Abstract
To evaluate methods for analysis of genotoxic effects on mammalian cell lines, we tested the effect of three common genotoxic agents on Chinese hamster ovary (CHO) cells by single-cell gel electrophoresis (comet assay) and gas chromatography-tandem mass spectrometry (GC-MS/MS). Suspension-grown CHO cells were separately incubated with etoposide, bleomycin, and ethyl methanesulfonate and analyzed by an alkaline comet assay and GC-MS/MS. Although DNA strand breaks were detected by the comet assay after treatment with all three agents, GC-MS/MS could only detect DNA nucleobase lesions oxidatively induced by bleomycin. This demonstrates that although GC-MS/MS has limitations in detection of genotoxic effects, it can be used for selected chemical genotoxins that contribute to oxidizing processes. The comet assay, used in combination with GC-MS/MS, can be a more useful approach to screen a wide range of chemical genotoxins as well as to monitor other DNA-damaging factors.
Highlights
Exposure of mammalian cells to genotoxic agents and ionizing radiation can result in a wide range of adverse effects, including lipid, protein, and DNA damage
We compared gas chromatography-tandem mass spectrometry (GC-Mass spectrometry (MS)/MS) and the comet assay in detecting DNA damage from three commonly used genotoxic agents: etoposide, bleomycin, and ethyl methanesulfonate (EMS), which are known to have different mechanisms of action and levels of toxicity
By comparing the effectiveness of the alkaline comet assay to gas chromatography-mass spectrometry (GC-MS)/MS, we show that paired measurements of DNA strand breaks and DNA lesions, using these two methods, provide insight into the proportions of these types of damage produced by toxins with a widely varying mechanism of action and potentially by ionizing radiation
Summary
Exposure of mammalian cells to genotoxic agents and ionizing radiation can result in a wide range of adverse effects, including lipid, protein, and DNA damage. DNA damage is implicated in the development of mutagenesis, carcinogenesis, and pathogenesis of several diseases including, Huntington’s, Parkinson’s, Alzheimer’s, and acquired immune deficiency syndrome [1]. Ionizing radiation is another DNA damage source, which is widely used in clinical practice for cancer treatment [2]. Experimental in vitro evidence led to the hypothesis that complex damage that includes both strand breaks and base lesions is more difficult to repair by repair enzymes [3] In this regard, it is critical to have reliable methods to assess both base lesions and strand breaks as induced by chemical agents by detecting the resulting damage to cellular DNA. We compared gas chromatography-tandem mass spectrometry (GC-MS/MS) and the comet assay in detecting DNA damage from three commonly used genotoxic agents: etoposide, bleomycin, and ethyl methanesulfonate (EMS), which are known to have different mechanisms of action and levels of toxicity
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
