j.1365-2249.1994.tb07015.x - copia

  • Upload
    rrokel

  • View
    215

  • Download
    0

Embed Size (px)

Citation preview

  • 8/13/2019 j.1365-2249.1994.tb07015.x - copia

    1/7

    Clin Exp Immunol 1994; 95:436-441

    Selective increase of activation antigens HLA-DR and CD38 onCD45RO+ T lymphocytes during HIV-1 infectionL, KESTENS. G. VANHAM, C. VEREECKEN. M. VANDENBRUAENE*t . G. VERCAUTEREN*,R . L . C O L E B U N D E R S * t & P . L , G I G A S E Lah oralory of Pathology & Immuno logy. *M icrohiohgy and

    fOtilpiilient Clinic. Institute of T ropical Medicine. Antwerpen. B elgium

    (Acceptedfor publication 5 November 1993)

    S U M M A R YInfection with HIV results in a progressive depletion of CD4* T cells and leads to significant in vivolymphocyte phenotype changes. In this regard, the expression of HLA-DR and CD38 on CD8^ Tceils has been shown to increase dramatically with disease progression. We investigated theexpression of both aetiva tion ma rker s on CD4+ T cells in HIV-1-infected subjeets at diliferent clinicalstages of infection and compared the in vivoactivation of CD4 ^ T cells with param eters of viralactivity and CD8^ T cell activation. Eresh peripheral venous blood was obtained from 54 HIV-infeeted subjects and from 28 uninfeeted healthy con trols. Three -colour im mu noph enoty ping of theC D 4 ' Tee ll subset showed that the prop ortion of C D 4 ' T cells expressing HL A-D R (10% in HIV -negativecontr ols) or CD 38 (62"/l. inHIV-negati ve controls) was higher in a symp tomatic (P

  • 8/13/2019 j.1365-2249.1994.tb07015.x - copia

    2/7

    HIV-1 infection and CD4^ T cell activationTable I . Study populat ion, anii-relroviral i rf i i lmeiu. HIV antigen and CD4 count

    437

    Male/femaleMed ian age (years)Received anti-ret rov iral treatmentHIV antigen*positiveTotal lymphocyte count* (celis/mm')CD4 percentage*Tola CD4 count* (cells;i iim')

    H IV (/j = 28)NAN An7X (i 163-1723)46(40-50)635(476 774)

    W HO I (/i = 27)21/636

    6 (22'H,)2 ( 7 % )

    1957(1644 2489)24 (IS 33)

    459 (307 637)

    W H O 2 +3 (/ = 15)14/143

    7 (4 7 % )9 (60%)

    1290(906-1768)14(7 32)199(6 400)

    W H O 4 ( H = I 2 )1 /I4212(100%)7 (58%)IO27(93-I5(H))8(5-6-13)104(45 208)

    * Results are expressed as median an d interqua rtile ranges (p25 p75).NA . No available.

    between li) /.) and 30H;, and with more ihan 30' /,, were H IV ' subjects were clinically classified ac cord ing to

    haden opathy (W HO stage 1). 15 were symp tomatic non -2+3) . and 12 were AIDS patients

    // (r /i (' C D 4' T cell activation was examined by three-colourunop heno typing using peridinyl chlorophyll-A proteinof FITC-conjtigated anti HL A-D R and an ti-CD 45R O

    Leu-2a-FITC) in combinat ion with ant i-H LA -DR -FIT C

    The lymphocyte staining was performed according to theThe stained samples were stored at 4 C and analysed using aan flow cytometer witb LYSYS software within 24 hve gate was set aro und the C D 4 ' cluster inr to acqu ire a min imum of IOOO CD 4* cells dur ing cell

    of total HfV-l antigenHIV 1 antigen

    with 50 /d I N HCl and in cub ated for I h atples were then neutralized with 50 ;/l I N Na OH

    on of .soluble CDH antif^en

    stored at 20 C was thawed slowly and 5 /d were assayedundiluted. C'D8 antigen reference standards provided In the kitwere assayed in parallel to generate a standard curve of units ofsCD8 per ml serum.

    Statistical amily.\isDifferences between groups were tested i\ir statistical signifi-cance using the non-parametric Wilcoxon two-sample test.Where appropriate, P values were corrected applying theBonferroni adjustment for multiple comparisons. For eachcombination of activation antigens studied on CD4 ' Tcells, tbeP value was multiplied by 4 Utadjust lor com parin g results ofthe four dependent fiuorescenee qua dra nts. The 9 5% confidenceintervals of the median were calculated using the formula:l-56(p75-p25)/SORT(,V) in which p75 and p25 represent the75th and the 25th perccntile about the median and A'the numberol 'subjects [15]. Cor relatio n analysis was performed with theSpearman's rank-correlation test.

    RESULTSSubjects ami treatmentThe male to female ratio, the median age. the use of anti-retroviral treatment (zidovudine), the presence o[ HIV antigenin serum, the lotal lymphocyte count and the relative andabsolute C D 4 ' T cell coun ts are presented in Table 1.Three-colour imtniinopheno/ypin^ of CD4 cellsHLA-DR expression was generally low in HIV-negalive con-trols (median 104%). bul 50"/,, or more of the CD 4' T cellsexpressed CD38 (median 62%) or CD45RO (median 54';.>) (Fig.1). All H I V groups showed a relative increase of HLA -D Rand CD38 expression on CD 4' Tcel ls compared with controls.HLA-DR and CD38 expression increased from WHO stage I tosubsequent WHO disease stages. Conversely, the expression ofCD 45 RO o n C D 4 ' T cells did not differ significantly betw eenHIV-infected subjects and controls, and was only slightlyincreased in patients from tbe WHO stages 2, 3 and 4 {Fig. 1).

    In HIV-negative control subjects, mosl CD4' T cellsexpressed CD38 but no HLA-DR (median 59'^) or werenegative for bolh markers (median 34".,). whereas only a smallproport ion of CD 4 ' T cel ls were CD38 ' HL A-D R *(median23%) or CD38 HLA-DR* (median 38%) {Fig. 2a). Theproport ion ofC D 4 ' Tcells thai expressed CD 38 alone In HIV*

  • 8/13/2019 j.1365-2249.1994.tb07015.x - copia

    3/7

    43 8 L. Kestens et al.too

    80

    6 0

    4 0

    20

    0

    * *

    WHOI WH02+3 WH04Fig. I. Proporli i in o r C D 4 ' T ucil.s ihal expresses HLA-D R (A ).CD45R(1 (V) or C'D.'IS (O) antigen in rekition lo ihc cliniciil stage ofMIV inTection. H I V ( - ) . Unintccted cnn tml s; W H O I. ji.syinplonKiticHIV infeclion; W H 02 + 1 illV-infccled subjects with non-AIDSdefining signs or symptoms: WHO4. AIDS patients. The restilts arepresented as ihe median and the"^5''..confidence intervals of the mediiin.Differences in expression levels were tested for statistical significanceusing the non-param elric Mann Whitney ( -lest. * / '< 0 ' l ) 5 :

    subjects retnaincd unchanged ihroughoui ihc various diseasestages, but a signilicani proportional increase ofCD^X ' HLA-DR* o nCD 4* Tccl l sw asnol iccd . Al though the CD 4' T cel lswith the CD 38 "H LA -D R^ pheno type increased significantly(/ '

  • 8/13/2019 j.1365-2249.1994.tb07015.x - copia

    4/7

    HIV-1 infection ami CD4 ^ T cell activation 4391008 0

    r 604 02 0

    1(b )

    HIV(-) WH01 WH02 3 WH04

    cl

    HIV -1 WHOl WHO2-t-3 WH04 HIV -) WHOl WH O2+ WH04Fit;. 2. Proportio n of CD 4 ' Tc ells ihal cipr css (a) C'D.'IX and IIL A- DR . |b) ilL A- DR :ind C'D45RO, dnd (c) CD.IX and fD 45 R O inrclaiion to llu'cliniual slagc o lH lV inCcL-Uoii, HIV( - ), Uninfcck-d conlrol.s: W H O l. asy mp lom alic HIV inreclion: WH O2 + 1. HIV-mleclcd subjctls wilh rn)n-AIDS defining signs or symptoms: WHO4, AIDS piilients. The rcsulls arc presented as the median andthe 95' - , confidence in terva ls of lhe med ian. Dilleren ces in c.vprcssion levcN were tested Tor stalislical significance using the non -pa r am e t ri c Mann Whi tney fMcsl . *P< 0 ( ) 5 : P < 0 00 5 ; ***f < ( I O ( H . ( a) A . C D 38 ' D R : , C D 3K 'D R ^ T , C D 38 D R ' .( b ) O . D R ' C D 4 5 R O - ; . D R C D 4 5 R O . (c ) O . CD.1H+ C D 45R O ' ; . C D 38 ' C D 45R O V

    HIV*" control Asymp HtV-r Symp. HIV-I + AIDSHIV-1*

    1

    13

    ' I

    . ; , " r - . "--vr-,v..i;;. .

    CD45R0 CD45R0 CD45R0 C045R0

    1CD45R0 CD45R0 CD49R0

    . - " . ' " ' ' . ' . '

    ; v .

    I

    \

    } .

    ' 1 IP lip

    J ' ' " J . ' " '- N* ^

    - I ' - T i r (

    HLA-DR HLA-DR HLA-DR HLA-DRFig. 3. Qua litative phe notyp ic changes on CD 4 ' T cells durin g HIV-1 infeclion. Th e relative increase ol" HL A-D R and C D38 co-expression with disease progression (botto w row) is confined to C D 4 ' T cells expressing lhe CD 45 RO .

  • 8/13/2019 j.1365-2249.1994.tb07015.x - copia

    5/7

    440 L. estens et al.Th eincreasing degree of CD 4' T cell activation on one handand the correlation between ihe activation of C D 4 ' and CDS 'T cells on the other, suggest that the progressive proportionalincrease of HLA-DR and CD38 expression on CD4 ' Teelis co-expressing CD45RO refleets an increasing state of immuneaetivation with disease progression. In this respect, our findingssupport the hypothesis of AIDS as immune system activation

    [17].The proport ion of C D 4' Tcel ls with the CD38 ' HL A-D R ' .C D3 8 ^C D 4 5 R O" and HL A- DR 'C D 4 5 R O' p h en o ty p e d idnot differ significantly in subjects from the various symptomaticdisease stages (WHO 2 and 3 versus WHO 4). This lack ofditlerence eouid have been biased by the elinieal classification

    itself.When these C D 4 ' T cell activation marke rs in symptom a-tic subjects were calculated and classified according to decrea.s-ing CD4'*" T cell percentage intervals, a clear increase wasobserved with decreasing CD4 percentages (data not shown).No selective lossofC D4 5RO or CD 4? RO cells within theC D 4 ' T cell subset was observed vt-ith disease progression.,which is in agreement with previous studies [7-10]. In thisregard, the preferential infection of memory CD4' T cells by

    HIV-1 [18] and th e finding tha t activated but not necessarilyinfected CD 4 ' lymphocytes from HlV-l-infected subjects arethe target of autoreactive eytotoxie T lymphocytes [10,19] arenot reflected in a selective depletion of CD45 RO ' C D 4' Tce lls.Nevertheless, (he CD 45 RO ' subset of CD 4 lytiiphocytes dis-plays predominant proliferative defects and is several timesmore vulnerable to activation-associated lymphocyte deaththan the CD 45R O subset [20]. This parad ox tnight beexplained by a dynamic equilibriutn in which the spontaneouslydying or actively destroyed CD45RO'CD4'^ T cells are re-placed by other CD 4+ Tcells, continuously being switched fromCD 45 RA ' to CD 45RO * by non-specific activation [21].Although reconciling seemingly contradictory data, ourhypothesis is of course difficult to prove in rivo.

    The present study showed that the relative expression ofHLA-DR and CD38 on CD4* T cells is increased in HIV-positive subjects and becomes relatively more important withprogression from the asymptotnatic lo the syniplotiiatic diseasestage. It has been shown that anti-retroviral therapy cantemporarily d own-regu late the expression levels of H LA -DRand CD38 on CDH eells [4,22]. To examine the effect of anti-retroviral treatment on the expression levels of these antigens onCD 4eells, we compa red a group of treated p atients with a groupof untreated patients, matched for CD4 percentages andabsolute numbers of CD4' T cells; however, no significantdifferences could be demonstrated. To explore the possibilityihat treatment has a temporary effect on activation antigens onCD4 eells as previously shown for CD8 cells, a longitudinalstudy would be needed.

    In conclusion, this study showed that the expression ofact ivat ion ant igens CD38 and HLA -DR within ih eC IM ' Tcel lsubset changed significantly after HIV infection and increasedwith disease progression. A signiiicani correlation was observedbetween activation of CD4* and CD8^ Tcells on one hand andbetween activation of C D 4 ' T cells and the presence of HIVantigen on the other. The usefulness of assessing ihese twoaetivation antigens on CD4' T cells as surrogate markers fortherapy or as prognostic markers for disease progressionremains to be determined.

    ACKNOWLEDGMENTSThe uttlhors wish to thiiiik Dr Ludo MLIVIIC froni the AntwerpTransfusion Centre lor providing the control blood samples, aGreet Verhulst tor secretarial assistance.

    REFERENCES1 Fahey JL. Taylor JMG. Dctcls R et al. The prognostic vi

    cellular and serologic markers in infection willi human immuciency virus type I. N Engl J Med 1990; 322:166 72.

    2 Prince HF. Jensen ER. Three-color cytofluoromelric analyCDS cell subsets in (IlV-l infection. J Acqiiir Immun Defic1991:4:1227 32.

    y Kestens L, Vnnham G. Gigase P el al. Fxpres.sion of iK-antigens. HLA-DR and CD38 on CD8 lympfiocylcs duringinfection. A IDS i')92: 6:793 7.

    4 Levacher M . Ihilstaert F. Tallet S. Uilery S. Pocidalo JJ. BaThe significance of activation markers on Iymphocyles-Chuman immunodeficiency syndrome staging and prognosticClin Exp Imm unol 1992: 90:376 82 .

    .1 Prince HE, York J.Jensen FR. Phenolypic comparison of thpopulat ions of human lymphocytes defined by CD45RCD45RA expression. Cell Immunol 1992: 145:254-62.6 Jackson DG. Belt JI . Isolat ion of a cDNA encoding the CD38 (TIO) molecule, a cell surface glycoprotein with an udiscontinuous pattern ofexpression during lympliocyte diiVtion. J Immunol 1990: 144:2811-5.

    7 Vuillier F, Laprcsic C. Dighiero G. Comparative analysts o4B4 and CD4-2H4 lymphocyte subpopulat ions in HIV nhomosexual, HIV seropositive and healthy subjects. ClImmunol 1988:71:8 12.

    8 Prince ME. Czaplicki CD . Preferential loss of leu-H . CDH L A -D R' CD 8 cell subsets during i/i i i /w culture of mon ocells from humcin immunodeficiency virus type-1 (lllV)-seropformer blood d ono rs. J Clin Imm unol 1989: 9:421 8.

    9 Reddy MM , Grieco M H, Quanti tat ive changes in T-helper (CD4^ CD45RA ) , T-suppressor inducer (CD4* C D45 RAsuppressor (CD8 ' C DII b+ )- and T-cylotoxic (CD 8' CDsubsets in human immunodeficiency virus infection, J Clin LaI'WI: 5:96 100,

    10 Vanham G. Kestens L, Penne G

  • 8/13/2019 j.1365-2249.1994.tb07015.x - copia

    6/7

    HIV-} infection and CD 4^ T eell activation 441ul . Increas-

    ing viral burden in C D 4 ' T cells from patients with huma nimmunodeficiency virus (HIV) infection reflecls rapidly progressiveimmunosuppression and clinii:;il disease. Ann Intern Med 1990:112:438-43.

    r l ing JM .Le db et terJ A,S iasJc /u/ . HlV-infce ted humans , but notchimpanzees, have circulating cytoloxic T lymphocyies that lyseuninTected CD 4+ cells. J Im mun ol 1990: 144:2992 S.Janossy G, Borthwick N, Lomnit /fr Rci ul.Lymphocyte act ivat ionin HIV-I infection, I, Predominant prolifcriitive defects among

    CD 4.1 RO ' cells of Ihe CD4 and C DS lineages, AID S 1993: 7:613-24.21 P intoL , Cov asM J, Vic lor in oRM M. L(W sorCD45RA and gain of

    CD45RO Liflerin rlirn activation of lymphocylcs from HIV-infectedpaiients. Immun ology 1991:73:147 50.

    22 Bass HZ. Hardy WD, Milsuyasu RT. Wang YX, Cumberland W.F'ahey JL. Fleven lymphoid phenolypic markers in HIV infection:.selective changes induced by Zidovudine trealment. J AcquirlmmunDefrc Syndr 1992; 5:890-7.

  • 8/13/2019 j.1365-2249.1994.tb07015.x - copia

    7/7