目的 研究三乙酰穿心莲内酯(ADA)对人乳腺癌MCF-7细胞的增殖抑制作用和凋亡的影响,并对其作用机制进行研究和探讨。方法 采用四甲基偶氮唑蓝(MTT)法检测细胞增殖。4',6-二脒基-2-苯基吲哚(DAPI)、吖啶橙/溴乙啶(AO/EB)双染法、以及Annexin V-FITC/PI双染法流式细胞仪测定细胞凋亡。Western bolt法检测NF-κB信号通路相关蛋白NF-κB p65核位移、磷酸化IκBɑ、磷酸化IKKβ的表达以及凋亡通路中相关蛋白Bcl-2、Bax和caspase 3的表达。结果 MTT结果显示,ADA对MCF-7细胞增殖有非常显著的生长抑制作用(P<0.01),并呈剂量依赖性;ADA可以诱导MCF-7细胞凋亡,具体可能机制为MCF-7细胞caspase 3被激活并表达增加(P<0.01);促凋亡蛋白Bax的表达增加(P< 0.01),抗凋亡Bcl-2的表达降低(P< 0.05),提示ADA参与Bax、Bcl-2、caspase 3的调控,同时磷酸化IκBɑ、磷酸化IKKβ、细胞核内NF-κB p65表达均降低(P< 0.01),细胞质内p65表达增加 (P<0.01),表明ADA也参与NF-κB信号通路的调控。结论 ADA可诱导MCF-7细胞凋亡,可能参与Bax、Bcl-2、caspase 3与NF-κB信号通路的调控。
Abstract
OBJECTIVE To investigate the anti-proliferation effects of andrographolide Derivative(ADA) and its potential anti-tumor mechanisms in MCF-7 cells. METHODS The anti-proliferation effects of ADA were assessed by MTT assay in MCF-7 cells. Apoptotic morphological changes of MCF-7 cells after ADA treatment were observed by high content screening (HCS) after AO/EB double staining and DAPI, Apoptotic number was investigated by using flow cytometry. The expression of Bcl-2, Bax,caspase 3,NF-κB p65,p-IκBɑ and p-IKKβ were measured using Western blot.RESULTS MTT assay demonstrated that ADA exerted potent anti-tumor effect in a significant concentration-dependent manner;ADA could promote the apoptosis of MCF-7 cells. Western blot results showed that Bcl-2 families and NF-κB p65 and caspase 3 involved in the process of apoptosis, Bcl-2 expression lowered(P<0.05),p-IκBɑ and p-IKKβ expression decreased(P<0.01), Bax, NF-κB p65 and caspase 3 expression increased(P<0.01). CONCLUSION ADA could inhibit the proliferation of MCF-7 cells and mechanism may be that it might induce apoptosis by regulated the expression of Bax, Bcl-2 and caspase 3 protein and NF-κB passway.
关键词
三乙酰穿心莲内酯 /
乳腺癌细胞MCF-7 /
增值 /
凋亡 /
NF-κB
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Key words
andrographolide derivative /
MCF-7 cells /
proliferation /
apoptosis /
NF-κB
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中图分类号：
R965
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参考文献
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脚注
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基金
河南省科技厅基础前沿项目(142300410369)
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