Cell culture-based blood-brain barrier (BBB) models are useful experimental tools for developing central nervous system drugs. Several endothelial cell sources exist for BBB models, including primary cultured brain endothelial cells and immortalized cell lines. Among them, primary cell-based models are considered suitable for the functional analysis of the BBB; however, little is known about the utility of low-passage brain endothelial cells for this purpose. In this study, we investigated the effect of passage on brain endothelial cells from human, mouse and rat brain tissue as BBB models. We established in vitro BBB models using primary brain endothelial cells (Passage 1-Passage 4) from humans, mice, and rats. To analyze the effect of cell type on BBB function, we evaluated transendothelial electrical resistance (TEER) and performed immunofluorescence staining of tight junction proteins. Among the brain endothelial cell models, TEER was highest in the Passage 1 (P1) cell-based BBB model. There was no adequate increase in TEER in other low-passage cultures (P2-P4). A confluent, non-overlapping, uniform monolayer of cells in all P1 cell-based models was visible on immunostaining of tight junction proteins, whereas it was weak or undetectable in more passaged cultures. Increasing passages cultured of brain endothelial cells did not exhibit restrictive BBB function regardless of the cell source and despite culturing with pericytes and astrocytes. Among the tested culture models, only the lowest cultured cell-based models are suitable for functional analysis of the BBB.

中文翻译：

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血脑屏障体外模型的去分化率与细胞来源之间增加的通道的比较。

基于细胞培养的血脑屏障 (BBB) 模型是开发中枢神经系统药物的有用实验工具。BBB 模型存在多种内皮细胞来源，包括原代培养的脑内皮细胞和永生化细胞系。其中，基于原代细胞的模型被认为适用于 BBB 的功能分析；然而，关于低通道脑内皮细胞用于此目的的效用知之甚少。在这项研究中，我们研究了传代对来自人类、小鼠和大鼠脑组织的脑内皮细胞的影响，作为 BBB 模型。我们使用来自人类、小鼠和大鼠的原代脑内皮细胞（第 1 代至第 4 代）建立了体外 BBB 模型。为了分析细胞类型对 BBB 功能的影响，我们评估了跨内皮电阻 (TEER) 并对紧密连接蛋白进行了免疫荧光染色。在脑内皮细胞模型中，TEER 在基于通道 1 (P1) 细胞的 BBB 模型中最高。在其他低传代培养物 (P2-P4) 中，TEER 没有足够的增加。在所有基于 P1 细胞的模型中，在对紧密连接蛋白进行免疫染色时都可以看到融合的、不重叠的、均匀的单层细胞，而在更多传代培养物中它很弱或无法检测到。无论细胞来源如何，尽管与周细胞和星形胶质细胞一起培养，但越来越多的脑内皮细胞培养传代并未表现出限制性 BBB 功能。在测试的培养模型中，只有最低的培养细胞模型适用于 BBB 的功能分析。