| Depression is a dangerous mental disorder with high rate of relapse.In the past few decades,researchers have focused on the pathogenesis such as heredity,immunity,endocrinology and so on.The development and research of clinical antidepressants,for example selective serotonin reuptake inhibitors(SSRIs),are based on these aspects,but these antidepressants have obvious adverse reactions and other disadvantages.Zhi-Zi-Chi decoction(ZZCD),a classic Chinese herbal formulation,shows that it has a good effect on improving depression,insomnia and other psychological sub-health diseases in the clinical application.Currently,most studies are focused on single medicinal herb,including Gardenia jasminoides Ellis and Semen sojae praeparatum.However,there are few studies reported on the anti-depression mechanism of ZZCD and its mechanism of compatibility which can attenuate toxicity and reserve efficacy.Additionally,recent studies have found that gut microbiota which may be a new target for treatment of depression is tightly related to the occurrence and development of depression.Therefore,our study aims to deeply explore the anti-depression and compatibility attenuating toxicity mechanism of ZZCD based on gut microbiota.First of all,the depression rat model was established by the way of internationally recognized CUMS coupled with separate feeding.Sucrose preference,tail suspension test and forced swimming test were used as evaluation indexes.After 6 weeks of stimulation,rats in the model group showed a decrease in the percentage of sucrose preference and a increase in the immobility time of tail suspension and forced swimming,which indicated that the CUMS-induced depression model in rats was successfully established.Meanwhile,the partially purified extraction of ZZCD could significantly improve the abnormalities of the above indexes.Subsequently,nissl staining was performed on the hippocampus of rats.Compared with the model group,the arrangement of hippocampus CA1 pyramidal neurons were more order and the nissl bodies in the cells were more clear in ZZCD group.This part clarified the anti-depression effect of the partially purified extraction of ZZCD from behavior and histological morphology of hippocampus,which laid a foundation for the subsequent research.Gut microbiota is the key target in the brain-gut axis.The brain-gut axis is a bidirectional regulating axis where the gastrointestinal tract communicate with the central nervous system via hormones related to hypothalamic-pituitary-adrenal(HPA)axis,vagus nerve,immune system and other endogenous substances or anatomic site.Thus,gut microbiota is closely related to the occurrence of inflammation,the disturbance of 5-Hydroxytryptamine(5-HT)metabolism and other aspects.Illumina Miseq high-throughput sequencing was used to sequence the V3-V4 hypervariable region of bacterial 16 S rRNA gene.The microbial differences of caecum contents in each group were compared comprehensively by diversity and microbial composition.The results showed that there was no significant difference in the alpha diversity in each group,whereas beta diversity analysis showed that partially purified extraction of ZZCD could regulate the disturbance of gut microbiota caused by CUMS.Further studies showed that partially purified extraction of ZZCD could improve the level of beneficial bacteria such as anti-inflammatory bacteria and bacteria producing short-chain fatty acids and could decrease the level of Inflammatory bacteria and bacteria related to tryptophan metabolism to reflect changes in inflammation and tryptophan metabolism from the perspective of gut microbiota.Based on the study of gut microbiota changes,we investigated the effect of partially purified extraction of ZZCD on proinflammatory cytokines,brain-derived neurotrophic factor(BDNF),5-HT and other substances in the different parts of brain-gut axis to fully study the anti-depression mechanism of ZZCD mediated by gut microbiota.The contents of pro-inflammatory cytokines and brain-derived neurotrophic factor(BDNF)in serum,ileum and hypothalamus of rats were detected by enzyme-linked immunosorbent assay(ELISA).It was found that the partially purified extraction of ZZCD could reduce the contents of pro-inflammatory cytokines and increase the content of BDNF in different parts of CUMS rats.Subsequently,the changes of 5-HT and its related substances in serum,caecum and hippocampus of rats were detected by UHPLC-Q-TOF/MS.The results showed that partially purified extraction of ZZCD could increase the level of 5-HT and 5-Hydroxytryptamine/5-Hydroxyindoleacetic acid(5-HT/5-HIAA)and decrease the level of 5-HIAA in different parts of rats caused by CUMS.Therefore,this part illuminated the regulatory effect of partially purified extraction of ZZCD on the disorders of pro-inflammatory cytokines,BDNF and 5-HT in different parts of brain-gut axis caused by CUMS.Gardenia jasminoides Ellis has toxicity.The profound meaning of compatibility is that it can play an important role in toxicity attenuation and efficacy reservation.To elucidate this mechanism of compatibility,Illumina Miseq high-throughput sequencing was used to investigate the effect of isoflavones,the main active ingredients in Semen sojae praeparatum,on gut microbiota.Meanwhile,combination with our previous study about the effect of Semen sojae praeparatum on the metabolism of Gardenia jasminoides Ellis,our study found that non-isoflavones diet could reduce the relative abundance of bacteria related to ?-glucosidase and phase-II metabolic enzymes and increase the relative abundance of sulfate reducing bacteria.Therefore,isoflavones diet could accelerate the biotransformation of genipin into phase-II metabolites in order to excrete quickly compared with the non-isoflavones diet.Then the excretion rate of gardenia blue from the rats fed with Semen sojae praeparatum was observed.It was found that the extraction of Semen sojae praeparatumgardenia could accelerate the excretion of blue-green faeces after a week of Semen sojae praeparatumgardenia intervention to rats.Therefore,this part explained the compatibility mechanism of ZZCD in attenuating toxicity by accelerating the metabolism of genipin to phase-II metabolite and gardenia blue. |
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