葫芦素 B 通过抑制粘着斑激酶减少胆管癌细胞的转移行为。
Cucurbitacin B Diminishes Metastatic Behavior of Cholangiocarcinoma Cells by Suppressing Focal Adhesion Kinase.
机构信息
Department of Pharmacology, Faculty of Medicine, and Cholangiocarcinoma Research Institute, Khon Kaen University, Khon Kaen, Thailand.
出版信息
Asian Pac J Cancer Prev. 2021 Jan 1;22(1):219-225. doi: 10.31557/APJCP.2021.22.1.219.
OBJECTIVE
Cholangiocarcinoma (CCA) is a malignant tumor with aggressive metastatic property resulted from dysregulation of metastasis-regulated signaling pathways. The aim of this study was to investigate the effect of cucurbitacin B on metastatic behavior of CCA cells through modulation of focal adhesion kinase (FAK) protein.
METHODS
KKU-452 cells were treated with a specific FAK inhibitor, FAK inhibitor-14, or cucurbitacin B at various concentrations for 24 h. Cell viability was assessed by sulforhodamine B assay. The migratory and invasive abilities of the cells were investigated using wound healing and transwell invasion assays, respectively. The fibronectin-coated plate was used for adhesion assay. The effects of the test compounds on FAK activation and the expression of metastasis-associated proteins were determined by Western blot analysis. The amount of MMP-9 was evaluated using a commercial ELISA Kit.
RESULTS
FAK inhibitor-14 and cucurbitacin B at concentrations which minimally affected KKU-452 cell viability could suppress FAK activation, evidently by decreased level of phospho-FAK protein after exposure to the compound. At these conditions, cucurbitacin B suppressed metastatic behavior including migration, invasion and adhesion abilities of CCA cells similar to FAK inhibitor-14. Further molecular studies demonstrated that FAK inhibitor-14 and cucurbitacin B downregulated the expression of metastasis-associated proteins including MMP-9, ICAM-1 and VEGF. Consequently, exposure to cucurbitacin B inhibited the production of MMP-9 enzyme in CCA cells similar to FAK inhibitor-14 treatment.
CONCLUSION
FAK participated in regulation of metastatic behavior of KKU-452 CCA cells. Cucurbitacin B suppressed FAK activation in the cells which was associated with inhibition of metastasis essential steps and their related metastatic proteins. The compound may be developed as a novel therapeutic agent for CCA metastasis therapy.
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目的
胆管癌（CCA）是一种恶性肿瘤，具有侵袭性转移特性，是由于转移调节信号通路失调所致。本研究旨在通过调节粘着斑激酶（FAK）蛋白，研究葫芦素 B 对 CCA 细胞转移行为的影响。
方法：用特定的 FAK 抑制剂 FAK 抑制剂-14 或葫芦素 B 处理 KKU-452 细胞，浓度为 24 小时。用磺酰罗丹明 B 法测定细胞活力。用划痕愈合和 Transwell 侵袭实验分别检测细胞的迁移和侵袭能力。用纤维连接蛋白包被板进行黏附实验。用 Western blot 分析测定测试化合物对 FAK 激活和转移相关蛋白表达的影响。用商业 ELISA 试剂盒评估 MMP-9 的量。
结果：FAK 抑制剂-14 和葫芦素 B 的浓度对 KKU-452 细胞活力的影响最小，可抑制 FAK 激活，明显表现为暴露于化合物后磷酸化 FAK 蛋白水平降低。在这些条件下，葫芦素 B 抑制了 CCA 细胞的转移行为，包括迁移、侵袭和黏附能力，与 FAK 抑制剂-14 相似。进一步的分子研究表明，FAK 抑制剂-14 和葫芦素 B 下调了包括 MMP-9、ICAM-1 和 VEGF 在内的转移相关蛋白的表达。因此，暴露于葫芦素 B 可抑制 CCA 细胞中 MMP-9 酶的产生，与 FAK 抑制剂-14 处理相似。
结论：FAK 参与了 KKU-452 CCA 细胞转移行为的调节。葫芦素 B 抑制了细胞中的 FAK 激活，这与抑制转移的基本步骤及其相关的转移蛋白有关。该化合物可能被开发为 CCA 转移治疗的新型治疗剂。
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