β-elemene reverses the drug resistance of lung cancer A549/DDP cells via the mitochondrial apoptosis pathway
β-elemene reverses the drug resistance of lung cancer A549/DDP cells via the mitochondrial apoptosis pathway
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DOI：
10.3892/or.2014.3083
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发表时间：
2014-05-01
期刊：
影响因子：
4.2
通讯作者：
Zhang, Yi
中科院分区：
文献类型：
--
作者：
Yao, Cheng-Cai;Tu, Yuan-Rong;Zhang, Yi
β-榄香烯(beta-elemene，β-ELE)是从莪术中提取的一种新型抗癌药物，广泛用于治疗恶性肿瘤。最近的研究表明，β-ELE可以逆转肿瘤细胞的耐药性。为了探讨β-ELE的作用机制，我们观察了β-ELE对顺铂耐药的人肺腺癌A549/DDP细胞的作用。用四甲基偶氮唑盐比色法检测β-ELE对A549/DDP细胞体外生长的影响。用Hoechst 33258染色的荧光显微镜和Annexin V-FITC/PI双重染色的流式细胞仪检测细胞凋亡。用JC-1荧光探针和激光共聚焦扫描显微镜检测线粒体膜电位，用2‘，7’-二氯荧光素-双乙酸酯染色和流式细胞仪检测细胞内氧自由基水平。用GSH试剂盒测定细胞内谷胱甘肽含量。免疫印迹法检测细胞色素c、半胱氨酸天冬氨酸氨基转移酶-3、原天冬氨酸氨基转移酶-3和Bcl2家族蛋白的表达。结果表明，β-ELE对A549/DDP细胞的增殖有抑制作用，且呈时间和剂量依赖性。此外，β-ELE增强了A549/DDP细胞对顺铂的敏感性，并逆转了A549/DDP细胞的耐药性。与激活细胞凋亡的作用一致，β-ELE以时间和剂量依赖的方式降低线粒体膜电位，增加细胞内活性氧浓度，降低细胞质谷胱甘肽水平。β-ELE与顺铂联合应用可上调A549/DDP肺癌细胞中细胞色素c、caspase-3和Bad的蛋白表达，降低Bcl2和proaspase-3的蛋白水平。这些结果确定了一条原天冬氨酸氨基转移酶-3-β-ELE功能的途径，该途径涉及线粒体膜电位降低，导致细胞色素c释放到细胞质内并调节与凋亡相关的基因而触发细胞凋亡。β-ELE对A549/DDP细胞耐药的逆转作用可能与其诱导细胞凋亡有关。
beta-elemene (beta-ELE) is a new anticancer drug extracted from Curcuma zedoaria Roscoe and has been widely used to treat malignant tumors. Recent studies have demonstrated that beta-ELE reverses the drug resistance of tumor cells. To explore the possible mechanisms of action of beta-ELE, we investigated its effects on cisplatin-resistant human lung adenocarcinoma A549/DDP cells. The effects of beta-ELE on the growth of A549/DDP cells in vitro were determined by MTT assay. Apoptosis was assessed by fluorescence microscopy with Hoechst 33258 staining and flow cytometry with Annexin V-FITC/PI double staining. Mitochondrial membrane potential was assessed using JC-1 fluorescence probe and laser confocal scanning microscopy, and intracellular reactive oxygen species levels were measured by 2',7'-dichlorofluorescein-diacetate staining and flow cytometry. Cytosolic glutathione content was determined using GSH kits. The expression of cytochrome c, caspase-3, procaspase-3 and the Bcl-2 family proteins was assessed by western blotting. The results demonstrated that beta-ELE inhibited the proliferation of A549/DDP cells in a time-and dose-dependent manner. Furthermore, beta-ELE enhanced the sensitivity of A549/DDP cells to cisplatin and reversed the drug resistance of A549/DDP cells. Consistent with a role in activating apoptosis, beta-ELE decreased mitochondrial membrane potential, increased intracellular reactive oxygen species concentration and decreased the cytoplasmic glutathione levels in a time-and dose-dependent manner. The combination of beta-ELE and cisplatin enhanced the protein expression of cytochrome c, caspase-3 and Bad, and reduced protein levels of Bcl-2 and procaspase-3 in the A549/DDP lung cancer cells. These results define a pathway of procaspase-3-beta-ELE function that involves decreased mitochondrial membrane potential, leading to apoptosis triggered by the release of cytochrome c into the cytoplasm and the modulation of apoptosis-related genes. The reversal of drug resistance of the A549/DDP cell line by beta-ELE may be derived from its effect in inducing apoptosis.