Increase In Micronucleus Frequency Research Articles

Frequencies of micronuclei in buccal cells and their spatial distribution in a population living in proximity to coal mining areas in southern Brazil

The extraction and burning of coal release genotoxic pollutants, and understanding the relationship between genetic damage and the spatial distribution of residences in coal-using regions is crucial. The study aimed to conduct a spatial analysis of genotoxic damage through the of micronuclei (MNs) number and their proximity to coal mining/burning in the largest coal exploration region in Brazil. In this study, the detection of genotoxic damage was performed using the MN assay in oral cells of residents exposed to coal mining activities. Spatial analysis was conducted using QGIS 3.28.10 based on information obtained from a questionnaire administered to the population. Multiple linear regression analysis was carried out to assess the influence of the distance from residential areas to polluting sources on the number of MNs found. Additionally, Spearman's correlation was performed to identify the strength and direction of the association between the frequency of MNs and each of the polluting sources. A total of 147 MNs were quantified among all participants in the coal mining region. Notably, residents living within 2 km and 10 km of pollution sources exhibited the highest prevalence of MNs. The analysis demonstrated a significant correlation between closer proximity to pollution sources and increased MN frequency, underscoring the spatial relationship between these sources and genotoxic damage. Environmental pollutants from anthropogenic sources present a major health risk, potentially leading to irreversible damage. The spatial analysis in this study highlights the importance of targeted public policies. These policies should aim for a sustainable balance between economic development and public health, promoting effective measures to mitigate environmental impacts and protect community health.

Cellular and Genomic Instability Induced by the Herbicide Glufosinate-Ammonium: An In Vitro and In Vivo Approach.

Glufosinate-ammonium (GLA), an organophosphate herbicide, is released at high concentrations in the environment, leading to concerns over its potential genotoxic effects. However, few articles are available in the literature reporting the possible cellular and nuclear effects of this compound. We assessed, by in vitro and in vivo micronucleus assays, the genotoxicity of GLA on cultured human lymphocytes and Lymnaea stagnalis hemocytes at six concentrations: 0.010 (the established acceptable daily intake value), 0.020, 0.050, 0.100, 0.200, and 0.500 µg/mL. In human lymphocytes, our results reveal a significant and concentration-dependent increase in micronuclei frequency at concentrations from 0.100 to 0.500 μg/mL, while in L. stagnalis hemocytes, significant differences were found at 0.200 and 0.500 μg/mL. A significant reduction in the proliferation index was observed at all tested concentrations, with the only exception of 0.010 μg/mL, indicating that the exposure to GLA could lead to increased cytotoxic effects. In L. stagnalis, a significant reduction in laid eggs and body growth was also observed at all concentrations. In conclusion, we provided evidence of the genomic and cellular damage induced by GLA on both cultured human lymphocytes and a model organism's hemocytes; in addition, we also demonstrated its effects on cell proliferation and reproductive health in L. stagnalis.

CDK4/6 inhibitor-induced bone marrow micronuclei might be caused by cell cycle arrest during erythropoiesis

BackgroundA micronucleus test is generally used to evaluate the genotoxic potential of chemicals. Exaggerated erythropoiesis, as occurs following bleeding, may induce an unexpected increase in micronucleus frequency. This false positive result would be typical in a genotoxicity study due to the enhanced progression of the cell cycle that restores decreased blood cells. The cyclin-dependent kinase (CDK) family is known to play an essential role in preventing genomic instability. Conversely, a selective CDK4/6 inhibitor PD0332991, clinically named Palbociclib, is reported to have genotoxic potential, shown by positive results in both in vitro and in vivo micronucleus studies. To clarify the mechanism by which cell cycle arrest induced by a CDK4/6 inhibitor increases micronucleus frequency, we investigated the positive results of the bone marrow micronucleus test conducted with PD0332991.ResultsRats treated with PD0332991 exhibited increased micronucleus frequency in an in vivo bone marrow micronucleus test whereas it was not increased by treatment in human lymphoblastoid TK6 cells. In addition, all other genotoxicity tests including the Ames test and the comet assay showed negative results with PD0332991. Interestingly, PD0332991 treatment led to an increase in erythrocyte size in rats and affected the size distribution of erythrocytes, including the micronucleus. The mean corpuscular volume of reticulocytes (MCVr) in the PD0332991 treatment group was significantly increased compared to that of the vehicle control (83.8 fL in the PD0332991, and 71.6 fL in the vehicle control.). Further, the average micronucleated erythrocytes (MNE) size of the PD0332991 group and vehicle control was 8.2 and 7.3 µm, respectively. In the histogram, the vehicle control showed a monomodal distribution with a peak near 7.3 µm. In contrast, the PD0332991 group showed a bimodal distribution with peaks around 7.5 and 8.5 µm. Micronucleated erythrocytes in the PD0332991 group were significantly larger than those in the vehicle control.These results suggest that the increase in micronucleus frequency induced by the CDK4/6 inhibitor is not due to genotoxicity, but is attributable to disturbance of the cell cycle, differentiation, and enucleation of erythroblasts.ConclusionsIt was suggested that the positive outcome of the in vivo bone marrow micronucleus test resulting from treatment with PD0332991 could not be attributed to its genotoxicity. Further studies to clarify the mechanism of action can contribute to the development of drug candidate compounds lacking intrinsic genotoxic effects.

Detection of early-stage gastrointestinal cancers by micronuclei DNA from erythrocytes.

382 Background: Gastrointestinal (GI) cancers account for over a third of cancer deaths. However, their early detection remains a challenge. Micronuclei (MN) are extranuclear bodies containing damaged chromosome segments indicative of genomic instability. Increased MN frequency in hematopoietic cells is observed in patients with solid tumors. We have developed a method (WO2021/228246 A1) for purifying and characterizing micronuclei DNA (mnDNA) in erythrocytes from peripheral blood. Here, we evaluated the potential of mnDNA for early-stage GI cancer detection, including colorectal (CC) and gastric (GC) cancers. Methods: Peripheral blood (1 ml) was collected from healthy donors (HD) and cancer patients mnDNA isolation and purification from erythrocytes. Participants were randomly divided into training, validation and independent test cohorts in a 7:2:1 ratio, maintaining similar cancer types, CC/GC/HD ratio, gender and age distribution. Distinct mnDNA features between cancer patients and HDs were identified using sequencing data. Logistic regression algorithms were applied using these features for cancer detection. Results: The study enrolled 1046 participants, including 419 HDs, 366 CC and 261 GC cases. Of these, over half were diagnosed with early-stage diseases; TNM stage I accounted for 38.7%, stage II for 22.8%, stage III for 30.5% and stage IV for 8.0%. The cancer detection model with mnDNA features yielded an overall accuracy of 84.8%, with a sensitivity of 84.1% and specificity of 85.7%. For individual caner types, sensitivity was 89.2% for CC and 76.9% for GC. For early-stage (stage I-II) CC and GC, the model demonstrated sensitivities of 88.9% and 76.5%, respectively, at 85.7% specificity. Conclusions: Unlike cell-free DNA, mnDNA are chromosomal fragments in the cytoplasm. The abundance of erythrocytes in peripheral blood provides an easily accessible source for mnDNA enrichment. This pilot study illustrates the potential of mnDNA as a tool for early GI cancer detection, contributing to large-scale efforts towards developing an effective GI cancer screening test.

Vicia faba Plant Suitability Assessment for Genotoxicity, Cytotoxicity, and Mutagenicity Testing of Pharmaceutical-Containing Wastewater

The article aimed to assess the Vicia faba plant’s suitability in the micronucleus test for determining toxicity of wastewater containing diclofenac and sulfamethoxazole. Additionally, the study evaluated the activity of the antioxidant enzymes catalase and superoxide dismutase in plant leaves. The assessment of wastewater was performed on laboratory-constructed wetland models. Both influent and effluent samples were tested, and the study examined two methods of plant root exposure: hydroponic culture and soil culture. The analysis showed a decrease in the mitotic index (57% inhibition on average in hydroponic and 42% in soil culture for influent and 39% and 19%, respectively, for treated wastewater), indicating the toxicity of the wastewater. The inhibition of the cell division frequency was lower in soil culture, and the frequency of aberrations of chromosomes was also lower. However, there was no increase in micronuclei frequency. An upsurge in catalase activity was observed upon analyzing the wastewater, with a 67% increase in the influent and a 20% increase in the treated wastewater. Additionally, there was a notable boost in superoxide dismutase activity, primarily in hydroponic culture with raw wastewater, averaging 186%. The results showed genotoxic and cytotoxic effects, but there were no mutagenic effects. The Vicia faba assay is advantageous for its simplicity and rapid results; it offers representative assessment of genotoxicity through its broad range of detected effects.

Genotoxicity assessment of eight nitrosamines using 2D and 3D HepaRG cell models.

N-nitrosamine impurities have been increasingly detected in human drugs. This is a safety concern as many nitrosamines are mutagenic in bacteria and carcinogenic in rodent models. Typically, the mutagenic and carcinogenic activity of nitrosamines requires metabolic activation by cytochromes P450 enzymes (CYPs), which in many in vitro models are supplied exogenously using rodent liver homogenates. There are only limited data on the genotoxicity of nitrosamines in human cell systems. In this study, we used metabolically competent human HepaRG cells, whose metabolic capability is comparable to that of primary human hepatocytes, to evaluate the genotoxicity of eight nitrosamines [N-cyclopentyl-4-nitrosopiperazine (CPNP), N-nitrosodibutylamine (NDBA), N-nitrosodiethylamine (NDEA), N-nitrosodimethylamine (NDMA), N-nitrosodiisopropylamine (NDIPA), N-nitrosoethylisopropylamine (NEIPA), N-nitroso-N-methyl-4-aminobutyric acid (NMBA), and N-nitrosomethylphenylamine (NMPA)]. Under the conditions we used to culture HepaRG cells, three-dimensional (3D) spheroids possessed higher levels of CYP activity compared to 2D monolayer cells; thus the genotoxicity of the eight nitrosamines was investigated using 3D HepaRG spheroids in addition to more conventional 2D cultures. Genotoxicity was assessed as DNA damage using the high-throughput CometChip assay and as aneugenicity/clastogenicity in the flow-cytometry-based micronucleus (MN) assay. Following a 24-h treatment, all the nitrosamines induced DNA damage in 3D spheroids, while only three nitrosamines, NDBA, NDEA, and NDMA, produced positive responses in 2D HepaRG cells. In addition, these three nitrosamines also caused significant increases in MN frequency in both 2D and 3D HepaRG models, while NMBA and NMPA were positive only in the 3D HepaRG MN assay. Overall, our results indicate that HepaRG spheroids may provide a sensitive, human-based cell system for evaluating the genotoxicity of nitrosamines.

36 An Investigation into Mechanisms Responsible for (geno)Toxicity Induced by Exposure to Industrially-Relevant Multi-Component Nanomaterials and high Aspect Ratio Nanomaterials

Abstract Production and application of multi-component nanomaterials (MCNMs) and high aspect ratio nanomaterials (HARN) has raised questions as to their inhalation risk. A PRISMA-orientated systematic literature review indicated a key knowledge gap related to the (geno)toxicity of metal oxide MCNMs and HARN. The aim of this study was to conduct a mechanistic-based toxicology assessment of a number of industrially-relevant MCNMs and HARN materials, in order to fill this knowledge gap and provide vital hazard assessment towards these materials’ safe and sustainable design. A549 monocultures (2.5x105), and A549/dTHP-1 co-cultures (1:10) were seeded in 12-well transwell-membranes at the air-liquid interface. Cell cultures were exposed, via aerosol, to Printex90, LaCo0.5Ni0.05O3, LaCo0.475Ni0.475Pt0.05O3, LaCo0.475Ni0.475Pd0.05O3, LaCo0.25Ni0.75O3, LaNiO3, NiZnFe4O8, ZnFe2O4, and NiFe2O4 to human exposure-relevant concentrations of 390, 780, 3100 ng/cm2 (24hrs). Aerosolised MCNM and HARN were characterised following acellular deposition via electron microscopy. Cellular cytotoxicity, barrier integrity, (pro)inflammatory/fibrotic response, oxidative stress and mutagenicity/DNA damage was assessed. Results indicate barrier integrity and IL-8 production were unaffected by Printex90, LaCo0.5Ni0.05O3, and ZnFe2O4 exposure. A dose-dependent increase in micronucleus frequency was observed for these four materials. Investigations are continuing to determine the mechanisms associated with this noted genotoxic effect, as well as upon further MCNMs/HARN (i.e. Fe2CoO4(Fe1/Co3), Silica-Ref-Std). Emphasis is being placed upon elucidating specific structure-activity relationships for each MCNM/HARN by testing specific key events of inhalation-based adverse outcome pathways relevant to the in vitro models used. This project was funded by EU H2020 research and innovation programme, grant agreement No. 953183 (HARMLESS).

Heavy metal accumulation, biochemical and transcriptomic biomarkers in earthworms Eisenia andrei exposed to industrially contaminated soils from south-eastern Tunisia (Gabes Governorate)

Heavy metal pollution is causing harmful consequences on soil fertility, and earthworms are frequently employed as test organisms to evaluate the ecotoxicity of polluted soils. In this study, Eisenia andrei was exposed for 7 and 14 days to polymetallic contaminated soils collected from an industrial zone in the south-eastern Tunisia. Earthworm growth, heavy metal accumulation, genotoxicity, cytotoxicity, biochemical and transcriptional responses were determined. Results revealed a higher accumulation of heavy metals in earthworms after 14 than 7 days of soil exposure, a reduction in lysosomal membrane stability (LMS), besides an increase in micronuclei frequency (MN). Moreover, earthworm oxidative status was affected in terms of increases in malondialdehyde (MDA) and metallothionein (MTs) content, and enhancement of catalase (CAT) and glutathione-S-transferase (GST) activities. An inhibition of acetylcholinesterase (AChE) activity was also observed in treated earthworms, whereas transcriptional data demonstrated an up-regulation of cat, gst, mt, p21 and topoisomerase genes. Overall, these findings support the use of earthworms as suitable bioindicator species for pollution monitoring and assessment, advance our understanding of the interaction between heavy metals and earthworms, and provide valuable information about the harmful impact of biota exposure to naturally contaminated soils.

Genotoxic and cytotoxic effects of thiomersal and paracetamol combination

Background: Thiomersal is used as a preservative of some vaccines or as a trace from the pathogen inactivation process in vaccine production. Prophylactic use of paracetamol upon vaccination is still common, even though paracetamol decreases immune response on some vaccines. Considering the cytotoxic and genotoxic potential of thiomersal and paracetamol and possible interaction in vivo, in vitro study was performed. Methods: The genotoxic and cytotoxic effects of thiomersal and paracetamol combination were examined on human lymphocyte cultures by using two methods: analysis of chromosomal aberrations and cytokinesis-block micronucleus cytome assay. Blood samples of three healthy donors were analyzed with the following concentrations of tested substances: thiomersal 1 µg/ml and 0.5 µg/ml, paracetamol 20 µg/ml, thiomersal 0.5 µg/ml with paracetamol 20 µg/ml and thiomersal 1.0 µg/ml with paracetamol 20 µg/ml. Results: The analysis of structural chromosomal aberrations was significantly increased in all treated cultures. In cells treated with the combination of thiomersal 1 µg/ml and 20 µg/ml of paracetamol, the number of aberrations was significantly decreased. Cytokinesis-block micronucleus cytome assay analyses showed significantly increased micronucleus frequency in lymphocytes cultivated with thiomersal 1 µg/ml compared to lymphocytes cultures exposed to thiomersal 0.5 µg/ml. Conclusions: Induction of structural chromosome aberrations and micronucleus is shown as a sign of genotoxicity for the examined concentrations of thiomersal and paracetamol. The suppressing effect of paracetamol on thiomersal genotoxicity in lymphocytes culture treated with thiomersal was shown to be indicative of further examination of paracetamol use in the prevention of genotoxicity.

Cytotoxic and genotoxic effects of leaking chemicals from serum infusion sets: an in-vitro study.

Safety concerns about medical devices playing important role in health sciences and bioengineering research are rising day by day. Although there are specific standards regarding disposable medical materials, the information is updating with the toxicological studies. In this study, cytotoxic/genotoxic effects of chemicals leaking from serum infusion sets that have an important place in the clinic were investigated. Media containing leakage chemicals were prepared from equal samples taken from the plastic line sections of 13 different brands of serum infusion sets containing phthalates and the effects on the cultured cells were compared with the untreated control groups. To obtain leaking chemicals, extracting period was selected as 72h, a routine set-change time in the clinic as indicated in various publications. Neutral red uptake and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide tests were performed in L929 cells to determine cytotoxicity, and cytokinesis blocked micronucleus technique was performed in lymphocytes to determine genotoxicity. Cytotoxic and genotoxic damage levels were compared by evaluating cell-viability rates relative to control, micronucleus frequency, and nuclear division index values. The results showed that all sets caused a decrease in cell viability revealing the effects both on lysosomal and mitochondrial activity and increase in micronucleus frequencies in general. The number of similar studies is extremely limited, and in this study in addition to the short-term effects of using the serum infusion sets, the information about the sample tests to determine the biosecurity of disposable medical materials is given.

Does cone-beam computed tomography examination increase the micronuclei frequency in the oral mucosa exfoliated cells? A systematic review and meta-analysis

ObjectiveThis systematic review (SR) with meta-analysis aimed to evaluate the frequency of micronuclei in the oral mucosa exfoliated cells after cone-beam computed tomography (CBCT) examination.MethodsWe performed language-independent computer-assisted data searches using PubMed databases, Cochrane, Embase, Web of Science all databases, and Google Scholar. The literature on micronucleus (MN) frequency of clinical trials before and after CBCT examination was included. The frequency of MN in exfoliated cells of the human oral mucosa was the primary outcome of the study. All statistical analyses were performed with R (version 4.1.0), RStudio (version 2022.02.2 + 485) software, and Meta packages (version 5.2–0). Two reviewers independently assessed the quality of the included studies by the EPHPP (Effective Public Health Practice Project) Modified scale with minor modifications. The heterogeneity of the data was analyzed using I2 statistics, in which I2 > 50% was considered substantial heterogeneity.ResultsA total of 559 articles were selected through the search strategy. After screening titles and abstracts, nine full-text manuscripts were assessed for eligibility, and six observational studies were included in the meta-analysis. The present study showed a significant increase in MN frequency of human oral mucosal exfoliated cells 10 days after CBCT examination compared to baseline (SMD = − 0.56, 95%-CI = − 0.99 ~ − 0.13, p = 0.01). Because of the high heterogeneity among the studies (I2 = 72%), after removing one study that was the main source of heterogeneity, excluding the study (I2 = 47%), the common-effect model was chosen, and the meta-analysis also showed that the frequency of MN in human oral mucosa exfoliated cells increased significantly 10 days after CBCT examination (SMD = − 0.35, 95%-CI = − 0.59 ~ − 0.11, p = 0.004).ConclusionThis review suggested that CBCT examination increases the frequency of micronuclei in oral mucosal exfoliated cells.

Sub-Lethal Effects Of Chlorpyrifos On Geno-Neurotoxic Profiles In Freshwater Fish Cyprinus Carpio(L.)

Pesticide toxicity is a global concern and chlorpyrifos is one of the most widely used organophosphate agrochemicals,it is frequently detected in surface ground waters of India. Chlorpyrifos(CPF) LC50 – 96 h was calculated as 0.318 mg/l using probit analysis based on LC50 value two sublethal concentrations of 1/5th of LC50 (0.0636 mg/l) and 1/10th of LC50 (0.0318 mg/l) were determined. The influence of CPF on the acetylcholinesterase (AChE) activity and Ach content and the Genotoxicity of Cyprinus carpio was evaluated. Results confer that significant inhibition in the AchE activity in brain tissue resulting in a build-up of Ach at the neuronal synapse resulting in a decrease of cholinergic transmission finally impairs the neurophysiological activity and ultimately result in the death of fish although studies on genotoxic effects of CPF were carried out to examine the prevalence of nuclear abnormalities in freshwater fish Cyprinus carpio using Comet and MN assay in blood erythrocytes and gill cells as percentile tail DNA damage as comets and micronuclei and other nuclear anomalies were scored in erythrocytes of Cyprinus carpio it was found that significant effect was observed for both concentrations and periods of exposure in the treated fishes. The highest DNA damage and increased micronuclei frequency and inhibitory AchE activity in Cyprinus carpio were found to be high on the highest sublethal 1/5th of LC50 followed by Sublethal 1/10th of LC50 concentration. Overall findings imply that AchE and genotoxic biomarkers may be useful diagnostic tools for chlorpyrifos poisoning in biomonitoring programs.

Assessment of Cytogenetic Damage in Flexographic Workers: Micronucleus Assay in Exfoliated Buccal Epithelial Cells.

Literature review revealed that chemicals used in the printing industry show an association between genotoxicity and occupational exposure. Flexography is one type of printing technique recently becoming popular because of its fast, cost-efficient, and high-quality label printing. The micronucleus (MN) is considered to be a reliable marker for genotoxic damage, and it has a close association with cancer incidences by determining the presence and the extent of the chromosomal damage. Because there are no studies on flexographic workers (FWs), this study was intended to analyze and evaluate the effect of occupational exposure on the MN frequency of buccal epithelial cells. The study comprised 100 FWs and 100 age-matched healthy controls with and without smoking habits. Buccal epithelial cells were collected from all subjects by using a cytobrush, followed by staining with Feulgen fast green. The MN frequency was recorded for each individual using the Tolbert et al. criteria. Data was statistically analyzed by using one-way analysis of variance and the posthoc test. FWs with smoking habits showed a significant increase in MN frequency (1.86 ± 1.77) than workers without the habit (1.02 ± 1.08) and controls (with the habit 1.26 ± 1.33 and without the habit 0.62 ± 0.92). However, there was no significant increase of MN in FWs without the habit when compared with controls. This study observed the cytogenetic damage in FWs and concludes that these workers are at greater risk for genotoxicity, and the MN assay can serve as a useful biomarker.

Evaluation of Clastogenic/Aneugenic Damage Using the FISH Micronucleus Assay in Mice Exposed to Chromium (VI).

Exposure to chromium (VI) [Cr(VI)] has been postulated to be associated with the induction of cancer. In vivo studies utilizing biomarkers of genotoxic damage could aid in elucidating the mechanisms underlying the genotoxic effects of Cr(VI) and their relationship with carcinogenesis. In this study, the origin (clastogenic and/or aneugenic damage) and kinetics of micronuclei (MN) induced by Cr(VI) were investigated. Hsd:ICR female mice were divided into groups of five individuals each. MN kinetics were measured in groups treated with 20 or 25 mg/kg CrO3 intraperitoneally using acridine orange-coated slides in peripheral blood obtained from the caudal vein 0, 12, 24, 36, 48, 60, and 72 h after treatment. Whereas identification of MN with centromeric DNA (MNK+) was measured at the dose of 20 mg/kg of CrO3, using fluorescence in situ hybridization (FISH) with a centromere-specific probe in peripheral blood obtained at 0, 12, and 48 h after treatment. Control groups were administered vehicle only. Total MN were quantified and the clastogenic/aneugenic effects of Cr(VI) were evaluated based on the proportion of MNK+ versus micronuclei without centromeric DNA (MNK-). There was a significant increase in MN frequencies beginning at 12 h in the Cr(VI)-treated groups demonstrating its genotoxicity. When calculating the MNK+ as a percentage of the total MN, the increase was significant beginning 12 h after treatment. The fact that the MNK+ and MNK- were observed at both evaluation times corroborates Cr(VI) as a genotoxic agent and demonstrates that both clastogenic and aneugenic damages are involved in the formation of MN.

Artemisia vulgaris L., Artemisia alba Turra and their constituents reduce mitomycin C-induced genomic instability in human peripheral blood lymphocytes in vitro

This study aimed to evaluate the effect of aqueous and acetone extracts from Artemisia vulgaris L. (AV) and Artemisia alba Turra (AA), and two major polyphenols compounds (3,5-dihydroxybenzoic acid and quercetin-3-O-glucopyranoside) presented in both extracts of the plants against mitomycin C (MMC)-induced genomic instability. Genomic instability was measured using cytokinesis block micronucleus (MN) assay in human peripheral blood lymphocytes (PBLs) in vitro by analyzing two biomarkers – MN and nuclear division index (NDI). Extracts were tested in a concentration-dependent manner (10–250 µg/mL), while 3,5-dihydroxybenzoic acid and quercetin-3-O-glucopyranoside were tested in three different concentrations, in combination with 0.5 µg/mL of MMC. Aqueous and acetone extracts obtained from both plants significantly reduced MMC-induced MN frequency in PBLs, compared to positive control cells (p < 0.05). Extracts from AV did not affect NDI, whereas the concentrations of 10–100 μg/mL of aqueous and acetone AA extracts significantly elevated MMC-decreased NDI values in comparison to positive control cells (p < 0.05). Combined treatment of 3,5-dihydroxybenzoic acid and MMC showed a significant reduction of MMC-induced MN frequency, while quercetin-3-O-glucopyranoside increased MN frequency compared to positive control cells (p < 0.05). Both compounds decreased NDI values but only at the highest tested concentration of quercetin-3-O-glucopyranoside it was of greater significance. In conclusion, all extracts from AV and AA and 3,5-dihydroxybenzoic acid showed protective effect, whereby aqueous AA demonstrated the highest protective effect on MMC- induced genomic instability, while quercetin-3-O-glucopyranoside showed co-mutagen effect.

A cross-sectional clinical study in women to investigate possible genotoxicity and hematological abnormalities related to the use of black cohosh botanical dietary supplements.

Black cohosh (BC; Actaea racemosa L.), a top-selling botanical dietary supplement, is marketed to women primarily to ameliorate a variety of gynecological symptoms. Due to widespread usage, limited safety information, and sporadic reports of hepatotoxicity, the Division of the National Toxicology Program (DNTP) initially evaluated BC extract in female rats and mice. Following administration of up to 1000 mg/kg/day BC extract by gavage for 90 days, dose-related increases in micronucleated peripheral blood erythrocytes were observed, along with a nonregenerative macrocytic anemia resembling megaloblastic anemia in humans. Because both micronuclei and megaloblastic anemia may signal disruption of folate metabolism, and inadequate folate levels in early pregnancy can adversely affect neurodevelopment, the DNTP conducted a pilot cross-sectional study comparing erythrocyte micronucleus frequencies, folate and B12 levels, and a variety of hematological and clinical chemistry parameters between women who used BC and BC-naïve women. Twenty-three women were enrolled in the BC-exposed group and 28 in the BC-naïve group. Use of any brand of BC-only supplement for at least 3 months was required for inclusion in the BC-exposed group. Supplements were analyzed for chemical composition to allow cross-product comparisons. All participants were healthy, with no known exposures (e.g., x-rays, certain medications) that could influence study endpoints. Findings revealed no increased micronucleus frequencies and no hematological abnormalities in women who used BC supplements. Although reassuring, a larger, prospective study with fewer confounders (e.g., BC product diversity and duration of use) providing greater power to detect subtle effects would increase confidence in these findings.

10-year time course of Hg and organic compounds in Augusta Bay: Bioavailability and biological effects in marine organisms.

In the last century, many Mediterranean coastal areas have been subjected to anthropogenic disturbances from industrial activities, uncontrolled landfills, shipyards, and high maritime traffic. The Augusta Bay (eastern Sicily, Italy) represents an example of a strongly impacted coastal environment with an elevated level of sediments contamination due to the presence of one of the largest European petrochemical plants, combined with an extensive commercial and military harbor. The most significant contaminants were represented by mercury (Hg) and hexachlorobenzene (HCB), derived from a former chlor-alkali plant, and other organic compounds like polycyclic aromatic hydrocarbons (PAHs) and polychlorobiphenyls (PCBs). Since the 1970s, Augusta Bay has become internationally recognized as a contaminated marine environment, although very little information is available regarding the temporal trend of contaminants bioavailability and biological impacts on aquatic organisms. In this study, the Hg and HCB concentrations were investigated over 10 years (from 2003 to 2013) in sediments and invertebrate and vertebrate organisms; these two contaminants' ecotoxicity was further evaluated at a biochemical and cellular level by analyzing the induction of organic biotransformation processes and DNA damages. The results showed high concentrations of Hg and HCB in sediments and their strong bioaccumulation in different species with significantly higher values than those measured in reference sites. This trend was paralleled by increased micronuclei frequency (DNA damage biomarker) and activity of the biotransformation system. While levels of chemicals in sediments remained elevated during the time course, their bioavailability and biological effects showed a gradual decrease after 2003, when the chlor-alkali plant was closed. Environmental persistence of Hg and HCB availability facilitates their bioaccumulation and affects the health status of marine organisms, with possible implications for environmental risk, pollutants transfer, and human health.

An overall assessment of the effects of antidepressant paroxetine on aquatic organisms and human cells

Paroxetine (PRX) is one of the most used antidepressants and an emerging contaminant with potential harmful effects to the environment and human health. The present study investigates in detail the toxic potential of PRX using a battery of bioassays on fresh- and marine species, marine bacteria, and human lymphocytes. All the tested organisms and human lymphocytes were exposed at concentrations ranging from μg L−1 to mg L−1. It was found that PRX can cause toxic effects to aquatic organisms at environmental relevant concentrations (μg L−1 level). A significant effect of PRX was observed in all tested algal species especially at the first 24 h. However, differences in responses and sensitivities among the tested algal species were observed. The most sensitive organism was found to be Dunaliella tertiolecta with IC50 = 0.092 mg L−1 (72 h). In the case of Aliivibrio fischeri, EC50 values were determined to be 16.65, 14.31 and 14.41 mg L−1 for 5, 15 and 30 min of exposure, respectively. PRX also induced cytotoxic and genotoxic effects in human lymphocytes. A dose-dependent increase in micronucleus frequencies was occurred at all tested concentrations with a statistically significant increase in micronucleus frequencies at the medium to high PRX tested concentrations. The findings of the present study expand the available toxicity profile of PRX on aquatic organisms and the knowledge about the potential risk of PRX to induce genotoxic effects in cultured human lymphocytes.

Genotoxicity evaluation of nitrosamine impurities using human TK6 cells transduced with cytochrome P450s.

Many nitrosamines are recognized as mutagens and potent rodent carcinogens. Over the past few years, nitrosamine impurities have been detected in various drugs leading to drug recalls. Although nitrosamines are included in a 'cohort of concern' because of their potential human health risks, most of this concern is based on rodent cancer and bacterial mutagenicity data, and there are little data on their genotoxicity in human-based systems. In this study, we employed human lymphoblastoid TK6 cells transduced with human cytochrome P450 (CYP) 2A6 to evaluate the genotoxicity of six nitrosamines that have been identified as impurities in drug products: N-nitrosodiethylamine (NDEA), N-nitrosoethylisopropylamine (NEIPA), N-nitroso-N-methyl-4-aminobutanoic acid (NMBA), N-nitrosomethylphenylamine (NMPA), N-nitrosodiisopropylamine (NDIPA), and N-nitrosodibutylamine (NDBA). Using flow cytometry-based assays, we found that 24-h treatment with NDEA, NEIPA, NMBA, and NMPA caused concentration-dependent increases in the phosphorylation of histone H2A.X (γH2A.X) in CYP2A6-expressing TK6 cells. Metabolism of these four nitrosamines by CYP2A6 also caused significant increases in micronucleus frequency as well as G2/M phase cell-cycle arrest. In addition, nuclear P53 activation was found in CYP2A6-expressing TK6 cells exposed to NDEA, NEIPA, and NMPA. Overall, the genotoxic potency of the six nitrosamine impurities in our test system was NMPA > NDEA ≈ NEIPA > NMBA > NDBA ≈ NDIPA. This study provides new information on the genotoxic potential of nitrosamines in human cells, complementing test results generated from traditional assays and partially addressing the issue of the relevance of nitrosamine genotoxicity for humans. The metabolically competent human cell system reported here may be a useful model for risk assessment of nitrosamine impurities found in drugs.

29. Increased micronuclei frequency and DNA methylation patterns associated with Down syndrome regression disorder

Down syndrome is associated with intellectual disability and physiological conditions, as well as an increased risk for developing behavioral disorders. A distinct pattern of profound behavioral deterioration in people with Down syndrome that differs from the clinical courses of autism spectrum disorder and/or Alzheimer-like dementia is termed “Down Syndrome Regression Disorder” (DSRD). We hypothesized that acquired somatic cell chromosomal instability and/or epigenetic alterations may contribute to the development of DSRD considering that: (1) DSRD does not typically occur until adolescence/early adulthood and might be associated with stressful events; and (2) increased frequencies of somatic cell chromosomal instability have been associated with stress and/or an aneuploidy chromosomal complement.