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目的 :设计靶向端粒酶逆转录酶(telomerase reverse transcriptase,h TERT)的mi RNA表达框架,探讨其对h TERT的表达、端粒酶活性及K562增殖的影响,以期为慢性髓系白血病提供新的基因治疗策略。方法:构建靶向h TERT的mi RNA表达框架,脂质体法转染K562细胞,银染法及荧光定量PCR法检测端粒酶活性及h TERT的表达情况。Annexin V/PI双染检测细胞凋亡。结果:mi RNA表达框架转染48 h后,h TERT的表达明显下降,端粒酶活性明显减低,K562细胞凋亡率增加。结论:靶向h TERT的mi RNA表达框架能够有效干扰h TERT的表达,抑制端粒酶活性,诱导K562细胞的凋亡。以mi RNA表达框架为基础的RNAi技术,有望成为分子水平治疗慢性髓系白血病的有效工具。
OBJECTIVE: To design a mi RNA expression framework targeting telomerase reverse transcriptase (h TERT), and to investigate its effect on the expression of h TERT, telomerase activity and K562 proliferation in order to provide a novel therapeutic strategy for chronic myeloid leukemia New gene therapy strategy. Methods: The miRNA targeting hTERT was constructed and transfected into K562 cells by lipofectamine 2000. The telomerase activity and hTERT expression were detected by silver staining and fluorescence quantitative PCR. Annexin V / PI double staining was used to detect apoptosis. Results: After 48 h of transfection, the expression of h TERT was significantly decreased, the telomerase activity was significantly decreased, and the apoptosis rate of K562 cells was increased. CONCLUSION: The miRNA expression vector targeting h TERT can effectively interfere with the expression of h TERT, inhibit telomerase activity and induce apoptosis in K562 cells. RNAi technology based on the mi RNA expression framework is expected to be an effective tool to treat chronic myelogenous leukemia at the molecular level.