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| Figure 3: The effect of the eIF2a-S51A mutant on E‾/B48 lipoprotein catabolism. Raw 264.7 cells were stably transfected with a plasmid vector expressing eIF2a-S51A (S51A) or an empty pEGFP vector. (A) The transfected cells were incubated with various concentrations of 125I-labeled E+/B48 or E‾/B48 lipoproteins on ice for 4 hrs. The level of radioactivity bound to the cells was determined. (B, C, and D) Cells were incubated with 20 µg/ml of 125I-labeled E+/B48 or E‾/B48 lipoproteins at 37°C. Culture medium was collected at the indicated incubation times to determine the amount of degraded 125I-labeled lipoprotein particles. Cells were lysed to estimate the amount of cell-associated 125I-labeled lipoproteins. The uptake of lipoproteins was calculated as the sum of the associated and degraded lipoproteins at the given incubation time point. (E and F) Raw 264.7 cells were incubated at 37°C for 48 hrs with 20 µg/ml of E+/ B48 or E‾/B48 lipoproteins or culture medium alone. Cellular cholesterol contents were determined. Values represent the mean ± SEM of five experiments. *P<0.05 as compared with the control, and †P<0.05 as compared with cells transfected with empty pEGFP vector and treated with E‾/B48 lipoproteins. |
