大定风珠治疗脑出血恢复期阴虚动风证证效关系的蛋白质组学研究

目的 通过鉴定并分析脑出血（intracerebral hemorrhage,ICH）恢复期阴虚动风证患者大定风珠对证、镇肝熄风汤非对证治疗与健康对照组比较外周血单个核细胞（peripheralblood mononuclear cells,PBMCs）差异表达的蛋白质,从方证对应的角度寻找脑出血恢复期阴虚动风证的证效关系蛋白质,并探讨脑出血恢复期阴虚动风证的本质内涵.方法 分离对照组、脑出血恢复期阴虚动风组、脑出血恢复期阴虚动风对证治疗及非对证治疗组外周血单个核细胞,利用双向凝胶电泳技术建立各组的PBMCs 蛋白双向凝胶电泳图谱,经PDQuest8.0 软件分析,选取差异表达的蛋白质点,应用MALDI-TOF-MS 质谱仪获得肽质量指纹图谱,用Mascot 软件搜索数据库鉴定部分差异表达蛋白质.结果 经双向凝胶电泳和图像分析,筛选出的24 个差异表达蛋白质点中共有12 个得到鉴定,其中脑出血恢复期阴虚风动证组与健康对照组比较,5 个蛋白表达上调,4 个蛋白表达下调,1 个蛋白表达缺失;脑出血恢复期阴虚风动证大定风珠对证治疗组与健康对照组比较,7 个蛋白表达无明显差异,2 个蛋白表达下调,2 个蛋白表达上调,1 个蛋白表达缺失;脑出血恢复期阴虚风动证镇肝熄风汤非对证治疗组与健康对照组比较,6 个蛋白表达下调,3 个蛋白表达上调,1 个蛋白表达缺失.结论 初步鉴定了脑出血恢复期阴虚动风证对证与非对证治疗方证效应的相关蛋白,这些蛋白功能涉及应激、细胞代谢、细胞迁移、信号转导、细胞骨架等;大定风珠对证治疗则能有效地调节多个异常表达的蛋白,非对证治疗则使蛋白表达明显异常,体现了中医辨证论治、方证相应的科学性.

Proteomics study on the syndrome-effect relationship of Dading Fengzhu decoction for intracerebral hemorrhage patients with Yinxu Dongfeng syndrome in recovery stage

Objective To explore proteins which reflect syndrome-effect relationship from the aspects of formula corresponding to symptoms and illuminate the essence of intracerebral hemorrhage （ICH） patients with Yinxu Dongfeng syndrome（YXDFS） in recovery stage by comparing the different proteins in peripheralblood mononuclear cells （PBMCs） from healthy people and ICH patients with YXDFS treated by Dading Fengzhu decoction （DDFZD） or by Zhengan Xifeng decoction（ZGXFD）. Methods ICH patients with YXDFS were randomly divided into DDFZD-treated group and ZGXFD-treated group.PBMCs were isolated from aforesaid groups and healthy subjects using lym- phocytes separation medium.The total proteins were extracted, and separated by two-dimension- al gel electrophoresis （2-DE） and analyzed by PDQuest software.The peptide mass fingerprint（PMF） was determined using matrix-assisted laser desorption ionization-time-of-flight mass spectrometry. The differentially expressed protein spots were identified using a Mascot software system. Results Gel-image analysis revealed that there were 24 protein dots expressed differentially in the ICH patients with YXDFS as compared with healthy control, of which 12 were identified. Compare with healthy subjects, there were 5 proteins up-regulated,4 proteins down-regulated and 1 protein lost in ICH patients with YXDFS. Compare with healthy subjects, 7 proteins were not differences, 2 proteins were down-regulated,2 proteins were up-regulated after treated by DDFZD; 6 proteins were down-regulated,3 proteins were up-regulated and 1 proteins were lost after treated by ZGXFD. Conclusion The proteins on the syndrome-effect relationship of treatment for ICH patients with LYFWS by DDFZD is identified, these proteins are related to stress re- sponse, cell metabolism, cell migration, signal transduction, cytoskeleton and so forth. DDFZD can adjust several abnormally expressed proteins of ICH patients with LYFWS. On the contrary, ZGXFD couldn＇t adjust such abnormally expressed proteins. It is provided the experimental basis for the principle of treatment according to differentiation of svmotoms and sizns.