The Synergistic Anti-depression Effects Of Different Efficacy Groups Of Xiaoyaosan As Demonstrated By The Rough Set,metabonomics And Network Pharmacology

Rationales: Traditional Chinese medicines(TCMs)owning classic TCM theories and unique treatment modes are attracting more and more attentions for their reliable curative effects but less side effects.However,the laws of TCM compatibility are the focus and the difficulty of TCM prescription researches.Our research group has been devoted to the research of Xiaoyaosan(XYS)for more than 15 years.The antidepressant effects and mechanisms of XYS have been explained from the perspectives of pharmacodynamics,pharmacokinetics,metabolomics,and network pharmacology.Although the compatibility of XYS has important research values,the compatibility researches of XYS are limited to the aspects of disassembled prescription and pharmacodynamic effects.Moreover,further explanations have not been mentioned.It is found that data mining technology has unique advantages for the complex and the non-linear "black box" data of TCMs.Metabolomics of systems biology can be used not only to comprehensively evaluate the endogenous metabolites of biological fluids or tissues,but also to systematically identify and quantify these metabolites,which coincides with the overall concept of TCMs.The characteristics of multi-components and multi-targets of network pharmacology are corresponding to the complex characteristics of component targets of TCMs.And it also has unique advantages in studying TCM prescriptions.The applications of modern scientific technologies to clarify the compatibility of XYS have great significances to the quantitative study of TCM prescriptions.The depression model in rats was constructed by chronic unpredictable mild stress(CUMS).The effects of XYS and its efficacy groups on CUMS rats were investigated in the sucrose preference test,the open field test and the forced swimming test.Afterwards,based on the behavioral data,the rough set was applied to establish decision attributes and decision rules to quantitatively characterize the relationships between the efficacy groups.And then,the metabolomic was used to reveal the compatibility relationship of efficacy groups of XYS from the perspectives of differential metabolites and metabolic pathways.Finally,the potential mechanisms of antidepressant compatibility of XYS were investigated by network pharmacology.Based on the multidisciplinary approaches abovementioned,revealing the antidepressant compatibility of XYSs efficacy groups will help us scientifically understand the essences of compatibility of XYS.And it will also lay a foundation for the modernization and the internationalization of TCM researches and developments.Objective: 1.Rough set analysis was established to quantitatively characterize the compatibility relationship of Xiaoyaosan’s efficacy groups on depression.And it provides an approach to quantitatively study compatibility relationship of TCM prescriptions.2.Metabolomics and network pharmacology were used to reveal the antidepressant compatibility of Xiaoyaosan’s efficacy groups through determining micro-indicators,so as to explain the contribution of different efficacy groups to the anti-depression effect of the whole prescription.Methods: 1.The depression model in rats was constructed by CUMS.The effects of XYS and its efficacy groups on CUMS rats were investigated in the open field test,the sucrose preference rate and the forced swimming test.2.Rough set was used to establish the decision conditions and the decision rules of behavioral data.And the interactions between the treatment groups and their antidepressant effects were analyzed through the decision rules.3.1H NMR serum metabolomics approach coupled with multivariate data analysis was used for determining the metabolic profiles,serum metabolites and metabolic pathways of rats.And Rd was calculated to quantitatively evaluate the efficacies to explore the relationships of antidepressant compatibility of XYS. 4.Network pharmacology was used to preliminarily reveal the antidepressant compatibility mechanism of XYS.First,we used the TCMSP and Gene Cards databases to identify the chemical components and targets of XYS and its efficacy groups.We then analyzed “herb-component-target” pathways and constructed PPI networks.Potential pathways were determined by gene ontology(GO)and pathway enrichment analyses.Finally,molecular docking was used to dock the effective components and targets of XYS and its efficacy groups.Results: 1.XYS and its efficacy groups significantly improved the depressive behaviors of CUMS rats.Compared with the model group,XYS and its efficacy groups significantly increased the number of crossings and rearing of the rats in the open field test(P < 0.05)and the sucrose preference rate(P < 0.05).XYS and its efficacy groups significantly reduced the immobility time of forced swimming(P < 0.05).The order of improving the sucrose preference rate was XYS group > Jianpi group > Shugan group.The order of improving the behaviors in the open field test was Shugan group > Jianpi group > XYS group.The order of decreasing the time of forced swimming index was XYS group > Shugan group > Jianpi group.The order of improving the index in the Morris water maze test was XYS group > Shugan group > Jianpi group 2.The results of decision rules showed that,when used alone,Shugan and Jianpi groups exhibited certain antidepressant effects,which however were weaker than that of XYS.Of note,Shugan and Jianpi groups exhibited antidepressant effects in a synergetic manner.Such a synergistic effect was dose-dependent,i.e.,the higher the dose was,the greater the effect was.According to the decision table of XYS and its individual herbs,a table of attribute reduction of 8 herbs in XYS was conducted.By doing so,we found that Radix Bupleuri and Radix Paeoniae Alba were the representative drugs of Shugan and Jianpi group,respectively.3.Metabolomics results show that 10 different metabolites were identified to be related to depression,involving 5 metabolic pathways.Compared with the control group,the level of lactic acid in the serum of the model group decreased,while the levels of 3-hydroxybutyric acid,acetoacetate,succinate,trimethylamine oxide(TMAO),taurine,glycerol,acetic acid,and citrulline and α-glucose,etc.,increased.XYS could regulate 6 differential metabolites,5 for Shugan group and 7 for Jianpi group,respectively.Meanwhile,the XYS and the Jianpi group adjusted all the five pathways while the Shugan group only regulated four pathways.We found that the XH group had the highest Rd than other groups.The results of metabonomic showed that XYS could increase energy metabolism and neurotransmitter level by regulating the metabolism of carbohydrates,amino acids and intestinal microorganisms.4.We identified a total of 121 components of XYS and 111 targets that were associated with depression.Additionally,we screened 105 targets of 24 components in Shugan group and 106 targets of 95 components in Jianpi group.13 targets were in common for all the three groups.PPI network analysis showed that CASP3 and NOS3 were nodes that overlapped with proteins that known to be associated with depression.Pathway enrichment analysis further indicated that the putative targets of XYS and its efficacy groups mostly participated in pathways in cancer.XYS and its efficacy groups mainly played an antidepressant role by participating in glucocorticoid response,drug response and nitric oxide biosynthesis,regulating calcium channel,tumor necrosis factor signaling pathway and neurotrophic signaling pathway.Conclusion: XYS and its efficacy groups can not only improve the core symptoms of depression,including index in open field test,sucrose preference rate and forced swimming index,but also significantly improve and alleviate the decline of spatial learning and memory ability of depression rats,showing remarkable antidepressant effects.In terms of data mining,metabonomic and network pharmacology,from the macro to micro comprehensive point of view, we found that as containing different effective components,the Shugan group and Jianpi group differently contributions to the antidepressant effects of Xiaoyaosan at different levels and perspectives,which suggested a reasonable compatibility of XYS.The results provide not only a theoretical basis for the scientific interpretation of antidepressant compatibility of XYS,but also methodology for the compatibility relationships of TCM prescriptions.The current study also demonstrates the scientific connotations and the compatibility rules of TCMs from an overall perspective.