| 黄酮联合阿霉素协同诱导HL60/ADR细胞凋亡机制的研究 |
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| 引用本文： | 陈芳源,张旻玥,蔡佳翌,沈莉菁,钟济华,曹兰芳. 黄酮联合阿霉素协同诱导HL60/ADR细胞凋亡机制的研究[J]. 诊断学理论与实践, 2014, 13(2): 159-165. DOI: 10.16150/j.1671-2870.a0361 |
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| 作者姓名： | 陈芳源 张旻玥 蔡佳翌 沈莉菁 钟济华 曹兰芳 |
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| 作者单位： | 上海交通大学医学院附属仁济医院血液科; |
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| 基金项目： | 国家自然科学青年基金（81300405）; 上海市卫生局重大课题（ZYSNXD-CC-ZDYJ001）; 上海市科委中医引导项目（12401906700） |
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| 摘 要： | 目的:探究黄酮在体外对多药耐药细胞系HL60/ADR的影响及联合阿霉素后对细胞生长抑制和凋亡的作用机制,探讨中药在白血病化疗中扶正、增效的可能性。方法:用CCK-8法检测黄酮和阿霉素对HL60/ADR细胞生长的抑制作用;用流式细胞仪检测药物作用后的细胞凋亡率,并在光镜下观察细胞形态变化;蛋白印迹法检测药物作用后凋亡信号通路蛋白表达的情况;流式细胞仪检测药物作用后HL60/ADR线粒体跨膜电位的变化。结果:黄酮能显著抑制HL60/ADR细胞的增殖,且其抑制细胞增殖效应呈现浓度-时间依赖性;不同浓度的黄酮及联合阿霉素1.5μg/mL(20%抑制浓度即IC20值)后对细胞的抑制作用更明显,具有协同和相加作用。75μg/mL和100μg/mL黄酮能促进HL60/ADR细胞凋亡,而黄酮联合阿霉素的促凋亡作用更显著。蛋白信号通路研究显示,单药黄酮及两药联合能使HL60/ADR细胞线粒体膜电位下降,抗凋亡蛋白Bcl-2、Bcl-XL表达明显下调,促凋亡蛋白Bim、Bad、Bax明显增加,激活caspase途径;同时磷酸化的P-JNK蛋白表达水平升高,而P-ERK明显下降。结论:黄酮联合阿霉素可通过线粒体跨膜电位的下降,依赖caspase活化调控Bcl-2家族中Bim、Bad和Bax蛋白表达,下调ERK细胞保护通路并上调JNK应激相关通路,最终协同抑制HL60/ADR细胞增殖并诱导其凋亡。
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| 关 键 词： | 黄酮 阿霉素 凋亡 HL60/ADR细胞 |
Study on mechanism of apoptosis induced by combination of flavonoid and adriamycin in HL60/ADR cells |
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| CHEN Fangyuan,ZHANG Minyue,CAI Jiayi,SHEN Lijing,ZHONG Jihua,CAO Lanfang. Study on mechanism of apoptosis induced by combination of flavonoid and adriamycin in HL60/ADR cells[J]. Journal of Diagnostics Concepts & Practice, 2014, 13(2): 159-165. DOI: 10.16150/j.1671-2870.a0361 |
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| Authors: | CHEN Fangyuan ZHANG Minyue CAI Jiayi SHEN Lijing ZHONG Jihua CAO Lanfang |
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| Affiliation: | . (Department of Hematology, Renji Hospital, Shanghai Jiaotong University School of Medicine, Shanghai 200127, China) |
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| Abstract: | Objective： To study the effect and mechanism of flavonoid and flavonoid in combination with adrimycin on muhidrug resistant HL60/ADR cells in vitro for exploring the possible effect of Chinese traditional medicine in leukemia chemotherapy. Methods： HL60/ADR cells were treated with adriamycin, flavonoid and flavonoid combined with adriamycin, respectively. CCK-8 kit was used to detect the growth inhibition effect on HL60/ADR cells. Apoptosis rate and change of mitochondrial transmembrane potential were detected by flow cytometry, morphologic change was examined under light microscope, and the expression of apoptosis signaling pathway was determined by Western blotting. Results： Flavonoid could significantly inhibit the proliferation of HL60/ADR cells in a dose-and time-dependent manner. The inhibitory effect on cell proliferation was enhanced when cells were treated with combined use of different concentrations of flavonoid and 1.5 μg/mL adriamycin （IC20）, showing a synergistic and additive effect. Flavonoid at 75 μg/mL and 100 txg/mL could induce cell apoptosis in HL60/ADR cells, and the apoptosis was enhanced when flavonoid was used in combination with adriamycin. Flavonoid alone and flavonoid combined with adriamycin could decrease the mitochondrial membrane potential of HL60/ADR cells, down-regulate the expression of anti-apoptotic protein Bcl-2, Bel-XL and upregulate the expression of pro-apoptotie protein Bim, Bad, Bax. The easpase pathway was activated. The expression of phosphorylated P-JNK protein level was increased and P-ERK was decreased significantly. Conclusions： Combined use of flavonoids and adriamycin could synergistically inhibit cell proliferation and induce apoptosis in HL60/ADR cells by decreasing mitochondrial membrane potential, activating caspase pathway, regulating expression of Bcl-2 family and Bim, Bad, and Bax proteins, down-regulating ERK cell protective pathway and up-regulating JNK stress related pathway. |
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| Keywords: | Flavonoid Adriamycin Apoptosis HL60/ADR cells |
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