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  • The endosomal system of primary human vascular endothelial cells and albumin-FcRn trafficking.

The endosomal system of primary human vascular endothelial cells and albumin-FcRn trafficking.

Journal of cell science (2023-08-11)
Andreas Pannek, Janine Becker-Gotot, Steven K Dower, Anne M Verhagen, Paul A Gleeson
ABSTRACT

Human serum albumin (HSA) has a long circulatory half-life owing, in part, to interaction with the neonatal Fc receptor (FcRn or FCGRT) in acidic endosomes and recycling of internalised albumin. Vascular endothelial and innate immune cells are considered the most relevant cells for FcRn-mediated albumin homeostasis in vivo. However, little is known about endocytic trafficking of FcRn-albumin complexes in primary human endothelial cells. To investigate FcRn-albumin trafficking in physiologically relevant endothelial cells, we generated primary human vascular endothelial cell lines from blood endothelial precursors, known as blood outgrowth endothelial cells (BOECs). We mapped the endosomal system in BOECs and showed that BOECs efficiently internalise fluorescently labelled HSA predominantly by fluid-phase macropinocytosis. Pulse-chase studies revealed that intracellular HSA molecules co-localised with FcRn in acidic endosomal structures and that the wildtype HSA, but not the non-FcRn-binding HSAH464Q mutant, was excluded from late endosomes and/or lysosomes. Live imaging revealed that HSA is partitioned into FcRn-positive tubules derived from maturing macropinosomes, which are then transported towards the plasma membrane. These findings identify the FcRn-albumin trafficking pathway in primary vascular endothelial cells, relevant to albumin homeostasis.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Anti-FCGRT antibody produced in rabbit, Prestige Antibodies® Powered by Atlas Antibodies, affinity isolated antibody, buffered aqueous glycerol solution
Sigma-Aldrich
PMA, for use in molecular biology applications, ≥99% (HPLC)
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5-(N-Ethyl-N-isopropyl)amiloride