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3D cell culture models exposed at the air-liquid interface (ALI) represent a potential alternative to animal experiments for hazard and risk assessment of inhaled compounds. This study compares cocultures composed of either Calu-3, A549 or HBEC3-KT lung epithelial cells, cultured together with THP-1-derived macrophages and EA.hy926 endothelial cells, in terms of barrier capacity and responses to a standard reference sample of fine particulate matter (SRM 2786). High-content imaging analysis revealed a similar cellular composition between the different cell models. The 3D cell cultures with Calu-3 cells showed the greatest barrier capacity, as measured by transepithelial electrical resistance and permeability to Na-fluorescein. Mucus production was detected in 3D cell cultures based on Calu-3 and A549 cells. Exposure to SRM 2786 at ALI increased cytokine release and expression of genes associated with inflammation and xenobiotic metabolism. Moreover, the presence of THP-1-derived macrophages was central to the cytokine responses in all cell models. While the different 3D cell culture models produced qualitatively similar responses, more pronounced pro-inflammatory responses were observed in the basolateral compartment of the A549 and HBEC3-KT models compared to the Calu-3 model, likely due to their reduced barrier capacity and lower retention of secreted mediators in the apical compartment., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest. The following are the supplementary data related to this article. Supplementary data to this article can be found online at https://doi.org/10.1016/j.tiv.2024.105841., (Copyright © 2024. Published by Elsevier Ltd.)

The aims were to characterize the content of elements and polycyclic aromatic hydrocarbons (PAHs) in size-separated particulate matter (PM) sampled in a road tunnel, estimate the contribution of PAHs to the toxic potential, and measure the pro-inflammatory potential of PM samples and extracts with increasing polarity. Several elements/metals previously associated with cytokine responses were found. Based on PAHs levels and published PAHs potency, the calculated mutagenic and carcinogenic activities of size-separated samples were somewhat lower for coarse than fine and ultrafine PM. The AhR-activity of the corresponding PM extracts measured in an AhR-luciferase reporter model (human hepatocytes) were more similar. The highest AhR-activity was found in the neutral (parent and alkylated PAHs) and polar (oxy-PAHs) fractions, while the semi-polar fractions (mono-nitrated-PAHs) had only weak activity. The neutral and polar aromatic fractions from coarse and fine PM were also found to induce higher pro-inflammatory responses and CYP1A1 expression in human bronchial epithelial cells (HBEC3-KT) than the semi-polar fractions. Fine PM induced higher pro-inflammatory responses than coarse PM. AhR-inhibition reduced cytokine responses induced by parent PM and extracts of both size fractions. Contributors to the toxic potentials include PAHs and oxy-PAHs, but substantial contributions from other organic compounds and/or metals are likely., Competing Interests: Declaration of Competing Interest The authors declare that they have no competing interests. The authors alone are responsible for the content and writing of the paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The toxicity of some per- and polyfluoroalkyl substances (PFASs), such as perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA) has been studied thoroughly, showing that systemic PFASs targets the lungs. However, regulators lack data to assess the impact of other PFASs on the lungs and alternative methods to test substances for lung toxicity are needed. We combined two in vitro models to assess toxicity to the respiratory system; i) a lung surfactant (LS) function assay to assess the acute inhalation toxicity potential, and ii) a cell model with human bronchial epithelial cells to study pro-inflammatory potential and modulation of inflammatory responses. We tested salts of four PFASs: perfluorobutane sulfonate (PFBS), perfluorohexane sulfonate (PFHxS), PFOS, and PFOA as well as the fluorotelomer 8:2 FTOH. The results show that PFHxS, PFOA and PFOS can inhibit LS function. High PFOS concentrations induced a pro-inflammatory response, measured as increased IL-1α/β release. Moderate concentrations of PFOS suppressed release of the chemokines CXCL8 and CXCL10, whereas both PFOS and PFOA stimulated the release of the pro-inflammatory cytokine IL-1β in immune stimulated human bronchial epithelial cells. These findings support the concern that some PFASs may increase the risk of acute lung toxicity and of airway infections., (Copyright © 2019 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Different transition metals have been shown to induce inflammatory responses in lung. We have compared eight different metal ions with regard to cytokine responses, cytotoxicity and signalling mechanisms in a human lung epithelial cell model (BEAS-2B). Among the metal ions tested, there were large differences with respect to pro-inflammatory potential. Exposure to Cd(2+), Zn(2+) and As(3+) induced CXCL8 and IL-6 release at concentrations below 100μM, and Mn(2+) and Ni(2+) at concentrations above 200μM. In contrast, VO4(3-), Cu(2+) and Fe(2+) did not induce any significant increase of these cytokines. An expression array of 20 inflammatory relevant genes also showed a marked up-regulation of CXCL10, IL-10, IL-13 and CSF2 by one or more of the metal ions. The most potent metals, Cd(2+), Zn(2+) and As(3+) induced highest levels of oxidative activity, and ROS appeared to be central in their CXCL8 and IL-6 responses. Activation of the MAPK p38 seemed to be a critical mediator. However, the NF-κB pathway appeared predominately to be involved only in Zn(2+)- and As(3+)-induced CXCL8 and IL-6 responses. Thus, the most potent metals Cd(2+), Zn(2+) and As(3+) seemed to induce a similar pattern for the cytokine responses, and with some exceptions, via similar signalling mechanisms., (Copyright © 2016 Elsevier Ltd. All rights reserved.)

The present study compared non-crystalline silica particles of nano (50nm)- and submicro (500nm)-size (Si50 and Si500) for the potential to induce cytokine responses in bronchial epithelial lung cells (BEAS-2B). The cell cultures were exposed to equal mass and surface area concentrations of the two particles in different exposure media; LHC-9 and DMEM:F12. The state of agglomeration was different in the two media; with marked agglomeration in LHC-9 and nearly no agglomeration in DMEM:F12. On a mass basis, Si50 was more potent than Si500 in inducing cytokine responses in both exposure media. In contrast, upon exposure to similar surface area concentrations, Si500 was more potent than Si50 in DMEM:F12. This might be due to different agglomeration/sedimentation properties of Si50 versus Si500 in the two media. However, influence of differences in particle reactivity or particle uptake cannot be excluded. The data indicated no qualitative changes in the cytokine gene-expression patterns induced by the two particles, suggesting effects through similar mechanisms. These aspects might be of importance for interpretation of in vitro studies of nanomaterials., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The aims of the present study were to establish to what extent IL-1, and intracellular pathways involving mitogen-activated protein kinases (MAPKs) and nuclear factor-kappa B (NF-kappaB), play a role in ultrafine particle-induced release of IL-6 by primary rat epithelial lung cells. Ultrafine carbon black (Printex 90) induced a concentration- and time-dependent increase in the release of IL-1alpha, IL-1beta and IL-6. The ultrafine carbon black-induced release of IL-6 was completely eliminated by an IL-1 receptor antagonist (IL-1ra). Cellular release of IL-1alpha, IL-1beta and IL-6 was significantly attenuated by curcumin and by inhibitors of the MAPKs ERK1/2 (PD98069), p38 (SB202190) and JNK (SP600125), whereas pyrrolidine dithiocarbamate (PDTC) attenuated the release of IL-6, but not of IL-1alpha and IL-1beta. The effects of curcumin and PDTC may indicate an involvement of NF-kappaB. Furthermore, ultrafine carbon black induced degradation of IkappaBalpha, used as an indicator of NF-kappaB activation, and induced phosphorylation of ERK1/2, p38 and JNK1/2. This degradation and phosphorylation was attenuated by IL-1ra. The present findings provide more insight into the largely unknown mechanisms involved in ultrafine particle-induced release of cytokines from lung cells. The findings suggest that ultrafine carbon black-induced release of IL-6 strongly depends on IL-1 and that activation of MAPKs and NF-kappaB is involved in this response.

The pro-inflammatory cytokines IL-1 beta, TNF-alpha and IL-6 are of great importance in the development of silica-induced lung damage and repair. In this study we investigated the role of IL-1 beta, TNF-alpha and COX2 in silica-induced regulation of IL-6 release and pneumocyte loss in various mono- and co-cultures of monocytes, pneumocytes and endothelial cells. All co-cultures with monocytes, and especially cultures including endothelial cells, showed an increase of silica-induced release of IL-6 compared to the respective monocultures. Treatment with the antagonist IL-1 ra strongly decreased IL-1 beta and IL-6 release in contact co-cultures of monocytes and pneumocytes. COX2 up-regulation by silica and IL-1 beta was eliminated by IL-1 ra. Inhibition of COX2 markedly reduced both IL-1 beta and IL-6 release. IL-1 ra was more effective than COX2-inhibition in reduction of IL-6, but not of IL-1 beta. Silica-induced pneumocyte loss was reduced by IL-1 beta, but this effect was not counteracted by the IL-1 receptor antagonist. Our findings suggest that silica-induced IL-6 release from pneumocytes is mainly mediated via IL-1 beta release from the monocytes, via both COX2-dependent and -independent pathways. Notably, COX2-derived mediators seem crucial for a positive feed-back regulation of IL-1 beta release from the monocytes. In contrast to silica-induced IL-6, the reduction in pneumocyte loss by IL-1 beta does not seem to be regulated through an IL-1R1-dependent mechanism.

Several studies have shown that particles of smaller size may be more potent than larger to induce inflammatory and toxic responses in cultured lung cells. However, the relative importance of different size fractions of ambient PM to induce such effects is still not known. In this study, we investigated the potency of different size fractions of urban ambient air particles to induce release of inflammatory cytokines in the human alveolar cell line A549 and primary rat type 2 cells. A mineral-rich ambient air PM10 sample collected in a road tunnel (road PM10) was also included. The coarse fraction of the urban ambient air particles demonstrated a similar or higher potency to induce release of the proinflammatory cytokines IL-8/MIP-2 and IL-6 compared to the fine and ultrafine fractions. The coarse fraction was also the most toxic in both cell systems. In contrast to the A549 cells, no induction of cytokine release was induced by the ultrafine particles in the primary type 2 cells. The mineral-rich road PM10 may be equally or more potent than the various size fractions of the ambient air particles to induce cytokines in both cell types. In conclusion, the coarse fraction of ambient particles may be at least as potent by mass as smaller fractions to induce inflammatory and toxic effects in lung cells.