快中子与光子混合照射生物效应的实验研究

Radiobiological effects of fast neutron/photon mixed irradiation on nasopharyngeal cancer cell line CNE-1

目的探讨快中子与光子混合照射的生物效应及作用机制。方法利用35MeVp→Be快中子(N)和6MVX射线(X),分别以0,40,80,120,160,240,320,400cGy和0,100,200,300,400,600,800,1000cGy的不同剂量照射鼻咽癌细胞株CNE1,经克隆形成分析绘制细胞存活曲线,计算快中子的相对生物学效应(RBE),再根据等效剂量设置两个照射方案(1)1周分次照射分别采用X×5次、N×2次和XNXXN的照射方式,照射后进行克隆形成分析,比较存活分数;(2)两次照射分别采用X+N、N+X和X+X的照射方式,比较照射后不同时间点的细胞周期时相分布和细胞凋亡率。结果快中子的RBE为2.4,6MVX射线单次照射200cGy的快中子等效剂量近似为80cGy。克隆形成分析表明,XNXXN照射的细胞存活分数(0.0079)明显低于X×5次(0.018)和N×2次(0.017)照射(P<0.01)。流式细胞分析发现,混合照射诱发的凋亡率较高,两种射线不同顺序照射后细胞周期阻滞的变化不同,X+N组出现了更明显的G2期阻滞。结论快中子和光子混合体外照射对CNE1细胞有协同杀伤效应,细胞周期阻滞和凋亡可能在混合射线的损伤修复机制中起部分作用。

Objective To study the radiobiological effects of fast neutron/photon mixed irradiation on human cancer cell in vitro and to discuss the mechanism in relation with cell cycle and apoptosis, thus to provide experimental support for the further application of fast neutron radiotherapy of cancer. Methods Exponentially growing human nasopharyngeal cancer cell line CNE-1 was irradiated in vitro with 35 MeV p→Be fast neutron and 6 MV-X ray in grading doses （0 cGy, 40 cGy, 80 cGy, 120 cGy, 160 cGy, 240 cGy, 320 cGy and 400 cGy for neutron, and 0 cGy, 100 cGy, 200 cGy, 300 cGy, 400 cGy, 600 cGy, 800 cGy and 1000 cGy for X ray）.Clonogenic assay was performed, and relative biological effectiveness （RBE） of fast neutron was determined with D10 by means of cell survival curves. Isoeffective doses of 35 MeV p→ Be fast neutron and 6 MV-X my were obtained according to the RBE. The cells were assigned into two irradiation regimens, （1） the one-weekfractionation regimen, which adopted the radiation pattern of X × 5, N × 2 and X-N-X-X-N. After irradiation the clonogenic assay was performed to compare their survival fractions; （2） the two-dose regimen, with the radiation pattern of X ＋ N, N ＋ X and X ＋ X. Flow cytometry was done at different time points after irradiation to analyze cell cycle distribution and apoptosis. Fast neutron dose was delivered on Tuesday and Friday, and all the other irradiation intervals were 24 h. Results The RBE of fast neutron to X ray in CNE-1 cells according to the D10 ratio was 2.40. The neutron isoeffective dose for a single dose of 200 cGy of 6 MV-X ray was approximately 80 cGy. In clonogenic assay, the cell survival fractions were significantly lower in X-N-X-X-N group （0.0079） than those in X × 5 （0.018） and N × 2 （0.017） groups. The flow cytometry suggested a higher percentage of apoptotic cells after mixed irradiation, and different sequence of X ray and neutron irradiations caused varying changes in cell cycle arrest. Conclusion Mixed irradiation of fast neutron and X ray showed a synergic effect in vitro on CNE-1 cell killing. Cell cycle arrest and apoptosis may play some role in the radiation damage repair mechanisms of mixed beam irradiation.