2-甲氧基雌二醇对慢性髓系白血病K562细胞Caspase-3及Survivin表达的影响

Effects of 2-methoxyestradiol on the Expression of Caspase-3 and Survivin in Chronic Myelocytic Leukemia K562 Cells

本研究探讨2-甲氧基雌二醇(2-ME)对慢性髓系白血病(CML)K562细胞caspase-3和survivin表达的影响。实验分3组:对照组,培养基中不含2-ME;实验组,分别用1、2、4、8及16μmol/L的2-ME处理K562细胞;阴性对照组,培养液中以不含RNase的灭菌蒸馏水代替K562细胞。应用TUNEL、流式细胞术(FCM)、半定量RT-PCR分别检测K562细胞凋亡率、caspase-3及survivin蛋白及其基因表达水平。结果表明:2-ME在一定的浓度范围内呈剂量依赖形式诱导K562细胞凋亡,与对照组比较差异均有统计学意义(p均<0.01),且TUNEL和FCM方法检测的K562细胞凋亡率呈正相关(γ=0.845,p=0.034)。随着2-ME浓度增加,caspase-3蛋白表达量逐渐增加,而survivin蛋白表达量逐渐降低,两者与对照组比较差异均有统计学意义(p均<0.05);且caspase-3与survivin蛋白表达量之间呈负相关(γ=-0.956,p=0.001)。随着2-ME浓度增加,caspase-3基因表达量逐渐增加,而survivin基因表达量逐渐降低,两者与对照组比较差异均有统计学意义(p均<0.01),且caspase-3与survivin基因表达量之间呈负相关(γ=-0.966,p=0.001)。结论:2-ME能够以剂量依赖形式诱导K562细胞凋亡,显示了其对CML有潜在的治疗价值。

The aim of this study was to investigate the effects of 2-methoxyestradiol （2-ME） on expressions of caspase-3 and survivin in chronic myelocytic leukemia （CML） K562 cells. The expreriment was divided into 3 groups： control group, in which K562 cells were cultured in medium without 2-ME; the experimental group, in which K562 cells were cultured in medium containing different concentrations of 2-ME （1, 2, 4, 8 and 16 μmol/L） for 36 hours; the negative control group, in which K562 cells were replaced by distilled water without RNase in medium. The apoptosis rate, the protein and its mRNA expressions of caspase-3 and survivin of K562 cells was detected by TUNEL, flow cytometry （FCM）, half-quantitative RT-PCR respectively. The results showed that the apoptosis rate of K562 cells in experimental group was significantly higher than that in control group （p 〈 0.05）. The apoptosis rate of K562 cells detected by FCM was almost the same as that detected by TUNEL method （p 〈0.01 ）. The result detected by TUNEL methods was positively correlated with that detected by FCM （ γ = 0. 845, p = 0. 034 ）. The expression of caspase-3 protein increased in a concentration-dependent manner, and also this expression level in the experimental group was higher than that in the control group（p 〈 0.05 ） ; the expression of survivin protein decreased along with the increasing of 2-ME con- centration, and the difference between the experimental group and the control group was statistically significant （p 〈 0. 05 ）. The expression of caspase-3 mRNA was higher in the experimental group than that in the control group （p 〈0. 01 ）, and the expression of survivin mRNA was lower in the experimental group than that in the control group （p 〈 0. 01）. The expression level of caspase-3 mRNA was negatively correlated with that of survivin （ γ = -0. 966, p = 0. 001 ）. It is concluded that the 2-ME can induce apoptosis of K562 cells in a concentration-dependent manner and indi- cate its promising potential in the treatment of CML patients.