Plasma Metabolomic Study On Hypertension Rats With Syndrome Of Pathogenic Fire Derived From The Stagnation Of Liver Qi And Intervention Of Traditional Chinese Medicine

Object:To explore the construction of a stable animal model of Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi,and to explore the biological basis of this model based on metabolomics,to find differential metabolites,and to explore the relationship between differential metabolites and macroscopic characterization;and to observe the effect of Danzhixiaoyao Powder,a classical therapeutic prescription,on syndromes and their metabolism.Methods1.The establishment of Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver QiSpontaneous hypertensive rats,which are most similar to human essential hypertension,were used to establish a Traditional Chinese Medicine model by applying external interfering factors,combined with the equivalent transformation of clinical symptoms and animal characterization,to evaluate them from three dimensions:macroscopic characterization combination,microscopic physical and chemical indicators and prescription counterevidence,to determine the attributes,and to determine the correlation between macroscopic characterization and animal characterization by correlation analysis.Rats were randomly divided into control group,model group,and Danzhixiaoyao powder group.Except for the control group,the other two groups were given chronic restraint stress for 4 weeks.Meanwhile,the Danzhixiaoyao powder group was given Danzhixiaoyao powder aqueous extract(5g/kg/d,i.g.)at a dose of 10 mL/kg,and the model group was given an equal volume of pure water.The data of body weight,blood pressure,residence time ratio of open arm of elevated plus maze,rotation tolerance time and irritability degree of animals were collected every week,and the data of tongue color r value were collected once every 2 weeks.Four weeks later,the animals were sacrificed and plasma was collected,adrenals and thymus were harvested.2.Plasma metabolomics analysisUHPLC-QTOF-MS metabolomics technology was used to analyze and identify endogenous metabolites in animal plasma,and multivariate statistical methods such as principal component analysis and orthogonal partial least squares discriminant analysis were used for differential metabolite analysis.Metabolic pathway analysis was performed for metabolites with a secondary mass spectrometry reliability greater than 0.8(full value of 1)using the Metaboanalyst database as well as the KEGG database.3.correlation analysis between differential metabolites and macroscopic characterization indicatorsSpearman’s method was used to correlate differential metabolites with macroscopic characterization indicators.Correlation metabolites satisfied P<0.05 and Spearman coefficients≥0.4using Spearman’s test.Result1.The establishment of Hypertension Rats with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver QiIn the model group,the blood pressure increased,the body weight decreased,the residence time ratio of the open arm of the elevated plus maze decreased,the degree of irritability increased,the rotation endurance time decreased,the tongue color r value increased,the thymus and adrenal gland indices decreased,and the plasma CORT concentration increased.Compared with the model group,the blood pressure,rotation tolerance time and tongue color r value of Danzhixiaoyao powder were significantly improved.Correlation analysis showed that body mass,blood pressure,ratio of open arm time of elevated plus maze,degree of irritability,rotation tolerance time,and tongue color r value had significant correlation with Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi,and the correlation degree was high.The correct rate of classification of Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi by the above indicators could reach 92.9%,indicating that the selection of indicators in this experiment was objective and credible.Therefore,we preliminarily concluded that the model met the diagnostic criteria for Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi.2.Plasma metabolomics analysisThe model control group was significantly dispersed on the PCA diagram,and the two groups were significantly separated in the OLPS-DA analysis,with a total of 23 differential metabolites,involving triglyceride metabolism,glycine serine and threonine metabolism,phenylalanine metabolism,tryptophan metabolism,and steroid hormone synthesis pathways;the model group was well dispersed on the PCA diagram of Danzhixiaoyao powder,and the two groups were well separated in the OLPS-DA analysis,with a total of 13 differential metabolites,involving glycine serine and threonine metabolism,tryptophan metabolism,and steroid hormone synthesis pathways.3.Correlation analysis of differential metabolites and macro-characterization indicatorsBlood pressure,rotational endurance time,and elevated open-arm time ratio were closely related to tryptophan metabolism;tryptophan was positively correlated with rotational endurance time and irritability degree;the tryptophan metabolite acetylcarnitine was extremely negatively correlated with irritability degree;and creatine was negatively correlated with elevated open-arm time ratio.Conclusion1.Selected the spontaneously hypertensive rats,combine with the way of Traditional Chinese Medicine model establishment,used the method of combining macroscopic characterization with microscopic indicators and drug counterevidence to determine the model of combining disease and syndrome,and successfully established Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi.2.Metabolomics analysis of the model control group suggested that the biological basis of Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi may be related to triglyceride metabolism,glycine serine and threonine metabolism,phenylalanine metabolism,tryptophan metabolism,and steroid hormone metabolism.Danzhixiaoyao powder can improve triglyceride metabolism,tryptophan metabolism,steroid hormone metabolism to achieve the therapeutic effect on Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi.3.Correlation analysis between differential metabolites and macroscopic characterization of Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi suggests that macroscopic characterization of Hypertension with Syndrome of Pathogenic Fire Derived from the Stagnation of Liver Qi may be related to the decrease in tryptophan,acetylcarnitine,and creatine.