Rev. Bras. Hematol. Hemoter. vol.37 número5

rev bras hematol hemoter. 2015;37(5):287–289

w w w . r b h h . o r g

Revista

Brasileira

de

Hematologia

e

Hemoterapia

Brazilian

Journal

of

Hematology

and

Hemotherapy

Scientific

Comment

The

importance

of

hemoglobin

A

2

determination

夽

Maria

Stella

Figueiredo

UniversidadeFederaldeSãoPaulo(UNIFESP),SãoPaulo,SP,Brazil

a

r

t

i

c

l

e

i

n

f

o

Articlehistory: Received24June2015 Accepted7July2015 Availableonline21July2015

Hemoglobin(Hb)A2(␣2␦2)constituteslessthan3%ofthetotal

hemoglobin(Hb)inadultsand hasalmostnophysiological importance.1_{Ontheotherhand,thedeterminationofHbA}

2

isanimportanttooltodiagnosethe beta-thalassemia trait (BTT).1,2 _{Althoughindividuals with BTTdonotneed} treat-ment,theaccuratedetectionofthecarrierstateisimportant ingeneticcounselingtodetermineriskofhavingachildwith amajordisease.3

ElevatedlevelsofHb A2andmicrocytosis aresuggestive

ofthediagnosisofBTT.However,BTTmaybepresent with normallevelsofHbA2 asafewcasesof␤-thalassemiaare

notassociatedwithelevatedHbA2,andbecauseofthe

asso-ciation of BTT with iron deficiency or with ␣-thalassemia (␣-Thal).1,2,4–6 _{There are many other factors, inherited or} acquired,thatcaninterfereinHbA2levels(Table1).3,4

HbA2canbemeasuredbyseverallaboratorialmethods,but

thesemethodshavedifferencesinaccuracy.7_{Cationexchange} high performanceliquid chromatography(HPLC), microcol-umnchromatography,andcelluloseacetateelectrophoresis withelutionareconsideredacceptablemethodstodiagnose BTT,whereas celluloseacetate electrophoresis followed by scanning densitometry is not.2 _{The accuracy of cellulose} acetateelectrophoresis withelution dependson the train-ingandexperienceofthelaboratorytechnicianwhoperforms

夽

SeepaperbyFonsecaetal.onpages296–301.

∗ _{Correspondenceto: Hematology andBlood TransfusionDivision, Escola Paulista de Medicina,UniversidadeFederal de SãoPaulo} (UNIFESP),RuaDr.DiogodeFaria,824,0437-002SãoPaulo,SP,Brazil.

E-mailaddress:stella.figueiredo@unifesp.br

thetest,andmicrocolumnchromatographycangiveproblems withco-elutionofsomeHbvariants.7

Recentstudieshaveconfirmedthehigherqualityof auto-matedHPLCinthemeasurementofHbA2comparedtothe

othermethods,8,9_{whichiswhythishasbecomethemethod} ofchoice.Ontheotherhand,inautomatedHPLC,the mea-surementofHbA2 isinaccuratewhenHbSispresent.2,3,10

AstheamountofHbSisrelatedtothedegreeofinaccuracy, levelsarehigherinpatientswithsicklecellanemia(SCA)or HbS/␤-thalassemia(S-␤Thal)thaninsicklecelltrait.2_Thus, theamountofHbA2doesnotindicateBTTwhenHbAand

betagenevariantsarefoundtogether.11_{Furthermore,when} betagenevariantsarepresentwithoutHbA,thediagnosisof concomitantBTTisnotnecessarilyassociatedtotheelevation ofHbA2andsofurtherinvestigationsusingfamilystudiesor

DNAanalysisarenecessary.10

Asmentionedabove,␣-Thaliscapableofinterferinginthe determinationofHbA2.4Individualswiththe␣0-thalassemia

traitorhomozygousfor␣+-thalassemiahavelowerlevelsof HbA2,buttheinfluenceofthecoinheritanceof␣-Thal and

BTTonHbA2levelsisuncertain.6

InBrazil,theincidenceof␣-Thalvariesfrom0.11to0.22% dependingonthegeographicalregionstudied.12–15_Itiswell knownthattheassociationof␣-ThalandSCAiscommonin

http://dx.doi.org/10.1016/j.bjhh.2015.06.002

288

Table1–Causesofvariationinthepercentageof hemoglobinA2.

HbA2 Inherited Acquired

Increased (>3.4%)

␤-Thalassemia heterozygosity DeletionalHPFHfrom Vietnamese/SouthEast Asian

HereditaryhighHbA2

Unstablehemoglobin Sicklecelltrait Sicklecellanemia (particularlycoexisting

␣-thalassemia) HbS/␤0-thalassemia Congenital

dyserythropoieticanemia (somecases)

Heterozygosityforother

␤-chainvariants

Thyrotoxicosis HIVinfection Zidovudinetherapy Megaloblastic anemia(somecases)

Decreased (<2.2%)

␣-thalassemia:␣+

homozygosity,a0

heterozygosity,andHbH disease

DeletionalHPFH(except Vietnamese/SouthEast Asiantype)

␦␤andA_{␥␦␤-thalassemia}

heterozygosity(somecases)

␦-Thalassemia HemoglobinLepore HemoglobinKenya

Severeiron deficiency Anemiaofchronic disease Sideroblasticanemia Leadpoisoning Juvenile myelomonocytic leukemia AcquiredHbH disease Acutemyeloid leukemia(some cases,particularly erythroleukemia) Aplasticanemia (somecases) Hypothyroidism Chemotherapy-inducedincreased HbFsynthesis

Source:ModifiedfromBainetal.4

Hb A2 – hemoglobin A2; HPFH: hereditary persistence of

fetal hemoglobin; HIV: human immunodeficiency virus; HbH: hemoglobinH;HbF:hemoglobinForfetalhemoglobin.

Brazil.16–18_{SinceSCAisconsideredapublichealthproblemin} Brazilandduetotheclinicalsignificanceof␣-Thalinrespect tothisanemia,diagnosisisimportant.19–21_{However,diagnosis} ismainlyachievedbymoleculartechniquesthatareexpensive andnoteasilyaccessible.Itisalsoimportanttoremember thattheco-inheritanceof␣-ThalandSCAresultsinincreased levelsofHbA2 asmeasuredbyautomatedHPLC,andcould

resultinamisdiagnosisofS-␤Thal.4

There lies the importance of the paper entitled “Hemoglobin A2 values quantified by high performance

liquid chromatography inpatients with sickle cell disease, and the influence of the presence of alpha-thalassemia” written by Fonseca et al. and published in this edition of theRevistaBrasileiradeHematologiaeHemoterapia.22 _The authorsdemonstratethatHbA2wasoverestimatednotonly

inindividualswithHbSbutalsoinpatientswithHbC,and thattheHbA2levelwasinfluencedbythegenotypeof␣-Thal.

In conclusion, ina country with a high degree of mis-cegenation suchasBrazil,notonlythediagnosis ofdouble heterozygousstates,suchasS-␤Thal,butalsothediagnosisof co-inheritanceofSCAwith␣-Thalshouldbecarriedout care-fully,takingintoconsiderationthelimitationsoftheavailable laboratorytechniques.FamilystudiesorDNAanalysis,when possible,aredesirabletoconfirmthecorrectdiagnosis.

Conflicts

of

interest

Theauthorsdeclarenoconflictsofinterest.

r

e

f

e

r

e

n

c

e

s

1.SteinbergMH,AdamsJG.HemoglobinA2:origin,evolution,

andaftermath.Blood.1991;78(9):2165–77.

2.HeadCE,ConroyM,JarvisM,PhelanL,BainBJ.Some observationsonthemeasurementofhaemoglobinA2andS

percentagesbyhighperformanceliquidchromatographyin thepresenceandabsenceofalphathalassaemia.JClin Pathol.2004;57(3):276–80.

3.GiambonaA,PassarelloC,RendaD,MaggioA.The significanceofthehemoglobinA(2)valueinscreeningfor hemoglobinopathies.ClinBiochem.2009;42(18):1786–96.

4.BainBJ,WildBJ,StephensAD,PhelanLA.Variant

hemoglobins:aguidetoidentification.1sted.WestSussex, UK:Wiley-Blackwell;2010.

5.Saleh-GohariN,KhademiBamiM,NikbakhtR,Karimi-Maleh H.Effectsofalpha-thalassaemiamutationsonthe

haematologicalparametersofbeta-thalassaemiacarriers.J ClinPathol.2015;68(7):562–6.

6.DenicS,AgarwalMM,AlDabbaghB,ElEssaA,TakalaM, ShowqiS,etal.HemoglobinA2loweredbyirondeficiencyand

alpha-thalassemia:shouldscreeningrecommendationfor beta-thalassemiachange?ISRNHematol.2013;2013:858294.

7.ColahRB,SurveR,SawantP,D’SouzaE,ItaliaK,

PhanasgaonkarS,etal.HPLCstudiesinhemoglobinopathies. IndianJPediatr.2007;74(7):657–62.

8.AnagnostopoulosK,TentesI,KalleasC,MargaritisD,ToliA, PendilasD,etal.EffectofHbSinthedeterminationofHbA2

withtheMenariniHA-8160analyzerandcomparisonwith otherinstruments.IntJLabHematol.2009;31(6):665–72.

9.PaleariR,GulbisB,CottonF,MoscaA.Interlaboratory comparisonofcurrenthigh-performancemethodsforHbA2.

IntJLabHematol.2012;34(4):362–8.

10.ShokraniM,TerrellF,TurnerEA,AguinagaMD.

ChromatographicmeasurementsofhemoglobinA2inblood

samplesthatcontainsicklehemoglobin.AnnClinLabSci. 2000;30(2):191–4.

11.GiordanoPC.Editorial:measurementofHbA2.IntJLab

Hematol.2012;34(4):335.

12.SilvaCdeA,BaldimLB,NhoncanseGC,EstevaoIdaF,Melo DG.NeonatalscreeningforhemoglobinopathiesinSao Carlos,SaoPaulo,Brazil:analysisofaseriesofcases.RevPaul Pediatr.2015;33(1):19–27.

13.deMedeirosAlcoforadoGH,BezerraCM,AraújoMouraLemos TM,deOliveiraDM,KimuraEM,FerreiraCostaF,etal. Prevalenceofalpha-thalassemia3.7kbdeletionintheadult populationofRioGrandedoNorte,Brazil.GenetMolBiol. 2012;35(3):594–8.

revbrashematolhemoter.2015;37(5):287–289

289

determinantsinsouthBrazil:importanceforthediagnosisof microcyticanemia.GenetMolBiol.2010;33(4):641–5.

15.AdornoEV,CoutoFD,MouraNetoJP,MenezesJF,RêgoM,Reis MG,etal.HemoglobinopathiesinnewbornsfromSalvador, Bahia,NortheastBrazil.CadSaudePublica.2005;21(1): 292–8.

16.LyraIM,Gonc¸alvesMS,BragaJA,GesteiraMdeF,CarvalhoMH, SaadST,etal.Clinical,hematological,andmolecular characterizationofsicklecellanemiapediatricpatientsfrom twodifferentcitiesinBrazil.CadSaudePublica.

2005;21(4):1287–90.

17.FigueiredoMS,KerbauyJ,Gonc¸alvesMS,ArrudaVR,SaadST, SonatiMF,etal.Effectofalpha-thalassemiaandbeta-globin geneclusterhaplotypesonthehematologicalandclinical featuresofsickle-cellanemiainBrazil.AmJHematol. 1996;53(2):72–6.

18.DeLemosCardosoG,GuerreiroJF.Molecularcharacterization ofsicklecellanemiaintheNorthernBrazilianstateofPara. AmJHumBiol.2010;22(5):573–7.

19.dePaivaeSilvaRB,RamalhoAS,CassorlaRM.Sicklecell diseaseasapublichealthprobleminBrazil.RevSaude Publica.1993;27(1):54–8.

20.DomingosIF,FalcãoDA,HatzlhoferBL,CunhaAF,SantosMN, AlbuquerqueDM,etal.Influenceofthebetashaplotypeand alpha-thalassemiaonstrokedevelopmentinaBrazilian populationwithsicklecellanaemia.AnnHematol. 2014;93(7):1123–9.

21.Camilo-AraujoRF,AmancioOM,FigueiredoMS, Cabanas-PedroAC,BragaJA.Molecularanalysisand associationwithclinicalandlaboratorymanifestationsin childrenwithsicklecellanemia.RevBrasHematolHemoter. 2014;36(5):334–9.

22.FonsecaSF,AmorimT,Purificac¸ãoA,GoncalvesMS,Boa-Sorte N.HemoglobinA2valuesquantifiedbyhighperformance