喉癌干细胞在放化疗抵抗中的作用机制

Neoplasms stem cells play an important role in resistance of laryngeal squamous cancer to chemoradiotherapy

目的:探讨喉癌干细胞的分选方法,分析顺铂、放射线联合应用对喉癌肿瘤干细胞的杀伤效应及机制。方法:应用流式细胞仪荧光活化细胞分选技术检测并分选出喉癌Hep-2细胞系中的CD133+细胞和CD133-细胞,并检测CD133+细胞亚群的干细胞特性;采用CCK-8试剂盒分别检测顺铂、放射线对两组细胞的生长抑制率;流式细胞仪检测不同干预方案对2组细胞凋亡率及细胞周期分布情况的影响。结果:CD133+细胞在喉癌Hep-2细胞系中占(2.43±0.77)%,在细胞增殖、分化及体内成瘤试验中CD133+细胞均表现出肿瘤干细胞特性;不同浓度剂量的顺铂、放射线对Hep-2细胞均有抑制作用,并在一定浓度范围内呈剂量依赖性;顺铂、放射线单独或联合应用时,CD133-细胞凋亡率显著高于CD133+细胞(P<0.01),并产生G0/G1期阻滞。结论:CD133+细胞较CD133-细胞更具有明显的肿瘤干细胞特性,对放化疗具有明显的抵抗作用,对凋亡诱导作用不敏感和细胞周期改变为其机制之一。

Objective：To determine an approach enriching cancer stem cells from laryngeal cancer cell line. lo investigate whether laryngeal cancer stem cells in chemoradiotherapy have the characteristic of resistance. Method.. CD133＋ cells and CD133- cells was detected and isolated from Hep-2 cell line by fluorescence activated cell sorting technology. The cytotoxicities of cisplatin and radiation were investigated by cell counting kit-8 （CCK-8） assay. The apoptosis and cell cycle was analyzed with flow cytometry. Result： CD133＋ cells accounted for a fraction of （2.43±0.77）% in Hep-2 cell line. CD133＋ cells have a more obvious characteristics of cancer stem cells. Different cisplatin and radiation concentrations of for two cell have inhibition, in a certain concentration range and the dosage dependence. Cisplatin and radiation had synergistic inhibitory effects with CD133- cells on the growth of two cell. Moreover, cell cycle arrest at G0/G1 phase and more apoptosis was induced by synergistic combination. Different concentrations of cetuximab for Hep-2 cells have inhibition, in a certain concentration range and time and the dosage dependence. The half maxial inhibitory concentration （IC50） of cetuximab to Hep-2 cells on 24 h was 1 036.84 ug/L. Cisplatin and radiation had synergistic inhibitory effects with cetuximab on the growth of Hep-2 cell line. Moreover, cell cycle arrest at G0/G1 phase and more apoptosis was induced by synergistic combination. Con clusion：Compared with CD133- cells, CD133＋ cells subpopulation exhibited extraordinary cancer stem.