四溴二苯醚对人甲状腺细胞功能的影响蒋潇

2008-12-232009-01-09No.LJ200619E-mail:[e-mail];*CorrespondingauthorE-mail:[e-mail]*210029BDE-4710-12、10-1010-8mol·L-1BDE-47TEC24hTgTg8Pax-8.10-12、10-10mol·L-1Tgp0.0510-8mol·L-1BDE-47Tgp0.05.TgPax-8BDE-47p0.05-.BDE-47TgPax-8.BDE-4781673-589720092-231-06X171.5X503AEffectsofBDE-47ontheFunctionofCulturedHumanThyroidCellsJIANGXiaoLIUCui-pingMAOXiao-dongZHUQunLIUChao*DepartmentofEndocrinology,TheFirstAffiliatedHospital,NanjingMedicalUniversity,Nanjing210029Received23December2008accepted9January2009AbstractInordertoinvestigatetheeffectsofBDE-47onthefunctionofhumanthyroidcellsinvitro,thyroidfollicularepitheliumcellsTECwereculturedinthedifferentconcentrationsofBDE-470,10-12,10-10,10-8mol·L-1for24h.ThesecretoryfunctionofTECwasdetectedviathemeasurementofconcentrationsofthyroglobulinTgbyelectrochemiluminescenceimmunoassayECLIA.Theexpressionsofthyroidfunction-relatedTggeneandpairedboxgene8Pax-8wereassessedbysemi-quantitativeRT-PCR.ResultsshowedthattheTgconcentrationsinsupernatantsdecreasedsignificantlywhentheconcentrationsofBDE-47were10-12and10-10mol·L-1comparedwiththecontrolgroup,p0.05.However,Therewasnosignificantdifferencebetweenthe10-8mol·L-1groupandthecontrolgroupp0.05.TheexpressionsofTgandPax-8genesdecreasedsignificantlywiththeincreaseofBDE-47concentrationsp0.05.Thereweredose-effectrelationshipsbetweentheconcentrationsofBDE-47andtheexpressionsofTgandPax-8genes.BDE-47couldinhibitthefunctionofTECsignificantly,andthemechanismwasprobablyrelatedtodownregulationofexpressionsofTgandPax-8genes.KeywordsBDE-47humanthyroidcellsenvironmentalendocrinedisruptorsfunctionthyroglobulinTgpairedboxgene8Pax-8200942231-236AsianJournalofEcotoxicologyVol.4,2009No.2231-23641IntroductionEnvironmentalendocrinedisruptors,EEDsColborn,2002.EEDs、、Julanderetal.,2005;Dasetal.,2006.EEDsPolychlorinatedbipheny1s,PCBsPolybrominateddiphenylethers,PBDEsBarberetal.,2006;Bietal.,2006;2007;2006.PBDEs、、PCBsDasetal.,2006;Fernieetal.,2005.PBDEsFernieetal.,2005;Stapletonetal.,2005.PBDEsPBDEs.PBDEs/2008//.PBDEs.2,2′,4,4′-BDE-47、、PBDEsBietal.,2006BDE-47.TgLacroixetal.,2006TgPax-8Tg、Joetal.,2008;Delado觕yetal.,2007.TECTgBDE-47.2Materialsandmethods2.1BDE-47AccuStandardAnnexinV-FITCApoptosisDetectionKitIB&DBiosciencesTrizolRocheRT-PCRToyoboTSH、DMSOSigmaDMEM、FCSGibco.2.22.2.130minD-Hank’s1mm×1mm×1mm0.25%300U37℃60~90minD-Hank’s1:1.200HG-DMEM10%FCS1mU·mL-1TSH100U·mL-1100U·mL-15×105·mL-1.37℃2d1~2d3~5d.2.2.2TgTEC62mL2.0×105537℃、5%CO224h12h.Barber2006Hamers2006DMSOBDE-4710-12、10-10、10-8mol·L-1DMSO24hTg2006.2.2.3RT-PCRTgPax-825mL5mL1.0×1062.2.2.TrizolRNARNA1.8~2.0RNA2322RT-PCRcDNAPCR.PCR1.94℃30sTg55.3℃/Pax-862℃/GAPDH59℃30s72℃45s3272℃10min.RT-PCR2%.GAPDH3.2.3±x±sSPSS16.0p0.05.3Results3.1BDE-47TgBDE-47Tg1.24hBDE-47Tg.10-12mol·L-110-10mol·L-1Tgp0.05p0.0510-8mol·L-1Tgp0.05.3.2BDE-47BDE-472.2A、2BBDE-47TgPax-8-p0.05.1Table1PrimersforRT-PCRGenePrimersequences5′-3′Fragment/bpTgACCTGGGGCTGTGAGATGTATGTTAGATTCCTGGCTTCCGGTAGATGT125Pax-8GTGACAATGACACTGTGCCCCTCGTAGCTAAGGGTCTCGT280GAPDHACCACAGTCCATGCCATCACTCCACCACCCTGTTGCTGTA456A200bp100bp500bp400bp300bp200bp500bp400bpTg125bpGAPDH456bpPax-8280bpGAPDH456bp010-1210-1010-8BDE-47ConcentrationofBDE-47/mol·L-123344Discussion.Welshons2006EEDs———、.BDE-47Julander2005She2007BDE-47.、--.PBDEs20055“”.PBDEsFernieetal.,2005;Bietal.,2006;Boasetal.,2006PBDEsTT4FT4PBDEsStaskaletal.,2006;Darnerudetal.,2007.、、、、--Boasetal.,2006;Staskaletal.,2006;Darnerudetal.,2007;TabbandBlumberg,2006.BDE-47.TgBDE-47BDE-47.BDE-4710-8mol·L-1Tgp0.05BDE-47Tg.BDE-47Tg.3BDE-47TgBDE-47Tg.2342Pax-8Tg、Delado觕yetal.,2007.Pax-8.BDE-47TgBDE-47———TgPax-8.BDE-47..、、.、7562SCI16234.ReferencesBarberJL,WalshMJ,HewittR,JonesKC,MartinFL.2006.Low-dosetreatmentwithpolybrominateddiphenylethersPBDEsinducealteredcharacteristicsinMCF-7cellsJ.Mutagenesis,215:351-360BiX,QuW,ShengG,ZhangW,MaiB,ChenD,YuL,FuJ.2006.PolybrominateddiphenylethersinSouthChinamaternalandfetalbloodandbreastmilkJ.EnvironmentalPollution,1443:1024-1030BoasM,Feldt-RasmussenU,SkakkebaekNE,MainKM.2006.EnvironmentalchemicalsandthyroidfunctionJ.EuropeanJournalofEndocrinology,1545:599-611ColbornT.2002.CluesfromwildlifetocreateanassayforthyroidsystemdisruptionJ.EnvironmentalHealthPrespectives,110Suppl.3:363-367DarnerudPO,AuneM,LarssonL,HallgrenS.2007.PlasmaPBDEandthyroxinelevelsinratsexposedtoBromkalorBDE-47J.Chemosphere,679:S386-S392DasK,VossenA,TolleyK,VíkingssonG,ThronK,MüllerG,Baumg覿rtnerW,SiebertU.2006.Interfollicularfibrosisinthethyroidoftheharbourporpoise:anendocrinedisruption?J.ArchivesofEnvironmentalContaminationandToxicology,514:720-729Delado觕yJ,VassartG,VanVlietG.2007.Possiblenon-MendelianmechanismsofthyroiddysgenesisJ.EndocrineDevelopment,10:29-42FernieKJ,ShuttJL,MayneG,HoffmanD,LetcherRJ,DrouillardKG,RitchieIJ.2005.ExposuretopolybrominateddiphenylethersPBDEs:changesinthyroid,vitaminA,glutathionehomeostasis,andoxidativestressinAmericanKestrelsFalcosparveriusJ.ToxicologicalSciences,882:375-383HamersT,KamstraJH,SonneveldE,MurkAJ,KesterMH,AnderssonPL,LeglerJ,BrouwerA.2006.Invitroprofilingoftheendoc