肺癌抑癌基因1对HepG2细胞生长的抑制效应

The growth inhibition effects of TSLC1 gene on human hepatocyte carcinoma cell line HepG2

目的探讨肺癌抑癌基因1（TSLC1）对人肝癌细胞株HepG2生长的影响。方法RT-PCR法制备TSLC1全长cDNA并克隆至真核表达载体pCI-neo，稳定转染至肝癌细胞系HepG2中。以转染空质粒pCI-neo的HepG2细胞为对照组，野生型HepG2细胞为空白组，显微镜下观察细胞形态，MTT法检测细胞增殖，FACSort流式细胞仪检测细胞周期，AnnexinV／PI双染法检测细胞凋亡情况。结果实验建立了高表达TSLCI蛋白的稳定细胞株。实验组细胞呈多角形，聚集成团，细胞之间的黏附非常紧密，对照组和空白组细胞呈梭形，细胞与细胞之间较疏散。与对照组和空白组相比，实验组细胞株细胞生长速度减慢，增殖受到明显抑制，G0／G1期细胞为63．66%±3．83％，高于对照组（47．45％±0．91％）和空白组（54．47％±0．96％）；S期细胞数为22．90％±6．04％，低于对照组（36．58％±0．61％）和空白组（33．61％±2．99％），P〈0．01，实验组细胞周期发生了G0/G1期阻滞。实验组细胞早期凋亡率和晚期凋亡率分别为17．09%±0．20％和16．11％±0．40％，与对照组和空白组细胞相比均明显升高（P〈0．01）。结论TSLC1基因明显抑制HepG2细胞生长，并诱导细胞发生凋亡。

Objective To study the effects of tumor suppressor in lung cancer- 1 （TSLC1） on human hepatocyte carcinoma cell line HepG2. Methods A full length of TSLC 1 cDNA was amplified from RNA of normal human liver cells by RT-PCR, and it was cloned into a pCI-neo expression vector and transfected into human hepatocellular carcinoma cell line HepG2. The HepG2 cells transfected with this plasmid （experimental group） and those treated with pCI-neo vector （control group） and without any treatment （blank group） were compared. Cell morphology was studied microscopically and cell growth was analyzed with MTT assay. FACSort flow cytometry analysis was performed to assess the cell cycle distribution and apoptosis. Results A stable cell line expressing TSLC1 protein was successfully established. Morphologically, cells of the experimental group were tightly aggregated when compared with those of the control and blank groups. The growth of TSLC 1-transfected cells was significantly suppressed in vitro compared with those of the control and blank groups. The amount of G0-G1 cells was 63.66% ± 3.83% （P 〈 0.01） in the experimental group, while those of the control and blank groups were 47.45% ± 0.91% and 54.47% ± 0.96% respectively. The amount of S phase cells in the experimental group, 22.90% ± 6.04%, was significantly lower （P 〈 0.05） than that of the control group （36.58% ± 0.61%） and the blank group （33.61% ± 2.99%）, which suggested a G0-Gl cell cycle arresting. The number of cells in early and late phase apoptosis （17.09% ± 0.20% and 16.11% ± 0.40% respectively） were significantly higher than those of the control and blank groups （P 〈 0.01）. Conclusions TSLC1 strongly inhibits the growth of HepG2 cells in vitro and induces apoptosis of the cells,suggesting that TSLC 1 may have a tumor suppressor function in HCC.