摘要
目的 探讨人端粒酶蛋白催化活性亚单位 (Humantelomerasereversetranscriptase ,hTRT)反义基因转染对HepG2 肝癌细胞株端粒酶亚单位及细胞周期的影响。方法 采用流式细胞仪检测hTRT正义基因转染的HepG2 细胞(HepG2 S)以及hTRT反义基因转染的HepG2 细胞 (HepG2 AS)的细胞周期 ,采用RT PCR法检测上述细胞hTRT、端粒酶RNA(hTR)、端粒酶相关蛋白 1(TP1)以及c myc和bcl 2基因的变化 ,同时采用Westernblot检测hTRT蛋白质的变化。结果 HepG2 AS出现G0 G1 期阻滞 ,增殖指数增加 ,凋亡率亦明显增加 ,进一步研究发现 ,HepG2 AS端粒酶亚单位hTRT在蛋白质和mRNA水平均表达减少 ,而TP1、hTR无明显变化 ,bcl 2和c mycmRNA的表达亦明显下调。结论 hTRT反义基因不仅可以下调HepG2 细胞hTRTmRNA和蛋白质的表达 ,而且可以下调c myc、bcl 2mRNA的表达 ,从而一方面降低端粒酶活性 ,另一方面一定程度增加凋亡细胞的比率 。
Objective To explore the effect of antisense gene to human telomerase reverse transcriptase(hTRT) on telomerase subunits and cell cycle of HepG 2. Methods Cell cycles in HepG 2, HepG 2 S and HepG 2 AS were examined by flow cytometry. Telomerase subunits(hTR, hTRT, TP1), c myc and bcl 2 mRNA were detected by RT PCR. Meanwhile, protein in transfected and untransfected cells was determined by Western blotting. Results Flow cytometric analysis displayed an accumulation of cells in the G 0/G 1 phase of cell cycle and a decreased percentage of proliferative index after transfection with antisense gene of hTRT. Further studies showed expression levels of hTRT, bcl 2 and c myc were down regulated in HepG 2 AS, but hTR and TP1 remained unchanged in all kinds of cells. Conclusion hTRT antisense gene can down regulate the expression levels of hTRT mRNA and protein as well as the expression levels of c myc and bcl 2 mRNA in HepG 2 cell, suggesting that hTRT antisense gene can not only decrease the telomerase activity but also partially induce apoptosis in cancer cells.
出处
《第三军医大学学报》
CAS
CSCD
北大核心
2003年第24期2193-2195,共3页
Journal of Third Military Medical University
关键词
人端粒酶蛋白催化活性亚单位
反义基因治疗
肝癌
human telomerase reverse transcriptase (hTRT)
antisense gene therapy
hepatocellular carcinoma cell line
