吉西他滨对人舌鳞癌Tca8113细胞的放射增敏作用及其机制研究

Radiosensitizing effect of gemcitabine on human tongue squamous cell carcinoma Tca8113 cells and its mechanism

目的研究吉西他滨(GEM)对人舌鳞癌Tca8113细胞的放射增敏作用及其相关机制。方法用不同浓度GEM处理人舌鳞癌Tca8113细胞24 h,测定细胞存活率并确定后续实验浓度。对Tca8113细胞进行单独放疗或联合GEM处理,MTT法测定细胞存活率;平板克隆形成实验研究各组细胞集落形成的能力;流式细胞术验证Tca8113细胞对放射的敏感性变化。Western blotting检测Bcl-2、Bax、 Cleaved Caspase-3及Cleaved Caspase-9蛋白的表达。结果不同浓度GEM作用时的细胞存活率比较,经方差分析,差异有统计学意义(P<0.05)。与0.00μmol/L组比较,GEM浓度为0.01、0.02、0.03、0.10及1.00μmol/L时细胞存活率降低(P<0.05)。照射剂量为4 Gy时,与单纯放疗组比较,GEM联合放疗组细胞存活率降低(P<0.05)。克隆形成实验显示,GEM联合放疗组集落形成能力最弱(P<0.05);流式细胞术进一步证实,GEM联合放疗组较单纯放疗组凋亡率升高(P <0.05)。GEM联合放疗组Tca8113细胞凋亡高于其他各组,且Tca8113细胞中Bax、Cleaved Caspase-3及Cleaved Caspase-9蛋白表达水平高于其他组(P<0.05);而Bcl-2蛋白表达则低于其他各组(P<0.05)。结论 GEM对舌鳞癌Tca8113细胞的增殖有抑制作用,联合放射治疗能有效提高放射敏感性;两者可能具有协同抗肿瘤作用。

Objective To study the radiosensitizing effect of gemcitabine on human tongue squamous cell carcinoma Tca8113 cells and its related mechanisms. Methods After 24 hours-treatment of human tongue squamous cell carcinoma Tca8113 cells with different concentrations of gemcitabine, cell viability and the concentration of subsequent experiments was determined. Tca8113 cells were treated with radiotherapy alone or combined with gemcitabine. MTT assay was used to determine cell viability. Plate colony formation assay was used to study cell colonies. Flow cytometry was used to verify the sensitivity of Tca8113 cells to radiation, and Western blotting was used to detect Bcl-2, Bax, Cleaved Caspase-3, Cleaved Caspase-9 protein expression. Results The cell viability of different concentrations of gemcitabine was analyzed by variance, and the difference was statistically significant (P < 0.05). Compared with the blank control group, the cell viability of the 0.01, 0.02, 0.03, 0.1, and 1 μmol/L gemcitabine concentrations were lower, and the difference was statistically significant (P < 0.05). When the irradiation dose was 4Gy, the cell survival rate of gemcitabine combined with radiotherapy group was lower than that of radiotherapy alone (P < 0.05). Cloning formation experiment showed the colony formation ability of gemcitabine combined with radiotherapy group was the weakest (P < 0.05);flow cytometry further confirmed that the apoptotic rate was higher in gemcitabine combined with radiotherapy group than that in radiotherapy alone group (P < 0.05);The apoptosis of Tca8113 cells in gemcitabine combined with radiotherapy group was higher than that in other groups, and the expression levels of Bax, Cleaved Caspase-3 and Cleaved Caspase-9 in gemcitabine combined with radiotherapy group were higher than those in other groups (P < 0.05), while Bcl-2 protein was opposite.(P < 0.05). Conclusions Gemcitabine has a proliferation-inhibiting effect on tongue squamous cell carcinoma Tca8113 cells, and combined radiotherapy can effectively improve radiosensitivity, and both may have synergistic anti-tumor effects.