3D neurovascular unit models

National Center For Advancing Translational Sciences

Investigators

Abstract

The neurovascular unit (NVU) acts as the major interface between the central nervous system (CNS) and the blood compartment and is involved in maintaining brain hemostasis by acting as a unique diusion barrier, restricting both immune migration and diusion of soluble molecules from blood to brain parenchyma. The NVU plays a key role in many acute and chronic neurological disorders, brain cancers, and encephalitic viral diseases such as neurovirulent virus infections. A 3D bioprinted NVU model is developed to study glioblastoma (GBM) tumor growth in a brain-like microenvironment. The NVU model includes human primary astrocytes, pericytes and brain microvascular endothelial cells, and patient-derived glioblastoma cells (JHH-520) are used for this study. Fluorescence reporters are used with confocal high content imaging to quantitate real-time microvascular network formation and tumor growth. Single cell RNA sequencing (scRNAseq) to establish physiologically relevant transcriptomics changes. The scRNAseq revealed changes in gene expression and cytokines secretion associated with wound healing/angiogenesis, including the appearance of an endothelial mesenchymal transition cell population. We have analyzed the scRNAseq data and identified several ligand-receptor interactions that we are now testing pharmacology as potential new targets for GBM. Flaviviruses can lead to severe neurotropic-encephalitic pathology, for which the viral nonstructural protein 1 (NS1) plays a major role. 2D in vitro cellular systems and animal models are currently used to investigate the effects of flavivirus NS1 in the NVU do not fully capture pathogenesis and drug responses seen in humans. Here is reported the development of a perfusable human brain microvascular network-on-a-chip (BMVasChip) model which incorporates brain endothelial cells and pericytes and recapitulates the cellular organization and expression of NVU-specific markers, including intracellular junction proteins. This BMVasChip model was used to investigate vasculopathy caused by exposure to a panel of flavivirus NS1 proteins, including Japanese encephalitis (JEV), West Nile (WNV), and dengue virus (DENV) NS1 proteins. The BMVasChip platform is able to demonstrate flavivirus NS1-induced pathophysiological phenotypes in the BMVasChip, including early and late endothelial activation and dysfunction features such as loss of barrier integrity, morphological features, and production of pro-inflammatory markers in a virus, dose, and time-dependent manner. In summary, the BMVasChip model recapitulates key features of endothelial dysfunction, potentially reflecting NS1’s role in viral pathogenesis and providing targets for antiviral therapeutics and vaccine development.