Mutagenesis[JOURNAL]
Sun
200 papers
Ames test study designs for nitrosamine mutagenicity testing: qualitative and quantitative analysis of key assay parameters.
Thomas DN, Wills JW, Tracey H … +6 more , Baldwin SJ, Burman M, Williams AN, Harte DSG, Buckley RA, Lynch AM
Mutagenesis · 2024 Mar · PMID 38112628 · Full text
The robust control of genotoxic N-nitrosamine (NA) impurities is an important safety consideration for the pharmaceutical industry, especially considering recent drug product withdrawals. NAs belong to the 'cohort of con...
The robust control of genotoxic N-nitrosamine (NA) impurities is an important safety consideration for the pharmaceutical industry, especially considering recent drug product withdrawals. NAs belong to the 'cohort of concern' list of genotoxic impurities (ICH M7) because of the mutagenic and carcinogenic potency of this chemical class. In addition, regulatory concerns exist regarding the capacity of the Ames test to predict the carcinogenic potential of NAs because of historically discordant results. The reasons postulated to explain these discordant data generally point to aspects of Ames test study design. These include vehicle solvent choice, liver S9 species, bacterial strain, compound concentration, and use of pre-incubation versus plate incorporation methods. Many of these concerns have their roots in historical data generated prior to the harmonization of Ames test guidelines. Therefore, we investigated various Ames test assay parameters and used qualitative analysis and quantitative benchmark dose modelling to identify which combinations provided the most sensitive conditions in terms of mutagenic potency. Two alkyl-nitrosamines, N-nitrosodimethylamine (NDMA) and N-nitrosodiethylamine (NDEA) were studied. NDMA and NDEA mutagenicity was readily detected in the Ames test and key assay parameters were identified that contributed to assay sensitivity rankings. The pre-incubation method (30-min incubation), appropriate vehicle (water or methanol), and hamster-induced liver S9, alongside Salmonella typhimurium strains TA100 and TA1535 and Escherichia coli strain WP2uvrA(pKM101) provide the most sensitive combination of assay parameters in terms of NDMA and NDEA mutagenic potency in the Ames test. Using these parameters and further quantitative benchmark dose modelling, we show that N-nitrosomethylethylamine (NMEA) is positive in Ames test and therefore should no longer be considered a historically discordant NA. The results presented herein define a sensitive Ames test design that can be deployed for the assessment of NAs to support robust impurity qualifications.
Inhibition of homologous recombination repair by Mirin in ovarian cancer ameliorates carboplatin therapy response in vitro.
Horak J, Vallusova D, Cumova A … +3 more , Holy P, Vodicka P, Opattova A
Mutagenesis · 2025 Mar · PMID 38099488 · Publisher ↗
Chemoresistance poses one of the most significant challenges of cancer therapy. Carboplatin (CbPt) is one of the most used chemotherapeutics in ovarian cancer (OVC) treatment. MRE11 constitutes a part of homologous recom...
Chemoresistance poses one of the most significant challenges of cancer therapy. Carboplatin (CbPt) is one of the most used chemotherapeutics in ovarian cancer (OVC) treatment. MRE11 constitutes a part of homologous recombination (HR), which is responsible for the repair of CbPt-induced DNA damage, particularly DNA crosslinks. The study's main aim was to address the role of HR in CbPt chemoresistance in OVC and to evaluate the possibility of overcoming CbPt chemoresistance by Mirin-mediated MRE11 inhibition in an OVC cell line. Lower expression of MRE11 was associated with better overall survival in a cohort of OVC patients treated with platinum drugs (TCGA dataset, P < 0.05). Using in vitro analyses, we showed that the high expression of HR genes drives the CbPt chemoresistance in our CbPt-resistant cell line model. Moreover, the HR inhibition by Mirin not only increased sensitivity to carboplatin (P < 0.05) but also rescued the sensitivity in the CbPt-resistant model (P < 0.05). Our results suggest that MRE11 inhibition with Mirin may represent a promising way to overcome OVC resistance. More therapy options will ultimately lead to better personalized cancer therapy and improvement of patients' survival.
Maternal exercise during pregnancy modulates genotoxicity caused by high fructose consumption in mice offspring.
Magenis ML, Damiani AP, Monteiro IO … +7 more , Dagostin LS, Silva NDS, Scussel R, Nagashima S, Langie SAS, Pinho RA, de Andrade VM
Mutagenesis · 2024 Mar · PMID 38019677 · Publisher ↗
Pregnancy is a period that is characterized by several metabolic and physiological changes and requires special attention, especially with regard to the relationship between feeding and foetal development. Therefore, the...
Pregnancy is a period that is characterized by several metabolic and physiological changes and requires special attention, especially with regard to the relationship between feeding and foetal development. Therefore, the objective of this study was to evaluate whether the practice of voluntary physical exercise (VPE) in combination with chronic consumption of fructose (FRU) from the beginning of life and/or until the gestational period causes genotoxic changes in pregnant females and in their offspring. Seventy Swiss female mice received FRU in the hydration bottle and/or practiced VPE for 8 weeks (prepregnancy/pregnancy). After the lactation period, the offspring groups were separated by sex. It was observed that the consumption of FRU affected the food consumption, serum concentration of FRU, and glycemic profile in the mothers and that the VPE decreases these parameters. In addition, FRU was genotoxic in the mothers' peripheral tissues and VPE had a preventive effect on these parameters. The offspring showed changes in food consumption, serum FRU concentration, and body weight, in addition to an increase in the adiposity index in male offspring in the FRU (FRU) group and a decrease in the FRU + VPE group. FRU leads to hepatic steatosis in the offspring and VPE was able to decrease the area of steatosis. In addition, FRU led to genotoxicity in the offspring and VPE was able to modulate this effect, reducing damages. In conclusion, we observed that all interventions with VPE had nutritional, genetic, and biochemical benefits of the mother and her offspring.
Long or short? Telomere length and pancreatic cancer and its precursor lesions, a narrative review.
Campa D, Felici A, Corradi C … +4 more , Peduzzi G, Gentiluomo M, Farinella R, Rizzato C
Mutagenesis · 2025 Mar · PMID 37976300 · Publisher ↗
Pancreatic ductal adenocarcinoma (PDAC) is the most common and lethal form of pancreatic cancer, with a survival approaching only 11% at 5 years after diagnosis. In the last 15 years, telomere length (TL) measured in leu...
Pancreatic ductal adenocarcinoma (PDAC) is the most common and lethal form of pancreatic cancer, with a survival approaching only 11% at 5 years after diagnosis. In the last 15 years, telomere length (TL) measured in leukocyte (LTL) has been studied in relation to PDAC risk. The majority of the studies reported an association between short LTL and increased PDAC risk, but the results are heterogeneous. Genome-wide association studies have identified several single-nucleotide polymorphisms (SNPs) in the telomerase reverse transcriptase (TERT) gene as susceptibility loci for PDAC. Polygenic risk scores computed using SNPs associated with LTL have been tested in relation to PDAC susceptibility with various methods and giving contrasting results. The aim of this review is to analyze all publications carried out specifically on LTL, considering LTL measured with qPCR and with genetic proxies, and PDAC risk. Additionally, we will give an overview of the most relevant associations between SNPs in telomere-associated genes and PDAC, to answer the question shorter or longer? Which one of the two is associated with PDAC risk?
Matrine and Oxymatrine: evaluating the gene mutation potential using in silico tools and the bacterial reverse mutation assay (Ames test).
Fischer BC, Musengi Y, König J … +5 more , Sachse B, Hessel-Pras S, Schäfer B, Kneuer C, Herrmann K
Mutagenesis · 2024 Feb · PMID 37877816 · Full text
The quinolizidine alkaloids matrine and its N-oxide oxymatrine occur in plants of the genus Sophora. Recently, matrine was sporadically detected in liquorice products. Morphological similarity of the liquorice plant Glyc...
The quinolizidine alkaloids matrine and its N-oxide oxymatrine occur in plants of the genus Sophora. Recently, matrine was sporadically detected in liquorice products. Morphological similarity of the liquorice plant Glycyrrhiza glabra with Sophora species and resulting confusion during harvesting may explain this contamination, but use of matrine as pesticide has also been reported. The detection of matrine in liquorice products raised concern as some studies suggested a genotoxic activity of matrine and oxymatrine. However, these studies are fraught with uncertainties, putting the reliability and robustness into question. Another issue was that Sophora root extracts were usually tested instead of pure matrine and oxymatrine. The aim of this work was therefore to determine whether matrine and oxymatrine have potential for causing gene mutations. In a first step and to support a weight-of-evidence analysis, in silico predictions were performed to improve the database using expert and statistical systems by VEGA, Leadscope (Instem®), and Nexus (Lhasa Limited). Unfortunately, the confidence levels of the predictions were insufficient to either identify or exclude a mutagenic potential. Thus, in order to obtain reliable results, the bacterial reverse mutation assay (Ames test) was carried out in accordance with OECD Test Guideline 471. The test set included the plate incorporation and the preincubation assay. It was performed with five different bacterial strains in the presence or absence of metabolic activation. Neither matrine nor oxymatrine induced a significant increase in the number of revertants under any of the selected experimental conditions. Overall, it can be concluded that matrine and oxymatrine are unlikely to have a gene mutation potential. Any positive findings with Sophora extracts in the Ames test may be related to other components. Notably, the results also indicated a need to extend the application domain of respective (Q)SAR tools to secondary plant metabolites.
The relationship between lymphocyte DNA damage, coronary artery disease, and blood trace elements.
Erdoğu İ, Dural E, Altundağ H … +2 more , Oymak T, Süzen HS
Mutagenesis · 2024 Feb · PMID 37818856 · Publisher ↗
Somatic DNA damage and causative factors (occupational exposures, foods, habits, etc.) are thought to contribute to the pathogenesis of atherosclerosis, although knowledge about their role in coronary artery disease (CAD...
Somatic DNA damage and causative factors (occupational exposures, foods, habits, etc.) are thought to contribute to the pathogenesis of atherosclerosis, although knowledge about their role in coronary artery disease (CAD) is still insufficient. This study aimed to determine the effects of lymphocyte-DNA damage and blood trace element concentrations on CAD. The single-cell alkaline comet was used in the measuring of the lymphocyte DNA damage in blood samples obtained from patients (n = 99) whose CAD grade was determined by the syntax score while the angiographic intervention was carried out. Blood trace element (n = 14) concentrations were monitored by the inductively coupled plasma-optical emission spectroscopy (ICP-OES) after microwave digestion. The relationship between the DNA damage frequencies of the participants and their syntax scores, blood trace element concentrations, and other demographic and clinic parameters were statistically analyzed. Significant correlations were detected between comet data and syntax score (r = 0.858, P < .001), age (r = 0.337, P < .001), blood-urea (r = 0.360, P < .001), creatinine (r = 0.388, P < .001), HbA1c (0.218, P < .05), ECG-QRS time (r = 0.286, P < .01), ECHO-EF (r = -0.377, P < .001), and platelet (r = -0.222, P < .05). The DNA damage frequencies of the groups formed according to their CAD scores were significantly different from the control group (P < .001) and also each other (P ≤ .01). Comet frequencies and CAD grades were found to be correlated with aging (P < .05). DNA damage frequency and syntax score values were significantly (P < .05) higher in males compared to females. Syntax scores were correlated with aging (r = 0.348, P < .01), ECHO-EF (r = 0.374, P < .001), blood-urea (r = 0.398, P < .001), creatinine (r = 0.433, P < .001), glucose (0.218, P < .05), and HbA1c (r = 0.200, P < .05). Significant correlations were observed between trace elements and demographic values, blood parameters, diseases, angio parameters, ECHO, and ECG parameters. It was observed that the concentrations of trace elements detected in the blood were 93.4% correlated with each other. Lymphocyte DNA damage is a strong biomarker for the atherosclerotic indicator of CAD. Aging is an effective factor both in the DNA damage frequency and CAD risk index. Creatinine and urea are factors that have the power to change the CAD risk index and DNA damage frequency. The higher DNA damage and CAD risk were monitored in males compared to females. The relationship between some biomarkers and blood trace element concentrations showed that further studies are needed to more accurately evaluate the relationship between trace elements, DNA damage frequencies, and CAD.
The known unknowns of mitochondrial carcinogenesis: de novo NUMTs and intercellular mitochondrial transfer.
Harutyunyan T
Mutagenesis · 2024 Feb · PMID 37804235 · Publisher ↗
The translocation of mitochondrial DNA (mtDNA) sequences into the nuclear genome, resulted in the occurrence of nuclear sequences of mitochondrial origin (NUMTs) which can be detected in nearly all sequenced eukaryotes....
The translocation of mitochondrial DNA (mtDNA) sequences into the nuclear genome, resulted in the occurrence of nuclear sequences of mitochondrial origin (NUMTs) which can be detected in nearly all sequenced eukaryotes. However, de novo mtDNA insertions can contribute to the development of pathological conditions including cancer. Recent data indicate that de novo mtDNA translocation into chromosomes can occur due to genotoxic influence of DNA double-strand break-inducing environmental mutagens. This confirms the hypothesis of the involvement of genome instability in the occurrence of mtDNA fragments in chromosomes. Mounting evidence indicates that mitochondria can be transferred from normal cells to cancer cells and recover cellular respiration. These exchanged mitochondria can facilitate cancer progression and metastasis. This review article provides a comprehensive overview of the potential carcinogenicity of mtDNA insertions, and the relevance of mtDNA escape in cancer progression, metastasis, and treatment resistance in humans. Potential molecular targets involved in mtDNA escape and exchange of mitochondria that can be of possible clinical benefits are presented and discussed. Understanding these processes could lead to improved diagnostic approaches, novel therapeutic strategies, and a deeper understanding of the intricate relationship between mitochondria, nuclear DNA, and cancer biology.
Effect of iron and calcium on radiation sensitivity in prostate cancer patients relative to controls.
Dhillon VS, Deo P, Fenech M
Mutagenesis · 2023 Dec · PMID 37779442 · Full text
High intake of red meat and/or dairy products may increase the concentration of iron and calcium in plasma-a risk factor for prostate cancer (PC). Despite our understandings of nutrients and their effects on the genome,...
High intake of red meat and/or dairy products may increase the concentration of iron and calcium in plasma-a risk factor for prostate cancer (PC). Despite our understandings of nutrients and their effects on the genome, studies on the effects of iron and calcium on radiation sensitivity of PC patients are lacking. Therefore, we tested the hypothesis that high plasma levels of iron and calcium could increase baseline or radiation-induced DNA damage in PC patients relative to healthy controls. The present study was performed on 106 PC patients and 132 age-matched healthy individuals. CBMN assay was performed to measure mi-cronuclei (MN), nucleoplasmic bridges (NPBs), and nuclear buds (NBuds) in lymphocytes. Plasma concentrations of iron and calcium were measured using inductively coupled plasma atomic emission spectroscopy. MN, NPBs, and NBuds induced by radiation ex vivo were significantly higher in PC patients with high plasma iron (P = .004, P = .047, and P = .0003, respectively) compared to healthy controls. Radiation-induced MN and NBuds frequency were also significantly higher in PC patients (P = .001 and P = .0001, respectively) with high plasma calcium levels relative to controls. Furthermore, radiation-induced frequency of NBuds was significantly higher in PC patients (P < .0001) with high plasma levels of both iron and calcium relative to controls. Our results support the hypothesis that high iron and calcium levels in plasma increases the sensitivity to radiation-induced DNA damage and point to the need of developing nutrition-based strategies to minimize DNA damage in normal tissue of PC patients undergoing radiotherapy.
In vitro chemopreventive and cytotoxic effects of Amazon mosses Leucobryum martianum (Hornsch.) and Leucobryum laevifolium (Broth) extracts.
Fernandes ADS, de Oliveira CG, Evangelista H … +3 more , Sulamita D S M, Araujo-Lima CF, Felzenszwalb I
Mutagenesis · 2024 Feb · PMID 37776161 · Publisher ↗
Several bioactive compounds, such as polyphenols, demonstrate low toxicity and prominent effects on cancer cells with antioxidant, anti-inflammatory, and antitumor activities. Such compounds can be found in Amazon mosses...
Several bioactive compounds, such as polyphenols, demonstrate low toxicity and prominent effects on cancer cells with antioxidant, anti-inflammatory, and antitumor activities. Such compounds can be found in Amazon mosses Leucobryum martianum (Hornsch.) Hampe ex Müll. Hal. (Hornsch.) and Leucobryum laevifolium (Broth). Antimutagenic assay with Salmonella enterica serovar Typhimurium and cytotoxicity with different eukaryotic cell lines were carried out to screen aqueous, hydroalcoholic, and ethanolic extracts of those Amazon mosses for anticancer potential. The results indicate the capacity of all extracts of both mosses to exert chemopreventive effects against 4-nitroquinoline-N-oxide (4NQO) and 2-aminoanthracene (2-AA), which are direct or indirect mutagens. In particular, the ethanolic and aqueous extract from L. martianum. The ethanolic extract from L. martianum induces significant cytotoxicity by mitochondrial metabolism and cell membrane disruption pathways to tumor or non-tumor cells. The aqueous extract from L. martianum showed a mainly cytotoxic response in the HepG2 cells, a human liver carcinoma, reaching ~90% cytotoxicity. The same extract did not induce significant damage to normal liver cells (F C3H cells) by membrane interaction pathway. The selective cytotoxicity in the aqueous extract of L. martianum makes it a candidate against liver cancer. Further studies, including in vivo models, are necessary to validate the efficacy and safety of the aqueous extract of L. martianum.
Tensin 1 regulated by hepatic leukemia factor represses the progression of prostate cancer.
Zhou H, Wang F
Mutagenesis · 2023 Dec · PMID 37712764 · Publisher ↗
Hepatic leukemia factor (HLF), a transcription factor, is dysregulated in many cancers. This study investigates the function of HLF in prostate cancer (PCa) and its relation to tensin 1 (TNS1). Clinical tissues were coll...
Hepatic leukemia factor (HLF), a transcription factor, is dysregulated in many cancers. This study investigates the function of HLF in prostate cancer (PCa) and its relation to tensin 1 (TNS1). Clinical tissues were collected from 24 PCa patients. Duke University 145 (DU145) and PC3 cells overexpressing HLF were established. HLF signaling was downregulated in PCa tissues compared to adjacent tissues and in DU145 and PC3 cells compared to prostate epithelial cells RWPE-1 or prostate stromal cells (WPMY-1). PCa cell lines with overexpression of HLF had reduced proliferative, migratory, and invasive activity, increased apoptosis, and cell mitosis mostly in the G0/G1 phase. HLF induced the TNS1 transcription to activate the p53 pathway. Depletion of TNS1 reversed the anti-tumor effects of HLF on PCa cells and tumor growth and metastasis in vivo. In summary, our findings suggest that HLF suppressed PCa progression by upregulating TNS1 expression and inducing the p53 pathway activation, which might provide insights into novel strategies for combating PCa.
Multi-endpoint analysis of cadmium chloride-induced genotoxicity shows role for reactive oxygen species and p53 activation in DNA damage induction, cell cycle irregularities, and cell size aberrations.
Stannard LM, Doherty A, Chapman KE … +2 more , Doak SH, Jenkins GJ
Mutagenesis · 2024 Feb · PMID 37555614 · Full text
Cadmium chloride (CdCl2) is a known genotoxic carcinogen, with a mechanism of action thought to partly involve the generation of reactive oxygen species (ROS). We applied here a multi-endpoint approach in vitro to explor...
Cadmium chloride (CdCl2) is a known genotoxic carcinogen, with a mechanism of action thought to partly involve the generation of reactive oxygen species (ROS). We applied here a multi-endpoint approach in vitro to explore the impact of CdCl2 on both the genome and on wider cell biology pathways relevant to cancer. Multi-endpoint approaches are believed to offer greater promise in terms of understanding the holistic effects of carcinogens in vitro. This richer understanding may help better classification of carcinogens as well as allowing detailed mechanisms of action to be identified. We found that CdCl2 caused DNA damage [micronuclei (MN)] in both TK6 and NH32 cells in a dose-dependent manner after 4 h exposure (plus 23 h recovery), with lowest observable effect levels (LOELs) for MN induction of 1 μM (TK6) and 1.6 μM (NH32). This DNA damage induction in TK6 cells was ROS dependent as pretreatment with the antioxidant N-Acetyl Cysteine (1 mM), abrogated this effect. However, 2',7'-dichlorofluorescin diacetate was not capable of detecting the ROS induced by CdCl2. The use of NH32 cells allowed an investigation of the role of p53 as they are a p53 null cell line derived from TK6. NH32 showed a 10-fold increase in MN in untreated cells and a similar dose-dependent effect after CdCl2 treatment. In TK6 cells, CdCl2 also caused activation of p53 (accumulation of total and phosphorylated p53), imposition of cell cycle checkpoints (G2/M) and intriguingly the production of smaller and more eccentric (elongated) cells. Overall, this multi-endpoint study suggests a carcinogenic mechanism of CdCl2 involving ROS generation, oxidative DNA damage and p53 activation, leading to cell cycle abnormalities and impacts of cell size and shape. This study shows how the integration of multiple cell biology endpoints studied in parallel in vitro can help mechanistic understanding of how carcinogens disrupt normal cell biology.
Action-at-a-distance mutations induced by 8-oxo-7,8-dihydroguanine are dependent on APOBEC3.
Fukushima R, Suzuki T, Kobayakawa A … +1 more , Kamiya H
Mutagenesis · 2024 Feb · PMID 37471265 · Publisher ↗
DNA oxidation is a serious threat to genome integrity and is involved in mutations and cancer initiation. The G base is most frequently damaged, and 8-oxo-7,8-dihydroguanine (GO, 8-hydroxyguanine) is one of the predomina...
DNA oxidation is a serious threat to genome integrity and is involved in mutations and cancer initiation. The G base is most frequently damaged, and 8-oxo-7,8-dihydroguanine (GO, 8-hydroxyguanine) is one of the predominant damaged bases. In human cells, GO causes a G:C→T:A transversion mutation at the modified site, and also induces untargeted substitution mutations at the G bases of 5'-GpA-3' dinucleotides (action-at-a-distance mutations). The 5'-GpA-3' sequences are complementary to the 5'-TpC-3' sequences, the preferred substrates for apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like 3 (APOBEC3) cytosine deaminases, and thus their contribution to mutagenesis has been considered. In this study, APOBEC3B, the most abundant APOBEC3 protein in human U2OS cells, was knocked down in human U2OS cells, and a GO-shuttle plasmid was then transfected into the cells. The action-at-a-distance mutations were reduced to ~25% by the knockdown, indicating that GO-induced action-at-a-distance mutations are highly dependent on APOBEC3B in this cell line.
Glutathione-related antioxidant defence, DNA damage, and DNA repair in patients suffering from post-COVID conditions.
Kankaya S, Yavuz F, Tari A … +6 more , Aygun AB, Gunes EG, Bektan Kanat B, Ulugerger Avci G, Yavuzer H, Dincer Y
Mutagenesis · 2023 Aug · PMID 37422797 · Publisher ↗
Post-COVID conditions are defined as the continuation of the symptoms of Coronavirus Disease 2019 (COVID-19) 3 months after the initial Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection, with no othe...
Post-COVID conditions are defined as the continuation of the symptoms of Coronavirus Disease 2019 (COVID-19) 3 months after the initial Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection, with no other explanation. Post-COVID conditions are seen among 30%-60% of patients with asymptomatic or mild forms of COVID-19. The underlying pathophysiological mechanisms of post-COVID conditions are not known. In SARS-CoV-2 infection, activation of the immune system leads to increased production of reactive oxygen molecules, depleted antioxidant reserve, and finally occurrence of oxidative stress. In oxidative stress conditions, DNA damage increases and DNA repair systems impair. In this study, glutathione (GSH) level, glutathione peroxidase (GPx) activity, 8-hydroxydeoxyguanosine (8-OHdG) level, basal, induced, and post-repair DNA damage were investigated in individuals suffering from post-COVID conditions. In the red blood cells, GSH levels and GPx activities were measured with a spectrophotometric assay and a commercial kit. Basal, in vitro H2O2 (hydrogen peroxide)-induced, and post-repair DNA damage (DNA damage after a repair incubation following H2O2-treatment, in vitro) were determined in lymphocytes by the comet assay. The urinary 8-OHdG levels were measured by using a commercial ELISA kit. No significant difference was found between the patient and control groups for GSH level, GPx activity, and basal and H2O2-induced DNA damage. Post-repair DNA damage was found to be higher in the patient group than those in the control group. Urinary 8-OHdG level was lower in the patient group compared to the control group. In the control group, GSH level and post-repair DNA damage were higher in the vaccinated individuals. In conclusion, oxidative stress formed due to the immune response against SARS-COV-2 may impair DNA repair mechanisms. Defective DNA repair may be an underlying pathological mechanism of post-COVID conditions.
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.
Mišík M, Kundi M, Worel N … +6 more , Ferk F, Hutter HP, Grusch M, Nersesyan A, Herrera Morales D, Knasmueller S
Mutagenesis · 2023 Aug · PMID 37418160 · Full text
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 tel...
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.
Long-term cryopreservation of potassium bromate positive assay controls for measurement of oxidatively damaged DNA by the Fpg-modified comet assay: results from the hCOMET ring trial.
Møller P, Azqueta A, Rodriguez-Garraus A … +23 more , Bakuradze T, Richling E, Bankoglu EE, Stopper H, Claudino Bastos V, Langie SAS, Jensen A, Ristori S, Scavone F, Giovannelli L, Wojewódzka M, Kruszewski M, Valdiglesias V, Laffon B, Costa C, Costa S, Paulo Teixeira J, Marino M, Del Bo' C, Riso P, Zheng C, Shaposhnikov S, Collins A
Mutagenesis · 2023 Oct · PMID 37357815 · Publisher ↗
The formamidopyrimidine DNA glycosylase (Fpg)-modified comet assay is widely used for the measurement of oxidatively generated damage to DNA. However, there has not been a recommended long-term positive control for this...
The formamidopyrimidine DNA glycosylase (Fpg)-modified comet assay is widely used for the measurement of oxidatively generated damage to DNA. However, there has not been a recommended long-term positive control for this version of the comet assay. We have investigated potassium bromate as a positive control for the Fpg-modified comet assay because it generates many Fpg-sensitive sites with a little concurrent generation of DNA strand breaks. Eight laboratories used the same procedure for the treatment of monocytic THP-1 cells with potassium bromate (0, 0.5, 1.5, and 4.5 mM) and subsequent cryopreservation in a freezing medium consisting of 50% foetal bovine serum, 40% RPMI-1640 medium, and 10% dimethyl sulphoxide. The samples were analysed by the Fpg-modified comet assay three times over a 3-year period. All laboratories obtained a positive concentration-response relationship in cryopreserved samples (linear regression coefficients ranging from 0.79 to 0.99). However, there was a wide difference in the levels of Fpg-sensitive sites between the laboratory with the lowest (4.2% Tail DNA) and highest (74% Tail DNA) values in THP-1 cells after exposure to 4.5 mM KBrO3. In an attempt to assess sources of inter-laboratory variation in Fpg-sensitive sites, comet images from one experiment in each laboratory were forwarded to a central laboratory for visual scoring. There was high consistency between measurements of %Tail DNA values in each laboratory and the visual score of the same comets done in the central laboratory (r = 0.98, P < 0.001, linear regression). In conclusion, the results show that potassium bromate is a suitable positive comet assay control.
DNA strand break levels in cryopreserved mononuclear blood cell lines measured by the alkaline comet assay: results from the hCOMET ring trial.
Møller P, Azqueta A, Rodriguez-Garraus A … +23 more , Bakuradze T, Richling E, Bankoglu EE, Stopper H, Claudino Bastos V, Langie SAS, Jensen A, Ristori S, Scavone F, Giovannelli L, Wojewódzka M, Kruszewski M, Valdiglesias V, Laffon B, Costa C, Costa S, Paulo Teixeira J, Marino M, Del Bo C, Riso P, Zheng C, Shaposhnikov S, Collins A
Mutagenesis · 2023 Oct · PMID 37357800 · Publisher ↗
The comet assay is widely used in biomonitoring studies for the analysis of DNA damage in leukocytes and peripheral blood mononuclear cells. Rather than processing blood samples directly, it can be desirable to cryoprese...
The comet assay is widely used in biomonitoring studies for the analysis of DNA damage in leukocytes and peripheral blood mononuclear cells. Rather than processing blood samples directly, it can be desirable to cryopreserve whole blood or isolated cells for later analysis by the comet assay. However, this creates concern about artificial accumulation of DNA damage during cryopreservation. In this study, 10 laboratories used standardized cryopreservation and thawing procedures of monocytic (THP-1) or lymphocytic (TK6) cells. Samples were cryopreserved in small aliquots in 50% foetal bovine serum, 40% cell culture medium, and 10% dimethyl sulphoxide. Subsequently, cryopreserved samples were analysed by the standard comet assay on three occasions over a 3-year period. Levels of DNA strand breaks in THP-1 cells were increased (four laboratories), unaltered (four laboratories), or decreased (two laboratories) by long-term storage. Pooled analysis indicates only a modest positive association between storage time and levels of DNA strand breaks in THP-1 cells (0.37% Tail DNA per year, 95% confidence interval: -0.05, 0.78). In contrast, DNA strand break levels were not increased by cryopreservation in TK6 cells. There was inter-laboratory variation in levels of DNA strand breaks in THP-1 cells (SD = 3.7% Tail DNA) and TK6 reference sample cells (SD = 9.4% Tail DNA), whereas the intra-laboratory residual variation was substantially smaller (i.e. SD = 0.4%-2.2% Tail DNA in laboratories with the smallest and largest variation). In conclusion, the study shows that accumulation of DNA strand breaks in cryopreserved mononuclear blood cell lines is not a matter of concern.
Genotoxicity of cytokines at chemotherapy-induced 'storm' concentrations in a model of the human bone marrow.
Asurappulige HSH, Thomas AD, Morse HR
Mutagenesis · 2023 Aug · PMID 37326959 · Full text
Donor cell leukaemia (DCL) is a complication of haematopoietic stem cell transplantation where donated cells become malignant within the patient's bone marrow. As DCL predominates as acute myeloid leukaemia, we hypothesi...
Donor cell leukaemia (DCL) is a complication of haematopoietic stem cell transplantation where donated cells become malignant within the patient's bone marrow. As DCL predominates as acute myeloid leukaemia, we hypothesized that the cytokine storm following chemotherapy played a role in promoting and supporting leukaemogenesis. Cytokines have also been implicated in genotoxicity; thus, we explored a cell line model of the human bone marrow (BM) to secrete myeloid cytokines following drug treatment and their potential to induce micronuclei. HS-5 human stromal cells were exposed to mitoxantrone (MTX) and chlorambucil (CHL) and, for the first time, were profiled for 80 cytokines using an array. Fifty-four cytokines were detected in untreated cells, of which 24 were upregulated and 10 were downregulated by both drugs. FGF-7 was the lowest cytokine to be detected in both untreated and treated cells. Eleven cytokines not detected at baseline were detected following drug exposure. TNFα, IL6, GM-CSF, G-CSF, and TGFβ1 were selected for micronuclei induction. TK6 cells were exposed to these cytokines in isolation and in paired combinations. Only TNFα and TGFβ1 induced micronuclei at healthy concentrations, but all five cytokines induced micronuclei at storm levels, which was further increased when combined in pairs. Of particular concern was that some combinations induced micronuclei at levels above the mitomycin C positive control; however, most combinations were less than the sum of micronuclei induced following exposure to each cytokine in isolation. These data infer a possible role for cytokines through chemotherapy-induced cytokine storm, in the instigation and support of leukaemogenesis in the BM, and implicate the need to evaluate individuals for variability in cytokine secretion as a potential risk factor for complications such as DCL.
Next Generation Sequencing Workshop at the Royal Society of Medicine (London, May 2022): how genomics is on the path to modernizing genetic toxicology.
Lynch AM, Zanoni TB, Salk JJ … +10 more , Martincorena I, Young RR, Kucab J, Valentine CC, Yauk C, Escobar PA, Witt KL, Frötschl R, Reed SH, Ashford A
Mutagenesis · 2023 Aug · PMID 37300447 · Full text
The use of error-corrected Next Generation Sequencing (ecNG) to determine mutagenicity has been a subject of growing interest and potentially a disruptive technology that could supplement, and in time, replace current te...
The use of error-corrected Next Generation Sequencing (ecNG) to determine mutagenicity has been a subject of growing interest and potentially a disruptive technology that could supplement, and in time, replace current testing paradigms in preclinical safety assessment. Considering this, a Next Generation Sequencing Workshop was held at the Royal Society of Medicine in London in May 2022, supported by the United Kingdom Environmental Mutagen Society (UKEMS) and TwinStrand Biosciences (WA, USA), to discuss progress and future applications of this technology. In this meeting report, the invited speakers provide an overview of the Workshop topics covered and identify future directions for research. In the area of somatic mutagenesis, several speakers reviewed recent progress made with correlating ecNGS to classic in vivo transgenic rodent mutation assays as well as exploring the use of this technology directly in humans and animals, and in complex organoid models. Additionally, ecNGS has been used for detecting off-target effects of gene editing tools and emerging data suggest ecNGS potential to measure clonal expansion of cells carrying mutations in cancer driver genes as an early marker of carcinogenic potential and for direct human biomonitoring. As such, the workshop demonstrated the importance of raising awareness and support for advancing the science of ecNGS for mutagenesis, gene editing, and carcinogenesis research. Furthermore, the potential of this new technology to contribute to advances in drug and product development and improve safety assessment was extensively explored.
Current status and future challenges of genotoxicity OECD Test Guidelines for nanomaterials: a workshop report.
Doak SH, Andreoli C, Burgum MJ … +13 more , Chaudhry Q, Bleeker EAJ, Bossa C, Domenech J, Drobne D, Fessard V, Jeliazkova N, Longhin E, Rundén-Pran E, Stępnik M, El Yamani N, Catalán J, Dusinska M
Mutagenesis · 2023 Aug · PMID 37234002 · Full text
Genotoxicity testing for nanomaterials remains challenging as standard testing approaches require some adaptation, and further development of nano-specific OECD Test Guidelines (TGs) and Guidance Documents (GDs) are need...
Genotoxicity testing for nanomaterials remains challenging as standard testing approaches require some adaptation, and further development of nano-specific OECD Test Guidelines (TGs) and Guidance Documents (GDs) are needed. However, the field of genotoxicology continues to progress and new approach methodologies (NAMs) are being developed that could provide relevant information on the range of mechanisms of genotoxic action that may be imparted by nanomaterials. There is a recognition of the need for implementation of new and/or adapted OECD TGs, new OECD GDs, and utilization of NAMs within a genotoxicity testing framework for nanomaterials. As such, the requirements to apply new experimental approaches and data for genotoxicity assessment of nanomaterials in a regulatory context is neither clear, nor used in practice. Thus, an international workshop with representatives from regulatory agencies, industry, government, and academic scientists was convened to discuss these issues. The expert discussion highlighted the current deficiencies that exist in standard testing approaches within exposure regimes, insufficient physicochemical characterization, lack of demonstration of cell or tissue uptake and internalization, and limitations in the coverage of genotoxic modes of action. Regarding the latter aspect, a consensus was reached on the importance of using NAMs to support the genotoxicity assessment of nanomaterials. Also highlighted was the need for close engagement between scientists and regulators to (i) provide clarity on the regulatory needs, (ii) improve the acceptance and use of NAM-generated data, and (iii) define how NAMs may be used as part of weight of evidence approaches for use in regulatory risk assessments.
Visual comet scoring revisited: a guide to scoring comet assay slides and obtaining reliable results.
Møller P, Azqueta A, Sanz-Serrano J … +22 more , Bakuradze T, Richling E, Eyluel Bankoglu E, Stopper H, Claudino Bastos V, Langie SAS, Jensen A, Scavone F, Giovannelli L, Wojewódzka M, Kruszewski M, Valdiglesias V, Laffon B, Costa C, Costa S, Teixeira JP, Marino M, Del Bo C, Riso P, Zheng C, Shaposhnikov S, Collins A
Mutagenesis · 2023 Oct · PMID 37233347 · Publisher ↗
Measurement of DNA migration in the comet assay can be done by image analysis or visual scoring. The latter accounts for 20%-25% of the published comet assay results. Here we assess the intra- and inter-investigator vari...
Measurement of DNA migration in the comet assay can be done by image analysis or visual scoring. The latter accounts for 20%-25% of the published comet assay results. Here we assess the intra- and inter-investigator variability in visual scoring of comets. We include three training sets of comet images, which can be used as reference for researchers who wish to use visual scoring of comets. Investigators in 11 different laboratories scored the comet images using a five-class scoring system. There is inter-investigator variation in the three training sets of comets (i.e. coefficient of variation (CV) = 9.7%, 19.8%, and 15.2% in training sets I-III, respectively). However, there is also a positive correlation of inter-investigator scoring in the three training sets (r = 0.60). Overall, 36% of the variation is attributed to inter-investigator variation and 64% stems from intra-investigator variation in scoring between comets (i.e. the comets in training sets I-III look slightly different and this gives rise to heterogeneity in scoring). Intra-investigator variation in scoring was also assessed by repeated analysis of the training sets by the same investigator. There was larger variation when the training sets were scored over a period of six months (CV = 5.9%-9.6%) as compared to 1 week (CV = 1.3%-6.1%). A subsequent study revealed a high inter-investigator variation when premade slides, prepared in a central laboratory, were stained and scored by investigators in different laboratories (CV = 105% and 18%-20% in premade slides with comets from unexposed and hydrogen peroxide-exposed cells, respectively). The results indicate that further standardization of visual scoring is desirable. Nevertheless, the analysis demonstrates that visual scoring is a reliable way of analysing DNA migration in comets.
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