去甲氧柔红霉素对耐药白血病细胞中Chk1磷酸化水平的影响

The study of effect of idarubicin on the expression of phosphorylated-Chk1 in drug-resistant acute leukemia cells

目的:观察去甲氧柔红霉素(IDA)对耐药急性白血病细胞中细胞周期检测点激酶1(Chk1)磷酸化水平的影响,探讨Chk1信号通路在去甲氧柔红霉素抗瘤作用中的机制。方法:MTT法检测柔红霉素(DNR)和IDA的IC50,耐药白血病细胞分别与DNR和IDA的IC50共孵育,Western-blot检测细胞内Chk1磷酸化水平(P-Chk1)的表达,流式细胞术检测其细胞周期及细胞凋亡率。结果:DNR的IC50约为0.40μg/ml,IDA约为0.12μg/ml,以此浓度分别与AML细胞共孵育,6h后两组间S期、G2/M期细胞的百分率差异无显著性;24h后,DNR组S期和G2/M期细胞百分率分别为(3.56±1.17)%、(53.66±1.75)%,IDA组为(41.42±1.17)%、(28.45±3.72)%,差异有显著性;P-Chk1在DNR组为1.78±1.10,IDA组为0.56±1.25,两者比较差异有显著性;24h后,DNR组和IDA组的细胞凋亡率分别为(24.53±1.30)%、(66.47±2.61)%,IDA组明显高于DNR组,差异有显著性。结论:IDA可能通过下调P-Chk1水平延迟耐药白血病细胞的S期进程,从而增加自身的药物敏感性。

Objective：To explore the mechanisms of Chk1 signaling pathway involved in anti-leukemia ctivity of Idarubicin.Methods： IC50 of daunorubicin（DNR） and idarubicin（IDA） was detected by MTT.After treatment with daunorubicin and idarubicin respectively,Chk1 phosphorylation levels in acute leukemia cells was detected by Western-blot.The cell cycle distribution and cell apoptosis were detected by flow cytometry.Results： The 50% inhibition concentration（IC50） of DNR was about 0.40μg/ml,IDA was about 0.12μg/ml.Six hours after incubation with IC50 of anthracycline antibiotics individually,the proportion of the cell in S and G2/M phase between two group had no significant difference.24 hours after treatment,the proportion of the cell in G2/M phase for DNR was significantly higher than for IDA,the proportion of the cell in S phase for IDA was significantly higher than for DNR;the decreased Chk1 phosphorylation following DNA damage induced by IDA was observed in refractory cells（0.56±1.25）,significantly lower than those induced by DNR（1.78±1.10）;the cell apoptosis rates induced by IDA was（66.47±2.61）%,significantly higher than those induced by DNR（24.53±1.30）%.Conclusion： Idarubicin possibly delay progression of S phase in drug-resistant leukemia cells through downregulation of phosphorylated-Chk1 level,thus increasing its susceptibility.