DIPHOSPHORYL LIPID-A DERIVED FROM THE LIPOPOLYSACCHARIDE (LPS) OF RHODOBACTER-SPHAEROIDES ATCC-17023 IS A POTENT COMPETITIVE LPS INHIBITOR IN MURINE MACROPHAGE-LIKE J774.1 CELLS

Pentaacyl diphosphoryl lipid A derived from the nontoxic lipopolysaccharide (LPS) of Rhodobacter sphaeroides ATCC 17023 (RsDPLA) did not induce tumour necrosis factor-alpha nor interleukin-6 release in the murine macrophage-like cell line J774.1. However, it effectively inhibited the induction of these two cytokines by LPS of Salmonella minnesota Re mutant R595 (ReLPS) in a concentration-dependent manner. Maximal inhibition and half-maximal inhibition occurred when the ReLPS to RsDPLA mass ratio was 1:30 and 1:1, respectively. A binding study was performed in the presence of serum to determine whether RsDPLA is competing with ReLPS for LPS binding sites on J774.1 cells. This assay allows the determination of LPS binding to J774.1 cells via a mechanism involving CD14, a receptor for complexes of LPS with LPS binding protein (LBP), and its possible inhibition. The results show that RsDPLA strongly inhibits the binding of(125) I-labelled ReLPS to J774.1 cells. Maximal and one-half maximal inhibition of binding occurred when the ReLPS to RsDPLA mass ratios were 1:2.5 and 1:0.5, respectively. It was found that the inhibition of binding by RsDPLA was much stronger than that by unlabelled ReLPS. These results suggest that RsDPLA is competing with ReLPS for CD14-dependent recognition of LPS on J774.1 cells.