CMZ Reversed Chronic Ethanol-Induced Disturbance o

CMZReversedChronicEthanol-InducedDisturbanceofPPAR-aPossiblybySuppressingOxidativeStressandPGC-1aAcetylation,andActivatingtheMAPKandGSK3bPathwayTaoZeng,Cui-LiZhang,Fu-YongSong,Xiu-LanZhao,Ke-QinXie*InstituteofToxicology,SchoolofPublicHealth,ShandongUniversity,JinanCity,ShandongProvince,People’sRepublicofChinaAbstractBackground:CytochromeP4502E1(CYP2E1)hasbeensuggestedtoplaycriticalrolesinthepathogenesisofalcoholicfattyliver(AFL),buttheunderlyingmechanismsremainsunclear.ThecurrentstudywasdesignedtoevaluatewhetherCYP2E1suppressionbychlormethiazole(CMZ)couldsuppressAFLinmice,andtoexploretheunderlyingmechanisms.Methods:MiceweretreatedwithorwithoutCMZ(50mg/kgbw,i.p.)andsubjectedtoliquiddietwithorwithoutethanol(5%,w/v)for4weeks.Biochemicalparametersweremeasuredusingcommercialkits.TheproteinandmRNAlevelsweredetectedbywesternblotandqPCR,respectively.Histopathologyandimmunohistochemicalassaywereperformedwithroutinemethods.Results:CYP2E1inhibitionbyCMZcompletelyblockedAFLinmice,shownasthedeclineofthehepaticandserumtriglyceridelevels,andthefewerfatdropletsintheliversections.ChronicethanolexposureledtosignificantdecreaseofthemRNAandproteinlevelsofperoxisomeproliferator-activatedreceptora(PPAR-a),whichwasblockedbyCMZco-treatment.CMZco-treatmentsuppressedethanol-inducedoxidativestress,overproductionoftumornecrosisa(TNF-a),anddecreaseofproteinlevelsofthePPAR-aco-activatorsincludingp300anddeacetylatedPGC1-a.Furthermore,CMZco-treatmentledtotheactivationofAMP-activatedproteinkinase(AMPK),mitogen-activatedproteinkinase(MAPK),andPI3K/Akt/GSK3bpathway.However,chronicethanol-induceddeclineofacyl-CoAcarboxylase(ACC)andfattyacidsynthase(FAS)proteinlevelswaspartiallyrestoredbyCMZ,whiletheactivationofautophagyappearedtobesuppressedbyCMZ.Conclusion:TheseresultssuggestedthatCMZsuppressedchronicethanol-inducedoxidativestress,TNF-aoverproduction,declineofp300proteinlevelanddeacetylationofPGC1-a,andactivatedAMPK,MAPK,andPI3K/Akt/GSK3bpathway,whichmightcontributetotheactivationofPPAR-aandaccountfortheprotectionofCMZagainstAFL.Citation:ZengT,ZhangC-L,SongF-Y,ZhaoX-L,XieK-Q(2014)CMZReversedChronicEthanol-InducedDisturbanceofPPAR-aPossiblybySuppressingOxidativeStressandPGC-1aAcetylation,andActivatingtheMAPKandGSK3bPathway.PLoSONE9(6):e98658.doi:10.1371/journal.pone.0098658Editor:IrinaU.Agoulnik,FloridaInternationalUniversity,UnitedStatesofAmericaReceivedMarch23,2014;AcceptedMay5,2014;PublishedJune3,2014Copyright:2014Zengetal.Thisisanopen-accessarticledistributedunderthetermsoftheCreativeCommonsAttributionLicense,whichpermitsunrestricteduse,distribution,andreproductioninanymedium,providedtheoriginalauthorandsourcearecredited.DataAvailability:Theauthorsconfirmthatalldataunderlyingthefindingsarefullyavailablewithoutrestriction.AlldataareincludedwithintheSupportingInformationfiles.Funding:ThisworkwassupportedbytheNationalScienceFoundationofChina(GrantNo.81102153),theSpecialFinancialGrantfromtheChinaPostdoctoralScienceFoundation(GrantNo.201104626),andPostdoctoralScienceFoundationfundedProjectofShandongProvince(GrantNo.201002020).Thefundershadnoroleinstudydesign,datacollectionandanalysis,decisiontopublish,orpreparationofthemanuscript.CompetingInterests:Theauthorshavedeclaredthatnocompetinginterestsexist.*E-mail:[e-mail]iverdisease(ALD)remainstobeoneofthemostcommonetiologiesofliverdiseasesandisamajorcauseofmorbidityandmortalityworldwide[1].ALDisapathologicalprocesscharacterizedbyprogressiveliverdamagefromsteatosistosteatohepatitis,fibrosisandfinallycirrhosis[2].Alcoholicfattyliver(AFL)istheearliestandmostcommonpathologicalformofALD.AlthoughAFLwasconsideredbenigninthepast,recentstudieshavedemonstratedthatfataccumulationrendersthelivermorevulnerabletotoxinssuchasendotoxin,andAFLiseasilyprogressedtohepatitis,fibrosisandevencirrhosis[3].ItwasreportedthatAFLdevelopedinabout90%ofindividualswhodrankmorethan60g/dayofalcohol,butmightalsooccurinindividualswhodrankless[4,5].Otherstudiessuggestedthatprogressiontofibrosisandcirrhosisoccurredin5%–15%ofAFLpatientsdespiteabstinence[6,7].Thus,AFLshouldbetheoptimalintervenephaseforthepreventionofALDtomoreseriousphasessuchasfibrosisandcirrhosis[8].ThemechanismsforAFLhavebeenextensivelystudiedinthepastdecades,andseveralimportantfactorsincludingperoxisomeproliferator-activatedreceptora(PPAR-a),sterolregulatoryelement-bindingprotein-1c(SREBP-1c),AMP-activatedproteinkinase(AMPK),autophagy,cytochromeP4502E1(CYP2E1),andcytokinessuchasadiponectinandtumornecrosisa(TNF-a)haveallbeenproposedtobeinvolvedinthedevelopmentofAFL[2,9–PLOSONE||Volume9|Issue6|e9865811].Amongthesefactors,CYP2E1hasgainedgreatattentionbecauseitcouldmediatethemetabolismofethanol,togeneratereactiveoxygenspecies(ROS)andtobeinducedbyethanol[12].CYP2E1isamemberofthecytochromeP450superfamily,whichisagroupofheme-containingproteinswithmultiplefunctionsincludingthemetabolismofxenobioticsincludingdrugs,toxins,carcinogens,andendogenoussubstrates[13].CYP2E1couldcatalyzethetwoelectronoxidationofethanoltoacetalde