建立血脑屏障体外模型并进行功能表征，该模型由脑微血管内皮细胞和星形胶质细胞共培养组成。

Establishment and functional characterization of an in vitro model of the blood-brain barrier, comprising a co-culture of brain capillary endothelial cells and astrocytes.

作者信息

Gaillard P J, Voorwinden L H, Nielsen J L, Ivanov A, Atsumi R, Engman H, Ringbom C, de Boer A G, Breimer D D

机构信息

Department of Pharmacology, Leiden/Amsterdam Center for Drug Research (LACDR), Leiden University, P.O. Box 9503, 2300 RA, Leiden, The Netherlands.

出版信息

Eur J Pharm Sci. 2001 Jan;12(3):215-22. doi: 10.1016/s0928-0987(00)00123-8.

DOI:10.1016/s0928-0987(00)00123-8

PMID:11113640

Abstract

OBJECTIVE

The aim was to establish a flexible, abundantly available, reproducible and functionally characterized in vitro model of the blood-brain barrier (BBB).

METHODS

In a first step, bovine brain capillaries and newborn rat astrocytes were isolated. Subsequently, a co-culture of primary brain capillary endothelial cells (BCEC) on semi-permeable filter inserts, with astrocytes on the bottom of the filter was established. The cell material was characterized on the basis of specific cell-type properties and (functional expression of) specific BBB properties.

RESULTS

BCEC displayed: (1) characteristic endothelial cell morphology; (2) expression of endothelial cell markers (i.e., CD51, CD62P, CD71 and cadherin 5); (3) marginal F-actin localization; (4) tight junction formation between the cells; (5) expression of gamma-glutamyl-transpeptidase (gamma-GTP); (6) expression of P-glycoprotein (Pgp); (7) functional transendothelial transferrin transport and uptake; (8) restriction of paracellular transport; and (9) high transendothelial electrical resistance (TEER). Astrocytes displayed characteristic astrocyte morphology and expressed glial fibrillary acidic protein (GFAP). Co-culture with astrocytes increased TEER and decreased paracellular transport. In addition, expression of the glucocorticoid receptor (GR) was demonstrated in the endothelial cells of the BBB, while no expression of the mineralocorticoid receptor (MR) was found.

CONCLUSIONS

A high quality and mass-production in vitro BBB model was established in which experiments with physiological (e.g., regulation of BBB permeability), pharmacological (e.g., pharmacokinetics and pharmacodynamics) and pathophysiological (e.g., disease influence on BBB permeability) objectives can be reproducibly performed.

摘要

目的

旨在建立一种灵活、易于获取、可重复且具有功能特征的血脑屏障（BBB）体外模型。

方法

第一步，分离牛脑毛细血管和新生大鼠星形胶质细胞。随后，在半透膜滤器上建立原代脑毛细血管内皮细胞（BCEC）与滤器底部星形胶质细胞的共培养体系。根据特定细胞类型特性和特定BBB特性（的功能表达）对细胞材料进行表征。

结果

BCEC表现出：（1）特征性内皮细胞形态；（2）内皮细胞标志物（即CD51、CD62P、CD71和钙黏蛋白5）的表达；（3）边缘F-肌动蛋白定位；（4）细胞间紧密连接形成；（5）γ-谷氨酰转肽酶（γ-GTP）的表达；（6）P-糖蛋白（Pgp）的表达；（7）功能性跨内皮转铁蛋白转运和摄取；（8）细胞旁转运受限；（9）高跨内皮电阻（TEER）。星形胶质细胞表现出特征性星形胶质细胞形态并表达胶质纤维酸性蛋白（GFAP）。与星形胶质细胞共培养可增加TEER并减少细胞旁转运。此外，在BBB的内皮细胞中证实了糖皮质激素受体（GR）的表达，而未发现盐皮质激素受体（MR）的表达。