夏枯草提取物对肺腺癌细胞A549蛋白质组的影响

Effects of extracts of Prunella Vulgaris L.on proteome of human lung adenocarcinoma cell line A549

目的 应用双向电泳和质谱技术研究夏枯草提取物对肺癌细胞A549蛋白质组的影响,从蛋白质组学水平探讨夏枯草抗肺腺癌的作用机制.方法 提取A549细胞和经过300μg/ml夏枯草提取物处理的A549细胞总蛋白,并用双向电泳技术进行分离,经银染后扫描得到蛋白质组图谱,用Image Master2D 6.0软件进行差异蛋白质组分析,筛选差异在2倍以上的蛋白质作为差异蛋白进行质谱鉴定,并用Western印迹方法进行初步验证.结果 经夏枯草提取物处理后,A549细胞中1,4,5-三磷酸肌醇受体相互作用样前体蛋白、热休克同源蛋白70、丝氨酸-苏氨酸激酶相关受体蛋白、1型原肌球蛋白2（β）、细胞周期蛋白B3、MED12L蛋白和微丝交联蛋白亚型2表达量增加（与对照组蛋白比值为：2.051 93、1 000 001、2.203 08、5.042 01、15.178 00、1 000001、1 000 001）,而烯醇酶1、M2型丙酮酸激酶、热休克蛋白27、Rho家族GDP分离抑制剂1、热休克蛋白β1、TapasinERP57异源二聚体A链、无机焦磷酸酶、线粒体半胱氨酰-tRNA合成酶2（假定的）表达量降低（与对照组蛋白比值为：0.485 18、0.491 53、0.465 43、0.454 71、0.499 34、0.450 36、0.494 62、0.437 33）.结论 夏枯草提取物具有多靶点、多途径地发挥抗肺腺癌作用,其机制可能与调节Ca2＋平衡、维护细胞周期、抑制肿瘤细胞增殖和转移有关.

Objective To explore the effect oi extracts of Prunella vulgaris L.on proteome of human lung adenocarcinoma cell line A.549 by two-dimensional electrophoresis and mass spectrometry and elucidate the mechanism of anti-lung adenocarcinom effect of Prunella vulgaris L.at the level of proteome.Methods The proliferative activity of human lung adenocarcinoma cell line A549 was evaluated by methyl thiazolyl tetrazolium （MTT） colorimetric assay.According to the difference of culture medium,all subjects were divided into the experimental group with culture medium of extracts of Prunella vulgaris L.（300 μg/ml） and the control group with culture medium of DMSO （0.3％）.Proteins were isolated by two-dimensional electrophoresis and proteomic maps acquired by silver staining.And proteomic analysis was processed by Image Master 2D Quant Platinum 6.0.The proteins with ＞ 2-fold differences were used to analyze by mass spectrometry and confirmed by Western blot.Results The expressions of inositol 1,4,5-triphosphate receptor-interacting protein-like 2 precursor,heat shock cognate protein 70,serine-threonine kinase receptorassociated protein,tropomyosin 2（β） isoform 1,cyclin B3,MED12L protein and macrophin 1 isoform 2 were higher in experimental group than those in control group（ratio （medicial/normal） 2.051 93,1 000 001,2.203 08,5.042 01,15.178 00,1 000 001,1 000 001）.And the expressions of enolase 1,M2-type pyruvate kinase,heat shock protein 27,Rho GDP-dissociation inhibitor 1,heat shock protein β1,TapasinERP57 heterodimer chain A,inorganic pyrophosphatase and mitochondrial Cysteinyl-tRNA synthetase 2 （putative） were lower in experimental group than those in control group（ratio （medicial/normal） 0.485 18,0.491 53,0.465 43,0.454 71,0.499 34,0.450 36,0.494 62,0.437 33）.Conclusions The extracts of Prunellavulgaris L.have multi-target and multi-pathway effects on anti-lung adenocarcinoma.And its possible mechanisms may be due to the regulation of steady state of calcium ion,cell cycle and its steady state and the inhibition of tumor cell proliferation and metastasis.