Hayashi Y, Nomura M, Yamagishi S, Harada S, Yamashita J, Yamamoto H
Department of Biochemistry, Kanazawa University School of Medicine, Japan.
Glia. 1997 Jan;19(1):13-26.
Vascular endothelial cells (EC) exhibit organ-to-organ heterogeneity in their functions and morphologies. In particular, brain capillary EC have unique characteristics exemplified by the blood-brain barrier (BBB). The formation and the maintenance of BBB have been ascribed to EC responses to inductive signal(s) or factor(s) from astrocytes that encircle microvessels in the central nervous system. These EC responses were demonstrated in numerous in vivo studies, exemplified by those of Janzer and Raff (Nature 325:253, 1987) and Tout et al. (Neuroscience 55:291, 1993) showing that transplanted astrocytes induced BBB properties in non-neural vascular EC. In this study, we constructed a heterologous co-culture system, in which rat fetal brain astrocytes were cultivated on one surface of a porous membrane and human umbilical vein EC on the opposite surface. Electron microscopic examination revealed that astrocytes passed their endfeet through the pores, making contact with EC. In this system, gamma-glutamyltranspeptidase (gamma-GTP) activity in EC was found to be significantly increased by contacting astrocytes in a density- and time-dependent manner, but not when the astrocyte feeder layer was apart from EC or replaced by COS cells; astrocyte-derived extracellular matrix partially activated gamma-GTP. mRNAs for some of the representative BBB markers, including transferrin receptor, P-glycoprotein, brain-type glucose transporter (GLUT-1), and gamma-GTP were also demonstrated by reverse transcription-polymerase chain reaction to be upregulated in EC co-cultured with astrocytes. Astrocyte inductions of close membrane apposition resembling a zonula occludens and of an increase in the content of mitochondria in EC were also noted in electron micrographs. Furthermore, an increased barrier activity against inulin was conferred on EC when they were lined with astrocytes. The results obtained with this heterologous co-culture system thus indicate that through contact with their feet, astrocytes are capable of transdifferentiating non-neural EC into the brain type, endowing them with the BBB properties.
血管内皮细胞（EC）在功能和形态上表现出器官间的异质性。特别是，脑毛细血管内皮细胞具有以血脑屏障（BBB）为代表的独特特征。血脑屏障的形成和维持归因于内皮细胞对来自环绕中枢神经系统微血管的星形胶质细胞的诱导信号或因子的反应。这些内皮细胞反应在众多体内研究中得到证实，例如扬泽和拉夫（《自然》325:253，1987年）以及图特等人（《神经科学》55:291，1993年）的研究，这些研究表明移植的星形胶质细胞可诱导非神经血管内皮细胞产生血脑屏障特性。在本研究中，我们构建了一种异源共培养系统，其中大鼠胎儿脑星形胶质细胞在多孔膜的一个表面培养，人脐静脉内皮细胞在相对的表面培养。电子显微镜检查显示，星形胶质细胞将其终足穿过孔隙，与内皮细胞接触。在该系统中，发现内皮细胞中的γ-谷氨酰转肽酶（γ-GTP）活性通过与星形胶质细胞接触以密度和时间依赖性方式显著增加，但当星形胶质细胞饲养层与内皮细胞分开或被COS细胞取代时则不会增加；星形胶质细胞衍生的细胞外基质部分激活了γ-GTP。通过逆转录-聚合酶链反应还证明，与星形胶质细胞共培养的内皮细胞中，包括转铁蛋白受体、P-糖蛋白、脑型葡萄糖转运蛋白（GLUT-1）和γ-GTP在内的一些代表性血脑屏障标志物的mRNA上调。电子显微镜照片中还注意到星形胶质细胞诱导内皮细胞膜紧密贴附，类似于紧密连接，并且内皮细胞中线粒体含量增加。此外，当内皮细胞与星形胶质细胞排列在一起时，其对菊粉的屏障活性增加。因此，用这种异源共培养系统获得的结果表明，通过与终足接触，星形胶质细胞能够将非神经内皮细胞转分化为脑型，赋予它们血脑屏障特性。
