Fig. 5.

A pulse-chase quantification of uptake, resecretion (intact) and degradation of endogenously labeled 35S-apoA-I from macrophages. A: RAW cells were incubated (pulse) with 3 μg/ml ³⁵S-apoA-I ± 0.3 mM cAMP for 1 h. They were washed thoroughly according to Methods to remove residual surface ³⁵S-apoA-I and then incubated with clean medium containing 50 μg/ml human HDL for 1.5 h (chase) to assess resecretion. Total uptake is defined as the sum of the residual cell label and the label in the chase medium at the end of the experiment. The amount of secreted, degraded label was determined from a TCA precipitation performed on the chase medium. Resecreted (intact) apoA-I was defined as the difference between the total label in the chase medium and the amount that was degraded. Mass values were determined from the specific activity of the initial ³⁵S-apoA-I. B: The pulse-chase experiment was performed exactly as stated above, except that the length of the chase incubation was varied as shown and cAMP was included in all samples. A one-way ANOVA failed to identify a significant difference among the ratios at any time point. The error bars represent 1 SD of triplicate samples from one experiment.