School of Pharmacy, Bengbu Medical University, 2600 Donghai Road, Bengbu, Anhui, China.
Department of Medicinal Chemistry, School of Pharmacy, Bengbu Medical University, 2600 Donghai Road, Bengbu, Anhui, China.
Asian Pac J Cancer Prev. 2020 Apr 1;21(4):1073-1080. doi: 10.31557/APJCP.2020.21.4.1073.
A dimeric neolignan, bishonokiol A (BHNKA) isolated from Magnolia grandiflora, significantly inhibits the proliferation of human breast cancer cells. However, the exact mechanism of BHNKA induced breast cancer cell death is unknown. In this study, we investigated the pharmacological mechanism underlying BHNKA induced MCF-7 cell death.
Cell viability measurement was performed by the MTT assay. Flow cytometry with PI staining, DAPI staining, and electron microscopy were used to analyze cellular death modes. In addition, western blotting, siRNA transfection, ATP assay, and fluorescence microscopy were used to determine the mechanism of BHNKA induced MCF-7 cell death.
BHNKA induced cell death by apoptosis, necroptosis and autophagy at the same concentration and time in MCF-7 cells, and electron microscopy confirmed these results. The mechanism of BHNKA triggered apoptosis and autophagy in MCF-7 cells was primarily due to an increase in the Bax/Bcl-2 ratio and simultaneous up-regulation of LC3-II protein expression, respectively. BHNKA induced necroptosis by activation of the RIP1-RIP3-MLKL necroptosis cascade, up-regulation of cyclophilin D (CypD) protein expression to stimulate ROS generation. We further demonstrated that siRNA-mediated down-regulation of CypD protected against BHNKA induced cell death.
These results suggest that BHNKA may be a potential lead compound for development as an anti-breast cancer agent for induction of multiple cell death pathways.
从木兰科植物中分离得到的二聚新木脂素，毕赤诺克醇 A（BHNKA），显著抑制人乳腺癌细胞的增殖。然而，BHNKA 诱导乳腺癌细胞死亡的确切机制尚不清楚。在这项研究中，我们研究了 BHNKA 诱导 MCF-7 细胞死亡的药理学机制。
通过 MTT 法测定细胞活力。用 PI 染色、DAPI 染色和电子显微镜分析细胞死亡模式。此外，还通过 Western blot、siRNA 转染、ATP 测定和荧光显微镜来确定 BHNKA 诱导 MCF-7 细胞死亡的机制。
BHNKA 在相同浓度和时间下诱导 MCF-7 细胞发生凋亡、坏死和自噬性细胞死亡，电子显微镜证实了这些结果。BHNKA 诱导 MCF-7 细胞凋亡和自噬的机制主要是由于 Bax/Bcl-2 比值增加和 LC3-II 蛋白表达同时上调。BHNKA 通过激活 RIP1-RIP3-MLKL 坏死性细胞死亡级联反应诱导坏死性细胞死亡，上调 CypD 蛋白表达以刺激 ROS 生成。我们进一步证明，CypD 的 siRNA 介导下调可防止 BHNKA 诱导的细胞死亡。
这些结果表明，BHNKA 可能是一种有潜力的先导化合物，可用于开发诱导多种细胞死亡途径的抗乳腺癌药物。
