1. The major pathological responses to Gram-negative bacterial
sepsis are triggered by
endotoxin or
lipopolysaccharide. As
endotoxin is shed from the bacterial outer membrane, it induces immunological responses that lead to release of a variety of
cytokines and other cellular mediators. As part of a program aimed at developing a therapeutic agent for
septic shock, we have developed
E5531, a novel synthetic
lipopolysaccharide antagonist. 2. As measured by release by tumour
necrosis factor-alpha, human monocytes or whole blood can be activated by
lipopolysaccharide,
lipid A, and
lipoteichoic acid (from Gram-positive bacteria).
E5531 potently antagonizes activation by all these agents while itself being devoid of agonistic activity. 3. The inhibitory activity of
E5531 was dependent on time of addition. When 10 nM
E5531 was added simultaneously with
lipopolysaccharide or 1 - 3 h before addition of
lipopolysaccharide, production of tumour
necrosis factor-alpha was inhibited by more than 98%. The addition of
E5531 1 h after
lipopolysaccharide reduced the efficacy of
E5531 by 47%. 4. Antagonistic activity of
E5531 was specific for
lipopolysaccharide as it was ineffective at inhibiting
interferon-gamma mediated NO release of RAW 264.7 cells, phorbor 12-myristate 13-acetate stimulated
superoxide anion production in human neutrophils,
concanavalin A stimulated mitogenic activity in murine thymocytes and
tumor necrosis factor-alpha induced
E-selectin expression in human umbilical vein endothelial cells. 5.
E5531 as well as MY4, an anti-CD14 antibody, inhibited radiolabelled
lipopolysaccharide binding in human monocytes. 6. These results support our contention that
E5531 is a potent antagonist of
lipopolysaccharide-induced release of tumour
necrosis factor-alpha and other cellular mediators and may be an effective therapeutic agent for human
septic shock due to Gram-negative bacteria.