| Objective: In order to observe the biological differences between cold body,heat body and normal body rats,we studied the whole characterization of cold body,heat body and normal body rats and the expression of TRP protein channel.In the aspect of metabolism,metabonomics technology is used to clarify the mechanism of metabolism pathway caused by the difference of cold and hot constitution and the related mechanism of regulating body function of traditional Chinese medicine,so as to provide biological basis for the theory of adjustable constitution and the thought of "treating disease before disease" of traditional Chinese medicine.Methods: 1.In this experiment,the rats with normal,heat and cold body constitution were screened out by measuring their palms temperature combined with their own activities and general conditions.The autonomic activities of the three groups of physical rats were observed by the autonomic activity detection system.Then,the thermal pain threshold of rats with normal body,heat body and cold body constitution was measured by the method of photothermal plantar tingling,and the correlation analysis was made on the palmar temperature and autonomic activity and thermal pain threshold of the three groups of rats.UPLC-MS was used to detect the urine of the three constitution rats,and the differences in urine metabolic profiles of the three constitution rats were compared.The principal component analysis(PCA)and partial least squares discriminant analysis were used.Partial least square-discriminant analysis(PLS-DA)was used for data mining,and PRT analysis was used to verify the method.And use Metabo Analyst online tool and KEGG(Kyoto Encyclopedia of Genes and Genomes)database for enriched topology analysis.2.The expression of TRPV1 and TRPV4 channels in the hypothalamus of the heat body group was significantly higher than that of the cold body group.3.In the non-target metabolomics,the urine samples of the heat body group and the cold body group were separated on the PCA map,but could not be completely separated.On the OPLS map,it can be concluded that the urine samples of the normal body group and the cold body group,the heat body group and the cold body group,the heat body group and the normal body group can be completely separated,and the urine metabolites of the normal body group,the cold body group and the heat body group can be displayed at the same time.Four final differential metabolites were identified as EPI,NE,Tyr and VMA.According to different biomarkers,two metabolic pathways are finally enriched,one is phenylalanine,tyrosine and tryptophan biosynthesis pathway,the other is tyrosine metabolism pathway.Through online platform analysis and literature review,it was found that the metabolic difference was concentrated in catecholamine metabolic pathway of tyrosine metabolic pathway.4.Based on the results of non targeted metabonomics,targeted metabonomics was carried out around the catecholamine metabolic pathway.It was found that there were differences in EPI,DA and NE between the cold body group and the heat body group.The contents of the three substances in the cold body group were lower than those in the normal body group and the heat body group,while those in the heat body group were higher than those in the normal body group.There were differences in the contents of Tyr,LEV,NMN and VMA in the urine between the cold body group and the heat body group.The contents of LEV,NMN(Tyr,VMA)in the urine of the cold body group were lower(higher)than those of the normal body group and the heat body group,while the heat body group was the opposite.5.The differences of palmar temperature,autonomic activity and thermal pain threshold between cold body group and heat body group disappeared after 12 weeks of Chinese medicine regulation,and all of them were close to the normal body group.The expression of TRPV1 and TRPV4 channels in the hypothalamus of rats increased in the cold body group,but decreased in the heat body group,but there was no significant difference between the normal body group and the cold body group.According to the target Metabonomics Study of catecholamine metabolism pathway,the differences of Epi,DA,NE in plasma between cold body group and heat body group disappeared and approached to the normal body group,and the differences of Tyr,LEV,NMN,VMA in urine also disappeared and approached to the normal body group.Result: 1.There are significant differences in the thermal pain threshold among the normal,cold and heat body rats,and the thermal pain threshold is negatively correlated with the palm temperature of the rats.The differences in the expression of TRPV1 and TRPV4 channels in the hypothalamus may be the molecular biological basis for the differences in the perception of palm temperature and thermal pain in the cold and heat body rats.2.There are different metabolites and metabolic pathways among the three groups,including Epi,NE,Tyr and VMA.The enriched differential metabolic pathway is catecholamine metabolic pathway according to the literature investigation.3.There were differences in catecholamine metabolism among the three groups.Under the natural condition,the activation level of catecholamine pathway in the heat body group was higher than that in the normal body group,and that in the cold body group was lower than that in the normal body group.4.12 weeks later,the difference of thermal pain threshold,autonomic activity and hypothalamic TRP channel disappeared,and the activation level of catecholamine metabolic pathway tended to normal rats,indicating that the effect of traditional Chinese medicine on regulating cold and heat constitution was related to the expression of hypothalamic TRP channel and the activation level of catecholamine metabolic pathway. |
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