Rapid Internalization and Nuclear Translocation of CCL5 and CXCL4 in Endothelial Cells

Area of Science:

• Immunology
• Cell Biology

Background:

• Platelets release chemokines CCL5 and CXCL4, which bind to endothelial cells and promote monocyte adhesion.
• The subsequent fate and cellular mechanisms of chemokine deposition on endothelial cells remain incompletely understood.

Purpose of the Study:

• To investigate the internalization mechanisms and intracellular localization of CCL5 and CXCL4 after endothelial cell deposition.
• To determine the functional impact of chemokine internalization on monocyte arrest.

Main Methods:

• Incubation of human umbilical vein endothelial cells (HUVECs) and EA.hy926 cells with CCL5 or CXCL4.
• Analysis of chemokine uptake using microscopy and ELISA.
• Visualization of intracellular calcium signaling.
• Evaluation of monocyte arrest under laminar flow conditions.
• Confocal microscopy and ELISA on cell organelle fractions.

Main Results:

• CCL5 was fully internalized, while CXCL4 was partially retained on the cell surface.
• Endocytosis of both chemokines was rapid, active, and dependent on dynamin, clathrin, and G protein-coupled receptors (GPCRs).
• Internalization led to increased intracellular calcium signals and accumulation of both chemokines in the nucleus.
• Chemokine internalization did not alter monocyte adhesion to endothelial cells.

Conclusions:

• Endothelial cells actively internalize CCL5 and CXCL4 through clathrin- and dynamin-dependent endocytosis.
• Internalized chemokines are trafficked to the nucleus, suggesting novel intracellular functions.
• Despite internalization, the initial leukocyte arrest function of these chemokines is preserved.