Significant Increase In DNA Damage Research Articles

Geno protective efficacy of D-Pinitol isolated from aerial parts of Glycine max L. (Merr.) on Doxorubicin-mediated genotoxicity assessed through in vivo comet assay

The goal of this investigation was to determine whether D-pinitol could protect against doxorubicin-mediated genotoxicity using the in vivo Comet test. Ten groups of Swiss albino mice (n = 6) were formed. Group I (Control) received a 0.9% NaCl treatment for fifteen days. Group II (Positive control) received doxorubicin (DOX) at 5 mg/kg intraperitoneally (i.p.) for three days on the first, eighth, and fifteenth days. Mice were administered D-pinitol (D-P) at various concentrations (100 mg/kg, 200 mg/kg, 300 mg/kg, and 400 mg/kg p.o.) for 15 days, categorized as Groups III, IV, V, and VI based on increasing D-P dosages. Group VII received DOX followed by D-P 100 mg/kg, while Group VIII received D-P 200 mg/kg before DOX. Group IX received D-P 300 mg/kg before DOX, and Group X received D-P 400 mg/kg before DOX. The in vivo Comet assay assessed DNA damage after collecting EDTA blood samples (50 µl) from all experimental and control groups. A highly significant increase in DNA damage (P

Influence of macrophages and neutrophilic granulocyte-like cells on crystalline silica-induced toxicity in human lung epithelial cells.

In many industrial activities, workers may be exposed by inhalation to particles that are aerosolized, To predict the human health hazard of these materials, we propose to develop a co-culture model (macrophages, granulocytes, and alveolar epithelial cells) designed to be more representative of the inflammatory pulmonary response occurring in vivo. Phorbol 12-myristate 13-acetate (PMA)-differentiated THP-1 cells were used as macrophages, All-trans retinoic acid (ATRA)-differentiated HL60 were used as granulocytes and A549 were used as epithelial alveolar type II cells. A crystalline silica sample DQ12 was used as a prototypical particle for its capabilities to induce DNA damage, inflammatory response, and oxidative stress in epithelial cells; its polyvinylpyridine-N-oxide (PVNO)-surface modified counterpart was also used as a negative particulate control. Cells in mono-, bi- or tri-culture were exposed to DQ12 or DQ12-PVNO for 24h. DQ12 but not DQ12-PVNO induced a significant increase in DNA damage in A549 cells. The presence of differentiated THP-1 reduced the genotoxic effects of this crystalline silica sample. The exposure of A549 to DQ12 but not DQ12-PVNO induced a significant change in interleukin-8 (IL-8) protein levels which was exacerbated when differentiated THP-1, and HL-60, were added. In addition, while no production of TNFα was detected in the A549 monoculture, elevated levels of this cytokine were observed in the co-culture systems. This work shows that a cell culture model that takes into consideration the complexity of the pulmonary inflammatory response might be more dependable to study the toxicological properties of particles than "simple" monoculture models.

Genetic instability of a single exposure to sevoflurane at different concentrations in monitored mice.

Sevoflurane is an inhalation anesthetic widely used for general anesthesia, but its genotoxic potential is controversial in clinical studies. It is unknown whether the effects are due to surgery or the anesthetic. Thus, for the first time, the present study investigated genotoxicity in peripheral blood cells and in target organs (liver, lung, and kidney) and micronucleus (MN) in the bone marrow of a single exposure to sevoflurane at three different concentrations in monitored mice. Ninety Swiss mice were distributed into the following groups: exposure to sevoflurane at 3.3% (low), 4.5% (intermediate), and 6.0% (high) in 40% oxygen (O2) for 2 h; negative control (no exposure); negative control with O2; and positive control. The exposed animals were heated, monitored for vital signs (temperature, O2 saturation, heart rate/pulse, and respiratory rate), and anesthetized via a modern low-flow digital system. Mice were euthanized 2 and 24 h after exposure for evaluation by the comet assay and MN test, respectively. No DNA damage occurred in the 3.3% group for any of the organs evaluated, and no genotoxic or mutagenic effects were observed at any sevoflurane concentration in the peripheral blood or liver cells. However, a significant increase in DNA damage was observed at higher concentrations in kidney (4.5%) and lung cells (6.0%) and in the MN frequency (groups 4.5% and 6.0%). No cytotoxicity or histological alterations were observed. In conclusion, high concentrations of sevoflurane induce DNA damage, but concentrations equivalent to those used in clinical practice do not demonstrate genotoxic or mutagenic effects.

DNAR-05. VALIDATING THE ERK5 PROTAC OS11 AS A POTENTIAL NOVEL THERAPEUTIC INEX VIVO GLIOMA STEM CELL MODELS OF INTRATUMOURAL HETEROGENEITY AND RESIDUAL DISEASE

Abstract Glioblastoma is a grade IV brain tumour with poor survival rates of 15 months post-diagnosis, and 5-year survival rates of 10%. Limited progress over the past 40 years to improve the current treatment regimen of surgery and chemo-/radio-therapy are mainly due to tumour heterogeneity that drive resistance mechanisms. The persistence of glioma stem-like cells (GSCs) sub-populations, particularly within post-surgical residual tumour tissue. ERK5 is an emerging oncology drug target that we previously highlighted as a potential target for combined therapy with temozolomide (TMZ) in glioblastoma due to heightened ERK5 expression, which is associated with poor survival. This current project aims to expand on our previous findings by establishing whether targeting ERK5 can sensitise glioblastoma cells to TMZ using primary derived ex vivo GSC models of intratumor heterogeneity and residual disease. Primary GSC models were derived from patients at the Royal Hallamshire Hospital. The PROTAC OS11 (CRUK & University of Manchester) was used to degrade ERK5. DNA damage and cell survival were evaluated following treatment of TMZ alone and in combination with OS11. Consistent with our findings in established glioma cell lines, OS11 was able to robustly degrade ERK5 in a 3D GSC model for at least 72hrs. Combining OS11 and TMZ in GSCs led to a significant increase in DNA damage, with MGMT negative cells exhibiting a greater increase in DNA damage compared to MGMT positive cells. However, unlike our previous findings in established cell lines, this did not lead to reduced cell survival in combination with TMZ. These findings further support the preclinical use of ex vivo models that better represent intratumoural heterogeneity and post-surgical GCS niches. Overall, these data highlight the value of preclinical validation in more clinically relevant models to identify the best possible novel therapeutic approaches to pursue further towards clinical delivery.

Knock-down of FOXO3, GATA2, NFE2L2 and AHR promotes doxorubicin-induced cardiotoxicity in human cardiomyocytes

Recent advances in cancer therapy have substantially increased survival rates among patients, yet the prolonged effect of current treatment regimens with anthracyclines (ACs) often include severe long-term complications, notably in the form of anthracycline-induced cardiotoxicity (AIC). Despite known associations between AC treatment and AIC, a comprehensive understanding of the underlying molecular pathways remains elusive. This gap is highlighted by the scarcity of reliable therapeutic interventions, with dexrazoxane being the sole FDA-approved drug to mitigate AIC risks. This study aims at elucidating the transcriptional response of human cardiomyocytes (hCMs) to AC exposure by analyzing a previously generated RNA-sequencing dataset of cardiac spheroids subjected to clinically relevant doses of ACs. The analysis revealed a robust transcriptional response identified across various time points. We aimed at identifying important transcription factors (TFs) mediating AIC by employing predictive algorithms to highlight key TFs for further experimental validation. Using shRNA constructs, we further assessed the impact of these TFs on hCM response to doxorubicin (DOX) and revealed that these TFs had a notable impact on hCM survival upon DOX exposure. TFs FOXO3, GATA2, AHR and NFE2L2 were further investigated for their role in AIC including cell viability, DOX uptake, DNA damage repair and induction of apoptosis through Cleaved-Caspase 3. Our study demonstrated that eliminating FOXO3 and GATA2 made hCMs more vulnerable to DOX and the lack of GATA2, NFE2L2 and AHR led to significantly higher intracellular levels of DOX. Additionally, FOXO3 played a role in the repair of hCM DNA damage as we observed markedly enhanced levels of CDKN1A. We also noted significant increases in DNA damage through COMET-assays when FOXO3, GATA2, NFE2L2 and AHR were absent. Furthermore, we investigated the clinical relevance by comparing our results with those from a study based on hiPSC-CMs derived from patients with doxorubicin-induced cardiotoxicity, identifying overlapping TFs and their regulatory roles in critical cellular processes like the cell cycle and DNA repair. This approach not only advances the understanding of the molecular mechanisms behind AIC but also opens possible windows for new therapeutic approaches to mitigate the negative side-effects from patient AC treatment.

Growth performance, blood health, and antioxidant status of freshwater carp under brackish water rearing system: Sustainable aquaculture approach in arid and semi-arid conditions

Global warming, water salinization, and increased anthropogenic usage of freshwater sources have had severe effects on water sources and aquaculture practices, which is consequently putting pressure on aquatic fauna and fish production. Hence, brackish water can be used as an alternative and sustainable approach to meet the increasing food requirements. Four groups with triplicates were designed to evaluate the impact of brackish water on Cirrhinus mrigala (n = 100) for 180 days; T0: control group (freshwater), T1: (5.45 ppt salinity), T2: (8.45 ppt salinity) and T3 (12.45 ppt salinity). Little behavioural alterations and normal feeding were observed in all experimental groups as compared to control group, except group T3. Survival rate was 100 % in all groups excluding group T3, which had shown 88 % survival rate (P < 0.05), while mean weight gain and length gain were observed better with low FCR in T1 and T2 groups (P < 0.05) but best growth rate observed at 5 ppt salinity level. Significant reduction in haemoglobin (Hb), hematocrit (HCT), red blood cells (RBCs) and increase in white blood cells (WBCs) and platelets (PLTs) was observed in group T3 compared to other experimental groups (P < 0.05). ALT, AST, ALP, urea, creatinine, and triglycerides, exhibited significant changes in T3 group after 180 days (P < 0.05). Erythrocytes had shown morphological alterations as; micronuclei, notching, blebbing, pear-shaped nuclei and spherocytes in T3 group. Significant increase in DNA damage in various tissues had indicated potential genotoxic impact of highly saline brackish water. Oxidative stress markers were increased significantly and antioxidants reduced (P < 0.05) at high salinity level brackish water (T3) until 180 days. In conclusion, C. mrigala can potentially grow well in brackish water system at 5 ppt salinity whereas, rise in salinity level (12 ppt) is detrimental to freshwater fish health and growth.

Designing and applying a methodology to assess sperm cell viability and DNA damage in a model amphipod

Sperm quality is defined as the sperm cell ability to successfully fertilize eggs and allow normal embryo development⁠. Few studies explore sperm quality using aquatic invertebrates. Parhyale hawaiensis is a marine amphipod with a circumtropical distribution and considered a model for evolution, development, and ecotoxicological studies. We aimed to develop a methodology to collect sperm cells of P. hawaiensis and evaluate their viability and DNA damage (comet assay). We directly exposed the sperm cells to different mutagenic agents to optimize/develop the protocols. Then, as a proof of concept, we exposed the males to mutagenic compounds (EMS, benzo[a]pyrene (BaP), azo and anthraquinone dyes) at non-lethal concentrations verified by the proposed viability test and analyzed their sperm cells for DNA damage (comet assay). Organisms exposed to EMS presented a clear concentration response in the DNA damage response. We also showed that BaP was able to induce a statistically significant increase in DNA damage of the sperm cells. For the two dyes, although DNA damage increased, statistically differences were not observed. We believe we successfully developed a test to detect genotoxicity of chemicals in sperm cells using an invertebrate model. The protocol for sperm cell viability needs to be further explored with different chemicals to verify its utility as a toxicity endpoint. The developed genotoxicity test has the advantages to employ organisms that are easily cultivated in reduced space, use simple laboratory resources and reduced amount of material and reagents. Positive responses with this model could be used to disclose new germ cell mutagen candidates which could be further confirmed in vertebrates' systems.

Effects of low-dose gamma radiation on DNA measured using a quartz tuning fork sensor

Despite the expected direct effects of radiation on DNA through its direct interaction with the biomolecule, research indicates that radiation also interacts with the DNA’s environment, resulting in indirect effects. Therefore, in this study, we explored the feasibility of using the quartz tuning fork (QTF) sensor system in biomedical applications, specifically in detecting DNA damage caused by low doses of gamma radiation, directly and indirectly. We differentiated between direct and indirect damage by analyzing the fork’s resonance frequency changes. This experiment was divided into three stages: before, during, and after irradiation. Each stage involved samples of pure DNA and DNA in a 60-µL aqueous solution, evaluated under identical conditions. Before irradiation, we measured frequency shifts (Δf) over a 20-min period, resulting in values of 19.34, 20.25, and 7.6 Hz for water, DNA, and DNA in water, respectively. Subsequently, the samples were irradiated with cesium-137 for the specified duration, resulting in frequency shifts of ∼ 39.21, 28.37, and 41.23 Hz for the same conditions. Our investigations showed an increase in Δf from 20.25 to 28.3 Hz at doses ranging from 7.5 to 30 µGy for pure DNA. Interestingly, DNA in aqueous solution exhibited hypersensitivity to radiation, with frequency shifts ranging from 7.6 to 41.23 Hz. Furthermore, we observed a significant difference in frequency shift after irradiation between pure DNA and DNA in water, with shifts of ∼ 70.75–98.45 Hz and 56.32–79.28 Hz for DNA and DNA in water, respectively. This result indicates a significant increase in DNA damage in aqueous environments, driven by the generation of active hydroxyl radicals (OH−), resulting in base damage and an associated increase in strand breaks. Consequently, our research indicates a lack of substantial direct impact on DNA repair owing to the absence of a conducive postirradiation environment. Therefore, QTF is a valuable biomarker for radiation sensitivity and is promising for future applications as a mass-sensitive biosensor.

Protective efficacy of ramelteon on methotrexate-induced DNA damage

Ramelteon (RMLT) is a melatonin receptor agonist that it has antioxidative and anti-inflammatory effects associated with DNA damage through different mechanisms of action. In this regard, we investigated the potential usefulness of RMLT as a protective agent against methotrexate (MTX)-induced DNA damage. Four groups were constituted from 32 Wistar albino rats: Negative control, RMLT, MTX, and MTX + RMLT. Twenty mg/kg MTX (i.p., single dose) and RMLT 10 mg/kg (oral, 7 days) was administered. Comet assay was used and the parameter %TailDNA was used to detect DNA damage. %TailDNA was 4.90 ± 0.19 in the control group, 7.85 ± 0.33 in the MTX group, 5.49 ± 0.24 in the RMLT group, and 5.86 ± 0.23 in the MTX + RMLT group. While there was a significant increase in DNA damage in the MTX-treated group compared to the control group, there was a significant reduction in DNA damage in the MTX + RMLT group, compared to the MTX group (p < 0.001). In conclusion, it was observed that combined treatment with RMLT significantly reduced MTX-induced DNA damage.

Assessment of cellular senescence potential of PM2.5 using 3D human lung fibroblast spheroids in vitro model.

Epidemiological studies demonstrate that particulate matter 2.5 (PM2.5) exposure closely related to chronic respiratory diseases. Cellular senescence plays an important role in many diseases. However, it is not fully clear whether PM2.5 exposure could induce cellular senescence in the human lung. In this study, we generated a three-dimensional (3D) spheroid model using isolated primary human lung fibroblasts (HLFs) to investigate the effects of PM2.5 on cellular senescence at the 3D level. 3D spheroids were exposed to 25-100 μg/ml of PM2.5 in order to evaluate the impact on cellular senescence. SA-β-galactosidase activity, cell proliferation, and the expression of key genes and proteins were detected. Exposure of the HLF spheroids to PM2.5 yielded a more sensitive cytotoxicity than 2D HLF cell culture. Importantly, PM2.5 exposure induced the rapid progression of cellular senescence in 3D HLF spheroids, with a dramatically increased SA-β-Gal activity. In exploiting the mechanism underlying the effect of PM2.5 on senescence, we found a significant increase of DNA damage, upregulation of p21 protein levels, and suppression of cell proliferation in PM2.5-treated HLF spheroids. Moreover, PM2.5 exposure created a significant inflammatory response, which may be at least partially associated with the activation of TGF-β1/Smad3 axis and HMGB1 pathway. Our results indicate that PM2.5 could induce DNA damage, inflammation, and cellular senescence in 3D HLF spheroids, which may provide a new evidence for PM2.5 toxicity based on a 3D model which has been shown to be more in vivo-like in their phenotype and physiology than 2D cultures.

Abstract B035: Developing the UValidate platform to measure DNA damage and repair capacity in isogenic donor-derived skin keratinocytes, fibroblasts and melanocyte cell-lines with different Fitzpatrick phototypes

Abstract DNA damage associated with ultraviolet (UV) radiation exposure causes accelerated skin aging and cancer-initiating mutations. Over-the-counter UVA and UVB filters are required in all sunscreens, but transdermal absorption of these compounds have been reported to impair development and reproduction and metabolize into carcinogenic/mutagenic chemical intermediates. To assess the effects of sunscreen active ingredients on UVR-induced DNA damage and repair, we developed a novel high throughput (HTP) screening platform, UValidate, employing mixed populations of isogenic skin cells, exposed to single or combinations of UV filters in the presence of UVB and UVA. UValidate contains a patented LED UVR DNA damage induction system (LUDIS:V3), facilitating programmable precise induction of DNA damage and repair at multiple time points and replicates, using a 96-well configuration capable of simultaneous delivering UVA and UVB. LUDIS:V3 optimization experiments surprisingly revealed that, unlike keratinocytes, skin fibroblasts only repaired UVA DNA damage in conditioned media. We then tested sunscreen active ingredient combinations for their capacity to alter DNA repair and found that: 1) Some sunscreen ingredients, including dioxybenzone, induce a significant increase in DNA damage above endogenous levels. 2) Fibroblasts and keratinocytes respond differently to the treatment protocols. 3) Some sunscreen active compounds contribute to reduction of UVA-induced DNA damage. 4) No sunscreen completely inhibits all UVA DNA damage. We are currently employing the LUDIS:V3 system with HTP comet assays to compare DNA damage and DNA repair capacities, +/- sunscreen active ingredients, of six isogenic sets of primary epidermal keratinocytes (NHEK), melanocytes (NHEM), and fibroblasts (NHDF)` with different Fitzpatrick phototypes I-VI, representative of the diverse American population. These cells were isolated from donor skin samples, and further purified to establish banks of isogenic NHEK, NHEM, and NHDF cell lines. UVA exposure with the LUDIS:V3 and comet assays revealed higher levels of DNA damage in NHEK and NHDF, compared to NHEM, likely due to the UVR-protective effects of melanin in NHEM. Consistently, NHEK with phototype V (from darkly pigmented skin with higher melanin levels) exhibit significantly less UVA-induced DNA damage compared with phototype II NHEK. To create cellular models of dermal disease for DNA repair assays, we have successfully knocked out XPA in multiple NHDF cell lines with different phototypes using a CRISPR-Cas9 KO strategy, as confirmed by immunoblot and NGS sequence analyses. These and other engineered DNA repair-deficient skin cells will be assayed as single or mixed cell-type 2D cultures, with fluorescent labeling allowing for identification of subpopulations in comet and alkaline diffusion assays. This technology addresses the need for better platforms for rapid genotoxicity testing of safer and more effective chemical UV filters to prevent cutaneous carcinogenesis in diverse populations. Citation Format: Dean S. Rosenthal, Elijah Finn, Devin Teehan, Nusrat Islam, Veerupaxagouda Patil, Bonnie Carney, Scott S. Rosenthal, Lucia Dussan, Cynthia M. Simbulan-Rosenthal, Peter Sykora. Developing the UValidate platform to measure DNA damage and repair capacity in isogenic donor-derived skin keratinocytes, fibroblasts and melanocyte cell-lines with different Fitzpatrick phototypes [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: DNA Damage Repair: From Basic Science to Future Clinical Application; 2024 Jan 9-11; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2024;84(1 Suppl):Abstract nr B035.

Pharmacokinetics of Glutapyrone and its Impact on Expression of the Genes Involved in DNA Repair and Free Radical Production in Intact and Diabetic Rats

Abstract Modification of expression of several genes encoding enzymes involved in radical production and DNA repair by a 1,4-DHP derivative glutapyrone was studied in intact rats and animals with streptozotocin diabetes mellitus. Glutapyrone stimulated iNos mRNA production in both kidneys and blood. The effect was stronger in kidneys of diabetic animals, however in blood the expression was down-regulated. The compound increased the Parp 1 gene expression in kidneys of both intact and diabetic animals; in blood the effect was adverse. Expression of XDh gene was significantly increased by glutapyrone in kidneys of intact and diabetic animals. Administration of the compound to intact animals triggered significant increase of DNA damage in white blood cells assayed by comet assay; in diabetic animals no effect was produced. To explain discrepancies with the formerly described effects of glutapyrone on cultured cells, metabolism of the compound was studied. Glutapyrone is either oxidised or the residue of glutamic acid is removed, glutapyrone turns into AV-153, and the latter is metabolised to smaller compounds. Formation of AV-153, a DNA binder and genotoxic compound in high concentrations, can explain DNA damage in white blood cells and stimulation of DNA repair.

Detection of DNA Damage in Fish using Comet Assay

Heavy metals have an enduring presence, risky characteristics, and the propensity to accumulate in the environment. This is why heavy metal toxics are widely acknowledged as harmful environmental pollutants. Heavy metals damage both aquatic and terrestrial ecosystems, posing a major risk to the environment and human health. Four freshwater fish species namely Labeo rohita, Catla catla, Hypophthalmichthys molitrix, and Ctenopharyngodon idella were the focus of this investigation. This study investigated the potential genotoxic effects of lead (Pb), copper (Cu), and cadmium (Cd) on the above fish species through the application of comet assay test. The fish were exposed to these metals at four distinct concentrations (19%, 24%, 31%, and 50% of the LC50) over the course of 40 days. All four fish species were exposed to metals to varying degrees, according to the genetic damage index, cumulative tail length of comets, and the proportion of damaged cells. In contrast to Catla catla, Hypophthalmichthys molitrix had the highest prevalence of DNA damage. The current study suggests that the presence of these particular metals in Pakistan's aquatic ecosystems may have an adverse effect on the DNA of the country's fish species. Metals cause damage to DNA in fibroblast cells through distinct mechanisms when present in water, air, and soil. Comet assay test has a remarkable sensitivity that helps to identify extremely low amounts of DNA damage. Out of the four fish species, Ctenopharyngodon idella showed higher levels of damaged cells, a higher genetic damage index, and a cumulative comet tail length as compared to others. All four fish species experienced a significant increase in DNA damage, genetic damage index, and comet tail length at 50% concentration of metals LC50.

Assessing the Efficacy of Fenugreek Saponin Nanoparticles in Attenuating Nicotine-Induced Hepatotoxicity in Male Rats.

During smoking, nicotine, the most bountiful compound in cigarettes, is absorbed into the body by the lungs and quickly metabolized in the liver, causing three major adverse impacts such as toxic, neoplastic, and immunomodulatory effects. Saponins extracted from several plants are reported to exhibit various biological actions, such as anticancer effects. So, the potential protective effect of fenugreek saponin and nanofenugreek saponin against toxicity induced by nicotine in male rats was investigated in this study. Animals were exposed to nicotine (1.5 mg/kg/day) and/or treated with fenugreek saponin (25, 50, and 100 mg/kg/day) and nanofenugreek saponin (20, 40, and 80 mg/kg/day). Comet assays, histopathological examination, and analyses for the expression levels of glutamate aspartate transporter (GLAST) and glutamate transporter-1 (GLT-1) genes in liver tissues as well as the activity of glutathione peroxidase (GPx) and glutathione-S-transferase (GST) were conducted. The results revealed that nicotine treatment induced a significant increase in DNA damage, decrease in the expression levels of (GLAST) and (GLT-1) genes, and increase in histopathological alterations in liver tissues. Moreover, nicotine treatment induced a significant reduction in the activity of antioxidant enzymes GPx and GST. On the other hand, administration of fenugreek saponin or nanofenugreek saponin with nicotine significantly decreased the DNA damage, increased the expression levels of (GLAST) and (GLT-1) genes, and decreased histopathological alterations in liver tissues. Additionally, a significant increase in the activities of GPx and GST was observed. The results suggested that DNA damage and histological injuries induced by nicotine were decreased by the administration of fenugreek saponin or nanofenugreek saponin; thus, fenugreek saponin and nanofenugreek saponin can be used as ameliorative agents against nicotine toxicity.

Assessment of DNA Damage Induced by Velum® Prime in Human Lymphocytes

Aim: Fluopyram is a mitochondrial complex II inhibitor with low water solubility and a relatively long half-life in soil. So it may also be dangerous for humans. The effect of fluopyram on DNA damage was evaluated in human lymphocytes using the comet assay.&#x0D; Materials and Methods: Lymphocytes of eight volunteers were isolated using histopaque-1077. Fluopyram was administered at doses of 0.05, 0.25, and 1.00 mg/mL for 1, 2, and 4 h. The comet assay was applied, and photographs of the slides were taken under a fluorescence microscope. 50 cells per slide were analyzed using the OpenComet software. The obtained results were statistically evaluated using one-way ANOVA.&#x0D; Results: Fluopyram treatments at 1.00 mg/mL for 1 h and 0.05, 0.25, and 1.00 mg/mL for 2 and 4 h resulted in a statistically significant increase in DNA damage compared to the internal control groups (p

Evaluation of Genotoxic and Cytotoxic Effects of Gluten in Male Albino Mice

Gluten is a protein commonly found in daily diets in the form of wheat, barley, rye, and other grains. It serves as the structural component in flour, providing the binding qualities that maintain the shape and texture of food items. This study aimed to investigate the genotoxic and cytotoxic effects of gluten on bone marrow chromosomes and DNA of male albino mice. The animals were divided into four groups: a control group, a negative control group that received an oral dose of 0.02M glacial acetic acid, and two groups that were treated with gluten dissolved in 0.02M glacial acetic acid at doses of 1.5 g/kg and 3.0 g/kg body weight. The treated animals received oral doses with non-consecutively three times a week for a period of four weeks. The study evaluated chromosomal aberrations in the bone marrow, micronucleus test, and DNA damage using the comet assay. The results of the study showed that treatment with 1.5 and 3.0g/kg body weight of gluten induced chromosomal aberrations and damage in DNA content, with an increase in the severity of effects at a higher dose of gluten. The chromosomal aberrations seen included deletion, fragment, centromeric attenuation, centric fusion, ring formation, end to end association, chromosomal gap, beaded chromosomes, and polyploidy. The micronucleus test revealed toxicity in the bone marrow, as shown by appearance of micronuclei in polychromatic erythrocytes and a reduction in the ratio of polychromatic erythrocytes. The comet assay showed a significant increase of DNA damage in the tail length of the comet cells. This study concluded that the treatment with gluten has detrimental effects on the bone marrow chromosomes and DNA of mice, as demonstrated by the increased chromosomal aberrations, micronuclei, and DNA damage observed in the treated mice. So, the use of gluten should be within an acceptable and safe range.

In vivo protective efficacy of astaxanthin against ionizing radiation-induced DNA damage.

Astaxanthin, a carotenoid pigment, is believed to be effective in the repair of DNA damage. Our study evaluates the effect of astaxanthin on DNA damage in rats exposed to whole-body radiotherapy using the comet assay. Thirty-two male rats were randomly divided into four groups (control, ionizing radiation, astaxanthin, and radiation+astaxanthin). The radiation and radiation+astaxanthin groups were exposed to X-rays at a dose of 8 gray (0.62 gray/min). Astaxanthin was administered at 4 mg/kg by gavage for 7 days starting from irradiation. The %TailDNA parameter was chosen as an indicator of DNA damage and the results were compared using one-way ANOVA. %TailDNA was 3.24 ± 3.12 in the control group, 2.85 ± 2.73 in the astaxanthin group, 4.11 ± 7.90 in the radiation group, and 3.59 ± 4.05 in the radiation+astaxanthin group. There was a significant increase in DNA damage in the radiation group, compared with the control and astaxanthin groups (p < .001). DNA damage was reduced in the radiation+astaxanthin group compared with the radiation group (p < .05). Although this decrease did not reduce damage to the level of the control group, it was significant. The decrease in radiation-induced DNA damage by astaxanthin administration in our study supports the hypothesis that astaxanthin is a promising agent for against/reducing DNA damage.

Impact of mobile phone-specific electromagnetic fields on DNA damage caused by occupationally relevant exposures: results of ex vivo experiments with peripheral blood mononuclear cells from different demographic groups.

The aim of this study was to investigate if age and body mass of humans have an impact on the DNA-damaging properties of high-frequency mobile phone-specific electromagnetic fields (HF-EMF, 1950 MHz, universal mobile telecommunications system, UMTS signal) and if this form of radiation has an impact on the genotoxic effects of occupationally relevant exposures. Pooled peripheral blood mononuclear cells (PBMC) from three groups [young normal weight, young obese (YO), and older age normal weight individuals] were exposed to different doses of HF-EMF (0.25, 0.5, and 1.0 W/kg specific absorption rate-SAR) and simultaneously or sequentially to different chemicals which cause DNA damage (CrO3, NiCl2, benzo[a]pyrene diol epoxide-BPDE, and 4-nitroquinoline 1-oxide-4NQO) via different molecular mechanisms. We found no difference in regard to the background values in the three groups but a significant increase of DNA damage (81% without and 36% with serum) in cells from old participants after radiation with 1.0 W/kg SAR 16 h. In combined treatment experiments we found no impact of the UMTS signal on chemically induced DNA damage in the different groups in general. However, a moderate decrease of DNA damage was seen in simultaneous treatment experiments with BPDE and 1.0 W/kg SAR in the YO group (decline 18%). Taken together our findings indicate that HF-EMF cause DNA damage in PBMC from older subjects (69.1 years). Furthermore, they show that the radiation does not increase induction of DNA damage by occupationally relevant chemicals.

Genotoxicity from metronidazole detected in vitro, but not in vivo, in healthy dogs in a randomized clinical trial.

To determine whether metronidazole (MTZ), at recommended therapeutic dosages in dogs, induces peripheral blood cell (PMBC) genotoxicity, using the γ-H2AX assay as a sensitive measure of DNA breaks. The secondary aim was to assess dose-dependent genotoxicity in vitro in dog and cat PBMCs exposed to increasing MTZ concentrations. 12 healthy employee- and student-owned dogs and blood samples from 2 other healthy untreated dogs and 2 healthy untreated cats. Screened dogs were randomized to receive lower-dose MTZ (7.5 mg/kg, PO, q 12 h) or higher-dose MTZ (20 mg/kg, PO, q 12 h) for 7 days. Blood was drawn at baseline, after the 1 week of treatment, and after a 1-week washout, for DNA damage assessment and serum MTZ concentration measurements. For in vitro studies, PBMCs from untreated healthy dogs and cats were exposed to 0 to 500 μg/mL MTZ. No dogs showed a significant increase in DNA damage at these MTZ dosages for 1 week. The highest serum MTZ concentration observed 1 hour after dosing was 36 μg/mL. In vitro, MTZ led to a significant increase in DNA damage at 100 μg/mL in both canine and feline PBMCs. Although MTZ was not significantly genotoxic in vivo in the healthy dogs in this study, MTZ was significantly genotoxic to canine PBMCs in vitro at 3-fold higher concentrations than those documented in vivo. The safety of MTZ in clinically ill dogs, which may have impaired MTZ clearance or DNA repair, should be assessed next.

Is a non-cytotoxic and non-genotoxic novel bioinspired dipeptide structure synthesis possible for theragnostic applications?

The diagnosis and treatment of the diseases in a certain coordination is a subject that has been emphasized in recent years. Theragnostics approaches allow simultaneous diagnosis and treatment of chronic diseases such as cancer. An ideal theragnostic should be biocompatible and can be used safely in humans. Although several types of theragnostics have been developed, none of yet satisfied these criteria. Bioinspired materials with noble metal centers encapsulating therapeutic and imaging agents were shown to possess theragnostic activities. In this study, it was aimed to synthesize, characterize, and evaluate the cytotoxic and genotoxic effects of self-assembly of diphenylalanine (Phe–Phe) dipeptides presence of mercury (Hg2+) ions to be used for theragnostic. Cytotoxicity and genotoxicity studies were done in mouse fibroblast (NIH/3T3) cells by 3-(4,5-Dimethylthiazol-2-yl)- 2,5-diphenyltetrazolium bromide (MTT) and single cell gel electrophoresis (Comet) assays, respectively. It was found that cell viability decreased in a dose-dependent manner in 24-, 48-, and 72-h treatment. Also, Phe–Phe dipeptides did not cause any significant changes in DNA damage at the concentrations of 1, 2, and 5 mg/mL in 4- and 24-h exposures. In the 48-h exposure, Phe–Phe peptide exposure at concentrations of 2 and 5 mg/mL caused a significant increase in DNA damage and in the 72-h of exposure, a significant increase in DNA damage was observed at all studied concentrations. According to the results of the study, it can be said that Phe–Phe dipeptides presence of Hg2+ ions are biocompatible and can be used safely for theragnostic purposes.