冠心病血瘀证血小板差异功能蛋白筛选、鉴定及功能分析

Screening,Identification and Analysis of Platelet Differential Functional Proteins in Patients with Coronary Heart Disease of Blood-stasis Pattern

目的寻找冠心病血瘀证患者冠脉事件关键血小板功能蛋白。方法纳入冠心病血瘀证组患者,设立正常对照组,每组分4个小组,即两组各有4次重复。抽取外周血,分离血小板并提取蛋白,将各个提取血小板蛋白样本按小组等体积混合、定量。用荧光差异显示二维凝胶电泳技术筛选、基质辅助激光解析/电离-飞行时间质谱技术鉴定血小板差异功能蛋白。对鉴定成功差异蛋白,用Western-blotting进一步验证。结果筛选出13个差异蛋白点,质谱成功鉴定7个:isoform 1 of integrinalpha-Ⅱb、isoform 2 of integrinalpha-Ⅱb、actin-cytoplasmic1、actin-cytoplasmic 2、cDNAFLJ52842、cDNAFLJ55253、cDNAFLJ43573fis。其中isoform2ofintegrinalpha-Ⅱb(CD41)和actin-cytoplasmic2(Actinγ)成功验证。结论 CD41和Actinγ是冠心病血瘀证的标志蛋白。其他血小板功能蛋白的异常表达可能在冠心病血瘀证事件发生发展中起关键作用。

Objective To seek the key platelet functional proteins (PFPs) for the occurrence of blood-stasis pattern (BP) in patients with coronary heart disease (CHD). Methods Peripheral blood platelet protein of 22 patients and 24 healthy person (for control) were extracted respectively in 4 batches for carrying 4 times of the test out. Differential PFPs in samples were screened out by two-dimensional fluorescence difference gel electrophoresis, and identified with matrix-assisted laser desorption/ionizating-time of flight mass spectrometry; then the identified proteins were further authenticated by Western-blotting. Results Thirteen differential PFPs were screened out, and among them the 7 identified by spectrometry were: isoform 1 of integrin alpha-Ⅱb, isoform 2 of integrin alpha-Ⅱb, actin-cytoplasmic 1, actin-cytoplasmic 2, cDNA FLJ52842, cDNA FLJ55253, and cDNA FLJ43573 fis. Among them isoform 2 of integrin alpha-Ⅱb (CD41) and actin-cytoplasmic 2 (Actinγ) were authenticated successfully. Conclusion CD41 and actinγ are the possible marker proteins, and the other PFPs might play crucial roles in the occurrence and development of BSS in CHD.