Document 0659O8G27o1xk51wQeXNzyJvx
ouzie
Perfluorooctane Sulfonic Acid Induced HMG-CoA Reductase
in
nt
and Rat Py
DeannCaorJp. oLruaetbekeTro,xiMc.oSl.o,gyA,dv3aMncMeeddiRceasleaDrecphaTrotxmiecnotlogist
3M Center, Building 220-2E-02, Saint Paul, MN 55144 Phone (651) 737-1373 FAX (651) 736-2285 Diluehker@mmm.com
Inhibition
oR-TF
PREFACE
"This project is designed toserveboth as a 3M research project and as aprojectto fulfill
the requirementsof a Ph.D. thesis in Toxicology through the University of Minnesota
"Twin Cities. This
the results will be
proposails being. `submitted with
subject to a first-right-of-review
the understanding
by 3M.
that
publication
of
PROPOSED PROJECT AND ITS PURPOSE
"The objectiveofthis study i to gain knowledge into the mechanism of action by which
perfluorooctane
development in
sulfonic
rats. The
acid
hypo
t(hPeFsOiSs)thiantdPucFeOsSadavcetrssveiaefifnehcitbsitoinopneroif-3/p-ohsytdnraotxayl-3-
c`hmoeltehsytlegrollutsayrntyh-eCsoiAs,rewidlulctbaeseex(aHmiMnGe-d.CoAAnreadtutcetmapste)w,iltlheberamtae-dleimtiotipnrgeevennztyPmeFOoSf
toxicity by co-administrating mevalonate, the immediate product of HMG-CoA
reductase. HMG-CoA reductase activity willbe compared between `dams and their pups
and to age matched positive and negative controls. Assays will be performed to.
determine if PFOS acts directly on HMG-CoA reductase, indirectly via activation of
OAtMhPe-racptairvaamteteedrsp,rowtheiinchkiwnialsleb(eAfMoPl-loPwKe)d worhetrheropuogshsiibnldeu,ciinncgluadneibnocrdeyasweeiignhAt,MlPivelrevels.
weight, liver and serum PFOS, liver and serum lipids and miocardial fiber injury and/or
inflammation.
INTRODUCHON
Perfluorooctane sulfonic acid (PFOS) is the ultimate metabolite of|`perfluoroalkyl acids and their derivatives, aclass of chemicals produced by 3M and `broadly utilized across `multiple markets. The large production volume and broad-based utilization of this class
oafdcvheermseichaelasltjhusetfiffeicetsstthheenyemeadyfopro`saceo.mBperceahuesnesPiFvOeSunidsetrhsetabnrdeiankgodofwtnhepprootdeuncttoiaflthese:
chemicals, research focusing onitseffects and mechanism(s)ofaction is of primary
04441
0e.1rLaoshser
01212000
concer. This proposal is PFOS induces effects on
paeirmi-e/dpoasttgnaatianlindgevkenloowplmeedngteiinnrtaotst.he
mechanism(s)
by
which
Figure 1 : Chemical Structure of PFOS
error io o
Gweeniegrhatlgaeifnf,ecitnscsreeeanseidn p3eMrcePnFtOsStilrlebporrondauncdtidoencsrteuadsieedspiun prastusravrievarle(dCuacseed, mparteesrenntaaltbioond,y P19F9O9)S. trTehaetmpernitmianryadtualrtgertaosrignacnluiddeendteifcireedasferdomboadduyltweriatghsttugdaiiens, iisntchreealisveedr.liEvfefrewctesigohft, dpreoclriefaesraetdiosne.ruTmhechfoilressttdeertoelc,ttarbilgelybcieorliodgeiscaalnedffbeiclitrsuebiennainndadmuilltdrapterPoFxOiSsosmteudies is rbeedcuocmeed sveerryumstceheopleasntderaonl.aAppLahriegnht dwoassetsinogfsPyFnOdSroimneraotcsc,utrosxiwchiitychrecsuplomnisneatceusrviensdeath (3M Corporate Toxicology, unpublished, 1999). HefafuecgthoofmPaFnOdSSipnyrdaetvs.olTdh(e1y99f2o)unidnvPeFstOiSgatteoddtohwenmerceghualnaitesm3-uhnydderrolxyyi-n3g-mtehtehhyylgploultiapreymli-c CaomiAcrreodsuocmtaalsen(zHyMmGe-wChoiAchrceodnutcrtoalsse)thaectriavtiet-yltiomi3ti5n%g sotfccpoinntrcohlo.leHsMteGro-lCsoynAthreesdiusc,ttahsee is cNoAnvDePrsHio"n +ofCHoAM)G.-CTohAe atomomuenvtaolofnHatMeG(-HCMoGA-CreodAuc+ta2seNpAreDsPenHt i+n 2a Hce'll=>ismceovnatlroonlaltede + V1i9a83n)e,gaitnihviebiftieoendboafcmkRreNguAlattriaonnsltahtrioonug(hNarkeapnriesssheiedfgean.e, t1r9a8n8s)craipntdiosnti(mEudlawtairodnosfetfhale.,rate ocofndtergorlaldedataicounte(lGyililserfelv.e,rsi1b9l85e)p.hoTsphheorpyrliamtairoynm(eGcihbsaonni,sm19b8y5;whBiecghetthiasl.,en1z9y8m7e; Hiasrdie `amnedcChaarnliisnmg,s i1n9c9l7u)dewirtehveprhsoisblpehotrhyiloal-tdiiosnulifniadcetfivoartmiantgitohne(eRnoziytmeelm.anOatnhderShceocnhttreolr, a1l9t8e4raa)t,ioanlsloisntemreimcbarctainveatiflounibdiytyN(AMDi(trPo)pHou(lRoosiatnedlmVaennkaantdesSahne,ch1e9r8,5)1.984b) and lAiMvPe-ra(cCtairlviantgedetparlo,te1i9n8k9i).nasIeti(sAcMonPt-rPolKl)edisbtyhreemvearjsoirblHeMpGho-spChoorAylraetdiuocnt,asweitkhinase in rat pishotshephuoprsytlraetaimoncaocmtpiovanteinngttihnethkiisnahsieg.hlAyMcPo-nascetrivveadtekipnraosteeicnaskciandaese(Hkianradsiee (anAdMPCa-rPliKnKg,) a1l9l9o7s;teHriacradliley,aCcatirvlaitnignganAdMCPa-rlPsKoKn,,1b9y98b)i.ndEilnegvattoiAonMoPf-APMKPancdanmaakctiinvgatiet aAMpoPo-rePrK by SSuubbssttrraattee ffoorr tphreotAeiMnPp-hoPsKphKataansdesb,ybayllboisntedriincgaltloy AacMtPiv-atPiKngaAnMdPm-akPiKng(Citoratboentteetral., a1c9t9i5v;atHeadwulndeeyretcoanl.d,it1i9o9n5s; wDhaevrieesAeMt aPli.,se1l99e5v)a.teTdhaenAdMAPT-PPiKs dceapsrcesasdeed.is tThhereefore 04442
DPJa.geL2ue0b1ke6r
01200 cpoantshewqauyesn(cee.sg.,offaAttMyPa-ciPdKsyanctthievsaitsiovniainpchlousdpehoinraycltaitviaotnioonfaocfeAtTyPl-CcoonAscuamribnogxyalnaasbeo)laicnd Sterol synthesis, via phosphorylation of HMG-CoA reductase (Corton ef al., 1995; Henin aetcaild.,ox1i9d9a5t)i.onTvhiiasdceapsrceasdseioanlsoofamctailvoanteysl-thCeoAATlPe-vgelesne(rMeartriinlgl ecfataab,ol1i9c9p7a;tHhawyaayshoif fetatatly., 1nu9c9l8e)o.tiAdeMP(H-aPrdKiecoeutlad.t,h1u9s9a9c)t.toThbiaslamnacye tbheeismuppoprltyanotfdAuTriPngtopethreioddesomfacnedlolfultahre.stress ahsespoactioactyetdew(iCtohrtAoTn,PGdielplleestpiioena,nsducHharadsict,re1a9t9m4e)n.t with metabolic poisonsin the Figure 2: Reversible Phosphorylation of HMG-CoA Reductase Solid lin=e activation, dotted line = inactivation
Ta Sx.
TInhhiisbirteisoulntosfinHaMnGi-nCcroeAasereidnuhcetpaasteicacltiovwitdyenisncirteyalsiepsotphroeiceeilnlu(lLarDLd)emraecnedptfoorrscahnoldesatnerol. cianrcrrieeassemionsctleofatrhaenccehoolfepstlearsomlainLhDuLmbaynst,heinlhiivbeirti(oKnoovfaHneMnGe-t Calo,A19r8e1d)u.ctaBseecareussueltLs DinLa dheocrrmeoanseesinnepeldaesdmafocrhotlheestmearoiln.teCnhaonlcesetoefrporle,ghnoawnecvye,r,pairstaurnitiimopno,rltaacnttatpiroencaunrsdomrattoernal bneohramvailorin(uNtuemraona,nd19p8o8s;tnFartaelemdaenv,el1o9p8m8e;nHtoidngmeanm&mailssko(vBitezl,kn1a9p88&) Daincdtsicshrye,qu1i9r8e8d;for dYeofuinctie&ncyMcoNfammeavarlao,na1t9e91a)n.dIinshoipbrietnioonidosfsHuMchGa-sCdoolAicrhoeld,ucitsaospeenatlyslo aldeeandsinteo,a (ubSioqmuaineotnael,.,h1a9e9m2;ABarnedwemraentya.p,rot1e9i9n3s; bCoorusnidnitoeffaar.n,es1y9l95a)n.dTgheersaenyilsgoeprraennyolidrsesairdeues pirnovtoelivnedgliyncomseymlbatriaonne(Qsyunetshneseiys-,HDunNeAeursepelfiacla.tio1n9,8c3e;llSuulraarnigreotwatlh.,a1n9d83m;eFtaabmoslwiosrmtahnedf al., 1987; Brewer et al., 1993).
04443
DPJa.gLeu3ettke1r6
Figure 3: HMG-CoA Reductase Inhibition
avai
[ered oprennidsythcts|
Deeresed
Dn e T
synthesis
cohohloesotCenororrlmo(eSwrotmea,err.
m omt||Tris | TDpeemeiraiiseat|||| Smymseist
Trvit GZ| eIis || eamehommaanmnail,,rusion, || |||| {FGEeoheianinnQEpouen
YTomo&mko | rFroemain,1988o,y ase || || rSaimirra.o19s8;
crore?
Brower ral, 1999)
plasmaLDLbythe
Decreased plasma concentratiofn
1l9i8ve1r)(Kovancn ef al.
LNaDwLrocchkodleesrtaelr,ol1(9H95e)nck et al., 1998;
`Statins, a class ofcompounds used totreat hyperlipidemia, act as competitive inhibitors
of HMG-CoA reductase (Stryer, 1995). Some statins, for example lovastatin (Mevinolin,
`MmEusVtAbCe OenRz)ymaatnidcsailmlvyaosrtacthienmi(scyanlvliynoolpienn,eZdOtCoOtRhe)i,r reexsipstecatsivienaccatribvoex"ylpartoedrfuogrsm"swthoich elicit inhibitory activity against HMG-CoA reductase (Corsinietal, 1995). Other
statins, for example pravastatin (eptastatin) and fluvastatin (LESCOL),aredosed in their active, open ring form (Corsini ef al., 1995). The active open-acid structures resemble that of HMG-CoA, the normal substrate for the reductase. Statins bind to the reductase, prevent it from binding to HMG-CoA and thus block the production of mevalonate.
004444 Da1ieLsucokresr
ou212000
Figure 4 : Chemical Structures -- Statins & HMG-CoA
ExampleStatins:
Lovastatin (Mevinolin, MEVACOR) Inactive Prodrug
Ho,
Mevinolinic Acid Active Inhibitor
Hom} 8~C--0H
>0
BE
Ha
Ha
CH'
aur"
PrAacvtaivsetaItnihinbitor HO,
> 0
H
CONa
Yi
Hy
HNoMrGm-aCloHAM:G-CoA reductase substrate
H of--o-
CoA
HO
"1T9h9e8)m.ajoErfifetcyotfssitnactliundse airnecrkenasoewdnmtaolfboerdmeavteiloonpsm(eMnitnalslkyertoextiacl,t,o1r9a8t3s)(;Hdeenccrkeaesteadl.p,ost-
r`weedaunciendgfseutravlibvoaldyanwdeidgehltay(eWdisdeeveeflaol.p,me19n9t02o)f;rerfeldeuxceesdanfedtableahnadvioofrfs(pFrDiAng, b1o9d8y7)w;eights
((MWiinssekeetrealf.,al.1,9901b9)90;)r;erdeudcuecdeodfoffsfpsrpirnignbgobdoydwyewiegihgthatnadndindcerceraseeadsesnwciomnamtaalzesuerrvriovrasl
((HFiDrAab,e1t9a9l.3,).19S9t4u)d;ieasndharveetafroduenddsktehlaettcaol-daedvmienliosptmreanttioannodfrmeedvuacleodnafteeta,ltbhoediymwmeeidgihattse
prersoudlutcitngoffrHoMmGH-MCGo-ACroeAdurcetadsuec,tapsreeviennhitbsitOiFoann(iaMgaocnDioznesalVdareiroaul.s, Or1g9a88n;toKxoimcibtrieusst ef al.,
1989; Minsker et administrationof
al., 1983; Hrab et al., cholesterol, however,
1994). had no
Minsker effect on
etfhea.te(r1a9to8g3e)nifcoiutnydotfhlaotvcaos-tatin
(Mevinolin, MEVACOR). Some statins are also known to produce myocardialD.efJf.eLctuse.bker
Page Sol 16
GO4448
01212000 2Hi4rmagb/ektga/l.dy(o1f99f4l)uvfaosutnatdinsi(gnLiEfSicCaOnLt)c.ardAilalcamnyiompalasthdyyiinngpirnetghniasntdorsaetsgdroosuepdhwaidthcardiac lceasuisoenosfwdheiacthhw(erreabcoenfsaild,er1e9d94t)r.eaTthmeenaturtehloartsedcoanncdlubdeelidetvheadttionhbiebiatsisooncioaftmedevtaoltohneate ``aanndd/cohrodleesattehroolf sthyentfhleusviasstwaatisnatrseiagtneidfircaatsn.t Cfoac-taodrmicnointsrtirbauttiionngotfomtehveamlyooncaatredicaolmpdlaemtaege bSeloonckweidthanfdl/uovraastmaetliino(rHarteadbtehteaLm,or1t9a9l4i)ty.,Icna-vridtiraoc smtyuodipcastbhyyGaunidjaortchoeretaadlv.e(r1s9e9e8f,fe1c9ts99) faonudnldovtahsetaltiipnop(hmielivcinsotlaitinn,sMatEoVrvAasCtOatRin)(LtoIPpIrToOduRc)e, aspiompvtaosstiastoinfr(asytnavinndolhiun,maZnOvCaOscRu)lar Sapmoopottohsimsu(sGculiejacrerlolse(tVaSL,MC1)9.98,Me19v9a9l)o.naSteeveardamlinmiesvtarlaotnioantecommeptlaebtoellityepsrweeverneteevdatlhuiasted 1i0sodpeetnetremniynleawdehniicnhe,weubrieqiunivnoolnvee(dcionetnhzeysmteatQi1n-0i)ndauncdedsqapuoaplteonseisf.ailCehdolteosptreervole.nt the Sgteartainnyilngdeurcaendyle-ffpeycrtosphwohsiplheatthee(iGsoGpPrPe)nofiudlslyfarrenveesrysle-dptyhreopehffoescpthsat(Geu(iFjParPr)oaentdal, 1998, i1n9h9i9b)i.toTrhsemsaeystbuedimesorseugagreesstulttheofaddveecrrseeaseeffdescytnstsheeesniswoiftcheHrtMaGin-iCsoopArerneodiudcstatshean of decrease cholesterol synthesis.
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D. J. Lucbker Page 60116
Figure 5: Isoprenoid Biosynthetic Pathway
Key:
A PP = pyrophosphate
+ = shown to prevent
statin
induced
VSMC
apoptosis
in-vitro'
% = shown NOT fo prevent statin induced VSMC apoptosis in-vitro'
= shown to prevent other statin induced adverse effects in-vivo andlor in-vitro
= shown NOT to prevent other statin induced adverse effects in-vivo and/orin-vitro?
!Guijamreof al.,1998 & 1999
Minsektael.r, 1983; Suranietal., 1983;Carson &Lennarz, 1979: Hrabetal., 1994 MacDoneaflald.,
ETT
TEE
--
[roe} [i Comymeediroieo
m= Boca e
[roost
=
FHalimec r]| Np
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fpPage 7 of 16
0112172000
`rSemdiutcht,aLseeaernaznydmeEraicctkisviotny(i1n9r9a5t)s lfeosusntdhannosoerxerqeulaaltetdo 2d8ifdfaeyresnocleds. inHoHwMeGv-eCro,A. Sfioglnliofwiicnagntplyubheirgthyer(Caacrtlisvoitny, lMeivteclhsealrleafnoduGnodlidnfafrebm,al1e97r8a)t.s aLsecvoimnepatrale.d (t1o9m8a9l)es binivoessytnitghaetseids tahneddleivpeidlotprmaensnptoarltcihnahnugmesaninaenxdprraetssfietoanl oafndgenneeosnaitnavlollivveedrsi.nLcehvoellesstoefrol H(MMcGN-amCaorAa,reQduuacctakseenbmuRsNh AanudnRdoedrwgeolll,ar1ge97f2l;ucLteuoatniionest daulr,i1n9g84ra)t,lbievierngonhtiogghepnryior to biinrctrhe,adseicnlgiantinwgeatonilnogw(ldeavyels19d-u2r1inpgosstuncaktalli)n.gI(ndahyusma1n-s1,3 phooswtenvaetarl,)tahnedsetrmanRsiNenAtlyappear iantacnoenacrelniterratpihoansdeuorfinfgetthalerliefset(obfy twheeekint8r)aautnedriunneadesrwgeololnalsypmoisntniamtaall(l<ife3(fLoelvd)inchefaanlg,es C19o8A9).reFducetaastnedaeacatrilvliytypowshtinlaetafletraaltasnmdacyartlhyuspobsetnvaetrayl sheunmsiatnisvemtaoyinnhoitb.itors ofHMGTadhmeinsitsutdryatoiuotnliinnepdriengtnhainstprraotpsorseaslulitssdiensriegdneudcetdomhaetleprdneatlebromidnyewheiogwhtPFgaOiSn, increased ipsertcheatntPsFtOilSlbaorcntsptuopsinahinbditdHecMrGea-sCedopAupresduurcvtiavsael aacfttievritbyiritnh.prTehgenahnytporatthsesainsd/toorbtehetiersted orefsfuslptrsionfgtahnids tsthuudsyc,aanlboneghawviethaswhaadteviselaolpremaednytaklntoowxnicaabntouatt hhiugmhaenr dHoMsGe-lCevoeAls.. The reductase activity, will helpdeterminethe potential risk PFOS holds to be a developmental toxicant in humans. Figure 6: Hypothesis
= &/ or
Reductase In Dams
Reductaseinpups
RESEARCH PLAN
OBJECTIVES
1. Determine whether hepatic HMG-CoA reductase activity is inhibited in PFOS treated pregnant rats. 2. Determine whether hepatic HMG-CoA reductase activity is inhibited in rat pups from PFOS treated dams.
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3. Investigate the mechanism by which PFOS acts to inhibit HMG-CoA reductase activity.
a) Determineif PFOSactsdirectly on HMG-CoA reductase.
b) )
DDeetteerrmmiinneeiiffPPFFOOSS iacntdsucoensAaMnPi-ncPrKe.ase in cellular AMP levels.
EXPERIMENTALAPPROACH
Ableltwreeseunltdsawmillefbfeecctoamnpdapruepd ebfeftecwte.eBnoddaymsweaingdhtthaenidr ptuotpasl tliovierdewnetiifgyhtanwyilrlelbaetrioencsohridped
fdoarmesacahnddapumpsanwdilliltbeeracsomipnadirceeds toofaggeenemraaltchheeadltphoasintdivheepcaotnotrmoelgsal(ldyo.seEdawcihthgraosutpationf),
npeagraatmievteercsontthraotlwsi(lldobseedfowliltohwevdehwichleer)eapnodssmiebvlaelionncaltuedechlaivlelrenagneddsceornturmolPsF. OOSthleervels,
liver and serum lipid levels and necessary, portionsofthe study
myocardial fiber injury maybesent to contract
and inflammation.As labs for completion.
deemed
Table 1: Dose Groups
PROS [1 Groups Positive Control [5 Groups [6
[pros
|
PFOS vehicle control
|smtaetinv+alona_t__e| | statinvehicleconirol __|
In-VitroPhase
HMG-CoA reductase
TlihveerambiilcirtoysoofmPesF.OSMitcordoirseocmtelysiwnihlilbibteHpMreGp-arCeodAfrroemduucnttarseeatweildlfbeemaelxearmaitnsewdituhsianngdrat
`pwhiotshpohutopsroodtieuimn pflhuoosrpihdaet(aNsaeFa)ctiinviatlyl (buKfafteorsanadndBiasshsoapy,me1d9i7a2.) aNnadFthiunsh,ibiintcslusion of NaF
a(tpahlolspshtoargeysloatfeids)olraetdiuocntaasnedtpoutrhiefiaccattiivoen (suenvperheolsyphionrhyibliattsedth)efcoornmve(rNsoirodnstorfoimn,acRtoivdewell
ainncdubMaitticohnelpeenr,io1d9,7a7)s.saMyiecdrfoosrotmoteasl wainldl bacetidvoeseHdMaGs-lCisoteAd irnedtuabclteas1eabnyd,omfioslsliowoinnagnadn
inclusion, respectively, of NaF. Activity will be calculated as the rate.
(opfimciocmroolseso/mmailnuprtoet)eionf. foLrimpaitdiloenveolfs[o"fHtlhmeevvaarlioonuastedofsreodm m[i'cHroJsHoMmGe-sCwoilAl pbeermmeialsliugrreadm
uhespiantgoccoytmemse,rcaibailolmyaravkaeirlaobflHe kMiGts-aCndo/AorreHdPuLcCta.seCahcotilveisttye,rmolayprboeduucsteidonasinacsuelctounrdedary
measure of "in-vitro HMG-COA reductase inhibition.
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DP.aJ.geLu9eotfke1r6
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AMP levels To determine whether PFOS induces an increase in cellular AMP in-vitro, AMP levels of the microsomes will be quantified using HPLC.
AMP-PK dTeovealssoepsesdtbhey aDbaivliietys,ofCaPrFlOinSg taondactHadridreicetl(y19t8o9a)ctwihviatcehAuMsePs-aPsKynitnh-evtiitcrop,eaptpiedpetisdubesatrsastaey, `SeSqAuMeSncpeep(tSiedre7,9w)iwllhibcehusisedp.hoSspAhMorSylpaetpetdideexcwlaussidveevleylboypeAdMbPa-sPeKd.on AthMePa-miPnKo acid `FSatpelciivemrenass wdielslcrbiebperdebpyarCeadrlbiyngpaerttiaall.ly(1p9u8r9i)f.yiTnhgetsheescpyetcoisomleincspwrioltleibneftrraecattieodnaosfocuotnltirnoeld in table 1. Following an incubation period, AMP-PK activity will be measured based on
the incorporation of radioactivity from [y-** PJATP into the SAMS peptide (Davies ef al,
1989).
In-VivoPhase
HMG-CoA Reductase Activity in Pregnant Rats and Rat Pups
Iinsodliastsioocnoiaftcieolnolufltahresdurbufrgacftrioonmstthoe deinrzecytmley,evwahliucahtereHtMurGns-cCaotaAlyrtiecdaulcltyaascetaicvteiv(iCtoyrsriesnuiletts
rale.du1c9t9a5s)e.isTbhyedmeotsetrmfianvionrgabtlhee mine-tvhiovdo rtaotedeotfecitncionr-pvoivroatiinohniboiftipornecoufrsHoMrsG,-oCroeAxample.
acetate, into cholesterol (Corsini ef al, 1995).
conversion of "C-acetate into "*C-cholesterol
The potency ofPFOS
in-vivo, therefore, will
( inhibit the
be used as a
binihoimbairtikoenrotfo HdeMtGer-mCinoeAHrMedGu-ctCaoseAirnehidbuicttiaosneifnoudnadmisn.tThheeisne-vriefsruoltassssahyos.ulOdtpharearllelthe
`biomarkers which have been used successfully include urinary and plasma mevalonate
levels (Corsini et al, 1995).
tParbolgena1.ntPrriaotrswtiolclubtehanriezpaetaitodno,sdeadmdsurwiilnlg bgeesdtoatsieodnwaictchor"d"iCn-agcteotatthee. dHoaslef gofraolulpsdalimsstewdiilnl
gbeesataltlioownedantdo pcuopmsewtiolltbeermreamnodhvaeldf.wiIlflPbFeOcSutihsaaniHzMedGt-hCe omoAmriendugcotfasdeaiyn2hi1biotofr, the
rperdevuicoeuds msattuedrineaslswheoiugldhtbegaeiln,imiinncarteeadseodr satmiellliboorrnataendd idnetchreeamseevdalpounpatsuersviuvpapllseemeenntiend
o`asddeitgironoaulp.biPolmaasrmkaerasndo/foHrMurGi-neCmoaAybreeduccotlalseectaecdtiavnidtyainnatlhyezeddamfso.r mPelvaaslmoanamteevatloounsaeteas
levels wil inhibition
serve asthebiomarker to in cach litter. Liver (from
determine the cach dam and
degree pooled
of HMG-CoA for cach litter)
reductase will be used
for
'coAlMlePc-tPedKfarctoimveiatychdetdearmmianantdiopno,olleidvferorliepaidchanallityesrisfoarnPdFAOMSPdedteetremrimniantaitioonna.nSdesrearwuimll be 0,344 50
PDaJg.e L1u0e0t(ke1r6
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(LiLpDidL)anaalnydsitso.talLicvheolreasntedroslerwuilml hbiegdhedteenrsmiitnyeldipuospirnogtecionm(mHerDcLi)a,lllyowavdaeinlsaibtlyelkiiptosparontde/ionr
HEPnvLiCr.onmLeinvteralanLadbsoerrautomrPyFFOluSordienteerAmnianlayttiiocnalwCilhlembiespterryfToremaemd
by the 3M (FACT) headed
by
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Hapapnrsoepnr,iaPthe.mD.ethMyoodcfaorrdfiiablertiisnsjuuerywialnldb/eorexinafmlianmemdatbiyone.lectron microscopy (EM) or other
Liver AMP Levels in Pregnant Rats and Pups To determine whether PFOS induces an increase in cellular AMP levels in-vivo, liver AMP levels willbedetermined using HPLC.
AMP-PK Activity in PregnantRatsand Pups aAsMdPe-scPrKibsedpebcyimCearnlsiwniglletbael.pr(e1p98a9r)e.d AbyMPpa-rtPiKallayctpiuvriitfyywiinlgltbhee mcyetaossuorleicdpbraosteedinofnratchteion
incorporationofradioactivity from [1 PJATP into the SAMS peptide (Davies et al.,
1989).
SUMMARY
TbhyewhsitucdhyPoFutOlSinleedaidnstthoisrepdruocpeodsamliastaerinmaeldboadtygawieniignhgtkgnaoiwn,leidncgreeaisnetod tpheercmeencthsatniilslmb(osm) CanodAdercedruecatsaesde,puthpesruartvei-vlailmiitnirnagtse.nTzhyemehoyfpocthhoelseisstetrhaotl PsyFnOthSesaicst,swviialibnehiebxiatmioinneodf.HMGP(HFaOuSghhaosmparnedviSopuysdleyvboeledn,s1h9o92w)n.tSoedveorwanl HreMgGul-atCeoHAMrGe-duCcotaAsereidnuhicbtiatsoersac(tsitvaittiynsi)nhraavtse bale.e, n1s98h3o;wFnDtAo,in1d9u8c7e; dWeivsecloeptmaeln.,ta1l99e0fafe;cWtsissieemitlaalr.,to1t9h9o0sbe;sMeiennswkietrh ePtFalO.,S 1(9M9i0n;sHkrerabet eirmamle,di1a9t9e4;prFoDdAu,ct1o9f93H)M.GSt-uCdiocAs hraevduecftaosuen,dptrheavtecnot-saodrmiannitsatgroantiizoensovfamreivouasloonragtaen, the tetoxailc,iti1e9s8r8e;suKlotribngrufsrtometHaMl.G, -19C8o9;AMriendsukcetarseetianlh,ib1i9ti8o3n;iHnrraabtseatnald.,ra1b9b9i4t)s.(AMancaDtotneampltd `rweidllucbteasmeaadcetitvoitpyrweivlelntbePcFoOmSpatorxeidcibteytbwyeecon-dadammisniasntdratthienigr pmeuvpaslaonnadtteo.agHeMmGa-tcChoeAd dpiorseicttilvye oanndHnMeGga-tCivoeAcornetdruocltsa.seS,piencdiimreecntslywviila baecteivxaatmiionneodftAoMPd-eatecrtmiivnaetiefdPpFroOtSeiancts K`wihniacshe w(iAllMPbe-PfKol)loowretdhrwohuegrheipnodsusciiblneg,ainncilnucdreeabsoediyn AweMigPhtl,evleilvse.r Owetihgehrtp,alriavmeertaenrds, stoernuomrmPaFlOdSe,vellivoeprmaenndtaslcdriufmfelriepnicdessanindHmMyoGc-arCdoiaAl rfeidbuecritnajsueraycatinvdityi,nffeltaamlmaantdioena.rlDyue pfeotsatlnaantdalearartlsympaoystbneatmaluchhummaonrse(sLeenvsiintievfeat.o,H1M98G9-).CoTAhereredsuuclttassoefitnhhiisbisttiuodny,thtaankeanrewith `wmheacthsanailsrmeabdyywkhniocwhnhiagbhoudtosHeMsGof-CPoOASrceaduucsteaasdevaecrtsiveiteyf,fewcitlslihnpelrpegdneatnetrrmaitnse athned their
puuppss and the anderepotential risk PFOS holds to be adevelop"mental toxicant in humanPDsa..gJe. Lluoetfke1r6
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on21n000
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