反义寡核苷酸对淋巴瘤细胞Livin基因表达及细胞凋亡的影响

Effects of antisense oligodeoxynucleotide on the expression of livin gene and the apoptosis of lymphoma cell

目的分析凋亡抑制蛋白Livin基因的反义寡核苷酸(ASODN)对人淋巴瘤细胞株(Raji)Livin mRNA表达的抑制作用及其对细胞凋亡的影响。方法合成Livin特异性全硫代修饰的ASODN及其对照错义寡核苷酸(MSODN),通过脂质体转染Raji细胞,MTT法检测细胞的增殖抑制率;RT-PCR法检测转染前后Livin mRNA表达水平的变化;Hoechst染色观察细胞凋亡的形态学改变;流式细胞仪检测细胞的凋亡率。结果 Livin ASODN能显著抑制Raji细胞的增殖(P<0.01),并呈现一定的剂量效应关系。0.6μmol/L的ASODN作用Raji细胞48h后,Livin mRNA的表达水平明显下调,细胞出现凋亡的形态学改变,细胞凋亡率为(37.54±2.65)%,与对照组比较,差异有统计学意义(P<0.01)。结论 Livin ASODN能有效降低Livin mRNA表达水平,抑制Raji细胞增殖,并增强细胞凋亡的敏感性,有望成为淋巴瘤基因治疗的分子靶向。

【Objective】This study was intended to explore the effects of antisense oligodeoxynucleotide（ASODN） on the inhibition of Livin gene expression and apoptosis of human lymphoma cell line（Raji）.【Methods】Livin specific phosphorothioate antisense oligodeoxynucleotide and missense oligodeoxynucleotide were synthesized and transfected into Raji cells with liposome.MTT assay was used to detect the proliferation inhibition rate of Raji cells.The expression level of Livin mRNA in Raji cells was detected by RT-PCR.The morphologic changes of cell apoptosis were observed by Hoechst fluorescein staining.Apoptosis rate of Raji cells was examined by flow cytometer.【Results】Livin ASODN obviously inhibited the proliferation of Raji cells in a dose-dependent manner（P 0.01）.After transfection of 0.6 μmol/L ASODN,the expression of Livin mRNA was decreased and the apoptosis rate was（37.54± 2.65）%.The differences were significant when compared with the other control groups（P 0.01）.【Conclusion】Livin ASODN remarkably down-regulates the expression of Livin gene and supresses the proliferation and induces the apoptosis of Raji cells,which may be a molecular target for gene therapy of lymphoma.