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目的:探讨雷公藤红素(celestrol)诱导KRAS驱动的结肠癌SW620细胞产生凋亡的作用和机制。方法:四甲基偶氮唑蓝(MTT)法和台盼蓝拒染法检测细胞增殖;免疫印迹法检测蛋白表达;流式细胞仪和荧光显微镜检测细胞凋亡、细胞周期、线粒体膜电位;荧光显微镜检测细胞内活性氧水平(reactive oxygen species,ROS)。结果:雷公藤红素明显抑制SW620细胞的增殖活性;雷公藤红素下调SW620胞内的p-Akt、NF-κB、Survivin表达,激活caspase-7、caspase-3和PARP;雷公藤红素增加SW620细胞内的ROS、降低线粒体膜电位、阻滞细胞周期于G2/M期和诱导凋亡。抗氧化剂N-乙酰半胱氨酸(NAC)抑制雷公藤红素引起的上述作用。结论:通过诱导细胞内ROS的累积导致细胞内线粒体膜电位的下降进而触发细胞发生凋亡是雷公藤红素诱导SW620细胞凋亡的作用机制之一。
Objective: To investigate the effect and mechanism of celestrol on apoptosis induced by KRAS in colon cancer SW620 cells. Methods: Cell proliferation was detected by MTT assay and trypan blue exclusion assay. Protein expression was detected by Western blotting. Apoptosis, cell cycle and mitochondrial membrane potential were detected by flow cytometry and fluorescence microscope. Fluorescence microscopy was used to detect intracellular reactive oxygen species (ROS). Results: Tripterine significantly inhibited the proliferation of SW620 cells. Tripterine down-regulated the expression of p-Akt, NF-κB and Survivin in SW620 cells and activated caspase-7, caspase-3 and PARP ROS in SW620 cells decreased mitochondrial membrane potential, arrested cell cycle in G2 / M phase and induced apoptosis. Antioxidant N-acetylcysteine (NAC) inhibits tripterine-induced effects. CONCLUSION: Triptolide induces apoptosis in SW620 cells by inducing the accumulation of intracellular ROS, resulting in the decrease of mitochondrial membrane potential and triggering cell apoptosis.