title: Blood-brain barrier properties in vitro depend on composition and assembly of endogenous extracellular matrices.
abstract: Brain capillary endothelial cells, which constitute the blood-brain barrier (BBB), are enveloped by the extracellular matrix (ECM) produced by endothelial cells, pericytes and astrocytes. The contribution of matrix components secreted by the various cell types at the neurovascular unit, however, remains unclear with respect to their effect on endothelial barrier function. In this study, a new in vitro model was established by growing endothelial cells on an ECM produced by pericytes, astrocytes or a serial combination of both. The last-mentioned was found to be more in vivo-like. We investigated the role of the composition and morphology of ECM supra-structures in maintaining BBB function. The composition was analysed by protein analysis (enzyme-linked immunosorbent assay) and the ultrastructure of generated matrices was analysed by transmission electron microscopy including immunogold labelling. We could show by electric cell-substrate impedance sensing measurements that pericytes and combined matrices significantly improved the barrier tightness of porcine brain capillary endothelial cells (PBCEC). The increase of the resistance was verified by enhanced expression of tight junction proteins. Thus, for the first time, we have shown that barrier integrity is strictly controlled by the ECM, which is a product of all cells involved in the secretion of ECM components and their modification by corresponding cells. Moreover, we have demonstrated that complex matrices by the various cells of the BBB induce barrier marker enzymes in PBCEC, such as alkaline phosphatase.
authors.affiliation: Institute of Biochemistry, University of Muenster, 48149, Muenster, Germany.
authors.forename: Kathrin
authors.initials: K
authors.lastname: Zobel
authors.affiliation: Institute of Experimental Musculoskeletal Medicine, University Hospital Muenster, 48149, Muenster, Germany.
authors.forename: Uwe
authors.initials: U
authors.lastname: Hansen
authors.affiliation: Institute of Biochemistry, University of Muenster, 48149, Muenster, Germany. gallah@uni-muenster.de.
authors.forename: Hans-Joachim
authors.initials: HJ
authors.lastname: Galla
mesh_terms: D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D001812:Blood-Brain Barrier / Q000378:metabolism*; D001921:Brain / Q000098:blood supply; D002196:Capillaries / Q000166:cytology; D062507:Claudin-5 / Q000378:metabolism; D024141:Collagen Type IV / Q000378:metabolism; D017097:Electric Impedance; D042783:Endothelial Cells / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D005109:Extracellular Matrix / Q000378:metabolism*; D005353:Fibronectins / Q000378:metabolism; D015151:Immunoblotting; D020782:Matrix Metalloproteinases / Q000378:metabolism; D062793:Occludin / Q000378:metabolism; D034421:Sus scrofa; D062826:Zonula Occludens-1 Protein / Q000378:metabolism
chemical_list: D015415:Biomarkers; D062507:Claudin-5; D024141:Collagen Type IV; D005353:Fibronectins; D062793:Occludin; D062826:Zonula Occludens-1 Protein; D000469:Alkaline Phosphatase; D020782:Matrix Metalloproteinases
keywords: Blood-brain barrier; Collagen IV; Electric cell–substrate impedance sensing; Electron microscopy; Fibronectin