尼妥珠单抗对食管鳞癌ECA-109细胞放疗敏感性的影响

Effect of Nimotuzumab on Radiosensitivity of Esophageal Squamous Cell Carcinoma ECA-109 Cells

目的探讨尼妥珠单抗对食管鳞癌ECA-109细胞放疗敏感性的作用及可能的机制。方法选取人食管鳞癌细胞ECA-109,采用克隆形成实验检测尼妥珠单抗对ECA-109细胞放射敏感性的影响;将细胞分为对照组、400μg·mL^(-1)尼妥珠单抗组、6 Gy放射组和400μg·mL^(-1)尼妥珠单抗+6 Gy放射组,采用流式细胞仪检测各组细胞周期及凋亡情况,采用Western blot检测各组DNA-PKcs、p-DNA-PKcs和γH2AX蛋白表达。结果 2~6 Gy照射后,50μg·mL^(-1)、200μg·mL^(-1)和400μg·mL^(-1)尼妥珠单抗组ECA-109细胞存活分数均明显低于对照组(P<0.05),50μg·mL^(-1)、200μg·mL^(-1)和400μg·mL^(-1)尼妥珠单抗组的放射增敏比(SER)分别为1.01、1.06和1.13;400μg·mL^(-1)尼妥珠单抗+6 Gy放射组ECA-109细胞G2/M期比例为(29.76±1.10)%,明显高于对照组、400μg·mL^(-1)尼妥珠单抗组和6 Gy放射组;400μg·mL^(-1)尼妥珠单抗+6 Gy放射组ECA-109细胞凋亡率为(40.43±1.60)%,明显高于对照组、400μg·mL^(-1)尼妥珠单抗组和6 Gy放射组(P<0.05);400μg·mL^(-1)尼妥珠单抗+6 Gy放射组ECA-109细胞p-DNA-PKcs蛋白相对表达为(0.103±0.064),明显低于对照组、400μg·mL^(-1)尼妥珠单抗组和6 Gy放射组(P<0.05),而γH2AX蛋白相对表达为(0.416±0.103),明显高于对照组、400μg·mL^(-1)尼妥珠单抗组和6 Gy放射组(P<0.05)。结论尼妥珠单抗可增加食管鳞癌ECA-109细胞的放射敏感性,可能与其阻滞细胞周期G2/M期、诱导细胞凋亡和阻碍DNA损伤修复有关。

Objective To investigate the radiosensitivity of nimotuzumab onesophageal squamous cell carcinoma cell line ECA-109and its possible mechanism.Methods Human esophageal squamous cell carcinoma cell line ECA-109was selected and clonogenic assay was used to detect the radiosensitivity of nimotuzumab on ECA-109cells;The cells were divided into control group,400μg·mL-1nimotuzumab group,6Gy radiation group and400μg·mL-1nimotuzumab+6Gy radiation group,the cell cycle and apoptosis of each group were detected by flow cytometry,and the expression of DNA-PKcs,p-DNA-PKcs andγH2AX protein in each group was detected by Western blot.Results After2~6Gy irradiation,the survival fraction of ECA-109cells of50μg·mL-1,200μg·mL-1and400μg·mL-1nimotuzumab group were significantly lower than that of the control group(P<0.05);radiosensitization ratio(SER)in50μg·mL-1,200μg·mL-1and400μg·mL-1nimotuzumab group were1.01,1.06and1.13;The percentage of ECA-109cells of G2/M phase in400μg·mL-1nimotuzumab+6Gy radiation group was(29.76±1.10)%,which was significantly higher than that in control group,400μg·mL-1nimotuzumab group and6Gy radiation group;The apoptosis rate of ECA-109cells in400μg·mL-1nimotuzumab+6Gyradiatio group was(40.43±1.60)%,which was significantly higher than that in control group,400μg·mL-1nimotuzumab group and6Gy radiation group(P<0.05);The p-DNA-PKcs protein relative expression of ECA-109cells in400μg·mL-1nimotuzumab+6Gy radiation group was(0.103±0.064),which was significantly lower than that in the control group,400μg·mL-1nimotuzumab group and6Gy radiation group(P<0.05),andγH2AX protein expression was(0.416±0.103),which was significantly higher than that in the control group,400μg·mL-1nimotuzumab group and6Gy radiation group(P<0.05).Conclusion Nimotuzumab can increase the radiosensitivity of esophageal squamous cell carcinoma ECA-109cells,which may be related to cell cycle arrest at G2/M phase,induction of apoptosis and inhibition of DNA damage repair.