【摘要】 目的探讨人端粒酶核糖核酸(humantelomeraseribonucleicacid,hTR)反义寡核苷酸(antisenseoligodeoxynucleotides,ASODN)对顺铂诱导原代白血病细胞凋亡的影响。方法采用与hTR模板互补的硫代ASODN处理原代白血病细胞,采用端粒酶重复扩增分析聚合酶链反应酶联免疫分析法检测端粒酶活性的变化;采用苔盼蓝拒染法观察经hTRASODN处理的原代白血病细胞对顺铂的反应;采用AnnexinV与碘化丙啶双染色观察凋亡细胞形态及用流式细胞仪检测凋亡细胞率。结果经hTRASODN作用后,原代白血病细胞端粒酶活性明显下降。ASODN作用24小时,再加入顺铂作用48小时,明显抑制原代白血病细胞,并使原代白血病细胞凋亡明显增加。ASODN作用24小时后再加入5μmol/L顺铂作用72小时的原代白血病细胞凋亡率为(41±9)%,分别高于正义寡核苷酸加顺铂组[(15±5)%]及单用顺铂组[(13±3)%],差异有统计学意义(均为P<001)。结论hTRASODN可明显抑制原代白血病细胞端粒酶活性,并增强顺铂诱导原代白血病细胞的凋亡,人端粒酶有可能成为白血病基因治疗的新靶点。更多还原

【Abstract】 Objective: To explore whether the antisense oligodeoxynucleotides (ASODN) of human telomerase ribonucleic acid (hTR) can enhance cisplatin-induced apoptosis of cultured primary leukemia cells. Methods:Telomerase activity was assayed by telomerase repeat amplification protocol and polymerase chain reaction and enzyme-linked immunoassay. Cell surviving fraction was determined by using the trypan blue dye exclusion assay. Apoptosis was assayed by morphological methods (Annexin V & PI) and flow cytometry analysis technology. Results:Telomerase activity of cultured primary leukemia cells was suppressed significantly by hTR ASODN. Treatment with cisplatin after 24h of exposure to hTR ASODN reduced the number of cultured primary leukemia cells significantly. Apoptosis rates of primary leukemia cells treated with cisplatin for 72h after 24h of exposure to hTR ASODN were(41±9)~%. There was significant difference in the percentage of apoptotic cells between hTR ASODN plus cisplatin and sense oligodeoxynucleotides plus cisplatin[(15±5)%] or cisplatin alone group [(13±3)%] (P<0.01). Conclusion:hTR ASODN can significantly suppress the telomerase activity and enhance cisplatin-induced apoptosis of primary leukemia cells, indicating that human telomerase can be a new target of gene therapy for leukemia. [更多还原