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目的探索放射抗拒鳞癌细胞株放射敏感性的改变与端粒酶活性、端粒长度变化的关系。方法以人喉鳞癌细胞株Hep2为实验对象,用γ射线反复照射获得具有放射抗拒性细胞株Hep2R,拟合细胞存活曲线比较两种细胞株的放射生物学参数及端粒酶抑制剂AZT(azidothymidine,叠氮胸苷)作用后的变化,用TRAP-ELISA法测定端粒酶活性(OD值),Southernblotting法分析端粒平均长度(TRF),比较端粒酶活性、端粒平均长度的变化。结果辐射诱导出一个具有放射抗拒性的Hep-2R细胞株,并已稳定传代培养30代以上且放射抗拒性能稳定,测得其SF2=0.6798、D0=3.24,Hep-2R细胞的端粒酶活性较Hep-2细胞升高(0.982±0.005vs0.604±0.015),端粒长度延长了2倍(11.12kbvs3.76kb),AZT作用后Hep2R细胞株SF2=0.4892、D0=2.51,端粒酶活性下降为0.708±0.011、端粒长度缩短为10.18kb。Hep2细胞对应的参数SF2=0.4148、D0=2.06,AZT作用后Hep2细胞SF2=0.3843、D0=1.81,端粒酶活性下降为0.364±0.003、端粒长度缩短为2.76kb。结论辐射诱导细胞获放射抗拒性后其端粒酶活性升高、端粒平均长度增长,端粒酶抑制剂能通过降低端粒酶活性、缩短端粒长度而对其有增敏效应。
Objective To explore the relationship between radiosensitivity of radiation-resisting squamous carcinoma cell lines and changes of telomerase activity and telomere length. Methods Human laryngeal squamous cell carcinoma cell line Hep2 was used as experimental object. Radiation-resistant cell line Hep2R was obtained by repeated irradiation with γ-rays. The survival curves of two cell lines were compared with those of telomerase inhibitor AZT azidothymidine and zidovudine). The telomerase activity (OD) was determined by TRAP-ELISA. The average telomere length (TRF) was analyzed by Southern blotting. The changes of telomerase activity, telomere average length . Results Radiation induced Hep-2R cell line with radioresistance and stable subculture for more than 30 passages with stable radioresistance. SF2 = 0.6798, D0 = 3.24, telomerase activity in Hep-2R cells (0.982 ± 0.005 vs0.604 ± 0.015), the telomere length was doubled (11.12kbvs3.76kb), the Hep2R cell line SF2 = 0.4892, D0 = 2.51, telomerase activity Down to 0.708 ± 0.011, telomere length shortened to 10.18kb. Corresponding parameters SF2 = 0.4148, D0 = 2.06, Hep2 cells SF2 = 0.3843, D0 = 1.81, telomerase activity decreased to 0.364 ± 0.003, telomere length shortened to 2.76kb. CONCLUSION: Radiation-induced cells show increased telomerase activity and longer telomere length. Radiosensitizing effect of telomerase inhibitors can be enhanced by reducing telomerase activity and shortening telomere length.