去铁胺诱导K562细胞凋亡研究

Deferoxamine induces apoptosis of K562 cells

摘要:

目的：探讨铁螯合剂去铁胺（DFO）诱导白血病细胞凋亡的分子机制。方法：实验分为DFO组(终浓度分别为10、50、100 μM)、DFO+FeCl3（各10 μmol/L）组及空白对照组。用钙黄绿素检测K562细胞可变铁池。锥虫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方法检测K562细胞凋亡；比色法检测caspase-3活性。结果：（1）DFO作用于K562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；而DFO+FeCl3组细胞凋亡率与空白对照组差异无统计学意义。（2）50 μmo/L、100 μmol/L DFO作用于K562细胞24 h时，caspase-3酶活性升高明显，与对照组相比，差异有统计学意义(P<0.01)；相关分析结果显示，K562细胞铁池的荧光改变与caspase-3活性变化呈负相关(r=-0.894, 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，降低细胞可变铁池，激活caspase-3有关。

Abstract:

OBJECTIVE: To study the molecular mechanism of apoptosis of leukemic cells (K562 cells) induced by iron chelating agent deferoxamine (DFO). METHODS: The exponentially growing K562 cells were used (1×106/mL) in this study. The K562 cells were treated with different concentrations of DFO (10, 50 and 100 mmol/L), DFO+FeCl3 (10 μmol/L each) or normal saline (blank control). The cellular labile iron pool was measured with a fluorimetric assay using the metalsensitive probe calcein-AM. The viable count and cell viability were determined by typanblue assay. Cell apoptosis was determined by morphological study and flow cytometry assay. Caspase-3 activity in K562 cells was detected by colorimetry. RESULTS: After DFO treatment, the cellular labile iron pool and the viability of K562 cells were reduced and the cell apoptosis increased in a time- and dose-dependent manner compared with the blank control group. The apoptosis rate of K562 cells in the DFO+FeCl3 treatment group was not significantly different from that in the blank control group. The caspase-3 activity in K562 cells increased significantly 24 hrs after 50 and 100 μmmol DFO treatment when compared with the blank control group (P<0.01). There was a negative correlation between cellular labile iron pool and caspase-3 activity of K562 cells (r=-0.894, P<0.05). CONCLUSIONS: DFO induces apoptosis of leukemic cells possibly through decreasing cellular labile iron pool and increasing caspase-3 activity of the cells.