Blood-brain barrier-specific properties of a human adult brain endothelial cell line

• Published in: The FASEB journal Vol. 19; no. 13; pp. 1872 - 1874
• Main Authors: Weksler, B. B, Subileau, E. A, Perrière, N, Charneau, P, Holloway, K, Leveque, M, Tricoire-Leignel, H, Nicotra, A, Bourdoulous, S, Turowski, P, Male, D. K, Roux, F, Greenwood, J, Romero, I. A, Couraud, P. O
• Format: Journal Article
• Language: English
• Published: United States Federation of American Societies for Experimental Biology 01-11-2005
• Subjects: Agar - chemistry; Animals; Antigens, CD; Antigens, Polyomavirus Transforming - biosynthesis; Antigens, Polyomavirus Transforming - genetics; Blood-Brain Barrier - drug effects; Blotting, Western; Brain - cytology; Brain - drug effects; Brain - metabolism; Brain - pathology; Cadherins - biosynthesis; Capillaries - pathology; Cattle; Cell Adhesion; Cell Culture Techniques - methods; Cell Line; Cell Proliferation; Cells, Cultured; cellular junctions; Cloning, Molecular; Collagen - pharmacology; Cytokines - metabolism; Drug Combinations; drug permeability; Drug Resistance, Multiple; Endothelial Cells - cytology; Endothelial Cells - pathology; Endothelium, Vascular - cytology; Endothelium, Vascular - pathology; Female; Flow Cytometry; Humans; Immunohistochemistry; in vitro model; Karyotyping; Laminin - pharmacology; Lentivirus - genetics; Lymphocytes - metabolism; Microscopy, Fluorescence; Models, Biological; multidrug resistance; Perfusion; Permeability; Phenotype; Platelet Endothelial Cell Adhesion Molecule-1 - biosynthesis; Proteoglycans - pharmacology; Rats; Reverse Transcriptase Polymerase Chain Reaction; RNA - metabolism; Signal Transduction; Telomerase - genetics; Telomerase - metabolism; Time Factors; Up-Regulation; von Willebrand Factor - biosynthesis
• ISSN: 0892-6638; 1530-6860
• DOI: 10.1096/fj.04-3458fje

ABSTRACTEstablishment of a human model of the blood‐brain barrier has proven to be a difficult goal. To accomplish this, normal human brain endothelial cells were transduced by lentiviral vectors incorporating human telomerase or SV40 T antigen. Among the many stable immortalized clones obtained by sequential limiting dilution cloning of the transduced cells, one was selected for expression of normal endothelial markers, including CD31, VE cadherin, and von Willebrand factor. This cell line, termed hCMEC/D3, showed a stable normal karyotype, maintained contact‐inhibited monolayers in tissue culture, exhibited robust proliferation in response to endothelial growth factors, and formed capillary tubes in matrix but no colonies in soft agar. hCMEC/D3 cells expressed telomerase and grew indefinitely without phenotypic dedifferentiation. These cells expressed chemokine receptors, up‐regulated adhesion molecules in response to inflammatory cytokines, and demonstrated blood‐brain barrier characteristics, including tight junctional proteins and the capacity to actively exclude drugs. hCMEC/D3 are excellent candidates for studies of blood‐brain barrier function, the responses of brain endothelium to inflammatory and infectious stimuli, and the interaction of brain endothelium with lymphocytes or tumor cells. Thus, hCMEC/D3 represents the first stable, fully characterized, well‐differentiated human brain endothelial cell line and should serve as a widely usable research tool.