高血压病肝火亢盛证血清代谢组学初步研究

Preliminary study on metabonomics of hypertension hyperactivity of liver yang syndrome

目的本文从代谢组学水平探讨原发性高血压病（EH）中医“证”的生物学基础。方法选择男性原发性高血压中医辨证为肝火亢盛证患者18例以及体检健康男性15例，入选的原发性高血压患者要求均没有服用任何降压药物，或者己停服各类药物1周以上。抽取受试者空腹静脉采血，分离血清。通过气相色谱质谱联用技术（GCMS）分析所获得的代谢指纹图谱，经谱图预处理并积分后，对积分值进行多元统计分析，根据各样本的主成分得分作图，可判别组间差异；进一步用PCA以分析进行主成分因子负荷矩阵分析。结果经过分析鉴定得出EH肝火亢盛证患者相比较健康对照组的显著性差异表达血清代谢物有13种（尸〈0．05）：棕榈酸（RT18．360）、尿酸（RT11．241）、柠檬酸盐（RT11．583）、亚油酸（RT30．139）、硬脂酸（RT25．117）、亮氨酸（RT8．478）、缬氨酸（RI13．077）、胆固醇（RT36．511）。表达降低的有：丙氨酸（RT6．444）、油酸（RT23．825）、花生四烯酸（RT28．907）、门冬氨酸（RT16．681）、甘氨酸（RT11．415）。结论EH肝火亢盛证患者血清代谢标志物群中：尿酸、尿素、棕榈酸、油酸、蛋氨酸、柠檬酸盐、亮氨酸、脯氨酸、胆固醇、硬脂酸、乳酸表达增高。甘油酸盐、丙氨酸、肉豆蔻酸、亚油酸、花生四烯酸、门冬氨酸表达降低。

Objective In the present study, we use GC/MS-based metabolomics approaches to make analysis of serum metabolic profiles of healthy people and the hyperactivity of liver yang type of constitution in patients with essential hypertension. Try to establish the discriminant model, to discover biomarkers （group） of the hyperactivity of liver yang type of essential hypertension, and to explore the essential material basis of Traditional Chinese medicine ＂syndrome＂ theory. Methods Classified according to the guiding principles of Chinese medicine research, the hyperactivity of liver yang type of constitution in male patients with essential hypertension （n = 18）, as well as health volunteers （n = 15） were randomly selected from Guang An Men Hospital clinic, wards and medical center in the first half of 2010, selected patients with essential hypertension requirements are not taking any drugs or Chinese herbs, or stop taking the various drugs more than one week. Extracted Venous Blood of subjects fasting for 12 hours, and serum was separated through centrifugation. Serum samples are stored and at --86~C refrigerator. Survey and evaluate endogenous metabolism in serum samples of health control group and types of syndrome mentioned above by gas chromatograph mass spectrometry （GCMS） analysis. Then, analyze the metabolites with Partial Least Squares-Discriminant Analysis. Further use PCA to analyze the principal component factor loadings matrix analysis, and for variable scatter plot （Loading plot）, significant increase or decrease the variables can be found from the figure. The combination of these variables is the lesion biomarkers group. Results Compared with the health control group, 13 differentially expressed metabolites in the essential hypertension hyperactivity of liver yang type group can be identified （P〈0.05）. 8 metabolites were up-regulated expression： Uric acid, citrate, Octadecanoic acid, Hexadecanoic acid, Octadecadienoic acid, Leucine, Cholesterol, Norvaline, and 5 metabolites were down-regulated expression： arachidonate, Oleate, Alanine, Aspartic acid, glycine. Conclusion We are incline to regard that the 13 of EH patient serum differentially expressed metabolites are EH hyperactivity of liver yang syndrome metabolic biomarkers： Uric acid, citrate, Octadecanoic acid, Hexadecanoic acid, Octadecadienoic acid, Leucine, Cholesterol, Norvaline, Arachidonate, Oleate, Alanine, Aspartic acid, glycine.