Hypergranular γδ T-Cell Lymphoma in a Heifer · 2018. 11. 13. · but slight erythrophagia was...
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79 Introduction Human T-cell large granular lymphocyte (LGL) leu- kemia is a heterogeneous disorder characterized by a per- sistent increase in the number of peripheral blood LGLs with fine or coarse azurophilic granules 1 . The granules contain a number of proteins that play a role in cytolysis such as perforin and granzyme B 1 . αβ T-cell receptors (TCRs) are expressed in most cases, and those with γδ TCRs are very rare 1,8 . A feline LGL lymphoma express- ing surface CD3 and perforin was thought to be of T-cell origin 3 . Although perforin-like activity was not observed in 6 cases of feline LGL neoplasm, 2 cases expressed αβ TCRs 2 . In cattle, γδ T-cell lymphoma is a neoplasm char- acterized by positive reactivity for WC1 and perforin, and epitheliotropism is seen in most cases 4,7,11 . Natural killer (NK)-like T-cell lymphoma has been reported in a calf, and many neoplastic cells had eosinophilic cytoplasmic granules that were of variable size and positive for per- forin 5 . Here we report a case of hypergranular γδ T-cell lymphoma in a heifer that did not resemble the human counterpart at all and was considered to be a cytologic variant of γδ T-cell lymphoma. Materials and methods 1. Animal A 16-month-old Holstein heifer was examined because of a 3-day history of anorexia. Clinical exami- nation revealed emaciation, bilateral exophthalmos with epiphora, and hyperthermia (41ºC). Despite antimicrobial treatment, the heifer’s condition worsened with progres- sive exophthalmos, and it was euthanized one week after the first examination. At necropsy, there were several tumor masses up to 5 cm in diameter on the vulva, perineum and perianal area. On cut section, they were homogeneous, white and fleshy. The skin of the udder showed diffuse swelling due to neoplastic involvement of the dermis and subcutis. The ocular muscles were almost completely replaced by white neoplastic tissue, and the glossal muscles were severely affected. There were sparsely distributed nodules up to Hypergranular γδ T-Cell Lymphoma in a Heifer Kazumi NAITOU 1 , Yoshiharu ISHIKAWA 2 and Koichi KADOTA 2 * 1 Tobu Livestock Hygiene Service Center,Yamanashi Prefecture (Fuefuki, Yamanashi 406–0034, Japan) 2 Hokkaido Research Station, National Institute of Animal Health (Toyohira, Sapporo 062–0045, Japan) Abstract Lymphoma with large granular lymphocyte (LGL) morphology in a heifer is described. The most noticeable macroscopic finding was neoplastic involvement of the skin, mucosae of the alimentary tract, urinary bladder, and uterus. Histologically, lymphoma cells with large numbers of fine eosinophilic granules predominated in the neoplastic tissue, and the granules were positive for perforin. Surface CD3, CD5 or WC1 was expressed by some neoplastic cells. In addition to the epitheliotropism that is a feature of ordinary γδ T-cell lymphoma, the neoplastic cells showed angiodestruction and erythropha- gocytosis. These phenomena were thought to result from marked activation of neoplastic γδ T-cells. The present lymphoma was distinct from the previously reported bovine NK-like T-cell lymphoma in its tissue distribution, cytomorphology, cytochemistry, and immunophenotype. Since this lymphoma was regarded as a cytologic variant of γδ T-cell lymphoma, the term hypergranular γδ T-cell lymphoma seemed to be more appropriate than γδ T-cell LGL lymphoma. Discipline: Animal health Additional key words: angiodestructive growth pattern, large granular lymphocyte lymphoma, perforin, WC1 JARQ 41 (1), 79 – 83 (2007) http://www.jircas.affrc.go.jp *Corresponding author: e-mail [email protected] Received 17 April 2006; accepted 30 May 2006.
Transcript of Hypergranular γδ T-Cell Lymphoma in a Heifer · 2018. 11. 13. · but slight erythrophagia was...
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Introduction
HumanT-celllargegranularlymphocyte(LGL)leu-kemiaisaheterogeneousdisordercharacterizedbyaper-sistentincreaseinthenumberofperipheralbloodLGLswithfineorcoarseazurophilicgranules1.ThegranulescontainanumberofproteinsthatplayaroleincytolysissuchasperforinandgranzymeB1. αβT-cellreceptors(TCRs)areexpressedinmostcases,andthosewithγδTCRsareveryrare1,8.AfelineLGLlymphomaexpress-ingsurfaceCD3andperforinwasthoughttobeofT-cellorigin3.Althoughperforin-likeactivitywasnotobservedin6casesoffelineLGLneoplasm,2casesexpressedαβTCRs2.
Incattle,γδT-celllymphomaisaneoplasmchar-acterizedbypositivereactivityforWC1andperforin,andepitheliotropismisseeninmostcases4,7,11.Naturalkiller(NK)-likeT-celllymphomahasbeenreportedinacalf,andmanyneoplasticcellshadeosinophiliccytoplasmicgranulesthatwereofvariablesizeandpositiveforper-forin5.HerewereportacaseofhypergranularγδT-cell
lymphomainaheiferthatdidnotresemblethehumancounterpartatallandwasconsideredtobeacytologicvariantofγδT-celllymphoma.
Materials and methods
1. AnimalA 16-month-old Holstein heifer was examined
becauseofa3-dayhistoryofanorexia.Clinicalexami-nationrevealedemaciation,bilateralexophthalmoswithepiphora,andhyperthermia(41ºC).Despiteantimicrobialtreatment,theheifer’sconditionworsenedwithprogres-siveexophthalmos,anditwaseuthanizedoneweekafterthefirstexamination.
Atnecropsy, therewereseveral tumormassesupto5cmindiameteronthevulva,perineumandperianalarea.Oncutsection,theywerehomogeneous,whiteandfleshy.Theskinoftheuddershoweddiffuseswellingduetoneoplasticinvolvementofthedermisandsubcutis.Theocularmuscleswerealmostcompletelyreplacedbywhiteneoplastictissue,andtheglossalmuscleswereseverelyaffected.Thereweresparselydistributednodulesupto
Hypergranular γδ T-Cell Lymphoma in a Heifer
Kazumi NAITOU1, Yoshiharu ISHIKAWA2 and Koichi KADOTA2*1TobuLivestockHygieneServiceCenter,YamanashiPrefecture
(Fuefuki,Yamanashi406–0034,Japan)2HokkaidoResearchStation,NationalInstituteofAnimalHealth
(Toyohira,Sapporo062–0045,Japan)
AbstractLymphomawithlargegranularlymphocyte(LGL)morphologyinaheiferisdescribed.Themostnoticeablemacroscopicfindingwasneoplasticinvolvementoftheskin,mucosaeofthealimentarytract,urinarybladder,anduterus.Histologically,lymphomacellswithlargenumbersoffineeosinophilicgranulespredominatedintheneoplastictissue,andthegranuleswerepositiveforperforin.SurfaceCD3,CD5orWC1wasexpressedbysomeneoplasticcells.InadditiontotheepitheliotropismthatisafeatureofordinaryγδT-celllymphoma,theneoplasticcellsshowedangiodestructionanderythropha-gocytosis.ThesephenomenawerethoughttoresultfrommarkedactivationofneoplasticγδT-cells.ThepresentlymphomawasdistinctfromthepreviouslyreportedbovineNK-likeT-celllymphomainitstissuedistribution,cytomorphology,cytochemistry,andimmunophenotype.SincethislymphomawasregardedasacytologicvariantofγδT-celllymphoma,thetermhypergranularγδT-celllymphomaseemedtobemoreappropriatethanγδT-cellLGLlymphoma.
Discipline:AnimalhealthAdditional key words:angiodestructivegrowthpattern,largegranularlymphocytelymphoma,
perforin,WC1
JARQ41(1),79–83(2007)http://www.jircas.affrc.go.jp
*Correspondingauthor:[email protected];accepted30May2006.
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1.5cmindiameterintheobliquusinternusabdominisandglutealmuscles.Similarnoduleswerealsodetectedinsomeotherskeletalmuscles,andappearedtobegeneral-izedindistribution.
Thereweremultiple tumormasses,5to10cmindiameter,onthemucosalsurfaceoftherumen.Similarbutsmalleronesweredetectedinthereticulumandoma-sum,andthewalloftheabomasumwasdiffuselythick-ened.Variouslysizedtumormasseswerescatteredinthejejunumandileum,andthemajoritywereonthemucosalsurface.Scatteredtumormasseswerefoundontheserosaofthecolonandrectum.Nodules1to2cmindiameterwereoccasionallyseenintheliver.
Multiplenodules,0.5to1cmindiameter,werepres-entonthemucosalsurfaceoftheurinarybladder.Almostallpartsoftheuretericwallwerethickened.Frequently,themucosalsurfacesoftheoviduct,uterus,vagina,andvestibulumwere elevatedbyneoplasticproliferationsupto5cmindiameter.Somesmalleroneswerepresentontheserosalsurfaces.Mostpartsofbothovarieswerereplacedbyneoplastictissue.
Thesuperficial,abdominalandthoraciclymphnodeswereenlargedtovaryingdegrees.Thecapsuleofhepaticlymphnodeswasinfiltratedwithneoplastictissue,whichextended into theadjacentpancreatic tissue. The leftkidney,spleenandlunghadsingleneoplasticnodules.Numerousneoplasticfociwerepresent throughout theheart.Severaltumornodulesupto1cmindiameterwereadherenttotheduramatter.
2. Histology, immunohistochemistry and electron microscopy
Tissuesampleswerefixedin10%phosphate-buff-eredformalin,embeddedinparaffin,sectionedat4μm,andstainedwithhematoxylinandeosin(HE),Giemsa,phosphotungsticacidhematoxylin(PTAH)andnaphtholAS-Dchloroacetateesterase(CAE). Selectedparaffinsectionsweredewaxedandlabeledbytheavidin-biotin-peroxidasecomplex(ABC)method.Theprimaryantibod-iesusedwererabbitpolyclonalantibodiestoCD3ε(Dako,Glostrup, Denmark) and CD5 (Lab Vision, Fremont,CA,USA),andmousemonoclonalantibodiestoCD79a(Dako),CD57(BioGenexLaboratories,SanRamon,CA,USA)andWC1-N3(VeterinaryMedicalResearchandDevelopment,Pullman,WA,USA).Subsequentproce-dureswerecarriedoutbymeansofanimmunoperoxidaselabelingsystem(Nichirei,Tokyo,Japan).Smallpiecesfromformalin-fixedtissueswerepost-fixedin1%osmiumtetroxide,andembeddedinepoxyresin.Ultrathinsec-tionswerestainedwithuranylacetateandleadcitrate,andexaminedbyelectronmicroscopy(EM).
Results
Histologically,thereweredensediffusegrowthsoflymphomacellsinallofthenodularormassivelesionsexamined,butanangiocentricorangiodestructivegrowthpatternwasfoundsporadically(Fig.1A).Themostout-standingfindingwasepitheliotropism,andintraepithelialtumorcellscouldbeascertainedintheskin,rumen(Fig.1B),reticulum,omasum,abomasum,jejunum,ileum,uri-narybladder,uterus,andbronchiolarepithelium.Variousdegreesofneoplasticinvasionwereobservedinthelymphnodes,andtherewereneoplasticinfiltratesintheparacor-ticalzonesofpartiallyinvolvedlymphnodes.
Theneoplasticcellswere6to15µmindiameter,withapredominanceof largercells. Thenucleiwerevesicularandround,ovoidorirregular,withinconspicu-oustomedium-sizednucleoli.Thecytoplasmwasabun-dant,andmanyneoplasticcellscontainedfineeosino-philicgranules (Fig.1C). Incertainplaces,however,agranular lymphomacellsproliferatedpreponderantlyorexclusively(Fig.1D).ThegranulesstainedredwithGiemsa,andwerepositiveforPTAHbutnotforCAE.Afairnumberoftumorcellsshowederythrophagocytosis.Mitoticfigureswereplentiful.
Immunohistochemically, occasional neoplasticcellsstainedpositivelyforsurfaceCD3,CD5orWC1.NotumorcellsshowedpositivereactivityforCD57andCD79a. The intracytoplasmic granules were weaklypositiveforperforin(Fig.2A).Cytokeratinstaining,forwhichepithelialcellsstainedpositively,highlightedtheepitheliotropismoftheneoplasticcells(Fig.2B).
Ultrastructurally,themostcharacteristicfeatureofthelymphomacellswasthepresenceofelectron-densegranulesinthecytoplasm(Fig.3),thoughagranularcellswererarelyseen.Thegranulesvariedbetween0.1–0.8μmindiameter.Thegranulesizetendedtobelargerincellswithabundantgranules.Thereweremoderatequan-titiesofroughendoplasmicreticulum(RER)incellswithmanygranules,whereascellswithfewornogranuleswerecharacterizedbythepaucityoforganelles. Mostnucleiwereirregularincontour,havingabundanteuchro-matin.
Discussion
In thecasedescribedhere,manyneoplasticcellsthathadnumerouseosinophilicgranulesinthecytoplasmwere reminiscentofmastcellsormyelocytes,butdidnotshowmetachromasiaanddidnotstainwithperoxi-daseorCAE9,10.OnthebasisoftheexpressionofCD3,CD5,WC1,andperforinbyneoplasticcellsaswellastheabundanceofcytoplasmicgranules,apresumptive
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A B
Fig. 1. Histology of neoplastic lesions of the heiferA:Portahepatis.Twobloodvesselsareheavilyinvadedanddestroyedbyneoplasticcells.HE.×100.B:Rumen.Tumorcellsareisolatedoringroupswithintheepithe-lium.HE.×200.C:Jejunum.Almostalltumorcellshavenumerousacidophilicgranulesinthecytoplasm,whichareresponsiblefordisplacementofthenuclei.HE.×600.D:Jejunum.AlthoughthisisthesamelesionasinFig.1C,lymphomacellswithgranules(arrows)areinconspicuous.HE.×600.
Fig. 2. Immunohistochemistry of neoplastic lesions of the heiferA:Jejunum.Arrowsindicateneoplasticcellsthatareweaklypositiveforperforin.ABC.×1,000.B:Urinarybladder.Thetransitionalepitheliumisclearlyseenwithcytokeratin staining,whichhighlights theprominentepitheliotropism of cytokeratin-negative lymphomacells(arrows).ABC.×600.
A B
C D
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diagnosisofγδT-cellLGLlymphomawasmade1,4.Inhumans,themajorityofT-cellLGLneoplasmsareposi-tiveforαβTCRs,andcasesexpressingγδTCRsarerare1,8.Comparedwithhumancases,thecytoplasmicgranulesweremuchmoreprominentinthecurrentcase,andwerereadilydiscernibleinhistologicsections.Moreover,thislymphomawasquitedissimilarclinicopathologicallytothehumancounterpart,whichisachroniclymphoidleu-kemiaandhasanindolentclinicalcourse1.
Inthesinusoidsofthenormalratliver,LGLshavenumeroussmallcytoplasmicgranules,whereasperipheralbloodLGLscontainafewlargergranules12.ThehepaticLGLsareconsideredtobeaspecificpopulationofhighlyactivatedorfurtherdifferentiatedLGLs12.Withthisinmind,thepresentlymphomawithneoplasticcellshavingnumeroussmallgranuleswasinterpretableasaneoplasmofpredominantlyactivatedLGLs,andthehypergranularcellsmightbeintheultimatestageofγδT-cellactivation.ThiscasewascytologicallyandcytochemicallydistinctfromabovineNK-likeT-celllymphomawhosecompo-nentcellshadCAE-positivegranulesofvarioussizesandnumbers5.Inaddition,thelymphomashowedlittleten-dencytoformtumormassesandtheliverwasseverelyaffected5.
Although the number of cytotoxic granules wasmuchgreaterthaninordinaryγδT-celllymphomas11,thepresentneoplasmwasverysimilartosomeoftheselym-phomasinitstissuedistributionandepitheliotropism4,7.Theneoplasmwasregardedastheircytologicsubtype,andwasdesignatedhypergranularγδT-celllymphoma.AnangiodestructivegrowthpatternischaracteristicofhumanNK-cellneoplasms,butfocalvasodestructivepro-liferationmayalsobepresentinhumanγδT-celllympho-mas13.Thepresentneoplasmshowedasimilargrowthpatternandconsiderableerythrophagocytosis.Thefor-merhasnotbeenseeninbovineγδT-celllymphomas,butslighterythrophagiawasdetectedinameningealγδT-celllymphoma11.ThesephenomenainthepresentcasemaybeduetomarkedactivationofneoplasticγδT-cells;abundantcytotoxicgranulesarepresumablyassociatedwithdestructionofbloodvessels6.
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Fig. 3. Electron microscopy of tumor cellsUterus.Inhabitingthecytoplasmoflymphomacells,agreatnumberofelectron-densegranulesareconcentratedtoonesideofthecells.Onelymphomacellwithfewgranules(arrow)isalsovisible.EM.×6,000.
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