ABSTRACT
Objective:To compare the effects on DNA strand break induced by ultra-high dose rate (FLASH) electron beam and conventional irradiation, and investigate whether FLASH effect was correlated with a reduction of radiation response.Methods:Aqueous pBR322 plasmid was treated with FLASH (125 Gy/s) and conventional irradiation (0.05 Gy/s) under physioxia (4% O 2) and normoxia (21% O 2). Open circle DNA and linear DNA were detected by agarose gel electrophoresis, and the plasmid DNA damage was quantified with an established mathematical model to calculate the relative biological effect (RBE) of DNA damage. In some experiments, Samwirin A (SW) was applied to scavenge free radicals generated by ionizing radiation. Results:Under physioxia, the yields of DNA strand breakage induced by both FLASH and conventional irradiation had a dose-dependent manner. FLASH irradiation could significantly decrease radiation-induced linear DNA compared with conventional irradiation ( t=5.28, 5.79, 7.01, 7.66, P<0.05). However, when the aqueous plasmid was pretreated with SW, there was no difference of DNA strand breakage between FLASH and conventional irradiation ( P>0.05). Both of the yields of open circle DNA and linear DNA had no difference caused by FLASH and conventional radiotherapy at normoxia, but were significantly higher than those under physioxia. In addition, the yields of linear DNA and open circle DNA induced by FLASH irradiation per Gy were (2.78±0.03) and (1.85±0.17) times higher than those of conventional irradiation, respectively. Conclusions:FLASH irradiation attenuated radiation-induced DNA damage since a low production yield of free radical in comparison with conventional irradiation, and hence the FLASH effect was correlated with oxygen content.
ABSTRACT
Reactive oxygen species (ROS) can induce lesions in different cellular targets, including DNA. Stannous chloride (SnCl2) is a ROS generator, leading to lethality in Escherichia coli (E. coli), with the base excision repair (BER) mechanism playing a role in this process. Many techniques have been developed to detect genotoxicity, as comet assay, in eukaryotic cells, and plasmid DNA agarose gel electrophoresis. In this study, an adaptation of the alkaline gel electrophoresis method was carried out to ascertain the induction of strand breaks by SnCl2 in bacterial DNA, from E. coli BER mutants, and its repair pathway. Results obtained show that SnCl2 was able to induce DNA strand breaks in all strains tested. Moreover, endonuclease IV and exonuclease III play a role in DNA repair. On the whole, data has shown that the alkaline gel electrophoresis assay could be used both for studying DNA strand breaks induction and for associated repair mechanisms.
Espécies reativas de oxigênio (ERO) podem induzir lesões em diferentes alvos celulares, incluindo o DNA. O cloreto estanoso (SnCl2) é um gerador de ERO que induz letalidade em E. coli, sendo o reparo por excisão de bases (BER) um mecanismo importante neste processo. Técnicas como o ensaio cometa (em eucariotos) e a eletroforese de DNA plasmidial em gel de agarose têm sido utilizadas para detectar genotoxicidade. No presente estudo, uma adaptação do método de eletroforese em gel alcalino de agarose foi usada para verificar a indução de quebras, pelo SnCl2, no DNA de E. coli, bem como a participação de enzimas do BER na restauração das lesões. Os resultados mostraram que o SnCl2 induziu quebras no DNA de todas as cepas testadas. Além disso, endonuclease IV e exonuclease III estão envolvidas na reparação dos danos. Em resumo, os dados obtidos indicam que a metodologia de eletroforese em gel alcalino de agarose pode ser empregada tanto para o estudo de quebras no DNA, quanto para avaliação dos mecanismos de reparação associados.
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Objective To investigate the effects of Tripterygium Hypoglaucum (Levl) Hutch alkaloids on the nuclear DNA strand breaks and DNA fragmantation in Jurkat T lymphoma cells to understand the mechanisms of the apoptosis. Methods After Jurkat cells were induced by the alkaloids, DNA stand breaks were labeled by TUNEL assay and DNA fragmentation was analyzed by DNA content analysis. Flow cytometry was performed to determine these phenomena. Results THH alkaloids could effectively induce DNA stand breaks and DNA fragmentation. Conclusion There are great changes in nuclear DNA in the apoptosis of Jurkat T lymphoma cells induced by THH alkaloids.
ABSTRACT
(-)-Epigallocatechin gallate (EGCG), a catechin polyphenol component, is the main ingredient of green tea extract. Although the anti-carcinogenic and cancer inhibitory effects of EGCG have been widely reported, its genotoxicity is not clear and seldom reported. In this study, we examined the effects of EGCG on DNA strand breaks in the isolated lymphocytes and whole blood lymphocytes obtained from two smoking subjects and a nonsmoking healthy subject using a single-cell gel electrophoresis (SCG) assay. The results showed that after 2 hrs of treating the isolated lymphocytes from the smokers, EGCG induced a significant increase in DNA strand breaks at concentrations from 2.5 × 10-5 M to 2.0 × 10-4 M, while after 2 hrs of treating the whole blood obtained from the same smokers, EGCG suppressed the DNA strand breaks in the lymphocytes at concentrations of 1.0 × 10-4 M and 2.0 × 10-4 M. A similar suppressive result was also shown in the whole blood lymphocytes from the nonsmoker at nearly the same concentrations, while at concentrations of 1.0 × 10-3 M or 2.0 × 10-3 M, EGCG induced a significant increase in DNA strand breaks in the whole blood lymphocytes from the nonsmoker. This result suggests that EGCG is not only inhibitory against DNA strand breaks in whole blood, but also genotoxic to the isolated or whole blood lymphocytes at high concentrations. Thus, more research is needed to comprehensively assess the effects of EGCG on genetic materials.
Subject(s)
LymphocytesABSTRACT
Objective To investigate the action of inorganic arsenic in the induction of DNA strand breaks in primary cultured human skin fibroblasts. Methods Sodium arsenate and sodium arsenite were used as test inorganic arsenics. DNA strand breaks were assessed by single cell gel electrophoresis assay (SCGE). Results Arsenate at 1～10 ?mol/L dose_dependently induced DNA strand breaks in cells. Arsenate at lower concentrations induced mainly degree I DNA strand breaks, while the proportion of cells with degree Ⅱ DNA strand breaks increased to 50% when treated with 10?mol/L arsenate,but no cells with degree ⅢDNA strand breaks were observed.Cells treated with 1?mol/L arsenite showed no significant increase in DNA strand breaks. At the concentration of 10?mol/L, however, arsenite induced DNA strand breaks with different degrees, and the apoptotic type DNA strand breaks were the major type. Conclusion Sodium arsenate mainly induced general type DNA strand breaks and sodium arsenite induced apoptotic type,beside general type of DNA strand breaks in primary cultured human skin fibroblasts. This could be explained by their different reaction modes with DNA, and further studies were needed.