三氧化二砷联合伊曲康唑对KG1a细胞的协同杀伤作用

Synergistic Killing Effects of Arsenic Trioxide Combined with Itraconazole on KG1a Cells

目的:探讨三氧化二砷联合伊曲康唑对急性髓系白血病KG1a细胞株的增殖与凋亡的影响及其可能的机制。方法:用瑞氏姬姆沙染色法观察细胞形态;CCK-8检测细胞抑制率;甲基纤维素集落形成实验检测细胞集落形成能力;流式细胞术检测细胞凋亡率及细胞周期阻滞变化;Western blot检测细胞BCL-2、caspase-3、BAX、SMO、Gli1、Gli2蛋白表达。结果:三氧化二砷和伊曲康唑单用对KG1a细胞均具有增殖抑制作用,且呈时间剂量依赖性。联合用药与单药对比,前者细胞存活率、集落形成数均明显减少(P<0.05),而细胞凋亡率增高。联合用药能够增加细胞的Caspase-3和BAX两种蛋白的表达、下调BCL-2、SMO、Gli1、Gli2的表达。结论:三氧化二砷联合伊曲康唑能抑制KG1a细胞增殖及诱导其凋亡,其机制可能与Hh信号通路受抑制,Caspase-3和BAX蛋白表达上调有关。对难治性AML进行三氧化二砷与伊曲康唑联合实验性治疗提供实验数据。

Objective： To explore the effect of arsenic trioxide combined with itraconazole on proliferation and apoptosis of KG1 a cells and its potential mechanism. Methods： The cell morphology was observed with Wrighe-Giemsa staining;cell survival rate was examined by CCK-8; and colony formation capacity was measured by methylcellulose colony formation test; the flow cytometry was used to analyse the cell apoptosis rate and cell cycle; the protein expressions of BCL-2, caspase-3,BAX,SMO,Gli1 and Gli2 were detected by Western-blot. Results： The arsenic trioxide and itraconazole alone both could inhibit the KG1a cell proliferation in dose-and time-dependent manner. In comparison between single and combined drug-treatment group,both the cell survival rate and the colony number of the single drug-treatment group were significantly lower（ P〈0. 05）,and the apoptosis rate was higher in the combined drug-treatment group. In the combined-treatment group,the protein expression of Caspase-3 and BAX was upregulated,while the protein expression of BCL-2,SMO,Gli1 and Gli2 was downregulated. Conclusion： Arsenic trioxide combined with itraconazole can inhibit the KG1a cell proliferation and induce apoptosis,which may be related with the inhibition of Hh signaling pathway and upregulation of both Caspase-3 and BAX protein expression,and provided experimental data of arsenic trioxide combined with itraconazole for the treatment of refractory AML.