| Depression is a common mental disorder,which is a public health and social problem that seriously affects the quality of human life and social and economic development.Up to now,the etiology and pathogenesis of depression are still not very clear.At present,most of the antidepressants commonly used in clinic are chemical drugs which are often accompanied by certain side effects,and the target of these drugs is single which easy to develop drug resistance.It is worth noting that traditional Chinese medicine prescription can simultaneously regulate multiple links of the disease network through its multi-components with high efficacy and low side effects,suitable for long-term use,and has a unique advantage in the treatment of depression.Zhi-Zi-Chi decoction comes from Treatise on febrile Diseases,which has the effect of dispelling annoyance.The clinical application study of modern Chinese medicine shows that Zhi-Zi-Chi decoction has a good effect on regulating and improving depression,but themechanism of antidepressant and material basis of efficacy are not clear.The previous study of our group found that Zhi-Zi-Chi decoction had an effect on brain function and behavior through intestinal flora-gut-brain axis,thus playing an antidepressant role.Among them,the main chemical component geniposide also has a certain antidepressant effect,which may be the main antidepressant substance of Zhi-Zi-Chi decoction.Based on this,the purpose of this study is to explore the effects of geniposide on the microbial ecology of gut microbiota from the point of view of molecular microbial ecology,molecular biology and metabonomics.to reveal the regulatory mechanism of gut microbiota on the level of neurotransmitters,inflammatory cytokines and brain-derived neurotrophic factor(brain-derived neurotrophic factor,BDNF)in different parts of the braingut axis under the intervention of geniposide.Thus,the antidepressant mechanism of geniposide mediated by intestinal flora-intestine-brain axis was revealed at the molecular level,and the material basis of Zhi-Zi-Chi decoction was clarified,so as to provide ideas and reference for screening new antidepressant drugs with intestinal microflora as a new target.Firstly,we used the classic chronic unpredictable mild stress(chronic unpredictable mild stress,CUMS)combined with orphanage to establish the depression model of SD rats.Low,middle and high dose groups were set up and fluoxetine was used as positive control.Sucrose preference test(sucrose preference test,SPT),tail suspension test(tail suspension test,TST)and forced swimming test(forced swimming test,FST)were used to evaluate the consequence.The results showed that after administration of geniposide,the degree of sucrose preference in depressed rats increased,and the effect of middle and high dose groups was better than that of low dose groups.This shows that middle and high doses of geniposide can improve the depressive symptoms of depressed rats to a certain extent,and have a certain antidepressant effect.Because there is no significant difference between the middle dose group and the high dose group,the follow-up experiment will only be carried out for the middle dose group.After nissl staining was performed on the hippocampus of rats.,it was found that the state of vertebral cells in hippocampal CA1 region of model group was abnormal.Geniposide and fluoxetine could repair the damage to some extent,and the repair effect of geniposide group was significantly better than that of fluoxetine.The results showed that geniposide could improve the behavior and hippocampal histomorphology of CUMS-induced depression rats.It was shown that geniposide is a medicinal substance that exerts an antidepressant effect of Zhi-Zi-Chi decoction.Through the metabonomic analysis of rat brain tissue,it can be seen that CUMS stimulation can cause disorders in the metabolism of arginine、 proline、 tryptophan、tyrosine 、 primary bile acid biosynthesis and glutathione.Geniposide can regulate the metabolism of tryptophan 、sphingolipid and glutathione in the brain of CUMS-induced depression rats.The ratio of glutathione(glutathione,GSH)to glutathione disulfide(glutathione Disulfide,GSSG)can reflect the body’s redox equilibrium state.Based on the regulatory effect of geniposide on the metabolism of glutathione,the activities of superoxide dismutase(superoxide dismutase,SOD),catalase(catalase),glutathione reductase(glutathione reductase,GR)and glutathione peroxidase(glutathione peroxidase,GPx)and the contents of reactive oxygen species(reactive oxygen species,ROS),GSH and GSSG in rat brain were determined.The results showed that geniposide could reduce the contents of ROS and GSSG and increase the content of GSH in depressed rats.At the same time,it can increase the enzyme activity of SOD,GR and GPx.The results shown that geniposide could enhance the antioxidant capacity of the body and improve the symptoms of depression by regulating oxidative stress.We found that geniposide has a significant regulatory effect on the metabolism of brain tissues as well as oxidative stress responses.However,due to the presence of the blood-brain barrier,the bioavailability of oral geniposide is low and it is difficult to act directly on the brain.The gut microbiota-gut-brain axis is a bidirectional regulatory axis of braingastrointestinal interactions.Therefore,we speculate that geniposide may act on gut microbiota and play an antidepressant effect through the brain-gut axis.The microorganisms in the cecal contents of rats in each group were analyzed by Illumina Miseq high-throughput sequencing,the diversity of gut microbiota in each group was analyzed by α-diversity andβ-diversity analysis,and the species composition was analyzed at the phylum and genus levels.The results showed that the diversity of gut microbiota in depressed rats was reduced,and the microflora structure was disturbed.The administration of geniposide can improve the disorder of gut microbiota in CUMS-induced depression rats.At the same time,it can affect the level of intestinal bacteria related to tryptophan metabolism,oxidative stress and inflammation,and affect depressive symptoms from the aspects of tryptophan metabolism,oxidative stress and inflammation.Based on the effect of geniposide on inflammatory bacteria and anti-inflammatory bacteria in gut microbiota,the levels of tumor necrosis factor-α(tumor Necrosis Factor,TNF-α),interleukin-1β(interleukin-1β,IL-1β),interleukin-6(interleukin-6,IL-6),interleukin-10(interleukin-10,IL-10)and BDNF in different parts of the brain-gut axis were detected.The results showed that there are similar trends in brain tissue,plasma,and cecum.Geniposide could decrease the levels of pro-inflammatory factors and increase the levels of anti-inflammatory factors and BDNF in CUMS-induced depression rats.It is clear that geniposide can play an antidepressant effect by regulating the disorders of pro-inflammatory factors,anti-inflammatory factors and BDNF in different parts of the brain-gut axis mediated by gut microbiota.Based on the regulatory effect of geniposide on gut microbiota related to tryptophan,the metabolism of tryptophan in different parts of the brain-gut axis was studied.It was found that CUMS can up-regulate the levels of kynurenine(kynurenine,Kyn)and 5-hydroxyindoleacetic acid(5-Hydroxyindoleacetic acid,5-HIAA),down-regulate the level of5-Hydroxytryptamine(5-Hydroxytryptamine,5-HT),and weaken the metabolism of tryptophan(tryptophan,Trp)to 5-HT,while geniposide can regulate the levels of Kyn,5-HIAA and 5-HT,and enhance the metabolism of Trp to 5-HT.In addition,different parts of the brain-gut axis had the same trend.It can be concluded that geniposide can act on the gut microbiota related to tryptophan metabolism,and then regulate the metabolism of tryptophan in the different parts of the brain-gut axis,and finally improve the symptoms of depression.This study revealed the effect of geniposide on tryptophan metabolism through brain-gut axis and the mechanism of improving depressive symptoms from the point of view of gut microbiota.In summary,this study studied the structural characteristics of physiological intestinal microecology of normal rats and depression model rats,and compared the differences of microflora before and after administration of geniposide to find the specific gut microbiota related to the antidepressant effect of geniposide.The traditional pharmacology,molecular biology and metabonomics methods were combined to reveal the regulation mechanism of gut microbiota under the intervention of geniposide on the levels of neurotransmitters,inflammatory cytokines and brain-derived neurotrophic factor in different parts of the braingut axis.The molecular mechanism of antidepressant effect of geniposide mediated by intestinal flora-gut-brain axis was revealed from the point of view of metabonomics and molecular biology,respectively,and the material basis of antidepressant effect of Zhi-ZiChi decoction was determined.The purpose of this study is to provide ideas and reference for screening antidepressant prescriptions with gut microbiota as a new target. |
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