Air-liquid Interface Exposure Research Articles

Fifteen day repeat air: liquid Interface air-only exposures can cause respiratory epithelium injury in MucilAir™ nasal respiratory epithelial cells that parallels chemically induced cytotoxicity

New Approach Methodologies (NAMs) are being widely used to reduce, refine, and replace, animal use in studying toxicology. For respiratory toxicology, this includes in silico and in vitro alternatives using air:liquid interface (ALI) exposures to replace traditional in vivo inhalation studies. In previous studies using 1,3-dichloropropene (1,3-DCP), a 5-day 4 h repeat exposures of MucilAir™ nasal cell culture models caused, dose-dependent cytotoxicity, depletion of GSH, changes in differential gene expression and histopathological transitions in cellular morphology from pseudostratified columnar epithelium to squamous epithelium. In this report we attempted to extend these studies using 15-day 1,3-DCP 4 h exposures to using MucilAir™ nasal cultures as outlined by an US EPA recent task order (US EPA 2023). For the 15-day repeat exposure, there were severe histopathologic changes in the MucilAir™ nasal mock-treatment (air-only) VITROCELL® chamber controls compared to incubator controls preventing any further analysis. The histopathological transitions in cellular morphology from pseudostratified columnar epithelium to squamous epithelium observed in the air only control in this study and previously with 1,3-DCP in MucilAir™ nasal cultures is also a hallmark of chemically induced cytotoxic responses in vivo in the respiratory tract. Histopathology assessments of 3D respiratory tract models used in ALI exposures can provide the linkage between in vitro to in vivo outcomes as part of the validation efforts of ALI use in regulatory toxicology. This report indicates that importance of histopathological assessments of incubator and mock-treatment (air-only) controls from each ALI exposure experiment along with exposed cell based model.

Toxicological evaluation of primary particulate matter emitted from combustion of aviation fuel

Recently, Sustainable Aviation Fuel (SAF) blends and novel combustion technologies have been introduced to reduce aircraft engine emissions. However, there is limited knowledge about the impact of combustion technology and fuel composition on toxicity of primary Particulate Matter (PM) emissions, comparable to regulated non-volatile PM (nvPM).In this study, primary PM was collected on filters using a standardised approach, from both a Rich-Quench-Lean (RQL) combustion rig and a bespoke liquid fuelled Combustion Aerosol Standard (CAST) Generator burning 12 aviation fuels including conventional Jet-A, SAFs, and blends thereof. The fuels varied in aromatics (0–25.2%), sulphur (0–3000 ppm) and hydrogen (13.43–15.31%) contents. Toxicity of the collected primary PM was studied in vitro utilising Air-Liquid Interface (ALI) exposure of lung epithelial cells (Calu-3) in monoculture and co-culture with macrophages (differentiated THP-1 cells). Cells were exposed to PM extracted from filters and nebulised from suspensions using a cloud-based ALI exposure system. Toxicity readout parameters were analysed 24 h after exposure.Results showed presence of genotoxicity and changes in gene expression at dose levels which did not induce cytotoxicity. DNA damage was detected through Comet assay in cells exposed to CAST generated samples. Real-Time PCR performed to investigate the expression profile of genes involved in oxidative stress and DNA repair pathways showed different behaviours after exposure to the various PM samples. No differences were found in pro-inflammatory interleukin-8 secretion. This study indicates that primary PM toxicity is driven by wider factors than fuel composition, highlighting that further work is needed to substantiate the full toxicity of aircraft exhaust PM inclusive of secondary PM emanating from numerous engine technologies across the power range burning conventional Jet-A and SAF.

An original device to assess the respiratory impact of indoor air VOCs mixture using an in vitro approach

Exposure to indoor air pollution, particularly volatile organic compounds (VOCs), has been recognized as a risk factor in the development of respiratory and allergic diseases. VOCs are mainly emitted continuously at low concentrations from construction furniture and decoration products. Measurement campaigns carried out in new dwellings in France have shown that aldehydes predominate with a tendency to decrease formaldehyde concentrations and to increase those in hexanal. As the main route of VOCs exposure is inhalation, this project assessed the impact of a mixture of 17 VOCs representative of indoor air (in quality and quantity) on respiratory health using an in vitro approach. This original work was based on the set-up of an experimental device, combining a gas generation and dilution bench and exposure to the air-liquid interface (ALI) adapted to the reconstructed human airway epithelium model. The VOC mixture was enriched with formaldehyde or hexanal in different proportions (from 20 to 240 µg.m−3) to study the biological impact of these aldehydes after repeated exposures of airway epithelium. After examination of the stability of the VOC concentrations in generated mixtures and the found of the optimal operating conditions for the dynamic gas generating system, the gaseous mixtures were distributed to the epithelium using the ALI exposure system providing direct contact between the epithelium and the tested mixtures. Our device lead to reproduce real conditions of human exposure. The results showed that the inflammatory response, assessed by the production of four cytokines, varied according to the nature of the aldehyde present in the VOC mixture (formaldehyde or hexanal), its concentration, and the duration and number of exposures applied. The most original and innovative results concern those obtained with hexanal, pollutant under-researched. Our results showed that this aldehyde could pose risks to respiratory health.

Air-liquid interface exposure of A549 human lung cells to characterize the hazard potential of a gaseous bio-hybrid fuel blend.

Gaseous and semi-volatile organic compounds emitted by the transport sector contribute to air pollution and have adverse effects on human health. To reduce harmful effects to the environment as well as to humans, renewable and sustainable bio-hybrid fuels are explored and investigated in the cluster of excellence "The Fuel Science Center" at RWTH Aachen University. However, data on the effects of bio-hybrid fuels on human health is scarce, leaving a data gap regarding their hazard potential. To help close this data gap, this study investigates potential toxic effects of a Ketone-Ester-Alcohol-Alkane (KEAA) fuel blend on A549 human lung cells. Experiments were performed using a commercially available air-liquid interface exposure system which was optimized beforehand. Then, cells were exposed at the air-liquid interface to 50-2000 ppm C3.7 of gaseous KEAA for 1 h. After a 24 h recovery period in the incubator, cells treated with 500 ppm C3.7 KEAA showed significant lower metabolic activity and cells treated with 50, 250, 500 and 1000 ppm C3.7 KEAA showed significant higher cytotoxicity compared to controls. Our data support the international occupational exposure limits of the single KEAA constituents. This finding applies only to the exposure scenario tested in this study and is difficult to extrapolate to the complex in vivo situation.

Modular air–liquid interface aerosol exposure system (MALIES) to study toxicity of nanoparticle aerosols in 3D-cultured A549 cells in vitro

We present a novel lung aerosol exposure system named MALIES (modular air–liquid interface exposure system), which allows three-dimensional cultivation of lung epithelial cells in alveolar-like scaffolds (MatriGrids®) and exposure to nanoparticle aerosols. MALIES consists of multiple modular units for aerosol generation, and can be rapidly assembled and commissioned. The MALIES system was proven for its ability to reliably produce a dose-dependent toxicity in A549 cells using CuSO4 aerosol. Cytotoxic effects of BaSO4- and TiO2-nanoparticles were investigated using MALIES with the human lung tumor cell line A549 cultured at the air–liquid interface. Experiments with concentrations of up to 5.93 × 105 (BaSO4) and 1.49 × 106 (TiO2) particles/cm3, resulting in deposited masses of up to 26.6 and 74.0 µg/cm2 were performed using two identical aerosol exposure systems in two different laboratories. LDH, resazurin reduction and total glutathione were measured. A549 cells grown on MatriGrids® form a ZO-1- and E-Cadherin-positive epithelial barrier and produce mucin and surfactant protein. BaSO4-NP in a deposited mass of up to 26.6 µg/cm2 resulted in mild, reversible damage (~ 10% decrease in viability) to lung epithelium 24 h after exposure. TiO2-NP in a deposited mass of up to 74.0 µg/cm2 did not induce any cytotoxicity in A549 cells 24 h and 72 h after exposure, with the exception of a 1.7 fold increase in the low exposure group in laboratory 1. These results are consistent with previous studies showing no significant damage to lung epithelium by short-term treatment with low concentrations of nanoscale BaSO4 and TiO2 in in vitro experiments.

An integrated new approach methodology for inhalation risk assessment of safe and sustainable by design nanomaterials

The production and use of nanomaterials (NMs) has increased over the last decades posing relevant questions on their risk after release and exposure of the population or sub-populations. In this context, the safe and sustainable by design (SSbD) approach framework requires to assess the potential hazard connected with intrinsic properties of the material along the whole life cycle of the NM and/or of the nano enabled products. Moreover, in the last years, the use of new advanced methodologies (NAMs) has increasingly gained attention for the use of alternative methods in obtaining relevant information on NMs hazard and risk. Considering the SSbD and the NAMs frameworks, within the ASINA H2020 project, we developed new NAMs devoted at improving the hazard and risk definition of different Ag and TiO2 NPs. The NAMs are developed considering two air liquid interface exposure systems, the Vitrocell Cloud-α and the Cultex Compact module and the relevant steps to obtain reproducible exposures are described. The new NAMs build on the integration of environmental monitoring campaigns at nano-coating production sites, allowing the quantification by the multiple-path particle dosimetry (MPPD) model of the expected lung deposited dose in occupational settings. Starting from this information, laboratory exposures to the aerosolized NPs are performed by using air liquid interface exposure equipment and human alveolar cells (epithelial cells and macrophages), replicating the doses of exposure estimated in workers by MPPD. Preliminary results on cell viability and inflammatory responses are reported. The proposed NAMs may represent possible future reference procedures for assessing the NPs inhalation toxicology, supporting risk assessment at real exposure doses.

A simple air-liquid interface exposure system for exposing cultured human 3D epidermis and cornea to PM2.5 collected through cyclonic separation.

Particulate matter (PM) is among the major air pollutants suspended in the atmosphere. PM2.5 has a particle size of 2.5 µm; it is known to cause inflammation, especially in the respiratory tract and skin. Since the skin acts a primary barrier against harmful environmental substances that may enter the body, it is highly exposed to PM2.5 present in the environment. However, the adverse health effects of PM2.5 exposure on human skin have not been accurately examined due to the lack of a system that exposes human epidermal tissue to the actual environmental concentration of PM2.5. In this study, we developed an air-liquid interface exposure system for exposing cultured human 3D epidermis and cornea to PM2.5 collected through cyclonic separation. PM2.5 suspension was nebulized in an acrylic chamber, and the resulting mist was pumped through a diffusion dryer into a glass exposure chamber. A particle counter was connected to the exposure chamber to continuously measure the spatial mass concentration of PM. Human 3D epidermis was cultured in the exposure chamber. Exposure of the human 3D epidermis to PM aerosol increased interleukin-8 release into the media around 50 µg/m3. Mass concentrations above 100 µg/m3 caused cell death. Furthermore, a human corneal model showed similar responses against PM2.5 exposure as 3D epidermis. The air-liquid interface exposure system developed in this study is considered useful for evaluating the health effects induced by environmental PM2.5 and can be used as an alternative to experiments involving actual human or animals.

The Cytotoxic Effects of Fine Particulate Matter (PM2.5) from Different Sources at the Air-Liquid Interface Exposure on A549 Cells.

The health of humans has been negatively impacted by PM2.5 exposure, but the chemical composition and toxicity of PM2.5 might vary depending on its source. To investigate the toxic effects of particulate matter from different sources on lung epithelial cells (A549), PM2.5 samples were collected from residential, industrial, and transportation areas in Nanjing, China. The chemical composition of PM2.5 was analyzed, and toxicological experiments were conducted. The A549 cells were exposed using an air-liquid interface (ALI) exposure system, and the cytotoxic indicators of the cells were detected. The research results indicated that acute exposure to different sources of particulate matter at the air-liquid interface caused damage to the cells, induced the production of ROS, caused apoptosis, inflammatory damage, and DNA damage, with a dose-effect relationship. The content of heavy metals and PAHs in PM2.5 from the traffic source was relatively high, and the toxic effect of the traffic-source samples on the cells was higher than that of the industrial- and residential-source samples. The cytotoxicity of particulate matter was mostly associated with water-soluble ions, carbon components, heavy metals, PAHs, and endotoxin, based on the analysis of the Pearson correlation. Oxidative stress played an important role in PM2.5-induced biological toxicity.

Cytotoxicity and Characterization of Ultrafine Particles from Desktop Three-Dimensional Printers with Multiple Filaments.

Previous research has indicated that ultrafine particles (UFPs, particles less than 100 nm) emitted from desktop three-dimensional (3D) printers exhibit cytotoxicity. However, only a limited number of particles from different filaments and their combinations have been tested for cytotoxicity. This study quantified the emissions of UFPs from a commercially available filament extrusion desktop 3D printer using three different filaments, including acrylonitrile butadiene Styrene (ABS), thermoplastic polyurethane (TPU), and polyethylene terephthalate glycol (PETG). In this study, controlled experiments were conducted where the particles emitted were used to expose cells grown in an air-liquid interface (ALI) system. The ALI exposures were utilized for in vitro characterization of particle mixtures, including UFPs from a 3D printer. Additionally, a lactate dehydrogenase (LDH) assay was used to evaluate the cytotoxic effects of these UFPs. A549 cells were exposed at the ALI to UFPs generated by an operational 3D printer for an average of 45 and 90 min. Twenty-four hours post-exposure, the cells were analyzed for percent cytotoxicity in a 24-well ALI insert (LDH assay). UFP exposure resulted in diminished cell viability, as evidenced by significantly increased LDH levels. The findings demonstrate that ABS has the most significant particle emission. ABS was the only filament that showed a significant difference compared to the high efficiency particulate arrestance (HEPA) following 90 min of exposure (p-value < 0.05). Both ABS and PETG exhibited a significant difference compared to the HEPA control after 45 min of exposure. A preliminary analysis of potential exposure to these products in a typical environment advises caution when operating multiple printer and filament combinations in poorly ventilated spaces or without combined gas and particle filtration systems.

Increased oxidative stress responses in murine macrophages exposed at the air-liquid interface to third- and fourth-generation electronic nicotine delivery system (ENDS) aerosols

BackgroundNew fourth generation electronic nicotine delivery system (ENDS) devices contain high levels of nicotine salt (up to 60 mg/mL), whose cellular and molecular effects on immune cells are currently unknown. Here, we used a physiologically-relevant in vitro air-liquid interface (ALI) exposure model to assess the toxicity of distinct ENDS, a 3rd-generation electronic-cigarette (e-cig) and two 4th-generation ENDS devices (JUUL and Posh Plus). MethodsMurine macrophages (RAW 264.7) were exposed at the ALI to either air, Menthol or Crème Brûlée-flavored ENDS aerosols generated from those devices for 1-hour per day for 1 or 3 consecutive days. Cellular and molecular toxicity was evaluated 24 h post-exposure. Results1-day of Menthol-flavored JUUL aerosol exposure significantly decreased cell viability and significantly increased lactate dehydrogenase (LDH) levels compared to air controls. Further, JUUL Menthol elicited significantly increased reactive oxygen species (ROS) and nitric oxide (NO) production compared to air controls. Posh Crème Brûlée-flavored aerosols displayed significant cytotoxicity – decreased cell viability and increased LDH levels –after 1- and 3-day exposures, while the Crème Brûlée-flavored aerosol produced by the 3rd-generation e-cig device only displayed significant cytotoxicity after 3 days compared to air controls. Further, both Posh and third-generation e-cig Crème Brûlée flavored-aerosols elicited significantly increased ROS plus high levels of 8-isoprostane after 1 and 3 days compared to air controls, indicating increased oxidative stress. Posh and third-generation e-cig Crème Brûlée flavored-aerosols elicited reduction in NO levels after one day, but elicited increase in NO after 3 days. Genes in common dysregulated by both devices after 1 day included α7nAChR, Cyp1a1, Ahr, Mmp12, and iNos. ConclusionOur results suggest that ENDS Menthol and Crème Brûlée-flavored aerosol exposures from both 3rd- and 4th-generation ENDS devices are cytotoxic to macrophages and cause oxidative stress. This can translate into macrophage dysfunction. Although 4th-generation disposable ENDS devices have no adjustable operational settings and are considered low-powered ENDS devices, their aerosols can induce cellular toxicity compared to air-exposed control cells. This study provides scientific evidence for regulation of nicotine salt-based disposable ENDS products.

Downscaling of a Three-Dimensional Alveolar In Vitro Model for High-Throughput Screening of Inhalable Respiratory Sensitizers in the Food Industry

Allergic diseases represent an increasing global health problem, among which food and respiratory allergies play an important part. Inhalation of sensitizers from food sources primes the immune system for an exacerbated response upon subsequent exposure via the ingestion route. With the increasing emergence of novel food additives and proteins and growing morbidity in both food and respiratory allergies, the hazard assessment of the allergenicity of food components is of paramount importance. A three-dimensional (3D) alveolar in vitro model for respiratory sensitization prediction was downscaled from the 6- to the 24-well plate insert format, to meet the industry’s high-throughput (HTS) screening requirements. The resemblance of the cellular architecture of the downscaled model to the alveolar region of the lung was evaluated through microscopy techniques and the functionality of the in vitro model was assessed following air-liquid interface (ALI) exposure to ammonium persulfate (APS). The model represents a promising in vitro tool to screen for the respiratory sensitization potential of food components in a fast and reliable approach.

Toxicity of exhaust emissions from high aromatic and non-aromatic diesel fuels using in vitro ALI exposure system

The differences in the traffic fuels have been shown to affect exhaust emissions and their toxicity. Especially, the aromatic content of diesel fuel is an important factor considering the emissions, notably particulate matter (PM) concentrations. The ultra-fine particles (UFP, particles with a diameter of <100 nm) are important components of engine emissions and connected to various health effects, such as pulmonary and systematic inflammation, and cardiovascular disorders. Studying the toxicity of the UFPs and how different fuel options can be used for mitigating the emissions and toxicity is crucial. In the present study, emissions from a heavy-duty diesel engine were used to assess the exhaust emission toxicity with a thermophoresis-based in vitro air-liquid interface (ALI) exposure system. The aim of the study was to evaluate the toxicity of engine exhaust and the potential effect of 20 % aromatic fossil diesel and 0 % aromatic renewable diesel fuel on emission toxicity. The results of the present study show that the aromatic content of the fuel increases emission toxicity, which was seen as an increase in genotoxicity, distinct inflammatory responses, and alterations in the cell cycle. The increase in genotoxicity was most likely due to the PM phase of the exhaust, as the exposures with high-efficiency particulate absorbing (HEPA)-filtered exhaust resulted in a negligible increase in genotoxicity. However, the solely gaseous exposures still elicited immunological responses. Overall, the present study shows that decreasing the aromatic content of the fuels could be a significant measure in mitigating traffic exhaust toxicity.

21 Toxicity of Aircraft Engine Emissions in Calu-3 Human Cells in Air-Liquid Interface Condition.

Abstract Particulate Matter (PM) emissions from aircraft engines adversely affect air quality around airports, contributing to public health concerns for airport workers and neighbouring communities. In this research, PM was collected from the exhaust of turbine engine after combustion of various types of aircraft fuels, including conventional fuel types Jet A and Jet A1, and more sustainable alternatives with relative low levels of aromatics. PM samples’ toxicity was studied in vitro by air-liquid interface (ALI) exposure of Calu-3 cells in monoculture and co-culture with macrophages (differentiated THP-1 cells). Cells were exposed to PM suspensions (250 µg/mL) using a cloud-based ALI exposure system and analysed 24-hours after exposure. DNA damage was detected through Comet assay in cells exposed to Jet A and Jet A1 samples in absence of cytotoxicity; co-incubation with endonucleases-III and formamidopyrimidine-DNA glycosylase showed that this was not associated with oxidation of DNA bases. Real-Time PCR was performed to investigate the expression profile of genes involved in oxidative stress and DNA repair pathways. Heme Oxygenase 1 gene, Mucin 5AC, and Angiotensin-Converting Enzyme 2 were modulated in the same samples where DNA damage was present. No differences were shown in the pro-inflammatory interleukine-8 secretion among the samples. Most of the PM samples generated using jet fuel with high content of aromatic compounds induced enhanced genotoxicity, suggesting that strategies lowering aromatic content may also result in less toxic PM emissions. Funded by Horizon2020 – Research of Aviation PM Technologies, mOdelling and Regulation (RAPTOR), grant number 863969.

123 Finding Optimal Methods for SbD Hazard Testing of Nanomaterials: The Effect of Cell Model and Exposure Method on Cytokine Response

Abstract The concept of safe-by-design (SbD) can help toxicologists and risk assessors to keep pace with the rapidly expanding field of nanotechnology. As part of SbD, the hazard potential of a new nanomaterial (NM) is identified at the first stages of product development. For this, simple yet predictive toxicity assays are crucial. We investigated the suitability of several in vitro models and exposure methods to predict the human (pulmonary) inflammation potential of NMs. Four silica NMs were selected of which in vivo pulmonary toxicity ranking had previously been established: DQ12 (quartz) &amp;gt; NM-203 (fumed silica) &amp;gt; Colloidal silica &amp;gt; Silanized colloidal silica. Several cell types including a 3D human airway epithelial model, lung epithelial cell lines, and immune cell lines, either in mono- or co-culture were exposed to the four materials either in a submerged setting or at the air-liquid interface (ALI). After 24 hours of exposure, cytokine response was assessed using either 13-plex LegendPlex, ELISA, or qPCR, and toxicity rankings were compared. The results indicate differences in sensitivity between cell models and exposure methods. Submerged exposure of a cell-line seems a suitable first tier for SbD hazard testing in case of NMs that can easily be dispersed. For NMs that are not compatible with submerged testing, ALI exposure might be needed. Higher tier testing might be used to confirm the ranking.

New insights into how popular electronic cigarette aerosols and aerosol constituents affect SARS-CoV-2 infection of human bronchial epithelial cells

The relationship between the use of tobacco products and SARS-CoV-2 infection is poorly understood and controversial. Few studies have examined the effect of electronic cigarettes (ECs) on SARS-CoV-2 infection. We tested the hypothesis that EC fluids and aerosols with nicotine promote SARS-COV-2 infection by increasing viral entry into human respiratory epithelial cells. Responses of BEAS-2B cells to JUUL aerosols or their individual constituents were compared using three exposure platforms: submerged culture, air–liquid-interface (ALI) exposure in a cloud chamber, and ALI exposure in a Cultex system, which produces authentic heated EC aerosols. In general, nicotine and nicotine + propylene glycol/vegetable glycerin aerosols increased ACE2 (angiotensin converting enzyme 2) levels, the SARS-CoV-2 receptor; and increased the activity of TMPRSS2 (transmembrane serine protease 2), an enzyme essential for viral entry. Lentivirus pseudoparticles with spike protein were used to test viral penetration. Exposure to nicotine, EC fluids, or aerosols altered the infection machinery and increased viral entry into cells. While most data were in good agreement across the three exposure platforms, cells were more responsive to treatments when exposed at the ALI in the Cultex system, even though the exposures were brief and intermittent. While both nicotine and JUUL aerosols increased SARS-CoV-2 infection, JUUL significantly decreased the effect of nicotine alone. These data support the idea that vaping can increase the likelihood of contracting COVID-19 and that e-liquid composition may modulate this effect.

Modeling Human Lung Cells Exposure to Wildfire Uncovers Aberrant lncRNAs Signature.

Emissions generated by wildfires are a growing threat to human health and are characterized by a unique chemical composition that is tightly dependent on geographic factors such as fuel type. Long noncoding RNAs (lncRNAs) are a class of RNA molecules proven to be critical to many biological processes, and their condition-specific expression patterns are emerging as prominent prognostic and diagnostic biomarkers for human disease. We utilized a new air-liquid interface (ALI) direct exposure system that we designed and validated in house to expose immortalized human tracheobronchial epithelial cells (AALE) to two unique wildfire smokes representative of geographic regions (Sierra Forest and Great Basin). We conducted an RNAseq analysis on the exposed cell cultures and proved through both principal component and differential expression analysis that each smoke has a unique effect on the LncRNA expression profiles of the exposed cells when compared to the control samples. Our study proves that there is a link between the geographic origin of wildfire smoke and the resulting LncRNA expression profile in exposed lung cells and also serves as a proof of concept for the in-house designed ALI exposure system. Our study serves as an introduction to the scientific community of how unique expression patterns of LncRNAs in patients with wildfire smoke-related disease can be utilized as prognostic and diagnostic tools, as the current roles of LncRNA expression profiles in wildfire smoke-related disease, other than this study, are completely uncharted.

Construction of an In Vitro Air-Liquid Interface Exposure System to Assess the Toxicological Impact of Gas and Particle Phase of Semi-Volatile Organic Compounds.

Anthropogenic activities and industrialization render continuous human exposure to semi-volatile organic compounds (SVOCs) inevitable. Occupational monitoring and safety implementations consider the inhalation exposure of SVOCs as critically relevant. Due to the inherent properties of SVOCs as gas/particle mixtures, risk assessment strategies should consider particle size-segregated SVOC association and the relevance of released gas phase fractions. We constructed an in vitro air-liquid interface (ALI) exposure system to study the distinct toxic effects of the gas and particle phases of the model SVOC dibutyl phthalate (DBP) in A549 human lung epithelial cells. Cytotoxicity was evaluated and genotoxic effects were measured by the alkaline and enzyme versions of the comet assay. Deposited doses were assessed by model calculations and chemical analysis using liquid chromatography tandem mass spectrometry. The novel ALI exposure system was successfully implemented and revealed the distinct genotoxic effects of the gas and particle phases of DBP. The empirical measurements of cellular deposition and the model calculations of the DBP particle phase were concordant.The model SVOC DBP showed that inferred oxidative DNA damage may be attributed to particle-related effects. While pure gas phase exposure may follow a distinct mechanism of genotoxicity, the contribution of the gas phase to total aerosol was comparably low.

Investigation of PM2.5-induced carcinogenic effects through mediation of ErbB family based on DNA methylation and transcriptomics analysis by a lung-mimicking microfluidic platform

Fine particle (PM2.5, less than 2.5 micrometers in diameter) is regarded as a harmful carcinogen. However, the molecular mechanisms of the carcinogenic effects of ambient fine particles have not been fully elucidated, and therapeutic options to address this major public health challenge are lacking. Here, we present global gene-specific DNA methylation and transcriptomic (RNA-Seq) analyses after HBE cells were exposed to fine particles on a portable, small, and all-in-one organ-level lung-mimicking air-liquid interface exposure (MALIE) microfluidic platform. A series of cancer-related signal transduction pathways were activated. ErbB1, ErbB2, and ErbB3 gene expression altered by fine particle exposure was the result of changes in the cellular DNA methylome. The protein expression of ErbB family was inhibited by drugs and could regulate downstream Grb2/Raf pathway and Akt/MDM2 pathway. All of the above results indicated that ErbB family may be promising drug targets for air pollution-related diseases and that inhibitor drugs can be used as therapeutic options to treat these diseases.

A mobile platform for characterizing on-road tailpipe emissions and toxicity of ultrafine particles under real driving Conditions

Acute exposure to fresh traffic-related air pollutants (TRAPs) can be high for road users, including motorbike drivers, cyclists, and pedestrians. However, evaluating the toxicity of fresh traffic emissions from on-road vehicles is challenging since pollution properties can change dynamically within a short distance and time. This study demonstrated a mobile platform equipped with an On-Board Diagnostic II (OBDII) system, a tailor-made portable emission measurement system, and an electrostatic air-liquid interface exposure system with human monocytic THP-1 cells to characterize on-road tailpipe emissions under real driving conditions. High number concentrations up to 106–107 # cm−3 of ultrafine particles (UFPs) were observed for a gasoline engine at the cold-start stage and a diesel engine during particulate filter regeneration. In particular, a substantial fraction of freshly emitted UFPs within the size less than 23 nm were observed and should be cautioned. The potential toxicity of fresh TRAPs was quantified by cell viability, cytotoxicity, oxidative stress, and inflammatory biomarkers. Results show that the decreased cell viability, increased lactate dehydrogenase (LDH) activity, and high oxidative stress induced by the fresh TRAPs were potentially contributed by gaseous pollutants as well as particles, especially driving with the high idling frequency. Moreover, the dominant contributor to the toxicity is different for gasoline's and diesel's TRAPs. Characterizing on-road air pollutant toxicity as well as physicochemical properties using an innovative mobile platform can fill this knowledge gap.

P12-15 Characterisation and biological effects of three types of TiO2 nanoparticles using air-liquid interface exposure

P12-15 Characterisation and biological effects of three types of TiO2 nanoparticles using air-liquid interface exposure