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目的本研究采用针对人端粒酶hTR基因的反义寡聚脱氧核苷酸,探讨端粒酶反义寡聚脱氧核苷酸(antisense oligodeoxy nucleotides,ASODN)对脑胶质瘤HO-8910细胞端粒酶活性及细胞增殖的影响。方法本实验将互补于hTR模板区的全硫代修饰型反义寡核苷酸(PS-ASODN)经lipotap脂质体转染作用于脑胶质瘤细胞系C6,然后分别采用四甲基偶氮唑蓝(MTT)比色实验、酶联免疫吸附试验(ELISA)法和流式细胞术检测C6脑胶质瘤细胞的体外增殖、端粒酶活性、细胞凋亡和细胞周期的改变。结果经MTT法检测,PS-ASODN对C6细胞生长具有明显的抑制作用和诱导细胞凋亡,1.0μMPS-ASODN处理后24h细胞生长抑制率为12.93%,且随着时间的延长,抑制作用逐渐增强;给药72h对细胞生长抑制率达到最高,为50.81%,有相应的浓度依赖关系。ELISA法检测结果 ,0.5～1.5μM的PS-ASODN对C6细胞作用72h后端粒酶皆为阴性,表明PS-ASODN能够抑制端粒酶活性,对照组端粒酶皆为阳性。结论端粒酶PS-ASODN对体外培养的脑胶质瘤C6细胞的增殖有明显的浓度、时间依赖性抑制作用;抑制脑胶质瘤C6的端粒酶活性和诱导C6脑胶质瘤细胞发生凋亡,可能是端粒酶PS-ASODN抑制C6细胞增殖的主要机制。
Objective To investigate the effect of antisense oligodeoxy nucleotides (ASODN) on the telomerase activity of human glioma HO-8910 cells by using antisense oligodeoxynucleotides targeting human telomerase hTR gene Granzyme Activity and Cell Proliferation. Methods In this study, the full-thio-modified antisense oligonucleotide (PS-ASODN) complementary to hTR template region was transfected into glioma cell C6 by lipotap liposome. MTT assay, enzyme-linked immunosorbent assay (ELISA) and flow cytometry were used to detect C6 glioma cells in vitro proliferation, telomerase activity, apoptosis and cell cycle changes. Results The results of MTT assay showed that PS-ASODN significantly inhibited the growth of C6 cells and induced apoptosis. The inhibition rate of PS-ASODN was 12.93% after treated with 1.0μMPS-ASODN for 24h, and the inhibitory effect was gradually enhanced with the prolongation of time ; 72h on the cell growth inhibition rate reached the highest, was 50.81%, a corresponding concentration-dependent relationship. ELISA results showed that telomerase activity of PS-ASODN of 0.5-1.5μM was negative after 72h, indicating that telomerase activity was inhibited by PS-ASODN and telomerase activity of control group was positive. CONCLUSION: Telomerase PS-ASODN inhibits the proliferation of C6 glioma cells in vitro in a concentration-dependent and time-dependent manner, inhibits the telomerase activity of C6 glioma cells and induces C6 glioma cells Apoptosis may be the main mechanism by which telomerase PS-ASODN inhibits the proliferation of C6 cells.