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人诱导多能干细胞衍生的脑微血管在多孔板中可用于抗炎药物的通透性筛选。

Human iPSC-derived brain endothelial microvessels in a multi-well format enable permeability screens of anti-inflammatory drugs.

机构信息

Laboratory Automation Technologies (CRFS-LAT), German Center for Neurodegenerative Diseases (DZNE), 53127, Bonn, Germany.

Light Microscopy Facility (CRFS-LMF), German Center for Neurodegenerative Diseases (DZNE), 53127, Bonn, Germany.

出版信息

Biomaterials. 2022 Jul;286:121525. doi: 10.1016/j.biomaterials.2022.121525. Epub 2022 Apr 30.

Abstract

Optimizing drug candidates for blood-brain barrier (BBB) penetration remains one of the key challenges in drug discovery to finally target brain disorders including neurodegenerative diseases which do not have adequate treatments so far. It has been difficult to establish state-of-the-art stem cell derived in vitro models that mimic physiological barrier properties including a 3D microvasculature in a format that is scalable to screen drugs for BBB penetration. To address this challenge, we established human induced pluripotent stem cell (iPSC)-derived brain endothelial microvessels in a standardized and scalable multi-well plate format. iPSC-derived brain microvascular endothelial cells (BMECs) were supplemented with primary cell conditioned media and grew to microvessels in 10 days. Produced microvessels show typical BBB endothelial protein expression, tight-junctions and polarized localization of efflux transporter. Microvessels exhibited physiological relevant trans-endothelial electrical resistance (TEER), were leak-tight for 10 kDa dextran-Alexa 647 and strongly limited the permeability of sodium fluorescein (NaF). Permeability tests with reference compounds confirmed the suitability of our model as platform to identify potential BBB penetrating anti-inflammatory drugs. The here presented platform recapitulates physiological properties and allows rapid screening of BBB permeable anti-inflammatory compounds that has been suggested as promising substances to cure so far untreatable neurodegenerative diseases.

摘要

优化候选药物透过血脑屏障(BBB)的能力仍然是药物发现中的关键挑战之一,旨在最终针对包括神经退行性疾病在内的脑部疾病,这些疾病迄今尚无有效的治疗方法。建立能够模拟生理屏障特性的最先进的基于干细胞的体外模型一直具有挑战性,包括以可扩展的方式筛选穿透 BBB 的药物的 3D 微血管结构。为了解决这一挑战,我们建立了人诱导多能干细胞(iPSC)衍生的脑内皮微血管,采用标准化和可扩展的多孔板格式。添加原代细胞条件培养基后,iPSC 衍生的脑微血管内皮细胞(BMEC)生长为微脉管,10 天即可形成。生成的微血管显示出典型的 BBB 内皮蛋白表达、紧密连接和外排转运蛋白的极化定位。微血管表现出生理相关的跨内皮电阻(TEER),对 10 kDa 的葡聚糖-Alexa 647 具有不透性,并且强烈限制了荧光素钠(NaF)的通透性。用参考化合物进行的渗透测试证实了我们模型的适用性,可作为识别潜在穿透 BBB 的抗炎药物的平台。该平台再现了生理特性,并允许快速筛选穿透 BBB 的抗炎化合物,这些化合物被认为是有希望治愈迄今无法治疗的神经退行性疾病的物质。

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