Ledwig Verena, Reichl Stephan
Center of Pharmaceutical Engineering (PVZ), Technische Universität Braunschweig, Franz-Liszt-Straße 35A, 38106 Braunschweig, Germany.
Institute of Pharmaceutical Technology and Biopharmaceutics, Technische Universität Braunschweig, Mendelssohnstraße 1, 38106 Braunschweig, Germany.
Pharmaceutics. 2023 Jun 8;15(6):1688. doi: 10.3390/pharmaceutics15061688.
The blood-brain barrier (BBB) is the bottleneck in the development of new drugs to reach the brain. Due to the BBB, toxic substances cannot enter the brain, but promising drug candidates also pass the BBB poorly. Suitable in vitro BBB models are therefore of particular importance during the preclinical development process, as they can not only reduce animal testing but also enable new drugs to be developed more quickly. The aim of this study was to isolate cerebral endothelial cells, pericytes, and astrocytes from the porcine brain to produce a primary model of the BBB. Additionally, as primary cells are well suited by their properties but the isolation is complex and better reproducibility with immortalized cells must be ensured, there is a high demand for immortalized cells with suitable properties for use as a BBB model. Thus, isolated primary cells can also serve as the basis for a suitable immortalization technique to generate new cell lines. In this work, cerebral endothelial cells, pericytes, and astrocytes were successfully isolated and expanded using a mechanical/enzymatic method. Furthermore, in a triple coculture model, the cells showed a significant increase in barrier integrity compared with endothelial cell monoculture, as determined by transendothelial electrical resistance measurement and permeation studies using sodium fluorescein. The results demonstrate the opportunity to obtain all three cell types significantly involved in BBB formation from one species, thus providing a suitable tool for testing the permeation properties of new drug candidates. In addition, the protocols are a promising starting point to generate new cell lines of BBB-forming cells as a novel approach for BBB in vitro models.
血脑屏障（BBB）是新药进入大脑研发过程中的瓶颈。由于血脑屏障的存在，有毒物质无法进入大脑，但有潜力的候选药物透过血脑屏障的能力也很差。因此，合适的体外血脑屏障模型在临床前研发过程中尤为重要，因为它们不仅可以减少动物实验，还能使新药研发得更快。本研究的目的是从猪脑中分离脑内皮细胞、周细胞和星形胶质细胞，以构建血脑屏障的原代模型。此外，原代细胞因其特性非常适合，但分离过程复杂，必须确保永生化细胞具有更好的可重复性，因此对具有合适特性用作血脑屏障模型的永生化细胞有很高的需求。因此，分离出的原代细胞也可作为合适的永生化技术的基础，以产生新的细胞系。在这项工作中，使用机械/酶法成功分离并扩增了脑内皮细胞、周细胞和星形胶质细胞。此外，在三重共培养模型中，通过跨内皮电阻测量和使用荧光素钠的渗透研究确定，与内皮细胞单培养相比，这些细胞的屏障完整性显著提高。结果表明，有机会从一个物种中获得所有三种对血脑屏障形成有重要作用的细胞类型，从而为测试新药候选物的渗透特性提供了合适的工具。此外，这些方案是生成血脑屏障形成细胞新细胞系的一个有前景的起点，作为血脑屏障体外模型的一种新方法。
