Hydrodynamic Focusing-Enabled Blood Vessel Fabrication for in Vitro Modeling of Neural Surrogates

The ability to interrogate the brain’s vasculature with improved resolution and sensitivity would be a major advancement for tissue engineering. Most in vitro brain models rely on 2-dimensional cell cultures or brain organoid cultures, yet neither system can assess the influence played by fluid perfusion dynamics in brain vascular development and maintenance as both models are avascular. Hydrodynamic focusing, the use of fluid dynamics to controllably mix biological materials including cells, has emerged as a powerful technique with which to organize living cells into complex, ordered 3-dimensional architectures. In the present work, we employ hydrodynamic focusing to fabricate blood vessels comprised of primary human endothelial cells and normal human primary astrocytes. Following maturation, the microvessels exhibit a fully developed endothelial cell-lined lumen, astrocytic outgrowth, and improved tight junction protein expression. Barrier development and gene analyses in response to perfusion are also described. These results demonstrate this fabrication technique to be an enabling technology for the interrogation of blood vessel dynamics in 3-dimensional in vitro neural surrogates that mimic the blood–brain barrier.