Characterization of the mechanism of the immunosuppressive activity of Interleukin-10 on human CD4?T cells

77 Pages·1999·0.98 MB·English

by  Perrin, George Q. III

CHARACTAECRTIIZVAITTYIOONFOIFNTTEHRELEMUEKCIHN-A1N0IOSNMHOFUMTHAENICMDM4U*NTOCSEULPLPSRESSIVE
GEORGEQ.PERRINIII
OAFDTIHSESOEUFRNTITAVHTEEIRORSNEIDQTPOYURCIOETRFSOEEFRMNLETOONEFRTDIPSDTHAFOIOLITRONHSPTEOAHPGREHRTDYIAEADGLURFAEUTELEFOSIFCLLHMOEONLT
UNIVERSITYOFFLORIDA
1999
ACKNOWLEDGMENTS
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TABLEOFCONTENTS
ALICSKTNOOFWTLAEBDLGESMENTS.
LISTOFF1GURF.S
ABSTRACT
CHAPTERS
1INTRODUCTION
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SIInunhhpiipbbriiettsiisoonniooonffoCSfeEltBlheCIyMncidlxueceePddroLPgyrrmoelpsihsfoieocrnayttiienonSRoFefBa-cPStBitouMnnCuslsicdCD4'Tcells
4DISSSTMIII1R1RuuLnL11aeCiE-hpp*--idmUiFpp2211nuebSrr00EtciRPeeSCeatReeIDssrinoInivdEsstoharueOiiioNienrarooNblclcoCssnniteefeeEtlRvoossyCeeSooffvlynfBfB.eRsFdleTAIra.ouoncsuf1fhctta-m0riPklIab1oUIrsEciop.ArofetP•fcrdflir?tcSuelooiIucguclorjvtLupitruoisf-upia-tcf1ioronryS0lneCasniisiecgEiosClrnfoneoliafilrplieCl-bcPyotoCRrcfsneyolCgtoceReuDnleilPegSnCaru'ePtopllrgea2Tlordt7geCoC*Krsrye*eussclsiullscotsLe-insefoePvonrseAotlcaets-ni.d'i ,.....3J34S5I48I24
BIOGRAPHICALSKETCH. -66
OFTABLES
1.BiologicalPropertiesofIL-10 5
2.IL-10SuppressesStimulationofCD4'TCells ... JO
3.IL-10DoseResponse.....,.,.,. ., 32
4.TimecourseofIL-10Addition 33
5.IL-10SuppressesIL-2Production „J7
6.EarlyIL-10SuppressionofIL-2Production 39
7.AdditionofIL-2ReversesIL-10Immunosuppression:
NoIL-10Pretrcatment .40
8.AdditionofIL-2ReversesIL-10Immunosuppression:
2DayIL-10Pretreatment 41
LISTOFFIGURES
24531.SMISMLuuaA-ppm1Peem0rraaaKSlnuinipnnnapaggrsncceennCsPeBsBlaieiltnsnhddCIwiiyhancnelyggMei.inxtehdeALbysmepnhcoecyotfeMRHeaCctCiloanssII.. ...2|17892?
6.IL-10SuppressesSuperantigenInducedStimulation 29
7IL-10InhibitsSuperanugenInducedChangesinCellCycleProtons .35
8AdditionofIL-2PartiallyReversesChangesinCellCycleProteins 43
9II.-10ReducesInductionolc-fosandc-jun .45
10IL-10DirectlyBlocksProliferationofCD4'TCells 47
11IL-10SuppressesAcuvanonofFrkinSFR-ShmulatedCeils 49
12.IL-10SuppressesActivationofErkinPMA/ionomycin-StimulatedCells 50
13.IL-10SuppressesActivationofRafinSEB-StimulatcdCells 53
AboRsftetrqhauecitUrnoeimfveeDnritssssietfryotroatftihFoelnoDrPeirgderaseeienntoPefadrDttoioacltthoFeurGlofrfialdPluhmiaeltnoetsoSofcphhtohyoel
CHARACTAECRTIIZVAITTYIOONFOIFNTTEHRELEMUEKCIHN-AB1yN0IOSNMOHFUMTHAENICMDM4U*NTOCSEULPLPSRESSIVE
GeorgeQ.PerrinIII
May1999
CMhaajiorrmDaenp:aHrtomweanrtd:MMi.cJroobhinosloongyandCellScience
Interleukinsarcproteinsproducedbycellsoftheimmunesystemwhichhelp
regulatetherelativestateofactivationoftheimmunesystem.Interlcukin-10(IL-10)isa
multifimctionalcytokinewhichhasimmunostimulatoryorimmunosuppressive
properties,dependingonthetargetcelltype,Ifirstinvestigatedtheantiproliferative
effectsofIL-10inamixedlymphocytereaction(MLR)inwhichperipheralblood
mononuclearcells(PBMC)ftomtwoseparatedonorsareculturedtogether.Theresulting
proliferativeresponsewasconsistentlysuppressedinthepresenceofIL-10.Therewere,
however,inconsistenciesinboththeintensityoftheimmuneresponseandthedegreeof
TofurtherstudyIL-10'simmunosuppressiveeffects,IanalyzedtheeffectsofIL-
10ontheentryofquiescentCD4"Tcellsintothecellcycleuponstimulationwiththe
uperanligcnstaphylc B(SEB).IL-IO <VG,.
tprreoagtrmeesnstionproeuvtenotfedthethGeodopwhnarseeguolfatthieocnelolfcpyc2l7eK.lpIlL,-aInOailnshoibpitroervyenptreodtetihnetuhpartcgcuolnattrioolns
oftheGicyclinsD2andD3,proteinsnecessaryforentryandprogressionthroughtheGi
phaseofthecellcycle.Associatedwiththeinhibitionofthecellcycle,IL-10suppressed
SEBinductionofintcrlcukin-2(IL-2).AdditionofexogenousIL-2toIL-10-lrealedcells
significantlyreversedtheantiproliferativeeffectofIL-10aswellasIL-10'seffectsonthe
earlyGiproteinsp27K,plandcyclinD2.AlthoughthisreversalbyIL-2waspronounced.
effectpurifiedCD4*Tcellsdirectly,providingfunctionalevidenceforthepresenceof
IL-10receptorsonCD4’Tcells.IL-10alsoinhibitedexpressionofIL-2transcriptional
regulatorsc-fosandc-jun.whichalsoinhibitothercellfunctions.Thesestudiesalsoshow
thatthemechanismofIL-10regulationofquiescentCD4*Tcellactivationismainlyby
blockinginductionofIL-2thatiscriticaltodownregulationofp27Ki*"andupregulation
Finally,mystudiesshowthatthisblockageofIL-2inductionisdirectlyrelatedto
theinhibitoryactivityofIL-10ontheproteinsofthemitogen-activatedprotein(MAP)
kinasepathway,whichnormallyleadtoIL-2inductioninactivatedCD4*Tcells.
INCTHRAOPDTUECRTIION
CytokinesandtheImmuneSystem
Cytokinesaresmallsolubleproteinsproducedmainlybythecellsoftheimmune
system.Theseproteinsactprimarilytoregulatetheactivation,growthand
differentiationofvariouslymphocytepopulations.Cytokinesmayhavevaryingeffects
onthecellsoftheimmunesystemrangingfromstimulatinganimmuneresponse,to
suppressinganimmuneresponse,toinducingthedifferentiationofimmunesystem
cells.Theseeffectsarcdeterminedbywhichcytokinesarcactingandwhichtargetcells
theyareactingupon.
synthesizedwhenneeded.Cytokinesynthesismaybeinducedinanumberofways
includingbindingofextracellularcytokinestocellsurfacereceptors.Cytokinesoften
cytokinesmaymediatethebiologicalactionsofthefirstcytokine.Thisabilityofone
cytokinetoenhanceorsuppresstheproductionofothersmayprovideimportantpositive
Cytokinesofteninfluencethebiologicalactivityofothercytokines(Lengycl
1982).Thiscanoccurcitherbyactinginanantagonisticfashion,producinganadditive