Fumonisin B1-induced toxicity and oxidative damage in U-118MG glioblastoma cells
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Fragmentation
Viability assay
Comet Assay
Sphingolipid
Fumonisin
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In recent years the Comet assay (or single cell gel electrophoresis assay) has been established as a rapid and sensitive method for the detection of DNA damage. For early genotoxicity screening of new chemical entities in industrial toxicology, the Comet assay is more and more used for assessment of the DNA damaging potential of a test compound. In order to increase compound screening throughput, we have established an image analysis system for fully automated measurement of microscope slides processed in the Comet assay. For the comparative investigation various cell types, such as V79 Chinese hamster cells, mouse lymphoma cells and human leukocytes, were treated with several test compounds. Using tail moment as the quantitative parameter for comet formation, we show a very high correlation between our automatic image analysis system and a commercially available, interactive system (Comet Assay II of Perceptive Instruments). The possibility of analyzing 50 samples within 1 day and the high reproducibility of results make automated image processing a powerful tool for automatic analysis of slides processed in the Comet assay.
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Comet Assay
In vitro toxicology
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The analysis of sperm DNA fragmentation has become a new marker to predict male infertility, and many techniques have been developed. The sperm Comet assay offers the possibility of differentiating single- and double-stranded DNA (ssDNA and dsDNA) breaks, which could have different effects on fertility. The objective of this study was to perform a descriptive characterization of different groups of patients, such as those with asthenoteratozoospermic (ATZ) with or without varicocele, oligoasthenoteratozoospermic (OATZ) or balanced chromosome rearrangements, as compared with fertile donors. The Comet assay was used to investigate sperm samples for ssDNA and dsDNA breaks. The analysis of alkaline and neutral Comet assays in different groups of patients showed different sperm DNA damage profiles. Most fertile donors presented low values for ssDNA and dsDNA fragmentation (low-equivalent Comet profile), which would be the best prognosis for achieving a pregnancy. OATZ, ATZ and ATZ with varicocele presented high percentages of ssDNA and dsDNA fragmentation (high-equivalent Comet assay profile), ATZ with varicocele being associated with the worst prognosis, due to higher levels of DNA fragmentation. Rearranged chromosome carriers display a very high variability and, interestingly, two different profiles were seen: a high-equivalent Comet assay profile, which could be compatible with a bad prognosis, and a non-equivalent Comet assay profile, which has also been found in three fertile donors. Comet assay profiles, applied to different clinical groups, may be useful for determining prognosis in cases of male infertility.
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Fragmentation
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Fumonisin
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The present study investigated the genotoxic effects of flumorph in various organs (brain, liver, spleen, kidney and sperm) of mice. The DNA damage, measured as comet tail length (µm), was determined using the alkaline comet assay. The comet assay is a sensitive assay for the detection of genotoxicity caused by flumorph using mice as a model. Statistically significant increases in comet assay for both dose-dependent and duration-dependent DNA damage were observed in all the organs assessed. The organs exhibited the maximum DNA damage in 96 h at 54 mg/kg body weight. Brain showed maximum DNA damage followed by spleen > kidney > liver > sperm. Our data demonstrated that flumorph had induced systemic genotoxicity in mammals as it caused DNA damage in all tested vital organs, especially in brain and spleen.
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The morphological changes observed by electron microscopy indicate that after internal irradiation with ^153Sm-ESTMP bone tumor cells displayed feature of apoptosis,such as margination of condensed chromatin,chromatin fragmentation.as well as the membranebouded apoptotic bodies formation.THe quantification analysis of fragmentation DNA for bone tumor cells induced by ^153Sm-EDTMP shows that the DNA fragmentation is enhanced with the prolongation of internally irradiated time.These characteristics suggest that ^153Sm-EDTMP internal irradiation could induce bone tumor cells to go9 to apoptosis.
Fragmentation
Apoptotic DNA fragmentation
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Although the Comet assay, a procedure for quantitating DNA damage in mammalian cells, is considered sensitive, it has never been ascertained that its sensitivity is higher than the sensitivity of other genotoxicity assays in mammalian cells. To determine whether the power of the Comet assay to detect a low level of genotoxic potential is superior to those of other genotoxicity assays in mammalian cells, we compared the results of Comet assay with those of micronucleus test (MN test). WTK1 human lymphoblastoid cells were exposed to methyl nitrosourea (MNU), ethyl nitrosourea (ENU), methyl methanesulfonate (MMS), ethyl methanesulfonate (EMS), bleomycin (BLM), or UVC. In Comet assay, cells were exposed to each mutagen with (Comet assay/araC) and without (Comet assay) DNA repair inhibitors (araC and hydroxyurea). Furthermore, acellular Comet assay (acellular assay) was performed to determine how single-strand breaks (SSBs) as the initial damage contributes to DNA migration and/or to micronucleus formation. The lowest genotoxic dose (LGD), which is defined as the lowest dose at which each mutagen causes a positive response on each genotoxicity assay, was used to compare the power of the Comet assay to detect a low level of genotoxic potential and that of MN test; that is, a low LGD indicates a high power. Results are summarized as follows: (1) for all mutagens studied, LGDs were MN test ≦ Comet assay; (2) except for BLM, LGDs were Comet assay/araC ≦ MN test; (3) except for UVC and MNU, LGDs were acellular assay ≦ Comet assay/araC ≦ MN test ≦ Comet assay. The following is suggested by the present findings: (1) LGD in the Comet assay is higher than that in MN test, which suggests that the power of the MN test to detect a low level of genotoxic potential is superior to that of the Comet assay; (2) for the studied mutagens, all assays were able to detect all mutagens correctly, which suggests that the sensitivity of the Comet assay and that of the MN test were exactly identical; (3) the power of the Comet assay to detect a low level of genotoxic potential can be elevated to a level higher than that of MN test by using DNA resynthesis inhibitors, such as araC and HU.
Comet Assay
Ethyl methanesulfonate
Methyl methanesulfonate
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