Tengku Din Tengku Ahmad Damitri Al-Astani, Seeni Azman, Khairi Wirdatul-Nur Mohd, Shamsuddin Shaharum, Jaafar Hasnan
Department of Pathology, School of Medical Sciences, Universiti Sains Malaysia, Kubang Kerian, Malaysia E-mail :
Asian Pac J Cancer Prev. 2014;15(24):10659-63. doi: 10.7314/apjcp.2014.15.24.10659.
Rapamycin is an effective anti-angiogenic drug. However, the mode of its action remains unclear. Therefore, in this study, we aimed to elucidate the antitumor mechanism of rapamycin, hypothetically via apoptotic promotion, using MCF-7 breast cancer cells.
MCF-7 cells were plated at a density of 15105 cells/well in 6-well plates. After 24h, cells were treated with a series of concentrations of rapamycin while only adding DMEM medium with PEG for the control regiment and grown at 37oC, 5% CO2 and 95% air for 72h. Trypan blue was used to determine the cell viability and proliferation. Untreated and rapamycin-treated MCF-7 cells were also examined for morphological changes with an inverted-phase contrast microscope. Alteration in cell morphology was ascertained, along with a stage in the cell cycle and proliferation. In addition, cytotoxicity testing was performed using normal mouse breast mammary pads.
Our results clearly showed that rapamycin exhibited inhibitory activity on MCF-7 cell lines. The IC50 value of rapamycin on the MCF-7 cells was determined as 0.4μg/ml (p<0.05). Direct observation by inverted microscopy demonstrated that the MCF-7 cells treated with rapamycin showed characteristic features of apoptosis including cell shrinkage, vascularization and autophagy. Cells underwent early apoptosis up to 24% after 72h. Analysis of the cell cycle showed an increase in the G0G1 phase cell population and a corresponding decrease in the S and G2M phase populations, from 81.5% to 91.3% and 17.3% to 7.9%, respectively.
This study demonstrated that rapamycin may potentially act as an anti-cancer agent via the inhibition of growth with some morphological changes of the MCF-7 cancer cells, arrest cell cycle progression at G0/G1 phase and induction of apoptosis in late stage of apoptosis. Further studies are needed to further characterize the mode of action of rapamycin as an anti-cancer agent.
雷帕霉素是一种有效的抗血管生成药物。然而，其作用方式仍不清楚。因此，在本研究中，我们旨在使用MCF-7乳腺癌细胞，假设通过促进凋亡来阐明雷帕霉素的抗肿瘤机制。
将MCF-7细胞以15×10⁵个细胞/孔的密度接种于6孔板中。24小时后，用一系列浓度的雷帕霉素处理细胞，而对照组仅添加含有聚乙二醇的DMEM培养基，并在37℃、5%二氧化碳和95%空气条件下培养72小时。用台盼蓝测定细胞活力和增殖情况。还使用倒置相差显微镜检查未处理和经雷帕霉素处理的MCF-7细胞的形态变化。确定细胞形态的改变以及细胞周期和增殖阶段。此外，使用正常小鼠乳腺垫进行细胞毒性测试。
我们的结果清楚地表明，雷帕霉素对MCF-7细胞系具有抑制活性。雷帕霉素对MCF-7细胞的IC50值确定为0.4μg/ml（p<0.05）。倒置显微镜直接观察表明，经雷帕霉素处理的MCF-7细胞表现出凋亡的特征，包括细胞收缩、血管化和自噬。72小时后，细胞发生早期凋亡的比例高达24%。细胞周期分析显示，G0/G1期细胞群体增加，S期和G2/M期群体相应减少，分别从81.5%降至91.3%和从17.3%降至7.9%。
本研究表明，雷帕霉素可能通过抑制生长、使MCF-7癌细胞发生一些形态变化、使细胞周期停滞在G0/G1期以及诱导晚期凋亡来潜在地发挥抗癌作用。需要进一步研究以进一步阐明雷帕霉素作为抗癌药物的作用方式。
