藤黄酸通过MAPK通路诱导Jurkat细胞凋亡的机理

摘要

The aim of this study was to investigate the apoptosis-inducing effect of gambogic acid (GA) on Jurkat cells and its underlying signaling pathway. Apoptosis induced by GA and some inhibitors was assayed by Annexin V/PI doubling staining. The levels of caspase 3, caspase 8 and caspase 9 activated in living Jurkat cells were measured by flow cytometry. The expressions of caspase 3, caspase 9, β-JNK and P38 were detected by Western blot. The results showed that GA induced apoptosis of Jurkat cells in a dose-dependent manner. The positive cell number of activated caspase 3, caspase 8, caspase 9 and the levels of activated caspase 3 .caspase 9,p-JNK, P38 increased after Jurkat cells were treated with GA. ROS, CaMK II , caspase 3, caspase 9, MAPKK, JNK1/2 and P38 inhibitors had some significant effect on GA-induced apoptosis. ROS, CaMK H , MAPKK, JNK1/2 and P38 inhibitors decreased the levels of activated caspase 3, caspase 9 by GA. ROS, CaMK II , MAPKK, JNK1/2 inhibitors decreased the levels of β-JNK by GA. ROS, CaMK II, MAPKK, P38 inhibitors decreased the levels of P38 by GA. It is concluded that GA induced apoptosis of Jurkat cells by activated caspases through activating of ROS-CaMKII - MAPKK-JNK/P38 pathway.%本研究探讨藤黄酸对Jurkat细胞的抑制作用及可能机理.以不同浓度藤黄酸与抑制剂处理Jurkat细胞,Annexin V/PI双染法检测细胞的凋亡；流式细胞仪检测荧光素激活的caspase3、caspase 8、caspase 9阳性细胞水平；免疫印迹检测pro-caspase 3、pro-caspase 9、磷酸化JNK及P38蛋白水平.结果表明,藤黄酸呈浓度依赖性诱导Jurkat细胞凋亡；藤黄酸作用Jurkat细胞后caspase3、caspase8、caspase9阳性细胞水平升高,caspase3、caspase9活化蛋白及磷酸化JNK及P38蛋白水平升高；ROS、CaMKⅡ、caspase 3、caspase 9、MAPKK、JNK1/2及P38抑制剂减弱藤黄酸诱导Jurkat细胞凋亡的作用；ROS、CaMKⅡ、MAPKK、JNK1/2及P38抑制剂降低藤黄酸作用Jurkat细胞后caspase 3、cmaspase9活化蛋白水平；ROS、CaMKⅡ、MAPKK、JNK1/2抑制剂降低藤黄酸作用Jurkat细胞后磷酸化JNK蛋白水平；ROS、CaMKⅡ、MAPKK、P38抑制剂减弱藤黄酸作用Jurkat细胞后磷酸化P38蛋白水平.结论:藤黄酸通过激活caspase途径诱导Jurkat细胞凋亡,该过程可能与ROS- CaMKⅡ-MAPKK-JNK/P38通路有关.