基于代谢组学研究黄连-黄芩药对治疗脑缺血大鼠的作用机制

Mechanism of treatment effect of Huanglian-Huangqin herb pairs on cerebral ischemia rats based on metabolomic approach

运用代谢组学方法研究黄连-黄芩药对对脑缺血模型大鼠代谢的作用机制。采用线栓法致大脑中动脉阻塞复制缺血再灌脑损伤大鼠模型(MCAO)。通过超高效液相色谱-串联四级杆飞行时间质谱(UPLC-Q-TOF/MS)的方法、Markerlynx软件、以及主成分分析法和偏最小二乘判别法分析正常大鼠、脑缺血模型大鼠以及分别给予黄连(HL)、黄芩(HQ)和黄连-黄芩药对(LQ)干预后大鼠尿液样本中的内源性代谢物的差异,结合内源性代谢物的一级及二级碎片离子的精确信息,检索并鉴定可能的生物标志物,进而在MetPA数据库中构建代谢通路。结果在脑缺血模型大鼠的尿液中发现并鉴定出了20种生物标志物,并对其相对含量及代谢途径进行分析。主要涉及神经递质调节、氨基酸代谢、能量代谢、脂质代谢等。这些代谢通路在脑缺血模型大鼠的体内发生紊乱,主成分分析表明,正常与脑缺血模型得到明显区分,而给予HL,HQ,LQ后可以使之向着正常状态转变。此研究对脑缺血大鼠代谢组及黄连-黄芩药对干预的作用机制进行阐释,体现代谢组学可反映生物体的生理及代谢状态,能较好地体现药对是中医遣方用药的特色优势,具有内在的组合变化规律以及中医药的整体观、系统观以及多成分协同或拮抗作用的特点。

The metabolic effect of Huanglian-Huangqin herb pairs on cerebral ischemia rats was studied by using metabolomic method. The rat model of ischemia reperfusion injury induced by introduction of transient middle cerebral artery occlusion （MCAO） followed by reperfusion. Ultra high performance liquid chromatography-series four pole time of flight mass spectrometry method（ UPLC-Q- TOF/MS） , Markerlynx software, and principal component analysis and partial least-squares discriminant analysis were used to analyze the different endogenous metabolites among the urine samples of sham rats, cerebral ischemia model rats, Huanglian groups （ HL）, Huangqin groups （HQ） and Huanglian-Huangqin herb pairs groups （LQ） was achieved, combined with accurate information about the endogenous metabolites level and secondary fragment ions, retrieval and identification of possible biological markers, metabolic pathway which build in MetPA database. The 20 potential biomarkers were found in the urine of rats with cerebral ischemia, which mainly in-volved in the neurotransmitter regulation, amino acid metabolism, energy metabolism, lipid metabolism and so on. Those metabolic pathways were disturbed in cerebral ischemia model rats, the principal component analysis showed that the normal and cerebral ischemia model is clearly distinguished, and the compound can be given to the normal state of change after HL, HQ, LQ administration. This study index the interpretation of cerebral ischemia rat metabolism group and mechanism, the embodiment of metabonomics can reflect the physiological and metabolic state, which can better reflect the traditional Chinese medicine as a whole view, system view and the features of multi ingredient synergistic or antagonistic effects.