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目的:探讨三氧化二砷(Arsenictrioxide,As2O3)诱导尤文肉瘤细胞凋亡及对融合蛋白EWS-FLi1表达的影响。方法:应用噻唑蓝(MTT)法、形态学观察、原位末端标记法(TUNEL)、流式细胞术(FCM)观察As2O3对体外生长的尤文肉瘤RD-ES细胞系生物行为的影响;应用半定量RT-PCR测定应用As2O3前后c-myc基因mRNA水平的变化;应用免疫细胞化学及固定化蛋白印迹法(Westernblot)观察用药前后EWS-FLi1融合蛋白表达水平的变化。结果:As2O3对体外生长的RD-ES细胞系具有明显抑制作用,并可诱导细胞凋亡。c-myc基因mRNA表达水平和EWS-FLi1融合蛋白表达量随As2O3作用时间延长逐渐降低。结论:常规治疗浓度的As2O3对体外生长的尤文肉瘤细胞具有明显的杀伤作用,其作用机制与改变线粒体膜通透性和抑制EWS-Fli1融合蛋白、降低c-myc基因表达有关。
Objective: To investigate the apoptosis of Ewing sarcoma cells induced by Arsenictrioxide (As2O3) and its effect on the expression of fusion protein EWS-FLi1. Methods: The effects of As2O3 on the biological behavior of RD-ES cell lines in vitro were observed by MTT, morphology, TUNEL and FCM. The level of c-myc mRNA was detected by quantitative RT-PCR. The expression of EWS-FLi1 fusion protein was detected by immunocytochemistry and Western blotting. Results: As2O3 inhibited the growth of RD-ES cells in vitro and induced apoptosis. The expression of c-myc mRNA and the expression of EWS-FLi1 fusion protein gradually decreased with the prolongation of As2O3. CONCLUSION: As2O3 treatment with conventional concentration has a significant killing effect on Ewing’s sarcoma cells in vitro. Its mechanism is related to the change of mitochondrial membrane permeability, inhibition of EWS-Fli1 fusion protein and reduction of c-myc gene expression.