Rev. Bras. Hematol. Hemoter. vol.38 número1

rev bras hematol hemoter. 2016;38(1):88–89

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

Revista

Brasileira

de

Hematologia

e

Hemoterapia

Brazilian

Journal

of

Hematology

and

Hemotherapy

Letter

to

the

Editor

What

is

the

adequate

mononuclear

cell

content

for

selecting

umbilical

cord

blood

units

for

cryopreservation?

Overthe past decade umbilicalcord blood (UCB) hasbeen used as an alternative source of hematopoietic stem cells (HSCs).Worldwide,morethan500000cordbloodunits(CBUs) havebeencryopreservedand storedinbanks.1 _Ithasbeen

shown thatthe CD34+ _{cell content influencesengraftment}

andsurvivalafterUBCtransplants.2 _{Althoughthereareno}

fullyagreeduponstandardizedcriteriaforselectingunitsfor cryopreservation,mostcordbloodbanksuseacombinationof thetotalnucleatedcell(TNC)countandvolume;aTNCcount

≥8×108 _{hasbeen considered the best predictor for CD34}+

HSCcontent.3_{Nosuchparameterhasbeenestablishedforthe}

mononuclearcellcount(MNC).

Recentlyithasbeenproposedthatinordertoassurethat stemcellswillperformasintended,thequalityandpotency ofUCBhematopoieticprogenitorcellsshouldbemeasured. ThiscanonlybedoneusingtheMNCfraction,nottheTNC fraction.4,5 _{Giventhisrecentdevelopment, weassessedthe}

possibilityofusingtheMNCtopredicttheCD34+_cellcontent

incordbloodanditsrelationshiptotheTNCcount.

Weanalyzed 857CBUs received atthe cordblood bank oftheHematologyDepartmentoftheHospitalUniversitario ‘Dr.JoseE.Gonzalez’,UniversidadAutónomadeNuevoLeón inMonterrey,México.TNCcountsweredeterminedwithan automatedhematologyanalyzer (SysmexXT 2000,Sysmex, Mundelein,IL,USA)usingamultianglepolarizedscatter sepa-rationtechniquewhichprovidestheprimaryTNCcount.Total TNCcountswerecalculatedbymultiplyingtheprimaryTNC count(permicroliter)bythe totalvolumeoftheCBU.Total MNCcountswerecalculatedbyaddingtheabsolute lympho-cyte and monocyte counts(per microliter)reported in the completebloodcountandthenmultiplyingthisvaluebythe totalvolumeofthebag.CD34+_{cellcountsweredetermined}

byemployingfluoresceinisothiocyanatelabeled anti-CD45+

monoclonal antibodies (FITC; Becton Dickinson, San Jose, CA,USA)andmonoclonalanti-humanCD34+_{ClassIII/FITC,}

Clone BIRMA K-3 (DAKO, Denmark) in a FACSCalibur flow cytometer (Becton Dickinson).The mean collected volume oftheCBUswas 96.98±28.88mLandthe meanTNC, MNC andCD34+_{countswere9.95±4.97×10}8_{,5.10±2.65×10}8_and

2.95±2.33×106_{,respectively.WedividedtheCBUsinto two}

groupsdependingontheCD34+_{cellcount(lessthanorgreater}

than 2×106_{). Receiver operating characteristic(ROC) curve}

analysiscomparingTNCandMNCtoselectCBUswithaCD34+

cellcountof≥2×106_{weredesigned.Theoptimaloperating}

pointsforTNCandMNCthatbestcorrelated withaCD34+

count ≥2×106 _{were determined (Table 1; Figure 1). A}

sig-nificant difference between TNC and MNC was not found (p-value=0.059).Inaddition,aTNC>7.94×108_wasselected

byROCanalysisasthepointthatbestreflectstheMNC guar-anteeingaCD34+_{count≥2×10}6_(Table1).

Previously,weappliedaROCmodeltoanalyzethe correla-tionbetweenvolume,TNCandMNCwiththeCD34+ _count,

and established the TNC content as the most significant parameter.3_{Inthepresentreport,weevaluatedthedegreeto}

100

80

60

40

Sensitivity

20

0

0 20 40 60 80

100-specificity

100

MNC count TNC count

Figure1–Receiveroperatingcharacteristic(ROC)curve analysiscomparingthetotalnucleatedcell(TNC)count [areaunderthecurve(AUC)=0.822]vs.theabsolute mononuclearcell(MNC)count(AUC=0.806)ascriteriafor selectingcordbloodunitssuitableforcryopreservation, thatis,aCD34+_{cellcontent≥2.0×10}6_{.Nosignificant}

revbrashematolhemoter.2016;38(1):88–89

89

Table1–Acomparisonbetweentotalnucleatedcellsandmononuclearcellsfortheselectionofcordbloodunitswith CD34+_{counts≥2×10}6_.

Performanceevaluation ComparisonofROCcurves

Sensitivity Specificity −PV +PV +LR −LR AUC Difference

betweenareas

p-Value

TNCcutoff(×108_{) 0.822}

>8.33 77.86 69.88 67.70 79.60 2.59 0.32

>7.94* _{80.97 64.33 69.20 77.40 2.27 0.30}

MNCcutoff(×108_{) 0.806}

>4.05 80.78 66.37 69.60 78.30 2.40 0.29 0.0159 0.059

TNC:totalnucleatedcell;MNC:mononuclearcell;ROC:receiveroperatingcharacteristic;−PV:negativepredictivevalue;+PV:positivepredictive value;+LR:positivelikelihoodratio;−LR:negativelikelihoodratio;AUC:areaunderthecurve.

∗ _{OptimaloperatingpointchosenbyROCanalysisforTNC.}

whichaTNCcount≥8×108_{reflectsaMNCcontenttomeet}

therequiredCD34+_{countandfoundthatthemeanMNCcount}

was5.10×108_{(63.75%ofthemeanTNCcount).Moreover,ROC}

analysisselectedaMNCof4.05×108 _{asthecutoffpointto}

achieveanoptimalCD34+_{count(≥2×10}6_).

Thus,ourfindingsconfirmthatthecurrentTNCstandard forUCB cryopreservationgives reliable informationon the MNCcontentandsuggestsacutoffpointforthelowest frac-tionfortransplantationpurposes.

Conflicts

of

interest

Theauthorsdeclarenoconflictsofinterest.

Acknowledgement

WethankSergioLozano-RodriguezM.D.forhisreviewofthe manuscript.

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1.MazzocchettiD,BertiAM,SartiniR,LucariniA,RagusaG, CaroliM,etal.Totalnucleatedcellsasasolepredictorof distincttargetsofhematopoieticpotential(CD34+_cells)incord bloodunits:theresultsofalargeseriesanalysisinautologous cordbloodunits.Transfusion.2014;54(5):1256–62.

2.SolvesP,Carbonell-UberosF,MirabetV,RoigR.CD34+_cell contentforselectingumbilicalcordbloodunitsfor cryopreservation.Transfusion.2007;47(3):552–3.

3.Jaime-PerezJC,Monreal-RoblesR,Rodriguez-RomoLN, Mancias-GuerraC,Herrera-GarzaJL,Gomez-AlmaguerD. Evaluationofvolumeandtotalnucleatedcellcountascord bloodselectionparameters:areceiveroperatingcharacteristic curvemodelingapproach.AmJClinPathol.2011;136(5): 721–6.

4.PattersonJ,MooreCH,PalserE,HearnJC,DumitruD,Harper HA,etal.Detectingprimitivehematopoieticstemcellsintotal nucleatedandmononuclearcellfractionsfromumbilicalcord bloodsegmentsandunits.JTranslMed.2015;13:94.

5.RichIN.Improvingqualityandpotencytestingforumbilical cordblood:anewperspective.StemCellsTranslMed. 2015;4(9):967–73.

JoséCarlosJaime-Pérez∗_{,GiselaGarcía-Arellano,}

AlejandraCelinaEsparza-Sandoval

UniversidadAutónomadeNuevoLeón,Monterrey,Mexico

∗_{Corresponding author at: Servicio de Hematologia, Edificio}

“Dr.RodrigoBarragan”,2◦_{piso,HospitalUniversitario“Dr.Jose}

E.Gonzalez”,Av.MaderoyGonzalitosS/N,ColoniaMitras Cen-tro,Monterrey,NuevoLeonC.P.64460,Mexico.

E-mailaddress:carjaime@hotmail.com(J.C.Jaime-Pérez).

Received18December2015 Accepted9January2016 Availableonline9February2016

http://dx.doi.org/10.1016/j.bjhh.2016.01.003