Comparative in silico and in vitro evaluation of possible toxic effects of bisphenol derivatives in HepG2 cells.

Introduction: Bisphenols are widely used in the production of polycarbonate plastics and resin coatings. Bisphenol A (BPA) is suggested to cause a wide range of unwanted effects and "low dose toxicity". With the search for alternative substances to BPA, the use of other bisphenol derivatives namely bisphenol F (BPF) and bisphenol S (BPS) has increased. Methods: In the current study, we aimed to evaluate the in silico predicted inhibitory concentration 50s (pIC50s) of bisphenol derivatives on immune and apoptotic markers and DNA damage on HepG2 cells. Moreover, apoptotic, genotoxic and immunotoxic effects of BPA, BPF and BPS were determined comparatively. Effects of bisphenols on apoptosis were evaluated by detecting different caspase activities. The genotoxic effects of bisphenols were evaluated by measuring the levels of 8-hydroxy-2'-deoxyguanosine (8-OHdG) and 8-oxoguanine glycosylase (OGG1). To determine the immunotoxic effect of bisphenol derivatives, the levels of interleukin 4 (IL-4) and interleukin 10 (IL-10), transforming growth factor beta (TGF-ß) and tumor necrosis factor-alpha (TNF-α), which are known to be expressed by HepG2 cells, were measured. Results: In silico data indicate that all of the bisphenols may cause alterations in immune and apoptotic markers as well as DNA damage at low doses. In vitro data revealed that all bisphenol derivatives could affect immune markers at inhibitory concentration 30s (IC30s). In addition, BPF and BPS may also have apoptotic immunotoxic effects. Conclusion: Both in silico and in vivo research are needed further to examine the toxic effects of alternative bisphenol derivatives.

Biomonitoring of Oxidative-Stress-Related Genotoxic Damage in Patients with End-Stage Renal Disease.

Chronic kidney disease (CKD), a common progressive renal failure characterized by the permanent loss of functional nephrons can rapidly progress to end-stage renal disease, which is known to be an irreversible renal failure. In the therapy of ESRD, there are controversial suggestions about the use of regular dialysis, since it is claimed to increase oxidative stress, which may increase mortality in patients. In ESRD, oxidative-stress-related DNA damage is expected to occur, along with increased inflammation. Many factors, including heavy metals, have been suggested to exacerbate the damage in kidneys; therefore, it is important to reveal the relationship between these factors in ESRD patients. There are very few studies showing the role of oxidative-stress-related genotoxic events in the progression of ESRD patients. Within the scope of this study, genotoxic damage was evaluated using the comet assay and 8-OHdG measurement in patients with ESRD who were undergoing hemodialysis. The biochemical changes, the levels of heavy metals (aluminum, arsenic, cadmium, lead, and mercury) in the blood, and the oxidative biomarkers, including superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and malondialdehyde (MDA) levels were evaluated, and their relationship with genotoxic damages was revealed. Genotoxicity, oxidative stress, and heavy-metal levels, except mercury, increased significantly in all renal patients. DNA damage, 8OHdG, and MDA significantly increased, and GSH significantly decreased in patients undergoing dialysis, compared with those not having dialysis. The duration and the severity of disease was positively correlated with increased aluminum levels and moderate positively correlated with increased DNA damage and cadmium levels. In conclusion, this study revealed that the oxidative-stress-related DNA damage, and also the levels of Al and Cd, increased in ESRD patients. It is assumed that these changes may play an important role in the progression of renal damage. Approaches for reducing oxidative-stress-related DNA damage and heavy-metal load in ESRD patients are recommended.

Evaluation of genotoxicity, oxidative stress and immune parameters of auto-paint workers.

The automotive industry is a very wide area from the manufacturing of the pieces of the engine, the body, plastics to the assembly of the car. There is a chemical risk at different stages of production because of the requirement of the use of many corrosive and irritant chemicals such as paints, adhesives, acids, and bases. The aim of the study was to determine the genotoxicity, oxidative stress and immune parameters of automotive paint workers in Ankara, Türkiye. DNA damage of workers mainly responsible from the painting of the automotives were evaluated using the alkaline comet assay and the levels of some oxidative stress and immune biomarkers were also investigated. Increased lymphocyte DNA damage and also higher 8-hydroxy-2'-deoxyguanosine (8-OHdG) and malondialdehyde (MDA) levels were observed while decreased glutathione (GSH), glutathione peroxidase (GPx), and glutathione reductase (GR) levels were found in the workers compared to their controls There were no significant differences between the study groups in the levels of interleukin (IL)- 1beta, IL-17, IL-23, Clara cell secretory protein (CC16), tumor necrosis factor-alpha (TNF-alpha), catalase (CAT), and superoxide dismutase (SOD). The results show that occupational exposure to chemicals in automotive industry may cause DNA damage in workers due to oxidative stress.

What can we learn from epidemiological studies on chronic boron exposure?

Boric acid and sodium borates are classified as toxic to reproduction and development under category 1B, with the hazard statement of H360FD (May damage fertility. May damage the unborn child) in the EU-CLP regulation. This classification triggered environmental and occupational epidemiology studies on boron in China, Türkiye, and Argentina. The highest mean total daily boron exposure (DBE) levels in mining areas or processing plants in China and Türkiye were 41.2 and 47.17 mg/day, respectively. These DBE levels are higher than the proposed reference doses (RfDs) for the developmental (9.6 mg B/day) and reproductive effects (20.3 mg B/day) of boron in females and males. Despite these high exposure levels in occupational settings, boron-mediated reproductive and developmental effects have not been reported. Blood (or serum) boron concentrations were also reported in these studies. The highest mean blood-boron concentration reported for the boron-exposed male workers in Türkiye is 570.6 ng B/g (ppb). This blood-boron concentration is still much lower than the blood boron concentrations corresponding to the no observed adverse effect level (NOAEL) for both developmental and reproductive effects in rats. Given the blood boron concentrations reported in current epidemiological studies, the lack of reproductive and developmental effects in humans is not surprising. Recent epidemiological studies have proven that it will not be possible to reach the critical blood boron concentrations in humans for the reproductive and developmental effects, especially in daily life.

Effects of COVID-19 Disease on DNA Damage, Oxidative Stress and Immune Responses.

Coronavirus disease 2019 (COVID-19) has posed a great threat to public health and has caused concern due to its fatal consequences over the last few years. Most people with COVID-19 show mild-to-moderate symptoms and recover without the need for special treatment, while others become seriously ill and need medical attention. Additionally, some serious outcomes, such as heart attacks and even stroke, have been later reported in patients who had recovered. There are limited studies on how SARS-CoV-2 infection affects some molecular pathways, including oxidative stress and DNA damage. In this study, we aimed to evaluate DNA damage, using the alkaline comet assay, and its relationship with oxidative stress and immune response parameters in COVID-19-positive patients. Our results show that DNA damage, oxidative stress parameters and cytokine levels significantly increased in SARS-CoV-2-positive patients when compared with healthy controls. The effects of SARS-CoV-2 infection on DNA damage, oxidative stress and immune responses may be crucial in the pathophysiology of the disease. It is suggested that the illumination of these pathways will contribute to the development of clinical treatments and to reduce adverse effects in the future.

Comparative evaluation of the effects of bisphenol derivatives on oxidative stress parameters in HepG2 cells.

Bisphenol A (BPA) BPA is an endocrine-disrupting chemical that has a wide range of uses. Exposure to BPA can be by oral, inhalation, and parenteral routes. Although its use in several products is limited, there is still concern on its adverse health effects, particularly for susceptible populations like children. Alternative bisphenols, such as bisphenol S (BPS) and bisphenol F (BPF), are now being used instead of BPA, although there is little information on the toxicity of these bisphenols. BPF is used as a plasticizer in the production of several industrial materials as well as in the coating of drinks and food cans. BPS is used in curing fast-drying epoxy glues, as a corrosion inhibitor and as a reactant in polymer reactions. In this study, the possible toxic effects of BPA, BPS, and BPF in HepG2 cells were evaluated comparatively. For this purpose, their effects on cytotoxicity, production of intracellular reactive oxygen species (ROS), oxidant/antioxidant parameters, and DNA damage have been examined. The cytotoxicity potentials of different bisphenols were found to be as BPS > BPF > BPA. All bisphenol derivatives caused increases in intracellular ROS production. We observed that all bisphenol derivatives cause an imbalance in some oxidant/antioxidant parameters. Bisphenols also caused significant DNA damage in order of BPF > BPA > BPS. We can suggest that both of the bisphenol derivatives used as alternatives to BPA also showed similar toxicities and may not be considered as safe alternatives. Mechanistic studies are needed to elucidate this issue.

Evaluation of Oxidative Stress Parameters and Genotoxic Effects in Patients With Work-Related Asthma and Silicosis.

OBJECTIVE: To determine the mechanistic roles of oxidative stress, inflammation, and genotoxicity parameters in patients with work-related asthma (WRA) and silicosis. METHODS: Thirty-eight healthy office workers, 27 employees with a history of exposure and no disease, 24 employees with WRA, and 23 employees with silicosis were included in this study. Superoxide dismutase, catalase, glutathione peroxidase, malondialdehyde, and interleukins (IL) 17, 23, and 27 levels were measured in the serum. Genotoxic damage was evaluated by calculating the frequency of micronuclei in swab samples and 8-hydroxy-2'-deoxyguanosine in serum. RESULTS: Serum superoxide dismutase, catalase, glutathione peroxidase, malondialdehyde, 8-hydroxy-2'-deoxyguanosine, and IL-17, IL-23, and IL-27 levels were found to be statistically significantly higher in the exposure, WRA, and silicosis groups compared with the control group. The frequency of micronuclei in buccal epithelial cells of the patient group was found to be significantly higher than that of the control group. CONCLUSION: These results may provide information for molecular mechanisms and early diagnosis of WRA and silicosis and will be a guide for taking precautions in the early period.

Unpredictable adverse effects of herbal products.

Herbal products are being increasingly used all over the world for preventive and therapeutic purposes because of the belief of their safety. They have become an important part of health care system in many countries since they can easily be purchased in the health food stores or online. However, the lack of sufficient study on their efficacy and toxicity, inadequate controls of their availability, reduce their safety. Unlike conventional drugs, herbal products are not regulated for purity and potency. Herbal products contain substances which can induce or inhibit enzymes that take part in drug metabolism. Therefore the concurrent use of drugs with some medicinal plants can cause serious adverse effects and can also decrease the efficacy of the therapy. Particularly, drugs with narrow therapeutic index and plants which can affect drug metabolizing enzymes when used together, may lead to unpredictable adverse reactions. Impurities, contaminants and adulterants found in the herbal products, are the most common malpractises in herbal raw-material trade. In this review the unpredictable adverse effects of herbal products due to their possible interactions with drugs and also due to the adulteration and contamination with prohibited chemicals will be discussed in detail.

DNA damage in circulating leukocytes measured with the comet assay may predict the risk of death.

The comet assay or single cell gel electrophoresis, is the most common method used to measure strand breaks and a variety of other DNA lesions in human populations. To estimate the risk of overall mortality, mortality by cause, and cancer incidence associated to DNA damage, a cohort of 2,403 healthy individuals (25,978 person-years) screened in 16 laboratories using the comet assay between 1996 and 2016 was followed-up. Kaplan-Meier analysis indicated a worse overall survival in the medium and high tertile of DNA damage (p < 0.001). The effect of DNA damage on survival was modelled according to Cox proportional hazard regression model. The adjusted hazard ratio (HR) was 1.42 (1.06-1.90) for overall mortality, and 1.94 (1.04-3.59) for diseases of the circulatory system in subjects with the highest tertile of DNA damage. The findings of this study provide epidemiological evidence encouraging the implementation of the comet assay in preventive strategies for non-communicable diseases.

The hCOMET project: International database comparison of results with the comet assay in human biomonitoring. Baseline frequency of DNA damage and effect of main confounders.

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.

Does storage conditions of whole blood or blood cells effect genotoxicity assessment by comet assay?

Comet assay, applied to in vitro, in vivo and ex vivo systems, is a quick, simple, and sensitive method for the detection of genotoxicity. In general, fresh whole blood or peripheral blood mononuclear cells (PBMCs) are used in the assay for the determination of DNA damage and repair. In this study, the effects of storage conditions on genotoxicity assessed by Comet assay in human whole blood and lymphocyte samples, were evaluated. Whole blood and lymphocyte samples were stored at 4 °C for 1, 2, 3, 4, 5 and 7 days; at -20 °C for 1 month and at -80 °C for 3, 6 and 12 months. 1% DMSO was used as cryoprotectant. No significant differences in DNA damage were demonstrated in all of the storage conditions and durations, and the results were similar according to the median values (p < 0.05). According to Spearman or Pearson correlations, an important correlation was found between the DNA damage of the fresh samples and the samples which were kept at -80 °C for 6 months with temperature and time (p < 0.01 for Pearson and p < 0.05 for Spearman). The results of this study indicated that blood and lymphocyte samples stored in +4 °C, -20 °C and -80 °C up to 12 months can be used instead of fresh samples especially in human biomonitoring studies.

Minimum Information for Reporting on the Comet Assay (MIRCA): recommendations for describing comet assay procedures and results.

The comet assay is a widely used test for the detection of DNA damage and repair activity. However, there are interlaboratory differences in reported levels of baseline and induced damage in the same experimental systems. These differences may be attributed to protocol differences, although it is difficult to identify the relevant conditions because detailed comet assay procedures are not always published. Here, we present a Consensus Statement for the Minimum Information for Reporting Comet Assay (MIRCA) providing recommendations for describing comet assay conditions and results. These recommendations differentiate between 'desirable' and 'essential' information: 'essential' information refers to the precise details that are necessary to assess the quality of the experimental work, whereas 'desirable' information relates to technical issues that might be encountered when repeating the experiments. Adherence to MIRCA recommendations should ensure that comet assay results can be easily interpreted and independently verified by other researchers.

Evaluation of the Possible Role of miRNAs in Chemical Allergen Potency.

OBJECTIVES: MicroRNAs (miRNAs) are short, endogenous noncoding RNA molecules that can bind to certain parts of target mRNAs, thereby regulating gene expression. Studies showed that miRNAs could be up- or downregulated in different allergic skin conditions but there is still need for further studies. The aim of this study was to investigate the expression of miRNAs in response to the common contact allergen Bandrowski's base (BB), the principal allergen in patients reacting to p-phenylenediamine (PPD). MATERIALS AND METHODS: The human promyelocytic cell line THP-1 was exposed to BB at a concentration of 1 µg/mL for 24, 48, and 72 h. The dose was selected from the results of cytotoxicity assays. RNA was purified and miRNA expression profile and real-time polymerase chain reaction (RTPCR) were performed to identify up- or downregulated miRNAs and confirm their modulations. RESULTS: Among the different modulated miRNAs, the upregulation of miRNA-155 and the downregulation of miRNA-21 were found to be important because these are related to immune system. This expression profile of miRNAs was also confirmed by RT-PCR. CONCLUSION: These preliminary results showed that miR-155 and miR-21 may play a role in the pathogenesis of allergic contact dermatitis, but further studies are needed to clarify their definite roles.

Occupational Exposure to Metals and Solvents: Allergy and Airway Diseases.

PURPOSE OF REVIEW: Occupational allergic diseases (OAD) such as occupational contact dermatitis (OCD), occupational asthma (OA), and occupational rhinitis (OR) are the most prevalent occupational diseases in industrialized countries. The purpose of this review is to provide an update about the main occupational metal and solvent exposures related to allergy and airway diseases and to discuss newly defined causative agents and industries in this field. RECENT FINDINGS: Currently for over 400 causative agents for OA and OCD, several hundreds of agents for OR have been identified. Although many studies have reported an overall decline in OAD related to known agents after implementation of efficient and effective workplace preventive measures, the constant development of new products continuously introduces to the market potential unknown respiratory hazards. Workplace allergens are often high molecular weight (HMW) agents that are > 10 kDa molecular weight and capable of eliciting IgE sensitization. Sensitizing low molecular weight (LMW) agents are often reactive chemicals. Metals and solvents are two large causative agent groups related to OADs that mainly behave as LMW (< 10 kDa) sensitizers and/or irritants. Avoidance of causative exposures through control strategies is the primary prevention approach for OADs. These strategies must be applied and covered for all known and newly defined causative agents. This review aims to summarize current status of known occupational metal and solvent exposures related to allergy and airway diseases and to discuss newly defined causative agents and industries in this field.

Evaluation of oxidative stress and immune parameters of boron exposed males and females.

There is growing evidence that boron (B) and B compounds are essential nutrients for animals and humans. Besides, B compounds have been suggested to treat inflammation and oxidative stress. As a part of our "Boron Project II" on B-exposed persons in Bandirma and Bigadic (Turkey) between 2014 and 2017, anti-oxidant/pro-oxidant and inflammatory parameters were assessed. In this first large-scale human study biomarkers of oxidative stress such as the enzyme activities of superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), glutathione reductase (GR) and the levels of malondialdehyde (MDA), glutathione (GSH) and 8-hydroxy-2'-deoxy-guanosine (8-OH-dG) were investigated, in relation to B exposure. The immune biomarkers interleukin (IL)-1ra, IL-6, IL-8 and nuclear factor kappa B (NF-κB) levels were included. There was no influence of human exposure to B on the parameters of oxidative stress and inflammation.

Oxidative stress status of Turkish welders.

Welding, a fabrication process that joins metals or thermoplastics by causing coalescence, is indispensable in modern society and ubiquitous in industry. Welding generates fumes that contain several metals and gases that comprise fine and ultrafine particles with the potential for adverse effects. Although health risks of welders have been evaluated in different populations, occupational exposure to welding fumes is still considered to be an important health problem, especially in developing countries. The aim of this study was to investigate the effects of welding fume exposure on important oxidative stress parameters such as superoxide dismutase (SOD), catalase (CAT), glutathione reductase (GR), total glutathione (GSH), glutathione peroxidase (GPx), malondialdehyde (MDA), and 8-hydroxy-2'-deoxyguanosine (8-OHdG) in Turkish welders (n = 48). The influence of confounding factors such as age, smoking habits, alcohol consumption, and duration of exposure on the studied parameters was also analyzed. In our study, significant decreases in the levels of GSH and activities of CAT, SOD, and GPx and significant increases of MDA, 8-OHdG levels and GR activity were found in the workers compared to the controls. There was a negative correlation between GSH levels and alcohol usage. Also, older workers (≥35 years) had significantly higher GR levels than younger workers. But smoking and alcohol usage, duration of exposure, and utilization of protective measures had no significant effect on the studied parameters in the workers. These results indicate that occupational exposure to welding fumes appears to induce oxidative stress and oxidative DNA damage.

Effects of boron compounds on human reproduction.

Because of the high pKa of boric acid, inorganic borates, when taken up by the human or animal organism, exist in the body almost exclusively in the form of non-dissociated boric acid. Therefore, the variety of inorganic boron compounds is commonly addressed in the toxicological literature as "boron" (B). There is a discussion concerning categorisation of inorganic boron compounds as reproductive toxins. Boron treatment of rats, mice and dogs was dose-dependently associated with testicular toxicity, characterised by inhibited spermiation at lower dose levels and by reduction of epididymal sperm counts at higher dose levels. The NOAEL for such fertility effects of boric acid in male rats (oral feeding, Sprague Dawley strain) was evaluated to be 17.5 mg B/kg bw per day. As far as developmental toxicity is concerned, oral dosing of 9.6 mg B/kg bw daily to female pregnant Sprague Dawley rats remained without effects, with foetal skeletal effects observed at higher doses. Therefore, 9.6 mg B/kg bw (oral dosing) was evaluated as NOAEL for developmental effects of boric acid. The blood level in rats, equivalent to this NOAEL, is 1270 ng B/g. As far as B-exposed humans are concerned, field studies on the effect of boron on human reproduction are possible only in a few boron-rich geographical areas. Published field studies were conducted in China's Liaoning province, the Argentinian Andes and Western Anatolia/Turkey. Particularly relevant are studies on occupationally B-exposed groups, because the potential exposure to boron is much higher in occupational compared to environmental settings. Comparison of estimated daily B exposure levels in humans and actually measured B blood levels confirms the preference of biomonitoring for exposure assessment in environmental and occupational studies. A boron blood level scaling shows that the levels of high occupational B exposures reported in China and in Turkey are compatible. Compared to the experimental B blood levels at boron-related NOAELs for male fertility and for developmental toxicity in rats, the human blood level means of the highest occupational exposure groups in China and in Turkey are lower by factors of > 4 and > 2, respectively. Basically, concentrations of B within the body that exert reproductive toxicity in humans are not reached under the conditions of human normal handling and use, including conditions of extreme occupational exposures. In consequence, all relevant results of studies into human reproductive toxicity of B are basically negative. Considering the effective doses, there is no scientific contradiction between experimental and human results of B reproductive toxicity.

Application of the comet assay in human biomonitoring: An hCOMET perspective.

The comet assay is a well-accepted biomonitoring tool to examine the effect of dietary, lifestyle, environmental and occupational exposure on levels of DNA damage in human cells. With such a wide range of determinants for DNA damage levels, it becomes challenging to deal with confounding and certain factors are inter-related (e.g. poor nutritional intake may correlate with smoking status). This review describes the effect of intrinsic (i.e. sex, age, tobacco smoking, occupational exposure and obesity) and extrinsic (season, environmental exposures, diet, physical activity and alcohol consumption) factors on the level of DNA damage measured by the standard or enzyme-modified comet assay. Although each factor influences at least one comet assay endpoint, the collective evidence does not indicate single factors have a large impact. Thus, controlling for confounding may be necessary in a biomonitoring study, but none of the factors is strong enough to be regarded a priori as a confounder. Controlling for confounding in the comet assay requires a case-by-case approach. Inter-laboratory variation in levels of DNA damage and to some extent also reproducibility in biomonitoring studies are issues that have haunted the users of the comet assay for years. Procedures to collect specimens, and their storage, are not standardized. Likewise, statistical issues related to both sample-size calculation (before sampling of specimens) and statistical analysis of the results vary between studies. This review gives guidance to statistical analysis of the typically complex exposure, co-variate, and effect relationships in human biomonitoring studies.

Assessment of DNA damage in welders using comet and micronucleus assays.

Welding technology is widely used in pressurized containers, thermal power plants, refineries, chemical facilities and steel structures. Welders are exposed to a number of hazardous compounds such as ultraviolet (UV) radiation, electromagnetic fields, toxic metals and polycyclic aromatic hydrocarbons (PAHs). In the present study, 48 welders and an equal number of control subjects were evaluated for DNA damage in the whole blood and isolated lymphocytes using the comet assay. The genotoxic damage in buccal epithelial cells of subjects was determined by micronucleus (MN) assay. Metal(loids) such as Cr, Mn, Ni, Cu, As, Cd and Pb levels in blood samples were evaluated by using Inductively Coupled Plasma-Mass Spectrometer (ICP-MS). Results of this study showed that DNA damage in blood, isolated lymphocytes, and buccal epithelial cells were significantly higher in workers compared to the controls. Also, these workers had remarkably higher blood Cr, Cu, Cd, Ni and Pb levels. These results showed that occupational exposure to welding fumes may cause genotoxic damage that can lead to important health problems in the workers. More extensive epidemiological studies should be performed that enable the assessment of health risk in welding industry.