Department of Hematology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
J BUON. 2020 May-Jun;25(3):1607-1613.
PURPOSE: Acute myeloid leukemia (AML) is the most frequent leukemia identified in 25% of adults and in 15-20% of children. In the current study, the cytotoxicity and apoptosis-inducing properties of davanone - a terpenoid, were examined on human AML cell line NCI-H526 and normal AML-193 cell line. METHODS: The cytotoxic effects were examined through MTT assay and apoptosis was studied through DAPI and Annexin V/PI staining. Further, the effect on MMP and ROS levels were investigated through flow cytometry. Cell migration and invasion were determined by wound healing and cell invasion assays respectively. Western blotting analysis was performed to study the expressions of apoptosis and PI3K/AKT/MAPK signalling pathway associated proteins. RESULTS: The results revealed that Davanone induced cytotoxicity in NCI-H526 cells in a dose-dependent manner without causing too much toxicity to the normal AML-193 cells. Further investigations were done in order to validate whether the cytotoxicity was apoptosis-mediated, and the results revealed that cytotoxicity of the test molecule was apoptosis-dependent. On further investigations through western blotting analysis, cytotoxicity was shown to be due to caspase-dependent apoptosis with increased expressions of caspase-3 and Bax and decreased expressions of Bcl-2. Next, it was seen that Davanone treatment led to decrease in mitochondrial membrane potential (MMP) and an increase in reactive oxygen species formation (ROS). The tested molecule also significantly suppressed cell invasion and migration of leukemia cells. Finally, the effect on PI3K/AKT/MAPK signalling pathway was examined and the expression of related proteins was altered significantly. CONCLUSIONS: The present study outcomes propose that Davanone terpenoid could be considered as a promising anticancer agent for AML.
目的：急性髓细胞白血病（AML）是成年人中最常见的白血病，占 25%，在儿童中占 15-20%。在本研究中，考察了倍半萜达瓦酮对人 AML 细胞系 NCI-H526 和正常 AML-193 细胞系的细胞毒性和诱导细胞凋亡的特性。
方法：通过 MTT 测定法检测细胞毒性，通过 DAPI 和 Annexin V/PI 染色研究细胞凋亡。进一步通过流式细胞术研究对 MMP 和 ROS 水平的影响。通过划痕愈合和细胞侵袭测定分别确定细胞迁移和侵袭。通过 Western blot 分析研究凋亡和 PI3K/AKT/MAPK 信号通路相关蛋白的表达。
结果：结果表明，达瓦酮以剂量依赖的方式诱导 NCI-H526 细胞的细胞毒性，而对正常 AML-193 细胞没有太大毒性。进一步的研究旨在验证细胞毒性是否为凋亡介导的，结果表明该测试分子的细胞毒性是凋亡依赖性的。通过 Western blot 分析进一步研究，细胞毒性是由于 caspase 依赖性凋亡引起的，caspase-3 和 Bax 的表达增加，Bcl-2 的表达减少。接下来，研究表明达瓦酮处理导致线粒体膜电位（MMP）降低和活性氧（ROS）形成增加。测试分子还显著抑制白血病细胞的侵袭和迁移。最后，研究了对 PI3K/AKT/MAPK 信号通路的影响，相关蛋白的表达发生了明显改变。
结论：本研究结果表明，达瓦酮倍半萜可被视为 AML 的一种有前途的抗癌药物。
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