Comparative Proteomic Analysis of Endocytic Cell-Surface Proteins in Vascular and Lymphatic Endothelial Cells

Vascular endothelial cells (VECs) and lymphatic endothelial cells (LECs) regulate the homeostasis of fluids, nutrients, and immune cells in the blood and lymphatic systems, respectively. These specialized functions depend on distinct cell-surface proteins and tightly regulated endocytic pathways, but the molecular determinants underlying the cell-type-specific endocytic profiles of these cells remain unclear. We sought to quantitatively characterize and compare the endocytic cell-surface proteomes of human umbilical vein endothelial cells (HUVECs) and human dermal LECs (HDLECs) using cell-surface biotinylation and internalization assays combined with sequential window acquisition of all theoretical fragment ion spectra-MS-based quantitative proteomics. HUVECs and HDLECs had a similar number of cell-surface and internalized proteins, with substantial overlap, but also included sets unique to each cell type. We identified 32 HUVEC- and HDLEC-enriched endocytic cell-surface proteins, respectively; these proteins were present in both enriched cell-surface and internalized fractions, representing cell-type-selective endocytic proteins. Functional enrichment analysis showed that HUVEC-enriched proteins were associated with angiogenesis, nutrient uptake, and metabolism, whereas HDLEC-enriched proteins were linked to immune regulation, extracellular matrix organization, and lymphangiogenesis. In conclusion, our present study demonstrates that VECs and LECs possess distinct endocytic cell-surface protein profiles that define their specialized functions and represent promising targets for endothelial-selective drug delivery and imaging.