论文部分内容阅读
[目的]探讨镉致人支气管上皮(16HBE)细胞恶性转化中细胞周期的变化,分析去甲基化处理后细胞周期蛋白基因(CyclinD1)的表达情况。[方法]采用流式细胞术检测正常16HBE细胞和氯化镉恶性转化16HBE细胞不同阶段(第5、15、35代细胞)及镉恶性转化第35代细胞接种裸鼠成瘤细胞(简称成瘤细胞)的细胞周期变化情况;采用实时荧光定量聚合酶链反应和Western blot法检测上述细胞及经去甲基化转移酶抑制剂5-氮杂-2-脱氧胞苷(5-Aza-dC)处理后的成瘤细胞CyclinD1的mRNA和蛋白表达情况。[结果]与正常16HBE细胞比较,氯化镉恶性转化16HBE细胞系第35代细胞及成瘤细胞在G0/G1期的细胞比例随着转化代数的增加而增大,分别为60.04%、64.12%;而在S期和G2/M期细胞比例则随着转化代数的增加而减少——在S期细胞比例分别为35.03%、31.83%,在G2/M期分别为4.93%、4.01%——差异均有统计学意义(均P<0.05)。与正常16HBE细胞相比,氯化镉恶性转化16HBE细胞不同阶段(第5、15、35代细胞及成瘤细胞)的CyclinD1的mRNA和蛋白表达量呈现随着代数增加而上调的趋势;其中第35代细胞(2.34倍和2.41倍)及成瘤细胞(2.33倍和2.35倍)明显增加(均P<0.05);5-Aza-dC处理后,成瘤细胞的CyclinD1的mRNA和蛋白表达增加量有所下降,分别为处理前16HBE细胞的1.815倍和1.742倍(P<0.05)。[结论]镉恶性转化16HBE细胞中细胞周期发生变化,而这些改变可能与DNA甲基化相关。
[Objective] To investigate the changes of cell cycle in malignant transformation of cadmium-induced human bronchial epithelial (16HBE) cells and to analyze the expression of cyclinD1 after demethylation treatment. [Methods] Flow cytometry was used to detect the tumorigenicity of nude mice in normal 16HBE cells and malignant transformed cadmium chloride 16HBE cells (5th, 15th, 35th generation) and 35th generation of cadmium Cell cycle changes were detected by real-time fluorescence quantitative polymerase chain reaction and Western blot. The above cells and 5-Aza-2-deoxycytidine (5-Aza-dC) The mRNA and protein expression of CyclinD1 in the treated tumorigenic cells. [Results] Compared with normal 16HBE cells, the percentage of cells in the 35th passage and the tumorigenicity of 16HBE cells transformed with cadmium chloride increased with the number of transformed algebra at 60.04%, 64.12% While the proportion of cells in S phase and G2 / M phase decreased with the increase of transformation algebra - the proportion of cells in S phase was 35.03% and 31.83% respectively, 4.93% and 4.01% respectively at G2 / M phase, The differences were statistically significant (all P <0.05). Compared with normal 16HBE cells, the expression of CyclinD1 mRNA and protein in malignant transformed cadmium chloride-16HBE cells at different stages (5th, 15th, 35th generation and tumorigenic cells) showed an upward trend as algebraic increase. 35 cells (2.34- and 2.41-fold) and tumorigenic cells (2.33- and 2.35-fold) increased significantly (all P <0.05). After 5-Aza-dC treatment, the increase of mRNA and protein expression of CyclinD1 in tumorigenic cells Decreased by 1.815-fold and 1.742-fold (P <0.05) respectively, compared with the 16HBE cells before treatment. [Conclusion] The cell cycle changes in cadmium malignant transformed 16HBE cells, and these changes may be related to DNA methylation.