肝阳上亢与肝阳化风证大鼠模型脾淋巴细胞的蛋白质组学分析

Proteomics analysis of spleniclymphocytes in SD rats with liver-yang hyperactivity syndrome and liver-yang forming wind syndrome

目的建立高血压肝阳上亢证(简称肝阳上亢证)与出血性中风、缺血性中风肝阳化风证脾淋巴细胞双向凝胶电泳图谱,并识别鉴定肝阳上亢证与肝阳化风证相同与差异表达的蛋白,探讨二证的本质内涵。方法将SD大鼠随机分为正常组、肝阳上亢证、缺血化风组及出血化风组,从大鼠一般情况、血压测定、行为学与形态学及氯化三苯基四氮唑(TTC)染色评价模型,分离大鼠脾淋巴细胞,进行双向凝胶电泳,蛋白质组双向电泳图像(PDQUEST)软件分析,目标蛋白质点用基质辅助激光解析电离质谱进行鉴定。结果成功复制3种证型动物模型。肌动蛋白β等4个蛋白在3组模型组中均较正常组表达上调,肝阳化风组较肝阳上亢组表达上调;膜联蛋白Ⅲ等3个蛋白在3组模型组中均较正常组表达下调,肝阳化风组较肝阳上亢组表达下调;膜联蛋白Ⅲ等7个蛋白在出血性化风组与缺血性化风组比较表达无明显差异;还原型烟酰胺腺嘌呤二核苷酸(NADH)脱氢酶在缺血性化风组较出血性化风组表达下调。结论肝阳上亢证与化风证既有相同的蛋白质表达又有差异的蛋白质表达,提示二证既有相同的物质基础又有不同的本质内涵,为肝阳上亢证与肝阳化风证本质研究提供了科学依据。

Objective To establish the two-dimensional gel electrophoresis（2-DE） map of the spleniclymphocytes in animal models of hypertensive with liver-yang syndrome,hemorrhagic and ischemic stroke with liver-yang forming wind syndrome,and then to elucidate the essence of the two syndromes by analyzing the identical and differential proteins between them.Methods The SD rats were randomly divided into 4 groups including the normal control group,hypertensive with liver-yang hyperactivity syndrome group,hypertensive intracerebral hemorrhage（HICH） and cerebral infarction（CI） with liver-yang forming wind syndrome group.Models were evaluated by their general conditions and behavior,the blood pressure measurement and rotating time,histological morphology and biological behavior,and the TTC staining.The spleniclymphocytes were separated by 2-DE and analyzed by PDQuest software.The target protein spots were identified by matrix-assisted laser desorption ionization-time-of-flight mass spectrometry（MALDI-TOF-MS）.Results Beta-actin and other 3 protein dots were up-regulated in all the three groups as compared with healthy control,more obviously up-regulated in liver-yang forming wind syndrome group than in liver-yang hyperactivity syndrome group.Compared with the healthy control,Annexin Ⅲ and other 2 protein dots were down-regulated in the three groups,more obviously down-regulated in liver-yang forming wind syndrome group than in liver-yang hyperactivity syndrome group.There were no significant differences between HICH and IC with liver-yang forming wind syndrome groups in the expression levels of Annexin Ⅲ and other 7 protein dots.NADH dehydrogenase was down-regulated in IC with liver-yang forming wind syndrome group as compared with HICH with liver-yang forming wind syndrome group.Conclusion There are identical and differential proteins between liver-yang hyperactivity syndrome and liver-yang forming wind syndrome,which suggests the two syndromes have the same material base but different essence.