The Impacts Of Peroxisome Proliferator-activated Receptors On Depression

Depression is the most common mental disease worldwidecaused by emotional disorder,and also the largest cause of disability worldwide.According to the latest epidemiological statistical analysis,the number of people suffering from depression is increasing year by year,and there are 800,000 people committing suicide due to depression every year,which brings huge burden to the development of today's society.Although there are many antidepressants available,but drugs commonly used in the treatment of depression are based on the regulation of monoamine neurotransmitters in clinic,such as selective serotonin(5-hydroxytryptamine,5-HT)reuptake inhibitors and 5-HT receptor transport inhibitors.Although these drugs have been widely used in clinic for more than 60 years,most of them havetookalong time to work and failed to treat 30% of patientswith depression.Therefore,it is urgent to seek forthesafe and effective antidepressants.It has been reported that patients with depression show the characteristics of low glucose metabolism in the prefrontal limbic system,which are more likely to suffer from metabolic syndrome than normal people,while people with metabolic syndrome have a higher risk of depression.Therefore,the concept of“depression is ? metabolic syndrome”wasput forward.Studies have shown that there are many common pathological features and pathogenesis between depression and metabolic syndrome,such as chronic inflammation.In the central nervous system(CNS),the activation of microglia,astrocytes or other myeloid cells can induce neuroinflammation,thereby promoting the release of inflammatory cytokines,chemokines and other inflammatory mediators,and ultimately aggravating the nerve damage.Although the general M1 and M2 classificationof microglia have been controversial,microglia with different activation states still have corresponding functional manifestations.Therefore,the study of microglia M1/M2 transformation under the pathological changes of depression is helpful to further explore the molecular mechanism of depressionand explore novel therapeutic strategies.Peroxisome proliferator-activated receptors(PPARs)are ligand-activated transcription factors which belong to the nuclear receptor superfamily,whichare involved in sugar and lipid metabolism,cell differentiation and inflammatory response.More and more literatures show that PPARs play an important neuroprotective role in a variety of neurological diseases.For example,PPAR?/?dualantagonists inhibited the inflammatory response in the brain of mice,thereby alleviating thebrain injury induced by ischemia reperfusion;PPAR?/? agonist inhibitedmicroglial activation in the brain of experimental autoimmune encephalomyelitis mice,reduced pathological damage and mortality.These findings suggest that PPARs may also play an important role in the pathological process of depression.It has been reported that rosiglitazone,a PPAR? agonist,improvedthedepression-like behavior in mice induced by chronic mild unpredictable stress(CMS),butthe mechanism of action is unclear.Besides,PPAR?/? is themost expressed receptor subtype in the brain,but the mechanism of which in the brain is rarely reported.Therefore,the antidepressant effect and its mechanism need further research and explore,so as toprovide a novel strategy for the development of new anti-depressants based on "non-monoamines".Based onourprevious studies,the first part of this paper is to study the protective effect and molecular mechanism of PPAR?/? on astrocytes in the pathological process of depression.Wild-type C57BL/6 mouse were used to establish the depression model induced by CMS,and the methods ofclassical behavior,immunohistochemistry and transmission electron microscopewere used to demonstrated that PPAR?/?activation could improve depression-like symptomsinduced by CMS in mouse,reduce the injury of hippocampal neurons and astrocytes,and significantly inhibit the damage of endoplasmic reticulum of hippocampal astrocytes.In order to further elucidate the protective effect and molecular mechanism of activated PPAR?/?,corticosterone was used to stimulate primary cultured astrocytes to establish thestress model in vitro,and astrocytes were treated with PPAR?/?agonist GW0742.The second part of this paper mainly explores the regulatory effect and molecular mechanism of PPAR? on microglial phenotype transformation during the pathological process of depression.In this part,lipopolysaccharid(LPS)was used to stimulate primary microglia cells to establish anneuroinflammatory modelin vitro,which was used toobserve the effect of PPAR?on microglial phenotypic transformation.Part ? The impacts and the involved mechanisms of PPAR?/? activation on chronic mild unpredictable stress-induced depressive miceAIM:To investigate and clarify thetherapeutic effectsand the involved mechanisms of PPAR?/? activation on CMS-induced depressive mice.This research provided thetheoretical basis for the exploration of new strategies for the treatment of depression and the development of new effective antidepressants,and broadened the pharmacological effect of PPAR?/?.METHODS: 1.Wild-type C57BL/6 mouse were used to establish the depression model by CMS;2.The behavior test was carried outbysucrose preference test(SPT),forced swimming test(FSF),open field test(OPT)and tail suspension test(TST).3.The number and morphology of neurons and astrocytes and the expression of related proteins were observed in hippocampal by Immunofluorescence(IF)assay.4.The structure and morphology of endoplasmic reticulum of astrocytes were observed by transmission electron microscope.5.Primary cultured astrocytes were obtained from mouse,and the purity of primary cultured astrocytes was identified by immunofluorescence staining combined with counting method.6.Corticosterone was used to stimulate the primary astrocytes,which was establishedchronic stress model in vitro.7.Methyl thiazolyl tetrazolium(MTT)test was used to observe the protective effect of PPAR?/? activationon corticosterone-induced astrocytic injury.8.Flow cytometry(FCM)was used to observe the effect of PAR?/? activation on corticosterone-induced astrocytic apoptosis.9.Western blotting(WB)was used to detect the expression of endoplasmic reticulum-related proteins and apoptosis-related proteins,which was observed the effect of PPAR?/? activationon corticosterone-induced endoplasmic reticulum stress and apoptosis in astrocytes.10.The effect of PPAR?/?activationon the binding of GRP78 and p-IRE1? was observed by co-immunoprecipitation(Co-IP).RESULTS:1.The expression of PPARs subtypes are down-regulated in the hippocampal of CMS-induced mice;2.PPAR?/?activation significantly improves CMS-induced the depression-like symptoms in mouse;3.There was no change in the shape and number of nissl bodies in the hippocampal of CMS-induced mice;4.PPAR?/?activation inhibits CMS-inducedthe astrocyticendoplasmic reticulum stress and injurys in hippocampal;5.PPAR?/?activation promotesastrocytic survival by inhibiting corticosterone-induce the endoplasmic reticulum stress in astrocytes;6.PPAR?/?activation promotes the binding of GRP78 to p-IRE1?in astrocytes,and inhibitsthe endoplasmic reticulum stress(ERs)-IRE1?-mediated apoptosis pathway of astrocytes.CONCLUSIONS:1.PPAR?/? activation improves CMS-induced depression-like symptoms in mouse,and exertsthe protective role of astrocyte in hippocamp;2.PPAR?/? activation exerts a protective effect on astrocytes by inhibitingthe endoplasmic reticulum stress(ERs)-IRE1?-mediated apoptosis pathway of astrocytes.Part ? Antagonizing PPAR? facilitates M1-to-M2 shift of microglia by enhancing autophagy via the LKB1-AMPK signaling pathwayAIM:To investigate and clarify the effectsand the involved mechanisms of antagonizing PPAR? facilitates the LPS-induced switch of microglial polarization from the M1 phenotype to the M2 phenotype.This research broadened the pharmacological role of PPAR?,which may provide a potent therapeutic strategy for related neuroinflammatory diseases.METHODS: 1.Primary cultured microglia cellswere obtained from rats,and the purity of primary cultured microglia cells was identified by immunofluorescence staining combined with counting method.2.Lipopolysaccharide(LPS)was used to stimulate microglia cells,which was used to establish neuroinflammatory modelin vitro;3.The microscope was used to observe the effects of antagonizing PPAR?on the ameba-like state of microglia induced by LPS,which was used to determine the effective concentration.4.The real-time quantitative polymerase chain reaction(RT-q PCR)was used to observethe effects of PPAR?agonist rosiglitazone and PPAR?antagonist T0070907 on the expression of M1 phenotype and M2 phenotype markers in microglia after LPS stimulation.5.Immunofluorescence co-localization was used to observethe effects of PPAR?antagonist T0070907 on the expression of M1 phenotype and M2 phenotype markers in microglia after LPS stimulation.6.Flow cytometry(FCM)was used to observethe effects of PPAR?antagonist T0070907 on the expression of M1 phenotype and M2 phenotype markers in microglia after LPS stimulation.7.Western blotting(WB)was used to detect the expression of autophagy related proteins and autophagy upstream related proteins,which was observedthe effect of PPAR?antagonist T0070907 on the autophagy in microglia after LPS stimulation.8.The electron microscope was used to observethe effect of PPAR?antagonist T0070907 on the autophagy in microglia after LPS stimulation.9.Primary microglia cells were transfected with mcherry-EGFP-LC3 plasmid,which was observed to the effect of PPAR antagonist on the autophagy in microglia after LPS stimulation by laser confocal scanning.10.The phagocytosis experiment was used to observe the effect of PPAR antagonist on the autophagy in microglia after LPS stimulation.11.Co-immunoprecipitation(Co-ip)was used to observe the effect of PPAR antagonists on the transcriptional activation of LKB1.12.Primary microglial cells and BV2 cells were transfected with si RNA to determine the related regulatory effect of PPAR?.RESULTS:1.Activating PPAR?by rosiglitazone fails to promote LPS-induced the switch of microglial polarization from the M1 phenotype to the M2 phenotype.2.Antagonizing PPAR? by T0070907 facilitats the LPS-induced the switch of microglial polarization from the M1 phenotype to the M2 phenotype.3.Antagonizing PPAR? by T0070907 reverses LPS-induced the inhibition of autophagy in primary microglia cells.4.The effects of antagonizing PPAR?on the autophagy in primary microglia,which depended on the activation of LKB1-AMPK signaling pathway.5.Antagonizing PPAR?facilitates the LPS-induced the switch of microglial polarization from the M1 phenotype to the M2 phenotypeby activating LKB1;6.PPAR?knockdown in microglia cells activates the LKB1-AMPK signaling pathway and inhibites LPS-induced microglial M1 polarization.CONCLUSIONS:1.Antagonizing PPAR? by T0070907 facilitates the LPS-induced the switch of microglial polarization from the M1 phenotype to the M2 phenotype;2.Antagonizing PPAR? by T0070907 facilitates the LPS-induced the switch of microglial polarization from the M1 phenotype to the M2 phenotypeby increasingthe intracellular autophagy.3.Antagonizing PPAR? by T0070907 reverses the LPS-induced the inhibition of intracellular autophagy by activating LKB1-AMPK signaling pathway.The major contributions of the present study lie in:1.To confirm the antidepressant effectof PPAR?/? activation.It is further elucidated that PPAR?/? activation promotes the protective effect on astrocytesby inhibiting the endoplasmic reticulum stress(ERs)-IRE1?-mediated apoptosis pathway.It provides the new idea and experimental basis for the development of new antidepressant drugs based on "non-monoamine".2.It is found that antagonizing PPAR?facilitated the LPS-induced the switch of microglial polarization from the M1 phenotype to the M2 phenotype.It was further revealed that antagonistic antagonizing PPAR? by T0070907 reversed the LPS-induced the inhibition of intracellular autophagy by activating LKB1-AMPK signaling pathway.It provides the important theoretical basis for the anti-inflammatory treatment and clinical application of anti-depression.