目的:探讨榄香烯乳(ElEMEnE,ElE)逆转人肺腺癌耐顺铂(CISPlATIn,ddP)细胞A549/ddP的耐药性及作用机制。方法:采用MTT法检测榄香烯乳单用的细胞毒作用及与ddP合用时耐药逆转作用。荧光探针JC-1结合激光共聚焦显微镜检测线粒体膜电位的变化。dCfH-dA荧光探针结合流式细胞仪检测细胞内活性氧(rEACTIVE OXygEn SPECIES,rOS)水平。用谷胱甘肽试剂盒结合分光光度法检测计算gSH/(gSSg+gSH)比值。蛋白质印迹法检测胞质中CyTO C、PrO-CASPASE-3、CASPASE-3和bCl-2家族蛋白表达情况。结果:不同浓度榄香烯乳抑制A549/ddP细胞株生长,呈时间-剂量依赖性效应,联合顺铂能提高A549/ddP细胞株对顺铂的敏感性而逆转耐药。不同浓度榄香烯乳联合顺铂使A549/ddP细胞株线粒体膜电位下降,rOS浓度增加,gSH/(gSSg+gSH)比值降低,上调胞质中CyTO C、CASPASE-3、bAd蛋白表达,下调PrO-CASPASE-3、bCl-2蛋白表达。结论:榄香烯乳逆转A549/ddP细胞株耐药性可能与其损伤线粒体膜,活化胞内氧化还原体系,诱导线粒体凋亡路径有关。Objective: To explore the mechanism that elemene( ELE) reversed the multidurg resistance( MDR)of A549 /DDP lung adenocarcinoma cell.Methods: MTT assay was used to determine the growth inhibition of human lung adenocarcinoma A549 /DDP cells in vitro.Mitochondrial membrane potential( MMP) was monitored by JC- 1fluorescence probe with laser confocal scanning microscopy,the intercellular reactive oxygen species( ROS) level was measured by 2',7'- dichlorfluorescein- diacetate( DCFH- DA) staining and flow cytometry and the ratio of GSH /( GSSG +GSH) was calculated according to detection results of GSH kit.The expression of Cytochrome C,Caspase-3 and the Bcl- 2 family proteins and in the case of cyclosporine A and DEVD- CHO,the expression of Caspase- 3expression were measured by Western blot.Results: MTT results showed that different concentrations ELE could inhibit the proliferation of A549 /DDP cells in a time- and dose- dependent manner.Intriguingly,ELE plus cisplatin enhanced the sensitivity of A549 /DDP cells to cisplatin and reversed A549 /DDP cells dury resistance.Different concentrations ELE decreased mitochondrial membrane potential,increased intracellular ROS concentration and decreased GSH /( GSSG + GSH) ratio of A549 /DDP cells in a time- and dose- dependent manner.Furthermore,the combination with both ELE and cisplatin also enhanced the protein expression of Cytoplasmic C,Caspase- 3 and Bad,and reduced the protein levels of Bcl- 2 and Pro- caspase- 3 in the cisplatin- resistant A549 /DDP cancer cells.Conclusion: ELE reversed the MDR of A549 /DDP cell line may demage mitochondrial membrane,active intracellular redox system and induce the mitochondrial apopotosis pathway.福建省自然科学基金面上项目(编号:2010D014
