Hypoxia-enhanced Blood-Brain Barrier Chip recapitulates human barrier function and shuttling of drugs and antibodies

Tae-Eun Park,Henry Sanchez,Donald E Ingber,Eric Shusta,Hannah W Song,Liliana Christova Goumnerova,Nur Mustafaoglu,Ryan Hasselkus,Alexander Watters,Edward A Fitzgerald,Maximilian Benz,Robert Mannix,Sean P Palecek,Heather J Mccrea,Anna Herland,Rachelle Prantil-Baun,Olivier Henry

doi:10.1038/s41467-019-10588-0

Abstract

The high selectivity of the human blood-brain barrier (BBB) restricts delivery of many pharmaceuticals and therapeutic antibodies to the central nervous system. Here, we describe an in vitro microfluidic organ-on-a-chip BBB model lined by induced pluripotent stem cell-derived human brain microvascular endothelium interfaced with primary human brain astrocytes and pericytes that recapitulates the high level of barrier function of the in vivo human BBB for at least one week in culture. The endothelium expresses high levels of tight junction proteins and functional efflux pumps, and it displays selective transcytosis of peptides and antibodies previously observed in vivo. Increased barrier functionality was accomplished using a developmentally-inspired induction protocol that includes a period of differentiation under hypoxic conditions. This enhanced BBB Chip may therefore represent a new in vitro tool for development and validation of delivery systems that transport drugs and therapeutic antibodies across the human BBB.

Highlights

The high selectivity of the human blood-brain barrier (BBB) restricts delivery of many pharmaceuticals and therapeutic antibodies to the central nervous system
We found that exposure to hypoxia during this differentiation protocol produced significant (2–6-fold) increases in the mRNA levels for the endothelial cell–cell adhesion molecules, VEcadherin and PECAM-1, as well as the influx transporter GLUT-1 (BBB-specific glucose transporter), efflux transporter P-gp, and VEGF-A, relative to control induced pluripotent stem (iPS)-brain microvascular endothelial cells (BMVECs) induced under normoxic conditions (Supplementary Figure 1b and Supplementary Table 1)
BBB Chips formed with iPS-BMVECs induced by exposure to CoCl2, exhibited similar differential transcellular transport, again capturing the threefold difference between the two anti-TFR antibodies (Fig. 5b) without disrupting barrier integrity (Supplementary Figure 11c). These results demonstrate that mimicking the hypoxic microenvironment of the developing brain during differentiation of human iPSCs into BMVECs, by either lowering oxygen levels or adding hypoxia-induced factor 1α (HIF1α)-inducing mimetics (e.g., CoCl2 or DMOG), enables differentiation of endothelial cells that recapitulate human-relevant physiological BBB properties when the cells are interfaced with human brain astrocytes and pericytes in a 2-channel microfluidic BBB Chip

Summary

Introduction

The high selectivity of the human blood-brain barrier (BBB) restricts delivery of many pharmaceuticals and therapeutic antibodies to the central nervous system. We describe an in vitro microfluidic organ-on-a-chip BBB model lined by induced pluripotent stem cellderived human brain microvascular endothelium interfaced with primary human brain astrocytes and pericytes that recapitulates the high level of barrier function of the in vivo human BBB for at least one week in culture. Increased barrier functionality was accomplished using a developmentally-inspired induction protocol that includes a period of differentiation under hypoxic conditions This enhanced BBB Chip may represent a new in vitro tool for development and validation of delivery systems that transport drugs and therapeutic antibodies across the human BBB. The resulting human BBB Chip exhibits physiologically relevant levels of human BBB function for at least 1 week in vitro, including low barrier permeability and expression of multiple efflux pumps and transporter functions that are required for analysis of drug and therapeutic antibody transport

Methods

Results

Conclusion