Modeling hyperosmotic blood-brain barrier opening within human tissue-engineered in vitro brain microvessels.

As the majority of therapeutic agents do not cross the blood-brain barrier (BBB), transient BBB opening (BBBO) is one strategy to enable delivery into the brain for effective treatment of CNS disease. Intra-arterial infusion of the hyperosmotic agent mannitol reversibly opens the BBB; however, widespread clinical use has been limited due to the variability in outcomes. The current model for mannitol-induced BBBO assumes a transient but homogeneous increase in permeability; however, the details are poorly understood. To elucidate the mechanism of hyperosmotic opening at the cellular level, we developed a tissue-engineered microvessel model using stem cell-derived human brain microvascular endothelial cells (BMECs) perturbed with clinically relevant mannitol doses. This model recapitulates physiological shear stress, barrier function, microvessel geometry, and cell-matrix interactions. Using live-cell imaging, we show that mannitol results in dose-dependent and spatially heterogeneous increases in paracellular permeability through the formation of transient focal leaks. Additionally, we find that the degree of BBB opening and subsequent recovery is modulated by treatment with basic fibroblast growth factor. These results show that tissue-engineered BBB models can provide insight into the mechanisms of BBBO and hence improve the reproducibility of hyperosmotic therapies for treatment of CNS disease.

Human iPSC-derived blood-brain barrier microvessels: validation of barrier function and endothelial cell behavior.

Microvessels of the blood-brain barrier (BBB) regulate transport into the brain. The highly specialized brain microvascular endothelial cells, a major component of the BBB, express tight junctions and efflux transporters which regulate paracellular and transcellular permeability. However, most existing models of BBB microvessels fail to exhibit physiological barrier function. Here, using (iPSC)-derived human brain microvascular endothelial cells (dhBMECs) within templated type I collagen channels we mimic the cylindrical geometry, cell-extracellular matrix interactions, and shear flow typical of human brain post-capillary venules. We characterize the structure and barrier function in comparison to non-brain-specific microvessels, and show that dhBMEC microvessels recapitulate physiologically low solute permeability and quiescent endothelial cell behavior. Transcellular permeability is increased two-fold using a clinically relevant dose of a p-glycoprotein inhibitor tariquidar, while paracellular permeability is increased using a bolus dose of hyperosmolar agent mannitol. Lastly, we show that our human BBB microvessels are responsive to inflammatory cytokines via upregulation of surface adhesion molecules and increased leukocyte adhesion, but no changes in permeability. Human iPSC-derived blood-brain barrier microvessels support quantitative analysis of barrier function and endothelial cell dynamics in quiescence and in response to biologically- and clinically-relevant perturbations.