Tissue-engineered blood-brain barrier models via directed differentiation of human induced pluripotent stem cells

Grifno, Gabrielle N.; Farrell, Alanna; Linville, Raleigh M.; Arevalo, Diego; Kim, Joo Ho; Gu, Luo; Searson, Peter C.

doi:10.1038/s41598-019-50193-1

Cited by 76 publications

(75 citation statements)

References 66 publications

(98 reference statements)

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“…Importantly, the protocols mentioned above continue to benchmark differentiated cells using TEER, efflux transporter activity (ETA), and expression of essential BBB junctional proteins (Claudin-5, ZO-1, Occludin, VE-Cadherin), transporters (P-gp, GLUT1), and other factors (PECAM-1, VEGFR2, vWF). Additional recent improvements in the differentiation of iBMECs include the use of fully defined media [20], sorting strategies to increase iBMEC purity [21], and effective methods for the cryopreservation of differentiated cells [22,23], which collectively are improving the reproducibility and scalability of iBMECs for laboratory and potential clinical use.…”

Section: Improvements In Bmec Differentiation Methodsmentioning

confidence: 99%

“…Additionally, as new methods to generate iBMECs continue to evolve, the reproducibility of differentiation protocols needs to be considered. iBMEC differentiation efficiency and barrier formation have been shown to vary based on cell line [12], cell seeding density [16], reagent source [84], and response to media components [23,62]. Developing robust protocols that are less sensitive to these variables will undoubtedly improve intra-and interlab reproducibility.…”

Section: Challenges and Future Directionsmentioning

confidence: 99%

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Recent advances in human iPSC-derived models of the blood–brain barrier

Workman

Svendsen

2020

Fluids Barriers CNS

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The blood-brain barrier (BBB) is a critical component of the central nervous system that protects neurons and other cells of the brain parenchyma from potentially harmful substances found in peripheral circulation. Gaining a thorough understanding of the development and function of the human BBB has been hindered by a lack of relevant models given significant species differences and limited access to in vivo tissue. However, advances in induced pluripotent stem cell (iPSC) and organ-chip technologies now allow us to improve our knowledge of the human BBB in both health and disease. This review focuses on the recent progress in modeling the BBB in vitro using human iPSCs.

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Section: Improvements In Bmec Differentiation Methodsmentioning

confidence: 99%

Section: Challenges and Future Directionsmentioning

confidence: 99%

Recent advances in human iPSC-derived models of the blood–brain barrier

Workman

Svendsen

2020

Fluids Barriers CNS

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“…The human immortalized endothelial cell line hCMEC/D3 [153,154] is widely used for the study of drug transport [163][164][165][166] because it has been thoroughly characterized and it expresses ABC and SLC transporters, as well as tight junctions [167][168][169], despite the fact that the tightness of its monolayer is lower than in intact microvessels due to a lower expression of claudin-5 [169]. Recently, human brain endothelial cells have been obtained from stem cells such as human cord blood-derived stem cells of circulating endothelial progenitor and hematopoietic lineages [156,157], human pluripotent stem cells (hPSCs) [158], and induced pluripotent stem cells (iPSCs) [170,171]. After differentiation and isolation, the hPSC-derived brain endothelial cells monolayers present key BBB characteristics, including tight junctions and functional ABC transporters [158,159,172].…”

Section: In Vitro Models Of the Bbbmentioning

confidence: 99%

ABC Transporters at the Blood–Brain Interfaces, Their Study Models, and Drug Delivery Implications in Gliomas

et al. 2019

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Drug delivery into the brain is regulated by the blood–brain interfaces. The blood–brain barrier (BBB), the blood–cerebrospinal fluid barrier (BCSFB), and the blood–arachnoid barrier (BAB) regulate the exchange of substances between the blood and brain parenchyma. These selective barriers present a high impermeability to most substances, with the selective transport of nutrients and transporters preventing the entry and accumulation of possibly toxic molecules, comprising many therapeutic drugs. Transporters of the ATP-binding cassette (ABC) superfamily have an important role in drug delivery, because they extrude a broad molecular diversity of xenobiotics, including several anticancer drugs, preventing their entry into the brain. Gliomas are the most common primary tumors diagnosed in adults, which are often characterized by a poor prognosis, notably in the case of high-grade gliomas. Therapeutic treatments frequently fail due to the difficulty of delivering drugs through the brain barriers, adding to diverse mechanisms developed by the cancer, including the overexpression or expression de novo of ABC transporters in tumoral cells and/or in the endothelial cells forming the blood–brain tumor barrier (BBTB). Many models have been developed to study the phenotype, molecular characteristics, and function of the blood–brain interfaces as well as to evaluate drug permeability into the brain. These include in vitro, in vivo, and in silico models, which together can help us to better understand their implication in drug resistance and to develop new therapeutics or delivery strategies to improve the treatment of pathologies of the central nervous system (CNS). In this review, we present the principal characteristics of the blood–brain interfaces; then, we focus on the ABC transporters present on them and their implication in drug delivery; next, we present some of the most important models used for the study of drug transport; finally, we summarize the implication of ABC transporters in glioma and the BBTB in drug resistance and the strategies to improve the delivery of CNS anticancer drugs.

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“…For example, 3D BBB models commonly utilize nonbrain ECM components including collagen I [27,[54][55][56][57][58] and fibrin [20][21][22]59]. We previously characterized and compared the stiffness of collagen I hydrogels to native mouse brain, and showed that 6 mg mL − 1 collagen is a reasonable proxy for brain stiffness [7]. Additionally, materials with stiffnesses much lower than native brain were not conducive to the formation of stable BBB microvessels [7].…”

Section: Critical Chemical Cues Implicated In Developmental Brain Angmentioning

confidence: 99%

“…To enable imaging of angiogenesis in 3D, we have adapted the fibrin bead angiogenesis assay [6], forming a confluent monolayer of iPSC-derived brain microvascular endothelial cells (dhBMECs) on microbeads (BBB beads). The beads are then embedded in collagen I hydrogels, which mimic the native stiffness of brain parenchyma [7]. We then explore how changes in the chemical and extracellular matrix microenvironment influence in vitro brain angiogenesis.…”

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confidence: 99%

Three-dimensional induced pluripotent stem-cell models of human brain angiogenesis

Searson

Linville

Arevalo

et al. 2019

Preprint

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Background: During brain development, chemical cues released by developing neurons, cellular signaling with pericytes, and mechanical cues within the brain extracellular matrix (ECM) promotes angiogenesis occurs of brain microvascular endothelial cells (BMECs). During brain disease, angiogenesis can also occur due to pathological chemical, cellular, and mechanical signaling. Existing in vitro and in vivo models of brain angiogenesis have key limitations. Methods: Here, we develop a high-throughput in vitro BBB bead assay of brain angiogenesis utilizing 150 μm diameter beads coated with induced pluripotent stem-cell (iPSC)-derived human BMECs (dhBMECs). After embedding the beads within a 3D matrix, we introduce various chemical cues and extracellular matrix components to explore their effects on angiogenic behavior. Based on the results from the bead assay, we generate a multi-scale model of the human cerebrovasculature within perfusable three-dimensional tissue-engineered blood-brain barrier (BBB) microvessels. Results: A sprouting phenotype is optimized in confluent monolayers of dhBMECs using chemical treatment with vascular endothelial growth factor (VEGF) and wnt ligands, and the inclusion of pro-angiogenic ECM components. As a proof-of-principle that the bead angiogenesis assay can be applied to study pathological angiogenesis, we show that oxidative stress can exert concentration-dependent effects on angiogenesis. Finally, we demonstrate the formation of a hierarchical microvascular model of the human blood-brain barrier displaying key structural hallmarks. Conclusions: We develop two in vitro models of brain angiogenesis: the BBB bead assay and the tissue-engineered BBB microvessel model. These platforms provide a tool kit for studies of physiological and pathological brain angiogenesis, with key advantages over existing two-dimensional models.

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Tissue-engineered blood-brain barrier models via directed differentiation of human induced pluripotent stem cells

Cited by 76 publications

References 66 publications

Recent advances in human iPSC-derived models of the blood–brain barrier

Recent advances in human iPSC-derived models of the blood–brain barrier

ABC Transporters at the Blood–Brain Interfaces, Their Study Models, and Drug Delivery Implications in Gliomas

Three-dimensional induced pluripotent stem-cell models of human brain angiogenesis

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