survivin反义寡核苷酸抑制HL-60细胞增殖并诱导细胞凋亡

Induction of apoptosis and inhibition of HL-60 cell proliferation by survivin antisense oligonucleotide

摘要:

目的：探讨survivin反义寡核苷酸(ODN)对人白血病细胞系HL-60细胞增殖和凋亡的影响。方法：人工合成survivin基因反义和正义ODN,并进行硫代磷酸化修饰,通过脂质体途径分别转染HL-60细胞;应用RT-PCR和W estern B lot检测survivin mRNA和蛋白表达;应用MTT法检测survivin反义ODN对HL-60细胞增殖的影响;流式细胞仪检测细胞周期变化及细胞凋亡比率。结果：体外培养的HL-60细胞可表达较强的survivinmRNA和蛋白;survivin反义ODN可呈浓度依赖性地抑制survivin mRNA和蛋白表达,1 000 ng/mL AS-ODN几乎完全抑制survivin mRNA和蛋白表达。MTT研究结果表明,survivin反义ODN可呈浓度依赖性地抑制HL-60细胞增殖,1 000 ng/mL AS-ODN对细胞生长的抑制率可达78.14%;诱导细胞凋亡,使细胞阻滞于G2/M期。survivin正义ODN对survivin mRNA和蛋白以及HL-60细胞的增殖和细胞周期无明显的抑制作用。结论：脂质体介导转染survivin反义寡核苷酸可抑制细胞增殖、诱导细胞发生G2/M期阻滞而促进细胞凋亡。

Abstract:

OBJECTIVE: To investigate the effects of survivn antisense oligonucleotide(ODN) on cell proliferation and apoptosis of HL-60 cells. METHODS: Synthetic ODN was completely phosphorothioate-modified. Cationic lipid-mediated antisense ODN was transferred into HL-60 cells. The expression of survivin mRNA and protein was detected by RT-PCR and Western Blot.The incorporation of MTT was used as the measurement of HL-60 proliferation. The cell-cycle and apoptosis were analyzed by flow cytometry. RESULTS: HL-60 cells spontaneously expressed survivin mRNA and protein. Both mRNA and protein expression of survivin decreased significantly in the antisense ODN transfected cells in comparison to that in the original cells and cells transfected with sense ODN.Survivin antisense ODN significantly inhibited cell proliferation and induced apoptosis in a dose-dependent manner. The cell-cycle in the antisense ODN-transfected cells stopped at the G_2/M phase. CONCLUSIONS: Antisense ODN targeting at survivin mRNA can inhibit HL-60 cell proliferation and induce G_2/M stop and apoptosis.