摘要
目的探讨黄芩素和U0126诱导人乳腺癌细胞株MCF-7凋亡的分子机制。方法单独及联合使用20μmol黄芩素、10μmol U0126处理MCF-7细胞24 h;光学显微镜观察细胞数量变化,流式细胞术检测MCF-7细胞周期变化,CCK8法检测细胞增殖变化,TUNEL法和流式细胞术检测细胞凋亡,实时聚合酶链反应(Real Time PCR)和Western blot检测凋亡相关因子m RNA和蛋白的表达水平。结果与黄芩素单独刺激MCF-7细胞相比,黄芩素联合U0126刺激MCF-7细胞时,S期细胞比例降低更明显;MCF-7细胞经不同浓度黄芩素或U0126处理后,增殖抑制,且具有浓度依赖性(P<0.05);与黄芩素单独处理MCF-7细胞相比,黄芩素与U0126联合处理时,细胞凋亡更加显著(P<0.05),早期凋亡和晚期凋亡均增多(P<0.05);与黄芩素或U0126单独处理MCF-7细胞相比,黄芩素和U0126联合处理MCF-7细胞时,凋亡抑制因子Bcl-2的m RNA水平降低(P<0.05),凋亡促进因子Bax的m RNA水平增高(P<0.05),ERK1/2、GSK-3β和P38的磷酸化水平降低(P<0.05),凋亡促进因子Bax表达量增高(P<0.05),凋亡抑制因子Bcl-2表达量降低(P<0.05)。结论黄芩素或U0126通过调控Bcl-2/Bax的表达水平以及ERK1/2、GSK-3β和P38的磷酸化水平抑制MCF-7细胞增殖,促进细胞凋亡,且具有协同效应。因此,黄芩素和U0126可联合用于临床治疗乳腺癌,具有重要临床意义。
Objective To investigate the molecular mechanisms of apoptosis of human breast cancer cell line MCF-7 induced by baicalein and U0126. Methods The human breast cancer cell line MCF-7 cells were treated by 20 μM baicalein, 10 μM U0126, and 20 μM baicalein combined with 10 μM U0126 respectively. Flow cytometry was used to test the changes of MCF-7 cell cycle. CCK8 assay was applied to measure proliferation of MCF-7 cells. The number of ceils was observed under light microscope. Flow cytometry and TUNEL were used to evaluate the apoptosis of MCF-7 cells. RT-PCR and Western blot were adopted to detect the mRNA and protein levels of apoptosis-related proteins. Results Compared to the MCF-7 cells stimulated with 20 μM baicalein alone, the ratio of S phase MCF-7 cells decreased after treatment by baicalein combined with U0126 （P 〈 0.05）. When the MCF-7 cells were treated by different concentrations of baicalein or U0126, the proliferation was inhibited dramatically in a dose-dependent way （P 〈 0.05）. Compared to the MCF-7 cells treated with baicalein alone, the early and late apoptotic ceils were augmented dramatically when they were treated by both baicalein and U0126 （P 〈 0.05）. When the MCF-7 cells were treated by baicalein combined with U0126, the level of Bcl-2 decreased （P 〈 0.05）, the level of Bax greatly increased （P 〈 0.05）, and the phosphorylation levels of ERK1/2, GSK-313 and P38 obviously reduced （P 〈 0.05） compared with the cells with single treatment of baicalein or U0126. Conclusions Baicalein and U0126 can dramatically inhibit the proliferation and increase the apoptosis of MCF-7 ceils through decreasing the level of Bcl-2, reducing the phosphorylation levels of ERK1/2, GSK-313 and P38, and increasing the level of Bax. Hence, baicalein and U0126 can be used to treat breast cancer, both are very important in clinic.
出处
《中国现代医学杂志》
CAS
北大核心
2017年第10期6-13,共8页
China Journal of Modern Medicine
基金
四川省科技厅项目(14JC01353-LH67)
四川省泸州市科技局项目(2014-S-44)
