High-density lipoprotein endocytosis in endothelial cells

Figure 3 Quantification of high density lipoprotein uptake. A: Time course of HDL uptake; HUVECs were incubated with iodinated HDL for up to 6 h and specific cell association was measured using a gamma counter. Measurements were normalized to protein content, determined by the Bradford protein assay; B: HUVECs were incubated with Alexa 568 labeled HDL for 1 h. Cells were fixed and immunofluorescene was performed using an antibody against LIMPII and visualized using fluorescence microscopy. No colocalization of LIMPII, a marker for the lysosomal compartment, and HDL-Alexa 568 was seen; C: HUVECs were incubated with HDL-Alexa-BP-C or -BP-CE for 60 min. Cells were then lyzed and the fluorescence intensity for each label was measured using a fluorometer. Results are expressed as the Alexa: Bodipy ratio initially and after the 60 min incubation period (n = 3). ^aP < 0.05 between groups. HDL: High density lipoprotein; BP-C: Bodipy-cholesterol; BP-CE: Bodipy-cholesteryl oleate; HUVECs: Human umbilical vein endothelial cells.