反义寡核苷酸对K562细胞的抑制作用及诱导细胞凋亡的研究

THE STUDY OF INHIBITORY AND APOPTOSIS OF K562 CELLS BY ANTISENSE OLIGODEOXYNUCLEOTIDES

目的 :研究bcr abl硫代磷酸反义寡脱氧核糖核酸 (Aspo)作用于K5 62细胞后 ,对细胞mRNA、蛋白水平的影响 ,以及诱导细胞凋亡情况。方法 :Aspo与K5 62细胞共培养后 ,用流式细胞仪检测P2 10蛋白表达及细胞凋亡率 ,RT PCR半定量检测bcr ablmRNA表达情况 ,电镜观察细胞凋亡的形态学改变。结果 :K5 62细胞经浓度大于 5 μmol Lbcr ablAspo处理 2 4h ,流式细胞仪检测细胞P2 10蛋白表达下调甚至完全受抑制 ,10 μmol Lbcr ablAspo作用 48h ,细胞bcr ablmRNA下降 45 %左右 ,当细胞初始浓度为 1× 10 4 ml时 ,Aspo作用 12 0h细胞凋亡率 2 0 %～ 30 % ,当细胞数增加至 1× 10 5 ml时 ,Aspo作用 48h即可使 30 %细胞发生凋亡 ,电镜下观察到典型凋亡细胞形态学改变。结论 :bcr abl反义核酸对K5 62细胞mRNA水平具有抑制作用 。

Aim:To study the effect of bcr abl fusion gene antisense phosphorothioate oligodeoxynucleotide (Aspo) on bcr abl mRNA and apoptosis of K562 cells.Methods: Cells were exposed to Aspo.P210 was measured by Flow Cytometry.Cellular bcr abl mRNA was detected by RT PCR mediquantitive analysis.Cell apoptosis was measured by Flow Cytometry and observed by electron microscope (EM).Results:The P210 was down regulated or completely suppressed after 24h treatment with more than 5 μmol/L Aspo.The decrease of bcr abl mRNA was about 45%.After incubation 48 h with 10 μmol/L Aspo.Also,20%～30% K562 cells were induced to apoptosis at 120 h when the cell number was 1×10 4/ml at the beginning. While the cell number was 1×10 5/ml,the apoptosis rate was 30% after 48 h culture and the typical morphology of apoptosis cell was observed under EM.Conclusion:bcr abl Aspo could inhibit the expression of bcr abl mRNA and P210.Also,it could induce apoptosis of K562 cells.