Discipline: Technology and Engineering
Subcategory: Biomedical Engineering
Bethsymarie Soto Morales - University of Puerto Rico–Mayaguez
Co-Author(s): Scott Canfield, Eric Shusta, and Sean Palecek, University of Wisconsin-Madison, WI
The blood brain barrier (BBB) restricts the delivery of toxins, bacteria, viruses and ions to the brain. Additionally, the BBB is affected in a number of pathological states. The BBB is comprised of brain microvascular endothelial cells (BMECs) that line the brain capillaries and are supported by astrocytes, pericytes, neurons, and neural stem cells. There are several limitations to current BBB models including: species variation, loss of BBB properties over time, and sub-par BBB phenotypes. We utilized stem cells to derive the neurovascular unit comprised of BMECs, neurons and astrocytes in an attempt to have the most physiologically relevant BBB model currently available. These iPSC-derived BMECs exhibit greater BBB properties than other models currently available, such as a greater transendothelial electrical resistance (TEER), reduced permeability, polarized efflux transport, and expression of tight junctional markers. However, to further enhance our BMEC model and recapitulate the in vivo setting we co-cultured our BMECs with human neurons and astrocytes from stem cells. To monitor BBB properties we utilized several techniques. TEER measurements were conducted to measure the tightness of the barrier as recorded as an increase in electrical resistance. Fluorescein permeability was monitored to additionally detect barrier tightening. The transport of Rhodamine 1,2,3 was monitored with and without the presence of a PGP efflux transporter inhibitor to directly measure PGP efflux transporter activity. Finally, the presences of tight junctional proteins were observed with immunocytochemistry. All changes in barrier properties due to co-culture were directly compared to monoculture (BMECs only). The co-culture of BMECs with both stem cell-derived astrocytes and neurons further enhanced our BBB phenotype, specifically a substantial elevation in the transendothelial electrical resistance, reduction in fluorescein permeability, and an increase in tight junctional proteins visualized by immunocytochemistry. Future research involves quantifying tight junctional protein changes, verifying observations in multiple cell lines, and the investigation of signaling pathways involved in the co-culture induced barrier tightening. The development of an entirely human stem cell derived BBB neurovascular unit will be critical in understanding the physiological and pathological states of the BBB.
Funder Acknowledgement(s): This study was supported by a generous gift from The University of Wisconsin-Madison Graduate School and the University of Wisconsin-Madison College of Engineering. Finally I thank Kelly Burton for this opportunity and the Sean and Eric Laboratory Group for help in the laboratory work.
Faculty Advisor: Sean Palecek, Eric Shusta, Scott Canfield, firstname.lastname@example.org