Copper-dependent biological effects of particulate matter produced by brake systems on lung alveolar cells.

Road traffic is one of the main sources of particulate emissions into the environment and has an increasing, negative impact on the release of potentially dangerous materials. Vehicle brakes release a significant amount of wear particles, and knowledge regarding their possible adverse effects is limited. One of the most dangerous elements contained in brake pads is copper (Cu), known to be toxic for human health. Therefore, our aim was to study the cell toxicity of particulate matter (PM) produced by different combinations of braking discs and pads containing different amounts of Cu. We investigated whether brake-derived microparticles have toxic effects on lung cells proportionally to their Cu content. Analyte content was measured in friction materials by XRFS and in PM2.5 captured during braking tests using SEM/EDX. The biological impact of brake-derived PM2.5 was investigated on a human epithelial alveolar cell line (A549). Cell viability, oxidative stress, mitochondrial membrane potential, apoptosis, and the pro-inflammatory response of the cells, as well as gene expression, were assessed following exposure to increasing PM2.5 concentrations (1, 10, 100, 200, and 500 µg/ml). The brake debris with the lowest Cu content did not induce significant changes in biological effects on A549 cells compared to normal controls, except for ROS production and IL6 gene expression. PM2.5 containing higher Cu quantities induced cell toxicity that correlated with Cu concentration. Our data suggest that the toxicity of PM2.5 from the brake system is mainly related to Cu content, thus confirming that eliminating Cu from brake pads will be beneficial for human health in urbanized environments.

Ecotoxicological effects of atmospheric particulate produced by braking systems on aquatic and edaphic organisms.

Vehicles generate particulate matter (PM) in significant amounts as their brake systems wear. These particles can influence air quality and their transport/deposition may affect the edaphic and aquatic ecosystems. As part of the LOWBRASYS H2020 project, new more eco-friendly brake disc and pad formulations were developed. PMs generated from traditional (FM1-BD1) and innovative (FM4-BD2, FMB-BD7) brake systems in bench tests were studied. The PMs' physical/chemical characteristics were preliminarily investigated. To study the possible environmental impact of the nano-micro particulate, we used a battery of ecotoxicological tests. We employed the microalga Pseudokirchneriella subcapitata, the crustacean Daphnia magna and the bacteria Vibrio fischeri as aquatic bioindicators, while for the edaphic ecosystem we used the seeds of Lepidium sativum and Sorghum saccharatum, the nematode Caenorhabditis elegans, the earthworm Eisenia andrei and the ameba Dictyostelium discoideum. The results showed a higher sensitivity of the freshwater organisms exposed to the soluble PM fraction, with respect to the edaphic ones. FM4-BD2 brake formulation was slightly more toxic for algae (200 mg/L) than FM1-BD1 (500 mg/L). The new system FMB-BD7 particulate was not harmful for crustacean survival, and resulted weakly toxic for algal reproduction only at 500 mg/L. The particulate material per se was found to affect the algal reproduction. No toxic effects were found on nematodes, earthworms and seeds up to 1000 mg/L. However, in D. discoideum the reproduction rate was significantly reduced starting from 100 mg/L; and the lysosomal membrane stability showed a relevant alteration also at minimal concentration (0.1 mg/L). The results demonstrated a minimal risk for biodiversity of the particulates from the different brake systems and highlighted a more eco-friendly performance the new brake-pad FMB-BD7. However, the occurrence of sublethal effects should be considered as a possible contribution of the particle toxicity to the biological effects of the environmental pollution.

Phytotoxicity of wear debris from traditional and innovative brake pads.

Traffic-related emissions include gas and particles that can alter air quality and affect human and environmental health. Limited studies have demonstrated that particulate debris thrown off from brakes are toxic to higher plants. The acute phytotoxicity of brake pad wear debris (BPWD) investigated using cress seeds grown in soil contaminated with increasing concentrations of debris. Two types of pads were used: a commercially available phenol based pad and an innovative cement-based pad developed within of the LIFE+ COBRA project. The results suggested that even through the BPWD generated by the two pads were similar in and morphology, debris from traditional pads were more phytotoxic than that from cementitious pads, causing significant alterations in terms of root elongation and loss of plasma membrane integrity.

Air quality in the Olona Valley and in vitro human health effects.

Air quality is a major point in current health policies in force globally to protect human health and ecosystems. Cardiovascular and lung diseases are the pathologies most commonly associated with air pollution and it has been estimated that exposure to particulate matters and ground-level ozone and nitric oxides caused >500.000 premature deaths in Europe. Although air quality was generally improved in the recent years, further efforts are required to reduce the impact of air pollution on humans. The present study applied a multidisciplinary approach to estimate the adverse effects on the health of the inhabitants of the Olona Valley in the north of Italy. Chemical analyses quantified the air levels of metals, dioxins, PCBs, PAHs and some macropollutants, including total, fine and coarse airborne particles. These results were used as input for the health risk assessment and in vitro bioassays were used to evaluate possible adverse effects on the respiratory tract due to the organic pollutants adsorbed on the airborne particulate matter. Critical alerts were identified from the air characterization and from the chemical-based risk assessment in view of the levels of arsenic, nickel, benzene, fine and coarse particulate matters found in the investigated zone, which can induce severe adverse effects on human health. These findings were confirmed by bioassays with A549 and BEAS-2B cells. We also used the cell transformation assay with BALB/c 3T3 cells to assess the carcinogenicity of the organic extracts of collected particles as an innovative tool to establish the possible chronic effects of inhaled pollutants. No significant changes in morphological transformation were found suggesting that, although the extracts contain compounds with proven carcinogenic potential, in our experimental conditions the levels of these pollutants were too low to induce carcinogenesis as resulted also by the chemical-based risk assessment.

Soil quality in the Lomellina area using in vitro models and ecotoxicological assays.

Soil quality is traditionally evaluated by chemical characterization to determine levels of pollutants. Biological tools are now employed for soil monitoring since they can take account of the global biological effects induced by all xenobiotics. A combined monitoring of soils based on chemical analyses, human-related in vitro models and ecotoxicological assay was applied in the Lomellina, a semirural area of northern Italy. Chemical characterization indicated overall good quality of the soils, with low levels of toxic and carcinogenic pollutants such as heavy metals, PAHs, PCDD/Fs and PCBs. HepG2 cells were used as a model for the human liver and BALB/c 3T3 cells to evaluate carcinogenic potential. Cells were treated with soil extractable organic matter (EOM) and the MTS assay, DNA release and morphological transformation were selected as endpoints for toxicity and carcinogenicity. Soil EOMs induced dose-dependent inhibition of cell growth at low doses and cytotoxicity only at doses of 500 and 1000 mg soil equivalents/ml. Potential issues for human health can be hypothesized after ingestion of soil samples from some sites. No statistically significant inductions of foci were recorded after exposure to EOMs, indicating that the levels of the soil-extracted organic pollutants were too low to induce carcinogenesis in our experimental conditions. An acute phytotoxicity test and studies on Caenorhabditis elegans were used as ecotoxicological assays for plants and small invertebrates. No significant alerts for ecotoxicity were found. In this proposed case study, HepG2 cells detected differences in the toxicity of soil EOMs, indicating that this cell line could be appropriate to assess the potential harm caused by the ingestion of contaminated soil. Additional information on the carcinogenic potential of mixtures was provided by the cell transformation assay, strengthening the combined approach.

Chemical-based risk assessment and in vitro models of human health effects induced by organic pollutants in soils from the Olona Valley.

Risk assessment of soils is usually based on chemical measurements and assuming accidental soil ingestion and evaluating induced toxic and carcinogenic effects. Recently biological tools have been coupled to chemical-based risk assessment since they integrate the biological effects of all xenobiotics in soils. We employed integrated monitoring of soils based on chemical analyses, risk assessment and in vitro models in the highly urbanized semirural area of the Olona Valley in northern Italy. Chemical characterization of the soils indicated low levels of toxic and carcinogenic pollutants such as PAHs, PCDD/Fs, PCBs and HCB and human risk assessment did not give any significant alerts. HepG2 and BALB/c 3T3 cells were used as a model for the human liver and as a tool for the evaluation of carcinogenic potential. Cells were treated with soil extractable organic matters (EOMs) and the MTS assay, LDH release and morphological transformation were selected as endpoints for toxicity and carcinogenicity. Soil EOMs induced dose-dependent inhibition of cell growth at low doses and cytotoxicity after exposure to higher doses. This might be the result of block of cell cycle progression to repair DNA damage caused by oxidative stress; if this DNA damage cannot be repaired, cells die. No significant inductions of foci were recorded after exposure to EOMs. These results indicate that, although the extracts contain compounds with proven carcinogenic potential, the levels of these pollutants in the analyzed soils were too low to induce carcinogenesis in our experimental conditions. In this proposed case study, HepG2 cells were found an appropriate tool to assess the potential harm caused by the ingestion of contaminated soil as they were able to detect differences in the toxicity of soil EOMs. Moreover, the cell transformation assay strengthened the combined approach giving useful information on carcinogenic potential of mixtures.