Study On The Mechanism Of Improving Depressive-like Behavior In CUMS Rats By Down-regulating The Level Of MKP-1 In The Hippocampus

BackgroundDepression,is one of a common mental disorder,seriously affects the quality of life of patients and their families,and brings a heavy burden to families and society.Studies have shown that mitogen-activated protein kinase phosphatase-1(MKP-1)is one of the molecules of depression and involved in the regulation of the inflammatory activation of microglia through activating mitogen-activated protein kinase(MAPK).Therefore,the down-regulation of MKP-1 may alleviate the depressive-like behaviors induced by chronic unpredictable mild stress(CUMS).This may be related to MKP-1 inhibit the expression proteins of MAPK signaling pathway and the inflammatory activation of microglia in the hippocampus.ObjectivesTo explore the expression levels of MKP-1,MAPK signaling pathway,inflammatory factors and apoptotic factors in the hippocampus of CUMS rats,in which MKP-1 was downgraded.Further more,to explore the molecular mechanisms of MKP-1 in regulating the pathogenesis of depression,and provide new options for the treatment of depression.Methods1.Group: In experiment one,30 male SD rats(180-220g)were randomly divided into control group and CUMS group.In experiment two,31 male SD rats(180-220g)were randomly divided into one virus control group and another MKP-1 down-regulated group.2.Behaviors before stress was evaluated after 1 week acclimatized.Subsequently,the control group was maintained feeding and handled for 3 to 5 minutes per day;the CUMS group was stressed for 6 weeks.The virus control group and the MKP-1down-regulated group recovered for 1 week after the virus injection,followed by CUMS stress for 6 weeks.3.CUMS stress: 2-3 kinds of stimuli were randomly given to rats every day for 6 weeks,and the CUMS stress were raised alone.4.NC-sh RNA adeno-associated virus were injected to the virus control group,while Dusp1-sh RNA adeno-associated virus were injected to the MKP-1 down-regulated group,to the CA1 and CA3 areas of hippocampus.5.Sucrose preference test,forced swimming test and open field test were used to evaluated depressive-like behavioral.6.Western blot was used to detect the relative expression levels of MKP-1,extracellular signal-regulated kinase(ERK),p-ERK,p38,P-p38,c-Jun N-terminal kinases(JNK),P-JNK,IL-6,IL-1β,TNF-ɑ,B-cell lymphoma-2(Bcl-2)and B-cell lymphoma-2(Bax)in the hippocampus of rats.7.Real-time PCR was used to detect the expression levels of MKP-1 m RNA,IL-6m RNA,IL-1β m RNA,and TNF-ɑ m RNA in the hippocampus of each group of rats.8.Immunofluorescence was used to detect the expression of Iba-1 in the hippocampus of rats.9.All data was analyzed with SPSS 22.0 software.The results were presented as mean±standard deviation.Repeated measures analysis of variance was used for the weight and behavioral test results of rats.The results of protein and PCR levels were analyzed by independent sample t test.Results1.Weight Change Compared with the control group,the CUMS group gained weight slower stress.There was no significant difference in body weight between the virus control group and the MKP-1 down-regulated group during the CUMS stress.2.Behavioral assessment results Compared with the control group after stress,the sucrose preference rate in the CUMS group decreased,the immobile time of forced swimming increased,and the number of defecation particles in the open field experiment decreased.The MKP-1 down-regulation group had a higher sucrose preference rate,and reduced forced swimming immobile time compared with the virus control group.In the open field text,rats spend time percent increased in the central area,and the number of uprights increased.3.MKP-1,P-ERK,ERK,P-p38,p38,P-JNK,JNK,IL-6,IL-1β,TNF-ɑ relative expression levels of proteins in the hippocampus Compared with the control group,the expression level of MKP-1、IL-6、IL-1β、TNF-ɑand Bax/Bcl-2 all increased,P-ERK/ERK and P-p38/p38 decreased,while P-JNK/JNK did not change significantly in the CUMS group.The relative expression level of MKP-1 、 IL-6 、 IL-1β and Bax/Bcl-2 decreased,P-ERK/ERK、P-JNK/JNK and TNF-ɑ increased,while P-p38/p38 changed not significant in the MKP-1 down-regulated group compared with the virus control group.4.Relative expression levels of MKP-1 m RNA,IL-6 m RNA,IL-1βm RNA,and TNF-ɑm RNA in the hippocampus Compared with the control group,the CUMS group expression level of MKP-1m RNA,IL-6 m RNA,IL-1β m RNA and TNF-ɑ m RNA increased in the hippocampus. Compared with the virus control group,MKP-1 down-regulated group MKP-1m RNA,IL-6 m RNA and IL-1β m RNA relative expression level decreased,but the expression level of TNF-ɑ m RNA had no statistical difference. 5.The expression of Iba-1 in the hippocampus Compared with the control group,the number of microglia in the CA1 area of the hippocampus in the CUMS group was increased,suggesting that the activity of microglia in the CA1 of hippocampus can be activated after chronic stress.Compared with the virus control group,the number of microglia in the CA1 area of the MKP-1 down-regulation group was reduced,indicating that the interference of MKP-1 could alleviate the activation of microglia in the hippocampus by CUMS.Conclusion1.MKP-1 activated microglia and induced apoptosis of hippocampus neurons through inhibiting the MAPK pathway proteins.2.The down-regulation of MKP-1 alleviated the depressive-like behaviors of CUMS rats,which could through up-regulating MAPK signaling pathway and down-regulating the inflammatory activation of microglia.3.The results of this study provide preclinical evidence for the treatment of depression by targeting key molecules that regulate the MKP-1 and MAPKs signaling pathways.