活性氧在荆花牡荆素诱导人肺癌A549细胞凋亡中的作用

The role of reactive oxygen species in casticin-induced apoptosis of human lung cancer A549 cells

目的探讨紫花牡荆素(CAS)诱导人肺腺癌A549细胞凋亡及其机制。方法体外培养A549细胞。MTT法测定CAS对A549细胞增殖的抑制;Annexin V/PI双染色分析细胞凋亡率;H2DCFH-DA探针流式细胞术分析活性氧(ROS)生成。结果 CAS能抑制人肺癌A549细胞增殖,呈浓度依赖性。Annexin V/PI法检测结果显示10μmol/L的CAS作用A549细胞12h、24h、48h后,其凋亡率分别为22.39%、38.66%、64.82%。H2DCFH-DA探针流式细胞术分析表明,完全培养基组、溶媒(0.1%DMSO)组对A459细胞作用0h;CAS(10μmol/L)对A549细胞作用3h、6h、12h;N-乙酰-L-半胱氨酸(NAC,10mmol/L)+CAS(10μmol/L)组对A549细胞作用6h的ROS生成水平分别为8.47、15.26、66.2、74.1、82.2、67.3,随着CAS作用时间延长,细胞内ROS水平增加。NAC对细胞凋亡有抑制作用。结论 CAS可诱导A549细胞凋亡,其作用机制可能与提高细胞内ROS产生增加有关。

Objective To investigate the apoptosis induced by casticin（CAS）on human lung cancer A549 cells and the mechanism involved.Methods Human lung cancer A549 cells line was cultured in vitro.The inhibitory effect of CAS on the proliferation of human lung cancer A549 cells was measured by MTT assay.CAS-induced apoptosis rates of A549 cells were observed by flow cytometry with annexin V/PI fluorescence staining.The flow cytometry with the probe of H2DCFH-DA was used to detect the intracellular reactive oxygen species（ROS）level.Results The MTT assay showed that CAS had a significantly inhibiting effect on the cell proliferation in A549 cells in a concentration-dependent manner.The flow cytometry with annexin V/PI fluorescence staining indicated that the apoptosis rates of A549 cells treated with 10 μmol/L CAS for 12 h,24 h,48 h were 22.39%,38.66%,64.82% respectively.The flow cytometry with the probe of H2DCFH-DA indicated that CAS could increase in the intracellular ROS level.However,after treatment with N-acetyl-L-cystein（NAC）,the apoptotic rate of the cells and the intracellular ROS level also decreased.Conclusions CAS possesses a significant function for inducing apoptosis of A549 cells,which seems to be due to associated with intracellular generation of ROS.