Introduction:: The blood-brain barrier (BBB) is a highly selective structure and provides a physical barrier between the central nervous system (CNS) and the peripheral circulation, preventing harmful substances like pathogens, toxins, and even many medications from passing through. BBB disruption has been associated with various diseases such as stroke, Alzheimer's disease, and multiple sclerosis. In vitro platforms involving human cells and recapitulating some of the cardinal features of BBB will significantly contribute to new understandings and development of new therapeutic interventions. We have leveraged microfluidics to create a perfusable BBB. Our model consists of a microfluidic device with two compartments separated by a porous membrane. The model allows for the co-culture of endothelial cells along with other cell types, such as astrocytes and pericytes, the key cells involved in BBB function. Employing this platform, we have examined the effect of inflammatory molecules on BBB integrity and function by using IL-1b as a model system. We also investigated the protective effects of omega-3 fatty acids on the BBB. Omega-3 fatty acids are important polyunsaturated fatty acids with anti-inflammatory, antioxidant, and neuroprotective effects.
Materials and Methods:: The device consists of two compartments (vascular channels and brain chamber) separated by a thin, porous polyester track etch (PETE) membrane (pore size: 0.4 μm). The PETE membrane was coated with Matrigel and seeded serially with human brain microvascular endothelial cells (HBMVECs) in the vascular channel, and primary human astrocytes and brain vascular pericytes in the brain chamber. A peristaltic pump was used to perfuse the device with medium (40 μL/min). Co-culture was maintained for 4 days before exposing to IL-1b and/or Omegaven® through the vasculature channel. Changes to the BBB integrity was examined by TEER measurement, tracer diffusion, and immunofluorescent staining.
Results, Conclusions, and Discussions:: TEER measurements showered lower TEER value following IL-1β treatment compared to control and Omegaven® treated groups. However, the endothelial cell layer pretreated with Omegaven® prior to IL-1β exposure, the TEER values were recovered to the levels similar to the control and Omegaven® groups. Immunofluorescent staining was carried out for the tight junction protein Zonula occludens-1 (ZO-1) and vascular cell adhesion molecule 1 (VCAM-1). The results suggested impaired BBB integrity resulting from the IL-1β exposure, which was attenuated by the Omega-3 treatment. Treatment specific changes to the BBB integrity was also confirmed assessing the diffusion of FITC-labeled dextran (70kDa) across the BBB. In addition, changes to astrocyte morphology and network density were observed with IL-1β treatment. In contrast, pretreatment with Omega-3 fatty acids rescued the astrocyte morphology and the disruption in network density caused by IL-1β. This model is useful for understanding the changes to BBB structure and function, as well as screening potential therapeutic drugs. Our findings highlight the potential use of in vitro models to understand the complex interactions between BBB components. Future research will expand the model to include additional brain cells and validate its applicability to study neuroinflammation.
Acknowledgements (Optional): : This research was supported by the National Institutes of Health (NIH) (R01AR79189-02S1 PA034673, RF1AG076203-01) and The Alfred P. Sloan Foundation. We gratefully acknowledge their support in funding this project.
