hTERT反义寡核苷酸对HL-60细胞凋亡及细胞周期的影响

目的：研究hTERT基因反义寡核苷酸（ASODN）对HL-60细胞诱导凋亡作用及细胞周期的影响.方法：应用hTERT ASODN封闭HL-60细胞hTERT基因,RT-PCR方法检测目的基因的表达;流式细胞仪检测细胞凋亡及细胞周期分析,TUNEL方法检测细胞凋亡,琼脂糖凝胶电泳检测凋亡细胞DNA梯带.结果：hTERT ASODN作用细胞72h后,hTERT基因表达明显受到抑制（P＜0.01）.流式细胞仪检测显示：ASODN组细胞阻滞于G0/G1期,S、G2/M期细胞减少（P＜0.05）;细胞增殖指数（PI）明显降低（P＜0.05）;检测出早期凋亡峰.Annexin V/PI检测及TUNEL检测均显示：ASODN组凋亡细胞阳性率明显高于SODN组（P＜0.01）.琼脂糖凝胶电泳显示：ASODN组出现凋亡DNA梯带.结论：hTERT ASODN能够封闭目的基因表达,阻滞HL-60细胞于G0/G1期,并诱导细胞凋亡,对治疗白血病具有潜在应用价值.

Effects of hTERT ASODN on the Induction of Apoptosis and the Cell Cycle of HL-60 Cells

Objective:To investigate the effects of hTERT antisense oligodeoxynucleotide(ASODN)on the cell cycle and the induction of apoptosis in HL-60 cells.Methods:After treatment with hTERT ASODN the expression of hTERT was assessed by RT-PCR and agarose gel electrophoresis,apoptosis was detected with TUNEL,and cell cycle ratios were determined using flow cytometry(FCM).Results:After 72h of treatment with hTERT ASODN,expression of the hTERT gene was inhibited significantly(P<0.01).Analysis of the FCM data revealed that the ASODN treated cells were blocked at G0/G1 phase and that the number of cells in S phase and G2/M phase was reduced(P<0.05).The proliferation index of the ASODN-treated group was markedly decreased(P<0.05).In the ASODN-treated group the DNA ladder and early apoptosis peak appeared when samples were analyzed by agarose gel electrophoresis and FCM,respectively.Both Annexin V/PI FCM data and TUNEL results indicated that the number of apoptotic cells in the ASODN-treated group was much higher than that found in the SODN-treated group(P<0.01).Conclusion:The hTERT ASODN inhibits expression of the hTERT gene,blocks HL-60 cells at G0/G1 phase,induces apoptosis and has a potential therapeutic effect for leukemia.