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目的观察食管癌细胞经射线反复照射后放射敏感性的变化,应用基因芯片技术分析放射抗拒性食管癌细胞基因表达变化。方法食管鳞状细胞癌细胞株TE13经反复γ射线照射(累积剂量120Gy),逐步筛选出具有放射抗拒性的细胞TE13R120。相差显微镜下观察TE13及TE13R120的形态学差异;应用细胞克隆形成实验,验证TE13及TE13R120两种细胞的不同放射敏感性,用流式细胞术检测它们的细胞周期分布特征;基因芯片分析两种细胞基因表达差异。结果TE13R120的细胞群体倍增时间为39·93h,长于亲代TE13(33·94h)。TE13及TE13R120的放射敏感性明显不同(D0值分别为1·63和2·85Gy,SF2值分别为0·55和0·64,Dq值分别为1·38和1·15Gy,n值分别为2·33和1·49)。两种细胞经4Gy放射线照射后,出现不同的细胞周期分布改变,12~48hTE13R120细胞周期变化不明显,而其亲代细胞TE13发生明显G1期阻滞。应用DNA芯片对2159个基因进行了筛选,TE13R120与TE13相比,上调基因96个,下调基因80个。结论新的食管癌细胞系TE13R120比其亲本对射线更加抗拒,并且在基因水平上发生明显变化。4Gy照射后12~48hTE13R120细胞周期变化不明显,而其亲代细胞TE13发生明显G1期阻滞。
Objective To observe the changes of radiosensitivity of esophageal cancer cells after repeated exposure to radiation and analyze the gene expression changes of esophageal cancer cells by gene chip technique. Methods esophageal squamous cell carcinoma cell line TE13 was irradiated by repeated γ-rays (cumulative dose of 120Gy), and gradually screened TE13R120 with radioresistant cells. The morphological differences of TE13 and TE13R120 were observed by phase-contrast microscopy. The different radiosensitivity of TE13 and TE13R120 cells was verified by cell clone formation assay. The cell cycle distribution was analyzed by flow cytometry. Differences in gene expression. Results The population doubling time of TE13R120 was 39.93 h, longer than that of the parental TE13 (33.94 h). The radiosensitivity of TE13 and TE13R120 were significantly different (D0 values were 1.63 and 2.85Gy, SF2 values were 0.55 and 0.64 respectively, Dq values were 1.38 and 1.15Gy, respectively, with n values of 2 · 33 and 1 · 49). After 4Gy radiation, the cell cycle distribution of the two kinds of cells changed. The cell cycle changes of 12 ~ 48hTE13R120 cells were not obvious, while the G1 cell cycle of TE13 cells was obviously blocked. A total of 2159 genes were screened by DNA chip. Compared with TE13, TE13R120 up-regulated 96 genes and down-regulated 80 genes. Conclusion The new esophageal cancer cell line TE13R120 is more resistant to radiation than its parent and has significant changes at the gene level. The changes of cell cycle of TE13R120 cells were not obvious after 12 ~ 48 hours of 4Gy irradiation, while G1 cell cycle of TE13 cells was obviously arrested in 4GY cells.