文章：

shRNA介导的hTERT沉默抑制骨肉瘤细胞增殖并促进细胞凋亡

shRNA-mediated silencing of hTERT suppresses proliferation and promotes apoptosis in osteosarcoma cells 

原文发布日期：2017-08-11 

英文摘要：

In this study, we aim to explore the effects of short hairpin RNAs (shRNAs) targeting human telomerase reverse transcriptase (hTERT) on the proliferation and apoptosis of osteosarcoma cells. After the synthesis of shRNA that target hTERT, osteosarcoma cells were assigned into three experimental groups—shRNA group, scramble group and blank group. The transcription and expressions of the hTERT gene in transfected cells were measured with quantitative real-time polymerase chain reaction and western blotting. Cell proliferation in each group was detected by Cell Counting Kit-8 assay. Cell cycle and rates of apoptosis were measured by flow cytometry. Expressions of apoptosis-related proteins, caspase-9 and caspase-3, were detected by western blotting. Telomerase activity was measured by PCR enzyme-linked immunosorbent assay. Results show that both the mRNA and protein expressions of hTERT were significantly lowered after the transfection of hTERT-shRNA. The proliferation capacity of transfected osteosarcoma MG-63, SaOS2 and U2OS cells in the shRNA group was lower than that in the blank group. We also found changes and differences in the amount of cells throughout the cell cycle. All cells in the G0/G1 phase increased in numbers, whereas the number of cells in the S phase were reduced, with elevated apoptosis rates. Expressions of apoptosis-related proteins, caspase-9 and caspase-3, were increased and telomerase activity was decreased in the transfected shRNA group (all P<0.05). Our results showed that shRNA targeting of the hTERT gene was able to inhibit cell proliferation and promote apoptosis of osteosarcoma cells by reducing the telomerase activity. 

摘要翻译： 

本研究旨在探讨靶向人端粒酶逆转录酶（hTERT）的短发夹RNA（shRNA）对骨肉瘤细胞增殖与凋亡的影响。通过合成靶向hTERT的shRNA后，将骨肉瘤细胞分为shRNA组、乱序序列组和空白组三组实验组。采用实时荧光定量PCR和蛋白质印迹法检测转染细胞中hTERT基因的转录及表达水平；通过CCK-8法检测各组细胞增殖能力；流式细胞术检测细胞周期与凋亡率；蛋白质印迹法检测凋亡相关蛋白caspase-9与caspase-3的表达；PCR酶联免疫吸附法测定端粒酶活性。结果显示：转染hTERT-shRNA后，hTERT的mRNA及蛋白表达均显著降低；shRNA组骨肉瘤MG-63、SaOS2和U2OS细胞的增殖能力均低于空白组；细胞周期各时相细胞数量出现变化与差异，G0/G1期细胞数量增加，S期细胞数量减少，且细胞凋亡率升高；shRNA组凋亡相关蛋白caspase-9与caspase-3表达升高，端粒酶活性降低（均P<0.05）。结果表明：靶向hTERT基因的shRNA可通过降低端粒酶活性，抑制骨肉瘤细胞增殖并促进其凋亡。

原文链接：

shRNA-mediated silencing of hTERT suppresses proliferation and promotes apoptosis in osteosarcoma cells