| Glyoxal is an a - keto aldehyde, which can synthesize glyoxalic acid. It is extensively used in industry such as paper making, spin, pharmacy, dyestuff manufacture, and so on. It is an intermediate product of the synthetic resin. It is found in ozonated drinking water, ozonated products of atmospheric aromatic hydrocarbons, heated glucose and coffee as well as cigarette. In the past it has been thought that the toxicity of glyoxal is low and mainly associated with simulation in the eyes and the mucosa of the respiratory tract. It has also been reported that glyoxal has cytotoxicity. In recent years it has been found that glyoxal has mutagenicity and carcinogenicity. In addition, it has a close relationship with lipid peroxidation, and it can induce the formation of active oxygen free radicals. In the present study we determined the effect of different concentrations of glyoxal on the activity of the lymphocytes in rat by using AlamarBlue?assay. The genotoxicity of glyoxal was studied through the detection of the effect on DNA damage in rats by exposure to the agent in vivo and in vitro by using single cell gel electrophoresis assay ( SCGE). This investigation may provide experimental basis for the mechanism of toxicity of glyoxal.Methods1. Measurement of activity of lymphocytesLymphocytes were isolated and suspended by RPMI1640, then plated at a density of 1 x 10 cells/ml in 96 - well flatbottom plates. After being treated with different levels (0, 31.25, 62.5, 125, 250, 500, 1000 and 2000jxmol/ L) of glyoxal, the cell suspensions were incubated at 37℃, 5% C02 for 24 hours. At the twentieth hour of incubation, AlamarBlue was added to each well.cell suspensions were then reincubated for four hours. Read absorbance of each well at 570nm and 620nm wavelengths respectively.2. Determination of the DNA damage2.1 In vivo exposure of ratsA total of 20 male Wistar rats, weighing (200 10) g , were divided into 4 groups evenly on random. There were 5 rats in each group. The experimental animals were administrated with aqueous solution of glyoxal at different concentrations by intraperitoneal injection at the dose of 12. 5, 25 and 50mg/kg body weight. Control animals were injected the same volume of saline. Took blood from the rats after they were treated with glyoxal for 16 hours. The lymphocytes were isolated and then used single cell gel electrophoresis assay to detect the DNA damage.2.2 In vitro exposure of lymphocytesIsolated lymphocytes and regulated the density for 3 x 106 cells/ml. After being treated with different levels (0, 0. 25 , 0. 5 and 1mmol/L) of glyoxal, the cell suspensions were incubated at 37 for 1 hour. Then the DNA lesion was determined by single cell gel electrophoresis assay.3. Statistical analysis of dataThe data were analyzed by using SPSS 10. 0 Software. The chi - square test and One - way ANOVA were used to test the significance.Results1. The effect of glyoxal on the activity of lymphocytesAfter being treated with glyoxal for 24 hours, there was no effect on activity of lymphocytes at the dose of 31. 25 and 62. 5mol/L. The activity of lymphocytes was significantly decreased at the dose of 125 ~2000mol/L.2. DNA damage of lymphocytes by glyoxal 2.1 SCGE imageThe peripheral blood lymphocytes of rats was stained for orange. The normal cell was round and the texture was symmetrical and compact. When lymphocytes were damaged by being treated with certain levels of glyoxal, the DNAfragments migrated to the positive pole under alkaline condition and then shaped into tail ( comet image).2. 2 DNA damage in rat lymphocytes following in vivo exposure DNA lesions were detected in lymphocytes of the dose of 25 and 50mg/kg body weight. And the percent of comet cell and the length of comet tail were higher than control group. There was a significant difference between the experimental groups.2. 3 DNA damage in rat lymphocytes following in vitro exposure Exposure of lymphocytes to 0.5 and Immol/L glyoxal resulted in DN...
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