Role of different mechanisms in pro-inflammatory responses triggered by traffic-derived particulate matter in human bronchiolar epithelial cells.

BACKGROUND: Traffic-derived particles are important contributors to the adverse health effects of ambient particulate matter (PM). In Nordic countries, mineral particles from road pavement and diesel exhaust particles (DEP) are important constituents of traffic-derived PM. In the present study we compared the pro-inflammatory responses of mineral particles and DEP to PM from two road tunnels, and examined the mechanisms involved. METHODS: The pro-inflammatory potential of 100 µg/mL coarse (PM10-2.5), fine (PM2.5-0.18) and ultrafine PM (PM0.18) sampled in two road tunnels paved with different stone materials was assessed in human bronchial epithelial cells (HBEC3-KT), and compared to DEP and particles derived from the respective stone materials. Release of pro-inflammatory cytokines (CXCL8, IL-1α, IL-1ß) was measured by ELISA, while the expression of genes related to inflammation (COX2, CXCL8, IL-1α, IL-1ß, TNF-α), redox responses (HO-1) and metabolism (CYP1A1, CYP1B1, PAI-2) was determined by qPCR. The roles of the aryl hydrocarbon receptor (AhR) and reactive oxygen species (ROS) were examined by treatment with the AhR-inhibitor CH223191 and the anti-oxidant N-acetyl cysteine (NAC). RESULTS: Road tunnel PM caused time-dependent increases in expression of CXCL8, COX2, IL-1α, IL-1ß, TNF-α, COX2, PAI-2, CYP1A1, CYP1B1 and HO-1, with fine PM as more potent than coarse PM at early time-points. The stone particle samples and DEP induced lower cytokine release than all size-fractionated PM samples for one tunnel, and versus fine PM for the other tunnel. CH223191 partially reduced release and expression of IL-1α and CXCL8, and expression of COX2, for fine and coarse PM, depending on tunnel, response and time-point. Whereas expression of CYP1A1 was markedly reduced by CH223191, HO-1 expression was not affected. NAC reduced the release and expression of IL-1α and CXCL8, and COX2 expression, but augmented expression of CYP1A1 and HO-1. CONCLUSIONS: The results indicate that the pro-inflammatory responses of road tunnel PM in HBEC3-KT cells are not attributed to the mineral particles or DEP alone. The pro-inflammatory responses seem to involve AhR-dependent mechanisms, suggesting a role for organic constituents. ROS-mediated mechanisms were also involved, probably through AhR-independent pathways. DEP may be a contributor to the AhR-dependent responses, although other sources may be of importance.

Road tunnel-derived coarse, fine and ultrafine particulate matter: physical and chemical characterization and pro-inflammatory responses in human bronchial epithelial cells.

BACKGROUND: Traffic particulate matter (PM) comprises a mixture of particles from fuel combustion and wear of road pavement, tires and brakes. In countries with low winter temperatures the relative contribution of mineral-rich PM from road abrasion may be especially high due to use of studded tires during winter season. The aim of the present study was to sample and characterize size-fractioned PM from two road tunnels paved with different stone materials in the asphalt, and to compare the pro-inflammatory potential of these fractions in human bronchial epithelial cells (HBEC3-KT) in relation to physicochemical characteristics. METHODS: The road tunnel PM was collected with a vacuum pump and a high-volume cascade impactor sampler. PM was sampled during winter, both during humid and dry road surface conditions, and before and after cleaning the tunnels. Samples were analysed for hydrodynamic size distribution, content of elemental carbon (EC), organic carbon (OC) and endotoxin, and the capacity for acellular generation of reactive oxygen species. Cytotoxicity and pro-inflammatory responses were assessed in HBEC3-KT cells after exposure to coarse (2.5-10 µm), fine (0.18-2.5 µm) and ultrafine PM (≤ 0.18 µm), as well as particles from the respective stone materials used in the pavement. RESULTS: The pro-inflammatory potency of the PM samples varied between road tunnels and size fractions, but showed more marked responses than for the stone materials used in asphalt of the respective tunnels. In particular, fine samples showed significant increases as low as 25 µg/mL (2.6 µg/cm2) and were more potent than coarse samples, while ultrafine samples showed more variable responses between tunnels, sampling conditions and endpoints. The most marked responses were observed for fine PM sampled during humid road surface conditions. Linear correlation analysis showed that particle-induced cytokine responses were correlated to OC levels, while no correlations were observed for other PM characteristics. CONCLUSIONS: The pro-inflammatory potential of fine road tunnel PM sampled during winter season was high compared to coarse PM. The differences between the PM-induced cytokine responses were not related to stone materials in the asphalt. However, the ratio of OC to total PM mass was associated with the pro-inflammatory potential.

Potential role of polycyclic aromatic hydrocarbons in air pollution-induced non-malignant respiratory diseases.

Epidemiological studies have found strong associations between air pollution and respiratory effects including development and/or exacerbation of asthma and chronic obstructive pulmonary disease (COPD) as well as increased occurrence of respiratory infections and lung cancer. It has become increasingly clear that also polycyclic aromatic hydrocarbons (PAHs) may affect processes linked to non-malignant diseases in the airways. The aim of the present paper was to review epidemiological studies on associations between gas phase and particle-bound PAHs in ambient air and non-malignant respiratory diseases or closely related physiological processes, to assess whether PAH-exposure may explain some of the effects associated with air pollution. Based on experimental in vivo and in vitro studies, we also explore possible mechanisms for how different PAHs may contribute to such events. Epidemiological studies show strongest evidence for an association between PAHs and asthma development and respiratory function in children. This is supported by studies on prenatal and postnatal exposure. Exposure to PAHs in adults seems to be linked to respiratory functions, exacerbation of asthma and increased morbidity/mortality of obstructive lung diseases. However, available studies are few and weak. Notably, the PAHs measured in plasma/urine also represent other exposure routes than inhalation. Furthermore, the role of PAHs measured in air is difficult to disentangle from that of other air pollution components originating from combustion processes. Experimental studies show that PAHs may trigger various processes linked to non-malignant respiratory diseases. Physiological- and pathological responses include redox imbalance, oxidative stress, inflammation both from the innate and adaptive immune systems, smooth muscle constriction, epithelial- and endothelial dysfunction and dysregulated lung development. Such biological responses may at the molecular level be initiated by PAH-binding to the aryl hydrocarbon receptor (AhR), but possibly also through interactions with beta-adrenergic receptors. In addition, reactive PAH metabolites or reactive oxygen species (ROS) may interfere directly with ion transporters and enzymes involved in signal transduction. Overall, the reviewed literature shows that respiratory effects of PAH-exposure in ambient air may extend beyond lung cancer. The relative importance of the specific PAHs ability to induce disease may differ between the biological endpoint in question.

Effects of particulate matter on atherosclerosis: a link via high-density lipoprotein (HDL) functionality?

BACKGROUND: Exposure to air pollution has been associated with adverse effects on human health, and ultimately increased morbidity and mortality. This is predominantly due to hazardous effects on the cardiovascular system. Exposure to particulate matter (PM) is considered to be responsible for the most severe effects. MAIN BODY: Here we summarize current knowledge from existing epidemiological, clinical and animal studies on the influence of PM exposure on high-density lipoprotein (HDL) functionality and the potential initiation and progression of atherosclerosis. We highlight experimental studies that bring support to the causality and point to possible mechanistic links. Recent studies indicate that the functional properties of HDL are more important than the levels per se. Fine (PM2.5-0.1) and ultrafine (UFP) PM are composed of chemicals as well as biological elements that are redox-active and may trigger pro-inflammatory responses. Experimental studies indicate that these properties and responses may promote HDL dysfunction via oxidative pathways. By affecting protein and lipid components of the HDL particle, its anti-atherosclerotic characteristics including cholesterol efflux capacity, as well as other anti-oxidative and anti-inflammatory features might be impaired. CONCLUSION: Current literature suggests that PM promotes HDL dysfunction via oxidative pathways. However, as relatively few studies so far have evaluated the impact of particulate air pollution on HDL functionality, more human epidemiological as well as experimental studies are needed to strengthen any possible causal relationship and determine any relevance to atherosclerosis.

Characterization and pro-inflammatory potential of indoor mold particles.

A number of epidemiological studies find an association between indoor air dampness and respiratory health effects. This is often suggested to be linked to enhanced mold growth. However, the role of mold is obviously difficult to disentangle from other dampness-related exposure including microbes as well as non-biological particles and chemical pollutants. The association may partly be due to visible mycelial growth and a characteristic musty smell of mold. Thus, the potential role of mold exposure should be further explored by evaluating information from experimental studies elucidating possible mechanistic links. Such studies show that exposure to spores and hyphal fragments may act as allergens and pro-inflammatory mediators and that they may damage airways by the production of toxins, enzymes, and volatile organic compounds. In the present review, we hypothesize that continuous exposure to mold particles may result in chronic low-grade pro-inflammatory responses contributing to respiratory diseases. We summarize some of the main methods for detection and characterization of fungal aerosols and highlight in vitro research elucidating how molds may induce toxicity and pro-inflammatory reactions in human cell models relevant for airway exposure. Data suggest that the fraction of fungal hyphal fragments in indoor air is much higher than that of airborne spores, and the hyphal fragments often have a higher pro-inflammatory potential. Thus, hyphal fragments of prevalent mold species with strong pro-inflammatory potential may be particularly relevant candidates for respiratory diseases associated with damp/mold-contaminated indoor air. Future studies linking of indoor air dampness with health effects should assess the toxicity and pro-inflammatory potential of indoor air particulate matter and combined this information with a better characterization of biological components including hyphal fragments from both pathogenic and non-pathogenic mold species. Such studies may increase our understanding of the potential role of mold exposure.

Pro-inflammatory effects of crystalline- and nano-sized non-crystalline silica particles in a 3D alveolar model.

BACKGROUND: Silica nanoparticles (SiNPs) are among the most widely manufactured and used nanoparticles. Concerns about potential health effects of SiNPs have therefore risen. Using a 3D tri-culture model of the alveolar lung barrier we examined effects of exposure to SiNPs (Si10) and crystalline silica (quartz; Min-U-Sil) in the apical compartment consisting of human alveolar epithelial A549 cells and THP-1-derived macrophages, as well as in the basolateral compartment with Ea.hy926 endothelial cells. Inflammation-related responses were measured by ELISA and gene expression. RESULTS: Exposure to both Si10 and Min-U-Sil induced gene expression and release of CXCL8, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), interleukin-1α (IL-1α) and interleukin-1ß (IL-1ß) in a concentration-dependent manner. Cytokine/chemokine expression and protein levels were highest in the apical compartment. Si10 and Min-U-Sil also induced expression of adhesion molecules ICAM-1 and E-selectin in the apical compartment. In the basolateral endothelial compartment we observed marked, but postponed effects on expression of all these genes, but only at the highest particle concentrations. Geneexpressions of heme oxygenase-1 (HO-1) and the metalloproteases (MMP-1 and MMP-9) were less affected. The IL-1 receptor antagonist (IL-1RA), markedly reduced effects of Si10 and Min-U-Sil exposures on gene expression of cytokines and adhesion molecules, as well as cytokine-release in both compartments. CONCLUSIONS: Si10 and Min-U-Sil induced gene expression and release of pro-inflammatory cytokines/adhesion molecules at both the epithelial/macrophage and endothelial side of a 3D tri-culture. Responses in the basolateral endothelial cells were only induced at high concentrations, and seemed to be mediated by IL-1α/ß released from the apical epithelial cells and macrophages.

Potential role of polycyclic aromatic hydrocarbons as mediators of cardiovascular effects from combustion particles.

Air pollution is the most important environmental risk factor for disease and premature death, and exposure to combustion particles from vehicles is a major contributor. Human epidemiological studies combined with experimental studies strongly suggest that exposure to combustion particles may enhance the risk of cardiovascular disease (CVD), including atherosclerosis, hypertension, thrombosis and myocardial infarction.In this review we hypothesize that adhered organic chemicals like polycyclic aromatic hydrocarbons (PAHs), contribute to development or exacerbation of CVD from combustion particles exposure. We summarize present knowledge from existing human epidemiological and clinical studies as well as experimental studies in animals and relevant in vitro studies. The available evidence suggests that organic compounds attached to these particles are significant triggers of CVD. Furthermore, their effects seem to be mediated at least in part by the aryl hydrocarbon receptor (AhR). The mechanisms include AhR-induced changes in gene expression as well as formation of reactive oxygen species (ROS) and/or reactive electrophilic metabolites. This is in accordance with a role of PAHs, as they seem to be the major chemical group on combustion particles, which bind AhR and/or is metabolically activated by CYP-enzymes. In some experimental models however, it seems as PAHs may induce an inflammatory atherosclerotic plaque phenotype irrespective of DNA- and/or AhR-ligand binding properties. Thus, various components and several signalling mechanisms/pathways are likely involved in CVD induced by combustion particles.We still need to expand our knowledge about the role of PAHs in CVD and in particular the relative importance of the different PAH species. This warrants further studies as enhanced knowledge on this issue may amend risk assessment of CVD caused by combustion particles and selection of efficient measures to reduce the health effects of particular matters (PM).

Pro-inflammatory responses induced by A. fumigatus and A. versicolor in various human macrophage models.

Exposure to mold-contaminated indoor air has been associated with various respiratory diseases, and there is a need for experimental data to confirm these associations. The pro-inflammatory properties of well-characterized aerosolized spores and hyphal fragments from Aspergillus fumigatus and Aspergillus versicolor were examined and compared using various human macrophage cell models including phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 macrophages (THP-1 Ma), primary peripheral blood monocyte-derived macrophages (MDM), and primary airway macrophages (AM) from induced sputum. X-ray treated samples of the two mold species induced different responses with A. fumigatus displaying the most potent induction of pro-inflammatory responses. While hyphal fragments from A. fumigatus were more potent than spores, similar responses were produced by the two growth stages of A. versicolor. THP-1 Ma was the most sensitive model releasing a broad range of cytokines/chemokines. MDM exhibited a similar cytokine/chemokine profile as THP-1 Ma, except for a low-quantity release of interleukin-1ß (IL-1ß). In contrast, AM appeared to be nonresponsive and yielded a different pattern of pro-inflammatory markers. Toll-like receptor (TLR)4, but also to a certain degree TLR2, was involved in several responses induced by spores and aerosolized hyphal fragments of A. fumigatus in MDM. Taken together, MDM seems to be the most promising experimental macrophage model. Abbreviations: AF: A. fumigatus, Aspergillus fumigatus; AV: A. versicolor, Aspergillus versicolor; AM: Airway Macrophage; CBA: Cytometric Bead Array; CD: Cluster of Differentiation; DTT: dithiothreitol; ELISA: Enzyme Linked Immunosorbent Assay; FBS: fetal bovine serum; GM-CSF: Granulocyte macrophage colony-stimulating factor; IL-1ß: Interleukin-1beta; MDM: Monocyte-Derived Macrophages; NF-κB: Nuclear Factor kappa light chain enhancer of activated B cells; NLR: NOD-like Receptor; PAMP: Pathogen Associated Molecular Pattern; PMA: Phorbol 12-myristate 13-acetate; PRR: Pattern Recognition Receptor; THP-1: Human leukemia monocyte cell line; TLR: Toll-like Receptor; TNF-α: Tumor Necrosis Factor- alpha.

Environmental carcinogenesis and pH homeostasis: Not only a matter of dysregulated metabolism.

According to the World Health Organization, around 20% of all cancers would be due to environmental factors. Among these factors, several chemicals are indeed well recognized carcinogens. The widespread contaminant benzo[a]pyrene (B[a]P), an often used model carcinogen of the polycyclic aromatic hydrocarbons' family, has been suggested to target most, if not all, cancer hallmarks described by Hanahan and Weinberg. It is classified as a group I carcinogen by the International Agency for Research on Cancer; however, the precise intracellular mechanisms underlying its carcinogenic properties remain yet to be thoroughly defined. Recently, the pH homeostasis, a well known regulator of carcinogenic processes, was suggested to be a key actor in both cell death and Warburg-like metabolic reprogramming induced upon B[a]P exposure. The present review will highlight those data with the aim of favoring research on the role of H+ dynamics in environmental carcinogenesis.

Early life exposure to air pollution particulate matter (PM) as risk factor for attention deficit/hyperactivity disorder (ADHD): Need for novel strategies for mechanisms and causalities.

Epidemiological studies have demonstrated that air pollution particulate matter (PM) and adsorbed toxicants (organic compounds and trace metals) may affect child development already in utero. Recent studies have also indicated that PM may be a risk factor for neurodevelopmental disorders (NDDs). A pattern of increasing prevalence of attention deficit/hyperactivity disorder (ADHD) has been suggested to partly be linked to environmental pollutants exposure, including PM. Epidemiological studies suggest associations between pre- or postnatal exposure to air pollution components and ADHD symptoms. However, many studies are cross-sectional without possibility to reveal causality. Cohort studies are often small with poor exposure characterization, and confounded by traffic noise and socioeconomic factors, possibly overestimating the study associations. Furthermore, the mechanistic knowledge how exposure to PM during early brain development may contribute to increased risk of ADHD symptoms or cognitive deficits is limited. The closure of this knowledge gap requires the combined use of well-designed longitudinal cohort studies, supported by mechanistic in vitro studies. As ADHD has profound consequences for the children affected and their families, the identification of preventable risk factors such as air pollution exposure should be of high priority.

Lipophilic components of diesel exhaust particles induce pro-inflammatory responses in human endothelial cells through AhR dependent pathway(s).

BACKGROUND: Exposure to traffic-derived particulate matter (PM), such as diesel exhaust particles (DEP), is a leading environmental cause of cardiovascular disease (CVD), and may contribute to endothelial dysfunction and development of atherosclerosis. It is still debated how DEP and other inhaled PM can contribute to CVD. However, organic chemicals (OC) adhered to the particle surface, are considered central to many of the biological effects. In the present study, we have explored the ability of OC from DEP to reach the endothelium and trigger pro-inflammatory reactions, a central step on the path to atherosclerosis. RESULTS: Exposure-relevant concentrations of DEP (0.12 µg/cm2) applied on the epithelial side of an alveolar 3D tri-culture, rapidly induced pro-inflammatory and aryl hydrocarbon receptor (AhR)-regulated genes in the basolateral endothelial cells. These effects seem to be due to soluble lipophilic constituents rather than particle translocation. Extractable organic material of DEP (DEP-EOM) was next fractionated with increasing polarity, chemically characterized, and examined for direct effects on pro-inflammatory and AhR-regulated genes in human microvascular endothelial (HMEC-1) cells and primary human endothelial cells (PHEC) from four healthy donors. Exposure-relevant concentrations of lipophilic DEP-EOM (0.15 µg/cm2) induced low to moderate increases in IL-1α, IL-1ß, COX2 and MMP-1 gene expression, and the MMP-1 secretion was increased. By contrast, the more polar EOM had negligible effects, even at higher concentrations. Use of pharmacological inhibitors indicated that AhR and protease-activated receptor-2 (PAR-2) were central in regulation of EOM-induced gene expression. Some effects also seemed to be attributed to redox-responses, at least at the highest exposure concentrations tested. Although the most lipophilic EOM, that contained the majority of PAHs and aliphatics, had the clearest low-concentration effects, there was no straight-forward link between chemical composition and biological effects. CONCLUSION: Lipophilic and semi-lipophilic chemicals seemed to detach from DEP, translocate through alveolar epithelial cells and trigger pro-inflammatory reactions in endothelial cells at exposure-relevant concentrations. These effects appeared to be triggered by AhR agonists, and involve PAR-2 signaling.

Lipophilic Chemicals from Diesel Exhaust Particles Trigger Calcium Response in Human Endothelial Cells via Aryl Hydrocarbon Receptor Non-Genomic Signalling.

Exposure to diesel exhaust particles (DEPs) affects endothelial function and may contribute to the development of atherosclerosis and vasomotor dysfunction. As intracellular calcium concentration [Ca2+]i is considered important in myoendothelial signalling, we explored the effects of extractable organic matter from DEPs (DEP-EOM) on [Ca2+]i and membrane microstructure in endothelial cells. DEP-EOM of increasing polarity was obtained by pressurized sequential extraction of DEPs with n-hexane (n-Hex-EOM), dichloromethane (DCM-EOM), methanol, and water. Chemical analysis revealed that the majority of organic matter was extracted by the n-Hex- and DCM-EOM, with polycyclic aromatic hydrocarbons primarily occurring in n-Hex-EOM. The concentration of calcium was measured in human microvascular endothelial cells (HMEC-1) using micro-spectrofluorometry. The lipophilic n-Hex-EOM and DCM-EOM, but not the more polar methanol- and water-soluble extracts, induced rapid [Ca2+]i increases in HMEC-1. n-Hex-EOM triggered [Ca2+]i increase from intracellular stores, followed by extracellular calcium influx consistent with store operated calcium entry (SOCE). By contrast, the less lipophilic DCM-EOM triggered [Ca2+]i increase via extracellular influx alone, resembling receptor operated calcium entry (ROCE). Both extracts increased [Ca2+]i via aryl hydrocarbon receptor (AhR) non-genomic signalling, verified by pharmacological inhibition and RNA-interference. Moreover, DCM-EOM appeared to induce an AhR-dependent reduction in the global plasma membrane order, as visualized by confocal fluorescence microscopy. DCM-EOM-triggered [Ca2+]i increase and membrane alterations were attenuated by the membrane stabilizing lipid cholesterol. In conclusion, lipophilic constituents of DEPs extracted by n-hexane and DCM seem to induce rapid AhR-dependent [Ca2+]i increase in HMEC-1 endothelial cells, possibly involving both ROCE and SOCE-mediated mechanisms. The semi-lipophilic fraction extracted by DCM also caused an AhR-dependent reduction in global membrane order, which appeared to be connected to the [Ca2+]i increase.

AhR and Arnt differentially regulate NF-κB signaling and chemokine responses in human bronchial epithelial cells.

BACKGROUND: The aryl hydrocarbon receptor (AhR) has gradually emerged as a regulator of inflammation in the lung and other tissues. AhR may interact with the p65-subunit of the nuclear factor (NF)-κB transcription factors, but reported outcomes of AhR/NF-κB-interactions are conflicting. Some studies suggest that AhR possess pro-inflammatory activities while others suggest that AhR may be anti-inflammatory. The present study explored the impact of AhR and its binding partner AhR nuclear translocator (Arnt) on p65-activation and two differentially regulated chemokines, CXCL8 (IL-8) and CCL5 (RANTES), in human bronchial epithelial cells (BEAS-2B). RESULTS: Cells were exposed to CXCL8- and CCL5-inducing chemicals, 1-nitropyrene (1-NP) and 1-aminopyrene (1-AP) respectively, or the synthetic double-stranded RNA analogue, polyinosinic-polycytidylic acid (Poly I:C) which induced both chemokines. Only CXCL8, and not CCL5, appeared to be p65-dependent. Yet, constitutively active unligated AhR suppressed both CXCL8 and CCL5, as shown by siRNA knock-down and the AhR antagonist α-naphthoflavone. Moreover, AhR suppressed activation of p65 by TNF-α and Poly I:C as assessed by luciferase-assay and p65-phosphorylation at serine 536, without affecting basal p65-activity. In contrast, Arnt suppressed only CXCL8, but did not prevent the p65-activation directly. However, Arnt suppressed expression of the NF-κB-subunit RelB which is under transcriptional regulation by p65. Furthermore, AhR-ligands alone at high concentrations induced a moderate CXCL8-response, without affecting CCL5, but suppressed both CXCL8 and CCL5-responses by Poly I:C. CONCLUSION: AhR and Arnt may differentially and independently regulate chemokine-responses induced by both inhaled pollutants and pulmonary infections. Constitutively active, unligated AhR suppressed the activation of p65, while Arnt may possibly interfere with the action of activated p65. Moreover, ligand-activated AhR suppressed CXCL8 and CCL5 responses by other agents, but AhR ligands alone induced CXCL8 responses when given at sufficiently high concentrations, thus underscoring the duality of AhR in regulation of inflammation. We propose that AhR-signaling may be a weak activator of p65-signaling that suppresses p65-activity induced by strong activators of NF-κB, but that its anti-inflammatory properties also are due to interference with additional pathways.

The occurrence of polycyclic aromatic hydrocarbons and their derivatives and the proinflammatory potential of fractionated extracts of diesel exhaust and wood smoke particles.

Exposure to combustion emissions, including diesel engine exhaust and wood smoke particles (DEPs and WSPs), has been associated with inflammatory responses. To investigate the possible role of polycyclic aromatic hydrocarbons (PAHs) and PAH-derivatives, the DEPs and WSPs methanol extracts were fractionated by solid phase extraction (SPE), and the fractions were analyzed for more than ∼120 compounds. The pro-inflammatory effects of the fractionated extracts were characterized by exposure of bronchial epithelial lung cells (BEAS-2B). Both native DEPs and WSPs caused a concentration-dependent increase in IL-6 and IL-8 release and cytotoxicity. This is consistent with the finding of a rather similar total content of PAHs and PAH-derivatives. Yet, the samples differed in specific components, suggesting that different species contribute to the toxicological response in these two types of particles. The majority of the IL-6 release and cytotoxicity was induced upon exposure to the most polar (methanol) SPE fraction of extracts from both samples. In these fractions hydroxy-PAHs, carboxy-PAHs were observed along with nitro-amino-PAHs in DEP. However, the biological effects induced by the polar fractions could not be attributed only to the occurrence of PAH-derivatives. The present findings indicate a need for further characterization of organic extracts, beyond an extensive analysis of commonly suspected PAH and PAH-derivatives. Supplemental materials are available for this article. Go to the publisher's online edition of Journal of Environmental Science and Health, Part A, to view the supplemental file.

Cell cycle alterations induced by urban PM2.5 in bronchial epithelial cells: characterization of the process and possible mechanisms involved.

BACKGROUND: This study explores and characterizes cell cycle alterations induced by urban PM2.5 in the human epithelial cell line BEAS-2B, and elucidates possible mechanisms involved. METHODS: The cells were exposed to a low dose (7.5 µg/cm(2)) of Milan winter PM2.5 for different time points, and the cell cycle progression was analyzed by fluorescent microscopy and flow cytometry. Activation of proteins involved in cell cycle control was investigated by Western blotting and DNA damage by (32)P-postlabelling, immunostaining and comet assay. The formation of reactive oxygen species (ROS) was quantified by flow cytometry. The role of PM organic fraction versus washed PM on the cell cycle alterations was also examined. Finally, the molecular pathways activated were further examined using specific inhibitors. RESULTS: Winter PM2.5 induced marked cell cycle alteration already after 3 h of exposure, represented by an increased number of cells (transient arrest) in G2. This effect was associated with an increased phosphorylation of Chk2, while no changes in p53 phosphorylation were observed at this time point. The increase in G2 was followed by a transient arrest in the metaphase/anaphase transition point (10 h), which was associated with the presence of severe mitotic spindle aberrations. The metaphase/anaphase delay was apparently followed by mitotic slippage at 24 h, resulting in an increased number of tetraploid G1 cells and cells with micronuclei (MN), and by apoptosis at 40 h. Winter PM2.5 increased the level of ROS at 2 h and DNA damage (8-oxodG, single- and double stand breaks) was detected after 3 h of exposure. The PM organic fraction caused a similar G2/M arrest and augmented ROS formation, while washed PM had no such effects. DNA adducts were detected after 24 h. Both PM-induced DNA damage and G2 arrest were inhibited by the addition of antioxidants and α-naphthoflavone, suggesting the involvement of ROS and reactive electrophilic metabolites formed via a P450-dependent reaction. CONCLUSIONS: Milan winter PM2.5 rapidly induces severe cell cycle alterations, resulting in increased frequency of cells with double nuclei and MN. This effect is related to the metabolic activation of PM2.5 organic chemicals, which cause damages to DNA and spindle apparatus.

Effects of organic chemicals from diesel exhaust particles on adipocytes differentiated from human mesenchymal stem cells.

Exposure to fine particulate matter (PM2.5 ) from incomplete fossil fuel combustion (coal, oil, gas and diesel) has been linked to increased morbidity and mortality due to metabolic diseases. PM2.5 exaggerate adipose inflammation and insulin resistance in mice with diet-induced obesity. Here, we elucidate the hypothesis that such systemic effects may be triggered by adhered particle components affecting adipose tissue directly. Studying adipocytes differentiated from primary human mesenchymal stem cells, we found that lipophilic organic chemicals (OC) from diesel exhaust particles induced inflammation-associated genes and increased secretion of the chemokine CXLC8/interleukin-8 as well as matrix metalloprotease 1. The oxidative stress response gene haem oxygenase-1 and tumour necrosis factor alpha were seemingly not affected, while aryl hydrocarbon receptor-regulated genes, cytochrome P450 1A1 (CYP1A1) and CYP1B1 and plasminogen activator inhibitor-2, were clearly up-regulated. Finally, expression of ß-adrenergic receptor, known to regulate adipocyte homoeostasis, was down-regulated by exposure to these lipophilic OC. Our results indicate that low concentrations of OC from combustion particles have the potential to modify expression of genes in adipocytes that may be linked to metabolic disease. Further studies on mechanisms linking PM exposure and metabolic diseases are warranted.

Lung cancer associated with combustion particles and fine particulate matter (PM2.5) - The roles of polycyclic aromatic hydrocarbons (PAHs) and the aryl hydrocarbon receptor (AhR).

Air pollution is the leading cause of lung cancer after tobacco smoking, contributing to 20% of all lung cancer deaths. Increased risk associated with living near trafficked roads, occupational exposure to diesel exhaust, indoor coal combustion and cigarette smoking, suggest that combustion components in ambient fine particulate matter (PM2.5), such as polycyclic aromatic hydrocarbons (PAHs), may be central drivers of lung cancer. Activation of the aryl hydrocarbon receptor (AhR) induces expression of xenobiotic-metabolizing enzymes (XMEs) and increase PAH metabolism, formation of reactive metabolites, oxidative stress, DNA damage and mutagenesis. Lung cancer tissues from smokers and workers exposed to high combustion PM levels contain mutagenic signatures derived from PAHs. However, recent findings suggest that ambient air PM2.5 exposure primarily induces lung cancer development through tumor promotion of cells harboring naturally acquired oncogenic mutations, thus lacking typical PAH-induced mutations. On this background, we discuss the role of AhR and PAHs in lung cancer development caused by air pollution focusing on the tumor promoting properties including metabolism, immune system, cell proliferation and survival, tumor microenvironment, cell-to-cell communication, tumor growth and metastasis. We suggest that the dichotomy in lung cancer patterns observed between smoking and outdoor air PM2.5 represent the two ends of a dose-response continuum of combustion PM exposure, where tumor promotion in the peripheral lung appears to be the driving factor at the relatively low-dose exposures from ambient air PM2.5, whereas genotoxicity in the central airways becomes increasingly more important at the higher combustion PM levels encountered through smoking and occupational exposure.

Characterization of elements, PAHs, AhR-activity and pro-inflammatory responses of road tunnel-derived particulate matter in human hepatocyte-like and bronchial epithelial cells.

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.

Use of Swedish smokeless tobacco during pregnancy: A systematic review of pregnancy and early life health risk.

BACKGROUND AND AIMS: Smokeless tobacco is a heterogeneous product group with diverse composition and prevalence globally. Tobacco use during pregnancy is concerning due to the risk of adverse pregnancy outcomes and effects on child health. Nicotine may mediate several of these effects. This systematic review measured health outcomes from Swedish smokeless tobacco (snus) use during pregnancy. METHOD: Literature search was conducted by an information specialist in May 2022. We included human studies of snus use during pregnancy compared with no tobacco use, assessed risk of bias, conducted a meta-analysis and assessed confidence in effect-estimates using Grading of Recommendations, Assessment, Development and Evaluations (GRADE). RESULTS: We included 18 cohort studies (42 to 1 006 398 participants). Snus use during pregnancy probably (moderate confidence in risk estimates) increase the risk of neonatal apnea, adjusted odds ratio 95% confidence interval [aOR (95% CI)] 1.96 (1.30 to 2.96). Snus use during pregnancy possibly (low confidence in risk estimates) increase the risk of stillbirths aOR 1.43 (1.02 to 1.99), extremely premature births aOR 1.69 (1.17 to 2.45), moderately premature birth aOR 1.26 (1.15 to 1.38), SGA aOR 1.26 (1.09 to 1.46), reduced birth weight mean difference of 72.47 g (110.58 g to 34.35 g reduction) and oral cleft malformations aOR 1.48 (1.00 to 2.21). It is uncertain (low confidence in risk estimates, CI crossing 1) whether snus use during pregnancy affects risk of preeclampsia aOR 1.11 (0.97 to 1.28), antenatal bleeding aOR 1.15 (0.92 to 1.44) and very premature birth aOR 1.26 (0.95 to 1.66). Risk of early neonatal mortality and altered heart rate variability is uncertain, very low confidence. Snus using mothers had increased prevalence of caesarean sections, low confidence. CONCLUSIONS: This systematic review reveals that use of smokeless tobacco (snus) during pregnancy may adversely impact the developing child.