小胶质细胞p2y6受体激活与帕金森病发病相关性的分析-correlation between activation of p2y6 receptor in microglia and parkinsons disease.docxVIP

标记。关键词：P2Y6受体，小胶质细胞，帕金森病，IL-8IIResearchontherelationshipbetweenactivationofP2Y6receptoronmicrogliaandthepathogenesisofParkinsondiseaseAbstractObjective:TostudythemechanismonactivationofP2Y6receptoronmicrogliaanditseffectondopaminergicneurons.ToinvestigatewhetherP2Y6receptorcouldbeusedasadiagnosticbiomarkerforParkinsondiseasebydetectingIL-8levelsreleasedfromperipheralbloodmononuclearcellsofpatientswithParkinsondiseaseincaseofinflammation.Methods:MeasuretheexpressionlevelofP2Y6receptoronmicrogliaafterLPStreatmentaswellasthatofMIP-2mRNAandiNOS,COX-2proteininducedbyUDPandLPSthroughRT-PCRandwesternblot.ToconfirmtheinfluenceofP2Y6receptorontheapoptosisandvitalityofSH-SY5YcellsbywesternblotandMTTasssay.NotonlythephosphorylatedlevelofMAPKsignaltransductionpathwayafterincubationofBV-2cellswithLPSorUDPbutalsothechangeofNF-κBofBV-2cellsinducedbyUDPwastestedbyWesternblot.ExpressionlevelofiNOSandCOX-2proteinfollowingLPSstimulationafterinhibitionoftheMAPKsignaltransductionpathwaywasanalyzed,ThelevelsofIL-8inthesupernatantofperipheralbloodmomonuclearcells(PBMC)frompatientswithParkinsonsdiseaseandhealthycontrolsbeforeandafterLPSstimulationwasanalyzedbyELISAmethods.Results:LPScanupregulatetheexpressionofP2Y6onBV-2microglia.LPSalonebutnotUDPcaninducemicrogliatoreleaseMIP-2,iNOSandCOX-2,howeverUDPcandownregulatetheexpressionofiNOSinducedbyLPS.KnockingdownP2Y6receptorcanreducedtheapoptosisandtoxicityofSH-SY5Ycellsaddedbyconditionalmediumofmicroglia.P2Y6receptoractivatemicrogliapartiallydependedonactivationofERKandNF-κBpathway.ThePBMCcanexpressandreleasemoreIL-8afterLPSstimulationinIIIbothPDpatientsandhealthycontrols.ThePBMCofPDpatientscanexpressandreleasemoreIL-8beforeandafterLPSstimulationthancontrols,aswellastheincreasingrateofIL-8,althoughthesewasnotsignificantly.Conclusion:P2Y6receptorcanpartiallyparticipateintheactivationofmicrogliaandproductionofpro-inflammatoryfactorsinducedbyLPSthroughERKandNF-κBpathway,causingapoptosisanddopaminergicneurondeath.P2Y6receptormightplayanimportant