抑制自噬通过促进Caspase依赖性细胞凋亡逆转鼻咽癌紫杉醇耐药性的研究  

Inhibition of Autophagy Sensitizes Taxol-resistant Nasopharyngeal Carcinoma Cells to Taxol Treatment by Enhancing Taxol-induced Caspase-dependent Apoptosis

作　　者：陈亦龙 李维[1] 苏雪萍 谭国林[1] 李和清[1] CHEN Yi-long;LI Wei;SU Xue-ping;TAN Guo-lin;LI He-qing(Third Xiangya Hospital of Central South University,Changsha 410000,China;PLA 303 Hospita,Nanning 530000,China)

机构地区：[1]长沙市中南大学湘雅三医院,湖南长沙410000 [2]南宁市中国人民解放军第三〇三医院,广西南宁530000

出　　处：《中国肿瘤》2018年第8期626-633,共8页China Cancer

基　　金：国家自然科学基金青年项目(81702706)

摘　　要：[目的]探讨细胞自噬在鼻咽癌紫杉醇耐药中的作用及其机制。[方法]以鼻咽癌亲本细胞和紫杉醇耐药细胞为研究对象。自噬抑制剂3-MA抑制自噬,si RNA下调自噬蛋白Beclin-1的表达,分别应用集落形成实验检测细胞的增殖能力,流式细胞术检测凋亡水平,蛋白印迹分析检测自噬相关蛋白和Caspase蛋白的表达水平,透射电子显微镜和荧光显微镜进行细胞形态学观察。通过基因芯片技术分析Caspase家族基因在亲本和耐药细胞中的不同表达。[结果]紫杉醇耐药细胞中自噬蛋白Beclin-1、LC3-Ⅱ的表达水平高于亲本细胞（P〈0.05）。用自噬抑制剂3-MA预处理鼻咽癌耐药细胞后,CNE-1/Taxol＋3-MA和HNE-2/Taxol＋3-MA细胞的IC50值分别为（6.25±1.0）nmol/L、（2.19±0.19）nmol/L,与对照组相比,细胞的生长抑制率明显增加（P〈0.01）。鼻咽癌紫杉醇耐药细胞转染Beclin-1-si RNA后,CNE-1/Taxol细胞和HNE-2/Taxol细胞的IC50值分别为（5.69±1.23）nmol/L、（3.08±0.67）nmol/L,转染后的细胞对紫杉醇的敏感性明显高于阴性对照组。通过3-MA和Beclin-1-si RNA抑制自噬能增加紫杉醇诱导的Caspase依赖的细胞凋亡,同时能部分逆转鼻咽癌细胞紫杉醇耐药。[结论]紫杉醇诱导的鼻咽癌细胞自噬可能是通过降低Caspase依赖的细胞凋亡,从而部分参与了紫杉醇耐药,抑制自噬能通过促进凋亡从而增加鼻咽癌细胞对紫杉醇的敏感性。[Purpose] To investigate the effects and mechanisms of autophagy in the taxol resistance of nasopharyngeal carcinoma（NPC） cells. [Methods] The NPC parental（CNE-1, HNE-2） and taxolresistant（CNE-1/Taxol, HNE-2/Taxol） cells were used in the study;3-MA was used as autophagy inhibitors;si RNA to Beclin-1 was used to inhibit expression of Beclin-1. Colony formation assay was utilized to detect cell proliferation;flow cytometry assay was used to detect apoptosis. Expression of autophagy-related protein and cleaved Caspase 3 were measured by Western blot analysis. Morpholo-gy studies were performed using transmission electron microscopy and fluorescent microscopy. The c DNA microarray was used to analyze the different expression levels of caspase family genes between parental and taxol-resistant cells. [Results] The expression levels of autophagy protein Beclin-1,LC3-Ⅱ in taxol-resistant cells were higher than those in parental cells（P〈0.05）. The IC50 values of taxol in CNE-1/Taxol and HNE-2/Taxol were（6.25±1.0）nmol/L and（2.19±0.19）nmol/L after pretreatment with 3-MA, which were significantly lower than that in control groups（P〈0.01）.The IC50 value of taxol in CNE-1/Taxol and HNE-2/Taxol were（5.69±1.23）nmol/L and（3.08±0.67）nmol/L after transfection with Beclin-1-si RNA, which were significantly lower than those in control groups（P〈0.05）. Inhibition of autophagy through 3-MA and Beclin-1-si RNA enhanced taxol-induced caspase-dependent apoptosis,resulted in partial reversal of the acquired taxol resistance in taxol-resistant cells. [Conclusion] The taxol-induced autophagy may protect nasopharyngeal carcinoma cells from caspase-dependent apoptotic death, and inhibition of autophagy can increase the taxol sensitivity by increasing apoptosis.

关 键 词：鼻咽肿瘤 紫杉醇 自噬 凋亡 耐药 

分 类 号：R739.62[医药卫生—肿瘤]