冬凌草甲素诱导结肠癌细胞凋亡的机制研究

Study on the mechanism for oridonin-induced apoptosis of human colon cancer cells

目的:观察冬凌草甲素(oridonin)对人结肠癌细胞HCT-116和HT-29的凋亡抑制作用,同时探讨其诱导结肠癌细胞凋亡作用的机制。方法:应用Cell-counting kit-8检测冬凌草甲素对HCT-116和HT-29细胞的生长抑制作用及对细胞活力的影响;采用烟酸己可碱(hoechst)33342和碘化丙啶(propidium lodine,PI)荧光染色法,观察冬凌草甲素诱导结肠癌HCT-116和HT-29细胞凋亡的形态学变化;采用总NO检测试剂盒与活性氧(reactive oxygen species,ROS)检测试剂盒检测冬凌草甲素处理HCT-116和HT-29细胞时,细胞内NO与ROS水平的变化;同时应用ROS阻断剂N-乙酰基半胱氨酸(N-acetyl-cysteine,NAC)预处理HCT-116、HT-29细胞后再加入冬凌草甲素,观察细胞凋亡情况。结果:冬凌草甲素可明显抑制HCT-116和HT-29细胞的增殖、降低细胞活力;Hoechst 33342和PI双荧光染色发现冬凌草甲素可诱导HCT-116和HT-29细胞的凋亡。虽然冬凌草甲素不能改变HCT-116、HT-29细胞内的NO水平,但是可提高HCT-11和HT-29细胞内ROS水平;ROS阻断剂NAC可阻断冬凌草甲素诱导HCT-116和HT-29细胞凋亡。结论:冬凌草甲素通过提高ROS水平诱导HCT-116和HT-29细胞的凋亡。

Objective：To study the inhibitory effect of oridonin on human colon cancer HCT-116 and HT-29 cells and investigate the molecular mechanism for apoptosis-inducing effect of oridonin.Methods：Cell-counting kit-8 was used to detect the inhibitory effects of oridonin on proliferation and viability of colon cancer cells.Hoechst 33342 and propidium iodide（PI） fluorescent staining was used to investigate the apoptotic morphology of HCT-116 and HT-29 cells.The intracellular levels of nitric oxide（NO） and reactive oxygen species（ROS） after oridonin treatment were detected by using NO detection kit and ROS detection kit,respectively.The colon cancer HCT-116 and HT-29 cells were pretreated with N-acetyl cysteine（NAC） and then treated with oridonin to observe the apoptotic changes.Results：Oridonin significantly inhibited the growth and viability of HCT-116 and HT-29 cells.Hoechst 33342 and PI double fluorescent staining showed that the oridonin induced apoptosis of HCT-116 and HT-29 cells.Although oridonin could not change the intracellular NO level in HCT-116 and HT-29 cells,but it increased the intracellular ROS level.The ROS blocker,NAC,reversed the effects of oridonin on apoptosis of HCT-116 and HT-29 cells.Conclusion：Oridonin induces apoptosis of HCT-116 and HT-29 cells by elevating ROS level.