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Expression and localization of claudins-3 and -12 in transformed human brain endothelium.doc

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Expression and localization of claudins-3 and -12 in transformed human brain endothelium

Schradeetal.FluidsandBarriersoftheCNS2012,9:6 /content/9/1/6 FLUIDS AND BARRIERS OF THE CNS SHORT PAPER OpenAccess Expressionandlocalizationofclaudins-3and-12 intransformedhumanbrainendothelium AnjaSchrade1,7,HadassahSade2,Pierre-OlivierCouraud3,4,IgnacioARomero5,BabetteBWeksler6and JensNiewoehner2* Abstract Background:TheaimofthisstudywastocharacterizethehCMEC/D3cellline,aninvitromodelofthehuman BloodBrainBarrier(BBB)fortheexpressionofbrainendothelialspecificclaudins-3and-12. Findings:hCMEC/D3cellsexpressclaudins-3and-12.Claudin-3isdistinctlylocalizedtotheTJwhereasclaudin-12 isobservedintheperinuclearregionandcompletelyabsentfromTJs.Weshowthattheexpressionofboth proteinsislostincellpassagenumberswheretheBBBpropertiesarenolongerfullyconserved.Expressionand localizationofclaudin-3isnotmodulatedbysimvastatinshowntoimprovebarrierfunctioninvitroandalso recommendedforroutinehCMEC/D3culture. Conclusions:Theseresultssupportconservationofclaudin-3and-12expressioninthehCMEC/D3celllineand makeclaudin-3apotentialmarkerforBBBcharacteristicsinvitro. Keywords:BloodBrainBarrier(BBB),hCMEC/D3,Claudins,Tightjunction,Statins Background The hCMEC/D3 cell line is the best-characterized in Membersoftheclaudinsuperfamilyareimportantpro- vitro model system of the human blood brain barrier. tein constituents of tight junctions (TJs) in polarized The cells express endothelial specific and TJ markers epithelial and endothelial monolayers and play an includingclaudins-1and-5[8,9].Theaimofthepresent important role in maintaining the paracellular barrier. studywastoestablishtheproteinexpressionandlocali- At the blood brain barrier (BBB), claudins-1,-3, -5 and zation pattern of the other brain endothelial specific -12havebeenshowntoparticipate intheformation of claudins-3and-12inthehCMEC/D3cellline. TJs between brain microvascular endothelial cells in vivo[1-4].Ofthesemembers,therehasbeenaparticu- Methods lar interest in claudin-5, as mice deficient for this pro- Cell culture