Study Of Regulated Hippocampal MKP-1-induced Depressive Behavior And Mechanism In Rats

BackgroundDepression disorder has become an important global public health problem.Previous studies have shown that there existed microglia inflammatory activation in depression and was closely related to mitogen-activated protein kinases(MAPKs)signaling pathway regulated by mitogen activated protein kinase phosphatase-1(MKP-1).We used SD male rats as research subjects with which depression model and MPK-1 over-expression model were created through CUMS modeling methods and adeno-associated virus transfection technology.MKP-1 expression and inflammatory activation of microglia were examined,and the mechanism of regulated MKP-1 expression on microglia inflammatory activation in depression was explored.ObjectivesTo explore the relationship between MAPK family proteins with microglia inflammatory activation under the influence of regulated hippocampal MKP-1 activity,and to investigate the mechanism of MKP-1 and MAPK signaling pathway in promoting the microglial cell releasing inflammatory cytokines.We aimed to illuminate the preliminary molecular mechanism of microglia inflammatory activation mediated by MKP-1 through MAPK signaling pathway in the onset of depression,and to provide supportive evidence for neuro-immunological hypothesis in depression.Methods1.Laboratory animal80 sprague-dawley rats(male,SPF grade)were accommodated in certified animal laboratory for one week,then were divided into a normal group,CUMS group,controlgroup and over-expression group(MKP-1)based on the results of initial behavioral assessments.2.Cerebral stereotaxic injectionAfter behavioral assessment,the rats in control group and MKP-1 group underwent ventral hippocampal micro-injection at stereotaxic apparatus.The control group and MKP-1 group did not undergo the CUMS modeling processes.3.CUMS modelingThe CUMS group received 5 weeks of modeling process.Statistically,single kind of stimulation was administrated on daily basis,without any repetition within a week,thus,the rats could not predict the next stimulus.4.Behavioral assessment: open field test,sucrose preference test and forced swimming were used as behavioral evaluation indexes for all animals.5.Expression of hippocampal MKP-1,FLAG,p-ERK,ERK,p-JNK,JNK,p-p38 and p38 were detected with Immunoblotting(Western blotting)in four groups.6.Expression of hippocampal calcium binding protein-1(ionized calcium binding adaptor molecule 1,Iba1)in each group was detected by immunohistochemistry.7.Inflammatory cytokines,such as IL-6,IL-1?,TNF-? in hippocampal cerebral tissues in each group were detected by fluorescence quantitative PCR(Quantitative RTPCR).8.Statistical methodThe measurement data were demonstrated as mean±standard deviation();Changes of behavioral data,microglia,and cytokine levels in each group were compared with one-way ANOVA;data comparison between two groups was conducted with two independent samples t test;With two-sided test,once P<0.05,significant differences were noticed.Results1.MKP-1 adeno-associated virus transfection(1)The hippocampal MKP-1 mRNA expression and protein in MKP-1 overexpression group were significantly higher than those in control group(P<0.05).(2)Hippocampal GFP-labeled fluorescent expression was found in both MKP-1 over expression group and control group.2.Behavioral assessmentCompared with normal group,the behavioral test results in over-expression group and CUMS group were both significantly different(P<0.05);compared with control group,the behavioral test results in over-expression group and CUMS group were both significantly different(P<0.05).3.Western blotting(1)MKP-1 protein: compared with normal group,expression of hippocampal MKP-1protein in CUMS group was significantly higher(P<0.05).(2)p-ERK/ERK protein: compared with normal group,expression of hippocampal p-ERK/ERK protein in CUMS group and over-expression group was significantly lower(P<0.05);Compared with control group,expression of hippocampal p-ERK/ERK protein in CUMS group and over-expression group was significantly lower(P<0.05).4.Morphological changes of microglia marker IBA-1The hippocampal microglia activation,such as increased cell body and decreased synapse,were observed in over-expression group and CUMS group,turning into amoeba-like morphology.5.Expression of inflammatory cytokinesCompared with normal group,expressions IL-1?,IL-6 and TNF-? mRNA in over-expression group and CUMS group were significantly higher(P<0.05);Compared with control group,expressions of IL-1? and IL-6 mRNA in over-expression group and CUMS group were significantly higher(P<0.05);compared with CUMS group,expressions of IL-1?,IL-6,and TNF-? mRNA were significantly higher(P<0.05);TNF-? mRNA in control group was higher than that in normal group.Conclusions1.MKP-1 may be an important gene in pathogenesis of depression.2.MAPK-ERK signaling pathway may be related to pathogenesis of depression.3.In over-expression group and CUMS group,abnormal hippocampal microglia activation and inflammatory cytokines change were observed,which may be an important factor in depressive behavior of rats.