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目的探讨辐射对肺腺癌A549细胞细胞周期的影响,观察其细胞上清外泌体TMEM88蛋白(transmembrane protein 88)的表达情况。方法取对数生长期肺腺癌A549细胞,按0、0.5、1、1.5、2 Gy剂量(剂量率1.75 Gy/min,300 Mu/min)照射细胞,培养6和12 h后,流式细胞术检测不同辐射剂量对细胞周期的影响;收集细胞上清,超速离心提取外泌体蛋白,Western blot法分析外泌体TMEM88蛋白的表达情况。结果 G2期细胞6 h检测结果显示,与0 Gy组相比,各剂量组细胞比例显著增加(P<0.001),且随照射剂量增加,细胞比例增加越明显,除0.5 Gy组与1 Gy组外,其余各组间差异均有统计学意义(P<0.001);12 h检测结果显示,G2期细胞比例随照射剂量增加而显著增加,各组间差异均有统计学意义(P<0.05)。S期细胞6 h检测结果显示,与0 Gy组相比,0.5、1.0、1.5 Gy组S期细胞比例显著增加(P<0.05);12 h检测结果显示,与0、0.5 Gy组相比,1.0、1.5 Gy组S期细胞比例显著增加(P<0.05)。随照射剂量的增加,外泌体TMEM88蛋白的表达逐渐降低,6 h检测结果显示,0 Gy组与1.5、2.0 Gy组以及0.5 Gy组与2.0 Gy组相比,差异均有统计学意义(P<0.05));12 h检测结果显示,0.5、1.0、1.5、2.0 Gy组与0 Gy组相比,差异均有统计学意义(P<0.001)。结论辐射能够诱导肺腺癌A549细胞细胞周期阻滞,并能抑制细胞上清外泌体TMEM88蛋白的表达。
Objective To investigate the effect of radiation on the cell cycle of lung adenocarcinoma A549 cells and to observe the expression of TMEM88 protein (TMEM88) in its supernatant. Methods Lung adenocarcinoma A549 cells in logarithmic growth phase were irradiated with 0, 0.5, 1, 1.5 and 2 Gy doses (dose rate of 1.75 Gy / min, 300 Mu / min) and cultured for 6 and 12 h. Flow cytometry The effects of different doses of radiation on the cell cycle were examined. The supernatant of the cells was collected and the exosomes were extracted by ultracentrifugation. The expression of exosome TMEM88 protein was analyzed by Western blot. Results Compared with 0 Gy group, the percentage of cells in each dose group increased significantly (P <0.001) at 6 h. The percentage of cells in G2 group increased more significantly than that in 0 Gy group, except 0.5 Gy group and 1 Gy group (P <0.001). The results of 12 h showed that the proportion of cells in G2 phase increased significantly with the increase of radiation dose, the differences among the groups were statistically significant (P <0.05) . Compared with 0 Gy group, the proportion of S phase cells in 0.5, 1.0, 1.5 Gy group increased significantly (P <0.05) at 6 h in S phase. Compared with 0 and 0.5 Gy group, The proportion of S phase cells in 1.0 and 1.5 Gy group increased significantly (P <0.05). With the increase of irradiation dose, the expression of exosome TMEM88 protein gradually decreased. The results of 6 h showed that there was significant difference between 0 Gy group and 1.5 Gy group and 0.5 Gy group and 2.0 Gy group (P <0.05)). The result of 12 h showed that the difference was statistically significant (P <0.001) in 0.5,1.0,1.5,2.0 Gy group and 0 Gy group. Conclusion Radiation can induce the cell cycle arrest of lung adenocarcinoma A549 cells and inhibit the expression of TMEM88 protein in the supernatant of the cell supernatant.