The cellular consequences of aneuploidy are largely dependent on the cell types examined. Aneuploid yeasts and mouse embryonic fibroblasts exhibit cell proliferation defects and can be selectively inhibited by compounds that cause proteotoxic or energy stress. By contrast, most aneuploid pluripotent stem cells proliferate rapidly and reach higher saturation densities. The responses of aneuploid pluripotent stem cells to the stress-inducing compounds remain uncharacterized. Here, we tested the response of aneuploid embryonic stem cells to several compounds that caused proteotoxic, energy and genotoxic stress using previously established mouse embryonic stem cell lines trisomic for chromosome 6, 8, 11, or 15. Not all trisomic embryonic stem cells were selectively inhibited by compounds that cause proteotoxic or energy stress. However, most of these cells exhibited increased sensitivity to genotoxins. They displayed elevated DNA damage response as characterized by increased γH2A.X foci under genotoxic stress. Further investigations indicated that elevated autophagy levels might contribute to the increased cytotoxic effects of genotoxins on trisomic embryonic stem cells. Our study laid the foundation for eliminating aneuploidy that might be an effective approach for controlling cancer progression.
In the present study, a disposable electrochemical DNA nano-biosensor is proposed for the rapid detection of genotoxic compounds and bio-analysis of water pollution. The DNA nano-biosensor is prepared by immobilizing DNA on Au nanoparticles and a self-assembled monolayer of cysteamine modified Au electrode. The assembly processes of cysteamine, Au nanoparticles and DNA were characterized by cyclic voltammetry （CV）. The Au nanoparticles enhanced DNA immobilization resulting in an increased guanine signal. The interaction of the analyte with the immobilized DNA was measured through the variation of the electrochemical signal of guanine by square wave voltammetry （SWV）. The biosensor was able to detect the known genotoxic compounds： 2-anthramine, acridine orange and 2- naphthylamine with detection limits of 2, 3 and 50 nmol/L, respectively. The biosensor was also used to test actual water samples to evaluate the contamination level. Additionally, the comparison of results from the classical genotoxiciw bioassay has confirmed the applicability of the method for real samoles.
An E. coli SOS-EGFP biosensor which expresses enhanced green fluorescent protein as a reporter protein under the control of recA gene promoter in SOS response was constructed for detection of DNA damage and evaluation of DNA damaging chemicals. The chemicals that may cause substantial DNA damage will trigger SOS response in the constructed bacterial biosensor, and then the reporter egfp gene under the control of recA promoter is stimulated to express as a fluorescent protein, allowing fast and sensitive fluorescence detection. Interestingly, this biosensor can be simultaneously applied for evaluation of genotoxicity and cytotoxicity. The SOS-EGFP bacterial biosensor provides a sensitive, specific and simple method for detecting known and potential DNA damaging chemicals.
Two highly sensitive methods for the determination of genotoxic alkyl methane sulfonates (AMSs) and alkyl paratoluene sulfonates (APTSs) in lamivudine using hyphenated techniques have been presented. AMSs were determined by GC-MS method using GSBPINOWAX (30 m 0.25 mm 0.25 mm) column. Temperature program was set by maintaining at 100 1C initially for 3 min, then rised to 220 1C at the rate of 15 1C/min and maintained at 220 1C for 16 min. N,N-dimethyl formamide was used as diluent. APTSs were determined by LC-MS using Zorbax, Rx C8, 250 mm 4.6 mm, 5 mm column as stationary phase. 0.01 M ammonium acetate is used as buffer. The mixture of buffer and methanol in 75:25 (v/v) ratio was used as mobile phase A and mixture of buffer and methanol in 5:95 (v/v) ratio was used as mobile phase B. The gradient program (T/%B) was set as 0/28, 16/50, 17/100, 23/100, 27/28 and 40/28. Both the methods were validated as per International Conference on Harmonization guidelines. Limit of quantitation was found 1.5 mg/mL for AMSs and was in the range of 1.0-1.5 mg/mL for APTSs.
