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The amphibian micronucleus test has been widely used during the last 30 years to test the genotoxic properties of several chemicals and as a tool for ecogenotoxic monitoring. The vast majority of these studies were performed on peripheral blood of urodelan larvae and anuran tadpoles and to a lesser extent adults were also used. In this study, we developed protocols for measuring micronuclei in adult shed skin cells and larval gill cells of the Italian crested newt (Triturus carnifex). Amphibians were collected from ponds in two protected areas in Italy that differed in their radon content. Twenty-three adult newts and 31 larvae were captured from the radon-rich pond, while 20 adults and 27 larvae were taken from the radon-free site. The animals were brought to the laboratory and the micronucleus test was performed on peripheral blood and shed skins taken from the adults and on larval gills. Samples from the radon-rich site showed micronucleus frequencies higher than those from the radon-free site and the difference was statistically significant in gill cells (P < 0.00001). Moreover, the larval gills seem to be more sensitive than the adult tissues. This method represents an easy (and noninvasive in the case of the shed skin) application of the micronucleus assay that can be useful for environmental studies in situ. Environ. Mol. Mutagen. 56:412-417, 2015. (c) 2014 Wiley Periodicals, Inc.

Telomeres are protein-bound tandemly repeated simple DNA sequences placed at the chromosome ends, their role being essential for maintaining genome integrity. Severe telomere damage can trigger potential carcinogenic events such as chromosome fusions, whilst telomere shortening results (at least in mammals) in a protective mechanism known as 'replicative senescence'. In most Metazoa, telomere shortening is avoided by the ribonucleoprotein telomerase. In this brief overview, we focused on the evolutionary conservation of telomeres and telomerase in basal Metazoa (Ctenophora, Porifera, Placozoa, Cnidaria). In all these taxa, telomerase seems to be active and telomeres show the canonical TTAGGG sequence. Presence of telomerase activity and absence of telomere shortening is in accordance with the lack of senescence seen in basal Metazoa, although a clear correspondence between the “demographic senescence” and “replicative senescence” remains to be elucidated. Nonetheless, basal Metazoa can be useful as model organisms in studies on the evolution of telomere biology, Evo-Devo investigations on the control of telomerase activity, the emergence of senescence, as well as telomere-independent effects of telomerase., Invertebrate Survival Journal, Vol 14 No 1 (2017)

Djungarian hamsters (Phodopus sungorus) are increasingly used as a laboratory model, but published information on their blood cell morphology has not been available. The aim of this study was to describe the morphologic characteristics of peripheral blood cells of P. sungorus. Erythrocytes had an average diameter of 6.02 ?m, ranging between 4 and 7.5 ?m; thus they were smaller than those of other Cricetine (hamster) species. Polychromatic (immature) erythrocytes were relatively abundant, as is common in rodent species. The differential count of leukocytes was similar to that of the golden hamster. The percentage of neutrophils and monocytes was slightly higher than in the golden hamster, while that of lymphocytes was somewhat lower. Neutrophils ranged in size from10-12 ?m, containing neutrophilic granules and well-defined nuclear lobes. Lymphocytes diameters were 9-12 ?m. Small lymphocytes were the most abundant. Monocytes ranged in size from 10-16 ?m, being the largest cells. Eosinophils varied in size from 10-14 ?m, and displayed a ringed nucleus., Scandinavian Journal of Laboratory Animal Sciences, Vol 41 (2015)

Many authors have examined the genotoxic properties of magnetic fields. Some studies detected increases in micronuclei frequencies and chromosomal aberrations in samples taken from individuals professionally exposed, such as photocopying machine workers, power-line operators and railwaymen. More abundantly, laboratory studies validated the hypothesis that magnetic fields would induce DNA damage. Genotoxicity studies included detection of Sister Chromatid Exchange (SCE), Chromosomal Aberrations (CA), presence of 8- hydroxy-2’-deoxyguanosine, the alkaline single cell gel electrophoresis (Comet test) and the Micronucleus test. Among genotoxicity assays, one of the most popular is the micronucleus test, because of its simplicity, sensitivity and reliability. Micronuclei are nuclear remains produced during mitosis (or meiosis) when a chromosome fragment or an entire chromosome fails to migrate with one of the two daughter nuclei formed. Basically, this assay consists in the observation of the variations of the frequencies of micronucleated cells. Investigations have been conducted both with in vitro and in vivo exposure. Several works denied the hypothesis that Extremely Low Frequency (ELF) magnetic fields have genotoxic properties, while other studies have detected positive results only in conditions of co-exposure with other mutagenic agents, such as static magnetic fields, X and gamma rays, benzopyrene, aflatoxine and vinblastine. These results led to the hypothesis that ELF magnetic fields are able to enhance, but not to start, a mutagenic event. This statement could be strengthened when you consider the combined action of ELF and static magnetic fields. In the last years, however, an increasing number of works detected genotoxic properties of ELF magnetic fields, both with in vivo and in vitro exposure.

The last 25 years have seen an ever-growing use of the erythrocyte micronucleus test for measuring damage to mammalian chromosomes in vivo. In addition, staining micronuclei with antikinetochore antibodies from CREST serum discriminates aneugenic from clastogenic damage. The use of the micronucleus test in rats, however, has been problematic because the spleen of adult rats efficiently removes micronucleated erythrocytes from the blood. In the present study, we have treated 5-day-old rats with either X-rays (a clastogen) or vinblastine (an aneugen) and measured micronuclei in erythrocytes from the blood and liver. Each treatment increased the frequency of micronuclei in both tissues, with the percentages of CREST-staining micronuclei reflecting the mechanism of micronucleus induction by the two agents. The results indicate that performing the micronucleus assay in the liver and peripheral blood of 5-day-old rats may be a useful approach for detecting the in vivo genotoxicity of chemical and physical agents

Exposure of human alimentation-destined animals to toxic substances can be an important risk factor for human health. Mutagenicity tests represent a good method for genotoxic effect evaluation of environmental pollutants. The micronucleus frequency in peripheral blood and bone marrow erythrocytes has been evaluated in four domestic Ungulate species (ox, sheep, swine and horse). For each species two or three groups of animals coming from Italy and other European countries were analysed and the results indicate a relatively low mean frequency of micronucleated erythrocytes (ME), both in peripheral blood and bone marrow. The comparison between these two frequencies in the four species studied shows that theMEfrequency in sheep and horse is significantly higher in peripheral blood than in bone marrow, whereas in bovines it is higher in the bone marrow, and in swine the difference is not significant. These results could indicate that in ox and swine the spleen is involved in MEremoval from the peripheral circulation, as is known for other species including man. Nevertheless, it cannot be excluded that the same occurs in the other two species, since they exhibit a relatively low peripheral blood ME frequency as well. Further studies on domestic mammals are needed to clarify the spleen function and verify the use of the peripheral blood micronucleus test for genotoxic biomonitoring. At present, the application of the micronucleus test to the bone marrow seems a more suitable method, but, being invasive, it can be used only as a control system of animal hygiene and health, in addition to the routine tests, rather than for biomonitoring

The Directive 2013/59/Euratom established a reduction of the occupational exposure limits to the lens. Since it is become crucial to estimate the lens absorbed dose, the individual variability of exposed worker's ocular conformations with respect to the data estimated with their personal dosimetry has been studied. The anterior eye conformation of 45 exposed workers was acquired with Scheimpflug imaging and classified according to eye vision conditions (emmetropia, myopia or hypermetropia). Three eye models were computed, with two lens reconstructions, and implemented in an interventional radiology scenario by using Monte Carlo (MC) code. The models were dosimetrically analysed simulating setup A, a theoretical monoenergetic and isotropic photon's source (10-150 keV) and setup B, a more realistic interventional conditions with an angiographic X-ray unit (50, 75, 100 kV-peak). Scheimpflug imaging provided an average anterior chamber depth of (6.4 ± 0.5) mm and a lens depth of (3.9 ± 0.3) mm, together with a reconstructed equatorial lens length of (7.1 - 10.1) mm. Using this data for model's reconstruction, the dose conversion coefficients (DCs) for all ocular structures were simulated. Regardless of the eye model used, DCs show a similar trend with radiation energy which highlights, for the same energy and setup used, no significant dependence on ocular morphology and worker's visual conditions. The maximum difference obtained does not exceed 1% for all eye models or structures analysed. Therefore, the individual variabilities of worker ocular anatomy do not require any additional correction compared to the personal dosimetry data measured with a dedicated lens dosimeter. To estimate the absorbed dose to the other eye structures, it is essential instead to know the spectrum of the source that has generated the irradiation since there are differences considering monoenergetic sources or more realistic angiographic units.

Mandibular hypoplasia, Deafness and Progeroid features with concomitant Lipodystrophy define a rare systemic disorder, named MDPL Syndrome, due to almost always a de novo variant in POLD1 gene, encoding the DNA polymerase δ. We report a MDPL female heterozygote for the recurrent p.Ser605del variant. In order to deepen the functional role of the in frame deletion affecting the polymerase catalytic site of the protein, cellular phenotype has been characterised. MDPL fibroblasts exhibit in vitro nuclear envelope anomalies, accumulation of prelamin A and presence of micronuclei. A decline of cell growth, cellular senescence and a blockage of proliferation in G0/G1 phase complete the aged cellular picture. The evaluation of the genomic instability reveals a delayed recovery from DNA induced-damage. Moreover, the rate of telomere shortening was greater in pathological cells, suggesting the telomere dysfunction as an emerging key feature in MDPL. Our results suggest an alteration in DNA replication/repair function of POLD1 as a primary pathogenetic cause of MDPL. The understanding of the mechanisms linking these cellular characteristics to the accelerated aging and to the wide spectrum of affected tissues and clinical symptoms in the MDPL patients may provide opportunities to develop therapeutic treatments for progeroid syndromes.

The impact of emerging chemical pollutants, on both status and functionality of aquatic ecosystems is worldwide recognized as a relevant issue of concern that should be assessed and managed by researchers, policymakers, and all relevant stakeholders. In Europe, the Reach Regulation has registered more than 100.000 chemical substances daily released in the environment. Furthermore, the effects related to the mixture of substances present in aquatic ecosystems may not be predictable on the basis of chemical analyses alone. This evidence, coupled with the dramatic effects of climate changes on water resources through water scarcity and flooding, makes urgent the application of innovative, fast and reliable monitoring methods. In this context, Effect-Based Methods (EBMs) have been applied in the urban stretch of the Tiber River (Central Italy) with the aim of understanding if detrimental pressures affect aquatic environmental health. In particular, different eco-genotoxicological assays have been used in order to detect genotoxic activity of chemicals present in the river, concurrently characterized by chemical analysis. Teratogenicity and embryo-toxicity have been studied in order to cover additional endpoints. The EBMs have highlighted the presence of diffuse chemical pollution and ecotoxicological effects in the three sampling stations, genotoxicological effects have been also detected through the use of different tests and organisms. The chemical analyses confirmed that in the aquatic ecosystems there is a diffuse presence, even at low concentrations, of emerging contaminants such as pharmaceuticals, not routinely monitored pesticides, personal care products, PFAS. The results of this study can help to identify an appropriate battery of EBMs for future studies and the application of more appropriate measures in order to monitor, mitigate or eliminate chemical contamination and remediate its adverse/detrimental effects on the ecosystem health.

Mancozeb (MZ) and zoxamide (ZOX) are fungicides commonly used in pest control programs to protect vineyards. Their toxic and genotoxic potential were investigated in vitro on HepG2 and A549 cell lines at environmentally relevant concentrations. Cytotoxicity, apoptosis, necrosis and intracellular reactive oxygen species (ROS), comet assay and a micronucleus test with CREST immunofluorescence were used. The expression of a panel of genes involved in apoptosis/necrosis (BAX/BCL2), oxidative stress (NRF2), drug metabolism (CYP1A1) and DNA repair (ERCC1/OGG1) was evaluated by real-time PCR. Both fungicides were cytotoxic at the highest tested concentrations (295.7 and 463.4 µM, respectively), MZ induced necrosis, ZOX did not increase apoptosis but modulated BAX and BCL2 expression, suggesting a different mechanism. Both compounds did not increase ROS, but the induction of CYP1A1 and NRF2 expression supported a pro-oxidant mechanism. The comet assay evidenced MZ genotoxicity, whereas no DNA damage due to ZOX treatment was observed. Positive micronuclei were increased in both cell lines treated with MZ and ZOX, supporting their aneugenic potential. ERCC1 and OGG1 were differently modulated, indicating the efficient activation of the nucleotide excision repair system by both fungicides and the inhibition of the base excision repair system by MZ. Overall, MZ confirmed its toxicity and new ZOX-relevant effects were highlighted.

Telomere length is maintained by either telomerase, a reverse transcriptase, or alternative lengthening of telomeres (ALT), a mechanism that utilizes homologous recombination (HR) proteins. Since access to DNA for HR enzymes is regulated by the chromatin status, it is expected that telomere elongation is linked to epigenetic modifications. The aim of this review is to elucidate the epigenetic features of ALT-positive cells. In order to do this, it is first necessary to understand the telomeric chromatin peculiarities. So far, the epigenetic nature of telomeres is still controversial: some authors describe them as heterochromatic, while for others, they are euchromatic. Similarly, ALT activity should be characterized by the loss (according to most researchers) or formation (as claimed by a minority) of heterochromatin in telomeres. Besides reviewing the main works in this field and the most recent findings, some hypotheses involving the role of telomere non-canonical sequences and the possible spatial heterogeneity of telomeres are given.

Purpose: To examine the phenomena governing the quantitative relationships between acentric fragments and micronuclei and understand which formulas are useful for curve-fitting of experimental data of micronuclei. Materials and methods: A stochastic model, including the phenomena of inclusion, coalescence and culling out, was developed and applied to experimental data. Results: Probabilities for inclusion/exclusion of acentric fragments into daughter nuclei and for coalescence of many fragments into a single micronucleus were found to be not cell type-specific. The biological basis for this result is explained with the lack of DNA damage checkpoints between metaphase (when acentric fragments are scored) and telophase (when micronuclei are formed). The phenomenon of “culling out” cells with high numbers of acentric fragments is also described, along with its proposed biological mechanism. Conclusions: Apart from complex formulas that describe these phenomena, we discuss which simple formulas can best approximate them and when is the case to use them for curve fitting of micronuclei data.