Regulation of gamma‐glutamyl transpeptidase and alkaline phosphatase activities in immortalized rat brain microvessel endothelial cells

Abstract

Rat brain microvessel endothelial cells were immortalized by transfection with a plasmid containing the E1A adenovirus gene. One clone, called RBE4, was further characterized. These cells display a nontransformed phenotype and express typical endothelial markers, Factor VIII-related antigen and Bandeiraea simplicifolia binding sites. When RBE4 cells were grown in the presence of bFGF and on collagen-coated dishes, confluent cultures developed sprouts that extend above the monolayer and organized into three-dimensional structures. The activity of the blood-brain barrier-associated enzyme, gamma-glutamyl transpeptidase (gamma GTP), was expressed in these structures, not in the surrounding monolayer. Similar results were obtained with the microvessel-related enzyme alkaline phosphatase (ALP). Addition of agents that elevate intracellular cAMP reduced the formation of three-dimensional structures, but every cell inside the aggregates still expressed gamma GTP and ALP activities. Such structures, associated with high levels of gamma GTP and ALP activities, were also induced by astroglial factors, including (1) plasma membranes from newborn rat primary astrocytes or rat glioma C6 cells, (2) C6 conditioned media, or (3) diffusible factors produced by primary astrocytes grown in the presence of, but not in contact with RBE4 cells. RBE4 cells thus remain sensitive to angiogenic and astroglial factors for the expression of the blood-brain barrier-related gamma GTP activity, as well as for ALP activity, and could constitute the basis of a valuable in vitro model of the blood-brain barrier.

Stimulatory Effects of Thyroid Hormone on Brain Angiogenesis in vivo and in vitro

“…The immortalized rat brain EC line RBE4 was a kind gift from Dr F Roux, and was cultured as described previously (Roux et al, 1994). Briefly, culture plates or flasks were coated with 100 mg/mL type I collagen (Roche Molecular Biochemical, Stockholm, Sweden).…”

ICAM-1-Coupled Cytoskeletal Rearrangements and Transendothelial Lymphocyte Migration Involve Intracellular Calcium Signaling in Brain Endothelial Cell Lines

“…RBE4 cells were isolated from rat brain cortex and immortalized with the plasmid pE1A-neo, containing the adenovirus E1A-encoding sequence followed by a neomycin-resistance gene (18). GP8 cells were isolated from rat brain cortex and immortalized with SV40 large T-Ag (19).…”

Manganese accumulates primarily in nuclei of cultured brain cells

“…Thus, the purpose of this study was to determine and compare the subcellular distribution of Mn in cell types that are important for Mn transport and toxicity. We chose choroidal epithelial Z310 cells established from the choroid plexus of the BCB (Zheng and Zhao, 2002), brain endothelium RBE4 cells derived from rat brain endothelium of the BBB (Roux et al, 1994), mesencephalic dopaminergic neuronal (N27) cells (Prasad et al, 1994), and pheochromocytoma PC12 cells derived from rat adrenal medulla (Greene and Tischler, 1976), for this study.…”