白花蛇舌草对香豆酸组合物对急性髓系白血病细胞Kasumi-1生长抑制作用

Inhibitory Effect of Total Coumarins of Hedyotis diffusa Willd on the Growth of AML Cell Line Kasumi-1

目的研究白花蛇舌草对香豆酸组合物(total coumarins of Hedyotis diffusa Willd,TCHDW)对急性髓系白血病Kasumi-1细胞株生长抑制作用。方法采用不同浓度的TCHDW(0.02、0.04、0.06、0.08、0.10 mg/m L),作用于对数生长期的Kasumi-1细胞,MTT法检测TCHDW对Kasumi-1细胞株作用24、48 h后的生长抑制作用。再以经一定浓度(0、0.02、0.04、0.06 mg/m L)的TCHDW处理2 4 h后的Kasumi-1细胞为对象,采用Hoechst33258染色法观察Kasumi-1细胞凋亡的形态学改变,Western Blot检测Kasumi-1细胞中有关凋亡信号传导通路蛋白Caspase家族、Bcl-2家族、凋亡抑制蛋白IAPs家族、MAPKs信号通路蛋白的变化情况。结果 0.0 2~0.1 0 mg/m L的TCHDW可明显抑制白血病Kasumi-1细胞株的增殖,与空白组比较,细胞存活率明显降低(P<0.01),呈时间-浓度依赖性;Kasumi-1细胞核内凋亡小体随着药物浓度的增加逐渐增多。与空白组比较,TCHDW0.06 mg/m L组Caspase家族蛋白、Bak、Bax、Smac、p-ERK、Ras、p-P38、p-JNK升高(P<0.01,P<0.05);Bcl-2、Bcl-XL、Bim、Mcl-1、XIAP、CIAP1、CIAP2、Survivin、JNK降低(P<0.01,P<0.05)。结论 TCHDW可抑制Kasumi-1细胞增殖并诱导其凋亡,其凋亡机制与Caspase家族蛋白、Bcl-2家族蛋白和IAPs家族蛋白表达变化以及MAPKs信号通路的激活相关。

Objective To explore the growth-inhibition of total coumarins of Hedyotis diffusa Willd（ TCHDW） on AML Cell Line Kasumi-1. Methods Kasumi-1 cells at logarithmic growth phase were treated with TCHDW in various concentrations（ 0. 02,0. 04,0. 06,0. 08,0. 10 mg/m L） for different duration（ 24 or 48 h）. The inhibitive effect of TCHDW on cell growth was determined by MTT method. After Kasumi-1 cells were incubated with TCHDW（ 0 、0. 02 、0. 04 、0. 06 mg/m L） for 24 h,the apoptosis of cells were observed by Hoechst33258 staining. The expression levels of Caspase family,Bcl-2 family,IAPs family and MAPKs survival signaling pathways were examined by Western Blot. Results The proliferation of Kasumi-1cells was significantly inhibited after treatment with TCHDW（ 0. 02-0. 10 mg/m L）,which was lower than that of the control group（ P 0. 01）,and showed a dose-and time-dependent manner. After treated with TCHDW,apoptotic body appeared in Kasumi-1 cells nucleus and the number of apoptotic body increased in adose-dependent manner. Compared with control group,Caspase family proteins,Bak,Bax,Smac,p-ERK,Ras,p-P38,p-JNK inceased（ P 0. 01,P 0. 05）; Bcl-2,Bcl-XL,Bim,Mcl-1,XIAP,CIAP1,CIAP2,Survivin,JNK decreased（P 0. 01,P 0. 05）. Conclusion TCHDW could significantly inhibit the proliferation of Kasumi-1 cells and induce its apoptosis. Its mechanism may involve the Caspase,Bcl-2 and IAPs family proteins,and the MAPK signaling pathways.