An isogenic neurovascular unit model comprised of human induced pluripotent stem cell-derived brain microvascular endothelial cells, pericytes, astrocytes, and neurons

Scott G. Canfield,Eric V. Shusta,Sean P. Palecek,Matthew J. Stebbins,Madeline G. Faubion,Benjamin D. Gastfriend

doi:10.1186/s12987-019-0145-6

Scott G. Canfield, Eric V. Shusta + Show 4 more

Open Access

https://doi.org/10.1186/s12987-019-0145-6

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Abstract

BackgroundBrain microvascular endothelial cells (BMECs) astrocytes, neurons, and pericytes form the neurovascular unit (NVU). Interactions with NVU cells endow BMECs with extremely tight barriers via the expression of tight junction proteins, a host of active efflux and nutrient transporters, and reduced transcellular transport. To recreate the BMEC-enhancing functions of NVU cells, we combined BMECs, astrocytes, neurons, and brain pericyte-like cells.MethodsBMECs, neurons, astrocytes, and brain like pericytes were differentiated from human induced pluripotent stem cells (iPSCs) and placed in a Transwell-type NVU model. BMECs were placed in co-culture with neurons, astrocytes, and/or pericytes alone or in varying combinations and critical barrier properties were monitored.ResultsCo-culture with pericytes followed by a mixture of neurons and astrocytes (1:3) induced the greatest barrier tightening in BMECs, supported by a significant increase in junctional localization of occludin. BMECs also expressed active P-glycoprotein (PGP) efflux transporters under baseline BMEC monoculture conditions and continued to express baseline active PGP efflux transporters regardless of co-culture conditions. Finally, brain-like pericyte co-culture significantly reduced the rate of non-specific transcytosis across BMECs.ConclusionsImportantly, each cell type in the NVU model was differentiated from the same donor iPSC source, yielding an isogenic model that could prove enabling for enhanced personalized modeling of the NVU in human health and disease.

Highlights

The blood–brain barrier (BBB) is both a passive and active barrier between the CNS and its surrounding vasculature [1]
Following seeding onto Transwells, Brain microvascular endothelial cells (BMECs) were co-cultured with either iPSCderived pericytes alone (P), a neuron and astrocyte mixture (1:3 NA) or a sequential pericyte co-culture followed by a neuron and astrocyte co-culture (PNA) (Fig. 1a)
Monocultured BMECs exhibited a maximum Trans‐endothelial electrical resistance (TEER) value of 310 ± 19 Ω × cm2, whereas BMECs co-cultured with pericytes reached a maximum TEER value of 564 ± 21 Ω × cm2 (p < 0.05) and remained significantly elevated above monoculture and fibroblast co-culture for the duration of the experiment (p < 0.05)

Summary

Introduction

The blood–brain barrier (BBB) is both a passive and active barrier between the CNS and its surrounding vasculature [1]. A number of in vitro models have been utilized to understand the role of the BBB in both physiological and pathological states [9,10,11,12]. Human induced pluripotent stem cell (iPSC)-derived BMECs have recently become available and exhibit multiple critical phenotypes: elevated TEER, reduced permeability, and the expression of tight junction proteins, nutrient transporters and efflux transporters [21,22,23,24,25,26]. Brain microvascular endothelial cells (BMECs) astrocytes, neurons, and pericytes form the neurovascular unit (NVU). Interactions with NVU cells endow BMECs with extremely tight barriers via the expression of tight junction proteins, a host of active efflux and nutrient transporters, and reduced transcellular transport. To recreate the BMEC-enhancing functions of NVU cells, we combined BMECs, astrocytes, neurons, and brain pericyte-like cells

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Discussion

Conclusion