鼻咽癌肿瘤干细胞中mTOR信号通路的激活及雷帕霉素对其的抑制作用  

Activation of mTOR signaling pathway in cancer stem cells of nasopharyngeal carcinoma and inhibitive effect of rapamycin against the cancer stem cells

作　　者：张宇[1] 林刃舆[2] 张子恒[2] 陈健[2] 杨春光[3] 周磊[2] 张悦[2] 

机构地区：[1]湖北医药学院附属东风医院耳鼻咽喉头颈外科,湖北十堰442000 [2]温州医科大学附属第一医院耳鼻咽喉头颈外科 [3]中南大学湘雅医学院附属第一医院耳鼻咽喉头颈外科

出　　处：《临床耳鼻咽喉头颈外科杂志》2015年第13期1179-1184,共6页Journal of Clinical Otorhinolaryngology Head And Neck Surgery

摘　　要：目的:研究鼻咽癌肿瘤干细胞中mTOR信号通路蛋白的表达,初步探讨雷帕霉素抑制鼻咽癌肿瘤干细胞增殖的机制。方法:用无血清培养法收集鼻咽癌肿瘤球细胞,用CCK8检测成球细胞及CNE2细胞的增殖活性,Western blot检测干细胞标记物蛋白CD44、OCT4、SOX2和mTOR及其信号通路下游相关蛋白P70S6、4EBP1的表达;CCK8法测不同浓度雷帕霉素(0、0.1、1.0、10.0、100.0、1 000.0nmol/L)对鼻咽癌肿瘤球细胞及CNE2细胞抑制率的差异,Western blot检测雷帕霉素作用肿瘤球细胞后mTOR及其信号通路下游相关蛋白P70S6、4EBP1的表达。结果:成球细胞在含血清培养液中增殖较CNE2细胞明显,与CNE2细胞相比,成球细胞OCT4、SOX2呈过度表达(均P<0.05),而在CD44的表达上,2组间差异无统计学意义(P>0.05);在mTOR、P70S6、4EBP1的表达上,2组细胞无明显差异(P>0.05),而在各自激活态的磷酸化水平(P-mTOR、P-P70S6、P-4EBP1)的表达上,成球细胞较CNE2明显呈过表达状况(均P<0.05);不同浓度雷帕霉素作用鼻咽癌成球细胞及CNE2细胞,对比两者半数抑制率浓度,成球细胞为27.59nmol/L,CNE2细胞为78.12nmol/L,雷帕霉素对成球细胞具有更强的抑制作用;不同浓度雷帕霉素作用成球细胞后,P-mTOR、P-P70S6、P-4EBP1的表达随浓度的升高而呈逐渐下降趋势,而mTOR、P70S6、4EBP1的表达差异无统计学意义。结论:鼻咽癌肿瘤干细胞mTOR信号通路蛋白呈过度表达,雷帕霉素可通过干预mTOR信号通路来抑制其增殖活性。Objective：To study the mTOR expression of cancer stem cells（CSCs）in nasopharyngeal carcinoma and preliminarily explore the mechanism of inhibiting its proliferation with rapamycin.Method：Nasopharyngeal carcinoma spherical cells were gathered by using serum-free suspension culture method,CCK8 assay was used to detect cell proliferation,Western blot assay was used to detect the expression of CD44,OCT4,SOX2 and mTOR signaling.The spherical cells and CNE2 were treated with rapamycin in concentrations of 0,0.1,1.0,10.0,100.0,1 000.0nmol/L,CCK8 assay was used to detect cell inhibition ratio,Western blot assay was used to detect the expression of mTOR signaling of nasopharyngeal carcinoma spherical cells.Result：Compared with CNE2,the spherical cells exhibited a high proliferation rate in RPMI 1640 medium supplemented with fetal bovine serum,and overexpressed in OCT4,SOX2（P〈0.05）,but not that of CD44（P〉0.05）.Although the expression levels of mTOR,P70S6,4EBP1 were not significantly different between the two kinds of cells（P〉0.05）,the proteins of phosphorylation activation form of them（P-mTOR,P-P70S6,P-4EBP1）were highly expressed in spherical cells（P〈0.05）.The spherical cells and CNE2 were treated with rapamycin in different concentrations,the concentrations for 50% of maximal effect of spherical cells and CNE2 were 27.59nmol/L and 78.12nmol/L respectively,rapamycin inhibited the spherical cells more strongly compared with CNE2.The expression levels of P-mTOR,P-P70S6,P-4EBP1 in spherical cells were gradually decreased with increasing of the concentrations of rapamycin,but the difference of the expression levels of mTOR,P70S6,4EBP1 were not significant.Conclusion：The proteins of mTOR signaling pathway of CSCs in nasopharyngeal carcinoma are overexpressed,and rapamycin can effectively inhibit cell proliferation of CSCs in nasopharyngeal carcinoma by blocking mTOR signaling pathway.

关 键 词：鼻咽癌 肿瘤干细胞 MTOR信号通路 雷帕霉素 

分 类 号：R739.6[医药卫生—肿瘤]