大分割电离辐射对放射抗性鼻咽癌细胞免疫原性死亡的影响

Effect of hypofractionated ionizing radiation on immunogenic death of radioresistant nasopharyngeal carcinoma cells

目的研究大分割电离辐射对放射抗性鼻咽癌(NPC)细胞系C666-1R细胞活力、细胞周期及免疫原性细胞死亡(ICD)的影响。方法采用0、2、4、6、8、10、12及14 Gy剂量电离辐射照射NPC细胞株C666-1和C666-1R后、常规培养48 h,采用CCK-8法检测细胞活力,流式细胞术检测细胞周期,RT-PCR法检测钙网蛋白(CRT)、高迁移率族蛋白1(HMGB1)及热休克蛋白70(HSP70)mRNA的表达水平,ELISA和Western blot法检测细胞CRT、HMGB1及HSP70蛋白表达水平,采用试剂盒检测细胞内腺嘌呤核苷三磷酸(ATP)含量。结果经放射处理后不同照射剂量的C666-1R细胞活力均高于C666-1(P<0.05);与未放射细胞相比,经2~14 Gy电离辐射照射的2种NPC细胞株活力均明显下降(P<0.05),8 Gy照射的细胞活性最低(P<0.05),细胞周期阻滞于G_(2)期,CRT、HMGB1及HSP70 mRNA或蛋白表达水平、细胞内的ATP含量均上调(P<0.05)。结论大分割电离辐射能够诱导放射抗性NPC细胞发生ICD。

Objective To investigate the effects of hypofractionated ionizing radiation on cell viability,cell cycle and immunogenic cell death(ICD)of radioresistant nasopharyngeal carcinoma(NPC)cell line C666-1R.Methods NPC cell lines(C666-1 and C666-1R)were irradiated at doses of 0,2,4,6,8,10,12,and 14 Gy and cultured for 48 h routinely.The cell viability was detected by CCK-8.The cell cycle was detected by flow cytometry.The mRNA expression levels of calreticulin(CRT),high mobility group protein 1(HMGB1)and heat shock protein 70(HSP70)were detected by RT-PCR;the protein expression levels of CRT,HMGB1,and HSP70 were detected by ELISA and Western blot.The content of adenine nucleoside triphosphate(ATP)in cells was detected by a kit.Results After radiation treatment,the cell activity of C666-1R was higher than that of C666-1 in all groups treated with different dosages(P<0.05).Compared with the non-radiation group,the viability of NPC cells irradiated by 2 to 14 Gy ionizing radiation was significantly decreased(P<0.05),and the cells with 8 Gy radiation had the lowest viability(P<0.05).The cell cycle was arrested in G_(2) phase.The mRNA or protein expression levels of CRT,HMGB1,and HSP70 and intracellular ATP content were all up-regulated(P<0.05).Conclusion Hypofractionated ionizing radiation can induce ICD in radioresistant NPC cells.