专业英语 Unit 3教案学习资料

UnitThreeULTRASTRUCTURALANDIMMUNOHISTOCHEMICALFINDINGSINORALHAIRYLEUKOPLAKIAVirchowsArchivAPatholAnatHistopathol(1988)412:533-542Summary.ThreecasesofHLfromthelateralborderofthetongueofmalehomosexualAIDSpatientswereinvestigatedbythinsectionelectronmicroscopy.Keratinocytescontainedcondensedchromatinintheirpyknoticnucleiandafeworganellesintheedematouscytoplasm.Chromatinwasincloseassociationtothenuclearmembraneandshowedapunched-outappearance.Particletypicaloftheherpesvirusgroupwasabundantintheuppertwothirdsoftheepitheliuminallthreecases.Virusparticleswereseenfrequentlyinthenucleioftheballoonedkeratinocytes,butrarelyincellscontainingCandidaalbicans.Viralnucleocapsidswereobservedbuddingattheinner-nuclearmembrane,therebyacquiringtheprospectiveviralenvelope.Complete,envelopedvirionswerefoundintheendoplasmicreticulumandintheextracellularspace.ThesevirionswereidentifiedimmunohistochemicallyasEpstein-Barrvirus(EBV)usingtwomonoclonalantibodiesdirectedagainstEBVcapsidandmembraneantigen,respectively.Candidaalbicanswasobservedinthestratumcorneumandintheupperlayerofthestratumspinosum.Specialcytoplasmictubularstructuresarrangedinparallelbundleswerefoundinkoilocytoticcellsinadditiontocharacteristicmembranestructurescomposedofundulatingconvolutedmembranes.Epithelialbasementmembraneswerealwaysintact.Keywords:Hairyleukoplakia,Electronmicroscopy,Epstein-Barrvirus,CandidaalblcansIntroductionOralhairyleukoplakia(HL),anewclinicalentity,wasfirstdescribedbyGreeenspanetal.(1984)aspathognomonicfortheHIV-infection.HLischaracterizedbywhitishplaquesatthelateralborderofthetongue,whichcannotberubbedoff.Thelesion,mainlyobservedinHIV-infectedhomosexualmales,wasalsoreportedinheterosexualHIV-infectedindividuals,hemophiliacs,atransfusionrecipient(Greenspanetal.1986;Rindumetal.1987),andintravenous(iv)drugabusers(Reichartetal.1986,L987).AsurvivalanalysisshowedthattheprobabilityofdevelopingAIDSinpatientswithHLwas40%during16monthsand83%during31months(Greenspanetal.1987)Lightmicroscopicinvestigations(Greenspanetal.1984;Greenspanetal.1985;Eversoleetal.1986)indicated,thatthelesionischaracterizedbyparakeratosis,acanthosis,koilocytosisandlackofinflammationinthesubepithelialtissue.Usingelectronmicroscopy(EM)particlesoftheherpesvirusgroupwerefoundinHL(Greenspanetal.1984,1985;BeltonandEversole1986;Konrad1986).ImmunofluorescencestudiesdemonstratedEpstein-Barrvirus(EBV)antigen,andanucleicacid-hybridizationprocedurerevealedEBVviralDNAinahighcopynumber(Greenspanetal.1985;Loningetal.1987)PurposeofthepresentstudywastodescribeultrastructuralfeaturesofHLandtoidentifythevirionparticlesusingmonoclonalantibodiesagainstEBVcoreandmembraneantigens.MaterialsandmethodsFromJuly1984toJuly1987among195HIVseropositiveindividualsatotalof61patients(malen=59,femalen=2;homo-/bisexualn=55,i.v.drugabusetsn=5,bloodtransfusionrecipientn=l)withtheclinicaldiagnosisofHLhavebeenobserved.TheaverageageofHLpatientswas34.2years.AtthetimeofserodiagnosisfourHIVpositiveindividualswereclinicallysymptomless,18patientsshowedARC,and39patientsAIDSsymptoms.ForthepresentstudybiopsiesofHLwereobtainedfrom3malehomosexualpatients(age39,46and83years),showingdifferentAIDSmanifestations(multipleKaposisarcoma:n=l,opportunisticinfections:n=2).Allthreepatientsdiedwithin2.8monthsafterbiopsy.Biopsiesweretakenfromthelateralmarginofthetongueunderlocalanesthesia(UltracainDSR).Priortobiopsyallthreepatientshadbeentreatedfortwoweekswithtopical(Mikonazole),twopatientsadditionallywithsystemicantimycotictherapy(Ketokonamle).Tissuesweredividedandonepartwasfixedin2.5%glutaraldehydeinPBSfor3hours.AfterwashingwithPBSspecimenswerepostfixedinl%OSO4forlhat4ْC,andafterwardstreatedforlhatroomtemperaturewithinl%uranylacetate.Afterdehydrationinagradedseriesofethanol,specimenswereinfiltratedby3changesofpropyleneoxideandembeddedinEpon812followingroutinetechniques(Gelderblometal.1974).Semithinsections(0.5-1minthicknessofeachspecimenswerecut,stainedwithtoluidineblue,andevaluatedlightmicroscopicallyforpathognomonicchanges.Afterfindingsuspectiveareasinlightmicroscopy,ultrathinsections(40-60nm)werecut.Theseweremountedonbaregrids,poststainedwithleadcitrate,stabilizedwithcarbonandexaminedusingaZeissEM10Aat60kV.Theotherpartofthebiopsywasfixedindimethylsuberimidate(DMS,HasselandHand,1974)for2h,washedinPBSandinfiltratedstepwisewith5%,l.0,l.2Mand2.3MsucroseinPBS.Biopsiesweredeepfrozeninliquidnitrogen.Semithincryosectionsof0.5mwerecutusingtheReichertFC4cryoattachment(ReichertAG,Wien).SectionswereincubatedwithmonoclonalantibodiesdirectedagainstEBVcapsid(VCA)andmembraneantigen(MA)(dilutionl:1000inPBS).Immunobindingwasvisualizedusingamodificationofthealkaline-phosphatase-mouse-anti-alkaline-phosphatasetechnique(APAAP;Beckeretal.1987).Controlsincludedtheuseofsecondandthirdstepantibodies,aswellasnormaltissuesofHIVseronegativepersons.ResultCommonfeaturesinallcaseswerehyperkeratosis,parakeratonsisandacanthosisofepithelium.Abovethebasalcelllayeranincreaseddegreeofintracellularedemaofspinouscellswasnoted.Theballoonedkeratinocytesoftenlosttheirchromatinpatternandsometimesbasophilicintranuclearinclusionswereobserved.Candidaalbicanswasfoundwithinepithelialcellofthesuperficiallayers.Infiltrationbyinflammatorycellswasnotapparent,neitherinthelaminapropriaofthemucosanorintheepithelium.Inallcases,balloonedkeratinocyteswereseeninclusterswithinthespinouslayeroftheepitheliumonelectronmicroscopy.Thesecellswereincreasedinsizeandcontainedonlyfeworganelles,includingdegeneratedmitochondriaandabundantintermediatesizedkeratinfibrils.Fociofcondensedchromatinwithapunched-outappearancewerefoundinpyknoticnucleimainlyincloseassociationwiththenuclearmembrane.Candidaalbicanswasfrequentlyobservedinthestratumcorneumaswellasintheupperlayersofthestratumspinosum.Virusparticlesoftheherpesvirusgroupwerepresentintheupperspinouslayermainlyinandaroundkoilocytoticcells,andintheintercellularspaceofflattenedcellsofthesuperficiallayers.ViruswasalsoobservedoccasionallyincellsinfectedbyCandidaalbicans.Thenucleiofinfectedcellscontainedvaryingamountsofviralnucleocapsidswithadiameterof100nm;whilesomecapsidsalreadycontainedtheelectron-densetoroidaldesoxyribonucleoproteinportiontypicalforherpesviruses,othersrepresentedemptyproteincapsids.Occasionally,theegressofherpesviruscapsidsfromthenucleiwasobserved.Duringegressthenucleocapsidacquiresitsfutureviralenvelopemostlyfromtheinnernuclearmembraneinabuddingprocess.Themembranewasthickenedinregionsofnucleocapsidenvelopment.Virusparticleswerealsofoundinthecytoplasm,eitherfreeasnucleocapsidsorascompleteenvelopedparticlescontainedwithintheendoplasmicreticulum.Virionswerefinallyreleasedinabundantnumbersintotheextracellularspace.Enveloped,matureherpestypevirionswithinthecytoplasmandintheextracellularspaceshowedidenticaldiametersof150nm.Twocharacteristicstructuresoccurredwithinthecytoplasmofballoonedkeratinocytesinfectedbythevirus.Oneconsistedoftubules,35nmindiameter,arrangedinparallelbundlesaboutlpminlength.Anotherinclusionbodywascomposedofhighlyundulating,convolutedmembranes.Thesewereobservedclosetothenucleiofkoilocytoticcellsconsistingoftwo,partlyfusedlipidbilayers.Theinnerfusedpartofthisstructurewaselectrondenseand25nminwidth.Thesubepithelialbasementmembranedidnotrevealanyirregularities.LangerhanscellswerenotobservedwithintheHLlesions.AspecificimmunostainingwasnotedintheuppertwothirdsoforalepitheliausingsemithincryosectionsandEBV-specificmonoclonalantibodies.AntibodiesagainstEBVcapsid(VCA)andmembraneantigens(MA)revealedacompletelydifferentstainingpattern.WhileVCAwaspredominantinandaroundthenucleiofkeratinocytes,MAwasobservedinhighconcentrationintheinterepithelialspacesinthesametissueareas,whereEBVVCAwasobservedinconsecutivesections.DiscussionHL,anexclusivelyorallyoccuringlesion,wasreportedtobeanearlyindicatorofHIV-associatedimmunosuppression(Eversoleetal.1986;Silvermannetal.1986).TheoccurrenceofHLindifferentriskgroups,asobservedinthepresentstudy,haswellbeendocumentedintheliterature(Greenspanetal.1984,1986;Reichartetal.1987;Rindumetal.1987).TheaetiologyofHLhaslongbeensupposedtobevirus-relatedandinmostinvestigationsaherpestypeviruswasdetected(Greenspanetal.1984.1985;Konradetal.1986).Furthermore,insomeofthesestudiespapillomavirus-likeparticleswereobservedinadditiontotheherpestypevirus(Greenspanetal.1984,1985).WhileCreenspanetal.(1984,1985)reporteddoubleinfectionsinHLbyEBV(usinginsituhybridization)andHPV(EM,immunohistochemistry),wehavenotbeenabletodetectpapillomavirusinHLontheEMlevelinthisinvestigation.InafurtherinsituhybridizationstudyofsevenHLallspecimenswerenegativewiththeappliedHPVtype6,11,13,16,18probeswhilefiveofthesespecimenswereEBVpositive(Loningetal.1987)Fromtheobservationthatpapillomavirus-likeparticleswerepresentinall23casesofHLattheEMlevelGreenspanetal.(1985)consideredwhetherthecoexistenceofEBVandHPVinHLisfortuitous.SinceHPVwasnotdemonstratedinourthreecasesorinafurther7cases(Loningetal.1987)ourfindingsdonotsupporttheconceptofsimultaneousinfectionofHL.However,negativefindingsregardingHPVdonotruleoutapossibleinvolvementofasecondvirus.IndifferenttissuesfrompatientswithARCorAIDSintracytoplasmicinclusionshavebeendescribed,appearingasvesicularrosettes,testtubeandring-shapedforms(TRF),ortubuloreticularstructures(TRS)(Onerheimetal.1984;Kuntzetal.1987).WithinthecytoplasmofEBVproducingcells,weobservedonlyonetypeofparalleltubularstructures,whichresembledinbothsizeandappearancetheinclusionsdescribedbyBeltonandEversole(1986).Theoriginandnatureoftheseaggregatesisasyetunknown.Whiletheoccurrenceoftheseparallelarranged,intracytoplasmictubularstructureshasonlybeendescribedinHL.TRFinclusionshavealsobeenfoundinassociationwithdifferentimmunologicalandneurodegenerativediseasesaswellasinviralinfections.Thesemembranederivedinclusionsappearedtobeassociatedwiththeproductionofalpha-andbeta-interferon,butnotofgamma-interferon(Onerheimetal.L984).Theoccurrenceoftheseaggregatesmaybeamorphologicalindicatorofinterferonproduction,inducedbyviralinfection,andmayreflecttheself-perpetuatingimmunologicaldysfunction.Inaddition,undulatingmembranestructureswereobservedwithinthecytoplasmofkoilocytoticcells,consistingofpartlyfusedmembranessimilartotheabovementionedTRF/TRS.ItisprobablethatthesemembranestructuresareindeedprecursorsofTRF/TRS.WedidnotobserveLangerhanscellsorothersignsofcellularimmunereactionintheepithelium.ClearanceofCandidaalbicansbycellularimmunemechanismswasnotfoundinourstudynorinotherreports(BeltonandEversole1986).ThisisprobablyduetodefectivemonocytemigratoryfunctioninAIDS-patientsandgivesreasontoexpectfurtherfunctionaldisturbancesinantigenprocessingandpresentationbymonocytesandLangerhanscells(Danielsetal.1987;Beckeretal.1987).RegardingtheetiologyofHL,ourEMandimmunohistochemicalfindingscorroboratethecloseassociationofEBVwiththislesion.Fromclinicalobservationsthevirusseemstobeinvolvedinthepathogenesis.ofHL,sinceduringtreatmentwithhighdosesofacyclovir(4x800mg/dayduring14days)clinicalregressionofthelesionhasbeenfound(Freidman-Kien1986).TheinvolvementofEBVinHLmaybeimportantforthefurtherprogressoftheundelyingHIVdisease,becauseEBVisknowntobecapableoftransformingB-lymphocytessothatHIV(Montagnieretal.1984)caninfectthem.TherecruitmentofsuchtransformedB-cellsinadditiontotheCD4+T-helperlymphocytes,monocytes/macrophagesandLangerhanscellsmayenhanceHIVproductionconsiderably.ThisthendirectlyleadstoahigherlevelofinfectedcellsengagedinHIVreplication,andconcomitantlytoacontinuouslyincreasinglossofCD4+-lymphocytesbydirectcytotoxicityofHIVand/orbythemanifoldHIVinducedimmunopathogenicmechanisms(Gelderbolometal.1985;KlatzmannandGluckman1986).Finally,themassesofEBVproducingcellsinHLareindicativeofasevereimmunodeficiencyobservedinthesepatients.VOCABULARYl.hairyleukoplakia毛状白斑2.homosexual同性交的n．同性恋者3.AIDS(AcquiredImmunodeficiencySyndrome)艾滋病4.thinsection超薄切片的别称(电镜技术用语)5.semithinsection半薄切片(电镜技术用语)6.condensedchromatin“浓缩的”染色质7.punched-outappearance穿孔样表现(改变)8.pyknoticnuclei“皱缩的”细胞核9.herpesvirusgroup疱疹病毒族10.balloonedkeratinocyte“汽球状”角质细胞11.Candidaalbicans白色念珠菌12.nucleocapsid核壳，外壳13.bud芽，vi．芽生14.viralenvelop病毒包膜15.endoplasmicreticulum内质网16.extracellularspace细胞间隙17.Epstein-BarrvirusEB病毒(γ组疱疹病毒)18.monoclonalantibody单克隆抗体19.against抗…20.stratumcorneum角质细胞层21.stratumspinosum棘细胞层22.koilocytoticcell“凹状”细胞23.undulatingconvoluted波状的，折迭状的24.basementmembrane基膜25.pathognomonic特定的症状，有确诊意义的症状、表现26.HIV-infectionHIV感染27.plaquen．斑，溶菌斑；血小板28.ruboff擦掉(撕脱)29.heterosexual异性恋的n异性恋者30.hemophiliac血友病人31.intra-venousdrugabuser静脉吸毒者32.survival生存，成活33.parakeratosis不全角化症34.acanthosis棘层增生症35.subepithelialtissue上皮下组织36.nucleicacid-hybridization核酸杂交37.EBVcoreEB病毒的“核”38.seropositive血清阴性的39.homo-/bisexual同性／同性、异性恋的40.serodiagnosis血清学诊断41.ARC(AIDSRelatedComplex)艾滋相关症候群42.biopsy活体组织的病理检查43.multiple多重的，多倍的44.KaposiSacoma卡波济肉瘤45.opportunisticinfection机遇性感染46.prior在前面的47.glutaraldehyde戊二醛48.washing刷洗49.postfixed后固定的50.uranylacetate醋酸双氧铀51.dehydration脱水52.gradedseriesofethanol梯度酒精53.infiltrate浸润54.propyleneoxide氧化丙烯55.Epon812包埋电镜标本的一种环氧树脂56.toluidineblue甲苯胺蓝57.ultrathinsection超薄切片58.nm(nanometre)纳米，毫微米59.mounted裱的60.baregrid裸(铜)网，超薄切片的戴网61.leadcitrate柠檬酸铅62.stabilizedwithcarbon(喷)镀碳膜63.dimethylsuberimidate(DMS)类似于二甲基亚矾的一种冷冻保护剂64.stepwise逐步的65.sucrose蔗糖66.cryosection冷冻切片67.cryoattachment作冷冻切片的附加设备68.incubate孵育69.immunobinding免疫结合(粘合)70.visualize显现71.modification改良72.control对照73.secondantibody第二抗体74.seronegative血清阴性的75.adhering粘附76.hyperkeratosis过度角化症77.basophilic嗜碱性的78.intranuclearinclusion核内包涵体79.superficiallayer浅层80.laminapropria