Visualizing DNA single- and double-strand breaks in the Flash comet assay by DNA polymerase-assisted end-labelling.

In the comet assay, tails are formed after single-cell gel electrophoresis if the cells have been exposed to genotoxic agents. These tails include a mixture of both DNA single-strand breaks (SSBs) and double-strand breaks (DSBs). However, these two types of strand breaks cannot be distinguished using comet assay protocols with conventional DNA stains. Since DSBs are more problematic for the cells, it would be useful if the SSBs and DSBs could be differentially identified in the same comet. In order to be able to distinguish between SSBs and DSBs, we designed a protocol for polymerase-assisted DNA damage analysis (PADDA) to be used in combination with the Flash comet protocol, or on fixed cells. By using DNA polymerase I to label SSBs and terminal deoxynucleotidyl transferase to label DSBs with fluorophore-labelled nucleotides. Herein, TK6-cells or HaCat cells were exposed to either hydrogen peroxide (H2O2), ionising radiation (X-rays) or DNA cutting enzymes, and then subjected to a comet protocol followed by PADDA. PADDA offers a wider detection range, unveiling previously undetected DNA strand breaks.

Chemotherapeutic Mechanism of Action of the Synthetic Resorcinolic Methyl 3,5-dimethoxy-2-octanoylbenzoate.

Resorcinolic lipids are described as potential examples of selective chemotherapeutic adjuvants that can enhance the effects of cyclophosphamide (CYC) while promoting cell death without causing DNA damage. Therefore, the current study attempted to describe how the resorcinolic lipid methyl 3,5-dimethoxy-2-octanoylbenzoate (AMS35BB) interacted with DNA (DNA docking) and how this compound affected genetic toxicology models and other biological characteristics when combined with CYC. We observed that AMS35BB, used alone (7.5 and 10 mg/kg), increases the frequency of genomic damage (comet assay) but not chromosomal damage (micronuclei assay), lowers phagocytosis, and promotes cell death in Swiss male mice. When used in association with CYC, AMS35BB can reduce the risk of genomic damage by up to 33.8% as well as chromosomal damage, splenic phagocytosis, cell death, and lymphocyte frequency. Molecular docking showed that AMS35BB had a higher affinity than the active metabolite of CYC for binding to the DNA double helix major groove. As a result, AMS35BB has the potential to be both an adjuvant when used in association with CYC and a therapeutic candidate for the development of a selective chemotherapeutic drug.

Assessment of the three-test genetic toxicology battery for groundwater metabolites.

The two-test in vitro battery for genotoxicity testing (Ames and micronucleus) has in the majority of cases replaced the three-test battery (as two-test plus mammalian cell gene mutation assay) for the routine testing of chemicals, pharmaceuticals, cosmetics, and agrochemical metabolites originating from food and feed as well as from water treatment. The guidance for testing agrochemical groundwater metabolites, however, still relies on the three-test battery. Data collated in this study from 18 plant protection and related materials highlights the disparity between the often negative Ames and in vitro chromosome aberration data and frequently positive in vitro mammalian cell gene mutation assays. Sixteen of the 18 collated materials with complete datasets were Ames negative, and overall had negative outcomes in in vitro chromosome damage tests (weight of evidence from multiple tests). Mammalian cell gene mutation assays (HPRT and/or mouse lymphoma assay (MLA)) were positive in at least one test for every material with this data. Where both MLA and HPRT tests were performed on the same material, the HPRT seemed to give fewer positive responses. In vivo follow-up tests included combinations of comet assays, unscheduled DNA synthesis, and transgenic rodent gene mutation assays, all gave negative outcomes. The inclusion of mammalian cell gene mutation assays in a three-test battery for groundwater metabolites is therefore not justified and leads to unnecessary in vivo follow-up testing.

Measurement of chromosomal instability and level of DNA damage in peripheral blood mononuclear cells of endometrial cancer patients.

Endometrial cancer is one of the most common invasive gynecologic malignancies in developed countries. The aim of this study was to evaluate chromosomal instability and level of DNA damage in peripheral blood mononuclear cells (PBMCs) of newly diagnosed endometrial cancer patients in relation to health status (diagnosis), age, histological grade of cancer, residence, smoking, number of pregnancies, miscarriages, and abortions. The analyzed sample consisted of 60 individuals, 30 endometrial cancer patients with an average age of 64.37 ±â€…7.08, and 30 healthy control women with an average age of 60.23 ±â€…11.55. Chromosomal instability was evaluated by the cytokinesis-block micronucleus (CBMN) assay, and the level of DNA damage by the single-cell gel electrophoresis (comet) assay in PBMCs. The average frequencies of micronuclei (MNi), nucleoplasmic bridges (NPBs) as well as nuclear buds (NBUDs) were significantly higher in cancer patients compared to controls (P < .0005). There was no difference in the nuclear division index (NDI) among the analyzed samples. The comet assay showed that the patients had a significantly increased genetic damage index (GDI) compared with controls (P < .0005). Using linear regression analysis, we found that health status (diagnosis) had the strongest influence on the MN frequency as well as GDI (P < .0005). Our results indicated that there is a high level of genetic damage in both the level of DNA and the level of chromosomes in the PBMCs of newly diagnosed patients with endometrial cancer, where the frequency and level of damage were significantly affected by health status, grade of cancer, residence, number of pregnancies, miscarriages, and abortions.

Baseline and oxidatively damaged DNA in end-stage renal disease patients on varied hemodialysis regimens: a comet assay assessment.

Global estimates exhibit that one million people have end-stage renal disease, a disease-state characterized by irreversible loss of kidney structure and function, thus necessitating renal replacement therapy. The disease-state, oxidative stress, inflammatory responses, as well as the treatment procedure can have damaging effects on the genetic material. Therefore, the present study was carried out to investigate DNA damage (basal and oxidative) using the comet assay in peripheral blood leukocytes of patients (n = 200) with stage V Chronic Kidney Disease (on dialysis and those recommended but yet to initiate dialysis) and compare it to that in controls (n = 210). Basal DNA damage was significantly elevated (1.13x, p ≤ 0.001) in patients (46.23 ± 0.58% DNA in tail) compared to controls (40.85 ± 0.61% DNA in tail). Oxidative DNA damage was also significantly (p ≤ 0.001) higher in patients (9.18 ± 0.49 vs. 2.59 ± 0.19% tail DNA) compared to controls. Twice-a-week dialysis regimen patients had significantly elevated % tail DNA and Damage Index compared to the non-dialyzed and to the once-a-week dialysis group implying dialysis- induced mechanical stress and blood-dialyzer membrane interactions as probable contributors to elevated DNA damage. The present study with a statistically significant power implies higher disease-associated as well as maintenance therapy (hemodialysis)-induced basal and oxidatively damaged DNA, which if not repaired has the potential to initiate carcinogenesis. These findings mark the need for improvement and development of interventional therapies for delaying disease progression and associated co-morbidities so as to improve life expectancy of patients with kidney disease.

Molecular mechanism of high glucose-induced mitochondrial DNA damage in retinal ganglion cells.

Mitochondrial DNA damage in retinal ganglion cells (RGCs) may be closely related to lesions of glaucoma. RGCs were cultured with different concentrations of glucose and grouped into 3 groups, namely normal control (NC) group, Low-Glu group, and High-Glu group. Cell viability was measured with cell counting kit-8, and cell apoptosis was measured using flow cytometry. The DNA damage was measured with comet assay, and the morphological changes of damaged mitochondria in RGCs were observed using TEM. Western blot analyzed the expression of MRE11, RAD50, and NBS1 protein. Cell viability of RGCs in Low-Glu and High-Glu groups were lower than that of NC group in 48 and 96 h. The cell apoptosis in NC group was 4.9%, the Low-Glu group was 12.2% and High-Glu group was 24.4%. The comet imaging showed that NC cells did not have tailings, but the low-Glu and high-Glu group cells had tailings, indicating that the DNA of RGCs had been damaged. TEM, mitochondrial membrane potential, ROS, mitochondrial oxygen consumption, and ATP content detection results showed that RGCs cultured with high glucose occurred mitochondrial morphology changes and dysfunction. MRE11, RAD50, and NBS1 protein expression associated with DNA damage repair pathway in High-Glu group declined compared with Low-Glu group. Mitochondrial DNA damage caused by high glucose will result in apoptosis of retinal ganglion cells in glaucoma.

Toxicological inhalation studies in rats to substantiate grouping of zinc oxide nanoforms.

BACKGROUND: Significant variations exist in the forms of ZnO, making it impossible to test all forms in in vivo inhalation studies. Hence, grouping and read-across is a common approach under REACH to evaluate the toxicological profile of familiar substances. The objective of this paper is to investigate the potential role of dissolution, size, or coating in grouping ZnO (nano)forms for the purpose of hazard assessment. We performed a 90-day inhalation study (OECD test guideline no. (TG) 413) in rats combined with a reproduction/developmental (neuro)toxicity screening test (TG 421/424/426) with coated and uncoated ZnO nanoforms in comparison with microscale ZnO particles and soluble zinc sulfate. In addition, genotoxicity in the nasal cavity, lungs, liver, and bone marrow was examined via comet assay (TG 489) after 14-day inhalation exposure. RESULTS: ZnO nanoparticles caused local toxicity in the respiratory tract. Systemic effects that were not related to the local irritation were not observed. There was no indication of impaired fertility, developmental toxicity, or developmental neurotoxicity. No indication for genotoxicity of any of the test substances was observed. Local effects were similar across the different ZnO test substances and were reversible after the end of the exposure. CONCLUSION: With exception of local toxicity, this study could not confirm the occasional findings in some of the previous studies regarding the above-mentioned toxicological endpoints. The two representative ZnO nanoforms and the microscale particles showed similar local effects. The ZnO nanoforms most likely exhibit their effects by zinc ions as no particles could be detected after the end of the exposure, and exposure to rapidly soluble zinc sulfate had similar effects. Obviously, material differences between the ZnO particles do not substantially alter their toxicokinetics and toxicodynamics. The grouping of ZnO nanoforms into a set of similar nanoforms is justified by these observations.

Evaluating the mutagenicity of N-nitrosodimethylamine in 2D and 3D HepaRG cell cultures using error-corrected next generation sequencing.

Human liver-derived metabolically competent HepaRG cells have been successfully employed in both two-dimensional (2D) and 3D spheroid formats for performing the comet assay and micronucleus (MN) assay. In the present study, we have investigated expanding the genotoxicity endpoints evaluated in HepaRG cells by detecting mutagenesis using two error-corrected next generation sequencing (ecNGS) technologies, Duplex Sequencing (DS) and High-Fidelity (HiFi) Sequencing. Both HepaRG 2D cells and 3D spheroids were exposed for 72 h to N-nitrosodimethylamine (NDMA), followed by an additional incubation for the fixation of induced mutations. NDMA-induced DNA damage, chromosomal damage, and mutagenesis were determined using the comet assay, MN assay, and ecNGS, respectively. The 72-h treatment with NDMA resulted in concentration-dependent increases in cytotoxicity, DNA damage, MN formation, and mutation frequency in both 2D and 3D cultures, with greater responses observed in the 3D spheroids compared to 2D cells. The mutational spectrum analysis showed that NDMA induced predominantly A:T → G:C transitions, along with a lower frequency of G:C → A:T transitions, and exhibited a different trinucleotide signature relative to the negative control. These results demonstrate that the HepaRG 2D cells and 3D spheroid models can be used for mutagenesis assessment using both DS and HiFi Sequencing, with the caveat that severe cytotoxic concentrations should be avoided when conducting DS. With further validation, the HepaRG 2D/3D system may become a powerful human-based metabolically competent platform for genotoxicity testing.

Genotoxicity and safety pharmacology of the rVSVInd(GML)-mspSGtc vaccine against SARS-CoV-2 in Sprague-Dawley rats and Beagle dogs.

The emergence of coronavirus disease (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) led to a pandemic, prompting rapid vaccine development. Although vaccines are effective, the occurrence of rare adverse events following vaccination highlights the necessity of determining whether the benefits outweigh the risks posed by the infection itself. The recombinant Vesicular Stomatitis Virus (rVSV) platform is a promising vector for vaccines against emerging viruses. However, limited studies have evaluated the genotoxicity and safety pharmacology of this viral vector vaccine, which is crucial to ensure the safety of vaccines developed using this platform. Hence, the present study aimed to assess the genotoxicity and safety pharmacology of the rVSVInd(GML)-mspSGtc COVID-19 vaccine using micronucleus and comet assays, as well as neurobehavioral, body temperature, respiratory, and cardiovascular assessments in Sprague-Dawley rats and beagle dogs. The intramuscular administration of rVSVInd(GML)-mspSGtc at doses up to 1.5 × 109 PFU/animal did not increase the number of bone marrow micronucleated polychromatic erythrocytes or cause liver DNA damage. Additionally, it had no significant impact on neurobehavioral functions in rats and showed marginal temporary changes in body temperature, respiratory rate, heart rate, and electrocardiogram parameters in rats and dogs, all of which resolved within 24 h. Overall, following genotoxicity and pharmacological safety assessments, rVSVInd(GML)-mspSGtc displayed no notable systemic adverse effects in rats and dogs, suggesting its potential as a vaccine candidate for human clinical trials.

Study of the DNA damage and cell death in human peripheral blood mononuclear and HepG2/C3A cells exposed to the synthetic 3-(3-hydroxyphenyl)-7-hydroxycoumarin.

Hydroxycoumarins are an important source of biologically active compounds. Previous studies have shown that the number and position of the hydroxyl substituents in the scaffold play an important role for the observed biological activity. In the present study, 3-(3-hydroxyphenyl)-7-hydroxycoumarin was synthesized, and potential cytogenotoxic effects determined in human HepG2/C3A cells displaying phase 1 and phase 2 enzymes (metabolizing cell ability) and compared to human peripheral blood mononuclear cells (PBMC) without xenobiotics metabolizing capacity. Cell viability was determined with concentrations between 0.01 and 10 µg/ml of 3-(3-hydroxyphenyl)-7-hydroxycoumarin using MTT (3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyl tetrazolium bromide) and trypan blue tests. Genotoxicity was determined utilizing the comet assay, and the clastogenic/aneugenic potential employing the micronucleus (MN) test. The results of the in vitro cytotoxicity assays showed a significant decrease in cell viability of PBMC following exposure to 10 µg/ml concentration of the studied compound after 48 and 72 hr. Comet assay observations noted significant DNA damage in PBMC after 4 hr treatment. No marked cytogenotoxic effects were found in HepG2/C3A cells. No chromosomal mutations were observed in both cell lines. It is important to note that 3-(3-hydroxyphenyl)-7-hydroxycoumarin may exert beneficial pharmacological actions at the low micromolar range and with half-life less than 24 hr. Therefore, the results obtained encourage the continuation of studies on this new molecule for medicinal purposes, but its potential toxicity at higher concentrations and longer exposure times needs to be investigated in further studies.

Evaluation of infliximab-induced genotoxicity and possible action on BCL-2 and P53 genes.

Although the last pandemic created an urgency for development of vaccines, there was a continuous and concerted effort to search for therapeutic medications among existing drugs with different indications. One of the medications of interest that underwent this change was infliximab (IFM). This drug is used as an anti-inflammatory, predominantly in patients with Crohn 's disease, colitis ulcerative, and rheumatoid arthritis. In addition to these patients, individuals infected with Coronavirus Disease (COVID-19) were administered this chimeric monoclonal antibody (IMF) to act as an immunomodulator for patients in the absence of comprehensive research. Consequently, the present study aimed to examine the genotoxic effects attributed to IFM treatment employing different assays in vivo using mouse Mus musculus. Therefore, IFM was found to induce genotoxic effects as evidenced by the comet assay but did not demonstrate genotoxic potential utilizing mouse bone marrow MN test. The results of evaluating the expression of the P53 and BCL-2 genes using RT-qPCR showed stimulation of expression of these genes at 24 hr followed by a decline at 48 hr. Although the comet assay provided positive results, it is noteworthy that based upon negative findings in the micronucleus test, the data did not demonstrate significant changes in the genetic material that might affect the therapeutic use of IFM. The stimulation of expression of P53 and BCL-2 genes at 24 hr followed by a decline at 48 hr suggest a transient, if any, effect on genetic material. However, there is still a need for more research to more comprehensively understand the genotoxic profile of this medication.

Phytochemical profile and determination of cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity of aqueous and ethanolic extracts of Pseudobombax marginatum (A. St.-Hil.) A. Robyns.

Pseudobombax marginatum, popularly known as "embiratanha," is widely used by traditional communities as anti-inflammatory and analgesic agent. This study aimed to determine the phytochemical profile as well as cytotoxicity, acute oral toxicity, genotoxicity, and mutagenicity attributed to exposure to aqueous (AqEx) and ethanolic (EtEx) extracts of embiratanha bark. Phytochemical screening was conducted using thin-layer chromatography (TLC). Cell viability was analyzed using MTT assay with human mammary gland adenocarcinoma (MDA-MB-231) and macrophage (J774A.1) cell lines, exposed to concentrations of 12.5, 25, 50, or 100 µg/ml of either extract. For acute oral toxicity, comet assay and micronucleus (MN) tests, a single dose of 2,000 mg/kg of either extract was administered orally to Wistar rats. TLC analysis identified classes of metabolites in the extracts, including cinnamic acid derivatives, flavonoids, hydrolyzable tannins, condensed tannins, coumarins, and terpenes/steroids. In the cytotoxicity assay, the varying concentrations of extracts derived from embiratanha induced no significant alterations in the viability of MDA-MB-231 cells. The lowest concentration of EtEx significantly increased macrophage J774A.1 viability. However, the higher concentrations of AqEx markedly lowered macrophage J774A.1 viability. Animals exhibited no toxicity in the parameters analyzed in acute oral toxicity, comet assay, and MN tests. Further, EtEx promoted a significant reduction in DNA damage index and DNA damage frequency utilizing the comet assay, while the group treated with AqEx exhibited no marked differences. Thus, data demonstrated that AqEx or EtEx of embiratanha may be considered safe at a dose of 2,000 mg/kg orgally under our experimental conditions tested.

Determination of genoprotection against cyclophosphamide induced toxicity in bone marrow of Swiss albino mice by Moringa oleifera leaves and Tinospora cordifolia stem.

The present study aimed to determine the genoprotective activity and safety of Moringa oleifera leave and Tinospora cordifolia stem extracts against cyclophosphamide (CP)-induced genotoxicity utilizing Swiss albino mice. Animals were divided into 14 groups for subacute treatment with either M. oleifera or T. cordifolia extracts daily for 28 days. The extract doses selected were 100, 200 or 400 mg/kg b.w administered orally alone or combined with CP (50 mg/kg b.w. intraperitoneally daily for 5 days). Analyses performed included the comet assay, micronucleus test (MN) in bone marrow cells and sperm head abnormality assay (SHA). M. oleifera and T. cordifolia extracts induced no significant genotoxic effects on somatic and germ cells. In contrast, for all cells examined M. oleifera and T. cordifolia extracts inhibited DNA damage initiated by CP. Taken together data demonstrated that both plant extracts did not exhibit marked genotoxic effects but displayed potential chemoprotective properties against CP-induced genotoxicity in Swiss mice.

Agricultural exposures and DNA damage in PBMC of female farmers measured using the alkaline comet assay.

OBJECTIVE: Several studies investigated the link between agricultural occupational exposures and DNA damage, in an attempt to bring elements of biological plausibility to the increased cancer risk associated with them. However, only a few of these studies focused on females. METHODS: The comet assay was performed on PBMC (Peripheral Blood Mononuclear Cells) samples from 245 females working in open field farming and cattle raising, located in the Normandy area of France. Individual questionnaires on tasks performed were administered at the time of sampling to directly assess exposures. Environmental exposures were issued from a questionnaire assessing the farm productions. Linear regression analyses were done using the DNA damage scores. RESULTS: Regarding direct exposures, several tasks associated with exposure to potentially harmful chemicals were not associated with DNA damage, but a longer duration of use of herbicide on meadows (p = 0.05) or of cleaning and upkeep of agricultural equipment (p = 0.06) revealed higher DNA damage levels, although the number of exposed women was low. Several indirect and/or environmental exposures were associated with DNA damage in multivariate analyses: a larger surface of meadows (p = 0.006) or the presence of poultry (p = 0.03) was associated with less DNA damage, while the presence of swine (p = 0.01) was associated with higher DNA damage. Smokers and former smokers had less DNA damage than non-smokers (p = 0.0008 and p = 0.03). CONCLUSIONS: We report modified levels of DNA damage for those environmentally exposed to meadows, poultry and pig farming, underlining the need for a better knowledge of the potential health risks experienced by females in this setting.

In vivo alkaline comet assay: Statistical considerations on historical negative and positive control data.

The alkaline comet assay is frequently used as in vivo follow-up test within different regulatory environments to characterize the DNA-damaging potential of different test items. The corresponding OECD Test guideline 489 highlights the importance of statistical analyses and historical control data (HCD) but does not provide detailed procedures. Therefore, the working group "Statistics" of the German-speaking Society for Environmental Mutation Research (GUM) collected HCD from five laboratories and >200 comet assay studies and performed several statistical analyses. Key results included that (I) observed large inter-laboratory effects argue against the use of absolute quality thresholds, (II) > 50% zero values on a slide are considered problematic, due to their influence on slide or animal summary statistics, (III) the type of summarizing measure for single-cell data (e.g., median, arithmetic and geometric mean) may lead to extreme differences in resulting animal tail intensities and study outcome in the HCD. These summarizing values increase the reliability of analysis results by better meeting statistical model assumptions, but at the cost of information loss. Furthermore, the relation between negative and positive control groups in the data set was always satisfactorily (or sufficiently) based on ratio, difference and quantile analyses.

Genotoxicity evaluation of food additive titanium dioxide using a battery of standard in vivo tests.

The increasing use of titanium dioxide (TiO2) nanoparticles (NPs) has raised concern about the safety of food additive TiO2. TiO2 has been considered no longer safe by EFSA due to concerns over genotoxicity, however, there are conflicting opinions upon the safety of TiO2 as a food additive, and the number of in vivo genotoxicity studies conducted on food additive TiO2 was limited. In order to investigate the potential genotoxicity of food additive TiO2, we evaluated the genotoxicity of a commercial food additive TiO2 (average size of 135.54 ± 41.01 nm, range from 60.83 to 230.16 nm, NPs account for 30% by number) using a battery of standard in vivo tests, including mammalian erythrocyte micronucleus test, mammalian bone marrow chromosomal aberration test and in vivo mammalian alkaline comet test. After 15 days of consecutive intragastric administration at doses of 250, 500, and 1000 mg/kgBW, food additive TiO2 neither increased the frequencies of bone marrow micronuclei or chromosomal aberration in mice, nor induced DNA strand breakage in rat liver cells. These results indicate that under the condition of this study, food additive TiO2 does not have genotoxic potential although it contains a fraction of NPs.

Lanthanum nitrate demonstrated no genotoxicity in the conducted tests.

Given the widespread applications in industrial and agricultural production, the health effects of rare earth elements (REEs) have garnered public attention, and the genotoxicity of REEs remains unclear. In this study, we evaluated the genetic effects of lanthanum nitrate, a typical representative of REEs, with guideline-compliant in vivo and in vitro methods. Genotoxicity assays, including the Ames test, comet assay, mice bone marrow erythrocyte micronucleus test, spermatogonial chromosomal aberration test, and sperm malformation assay were conducted to assess mutagenicity, chromosomal damage, DNA damage, and sperm malformation. In the Ames test, no statistically significant increase in bacterial reverse mutation frequencies was found as compared with the negative control. Mice exposed to lanthanum nitrate did not exhibit a statistically significant increase in bone marrow erythrocyte micronucleus frequencies, spermatogonial chromosomal aberration frequencies, or sperm malformation frequencies compared to the negative control (P > 0.05). Additionally, after a 24-h treatment with lanthanum nitrate at concentrations of 1.25, 5, and 20 µg/ml, no cytotoxicity was observed in CHL cells. Furthermore, the comet assay results indicate no significant DNA damage was observed even after exposure to high doses of lanthanum nitrate (20 µg/ml). In conclusion, our findings suggest that lanthanum nitrate does not exhibit genotoxicity.

Biocompatibility of nano/micro-sized pyrophyllite particles by pulmo, liver, kidney and gastric mucosis cells.

Pyrophyllite is the least studied natural clay in terms of its potential in biomedical applications, although there are many deposits of this aluminosilicate around the world. Genotoxicity study was performed in vitro for this mineral. Subsequently, Wister rats were exposed to the pyrophyllite micronized to below 100 µm. After the exposure period, histology of the lung, liver, kidney and gastric tissues were performed, followed by the stereological and hematological analysis. The physicochemical analyses revealed typical XRD characteristics of pyrophyllite clay with particle-size distribution ranging 50 nm-100 µm with stable mineral composition and unique buffering property to pH around 8. The results showed that there were no cytotoxic effects on to THP-1 cells, or genotoxicity of pyrophyllite measured by the Comet assay. In vivo studies are accompanied by the thorough physicochemical characterization of the micronized pyrophyllite. Histology of the lung tissue proved presence of an inflammatory reaction. On the other hand, gastric tissue has shown the selective accumulation of nanoparticles in enterocytes of the stomach only, as supported by ultrastructural analysis. Liver and kidney tissues have shown tolerability for pyrophyllite particles. The results give directions for further comprehensive studies of potential biomedical applications of the pyrophyllite.

Genetic damage among children living in agricultural areas in the North of Colombia.

In recent years, the use of pesticides has increased considerably for pest control and to improve agricultural production. The rural areas of several municipalities of department of Cordoba, north of Colombia, are highly dependent on agriculture. In this study, a questionnaire and field observations about pesticide use and genotoxic damage through the comet assay in peripheral blood lymphocytes of children who live near crop fields was evaluated. Damage Index for Comet Assay (DICA) of five children populations exposed to pesticides (mean of 94.73±53.95 for the municipality of Monteria, the higher damage in this study) were significantly Higher than control children population (mean of 7.56±7.39). Results showed the damage index in children exposed group was higher than in the control group. An inadequate management of pesticides, as well as incorrect disposal of toxic wastes was observed in the study zone.

Assessment of imidacloprid induced genotoxicity in Pethia conchonius (Rosy barb), a common freshwater fish of India.

Imidacloprid is one of the highly efficient, globally used neonicotinoid groups of insecticides. The indiscriminate use of imidacloprid is contaminating large water bodies affecting not only the target organisms but also non-target organisms including fish. The present study aimed to assess the extent of nuclear DNA damage by imidacloprid in Pethia conchonius a freshwater fish in India using comet and micronucleus assays. The LC50 value of imidacloprid was estimated to be 227.33 mg L-1. Based on the LC50-96 h value, three sub-lethal concentrations of imidacloprid, SLC I -18.94 mg L-1, SLC II -28.41 mg L-1 and SLC III -56.83 mg L-1 were used to detect its genotoxic effect at DNA and cellular level. The imidacloprid exposed fishes exhibited higher DNA damage and nuclear abnormalities (p < 0.05) than the control. The %head DNA, %tail DNA, tail length and the frequency of micronuclei with other nuclear abnormalities like blebbed and notched nuclei were significantly higher than the control in a time and concentration-dependent manner. The DNA damage parameters such as %head DNA (29.107 ± 1.843), %tail DNA (70.893 ± 1.843), tail length (361.431 ± 8.455) micronucleus (1.300 ± 0.019), notched (0.844 ± 0.011) and blebbed (0.811 ± 0.011) nuclei were found to be highest for SLC III (56.83 mg L-1) at 96 h. The findings indicate that IMI is highly genotoxic in fish and other vertebrates leading to mutagenic/clastogenic effects. The study will be helpful in optimization of the imidacloprid use.