Serum-Induced Inhibition of the Phagocytic Activity of Cultured Macrophages IC-21

Fetal bovine serum has been shown to considerably inhibit phagocytosis of non-opsonized 2-mu m fluorescent latex beads by cultured macrophages IC-21. Phagocytic activity was assessed using fluorescent microscopy and specially devised ImageJ plugins. Phagocytosis percent, PP (percentage of the bead-containing cells in the cell population under study), and phagocytosis index, PI (mean number of beads per cell in the beadcontaining population), were about 2 times lower in the cells incubated in the presence of 10% serum as compared to the respective parameters for the cells incubated in serum-free medium (55 - 5% vs. 92 +/- 1% and 2.0 +/- 0.2 vs. 4.3 +/- 0.2 beads/cell). The effect of serum was dose-dependent. Albumin (10 mg/ml) did not mimic the effect of serum, suggesting that fatty acid extraction was not the cause of the serum-induced inhibition. Serum is a source of exogenous cholesterol, therefore we checked if cholesterol removal could stimulate phagocytosis. Cholesterol-sequestering agent methyl-beta-cyclodextrin (m - CD) in concentrations of 5-7 mM indeed caused an increase of the phagocytic activity but at 10 mM exerted an inhibitory effect in serum-free medium. Connexin channel blocker carbenoxolone (CBX, 250-500 mu M) in most cases inhibited phagocytosis; the presence of serum or m beta CD modulated the CBX effects. The data indicate an important role of serum in regulation of the macrophage phagocytic activity. Stimulating effect produced by serum removal may partly be accounted for by a decrease of cholesterol concentration, which in turn may alter the functioning of integral proteins involved in the mechanisms of phagocytosis.