Liu Su-Xia, Sun Wen-Sheng, Cao Ying-Lin, Ma Chun-Hong, Han Li-Hui, Zhang Li-Ning, Wang Zhen-Guang, Zhu Fa-Liang
Institute of Immunology, Medical School of Shandong University, Wenhua West Road 44, Jinan 250012, Shandong Province, China.
World J Gastroenterol. 2004 Feb 1;10(3):366-70. doi: 10.3748/wjg.v10.i3.366.
To evaluate the inhibitory effect of antisense phosphorothioate oligonucleotide (asON) complementary to the initiator of human telomerase catalytic subunit (hTERT) on the growth of hepatoma cells.
The as-hTERT was synthesized by using a DNA synthesizer. HepG2.2.15 cells were treated with as-hTERT at the concentration of 10 micromol/L. After 72 h, these cells were obtained for detecting growth inhibition, telomerase activity using the methods of MTT, TRAP-PCR-ELISA, respectively. BALB/c(nu/nu) mice were injected HepG2.2.15 cells and a human-nude mice model was obtained. There were three groups for anti-tumor activity study. Once tumors were established, these animals in the first group were administered as-hTERT and saline. Apoptosis of tumor cells was detected by FCM. In the 2nd group, the animals were injected HepG2.2.15 cells together with as-hTERT. In the third group, the animals were given as-hTERT 24 hours postinjection of HepG2.2.15 cells. The anti-HBV effects were assayed with ELISA in vitro and in vivo.
Growth inhibition was observed in cells treated with as-hTERT in vitro. A significant different in the value of A570-A630 was found between cells treated with as-hTERT and control (P<0.01) by MTT method. The telomerase activity of tumor cells treated with as-hTERT was reduced, the value of A450 nm was 0.42 compared to control (1.49) with TRAP-PCR-ELISA. The peak of apoptosis in tumor cells given as-hTERT was 21.12%, but not seen in saline-treated control. A prolonged period of carcinogenesis was observed in the second and third group animals. There was inhibitory effect on the expression of HBsAg and HBeAg in vivo and in vitro.
As-hTERT has an anti-tumor activity, which may be useful for gene therapy of tumors.
评估与人类端粒酶催化亚基（hTERT）启动子互补的硫代磷酸反义寡核苷酸（asON）对肝癌细胞生长的抑制作用。
使用DNA合成仪合成as-hTERT。用浓度为10微摩尔/升的as-hTERT处理HepG2.2.15细胞。72小时后，分别采用MTT法、TRAP-PCR-ELISA法检测这些细胞的生长抑制情况和端粒酶活性。将HepG2.2.15细胞注射到BALB/c（nu/nu）小鼠体内，建立人裸鼠模型。进行抗肿瘤活性研究分为三组。肿瘤形成后，第一组动物给予as-hTERT和生理盐水。通过流式细胞术检测肿瘤细胞凋亡情况。第二组动物注射HepG2.2.15细胞的同时注射as-hTERT。第三组动物在注射HepG2.2.15细胞24小时后给予as-hTERT。采用ELISA法在体外和体内检测抗乙肝病毒效果。
体外实验中，as-hTERT处理的细胞出现生长抑制。MTT法检测显示，as-hTERT处理的细胞与对照组之间A570-A630值存在显著差异（P<0.01）。as-hTERT处理的肿瘤细胞端粒酶活性降低，TRAP-PCR-ELISA法检测显示，与对照组（1.49）相比，A450纳米值为0.42。给予as-hTERT的肿瘤细胞凋亡峰值为21.12%，而生理盐水处理的对照组未见凋亡。第二组和第三组动物的肿瘤发生期延长。在体内和体外对乙肝表面抗原（HBsAg）和乙肝e抗原（HBeAg）的表达均有抑制作用。
As-hTERT具有抗肿瘤活性，可能对肿瘤的基因治疗有帮助。
