姜黄素联合阿糖胞苷对人急性髓系白血病细胞KG1a增殖、自噬及凋亡的影响

Effect of curcumin combined with cytarabine on proliferation,autophagy and apoptosis of human acute myeloid leukemia cell line KG1a

目的探讨姜黄素(CUR)联合阿糖胞苷(Ara-C)对人急性髓系白血病KG1a细胞株增殖、凋亡的影响,及与自噬间的联系。方法 MTT法筛选CUR及Ara-C的最佳联合浓度并检测联合效应,分析CUR和Ara-C对KG1a细胞的增殖、自噬、凋亡和细胞周期的影响,评估CUR和Ara-C的联合用药效果。结果 CUR和Ara-C均对KG1a细胞的增殖有明显抑制作用(P<0.05),且呈一定的剂量、时间依赖性。经40μmol·L^(-1)CUR和0.5μmol·L^(-1)Ara-C联合处理的细胞抑制率明显高于各自剂量的单用组,经3-MA预处理的细胞存活率明显降低(P<0.05)。吖啶橙染色观察到细胞内出现自噬泡,联合用药的表达率高于单用组,且能被3-MA抑制。联合组细胞的凋亡率高于单用组,经3-MA预处理组细胞凋亡率均高于各自单用组(P<0.05)。细胞在G_0/G_1期比例明显多于S期。caspase-3、LC3、Beclin-1基因的表达上调,Bcl-2基因则下调(P<0.05)。联合组caspase-3和Beclin-1的蛋白量明显高于单用组(P<0.05),LC3-Ⅱ/LC3-Ⅰ的比值升高。3-MA预处理组的Beclin-1表达降低,caspase-3的表达则升高(P<0.05)。结论姜黄素能诱导KG1a细胞自噬与凋亡,并能提高白血病细胞对阿糖胞苷的敏感性,自噬抑制剂3-MA不仅能抑制该自噬,而且能促进其凋亡。

Aim To investigate the effect of curcumin（ CUR） combined with cytarabine（ Ara-C） on the proliferation and apoptosis of human acute myeloid leukemia cell line KG1a and its relationship with autophagy.Methods The optimal combination concentration of curcumin and cytarabine was screened by MTT method and the combined effects were detected.The effects of CUR and Ara-C on the proliferation,autophagy,apoptosis and cycle of KG1a cells were analyzed.Results Both CUR and Ara-C significantly inhibited the proliferation of KG1a cells（ P〈0.05）,and showed a doseand time-dependent manner.The inhibition rate of cells treated with 40 μmol·L^（-1） CUR and 0.5 μmol·L^（-1） Ara-C was significantly higher than that of the other doses alone.The survival rate of cells pretreated with3-MA was significantly decreased（ P〈0.05）.Autophagic vacuoles was observed in cells with Acridine orange staining methods,the expression rate of the combination group was higher than the single group,and can be inhibited by 3-MA.The apoptosis rate of the combined group was higher than that of the single group.The apoptosis rate of the 3-MA pretreatment group was higher than that of the single group（ P〈0.05）.Cell numbers of the G_0/G_1 phase were significantly more than the S phase.The expression of caspase-3,LC3 and Beclin-1 were up-regulated while the Bcl-2 was down-regulated（ P〈0.05）.The protein level of caspase-3 and Beclin-1 of the combination group was significantly higher than that of the single group（ P〈0.05）,and the ratio of LC3-Ⅱ/LC3-Ⅰ was increased.The Beclin-1 expression and caspase-3 expression in 3-MA pretreatment group decreased（ P〈0.05）.Conclusion Curcumin can induce autophagy and apoptosis of KG1a cells and increase the sensitivity of leukemic cells to cytarabine.Autophagy inhibitor 3-MA can not only inhibit the autophagy but also promote apoptosis.