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目的探讨茶多酚特异性诱导胰腺癌细胞死亡的线粒体机制。方法电子显微镜下观察茶多酚处理后胰腺癌细胞的形态学变化。三磷酸腺苷生物发光法检测茶多酚作用下胰腺癌细胞ATP 含量的变化,MitoCapture 法检测茶多酚作用下胰腺癌细胞的线粒体膜电位的变化。探讨细胞ATP 含量与胰腺癌细胞凋亡与坏死之间的关系。Western blot 技术检测茶多酚作用后胰腺癌细胞胞液中细胞色素 C,caspase 蛋白质含量的变化。结果随着茶多酚浓度增高和不同浓度下作用时间延长,电镜下胰腺癌细胞线粒体肿胀,部分细胞呈坏死样改变,细胞 ATP 含量降低,MitoCapture 阳性细胞百分数增加。ATP 含量与 BxPC-3细胞细胞 MitoCapture 阳性细胞呈负相关(r=0.808,P<0.01)。细胞色素 C 和 procaspase-9释放到胞液中,胞液中可检测出活化的 caspase-9,和 caspase-3。结论茶多酚具有特异性诱导胰腺癌细胞死亡的功能,机制与细胞 ATP 含量降低,线粒体膜电位破坏,通透性增高,细胞色素 C 和 procaspase-9释放,活化 caspase-3启动胰腺癌细胞细胞凋亡过程,当ATP 含量极度降低时部分胰腺癌细胞细胞出现坏死。
Objective To investigate the mitochondrial mechanism of specific induction of pancreatic cancer cell death by tea polyphenols. Methods The morphological changes of pancreatic cancer cells treated with tea polyphenols were observed under electron microscope. The ATP content of pancreatic cancer cells was detected by adenosine triphosphate bioluminescence assay. The changes of mitochondrial membrane potential of pancreatic cancer cells were detected by MitoCapture assay. To explore the relationship between cellular ATP content and apoptosis and necrosis of pancreatic cancer cells. Western blot technique was used to detect the changes of cytochrome C and caspase protein contents in the cytosol of pancreatic cancer cells after tea polyphenols treatment. Results With the increase of the concentration of tea polyphenols and the prolongation of the action time under different concentrations, the mitochondria of pancreatic cancer cells swelled under electron microscope, some of the cells showed necrosis-like changes, the cellular ATP content decreased, and the percentage of MitoCapture positive cells increased. The ATP content was negatively correlated with MitoCapture positive cells in BxPC-3 cells (r=0.808, P<0.01). Cytochrome C and procaspase-9 are released into the cytosol, and caspase-9 and caspase-3 are detected in the cytosol. Conclusion Tea polyphenols have the function of specifically inducing the death of pancreatic cancer cells. The mechanism is related to the decrease of cellular ATP content, destruction of mitochondrial membrane potential, increased permeability, release of cytochrome C and procaspase-9, and activation of caspase-3 to activate pancreatic cancer cells. During apoptosis, some pancreatic cancer cells appear necrotic when the ATP content is extremely reduced.