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Application potential of screening in vitro toxicological assays in qualitative risk assessment of nanomaterials
Závodná, Táňa ; Topinka, Jan ; Danihelka, J.
Undeniable benefits of engineered nanomaterials might be discredited by their potential enhanced or unexpected toxicity arising from nano-specific properties and behavior. An analysis of the applicability of the traditional chemical risk assessment approach in nanomaterials revealed high levels of uncertainty in both hazard characterization and exposure assessment due to the lack of relevant validated methods and reliable data. This indicates the limited capability of the conventional risk assessment approach to ensure the safe use of nanomaterials. Based on the identified uncertainties, the control banding approach was proposed as a suitable tool for preliminary qualitative risk assessment of nanomaterials in occupational settings. Control banding categorizes hazard and exposure into levels referred to as bands. The combination of the hazard and exposure bands results in a risk band determining the necessary degree of control and regulatory measures. To decrease the number of cases where, based on the precautionary principle, unavailable experimental or field data would lead to the assignment to the highest hazard category requiring costly exposure control, screening evaluation of nanomaterial toxicity was proposed as an additional decision criterion. For this purpose, a battery of in vitro toxicological assays enabling screening evaluation of potential toxic effects of NMs was proposed. The assays evaluate endpoints covering basic toxic effects of substances (cytotoxicity, genotoxicity), as well as known nonspecific mechanisms of toxicity typical for nanomaterials (oxidative stress, inflammation). The proposed risk management strategy is intended to assist small and medium-sized enterprises to implement adequate measures to ensure employee safety.
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The role of dispersion medium on nanoparticle aggregation and size in biological systems
Červená, Tereza ; Rössnerová, Andrea ; Závodná, Táňa ; Vrbová, Kristýna ; Sikorová, Jitka ; Topinka, Jan ; Rössner ml., Pavel
The use of nanomaterials (NMs) in different areas has been rising for more than a decade. Along with this growth, there is visible development of different testing tools and approaches for measuring the actual size of nanomaterials in biological systems. Test conditions during in vitro toxicological assays are different from the standard conditions under which nanomaterials are characterized and careful evaluation of results is needed. The unique properties and range variety of NMs require the close look how the NMs behave in different dispersion medium over time. In this study we present the results of five types of well-characterized NMs (TiO2: NM-101 and NM-103, SiO2: NM-200, Ag: NM-300K and NM-302) of specific size and shape. The hydrodynamic size and Zeta potentials in suspensions were measured using a dynamic light scattering technique (DLS) (Zetasizer Nano ZS, Malvern, UK). The DLS method is suitable for spherical particles, nevertheless, all samples were measured in order to obtain a rough insight into agglomerate formation in the medium. NM300, NM302, and NM200 aggregated rapidly in the media, thus the cells would be most likely exposed to settled big aggregates then small clusters or individual particles. More stable NMs (NM100 and NM103) showed slight grow along with cultivation time or concentration corresponding to cluster formation. Cells exposed to those NMs would be in contact with small clusters and aggregates of NMs. Measured zeta potentials fluctuated around the stability limit corresponding to observed aggregation.
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Ultrafine particles and their possible role in etiology and development of neurodegenerative diseases
Topinka, Jan ; Závodná, Táňa ; Rössnerová, Andrea ; Rössner ml., Pavel
Air pollutants have been shown to cause a vast amount of different adverse health effects. These effects include impairment of many respiratory (e.g. asthma, chronic obstructive pulmonary disease) and cardiovascular (ischemic heart disease, infarction, stroke) diseases. However, in recent years, the evidence showing effects beyond the lungs and circulatory system are becoming more evident. Neurological diseases, namely Alzheimer's disease (AD) has shown to be associated with living near traffic. However, reason for this has remained unresolved until today. Our new H2020 project TUBE aims on revealing the mechanisms of action of ultrafine particles involved in neurological diseases. The TUBE consortium includes experts in areas of aerosol technology, emission research, engine and fuel research, human clinical studies, epidemiology, emission inventories, inhalation toxicology, neurotoxicology and disease mechanism studies. This enables research of resolving the effects of nanoparticles from different traffic modes for both air quality and concomitant toxic effect of these air pollutants. We will investigate adverse effects of air pollutants using cell cultures, animal exposures and volunteered human exposures as well as the material from epidemiological cohort study. These are going to be compared according to inflammatory, cytotoxic and genotoxic changes and furthermore beyond the current state of the art to neurotoxic and brain health effects. With this approach, we are aiming to a comprehensive understanding of the adverse brain effects of nanoparticles from traffic.
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Toxic responses in human lung epithelial cells (BEAS-2B) exposed to particulate matter exhaust emissions from gasoline and biogasoline
Závodná, Táňa ; Líbalová, Helena ; Vrbová, Kristýna ; Sikorová, Jitka ; Vojtíšek-Lom, M. ; Beránek, V. ; Pechout, M. ; Kléma, J. ; Cigánek, M. ; Machala, M. ; Neča, J. ; Rössner ml., Pavel ; Topinka, Jan
Motor vehicle emissions substantially contribute to air pollution worldwide and cause serious health problems. While the deleterious effects of diesel exhaust particulate matter (PM) have been widely studied, much less attention is paid to toxicity of PM emitted by gasoline engines although they also produce considerable amount of PM. The primary objective of this research was to assess toxic potencies of exhaust PM released by conventional gasoline engine fueled with neat gasoline (EU) or gasoline-ethanol blend (15% ethanol, v/v, E15). Despite a similar particle mass (mu g PM/kg fuel) produced by both fuels, PM emitted by E15 contained higher amount of harmful polycyclic aromatic hydrocarbons (PAH) as suggested by chemical analysis. To examine the toxicity of organic PM constituents, human lung BEAS-2B cells were exposed for 4h and 24h to a subtoxic dose of E0 and E15 PM organic extracts. We used genome scale transcriptomic analysis to characterize the toxic response and to identify modulated biological process and pathways. Whereas 4h exposure to both PM extracts resulted in modulation of similar genes and pathways related to lipid and steroid metabolism, activation of PPAR alpha, oxidative stress and immune response, 24h exposure was more specific for each extract, although both induced expression of PAH-metabolic enzymes, modulated metabolism of lipids or activated PPAR alpha, E15 additionally deregulated variety of other pathways. Overall, the PM mass produced by both fuels was similar, however, higher PAH content in E15 PM organic extract may have contributed to more extensive toxic response particularly after 24h exposure in BEAS-2B cells.
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Males-females differences in the spectrum of chromosomal aberrations in the group of nanocomposites production workers
Rössnerová, Andrea ; Pelcová, D. ; Ždímal, Vladimír ; Elzeinova, Fatima ; Margaryan, Hasmik ; Chvojková, Irena ; Topinka, Jan ; Schwarz, Jaroslav ; Ondráček, Jakub ; Koštejn, Martin ; Komarc, M. ; Vlčková, Š. ; Fenclová, Z. ; Lischková, L. ; Dvořáčková, Š. ; Rössner ml., Pavel
An increase in the use of nanomaterials (NM) has been witnessed in many areas of human life. Therefore, assessment of genotoxicity of NM and nanoparticles (NP) is one of the main objectives of genetic toxicology. Despite this fact, human cytogenetic studies following the exposure to NP are still rare. Moreover, no relevant information on possible differences in sensitivity to NP related to gender is available.\n\nIn this study we periodically (in September 2016, 2017 and 2018; pre-shift and post-shift each year) analyzed a group of workers (both genders), working long time in nanocomposites research, and matched controls. Aerosol exposure monitoring of particulate matter including nano-sized fractions was carried out during working shift. Micronucleus assay using Human Pan Centromeric probes, was applied to distinguish, besides the frequency of total MN in binucleated cells (BNC), also other types of chromosomal damage (losses and breaks). Moreover, whole-chromosome painting (WCP) for autosome #1 and both gonosomes (X and Y) were applied in third sampling period (2018) with the aim to identify the particular structural and numerical chromosomal aberrations.\n\nObtained results showed: (i) differences in the risk of exposure to NP related to individual working processes (welding, smelting and machining); (ii) differences in chemical composition of nano-fraction; (iii) no effect of chronic exposure of NP (total MN) opposite to significant effect of acute exposure; (iv) gender-related DNA damage differences (females seem to be more sensitive to chromosomal losses). Additional data from WCP suggested increased frequency of numerical aberrations in gonosomes.
Plný tet:  PDF
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Measurement and toxicity of combustion generated nanoparticles
Sikorová, Jitka ; Topinka, Jan (advisor) ; Klusoň, Petr (referee) ; Bláha, Luděk (referee)
This thesis is focused on nanoparticles produced by internal combustion engines utilized in vehicles. It deals with spatial distribution of nanoparticles within urban areas, impact of alternative fuels usage on particle production and toxicity, and a particle toxicological testing methodology. Monitoring of airborne nanoparticles identified traffic as the main source of airborne nanoparticles in places with heavy traffic load (Prague), as well as in a small city with only local traffic (Čelákovice). Most particles were likely emitted during short episodes of high emissions (e.g. uphill acceleration). During the measurements, high-emission vehicles responsible for a large fraction of the air pollution were also identified. On the other hand, small non-road internal combustion engines, which are not subject to any limit on particle emissions, such as a lawn mower, were operated during the measurement and generated a large number of nanoparticles. The amount and characteristics of the particles produced by combustion depend on the combustion technology and the fuel composition. A large part of the thesis deals with alternative fuels and their effects on the quantity of produced particles and toxicity of organic matter adsorbed on the particles. Hydrotreated vegetable oil (HVO) exhibited the lowest...
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Use of transcriptomics to study mechanism of the action of complex mixtures of organic compounds occurring in the ambient air focusing on polycyclic aromatic hydrocarbons
Líbalová, Helena ; Topinka, Jan (advisor) ; Krásný, Libor (referee) ; Postlerová, Pavla (referee)
Polycyclic aromatic hydrocarbons (PAH) represent a large group of organic compounds occuring as pollutants in ambient air. Besides their genotoxic effect, some of them are known to be complete carcinogens and act via nongenotoxic and tumor promoting mechanism. Although effects of many individual compounds are well-documented, human exposure to polycyclic aromatic hydrocarbons in ambient air occurs through complex mixtures and only few studies describe the behavior of PAH in real complex mixtures. The first part of the thesis is dealing with the global gene expression changes in human embryonic lung fibroblasts (HEL) as a consequence of the effect of complex mixtures containing PAH extracted from the respirable airborne particles PM2.5. These particles were collected in 4 localities in the Czech republic (Ostrava - Bartovice, Ostrava - Poruba, Karviná, Třeboň) differing in the level of the air pollution. Gene expression changes induced by three subtoxic concentrations of organic extracts (EOM - extractable organic matter) from each locality after 24 hour incubation were examined by microarray analysis. Pathway analysis using the Kyoto Encyclopedia of Genes and Genomes (KEGG) database was applied to interpret gene expression data. In each locality we identified several deregulated signaling pathways...
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Interactions of cells with nanoparticles for bio-medical applications
Bělinová, Tereza ; Hubálek Kalbáčová, Marie (advisor) ; Benson, Veronika (referee) ; Topinka, Jan (referee)
In the past decades, nanoparticles have been viewed as a potentially powerful platform for various applications in biomedical sciences. The possible application of nanoparticles varies from drug delivery agents to novel imaging platforms and surely, some application potential still remains hidden. Thus, it is necessary to broadly study their in vitro behavior in order to assess the precise theranostic potential as well as to distinguish possible threats to human health. Even though nanoparticles are getting more and more attention in current research, still only a limited amount of information is available, especially regarding interactions of ultra-small (< 5 nm) nanoparticles with biological environment and cells. The aim of the work presented herein is to provide the reader with information concerning interactions of various ultra-small nanoparticles (silicon-based, gold, nanodiamonds) with biological environment and human cells. Dose- and time-dependent influence of the various nanoparticles on behavior of different human cells (osteoblasts, monocytes, keratinocytes, mesenchymal stem cells) was established under different conditions, stressing out the importance of protein corona (a layer of proteins originating from cultivation medium attached to nanoparticles). Biocompatibility of two...
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Interactions of cells with nanoparticles for bio-medical applications
Bělinová, Tereza ; Hubálek Kalbáčová, Marie (advisor) ; Benson, Veronika (referee) ; Topinka, Jan (referee)
In the past decades, nanoparticles have been viewed as a potentially powerful platform for various applications in biomedical sciences. The possible application of nanoparticles varies from drug delivery agents to novel imaging platforms and surely, some application potential still remains hidden. Thus, it is necessary to broadly study their in vitro behavior in order to assess the precise theranostic potential as well as to distinguish possible threats to human health. Even though nanoparticles are getting more and more attention in current research, still only a limited amount of information is available, especially regarding interactions of ultra-small (< 5 nm) nanoparticles with biological environment and cells. The aim of the work presented herein is to provide the reader with information concerning interactions of various ultra-small nanoparticles (silicon-based, gold, nanodiamonds) with biological environment and human cells. Dose- and time-dependent influence of the various nanoparticles on behavior of different human cells (osteoblasts, monocytes, keratinocytes, mesenchymal stem cells) was established under different conditions, stressing out the importance of protein corona (a layer of proteins originating from cultivation medium attached to nanoparticles). Biocompatibility of two...
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Measurement and toxicity of combustion generated nanoparticles
Sikorová, Jitka ; Topinka, Jan (advisor) ; Klusoň, Petr (referee) ; Bláha, Luděk (referee)
This thesis is focused on nanoparticles produced by internal combustion engines utilized in vehicles. It deals with spatial distribution of nanoparticles within urban areas, impact of alternative fuels usage on particle production and toxicity, and a particle toxicological testing methodology. Monitoring of airborne nanoparticles identified traffic as the main source of airborne nanoparticles in places with heavy traffic load (Prague), as well as in a small city with only local traffic (Čelákovice). Most particles were likely emitted during short episodes of high emissions (e.g. uphill acceleration). During the measurements, high-emission vehicles responsible for a large fraction of the air pollution were also identified. On the other hand, small non-road internal combustion engines, which are not subject to any limit on particle emissions, such as a lawn mower, were operated during the measurement and generated a large number of nanoparticles. The amount and characteristics of the particles produced by combustion depend on the combustion technology and the fuel composition. A large part of the thesis deals with alternative fuels and their effects on the quantity of produced particles and toxicity of organic matter adsorbed on the particles. Hydrotreated vegetable oil (HVO) exhibited the lowest...
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