卤地菊乙醇提取物W40单体诱导GLC-82细胞凋亡的分子机制研究  

Molecular Mechanism of Inducing GLC-82 Cells Apoptosis by Ethanol Extract from Wedelia prostrate(Hook. et Arn.) Hemsl

作　　者：代立婷 吴忠南 黄翔[1] 杨杰[1] 曾慧兰[1] 王国才[2] 蒋建伟[1] 

机构地区：[1]暨南大学医学院,广州510632 [2]暨南大学药学院中药及天然产物研究所,广州510632

出　　处：《中国生物工程杂志》2017年第8期1-7,共7页China Biotechnology

基　　金：广东省自然科学基金(2014A030313356);广东省医学科研基金(2015130164321691)资助项目

摘　　要：目的:探讨卤地菊Wedelia prostrate(Hook.et Arn.)Hemsl.的乙醇提取物W40单体对非小细胞肺癌GLC-82细胞的抗肿瘤作用分子机制。方法:通过MTT和克隆形成抑制实验检测W40对非小细胞肺癌GLC-82细胞增殖的影响;通过细胞划痕实验检测W40对细胞迁移的影响;通过Annexin V-FITC/PI双染法检测W40对细胞凋亡的诱导;通过Western blotting分析细胞增殖和凋亡相关蛋白的水平。结果:MTT实验表明,W40对GLC-82细胞有较为明显的细胞毒作用;克隆形成抑制实验表明,W40可以显著抑制GLC-82细胞增殖,且抑制程度呈现浓度依赖效应;细胞划痕实验表明,W40能够在一定程度上抑制GLC-82细胞的迁移能力;Annexin V-FITC/PI双染的结果显示,随着药物浓度的增加,细胞凋亡率逐渐增加;Western blotting结果表明,随着药物浓度的增加,p-Stat3蛋白的表达水平下降,Stat3下游蛋白Bcl-2、Mcl-1的表达减少。同时,促凋亡蛋白Bax的表达增多,并伴随Caspase-9、Caspase-3活化及多聚ADP-核糖聚合酶PARP酶切失活;p-BRAF蛋白的表达水平下降,p-BRAF下游蛋白p-MEK、p-ERK表达减少。结论:W40通过抑制BRAF/MAPK/ERK及Stat3信号通路来诱导细胞凋亡。Objective： To study the antitumor mechanism of W40,a monomer purified from Wedelia prostrate（ Hook. et Arn.） Hemsl. Methods： The effects of W40 on the cell proliferative of GLC-82 cells were detected by MTT assay and colony formation assay. The migratory abilities of GLC-82 cells were observed by wound healing assay. Cell apoptosis was evaluated by Annexin V-FITC/PI staining analysis. The levels of apoptosis-relative proteins and cell proliferation-related proteins,such as Caspase-3,PARP,Stat3 and ERK,were detected by Western blotting. Results： MTT assay showed that W40 had a significant cytotoxic effect on non-small cell lung cancer GLC-82 cells. Colony formation assays showed that W40 significantly inhibited GLC-82 cells proliferation.The migration of GLC-82 cells was inhibited by W40 in a dose-dependent manner. Flow cytometry showed that the apoptotic rate increased gradually in a concentration-dependent manner. W40 down-regulated Stat3 as decreasing p-Stat3 and downstream proteins of Bcl-2 and Mcl-1. At the same time,W40 up-regulated the expression of pro-apoptotic protein Bax,and increased the cleavaged Caspase-9,Caspase-3 and PARP. W40 also down-regulated BRAF/MAPK/ERK signal pathway as decreasing p-BRAF,p-MEK and p-ERK. Conclusions：W40 induced apoptosis by inhibiting BRAF/MAPK/ERK and Stat3 signaling pathways.

关 键 词：卤地菊 肺癌 细胞凋亡 STAT3 BRAF/MAPK/ERK信号通路 

分 类 号：Q819[生物学—生物工程]