乳腺癌细胞株MDA-MB-231获得性放疗抵抗机制探讨

目的探讨乳腺癌细胞株MDA-MB-231产生获得性放疗抵抗的可能机制。方法采用CCK8及流式细胞术检测产生乳腺癌细胞株MDA-MB-231获得放疗抵抗能力后细胞增殖及细胞周期的变化,采用蛋白质印迹法检测放疗抵抗后相关信号通路蛋白的变化,初步探讨乳腺癌细胞产生放疗抵抗的可能机制。结果 CCK8法检测结果显示,第1天实验组吸光度值为0.305 7±0.013 1,明显高于对照组的0.277 8±0.011 8,差异无统计学意义,F=7.571,P=0.051;第2天实验组为0.401 7±0.048 0,明显高于对照组的0.289 0±0.020 9,差异有统计学意义,F=13.907,P=0.020;第4天实验组为0.635 7±0.026 1,明显高于对照组的0.434 2±0.080 6,差异有统计学意义,F=16.994,P=0.015;第6天实验组为1.5347±0.0391,显著高于对照组的1.075 7±0.036 8,差异有统计学意义,F=218.953,P〈0.001。提示231/RR10细胞株在获得放疗抵抗能力的同时,增殖能力也增强,差异有统计学意义,P〈0.05。流式细胞术检测结果显示,放疗抵抗细胞株中,G0-G1及S期的细胞比例减少,但差异无统计学意义,P值分别为0.083和0.165;G2-M期的细胞比例明显增加,差异有统计学意义,P值分别为0.002和〈0.01。蛋白质印迹法检测结果显示,抑癌基因PTEN的表达明显减少,表皮生长因子的活化状态pEGFR的表达明显增加,AKT蛋白的表达水平无变化,而AKT磷酸化蛋白的表达明显增加。结论乳腺癌细胞株MDA-MB-231中EGFR/AKT信号通路激活,促进细胞生长和生存,从而导致放疗抵抗的产生。这一信号通路可能由抑癌基因PTEN负性调控。

Acquired resistance to radiotherapy of breast cancer cell line MDA-MB-231 and its possible mechanism

OBJECTIVE To investigate the possible mechanism for acquired radioresistance of breast cancer cell line MDA-MB-231. METHODS The cell proliferation was measured by CCK-8. Cell cycle percentage was observed by flow cytometry. Protein expression was analyzed by western blot. RESULTS CCK8 result revealed that cell proliferation of MDA-MB-231/RR10 was significantly promoted in comparison with those in control cells. The OD value of 231/ RR10 group and control group in the {rist day were 0. 305 7±0. 013 1 and 0. 277 8±0. 011 8（F=7. 571, P=0. 051）, in the second day were 0. 401 7±0. 048 0 and 0. 289 0±0. 020 9（F=13. 907,P=0. 020）, in the fourth day were 0. 635 7± 0. 026 1 and 0. 434 2±0. 080 6（F=16. 994,P=0. 015）. The greatest difference occurred in day 6, OD value of 231/RR10 was 1. 534 7±0. 039 1, while control group was 1. 075 7±0. 036 8, F=218. 953,P〈0. 001. Cell cycle analysis indicated that distribution of cells in the G0/G1 and S phases in radioresistant cells was less than that in control cells. However, the differences were not statistically significant （P were 0. 083 and 0. 165, respectively）. There were significantly more cells populated in the G2/M phase （P= 0. 002, P〈0.01）. Western blot result showed that with acquired radioresistance in MDA-MB-231/RR10, the expression of PTEN was remarkably decreased, the expression of phosphorylation EGFR was markedly increased, AKT expression has no change, while phosphorylation AKT expression was significantly increased. CONCLUSION Breast cancer radioresistance was regulated by activated EGFR/AKT signaling pathway-mediated promoted cell proliferation and survival, which could be negatively regulated by tumor suppressor gene PTEN.