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目的探讨铁螯合剂去铁胺(DFO)对诱导白血病细胞HL-60的分子机制。方法 2003年712月用钙黄绿素(calcein)检测HL-60细胞LIP。台盼蓝活细胞拒染实验进行活细胞计数及细胞存活率测定;光镜形态学观察及流式细胞仪(FCM)等方法检测HL-60细胞凋亡;比色法检测caspase-3(基于pNA标记底物的比色法)活性。结果①不同浓度的DFO作用于HL-60细胞后,随培养时间延长及DFO浓度的增加,动态铁池降低,细胞生存率逐渐下降,凋亡率增加,显示一定的时间剂量依赖性。②HL-60细胞在不同浓度的DFO作用下,caspase-3的活性逐渐升高。50、100μmol/LDFO作用于HL-60细胞24h,caspase-3酶活性升高明显,与对照组相比,有统计学意义(P<0.001);相关分析结果显示,HL-60细胞LIP的改变与caspase-3活性变化呈负相关系(r=-0.887,P<0.05)。结论 DFO诱导白血病细胞凋亡的作用可能与螯合细胞内铁,降低细胞LIP,激活caspase-3,最终实施细胞凋亡密切相关。
Objective To investigate the molecular mechanism of inducing leukemia cell line HL-60 by iron chelator deferoxamine (DFO). Methods In Dec. 2003, LIP of HL-60 cells was detected by calcein. Trypan blue exclusion test was used to determine the viable cell count and cell viability. The apoptosis of HL-60 cells was detected by light microscopy and flow cytometry (FCM). The levels of caspase-3 pNA labeled substrate colorimetric method) activity. Results ① After different concentrations of DFO were added into HL-60 cells, the dynamic iron pool decreased, the cell viability decreased and the apoptosis rate increased with the prolongation of culture time and the increase of DFO concentration, indicating a certain time-dependent dose-dependent manner. ②HL-60 cells under different concentrations of DFO, caspase-3 activity gradually increased. The activities of caspase-3 in HL-60 cells treated with 50 and 100μmol / L LDF for 24h were significantly higher than those in control group (P <0.001). The correlation analysis showed that the changes of LIP in HL-60 cells Negatively correlated with the activity of caspase-3 (r = -0.887, P <0.05). CONCLUSION DFO-induced apoptosis in leukemic cells may be related to chelating intracellular iron, decreasing cellular LIP, activating caspase-3, and ultimately effecting apoptosis.