Figure 1.

Altered recycling of BODIPY-LacCer in NPFs vs. normal HSFs. (A) Cells were incubated with 2.5 μM BODIPY-LacCer for 1 h at 16°C to selectively label EEs, washed, and back-exchanged with 5% DF-BSA to remove any fluorescent lipid remaining at the PM. The samples were then incubated for 0, 10, or 40 min at 37°C in the presence of 5% DF-BSA to remove any fluorescent LacCer that was recycled back to the PM. G, Golgi apparatus. Bar, 10 μm. (B) Cells were incubated with fluorescent LacCer as in A, and the amount of cell-associated fluorescence was determined by lipid extraction and analysis. Inset, the first 10 min of LacCer recycling was examined in detail. In some experiments (filled symbols), cells were grown in lipoprotein-deficient serum to deplete cholesterol (see Materials and Methods). All values are expressed as percentage of cell-associated fluorescence at 0 min for each cell type and are mean ± SD of three or more independent experiments.