Induction of blood-brain barrier properties in immortalized bovine brain endothelial cells by astrocytic factors.
The blood-brain barrier (B-BB) protects the free passage of substances into the brain and maintains the homeostasis of the central nervous system. It is commonly accepted that astrocytes surrounding brain endothelial cells influence the B-BB formation and the exhibition of B-BB function of capillaries. To begin the in vitro study on the B-BB, it is essential to obtain a homogenous and sufficient supply of brain endothelial cells as well as astrocytes. We thus immortalized the bovine brain endothelial cell (BBEC) by transfection of the SV40 large T antigen and obtained a single clone, t-BBEC-117, which retained the brain endothelial cell phenotype. Astrocyte in co-culture was found to tighten the intercellular contacts of the immortal cells resulting in a reduced L-glucose permeability, and its conditioned medium (CM) augmented a B-BB phenotype, alkaline phosphatase (ALP) activity. Among known astrocytic factors, only fibroblast growth factor-basic (bFGF) could mimic the actions of astrocytes as measured by L-glucose permeability and ALP activity. Moreover, anti-bFGF antibody canceled 90% of ALP activation by astrocyte CM. Basic FGF, however, failed to induce other B-BB phenotypes such as the expressions of multidrug resistance (mdr) and glucose transporter (GLUT-1) genes. These data suggest that bFGF is one of the most plausible astrocytic factors to induce the B-BB properties of immortal brain endothelial cells together with some unknown factors in the astrocyte CM.
Sobue
K
K
Department of Anesthesiology and Resuscitology, Nagoya City University Medical School, Nagoya, Japan. kazsobue@med.nagoya-cu.ac.jp
Yamamoto
N
N
Yoneda
K
K
Hodgson
M E
ME
Yamashiro
K
K
Tsuruoka
N
N
Tsuda
T
T
Katsuya
H
H
Miura
Y
Y
Asai
K
K
Kato
T
T
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000235:genetics; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D000943:Antigens, Differentiation / Q000096:biosynthesis; D000957:Antigens, Viral, Tumor / Q000235:genetics; D001253:Astrocytes / Q000166:cytology / Q000378:metabolism*; D001483:Base Sequence; D001812:Blood-Brain Barrier / Q000187:drug effects / Q000502:physiology*; D001921:Brain / Q000098:blood supply / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D002417:Cattle; D002463:Cell Membrane Permeability / Q000187:drug effects / Q000502:physiology; D002478:Cells, Cultured; D018920:Coculture Techniques; D017077:Culture Media, Conditioned / Q000378:metabolism; D004730:Endothelium, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D004789:Enzyme Activation / Q000187:drug effects; D005947:Glucose / Q000378:metabolism; D006133:Growth Substances / Q000378:metabolism* / Q000494:pharmacology; D008969:Molecular Sequence Data; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017386:Sequence Homology, Amino Acid; D014162:Transfection
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D000943:Antigens, Differentiation; D000957:Antigens, Viral, Tumor; D017077:Culture Media, Conditioned; D006133:Growth Substances; D000469:Alkaline Phosphatase; D005947:Glucose