Climate change-driven increases in forest fires pose a major global health risk due to exposure to smoke containing hazardous gases and fine particulates, emphasizing the need for physiologically relevant in vitro airway models for studying smoke-induced responses. Microfluidic lung-on-a-chip technologies provide a strong foundation for in vitro airway modeling and ongoing developments are expanding their ability to incorporate multicellular organization, extracellular matrix complexity, and physiologically relevant exposure methods. This work presents the optimization and integration of a photopolymerizable gelatin methacrylate (GelMA)-based hydrogel into a microfluidic airway-on-a-chip that models the human small conducting airways and supports controlled aerosol exposure to wood smoke. The GelMA hydrogel was optimized to support fibroblast encapsulation, endothelial adhesion, and robust mechanical stability. The device combines the hydrogel with a compartmentalized microchannel layout, and sacrificial molding to create a 3D organotypic airway culture featuring a multilayer architecture, 3D stromal matrix, and a perfusable vasculature-like lumen. Coupling the platform with a custom aerosol exposure system enables precise, biomimetic exposure to whole wood smoke. Proof-of-concept studies using transforming growth factor beta 1 (TGF-β1) and whole wood smoke elicited expected inflammatory and fibrotic responses, validating the platform’s physiological relevance for inhalation studies and investigating smoke-induced airway remodeling and inflammation.

Traffic-related air pollution (TRAP) exposure is associated with adverse health effects, including chronic inflammation and the exacerbation of respiratory diseases. Although the effects of TRAP exposure begin locally in the lungs, they can spread systemically throughout the body via the blood. To understand blood-lung dynamics upon TRAP exposure, we interrogated blood and airway gene expression and performed a concordant analysis of these datasets. Specifically, we investigated the transcriptomic response of the blood and airway epithelium to inhaled diesel exhaust (DE), an experimental model of TRAP, across thirty-two research participants. In this double-blinded, crossover, controlled human exposure study, participants were exposed to DE and filtered air for two hours on two separate occasions. Paired-end RNA-sequencing was conducted on blood and bronchial brushing samples. Differential gene expression analysis revealed significant gene expression changes in airways, including upregulation of GPX2 and NQO1, genes essential to antioxidant defense. Differentially expressed genes (DEGs) were mapped to hallmark pathways and cell types using the Human Molecular Signatures Database (MSigDB). In the blood and airway epithelium, pathways with altered activity included interferon alpha (IFN-α) response, interferon gamma (IFN-γ) response, inflammatory response, and reactive oxygen species (ROS) pathway. Concordant analysis revealed correlated cellular activity between blood and airways, providing new evidence of a potential blood-lung relationship in response to DE exposure, which was not detectable through independent analysis alone. Combining local and systemic data may enrich our understanding of the downstream biological pathways affected by DE exposure, allowing for new avenues to monitor lung responses through blood analysis.

Climate change is increasing the frequency and severity of wildfires globally, causing significant woodsmoke (WS) emissions. Vehicles emit sizable amounts of toxic traffic-related air pollution (TRAP), for which diesel exhaust (DE) is a model. Both WS and DE contain particulate matter < 2.5 microns (PM2.5), which deeply penetrates the lungs causing respiratory epithelial inflammation that drives health effects. Regulations focus on PM2.5 concentration, despite emerging research that highlights how composition mediates health effects. As WS and DE are compositionally distinct, we conducted the first head-to-head comparison of effects on the transcriptomes of air-liquid interface cultured primary human bronchial epithelial cells (HBEC). Differentiated donor-matched HBEC transwells were exposed for 2-hours to filtered air (FA; control), or WS (furnace tube burning pine) or DE (Hatz 1B30E generator) both diluted to 300 µg/m3 of PM2.5. WS had higher ultrafine PM, whereas DE exposure contained significantly higher NO2, CO, and O3. RNA sequencing showed that WS exposure resulted in 159 (↑50, ↓109) differentially expressed genes, while DE modulated 439 (↑264, ↓175) compared to FA exposure. WS was associated with small ribosomal subunit and cytochrome complex related genes, while DE exposure was associated with HIF-1 signaling, respiratory chain complex and interferon alpha/beta signaling/ISG15-protein conjugation, suggesting how TRAP exposure may enhance infection risk. We also analyzed exposure effects on protein immune-mediators. We demonstrate that two major air pollution sources modulate different genes and pathways in HBECs, with minimal overlap. This informs the debate regarding the regulatory focus on concentration and assumptions that similar concentrations of air pollution have indistinct effects.

Respiratory tract infections have been linked to air pollution exposure, and both are known exogenous risk factors of asthma exacerbations. The current frontline therapy used to minimize exacerbations are inhaled corticosteroids (ICS), but these medications increase the risk of respiratory tract infections in patients due to immunosuppressive side-effects. Virally-triggered exacerbations are most commonly associated with rhinovirus, and the ICS fluticasone propionate (FP) has been demonstrated to increase rhinovirus viral load by 2.5-fold in infected epithelial cells. We hypothesized that air pollution exposure combined with ICS would magnify effects on host-defence antiviral responses and viral replication. Using rhinovirus (RV16), we infected human bronchial epithelial (BEAS-2B) cells that were pre-treated with FP (250 nM) and exposed to diesel exhaust particles (DEP) SRM2975 (50 μg/cm2). We show that DEP exposure significantly increased RV16 viral RNA by 12-fold over untreated cells 24 h post-infection, while FP alone induced a 2-fold rise. However, when combined, FP and DEP induced a significant supra-additive 17-fold increase in RV16 RNA. In addition, we demonstrate that DEP induces the expression of the RV16 entry receptor ICAM-1 through the canonical NF-κB pathway and that suppression of this pathway results in attenuation of RV16 viral expression. Our findings suggest a mechanism through which combined epithelial exposure to DEP and ICS increases RV16 infectivity through the suppression of innate antiviral immune responses and induction of the RV16 entry receptor ICAM-1. These findings suggest a need for caution during periods of high air pollution by those managing chronic lung diseases with corticosteroids.

Background: The airway epithelium plays a crucial role as a mucosal barrier against environmental challenges, including traffic-related air pollution and wildfire smoke – the two most common exposures in North America. By utilizing an in-vitro exposure cell culture model and transcriptomic analysis to assess the effects of diesel exhaust (DE; a model for traffic-related air pollution) and wood smoke (WS; a model for wildfires), we can gain a comprehensive understanding of the similarities and differences in how each exposure impacts the respiratory tract. Methods: Human bronchial epithelial cells (hBEC) were collected from six healthy never-smokers undergoing a research bronchoscopy. Cells were cultured, expanded, and differentiated at air-liquid interface (ALI) for >21 days. Differentiated hBECs were exposed to filtered air (control condition), diesel exhaust (diluted to PM2.5 = 300 μg/m3), or woodsmoke (diluted to PM2.5 = 300 μg/m3) for 2 hours using a CULTEX in-vitro exposure system. Twenty-four hours after each exposure, the cells were harvested for RNA sequencing. Total RNA was extracted, followed by PolyA mRNA enrichment, cDNA synthesis, and sequencing library generation. Paired-end 150bp Illumina NovaSeq sequencing targeting 50 million read-pairs per library was then performed. After preprocessing, we conducted differential gene expression and pathway analysis through the nf-core RNA-seq pipeline before conducting DESeq2 differential expression analysis and Gene Set Enrichment Analysis (GSEA). Results: Transcriptome analysis revealed 283 (Up:162; Down:121) differentially expressed genes (DEGs) in DE-exposed cells compared to FA exposure (false discovery rate (FDR) <0.05). In contrast, there were 83 (Up:24; Down:59) DEGs in WS-exposed cells compared to control. Both DE and WS exposures elicited changes in 18 common differentially expressed transcripts (Up:5; Down:13). GSEA analysis showed enrichment of 33 and 12 KEGG Pathways in the DE and WS exposed cells, respectively, based on DEGs. GSEA further revealed shared enriched pathways (including oxidative phosphorylation and chemical carcinogenesis) between the DE and WS-exposed cells. No significant differences in cytotoxicity (LDH assay) or barrier function (transepithelial electrical resistance) were observed when comparing the FA condition to the exposure groups. Conclusions: This study demonstrates the utility of a systems biology approach in uncovering distinct and shared transcriptional responses of bronchial epithelial cells to two extremely common environmental exposures. Given some public perception of WS as inconsequential, along with alternative assumptions that similar concentrations of DE and WS likely have indistinct effects, these findings highlight the importance of understanding the intricacies of complex pollution-induced changes in airway health.

BACKGROUND:Loss-of-function (LOF) alterations in tumour suppressor genes cannot be directly targeted. Approaches characterising gene function and vulnerabilities conferred by such mutations are required. METHODS:Here, we computationally map genetic networks of KMT2D, a tumour suppressor gene frequently mutated in several cancer types. Using KMT2D loss-of-function (KMT2DLOF) mutations as a model, we illustrate the utility of in silico genetic networks in uncovering novel functional associations and vulnerabilities in cancer cells with LOF alterations affecting tumour suppressor genes. RESULTS:We revealed genetic interactors with functions in histone modification, metabolism, and immune response and synthetic lethal (SL) candidates, including some encoding existing therapeutic targets. Notably, we predicted WRN as a novel SL interactor and, using recently available WRN inhibitor (HRO761 and VVD-133214) treatment response data, we observed that KMT2D mutational status significantly distinguishes treatment-sensitive MSI cell lines from treatment-insensitive MSI cell lines. CONCLUSIONS:Our study thus illustrates how tumour suppressor gene LOF alterations can be exploited to reveal potentially targetable cancer cell vulnerabilities.

The evidence associating traffic-related air pollution (TRAP) with allergic asthma is growing, but the underlying mechanisms for this association remain unclear. The airway epithelium is the primary tissue exposed to TRAP, hence understanding its interactions with TRAP and allergen is important. Diesel exhaust (DE), a paradigm of TRAP, consists of particulate matter (PM) and gases. Modern diesel engines often have catalytic diesel particulate filters to reduce PM output, but these may increase gaseous concentrations, and their benefits on human health cannot be assumed. We conducted a randomized, double-blinded, crossover study using our unique in vivo human exposure system to investigate the effects of DE and allergen co-exposure, with or without particle depletion as a proxy for catalytic diesel particulate filters, on the airway epithelial transcriptome. Participants were exposed for 2 h before an allergen inhalation challenge, with each receiving filtered air and saline (FA-S), filtered air and allergen (FA-A), DE and allergen (DE-A), or particle-depleted DE and allergen (PDDE-A), over four different occasions, each separated by a 4-week washout period. Endobronchial brushings were collected 48 h after each exposure, and total RNA was sequenced. Differentially expressed genes (DEGs) were identified using DESeq2, followed by GO enrichment and pathway analysis. FA-A, DE-A, and PDDE-A exposures significantly modulated genes relative to FA-S, with 462 unique DEGs identified. FA-A uniquely modulated the highest number (↑178, ↓155), followed by DE-A (↑44, ↓23), and then PDDE-A exposure (↑15, ↓2); 6 DEGs (↑4, ↓2) were modulated by all three conditions. Exposure to PDDE-A resulted in modulation of 285 DEGs compared to DE-A exposure, further revealing 26 biological process GO terms, including "cellular response to chemokine" and "inflammatory response". The transcriptional epithelial response to diesel exhaust and allergen co-exposure is enriched in inflammatory mediators, the pattern of which is altered upon particle depletion.

AbstractLoss-of-function (LOF) alterations in tumour suppressor genes cannot be directly targeted. Approaches characterising gene function and vulnerabilities conferred by such mutations are required. Here, we computationally map genetic networks ofKMT2D, a tumour suppressor gene frequently mutated in several cancer types. UsingKMT2Dloss-of-function (KMT2DLOF) mutations as a model, we illustrate the utility ofin silicogenetic networks in uncovering novel functional associations and vulnerabilities in cancer cells with LOF alterations affecting tumour suppressor genes. We revealed genetic interactors with functions in histone modification, metabolism, and immune response, and synthetic lethal (SL) candidates, including some encoding existing therapeutic targets. Analysing patient data from The Cancer Genome Atlas and the Personalized OncoGenomics Project, we showed, for example, elevated immune checkpoint response markers inKMT2DLOFcases, possibly supportingKMT2DLOFas an immune checkpoint inhibitor biomarker. Our study illustrates how tumour suppressor gene LOF alterations can be exploited to reveal potentially targetable cancer cell vulnerabilities.

BackgroundEnvironmental co-exposure to allergen and traffic-related air pollution is common globally and contributes to the exacerbation of respiratory diseases. Individual responses to environmental insults remain variable due to gene-environment interactions.ObjectiveThis study examined whether single nucleotide polymorphisms (SNPs) in lung cell surface receptor genes modifies lung function change and immune cell recruitment in allergen-sensitized individuals exposed to diesel exhaust (DE) and allergen.MethodsIn this randomized, double-blinded, four-arm, crossover study, 13 allergen-sensitized participants underwent allergen inhalation challenge following a 2-hour exposure to DE, particle-depleted diesel exhaust (PDDE) or filtered air (FA). Lung function tests and bronchoscopic sample collection were performed up to 48 h after exposures. Transient receptor potential channel (TRPA1 and TRPV1) and toll-like receptor (TLR2 and TLR4) risk alleles were used to construct an unweighted genetic risk score (GRS). Exposure-by-GRS interactions were tested using mixed-effects models.ResultsIn participants with high GRS, allergen exposure was associated with an increase in airway hyperresponsiveness (AHR) when co-exposed to PDDE (p = 0.03) but not FA or DE. FA and PDDE also were associated with a relative increase in macrophages and decrease in lymphocytes in bronchoalveolar lavage.ConclusionsTRPs and TLRs variants are associated with increased AHR and altered immune cellularity in allergen-exposed individuals. This effect is blunted by DE exposure, suggesting greater influence of unmeasured gene variants as primary meditators of a particulate-rich co-exposure.Trial registrationThe study was registered with ClinicalTrials.gov on December 20, 2013 (NCT02017431).

A complete understanding of how exposure to environmental substances promotes cancer formation is lacking. More than 70 years ago, tumorigenesis was proposed to occur in a two-step process: an initiating step that induces mutations in healthy cells, followed by a promoter step that triggers cancer development 1 . Here we propose that environmental particulate matter measuring ≤2.5 μm (PM 2.5 ), known to be associated with lung cancer risk, promotes lung cancer by acting on cells that harbour pre-existing oncogenic mutations in healthy lung tissue. Focusing on EGFR-driven lung cancer, which is more common in never-smokers or light smokers, we found a significant association between PM 2.5 levels and the incidence of lung cancer for 32,957 EGFR-driven lung cancer cases in four within-country cohorts. Functional mouse models revealed that air pollutants cause an influx of macrophages into the lung and release of interleukin-1β. This process results in a progenitor-like cell state within EGFR mutant lung alveolar type II epithelial cells that fuels tumorigenesis. Ultradeep mutational profiling of histologically normal lung tissue from 295 individuals across 3 clinical cohorts revealed oncogenic EGFR and KRAS driver mutations in 18% and 53% of healthy tissue samples, respectively. These findings collectively support a tumour-promoting role for PM 2.5 air pollutants and provide impetus for public health policy initiatives to address air pollution to reduce disease burden.

The management of asthma and chronic obstructive pulmonary disease (COPD) is guided by the Global Initiative for Asthma and Global Obstructive Lung Disease guidelines, respectively (1, 2). In both the Global Initiative for Asthma and Global Obstructive Lung Disease guidelines, pharmacological interventions that increase levels of intracellular cAMP are used with the primary intention of relaxing airway smooth muscle and opening the airways. Indeed, the earliest asthmamedications reported include adrenaline, which activates the b-2-adrenoceptor and downstream adenylyl cyclase production of cAMP, and caffeine, which functions as a PDE inhibitor to prevent cAMP degradation (3). Elevations in intracellular cAMP levels lead to smooth muscle relaxation by multiple mechanisms that may include cAMP-dependent PKA (protein kinase A) activation, PKAindependent activation of exchange proteins, and decrease of intracellular calcium concentrations and associated signaling (4). In this issue of the Journal (pp. 96–106), Cao and colleagues report that ABCC1 (MRP1), a member of the ATP-binding cassette transporter family, represents an additional candidate target potentially important in normalization of cAMP signaling and second messenger function in airway smooth muscle cells, with potential implications for patients with asthma and COPD (5). Cao and colleagues used primary human airway smooth muscle cells from tracheas in observational and mechanistic experimental approaches to characterize the expression and function of ABCC1. Specifically, ABCC1 gene and protein expression were confirmed by real-time PCR, immunohistochemistry, and immunoblot of cultured primary human airway smooth muscle cells and corroborated with publicly available RNA sequencing data generated from a distinct set of human airway smooth muscle cells. These foundational observational steps were complemented by mechanistic approaches with siRNA knockdown of ABCC1 transcripts and pharmacological interventions. Using this multifaceted approach, the authors demonstrate prominent ABCC1 expression in primary human airway smooth muscle cells. Furthermore, functional cAMP transport assays with pharmacological interventions and siRNA knockdown were used to mechanistically implicate ABCC1 in cAMP efflux after activation of adenylyl cyclase. The cytoskeleton properties of human airway smooth muscle cells also were explored with magnetic twisting cytometry, another mechanistic approach. Collectively, their findings provide a compelling foundational demonstration of the expression and function of ABCC1 in human airway smooth muscle. The major translational findings of ABCC1 function as a cAMP exporter in human airway smooth muscle present a new diseasemodifying target that could be relevant in both asthma and COPD. Importantly, the principle behind ABCC1 as a new target is grounded in decades of clinical asthma and COPDmanagement that focused on cAMPmodulation. Building on that concept, the authors expand the two-dimensional approach of cAMPmodulation (elevating and blocking breakdown) by adding the third dimension of extracellular transport. The novel concept that cAMP signaling is regulated by mechanisms that go beyond receptor activation and enzymatic degradation to include intracellular compartmentalization and transport requires deeper exploration (6), which will likely arise from the presented work. A major strength of the study is the use of primary human airway smooth muscle cells and publicly available data sets for complementary confirmation of ABCC1 expression, which provide a solid foundation for the authors to perform their mechanistic studies. Their demonstration of the specificity of the antibodies used for detecting ABCC1 protein and the localization of ABCC1 at the cell membrane represent important validations. The use of bothmolecular (siRNA) and pharmacological (e.g., MK-571) interventions to both identify ABCC1 as the primary cAMP efflux transporter and target the protein’s function are also strengths of the study. Their complementary approaches are valuable because neither approach alone is absolute— complete knockdown efficiency is challenging in primary cell culture systems andmany pharmacological interventions have off-target effects. Their study does have some limitations due to the exclusive focus on airway smoothmuscle cells. The demonstration that ABCC1 functions as a cAMP transporter in human airway smoothmuscle cells warrants the exploration of this molecule’s expression and function in other lung and immune cells that may be important in asthma and COPD, including airway epithelial cells, where we have observed robust gene expression of this transporter (7). Importantly, ABCC1 has broad substrate specificity and can transport a variety of endogenous and exogenous molecules (8). The consequences of interventions targeting ABCC1may therefore extend beyond airway smoothmuscle relaxation and could impair other biological processes important in normal health. In addition, although the elevation of cAMP in airway smooth muscle via ABCC1 is grounded in decades of historical clinical practice with cAMPmodulators (3), more recent evidence suggests that other cells, including airway epithelial cells, can respond to cAMP elevation with potentially deleterious proinflammatory consequences (9). Finally, of interest is the exploration of cAMP as a serum biomarker, although it remains to be determined if a mechanistic link exists between the modulation of cAMP efflux in the lung tissue and levels in circulation. The use of cAMP elevating agents alone for the management of asthma has recently been revised in a change that strongly supports combinatorial therapies that include glucocorticoids (1). The interaction between cAMP elevating agents and glucocorticoids is additive and potentially synergistic, as suggested by studies exploring mechanisms of bronchodilation and antiinflammatory responses (10, 11).

Background and objectives: Neutrophils are key mediators of the innate immune response but also contribute to deleterious inflammation in the lungs. Particulate matter (PM) in traffic-related air pollution (TRAP) recruits neutrophils to the airways and is associated with increased respiratory morbidity. The mechanisms and consequences of TRAP-neutrophil interactions within the airways are incompletely understood. This study aims to investigate how PM may contribute to neutrophil-driven airways pathology. Methods: Neutrophils were isolated from the blood of healthy volunteers and cultured with diesel particulate matter (PM). They were analysed for activation status, reactive oxygen species (ROS) generation and lysosomal activity. Results: Exposure to PM results in neutrophil activation (Fig 1A), priming for ROS generation (Fig 1B) and inhibited LPS induced lysosomal protein catabolism (Fig 1C). Conclusions: Taken together, these results show that exposure to TRAP-related PM results in neutrophil activation and priming. Furthermore, PM impairs neutrophil lysosomal activity. This is likely to have important consequences for a range of neutrophil functions, including metabolism and bacterial killing. Along with our previous findings on TRAP-altered host defence peptides, these changes are likely to ultimately upset the delicate balance of immunity and inflammation in the airways.

Phthalates are ubiquitous environmental contaminants associated with allergic disease in epidemiological and animal studies. This investigation aims to support these associations by interrogating systemic immune effects in allergen-sensitized volunteers after controlled indoor air exposure to a known concentration of dibutyl phthalate (DBP). The phthalate-allergen immune response (PAIR) study enrolled 16 allergen-sensitized participants to a double-blinded, randomized, crossover exposure to two conditions (DBP or control air for 3 hr), each followed immediately by inhaled allergen challenge. Peripheral blood immune cell composition and activation along with inflammatory mediators were measured before and after exposure. DBP exposure prior to the inhaled allergen challenge increased the percentage of CD4+ T helper cells and decreased the percentage of regulatory T cells (3 hr and 20 hr post-exposure), while only modest overall effects were observed for inflammatory mediators. The cells and mediators affected by the phthalate exposure were generally not overlapping with the endpoints affected by allergen inhalation alone. Thus, in distinction to our previously published effects on lung function, DBP appears to alter endpoints in peripheral blood that are not necessarily enhanced by allergen alone. Further studies are needed to clarify the role of phthalate-induced systemic effects in disease pathogenesis.

With prevalent global air pollution, individuals with certain genetic predispositions and sensitivities are at of higher risk of developing respiratory symptoms including chronic cough. Studies to date have relied on patient-filled questionnaires in epidemiological studies to evaluate the gene-by-environment interactions. In a controlled human exposure study, we evaluated whether genetic risk score (GRS) based on cough-related single-nucleotide polymorphisms (SNPs) are associated with a cough count over 24 h post-exposure to diesel exhaust (DE), a model for traffic-related air pollution. DE is a mixture of several known air pollutants including PM2.5, CO, NO, NO2, and volatile organic compounds. Under closely observed circumstances, we determined that GRS constructed from 7 SNPs related to TRPA1, TRPV1, and NK-2R were correlated with cough count. Selection of channels were based on prior knowledge that SNPs in these channels lead to acute airway inflammation as a result of their increased sensitivity to particulate matter. We performed a linear regression analysis and found a significant, positive correlation between GRS and cough count following DE exposure (p = 0.002, R2 = 0.61) and filtered air (FA) exposure (p = 0.028, R2 = 0.37). Although that correlation was stronger for DE than for FA, we found no significant exposure-by-GRS interaction. In summary, cough-relevant GRS was associated with a higher 24 h cough count in a controlled setting, suggesting that individuals with a high GRS may be more susceptible to developing cough regardless of their exposure. The trend towards this susceptibility being more prominent in the context of traffic-related air pollution remains to be confirmed. Trial registration: ClinicalTrial.gov NCT02236039; NCT0223603. Registered on August 11, 2014, https://clinicaltrials.gov/ct2/show/NCT02236039 .

Background: Environmental co-exposure to traffic related air pollution and allergen is common globally. However, individual responses to these environmental insults remain variable, possibly due to individual gene-environment interactions. Objective: Determine whether a genetic risk score (GRS) calculated from single nucleotide polymorphisms in lung cell surface receptor genes implicated in mediating inflammatory responses to environmental pollutants modified lung function in individuals exposed to diesel exhaust (DE) and allergen. Methods: In a randomized crossover study, 13 allergen-sensitized participants were exposed to diesel exhaust (DE; PM2.5 concentration of 300 µg/m3), particle-depleted diesel exhaust (PDDE), or filtered air (FA) for 2 hours prior to undergoing an inhaled allergen challenge. Blood samples, spirometry, methacholine challenge, and bronchoscopy were performed up to 48 hours after exposures. Transient receptor potential channel (TRPA1 and TRPV1) and toll-like receptor (TLR2 and TLR4) risk alleles were used to construct an unweighted GRS. Effect modification by condition and GRS was determined relative to the baseline FA and saline co-exposure using mixed effect models. Results: High GRS and allergen was associated with a significant increase in 24 hour airway hyperresponsiveness (AHR) when co-exposed to FA and PDDE (P = 0.04 and 0.02 respectively) but not DE. Increased AHR in FA and PDDE conditions was associated with increased bronchoalveolar macrophages and decreased lymphocytes. Conclusions: Variant alleles of the TRP and TLR receptors predispose allergen-exposed individuals to increased AHR and changes in airway immune cells.

Background: Exposure to traffic-related air pollution (TRAP) is linked to COPD morbidity and mortality, suggesting airway susceptibility in those with COPD exposed to TRAP. Objective: Evaluate the impact of TRAP exposure on the epithelial transcriptome in human bronchioles. Methods: We recruited healthy never-smokers (NS; N=9), ex-smokers (ES; N=7) with >10 pack-year smoking but not COPD, and GOLD stage 1&2 COPD participants (COPD; N=9). In an order-randomized crossover study, each participant was exposed to diesel exhaust (DE; PM2.5 concentration of 300 µg/m3) and filtered air (control) for 2 hours. 24 hours post-exposure, bronchoscopy was performed to collect endobronchial brushes. PolyA RNA sequencing was performed. Results: DE significantly modulated expression of 35, 131 and 10 genes in NS, ES and COPD epithelium, respectively (Fig 1). The effect of DE on139 genes was significantly modified by COPD status. Differentially expressed genes common to NS and ES were GPX2, PSMC1P1 and TRIM16L, and functional enrichment analysis revealed NRF2 pathway activation. Conclusions: Exposure to TRAP, typical of congested urban areas, increases expression of antioxidant genes in the airway epithelium in vivo, and this expression is modified in COPD epithelium.