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Humans are chronically exposed to furan, a potent liver toxicant and carcinogen that occurs in a variety of heat-processed foods. Assessment of human exposure based on the furan content in foods is, however, subject to some uncertainty due to the high volatility of furan. Biomarker monitoring is thus considered an alternative or complementary approach to furan exposure assessment. Previous work suggested that urinary furan metabolites derived from the reaction of cis-2-butene-1,4-dial (BDA), the reactive intermediate of furan, with glutathione (GSH) or amino acids may serve as potential biomarkers of furan exposure. However, some metabolites were also reported to occur in urine of untreated animals, indicating either background contamination via animal feed or endogenous sources, which may limit their suitability as biomarkers of exposure. The overall aim of the present study was to accurately establish the correlation between external dose and concentration of furan metabolites in urine over time and to discriminate against endogenous formation and furan intake via feed. To this end, the furan metabolites GSH-BDA (N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-L-cysteinylglycine), NAcLys-BDA (R-2-(acetylamino)-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)-1-hexanoic acid), NAcCys-BDA-NAcLys (N-acetyl-S-[1-[5-(acetylamino)-5-carboxypentyl]-1H-pyrrol-3-yl]-L-cysteine) and NAcCys-BDA-NAcLys sulfoxide (N-acetyl-S-[1-[5-(acetylamino)-5-carboxypentyl]-1H-pyrrol-3-yl]-L-cysteine sulfoxide) were simultaneously analyzed by stable isotope dilution ESI-LC-MS/MS as unlabeled and [ 13 C 4 ]-furan dependent metabolites following oral administration of a single oral dose of isotopically labelled [ 13 C 4 ]-furan (0.1, 1, 10, 100 and 1000 µg/kg bw) to male and female F344/DuCrl rats. Although a linear correlation between urinary excretion of [ 13 C 4 ]-furan-dependent metabolites was observed, analysis of unlabeled NAcLys-BDA, NAcCys-BDA-NAcLys and NAcCys-BDA-NAcLys sulfoxide revealed substantial, fairly constant urinary background levels throughout the course of the study. Analysis of furan in animal feed excluded feed as a source for these background levels. GSH-BDA was identified as the only furan metabolite without background occurrence, suggesting that it may present a specific biomarker to monitor external furan exposure. Studies in humans are now needed to establish if analysis of urinary GSH-BDA may provide reliable exposure estimates., (© 2024. The Author(s).)

1,1,2-Trifluoroethene (HFO-1123) is anticipated for use as a refrigerant with low global warming potential. Inhalation studies on HFO-1123 in rats indicated a low potential for toxicity (NOAELs ≥ 20,000 ppm). In contrast, single inhalation exposure of Goettingen ® minipigs (≥ 500 ppm) and New Zealand white rabbits (≥ 1250 ppm) resulted in severe toxicity. It has been suggested that these pronounced species-differences in toxicity may be attributable to species-differences in biotransformation of HFO-1123 via the mercapturic acid pathway. Therefore, the overall objective of this study was to evaluate species-differences in glutathione (GSH) dependent in vitro metabolism of HFO-1123 in susceptible versus less susceptible species and humans as a basis for human risk assessment. Biotransformation of HFO-1123 to S-(1,1,2-trifluoroethyl)-L-glutathione (1123-GSH) and subsequent cysteine S-conjugate β-lyase-mediated cleavage of the corresponding cysteine conjugate (1123-CYS) was monitored in hepatic and renal subcellular fractions of mice, rats, minipigs, rabbits, and humans. While 1123-GSH formation occurred at higher rates in rat and rabbit liver S9 compared to minipig and human S9, increased β-lyase cleavage of 1123-CYS was observed in minipig kidney cytosol as compared to cytosolic fractions of other species. Increased β-lyase activity in minipig cytosol was accompanied by time-dependent formation of monofluoroacetic acid (MFA), a highly toxic compound that interferes with cellular energy production via inhibition of aconitase. Consistent with the significantly lower β-lyase activity in human cytosols, the intensity of the MFA signal in human cytosols was only a fraction of the signal obtained in minipig subcellular fractions. Even though the inconsistencies between GSH and β-lyase-dependent metabolism do not allow to draw a firm conclusion on the overall contribution of the mercapturic acid pathway to HFO-1123 biotransformation and toxicity in vivo, the β-lyase data suggest that humans may be less susceptible to HFO-1123 toxicity compared to minipigs., (© 2023. The Author(s).)

The Senate Commission on Food Safety (SKLM) of the German Research Foundation (DFG) has reviewed the currently available data in order to assess the health risks associated with the use of acetaldehyde as a flavoring substance in foods. Acetaldehyde is genotoxic in vitro. Following oral intake of ethanol or inhalation exposure to acetaldehyde, systemic genotoxic effects of acetaldehyde in vivo cannot be ruled out (induction of DNA adducts and micronuclei). At present, the key question of whether acetaldehyde is genotoxic and mutagenic in vivo after oral exposure cannot be answered conclusively. There is also insufficient data on human exposure. Consequently, it is currently not possible to reliably assess the health risk associated with the use of acetaldehyde as a flavoring substance. However, considering the genotoxic potential of acetaldehyde as well as numerous data gaps that need to be filled to allow a comprehensive risk assessment, the SKLM considers that the use of acetaldehyde as a flavoring may pose a safety concern. For reasons of precautionary consumer protection, the SKLM recommends that the scientific base for approval of the intentional addition of acetaldehyde to foods as a flavoring substance should be reassessed., (© 2023 The Authors. Molecular Nutrition & Food Research published by Wiley-VCH GmbH.)

This opinion of the Senate Commission on Food Safety (SKLM) of the German Research Foundation (Deutsche Forschungsgemeinschaft, DFG) presents arguments for an updated risk assessment of diet-related exposure to acrylamide (AA), based on a critical review of scientific evidence relevant to low dose exposure. The SKLM arrives at the conclusion that as long as an appropriate exposure limit for AA is not exceeded, genotoxic effects resulting in carcinogenicity are unlikely to occur. Based on the totality of the evidence, the SKLM considers it scientifically justified to derive a tolerable daily intake (TDI) as a health-based guidance value., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Remdesivir is a prodrug of a nucleoside analog and the first antiviral therapeutic approved for coronavirus disease. Recent cardiac safety concerns and reports on remdesivir-related acute kidney injury call for a better characterization of remdesivir toxicity and understanding of the underlying mechanisms. Here, we performed an in vitro toxicity assessment of remdesivir around clinically relevant concentrations (C max 9 µM) using H9c2 rat cardiomyoblasts, neonatal mouse cardiomyocytes (NMCM), rat NRK-52E and human RPTEC/TERT1 cells as cell models for the assessment of cardiotoxicity or nephrotoxicity, respectively. Due to the known potential of nucleoside analogs for the induction of mitochondrial toxicity, we assessed mitochondrial function in response to remdesivir treatment, early proteomic changes in NMCM and RPTEC/TERT1 cells and the contractile function of NMCM. Short-term treatments (24 h) of H9c2 and NRK-52E cells with remdesivir adversely affected cell viability by inhibition of proliferation as determined by significantly decreased 3 H-thymidine uptake. Mitochondrial toxicity of remdesivir (1.6-3.1 µM) in cardiac cells was evident by a significant decrease in oxygen consumption, a collapse of mitochondrial membrane potential and an increase in lactate secretion after a 24-48-h treatment. This was supported by early proteomic changes of respiratory chain proteins and intermediate filaments that are typically involved in mitochondrial reorganization. Functionally, an impedance-based analysis showed that remdesivir (6.25 µM) affected the beat rate and contractility of NMCM. In conclusion, we identified adverse effects of remdesivir in cardiac and kidney cells at clinically relevant concentrations, suggesting a careful evaluation of therapeutic use in patients at risk for cardiovascular or kidney disease., (© 2022. The Author(s).)

Reactive oxygen species (ROS) and reactive nitrogen species (RNS) are formed as a result of natural cellular processes, intracellular signaling, or as adverse responses associated with diseases or exposure to oxidizing chemical and non-chemical stressors. The action of ROS and RNS, collectively referred to as reactive oxygen and nitrogen species (RONS), has recently become highly relevant in a number of adverse outcome pathways (AOPs) that capture, organize, evaluate and portray causal relationships pertinent to adversity or disease progression. RONS can potentially act as a key event (KE) in the cascade of responses leading to an adverse outcome (AO) within such AOPs, but are also known to modulate responses of events along the AOP continuum without being an AOP event itself. A substantial discussion has therefore been undertaken in a series of workshops named "Mystery or ROS" to elucidate the role of RONS in disease and adverse effects associated with exposure to stressors such as nanoparticles, chemical, and ionizing and non-ionizing radiation. This review introduces the background for RONS production, reflects on the direct and indirect effects of RONS, addresses the diversity of terminology used in different fields of research, and provides guidance for developing a harmonized approach for defining a common event terminology within the AOP developer community., Competing Interests: KL and HY were employed by the company Philip Morris International (PMI). The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Tanabe, O’Brien, Tollefsen, Kim, Chauhan, Yauk, Huliganga, Rudel, Kay, Helm, Beaton, Filipovska, Sovadinova, Garcia-Reyero, Mally, Poulsen, Delrue, Fritsche, Luettich, La Rocca, Yepiskoposyan, Klose, Danielsen, Esterhuizen, Jacobsen, Vogel, Gant, Choi and FitzGerald.)

Food safety problems are a major hindrance to achieving food security, trade, and healthy living in Africa. Fungi and their secondary metabolites, known as mycotoxins, represent an important concern in this regard. Attempts such as agricultural, storage, and processing practices, and creation of awareness to tackle the menace of fungi and mycotoxins have yielded measurable outcomes especially in developed countries, where there are comprehensive mycotoxin legislations and enforcement schemes. Conversely, most African countries do not have mycotoxin regulatory limits and even when available, are only applied for international trade. Factors such as food insecurity, public ignorance, climate change, poor infrastructure, poor research funding, incorrect prioritization of resources, and nonchalant attitudes that exist among governmental organisations and other stakeholders further complicate the situation. In the present review, we discuss the status of mycotoxin regulation in Africa, with emphasis on the impact of weak mycotoxin legislations and enforcement on African trade, agriculture, and health. Furthermore, we discuss the factors limiting the establishment and control of mycotoxins in the region.

In line with recent OECD activities on the use of AOPs in developing Integrated Approaches to Testing and Assessment (IATAs), it is expected that systematic mapping of AOPs leading to systemic toxicity may provide a mechanistic framework for the development and implementation of mechanism-based in vitro endpoints. These may form part of an integrated testing strategy to reduce the need for repeated dose toxicity studies. Focusing on kidney and in particular the proximal tubule epithelium as a key target site of chemical-induced injury, the overall aim of this work is to contribute to building a network of AOPs leading to nephrotoxicity. Current mechanistic understanding of kidney injury initiated by 1) inhibition of mitochondrial DNA polymerase γ (mtDNA Polγ), 2) receptor mediated endocytosis and lysosomal overload, and 3) covalent protein binding, which all present fairly well established, common mechanisms by which certain chemicals or drugs may cause nephrotoxicity, is presented and systematically captured in a formal description of AOPs in line with the OECD AOP development programme and in accordance with the harmonized terminology provided by the Collaborative Adverse Outcome Pathway Wiki. The relative level of confidence in the established AOPs is assessed based on evolved Bradford-Hill weight of evidence considerations of biological plausibility, essentiality and empirical support (temporal and dose-response concordance)., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Mally and Jarzina.)

Recent analyses conducted by German official food control reported detection of the aromatic amides N-(2,4-dimethylphenyl)acetamide (NDPA), N-acetoacetyl-m-xylidine (NAAX) and 3-hydroxy-2-naphthanilide (Naphthol AS) in cold water extracts from certain food contact materials made from paper or cardboard, including paper straws, paper napkins, and cupcake liners. Because aromatic amides may be cleaved to potentially genotoxic primary amines upon oral intake, these findings raise concern that transfer of NDPA, NAAX and Naphthol AS from food contact materials into food may present a risk to human health. The aim of the present work was to assess the stability of NDPA, NAAX and Naphthol AS and potential cleavage to 2,4-dimethylaniline (2,4-DMA) and aniline during simulated passage through the gastrointestinal tract using static in vitro digestion models. Using the digestion model established by the National Institute for Public Health and the Environment (RIVM, Bilthoven, NL) and a protocol recommended by the European Food Safety Authority, potential hydrolysis of the aromatic amides to the respective aromatic amines was assessed by LC-MS/MS following incubation of the aromatic amides with digestive fluid simulants. Time-dependent hydrolysis of NDPA and NAAX resulting in formation of the primary aromatic amine 2,4-DMA was consistently observed in both models. The highest rate of cleavage of NDPA and NAAX was recorded following 4 h incubation with 0.07 M HCl as gastric-juice simulant, and amounted to 0.21% and 0.053%, respectively. Incubation of Naphthol AS with digestive fluid simulants did not give rise to an increase in the concentration of aniline above the background that resulted from the presence of aniline as an impurity of the test compound. Considering the lack of evidence for aniline formation from Naphthol AS and the extremely low rate of hydrolysis of the amide bonds of NDPA and NAAX during simulated passage through the gastrointestinal tract that gives rise to only very minor amounts of the potentially mutagenic and/or carcinogenic aromatic amine 2,4-DMA, risk assessment based on assumption of 100% cleavage to the primary aromatic amines would appear to overestimate health risks related to the presence of aromatic amides in food contact materials., (© 2022. The Author(s).)

Application of adverse outcome pathways (AOP) and integration of quantitative in vitro to in vivo extrapolation (QIVIVE) may support the paradigm shift in toxicity testing to move from apical endpoints in test animals to more mechanism-based in vitro assays. Here, we developed an AOP of proximal tubule injury linking a molecular initiating event (MIE) to a cascade of key events (KEs) leading to lysosomal overload and ultimately to cell death. This AOP was used as a case study to adopt the AOP concept for systemic toxicity testing and risk assessment based on in vitro data. In this AOP, nephrotoxicity is thought to result from receptor-mediated endocytosis (MIE) of the chemical stressor, disturbance of lysosomal function (KE1), and lysosomal disruption (KE2) associated with release of reactive oxygen species and cytotoxic lysosomal enzymes that induce cell death (KE3). Based on this mechanistic framework, in vitro readouts reflecting each KE were identified. Utilizing polymyxin antibiotics as chemical stressors for this AOP, the dose-response for each in vitro endpoint was recorded in proximal tubule cells from rat (NRK-52E) and human (RPTEC/TERT1) in order to (1) experimentally support the sequence of key events (KEs), to (2) establish quantitative relationships between KEs as a basis for prediction of downstream KEs based on in vitro data reflecting early KEs and to (3) derive suitable in vitro points of departure for human risk assessment. Time-resolved analysis was used to support the temporal sequence of events within this AOP. Quantitative response-response relationships between KEs established from in vitro data on polymyxin B were successfully used to predict in vitro toxicity of other polymyxin derivatives. Finally, a physiologically based kinetic (PBK) model was utilized to transform in vitro effect concentrations to a human equivalent dose for polymyxin B. The predicted in vivo effective doses were in the range of therapeutic doses known to be associated with a risk for nephrotoxicity. Taken together, these data provide proof-of-concept for the feasibility of in vitro based risk assessment through integration of mechanistic endpoints and reverse toxicokinetic modelling., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Jarzina, Di Fiore, Ellinger, Reiser, Frank, Glaser, Wu, Taverne, Kramer and Mally.)

Prerequisite to any biological laboratory assay employing living animals is consideration about its necessity, feasibility, ethics and the potential harm caused during an experiment. The imperative of these thoughts has led to the formulation of the 3R-principle, which today is a pivotal scientific standard of animal experimentation worldwide. The rising amount of laboratory investigations utilizing living animals throughout the last decades, either for regulatory concerns or for basic science, demands the development of alternative methods in accordance with 3R to help reduce experiments in mammals. This demand has resulted in investigation of additional vertebrate species displaying favourable biological properties. One prominent species among these is the zebrafish ( Danio rerio ), as these small laboratory ray-finned fish are well established in science today and feature outstanding biological characteristics. In this review, we highlight the advantages and general prerequisites of zebrafish embryos and larvae before free-feeding stages for toxicological testing, with a particular focus on cardio-, neuro, hepato- and nephrotoxicity. Furthermore, we discuss toxicokinetics, current advances in utilizing zebrafish for organ toxicity testing and highlight how advanced laboratory methods (such as automation, advanced imaging and genetic techniques) can refine future toxicological studies in this species.

Ochratoxin A (OTA) is a widespread food contaminant, with exposure estimated to range from 0.64 to 17.79 ng/kg body weight (bw) for average consumers and from 2.40 to 51.69 ng/kg bw per day for high consumers. Current exposure estimates are, however, associated with considerable uncertainty. While biomarker-based approaches may contribute to improved exposure assessment, there is yet insufficient data on urinary metabolites of OTA and their relation to external dose to allow reliable estimates of daily intake. This study was designed to assess potential species differences in phase II biotransformation in vitro and to establish a correlation between urinary OTA-derived glucuronides and mercapturic acids and external exposure in rats in vivo. In vitro analyses of OTA metabolism using the liver S9 of rats, humans, rabbits and minipigs confirmed formation of an OTA glucuronide but provided no evidence for the formation of OTA-derived mercapturic acids to support their use as biomarkers. Similarly, OTA-derived mercapturic acids were not detected in urine of rats repeatedly dosed with OTA, while indirect analysis using enzymatic hydrolysis of the urine samples prior to LC-MS/MS established a linear relationship between urinary glucuronide excretion and OTA exposure. These results support OTA-derived glucuronides but not mercapturic acids as metabolites suitable for biomonitoring.

This study assessed relationship between hair loss and professional success by characterizing severity of hair loss among current white male chief executive officers (CEOs) in the United States. This observational study of androgenic alopecia among business executives was conducted by grading anterior and lateral view photographs of CEOs aged 50-69 years. Blinded, independent two-reviewer assessments of androgenic alopecia were performed based on the Hamilton-Norwood Hair Loss Scale. In all, 68 CEOs were included in this review, with 34 individuals each in the age groups of 50-59 and 60-69 years. Significant hair loss occurred in 18% (6/34) of the CEOs aged 50-59 years and in 35% (12/34) of those aged 60-69 years. The findings of the study established that among white male CEOs aged 50-69 years, significant hair loss occurred less frequently than previously reported for the general white male population.

Due to an increasing demand for testing of new and existing chemicals and legal restrictions for the use of animals, there is a strong need for alternative approaches to assess systemic toxicity. Embryonic and larval zebrafish (Danio rerio) are increasingly recognized as a promising alternative whole-animal model that may be able to overcome limitations of cell-based in vitro assays and bridge the gap between high-throughput in vitro screening and low-throughput in vivo tests in animals. Despite the relatively simple anatomical structure of the zebrafish larval kidney (pronephros) - composed of only two nephrons - the pronephros shares major functions and cell types with mammalian nephrons. Glomerular filtration begins at 48 h post fertilization. The aim of the present study was to investigate if early zebrafish larvae might be a suitable model for nephrotoxicity testing. On day 3 post fertilization, larval zebrafish were treated with selected nephrotoxins (aristolochic acid, cadmium chloride, potassium bromate, ochratoxin A, gentamicin) for 48 h. Histological evaluation of zebrafish larvae exposed to model nephrotoxins revealed tubule injury as evidenced by dilated tubules with loss of the brush border, tubule cell necrosis and disorganization of the tubular epithelium. These changes were most severe after treatment with gentamicin, which also impaired pronephros function as evidenced by reduced clearance of FITC-dextran. Whole-mount in situ hybridization showing loss of cdh17 expression revealed site-specific injury to the proximal tubule segment. Analysis of genes previously identified as novel biomarkers of kidney injury in mammals showed upregulation of the kidney injury marker genes heme oxygenase 1 (hmox1), clusterin (clu), secreted phosphoprotein/osteopontin (spp1), connective tissue growth factor (ctgf) and kim-1 (havcr-1) in response to nephrotoxin treatment, although the response of individual genes varied across compounds. Consistent with the severity of lesions and impaired kidney function, the most prominent gene expression changes occurred in larvae exposed to gentamicin. Overall, our results suggest that larval zebrafish may be a suitable alternative model organism for nephrotoxicity screening, yet further improvements and integration with quantitative in vitro to in vivo extrapolation will be needed to predict human toxicity., Competing Interests: Declaration of Competing Interest The authors declare no conflict of interests., (Copyright © 2021 Elsevier B.V. All rights reserved.)

It was generally accepted as a default assumption that No-Observed-Adverse-Effect Levels (NOAELs) or Lowest-Observed-Adverse-Effect Levels (LOAELs) in long-term toxicity studies are lower than in short-term ones, i.e. the toxic potency increases with prolonged exposure duration. Recent studies on pesticides and industrial chemicals reported that subacute, subchronic or chronic NOAELs/LOAELs are similar when study design factors are appropriately considered. We investigated whether these findings also apply to certain food constituents. After reviewing subchronic and chronic toxicity studies on more than 100 compounds, a total of 32 compounds could be included in the analysis. Geometric mean (GM) values of subchronic vs. chronic NOAEL or LOAEL ratios ranged from 1.0 to 2.0, with a geometric standard deviation from 2.2 to 4.2, which is consistent with data reported in the literature. While for many of the investigated compounds the ratio is around 1 - suggesting that health-based guidance values could appropriately be derived from subchronic toxicity studies - our study also identified some substances with higher ratios leading to a GM of around 2. The EFSA Scientific Committee suggested to apply an uncertainty factor of 2 to extrapolate from subchronic to chronic studies and, as a precautionary approach, we concur with this suggestion., (Copyright © 2020 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Infants and young children (IYC) remain the most vulnerable population group to environmental hazards worldwide, especially in economically developing regions such as sub-Saharan Africa (SSA). As a result, several governmental and non-governmental institutions including health, environmental and food safety networks and researchers have been proactive toward protecting this group. Mycotoxins, toxic secondary fungal metabolites, contribute largely to the health risks of this young population. In SSA, the scenario is worsened by socioeconomic status, poor agricultural and storage practices, and low level of awareness, as well as the non-establishment and lack of enforcement of regulatory limits in the region. Studies have revealed mycotoxin occurrence in breast milk and other weaning foods. Of concern is the early exposure of infants to mycotoxins through transplacental transfer and breast milk as a consequence of maternal exposure, which may result in adverse health effects. The current paper presents an overview of mycotoxin occurrence in foods intended for IYC in SSA. It discusses the imperative evidence of mycotoxin exposure of this population group in SSA, taking into account consumption data and the occurrence of mycotoxins in food, as well as biomonitoring approaches. Additionally, it discusses the health implications associated with IYC exposure to mycotoxins in SSA.

Furan is a liver toxicant and carcinogen that occurs in heat-processed foods. Due to its volatility, analysis of furan in food does not provide reliable estimates of exposure. Biomarker-based approaches offer the opportunity to more accurately assess human exposure, but a correlation between concentrations of potential biomarkers of furan exposure and external dose has not been established. Bioactivation of furan and subsequent reaction of cis-2-butene-1,4-dial (BDA) with cellular nucleophiles gives rise to a range of metabolites that may serve as biomarkers of furan exposure. In this study, N-[4-carboxy-4-(3-mercapto-1H-pyrrol-1-yl)-1-oxobutyl]-L-cysteinylglycine cyclic sulfide, a mono-glutathione adduct of BDA (GSH-BDA), and R-2-acetylamino-6-(2,5-dihydro-2-oxo-1H-pyrrol-1-yl)-1-hexanoic acid, an adduct of BDA with N α -acetyl-L-lysine (NAcLys-BDA), were synthesized and analysed by LC-MS/MS in urine of rats treated with furan at 0, 0.1, 0.5 and 2.0 mg/kg bw for 5 and 28 days. GSH-BDA and NAcLys-BDA were both excreted in a dose-related manner. 24 h excretion rates ranged between 0.6 and 1.1% of the administered dose for GSH-BDA, and 1.4-2.1% for NAcLys-BDA. In contrast to GSH-BDA, NAcLys-BDA was also present in urine of controls, suggesting either endogenous formation or background exposure. Overall, the close correlation between urinary furan metabolites and external dose provides experimental support for biomarker-based approaches to monitor human exposure to furan., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

The European Commission asked EFSA to update their 2006 opinion on ochratoxin A (OTA) in food. OTA is produced by fungi of the genus Aspergillus and Penicillium and found as a contaminant in various foods. OTA causes kidney toxicity in different animal species and kidney tumours in rodents. OTA is genotoxic both in vitro and in vivo ; however, the mechanisms of genotoxicity are unclear. Direct and indirect genotoxic and non-genotoxic modes of action might each contribute to tumour formation. Since recent studies have raised uncertainty regarding the mode of action for kidney carcinogenicity, it is inappropriate to establish a health-based guidance value (HBGV) and a margin of exposure (MOE) approach was applied. For the characterisation of non-neoplastic effects, a BMDL 10 of 4.73 μg/kg body weight (bw) per day was calculated from kidney lesions observed in pigs. For characterisation of neoplastic effects, a BMDL 10 of 14.5 μg/kg bw per day was calculated from kidney tumours seen in rats. The estimation of chronic dietary exposure resulted in mean and 95th percentile levels ranging from 0.6 to 17.8 and from 2.4 to 51.7 ng/kg bw per day, respectively. Median OTA exposures in breastfed infants ranged from 1.7 to 2.6 ng/kg bw per day, 95th percentile exposures from 5.6 to 8.5 ng/kg bw per day in average/high breast milk consuming infants, respectively. Comparison of exposures with the BMDL 10 based on the non-neoplastic endpoint resulted in MOEs of more than 200 in most consumer groups, indicating a low health concern with the exception of MOEs for high consumers in the younger age groups, indicating a possible health concern. When compared with the BMDL 10 based on the neoplastic endpoint, MOEs were lower than 10,000 for almost all exposure scenarios, including breastfed infants. This would indicate a possible health concern if genotoxicity is direct. Uncertainty in this assessment is high and risk may be overestimated., (© 2020 European Food Safety Authority. EFSA Journal published by John Wiley and Sons Ltd on behalf of European Food Safety Authority.)

Recently, epidemiological studies have suggested that fluoride is a human developmental neurotoxicant that reduces measures of intelligence in children, placing it into the same category as toxic metals (lead, methylmercury, arsenic) and polychlorinated biphenyls. If true, this assessment would be highly relevant considering the widespread fluoridation of drinking water and the worldwide use of fluoride in oral hygiene products such as toothpaste. To gain a deeper understanding of these assertions, we reviewed the levels of human exposure, as well as results from animal experiments, particularly focusing on developmental toxicity, and the molecular mechanisms by which fluoride can cause adverse effects. Moreover, in vitro studies investigating fluoride in neuronal cells and precursor/stem cells were analyzed, and 23 epidemiological studies published since 2012 were considered. The results show that the margin of exposure (MoE) between no observed adverse effect levels (NOAELs) in animal studies and the current adequate intake (AI) of fluoride (50 µg/kg b.w./day) in humans ranges between 50 and 210, depending on the specific animal experiment used as reference. Even for unusually high fluoride exposure levels, an MoE of at least ten was obtained. Furthermore, concentrations of fluoride in human plasma are much lower than fluoride concentrations, causing effects in cell cultures. In contrast, 21 of 23 recent epidemiological studies report an association between high fluoride exposure and reduced intelligence. The discrepancy between experimental and epidemiological evidence may be reconciled with deficiencies inherent in most of these epidemiological studies on a putative association between fluoride and intelligence, especially with respect to adequate consideration of potential confounding factors, e.g., socioeconomic status, residence, breast feeding, low birth weight, maternal intelligence, and exposure to other neurotoxic chemicals. In conclusion, based on the totality of currently available scientific evidence, the present review does not support the presumption that fluoride should be assessed as a human developmental neurotoxicant at the current exposure levels in Europe.

Current reports increasingly associate dietary "advanced glycation end products" ("AGEs") resulting from the Maillard reaction (glycation) between reducing sugars and amino compounds in foods with pathophysiological consequences, such as chronic inflammation, atherosclerosis, and metabolic syndrome. Heated foods are therefore suggested to pose a potential risk for human health. However, studies in this field are very often based on questionable quantitative data and inadequate structural characterization. To improve the situation, the present perspective suggests quality criteria for future studies and the assessment of the currently available literature.

A comprehensive definition introducing the term "modified mycotoxins" to encompass all possible forms in which mycotoxins and their modifications can occur was recently proposed and has rapidly gained wide acceptance within the scientific community. It is becoming increasingly evident that exposure to such modified mycotoxins due to their presence in food and feed has the potential to pose a substantial additional risk to human and animal health. Zearalenone (ZEN) is a well-characterized Fusarium toxin. Considering the diversity of modified forms of ZEN occurring in food and feed, the toxicologically relevant endocrine activity of many of these metabolites, and the fact that modified forms add to a dietary exposure which approaches the tolerable daily intake by free ZEN alone, modified forms of ZEN present an ideal case study for critical evaluation of modified mycotoxins in food safety. Following a summary of recent scientific opinions of EFSA dealing with health risk assessment of ZEN alone or in combination with its modified forms, uncertainties and data gaps are highlighted. Issues essential for evaluation and prioritization of modified mycotoxins in health risk assessment are identified and discussed, including opportunities to improve exposure assessment using biomonitoring data. Further issues such as future consideration of combinatory effects of the parent toxin with its modified forms and also other compounds co-occurring in food and feed are addressed. With a particular focus on ZEN, the most pressing challenges associated with health risk assessment of modified mycotoxins are identified and recommendations for further research to fill data gaps and reduce uncertainties are made.

Isoflavones are secondary plant constituents of certain foods and feeds such as soy, linseeds, and red clover. Furthermore, isoflavone-containing preparations are marketed as food supplements and so-called dietary food for special medical purposes to alleviate health complaints of peri- and postmenopausal women. Based on the bioactivity of isoflavones, especially their hormonal properties, there is an ongoing discussion regarding their potential adverse effects on human health. This review evaluates and summarises the evidence from interventional and observational studies addressing potential unintended effects of isoflavones on the female breast in healthy women as well as in breast cancer patients and on the thyroid hormone system. In addition, evidence from animal and in vitro studies considered relevant in this context was taken into account along with their strengths and limitations. Key factors influencing the biological effects of isoflavones, e.g., bioavailability, plasma and tissue concentrations, metabolism, temporality (pre- vs. postmenopausal women), and duration of isoflavone exposure, were also addressed. Final conclusions on the safety of isoflavones are guided by the aim of precautionary consumer protection.

Ochratoxin A (OTA) is a potent renal carcinogen but its mechanism has not been fully resolved. In vitro and in vivo gene expression studies consistently revealed down-regulation of gene expression as the predominant transcriptional response to OTA. Based on the importance of specific histone acetylation marks in regulating gene transcription and our recent finding that OTA inhibits histone acetyltransferases (HATs), leading to loss of acetylation of histones and non-histone proteins, we hypothesized that OTA-mediated repression of gene expression may be causally linked to HAT inhibition and loss of histone acetylation. In this study, we used a novel mass spectrometry approach employing chemical 13 C-acetylation of unmodified lysine residues for quantification of post-translational acetylation sites to identify site-specific alterations in histone acetylation in human kidney epithelial cells (HK-2) exposed to OTA. These results showed OTA-mediated hypoacetylation at almost all lysine residues of core histones, including loss of acetylation at H3K9 and H3K14, which are hallmarks of gene activation. ChIP-qPCR used to establish a possible link between H3K9 or H3K14 hypoacetylation and OTA-mediated down-regulation of selected genes (AMIGO2, CLASP2, CTNND1) confirmed OTA-mediated H3K9 hypoacetylation at promoter regions of these genes. Integrated analysis of OTA-mediated genome-wide changes in H3K9 acetylation identified by ChIP-Seq with published gene expression data further demonstrated that among OTA-responsive genes almost 80% of hypoacetylated genes were down-regulated, thus confirming an association between H3K9 acetylation status and gene expression of these genes. However, only 7% of OTA repressed genes showed loss of H3K9 acetylation within promoter regions. Interestingly, however, GO analysis and functional enrichment of down-regulated genes showing loss of H3K9 acetylation at their respective promoter regions revealed enrichment of genes involved in the regulation of transcription, including a number of transcription factors that are predicted to directly or indirectly regulate the expression of 98% of OTA repressed genes. Thus, it is possible that histone acetylation changes in a fairly small set of genes but with key function in transcriptional regulation may trigger a cascade of events that may lead to overall repression of gene expression. Taken together, our data provide evidence for a mechanistic link between loss of H3K9 acetylation as a consequence of OTA-mediated inhibition of HATs and repression of gene expression by OTA, thereby affecting cellular processes critical to tumorigenesis.

Exposure assessment is a fundamental part of the risk assessment paradigm, but can often present a number of challenges and uncertainties. This is especially the case for process contaminants formed during the processing, e.g. heating of food, since they are in part highly reactive and/or volatile, thus making exposure assessment by analysing contents in food unreliable. New approaches are therefore required to accurately assess consumer exposure and thus better inform the risk assessment. Such novel approaches may include the use of biomarkers, physiologically based kinetic (PBK) modelling-facilitated reverse dosimetry, and/or duplicate diet studies. This review focuses on the state of the art with respect to the use of biomarkers of exposure for the process contaminants acrylamide, 3-MCPD esters, glycidyl esters, furan and acrolein. From the overview presented, it becomes clear that the field of assessing human exposure to process-related contaminants in food by biomarker monitoring is promising and strongly developing. The current state of the art as well as the existing data gaps and challenges for the future were defined. They include (1) using PBK modelling and duplicate diet studies to establish, preferably in humans, correlations between external exposure and biomarkers; (2) elucidation of the possible endogenous formation of the process-related contaminants and the resulting biomarker levels; (3) the influence of inter-individual variations and how to include that in the biomarker-based exposure predictions; (4) the correction for confounding factors; (5) the value of the different biomarkers in relation to exposure scenario's and risk assessment, and (6) the possibilities of novel methodologies. In spite of these challenges it can be concluded that biomarker-based exposure assessment provides a unique opportunity to more accurately assess consumer exposure to process-related contaminants in food and thus to better inform risk assessment.

Recently, bile acids (BAs) were reported as promising markers for drug-induced liver injury (DILI). BAs have been suggested to correlate with hepatocellular and hepatobiliary damage; however a clear connection of BA patterns with different types of DILI remains to be established. To investigate if BAs can improve the assessment of liver injury, 20 specific BAs were quantitatively profiled via LC-MS/MS in plasma and liver tissue in a model of methapyrilene-induced liver injury in rats. Methapyrilene, a known hepatotoxin was dosed daily over 14-days at doses of 30 and 80mg/kg, followed by a recovery phase of 10days. Conventional preclinical safety endpoints were related to BA perturbations and to hepatic gene expression profiling for a mechanistic interpretation of effects. Histopathological signs of hepatocellular and hepatobiliary damage with significant changes of clinical chemistry markers were accompanied by significantly increased levels of indivdual BAs in plasma and liver tissue. BA perturbations were already evident at the earliest time point after 30mg/kg treatment, and thereby indicating better sensitivity than clinical chemistry parameters. Furthermore, the latter markers suggested recovery of liver injury, whereas BA levels in plasma and liver remained significantly elevated during the recovery phase, in line with persistent histopathological findings of bile duct hyperplasia (BDH) and bile pigment deposition. Gene expression profiling revealed downregulation of genes involved in BA synthesis (AMACR, BAAT, ACOX2) and hepatocellular uptake (NTCP, OATs), and upregulation for efflux transporters (MRP2, MRP4), suggesting an adaptive hepatocellular protection mechanism against cytotoxic bile acid accumulation. In summary, our data suggests that specific BAs with high reliability such as cholic acid (CA) and chenodeoxycholic acid (CDCA) followed by glycocholic acid (GCA), taurocholic acid (TCA) and deoxycholic acid (DCA) can serve as additional biomarkers for hepatocellular/hepatobiliary damage in the liver in rat toxicity studies., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Furan hepatotoxicity is thought to be linked to covalent binding of its reactive metabolite, cis-2-butene-1,4-dial, to hepatic proteins critical for cell homeostasis and survival. We previously identified 61 putative furan target proteins, which participate in various cellular processes including carbohydrate metabolism, fatty acid β-oxidation, adenosine triphosphate (ATP) synthesis, protein folding and maintenance of redox homeostasis. To further investigate the biological significance of target protein modification, this study was designed to determine the impact of furan on the activity of key target enzymes involved in glycolysis, β-oxidation, ATP synthesis, and redox regulation in rat liver, and to link these functional changes to alterations in cellular processes. While cis-2-butene-1,4-dial inhibited thioredoxin 1 (Txn1) in a cell-free assay, in livers of rats treated with a single high dose of furan Txn1 activity was markedly increased due to rapid up-regulation of Txn1 mRNA expression. Significant inhibition of glyceraldehyde-3-phosphate dehydrogenase and metabolic changes consistent with blocked glycolytic breakdown of glucose were observed in rat liver in response to a single high dose of furan. In contrast, furan treatment resulted in increased activity of enoyl-CoA hydratase and enhanced production of ketone bodies, indicative of increased utilization of fatty acids as energy source. Consistent with changes in TCA cycle metabolites, furan treatment resulted in a reduction of succinate dehydrogenase activity, supporting mitochondrial dysfunction as a critical event in furan toxicity. No significant changes in target protein function were observed following repeated administration of furan at lower dose (0.1 and 0.5mg/kg bw for 4 weeks) closer to estimated human exposure to furan via food. Although the relative contribution of furan mediated alterations in metabolic pathways and antioxidant defense to the overall toxic response to furan, including considerations of dose and time, remains to be established, our work contributes to mapping biological processes and toxicity pathways modulated by reactive electrophiles., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Zearalenone (ZEN), a mycotoxin with high estrogenic activity in vitro and in vivo, is a widespread food contaminant that is commonly detected in maize, wheat, barley, sorghum, rye and other grains. Human exposure estimates based on analytical data on ZEN occurrence in various food categories and food consumption data suggest that human exposure to ZEN and modified forms of ZEN may be close to or even exceed the tolerable daily intake (TDI) derived by the European Food Safety Authority (EFSA) for some consumer groups. Considering the inherent uncertainties in estimating dietary intake of ZEN that may lead to an under- or overestimation of ZEN exposure and consequently human risk and current lack of data on vulnerable consumer groups, there is a clear need for more comprehensive and reliable exposure data to refine ZEN risk assessment. Human biomonitoring (HBM) is increasingly being recognized as an efficient and cost-effective way of assessing human exposure to food contaminants, including mycotoxins. Based on animal and (limited) human data on the toxicokinetics of ZEN, it appears that excretion of ZEN and its major metabolites may present suitable biomarkers of ZEN exposure. In view of the limitations of available dietary exposure data on ZEN and its modified forms, the purpose of this review is to provide an overview of recent studies utilizing HBM to monitor and assess human exposure to ZEN. Considerations are given to animal and human toxicokinetic data relevant to HBM, analytical methods, and available HBM data on urinary biomarkers of ZEN exposure in different cohorts.

Altered expression of tumor suppressor genes and oncogenes, which is regulated in part at the level of DNA methylation, is an important event involved in non-genotoxic carcinogenesis. This may serve as a marker for early detection of non-genotoxic carcinogens. Therefore, we evaluated the effects of non-genotoxic hepatocarcinogens, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), hexachlorobenzene (HCB), methapyrilene (MPY) and male rat kidney carcinogens, d-limonene, p-dichlorobenzene (DCB), chloroform and ochratoxin A (OTA) on global and CpG island promoter methylation in their respective target tissues in rats. No significant dose-related effects on global DNA hypomethylation were observed in tissues of rats compared to vehicle controls using LC-MS/MS in response to short-term non-genotoxic carcinogen exposure. Initial experiments investigating gene-specific methylation using methylation-specific PCR and bisulfite sequencing, revealed partial methylation of p16 in the liver of rats treated with HCB and TCDD. However, no treatment related effects on the methylation status of Cx32, e-cadherin, VHL, c-myc, Igfbp2, and p15 were observed. We therefore applied genome-wide DNA methylation analysis using methylated DNA immunoprecipitation combined with microarrays to identify alterations in gene-specific methylation. Under the conditions of our study, some genes were differentially methylated in response to MPY and TCDD, whereas d-limonene, DCB and chloroform did not induce any methylation changes. 90-day OTA treatment revealed enrichment of several categories of genes important in protein kinase activity and mTOR cell signaling process which are related to OTA nephrocarcinogenicity., (Copyright © 2015 Elsevier Inc. All rights reserved.)

Increasing experimental and clinical evidence suggest a contribution of non-drug related risk factors (e.g., underlying disease, bacterial/viral infection) to idiosyncratic drug reactions (IDR). Our previous work showed that co-treatment with bacterial endotoxin (LPS) and therapeutic doses of diclofenac (Dcl), an analgesic associated with drug idiosyncrasy in patients, induced severe hepatotoxicity in rats. Here, we used an integrated discovery to targeted LC-MS proteomics approach to identify mechanistically relevant liver and plasma proteins modulated by LPS/Dcl treatment, potentially applicable as early markers for IDRs. Based on pre-screening results and their role in liver toxicity, 47 liver and 15 plasma proteins were selected for targeted LC-MS analysis. LPS alone significantly changed the levels of 19 and 3 of these proteins, respectively. T-kininogen-1, previously suggested as a marker of drug-induced liver injury, was markedly elevated in plasma after repeated Dcl treatment in the absence of hepatotoxicity, possibly indicating clinically silent stress. Dcl both alone and in combination with LPS, caused up-regulation of the ATP synthase subunits (ATP5J, ATPA, and ATPB), suggesting that Dcl may sensitize cells against additional stress factors, such as LPS through generation of mitochondrial stress. Additionally, depletion of plasma fibrinogen was observed in the co-treatment group, consistent with an increased hepatic fibrin deposition and suspected contribution of the hemostatic system to IDRs. In contrast, several proteins previously suggested as liver biomarkers, such as clusterin, did not correlate with liver injury in this model. Taken together, these analyses revealed proteomic changes in a rat model of LPS/Dcl co-administration that could offer mechanistic insight and may serve as biomarkers or safety alert for a drug's potential to cause IDRs., (Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.)

Based on the outcome of a number of experimental studies, progesterone (PROG) holds promise as a new therapy for stroke. To understand more about the mechanisms involved, we administered PROG (or the major metabolite, allopregnanolone, ALLO), intra-peritoneally, for a period of 24 h after transient middle cerebral artery occlusion to male mice variably expressing intracellular progesterone receptors (iPR) A/B. Effects on infarct volume and neurogenesis were then assessed up to 1 month later. Predictably, infarct volume in wild-type mice receiving either drug was significantly smaller. However, mice heterozygous for iPRs A/B showed protection by ALLO but not by PROG. There was robust amplification of cell division in the wall of the lateral ventricle on the injured side of the brain, these cells migrated into the striatum and lateral cortex, and a significant number survived for at least 3 weeks. However, very few doublecortin-positive cells emerged from the subventricular zone and subsequent expression of NeuN in these newborn neurons was extremely rare. Neither PROG nor ALLO amplified the rate of neurogenesis, suggesting that the long-term benefits of acute drug administration results from tissue preservation. Male mice derive long-lasting benefit from progesterone and allopregnanolone after ischemic stroke. In mice heterozygous for iPRs, only allopregnanolone proved effective, suggesting distinct mechanisms. Abundant newborn cells were found in the wall of the lateral ventricle on the injured side (many doublecortin-positive), some migrated into the striatum and lateral cortex, but very few survived as mature neurons. Neurosteroid administration did not amplify this process., (© 2014 International Society for Neurochemistry.)

Inhibition of the glucagon receptor (GCGR) has been identified as a potential therapeutic approach for the treatment of type 2 diabetes. However, a small molecule drug candidate antagonizing GCGR (BAY16) failed during preclinical drug development, in part due to drug induced hepatotoxicity in animals. Since there is evidence to suggest that endogenous GCGR signaling might be important for hepatocyte survival, we hypothesized that on-target effects, i.e., modulation of GCGR activity by BAY16, may contribute to BAY16 hepatotoxicity and associated gene expression changes in rats. To understand the role of GCGR inhibition in BAY16 toxicity, we analyzed cell viability and gene expression profiles in non-silenced and GCGR-targeting siRNA transfected primary rat hepatocytes with and without exposure to BAY16 to discriminate between on- and off-target effects of BAY16. siRNA-mediated silencing of the GCGR did not affect cell viability in primary rat hepatocytes, indicating that cytotoxicity of BAY16 occurs independent of its pharmacological effects. In support of this, gene expression analysis of GCGR silenced hepatocytes revealed no transcriptional alterations relevant to toxicity. In contrast, BAY16 caused a concentration-dependent decrease in cell viability, along with changes in the expression of genes associated with altered xenobiotic metabolism, oxidative stress, increased fatty acid synthesis, and alterations in cholesterol and bile acid metabolic processes. Based on gene expression data, it appears that hepatocytes inhibit cholesterol synthesis and increase detoxifying and eliminating processes in order to protect themselves from accumulation of bile acids, cholesterol or drug intermediates. Importantly, comparison of transcriptional changes in the absence and presence of GCGR revealed that the same pathways were affected in both silenced and non-silenced hepatocytes, indicating that BAY16 toxicity occurs independent of the GCGR receptor., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

As the term "masked mycotoxins" encompasses only conjugated mycotoxins generated by plants and no other possible forms of mycotoxins and their modifications, we hereby propose for all these forms a systematic definition consisting of four hierarchic levels. The highest level differentiates the free and unmodified forms of mycotoxins from those being matrix-associated and from those being modified in their chemical structure. The following lower levels further differentiate, in particular, "modified mycotoxins" into "biologically modified" and "chemically modified" with all variations of metabolites of the former and dividing the latter into "thermally formed" and "non-thermally formed" ones. To harmonize future scientific wording and subsequent legislation, we suggest that the term "modified mycotoxins" should be used in the future and the term "masked mycotoxins" to be kept for the fraction of biologically modified mycotoxins that were conjugated by plants.

The predictive value of different urinary and transcriptional biomarkers was evaluated in a proof-of-principle toxicology study in rats using aristolochic acid (AA), a known nephrotoxic agent. Male Wistar rats were orally dosed with 0.1, 1, or 10 mg/kg for 12 days. Urine was collected on days 1, 5, and 12 over 24 hours. Gene expression analysis was also conducted using quantitative real-time polymerase chain reaction and Illumina whole-genome chips. Protein biomarkers (Kim-1, Timp-1, vascular endothelial growth factor, osteopontin, clusterin, cystatin C, calbindin D-28K, β2-microglobulin, α-glutathione S-transferase, GSTY1b, RPA-1, and neutrophil gelatinase-associated lipocalin) were measured in these urine samples. Treatment with AA resulted in a slight dose- and/or time-dependent increase in urinary β2-microglobulin, lipocalin 2, and osteopontin before an increase in serum creatinine or serum urea nitrogen was observed. A strong decrease in urinary calbindin D-28K was also detected. The Compugen Ltd. prediction model scored both the 1- and 10-mg/kg AA dose groups as positive for nephrotoxicity despite the absence of renal histopathological changes. In addition, several previously described transcriptional biomarkers were identified as early predictors of renal toxicity as they were detected before morphological alterations had occurred. Altogether, these findings demonstrated the predictive values of renal biomarkers approved by the Food and Drug Administration, European Medicines Agency, and Pharmaceuticals & Medical Devices Agency in AA-induced renal injury in rats and confirmed the utility of renal transcriptional biomarkers for detecting progression of compound-induced renal injury in rats. In addition, several transcriptional biomarkers identified in this exploratory study could present early predictors of renal tubular epithelium injury in rats.

Although a basic understanding of the chemical and biological events leading to idiosyncratic drug toxicity is still lacking, it appears that drug-independent risk factors that increase reactive metabolite formation or alter cellular stress and immune response may be critical determinants in the response to an otherwise non-toxic drug. Thus, we were interested to determine the impact of various drug-independent stress factors - lipopolysaccharide (LPS), poly I:C (PIC) or glutathione depletion via buthionine sulfoximine (BSO) - on the toxicity of diclofenac (Dcl), a model drug associated with rare but significant cases of serious hepatotoxicity, and to understand if enhanced toxicity occurs through alterations of drug metabolism and/or modulation of stress response pathways. Co-treatment of rats repeatedly given therapeutic doses of Dcl for 7 days with a single dose of LPS 2h before the last Dcl dose resulted in severe liver toxicity. Neither LPS nor diclofenac alone or in combination with PIC or BSO had such an effect. While it is thought that bioactivation to reactive Dcl acyl glucuronides (AG) and subsequent protein adduct formation contribute to Dcl induced liver injury, LC-MS/MS analyses did not reveal increased formation of 4'- and 5-hydroxy-Dcl, Dcl-AG or Dcl-AG dependent protein adducts in animals treated with LPS/Dcl. Hepatic gene expression analysis suggested enhanced activation of NFκB and MAPK pathways and up-regulation of co-stimulatory molecules (IL-1β, TNF-α, CINC-1) by LPS/Dcl and PIC/Dcl, while protective factors (HSPs, SOD2) were down-regulated. LPS/Dcl led to extensive release of pro-inflammatory cytokines (IL-1β, IL-6, IFN-γ, TNF-α) and factors thought to constitute danger signals (HMGB1, CINC-1) into plasma. Taken together, our results show that Dcl enhanced the inflammatory response induced by LPS - and to a lesser extent by PIC - through up-regulation of pro-inflammatory molecules and down-regulation of protective factors. This suggests sensitization of cells to cellular stress mediated by non-drug-related risk factors by therapeutic doses of Dcl, rather than potentiation of Dcl toxicity by the stress factors., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

The mycotoxin fumonisin B1 (FB1) is an important contaminant of maize and maize-based products. In rodent toxicity studies, FB1 was shown to be hepato- and nephrotoxic, and to induce renal tumors in rats when administered via the diet. Of particular note are the aggressive growth characteristics of FB1-induced tumors with a high potential to metastasize. While genotoxicity does not appear to contribute to FB1 carcinogenicity, it is well established that FB1-mediated disruption of sphingolipid metabolism plays a key role in FB1 toxicity. This review provides an overview on human dietary exposure to FB1, FB1 toxicity and carcinogenicity, and potential mechanisms involved in FB1-mediated tumor formation, with a particular focus on cellular functions of sphingolipids and biological consequences of FB1-mediated perturbation of sphingolipid metabolism., (Copyright © 2012 Elsevier Ltd. All rights reserved.)

Scope: Furan is a potent hepatotoxicant and liver carcinogen in rodents. However, short-term tests for genotoxicity of furan are inconclusive. The aim of this study was to assess the potential of furan to covalently bind to DNA, and to assess furan genotoxicity in rats in vivo., Materials and Methods: Accelerator mass spectrometry was used to determine the (14) C-content in DNA following administration of [3,4-(14) C]-furan (0.1 and 2.0 mg/kg bw) to F344 rats. DNA damage, micronuclei, chromosomal aberrations, and sister chromatid exchanges were analyzed in F344 rats treated with furan for up to 28 days., Conclusion: The (14) C-content in liver DNA was significantly increased in a dose-dependent manner, with mean concentrations of 7.9 ± 3.5 amol (14) C/μg DNA and 153.3 ± 100.2 amol (14) C/μg DNA, corresponding to 16.5 ± 7.4 and 325.2 ± 212.7 adducts/10(9) nucleotides at 0.1 and 2.0 mg/kg bw, respectively. There was no evidence for genotoxicity of furan in peripheral blood and bone marrow cells. However, a dose-related increase in the incidence of chromosomal aberrations in rat splenocytes and some indication of DNA damage in liver were observed. Collectively, results from this study indicate that furan may operate-at least in part-by a genotoxic mode of action., (© 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)