小型猪心肌缺血血瘀证的血浆蛋白质组学研究

目的：寻找血瘀证（心肌缺血）相关的血浆蛋白质组学特征。方法：采用前期建立的方法,以小型猪前降支主干放置Ameroid缩窄环结合动态观察的方法制备小型猪血瘀证模型。在证候时间窗内前腔静脉采血,枸橼酸钠抗凝,离心收集上清备用。血浆总蛋白上样量1.3mg,同组样品等量混合后,采用双向凝胶电泳分析差异蛋白点。等电聚焦用IPG胶条24cm、PH范围3-11NL,SDS-PAGE浓度13%,考马斯亮蓝R350染色,凝胶扫描后,采用Imagemaster 2D Platinum 6.0软件进行凝胶的蛋白点检测、匹配。对匹配蛋白点的体积百分比进行比较,确定差异蛋白点。差异蛋白点胰蛋白酶酶切后,MALDI-TOF检测,数据库搜索,确定其相关信息。结果：模型组有10个明显的上调蛋白点。质谱鉴定结果提示,部分上调的差异蛋白为α1-酸性糖蛋白、α1-抗胰蛋白酶、胎球蛋白A、载脂蛋白A-I、载脂蛋白A-IV。结论：已鉴定的差异蛋白质主要为急性期反应蛋白及载脂蛋白等,炎症/抗炎是其互相联系的核心环节,提示在该心肌缺血血瘀证同时存在着炎症反应这一病理进程,炎症反应可能是血瘀证的病理机制之一,但仍有待进一步研究。

Research on the Plasma Proteomic Characteristics of Blood Stasis Syndrome with Myocardial Ischemia by Using an Integrated Animal Model of Mini Swine

To discover the plasma proteomic characteristics of blood stasis syndrome with myocardial ischemia.Methods：An integrated animal model of mini swine blood stasis syndrome with myocardial ischemia was established as we reported before and sham operated group was used as control.Plasma samples obtained were quantitated by Bradford assay,and then pooled samples were separated by two-dimensional electrophoresis（2-DE） to identify differentially expressed proteins.Precast IPG strips（pH3-11 nonlinear,24cm） were used in the first dimension,and SDS-polyacrylamide gel electrophoresis（PAGE） was performed using 1 mm thick,13% SDS-PAGE gels.The Coomassie Brilliant Blue R350-stained 2-DE were scanned with digital scanner and analyzed with ImageMaster 2D Platinum 6.0 software.A cluster of protein spots differentially expressed were selected and identified with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry（MALDI-TOF-MS）.Results：10 up-regulated protein spots were detected in the 2-DE separation of pooled samples.MS results suggest that the upregulated proteins including α1-acid glycoprotein,α1-antitrypsin,fetuin A,apolipoprotein A-IV,apolipoprotein A-I.Other changes were still unknown.Conclusions：The detected differential expressed plasma proteins of animals with blood stasis syndrome were associated with inflammatory reactions.Thus,we supposed that inflammation may be one of the important pathologic processes in blood stasis syndrome.However,their exact relationship was still unknown.