Interleukin 1 gene expression in adult T cell leukemia

Interleukin 1 gene expression in adult T cell

leukemia.

Y Wano, … , U Gubler, W C Greene

J Clin Invest.

1987;

80(3)

:911-916.

https://doi.org/10.1172/JCI113152

.

The adult T cell leukemia (ATL) is a T cell neoplasm etiologically associated with human T

lymphotropic virus type I (HTLV-I) infection. ATL cells often abnormally express interleukin 2

(IL-2) receptors, and ATL patients may show clinical evidence of hypercalcemia, osteolytic

bone lesions, or increased bone turnover. Whereas interleukin 1 (IL-1) is not generally

recognized as a product of T cells, this cytokine is capable of both altering IL-2 receptor

expression and activating osteoclasts. Thus, we investigated the possibility that primary

ATL leukemic T cells and HTLV-I-infected long-term ATL cell lines produce IL-1. S1

nuclease protection assays demonstrated that primary leukemic ATL cells from five out of

six patients, as well as one patient with T4+ chronic lymphocytic leukemia, contained

considerable quantities of IL-1 beta messenger RNA (mRNA) and small amounts of IL-1

alpha mRNA. These primary leukemic T cells also released biologically active IL-1 protein

as evaluated in the murine thymocyte comitogenesis bioassay. In contrast to primary tumor

cells, four out of six long-term ATL cell lines produced variable amounts of IL-1 alpha mRNA

in the absence of detectable IL-1 beta mRNA as measured by S1 nuclease protection.

These data demonstrate that IL-1 gene (especially IL-1 beta) expression occurs in many

primary HTLV-I-infected leukemic T cells raising the possibility that this mediator may play a

role in the pathological […]

Research Article

Rapid

Publication

Interleukin

1

Gene

Expression

in Adult T Cell Leukemia

Yuji Wano,*ToshioHattori,*Masao Matsuoka,t Kiyoshi Takatsuki,t Anne 0.Chua,lUeliGubler,§andWarnerC. Greene*

*HowardHughes Medical Institute, Department ofMedicine, DukeUniversitySchoolofMedicine,Durham, North Carolina27710;

*Second

Department ofMedicine, Kumamoto University Medical School,Kumamoto860, Japan; and DepartmentofMolecular

Genetics, Hoffmann-LaRoche, Inc., Nutley, New Jersey 07110

Introduction

The adult T cell leukemia(ATL) is a T cell neoplasm etiologic-ally associated with human T lymphotropic virus type I (HTLV-I) infection. ATL cells often abnormally express in-terleukin 2(IL-2)receptors, and ATLpatientsmayshow clin-ical evidence ofhypercalcemia,

osteolytic

bonelesions, or in-creased bone turnover. Whereas interleukin l (IL-i) is not generallyrecognized asa product ofT cells, this cytokine is capableof both altering IL-2 receptor expression and

activat-ingosteoclasts. Thus, weinvestigatedthe possibility that pri-maryATL leukemic T cells and HTLV-I-infectedlong-term ATL cell lines produce IL-1. Si nuclease protection assays demonstrated thatprimaryleukemic ATL cells fromfiveoutof

six patients,as wellas onepatientwithT4' chronic lympho-cytic leukemia, contained considerable quantities of

IL-i8

messenger RNA(mRNA)and smallamountsofIL-la mRNA. Theseprimary leukemicTcells also released biologically

ac-tiveIL-1proteinasevaluated in the murine thymocyte

comito-genesis bioassay.Incontrast toprimarytumorcells,fouroutof

six long-term ATL cell lines produced variable amounts of

IL-ia

mRNA in the absence ofdetectable

_IL-Ilf

mRNA as

measuredby

Si

nucleaseprotection. These data demonstrate that IL-1 gene (especially

IL-liN)

expression

occursin many

primary HTLV-I-infected leukemicTcellsraisingthe possi-bilitythat this mediator may play arole in the

pathological

changes associated with this leukemia.

Also,

these studies

show that the pattern ofIL-1a and

IL-IS

gene expression differs between

primary

ATLtumor cellsandlong-term cul-turedATL celllines,

indicating

aninteresting biological dif-ferencein thesetwoHTLV-I-infectedcellpopulations.

Part of thisstudywaspresentedattheannualmeetingof Federation of American Society for Experimental Biology,

Washington

D. C.,

March-April 1987, andwas_publishedas an abstract

(Fed.

Proc.

46:1344.).

Address allcorrespondencetoDr. Y.Wano,HowardHughes Med-icalInstitute,DukeUniversityMedicalCenter,P.0.Box_3705, Dur-ham, NC 27710.

Receivedfor publication 21January1987andinrevised

form

20

April1987.

The adult T cell leukemia (ATL)' is a usually fatal T cell malignancy produced by human T lymphotropic virus type I (HTLV-I) infection ofT3'and T4' lymphocytes

(helper/in-ducerphenotype) (1-3). These leukemic T cells are often

char-acterized by the abnormal constitutive expression of mem-brane receptors forinterleukin 2 (IL-2) (4, 5). Clinically, this leukemia may present in acute, chronic, or smoldering forms (6).The acute formofthis disease, and thedevelopment of a leukemic crisis in the other less common forms, is frequently associated with hypercalcemia (6-8), evidence of increased bone turnover or osteolysis. Several investigators have

re-ported that HTLV-I-infected primaryleukemic T cells pro-duce osteoclast-activating factor (OAF) or OAF-like sub-stance(s) that promotebone resorption and may contribute to the development of hypercalcemia (8-12). However, this

ATL-associated OAF has not been purified and remains

poorlycharacterized. Recently,Dewhirstetal. (13)purified a human OAF (Mr of 17,800; isoelectric point

[p1]6.8)

from

mitogenandphorbolester-activated peripheral blood mono-nuclear cells (PBMC) and unexpectedlyfoundthatits NH2-terminal amino acid sequence wasidentical to that of

inter-leukin

Il#

(IL-18) (14, 15).This finding, coupled with the

rec-ognized ability ofIL-1 toinduceIL-2 receptor gene expression (16, 17) and to enhance bone resorption in vitro (18, 19),

promptedus toinvestigatewhether primary ATL tumor cells

andHTLV-I-infectedlong-term ATL celllinesproduce IL-1.

Methods

Patients.Sevenpatients with HTLV-I-inducedATLand onepatient with T4' chroniclymphocytic leukemia (CLL) not associated with HTLV-I infectionwerestudied.AlltheATL cases,butnotthe T4' CLLcase, werefoundto containamonoclonal patternof HTLV-I provirus integration in thetumorcellDNA(2).Allbutoneof theATL casesrepresentedpatientswith theacuteform of leukemia. The other ATLpatient(case5)hadhighleukemic cellcountsin theperipheral

blood

(34,000-50,1O001.),

butwasclinicallystablewithouttreatment,

and thuswascategorizedas"chronic"ATL.

Cells. The long-term cultured HTLV-I-infected T celllines 1.Abbreviations usedinthispaper: ATL,adultTcellleukemia; CLL,

chroniclymphocytic leukemia; HTLV-I,human Tlymphotropicvirus

typeI;nt, nucleotides; OAF,osteoclast-activating factor, PBMC, pe-ripheral blood mononuclear cells; rIL-la and rIL-l, recombinant interleukin Ihaand 1 ,.

Abstract

J.Clin.Invest.

K TheAmericanSocietyfor ClinicalInvestigation,Inc.

0021-9738/87/09/0911/06 $2.00

HUT-102B2, MT-1, MW,C5/MJ, PLP6, and C91/PL(20-22)were

cultured incomplete mediumconsistingof RPMI 1640supplemented

with 10% heat inactivatedfetal calfserum(FCS), 2.0 mM L-glutamine, 100 U/mlpenicillin and100,g/mi streptomycin.PBMCfrom normal volunteers and the leukemicpatients were separated by Ficoll-Hypa-quedensitygradientcentrifugation. PBMCpopulationsfrom the leu-kemicpatients consisted of>92% of TI1I+cells, 2 76% of T4'cells,

and <2% (< 1% in most cases) ofMol' cells as assessed by flow

cytofluorometry, which indicate minimal monocyte contamination. Cells from thebuffycoatsof normal donorswereculturedfor 18 h in the presence oflipopolysaccharide (LPS, 10ug/ml).

IL-IcomplementaryDNA(cDNA)and protein.ThehumanIL- Ia

(23) andIL-I, (14)cDNAs used have beenpreviouslydescribed. Puri-fied recombinant IL-la(rILla)andIL-1I3(rIL-Ij3)werekindly pro-vided by the Otsuka Pharmaceutical Co.(Tokushima, Japan)and the Dainippon Pharmaceutical Co.(Osaka,Japan),respectively.

Murinethymocyteproliferation assayforIL-I activity.

Superna-tantsfrom cultures of the ATL,T4' CLL,ornormal cells(106/ml)

were harvested after 72hof incubation incomplete medium. After sterile filtering, serial dilutions of these supernatantswereaddedto

C3H/HeN murinethymocytesculturedincompletemedium in the presenceof5 ug/mIofPHA(Difco, Inc.,Findlay,OH),as_previously

described (24). [3H]Thymidinewasaddedduringthe final 6 hof the 72-h culture.

SI nuclease protection assayofleukemic T cell RNA. RNAwas

purified by theguanidine isothiocyanatemethod(25). 10 Ag of total

RNAobtained from the different leukemic and normal cell

popula-tionswerehybridizedwith either theIL-laorIL-1NcDNAsand di-gested with S Inuclease,aspreviouslydescribed(26).Inthis assay,fully

splicedIL-la messenger RNA(mRNA)protectsa210-nucleotide_(nt) fragment(amino acids 64-134 of the IL-la precursorprotein)derived fromexons 4and 5 (27). IL-I( mRNA protectsa525-nt fragment

(amino acids 133-269 of theIL-1j3precursorplus 116ntfrom the 3' untranslated region).Thespecific activityof the end-labeledprobes

usedwas1 X 104-2 X_104dpm/fmol.

Results

PrimaryATL tumorcellscontainIL-iaand

IL-1j3

mRNA.To

investigate thepossibility ofIL-i gene expression in the pri-mary leukemia T cell populations, SI nuclease protection assays were performed with total RNA isolated from these

neoplasticcells. As shownin Fig. 1 (left),the expected 210-nt

fragment protectedby IL-la mRNA was readily detected in

the LPS-induced buffycoat cell RNA (positive control), but not inthepig gut RNA sample (negative control) nor in the

absenceofadded RNA. Large quantities of

IL-la

mRNA were present inthe T4+ CLL RNA sample, whereas very low, but

detectable, levels ofIL-la mRNA were present in RNA

iso-latedfrom ATL cases, 1, 2, 3, and 4. IL-la mRNA was not

detected inATL cases S and 6 norin RNA isolated from the

HTLV-I-infected HUT-102B2 cell line. RNA from the in-duced human peripheral blood leukocytes (buffy coat cells)

consistently yieldedtwo protectedfragments, 210 and 125 nt in length (26, 27). This finding is explained by the fact that this S1 probeincludessequences from both exons 4 and 5 of the

IL-ia gene (27, 28). The 210-nt fragment is produced after

hybridization of the probe to fully spliced or mature IL-la mRNA, whereas the 125-nt fragment, by hybridization of the

probetounsplicedIL-lamRNA. The 3' 85-nt fragment was notdetected in our procedure as a 5' end-labeled fragment was used. Thedifference in relative ratio of the 210- and 125-nt

fragmentsbetween the T4+ CLL and the buffy coat cell

sam-ples, thus, may reflect a difference in relative amounts of

splicedand unspliced IL-la mRNA presentinthesetwo dif-ferent cell populations.

Muchgreaterquantities of IL- IrmRNAweredetected in

fiveoutof sixATLcases(cases 1-4 and6)asindicated by the intensities of the protected 525-nt fragments (right, Fig. 1). ATLcase5wasthe only leukemic sample that didnotcontain either IL-la orIL-l1: mRNA. Interestingly, these cells were

obtained fromapatient withachronic rather thanacuteform of this leukemia. The

T4'

CLL cells contained large quantities of IL-I(3 mRNA, whereas HUT-102B2 T cells again lacked this mRNA species. Buffy coat RNA, piggutRNA, and no RNAcontrols yielded the appropriate patterns of protection. The multiple smaller bands observed in the IL-

_1#

assayreflect partial escapefromcomplete protectionthat iscommonly

ob-served with this particular probe and the conditions of Si nucleasedigestion used.

These findings indicate that many of the primary ATL

tumorcellsexpressbothIL- la and IL-I mRNAand that the

levels of IL-1

#

mRNA are considerably greaterthan those of

IL-la mRNA. Northern blotting studies confirmed the pres-enceof IL-13mRNAinfour out ofsix ATLleukemiasand in

the

T4'

CLL sample. However, usingthis less sensitive tech-nique, IL-la mRNA production wasweaklydetected in the

T4'CLLRNA sample only and undetectableinall oftheATL

samples (data not shown).

Thymocytecomitogenic activity inculturesupernatantsof

primary tumor Tcells. To examine whether these tumor T cells actually release biologically active IL-I protein,thecells were cultured for 72 h in complete medium and the culture supernatants assayed for the presence ofthymocyte comito-genic activity. Assummarized in Table I, acontrol

superna-tantobtained from normal PBMC didnotpromotesignificant

[3H]thymidine

incorporation by murine thymocytes, whereas both purified recombinant IL-la and IL-13 significantly

en-hancedthymoctyeproliferation in adose-dependent manner. Supernatants from the T4+ CLL and four of the ATLcases

that contained IL-I mRNA, as well as an additional ATL sample not studied at the RNA level (case 7),containedIL-I biologicalactivity as measured in this assay. These results sug-gestthat theprimary tumor cells from the ATL and T4+ CLL patients secrete or release biologically active IL-I protein.

The absence ofIL-1(3 mRNA in HTLV-I-infected

long-term celllines. Whereas IL-,1 gene expression was evident in mostof theI-infected primary ATL cells, the HTLV-I-infected long-term ATL line, HUT-102B2, did not appear to contain either IL-Ia or IL-1,B mRNA (Fig. 1). To determine whetherthis was a general property of long-term cultured ATL cell lines, RNA samples from five additional ATL cell lines werestudied for the presence of IL-la or IL-I

_#

mRNA. IL-I

_#

geneexpression was undetectable inall of theATLcelllines

(right, Fig. 2). In contrast, considerable quantities of IL-la mRNA were presentin the C9 1/PL and C5/MJ RNA samples, whereas small amounts were detected in the PLP6 and MJ cell lines. No IL-lIa mRNA was found in the HUT-102B2 orMT-l

RNA samples. Thus, thesefindings suggest that the pattern of IL-Igeneexpressiondiffers inprimary and long-term cultured celllines infected withHTLV-I.

Discussion

rV~~~~~~~~%

I

-. K.("V

'b

b06

4o

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N s

C.)

AZ

se

a;

a

)

0

,rs

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v

_Te

_T

_Ts

_tf4?

525

-_

iI04

*1

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153-

_

_;

-_

125

IL-ia-Figure1. SI nucleaseprotectionassay of RNA isolated fromprimary leukemicTcells. End-labeled cDNAprobes specificforIL-la(left) andIL-Ifl(right)mRNAwerehybridizedwith 10,gof totalRNA

from thenormal andprimaryleukemiccells followedby digestion with SI nuclease (26).Asapositiveandanegative control,RNAwas

aswellasinasinglecaseofT4'CLL. It is unlikely thatthese IL-1 mRNAs are derived from contaminating monocytes

since, inmostcases,the

Mol'

cellsrepresented< 1%(atmost

<2%) of the tumorcell population and no specific

macro-phage-inducing agents were used. Furthermore, identically preparednormal T cellpopulations andoneATLsample

con-tainingasimilar number of

Mol'

cellslacked detectable

evi-dence of IL-l geneexpression. Inall populationsof the

posi-tiveprimaryATLtumorcells studied, the relativeamountsof IL-1,8mRNAwerefargreaterthan that ofIL-lamRNA. Also, these cells secretedbiologicallyactive IL-1proteinasmeasured inthe murinethymocyte costimulatorassay.Insharpcontrast,

noneof the sixHTLV-I-infected long-term culturedATLcell lines contained IL-13 mRNA; however, four of these cell lines

produced variable amountsof IL-la mRNA. These findings reveal anintriguingdifference in these twoHTLV-I-infected

cell populuations. Interestingly,recent studies have suggested

that many long-term cultured ATL cell lines do not

corre-IL-1P

isolated from LPS-stimulated peripheral blood leukocytes (buffy coat) and piggut,respectively. SI nucleasedigestionof eachprobein

theabsence of addedRNAwasalsoperformed.Thedigestion

prod-uctsweresize-fractionatedon a5%denaturing acrylamide

sequenc-ing gel, and exposedtoKodakXAR filmfor 55hat-70'C.

spondtotheoriginal primarytumorcells basedontheclonal

pattern ofrearrangementswithin the T cellreceptor chain

gene locusor in theintegration site of the HTLV-I provirus (29). Also, expression of thetransactivator(tat)geneproduct

of HTLV-Iisusuallyundetectable in unculturedprimary leu-kemiacells, whereas long-term cultured ATLcell lines often containrelatively large quantitiesof thisprotein (30).

Further-more, the primary leukemic T cells often require IL-2 for short-termgrowth,whereas theATLcell linesgenerally

prolif-eratereadilyinthe absence of IL-2.Thus,it is clear that these two HTLV-I-infected T cell populations exhibit potentially important geneticand functional differences.

Marked IL-1 production was also observed in the single

caseofT4'CLL studied. These cellsdo not containintegrated

HTLV-I provirus, thus, IL-1 production byleukemic T cells

doesnotappeartobesolelylimitedtoHTLV-I-infectedcells.

Interestingly, these T4' cells also display IL-2 receptors and

respond to exogenousIL-2 with increased proliferation. It is

(nt)

517

506-%W

I

N&**.4

N

_Al/

₀₃

_Itp.

₄₀

_co

A.,'

-₄

.v

.,.

C.)

&

& &

,Z-'

..

T T T

T T

344

221

220-(nt)

517

-506

-396

-344

-298

...

F..

_

-_

-A.;,.

-210

221

-220

-153

4-

TableI. Thymocyte Comitogenic Activity in the TumorCellCulture Supernatants

Fmaldilution

Experiments orconcentration [3H]TdRincorporation mean±SD cpm

Medium 1:4 650±100

rIL-la 10U/mi 8,400±850

100U/mi 26,650±880 rIL-I 1 U/ml 3,710±120 10 U/ml 14,910±2,610 100 U/ml 27,760±5,470

T4CLL 1:4 31,100±2,210

1:12 23,800±1,510 1:36 9,620±570 ATLcase1 1:4 32,250±6,710

1:12 19,250±1,450 ATLcase2 1:4 8,370±2,880 1:12 1,050±190

ATLcase3 1:4 3,250±1,790

ATLcase 6 1:4 4,620±820 1:12 2,840±990 1:36 1,220±230 ATLcase7 1:4 20,200±1,440

possible

that the

production

ofIL-I

_by

these cellsmay

play

a

role in the

unexpected

IL-2

receptor

expression

observed.

Ad-ditionalcasesof

T4'

CLLare now

being

examinedto

investi-gate

the

_generality

ofIL-I

production

inthis form of leukemia andtodetermine the

_relationship

of this mediatortothe

ab-normal

_expression

ofIL-2

_receptors.

Whereas_{IL-I gene}

_expression

has been

previously

demon-strated ina

_variety

of cell

types,

including

monocytes-macro-phages,

B

_cells,

andendothelial cells

(31),

Tcell

_production

of this

_cytokine

has not been

_generally

_recognized.

_Recently,

Tartakovsky

and

_{colleagues (32)}

have

_reported

IL-la

produc-tion

_by

certain

_{antigen-specific}

murineTcell clonesafter acti-vation with

antigen-presenting

Bcells. Nomaand

colleagues

(33)

alsohavedescribed IL-Ia butnotIL-I(3 mRNA

produc-tion ina

few

long-term

culturedhumanT celllines infected with HTLV-I. Our

finding

of the IL-Ia and IL-

IB

gene

ex-pression

in

primary

ATLand

T4'

CLLcells

coupled

with the

finding

thatfouroutof six HTLV-I-infected

_long-term

Tcell

lines

expressed

the IL-la gene

provides

additional evidence that_{IL-I gene}

_expression

canoccurin activated cells of theT

lymphocyte

lineage.

Although

IL-i

production

by

primary

ATL tumorcells appearstorepresentverycommon

_finding

_(thymocyte

comi-togenic activity

has also been detected in the leukemic cells isolated from

_eight

outof nine additionalATLcases not

dis-cussedin this

_paper),

it is unclearwhat rolethismediator

plays

in the

pathological

changes

associated with this leukemia.

IL-I

hasbeen

_reported

toenhance_{IL-2 receptor}

_expression

_(16,

_17,

31),

and

abnormal

IL-2 receptor

display

isacommonfeature inATL

_{(4, 5).}

Whereasit is

_possible

thatIL-I

production plays

a direct role in

_altering

IL-2 _receptor gene

expression,

our

preliminary

results

indicate

that

neutralizing

anti-rIL-la or

anti-rIL-

I#

rabbit

heteroantibodies

failtoinhibitthe

sponta-neousincrease in IL-2 receptor expression that occurs when

primaryATL tumorcellsareplacedin short-term culture(3).

Furthermore, in contrast to a previous report (34), we have been unable to demonstrate that rIL-la or rIL-j3 enhances

either IL-2receptordisplayortheproliferationoffreshly

iso-latedATLcells. Moreover,wedid not detect IL-la orIL-l,8

gene expression in HTLV-I-infected HUT-102B2 or MT-i cells that constitutively express

large

numbers of IL-2 recep-tors. Thus, while not completely excluded, acausal role for IL-la orIL-1,8 in the abnormal expression of IL-2 receptors in HTLV-I-induced ATL has not been established.

Asecond prominent feature of HTLV-I-induced ATL is itsfrequentassociation withhypercalcemia (6-12).The com-monfindingsof osteoclast activation on bone biopsy and in-creased bone turnover without evidence of frank tumor cell infiltration (8, 10-12) has suggested the possible involvement of circulating humoral factor(s) in ATL-associated hypercalce-mia. The abnormal production of such humoral factors as parathyroid hormone, 1, 12-(OH)2 vitaminD3,or

prostaglan-dins,implicated as the cause of hypercalcemia in other malig-nancies(35), has not been detectedin ATL(8-12).Our

find-ings of IL-l production byprimaryATLtumorcellscoupled

withprevious reports of OAF productionbythese cells (8-12) is particularly intriguing in view ofthe report byDewhirstetal. (13) that OAF and IL-

_I#

areidentical insize,isoelectricpoint,

functional properties(thymocytecomitogenesisand bone re-sorption), and NH2-terminal amino acid sequence. It seems

likely that at least one OAFproduced by primaryATL tumor cells corresponds to

IL-lIP.

Inthisregard,hypercalcemiawasdocumentedatthe time of tumor cell isolation in two of the ATL patients studied (cases 1 and 2). The lack ofhypercalcemiain the other cases may reflect compensatory changes in renal and endocrine

function, whichserve to normalize the level ofserumcalcium (35). Consistent with thispossibility,Bunnetal.(8)have noted that ATLpatients oftenshow evidence of osteoclast activation andaccelerated bone turnover in the absence of frank

hyper-calcemia. Neither bonebiopsiesnormetabolic bone turnover

studies were available from the present group ofpatients.

Thus, toestablishfirmly thepossiblerelationshipof IL-la and

IL-1, production to ATL-associated changes in calcium

me-tabolism, serialprospective studies correlatingIL-Iproduction

with changes in bone turnover, osteoclast activation, and

serumandurine calcium levelswillbe required.

IL-I can also produce

neutrophilia,

acute

inflammatory

protein secretionbyhepatocytes, andfever in vivo (31, 36).In

thisregard, Yamamotoetal. (37) have described the frequent

presence of neutrophilia in ATL patients. It is possible that

ATLcell-derived IL-I maymediate this response. Also, ATL

patients, including those we have studied, often exhibit

ele-vated serum levels of lactic dehydrogenase, alkaline

phospha-tase, and C-reactive protein (1, 6, 8). Thesechanges may be

consistent with arole forIL-1; however, direct leukemic cell

infiltration oftheliver (afrequent

finding

in ATL) couldalso produce these changes (1, 8).Finally, fever isafrequent

clini-calfinding inATL;however, it is difficultto

implicate

tumor

cell-derived IL-I asits solecausegiven thefrequency of inter-current opportunistic infections and the administration of multiple drugs.

Ofnote,the

single

ATL case(case

5)

whosetumorcellsdid

diag-(nt)

^aIWUz

517

525506 -396 344

298-221

210

% co

te\+~

~~~~N'b

-mwswwrt

(nt)

0

517 506

-396

-344

-298

221 210 -210

153

-153

-125

IL-la

IL-

1i3

Figure 2. SInucleaseprotectionassayofRNAfrom long-term cultured ATL(HTLV-Iinfected)celllines.For details, see the legendfor Fig. 1.

Left,IL-la;right,IL-l.

nosed as chronic ATL. Preliminary studies of leukemic cells from a second chronic ATL case also has revealed a lack of IL-I mRNA production. Furtherstudiesof thepotential

dif-ferences in IL-l gene expression in the acute vs. chronic or

smolderingforms of ATLarepresently underway. Acknowledaments

WewishtothankDr.I. Miyoshi of Kochi Medical School, Japan fora

generousgift ofMT- I cellline.

This workwaspartially supported byagrantinaid for scientific

researchfromtheMinistry of Education, Science,and Culture, Japan. References

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