细胞色素C对HL-60细胞促凋亡作用及其与相关凋亡基因的研究

Study of apoptosis effect of cytochrome C on HL-60 cells and its relationship with the relevant apoptotic gene

目的 探讨细胞色素C在体外作用于HL - 6 0细胞时细胞发生效应及其相关凋亡基因的表达变化的机制。方法 用不同浓度的细胞色素C作用于HL - 6 0细胞 2 4小时 ,然后应用光镜、荧光显微镜检测HL - 6 0细胞形态的变化 ;用流式细胞仪、DNA凝胶电泳对HL - 6 0细胞凋亡进行检测 ;用RT -PCR检测Bcl- 2、Fas基因表达的变化。结果 当细胞色素C浓度在 0～ 37.5mg L作用HL - 6 0细胞 2 4小时 ,细胞发生的凋亡逐渐增加 ,可见典型的凋亡细胞和明显的DNA梯度 ;同时 ,在该浓度范围内 ,Bcl- 2表达逐渐减少 ,而Fas表达逐渐增加 ;当细胞色素C浓度大于 37 5mg L时 ,细胞凋亡率并不增加 ,而是下降 ,且出现细胞坏死。结论 一定浓度细胞色素C能诱导HL - 6 0细胞发生凋亡 ,并且细胞凋亡率、Bcl- 2、FAS基因表达的变化与细胞色素C浓度呈一定的量效依赖关系 ,细胞色素C诱导HL - 6 0细胞凋亡可能与抑制Bcl- 2基因的表达和激活Fas基因的表达有一定的关系。

Objective To study the effect of cytochrome C on HL-60 cells in vitro and the mechanism of expression changes of relevant apoptotic genes. Methods The morphology of cell was observed by microscope and fluorescence microscope; The apoptosis was essayed by flow cytometry (FCM) and DNA electrophoresis; the expression changes of Bcl-2 and Fas mRNA were examined by RT-PCR, after HL-60 cells were treated with different concentrations of cytochtome C for 24 hours. Results When treated with 0～37.5 mg/L cytochtome C for 24 hours, the percentage of apoptotic HL-60 cells increased with the dose increasing, and the typical cells and the appearance of apoptotic DNA ladder were observed. At the same time, within this range of concentration, the expression of Bcl-2 mRNA decreased gradually and the expression of Fas increased gradually. When higher than 37.5 mg/L cytochrome C, the percentage of apoptotic HL-60 cells did not increase, but decreased, also the cells necrosised. Conclusion It suggested from the results that at certain range of concentration, apoptosis or necrosis could be induced by Cytochrome C on HL-60 cells, the percentage of apoptotic, the changes of expression of bcl-2 and Fas depended on the dose of cytochrome C. The mechanism that cytochtome C induced apoptosis on HL-60 cells may relate to suppressing the expression of bcl-2 and activating the expression of Fas.