沉默EZH2诱导舌癌Tca-8113细胞凋亡及其对放射敏感性的影响

Effects of EZH2 siRNA on apoptosis induction and radiosensitivity of tongue cancer Tca-8113 cells

目的 探讨zeste基因增强子同源物2（enhancer of zeste homolog 2，EZH2）对舌鳞癌细胞凋亡及放射敏感性的影响。方法 以舌鳞癌Tca-8113细胞为研究对象，细胞转染EZH2小干扰RNA（EZH2 siRNA1、EZH2 siRNA2）及小干扰RNA阴性对照（siRNA-NC），RT-PCR和Western blot检测EZH2表达水平，筛选干扰效果较好的EZH2 siRNA2继续研究。将转染EZH2 siRNA2后的Tca-8113记为EZH2 siRNA2组，同时以不做处理的细胞为对照组，用8 Gy剂量照射转染siRNA对照组和EZH2 siRNA2细胞，并依次命名为照射组和联合组，四甲基偶氮唑盐（MTT）检测细胞增殖，流式细胞术检测细胞凋亡，Western blot检测信号转导与转录因子3（STAT3）、磷酸化的STAT3（p-STAT3）、活性Caspase-3（Cleaved Caspase-3）表达。siRNA-NC组和EZH2 siRNA2组细胞用0、2、4、6、8 Gy剂量照射处理后，细胞克隆实验检测放射敏感性。结果 EZH2 siRNA1、EZH2 siRNA2均能够干扰舌鳞癌细胞中EZH2的表达，且EZH2 siRNA2干扰效果最好（tmRNA=8.660，PmRNA〈0.01；t蛋白=2.883，P蛋白〈0.05）。下调EZH2 siRNA2组细胞凋亡率为（29.90±1.64）%，联合组凋亡率为（38.17±1.59）%，二者比较，差异有统计学意义（t=4.742，P〈0.05），同时下调EZH2还可以降低p-STAT3水平，促进Cleaved Caspase-3蛋白表达。干扰EZH2表达后Tca-8113细胞辐射增敏比为1.668。结论 干扰EZH2表达能够协同放疗促进舌鳞癌细胞凋亡，抑制舌鳞癌细胞增殖，增加舌鳞癌细胞放射敏感性。

Objective To investigate the effect of EZH2 on apoptosis and radiosensitivity of squamous cell carcinoma of tongue.Methods Tongue squamous carcinoma cells Tca-8113 were transfected with small interfering RNA of EZH2 （EZH2 siRNA1, EZH2 siRNA2） and its negative controls （siRNA-NC）, the expression levels of EZH2 were detected by RT-PCR and Western blot and EZH2 siRNA2 was used for further studied since its better interference efficiency. The cells with siRNA transfection were irradiated with 8 Gy doses, and cell proliferation was detected by MTT, apoptosis was detected by flow cytometry, the expression of p-STAT3, STAT3 and Cleaved Caspase-3 was detected by Western blot. In addition, the cells were irradiated with 0, 2, 4, 6, and 8 Gy to detect radiosensitivity by cell colony formation assay.Results EZH2 siRNA1 and EZH2 siRNA2 decreased the expression of EZH2 in Tca-8113 cells and EZH2 siRNA2 had a better interference efficiency （tmRNA=8.660, PmRNA〈0.01; tprotein=2.883, Pprotein〈0.05）. The apoptotic rate in the EZH2 siRNA group was （29.90±1.64）%, and was increased by 8 Gy irradiation to （38.17±1.59）% （t=4.742, P〈0.05）. At the same time, EZH2 siRNA reduced the level of p-STAT3, but promoted the expression of Cleaved Caspase-3 protein, and enhanced the sensitivity of Tca-8113 cells to 1.668-times of control.Conclusions Interfering EZH2 could promote apoptosis, inhibit proliferation and increase radiosensitivity of squamous cell carcinoma of tongue.