摘要
目的 探讨电离辐射对口腔鳞状细胞癌细胞SCC25表面NKG2D配体表达的影响及其对肿瘤细胞的杀伤作用.方法 SCC25细胞培养至对数生长期时,通过抽签的方式完全随机设计为对照组（未作处理）和实验组（2Gy电离辐射处理）.对照组和实验组细胞培养24 h后,流式细胞术检测SCC25表面NKG2D配体主要组织相容性复合体Ⅰ类链相关分子（MIC）A、MICB、UL16结合蛋白（ULBP）1的表达量.实验组和对照组细胞培养24 h后,实时荧光定量PCR （RT-PCR）法检测SCC25细胞表面NKG2D配体mRNA表达的变化;制备细胞,将细胞分为空白对照组（NC）、2Gy电离辐射（R）组、NK1组（效靶比为5∶1）、NK2组（效靶比为20∶1）、NK1＋R组（效靶比5∶1,2Gy电离辐射）、NK2＋R组（效靶比20∶1,2Gy电离辐射）,各组细胞培养24 h后,细胞增殖-毒性试验（CCK8）法检测电离辐射和自然杀伤（NK）细胞对口腔鳞状细胞癌SCC25细胞的杀伤能力.结果 流式细胞术结果显示,NKG2D配体MICA对照组和实验组的荧光值分别为21.04±0.39、22.90±0.40（江2.465,P=0.069）,MICB荧光值分别为27.58±0.50、29.83±1.05（t=1.936,P=0.125）,ULBP1的荧光值分别为21.04±0.40、21.78±0.50（t=1.154,P=0.313）,表明SCC25经过电离辐射后,其表面NKG2D配体MICA、MICB、ULBP1的表达量稍有上升,但差异无统计学意义.RT-PCR显示,MICB、ULBP1 mRNA表达对照组和实验组差异均有统计学意义（t=18.334,P=0.000;t=6.381,P=0.008）,实验组的表达量分别是对照组的6.49、1.64倍;CCK8结果显示,NK1、NK2组和NC组相比,细胞杀伤能力的差异有统计学意义（F=344.600,P=0.000）,表明NK细胞可以明显杀死肿瘤细胞,且提高NK细胞与SCC25的比例后杀伤作用更强.R组和NC组相比,细胞杀伤能力差异无统计学意义（P =0.567）,NK1＋R组和NK1组相相比,差异无统计学意义（P=0.915）,NK2＋R组和NK2组相比,差异亦无统计学意义（P =0.678）,表明电离辐射杀伤作用不明显.结论 电离辐射可以使NKG2D配体MICB、ULBP1的mRNA表达增强,可能为肿瘤的免疫治疗提供新的途径.电离辐射对细胞的杀伤作用不明显,可能与辐射剂量低且细胞仅培养24h有关.
Objective To investigate the effect of ionizing radiation on the expression of NKG2D ligand on the surface of oral squamous cell carcinoma cell line SCC25 and its cytotoxicity to tumor cells.Methods When SCC25 cells were cultured into logarithmic growth phase,they were randomly designed as control （without treatment） and experimental group （2 Gy ionizing radiation treatment） by drawing lots.Flow cytometry was used to detect the expressions of NKG2D ligands major histocompatibility complex class Ⅰ chainrelated molecule （MIC）A,MICB,UL16 binding protein （ULBP）1 on the surface of SCC25 in the control group and the experimental group cultured for 24 h.Real-time fluorescence quantification polymerase chain reaction （RT-PCR） was used to detect the changes of NKG2D ligand mRNA expression on SCC25 cell surface after 24 h culture in the experimental group and the control group.The cells were prepared and divided into blank control group （NC）,2 Gy ionizing radiation group （R）,NK1 group （target ratio was 5 ∶ 1）,NK2 group （target ratio was 20 ∶ 1）,NK1 ＋ R group （target ratio was 5 ∶ 1,2 Gy ionizing radiation）,NK2 ＋ R group （target ratio was 20 ∶ 1,2 Gy ionizing radiation）.After each group was cultured for 24 h,the killing abilities of ionizing radiation and natural killer （NK） cells to oral squamous cell carcinoma SCC25 cells were detected by CCK8.Results Flow cytometry experiment showed that,among the NKG2D ligands,the MICA fluorescence values of experimental group and control group were respectively 21.04 ± 0.39,22.90 ± 0.40 （t =2.465,P =0.069）,MICB fluorescence values were 27.58 ± 0.50,29.83 ± 1.05 （t =1.936,P =0.125）,and ULBP1 fluorescence values were 21.04 ± 0.40,21.78 ± 0.50 （t =1.154,P =0.313）.This indicated that after ionizing radiation on SCC25,the NKG2D ligand MICA,MICB,ULBP1 expression increased slightly,but the differences were not statistically significant.RT-PCR indicated that mRNA expressions of MICB,ULBP1 were significantly different between the control group and the experimental group （t =18.334,P =0.000;t =6.381,P =0.008）.The expressions of the experimental group were respectively 6.49,1.64 times as those of the control group.The results of CCK8 showed that,there was a significant difference in cell killing ability among NK1 group,NK2 group and NC group （F =344.600,P =0.000）,suggesting that NK cells could kill tumor cells,and the higher ratio of NK cells and SCC25,the stronger killing effect.The comparison between R group and NC group showed that the difference in cell killing ability was not statistically significant （P =0.567）.NK1 ＋ R group and NK1 group were compared and the difference was not statistically significant （P =0.915）.There was no significant difference between NK2 ＋ R group and NK2 group （P =0.678）.This showed that the killing effect of ionizing radiation was weak.Conclusion Ionizing radiation can increase the mRNA expression of NKG2D ligands MICB and ULBP1.This may provide a new way for tumor immunotherapy.The killing effect of ionizing radiation on cells is not obvious.It may be related to low radiation dose and only 24 h for cell culture.
出处
《国际肿瘤学杂志》
CAS
2017年第8期561-564,共4页
Journal of International Oncology
