bcl-2硫代反义寡核苷酸对Raji细胞增殖和凋亡的影响

Effects of bcl-2 antisense oligodeoxynucleotide on proliferation and apoptosis of Raji cells

目的 研究bcl 2硫代反义寡核苷酸 (bcl 2ASODN)对淋巴系统恶性肿瘤的作用及机制。方法 将bcl 2ASODN与Raji细胞共孵育 ,应用MTT检测、电镜超微结构、流式细胞仪 (FCM)分析和RT PCR方法 ,观察bcl 2ASODN对Raji细胞增殖及凋亡的影响 ,以及bcl 2蛋白及mRNA水平的变化。结果 MTT检测显示bcl 2ASODN可以部分抑制Raji细胞增殖 ;经ASODN作用 4 8h ,电镜超微结构观察细胞呈现典型凋亡特征 ,包括胞膜出芽、异染色质核膜下浓聚形成凋亡小体及胞核碎裂 ;FCM检测AnnexinⅤ + PI- 的凋亡细胞比例升高。 2 0 μmol Lbcl 2ASODN处理Raji细胞 72h ,细胞凋亡率达4 3 86 % ,比对照组 (10 .0 5 % )明显增高。bcl 2阳性细胞率逐渐下降 ,于 72h达低谷为 0 88% ,明显低于对照组 (79.5 4 % )。bcl 2mRNA水平亦显著下降。结论 bcl 2ASODN通过下调bcl 2蛋白表达水平 ,诱导Raji细胞凋亡。

Objective To study the in vitro antitumor activity of bcl 2 fully phosporothioated antisense oligodeoxynucleotide (bcl 2 ASODN) to malignant lymphoblastic cells. Methods Proliferation and apoptosis of Raji cells incubated with bcl 2 ASODN were evaluated by MTT assay, flow cytometry (FCM) and electron microscopy, and the level of bcl 2 protein and mRNA expression were assessed by FCM and RT PCR, respectively. Results MTT assay demonstrated that bcl 2 ASODN could partially inhibit the growth of Raji cells. After incubated with ASODN for 48 hours, Raji cells exhibited characteristic morphologic changes of apoptosis, including cytoplasm membrane blebbing, chromatin condensation crescents formation and nuclear fragmentation. The apoptosis rate of Raji cells treated with 20 μmol/L bcl 2 ASON for 72 hrs was 43.86% which is significantly higher than that of control (10.05%). The bcl 2 ASODN induced apoptosis of Raji cells was accompanied by declined expression of bcl 2 mRNA , which decreased to 0.88% at 72 hrs and was significantly lower than that of control(79.54%). Conclusion bcl 2 ASODN induced Raji cells apoptosis by downregulating bcl 2 protein.