总状绿绒蒿醇提物对人白血病K562细胞DNA损伤及细胞周期的影响

Influence of Meconopsis racemosa Maxim.alcohol extract in DNA injury and cell cycle of human leukemia K562 cells

目的:探讨传统藏药总状绿绒蒿体外对人慢性髓系白血病细胞株K562细胞增殖活性的影响,并阐明其相关作用机制。方法:不同浓度(30、60、90、120和150mg.L-1)总状绿绒蒿醇提物作用于体外培养的白血病K562细胞,同时设置空白对照组,MTT法检测各组细胞增殖抑制率,倒置显微镜观察细胞形态学改变,单细胞凝胶电泳检测细胞DNA损伤,流式细胞术检测细胞周期时相变化。结果:作用于K562细胞24、48和72h后,不同浓度总状绿绒蒿醇提物组K562细胞生长受到明显抑制,与空白对照组比较,其增殖抑制率明显增加(P<0.05),并呈浓度和时间依赖性。倒置显微镜下观察,随着总状绿绒蒿醇提物浓度增加,K562细胞数量减少,细胞形态不规则,轮廓模糊,细胞碎片增多。单细胞凝胶电泳(SCGE)检测,与空白对照组比较,60、90和120mg.L-1总状绿绒蒿醇提物组K562细核内DNA片段化、TailDNA%升高,差异有统计学意义(P<0.05或P<0.01)。细胞周期检测,60、90和120mg.L-1总状绿绒蒿醇提物作用于K562细胞24h后,G0/G1期、S期细胞所占比例逐渐下降,G2/M期细胞所占比例逐渐升高,分别为(1.88±0.30)%、(5.46±0.57)%和(19.80±1.03)%,各浓度组与空白对照组[(1.10±0.12)%]比较差异均有统计学意义(P<0.05),总状绿绒蒿醇提物组K562细胞周期发生了G2/M期阻滞。结论:总状绿绒蒿醇提物对K562细胞有显著的增殖抑制作用,其机制可能与诱导DNA损伤引发G2/M期阻滞有关联。

Objective To investigate the influence of Meconopsis racemosa Maxim.of a traditional Tibetan medicine in the biological activity of human leukemia cell line K562cells in vitro and to clarify its related mechanism.Methods The K562cells were treated with different concentrations（30,60,90,120,150 mg.L-1）of Meconopsis racemosa Maxim.alcohol extract in vitro,meanwhile blank control group was set up.MTT assay was used to detect the growth inhibitory rates of K562cells treated with Meconopsis racemosa Maxim.alcohol extract.The morphological changes of K562cells were observed by inverted microscope.DNA injury was evaluated by single cell gel electrophoresis assay（SCGE）.The changes of cell cycle were detected using flow cytometry.Results The inhibitory rates of K562cells proliferation in Meconopsis racemosa Maxim.alcohol extract groups were significantly higher than those in blank control group after 24,48and 72h（P0.01）in dose-dependent and time-dependent manner.The numbers of K562cells was reduced,and the cells shapes were irregular with cuntour fuzzy under inverted microscope.The SCGE results showed that nucleus DNA fragmentation and TailDNA% were increased after treated with 60,90,and 120mg.L-1 Meconopsis racemosa Maxim.alcohol extract,there were significant differences compared with blank control group（P0.05,P0.01）.The cell cycle detection results showed that in 60,90and 120mg.L-1 Meconopsis racemosa Maxim.alcohol extract groups,the percentages of K562cells at G0/G1phase and S phase were decreased,the percentages of K562cells at G2/M phase were increased（1.88%±0.30%,5.46%±0.57%,19.8%±1.03%）,they were significantly higher than that in blank control group（1.10%±0.12%）（P0.05）,which suggested a G2/M cell cycle arresting in Meconopsis racemosa Maxim.alcohol extract groups.Conclusion Meconopsis racemosa Maxim.alcohol extract could obviously inhibit the proliferation of K562cells,and the mechanism may be related to inducing DNA injury of K562cells and arresting cell cycle in G2/M phase.