基于儿茶酚胺代谢途径的寒、热体质大鼠靶向代谢组学研究及中药的调整

Constitution of TCM what is adjustable is the inherent qualities and personality traits of human. It is the base of preventive treatment of disease, relate to the pathogenesis, prevention and cure. Cold and Heat Constitution is the basic type of various kinds of constitution classification. To identify the Normal, Cold and Heat constitution rats from a group of natural rats, Cold constitution rats are treated with Rougui (cinnamon) with nature of hot, and Heat constitution rats are treated with Zhimu (anemarrhenae) with nature of cold. LC/MS is used to the targeted analysis of precussor substances and metabolites which belong to catecholamine in urine. In order to find out the key protease, which leads to the differences between the two constitutions, Catecholamine, synthetase and metabolic enzymes are observed by HPLC、ELISA、RT-PCR and Western blotting. Based on the targeted metabolomics, we target at the pathway of catecholamine that adjust energy metabolism of Cold and Hot constitution, explore the key enzymes that result in difference of two constitutions, explain the material basis and mechanism. From the impact of catecholamine pathway in rats, the adjustable action of chinese herbal medicine will be confirmed, and the mechanism of adjustable action could be found out. The research will provide objective biological basis for adjustable constitution theory and achieve application of preventive treatment of disease thought in modern society.

中医体质是人体固有特质和个性特征，具有可调性，与发病、防病及治病之间密切相关，是“治未病”的基础。寒、热体质是各种体质分类法中最基本的类型。本研究从自然群体大鼠中筛选出常体、寒体和热体体质大鼠，通过热性药肉桂、寒性药知母分别对寒体和热体大鼠进行体质调整，运用LC-MS对尿液儿茶酚胺的前体物质和代谢物进行靶向分析，利用HPLC、ELISA、RT-PCR和WB等方法，观察中药调整前后大鼠儿茶酚胺、递质合成酶与代谢酶的变化，找出通路中造成寒、热体质差异的关键性蛋白酶。从靶向代谢组学的角度，对参与调节寒、热体质大鼠能量代谢的儿茶酚胺途径进行针对性研究，探寻该途径中造成寒、热体质差异的关键蛋白酶，阐释产生体质差异的物质基础和机制。从对大鼠儿茶酚胺途径的影响，明确中药对寒、热体质调整作用，阐明调整作用的机制，为体质可调性理论及实现“治未病”思想在现代社会的应用，提供客观生物学依据。

寒、热体质是最基础的体质类型。对造成寒、热体质差异的生物学基础及中药对寒、热体质的调整作用进行研究，对于指导辨体论治和“治未病”都具有重要的意义。本课题从自然群体大鼠中筛选常体、寒体和热体体质大鼠，通过热性药肉桂、寒性药知母分别对寒体和热体大鼠进行为期12周的体质调整，运用LC-MS对儿茶酚胺代谢途径分析，发现寒、热体质大鼠血浆中DA、NA、Epi和尿液中Tyr、Lev、VMA存在差异；通过分子生物学验证，发现是代谢途径上的关键合成酶DβH、PNMT和代谢酶COMT、MAO在mRNA转录水平和蛋白表达上存在差异。在使用中药调整体质后，上述指标都不同程度的向正常体质趋近，提示儿茶酚胺通路的激活水平差异是造成寒、热体质大鼠体质差异的重要原因。中药对寒、热体质具有调整作用，作用机制是调节儿茶酚胺代谢通路上关键酶表达，继而影响儿茶酚胺类物质的合成与代谢，最终对寒、热体质大鼠整体状态造成影响。