石榴酸诱导子宫内膜样腺癌细胞凋亡的作用机理

Mechanisms of apoptosis induction of endometrioid adenocarcinoma cells by punicic acid

研究了石榴酸诱导子宫内膜样腺癌RL-952细胞凋亡的作用机理。以α-亚麻酸和α-桐酸为对照,通过MTT法检测石榴酸对细胞相对活力的影响,并通过荧光显微镜观察细胞核的形态学变化,应用流式细胞术分析细胞的凋亡率。应用蛋白免疫印迹检测凋亡相关蛋白的表达,分析石榴酸对PPARγ和p53蛋白表达的影响。应用荧光染料DCFH-DA染色,通过荧光分光光度法分析了细胞内ROS水平的变化。应用PPARγ抑制剂T0070907和ROS清除剂NAC对石榴酸的作用机理进行了初步探讨。结果表明,与α-亚麻酸和α-桐酸相比,石榴酸抑制RL-952细胞相对细胞活力的作用最强;石榴酸处理后细胞出现明显的凋亡形态学改变,凋亡细胞的比例明显增加。石榴酸能够明显促进凋亡相关蛋白酶Caspase-3的表达,并上调凋亡相关蛋白Bax/Bcl-2的表达比例,且明显增加PPARγ和p53蛋白的表达水平。石榴酸还能够显著增加RL-952细胞内ROS的产生。石榴酸抑制RL-952细胞活力的作用可以被T0070907和NAC所拮抗,结果提示,石榴酸对子宫内膜样腺癌RL-952细胞的凋亡诱导作用可能与PPARγ信号通路的活化及ROS的产生有关。

The mechanisms of apoptosis of endometrioid adenocarcinoma RL-952 cells induced by punicic acid were studied.Compared withα-linolenic acid andα-eleostearic acid,the effects of punicic acid on the cell relative activity were detected by MTT method.The morphological changes of nuclei were observed by fluorescence microscope,and the apoptosis rate of cells was analyzed by flow cytometry.The expression of apoptosis related proteins was detected by Western Blot,and the effect of punicic acid on the expression of PPARγand p53 were analyzed.The change of ROS level in cells was analyzed by fluorescence spectrophotometry with fluorescent dye DCFH-DA.The mechanism of PPARγinhibitor T0070907 and ROS scavenger NAC on punicic acid was evaluated.The results showed that compared withα-linolenic acid andα-eleostearic acid,punicic acid had the most effective inhibitory effect on the relative cell viability of RL-952 cells.After treated by punicic acid,cells showed obvious morphological changes of apoptosis and the proportion of apoptotic cells increased significantly.The punicic acid could significantly promote the expression of Caspase-3,up-regulate the expression ratio of Bax/Bcl-2,and significantly increase the expression level of PPARγand p53.The punicic acid could also significantly increase ROS production in RL-952 cells.The inhibitory effect of pomegranate acid on the activity of RL-952 cells can be antagonized by T0070907 and NAC.The results suggested that the apoptosis inducing effect of punicic acid on RL-952 cells may be related to the activation of PPARγsignal pathway and the production of ROS.