Study on DNA damage of retinal pigment epithelium cell induced by arsenic trioxide in rabbit

Abstract

Background Researches have reported that arsenic trioxide (As_2O_3) is an effective method of treating solid tumor,and its mechanism is to inhibit the cellular proliferation and induce cellular apoptosis.However,the research on DNA damage caused by As_2O_3 is not clear.Objective This study is to investigate the roles of arsenic trioxide (As_2O_3) on DNA damage of retinal pigment epithelial cell in rabbit.Methods Retinal pigment epithelial cells were isolated from pigmented rabbits and cultured in vitro according to the method of Singh NP[2].Cultured cells were identified by cytokeratin.The third or fourth generation of cells were used to this study.50.00,5.00,2.00,1.00,0.50 and 0.25 μmol/L of As_2O_3 were added into medium respectively for 1 hour,and the same volume of PBS was added as negative control group.Cultured cells were exposed to ultraviolet (UV) rays with wave length of 254 nm for 10 minutes as positive control group.The cell vitality was detected by trypan-blue staining.The comet assay was applied to evaluate the DNA damage.SPSS 13.0 software was used for statistical analysis.Results The black granule were seen in cultured RPE cells.Cultured cells showed the positive brown-yellow particles in cytoplasm for cytokeratin with the positive rate over 90%.The morphology of the cells was similar among experimental group,UV irradiation group and PBS group under the inverted microscope.The normal cell activity was exhibited by trypan-blue staining in these three groups.Compared with PBS group,As_2O_3 caused obvious abnormality of DNA of RPE cell in a dose-dependent manner.Both the percentage of tail DNA and tail moment were gradually increased with the enhance of As_2O_3 concentration (F=88.548,P=0.000; F= 129.704,P=0.000).Significant differences also were found between different concentrations of As_2O_3 groups and UV irradiation group compared with PBS group (P＜0.05).As_2O_3 caused the obvious breaking up of DNA strands of RPE cells at the concentration of 50.00 μmol/L,but there was no statistical difference in spontaneous DNA damage of RPE cells between 0.50 and 0.25 μmol/L of As_2O_3 (P＞0.05).Conclusion As_2O_3 has a potential genotoxicity for rabbit retinal pigment epithelial cells in vitro. Key words: retinal pigment epithelia; As2O3; arsenic trioxide DNA damage