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目的研究苯并(a)芘[B(a)P]对静止期的二倍体人胚肺成纤维细胞(HELF)细胞周期的影响。方法血清饥饿法使细胞同步化于 G_0期;20μmol/L B(a)P 作用细胞4h 后,采用流式细胞术分别检测遗传损伤发生后0、24、48 h 细胞周期分布的改变,并采用 Western blotting 分析0、5、10、20μmol/L B(a)P 作用24 h 后及20 μmol/L B(a)P 作用4 h 后24 h 内细胞周期主要调节基因 p53、p21和 p16表达的改变。结果 0.5%低血清培养48 h 能够较好的达到 G_0同步化效果,处于 G_0期细胞占78%;正常培养细胞持续进入细胞周期进行 DNA 合成,24h 时 S 期细胞达43.9%,而20μmol/LB(a)P 处理组细胞发生 G_1期阻滞;48 h 后对照组细胞完成一个细胞周期回到以 G_1期为主的状态,B(a)P 处理组细胞则从 G_1期阻滞中恢复,继续进入细胞周期,S 期细胞达到了26.5%,延迟约24h。不同浓度 B(a)P 作用后引起 P53、P21蛋白表达明显增加,20μmol/L B(a)P 处理后4 h 即可诱导蛋白表达增加,P53在12 h 后逐渐恢复,P21直至24 h 仍未下降。P16初始略有下降,24 h 恢复。结论B(a)P 引起同步化于 G_0期的细胞发生可逆转的 G_1期阻滞,该阻滞和 p53-p21途径相关。
Objective To investigate the effect of benzo (a) pyrene [B (a) P] on the cell cycle of diploid human embryonic lung fibroblasts (HELF). Methods Cells were synchronized to G 0 by serum starvation method. After treated with 20 μmol / L (a) P for 4 hours, the changes of cell cycle distribution at 0, 24 and 48 h after genetic damage were detected by flow cytometry. The expression of p53, p21 and p16 in the cell cycle regulated by 0, 5, 10, 20μmol / L (a) P for 24 h and 20 μmol / L (a) P for 4 h were analyzed by Western blotting. Results 0.5% hypokalemia could reach the G_0 synchronization effect at 48 h, 78% of cells in G_0 stage. Normal cultured cells continued to enter the cell cycle for DNA synthesis, and reached 43.9% in S phase at 24h, while 20 μmol / L (a) G 1 phase arrest occurred in cells treated with P (P <0.05); cells in the control group returned to G 1 phase after 48 h, cells in B (P) treated group recovered from G 1 phase arrest, Continue to enter the cell cycle, S phase cells reached 26.5%, delayed about 24h. After treated with different concentrations of B (a) P, the expression of P53 and P21 protein were significantly increased, while the expression of P53 and P21 protein was increased 4 h after treatment with 20 μmol / L (P) P, P53 gradually recovered after 12 h, but not up to 24 h after P21 decline. P16 initially decreased slightly, 24 h recovery. Conclusions B (a) P causes reversible G_1 phase arrest in cells synchronized to G_0 phase, which is related to the p53-p21 pathway.