tissue culture‐treated and collagen I‐fibronectin coated surfaces within 3 h. The effects of the paramagnetic agent on cell viability, metabolism, and gene expression are investigated. A mathematical model reveals the dynamics of the cell‐free area closure. The magnetic exclusion technique is inexpensive, easy to use, and amenable to automation. Potential applications include cancer research, high throughput drug discovery, and screening.
Publisher: MDPI AG
Date: 18-06-2021
Abstract: Decellularization efforts must balance the preservation of the extracellular matrix (ECM) components while eliminating the nucleic acid and cellular components. Following effective removal of nucleic acid and cell components, decellularized ECM (dECM) can be solubilized in an acidic environment with the assistance of various enzymes to develop biological scaffolds in different forms, such as sheets, tubular constructs, or three-dimensional (3D) hydrogels. Each organ or tissue that undergoes decellularization requires a distinct and optimized protocol to ensure that nucleic acids are removed, and the ECM components are preserved. The objective of this study was to optimize the decellularization process for dECM isolation from human lung tissues for downstream 2D and 3D cell culture systems. Following protocol optimization and dECM isolation, we performed experiments with a wide range of dECM concentrations to form human lung dECM hydrogels that were physically stable and biologically responsive. The dECM based-hydrogels supported the growth and proliferation of primary human lung fibroblast cells in 3D cultures. The dECM is also amenable to the coating of polyester membranes in Transwell™ Inserts to improve the cell adhesion, proliferation, and barrier function of primary human bronchial epithelial cells in 2D. In conclusion, we present a robust protocol for human lung decellularization, generation of dECM substrate material, and creation of hydrogels that support primary lung cell viability in 2D and 3D culture systems
Publisher: XMLink
Date: 2014
Publisher: Elsevier BV
Date: 2022
Publisher: Elsevier BV
Date: 12-2021
DOI: 10.1016/J.TIV.2021.105253
Abstract: The airway epithelium is exposed to a variety of air pollutants, which have been associated with the onset and worsening of respiratory diseases. These air pollutants can vary depending on their composition and associated chemicals, leading to different molecular interactions and biological effects. Mucociliary clearance is an important host defense mechanism against environmental air pollutants and this process is regulated by various ion transporters including the cystic fibrosis transmembrane conductance regulator (CFTR). With evidence suggesting that environmental air pollutants can lead to acquired CFTR dysfunction, it may be possible to leverage therapeutic approaches used in cystic fibrosis (CF) management. The aim of our study was to test whether environmental air pollutants tobacco smoke extract, urban particulate matter, and diesel exhaust particles lead to acquired CFTR dysfunction and whether it could be rescued with pharmacological interventions. Human airway epithelial cells (Calu-3) were exposed to air pollutant extracts for 24 h, with and without pharmacological interventions, with readouts of CFTR expression and function. We demonstrate that both tobacco smoke extract and diesel exhaust particles led to acquired CFTR dysfunction and that rescue of acquired CFTR dysfunction is possible with pharmacological interventions in diesel exhaust particle models. Our study emphasizes that CFTR function is not only important in the context of CF but may also play a role in other respiratory diseases impacted by environmental air pollutants. In addition, the pharmacological interventions approved for CF management may be more broadly leveraged for chronic respiratory disease management.
Publisher: Elsevier BV
Date: 03-2018
DOI: 10.1016/J.AJPATH.2017.11.006
Abstract: Marfan syndrome (MFS) is a genetic disorder that frequently leads to aortic root dissection and aneurysm. Despite promising preclinical and pilot clinical data, a recent large-scale study using antihypertensive angiotensin II (AngII) receptor type 1 (ATR1) blocker losartan has failed to meet expectations at preventing MFS-associated aortic root dilation, casting doubts about optimal therapy. To study the deleterious role of normal ATR1 signaling in aortic root widening, we generated MFS mice lacking ATR1a expression in an attempt to preserve protective ATR2 signaling. Despite being hypotensive and resistant to AngII vasopressor effects, MFS/ATR1a-null mice showed unabated aortic root enlargement and remained fully responsive to losartan, confirming that blood pressure lowering is of minor therapeutic value in MFS and that losartan's antiremodeling properties may be ATR1 independent. Having shown that MFS causes endothelial dysfunction and that losartan can activate endothelial function in mice and patients, we found that nitric oxide synthase (NOS) inhibition renders losartan therapeutically inactive, whereas multiple transgenic and pharmacologic models of endothelial NOS activation block aortic root dilation by correcting extracellular signal-regulated kinase signaling. In vitro, losartan can increase endothelial NO release in the absence of AngII and correct MFS NO levels in vivo. Our data suggest that increased protective endothelial function, rather than ATR1 inhibition or blood pressure lowering, might be of therapeutic significance in preventing aortic root disease in MFS.
Publisher: Informa UK Limited
Date: 12-05-2020
Publisher: European Respiratory Society (ERS)
Date: 2020
Publisher: Elsevier BV
Date: 04-2017
DOI: 10.1016/J.TRSL.2016.11.001
Abstract: Adipokines are mediators released from adipose tissue. These proteins are regarded as active elements of systemic and pulmonary inflammation, whose dysregulation can alter an in idual's risk of developing allergic lung diseases. Despite this knowledge, adipokine responses to inhaled stimuli are poorly understood. We sought to measure serum and lung adiponectin, leptin, and resistin in an atopic adult study population following exposure to allergen and diesel exhaust (DE). Two types of lung s les including bronchoalveolar lavage (BAL) and bronchial wash (BW), and a time course of serum s les, were collected from the 18 subjects who participated in the randomized, double-blinded controlled human study. The two crossover exposure triads in this study were inhaled DE and filtered air each followed by instilled allergen or saline. Serum and lung adipokine responses to these exposures were quantified using enzyme-linked immunosorbent assay. Allergen significantly increased adiponectin and leptin in BAL, and adiponectin in the BW 48 hours after exposure. Serum leptin and resistin responses were not differentially affected by exposure, but varied over time. Coexposure with DE and allergen revealed significant correlations between the adiponectin/leptin ratio and FEV
Publisher: Informa UK Limited
Date: 03-07-2015
DOI: 10.3109/01902148.2015.1040528
Abstract: The airway epithelium represents the first line of defense against inhaled environmental insults including air pollution, allergens, and viruses. Epidemiological and experimental evidence has suggested a link between air pollution exposure and the symptoms associated with respiratory viral infections. We hypothesized that multiple insults integrated by the airway epithelium NLRP3 inflammasome would result in augmented IL-1β release and downstream cytokine production following respiratory virus exposure. We performed in vitro experiments with a human airway epithelial cell line (HBEC-6KT) that involved isolated or combination exposure to mechanical wounding, PM10, house dust mite, influenza A virus, and respiratory syncytial virus. We performed confocal microscopy to image the localization of PM10 within HBEC-6KT and ELISAs to measure soluble mediator production. Airway epithelial cells secrete IL-1β in a time-dependent fashion that is associated with internalization of PM10 particles. PM10 exposure primes human airway epithelial cells to subsequent models of cell damage and influenza A virus exposure. Prior PM10 exposure had no effect on IL-1β responses to RSV exposure. Finally we demonstrate that PM10-priming of human airway epithelial cell IL-1β and GM-CSF responses to influenza A exposure are sensitive to NLRP3 inflammasome inhibition. Our results suggest the NLRP3 inflammasome may contribute to exaggerated immune responses to influenza A virus following periods of poor air quality. Intervention strategies targeting the NLRP3 inflammasome in at risk in iduals may restrict poor air quality priming of mucosal immune responses that result from subsequent viral exposures.
Publisher: American Chemical Society (ACS)
Date: 25-05-2023
Publisher: American Thoracic Society
Date: 30-10-2023
Publisher: Elsevier BV
Date: 04-2021
Publisher: Springer Science and Business Media LLC
Date: 10-02-2021
DOI: 10.1038/S41598-021-83189-X
Abstract: In many biological systems, pH can be used as a parameter to understand and study cell dynamics. However, measuring pH in live cell culture is limited by the sensor ion specificity, proximity to the cell surface, and scalability. Commercially available pH sensors are difficult to integrate into a small-scale cell culture system due to their size and are not cost-effective for disposable use. We made PHAIR—a new pH sensor that uses a micro-wire format to measure pH in vitro human airway cell culture. Tungsten micro-wires were used as the working electrodes, and silver micro-wires with a silver/silver chloride coating were used as a pseudo reference electrode. pH sensitivity, in a wide and narrow range, and stability of these sensors were tested in common standard buffer solutions as well as in culture media of human airway epithelial cells grown at the air–liquid interface in a 24 well cell culture plate. When measuring the pH of cells grown under basal and challenge conditions using PHAIR, cell viability and cytokine responses were not affected. Our results confirm that micro-wire-based sensors have the capacity for miniaturization and detection of erse ions while maintaining sensitivity. This suggests the broad application of PHAIR in various biological experimental settings.
Publisher: Springer Science and Business Media LLC
Date: 02-03-2021
DOI: 10.1186/S12931-021-01669-0
Abstract: Asthma was identified as the most common comorbidity in hospitalized patients during the 2009 H1N1 influenza pandemic. We determined using a murine model of allergic asthma whether these mice experienced increased morbidity from pandemic H1N1 (pH1N1) viral infection and whether blockade of interleukin-4 receptor α (IL-4Rα), a critical mediator of T h 2 signalling, improved their outcomes. Male BALB/c mice were intranasally sensitized with house dust mite antigen (Der p 1) for 2 weeks the mice were then inoculated intranasally with a single dose of pandemic H1N1 (pH1N1). The mice were administered intraperitoneally anti-IL-4Rα through either a prophylactic or a therapeutic treatment strategy. Infection with pH1N1 of mice sensitized to house dust mite (HDM) led to a 24% loss in weight by day 7 of infection (versus 14% in non-sensitized mice p .05). This was accompanied by increased viral load in the airways and a d ened anti-viral host responses to the infection. Treatment of HDM sensitized mice with a monoclonal antibody against IL-4Rα prior to or following pH1N1 infection prevented the excess weight loss, reduced the viral load in the lungs and ameliorated airway eosinophilia and systemic inflammation related to the pH1N1 infection. Together, these data implicate allergic asthma as a significant risk factor for H1N1-related morbidity and reveal a potential therapeutic role for IL-4Rα signalling blockade in reducing the severity of influenza infection in those with allergic airway disease.
Publisher: Elsevier BV
Date: 2005
DOI: 10.1016/J.JACI.2004.09.022
Abstract: There is increasing evidence that hemopoietic progenitor cells may traffic from bone marrow to sites of allergen exposure in asthma and undergo in situ differentiation, contributing to ongoing airway inflammation. However, the isolation and detailed phenotyping of true CD34 + progenitors from lung tissue during an allergen-induced airway eosinophilia has not been performed. We attempted to isolate and investigate the in vivo kinetics of hemopoietic progenitor cells and production of eosinophilopoietic mediators in the lung. In a mouse model of allergic airway inflammation, cells were extracted from lung tissue by enzymatic digestion. Total (CD34 + 45 + ) and eosinophil lineage committed (CD34 + 45 + IL-5Ralpha + ) progenitors were enumerated by flow cytometry. Outcome measurements were made 2, 6, 12, 24, 48, and 72 hours and 7 and 14 days after allergen challenge. Compared with saline control, CD34 + 45 + progenitors were elevated between 6 and 48 hours ( P .05). CD34 + 45 + IL-5Ralpha + progenitors were transiently elevated at 6 hours ( P < .05) before a return to preallergen levels by 12 hours and a subsequent increase at 14 days ( P < .05). Bronchoalveolar lavage eosinophils were increased at 2 hours, peaking at 72 hours ( P < .00625) and declining by 14 days. Both IL-5 and eotaxin levels were increased by 2 hours, peaking at 12 hours ( P < .05) and 24 hours ( P < .05), respectively. We propose that the increase in CD34 + 45 + IL-5Ralpha + cells and the eosinophilopoietic mediators IL-5 and eotaxin in the lung after allergen exposure may promote in situ differentiation of eosinophils that contribute to ongoing allergic airway inflammation.
Publisher: Elsevier BV
Date: 04-2014
DOI: 10.1016/J.JACI.2014.02.033
Abstract: Allergic inflammation involves the sensitization of naive CD4(+) T cells to allergens, resulting in a TH2-skewed inflammatory response. Although antigen presentation by dendritic cells to T cells in the lymph node is crucial for TH2 cell development, the innate signals that initiate adaptive type 2 inflammation and the role of group 2 innate lymphoid cells (ILC2s) are poorly understood. We sought to investigate the influence of ILC2s and the route of priming on the development of an adaptive type 2 immune response to lung allergens. Wild-type and ILC2-deficient mice were exposed intranasally or systemically to the TH2-inducing antigens house dust mite or ovalbumin in a model of allergic airway inflammation or the TH17-inducing bacterial antigen Saccharopolyspora rectivirgula in a model of hypersensitivity pneumonitis. The formation of an adaptive immune response was evaluated based on serum antibody titers and production of T cell-derived cytokines (IL-4, IL-5, IL-13 and IL-17A). We find that lung ILC2s play a critical role in priming the adaptive type 2 immune response to inhaled allergens, including the recruitment of eosinophils, TH2 cytokine production and serum IgE levels. Surprisingly, systemic priming with ovalbumin, with or without adjuvants, circumvents the requirement for ILC2s in inducing TH2-driven lung inflammation. ILC2s were also found to be dispensable for the sensitization to TH1- or TH17-inducing antigens. These data highlight a critical role for ILC2s in the development of adaptive type 2 responses to local, but not systemic, antigen exposure.
Publisher: Elsevier BV
Date: 12-2016
DOI: 10.1016/J.JACI.2016.02.038
Abstract: Air pollution's association with asthma may be due to its augmentation of allergenic effects, but the role of microRNA (miRNA) and gene expression in this synergy is unknown. We sought to determine whether exposure to allergen, exposure to diesel exhaust (DE), or coexposures modulate miRNA, gene expression, or inflammatory pathways and whether these measurements are correlated. Fifteen participants with atopy completed this controlled study of 2 hours of filtered air or DE (300 μg PM Robust linear models indicated that DE plus saline and DE plus allergen significantly modulated the highest number of miRNAs and messenger RNAs, respectively, relative to control (filtered air plus saline). In mixed models, allergen exposure modulated (q ≤ 0.2) miRNAs including miR-183-5p, miR-324-5p, and miR-132-3p and genes including NFKBIZ and CDKN1A, but DE did not significantly modify this allergenic effect. Repression of CDKN1A by allergen-induced miR-132-3p may contribute to shedding of bronchial epithelial cells. Expression of specific miRNAs and genes associated with bronchial immune responses were significantly modulated by DE or allergen. However, DE did not augment the effect of allergen at 48 hours, suggesting that adjuvancy may be transient or require higher or prolonged exposure. In silico analysis suggested a possible mechanism contributing to epithelial wall damage following allergen exposure.
Publisher: Elsevier BV
Date: 04-2012
DOI: 10.1016/J.JACI.2011.11.033
Abstract: The airway epithelium is the first line of defense against inhaled insults and therefore must be capable of coordinating appropriate inflammatory and immune responses. We sought to test the hypothesis that the nucleotide-binding domain and leucine-rich repeat protein 3 (NLRP3) inflammasome, an intracellular danger-sensing complex, plays a critical role in airway epithelium-mediated immune responses to urban particulate matter (PM) exposure. In this study we (1) identified NLRP3 and caspase-1 expression in human airway epithelium bronchus and primary cells, (2) characterized NLRP3 inflammasome-mediated IL-1β production from human airway epithelium in response to PM, and (3) performed in vivo PM exposure experiments with wild-type and Nlrp3(-/-) mice. Our results demonstrate that human airway epithelium contains a functional NLRP3 inflammasome that responds to PM exposure with caspase-1 cleavage and production of IL-1β. Exposure of Nlrp3(-/-) and wild-type mice to PM in vivo demonstrates NLRP3-dependent production of IL-1β in the lung, airway neutrophilia, and increases in CD11c(+hi)/MHC class II(+hi) cell numbers in intrathoracic lymph nodes. Our study is the first to characterize airway epithelial NLRP3 inflammasome-mediated immune responses to PM exposure, which might have implications in patients with asthma and other lung diseases.
No related grants have been discovered for Jeremy Hirota.