特异性人端粒酶催化亚单位基因小干扰RNA对HeLa细胞生长的抑制作用

通过设计并化学合成人端粒酶催化亚单位(hTERT)特异性siRNA,观察其对hTERT表达水平及肿瘤细胞生长的影响。将hTERT-siRNA以脂质体法转染入HeLa细胞,应用RT-PCR、实时定量TRAP、Western印迹、软琼脂克隆形成实验、荷瘤裸鼠肿瘤内注射等方法检测细胞内hTERTmRNA、蛋白质表达水平及对肿瘤细胞生长的影响。RT-PCR、实时定量TRAP和Western印迹的结果显示hTERT-siRNA明显降低了HeLa细胞内hTERT的mRNA及蛋白质表达水平并伴随有端粒酶活性的下降。克隆形成实验表明hTERT-siRNA组的体外肿瘤形成能力受到抑制。荷瘤裸鼠肿瘤内注射hTERT-siRNA使肿瘤平均体积显著小于对照组。TUNEL凋亡检测表明hTERT-siRNA转染组的凋亡率明显高于对照组。研究表明hTERT特异性siRNA可以明显抑制HeLa细胞内hTERT的表达水平,对其生长有明显抑制作用,是一种有前途的肿瘤治疗新方法。

Growth Inhibition of Uterine Cervix Cancer Cells (HeLa) by an siRNA Targeting Human Telomerase Catalytic Subunit Gene

In this study, we aimed to observe the effect of chemically synthesized human telomerase catalytic subunit (hTERT)-oriented siRNA in cultured cell line and in a mouse xenograft model. Transfected hTERT- siRNA into HeLa cells, then using the RT-PCR, RQ TRAP, Western blot, soft agar colony forming assay, and intratumoral injection of hTERT-siRNA in a mouse xenograft model to investigate the level of hTERT mRNA and protein, the activity of telomerase, energy for growth of HeLa in vitro and in vivo. The result of RT-PCR, Western blot and RQ TRAP had showed that after transfected with hTERT-siRNA, the level of hTERT mRNA and protein decreased, and the activity of the telomerase was depressed obviously. The colony forming assay showed that energy for growth of hTERT-siRNA group was lower than that of the two control groups. The animal experiments showed that the cancer growth of hTERT-siRNA group was depressed. TUNEL assay showed an increased apoptosis rate in hTERT-siRNA group. Our study have showed that hTERT-siRNA can inhibit the expression of hTERT and depress the growth of the HeLa cells by inducing apoptosis in vitro and in vivo.