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目的:探讨肿瘤坏死因子(TNF)与白血病U937细胞的bcl—2基因表达和细胞周期的关系。方法:首先用TNF处理U937细胞,提取小片段DNA进行DNA断裂分析,继而用RT—PCR和S—P免疫组化染色法检测U937细胞的bcl—2基因表达变化,并通过流式细胞术分析TNF对U937细胞的DNA含量和细胞周期的影响。最后,用磷酸钙沉淀法将bcl—2基因转染到U937细胞,经G418筛选稳定转染子,用不同剂量TNF处理后观察转染与未转染细胞的存活率。结果:TNF可诱发U937细胞凋亡,经TNF处理的U937细胞bcl—2基因表达下降,细胞周期阻滞于G0/G1期。用bcl—2基因稳定转染的U937细胞经TNF处理,存活率明显高于未转染细胞。结论:TNF下调U937细胞bcl—2基因表达、改变细胞周期是其诱导细胞凋亡的可能作用机制。
Objective: To investigate the relationship between tumor necrosis factor (TNF) and bcl-2 gene expression and cell cycle in leukemia U937 cells. Methods: First, U937 cells were treated with TNF and DNA fragments were extracted for DNA fragmentation analysis. The expression of bcl-2 gene in U937 cells was detected by RT-PCR and S-P immunohistochemical staining, and analyzed by flow cytometry Effect of TNF on DNA content and cell cycle of U937 cells. Finally, the bcl-2 gene was transfected into U937 cells by calcium phosphate precipitation method. Stable transfectants were screened by G418 and treated with different doses of TNF to observe the survival rate of transfected and non-transfected cells. Results: TNF induced the apoptosis of U937 cells. The expression of bcl-2 gene in U937 cells treated with TNF decreased and the cell cycle arrested in G0 / G1 phase. U937 cells stably transfected with bcl-2 gene were treated with TNF and the survival rate was significantly higher than that of untransfected cells. Conclusion: TNF down-regulated the expression of bcl-2 gene in U937 cells and changed the cell cycle as a possible mechanism of its induction of apoptosis.