bcl-2反义寡核苷酸增强足叶乙苷诱导原代白血病细胞凋亡的研究

bcl-2 ANTISENSE OLIGONUCLEOTIDE ENHANCES ETOPOSIDE-INDUCED APOPTOSIS IN AL CELLS

目的 :研究 bcl- 2的反义寡核苷酸对足叶乙苷 (VP1 6 )诱导原代培养的急性白血病(AL)细胞凋亡的影响。方法 :用细胞计数法观察细胞的生存情况 ;用免疫荧光标记法通过流式细胞仪观测细胞 bcl- 2蛋白水平 ;用形态学观察及流式细胞仪检测细胞凋亡。结果 :靶向 bcl- 2 m RNA翻译起始区与靶向蛋白编码区的两个反义寡核苷酸分别与 VP1 6 联合作用 AL 细胞 4 8h,细胞的生存受到明显的抑制 ,分别同无关寡核苷酸 (NS- ODN)联合 VP1 6 组、单用 VP1 6 组进行比较 ,差异有显著性 (P<0 .0 5 )。这两个不同靶点的反义寡核苷酸分别与 VP1 6 联用 ,均能明显下调 AL 细胞 bcl- 2蛋白的表达 ,并且联合作用 AL 细胞 4 8h的凋亡细胞百分率分别与 NS- ODN联合 VP1 6 组、单用 VP1 6 组进行比较 ,差异有显著性 (P<0 .0 5 )。结论 :针对 bcl- 2 m RNA翻译起始区和蛋白编码区两个靶点的反义寡核苷酸能增强 VP1 6 诱导急性白血病细胞的凋亡。

Objective:To investigate whether bcl 2 antisense oligonucleotide increase etoposide (VP 16 ) induced apoptosis in cultured primary acute leukemia (AL) cells.Methods:Cell surviving fraction was determined using the trypan blue dye exclusion assay. The expression levels of bcl 2 protein were assayed by immunofluorescence using fluoresce isothiocyanate label. Apoptosis was detected by morphological observation and flow cytometric analysis.Results:Treatment with two oligonucleotides directed against the coding region and the translation initiation region of bcl 2 messenger RNA/VP 16 combination respectively for 48 h had significantly reduced the number of viable AL cells. However, there was no difference on AL survival between nosense oligodeoxynucleotide/VP 16 combination and VP 16 treated cells alone. It was found that the two antisense oligonucleotides combined respectively with VP 16 could inhibit expression of bcl 2 protein, increase apoptosis in AL cells, significantly ( P <0.05).Conclusion:The two bcl 2 antisense sequences could enhance the VP 16 induced apoptosis of AL cells in sequence specific manner.