Effects of recombinant soluble CD14 on association of bacterial lipopolysaccharide with bovine alveolar macrophages in vitro

Abstract

The CD14 receptor on the cell membrane of macrophages is well-known as a receptor for bacterial lipopolysaccharide (LPS) and contributes to cellular activation, but the potential role of the soluble CD14 receptor (sCD14) in macrophage activation is less understood. In this study, CD14-dependent binding or uptake of [3H]-LPS by bovine alveolar macrophages (AM) was determined in vitro, and LPS-mediated activation of AM was evaluated using the tissue factor-procoagulant assay. In the absence of LPS-binding protein (LBP) or other serum components, recombinant human sCD14 (rsCD14) enhanced binding of [3H]-LPS to AM and also increased LPS-mediated activation of AM in a concentration-dependent manner. These effects were inhibitable by anti-CD14 monoclonal antibodies; the antibodies decreased, but did not completely prevent, association of [3H]-LPS with cells. Binding of [3H]-LPS to AM was greater in the presence of rsCD14 + bovine LBP (5-40 ng/ml), as compared to the moderately lower quantity of bound lipopolysaccharide with [3H]-LPS + LBP alone, or compared to the much lower quantity of lipopolysaccharide bound when [3H]-LPS was used alone. In the presence of whole serum, the effects of rsCD14 were dichotomous and depended upon the serum concentration. rsCD14 (4 μg/ml) added to medium containing ≥ 1% (v/v) fetal bovine serum resulted in reduced binding/uptake of [3H]-LPS by AM. However, rsCD14 slightly enhanced binding of [3H]-LPS to AM at a very low concentration of serum (0.1%) and in the absence of serum. These results suggest that sCD14 enhances binding of [3H]-LPS to macrophages when serum-deficient conditions prevail, and sCD14 also enhances binding of LPS in the presence of low concentrations of purified LBP. However, sCD14 decreases association of LPS with AM in the presence of serum (≥ 1 %).