摘要
几乎所有类型的人类肿瘤都可以检测到端粒酶的活性,通过抑制端粒酶逆转录酶基因(TERT)表达降低端粒酶活性可能是进行抗肿瘤治疗的一个新靶点。本试验通过构建靶向端粒酶TERT基因的shRNA(Sh1,Sh2和Sh3)表达质粒载体转染MDCC-MSB1细胞抑制chTERT基因表达,从而降低端粒酶活性及细胞增殖。Real-time PCR结果显示,Sh1和Sh3在质粒载体转染细胞后48,72,96h显著抑制chTERT基因表达(P<0.05),且呈持续效果;TRAP法端粒酶活性分析显示,Sh1和Sh3在质粒转染细胞72h显著降低端粒酶活性;流式细胞分析显示,Sh3在转染后72h显著降低了S期细胞的比例(P<0.01),G2/M期细胞比例显著上升(P<0.01),G0/G1期细胞比例没有明显变化(P>0.05),抑制了MDCC-MSB1细胞的增殖。结果提示,shRNA通过抑制TERT基因表达可有效降低端粒酶活性,阻滞肿瘤细胞的增殖,为抗癌症治疗提供了新的备选方案及具有参考价值的基础科学数据。
Telomerase activity has been showed in almost human tumors,inhibition of telomerase activation by TFRT silencing may be a new target in anticancer therapeutics. Here,shRNAs （Shl, Sh2 and Sh3） expression plasmids targeting chTERT were constructed,and then transfected into MDCC-MSB1 cells. The expression level of chTERT was monitored by real-time PCR,telomerase activity were examined by TRAP assay,and cells proliferation were measured by flow cytometry. Our findings showed that the expression of chTERT was significantly inhibited by Shl and Sh3 at 48 h,72 h and 96 h post-transfection （P〈0.05）,Shl and Sh3 suppressed telomerase activation at 72 h post-transfection,and Sh3 inhibited MDCC-MSB1 cells proliferation by significantly decreasing numbers of cells in S phase （P〈0.01） ,increasing numbers of cells in G0/G1 phase （P〈0.01） at 72 h post-transfection, moreover, numbers of cells in G0/G1 phase had no change. These results indicated that chTERT silencing mediated by shRNA decreased telomerase activation,and inhibited MDCC-MSB1 cell proliferation,which provided a new alternative for anticancer therapeutics.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2017年第12期2350-2357,共8页
Chinese Journal of Veterinary Science
基金
国家自然科学基金资助项目(30471284)
