Fabrication and Optimization of a Tubular Blood Brain Barrier Model from Polycaprolactone Electrospun Fibers

Abstract

The blood-brain barrier (BBB) regulates the exchange of nutrients and molecules between the brain and the circulating blood. The control over the passage of molecules into and out of the brain is achieved through a layer of endothelial cells interconnected through tight junctions that protect the brain by allowing only certain substances to cross into the brain milieu. However, the BBB also impedes drug delivery to the brain making it harder to treat common brain diseases including but not limited to Alzheimer’s and Parkinson’s. In order to facilitate research in brain-related diseases, in vitro models that mimic the BBB are being developed, but the challenge is to create a barrier integrity similar to that of in vivo tissues. Previous studies have been conducted to create two-dimensional (2D) and ductular models that mimic the structure of the BBB, enhance cell attachment, and support the formation of tight junctions. However, these models have a number of biomimicry shortcomings such as the absence of dynamic blood flow conditions on cells, the lack of interaction between the duct lumen and the external environment and having rectangular ducts unlike the circular in vivo geometry all of which can alter the properties of the BBB. To address the above limitations, we engineered a BBB model by electrospinning a thin polycaprolactone (PCL) fibrous and porous circular duct that favors cell attachment in monolayers and enables exchange of media and nutrients between the inside and the outside of the duct. The biological and mechanical properties of the ducts were optimized for cell culture, specifically endothelial cells (ECV-304), to study the cells’ ability to cover the ducts and form tight junctions. It was found that 25 w/v% PCL solution, with 1 hr duration of electrospinning, and 20 x 10^6 cell/mL seeding density resulted in the highest cell coverage of 45.3% on day 7 compared to 5 x 10^6 cell/mL (p=0.0014) and reaching 85.5% cell coverage after 14 days. Fiber alignment was achieved longitudinally on thin wires after decreasing the flow rate to 0.25 mL/hr. These optimized conditions were used to evaluate the ability of the model to mimic the BBB functionality by measuring the trans-endothelial electric resistance (TEER) over 14 days in comparison to the Transwell® model. TEER significantly increased after 14 days from 23 Ω.cm^2 to 186 Ω.cm^2 compared to the Transwell® alternative that only reached 67 Ω.cm^2 after 14 days (p=0.0105). The developed model holds a promising potential for mimicking in vivo ductular tissues such as the BBB, breast ducts, and tissues or organs with a cylindrical architecture. Furthermore, it allows the incorporation of mimetic blood flow over cells to better mimic the in vivo conditions and increase extracellular matrix deposition by cells thus resulting in a more biomimetic barrier.