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Objective To investigate the genotoxicity of aniline and repair dynamics in hepatocytes and lymph-cytes.Methods Aniline was administered intragastrically to SPF Kunming mice ( five mice in each group) in a single dose of 100 mg/kg body weight.The hepatocytes and peripheral blood lymphocytes were obtained at 3, 8, 16, 24, and 32 hours after aniline administration, respectively.The control mice received tap water only.The DNA damages were detected by single cell gel electrophoresis assay ( SCGE) and the time-effect relationship was analyzed.Results The results of SCGE experiment showed that both the tail lenth and tail moment of the hepatocyte DNA were increased gradually from 8 h, and reached the maximum at 16 h ( P0.05).The two DNA damage indexes of peripheral blood lymphocytes started to increase at 16 h, reached the maxi-mum at 24 h ( P<0.01) , and began to recover at 32 h after aniline administration.Conclusions Our findings suggeste that aniline may be a potential genotoxicant to hepatocytes and peripheral blood lymphocytes.There is a clear time-response relationship in terms of the two DNA damage indexes, indicating that hepatocytes and lymphocytes in mice possess an effi-cient DNA repair mechanism against aniline toxicity.
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Objective To investigate the characteristics of cisplatin resistance in human gastric cancer cell line SGC-7901 . Methods Single cell gel electrophoresis ( SCGE ) was used to measure the level of DNA damage and repair in gastric cancer cell line SGC-7901 and gastric cancer cisplatin resistance cell line SGC-7901/DDP by ob-serving the tail length. Morphological changes of SGC-7901 and SGC-7901/DDP were recorded to evalutate the differences between the two lines. The degree of migration of SGC-7901/DDP and SGC-7901 measured by cell wound scratch assay was used to estimate the ability of invasion. MTT assay was performed to determine the drug sensitivity, IC50 values and cross-resistance of SGC-7901/DDP treated with 5-FU, VP-16, ADM, TAX and LOHP separately. Results The level of DNA damage and repair in SGC-7901/DDP was higher than that in SGC-7901 ac-cording to the tail length of SCGE tests ,suggesting the relationship between cisplatin resistance and the abiity of DNA damage and repair. There was a much smaller volume in SGC-7901/DDP compared with SGC-7901,and clone aggregation always appeared in SGC-7901/DDP. Cell wound scratch assay showed that the migration of SGC-7901/DDP was weaker than that of SGC-7901. MTT showed the significant increase of IC50 and cross resitance in SGC-7901/DDP compared with SGC-7901 treated with 5-FU, VP-16, ADM, TAX and LOHP simultaneously. Conclu-sion The ability of DNA damage and repair in gastric cancer cisplatin resistance cell line SGC-7901 is enhanced significantly. The SGC-7901/DDP shows a notable promotion on multidrug resistance in vivo, and the migration of SGC-7901/DDP is weaker than that of SGC-7901 .
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Zearalenone (ZEN) is a non-steroidal estrogenic mycotoxin produced by several species of Fusarium that are found in cereals and agricultural products. ZEN has been implicated in mycotoxicosis in farm animals and in humans. The toxic effects of ZEN are well known, but the ability of an alkaline Comet assay to assess ZEN-induced oxidative DNA damage in Chang liver cells has not been established. The first aim of this study was to evaluate the Comet assay for the determination of cytotoxicity and extent of DNA damage induced by ZEN toxin, and the second aim was to investigate the ability of N-acetylcysteine amide (NACA) to protect cells from ZEN-induced toxicity. In the Comet assay, DNA damage was assessed by quantifying the tail extent moment (TEM; arbitrary unit) and tail length (TL; arbitrary unit), which are used as indicators of DNA strand breaks in SCGE. The cytotoxic effects of ZEN in Chang liver cells were mediated by inhibition of cell proliferation and induction of oxidative DNA damage. Increasing the concentration of ZEN increased the extent of DNA damage. The extent of DNA migration, and percentage of cells with tails were significantly increased in a concentration-dependent manner following treatment with ZEN toxin (p < 0.05). Treatment with a low concentration of ZEN toxin (25 microM) induced a relatively low level of DNA damage, compared to treatment of cells with a high concentration of ZEN toxin (250 microM). Oxidative DNA damage appeared to be a key determinant of ZEN-induced toxicity in Chang liver cells. Significant reductions in cytolethality and oxidative DNA damage were observed when cells were pretreated with NACA prior to exposure to any concentration of ZEN. Our data suggest that ZEN induces DNA damage in Chang liver cells, and that the antioxidant activity of NACA may contribute to the reduction of ZEN-induced DNA damage and cytotoxicity via elimination of oxidative stress.
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Humans , Acetylcysteine , Animals, Domestic , Cell Proliferation , Edible Grain , Comet Assay , DNA , DNA Damage , Electrophoresis , Estrogens , Fusarium , Liver , Mycotoxicosis , Oxidative Stress , ZearalenoneABSTRACT
The plant Piper cubeba is widely distributed in tropical and subtropical regions and is used medically for various purposes but has not yet been evaluated for genotoxicity. We used male and female Swiss mice and Wistar rats and the comet assay and micronucleus test to investigate the mutagenic potential of a crude extract of P. cubeba seeds. The rodents were administered 0.5 g kg-1, 1.0 g kg-1 and 1.5 g kg-1 of the extract by gavage. For the Swiss mice, peripheral blood was collected 24 h after treatment for the comet assay, and at 48 and 72 h for the micronucleus test. For the Wistar rats, peripheral blood and hepatic cells were collected for the comet assay and bone marrow cells were collected for the micronucleus test 24 h after treatment. At 1.5 g kg-1, the highest dose tested, the extract induced a statistically significant increase in both the mean number of micronucleated polychromatic erythrocytes and the level of DNA damage in the rodent cell types analyzed. Under our experimental conditions, the P. cubeba seed extract was genotoxic in vivo when administered orally to mice and rats.
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Objective To detect nuclear DNA degradation of bone marrows and brains in rat cadavers at different temperatures,and develop a new parameter for estimating early postmortem interval(PMI).Methods The brain and bone marrow were taken out for every 4h,during 0~40h after death at 10℃ and 20℃,respectively.And the single cell gel electrophoresis(SCGE) was carried out to detect the nuclear DNA degradation.Linear regression analysis was used to assay the relationship of the comet parameter HeadDNA%,Tail Length(TL) and Olive TailMoment(TM) with PMI.Results Different decline degrees of comet HeadDNA% were found in both brain cells and bone marrow cells after death,the decline of HeadDNA% in brain cells at 20℃ was faster.Compared with degradation in marrow cells,the linear relation between degradation of brain cells and PMI was better.Conclusion with that of comet parameters TL and TM,the perfect linear relationship between HeadDNA% and PMI was also observed.Conclusion Brain tissues are more suitable for PMI estimation by detecting degradation of DNA with SCGE.The HeadDNA% is more valuable for PMI estimation than TL and TM.
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Objective To detect DNA damage in rat lymphocytes and brain cells induced by tetramine and study the toxicological mechanism of tetramine.Methods Lymphocytes and brain cells were separated and collected from healthy rats.DNA damages of cells which were exposed to various doses of tetramine for 60 min was detected using the single cell gel electrophoresis (SCGE) or comet assay.Results These are different degree DNA damages of lymphocytes and brain cells exposed from doses 1/20 LD 50 doses of tetramine to 1/2 LD 50 doses of tetramine.The test groups are very significantly statistical different to the control group(P
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Objective:To study the genetic toxic effect of smoking on human oral mucosa,and the DNA damage of exfoliated oral mucosa cells.Methods:The DNA damage of exfoliated oral mucosa cells was investigated by single cell gel electrophoresis(SCGE)in 12 cases of malignant tumor,19 cases of benign tumor and 10 health controls.There were 24 smokers and 17 non-smokers among them.The tail length and frequency of comet cells were used to measure DNA damage.SPSS and SAS software were used for statistical analysis.Results:Malignant tumors had a longer tail length and higher frequency of comet cells than benign tumors and health controls,and the difference was statistically significant(P0.05).The DNA damage of exfoliated oral mucosa cells in smokers was more serious than that of non-smokers(P
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To elucidate arsenic-induced oxidative DNA damage, the genotoxicity of arsenic in human cells was comparatively studied with single cell gel electrophoresis (SCGE) assay in combination with the observation of the protective effects of dimethyl sulfoxide (DMSO) and catalase. Arsenic, at the concentration of 2.4 μM by coincubation for 24 hours, significantly induced DNA damage in HL60, a human promyelocytic leukemia cell line. In contrast, significant DNA damage was found in human mononucleocytes at the concentration of 4.8 μM or above. The cells were incubated separately with DMSO (12 mM/l), a well-known hydroxyl radical (OH-) scavenger, and catalase (1,300 U/ml), a hydrogen peroxide (H2O2) scavenger, for 6 hours and then further coincubated with various concentrations of arsenic for 24 hours at 37°C and 5% CO2. The findings showed that both DMSO and catalase significantly reduced the arsenic-induced tail moment, a parameter of total damaged DNA, in HL60 and mononucleocytes. Hence our findings indicate that arsenic, with micromolar concentrations, induces typical and various extents of DNA damage in human cells via reactive oxygen species in a dose-dependent manner.
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Humans , DNA Damage , Arsenic , Dimethyl SulfoxideABSTRACT
Objective To study H2O2-induced single strand breaks(SSB) formation in DNA and the reparation in NIH3T3 mouse fibroblasts by single cell gel electrophoresis technique(SCGE,Comet Assay) and to introduce SCGE.Methods The NIH3T3 mouse fibroblasts were incubated,and cell damage was induced by H2O2 and determined by SCGE.Results ① H2O2-induced damage class graph of the DNA Comet in the NIH3T3 mouse fibroblasts was established.② H2O2-induced SSB was H2O2 dose-dependent.③ The reparation of the damaged DNA was time-dependent and could be achieved 15min after incubation.The data indicated that most reparations were completed within 1 hour,but a few might take longer time.Conclusion Single cell gel electrophoresis technique is a simple and sensitive method to determine oxidation-induced single strand breaks(SSB) formation in DNA.
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Objective: The study was to exlpore the nutritional, antioxidative functions and toxicology of ascorbic acid in different levels in vitro. [WT5FZ]Methods: Hela (human transformed epithelial) cells incubated with three levels of ascorbic acid, i.e. 0.1 mmol/L, 0.25 mmol/L and 0.5 mmol/L, were calculated and a single cell gel electrophoresis (SCGE) was used for measuring DNA oxidative damage. [WT5FZ]Results: The results showed that there were no differences in spontaneous DNA damage of Hela cells incubated with three levels of ascorbic acid. However, there was a less DNA oxidative damage induced by H 2O 2 in 0.1 mmol/L and 0.25 mmol/L of ascorbic acid supplemented groups respectively than in control group. In contrast, more serious DNA damage was found in 0.5 mmol/L ascorbic acid supplemented group. [WT5FZ]Conclusion: It is suggested that the higher levels of ascorbic acid might not directly damage DNA; the moderate supplementation of ascorbic acid may increase antioxidative ability of cells; excess ascorbic acid is harmful to DNA and enhances the susceptibility to H 2O 2 potentially.
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Objective: To study the effect of dietary nucleotides on DNA damage of thymocytes in mice. Methods: KM mice (n=30) , 5-6 weeks, were randomized into 3 groups,negative group (group 1), positive group (group 2) and nucleotides group (group 3). Mice in group 1 and 2 were fed nucleotide-free diet and group 3 nucleotide-supplemented diet (0.25% nucleotides). Mice in group 2, 3 were given single cyclophosphamide intraperitoneal injection of 150 mg/kg bw at day 21. DNA damage in thymus lymphocytes was evaluated 18 h after injection by single-cell gel electrophoresis (SCGE) assay. Thymus and spleen were weighed. Results: Dietary nucleotides had no effect on weights of thymus and spleen, but significantly decreased the number of damaged lymphocytes and the degree of damage. Conclusion: Dietary nucleotides may protect thymocytes DNA in mice from damage.