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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 study the inhibitory effect of total coumarins of Hedyotis diffusa Willd (TCHDW) on the growth of Kasumi-1 cell line of acute myeloid leukemia. Methods Kasumi-1 cells were treated with different concentrations of TCHDW (0.02, 0.04, 0.06, 0.08 and 0.10 mg / mL) in logarithmic growth phase. MTT assay was performed on Kasumi-1 cells for 24 and 48 h Growth inhibitory effect. Kasumi-1 cells were treated with TCHDW (0, 0.02, 0.04 and 0.06 mg / mL) for 24 h. The morphological changes of Kasumi-1 cells were observed by Hoechst 33258 staining. Blot was used to detect the changes of apoptosis-related signal pathway protein Caspase family, Bcl-2 family, apoptosis-inhibitory protein IAPs family and MAPKs signaling pathway in Kasumi-1 cells. Results TCHDW 0.0-2 ~ 0.1 0 mg / m L could significantly inhibit the proliferation of Kasumi-1 leukemia cell line. Compared with the blank group, the cell survival rate was significantly decreased (P <0.01) Apoptotic bodies in the nucleus gradually increased with the increase of drug concentration. Compared with blank group, Caspase family protein, Bak, Bax, Smac, p-ERK, Ras, p-P38 and p-JNK in TCHDW0.06 mg / m L group were significantly increased (P < 2, Bcl-XL, Bcl-2, Mcl-1, XIAP, CIAP1, CIAP2, Survivin and JNK decreased (P <0.01, P <0.05). Conclusion TCHDW can inhibit the proliferation and induce the apoptosis of Kasumi-1 cells. The mechanism of apoptosis is related to the changes of Caspase family proteins, Bcl-2 family proteins and IAPs family proteins and the activation of MAPKs signaling pathway.