摘要
目的探讨人端粒酶逆转录酶(hTERT)基因反义寡核苷酸对皮肤T细胞淋巴瘤(CTCL)细胞株端粒酶活性及细胞生长的影响,为CTCL基因治疗提供新的基因靶点。方法采用脂质体介导的基因转染方法,将不同浓度(10 μmol／L、20 μmol／L、30 μmol／L)的寡核苷酸分别导入CTCL细胞株Hut78中; 分别于不同的时间,应用端粒酶重复序列扩增及酶联免疫吸附方法(PCR-ELISA)、逆转录-聚合酶链反应 (RT-PCR)、流式细胞仪技术等动态观察转染细胞中端粒酶活性、hTERT mRNA表达以及细胞凋亡的变化。结果经hTERT反义寡核苷酸(AODN)作用72 h后,细胞端粒酶的活性明显下降,hTERT mRNA的表达减弱,并可观察到细胞凋亡,凋亡细胞发生率为13．05％。细胞生长抑制作用有明显的时效性,以 30μmol／L、72 h时下降最明显。而有义寡核苷酸(SODN)组和对照组以上指标均无明显变化(P>0．05)。结论 hTERT反义寡核苷酸可显著抑制CTCL细胞的端粒酶活性,抑制细胞生长增殖,诱导细胞凋亡。
Objective To investigate the effect of human telomerase reverse transcriptase （hTERT） gene antisense oligodeoxynucleotide （AODN） on telomerase activity and cell apoptosis in a cutaneous T-cell lymphoma （CTCL） cell line, Hut78. Methods Different concentrations （10 μmol/L, 20 μmol/L, 30 μmol/L） of telomerase antisense oligodeoxynueleotide were introduced into Hut78 cells by lipofectamine-mediated DNA transfection technique. The expressions of hTERT mRNA and telomerase activity were assessed by reverse transcription-polymerase chain reaction and telomeric repeat amplification protocol, respectively. The proliferation and apoptosis of Hut78 cells were detected by flow cytometry. Results After 72 h of incubation, AODN down-regulated the expression of hTERT mRNA, inhibited telomerase activity significantly, and suppressed the viability of Hut78 cells in a time-dependent manner. Cell growth was most clearly suppressed with 30 μmol/L of AODN after 72 h of incubation. The apoptotic rate was 13.05%. Conclusion Telomerase antisense oligodeoxynucleotide could suppress the viability and proliferation of CTCL cell line by inducing apoptosis of these cells.
出处
《中华皮肤科杂志》
CAS
CSCD
北大核心
2005年第12期751-754,共4页
Chinese Journal of Dermatology
基金
国家教委高校博士点基金资助项目(20030023055)
