Molecular and cellular mechanisms underlying adult blood-brain barrier maintenance

Abstract

To ensure optimal neuronal function, the blood-brain barrier (BBB) maintains a homeostatic brain microenvironment free from harmful agents and delivers nutrients to the brain to support metabolic needs. Although the BBB was first identified more than a century ago, the molecular and cellular mechanisms underlying its formation and maintenance have only recently begun to be uncovered. To date, studies that have examined BBB function in adult mice have largely focused on injury, aging, pathological, or therapeutic models. Thus, the molecular and cellular mechanisms underlying physiological BBB maintenance in adulthood remain largely unknown.

The goal of this dissertation is to uncover novel mechanisms of adult BBB maintenance. First, we provide a mechanism of how canonical Wnt signaling regulates BBB integrity in adulthood. Following acute attenuation of canonical Wnt signaling in mouse brain endothelial cells (ECs) in vivo, we find that there is an increase in transcytosis, which correlates with a rapid loss of the caveolae-mediated transcytosis inhibitor MFSD2A. Strikingly, EC-specific overexpression of MFSD2A in Wnt signaling-deficient brain ECs is not sufficient to rescue increased transcytosis, indicating that canonical Wnt signaling regulates multiple transcytosis pathways to maintain BBB integrity. Second, we make progress toward the development of small molecular weight tracers to evaluate the permeability of tight junctions (TJs) at the ultrastructural level. In Wnt signaling-deficient brain ECs, we find that TJs are impermeable to the smallest molecular weight tracer assayed (1.9 kDa) despite reductions in TJ protein levels.

Third, we investigate the role of nutrient transporters in establishing and maintaining BBB integrity. This class of genes, which is highly expressed and enriched in brain ECs compared to peripheral ECs, delivers a wide variety of nutrients and substrates to the brain to meet the metabolic and energetic needs of neurons. We use a combination of mouse genetics, BBB leakage assays, and electron microscopy to investigate the roles of Slc16a1 (monocarboxylate transporter 1, MCT1) and Slc7a5 (large neutral amino acid transporter 1, LAT1) in regulating BBB integrity, finding that neither are required for BBB integrity.