The blood-brain barrier (BBB) is a tight barrier formed by microvessels and capillaries that control the passageof nutrients, fluids, metabolic products, and drugs between the blood and the brain. Impaired function of theBBB is involved in a number of major pathologies afflicting the brain, such as Alzheimer?s disease, multiplesclerosis, Parkinson?s disease, brain manifestations of AIDS, stroke, and cancer. Although theneurotherapeutics sector is among the largest and fastest growing markets in the pharmaceutical industry,progress is currently impaired by the lack of in vitro assays that reliably predict in vivo BBB permeability. Noneof the existing models adequately replicates the organotypic microenvironment of the BBB, in which brainendothelial cells (ECs), pericytes (PCs) and astrocytes (ACs) are arranged in a characteristic architecture.The proposed work utilizes organ-on-chip technology recently developed by Nortis, Inc. for creating 3D tissuemicroenvironments in disposable microfluidic chips. The chip design enables the integration of living, lumenallyperfused microvasculature, making it suitable for studying barrier function. Strikingly, extensive preliminarydata indicate that human brain ECs, PCs, and ACs have the capacity to self-assemble into a BBB-likearchitecture within the Nortis chip. This data will be leveraged to further develop and eventually commercializeBBB models of mouse and human. The objective of Phase I is to achieve a model that replicates critical BBBfunctions of the mouse brain. The mouse model will be developed and optimized for viability, structure, andfunction. Expression of tight-junction (TJ) proteins and the transporter P-glycoprotein, an important functionalcharacteristic of the BBB, will be measured. Microvessel permeability will be assessed by perfusion withfluorescently labelled molecules (Aim 1). The model will then be challenged with the barrier-modulatingcompound lipopolysaccharide (LPS), and evaluated for associated changes in TJ protein expression, moleculepermeability, and leukocyte transendothelial migration (Aim 2). During Phase II, the mouse BBB chip will beused to develop and qualify specific BBB assays, such as transferrin receptor transporter activity, BBBpermeability challenge with LPS, and stimuli-induced leukocyte transmigration (Aim 1). Success criteria is anassay robustness of Z? ? 0.2.
Aim 2 of Phase II is to develop a human BBB model. The human model will beoptimized to recapitulate key structural and functional features of the BBB, including TJ formation, permeability,and transporter activity. To demonstrate utility, the model will be treated with LPS, mannitol, and angiotensin IIand evaluated for associated changes in BBB structure and function. Each of these compounds has clinicalrelevance but acts by a different mechanism.
Aim 3 is to qualify specific human BBB assays and establishrelevance to clinical data.The products developed with support from this grant will significantly enhance progress in basic, translational,and clinical neuroscience research and will significantly advance therapy for numerous devastating diseases.
