| ObjectiveChronic restraint stress(CRS)was used to establish the depression model with liver depression and spleen deficiency in C57BL/6 male rice.To investigate the effect as well as the mechanisms of Jiawei Xiaoyao Powder in depression mice,using the techniques and methods of behavior,pathology and intestinal microbiology.Methods1.The 60 C57BL/6 male mice were randomly divided into normal control group,model control group,Jiawei Xiaoyao Powder high dosage group,Jiawei Xiaoyao Powder medium dosage group,Jiawei Xiaoyao Powder low dosage group,and the sertraline group.There were 10 samples in each group.Chronic restraint stimulus(CRS)was used to make a mouse model of depression with liver depression and spleen deficiency.A self-made restraint tube was used to bind mice for 3 hours a day for 8 weeks.The occurrence and development of depression with liver depression and spleen deficiency were observed dynamically.The low,medium and high dose groups of Jiawei Xiaoyao Powder were given 1.0g/Kg,2.0g/Kg and 4.0g/Kg Jiawei Xiaoyao Powder aqueous solution respectively.In the sertraline group,each mouse was given sertraline 10mg/Kg daily;the model group given equal and the normal group were given the same amount of distilled water.For the above groups,gavage was performed once a day for 8 weeks.Relevant indicators were observed at the beginning,2 weeks,4 weeks,6 weeks and 8 weeks(at the end)of the experiment,which include mental state,activity,diet,stool and body weight of mice.Sucrose preference test and forced swimming experiments also were conducted at the same time.Feces were collected at the beginning of the experiment and every weekend.And put faces in sterile centrifuge tube,and then frozen in-80℃ refrigerator.After the modeling,all animals were tested and evaluated by tests such as locomotor activity,the elevated plus maze test,Y-maze test,the light-dark box test.2.Hematoxylin-eosin staining was used to observe the changes of neurons in hippocampus and amygdala of mice in each group.And the expression of microglia,which labeled by IBA-1,and astrocytes,which labeled by GFAP were determined using immunofluorescence was used to observe the changes of neurons in hippocampus and amygdala of mice in each group.And the expression of microglia,which labeled by IBA-1,and astrocytes,which labeled by GFAP were determined using immunofluorescence.3.PacBio SMRT sequencing was used to sequence the full-length amplifier of 16S rDNA in the feces of mice.To investigate the changes of gut microbiota as well as the mechanisms of Jiawei Xiaoyao Powder in depression mice,using the techniques and methods of behavior,pathology and intestinal microbiology.Results1.The body weight of mice under chronic restraint stress increased slowly,their food intake decreased significantly.What is more,CRS mice showed depressive like behavior.Compared with the normal group,the total motor distance and central motor distance of the model group decreased significantly,and model mice spent less time in center area.The rate of sucrose preference in SPT decreased significantly,while the immobility time in FST increased.In Y-maze test,the alternation rate of CRS mice was lower than that in the normal group.In the EMP and LBD,OT%,LBT%and LBD%were significantly reduced.Jiawei Xiaoyao Powder can improve weight gain and food intake of CRS mice,which showed significantly difference with the model group.And Jiawei Xiaoyao Powder can improve the behavior of depressive mice with liver depression and spleen deficiency syndrome.2.Jiawei Xiaoyao Powder regulated microglia and astrocytes in hippocampus of CRS mice.It reversed the over activation of microglia and the apoptosis of astrocytes.Compared with the normal control group,the hippocampal area of the model group showed a higher level of IBA-1 expression(p<0.05),and Jiawei Xiaoyao powder and sertraline could down-regulate the expression of IBA-1(p<0.05).The expression of GFAP(+)cells in model mice was lower than that in the normal group,and the number of astrocytes was increased after administration.3.Compared with the normal group,the CRS mice showed a significant change of gut microbiota.The relative abundance of harmful bacteria,such as Dubosiella,Turicibacter,increased;while the abundance of beneficial bacteria such as Paracteroidetes and Odoribacter reduced.The intervention of Jiawei Xiaoyao Powder reversed the imbalance of gut microbiota.Conclusions1.The C57 showed depression with liver depression and spleen deficiency after 8 weeks of chronic restraint stress.Jiawei Xiaoyao Powder can improve the body weight,food intake,general state and behavioral indexes of model mice,and regulate the symptoms of liver depression and spleen deficiency in depression.2.Jiawei Xiaoyao Powder can improve hippocampal damage,ameliorate the neuro-inflammation in hippocampus,and inhibit the apoptosis of neurons and astrocytes.3.The occurrence and development of depression with liver depression and spleen deficiency are related to the imbalance of intestinal microbial homeostasis.There were significant differences in intestinal microbial diversity and community structure between model and normal mice,with more harmful bacteria and less beneficial bacteria.Jiawei Xiaoyao Powder can achieve anti-depression by reversing the imbalance of gut microbiota. |
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