Fig. 4.

HDL can extract freFC directly from agLDL in absence of cells and during agLDL catabolism. (A–O) Alexa546–agLDL in a collagen matrix was left untreated for (A–C) 0 or (D–F) 4 h, or (G–O) treated with the given concentrations of HDL for 4 h prior to staining of FC or CE using filipin or LipidTOX Green, and analysis by confocal microscopy. Images were used to quantify (P) filipin, (Q) Alexa546–agLDL and (R) LipidTOX and Alexa546 intensity for at least 10 fields containing >500 agLDL aggregates relative to that in control cells (set at 1). (S) Alexa633–AgLDL was reconstituted with DHE or DHE-oleate and then incubated with or without 500 µg/ml HDL for 4 h. AgLDL was pelleted by centrifugation, resuspended and measured by fluorescence spectroscopy. (T) J774 macrophages were incubated for 4 h with DHE-labeled agLDL or DHE-labeled agLDL plus 500 μg/ml HDL or 40 mM cholesterol-balanced HPCD. The medium was collected, agLDL removed by centrifugation and DHE fluorescence measured by fluorescence spectroscopy. Data compiled from at least three independent experiments. Error bars show s.e.m. *P≤0.05; ***P≤0.001; n.s. not statistically significant.