title: A new blood-brain barrier model using primary rat brain endothelial cells, pericytes and astrocytes.
abstract: Blood-brain barrier (BBB) characteristics are induced and maintained by cross-talk between brain microvessel endothelial cells and neighbouring elements of the neurovascular unit. While pericytes are the cells situated closest to brain endothelial cells morphologically and share a common basement membrane, they have not been used in co-culture BBB models for testing drug permeability. We have developed and characterized a new syngeneic BBB model using primary cultures of the three main cell types of cerebral microvessels. The co-culture of endothelial cells, pericytes and astrocytes mimick the anatomical situation in vivo. In the presence of both pericytes and astrocytes rat brain endothelial cells expressed enhanced levels of tight junction (TJ) proteins occludin, claudin-5 and ZO-1 with a typical localization at the cell borders. Further morphological evidence of the presence of interendothelial TJs was provided by electron microscopy. The transendothelial electrical resistance (TEER) of brain endothelial monolayers in triple co-culture, indicating the tightness of TJs reached 400Omegacm(2) on average, while the endothelial permeability coefficients (P(e)) for fluorescein was in the range of 3x10(-6)cm/s. Brain endothelial cells in the new model expressed glucose transporter-1, efflux transporters P-glycoprotein and multidrug resistance protein-1, and showed a polarized transport of rhodamine 123, a ligand for P-glycoprotein. To further characterize the model, drug permeability assays were performed using a set of 19 compounds with known in vivo BBB permeability. Good correlation (R(2)=0.89) was found between in vitroP(e) values obtained from measurements on the BBB model and in vivo BBB permeability data. The new BBB model, which is the first model to incorporate pericytes in a triple co-culture setting, can be a useful tool for research on BBB physiology and pathology and to test candidate compounds for centrally acting drugs.
authors.affiliation: Department of Pharmacology, Nagasaki University Graduate School of Biomedical Sciences, Sakamoto, Japan.
authors.forename: Shinsuke
authors.initials: S
authors.lastname: Nakagawa
authors.forename: Mária A
authors.initials: MA
authors.lastname: Deli
authors.forename: Hiroko
authors.initials: H
authors.lastname: Kawaguchi
authors.forename: Takeshi
authors.initials: T
authors.lastname: Shimizudani
authors.forename: Takanori
authors.initials: T
authors.lastname: Shimono
authors.forename: Agnes
authors.initials: A
authors.lastname: Kittel
authors.forename: Kunihiko
authors.initials: K
authors.lastname: Tanaka
authors.forename: Masami
authors.initials: M
authors.lastname: Niwa
mesh_terms: D018435:ATP Binding Cassette Transporter, Subfamily B / Q000378:metabolism; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism; D000818:Animals; D001253:Astrocytes / Q000378:metabolism* / Q000648:ultrastructure; D001812:Blood-Brain Barrier / Q000378:metabolism* / Q000648:ultrastructure; D002196:Capillaries / Q000378:metabolism / Q000648:ultrastructure; D002352:Carrier Proteins / Q000378:metabolism; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D002536:Cerebral Arteries / Q000378:metabolism / Q000648:ultrastructure; D062507:Claudin-5; D042783:Endothelial Cells / Q000378:metabolism* / Q000648:ultrastructure; D008564:Membrane Potentials / Q000502:physiology; D008565:Membrane Proteins / Q000378:metabolism; D008833:Microcirculation / Q000502:physiology; D046529:Microscopy, Electron, Transmission; D008954:Models, Biological; D062793:Occludin; D020286:Pericytes / Q000378:metabolism* / Q000648:ultrastructure; D010750:Phosphoproteins / Q000378:metabolism; D051381:Rats; D020112:Rhodamine 123 / Q000493:pharmacokinetics; D019108:Tight Junctions / Q000378:metabolism / Q000648:ultrastructure; D062826:Zonula Occludens-1 Protein
chemical_list: D018435:ATP Binding Cassette Transporter, Subfamily B; D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; C107466:CLDN5 protein, human; D002352:Carrier Proteins; D062507:Claudin-5; D008565:Membrane Proteins; C568985:OCLN protein, human; D062793:Occludin; C568987:Ocln protein, rat; D010750:Phosphoproteins; C569042:TJP1 protein, human; C569044:Tjp1 protein, rat; D062826:Zonula Occludens-1 Protein; D020112:Rhodamine 123; C095122:multidrug resistance protein 3