小白菊内酯对人肺癌A-549细胞株的抗增殖及诱导凋亡作用

Effects of parthenolide on inhibiting proliferation and inducing apoptosis of human lung cancer A-549 cell line

[目的]探讨小白菊内酯(Par)对人肺癌A-549细胞株的抗增殖及诱导凋亡作用及其机制.[方法]应用倒置显微镜、HE染色光学显微镜、吖啶橙(AO)/溴化乙锭(EB)双染色荧光显微镜观察Par作用前后人肺癌A-549细胞的形态学变化;应用流式细胞术(FCM)检测Par作用之后肿瘤细胞周期分布及诱导凋亡的情况;应用免疫细胞化学技术检测Par作用前后蛋白阳性表达变化情况,并观察Par对人肺癌A-549细胞株抑制增殖及诱导凋亡的作用机制.[结果]倒置显微镜观察结果显示,实验组细胞体积缩小、变圆,核染色质聚集成团,细胞之间的连接疏松,贴壁不牢,易脱落.HE染色光学显微镜观察结果显示,实验组细胞体积变小,数目减少,可见部分细胞核固缩碎裂,部分细胞凋亡.AO/EB双染色荧光显微镜观察结果显示,对照组细胞呈绿色荧光,而实验组中早期凋亡细胞呈黄绿色荧光,随着Par作用时间延长,可见呈橘红色荧光的晚期凋亡细胞及坏死细胞.流式细胞术检测结果显示,G0/G1期细胞数量减少,而S期和G2/M期细胞数量明显增多(P<0.05).免疫细胞化学技术检测结果显示,实验组细胞Bcl-2,Ki-67,Livin,Survivin及核转录因子kappa B(NF-κB)表达明显降低(P<0.05).[结论]Par对人肺癌A-549细胞有明显的抑制增殖作用,其机制可能与细胞周期发生S期和G2/M期阻滞有关.Par作用于人肺癌A-549细胞株后,可诱导细胞凋亡,其机制是通过死亡受体通路及线粒体通路来完成的,NF-κB可能参与Par诱导人肺癌A-549细胞凋亡的作用.

OBJECTIVE To study the effects and its mechanism of parthenolide（Par）on inhibiting proliferation and inducing apoptosis of human lung cancer A-549 cell line in vitro.METHODS Morphological changes of human lung cancer A-549 cells before and after treated with the Par were observed by inverted microscope,light microscope with HE staining,fluorescent microscope with AO/EB double staining,and the tumor cell cycle distribution and inducing apoptosis after treated with the par were observed by flow cytometry,and changes of protein expression before and after treated with the par were detected by immunocytochemistry to further explore the mechanism of par on inhibiting proliferation and inducing apoptosis of human lung cancer A-549 cell line.RESULTSIn experiment group,it was showed by inverted microscope that the A-549 cells were volume shrinkage,turning round,nuclear chromatin gathered together,loose connections between cells and easy to fall off from wall,and by light microscope with HE staining that the A-549 cells were smaller,decrease of cell number,part of karyopycnosis and karyorrhexis and part of the cell apoptosis,and by fluorescent microscope with AO/EB double staining that the early cell apoptosis with yellow-green fluorescence,late cell apoptosis and necrotic cells with orange fluorescent along with treating time extension with Par as with green fluorescence in control group.It was showed by flow cytometry that percentage of cells in G0/G1 phase decreased and percentage of cells in S G2/M phase increased after treated with Par（P〈0.05）.It was showed by immunocytochemistry that the expression of Bcl-2,Ki-67,Livin,surviving and NF-κB in experimental groups（P〈0.05）.CONCLUSION The Par has obviously inhibitive effects for proliferation of A-549 cells in vitro,and its mechanism maybe relate to make the cell cycle of A-549 cells arrest in the S G2/M phase.The Par induces apoptosis of A-549 cells,and its mechanism is completed by the death receptor pathways and mitochondrial pathways,and NF-κB may participate in the process of inducing apoptosis of A-549 cells.