Figure 3.

Effect of IFG on enzyme uptake by macrophages. (A) Uptake of GLA with or without IFG-preincubation. Intracellular enzyme was assessed by enzyme activity (upper) and Western blot analysis (lower). Data are presented as mean ± SE (n = 3). *P < 0.005, **P<0.05, activity of the cells loaded with IFG-preincubated enzyme compared to that with untreated enzyme, t test. (B) Stability of endocytosed GLA with or without IFG-preincubation. Intracellular enzyme was assessed by enzyme activity (upper) and Western blot analysis (lower). Data are presented as mean ± SE (n = 3). (C) Comparison of activity and protein level of endocytosed GLA through 20 hrs after removal of enzyme from medium. Protein level was quantified by densitometry. Enzyme activity was obtained by subtracting the endogenous GLA activity in control cells from the activity in enzyme-loaded cells. (D) Effect of IFG on stability of the enzyme in the culture medium. GLA with or without IFG-preincubation was incubated in the medium at 37°C, and the enzyme amount was assessed by Western blotting. (E) Effect of IFG on binding and internalization of the enzyme to the cells. Binding was conducted at 4°C for 2 hrs. To assess internalization of surface-bound protein, the cells were transferred to 37°C and incubated for 45 min.