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

Abstract

Optimizing drug candidates for blood-brain barrier (BBB) penetration in humans remains one of the key challenges and many devastating brain diseases including neurodegenerative diseases still do not have adequate treatments. So far, it has been difficult to establish state-of-the-art human stem cell derived in vitro models that mimic physiological barrier properties including a 3D microvasculature in a format that is scalable enough to screen drugs for BBB penetration in early drug development phases. 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 intact microvessels in 10 days of culturing. Produced microvessels show a typical BBB phenotype including 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. In summary, the here presented brain microvessel 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.

From bedside to the bench: patient-specific hiPSC-EC models uncover endothelial dysfunction in genetic cardiomyopathies

“…Then, functional assays have to be conducted to evaluate the angiogenic potential of hiPSC-ECs, such as their ability to form tube-like structures on Matrigel or hydrogel, and/ or to support vasculogenesis in vivo in xenograft models (DeCicco-Skinner et al, 2014;Orlova et al, 2014;Belair et al, 2016). Functional validation should also include assays for nitric oxide (NO) synthesis, low-density lipoprotein (LDL) incorporation, and for endothelial barrier function, such as transendothelial electrical resistance (TEER) or permeability assays using fluorescent tracers (Ritter et al, 2018;Goshi et al, 2019;Fengler et al, 2022). Moreover, hiPSC-ECs should respond to relevant stimuli, such as VEGF, by activating downstream signalling pathways involved in angiogenesis (Zachary, 2003).…”

Biosensors Integration in Blood–Brain Barrier-on-a-Chip: Emerging Platform for Monitoring Neurodegenerative Diseases

“…25,39 Also, previous work where TEER was monitored over the days of culture, showed significance difference in values due to the optimal incubation time for EC to develop TJs properly. 21,49 Moreover, several studies demonstrated that shear stress has a mechanotransductive effect that up-regulates the TJs expressions and RNA levels of BBB transporters. 50,51 As well, most BBB-oCs with applied shear stress showed higher electrical resistance (Figure 2E−H).…”

“…20,22,23 This fabrication approach is easier to industrialize than other configurations, which are currently commercialized by companies such as MIMETAS. 21 Usually, a positive electrode at the input and a negative electrode at the output are used for TEER measurements in this distribution (Figure 1C). The third arrangement in BBC-oC is based on tubular structures to mimic the cylindrical-like brain capillaries geometry based on the formation of hollow fibers as scaffolds that allows the tubular shape.…”

“…This system allows one to measure up to 40 samples at once in less than a minute, providing an efficient platform to evaluate the barrier permeability. 21…”

“Focused Ultrasound-mediated Drug Delivery in Humans – a Path Towards Translation in Neurodegenerative Diseases”

“…hiPSC-derived BBB models may become an important tool for simulation of potential adverse responses in human BBB to FUS+MB, and identify co-treatments or settings that could attenuate these effects. The high-throughput potential of novel human BBB models [83,102] and applicability of FUS+MB in vitro [84] could also facilitate screening of various combinations of disease-targeting drug and adjuvant therapies alleviating effects of FUS in the context of distinct neurodegenerative diseases or underlying mutations, before clinical application. For example, glucocorticoids (GC) have known immunosuppressive effect and were shown to reduce brain edema and improve tightness of the BBB by upregulating expression of occludin, claudins and VE-cadherin [103].…”