百草枯通过激活线粒体凋亡通路诱导人肺Ⅱ型上皮样A549细胞凋亡

发布时间：2018-04-14 01:31

本文选题：百草枯 + 肺泡上皮细胞　；参考：《中国医科大学学报》2017年11期

【摘要】：目的探讨百草枯(PQ)诱导人肺Ⅱ型上皮样A549细胞凋亡过程中线粒体凋亡通路的发生机制。方法体外培养A549细胞,对照组加入RPMI 1640培养液,实验组加入不同浓度的PQ(50、100、150和200μmol/L),2组细胞继续培养24和48 h。MTT法检测细胞活性;Hoechst 33258染色法观察细胞核的形态;流式细胞术检测细胞凋亡率以及线粒体膜电位;分光光度法检测caspase-3和caspase-9的活化程度;Western blotting检测线粒体凋亡相关蛋白Bcl-2、Bcl-x L、Bax和Bak的表达。结果 MTT结果显示,PQ对A549细胞具有显著的生长抑制作用。Hoechst染色显示,不同浓度的PQ作用于A549细胞24和48 h后,可发现细胞凋亡的特征性改变,如细胞核固缩、核碎裂、凋亡小体形成,并且细胞的凋亡程度随着PQ浓度的增大和作用时间的延长而加重。细胞凋亡率升高也证实了这一结果。线粒体膜电位出现不同程度的降低;caspase-3和caspase-9活性不同程度的增加;抗凋亡蛋白Bcl-2、Bcl-x L的表达量降低;促凋亡蛋白Bax、Bak的表达量升高。以上结果均呈时间与浓度依赖性。结论 PQ通过激活线粒体凋亡通路诱导A549细胞凋亡。
[Abstract]:Objective to investigate the pathogenesis of paraquat (PQ) mitochondrial apoptosis pathway induced apoptosis of human pulmonary epithelioid A549 cells. Methods A549 cells were cultured in vitro, the control group added RPMI 1640 medium, different concentration of experimental group treated with PQ (50100150 and 200 mol/L), 2 groups of cells to detect cell activity and 24 48 h.MTT culture method; nuclear morphology was observed by Hoechst 33258 staining; detection of apoptosis rate and mitochondrial membrane potential flow cytometry; the activation level of Caspase-3 and caspase-9 detected by spectrophotometry; mitochondrial apoptosis detection Western blotting related protein Bcl-2, Bcl-x L, the expression of Bax and Bak. The results of MTT showed that PQ has significant growth inhibition of.Hoechst staining of A549 cells, different concentrations of PQ in A549 cells 24 and 48 h, reveals the characteristic changes of cell apoptosis, such as nuclear pyknosis, nuclear fragmentation The degree of apoptosis, apoptotic body formation, and the cells were aggravated with the increase of PQ concentration and reaction time. The apoptosis rate increased also confirmed this result. The mitochondrial membrane potential decreased differently; increased caspase-3 and caspase-9 activity in different degree; the anti apoptosis protein Bcl-2, expression of Bcl-x L decreased; the pro apoptotic protein Bax, the expression of Bak was increased. The above results were in a time and concentration dependent manner. Conclusion PQ can induce apoptosis in A549 cell activation of the mitochondrial apoptosis pathway.

【作者单位】：中国医科大学附属第一医院急诊科;
【基金】：国家自然科学基金(81471851) 辽宁省博士启动基金(20141033)
【分类号】：R595.4

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