人鼻咽癌放射抗拒细胞株建立及其细胞周期的观察

Establishment of a radioresistant subline from human nasopharyngeal carcinoma cell line and its cell cycle distribution pattern

目的:建立人鼻咽癌放射抗拒细胞株CNE-2R,为研究鼻咽癌放射抗拒机制提供具有可比性的成对细胞。方法:用γ射线反复照射CNE-2细胞,筛选出放射抗拒细胞株CNE-2R;检测细胞生长的倍增时间;应用细胞克隆形成实验检测细胞的放射敏感性;MTT法检测照射后不同时间的存活分数;用流式细胞术检测细胞的周期分布特征。结果:CNE-2R的细胞群体倍增时间长于亲代CNE-2(4.5dvs3.6d),P=0.047;CNE-2及CNE-2R的放射敏感性明显不同,CNE-2R的放射抗拒性增加;照射后12hCNE-2R的存活率大于CNE-2(89.0%vs71.7%),P=0.001;细胞分布周期改变,CNE-2R的S期比例增高,G0/G1和G2/M期细胞减少,χ2=7.312,P=0.034;CNE-2R在照射后12～36h出现明显的G2/M期阻滞,亲本CNE-2仅在照射后12～24hG2/M期略有增加。结论:人鼻咽癌细胞株CNE-2经间歇性大剂量γ射线多次照射后得到的CNE-2R具有稳定放射抗拒性,并且显示出与亲代不同的细胞周期特征。

OBJECTIVE：To establish a radioresistant suhline from human nasopharyngeal carcinoma cell line CNE-2, so as to supply comparable pair cells for investigating the mechanism of radioresistance. METHODS： CNE-2 human nasopharyngeal carcinoma cells were exposed repeatedly to γ-rays and then screened for a new radioresistant subline named CNE-2R. Double times of CNE-2 and CNE-2R cells were determined by MTT assays. The relative radiosensitivities of tumor cells were assessed by standard colony formation assays. Survival rates after exposed to irradiation were measured by MTT assays. The time courses of cell cycle distribution before and after irradiation were investigated by flow cytometry （FCM）. RESULTS： The double time of CNE- 2R cells was longer than that of parental generation CNE-2 cells（4.5 d vs 3.6 d, P=0. 047）. By using the colony-forming assay and line-quad- ratic model, the parameters of CNE-2 and CNE-2R were obtained. The radioresistance of CNE-2R cells was stronger than that of CNE-2 cells. After exposed to irradiation respectively, the survival rate in CNE-2R was also higher than that in CNE-2 （89.0% vs 71.7%, P=0. 001）. The CNE-2R subline showed higher percentage of cells in S phase, and lower percentage of cells in G0/G1 and G2/M phases （χ^2 =7. 312, P= 0. 034）. A marked accumulation of cells in G2/M phase was observed in CNE-2R ceils during the period from 12 to 36 h after irradiation A little increase of cells in G2/M phase was observed in CNE-2 cells from 12 to 24 h after irradiation. CONCLUSION： The new subline CNE-2R is more radioresistant comparable to its parental CNE-2 cell line, and has a different cell cycle distribution.