Abstract
Neurovascular inflammation is a major contributor to many neurological disorders, but modeling these processes in vitro has proven to be difficult. Here, we microengineered a three-dimensional (3D) model of the human blood-brain barrier (BBB) within a microfluidic chip by creating a cylindrical collagen gel containing a central hollow lumen inside a microchannel, culturing primary human brain microvascular endothelial cells on the gel’s inner surface, and flowing medium through the lumen. Studies were carried out with the engineered microvessel containing endothelium in the presence or absence of either primary human brain pericytes beneath the endothelium or primary human brain astrocytes within the surrounding collagen gel to explore the ability of this simplified model to identify distinct contributions of these supporting cells to the neuroinflammatory response. This human 3D BBB-on-a-chip exhibited barrier permeability similar to that observed in other in vitro BBB models created with non-human cells, and when stimulated with the inflammatory trigger, tumor necrosis factor-alpha (TNF-α), different secretion profiles for granulocyte colony-stimulating factor (G-CSF) and interleukin-6 (IL-6) were observed depending on the presence of astrocytes or pericytes. Importantly, the levels of these responses detected in the 3D BBB chip were significantly greater than when the same cells were co-cultured in static Transwell plates. Thus, as G-CSF and IL-6 have been reported to play important roles in neuroprotection and neuroactivation in vivo, this 3D BBB chip potentially offers a new method to study human neurovascular function and inflammation in vitro, and to identify physiological contributions of individual cell types.
Highlights
The blood vessels in the brain are of major physiological importance because they maintain the blood-brain barrier (BBB), support molecular transport across this tight barrier, control local changes in oxygen and nutrients, and regulate the local immune response in the brain [1]
To build a 3D BBB chip containing a hollow endothelium lined microvessel surrounded by a compliant extracellular matrix (ECM), we first formed a cylindrical collagen gel within a single square-shaped microchannel (1 mm high × 1 mm wide × 2 cm long) (Fig 1A) in an optically clear polydimethysiloxane (PDMS) chip mounted on a standard glass microscope slide (Fig 1B) using soft lithography, as previously described [39]
This method for establishing co-cultures of multiple types of primary human brain-derived vascular cells in microfluidic chips that reconstitute their normal 3D spatial relationships has permitted us to dissect the contributions of these cells to the neuroinflammatory response in vitro
Summary
The blood vessels in the brain are of major physiological importance because they maintain the blood-brain barrier (BBB), support molecular transport across this tight barrier, control local changes in oxygen and nutrients, and regulate the local immune response in the brain [1]. The BBB is formed by the continuous brain microvascular endothelium, its underlying basement membrane, pericytes that tightly encircle the endothelium, and astrocytes in the surrounding tissue space that extend their cell processes towards the endothelium and insert on the basement membrane [5]. Together, these cells maintain a highly selective permeability barrier between the blood and the brain compartments that is critical for normal brain physiology. The complex interaction between these cell types and the microvascular endothelium make it extremely difficult to analyze their individual contribution to neuroinflammation in vivo
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