Study On The Effect Of Paeoniflorin In The Treatment Of Parkinson Disease Depression Based On NLRP3 Inflammasome Of Microglia

Parkinson Disease(PD)is a common chronic neurodegenerative disease in the elderly.Among them,depression is the most common non-motor symptom of PD.A large number of clinical studies have shown that Parkinson Disease is associated with depression(PDD).The incidence is over 50%.Paeoniflorin is extracted from the root of peony,and has anti-inflammatory,antioxidant and memory-enhancing effects.The role and mechanism of paeoniflorin in the development of depression in Parkinson's disease are unclear.Objective:This study elucidates paeoniflorin treats depression in Parkinson's disease by inhibiting microglial NLRP3 inflammasome mediated neuroinflammation.Methods:1.55 C57/b6J adult male mice were randomly divided into control group(Con,10 mice),1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP)subacute model group(MPTP,35 mice),and single peoniflorin group(Pae,10 mice).Control group will not be any stimulus,Pae group daily lavage paeoniflorin(30 mg/kg*7d),MPTP intraperitoneal injection of subacute model group MPTP(30 mg/kg*7d),after choose through forced swimming(FST)63.3%in depression mice,select continue to 10 mice gastric Pae(30 mg/kg*7d),the amount of normal saline as control,the last delivery after 1 days,by the sugar water preference(SPT),forced swimming(FST)and the tail suspension test(TST)detection in depression in mice behavior;The expression of synaptic proteins psd-95,synapsin-1 and GluRl in mPFC region and the expression of inflammasome subunits NLRP3,Caspase-1 and ASC in VTA region were detected by western-blotting.The morphological changes of dopaminergic neurons in VTA were observed by immunohistochemistry.Changes in levels of inflammatory cytokines IL-1?,TNF-? and IL-6 in the VTA region were detected by enzyme-linked immunosorbent assay kit(ELISA).2.The model of N2a cell Parkinson's disease was established with 1-methyl-4-phenylpyridine(MPP+).The experiment was divided into 4 groups:control group,model group(MPP+),model group+paeoniflorin group,and paeoniflorin group.CCK8 kit was used to test the survival rate of cells in each group of the experiment;BV-2 cells were used to prepare LPS+ATP/MPP++ATP inflammasome models.The experiment was divided into 4 groups:normal group,model group,model group+paeoniflorin group,Paeoniflorin group.CCK8 was used to detect microglial apoptosis,ELISA was used to detect changes in the contents of IL-1?,TNF-?,and IL-6 in cell supernatants,and Western-Blotting was used to determine changes in inflammatory corpuscle-related proteins in cell supernatants.Confocal microscope observation of morphological changes of microglia and the distribution of NLRP3 and Caspase-1.Results:1.In vivo experiments show that paeoniflorin can reverse the prolongation of FST and TST immobility in mice and decrease the sugar and water preference rate caused by MPTP.MPTP modeling and paeoniflorin administration had no significant effect on the behavior of mice in OFT.In addition,paeoniflorin can reverse the reduction of synaptic proteins PSD-95,synapsin-1,and GluR1 in mPFC region of MPTP-induced PDD model mice.It was further found that after MPTP modeling,DA neurons in the VTA region of mice were reduced,which was 78%of the control group.Microglial cells significantly proliferated and activated.Paeoniflorin could improve DA neuron damage in the VTA region of PDD mice.Inhibits excessive activation of microglia.Finally,the results of Western blotting showed that paeoniflorin could inhibit the expression of NLRP3,ASC,and Caspase-1 in the VTA region of PDD model mice.2.In vitro experiments showed that 10?M MPP+had no significant effect on N2a cells.50,100,250,500,1000,2000 ?M MPP+ concentration-dependently damaged N2a cells.When the MPP+concentration reached 250?M,the cell survival rate was 71.1%.Glycosides(1,5,25,50,100?M)can improve the survival rate of N2a cells.Under the LPS+ATP/MPP++ATP model,the survival rate of BV-2 cells was reduced,while paeoniflorin significantly inhibited the death of BV-2 cells caused by the activation of inflammatory bodies.It was further found that under the LPS+ATP/MPP++ATP model,BV-2 secreted IL-1?,TNF-? and IL-6 significantly increased.Paeoniflorin only inhibited the secretion of IL-1?.For TNF-? and IL-6 secretion has no effect.In addition,paeoniflorin inhibited the proliferation and activation of BV-2 cells induced by LPS+ ATP/MPP++ATP,and reduced the expression of NLRP3 and Caspase-1 in the cytoplasm.Finally,paeoniflorin can inhibit the activation of Caspase-1 and the maturation and secretion of IL-1? by BV-2 cells induced by LPS+ATP/MPP++ ATP.Conclusion:1.In PDD model mice,paeoniflorin protects DA neurons and exerts antidepressant effects by inhibiting microglial-mediated neuroinflammation in the VTA region.2.Paeoniflorin can inhibit the proliferation and activation of microglia by targeting the inhibition of NLRP3 inflammasome-mediated IL-1? secretion,and exert anti-inflammatory effects.