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目的:对吉祥草乙醇提取物(Reineckia carnea ethanol extracts,RCEE)诱导人肺癌A549细胞凋亡及其机制做初步研究。方法:采用cell counting kit-8试剂盒测定细胞存活率,AnnexinⅤ-FITC/PI双标后流式细胞仪检测细胞凋亡率;分别采用FLUO-8 AM、JC-1染色标记后流式细胞仪测定细胞内钙离子浓度和细胞线粒体膜电位,化学发光法检测Casapse-3/7活性。结果:RCEE对A549细胞增值的抑制作用较强且呈量效关系,作用8 h时IC50值为48.14 mg·L-1;RCEE能诱导A549细胞凋亡并呈剂量依赖关系;RCEE作用后A549细胞内游离钙离子水平升高、线粒体膜电位下降、Caspase-3/7活性升高;细胞内钙离子螯合剂BAPTA-AM和Caspase广谱抑制剂Z-VAD-FMK可以明显抑制RCEE诱导的A549细胞凋亡。结论:RCEE能抑制A549细胞增殖并诱导细胞凋亡,其机制可能与细胞内钙超载、线粒体膜电位下降、Caspase级联酶激活有关。
OBJECTIVE: To study the apoptosis of human lung cancer A549 cells induced by Reineckia carnea ethanol extract (RCEE) and its mechanism. Methods: The cell survival rate was determined by cell counting kit-8 kit. Flow cytometry was used to detect the cell apoptosis rate by AnnexinⅤ-FITC / PI double labeling. FLUO-8 AM, JC- The intracellular calcium concentration and mitochondrial membrane potential were measured. The chemiluminescence method was used to detect the activity of Casapse-3/7. Results: The inhibitory effect of RCEE on proliferation of A549 cells was strong and dose-dependent. IC50 value was 48.14 mg · L-1 at 8 h. RCEE could induce A549 cell apoptosis in a dose-dependent manner. After RCEE treatment, A549 cells The level of intracellular free calcium increased and the mitochondrial membrane potential decreased. The activity of Caspase-3/7 increased. The intracellular Ca2 + chelator BAPTA-AM and Z-VAD-FMK could inhibit the proliferation of A549 cells induced by RCEE Apoptosis. CONCLUSION: RCEE can inhibit the proliferation of A549 cells and induce the apoptosis of A549 cells. The mechanism may be related to the overload of calcium, the decrease of mitochondrial membrane potential and the activation of Caspase cascade.