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UBC Theses and Dissertations

Podocalyxin promotes vascular barrier function Cait, Jessica

Abstract

The CD34-family sialomucin, podocalyxin (Podxl), is broadly expressed on the luminal face of blood vessels in adult mammals; however, its biological function on vascular endothelial cells (vEC) is not well-defined. Here, we reveal specific functions for podocalyxin in maintaining endothelial barriers using HUVEC monolayers as a model in vitro. Detailed analysis of barrier HUVEC characteristics using electrical cell-substrate impedance sensing (ECIS) and live cell imaging revealed essential roles for podocalyxin in maintaining cell-cell and cell-matrix interactions. Thus, podocalyxin-deficient HUVEC fail to form a functional barrier when plated on several extracellular matrix (ECM) substrates. Regardless of ECM substrate, these monolayers lack adherens junctions and focal adhesions; and display a disorganized cortical actin cytoskeleton. To explore an in vivo function of podocalyxin, we conditionally deleted Podxl in vEC using the Tie2Cre strain (PodxlΔTie2Cre). Although we did not detect altered permeability in naïve mice at steady state, systemic priming with lipopolysaccharides (LPS) disrupted the blood-brain barrier (BBB) in PodxlΔTie2Cre but not WT mice. To study the potential consequence of this BBB breach, we used a selective agonist of PAR-1, a thrombin receptor expressed by neurons and glial cells. As a polar peptide, the PAR-1 agonist (TFLLRN), is normally excluded from CNS parenchyma by the BBB. In response to systemic administration of TFLLRN, LPS-primed PodxlΔTie2Cre mice experienced a dramatic behavioral change marked by a severely dampened neurological electrical activity. We conclude that podocalyxin expression by CNS vECs is required to maintain BBB integrity under inflammatory conditions. Supplementary materials available at: http://hdl.handle.net/2429/68998

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Attribution-NonCommercial-NoDerivatives 4.0 International