| Background: Depression,as a common neuropsychiatric disease in the world,has attracted much attention.According to the latest report of the World Health Organization(WHO),3.8% of the world’s population suffers from depression,and adults account for about 5%.Studies have found that the composition and structure of the gut microbiota in patients with depression are significantly different from those in healthy people,and there have been clinical cases of successful treatment of neuropsychiatric diseases by transplanting fecal microbiota.In recent years,the role of gut microbes in the human body cannot be underestimated,and its influence on central nervous system diseases such as depression has gradually entered the public eye,and the microbegut-brain axis theory has emerged as the times require.Tryptophan metabolism has been proposed to affect the microbiota-gut-brain axis at multiple levels,and the two pathways of 5-HT metabolism and kynurenine metabolism play a particularly significant role in it.There is also a complex relationship between exercise and the microbiome-gut-brain axis,which exerts its benefits mainly by affecting the composition and diversity of the gut microbiota,the endocrine system,the autonomic nervous system,and the immune system.However,the effects of different exercise intensities and exercise patterns on neuropsychiatric diseases have always been controversial and the specific mechanism has not yet been clarified.Objective: From the perspective of microbial-gut-brain axis,based on chronic restraint stress(CRS)model of depressed mice,this study adopted fecal bacteria transplantation(FMT),to explore the possible association mechanism between intestinal microbiota,tryptophan metabolism and depression,and to explore the effect of changes in intestinal microbiota caused by different exercise intensities on depressive behavior of CRS mice and the possible mechanism,so as to provide reference for improving the effectiveness and accuracy of exercise intervention.Methods: In this experiment,a total of 117 healthy male mice of SPF grade C57BL/6 were selected,and the mice were randomly divided into two parts according to the experimental design:feces donor mice and feces recipient mice.Donor mice were randomly divided into the following4 groups:(1)SED group(n=6),i.e.quiet control group,which was fed normally for 6 weeks without any intervention;(2)LE group(n=5),i.e.low In the intensity exercise group,the experimental mice underwent low-intensity treadmill exercise(6m/min)for 6 weeks;(3)ME group(n=5),that is,the moderate-intensity exercise group,the experimental mice underwent moderate-intensity treadmill exercise for 6 weeks Exercise(12m/min);(4)HE group(n=5),namely the high-intensity exercise group,the experimental mice underwent high-intensity intermittent treadmill exercise(18-20m/min + 4m/min)for 6 weeks.Recipient mice were randomly divided into the following 8 groups:(1)SED+CON group(SC,n=12),control group receiving fecal microbiota transplantation from quiet group donor mice,(2)LE+CON group(LC,n=12),the control group receiving fecal microbiota transplantation from donor mice in the lowintensity exercise group,(3)ME+CON group(MC,n=12),the control group receiving fecal microbiota transplantation from donor mice in the moderate-intensity exercise group(4)HE+CON group(HC,n=12),a control group that received fecal bacteria transplantation from donor mice in the high-intensity intermittent exercise group.The mice in the above four groups were fed normally for 4 weeks,and then treated with antibiotics for 1 week,and then received FMT for 3weeks;(5)SED+CRS group(SR,n=12),restraint stress group receiving fecal bacteria transplantation from quiet group donor mice,(6)LE+CRS group(LR,n=12),receiving lowintensity exercise Restraint stress group of donor mice transplanted with fecal bacteria,(7)ME+CRS group(MR,n=12),restraint stress group of donor mice transplanted with moderateintensity exercise,(8)HE+CRS group(HR,n=12),restraint stress group receiving fecal bacteria transplantation from donor mice of high-intensity intermittent exercise group,recipient mice of the above four groups underwent CRS for 4 weeks,and then received antibiotic treatment for 1week,followed by FMT for 3 weeks.After all the experimental interventions,the mice in each group were tested for depression behavior,namely the sucrose preference test,the forced swim test,the tail suspension test and the open field test.Afterwards,the mice were anesthetized with gas and blood was taken,and the hippocampus were collected after neck dislocation.16 Sr RNA sequencing technology was used to detect the composition and structure of the bacterial flora in mouse fecal samples;Elisa technology was used to detect the contents of 5-HT,TRP,KYN,IFN-γ,3-HK,and KYNA in mouse serum;Real-time PCR technology was used to detected the m RNA expression levels of TPH2,MAO,IFN-γ,IDO,KAT,and KMO in the hippocampus;the protein levels of TPH2,IDO,KAT,and KMO in the hippocampus were detected by automatic western blotting(WES).Results:(1)Effects of different intensity of exercise on intestinal flora of mice(16Sr RNA gene sequencing results): The community diversity of intestinal microbiome of mice showed that there was a significant difference in α diversity between mice in HE group and those in ME group(P<0.05).The species dilution curve showed that the species abundance and distribution of intestinal flora in SED,LE,ME and HE groups were relatively high.In terms of β diversity,PCo A analysis showed that the diversity of intestinal flora among the four groups was different,and the difference between SED group and LE group was great.The relative abundance of intestinal flora of mice showed that the fecal abundance of Bacteroidetes,Firmicutes and actinomyces was higher in the four groups of mice at the phylum level.As for Bacteroidetes,the expression abundance of LE group was significantly decreased compared with SED group(P<0.05).At the genus level,Muribaculaceae,Dubosiella and lleibacterium were relatively abundant in feces of the four groups.In terms of Muribaculaceae,the expression abundance of mice in LE group was significantly decreased compared with that in SED group(P<0.01).The expression abundance of mice in LE group was significantly increased compared with SED group(P<0.05),and the expression abundance of mice in ME group and HE group was significantly decreased compared with LE group(P<0.05).In terms of lleibacterium,the expression abundance of mice in ME group was significantly increased compared with HE group(P<0.05).The phylogenetic branch of different species of mice intestinal flora showed that the main microorganisms in SED group were Euobacteriaceae,Euobacteriaceae,and Vibrio desulphurica.The main microorganisms in LE group were Coprococcus,Lactobacillus,Nocardia and Flavobacteria.In the ME group,Bifidobacteria,Bifidobacteria,Bifidobacteria,Actinobacteria,Coriobacteriaceae＿UCG＿002,Coriobacteria,Adlercreutzia and Oribacterium played the main roles.Streptococcus and Orussenella played major roles in HE group.(2)Effects of fecal bacteria from mice exercising at different intensities on depressive behavior of CRS mice(depressive behavioral results): compared with SC group,the central distance of mice in LR group and MR group in OFT was significantly increased(P <0.05),the sugar water preference of the mice in the HC group decreased significantly in SPT(P<0.05),and the immobility time of the mice in the HR group increased dramatically in the FST(P<0.05);compared with the SR group,the HR group The immobility time of the mice in the TST was significantly increased(P<0.05);compared with the LR group,the immobility time of the mice in the HC group was significantly increased(P<0.05).(3)Effects of fecal bacteria on serum tryptophan metabolites of CRS mice(ELISA results):Compared with SC group,IFN-γ concentration of mice in LR group(P<0.01)and MC group(P<0.05)was significantly decreased;Compared with SR group,5-HT concentration in LC group(P<0.001)and MC group(P<0.05)was significantly increased,TRP concentration in HR group was significantly increased(P<0.01)and KYNA concentration was significantly decreased(P<0.001).Compared with LC group,TRP concentration in HC group was significantly increased(P<0.01),and 5-HT concentration in HR group was significantly decreased(P<0.05).Compared with LR group,KYNA concentration of mice in HC group was significantly decreased(P<0.01),and KYN concentration of mice in HR group was significantly increased(P<0.05).(4)Effects of fecal bacteria on tryptophan metabolism related enzymes in the hippocampus of CRS mice(PCR and WES results): compared with SC group,the expression levels of TPH2 and KAT in the hippocampus of mice in HC group were significantly decreased(P<0.05).The m RNA level of IFN-γ in HC group was significantly increased(P<0.05),the m RNA expression level of KMO in HR group was significantly increased(P<0.05),and the protein expression level of KAT was significantly decreased(P<0.05).Compared with the LC group,the IFN-γ m RNA level in HC group was significantly increased(P<0.05),and the KAT m RNA expression level in HR group was significantly decreased(P<0.05).Compared with HR group,TPH2 protein level in LR group(P<0.01)and MR Group(P<0.05)was significantly increased.Conclusion:(1)Different intensity of exercise could cause different changes in intestinal flora composition and diversity.After CRS mice received FMT,the depression behavior of lowintensity and moderate-intensity exercise receptor mice was improved,but the depression behavior of high-intensity exercise receptor mice was aggravated.(2)Different intensities of exercise can mediate the intervention of gut microbiota from the level of different taxa and the 5-HT metabolism of tryptophan and the metabolism of kynurenine by different microorganisms.It is suggested that the possible mechanism of exercise improving depressionis to regulate the intestinal microbiota,increases the level of circulating and central 5-HT,reduces the level of inflammation in the body,regulates the metabolism of kynurenine and increases the expression of KAT. |
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