Kinetics of the endocytotic pathway of Low Density Lipoprotein (LDL) in human endothelial cells line under shear stress: An in vitro confocal microscopy study

Abstract

We studied the effect of mechanical forces (shear stress) on the kinetics of internalization of native LDL and ox-LDL in endothelial cell line ECV304. This study was performed by using Confocal microscopy and FRET with two carbocyanine dyes, 1,1′-dioctadecyl-3,3,3′,3′-tetramethylindocarbocyanine perchlorate (DiO) as the donor and 3,3′-dioctadecyloxacarbocyanine perchlorate (DiI) as the receptor. The cells were incubated with a culture medium containing either 10 μg/ml DiI-LDL or DiO-LDL in static conditions or subjected to a laminar flow under a Confocal Laser Scanning Microscope (SP2 Leica, Germany). The results showed: (1) the possibility to evaluate the kinetics of LDL endocytosis in living cells, (2) shear stress in comparison with control group more effectively enhanced LDL uptake, (3) ox-LDL (>50 μg/ml) >4 hours incubation was found to affect the cells as reflected by their detachment at low shear stress.