Comparing The Effects Of Antidepressant Chinese Herbal Medicine And Chemical Drugs On The Improvement Of Depressive Symptoms And Intestinal Microbiota In Rats Exposed To Chronic Unpredictable Mild Stress

In recent years,numerous evidences have shown that gut microbes are closely related to central nervous system diseases(depression,Alzheimer’s disease,Parkinson’s disease,etc.).Explaining the mechanism of drug intervention in diseases from the perspective of intestinal microbiota is a research hot spot of common concern in the field of medicine and microbiology.Drugs may affect the composition and function of the gut microbiota,which can be seen as an important "organ" for maintaining host health.Given the influence of intestinal flora on the biotransformation,clinical efficacy,and toxicity of drugs,the gut microbiome has become an important factor to consider when evaluating drug.However,the extent to which antidepressants affect the gut microbiome is still unknown.Chinese herbal medicine,such as Semen Ziziphi Spinosae(SZS)and Xiaoyao San(XYS)has been used in the treatment of depression in China for many years,but their active ingredients are complex and their mechanism of action is not clear.Therefore,a randomized,controlled trial was conducted to compare the effects of SZS,XYS,amitriptyline(Ami)and fluoxetine(Flu)on the improvement of depressive symptoms and intestinal microbiota based on the 16 S rRNA V4-V5 amplicon sequencing and metagenome sequencing technology using chronic unpredictable mild stress(CUMS)rats,a well-established rodent model for depression.The experiments and results are as follows:a)Body weight and an array of behavior tests(sucrose preference test(SPT),open filed test(OPT)and light/dark test(LDT))showed that CUMS stimulation successfully constructed a rat model of depression.The 16 S rRNA gene sequencing of rat feces showed that CUMS significantly reduced the alpha-diversity and altered the structure of fecal microbiota.Statistical analysis of the relative abundance of each taxa showed that there were significant differences between several taxa from phylum to genus in CUMS and HC groups,especially Bacteroidetes,Firmicutes and their ratios.At the genus level,CUMS stimulation significantly changed the abundances of Bacteroides,Parabacteroides and Ruminococcus＿2.Finally,functional prediction based on 16 S rRNA gene sequences revealed significant differences in pathways related to amino acid,carbohydrate and lipid metabolism.b)To compare the behavior improvement effects of SZS,XYS,Ami and Flu on the model rats of depression with CUMS.The results showed that the four drugs all improved the depression symptoms of CUMS rats,and the improvement effect of SZS was lower than that of the other three groups.The fluoxetine group had the best performance in open field test and light-dark test.However,in terms of body weight,the SZS and XYS improved the body weight of rats significantly better than Ami and Flu.c)Based on 16 S rRNA gene sequencing,the effects of SZS,XYS,Ami and Flu on intestinal microbiota in rats with CUMS were compared.The results showed that XYS,Ami and Flu could increase the alpha-diversity of microbiota.In addition,four drugs all altered the structure of the microbiota.In terms of microbiaota abundance,fluoxetine intervention produced the most different species,followed by amitriptyline.According to previous studies,it was found that the species that had positive effects on depression,such as Bacteroides,Butyricimonas and Parabacteroides,were all regulated to a certain extent under the action of the four drugs,and their abundance reached a level similar to that of the healthy control group(HC).However,the abundance of some species that may be harmful to human body,such as Porphyromonadaceae and Alistipes,was significantly increased only under the effects of Ami and Flu,while no difference was found between HC and CUMS groups.Finally,based on the function prediction of 16 S rRNA gene sequence,it was found that the four drugs regulated some metabolic pathways,such as carbohydrate metabolism and amino acid metabolism.d)Based on metagenomic data,the effects of XYS,Ami and Flu on fecal microbial function were compared.In terms of KEGG metabolic function,it was found that all three drugs could regulate KEGG orthologues(KOs)related to membrane transport,carbohydrate metabolism and signal transduction.In the profile of antibiotic resistance genes(ARGs),we used linear discriminant analysis effect size(LEf Se)to find that both Chinese herbal medicines and antidepressants could significantly change some ARGs.After XYS treatment,it was found that a variety of multidrug resistance related ARGs(mdtN、mdtO、mdtM、mdtK、mdtL、mdtG、mdtH、mdtP、bcr、emrD)were significantly reduced.After Ami treatment,the ARGs associated with aminoglycosides,tetracycline and chloramphenicol(aph3iiiA、tetM、lsA、catb2)were significantly enriched,while the ARGs associated with multidrug resistance(bcr、emrD)were significantly decreased.Nine of ARGs were significantly different after Flu intervention,including multidrug resistance(mdtK、mdtP、mdtH、mdtG 和 acrA),aminoglycoside(aph3iiiA),tetracycline(tetM),and chloramphenicol(cml＿e8).Among them,the abundance of ARGs associated with multidrug resistance was still significantly reduced.Finally,co-occurrence network analysis showed that there was a strong correlation between the gut microbiota and ARGs.