自噬抑制剂磷酸氯喹增加鼻咽癌CNE-2细胞株的放射敏感性  

Contribution of autophagy inhibitor to radiation sensitization in nasopharyngeal carcinoma cells

作　　者：周支瑞[1] 朱小东[1] 赵伟[1] 曲颂[1] 潘闻燕[1] 郭亚[1] 苏芳[1] 李小宇[1] 

机构地区：[1]广西医科大学附属肿瘤医院广西壮族自治区肿瘤研究所放疗科,南宁530021

出　　处：《中华放射医学与防护杂志》2012年第5期449-454,共6页Chinese Journal of Radiological Medicine and Protection

基　　金：基金项目：国家自然科学基金（81160285）;广西壮族自治区自然科学基金（桂科自2010gxnsfa013240）

摘　　要：目的探讨自噬现象在照射致人鼻咽低分化鳞癌（CNE-2）细胞死亡过程中的作用。方法采用Westernblot技术检测CNE-2细胞经射线照射后自噬标志物LC3、P62的变化，透射电镜检测自噬小体；流式细胞术检测CNE．2细胞凋亡率；MTT法检测CNE-2细胞照射后不同时间的存活率；细胞克隆形成实验检测CNE．2细胞的存活能力及放射敏感性。结果透射电镜检测示自噬抑制剂磷酸氯喹（CDP）抑制照射所致自噬体的形成，自噬诱导剂雷帕霉素（RAPA）增加照射致自噬体数量（F=105．15，P〈0．05）；CDP抑制射线引起的LC3．I向LC3-Ⅱ的转化，而RAPA促进LC3-I向LC3-II的转化（F=231．68，P〈0．05）；CDP使P62表达上调，RAPA使P62表达下调（F=117．52，P〈0．05）；CDP＋照射组和RAPA＋照射组的CNE-2细胞凋亡率明显高于单纯照射组（F=143．72，P〈0．05）；CDP、RAPA降低了照射后CNE-2细胞的存活率（F=25．88，P〈0．05）；二次线性模型拟合剂量存活曲线示自噬抑制剂CDP、RAPA可增加CNE-2细胞株的放射敏感性（F=167．32，P〈0．05）。结论照射联合自噬抑制剂CDP显著增加了CNE-2细胞株对射线的敏感性，提示自噬抑制剂或可用于鼻咽癌的辅助治疗。Objective To investigate the role of autophagy in radiation-induced death response of human nasopharyngeal carcinoma cells. Methods MTT method was used to detect cell viability of CNE-2 cells in different time after irradiation. Clonogenic survival assay was used to evaluate the effect of autophagy inhibitor （chloroquine phosphate） and autophagy inductor （rapamycin） on radiosensitivity of nasopharyngeal carcinoma cells. Cell apoptosis was assessed by flow cytometry. The expressions of LC3 and P62 were measured with Western blot. Cell uhrastructural analysis was performed under an electron microscope. Results Irradiation with 10 Gy induced a massive accumulation of autophagosomes accompanied with up-regulation of LC3-Ⅱ expression in CNE-2 cells. Compared with radiation alone, chloroquine phosphate （CDP） enhanced radiosensitivity significantly by decreasing cell viability （F = 25.88 ,P 〈0. 05）, autophagic ratio（F = 105.15 ,P 〈0. 05）, and LC3- Ⅱ protein level（F =231.68,P 〈 0. 05）, while up-regulating the expression of P62 （ F = 117.52, P 〈 0. 05 ）. Inhibition of autophagy increased radiation-induced apoptosis （ F = 143.72, P 〈 0. 05 ）. Rapamycin （RAPA） also significantly decreased cell viability, but increased autophagic ratio and LC3-Ⅱ protein level while down-regulated the expression of P62. Induction of autophagy increased radiation-induced apoptosis（ F = 167.32, P 〈 0. 05 ） . Conclusions Blockage of autophagy with CDP could enhance radiosensitivity in human nasopharyngeal carcinoma cells, suggesting that inhibition of autophagy could be used as an adjuvant treatment to nasopharyngeal carcinoma.

关 键 词：自噬 鼻咽癌 放射敏感性 磷酸氯喹 雷帕霉素 

分 类 号：R739.63[医药卫生—肿瘤]