马蔺子素以IAPs为靶点诱导人非小细胞肺腺癌NCI-H1975细胞凋亡的机制研究

Study on the Mechanism of Irisquinone in Inducing Apoptosis of Human Non-Small Cell Lung Adenocarcinoma NCI-H1975 by Targeting IAPs

目的:研究马蔺子素对人非小细胞肺腺癌NCI-H1975细胞的体外抗肿瘤作用及其以IAPs为靶点诱导凋亡的机制。方法:培养NCI-H975细胞,在显微镜下观察马蔺子素对NCI-H1975细胞形态学的影响;采用MTT法检测不同浓度马蔺子素对NCI-H1975细胞增殖的影响;流式细胞术检测马蔺子素对NCI-H1975细胞凋亡的影响;Western Blot法检测Smac蛋白和凋亡相关蛋白Caspase-3、Caspase-9、PARP、Bcl-2、Bax及IAPs家族相关蛋白CIAP1、CIAP2、XIAP、Survivin、Livin的表达。结果:细胞形态学显示不同浓度马蔺子素作用不同时间下细胞增殖受到不同程度抑制,细胞数量减少;MTT实验表明马蔺子素能呈浓度和时间依赖性地抑制NCI-H1975细胞的增殖(P<0.05);流式细胞术结果表明马蔺子素能促进NCI-H1975细胞凋亡(P<0.05);Western Blot结果显示随着马蔺子素浓度的升高,凋亡相关蛋白PARP被切割、Caspase-3/9原蛋白和抗凋亡蛋白Bcl-2的表达下调同时促凋亡蛋白Bax表达上调,Smac蛋白表达上调,IAPs家族相关蛋白CIAP1、CIAP2、XIAP、Survivin、Livin的表达下调(P<0.05)。结论:马蔺子素能抑制NCI-H1975细胞增殖并诱导其凋亡,其机制可能与抑制IAPs家族蛋白有关。

Objective:To study the in vitro antitumor effect of irisquinone on human non-small cell lung adenocarcinoma NCI-H1975 cells and its mechanism of inducing apoptosis by targeting IAPs.Methods:NCI-H975 cells were cultured and the morphological effects of irisquinone on NCI-H1975 cells were observed under microscope.MTT assay was used to detect the effect of different contentrations of irisquinone on proliferation of NCI-H1975 cells.Flow cytometer was used to detect the effect of irisquinone on apoptosis of NCI-H1975 cells.Western Blot was used to detect the expressions of Smac protein and apoptosis related proteins Caspase-3,Caspase-9,PARP,Bcl-2,Bax and the IAPs family related proteins CIAP1,CIAP2,XIAP,Survivin,Livin.Results:The different contentrations irisquinone inhibited the proliferation of NCI-H1975 cells in a time-dependent manner,and the number of cells decreased.MTT assay showed that irisquinone inhibited the proliferation of NCI-H1975 cells in a concentration-dependent and time-dependent manner(P<0.05).Flow cytometry showed that irisquinone could promote apoptosis of NCI-H1975 cells(P<0.05).Western Blot results showed that with the increase of irisquinone concentration,apoptosis-related protein PARP was cleaved,the expressions of Caspase-3/9 proprotein and anti-apoptotic protein Bcl-2 were down-regulated,and the expression of pro-apoptotic protein Bax was up-regulated,the protein expression of Smac was increased,the protein expressions of IAPs family related proteins CIAP1,CIAP2,XIAP,Survivin,Livin were down-regulated(P<0.05).Conclusion:Irisquinone can inhibit the proliferation and induce apoptosis of NCI-H1975 cells,the mechanism may be related to inhibition of IAPs family proteins.