Bisphenol A (BPA), a synthetic chemical widely used in the production of polycarbonate plastic and epoxy resins, has been associated with a variety of adverse effects in humans including metabolic, immunological, reproductive, and neurodevelopmental effects, raising concern about its health impact. In the EU, it has been classified as toxic to reproduction and as an endocrine disruptor and was thus included in the candidate list of substances of very high concern (SVHC). On this basis, its use has been banned or restricted in some products. As a consequence, industries turned to bisphenol alternatives, such as bisphenol S (BPS) and bisphenol F (BPF), which are now found in various consumer products, as well as in human matrices at a global scale. However, due to their toxicity, these two bisphenols are in the process of being regulated. Other BPA alternatives, whose potential toxicity remains largely unknown due to a knowledge gap, have also started to be used in manufacturing processes. The gradual restriction of the use of BPA underscores the importance of understanding the potential risks associated with its alternatives to avoid regrettable substitutions. This review aims to summarize the current knowledge on the potential hazards related to BPA alternatives prioritized by European Regulatory Agencies based on their regulatory relevance and selected to be studied under the European Partnership for the Assessment of Risks from Chemicals (PARC): BPE, BPAP, BPP, BPZ, BPS-MAE, and TCBPA. The focus is on data related to toxicokinetic, endocrine disruption, immunotoxicity, developmental neurotoxicity, and genotoxicity/carcinogenicity, which were considered the most relevant endpoints to assess the hazard related to those substances. The goal here is to identify the data gaps in BPA alternatives toxicology and hence formulate the future directions that will be taken in the frame of the PARC project, which seeks also to enhance chemical risk assessment methodologies using new approach methodologies (NAMs).
Carcinogenic chemicals, or their metabolites, can be classified as genotoxic or non-genotoxic carcinogens (NGTxCs). Genotoxic compounds induce DNA damage, which can be detected by an established in vitro and in vivo battery of genotoxicity assays. For NGTxCs, DNA is not the primary target, and the possible modes of action (MoA) of NGTxCs are much more diverse than those of genotoxic compounds, and there is no specific in vitro assay for detecting NGTxCs. Therefore, the evaluation of the carcinogenic potential is still dependent on long-term studies in rodents. This 2-year bioassay, mainly applied for testing agrochemicals and pharmaceuticals, is time-consuming, costly and requires very high numbers of animals. More importantly, its relevance for human risk assessment is questionable due to the limited predictivity for human cancer risk, especially with regard to NGTxCs. Thus, there is an urgent need for a transition to new approach methodologies (NAMs), integrating human-relevant in vitro assays and in silico tools that better exploit the current knowledge of the multiple processes involved in carcinogenesis into a modern safety assessment toolbox. Here, we describe an integrative project that aims to use a variety of novel approaches to detect the carcinogenic potential of NGTxCs based on different mechanisms and pathways involved in carcinogenesis. The aim of this project is to contribute suitable assays for the safety assessment toolbox for an efficient and improved, internationally recognized hazard assessment of NGTxCs, and ultimately to contribute to reliable mechanism-based next-generation risk assessment for chemical carcinogens.
- Publikační typ
- časopisecké články MeSH
- přehledy MeSH
Titanium dioxide nanoparticles (TiO2 NPs) are used in a wide range of applications. Although inhalation of NPs is one of the most important toxicologically relevant routes, experimental studies on potential harmful effects of TiO2 NPs using a whole-body inhalation chamber model are rare. In this study, the profile of lymphocyte markers, functional immunoassays, and antioxidant defense markers were analyzed to evaluate the potential adverse effects of seven-week inhalation exposure to two different concentrations of TiO2 NPs (0.00167 and 0.1308 mg TiO2/m3) in mice. A dose-dependent effect of TiO2 NPs on innate immunity was evident in the form of stimulated phagocytic activity of monocytes in low-dose mice and suppressed secretory function of monocytes (IL-18) in high-dose animals. The effect of TiO2 NPs on adaptive immunity, manifested in the spleen by a decrease in the percentage of T-cells, a reduction in T-helper cells, and a dose-dependent decrease in lymphocyte cytokine production, may indicate immunosuppression in exposed mice. The dose-dependent increase in GSH concentration and GSH/GSSG ratio in whole blood demonstrated stimulated antioxidant defense against oxidative stress induced by TiO2 NP exposure.
- Publikační typ
- časopisecké články MeSH
Copper oxide nanoparticles (CuO NPs) are increasingly used in various industry sectors. Moreover, medical application of CuO NPs as antimicrobials also contributes to human exposure. Their toxicity, including toxicity to the immune system and blood, raises concerns, while information on their immunotoxicity is still very limited. The aim of our work was to evaluate the effects of CuO NPs (number concentration 1.40×106 particles/cm3, geometric mean diameter 20.4 nm) on immune/inflammatory response and antioxidant defense in mice exposed to 32.5 μg CuO/m3 continuously for 6 weeks. After six weeks of CuO NP inhalation, the content of copper in lungs and liver was significantly increased, while in kidneys, spleen, brain, and blood it was similar in exposed and control mice. Inhalation of CuO NPs caused a significant increase in proliferative response of T-lymphocytes after mitogenic stimulation and basal proliferative activity of splenocytes. CuO NPs significantly induced the production of IL-12p70, Th1-cytokine IFN-γ and Th2-cytokines IL-4, IL-5. Levels of TNF-α and IL-6 remained unchanged. Immune assays showed significantly suppressed phagocytic activity of granulocytes and slightly decreased respiratory burst. No significant differences in phagocytosis of monocytes were recorded. The percentage of CD3+, CD3+CD4+, CD3+CD8+, and CD3-CD19+ cell subsets in spleen, thymus, and lymph nodes did not differ between exposed and control animals. No changes in hematological parameters were found between the CuO NP exposed and control groups. The overall antioxidant protection status of the organism was expressed by evaluation of GSH and GSSG concentrations in blood samples. The experimental group exposed to CuO NPs showed a significant decrease in GSH concentration in comparison to the control group. In summary, our results indicate that sub-chronic inhalation of CuO NPs can cause undesired modulation of the immune response. Stimulation of adaptive immunity was indicated by activation of proliferation and secretion functions of lymphocytes. CuO NPs elicited pro-activation state of Th1 and Th2 lymphocytes in exposed mice. Innate immunity was affected by impaired phagocytic activity of granulocytes. Reduced glutathione was significantly decreased in mice exposed to CuO NPs.
Levels of DNA damage represent the dynamics between damage formation and removal. Therefore, to better interpret human biomonitoring studies with DNA damage endpoints, an individual's ability to recognize and properly remove DNA damage should be characterized. Relatively few studies have included DNA repair as a biomarker and therefore, assembling and analyzing a pooled database of studies with data on base excision repair (BER) was one of the goals of hCOMET (EU-COST CA15132). A group of approximately 1911 individuals, was gathered from 8 laboratories which run population studies with the comet-based in vitro DNA repair assay. BER incision activity data were normalized and subsequently correlated with various host factors. BER was found to be significantly higher in women. Although it is generally accepted that age is inversely related to DNA repair, no overall effect of age was found, but sex differences were most pronounced in the oldest quartile (>61 years). No effect of smoking or occupational exposures was found. A body mass index (BMI) above 25 kg/m2 was related to higher levels of BER. However, when BMI exceeded 35 kg/m2, repair incision activity was significantly lower. Finally, higher BER incision activity was related to lower levels of DNA damage detected by the comet assay in combination with formamidopyrimidine DNA glycosylase (Fpg), which is in line with the fact that oxidatively damaged DNA is repaired by BER. These data indicate that BER plays a role in modulating the steady-state level of DNA damage that is detected in molecular epidemiological studies and should therefore be considered as a parallel endpoint in future studies.
- MeSH
- DNA-formamidopyrimidinglykosylasa MeSH
- epidemiologické studie MeSH
- kometový test MeSH
- lidé středního věku MeSH
- lidé MeSH
- oprava DNA * genetika MeSH
- poškození DNA * MeSH
- Check Tag
- lidé středního věku MeSH
- lidé MeSH
- mužské pohlaví MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- metaanalýza MeSH
DNA damage and unrepaired or insufficiently repaired DNA double-strand breaks as well as telomere shortening contribute to the formation of structural chromosomal aberrations (CAs). Non-specific CAs have been used in the monitoring of individuals exposed to potential carcinogenic chemicals and radiation. The frequency of CAs in peripheral blood lymphocytes (PBLs) has been associated with cancer risk and the association has also been found in incident cancer patients. CAs include chromosome-type aberrations (CSAs) and chromatid-type aberrations (CTAs) and their sum CAtot. In the present study, we used data from our published genome-wide association studies (GWASs) and extracted the results for 153 DNA repair genes for 607 persons who had occupational exposure to diverse harmful substances/radiation and/or personal exposure to tobacco smoking. The analyses were conducted using linear and logistic regression models to study the association of DNA repair gene polymorphisms with CAs. Considering an arbitrary cutoff level of 5 × 10-3, 14 loci passed the threshold, and included 7 repair pathways for CTA, 4 for CSA, and 3 for CAtot; 10 SNPs were eQTLs influencing the expression of the target repair gene. For the base excision repair pathway, the implicated genes PARP1 and PARP2 encode poly(ADP-ribosyl) transferases with multiple regulatory functions. PARP1 and PARP2 have an important role in maintaining genome stability through diverse mechanisms. Other candidate genes with known roles for CSAs included GTF2H (general transcription factor IIH subunits 4 and 5), Fanconi anemia pathway genes, and PMS2, a mismatch repair gene. The present results suggest pathways with mechanistic rationale for the formation of CAs and emphasize the need to further develop techniques for measuring individual sensitivity to genotoxic exposure.
- Publikační typ
- časopisecké články MeSH
PURPOSE: To assess whether the micronucleus cytome assay (MCyt) reliably detects DNA damage occurring in control and pathological superficial epithelial cells from human conjunctiva. METHODS: Impression cytology samples from the bulbar conjunctiva of 33 healthy controls, eight patients with conjunctival intraepithelial neoplasia (CIN) and eight with mucous membrane pemphigoid (MMP) were examined using the MCyt modified for the ocular surface. RESULTS: The mean number of micronuclei (MNi) in control samples was 0.94 MNi/1000 epithelial cells, with no significant difference between conjunctival quadrants and independent of sex and age. The MCyt assay applied to CIN-affected eyes showed a significantly higher frequency of MNi (18.63/1000 cells), apoptotic cells, nuclear enlargement, multinucleated cells, and keratolysis compared with the corresponding unaffected paired eyes and with the control value. Although the mean MNi frequency in MMP eyes was also higher (1.73 MNi/1000 cells), it did not prove to be statistically different from the control samples. On the other hand, the MMP-affected eyes revealed significantly elevated percentages of cells with snake-like chromatin, multinucleated cells, apoptotic cells, and nuclear buds compared with controls. CONCLUSIONS: Micronucleus cytome assay was adapted as a rapid screening test for genomic instability on the ocular surface. We have determined reference levels for MNi and other nuclear alterations on healthy conjunctiva and demonstrated that particularly frequencies of MNi are significantly elevated in conjunctiva affected by CIN. We demonstrate that MNi are more specific than other nuclear abnormalities and thus can be used for screening of ocular surface neoplasia.
- MeSH
- buněčné jádro MeSH
- epitelové buňky * MeSH
- konjunktiva MeSH
- lidé MeSH
- mikrojaderné testy MeSH
- poškození DNA * MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
Nonspecific structural chromosomal aberrations (CAs) can be found at around 1% of circulating lymphocytes from healthy individuals but the frequency may be higher after exposure to carcinogenic chemicals or radiation. The frequency of CAs has been measured in occupational monitoring and an increased frequency of CAs has also been associated with cancer risk. Alterations in DNA damage repair and telomere maintenance are thought to contribute to the formation of CAs, which include chromosome type of aberrations and chromatid type of aberrations. In the present study, we used the result of our published genome-wide association studies to extract data on 153 DNA repair genes from 866 nonsmoking persons who had no known occupational exposure to genotoxic substances. Considering an arbitrary cut-off level of P< 5 × 10-3, single nucleotide polymorphisms (SNPs) tagging 22 DNA repair genes were significantly associated with CAs and they remained significant at P < 0.05 when adjustment for multiple comparisons was done by the Binomial Sequential Goodness of Fit test. Nucleotide excision repair pathway genes showed most associations with 6 genes. Among the associated genes were several in which mutations manifest CA phenotype, including Fanconi anemia, WRN, BLM and genes that are important in maintaining genome stability, as well as PARP2 and mismatch repair genes. RPA2 and RPA3 may participate in telomere maintenance through the synthesis of the C strand of telomeres. Errors in NHEJ1 function may lead to translocations. The present results show associations with some genes with known CA phenotype and suggest other pathways with mechanistic rationale for the formation of CAs in healthy nonsmoking population.
- MeSH
- běloši genetika MeSH
- celogenomová asociační studie MeSH
- chromozomální aberace * MeSH
- DNA vazebné proteiny genetika MeSH
- dospělí MeSH
- enzymy opravy DNA genetika MeSH
- helikasy RecQ genetika MeSH
- helikáza Wernerova syndromu genetika MeSH
- jednonukleotidový polymorfismus * MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- nekuřáci * MeSH
- oprava chybného párování bází DNA genetika MeSH
- oprava DNA genetika MeSH
- počítačová simulace MeSH
- poly(ADP-ribosa)polymerasy genetika MeSH
- replikační protein A genetika MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- zdraví dobrovolníci pro lékařské studie MeSH
- Check Tag
- dospělí MeSH
- lidé středního věku MeSH
- lidé MeSH
- mladiství MeSH
- mladý dospělý MeSH
- mužské pohlaví MeSH
- senioři nad 80 let MeSH
- senioři MeSH
- ženské pohlaví MeSH
- Publikační typ
- časopisecké články MeSH
- multicentrická studie MeSH
- práce podpořená grantem MeSH
- Geografické názvy
- Česká republika MeSH
- Slovenská republika MeSH
The alkaline comet assay, or single cell gel electrophoresis, is one of the most popular methods for assessing DNA damage in human population. One of the open issues concerning this assay is the identification of those factors that can explain the large inter-individual and inter-laboratory variation. International collaborative initiatives such as the hCOMET project - a COST Action launched in 2016 - represent a valuable tool to meet this challenge. The aims of hCOMET were to establish reference values for the level of DNA damage in humans, to investigate the effect of host factors, lifestyle and exposure to genotoxic agents, and to compare different sources of assay variability. A database of 19,320 subjects was generated, pooling data from 105 studies run by 44 laboratories in 26 countries between 1999 and 2019. A mixed random effect log-linear model, in parallel with a classic meta-analysis, was applied to take into account the extensive heterogeneity of data, due to descriptor, specimen and protocol variability. As a result of this analysis interquartile intervals of DNA strand breaks (which includes alkali-labile sites) were reported for tail intensity, tail length, and tail moment (comet assay descriptors). A small variation by age was reported in some datasets, suggesting higher DNA damage in oldest age-classes, while no effect could be shown for sex or smoking habit, although the lack of data on heavy smokers has still to be considered. Finally, highly significant differences in DNA damage were found for most exposures investigated in specific studies. In conclusion, these data, which confirm that DNA damage measured by the comet assay is an excellent biomarker of exposure in several conditions, may contribute to improving the quality of study design and to the standardization of results of the comet assay in human populations.
- MeSH
- biologické markery krev MeSH
- kometový test metody MeSH
- lidé MeSH
- poškození DNA genetika fyziologie MeSH
- Check Tag
- lidé MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- přehledy MeSH
This optimized protocol (including links to instruction videos) describes a comet-based in vitro DNA repair assay that is relatively simple, versatile, and inexpensive, enabling the detection of base and nucleotide excision repair activity. Protein extracts from samples are incubated with agarose-embedded substrate nucleoids ('naked' supercoiled DNA) containing specifically induced DNA lesions (e.g., resulting from oxidation, UVC radiation or benzo[a]pyrene-diol epoxide treatment). DNA incisions produced during the incubation reaction are quantified as strand breaks after electrophoresis, reflecting the extract's incision activity. The method has been applied in cell culture model systems, human biomonitoring and clinical investigations, and animal studies, using isolated blood cells and various solid tissues. Once extracts and substrates are prepared, the assay can be completed within 2 d.
- MeSH
- buněčné linie MeSH
- kometový test metody MeSH
- lidé MeSH
- oprava DNA * MeSH
- zvířata MeSH
- Check Tag
- lidé MeSH
- zvířata MeSH
- Publikační typ
- časopisecké články MeSH
- práce podpořená grantem MeSH
- Research Support, U.S. Gov't, Non-P.H.S. MeSH
