齐墩果酸通过MAPK／ERK信号传导通路对U87MG细胞增殖及凋亡的体外实验研究

Oleanolic acid by MAPK/ERK signal transduction pathway in the proliferation and apoptosis of U87 MG cell in vitro

目的探讨齐墩果酸（OA）对神经胶质母细胞瘤U87MG细胞增殖及细胞凋亡的影响。方法以U87MG细胞为研究对象，使用25、50、80、100、150和200μg／ml的OA处理24、48和72h后，采用MTF实验检测OA对体外培养U87MG细胞增殖抑制作用，计算其抑制率；采用划痕法检测OA对U87MG细胞迁移能力的影响；用流式细胞仪分析OA对U87MG细胞周期与凋亡的影响；通过Westernblot检测OA对MAPK／ERK信号传导通路活性的影响。结果50μg/ml以上浓度的OA处理48h后，U87MG细胞的形态发生不同程度的变化，大量细胞脱落。150μg／ml和200μg／ml的OA处理U87MG细胞72h后，生长抑制率可达100％，与对照组相比差异有统计学意义（P〈0．01）。25μg/ml的OA处理48h后，迁移至划痕区的细胞数量相比于对照组明显减少。经50μg/mlOA作用48h后，处于G0／G0期的细胞从64．23％上升至75．03％，说明发生G0／G1期阻滞。同一浓度下，细胞凋亡率达到18．77％。U87MG细胞中MEK和ERK蛋白的磷酸化水平受到明显的抑制。结论OA可显著抑制U87MG细胞的生长增殖，呈一定的量效关系；OA可明显抑制U87MG细胞的迁移，并能抑制MAPK／ERK信号传导通路的激活。OA通过抑制MAPK／ERK信号传导通路的激活，可以抑制神经胶质母细胞瘤细胞的增殖生长。

Objective To investigate the role of oleanolic acid on the proliferation and Suppression of the migration of Malignant Glioma through the Deactivating MAPK/ERK Signaling Pathway in human Glioblastoma cell line U87. Methods The human C, lioblastoma cells of U87 were treated with 25 μg/ml,50 μg/ml,80 μg/ml, 100 μg/ml,150 μg/ml, and 200 μg/ml of OA for 24 h,48 h,and 72 h. Cell viability of the proliferation was assessed with MTT assay. The migration capabilities were observed with the scratch test. Flow cytometry was adopted to analyze cell cycle and apoptosis. Western blotting was used to detect the activation of the MAPK/ERK Signaling Pathway. Results The morphology of Ug7 MG ceils changed with extensive cell detachment, after 48 h treatment of 50 μg/ml and a higher concentration of OA. Its growth inhibitory rates on U87 MG cells were up to 100% under 72 h treatment on 150 μg/ml and 200 μg/ml, and compared with control group P 〈0. 01. The 48 h treatment of 25 μg/ml OA reduced the amount of cells nligrating into the wounding area. The percentage of cells undergoing G0/G1 phase rose from 64. 23% to 75. 03% after the 48 h treatment of 50μg/ral OA, indicating that G0/G1 arrest took place. At the same concentration, the apoptotic rate of U87 MG cells was 18. 77%. The phosphorylations of MEK and ERK were both suppressed in U87 MG ceils. Conclusion OA can significantly inhibit the growth of U87 cell proliferation, a certain quantity effect characteristic; it can obviously inhibit U87 cell migration, and inhibit the MAPK/ERK signaling pathways leading to the activation. Oleanolic acid has the effects of inhibition on U87 cell proliferation and migration capabilities, also it can induct cell apoptosis through deactivating the MAPK/ERK signaling pathway.