放疗耐受性宫颈癌Hela细胞的生物学特性

Biological Characteristics in Cervical Cancer Cell Line Hela Tolerated to Radiation

【目的】研究耐放疗的宫颈癌Hela细胞生物学特性的改变,并探讨其与宫颈癌肿瘤干细胞间的关系。【方法】采用多次分割剂量照射技术建立宫颈癌Hela细胞的耐放疗模型(Hela-R),实验分4组:Hela-R1组,Hela-R2组,Hela-R3组和对照组。四甲基偶氮唑蓝法检测细胞生长情况,克隆形成实验测定放射敏感性和克隆能力,流式细胞术检测细胞周期分布和增殖能力,球囊培养法检测细胞自我更新能力。【结果】Hela、Hela-R细胞接受照射后均呈现先加速增殖后出现生长抑制的现象。Hela-R1、Hela-R2、Hela-R3的细胞倍增时间分别为(43.4±1.0)h、(49.2±2.0)h和(48.7±3.3)h,克隆形成率分别为(20.3±4.0)%、(49.3±11.6)%和(6.3±5.9)%,S期细胞比例分别为(9.9±0.4)%、(13.0±0.9)%和(9.6±0.7)%,增殖指数(PI)分别为(27.3±2.6)%、(31.8±4.9)%和(37.4±8.0)%。与对照组比较,Hela-R3组的放射抗拒性增强。非粘附性球囊培养法培养Hela及Hela-R细胞可得到肿瘤细胞球,四组的球囊形成率分别为(9.9±0.4)%、(13.0±0.9)%、(9.6±0.7)%和(5.0±0.3)%。【结论】多次分割剂量照射可在体外建立宫颈癌Hela细胞的耐放疗模型,并可富集肿瘤干细胞;多次分割照射后,Hela细胞生长速度减慢,增殖能力有升高趋势,自我更新能力、克隆能力增强,细胞周期无明显变化。

[ Object ] The aim of the study was to investigate biological behaviors of radioresistant cervical cancer cells. The relationship between radioresistant cancer cells and cancer stem cells would be discussed. [ Methods ] Hela cells were treated with fractionated irradiation, yielding Hela-R, the radioresistant model. There were four groups ： Hela-R1, Hela-R2, Hela-R3 and control. Biological characteristics, including cell growth, clone-generating capability, cell cycle distribution and proliferation, and tumor sphere-forming rate were detected by MTI＂ assay, clone formation assay, flow cytometry assay and serum-free medium culturing, respectively. [Results] Before growth inhibition, both the parental cells and the sub-lines Hela-R were accelerated proliferation by exposure to irradiation. In the three groups of Hela-R, cell doubling times were （43.4± 1.0）h, （49.2 ± 2.0）h, and （48.7 ±3.3）h; the colony forming rates were （20.3 ± 4.0）%, （49.3 ±11.6）%, and （6.3 ±5.9）%; S-Phase Fraction were （17.6 ± 1.3）%, （20.0 ± 2.4）%, and （23.9± 2.3）%; Proliferation index （PI） were （27.3 ±2.6）%, （31.8 ± 4.9）%, and （37.4± 8.0）%, respectively. Compared with the parental cells, cells in Hela-R3 were more radioresistant to electronic rays. The formation of tumor spheres could be observed when Hela cells and Hela-R cells were cultured in non-adherent serum-free medium, and tumor sphere-forming rates were（9.9 ± 0.4）%, （13.0 ± 0.9）%, （9.6 ± 0.7）%, and （5.0 ± 0.3）%, respectively. [Conclusion]Long-term fractionated radiation exposure conferred tolerance of ionizing radiation to the human tumor cells Hela, which is a method to establish radioresistant model and obtain tumor stem cells by radiation enrichment. Hela-R cells were more powerful in self-renew and cloning-generating capability, with a trend of proliferative capability increased, while the cellular growth was slower and cell cycle showed no significant change.