Laminin 221 fragment is suitable for the differentiation of human induced pluripotent stem cells into brain microvascular endothelial-like cells with robust barrier integrity

Aoki, Hiromasa; Yamashita, Misaki; Hashita, Tadahiro; Iwao, Takahiro; Matsunaga, Tamihide

doi:10.1186/s12987-020-00186-4

Cited by 21 publications

(24 citation statements)

References 22 publications

(27 reference statements)

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“…The present model presents improved tightness of the cell monolayer in comparison to cell line models such as hCMEC/D3, which displays low TEER values (~40 Ω·cm 2 , [ 3 ]) and for which an evaluation of the transcytosis of macromolecules across the model may only be carried using the so-called pulse-chase assay [ 105 ]. On the other hand, iPSC-derived BBB models have reported high TEER values (up to 2500 Ω·cm 2 , [ 106 ]), close to the ones reported in a physiological human BBB (~5000 Ω·cm 2 , [ 107 ]), however their differentiation conditions need to be carefully controlled to avoid an epithelial rather than endothelial phenotype [ 108 , 109 ]. In addition, no report can be found demonstrating transcytosis of anti-TfR antibodies in iPSC-derived BBB models, and comparable permeabilities for transferrin vs. human IgG have been shown [ 110 ].…”

Section: Discussionmentioning

confidence: 99%

Non-Human Primate Blood–Brain Barrier and In Vitro Brain Endothelium: From Transcriptome to the Establishment of a New Model

Chaves

Cegarra

et al. 2020

Pharmaceutics

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The non-human primate (NHP)-brain endothelium constitutes an essential alternative to human in the prediction of molecule trafficking across the blood–brain barrier (BBB). This study presents a comparison between the NHP transcriptome of freshly isolated brain microcapillaries and in vitro-selected brain endothelial cells (BECs), focusing on important BBB features, namely tight junctions, receptors mediating transcytosis (RMT), ABC and SLC transporters, given its relevance as an alternative model for the molecule trafficking prediction across the BBB and identification of new brain-specific transport mechanisms. In vitro BECs conserved most of the BBB key elements for barrier integrity and control of molecular trafficking. The function of RMT via the transferrin receptor (TFRC) was characterized in this NHP-BBB model, where both human transferrin and anti-hTFRC antibody showed increased apical-to-basolateral passage in comparison to control molecules. In parallel, eventual BBB-related regional differences were investigated in seven-day in vitro-selected BECs from five brain structures: brainstem, cerebellum, cortex, hippocampus, and striatum. Our analysis retrieved few differences in the brain endothelium across brain regions, suggesting a rather homogeneous BBB function across the brain parenchyma. The presently established NHP-derived BBB model closely mimics the physiological BBB, thus representing a ready-to-use tool for assessment of the penetration of biotherapeutics into the human CNS.

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Section: Discussionmentioning

confidence: 99%

Non-Human Primate Blood–Brain Barrier and In Vitro Brain Endothelium: From Transcriptome to the Establishment of a New Model

Chaves

Cegarra

et al. 2020

Pharmaceutics

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“…Recently, the use of laminin isoforms during the differentiation of iBMECs has been studied, suggesting laminin 221 (LN 221) as an enhancer of iBMEC barrier properties, but this study did not evaluate the effect of laminin isoforms of the endothelial BM on the culture of differentiated iBMECs [26]. Another study reported the effect of different ECM proteins including laminin on TEER of differentiated iBMECs; however, specific laminin isoforms of the endothelial BM were not investigated [27].…”

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confidence: 97%

Chemically defined human vascular laminins for biologically relevant culture of hiPSC-derived brain microvascular endothelial cells

Motallebnejad

Azarin

2020

Fluids Barriers CNS

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Background In recent years, differentiation of human induced pluripotent stem cells (hiPSCs) into brain-specific microvascular endothelial cells (iBMECs) has frequently been used to model the blood–brain barrier (BBB). However, there are limitations in the use of iBMECs for in vitro studies, such as transendothelial electrical resistance (TEER) instability, weak junctional expression of VE-cadherin, and lack of proper fluid shear stress response. In vivo, the basement membrane (BM) composition of the BBB evolves throughout development, and laminins become the dominant component of the mature vascular BM. However, laminin isoforms of the endothelial BM have not been used for culture of differentiated iBMECs. The main goal of this study is to investigate the effect of different laminin isoforms of the endothelial BM on iBMEC functionality and to determine whether better recapitulation of the physiological BM in vitro can address the aforementioned limitations of iBMECs. Methods Using a previously reported method, hiPSCs were differentiated into iBMECs. The influence of main laminins of the endothelial BM, LN 411 and LN 511, on iBMEC functionality was studied and compared to a collagen IV and fibronectin mixture (CN IV-FN). Quantitative RT-PCR, immunocytochemistry, and TEER measurement were utilized to assess gene and protein expression and barrier properties of iBMECs on different extracellular matrices. Single-channel microfluidic devices were used to study the effect of shear stress on iBMECs. Results LN 511, but not LN 411, improved iBMEC barrier properties and resulted in more sustained TEER stability. Immunocytochemistry showed improved junctional protein expression compared to iBMECs cultured on CN IV-FN. iBMECs cultured on LN 511 showed a reduction of stress fibers, indicating resting endothelial phenotype, whereas gene expression analysis revealed upregulation of multiple genes involved in endothelial activation in iBMECs on CN IV-FN. Finally, culturing iBMECs on LN 511 enhanced physiological responses to shear stress, including morphological changes and enhanced junctional protein association. Conclusion LN 511 improves the functionality and long-term barrier stability of iBMECs. Our findings suggest that incorporation of physiologically relevant LN 511 in iBMEC culture would be beneficial for disease modeling applications and BBB-on-a-chip platforms that accommodate fluid flow.

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“…The recognized lot-to-lot variability in Matrigel ® composition has led researchers to examine specific components of ECM to generate a more controlled and reproducible environment with less lot-to-lot variation. A recent study compared defined coatings to Matrigel ® for the differentiation of iPSC-derived BMECs [ 279 ] and found that cells cultured on laminin-221 had superior barriers, with higher TEER values, and the barrier was maintained for longer. Though laminin-221 is slightly different from the laminin-211 secreted by astrocytes, they both share the alpha-2 chain, which is suggested to be the main regulator of the maturation and function of the BBB [ 107 ].…”

Section: Decision Workflow: Factors To Consider When Selecting a Model Systemmentioning

confidence: 99%

Review of Design Considerations for Brain-on-a-Chip Models

et al. 2021

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In recent years, the need for sophisticated human in vitro models for integrative biology has motivated the development of organ-on-a-chip platforms. Organ-on-a-chip devices are engineered to mimic the mechanical, biochemical and physiological properties of human organs; however, there are many important considerations when selecting or designing an appropriate device for investigating a specific scientific question. Building microfluidic Brain-on-a-Chip (BoC) models from the ground-up will allow for research questions to be answered more thoroughly in the brain research field, but the design of these devices requires several choices to be made throughout the design development phase. These considerations include the cell types, extracellular matrix (ECM) material(s), and perfusion/flow considerations. Choices made early in the design cycle will dictate the limitations of the device and influence the end-point results such as the permeability of the endothelial cell monolayer, and the expression of cell type-specific markers. To better understand why the engineering aspects of a microfluidic BoC need to be influenced by the desired biological environment, recent progress in microfluidic BoC technology is compared. This review focuses on perfusable blood–brain barrier (BBB) and neurovascular unit (NVU) models with discussions about the chip architecture, the ECM used, and how they relate to the in vivo human brain. With increased knowledge on how to make informed choices when selecting or designing BoC models, the scientific community will benefit from shorter development phases and platforms curated for their application.

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Laminin 221 fragment is suitable for the differentiation of human induced pluripotent stem cells into brain microvascular endothelial-like cells with robust barrier integrity

Cited by 21 publications

References 22 publications

Non-Human Primate Blood–Brain Barrier and In Vitro Brain Endothelium: From Transcriptome to the Establishment of a New Model

Non-Human Primate Blood–Brain Barrier and In Vitro Brain Endothelium: From Transcriptome to the Establishment of a New Model

Chemically defined human vascular laminins for biologically relevant culture of hiPSC-derived brain microvascular endothelial cells

Review of Design Considerations for Brain-on-a-Chip Models

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