放射诱导多倍体结肠癌细胞凋亡与自噬的研究

Radiation-induced apoptosis and autophagy of polyploid colon cancer cells

目的探讨放射诱导多倍体结肠癌细胞的凋亡和自噬。方法采用14 Gy剂量的X射线处理SW1116细胞, 分别在放射处理后的第3天、第5天、第7天、第10天及第25天观察细胞的形态变化, 分别记为A组、B组、C组、D组和E组;在第5天、第25天用流式细胞术检测细胞倍性、细胞凋亡的变化;采用Western blot检测细胞周期、凋亡和自噬相关蛋白的表达。结果经14 Gy处理后, SW1116细胞出现多倍体化并伴随细胞体积增大, 细胞发生凋亡和自噬。B组多倍体细胞亚群(DNA含量>4 N)比例[(57.50±2.33)%]和凋亡细胞亚群比例[(14.63±1.90)% ]均高于未放射组细胞[(1.90±0.17)%、(7.57±1.42)%], 差异有统计学意义(P<0.05)。B组细胞周期相关蛋白周期素B1、细胞分裂周期基因2(cdc 2)表达上调, 凋亡抵抗相关蛋白存活蛋白表达上调, 自噬相关蛋白自噬微管相关蛋白轻链3(LC3)-Ⅱ/LC3-Ⅰ比值升高, 自噬特异性底物选择性自噬接头蛋白(p62)蛋白表达下调, 差异有统计学意义(P<0.05)。结论结肠癌多倍体细胞可能通过上调细胞周期相关蛋白、抗凋亡相关蛋白和激活自噬, 来维持多倍体细胞存活并发生去倍化增殖, 参与多倍体细胞的放射抵抗性和治疗后肿瘤的复发。

Objective To investigate the radiation-induced apoptosis and autophagy in polyploid colon cancer cells.Methods SW1116 cells were treated with 14 Gy dose of X-ray,and morphological changes of cells were observed on Day 3,5,7,10 and 25 after radiation treatment,which were classified as Group A,Group B,Group C,Group D and Group E,respectively.The changes of cell ploidy and apoptosis were detected by flow cytometry on Day 5 and 25.Western blot was used to detect the expression of cell cycle,apoptosis and autophagy related proteins.Results After 14 Gy treatment,SW1116 cells showed polyploidy accompanied by increased cell volume,apoptosis and autophagy.The proportion of polyploid cell subsets(DNA content>4 N)(57.50±2.33)%and the proportion of apoptotic cell subsets(14.63±1.90)%in Group B were higher than those of cells in the non-radiation group[(1.90±0.17)%and(7.57±1.42)%],and the differences were statistically significant(P<0.05).In Group B,the expressions of cyclinB1,cell division cycle 2(cdc 2)and apoptosis resistance-related protein Survivin were upregulated,the ratio of autophagy-related proteins microtubule-associated protein light chain 3(LC3)-Ⅱ/LC3-Ⅰwas increased,and the expression of autophagy-specific substrate p62 protein was down-regulated(P<0.05).Conclusions Polyploid colon cancer cells may maintain the survival and deploid proliferation of polyploid colon cancer cells by up-regulating cycle-associated proteins,anti-apoptosis associated proteins and activating autophagy,and participate in the radiation resistance of polyploid colon cancer cells and tumor recurrence after treatment.