模拟调强技术照射鼻咽低分化鳞癌细胞的生物效应机制研究

Radiobiological effect of simulative intensity-modulated radiotherapy in poor differentiated nasopharyngeal carcinoma cells

目的探讨延长时间的调强照射对鼻咽低分化鳞癌细胞系（CNE-2）放射生物效应的影响以及初步机制研究。方法实验分为空白对照组、单次照射组、模拟调强照射组，照射采用6MV X线照射2、4、6、8Gy（成克隆实验增加1Gy剂量点），剂量率3Gy／min。单次照射组完成时间1—3min，模拟调强照射组各剂量点分别等分割5次，每次间隔8．0～8．5min。采用克隆分析法检测不同照射模式下CNE-2细胞的放射敏感性；流式细胞仪检测细胞凋亡率；RT—PCR法检测细胞凋亡相关因子Box、Bcl-2的转录水平。结果模拟调强照射组不同剂量的存活分数均高于单次照射组，其仪单次照射〉d模拟调强照射（0．675Gy^-1：0．538Gy^-1）、p单次照射〉p模拟调强照射（0．051Gy^-2：0．027Gy^-2）、D0单次照射〈D0模拟调强照射（0．885Gy：0．986Gy）、Dq单次照射〈Dq模拟调强照射（0．563Gy：0．946Gy）。模拟调强照射组CNE-2细胞早、晚期平均凋亡率均低于单次照射组[21．20％：15．89％（F=18．51，P=0．020）、13．00％：10．20％（F=15．67，P=0．040）]。模拟调强照射组Bax平均转录水平低于单次照射组[76．75％：62．50％（F=36．57，P=0．000）]并呈剂量依赖性关系；Bcl-2平均转录水平两组差异无统计学意义[29．25％：29．75％（F=0．74，P=0．800）]；Bax／Bcl-2比平均值模拟调强照射组低于单次照射组[26．50％：21．10％（F=16．02，P=0．020）]。结论模拟调强照射模式下随分次照射时间延长，相对剂量率降低，从而导致生物效应下降。

Objective To study the altered radiobiological effect of simulative intensity-modulated radiotherapy （SIMR） in cultured human nasopharyngeal carcinoma （NPC） cells and the related mechanism. Methods Single cell suspension of exponentially growing CNE-2 cells, a poor differentiated NPC cell line, was seeded and cultured for 12 hours, then the cells were irradiated in two different models by 6 MV X-ray beams at 3 Gy/min, In single fraction irradiation （SFR） model, cells were irradiated a single fraction of 0, 2, 4, 6 or 8 Gy within 0 to 3 minutes. In SIMR model, cells were irradiated 0, 2, 4, 6 or 8 Gy in 5 fractions with interval of 8.0 - 8.5 minutes between. Clonogenic assay was performed to determine the radiosen- sitivity. Cellular apoptosis was measured by flow cytometry. RT-PCR was used to detect mRNA expressions of Bax and Bcl-2, Respectively. Results Compared with SFR group, the survival fraction in SIMR group was higher at all the dose levels. The values of α,β, Do and Dq were higher in SIMR group than in SFR group. At dose levels of 2 Gy, 4 Gy and 6 Gy, The early and late apoptotic cells in SIMR group were lower than in SFR group （21.20%： 15.89%, F=18.51, P =0.020;13.00%： 10.20, F=15.67, P =0.040）. The mRNA expression of Bax was upregulated in a dose-dependent manner in the both groups. Compared with SFR group, the mRNA expression of Bax in SIMR group was lower at all the dose levels （ Mean value of 76.75%：62.50%, F = 36.57, P =0. 000）. Bcl-2 mRNA expression at every dose level had no significant difference between the two groups （ Mean value of 29.25 % ： 29.75 %, F = 0.74, P = 0.800）. Conclusions Prolonged delivery time in SIMR model can decrease the radiobiological effects.