Pang Jian-Xin, Cheng Xi-Yuan, Xu Wei, Wu Shu-Guang
Institute of Pharmaceutic Sciences, First Military Medical University, Guangzhou 510515, China.
Acta Pharmacol Sin. 2003 Jan;24(1):91-6.
To study the effects of transcriptional factor Sp1 antisense oligodeoxynucleotide (ODN) on telomerase activity and human telomerase reverse transcriptase (hTERT) expression.
Antisense oligodeoxynucleotide (ODN) was designed to inhibit Sp1 expression and transferred to Jurkat T cells by lipofectamin. Telomerase PCR-ELISA was used to detect telomerase activity. RT-PCR analysis was used to assess the mRNA expression of Sp1 and hTERT, and Western blot was used to analyze the levels of Sp1 protein.
Treatment of Jurkat T cells with Sp1 antisense ODN (1 micromol/L) dramatically reduced Sp1 mRNA and protein levels. The inhibition rate was 44.8 % (P <0.05) and 57 % (P <0.01), respectively. Following the transcriptional factor Sp1 functionally altering, hTERT mRNA expression were suppressed with a 43.7 % inhibition rate (P <0.01). A dose-dependent inhibition of telomerase activity by antisense Sp1 ODN was also discovered. From 0.25 to 2.0 micromol/L, telomerase activity was reduced from 27.1 % to 64.6 %.
Antisense Sp1 ODN decreases telomerase activity by inhibiting hTERT mRNA expression in Jurkat T cells.
研究转录因子Sp1反义寡脱氧核苷酸(ODN)对端粒酶活性及人端粒酶逆转录酶(hTERT)表达的影响。
设计反义寡脱氧核苷酸(ODN)抑制Sp1表达,并通过脂质体转染法将其转入Jurkat T细胞。采用端粒酶PCR-ELISA法检测端粒酶活性。运用RT-PCR分析法评估Sp1和hTERT的mRNA表达,并用蛋白质免疫印迹法分析Sp1蛋白水平。
用Sp1反义ODN(1 μmol/L)处理Jurkat T细胞后,Sp1的mRNA和蛋白水平显著降低。抑制率分别为44.8%(P<0.05)和57%(P<0.01)。转录因子Sp1功能改变后,hTERT mRNA表达受到抑制,抑制率为43.7%(P<0.01)。还发现反义Sp1 ODN对端粒酶活性有剂量依赖性抑制作用。在0.25至2.0 μmol/L范围内,端粒酶活性从27.1%降至64.6%。
反义Sp1 ODN通过抑制Jurkat T细胞中hTERT mRNA表达降低端粒酶活性。