Lab 20 – Aplastic Anemias
GOAL
The studentwill be able tooutline the causes,pathogenesis,pathology,clinical picture andout...
and megakaryocytes.The biopsyalsoindicated decreasedmarrow cellularity(~35-40%) withfattycell replacement.
Diagnosticconsi...
3. Aplastic Anemia: Pathogenesis
Theoretically,the bone marrow failure mayresultfromtwo
mechanisms:eithera) damage toor de...
Aplasticanemiasharesnumerousclinical andpathophysiologicfeatureswith otherautoimmunedisorders.Eachof these
disordersischar...
5. Aplastic Anemia: Pathology
The bone marrow inaplasticanemiashowsvariablyreducedcellularity,dependingonthe clinical stag...
8. Aplastic Anemia Bone MarrowBiopsy:Low & HighMagnification
Thisinteractive image depictsabone marrowbiopsy
specimeninapl...
11. Aplastic Anemia: Treatment
Transfusion:Patientswithaplasticanemiarequiretransfusionsupportuntilthe diagnosisisestablis...
15. Anemia ofRenal Disease:Peripheral BloodSmear,Medium Magnification
The anemiaof renal failure resultsdirectlyfroma
defi...
storage ironsuch inthisimage of bone marrowaspirationanddecreasedserumiron.These abnormalitiesof iron
distributionare gene...
Quiz
1) Which of the followingfeaturesinthe CBCcountdoesnotagree witha diagnosisof aplasticanemia?
A) Neutropenia
B) Normo...
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Lab 20 – Aplastic Anemias

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Lab 20 – Aplastic Anemias

  1. 1. Lab 20 – Aplastic Anemias GOAL The studentwill be able tooutline the causes,pathogenesis,pathology,clinical picture andoutcome of aplasticanemia, anemiaof chronicdisease andanemiaof renal failure Objectives: Upon completionof thiscase the studentwill be able to: 1. List the majorcausesof aplasticanemia.Describe the twomechanismspostulatedtoexplainthe failureof stem cellsinaplasticstates 2. List the causesof anemiadue to bone marrow infiltrations,anddescribethe expectedbloodfindings 3. Define myelophthisicanemia 4. Describe the epidemiologyof aplasticanemia 5. Describe the clinical presentationof aplasticanemia 6. Understandimportance of bone marrowaspirationandbiopsyinthe diagnosisof aplasticanemia 7. Describe the bone marrowcellularityandperipheral bloodcountsinaplasticanemia 8. Discussthe prognosisof aplasticanemia 9. Reviewthe pathogenesisandhematologicfindingsof anemiaof chronicdisease 10. Understandthe mechanismof anemiaof renal failure andhow thisdiffersfromall otherformsof anemia RESOURCES Robbins& CotranPathologicBasisof Disease (7thedition)Chapter13,pp. 646-649 RonaldHoffman(editor) Hematology:BasicPrinciplesandPractice (3rdedition) ChurchillLivingstone,2000, pp.383-388 CASE HISTORY PatriciaD., age 62, comesto youcomplainngthatshe'sexperiencedgraduallyincreasingtirednessandlackof energy overthe past month.She'sseekingmedical helpafterunsuccessful self-medicationwithstress-formulavitamins,ather husband'sinsistence. Her onlyothermedical problemisosteoarthritis,forwhichshe hasbeentakingphenylbutazone (100mg TID) forthe past 6 months,as well asaspirinona PRN basis(generally4-6regularstrengthtabletsdaily). The physical examinationisunremarkable exceptforpallor,several bruisesonvariouspartsof the bodyandfindings typical forosteoarthritis.However,the laboratorytestsreveal aredbloodcell countof 2.5 million/mm3(normal 4.2- 6.2), a hemoglobinof 6.2gr/dL (normal 13-18), a reticulocyte countof 0.5% (normal 0.5-1.5% of redbloodcells),awhite bloodcountof 1800/mm3 (normal 4,500-11,000) with50% neutrophils(normal 57-67% of leukocyte differential) anda plateletcountof 35,000/mm3(normal 150,000-350,000). On the basisof the above findings,abone marrow aspirationandbiopsywere performedatthe posterioriliaccrestand spine.The aspirate revealedmoderatelyhypocellularmarrow withreducedgranulocyteandredbloodcell precursors
  2. 2. and megakaryocytes.The biopsyalsoindicated decreasedmarrow cellularity(~35-40%) withfattycell replacement. Diagnosticconsiderationsinclude drug-inducedaplasticanemia. Clinical diagnosis:Aplasticanemia,mostprobably induced by phenylbutazone 1. Aplastic Anemia: Definition Aplasticanemiaisadisorderof pluripotential stemcellsthatleadstobone marrow failure.The disorderfeatures hypocellularbone marrowandpancytopenia(decreasedcirculatinglevelsof all formedelementsinthe blood).The term “pancytopenia”isusedwhenthere isareductioninall three marrow lineages(granulocytes,plateletsandredblood cells).“Bicytopenia”isareductionintwoof three bone marrow cell lines.“Monocytopenia”or“single cytopenia”is reductioninonlyone of three cell lines. Aplasticanemiaisusuallycharacterizedbyasevere diminuitioninbone marrow functionthataffectsall the hematopoieticlineages(pancytopenia).Granulocyte,plateletandredbloodcell levelsmaynotbe depresseduniformly and sometimespancytopeniaisnotobservedbutoddcombinationsof bicytopeniaandevenmonocytopeniasare observedinstead. 2. Aplastic Anemia: Etiology There are three etiologicgroupsof aplasticanemia: 1) Idiopathicaplasticanemia(2/3of all cases) 2) Acquiredaplasticanemia,i.e. secondaryto:  Drugs usedincancer chemotherapy  Idiosyncraticeffectof drugssuchas phenylbutazone,gold(usedtotreatjoint diseases) andchloramphenicol (antibiotic)  Chemicals(benzene,insecticides)  Viral infections(hepatitisviruses,Ebstein-Barr virus,HIV,parvovirus) 3) Inherited(rare)  Fanconi anemia
  3. 3. 3. Aplastic Anemia: Pathogenesis Theoretically,the bone marrow failure mayresultfromtwo mechanisms:eithera) damage toor defectsinthe stemand progenitorcellsinbone marrow;orb) damage toor defectsinthe bone marrow stromal cells.Damagedstemcellscanproduce progeny expressingneo-antigensthatevoke anautoimmune reaction,orgive rise to a clonal populationwithreducedproliferativecapacity.Either pathwaycouldleadto marrow aplasia. A consistentlaboratoryfindinginaplasticanemiaisthe verylow numbersof assayable bone marrow progenitorandstemcells.Their total numbersare dramaticallydecreased.Colonyformationbybone marrow cellsof patientswithaplasticanemiaremainsunresponsive eventohighlevelsof hepatopoieticgrowthfactors. The survival andproliferationof hematopioeticcellsare dependenton stromal marrow cells.Stromal cell functionisnotdefective inaplastic anemiaandtheyproduce normal quantitiesof hematopoieticgrowthfactors.Adequate stromal functionisimplicitin the successof bone marrowtransplantationinaplasticanemia,because importantstromal elementsremainof host origin. The pathogenesisof aplasticanemiaisnotfullyunderstood.Indeeditisunlikelythatasingle mechanismunderliesall causesof marrowaplasia.Twomajor mechanismshave beeninvokedtoexplainacquiredaplasticanemia: 1) Directhematopoieticinjury: Certainchemical orphysical agentsinjurydirectlybothproliferatingandquiescenthematopoieticcells.Suchagents cause significantdamage toDNA to resultinapoptosis.  Approximately25%of all the cases of aplasticanemiacouldbe blamedondruguse.There are twoclassesof drugsthat cause bone marrowfailure: A) Drugs usedincancer chemotherapy(cisplatin,mercaptopurine,vincristine,etc.) regularlyinduce,inadose - dependentmanner,marrowaplasia;thisisanexpectedeffectanditis counterbalancedbythe benefitthese drugs provide incancertreatment. B) Most of aplasticanemiasassociatedwithmedical drugsisdescribedasidiosyncratic, i.e. itisunexpected,andcaused by drugsthat are notknownto induce regularlymarrow failure.The term“idiosyncracy”inthiscontextmeansan unusual individualreactiontoa drug or a food.Most marrow failure complicationsfromthese drugsoccuraftera few weeksof initiationof druguse.  Viruseshave alsobeenhypothesizedtobe involvedinthe pathophysiologyof aplasticanemias.Inadditionto theirdirecttoxiceffects,chemicalsandvirusesmayinduce compleximmunereactionsleadingtobone marrow failure inaplasticanemia. 2) Immune-mediatedbone marrowfailure.
  4. 4. Aplasticanemiasharesnumerousclinical andpathophysiologicfeatureswith otherautoimmunedisorders.Eachof these disordersischaracterizedbyT-cell mediated,specificorgandestruction.Commontoall these disordersthere isa cytotoxicT-lymphocyteactivation,cytokine productionandspecifictargetcell elimination.  In aplasticanemias,chemicalsandviral antigensare thoughttoinitiate the autoimmuneprocess.  It ispostulatedthatstemcellsare firstantigenicallyalteredbyexposure todrugs,infectiousagents,orother unidentifiedenvironmental insults.Thisevokesacellularimmuneresponse,duringwhichactivatedTcells produce cytokinessuchasinterferon-γ andTNFthat preventnormal stemcell growthanddevelopment.  Thisscenarioissupportedbyseveral observations.Immunosuppressive therapywithantithymocyte globulin combinedwithdrugssuchas cyclosporine producesresponsesin60% to 70% of patients.Successfulbone marrow transplantationrequires"conditioning"withhighdosesof myelotoxicdrugsorradiation.Inboth instances,itishypothesizedthese therapiesworkbysuppressingorkillingautoreactiveT-cell clones.The target antigensforT-cell attackare notwell defined.  The bloodmarrow andbloodfromaplasticanemiapatientscontainelevatednumbersof activatedcytotoxicT- lymphocytes.ActivatedcytotoxicT-cellsoverproduce bothinterferongamma(IFN-γ) andtumornecrosisfactor (TNF).These twocytokinesinhibithematopoieticproliferationof progenitorandstemcells.  The accumulationof activatedcytotoxicT-lymphocytesandthe secretion of lymphokinesare specifically localizedtothe affectedtargettissue, i.e.,the bone marrow.The inhibitoryeffectsonhematopoietic proliferationare farmore potentwhenthose lymphokinesare secretedintothe bone marrow microenvironmentthanwhentheyare simplyaddedtothe cultures.  INFγ and TNFnot onlycan suppresshematopoiesisbyinhibitoryeffectsoncell proliferation,butanimportant componentof theirinhibitoryactivityiscell deathbyinductionof apoptosis.Apparently,immune-mediatedcell cycle blockade andapoptosiscanleadto the dramaticeliminationof hematopoieticprogenitorandstemcellsin aplasticanemias.  The earlyimmune systemeventsthatprecede the global destructionof hematopoieticcellsare muchlessclear. The initial antigenandthe eventsthatleadtobreachedtolerance andthe subsequentprocessof spreading autoimmunityremainundetermined. 4. Aplastic Anemia: Epidemiology The annual incidence of aplasticanemiainNorthAmericais2 casespermillionpopulation. Aplasticanemiaismuchmore frequentlyseeninthe Orient(6casespermillionpopulationinThailand;14casesper millionpopulationinJapan). The male-to-female ratioinacquiredaplasticanemiaisapproximately1:1. Aplasticanemiaoccursinall age groups,withtwopeaksof incidence.One isobservedinpeople aged20-25 years;the subsequentpeakisobservedinpeople olderthan60 years.
  5. 5. 5. Aplastic Anemia: Pathology The bone marrow inaplasticanemiashowsvariablyreducedcellularity,dependingonthe clinical stage of the disease. There isa decrease inthe numberof cellsof myeloid,erythroid,andmegakaryocyticlineage,witharelative increasein lymphocytesandplasmacells.Asthe cellularitydecreases,thereisacorrespondingincreaseinbone marrowfat. Anemia,leukopenia(primarilygranulocytopenia) andthrombocytopeniacharacterize aplasticanemia.Circulatingred cellshave a normal shape butare oftenmildlymacrocytic.Despite elevatederythropoietinlevels,reticulocytosisisnot present.Increasedfetalhemoglobincanbe demonstratedinthe redcellsinsome cases.Asinthe bone marrow,a relative lymphocytosisisobservedinthe blood. 6. Aplastic Anemia: Laboratory Studies Peripheral bloodsmearswillapaucityof platelets, redbloodcells,granulocytesandmonocytes,butwithnormal red bloodcell morphology(incontrasttohemolyticanemiasandsickle cell disease).The reticulocytecountislow,lessthan 0.5% of redbloodcells(normal is0.5%-1.5%). Bone marrow aspirationsandbiopsyare assessedqualitatively(bone marrow aspiration) andquantitatively(bone marrow biopsy) forcellularityandmorphologyof residualcells.  To assessbone marrowcellularity,the percentage of marrow space occupiedbyhematopoieticelementsplus stroma isvisuallyestimatedinbone marrow biopsy.  The reference age formarrowcellularitydecreaseswithage (from90% ininfantsto50% inthe elderly) andso the value mustbe interpretedinlightof the patient’sage.  In aplasticanemiaboththe bone marrow aspirationandbiopsyspecimenare hypocellularoracellular.The disease isconsideredsevere if the bone marrow cellularityisreducedlessthan30%.  Pathological examinationof bone marrow biopsyspecimenshowsfatcell replacementof the bone marrow. 7. Normal Bone MarrowBiopsy Specimen: LowMagnification A bone marrow biopsyof a normal specimen(above) will normallyshow approximately50% to 60% of the marrow space as hematopoieticelements;the fatcellsmake up the remaining40%.These percentagesare age- dependent,the proportionof marrow fatincreasingwith age.This biopsysectionisstainedwithhematoxylinand eosin,whichallowsrecognitionof the differentcell lineagespresente.g.,erythroid,granulocyticand megakaryocytic.
  6. 6. 8. Aplastic Anemia Bone MarrowBiopsy:Low & HighMagnification Thisinteractive image depictsabone marrowbiopsy specimeninaplasticanemia.Atlow magnification (10x) the marrow space isseentobe virtuallydevoid of hematopoieticelements.The highermagnification image (40x) corroboratesthe hypocellularity(<5%) of the marrow.The residual cellularcompositionincludes some myeloidelements,plasmacells,and lymphocytes. 9. Aplastic Anemia: Peripheral BloodSmear The hypocellularityof the marrow isreflectedin the peripheral bloodsmear,whichshows granulocytopenia(the nucleatedcellshownisa lymphocyte) andleukopenia.Normally,atthis magnification(about40x) youshouldbe able to identify5to 7 leukocytes.Alsonotice thatthere isample space betweenredcells,areflectionof thispatient'sanemia;andthere are few,if any, platelets.Allthesefeaturesreflectthe suppressionof formationof all the cell linesinthe marrow. 10. Aplastic Anemia: Clinical Features The clinical presentationof aplasticanemiaincludessymptomsrelatedtothe decrease inbone marrow productionof hematopoieticcells.The onsetisinsidious,withthe initial symptomrelatingtoanemiaorbleedingorneutropenia. Anemiamaymanifestaspallor,dyspnea(breathingdifficulty)orfatigue.Thrombocytopeniamaypresentasmucosal or gingival bleedingorskinbruises.Neutropeniamaymanifestasovertorrecurrentinfection.Levelsof serum erythropoietinare elevatedproportionallytothe degree of anemia.
  7. 7. 11. Aplastic Anemia: Treatment Transfusion:Patientswithaplasticanemiarequiretransfusionsupportuntilthe diagnosisisestablishedandspecific therapycan be instituted. Treatmentof infectionswithbroad-basedempirical antibiotictherapy. Bone marrow transplantationfromaHLA-matchedsiblingdonoristhe treatmentof choice forpatientslessthan60 yearsold.Immunosuppressive therapyisusedif the patientisolderthan60 years. 12. Aplastic Anemia: Prognosis Aplasticanemiahasgreaterthana 75% mortalityrate withsupportive care alone.Itrepresents ahematological emergencyandcare shouldbe institutedpromptly. The 5-year survival rate forthe patientsreceivingmatchedsiblingdonorbone marrow transplantationisgreaterthan 90%. The 5-yearsurvival rate forpatientsreceivingonlyimmunosuppressive therapyis75%. 13. Which ofthis patient's drugs is most likely to have causedher aplastic anemia? Phenylbutazone,anonsteroidalantiinflammatorydrug(NSAID) usedforthe treatmentof painof osteoarthritis,is probablythe culprit. Phenylbutazone isawell knowncause of aplasticanemia,andthe riskisgreaterwhen phenylbutazone istakenregularlyandfora sustainedperiod. 14. Anemia ofRenal Disease Anemiaof chronicrenal insufficiencyreflectsdecreasedproductionof erythropoietinbythe damagedkidneys.The severityof anemiaisproportionaltothe underlyingdegreeof renal insufficiency. In additiontorenal disease (fromvaryingcauses) causingdecreasedproductionof erythropoietinandsubsequent developmentof anemia,ithasbeensuggestedthatthere isa“uremictoxin,”capable of suppressingerythroid precursors,as well asa minorhemolyticcomponentproduced.These have beenimplicatedascontributingtothe anemiaof chronicrenal disease,butthe proof of thishypothesisislacking. The anemiaof chronic renal disease isnormocyticandnormochromic.Insome cases,echinocytes(crenatedcells,burr cells) withscallopedcell membranescanbe seen.If the renal insufficiencyissecondarytomalignanthypertension,red cell fragmentationwithformationof schistocytesmayalsobe seenonsmears. Administrationof recombinanterythropoietinisthe treatmentof choice.
  8. 8. 15. Anemia ofRenal Disease:Peripheral BloodSmear,Medium Magnification The anemiaof renal failure resultsdirectlyfroma deficiencyinerythropoietinproduction,makingitunique amongthe manytypesof anemia.Because of the decreasederythropoietin,there isdecreasedproliferation of erythroidprecursorsinthe marrow.Thus,anemiaof renal failure fallsintothe broadcategoryof hypoproliferative anemias.The anemiaisgenerally normochromic/normocyticwithlittle anisopoikilocytosis, as illustratedinthe above image. 16. Anemia ofChronic Disease: Pathology & Abnormalities ofIronDistribution Anemiaof chronicdisease arisesin associationwithchronicinflammatoryand malignantconditions.The pathogenesisof nnemiaof chronicdisease resultsfromthe release of inflammatorycytokines(IL-1, TNF,IFN-gamma) ininflammatoryor neoplasticprocesses.Itisthoughtthat these cytokinessuppresserythropoiesis throughmultiple mechanisms. Thus,anemiaof chronic disease representsanotherhypoproliferative anemia.The cytokinesalsocause abnormalitiesof ironmetabolism,includingincreasedstorage iron(secondfeature) anddecreasedserumiron.These abnormalitiesof irondistributionare generallynotfelttobe asignificantcause of the anemiainthisdisorder,butare useful as laboratorymarkers.The increased storage ironisreflectedasincreasedserumferritin;thus,increasedserumferritinin conjunctionwithdecreasedserumironischaracteristicof anemiaof chronicdisease. The red cellsinanemiaof chronicdisease are reducedin numberandmay be normocyticas shownabove left,or mildlymicrocyticandminimallyhypochromicasshownat right.The anemiaof chronicdisease ismildtomoderate, and the redcellsare oftenmicrocytic.Prussianblue staining(left)demonstratesnormal oroftenincreased amountsof storage iron.Serumiron levelstendtobe reduced. However,incontrastto irondeficiencyanemia,total iron bindingcapacityalsotendstobe decreased(asisthe serumalbuminlevel).Successful treatmentof the underlyingchronicdisease isassociatedwithrestorationof normal hemoglobinlevels.The cytokinesalsocause abnormalitiesof ironmetabolism, includingincreased
  9. 9. storage ironsuch inthisimage of bone marrowaspirationanddecreasedserumiron.These abnormalitiesof iron distributionare generallynotfelttobe a significantcause of the anemiaof chronicdisease,butare useful aslaboratory markers.The increasedstorage ironisreflectedasincreasedserumferritin;thus,increasedserumferritininconjunction withdecreasedserumironischaracteristicof anemiaof chronicdisease. 17. Myelophthisic Anemia - Anemia Associatedwith Bone MarrowInfiltration Myelophthisicanemiareferstohypoproliferative anemiaassociatedwithinfiltrationof the bone marrow bya varietyof processes. Pathogenesis:Anyinfiltrativeprocess(suchasmyelofibrosis,hematological malignancies,metastaticcarcinoma,or granulomatousdisease) canreplace normal hematopoieticelementsandcause anemia(andoftenleukopeniaand thrombocytopenia).Inanattempttomaintainbloodcell production,foci of extramedullaryhematopoiesismayappear, mostprominentlyinthe spleenandliver. Pathology:Bone marrowinfiltrationresultsinmoderate toseverenormocyticanemia,withanisopoikilocytosisand teardropcells.Circulatingimmature granulocytesandnucleatedredbloodcells(leukoerythroblastosis) are frequently identified.
  10. 10. Quiz 1) Which of the followingfeaturesinthe CBCcountdoesnotagree witha diagnosisof aplasticanemia? A) Neutropenia B) Normocyticanemia C) Microcyticanemia D) Thrombocytopenia E) Lack of reticulocytosis 2) The diagnosisof aplasticanemiacanbe made withoutabone marrow biopsyif the CBC count showssevere pancytopenia. A) True B) False 3) A 35-year-oldmanpresentswitha2-monthhistoryof headache andwas notedtobe pale bya friend.On examination,petechia,hepatosplenomegalyandsupraclavicularadenopathyare present.The CBCcountconfirms severe pancytopenia.Aplasticanemiaisthe likelydiagnosis. A) True B) False 4) Which of the followingisneitheracomplicationof aplasticanemianeitheraconsequenceof pancytopenia? A) Thrombosis B) Pallor C) Sepsis D) Tirednessandmalaise E) Ecchymoses 5) A 35-year-oldmanpresentswithpetechia,ecchymoses,fatigue,andalow-grade fever.Hisphysical examinationis otherwise unremarkable.A completebloodcellcountrevealsamoderatelysevere normocyticanemia, thrombocytopenia,andabsolute neutropenia.The reticulocyte countislessthan0.5 %. The serumbloodureanitrogen and creatinine are normal.The nextstepinthe managementof thispatientistoorderwhichone of the followingtests? A) Direct Coombstest B) Sugar waterhemolysistest C) SerumvitaminB12 level measurement D) Bone marrowaspirate and biopsy E) Serumironand ferritintest

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