固有免疫研究进展课件

1 1固有免疫研究进展固有免疫研究进展固有免疫研究固有免疫研究进进展展2 2Contents1、Innateimmunesystem2、Innateimmunerecognition3、Innateimmuneresponse4、ProgressofinnateimmunityContents1、Innate immune system3 3固有免疫研究固有免疫研究进进展展课课件件4 4固有免疫研究固有免疫研究进进展展课课件件5 5InnateImmunitySystem一、固有免疫系统一、固有免疫系统Innate Immunity System6 6宿主固有免疫系统宿主固有免疫系统固有免疫系统固有免疫系统固有屏障结构固有屏障结构皮肤与粘膜皮肤与粘膜血脑屏障血脑屏障胎盘屏障胎盘屏障固有免疫分子固有免疫分子补体补体溶菌酶溶菌酶防御素防御素固有免疫细胞固有免疫细胞PMNDCsMf f宿主固有免疫系宿主固有免疫系统统固有免疫系固有免疫系统统固有屏障固有屏障结结构皮肤与粘膜血构皮肤与粘膜血脑脑屏障胎屏障胎7 7（一）固有屏障结构（一）固有屏障结构（一）固有屏障（一）固有屏障结结构构8 8Figure2-4Intactskin/MucusmembraneMucosalimmunesystem(MIS)Secretionofanti-microbialfactorsNormalfloraBlood-brainandplacentabarriersAnatomicandPhysiologicBarriersFigure 2-4Intact skin/Mucus me9 9BarriersofinnateimmunitysystemlSkinandmucosalbarrierlPhysical:skin&mucosa,epidermis-dermisconjunction,mucosalsecretion,lavagingoffluidsandciliaryactionlChemical:acidicsecretion,lysozymelMicroorganism:commensalbacteriaBarriers of innate immunity sy1010固有免疫研究固有免疫研究进进展展课课件件1111LocalbarrierstructurelBlood-thymus-barrierlBlood-gas-barrierlBlood-urine-barrierlBlood-testis-barrierlBlood-epididymis-barrierlCerebral barrier:BBB,etc.lBlood-fetus-barrierlMucosa-bicarbonatebarrierLocal barrier structureBlood-t1212（二）固有免疫细胞（二）固有免疫细胞（二）固有免疫（二）固有免疫细细胞胞1313ComponentsoftheImmuneSystemComponents of the Immune Syste1414固有免疫细胞固有免疫细胞固有免疫固有免疫细细胞胞1515固有免疫研究固有免疫研究进进展展课课件件1616固有免疫研究固有免疫研究进进展展课课件件1717PhagocytosisTypeofphagocytes:Smallphagocytes(neutrophils)largephagocytes(macrophage)DendriticCellsChemotaxisContactPhagocytosisIntracellularkillingPhagocyticeffectsenhancedbyantibodyorcomplementPhagocytosisType of phagocytes1818吞噬和杀菌过程：吞噬和杀菌过程：趋化趋化黏附黏附 吞入吞入 杀灭与消化杀灭与消化(CD14)吞噬和吞噬和杀杀菌菌过过程：程：(CD14)1919Real-timemovieofphagocytosisReal-time movie of phagocytosi2020（三）固有免疫分子（三）固有免疫分子（三）固有免疫分子（三）固有免疫分子2121固有免疫分子固有免疫分子固有免疫分子固有免疫分子2222固有免疫分子固有免疫分子1.Complment:alternative&MBLpathway2.Lysozyme:tissueandbodyfluid,secre-tion,lysosomeinphagocytes3.Antimicrobial peptides:defensin4.Cytokines:interferon,IL-1,IL-6,IL-12,TNF-,chemotaxin5.Acute phase protein:CRP,MBP,SAP,1-acidglycoprotein,FN,etc.固有免疫分子固有免疫分子Complment:alternative&2323Lysozyme1.KillingG+bacteria,splitstheexposedpeptidoglycanwall2.Category:3.Bacterial lysozyme4.Acton-1,4glycosidicbond5.Actonthepeptidetailandamide6.Fungal lysozyme:Saccharomycetic,AspergillarLysozymeKilling G+bacteria,s2424Cytokines1.Interferon:virallyinfectedcellscansynthesizetypeIFN.ReducemRNAtranslation,degrademRNA,inhibitviralreplication2.Interleukin:IL-1,IL-6,IL-123.Chemotacticfactors4.TNF:TNF-CytokinesInterferon:virally i2525InnateRecognition二、固有免疫识别二、固有免疫识别Innate Recognition2626FatherofInnateImmunity：Prof.JanewayCConcept:PRR:PathogenrecognizingreceptorsPAMPs:PathogenassociatedmolecularpatternsFather of Innate Immunity：Prof2727（一）识别什么？（一）识别什么？PAMPs（一）（一）识别识别什么？什么？PAMPs2828PAMPandPRR1、PAMP:PathogenAssociatedMolecularPatternsLippolysaccharide(LPS),dsRNA,unmethylatedCpG,mannose,glucan,Lipoteichoicacid(LTA),Pepidoglycan(PGN)2、PRR:receptorsonprofessionalAPCandMwhichenablethemtorecognizePAMP.lmannosereceptorlToll-likereceptorlscavengerreceptorlphosphatidylserinereceptorPAMP and PRR1、PAMP:Pathogen A292929PathogenAssociatedMolecularPatternlPAMPareconserved,sharedbyalargegroupofinfectiousagents,andclearlydistinguishablefromselfpatterns.lTypicalPAMP:lipopolysaccharide(LPS),mannose,glucan,dsRNA,lipoteichoicacid(LTA),peptidoglycan(PGN)unmethylatedCpGDNAetc29Pathogen Associated Molecula3030Pathogen-AssociatedMolecularPatterns(PAMPs)Pathogen-Associated Molecular 3131黏附素（黏附素（adhesin)adhesin)黏附素（黏附素（adhesin)3232fimbrial adhesins(E.coli)non-fimbral adhesin（Bartonella）黏附素黏附素fimbrial adhesins(E.coli)no3333固有免疫研究固有免疫研究进进展展课课件件3434LigandSpecificitiesofTLRsLigand Specificities of TLRs3535（二）用什么去识别？（二）用什么去识别？PRR（二）用什么去（二）用什么去识别识别？PRR36363636Pattern-RecognitionReceptor，PRRlExpressedmainlyonDCandMfwhichenablethemtorecognizePAMP.lThreetypesofPRR:secretedPRR:MBL,C-reactiveproteinendocyticPRR:lscavengerreceptor,lmannosereceptorsignalingPRR:Toll-likereceptor,NOD-likereceptor,RIG-likereceptor3636Pattern-Recognition Recept37373737一）一）TLR的发现的发现lDrosophilaembryogenesis.(1980s)lAntifungalresponseinDrosophilaadults.(Hoffmannet al.,Cell,1996)lAhumanhomologueofToll(Janewayet al.,Nature,1997)3737一）一）TLR的的发现发现Drosophila embryog38383838The2011NobelPrizeJulesA.HoffmannBruceBeutlerTheyreceivedone-halfofthe2011NobelPrizeinPhysiologyorMedicinethe2011NobelPrizeinPhysiologyorMedicine,fortheirdiscoveriesconcerningtheactivationofinnateimmunitytheactivationofinnateimmunity3838The 2011 Nobel Prize Jul3939Toll-likeReceptors(TLRs)Afamilyofproteins,whichareAfamilyofproteins,whicharestructurallyrelatedtoDrosophilastructurallyrelatedtoDrosophilaTollandIL-1receptor.TollandIL-1receptor.TLR1-10inhuman,TLR1-13inTLR1-10inhuman,TLR1-13inmouse.mouse.RecognizespecificpathogenRecognizespecificpathogenassociatedmolecularpatternsassociatedmolecularpatterns(PAMPsPAMPs)suchasendotoxin)suchasendotoxin(LPS).(LPS).RegulatetheactivationofbothRegulatetheactivationofbothinnateinnateandandadaptiveadaptiveimmunity.immunity.Toll-like Receptors(TLRs)A fa4040固有免疫研究固有免疫研究进进展展课课件件4141TLRsignaling AkiraS.etal.(2005)AkiraS.etal.(2005)Int.Immunol.Int.Immunol.17:1-1417:1-14TLR signaling Akira S.et al.4242固有免疫研究固有免疫研究进进展展课课件件434343434444TLRsininnateandadaptiveimmunityTLRs in innate and adaptive im4545TLRscontroladaptiveimmunityAkiraS.etal.AkiraS.etal.(2001)(2001)Nature ImmunologyNature Immunology2:675-6802:675-680 TLRs control adaptive immunity4646固有免疫研究固有免疫研究进进展展课课件件4747固有免疫研究固有免疫研究进进展展课课件件4848参与固有免疫识别的受体参与固有免疫识别的受体模式识别受体（模式识别受体（PRR)PRR)参与固有免疫参与固有免疫识别识别的受体的受体4949抗感染免疫与固有免疫识别抗感染免疫与固有免疫识别抗感染免疫与固有免疫抗感染免疫与固有免疫识别识别5050NOD-likereceptors,NLRlRecognizecytoplasmicPAMPsand/orendogenousdangersignals,inducingimmuneresponses.lLigands:MDPandmeso-DAPlThreedomains:C-terminalLRRssenseligandscentrallylocatedNODdomainN-terminaleffectorregioncomprisingaprotein-proteininteractiondomainsuchastheCARD,PyrinorBIRdomain.NOD-like receptors,NLRRecogni5151NLRsubfamilieslSeveralsubfamilies(onthebasisoftheireffectordomains)CARDsubfamily:NODs,IPAF,CIITA,PYDsubfamily:NALPsBIRsubfamily:NAIPsNLR subfamiliesSeveral subfami525252NLRDomains52NLR Domains535353NLRfamilies53NLR families5454FunctionsofNLRlTheymaythenformoligomersthatactivateinflammatorycaspase1causingcleavageandactivationofimportantinflammatorycytokines（IL-1）lNLRsmayalsoactivatetheNF-kBsignalingpathwaytoinduceproductionofinflammatorymolecules.Functions of NLRThey may then 55555555NLRsignaling5555NLR signaling 5656NLRandinflammasome inflammasome,amultiproteincomplexthatcontains,inadditiontoNLR,theadaptormoleculeASCandtheproteasecaspase-1.inflammasomeisresponsiblefortheproteolyticprocessingoftheimmatureformofthecytokinesbelongingtotheIL-1 family(IL-1b,IL-18,andIL-33),56NLR and inflammasome inflammas5757IL-1/IL-18inflammosome57IL-1/IL-18 inflammosome575858NALP3andIPAFinflammasomeFabio Martinon，Annu.Rev.Immunol.2009.27:229NALP3 and IPAF inflammasomeFab5959RIG-Ilikereceptors，RLRlDetectintracellulardsRNAofviralorigin:RIG-I(retinoicacidinducedgene)MDA-5(melanomadifferentiationassociatedgene)LGP2(laboratoryofgeneticsandphysiology)lComprisesN-terminalcaspaseactivationandrecruitmentdomains(CARDs),aDECHhelicase,andaC-terminaldomain(CTD).RIG-I like receptors，RLRDetect606060RLRDomainsRedbox,CARD1;bluebox,CARD2;greenbox,Helicasedomain;purplebox,DEAD/DEAHdomain60RLR DomainsRed box,CARD1;616161SignalingofRLR61Signaling of RLR6262e.g.,HCVproteinNS3/4Aeliminatesantivirale.g.,HCVproteinNS3/4AeliminatesantiviralsignalingbycleavageofMAVSandTRIF.signalingbycleavageofMAVSandTRIF.e.g.,HCV protein NS3/4A elimin636363SummaryofPatternRecognitionThe Journal of Clinical Investigation,118(2),200863Summary of Pattern Recogniti6464InnateImmuneResponse固有免疫应答固有免疫应答Innate Immune Response6565固有免疫研究固有免疫研究进进展展课课件件6666固有免疫研究固有免疫研究进进展展课课件件6767固有免疫研究固有免疫研究进进展展课课件件6868固有免疫研究固有免疫研究进进展展课课件件696969697070固有免疫研究固有免疫研究进进展展课课件件7171固有免疫研究固有免疫研究进进展展课课件件7272视频视频(ImmuneResponse)视频视频 7373固有免疫研究固有免疫研究进进展展课课件件7474固有免疫研究固有免疫研究进进展展课课件件7575固有免疫研究固有免疫研究进进展展课课件件7676固有免疫研究固有免疫研究进进展展课课件件7777IntracellularkillingmechanismsO2-dependent:ROI,RNI(reactivenitrogenintermediate),SOD(superoxidasedismutase)O2independent:MPO(myeloperoxidase,)Enzymes(lysozyme)DefensinsIntracellular killing mechanis7878ProgressofInnateImmunity固有免疫研究进展固有免疫研究进展Progress of Innate Immunity7979抗细菌感染免疫的抗细菌感染免疫的固有免疫识别与免疫应答固有免疫识别与免疫应答抗抗细细菌感染免疫的菌感染免疫的8080抗病毒免疫的抗病毒免疫的固有免疫识别与免疫应答固有免疫识别与免疫应答抗病毒免疫的抗病毒免疫的8181P.aeruginosakeratitisNormalcorneaBacterialKeratitis:ClinicalPictureP.aeruginosa keratitisNormal 8282研究依据研究依据WHOWHO（20062006）:在影响身心健康的疾病中，在影响身心健康的疾病中，眼科疾病仅次于肿瘤与心血管疾病眼科疾病仅次于肿瘤与心血管疾病角膜病主要的致盲性眼病角膜病主要的致盲性眼病 中国角膜盲患者中国角膜盲患者350350500500万万 绝大部分与角膜溃疡相关绝大部分与角膜溃疡相关广东省是角膜溃疡的高发区广东省是角膜溃疡的高发区研究依据研究依据WHO（2006）:在影响身心健康的疾病中，眼科疾病在影响身心健康的疾病中，眼科疾病8383立项依据立项依据角膜感染角膜感染 溃疡穿孔溃疡穿孔致盲致盲感染性角膜溃疡：感染性角膜溃疡：青壮年劳动人口、绿脓杆菌青壮年劳动人口、绿脓杆菌青壮年劳动人口、绿脓杆菌青壮年劳动人口、绿脓杆菌病理本质：免疫病理损害病理本质：免疫病理损害溃疡坏死溃疡坏死临床治疗：抗感染临床治疗：抗感染耐药性、过度免疫反应耐药性、过度免疫反应耐药性、过度免疫反应耐药性、过度免疫反应 溃疡穿孔溃疡穿孔溃疡穿孔溃疡穿孔致盲致盲致盲致盲立立项项依据角膜感染依据角膜感染 溃疡溃疡穿孔穿孔致盲致盲8484免疫病理损害机制免疫病理损害机制激活机体的激活机体的抗感染免疫抗感染免疫 清除病原体清除病原体病原体感染病原体感染宿主免疫宿主免疫激活激活PRRPRR受体受体 过度炎症反应过度炎症反应过度炎症反应过度炎症反应 免疫病理损害免疫病理损害免疫病理损害免疫病理损害 LPSLPSLPSLPS、DNADNADNADNA、鞭毛素鞭毛素鞭毛素鞭毛素 负调控负调控负调控负调控免疫病理免疫病理损损害机制激活机体的抗感染免疫病原体感染宿主免疫害机制激活机体的抗感染免疫病原体感染宿主免疫 8585 热热 红红 肿肿 痛痛 功能丧失功能丧失炎症炎症 热热 红红 肿肿 痛痛 功能功能丧丧失失 炎症炎症8686BacteriaPseudomonas aeruginosaPseudomonas aeruginosaATCCstrain19660ATCCstrain196601x101x106 6CFU/mlappliedtoscarifiedcorneaCFU/mlappliedtoscarifiedcorneaBacteriaPseudomonas aeruginosa8787AnimalmodelofP.aeruginosainfectionC57BL/6(B6)miceSusceptible(cornealperforationwithin5-7daysp.i.)Susceptible(cornealperforationwithin5-7daysp.i.)BALB/cmiceResistant(restorecornealclaritywithin1-2wkp.i.)Resistant(restorecornealclaritywithin1-2wkp.i.)Animal model of P.aeruginosa 8888MicroarrayanalysisWhatistheoverallgeneexpressionprofileinB6vs.WhatistheoverallgeneexpressionprofileinB6vs.BALB/cmiceafterBALB/cmiceafterP.aeruginosaP.aeruginosacornealinfection?cornealinfection?B6andBALB/cmicewereinfectedwithB6andBALB/cmicewereinfectedwithP.aeruginosaP.aeruginosa Uninfected(N)corneastakenandat1dayp.i.Uninfected(N)corneastakenandat1dayp.i.TotalRNAextractedforlabelingTotalRNAextractedforlabelingRNAhybridizedwithamurinearrayMG-U74(Affymetrix)RNAhybridizedwithamurinearrayMG-U74(Affymetrix)ArraychipsscannedforrawdataArraychipsscannedforrawdataDataanalyzedusingbioinformaticssoftwarepackageDataanalyzedusingbioinformaticssoftwarepackageHuangX.andHazlettLD.HuangX.andHazlettLD.(2003)(2003)IOVS IOVS 44:3409-341644:3409-3416Microarray analysisB6 and BAL8989Type-1(Red)vs.Type-2(Green)B61dpiB61dpiCC CEE EDD DFF FB6NormalB6NormalB6NormalB6NormalBALB/cNormalBALB/cNormalBALB/cNormalBALB/cNormalB61dpiB61dpiB61dpiB61dpiBALB/c1dpiBALB/c1dpiBALB/c1dpiBALB/c1dpiBB BBALB/cNormalBALB/cNormalBALB/c1dpiBALB/c1dpiAA AB6NormalB6NormalBlue=DetectableBlue=DetectableRedRed=type-1genes=type-1genesGreenGreen=type-2genes=type-2genesHuangX.andHazlettLD.HuangX.andHazlettLD.(2003)(2003)IOVS IOVS 44:3409-341644:3409-3416Type-1(Red)vs.Type-2(Green9090GeneexpressiondifferencesconfirmedbyPCRHuangX.andHazlettLD.(2003)IOVS44:3409-3416Gene expression differences co9191RNAexpressionofcellulargenesup-(ordown-)RNAexpressionofcellulargenesup-(ordown-)regulated4-foldorhigherinbothmousestrainsregulated4-foldorhigherinbothmousestrainsGeneNameGeneNameAccessionNo.AccessionNo.TLRs/Signaltransduction:TLRs/Signaltransduction:1.CD141.CD14X13333X133332.IL-1R2.IL-1RL32838L328383.TLR-63.TLR-6AB020808AB0208084.IKKi4.IKKiAB016589AB0165895.NIK5.NIKU88984U889846.Fos-likeantigen16.Fos-likeantigen1AF017128AF0171287.Fasantigen7.FasantigenM83649M836498.TNFReceptor18.TNFReceptor1X87128X871289.TRAF-19.TRAF-1L35302L35302HuangX.andHazlettLD.HuangX.andHazlettLD.(2003)(2003)IOVS IOVS 44:3409-341644:3409-3416RNA expression of cellular gen9292TLR4inbacterialkeratitisP.aeruginosa P.aeruginosa LPSLPS,aspecificligandofTLR4.aspecificligandofTLR4.AkiraS.etal.(2005)AkiraS.etal.(2005)Int.Immunol.Int.Immunol.17:1-1417:1-14TLR4 in bacterial keratitisP.9393DayspostinfectionDayspostinfectionN N113355770 0303060609090120120150150RelativemRNAofTLR4RelativemRNAofTLR4TLR4mRNAexpressioninBALB/cmiceHuangX.etal.(2006)HuangX.etal.(2006)IOVS IOVS 47:4910-491647:4910-4916Days postinfectionN 1 3 5 703094940 01 12 23 34 45 5BALB/cBALB/cTLR4TLR4lps-dlps-d1d1d3d3d5d5d DayspostinfectionDayspostinfectionClinicalscoreClinicalscoreDiseaseresponseafterP.aeruginosainfectionA AB BC CD DHuangX.HuangX.etal.etal.(2006)(2006)IOVS IOVS 47:4910-491647:4910-4916012345BALB/cTLR4lps-d1d3d5d Da9595DayspostinfectionDayspostinfectionLogLog1010CFU/corneaCFU/cornea3d3d5d5d0 02 24 46 68 810101212BALB/cBALB/cTLR4TLR4lps-dlps-dp0.001p0.001p0.001p0.001DayspostinfectionDayspostinfectionUnitsMPO/corneaUnitsMPO/cornea3d3d5d5d0 06060120120180180240240300300BALB/cBALB/cTLR4TLR4lps-dlps-dp0.01p0.01p0.001p0.001HuangX.etal.HuangX.etal.(2006)(2006)IOVS IOVS 47:4910-491647:4910-4916Bacterialload&MPOassayDays postinfectionLog10 CFU/co9696Pro-inflammatorycytokineproductionHuangX.etal.(2006)HuangX.etal.(2006)IOVS IOVS 47:4910-491647:4910-4916DayspostinfectionDayspostinfectionDayspostinfectionDayspostinfectionIL-1IL-1b b b bprotein(pg/ml)protein(pg/ml)3d3d5d5d0 030003000600060009000900012000120001500015000BALB/cBALB/cTLR4TLR4lps-dlps-dp0.01p0.01p0.001p0.001DayspostinfectionDayspostinfectionMIP-2protein(pg/ml)MIP-2protein(pg/ml)3d5d030006000900012000BALB/cBALB/cTLR4TLR4lps-dlps-dp0.001p0.001p0.001p0.001RelativemRNAofIFN-RelativemRNAofIFN-g g g g1d1d3d3d5d5d0 040408080120120160160200200240240280280BALB/cBALB/cTLR4TLR4lps-dlps-dp0.01p0.01p=0.35p=0.35p0.01p0.01DayspostinfectionDayspostinfectionRelativemRNAofIL-18RelativemRNAofIL-181d1d3d3d5d5d0 05050100100150150200200250250BALB/cBALB/cTLR4TLR4lps-dlps-dp0.01p0.01p0.01p0.01p0.01p0.01Pro-inflammatory cytokine prod9797DayspostinfectionDayspostinfectionRelativemRNAofiNOsRelativemRNAofiNOs1d1d3d3d5d5d0 08 81616242432324040BALB/cBALB/cTLR4TLR4lps-dlps-dp0.05p0.05p0.01p0.01p0.05p0.05DayspostinfectionDayspostinfectionRelativemRNAofRelativemRNAofb b b b-Defensin-2-Defensin-21d1d3d3d5d5d0 010102020303040405050BALB/cBALB/cTLR4TLR4lps-dlps-dp0.05p0.05p=0.83p=0.83p0.05p0.05iNOsandb b-defensin-2productionHuangX.etal.(2006)HuangX.etal.(2006)IOVS IOVS 47:4910-491647:4910-4916Days postinfectionRelative mRN9898SummaryofTLR4inBacterialKeratitisIntheinfectedcornea:Intheinfectedcornea:TLR4-deficiencyincreasescornealTLR4-deficiencyincreasescornealinflammation;inflammation;IncreasesTh1-typecytokineproduction;IncreasesTh1-typecytokineproduction;DecreasesiNOsandDecreasesiNOsandb b b b-defensin-2-defensin-2production;production;Reducesbacterialkilling.Reducesbacterialkilling.Summary of TLR4 in Bacterial K9999TLR9inbacterialkeratitis AkiraS.etal.(2005)AkiraS.etal.(2005)Int.Immunol.Int.Immunol.17:1-1417:1-14TLR9 in bacterial keratitis Ak100100BALB/cB601020304050601dayp.i.NormalATLR9mRNAlevelsTLR9IL-1b b MIP-20102030405060GpCcontrolCpGtreatmentBRalativemRNAlevelsIL-12IL-12 IL-18IL-18 IFN-IFN-g g0102030405060GpCcontrolGpCcontrolCpGtreatmentCpGtreatmentCRelativemRNAlevelsIL-4IL-4IL-10IL-100102030405060CpGtreatmentGpCcontrolDRelativemRNAlevelsRelativemRNAlevelsTLR9expressionandsignalingactivationinthecorneaTLR9expressionandsignalingactivationinthecorneaHuangX.etal.(2005)HuangX.etal.(2005)IOVS IOVS 46:4209-421646:4209-4216 BALB/cB601020304050601 day p.101101TLR9templateTLR9templatePCRreactionPCRreactionT7-TLR9T7-TLR9TranscriptionTranscriptionTLR9dsRNATLR9dsRNARNaseIIIRNaseIIITLR9siRNATLR9siRNATLR9tempTLR9tempT7-TLR9T7-TLR9TLR9dsRNATLR9dsRNAOverviewofTLR9-siRNAProductionHuangX.etal.(2005)HuangX.etal.(2005)IOVS IOVS 46:4209-421646:4209-4216RNAinterference(RNAi):RNAinterference(RNAi):anaturallyoccurringbiologicalprocessforanaturallyoccurringbiologicalprocessforsilencinggeneexpressionandfunctionbysiRNA.silencinggeneexpressionandfunctionbysiRNA.SmallinterferingRNA(siRNA):SmallinterferingRNA(siRNA):asmallssRNAfragmentspecificallydegradeasmallssRNAfragmentspecificallydegradehomologousRNAinmammaliancells.homologousRNAinmammaliancells.TLR9 templateATLR9 temp 102102TLR9-siRNAinhibitsmRNAexpressioninvitroHuangX.etal.(2005)HuangX.etal.(2005)IOVS IOVS 46:4209-421646:4209-4216TLR9-siRNA inhibits mRNA expre103103TLR9silencinginvitro(Rho-CpGasareadout)HuangX.etal.(2005)HuangX.etal.(2005)IOVS IOVS 46:4209-421646:4209-4216TLR9 silencing in vitro Huang 104104TLR9SilencingIn Vivo0 01 12 23 34 45 56 6Control-siRNAControl-siRNATLR9-siRNATLR9-siRNA1d1d3d3d5d5dDayspostinfectionDayspostinfectionClinicalscoreClinicalscoreCIL-12IL-12IL-18IL-18IFN-IFN-g g0 0101020203030404050506060Control-siRNATLR9-siRNARelativemRNAlevelsRelativemRNAlevelsDIL-4IL-4IL-10IL-100 0101020203030404050506060TLR9-siRNAControl-siRNARelativemRNAlevelsRelativemRNAlevelsEABTLR9-siRNAControl-siRNATLR9-siRNAControl-siRNAHuangX.etal.(2005)HuangX.etal.(2005)IOVS IOVS 46:4209-421646:4209-4216TLR9 Silencing In Vivo0123456C105105ConfirmingSilencingWorksIn vivoHuangX.etal.(2005)HuangX.etal.(2005)IOVS IOVS 46:4209-421646:4209-4216Confirming Silencing Works In 106106IL-1IL-1 b b b bELISAELISAControlControl TLR9-siRNATLR9-siRNA02000400060008000AIL-1IL-1b b b b(pg/ml)(pg/ml)MIP-2ELISAMIP-2ELISAControlControl TLR9-siRNATLR9-siRNA0400080001200016000BMIP-2(pg/ml)MIP-2(pg/ml)ControlControl TLR9-siRNATLR9-siRNA050100150200250CUnitsMPO/corneaUnitsMPO/corneaControlControl TLR9-siRNATLR9-siRNA0246810DLog10CFU/corneaELISAandMPOAnalysesHuangX.etal.(2005)HuangX.etal.(2005)IOVS IOVS 46:4209-421646:4209-4216I L-1 b ELISAControlTLR9-siRN107107SummaryofTLR9dataIntheinfectedcornea,TLR9-siRNAtreatment:Intheinfectedcornea,TLR9-siRNAtreatment:Reducesinflammation;Reducesinflammation;Down-regulatesTh1-typecytokines;Down-regulatesTh1-typecytokines;Down-regulatesPMNinfiltrate;Down-regulatesPMNinfiltrate;Increasesbacterialload.Increasesbacterialload.Summary of TLR9 dataIn the inf108108科研课题：固有免疫识别与免疫应答科研课题：固有免疫识别与免疫应答TLR9:TLR9:TLR9:TLR9:被细菌被细菌被细菌被细菌DNADNADNADNA激活激活激活激活 导致过度炎症反应导致过度炎症反应导致过度炎症反应导致过度炎症反应 角膜溃疡的关键分子之一角膜溃疡的关键分子之一角膜溃疡的关键分子之一角膜溃疡的关键分子之一HuangXetal,IOVS,2005;46:4209-4216 （第一和通讯作者：黄曦）（第一和通讯作者：黄曦）TLR4:TLR4:TLR4:TLR4:被内毒素被内毒素被内毒素被内毒素LPSLPSLPSLPS激活激活激活激活 免疫病理损害的分子基础免疫病理损害的分子基础免疫病理损害的分子基础免疫病理损害的分子基础 角膜抗感染免疫所必需角膜抗感染免疫所必需角膜抗感染免疫所必需角膜抗感染免疫所必需HuangXetal,IOVS,2006;47:4910-4916 （第一和通讯作者：黄曦）（第一和通讯作者：黄曦）科研科研课题课题：固有免疫：固有免疫识别识别与免疫与免疫应应答答TLR9:TLR4:109109NegativeregulationofTLRsignaling1 1ststSolubleTLRsSolubleTLRssuchassTLR2andsTLR4,actassuchassTLR2andsTLR4,actasdecoyreceptorsforTLR2andTLR4signaling.decoyreceptorsforTLR2andTLR4signaling.2 2ndndTransmembraneproteinsTransmembraneproteinssuchasSIGIRRandsuchasSIGIRRand ST2,sequesterrecruitmentofadaptorsandinhibitST2,sequesterrecruitmentofadaptorsandinhibitformationofTLRsignalingcomplex.formationofTLRsignalingcomplex.3 3rdrdIntracellularregulatorsIntracellularregulatorssuchasMyD88s,SOCS,suchasMyD88s,SOCS,IRAK-M.IRAK-M.4 4ththReductionofTLRReductionofTLRexpressionorincreasedTLRexpressionorincreasedTLRdegradation.degradation.5 5ththTLR-inducedTLR-inducedapoptosisapoptosis.6 6ththTLRTLRtolerancetolerance.Negative regulation of TLR sig110110NegativeregulationofTLR4signalingNegative regulation of TLR4 si111111SIGIRR and ST2:l lMembers of immunoglobulin domain subgroup of TLR/IL-1R Members of immunoglobulin domain subgroup of TLR/IL-1R super-familysuper-familyl lNegatively regulate TLR signalingNegatively regulate TLR signalingl lLigand unknown(SIGIRR)Ligand unknown(SIGIRR)l lLigand for ST2(IL-33)Ligand for ST2(IL-33)Li et al.(2005)J.Mol.Med.83:258-266Li et al.(2005)J.Mol.Med.83:258-266SIGIRR and ST2:Li et al.(200112112TLRTLR负调控：抑制过度炎症反应负调控：抑制过度炎症反应ST2:ST2:ST2:ST2:调控调控调控调控Th1/Th2Th1/Th2Th1/Th2Th1/Th2细胞因子平衡细胞因子平衡细胞因子平衡细胞因子平衡 抑制抑制抑制抑制TLRTLRTLRTLR信号转导通路信号转导通路信号转导通路信号转导通路 抑制免疫病理损害抑制免疫病理损害抑制免疫病理损害抑制免疫病理损害IOVS,2007;48:4626-4633（第一作者：黄曦）（第一作者：黄曦）SIGIRR:SIGIRR:SIGIRR:SIGIRR:直接抑制直接抑制直接抑制直接抑制TLR4TLR4TLR4TLR4表达表达表达表达 角膜抗感染免疫所必需角膜抗感染免疫所必需角膜抗感染免疫所必需角膜抗感染免疫所必需 抑制免疫病理损害抑制免疫病理损害抑制免疫病理损害抑制免疫病理损害J Immunol.2006;177:548-556 （通讯作者：黄曦）（通讯作者：黄曦）TLR负调负调控：抑制控：抑制过过度炎症反度炎症反应应ST2:SIGIRR:113113RoleofSIGIRRinresistanceagainstP.aeruginosakeratitis Role of SIGIRR in resistance 114114Bacteria Pseudomonas aer