REGULATION OF GAMMA-GLUTAMYL-TRANSPEPTIDASE AND ALKALINE-PHOSPHATASE ACTIVITIES IN IMMORTALIZED RAT-BRAIN MICROVESSEL ENDOTHELIAL-CELLS
REGULATION OF GAMMA-GLUTAMYL-TRANSPEPTIDASE AND ALKALINE-PHOSPHATASE ACTIVITIES IN IMMORTALIZED RAT-BRAIN MICROVESSEL ENDOTHELIAL-CELLS
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DOI：
10.1002/jcp.1041590114
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发表时间：
1994-04-01
影响因子：
5.6
通讯作者：
COURAUD, PO
中科院分区：
文献类型：
--
作者：
ROUX, F;DURIEUTRAUTMANN, O;COURAUD, PO
将含有E1A腺病毒基因的质粒导入大鼠脑微血管内皮细胞，使其永生化。对一个名为RBE4的克隆进行了进一步鉴定。这些细胞表现为未转化的表型，表达典型的内皮标记物、因子VIII相关抗原和单叶巴氏杆菌结合部位。当RBE4细胞在碱性成纤维细胞生长因子的存在和胶原蛋白覆盖的培养皿中生长时，融合培养物产生的芽延伸到单层以上并组织成三维结构。血脑屏障相关酶，γ-谷氨酰转肽酶(GammaGTP)的活性在这些结构中表达，而不是在周围的单层中表达。微血管相关酶碱性磷酸酶(ALP)也得到了类似的结果。增加细胞内cAMP的药物减少了三维结构的形成，但聚集体内的每个细胞仍然表达伽马GTP和碱性磷酸酶的活性。这些结构与高水平的GTP和ALP活性有关，也被星形胶质细胞因子诱导，包括(1)新生大鼠原代星形胶质细胞或大鼠胶质瘤C6细胞的质膜，(2)C6条件培养液，或(3)在RBE4细胞存在下生长的原代星形胶质细胞产生的扩散因子。因此，RBE4细胞对血管生成和星形胶质细胞因子表达血脑屏障相关的GTP活性和ALP活性仍然敏感，并可能构成有价值的血脑屏障体外模型的基础。(C)1994年Wiley-Liss，Inc.
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 (gammaGTP), 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 gammaGTP and ALP activities. Such structures, associated with high levels of gammaGTP 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 gammaGTP activity, as well as for ALP activity, and could constitute the basis of a valuable in vitro model of the blood-brain barrier. (C) 1994 Wiley-Liss, Inc.